Study On Antibiotic-resistant Genes Of Aminoglyeosides Modifying Enzymes And16S RRNA Methytases Among Gram-negative Bacilli

ObjectiveThis study was conducted to detect the drug susceptibility of aminoglycoside antibiotics ingram-negative bacilli isolated from some hospitals in Changchun and to investigate theoccurrence of both aminoglycoside modifying enzyme genes and16S rRNA methylase genes inclinical isolates in order to provide a basis for the rational use of aminoglycoside antibiotics.Methods1.116non-repetitive clinical isolates of gram-negative bacilli strains were collected fromclinical laboratory of some hospitals in Changchun aera.33strains of Escherichia coli,25strainsof Klebsiella pneumoniae,14strains of Enterobacter cloacae,15strains of Acinetobacterbaumannii,19strains of Pseudomonas aeruginosa and10strains of Serratin marcescens wereincluded. All of the clinical isolates were identified by BioMerieux automatemicrobiological-assay system VITEK-32. Strains were screened by using agar dilution tests andto analyse the susceptibility of amikacin and gentamicin.2. Aminoglycoside modifying enzyme genes (aac(3)-I、aac(3)-II、aac(6’)-Ib、aac(6’)-II、ant(3")-I�'�ant(2")-I) and16S rRNA methylase genes (armA、rmtA、rmtB、rmtC、rmtD andnpmA) in gram-negative bacilli isolates were amplified by PCR.3. The PCR resistance gene fragments were purified and sequenced by automatic sequencer.The sequences acquired were compared with the known sequences in GenBank datebase in orderto determine the gene subtype by BLAST searching.4. Investigate the relation between resistance genes and resistance phenotypes bycomparing116strains of gram-negative bacilli resistant genotypes and phenotypes.Results1. The resistant rates of116strains of gram-negative bacilli to amikacin and gentamicinwere44.0%and75.0%, respectively. Strains resistant to amikacin were resistant to gentamicin.2.81strains of gram-negative bacilli were positive for aminoglycoside modifying enzymegenes with the positive rate of69.8%(81/116). The genotypes included aac(3)-I, aac(3)-II, aac(6’)-Ib, ant(3")-I and ant(2")-I with positive rates of2.6%(3/116),63.8%(74/116),28.4%(33/116),19.8%(23/116) and9.5%(11/116), respectively. But the aac(6’)-Ⅱwas not found. Onlyone strain of116isolates harboured four types of AME genes simutaneously,16strains ofgram-negative bacillus isolates carried three types of AME genes, and28strains were positivefor two types of AME genes.3.46strains of gram-negative bacilli were positive for16S rRNA methylase genes with thepositive rate of39.7%(46/116), including armA gene12.1%(14/116) and rmtB gene31.0%(36/116).4strains of116isolates carried both armA and rmtB genes. But the rmtA, rmtC, rmtDand npmA genes were not found.Conclusions1. High levels of aminoglycoside resistance among gram-negative bacillus isolates werefound in some hospitals of Changchun area.2. Five genotypes of aminoglycoside modifying enzyme genes were detected among116strains of gram-negative bacilli，including aac(3)-II, aac(6’)-Ib, ant(3")-I, ant(2")-I, and aac(3)-Iarranged in order according to the level of detection rate.3. armA and rmtB16S rRNA methylase genes were predominated in local area.