The prevalence of proviral bovine leukemia virus in peripheral blood mononuclear cells at two subclinical stages of infection.

The bovine leukemia virus (BLV) is an oncogenic retrovirus that is associated with the development of persistent lymphocytosis (PL) and lymphoma in cattle. While B lymphocytes have been shown to be the primary cellular target of BLV, recent studies suggest that some T lymphocytes and monocytes may be infected by the virus. Because virally altered functions of monocytes and/or T cells could contribute to the development of lymphoproliferative disease, we sought to clarify the distribution of the BLV provirus in subpopulations of peripheral blood mononuclear cells in seropositive cows with and without PL. CD2+ T cells, monocytes, and CD5+ and CD5- B cells were sorted by flow cytometry and tested for the presence of BLV by single-cell PCR. We did not obtain convincing evidence that peripheral blood monocytes or T lymphocytes contain the BLV provirus in seropositive cows with or without PL. In seropositive cows without PL (n=14), BLV-infected CD5+ and CD5- B cells accounted for 9.2% +/- 19% and 0.1% +/- 1.8% of circulating B lymphocytes, respectively. In cows with PL (n=5), BLV-infected CD5+ and CD5- B cells accounted for 66% +/- 4.8% and 13.9% +/- 6.6% of circulating B lymphocytes, respectively. The increase in lymphocyte numbers in cows with PL was entirely attributable to the 45-fold and 99-fold expansions of infected CD5+ and CD5- B-cell populations, respectively. Our results demonstrate that B cells are the only mononuclear cells in peripheral blood that are significantly infected with BLV. On the basis of the absolute numbers of infected cells in seropositive, hematologically normal animals, there appear to be differences in susceptibility to viral spread in vivo that may be under the genetic control of the host.     

BoLA class I allele diversity and polymorphism in a herd of cattle

Major histocompatibility complex class I genes are among the most polymorphic genes characterized. The high level of polymorphism is essential for generating host immune responses. In humans, three distinct genomic loci encode human leukocyte antigen (HLA) class I genes, allowing individuals to express up to six different HLA class I molecules. In cattle, the number of distinct genomic loci are currently at least six, and the number of different bovine leukocyte antigens (BoLA) class I molecules that are expressed in individual animals are variable. The extent of allele variation within the cattle population is unknown. In this study, the number and variety of BoLA class I sequences expressed by 36 individuals were determined from full-length BoLA class I cDNA clones. Twenty distinct BoLA class I alleles were identified, with only four being previously reported. The number of expressed BoLA class I alleles in individual animals ranged between one and four, with none of the animals having an identical complement of BoLA class I molecules. Variation existed in the number of BoLA class I alleles expressed as well as the composition of expressed alleles, however, several BoLA class I alleles were found in multiple individual animals. Polymorphic amino acid sites were analyzed for positive and negative selection using the ADAPTSITE program. In the antigen recognition sites (ARS), there were eight positions that were predicted to be under positive selection and three positions that were predicted to be under negative selection from 62 positions. In contrast, for non-antigen recognition sites (non-ARS), there were three positions that were predicted to be under positive selection and 20 that were predicted to be under negative selection from 278, indicating that positive selection of amino acids occurs at a greater frequency within the antigen recognition sites.     

Associations between strain,herd size,age at first calving,culling reason and lifetime performance characteristics in Holstein-Friesian cows

Cow longevity and lifetime performance traits are good indicators of breeding effectiveness and animal welfare. They are also interrelated with the economics of dairy herd. Unfortunately, a high milk yield is often associated with deteriorated cow health and fertility and, consequently, with an increased culling rate. This situation, observed also in the Polish population of Holstein-Friesian cattle, inspired us to undertake a study on the associations between some factors and lifetime performance characteristics. The data set consisted of the records on 135 496 cows, including 131 526 of the Black and White strain (BW), and 3970 of the Red and White strain (RW) covered by performance recording and culled in 2012. It was found that cows of the BW strain and those from the largest herds (>100 cows) reached higher lifetime and mean daily energy-corrected milk (ECM) yields than cows of the RW strain and those from smaller herds culled at a similar age. Cows youngest at first calving (<2.0 years) were characterised by the highest lifetime ECM yield. It indicates that heifers can be bred even when they are younger than 15 to 16 months with no significant negative effect on their later performance. Infertility and reproduction problems (39.6%) and udder diseases (15.5%) constituted the most frequent reasons for cow culling. Cow longevity and lifetime productivity were considerably affected by the interactions between the studied factors.     

Evidence for BoLA-linked resistance and susceptibility to subclinical progression of bovine leukaemia virus infection

The role of the bovine major histocompatibility complex in bovine leukaemia virus (BLV) infection and disease progression was investigated in a herd of Shorthorn cattle (n = 117). The frequency of cows that were seropositive to BLV-glycoprotein antigen was 51%. Twenty-three per cent of the seropositive cows were lymphocytotic. At the herd level, relative resistance to BLV-dependent B-cell proliferation and lymphocytosis among seropositive cows was associated with bovine lymphocyte antigen (BoLA)-DA7, whereas susceptibility was associated with BoLA-DA12.3. These associations were also confirmed at the family level, where BoLA phenotypes were used as haplotypic markers. Among the offspring of one BoLA-heterozygous sire (n = 33), resistance segregated with the DA7 haplotype and susceptibility with the DA12.3 haplotype. In this sire group, maternal transmission of the BoLA-w8 allele was associated with increased susceptibility to B-cell proliferation and lymphocytosis in w8/DA12.3 heterozygotes. These data provide the first evidence that subclinical progression of BLV infection is under the control of the BoLA complex, and suggest that the BoLA system can be used to select for resistance to B-cell proliferation and the development of lymphocytosis in BLV-infected herds.     

In vivo infection of sheep by bovine leukemia virus mutants.

Direct inoculation of a cloned bovine leukemia virus (BLV) provirus into sheep has allowed study of the viral infectivity of genetic mutants in vivo. Three BLV variants cloned from BLV-induced tumors and 12 in vitro-modified proviruses were isolated and analyzed for viral expression in cell culture. The proviruses were then inoculated into sheep in order to assess viral infectivity in vivo. Of three variants cloned from BLV-induced tumors (344, 395, and 1345), one (344) was found infectious in vivo. This particular provirus was used to engineer 12 BLV mutants. A hybrid between the 5' region of the complete but noninfectious provirus 395 and the 3' end of mutant 344 was infectious in vivo, suggesting that the tax/rex sequences were altered in virus 395. As expected, several regions of the BLV genome appeared to be essential for viral infection: the protease, pol, and env genes. Even discrete modifications in the fusion peptide located at the NH2 end of the transmembrane gp30 glycoprotein destroyed the infectious potential. In contrast, mutations and deletions in the X3 region present between the env gene and the 3' tax/rex region did not interfere with viral infection in vivo. This region of unknown function could thus be used to introduce foreign sequences. A BLV recombinant carrying a ribozyme directed against the tax/rex sequences was still infectious in vivo. Cotransfection of two noninfectious mutants carrying deletions led to infection in two of four independent injections, the infectious virus being then a recombinant between the two deletants. The experimental approach described here should help to gain insight into essential mechanisms such as in vivo viral replication, cooperation between deletants for viral infectivity, and viral superinfections. The gene products in the X3 and X4 region which are dispensable for in vivo infection could be involved in leukemogenesis, and thus proviruses deleted in these sequences could constitute the basis for a live attenuated vaccine.     

The relationship between antibody status to bovine corona virus and bovine respiratory syncytial virus and disease incidence,reproduction and herd characteristics in dairy herds

Background  

Bovine respiratory syncytial virus (BRSV) and bovine corona virus (BCV) affects cattle worldwide. Our objective was to evaluate the effects of these infections on general health and reproduction parameters measurable on herd level and to explore the association between antibody status and some herd characteristics.     

Structure of a defective provirus of bovine leukemia virus

Defective proviruses of bovine leukemia virus (BLV) in the genomes of infected cells were investigated by using Southern blotting hybridization analysis with various portions of a cloned BLV DNA as probes. When nine independent tumors of enzootic bovine leukosis with a single proviral copy per cell were examined, a single defective provirus of BLV was found in one tumor and also in a bovine B cell line derived from this tumor. Hybridization analysis of this defective provirus revealed that it underwent deletion between the pol and env genes and contained no major deletion in the other regions.     

No involvement of bovine leukemia virus in sporadic bovine lymphosarcoma

Using various portions of a molecularly cloned bovine leukemia virus (BLV) DNA as probes, the possible integration of a BLV genome or a BLV-related sequence into the chromosomal DNA of sporadic bovine leukosis (SBL) tumor cells was investigated by Southern blotting analysis. Under stringent as well as nonstringent conditions of hybridization, neither BLV nor BLV-related sequence specific to SBL DNAs was detected in any SBL tumor examined. These results provide conclusive evidence for lack of the relation of BLV or a BLV-related agent to SBL.     

Using PCR for early diagnosis of bovine leukemia virus infection in some native cattle

Bovine leukemia virus (BLV), the causative agent of enzootic bovine leukosis, is an exogenous, B lymphotropic retrovirus belonging to the Retroviridae family that induces persistent lymphocytosis in cattle and sheep. PCR has proven to be particularly suitable for investigating herds of cattle with a very low incidence of BLV infection and for clarifying doubtful serological results obtained by immunodiffusion or ELISA. The native Iranian and Russian cattle have a series of valuable traits that discriminate them as unique breeds that are well able to compete with western analogues. However, their gene pools have not been analyzed with molecular markers, including detection of BLV by PCR. Two pairs of primers were used: gag1 and gag2, and pol1 and pol2, which encompass 347- and 599-bp fragments of the BLV gene, respectively. Sixty-five Iranian Sistani, 120 Yaroslavl, 50 Mongolian, and 35 Black Pied cows were investigated. Among these 270 animals, we obtained 42 positive and 15 doubtful results in the first PCR. The second PCR was very effective in increasing BLV test reliability data to support detection of BLV.     

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.     

Frequency of twin births among Holstein-Friesian cows in a warm dry climate

The objectives of this study were to examine 1) the effect of parity of dam and season of birth on the incidence of twins in Holstein-Friesian cattle in a warm, dry climate and 2) the sex ratio, gestation period and birth weight of calves born. Natural calvings (n = 24,843) from nine dairy farms in Saudi Arabia were evaluated over an 8-year period. The calendar year was divided into six bimonthly intervals to identify a possible seasonal effect on the number of natural twins born (twinning rate). The twinning rate increased (P < 0.05) from 1.0% (103 10,094 ) at first calving to 8.0% (1,178 14,749 ) for all the subsequent calvings. The twinning rate was similar with the second calving (P(2)) at 7.0% (417 5,929 ), the third calving (P(3)) at 7.5% (218 2,896 ), and the fourth calving (P(4)) at 7.8% (147 1,864 ). Correspondingly, cows having >/= five calves had a higher (P < 0.01) twinning rate at 9.0% (164 1,805 ) than cows with too, three or four calvings collectively at 7.3% (782 10,689 ). The seasonal effect on twinning rate was significant within all parity calvings except in primiparous females. The peak twinning rate occurred in the third season interval (May/June) at 9.3% (66 708 ) with P(2), at 13.0% (34 262 ) with P(3), at 13.8% (27 196 ) with P(4), and at 12.5% (23 184 ) with P(5) females. The male to female ratio was different (P < 0.01) for multiparous females (55:46; n = 12,305). The ratio of males to females (55:45) was also different (P < 0.01) for all twin pairs (n = 1,151), evaluated in the study. The gestation period for cows with single calves was 8.5 days longer (P < 0.05) than that of cows carrying twin calves. The birth weight of single calves was 13.1 kg greater (P < 0.05) than that of twin calves. The reason for the marked increase in the twinning rate during the warmest months may be explained by greater embryo mortality in the hot vs cool months.     

Characterization of infertility and bovine leukemia virus infection in beef bulls on southwestern Louisiana coastal range

One hundred and twenty-six beef bulls on southwestern Louisiana coastal range were evaluated for breeding soundness. Samples were taken to determine the incidence of bovine leukemia virus (BLV) infection, and the prepuce was cultured for potential pathogens. A high incidence (47.6%) of questionable and unsatisfactory potential breeders resulted mainly from 37.0% of the bulls exhibiting high numbers of abnormal sperm cells in the semen. Only bulls in the 4-to 5-yr age group exhibited the expected incidence of normal spermiograms. Genital campylobacteriosis was not diagnosed but there was genital trichomoniasis in three of the seven herds. Hemophilus somnus , mycoplasma and ureaplasma were isolated from the prepuce of 13.3, 48.8 and 36.7% of the bulls, respectively. Isolation of these organisms from the prepuce did not appear to be associated with abnormal spermiograms. Of the bulls studied, 34.4% had positive AGID reactions for BLV. Bulls seropositive to BLV had an increased incidence of leukocyte counts that were above the normal range. There was no apparent relationship between BLV infection and abnormal spermiograms.     

Evaluation of radioimmunoprecipitation for the detection of bovine leukemia virus infection in domestic cattle.

The potential application of a recently developed radioimmunoprecipitation test for antibody directed against the major structural protein of bovine leukemia virus was evaluated for use in detection of BLV-infection in domestic cattle. This technique was found to be considerably more sensitive than serologic procedures currently being utilized for this purpose. Radioimmunoprecipitation was also shown to have distinct advantages as compared to hematologic criteria, such as specified by Bendixen's index, for identification of BLV-infected animals. By the use of radioimmunoprecipitation, high levels of antibody to BLV were demonstrated in sera of animals with confirmed adult lymphosarcoma, but not in animals with a less common sporadic form of the disease which occurs in calves.     

Expression and genetic segregation of parental BoLA serotypes in bovine embryos

The detection of parentally derived BoLA serotypes was attempted in 68 bovine embryos. 23 bovine embryos were tested for genetic segregation of maternally derived BoLA serotypes. 45 bovine embryos were tested for genetic segregation of paternally derived BoLA serotypes. The expected parentally derived BoLA gene products were detectable on approximately 50% of the embryos tested. A 1:1 segregation ratio of expression or non-expression of parental BoLA serotypes in 7-day-old preimplantation bovine embryos, which is expected for codominant alleles, could not be rejected.     

Association between Epstein-Barr virus infection and chemoresistance to docetaxel in gastric carcinoma

Epstein-Barr virus (EBV) is associated with human cancers such as nasopharyngeal carcinoma, Burkitt’s lymphoma, Hodgkin’s disease, and gastric carcinoma (GC). EBV is associated with about 10% of all GC cases globally. EBV-associated GC has distinct features from EBV-negative GC. However, it is still unclear if EBV infection has any effect on GC chemoresistance. Cell proliferation assay, cell cycle analysis, and active caspase Western blot revealed that the EBV-positive GC cell line (AGS-EBV) showed chemoresistance to docetaxel compared to the EBV-negative GC cell line (AGS). Docetaxel treatment increased expression of Bax similarly in AGS and AGS-EBV cell lines. However, Bcl-2 induction was markedly higher in AGS-EBV cells, after docetaxel treatment. Although docetaxel increased the expression of p53 to a similar extent in both cell lines, induction of p21 in AGS-EBV cells was lower than in AGS cells. Furthermore, expression of survivin was higher in AGS-EBV cells than in AGS cells following docetaxel treatment as well as at basal state. EBVlytic gene expression was induced by docetaxel treatment in AGS-EBV cells. The results suggest that EBV infection and lytic induction confers chemoresistance to GC, possibly by regulating cellular and EBV latent and lytic gene expression.     

Is there a relationship between genetic merit and enteric methane emission rate of lactating Holstein-Friesian dairy cows?

The present study was undertaken to examine the effect of cow genetic merit on enteric methane (CH₄) emission rate. The study used a data set from 32 respiration calorimeter studies undertaken at this Institute between 1992 and 2010, with all studies involving lactating Holstein-Friesian dairy cows. Cow genetic merit was defined as either profit index (PIN) or profitable lifetime index (PLI), with these two United Kingdom genetic indexes expressing the expected improvement in profit associated with an individual cow, compared with the population average. While PIN is based solely on milk production, PLI includes milk production and a number of other functional traits including health, fertility and longevity. The data set had a large range in PIN (n=736 records, −£30 to +£63) and PLI (n=548 records, −£131 to +£184), days in milk (18 to 354), energy corrected milk yield (16.0 to 45.6 kg/day) and CH₄ emission (138 to 598 g/day). The effect of cow genetic merit (PIN or PLI) was evaluated using ANOVA and linear mixed modelling techniques after removing the effects of a number of animal and diet factors. The ANOVA was undertaken by dividing each data set of PIN and PLI into three sub-groups (PIN:<£3, £3 to £15 and >£15, PLI:<£23, £23 to £67 and >£67) with these being categorised as low, medium and high genetic merit. Within the PIN and PLI data sets there was no significant differences among the three sub-groups in terms of CH₄ emission per kg feed intake or per kg energy corrected milk yield, or CH₄ energy (CH₄-E) output as a proportion of energy intake. Linear regression using the whole PIN and PLI data sets also demonstrated that there was no significant relationship between either PIN or PLI, and CH₄ emission per kg of feed intake or CH₄-E output as a proportion of energy intake. These results indicate that cow genetic merit (PIN or PLI) has little effect on enteric CH₄ emissions as a proportion of feed intake. Instead enteric CH₄ production may mainly relate to total feed intake and dietary nutrient composition.     

Association between HLA class II antigens and hepatitis C virus infection

The aim was to confirm the influence of HLA Class II antigens on the progression of HCV infection and to assess the relationship between these antigens and histological damage, HCV viral load and HCV genotype. 143 patients were enrolled and divided into three groups. Group A included 34 anti-HCV positive, HCV-RNA negative patients with ALT persistently normal; group B included 39 patients with HCV-RNA positive and abnormal ALT level; group C included 70 normal subjects. Serological HCL typing was performed with lymphocytotoxicity test by Terasaky and McClelland, using lymphobeads HLC class II. The frequency of HLA DR11 (5) was significantly higher in the control group (52.9%) and in group A (64.7%), than in group B (28.2%). Allele HLA DR6 was demonstrated in a similar proportion (26%) among control group and group B, while HLA DR14 (6) was less frequent among controls (18% vs 1.4%). In group A the frequency of HLA DR14 (6) was 3% compared to group B, HLA DR17 (3) was prevalent (15.4%) in group B. Liver damage was associated with the detection of HLA DR14 (6) and HLD DR17 (3) antigens. Significantly lower levels of HCV-RNA were measured in subjects with HLA DR11 (5) than in these with either DR6 or DR17 (3). HLA class II antigens appear crucial for resolution or progression of HCV hepatitis. The punctual identification of these genetic factors may, therefore, prove to be useful in predicting disease evolution, in guiding the appropriate therapy for patients with poor prognosis, and in encouraging the development of now therapeutic strategies.     

Estrus detection and estrus characteristics in housed and pastured Holstein-Friesian cows

This study compared three methods of estrus detection and characteristics of standing estrus between dairy cows kept in cubicle housing and fed a total mixed ration diet (HOUSED treatment) and those kept at pasture with concentrate ration supplementation (PASTURE treatment). The 23 spring-calved Holstein-Friesians in each treatment were monitored by three estrus detection methods simultaneously—visual observations, tail paint and radiotelemetry (HeatWatch)—for 9 wk. Milk progesterone profiles were used to determine the dates of true standing estrus events. All three detection methods had a higher efficiency of estrus detection in the PASTURE treatment than in the HOUSED treatment (P < 0.001), but there was no difference in the accuracy of estrus detection between the two treatments (P > 0.05). Within each treatment there was no difference between the efficiency and accuracy of the three methods (P > 0.05). HeatWatch was as efficient as visual observations at detecting standing estrus events. However, during visual observation sessions all occasions when animals were observed standing to be mounted were not recorded by HeatWatch. More cows expressed sub-estrus events and fewer expressed standing estrus events in the HOUSED than in the PASTURE treatment (P < 0.05). The interval between parturition and the second standing estrus was longer in the HOUSED treatment than in the PASTURE treatment (P < 0.05). All three detection methods were much less effective in the HOUSED than in the PASTURE treatment. This is because all of the detection methods tested relied solely on standing to be mounted and this was reduced in the HOUSED cows. Alternative approaches to estrus detection are needed for cows kept indoors on concrete.