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In recent years, the emerging avian orthoreovirus (ARV) strains that led to viral arthritis have attracted much attention from the chicken industry worldwide, due to the significant economic losses suffered. In mid-2020, with the assistance of next-generation sequencing technology, we achieved success in characterizing two divergent avian orthoreovirus (ARV) variants (0543/SDYT) and isolating them from the broiler tendons characterized by arthritis. As suggested by the genome characterization of the 0543/SDYT strains, they belong separately to clusters Ⅰ and Ⅳ. As revealed by sequence comparison, phylogenetic, and recombination analysis, for μA, μB, and σNS genes, considerable genetic divergences were also observed in the two new isolates. However, in the case of λA, λB, μNS, σA, and σB genes, very clear clustering patterns were observed for SDYT and 0543 field strains, respectively. In terms of the μA, μNS, p10, p17 genes of SDYT isolate and μNS, p17, σC, σA genes of 0543 isolate, the lower similarity was observed with NCBI stored sequences, with nt highest identity values below 90 %. In addition, there is an intra-fragmental recombination event in the M1 gene of the SDYT strain. In regard, the multiple segmental recombination and accumulation of point mutations play a role in the newly-emerging ARV strains. Not only did the isolates strain exhibit strong replication ability in vivo , but they also displayed strong arthritogenicity in broilers with low neutralizing maternal antibodies, indicating that maternal antibody treatment may not effectively reduce the oral infection of avian orthoreovirus. These findings suggest that it is necessary to develop a new strategy for enhanced effectiveness in preventing and controlling ARV infection. • We successfully isolated and identified two broiler-derived Avian orthoreovirus strains, namely SDYT and 0543. • Genetic characterization revealed that SDYT was an intra-segmental recombinant strain, classified as genotype IV, and 0543 was a genotype 1 variant, distinct from the S1133 vaccine strain. • We assessed pathogenicity in vivo by challenging S1133, SDYT, 0543 strains in chicks containing maternally derived antibodies (mainly from S11333 vaccinated breeders). [ABSTRACT FROM AUTHOR]

• Using the NGS technology, we identified a recombinant cluster 2 ARV variant named PHC-2020-0545, isolated from tendons of broilers in China. • The isolate was genetically significantly distinct from known ARV strains in M1 and M3 genes. • Multiple segmental recombination, intra-segmental recombination and point mutations might contribute to the emergence of this isolate. • The isolate had a strong replication ability in 1-day-old broilers, and mainly affected the movement, digestion and metabolism of broilers. In mid-2020, using next-generation sequencing (NGS) technology, we identified a recombinant cluster 2 avian orthoreovirus (ARV) variant named PHC-2020−0545, isolated from tendons of 33-day-old broilers with leg swelling in China. Complete genomic sequencing and analyses demonstrated that the isolate was genetically significantly distinct from known ARV strains in M1 and M3 genes and its σC coding gene had an extremely high variability, compared with the identified ARV strains grouped into other genotyping cluster. Further analysis showed that many base substitutions were silent and non-silent substitutions are most likely to occur in the first positions of codons. Multiple segmental recombination, intra-segmental recombination and accumulation of point mutations might contribute to the emergence of this isolate. The PHC-2020−0545 strain had a strong replication ability in 1-day-old broilers, and mainly affected the movement, digestion and metabolism of broilers. In addition, the infection route of the isolate is related to its pathogenicity to broilers. Therefore, combined with its unique genetic characteristics and potential origin, we determined that the PHC-2020−0545 field strain is a novel recombinant ARV strain, which has certain reference value for the preparation and evaluation of new vaccines. [ABSTRACT FROM AUTHOR]

• A model building method was used to simulate the pathogenicity of FAdV-4 and ARV coinfection. • ARV can promote FAdV-4 replication, whereas FAdV-4 has no significant effect on ARV replication. • Compared with singular FAdV-4 infection or singular ARV infection, FAdV-4 and ARV coinfection increased the pathogenicity to SPF chicken embryos and SPF chickens. • Pathogenicity of FAdV-4 and ARV coinfection was closely related to the exact timing and sequence of infection of SPF chicken embryos and SPF chickens with these two pathogens. Hydropericardium hepatitis syndrome (HHS) is a fatal disease caused by fowl adenovirus serotype 4 (FAdV-4). Avian viral arthritis is an infectious disease characterized by movement disorders caused by avian orthoreovirus (ARV). In the early 2019, our epidemiologic survey on poultry diseases in eight commercial broiler farms in China showed that FAdV-4 and ARV have a high coinfection rate, accounting for 63 % of all ARV-positive samples. We designed chicken embryo and animal models to investigate the synergistic pathogenicity of FAdV-4 and ARV. Weakness and inappetence were observed in all specific-pathogen-free (SPF) chickens of the experimental group. FAdV-4 and ARV coinfection caused severe embryonic body and hepatic hemorrhage in SPF chicken embryos. Compared with the singular ARV-infected group, joint swelling was more severe in all coinfected groups. Compared with single virus infection, the coinfection of the two viruses increased the mortality of SPF chicken embryos and chickens. FAdV-4 and ARV coinfection resulted in significantly severe macroscopic and microscopic lesions of the liver, spleen, and kidney of SPF chickens. The detection results of viral load in allantoic fluid, liver, and cloacal swabs indicated that ARV enhanced FAdV-4 replication in SPF chicken embryos and chickens. Cytokine detection showed a significant change in interleukin-1 (IL-1), IL-6, and interferon-α (IFN-α) levels in coinfected groups compared with those in the single-infected groups. Additionally, FAdV-4 and ARV coinfection caused severe damage to the SPF chicken's immune system. In summary, these findings provide insights into the pathology, prevention, and treatment of FAdV-4 and ARV coinfection. [ABSTRACT FROM AUTHOR]