Objective To investigate antimicrobial susceptibility of Klebsiella pneumoniae (K .pneumoniae )to aminoglycosides and detection of 16S rRNA methylase genes in K .pneumoniae .Methods Ninety-six non-repetitive clinical K .pneumoniae isolates were collected from Xiangya hospital of Central South University from January to Ju-ly 2009,minimal inhibitory concentrations (MICs)of gentamycin,amikacin and tobramycin were determined by agar dilution method ;genotype of 16S rRNA methylase genes (armA,rmtA,rmtB ,rmtC,rmtD ,npmA)were detec-ted by polymerase chain reaction(PCR).Results MIC50 of amikacin,gentamycin and tobramycin was 256μg/mL, 512μg/mL and 512μg/mL respectively;and MIC90 were all>512μg/mL;antimicrobial resistance rate was 21 .88%, 63.54%,and 41 ,67% respectively.68 isolates (70.83%)were resistant to at least one kind of antimicrobial agent, 21 isolates(21 .88%)were resistant to three kinds of antimicrobial agents.22 isolates(22.92%)carried armA,but rmtA,rmtB ,rmtC,rmtD and npmA were not detected;of 22 isolates harboring armA 16S rRNA methylase genes, 17(77.27%)were highly resistant to gentamicin,amikacin and tobramycin,the homology of armA positive isolate and armA (FJ410928.1 )was 100%.Conclusion armA 16S rRNA methylase gene harbored in K .pneumoniae plays an important role in aminoglycoside resistance.