Identification of a multiple drug-resistance gene island in the Haemophilus parasuis chromosome

Objectives: The aim of this study was to determine the mechanisms and molecular characterisation of one strain (HPS412) of Haemophilus parasuis, which exhibited high MICs of antimicrobial susceptibility. Methods: A total of 113 H. parasuis strains isolated from pigs suffering from polyserositis, pneumonia or meningitis in China and screened them for antimicrobial susceptibility. Susceptibility testing of the minimal inhibitory concentrations (MIC) was determined in fastidious medium consisting of tryptone soya broth (TSB) containing 5% bovine serum and 10 mu g/mL NAD in 96-well microtiter plates. The genomic DNA was completely sequenced by combining PacBio RS II and Illumina HiSeq 4000 platforms. Gene prediction was performed using Glimmer v.3.02 with Hidden Markov models. Results: One strain (HPS412) exhibited high MICs of sulfamethoxazole (256 mu g/mL), tetracycline (1281 mu g/mL), streptomycin (1281 mu g/mL), gentamicin (1281 mu g/mL), amoxicillin (128 mu g/mL), chloramphenicol (641 mu g/mL), penicillin (641 mu g/mL) and cefaclor (641 mu g/mL). Sequence analysis showed that numerous drug-resistance genes including tet(B), bla(ROB-1), sul2, catIII, aph(3 '')-Ib, aph(6)-Id and aph(3')-la were present in a chromosomal gene island as adjacent duplicate copies and the rep-orf3-bla(ROB-1) structure most likely had a direct plasmid origin. The tet(B) and bla(ROB-1) were flanked on one or both by ISApll elements. Conclusions: The acquisition of bla(ROB-1) and the other antibiotic resistance genes was related to the presence of ISApl1. ISApl1 plays important roles in the horizontal transmission of antibiotic resistance genes. (C) 2020 The Author(s). Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy.