AbstractThe gram-negative, aerobic, rod-shaped bacteriumAeromonas hydrophila, the causative agent of motile aeromonad septicaemia, has attracted increasing attention due to its high pathogenicity. Here, we constructed the complete genome sequence of a virulent strain,A. hydrophilaHX-3 isolated fromPseudosciaena croceaand performed comparative genomics to investigate its virulence factors and quorum sensing features in comparison with those of otherAeromonasisolates. HX-3 has a circular chromosome of 4,941,513 bp with a 61.0% G + C content encoding 4483 genes, including 4318 protein-coding genes, and 31 rRNA, 127 tRNA and 7 ncRNA operons. Seventy interspersed repeat and 153 tandem repeat sequences, 7 transposons, 8 clustered regularly interspaced short palindromic repeats, and 39 genomic islands were predicted in theA. hydrophilaHX-3 genome. Phylogeny and pan-genome were also analyzed herein to confirm the evolutionary relationships on the basis of comparisons with other fully sequencedAeromonasgenomes. In addition, the assembled HX-3 genome was successfully annotated against the Cluster of Orthologous Groups of proteins database (76.03%), Gene Ontology database (18.13%), and Kyoto Encyclopedia of Genes and Genome pathway database (59.68%). Two-component regulatory systems in the HX-3 genome and virulence factors profiles through comparative analysis were predicted, providing insights into pathogenicity. A large number of genes related to the AHL-type 1 (ahyI,ahyR), LuxS-type 2 (luxS,pfs,metEHK,litR,luxOQU) and QseBC-type 3 (qseB,qseC) autoinducer systems were also identified. As a result of the expression of theahyIgene inEscherichia coliBL21 (DE3), combined UPLC-MS/MS profiling led to the identification of several new N-acyl-homoserine lactone compounds synthesized by AhyI. This genomic analysis determined the comprehensive QS systems ofA. hydrophila, which might provide novel information regarding the mechanisms of virulence signatures correlated with QS.