Identification of Aeromonas hydrophila PS-1 and GFP-labeled tracking of its infection in Pelodiscus sinensis

Abstract

The Chinese soft-shelled turtle (Pelodiscus sinensis) is a crucial economically significant aquatic species, yet its aquaculture is severely impacted by disease-related challenges. Among these, Aeromonas hydrophila stands out as one of the most prevalent pathogenic bacteria, responsible for the highest diversity of disease manifestations and detection rates in farmed P. sinensis. In this study, a pathogenic strain A. hydrophila PS-1 from diseased P. sinensis was isolated and identified. A green fluorescent protein (GFP) labeled PS-1 (PS-1/GFP) was constructed with the pAPR-EGFP plasmid. The growth, motility, and pathogenicity of PS-1/GFP showed no difference from that of the wild type strain. After artificial infection, green fluorescence was only observed in the intestine and liver in a paraffin section at 72 h of infection, which indicated that PS-1 infection exhibited tissue specificity. Accordingly, the colonization sites of PS-1 in P. sinensis were intestine and liver, with bacterial loads reaching as high as 5 × 10⁹ CFU/g and 7 × 10⁸ CFU/g, respectively. Moreover, PS-1 could internalize into intestinal fibroblast-like cells and induce similar cell morphological changes akin to pyroptosis. This work established methodological tools and fundamental insights for investigating diseases in P. sinensis, and laid the groundwork for further elucidation of the pathogen-host interaction mechanisms between A. hydrophila and P. sinensis.