scholarly journals Rapid Detection of Quinolone Resistance Mutations in gyrA of Helicobacter pylori by Real-Time PCR

Pathogens ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 59
Author(s):  
Franziska Haumaier ◽  
Anna Schneider-Fuchs ◽  
Steffen Backert ◽  
Michael Vieth ◽  
William Sterlacci ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Real Time ◽  
Melting Curve ◽  
Quinolone Resistance ◽  
Second Line ◽  
Wild Type ◽  
Resistance Mutations ◽  
Gyra Gene ◽  
Line Therapy ◽  
H Pylori

The treatment of infections by the gastric pathogen Helicobacter pylori (H. pylori) has become more difficult due to increased rates of resistances against various antibiotics. Typically, atriple therapy, employing a combination of at least two antibiotics and a proton pump inhibitor, is used to cure H. pylori infections. In case of first-line therapy failure, quinolones are commonly applied in a second-line therapy. To prevent second-line treatment failures, we developed an improved method to detect the most common quinolone-resistance mutations located in the quinolone-resistance-determining region (QRDR) of the bacterial gyrA gene. Biopsy material from the gastric mucosa of infected patients was used to identify quinolone-resistant strains before the onset of drug administration. Two different wild-type and six mutant QRDR sequences were included. Melting curve analyses were performed with corresponding gyrA plasmid DNAs using a real-time polymerase chain reaction (RT-PCR) assay. By applying a combination of only two different fluorescent probes, this assay allows wild-type sequences to be unambiguously distinguished from all known mutant QRDR sequences of H. pylori. Next, the Tm values of patient DNAs were established, and the genotypes were confirmed by sequencing. Thus, quinolone-resistant H. pylori strains can be easily and quickly diagnosed before treatment, which will help to avoid the administration of ineffective drug regimes.

scholarly journals Helicobacter pylori HP1512 Is a Nickel-Responsive NikR-Regulated Outer Membrane Protein

2006 ◽  
Vol 74 (12) ◽  
pp. 6811-6820 ◽  
Author(s):  
Gregg S. Davis ◽  
Erika L. Flannery ◽  
Harry L. T. Mobley
Keyword(s):  
Helicobacter Pylori ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Real Time ◽  
Outer Membrane Protein ◽  
Intergenic Region ◽  
Calf Serum ◽  
Nickel Concentration ◽  
Wild Type ◽  
H Pylori

ABSTRACT Helicobacter pylori is dependent upon the production of the highly abundant and active metalloenzyme urease for colonization of the human stomach. Thus, H. pylori has an absolute requirement for the transition metal nickel, a required cofactor for urease. To investigate the contribution of genes that are factors in this process, microarray analysis comparing the transcriptome of wild-type H. pylori 26695 cultured in brucella broth containing fetal calf serum (BBF) alone or supplemented with 100 μM NiCl2 suggested that HP1512 is repressed in the presence of 100 μM supplemental nickel. When measured by comparative real-time quantitative PCR (qPCR), HP1512 transcription was reduced 43-fold relative to the value for the wild type when cultured in BBF supplemented with 10 μM NiCl2. When grown in unsupplemented BBF, urease activity of an HP1512::cat mutant was significantly reduced compared to the wild type, 4.9 ± 0.5 μmol/min/mg of protein (n = 7) and 17.1 ± 4.9 μmol/min/mg of protein (n = 13), respectively (P < 0.0001). In silico analysis of the HP1511-HP1512 (HP1511-1512) intergenic region identified a putative NikR operator upstream of HP1512. Gel shift analysis with purified recombinant NikR verified nickel-dependent binding of H. pylori NikR to the HP1511-1512 intergenic region. Furthermore, comparative real-time qPCR of four nickel-related genes suggests that mutation of HP1512 results in reduced intracellular nickel concentration relative to wild-type H. pylori 26695. Taken together, these data suggest that HP1512 encodes a NikR-nickel-regulated outer membrane protein.

scholarly journals European Registry on Helicobacter pylori Management: Effectiveness of First and Second-Line Treatment in Spain

Antibiotics ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 13
Author(s):  
María Caldas ◽  
Ángeles Pérez-Aisa ◽  
Manuel Castro-Fernández ◽  
Luis Bujanda ◽  
Alfredo Lucendo ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Second Line ◽  
Cure Rate ◽  
Management Effectiveness ◽  
First Line ◽  
Intention To Treat ◽  
First Line Therapy ◽  
Line Therapy ◽  
H Pylori ◽  
European Registry

The management of Helicobacter pylori infection has to rely on previous local effectiveness due to the geographical variability of antibiotic resistance. The aim of this study was to evaluate the effectiveness of first and second-line H. pylori treatment in Spain, where the empirical prescription is recommended. A multicentre prospective non-interventional registry of the clinical practice of European gastroenterologists concerning H. pylori infection (Hp-EuReg) was developed, including patients from 2013 until June 2019. Effectiveness was evaluated descriptively and through a multivariate analysis concerning age, gender, presence of ulcer, proton-pump inhibitor (PPI) dose, therapy duration and compliance. Overall, 53 Spanish hospitals were included, and 10,267 patients received a first-line therapy. The best results were obtained with the 10-day bismuth single-capsule therapy (95% cure rate by intention-to-treat) and with both the 14-day bismuth-clarithromycin quadruple (PPI-bismuth-clarithromycin-amoxicillin, 91%) and the 14-day non-bismuth quadruple concomitant (PPI-clarithromycin-amoxicillin-metronidazole, 92%) therapies. Second-line therapies were prescribed to 2448 patients, with most-effective therapies being the triple quinolone (PPI-amoxicillin-levofloxacin/moxifloxacin) and the bismuth-levofloxacin quadruple schemes (PPI-bismuth-levofloxacin-amoxicillin) prescribed for 14 days (92%, 89% and 90% effectiveness, respectively), and the bismuth single-capsule (10 days, 88.5%). Compliance, longer duration and higher acid inhibition were associated with higher effectiveness. “Optimized” H. pylori therapies achieve over 90% success in Spain.

Helicobacter pylori eradication rates in Slovenia in the period from 2017 to 2019 – data from the European Registry on Helicobacter pylori Management (Hp-EuReg)

2020 ◽  
Author(s):  
Bojan Tepes ◽  
Natasa Brglez Jurecic ◽  
Katja Tepes ◽  
Marta Espada Sanchez ◽  
Olga Perez Nyssen ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Triple Therapy ◽  
Second Line ◽  
First Line ◽  
Second Line Therapy ◽  
Intention To Treat ◽  
Line Therapy ◽  
Medical Institutions ◽  
H Pylori ◽  
European Registry

Abstract Background. Helicobacter pylori (H pylori) is the most common chronic bacterial infection in the world, affecting over 50% of the world’s population. H pylori is a grade I carcinogen, responsible for the development of 89% of non-cardia gastric cancers. In the present study, we analyzed the data for H pylori eradication treatments in Slovenia after the 3rd National recommendations were implemented. Patients and methods. Slovenia is part of the European Registry on H pylori Management (Hp-EuReg) since the study was launched in 2013. Data were collected at AEG-REDCap e-CRF from September 2017 to December 2019. H pylori eradication treatment was assessed by modified intention-to-treat (mITT) and per-protocol (PP) analyses. Results. Overall, 823 patients from 3 medical institutions were included. Effectiveness with first line 14-day triple therapy with a proton-pomp-inhibitor (PPI), clarithromycin 500 mg, amoxicillin 1000 mg, all BID, was 93% by mITT (714 patients). In patients allergic to penicillin, first line 14-day triple therapy with PPI-clarithromycin-metronidazole achieved 83% effectiveness by mITT (35 patients). Second-line 14-day triple therapy with a PPI-amoxicillin-levofloxacin achieved 89% mITT eradication rate (51 patients). Second-line therapy with the 10-day three-in-one single capsule containing bismuth-tetracycline- metronidazole achieved optimal effectiveness (100% mITT) in 10 patients ( p=0.02). Conclusions. Slovenia is a country with < 15% H pylori resistance to clarithromycin. Triple therapy with a PPI plus two antibiotics during 14 days reported optimal effectiveness (over 90%). Ten-day quadruple bismuth second-line therapy had better results than 14-day triple therapy with levofloxacin).

scholarly journals The Helicobacter pylori Autotransporter ImaA (HP0289) Modulates the Immune Response and Contributes to Host Colonization

2012 ◽  
Vol 80 (7) ◽  
pp. 2286-2296 ◽  
Author(s):  
William E. Sause ◽  
Andrea R. Castillo ◽  
Karen M. Ottemann
Keyword(s):  
Immune Response ◽  
Helicobacter Pylori ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Dependent Manner ◽  
Wild Type ◽  
Content Type ◽  
H Pylori ◽  
Murine Infections

ABSTRACTThe human pathogenHelicobacter pyloriemploys a diverse collection of outer membrane proteins to colonize, persist, and drive disease within the acidic gastric environment. In this study, we sought to elucidate the function of the host-induced geneHP0289, which encodes an uncharacterized outer membrane protein. We first generated an isogenicH. pylorimutant that lacksHP0289and found that the mutant has a colonization defect in single-strain infections and is greatly outcompeted in mouse coinfection experiments with wild-typeH. pylori. Furthermore, we used protease assays and biochemical fractionation coupled with an HP0289-targeted peptide antibody to verify that the HP0289 protein resides in the outer membrane. Our previous findings showed that theHP0289promoter is upregulated in the mouse stomach, and here we demonstrate thatHP0289expression is induced under acidic conditions in an ArsRS-dependent manner. Finally, we have shown that theHP0289mutant induces greater expression of the chemokine interleukin-8 (IL-8) and the cytokine tumor necrosis factor alpha (TNF-α) in gastric carcinoma cells (AGS). Similarly, transcription of the IL-8 homolog keratinocyte-derived chemokine (KC) is elevated in murine infections with the HP0289 mutant than in murine infections with wild-typeH. pylori. On the basis of this phenotype, we renamed HP0289 ImaA forimmunomodulatoryautotransporter protein. Our work has revealed that genes inducedin vivoplay an important role inH. pyloripathogenesis. Specifically, the outer membrane protein ImaA modulates a component of the host inflammatory response, and thus may allowH. pylorito fine tune the host immune response based on ImaA expression.

scholarly journals Systems Modeling of the Role of Interleukin-21 in the Maintenance of Effector CD4+ T Cell Responses during Chronic Helicobacter pylori Infection

mBio ◽  
2014 ◽  
Vol 5 (4) ◽  
Author(s):  
Adria Carbo ◽  
Danyvid Olivares-Villagómez ◽  
Raquel Hontecillas ◽  
Josep Bassaganya-Riera ◽  
Rupesh Chaturvedi ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Helicobacter Pylori ◽  
T Cell ◽  
Wild Type ◽  
Content Type ◽  
Interleukin 21 ◽  
H Pylori ◽  
Deficient Mice

ABSTRACTThe development of gastritis duringHelicobacter pyloriinfection is dependent on an activated adaptive immune response orchestrated by T helper (Th) cells. However, the relative contributions of the Th1 and Th17 subsets to gastritis and control of infection are still under investigation. To investigate the role of interleukin-21 (IL-21) in the gastric mucosa duringH. pyloriinfection, we combined mathematical modeling of CD4+T cell differentiation within vivomechanistic studies. We infected IL-21-deficient and wild-type mice withH. pyloristrain SS1 and assessed colonization, gastric inflammation, cellular infiltration, and cytokine profiles. ChronicallyH. pylori-infected IL-21-deficient mice had higherH. pyloricolonization, significantly less gastritis, and reduced expression of proinflammatory cytokines and chemokines compared to these parameters in infected wild-type littermates. Thesein vivodata were used to calibrate anH. pyloriinfection-dependent, CD4+T cell-specific computational model, which then described the mechanism by which IL-21 activates the production of interferon gamma (IFN-γ) and IL-17 during chronicH. pyloriinfection. The model predicted activated expression of T-bet and RORγt and the phosphorylation of STAT3 and STAT1 and suggested a potential role of IL-21 in the modulation of IL-10. Driven by our modeling-derived predictions, we found reduced levels of CD4+splenocyte-specifictbx21androrcexpression, reduced phosphorylation of STAT1 and STAT3, and an increase in CD4+T cell-specific IL-10 expression inH. pylori-infected IL-21-deficient mice. Our results indicate that IL-21 regulates Th1 and Th17 effector responses during chronicH. pyloriinfection in a STAT1- and STAT3-dependent manner, therefore playing a major role controllingH. pyloriinfection and gastritis.IMPORTANCEHelicobacter pyloriis the dominant member of the gastric microbiota in more than 50% of the world’s population.H. pyloricolonization has been implicated in gastritis and gastric cancer, as infection withH. pyloriis the single most common risk factor for gastric cancer. Current data suggest that, in addition to bacterial virulence factors, the magnitude and types of immune responses influence the outcome of colonization and chronic infection. This study uses a combined computational and experimental approach to investigate how IL-21, a proinflammatory T cell-derived cytokine, maintains the chronic proinflammatory T cell immune response driving chronic gastritis duringH. pyloriinfection. This research will also provide insight into a myriad of other infectious and immune disorders in which IL-21 is increasingly recognized to play a central role. The use of IL-21-related therapies may provide treatment options for individuals chronically colonized withH. pylorias an alternative to aggressive antibiotics.

scholarly journals Vertical Helicobacter pylori transmission from Mongolian gerbil mothers to pups

2009 ◽  
Vol 58 (5) ◽  
pp. 656-662 ◽  
Author(s):  
Ichiro Oshio ◽  
Takako Osaki ◽  
Tomoko Hanawa ◽  
Hideo Yonezawa ◽  
Cynthia Zaman ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Gastric Mucosa ◽  
16S Rrna ◽  
Real Time ◽  
Mongolian Gerbil ◽  
Rt Pcr ◽  
Antibody Titres ◽  
Mother's Milk ◽  
Mother’S Milk ◽  
H Pylori

To identify the time frame and route of mother-to-child Helicobacter pylori infection, a Mongolian gerbil model was used. Four-week-old female Mongolian gerbils were infected with H. pylori, and then mated with uninfected males 2 months after infection. The offspring were sacrificed weekly after birth, and then serum, mother's milk from the stomach and gastric tissues were obtained from pups. Anti-H. pylori antibody titres were measured in sera and maternal milk using an ELISA. The stomach was cut in two in the sagittal plane, and then H. pylori colonization in mucosa was confirmed by culture and real-time RT-PCR in one specimen and by immunochemical staining in the other. Faeces and oral swabs were obtained from infected mothers, and H. pylori 16S rRNA was measured using real-time RT-PCR. H. pylori was not identified in cultures from the gastric mucosa of pups delivered by infected mothers, but H. pylori 16S rRNA was detected from 4 weeks after birth, suggesting that Mongolian gerbil pups become infected via maternal H. pylori transmission from 4 weeks of age. The anti-H. pylori antibody titre in sera of pups from infected mothers was maximum at 3 weeks of age and then rapidly decreased from 4 weeks of age. High antibody titres in mother's milk were detected during the suckling period, and GlcNAcα was detectable at 2–4 weeks of age, but disappeared as the offspring aged. Thus H. pylori seems to infect Mongolian gerbil pups from 4 weeks of age, in parallel with decreasing GlcNAcα expression in the gastric mucosa. These results suggested that H. pylori infection of Mongolian gerbil pups occurs via faecal–oral transmission from an infected mother.

Determination of Helicobacter pylori cagA, vacA genotypes with real-time PCR melting curve analysis

2001 ◽  
Vol 95 (1-6) ◽  
pp. 369-377 ◽  
Author(s):  
Agnes Ruzsovics ◽  
Bela Molnar ◽  
Zsuzsa Unger ◽  
Zsolt Tulassay ◽  
Laszlo Pronai
Keyword(s):  
Helicobacter Pylori ◽  
Real Time ◽  
Real Time Pcr ◽  
Melting Curve ◽  
Curve Analysis ◽  
Melting Curve Analysis

scholarly journals Empiric “Three-in-One” Bismuth Quadruple Therapy for Second-Line Helicobacter pylori Eradication: An Intervention Study in Southern Italy

Antibiotics ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 78
Author(s):  
Giuseppe Losurdo ◽  
Ilaria Lacavalla ◽  
Francesco Russo ◽  
Giuseppe Riezzo ◽  
Irene Vita Brescia ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Adverse Events ◽  
Eradication Rate ◽  
Sequential Therapy ◽  
Second Line ◽  
Capsule Formulation ◽  
Second Line Therapy ◽  
Quadruple Therapy ◽  
Intention To Treat ◽  
Line Therapy

The eradication of Helicobacter pylori (H. pylori) may be difficult due to antibiotic resistance. Indeed, after one failure, a second-line therapy is needed and a bismuth containing quadruple therapy (BQT) with a three-in-one capsule formulation is becoming very popular. Therefore, we aimed to evaluate effectiveness and safety of BQT as a second-line therapy. We recruited consecutive patients with one therapy failure. For ten days patients received the three-in-one BQT Pylera® therapy, in combination with a proton-pump inhibitor (PPI), decided at the choice of the investigator, at full dose bid. The eradication rate was calculated by intention-to-treat (ITT) and per-protocol (PP)analyses and 95% confidence intervals (CI) were calculated. Seventy-three patients were recruited, 41 females and 32 males (mean age 53.0±13.1 years). Fifty-five patients failed triple therapy with amoxicillin and clarithromycin and the remaining 18 received sequential therapy. Seventy-two patients consumed at least 90% of the capsules, while only one did not complete the therapy due to adverse events (nausea and diarrhea). By ITT analysis, BQT was successful in 62 subjects (eradication rate 84.9%, 95%CI 76.7–93.1%). By PP analysis, the eradication rate was 86.1% (95%CI 78.1–94.1%).Adverse events were observed in 14 subjects (20.5%).In conclusion, our report confirmed that BQT is effective as an empiric second-line regimen.

scholarly journals Evaluation of DNA extraction methods for quantifying Helicobacter pylori in water with PCR inhibitors

2021 ◽  
Author(s):  
Eun-Sook Lee ◽  
So-Yang Cha ◽  
Jong-Soon Jung
Keyword(s):  
Helicobacter Pylori ◽  
Real Time ◽  
Dna Extraction ◽  
Water Samples ◽  
Real Time Pcr ◽  
Heavy Rain ◽  
Extraction Methods ◽  
Pcr Inhibitors ◽  
H Pylori ◽  
Dna Extraction Methods

Abstract DNA extraction methods were evaluated to reduce PCR inhibitors and quantify Helicobacter pylori directly from water samples using real-time PCR. Three nucleic acid extraction methods were evaluated for different types of water samples. While the QIAamp DNA mini kit for tissue was suitable for DNA extraction from treated water, the QIAamp DNA stool mini kit was still efficient in analyzing samples from river water after heavy rain and with high concentration of PCR inhibitors. The FastDNA SPIN Kit for Soil could extract DNA effectively from microbes in river and stream waters without heavy rain. Immunomagnetic separation (IMS) was used prior to DNA extraction and was a useful tool for reducing PCR inhibitors in influent and stream samples. H. pylori in various waters could be quantified directly by real-time PCR while minimizing the effect of PCR inhibitors by an appropriate method through the evaluation of DNA extraction methods considering the characteristics of the matrix water. The findings of the present study suggest that the types or characteristics of water sample by source and precipitation are an important factor in detecting H. pylori and they can be applied when detecting and monitoring of other pathogens in water.

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