key: cord-020235-stcrozdw
authors: nan
title: Abstracts of Papers Presented at the 38th Meeting of the Deutsche Gesellschaft für Hygiene und Mikrobiologie, Virology Section, Göttingen, 5.–8.10.1981
date: 2012-03-15
journal: Zentralbl Bakteriol Mikrobiol Hyg A
DOI: 10.1016/s0174-3031(82)80128-5
sha: 
doc_id: 20235
cord_uid: stcrozdw

nan

SCHAEFER, A., ZIBIRRE, R., KABUS, P., KOHNE, JUTTA, and KOCH. G. Na +, K + -ATPase activity was studied in a plasma membrane rich fraction isolated from control and poliovirus infected He La cells and compared to membrane potential and amino acid uptake in parallel cultures of intact cells. Na + , K + -AT Pase activity in membranes from infected HeLa cells increased relative to control with a maximum at 90 minutes post infection (+ 30 %) and decreased again (180 rnin.: -30 %). Similar but slighter changes were observed in the membrane potential dependent tetraphenylphosphonium (T PP+ ) uptake, indicating a correlation between membrane porential in intact cells and our measurment of plasma membrane Na + , K +-AT Pase. At approximately 1 h post infection we observed a decrease in the uptake of amino acid (methionine, leucine) in infected cells relative to controls. These results suggest that the decline in amino acid upt ake is not mediated by virus-induced changes in the Na + , K + -ATPase activity or membrane potential. MPI f. Biochemic, Abt. Virologic, Sequence Homology in Different Strains of FMDV and other Picoma Viruses MARQUARDT, O. Restriction enzyme generated subgenomic fragments of cloned cDNA prepared from RNA of the strain 01K of FMDV were compared quantitatively for sequence complementarity with radioactive RNA from strains CObb and A2S or with RNA from mengo or polio virus in hybridization experiments by use of the Southern technique. Nucleic acid sequences neighbouring or including the 3' end of viral genomes are demonstrated to be 80 Ofo homologous in FMDV. In contrast, sequences coding for the capsid proteins VPl and VP3 were remarkably heterologous (less than 20010) in FMDV. Sequences coding for non-structural proteins showed 35-50010 homology. Thus no highly conserved coding sequence was detectable in FMDV by this technique. No hybridization could .be detected between 01K specific DNA and polio RNA, while weak hybridization was observed with mengo RNA at areas including the 3' end and with a part of the gene for precursor protein P52.

Abr, Molekularbiologie, Physiol-Chern. Inst., Grindelallee 117, D-2000 Hamburg 13 Modification of Poliovirus Capsid Proteins SCHARLI, CLAUDIA and KOCH, G.

Poliovirus type 1, strain Mohoney contains a protein kinase activity. A highly purified (two cycles of CsCI gradient centrifugation) poliovirus preparation is able to transfer the y-phosphoryl-group of [32P]_ATP to acid precipitable material. When the reaction product is anal yzed by SDS PAGE, all structural proteins of polio are phoshorylated. Most of the label is incorporated in the minor capsid protein VPO and to a lesser extend in VP1, VP2 and VP3. Hepatitis A virus (HAV) was isolated directly from human stool in diploid human fibroplasrs, Viral antigen was expressed only after 210 days of incubation of the infected cultures. In contrast, HAV first adapted to growth in human hepatocellular carcinoma cells caused antigen expression in fibroplast cultures already 90 days after infection. During further passage the time of first appearance of antigen in infected fibroblasts decreased to about 14 days in both passage series (i. e. cultures inoculated with virus recovered directly from human stool or after adaption to human hepatoma cells). Antigen was predominantly cell-associated and was shown by immunofluorescence to be located exclusively in the cytoplasm. -Biophysical properties of HAV particles extracted from infected cells were comparable to those described for HAV extracted from human stool. Those findings are of importance for preparation of large amounts of HAV for vaccine. production.

Max von Pettenkofer-Inst. u. Inst. f. Biochemie d. Univ. D-8000 Miinchen 2 Cloning of Hepatitis A Virus Genomes HELM, K. VON DER, WINNACKER, E. L., and DEINHARDT, F.

Subgenomic fragments of the genomic RNA of hepatitis A virus (HAV) were cloned via eDNA into Pst 1 site of pBR 322. By restriction enzyme pattern analysis and hybridization of the cloned HAV DNA with selected fragments of the HA V RNA it was determined that the major part of the cloned HAV DNA fragments are located at the 3' end of the genome but a few clones are distributed over the entire genome.

we have now identified subtype specific sites in HBV-restriction map s. The maps were aligned to the Eco RI site. Only subtype ay revealed one subtype specific site of each XbaI (2000 BP), H inelI (2200 BP) and Bam HI (2900 BP). They are situ ated outside the s-gene. Employing rhis system the subty pes of three HBV-ONA sequences that ha ve been published so far , could be determined . Thereupon the first 54 triplets of a HBV (ad)-DNA sequence of the s-gene could be com pared with pub lished data . Nucleotide exchanges in triplets 12 and 13 did not cau se an amino-acid (aa)-exchange, bu t cha nges in the triplets 45 and 46 do cause such exchange. Thus, in subtype ay, aa 45 is thr, and aa 46 is thr, wh ile in subtype ad, aa 45 is ser and aa 46 is pro. These exchanges may produce different conf ormation of th e peptides.

Dept. M ed. Mic robio l., Univ. 0 -3400 Gottingen Characterization of the Hepatitis B Virus (HBV) Associated Proteinkinase GERLICH, W. H.

Purified HBV-core preparations were shown to contain a pr ot eink inase which phosphory lates the major core protein (Albin and Robinson, 1980) . In this study it was found th at th is enzyme copurifies with the light (d = 1.31) but not with the heavy subfraction (d = 1.35) of the core-particles. The enzyme has a high affinity for ATP and it transfers approx. one ph osphat e per particle within minutes. The 32P-phosphate introd uced in vitro cannot be remove d from core-particles by digestion with alkaline ph osph at ase. After lysis of the cores with SOS the 32p can be cleaved from rhe protein.

T his suggests th at th e enzyme and its ph osphate acceptor site are located with in the pa rticle. After acid hydrolysis the incor porated 3!P co-migrates with ph osph oserin but not with phosphor hreonin or phos pho tryosin.

M ax von Petrenk ofer-Inst., Univ. 0 -8000 Miin chen 2 A Phosphokinase Activity Associated With the Hepatitis B Virus (HBV) Core HELM, K. VON DER, ROGGENDORF, M., and SIEGERT, W.

Prepar ation s of core antigen positive (HBcAg) fractions obta ined from HBV positive hu man liver tissue are associated with a ph osphokinase wh ich copurifies with the HBcAg in CsCl density-and sucrose velocity grad ient centrifugations. The acceptor protein for this kina se is the 19000 d HBcAg protein; casein as an exogenous acceptor is also phospho rylate d to a minor extend. The ph osphorylated amino acid is serine or threonine and not tyrosi ne, as it is frequently the case with tumor virus specific prot ein kinases.

Dept. Med . Microbiol., Univ. 0 -3400 Gott ingen Effect of Glycosidases on the Proteins of Hepatitis B Surface Antigen (HBsAg) STIBBE, W. and GERLICH, W. H .

The protein composition of purified HBsAg was stud ied by SOS·gc1e1ectrophoresis and staining with silver. In additio n to P25 and GP28 of HBsAg two furt her proteins, GP33 and GP36, were found consistently in preparations from 9 different blood donors. The glycoprotein nature of GP 33 and GP 36 was shown by the increase of electrophoretic mobility after treatment with endoglycosidase H or neuraminidase. Immune precipitation with anti-HBs in RIPA-buffer confirmed the specifity of the glycoproteins, The change in apparent mol. weight after treatment with neuraminidase was larger for GP36 (900) than for GP33 (600) or GP28 (300). The data suggest that GP 33 and GP36 are multiply glycosylated proteins with N-glycosidic linked carbohydrate side chains of the mixed type. Inst, f. Virologie, Univ., HB s antigen screening was done in the blood samples of 1,790 pregnant women. 33 blood samples were found to be HB s antigen positive. 32 women were asymptomic carriers, 1 woman suffered from chronic hepatitis. 31 children have been born until now. The examination of cord blood showed HB s antigen in 6 samples; the venous blood samples taken thereupon demonstrated HB s antigen only in two samples. The HB s antigen negative children were treated with HBIG. The follow-up examination of the treated children showed no signs of infection up to now. -One child born to an HB e antigen positive mother was already infected at the time of birth. Another child born to an anti HB e positive mother became HBs antigen positive at the age of 4 months. Unfortunately, this child had not been treated with HBIG at the time of birth.

WILLERS, H., PONNATH, H., SIPOS, S., and MOLLER, R. Sera of 5,847 pregnant women living in the Hannover area were investigated for the presence of HBsAg. 56 healthy HBsAg carriers were found: 17 (0.340/0) among 4,962 women of German origin and 39 (4.280/0) among 912 foreign-borne women from Southern European and non-European countries. 10 out of the 56 HBsAg carriers (114,962 German women and 9/912 foreign-borne women) were recognized to be HBeAg-positive. -The risk of perinatal transmission of hepatitis B virus in infants of HBeAg-positive mothers is suggested to be 90 % and of HBeAg-negative mothers 12 0/0. -Thus in the Hannover area 6 out of 10,000 infants born to German mothers and 129 out of 10,000 infants born ro foreign mothers become HBsAg carriers due to perinatal hepatitis B virus-infection.

Max von Pettenkofer Insr., Univ. D-8000 Munich 2

A radioimmunoassay for anti-HBc IgM using higher concentration for the dilution of serum (5 mg/ml) and HBcAg (10 mg/ml) was compared with an ELISA test ]. din. Microb iol. 13 (1981) , 618. Onl y 10 of 157 (6 0 /0) HBsAg positive blood don ors were positive (40 in ELISA) and no correl ation with the total anti -HBc titer could be found . After fraction ation of a serum with discrepant results a positi ve result was found in the ELISA only in the IgG fractions. In a collaborative stu dy with A/torfer et al. (presented also at Lisbo n, EASL 1981 (1981) , who show ed a 10 6 fold reduction in hepatitis B titer by means of !i-PLlUV treatme nt. The effectiveness of the cold sterilization procedure as regards reducing virus infectivity is considera bly greater than that of pasteurization (10 h 60 DC) for which Shikata et al. (1977) sho wed a 10 4 fold reduction of hep atitis B virus infectivity. It has also been found that factor IX conce ntrate and stab ilized serum (Biseko®) der ived from pooled. cold-sterilization hu man plasma transmitted neither hepatitis B nor hepatit is no n-A, non-B in chimpa nzees.

Max von Pettenk ofer-Insr., 0 -8000 Miinchen 2 Comparison of Different Evaluation Systems for Determination of Viral Antibodie s of the IgG and IgM Class in the Enzymeimmunoassay R O G G ENDORF, M. , ZACHOVAL, R ., ZOULEK, G., a nd DEINHAROT, F. T he enzymeimmunoassays used to demonstrate antibodies of the IgG and IgM class against viral antigens reveal a high sensitivity. At onset of acute viral infections IgM antibodies can be demonstrated up to serum dilutions of 10-7 • Due to th is high sensitivity, the determin ation of antibody tite rs is not accurate and reproducible, because titer end points are determ ined as that dilution of sera giving an 0.0. sample/O .O. negative control equal or greater than 2.1. -For the dete rmination of antibody con centr ations an evaluation method is proposed which correlates, the measured 0 .0. of sera at one dilut ion step to the 0 .0. of a reference serum which is defined arbitrarily to contain 100 anti body units. Using an ELISA for detection of antibodies to adenovirus, a significant increase in antibod y units of acute phase sera over that of convalescent pha se sera is observed. Low day to da y varia tions are seen in tests car ried out on different days. In an ELISA which is designed to detect anti bodies to tick-born e encephalitis virus of th e IgM class, the day to day variatio n using antibody units was significantly lower th an using PIN ratios.

Inst. f. Virolog ie, jusrus-Liebi g-Un iv., 0 -6300 GieBen;

Max-Planck-Insr, f. M olekulare Genet ik, D -lOoo Berlin Structure and Function of the Core Protein of Alphaviruses BOEGE, ULRIKE and WITT MAN N-LIEBOLD, BRIGITTE The com plete primary struc tures of the core proteins of Sindbis and Semliki Forest virus have been established by pr ote inchemical methods. Both proteins contain clusters of basic amino acids and proline in th e N-terminal part, suggesting th at its function is to bind to the nucle ic acid. The C-termi nal pa rts have ext ensive iden tical sequence regions, pr ob abl y p roviding the recognition sites for pr otein-protein interactions . Both proteins contain within these highl y conserved portion s the sequence Gly-Asp-Ser-Gly wh ich is typical for serine proteases and most likely is related to the cata lytic function of the core prot ein as a protease wh en cleaving its own peptide chai n from the nascent protein precursor. Experimental studies using peptides which contain certai n sequence regions will help to elucidate the relationships of struc ture and functions. An ac ute and a persistent infectio n with Rab ies virus (H EP Flury) was established in th e CN S-derive d cell 1ine 108-CC-I5 (NG 108-15). These cells possess specific membrane recept ors to many hormon es and neurotran smitters including opiate receptors. In both cases we found increases in th e dissociation constant (Kn) for the agonist 3H-etorp hine as estimated by Scatcha rd plot ana lysis. H owe ver, in both cases th ere was no change in th e nu mb er of op iat e receptors per cell compa red to uninfected cells. These studies complete our previou s published observations of the impai rme nt o f receptor func tions in rabies virus infe cted cells (1) .

(1) Kosch el, K. an d M. Halb ach: J. Gen. Virol. 42 (1979) , 627-632.

Insr, f. Virologic u. Immunobi olo gie d. Univ., 0-8700 Wiirzburg Effect of Measles (SSPE) Antiserum on Viral Surface Proteins and Hormone Receptor Activity in C6/SSPE Persistently Infected Cells BARRETT, P. N. and KOSCHEL , K.

It has been reported (1) th at measles antis erum ca n modulate the expression of certain viru s prot eins in acu tely measles infected cells. We have exam ined th e effect of measles (SSPE) antiserum on the exp ressio n of vira l surface proteins in C6ISSPE infected cells. We have shown an over 50 % redu ction in the am ount of viral antigen present on the memb ranes of these cells following incuba tion with SSPE antiserum. This was demon-strated both by immunofluorescence and rad ioimmunoassay. However this loss of antigen had no effect on memb rane receptor linked cAMP synthesis. This is discussed with respect to the effect of virus antigen insertion in cell membranes on specialised membrane bound functions. (1) We have analyzed the effect of phosphorylation and dephosphorylation on the structure and DNA binding of D2-T antigen. On non-denaturing pore-gradient gels the purified protein migrated with an apparent size of 135,000 daltons, In vitro phosphorylation by the protein kinase associated with the purified protein resulted in a shift of most of the protein to a size of 740,000 daltons, and it was this form that contained most of the phosphate incorporated. This aggregation was completely reversible by treatment of phosphorylated D2-T antigen with alkaline phosphatase. Partial tryptic digestion indicated that phosphorylation of sites in the Nvrerminal part of the protein is responsible for the observed aggregation. -As show n by protein blotting onto nitrocellulose filters predominantly the 740,000 dalton form bound to SV40 DNA. However, onl y a fraction of the in vitro phosphorylated protein did bind to DNA, suggesting that aggregation alone is not sufficient for DNA bind ing. Subclasses of SV40 large T antigen were separated by zone velocity sedimentation. Three major forms, which sedirnented at about 5-6S, 14-16S and 23-255, have been shown to differ biologically and biochemically (1) . Each subclass was tested for specific binding to restriction fragments of SV40 DNA using an immunoprecipitation assay. -All three forms of T antigen bound specifically to a restriction fragment containing the SV40 origin of replication. However, the 5-6S form bound more origin fragment per unit T antigen than the other forms . The 5-6S form bound equally well to origin DNA in the presence and absence of excess cellul ar DNA , whereas the binding of the 14-16S form was reduced in the presence of cellular DNA . All forms of T antigen from SV40-transformed SV80 cells bound much less origin DNA than that from infected cells.

(1) Fanning, Nowak , Burger: J. Virol37 (1981) Wart scrapings from several small skin regions of an Epidermodysplasia verruciformis patient were tested for papilomavirus-specific sequences by means of a 32P-labelled HPV8 DNA. Uncleaved wart DNA contained uniforme full length HPV genomes. Cleavage with Bam HI revealed six different patterns and a surprising heterogeneity even within small biopsies. At least two virus types showed only limited cross-hybridization with HPV 8a. One of these closely resembles HPV 5b (Bam HI fragments 2,9 and 2,1; Eco RI fragments 3,6 and 1,1). Hind II fragments A and C of his virus perfectly hybridize with HPV 8a; B, D and 6 anneal only partially and F, G show no detectable hybridization. The DNAs of four subtypes were partially characterized and mapped. Only DNA of the HPV 5b-like virus was detected in a probe from the center of a carcinoma at the patient's forehead. This DNA persists extrachromosomally with more than 100 genome equivalents per cell.

Inst, f. Virologie, Zentrum Hygiene, Univ., Hermann-Herder-Str. 11, D-7800 Freiburg Gene Expression of Papillomaviruses in Hamster Tumors FREESE, U. K.,SCHULTE, P., and PFISTER, H. BPV 1 includes fibrosarcomas in hamsters. The tumors contain large amounts of complete virus genomes which persist extrachromosomally but there is no evidence for capsid protein synthesis or virus particle production. We used this system to study early viral gene expression. RNA from the tumors contained a single virus-specific RNA with about 1300 nucleotides which was shown to be polyadenylated by affinity chromatography on poly-Uesepharose. The transcribed DNA region of BPV 1 was mapped within the two Hpa II fragments, which are next to the Bam HI cleavage site within the 1.4 X 10 6 d Bam HIIEco RI fragment. Cross-hybridization studies under low stringency revealed some sequence relationship of HPV 1 and HPV 4 DNA to the transcribed region of BPV 1.

Inst. of Genetics, Univ., D-5000 Cologne

The Adenovirus Type 12-Mouse Cell System: Permissivity and Analysis of Viral DNA in Tumor Cells STARZINSKI-POWITZ, A., SCHULZ, M., and DOERFLER, W.

Interactions between viruses and eucaryotic cells can be studied in a genetically well defined system like the mouse system. We have investigated whether Ad 12 DNA is able to replicate in primary mouse kidney cells or in the mouse cell line L929. It was shown by Southern blot experiments that Ad12 DNA was not able to replicate in L929 cells, w hereas in prima ry mouse kidne y cells of (BALB/c X CS7/B16) Fl origin, viral DNA repl icated. Moreover , we subcutaneously injected Ad12 into mice of various genetic origins. In ab out 100 mice injected, one rumor emerged in a BALBlc mouse almost 8 months after injection. Restriction pattern anal ysis of the rumor or DNA indicated that abou t 2-3 copies of Ad12 DN A were integrated and covalently bound to cellular DNA. With th e techniques available no deletion or rearrangement of viral DNA could be found. T he sites of jun ction betw een viral and cellular DNA will now be cloned and sequenced.

Inst, of Genetics, Univ., 0-5000 Co logne Virus-Cell DNA Recombinants in Human Cells Lytically Infected by Ad2 NEUMANN, R., WEYER, U., and DOERFLER, W.

There is ample evidence for the notion that virus-cell DNA recombinants are formed in human cells productively infected with adenovirus type 2 (Ad2). These high molecular weight forms were detected at 1-2 h postinfection and were generated at high frequency, A limited set of rather specific recombinants was produced (Neumann and Doerfler, J. Virol. (1981) 887 suggesting th at recomb ination exhibited a cert ain specificity. We now succeeded in molecularly cloning DNA frag ments excised from the high mole cular weight DN A of Ad2-infected hum an cells by th e restri ction endonuclease EcoRI in AgtWES . AB or in ACharon 4B. Some of these clon ed frag ments had sequence ho mology to both viral and cellular D NA. Th is result provided proof fo r the occurrence of virus-cell DNA recombi na nts .

Inst, of Genetics, Univ., 0-5000 Cologne Unmethylated DNA Sequences in the Promoter Regions of Integrated Adenovirus Genes Correlate with Gene Expression KRUCZEK, 1. and DOERFLER, W.

An inverse correlation was established between the levels of DN A met hylation at 5'-CCG G-3' sites in specific segments of integrated adenovirus DNA and the extent to wh ich the se segments were expressed (Sutt er and Doerfler, PNAS 77 (1980) 253. Similar correlatio ns were reported in other viral and non-viral systems. More recently, the results of in vitro experiments prov ided direct evidence for the notion th at DNA methylation at specific sites led to gene inactivation (Vardimon et al., PNAS, in press ). -In the present stud y a detai led methylation map at 5'-CC GG-3' (H paIIlM spI) sites in the expressed early and the silent late genes of Ad12 DN A was determined in three Adl2-tr ansformed hamster cell lines. The early region s of inte grated Ad12 DNA were unmethylated; in particular their promot er regions were unrnethylated ar the Hpall sites in all three lines investigated. T he late regio ns including the promot er sites were completely meth ylated.

GAHLMANN, R., DEURING, R., STABEL, S., WINTERHOFF, U. VARDIMON,1., and DOERFLER, W.

The sites of junction between viral and cellular DNA were sequenced to investigate two problems: 1. Are the sites of insertion of viral DNA specific? 2. What type of recombinatorial events occur in viral DNA integration? We have studied junction sites from the Ad2-transformed hamster line HES, and from the Ad12-induced hamster tumor lines CLACI and CLAC3. The virus-cell DNA junctions were cloned in the DNA of bacteriophage 2gt· J.B. Appropriate restriction fragments were sequenced by the Maxam-Gilbert technique. There was no direct sequence identity apparent at the viral and cellular junction sequences. Deletions at the termini of the viral genome were seen involving 5 (HE5), 45 (CLAC3) or 174 (CLACl) nucleotides. Peculiar patch type homologies between the cellular and viral sequence adjacent to the site of junction and also in remote areas were observed. These patches may have an important function in integrational recombination.

BUTTNER, W., VERES-MaLNAR,S., and BLOCK, J.

As a first step to understand the relationship between structure and function of the adenovirus type 2 DNA binding protein (Ad2 DBP) we have begun to determine the primary structure of the DBP gene. The isolation and characterization of Tupaia adenovirus (TAV) has been reported. In order to construct a genetic map of the TAV genome the use of temperature-sensitive mutants (ts) of TAV was necessary. -A variety of ts mutants of T AV, which were generated by treatment of T AV virions (1 X 10 10 pfu) with hydroxylamine, were isolated and preliminary characterized. Six of these mutants its: 1, 4, 11, 12, 13 , and 54) did not replicate in Tupaia embryonic kidney cells at 39.5 DC, but did replicate well at 32°C. The characterization of these mutants was carried out using complementation tests, host range studies and DNA anal ysis using different restriction enzymes. According to complementation analysis four groups were determined : group I = ts 1, group II = ts 12, group III = ts 13, and group IV = ts 4, 11, and 54. The host rage study revealed that ts 1 and 54 also had properties of host range mutants. These mutants did not replicate in Tupaia bab y fibroblasts in contrast to wild-type virus. Genome analysis of these mutants revealed that the mutated region is located between 69.9 to 74.5 map units. In addition, in ts 1 and 54 mutants a deletion from 0.77 Kb to 1.39 Kb (map unit 76.5 to 100) was detected.

Interaction of Viral Substructures with Serum and Serumcomponents resp.

During the final stage in the course of many viral infections complete virus particles as well as viral substructures enter the intercellular space, e. g. HBsAG, HBcAG in hepatitis B virus infections . The present study deals on the interaction of the potentially infectious adenovirus cores with DNA-specific antibodies and immunoglobulin solutions which had been absorbed by DNA-antigens. Cores were prepared by sarcosyl treatment of purified adenovirus typ 5. By electron microscopy immunocomplexes could be demonstrated which are composed of several individual core units. By buoyant density centrifugation in metrizamide gradients a drastic rise in the density of cores could be shown too. With immunoglobulin solutions, absorbed with native as well as with denatured DNA, so far no complexes could be assessed. A cell line designated SBL-H12578 and several cell clones were established from skin tumours of the sporadic leukosis form. The cells were proved to be free of bovine leukemia (BLV), and some other bovine viruses. By indirect immunofluorescence macroschizonts of a Theileria species were seen in the cytoplasm. The cells originated from a BLVantibody negative animal and carried a female karyotype with some morphological aberrations. Evidence for a possible T cell origin of SBL-H12578 cells was obtained. After inoculation of 8 X lOa cells into 'nude mice' transplantable SBL-H12578 tumours developed. By incorporarion of 3H-uridine and electron microscopy, the production of retrovirus particles by the cultured cells was detected . In the simultaneous detection test a highmolecular weight RNA co-migrating with FeLV 70 S RNA was demonstrated. No antigenic or genetic relationship between th e skin tumour virus isolate and BLV or other major mammalian retrov iruses has been found. -(Supported by Stiftung Volkswagenwerk ).

Abt. f. Pathologie, Gesellschafl: f. Strahlen-u. Umweldorschung, D-8042 Neuherberg Insectpathogenic Baculoviruses: Studies of the Activation of Endogenous C-type Retroviruses in Mammalian Cell Cultures SCHMIDT, J. and ERFLE, V.

Activation of endogenous C-type retroviruses by Baculoviruses was studied in "in vitro" cell culture systems of four mammalian species: mouse, rat, monkey and man. Cells were treated with Baculoviruses (from larvae and insect cell cultures), Baculovirus-DNA, C-rype retrovirus-activating chemicals and chemical insecticides alone and in combination. The activation of retroviral genomes was tested by the determination of reverse transcriptase activiry in concentrated cell culture supernatants and by the demonstration of the intracellular localisation of retrovirus structural protein p30 applying the indirect immunoperoxydase technique. -C-type retroviruses were activated in mouse cells only by the halogenated pyrimidine analogue iododeoxyuridine. In Baculovirus-treated cell cultures no C-type retrovirus activation was detectable. In simultaneous treatments of the cells with Baculoviruses and chemicals no potentiating effects could be detected. Virions of Baculoviruses in mammalian cell cultures showed unaltered morphology and upon reisolation their infectivity in homologous insect cell cultures was lowered by approximately one log. No influence upon growth or morphology of the cells could be observed. The gene products of replication-defective oncornaviruses are high molecular weight proteins which comprise a gag related and one gene product. They are probably not cleaved because the p15 protease gene is lacking. In vitro, however, the gag-specific peptide sequences are cleaved off upon addition of the purified viral piS protease; in the case of the replication-defective, transforming avian sarcoma virus PRC II the cleaved nongag part has a ryrosine-phosphorylaring kinase activity similar to that described for the RSV src-gene product pp60 src .

Processing of Pr92 gp , the Precursor to the Viral Glycoproteins of Rous Sarcoma Virus BOSCH, V., SCHWARZ, R. T., ZIEMIECKI, A., and FRIIS, R. R.

The viral glycoproteins of Rous sarcoma virus gp85 and gp35 are synthesized via a precursor polyprotein Pr92". This precursor is already glycosylated and contains the polypeptides of both gp85 and gp35. We have studied the nature and site of processing of Pr92'~to mature disulfide-linked gp85 and gp35 (VGP). We could show that in addition to proteolytic cleavage, processing involves conversion of the high-mannose oligosac-charides found in Pr92' " to the complex, sialidated oligosaccharides found in VGP. Experiments pertaining to the site of processing indicate that processing does not occur extracellularly as has been proposed by others iKlemenz and Diggelmann, J. Virol. 29 (1979) 285-292. We have determined that the small amount of mature VGP found in infected cell lysates is localized chiefly within the cell, not at the cell surface. We favour the view that further glycosylation and proteolytic cleavage occur concomitantly on smooth membranes within the cell and that subsequent to this, export in virus is rapid. 1 Paul-Ehrlich-Inst., D-6000 Frankfurt;

2 lnst. f. Virologie, Univ., D·6300 Giefen, 3 A protein of a molecular weight of about 38.000 d has been found to be phosphorylated 1 h after the onset of cell transformation by Rous sarcoma virus (RSV). It is assumed to be a physiological target of the pp60'" kinase, since, apart from being phosphorylated in the transformed cell, it can be phosphorylated in vitro by the pp60"· kinase in tyrosine (1, 2) . -Using 6 different chromatographic procedures (chromatography on DEAE-Sephacel, poly (A)-Sepharose, Blue Sepharose, and hydroxylapatite, isoelectric focusing and gel filtration ) the 38,000 d protein could not be separated from cytosolic malic dehydrogenase activity (c-M DH ). Antiserum against the 38,000 d protein inhibited c-MDH. The Transformation-specific Protein "v-myc" of the Acute Avian Leukemia Virus MC29 DONNER, P., GREISER-WILKE, IRENE, and MOELLING, KARIN Avian acute leukemia viruses transform cells through a virus-coded oncogene. In the case of the rnyelocytornatosis virus, MC29, which transforms fibroblasts as well as bone marrow cells in vitro , this oncogene is fused to a viral structural component, p19. The fusion protein, v-rnyc, was characterized by using monoclonal antibodies against p19. MC29 transformed quail fibroblasts which do not produce any virus, MC29-Q8, were analyzed b y immunofluorescence. A nuclear fluorescence was observed which was not detected in normal cells or virus-producing cells. The monoclonal antibodies were used to purify the v-myc protein by immuno-affinity chromatography. The purification achieved by this single-step purification was 3,500 fold . The eluted protein was precipitable by anti-sera and will be further characterized for its biological properties.

Med. Poliklinik, Univ., D-8000 Miinchen; Abr, f. Pathologie d. GSF, Neuherberg Biochemical Characterization of Antigens in Human Leukemic Sera that Crossreact with SiSV p30 and BAEVp30 LEIB, c., SCHETTERS, H., ERFLE, V., and HEHLMANN, R.

Antigens crossreacting with the core proteins p30 of baboon endogenous virus (BaEV) and/or of simian sarcoma associated virus (SiSV) have been isolated from human leukemic sera by immunoaffinity chromatography. The antigens have an apparent molecular weight of 70,000 in SDS-polyacrylamide gel electrophoresis. Peptide maps performed with the antigens from two different leukemic sera show that the two antigens are identical. Peptide analyses of SiSV p30 and of BaEV p30 and simultaneous mapping of p30 proteins mixed with human antigens show that 11 out of 21 major peptides of SiSV p30 and 10 out of 20 major peptides of BaEV p30 have identical mobility with peptides of the human antigens. Human serum albumin, transferrin, fibrinogen, IgG and IgM share clearly less peptides of identical mobility with the human antigens. The isoglycoproteins gp69 and gp71 were purified from F-MuLV particles (propagated in Eveline cells) by solubilization (freezing and thawing), ion exchange chromatography (phosphocellulose) and preparative SDS-PAGE. Prior to amino acid and NH 2-terminal amino acid sequence analysis, the purified glycoproteins were subjected to high performance liquid chromatography (HPLC) to remove contaminants. -The NH 2-terminal amino acid sequences (23 residues) of gp71 and gp69 were found to be different (in 10 positions) but highly related. F-MuLV gp69 shows 410f0 homology to gp71 but lacks the potential glycosylation site (sequon) at position 12 in both F-MuLV gp71 and R-MuLV gp70. et al., 1980) . In our laborato ry 99 breast can cers, 60 normal breast tissues, 44 benign breast lesions and 10 other carcin om as from south german patients were tested for crossrea ctivit y w ith MMTV-gp52. The tests were done by indirect immunoperoxidase staining on paraffin-sections using antise ra against gp52 prov ided by Dr. Spiegelman, N. Y. Specificity of positive reactions wa s controlled by preabsorption with purified MMTV-gp52. 86 breast cancers (87 0 /0) and 13 benign breast lesions (30 0 /0) gave positive reactions, whereas normal bre ast tissue and other carcinomas were negative. -The test might be an additional useful diagnostic too l for the earl y detection of micrometastases, for the assignment of metastases from unknown pr imary tumors and in doubtful cases of breast cancer. It can possibly serve as an add itional criterion for the classification of mastopath ies. Viruses wer e found in 3,691 patients. Nearly all illnesses were caused by mumps-and entero-viruses , Other viru ses were found in less than 5 % of all cases. The mumpsmeningitides were ascertained constantly over all these year s. In 1967 In , 1974 In and 1980 an increased incide nce of meningitides caused by Echo-virus type 30 wa s seen. -There was no seaso na l dep endence on the occurrence of mumps virus meningitides. Meningitides, caused by ent ero-viruses was found mor e often in the autumn. Male patients fell ill twice as ofte n as female patients. Mumps-meningitides were rarel y found in the first year of life. In contras t, enterovirus meningitides could be dete cted during the first year and caused men ingitides to age of 14. Virus meningitides among adults were rarel y found. We suspected that a number of peripheral facial pareses (P.F.P.) considered "idiopathic" might, in fact, have a viral aetiology . 71 patients of the Cologne University E.N.T. Clinic with so-called idiopathic P.F.P. were examined under the aspect of a viral aetiology. In 32 cases conditions for virological investigations were optimal (paired sera, first serum within the first week after onset of disease). In 10 of these 32 cases a viral infection could be proven (varicella zoster virus: 7, herpes simplex virus: 2, coxsackie B 4 :1). In 5 of 7 VZV cases minimal zoster lesions were observed, 4 facial and 1 thoracic. Among the 39 oth er patients (with late sera) no viral infection could be proven. In conclusion, by means of alert clinical and virological examinat ions, a considerable fract ion of idiopathic P.F.P. could be associated with a virus infection. 152 neonates were screened from the delivery to the time of discharge for rotavirus infections. Daily faecal specimens were examined by an enzyme-linked immunosorbent assay (ELISA) and a subgroup of positive specimens were also tested by a negative staining electromicro scopic method. -22 babies (14.5 0/0) were found to excrete rotavirus. 20 of them were asymptomatic infected and 2 showed mild gastrointestinal symptoms. With one exception viruses were not detected in babies less than 24 h old, but 10.9 % of them excreted virus during the second day of life and after 10 days 30 % of the neonates were positive for rotavirus. -Excretion persisted for 1 to 5 days. According to our study most babies (40 0 /0) excreted roravirus for 3 days. -A great number of the stools (29%) from the first da y which were tested by ELISA were found to have nonspecific activity in the absence of rotaviral antigen. Theref ore such stool specimens should only be examined by electron microscopy. The single radial hemolysis test (SRH-test = hemolysis-in-gel test) is a suitable technique for detection of rubella and influenza antibodies in a large number of sera. In our stud y we looked for the effect of att achment of the viral antigens to the erythrocytes by different coupling reagent s. Chrom ic chloride, cyanogen chloride, glutaraldehyd and tetraazotized -0 -dianisid ine (TOD) were used for the sensitization of the erythrocytes. -In the rubella SRH-test no improvement on regard to sensitivity of the test was seen. Moreover it is not possible to detect IgM specific antibodies after different kinds of attachment in the SRH-test. -In the influenza SRH-test with allantoic fluid of eggs infected with H 3N2 virus it was possible to increase the sensitivity with TOD, chromic chloride and potassium periodate. If tween-ether treated hemagglutinin was used only after sensitization with TOD, chromic chloride and periodate hemolytic zones were detectable .

Univ.-KinderkIinik, Mathildenstr. 1, D-7800 Freiburg RNA-Electropherotyping of Human Rotaviruses 1978 . and PASTOR, S.

Rotaviruses contain a double-stranded ribonucleic acid genome consisting of 11 segments. This can easily be extracted from crude stool suspension (method by Rodger et al., J. Clin. Microbial. 1981) . 55 out of 210 Rota antigen positive samples contained sufficient RNA to produce a satisfying pattern in the SDS-acrylamide-electrophoresis. We demonstrate six electropherotypes with differences in the relative mobility of segments 2, 3, 4, 5, 7, 8, 9, 11 . Of these diarrhea producing strains at least two different electropherotypes were found during every outbreak of Rotavirus gastroenteritis. Our findings are in good agreement with the results reported by Rodger ), Espejo et al. (1980 ), and Kalica et al. (1978 , 1976 . The so-called M-type of EMC virus is capable of inducing diabetes mellitus in mice by a selective B cell damage. The M-EMC strain used in our laboratory has partially lost this capacity. We attempted to reisolate a diabetogenic variant and to elucidate the causes of the change. Seven serial heart passages in mice did not enrich such a variant. Cloning of the virus stock yielded one clone diabetogenic in two of ten animals (5 clones tested so far), -In a different substrain of M-ECM virus (the highly diabetogenic Dvariant, obtained from Dr. Petersen) we found both diabetogenic and non-diabetogenic clones. Incidence and severity of diabetes has been shown to be age-related. T his safety study was to demonstrate whether or not granulosis virus (GV) of Laspeyresia pomonella can rep licate in vertebrates. After feeding GV to NMRI-mice, no virus induced antibodies could be detected within eighty days by radioimmunoassay (RIA) and no vertical virus transmission was observed. GV did not replicate in mice. Human sera, as well as sera from horses, cattle, sheep and swine reacted with GV in the RIA. By characterization the positive reacting human immunoglobulin c1ass(es), IgG showed the strongest positive reaction, less positive reactions were shown by IgE and IgM and no reaction by IgA and IgD. By concentrating the immunoglobulins of the negative reacting sera to 250 pg IgG/ml, all sera could be recognized as positive. Thus a non-immunological reaction has been suggested. Infection of human fibroblasts with cytomegalovirus induces typical cytopathic alterations. Cell rounding within 5-6 h postinfection (p.i.) is followed by an increase in cell size, appearance of cytoplasmic and nuclear inclusions and a morphological change to an epitheloid cell shape by 48 h p.i. In order to elucidate the participation of the cytoskeleton in this alteration of cell morphology experiments were initiated in serum-starved human fibroblasts to visualize changes in actin arrangement by indirect immunofluorescence. A early as 3 h p.i. cytoplasmic microfilamenrs had shortened and were rearranged to a more irregular pattern. At 12 h p.i. actin fibers were absent from rounded cells. The same was observed in epitheloid cells at 48 h p.i. Cultivation of infected cells with phosphonoacetic acid resulted in a partial preservation of the normal actin fiber distribution. In contrast, infected cells did not exhibit major changes in actin synthesis as estimated from the specific radioactivity of cytoplasmic actin isolated from 3H-leucine pulse labelled infected cells by DNAase I-Sepharose affinity chromatography and SDSpolyacrylamide electrophoresis with fluorography. Chicken erythrocytes were coated with glycine-extracted CMV antigen and negative control antigen by treatment with formaldehyde and CrCl s and were used for detection of CMV specific serum antibodies in PHA tests. Their sensitivity was proven to be in good agreement and their specificity superior to results seen in CFT. IgM-specific CMV antibody detection was performed either after a simple and rapid DEAE-cellulose exchange chromatography of serum samples or by IgG immunoprecipitation combined with ME-reduction controls. The main advantage of the modified CMV-PHAT was seen in the stability of sensitized erythrocytes, which can be lyophilized.

LANVERS, A., MERTENS, TH., and EGGERS, H. J.

There are reports that herpes simplex virus (HSV) could be isolated from the genitourinary tract of up to 15 Ofo of males without manifest herpes. In order to confirm these results we first wanted to establish a method for typing possible HSV isolates. We used 3 published methods: a plaque test in chick embryo cells, a neutralisation test, and the analysis of the early HSV-proteins (SDS-PAGE). All 3 methods yielded identical results. We then tried to isolate HSV from 192 materials of 181 asymptomatic males (89 urethral swabs, 55 seminal fluids, 48 fresh tissue probes). All materials were immediately inoculated into tube cultures of two cell lines shown to be highly HSV-susceptible. Additionally, the tissue probes were co-cultivated with permissive cells. We also induced cell fusion (PEG) in such cocultures. Despite all efforts we did not isolate any virus from all these materials. For an experimental approach to answer the question as to which viral genes may control pathogenesis after peripheral infection of inbred mice with HSV, we have isolated a variant strain (ANG-path) that proved highly neuropathogenic both upon i.p. -or intravaginal infection from the apathogenic strain HSV-1 ANG. Two alterations of the ANG genome have been detected by restriction enzyme analysis: the loss of the amplifiable 500 b.p. nucleotide sequence typical for ANG and a 400 b.p, deletion approximately at the position mapped for the structural viral glyco-protein D. Despite the induction of interferon and NK-cells both variants multiply to a similar extent, probably in the same target cells of the peritoneum. The spread from the peritoneum, spleen, liver, thymus and local lymph nodes to the eNS seems, however, to be controlled by the action of gene products coded for in only one of the variants. Div, of expo Virology, Insr, for Med. Microbiology, Univ., D-6500 Mainz

KNOBLICH, A., FRIEDRICH, D., GOERTZ,] ., and FALKE, D.

Herpesviruses are known to cause infections in men and animals. Strong differences in resistance to Herpes simplex virus (HSV-1) are noted among mice of various strains. Anti-HSV-activity of T-Iymphocytes, macrophages and NK cells has been demonstrated in the last years. Our interest was focused on neutralizing antibodies in primary HSV-1

Abstracts of the 38 th Meeting of the OGHM infections of mice. Antibodies become detectable by day 5 after infection and reach a plateau level at day 21, the day we chose to test the sera. Comparison of antibody titers in 18 strains of mice revealed titers always to be higher in female mice, whereas no clear influence of either H-2-haplotype or background genome could be detected. Sexual steroids produced in ovaries and testes were identified to exert influence on antibody formation by castration experiments. Treatment of mice with silica once between day 1 before and day 12 after infection resulted in a strong increase of antibody titers both in females and males at the same time abolishing the difference in antibody titers between the sexes. Silica could enhance antibody levels also after immunisation of mice with a formol-inactivated HSV-vaccine. Bestatin is a small peptide known-selectively to stimulate DNA metabolism in T-lymphocytes and to enhance HSV-antibody -titers maximally when given at day 5 after infection . After pretreatment of mice with silica the antibody augmenting effect is already achieved at day 1 after infection . In secondary HSV-l infection Bestatin acts best 1 day after infection, too.

Insr. f. Med. Virologie d. Univ., 0-6900 Heidelberg Induction and Characterization of Herpes Simplex Virus Reisolates, Isolated after Intertypic Superinfection of Latent Infected Tupaias DARAI, G. and SCHOLZ, J.

The susceptibility of juvenile and adult Tupaias to Herpes simplex virus type 1 and 2 had been reported. The intertypic recombination of herpes simplex virus using temperature-sensitive mutants of HSV-l and 2 and superinfection with wild-type HSV-l and 2 was studied. It was found that animals survived an infection of 1 X 10 7 to 1 X l()8 pfu of rs mutants of HSV-l and /or 2 which were inoculated intravenously (L0 50 for HSV-l = 10-3 • 75 and for HSV-2 = 10-2 • 25 ) . The inoculated animals were protected against a superinfection of HSV-l or 2 (5.0 X 10 6 pfu/animals). The state of viral latency in surviving animals was investigated. It was found that infectious virus was recovered from ganglia of those animals which had initially been infected with wild-type HSV-l or 2 and /or superinfected with wild-type viruses. In contrast, the infectious viruses were recovered only from spleens of those animals which had initially been infected with ts mutants of HSV-l or 2 and superinfected with wild-type HSV-2 and /or 1. It was found that recovered viruses from the spleen of the animals lost their pathogenicity and their natural tissue tropism . Significant changes in the genome of the recovered viruses from the spleen were detected. Recombination between ts mutants of HSV-l and 2 and challenged wild-type viruses was observed. Thus, the pathogenicity and genomic properties of recovered viruses were altered . This observation is the first description of generation of inrerrypic recombinants of HSV-l and 2 in in vivo.

Oiv. of expo Virology, Inst. for Med. Microbiology, Univ., 0-6500 Mainz

The Functions of the HSV-Coded dPyK-Complex Enzyme LABENZ, j., BRAUER, D., MOLLER, W. E. G., and FALKE, D.

Analysing the phosphorylating capacity of the HSV-coded dPyK of HSV-l and 2 by glycerol gradient centrifugation we detected each three peaks differing in molecular weights using ATP, AOP or AMP as phosphate donors. Also by PAGE peaks with differ-ing Rj-values could be seen. Indeed, 32P_AMP and 32P_ADP were used for phosphorylation of dThd. The AMP-dependent activiry could be purified BOO-fold. Two antisera against HSV-coded PdyK neutralized all three activities. The TK-mutant MDK (B2006) did not induce in'I'Kr cells the respective activities, only the 3 cellular TK's were detected and identified by their Rj-values. Further experiments have shown that only the HSV-l-dPyK has thymidylate kinase activity, but not that of HSV-2. The HSV-l thymidylate kinase activity could be neutralized by a TK-antiserum. -The pH-optimum, sensitivity to Mg'", Fe'", Zn'", Co" and Mri'" ions differed. The susceptibility to thiol reagents was different, the AMP-dependent activity proved to be susceptible to phenanthroline. Also the K m and Vrnax-values were detected. A diagram summarizes the biochemical function of the HSV-coded dPyK-complex. Finally there is some indication that the enzyme phosphorylates Ado by using dTMP or ATP as a phosphate donor. The importance of the enzyme complex in HSV-infected cells is discussed.

DKFZ, Inst. f. Virusforschung, 1m Neuenheimer Feld 280, Deletion of N uc1eotide-Sequences in Cloned HSV-1 Fragments during Propagation in the rec A E. coli, HB 101 GRAY, C. P., JELLINGHAUS, U., and KAERNER, H. C.

Passage of HSV at high MOl results in the generation of defective genomes which are of full length, consisting of a relatively short region of the wild type genome that is repeated. They are packaged into mature virus particles and thus leave the cell in a state that is capable of entering a new host. Such defective molecules are of interest as they represent a simpler model in which to study replication, recombination and packaging of HSV. -Such a defective molecule arising from the serial passage of HSVtype l-ANG has been mapped, and restriction fragments have been cloned into pBR 322 using the rec A-E.coH, HB 101, as host. -All the resulting clones were found to be unstable and to contain deletions, one of which has been characterised in more detail. The isolation and characterization of Tupaia Herpesviruses (THY 1 to 4) has been reported. The analysis of DNA of these viruses showed the absence of submolar DNA fragments, when the DNA was cleaved with different restriction enzymes, as well as of a stem loop structure, when analysed by electron microscopy. With respect to this genomic structure it was of interest to study he recombination events between THV 1 to 4. Recombinants were generated between these viruses using a co-infection technique in vitro on Tupaia fibroblasts. Recombinants were selected after the stocks of new progency virus were treated with specific antiserum against each parental virus. Different recombinants were isolated, plaque-purified and characterized. Results for one recombinant THV-R-26 between THV-2 and 3 were as follows: (i) The in vitro host range and the in vivo pathogenicity in juvenile Tupaia was altered compared to parental viruses. Phosphorylation of proteins is a posttranslational modification which is regulatory for the activity of several enzymes. Most animal viruses code for phosphoproreins and the degree of their phosphorylation is thought to be a controlling factor during viral macromolecular synthesis. Recent studies on protein kinases from a number of tumor viruses have raised the possibility that the phosphorylation of cell proteins is involved in the processes leading to cell transformation. -Incubation of purified tree shrew (Tupaia) herpesvirus (THV) particles with y.32p ATP resulted in the incorporation of 32p labelled phosphate into proteins. A nonionic detergent such as NP-40 was necessary for the detection of protein kinase activity. The incorporation of 32phosphate was proportional to the quantity of TH viral proteins, indicating that the viral proteins can serve as substrates for the viral enzyme. a_ 32p dATP or a_ 32P ATP did not function as phosphate donors. A divalent cation such as Mg2+ or Mn'" is essential for the enzymatic activity. A product analysis revealed that six viral polypeptides are phosphorylated. The predominant sites of phosphorylation are the (f-OH groups of serine and threonine. In 1972 a herpesvirus was isolated from young goats with a severe generalized infection in California. This isolate has been characterized in some detail and named caprine herpesvirus 1. Recently a herpesvirus was isolated in Switzerland from goats with a similar infection. We report a further characterization of both isolates and propose their classification as Bovid Herpesvirus 6 (BHV-6). -BHV-6 multiplies rapidly and shows a broad hostcell range. Crossneutralization onl y could be observed with BHV-1 (IBR/IPV-Virus), in a one way reaction. In gel immunoelectrophoresis the serologic relationship envolved the major antigenic components of BHV-1. We have evaluated two methods for the analysis of antibodies directed against EBVspecified proteins: 1. Indirect immunoprecipitation (IP) and 2. Radioimmunoassay of electroblots of SDS PAGE separated EBV-specified proteins (RIAB). Using IP we have identified 20 proteins against which antibodies are made during infection, some of these proteins have also been found using the latter technique. Many of the proteins are only reactive with EA'VCA+ sera and not ractive with EA-VCA + sera and may therefore be candidates for the EA specifying proteins. It seems likely that the failure to identify all proteins using the RIAB technique is probably due to the strong denaturating conditions used during the SDS PAGE step. Only those antibodies directed towards the primary sequence of the protein will react with the blotted proteins, whereas with IP analysis the native proteins are available for interaction with the sera. Previousl y we demonstrated that the synthesis of EBV-induced proteins in superinfected Raji cells (Raji 51) could be divided into 3 phases: Primary, secondary and tertiary (Bay-liss and Wolf, J. Gen. Virol., 1981, in press) . Recent experiments show that incubation of Raji SI in the presence of canavanine and the absence of arginine allows only a limited expression of the viral genome. If the cells are released from a canavanine block (applied from 0 to 8 h post infection) then between 2 and 4 h later several new proteins are synthesized, however, if m-RNA synthesis is inhibited (with actinomycin D) after removal of the canavanine then these new proteins fail to appear. Amongst these proteins are members of the EBV-specified early antigen complex (EA). These results indicate that an arginine-containing protein is synthesized soon after infection and that this protein is required in an active form in order to synthesize m-RNA for the secondary group of proteins. Amongst these proteins are the major components of the early antigen (EA) complex.

Max von Pettenkofer Inst., Univ., D-8000 Miinchen 2; ENT-Klinik, Univ., D-8700 Wiirzburg; Dept. Tumor Virology, Chinese Acad. Med. Sci., Beijing, China

SEIBL, R., RICHTER, W., ZENG, Y., GU,S.-Y., and WOLF, H.

Antibody titers of IgA antivirus capsid antigen (VCA) can be used for screening early tumor cases, confirming histological diagnoses and longtime surveillance of therapy. However, in high risk areas (S. China, incidence of NPC: 20/10 5 ) 2 % of the population have IgA anti VCA antibody indicating a need for methods which allow monitoring of additional parameters for deciding on the need for therapy. The same need exists in case of long term (1-2 years) survivors of NPC with constant IgA anti VCA titers. -We have developed a system to collect cell specimens by application of buffer to the tumor site and collection of cells directly on to nitrocellulose filters. These cells can be examined cytologically or for EBV DNA in nucleic acid hybridization. For the latter tests a modification of the Grunstein Hogness colony hybridization test and cloned EBV DNA have been used. In reconstruction experiments 25 virus-producing cells could be detected.

Inst. f. klin. Virologie, Univ., D-8520 Erlangen-Niimberg

KEIL, G. and FLECKENSTEIN, B.

After infection of permissive cell cultures with overlapping restriction-fragments of viral DNA derived from different strains of Herpesvirus saimiri (H. saimiri) a number of recombinants could be isolated. The analysis of the viral proteins of the wt-strains 11, OMI and S295C led to the identification of four proteins that differed within these strains with respect to molecular weight. We are able to localize these proteins in the internal region of the M-DNA by comparing the protein patterns of recombinant and parent strains. The exact localization was so far not possible because the recombination events occured mainly in regions near the ends of the L-DNA. -Furthermore recombinants were constructed to identify the genomic region responsible for oncogenicity. The wt-strains of H. saimiri are oncogenic while attenuated strain 11-att, that was obtained from strain 11 has lost this property. This may be due to a deletion of 1,1 md at the left end of the L-DNA. Recombinants between this attenuated strain and wt-strains were constructed. The test for oncogenicity in vivo is in progress. The T-Iymphoid cells can contain up to 300 genome copies per cell as episomes. We have begun to study translation and transcription in transformed cells in comparison to lytically infected OMK-cells. About 25 new proteins can be detected in lyrically infected cells having apparent molecular weights between 12-200 kd. -At early and late stages of infection the right part of the genome is preferentially transcribed and each DNA-fragment encodes a series of specific RNAs. -In contrast, we do not find any virus-specific proteins in the transformed cells after labeling with 3sS-methionine and subsequent immunoprecipitation. Preliminary results may suggest that the only virus-specific RNA found in the transformed cells are small RNAs with molecular weights of about 0.13 kb. -A more detailed analysis has to be performed in order to confirm these hybridization data.

Max von Pettenkofer Inst., D-8000 Mi.inchen 2 Characterization of Herpesvirus saimiri Glycoproteins MODROW, S. and WOLF, H.

For the identification of Herpesvirus saimiri glycoproteins we seperated H. saimiri 11 induced cell proteins on SDS-polyacrylamide gels and transfered the polypeptides by elec-trophoretic blotting (2 h, 3,7 mA/cm 2 ) to nitrocellulose paper using carbon electrodes and buffer soaked sponge to cover the gel/filter layer. Lectins, which are known to bind very specifically to certain sugar residues (Concanavalin A to D-glucose and D-mannose derivates, Soy bean agglutinin to N-acetyl-D-galactosamine and D-galactose, Dolichos biflorus agglutinin to N-acetyl-D-galactosamine, Ulex europaeus agglutinin to L-fucose) were iodinated using the NEN-Iactoperoxidase system. The glycoproteins bound to the nitrocellulose sheets were detected by incubation with the 1Z5J-labelled lectins. By this, eight viral glycoproteins could be identified, two of them were synthesized in the presence of canavanine, and characterized according to the type of glycosylation. Lymphocystis disease (LD) is a virus disease of marine fish with an almost world-wide geographical distribution. This disease is characterised by papilloma-like tumour lesions. Lymphocystis disease virus (FDLV) has been tentatively classified as belonging to the family of Iridoviridae. This report describes the anatomy of FDLV: The FDLV virions were isolated from a total of 22 fish with LD lesions, caught near the Doggerbank, including 12 flounders, 6 dabs, and 4 plaice, which were analysed individually. The purity of the virus preparations was examined by a negative-staining technique and the virion diameters were determined: flounder 227.5 ± 12.5 nm, LDV-plaice 198.8 ± 12.9 nm, and LDV-dab 200.5 ± 12 nm. DNAs of these different LDV isolates were cleaved with different restriction endonucleases and the resulting DNA fragments were separated electrophoretically on agarose slab gels. The fragment patterns demonstrate that LDV DNA of flonders and of plaice are indistinguishable, but clearly different from those of dab. The determination of the molecular weights of FDLV DNAs using contour length measurements by electron microscopy resulted in a value ranging from 60 to 150 X 10 6 daltons. In contrast the molecular weight estimations by restriction enzyme analysis resulted in lower values ranging from 60 to 90 X 10 6 daltons. This discrepancy is probably due to a restricted, permutated structure of the LDV genome similar to the genome of frog virus 3 as reported by Granoff. The structure of FLDV constituents and its interaction with host cells remains obscure. In an effort to clarify the viral components and their functions we have studied the proteins and purified FLDV and searched for virion-associated enzymes. -At least 33 distinct viral polypeptides were detected by polyacrylamide gel electrophoresis under denaturing conditions. The polypeptide patterns are remarkably specific for a given fish species (flounder, plaice, and dab) although some heterogeneity was found when proteins of individual fish of the same species were analysed. -It was found that a nucleoside triphosphate phosphohydrolase activity is closely associated with FLDV particles. This activity hydrolyses ATP with a high preference. The reaction requires a non-ionic detergent and a divalent cation, such as Mg2+. Reaction rates and substrate specifities were determined. The products of the reaction are nucleosides disphophares and inorganic phosphate.

Characterization of a Poxvirus Isolated from White Rhinoceros (Ceratotherium s. simum) PILASKI,] ., SCHALLER, K., OLBERDING, P., and FINKE, HANNELORE In September 1977 an outbreak of poxdisease occurred in White Rhinoceroses (Ceratotherium s. simum) in the Munster zoo. At the same time a similar outbreak was observed in elephants (Elephas maximus, Loxodonta africana) of the zoo in Frankfurt (airline distance about 230 km). In both cases Orthopoxvirus strains could be isolated which were similar but not identical in their biological properties (small efflorescences on the CAM with hemorrhagic center, inclusion bodies of type A V+, high pathogenicity for rabbit skin, characteristic skin lesions in adult mice) and their DNA restriction patterns (Xho I, Eco R I, Hin d III). Both virus strains were incorporated into the group of "cowpoxlike viruses" (Baxby and Ghaboosi, 1977) . The results indicate that both outbreaks had occurred independently from each other.

Characterization of a 37K Protein Specifically Associated with Released Extracellular Vaccinia Particles HILLER G. and AULBACH, H.

Infectious vaccinia virus can be isolated from infected cells after experimentally induced lysis (intracellular virus) or from the growth medium of infected cultures (extracellular virus). We have characterized a 37K protein only present on extracellular particles by its amino acid composition and its behaviour on isoelectric focusing. In addition we have used a 37K-specific antiserum to detect its distribution within infected cells. -37K protein is a late viral protein appearing 5-6 h p.i. It is predominantly found associated with the cellular Golgi complex but never with structures representing pox virus "factories". Later in infection 37K protein is incorporated into single viral particles preferentially found in the cell periphery. Upon electron microscopy approximately 3040 % of morphologically mature virions inside the cell are enwrapped by a double-membranate vesicular structure. Thus 37K viral protein is probably a component of this vesicular structure and only vesiculated virions can be released by the cell before lysis occurs.

Epidemiology of Influenza in Lower Saxony WILLERS, H. and HOPKEN, W.

The influenza surveillance in Lower Saxony is mainly based on laboratory investigations especially on the attempts to isolate influenza viruses throughout the year. -In the winter 1977-78 and in the winter 1980-81 the two influenza subtypes H3N2 and HINI circulated at the same time. The H3N2 subtype affected during the last years persons of all age-groups whereas the HINI subtype affected only persons younger than 30 years. -In the winter 1978-79 influenza B was found in an epidemiological extent. The disease caused outbreaks mostly in schools and kindergartens but also affected adults. -In 1980-81 from mid-January to mid-February scattered outbreaks were caused by the A subtype HINl, which particularly affected schoolchildren contrarily to 1978 where the HINI subtype mostly infected young adults. Influenza of the H3N2 subtype circulated from January to March. Inst. f. med. Mikrobiologie. Abr, Virologie d. TU, Biedersteiner Str. 29, Antibodies against Influenza C Virus in the Population of Germany, Kenia and Australia PFEIL-PUTZIEN, C. and MEIER-EWERT, H.

Over one hundred human sera from each of the three countries Germany, Kenia and Australia were tested for antibodies against influenza C virus, using the conventional hemagglutination inhibition test (HI). The rate of positive sera, showing a HI titer 8 amounted to 59 Ofo for Germany, 93 Ofo for Kenia and 96 Ofo for Australia. In the age group up to 5 years, 34 Ofo of German and 94 Ofo of kenian sera had already antibody titers against influenza C virus. The australian sera were tested for the age group of 16-25 years and showed 95 Ofo seropositivity. The results show that influenza C virus is circulating to a higher extend in the populations of countries with subtropical climate, as compared to the more temperate middle european zones.

Inst, f. Virologie, justus-Liebig-Univ., D-6300 Giessen

The Proteolytic Activation of Influenza Hemagglutinin, Structure of the Cleavage Site and the Mechanism of Cleavage GARTEN, W. and BOSCH, F. X.

The hemagglutinin precursor HA is posttranslationally cleaved by proteases to the complex H I , 2' In vitro cleavage of HA by trypsin and trysin-like proteases yields infectious virus. Cleavage by thermo lysin or chymotrypsin yields non-infectious virus. We have analyzed the cleavage sites of HI, H3 and HlO hemagglutinins. The amino acid sequences at the cleavage site, i. e. C-terminus of HAl and the N-terminus of HA 2 are identical when virus is activated in vivo and in vitro. Under both conditions, an arginine residue connecting HAl and HA 2 in the precursor is eliminated. The elimination of argmme results in a shift of the isoelectric point of the hemagglutinin as demonstrated by isoelectric focusing. Non-activating enzymes cleave only one peptide bond in the HA 2-Nterminal region of activated HA and thus do not affect the isoelectric point. We have also analyzed the cleavage site of the hemagglutinin of fowl plaque virus (H7). A connecting peptide containing several basic amino acids is eliminated. -The data show that activation of the influenza hemagglutinin involves the action of a cellular protease with trypsin-like specificity followed by the action of an exopeptidase of the carboxy-peptidase B-type. The latter enzyme activity is associated with purified virus and can be analyzed by an assay employing peptides bearing 3H-arginine at the C-terminus. The carbohydrates of the glycoproteins of 21 influenza A strains containing hemagglutinin and neuraminidase of all serotypes known to date have been compared by analysis of glycopeptides labeled with radioactive sugars. Analysis of incompletely glycosylated glycoproteins synthesized in the presence of glycosylation inhibitors allowed the determination of the number of oligosaccharide side chains on HA 2 • With all strains, the neuraminidase contains side chains of both the complex type I and the mannose-rich type II. There are distinct quantitative and qualitative differences between the strains in the distribution of type I and type II side chains on the hemagglutinin fragments HAl and HA 2 • The majority of the hemagglutinin oligosaccharides is located on HAl' These side chains are usually of type 1. Only the hem agglutinins of serotype H3 have, in addition, a substantial amount of type II side chains on HAl' Most strains have on HA 2 a single side chain which is usually of type I. With serotype HS this side chain is free of fucose, and with serotype H8 it appears to be missing completely. Serotypes H7 and H10 have, in addition to the type I, a type II side chain on HA 2 • These observations strengthen the concept that the primary structure of the polypeptide chain is an important determinant for the carbohydrate moiety of the hemagglutinin.

Inst. f. Virologie, ]ustus-Liebig-Univ., D-6300 Giessen Acylation of Viral Glycoproteins SCHMIDT, M. F. G. Covalent binding of fatty acids to viral glycoproteins was first detected with Sindbis virus and vesicular stomatitis virus (1, 2) . Studies on acylation in other enveloped viruses revealed that covalent addition of fatty acid to spike glycoproteins is a more general feature. While in Sindbis and in Semliki Forest virus both species of spike glycoprotein (El and E2) carry fatty acid chains, acylation with the other viruses studied (corona-, influenza A-and paramyxovirus family) seems restricted to those glycoproteins that are known to carry fusion activity. This new type of modification of viral glycoproteins occurs in a wide variety of host cells including those of human, bovine, mouse, hamster, avian and insect origin (3) . -With the aid of controlled digestion of 3H-palmitic acid labelled virus particles and by the analysis of cyanogen bromide fragments of fatty acid labelled glycoprotein the fatty acid bind ing site in influenza hemagglutinin (HAt), VSV G-protein and Sindbis virus E1 and E2 could be located to the membrane spanning portion of the respective proteins (3).

Heinrich-Pette-Institut f. Exp. Virologie u. Immunologie an d. Univ., Martinistr. 52, D-2000 Hamburg 20 

MANNWEILER, K., BOHN, W., RUTTER, G., and HOHENBERG, H.

In replica-immunocytochemical (RIC) -and ultrathin section (US) preparations the ultrastructures of the specific alterations and of virus antigens, which appear at the plasma membrane of He La cell coverslip cultures after infection with an adapted measles virus strain were investigated. As immunomarker protein A-coated gold particles were used (1) . This method is sufficiently sensitive to enable labeling of even small altered areas of the plasma membrane (70-100 nm Cb). Due to the high atomic number contrast in the TEM and the small size of the marker (-<:: 10 nm) the ultrastructure of characteristic alterations morphologically still remains visible with ease in a three-dimensional aspect. Data obtained by RIC and US preparations after labeling with antimeasles immune serum or with monoclonal antibodies against HA (2) are demonstrated, compared and discussed.

BOHN, W., RUTTER, G., and MANNWEILER, K.

By use of the mouse hybridoma technique, monoclonal antibodies were obtained with specificity for the HA (79K), P (72K) and M (36K) polypeptides of measles virus. Balblc mice were immunized with native measles virus and measles virus treated with detergents and heat. Clones obtained after immunization of mice with native measles virus showed specificity for the HA polypeptide only. After immunization with measles virus, treated with 1 0/0 sodium sarkosyl sulfate (SSS) at 20°C a clone was obtained producing antibodies to the M polypeptide. Heating of measles virus in the presence of 1 0/0 SDS under reducing conditions elicited a selective immune response to the P and NP polypeptides. Thus, clones producing antibodies to the P polypeptides were isolated. contains a 50s RNA, but it does not show any infectivity even after trypsin treatment. An activation of the 6/94 cl virus can be obtained by i) cocultivation of the CL-E-8 cells with standard cells (e. g. BHK-21) and ii) serial passaging of the 6/94 cl virus in several cell lines (e. g. BSC-1). Infectious 6/94 cl virus can be detected in the cell supernatant after a period of 5-10 days and 25-30 days respectively. This virus cannot be propagated in chicken eggs, but it can replicate in serial cell cultures without trypsin treatment; moreover trypsin treatment does not influence the viral replicaiton. In comparison 6/94 virus released from an in-vitro generated persistent infection (BSC-1 cells infected with egggrown 6/94 standard virus) can be propagated in chicken eggs. This virus termed 6/94 pi virus also can grow on serial cell passages without trypsin treatment, whereas the serial replication of 6/94 standard virus on cell cultures depends on the trypsin-activation.

Inst, f. Virologie u. Immunbiologie, Versbacher Str. 7, D-8700 Wiirzburg

MONZEL, P. and KOSCHEL, K.

The paramyxvviruses measles (SSPE) virus and canine distemper virus (CDV) cause an impairment of the catecholamine induced p-adrenergic receptor dependent c-AMP generation in persistently infected C6 rat glioma cells. In CDV persistently infected C6 cells the number of receptors is greatly reduced. Hirata and Axelrod have shown that the number of Ii-adrenergic receptors could be regulated by methylation of phosphatidyl ethanolamine (PE) resulting in lecithin synthesis (1) . We have therefore studied the methylation of PE in persistently infected cells by the incorporation of (3H) methyl groups from (3H-methyl)-Methionine into PE. In both infected systems, C6/SSPE and C6/CDV, we observed a total loss of catecholamine stimulated p-adrenergic receptor dependent methylation whereas the p-receptor independent methylation of phospholipids is unchanged.

Inst. f. Med. Virologie d. Univ., D-6900 Heidelberg; t Robert-Koch-Institut, D-1000 Berlin 65; 2 Inst. f. Virusforschung, DKFZ, D-6900 Heidelberg KURZ, W., GELDERBLOM, H.t, FLOGEL, R. M.2, and DARAI, G. A paramyxovirus was isolated from a kidney biopsy of a Tupaia (three shrew) and termed (TPV). The detailed host range study revealed that only Tupaia embryonic fibroblasts and Tupaia kidney cells are the cells of choice for the efficient propagation of TPV. TPV can be plaque-assayed on Tupaia embryonic fibroblasts and this cell line was used for the continued propagation of TPV. Electron microscopy of purified TPV revealed the presence of typical paramyxovirus particles. -The hemagglutination test was performed with erythrocytes with a variety of different species. It was found that guinea pig erythrocytes were agglutinated with TPV. The buoyant density of purified virions was determined in sucrose gradient and found to be 1.19 g X ml", -The biological chara cterization of TPV which was perfo rmed by host range study in vivo revealed that TPV is highly pathogenic for new born mice and hamsters. -The characterization of viral RNA and proteins of th is new member of paramyxo viridae is now in progress. Coronaviruses contain two glycoprotein species £2 (180 K) and £1 (23 K) which are both synthesized in the R£R of the infected host cell. Glycosylation of £2 is initiated at the cotranslational level and it can be inhibited by 2-deoxyglucose and tunicamycin indicating the presence of N-glycosidic carbohydra te protein linkages. Particles formed in the presen ce of these inh ibit ors are non infectious and lack detect able amounts of £2. -Cell fractionation experiments show that glycosylation of glycoprotein £1 occur s posttranslationally in smooth memb ranes. The carb ohydrate protein linkages in £1 are susceptible to mild alkaline reductive conditions and N-acetylgalactosamine was determined to be the reduc ing sugar of the released oligo saccharides . This together with the finding that glycosylation of £1 is not sensitive to inhibito rs of N-glycosylation suggests that glycoprotein in £1 of coronaviruses is the first structural virus glycoprotein containing O-glycosidic side chains exclusively.

Insr, £. Virologie, jusrus-Liebig-Univ., D-6300 Giessen Target Cells of Infectious Bursal Disease Virus (IBDV) of Chickens MOLLER, H. and BECHT, H. Infectious bursal disease virus (IBDV), the causative agent of a highl y contagious disease of young chickens result ing in severe necrotic lesions in the bur sa of Fabricius (Gumboro disease), is a non-enveloped icosahedral particle with a diameter of about 60 nm. Its genome con sists of 2 segments of double-stranded RNA with mol ecular weight s of 2.2 X 10· and 2.5 X 10· daltons. Th e virion is composed of 5 structural pol ypeptides with mole cular weights of 90 kd, 48 kd. 40 kd, 32 kd and 28 kd. The 40 kd pol ypeptide, one of the two main structural proteins, is derived from the 48 kd pol ypeptide, perh aps by proteolytic modification (1) . With immunofluorescence and "i nfectious center assays" we were able to show th at 1. after infection of isolated lymphoid cells in vitro only 20 Ofo of bur sa cells, 2 % of thymus cells and 5 Ofo of spleen cells produce infective virus (i, e. plaques in infectious center assays) alth ough the donor chickens were in the most susceptible age of 4 to 5 weeks. 2. the number of virus producing cells is not cor related with the appe arance of slgM or sIgG. 3. virus yields seem to be influenced by the cell cycle: the number of chick emb ryo fibro blasts producing plaques in "infectious center assays" is increased after synchro nisa tion of the cells before infection. 4. cells that produce infective virus show an increased uptake of 3H-thymidine. 5. isolated lymphoid cells from thymus or spleen as well as blood lymphocytes can be stimulated by mitogens to produce higher virus yields. infection of Borna virus without any influence of the immunoresponse. The antigens first appeared in the nucleus of neurons and sometimes fibroblasts, then filled the cytoplasm, most brilliantly 7 days post infection. Thereafter they disappeared from the cytoplasm, but remained persistently only in the nucleus in point shape. No morphological changes were seen in the infected cells during 60 days post infection. We can say that the virus does not kill the cell. For the destruction of nerve cells in in vivo conditions it can be pointed to the importance of the immunological events, which might cause the clinical pictures.

Abr, Molekularbiologie, Physiol.-Chem. lnst., Grindelallee 117, D-2000 Hamburg 13; Heinrich Pette-Inst., Martinistr. 52, D-2000 Hamburg 20 Large Scale Production of Biologically Active VSV in EAT-Cells MACK, D., KRUPPA, J., and BREINDL, M.I Ehrlich ascites tumor cells maintained in mice were used to prepare milligram quantities of biologically active VSV. At the 6th day after passage when the cell number reached approx. 7 X 10 8 cells/mouse, mice were infected by intraperitoneal injection of appropriate concentrations of VSV. Ascites fluid was harvested after 20 h. Virus production was exponential for at least 16 h and continued for at least 20 h p.i. -Approximately 3-4 mg of viral protein/mouse and 2 X 1011 PFUlmouse were routinely obtained. The specific infectivity of VSV isolated from EAT-cells reached nearly 1.5 X 10 8 PFUI,ug protein. The endogenous transcriptase activity of VSV produced in EAT, BHK, and HeLa cells showed no significant differences. Large amounts of biologically active VSV may be produced rapidly and much less costly with the described procedure than using tissue culture cells. It should be possible to adapt the procedure for the production of other viruses.

Lonza A.G., CH-4000 Basel, Schweiz

The Quantitative Determination of Bardac-22, Formaldehyde, Glyoxal and Glutardialdehyde in Disinfectants WEINREIS, P. and GOLLER, S.

Several institutes of hygiene have been provided with two disinfected formulations, A and B, with the intention to analyse these mixtures, containing different amounts of quaternary ammonium compounds, formaldehyde, glyoxal and glutardialdehyde, -The quantitative analysis of the components of the two mixtures depends on well known volumetric and spectrophotometric methods. -The results of this analysis have shown that it is possible to use the described procedure for routine check ups of disinfectants without using microbiological tests.

Proc . nat. Acad. Sci

Virolog y (1982)

Previous studies have shown, that mouse-neurovirulent recombinants can be obtained from mixtures of avirulent influenza A-viruses provided one of the parents had previously been adapted to that host. This study shows that prior adaptation of parental strains is not necessary and that generation of neurovirulent recombinants is frequent. The gene constellation for neurovirulence was predictible for recombinants derived from a particular pair of parental strains

In influenza C virus -infected cells a virus-specific protein of a molecular weight of 65,000 Dalton can be detected when glycosylation is inhibited by Tunicamycin. Since this protein can not be found in untreated control cells, it probably constitutes the unglycosylated precursor of the viral glycoprotein gp 88

Since the petide pattern is identical for the doublet bands, it remains to be established whether this reflects a differential glycosylation or dissimilar proteolytic cleavage sites. The coding capacity of the viral genome RNA-segment No

Abstracts of the 38th Meeting of the OGHM 1 Augenklinik d. justus-Liebig-Univ., 0-6300 Giessen; 2 Inst. f. Neuropathologie u. Virologie, Freie Univ., 0-1000 Berlin Retino-Cerebral Manifestation of Experimental Borna Disease In Rhesus Monkeys KREY, H.t, ROGGENDORF, W.2, and LUDWIG, H.3 Inflammation of the uveal tract, the retina meninges and brain represent uveo-meningoencephalitic syndromes of unknown origin. One of these, the Vogt-Koyanagi-Harada syndrome is primarily manifested by inflammation of the retinal pigment epithelium, the uveal tract and meninges. Severe visual and neurological impairment can occur. In our experimental studies 14 rhesus monkeys were experimentally infected with Borna disease virus. After a 4-7 week incubation period a progressive retino-cerebral syndrome was observed. Focal inflammatory lesions in the retinal pigmentepithelium and the uveal tract were accompanied by encephalitic and meningeal infiltrates. Infectious virus could be demonstrated in the retina and in the brain. Experimental Borna disease can serve as an appropriate model for uveo-meningo-encephalitic syndromes in men. Borna Disease (BO) virus induced encephalitis in horses, sheep, rodents and primates shows similarities in many aspects with other so-called slow virus diseases. The EEG has shown to be a specific tool in studying encephalitides of different types in man. This is a report on the EEG of the BO virus specific encephalitis. Twenty-eight rabbits inoculated by different routes and with different virus preparations were screened for EEG changes. The basic frequency was measured optically and an analysis of the EEG was performed. The following conclusions were made: 1. A significant slowing down of the basic frequency was observed in the EEGs of BO virus infected rabbits. 2. Spikes and spike-waves were present rather regularly and correlated with epileptic seizures at the end of the disease when they appeared rhythmically in intervalls of twenty seconds. Rademakercomplexes appeared from the third week on. Virological and serological data collected from all animals demonstrated a strong correlation of BO virus specific reactions with the EEG alterations. The patterns of EEG changes are reminiscent of those in SSPE. EEG features in this kind of slow virus diseases may have rather similar characteristics which could suggest that they may underlie common pathophysiological mechanism. We established the primary cultures of neural retina and pigment epithelium of rabbit and of the brain of chicken embryo to study the sensitivity of each kind of cells to the