Isolation and characterization of an enterovirus G strain causing diarrhea and gut microbiota dysbiosis in piglets

Enterovirus G (EV-G) is a significant causative agent of diarrhea in piglets, complicating differential diagnosis and impeding effective control of porcine diarrheal diseases. Enhanced clinical surveillance and pathogenicity assessment of EVG are essential to inform disease prevention strategies.

The EV-G strain was isolated and identified in MA-104 cells using plaque assay, immunofluorescence staining, viral replication kinetics analysis, and transmission electron microscopy. The complete genomic sequence was obtained for phylogenetic and recombination analysis. Five-day-old piglets were orally challenged with the prevalent EV-G strain, and pathogenicity was assessed based on diarrhea scoring, viral load quantification in feces and tissues, and histopathological examination. Gut microbiota dynamics were profiled via high-throughput 16S rRNA sequencing pre- and post-infection.

A recombinant G1 genotype EV-G strain, designated EVG-SH-2024, was successfully isolated. Phylogenetic analysis indicated its closest amino acid identity (94.7%) with KF985175, while recombination analysis suggested origin from genetic exchange between KF985175 and MN734577. Inoculation with EVG-SH-2024 induced mild diarrhea and persistent viral shedding in piglets, accompanied by significant tissue damage including neuronal vacuolation in the brain, pulmonary consolidation, and intestinal inflammatory infiltration. 16S rRNA sequencing revealed substantial gut microbiota dysbiosis post-infection, marked by an increase in pathogenic bacteria and a decline in beneficial microbes. Notably, the top ten differentially abundant species were predominantly short-chain fatty acid (SCFA)-producing bacteria, whose abundance showed a significant Spearman's rank correlation with pathogenicity indicators.

This study reports the first isolation and characterization of a G1 subtype EV-G strain in Shanghai, China. Integrated pathogenicity and microbiota analyses demonstrated that EV-G infection reduces the abundance of SCFA-producing bacteria, with this reduction significantly correlated with disease severity. These findings suggest a potential role of SCFAs in EV-G pathogenesis and provide a basis for developing microbiota-targeted interventions against EV-G infection.