Impact of bovine leukemia virus infection on beef cow longevity.

Bovine leukosis is a chronic lymphoproliferative disorder caused by bovine leukemia virus (BLV). Previous studies estimate that 38 % of cow-calf beef herds and 10.3 % of individual beef cows in the US are BLV seropositive. About 70 % of BLV infected animals are asymptomatic carriers of the virus, while less than 5% develop lymphosarcoma, the leading reason for carcass condemnation at the US slaughterhouses. Studies provide evidence that BLV infection leads to decreased immune function making animals more vulnerable to other diseases, which could shorten their productive lifespan and increase economic losses in the cattle industry. BLV seropositive dairy cows are reportedly more likely to be culled sooner compared with their uninfected herd mates. Beyond simple prevalence studies, little is known about the impact of BLV infection in beef cattle production or specifically on beef cow longevity. Our objective was to determine the association between BLV infection and cow longevity in beef cow-calf operations. Twenty-seven cow-calf herds from the Upper Midwest volunteered to participate in this study. Female beef cattle (n = 3146) were tested for serum BLV antibodies by ELISA. A subsample of 648 cows were also tested for BLV proviral load (PVL). Culling data was collected for the subsequent 24 months. Twenty-one herds (77.7 %) had at least one BLV-infected animal, and 29.2 % (930/3146) of tested animals were BLV seropositive. Of the BLV-positive cows, 33.7 % (318/943) were culled compared with 32.1 % (541/1682) of the seronegative cows. BLV status did not affect cows' longevity within herds (P = 0.062). However, cows with high BLV PVL had decreased survival within the herd compared with ELISA- negative cows (P = 0.01). Overall, infection with BLV did not impact beef cow longevity unless the disease had progressed to a point of high BLV PVL.

Early events following bovine leukaemia virus infection in calves with different alleles of the major histocompatibility complex DRB3 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.

Lifetime effects of infection with bovine leukemia virus on longevity and milk production of dairy cows.

Enzootic bovine leukosis (EBL) is an economically important disease of dairy cattle caused by bovine leukemia virus (BLV). The economic impacts of the infection have been debated in the literature. The present study was conducted to determine the lifetime effects of BLV infection on longevity and milk production of dairy cows in Canada. The data were aggregated from a combination of two data sets: 1) BLV serum-ELISA test results from Canada-wide surveys of production limiting diseases, which took place between 1998 and 2003 in 8 provinces, and 2) longitudinal production data for all cows in the former study, extracted from the Canadian dairy herd improvement database. All participant cows had been culled or died by the onset of this study. A historical cohort study was designed, including cows which tested positive to BLV-antibodies in their first lactation (positive cohort, n=1858) and cows which tested negative in their second or later lactations (negative cohort, n=2194). To assess the impacts of infection with BLV on longevity (the number of lifetime lactations), a discrete-time survival analysis was carried out. The effect of BLV on the lifetime milk production (the sum of all life 305-day milk production) was evaluated using a multilevel linear regression model. Overall, 4052 cows from 348 herds met the eligibility criteria and were enrolled in the study. In the longevity model, the interaction term between time (lactation number) and BLV-status was highly significant. Cows which were positive to BLV had consistently greater probabilities of being culled (or dying) than the test-negative cows. In the milk production model, the interaction term between BLV-status and longevity of the cows was highly significant; indicating that lifetime BLV effects on the total milk production was dependent on the lactation in which the study cows were culled/died. Infected cows with 2 and 3 lactations showed significantly lower life milk productions [-2554kg (-3609 to -1500) and -1171kg (-2051 to -292), respectively] compared with their negative counterparts with 2 and 3 lactations. As the cows lived longer (>3 lactations), the differences in life milk production between the two cohorts were no longer significant. Overall, it was predicted that the test-positive cows produced substantially lower milk compared to the test-negative cows throughout their study lifespans. With the high prevalence of BLV in Canadian dairy cows and its detrimental economic impacts, pursuing broad-based control programs in Canada should be evaluated.

Dynamics of perinatal bovine leukemia virus infection.

BACKGROUND: Bovine leukemia virus (BLV) is highly endemic in many countries, including Argentina. As prevention of the spread from infected animals is of primary importance in breaking the cycle of BLV transmission, it is important to know the pathophysiology of BLV infection in young animals, as they are the main source of animal movement. In this work, we determined the proviral load and antibody titers of infected newborn calves from birth to first parturition (36 months). RESULTS: All calves under study were born to infected dams with high proviral load (PVL) in blood and high antibody titers and detectable provirus in the colostrum. The PVL for five out of seven calves was low at birth. All animals reached PVLs of more than 1% infected peripheral blood mononuclear cells (PBMCs), three at 3 months, one at 6 months, and one at 12 months. High PVLs persisted until the end of the study, and, in two animals, exceeded one BLV copy per cell. Two other calves maintained a high PVL from birth until the end of the study. Antibody titers were 32 or higher in the first sample from six out of seven calves. These decayed at 3-6 months to 16 or lower, and then increased again after this point. CONCLUSIONS: Calves infected during the first week of life could play an active role in early propagation of BLV to susceptible animals, since their PVL raised up during the first 12 months and persist as high for years. Early elimination could help to prevent transmission to young susceptible animals and to their own offspring. To our knowledge, this is the first study of the kinetics of BLV proviral load and antibody titers in newborn infected calves.

Association between bovine leukemia virus, production, and population age in Michigan dairy herds.

The objective of this study was to determine the herd-level effect of bovine leukemia virus (BLV) infection on dairy production, culling, and cow longevity. During routine herd testing, Dairy Herd Improvement Association technicians collected milk samples from about 40 cows from each of 104 randomly selected Michigan dairy herds averaging ≥120 milking cows and 11,686 kg of milk/yr. Milk samples were analyzed for the presence of anti-BLV antibodies by ELISA, and herd- and lactation-specific estimates of BLV prevalence were computed to determine which were the most predictive of herd milk production, culling rate, and cow longevity (proportion of cows in their third or greater lactation). On this basis, the herd BLV index (an unweighted mean BLV prevalence rate for lactation number 1, 2, 3, and ≥4) was selected as the measure of BLV prevalence that was the most highly associated with BLV economic impact. Step-down multivariate analysis was used to determine the extent to which any of 19 herd-level management variables may have confounded the association of BLV index and measures of herd economic impact (milk production and cow longevity). The BLV index was not associated with the 12-mo culling rate, but was negatively associated in the final multivariable model with the proportion of cows that were ≥third lactation, and was negatively associated with herd milk production. In summary, increased prevalence of BLV within Michigan dairy herds was found to be associated with decreased herd milk production and decreased cow longevity. Our results provide evidence that BLV infection is associated with herd-level economic impacts in high-performing dairy herds.

Associations between reproductive performance and seropositivity for bovine leukemia virus, bovine viral-diarrhea virus, Mycobacterium avium subspecies paratuberculosis, and Neospora caninum in Canadian dairy cows.

Our objective was to investigate effects of seropositivity for bovine leukemia virus (BLV), Type 1 bovine viral-diarrhea virus (BVDV), Mycobacterium avium subspecies paratuberculosis (MAP), and Neospora caninum (NC), and their possible interactions, on reproductive efficiency (specifically, first-service conception [FSC], and calving interval [CI]) in dairy cows. The sample population included up to 30 randomly selected animals from 179 randomly selected farms in five provinces in Canada, from which 23 farms did not meet the inclusion criteria for the final dataset. Serum samples were tested for antibodies against the stated pathogens using commercially available diagnostic tests. A Cox proportional hazards model with shared (herd-level) frailty was utilized to analyze the CI data. In this model, BLV-seropositive cows had a 7% lower rate of conception compared to seronegative cows (P=0.06). Mixed logistic regression models of CI>484 days, CI>534 days, and CI>584 days were built to explore factors of long CIs. These cut-offs were selected to represent calving-to-conception intervals of >200 days, >250 days, and >300 days. BLV-seropositive cows had higher odds of having a CI>484 days compared to BLV-seronegative cows, and BLV serostatus interacted with lactation number in this model, with 1st lactation seropositive cows being more likely to have a CI>484 days than older seropositive cows. NC-seropositive cows had a 1.27 times higher odds of exhibiting a CI>484 days, a 1.37 times higher odds of a CI>534 days, and a 1.54 times higher odds of a CI>584 days, compared to NC-seronegative cows. Neither BVDV nor MAP seropositivity showed any significant effect in these models. For the FSC models, a first service was classified successful (pregnancy=1) if it was the cow's last service and she calved 270-290 days later. A mixed logistic regression model of FSC revealed an interaction between NC and BVDV-seropositivity at the herd level, with odds ratios of 0.64, 1.06 and 0.85 for NC-seropositive cows (compared to NC-seronegative cows) in BVDV-seronegative, BVDV-seropositive and BVDV-missing herds, respectively. BLV and MAP seropositivity had no significant impact on FSC. All models controlled for herd-clustering effects, and included parity, linear score of somatic cell counts, peak milk, and province to control for confounding. The overall FSC was 51%, the average CI was 393 days, and 18%, 9% and 5% of lactations had CI>484 days, >534 days, and >584 days, respectively.

Host soluble factors that regulate the synthesis of the major core protein of the bovine leukemia virus (BLV) in a naturally infected neoplastic B-cell line.

Bovine leukemia virus (BLV) is a B-cell tropic Deltaretrovirus that induces a lifelong infection and causes a fatal lymphosarcoma in less than 10% of the infected cattle. BLV is usually present in its host in a transcriptional repressed state but becomes de-repressed a few hours after the infected lymphocytes are cultured in vitro. In the present study we have examined the effect of soluble host factors and various substances on the synthesis of the major BLV protein (p24) in a permanent culture (cell line NBC-10) of neoplastic B-lymphocytes derived from BLV-infected cattle. Certain batches of fetal calf serum (FCS) and bovine platelet lysates (PLy) induced a rapid and drastic increase of the synthesis of BLVp24 in the NBC-10 cells. Neutralization experiments with specific antibodies demonstrated that the transforming growth factor-beta (TGF-beta) was responsible for the stimulatory activity of FCS and PLy on the synthesis of BLVp24 in the NBC-10 cells. Recombinant TGF-beta also stimulated the synthesis of BLVp24 in cultures of peripheral blood mononuclear cells (PBMCs) obtained from BLV-infected cattle. Mitogens, phorbol-myristate-acetate and prostaglandin E(2), previously shown to stimulate the expression of BLV in cultures of PBMC, did not induce the synthesis of BLVp24 in cultures of NBC-10 cells. Plasma, serum and milk from BLV-negative cattle inhibited the synthesis of BLVp24 induced by FCS, PLy or TGF-beta in the NBC-10 cells. The blocking activity was found in the whey and the beta-casein fractions of bovine milk. The relevance of these findings with regard to the previously reported plasma factor (PBB) with blocking activity on the expression of BLV in short-term PBMC cultures is discussed. Based on the information obtained in the present study we have standardized a reproducible and rapid assay system for the identification of factors that regulate the synthesis of BLVp24 in naturally infected neoplastic B cells.

Mechanisms of leukemogenesis induced by bovine leukemia virus: prospects for novel anti-retroviral therapies in human.

In 1871, the observation of yellowish nodules in the enlarged spleen of a cow was considered to be the first reported case of bovine leukemia. The etiological agent of this lymphoproliferative disease, bovine leukemia virus (BLV), belongs to the deltaretrovirus genus which also includes the related human T-lymphotropic virus type 1 (HTLV-1). This review summarizes current knowledge of this viral system, which is important as a model for leukemogenesis. Recently, the BLV model has also cast light onto novel prospects for therapies of HTLV induced diseases, for which no satisfactory treatment exists so far.

Production effects of pathogens causing bovine leukosis, bovine viral diarrhea, paratuberculosis, and neosporosis.

The primary purpose of this research was to determine associations among seropositivity for bovine leukemia virus (BLV), bovine viral diarrhea virus (BVDV), Mycobacterium avium ssp. paratuberculosis (MAP), and Neospora caninum (NC) and each of 3 outcome variables (305-d milk, fat, and protein production) in Canadian dairy cattle. Serum samples from up to 30 randomly selected cows from 342 herds on monthly milk testing were tested for antibodies against BLV (IDEXX ELISA; IDEXX Corporation, Westbrook, ME), MAP (IDEXX or Biocor ELISA; Biocor Animal Health, Inc., Omaha, NE), and NC (IDEXX or Biovet ELISA; Biovet Inc., St. Hyacinthe, Quebec, Canada). Up to 5 unvaccinated cattle over 6 mo of age were tested for virus-neutralizing antibodies to the Singer strain of type 1 BVDV. Dairy Herd Improvement records were obtained electronically for all sampled cows. Linear mixed models with herd and cow as random variables were fit, with significant restricted maximum likelihood estimates of outcome effects being obtained, while controlling for potential confounding variables. Bovine leukemia virus seropositivity was not associated with 305-d milk, 305-d fat, or 305-d protein production. Cows in BVDV-seropositive herds (at least one unvaccinated animal with a titer > or =1:64) had reductions in 305-d milk, fat, and protein of 368, 10.2, and 9.5 kg, respectively, compared with cows in BVDV-seronegative herds. Mycobacterium avium ssp. paratuberculosis seropositivity was associated with lower 305-d milk of 212 kg in 4+-lactation cows compared with MAP-seronegative 4+-lactation cows. Neospora caninum seropositivity in primiparous cows was associated with lower 305-d milk, fat, and protein of 158, 5.5, and 3.3 kg, respectively, compared with NC-seronegative primiparous cows. There were no interactions among seropositivity for any of the pathogens and their effects on any of the outcomes examined, although the low MAP seroprevalence limited this analysis. Results from this research will contribute to understanding the economic impacts of these pathogens and justify their control.

Involvement of glutathione as a mechanism of indirect protection against spontaneous ex vivo apoptosis associated with bovine leukemia virus.

Viruses have developed strategies to counteract the apoptotic response of the infected host cells. Modulation of apoptosis is also thought to be a major component of viral persistence and progression to leukemia induced by retroviruses like human T-lymphotropic virus type 1 (HTLV-1) and bovine leukemia virus (BLV). Here, we analyzed the mechanism of ex vivo apoptosis occurring after isolation of peripheral blood mononuclear cells from BLV-infected sheep. We show that spontaneous apoptosis of ovine B lymphocytes requires at least in part a caspase 8-dependent pathway regardless of viral infection. Cell death is independent of cytotoxic response and does not involve the tumor necrosis factor alpha/NF-kappaB/nitric oxide synthase/cyclooxygenase pathway. In contrast, pharmaceutical depletion of reduced glutathione (namely, gamma-glutamyl-l-cysteinyl-glycine [GSH]) by using ethacrynic acid or 1-pyrrolidinecarbodithioic acid specifically reverts inhibition of spontaneous apoptosis conferred indirectly by protective BLV-conditioned media; inversely, exogenously provided membrane-permeable GSH-monoethyl ester restores cell viability in B lymphocytes of BLV-infected sheep. Most importantly, intracellular GSH levels correlate with virus-associated protection against apoptosis but not with general inhibition of cell death induced by polyclonal activators, such as phorbol esters and ionomycin. Finally, inhibition of apoptosis does not correlate with the activities of GSH peroxidase and GSH reductase. In summary, our data fit into a model in which modulation of the glutathione system is a key event involved in indirect inhibition of apoptosis associated with BLV. These observations could have decisive effects during therapeutic treatment of delta-retroviral pathogenesis.

Reduced cell turnover in bovine leukemia virus-infected, persistently lymphocytotic cattle.

Although nucleotide analogs like bromodeoxyuridine have been extensively used to estimate cell proliferation in vivo, precise dynamic parameters are scarce essentially because of the lack of adequate mathematical models. Besides recent developments on T cell dynamics, the turnover rates of B lymphocytes are largely unknown particularly in the context of a virally induced pathological disorder. Here, we aim to resolve this issue by determining the rates of cell proliferation and death during the chronic stage of the bovine leukemia virus (BLV) infection, called bovine persistent lymphocytosis (PL). Our methodology is based on direct intravenous injection of bromodeoxyuridine in association with subsequent flow cytometry. By this in vivo approach, we show that the death rate of PL B lymphocytes is significantly reduced (average death rate, 0.057 day(-1) versus 0.156 day(-1) in the asymptomatic controls). Concomitantly, proliferation of the PL cells is also significantly restricted compared to the controls (average proliferation rate, 0.0046 day(-1) versus 0.0085 day(-1)). We conclude that bovine PL is characterized by a decreased cell turnover resulting both from a reduction of cell death and an overall impairment of proliferation. The cell dynamic parameters differ from those measured in sheep, an experimental model for BLV infection. Finally, cells expressing p24 major capsid protein ex vivo were not BrdU positive, suggesting an immune selection against proliferating virus-positive lymphocytes. Based on a comparative leukemia approach, these observations might help to understand cell dynamics during other lymphoproliferative disease such as chronic lymphocytic leukemia or human T-cell lymphotropic virus-induced adult T-cell leukemia in humans.

Association between bovine-leukosis virus seroprevalence and herd-level productivity on US dairy farms.

Bovine-leukosis virus (BLV; also termed 'bovine-leukemia virus') is a retrovirus that primarily affects lymphoid tissue of dairy and beef cattle. Our objective was to investigate the association between BLV infection and annual value of production (AVP) on dairy herds within the United States, as part of the USDA National Animal Health Monitoring System's 1996 dairy study. 1006 herds (in 20 states) with at least 30 dairy cows were interviewed during 1996. The agar-gel immunodiffusion test was used to detect serum antibodies to BLV. 10-40 cows from each herd were tested and each tested cow was classified as negative or positive based on results of a single test. A multivariable regression model was used with the 976 herds with complete data for analysis. When compared to herds with no test-positive cows, herds with test-positive cows produced 218 kg per cow (i.e. 3%) less milk. The average reduction in AVP was $59 per cow for test-positive herds relative to test-negative herds. For the dairy industry as a whole, BLV seropositivity was associated with loss to producers of $285 million and $240 million for consumers. Most of this $525 million industry loss was due to reduced milk production in test-positive herds.

Translocation of the B cell receptor to lipid rafts is inhibited in B cells from BLV-infected, persistent lymphocytosis cattle.

Bovine leukemia virus (BLV) infection causes a significant polyclonal expansion of CD5(+), IgM+ B lymphocytes known as persistent lymphocytosis (PL) in approximately 30% of infected cattle. There is evidence that this expanded B cell population has altered signaling, and resistance to apoptosis has been proposed as one mechanism of B cell expansion. In human and murine B cells, antigen binding initiates movement of the B cell receptor (BCR) into membrane microdomains enriched in sphingolipids and cholesterol, termed lipid rafts. Lipid rafts include members of the Src-family kinases and exclude certain phosphatases. Inclusion of the BCR into lipid rafts plays an important role in regulation of early signaling events and subsequent antigen internalization. Viral proteins may also influence signaling events in lipid rafts. Here we demonstrate that the largely CD5(+) B cell population in PL cattle has different mobilization and internalization of the BCR when compared to the largely CD5-negative B cells in BLV-negative cattle. Unlike B cells from BLV-negative cattle, the BCR in B cells of BLV-infected, PL cattle resists movement into lipid rafts upon stimulation and is only weakly internalized. Expression of viral proteins as determined by detection of the BLV transmembrane (TM) envelope glycoprotein gp30 did not alter these events in cells from PL cattle. This exclusion of the BCR from lipid rafts may, in part, explain signaling differences seen between B cells of BLV-infected, PL, and BLV-negative cattle and the resistance to apoptosis speculated to contribute to persistent lymphocytosis.

CD5 is dissociated from the B-cell receptor in B cells from bovine leukemia virus-infected, persistently lymphocytotic cattle: consequences to B-cell receptor-mediated apoptosis.

Bovine leukemia virus (BLV), a retrovirus related to human T-cell leukemia virus types 1 and 2, can induce persistent nonneoplastic expansion of the CD5(+) B-cell population, termed persistent lymphocytosis (PL). As in human CD5(+) B cells, we report here that CD5 was physically associated with the B-cell receptor (BCR) in normal bovine CD5(+) B cells. In contrast, in CD5(+) B cells from BLV-infected PL cattle, CD5 was dissociated from the BCR. In B cells from PL cattle, apoptosis decreased when cells were stimulated with antibody to surface immunoglobulin M (sIgM), while in B cells from uninfected cattle, apoptosis increased after sIgM stimulation. The functional significance of the CD5-BCR association was suggested by experimental dissociation of the CD5-BCR interaction by cross-linking of CD5. This caused CD5(+) B cells from uninfected animals to decrease apoptosis when stimulated with anti-sIgM. In contrast, in CD5(+) B cells from PL animals, in which CD5 was already dissociated from the BCR, there was no statistically significant change in apoptosis when CD5 was cross-linked and the cells were stimulated with anti-sIgM. Disruption of CD5-BCR interactions and subsequent decreased apoptosis and increased survival in antigenically stimulated B cells may be a mechanism of BLV-induced PL.

Experimental infection of calves with bovine leukemia virus (BLV): an applicable model of a retroviral infection.

An experimental model of chronic infection with bovine leukemia virus (BLV) was established in young calves within a relatively short time. In the sera of all infected calves, precipitating antibodies were detected within 5 weeks after infection but upon disease progression pattern of cellular profiles varied. Three calves exhibited transient lymphocytosis 3-5 weeks after infection, two became persistent lymphocytotic (PL+) by that time and one stayed non-lymphocytotic (PL-) for 11 weeks and became PL+ after 4.5 months. Eventually all infected calves became PL+ by the end of the experiment, 6-12 months after infection. Increase of total counts of peripheral blood mononuclear cells (PBMC) related to polyclonal expansion of B-cells. The latter was assessed in all infected calves where the expansion of CD5-bearing cells (B+ CD5+) correlated with increase or decrease of total PBMC counts. Other cell populations such as CD4 and CD8 were also affected. Percentages decreased by 5 weeks after experimental infection to about half their original values though actual cell numbers stayed relatively stable. The experimental model we established compared well with field cases of naturally BLV-infected cattle and thus permitted the investigation of the disease at early stages of infection.

Short-termed expression of interleukin-12 during experimental BLV infection may direct disease progression to persistent lymphocytosis.

In this study an attempt was made to elucidate cellular response cytokine expression upon experimental bovine leukemia virus (BLV) infection in cattle. Progression of infection was monitored by BLV gp51 mRNA expression or DNA amplification by RT-PCR or PCR, respectively, to detect provirus infected cells. Antibodies to BLV were detected by an agar gel immuno-diffusion (AGID) test in 5 weeks and persistent lymphocytosis (PL+) was established in all four BLV-infected animals in 24 weeks after infection. At the initial stage of infection a strong cellular immune response was induced mediated by IL-12p40 mRNA expression. Short-termed IL-12p40 expression was observed in peripheral blood mononuclear cells (PBMC) in two out of four infected animals following 1-3 weeks after infection, while viral mRNA expression was observed 2 weeks following infection. Expression of genes coding for the pro-inflammatory TNFalpha, IL-1beta and cellular response cytokines IFNgamma and IL-2 was detected beginning with the second and third week after infection in all BLV-infected animals. However, IFNgamma expression significantly decreased in 12 weeks after infection in three animals while IL-10 message initially detected 3 weeks after infection increased by 12 weeks and persisted. The observed immediate short-termed cell mediated immune response characterized by IL-12p40 and IFNgamma expression followed by an early shift to an IL-10 induced humoral response, may change the cytokine balance and direct disease progression to the PL+ stage.

Interleukin-12 p40 mRNA expression in bovine leukemia virus-infected animals: increase in alymphocytosis but decrease in persistent lymphocytosis.

Interleukin-12 (IL-12), a key cytokine in immune regulation, has an important role in activating the cell-mediated immune response in infectious diseases. Recently, a dichotomy between IL-12 and IL-10 regarding progression of a variety diseases has emerged. IL-12 activates type 1 cytokine production and has an antagonistic effect on type 2 cytokines. Here, by using quantitative competitive PCR, we show that peripheral blood mononuclear cells from bovine leukemia virus-infected animals in the alymphocytotic stage of disease express an increased amount of IL-12 p40 mRNA. In contrast, IL-12 p40 mRNA expression by cells from animals with late-stage disease, termed persistent lymphocytosis, was significantly decreased compared to that by normal and alymphocytotic animals. Interestingly, IL-12 p40 mRNA was also detected in tumor-bearing animals. IL-12 p40 expression occurred only in monocytes/macrophages, not B or T lymphocytes. The present study combined with previous findings suggest that IL-12 in bovine leukemia virus-infected animals may regulate production of other cytokines such as gamma interferon and IL-10 and the progression of bovine leukosis in animals that develop more advanced disease such as a persistent lymphocytosis of B cells or B-cell lymphosarcoma.

Bovine leukemia virus-induced lymphocytosis and increased cell survival mainly involve the CD11b+ B-lymphocyte subset in sheep.

In this study, we show that bovine leukemia virus (BLV)-induced persistent lymphocytosis (PL) results from the in vivo expansion of the CD11b+ B-lymphocyte population. This subset shares phenotypic characteristics with murine and human B-1 cells. BLV interactions with the sheep B-1-like subset were explored. We found that B-1- and B-2-like cells are initially infected to similar extents. However, in long-term-infected sheep, the viral load is higher in B-1-like cells and only B-1- and not B-2-like cells show increased ex vivo survival compared to that in uninfected sheep. Ex vivo viral expression was found in both B-1- and B-2-like cells, indicating that both cell types support viral replication. Finally, cycloheximide and a protein kinase C inhibitor (H7) that blocks the ex vivo activation of viral expression did not affect the increased survival in B-1-like cells, suggesting that resistance to apoptosis is acquired in vivo. Collectively, these results indicate a peculiar susceptibility of sheep B-1-like cells to BLV transforming effects and further support the involvement of increased survival in BLV pathogenesis.

Increased ratio of bcl-2/bax expression is associated with bovine leukemia virus-induced leukemogenesis in cattle.

To further investigate the molecular basis underlying the dysregulation of B cell homeostasis associated with bovine leukemia virus disease progression in cattle, bovine bax was cDNA cloned and sequenced. The predicted amino acid sequence of bovine Bax revealed a 192-amino-acid protein having extensive identity with the human (97%), murine (93%), and rat (94%) homologues. Because the ratio of Bcl-2 to Bax is believed to predetermine the susceptibility to a given apoptotic stimulus, the relative expression of the genes encoding these oncoproteins was evaluated in cattle naturally infected with BLV. In BLV-infected cattle an increase in the ratios of bcl-2/bax mRNA and protein expression correlated with advancing stages of disease. These findings suggest that in addition to the maintenance of BLV-associated hematopoietic malignancies, the reciprocal expression of Bcl-2/Bax may modulate the induction of B cell expansion typical of BLV disease progression.