Analysis of Equid Herpesvirus 1 Strain Variation Reveals a Point Mutation of the DNA Polymerase Strongly Associated with Neuropathogenic versus Nonneuropathogenic Disease Outbreaks

Abstract

Equid herpesvirus 1 (EHV-1) can cause a wide spectrum of diseases ranging from inapparent respiratory infection to the induction of abortion and, in extreme cases, neurological disease resulting in paralysis and ultimately death. It has been suggested that distinct strains of EHV-1 that differ in pathogenic capacity circulate in the field. In order to investigate this hypothesis, it was necessary to identify genetic markers that allow subgroups of related strains to be identified. We have determined all of the genetic differences between a neuropathogenic strain (Ab4) and a nonneuropathogenic strain (V592) of EHV-1 and developed PCR/ sequencing procedures enabling differentiation of EHV-1 strains circulating in the field. The results indicate the occurrence of several major genetic subgroups of EHV-1 among isolates recovered from outbreaks over the course of 30 years, consistent with the proposal that distinct strains of EHV-1 circulate in the field. Moreover, there is evidence that certain strain groups are geographically restricted, being recovered predominantly from outbreaks occurring in either North America or Europe. Significantly, variation of a single amino acid of the DNA polymerase is strongly associated with neurological versus nonneurological disease outbreaks. Strikingly, this variant amino acid occurs at a highly conserved position for herpesvirus DNA polymerases, suggesting an important functional role.Equid herpesvirus 1 (EHV-1), a member of the subfamily Alphaherpesvirinae, is a highly prevalent equine pathogen that can cause a range of clinical signs, from respiratory distress to the induction of abortion, neonatal foal death, and occasionally neurological damage resulting in paralysis (9,14,19,33,37,61). The severity of disease resulting from EHV-1 infection is likely to be influenced by a number of factors, including the age and physical condition of the host; whether the infection is primary, a reinfection, or a reactivation of latent virus; the immune status of the host; and the pathogenic potential of the strain involved. In order to assess the relative importance of EHV-1 strain variation regarding disease outcome, it is necessary to develop methods enabling precise discrimination between genetic subgroups of interrelated strains. Previous studies have utilized DNA restriction fragment length polymorphism (RFLP) to separate field isolates of EHV-1 into subgroups according to characteristic restriction enzyme site changes and the presence of variable numbers of copies of short sequence repeats. These studies demonstrated a relatively low frequency of genetic polymorphism for EHV-1 and suggested that distinct strains of EHV-1 do exist in the field (3,4,8,25,32,41,54,57). However, the relative lack of variation of EHV-1 sequences between strains has resulted in too few RFLP variants to be identified for detailed epidemiological studies. Furthermore, although such analyses may be used for tracing the genetic relatedness of strains, they allow identification only of those genetic changes resul...

Genetic diversity of equine herpesvirus 1 isolated from neurological, abortigenic and respiratory disease outbreaks

“…The tandem repeat regions were difficult to sequence as they were often longer than the read length. Even when tandem repeats were short enough to be covered within a single read, they were observed to vary in length among strains, and sometimes within strains, as reported previously for EHV‐1 (Nugent et al., 2006) and other alphaherpesviruses (Depledge et al., 2014; Hondo & Yogo, 1988). They also tended to be G+C‐rich and thus seemed to be under‐represented in the sequence libraries, making it inadvisable to estimate their lengths from coverage density data.…”

“…Sometimes overlooked is the finding in the field that the association of abortion with the EHV‐1 A2254/N752 strain variant was more statistically significant (Nugent et al., 2006). The majority of strains analysed in the present study had been isolated from abortion cases and had the A2254/N752 genotype, and most of the remaining strains that had been isolated from neurological disease outbreaks belonged to the G2254/D752 genotype, similar to published conclusions.…”

“…This substitution was also found in the strain RacL11, along with two further changes in ORF30, the neurological marker N752D and Y753S. The polymorphism at position 753 has been reported previously in a subset of strains isolated in central Kentucky (Smith et al., 2010) and in the strain RacH used in the live attenuated vaccines Rhinomune (Boehringer Ingelheim Vetmedica) and Prevaccinol (MSD Animal Health) (Nugent et al., 2006). …”

“…More recent genetic studies include determination of the whole genome sequences for two well‐characterized strains, Ab4 and V592 (Telford et al., 1992; Nugent et al., 2006). Comparison of the sequence of the neuropathogenic strain Ab4 (Crowhurst, Dickinson, & Burrows, 1981; Telford et al., 1992) with that of the abortigenic strain V592 (Mumford et al., 1987; Nugent et al., 2006) identified 43 amino acid residue differences distributed among 31 ORFs (Nugent et al., 2006).…”

In vitro susceptibility of six isolates of equine herpesvirus 1 to acyclovir, ganciclovir, cidofovir, adefovir, PMEDAP and foscarnet

“…The drug should be effective against isolates of EHV-1 associated with outbreaks of abortion as well as those associated with outbreaks of neurological disorders. Recently, it was postulated that a variation of a single amino acid of the DNA polymerase is strongly associated with neurological versus non-neurological disease outbreaks (Nugent et al, 2006). This may have an implication on the susceptibility of various isolates to anti-herpetic compounds as many of these compounds act on the DNA polymerase (De Clercq, 2004).…”