Rapid Detection of
            <i>aac(6</i>
            ′
            <i>)-Ib-cr</i>
            Quinolone Resistance Gene by Pyrosequencing

Guillard, Thomas; Duval, Véronique; Moret, Hélène; Brasme, Lucien; Vernet-Garnier, Véronique; Champs, Christophe de

doi:10.1128/jcm.01498-09

Cited by 43 publications

(22 citation statements)

References 23 publications

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“…According to the theory of the heterozygote, genetic heterozygote means that different mutations within a single gene locus cause the same phenotypic expression (10). These coexistence of wild-type aac(6')-Ib and aac(6')-Ib-cr variant (at nucleotide 304 by a T→C) alleles in an isolate was a surprisingly common phenomenon, a survey which was very recently supported in the other study (11). Also, these isolates where a proportion of the cells have acquired a cr allele, and under sustained antibiotic exposure these cells would quickly dominate the population (12).…”

Section: Introductionmentioning

confidence: 72%

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

Kim¹,

Jang²,

Kim³

et al. 2011

BMB Reports

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Section: Introductionmentioning

confidence: 72%

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

Kim¹,

Jang²,

Kim³

et al. 2011

BMB Reports

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“…qnrA, qnrB, and qnrS genes were detected using a multiplex PCR method and the primers listed in Table 1 (24). The aac(6=)-Ib-cr gene, which differed from aac(6=)-Ib by two singlenucleotide polymorphisms, namely, T304C/A and G535T, was detected by using a pyrosequencing method and the primers listed in Table 1 (16).…”

Section: Methodsmentioning

confidence: 99%

“…Because pyrosequencing is less labor-and time-intensive than the conventional Sanger method for nucleotide sequence analysis, this method has already been used successfully to identify the resistance-conferring genes of several bacterial species (14)(15)(16)(17). This method appears to be especially suitable as a tool for identifying "hot spot" mutations of QRDRs, namely, at amino acid positions 83 and 87 in GyrA and at positions 80 and 84 in ParC of E. coli.…”

mentioning

confidence: 99%

Molecular Characterization of Extraintestinal Escherichia coli Isolates in Japan: Relationship between Sequence Types and Mutation Patterns of Quinolone Resistance-Determining Regions Analyzed by Pyrosequencing

et al. 2013

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bInfection 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 highthroughput method for analyzing QRDR mutations by pyrosequencing is a powerful tool for epidemiological studies of fluoroquinolone resistance in bacteria.

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“…Detection of aac(6Ј)-Ib-cr has so far depended mainly on genotyping, PCR, and sequencing (2). Recently, simultaneous high-resolution melting analysis and pyrosequencing were developed for detection (1,3). However, these methods are costly and need specialized equipment.…”

mentioning

confidence: 99%

Practical Disk-Based Method for Detection of Escherichia coli Clinical Isolates Producing the Fluoroquinolone-Modifying Enzyme AAC(6′)-Ib-cr

Wachino¹,

Yamane²,

Arakawa

2011

J Clin Microbiol

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Rapid Detection of aac(6 ′ )-Ib-cr Quinolone Resistance Gene by Pyrosequencing

Cited by 43 publications

References 23 publications

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

Molecular Characterization of Extraintestinal Escherichia coli Isolates in Japan: Relationship between Sequence Types and Mutation Patterns of Quinolone Resistance-Determining Regions Analyzed by Pyrosequencing

Practical Disk-Based Method for Detection of Escherichia coli Clinical Isolates Producing the Fluoroquinolone-Modifying Enzyme AAC(6′)-Ib-cr

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