UNIVERSITY OF CALIFORNIA COLLEGE OF AGRICULTURE AGRICULTURAL EXPERIMENT STATION BERKELEY, CALIFORNIA BOVINE TRICHOMONIASIS HUGH S. CAMERON BULLETIN 624 JULY, 1938 UNIVERSITY OF CALIFORNIA BERKELEY, CALIFORNIA CONTENTS PAGE Distribution and economic importance 3 The causal organism 4 Cultivation 5 Pathogenicity 5 The course of the disease 7 Symptoms and diagnosis 8 Treatment 16 Control 16 Literature Cited 19 BOVINE TRICHOMONIASIS 1 2 HUGH S. CAMERON 3 Among cattle diseases, those causing such genital disturbances as abor- tion and sterility probably result in the heaviest losses. In recent years, a protozoan, Trichomonas foetus (Riedmiiller), has been incriminated as one cause of these two conditions. Although the incidence of this organ- ism has not been determined, it probably ranks next to Brucella abortus as a cause of genital disease in cattle. Bovine trichomoniasis may be de- fined as a venereal disease caused by Trichomonas foetus and character- ized by abortion, sterility, and pyometra (a condition characterized by an accumulation of pus in the uterus). Genital diseases of unknown cause in herds free from Bang's disease have caused considerable loss in several sections of California. Because of numerous requests to the University of California, an investigation, which later became cooperative between the College of Agriculture and the United States Department of Agriculture Bureau of Animal Indus- try, was initiated in the San Joaquin Valley. Trichomoniasis was diag- nosed in five herds that had reported breeding difficulties. Control measures for the elimination of the trichomonads increased the breeding efficiency. Because of the apparently growing importance of this disease in dairy herds, especially herds free from Bang's disease, the information about it has been compiled for the use of veterinarians and cattle owners. Although some of the information has already been published, a part is original and is largely based on recent investigations in the San Joaquin Valley. DISTRIBUTION AND ECONOMIC IMPORTANCE Bovine trichomoniasis has been reported from many European countries. In the United States, it was first recorded in 1932 by Emmerson in Penn- sylvania (i). 4 Since then, it has appeared in numerous states, ranging from the eastern seaboard to the Pacific. In California it has been diag- nosed by Traum 5 in one herd in Siskiyou County and by Schroeder 6 in 1 Eeceived for publication March 29, 1938. 2 The investigational work reported in this paper became cooperative with the United States Department of Agriculture Bureau of Animal Industry, February 1, 1937. 3 Assistant Professor of Veterinary Science and Assistant Veterinarian in the Experiment Station. 4 Italic numbers in parentheses refer to "Literature Cited" at the end of this bulle- tin. 5 Traum, J. Personal communication. 6 Schroeder, C. R. Personal communication. [3] 4 University of California — Experiment Station two herds in San Diego County. The writer has observed it in one herd in Alameda County and in five herds in Tulare County. In these in- stances the organism was observed in genital exudate. Certain other herds in which the organism was not found presented a typical picture of trichomoniasis. Apparently the condition is widespread in California. Being insidious, the infection is usually well established in a herd before it is observed. Losses, consequently, are heavy. Abelein, according to Stableforth and Scrogie (2), pronounces it the most common cause of sterility in his district of southwest Germany. Out of 168 cases of pyo- metra, trichomonads were observed in 144. In another herd, where no cause of breeding difficulties could be found except trichomonads, he re- ported that the percentage of cows with calf dropped from 96 to 26. The remaining 74 per cent, representing 49 cases, had breeding trouble. There were 6 cases of observed early abortion and 12 cases of pyometra. In a herd of 75 mature animals in New York (3) , there were 9 observed abor- tions and 7 unobserved (a previous examination had revealed preg- nancy) in 15 months. In an outbreak observed by Walsh, McNutt, and Murray (4), 133 services resulted in only 25 calves, an average of 5.3 services per calf. This compared with a previous record of 195 services for 128 calves, an average of 1.5 per calf. After the control of the out- break, 97 services produced 50 calves, an average of 1.9 per calf. Gould (5) estimated the loss in a valuable herd of about 30 animals at over $10,000. THE CAUSAL ORGANISM Trichomonas foetus is a flagellated protozoan. Normally the body is some- what spindle-shaped, 10 to 25 microns 7 in length, and about a third as wide. In other words, it is about a third larger than the head of a bovine sperm cell. It has three anterior flagella (fig. 1, a) arising from a common blepharoplast (fig. 1, 6), and one posterior flagellum (fig. 1, g), a con- tinuation of the edge of an undulating membrane (fig. 1, e) . In fresh preparations, the organism is actively motile, later becoming sluggish and more rounded. It is rapidly killed by drying and quickly loses its morphology so that it cannot be distinguished from cellular debris. Clearly stained specimens, therefore, are difficult to prepare un- less one takes particular pains in the process of fixing. Figure 1 shows a diagrammatic sketch of Trichomonas foetus; figure 2, a photomicrograph of a stained specimen obtained from a case of pyometra. The genus Trichomonas was first described in 1836, when one of its species (Tr. vaginalis) was observed in vaginitis of women. Since then many species have been described, the majority being saprophytes. The 7 1 micron = 39.37 millionths of an inch. Bul. 624] Bovine Trichomoniasis genus is widespread, being found in many different forms of life ; it is described in Wenyon's Protozoology (6). Tr. foetus, the species that pro- duces bovine trichomoniasis, has been studied in detail by Wenrich and Emmerson (7), who concluded that it was distinct from Tr. vaginalis. Cultivation. — Although several workers report success, Trichomonas foetus is not easy to cultivate. Apparently Witte (8) was first to obtain the organism in pure culture. He concluded that he obtained the best Fig. 1. — Diagram of Trichomonas foetus (Tviedmiiller) : a, one of the three anterior flagella; o, blepharoplast ; c, nucleus; d, parabasal body; e, undulating membrane; /, chromatic basal rod; g, posterior rlagellum; h, cytostorne; Tc, chromatic granules ; I, karyosome ; m, axostyle. growth in broth containing 5 to 10 per cent blood. Gehring and Murray (9) maintained cultures in Locke's egg medium with blood and 1-10,000 crystal violet. Glasser and Coria (10) purified trichomonads from bac- teria by inoculating with the mixed culture one arm of a V tube contain- ing 15 cc of sterile semisolid agar and one drop of defibrinated blood. After 16 to 20 hours at room temperature, a pure culture of trichomonads could be obtained from the other arm. Then it was cultivated on Locke's egg blood medium incubated at 37° C. Transfers were made every two weeks. Pathogenicity. — Although most species of the genus are considered harmless, Trichomonas foetus is generally recognized as disease-produc- ing. Drescher and Hopfengartner (11) in 1933 considered the disease 6 University of California — Experiment Station one of minor importance. They found trichomonads in only 9 per cent of 655 samples received on account of genital disease. Over half of the remaining cases, however, were not diagnosed. Abelein — according to Stableforth and Scrogie (2) — and others have not been able to observe the organism in a healthy genital tract. Witte (12) produced sterility Fig. 2. — Trichomonas foetus (Riedmiiller) accompanied by a piece of cellular debris. Photomicrograph of uterine exudate from a case of pyometra. (x 3,200.) Note undulat- ing membrane on left side. in three heifers with intra vaginal injections of bacteria-free cultures of trichomonads. The writer (3), in a previous investigation, obtained uterine exudate aseptically from a case of pyometra. Although it was heavily infected with trichomonads, cultural methods revealed no bacteria. When similar methods were used in a case in Alameda County, again no bacteria were observed. Actively motile trichomonads obtained directly from a case of abortion produced temporary sterility in one heifer and abortion follow- ing temporary sterility in another; Trichomonas foetus was obtained from the uterine exudate at the time of abortion. Success in controlling the disease by eliminating the bulls that are spreading trichomonads further supports the view that Trichomonas foetus is a pathogen. Bul. 624] Bovine Trichomoniasis THE COURSE OF THE DISEASE Apparently the principal means of transmission is by the bull during coitus. This conclusion (3) is based on findings in a large breeding herd free from Brucella infection. The herd was composed of six breeding units, which were stabled and pastured together so that, from a disease standpoint, there was no segregation. One or more bulls were assigned to each unit, and under no circumstances were these bulls changed from one unit to another ; there was no crossbreeding between the units. Trichom- onad infection was found only in the one unit that developed breeding difficulties. When Walsh, McNutt, and Murray (4) attempted to transmit the in- fection to normal cows by pasturing them with a heifer that was discharg- ing heavily infected pus, they found no evidence of spread. Andrews and Miller (13), on the other hand, observed the organism in the vaginas of virgin heifers that were mingling with infected cows. As with other vene- real diseases, infection may perhaps occur in this way ; but such an occur- rence is probably rare. Infrequent elimination and the brief duration of viability in the vagina support the assumption that transmission by other than venereal means is rare. Futamura (14) observed, at autopsy, Trichomonas foetus in the am- pulla of the vas deferens of an infected bull. It can readily be found at the fornix of the sheath on autopsy and occasionally by swabbing the sheath of the living animal. Apparently the organism does not interfere with conception, but the formation of a cervical seal creates conditions favorable for the propaga- tion of trichomonads. Eventually the latter become so numerous that the fetus dies and may be expelled immediately. Frequently it is so small as to be unobserved. On the other hand, it may be retained in the uterus, undergo maceration, and result in pyometra. During pyometra, estrum is usually inhibited ; and in its absence the cow is assumed to be pregnant. With the termination of pyometra, estrum occurs ; and if bred while still infected, the cow may transmit the organism to a clean bull. The source of the infection in a herd can usually be traced to the intro- duction into the herd of a bull used elsewhere or of a cow supposedly pregnant but really open, or to the acceptance of neighboring cows for service. The prognosis of bovine trichomoniasis differs in the sexes : In the male it is very unfavorable, and recovery is rare. In the female the opposite obtains. The cow will discard the organism rapidly after the opening of the cervix. If not exposed to reinfection, she will completely recover. 8 University of California — Experiment Station SYMPTOMS AND DIAGNOSIS Of the several symptoms associated with bovine trichomoniasis, none alone is diagnostic. Abortion — the most pronounced symptom — differs from that seen in Bang's disease : the fetus is usually in a macerated con- dition, having been dead in the uterus for some time, whereas in Bang's disease it usually appears normal and apparently has died only recently. The fetus may be so small that the abortion is unnoticed, and nothing is suspected until estrum reappears. In this case the condition is diagnosed as sterility rather than abortion. The predominating feature of the ap- parent sterility is delayed estrum. A period ranging usually from 30 to 90 days elapses between service and the return of estrum. Occasionally the period is longer — sometimes 8 or 9 months. In two cases during the Tulare investigations, the period extended beyond the date on which parturition was due. Rectal examination for determining pregnancy usually reveals pyome- tra in the herd. Mucopus at the vulva, especially when the animal is lying down, is frequently observed. In character the discharge may vary from clear mucus with a few white spots to a thick brownish pus. No apparent lesions are produced in the male genitalia. As mentioned in the discussion of transmission, the organism is readily found on au- topsy in the fornix of the sheath. Quantity, motility, and morphology of the spermatozoa are not affected. Any of the foregoing symptoms, especially if they occur sporadically, may be caused by agents other than Trichomonas foetus. If they appear as an epidemic, a presumptive diagnosis of trichomoniasis is justified. Since, however, the disease is insidious in its spread, accurate breeding records covering at least two years are essential before the herd history can be relied on for such a diagnosis. The following data accumulated from one of the herds in the San Joaquin Valley investigations presents a typical picture of bovine trichomoniasis. Detailed breeding records were compiled from several herds reporting considerable sterility. These herds were free from Brucella infection and tuberculosis. Free access to breeding records and a willingness to follow recommendations were requested. Figure 3 shows the breeding chart used in collecting those data. When it was brought up to date from existing records, the owner was instructed to record all breedings, and calvings. If an animal was in estrum but not bred, this was recorded in the breed- ing column and distinguished by a circle around the date. From such a record, the breeding efficiency of each bull was deter- mined. Tables 1 and 2 give detailed breeding records of two bulls in Bul. 624] Bovine Trichomoniasis < 5? m 5 O z 5 ■ r ■ J u < ° • 2-^ u 2 3f 1 a t- > < j o < H u u m 8 »- < a o z 5 ■ ■ S*3 rt ** « t- °S3 2°i ■ rJ Co 2 3 o ** ei_, M £ a ^ 2 £ Pi ^ a a^ O c3 J-l >H „ o 03 «H O c8 £0 03 «!_■ 03 E > ^ .a a ^ © © a i— i a to TO f_ © 3 ►"3 «H © © • i-H > - © OQ P o OQ © ,P P © © as p •>-H ^ -U r-w • |_l '-"' 83 § t:,p