Experimental infection of sheep with bovine herpesvirus type-5 (BHV-5): acute and latent infection

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Abstract

We demonstrated that sheep are susceptible to acute and latent infection by bovine herpesvirus type-5 (BHV-5). Lambs inoculated intranasally with two South American BHV-5 isolates replicated the virus with titers up to 107.1 TCID50/ml for up to 15 days and showed mild signs of rhinitis. Four lambs in contact with the inoculated animals acquired the infection and excreted virus for up to seven days. One lamb developed progressive signs of neurological disease and was euthanized in extremis. Clinical signs consisted of tremors of the face, bruxism, ptyalism, incoordination, lateral flexion of the neck and head, circling, walking backwards, recumbency and paddling. The virus was detected in the anterior and posterior cerebrum, dorso- and ventro-lateral cortex, cerebellum, pons, midbrain and olfactory bulb. Viral nucleic acids were demonstrated in neurons and astrocytes of the anterior and ventro-lateral cortex by in situ hybridization. Histological changes consisting of non-suppurative meningitis, perivascular mononuclear cuffing, focal gliosis, neuronal necrosis and intranuclear inclusions were observed in the anterior cerebrum, ventro-lateral cortex and midbrain. Dexamethasone treatment at Day 50 pi resulted in reactivation of the latent infection and virus shedding in 13/16 (81%) of the lambs. Together with previous reports of BHV-5 antibodies in sheep, these findings show that sheep are fully susceptible to BHV-5 suggesting that infection by BHV-5 in sheep may occur naturally.

Introduction

Bovine herpesvirus type-5 (BHV-5), also known as bovine encephalitis herpesvirus (BEHV), is an alphaherpesvirus associated with fatal meningoencephalitis in young cattle (Studdert, 1989; Roizman, 1992). Antigenically, genetically and biologically, BHV-5 is closely related to the respiratory and genital BHV-1 strains (Bagust, 1972; Metzler et al., 1986; Bulach and Studdert, 1990; Bratanich et al., 1991). The major difference between BHV-5 and BHV-1 is the distinct ability of the former to cause neurological disease in cattle (Bagust, 1972; Studdert, 1989; Belknap et al., 1994). Isolation of BHV-1 has been largely restricted to cases of respiratory and genital disease, whereas BHV-5 has been almost exclusively associated with meningoencephalitis (Studdert, 1989). Clinical signs of BHV-5-associated neurological disease include tremors, circling, bruxism, incoordination, nystagmus, recumbency followed by convulsions, paddling and inevitably death (Hall et al., 1966; Bagust and Clark, 1972; Carrillo et al., 1983).

The natural history, serology and geographic distribution of BHV-5 infections are largely unknown, partially due to the historic unprecise taxonomic classification of the virus. Only recently, serologic and virologic means of identifying and differentiating BHV-5 from BHV-1 on a field scale have been developed (Lindner et al., 1993a, Lindner et al., 1993b; d'Offay et al., 1995). Severe outbreaks of meningoencephalitis by BHV-5 have been frequently described in well-defined geographical locations, mainly in Argentina and southern Brazil (Carrillo et al., 1983; Schudel et al., 1986; Riet-Correa et al., 1989; Weiblen et al., 1989, Weiblen et al., 1996; Roehe et al., 1997; Odeon, 1998; Salvador et al., 1998). Elsewhere, BHV-5-associated neurological disease has been only sporadically reported (Johnston and McGavin, 1962; Bartha et al., 1969; Eugster et al., 1974). Cross-protection by naturally occurring or vaccine-induced BHV-1 antibodies has been suggested as a possible explanation for the rare occurrence of BHV-5-associated disease in BHV-1 endemic areas (d'Offay et al., 1995). The rare occurrence and high fatality of BHV-5-induced neurological disease have led to speculations about the origin of the virus. It has been suggested that BHV-5 might be a virus naturally infecting zebu cattle or another closely related ruminant species (Studdert, 1989; Bulach and Studdert, 1990).

In this study, we investigated the susceptibility of sheep to infection with two South American BHV-5 isolates. Our results demonstrate that sheep are fully susceptible to acute and latent BHV-5 infection and may even develop neurologic disease. Taken together with recent reports of BHV-5 antibodies in sheep (Lindner et al., 1993a, Lindner et al., 1993b), these findings suggest that natural infections by BHV-5 in this species may occur.

Section snippets

Experimental design

Two inoculations were performed independently to evaluate the susceptibility of sheep to BHV-5 infection. Lambs were inoculated intranasally with either of two South American BHV-5 isolates (A663 and EVI-88) and submitted to clinical and virological monitoring. In the first experiment (isolate A663), nine lambs were inoculated and three remained as non-inoculated controls. In the second experiment (isolate EVI-88), 12 lambs were inoculated and four non-inoculated lambs were maintained in close

Acute infection

Lambs inoculated with BHV-5 shed virus in nasal secretions from Day 1 to Day 16 pi (Table 1). Peaks of virus shedding were observed between Days 3 and 5 pi, declining progressively thereafter (data not shown). Maximum individual titers reached 107.1 and 105.9 TCID50/ml for isolates EVI-88 and A663, respectively. Some lambs inoculated with isolate EVI-88 shed virus in titers above 106 TCID50/ml for two to three days. The extent of virus shedding was capable of spreading virus to lambs maintained in

Discussion

The present study was conducted to investigate the susceptibility of sheep to infection with bovine herpesvirus type-5 (BHV-5). Recent studies have reported the natural occurrence of BHV-5 antibodies in sheep (Lindner et al., 1993a, Lindner et al., 1993b). In many regions, including some of those experiencing frequent outbreaks of meningoencephalitis due to BHV-5, cattle and sheep are raised together, sharing pastures and facilities. These mixed herds may provide conditions for interspecies

Acknowledgements

This work was supported by a MCT, CNPq, CAPES and FINEP grant (PRONEX em Virologia Veterinária, 215/96). E.F. Flores (352386/96), R. Weiblen (520161/97-1) and L.F. Irigoyen (523134/96-7) have scholarships from the Brazilian Council for Research (CNPq). F.A. Osorio's participation was partially supported by a fellowship by the Fullbright Commission in Brazil (Comissão Fullbright Brazil) and by a grant from USDA/FAS/ICD/RSED of the United States of America.

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