BoLA-DRB3 Alleles Research Articles

First characterization of major histocompatibility complex class II DRB3 diversity in cattle breeds raised in Egypt

Genes encoding bovine leukocyte antigen (BoLA) enable the immune system to identify pathogens. Therefore, these genes have been used as genetic markers for infectious and autoimmune diseases as well as for immunological traits in cattle. Although BoLA polymorphisms have been reported in various cattle breeds worldwide, they have not been studied in cattle populations in Egypt. In this study, we characterized BoLA-DRB3 in two local Egyptian populations and one foreign population using polymerase chain reaction-sequence-based typing (PCR-SBT) method. Fifty-four previously reported BoLA-DRB3 alleles and eight new alleles (BoLA-DRB3*005:08, *015:07, *016:03, *017:04, *020:02:02, *021:03, *164:01, and *165:01) were identified. Alignment analysis of the eight new alleles revealed 90.7–98.9 %, and 83.1–97.8 % nucleotide and amino acid identities, respectively, with the BoLA-DRB3 cDNA clone NR-1. Interestingly, BoLA-DRB3 in Egyptian cattle showed a high degree of allelic diversity in native (na = 28, hE > 0.95), mixed (na = 61, hE > 0.96), and Holstein (na = 18, hE > 0.88) populations. BoLA-DRB3*002:01 (14.3 %), BoLA-DRB3*001:01 (8.5 %), and BoLA-DRB3*015:01 (20.2 %) were the most frequent alleles in native, mixed, and Holstein populations, respectively, indicating that the genetic profiles differed in each population. Based on the allele frequencies of BoLA-DRB3, genetic variation among Egyptian, Asian, African, and American breeds was examined using Nei’s distances and principal component analysis. The results suggested that native and mixed cattle populations were most closely associated with African breeds in terms of their gene pool, whereas Holstein cattle were more distinct from the other breeds and were closely related to Holstein cattle populations from other countries.

Modeling of PCR methods for primary screening of BoLA-DRB3 alleles

Modeling of PCR methods for primary screening of BoLA-DRB3 alleles

An immunoinformatics study reveals a new BoLA-DR-restricted CD4+ T cell epitopes on the Gag protein of bovine leukemia virus

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leucosis (EBL), which has been reported worldwide. The expression of viral structural proteins: surface glycoprotein (gp51) and three core proteins - p15 (matrix), p24 (capsid), and p12 (nucleocapsid) induce a strong humoral and cellular immune response at first step of infection. CD4+ T-cell activation is generally induced by bovine leukocyte antigen (BoLA) region– positive antigen-presenting cells (APC) after processing of an exogenous viral antigen. Limited data are available on the BLV epitopes from the core proteins recognized by CD4+ T-cells. Thus, immunoinformatic analysis of Gag sequences obtained from 125 BLV isolates from Poland, Canada, Pakistan, Kazakhstan, Moldova and United States was performed to identify the presence of BoLA-DRB3 restricted CD4+ T-cell epitopes. The 379 15-mer overlapping peptides spanning the entire Gag sequence were run in BoLA-DRB3 allele-binding regions using a BoLA-DRB- peptide binding affinity prediction algorithm. The analysis identified 22 CD4+ T-cell peptide epitopes of variable length ranging from 17 to 22 amino acids. The predicted epitopes interacted with 73 different BoLA-DRB3 alleles found in BLV-infected cattle. Importantly, two epitopes were found to be linked with high proviral load in PBMC. A majority of dominant and subdominant epitopes showed high conservation across different viral strains, and therefore could be attractive targets for vaccine development.

Low proviral load in the Kumamoto strain of Japanese Brown cattle infected with the bovine leukemia virus

BackgroundThe Kumamoto strain of Japanese Brown (JBRK) cattle is a sub-breed of Wagyu and has a different genetic background than that of Japanese Black (JB) cattle. Bovine leukemia virus (BLV) is the pathogen causing enzootic bovine leukosis (EBL), the predominant type of bovine leukosis (BL). EBL is one of the most common bovine infectious diseases in dairy countries, including Japan. Some host genetic factors, including the bovine leukocyte antigen (BoLA)-DRB3 gene, have been associated with the proviral load (PVL) of BLV and/or onset of EBL. Here, we determined the number of BL cases by analyzing prefectural case records in detail. We measured the PVL of BLV-infected JBRK cattle and compared it with that obtained for other major breeds, JB and Holstein-Friesian (HF) cattle. Finally, the relationship between PVL levels and BoLA-DRB3 haplotypes was investigated in BLV-infected JBRK cattle.ResultsWe determined the number of BL cases recorded over the past ten years in Kumamoto Prefecture by cattle breed. A limited number of BL cases was observed in JBRK cattle. The proportion of BL cases in the JBRK was lower than that in JB and HF. The PVL was significantly lower in BLV-infected JBRK cattle than that in the JB and HF breeds. Finally, in BLV-infected JBRK cattle, the PVL was not significantly affected by BoLA-DRB3 alleles and haplotypes. BoLA-DRB3 allelic frequency did not differ between BLV-infected JBRK cattle with low PVL and high PVL.ConclusionsTo our knowledge, this is the first report showing that BL occurred less in the JBRK population of Kumamoto Prefecture. After BLV-infection, the PVL was significantly lower in JBRK cattle than that in JB and HF breeds. The genetic factors implicated in maintaining a low PVL have yet to be elucidated, but the BoLA-DRB3 haplotypes are likely not involved.

Influence of BoLA-DRB3 Polymorphism and Bovine Leukemia Virus (BLV) Infection on Dairy Cattle Productivity

Enzootic bovine leukosis caused by the bovine leukemia virus (BLV) results in substantial damage to the livestock industry; however, we lack an effective cure or vaccine. BoLA-DRB3 polymorphism in BLV-infected cattle is associated with the proviral load (PVL), infectivity in the blood, development of lymphoma, and in utero infection of calves. Additionally, it is related to the PVL, infectivity, and anti-BLV antibody levels in milk. However, the effects of the BoLA-DRB3 allele and BLV infection on dairy cattle productivity remain poorly understood. Therefore, we investigated the effect of BLV infection and BoLA-DRB3 allele polymorphism on dairy cattle productivity in 147 Holstein dams raised on Japanese dairy farms. Our findings suggested that BLV infection significantly increased milk yield. Furthermore, the BoLA-DRB3 allele alone, and the combined effect of BLV infection and the BoLA-DRB3 allele had no effect. These results indicate that on-farm breeding and selection of resistant cattle, or the preferential elimination of susceptible cattle, does not affect dairy cattle productivity. Additionally, BLV infection is more likely to affect dairy cattle productivity than BoLA-DRB3 polymorphism.

BOLA Gene Polymorphism and Determination of Disease Severity and Susceptibility: Review

Major histocompatibility complex is the gene complex that exists in all vertebrae like humans (HLA), dogs (DLA), bovine (BLA), ovine (OLA), swine (SLA), and equine (ELA). MHC possesses three regions (I, II, III) that control major specific immune responses and contain variety of genes which influence growth, development, reproduction, odor, and olfaction. Measuring levels of polymorphism of these genes can provide indirect measures of the immunological fitness of populations. As humans and mice, cattle have three MHC gene classes (class I, II, and III). Both BoLA class I and II play a role in antigen presentation, however the function of BoLA class III has been releated with components of the complement system. BoLA class II region is extra separated into class IIa and class IIb. In this gene, the genetic polymorphism of class II α and β genes occurs predominantly in exon 2 encoding the antigen binding site. The BoLa-A locus has 32 serologically defined alleles and minimum four further putative alleles in addition to a high frequency of null alleles. With association allele hold DRB3.2*8 (DRB3*1201) and DRB3.2*16 (DRB3*1501) genes are associated with mastitis susceptibility, and DRB3.2*22 (DRB3*1101), DRB3.2*23 (DRB3*2703) and DRB3.2*24 (DRB3*0101) with mastitis resistance. The A8 specificity and the EIAY sequence are significant markers of resistance, while the serine residue is a marker of susceptibility to dermatophlosis. According to the report 2004, BoLA-DRB3 alleles with the amino acid residues Glu, Arg, and valine (Val) at positions 74, 77, and 78, respectively, give resistance to tumor development by BLV infection. On the other hand, HaeIII CC and HaeIII BC genotypes were actually associated with resistance to FMD in contrast, the HaeIII AA genotype was associated with susceptibility to FMD. Another report on infected Egyptian buffaloes with serotype O FMDV genotype AA for FMD resistance while genotype AC for susceptibility to FMD.

Allelic polymorphisms of the BoLA-DRB3 gene and resistance to brucellosis in Kazakh cattle.

The Authors investigated polymorphism of the bovine BoLA‑DRB3 gene in connection with resistance or susceptibility to brucellosis of two Kazakh meat breeds, Auliekol and Kazakh Whiteheaded breeds, using PCR‑RFLP. In Auliekol cattle (n = 158), 22 alleles were detected in the brucellosis group, and 24 alleles were shown in the healthy group. BoLA-DRB3 alleles *3, *4, *19, *21 were more common in healthy animals, while Brucella‑positive cattle were more frequently carriers of alleles *7, *10, *18. In Kazakh Whiteheaded cattle (n = 146), 21 alleles were detected in infected and 23 alleles in healthy cattle. Alleles *3, *8, *21 significantly predominate in healthy cattle, while alleles *7, *11, *16 are typical for animals with brucellosis. This study identified BoLA-DRB3 alleles associated with genetic resistance (*3 and *21) and susceptibility (*7) to brucellosis; remarkably, resistance alleles are shared by two important meat breeds of Kazakhstan.

Phenotypic Selection of Dairy Cattle Infected with Bovine Leukemia Virus Demonstrates Immunogenetic Resilience through NGS-Based Genotyping of BoLA MHC Class II Genes.

Characterization of the bovine leukocyte antigen (BoLA) DRB3 gene has shown that specific alleles associate with susceptibility or resilience to the progression of bovine leukemia virus (BLV), measured by proviral load (PVL). Through surveillance of multi-farm BLV eradication field trials, we observed differential phenotypes within seropositive cows that persist from months to years. We sought to develop a multiplex next-generation sequencing workflow (NGS-SBT) capable of genotyping 384 samples per run to assess the relationship between BLV phenotype and two BoLA genes. We utilized longitudinal results from milk ELISA screening and subsequent blood collections on seropositive cows for PVL determination using a novel BLV proviral load multiplex qPCR assay to phenotype the cows. Repeated diagnostic observations defined two distinct phenotypes in our study population, ELISA-positive cows that do not harbor detectable levels of provirus and those who do have persistent proviral loads. In total, 565 cows from nine Midwest dairy farms were selected for NGS-SBT, with 558 cows: 168 BLV susceptible (ELISA-positive/PVL-positive) and 390 BLV resilient (ELISA-positive/PVL-negative) successfully genotyped. Three BoLA-DRB3 alleles, including one novel allele, were shown to associate with disease resilience, *009:02, *044:01, and *048:02 were found at rates of 97.5%, 86.5%, and 90.3%, respectively, within the phenotypically resilient population. Alternatively, DRB3*015:01 and *027:03, both known to associate with disease progression, were found at rates of 81.1% and 92.3%, respectively, within the susceptible population. This study helps solidify the immunogenetic relationship between BoLA-DRB3 alleles and BLV infection status of these two phenotypic groupings of US dairy cattle.

Association between BoLA-DRB3 polymorphism and bovine leukemia virus proviral load in Vietnamese Holstein Friesian cattle.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis. Polymorphism in bovine leukocyte antigen (BoLA)-DRB3 allele can influence the host immune response to pathogens, including BLV. However, association between specific BoLA-DRB3 alleles and BLV proviral load (PVL), which is a useful index for estimating disease progression and transmission risk, in Vietnamese cattle are unknown. Here, association study of BoLA-DRB3 allele frequency between cattle with high or low PVL demonstrated BoLA-DRB3*12:01 associates with high PVL in Vietnamese Holstein Friesian (HF) crossbred cattle. This is the first study to demonstrate that BoLA-DRB3 polymorphism confers susceptibility to BLV high PVL in HF crossbred kept in Vietnam. Our results may be useful in disease control and eradiation for BLV through genetic selection.

Kinetic Study of BLV Infectivity in BLV Susceptible and Resistant Cattle in Japan from 2017 to 2019.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis. Polymorphism in bovine lymphocyte antigen (BoLA)-DRB3 alleles is related to susceptibility to BLV proviral load (PVL), which is a useful index for estimating disease progression and transmission risk. However, whether differential BoLA-DRB3 affects BLV infectivity remains unknown. In a three-year follow-up investigation using a luminescence syncytium induction assay for evaluating BLV infectivity, we visualized and evaluated the kinetics of BLV infectivity in cattle with susceptible, resistant and neutral BoLA-DRB3 alleles which were selected from 179 cattle. Susceptible cattle showed stronger BLV infectivity than both resistant and neutral cattle. The order of intensity of BLV infectivity was as follows: susceptible cattle > neutral cattle > resistant cattle. BLV infectivity showed strong positive correlation with PVL at each testing point. BLV-infected susceptible cattle were found to be at higher risk of horizontal transmission, as they had strong infectivity and high PVL, whereas BLV-infected resistant cattle were low risk of BLV transmission owing to weak BLV infection and low PVL. Thus, this is the first study to demonstrate that the BoLA-DRB3 polymorphism is associated with BLV infection.

Genetic Variation and Population Differentiation in the Bovine Lymphocyte Antigen DRB3.2 Locus of South African Nguni Crossbred Cattle.

Simple SummaryIndigenous cattle breeds are important to their native environment as they confer significant and often unique adaptability traits. The Nguni is one such breeds that is indigenous to the Southern African region. This breed confers resistance to thermal stress and diseases, amongst other factors. The bovine major histocompatibility complex is an important region, which codes for alleles that have been associated with a plethora of diseases. In the current study, the genetic diversity within this region was assessed in Nguni crossbred cattle. This was done to detect the gene pool of the Nguni breed, and to identify genes that might be important within this breed. The populations displayed a high degree of genetic diversity, and some alleles were common throughout the populations and accounted for a significant portion of the total alleles. This high genetic diversity could account for the great adaptability of the Nguni breed to Southern Africa. The bovine lymphocyte antigen (BoLA-DRB3) gene is an important region that codes for glycoproteins responsible for the initiation of an immune response. BoLA-DRB3 alleles have been demonstrated to be associated with disease resistance/tolerance. Therefore, great genetic diversity is correlated with better adaptation, fitness, and robustness. The current study was conducted to assess the population genetic structure of the BoLA-DRB3 gene in Nguni crossbred cattle using polymerase chain reaction-sequence based typing (PCR-SBT). High genetic diversity was detected, with 30 alleles, 11 of which are novel to the study. Alleles DRB3*0201, DRB3*0701, DRB*0901, and DRB*1601 were present in all populations and accounted for nearly around 50% of all observed alleles. A mean genetic diversity (HE) of 0.93 was detected. The high overall genetic diversity is possibly associated with pathogen-assisted selection and heterozygote advantage. Such high diversity might explain the hardiness of the Nguni crossbred cattle to the Southern African region. Low population genetic structure was identified (FST = 0.01), suggesting possible gene flow between populations and retention of similar alleles. The study was undertaken to bridge the dearth of such studies in South African breeds and it is imperative for effective sustainability of indigenous breeds and the implementation of effective breeding strategies.

Risk Assessment of Bovine Major Histocompatibility Complex Class II DRB3 Alleles for Perinatal Transmission of Bovine Leukemia Virus.

Perinatal transmission plays a critical role in the spread of bovine leukemia virus (BLV) infection in cattle herds. In the Holstein breed, we previously identified BLV resistant and susceptible bovine leukocyte antigen (BoLA)-DRB3 alleles, including BoLA-DRB3*009:02 and *014:01:01 with a low BLV proviral load (PVL), and *015:01 and *012:01 with a high PVL. Here, we evaluated the perinatal BLV transmission risk in dams with different BoLA-DRB3 alleles. BoLA-DRB3 alleles of 120 dam-calf pairs from five dairy farms in Japan were identified; their PVL was quantified using the BLV-Coordination of Common Motifs (CoCoMo)-qPCR-2 assay. Ninety-six dams were BLV-positive, and 29 gave birth to BLV-infected calves. Perinatal transmission frequency was 19% in dams with resistant alleles suppressed to a low PVL level, and 38% and 25% in dams with susceptible and neutral alleles that maintained high PVL levels, respectively. Notably, all calves with resistant alleles were BLV free, whereas 30% of calves with susceptible genes were infected. Thus, vertical transmission risk was extremely lower for dams and calves with resistant alleles compared to those with susceptible alleles. Our results can inform the development of effective BLV eradication programs under field conditions by providing necessary data to allow for optimal selection of dams for breeding.

Relationship between Allelic Heterozygosity in BoLA-DRB3 and Proviral Loads in Bovine Leukemia Virus-Infected Cattle.

Simple SummaryBovine leukemia virus (BLV) caused a severe cattle neoplastic disease called enzootic bovine leukosis (EBL). EBL causes significant economic losses in farming by reducing milk production, reproductive performance, and fertility, and through cattle culling or death. The BLV proviral load (PVL) represents the quantity of BLV genome that has integrated into the host’s genome in BLV-infected cells. It has been reported that PVLs differ according to the genetic background of the host, and some studies of BLV-associated host factors have reported on polymorphisms within the bovine major histocompatibility complex (MHC), namely bovine MHC is bovine leukocyte antigen (BoLA-DRB3). However, there is a great diversity in the PVLs associated with carrying various combinations of these alleles, especially for heterozygous alleles. Therefore, this research investigated whether heterogeneity in BoLA-DRB3 allele combinations would affect PVLs during BLV infections in different ages and breeds of cattle in Japan. This is the first report where the association between heterozygous allelic combinations and BLV PVLs phenotypes (HPLs, LPLs) was analyzed. Our findings augment current understanding about the immunological role played by BoLA heterozygosity in BLV-associated PVLs and biocontrol in BLV infections.Enzootic bovine leukosis is a lethal neoplastic disease caused by bovine leukemia virus (BLV), belongs to family Retroviridae. The BLV proviral load (PVL) represents the quantity of BLV genome that has integrated into the host’s genome in BLV-infected cells. Bovine leukocyte antigen (BoLA) class II allelic polymorphisms are associated with PVLs in BLV-infected cattle. We sought to identify relationships between BoLA-DRB3 allelic heterozygosity and BLV PVLs among different cattle breeds. Blood samples from 598 BLV-infected cattle were quantified to determine their PVLs by real-time polymerase chain reaction. The results were confirmed by a BLV-enzyme-linked immunosorbent assay. Restriction fragment length polymorphism-polymerase chain reaction identified 22 BoLA-DRB3 alleles. Multivariate negative binomial regression modeling was used to test for associations between BLV PVLs and BoLA-DRB3 alleles. BoLA-DRB3.2*3, *7, *8, *11, *22, *24, and *28 alleles were significantly associated with low PVLs. BoLA-DRB3.2*10 was significantly associated with high PVLs. Some heterozygous allele combinations were associated with low PVLs (*3/*28, *7/*8, *8/*11, *10/*11, and *11/*16); others were associated with high PVLs (*1/*41, *10/*16, *10/*41, *16/*27, and *22/*27). Interestingly, the BoLA-DRB3.2*11 heterozygous allele was always strongly and independently associated with low PVLs. This is the first reported evidence of an association between heterozygous allelic combinations and BLV PVLs.

BoLA-DRB3 genetic diversity in Highland Creole cattle from Bolivia.

The genetic diversity of the BoLA-DRB3 gene has been reported in different cattle breeds owing to its central role in the immune response. However, it is still unknown in hundreds of cattle breeds, especially native populations. Here, we studied BoLA-DRB3 genetic diversity in Highland Creole cattle (CrAl) from Western Bolivia, raised at altitudes between 3800 and 4200 m. DNAs from 48 CrAl cattle were genotyped for BoLA-DRB3 exon 2 alleles using polymerase chain reaction-sequence-based typing (PCR-SBT). The results were compared with 1341 previously reported data from Tropical Creole cattle and other breeds raised in the region. Twenty-three BoLA-DRB3 alleles were identified in CrAl, including the BoLA-DRB3*029:02 variant previously detected in other Creole cattle. Observed and expected heterozygosity were 0.87 and 0.93, respectively. Nucleotide diversity and the number of pairwise difference values were 0.078 and 19.46, respectively. The average number of nonsynonymous and synonymous substitutions were 0.037 and 0.097 for the entire BoLA-DRB3 exon 2, and 0.129 and 0.388 for the antigen-binding site, respectively. Venn analysis and the review of the IPD-MHC database and the literature showed that 2 of 64 alleles were only detected in CrAl, including BoLA-DRB3*029:01 previously reported in African cattle and *048:01 detected in Philippine cattle. Two additional alleles, BoLA-DRB3*007:02 and *029:02, were only present in CrAl and Lowland Creole cattle. Principal Component Analysis (PCA) showed that Bolivian Creole cattle breeds were closely located but they were distant from the Colombian Hartón del Valle Creole. FST analysis showed a low degree of genetic differentiation between Highland and Lowland Bolivian Creole cattle (FST = 0.015). The present results contribute to increasing our knowledge of BoLA-DRB3 genetic diversity in cattle breeds.

Characterization of bovine MHC DRB3 diversity in global cattle breeds, with a focus on cattle in Myanmar

BackgroundMyanmar cattle populations predominantly consist of native cattle breeds (Pyer Sein and Shwe), characterized by their geographical location and coat color, and the Holstein-Friesian crossbreed, which is highly adapted to the harsh tropical climates of this region. Here, we analyzed the diversity and genetic structure of the BoLA-DRB3 gene, a genetic locus that has been linked to the immune response, in Myanmar cattle populations.MethodsBlood samples (n = 294) were taken from two native breeds (Pyer Sein, n = 163 and Shwe Ni, n = 69) and a cattle crossbreed (Holstein-Friesian, n = 62) distributed across six regions of Myanmar (Bago, n = 38; Sagaing, n = 77; Mandalay, n = 46; Magway, n = 46; Kayin, n = 43; Yangon, n = 44). In addition, a database that included 2428 BoLA-DRB3 genotypes from European (Angus, Hereford, Holstein, Shorthorn, Overo Negro, Overo Colorado, and Jersey), Zebuine (Nellore, Brahman and Gir), Asian Native from Japan and Philippine and Latin-American Creole breeds was also included. Furthermore, the information from the IPD–MHC database was also used in the present analysis. DNA was genotyped using the sequence-based typing method. DNA electropherograms were analyzed using the Assign 400ATF software.ResultsWe detected 71 distinct alleles, including three new variants for the BoLA-DRB3 gene. Venn analysis showed that 11 of these alleles were only detected in Myanmar native breeds and 26 were only shared with Asian native and/or Zebu groups. The number of alleles ranged from 33 in Holstein-Friesians to 58 in Pyer Seins, and the observed versus unbiased expected heterozygosity were higher than 0.84 in all the three the populations analyzed. The FST analysis showed a low level of genetic differentiation between the two Myanmar native breeds (FST = 0.003), and between these native breeds and the Holstein-Friesians (FST < 0.021). The average FST value for all the Myanmar Holstein-Friesian crossbred and Myanmar native populations was 0.0136 and 0.0121, respectively. Principal component analysis (PCA) and tree analysis showed that Myanmar native populations grouped in a narrow cluster that diverged clearly from the Holstein-Friesian populations. Furthermore, the BoLA-DRB3 allele frequencies suggested that while some Myanmar native populations from Bago, Mandalay and Yangon regions were more closely related to Zebu breeds (Gir and Brahman), populations from Kayin, Magway and Sagaing regions were more related to the Philippines native breeds. On the contrary, PCA showed that the Holstein-Friesian populations demonstrated a high degree of dispersion, which is likely the result of the different degrees of native admixture in these populations.ConclusionThis study is the first to report the genetic diversity of the BoLA-DRB3 gene in two native breeds and one exotic cattle crossbreed from Myanmar. The results obtained contribute to our understanding of the genetic diversity and distribution of BoLA-DRB3 gene alleles in Myanmar, and increases our knowledge of the worldwide variability of cattle BoLA-DRB3 genes, an important locus for immune response and protection against pathogens.

BoLA-DRB3 Polymorphism is Associated with Differential Susceptibility to Bovine Leukemia Virus-Induced Lymphoma and Proviral Load.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leucosis. However, less than 5% of BLV-infected cattle will develop lymphoma, suggesting that, in addition to viral infection, host genetic polymorphisms might play a role in disease susceptibility. Bovine leukocyte antigen (BoLA)-DRB3 is a highly polymorphic gene associated with BLV proviral load (PVL) susceptibility. Due to the fact that PVL is positively associated with disease progression, it is believed that controlling PVL can prevent lymphoma development. Thus, many studies have focused on the relationship between PVL and BoLA-DRB3. Despite this, there is little information regarding the relationship between lymphoma and BoLA-DRB3. Furthermore, whether or not PVL-associated BoLA-DRB3 is linked to lymphoma-associated BoLA-DRB3 has not been clarified. Here, we investigated whether or not lymphoma-associated BoLA-DRB3 is correlated with PVL-associated BoLA-DRB3. We demonstrate that two BoLA-DRB3 alleles were specifically associated with lymphoma resistance (*010:01 and *011:01), but no lymphoma-specific susceptibility alleles were found; furthermore, two other alleles, *002:01 and *012:01, were associated with PVL resistance and susceptibility, respectively. In contrast, lymphoma and PVL shared two resistance-associated (DRB3*014:01:01 and *009:02) BoLA-DRB3 alleles. Interestingly, we found that PVL associated alleles, but not lymphoma associated alleles, are related with the anti-BLV gp51 antibody production level in cows. Overall, our study is the first to demonstrate that the BoLA-DRB3 polymorphism confers differential susceptibility to BLV-induced lymphoma and PVL.

Early events following bovine leukaemia virus infection in calves with different alleles of the major histocompatibility complex \xa0DRB3 gene

Cattle maintaining a low proviral load (LPL) status after bovine leukaemia virus (BLV) infection have been recognized as BLV controllers and non-transmitters to uninfected cattle in experimental and natural conditions. LPL has been associated with host genetics, mainly with the BoLA class II DRB3 gene. The aim of this work was to study the kinetics of BLV and the host response in Holstein calves carrying different BoLA-DRB3 alleles. Twenty BLV-free calves were inoculated with infected lymphocytes. Two calves were maintained uninfected as controls. Proviral load, total leukocyte and lymphocyte counts, anti-BLVgp51 titres and BLVp24 expression levels were determined in blood samples at various times post-inoculation. The viral load peaked at 30 days post-inoculation (dpi) in all animals. The viral load decreased steadily from seroconversion (38 dpi) to the end of the study (178 dpi) in calves carrying a resistance-associated allele (*0902), while it was maintained at elevated levels in calves with *1501 or neutral alleles after seroconversion. Leukocyte and lymphocyte counts and BLVp24 expression did not significantly differ between genetic groups. Animals with < 20 proviral copies/30 ng of DNA at 178 dpi or < 200 proviral copies at 88 dpi were classified as LPL, while calves with levels above these limits were considered to have high proviral load (HPL) profiles. All six calves with the *1501 allele progressed to HPL, while LPL was attained by 6/7 (86%) and 2/6 (33%) of the calves with the *0902 and neutral alleles, respectively. One calf with both *0902 and *1501 developed LPL. This is the first report of experimental induction of the LPL profile in cattle.

Bovine leukemia virus proviral load is more strongly associated with bovine major histocompatibility complex class II DRB3 polymorphism than with DQA1 polymorphism in Holstein cow in Japan

Bovine leukemia virus (BLV) causes enzootic bovine leukosis and is closely related to the human T-lymphotropic virus. Bovine major histocompatibility complex (BoLAs) are used extensively as markers of disease and immunological traits in cattle. For BLV diagnosis, proviral load is a major diagnosis index for the determination of disease progression and transmission risk. Therefore, we investigated the frequency of BoLA-DRB3 alleles, BoLA-DQA1 alleles, and haplotypes of BoLA class II isolated from the heads of 910 BLV-infected cows out of 1290 cows assessed from BLV-positive farms, in a nationwide survey from 2011 to 2014 in Japan. Our aim was to identify BoLA class II polymorphisms associated with the BLV proviral load in the Holstein cow. The study examined 569 cows with a high proviral load and 341 cows with a low proviral load. Using the highest odds ratio (OR) as a comparison index, we confirmed that BoLA-DRB3 was the best marker for determining which cow spread the BLV (OR 13.9 for BoLA-DRB3, OR 11.5 for BoLA-DQA1, and OR 6.2 for BoLA class II haplotype). In addition, DRB3*002:01, *009:02, *012:01, *014:01, and *015:01 were determined as BLV provirus associated alleles. BoLA-DRB3*002:01, *009:02, and *014:01 were determined as resistant alleles (OR > 1), and BoLA-DRB3*012:01 and *015:01 were determined as susceptible alleles (OR < 1). In this study, we showed that BoLA-DRB3 was a good marker for determining which cow spread BLV, and we found not only one resistant allele (BoLA-DRB3*009:02), but also two other disease-resistant alleles and two disease-susceptible alleles. This designation of major alleles as markers of susceptibility or resistance can allow the determination of the susceptibility or resistance of most cows to disease. Overall, the results of this study may be useful in eliminating BLV from farms without having to separate cows into several cowsheds.

Population genetic structure of the Ukrainian black-pied dairy breed with the genome BoLA-DRB3

The second exon of the BoLA-DRB3 gene has the highest level of polymorphism among all studied loci of the major histocompatibility complex (MHC) in cattle, which allows it to be used for studying population-genetic structure and assessing the level of biodiversity of populations or comparing the biodiversity of particular herds. According to the results of typing the blood samples of 293 cows using the method of PCR-RLFP, we determined allele frequencies of the BoLA-DRB3 gene for the Ukrainian black-pied dairy breed. The study was conducted on three herds in Khmelnytskyi Oblast: LLC “Kozatska Dolyna 2006” (herd A, n = 122), agrofirm “Perlyna Podillya” (herd B, n = 82) and branch “Ridnyy kray” (herd C, n = 89). In total, 37 alleles were found: herd A 31, herd B 25 and herd C 28. In total, in the three subpopulations seven alleles were found with frequency of over 5%, the total share of which equaled 55.8%. The most widely distributed allele was BoLA-DRB3.2*24, which composed 22.2% of the allele pool of the breed. We determined a high level of observed (0.89 to 0.95) and expected (0.93 to 0.94) heterozygosity. In herds A and B, there was determined domination of homozygotes. Deviation from HWE, calculated using the value of Wright`s individual fixation index, equaled FIS(A) = 0.016 (χ2 = 0.03; P &gt; 0.05) and FIS(B) = 0.044 (χ2 = 0.076; P &gt; 0.05). In herd C, we found excess of heterozygotes FIS(C) = -0.017 (χ2 = 0.026; P &gt; 0.05). Rather low values were determined for the subpopulation fixation index: FST(A) = 0.009 (χ2 = 65.9; P &lt; 0.01), FST(B) = 0.012 (χ2 = 47.2; P &lt; 0.05) and FST(C) = 0.003 (χ2 = 14.4; P &gt; 0.05), which were significantly different from the mean value for cattle (FST = 0.078), indicating insignificant reduction of heterozygosity and divergence between the subpopulations by the BoLA-DRB3 gene. To assess genetic diversity, we calculated parameters of effective allele number (Ae) and Shannon’s information index (I). In spite of the different numbers of alleles found in the selections, it was suggested that for assessing their diversity, an efficiency index will be used which shows the share of effective alleles among all alleles found in a subpopulation (Ae/Nа). The calculated values of the parameters equaled: herd A Ae = 14.9, Ae/Nа = 0.48, I = 3.05; herd B Ae = 14.5, Ae/Nа = 0.58, I = 2.87; herd C Ae = 16.4, Ae/Nа = 0.59, I = 3.01. Frequencies of BoLA-DRB3 alleles were used for calculating genetic similarity and standard genetic distances according to Nei. Cows of herds B and C were found to be more genetically affinitive by the BoLA-DRB3 gene. Standard genetic distance between them was the lowest D = 0.13, which coincides with the geographic locations and historical development of these populations. The results of the study prove that the studied herds have a high level of polymorphism. Frequency characteristics, values of expected heterozygosity, effective allele number, efficiency index and Shannon’s information index compared to the similar parameters for Holstein, black-pied and some other local breeds of cattle indicate the high genetic diversity of the studied subpopulations of the Ukrainian black-pied dairy breed.

The influence of BoLA-DRB3 alleles on incidence of clinical mastitis, cystic ovary disease and milk traits in Holstein Friesian cattle.

The major histocompatibility complex in cattle (BoLA) is regulated by genes that are closely related to the development of the immunological response to pathogens. The most polymorphic BoLA-DRB3.2 locus was analysed in 209 black-and-white Holstein-Friesian cows in Poland in order to a better explanation of influence of MHC on immunity to diseases in dairy cattle. A total of 23 alleles were identified, among which the *24, *16 and *22 alleles were observed with the highest frequency. These alleles were analysed in terms of their association with the occurrence of mastitis, ovarian cysts, retained placenta and uterine abscesses as well as their contribution to production traits (milk yield, protein and fat percentage in milk). It was determined that the BoLA-DRB3.2 *22 and *16 alleles were associated with a lower risk of clinical mastitis; however, a statistical significance was observed only for the *22 allele. Clinical mastitis was observed at a frequency lower by 8% in cows with one copy of the *22 allele compared to cows with 0 copies of the allele. The presence of the *22 allele in the genotype was also associated with higher milk yield, although this association was not statistically significant.