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BackgroundAeromonas hydrophila is a zoonotic and human opportunistic pathogen. Increasing antibiotic resistance profiles have been discovered in both clinical and environmental A. hydrophila isolates in recent years. However, there are still very few in-depth studies regarding the role of plasmids in antibiotic resistance of A. hydrophila. Hence, we investigate the molecular characterization and functional characterization of a multidrug resistant plasmid which encoding a novel NDM variant, AFM-1, in clinical A. hydrophila SS332.MethodsThe minimum inhibitory concentrations (MICs) of 24 antibiotics for A. hydrophila SS332 were measured by the agar dilution method. The genome of A. hydrophila SS332 was sequenced with Pacific and Illumina platforms. Six plasmid-borne antimicrobial resistance genes were chosen for cloning, including blaAFM-1, blaOXA-1, msr(E), mph(E), aac(6')-Ib10, and aph(3')-Ia. Phylogenetic analysis, amino acid sequence alignment, and comparative genomic analysis were performed to elucidate the active site requirements and genetic context of blaAFM-1 gene.ResultsA. hydrophila SS332 showed high levels of resistance to 15 antibiotics, especially those with MIC levels up to or higher than 1024 μg/mL, including ampicillin, cefazolin, ceftriaxone, aztreonam, spectinomycin, roxithromycin. Six plasmid-borne resistance genes were verified to be functional in E. coli DH5α. The blaAFM-1 gene, a novel blaNDM variant, encodes a subclass B1 metallo-β-lactamase AFM-1 which shares 86% amino acid identity with NDM-1. AFM-1 showed resistance to penicillins, cephalosporins and carbapenems. Besides, blaAFM-1 gene was associated with three different novel ISCR19-like elements, designated ISCR19-1, ISCR19-2 and ∆ISCR19-3, which may be involved in the acquisition and mobilization of blaAFM-1 gene. Conclusions Our investigation showed that plasmid-borne resistance genes played a major role in the resistance profiles of A. hydrophila SS332. A novel blaNDM variant, blaAFM-1, was verified to be functional and disseminated among different bacteria species mediated by novel ISCR19-like elements. The fact indicated that the risk of spread of novel resistance genes and novel ISCR elements.

Background: With the wide use of florfenicol to prevent and treat the bacterial infection of domestic animals, the emergence of the florfenicol resistance bacteria is increasingly serious. It is very important to elucidate the molecular mechanism of the bacteria’s resistance to florfenicol.Methods: The minimum inhibitory concentration (MIC) levels were determined by the agar dilution method, and polymerase chain reaction (PCR) was conducted to analyze the distribution of florfenicol resistance genes in 39 CoNS strains isolated from poultry and livestock animals and seafood. The whole genome sequence of one multidrug resistant strain, Staphylococcus lentus H29, was characterized, and comparative genomics analysis of the resistance gene-related sequences was also performed.Results: As a result, the isolates from the animals showed a higher resistance rate (23/28, 82.1%) and much higher MIC levels to florfenicol than those from seafood. Twenty-seven animal isolates carried 37 florfenicol resistance genes (including 26 fexA, 6 cfr and 5 fexB genes) with one carrying a cfr gene, 16 each harboring a fexA gene, 5 with both a fexA and a fexB genes and the other 5 with both a fexA and a cfr genes. On the other hand, all 11 isolates from seafood were sensitive to florfenicol, and only 3 carried a fexA gene each. The whole genome sequence of S. lentus H29 was composed of a chromosome and two plasmids (pH29-46, pH29-26) and harbored 11 resistance genes, including 6 genes [cfr, fexA, ant(6)-Ia, aacA-aphD, mecA and mph(C)] encoded on the chromosome, 4 genes [cfr, fexA, aacA-aphD and tcaA] on pH29-46 and 1 gene (fosD) on pH29-26. We found that the S. lentus H29 genome carried two identical copies of the gene arrays of radC-tnpABC-hp-fexA (5,671 bp) and IS256-cfr (2,690 bp), of which one copy of the two gene arrays was encoded on plasmid pH29-46, while the other was encoded on the chromosome.Conclusions: The current study revealed the wide distribution of florfenicol resistance genes (cfr, fexA and fexB) in animal bacteria, and to the best of our knowledge, this is the first report that one S. lentus strain carried two identical copies of florfenicol resistance-related gene arrays.