Comparative evaluation of bovine immunodeficiency-like virus infection by reverse transcriptase and polymerase chain reaction.

Infection of embryonic bovine lung (EBL) cells by bovine immunodeficiency-like virus (BIV) were monitored by reverse transcriptase (RT), syncytia formation and polymerase chain reaction (PCR). Infection can be detected by PCR at 24 h while the presence of syncytia and RT were not detected until much later. The detection of BIV RT can be optimized by changing the pH and salt conditions. The enzyme is very sensitive to changes in pH but can tolerate a wider range of salt and MgCl2 concentrations. Infection of primary human cell cultures by BIV was monitored by both PCR and RT. No active infection of human cells were detectable.

Bovine immunodeficiency virus in experimentally infected rabbit: tropism for lymphoid and nonlymphoid tissues.

The bovine immunodeficiency virus (BIV)/New Zealand (Oryctolagus cuniculus) rabbit model was used to study events that underlie the early and chronic stages of viral replication, routes and time course of viral dissemination and the distribution of the virus in the lymphoid. nonlymphoid and mucosa associated tissues. The results indicated that BIV, a lentivirus with genetic relatedness to the HIV, induced changes of clinical (anorexia, weight loss, muscular wasting, diarrhea, hypoalgesia, torticollis), immunological (recurrent T- and B-cell dysfunctions) and histopathological (lymphadenopathy, splenomegaly) nature that closely parallels those described for cat (Fly), monkey (SIV) and human (HIV) lentiviral diseases. These findings showing that BIV induces both splenomegaly and lymphadenopathy syndromes with associated fatal immune dysfunctions and the ability of the virus to replicate productively at the mucosal surfaces in rabbits, emphasize the importance of the BIV/rabbit system as a good small-animal model for the study of retrovirus-induced AIDS and offers the opportunity to evaluate prophylactic and therapeutic anti-retroviral agents of relevance to HIV-1 as well as the opportunity to study mechanisms of drug resistance phenomena.

Bovine immunodeficiency-like virus: inactivation in milk by pasteurisation.

Bioassay was used to determine whether bovine immunodeficiency-like virus (BIV) in milk was inactivated by pasteurisation. Three groups of three calves were inoculated with virus (BIV isolate FL112), milk seeded with virus and milk seeded with virus that had been pasteurised before inoculation, respectively. Seroconversion to BIV was monitored for 12 months by an indirect immunofluorescence assay. The presence of BIV proviral DNA in peripheral blood was determined by a nested polymerase chain reaction (PCR). The animals were euthanized and virus isolation and PCR were attempted on peripheral blood mononunclear cells, prescapular lymph node and spleen. Transmission of BIV was confirmed in the groups that were inoculated with the virus and with the virus in milk, but no evidence of its transmission was demonstrated in the group that received the pasteurised inoculum.

Bovine immunodeficiency virus expression in vitro is reduced in the presence of beta-chemokines, MIP-1alpha, MIP-1beta and RANTES.

The inhibition of HIV expression in vitro by a cocktail of the beta-chemokines MIP-1alpha, MIP-1beta and RANTES provided the initial evidence that HIV utilizes chemokine receptors as co-receptors for infection of cells. Bovine immunodeficiency virus (BIV), a lentivirus, infects a wide variety of leukocyte populations, but the cellular receptor(s) utilized by this virus for infection of cells is not known. The purpose of this study was to determine whether MIP-1alpha, MIP-1beta and RANTES affect BIV expression in vitro, as a prelude to identifying the cellular receptors utilized by this virus. Fetal bovine lung (FBL) cells were pretreated with serial dilutions of a cocktail of the chemokines, and then the cells were infected with BIV. Virus expression in these cells was determined by counting the syncytia that had developed in the cultures by five days after infection. A significant decrease in syncytium formation, corresponding to increasing concentrations of the chemokines, was the result. Reacting the chemokines with chemokine-specific neutralizing antibodies prior to treatment of the cells neutralized the effect of the chemokines on virus replication in a dose-dependent manner, restoring viral expression to a level similar to that of untreated cells. The presence of a CCR5 homologue on the surface of FBL cells was confirmed using an anti-CCR5 monoclonal antibody and FACS analysis. Collectively, these data provide preliminary evidence that BIV may utilize the CCR5 receptor for infection of cells in vitro, but additional studies are necessary to confirm this.

Bovine immunodeficiency virus in relation to embryos fertilized in vitro.

The association of bovine immunodeficiency virus (BIV) with embryos derived by in vitro fertilization from oocytes of experimentally infected heifers or oocytes/embryos exposed to the virus in vitro was investigated. Using a nested-PCR assay, proviral DNA of BIV was not detected in follicular fluid or in embryos derived from BIV-infected donors. In vitro exposure of oocytes to BIV during maturation or insemination with BIV-infected semen resulted in zona pellucida-intact embryos testing negative for BIV provirus. However, exposure of zona pellucida-free day-7 embryos to the virus resulted in a positive BIV assay for 28% of the batches of embryos, suggesting that the zona pellucida has a role in protecting against BIV infection. The presence of BIV in the IVF system had no apparent effect on the development of bovine embryos to the blastocyst stage.

Development of the bovine immunodeficiency-like virus as a model of lentivirus disease.

The bovine immunodeficiency-like virus (BIV) is morphologically, serologically, and genetically related to the lentivirus subfamily of retroviruses which includes human and simian immunodeficiency viruses and other lentiviruses causally associated with debilitating diseases of domestic animals. There are many parallels in the biology and pathologic characteristics of BIV infections with those of HIV that make its development as a model of HIV-like infection and disease potentially attractive. In order to obtain a better understanding of the molecular basis of BIV-induced disease, two biologically active proviruses of BIV were molecularly cloned and sequenced. The BIV genome is 9.0 kilobases in the form of the proviral DNA. It contains the obligate retroviral structural genes, gag, pol, and env. In addition, in the BIV central region, between and overlapping pol and env, there are five potential coding regions for non-structural/regulatory genes; three are analogous to vif, tat, and rev in HIV and two, called W and Y, are unique to BIV. There is no coding region analogous to nef in BIV. Sequence comparisons of two functional proviruses obtained from the DNA of cells carrying an infection from a single virus isolation indicate that the genome of BIV is highly variable within a single biological isolate. Moreover, the greatest number of substitutions occur in the env gene. The results suggest the presence of multiple genotypes which may be of significance in defining the disease potential of a BIV isolate. These clones will be useful in dissecting the replicative cycle and mechanisms of pathogenesis of BIV in various animal models.

A western blot assay for the detection of antibodies to bovine immunodeficiency-like virus in experimentally inoculated cattle, sheep, and goats.

A cocultivation method was used to establish a cytocidal bovine immunodeficiency-like virus (BIV) infection in primary fetal bovine lung (FBL) cell cultures. Cultures were monitored for virus production using radial immunodiffusion and agar gel immunodiffusion. Pelleted virus and detergent (CHAPS)-solubilized infected cell lysates from BIV-infected cell cultures were compared as sources of antigen for Western blots. Pelleted virus preparations from FBL-BIV cell cultures produced the best antigen for Western blot. Sheep and goats were inoculated with BIV and serum antibody responses were monitored up to 1 year post inoculation (PI). Sera from experimentally infected cattle, sheep, and goats reacted in Western blot assay with BIV viral induced polypeptides gp 110, p 72, p 55, p 50, gp 42, p 38, p 26, p 24, p 18, p 15, and p 13. Antibodies to p 26 were detected as early as 2 weeks PI in cattle, sheep, and goats. Antibodies to gp 110 were detected by 4 to 6 weeks PI in cattle, and by 9 months PI in sheep and goats. Antibodies to BIV proteins were still evident in cattle sera 2 1/2 years PI, and in sheep and goat sera 1 year PI.