Phenotypic differences between GCRV resistant and susceptible grass carp (Ctenopharyngodon idella) based on tlr19 SNP genotypes

Grass carp reovirus (GCRV) infection causes severe hemorrhagic disease in grass carp (Ctenopharyngodon idella), resulting in substantial economic losses in aquaculture. Understanding the genetic basis of resistance is crucial for the selective breeding of robust strains. This study investigated single nucleotide polymorphisms (SNPs) within the tlr19 and their association with GCRV resistance. Phylogenetic analysis revealed that tlr19 is highly conserved in fish and closely related to other Cyprinidae species. Basal expression of tlr19 was highest in the head kidney, and significant upregulation occurred in multiple immune tissues such as head kidney, spleen, hepatopancreas, intestine and gill within 12 h post GCRV infection, indicating its critical role in the early antiviral response. Among three identified SNPs, the +3408 T/A locus showed a significant correlation with resistance. Secondary immersion challenge experiments using homozygous TT (resistant) and AA (susceptible) genotypes at this locus indicated that TT homozygotes had significantly higher survival rates compared to AA homozygotes, suggesting that the +3408 T/A locus could serve as a key genetic marker for GCRV resistance. Molecular and histopathological analyses of both phenotypic groups revealed that resistant grass carp mounted a more robust immune response following GCRV infection, characterized by marked upregulation of antiviral genes such as tlr19, irf3, nf-κb1, and ifn1. Furthermore, resistant individuals displayed enhanced activities of antioxidant enzymes, including catalase and total superoxide dismutase, which facilitated viral clearance and tissue repair. Histopathological examination showed severe hemosiderin deposition, tissue necrosis, and cellular vacuolation in the head kidney and spleen of susceptible fish, whereas tissue damage in resistant fish was notably attenuated. By elucidating the association between the tlr19 + 3408 T/A SNP locus and GCRV resistance phenotypes, this study not only identified a genetic marker applicable to disease-resistant molecular breeding but also revealed the immunological and pathological differences between GCRV-resistant and susceptible populations. These findings provide theoretical insights and molecular marker resources for the development of disease-resistant grass carp strains.