Sandra E. Pérez

Toll-like receptor activation and expression in bovine alpha-herpesvirus infections

The involvement of Toll-like receptors (TLRs) in bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) infections has not been analyzed. In this study, the role of TLR signaling on virus replication was investigated. Blood leukocytes consistently express TLRs. Thus, our approach was to study in vitro the effects of agonist stimulation of TLRs expressed by peripheral blood leukocytes on BoHV-1 and BoHV-5 replication. Furthermore, the patterns of TLRs 3, 7-9 expression on virus-infected-bovine leukocytes were analyzed. Only Imiquimod (TLR7/8 agonist) showed anti-viral activity on infected MDBK cells. This is the first evidence that the timely activation of TLR7/8 signaling is effective in impairing BoHV-1 and 5 replication, thereby providing an experimental indication that Imiquimod may be a promising immune modulator. This work describes, for the first time, the expression patterns of TLRs in BoHV-1- or BoHV-5-infected-bovine leukocytes, suggesting the involvement of TLR7 and TLR9 in the recognition of these viruses.

Involvement of toll-like receptors 3 and 7/8 in the neuropathogenesis of bovine herpesvirus types 1 and 5

Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are closely related alpha-herpesviruses. BoHV-5 is the causal agent of non-suppurative meningoencephalitis in calves. BoHV-1 causes respiratory disease, abortions, genital disorders and, occasionally, encephalitis in cattle. Both viruses are neurotropic and they share similar biological properties. Nevertheless, they differ in their ability to cause neurological disease. Toll-like receptors (TLRs) are involved in the innate immune response to pathogens. In this study, the variations in the expression levels of TLRs were evaluated in different regions of the bovine central nervous system during the acute infection and reactivation of BoHV-1 and BoHV-5- infected cattle. With the exception of TLR9, significant up-regulation of all TLRs was detected following primary infection of neural tissues by both bovine alpha-herpesviruses. Furthermore, the stages of acute infection and reactivation were characterized by a distinguishable TLR expression pattern. Important differences in TLR expression upon infection of the central nervous system by BoHV-1 or BoHV-5 were not detected. The striking differences in TLR mRNA levels during acute infection and reactivation provide evidence that the innate immune response may be involved in the clinical outcomes observed at each stage. Further research is required to analyze the mechanisms that initiate TLR activation and the signaling cascade mediated by each TLR to elucidate the precise role these receptors play in bovine herpesvirus encephalitis.

Molecular Characterization of the First Bovine Herpesvirus 4 (BoHV-4) Strains Isolated from In Vitro Bovine Embryos production in Argentina

Bovine herpesvirus 4 (BoHV-4) is increasingly considered as responsible for various problems of the reproductive tract. The virus infects mainly blood mononuclear cells and displays specific tropism for vascular endothelia, reproductive and fetal tissues. Epidemiological studies suggest its impact on reproductive performance, and its presence in various sites in the reproductive tract highlights its potential transmission in transfer-stage embryos. This work describes the biological and genetic characterization of BoHV-4 strains isolated from an in vitro bovine embryo production system. BoHV-4 strains were isolated in 2011 and 2013 from granulosa cells and bovine oocytes from ovary batches collected at a local abattoir, used as “starting material” for in vitro production of bovine embryos. Compatible BoHV-4-CPE was observed in the co-culture of granulosa cells and oocytes with MDBK cells. The identity of the isolates was confirmed by PCR assays targeting three ORFs of the viral genome. The phylogenetic analyses of the strains suggest that they were evolutionary unlinked. Therefore it is possible that BoHV-4 ovary infections occurred regularly along the evolution of the virus, at least in Argentina, which can have implications in the systems of in vitro embryo production. Thus, although BoHV-4 does not appear to be a frequent risk factor for in vitro embryo production, data are still limited. This study reveals the potential of BoHV-4 transmission via embryo transfer. Moreover, the high variability among the BoHV-4 strains isolated from aborted cows in Argentina highlights the importance of further research on the role of this virus as an agent with the potential to cause reproductive disease in cattle. The genetic characterization of the isolated strains provides data to better understand the pathogenesis of BoHV-4 infections. Furthermore, it will lead to fundamental insights into the molecular aspects of the virus and the means by which these strains circulate in the herds.

Infecção simultânea de Neospora caninum e Herpesvirus bovino tipo 5 em casos espontâneos de aborto bovino

ABSTRACT.- Marin M.S., Morrell E.L., Perez S.E., Leunda M.R., Moore D.P., Jones L.R., Cam-pero C.M. & Odeon A.C. 2013. Concomitant infection of Neospora caninum and Bovine Herpesvirus type 5 in spontaneous bovine abortions. Pesquisa Veterinaria Brasileira 33(11):1291-1295. Area de Investigacion en Produccion y Sanidad Animal, Instituto Nacional de Tecnologia Agropecuaria (INTA), Estacion Experimental Agropecuaria Balcarce, Ruta 226 Km 73.5 (7620), Balcarce, Buenos Aires, Argentina. E-mail: aodeon@balcarce.inta.gov.arBovine Herpesvirus type 5 (BoHV-5) has not been conclusively demonstrated to cau-se bovine abortion. Brain lesions produced by Neospora caninum and Bovine Herpesvirus type 1 (BoHV-1) exhibit common features. Therefore, careful microscopic evaluation and additional diagnostic procedures are required to achieve an accurate final etiological diag-nosis. The aim of the present work was to investigate the occurrence of infections due to BoHV-1, BoHV-5 and N. caninum in 68 cases of spontaneous bovine abortions which sho-wed microscopic lesions in the fetal central nervous system. This study allowed the identi -fication of 4 (5.9%) fetuses with dual infection by BoHV-5 and

Herpesvirus bovino 4 (BoHV-4): aspectos generales de su biología y situación en la República Argentina

Bovine herpesvirus 4 (BoHV-4) has been isolated from cattle with respiratory infections, vulvovaginitis, mastitis, abortions, endometritis and from apparently healthy animals throughout the world. Although it has not yet been established as causal agent of a specific disease entity, it is primarily associated with reproductive disorders of cattle. This virus can infect a wide range of species, either in vivo or in vitro. Two groups of prototype strains were originated from the first isolates: the DN599-type strains (American group) and the Movar-type strains (European group). In Argentina, BoHV-4 was isolated and characterized in 2007 from vaginal discharge samples taken from cows that had aborted. So far, more than 40 isolates, mainly associated with aborting bovine females have been registered in our country.

Bovine herpesvirus type 5 replication and induction of apoptosis in vitro and in the trigeminal ganglion of experimentally-infected cattle

Bovine herpesvirus (BoHV) types 1 and 5 are neuroinvasive. Cases of BoHV-1-induced encephalitis are not as frequent as those caused by BoHV-5. In this study, the capability of BoHV-5 to induce apoptosis in cell cultures and in the trigeminal ganglion during acute infection of experimentally-infected cattle was analyzed. Apoptotic changes in cell cultures agree with the ability of the viral strains to replicate in each cell line. Marked differences were observed between the in vitro induction of apoptosis by BoHV-1Cooper and BoHV-5 97/613 strains. Apoptotic neurons were clearly evident in the trigeminal ganglion of BoHV-1-infected calves. For BoHV-5 a fewer number of positive neurons was observed. There is an association between the magnitude of bovine herpesviruses replication and the induction of apoptosis in trigeminal ganglion. These findings suggest that the induction of apoptosis and the innate immune response orchestrate the final outcome of alpha herpesviruses infection of the bovine nervous system.