Jin-an Huang

Equine rhinitis A virus: structural proteins and immune response.

Equine rhinitis A virus (ERAV) is a picornavirus that has been reclassified as a member of the Aphthovirus genus because of its resemblance to foot-and-mouth disease virus at the level of nucleotide sequence and overall genomic structure. The N-terminal amino acid sequence of three of the four capsid proteins of ERAV was determined and showed that the proteolytic cleavage sites within the precursor P1 polypeptide occur exactly as those predicted for an aphthovirus-like 3C protease, which generates the capsid proteins VP1 and VP3. However, the autocatalytic cleavage site between VP4 and VP2, which is independent of 3C protease cleavage, was different from that predicted previously. ERAV.393/76 antisera from horses and rabbits showed different reactivity to the viral structural proteins in both serum neutralization assays and Western blots. High neutralizing antibody titres appeared to correlate with strong reactivity to VP1 in Western blots.

Sequence Conservation and Antigenic Variation of the Structural Proteins of Equine Rhinitis A Virus

ABSTRACT The nucleotide and deduced amino acid sequences of the P1 region of the genomes of 10 independent equine rhinitis A virus (ERAV) isolates were determined and found to be very closely related. A panel of seven monoclonal antibodies to the prototype virus ERAV.393/76 that bound to nonneutralization epitopes conserved among all 10 isolates was raised. In serum neutralization assays, rabbit polyclonal sera and sera from naturally and experimentally infected horses reacted in a consistent and discriminating manner with the 10 isolates, which indicated the existence of variation in the neutralization epitopes of these viruses.

Comparative sequence analysis of VP7 genes from five Australian porcine rotaviruses

SummaryThe genes coding for the rotavirus major neutralizing protein, VP7, from 5 Australian porcine rotaviruses representing glycoprotein (i.e. VP7 or G) serotypes 3, 4, and 5, were sequenced. The genes were each 1,062 nucleotides long with two long open reading frames for proteins of either 326 or 297 amino acids and containing only one potential glycosylation site at amino acid position 69. When compared to the corresponding genes of human viruses, the porcine genes showed very high nucleotide and deduced amino acid homology. Sequence comparison also revealed that Australian porcine rotaviruses of G serotype 4 and 5 were similar to the corresponding porcine strains found in the U.S.A. and U.K., while G serotype 3 and 4 porcine rotaviruses were closely related to human G serotype 3 strain, RV-3 and serotype 4 strain, ST-3, respectively. These Australian rotavirus VP7 sequences were found to correlate with serological data we reported previously.

Polymorphism of open reading frame 71 of equine herpesvirus-4 (EHV-4) and EHV-1

Open reading frame (ORF) 71 genes of both equine herpesvirus-1 (EHV-1) and EHV-4 encode a unique glycoprotein, which has been described to vary in molecular mass from 200 to 450 kDa. Using PCR and nucleotide sequence analysis, it was shown that the ORF 71 genes of EHV-1 and EHV-4 are polymorphic due to a variable number of reiterated sequences in two regions, designated regions A and B. Region A was threonine-rich and was located near the N terminus. Region B comprised a 38 amino acid repeat near the C terminus that expanded following cell culture adaptation. Western blot analysis of viruses showed that EHV-4 gp2 was modified by glycosylation and that variation in region A resulted in the marked differences in the molecular mass of EHV-4 gp2.

Persistence and chronic urinary shedding of the aphthovirus equine rhinitis A virus.

Equine rhinitis A virus (ERAV) is a member of the Aphthovirus genus, and has many physical and structural similarities to the prototype Aphthovirus foot-and-mouth disease virus (FMDV). The pathogenesis of FMDV has been extensively studied, however, the similarities in the pathogenesis of ERAV and FMDV disease has not been well documented. This study describes and compares the pathogenesis of ERAV both in the natural host and a small animal model alternative (CBA mice). Distinct parallels in the pathogenesis of the acute infection of these two viruses are described where infection in the upper respiratory tract precedes shedding of high levels of virus from the nasopharynx and a transient viraemic phase before dissemination to distal sites. The finding that ERAV is maintained at high levels in the urine of infected horses for at least 37 days post infection, however, is a feature unique to ERAV amongst all of the picornaviruses.

Equine rhinitis A virus-like particle expressing DNA vaccine induces a virus neutralising immune response in mice.

Equine rhinitis A virus (ERAV) is a respiratory pathogen of horses. Candidate vaccines to date have been hindered by low expression levels and the induction of non-neutralising antibodies. The immunodominant epitope of ERAV is conformational and is located within the quaternary structure of the capsid. This site should be retained in ERAV virus-like particles (VLPs) to stimulate the induction of neutralising antibodies. The immunogenicity of a plasmid-based DNA vaccine designed to express ERAV VLPs was assessed. The plasmid construct, pcD.P12A.3C, contained the capsid precursor (P1-2A) and the viral protease 3C, under the transcriptional control of a cytomegalovirus (CMV) promoter. Mature viral capsid proteins and VLPs were detected in vitro in transfected COS7 cells. Immunisation of BALB/c mice with pcD.P12A.3C induced virus neutralising antibodies and enhanced the virus neutralising antibody response to purified, UV-inactivated ERAV. This study further supports the use of DNA vaccines to elicit neutralising antibodies to complex antigenic proteins.

Glycoprotein G deletion mutants of equine herpesvirus 1 (EHV1; equine abortion virus) and EHV4 (equine rhinopneumonitis virus)

Summary.Glycoprotein G (gG) deletion mutants of EHV1 and EHV4, designated EHV1ΔgG and EHV4ΔgG, were constructed. The growth characteristics of the EHV1ΔgG mutants were similar to the parent virus. All of the EHV4ΔgG mutants grew more slowly in cell culture and produced plaques of different morphology including smaller size. The yields of both gG deletion mutant viruses in cell culture were similar to the parent viruses. Sequencing of the genes flanking gG, Southern blot, PCR and western blot analyses of the mutant viruses demonstrated that the deletions were as expected, except for EHV4ΔgG mutants, which in addition to deletion of gG contained unexpected deletions in the adjacent down stream gene ORF 71 (glycoprotein 2). Antisera to EHV1ΔgG and EHV4ΔgG neutralised the respective mutant and the parent viruses to the same titre and these antisera could be distinguished from antisera to the wild type viruses in a gG antibody detection ELISA. The mutant viruses may be useful as vaccine candidates and the deletion of gG may act as a marker to distinguish vaccinated from the naturally infected horses.

Duration of serological response to canine parvovirus-type 2, canine distemper virus, canine adenovirus type 1 and canine parainfluenza virus in client-owned dogs in Australia.

OBJECTIVE To determine whether client-owned dogs in Australia, last vaccinated with Canvac(®) vaccines containing canine parvovirus-type 2 (CPV-2), canine distemper virus (CDV), canine adenovirus type 2 (CAV-2) ± canine parainfluenza virus (CPiV) at least 18 months ago, were seropositive or responded serologically to revaccination. METHODS A total of 235 dogs were recruited from 23 veterinary clinics, representing a variety of breeds, ages and time since last vaccination (TSLV: range 1.5-9 years, mean 2.8 years). Dogs had a blood sample taken and were revaccinated on day 0. A second blood sample was taken 7-14 days later. Blood samples were assessed for antibody titres to CPV-2 (by haemagglutination inhibition) and CDV, CAV type 1 (CAV-1) and CPiV (by virus neutralisation). Dogs with a day 0 titre >10 or a four-fold increase in titre following revaccination were considered to be serological responders. RESULTS The overall percentage of dogs classified as serological responders was 98.7% for CPV-2, 96.6% for CDV, 99.6% for CAV-1 and 90.3% for CPiV. CONCLUSIONS These results suggest that the duration of serological response induced by modified-live vaccines against CPV-2, CDV, CAV-1 and CPiV, including Canvac(®) vaccines, is beyond 18 months and may extend up to 9 years. Accordingly, these vaccines may be considered for use in extended revaccination interval protocols as recommended by current canine vaccine guidelines.