The AsnC family regulator VU14_03665 modulates antibiotic resistance and acid tolerance in Aeromonas hydrophila AH10.

To investigate the biological function of the AsnC family transcriptional regulator VU14_03665 in Aeromonas hydrophila (A. hydrophila), this study used strain AH10 as the research model. A gene deletion mutant (ΔVU14_03665) was constructed via homologous recombination using the recombinant suicide plasmid pCVD442-ΔVU14_03665::Cm, which carried a chloramphenicol resistance marker assembled by fusion PCR. The successful knockout was confirmed by PCR. Phenotypic comparisons between the mutant and wild-type strains included antibiotic susceptibility, acid tolerance, motility, hemolytic activity, growth kinetics, biofilm formation, protease activity, liver colonization in a crucian carp infection model, median lethal dose (LD50), and histopathological changes. Transcriptome sequencing was performed on both strains, and differentially expressed genes were functionally annotated using GO and KEGG analyses.The ΔVU14_03665 strain was successfully generated and confirmed by PCR and sequencing. Phenotypic assays revealed that the mutant exhibited significantly increased sensitivity to six antibiotics (P < 0.05) and markedly reduced acid tolerance (P < 0.01). However, no significant differences were observed in motility, hemolytic activity, growth, biofilm formation, or protease activity compared to the wild-type strain (P > 0.05). In the crucian carp infection model, liver colonization was significantly lower in the mutant (P < 0.05), although LD50 values did not differ significantly. Histopathological examination indicated that infection with ΔVU14_03665 resulted in more severe liver vacuolation, renal hyaline degeneration, and splenic hemosiderin deposition than infection with the wild-type strain.Transcriptome analysis identified 100 differentially expressed genes (33 up-regulated, 67 down-regulated) in the mutant. These genes were primarily associated with biological processes such as organic hydroxy compound metabolism, protein folding, and carbohydrate transport, and were enriched in functions including oxidoreductase activity and metal ion binding. KEGG pathway analysis further revealed significant enrichment in glycerol ester metabolism, selenium compound metabolism, and purine metabolism. Collectively, these results demonstrate that VU14_03665 plays a key role in antibiotic resistance and acid tolerance in A. hydrophila. This study provides foundational insights into the regulatory mechanisms of AsnC family transcription factors in bacterial adaptation and pathogenicity.