Diagnosis, Pathogenesis And Control Of Chicken And Turkey Arthritis Reoviruses

Abstract

Avian reoviruses are the causative agent of arthritis/tenosynovitis in broilers and turkeys. Recently, variants of chicken arthritis reovirus (CARV) and turkey arthritis reovirus (TARV) have occurred worldwide leading to vaccine failures, causing huge economic loses, and increasing animal welfare concerns. Whole genome sequencing of 35 and 14 isolates of CARV from the US and Germany, respectively, indicated significant divergence in these strains. In fact, US strains had more divergence among themselves than the German strains. The US strains clustered in six previously established genotype clusters (GCs) while the German strains clustered in only four of the six GCs. Strains from both countries were highly divergent from the currently used vaccine strains. Based on distinct evolutionary trees of each gene, we discovered that all genes evolve in an independent manner and contribute to the evolutionary process. The S1 (σC) genome segment showed noticeably higher divergence followed by M2 (μB) and L3 (λC) genes. Congruent topologies of these isolates indicated frequent genetic re-assortment among multiple co-circulating variants. We believe that the genetic variability among CARVs is due to a combination of evolutionary mechanisms involving multiple cocirculating lineages and genetic reassortments.We developed a recombinant pichinde virus-vectored vaccine (rPICV-CARV) that expresses the sigma C (SC) and sigma B (SB) antigenic proteins of CARVs. Several combinations of monovalent and bivalent rPICV-CARV vaccines were developed carrying codon-optimized SC and SB genes from two divergent CARV strains. The S1 and S3 genes and antigens were found to be expressed in virus-infected cells via reverse transcriptase polymerase chain reaction (RT-PCR) and direct fluorescent antibody (DFA) technique, respectively. We conducted an in vivo study to determine safety and efficacy of two vaccine formulations against a wide spectrum of virus challenges. The vaccinated birds produced serum-neutralizing antibodies, which were responsible for early clearing of the virus from the host, inhibited virus replication in intestine and tendons, and decreased fecal shedding of the virus relative to non-vaccinated controls. The vaccine is a promising candidate that needs to be further evaluated in breeders. The survival of bivalent codon optimized rPICV-CARV vaccine was studied in poultry litter and water at room temperature (approx. 25°C). In spiked samples, it was found that the vaccine virus survived for approximately six hours in litter and drinking water. Reoviruses have been isolated from three different disease syndromes in turkeys, e.g., turkey enteric reovirus (TERV) from cases of turkey enteritis, turkey arthritis reovirus (TARV) from cases of tenosynovitis/arthritis in turkeys, and turkey hepatitis reovirus (THRV) from cases of hepatitis in turkeys. The comparative pathogenesis of these viruses, and correlation with their genetic make-up (if any), is not known. All nine viruses were found to be enterotropic; the virus gene copy number in the intestine reached a peak at 5 dpi followed by a sharp decline at 7 dpi. All viruses caused significant decrease in body weight gain of birds compared to the negative control group. Both TARV and THRV strains replicated in tendons and produced histologic lesions consistent with tenosynovitis. Hepatic lesions were produced by THRV only and the virus was re-isolated from liver and spleen of inoculated birds thereby fulfilling Koch’s postulates. We then conducted a study to determine the age at which turkey poults become susceptible to infection with TARV. All turkeys were susceptible to TARV infection at all ages studied. However, virus replication was more pronounced in the intestine and gastrocnemius tendons of turkeys at 2-weeks of age or less. Additionally, turkeys at all ages of TARV challenge developed typical lesions of lymphoplasmacytic tenosynovitis, shed TARV in feces, and transmitted TARV to sentinels. The sentinels, in turn, also showed virus replication in their intestines and tendons leading to histological lesions of arthritis/tenosynovitis. These findings indicate that turkeys at the age of 28 days or less are susceptible to infection with TARV following oral challenge. It was also found that TARV-infected birds could transmit the infection to naïve sentinel turkeys of the same age. We also created a recombinant live pichinde virus-vectored bivalent subunit vaccine that expresses Sigma C and Sigma B proteins of TARV SKM121. The efficacy of this vaccine was tested against both homologous (TARV SKM121) and heterologous (TARV O’Neil) virus challenges. Immunized poults produced serum-neutralizing antibodies that neutralized both viruses. The body weights of vaccinated and non-vaccinated birds were similar indicating no adverse effect of the vaccine on feed efficiency. Comparison of virus gene copy numbers in intestine and histologic lesion scores in tendons of vaccinated and non-vaccinated birds showed a decrease in the replication of challenge viruses in the intestines and tendons of vaccinated birds. These results indicate the potential usefulness of this vaccine. In addition, the vaccine virus was found to be transmissible horizontally to non-vaccinated pen mates of vaccinated birds inducing serum neutralizing antibodies.