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Al Qady A, Aldhaleei W, Salih M, Ali M, Menakuru S, Nayar KD, Wang Z, Stancampiano FF, Harris D, Bi Y. Accuracy of Fecal Polymerase Chain Reaction Testing in Clarithromycin-Resistant Helicobacter Pylori: A Systematic Review and Meta-Analysis. Clin Transl Gastroenterol 2025; 16:e00792. [PMID: 39620581 PMCID: PMC11845177 DOI: 10.14309/ctg.0000000000000792] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/26/2024] [Accepted: 11/09/2024] [Indexed: 12/21/2024] Open
Abstract
INTRODUCTION The increasing prevalence of clarithromycin (CLA)-resistant Helicobacter pylori(H. pylori) strains poses a significant challenge in the management of H. pylori infections. This systematic review and meta-analysis investigates the diagnostic accuracy of polymerase chain reaction (PCR) in identifying CLA-resistant H. pylori strains in stool. METHODS A comprehensive literature search was conducted using PubMed, Embase, and Cochrane databases from database inception to April 30, 2023. Eligible studies evaluated the effectiveness of PCR stool tests in detecting CLA-resistant H. pylori strains in adults (>18-year-old). Studies of pediatric populations, alternative methods to PCR or stool samples, and reference tests other than gastric biopsy were excluded. The bivariate random-effects model was used to pool diagnostic accuracy from the included studies. RESULTS The analysis of 11 prospective diagnostic studies with a total of 866 patients showed a pooled sensitivity of 0.97 (95% CI: 0.9-0.99) and a pooled specificity of 0.98 (95% CI: 0.81-1.00). Subgroup analysis based on the used technique demonstrated consistent findings without notable variations. The diagnostic odds ratio was calculated at 1843.92 (95% CI: 134.28-25,321.3). The positive likelihood ratio was determined as 51.02 (95% CI: 4.61-564.5), while the negative likelihood ratio was found to be 0.03 (95% CI: 0.01-0.1). DISCUSSION PCR testing for clarithromycin-resistant H. pylori was highly sensitive and specific across studies with proven reliability in clinical practice, particularly in outpatient settings. Their implementation offers cost-effectiveness and the potential for tailored treatment strategies, holding promise for improved patient outcomes.
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Affiliation(s)
- Ahmed Al Qady
- School of Medicine, Indiana University, Muncie, Indiana, USA
| | - Wafa Aldhaleei
- Division of Gastroenterology and Hepatology, Mayo Clinic, Jacksonville, Florida, USA
| | | | - Marriam Ali
- School of Medicine, Indiana University, Muncie, Indiana, USA
| | | | - Kapil Dev Nayar
- Division of Gastroenterology and Hepatology, Mayo Clinic, Jacksonville, Florida, USA
| | - Zhen Wang
- Health Care Policy and Research, Mayo Clinic, Rochester, New York, USA
| | | | - Dana Harris
- Department of Medicine, Mayo Clinic, Jacksonville, Florida, USA
| | - Yan Bi
- Division of Gastroenterology and Hepatology, Mayo Clinic, Jacksonville, Florida, USA
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Ren X, Suo B, Li C, Ping G, Ma L, Shi Y, Zhou K, Wang Y, Tian X, Zhou L, Song Z. Comparative analysis of the detection of antibiotic genotypic resistance with gastric mucosa, gastric fluid, and fecal samples in patients with Helicobacter pylori infection. J Clin Microbiol 2025; 63:e0103424. [PMID: 39679670 PMCID: PMC11784280 DOI: 10.1128/jcm.01034-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/10/2024] [Accepted: 11/23/2024] [Indexed: 12/17/2024] Open
Abstract
Genotypic methods for detecting antibiotic resistance in Helicobacter pylori infection offer a rapid, convenient, and accurate approach for tailored therapy. However, existing studies predominantly examine single sample types and lack comparative analyses across different samples. This study comprehensively detects and compares genotypic resistance to clarithromycin and levofloxacin in gastric mucosa, gastric fluid, and fecal samples from the same patients. The study enrolled 183 participants, comprising 124 H. pylori-positive and 59 H. pylori-negative patients. All participants provided fecal samples and underwent gastroscopy for the collection of gastric mucosa and gastric fluid. Real-time PCR was employed to detect genotypic resistance to clarithromycin and levofloxacin in conjunction with bacterial culture and antibiotic susceptibility testing. Genotypic resistance detection rates for clarithromycin were 100% in gastric mucosa, 99.2% in gastric fluid, and 79.8% in fecal samples. For levofloxacin, detection rates were 97.6%, 96.8%, and 72.6%, respectively. The results showed that PCR detection for clarithromycin exhibited high sensitivity (0.94-0.95) and specificity (0.88-0.89) across all sample types. However, PCR detection for levofloxacin demonstrated slightly lower sensitivity (0.79-0.89) and specificity (0.79-0.83). The comparison of genotypic resistance results by PCR among the three sample types showed that gastric mucosa and gastric juice exhibited higher consistency, while the consistency between feces and both gastric mucosa and gastric juice was lower. This study confirmed good consistency between genotypic and phenotypic resistance in clarithromycin and levofloxacin. While both gastric mucosa and gastric fluid samples demonstrated high detection performance, the efficiency of detecting fecal samples was constrained by challenges in DNA extraction. IMPORTANCE This study, with a large sample size, comprehensively tested both Helicobacter pylori-negative and -positive patients, including rapid urease test, histopathological evaluation and staining, bacterial culture, susceptibility testing, and resistance gene mutation analysis. By simultaneously examining gastric mucosa, gastric juice, and fecal samples from the same individuals, we minimized confounding factors arising from different sample sources, ensuring the reliability of our results. This approach effectively delineated the differences and characteristics in detection performance among different sample types, offering crucial reference data for selecting appropriate detection samples and identifying areas for improvement. The findings revealed robust concordance between genotypic and phenotypic resistance, with both gastric mucosa and gastric juice samples demonstrating excellent detection performance. However, the efficiency of detecting resistance in fecal samples was hampered by challenges in DNA extraction.
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Affiliation(s)
- Xinlu Ren
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Baojun Suo
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Cailing Li
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Guangjie Ping
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Lingling Ma
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Yanyan Shi
- Research Center of Clinical Epidemiology, Peking University Third Hospital, Beijing, China
| | - Kai Zhou
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Yuxin Wang
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Xueli Tian
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Liya Zhou
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
| | - Zhiqiang Song
- Department of Gastroenterology, Peking University Third Hospital, Beijing, China
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Kekic D, Jovicevic M, Kabic J, Lolic I, Gajic I, Stojkovic S, Ranin L, Milosavljevic T, Opavski N, Rankovic I, Milivojevic V. Genetic Determinants of Clarithromycin and Fluoroquinolones Resistance in Helicobacter pylori in Serbia. Antibiotics (Basel) 2024; 13:933. [PMID: 39452199 PMCID: PMC11505191 DOI: 10.3390/antibiotics13100933] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2024] [Revised: 09/20/2024] [Accepted: 09/29/2024] [Indexed: 10/26/2024] Open
Abstract
BACKGROUND/OBJECTIVES Stomach infections by Helicobacter pylori can cause acute or chronic gastritis, peptic ulcers, and gastric cancer. The rise in antibiotic resistance is a significant health issue highlighted by the World Health Organization. The increasing number of treatment failures underscores the necessity for antibiotic susceptibility testing (AST). The study aimed to investigate the current prevalence and resistance to fluoroquinolones and clarithromycin with their detected mutations. METHODS Stomach biopsies from symptomatic patients were subjected to molecular testing by GenoType Helico DR kit (Hain Lifescience GmbH, Nehren, Germany). RESULTS Positive findings on the presence of H. pylori were detected in 42.4% of symptomatic patients, with the significant majority of patients (69%) having previously failed treatments. The resistance rates to fluoroquinolones and clarithromycin were 53.9% and 58.5%, respectively, with significantly higher rates in secondary resistant strains. The main resistance markers in fluoroquinolones and clarithromycin were N87K (27.4%) and A2147G (78.6%), respectively. Hetero-resistance or mixed genotypes were detected in over 20% of tested patients. During the study period, a significant increase in trends in both fluoroquinolones and clarithromycin resistance rates was observed. CONCLUSIONS Results indicate the need for the implementation of the latest Maastricht VI Consensus recommendations for both AST whenever possible and the use of tailored guided therapy options due to high resistance rates and possible treatment failures. The GenoType Helico DR kit is a useful tool for AST, especially in cases of mixed H. pylori genotypes.
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Affiliation(s)
- Dusan Kekic
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | - Milos Jovicevic
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | - Jovana Kabic
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | - Iva Lolic
- Emergency Center, University Clinical Center of Serbia, 11 000 Belgrade, Serbia
| | - Ina Gajic
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | - Stefan Stojkovic
- Clinic for Gastroenterology and Hepatology, University Clinical Centre of Serbia, 11 000 Belgrade, Serbia
| | - Lazar Ranin
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | | | - Natasa Opavski
- Institute for Microbiology and Immunology, Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
| | - Ivan Rankovic
- Department of Gastroenterology and Liver Care, Royal Cornwall Hospitals NHS Trust University of Exeter, Treliske, Truro, Cornwall TR1 3LQ, UK
| | - Vladimir Milivojevic
- Clinic for Gastroenterology and Hepatology, University Clinical Centre of Serbia, 11 000 Belgrade, Serbia
- Faculty of Medicine, University of Belgrade, 11 000 Belgrade, Serbia
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Losurdo G, Mezzapesa M, Ditonno I, Piazzolla M, Pricci M, Girardi B, Celiberto F, Galeano G, Riezzo G, Russo F, Iannone A, Ierardi E, Di Leo A. Helicobacter pylori Secondary Antibiotic Resistance after One or More Eradication Failure: A Genotypic Stool Analysis Study. Antibiotics (Basel) 2024; 13:336. [PMID: 38667013 PMCID: PMC11047300 DOI: 10.3390/antibiotics13040336] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2024] [Revised: 04/04/2024] [Accepted: 04/05/2024] [Indexed: 04/29/2024] Open
Abstract
Helicobacter pylori (H. pylori) antibiotic resistance is the leading cause for unsuccessful eradication therapy. After one or more failures, the chance of encountering secondary antibiotic resistance increases. The aim of this study was to characterize genotypic secondary resistance in a cohort of southern Italian H. pylori patients with at least one previous failure. Such patients collected stool samples using a dedicated kit (THD fecal testTM), and bacterial DNA was extracted and amplified using RT-PCR. Resistance to clarithromycin, amoxicillin, metronidazole, levofloxacin, and tetracycline was assessed using a high-resolution melting curve. We enrolled 50 patients. A total of 72% of patients failed one previous antibiotic course, 16% failed two, 10% failed three, and 2% failed four. The rate of secondary antibiotic resistance was 16% for clarithromycin, 18% for metronidazole, 14% for amoxicillin, 14% for levofloxacin, and 2% for tetracycline. Among the eight clarithromycin-resistant patients, five (62.5%) previously received a clarithromycin-based regimen. The same rate was 33.3% (3/9) for metronidazole. The only tetracycline-resistant patient had received Pylera. In conclusion, our data seem to show that, even though secondary resistance is not very high, resistance to clarithromycin could be very likely related to previous exposure to this antibiotic.
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Affiliation(s)
- Giuseppe Losurdo
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Martino Mezzapesa
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Ilaria Ditonno
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Mariapaola Piazzolla
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | | | | | - Francesca Celiberto
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
- Ph.D. Course in Organs and Tissues Transplantation and Cellular Therapies, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Grazia Galeano
- Functional Gastrointestinal Disorders Research Group, National Institute of Gastroenterology IRCCS “Saverio de Bellis”, 70013 Castellana Grotte, Italy
| | - Giuseppe Riezzo
- Functional Gastrointestinal Disorders Research Group, National Institute of Gastroenterology IRCCS “Saverio de Bellis”, 70013 Castellana Grotte, Italy
| | - Francesco Russo
- Functional Gastrointestinal Disorders Research Group, National Institute of Gastroenterology IRCCS “Saverio de Bellis”, 70013 Castellana Grotte, Italy
| | - Andrea Iannone
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Enzo Ierardi
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Alfredo Di Leo
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
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Zhang Q, Yang S, Zhou J, Li Z, Wang L, Dong Q. Diagnostic accuracy of stool sample-based PCR in detecting Helicobacter pylori infection: a meta-analysis. J LAB MED 2023; 47:187-197. [DOI: 10.1515/labmed-2023-0004] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2025] Open
Abstract
Abstract
The present study was to evaluate the diagnostic accuracy of different types of PCR tests with the aim of determining which one performs best for detecting Helicobacter pylori in stool samples. Related articles were searched from PubMed, Embase, Web of Science databases, Scopus, and Scholar Google. The quality of included studies was assessed using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool and RevMan5.4 software. Pooled sensitivity, specificity, DOR, PLR and NLR for the stool PCR test in detecting H. pylori infection were performed by Stata 15.0 software. Subgroup meta-analysis was performed by Open Meta-analyst software. Ten studies were selected in this study. Stool PCR test had 92.0 % (83.0, 96.0 %) pooled sensitivity, 96.0 % (84.0, 99.0 %) pooled specificity, 296.0 (51.6, 1,696.9) pooled DOR, 26.1 (5.3, 128.7) pooled PLR and 0.09 (0.04, 0.18) NLR in the diagnosis of H. pylori infection, and summary receiver operating characteristic curve (SROC) illustrated an area under the curve (AUC) of 0.98. Subgroup meta-analysis showed rtPCR as having the highest diagnostic accuracy. Our results identify rtPCR as having the highest diagnostic accuracy for the detection of H. pylori in stool samples, and the stool PCR test as a reliable diagnostic tool for H. pylori infection.
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Affiliation(s)
- Qinglong Zhang
- Central Laboratories , Qingdao Municipal Hospital , Qingdao , P.R. China
- Shandong First Medical University , Jinan , P.R. China
| | - Shuang Yang
- Fever Clinic , Qingdao Municipal Hospital , Qingdao , P.R. China
| | - Jianhua Zhou
- Department of Stomatology , Qingdao Municipal Hospital , Qingdao , P.R. China
| | - Zhipeng Li
- Qingdao University , Qingdao , P.R. China
| | - Lili Wang
- Central Laboratories , Qingdao Municipal Hospital , Qingdao , P.R. China
| | - Quanjiang Dong
- Central Laboratories , Qingdao Municipal Hospital , Qingdao , P.R. China
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Šamanić I, Dadić B, Sanader Maršić Ž, Dželalija M, Maravić A, Kalinić H, Vrebalov Cindro P, Šundov Ž, Tonkić M, Tonkić A, Vuković J. Molecular Characterization and Mutational Analysis of Clarithromycin- and Levofloxacin-Resistance Genes in Helicobacter pylori from Gastric Biopsies in Southern Croatia. Int J Mol Sci 2023; 24:14560. [PMID: 37834008 PMCID: PMC10572715 DOI: 10.3390/ijms241914560] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2023] [Revised: 09/17/2023] [Accepted: 09/20/2023] [Indexed: 10/15/2023] Open
Abstract
Point mutations in the 23S rRNA, gyrA, and gyrB genes can confer resistance to clarithromycin (CAM) and levofloxacin (LVX) by altering target sites or protein structure, thereby reducing the efficacy of standard antibiotics in the treatment of Helicobacter pylori infections. Considering the confirmed primary CAM and LVX resistance in H. pylori infected patients from southern Croatia, we performed a molecular genetic analysis of three target genes (23S rRNA, gyrA, and gyrB) by PCR and sequencing, together with computational molecular docking analysis. In the CAM-resistant isolates, the mutation sites in the 23S rRNA gene were A2142C, A2142G, and A2143G. In addition, the mutations D91G and D91N in GyrA and N481E and R484K in GyrB were associated with resistance to LVX. Molecular docking analyses revealed that mutant H. pylori strains with resistance-related mutations exhibited a lower susceptibility to CAM and LVX compared with wild-type strains due to significant differences in non-covalent interactions (e.g., hydrogen bonds, ionic interactions) leading to destabilized antibiotic-protein binding, ultimately resulting in antibiotic resistance. Dual resistance to CAM and LVX was found, indicating the successful evolution of H. pylori resistance to unrelated antimicrobials and thus an increased risk to human health.
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Affiliation(s)
- Ivica Šamanić
- Department of Biology, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia; (B.D.); (M.D.); (A.M.)
| | - Blanka Dadić
- Department of Biology, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia; (B.D.); (M.D.); (A.M.)
| | - Željka Sanader Maršić
- Department of Physics, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia;
| | - Mia Dželalija
- Department of Biology, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia; (B.D.); (M.D.); (A.M.)
| | - Ana Maravić
- Department of Biology, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia; (B.D.); (M.D.); (A.M.)
| | - Hrvoje Kalinić
- Department of Compute Science, Faculty of Science, University of Split, Ruđera Boškovića 33, 21000 Split, Croatia;
| | - Pavle Vrebalov Cindro
- Department of Gastroenterology, University Hospital of Split, 21000 Split, Croatia; (P.V.C.); (Ž.Š.); (A.T.)
| | - Željko Šundov
- Department of Gastroenterology, University Hospital of Split, 21000 Split, Croatia; (P.V.C.); (Ž.Š.); (A.T.)
- Department of Internal Medicine, School of Medicine, University of Split, 21000 Split, Croatia
| | - Marija Tonkić
- Department of Medical Microbiology and Parasitology, School of Medicine, University of Split, 21000 Split, Croatia;
| | - Ante Tonkić
- Department of Gastroenterology, University Hospital of Split, 21000 Split, Croatia; (P.V.C.); (Ž.Š.); (A.T.)
- Department of Internal Medicine, School of Medicine, University of Split, 21000 Split, Croatia
| | - Jonatan Vuković
- Department of Gastroenterology, University Hospital of Split, 21000 Split, Croatia; (P.V.C.); (Ž.Š.); (A.T.)
- Department of Internal Medicine, School of Medicine, University of Split, 21000 Split, Croatia
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Celiberto F, Losurdo G, Pricci M, Girardi B, Marotti A, Di Leo A, Ierardi E. The State of the Art of Molecular Fecal Investigations for Helicobacter pylori ( H. pylori) Antibiotic Resistances. Int J Mol Sci 2023; 24:4361. [PMID: 36901792 PMCID: PMC10002064 DOI: 10.3390/ijms24054361] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2023] [Revised: 02/18/2023] [Accepted: 02/21/2023] [Indexed: 02/25/2023] Open
Abstract
A new paradigm shift for the treatment of Helicobacter pylori (H. pylori) infection would be timely due to a progressive increase in antibiotic resistance. Such a shift in the perspective of the H. pylori approach should include the preliminary assessment of antibiotic resistance. However, the availability of sensitivity tests is not widespread and the guidelines have always indicated empirical treatments without taking into account the need to make sensitivity tests accessible, i.e., the necessary starting point for improving results in different geographical areas. Currently, the traditional tools for this purpose (culture) are based on performing an invasive investigation (endoscopy) and often involve technical difficulties; thus, they were only confined to the settings where multiple attempts at eradication have failed. In contrast, genotypic resistance testing of fecal samples using molecular biology methods is much less invasive and more acceptable to patients. The purpose of this review is to update the state of the art of molecular fecal susceptibility testing for the management of this infection and to extensively discuss the potential benefits of their large-scale deployment, i.e., novel pharmacological opportunities.
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Affiliation(s)
- Francesca Celiberto
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
- Course in Organs and Tissues Transplantation and Cellular Therapies, Department of Precision Medicine Jonic Area, University “Aldo Moro” of Bari, 70124 Bari, Italy
| | - Giuseppe Losurdo
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | | | | | - Angela Marotti
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Alfredo Di Leo
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
| | - Enzo Ierardi
- Section of Gastroenterology, Department of Precision and Regenerative Medicine and Ionian Area, University of Bari, 70124 Bari, Italy
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Karmakar BC, Paul S, Basak S, Ghosh M, Mukherjee P, Das R, Chaudhuri S, Dutta S, Mukhopadhyay AK. Development and evaluation of a simple PCR assay and nested PCR for rapid detection of clarithromycin-resistant Helicobacter pylori from culture and directly from the biopsy samples in India. Gut Pathog 2023; 15:7. [PMID: 36782212 PMCID: PMC9925366 DOI: 10.1186/s13099-023-00530-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/28/2022] [Accepted: 01/23/2023] [Indexed: 02/15/2023] Open
Abstract
BACKGROUND Eradication of Helicobacter pylori provides the most effective treatment for gastroduodenal diseases caused by H. pylori infection. Clarithromycin, a member of the macrolide family, still remains the most important antibiotic used in H. pylori eradication treatment. But the increasing prevalence of clarithromycin resistant H. pylori strains due to point mutations in the V region of the 23S rRNA, poses a great threat in treating the ailing patients. So, we aimed for PCR-mediated rapid detection of the point mutation at 2143 position of 23S rRNA gene in H. pylori that is relevant to clarithromycin resistance from culture and simultaneously from biopsy specimens to avoid the empirical treatment. RESULTS Newly developed PCR assay using DNA of pure culture detected point mutation in 23S rRNA gene in 21 (8.04%) of 261 clinical strains tested. The agar dilution method showed that all these 21 strains were resistant to clarithromycin indicating the perfect match of the PCR based results. Additionally, the sequencing study also identified the A to G mutation at 2143 position in 23S rRNA gene of the resistant strains only. Consequently, the newly developed Nested-ASP-PCR dealing directly with 50 biopsy specimens demonstrated 100% sensitivity and specificity with the findings of agar dilution method taken as Gold standard. Bioinformatics based analysis such as accessibility analysis and dot plot clearly stated that the base pairing probability has increased due to mutation. Computational studies revealed that the point mutation confers more stability in secondary structure due to conversion of loop to stem. Furthermore, interaction studies showed binding affinity of the CLR to the mutant type is weaker than that to the wild type. CONCLUSION This assay outlines a rapid, sensitive and simple approach to identify point mutation that confers clarithromycin resistance as well as clarithromycin sensitive strains, providing rapid initiation of effective antibiotic treatment. Additionally, it is simple to adopt for hospital based diagnostic laboratories to evaluate the degree of regional clarithromycin resistance from biopsy specimens itself. Furthermore, in silico studies provide evidence or a signal that the prevalence of clarithromycin resistance may rise in the near future as a result of this point mutation.
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Affiliation(s)
- Bipul Chandra Karmakar
- Division of Bacteriology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Sangita Paul
- Division of Bacteriology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Surajit Basak
- Division of Bioinformatics, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Manisha Ghosh
- Division of Bioinformatics, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Piyali Mukherjee
- Division of Bacteriology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Rajashree Das
- Amity Institute of Biotechnology, Amity University, Noida, Uttar Pradesh, India
| | | | - Shanta Dutta
- Division of Bacteriology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India
| | - Asish Kumar Mukhopadhyay
- Division of Bacteriology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, 700010, India.
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Ansari S, Yamaoka Y. Helicobacter pylori Infection, Its Laboratory Diagnosis, and Antimicrobial Resistance: a Perspective of Clinical Relevance. Clin Microbiol Rev 2022; 35:e0025821. [PMID: 35404105 PMCID: PMC9491184 DOI: 10.1128/cmr.00258-21] [Citation(s) in RCA: 54] [Impact Index Per Article: 18.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Despite the recent decrease in overall prevalence of Helicobacter pylori infection, morbidity and mortality rates associated with gastric cancer remain high. The antimicrobial resistance developments and treatment failure are fueling the global burden of H. pylori-associated gastric complications. Accurate diagnosis remains the opening move for treatment and eradication of infections caused by microorganisms. Although several reports have been published on diagnostic approaches for H. pylori infection, most lack the data regarding diagnosis from a clinical perspective. Therefore, we provide an intensive, comprehensive, and updated description of the currently available diagnostic methods that can help clinicians, infection diagnosis professionals, and H. pylori researchers working on infection epidemiology to broaden their understanding and to select appropriate diagnostic methods. We also emphasize appropriate diagnostic approaches based on clinical settings (either clinical diagnosis or mass screening), patient factors (either age or other predisposing factors), and clinical factors (either upper gastrointestinal bleeding or partial gastrectomy) and appropriate methods to be considered for evaluating eradication efficacy. Furthermore, to cope with the increasing trend of antimicrobial resistance, a better understanding of its emergence and current diagnostic approaches for resistance detection remain inevitable.
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Affiliation(s)
- Shamshul Ansari
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu City, Oita, Japan
| | - Yoshio Yamaoka
- Department of Environmental and Preventive Medicine, Oita University Faculty of Medicine, Yufu City, Oita, Japan
- Department of Medicine, Gastroenterology and Hepatology Section, Baylor College of Medicine, Houston, Texas, USA
- Institute of Tropical Disease, Universitas Airlangga, Surabaya, Indonesia
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Gareayaghi N, Kocazeybek B. Detection of A2143G, A2142C, and A2142G Point Mutations with Real-Time PCR in Stool Specimens from Children Infected with Helicobacter pylori. Diagnostics (Basel) 2022; 12:2119. [PMID: 36140521 PMCID: PMC9497693 DOI: 10.3390/diagnostics12092119] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2022] [Revised: 08/26/2022] [Accepted: 08/29/2022] [Indexed: 11/16/2022] Open
Abstract
Reports have indicated an increasing prevalence of clarithromycin resistance in children relative to adults. Thus, it is important to investigate primary clarithromycin resistance before therapy to avoid treatment failure. A2142G, A2143G, and A2142C point mutations in the peptidyltransferase region of the 23S ribosomal RNA (rRNA) of Helicobacter pylori (H. pylori) strains isolated from children with gastrointestinal symptoms and asymptomatic children were evaluated via real-time polymerase chain reaction (RT-PCR) using fecal DNA samples. The presence of H. pylori was determined using a fecal H. pylori antigen enzyme-linked immunosorbent assay (ELISA) kit from the stools of children (n = 543). A2143G, A2142C, and A2142G point mutations were detected via RT-PCR and confirmed by sequencing the 23S rDNA. Fecal H. pylori antigen testing was positive in 101 symptomatic (49) and asymptomatic (52) children. A significant difference was found between the 0-5- and 5-18-year-old groups in terms of the A2143G and A2142G point mutations (p = 0.001). The A2142C mutation was not detected. There was a significant difference in the A2143G mutation between the symptomatic and asymptomatic 5-18-year-old children (p = 0.019). Macrolides are frequently used to treat upper respiratory tract infections in children due to their selective pressure effect. We suggest that H. pylori strains carrying mutations in the 23S RNA subunit conferring clarithromycin resistance may lead to an intense inflammatory response in the gastric epithelial cells, allowing them to proliferate more rapidly and causing possible diarrhea, halitosis, or abdominal pain in children.
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Affiliation(s)
- Nesrin Gareayaghi
- Istanbul Sisli Hamidiye Etfal Training and Research Hospital, Center for Blood, University of Health Sciences, Istanbul 34098, Turkey
| | - Bekir Kocazeybek
- Department of Medical Microbiology, Cerrahpasa Medical Faculty, Istanbul University-Cerrahpasa, Istanbul 34098, Turkey
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11
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Keikha M, Karbalaei M. Prevalence of antibiotic heteroresistance associated with Helicobacter pylori infection: A systematic review and meta-analysis. Microb Pathog 2022; 170:105720. [PMID: 35964816 DOI: 10.1016/j.micpath.2022.105720] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2022] [Revised: 08/05/2022] [Accepted: 08/08/2022] [Indexed: 10/15/2022]
Abstract
BACKGROUND Heteroresistance is a general term to describe diverse responses to specific antibiotics that can occur due to infection with either multiple bacterial strains or microevolution of a single strain during chronic infection. Due to limited information regarding heteroresistance Helicobacter pylori (H. pylori) infection, the current study was carried out to evaluate the prevalence of this phenomenon. METHODS For this study, all potential relevant studies were collected by searching international databases such as ISI Web of Science, PubMed, Scopus, ScienceDirect, ProQuest, Embase, DOAJ, China National Knowledge Infrastructure (CNKI), and Google Scholar. Finally, the frequency of heteroresistance H. pylori infection was measured using the event rate corresponding to 95% confidence intervals. RESULTS A total of 26 studies met our criteria; the eligible studies were related to the years 2001-2022. Our results showed that the prevalence of heteroresistance H. pylori strains was 60.1% to clarithromycin, 61.1% to metronidazole, 46.1% to levofloxacin, 3.8% to amoxicillin, and 21.1% to tetracycline. Our literature review also showed discrepancy of antimicrobial susceptibility test in strains isolated from different anatomical sites of the stomach. Heteroresistance H. pylori infection in developing countries is mostly due to infection with multiple H. pylori strains, while in developed countries it is due to microevolution of a single H. pylori strain in response to antibiotic pressure. CONCLUSIONS Heteroresistance H. pylori infection interferes with successful therapy and eventually can lead to the treatment failure. If a biopsy is taken from only one gastric site, resistant strains of H. pylori may be underestimated. Considering the role of heteroresistance H. pylori infection in treatment failure, it is very important for gastroenterologists to improve their knowledge about this fact. Regardingly, new guidelines should be developed and designed for the management and treatment of heteroresistance H. pylori infection.
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Affiliation(s)
- Masoud Keikha
- Department of Microbiology and Virology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Mohsen Karbalaei
- Department of Microbiology and Virology, School of Medicine, Jiroft University of Medical Sciences, Jiroft, Iran.
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12
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Yokota K, Osaki T, Hayashi S, Yokota S, Takeuchi H, Rimbara E, Ojima H, Sato T, Yonezawa H, Shibayama K, Tokunaga K, Kamiya S, Murakami K, Kato M, Sugiyama T. Establishment of a reference panel of Helicobacter pylori strains for antimicrobial susceptibility testing. Helicobacter 2022; 27:e12874. [PMID: 35255160 PMCID: PMC9286379 DOI: 10.1111/hel.12874] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/03/2021] [Revised: 12/28/2021] [Accepted: 12/29/2021] [Indexed: 12/17/2022]
Abstract
BACKGROUND Eradication treatment for Helicobacter pylori gastritis is covered by national health insurance since 2013 in Japan. However, eradication failure due to the increase of antimicrobial resistance has become a serious problem. The present study aims to establish a reference panel of Japanese H. pylori strains for antimicrobial susceptibility testing. METHOD A total of 28 strains were collected from 4 medical facilities in Japan. Antimicrobial susceptibility tests (ASTs) to clarithromycin (CLR), amoxicillin (AMX), and metronidazole (MNZ), were used to select standard reference strains. Complete genome sequences were also determined. RESULTS Three H. pylori strains (JSHR3, JSHR6 and JSHR31) were selected as standard reference strains by the Japanese Society for Helicobacter Research (JSHR). The minimum inhibitory concentrations (MICs) of the antibiotics against these 3 strains by agar dilution method with Brucella-based horse-serum-containing agar medium were as follows: JSHR3 (CLR 16 μg/ml, AMX 0.032 μg/ml and MNZ 4 μg/ml), JSHR6 (CLR 0.016 μg/ml, AMX 0.032 μg/ml and MNZ 4 μg/ml), and JSHR31 (CLR 16 μg/ml, AMX 1 μg/ml and MNZ 64 μg/ml). CONCLUSIONS A reference panel of H. pylori JSHR strains was established. The panel consisted of JSHR6, which was antibiotic-susceptible, JSHR3, which was CLR-resistant, and JSHR31, which was multi-resistant. This reference panel will be essential for standardized ASTs before the optimal drugs are selected for eradication treatment.
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Affiliation(s)
- Kenji Yokota
- Working Group of the Reference Panel of Helicobacter pylori Strains for Antimicrobial Susceptibility Tests in Japanese Society for Helicobacter ResearchTokyoJapan,Graduate School of Health ScienceOkayama UniversityOkayamaJapan
| | - Takako Osaki
- Working Group of the Reference Panel of Helicobacter pylori Strains for Antimicrobial Susceptibility Tests in Japanese Society for Helicobacter ResearchTokyoJapan,Department of Infectious DiseasesKyorin University School of MedicineMitakaJapan
| | - Shunji Hayashi
- Working Group of the Reference Panel of Helicobacter pylori Strains for Antimicrobial Susceptibility Tests in Japanese Society for Helicobacter ResearchTokyoJapan,Department of MicrobiologyKitasato University School of MedicineSagamiharaJapan
| | - Shin‐ichi Yokota
- Working Group of the Reference Panel of Helicobacter pylori Strains for Antimicrobial Susceptibility Tests in Japanese Society for Helicobacter ResearchTokyoJapan,Department of MicrobiologySapporo Medical University School of MedicineSapporoJapan
| | - Hiroaki Takeuchi
- Working Group of the Reference Panel of Helicobacter pylori Strains for Antimicrobial Susceptibility Tests in Japanese Society for Helicobacter ResearchTokyoJapan,Department of Medical Laboratory Sciences, Health SciencesInternational University of Health and Welfare Graduate SchoolNaritaJapan
| | - Emiko Rimbara
- Department of Bacteriology IINational Institute of Infectious Diseases (NIID)MusashimurayamaJapan
| | - Hinako Ojima
- Graduate School of Health ScienceOkayama UniversityOkayamaJapan
| | - Toyotaka Sato
- Department of MicrobiologySapporo Medical University School of MedicineSapporoJapan
| | - Hideo Yonezawa
- Department of Infectious DiseasesKyorin University School of MedicineMitakaJapan
| | - Keigo Shibayama
- Department of BacteriologyNagoya University Graduate School of MedicineNagoyaJapan
| | - Kengo Tokunaga
- Department of General MedicineKyorin University School of MedicineMitakaJapan
| | - Shigeru Kamiya
- Department of Infectious DiseasesKyorin University School of MedicineMitakaJapan
| | - Kazunari Murakami
- Department of GastroenterologyFaculty of MedicineOita UniversityOitaJapan
| | - Mototsugu Kato
- Department of GastroenterologyNational Hospital Organization National Hakodate HospitalHakodateJapan
| | - Toshiro Sugiyama
- Research Division of Molecular Target Therapeutics and Prevention of GI CancerHokkaido University HospitalSapporoJapan
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13
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Pichon M, Freche B, Burucoa C. New Strategy for the Detection and Treatment of Helicobacter pylori Infections in Primary Care Guided by a Non-Invasive PCR in Stool: Protocol of the French HepyPrim Study. J Clin Med 2022; 11:jcm11051151. [PMID: 35268242 PMCID: PMC8911369 DOI: 10.3390/jcm11051151] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2021] [Revised: 02/14/2022] [Accepted: 02/18/2022] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori (Hp) infects half of the world population and is responsible for gastric, duodenal ulcers and gastric cancer. The eradication of Hp cures ulcers and prevents ulcer recurrences and gastric cancer. Antibiotic resistance of Hp, and particularly clarithromycin resistance, is the primary cause of treatment failure and is a major concern identified by the WHO as a high priority requiring research into new strategies. Treatments guided by the detection of antibiotic resistance have proven their medical and economical superiority. However, this strategy is severely hampered by the invasive nature of the fibroscopy, since antibiotic resistance detection requires gastric biopsies. The eradication of Hp involves primary care physicians. The objective of this study will be to evaluate the feasibility of a strategy for the management of Hp infection in primary care by a recently developed non-invasive procedure and its non-inferiority in eradication rates compared with the strategy recommended by the French National Authority of Health. The non-invasive procedure is a PCR on stool to detect Hp infection and mutations conferring resistance to clarithromycin allowing a treatment guided by the results of the PCR. We present the protocol of a prospective, multicenter, randomized, controlled interventional study in two arms.
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Affiliation(s)
- Maxime Pichon
- Bacteriology Laboratory, Infectious Agents Department, CHU Poitiers, 86021 Poitiers, France
- INSERM U1070 Pharmacology of Antimicrobial Agents and Resistances, University of Poitiers, 86073 Poitiers, France;
- Correspondence: (M.P.); (C.B.); Tel.: +33-(0)5-49-44-41-43 (M.P.); +33-(0)5-49-44-64-68 (C.B.)
| | - Bernard Freche
- INSERM U1070 Pharmacology of Antimicrobial Agents and Resistances, University of Poitiers, 86073 Poitiers, France;
- Department of General Medicine, Faculty of Medicine and Pharmacy, University of Poitiers, 86073 Poitiers, France
| | - Christophe Burucoa
- Bacteriology Laboratory, Infectious Agents Department, CHU Poitiers, 86021 Poitiers, France
- INSERM U1070 Pharmacology of Antimicrobial Agents and Resistances, University of Poitiers, 86073 Poitiers, France;
- Correspondence: (M.P.); (C.B.); Tel.: +33-(0)5-49-44-41-43 (M.P.); +33-(0)5-49-44-64-68 (C.B.)
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14
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Sulo P, Šipková B. DNA diagnostics for reliable and universal identification of Helicobacter pylori. World J Gastroenterol 2021; 27:7100-7112. [PMID: 34887630 PMCID: PMC8613642 DOI: 10.3748/wjg.v27.i41.7100] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/20/2021] [Revised: 07/11/2021] [Accepted: 09/19/2021] [Indexed: 02/06/2023] Open
Abstract
Reliable diagnostics are a major challenge for the detection and treatment of Helicobacter pylori (H. pylori) infection. Currently at the forefront are non-invasive urea breath test (UBT) and stool antigen test (SAT). Polymerase chain reaction (PCR) is not endorsed due to nonspecific primers and the threat of false-positives. The specificity of DNA amplification can be achieved by nested PCR (NPCR), which involves two rounds of PCR. If the primers are properly designed for the variable regions of the 16S rRNA gene, it is not difficult to develop an NPCR assay for the unambiguous identification of H. pylori. Elaborate NPCR for a 454 bp amplicon was validated on 81 clinical biopsy, stool, and saliva samples, each from the same individuals, and compared with available H. pylori assays, namely histology, rapid urease test, SAT, and 13C-UBT. The assay was much more sensitive than simple PCR, and it was equally sensitive in biopsy samples as the 13C-UBT test, which is considered the gold standard. In addition, it is sufficiently specific because sequencing of the PCR products exclusively confirmed the presence of H. pylori-specific DNA. However, due to the threshold and lower abundance, the sensitivity was much lower in amplifications from stool or saliva. Reliable detection in saliva also complicates the ability of H. pylori to survive in the oral cavity aside from and independent of the stomach. The reason for the lower sensitivity in stool is DNA degradation; therefore, a new NPCR assay was developed to obtain a shorter 148 bp 16S rRNA amplicon. The assay was validated on stool samples from 208 gastroenterological patients and compared to SAT results. Surprisingly, this NPCR revealed the presence of H. pylori in twice the number of samples as SAT, indicating that many patients are misdiagnosed, not treated by antibiotics, and their problems are interpreted as chronic. Thus, it is unclear how to properly diagnose H. pylori in practice. In the first approach, SAT or UBT is sufficient. If samples are negative, the 148 bp amplicon NPCR assay should be performed. If problems persist, patients should not be considered negative, but due to threshold H. pylori abundance, they should be periodically tested. The advantage of NPCR over UBT is that it can be used universally, including questionable samples taken from patients with achlorhydria, receiving proton pump inhibitors, antibiotics, bismuth compound, intestinal metaplasia, or gastric ulcer bleeding.
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Affiliation(s)
- Pavol Sulo
- Department of Biochemistry, Comenius University, Bratislava 842 15, Slovakia
| | - Barbora Šipková
- Department of Biochemistry, Comenius University, Bratislava 842 15, Slovakia
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15
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Binmaeil H, Hanafiah A, Mohamed Rose I, Raja Ali RA. Development and Validation of Multiplex Quantitative PCR Assay for Detection of Helicobacter pylori and Mutations Conferring Resistance to Clarithromycin and Levofloxacin in Gastric Biopsy. Infect Drug Resist 2021; 14:4129-4145. [PMID: 34675558 PMCID: PMC8502538 DOI: 10.2147/idr.s325056] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2021] [Accepted: 09/08/2021] [Indexed: 12/12/2022] Open
Abstract
Aims and Objectives More than half of the world's population is infected with Helicobacter pylori, which can cause chronic gastritis. WHO has regarded clarithromycin-resistant H. pylori as a high priority pathogen. Hence, accurate diagnosis and detection of clarithromycin- and levofloxacin-resistant H. pylori strains is essential for proper management of infection. The objective of this study was to develop and optimize multiplex quantitative PCR assay for detection of mutations associated with clarithromycin and levofloxacin resistance in H. pylori directly from the gastric biopsies. Materials and Methods Specific primers and probes were designed to amplify ureA and mutations in 23S rRNA and gyrA genes. Singleplex and triplex qPCR assays were optimized and the assay's sensitivities and specificities were determined. The optimized multiplex qPCR assay was performed on 571 gastric biopsies. Results In this study, 14.7% (84/571) of the gastric biopsies were positive for H. pylori by conventional methods and 23.8% (136/571) were positive by the ureA-qPCR with 96.4% sensitivity and 88.5% specificity, while the +LR and -LR were 8.72 and 0.04, respectively. The ureA-positive samples (n=136) were subjected to multiplex qPCR which detected A2142G and A2143G mutations in the 23S rRNA gene (20.6%, 28/136) conferring clarithromycin resistance and gyrA mutations N87K, N87I, D91N, and D91Y (11.8%, 16/136) leading to levofloxacin resistance. The sensitivity and specificity of qPCR of 23S rRNA gene were 100% and 98.7%, respectively, while 100% and 99.8% for qPCR of gyrA, respectively. Conclusion The effectiveness of this qPCR is that it is sensitive in detecting low bacterial load and will help in timely detection of clarithromycin- and levofloxacin-resistant strains, especially in case of mixed infections. Since it is culture independent, it can inform clinicians about antibiotics to be included in the first-line therapy, thereby improving the management of H. pylori infection at a much greater pace.
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Affiliation(s)
- Hasyanee Binmaeil
- Department of Medical Microbiology and Immunology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia
| | - Alfizah Hanafiah
- Department of Medical Microbiology and Immunology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia.,GUT Research Group, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia
| | - Isa Mohamed Rose
- Department of Pathology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia
| | - Raja Affendi Raja Ali
- GUT Research Group, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia.,Department of Medicine, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, 56000, Malaysia
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16
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Li SY, Li J, Dong XH, Teng GG, Zhang W, Cheng H, Gao W, Dai Y, Zhang XH, Wang WH. The effect of previous eradication failure on antibiotic resistance of Helicobacter pylori: A retrospective study over 8 years in Beijing. Helicobacter 2021; 26:e12804. [PMID: 33860967 DOI: 10.1111/hel.12804] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/03/2020] [Revised: 02/23/2021] [Accepted: 03/17/2021] [Indexed: 12/26/2022]
Abstract
BACKGROUND Antibiotic resistance is the main cause of Helicobacter pylori (H. pylori) treatment failure. This study aimed to explore the characteristics of antibiotic resistance of H. pylori isolates in Beijing in the last 8 years and to estimate the impact of previous eradication failure on resistance patterns. MATERIALS AND METHODS This retrospective study included data from a single center in Beijing from 2013 to 2020. Antibiotic susceptibility of 365 clinical H. pylori isolates was tested for amoxicillin, clarithromycin, metronidazole, levofloxacin, moxifloxacin, and tetracycline. The characteristics of the included patients and their previous eradication history were collected. Primary and secondary resistance rates of H. pylori to the six antibiotics and the impact of previous eradication failure on antibiotic resistance patterns were analyzed. RESULTS The overall primary resistance rates of amoxicillin, clarithromycin, metronidazole, levofloxacin, moxifloxacin, and tetracycline were 0.7%, 55.2%, 68.0%, 49.7%, 64.5%, and 0%, with no significant increase during the observed period; while the secondary resistance rates were 3.2%, 96.7%, 90.7%, 93.1%, 80.0%, and 0%, respectively. The secondary resistance rate of clarithromycin (p < .001), metronidazole (p = .001), and levofloxacin (p < .001) significantly increased to 100% as the number of previous eradication therapies increased and exhibited a linear association. For strains naive to eradication, only 6.8% were susceptible to all the antibiotics, while 32.4% were single resistant, and 60.8% dual or multiple resistant. Clarithromycin+metronidazole+fluoroquinolone multiple resistance was the predominant pattern (0 course: 21.6%, 1 course: 37.5%, 2 courses: 56.1%, ≥3 courses: 71.1%; p < .001) for patients with treatment failure. The prevalence of dual or multiple-resistance patterns increased significantly as the number of previous therapies increased. CONCLUSIONS The prevalence of primary and secondary resistance rates of clarithromycin, metronidazole, moxifloxacin, and levofloxacin were high in Beijing. Multiple-resistance patterns were common after treatment failure. Resistance rates of amoxicillin and tetracycline remained low and stable.
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Affiliation(s)
- Si-Yu Li
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Jiang Li
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Xin-Hong Dong
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Gui-Gen Teng
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Wei Zhang
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Hong Cheng
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Wen Gao
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Yun Dai
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Xiao-He Zhang
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
| | - Wei-Hong Wang
- Department of Gastroenterology, Peking University First Hospital, Beijing, China
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17
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Clinical Evaluation of a Real-Time PCR Assay for Simultaneous Detection of Helicobacter pylori and Genotypic Markers of Clarithromycin Resistance Directly from Stool. J Clin Microbiol 2021; 59:JCM.03040-20. [PMID: 33536295 DOI: 10.1128/jcm.03040-20] [Citation(s) in RCA: 22] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2020] [Accepted: 02/02/2021] [Indexed: 12/28/2022] Open
Abstract
Helicobacter pylori infection is mainly diagnosed noninvasively, with susceptibility testing traditionally requiring endoscopy. Treatment is empirical, with clarithromycin-based triple therapy recommended where resistance rates are below 15%. Rising rates of clarithromycin resistance, resulting in high clarithromycin-based therapy failure rates, are seen worldwide, but U.S. data are limited. We developed a real-time PCR assay for simultaneous detection of H. pylori and genotypic markers of clarithromycin resistance directly from stool specimens. The assay was validated by testing 524 stool samples using an H. pylori stool antigen test as the reference method for detection accuracy and Sanger sequencing to confirm genotypic susceptibility results. A separate set of 223 antigen-positive stool samples was tested and retrospective medical record review conducted to define clinical utility. PCR resulted in 88.6% and 92.8% sensitivity in the validation and clinical study sets, respectively. Sequencing confirmed correct detection of clarithromycin resistance-associated mutations in all positive validation samples. The PCR-predicted clarithromycin resistance rate was 39% in the clinical data set overall and 31% in treatment-naive patients; the clarithromycin-based triple therapy eradication rate in treatment-naive patients was 62%. The clarithromycin-based triple therapy success was lower when resistance was predicted by PCR (41%) than when no resistance was predicted (70%; P = 0.03). PCR results were positive in 98% of antigen-positive stools from patients tested for eradication. The described PCR assay can accurately and noninvasively diagnose H. pylori, provide genotypic susceptibility, and test for eradication. Our findings support the need for susceptibility-guided therapy in our region if a clarithromycin-based regimen is considered.
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18
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Gong RJ, Xu CX, Li H, Liu XM. Polymerase chain reaction-based tests for detecting Helicobacter pylori clarithromycin resistance in stool samples: A meta-analysis. World J Clin Cases 2021; 9:133-147. [PMID: 33511178 PMCID: PMC7809662 DOI: 10.12998/wjcc.v9.i1.133] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/06/2020] [Revised: 11/07/2020] [Accepted: 11/14/2020] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Helicobacter pylori (H. pylori) infection is closely associated with the etiology of a variety of gastric diseases. The effective eradication of H. pylori infection has been shown to reduce the incidence of gastric carcinoma. However, the rate of H. pylori eradication has significantly declined due to its increasing resistance to antibiotics, especially to clarithromycin. Therefore, the detection of clarithromycin resistance is necessary prior to the treatment of H. pylori. Although many studies have been conducted on the use of polymerase chain reaction (PCR)-based tests to detect clarithromycin resistance in stool samples, no accurate data on the feasibility of these tests are available. Here, we performed a meta-analysis to assess the feasibility of these noninvasive tests.
AIM To evaluate the reliability of PCR-based tests for detecting H. pylori clarithromycin resistance in stool samples.
METHODS We searched PubMed, Medline, Embase, and other databases for articles that evaluated the value of the PCR analysis of stool samples for detecting the resistance of H. pylori to clarithromycin. We collected cross-sectional studies that met the inclusion criteria. Diagnostic accuracy measures were pooled using a random-effects model. The risk of bias was assessed using the Quality Assessment of Diagnostic Accuracy Studies-2 tool. Subgroup analysis was also conducted according to PCR type, purification technique, reference standard, mutation site, sample weight, number of patients, and age group, and the clinical utility of diagnostic tests was evaluated using the Likelihood Ratio Scatter Graph.
RESULTS Out of the 1818 identified studies, only 11 met the eligibility criteria, with a total of 592 patients assessed. A meta-analysis of the random-effect model showed that PCR-based analysis of stool samples had high diagnostic accuracy for detecting clarithromycin resistance in patients infected with H. pylori. The combined sensitivity was 0.91 [95% confidence interval (CI): 0.83-0.95], Q = 30.34, and I2 = 67.04, and the combined specificity was 0.97 (95%CI: 0.62-1.00), Q = 279.54, and I2 = 96.42. The likelihood ratio for a positive test was 33.25 (95%CI: 1.69-652.77), and that for a negative test was 0.10 (95%CI: 0.05-0.18), with an area under the curve of 0.94. The diagnostic odds ratio was 347.68 (95%CI: 17.29-6991.26). There was significant statistical heterogeneity, and the sub-analyses showed significant differences in the number of patients, sample weight, purification methods, PCR types, mutation points, and reference standards. The included studies showed no risk of publication bias.
CONCLUSION PCR-based tests on stool samples have high diagnostic accuracy for detecting H. pylori clarithromycin resistance.
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Affiliation(s)
- Ren-Jie Gong
- Department of Gastroenterology, The Third Xiangya Hospital of Central South University, Changsha 410013, Hunan Province, China
| | - Can-Xia Xu
- Department of Gastroenterology, The Third Xiangya Hospital of Central South University, Changsha 410013, Hunan Province, China
| | - Huan Li
- Department of Gastroenterology, The Third Xiangya Hospital of Central South University, Changsha 410013, Hunan Province, China
| | - Xiao-Ming Liu
- Department of Gastroenterology, The Third Xiangya Hospital of Central South University, Changsha 410013, Hunan Province, China
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19
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Kakiuchi T, Hashiguchi K, Imamura I, Nakayama A, Takamori A, Okuda M, Matsuo M. Assessment of a novel method to detect clarithromycin-resistant Helicobacter pylori using a stool antigen test reagent. BMC Gastroenterol 2020; 20:397. [PMID: 33228552 PMCID: PMC7682763 DOI: 10.1186/s12876-020-01549-9] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/02/2020] [Accepted: 11/19/2020] [Indexed: 02/08/2023] Open
Abstract
BACKGROUND The resistance rate of Helicobacter pylori to clarithromycin (CAM) is high among infected children in Japan. Therefore, a new method for detecting CAM-resistant H. pylori using a minimally invasive technique is strongly desired. We aimed to investigate the clinical usefulness of our newly developed nested polymerase chain reaction-quenching probe (Nested PCR-QP) method using stool specimens. METHODS We first evaluated our method using a residual solution of the H. pylori stool antigen test for adolescents. Then, we evaluated our method using culture testing for adults. RESULTS Among 57 middle school students with H. pylori, the Nested PCR-QP test results of 53 (90.3%) were able to be analyzed. A total of 28 students had CAM resistance mutations. We found a genetic mutation in 28 students and no mutation in 23 students, and these results were consistent with those of PCR-direct sequencing. In the 23 adults who were diagnosed with H. pylori infection using the rapid urease test and culture testing, we were able to use Nested PCR-QP for analyzing 21 adults who tested positive in the stool H. pylori antigen test. The results obtained for all 21 adults were consistent with those obtained via the drug susceptibility test. CONCLUSIONS Our novel method could be useful for non-invasively detecting CAM resistance mutations in H. pylori. This may help select a drug to reduce eradication failure rates against H. pylori. Trial registration This study was registered with the University Hospital Medical Information Network Clinical Trials Registry (no. UMIN000030632, https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000034977 ) on 29 December 2017.
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Affiliation(s)
- Toshihiko Kakiuchi
- Department of Pediatrics, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga-shi, Saga, 849-8501, Japan.
| | - Kazutoshi Hashiguchi
- Division of Gastroenterology, Department of Internal Medicine, Imamura Hospital, Tosu, Japan
| | - Ichiro Imamura
- Division of Gastroenterology, Department of Internal Medicine, Imamura Hospital, Tosu, Japan
| | - Aiko Nakayama
- Department of Pediatrics, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga-shi, Saga, 849-8501, Japan
| | - Ayako Takamori
- Division of Clinical Research Center, Saga University Hospital, Saga, Japan
| | - Masumi Okuda
- Department of Pediatrics, Hyogo College of Medicine, Nishinomiya, Japan
| | - Muneaki Matsuo
- Department of Pediatrics, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga-shi, Saga, 849-8501, Japan
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20
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Leonardi M, La Marca G, Pajola B, Perandin F, Ligozzi M, Pomari E. Assessment of real-time PCR for Helicobacter pylori DNA detection in stool with co-infection of intestinal parasites: a comparative study of DNA extraction methods. BMC Microbiol 2020; 20:131. [PMID: 32448186 PMCID: PMC7247253 DOI: 10.1186/s12866-020-01824-5] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2020] [Accepted: 05/14/2020] [Indexed: 12/19/2022] Open
Abstract
Background Many studies reported high prevalence of H. pylori infection among patients co-infected with intestinal parasites. Molecular approach for the DNA detection of those microbes in stool have been proposed. However there are a few reports that evaluated the effect of bead-beating in relation to the H. pylori outcome. Therefore, we developed and evaluated two TaqMan-based real-time PCR (rt-PCR) qualitative assays for the detection of ureC (glmM) and cagA of Helicobacter pylori on DNA extracted by three procedures. Results The two PCRs were analysed on 100 stool samples from patients who were screened for intestinal parasites. Three DNA extraction procedures were used: 1) automation with bead beating, 2) automation without bead beating and 3) hand column. The specificity of the new assays was confirmed by sequencing the PCR products and by the lack of cross-reactivity with other bacteria or pathogens DNA. Rt-PCR assays showed a detection limit of 10^4 bacteria/200 mg stool. The ureC_PCR with bead beating process was compared to conventional stool antigen test (SAT), with 94.12 and 93.75% of respectively sensitivity and specificity. However, the discordant samples were confirmed by DNA sequencing suggesting a potential higher sensitivity and specificity of PCR. Conclusions Our findings showed that the automation with bead-beating –suggested procedure for intestinal parasitic infections- can reach highly sensitive results in H. pylori detection on stool compared also with SAT. Thus, this work can provide new insights into the practice of a clinical microbiology laboratory in order to optimize detection of gastro-intestinal infections. Further studies are needed to better define the clinical value of this technique.
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Affiliation(s)
- Martina Leonardi
- Department of Infectious-Tropical Diseases and Microbiology, IRCCS Sacro Cuore Don Calabria Hospital, Via Don A. Sempreboni, 5 - 37024 Negrar di Valpolicella, Verona, Italy
| | - Giulia La Marca
- Department of Infectious-Tropical Diseases and Microbiology, IRCCS Sacro Cuore Don Calabria Hospital, Via Don A. Sempreboni, 5 - 37024 Negrar di Valpolicella, Verona, Italy
| | - Barbara Pajola
- Department of Infectious-Tropical Diseases and Microbiology, IRCCS Sacro Cuore Don Calabria Hospital, Via Don A. Sempreboni, 5 - 37024 Negrar di Valpolicella, Verona, Italy
| | - Francesca Perandin
- Department of Infectious-Tropical Diseases and Microbiology, IRCCS Sacro Cuore Don Calabria Hospital, Via Don A. Sempreboni, 5 - 37024 Negrar di Valpolicella, Verona, Italy
| | - Marco Ligozzi
- Department of Diagnostics and Public Health, University of Verona, Verona, Italy
| | - Elena Pomari
- Department of Infectious-Tropical Diseases and Microbiology, IRCCS Sacro Cuore Don Calabria Hospital, Via Don A. Sempreboni, 5 - 37024 Negrar di Valpolicella, Verona, Italy.
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Šeligová B, Lukáč Ľ, Bábelová M, Vávrová S, Sulo P. Diagnostic reliability of nested PCR depends on the primer design and threshold abundance of Helicobacter pylori in biopsy, stool, and saliva samples. Helicobacter 2020; 25:e12680. [PMID: 32057175 DOI: 10.1111/hel.12680] [Citation(s) in RCA: 31] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/29/2019] [Revised: 12/23/2019] [Accepted: 01/25/2020] [Indexed: 12/19/2022]
Abstract
BACKGROUND The aim of this work was to find a reliable nested PCR for the detection of Helicobacter pylori in biopsy, stool, and saliva specimens. MATERIALS AND METHODS Novel nested PCR was elaborated and validated on 81 clinical biopsy, stool, and saliva samples from the same individual and compared to available H pylori assays: histology, rapid urease test (RUT), stool antigen test (SAT), 13 C-urea breath test (UBT). RESULTS The efficiency and selectivity of 17 published nested polymerase chain reactions (PCR) available for Helicobacter pylori detection were re-evaluated. Most of them had serious limitations and mistakes in primer design. Hence, we elaborated a nested PCR for the unambiguous identification of H pylori in biopsy, stool, and saliva, using primers targeted to variable regions of the 16S ribosomal RNA (rRNA) gene. Moreover, we determined the detection limit by adding a known number of cells. This number was as low as 0.5 cells in a PCR vial, but due to the DNA isolation procedures, it required 1-5 × 103 cells/g or ml of specimen. The sensitivity for nested PCR from stomach biopsies was on the same scale as 13 C-UBT (93.8%), but it was much lower in amplifications from stool (31.3%). Sequencing of all obtained PCR products exclusively confirmed H pylori-specific DNA sequences. CONCLUSIONS Elaborated nested PCR assay can serve as an auxiliary method for controversial samples (patients with bleeding or taking proton-pump inhibitor) in laboratories with basic equipment. The sensitivity and specificity for the amplification from gastric biopsies was almost like 13 C-UBT. Despite the good sensitivity, the threshold occurrence and the ability to survive in the oral cavity aside from and independent of the stomach is the reason why H pylori DNA cannot be reliably detected in saliva, stool, and some biopsy samples.
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Affiliation(s)
- Barbora Šeligová
- Department of Biochemistry, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia
| | - Ľudovít Lukáč
- First Department of Internal Medicine, Faculty of Medicine, Comenius University, Bratislava, Slovakia
| | - Michaela Bábelová
- Department of Biochemistry, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia
| | - Silvia Vávrová
- Department of Molecular Biology, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia
| | - Pavol Sulo
- Department of Biochemistry, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia
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Diagnostic Accuracy of a Noninvasive Test for Detection of Helicobacter pylori and Resistance to Clarithromycin in Stool by the Amplidiag H. pylori+ClariR Real-Time PCR Assay. J Clin Microbiol 2020; 58:JCM.01787-19. [PMID: 31996442 DOI: 10.1128/jcm.01787-19] [Citation(s) in RCA: 53] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2019] [Accepted: 01/19/2020] [Indexed: 02/07/2023] Open
Abstract
The noninvasive detection of Helicobacter pylori and its resistance to clarithromycin could revolutionize the management of H. pylori-infected patients by tailoring eradication treatment without any need for endoscopy when histology is not necessary. Several real-time PCR tests performed on stools have been proposed, but their performances were either poor or they were tested on too few patients to be properly evaluated. We conducted a prospective, multicenter study including 1,200 adult patients who were addressed for gastroduodenal endoscopy with gastric biopsies and who were naive for eradication treatment in order to evaluate the performance of the Amplidiag H. pylori+ClariR assay recently developed by Mobidiag (Espoo, Finland). The results of the Amplidiag H. pylori+ClariR assay performed on DNA from stools (automatic extraction with the EasyMag system [bioMérieux]) were compared with those of culture/Etest and quadruplex real-time PCRs performed on two gastric biopsy samples (from the antrum and corpus) to detect the H. pylori glmM gene and mutations in the 23S rRNA genes conferring clarithromycin resistance. The sensitivity and specificity of the detection of H. pylori were 96.3% (95% confidence interval [CI], 92 to 98%) and 98.7% (95% CI, 97 to 99%), respectively. The positive and negative predictive values were evaluated to be 92.2% (95% CI, 92 to 98%) and 99.3% (95% CI, 98 to 99%), respectively. In this cohort, 160 patients (14.7%) were found to be infected (positive by culture and/or PCR). The sensitivity and specificity for detecting resistance to clarithromycin were 100% (95% CI, 88 to 100%) and 98.4% (95% CI, 94 to 99%), respectively.
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Helicobacter pylori Colonization in Patients with Adenotonsillar Hypertrophy: Study on Prevalence and Clinical Characteristics of Colonized Patients and Possible Association with Complications. ARCHIVES OF PEDIATRIC INFECTIOUS DISEASES 2020. [DOI: 10.5812/pedinfect.97561] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
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Feng ZH, Fan L, Yang J, Huo XY, Guo Y, Zhang Y, Lan CH. Mutant selection window of clarithromycin for clinical isolates of Helicobacter pylori. BMC Microbiol 2019; 19:176. [PMID: 31382897 PMCID: PMC6683470 DOI: 10.1186/s12866-019-1558-8] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2019] [Accepted: 07/29/2019] [Indexed: 12/18/2022] Open
Abstract
Background Clarithromycin-resistance is becoming a global health concern in the treatment of Helicobacter pylori (H. pylori). The mutant prevention concentration (MPC) represent the propensities of antimicrobial agents to select resistant mutants. The concentration range between the minimum inhibitory concentration (MIC) and the MPC is defined as mutant selection window (MSW). In this study, we aimed to determine the cause of increasing clarithromycin resistance by investigating the MSW for clinical isolates of H. pylori. Results A retrospective subgroup, which included 68 clarithromycin-sensitive H. pylori strains, was selected from a double-blind trial. The MICs and MPCs were determined using agar plate assays. Genotypic tests were performed using Sanger sequencing. All isolates were wild-type, and 33.82% (23/68) had a 0.016 mg/L MIC, 45.59% (31/68) had a 0.031 mg/L MIC, 16.18% (11/68) had a 0.062 ≤ MIC ≤ 0.125 mg/L, and 4.41% (3/68) had a 0.25 mg/L MIC. The MPC50/90 (mg/L) of the isolates were: 0.062/0.125, 0.125/0.5, 0.25/0.25 and 1/2, respectively. The MPCs showed a moderate correlation with the MICs (rs = 0.65, P < 0.0001). Using published data and MPC90, we calculated the time inside the MSW (TMSW) for low- and high-dose (200 or 500 mg bid) clarithromycin that were 6 and 0 h, 24 and 4 h, 15 and 2 h, 5 and 17 h for the strains with MICs (mg/L) of 0.016, 0.031, 0.062–0.125, and 0.25, respectively. Conclusions This study showed that in the clarithromycin-sensitive clinical isolates of H. pylori, low-dose clarithromycin may lead to decreased drug sensitivity or even clarithromycin resistance; strains with a 0.25 mg/L MIC display a high risk of treatment failure.
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Affiliation(s)
- Zi-Han Feng
- College of Basic Medical Sciences, Army Medical University, Chongqing, 400042, China
| | - Ling Fan
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China
| | - Jing Yang
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China
| | - Xing-Yue Huo
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China
| | - Yan Guo
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China
| | - Yi Zhang
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China
| | - Chun-Hui Lan
- Department of Gastroenterology, Daping Hospital, Army Medical University, 10 Changjiang Branch Road, Chongqing, 400042, China.
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25
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Arévalo-Jaimes BV, Rojas-Rengifo DF, Jaramillo CA, de Molano BM, Vera-Chamorro JF, Del Pilar Delgado M. Genotypic determination of resistance and heteroresistance to clarithromycin in Helicobacter pylori isolates from antrum and corpus of Colombian symptomatic patients. BMC Infect Dis 2019; 19:546. [PMID: 31226948 PMCID: PMC6587245 DOI: 10.1186/s12879-019-4178-x] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2018] [Accepted: 06/10/2019] [Indexed: 02/10/2023] Open
Abstract
BACKGROUND The effectiveness of Helicobacter pylori first-line treatment has decreased drastically with the rise of strains resistant to clarithromycin. Therapy failure has also been described in patients with infections by strains with dissimilar antimicrobial susceptibilities. The present study aims to estimate the prevalence of resistance and heteroresistance to clarithromycin in H. pylori isolates from antrum and corpus of Colombian patients. METHODS The study material included 126 isolates from antrum and corpus biopsies from 63 symptomatic patients over 18 years old who had a gastric endoscopy performed on them between June 2014 to August 2016. PCR amplification and sequencing of the H. pylori 23S rDNA gene was performed to determine the presence of mutations associated with clarithromycin resistance. Random amplified polymorphic DNA analysis was implemented in cases of resistance and heteroresistance. RESULTS The overall frequency of resistance to clarithromycin was 38.1% (24/63 patients), of which 19 patients had resistant isolates in both stomach segments (14 with A2143G mutation and 5 with A2142G mutation), and 5 patients had a heteroresistant status. The remaining 61.9% (39/63 patients) presented only susceptible isolates. DNA fingerprinting analysis showed different patterns in 4/22 paired isolates. CONCLUSIONS The high prevalence of H. pylori clarithromycin-resistance obtained (> 15%) constitutes an alert for gastroenterologists and suggests the need for reconsideration of the current eradication regimen for H. pylori in the studied population. The data show that heteroresistance status is an additional factor to be considered in the assessment of resistance. In consequence, it is advisable to examine at least two biopsies from different gastric segments.
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Affiliation(s)
- Betsy Verónica Arévalo-Jaimes
- Molecular Diagnostics and Bioinformatics Laboratory, Department of Biological Sciences, Los Andes University, Cra 1 # 18A- 10 Office J211, Zip Code, 111711, Bogotá, Colombia
| | - Diana F Rojas-Rengifo
- Molecular Diagnostics and Bioinformatics Laboratory, Department of Biological Sciences, Los Andes University, Cra 1 # 18A- 10 Office J211, Zip Code, 111711, Bogotá, Colombia
| | - Carlos Alberto Jaramillo
- Molecular Diagnostics and Bioinformatics Laboratory, Department of Biological Sciences, Los Andes University, Cra 1 # 18A- 10 Office J211, Zip Code, 111711, Bogotá, Colombia
| | - Belén Mendoza de Molano
- Gastroenterology Department, University Hospital Foundation Santa Fe de Bogotá, Bogotá, Colombia
| | | | - María Del Pilar Delgado
- Molecular Diagnostics and Bioinformatics Laboratory, Department of Biological Sciences, Los Andes University, Cra 1 # 18A- 10 Office J211, Zip Code, 111711, Bogotá, Colombia.
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Boltin D, Ashorov O, Benejat L, Hamouda D, Belfer RG, Niv Y, Dickman R, Perets TT. Novel high resolution melt curve assay for the analysis of predominance of Helicobacter pylori clarithromycin resistance. Pathog Dis 2019; 77:ftz042. [PMID: 31374569 DOI: 10.1093/femspd/ftz042] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2019] [Accepted: 07/31/2019] [Indexed: 01/05/2025] Open
Abstract
Clarithromycin resistance is the most common cause of Helicobacter pylori treatment failure and it is attributed to three point mutations, A2142G, A2142C and A2143G, within the 23S rRNA gene. We aimed to determine the prevalence of H. pylori clarithromycin resistance using a novel high resolution melt assay. A total of 151 stool samples were collected from treatment-naïve patients with general gastric discomfort who also performed 13CO2 breath tests. Stool antigen tests were also performed on 126 of the 151 stool samples collected. Bacterial DNA was extracted from the stool and analyzed by comparing it with four reference plasmids incorporating the three mutations and the wild type (WT) sequences. The melt assay detected 106 H. pylori positive samples, of which 54 had a WT sequence, and 52 had a point mutation associated with clarithromycin resistance, including A2142G in 10, A2142C in 13, A2143G in 18 and heterozygosity (multiple peaks) in 11. Compared with the gold standards (13CO2 breath and stool antigen tests), the melt assay had a sensitivity of 100% and 99% and a specificity of 82% and 78%, respectively. Therefore, our stool-based molecular assay is able to identify H. pylori infection and clarithromycin resistance. It could be used for screening prior to administration of clarithromycin eradication therapy.
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Affiliation(s)
- Doron Boltin
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
- Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Olga Ashorov
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
| | - Lucie Benejat
- Centre National de Référence des Campylobacters et Hélicobacters, Laboratoire de Bactériologie, CHU Pellegrin, Bordeaux, France
| | - Dalal Hamouda
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
| | - Rachel Gingold Belfer
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
- Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Yaron Niv
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
- Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Ram Dickman
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
- Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Tsachi Tsadok Perets
- Gastroenterology Laboratory and the Division of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petach Tikva, Israel
- Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
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Evaluation of a Novel Stool Antigen Rapid Test Kit for Detection of Helicobacter pylori Infection. J Clin Microbiol 2019; 57:JCM.01825-18. [PMID: 30567746 DOI: 10.1128/jcm.01825-18] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2018] [Accepted: 12/12/2018] [Indexed: 12/21/2022] Open
Abstract
The Quick Chaser H. pylori (QCP), which is a novel antigen detection kit for Helicobacter pylori, was developed recently. The previous examination kits targeted the catalase of H. pylori; however, this new test kit, to the best of our knowledge, is the first kit to target the flagellar protein of H. pylori The present study aimed to evaluate the efficacy of QCP compared with that of the already commercially available rapid test kit Testmate Rapid Pylori Antigen (TRP). TRP and QCP were utilized in 71 participants. The positive and negative concordance ratios of QCP to TRP were 100% (57/57) and 92.9% (13/14), respectively. Sensitivity based on the rapid urease test and culture test was 92.3%. Results of the dilution sensitivity test showed that QCP was eight times more sensitive to an H. pylori standard strain and the clarithromycin (CAM)-resistant clinical isolate and four times more sensitive to a CAM-susceptible clinical isolate than TRP. For the cross-reactivity test, 27 strains that exist in human feces, such as the Helicobacter genus, Bifidobacterium genus, Lactobacillus genus, and other resident bacteria, were selected. The results obtained using QCP were all negative, and no cross-reaction was observed. In conclusion, compared with TRP, QCP can detect H. pylori using clinical specimens with high sensitivity, regardless of CAM susceptibility and tolerance. No cross-reactivity was observed with other intestinal bacteria in humans. The kit is considered extremely useful in clinical settings.
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Iannone A, Giorgio F, Russo F, Riezzo G, Girardi B, Pricci M, Palmer SC, Barone M, Principi M, Strippoli GFM, Di Leo A, Ierardi E. New fecal test for non-invasive Helicobacter pylori detection: A diagnostic accuracy study. World J Gastroenterol 2018; 24:3021-3029. [PMID: 30038469 PMCID: PMC6054951 DOI: 10.3748/wjg.v24.i27.3021] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/28/2018] [Revised: 06/12/2018] [Accepted: 06/27/2018] [Indexed: 02/06/2023] Open
Abstract
AIM To assess the diagnostic accuracy of a new fecal test for detecting Helicobacter pylori (H. pylori), using13C-urea breath test as the reference standard, and explore bacterial antibiotic resistance. METHODS We conducted a prospective two-center diagnostic test accuracy study. We enrolled consecutive people≥ 18 years without previous diagnosis of H. pylori infection, referred for dyspepsia between February and October 2017. At enrollment, all participants underwent 13C-urea breath test. Participants aged over 50 years were scheduled to undergo upper endoscopy with histology. Participants collected stool samples 1-3 d after enrollment for a new fecal investigation (THD fecal test). The detection of bacterial 23S rRNA subunit gene indicated H. pylori infection. We also used the index diagnostic test to examine mutations conferring resistance to clarithromycin and levofloxacin. Independent investigators analyzed index test and reference test standard results blinded to the other test findings. We estimated sensitivity, specificity, positive (PPV) and negative (NPV) predictive value, diagnostic accuracy, positive and negative likelihood ratio (LR), together with 95% confidence intervals (CI). RESULTS We enrolled 294 consecutive participants (age: Median 37.0 years, IQR: 29.0-46.0 years; men: 39.8%). Ninety-five (32.3%) participants had a positive13C-urea breath test. Twenty-three (7.8%) participants underwent upper endoscopy with histology, with a full concordance between 13C-urea breath test and histology in detecting H. pylori infection. Four (1.4%) out of the 294 participants withdrew from the study after the enrollment visit and did not undergo THD fecal testing. In the 290 participants who completed the study, the THD fecal test sensitivity was 90.2% (CI: 84.2%-96.3%), specificity 98.5% (CI:96.8%-100%), PPV 96.5% (CI: 92.6%-100%), NPV 95.6% (CI: 92.8%-98.4%), accuracy 95.9% (CI: 93.6%-98.2%), positive LR 59.5(CI: 19.3-183.4), negative LR 0.10 (CI: 0.05-0.18). Out of 83 infected participants identified with the THD fecal test, 34 (41.0%) had bacterial genotypic changes consistent with antibiotic-resistant H. pylori infection. Of these, 27 (32.5%) had bacterial strains resistant to clarithromycin, 3 (3.6%) to levofloxacin, and 4 (4.8%) to both antibiotics. CONCLUSION The THD fecal test has high performance for the non-invasive diagnosis of H. pylori infection while additionally enabling the assessment of bacterial antibiotic resistances.
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Affiliation(s)
- Andrea Iannone
- Section of Gastroenterology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
- Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
| | | | - Francesco Russo
- National Institute of Gastroenterology, “S De Bellis” Research Hospital, CastellanaGrotte (Bari) 70013, Italy
| | - Giuseppe Riezzo
- National Institute of Gastroenterology, “S De Bellis” Research Hospital, CastellanaGrotte (Bari) 70013, Italy
| | | | | | - Suetonia C Palmer
- Department of Medicine, University of Otago Christchurch, Christchurch 8011, New Zealand
| | - Michele Barone
- Section of Gastroenterology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
| | - Mariabeatrice Principi
- Section of Gastroenterology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
| | - Giovanni FM Strippoli
- Diaverum Academy, Lund 22229, Sweden
- Diaverum Medical Scientific Office, Lund 22229, Sweden
- Sydney School of Public Health, University of Sydney, Sydney NSW-2000, Australia
- Section of Nephrology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
| | - Alfredo Di Leo
- Section of Gastroenterology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
| | - Enzo Ierardi
- Section of Gastroenterology, Department of Emergency and Organ Transplantation, University of Bari, Bari 70124, Italy
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Wang YH, Li Z, Wang L, Zhu-Ge LY, Zhao RL, Wu S, Wang Y, An Y, Xie Y. A systematic review and meta-analysis of genotypic methods for detecting antibiotic resistance in Helicobacter pylori. Helicobacter 2018; 23:e12467. [PMID: 29405526 DOI: 10.1111/hel.12467] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
BACKGROUND Antibiotic susceptibility testing is essential for tailored treatments to cure Helicobacter pylori (H. pylori) infection. However, phenotypic methods have some limitations. OBJECTIVES To evaluate the feasibility of genotypic detection methods compared with phenotypic detection methods using samples taken from H. pylori-infected patients. METHODS Literature searches were conducted in the following databases (from January 2000 to November 2016): PubMed, Embase, the Cochrane Library, and Web of Science. A meta-analysis and systematic review was performed for studies that compared genotypic methods with phenotypic methods for the detection of H. pylori antibiotic susceptibility. RESULTS This meta-analysis showed that the pooled sensitivity, specificity, and diagnostic odds ratio (DOR) for the A2142G/C and/or A2143G combination for the detection of clarithromycin resistance in the strain samples were 0.97 (95% CI: 0.94-0.99), 1.00 (95% CI: 0.99-1.00), and 13 742 (95% CI: 1708-110 554), respectively. The pooled sensitivity, specificity, and DOR for the A2142G/C and/or A2143G combination for the detection of clarithromycin resistance in biopsy samples were 0.96 (95% CI: 0.90-0.99), 0.96 (95% CI: 0.91-0.99), and 722 (95% CI: 117-4443), respectively. The summarized sensitivity, specificity, and DOR value for the ability of the genotypic methods to detect quinolone resistance in biopsy specimens were 0.97 (95% CI: 0.87-0.99), 0.99 (95% CI: 0.92-1.00), and 6042 (95% CI: 486-75 143), respectively. CONCLUSION The genotypic detection methods were reliable for the diagnosis of clarithromycin and quinolone resistance in the strain and biopsy specimens. The A2142G/C and/or A2143G combination had the best sensitivity and specificity for the detection of clarithromycin resistance.
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Affiliation(s)
- You-Hua Wang
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China
| | - Zhen Li
- Department of Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Le Wang
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China.,Jiangxi Provincial Key Laboratory of Translational Medicine and Oncology, Jiangxi Cancer Hospital, Nanchang, Jiangxi Province, China
| | - Li-Ya Zhu-Ge
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China
| | - Ru-Lin Zhao
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China.,Department of Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Shuang Wu
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China
| | - Ya Wang
- Department of Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Ying An
- Department of Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Yong Xie
- Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China
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Best LMJ, Takwoingi Y, Siddique S, Selladurai A, Gandhi A, Low B, Yaghoobi M, Gurusamy KS. Non-invasive diagnostic tests for Helicobacter pylori infection. Cochrane Database Syst Rev 2018; 3:CD012080. [PMID: 29543326 PMCID: PMC6513531 DOI: 10.1002/14651858.cd012080.pub2] [Citation(s) in RCA: 90] [Impact Index Per Article: 12.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
BACKGROUND Helicobacter pylori (H pylori) infection has been implicated in a number of malignancies and non-malignant conditions including peptic ulcers, non-ulcer dyspepsia, recurrent peptic ulcer bleeding, unexplained iron deficiency anaemia, idiopathic thrombocytopaenia purpura, and colorectal adenomas. The confirmatory diagnosis of H pylori is by endoscopic biopsy, followed by histopathological examination using haemotoxylin and eosin (H & E) stain or special stains such as Giemsa stain and Warthin-Starry stain. Special stains are more accurate than H & E stain. There is significant uncertainty about the diagnostic accuracy of non-invasive tests for diagnosis of H pylori. OBJECTIVES To compare the diagnostic accuracy of urea breath test, serology, and stool antigen test, used alone or in combination, for diagnosis of H pylori infection in symptomatic and asymptomatic people, so that eradication therapy for H pylori can be started. SEARCH METHODS We searched MEDLINE, Embase, the Science Citation Index and the National Institute for Health Research Health Technology Assessment Database on 4 March 2016. We screened references in the included studies to identify additional studies. We also conducted citation searches of relevant studies, most recently on 4 December 2016. We did not restrict studies by language or publication status, or whether data were collected prospectively or retrospectively. SELECTION CRITERIA We included diagnostic accuracy studies that evaluated at least one of the index tests (urea breath test using isotopes such as 13C or 14C, serology and stool antigen test) against the reference standard (histopathological examination using H & E stain, special stains or immunohistochemical stain) in people suspected of having H pylori infection. DATA COLLECTION AND ANALYSIS Two review authors independently screened the references to identify relevant studies and independently extracted data. We assessed the methodological quality of studies using the QUADAS-2 tool. We performed meta-analysis by using the hierarchical summary receiver operating characteristic (HSROC) model to estimate and compare SROC curves. Where appropriate, we used bivariate or univariate logistic regression models to estimate summary sensitivities and specificities. MAIN RESULTS We included 101 studies involving 11,003 participants, of which 5839 participants (53.1%) had H pylori infection. The prevalence of H pylori infection in the studies ranged from 15.2% to 94.7%, with a median prevalence of 53.7% (interquartile range 42.0% to 66.5%). Most of the studies (57%) included participants with dyspepsia and 53 studies excluded participants who recently had proton pump inhibitors or antibiotics.There was at least an unclear risk of bias or unclear applicability concern for each study.Of the 101 studies, 15 compared the accuracy of two index tests and two studies compared the accuracy of three index tests. Thirty-four studies (4242 participants) evaluated serology; 29 studies (2988 participants) evaluated stool antigen test; 34 studies (3139 participants) evaluated urea breath test-13C; 21 studies (1810 participants) evaluated urea breath test-14C; and two studies (127 participants) evaluated urea breath test but did not report the isotope used. The thresholds used to define test positivity and the staining techniques used for histopathological examination (reference standard) varied between studies. Due to sparse data for each threshold reported, it was not possible to identify the best threshold for each test.Using data from 99 studies in an indirect test comparison, there was statistical evidence of a difference in diagnostic accuracy between urea breath test-13C, urea breath test-14C, serology and stool antigen test (P = 0.024). The diagnostic odds ratios for urea breath test-13C, urea breath test-14C, serology, and stool antigen test were 153 (95% confidence interval (CI) 73.7 to 316), 105 (95% CI 74.0 to 150), 47.4 (95% CI 25.5 to 88.1) and 45.1 (95% CI 24.2 to 84.1). The sensitivity (95% CI) estimated at a fixed specificity of 0.90 (median from studies across the four tests), was 0.94 (95% CI 0.89 to 0.97) for urea breath test-13C, 0.92 (95% CI 0.89 to 0.94) for urea breath test-14C, 0.84 (95% CI 0.74 to 0.91) for serology, and 0.83 (95% CI 0.73 to 0.90) for stool antigen test. This implies that on average, given a specificity of 0.90 and prevalence of 53.7% (median specificity and prevalence in the studies), out of 1000 people tested for H pylori infection, there will be 46 false positives (people without H pylori infection who will be diagnosed as having H pylori infection). In this hypothetical cohort, urea breath test-13C, urea breath test-14C, serology, and stool antigen test will give 30 (95% CI 15 to 58), 42 (95% CI 30 to 58), 86 (95% CI 50 to 140), and 89 (95% CI 52 to 146) false negatives respectively (people with H pylori infection for whom the diagnosis of H pylori will be missed).Direct comparisons were based on few head-to-head studies. The ratios of diagnostic odds ratios (DORs) were 0.68 (95% CI 0.12 to 3.70; P = 0.56) for urea breath test-13C versus serology (seven studies), and 0.88 (95% CI 0.14 to 5.56; P = 0.84) for urea breath test-13C versus stool antigen test (seven studies). The 95% CIs of these estimates overlap with those of the ratios of DORs from the indirect comparison. Data were limited or unavailable for meta-analysis of other direct comparisons. AUTHORS' CONCLUSIONS In people without a history of gastrectomy and those who have not recently had antibiotics or proton ,pump inhibitors, urea breath tests had high diagnostic accuracy while serology and stool antigen tests were less accurate for diagnosis of Helicobacter pylori infection.This is based on an indirect test comparison (with potential for bias due to confounding), as evidence from direct comparisons was limited or unavailable. The thresholds used for these tests were highly variable and we were unable to identify specific thresholds that might be useful in clinical practice.We need further comparative studies of high methodological quality to obtain more reliable evidence of relative accuracy between the tests. Such studies should be conducted prospectively in a representative spectrum of participants and clearly reported to ensure low risk of bias. Most importantly, studies should prespecify and clearly report thresholds used, and should avoid inappropriate exclusions.
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Affiliation(s)
- Lawrence MJ Best
- Royal Free Campus, UCL Medical SchoolDepartment of SurgeryRowland Hill StreetLondonUKNW32PF
| | - Yemisi Takwoingi
- University of BirminghamInstitute of Applied Health ResearchEdgbastonBirminghamUKB15 2TT
| | | | | | | | | | - Mohammad Yaghoobi
- McMaster University and McMaster University Health Sciences CentreDivision of Gastroenterology1200 Main Street WestHamiltonONCanada
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Simultaneous detection of human CYP2C19 polymorphisms and antibiotic resistance of Helicobacter pylori using a personalised diagnosis kit. J Glob Antimicrob Resist 2018; 13:174-179. [PMID: 29444465 DOI: 10.1016/j.jgar.2017.12.018] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2017] [Revised: 12/22/2017] [Accepted: 12/30/2017] [Indexed: 02/08/2023] Open
Abstract
OBJECTIVES A personalised diagnosis kit for Helicobacter pylori that employs visual gene chip technology for the simultaneous detection of CYP2C19 polymorphisms and clarithromycin/levofloxacin antibiotic resistance was evaluated. METHODS Gastric antrum mucosa biopsy specimens of 394 patients were tested using the kit. DNA sequencing and antibiotic susceptibility testing of the H. pylori were also performed. RESULTS In total, 267 (67.8%) of the 394 specimens were positive for H. pylori using the kit and DNA sequencing, and 136 (34.5%) were positive by culturing. For human CYP2C19 and the bacterial 23S rRNA and gyrA genes, the concordance rates were 92.4% (364/394), 96.6% (258/267) and 97.0% (259/267) between the kit and DNA sequencing results, respectively. For clarithromycin and levofloxacin resistance, the concordance rates were 90.4% (123/136) and 81.6% (111/136) between the kit and antibiotic susceptibility testing results. CONCLUSIONS The personalised diagnosis kit for H. pylori provides useful information for the choice of proton pump inhibitor and antibiotic in combination therapy.
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Beer-Davidson G, Hindiyeh M, Muhsen K. Detection of Helicobacter pylori in stool samples of young children using real-time polymerase chain reaction. Helicobacter 2018; 23. [PMID: 29181860 DOI: 10.1111/hel.12450] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/15/2023]
Abstract
BACKGROUND The aims of this study were to develop and validate a multiplex real-time polymerase chain reaction (q-PCR) assay of Helicobacter pylori in stool samples of healthy children. Additionally, we determined the prevalence of clarithromycin resistance and cagA gene in H. pylori-positive samples. MATERIALS AND METHODS Archived stool samples from 188 children aged 6-9 years and 272 samples of 92 infants aged 2-18 months were tested for H. pylori antigens using enzyme immunoassay (EIA). A multiplex q-PCR assay was designed to detect H. pylori 16S rRNA and urease and the human RNase P gene as an internal control. Kappa coefficient was calculated to assess the agreement between q-PCR and EIA. RESULTS Laboratory validation of the q-PCR assay using quantitated H. pylori ATCC 43504 extracted DNA showed S-shaped amplification curves for all genes; the limit of detection was 1 CFU/reaction. No cross-reactivity with other bacterial pathogens was noted. Applying the multiplex q-PCR to DNA extracted from fecal samples showed clear amplification curves for urease gene, but not for 16S rRNA. The prevalence of H. pylori infection was 50% (95% CI 43%-57%) by q-PCR (urease cycle threshold <44) vs 59% (95% CI 52%-66%) by EIA. Kappa coefficient was .80 (P < .001) and .44 (P < .001) for children aged 6-9 years and 2-18 months, respectively. Sixteen samples were positive for cagA and three were positive for clarithromycin resistance mutation (A2143G) as confirmed by sequencing. CONCLUSIONS The developed q-PCR can be used as a cotechnique to enhance the accuracy of H. pylori detection in epidemiological studies and in clinical settings.
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Affiliation(s)
- Gany Beer-Davidson
- Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Musa Hindiyeh
- Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.,Molecular Diagnostic Unit, Israel Central Virology Laboratory, Chaim Sheba Medical Center, Tel-Hashomer, Israel
| | - Khitam Muhsen
- Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
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Pastukh N, Binyamin D, On A, Paritsky M, Peretz A. GenoType® HelicoDR test in comparison with histology and culture for Helicobacter pylori detection and identification of resistance mutations to clarithromycin and fluoroquinolones. Helicobacter 2017; 22. [PMID: 29058343 DOI: 10.1111/hel.12447] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
BACKGROUND There are several methods for Helicobacter pylori infection diagnosis. AIM The efficacies of three methods for H. pylori identification directly from a biopsy were compared: histology, culture, and molecular GenoType® HelicoDR test. MATERIALS & METHODS Eighty-five triplicates of stomach antrum biopsies were obtained during gastroscopy procedures for culture, histology, and molecular assay. In addition, we performed molecular identification of genes encoding resistance to clarithromycin and fluoroquinolones. RESULTS The results have shown that the most specific method with the highest number of positive specimens was by molecular kit, compared to culture and histology (94.3%, 77.1%, and 71.4%, respectively). There was a higher rate of resistance mutations to clarithromycin than to fluoroquinolones (68.26% vs 20%). The most common mutations for clarithromycin and fluoroquinolones resistance were found in alleles A2143G and N87K, respectively. The highest rate of positive specimens was identified by the molecular. DISCUSSION GenoType HelicoDR kit (94.3%), which has several advantages: direct identification, strain resistance characterization, mixture of genotypes detection, and no transport or storage limitations; thus, it is an excellent epidemiological screening tool. This work has demonstrated a lower resistance rate to fluoroquinolones; it is possible that in the investigated geographic area treatment with fluoroquinolones may be preferable to clarithromycin. GenoType® HelicoDR test eliminates the need for culture performance and susceptibility tests for several common antibiotic agents and enables optimal and specific antibiotic treatment adjustment. CONCLUSION We recommend a combination of PCR assay and bacterial culture for a quick method of screening and more efficient identification of H. pylori strains and resistance patterns.
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Affiliation(s)
- Nina Pastukh
- Clinical Microbiology Laboratory Baruch Padeh Medical Center, Poriya, affiliated with the Faculty of Medicine, Bar Ilan University, Galilee, Israel
| | - Dana Binyamin
- Clinical Microbiology Laboratory Baruch Padeh Medical Center, Poriya, affiliated with the Faculty of Medicine, Bar Ilan University, Galilee, Israel.,The Faculty of Medicine in the Galilee, Bar Ilan University, Zefat, Israel
| | - Avi On
- The Faculty of Medicine in the Galilee, Bar Ilan University, Zefat, Israel.,Pediatric Gastrointestinal Unit, Baruch Padeh Medical Center, Poriya, affiliated with the Faculty of Medicine, Bar Ilan University, Galilee, Israel
| | - Maya Paritsky
- The Faculty of Medicine in the Galilee, Bar Ilan University, Zefat, Israel.,Gastrointestinal Unit, Baruch Padeh Medical Center, Poriya, affiliated with the Faculty of Medicine, Bar Ilan University, Galilee, Israel
| | - Avi Peretz
- Clinical Microbiology Laboratory Baruch Padeh Medical Center, Poriya, affiliated with the Faculty of Medicine, Bar Ilan University, Galilee, Israel.,The Faculty of Medicine in the Galilee, Bar Ilan University, Zefat, Israel
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Khadangi F, Yassi M, Kerachian MA. Review: Diagnostic accuracy of PCR-based detection tests for Helicobacter Pylori in stool samples. Helicobacter 2017; 22. [PMID: 28961384 DOI: 10.1111/hel.12444] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
BACKGROUND Although different methods have been established to detect Helicobacter pylori (H. pylori) infection, identifying infected patients is an ongoing challenge. The aim of this meta-analysis was to provide pooled diagnostic accuracy measures for stool PCR test in the diagnosis of H. pylori infection. METHODS In this study, a systematic review and meta-analysis were carried out on various sources, including MEDLINE, Web of Sciences, and the Cochrane Library from April 1, 1999, to May 1, 2016. This meta-analysis adheres to the guidelines provided by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses report (PRISMA Statement). The clinical value of DNA stool PCR test was based on the pooled false positive, false negative, true positive, and true negative of different genes. RESULTS Twenty-six of 328 studies identified met the eligibility criteria. Stool PCR test had a performance of 71% (95% CI: 68-73) sensitivity, 96% (95% CI: 94-97) specificity, and 65.6 (95% CI: 30.2-142.5) diagnostic odds ratio (DOR) in diagnosis of H. pylori. The DOR of genes which showed the highest performance of stool PCR tests was as follows: 23S rRNA 152.5 (95% CI: 55.5-418.9), 16S rRNA 67.9 (95%CI: 6.4-714.3), and glmM 68.1 (95%CI: 20.1-231.7). CONCLUSIONS The sensitivity and specificity of stool PCR test are relatively in the same spectrum of other diagnostic methods for the detection of H. pylori infection. In descending order of significance, the most diagnostic candidate genes using PCR detection were 23S rRNA, 16S rRNA, and glmM. PCR for 23S rRNA gene which has the highest performance could be applicable to detect H. pylori infection.
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Affiliation(s)
- Fatemeh Khadangi
- Cancer Genetics Research Unit, Reza Radiotherapy and Oncology Center, Mashhad, Iran
| | - Maryam Yassi
- Cancer Genetics Research Unit, Reza Radiotherapy and Oncology Center, Mashhad, Iran
| | - Mohammad Amin Kerachian
- Cancer Genetics Research Unit, Reza Radiotherapy and Oncology Center, Mashhad, Iran.,Cancer Genetics Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.,Department of Medical Genetics, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
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Oikawa R, Watanabe Y, Miyamoto S, Sato Y, Ono S, Mabe K, Yamamoto H, Kato M, Itoh F. Enrichment of Helicobacter pylori mutant strains after eradication therapy analyzed by gastric wash-based quantitative pyrosequencing. Tumour Biol 2017; 39:1010428317734865. [PMID: 28990461 DOI: 10.1177/1010428317734865] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
The eradication of Helicobacter pylori reduces the risk of gastric cancer. A clear understanding of the factors underlying mixed infection with multiple clarithromycin-susceptible and clarithromycin-resistant H. pylori strains is necessary to design more effective therapies against H. pylori. We aimed to assess how the abundance and prevalence of H. pylori strains vary after clarithromycin-based eradication therapy. Using gastric wash samples, which represent the entire stomach, we sequentially analyzed the abundance and prevalence of H. pylori DNA by 23S ribosomal RNA pyrosequencing before and 1, 2, and 3 years after eradication therapy. Low levels of H. pylori DNA were still detectable at the first-year follow-up in all samples with negative post-treatment urea breath test results. The abundance of H. pylori DNA decreased significantly until the 2-year follow-up, but it switched to an increase at the 3-year follow-up. Importantly, the ratio of the prevalence of mutant strains to the prevalence of wild-type strains had already increased at the first-year follow-up and continued to increase, suggesting the selection and growth of clarithromycin-resistant strains during the follow-up periods. Being sensitive and representative, our assay will be useful in effectively addressing gastric cancer development by enhancing the long-term success of intervention strategies and consecutive surveillance for H. pylori eradication.
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Affiliation(s)
- Ritsuko Oikawa
- 1 Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan
| | - Yoshiyuki Watanabe
- 1 Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan.,2 Department of Internal Medicine, Kawasaki Rinko General Hospital, Kawasaki, Japan
| | - Shuichi Miyamoto
- 3 Department of Gastroenterology and Hepatology, Hokkaido University Graduate School of Medicine, Sapporo, Japan
| | - Yoshinori Sato
- 1 Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan
| | - Shoko Ono
- 4 Division of Endoscopy, Hokkaido University Hospital, Sapporo, Japan
| | - Katsuhiro Mabe
- 5 Department of Gastroenterology, National Hospital Organization Hakodate Hospital, Hakodate, Japan
| | - Hiroyuki Yamamoto
- 1 Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan
| | - Mototsugu Kato
- 5 Department of Gastroenterology, National Hospital Organization Hakodate Hospital, Hakodate, Japan
| | - Fumio Itoh
- 1 Division of Gastroenterology and Hepatology, Department of Internal Medicine, St. Marianna University School of Medicine, Kawasaki, Japan
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Osaki T, Mabe K, Zaman C, Yonezawa H, Okuda M, Amagai K, Fujieda S, Goto M, Shibata W, Kato M, Kamiya S. Usefulness of detection of clarithromycin-resistant Helicobacter pylori from fecal specimens for young adults treated with eradication therapy. Helicobacter 2017; 22. [PMID: 28544222 DOI: 10.1111/hel.12396] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
BACKGROUND To prevent Helicobacter pylori infection in the younger generation, it is necessary to investigate the prevalence of antibiotic-resistant H. pylori. OBJECTIVE The aim of this study was to evaluate the method of PCR-based sequencing to detect clarithromycin (CAM) resistance-associated mutations using fecal samples as a noninvasive method. METHODS DNA extracted from fecal specimens and isolates from gastric biopsy specimens were collected from patients with H. pylori infection. Antibiotic resistance to CAM was analyzed by molecular and culture methods. The detection rates of CAM resistance-associated mutations (A2142C or A2143G) were compared before and after eradication therapy. RESULTS With CAM resistance of H. pylori evaluated by antibiotic susceptibility test as a gold standard, the sensitivity and the specificity of gene mutation detection from fecal DNA were 80% and 84.8%, respectively. In contrast, using DNA of isolated strains, the sensitivity and the specificity were 80% and 100%. Of the seven cases in which eradication was unsuccessful by triple therapy including CAM, CAM-resistant H. pylori, and resistance-associated mutations were detected in three cases, CAM-resistant H. pylori without the mutation was detected in two patients, and resistance-associated mutation was only detected in one patient. CONCLUSION PCR-based sequencing to detect CAM resistance-associated mutations using isolates or fecal samples was useful for finding antibiotic-resistant H. pylori infection. Although the specificity of the detection from fecal samples compared with antibiotic susceptibility testing was lower than that from isolates, this fecal detection method is suitable especially for asymptomatic subjects including children. Further improvement is needed before clinical application.
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Affiliation(s)
- Takako Osaki
- Department of Infectious Diseases, Kyorin University School of Medicine, Mitaka, Tokyo, Japan
| | - Katsuhiro Mabe
- Department of Gastroenterology, National Hospital Organization Hakodate Hospital, Hakodate, Japan.,Division of Endoscopy, Hokkaido University Hospital, Sapporo, Japan
| | - Cynthia Zaman
- Department of Infectious Diseases, Kyorin University School of Medicine, Mitaka, Tokyo, Japan
| | - Hideo Yonezawa
- Department of Infectious Diseases, Kyorin University School of Medicine, Mitaka, Tokyo, Japan
| | - Masumi Okuda
- Department of General Medicine and Community Health Science, Hyogo College of Medicine, Sasayama, Hyogo, Japan
| | - Kenji Amagai
- Ibaraki Prefectural Central Hospital, Ibaraki, Japan
| | | | | | - Wataru Shibata
- Department of Gastroenterology, Yokohama City University, Yokohama, Japan
| | - Mototsugu Kato
- Department of Gastroenterology, National Hospital Organization Hakodate Hospital, Hakodate, Japan.,Division of Endoscopy, Hokkaido University Hospital, Sapporo, Japan
| | - Shigeru Kamiya
- Department of Infectious Diseases, Kyorin University School of Medicine, Mitaka, Tokyo, Japan
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Binyamin D, Pastukh N, On A, Paritsky M, Peretz A. Phenotypic and genotypic correlation as expressed in Helicobacter pylori resistance to clarithromycin and fluoroquinolones. Gut Pathog 2017. [DOI: 10.1186/s13099-017-0198-5] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
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Ierardi E, Giorgio F, Iannone A, Losurdo G, Principi M, Barone M, Pisani A, Di Leo A. Noninvasive molecular analysis of Helicobacter pylori: Is it time for tailored first-line therapy? World J Gastroenterol 2017; 23:2453-2458. [PMID: 28465629 PMCID: PMC5394508 DOI: 10.3748/wjg.v23.i14.2453] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/16/2016] [Revised: 02/21/2017] [Accepted: 03/20/2017] [Indexed: 02/06/2023] Open
Abstract
The main problem of Helicobacter pylori (H. pylori) infection management is linked to antibiotic resistances. This phenomenon has grown in the last decade, inducing a dramatic decline in conventional regimen effectiveness. The causes of resistance are point mutations in bacterial DNA, which interfere with antibiotic mechanism of action, especially clarithromycin and levofloxacin. Therefore, international guidelines have recently discouraged their use in areas with a relevant resistance percentage, suggesting first-line schedules with expected high eradication rates, i.e., bismuth containing or non-bismuth quadruple therapies. These regimens require the daily assumption of a large number of tablets. Consequently, a complete adherence is expected only in subjects who may be motivated by the presence of major disorders. However, an incomplete adherence to antibiotic therapies may lead to resistance onset, since sub-inhibitory concentrations could stimulate the selection of resistant mutants. Of note, a recent meta-analysis suggests that susceptibility tests may be more useful for the choice of first than second-line or rescue treatment. Additionally, susceptibility guided therapy has been demonstrated to be highly effective and superior to empiric treatments by both meta-analyses and recent clinical studies. Conventional susceptibility test is represented by culture and antibiogram. However, the method is not available everywhere mainly for methodology-related factors and fails to detect hetero-resistances. Polymerase chain reaction (PCR)-based, culture-free techniques on gastric biopsy samples are accurate in finding even minimal traces of genotypic resistant strains and hetero-resistant status by the identification of specific point mutations. The need for an invasive endoscopic procedure has been the most important limit to their spread. A further step has, moreover, been the detection of point mutations in bacterial DNA fecal samples. Few studies on clarithromycin susceptibility have shown an overall high sensitivity and specificity when compared with culture or PCR on gastric biopsies. On these bases, two commercial tests are now available although they have shown some controversial findings. A novel PCR method showed a full concordance between tissue and stool results in a preliminary experience. In conclusion, despite poor validation, there is increasing evidence of a potential availability of noninvasive investigations able to detect H. pylori resistances to antibiotics. These kinds of analysis are currently at a very early phase of development and caution should be paid about their clinical application. Only further studies aimed to evaluate their sensitivity and specificity will afford novel data for solid considerations. Nevertheless, noninvasive molecular tests may improve patient compliance, time/cost of infection management and therapeutic outcome. Moreover, the potential risk of a future increase of resistance to quadruple regimens as a consequence of their use on large scale and incomplete patient adherence could be avoided.
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Brennan DE, Omorogbe J, Hussey M, Tighe D, Holleran G, O’Morain C, Smith SM, McNamara D. Molecular detection of Helicobacter pylori antibiotic resistance in stool vs biopsy samples. World J Gastroenterol 2016; 22:9214-9221. [PMID: 27895408 PMCID: PMC5107602 DOI: 10.3748/wjg.v22.i41.9214] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/09/2016] [Revised: 09/27/2016] [Accepted: 10/19/2016] [Indexed: 02/06/2023] Open
Abstract
AIM To compare (1) demographics in urea breath test (UBT) vs endoscopy patients; and (2) the molecular detection of antibiotic resistance in stool vs biopsy samples. METHODS Six hundred and sixteen adult patients undergoing endoscopy or a UBT were prospectively recruited to the study. The GenoType HelicoDR assay was used to detect Helicobacter pylori (H. pylori) and antibiotic resistance using biopsy and/or stool samples from CLO-positive endoscopy patients and stool samples from UBT-positive patients. RESULTS Infection rates were significantly higher in patients referred for a UBT than endoscopy (overall rates: 33% vs 19%; treatment-naïve patients: 33% vs 14.7%, respectively). H. pylori-infected UBT patients were younger than H. pylori-infected endoscopy patients (41.4 vs 48.4 years, respectively, P < 0.005), with a higher percentage of H. pylori-infected males in the endoscopy-compared to the UBT-cohort (52.6% vs 33.3%, P = 0.03). The GenoType HelicoDR assay was more accurate at detecting H. pylori infection using biopsy samples than stool samples [98.2% (n = 54/55) vs 80.3% (n =53/66), P < 0.005]. Subset analysis using stool and biopsy samples from CLO-positive endoscopy patients revealed a higher detection rate of resistance-associated mutations using stool samples compared to biopsies. The concordance rates between stool and biopsy samples for the detection of H. pylori DNA, clarithromycin and fluoroquinolone resistance were just 85%, 53% and 35%, respectively. CONCLUSION Differences between endoscopy and UBT patients provide a rationale for non-invasive detection of H. pylori antibiotic resistance. However, the GenoType HelicoDR assay is an unsuitable approach.
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Luo XF, Jiao JH, Zhang WY, Pu HM, Qu BJ, Yang BY, Hou M, Ji MJ. Establishment of a nested-ASP-PCR method to determine the clarithromycin resistance of Helicobacter pylori. World J Gastroenterol 2016; 22:5822-5830. [PMID: 27433095 PMCID: PMC4932217 DOI: 10.3748/wjg.v22.i25.5822] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/31/2015] [Revised: 04/08/2016] [Accepted: 04/20/2016] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate clarithromycin resistance positions 2142, 2143 and 2144 of the 23SrRNA gene in Helicobacter pylori (H. pylori) by nested-allele specific primer-polymerase chain reaction (nested-ASP-PCR).
METHODS: The gastric tissue and saliva samples from 99 patients with positive results of the rapid urease test (RUT) were collected. The nested-ASP-PCR method was carried out with the external primers and inner allele-specific primers corresponding to the reference strain and clinical strains. Thirty gastric tissue and saliva samples were tested to determine the sensitivity of nested-ASP-PCR and ASP-PCR methods. Then, clarithromycin resistance was detected for 99 clinical samples by using different methods, including nested-ASP-PCR, bacterial culture and disk diffusion.
RESULTS: The nested-ASP-PCR method was successfully established to test the resistance mutation points 2142, 2143 and 2144 of the 23SrRNA gene of H. pylori. Among 30 samples of gastric tissue and saliva, the H. pylori detection rate of nested-ASP-PCR was 90% and 83.33%, while the detection rate of ASP-PCR was just 63% and 56.67%. Especially in the saliva samples, nested-ASP-PCR showed much higher sensitivity in H. pylori detection and resistance mutation rates than ASP-PCR. In the 99 RUT-positive gastric tissue and saliva samples, the H. pylori-positive detection rate by nested-ASP-PCR was 87 (87.88%) and 67 (67.68%), in which there were 30 wild-type and 57 mutated strains in gastric tissue and 22 wild-type and 45 mutated strains in saliva. Genotype analysis showed that three-points mixed mutations were quite common, but different resistant strains were present in gastric mucosa and saliva. Compared to the high sensitivity shown by nested-ASP-PCR, the positive detection of bacterial culture with gastric tissue samples was 50 cases, in which only 26 drug-resistant strains were found through analyzing minimum inhibitory zone of clarithromycin.
CONCLUSION: The nested-ASP-PCR assay showed higher detection sensitivity than ASP-PCR and drug sensitivity testing, which could be performed to evaluate clarithromycin resistance of H. pylori.
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Lim SG, Park RW, Shin SJ, Yoon D, Kang JK, Hwang JC, Kim SS, Kim JH, Lee KM. The relationship between the failure to eradicate Helicobacter pylori and previous antibiotics use. Dig Liver Dis 2016; 48:385-90. [PMID: 26856963 DOI: 10.1016/j.dld.2015.12.001] [Citation(s) in RCA: 45] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/07/2015] [Revised: 11/30/2015] [Accepted: 12/03/2015] [Indexed: 12/11/2022]
Abstract
BACKGROUND The previous use of antibiotics is known to correlate positively with antibiotic resistance; whether this is also the case in the eradication of Helicobacter pylori infection is unclear. AIM To investigate the relationship between the previous use of antibiotics and the failure of eradication therapy in H. pylori infection. METHODS The relationship between the clinical parameters and the failure of H. pylori eradication was analyzed in patients administered standard triple therapy and then assessed for the eradication of H. pylori based on a C13-urea breath test. RESULTS In a multivariate analysis, failure rates increased significantly in patients with a history of clarithromycin (odds ratio [OR], 4.445) or other macrolides (OR, 2.407) use, who were female (OR, 1.339), or who were older than 60 years of age (OR, 1.326). The eradication failure rate in patients with a history of macrolides use for >2 weeks was significantly higher than if the duration of use was <2 weeks (44.8% vs. 29.3%, p=0.047). CONCLUSIONS A patient's history of macrolides is a useful predictor of the likelihood of standard triple therapy failure in H. pylori eradication. The alternatives such as a bismuth-based quadruple or a levofloxacin-containing therapy should be considered in patients treated with macrolides for >2 weeks.
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Affiliation(s)
- Sun Gyo Lim
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Rae Woong Park
- Department of Biomedical Informatics, Ajou University School of Medicine, Suwon, South Korea
| | - Sung Jae Shin
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Dukyong Yoon
- Department of Biomedical Informatics, Ajou University School of Medicine, Suwon, South Korea
| | - Joon Koo Kang
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Jae Chul Hwang
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Soon Sun Kim
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Jin Hong Kim
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea
| | - Kee Myung Lee
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, South Korea.
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Xuan SH, Wu LP, Zhou YG, Xiao MB. Detection of clarithromycin-resistant Helicobacter pylori in clinical specimens by molecular methods: A review. J Glob Antimicrob Resist 2016; 4:35-41. [PMID: 27436390 DOI: 10.1016/j.jgar.2016.01.002] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2015] [Revised: 01/06/2016] [Accepted: 01/08/2016] [Indexed: 12/17/2022] Open
Abstract
Various molecular methods have been developed to rapidly detect clarithromycin (CLR) resistance in Helicobacter pylori isolates in clinical specimens. All of these assays for detecting CLR resistance in H. pylori are based on detection of mutations in the 23S rRNA gene. In this article, we summarise current knowledge regarding the detection of H. pylori CLR resistance in clinical specimens by molecular tests. The available data showed that restriction fragment length polymorphism (RFLP), 3'-mismatch PCR, DNA sequencing, the PCR line probe assay (PCR-LiPA) and fluorescence in situ hybridisation assay (FISH) are able to detect CLR-resistant H. pylori in clinical specimens with excellent specificity and sensitivity. However, several factors limit their clinical application, including fastidious, time-consuming preparation and low-throughput as well as carrying a risk of contamination. Furthermore, as an invasive method, FISH is not suitable for children or the elderly. Among the molecular methods, one that is most promising for the future is real-time PCR probe hybridisation technology using fluorescence resonance energy transfer (FRET) probes, which can rapidly detect CLR resistance with high sensitivity and specificity in biopsies and stool specimens, even though mixed infections are present in clinical specimens. Moreover, due to the advantages that this method is simple, rapid and economical, real-time PCR is technically feasible for clinical application in small- and medium-sized hospitals in developing countries. Second, with high sensitivity, specificity and throughput, DNA chips will also be a valuable tool for detecting resistant H. pylori isolates from cultures and clinical specimens.
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Affiliation(s)
- Shi-Hai Xuan
- Department of Clinical Laboratory, The Affiliated Dongtai Hospital of Nantong University, Dongtai 224200, China
| | - Li-Pei Wu
- Department of Clinical Laboratory, The Affiliated Dongtai Hospital of Nantong University, Dongtai 224200, China
| | - Yu-Gui Zhou
- Department of Clinical Laboratory, The Affiliated Dongtai Hospital of Nantong University, Dongtai 224200, China
| | - Ming-Bing Xiao
- Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong 226001, China.
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Mégraud F, Bénéjat L, Ontsira Ngoyi EN, Lehours P. Molecular Approaches to Identify Helicobacter pylori Antimicrobial Resistance. Gastroenterol Clin North Am 2015; 44:577-96. [PMID: 26314669 DOI: 10.1016/j.gtc.2015.05.002] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/21/2023]
Abstract
Antimicrobial susceptibility testing is needed to adapt Helicobacter pylori treatment to obtain the best results. Beside the standard phenotypic methods, molecular methods are increasingly used. The value of these molecular tests is that they are quick, independent of the transport conditions, easy to standardize, and commercial kits are available. In this article, these methods are reviewed, focusing on the determination of H pylori resistance to macrolides and fluoroquinolones, and mentioning also the methods used for tetracycline and rifampin.
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Affiliation(s)
- Francis Mégraud
- Bacteriology Laboratory, INSERM U853, University of Bordeaux, Bordeaux F-33000, France.
| | - Lucie Bénéjat
- Bacteriology Laboratory, INSERM U853, University of Bordeaux, Bordeaux F-33000, France
| | | | - Philippe Lehours
- Bacteriology Laboratory, INSERM U853, University of Bordeaux, Bordeaux F-33000, France
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Yonezawa H, Osaki T, Hanawa T, Kurata S, Ochiai K, Kamiya S. Impact of Helicobacter pylori biofilm formation on clarithromycin susceptibility and generation of resistance mutations. PLoS One 2013; 8:e73301. [PMID: 24039906 PMCID: PMC3765302 DOI: 10.1371/journal.pone.0073301] [Citation(s) in RCA: 59] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2013] [Accepted: 07/18/2013] [Indexed: 12/11/2022] Open
Abstract
The human gastric pathogen Helicobacter pylori forms biofilms in vitro and in vivo. The purpose of this study was to evaluate the effects of H. pylori biofilm formation in vitro on clarithromycin (CLR) susceptibility. CLR susceptibility of H. pylori intermediate (2-day) and mature (3-day) biofilms on glass coverslips was determined at concentrations from 0.03 to 0.5 µg/ml. H. pylori biofilm biomass was increased after treatment with CLR at minimum inhibitory concentration levels by up to 4-fold (2-day biofilm) and 16-fold (3-day biofilm). Minimum bactericidal concentrations of CLR against cells in a biofilm were higher (1.0 µg/ml) than that for planktonic cells (0.25 µg/ml). It was shown that the expression of efflux pump genes was significantly increased in biofilm cells. In addition, exposure of biofilms to CLR resulted in high level resistance generation compared to planktonic cells with increased resistance associated with the presence of a point mutation at either position 2142 or 2143 in the domain V loop of the 23S rRNA gene. These results demonstrate that H. pylori biofilm formation decreases the susceptibility to CLR and that H. pylori CLR resistance mutations are more frequently generated in biofilms than in planktonic cells.
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Affiliation(s)
- Hideo Yonezawa
- Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan
- * E-mail:
| | - Takako Osaki
- Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan
| | - Tomoko Hanawa
- Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan
| | - Satoshi Kurata
- Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan
| | - Kuniyasu Ochiai
- Department of Microbiology, Nihon University School of Dentistry, Tokyo, Japan
| | - Shigeru Kamiya
- Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan
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Xiong LJ, Tong Y, Wang Z, Mao M. Detection of clarithromycin-resistant Helicobacter pylori by stool PCR in children: a comprehensive review of literature. Helicobacter 2013; 18:89-101. [PMID: 23067446 DOI: 10.1111/hel.12016] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
BACKGROUND Helicobacter pylori infection is acquired mainly during childhood. To eradicate H. pylori, clarithromycin-based triple therapy has been recommended in children and adults by the latest Maastricht Consensus. However, the prevalence of clarithromycin-resistant H. pylori was higher in children than that in adults. Therefore, rapid, reliable and noninvasive methods for detecting clarithromycin-resistant H. pylori strains should be developed for children. MATERIALS AND METHODS Studies on evaluating stool PCR in detecting clarithromycin-resistant H. pylori and epidemiological surveys of the prevalence of clarithromycin-resistant H. pylori in children were searched in PubMed (from 1966 to December, 2011) for reviewing. RESULTS The average rates of primary clarithromycin-resistant H. pylori ranged from less than 10% to more than 40% in different regions. The rates of secondary resistance to clarithromycin were higher than primary resistance in the same population. In H. pylori isolated from children, the frequent point mutations that are responsible for the clarithromycin resistance included A2143G, A2142G, A2142C and A2144G, and they varied geographically. Comparing with culture-based susceptibility tests, stool PCR performed excellently for their rapidity, independence of bacterial growth, reproducibility and easy standardization. However, stool PCR showed lower sensitivity but perfect specificity in detection of clarithromycin-resistant H. pylori in children. Methodology and mixed infections of resistant H. pylori strains might contribute to the considerable discrepancies of stool PCR results. CONCLUSION Detection of clarithromycin-resistant H. pylori by stool PCR for children are reliable, rapid, noninvasive methods that are worthy of further clinical promotion. However, more evaluations of stool PCR in detection of clarithromycin-resistant H. pylori in children need to be conducted.
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Affiliation(s)
- Li Jing Xiong
- Department of Pediatrics, West China Second University Hospital, Sichuan University, Chengdu, China
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Abstract
Helicobacter pylori is an important pathogen whose primary niche is the human stomach. H. pylori is etiologically associated with gastric inflammation (gastritis), peptic ulcer disease, and gastric cancer. Both noninvasive (e.g., urea breath and stool antigen tests) and invasive (gastric biopsy for histology, culture, or PCR) tests are used for diagnosis. PCR detection of H. pylori has been reported using a variety of clinical samples including gastric biopsy, gastric juice, saliva, dental plaque, and stools as well as environmental samples. Whenever possibly, noninvasive tests are preferred over invasive tests. H. pylori are excreted in the stool. Culture from stool is variable whereas stool antigen testing is widely used. Stool consists of a complicated mixture of commensal bacteria and chemicals and often includes inhibitors of PCR. Nevertheless, simple extraction methods are available to efficiently extract DNA from human stools and nested-PCR targeting the 23S rRNA gene have proven to be highly sensitive for the detection of H. pylori. Detection of clarithromycin susceptibility/resistance is important clinically and the mutation of the 23S rRNA gene responsible for resistance can also be detected using stool. This described method can be modified for other clinical samples such as gastric juice or biopsy material.
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Affiliation(s)
- Emiko Rimbara
- Department of Bacteriology II, National Institute of Infectious Diseases, Tokyo, USA
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Deguchi R, Nakaminami H, Rimbara E, Noguchi N, Sasatsu M, Suzuki T, Matsushima M, Koike J, Igarashi M, Ozawa H, Fukuda R, Takagi A. Effect of pretreatment with Lactobacillus gasseri OLL2716 on first-line Helicobacter pylori eradication therapy. J Gastroenterol Hepatol 2012; 27:888-92. [PMID: 22098133 PMCID: PMC3504346 DOI: 10.1111/j.1440-1746.2011.06985.x] [Citation(s) in RCA: 51] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
BACKGROUND AND AIM Helicobacter pylori eradication clearly decreases peptic ulcer recurrence rates. H. pylori eradication is achieved in 70-90% of cases, but treatment failures due to poor patient compliance and resistant organisms do occur. Lactobacillus gasseri can suppress both clarithromycin-susceptible and -resistant strains of H. pylori in vitro. The aim of this study was to determine the effect of pretreatment with L. gasseri- containing yogurt on H. pylori eradication. We conducted a randomized, controlled clinical trial in patients with H. pylori infection. METHODS A total of 229 patients were randomized into either a 1-week triple therapy of rabeprazole (10 mg bid), amoxicillin (750 mg bid), and clarithromycin (200 mg bid) or triple therapy plus L. gasseri-containing yogurt. In the yogurt-plus-triple therapy groups, yogurt containing L. gasseri OLL2716 (112 g) was given twice daily for 4 weeks (3 weeks pretreatment and also 1 week during eradication therapy). Clarithromycin resistance was determined by the detection of a mutation in 23S rRNA using nested polymerase chain reaction and the direct sequencing of DNA from pretreatment feces. H. pylori eradication was diagnosed based on the urea breath test and a stool antigen test after 8 weeks of eradication. RESULTS The status of H. pylori susceptibility to clarithromycin was successively determined in 188 out of 229 samples. The rate of infection with clarithromycin-resistant strains of H. pylori was 27.1%. Overall eradication (intention to treat/per protocol) was 69.3/74.5% for the triple-only group, and 82.6/85.6% for the yogurt-plus-triple group (P = 0.018/P = 0.041). Eradication of primary clarithromycin-resistant strains tended to be higher for yogurt-plus-triple therapy than triple-only therapy (38.5 vs 28.0%, respectively, P = 0.458). CONCLUSION This study confirmed that the major cause of treatment failure is resistance to clarithromycin. A 4-week treatment with L. gasseri-containing yogurt improves the efficacy of triple therapy in patients with H. pylori infection.
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Affiliation(s)
- Ryuzo Deguchi
- Gastroenterology, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Hidemasa Nakaminami
- Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life ScienceHorinouchi, Hachioji, Tokyo, Japan
| | - Emiko Rimbara
- Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life ScienceHorinouchi, Hachioji, Tokyo, Japan
| | - Norihisa Noguchi
- Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life ScienceHorinouchi, Hachioji, Tokyo, Japan
| | - Masanori Sasatsu
- Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life ScienceHorinouchi, Hachioji, Tokyo, Japan
| | - Takayoshi Suzuki
- Gastroenterology, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Masashi Matsushima
- Gastroenterology, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Jun Koike
- Gastroenterology, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Muneki Igarashi
- Gastroenterology, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Hideki Ozawa
- General Internal Medicine, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Ryuki Fukuda
- General Internal Medicine, Tokai University School of MedicineIsehara, Kanagawa, Japan
| | - Atsushi Takagi
- General Internal Medicine, Tokai University School of MedicineIsehara, Kanagawa, Japan
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Sicinschi LA, Correa P, Bravo LE, Peek RM, Wilson KT, Loh JT, Yepez MC, Gold BD, Thompson DT, Cover TL, Schneider BG. Non-invasive genotyping of Helicobacter pylori cagA, vacA, and hopQ from asymptomatic children. Helicobacter 2012; 17:96-106. [PMID: 22404439 PMCID: PMC3305281 DOI: 10.1111/j.1523-5378.2011.00919.x] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
BACKGROUND Helicobacter pylori infection is usually acquired in childhood, but little is known about its natural history in asymptomatic children, primarily due to the paucity of non-invasive diagnostic methods. H. pylori strains harboring cagA and specific alleles of hopQ and vacA are associated with increased risk for gastric cancer. Many studies of H. pylori virulence markers in children have the bias that symptomatic subjects are selected for endoscopy, and these children may harbor the most virulent strains. Our aim is to genotype cagA, hopQ, and vacA alleles in stool DNA samples of healthy Colombian children residing in an area with high incidence of gastric cancer, to avoid selection bias resulting from endoscopy. METHODS H. pylori status of 86 asymptomatic children was assessed by (13) C-urea breath test (UBT) and PCR. H. pylori 16S rRNA, cagA, hopQ, and vacA genes were amplified from stool DNA samples and sequenced. RESULTS UBT was positive in 69 (80.2%) of 86 children; in stool DNA analysis, 78.3% were positive by 16S rRNA PCR. cagA, vacA, and hopQ were detected in 66.1%, 84.6%, and 72.3% of stool DNA samples from 16S rRNA-positive children. Of the children's DNA samples, which revealed vacA and hopQ alleles, 91.7% showed vacA s1 and 73.7% showed type I hopQ. Type I hopQ alleles were associated with cagA positivity and vacA s1 genotypes (p < 0.0001). CONCLUSIONS Using stool DNA samples, virulence markers of H. pylori were successfully genotyped in a high percentage of the asymptomatic infected children, revealing a high prevalence of genotypes associated with virulence. Type I hopQ alleles were associated with the presence of cagA and the vacA s1 genotype.
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Affiliation(s)
- Liviu A. Sicinschi
- Division of Gastroenterology, Dept. of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232,Department of Microbiology and Immunology, Holmes Regional Medical Center, Melbourne, FL 32901, USA
| | - Pelayo Correa
- Division of Gastroenterology, Dept. of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Luis E. Bravo
- Department of Pathology, School of Medicine, Universidad del Valle, Pasto, Colombia
| | - Richard M. Peek
- Division of Gastroenterology, Dept. of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232
| | - Keith T. Wilson
- Division of Gastroenterology, Dept. of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232,Veterans Affairs Tennessee Valley Health Care System, Nashville, TN 37212, USA
| | - John T. Loh
- Division of Infectious Diseases, Department of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
| | - Maria C. Yepez
- Centro de Estudios de Salud, Universidad de Nariño, Pasto, Colombia
| | - Benjamin D. Gold
- Children's Center for Digestive Healthcare, Pediatric Gastroenterology, Hepatology and Nutrition, Atlanta, GA 30342
| | - Dexter T. Thompson
- Division of Gastroenterology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA
| | - Timothy L. Cover
- Division of Infectious Diseases, Department of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232, USA,Department of Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA,Veterans Affairs Tennessee Valley Health Care System, Nashville, TN 37212, USA
| | - Barbara G. Schneider
- Division of Gastroenterology, Dept. of Medicine, Vanderbilt University School of Medicine, Nashville, TN 37232
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Takiue K, Sugiyama H, Inoue T, Morinaga H, Kikumoto Y, Kitagawa M, Kitamura S, Maeshima Y, Wang DH, Masuoka N, Ogino K, Makino H. Acatalasemic mice are mildly susceptible to adriamycin nephropathy and exhibit increased albuminuria and glomerulosclerosis. BMC Nephrol 2012; 13:14. [PMID: 22443450 PMCID: PMC3329410 DOI: 10.1186/1471-2369-13-14] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2011] [Accepted: 03/25/2012] [Indexed: 11/30/2022] Open
Abstract
Background Catalase is an important antioxidant enzyme that regulates the level of intracellular hydrogen peroxide and hydroxyl radicals. The effects of catalase deficiency on albuminuria and progressive glomerulosclerosis have not yet been fully elucidated. The adriamycin (ADR) nephropathy model is considered to be an experimental model of focal segmental glomerulosclerosis. A functional catalase deficiency was hypothesized to exacerbate albuminuria and the progression of glomerulosclerosis in this model. Methods ADR was intravenously administered to both homozygous acatalasemic mutant mice (C3H/AnLCsbCsb) and control wild-type mice (C3H/AnLCsaCsa). The functional and morphological alterations of the kidneys, including albuminuria, renal function, podocytic, glomerular and tubulointerstitial injuries, and the activities of catalase were then compared between the two groups up to 8 weeks after disease induction. Moreover, the presence of a mutation of the toll-like receptor 4 (tlr4) gene, which was previously reported in the C3H/HeJ strain, was investigated in both groups. Results The ADR-treated mice developed significant albuminuria and glomerulosclerosis, and the degree of these conditions in the ADR-treated acatalasemic mice was higher than that in the wild-type mice. ADR induced progressive renal fibrosis, renal atrophy and lipid peroxide accumulation only in the acatalasemic mice. In addition, the level of catalase activity was significantly lower in the kidneys of the acatalasemic mice than in the wild-type mice during the experimental period. The catalase activity increased after ADR injection in wild-type mice, but the acatalasemic mice did not have the ability to increase their catalase activity under oxidative stress. The C3H/AnL strain was found to be negative for the tlr4 gene mutation. Conclusions These data indicate that catalase deficiency plays an important role in the progression of renal injury in the ADR nephropathy model.
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Affiliation(s)
- Keiichi Takiue
- Department of Medicine and Clinical Science, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikatacho, Kita-ku, Okayama 700-8558, Japan
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Lins AK, Lima RA, Magalhães M. Clarithromycin-resistant Helicobacter pylori in Recife, Brazil, directly identified from gastric biopsies by polymerase chain reaction. ARQUIVOS DE GASTROENTEROLOGIA 2011; 47:379-82. [PMID: 21225149 DOI: 10.1590/s0004-28032010000400011] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/23/2009] [Accepted: 07/20/2010] [Indexed: 12/12/2022]
Abstract
CONTEXT Clarithromycin is the most effective drug used in the eradication of infection by Helicobacter pylori. Due to worldwide increase in resistance, pre-treatment susceptibility testing for clarithromycin is recommended. OBJECTIVES To evaluate the prevalence of clarithromycin resistance of H. pylori in Recife, a city in Northeast Brazil. METHODS From January 2006 to December 2007, 114 gastric biopsy samples positive for H. pylori at culture were directly assayed by polymerase chain reaction (PCR) to detect the most frequent point mutations involved in clarithromycin resistance. Results were compared with those obtained by Etests. RESULT Molecular and phenotypic methods showed 111 (97.4%) susceptible or resistant concordant results. PCR detected 3 (2.6%) biopsy specimens with H. pylori-resistant genotypes, which were misdiagnosed as susceptible by Etests. In Recife, based on PCR results, primary clarithromycin resistance was found in 15 (16.5%) patients, prevalence close to that observed in Southeast Brazil. Resistance increased to 52% among previously treated patients. The point mutation A2143G was present in 20 (71.4%) of specimens and A2142G, in 8 (28.6%) of specimens. A2142C was not found. CONCLUSION In Recife, the prevalence of primary clarithromycin resistance, 16.5%, showed the need for pretreatment susceptibility testing in H. pylori infections.
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