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Molecular characterization of extraintestinal Escherichia coli isolates in Japan: relationship between sequence types and mutation patterns of quinolone resistance-determining regions analyzed by pyrosequencing.
J Clin Microbiol. 2013 Jun; 51(6):1692-8.JC

Abstract

Infection from fluoroquinolone-resistant Enterobacteriaceae is an increasing health problem worldwide. In the present study, we developed a pyrosequencing-based high-throughput method for analyzing the nucleotide sequence of the quinolone resistance-determining regions (QRDRs) of gyrA and parC. By using this method, we successfully determined the QRDR sequences of 139 out of 140 clinical Escherichia coli isolates, 28% of which were nonsusceptible to ciprofloxacin. Sequence results obtained by the pyrosequencing method were in complete agreement with those obtained by the Sanger method. All fluoroquinolone-resistant isolates (n = 35; 25%) contained mutations leading to three or four amino acid substitutions in the QRDRs. In contrast, all isolates lacking a mutation in the QRDR (n = 81; 57%) were susceptible to ciprofloxacin, levofloxacin, and nalidixic acid. The qnr determinants, namely, the qnrA, qnrB, and qnrS genes, were not detected in the isolates, and the aac(6')-Ib-cr gene was detected in 2 (1.4%) of the isolates. Multilocus sequence typing of 34 randomly selected isolates revealed that sequence type 131 (ST131) (n = 7; 20%) is the most prevalent lineage and is significantly resistant to quinolones (P < 0.01). The genetic background of quinolone-susceptible isolates seemed more diverse, and interestingly, neighboring STs of ST131 in the phylogenetic tree were all susceptible to ciprofloxacin. In conclusion, our investigation reveals the relationship between fluoroquinolone resistance caused by mutations of QRDRs and the population structure of clinical extraintestinal E. coli isolates. This high-throughput method for analyzing QRDR mutations by pyrosequencing is a powerful tool for epidemiological studies of fluoroquinolone resistance in bacteria.

Authors+Show Affiliations

Department of Microbiology and Infectious Diseases, Toho University School of Medicine, Tokyo, Japan.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

23515543

Citation

Aoike, Nozomi, et al. "Molecular Characterization of Extraintestinal Escherichia Coli Isolates in Japan: Relationship Between Sequence Types and Mutation Patterns of Quinolone Resistance-determining Regions Analyzed By Pyrosequencing." Journal of Clinical Microbiology, vol. 51, no. 6, 2013, pp. 1692-8.
Aoike N, Saga T, Sakata R, et al. Molecular characterization of extraintestinal Escherichia coli isolates in Japan: relationship between sequence types and mutation patterns of quinolone resistance-determining regions analyzed by pyrosequencing. J Clin Microbiol. 2013;51(6):1692-8.
Aoike, N., Saga, T., Sakata, R., Yoshizumi, A., Kimura, S., Iwata, M., Yoshizawa, S., Sugasawa, Y., Ishii, Y., Yamaguchi, K., & Tateda, K. (2013). Molecular characterization of extraintestinal Escherichia coli isolates in Japan: relationship between sequence types and mutation patterns of quinolone resistance-determining regions analyzed by pyrosequencing. Journal of Clinical Microbiology, 51(6), 1692-8. https://doi.org/10.1128/JCM.03049-12
Aoike N, et al. Molecular Characterization of Extraintestinal Escherichia Coli Isolates in Japan: Relationship Between Sequence Types and Mutation Patterns of Quinolone Resistance-determining Regions Analyzed By Pyrosequencing. J Clin Microbiol. 2013;51(6):1692-8. PubMed PMID: 23515543.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Molecular characterization of extraintestinal Escherichia coli isolates in Japan: relationship between sequence types and mutation patterns of quinolone resistance-determining regions analyzed by pyrosequencing. AU - Aoike,Nozomi, AU - Saga,Tomoo, AU - Sakata,Ryuji, AU - Yoshizumi,Ayumi, AU - Kimura,Soichiro, AU - Iwata,Morihiro, AU - Yoshizawa,Sadako, AU - Sugasawa,Yasuyuki, AU - Ishii,Yoshikazu, AU - Yamaguchi,Keizo, AU - Tateda,Kazuhiro, Y1 - 2013/03/20/ PY - 2013/3/22/entrez PY - 2013/3/22/pubmed PY - 2013/11/19/medline SP - 1692 EP - 8 JF - Journal of clinical microbiology JO - J Clin Microbiol VL - 51 IS - 6 N2 - Infection from fluoroquinolone-resistant Enterobacteriaceae is an increasing health problem worldwide. In the present study, we developed a pyrosequencing-based high-throughput method for analyzing the nucleotide sequence of the quinolone resistance-determining regions (QRDRs) of gyrA and parC. By using this method, we successfully determined the QRDR sequences of 139 out of 140 clinical Escherichia coli isolates, 28% of which were nonsusceptible to ciprofloxacin. Sequence results obtained by the pyrosequencing method were in complete agreement with those obtained by the Sanger method. All fluoroquinolone-resistant isolates (n = 35; 25%) contained mutations leading to three or four amino acid substitutions in the QRDRs. In contrast, all isolates lacking a mutation in the QRDR (n = 81; 57%) were susceptible to ciprofloxacin, levofloxacin, and nalidixic acid. The qnr determinants, namely, the qnrA, qnrB, and qnrS genes, were not detected in the isolates, and the aac(6')-Ib-cr gene was detected in 2 (1.4%) of the isolates. Multilocus sequence typing of 34 randomly selected isolates revealed that sequence type 131 (ST131) (n = 7; 20%) is the most prevalent lineage and is significantly resistant to quinolones (P < 0.01). The genetic background of quinolone-susceptible isolates seemed more diverse, and interestingly, neighboring STs of ST131 in the phylogenetic tree were all susceptible to ciprofloxacin. In conclusion, our investigation reveals the relationship between fluoroquinolone resistance caused by mutations of QRDRs and the population structure of clinical extraintestinal E. coli isolates. This high-throughput method for analyzing QRDR mutations by pyrosequencing is a powerful tool for epidemiological studies of fluoroquinolone resistance in bacteria. SN - 1098-660X UR - https://www.unboundmedicine.com/medline/citation/23515543/Molecular_characterization_of_extraintestinal_Escherichia_coli_isolates_in_Japan:_relationship_between_sequence_types_and_mutation_patterns_of_quinolone_resistance_determining_regions_analyzed_by_pyrosequencing_ L2 - http://jcm.asm.org/cgi/pmidlookup?view=long&amp;pmid=23515543 DB - PRIME DP - Unbound Medicine ER -