Molecular characterization and phylogenetic analysis of quinolone resistance-determining regions (QRDRs) of gyrA, gyrB, parC and parE gene loci in viridans group streptococci isolated from adult patients with cystic fibrosis

Maeda, Yasunori; Murayama, Mayumi; Goldsmith, Colin E.; Coulter, Wilson A.; Mason, Charlene; Millar, B.C.; Dooley, James; Lowery, Colm; Matsuda, Motoo; Rendall, J.; Elborn, J. Stuart; Moore, John E.

doi:10.1093/jac/dkq485

Cited by 35 publications

(30 citation statements)

References 20 publications

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“…3, we offer a workflow for VGS species identification that can be used in either a research or clinical setting. Importantly, although gyrB encodes part of DNA gyrase, which is a target of fluoroquinolones, polymorphisms in GyrB are not a major mechanism by which VGS strains develop fluoroquinolone resistance (32,36). Therefore, fluoroquinolone resistance does not influence the ability of GyrB to assign VGS strains to a particular species.…”

Section: Discussionmentioning

confidence: 99%

“…The sodA sequence used for phylogenetic analysis was the same sequence fragment used for MLSA (base pairs 46 to 423), as described previously (3). Amplification and sequencing of the gyrB fragment for each of the VGS strains were done using previously published primers and the protocol of Maeda et al (36). The region of gyrB used for phylogenetic analyses and species identification was nucleotides 1113 to 1512, corresponding to amino acids 371 to 503.…”

Section: Methodsmentioning

confidence: 99%

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GyrB Polymorphisms Accurately Assign Invasive Viridans Group Streptococcal Species

et al. 2014

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Viridans group streptococci (VGS) are a heterogeneous group of medically important bacteria that cannot be accurately assigned to a particular species using conventional phenotypic methods. Although multilocus sequence analysis (MLSA) is considered the gold standard for VGS species-level identification, MLSA is not yet feasible in the clinical setting. Conversely, molecular methods, such as sodA and 16S rRNA gene sequencing, are clinically practical but not sufficiently accurate for VGS species-level identification. Here, we present data regarding the use of an ϳ400-nucleotide internal fragment of the gene encoding DNA gyrase subunit B (GyrB) for VGS species-level identification. MLSA, internal gyrB, sodA, full-length, and 5= 16S gene sequences were used to characterize 102 unique VGS blood isolates collected from 2011 to 2012. When using the MLSA species assignment as a reference, full-length and 5= partial 16S gene and sodA sequence analyses failed to correctly assign all strains to a species. Precise species determination was particularly problematic for Streptococcus mitis and Streptococcus oralis isolates. However, the internal gyrB fragment allowed for accurate species designations for all 102 strains. We validated these findings using 54 VGS strains for which MLSA, 16S gene, sodA, and gyrB data are available at the NCBI, showing that gyrB is superior to 16S gene and sodA sequence analyses for VGS species identification. We also observed that specific polymorphisms in the 133-amino acid sequence of the internal GyrB fragment can be used to identify invasive VGS species. Thus, the GyrB amino acid sequence may offer a more practical and accurate method for classifying invasive VGS strains to the species level.V iridans group streptococci (VGS) are a genetically heterogeneous group of commensal bacteria that cause a wide range of infections in humans, particularly in persons with hematologic disease or neutropenia as a result of chemotherapy (1, 2). Although Ͼ25 VGS species have been described, rapid and reliable identification to the species level eludes this group of organisms (3, 4). Accurate species identification is pertinent not only for epidemiological purposes but has potential implications for the clinical management of VGS infections. For example, recent studies show that Streptococcus mitis strains cause a higher incidence of severe clinical disease and have higher rates of resistance to penicillin and fluoroquinolones than do other VGS species (2, 4-6). Additionally, other VGS species have predilections to cause distinct types of infections, such as endocarditis by Streptococcus sanguinis and gastrointestinal infections by Streptococcus anginosus (2, 6).Routine tests and commercial kits used in clinical practice based on the phenotypic and biochemical traits of individual species, such as API Rapid ID32 Strep, Vitek 2, and Streptogram, have only a 30% to 80% accuracy rate for VGS, depending on the species (7-10). The inherent problems with these approaches lie in the variability of traits within V...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

GyrB Polymorphisms Accurately Assign Invasive Viridans Group Streptococcal Species

et al. 2014

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“…Antimicrobial susceptibility was tested by the disk diffusion method according to the Clinical and Laboratory Standards Institute guidelines, and MICs of ciprofloxacin, norfloxacin, cefotaxime, ceftriaxone, and nalidixic acid were determined by Etest strips (AB Biodisk, Solna, Sweden); E. coli ATCC 25922 was used as the quality control organism. Resistance genes gyrA, gyrB, parC, parE, qnr, acc, bla TEM , and bla CTX-M were detected using previously described PCR methods (4,5,8). Bidirectional sequence data were obtained for these resistance and virulence genes.…”

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confidence: 99%

“…In recent decades, an increasing number of E. coli isolates have acquired resistance to quinolones or extended-spectrum cephalosporins (5,13). However, few studies have described E. coli isolates that simultaneously display resistance to these two antimicrobials, which was primarily due to several ESBL-producing resistant genes (3,8). In contrast, the E. coli isolates in this study carried more resistant genes [i.e., two mutations in gyrA (Ser83¡Leu, Asp87¡Asn), two in parC (Ser80¡Ile, Ala108¡Val), one in parE (Ser458¡Ala), and aac(6=)-Ib-cr and bla CTX-M-15 ], which led to high-level resistance to both quinolones and extended-spectrum cephalosporins.…”

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confidence: 99%

Quinolone-Resistant Escherichia coli O127a:K63 Serotype with an Extended-Spectrum-Beta-Lactamase Phenotype from a Food Poisoning Outbreak in China

et al. 2012

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We report an atypical enteropathogenic Escherichia coli O127a:K63 strain with resistance to quinolones and extendedspectrum cephalosporins isolated from a 2010 food poisoning outbreak involving 112 adults in China. Two resistance genes [bla CTX-M-15 , aac(6=)-Ib-c] and five mutations (two in gyrA, two in parC, one in parE) coexisted in this enteropathogenic E. coli strain. Food poisoning from multidrug-resistant strains of diarrheagenic Escherichia coli has received considerable public attention, especially since the large outbreak of hemolytic uremic syndrome caused by E. coli O104:H4 in Germany in 2011 (12). Extended-spectrum cephalosporins and quinolones are widely used for diarrheagenic E. coli, but E. coli isolates are increasingly resistant to these antimicrobials (11). Here, we report the isolation of an E. coli strain harboring virulence genes and genes conferring resistance to quinolones and extended-spectrum cephalosporins from a 2010 food poisoning outbreak involving 112 adult students in China.On 16 September 2010, 112 students (18 to 23 years old) from a university in Zhengzhou, China, experienced diarrhea and other enterogastritis symptoms after eating in the same dining room the previous evening. Seventy-eight (70%) students were hospitalized, and 27 of these students experienced serious symptoms, including stomachache, fever, vomiting, and watery diarrhea (Ͼ5 times/day). Twenty-eight stool specimens were collected from these 27 inpatients within 24 h after admission for laboratory testing. The suspected pathogenic bacteria were isolated through standard procedures on conventional selective media, and the isolated strains were identified using a commercial biochemical test (API 20E system; bioMérieux Vitek, France). Serotyping was carried out by slide agglutination with a conventional E. coli antiserum kit (Denka Seiken, Tokyo, Japan). Sixteen strains of enteropathogenic E. coli serotype O127a:K63 were isolated from the stool specimens. Results of epidemiologic investigation and laboratory tests indicated that this food poisoning outbreak was due to the enteropathogenic E. coli O127a:K63.One 20-year-old male patient had especially severe symptoms, including persistent abdominal cramps with severe paroxysmal pain and abnormal blood test results. Between 16 September and 27 September, he was hospitalized twice for diarrhea. His symptoms were controlled with intravenous administration of levofloxacin and cefpiramide sodium and supportive treatment, and he was released from the hospital. We followed up with this patient for 1 year and found that he often experienced stomachaches, hiccups, nausea, and other symptoms of chronic enterogastritis. Further clinical investigation excluded other chronic or immunerelated diseases (e.g., cancer, hepatitis, and HIV infection); therefore, the chronic enterogastritis appeared to be related to the food poisoning caused by enteropathogenic E. coli.The two suspected isolates obtained from stool specimens during this patient's hospitalizations were identified as enter...

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“…The QRDRs, gyrA, gyrB, parC and parE gene loci, were analysed through sequence typing for all VGS isolates examined (Maeda et al, 2011), as well as carrying out detection and characterization of the macrolide resistance determinants, the ermB and mefA/E gene loci (Tazumi et al, 2009). These methods employed the sequencing of known gene resistance determinants associated with macrolide resistance, as well as fluoroquinolone resistance.…”

Section: Methodsmentioning

confidence: 99%

Comparison of minimum inhibitory concentration by broth microdilution testing versus standard disc diffusion testing in the detection of penicillin, erythromycin and ciprofloxacin resistance in viridans group streptococci

Maeda

Goldsmith

Coulter

et al. 2011

Journal of Medical Microbiology

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The aim of this study was to investigate the reliability of disc diffusion testing with penicillin, erythromycin and ciprofloxacin within the viridans group streptococci (VGS). In total, the antibiotic susceptibilities of 167 VGS isolates were compared by standard disc diffusion and broth microdilution methods, and these phenotypic data were compared to the carriage of the respective gene resistance determinants [ermB and mefA/E (macrolides); QRDR, gyrA, gyrB, parC and parE (quinolones)]. Overall, there were 35 discrepancies [resistant by MIC and susceptible by zone diameter (21.0 %)] between MIC and disc diameter when penicillin susceptibility was interpreted by Clinical and Laboratory Standards Institute criteria. Scattergrams showed a bimodal distribution between non-susceptible and susceptible strains when erythromycin susceptibility was tested by both methods. Thirty-four (20.4 %) isolates were categorized as resistant by MIC breakpoints, while disc diameter defined these as having intermediate resistance. With ciprofloxacin, three isolates (1.8 %) showed minor discrepancies between MIC breakpoints and disc diameter. Isolates nonsusceptible to all three antimicrobial agents tested were reliably distinguished from susceptible isolates by disc diffusion testing, except for the detection of low-level resistance to penicillin, where broth microdilution or an alternative quantitative MIC method should be used. Otherwise, we conclude that disc diffusion testing is a reliable method to detect strains of VGS non-susceptible to penicillin, erythromycin and ciprofloxacin, as demonstrated with their concordance to their gene resistance characteristics.

show abstract

Molecular characterization and phylogenetic analysis of quinolone resistance-determining regions (QRDRs) of gyrA, gyrB, parC and parE gene loci in viridans group streptococci isolated from adult patients with cystic fibrosis

Cited by 35 publications

References 20 publications

GyrB Polymorphisms Accurately Assign Invasive Viridans Group Streptococcal Species

GyrB Polymorphisms Accurately Assign Invasive Viridans Group Streptococcal Species

Quinolone-Resistant Escherichia coli O127a:K63 Serotype with an Extended-Spectrum-Beta-Lactamase Phenotype from a Food Poisoning Outbreak in China

Comparison of minimum inhibitory concentration by broth microdilution testing versus standard disc diffusion testing in the detection of penicillin, erythromycin and ciprofloxacin resistance in viridans group streptococci

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