Publications

The RNA segment (Gene 10) from a human group B rotavirus which encodes the homologue of the rotavirus enterotoxin (NSP4) has been cloned and sequenced. The gene is of the same length (751 nucleotides) as its better-characterized group A rotavirus counterpart but shows minimal homology (?10%) to it at the primary sequence level. Despite this low level of sequence homology, secondary structure predictions for the group B protein (ADRV-NSP4) showed a close similarity of structural features with the group A protein. Full-length ADRV-NSP4 was expressed in Escherichia coli with an amino terminal 6xHis tag that was used to purify it to homogeneity. The cytotoxicity of the purified protein was examined in a rapid dye-uptake assay that assesses membrane permeability and was found to be comparable to its group A counterpart.

Field strains of Foot-and-mouth disease virus (FMDV) use a number of alpha v-integrins as receptors to initiate infection on cultured cells, and integrins are believed to be the receptors used to target epithelial cells in animals. In this study, immunofluorescence confocal microscopy and real-time RT-PCR were used to investigate expression of two of the integrin receptors of FMDV, alpha v beta 6 and alpha v beta 3, within various epithelia targeted by this virus in cattle. These studies show that alpha v beta 6 is expressed constitutively on the surfaces of epithelial cells at sites where infectious lesions occur during a natural infection, but not at sites where lesions are not normally formed. Expression of alpha v beta 6 protein at these sites showed a good correlation with the relative abundance of beta 6 mRNA. In contrast, alpha v beta 3 protein was only detected at low levels on the vasculature and not on the epithelial cells of any of the tissues investigated. Together, these data suggest that in cattle, alpha v beta 6, rather than alpha v beta 3, serves as the major receptor that determines the tropism of FMDV for the epithelia normally targeted by this virus.

Fourteen pigs were inoculated with the 'Alfort 187' strain of classical swine fever (CSF) virus and killed in pairs at 2, 4, 7, 9, 11, 14 or 17 days post-inoculation for histopathological, ultrastructural and immunohistochemical examination. For the latter method, the antibodies used were those against viral antigen Gp55, porcine myeloid marker SWC3, IL-1 alpha, IL-6, TNF-alpha and Factor VIII-related antigen. Activation and increase in the number of hepatic macrophages was observed following viral detection in liver, as well as an increase in IL-1 alpha and IL-6 production, mainly by Kupffer cells. Maximum detection of viral antigen was observed in the middle stage of the experiment coinciding with overexpression of the three cytokines studied, with IL-6 production by interstitial macrophages prominent at the end. Additionally, the labelling of platelets for Factor VIII-related antigen and the ultrastructural study of the sinusoids revealed activation and aggregation of thrombocytes close to Kupffer cells at the beginning of the infection. The liver seems to play a prominent role in the origin of the thrombocytopenia that occurs in CSF and contributes to the overexpression of proinflammatory cytokines considered responsible for the disorders observed during the course of the disease.

Detection of antibodies to the non-structural proteins (NSP) of foot-and-mouth disease virus (FMDV) was compared with conventional serological and virological methods and with RT-PCR for the identification of FMDV carrier animals obtained after experimental contact challenge of vaccinated cattle. Transmission from carriers to sentinels was also monitored. Twenty FMDV vaccinated and five unvaccinated cattle were challenged by direct contact with five donor cattle excreting FMDV and monitored until 28 days post challenge-exposure [1]. Twelve vaccinated and three unvaccinated animals were retained up to 24 weeks post exposure to FMDV in order to monitor viral persistence, transmission and antibody responses. In nine vaccinated animals, infection persisted beyond 28 days post exposure, virus being detected more frequently and for longer in oesophagopharyngeal samples from these animals when examined by RT-PCR rather than by virus isolation. Although recovery of FMDV RNA became increasingly sporadic over time, the number of RNA copies detected in positive samples declined only slowly. Two naïve sentinel cattle housed with the persistently infected animals between 93 and 168 days after the latter had been challenge-exposed to FMDV did not become infected. There were differences in the ability of commercially available serological tests to detect antibodies to FMDV non-structural proteins (NSP) in vaccinated and subsequently challenged cattle. Although no single test could identify all of the vaccinated cattle that became persistently infected, the most poorly recognised animals were those with the least evidence of virus replication based on other tests. The potential of the detection of antibodies to the 2B NSP of FMDV for diagnosing persistent FMDV infection was demonstrated.