Failure to find an association between ocular squamous cell carcinoma and class I antigens of the bovine major histocompatibility system*

Summary. We tested 53 cattle with ocular squamous cell carcinoma (cancer-eye) and 53 paired, matched controls for 25 class I antigens of the bovine major histocompatibility system. The most common antigen was W5 which was present in 40% of the animals with cancer-eye and 36% of the controls. There were no statistically significant differences in BoLA antigen frequency between cattle with and cattle without cancer-eye.     

BoLA antigens are associated with increased frequency of persistent lymphocytosis in bovine leukaemia virus infected cattle and with increased incidence of antibodies to bovine leukaemia virus

The association between bovine major histocompatibility system (BoLA) type and persistent lymphocytosis in cattle with antibodies to bovine leukaemia virus was examined by comparing antigen frequencies in cattle with persistent lymphocytosis to controls matched for age, sex, breed and presence of antibodies to BLV. The cattle came from nine dairy herds in south-east Queensland, Australia; six herds were Australian Illawarra Shorthorn (AIS), two herds were Jersey and one herd was Friesian. Antigen W6 and Eu28R were more common in cattle with persistent lymphocytosis than in controls. Antigen W8 was less common in AIS cattle with persistent lymphocytosis. A study of 24 offspring from one sire, heterozygous for W10 and Eu28R, showed that offspring inheriting Eu28R from the sire were significantly more likely to have antibodies to BLV than offspring inheriting the opposing W10 haplotype.     

Identification of antigenic differences that discriminate between cattle vaccinated with Anaplasma centrale and cattle naturally infected with Anaplasma marginale

Monoclonal antibodies were raised against the vaccine strain of Anaplasma centrale used in Australia. A monoclonal antibody that reacted with an 80 kDa antigen was used to develop an A. centrale-specific fluorescent antibody test that will be useful for confirming species identity in patent infections. Another monoclonal antibody that reacted with a 116 kDa antigen was used to develop an A. centrale-specific competitive inhibition enzyme-linked immunosorbent assay (ELISA) for the serological identification of vaccinated cattle. The sensitivity of the ELISA was 100% in cattle experimentally infected with A. centrale, 97.1% in a vaccinated beef herd and 98.3% in a vaccinated dairy herd. The specificity of the ELISA was 98.6% in non-vaccinated cattle outside the Anaplasma marginale-endemic area, 97.9% in non-vaccinated cattle within the A. marginale-endemic area and 100% in cattle experimentally infected with A. marginale. The ELISA detected antibodies to A. centrale in cattle up to 9 years after vaccination with no apparent decrease in sensitivity. The assay has proved extremely valuable in Australia for investigating reported failures of multivalent live vaccines used to protect cattle against anaplasmosis and babesiosis, and should be similarly useful elsewhere in the world where these types of vaccines are used, e.g. Israel and South America.     

Japanese Graves' disease: association with HLA-Bw46

HLA-A, -B, and -C antigen frequency of 61 Japanese patients with Graves' disease, and HLA-DR and -DQ frequency of 53 patients with it were compared that of 1,998 controls. The prevalence of Bw46 was significantly higher in patients with Graves' disease (23.0% vs 8.4%, Pc less than 0.003). The result was quite different from previous reports on Japanese Graves' disease.     

Genetic characteristics of the Yakut aboriginal cattle and its hybrids

Materials are presented concerning the analysis of blood groups in the Yakutsk aboriginal cattle (n = 960) and its hybrids (n = 145) bred in Verkhoyansk and Yakutsk regions of Yakutskaya ASSR with respect to the problems of its origin. The genetic similarity of the Yakutsk cattle and its hybrids was evaluated; the cattle populations were studied in various years. The experiment also involved some hybrids bred in the northern part of the Soviet Union. The strongest similarity was found for the Yakutsk aboriginal breed and the cattle bred in Kalmytskaya and Buryatskaya Regions (0.841-0.839), i.e. those which are ascribed to the same group, according to their origin. Similarity with the Ashire and hybrids of Zebu and Black and White cattle was found to be less pronounced. The frequencies of antigens ranged from 0.2 (the antigen H") to 100% (the antigen F). The antigen frequencies were found to be higher in purebred animals, as compared to the hybrids almost within the whole antigen spectrum.     

Prevalence of Human Herpesvirus 6 Variants A and B in Patients with Chronic Fatigue Syndrome

Peripheral blood mononuclear cells collected from 13 patients with chronic fatigue syndrome and 13 healthy controls were analyzed for the presence of human herpesvirus 6 (HHV-6) DNA by variant-specific polymerase chain reaction and dot blot hybridization. HHV-6 DNA was detected in 7 of 13 (53%) patients, and of those 7 patients, 4 were positive for HHV-6 variant A DNA and 3 were for variant B. No HHV-6 DNA was detected in the controls. Serum antibody titers to the late antigen and antibody prevalence to the early antigen of HHV-6 were significantly higher in the patient group. These results suggest active replication of HHV-6 in patients with chronic fatigue syndrome.     

A survey of seropositivity to antigen B,an immunodiagnostic antigen for human cystic echinococcosis,in domestic animals in Mongolia

Cystic echinococcosis (CE) is well known to be an important zoonotic disease and national disease due to the traditional nomadic life style in Mongolia. The present study was carried out to obtain data on the seropositivity to antigen B, in domestic livestock, goats, sheep and cattle, in each province of Mongolia. The seropositivity to antigen B varied by province and ranged from 0% to 25.0% in goats, 0% to 12.5% in sheep, and 0% to 13.3% in cattle. In total, 9.2% of goats, 3.6% of sheep and 5.9% of cattle in Mongolia showed seropositivity.     

Class I antigens of the bovine major histocompatibility system are weakly associated with variation in faecal worm egg counts in naturally infected cattle

Three bulls selected for high faecal worm egg counts and three bulls selected for low faecal worm egg counts were mated to Africander-Hereford cross cows. Faecal worm egg counts were taken on four occasions from the 132 offspring. Also, each animal was typed for 32 class I antigens of the bovine major histocompatibility system (BoLA). Least squares analysis of variance showed that line, sex and some of the antigens were associated with differences in worm egg output in the faeces. After adjusting for the effects of line and sex, cattle with antigen W9 had about twice as many worm eggs in their faeces as cattle without W9; cattle with antigen CA45 had about half the concentration of faecal worm eggs as cattle without CA45. However, the antigen associations were of borderline significance at the 5% level and more work in additional populations is necessary to confirm these associations.     

Immunohistochemical expression of retinoblastoma gene product (Rb), p53 protein, MDM2, c-erbB-2, HLA-DR and proliferation indices in human urinary bladder carcinoma

Archival biopsy specimens from transitional cell bladder cancers (n=88) were analysed immunohistochemically for the expression of the retinoblastoma (Rb) gene protein, p53, mdm2, c-erbB-2, HLA-DR antigen and proliferation indices. An altered nuclear expression of Rb, p53 and mdm2 was observed in 55.2%, 33.3% and 18.2% of tumors respectively. Cytoplasmic membrane immunoreactivity (>25% tumor cells) for c-erbB-2 was detected in 14.1% of tumors and aberrant HLA-DR antigen cytoplasmic staining (>5% of tumor cells) in 22.2% of the cases. P53 overexpression was associated with higher tumor grade and stage. Aberrant HLA-DR antigen expression and PCNA were also correlated with the grade of differentiation and tumor stage. MIB1 was statistically correlated with stage. pRb scores and HLA-DR antigen expression were correlated with proliferation activity as determined by PCNA and MIB1 immunostaining. p53 protein was also strongly correlated with the proliferation index PCNA. A strong correlation between PCNA and MIB1 (p<0.0001) was also found. In addition a statistically positive correlation between p53 and HLA-DR antigen expression was observed. Our data show that, although pRb and p53 protein expressions are not associated between them, they may contribute to the growth fraction of the bladder cancer. In addition, p53 and HLA-DR antigen expression could be indicators of aggressive behavior of bladder cancer.     

Preliminary studies of the complement fixation test to confirm the diagnosis of bovine ephemeral fever

A strain of bovine ephemeral fever (BEF) virus isolated in China in 1976 was adapted to growth in tissue cultures. A baby hamster kidney complement fixing (CF) antigen, stable at -20 degrees C for at least 120 days, was prepared from the BEF virus grown in tissue culture and used to test bovine sera for antibodies to that virus. CF antibodies were detected in all of 31 cattle after convalescence from experimental infection with BEF virus, in 208 (98%) of 213 cattle observed to have shown clinical ephemeral fever in an epidemic, in 96 cattle in these herds which did not show clinical signs of ephemeral fever and 16 cattle from herds in northern China outside the epidemic area. The CF antibodies to BEF virus were found to persist in 34 (89%) of 38 cattle which were bled 6 years after natural exposure to ephemeral fever. The CF antigen is economical to prepare and is suitable to differentiate ephemeral fever from other viral infections with which it could possibly be confused on clinical appearance.     

Fasciola hepatica: influence of extrahepatic adult flukes on infections and immunity in rats

Surgical transfer of adult Fasciola hepatica from sheep, goats, and cattle to subcutis of rats 4 wk before infection with metacercariae resulted in a 50% decrease in infection rate as compared to nonoperated controls.Infection was established in 25 out of 77 rats with F. hepatica implants, while 54 out of 79 were infected in the control group. The protective effect of the fluke implantation is discussed. It is suggested that production of protective antibodies is stimulated by the undamaged living flukes, although the antigen itself has not been demonstrated.     

Production and biochemical characterization of the recombinant Boophilus microplus Bm95 antigen from Pichia pastoris

The new antigen Bm95 from the cattle tick Boophilus microplus was recently isolated, cloned and expressed in the methylotrophic yeast Pichia pastoris. The recombinant protein has shown to induce protection in cattle against infestations of B. microplus under controlled and production conditions. In this paper we report the production and large-scale purification of the Bm95 protein, following a simple and cost-effective process. The antigen was obtained highly aggregated, forming particles ranging from 26 to 30 nm and with purity higher than 80%. The process yield was 0.55 g of pure Bm95 protein per liter of culture. The 98% of the primary structure of the recombinant protein was verified by mass spectrometry. Three amino acid changes in comparison with the sequence deduced from cDNA were detected by LC-MS/MS. The antigen was also obtained N-glycosylated, as previously reported for heterologous protein expression in P. pastoris.     

A monoclonal antibody-based immunoradiometric assay for detection of circulating antigen in Bancroftian filariasis

A monoclonal antibody designated Gib 13 has been used in an immunoradiometric assay (IRMA) to detect circulating antigen in the sera of Wuchereria bancrofti-infected subjects from an endemic area of Papua New Guinea. A clear association between the presence of patent infection and the Gib 13 target epitope in serum was established because 93% of microfilaremic individuals were antigen-positive. Moreover, there was a significant correlation between levels of serum antigen and blood microfilarial counts. Detection of circulating antigen in amicrofilaremic subjects with acute symptoms of lymphatic filariasis, and 53% of asymptomatic amicrofilaremic subjects, but not in nonendemic controls, suggests that the Gib 13 IRMA will also be of value in the diagnosis of occult filariasis. However, as in all IRMA based on detection of potentially immunogenic molecules in man, antibodies can be expected to be the major contributor to reduced sensitivity of the assay.     

Rift valley fever on the east coast of Madagascar

In March 1990, a Rift Valley fever virus (RVFV) outbreak was suspected in the district of Fenerive on the east coast of Madagascar after an abnormally high incidence of abortions and disease in livestock. Sera from humans and cattle were tested for RVFV antibodies by immunofluorescence assay (IFA) and ELISA-IgM capture. Sera and mosquitoes collected in the same area were tested for virus isolation by tissue culture and suckling mouse intracerebral inoculation, and for antigen detection by an ELISA antigen capture assay. Among cattle from the area, RVFV antibody prevalence was 58.6 % by IFA and 29.6 % by ELISA-IgM. In contrast, human populations in the same area had a lower RVFV antibody prevalence, with 8.01 % IFA and 5.4 % IgM-positive sera. No RVFV antigen was detected and virus isolation was unsuccessful from the sera and mosquito pools tested. Different hypotheses concerning the emergence and diffusion of RVFV in this area and the occurrence of the outbreak are discussed.     

Autoantibody to glial fibrillary acidic protein in the sera of cattle with bovine spongiform encephalopathy

It is desirable to make the diagnosis in live cattle with bovine spongiform encephalopathy (BSE), and thus surrogate markers for the disease have been eagerly sought. Serum proteins from BSE cattle were analyzed by 2‐D Western blotting and TOF‐MS. Autoantibodies against proteins in cytoskeletal fractions prepared from normal bovine brains were found in the sera of BSE cattle. The protein recognized was identified to be glial fibrillary acidic protein (GFAP), which is expressed mainly in astrocytes in the brain. The antigen protein, GFAP, was also found in the sera of BSE cattle. The percentages of both positive sera in the autoantibody and GFAP were 44.0% for the BSE cattle, 0% for the healthy cattle, and 5.0% for the clinically suspected BSE‐negative cattle. A significant relationship between the presence of GFAP and the expression of its autoantibody in the serum was recognized in the BSE cattle. These findings suggest a leakage of GFAP into the peripheral blood during neurodegeneration associated with BSE, accompanied by the autoantibody production, and might be useful in understanding the pathogenesis and in developing a serological diagnosis of BSE in live cattle.     

Complex formation between the lymphotropic papovavirus large tumor antigen and the tumor suppressor protein p53

The simian B-lymphotropic papovavirus (LPV) encodes a large tumor antigen (T antigen) which is 45% identical to both the simian virus 40 (SV40) and the polyomavirus (PyV) large T antigens. In transgenic mice, the transforming properties of the LPV T antigen are similar to those of the SV40 T antigen. However, little is known about its biochemical activities. Since SV40 T antigen forms a complex with and stabilizes the host cell tumor suppressor protein p53 while the PyV large T antigen does not, we characterized the LPV T antigen for its ability to complex p53. We demonstrate an association between LPV T antigen and p53 in both a tumor-derived cell line and BALB/c 3T3 cells transformed in culture. A third protein of approximately 68 kDa which was found associated with the LPV T antigen-p53 complex in tumor-derived cells appears to be heat shock protein 70 (hsp70). The half-life of p53 in all LPV T-antigen-transformed cells was extended significantly; i.e., it was 3 to 7 h compared with 19 minutes in BALB/c 3T3 cells. The half-life of the LPV T antigen itself was 5 to 9 h depending on the cell line origin. That p53 was stabilized because of association with LPV T antigen and not because of mutation was demonstrated with the p53 conformation-dependent monoclonal antibody PAb246. This antibody distinguishes between wild-type p53 (PAb246+) and mutant, oncogenic p53 (PAb246-). Sequential immunoprecipitation showed all detectable p53 to be of the PAb246+ class in each LPV-transformed cell line, suggesting that the stable p53 was indeed wild type.     

Use of an immunodominant P17 antigenic fraction of Neospora caninum in detection of antibody response in cattle

A Neospora caninum 17 kDa protein fraction (p17) has been described as an immunodominant antigen (IDA) under reducing and non-reducing conditions. The aim of the present study was to investigate the diagnostic utility of p17 in cattle. In order to achieve this, p17 was purified by electroelution from whole N. caninum tachyzoite soluble extract and a p17-based Western blot (WB-p17) was developed. The p17 recognition was measured by densitometry and expressed as OD values to check the validity of the WB-p17. A total of 131 sera including sequential samples from naturally- and experimentally-infected calves and breeding cattle were analysed by WB-p17 and compared with IFAT using whole formalin-fixed tachyzoites as a reference test. The results obtained highlight the feasibility of using the N. caninum p17 in a diagnostic test in cattle. Firstly, the assay based on the p-17 antigen discriminated between known positive and negative sera from different cattle populations, breeding cattle and calves. Secondly, the p17 antigen detected fluctuations in the antibody levels and seroconversion in naturally- and experimentally-infected cattle. Significant differences in p-17 antigen recognition were observed between naturally infected aborting and non-aborting cattle, as well as significant antibody fluctuations over time in experimentally infected cattle, which varied between groups. Furthermore, the results obtained with WB-p17 are in accordance with the results obtained with the IFAT as high agreement values were obtained when all bovine subpopulations were included (kappa = 0.86).     

Chlamydiosis in cattle and in man: an epidemiologic and serologic study

In the Pardubice area, the systematic examinations for Chlamydia-specific antibodies in cattle were started in 1967. In the years 1967-1978 the detectable levels of antibody to Chlamydia were demonstrated in 16.1% of 38,394 sera from cattle and in 18.6% of 5,244 sera from slinking cows. In the year 1979 the percentage of seropositivity in cattle increased significantly to 23.5% of the 4,197 sera examined. Examinations in 1980 revealed that 23.1% of 4,042 sera from cattle, 34.4% of 319 sera from slinking cows and 17.9% of 209 sera from calves were found antibody positive. Over the period from 1974 to 1978 isolates of Chlamydia organisms were obtained from 11.7% of 452 bovine slinks and from 10% of 271 dead or prematurely slaughtered calves. In the years 1979-1981 a total of 775 individuals were subjected to medical and serological examinations for Chlamydia infection using the BIOVETA group antigen. The first group of examinees included 307 farm personnel, attendants to healthy breeds of cattle. Detectable levels of antibody to Chlamydia were recorded in 1.3% of clinically healthy individuals and in 12% of the 75 healthy, randomly selected cows taken care of by this group of personnel. The second group included 280 individuals who were in charge of slinking cows or calves with chlamydial pneumonia, or who were employed in a rendering plant. A total of 7.5% of antibody-positive individuals was found in this group of examinees. Of the 439 calves attended 17.9% had serologic evidence of infection and the 30 dead calves examined were isolate-positive: sera from 176 slinking cows showed positivity in 23.8% of cases. In the third group of 188 persons serving as controls 5.3% had serologic evidence of chlamydial infection. The statistical analysis of deviations from the Gaussian variance normal curve showed neither sex-related nor inter-group differences. At the time of serologic examinations of humans the epidemiologic situation in the given area was extremely favorable. The importance of a close cooperation between the public health and veterinary service personnel (notification of abortions in cattle) and the need of a due attention of gynecologists, dermatologists and urologists are emphasized.     

Relationship of phosphorylation to the oligomerization of SV40 T antigen and its association with p53

The potential significance of the phosphorylation of SV40 large T antigen for oligomers and complexes with the cellular protein p53 was investigated. We observed that T antigen oligomers remain stable after enzymatic dephosphorylation by alkaline phosphatase up to 80%. Separate analysis of free and p53-bound T antigen revealed a considerably lower phosphorylation of the p53-bound subclass. Therefore, a simple correlation between the overall phosphorylation of T antigen and the formation of oligomers and T-p53 complexes is highly unlikely.     

Oral infection of calves with Neospora caninum oocysts from dogs: humoral and cellular immune responses

Neospora caninum has been identified as a major cause of abortion in cattle in a number of countries throughout the world. Until the recent demonstration that dogs can serve as a definitive host of this parasite, it was not possible to study the infection in cattle orally exposed to oocysts. The aim of this study was to investigate the potential of N. caninum oocysts to infect calves, and to define initial immune responses that arise after oral infection. Seven calves were fed approximately 10(4)-10(5) N. caninum oocysts, three calves served as uninfected controls. Before infection, all calves were serologically negative for anti-Neospora antibodies and the calves were non-reactive to Neospora antigen in an in vitro lymphocyte proliferation assay. Peripheral blood lymphocytes from inoculated calves were able to mount in vitro proliferative responses to crude N. caninum antigen extract as early as 1 week p.i. Within 2 and 4 weeks p.i., Neospora-specific IgG1 and IgG2 antibodies were detected by IFAT and ELISA in serum from infected calves but not from sham-infected calves. The continued presence of reactive cells in the blood, spleen and mesenteric, inguinal, bronchial lymph nodes was seen as late as 2.5 months p.i., and parasite DNA was detected in the brain and spinal cord of the infected animals by PCR, indicating that the cattle were infected by oral inoculation of N. caninum oocysts collected from dogs, and that the animals were systematically sensitised by parasite antigen.