Hanna Ungar-Waron

Detrimental effect of bovine leukemia virus (BLV) on the immunological state of cattle

Bovine leukemia virus (BLV) is a retrovirus which seems to affect both the humoral and the cellular immune response. Cows affected by enzootic bovine leukemia (EBL) showed a reduction of IgM-producing cells in the spleen and lymph nodes. Experimentally infected calves had lower levels of secretory IgM and a decrease in T lymphocytes in the peripheral blood. The reduction in the amount of T cells was noticed mainly in cells bearing the CD4 markers. BLV-infected animals showed diminished responsiveness to newly encountered antigens. Cows naturally infected by BLV produced Igs with impaired structural or biological reactivity. The primary immune response was shown to be deficient in BLV-infected cows following vaccination with synthetic antigen. A marked shift in the proportion of PBL, especially of the CD5+ subset, was noticed. Peripheral blood mononuclear cells from BLV-infected cows secrete elevated levels of certain cytokines and contain increased levels of cytokine mRNA. High levels of cytokines are also found in the sera of BLV-infected cows compared to non-infected animals. A correlation was found between BLV infection and lack of spontaneous recovery from Trichophyton verrucosum infection. Moreover, some studies ascertained a significant association between the herd BLV infection status and disease incidence. The culling rate was higher and milk production lower in BLV-infected vs. BLV-free herds. It seems that BLV infection affects the immune system of a cow to such an extent that it ceases to be productive enough to be kept and, in most cases, the animal is culled before any symptoms of illness associated with persistent immunodeficiency become apparent.

Cellular immune response cytokine expression during the initial stage of bovine leukemia virus (BLV) infection determines the disease progression to persistent lymphocytosis.

We have established experimental models of bovine leukemia virus (BLV) infection followed by progression to persistent lymphocytosis (PL) positive (BLV+PL+) or PL negative (BLV+PL-) stages of infection. Two out of six BLV infected animals developed PL+ 4 weeks after BLV infection. One other animal became PL+ late in the course of infection and three infected animals stayed PL-. These animals (PL-) exhibited transient lymphocytosis 3-4 weeks after infection and sustained PL- lymphocyte counts up to 24 weeks after infection. Competitive RT-PCR analysis of IFN-gamma mRNA expression revealed that peripheral blood mononuclear cells (PBMC) of animals with PL+ status developed by 4 weeks after infection had augmented IFN-gamma mRNA expression 3-4 weeks after BLV infection. However PBMC of animals that sustained a long-termed PL- lymphocyte count had elevated IFN-gamma mRNA expression 1-24 weeks after infection. Competitive RT-PCR analysis of IL-2 mRNA expression showed an increase in the levels of IL-2 mRNA in PL animals. Interleukin-10 (IL-10) mRNAs expression were elevated both in PL+ and PL- animals from 3 and 12 weeks after infection respectively. We suggest that early and extended expression of cellular response cytokines may delay the progression to PL+ in enzootic bovine leukemia.

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.

IgG and IgM antibodies in normal and leukaemic cattle

Abstract Normal cattle were immunized with a native protein, human serum albumin (HSA), and a multichain synthetic polypeptide, poly(Tyr, Glu)-poly( dl Ala)-poly(Lys), (T,G)-A—L. The synthetic antigen provoked precipitable antibody formation when administered in complete Freunds adjuvant but not when injected in incomplete adjuvant. Intradermal and subcutaneous injections resulted in a similar pattern of antibody production. HSA was tested only in complete Freunds adjuvant. The temporal pattern of the immune response was similar in all cattle to both antigens. In the primary response antibodies of IgG and IgM classes could be detected. In anti-(T,G)-A—L sera there was a relatively high percentage of IgM antibodies after primary immunization. After secondary immunization the antibodies were elicited for a longer period, and were exclusively of the IgG class. Leukaemic cattle were injected with the same antigens. In each responding cow antibody appearance occurred late, was low, and the antibodies produced were mainly of the IgG class. Even though all the leukaemic cattle possessed IgM in their sera, they produced very little IgM class antibody upon antigenic stimulation, compared with normal cattle. A synthetic antigen, (T,G)-A—L, which is capable of eliciting antibodies of the IgM class in almost equal amounts to those of the IgG class in normal cattle, was not capable of causing the production of more than traces of IgM antibodies. It seems, therefore, that the mechanism of IgM synthesis must be significantly impaired in leukaemic cattle in addition to a general depression of the immune response.

Somatic cell hybridization between bovine leukemia virus-infected lymphocytes and murine plasma cell tumors: Cell fusion studies with bovine cells☆

Hybridization of peripheral blood lymphocytes from bovine leukemia virus-infected cows with murine myeloma cells resulted in the generation of hybrid cells secreting immunoglobulins composed of various combinations of heavy and light chains of both bovine and murine Ig origin. Some hybrid cells derived from the light-chain producer, but non-secretor murine myeloma NSI cell line, secreted IgM molecules composed of bovine mu-chain linked to bovine and/or murine light chains. Other hybridomas secreted mouse and bovine light-chain dimers and/or monomers, or failed to secrete any Ig polypeptide chain whatsoever. Immunoglobulins secreted by hybridomas obtained upon hybridization of bovine cells with the IgG-secreting murine myeloma P2X63 cell line, contained bovine mu-chain in one of the seven hybridomas obtained, and bovine light chain in two of them. All the cell lines secreted murine light- and gamma-chains.

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.

Purification and characterization of IgM and of its μ-chain from bovine serum

Abstract A method is described for the purification of bovine IgM, of its pure μ-chain, and for the production of their respective specific antisera.

γδ T-lymphocytes and anti-heat shock protein reactivity in bovine leukemia virus infected cattle

Bovine leukemia virus (BLV) induces a chronic infection in cattle that may result in persistent lymphocytosis (PL) and, sometimes, enzootic bovine leukosis. The cellular and humoral immune responses of the host following infection have been extensively investigated but little is known about the involvement of gamma delta T-cells in BLV pathogenesis. The affluence of these cells in cattle, and particularly in the peripheral blood of young ruminants, may suggest a particular role for them in defense mechanisms. In this study we have examined circulating gamma delta lymphocytes that express workshop clusters 1 (WC1) and 2 (WC2). In healthy cattle the WC1 cell count tends to decrease with age and adult cattle blood has statistically lower numbers (19.0 +/- 6.6%) than that of young animals (40.1 +/- 7.2%). However, in the blood of BLV-seropositive adult cattle and mainly in BLV+ PL+ animals the population of WC1 cells is elevated compared with uninfected animals (P < 0.007). Likewise, the WC2 cells count is increased (P < 0.01) in BLV+PL+. Furthermore, we have investigated whether BLV infection up-regulates the expression of heat shock proteins (HSP) which in turn could augment the humoral response. Anti-HSP70 activity was examined in the sera of 34 BLV-infected cattle and 40 healthy controls by ELISA. Significantly higher activities (P < 0.001) were observed in BLV-infected cattle.

Reactivity of IgM-rheumatoid factors of human, feline and bovine sera.

The reactions of IgM-rheumatoid factors (IgM-RFs) from human, feline and bovine sera were examined with their homologous monomeric IgGs and with IgGs of other species. Cross-reactivities with IgGs of other species were observed in each separate system. By the use of an enzyme-linked immunosorbent assay (ELISA), measurable differences in reactivities were obtained. The results suggest that IgM--RFs of human and feline origin recognize a similar determinant on the Fc region of IgG. A different moiety on the latter molecule appears to be involved in the reactivity of bovine IgM-RF.

Rheumatoid factors in natural retrovirus infections of the cat and cattle

The occurrence of IgM-rheumatoid factors (RFs) was investigated in natural retrovirus infection of cats with feline leukaemia virus and of cattle with bovine leukaemia virus. IgM-RFs of polyclonal character were detected. No significant differences were observed between the amounts of IgM-RFs in the retrovirus-infected animals and their respective controls.