|
1
|
Zhang S, Wang X, Wise MJ, He Y, Chen H, Liu A, Huang H, Young S, Tay CY, Marshall BJ, Li X, Chua EG. Mutations of Helicobacter pylori RdxA are mainly related to the phylogenetic origin of the strain and not to metronidazole resistance. J Antimicrob Chemother 2021; 75:3152-3155. [PMID: 32676634 DOI: 10.1093/jac/dkaa302] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2020] [Accepted: 06/09/2020] [Indexed: 12/29/2022] Open
Abstract
OBJECTIVES Drug resistance of Helicobacter pylori is a major clinical problem worldwide. The objective of the present study was to investigate the prevalence of antibiotic-resistant H. pylori in the city of Shenzhen in China, as well as to identify the genetic mutations specifically associated with drug resistance rather than unrelated phylogenetic signals. METHODS Antibiotic susceptibility testing was performed on 238 clinical strains successfully isolated from H. pylori-positive dyspeptic patients who underwent gastroscopy at the Department of Gastroenterology in Shenzhen People's Second Hospital. Following WGS of all strains using Illumina technology, mutation and phylogenetic analyses were performed. RESULTS The resistance rates were 84.9%, 35.3%, 25.2% and 2.1% for metronidazole, clarithromycin, ciprofloxacin and rifampicin, respectively. An A2143G conversion in the 23S rRNA gene was the primary mutation observed in clarithromycin-resistant strains, whilst N87K/I and D91G/N/Y in GyrA were detected in ciprofloxacin-resistant strains. In RdxA, our results demonstrated that only R16H/C and M21A are significant contributors to metronidazole resistance; there were 15 other sites, but these are phylogenetically related and thus unrelated to metronidazole resistance. CONCLUSIONS There is a high prevalence of metronidazole, clarithromycin and ciprofloxacin resistance and a low prevalence of rifampicin resistance in H. pylori from Shenzhen, China. Omission of phylogenetically related sites will help to improve identification of sites genuinely related to antibiotic resistance in H. pylori and, we believe, other species.
Collapse
Affiliation(s)
- Shuzhen Zhang
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China
| | - Xiangyu Wang
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China.,Department of Gastroenterology, The First Affiliated Hospital of Shenzhen University, Shenzhen People's Second Hospital, Shenzhen 518000, China
| | - Michael J Wise
- Department of Computer Science and Software Engineering, University of Western Australia, Perth 6009, Australia.,The Marshall Centre for Infectious Diseases Research and Training, University of Western Australia, Perth 6009, Australia
| | - Yongsheng He
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China
| | - Haiting Chen
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China
| | - Aijun Liu
- Department of Gastroenterology, Kuichong People's Hospital, Shenzhen 518119, China
| | - Haiyan Huang
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China
| | - Sylvia Young
- Harry Perkins Institute of Medical Research, The University of Western Australia, Perth 6009, Australia
| | - Chin Yen Tay
- The Marshall Centre for Infectious Diseases Research and Training, University of Western Australia, Perth 6009, Australia
| | - Barry J Marshall
- The Marshall Centre for Infectious Diseases Research and Training, University of Western Australia, Perth 6009, Australia
| | - Xuehong Li
- Department of Clinical Laboratory, Kuichong People's Hospital, Shenzhen 518119, China
| | - Eng Guan Chua
- The Marshall Centre for Infectious Diseases Research and Training, University of Western Australia, Perth 6009, Australia
| |
Collapse
|
|
2
|
Windham IH, Merrell DS. Analysis of fitness costs associated with metronidazole and amoxicillin resistance in Helicobacter pylori. Helicobacter 2020; 25:e12724. [PMID: 32677105 DOI: 10.1111/hel.12724] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/28/2020] [Revised: 06/17/2020] [Accepted: 06/21/2020] [Indexed: 12/12/2022]
Abstract
BACKGROUND Increasing rates of antibiotic resistance are a major concern for all pathogens, including H. pylori. However, increased resistance often coincides with a decrease in relative fitness of the pathogen in the absence of the antibiotic, raising the possibility that increased resistance can be mitigated for some antibiotics by improved antibiotic husbandry. Therefore, a greater understanding of which types of antibiotic resistance create fitness defects in H. pylori may aid rational treatment strategies. MATERIALS AND METHODS While a wealth of H. pylori literature reports mutations that correlate with increased resistance, few studies demonstrate causation for these same mutations. Herein, we examined fitness costs associated with metronidazole and amoxicillin resistance. Isogenic strains bearing literature reported point mutations in the rdxA and pbp1 genes were engineered and tested in in vitro competition assays to assess relative fitness. RESULTS None of the metronidazole resistance mutations resulted in a fitness cost under the tested conditions. In contrast, amoxicillin-resistant mutant strains demonstrated a defect in competition by 24 hours. This change in fitness was further enhanced by moderate osmotic stress. However, under extreme osmotic stress, the amoxicillin-resistant N562Y PBP1 mutant strain showed enhanced fitness, suggesting that there are some pbp1 mutations that can give a conditional fitness advantage. CONCLUSIONS Our results demonstrate the role of specific point mutations in rdxA and pbp1 in antibiotic resistance and suggest that amoxicillin-resistant strains of H. pylori show environmentally dictated changes in fitness. These later finding may be responsible for the relatively low rates of amoxicillin resistance seen in the United States.
Collapse
Affiliation(s)
- Ian H Windham
- Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, MD, USA
| | - D Scott Merrell
- Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, MD, USA
| |
Collapse
|
|
3
|
Chu A, Wang D, Guo Q, Lv Z, Yuan Y, Gong Y. Molecular detection of
H. pylori
antibiotic‐resistant genes and molecular docking analysis. FASEB J 2019; 34:610-618. [PMID: 31914672 DOI: 10.1096/fj.201900774r] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2019] [Revised: 11/04/2019] [Accepted: 11/04/2019] [Indexed: 12/23/2022]
Affiliation(s)
- Aining Chu
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| | - Dan Wang
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| | - Qianqian Guo
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| | - Zhi Lv
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| | - Yuan Yuan
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| | - Yuehua Gong
- Tumor Etiology and Screening Department of Cancer Institute and General Surgery the First Hospital of China Medical University Shenyang China
- Key Laboratory of Cancer Etiology and Prevention in Liaoning Education Department the First Hospital of China Medical University Shenyang China
- Key Laboratory of GI Cancer Etiology and Prevention in Liaoning Province the First Hospital of China Medical University Shenyang China
| |
Collapse
|
|
4
|
Helicobacter pylori: Multiple resistance in patients from Bogotá, Colombia. BIOMEDICA 2019; 39:125-134. [PMID: 31529855 DOI: 10.7705/biomedica.v39i3.4437] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/01/2018] [Indexed: 12/14/2022]
Abstract
Introduction: The main cause for Helicobacter pylori infection treatment failure is antibiotic resistance, where clarithromycin and metronidazole play the main role. In Colombia, primary resistance as a consequence of the use of these two antibiotics and excessive levofloxacin use is above the accepted limit (13.6%, 83%, and 16%, respectively). Despite this fact, empirical therapies that include the combination of these antibiotics are used in patients with previous therapeutic failure.
Objective: To determine antibiotic resistance in patients previously treated for H. pylori in Bogotá, Colombia.
Materials and methods: We conducted a descriptive study that included ten isolates obtained from five patients with three or four previous failed treatments for H. pylori.
Antibiotic resistance to amoxicillin, clarithromycin, levofloxacin, and metronidazole was investigated by agar dilution and confirmed by DNA sequencing (Magrogen, Korea).
Results: Eight isolates were resistant to two or more antibiotics. All isolates were resistant to levofloxacin. Susceptibility patterns in isolates from the gastric antrum and the body of the stomach were different in three patients.
Conclusion: As far as we know, this is the first evidence of multiple H. pylori resistance in Colombia in previously treated patients. Results demonstrated the consequences of using an ineffective antibiotic scheme and the need to assess antibiotic susceptibility in different anatomical sites of the stomach. The consequences of multiple resistance decrease possible antibiotic effectiveness to eradicate H. pylori in the future.
Collapse
|
|
5
|
Miftahussurur M, Cruz M, Subsomwong P, Jiménez Abreu JA, Hosking C, Nagashima H, Akada J, Yamaoka Y. Clarithromycin-Based Triple Therapy is Still Useful as an Initial Treatment for Helicobacter pylori Infection in the Dominican Republic. Am J Trop Med Hyg 2017; 96:1050-1059. [PMID: 28193745 DOI: 10.4269/ajtmh.16-0729] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
AbstractHelicobacter pylori antibiotic susceptibility in the Dominican Republic has not been monitored. We assessed H. pylori antibiotic susceptibility in the Dominican Republic, and analyzed H. pylori mutations associated with antibiotic resistance. We recruited 158 dyspeptic patients in Santo Domingo and used agar dilution to test susceptibility to five antibiotics. Polymerase chain reaction-based sequencing was used to assess gyrA, gyrB, rdxA, frxA, and 23S rRNA mutations; next-generation sequencing was used to identify other metronidazole resistance-associated genes. Among 64 H. pylori strains isolated, we identified two (3.1%), one (1.6%), and no strains with clarithromycin, amoxicillin, and tetracycline resistance, respectively. Moreover, high frequency of metronidazole resistance (53/64, 82.8%) was observed, whereas levofloxacin resistance is emerging (23/64, 35.9%). We identified many rdxA and frxA mutations in metronidazole-resistant strains, but no synergistic effect was apparent. We revealed novel mutations in dppA, dppB, fdxA, and fdxB, irrespective of rdxA and frxA mutations. Novel mutations at Ser-14 of trx1 and Arg-221 of dapF were associated with different levels of metronidazole resistance. Most levofloxacin-resistant strains had a substitution at Asn-87 of gyrA, including the strain with the highest levofloxacin resistance, whereas only three substitutions were found at Ser-479 of gyrB with no synergistic effect. Besides the 23S rRNA A2142G mutation, we observed another mutation at T1958G in both clarithromycin-resistant strains. We confirmed high metronidazole and levofloxacin resistance associated with genetic mutations in the Dominican Republic. However, prevalence of clarithromycin resistance was low, suggesting that standard clarithromycin-based triple therapy remains useful as initial treatment of H. pylori infection.
Collapse
Affiliation(s)
- Muhammad Miftahussurur
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan.,Department of Medicine, Gastroenterology and Hepatology Section, Baylor College of Medicine, Houston, Texas.,Gastroentero-Hepatology Division, Department of Internal Medicine, Faculty of Medicine-Dr. Soetomo Teaching Hospital-Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Modesto Cruz
- Institute of Microbiology and Parasitology, Faculty of Science, Autonomous University of Santo Domingo, Santo Domingo, Dominican Republic.,Department of Biomedical Research, National Institute of Medicine and Diagnostic Imaging, Santo Domingo, Dominican Republic
| | - Phawinee Subsomwong
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - José A Jiménez Abreu
- Dominican-Japanese Digestive Disease Center, Dr. Luis E. Aybar Health and Hygiene City, Santo Domingo, Dominican Republic
| | - Celso Hosking
- Institute of Microbiology and Parasitology, Faculty of Science, Autonomous University of Santo Domingo, Santo Domingo, Dominican Republic
| | - Hiroyuki Nagashima
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - Junko Akada
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - Yoshio Yamaoka
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan.,Department of Medicine, Gastroenterology and Hepatology Section, Baylor College of Medicine, Houston, Texas
| |
Collapse
|
|
6
|
Miftahussurur M, Syam AF, Nusi IA, Makmun D, Waskito LA, Zein LH, Akil F, Uwan WB, Simanjuntak D, Wibawa IDN, Waleleng JB, Saudale AMJ, Yusuf F, Mustika S, Adi P, Maimunah U, Maulahela H, Rezkitha YAA, Subsomwong P, Nasronudin, Rahardjo D, Suzuki R, Akada J, Yamaoka Y. Surveillance of Helicobacter pylori Antibiotic Susceptibility in Indonesia: Different Resistance Types among Regions and with Novel Genetic Mutations. PLoS One 2016; 11:e0166199. [PMID: 27906990 PMCID: PMC5131997 DOI: 10.1371/journal.pone.0166199] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2016] [Accepted: 10/24/2016] [Indexed: 02/05/2023] Open
Abstract
Information regarding Helicobacter pylori antibiotic resistance in Indonesia was previously inadequate. We assessed antibiotic susceptibility for H. pylori in Indonesia, and determined the association between virulence genes or genetic mutations and antibiotic resistance. We recruited 849 dyspeptic patients who underwent endoscopy in 11 cities in Indonesia. E-test was used to determine the minimum inhibitory concentration of five antibiotics. PCR-based sequencing assessed mutations in 23S rRNA, rdxA, gyrA, gyrB, and virulence genes. Next generation sequencing was used to obtain full-length sequences of 23S rRNA, infB, and rpl22. We cultured 77 strains and identified 9.1% with clarithromycin resistance. Low prevalence was also found for amoxicillin and tetracycline resistance (5.2% and 2.6%, respectively). In contrast, high resistance rates to metronidazole (46.7%) and levofloxacin (31.2%) were demonstrated. Strains isolated from Sumatera Island had significantly higher metronidazole resistance than those from other locations. Metronidazole resistant strains had highly distributed rdxA amino acid substitutions and the 23S rRNA A2143G mutation was associated with clarithromycin resistance (42.9%). However, one strain with the highest MIC value had a novel mutation in rpl22 without an A2143G mutation. Mutation at Asn-87 and/or Asp-91 of gyrA was associated with levofloxacin-resistance and was related to gyrB mutations. In conclusions, although this is a pilot study for a larger survey, our current data show that Indonesian strains had the high prevalence of metronidazole and levofloxacin resistance with low prevalence of clarithromycin, amoxicillin, and tetracycline resistance. Nevertheless, clarithromycin- or metronidazole-based triple therapy should be administered with caution in some regions of Indonesia.
Collapse
Affiliation(s)
- Muhammad Miftahussurur
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
- Department of Medicine, Gastroenterology and Hepatology Section, Baylor College of Medicine, Houston, Texas, United States of America
- Division of Gastroentero-Hepatology, Department of Internal Medicine, Universitas Airlangga Faculty of Medicine, Surabaya, Indonesia
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Ari Fahrial Syam
- Division of Gastroenterology, Department of Internal Medicine, Faculty of Medicine, University of Indonesia, Jakarta, Indonesia
| | - Iswan Abbas Nusi
- Division of Gastroentero-Hepatology, Department of Internal Medicine, Universitas Airlangga Faculty of Medicine, Surabaya, Indonesia
| | - Dadang Makmun
- Division of Gastroenterology, Department of Internal Medicine, Faculty of Medicine, University of Indonesia, Jakarta, Indonesia
| | - Langgeng Agung Waskito
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Lukman Hakim Zein
- Division of Gastroentero-Hepatology, Department of Internal Medicine, Faculty of Medicine, University of Sumatera Utara, Medan, Indonesia
| | - Fardah Akil
- Center of Gastroentero-Hepatology, Department of Internal Medicine, Faculty of Medicine, Hasanuddin University, Makassar, Indonesia
| | - Willy Brodus Uwan
- Department of Internal Medicine, Santo Antonius Hospital, Pontianak, Indonesia
| | - David Simanjuntak
- Department of Internal Medicine, Yowari Hospital, Jayapura, Indonesia
| | - I Dewa Nyoman Wibawa
- Division of Gastroentero-hepatology, Department of Internal Medicine, Faculty of Medicine University of Udayana, Denpasar, Indonesia
| | - Jimmy Bradley Waleleng
- Division of Gastroentero-hepatology, Department of Internal Medicine, Faculty of Medicine, University of Sam Ratulangi, Prof. Dr. RD Kandou Hospital, Manado, Indonesia
| | | | - Fauzi Yusuf
- Division of Gastroentero-hepatology, Department of Internal Medicine, Dr. Zainoel Abidin General Hospital, Banda Aceh, Indonesia
| | - Syifa Mustika
- Division of Gastroentero-hepatology, Department of Internal Medicine, Dr. Saiful Anwar General Hospital, Malang, Indonesia
| | - Pangestu Adi
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Ummi Maimunah
- Division of Gastroentero-Hepatology, Department of Internal Medicine, Universitas Airlangga Faculty of Medicine, Surabaya, Indonesia
| | - Hasan Maulahela
- Division of Gastroenterology, Department of Internal Medicine, Faculty of Medicine, University of Indonesia, Jakarta, Indonesia
| | | | - Phawinee Subsomwong
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - Nasronudin
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Dadik Rahardjo
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
| | - Rumiko Suzuki
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - Junko Akada
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
| | - Yoshio Yamaoka
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu, Japan
- Department of Medicine, Gastroenterology and Hepatology Section, Baylor College of Medicine, Houston, Texas, United States of America
| |
Collapse
|
|
7
|
Miftahussurur M, Shrestha PK, Subsomwong P, Sharma RP, Yamaoka Y. Emerging Helicobacter pylori levofloxacin resistance and novel genetic mutation in Nepal. BMC Microbiol 2016; 16:256. [PMID: 27809767 PMCID: PMC5096319 DOI: 10.1186/s12866-016-0873-6] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2016] [Accepted: 10/27/2016] [Indexed: 01/06/2023] Open
Abstract
Background The 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. Results This 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. Conclusions The 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. Electronic supplementary material The online version of this article (doi:10.1186/s12866-016-0873-6) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Muhammad Miftahussurur
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, 1-1 Idaigaoka, Hasama-machi, Yufu-City, Oita, 879-5593, Japan.,Department of Gastroenterology and Hepatology, Baylor College of Medicine and Michael DeBakey Veterans Affairs Medical Center, Houston, TX, 77030, USA.,Gastroentero-Hepatology Division, Department of Internal Medicine, Faculty of Medicine-Institute of Tropical Disease, Universitas Airlangga, Surabaya, 60115, Indonesia
| | - Pradeep Krishna Shrestha
- Gastroenterology Department, Maharajgunj Medical Campus, Tribhuvan University Teaching Hospital, Kathmandu, 44600, Nepal
| | - Phawinee Subsomwong
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, 1-1 Idaigaoka, Hasama-machi, Yufu-City, Oita, 879-5593, Japan
| | - Rabi Prakash Sharma
- Gastroenterology Department, Maharajgunj Medical Campus, Tribhuvan University Teaching Hospital, Kathmandu, 44600, Nepal
| | - Yoshio Yamaoka
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, 1-1 Idaigaoka, Hasama-machi, Yufu-City, Oita, 879-5593, Japan. .,Department of Gastroenterology and Hepatology, Baylor College of Medicine and Michael DeBakey Veterans Affairs Medical Center, Houston, TX, 77030, USA.
| |
Collapse
|
|
8
|
Hu Y, Zhang M, Lu B, Dai J. Helicobacter pylori and Antibiotic Resistance, A Continuing and Intractable Problem. Helicobacter 2016; 21:349-63. [PMID: 26822340 DOI: 10.1111/hel.12299] [Citation(s) in RCA: 102] [Impact Index Per Article: 11.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Helicobacter pylori, a human pathogen with a high global prevalence, is the causative pathogen for multiple gastrointestinal diseases, especially chronic gastritis, peptic ulcers, gastric mucosa-associated lymphoid tissue lymphoma, and gastric malignancies. Antibiotic therapies remain the mainstay for H. pylori eradication; however, this strategy is hampered by the emergence and spread of H. pylori antibiotic resistance. Exploring the mechanistic basis of this resistance is becoming one of the major research questions in contemporary biomedical research, as such knowledge could be exploited to devise novel rational avenues for counteracting the existing resistance and devising strategies to avoid the development of a novel anti-H. pylori medication. Encouragingly, important progress in this field has been made recently. Here, we attempt to review the current state and progress with respect to the molecular mechanism of antibiotic resistance for H. pylori. A picture is emerging in which mutations of various genes in H. pylori, resulting in decreased membrane permeability, altered oxidation-reduction potential, and a more efficient efflux pump system. The increased knowledge on these mechanisms produces hope that antibiotic resistance in H. pylori can ultimately be countered.
Collapse
Affiliation(s)
- Yue Hu
- Department of Gastroenterology, First Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Meng Zhang
- Department of Gastroenterology, First Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Bin Lu
- Department of Gastroenterology, First Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, China.
| | - Jinfeng Dai
- Department of Gastroenterology, First Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| |
Collapse
|
|
9
|
Search for novel candidate mutations for metronidazole resistance in Helicobacter pylori using next-generation sequencing. Antimicrob Agents Chemother 2015; 59:2343-8. [PMID: 25645832 DOI: 10.1128/aac.04852-14] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023] Open
Abstract
Metronidazole resistance is a key factor associated with Helicobacter pylori treatment failure. Although this resistance is mainly associated with mutations in the rdxA and frxA genes, the question of whether metronidazole resistance is caused by the inactivation of frxA alone is still debated. Furthermore, it is unclear whether there are other mutations involved in addition to the two genes that are associated with resistance. A metronidazole-resistant strain was cultured from the metronidazole-susceptible H. pylori strain 26695-1 by exposure to low concentrations of metronidazole. The genome sequences of both susceptible and resistant H. pylori strains were determined by Illumina next-generation sequencing, from which putative candidate resistance mutations were identified. Natural transformation was used to introduce PCR products containing candidate mutations into the susceptible parent strain 26695-1, and the metronidazole MIC was determined for each strain. Mutations in frxA (hp0642), rdxA (hp0954), and rpsU (hp0562) were confirmed by the Sanger method. The mutated sequence in rdxA was successfully transformed into strain 26695-1, and the transformants showed resistance to metronidazole. The transformants containing a single mutation in rdxA showed a low MIC (16 mg/liter), while those containing mutations in both rdxA and frxA showed a higher MIC (48 mg/liter). No transformants containing a single mutation in frxA or rpsU were obtained. Next-generation sequencing was used to identify mutations related to drug resistance. We confirmed that the mutations in rdxA are mainly associated with metronidazole resistance, and mutations in frxA are able to enhance H. pylori resistance only in the presence of rdxA mutations. Moreover, mutations in rpsU may play a role in metronidazole resistance.
Collapse
|
|
10
|
Mirzaei N, Poursina F, Moghim S, Rahimi E, Safaei HG. The mutation of the rdxA gene in metronidazole-resistant Helicobacter pylori clinical isolates. Adv Biomed Res 2014; 3:90. [PMID: 24761398 PMCID: PMC3988589 DOI: 10.4103/2277-9175.128469] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2013] [Accepted: 08/06/2013] [Indexed: 01/15/2023] Open
Abstract
BACKGROUNDS Antibiotic resistance is an increasing problem throughout the developed world, and knowledge about different resistance mechanisms is consequential for efficient treatment of bacterial infections. Although metronidazole has been frequently used in treatment regimens for H. pylori infection, but antibiotic resistance is now a major contributing factor in treatment failure. Nevertheless metronidazole has been greatly used as a critical component of combination therapies for H. pylori infection. OBJECTIVE This study is trying to describe the mutational mechanisms of metronidazole resistance in H. pylori in our clinical isolates in Isfahanian patients, Iran and compare with the findings of previous studies in world. MATERIALS AND METHODS MIC values of metronidazole for H. pylori strains were determined by E- test. Both rdxA and glmM genes used for confirmation of isolates as H. pylori and then amplification of another rdxA oligonucleotide pair was done. Finally, the six resistant strains were sent to sequencing for other processing and further analysis was done by software. RESULTS The result of six clinical isolates in comparison with 26695, J99 and 69A as a sensitive and resistant reference strains showed plenty of mutations. No frame shift and nonsense mutation was seen in our clinical isolates. CONCLUSION An interesting finding in metronidazole-resistant strains in our study was the detection of one mutation not previously described in the literature in the rdxA gene and this W(209)R substitution presumably plays a role in inducing metronidazole resistance.
Collapse
Affiliation(s)
- Nasrin Mirzaei
- Department of Biology, Tonekabon Branch, Islamic Azad University, Tonekabon, Iran
| | - Farkhondeh Poursina
- Department of Microbiology, Isfahan University of Medical Sciences, Isfahan, Iran
| | - Sharareh Moghim
- Department of Microbiology, Isfahan University of Medical Sciences, Isfahan, Iran
| | - Ebrahim Rahimi
- Department of Food Hygiene, Azad University of Shahrekord, Shahrekord, Iran
| | | |
Collapse
|
|
11
|
Mejía-Jaramillo AM, Fernández GJ, Palacio L, Triana-Chávez O. Gene expression study using real-time PCR identifies an NTR gene as a major marker of resistance to benzonidazole in Trypanosoma cruzi. Parasit Vectors 2011; 4:169. [PMID: 21892937 PMCID: PMC3185274 DOI: 10.1186/1756-3305-4-169] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2011] [Accepted: 09/05/2011] [Indexed: 01/10/2023] Open
Abstract
Background Chagas disease is a neglected illness, with limited treatments, caused by the parasite Trypanosoma cruzi. Two drugs are prescribed to treat the disease, nifurtimox and benznidazole, which have been previously reported to have limited efficacy and the appearance of resistance by T. cruzi. Acquisition of drug-resistant phenotypes is a complex physiological process based on single or multiple changes of the genes involved, probably in its mechanisms of action. Results The differential genes expression of a sensitive Trypanosoma cruzi strain and its induced in vitro benznidazole-resistant phenotypes was studied. The stepwise increasing concentration of BZ in the parental strain generated five different resistant populations assessed by the IC50 ranging from 10.49 to 93.7 μM. The resistant populations maintained their phenotype when the BZ was depleted from the culture for many passages. Additionally, the benznidazole-resistant phenotypes presented a cross-resistance to nifurtimox but not to G418 sulfate. On the other hand, four of the five phenotypes resistant to different concentrations of drugs had different expression levels for the 12 genes evaluated by real-time PCR. However, in the most resistant phenotype (TcR5x), the levels of mRNA from these 12 genes and seven more were similar to the parental strain but not for NTR and OYE genes, which were down-regulated and over-expressed, respectively. The number of copies for these two genes was evaluated for the parental strain and the TcR5x phenotype, revealing that the NTR gene had lost a copy in this last phenotype. No changes were found in the enzyme activity of CPR and SOD in the most resistant population. Finally, there was no variability of genetic profiles among all the parasite populations evaluated by performing low-stringency single-specific primer PCR (LSSP-PCR) and random amplified polymorphic DNA RAPD techniques, indicating that no clonal selection or drastic genetic changes had occurred for the exposure to BZ. Conclusion Here, we propose NTR as the major marker of the appearance of resistance to BZ.
Collapse
Affiliation(s)
- Ana M Mejía-Jaramillo
- Grupo Biología y Control de Enfermedades Infecciosas-BCEI-SIU, Instituto de Biología, Universidad de Antioquia, Medellín, Colombia
| | | | | | | |
Collapse
|
|
12
|
Francesco VD, Zullo A, Hassan C, Giorgio F, Rosania R, Ierardi E. Mechanisms of Helicobacter pylori antibiotic resistance: An updated appraisal. World J Gastrointest Pathophysiol 2011; 2:35-41. [PMID: 21860834 PMCID: PMC3158889 DOI: 10.4291/wjgp.v2.i3.35] [Citation(s) in RCA: 100] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/18/2011] [Revised: 05/29/2011] [Accepted: 06/05/2011] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori (H. pylori) antibiotic resistance is the main factor affecting the efficacy of the current eradicating therapies. The aim of this editorial is to report on the recent information about the mechanisms accounting for the resistance to the different antibiotics currently utilized in H. pylori eradicating treatments. Different mechanisms of resistance to clarithromycin, metronidazole, quinolones, amoxicillin and tetracycline are accurately detailed (point mutations, redox intracellular potential, pump efflux systems, membrane permeability) on the basis of the most recent data available from the literature. The next hope for the future is that by improving the knowledge of resistance mechanisms, the elaboration of rational and efficacious associations for the treatment of the infection will be possible. Another auspicious progress might be the possibility of a cheap, feasible and reliable laboratory test to predict the outcome of a therapeutic scheme.
Collapse
|
|
13
|
Functional analysis of the RdxA and RdxB nitroreductases of Campylobacter jejuni reveals that mutations in rdxA confer metronidazole resistance. J Bacteriol 2010; 192:1890-901. [PMID: 20118248 DOI: 10.1128/jb.01638-09] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Campylobacter jejuni is a leading cause of gastroenteritis in humans and a commensal bacterium of the intestinal tracts of many wild and agriculturally significant animals. We identified and characterized a locus, which we annotated as rdxAB, encoding two nitroreductases. RdxA was found to be responsible for sensitivity to metronidazole (Mtz), a common therapeutic agent for another epsilonproteobacterium, Helicobacter pylori. Multiple, independently derived mutations in rdxA but not rdxB resulted in resistance to Mtz (Mtz(r)), suggesting that, unlike the case in H. pylori, Mtz(r) might not be a polygenic trait. Similarly, Mtz(r) C. jejuni was isolated after both in vitro and in vivo growth in the absence of selection that contained frameshift, point, insertion, or deletion mutations within rdxA, possibly revealing genetic variability of this trait in C. jejuni due to spontaneous DNA replication errors occurring during normal growth of the bacterium. Similar to previous findings with H. pylori RdxA, biochemical analysis of C. jejuni RdxA showed strong oxidase activity, with reduction of Mtz occurring only under anaerobic conditions. RdxB showed similar characteristics but at levels lower than those for RdxA. Genetic analysis confirmed that rdxA and rdxB are cotranscribed and induced during in vivo growth in the chick intestinal tract, but an absence of these genes did not strongly impair C. jejuni for commensal colonization. Further studies indicate that rdxA is a convenient locus for complementation of mutants in cis. Our work contributes to the growing knowledge of determinants contributing to susceptibility to Mtz (Mtz(s)) and supports previous observations of the fundamental differences in the activities of nitroreductases from epsilonproteobacteria.
Collapse
|
|
14
|
Jones KR, Cha JH, Merrell DS. Who's Winning the War? Molecular Mechanisms of Antibiotic Resistance in Helicobacter pylori. CURRENT DRUG THERAPY 2008; 3:190-203. [PMID: 21765819 PMCID: PMC3136193 DOI: 10.2174/157488508785747899] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
The ability of clinicians to wage an effective war against many bacterial infections is increasingly being hampered by skyrocketing rates of antibiotic resistance. Indeed, antibiotic resistance is a significant problem for treatment of diseases caused by virtually all known infectious bacteria. The gastric pathogen Helicobacter pylori is no exception to this rule. With more than 50% of the world's population infected, H. pylori exacts a tremendous medical burden and represents an interesting paradigm for cancer development; it is the only bacterium that is currently recognized as a carcinogen. It is now firmly established that H. pylori infection is associated with diseases such as gastritis, peptic and duodenal ulceration and two forms of gastric cancer, gastric adenocarcinoma and mucosa-associated lymphoid tissue (MALT) lymphoma. With such a large percentage of the population infected, increasing rates of antibiotic resistance are particularly vexing for a treatment regime that is already fairly complicated; treatment consists of two antibiotics and a proton pump inhibitor. To date, resistance has been found to all primary and secondary lines of antibiotic treatment as well as to drugs used for rescue therapy.
Collapse
Affiliation(s)
- Kathleen R. Jones
- Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Rd., Bethesda, MD 20814, USA
| | - Jeong-Heon Cha
- Department of Oral Biology, Oral Science Research Center, BK21 project, Yonsei University College of Dentistry, Seoul, Korea
| | - D. Scott Merrell
- Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Rd., Bethesda, MD 20814, USA
| |
Collapse
|
|
15
|
Lee YC, Lee SY, Pyo JH, Kwon DH, Rhee JC, Kim JJ. Isogenic variation of Helicobacter pylori strain resulting in heteroresistant antibacterial phenotypes in a single host in vivo. Helicobacter 2005; 10:240-8. [PMID: 15904482 DOI: 10.1111/j.1523-5378.2005.00316.x] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
BACKGROUND Antibiotic-susceptible and -resistant Helicobacter pylori can be present simultaneously in the same host. The aim of this study was to evaluate the genomic diversity of H. pylori strains resulting in heteroresistant antibacterial phenotypes. MATERIALS AND METHODS Twenty-one pairs of H. pylori strains isolated from the antrum and body displaying heteroresistant antibacterial phenotypes were included. We compared the genotypes of paired-isolates by random arbitrarily primed polymerase chain reaction (PCR), flagella gene PCR-based restriction fragment length polymorphism, and flaA gene sequencing. In metronidazole-heteroresistant isolates, the sequence variation of rdxA and frxA genes was analyzed using phylogenetic analysis. RESULTS The DNA fingerprinting patterns of the paired isolates revealed that 12 pairs (57.1%) were identical, whereas one pair (3.8%) was different. The remaining eight pairs (38.1%) of isolates showed minor heterogenecity in fingerprinting patterns. In flaA gene sequencing, these identical and similar isolates showed close sequence similarity between the antrum and body, whereas different isolate showed 31 points of different nucleotide sequences. Phylogenetic analysis of the metronidazole-heteroresistant pairs showed consistent genetic relatedness of each paired isolates despite the sequence variation of the rdxA or frxA genes in five pairs (71.4%). CONCLUSIONS These results suggest that continuing genomic diversities in the same strain may play an important role in modulating the antibiotic-heteroresistant H. pylori in vivo.
Collapse
Affiliation(s)
- Yong Chan Lee
- Department of Internal Medicine and Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul, Korea
| | | | | | | | | | | |
Collapse
|
|
16
|
Simala-Grant JL, Taylor DE. Molecular biology methods for the characterization of Helicobacter pylori infections and their diagnosis. APMIS 2005; 112:886-97. [PMID: 15688524 DOI: 10.1111/j.1600-0463.2004.apm11211-1211.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Helicobacter pylori infects approximately half of the human population; however, the outcome of infection is affected by many factors, including strain and host genotype characteristics and bacterial density within the stomach. Many molecular methods have been developed to provide information with respect to these characteristics. Methods that provide results within 24 h of endoscopy may be used to develop individualized treatment that is more effective, results in fewer side effects, cuts costs,decreases the number of treatment failures and results in the development of fewer antibiotic-resistant H. pylori strains.
Collapse
Affiliation(s)
- Joanne L Simala-Grant
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Canada
| | | |
Collapse
|
|
17
|
Aldana LP, Kato M, Kondo T, Nakagawa S, Zheng R, Sugiyama T, Asaka M, Kwon DH. In vitro induction of resistance to metronidazole, and analysis of mutations in rdxA and frxA genes from Helicobacter pylori isolates. J Infect Chemother 2005; 11:59-63. [PMID: 15856372 DOI: 10.1007/s10156-004-0370-y] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2004] [Accepted: 12/28/2004] [Indexed: 10/25/2022]
Abstract
In clinical Helicobacter pylori isolates, metronidazole resistance has been associated with mutations in the rdxA and frxA genes. The aim of this study was to examine the role of the rdxA and frxA genes after the in vitro induction of metronidazole resistance. A total of five suscep-tible H. pylori isolates were initially exposed to different subinhibitory metronidazole concentrations to induce in vitro resistance to metronidazole. Susceptible and resistant strains after the in vitro induction of resistance were examined to evaluate mutations of the rdxA and frxA genes by sequence analysis. After the in vitro induction of resistance, analysis revealed that two and four susceptible strains developed resistance when cultured with 0.3 microg/ml and 0.6 microg/ml of metronidazole, respectively. Before and after the induction of resistance, none of the susceptible strains that developed low and moderate levels of resistance presented any mutation in either of the evaluated genes, whereas strains with high-level metronidazole resistance contained a simple mutation of the frxA gene, but no specific changes in the rdxA gene. Strains with moderate-level resistance contained both single and multiple mutations of rdxA and frxA, respectively, and the low-level-metronidazole-resistant strain contained a single mutation in the frxA gene, without any significant change in the rdxA gene. In this study, the strains that developed resistance were mainly associated with mutations of the frxA gene, suggesting the possibility that inactivation of this gene could originate metronidazole resistance. The results after the in vitro induction of resistance to metronidazole suggested the presence of additional metronidazole resistance mechanisms, other than mutations of the rdxA and/or frxA genes.
Collapse
Affiliation(s)
- Luis Perez Aldana
- Department of Gastroenterology, Hokkaido University Graduate School of Medicine, Sapporo, Japan
| | | | | | | | | | | | | | | |
Collapse
|
|
18
|
Hiyama T, Tanaka S, Masuda H, Shima H, Kose K, Tuncel H, Ito M, Kitadai Y, Sumii M, Uemura N, Yoshihara M, Shimamoto F, Haruma K, Chayama K. Prevalence of Helicobacter pylori resistance to clarithromycin and metronidazole determined by 23S ribosomal RNA and rdxA gene analyses in Hiroshima, Japan. J Gastroenterol Hepatol 2003; 18:1202-1207. [PMID: 12974909 DOI: 10.1046/j.1440-1746.2003.03140.x] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
BACKGROUND AND AIMS Resistance to antibiotics in Helicobacter pylori is increasing and becoming a serious problem in eradication treatment of H. pylori. The prevalence of H. pylori infections that are resistant to clarithromycin, metronidazole, or both were determined in H. pylori isolates in Hiroshima, Japan. METHODS Sixty Japanese patients with H. pylori infection were collected between 1999 and 2000. To detect the resistance to clarithromycin and metronidazole, mutations of the 23S ribosomal RNA (rRNA) and rdxA genes that are responsible for resistance in H. pylori, were examined by direct sequencing analysis. RESULTS Resistance to clarithromycin and metronidazole was detected in 12 (20.0%) and nine (15.0%) of the patients, respectively. Dual resistance to clarithromycin and metronidazole was detected in five (8.3%) patients. CONCLUSION These results indicate that the relatively high prevalence of the dual resistance in H. pylori isolates may need special attention and new therapeutic approaches in Japan.
Collapse
Affiliation(s)
- Toru Hiyama
- Health Service Center, Hiroshima University, Higashihiroshima, Japan.
| | | | | | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
19
|
Smeets LC, Arents NLA, van Zwet AA, Vandenbroucke-Grauls CMJE, Verboom T, Bitter W, Kusters JG. Molecular patchwork: Chromosomal recombination between two Helicobacter pylori strains during natural colonization. Infect Immun 2003; 71:2907-10. [PMID: 12704167 PMCID: PMC153253 DOI: 10.1128/iai.71.5.2907-2910.2003] [Citation(s) in RCA: 20] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Genetic analysis of two Helicobacter pylori strains isolated from a single gastric biopsy showed evidence of extensive horizontal gene transfer. Several large recombinations were identified in the rdxA gene, which is involved in metronidazole resistance.
Collapse
Affiliation(s)
- Leonard C Smeets
- Department of Medical Microbiology and Infection Control, Vrije Universiteit Medical Center, Amsterdam, The Netherlands.
| | | | | | | | | | | | | |
Collapse
|
|
20
|
Marais A, Bilardi C, Cantet F, Mendz GL, Mégraud F. Characterization of the genes rdxA and frxA involved in metronidazole resistance in Helicobacter pylori. Res Microbiol 2003; 154:137-44. [PMID: 12648728 DOI: 10.1016/s0923-2508(03)00030-5] [Citation(s) in RCA: 60] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Metronidazole (Mtz) resistance in Helicobacter pylori has been found to be associated with mutations in rdxA, a gene encoding an oxygen-insensitive NADPH nitroreductase, and enhanced by mutations in frxA, a gene encoding a NAD(P)H-flavin oxidoreductase. The roles of these two genes in Mtz resistance in H. pylori were examined in this study. The rdxA and frxA genes were sequenced in nine pairs of strains isolated from biopsies obtained from patients before and after failed eradication treatments which included Mtz and resulted in the appearance of resistant strains. Metronidazole resistance could be explained in seven of these pairs of strains by mutations in rdxA and frxA. However, in one pair of strains, rdxA was identical in the susceptible and resistant strains, and only changes in frxA were observed; and in another pair, neither rdxA nor frxA were different in the susceptible and resistant strains. Sequencing of the upstream region of frxA and of the recA gene in the latter pair of strains did not reveal any mutations. To establish whether mutations in frxA alone could be involved in Mtz resistance, a resistant Escherichia coli strain transformed with the frxA of a Mtz susceptible H. pylori strain was rendered susceptible, and transformation with a mutated H. pylori frxA gene under the same conditions did not change the resistant E. coli phenotype. The results suggested that a Mtz resistance phenotype may arise in H. pylori without mutations in rdxA or frxA, or with mutations only in frxA.
Collapse
Affiliation(s)
- Armelle Marais
- Laboratoire de Bactériologie, Université Victor Segalen Bordeaux 2, 146, rue Léo Saignat, 33076 Bordeaux, France
| | | | | | | | | |
Collapse
|
|
21
|
Abstract
Modern triple drug regimens are highly effective for treating Helicobacter pylori infection, but bacterial resistance to one of the most effective antibiotics, metronidazole, is a serious and increasing problem. The activity of metronidazole in H. pylori is dependent on reduction of its nitro moiety to highly reactive compounds that cause DNA strand breakage. The acquisition of resistance is highly associated with mutational inactivation of the rdxA gene, which encodes an oxygen-insensitive NADPH nitroreductase. Recent evidence has suggested that inactivation of frxA (NADPH flavin oxidoreductase), fdxB (ferredoxin-like protein) and possibly other reductase-encoding genes may also contribute to the resistant phenotype. Improved understanding of the mechanisms of metronidazole resistance in H. pylori is essential for the development and validation of biopsy-based tests for detection of resistance in clinical practice.
Collapse
Affiliation(s)
- Peter J Jenks
- Institute of Infections and Immunity, Queen's Medical Centre, University Hospital, Floor C, West Block, NG7 2UH, Nottingham, UK.
| | | |
Collapse
|
|
22
|
Latham SR, Owen RJ, Elviss NC, Labigne A, Jenks PJ. Differentiation of metronidazole-sensitive and -resistant clinical isolates of Helicobacter pylori by immunoblotting with antisera to the RdxA protein. J Clin Microbiol 2001; 39:3052-5. [PMID: 11526127 PMCID: PMC88295 DOI: 10.1128/jcm.39.9.3052-3055.2001] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2001] [Accepted: 06/14/2001] [Indexed: 01/06/2023] Open
Abstract
Antimicrobial resistance in Helicobacter pylori is a serious and increasing problem, and the development of rapid, reliable methods for detecting resistance would greatly improve the selection of antibiotics used to treat gastric infection with this organism. We assessed whether detection of the RdxA protein could provide the basis for determining the susceptibility of H. pylori to metronidazole. In order to raise polyclonal antisera to RdxA, we cloned the rdxA gene from H. pylori strain 26695 into the commercial expression vector pMAL-c2, purified the resultant fusion protein by affinity chromatography, and used this recombinant RdxA preparation to immunize rabbits. We then used this specific anti-RdxA antibody to perform immunoblotting on whole bacterial cell lysates of 17 metronidazole-sensitive and 27 metronidazole-resistant clinical isolates of H. pylori. While a 24-kDa immunoreactive band corresponding to the RdxA protein was observed in all metronidazole-sensitive strains, this band was absent in 25 of 27 resistant isolates. Our results indicate that testing for the absence of the RdxA protein would identify the majority of clinical isolates that will respond poorly to metronidazole-containing eradication regimens and have implications for the development of assays capable of detecting metronidazole resistance in H. pylori.
Collapse
Affiliation(s)
- S R Latham
- Department of Medical Microbiology, Royal Free and University College Medical School, London, United Kingdom
| | | | | | | | | |
Collapse
|
|
23
|
Kwon DH, Hulten K, Kato M, Kim JJ, Lee M, El-Zaatari FA, Osato MS, Graham DY. DNA sequence analysis of rdxA and frxA from 12 pairs of metronidazole-sensitive and -resistant clinical Helicobacter pylori isolates. Antimicrob Agents Chemother 2001; 45:2609-15. [PMID: 11502537 PMCID: PMC90700 DOI: 10.1128/aac.45.9.2609-2615.2001] [Citation(s) in RCA: 39] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
We previously reported that inactivation of rdxA and/or frxA converted Helicobacter pylori from metronidazole sensitive to metronidazole resistant. To examine the individual roles of rdxA and frxA in the development of metronidazole resistance in H. pylori, we examined the status of rdxA and frxA from 12 pairs of metronidazole-sensitive and -resistant H. pylori isolates obtained following unsuccessful therapy containing metronidazole. Arbitrary primed fingerprinting analyses revealed that the genotypes of 11 sensitive and resistant pairs of strains were essentially identical. Amino acid sequence identities of RdxA and FrxA from the 14 metronidazole-sensitive isolates ranged from 92 to 98% and 95 to 98%, respectively, compared to that of H. pylori J99 (MIC, 1 microg/ml). All strains with high-level metronidazole resistance (MICs, 128 microg/ml) contained premature truncation of both RdxA and FrxA caused by nonsense and/or frameshift mutations. Strains with intermediate resistance to metronidazole (MICs, 32 to 64 microg/ml) contained a single premature truncation and/or altered RdxA and FrxA caused by nonsense, frameshift, and unique missense mutations. The low-level metronidazole-resistant strains (MICs, 8 microg/ml) contained unique missense mutations in FrxA but no specific changes in RdxA. The results demonstrate that alterations in both the rdxA and frxA genes are required for moderate and high-level metronidazole resistance and that metronidazole resistance that develops during anti-H. pylori therapy containing metronidazole is most likely to involve a single sensitive strain infection rather than a coinfection with a metronidazole-resistant strain.
Collapse
Affiliation(s)
- D H Kwon
- Department of Medicine, Baylor College of Medicine and Veterans Affairs Medical Center, Houston, Texas 77030, USA.
| | | | | | | | | | | | | | | |
Collapse
|
|
24
|
Mégraud F. Resistance of Helicobacter pylori to antibiotics and its impact on treatment options. Drug Resist Updat 2001; 4:178-86. [PMID: 11768331 DOI: 10.1054/drup.2001.0203] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
The treatment of Helicobacter pylori infection is jeopardized by resistance to the antibiotics used, which turns out to be the main risk factor for failure. Resistance is due to point mutations. For clarithromycin only two sites in the 23S rRNA sequence are concerned and can be easily detected by molecular methods, while for metronidazole several mutations on rdxA and other genes can be responsible and so do not allow such detection. The situation for the rare cases of amoxicillin resistance is not fully determined. The impact of resistance on the clinical outcome is dramatic for clarithromycin while it only decreases the success by 20% for metronidazole.
Collapse
Affiliation(s)
- F Mégraud
- Laboratoire de Bactériologie, Hôpital Pellegrin, Bordeaux, France.
| |
Collapse
|