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Point Mutations at gyrA and gyrB Genes of Levofloxacin-Resistant Helicobacter pylori Isolates in the Esophageal Mucosa from a Venezuelan Population.

ABSTRACT: The treatment of Helicobacter pylori infection is complicated by antibiotic resistance. A high levofloxacin (LVX) resistance rate was previously demonstrated in H. pylori isolates from gastric mucosa (40%) and esophagus (19%) in individual hosts of a Venezuelan population. We aimed to assess the molecular mechanisms of LVX resistance and susceptibility in isolates from the gastroesophageal mucosa, by studying point mutations in the quinolone resistance-determining region of gyrA and gyrB genes. Sequencing of gyrA and gyrB genes (N = 120) helped to identify point mutations in 60 isolates (30 from antrum and 30 from esophagus) of five dyspeptic patients. Double (Asn87Thr and Asp91Asn) and single (Asn87Ile or Asn87Thr) mutations in the gyrA gene were identified in the esophageal mucosa. These mutations have been commonly found in the stomach. Occurrence of a single (Asn87Ile) mutation was associated with high resistance (minimum inhibitory concentration ? 32 ?g/mL) to LVX. Only a single (Ser479Gly) mutation was found in the gyrB gene in both mucosae. One patient presented isolates with no mutations in the two genes studied. Isolates with the same mutation pattern in individual hosts revealed identical genetic profiles for these genes, confirming that isolates identified in the esophageal mucosa come from isolates colonizing the stomach. Helicobacter pylori resistance to LVX in the esophagus is related to double- and single-point mutations in gyrA and gyrB genes, such as those found in the stomach. Levofloxacin should be applied with caution, because its antibiotic effect on H. pylori is decreasing in Latin America, perhaps owing to high prescription rates.

SUBMITTER: Lopez-Gasca M

PROVIDER: S-EPMC5928819 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Point Mutations at gyrA and gyrB Genes of Levofloxacin-Resistant Helicobacter pylori Isolates in the Esophageal Mucosa from a Venezuelan Population.

López-Gasca Mariela M Peña Jessy J García-Amado María-Alexandra MA Michelangeli Fabián F Contreras Monica M

The American journal of tropical medicine and hygiene 20180201 4

The treatment of Helicobacter pylori infection is complicated by antibiotic resistance. A high levofloxacin (LVX) resistance rate was previously demonstrated in H. pylori isolates from gastric mucosa (40%) and esophagus (19%) in individual hosts of a Venezuelan population. We aimed to assess the molecular mechanisms of LVX resistance and susceptibility in isolates from the gastroesophageal mucosa, by studying point mutations in the quinolone resistance-determining region of gyrA ...[more]

PMID: 29405113

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Project description:AimTo investigate the resistance of Helicobacter pylori (H. pylori) to ciprofloxacin (CIP), levofloxacin (LVX) and moxifloxacin (MOX) in the Beijing area and to elucidate the resistance mechanisms.MethodsSeventy-nine H. pylori clinical strains, isolated from patients who had undergone upper gastrointestinal endoscopy in Peking University First Hospital from 2007 to 2009, were tested for their susceptibility to CIP, LVX and MOX using the E-test method. H. pylori strain 26695 was included in the susceptibility testing as a control strain. According to the minimal inhibitory concentration (MIC) values, a strain was classified as resistant to CIP, LVX or MOX when the MIC was > 1 microg/mL. We amplified by polymerase chain reaction (PCR) and sequenced the quinolone resistance-determining regions of the gyrA and gyrB genes from 29 quinolone-resistant and 16 quinolone-susceptible H. pylori strains selected at random.ResultsIn this study, the resistance rates of H. pylori to CIP, LVX or MOX were 55.7% (44/79), and the primary resistance rates were 26.6% (21/79). Patients with secondary resistance had received LVX in previous eradication treatments, but not MOX or CIP. Forty-five strains, including 29 CIP, LVX or MOX-resistant strains (MIC: 1.5-32 microg/mL) and 16 susceptible strains, were selected randomly from the 79 strains and used in PCR analysis. Among these 45 strains, 27 resistant strains had mutations in the gyrA gene, including 11 strains with mutations corresponding to Asp-91 (MIC: 2-32 microg/mL), one of which also had a mutation corresponding to Val-150, and 16 strains had mutations at Asn-87 (MIC: 4-32 microg/mL), three of which also had mutations corresponding to Arg-140 or Val-150. In addition, Arg-140, Val-150 or Ala-97 mutations were separately detected in three susceptible strains. Analysis of the gyrB gene showed that one strain of low resistance had a mutation corresponding to Ser-457 that coexisted with an Asp-91 mutation. There was a significant difference in the occurrence of mutations in the gyrA gene between CIP, LVX and MOX-resistant and -susceptible strains (P < 0.05), but 2 resistant strains were found to possess no quinolone resistance-determining region mutations.ConclusionResistance is primarily mediated through point mutations in gyrA. Whether other mechanisms are responsible for resistance in strains without mutations in the QRDR should be detected.

Project description:IntroductionUnderstanding drug resistance is important in drug selection for Helicobacter pylori (H. pylori) eradication, and drug resistance data are lacking in Beijing.PurposeThis cross-sectional study aimed to isolate H. pylori from patients with gastroduodenal diseases and to analyze drug resistance to clarithromycin (CLA) and levofloxacin (LEV), which are used frequently in China.Patients and methodsOne hundred and seventy-six patients with gastroduodenal diseases undergoing gastroduodenoscopy were selected by convenient sampling. Gastric mucosa samples were cultured and sub-cultured using a new medium broth. Active H. pylori strains were confirmed by microscopy observation as Gram-negative curved bacilli with positive test results for urease, oxidase, and catalase, and H. pylori 16S rRNA amplification by polymerase chain reaction (PCR). CLA and LEV resistance was identified by minimum inhibitory concentration (MIC) tests and sequencing of 23S rRNA, gyrA, and gyrB genes.ResultsFrom the 176 clinical samples, 112 (112/176, 63.6%) were confirmed with H. pylori infection and 65 (65/176, 36.9%) active H. pylori strains were obtained and further confirmed by MIC assay. Overall, the rates of CLA-resistant and LEV-resistant mutations in the 112 samples were 50.9% and 33.0%, respectively. Mutation related to CLA resistance was A2143G in the 23S rRNA gene and mutations associated with LEV resistance were N87K, D91G, and D91Y in the gyrA gene. Of 112 samples, 22 (19.6%) presented dual resistance to CLA and LEV. Resistance of the H. pylori strains to CLA (r=0.846, P<0.001) and LEV (r=0.936, P<0.001) had a strong correlation in phenotypic and genotypic level.ConclusionThe results indicated that resistance of CLA and LEV is severe among patients with gastroduodenitis. A good consistency could be found as to drug resistance between genotypic or phenotypic assay, suggested extending the detection of H. pylori drug resistance from the MIC method to a genotypic assay.

Project description:BackgroundThe prevalence of Helicobacter pylori antibiotic susceptibility in the Nepalese strains is untracked. We determined the antibiotic susceptibility for H. pylori and analyzed the presence of genetic mutations associated with antibiotic resistance in Nepalese strains.ResultsThis study included 146 consecutive patients who underwent gastroduodenal endoscopy in Kathmandu, Nepal. Among 42 isolated H. pylori, there was no resistance to amoxicillin and tetracycline. In contrast, similar with typical South Asian patterns; metronidazole resistance rate in Nepalese strains were extremely high (88.1 %, 37/42). Clarithromycin resistance rate in Nepalese strains were modestly high (21.4 %, 9/42). Most of metronidazole resistant strains had highly distributed rdxA and frxA mutations, but were relative coincidence without a synergistic effect to increase the minimum inhibitory concentration (MIC). Among strains with the high MIC, 63.6 % (7/11) were associated with frameshift mutation at position 18 of frxA with or without rdxA involvement. However, based on next generation sequencing data we found that one strain with the highest MIC value had a novel mutation in the form of amino acid substituted at Ala-212, Gln-382, Ile-485 of dppA and Leu-145, Thr-168, Glu-117, Val-121, Arg-221 in dapF aside from missense mutations in full-length rdxA. Mutations at Asn-87 and/or Asp-91 of the gyrA were predominantly in levofloxacin-resistant strains. The gyrB mutation had steady relationship with the gyrA 87-91 mutations. Although three (44.4 %) and two (22.2 %) of clarithromycin resistant strains had point mutation on A2143G and A2146G, we confirmed the involvement of rpl22 and infB in high MIC strains without an 23SrRNA mutation.ConclusionsThe rates of resistance to clarithromycin, metronidazole and levofloxacin were high in Nepalese strains, indicating that these antibiotics-based triple therapies are not useful as first-line treatment in Nepal. Bismuth or non-bismuth-based quadruple regimens, furazolidone-based triple therapy or rifabutin-based triple therapy may become alternative strategy in Nepal.

Dataset Information

Point Mutations at gyrA and gyrB Genes of Levofloxacin-Resistant Helicobacter pylori Isolates in the Esophageal Mucosa from a Venezuelan Population.

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Point Mutations at <i>gyrA</i> and <i>gyrB</i> Genes of Levofloxacin-Resistant <i>Helicobacter pylori</i> Isolates in the Esophageal Mucosa from a Venezuelan Population.

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