ADP-heptose-LPS heptosyltransferase I (WaaC) as a key molecule maintains morphological structure and metabolic...

ADP-heptose-LPS heptosyltransferase I (encoded by waaC gene) is a crucial enzyme in the lipopolysaccharide (LPS) synthesis, maintaining the stability of LPS and cell walls in numerous Gram-negative bacteria. Vibrio mimicus is an epidemic pathogen that threatens aquatic animals and human health, resulting in high morbidity and mortality after infecting the fish. Currently, the role of waaC gene in V. mimicus is still unclear. In this study, waaC gene deletion and complementation strains of V. mimicus were constructed. Our results show that ΔwaaC exhibited a rough phenotype on LB agar, with elevated exocrine protein and a disordered cell wall observed by the electron microscope. Transcriptomic analysis revealed that following the deletion of waaC gene, the expression of 224 genes was drastically upregulated, while the expression of 229 genes was significantly downregulated. These genes involve various pathways, including material transport, metabolism, and environmental adaptation. The tricarboxylic acid cycle (TCA cycle) and pyruvate metabolism are the primary pathways affected by waaC gene. Our phenotypic analysis is consistent with transcriptomic findings, indicating that the decreased pathogenicity of ΔwaaC is related to the effect of waaC gene on these genes, which negatively impacts V. mimicus growth, motility, adhesion, and biofilm formation while enhancing its self-aggregation. The virulence of the ΔwaaC was 103-fold lower than that of the wild strain. Moreover, the expression levels of inflammatory cytokine and pro-apoptotic genes in epithelioma papulosum cyprini (EPC) cells were dramatically upregulated during the ΔwaaC infection. These results provide valuable insights into revealing the pathogenic mechanism of V. mimicus, further bringing more options for the candidate deletion targets of the V. mimicus attenuated vaccine.