Characterization of antimicrobial resistance in Klebsiella species isolated from chicken broilers

The prevalence of antimicrobial resistant Klebsiella pneumoniae in poultry products has been a public concern, as it severely endangers food safety and human health. In this study, we investigated 90 antimicrobial resistant Klebsiella strains that were isolated from a commercial broiler slaughter plant in Shandong province of China. Nearly all (89/90) of the isolates were identified as infectious phylogenetic group KpI-type K. pneumoniae. Out of these 90 strains, 87 (96.7%) were multidrug-resistant isolates, and 87 (96.7%) were extended-spectrum beta-lactamase (ESBL)-producing isolates. An analysis of the prevalence of quinolone resistance genes showed that 7.8%, 77.8%, 26.7%, and 2.2% of the strains carried the qnrA, qnrB, qnrS, and qepA genes, respectively. An analysis of beta-lactam resistance genes showed that a high percentage of the strains contain the blaTEM (76.7%), blaSHV (88.9%), and blaCTX-M (75.6%) genes, among which three blaSHV subtypes (blaSHV-1, n=30; blaSHV-11, n=38; blaSHV-12, n=12) and three blaCTX-M subtypes (blaCTX-M-14, n=14; blaCTX-M-15, n=35; blaCTX-M-55, n=19) were found. A further investigation of mobile genetic elements involved in horizontal multidrug resistance gene transfer showed the presence of class 1 and 2 integrons in 77 (85.6%) and five (5.6%) isolates, respectively, while no class 3 integrons were detected. Four types of class 1 integrons containing specific gene cassette arrays (dfrA12-orfF-aadA2, dfrA17-aadA5, dfrA1-aadA1, and empty) were identified. Only one gene cassette array (dfrA1-sat2-aadA1) was detected in the class 2 integrons. Furthermore, four different types of insertion sequence common region 1 (ISCR1)-mediated downstream structures were successfully identified in 46 class 1 integron-positive isolates, among which ISCR1-sapA-like-qnrB2-qacEΔ1 was the most commonly observed structure. Chi-square tests revealed a significant association between ESBL genes, plasmid-mediated quinolone resistance (PMQR) genes, and class 1 integrons (p<0.01). Additional conjugation experiments confirmed this relationship (p<0.01) in transconjugants by finding that a high percentage of PMQR genes (74.0%) and class 1 integrons (73.7%) were co-transferred with ESBL genes. Finally, multilocus sequence typing (MLST) was performed, and it revealed that the isolates from chickens are widely distributed in humans, and that antimicrobial resistance is not only disseminated by clonal spreading, but largely by horizontal gene transfer. These results suggest that horizontal transfer of antimicrobial resistance genes by mobile genetic elements, such as integrons, plays a major role in the spread of antimicrobial resistance. Therefore, elucidating the structures of drug resistance integrons is of great importance to the commercial broiler slaughter plant in Shandong, China.