High prevalence of plasmid-mediated 16S rRNA methylase gene rmtB among Escherichia coli clinical isolates from a Chinese teaching hospital.
BACKGROUND
Recently, production of 16S rRNA methylases by Gram-negative bacilli has emerged as a novel mechanism for high-level resistance to aminoglycosides by these organisms in a variety of geographic locations. Therefore, the spread of high-level aminoglycoside resistance determinants has become a great concern.


METHODS
Between January 2006 and July 2008, 680 distinct Escherichia coli clinical isolates were collected from a teaching hospital in Wenzhou, China. PCR and DNA sequencing were used to identify 16S rRNA methylase and extended-spectrum beta-lactamase (ESBL) genes, including armA and rmtB, and in situ hybridization was performed to determine the location of 16S rRNA methylase genes. Conjugation experiments were subsequently performed to determine whether aminoglycoside resistance was transferable from the E. coli isolates via 16S rRNA methylase-bearing plasmids. Homology of the isolates harboring 16S rRNA methylase genes was determined using pulse-field gel electrophoresis (PFGE).


RESULTS
Among the 680 E. coli isolates, 357 (52.5%), 346 (50.9%) and 44 (6.5%) isolates were resistant to gentamicin, tobramycin and amikacin, respectively. Thirty-seven of 44 amikacin-resistant isolates harbored 16S rRNA methylase genes, with 36 of 37 harboring the rmtB gene and only one harboring armA. The positive rates of 16S rRNA methylase genes among all isolates and amikacin-resistant isolates were 5.4% (37/680) and 84.1% (37/44), respectively. Thirty-one isolates harboring 16S rRNA methylase genes also produced ESBLs. In addition, high-level aminoglycoside resistance could be transferred by conjugation from four rmtB-positive donors. The plasmids of incompatibility groups IncF, IncK and IncN were detected in 34, 3 and 3 isolates, respectively. Upstream regions of the armA gene contained ISCR1 and tnpU, the latter a putative transposase gene,. Another putative transposase gene, tnpD, was located within a region downstream of armA. Moreover, a transposon, Tn3, was located upstream of the rmtB. Nineteen clonal patterns were obtained by PFGE, with type H representing the prevailing pattern.


CONCLUSIONS
A high prevalence of plasmid-mediated rmtB gene was found among clinical E. coli isolates from a Chinese teaching hospital. Both horizontal gene transfer and clonal spread were responsible for the dissemination of the rmtB gene.
Yu
Fang-you
FY
Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, Institute of Medical Microbiology and Institutes of Biomedical Sciences, Shanghai Medical School of Fudan University, Shanghai 200032, China.
Yao
Dan
D
Pan
Jing-ye
JY
Chen
Chong
C
Qin
Zhi-qiang
ZQ
Parsons
Chris
C
Yang
Le-he
LH
Li
Qiao-qiao
QQ
Zhang
Xue-qing
XQ
Qu
Di
D
Wang
Liang-xing
LX
D000617:Aminoglycosides / Q000494:pharmacology*; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015373:Bacterial Typing Techniques; D002681:China; D016000:Cluster Analysis; D003227:Conjugation, Genetic; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D024881:Drug Resistance, Bacterial; D016521:Electrophoresis, Gel, Pulsed-Field; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D004927:Escherichia coli Infections / Q000382:microbiology*; D029968:Escherichia coli Proteins / Q000235:genetics*; D022761:Gene Transfer, Horizontal; D006784:Hospitals, Teaching; D006801:Humans; D017403:In Situ Hybridization; D008780:Methyltransferases / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D015995:Prevalence; D017422:Sequence Analysis, DNA; D001618:beta-Lactamases / Q000235:genetics
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D008780:Methyltransferases; C540682:RmtB protein, E coli; C035181:rRNA (adenosine-O-2'-)methyltransferase; D001618:beta-Lactamases