'LINES OP Dairy Bacteriolo; ;-NRLF EMI MS3 RUSSELi JV_ii rv_n_n_'i_- " " " - " -' " - - ' '- - - - " n - - --_ - -..- i_JT_n_n_n_^n_ _n_n_ r REESE LIBRARY OF THE UNIVERSITY OF CALIFORNIA. l^ecewed , igo . BIOLOGY Accession No... 8^60^ - OatsNo.*?* u, ,u u u u u u u u u u ll u u if u u u u u u u u u u uir-u u u u ii ii u U--M--U u u J BEESg OUTLINES OF DAIRYBACTERIOLOGY A Concise Manual for the Use of Students in Dairying BY H. L. RUSSELL PROFESSOR OF BACTERIOLOGY University of Wisconsin UNIVERSITY FOURTH EDITION, THOROUGHLY REVISED MADISON, WIS. H. L. RUSSELL 1899 COPYRIGHTED 1894, 1896, AND i! BY H. L. RUSSELL. Tracy, Gibbs & Co., Printers, Madison, Wis. PREFACE TO FOURTH EDITION. The cordial reception with which these Outlines continue to be received shows that the effort to keep them abreast of bacteriological advance is appreciated by students of dairying. An exposition of the principles of bacteriology as applied to dairy problems is now recognized as an essential part of dairy science, for modern dairy practice is rapidly adopting the methods that have been developed through bacteriological study. Although a thoroughly satisfactory explanation has not as yet been given for all the phenomena connected with milk and its products, still a better understanding of these prob- lems has been rendered possible through the light that has been thrown upon these questions by scientific study from a bacteriological point of view. A brief glossary is appended. embracing those technical terms that are necessarily employed. My acknowledgments are due to the following parties for the loan of cuts: Wisconsin Agricultural Experiment Station: Western Creamery, San Francisco, Cal.; Cornish, Curtis and Greene, Ft. Atkinson, Wis.; and A. H. Reid and Co., Philadelphia, Pa. H. L. RUSSELL. Wisconsin Dairy School, Madison, Wis., Dec., 1898. 82602 INTRODUCTION. Bacteriology is the youngest member of the sisterhood of those biological sciences that deal with life in its varied man- ifestations and functions. While it has a strictly scientific side, this subject has a greater practical bearing than is found in many of its sister sciences. For a long time is was merely a protege of medicine; even now, the term bacteria is nearly always associated in the minds of many with some dread con- tagious disease, but with further study, the effect of bacteria in many other practical lines is becoming better known, and the circle of its influence is steadily widening. Its methods have revolutionized the brewing industries; on the presence of bacteria depends the success or failure of many of the industrial arts, such as butter and cheese-making, many of the operations in tanning, the manufacture of vine- gar, of wines, etc. Modern agriculture recognizes the effect of germ life in the various processes of fertilization by natural manures; in the accumulation of nitrogenous food in the soil as a result of the action of nitrification, and in the fixation of free nitrogen of the air by members of the clover family. Sometimes the bacteria are to us a scourge, but often, in- deed, they come in the character of a friend and helper. More especially is this true of that class that are associated with dairy products, for in both the butter and cheese indus- tries, success is only assured through the activities of certain favorable forms. The following pages attempt to show this group of organ- isms so infinitely small yet almost infinitely powerful in their relation to the dairy and dairy products. Knowledge in dairying, like all other technical industries has grown mainly out of experience. The facts have been learned by observation, but the why of each is frequently shrouded in mystery. vi Introduction. Modern dairying is attempting to build its more accurate knowledge upon a broader and surer foundation, and in doing this is seeking to ascertain the cause of well established pro- cesses. In this, bacteriology is destined to play an important role. Indeed, it may be safely predicted that future progress in dairying will, to a large extent, depend upon bacteriologi- cal research. As Fleischmann, the eminent German dairy scientist, says: "The gradual abolition of uncertainty sur- rounding dairy manufacture is the present important duty which lies before us, and its solution can only be effected by bacteriology." TABLE OF CONTENTS. PART 1. Synopsis of Bacteria in General. Chapter I. Structure and form 1 Chapter II. Physiology , 5 Chapter III. Methods of studying bacteria 17 PART II. Bacteria in Relation to Milk. Chapter IV. Contamination of milk 24 a. Infection on the farm 26 b. Infection in the factory 46 Chapter V. Milk fermentations and their treatment 54 Chapter VI. Disease-producing bacteria in milk '. 75 a. Infection direct from affected animal 76 b. Infection subsequent to milking 80 c. Poison-forming bacteria 85 Chapter VII. Principles of milk preservation 87 a. Chemical methods 88 b. Physical Methods 90 Sterilization 94 Pasteurization 95 Pasteurizing apparatus ..." 106 PART III. Bacteria in Relation to flilk Products. Chapter VIII. Bacteria in cream and factory by-products 118 Chapter IX. Bacteria in butter-making 127 a. Beneficent action in butter 128 b. Bacterial defects in butter 145 Chapter X. Bacteria in cheese industry 148 a. Bacteria in normal cheese-ripening 150 b. Bacteria in abnormal cheese processes 164 PART I. GENERAL PRINCIPLES OF BACTERIOLOGY. CHAPTER I. STRUCTURE AND FORM. BEFORE one can gain any intelligent conception of the manner in which bacteria affect dairying, it is first necessary to know something of the life history of these organisms in general, how they live, move, and react toward their environment. 1. What are bacteria? Toadstools, smuts, rusts, and mildews are known to even the casual observer, be- cause they are of such evident size. Their plant-like nature can be readily understood from their general struc- ture and habits of life. The bacteria, however, are so small that, under ordinary conditions, they only become evident to our unaided senses by the by-products of their activity. When Leeuwenhoek (pronounced Lave-en-hake) in 1675 first discovered these tiny, rapidly moving organ- isms, he thought they were animals. Indeed, under a microscope, many of them bear a close resemblance to those minute worms found in vinegar that are known as "vinegar eels." The idea that they belonged to the ani- mal kingdom continued to hold ground until after the middle of the present century ; but with the improvement in microscopes, a more thorough study of these tiny structures was made possible, and their vegetable nature i B. ' 2 Dairy Bacteriology. demonstrated. The bacteria as a class are separated from the fungi (smuts, molds, etc.) , mainly by their method of growth; from the lower green algae by the absence of chlorophyll, the green coloring matter of vegetable organisms . 2. Structure of bacteria. As far as structure is con- cerned the bacteria stand on the lowest plane of vegetable life. They are single- celled plants. In its inner struc- ture, the cell does not differ essentially from that of many other types of plant life. It is composed of a proto- plasmic body which is surrounded by a thin membrane that separates it from neighboring cells that are alike in form and size. 3. Form and size. Where a plant is composed of a single cell but little difference in form is to be expected. While there are intermediate stages that grade insensibly into one another, the bacteria may be grouped into three main types (so far as form is concerned). These are FIG. 1. Different forms of bacteria, a, b, c, represent different types as to form: a, coccus, b, bacillus, c, spirillum; d, diplococcus or twin coccus; e, staphy- lococcus or cluster coccus; / and g, different forms of bacilli, shows internal endospores within cell; h and *', bacilli with motile organs (cilia). spherical, elongated, and spiral, and to these different types are given the names, respectively, coccus, bacillus, and spirillum (plural, cocci, bacilli, spirilla), (fig. 1). Structure and Form. 3 A ball, a short rod, and a corkscrew serve as convenient models to illustrate these different forms. In size, the bacteria as a class are the smallest organ- isms that are known to exist. Relatively there is consid- erable difference in size between the different species, yet in absolute amount this is so slight as to require the highest powers of the microscope to detect it. As an average diameter, one-thirty-thousandth of an inch may be taken. If a hundred individual germs could be placed side by side, their total thickness would not equal that of a single sheet of paper upon which this page is printed. 4. Manner of growth. As the cell increases in size as a result of growth, it usually elongates in one direc- tion, and finally a new cell- wall is formed, dividing the -so- called mother- cell into two equal- sized daughter- cells. This process of cell division, which is called, fission, is continued almost indefinitely until growth ceases as a nat- ural sequence. 5. Cell arrangement. If this process of fission goes on in the same plane it results in the formation of a cell- row. A species forming such a chain of cells of the coccus type is called strepto - coccus (chain- coccus). If the second division plane is formed at right angles to the first, a cell -surface is formed. If growth takes place in three dimensions of space, a cell -mass is produced as in the sarcina group. In some cases these cell aggregates cohere so tenaciously that this character is of value in distinguishing different species. 6. Reproduction. The process of cell division known as fission enables the bacterial form to reproduce itself rapidly. ' Some species possess another method of vege- tative reproduction, viz., spore-formation (fig. 1, g) . The spores are usually formed within a mother-cell ; hence, 4 Dairy Bacteriology. they are called endospores. The protoplasm of the endo- spore contracts into a small ball, and finally acquires a thickened outer part that enables it to resist unfavorable surroundings; therefore, they are adapted to the perpetua- tion of the species under severe conditions, and are analo- gous in this respect to the seeds of higher plants. Many of the bacilli form spores, the cocci do not. Spores do not germinate and grow as readily as the ordinary cells. 7. Movement. Many bacteria are unable to move about from place to place. They have, however, a vibra- tory movement known as the Brownian motion, that is purely physical. Many other forms possess an ability to move about. This they do by means of fine thread-like processes on the edge of the cell known as cilia (sing. cilium) . Coccus forms in general are non-motile, while bacilli may or may not possess these locomotor organs. 8. Classification. In arranging bacteria according to their various relationships, difficulties are experienced that do not obtain with the higher forms of plants. There exists so little difference between the various spe- cies as regards form and size that it becomes necessary to employ other characters of a physiological nature, as for instance the way in which the germ develops in cul- ture media, the by-products that are formed as a result of growth and other characters of a similar nature. CHAPTER II. PHYSIOLOGY. 9. Conditions essential for growth. The growth of bacteria, like all other living organisms, bears a direct relation to their external surroundings. Certain condi- tions are absolutely essential before life can develop. Other conditions though often advantageous are not of such vital importance. 10. 1. Food-supply. Concerning the character of the food- supply necessary for different species there is a very great difference. Many of the disease- producing forms are very particular in the selection of their food. With those forms that are usually found in milk, such delicacy of choice is not common. A food substance to be available for bacteria must be in solution, as it must pass through the cell-wall by absorption. Bacteria can live upon solid substances, but they form chemical substances that enable them to render soluble the necessary food material. The essential food elements that must be present are nitrogen, carbon, and oxygen, together with minute quan- tities of mineral elements. The nitrogen and carbon are more available when in the form of organic compounds than as simple inorganic salts. Albuminous or proteid substances are best adapted for the nitrogen supply, while sugars are available for the carbonaceous part of the food. The nitrogenous element is, however, the most indispensable. Concentration of medium. If the fluid is too dense, bacteria cannot grow. Condensed milk or syrup is too [51 6 Dairy Bacteriology. concentrated to permit of bacterial growth, but if diluted considerably, rapidly undergoes fermentation. The keep- ing quality of these products is dependent upon this prop- erty. Chemical reaction of medium. As a rule, bacteria prefer a slightly alkaline to an acid medium, but there are so many exceptions to this rule, that the force of it as a gen- eral statement is not very strong. Those organisms that are normal inhabitants of milk are, as a rule, less sus- ceptible to slight variations in the reaction of the food medium than many others. 11. 2. Temperature. A certain degree of heat is ab- solutely necessary before the spores of bacteria can ger- minate, just as seed grain will not sprout when the ground is too cold. As the temperature of a fluid increases, the rapidity with which the bacteria multiply also increases for a time. Beyond a certain point, however, a heat rigor sets in that destroys the activity of the protoplasm. There is therefore, an optimum or best temperature for growth, and minimum and maximum points as well, below and above which, development is impossible. These three cardinal growth points vary considerably with different germs. The temperature limits of growth, i. e., the range be- tween the maximum and minimum points of development, are much wider with bacteria than with almost any other forms of living matter. For this reason, bacteria are more widely distributed than any other class of living organisms. Some forms thrive at 32 F., while others are able to grow at temperatures approximating 140 F. With the great majority of bacteria, especially those growing in milk, this range is not so great. Most milk bacteria fail to develop at a point below 40-50 F . , while the maximum growth point does not exceed 105-110 F., the optimum Physiology. 1 ranging from 80-100 F. Parasitic forms living in the animal or human body have a high optimum, usually ap- proximating the blood temperature (98-100 F.). 12. 3. Gaseous environment. To most forms of life, atmospheric air is a necessity, in order to supply the oxygen used in growth. With the bacteria, the -great majority require the free access of air the same as other living organisms, and if denied this, fail to grow. Such bacteria are called aerobic. Toward some, however, oxy- gen acts as a direct poison. Only when they are sur- rounded with an atmosphere other than air, such as hydrogen or nitrogen gas, can they grow. These forms are called anaerobic. All bacteria are not divided sharply into these two classes. While some of them grow strictly under one condition or the other, hence are called obligate aerobes or anaerobes, other forms are seemingly indiffer- ent to their gaseous surroundings. To this class, the name of facultative or optional aerobe or anaerobe is given, depending upon the relation of the germ to the oxygen supply (fig. 1, d, e) . Most milk bacteria belong to the aerobic class, but there are quite a number that are able to grow without free oxygen. 13. 4. Moisture. On a dry medium, bacteria can not grow, neither can the spores germinate. A certain amount of moisture is, therefore, necessary before bacterial growth can take place. Organic substances such as vegetable or animal tissue contain sufficient water to permit growth. 14. Rate of growth. The rate of growth of actively vegetating bacteria is in many instances perfectly astound- ing. With many species under favorable conditions, a single cell will divide in twenty minutes, and each of the daughter- cells will repeat this operation in the same 8 Dairy Bacteriology. time. This rapid rate cannot be maintained indefinitely, for the bacteria soon limit their own development by the production of by-products that are unfavorable to their own growth. The sour milk bacillus thrives readily in milk until the lactic acid that is formed by it exceeds a certain amount, then growth ceases. If this acid is neu- tralized by the addition of chalk, the lactic bacteria will start again to grow, and produce acid to the point of excess. There is a marked difference with different forms in their rate of growth, and the conditions under which development best occurs. 15. Effect of external conditions. Bacteria, even in a vegetating stage, possess a much greater resistance toward external forces than other forms of animal or veg- etable life. When they are in a spore stage, this resist- ance is still further increased. A thorough knowledge of the effect of these external forces is essential, for it is by their action that w r e are often able to destroy undesir- able forms of germ life. 16. Physical forces. 1. Heat. The bacteria possess a high power of resistance toward heat ; but this relation varies, depending upon the condition in which it is ap- plied, whether it is in a dry or moist state. At some temperatures moist heat, on account of its greater pene- trative power, is much more effective than dry in destroy- ing germ life. The temperature at which any form is killed is called its thermal death point. For the majority of germs in a spore-free, developing condition, a temper- ature from 130-140 F. for ten minutes in a liquid medium is fatal. A shorter exposure necessitates a some- what higher temperature. When in the more resistant spore- stage, many of them are able to withstand moist heat in the form of steam (212 F.) from one to three hours. To destroy spores Physiology. 9 by dry heat, a temperature varying from 260-300 F. is necessary for an hour or more. Germ life may be more rapidly destroyed in super-heated steam under pressure, a temperature of 230-240 F. for fifteen to twenty min- utes being sufficient to kill most species. Many of the milk bacteria, like the sour milk germ, are very easily destroyed by heat, as they do not form spores. Other forms like the hay and potato bacilli are difficult to eradicate on account of the great resistance that their spores offer toward heat. High temperatures are by far the most efficient means that can be used to render any substance germ-free and are most often employed for this purpose. 2. Cold. While the maximum temperature that the different spore-bearing species can stand has been accu- rately determined in many cases, the minimum tempera- ture has in most instances never been reached. But lit- tle reliance is to be placed upon this method in attempting to free any substance completely from bacterial life, although freezing is able to destroy a large percentage. Even an artificial degree of cold as low as 220 F. for a day has been found to be insufficient to destroy some forms, especially when in the spore stage. Bacterial growth is held in abeyance when a liquid is congealed, but development occurs very soon after it is melted. 3. Desiccation. Different bacteria behave very differ- ently when subjected to drying. The cholera germ dies in three hours if it is dried, while anthrax retains its viru- lence unimpaired for decades. Tuberculous sputum with- stands drying and is often found to be infectious after the lapse of eight or nine months. Those species that form spores naturally resist desiccation much better than those that do not form these latent structures. 10 Dairy Bacteriology. 4 . LigM . The influence of light on bacterial growth has not been generally appreciated until within a few years. Exposed to the rays of direct sunlight, many forms are killed in a few hours. Even diffused daylight often ex- erts a powerful inhibiting effect, if the exposure covers any considerable length of time. Bacterial spores are not as readily destroyed as the vegetative forms. 17. Chemical substances. Different chemical sub- stances exert a powerful toxic action on bacterial life with which they come in contact. Those that destroy or kill bacterial forms are known as disinfectants; those that merely inhibit, or retard growth are known as anti- septics. All substances possessing disinfecting power must of necessity be antiseptic in their action, but not all antiseptics are disinfectants even when used in strong doses. Most of the disinfectants, under ordinary cir- cumstances, have no use in creamery practice except as preservatives for samples. Of these corrosive sublimate and potassium bichromate are most frequently used. In some countries antiseptics, principally of boracic acid or formalin, are employed to "keep" milk, but in general, their use is prohibited by law. 18. The role of bacteria in nature. The great ma- jority of bacteria like animal life, belong to a class whose function it is to disintegrate organic matter and resolve it into its constituent elements. The value of this break- ing down process is evident at a glance when we consider what would be the result, if all decay, putrefaction, and decomposition were at once arrested. Not only would the supply of carbonic acid gas be very soon absorbed, stopping the development of chlorophyll-bearing plants, but all nature would be clogged and soon buried under its own debris. Dead and dying vegetable and animal Physiology. 11 matter, unable to rot and decay would soon bury all un- der the constantly accumulating mass. Owing to the energies of the lower types of plant and animal life, all of this dead effete matter is slowly consumed. The split- ting of these materials into the simpler elements is largely due to the activities of the bacteria. Almost without ex- ception, they prey upon organized matter, absorbing suf- ficient energy for their maintenance, and at the same time, giving off the unused elements to form new combinations that are again absorbed in different forms. 19. Specialization of bacteria. With the higher plant and animal forms, a certain specialization is in progress. The plant or the animal adapts itself to its environment, and in doing so, each group as a whole acquires certain characteristics that mark it in a greater or lesser degree. Usually, specialization accompanies a complexity of struc- ture, so that the higher developed plants and animals show this characteristic more fully than those that belong to the more primitive types. In the bacteria is seen a curious anomaly to this rule. Simple in structure as they appear to be, they surpass in functional variability many of the higher forms of plant and animal life. Ubiquitous in their distribution through- out the realm of nature, they have gradually adapted themselves to the different habitats in which they have found themselves, and as a result, more or less well marked groups are to be found. Thus, there is a normal bacte- rial flora of the mouth, another of the skin, and still another of the intestines. Again, there is a class known collectively as water bacteria, still another that might with propriety be classified as milk bacteria. Those forms that have become habituated to certain conditions, may be said to be indigenous or native to that habitat, while the presence of uncommon forms from some other source 12 Dairy Bacteriology. would be considered as adventive or introduced. Thus, the tubercle bacillus is adventive in milk, even though it maybe derived from the udder direct, while the sour milk bacillus is a natural and therefore indigenous organism. 20. Distribution of bacteria. If one classifies bac- teria according to the habitat in which they are found, they may be divided into various groups such as soil, air or water forms. 1 . Soil bacteria . The superficial layers of the soil teem with myriads of forms of bacteria. The amount of food present and other favorable growth conditions enable many forms to thrive luxuriantly. At a depth of a few feet most of them are filtered out, and the conditions are likewise unsuitable for their growth . so in deep layers the soil is practically sterile. 2. Air bacteria. The bacteria found in the air are orig- inally from the soil beneath. In the atmosphere they are unable to develop, but exist in a latent condition. Their prevalence in the air is measured by the condition of the soil below and the movement of dust particles. For this reason they are more numerous in summer than in win- ter; the atmosphere of cities contains a larger number of them than country air. They are very prevalent in illy- ventilated houses and out-buildings, particularly barns and stables, where dust from hay and dried manure par- ticles fill the air. 3. Water bacteria. Water when exposed to the air invariably contains a sufficient amount of organic matter to serve as bacterial food. Some of its germ life is de- rived from dust, or the washings of the land, but many species exist in this element which are not to be found in soil. Stagnant pools rich in organic matter always teem with bacteria, and even running water often has large numbers. The ground water layer is normally free, as Physiology. 13 the bacteria are filtered out by passing through the inter- vening soil layers. As a consequence spring water as it issues from the soil is relatively poor in bacteria, but quickly becomes contaminated after it reaches the sur- face. Some of the highly infectious diseases, such as cholera and typhoid fever are often transmitted by means of a contaminated water-supply. 21. Saprophytic bacteria. If bacteria are consid- ered from the manner in which they live, they may be divided into two very unequal divisions, known as sapro- phytes and parasites. Those belonging to the first class subsist on dead organic matter, while the parasitic forms are able to thrive in living tissues of either vegetable or animal nature. The great majority of different forms belong to the first class, and it is undoubtedly true that this condition is more fundamental than the parasitic method of life. The saprophytes find their food- supply in the vegetable and animal matter that has already ceased to live. They are the organisms concerned in the tear- ing down processes of nature and their beneficent func- tion in the world about them is in their scavenger char- acter as they make way with the offal and debris of organic life. 22. Parasitic bacteria. There is no sharp line sep- arating the parasitic bacteria from those that live on lifeless matter. In all probability these forms that are now able to thrive in living tissue came originally from ancestors of a saprophytic type. Some, as the tubercle bacillus, have gradually adapted themselves to the more restricted parasitic method of growth, just as many para- sitic plants have been produced, and as a result of this specialization, they have often lost the power to thrive under conditions generally favorable to saprophytic forms. In parasitism a marked variation in degree is to be 14 Dairy Bacteriology. noted. Some species, like leprosy, grow with only the greatest difficulty outside of their proper host. Such as this may be said to be an obligatory parasite. Then again, other forms like the colon or feces bacillus are normally saprophytes, but under certain conditions, as- sume a semi-parasitic mode of life, becoming therefore, facultative parasites, i. e., they possess at times the fac- ulty of developing under parasitic conditions. 23. Fermentation. Most of the saprophytic bacteria are concerned in the breaking down of organic matter, and consequently, they are often associated with the many different phases that this process assumes as seen in putrefaction, fermentation, decomposition or decay. These changes are of a complex character and vary much from one instance to another. The changes embraced under the special term, fermen- tation, are distinguished by such a prominent character- istic that they may well be considered separately. In fermentation, complex substances are transformed by regular steps into simpler compounds. In this class are to be included a large number of fer- mentative changes that occur in milk, such as the produc- tion of lactic acid in the souring of milk, the formation of butyric acid, and others that will be mentioned later. In fermentation, there are often large quantities of the fer- mentable substances changed, and, while a certain amount of the energy released in the breaking down of this ma- terial may be utilized by the bacterial cells, yet the dis- ruptive changes set in motion by the living germs are out of all proportion to the results that are seen in the end. Fermentation, although one of the best known pro- cesses that occurs in nature is even yet a partial mystery. Our knowledge of the changes that fermentable solutions Physiology. 15 undergo during this process has been vastly augmented during the latter half of this century, but even yet, the problem has not been completely solved. The results of fermentation have been made use of from time immemo- rial, but no adequate conception of the changes involved was ever recognized until this century. The earlier the- ories of this action were mainly chemical, and it was not until the important studies of Pasteur were made that the relation of these changes to the action of living or- ganisms was fully proven. He showed that fermentation was closely connected with the growth and multiplication of minute forms of organic life, and that the process was usually inaugurated by vital forces rather than by purely^ chemical activity. Previous to his work the action of rennet upon milk had been known, and the difference between this fermen- tation and other types had been recognized by Schroeder and Dusch. They showed that the rennet ferment was unaffected by alcohol and other chemicals, while other fermentations due to organisms were checked in these fluids. Pasteur emphasized the distinction between these two sets of fermentative action and soon was able to clas- sify these changes into two distinct types. 1. Organized or living ferments those in which the fermentative change takes place as a result of the activity of a living organism. 2. Unorganized or chemical ferments those in which the change is caused by a chemical substance, devoid of vitality, that is itself unchanged in the fermenting pro- cess. These unorganized, non- vital ferments are known as enzymes. Among the better known of these are ren- net, that has the power of coagulating milk; diastase, the enzyme that converts starch into sugar; pepsin and trypsin, the digestive ferments of the animal body. In 16 Dairy Bacteriology. their relation toward external influences such as heat, etc., many of these are affected in a manner similar to the vital ferments. Among the organized class of ferments are those that directly affect the character of sugar- containing fluids, such as the yeasts and most of the bacteria. A great many of these organized ferments accomplish their effect indirectly through the agency of the enzymes that they themselves excrete. For instance, many of the abnormal fermentations in milk are caused by bacteria that are able to form various enzymes . CHAPTER III. METHODS OF STUDYING BACTERIA. 24. Necessity of bacterial masses for study. Bac- teria are so infinitesimally small that it is impossible to study individual germs separately without the aid of first- class microscopes. For this reason, but little advance was made in the knowledge of these lower forms of plant life, until the introduction of culture methods, whereby a single organism could be cultivated and the progeny of this cell increased to such an extent in a short course of time, that they would be visible to the unaided eye. 25. Culture methods. The system of cultivating bacteria, known as the pure culture method, is based upon the supposition that the food medium in which the organism is grown is first freed completely from all pre- existing forms of life, or in other words, is perfectly sterile. The pure culture processes of the bacteriologist may be said to be in a sense, refined methods of seeding, such as the agriculturist employs . Just as the seed grain will in due season bring forth a harvest after its kind, so any kind of bacteria planted in a favorable food medium will produce a crop of its own. If the farmer's seed is foul, it shows in%is crop, and the same is true with bacterial farming. Bacteria, however, are so universally distributed that it becomes an impossibility to grow any special kind, unless the soil is first freed from all existing forms of germ life. To accomplish this, it is necessary to subject the nutrient medium used for a culture to some method of sterilization, such as by heat or filtration, whereby all 2-B. I 18 Dairy Bacteriology. forms of organic life are thoroughly eliminated. Germ- free culture material is kept in sterilized glass tubes and flasks, and is protected from outside infection by plugs of sterile cotton. Material thus prepared, if protected from evaporation, will keep indefinitely, as the cotton acts as an effectual filter against the passage of any par- ticles of matter. 26. Culture media. For culture media, many differ- ent substances are employed. In fact, bacteria will grow on almost any organic substance whether it is solid or fluid, provided the essential conditions of growth are fur- nished. The food substances that are used for culture purposes are divided into two classes; solids and liquids. Solid media may be either permanently solid like pota- toes or they may retain their solid properties only at cer- tain temperatures like gelatin or agar. These last are of utmost importance in bacteriological research, for their use, which was introduced b(y Koch, permits the separa- tion of the different forms that may happen to be in any mixture. Gelatin is used advantageously because the majority of bacteria present wider differences in their ap- pearance upon this medium than upon any other. It re- mains solid at ordinary temperatures, becoming liquid in the neighborhood of 70 F. Agar, a gelatinous product derived from a Japanese sea- weed, has a much higher melting point, and can be successfully used, especially with those organisms whose optimum growth point is above the melting point of gelatin. Besides these solid media, different liquid substances are extensively used, such as beef broth, milk, and in- fusions of various vegetable and animal tissues. Skim- milk is of especial value in studying the milk bacteria and may be used in its natural condition, or a few drops of litmus solution may be added in order to detect any change in its chemical reaction due to the bacteria. Methods of Studying Bacteria. 19 27. Methods of isolation. Suppose for instance one wishes to isolate the different varieties of bacteria found in milk. The method of procedure is as follows: Sterile gelatin in glass tubes is melted and cooled down so as to be barely warm. To this gelatin which is germ-free a drop of milk is added. The gelatin is then gently shaken so as to thoroughly distribute the milk particles, and poured out into a sterile flat glass dish and quickly cov- b--- FIG. 2. A gelatin plate culture showing appearance of different organisms from a sample of milk. Each mass represents a bacterial growth (colony) de- rived froia a single cell. Different forms react differently toward the gelatin, some liquefying the same, others growing in a restricted mass, a, represents a colony of the ordinary bread mold; b, a liquefying organism; c, and d, solid forms. ered. This is allowed to stand on a cool surface until the gelatin hardens. After the culture plate has been left for twenty- four to thirty- six hours at the proper temperature, tiny spots will begin to appear on the sur- face, or in the depth of the culture medium. These 20 Dairy Bacteriology. patches are called colonies and are composed of infinite numbers of individual germs, the result of the continued growth of the single organism that was in the drop of milk which was firmly held in place when the gelatin solidi- fied. The number of these colonies represent in gen- eral the number of germs that were present in the milk drop. If the plate is not too thickly sown with these germs, the colonies will continue to grow and increase in FIG. 3. Profile view of gelatin plate culture. Shaded part represents the gelatin medium in the covered glass dish; on the surface, different bacteria are developing; b, is a liquefying form that dissolves the gelatin while c and d grow on surface only and do not render gelatin soluble. size, and as they do, minute differences will begin to ap- pear. These differences may be in the color, the con- tour and the texture of the colony, or the manner in which it acts toward gelatin. (See fig. 3.) In order to make sure that the seeding is not too copious so as to interfere with continued study, an attenuation is usually made. This consists in taking a drop of the infected gelatin in the first tube, and transferring it by means of a sterile needle into another tube of sterile media. Usu- ally this operation is repeated again so that these culture plates are made with different amounts of seed with the expectation that in at least one plate the seeding will not be so thick as to prevent further study. To further study the peculiarities of different germs, the separate colonies are transferred to other sterile tubes Methods of Studying Bacteria. 21 of culture material and thus pure cultures of the various germs are secured. These cultures then serve as a basis for continued study and must be planted and grown upon all the different kinds of media that are obtainable. In this way, the slight variations in the growth of different 1 FIG. 4. Pure cultures of different kinds of bacteria in gelatin tubes, a, growth slight in this medium; b, growth copious at and near surface. Fine parallel filaments growing out into medium liquefying at surface; c, a rapid liquefying form; d, a gas-producing form that grows equally well in lower part of tube as at surface (facultative anaerobe); e, an obligate anaerobe, that devel- opes only in absence of air. forms are detected and the peculiar characteristics are determined, so that the student is able to recognize this form when he meets it again. 22 Dairy Bacteriology. These culture methods are of essential importance in bacteriology, as it is the only way in which it is possible to secure a quantity of germs of the same kind. 28. Use of the microscope in bacterial investiga- tion. The microscope is in constant demand throughout all the different stages of the isolating process in order to verify the purity of the cultures. For this purpose, it is essential that the instrument used shall be one of strong magnifying powers (600-800 diameters) combined with sharp definition, so that these tiny organisms shall stand out clear and distinct. The microscopical examination of any germ is quite as essential as the culture characteristics; in fact, the two must always go hand in hand. This examination reveals not only the form and size of the individual germ, but the manner in which they are united with each other, and any peculiarities of movement that they may possess. In carrying out the microscopical part of the work, not only is the organism examined in a living condition, but stained preparations are made by using solutions of anilin dyes as staining agents. These are of great service in bringing out almost imperceptible differences. The art of staining has been carried to the highest degree of per- fection in bacteriology, especially in the detection of germs that are found in diseased tissues in the animal or human body. In studying the peculiarities of any special organism, not only is it necessary that these cultural and micro- scopical characters should be closely observed, but special experiments 'must be carried out along different lines, in order to determine any special properties that the germ may possess. Thus, the ability of any form to act as a fermentative organism can be tested by fermentation ex- periments; the property of causing disease, studied by Methods of Studying Bacteria. 23 the inoculation of pure cultures into animals. A great many different methods have been devised for the pur- pose of studying special characteristics of different bacteria, but a full description of these would necessarily be so lengthy that in a work of this character they must be omitted. 1 i The following general works contain more or less complete descriptions of the various processes employed in studying bacteria: Sternberg, Manual of Bacteriology, 1898; Frankel, Bacteriology, 1891; Wood- head, Bacteria and their products, 1893; Abbott, Principles of Bacteriology, 1896; Pearmain and Moor, Applied Bacteriology, 1897; McFarland, Text-Book upon Pathogenic Bacteria, 1898. PART II. BACTERIA IN RELATION TO MILK. CHAPTER IV. CONTAMINATION OF MILK. 29. Milk as a food for bacteria. The fact that milk so readily undergoes decomposition changes shows that it is well suited for the nourishment of bacterial life. Its high content in organic matter, and the dilu- tion of the same in a watery medium makes it an excel- lent food for germ as well as mammalian life. Its dif- ferent constituents, however, possess different nutritive values. Of most importance are the nitrogen-containing compounds. The albumen which is in solution is readily available. Casein can not be appropriated on account of its insol- uble nature, unless it is first rendered soluble, a pro- cess which occurs with those bacteria that secrete en- zymes that act on proteids. Of the constituents of the milk that belong to the car- bon-containing compounds, only one can be utilized by bacteria. The fat possesses but little food value for these organisms, because it cannot be decomposed by them. The milk sugar, however, is an admirable food for many species, more especially those that are known as the lac- tic acid producing or natural milk bacteria. The bacterial cell contains so little mineral matter that the requirements of the cell for its growth are very lim- ited, yet the mineral elements of the milk are needed for the growth of any protoplasm, and are used by the bac- teria in the formation of new cell matter. [24] Contamination of Milk. 25 30. Milk, germ-free in udder. Under ordinary con- ditions, when examined in the proper manner, milk always reveals bacterial life. This germ content, how- ever, is due to infection from without, for in the udder of a healthy animal, as secreted, the milk like the other secretions and tissues of the body is normally sterile. 31. Contamination of milk. In withdrawing the milk from the udder, it invariably comes in contact with germ life. The same is true after it is milked. From the time of milking, until it is consumed in one form or another, it is continually subject to contamination from exterior sources. In the main, germ life gains access while the milk is on the farm, but even in the factory FIG. 5. Microscopic appearance of milk showing relative size of fat globules and bacteria. Group of bacteria on left are lactic acid organisms. the opportunities for infection are present in greater or lesser degree. Those forms that gain an early entrance generally predominate in the milk, as their early intro- duction enables them to develop for a longer period of time. 26 Dairy Bacteriology. A. INFECTION OF MILK ON THE FARM. 32. Sources of contamination* The bacterial life that finds its way into the milk while it is yet on the farm may be traced to several sources, which may be grouped under the following heads: Unclean dairy utensils, fore milk, coat of animal, and general atmospheric surround- ings. The relative importance of these various factors fluctuates in each individual instance. 33. Dairy utensils. Of first importance, are the ves- sels that are used during milking, and also all storage cans and other dairy utensils that come in contact with the milk after it is drawn. By unclean utensils, actually visible dirt need not always be considered, although its presence in cracks and joints of pails and cans is often evident. Unless cleansed with especial care, these places are apt to be filled with foul and decomposing material that suffice to abundantly seed the milk. Soxhlet 1 found that the addition of 0.1 per cent, of sour milk to fresh milk decreased the keeping quality of the latter from 15 30 per cent.; the addition of 1.5 per cent, diminished it 80 per cent. Where cans are not well cleaned the above amount could easily be added to the milk from the ma- terial that adhered to the walls of the can. Through negligence, vessels are often used that are either unfit or are in an improper condition for handling milk. A rusty milk- can often spoils more milk than sufficient to purchase a new vessel. Wooden pails are no longer to be tolerated in a well-regulated dairy. Where possible, vessels should be made of pressed tin. If joints are necessary, they should be well flushed with solder so that they may be easily and thoroughly cleaned. In much of the cheap tinware that is now to be found on 1 Soxhlet, Ber. d Wanderversammlung bayer. Landwirthe, Oct., 1894. Contamination of Milk. 27 the market, there is altogether too thin a veneer of solder to effectually cover the joints. 34. Use of milk-cans for transporting" factory by- products. The general custom of using the milk-cans to carry back to the farm the factory by-products (skim- milk or whey) has much in it that is to be deprecated. These by-products are generally rich in bacterial life, FIG. 6. The wrong and the right kind of a milk-pail. A, the ordinary type of pail showing sharp angle between sides and bottom; B, the same properly flushed with solder so as to facilitate thorough cleaning. The lower figure rep- resents a joint as ordinarily made in tinware. The depression a affords a place of refuge for bacteria from which they are not readily dislodged. This open joint should be filled completely with solder. more especially where the closest scrutiny is not given to the daily cleaning of the vats and tanks. Too frequently the cans are not cleaned immediately upon arrival at the farm, so that the conditions are favorable for rapid fer- mentation. This danger can be entirely obviated by a thorough cleansing process, but the patron who is shift- less in regard to his cans will generally be lax in his treatment of these vessels, and thus the opportunity for infection is present. Many of the taints that bother factories are directly traceable to such a cause. A few dirty patrons will thus seriously infect the whole supply. 28 Dairy Bacteriology. The responsibility of this defect should, however, not be laid entirely upon the shoulders of the producer. The factory operator should see that the refuse material does not accumulate in the waste vats from day to day and be transformed into a putrid mass. A dirty whey tank is not an especially good object lesson to the patron to keep his cans clean. It is possible that abnormal fermentations may thus be disseminated from one farm to another in this way. Suppose there appears in the dairy of A an infectious milk trouble, such as bitter milk. This milk is taken to the factory and passes unnoticed into the general milk- supply. The skim-milk from the separator is of course infected with the germ, and if conditions favor its growth, the whole lot soon becomes tainted. If this waste product is returned to the different patrons in the same cans that are used for the fresh milk, the probabilities are strongly in favor of some of the cans being contaminated and thus infecting the milk- supply of other patrons. If the organ- ism is endowed with spores so that it can withstand un- favorable treatment, this disease may spread from patron to patron simply through the infection of the vessels that are used for the transportation of the by-products. It would be possible to obviate any trouble arising from this source if a separate receptacle was used for this pur- pose, but the objection is frequently urged that this is impractical, yet many of the more progressive factories are following this practice with excellent results. 35. Effect of steaming 1 milk-pails. Even where utensils are in good condition and well cleaned, the germ content of the milk may be reduced, and therefore, the keeping quality enhanced by a brief steaming of the re- ceiving cans. For this experiment 1 two cans were taken, 1 Russell, nth Kept. Wis. Agr'l. Expt. Stat., p. 152, 1894. Contamination of Milk. 29 one of which had been cleaned in the ordinary way, while the other was sterilized by steaming. Before milking, the iidder of animal was thoroughly cleaned, and special pre- cautions taken to avoid raising of dust; the fore milk was also rejected. Milk was then drawn directly into these two cans and bacterial determinations made: Number of bacteria per cc. in steamed pail, 165. Number of bacteria per cc. in ordinary pail, 4265. Time before milk in steamed pail soured, 28-g- hours. Time before milk in ordinary pail soured, 23 hours. If an imperfectly cleaned pail had been used for this purpose, the difference in souring would have undoubt- edly been more apparent. To illustrate the varying germ content of cans cleaned in different ways, Harrison 1 rinsed out cans with 100 cc. of sterile water, and then determined the germ content of the same. The following data represents the number of bacteria per cc. in the rinsing water from cans improperly cleaned (series A), from cans washed in tepid water and then scalded the usual factory method ( series B ) and from cans washed in tepid water and steamed for five minutes (series C.). Effect of steaming on germ content of cans. Series A (Dirty cans) .. 238,525, 342,875, 215,400, 618,200, 806,320, 510,270, 230,100, 610,510, 418,810, 317,250. Series B (Ordinary cleaning method)... 89,320, 84,750, 26,800, 24,000, 38,400, 76,800, 15,200, 13,080, 44,160, 93,400. Series C (Approved method) 1,170, 1,792, 890, 355. 416. 36. Cleaning 1 dairy utensils. Milk vessels should never be allowed to become dry when dirty, for dried particles of milk residue are extremely difficult to remove. 1 Harrison, 22nd Kept. Ont. Agr'l. Coll., p. 113. 1896. 30 Dairy Bacteriology. In cleaning dairy utensils they should first be rinsed in lukewarm instead of hot water, so as to remove organic matter without coagulating the milk. Then wash thor- oughly in hot water, using soap or weak alkali. A borax solution is sometimes recommended for cleaning bottles. Strong alkalies should not be used. After washing, rinse thoroughly in . clean hot water ; then invert over a steam jet for a few minutes. A momentary application of hot or even boiling water is insufficient to destroy germ life that lurks in joints of vessels. Live steam is especially efficient as a germ- destroy ing agent. If steamed, the cans will dry more quickly. It is not often that steam is available on the farm, but under such conditions, it is possible to acquire practically the same results by using boiling water, although the length of exposure must be increased. Not only should the greatest care be paid to the condi- tion of the cans and milk-pails, but all dippers, strainers, and other utensils that come in contact with the milk, must be kept thoroughly clean. Cloth strainers, unless at- tended to, are objectionable, for the fine mesh of the cloth retains so much moisture that they become a veritable hot- bed of bacterial life, unless they are daily boiled or FIG. 7. Section of udder showing cf aarn g(J relation of milk-secreting tissue to milk duct (after Thanhoffer). a, ex- 37. Influence Of flPSt OP secreting tissue; e, sphincter mus- in the Contamination of milk, the importance of which is rarely recognized, comes from the bacteria that gain ac- cess to the milk, by mixing the first or fore milk with Contamination of Milk. 31 the remainder of the milk. Even when the milking is most thoroughly done, there remains in the milk ducts of the udder, a few drops of milk that afford sufficient nutriment for the development of any germs that may gain access through the opening of the teat. Accord- ing to Gernhardt, it is possible that they may penetrate the udder as far as the milk cisterns or gland tissue itself, but the evidence on this point is not decisive. The rela- tively high temperature of the teat facilitates a rapid growth. Under these conditions, a small number of organisms are able to increase in such numbers that the first few spurts of milk contain many more than those which are subsequently drawn. The following data by Harrison 1 strikingly illustrates this point: Number of bacteria per cc. in milk. Foremilk 26,070, 25,630, 38,420, 18,110, 54,800, 32,700, 43,520, 27,830, 18,500, 29,400, 45,630, 48,700. Milk after removal of fore milk 1,246, 1,150, 1,430, 3,420, 1,560, 890, 2,575, 4,820, 3,270, 1,285, 1,350. If the fore milk is received in a separate vessel and kept protected from the air, it will generally be noted that it sours more rapidly than the remainder of the milk. As a rule the number of different species found in the fore milk is usually small, not more than one or two forms being present at any time. As to the character of these forms data is conflicting. Harrison 2 reports finding peptonizing bacteria in the same, and Marshall 3 states that organisms are found that resist pasteurizing, 1 Harrison, 226! Kept. Ont. Agr'l. Coll., p. 108, 1896. M. c., p. 108. 8 Marshall, Mich. Expt. Stat., Bull. 147, p. 42. 32 Dairy Bacteriology. a characteristic not usually associated with the lactic acid class. Bolley 1 in thirty experiments found twelve out of six- teen species to belong to the lactic acid class. In no case were gas- generating species present. This fact is important in the selection of a milk from a single animal for the cultivation of a starter. This condition represents the milk of perfectly healthy animals; where the udder is diseased, as in the case of infectious garget or inflammation, bacteria may be pres- ent in much larger numbers and affect the milk seriously for food purposes. Not all species seem to be able to maintain themselves in the udder even if they should be introduced. Dinwid- die 2 injected into the milk cistern a lactic acid producing facultative anaerobe that grew luxuriantly at 99 F. An examination of the milk several times afterwards failed to show the presence of this organism in any case, although other species were isolated. 38. Dirt from animal. By many it is believed that much of the germ life that gets into the milk comes from the food of the animal. For this reason fermenting food- stuffs of any kind are regarded as unfit for use. While material of this class is not a suitable nutrient for any animal, the danger to the milk is not due to the bac- teria ingested with the food, but to a large extent to those that adhere to the animal's coat, and subsequently fall into the milk. Of course the udder may be infected if the animal is diseased as in inflammation (mammitis), 3 1 Assoc. Ag. Coll. and Expt. Stat.,1895, also Cent. f. Bakt. II Abt. 1: 795, 1895. Second article, Bull. No. 21, N. D. Expt. Station. 2 Dinwiddie, Ark Expt. Stat, Bull. 45, p. 57. 3 Guillebeau, Landw. Jahr. d. Schweiz, 1892, p. 27. Contamination of Milk. 33 or tuberculosis, in which case bacteria find their way di- rectly into the milk. Marshall 1 succeeded in isolating direct from the udder, a pure culture of Nocard's strep- tococcus, the cause of infectious udder inflammation. The hairy coat of the animal offers exceptional facili- ties for the harboring of dust and dirt. Cows wading in stagnant pools cover the udder with slime and dirt that is readily dislodged when dried. The hairy coat is, there- fore, extremely rich in the various forms of bacterial life that are derived from the particles of excreta that stick to the flanks and under parts of the animal. Where peat is used for bedding, favorable reports are made as to its value. The amount of actual impurities that are to be found in milk, even after it is strained, will surprise the casual observer, although it should be noted according to Backhaus, 2 that about one-half of fresh manure dis- solves in milk and thus does not appear as sediment. From a large number of determinations of the solid im- purities found in the market milk of different European cities, Renk 3 deduces the following rule: If a sample of milk shows any evidence of impurity settling on a trans- parent bottom within two hours, it is to be regarded as containing too much solid impurities. These solid par- ticles, composed largely of manure and dirt, are always teeming with bacteria, especially with putrefactive and decomposition organisms. It has recently been estimated that the city of Berlin consumes daily 300 pounds of dirt and filth in its milk- supply. Not only is the number of bacteria thus introduced into the milk very considerable, but the character of the same 1 Marshall, Mich. Expt. Stat., Bull. 146, p. 6. 'Backhaus, Milch Ztg., 26: 357, 1897. 3 Renk, Cent. f. Bakt., 10: 193. 3-B. 34 Dairy Bacteriology. is a question of even greater importance. Those species that are derived primarily from manure particles are, as a rule, the peptonizing or digestive species that cause a decomposition of the casein, and should, therefore, be avoided if possible. Improper stable conditions greatly favor the amount of filth that may adhere to the animal. The more highly nitrogeneous feeding that is practiced at present pro- duces a softer manure and one in which putrefactive bac- teria are much more likely to be abundant. Wiithrich and Freudenreich 1 have studied the influ- ence of feeding on the bacterial content of manure, and they find a markedly higher content in manure where animals are given dry feed than where kept on grass. The character of the manure, however, is different, it being much more liquid with moist than dry feed, and therefore, they believe more likely to find its way into the milk. They found as many as 375,000,000 bacteria per gram in fresh manure, the majority of which consisted of B. coli communis, the hay bacillus, and other species able to peptonize the casein. 39. Influence of tne milker. The condition of the milker is by no means an unimportant factor. If he per- forms the milking in the dust- laden garments that he has worn all day, he himself is covered with particles that are readily dislodged when he comes in contact with the cow. Particular attention should be paid to the hands of the milker. The habit of moistening the hands with a few drops of milk just before milking is to be deprecated from every standpoint, but especially so, when consid- ered from our present point of view. After having !Cent. f. Bakt., II Abt., 1: 873, 1895. ^ OF THB r UNIVERSITY Contamination of Milk. 35 washed his hands in clean water, a pinch of vaseline on his hands will enable him to obtain a firmer grasp, and at the same time, any scales or dirt rubbed from the teat would be held by the vaseline. Its healing effect on chapped or sore teats would also be helpful. Freuden- reich 1 reports some experiments in which the germ con- tent of milk was reduced from several thousand to 200 where the hands were well rubbed with vaseline before milking. Where the best of conditions are carried out it is worth while to have the milker clothed in a suit kept for this purpose, especially the upper portion of the body. An outer garment could easily be slipped over the regular working clothes. This garment should be laundried at frequent intervals. 40. Exclusion of dirt. A large amount of filth and dirt can be prevented from falling into the milk. Card- ing the udder and flanks to remove the loose hairs will remove a considerable source of dirt. So long, however, .as the coat of the animal is dry, dust particles with their adherent bacteria are readily dislodged. If the coat of the animal is moist, this deposition can be almost effect- ually prevented. The surface should, however, not be dripping wet. The objection has been urged by some that washing the udder starts the milk secretion, and un- less the animal is milked at once, the yield of milk is diminished thereby. Eckles 2 has recently reported a number of experiments from which he concludes that when the animal is accustomed to the treatment, no no- ticeable effect is produced either in amount of milk or butter fat. In order to show the effect of dirt and dust, the experi- ment described below teaches a valuable lesson. A cow ipreudenreich, Die Bakteriologie, p. 30. * Eckles, Hoard's Dairyman, Aug. 8, 1898. 36 Dairy Bacteriology. that had been pastured in a meadow was partially milked out of doors. During the operation a covered glass dish containing sterile gelatin was exposed for sixty seconds underneath the belly of the cow in close proximity to the milk pail. The udder, flank, and legs of the cow were then thoroughly cleaned with water, and all of the pre- cautions referred to before were carried out, and the milk- ing then resumed. A second plate was then exposed in the same place for an equal length of time; a control also being made at the same time at a distance of ten feet from the animal and six feet from the ground to as- certain the germ contents of the surrounding air. From this experiment the following instructive data were gathered. Where the animal was milked without any special precautions being taken, there were 3,250 bacterial germs per minute deposited on an area equal to the exposed top of a ten- inch milk pail. Where the cow received the precautionary treatment as suggested above, there were only 115 germs per minute deposited on the same area. In the plate that was exposed to the sur- rounding air at some distance from the cow, there were sixty- five bacteria. This indicates that a large number of organisms from the dry coat of the animal can be kept out of milk if such simple precautions as these are in- stituted. Another method of exclusion is to use a milk pail hav- ing a partially closed top that will prevent the introduc- tion of a large part of the dirt. Still another method that has much to recommend it, is passing the milk through a hand separator immedi- ately. This not only removes nearly all of the suspended particles oi; foreign matter, such as dirt and filth of vari- ous kinds, 1 but it eliminates a large part of the bacteria 1 Backhaus (Milch Ztg., 26: 358, 1897) found that 95.6 per cent, of impurities were removed by centrifugal separation. Contamination of Milk. 37 with the same, if it is done immediately after the milk is drawn. The only objection to this process is that the cream does not rise so thoroughly as where it is not cen- trifuged, but the other advantages where an especially clean milk is wanted more than compensate for this defect. 41. Influence of air. It is impossible to separate the influence of the air entirely from that of the animal, as the dust particles from the coat of the animal must of necessity pass through the air. FIG. 8. Effect of contaminated air. Each spot on this surface represents a developing colony that has grown from a germ that was deposited on surface of a sterile gelatin plate (3 inches in diam.) in 30 seconds. This exposure was made at time the cows were fed. Germ life cannot develop in the air, but in a dried con- dition, organisms retain their vitality for long periods of time. The use of dry fodder, the bedding of animals with straw adds greatly to the amount of dust particles, and consequently the germ life floating in the air, as seen in fig. 8. Taints in milk have frequently been traced to infection arising from this source. 38 Dairy Bacteriology. While the stable air cannot be freed entirely from dust, much can be done with a little forethought. Feeding before milking adds materially to the germ content, espe- cially if feed is of dry character. If moistened feed is given just before or during milking, the same objection does not inure. Harrison 1 gives some striking results on this point. The numbers cited indicate the num- ber of bacteria that were deposited per minute on a surface equal to that of a twelve- inch milk pail. In Series A, the exposure was made during bedding; in B, this operation was performed an hour before. Influence of dusty air on germ life. Series A 16,000, 13,536, 12,216, 12,890, 15,340, 19,200, 23,400, 27,342, 42,750, 18,730. Series B 483, 610, 820, 715, 1,880, . 2,112, 1,650, 990, 1,342, 2,370. These results indicate that the bacteria are for the most part attached to particles of considerable weight as they settle readily to the floor. 42. The relative importance of foregoing factors. The relative importance of these various factors differs so much in different cases that 110 uniform rule can be given as to the effect of their presence in different cases. In those dairies where no especial care is exercised over the methods of handling the milk, the factor of unclean utensils and filth from the animal are usually the great- est. Where a careful supervision is given to these details, the influence of the fore milk is of first importance. The effect of these various contaminating factors can be largely minimized, and in some instances eliminated with- out great difficulty. A goodly number of the bacteria present in the fore milk can be prevented from gaining access to the milk by rejecting the first few streams that are milked from each 1 Harrison, 22d Kept. Ont. Ag. Coll., 1896, p. 111. Contamination of Milk. 39 teat, although this factor can be controlled to a less de- gree than any of the others. The actual loss occasioned in doing this is very slight, as the first part of the milk- ing is very poor in butter fat. Dust in the air of the barn is a minor factor, yet this can be materially reduced by exercising care relative to feeding dry fodder, or bedding the animals just previous to, or during the milking. The number of germs derived from the animal itself and the milker can be largely decreased by keeping the animal thoroughly clean, and having the milker milk with clean hands. The effect of contamination arising from imperfectly cleaned milk vessels can be practically excluded by ster- ilizing all such utensils in steam, or even in scalding water for a short time. To determine what reduction could be accomplished by drawing and handling the milk under as clean conditions as possible, the following experiment was carried out: The udder was thoroughly carded, and then moistened; the milk received in steamed pails, the fore milk being rejected. The milk from a cow treated in this way contained 330 bacteria per cc., while that of a mixed herd taken under usual conditions was 15,500 bacteria for same volume. This carefully handled milk kept over twenty hours longer at room temperatures than the ordinary product. Backhaus estimates that the germ life in milk can be easily reduced to Wire of its original number by using care in milking. Methods of this sort must be instituted in those dairies that are furnishing the highest grade of product. Where such methods are in vogue, bacteria are excluded for the most part and pasteurization be- comes unnecessary. Such milks are frequently known as 40 Dairy Bacteriology. "sanitary" or "certified" as the method of handling the herd is often nnder the control of ontside parties (phy sicians or health board) who certify to the regulations that are followed. FIG. 9. Bacterial content of milk handled in ordinary way. Each spot rep- resents a colony growing on gelatin plate. Compare with fig. 10, where same quantity of milk is used in making culture. Over 15,000 bacteria per cc. in this milk. The question may arise as to the necessity of these precautionary measures if the milk is to be used for or- dinary factory purposes. While certain bacteria are essential in the butter and cheese industry to secure a normal and characteristic fermentation, yet it is better to have the germ life reduced to the lowest numbers than to have the raw milk infected through slovenly methods of handling. If this is done, the maker has the fer- mentation under his control, and by the addition of a starter which he can choose, he can vary the character of the product to suit the demands of the trade, which he cannot do, if the raw milk is brought to him in a dirty condition, and in an advanced stage of fermentation. Contamination of Milk. 41 Dairymen have learned many of these lessons in the severe school of experience, but the reason for the same is so palpably plain in the light of bacteriological explana- tion, that further discussion would seem unnecessary. It FIG. 10. Bacterial content of milk drawn with care. Diminished germ con- tent is shown by smaller number of colonies (330 bacteria per cc.). Compare this culture with that shown in fig. 9. remains to be seen whether the words of the eminent German authority, Prof. Fleischmann will much longer prove true, when he says that "all the results of sci- entific investigation which have found such great prac- tical application in the treatment of disease, in disinfec- tion and in the preservation of various products, are almost entirely ignored in milking. " 43. Effect of temperature on bacterial growth. After milk is once seeded with bacterial life, no one factor exerts so potent an effect upon the rate of change that takes place as that of temperature, for the rapidity of bacterial growth in milk is determined mainly by this factor as is seen in fig. 11. 42 Dairy Bacteriology. Although different species vary in their rate of devel- opment, yet moderately warm" temperatures from 75 to 90 F., encourage rapid growth. Unless the milk is PROGENY Or A SINGLE GERM IN TWELVE: HOURS FIG. 11 Showing the effect of cooling milk on the growth of bacteria. The beneficial results of early chilling are readily apparent. quickly deprived of its original heat, the rate of the fer- mentative changes will be much increased as is shown in following results obtained by Cnopf and Escherich : Rate of growth of single germ 2 hrs. 3 hrs. 4 hrs. 5 hrs. 6 hrs. 54 F 4 6 8 26 435 97 F 23 60 215 1830 3800 If a can of milk is allowed to cool naturally, it will take several hours before it reaches the temperature of the surrounding air. During this time, the organisms in the fore milk are continuing their rapid growth, while those forms that come from dust, and are presumably in a latent state on account of their condition, awake from their lethargy under the influence of these favora- ble surroundings. If bacteria once gain an entrance and begin to germinate, a considerably lower temperature is required to successfully check development than to hold Contamination of Milk. 43 latent organisms, like spores, in a condition where germi- nation will not occur. To hasten this lowering of temperature, artifical cool- ing is a necessity. Coolers using cold running water or ice water are efficient in reducing the temperature to a point where development is much checked. 44. Tainted milks. Not all taints in milk can be traced to the development of bacterial causes. In many cases they are produced by the direct absorption of odors in a purely physical way or to some unusual condi- tion of the system of the animal. In some cases with animals old in lactation, the milk becomes abnormal in that the cream does not rise readily. Such milks are known as "lazy n or "dead" milks. Then again, the dairyman may experience difficulty in churning, but more often these difficulties are attributable to the non- performance of important manufacturing details rather than to a perverted condition of the milk itself. Milk is very prone to absorb volatile odors, the fat especially having a great affinity for many of these substances. 45. Direct absorption from animal. Odors of this sort may be absorbed by the milk previous to or after the milking has occurred. The peculiar "cowy" or "an- imal odor 7 ' of fresh milk is an illustration of an inherent peculiarity of the milk. If certain strong flavored sub- stances as onions, certain root crops, or other vegetables- are consumed by the cow, the odor-yielding substances in the same may reappear in the milk, especially if the animal partakes of these a short time before milking. According to Kober and Busey, the milk of swill-fed cows has a peculiar taste, and is said to produce a highly acid urine and eczema. Brewers' grains and distil- lery slops when fed in large quantities frequently in- duce an abnormal chemical reaction of the milk. Taints 44 Dairy Bacteriology. of this sort can easily be prevented by taking care that dairy stock is not fed such feed. 46. Straining" milk in the barn. For convenience, milk is often strained in the barn, but the danger of tainting the same where this is done is so great that the custom is not to be recommended. In straining the milk so much surface is exposed to the air that considerable bacterial infection can occur, unless this is carried out in a room free from all dust particles. Where the process is done in the barn, the can with its strainer is often left uncovered. Under these conditions, a constant deposi- tion of germ life is taking place, and with every pailful of milk strained, this is washed through the meshes of the strainer into the milk below. 47. Absorption of odors in fresh milk. Not only is straining in the barn to be deprecated from the above standpoint, but the possibility of direct physical absorp- tion of existing taints in the stable should warn one against this custom. It is a commonly accepted idea that milk evolves odors and cannot absorb them so long as it is warmer than the surrounding air, but from experi- mental evidence, l the writer has definitely shown that the direct absorption of odors takes place much more rapidly when the milk is warm than when cold, although under either condition, it absorbs volatile substances with con- siderable avidity. In testing this, fresh milk was ex- posed to an atmosphere impregnated with odors of various essential oils and other peculiar aromatic sub- stance, whose odors could be readily identified. Under these conditions the cooled milk was tainted very much less than the milk at body temperatures, even where the exposure was for a half hour. Russell, 15th Kept. Wis. Expt. Stat., p. 104, 1898. Contamination of Milk. 45 48. Aeration. Practical experience has long demon- strated the advantage of aerating the milk as soon after milking as possible. This is accomplished in a variety of ways. In some cases, air is forced into the milk; in others, the milk is allowed to distribute itself in a thin sheet over a broad surface and fall some distance so that it is brought intimately in contact with the air. The benefit claimed for aeration is that foul odors and gases which may be present in the milk are thus allowed to escape by bringing the finely divided milk into contact with the air. As ordinarily practiced, aeration is usually combined with cooling, and it is noteworthy that the most effective aerators are those that cool simultane- ously. Under these conditions, the keeping quality of the milk is increased, but where milk is simply aerated without cooling, no material benefit in keeping quality is observed. A satisfactory scientific explanation of the advantages of aeration has not yet been made. It is dif- ficult to see how the process can have any effect on the bacterial life in the milk. Its influence, undoubtedly, is on the odors directly absorbed by the milk. 49. Distinction between bacterial and non-bac- terial defects in milk. In fresh milk it is relatively easy to distinguish between taints caused by the opera- tion of external biological forces and those due to direct absorption. If the taint grows more pronounced as the age of the milk increases, it is probably due to the living organisms, as the taint- producing bacteria usually gain an entrance after the milk is drawn from the cow, and require a cer- tain period of incubation before undesirable products are formed. Taints due to errors in feeding are more pro- nounced when the milk is first drawn. 46 Dairy Bacteriology. The absorption of flavors by old milk is sometimes puzzling to distinguish from a bacterial trouble, but if the surroundings are closely noted, the cause can usually be traced to delayed absorption of bad odors from the room where milk is stored. If the trouble is of bacterial origin, it can frequently be detected by transferring a small quantity of the sus- pected milk to a fresh lot, preferably that which has first been boiled. If due to a living organism, the abnormal fermentation will be propagated and so reproduce the difficulty in the infected milk. B. INFECTION OF MILK IN THE FACTORIES. 50. Treatment of milk in factories. Where milk is taken to a creamery or cheese factory, the method of treatment often modifies the bacterial life of the milk to -a marked degree. Butter and cheese-making both require the presence of bacteria, but the germ life must be of the right sort to be beneficial in its action. For this reason the maker wishes the milk to be as free as possible from all contam- inating influences. If the foregoing suggestions are in- telligently followed, the raw material will be turned over to the factory in prime condition ; but the rational treat- ment must be continued here if the best of products is to be expected. Numerous influences are at work in the factory that add their mite to the milk in different stages of its manufacture, and ultimately affect in a serious way the final product. Of course, very much depends upon the proper observance of physical conditions, but -a brief outline of some of the possible dangers will also be pertinent. The cardinal precept in the factory as well as on the farm should be cleanliness. Cleanliness here must not Contamination of Milk. 47 be taken to mean a mere absence of dirt and filth, but all utensils that come in actual contact with the milk should be rendered as germ-free as possible. From the time that the milk enters the weigh- can until the butter is in the tub, and the cheese on the curing- shelf, it should be remembered that many opportunities for infection are always present. 51. Factory utensils. In the factory, part of the same set of factors of infection are at work as are found in the farm dairy. The condition of factory utensils is always a point of prime importance. Where steam is accessible, as it is in the majority of cases, there is no excuse for uncleanliness of any sort, as most pieces can be steamed directly. Open vats can be thoroughly scalded if covered with a heavy canvas cloth. Separator bowls, churns, cans, and dippers should always receive a daily treatment. The rational nature of these methods is to be seen in those cases where the same utensil, say a dipper, is employed indiscriminately in handling all kinds of dairy products. All cans with rusty seams should be discarded. Permit no vat to be repaired by putting in a false covering over the old one. If a minute leak is established, such places become a harbor of refuge for all kinds of putrefactive organisms . In a number of cases ill- smelling factory odors have been traced to such a cause. 52. Infection from air. The influence of the air on the germ content of the milk is, as a rule, overesti- mated. If the air is quiet, and free from dust, the amount of germ life in the same is not relatively large. In a creamery or factory, infection from this source ought to be much reduced, for the reason that the floors and walls are, as a rule, quite damp, and hence germ life cannot easily be dislodged. The majority of organisms 48 Dairy Bacteriology. found under such conditions comes from the person of the operators and attendants. Any infection can easily be prevented by having the ripening cream- vats covered with a canvas cloth. The clothing of the operator should be different from the ordinary wearing- apparel. If made of white duck, the presence of dirt is more quickly rec- ognized, and greater care will therefore be taken than if ordinary clothes are worn. The surroundings of the factory have much to do with the danger of germ infection. Many factories are poorly constructed and the drainage is poor so that filth and slime collect about and especially under the factory. The emanations from these give the peculiar "factory odor" that indicates fermenting matter. Not only are these odors absorbed directly, but germ life from the same finds its way into the milk, contaminating the same. Connell 1 has recently reported a serious defect in cheese that was traced to germ infection from defec- tive factory drains. According to Robertson, sometimes it becomes necessary to remove cheese factories to new locations before the bad conditions can be controlled. 53. Water and ice-supply of factories. These sup- plies should be carefully controlled. Water, and to a less extent ice, always contain bacteria in varying num- bers, so that it is possible that undesirable organisms may be introduced from this source. Most creameries derive their water-supply from private wells, and in using these, care should be taken that they are arranged so as not to receive any surface drainage. A deep well from which the water is used in large quantities, if properly arranged, will contain the minimum number of germs as the ground water is practically free from them. 1 Connell, Kept, of Com. of Agric., part xvi, p. 15, 1897. Contamination of Milk. 49 Harrison 1 has recently traced an off- flavor in cheese in a Canadian factory to an infection arising from the water-supply. He found the same germ in both water and cheese and by inoculating a culture into pasteurized milk succeeded in producing the undesirable flavor. Milk can easily be infected through using such water for washing utensils. Some well waters containing iron in solution are a source of trouble in factories on account of the development of iron bacteria that cause the solu- ble iron to be precipitated in the form of rusty flakes of ferric oxid. In ice the majority of organisms (60-90%) are de- stroyed, but enough remain, so that if ice is secured from a highly polluted source, the living germ life in it may be an element of danger. It is not considered ad- visable to add ice directly to milk or cream, but as stor- age vessels are indiscriminately used to contain either ice or milk, the possibility of infection should be kept in mind. 54. Numbers of bacteria in milk. The germ con- tent of milk varies so greatly that unless the conditions are all known, it is impossible to foretell what may be found therein. An examination of milk will often reveal a difference in numbers, ranging from a few score of germs to hundreds of millions per cc. The presence of such a varying number is dependent upon certain factors, as the age of the milk, the care taken during the milking, and also the way in which it has been handled since that time. Disregarding milk of different ages, the number of germs present in any sample bears a general relation to the amount of dirt and filth with which it has come in contact since it was drawn from the cow. Bacteria and 1 Harrison, Hoard's Dairyman, Mch. 4, 1898. 4-B. 50 Dairy Bacteriology. filth of all kinds are so intimately associated with each other that the presence of one rightly presupposes that of the other. As to the numerical bacterial content of any milk, there is such a wide variation under different conditions that figures are of little worth. No exact relation can be maintained between number of bacteria in milk and the development of fermentative products. The studies by different observers have been carried on under such diverse conditions that no comparison of the results can well be made. Under American conditions but little work has been done in this direction, yet milk as it is sold here to the consumer usually contains less bacteria than that retailed in European cities, although as Conn has pointed out, it is materially older. As he in- timates this fact is explained by the relatively free use of ice in this country. A few determinations of the bac- terial contents of European milks that have been analysed biologically will illustrate this point. Renk 1 found in Halle milk- supply 6-30,000,000 germs percc.; Cnopf 2 in Munich milk- supply 200, 000-6, 000,- 000 per cc.; Uhl 3 in Giessen milk 83,000-170,000,000 per cc. ; Clauss 4 in Wurzburg 222,000-23,000,000 per cc. ; Bujwid in Warsaw an average of 4,000,000 per cc. and Knochensteirn 5 in Dorpat 25,000,000 per cc. Sedgwick and Batchelder 6 report fifty- seven samples of Boston milk as containing from 30,000-4,220,000 per cc. In the country, they found in the milk fresh from iRenk, Cent. f. Bakt., 10: 193. 5 Cnopf, Ibid. 6: 553. 3 Uhl, Zeit. f. Hyg., 12: 475 (1892). 4 Clauss, Diss. Wurzburg, 1889. 5 Knochensteirn, Chem. Cent., 11: 62. 6 Sedgwick & Batchelder, Boston Med. Surg. Journ., Jan. 14, 1892. Contamination of Milk. 51 the cow 30,000 and in the milk as used on the table, about 70,000 organisms per cc. Loveland and Watson 1 found in the supply of Middletown, Conn., from 11,000 to 85,500,000 per cc. McClatchie 2 found in Los Angeles, Cal., supply, an average of 11,700 per cc. in twenty-six trials. In my experience, the mixed milk of a herd that is kept with any reasonable degree of cleanliness, if ex- amined immediately after it is milked, usually will not contain more than 520,000 germs per cc. The number present in any milk is due to the influence of so many factors that it is practically impossible to establish any number as a normal, although Bitter 3 sets 50,000 germs per cc. as a maximum limit for a milk intended for hu- man food. The milk as delivered by the milkmen to their private customers in the city of Madison, Wis., ranges from 15,000-2,000,000 organisms per cc., varying mainly with the season of the year. The presence of such large numbers in a food product need not necessarily occasion alarm from a hygienic stand- point, although it is quite certain that putrefactive forms may have an irritating effect upon a deranged digestive tract, and thus produce intestinal disturbances, especially with infants during the summer months. If we compare the bacterial flora of milk with that of sewage, a fluid that is popularly and rightly supposed to be teeming with germ life, it will almost always be ob- served that milk when it is consumed, is richer in bacte- ria by far than the sewage of our large cities. Sedgwick 4 found that the sewage of Lawrence, Mass., contained at 1 Loveland and Watson, 7th Report, Storrs Sta. (Conn.), 1894, p, 72. 2 McClatchie, Bull. 3, Agr. Expt. Stat. (So. Cal. Acad. Sc.), Aug. 1897. 3 Bitter, Zeit. f. Hyg\, 8: 240. 4 Sedgwick, Kept. Mass. Bd. Health, 1890, p. 60. 52 Dairy Bacteriology. the lowest, 100,000 germs, while the maximum number was less than 4,000,000 per cc. This range in numbers is much less than is usually found in the milk- supply of our large cities. 55. Kinds of bacteria in milk. The number of bac- teria in milk is not of so much consequence as the kind present. While milk may contain forms that are inju- rious to man, still the great majority of them have no apparent effect on human health. In their effect on milk, the case is much different. Roughly, we may di- vide them into three classes, depending upon their action in milk. 1. Bacteria that exert no appreciable effect in milk. 2. Bacteria that are beneficial by reason of the products which they form. 3. Bacteria that are injurious on account of the effect which they produce in milk. A suprisingly large number of bacteria that are found in milk belong to the first class. Undoubtedly they af- fect the chemical characteristics of the milk somewhat, but not to the extent that it becomes physically percep- tible. Those species that are concerned in the production of proper flavor and aroma in butter, and which are also concerned in the development of acid and possibly asso- ciated with formation of cheese flavor represent the sec- ond type. Many of these organisms are lactic acid pro- ducing, but in addition to these, some of the casein ferments are also associated with aroma production in butter. The third class includes those species that are able to produce deleterious and undesirable flavors in milk and milk products. The majority of the abnormal fermenta- tions of milk referred to in this chapter come under this Contamination of Milk. 53 head. Most of these gain access to the milk through slovenly and careless methods of handling. Those spe- cies associated with animal excreta are particularly dan- gerous. The number of different kinds that have been found in milk is quite considerable, something over 200 species having been described more or less thoroughly . In all probability, however, many of these forms will be found to be identical when they are subjected to critical study. CHAPTER V. MILK FERMENTATIONS AND THEIR TREATMENT. 56. Classification of milk fermentations. If any sample of milk is allowed to stand at any ordinary tem- perature for several days, a profound physical and chemi- cal change inevitably takes place. As a rule the milk will sour. In this process, certain acids are formed at the expense of the milk-sugar. If it is still allowed to stand for some time after it has become thoroughly sour, a series of subsequent changes usually occur. Often it will evolve foul- smelling gases and undergo putrefactive changes. Sometimes, however, other ferment organisms gain the ascendency over the ordinary souring process, in which case the fermentation is said to be abnormal. These peculiar changes are the cause of various taints in milk. They are of great importance because a small quantity of milk tainted in this way is liable to infect a much larger quantity and spoil the same through the rapid development of the obnoxious organism. In some cases, the abnormal condition may not become marked until the milk is made up into some other product as butter or cheese. The difficulties that occur under these conditions will be discussed under their appropriate heads. It is impossible in the present state of our knowledge to classify these fermentations in any other than a most provisional way. While our knowledge of some of them from a bacteriological standpoint is fairly complete, so many, as yet, have been studied only superficially that we are not in a position to classify them as we would other chemical changes. [54] Milk Fermentations. 55 The various fermentations may be grouped according to the substances in the milk upon which they chiefly act, such as those affecting milk-sugar, or casein. This arrangement is followed in this connection, but it should be borne in mind that many organisms act upon several of the milk constituents, and hence, it is difficult to correctly classify the same. Another method of classification is based upon the products manufactured during the fermentation, but even here confusion is liable to enter to some extent, because in many cases there are several distinct substances formed. Milk is such a complex substance that the changes pro- duced by a single germ are often so numerous that the processes can not be separated in their reactions. It must be remembered then, in referring to the different types of fermentations, that perhaps a distinct by-product is being formed, but it is more than probable that there are a series of changes, in which the most marked decom- position process is alone taken into consideration. For example, there is a fermentation classed under the head of the butyric changes, a decomposition process in which butyric acid is the chief product formed, but this may be associated with an alkaline condition of the milk and the production of a bitter substance in the same. Thus, the subdivision followed here will of necessity be imperfect and occasional instances will be noted where some changes in milk might well be. described under several heads. Some of these fermentation changes occur so con- stantly that they are to be regarded as normal or natural in their character. Normal, however, only in the sense that the bacteria that cause them are so widely distributed that the change is inevitable and not that the milk itself would undergo any of these changes if it were not for the presence of the different germs that bring them 56 Dairy Bacteriology. about. Of such a nature is the lactic acid fermentation, by far the most common change that occurs in milk. 57. Souring 1 of milk. Milk naturally undergoes a change known as souring, if allowed to stand for several days at ordinary temperature. This is due to the forma- tion of lactic acid, which is produced by the decomposition of the milk-sugar. While this change is wellnigh uni- versal, it does not occur without a pre-existing cause, and that is the presence of certain living bacterial forms. These organisms develop in milk with great rapidity, and the decomposition changes that are noted in souring are due to the by-products of their development. The milk-sugar undergoes fermentation, the chief pro- duct being lactic acid, although various other by-pro- ducts as other organic acids (acetic, formic, and succinic) , different alcohols, and gaseous products, as CO 2 , H, N, and methane (CH 4 ) are produced in small amounts. In the souring of milk, the formation of acid does not continue until the sugar is all exhausted. When the acidity reaches about 0.4% , milk begins to taste sour. In- cipient curdling takes place at about 0.6% , and soon after this, bacterial development is checked altogether. The acid formation goes on, however, until from 0.8 to 1.0% is reached. The amount of acid formed varies consider- ably in different cases. 1 If the acid produced in any case is removed by neu- tralizing it with a carbonate, its development will begin anew, showing that it is suspended by the inability of the organisms to grow in such a medium. Cream never contains as much acid as milk for the reason that a considerable proportion of its volume is oc- cupied by the butter-fat which is not subject to this de- composition. 1 Warrington, Journ. Chem. Soc., 53: 727, 1888. .Milk Fermentations. 57 It is a wide- spread belief that thunder-storms cause the premature souring of milk. Numerous experiments have been conducted along these lines, and the general con- clusion is that neither the electric discharge nor shock due to thunder exert any effect on the development of acid, but that the atmospheric conditions usually incident to a thunder-storm are such as permit a more rapid bac- terial growth. The lactic acid fermentation is produced by a large number of different kinds of bacteria, although in the spontaneous coagulation of milk, it is now believed that a very widely distributed species is responsible for the most of it. This organism was first described in a com- plete manner by Hiippe 1 and called by him B. acidi lac- tici. Giinther & Thierf elder 2 working on the spontaneous souring of milk in the neighborhood of Berlin found what they think is the same germ. Esten 3 , in this country, studied milks from thirty different localities in New England and the Middle States. He found a germ in all but two cases that agreed in general with Giinther 's description. Dinwiddie 4 , studying the same question in Arkansas, arrives at the same conclusion. This pre- ponderance of evidence makes it quite probable that there is a widely distributed germ that is concerned in this change although there are numerous other forms 5 that are associated with this type of decomposition. Conn and Aikman refer to the fact that over 100 species are already known. It is fair to presume, however, that a careful comparative study of these would show that 1 Huppe, Mitt. a. d. k. Gesund. Amte, 2: 309, 1884. 2 Giinther & Thierfelder, Arch. f. Hyg., 25: 164. 3 Esten, 9th Kept. Storrs Expt. Stat., p. 44, 1896. 4 Dinwiddie, Ark. Expt. Stat., Bull. 45, May, 1897. 5 Kayser, Ann. de 1'Inst. Past., 10: 737. 58 Dairy Bacteriology. simply racial differences exist in many cases, and there- fore, that they are not distinct species. As a rule this class of bacteria is unable to liquefy gelatin or develop spores. On account of this latter characteristic they are easily destroyed when milk is pas- teurized or heated to a higher temperature. They live un- der aerobic or anaerobic conditions, many of them being able to grow in either environment. According to Hoft 1 spontaneous souring occurs more rapidly in vessels hav- ing a small surface exposed to the air, indicating that anaerobic activity was more pronounced. The temperature conditions as to growth vary some- what with different species. With most species growth occurs at 50 F., but appreciable amounts of acid are not produced until a higher temperature is reached 2 . While the souring of milk is a very wide- spread phe- nomenon, still lactic acid germs do not abound every- where. Esten finds them abundantly in the milk- ducts but not present on hay. From the milk dealer's stand- point, this fermentation like all others, is undesirable, as it destroys the food value of milk for direct consump- tion. The fermented product, sour milk, has some value for cooking purposes. From this fermented product, the Armenians make a palatable drink, matzoon, that has considerable dietetic value in certain stomach troubles. While these bacteria are undesirable in milk- supplies, they are essential to the production of butter and cheese. The aroma and flavor of butter is, as a rule, dependent upon their presence, and in cheese they are absolutely essential in various stages of its manufacture. Milch Ztg., 26: 212, 1897. 2 Kayser, Cent. f. Bakt., II. Abt., I: 436. Milk Fermentations. 59 58. " Gassy " milks. This very common and undesir- able type of fermentation is caused by the presence of a large number of different bacteria, the majority of which belong to the lactic acid group. The amount of acid formed is always considerably less than that which is present in the typical lactic formation. Besides this, there is also produced as a result of the decomposition of the milk-sugar, various gases such as CO 2 , H, and meth- ane. Accompanying these are also to be noted various FIG. 12. A cheese made from "gassy" milk. A severe type of this fermentation. other decomposition substances that impart to milk and its products an undesirable flavor and odor. These fermenta- tions are particularly undesirable in cheese-making, as they are the cause of "floating" and "pin-holey 7 ' curds. 1 The swelling or huffing of green cheese is invariably due to this type of ferment action. Organisms producing this abnormal change are particularly numerous in ma- nure particles 2 , the colon bacillus (B. coli communisj y 1 Freudenreich, Landw. Jahr. d. Schweiz, p. 17, 1890. Russell, 12th Kept. Wis, Expt. Stat., p. 139, 1895. 2 Bolley, N. D. Expt. Stat., Bull. 21. 60 Dairy Bacteriology. the common fecal inhabitant of the intestinal canal being able to produce large quantities of gas.. In some cases gas- generating bacteria belong to the casein- dissolving group, but for the most part they attack the milk-sugar. 59. Slimy or ropy milk. The viscosity of milk is often markedly increased as a result of bacterial fermenta- tions. This condition varies much in intensity, in some cases the milk merely becoming viscous, when it is known as sticky or slimy; then again the viscosity may be so increased, that particles of milk, when touched, will cohere and string out in long threads, in which condition it is known as ropy, thready, or stringy. This con- dition in milk generally occurs during warm weather. Its presence prevents perfect creaming, as the globules of butter-fat are unable to rise, owing to the viscous nature of the fluid. Nat- urally it also has an injurious effect upon the quality of the cream, owing to the by-products that are formed. While a number of different species of bacteria have been more or less thor- " oughly studied which possess the prop- erty of rendering milk slimy, these various species are not of equal im- FIG. is. siimy milk. p Or tance in affecting milk under nat- This milk would "string * to . out" several feet in ural conditions. The manner in which length> the slime-forming organisms are first introduced into milk is of considerable importance, as a knowledge of this often enables restrictive measures to be applied directly. Marshall 1 reports an outbreak in Michi- 1 Marshall, Mich. Expt. Stat., Bull. 140. Milk Fermentations. 61 gan which he traced to an external infection from the udder. In another case, he found the slimy germ in the dust on the floor of a barn. Guillebeau 1 has reported a number of cases in which slime-forming organisms were traced to a diseased condition of the udder. In Switzerland, the chief cause of ropy milk seems to be Mic. Freudenreichii, a large, immotile, non-liquefying coccus that grows well at ordinary temperatures. This organism is readily killed by heat, two minutes at 212 F. being sufficient, but in a dried condition it has great resisting powers. The .slimy substance formed in milk comes from vari- ous ingredients of the milk, and the chemical character of the slime produced also varies with different germs. In some cases the slimy material is merely the swollen outer cell membrane of the bacteria themselves; in others it is due to the decomposition of the proteids, but in general, the chief decomposition product appears to come from a viscous fermentation of the milk-sugar. Normally this class is repressed by the development of the lactic acid bacteria; in fact, putrefactive processes sel- dom occur where the lactic organisms are in the ascend- ency. In sterilized or pasteurized milk this competition is removed as the lactic forms are unable to withstand this treatment, while this enzyme-forming group sur- vive by virtue of the spores which they possess. They gain access to the milk not infrequently from manure particles 2 that are derived from the coat of the animal. 60. Favorable slimy fermentations. While in this country slimy milks are considered as undesirable, yet in certain parts of Europe changes of this character are put to good use. In Holland it has long been the custom to 1 Guillebeau, Landw. Jahr. d. Schweiz, p. 27, 1890. * Backhaus, abs. in Expt. Stat. Rec., 10: 89. 2 Dairy Bacteriology. add a starter known as "lange wei" (long or stringy whey) , in the manufacture of Edam cheese to control the gassy fermentations. Weigmann has isolated from this material a slime-producing organism known as Strepto- coccus Hollandicus, which renders milk stringy and acid in a few hours at ordinary temperatures. The clotted or thickened milk known as ' ' taettemjolk, ' ' that is a favorite beverage in Norway, and the film j 61k (ropy milk) of Finland, is produced by adding leaves of common butterwort (Pinguicula vulgaris) to the milk. Weigmann 1 has isolated a bacterial form from the leaves of this plant that is similar to St. Hollandicus that is able to cause a slimy change in milk, so it is probable the change is due to the germs on the surface rather than to the plant itself. 61. Alcoholic fermentations. While fluids contain- ing ordinary sugar or glucose are very apt to undergo alcoholic fermentation if exposed to the air, milk-sugar does not decompose readily in this way. The alcohol- producing ferments are mainly yeasts which class of or- ganisms do not thrive readily in milk, although Duclaux 2 reports a serious outbreak in a dairy due to an organism of this class. Among some of the Oriental tribes, alcoholic bever- ages are used that are make from milk. Kumiss (spelt also koumiss, kumys), is made by a fermentation of mare's milk induced by the addition of old kumiss. A similar preparation intended for invalids is now made in this country from cow's milk by the addition of a small amount of cane-sugar and the subsequent intro- duction of yeast. 1 Weigmann, Milch Ztg., Beilage, No. 48, 1889. 3 Duclaux, Principles de Laiterie, p. 60. Milk Fermentations. 63 Another alcoholic beverage that is in common use among the people of Caucasus is kephir (also kefyr, kefir) . This is a sour, effervescent alcoholic fluid prepared from the milk of goats, cows, or sheep. The direct cause of the fermentation is the so-called kefir grain, a yellowish mass about as large as a walnut that is added to the milk. These grains are left in the milk for about a day; the milk is then poured off, and the grain dried and pre- served for future use. This milk after being mixed with fresh milk is kept in leather flasks and soon a mixed fer- mentation sets in. This alcoholic change has been studied considerably from a biological point of view 1 , but even yet is not thoroughly understood . It is evidently a mixed fermenta- tion, and one in which no single organism can produce all the essential ingredients. The sugar of milk is par- tially converted into alcohol, but at the same time the casein is coagulated and digested to a certain extent, so that the process is quite complicated. Some of the or- ganisms isolated have been found to be able to produce a change allied to that seen under natural conditions. A yeast form is probably the main cause of the alcoholic fermentation, while bacteria change the other constitu- ents of the milk. 62. Butyric acid fermentations. The fermentation characterized by the production of butyric acid is also a class fermentation that is caused by the action of a num- ber of different aerobic and anaerobic bacterial species. This decomposition process is common in boiled milk and as a secondary fermentation in sour milk. 2 For a long time it was thought that the butyric acid change in sour milk was a continuation of the lactic fermentation, but 1 Freudenreich, Landw. Jahr. d. Schweiz, 10: 1, 1896. 2 Hofmann, Ann. de Sci. Nat., 11: 5, 1869. 64 Dairy Bacteriology. it is now believed that these organisms find more favor- able growth not so much on account of the lactic acid formed as in the absence of dissolved oxygen in the milk which is consumed by the sour milk organisms. Most of the butyric class of bacteria are spore-bearing, and hence, they are frequently found in boiled or steril- ized milk . The by-products formed in this series of changes are quite numerous. In most cases, butyric acid is prominent, but in addition to this, other organic acids as lactic, succinic, and acetic are produced, likewise different alcohols. Concerning the chemical origin of butyric acid there is yet some doubt. Duclaux 1 affirms that the fat, sugar, and casein are all decomposed by various forms. In some cases, the reaction of the milk is alkaline, with other species it may be neutral or acid. This type of fermentation has not yet received the study it deserves. In milk these organisms are not of great importance, as this fermentation does not readily gain the ascendency over the lactic bacteria. It has been supposed until re- cently, that the rancidity of butter was attributable to the action of these ferments, but the general belief now is that this is a purely chemical change that may be in- fluenced but not caused by these organisms. 2 63. Bitter milk. A bitter taste may be imparted to milk in a variety of ways. In some cases it is due to improper feeding caused by eating herbs such as lupines, wormwood, or chicory. Then again, at certain periods of lactation, a bitter salty taste is occasionally noted in the milk that is peculiar to individual animals. A considerable number of cases of bitter milk have, however, been traced to bacterial origin. For a number of years the bitter fermentation of milk was thought to 1 Duclaux, Principes de Laiterie, p. 67. 2 Duclaux, Compt. rendu, 102: 1077. Milk Fermentations. 65 be associated with the butyric fermentation, but Weig- mann 1 showed that the two conditions were not depend- ent upon each other. He found that the organism which produced the bitter taste acted upon the casein. Conn 2 found a coccus form in bitter cream that was able to impart a bitter flavor to milk. The writer sep- arated a lactic acid species from milk that also possessed a similar property. Sometimes a bitter condition does not develop in the milk, but may in the milk products later. Freudenreich 3 separated a micrococcus from cheese that was found to be the cause of bitterness. Cream ripened at low temperatures not infrequently develops a bitter flavor, showing that the optimum tem- perature for this type of fermentation is below the typical lactic acid change. It has long been a question how to account chemically for the bitter taste in milk. Various ideas have been advanced, but Freudenreich has demonstrated in one case that a bitter substance is formed in the milk that can be isolated by adding alcohol. Milk that has been cooked is likely to develop a bitter condition. The explanation of this is that the bacteria producing the bitter substances usually possess endospores and that while the boiling or sterilizing of milk easily kills the lactic acid germs, these forms on account of their greater resisting powers are not destroyed by the heat. 64. " Sweet curdling 1 " and digesting fermenta- tions. Not infrequently milk instead of undergoing spontaneous souring curdles in a weakly acid or neutral condition, in which state it is said to have undergone 1 Weigmann, Milch Ztg., 1890, p. 881. 2 Conn, Kept. Storrs' Expt. Stat., p. 158. 1890. 3 Freudenreich, Fiihl. Landw. Ztg., 43: 361. 5-B. 66 Dairy Bacteriology. " sweet curdling. " In some cases the curdled casein may remain intact; in others it steadily diminishes in volume, a turbid and somewhat colored watery fluid separating from it. In this stage, the milk is said to have "wheyed off.' 1 The physical appearance of milk undergoing these fer- mentations is materiall}* different from that where normal souring occurs. The curd of sour milk is hard and breaks with a fractured surface, while the coagulum in these cases is soft and somewhat slimy. In the later stages of the digestive process, the milk assumes a watery appear- ance. These fermentations assume two phases: 1. Curdling followed by a subsequent digestion of the casein; 2. Digestion or peptonization of the casein without any ap- parent previous coagulation. A great variety of bacteria are able to participate in these changes, particularly those belonging to the class represented by the hay and potato bacilli. This group of bacteria as a rule are able to liquefy gelatin, a fermenta- tive change of a similar nature to the digestion of the casein. 1 In fact they are frequently referred to as casein- ferments. The characteristic of these fermenta- tions is the production of certain unorganized ferments or enzymes that have the power of acting on the proteid molecule independent of vital activity. The two ferments that are best known are the rennet or curdling enzyme, and the tryptic or digesting enzyme. As a rule any or- ganism that possesses the digestive power, first -causes a coagulation of the casein in a manner comparable to ren- net. Conn 2 has separated this enzyme in a relatively pure condition, and Fermi 3 has isolated the digestive 1 Sterling, Cent. f. Bakt., II. Abt.,1: 473. 2 Conn, 5th Rep't. Storrs' Expt. Stat., 196, 1892. 3 Fermi, Arch. f. Hyg., 14: 1, 1892. Milk Fermentations. 67 principle from a number of different species. Duclaux 1 has given to this digesting enzyme the name casease or cheese ferment. These isolated ferments when added to fresh milk possess the power of causing the characteristic curdling and subsequent digestion quite independent of cell development. The quantity of ferment produced by different species differs materially in some cases, the amount of rennet ferment being so imperceptible as to be obscured in the reaction by the digestive process. In these fermentations, the chemical transformations are profound, the complex proteid molecule being broken down into albumoses, peptones, amido- acids (tyrosin and leucin), and ammonia as well as fatty acids. Not infrequently these fermentations gain the ascend- ency over the normal souring change, but under ordinary conditions they are repressed, as they are unable to tolerate the lactic acid group. They are, however, present in all milks to a greater or less extent, as can be seen from the fact that boiled, sterilized, or -pasteurized milks invari- ably undergo this type of fermentation on account of the resistance of the spore-bearing species that remain in the milk after the lactic forms are killed. They are present in milks to a larger extent in summer than winter, on which account it is much more difficult to sterilize milk thoroughly during this season. Germs of this class are not only undesirable if they gain the as- cendency in milk, but where milk is made up into cheese, considerable loss occurs from the digestion of the casein, the peptonized portions being lost in the whey. 65. Soapy milk. Weigmann and Zirn 2 isolated from a milk having a soapy flavor, a specific germ, B. lactis saponacei, capable of imparting a taste of this sort. Milk 1 Duclaux, Le Lait. p. 121. 2 Weigmann and Zirn, Milch Zeit., 22: 569. 68 Dairy Bacteriology. affected in this way foams readily, and is abnormally slow in souring. They traced the cause of the trouble in one case to the infection of the milk from the straw that was used for bedding; in another, it was present in the hay. Marshall 1 in this country has also isolated an or- ganism of this sort that acts upon casein and albumen. 66.. Bloody or red milk. This condition often arises from the actual presence of blood in the milk due to some wound in the udder. The ingestion of certain plants, as sedges and scouring rushes, is said to cause a bloody condition in milk; madders impart a reddish tinge on account of a coloring matter absorbed. These instances can always be separated from bacterial troubles, because if due to these sources the color will be noted at the time of milking. There are several chromogenic or pigment-bearing bac- teria that have been isolated from milk, that have the power of turning milk red, although this change is so slow that it does not amount to much in dairy practice. The most widely known form able to bring about this change is Bacillus prodigiosus, the so-called "bleeding bread ' ' bacillus. It is interesting from a historical stand- point, inasmuch, as it has been found to be the cause of the bloody bread that has been the source of much super- stitious fear. Its growth in milk is marked by the pro- duction of a coloring matter that is diffused throughout the milk, especially near the upper surface. Free con- tact with oxygen is required to produce the characteristic pigment. Bacillus lactis erythrogenes (bacillus of red milk), is another form that grows easily in milk, produ- cing a red color, but it has the curious property of being able to form the color only in the dark, and in milk 1 Marshall, Mich. Expt. Stat., Bull. 146, p. 16. Milk Fermentations. 69 that is not strongly acid in its reaction. When grown in the light, this germ forms a yellow pigment. In milk, the casein is slowly precipitated and gradually dis- solved. It thrives at a high temperature, from 80-95 F . Other cases of red milk have been reported that have been traced to other germs. 1 Menge 2 found a red sar- cina that was the cause of trouble in a milk-supply. Here in this country, it is not at all uncommon to find on old milk, red patches that' are caused by the growth of a yeast-like germ, Saccharomyces glutinis, that is often found in the air. 67. Blue milk. Blue milk is historically a better known disease than almost any other trouble in milk. As long ago as 1838, Steinhoff showed that the trouble was communicable from one lot of milk to another. It manifests itself in the course of one to three days by the appearance of isolated flecks of bluish or grayish color on the surface of the milk. In fresh milk that is only slightly acid, the gray tints prevail, but as the amount of lactic acid increases in the milk, the blue coloration becomes more marked. So far only one form (Bacillus cyanogenus) is known that is able to produce this change. It does not materially affect the milk, but the butter made from infected cream has very poor keeping quali- ties. In Mecklenburg an outbreak of this disease once persisted for a period of several years 3 . Heim 4 found that this bacillus was especially resistant toward drying, or the influence of chemical agents like soda and potash, but a temperature of 176 F. for a moment sufficed to kill it. 1 Keferstein, Cent. f. Bakt. I Abt., 21: 177. 2 Menge, Cent. f. Bakt., 6: 596. 3 Fleischmann, Book of the Dairy, p. 51. 4 Heim, Arb. a. d. Kais. Ges. Amte, 5: 518. 70 Dairy Bacteriology. 68. Other kinds of colored milk. Two or three chromogenic forms producing still other colors have been found in milk. Schroeter discovered in a sample of cooked milk, a peculiar form (Bacillus synxanthus) that produced a citron yellow appearance and which precipi- tated and finally dissolved the casein. Adametz, Conn, and List have described other species that confer tints of yellow on milk. Some of these are bright lemon, others orange, and some amber in color. Still other color- producing bacteria, such as those that produce violet or green changes in the milk have been observed. In fact, almost any of the chromogenic bacteria are able to produce their color changes in milk as it is such an excellent food medium. Under ordinary conditions, these do not gain access to milk in sufficient numbers so that they modify the appearance of it except in occasional instances. 69. Treatment of milk fermentations. Attention has already been drawn to the distinction that should be made between taints due to fermentative action caused by the absorption of some pre-existing odor. In treating any abnormal fermentation, the attempt should first be made to locate the cause of the trouble. In most in- stances where the difficulty is due to bacteria, it is caused by foreign matter gaining access to the milk. Scrupulous cleanliness will therefore in most cases eliminate the trouble and the suggestions made in Chapter IV as to the methods of preventing bacterial infection will be help- ful in this connection. So efficacious is this course that cleanliness in every detail in dairy pursuits is almost a panacea for troubles and taints of all sorts that occur in milk. If the taint is recognized in the mixed milk of the herd, it is necessary to ascertain, first, whether it is a general Milk Fermentations. 71 trouble, or whether it is restricted to one or more ani- mals. For this purpose the fermentation or curd test in some form or other is invaluable. Often the whole milk- ing may be infected from the milk of a single animal, as is frequently the case in udder inflammation. To prove whether the trouble is a general or incidental one, can easily be done by separating the milk of the dif- ferent cows, or if this is not feasible by massing that of a few together, and so gradually narrowing down the number. Where the trouble is a general one, and is not due to the spreading of the infection from a local source, the fault is usually to be traced to some error in handling the whole mass of milk. Imperfectly cleaned cans that are used in setting the milk often contaminate the entire lot; then, too, noxious germs derived from the coat of the animal often gain access to the milk. The herd, es- pecially in the late summer, when the upland pastures are dry and the grass is short, are generally pastured on marsh or lowland fields. The stock seek the low places frequently slime-covered mud-holes, and in passing through these, their coats are fouled with the scum and slime that is filled with putrefactive forms of bacteria. Sometimes the source of the filth may be in the barn itself. Dirty stalls filled with moist and decaying mat- ter may be the means by which the milk is often seeded, or it may come from manure particles that contain putre- factive organisms in abundance. Generally where the source of contamination can be discovered, it will be an easy matter to get rid of the obnoxious fermentation by using physical means of disinfection such as steam or hot water. In some cases pasteurizing the milk is of material help. Chemical disinfection can sometimes be employed advantageously, but the application of these agents should 72 Dairy Bacteriology. be confined to the treatment of surroundings, rather than used in the milk itself. 70. Overcoming 1 taints by use of starters. An- other method that is often fruitful, is that which rests upon the inability of one kind of bacteria to grow in the same medium in competition with other species. Some of the undesirable taints in factories can be in large part controlled by the introduction of starters made from certain organisms that are able to obtain the as- cendency over the taint-producing germ. Such a method is commonly followed when a lactic ferment, either a commercial pure culture, or a home-made starter, is added to milk to overcome the effect of gas- generating bacteria. A similar illustration is seen in the case of the "lange wei" (slimy whey), that is used in the manufacture of Edam cheese to control the character of the fermentation of the milk. This same method is sometimes applied in dealing with certain abnormal fermentations that are apt to occur on the farm. It is particularly useful with those tainted milks known as "sweet curdling. n The ferment organ- isms concerned in this change are unable to develop in the presence of lactic acid bacteria, so the addition of a clean sour milk as a starter restores the normal condi- tions by giving the ordinary milk bacteria the ascend- ency. 71. Chemical disinfection. In only exceptional in- stances will it be necessary to employ chemical disinfect- ants to restore the normal conditions. Of course with such diseases as tuberculosis, very stringent measures are required, as they are such a direct menace to human life, but with these abnormal or taint- producing fermenta- tions, care and cleanliness, well directed, will usually overcome the trouble. Milk Fermentations. 73 In case it becomes necessary to employ chemical sub- stances as disinfecting agents, their use should always be preceded by a thorough cleansing with hot water so that the germicide may come in direct contact with the surface to be disinfected. It must be borne in mind that many chemicals act as deodorants, i. e., destroy the offensive odor, without de- stroying the cause of the trouble. Sulfur is often recommended as a disinfecting agent, but its use should be carefully controlled, otherwise the vapors have but little germicidal power. The common practice of burning a small quantity in a room or any closed space for a few moments, has little or no effect upon germ life. The effect of sulfur vapor (802) alone, upon germ life is relatively slight, but if this gas is produced in the presence of moisture, sulfurous acid (H 2 S0 3 ) is formed, which is very much more efficient. To use this agent effectively, it must be burned in large quantities in a moist atmosphere (three Ibs. to every 1,000 cubic feet of space) for at least twelve hours. After this operation, the space should be thoroughly aired. Formalin, a watery solution of a gas known as form- aldehyde is a new disinfectant that recent experience has demonstrated to be very useful. It may be used as a gas where rooms are to be disinfected, or applied as a liquid where desired. It is much more powerful in its action than sulfur, and it has a great advantage over mercury and other strong disinfectants, as it is not so poisonous to man as it is to the lower forms of life. Bleaching powder or chloride of lime (Ca01 2 ) is often recommended where a chemical can be advantageously used. This substance is a good disinfectant as well as a deodorant, and if applied as a wash, in the proportion of four to six ounces of the powder to one gallon of water, 74 Dairy Bacteriology. it will destroy most forms of life. In many cases it is inapplicable on account of its odor. Corrosive sublimate (HgCl 2 ) for most purposes is a good disinfectant, but it is such an intense poison that its use is dangerous in places that are at all accessible to stock. For the disinfection of walls in stables and barns, com- mon thin whitewash (CaOH) if made from freshly burned quicklime is admirably adapted. It possesses strong germicidal powers, increases the amount of light in the barn, is a good absorbent of odors, and is exceedingly cheap. Carbolic acid, creosote, and such products, while excel- lent disinfectants, cannot well be used in factories on ac- count of their odor. For gutters, drains, and waste-pipes in factories, vitriol salts (sulfates of copper, iron, and zinc) are often used These are deodorants as well as mild disinfectants. CHAPTER VI. DISEASE-PRODUCING BACTERIA IN MILK. 73. Milk a medium for pathogenic bacteria. Not only is milk an excellent food for fermentative bacteria, but a not inconsiderable number of disease organisms also find in it a suitable substratum for development. In so far as these occur in milk under natural conditions, they become a serious menace to public health. Statis- tical evidence shows that quite a large number of epi- demics have been traced to a contaminated milk-supply, which has served as a vehicle for the transmission of dif- ferent disease organisms. The following table, prepared by Freeman, represents the outbreaks that have occurred since 1880. Number of disease epidemics traced to contaminated milk- supply. Typhoid fever 53 Foot and mouth disease 2 Scarlet fever 26 Throat trouble 3 Diphtheria 11 Cholera 1 While such evidence can only be approximately correct, still the data at hand is overwhelming as to the relation of disease bacteria to milk. These organisms may be considered under the follow- ing heads : 1. Pathogenic bacteria causing diseases common to man and beast, which may be transmitted directly to man through the medium of the milk, as in tuberculosis. 2. Pathogenic bacteria that can thrive in milk under saprophytic conditions, and which gain access to it sub- sequent to its withdrawal, as in typhoid fever. [751 76 Dairy Bacteriology. 3. Saprophytic bacteria that can form toxic or poison- ous substances in the milk itself or in the intestine after it is ingested, as in certain types of cholera infantum. A. DISEASE BACTERIA DERIVED DIRECTLY FROM AF- FECTED ANIMAL. 74. Tuberculosis. Of those diseases that are com- municable from the animal to man by means of the milk, tuberculosis is by far the most common. This term is now used to indicate a number of maladies that have heretofore been claimed as separate diseases and which affect warm-blooded animals. It is now known that these different manifestations are all caused by the growth of the tubercle bacillus, which was discovered by Koch in 1882. In this connection, reference can only be made to the bovine type of the disease, and the relation that this bears in milk and dairy products to the human race. The disease is caused by the same germ whether it is present in the human being or the lower animals, 1 and the danger of infection exists in the transmission of the virus from one to the other. The organism causing this disease is remarkable for the narrow temperature limits within which growth will take place, the minimum being, according to Koch, 86 F., while the maximum is 104 F. This fact is of importance as it indicates that the tuber- cle bacillus is unable to develop, under normal tempera- ture conditions, in milk after it is drawn. The organism withstands drying readily; in fact, by virtue of this prop- erty, it is most widely disseminated, as the tubercular material is discharged from the diseased animal and is distributed in a dried condition in the dust. 1 Th. Smith has recently determined that there are certain varietal differences between these two types of tuberculosis. Disease Bacteria in Milk. 77 Putrefaction and decomposition even, will not quickly destroy its vitality. Direct sunlight is, however, an effi- cient disinfecting agent, and, even in diffused light the bacilli are destroyed in a few days. 75. Prevalence among* cattle. Tuberculosis not infrequently affects many species of warm-blooded ani- mals, but it is particularly prevalent among cattle in cer- tain regions of the world. The recent introduction of the tuberculin of Koch as an aid in the diagnosis of the disease in cattle has shown the trouble to be more widely spread than was at first believed, but sufficient data have not yet been collated to enable an accurate estimate to be made. In Denmark the percentage of affected animals has been shown by Bang to be very high, ranging from 30-40% . Concerning the distribution of the disease in this country, data is still very meager. In 15,000 head tested under the auspices of the U. S. Dept. of Agriculture, 19% showed a reaction. Unquestionably it is more widely disseminated, and a larger percentage of stock are affected in the older dairy regions of the east than in the west. The amount of disease among the range stock of the western plains is relatively small. 76. Tuberculin test. The tuberculin test is made by injecting into the animal a small quantity of a liquid known as tuberculin, which is a sterile glycerine extract of the growth products of the tubercle bacillus. The re- sults obtained by the use of this test show that it is far superior to the physical methods of examination, it being possible to detect the disease in its earliest stages. When tuberculin is inoculated into an animal it causes a febrile reaction in those cases affected with this disease; the rise in temperature usually exceeding the average normal 78 Dairy Bacteriology. temperature three or four degrees; in healthy animals only a slight rise occurs. A positive reaction should be at least from 2.2-2.5 F. above average normal, and should be maintained with some fluctuations for several hours. Animals in the later stages of the disease some- times do not respond, but when the test is properly ap- plied, it is on the whole a most satisfactory aid in diag- nosis. 77. Relation of disease organism to milk-supply. While tuberculosis is widely distributed among cattle, it FIG. 14. Side view of tuberculous udder, showing extent of swelling in single quarter. must not be supposed that the milk of all reacting ani- mals is infected with the specific organism. In a small percentage of cases, the udder and related tissues are af- fected, in which case the milk generally possesses in- Disease Bacteria in Milk. 79 factious qualities. In the early stages of the disease, the milk appears perfectly normal; as the disease progresses the milk assumes a watery appearance and changes in color. A diseased condition of the udder can usually be diagnosed where a hard, painless swelling of the udder occurs that is confined to one quarter. Udder inflamma- tions other than tubercular are generally accompanied with pain. Milk from animals having udder tuberculo- sis should be unconditionally rejected for food purposes. Woodhead 1 found in fourteen out of nineteen cases that milk from a tuberculous animal or the sediment from it was sufficiently infectious to produce the disease in guinea-pigs inoculated with small quantities of it. The writer 2 found one case in which a single cc. of milk from a diseased animal sufficed to kill a rabbit in- oculated with it. In this same case, the bacilli were also demonstrated microscopically in the milk. Sometimes the udder contains tubercle bacilli and still does not show any external symptoms of the disease. Bang 3 , Ernst 4 , and others have demonstrated that in quite a percentage of animals with apparently healthy udders, the milk possesses infectious properties. Where the disease is localized in the lungs, the danger from the milk is probably but slight, but it is often im- possible to determine the exact condition of the disease in the animal. Even where the disease is local there is a danger that a previous chronic condition may suddenly become acute. With adults in normal health, the danger from an in- fected milk-supply is undoubtedly greatly minimized, as 1 Woodhead, Trans. 7th Intern. Hyg. Cong., London, 1891. 8 Russell, llth Kept. Wis. Expt. Stat., p. 196, 1894. 8 Bang, Cong, for Tuberculosis, 1888, p. 70. 4 Ernst, Hatch Expt. Stat., Bull. 8, 1890. 80 Dairy Bacteriology. the healthy digestive tract is relatively insusceptible, but with infants and invalids the case is far different. The difficulty of proving an infection in this way is very great, as it is impossible to exclude the more common in- fection by the air; yet a number of well authenticated cases have been traced to this source. The presence of the disease in many cases in the digestive organs alone is inexplicable except in this way. It is impossible to de- termine what percentage of human tuberculosis is ac- quired in this way, but even though the amount be small, it devolves upon us to restrict this possibility in every conceivable way. Not alone should it be considered from the human standpoint, but from the point of view of successful animal industry must every measure be taken to repress and extirpate this disease from our herds. Tubercle bacilli in other milk products. If milk con- taining numerous tubercle bacilli is made up into differ- ent products the specific organism still is able to exist in such for a considerable length of time. Heim 1 found that tubercle bacilli were able to live in butter for' a month and in cheese for a fortnight, but the probability of infection occurring in this way is very slight as the quantity of these materials consumed at any one time is not large. 78. Methods of treating- milk. The possible danger from tuberculosis being spread by a contaminated milk- supply may be greatly diminished or entirely eliminated in the following ways: 1. Dilution. To produce infection it requires the simultaneous introduction of a number of organisms. Bollinger and Gebhardt 2 showed that milk from tuber- culous animals that would produce the disease in guinea- 1 Heim, Arb. a. d. kais. Gesundh., 5: 294. 2 Bollinger, Tagebl. d. 62 Versamm. Deutsch. Naturf., Sept. 1889. Disease Bacteria in Milk. 81 pigs and rabbits was innocuous when diluted with healthy milk to 50-100 times its volume. Therefore, there is less danger in mixed herd milks, than in that of a single cow, unless it is positively known that she is unaffected with the disease. 2. Destruction by heat. If milk containing tubercle bacilli is boiled, the disease organism is destroyed. The same result may be accomplished by pasteurizing if this process is correctly carried out. A temperature of 155- 160 F. for fifteen to twenty minutes is sufficient for this purpose. 3. It has been suggested that inasmuch as centrifuging seems to throw out many of the tubercle bacilli in the slime 1 that this method of purification could be used, but Moore 2 has shown that no reliance can be placed on this method. 79. Foot and mouth disease. This disease is dis- tinctively an animal malady, but it is also transmissable to man, although it is not often fatal. Hertwig 3 records some experiments made on himself. After drinking milk from an animal suffering from this disease, the mucous membrane of the mouth became swollen, small vesicles appearing in the mouth. Not an inconsiderable number of epidemics have been reported that have been traced to this source. 4 Calves readily acquire the disease suckling affected mothers. The law in Prussia since 1894 requires that the milk from all infected herds shall be heated to 194 F., for fifteen minutes before it is taken from the dairy. 1 Bang, Milch Zeitung, 1893, p. 672. 8 Moore, Year Book, Dept. of Agriculture, p. 432, 1895. 3 Ziemssen, Cyclop, of Prac. of Med., 3: 521. 4 Freeman, Med. Record, March 28, 1896. 6 B. 82 Dairy Bacteriology. 80. Other diseases. There are a number of other bo - vine diseases such as anthrax, 1 lockjaw, 2 and hydrophobia in which it has been asserted that the virus of the disease is transmissible to man, but in most cases of these diseases the secretion of the udder soon becomes affected, so that no danger need be apprehended unless such milks are wilfully sold for human food. The only safe rule is to reject milk coming from animals that show any signs of sickness, even though the disease may be one that is not common to man. B. INFECTION OF MILK BY DISEASE BACTERIA AFTER IT IS DRAWN. 81. Typhoid Fever. This disease stands next to tuberculosis in importance in its relation to milk. The organism producing this fever does not develop in the animal itself, consequently, no danger need be appre- hended from milk, if it is properly cared for after it comes from the cow. The typhoid fever bacillus, however, finds in raw milk such favorable conditions for develop- ment, that if it is once introduced, it is often able to thrive for a considerable length of time. The disease usually spreads by means of the water-supply, yet a large number of epidemics have been traced directly to milk as the original and only source of infection. A striking case is the Stamford, Conn., epidemic of 1895. 386 cases of the disease developed in six weeks, and of this number, over 97% came from a single milk-supply. The milk was infected by rinsing out the cans with water from a shallow contaminated well. The contamination of milk by this disease has been 1 Stohmann, Milch u. Molk. produkte, p. 382. 2 Marx, Deutsche Viertelsjahr. f. offentl. Gesunds. Pflege, 20: 444, 1890. Disease Bacteria in Milk. 83 traced to numerous causes, among which the following are most important: 1. Direct transmission to the milk from person conva- lescing from the disease. 2. Indirect transmission by milker also serving as nurse to patient. 3. Indirect transmission through polluted water-sup- ply where same is used for cleansing milk vessels. All of these sources of infection can be readily gov- erned with a little care. If typhoid fever is present in the family of a milk-handler, especial care should be taken that no one has any access to the patient that has anything to do with the milk. In case of disease in fam- ily, all water used in cleaning cans should first be boiled or else secured from a source in which contamination is impossible. Wells in vicinity of dwellings are often infected with disease germs that are derived from the ex- creta, and if such water is used, infection of milk- supply is possible. In some cases, the disease has been spread by infect- ing a general creamery supply, the contaminated skim- milk distributing the virus of the disease. In the Wei- ply epidemic in England in 1893, twenty-three cases appeared among the patrons of a single factory. The typhoid fever organism is able to thrive in milk for a considerable time (20-35 days according to Heim) * , on account of its tolerance toward weak acids. Even in butter and cheese these organisms are able to live, but the conditions are so unfavorable and the probability of infection so slight as to practically eliminate such a source . 82. Cholera, diphtheria, scarlet fever, etc. 1. Cholera. Milk also functions as a medium for the 1 Heim, Arb. a. d. k. Ges. Amte, 5: 303. 84 Dairy Bacteriology. transmission of cholera, although the danger from this source is somewhat minimized on account of the inability of this germ to thrive luxuriantly in acid fluids. Kitasato 1 found that the cholera germ could live in raw milk from one to four days, depending upon the amount of acid that is present in the m ilk . In boiled , or sterilized milk , cholera can grow more freely as the acid-producing organisms are destroyed by the heat used. In butter it has been found after four or five days, but the acid reaction re- strains its growth and soon kills it. Cholera epidemics in India have more than once been traced directly to contaminated milk. Simpson 2 records a case of infection where ten sailors out of twenty- four partook of some milk that had been purchased from a milkman, in whose district cholera had recently broken out. Of these ten, four died, five were severely sick, and one that used the milk sparingly was slightly ill. Investigation proved that the milk had been adulterated with water that had been taken taken from an open pool in the infected district. 2. Diphtheria. This is another contagious disease, the specific organism of which finds in milk favorable condi- tions of growth, and there is abundant evidence to indicate that milk functions as a transmitter of contagion if it once becomes contaminated with the same. Whether the animal can actually serve directly to spread the disease is yet a question. Klein 3 affirms as a result of animal inoculations that diphtheria will develop in the cow, at- tacking among other organs the udder, and so infecting the milk, but both Abbott 4 and Vladimirow 5 failed to confirm these observations. 1 Kitasato, Arb. a. d. Kais. Gesund. Amte, 1: 470. 8 Simpson, London Practitioner, 39: 144 (1887). 3 Klein, 19th Kept. Soc. Gov. Bd. (Great Britain), 1889, 167. 4 Abbott, Vet. Mag., 1: 17. 5 Vladimirow, Arch. sci. biol. Inst. Med. St. Petersburg, p. 84, 1892. Disease Bacteria in Milk. 85 3. Scarlet Fever. The exact relation of scarlet fever to milk is still harder to study, inasmuch, as the specific organism of this disease has not yet been isolated. Nev- ertheless, the evidence in a constantly increasing number of epidemics is so strong in favor of the view that dis- semination of disease occurs by means of polluted milk, that the relation may be considered as practically estab- lished. Klein 1 holds that a certain eruptive udder dis- ease of cattle is the same as scarlet fever, and that it is communicable to man. His conclusions, however, are not generally accepted, although it is regarded as thor- oughly proven that the disease may be propagated in the milk after it is drawn. C. POISON-FORMING BACTERIA IN MILK. 83. Toxic or poisonous milk. Milk not infre- quently acquires poisonous properties by virtue of the development of various putrefactive bacteria that form poisonous by-products as a result of decomposition pro- cesses. In some cases the toxic products are formed in the milk outside of the body. When such milk is in- gested, symptoms of poisoning soon appear. In other cases, certain bacteria found in the milk develop in the intestine producing a toxic effect. Milk contaminated with filth frequently contains putre- factive bacteria that form substances that cause gastric and intestinal troubles, especially in infants. Vaughan asserts that the larger number of cases of summer diar- rhoea are due to putrefactive organisms in the milk. The much higher mortality of bottle-fed, compared with breast-fed infants is undoubtedly attributable to the ac- tion of these microbes in milk 2 . 1 Klein, 7th Intern. Hyg. Cong. (London), p. 130, 1891. 2 Baginsky, Hyg. Rund., p. 176, 1895. 86 Dairy Bacteriology. Many of the cases of poisoning due to ice cream, cheese, and milk come from such a source. Improperly handled milk may be made up into these various products, and in this condition, the poisonous principle remain un- changed for a considerable time. Vaughan 1 has isolated a poisonous substance from these materials, calling it tyrotoxicon (cheese poison). Our knowledge concerning the way in which it is produced is, as yet, very vague, but from his most recent researches 2 , it is quite probable that the organisms causing these troubles are putrefac- tive forms that gain access to the milk where it is im- properly kept. 1 Vaughan, 9th Intern. Hyg. Cong., (London), p. 118, 1891. 2 Vaughan and Perkins, Arch. f. Hyg., 27: 308. CHAPTER VII. PRINCIPLES OF MILK PRESERVATION. 84. Keeping* quality. Where milk is consumed in the form of milk or cream, it is desirable to retard or in- hibit the fermentative changes as much as possible. In the preceding chapters, the direct relation which exists between the keeping quality of the milk and the germ life of the same has been indicated, so that it is evident, in order to perfectly preserve milk, the bacteria must either be prevented from gaining an entrance, or de- stroyed after they have once established themselves. Bacteria are so widely distributed, and milk is such a nutritious medium for their development, that to entirely prevent their entrance and growth is not feasible. Scru- pulous care in the dairy both before and after the milking, will however, do much to reduce the germ content of milk as is seen in the ' ' sanitary " or " certified ' ' milk that is being put on the market in some cities. These milks are drawn under the most careful conditions, and then held during distribution in such a way as to retard the course of the fermentative changes in a marked de- gree. Milk so treated will remain sweet for several days, which period is sufficiently long for practical purposes. Such milk cannot well be produced in herds that are not under one management as the degree of care necessary to success is not often exercised unless under the direct su- pervision of a competent manager. 85. Preservation of milk. In considering methods of milk preservation, reference will only be made to [87] 88 Dairy Bacteriology. those that are applicable to milk intended for food. Sam- ples destined for analytical use, such as for fat tests, etc., would be treated in a different manner. The various methods that have been suggested may be classified un- der two heads, those attained by the use of chemical, or by physical agents. In some methods the preservation is accomplished by the operation of both physical and chemical factors. 86. Chemical agents. Under the head of antiseptics and disinfectants, the action of different chemicals on bacterial life has been discussed 1 . Those substances that are inimical to the development of bacteria are usually too strong for use in a food product as preservatives, because the protoplasm of bacteria as a rule is more re- sistant than other forms of living matter. For this reason, the application of strong disinfectants like carbolic acid, mercury salts, strong acids, or alka- lies is excluded. The chemical agents that are used in the preservation of milk fall into two classes. 1. Those that unite chemically with certain products of bacterial growth to form more or less inert substances in the milk. 2. Those that restrain or inhibit the development of fermentative organisms in the milk. To this first class belong those alkaline salts such as bicarbonate of soda, etc., that combine with the acids that are formed in the milk. While salts of this sort neutralize the acidity produced by the sour milk bacteria, they do not kill these organisms, their growth again being renewed after the milk is neutralized. As representatives of the second class, salicylic acid, boracic acid, and their derivatives may be mentioned. T Lazarus, Zeit. f. Hyg., 8: 207. Principles of Milk Preservation. 89 Formalin has been extensively advertised of late, but even though this agent does not exert as prejudicial an effect on human tissues as some of the other disinfectants, yet it must be regarded with the remainder of chemical preservatives as undesirable. These substances can be added to milk in quantities not recognizable to the taste, and they will materially in- crease the time that milk will remain sweet, but their general use in milk is to be strongly deprecated. In many European countries their use is prohibited entirely. A large number of the preservatives are sold under pro- prietary names, and as a rule for extremely high prices, but they all depend for their efficiency upon the anti- septic action of chemicals. 87. Detecting- preservaline. The detection of these various preservatives in milk does not come within the general province of this work, but Farrington 1 has suggested such a simple means for the determination of the presence of preservaline and other boracic acid compounds that is so intimately connected with the de- velopment of acid that it is given here. When the acidity of a normal milk reaches 0.3-0.4%, it tastes sour, but when boracic acid is added to fresh milk, the acidity is much increased without affecting the taste. In fact, for some reason not yet explained, the acidity of the milk is increased considerably more than that due to the chemical added. Therefore milks having a high acid reaction (ex- ceeding 0.3-0.4% ) and not tasting perceptibly sour have in all probability been treated with preservatives. Failure to sour, especially when milk is kept at room temperatures for several days, is pretty conclusive evidence that chem- icals have been used. The sale of these preparations for use in milk finds its only outlet with those dairymen who 1 Farrington, Journ. Amer. Chem. Soc., Sept., 1896. 90 Dairy Bacteriology. are anxious to escape the exactions that must be met lay all who attempt to handle milk in the best possible method. Dirty milk drugged with these chemicals is fre- quently able to falsely pass as an extra good product. 88. Physical agents. The physical methods of milk preservation as a rule are much more feasible than the chemical, as they do not injure the nutritive value of the fluid to such an extent. The efficiency of physical meth- ods of preserving milk is dependent upon the elimination or destruction of bacterial life, or at least an inhibition of their powers of growth. Some methods, such as the use of electricity, or of compressed air, or different gases have been suggested, but a .practical treatment of milk with these agents has not as yet been devised. Different methods of nitration have been tried. Gravel or sand niters have been recommended for purifying milk- supplies. In these the suspended foreign matter, and a part of the bacteria can undoubtedly be removed, but the incon- venience of sterilizing such a filter would seem to be considerable. Filters of this character have, however, been satisfactorily used by the Copenhagen Dairy Co., for a number of years. Germ-proof filters, such as the Pasteur filter that are so efficient in water purification, are not applicable to milk as they clog so quickly. Recently the use of cellulose as a filtering substance has been proposed, and according to Backhaus and Comlien, 1 this is satis- factory from a bacteriological as well as a mechanical stand-point. 89. Freezing-. Chilling the milk has always been practiced as a means of enhancing the keeping quality, but recently the attempt has been made to transport milk for long distances in a frozen condition, in which state bacterial growth is entirely suspended. Milk can be pre- 1 Ber. Landw. Inst. Univ. Konigsberg, 2: 12, 1897. Principles of Milk Preservation. 91 served best by first pasteurizing it, then freezing it and shipping in this condition. 1 Another process is Casse's method in which a quantity of milk is frozen and placed in large cans. The intervening space is then filled with fresh milk and the whole closed air tight. It is claimed this milk keeps for weeks in this condition. 2 One of the difficulties of simple freezing is that the milk constitu- ents separate out so thoroughly that it is difficult to per- fectly re- incorporate them upon melting. 90. Condensed milks. Milks may be preserved for an indefinite period of time by condensing them. The keeping quality of such milk often depends, however, upon the action of another principle, viz., the inhibition of bac- terial development by reason of the concentration of the medium. This degree of concentration is reached in either of two ways, by the addition of sugar, or by evap- orating the watery solution by boiling it down either in open air or preferably in a vacuum pan. In the milks preserved by addition of sugar, the bacteria are not nec- essarily destroyed but they are unable to grow on account of the concentrated condition of the medium. Such milks diluted with several volumes of water, even sterile water, not infrequently undergo the ordinary decomposi- tion changes. In some cases, the heating process also destroys in part the contained germ life. 9 1 . High temperatures. Heat has long been used as a preserving agent. Milk has been scalded or cooked from time immemorial to keep it. Heat may be used at different temperatures, and when so applied exerts a varying effect, depending upon temperature employed. All methods of preservation by heat rest, however, upon the use of the two following principles: 1 Milch Zeit.., No. 9, 1895. 2 Milch Zeit., No. 33, p. 527, 1897. 92 Dairy Bacteriology. 1. A temperature above the maximum growing- point (42-45 C.) and below the the thermal death-point (52- 68 C.) will prevent further growth, and consequently fermentative action. 2. A temperature above the thermal death-point de- stroys bacteria, and thereby stops all changes. This tem- perature varies, however, with the condition of the bac- teria being much higher for spore-bearing species. Attempts have been made to employ the first principle, viz., prolonged heating above growing temperature, but when milk is so heated, its physical appearance is changed. The methods of heating most satisfactorily used are known as sterilization and pasteurization, in which a degree of tem- perature is used approxi- mating the boiling- and scalding-points respect- ively. 92. Characteristics of heated milk. When milk is subjected to the action of heat, a num- ber of changes take place in its reactions. 1 . Thinner body . Milk , FIG. 15. Microscopic appearance of nor- mal milk. The fat globules are grouped but more especially together into tiny clots. The consistency cream heated to a tem- or body 01 cream is in part due to this char- m acteristic. perature exceeding 150 F., becomes much thin- ner, a condition due to a change in the grouping of the fat globules. 1 In normal milk, the butter-fat is massed together in microscopic clots as in fig. 15. In heated 1 Babcock & Russell, 13th Kept. Wis. Stat., p. 73, 1896. Principles of Milk Preservation. 93 cream these clumps are broken down and the globules are homogeneously distributed as in fig. 16. 2. Cooked flavor. If milk is heated to 160 F., it ac- quires a cooked taste that becomes more pronounced as the temperature is further raised. The cause of this change is not well known. Usually it has been explained ' as being produced by changes in the nitrogenous ele- ments in the milk, particularly in the albumen. Re- cently , Thoerner 1 has pointed out the coincidence that exists between the appearance of a cooked taste and the loss of certain gases that are expelled by heating. He finds that the milk heated in closed vessels from which the gas cannot escape has a much less pro nounced cooked flavoi than if heated in an open vessel. 3. Fermentative changes. Normally milk undergoes the lactic acid fermentation. If, ever, it is heated to temperature above thermal death-point these non-spore-bearing organisms, as is the case ., -i FIG. 16. Microscopic appearance of milk in pasteurizing, it does heated to 150 o F or above< The aggrega . not SOUr but CUrdleS by tionof ifat globules that is present in normal j, , milk is broken down, the globules being means Ot a rennet ter- homogeneously distributed throughout the ment Which is Secreted mil ^ serum. This lessens the consistency of , . , , , , , cream and makes it appear thinner. by those bacteria that resist the heating process. 4. Action toward rennet. The action of heat causes the soluble salts of lime in the milk to be precipitated, and 1 Thoerner, Chem. Ztg., 18: 845. "94 Dairy Bacteriology. as the curdling of milk by rennet is dependent upon the presence of these salts, their absence in heated milks retards greatly the rennet action . The character of the coagulum is also considerably different from what it is with raw milk, being much softer and less tenacious. 5. Hydrogen peroxid test (Storch's test). When a small quantity of potassium iodid and starch is added to milk and the same then treated with a few drops of dilute hydrogen peroxid, normal milk will turn blue, while that heated to 175 F. or above, will not change. This reaction is due to the fact that the inherent enzymes normally found in milk are destroyed by this degree of heat, and therefore the hydrogen peroxid is not decom- posed 1 . 93. Sterilization. To thoroughly sterilize milk, it is necessary to render it germ free. So far as the vegetating bacteria are concerned, this can be done by an applica- tion of heat such as is used in pasteurizing, but on ac- count of the presence of spore-bearing forms, a much higher temperature is necessary. To render milk per- fectly sterile, it is necessary to heat it for about two hours at 250 F., or thirty minutes at 270 F. At the temperature of boiling- water, milk often withstands, es- pecially during the summer months, a prolonged steam- ing for five to six hours. Another method of sterilizing is that used in what is k:nown as intermittent sterilization, a method employed by Dahl, of Norway. This consists in heating the milk to a temperature fatal to the contained bacteria, 158 F. or above, for one-half hour in order to kill the vegetating bacteria. The milk is then placed under conditions that will cause the latent spores to germinate. After this oc- 1 V. Storch, 4.0th Kept. Danish Expt. Stat., Copenhagen, 1898. Babcock and Russell, 15th Kept. Wis. Expt. Station, p. 84, 1898. Principles of Milk Preservation. 95 curs, a second application of heat is made on the suc- ceeding day, and so on for three or four days. By this intermittent process, all spores are given a chance to germinate, and thus become susceptible to a relatively low heat. These methods which necessitate so protracted a treat- ment are manifestly out of the question for commercial purposes unless a product is desired that will keep for a considerable period of time. When milk is sterilized commercially, it is usually treated for a single time at a high temperature, and in some of the better types of apparatus is rendered nearly sterile, or at least the small amount of germ-life remain- ing is so weakened by the treatment given that develop- ment is suspended, unless the conditions are very favor- able. In most cases sterilized milks acquire a more or or less pronounced cooked taste. This is more of an ob- jection in America than in Europe, where in most cases milk is usually heated before being consumed. Appar- atus of this class is usually expensive, so that the cost of sterilized milk is higher than the normal product. Machines for this purpose have not been readily adopted in this country, the tendency being to employ the follow- ing process for milk preservation. 94. Pasteurization. In this method the degree of heat ranges from 140-175 F. and the application is made for only a limited length of time. The process was first extensively used by Pasteur (from whom it derives its name) in combatting the various maladies of beer and wine. Its importance as a means of increasing the keeping quality of milk was not generally recognized until a few years ago ; but the method is now growing rapidly in favor as a means of purifying milk for com- mercial purposes. The method does not destroy all 96 Dairy Bacteriology. germ-life in milk; it affects only those organisms that are in a growing, vegetative condition, but if the same is quickly cooled, it enhances the keeping quality very ma- terially. The experiments of Bitter 1 indicate that when stored at 86 F., properly pasteurized milk will remain sweet from six to eight hours longer than raw milk. At 77 F., ten hours; at 73 F., twenty hours; at 58 F., from fifty to seventy hours. In our experience, pasteurized cream when kept in an ordinary refrigerator usually re- mains sweet for a period of four to six days, and some- times even longer. 95. Requirements of Pasteurized milks. Pasteur- ized products should possess the following requirements : 1. Absolute freedom from disease bacteria. Fortu- nately the disease bacteria that are apt to be transmitted by means of the milk (tuberculosis, typhoid, etc.), do not form spores. Therefore, a proper pasteurization will destroy the virus of these diseases. 2 . Ordinary milk bacteria should be diminished . Proper pasteurizing will destroy all non- spore-bearing or vegeta- tive bacteria. 3. Improved keeping quality. This destruction of the normal milk flora will result in lengthening the time that milk will remain sweet. 4. Normal in taste and appearance. Pasteurized pro- ducts should have no perceptible cooked taste. They will, however appear thinner in consistency, a condition that often leads to the belief that they contain less fat. This is not a serious objection in the case of milk, but in cream it is more apparent. It can, however, be easily remedied by the use of viscogen (98). 1 Bitter, Zeit. f. Hyg., 8: 240, 1890. Principles of Milk Preservation. 97 ' 96. Pasteurized vs. sterilized milk. For consump- tion within a few days, pasteurized products are as well adapted as sterilized. For export and long- keeping milk, sterilized is better. Pasteurized is less objectionable than sterilized on account of cooked taste and flavor. As to the relative digestibility, authorities disagree. For the healthy child there is probably but little choice; for infants, pasteurized is believed to be better adapted. Pas- teurized products can be prepared and sold at less ex- pense than sterilized. 97. Children's milk. Attention has already been called to the relation which exists between infant mor- tality and the use of cows' milk. The cause of this lies in the fact as shown by Soxhlet that breast milk is con- sumed directly and in a condition practically sterile, although the researches of Knochensteirn show that it is quite as impossible to secure germ free mother's milk as it is cow's milk. The method of handling cow's milk is such that germ life finds ready access and opportunity to grow, with the result that infantile disturbances are com- mon where ordinary milk is used. Soxhlet introduced a method of sterilizing designed for family use that has proven very successful in Germany, where it is widely used. In sterilizing or pasteurizing milk for infant feeding, it is desirable that only a sufficient quantity for each day's use be prepared at once, for any milk that is not perfectly sterile will gradually develop the latent germ life that is in the same, and as these forms belong to the peptonizing class of organisms, their presence is not de- sirable, as has been shown by Fliigge 1 . Fliigge, Zeit. f. Hyg , 17: 272. 7-B. 98 Dairy Bacteriology. 98. Restoration of "body" of pasteurized cream. The action of heat causes the tiny groupings of fat glob- ules in normal milk (fig. 15) to break up, and with this change which occurs in the neighborhood of 150 F., the consistency of the liquid is diminished, notwithstanding the fact that the fat-content remains unchanged. Bab- cock and the writer l devised the following ' ' cure J ' for this apparent defect. If a strong solution of cane-sugar is added to freshly slaked lime and the mixture allowed to stand, a clear fluid can be decanted off. The addition of this alkaline liquid, which they call " viscogen," to pasteurized cream in proportions of about one part of sugar-lime solution to 100-150 of cream, restores the con- sistency of the cream, as it causes the fat globules to cluster together in small groups. The relative viscosity of creams can be easily deter- mined by the following method (fig. 17) : Take a perfectly clean piece of glass (plate or picture glass is preferable, as it is less liable to be wavy) . Drop on one edge two or three drops of cream at intervals of an inch or so. Then incline piece of glass at such an angle as to cause the cream to flow down surface of glass. The cream having the heavier body or viscosity will move more slowly. If several samples of each cream are taken, then the aggregate lengths of the different cream paths may be taken, thereby eliminating slight differences due to condition of glass. 99. Pasteurizing* details. While the pasteurizing process is exceedingly simple, yet in order to secure the best results, certain conditions must be rigidly observed in the treatment before and after the heating process. It is a mistaken idea that any milk is fit for pasteurizing. 1 Babcock and Russell, Bull. 54, Wis. Expt. Stat., also 13th Kept. Wis. Expt. Stat, p. 81, 1896. Principles of Milk Preservation. 99 The fresher and better the milk, the less likely it is to contain deleterious spore-bearing bacteria which are not destroyed by pasteurizing. The unhindered development of these germs in the milk may sometimes give rise to the production of decomposition products that are toxic or at least highly irritating in their character 1 . FIG. 17. Relative consistency of pasteurized cream, before (.A) and after (B treatment with viscogen. Cream flowing down inclined glass plate. 1 00. Selection of milk. Milk for pasteurizing should be as clean as possible, for in general terms, the bacteria associated with filth and dirt, more especially manure particles, belong to the spore-bearing putrefactive class. 1 Flugge, Zeit. f. Hyg., 17: 272. 100 Dairy Bacteriology. For high grade pasteurizing, it is advisable to purify the milk by passing it through a separator, unless the raw product is secured under conditions that exclude the ad- mission of dirt. The age of the milk and the conditions under which it has been kept should also be carefully noted. Old milk is almost always richer, not only in bacterial germs but in the latent spore forms as well; so the fresher the milk the fewer bacteria it will have, and therefore the pasteur- izing process will be the more complete. 101. Selecting" milk by acid test. The true stand- ard for selecting milk for pasteurization should be to de- termine the actual number of bacterial spores that are able to resist the heating process, but this method is im- practicable under commercial conditions. The following method while only approximate in its results will be found helpful. Assuming that the age or treatment of the milk bears a certain relation to the pres- ence of spores, and that the acid increases in a general way with an increase in age or temperature, the amount of acid present may be taken as an approximate index of the suitability of the milk for pasteurizing purposes. Biological tests were carried out in the author's labora- tory 1 on milks having a high and low acid content, and it was shown that the milk with the least acid as a rule was the freest from spore-bearing bacteria. This acid determination can be made at the weigh-can by employing the Farrington alkaline tablet which is used in cream-ripening. Where milk is pasteurized under general creamery conditions, none should be used con- taining more than 0.2% acidity. If only perfectly fresh milk is used the amount of acid will generally be about 0.15% with phenolphthalein as indicator. i Shockley, Thesis, Univ. of Wis., 1896. Principles of Milk Preservation. 101 Circumstances may arise that might lead to an error if this method is blindly followed. It has been pointed out that if milk is allowed to stand in rusty cans for some time its acid content is diminished materially. Bid- dick 1 has reported the following interesting observations. The average of nine samples of milk brought to a factory in clean cans contained .228% acidity, while that of nine other patrons brought in rusty cans contained only .134% acid. Milk kept in rusty cans is sure to contain large quantities of bacteria, even though its acid content may be low. & OunceBottle. Measure FIG. 18. Apparatus used in making rapid acid test. 102. Making- the acid test. Fig. 18 gives the ap- paratus necessary to make an approximate determination of the acidity of the milk. A solution of the alkaline tablets is first prepared by dissolving same in clean soft water, one tablet for each ounce of water; thus eight 1 Biddick, Hoard's Dairyman, July 30, 1897. /102 Dairy Bacteriology. tablets to an eight ounce bottle of water. In determin- ing the acidity in each patron's milk, a number of com- mon white cups are used, one for each patron. Two measures full of the alkali solution are placed in each cup, and then as the milk is received at the weigh- can, one- half as much milk is added to the alkali solution in the cup, 1 and the whole gently, but thoroughly shaken. If the pink color of the alkali solution persists even faintly, it shows that there is not enough acid in the milk to neu- tralize the same; if it disappears altogether, leaving the milk white in color, it indicates that there is more acid in the milk than can be neutralized by the alkali of the tablet solution. Any standard desired can be chosen, but where the relation of the milk to the alkali solution is maintained in above ratio (two to one), it indicates that 0.2% acidity is present, if the alkali is completely neutralized. Extended experience has shown the neces- sity of selecting milks for pasteurizing that come within this standard. 103. Temperature and time limits. The time and temperature limits in pasteurizing are subject to consid- erable variation. The minimum temperature, however, must exceed the thermal death-point at which the milk bacteria are destroyed. As the tubercle bacillus is some- times found in milk, and as it is one of the most resist- ant organisms in its vegetative state that is known, the thermal death-point of this germ serves as a minimum standard for efficient pasteurizing. The determination of this point depends on the follow- ing conditions: 1. The temperature of heat used. 2. The length of exposure to the heat. 1 Farrington, Wis. Expt. Stat., Bull. 52. Principles of Milk Preservation. 103 With either condition fixed, the same result is accom- plished more rapidly by an increase either in temperature or duration of exposure, so that an exposure for a longer time at a lower temperature is quite as effective as a shorter exposure at a relatively higher temperature. With the great majority of bacterial species that have been individually tested as to their thermal death-point, 140 F. for ten minutes has been found fatal. This de- gree of heat suffices to kill all the disease-producing bac- teria that are found in milk, with the exception of the tubercle bacillus. According to Forster, heating thirty minutes at 149 F. , fifteen minutes at 155 F., or ten minutes at 167 F. suffices to destroy this germ. The maximum temperature that can be employed is just below the point at which the milk will permanently ac- quire a cooked flavor. The appearance of this peculiar flavor cannot be detected with absolute accuracy, but it is not far from 158 F. It appears in milk to some ex- tent before this temperature is reached but upon chilling disappears. It is not enough to heat milk to this tem- perature, but it should be maintained for the requisite length of time. 1 04. Subsequent chilling". The heating process de- stroys only the vegetative organisms, the spores resisting this temperature. To prevent the germination of these latent forms the milk should be quickly chilled, for if ger- mination once occurs, they can develop' at even low tem- peratures. The following experiments by Marshall 1 are of interest as showing the influence of refrigeration on germination of spores. 1 Marshall, Mich. Expt. Stat., Bull. 147, p. 47. 104 Dairy Bacteriology. Cultures of organisms that had been isolated from pasteurized milk were inoculated into bouillon. One set was left to grow at room temperature, another was pas- teurized and allowed to stand at same temperature, FIG. 19. Diagram showing temperature changes in pasteurizing, and the re- lation of same to bacterial growth. Shaded zone represents limits of bacterial growth, 10-43 C. (50-109 F.); the in- tensity of shading indicating rapidity of development. The solid black line shows temperature of milk during the process. The necessity for rapid cooling is evident as the milk falls in temperature to that of growing zone. while another heated set was kept in a refrigerator. The unheated cultures at room temperature showed evi- dence of growth in thirty trials in an average of 26 Principles of Milk Preservation. 105 hours; 29 heated cultures at room temperature all de- veloped in an average of 50 hours, while the heated cultures kept in refrigerator showed no growth in 45 days with but four exceptions. 105. Preparation of utensils. After the milk is pasteurized, it must of necessity be stored and handled H.S. S.TP FIG. 20. Steam sterilizer for sterilizing utensils (cans, bottles, etc.). St. p., steam pipe; St. p. v., vent for steam pipe; w. s., wire shelf; c, outlet cock for condensed water. in germ free receptacles. All utensils such as cans, dip- pers, bottles, etc., must be thoroughly sterilized. For this purpose a sterilizing oven should be had which is steam fitted, so that direct steam can be used. Material of this sort after being thoroughly cleansed, should be steamed for one-half to three-quarters of an hour. Ster- ilized bottles should be kept protected from dust until they are used. 106 Dairy Bacteriology. 106. Bottling" and handling- the product. In bot- tling the product it is necessary to keep the milk pro- tected from reinfection. It may be bottled from a large can with a bottom faucet, or, on a large scale, with com- mercial bottling machines that fill several bottles at once. If "viscogen" is added to restore consistency of cream, it should be done before bottling, but not before the cream is thoroughly cooled. The best bottles for the purpose are those that have a plain pulp cap. All metal fastenings or stoppers are dirt catchers and are likely to get out of order. It is bur practice to heat pulp caps in paraffin, thereby rendering them more pliable and at the same time sterilizing them. Bottles sealed with hot caps in this way are tightly closed. In delivering pasteurized products, it is always neces- sary to use care in handling to prevent the cream and milk from being warmed up, and thus inciting into activity the latent spores. 107. Pasteurizing 1 apparatus. If it is desired to pasteurize milk or cream for direct consumption, the treatment differs somewhat from that when pasteurizing for butter- making is the object in view. The equipment necessary for the first type of pasteurizing may be divided into two general classes. 1. Apparatus of limited capacity designed for private family use. 2. Apparatus of sufficient capac- ity to pasteurize on a commercial scale. 108. Domestic apparatus. Pasteurization can be easily and efficiently done in a limited way with the addi- FIG. 21. Sectional view of family pasteurizer.show- ing milk bottles immersed in water. Principles of Milk Preservation. 107 tion of an ordinary dairy thermometer to the common utensils found in any kitchen. Fig. 22 indicates a sim- ple contrivance than can be readily arranged for this purpose. FIG. 22. A home-made pasteurizer. The following suggestions indicate the different steps of the process: 1. Use only fresh milk for this purpose. 2. Place milk in clean bottles or fruit cans, filling to a uniform level. If pint and quart cans are used at the same time, an inverted dish or piece of wood will equal- ize the level. Set these in a flat-bottomed tin pail and fill with warm water to same level as milk. An inverted pie tin punched with holes will serve as a stand on which to place the bottles during the heating process. 3. Heat water in pail until the temperature of same reaches 160 F. ; then remove from source of direct heat, 108 Dairy Bacteriology. cover with a cloth or tin cover and allow the whole to stand for half an hour. 4. Remove bottles of milk and cool them as rapidly as possible without danger to bottles and store in a re- frigerator. 109. Apparatus for commercial pasteurizing 1 . Pasteurizing as applied to the preservation of milk, orig- inated in Germany and Denmark, where it is used largely in the treatment of skim-milk, and the heating of cream in butter- making; The type of machinery that has been devised for this purpose has therefore naturally been con- structed on somewhat different principles than would have been employed where milk is treated for direct con- sumption. For the treatment of milk for direct con- sumption, it is believed that the sanitary phase of the problem demands more attention than the economic oper- ation of the apparatus, and, therefore, it is more neces- sary to use a machine that pasteurizes so thoroughly that all possible disease bacteria are destroyed, than it is to secure apparatus that will permit of the handling of large quantities of milk. Pasteurizing involves considerable time and trouble, and it is better not to have the process done at all than to have it imperfectly performed. The various types of machinery that have been sug- gested for this use may be grouped as follows, depending upon their method of operation 1 . 1. Continuous flow machines. 2. Intermittent machines. 110. Continuous flow pasteurizers. Apparatus of this class varies much in detail, but they possess this common principle that the milk enters the machine in a 1 For the detailed description of pasteurizing machinery, reference should be made to Monrad's Pasteurization of Milk, or Weigmann's Conservierung der Milch. Principles of Milk Preservation. 109 continuous stream and is generally discharged in the same way. The objection to this type of apparatus is that the time of heating cannot be regulated with any certainty, although the temperature can be controlled FIG. 24. Hochmuth's combined heater and cooler. Cold water enters at k and circulates through coil, cooling the milk which is heated in upper part of ribbed surface. This warm water is used to warm the milk, and then steam is intro- duced through d. in part by varying the speed of flow. Another objection is that the rapid heating often necessitated by these ma- chines scalds the proteids of the milk, making them ad- here to the walls of the pasteurizer. 110 Dairy Bacteriology. In some of these machines (Thiel, Kuehne, Lawrence, De Laval, and Hochmuth), a ribbed surface is employed over which the milk flows, while the opposite surface is heated with hot water or steam. Monrad, Lefeldt and Lentsch, employ a centrifugal apparatus in which a thin layer of milk is heated in a revolving drum. FIG. 25. Monrad's centrifugal pasteurizer. Milk is introduced into center through^, and is thrown out on the walls of centrifuge d, the steam being ap- plied to the outside of the revolving drum. Reservoir m catches and holds the milk for a moment. Milk finally flows at h. Some of these machines are suitable for heaters, if the milk was held in vats that would retain the temperature at a pasteurizing point for a sufficient length of time. Other machines of the continuous flow type employ a Principles of Milk Preservation. Ill reservoir that is filled with milk and surrounded with an outer shell that contains the heating agent, steam or hot water. Some of this class are provided with an agitator in the milk reservoir so as to hasten the equalization of tem- perature in the inner chamber, and at the same time keep the milk in motion in order to prevent the coagulation of the proteids. Most of them are arranged for a continuous delivery, the milk flowing in at the lower end and displacing that already pasteurized, which flows out above into a cooler. In some of them the agitator even mixes the fresh supply with that which has already been heated, so that the effi- ciency of the process where milk is being treated for direct consumption, is much lessened. Where it is constructed for continuous delivery, the length of exposure must necessarily be quite limited, and as the temperature of the milk ought not to exceed 160 F. for fear of scalding, it very often happens that the pasteurizing process is not efficient. In the case of the sour milk organisms, from the hygienic standpoint, it is of little moment, but to insure absolute freedom from disease germs, the temperature and the time of exposure must be thoroughly under control. Very few of the ma- chines intended for this purpose have been subjected to a rigid bacteriological test, and the lack of this has allowed the introduction of many designs that may be adapted for the pasteurization of by-products intended for animal food, for which purpose many of them were originally designed, but they certainly do not deliver a product that can be relied upon for human food. 111. Intermittent pasteurizers. Inasmuch as the biological and physical requirements as to pasteurizing, necessitate milk being heated between the temperatures 112 Dairy Bacteriology. of about 145-160 F., it is desirable that the temperature should be under absolute control. Moreover the time limit must also be known. This requires the treatment of a definite quantity of material for a definite length of time at a definite temperature. A fulfillment of these condi- tions necessitates the use of the intermittent type of apparatus, or continuous apparatus arranged so as to practically conform to the discontinuous process. The simplest way in which these conditions can be car- ried out is to employ a number of shot-gun cans immersed in a tank of hot water. By means of this crude device milk or cream can be pasteurized more effectually than in many of the specially designed pieces of apparatus. Tanks surrounded with water spaces can also be used quite successfully, although from a commercial point of view apparatus of this sort is imperfect, unless it is spe- cially designed for this purpose. The use of the Boyd cream ripening vat has been sug- gested, and this fulfills the necessary conditions as to a commercial pasteurizer. The cream in this is heated by means of a swinging coil immersed in the same, through which hot water circulates. In some of the pasteurizers, steam is introduced di- rectly into the milk or cream, as in Bentley's apparatus. It is obvious that while such a method may be a cheaper way in which to heat the milk, still the proteids of the fluid must be scalded in part, although the temperature of the whole mass may not exceed the proper pasteur- izing point. The writer 1 in 1894 devised a tank pasteurizer that was made to conform to the bacteriological requirements. It consists of a long, narrow vat, surrounded by a water chamber, on the bottom of which is placed a row of per- 1 Russell, Wis. Expt. Stat., Bull. 44. Principles of Milk Preservation. 113 f orated steam pipes. To facilitate the heating of the milk, both the milk and water reservoirs are supplied with agitators having a to and fro movement. The milk stirrers are perforated so as to give an up and down FIG. 26. Russell's pasteurizing vat. movement, thus preventing the cream from rising to the top of the vat during the operation. The milk is with- drawn from the vat by means of a stop- cock that is placed inside of the water chamber. This cock has a circular bore so that when open, the outlet tube presents a continuous passage that can be easily and thoroughly cleaned. Placing the stop-cock within the water chamber prevents the accumulation of unpasteurized milk in the outlet tube at a point not reached by the heated water, and where it would contaminate the whole vat when the milk was withdrawn. The Potts pasteurizer is another machine of the inter- mittent type that has recently been introduced that con- forms to the necessary biological conditions. This ap- paratus has a central milk chamber that is surrounded with an outer shell containing hot water. The whole machine revolves on a horizontal axis, and the cream or milk is thus thoroughly agitated during the heating pro- cess. 8 B. 114 Dairy Bacteriology. 112. Coolers. A speedy cooling of the heated pro- duct is essential to success in pasteurizing. Some of the machines have been devised for a combination purpose, being used for the heating and subsequent cooling of the FIG. 27. Pott's pasteurizer. milk. This is an evident advantage in some ways, as it lessens the amount of apparatus necessary, also the work involved in cleaning the same, but at the same time the problems of quick heating and cooling involve somewhat different principles, so that for the most economical manipulation of the product, separate pieces of appara- tus are advisable where the business warrants such ex- pense. The simplest method of treatment in cooling is to draw off the milk in shot-gun cans and place these first in water, then in ice -water. To cool milk most economically, two coolers should be provided. One of these can use cold water, and by the aid of good well water, the temperature can be reduced to nearly that of the water in a short time. In order, however, to lower the temperature below a point where spore germination will readily occur, milk should be chilled by the aid of ice. This may be applied in the same cooler as is used for running cold water, by sup- plying ice- water for the latter part of the cooling process. Principles of Milk Preservation. 115 To use ice economically, the ice itself should be applied as closely as possible to the milk to be cooled, for the larger part of the chilling value of ice comes from the melting of the same. To convert a pound of ice at 32 FIG. 28. Water cooler, using either running cold water, or water and crushed ice. The milk is introduced at i. m. and is spread out in m. c. in a thin cylindri- cal sheet, flowing out at o. m. Cold water circulates through inside of milk cylinder in w. c., while ice may be used in outer water chamber, o. w. c. F. into a pound of water at the same temperature re- quires as much heat as would suffice to raise 142 pounds of water one degree F., or one pound of water 142 F. The absorptive capacity of milk for heat is not quite the same as it is with water. It fluctuates with the amount of solids in the milk, but for ordinary milk is about .85 while water is taken as a standard 1.0. Hot milk would therefore require somewhat less ice to cool it than would be required by an equal volume of water at same tem- perature. In the mere melting of a pound of ice, a large part of the heat in a pound of pasteurized milk will be absorbed. To take advantage of this, the ice should be brought in close contact with the milk rather than to spend all of this absorptive capacity on cooling water which is later applied to the milk. If broken ice is used directly, it should be arranged so that the milk surrounds it, as in this way the specific capacity for heat that is latent in the ice acts on the milk instead of being radiated in part to the outside. 116 Dairy Bacteriology. 113. Bacteriological study of pasteurized milk. An extended bacteriological examination of milk and cream pasteurized on a commercial scale in the Russell vat at the Wisconsin Dairy school showed that over 99.8 FIG. 29. Effect of pasteurizing on germ content of milk. Black square repre- sents bacteria of raw milk; small white square, those remaining after pasteuri- zation. % of the bacterial life in raw milk or cream was destroyed by the heat employed, i. e., 155 F. for twenty minutes duration 1 . In nearly one-half of the samples of milk, the germ content in the pasteurized sample fell below 1,000 bacteria per cc., and the average of twenty-five samples contained 6 , 140 bacteria per cc . In cream the germ content was higher, averaging about 25 , 000 bacteria per cc . When the high initial content of either milk or cream is taken into consideration, it indicates that the spore-bearing bacteria are relatively few in properly selected milk that 1 Russell, 12th Wis. Expt. Stat. Kept., p 160, 18 ( J5. Principles of Milk Preservation. 117 is efficiently pasteurized. The kinds that are destroyed are mainly the lactic acid species; those that resist the pasteurizing heat are generally of the enzyme-forming class, which in many cases are also able to liquefy gelatin. The acidity of pasteurized products always increases some- what, but even when it is curdled, it rarely contains over 0.3-0.4 % of acid. The organisms that remain belong then to the sweet curdling type of bacteria. While these organisms capable of resisting the pasteur- izing temperature are fermentative germs, it is also im- portant to determine if they have the power of forming toxic substances that exert a harmful effect on the ani- mal body. Fluegge 1 has found several forms. in sterilized milk that are able to produce highly toxic substances. A similar study has been made under the writer's direc- tion on the bacteria isolated in pasteurized milk under commercial conditions. Shockley 2 studied thirteen differ- ent forms found in pasteurized milk, but all of them with a single exception failed to cause any disturbance when inoculated into white mice and rabbits. In the single instance, fatal results were only obtained when large quantities were inoculated. 1 Fluegge, Zeit. f. Hyg., 17: 272, 1894. 2 Shockley, Thesis, Univ. of Wis., 1896. PART III. RELATION OF BACTERIA TO MILK PRODUCTS. CHAPTER VIII. BACTERIA IN CREAM AND FACTORY BY-PRODUCTS. 114. Bacteria in cream due to mechanical causes. Cream whether secured by the gravity process or by a cream separator is invariably richer in bacteria than the skim-milk of the same age. A sample of milk might contain less than 100,000 germs per cc. in the skimmed part, while the bacterial contents of the cream layer in the same would be several millions for the same unit of volume. This is largely due to the filtering out of the microbes during the process ' of cream- ing. It is a well-known fact in sewage filtration that the addition of some chemical that will cause the pre- cipitation of certain organic elements always present in the sewage, will take out the majority of bacteria in the set- tling of thispercipitate. The same principle seems to be operative in the process of creaming, except that the fat globules instead of sinking to the bottom rise to the surface. Gravity-raised cream is usually richer in bac- teria than separator cream, because it is materially older when gathered. In full milk separated by the centrifugal method, there are three well-marked products, the skim-milk, the cream, and the slime that adheres to the separator bowl. [118] Cream and Factory By-Products. 119 The bacterial content of each of these will vary materially in different cases. The slime which is composed of particles having a greater specific gravity than the milk serum is thrown out on the edge of the revolving fluid by centrifugal force. This material, if examined microscopically, will be found to contain large quantities of foreign matter as well as innumerable bacteria. The fact that it rapidly undergoes decomposition is evidence of its high germ content. The cream will almost always contain a larger number of bacteria than the skim-milk. Popp and Becker found in a sample of whole milk containing 73,000 germs per cc. , the following germ contents after it had been separated: Cream, 58, 275 germs; skim-milk, 21,700 germs, and the separator slime 43, 900 per cc. This centripetal niovement of a large number of germs with the cream indicates that they adhere to the tiny fat globules, for this peculiarity in distribution can hardly be explained on the ground of their specific gravity, as they remain in the skim-milk in considerable numbers in spite of the great centrifugal pressure . 115. Tubercle bacillus and separator slime. Ac- cording to Scheurlen 1 and Bang 2 , tubercle bacilli, if present in a milk are largely thrown out with the slime in the separating process. Moore 3 found in milk arti- ficially infected with tubercle bacilli that the separating process diminished them to such an extent that they could not be determined microscopically, but when this separated milk was inoculated into guinea-pigs, infection occurred. This indicates that while the removal was con- siderable, yet it was not complete enough to justify the 1 Scheurlen, Arb. a. d. k. Ges. Amte, 7: 269, 1891. 2 Bang, Land. Woch. f. Sch. Hoi., 1894, p. 47. 3 Moore, Year-book of U. S. Dept. of Agri. 1895, p. 432. 120 Dairy Bacteriology. use of this method for the purification of infected milk. Coupled with this peculiar relation of the tubercle germ to centrifuge slime, is the fact that tuberculosis among swine is much more prevalent in Denmark and North Germany where the centrifugal process in creaming is extensively used, and where, until recently, the swine were fed the uncooked separator slime. Ostertag 1 has pointed out this condition, and has drawn attention to the nu- merous cases of intestinal tuberculosis in hogs. 116. Bacterial changes in cream. Although cream is numerically much richer in bacteria than milk, yet the changes due to bacterial action are so much slower that the latter product usually spoils sooner than cream. For this reason, cream will sour sooner when it remains on the milk than it will if it is separated as soon as possible. This fact indicates the necessity of early creaming, so as to increase the keeping quality of the product, and is an- other argument in favor of the separator process. 117. Bacteria in different creaming' methods. The method used in creaming has an important bearing on the kind as well as the number of the bacteria that are to be found in the cream. The difference in species is largely determined by the difference in ripening tempera- ture, while the varying number is governed more by the age of the milk. 1. Primitive gravity methods. In the old shallow pan process, the temperature of the milk is relatively high, as the milk is allowed to cool naturally. This compara- tively high temperature favors especially the develop- ment of those forms whose optimum growing-point is near the air temperature. By this method the cream layer is exposed to the air for a longer time than any 1 Ostertag, Milch Ztg., 22: 672. Cream and Factory By -Products. 121 other, and consequently, the contamination from this source is greater. Usually, cream obtained by the shal- low pan process will contain a larger number of species and also have a higher add content. 2. Modern gravity methods. In the Cooley process, or any of the modern gravity methods where cold water or ice is used to lower the temperature, the conditions do not favor the growth of a large variety of species. The bacterial numbers in the cream will depend largely upon the way in which the milk is handled previous to setting. If milked with care, and kept so as to exclude outside contamination, the cream will be relatively poor in bac- teria. Only those forms will develop in abundance that are able to grow at the low temperature at which the milk is set. Cream raised by this method is less frequently infected with undesirable forms than that which is creamed at a higher temperature. 3. Centrifugal method. Separator cream should be freer from germ-life than that which is secured in any other way. It should contain only those forms that have found their way into the milk during and subsequent to the milking, for the cream is ordinarily separated so soon that there is but little opportunity of infection, if care is taken in the handling. As a large part of the infection of fresh milk is due to the contamination from the fore milk, which usually has but a few species, separated cream, if handled with caution, will contain mainly those species that are to be found in abundance in the milk while it is still fresh. Where milk is separated, it is always prudent to cool the cream, as the milk is generally heated before separa- ting in order to skim efficiently. 118. Factory by-products. While the by-products in the manufacture of butter and cheese are in a certain 122 Dairy Bacteriology. sense waste products, yet, all of them possess sufficient nutrient value to warrant their use as human food or in animal feeding. When handled carelessly, however, their nutritive value is much lessened by the continued fer- mentations that occur in them. All of these products are rich in bacteria owing either to their age or the treat- ment they have undergone in the manufacture of butter or cheese. It is, therefore, all the more essential that they should be kept in such a manner as to check the continued development of germ-life within them. 119. Skim-milk. Skimmed milk varies much in its bacterial content, depending upon the way in which the full milk has been treated. Milk from which the cream has been removed by the shallow setting process is usu- ally very rich in germs, and often has so much acid that it is easily recognized by the taste. Where the cream is gathered by the aid of ice-water, the temperature is re- duced to such an extent that the skimmed part is rela- tively poor in bacteria. Separator skim-milk is treated in such a radically different way that it is bacteriologic- ally quite a different product. The skimmed part is sep- arated from the fat when the milk is only a few hours old so that the opportunity for germ growth is relatively slight. 1 20. Buttermilk. Buttermilk contains a large amount of casein and sugar, and is, therefore, of considerable value for feeding purposes. It is usually very rich in bacteria, sometimes containing even more than ripened cream. Pammel 1 found 1,700,000 germs per cc. in but- termilk, while the organisms in butter were less than half a million. This high content is due to the age of the milk and temperature at which the cream is previously ripened. 1 Pammel, Iowa Expt. Stat., Bull. 21. Cream and Factory By -Products. 123 121. Whey. This by-product of cheese-making also has considerable value for feeding purposes. Like sep- arator skim-milk, it is drawn at a temperature that greatly favors bacterial development. Compared with the germ content of the raw milk, whey possesses fewer organisms than skim- milk or buttermilk kept under sim- ilar conditions, as a larger part of the bacteria present in the milk are caught in the coagulated curd. The pres- ence of these in whey, if left to cool naturally, soon sours the product, as the milk-sugar is further converted into various acids. Both whey and skim-milk for feeding purposes should be carefully handled in order to get the most out of them. If the ferments are allowed to develop, the sugar is changed into various acids, the albumen un- dergoes putrefactive changes, and much of the feeding value is lost. 122. Treatment of whey and skim-milk tanks. The vats for factory by-products are often made of wood; consequently, they are difficult to clean thoroughly, even if that procedure is attempted. If not carefully cleansed and sterilized by steam each day, the particles of milk or whey that adhere to the walls, quickly sour, and so infect the material that is stored in the vat on the succeeding day. In this way the vat becomes a center of bacterial infection. Often, too, this already contaminated waste product is allowed to stand in cans after it is taken back to the farms until it is thoroughly soured. Such fer- mented food has only a minimum value, as much of its nutritive worth is gone. Vats for these by-products should be constructed from galvanized iron and arranged to empty by gravity. The vats and supply pipes should be carefully cleaned each day as much as any other part of the factory. The trouble arising from sour whey and sour milk is 124 Dairy Bacteriology. made still worse by the practice of returning these highly contaminated fluids to the farm in the same set of cans that are used for the transportation of milk. Filthy whey- vats and cans are responsible for much of the tainted milk that is seen in factories. It is necessary that the cheese-maker should have his waste-vat free from sour and half-fermented whey before he can charge all of the blame upon the patron who brings bad milk. FIG. 30. A Swiss cheese factory, showing careless way in which the whey is handled. Each patron's share is placed in a barrel, from which it is removed by him. No attempt is made to cleanse these receptacles. Fig. 30 illustrates the manner in which the whey is distributed and cared for in many factories that make Swiss cheese particularly. The hot whey is run out through the trough from the factory into a large trough that is placed over a row of barrels, as seen in the fore- ground. Each patron thus has allotted to him his proper share, which he removes day by day. No attempt is made to clean out these receptacles, and the inevitable Cream and Factory By -Products. 125 result is that they become a foul, stinking mass in hot weather. This material is usually carried home in the same set of cans in which the milk is brought and as the cans are often insufficiently cleaned, the new milk is in- fected with undesirable organisms. That such a custom should have grown up in the Swiss cheese industry, where on account of the sweet curd nature of the cheese, all possible precautions should be taken to insure the best quality of milk, illustrates the necessity of bacteriologi- cal principles being thoroughly instilled into the dairy industry. 123. Preserving- factory by-products. Factory by- products on account of their chemical constitution and their bacterial flora soon decompose, and a considerable proportion of the nutrients are lost. This has led to the introduction in some factories of measures that tend to prolong the keeping quality of the whey and skim-milk, although the value of these products do not warrant much expense. The conditions under which skim-milk and whey are produced facilitate the rapid development of bacteria. The continued fermentation of these materials may be inhibited in part as follows : 1. Quick cooling to a temperature unfavorable for germ development. 2. Pasteurizing or scalding. Where fed immediately, it is hardly worth while to treat these by-products, as they will remain sweet for several hours. In some factories the skim-milk is heated by introducing live steam. Where milk is pasteurized for butter- making, the keeping quality of the skim-milk is increased 24-48 hours thereby. 124. Skim-milk a distributor of disease. In some countries, notably in Denmark and Germany, tubercu- losis is so common among cattle that skim-milk from 126 Dairy Bacteriology. factories becomes a serious menace when fed to young stock. To prevent distribution in this manner, compul- sory legislation requires that the skim-milk shall be pasteurized at a temperature of 176 F., in order to destroy the bacilli. Storch has recently devised a test that can be easily applied to milk to determine whether such treatment has been carried out. It rests upon the following principle: Milk contains a, certain substance, (presumably an enzyme, 1 ) that decomposes hydrogen peroxid. If milk is heated to 80 C. (176 F.,) or above, this reaction ceases. When potassium iodid and starch are added to unheated milk and the same treated with di- lute hydrogen peroxid, the fluid assumes a blue color due to the action of released iodin upon the starch. A striking illustration of the danger that may come from a skim-milk-supply that is infected with disease bac- teria is seen in the Welply epidemic of typhoid fever in England in 1893, where twenty- three cases of this disease developed in the families of patrons of a single factory, the milk- supply of which became infected and was spread by means of the skim -milk. Foot and mouth disease of cattle is often disseminated in the same way. During the last decade, this disease has been very severe in certain parts of Europe. The virus of the disease can, however, be destroyed by the use of heat. The regulation of the Prussian govern- ment require milk of all diseased animals to be heated to 212 F. or to 194 F. for fifteen minutes. 1 Storch believes it to be the unorganized ferment discovered in milk by Babcock and Russell, 14th Wis. Expt. Stat., p. 77, 1897. CHAPTER IX. BACTERIA IN BUTTER-MAKING. 125. Sweet and ripened cream butter. The pecu- liar qualities of ordinary butter are so dependent upon the relation of bacteria to cream, that in order to under- stand the subject aright, it is necessary to consider, first, their effect in cream. If butter is made from fresh sweet cream, it has a delicate, although unpronounced flavor that differs .considerably from the usual product. With the great bulk of the commercial product, the cream is allowed to stand for a certain length of time, during which it undergoes a series of fermentations technically known as "ripening." American consumers have become accustomed to the peculiar properties of acid cream butter, and as yet, there is but little demand for the sweet cream product. The keeping quality of the latter is relatively poor compared with that made from ripened cream, so that it is only adapted for immediate consumption. The germ content and the quality of ripened cream butter varies materially, depending upon the way the cream is handled. In dairy butter-making, the cream is usually gathered by the gravity process, and is ripened in various ways. In creamery butter two methods of securing cream are in vogue. The more primitive way is where the cream is separated by gravity and is taken to the factory when it is nearly ready for churning. The more modern method is to bring the whole milk to the creamery where the cream is removed by centrifugal separation. In the one [127] 128 Dairy Bacteriology. case, the fermentative changes are well under way when it reaclies the central station, and as the cream is secured from a number of different persons in a partially ripened condition, it is far from being uniform in character. Where centrifugal cream separators are used, the cream is secured sweet, thereby permitting a supervision of the ripening process at the central station where the butter is made. This control of the ripening process exerts a profound influence on the character of the fermentation that takes place, and naturally modifies the product. The uni- formity of methods employed necessarily has a tendency to render the product more uniform. A. BENEFICENT ACTION OF BACTERIA IN BUTTER. 126. Ripening- of cream. The ripening changes that occur in cream are exceedingly complex, and our knowl- edge concerning the same is as yet far from satisfactory. The present belief is that these fermentations are largely the result of bacterial action, as germ growth is very abundant. Conn 1 found in fresh cream 4,060,000 bac- teria, while the ripened cream contained 346,000,000. In the ripening of cream at least three different fac- tors are to be taken into consideration, the development of acid, flavor and aroma. Much confusion in the past has arisen from a failure to discriminate between these factors. While all three of these qualities are produced simultaneously in ordinary ripening, it does not neces- sarily follow that any single organism is able to develop all three qualities. If this ripening is allowed to go too far, undesirable rather than beneficial decomposition products are produced. These greatly impair the value of butter, so that in a sense the commercial value of this 1 Conn, Storr's Agric. Expt. Stat., 7: 72, 1894. Bacteria in Butter- Making . 129 product is dependent upon the character and the extent of the ripening changes. A careful study of large num- bers of dairy bacteria as has been done by Conn, shows that these problems can be separated, if pure cultures are used. 127. Development of acid. In the ripening of cream acid is almost invariably developed. This development is essential, as it renders churning easier, and increases the yield of butter. The acid formed is largely lactic, and is produced by a decomposition of the milk-sugar. So characteristic of cream-ripening is this production of acid that the process is frequently spoken of as souring, although it must be kept in mind that the process in- volves much more than mere acid production. Normal butter made from ripened cream always has a character- istic flavor, which quality is distinct from the acid for- mation. This difference is seen in the results obtained by Tiemann 1 when cream is ripened by the addition of hydrochloric acid. When so treated, cream can be easily churned, but the product is lacking in the aromatic qual- ities found in normally ripened cream. 128. Flavor. The flavor of butter is that quality that is judged by the sense of taste. Good, bad, or indiffer- ent flavors may be produced in butter as a result of bac- terial action. In fact, the production of flavor in ripened cream butter is in large part due to the fermentative pro- ducts of microbes, although it is to some extent modified by the inherent qualities of the milk that are induced by the character of the feed which is consumed by the animal. 129. Aroma. The aroma of butter is often confounded with that of flavor, but this quality is dependent upon 1 Tiemann, Milch Ztg., 23: 701. 9-B. 130 Dairy Bacteriology. the presence of volatile decomposition products that appeal to the sense of smell rather than taste. As a rule butter is judged by this characteristic, a good flavor ac- companying a desirable aroma, but while these two qualities are frequently present in the same butter made from naturally ripened cream, it does not by any means follow that one is directly dependent upon the other. 130. Origin of flavor and aroma. The source from which these delicate and evanescent qualities are derived is not yet definitely known. Two opposing views have been advanced. Storch 1 holds that the flavors are pro- duced from the decomposition of milk-sugar and the ab- sorption of the volatile flavors by the butter fat. Conn 2 believes that the nitrogenous elements in the cream func- tion as food materials from which are formed various decomposition products, among which is the desired aro- matic substance. The change is unquestionably a com- plex one, and cannot be explained as a single fermentation. There is no longer much doubt but that both acid-form- ing and casein- digesting species are concerned in the pro- duction of proper flavors as well as aromas. The re- searches of Conn 3 , who has studied this question most exhaustively, indicate that both of these types of decom- position participate in the production of flavor and aroma. He has shown that both flavor and aroma production are independent of acid, that many good flavor- producing forms belong to that class which renders milk alkaline, or does not change the reaction at all. Some of these species liquefied gelatin and would therefore belong to the casein- dissolving class. Those species that produced bad flavors are also included in both fermentative types. 1 Storch, Nogle. Unders. over Floed. Syrning, 1890. 2 Conn, 6th Storrs Expt. Stat., p. 66, 1893. 3 Conn, 9th Storrs Expt. Stat., p. 17, 1896. Bacteria in Butter-Making. 131 Conn has found a number of the organisms that are favorable flavor- producers ; in fact they were much more numerous than desirable aroma-yielding species. None of the favorable aroma forms were lactic acid species l . 131. Effect of bacteria on ripening". The majority of bacteria in ripening cream do not seem to exert any particular influence in butter. A considerable number are positively beneficial, inasmuch as they produce a good flavor and aroma, so far as these qualities are pronounced. A more limited number are concerned in the production of undesirable ripening changes 2 . A knowledge of the conditions that control the development of these respect- ive types of ripening is of greatest practical value to the butter-maker. If it were possible to have these condi- tions under exact control, then the butter industry would be reduced to the terms of an exact science. 132. Methods of cream-ripening". 1. Natural ri- pening. The simplest and oldest method is where the cream is allowed to ripen without any special control, the only artificial aid being a regulation in a crude way of the temperature. The whole process is a let-alone one. The ripened product varies much in degree of ripeness ; also in the kind of fermentation, depending upon the various species of bacteria that have happened to gain access to the milk. The results obtained by these cruder methods are usually fair, but simply because the majority of the or- ganisms normally present in ordinarily clean milk are such as are unable to produce marked flavors of an un- desirable character. 2. Natural starters. In the above method the rate of ripening is often irregular, and to overcome this, starters 1 Weigmann has reached these same results (Milch Ztg., p. 793, 1891). 2 Eckles. Cent. f. Bakt., II. Abt., 4: 730. 1898. 132 Dairy Bacteriology. have been introduced as a means of hastening and rendering the ripening process more nniform. In the use of these, bacterial growths are added to assist in the ripening of cream, although generally no knowledge of the bac- terial processes involved has until recently been taken into consideration. These methods are a great improve- ment over the previous let-alone policy of cream-ripening as one is often able to exclude in this way, undesirable fermentative changes. For starters of this sort, several different materials are used, as sour-milk, buttermilk, or whey, all of which are liquids rich in bacteria. 3. Artificial starters (bacterial cultures}. A much more scientific method of ripening cream has been intro- duced within the last few years and is now being exten- sively used in Scandinavian, Danish, and German dairies. This method dates from the labors of Storch, 1 who in 1890 proposed and introduced the use of pure cultures that have been selected on account of their ability to produce a desirable ripening change in cream. 4 His in- vestigations opened up a new field of research which has since been diligently prosecuted in both Europe and America. 133. Principles of pure culture cream-ripening'. This method rests on the principle that a selected culture that has been chosen on account of its favorable ripen- ing qualities will be able to produce a similar fermenta- tion in cream, if the proper conditions are present. To get the best results, it is evident that the selected culture will have favorable conditions for its development, if the pre-existing bacteria in the cream are first destroyed. In this way competition is reduced to a minimum. This preparation of the cream is accomplished by pasteurizing 1 Storch, Milch Zeit., 1890, p. 304. Bacteria in Butter-Making. 133 the same. This destroys for the most part the vegeta- tive bacteria, leaving the latent spore forms in the cream. The addition then of a properly propagated starter gives the selected organism snch an impetus that it should be able to overcome any other organism in the cream. Where pure cultures are employed extensively, as in Denmark and Germany, this method of cream-ripening is followed. The attempt has been made to use these culture start- ers in raw sweet cream, but it can scarcely be expected that the most beneficial results will be attained in this way. This method has been justified on the basis of the following experiments. Where cream is pasteurized and no starter is added, the spore-bearing forms frequently produce undesirable flavors. These can almost always be controlled if a culture starter is added, the obnoxious form being repressed by the presence of the added starter. This condition is interpreted as indicating that the addi- tion of a starter to cream which already contains devel- oping bacteria will prevent those originally present in the cream from growing. 1 This repressive action of one species on another is a well-known bacteriological fact, but it must be remembered that such an explanation is only applicable in those cases where the culture organ- ism exercises a direct prejudicial influence on the exist- ing flora of the cream. If the culture organism is added to raw milk or cream which already contains a flora that is eminently adapted for development in this medium, it is quite doubtful whether the culture organism would gain the supremacy in the ripening cream. The above method of adding a culture to raw cream renders cream-ripening details less burdensome, but at the same time Danish experience, 1 Conn, 9th Storrs Expt. Stat., p. 25, 1896. 134 Dairy Bacteriology. which is entitled to most credence on this question 1 , is opposed to this method. 1 34. Pasteurizing* milk or cream for butter. To pasteurize for butter-making, it is not necessary to carry out the process in the same stringent manner as when it is done to preserve milk or to free it from pos- sible disease-breeding bacteria. A temperature of 140 F. for ten minutes is fatal to most of the lactic acid group, but in pasteurizing for butter-making, it is cus- tomary to heat the milk to a higher temperature (155- 175 F.). This is done for two purposes. First, the milk is generally heated in a continuous-flow machine; consequently, the maximum temperature is not main- tained for more than part of the time required to go through the machine. Second, the wide- spread preva- lence of tuberculosis, and foot and mouth disease in some of the dairy regions of Europe makes it necessary to heat the skim-milk high enough to destroy the seeds of these diseases. In pasteurizing for butter-making, it is also possible to drive off many taints that have been absorbed directly from the cow or through exposure to foul odors 2 . Some states of the Atlantic sea-board are so infested with wild garlic that the milk-supply is rendered practically worth- less for dairy purposes. If this is treated by pasteur- ization, it is often possible to eliminate in part such taints, so that a fairly good product of butter can be made. In pasteurizing for butter-making, two methods are in vogue in Denmark where the greatest activity exists in regard to this matter. There, either the whole milk is 1 99$ of the Danish creameries have pasteurizers, and over 90$ pas- teurize the cream. 2 McKay, la. Expt. Stat., Bull. 32, p. 477. Bacteria in Butter- Making. 135 handled or the cream is pasteurized after separation. The necessity of heating the skim-milk to destroy the seeds of tubercle in the milk- supply makes it advisable to heat the whole milk, thus subserving two purposes by the same operation. 135. Pasteurizing" apparatus for butter- making*. Most of the European pasteurizing machinery has been devised from the butter-maker's standpoint. They are, therefore, machines that have a large capacity, as this factor is generally regarded as of chief importance in pasteurizing for butter- making. To fulfill this condition, most of them are arranged with a continuous inflow and outflow. Such an arrangement militates considerably against the efficiency of any machine as a bacteria- de- stroying agent. In a test made by the writer 1 of Reid's pasteurizer (fig. 31), which is a modification of a Danish machine, it was found that the bacterial content of the pasteurized milk varied greatly, rang- ing from 50-99%, although on the whole the germicidal effect was very marked. 136. Attributes desir- able in pure culture starters. In adding a pure Reid's pasteurizer for butter- culture starter to cream, it is desirable, if possible, to use a ferment that possesses the following character- istics : FIG. 31. making. 1 Wis. Expt. Stat., Bull. 69, p. 11. 136 Dairy Bacteriology. 1. Vigorous growth in milk at ordinary ripening tem- peratures. 2 . Ability to form acid so as to facilitate churning and increase the yield of butter. 3. Able to produce a clean flavor and desirable aroma. 4. Impart a good keeping quality to butter. 5. Not easily modified in its flavor- producing qualities by artificial cultivation. These different conditions are difficult to attain, for the reason that some of them seem to be in part incompat- able. Weigmann 1 found that a good aroma was gener- ally an evanescent property, and therefore, opposed to good keeping quality. Conn has shown that the functions of acid-formation, flavor and aroma production are not necessarily related, and therefore, the chances of finding a single organism that possesses all the desirable at- tributes are not very good. It may well be asked if the conditions are so peculiar, how is it that a good product is ever made where no special attention is given this matter of culture starters? The reason for this is that the ripening of cream is not a single type of fermentation, but a complex process in which various organisms may participate. In all probability no one germ possesses all of the de- sirable qualities, but the resultant of the action of several forms may produce a good product 2 . This idea has led to the attempt of mixing selected organisms that have been chosen on account of certain favorable characteris- tics which they might possess. The difficulty of main- taining such a composite culture in its correct proportions when it is being propagated in the creamery is seemingly well nigh insuperable, as one organism is very apt to develop more or less rapidly than the other. 1 Weigmann, Landw. Woch. f. Schl. Hoi., No. 2, 1890. 2 Weigmann, Cent. f. Bakt., II. Abt., 3: 497, 1897. Bacteria in Butter- Making. 137 137. Reputed advantages of culture starters. 1. Flavor and aroma. These are factors of greatest im- portance, as the market value is determined largely by this quality. Ferments of this sort rarely produce any higher or more pronounced flavor or aroma than the best of the natural product, but they possess the reputation of producing a clean, mild flavor that is very desirable when the market becomes accustomed to it. 2. Uniformity of product. Culture starters produce a more uniform product, because the type of fermentation does not vary from day to day. Even the best of butter- makers sometimes fail to make a standard product, owing to their inability to fully control the character of the ripening process. 3. Keeping quality of product. Butter made from pas- teurized cream to which a good, pure starter has been added will keep much better than the ordinary product, because the diversity of the flora is less and the milk is not likely to contain those organisms that produce an " off " condition. 4. Elimination of butter defects. One of the greatest advantages is its application to factories that are unable to make good butter on account of some hidden trouble due to bacteria. 138. Pure cultures vs. home-made starters. The question as to the relative merits of pure or domestic fer- ments as starters should be answered in the light of vary- ing conditions. In the present development of the pure culture system, no culture on the market possesses such superior advantages over a first-class domestic starter especially as to flavor and aroma, that it can be unquali- fiedly recommended as better. The main advantage of the culture system lies in the uniformity of the product. The elimination of the risk that always attends in a greater 138 Dairy Bacteriology. or smaller degree the use of an ordinary starter, is of material worth. If great care, however, is taken in the selection of the domestic starter, and it is propagated un- der as equally favorable conditions for the retention of its original purity, as can easily be done with the com- mercial- starter, then there is no reason why practically as good results cannot be obtained in the use of one as the other. 139. Imperfections in system. The commercial value of butter is so dependent upon the character of the flavor that the effect of a starter on this factor will always be of supreme importance . At the present time no pure culture starter is known that is capable of imparting a much higher flavor than is obtained in the regular way l . Consequently, the maker is unable to secure a higher price for his butter than in the better creameries where the older method is used. Another decided disadvantage that occurs where the cream is pasteurized in conjunction with the use of a pure culture, is the effect on the grain or body of the product. As judged by our present American standards, the grain of the butter is materially lowered where the cream has been heated. 2 This undoubtedly would not be so considered in the foreign, especially the English or Danish markets, but so long as our market standards exact the qualities now demanded on grain, this will to some extent handicap pasteurized butter. 140. Propagation of starter for cream-ripening-. The preparation and propagation of a starter for cream- ripening is a process involving considerable bacteriologi- cal knowledge, whether the starter is of domestic origin or prepared from a pure- culture ferment. In any event, 1 This statement is confined to American conditions. 2 Farrington and Russell, Wis. Expt. Stat., Bull. 69, p. 28. Bacteria in Butter- Making. 139 it is necessary that the starter should be handled in a way so as to prevent the introduction of foreign bacteria as far as this is possible. The following points should be kept in mind in manipulating the starter: FIG. 32. Apparatus for sterilizing and propagating the starter 1. If a pure-culture ferment is used, see that it is fresh and that the seal has not been disturbed. 2. If a home-made starter is employed, use the great- 140 Dairy Bacteriology. est possible care in selecting the milk that is to be used as a basis for the starter. 3. For the propagation and perpetuation of the starter from day to day, it is necessary that the same should be grown in milk that is as germ-free as it is possible to secure it. For this purpose sterilize some fresh skim- milk in a covered can that has previously been well steamed. This can be done easily by setting cans contain- ing skim-milk in a vat filled with water (fig. 32 A) and heating the same to approximately the boiling point. The temperature should be maintained for an hour or more. This destroys all but the most resistant spore- bearing organisms 1 . 4. After the heated milk is cooled down to about 70 F., it can be inoculated with desired culture. Some- times it is desirable to ' ' build up ' ' the starter by prop- agating it first in a smaller volume of milk, and then after this has developed, adding it to a larger amount. This method is of particular value where a large amount of starter is needed for the cream -ripening. 5. After the milk has been inoculated, it should be kept at a temperature that is suitable for the rapid devel- opment of the contained bacteria, 60-70 F., which tem- perature should be kept as constant as possible. 6. This can best be done by filling water- vat (A) with water and heating the same to desired temperature, cov- ering the vat with a wooden cover or heavy cloth during the night to maintain proper temperature. 7. The starter should not be thoroughly curdled and solid when it is needed for use, but should be well soured 1 A number of tests made by the writer at the Wis. Dairy School Creamery, showed that the bacteria in skim-milk were practically destroyed, only 7-30 bacteria per cc. remaining after this sterilizing process. Bacteria in Butter-Making. 141 and partially curdled. This point is of importance for the following reasons : a. It is difficult to thoroughly break up curd particles if the starter is completely curdled. If these curd masses are added to ripening cream, white specks may appear in the butter. b. The vigor of the starter is undoubtedly stronger when the milk is on the point of curdling than it is after the curd has been formed some time. The continued for- mation of lactic acid kills many of the bacteria and thus weakens the fermentative action. 8. The starter should be propagated from day to day by adding a small quantity to a new lot of milk. For this purpose two propagating cans should be provided (fig. 32 B) so that one starter may be in use while the other is being prepared. 9. After the starter has been used for a few days, the same should be emptied, and the can cleaned and steril- ized by steam before being used again. 141. How long 1 should a starter be propagated? No hard and fast rule can be given for this, for it de- pends largely upon how carefully the starter is handled during its propagation. If the starter is grown in steril- ized milk kept in steamed vessels and is handled with sterile dippers, it is possible to maintain it in a state of relative purity for a considerable period of time ; if, how- ever, no especial care is given it, it will soon become in- fected from the air and the retention of its purity will depend more upon the ability of the contained organism to choke out foreign growths than upon any other factor. While it is possible by bacteriological methods to deter- mine with accuracy the actual condition of a starter as to its germ content, still such methods are inapplicable in creamery practice. Here the maker must rely largely 142 Dairy Bacteriology. upon the general appearance of the starter as determined by taste and smell. Where it is possible to propagate the starter as has been described, it is hardly worth run- ning any risk in using an old starter when a fresh one -can be so easily kept in condition. 142. Bacteria in butter. As ripened cream is neces- sarily rich in bacteria, it follows that butter will also contain germ life in varying amounts, but as butter-fat is not well adapted for bacterial food, the number of germs in butter is usually less than in ripened cream. Sweet cream butter is naturally poorer in germ life than that made from ripened cream. Grotenfelt 1 reports in sweet cream butter, the so-called " Paris butter " only 120-300 bacteria per cc., while in butter from sour cream 2,000-55,000 germs per cc. were found. Pammel 2 found from 125,000-730,000 per gram, while Lafar 3 found in butter sold in Munich from 10-20,000,000 organisms per gram. The germ content of butter on the outside of a pack- age is much greater than it is in the middle of a mass ; this doubtless being due to the freer access of air favor- ing the growth of aerobic forms. 143. Changes in germ content. The bacteria that are incorporated with the butter as it first ' ' comes ' 7 un- dergo a slight increase for the first few days. The dura- tion of this period of increase is dependent largely upon the condition of the butter. If the buttermilk is well worked out of the butter, the increase is slight and lasts for a few days only, while the presence of so nutritious a medium as buttermilk affords conditions much more favorable for the continued growth of the organisms. 1 Grotenfelt-Woll, Prin. Mod. Dairy Practice, p. 244. 2 Pammel, Bull. 21, Iowa Expt. Stat., p. 801. 3 Lafar, Arch. f. Hyg., 13: 1, 1891. Bacteria in Butter- Making. 143 While there may be many varieties in butter when it is fresh, they are very soon reduced in kind as well as num- ber. The lactic acid group of organisms disappear quite rapidly ; the spore-bearing species remaining for a some- what longer time. Butter examined after it is several months old is often found to be almost free from germs. This fact is important, for in considering the after -changes in ' ' storage ' ' butter, the relation of these changes to the germ life would naturally be considered. In the manufacture of butter there is much that is de- pendent upon the mechanical processes of churning, washing, salting, and working the product. These pro- cesses do not involve any bacteriological principles other than those that are incident to cleanliness. The cream, if ripened properly, will contain such enormous numbers of favorable forms that the access of the few organisms that are derived from the churn, the air, or the. water in washing will have little effect, unless the conditions are abnormal. 144. Rancid change in butter. Fresh butter has a peculiar aroma that is very desirable and one that enhances the market price, if it can be retained; but this delicate flavor is more or less evanescent, soon disappearing, even in the best makes. While a good butter loses with age some of the peculiar aroma that it possesses when first made, yet a gilt-edged product should retain its good keeping qualities for some length of time. All butters, however, sooner or later undergo a change that render them worthless for table use. This change is usually a rancidity that is observed in all stale products of this class. The cause of this rancid condition in butter has been attributed to the action of living organisms, partic- ularly those that form butyric acid, to the influence of light, of air, etc. 144 Dairy Bacteriology. In rancid butter, butyric and allied acids are always found, and it was supposed for a long time that the change was a butyric fermentation inaugurated by some of the butyric organisms that are found so commonly in milk and cream. Ritsert 1 found that sterile butter became rancid in three days if exposed to the action of the light, while unsteril- ized, normal butter exposed under similar conditions, did not change in five months, if the air was excluded from it. Duclaux 2 has proven that the rancid change is largely a chemical action that takes place in butter-fat where it is exposed to light and oxygen; that it is not necessarily inaugurated by the vital functions of any special kind of bacteria. While the change goes on in most cases in a purely chemical way, there are, however, certain organ- isms that are able to hasten this process if they are present in the butter 3 . For this reason a soft butter containing considerable quantities of buttermilk, and therefore rich in nitrogenous material, undergoes a rapid change and quickly becomes rancid. 145. Defects due to manufacturing' methods. There are other defects in butter that are also attributa- ble to other than bacterial causes. These are for the most part due to errors in manufacture. Thus, mottled or wavy butter is generally caused by the uneven distribu- tion of the salt. White specks in butter are often pro- duced where the cream is allowed to ripen for too long a time, or where curdled milk is used as a starter. In such cases, the curd particles remain undissolved, and are held in the butter, where they appear as white specks. 1 Ritsert, Inaug. Diss. Berne, 1890. * Duclaux, Le Lait, p. 34. 3 Von Klecki, Cent. f. Bakt., 15: 354. DIVERSITY Bacteria in Butter- Making. 145 B. BACTERIAL DEFECTS IN BUTTER. 146. Lack of flavor. Often this may be due to im- proper handling of the cream in not allowing it to ripen far enough, but some times it is impossible to produce a high flavor. The lack of flavor in this case, is due to the ab- sence of the proper flavor- producing organisms. This condition can usually be overcome by the addition of a proper starter. The relation between flavor and desir- able bacteria is very intimate, and troubles of this kind usually arise, because the proper forms commonly found in the cream have been supplanted by other species that do not possess the ability of forming these aromatic sub- stances so necessary in sour cream butter. 147. Putrid butter. This specific butter trouble has been observed in Denmark, where it has been thoroughly studied by Jensen 1 . Butter affected by it rapidly ac- quires a peculiar putrid odor that ruins it for table use. Sometimes, this flavor may be developed in the cream previous to churning. Jensen found the trouble to be due to several different putrefactive bacteria. One form which he called Bacillus fcetidus lactis, a close ally of the common feces bacillus, produced this rotten odor and taste in milk in a very short time. Fortunately, this organism was easily killed by a comparatively low heat, so that pasteurization of the cream and use of a culture starter quickly eliminated the trouble, where it was tried. 1 48 . Turnip-flavored butter. Butter sometimes ac- quires a peculiar flavor recalling the odor of turnips, rutabagas, and other root crops. Often this trouble is due to feeding, there being in several of these crops, aromatic substances that pass directly into the milk, but in some instances the trouble arises from bacteria that 1 Jensen, Cent. f. Bakt., 11: 409, 1891. 10 B. 146 Dairy Bacteriology. are able to produce decomposition products 1 , the odor and taste of which strongly recalls these vegetables. 149. "Cowy" odor in butter. Frequently there is to be noted in milk a peculiar odor that resembles that of the cow stable and sometimes the odor of the animal itself. Usually this defect in milk has been ascribed to the absorption of impure gases by the milk as it cools, although the gases and odors naturally present in fresh milk have this peculiar property that is demonstrable by certain methods of aeration. Occasionally, it is trans- mitted to butter, and recently, Pammel 2 has isolated from butter, a bacillus that produced in milk the same peculiar odor so commonly present in stables. 150. Lardy and tallowy butter. The presence of this unpleasant taste in butter may be due to a variety of causes. In some instances, improper food seems to be the source of the trouble; then again, butter exposed to direct sunlight bleaches in color and develops a lardy flavor. 3 In addition to these, cases have been found in which the defect has been traced to the action of bacteria. S torch 4 has described a lactic acid form in a sample of tallowy butter that was able to produce this disagreeable odor. 151. Oily butter. Jensen has isolated one of the causes of the dreaded oily butter that is reported quite frequently in Denmark. The specific organism that he found belongs to the sour-milk bacteria. In twenty-four hours it curdles milk, the curd being solid like that of ordinary sour milk. There is produced, however, in ad- dition to this, an unpleasant odor and taste resembling 1 Jensen, Molk. Ztg., 6: Nos. 5 and 6, 1892. 8 Pammel, Bull. 21, Iowa Expt. Stat., p. 803. 3 Fischer, Hyg. Rund, 5: 573. 4 Storch, 18th Kept. Danish Agric. Expt. Stat., 1890. Bacteria in Sutler-Making. 147 that of machine oil, a peculiarity that is transmitted directly to butter made from affected cream. 152. Bitter butter. Now and then butter develops a bitter taste that may be due to a variety of different bac- terial forms. In most cases, the bitter flavor in the butter is derived primarily from the bacteria present in the cream or milk. Several of the fermentations of this character in milk are also to be found in butter. In ad- dition to these defects produced by a biological cause, bitter flavors in butter are sometimes produced by the milk being impregnated with volatile, bitter substances derived from weeds. 153. Moldy butter. This serious defect in butter is caused by the development of the common mold fungus (Penicillmm glaucum) on the inside of butter packages. Where tubs are made from green, sappy wood, the mold- spores that are practically everywhere, find favorable conditions for growth, especially, where moisture exists. The result is that as the wood molds, the outer layers of the butter are stained and at the same time acquire a moldy taste. This defect can be easily obviated by steaming the tubs thoroughly before packing. Butter tubs should always be kept in a cool, dry place under conditions unfavorable for the growth of the mold-spores. CHAPTER X. BACTERIA IN THE CHEESE INDUSTRY. 154. Relation of bacteria to cheese. The processes of cheese-making are much more affected by the operation of bacterial causes than almost any other dairy industry. Cheese contains so large a proportion of nitrogenous matter that, so far as food is concerned, bacteria find bet- ter conditions for development than in butter. The ripening or curing of cheese is a fermentative process in which bacteria are intimately concerned. Our knowledge of the actual changes induced by these organisms is, as yet, quite meager, but enough has already been deter- mined to indicate that the whole industry is based largely on phenomena of ferment action, and that the application of bacteriological principles and ideas to this phase of dairying is sure to yield more than ordinary results, in explaining in a rational way, the reasons underlying many of the operations of this industry. 155. Principles of cheese-making*. The general principle of cheese-making is to precipitate the casein, and then press this into a more or less solid mass," allow- ing it to stand for varying periods of time, and under various conditions, so that a wide range is presented, in which various fermentative actions may operate to pro- duce the widely different kinds of cheese. From the same kind of milk, a great variety of different cheeses can be prepared, depending upon the treatment of the milk during the manufacture, and the way in which the cheese is handled after it is made. [148] Bacteria in the Cheese Industry. 149 In precipitating the casein, the fat is caught in the curd particles, and is thus incorporated in the cheese. This curdling of the casein is brought about in two ways. 1. Addition of rennet, a chemical ferment extracted from calves' stomachs 1 , which has the specific property of rendering the casein of milk insoluble, e. g., cheddar and Swiss varieties of cheese. 2. Development of acid until the casein is precipitated, e.