Comparative genomic analysis of five high drug-resistance Aeromonas hydrophila strains induced by doxycycline in laboratory and nine reference strains in Genbank

Aeromonas hydrophila is an opportunistic pathogen and the leading cause of fatal haemorrhagic septicaemia in fish and shellfish. Doxycycline, one of the second generation tetracyclines, has been used in fish farming to fight against infectious diseases caused by A.hydrophila due to its broad-spectrum antimicrobial activity and lower cost. However, progressive increase in resistance of Aeromonas strains to doxycycline aroused serious concern. In this report, drug-resistant A.hydrophilaAH10 strains were induced and selected by using a consecutive batch culture system in Mueller-Hinton Broth (MHB) supplemented with varying concentrations of doxycycline. Five isolates (AH101-105) were obtained from the bacterial culture induced by 25g/ml doxycycline for drug-resistance analysis. Minimal inhibitory concentrations (MIC) values of all five isolates were 50 times higher than that of the parental strain AH10. All of them also displayed high-level resistance to sulphonamides and amides. We sequenced five isolates and performed comparative genomic analysis of these draft genomes with nine A.hydrophila complete genomes from GenBank. Results showed that the pan-genome of 14 strains contains 4,730 genes, 3,056 genes of which present in all strains. The drug-resistance genes also showed significant difference in these genomes, which indicated dangers of indiscriminate use of antibiotics in aquaculture and the necessity of understanding the variation of antibiotic resistance of A.hydrophila. Pan-genome analysis further revealed that no specific SNP (single nucleotide polymorphism) or InDel (insertion and deletion variation) was identified in any functional gene locus among the genomes of AH10 mutated strains, in contrast, significant CNVs (copy number variations) and SV (structure variations) for gene groups were identified in all the mutant genomes.