Phenotypic and genotypic analysis of clarithromycin-resistant Helicobacter pylori from Bogotá D.C., Colombia

Resistance of Helicobacter pylori to clarithromycin is the most common cause of treatment failure in patients with H. pylori infections. This study describes the MICs and the presence of 23S rRNA mutations of H. pylori isolates from Bogotá, D.C., Colombia. H. pylori were isolated from gastric biopsies from patients with functional dyspepsia. Clarithromycin susceptibility was investigated by agar dilution and strains were considered resistant if the MIC was ≥1 μg/ml. DNA sequences of the 23S rRNA gene of strains resistant and sensitive to clarithromycin were determined to identify specific point mutations. Clarithromycin resistance was present in 13.6% of patients by agar dilution. The A2143G, A2142G and A2142C mutations were found in 90.5, 7.1, and 2.4% of H. pylori strains with resistance genotype.The resistant phenotype was associated with 23S rRNA resistance genotype in 85.7% of isolates. The point mutations in 23S rRNA were well correlated with MICs values for clarithromycin.     

Real-time PCR improves Helicobacter pylori detection in patients with peptic ulcer bleeding

Background and Aims

Histological and rapid urease tests to detect H. pylori in biopsy specimens obtained during peptic ulcer bleeding episodes (PUB) often produce false-negative results. We aimed to examine whether immunohistochemistry and real-time PCR can improve the sensitivity of these biopsies.

Patients and Methods

We selected 52 histology-negative formalin-fixed paraffin-embedded biopsy specimens obtained during PUB episodes. Additional tests showed 10 were true negatives and 42 were false negatives. We also selected 17 histology-positive biopsy specimens obtained during PUB to use as controls. We performed immunohistochemistry staining and real-time PCR for 16S rRNA, ureA, and 23S rRNA for H. pylori genes on all specimens.

Results

All controls were positive for H. pylori on all PCR assays and immunohistochemical staining. Regarding the 52 initially negative biopsies, all PCR tests were significantly more sensitive than immunohistochemical staining (p<0.01). Sensitivity and specificity were 55% and 80% for 16S rRNA PCR, 43% and 90% for ureA PCR, 41% and 80% for 23S rRNA PCR, and 7% and 100% for immunohistochemical staining, respectively. Combined analysis of PCR assays for two genes were significantly more sensitive than ureA or 23S rRNA PCR tests alone (p<0.05) and marginally better than 16S rRNA PCR alone. The best combination was 16S rRNA+ureA, with a sensitivity of 64% and a specificity of 80%.

Conclusions

Real-time PCR improves the detection of H. pylori infection in histology-negative formalin-fixed paraffin-embedded biopsy samples obtained during PUB episodes. The low reported prevalence of H. pylori in PUB may be due to the failure of conventional tests to detect infection.     

Co-detection of Helicobacter pylori and of its resistance to clarithromycin by PCR

Our aim was to develop a rapid molecular test based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and making it possible to detect Helicobacter pylori directly from gastric biopsy samples, and to test its susceptibility to clarithromycin. A 629-bp fragment of the 23S rRNA gene of H. pylori was amplified by PCR and the mutations responsible for clarithromycin resistance were detected with BsaI and BbsI restriction endonucleases. Thirty-five gastric samples were tested in parallel by standard microbiologic methods (culture and clarithromycin susceptibility testing with E-test strips) and by PCR-RFLP. The 10 culture-negative samples were also PCR-negative. Sixteen out of the 25 culture-positive samples (64%) were PCR-positive. RFLP analysis could be done in 12 cases and the results were in agreement with those of the E-test: susceptibility in five cases, resistance in seven (six A2144G mutations and one A2143G mutation).     

Primary antibiotic resistance and associated mechanisms in Helicobacter pylori isolates from Senegalese patients

Background

Antibiotic combination therapy for Helicobacter pylori eradication must be adapted to local resistance patterns, but the epidemiology of H. pylori resistance to antibiotics is poorly documented in Africa. The aim was to determine the antibiotic resistance rates, as well as the associated molecular mechanisms, of strains isolated in Dakar, Senegal.

Methods

One hundred and eight H. pylori strains were isolated between 2007 and 2009 from 108 patients presenting with upper abdominal pain to the Gastroenterology Department of Le Dantec Hospital. Antimicrobial susceptibility testing was performed for amoxicillin, clarithromycin, metronidazole, levofloxacin and tetracyclin using the E-test method. Mutations in the 23S rRNA gene of clarithromycin-resistant strains and in gyrA and gyrB of levofloxacin-resistant strains were investigated.

Results

Isolates were characterized by no resistance to amoxicillin (0%), tetracycline (0%), and very low rate of resistance to clarithromycin (1%), but a high rate of resistance to metronidazole (85%). The clarithromycin-resistant strain displayed the A2143G mutation. A worrying rate of levofloxacin resistance was detected (15%). N87I and D91N were the most common mutations in the quinolone-resistance-determining region of gyrA.

Conclusions

The first-line empirical regimen for H. pylori eradication in Senegal should include clarithromycin. Increasing rates of fluoroquinolone resistance detected should discourage the use of levofloxacin-containing regimens without prior antimicrobial susceptibility testing.     

Detection of clarithromycin resistance and 23SrRNA point mutations in clinical isolates of Helicobacter pylori isolates: Phenotypic and molecular methods

Background and objectivesPeptic ulcer disease, chronic gastritis, and stomach cancer are all caused by H. pylori. The most notable drug for the treatment is the antibiotic clarithromycin, which is currently the drug of choice. H. pylori clarithromycin resistance has been associated with point mutations in 23srRNA, the most prominent of which are A2143 and A2144G. In H. pylori bacteria, methylase synthesis, macrolide-inactivating enzyme activity, and active efflux have all been found to be resistance mechanisms. The goal of the study is to determine how resistant H. pylori is to clarithromycin and what the minimum inhibitory concentration is for various antimicrobials. Furthermore, gastro-endoscopy will be performed on Iraqi patients to detect the presence of A2143G and A2144G point mutations in Helicobacter pylori infections, as diagnosed from the pyloric region and other anatomical regions.MethodsOne hundred fifteen samples were collected from patients strongly suspected of H. pylori infection presented for upper gastrointestinal endoscopy at Ramadi Teaching Hospitals and Private Clinics for the period from January 2020 until February 2021. Specimens were cultured on brain heart infusion agar containing various antibiotics and were incubated at 37 °C under microaerophilic conditions. For identification of H. pylori, isolates of the biochemical tests and RT-PCR assay were applied. The Epsilometer test was used in the antibiotic susceptibility testing as dependent on the CLSI standard. The Restriction Fragment Length Polymorphism technique was used to determine point mutations.ResultsIn total, 55 (47.8%) Helicobacter pylori isolates were cultured from the 115 biopsy specimens, among which 16 (29.1%), 38 (69.1%), 20 (36.4%), and 40 (72.7%) revealed some degree of resistance to levofloxacin, clarithromycin, ciprofloxacin, and metronidazole, respectively. The frequency of A2144G and A2143 point mutations were 23 (60.5%) and 19 (50%), respectively.ConclusionsAccording to our results, Helicobacter pylori showed high resistance to clarithromycin. Our results demonstrate the requirement for antibiotic susceptibility testing and molecular methods in selecting drug regimens.     

Development of a Highly Sensitive Method for Detection of Clarithromycin-Resistant Helicobacter pylori from Human Feces

Gastric infection of clarithromycin (CAM)-resistant Helicobacter pylori is one of the major causes of failure to eradicate this organism. A noninvasive and useful method for the detection of CAM-resistant H. pylori from human feces by restriction fragment length polymorphism (RFLP)-nested polymerase chain reaction (PCR) targeting the mutation of the 23S rRNA gene that confers CAM-resistance in H. pylori was developed in this study. Our nested PCR method detected DNA of H. pylori in feces with high sensitivity and specificity compared with both an enzyme-linked immunoadsorbent assay (ELISA) of H. pylori in feces and the isolation of H. pylori from gastric biopsy. Furthermore, the results of mutation analysis of the H. pylori 23S rRNA gene amplified from feces completely correlated with both that of the H. pylori 23S rRNA gene amplified from the isolates of gastric biopsy and the susceptibility of H. pylori isolates to CAM. Therefore, our results show that this RFLP/nested PCR method is useful for the accurate diagnosis of CAM-resistant H. pylori infection from feces.     

<Emphasis Type="Italic">A2144G</Emphasis> Is the Main Mutation in the <Emphasis Type="Italic">23S</Emphasis> rRNA Gene of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> Associated with Clarithromycin Resistance

To detect point mutations A2115C, A2143G/C, and A2144G in the 23S rRNA gene of Helicobacter pylori associated with resistance of the microorganism to clarithromycin, a new powerful way of analysis was used. This method involved the reaction of minisequencing followed by MALDI-TOF mass spectrometry of reaction products. In ten analyzed clarithromycin-resistant clinical isolates of H. pylori obtained in Russia, the resistance was found to be mediated only by mutation A2144G in the 23S rRNA gene.     

Identification of a novel mutation affecting domain V of the 23S rRNA gene in Helicobacter pylori

BACKGROUND: This study analyzes clarithromycin resistance status and 23S rRNA gene mutations in Helicobacter pylori strains from Central Italian patients. MATERIALS AND METHODS: H. pylori strains from 235 dyspeptic patients (205 with no history of clarithromycin exposure and 30 referred for failure of eradication therapy) were tested for clarithromycin resistance by screening agar method and E-test. Resistant strains were analyzed for mutations of the 23S rRNA gene by PCR-RFLP and sequencing. RESULTS: Primary resistance was observed in strains from 43/205 (21%) patients with no history of clarithromycin exposure and secondary resistance in 30/30 (100%) strains from previously treated patients. A single mutant strain was detected in 54/73 (74%) cases, a mixture of one or more mutant(s) plus the wild type in the remaining 19/73 (26%) cases. One 23S rRNA gene mutation (A-->T transversion at nucleotide 2144) in the peptidyltransferase region of domain V was novel. CONCLUSIONS: This study shows: (a) a high prevalence of H. pylori strains with primary or secondary clarithromycin resistance in an urban area of Central Italy; (b) colonization by both mutant and wild-type H. pylori in the same patient; (c) a novel variant of the H. pylori 23S rRNA gene.     

Primary antibiotic resistance of Helicobacter pylori isolated from Beijing children

Background: The antimicrobials resistance of Helicobacter pylori (H. pylori) was able to sharply decline the eradication rate of H. pylori both in adults and children, but there are limited studies about the primary antibiotic resistance and the related gene mutations, specifically in China. Materials and Methods: The primary resistance to 9 antibiotics of 73 H. pylori strains isolated from gastric biopsies of children recruited at Beijing Children’s Hospital was assessed, and the mutations in 23S rRNA gene of 65 macrolide‐resistant strains and in gyrA and gyrB of 12 quinolone‐resistant strains were investigated. Results: The resistance rate to clarithromycin, azithromycin, metronidazole, levofloxacin, moxifloxacin, and rifampicin was 84.9%, 87.7%, 61.6%, 13.7%, 15.1%, and 6.8%, respectively. No resistance to amoxicillin, gentamicin, and tetracycline was observed. Dual, triple, and quadruple antibacterial resistant percentage was 46.6% (34/73), 15.1% (11/73), and 2.7% (2/73), respectively. The gene mutation rate of A2142C, A2142G, and A2143G in 23S rRNA gene was 1.5% (1/65), 6.2% (4/65), and 84.6% (55/65), respectively. The detection rate of mutations of Asn87, Asp91, and Met191 in GyrA was 41.7% (5/12), 25% (3/12), and 25% (3/12), respectively. Conclusion: The high prevalence of primary antibiotic resistance was out of expectation in H. pylori strains isolated from the children in Beijing. Antibiotic susceptibility should be made clear before the antibiotic was used in the anti‐H. pylori therapy in this population. The A2143G was the most populated mutation in macrolide‐resistant strains, and Asn87 and Asp91 of GyrA were the most common mutation points in quinolone resistance strains.     

Functional and Molecular Surveillance of Helicobacter pylori Antibiotic Resistance in Kuala Lumpur

Background

Helicobacter pylori is the etiological agent for diseases ranging from chronic gastritis and peptic ulcer disease to gastric adenocarcinoma and primary gastric B-cell lymphoma. Emergence of resistance to antibiotics possesses a challenge to the effort to eradicate H. pylori using conventional antibiotic-based therapies. The molecular mechanisms that contribute to the resistance of these strains have yet to be identified and are important for understanding the evolutional pattern and selective pressure imposed by the environment.

Methods and Findings

H. pylori was isolated from 102 patients diagnosed with gastrointestinal diseases, who underwent endoscopy at University Malaya Medical Centre (UMMC). The isolates were tested for their susceptibility on eleven antibiotics using Etest. Based on susceptibility test, 32.3% of the isolates were found to have primary metronidazole resistance; followed by clarithromycin (6.8%) and fluoroquinolones (6.8%). To further investigate the resistant strains, mutational patterns of gene rdxA, frxA, gyrA, gyrB, and 23S rRNA were studied. Consistent with the previous reports, metronidazole resistance was prevalent in the local population. However, clarithromycin, fluoroquinolone and multi-drug resistance were shown to be emerging. Molecular patterns correlated well with phenotypic data. Interestingly, multi-drug resistant (MDR) strains were found to be associated with higher minimum inhibitory concentration (MIC) than their single-drug resistant (SDR) counterparts. Most importantly, clarithromycin-resistant strains were suggested to have a higher incidence for developing multi-drug resistance.

Conclusion

Data from this study highlighted the urgency to monitor closely the prevalence of antibiotic resistance in the Malaysian population; especially that of clarithromycin and multi-drug resistance. Further study is needed to understand the molecular association between clarithromycin resistance and multi-drug resistance in H. pylori. The report serves a reminder that a strict antibiotic usage policy is needed in Malaysia and other developing countries (especially those where H. pylori prevalence remained high).     

Molecular characterization of ticks and tick-borne piroplasms from cattle and camel in Hofuf,eastern Saudi Arabia

The aims of the present study were to characterize ticks infesting the dromedary camel and cattle in Hofuf, Eastern Saudi Arabia and to determine the piroplasms that they may harbor. DNA was extracted from ticks, collected from camels and cattle, using commercial kits and subjected to polymerase chain reaction using specific primers for the amplification of ticks and piroplasms DNA. The cytochrome oxidase subunit I mitochondrial gene (COI) was used for characterization of ticks whereas partial 18S rRNA gene (18S rRNA) was used for piroplasms characterization. Ticks were genetically identified as Hyalomma dromedarii and Hyalomma anatolicum. Both cattle and camel in Hofuf, were found to be infested with both species. Both ticks identified as H. dromedarii and H. anatolicum from camels and cows showed 100% identity to COI sequences from the same species available in GenBank. Only Theileria annulata DNA was amplified from both H. anatolicum and H. dromedarii infesting cattle. None of the ticks collected from camels revealed DNA of piroplasms. T. annulata DNA was reported for the first time from Hofuf and the role of both H. anatolicum and H. dromedarii as potential vectors for this parasite in cattle in Saudi Arabia has been documented for the first time.     

Detection of Helicobacter pylori in stool samples of young children using real‐time polymerase chain reaction

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.     

Novel genotypes in Helicobacter pylori involving domain V of the 23S rRNA gene

BACKGROUND: Helicobacter pylori is a pathogenic bacterium that infects a half of the human population. In Chile, between 55% and 79% of people are colonized by H. pylori. At present, therapeutic strategies to eradicate the bacterium depend on the knowledge of its resistance to antibiotics. The clarithromycin resistance in H. pylori is associated with point mutations in the 23S rRNA. This study analyzes 23S rRNA gene mutations and minimum inhibitory concentration (MIC) for clarithromycin in H. pylori isolates from patients of the metropolitan region of Chile. MATERIALS AND METHODS: H. pylori isolates from 50 dyspeptic patients with no history of clarithromycin exposure were tested for clarithromycin resistance by agar dilution method. Resistant strains were analyzed for mutations in the 23S rRNA gene by polymerase chain reaction-based restriction fragment length polymorphism and sequencing. RESULTS: Primary resistance was observed in 10 isolates (20%). A single mutation was detected in four of the 10 isolates and two or more mutations in the other six cases. The C2147G transversion and G1939A, T1942C, and A2142G transitions in the peptidyltransferase region of domain V were novel. CONCLUSIONS: The study shows: 1, novel variants of the H. pylori 23S rRNA gene; and 2, a high prevalence of H. pylori displaying primary clarithromycin resistance with low level of MIC in an urban area of the Metropolitan Region of Chile.     

Rapid combined characterization of microorganism and host genotypes using a single technology

Background. Genetic information is becoming increasingly important in diagnosis and prognosis of infectious diseases. In this study we investigated the possibility of using a single technology, the Pyrosequencing? technology (Biotage AB, Uppsala, Sweden), to gather several kinds of important genetic information from the human pathogen Helicobacter pylori, as well as from the carrier of the H. pylori infection. Materials and Methods. DNA from 87 clinical isolates of H. pylori, 50 isolates from H. pylori‐infected transgenic mice and nine gastric biopsies from H. pylori‐infected patients was analyzed for targets in the 16S rRNA, 23S rRNA and cytotoxin associated gene A (cagA) genes to determine species identity, clarithromycin susceptibility and virulence level, respectively. In addition, three single nucleotide polymorphisms in the human interleukin‐1B (IL‐1B) gene, reported to affect the risk of developing gastric cancer, were analyzed in the gastric biopsy samples. Results. All DNA targets were processed and analyzed in parallel, enabling convenient genetic characterization of both pathogen and host. All genotypes were easily and accurately assigned. In the 16S rRNA analysis, 99.83% of the bases were correctly called. Conclusions. We conclude that genetic analysis using Pyrosequencing? technology was nonlaborious, and gave highly accurate data for different kinds of target. We therefore believe that this technology has the potential to complement or in the future substitute the time‐consuming traditional microbial identification and typing methods, as well as enabling rapid typing of relevant host genetic markers.     

Molecular diagnosis of Helicobacter pylori antibiotic resistance in the Taif region,Saudi Arabia

Success in eradication of Helicobacter pylori is declining globally because H. pylori has developed resistance against most of the antibiotics proposed for eradication regimens, mainly through point mutations. The present study included 200 patients with dyspepsia attending Taif Hospital. Gastric biopsies were obtained during gastroscopy and subjected to rapid urease testing. Molecular methods were used to confirm diagnoses of H. pylori infection and to identify resistance gene variants of four antibiotics; namely, clarithromycin, metronidazole, fluoroquinolones and tetracycline (23S rRNA, gyrA, rdxA and 16S rRNA respectively). Of all investigated patients, Molecular diagnoses were made in 143 of all investigated patients; thus, the prevalence was .5%. The overall rate of resistance to clarithromycin among the H. pylori‐positive patients was high (39.9%) and the rate of resistance significantly greater (48.2%) among the secondary resistance group, secondary resistance being defined as resistance as a result of previous exposure to the relevant antibiotic. The rate of resistance to fluoroquinolones was considered moderate; the difference in rate of resistance between the primary and secondary resistance groups (8.4% and 9.5%, respectively) was not significant Also, there was a low prevalence of both primary and the secondary tetracycline resistance in the study cohort. In contrast, the prevalence of metronidazole resistance was considered high with no significant difference between the two resistance groups. H. pylori showed an increased prevalence of resistance to all four of the commonly used therapeutic agents. Thus, eradication therapy should be based on the regional results of susceptibility testing. Moreover, treatment tailored according to individually determined H. pylori susceptibility may be a reasonable future goal.     

Application of real-time PCR stool assay for Helicobacter pylori detection and clarithromycin susceptibility testing in Brazilian children

Background: Helicobacter pylori ClariRes assay is a novel commercially available real‐time PCR assay allowing H. pylori detection and clarithromycin susceptibility testing in either gastric biopsy or stool specimens. Objective: The aim of this study was to validate the novel biprobe real‐time assay in stool specimens from 217 dyspeptic children. Methods: DNA from gastric biopsies and stool specimens were obtained and submitted to the biprobe real time assay for H. pylori detection and clarithromycin susceptibility testing. Results: The sensitivity, specificity, and test accuracy were 69, 100 and 93.9% for the detection of H. pylori infection and 83.3, 100 and 95.6%, for detection of clarithromycin resistance. Conclusion: This assay proved to be appropriate for H. pylori clarithromycin susceptibility testing, particularly in children populations where a high prevalence of clarithromycin‐resistant strains is suspected.     

Helicobacter pylori antimicrobial resistance in a pediatric population

Background

The increasing prevalence of Helicobacter pylori (H. pylori) antimicrobial resistance, primarily for clarithromycin decreases the success of treatment. The aim of this study is to determine the local pattern of first‐line antimicrobials resistance and the eradication rate.

Material and Methods

Prospective cohort study of H. pylori infected patients (positive histological or cultural exams) treated at Centro Materno‐Infantil do Norte from January of 2013 to October of 2017. Susceptibility to 4 antibiotics: amoxicilin, metronidazole, clarithromycin, and levofloxacin were analyzed by E‐test (phenotypic resistance). The E‐test was chosen because it is simple and cost‐effective for routine susceptibility testing. Point mutations that confer clarithromycin resistance were surveyed (genotypic resistance). Eradication of H. pylori infection was defined by a negative urea breath test or fecal antigen 6‐8 weeks after the end of treatment.

Results

Of a total of 74 H. pylori infected patients, 16 were excluded because they had previous H. pylori treatment or severe systemic disease. Median age of infection cases was 15 years (3‐17 years). Eradication regimen used in all patients combined the use of 3 antibiotics (amoxicillin and metronidazole or clarithromycin) and proton pump inibhitor for 14 days and was tailored according antimicrobial susceptibility. 79.5% of the patients completed the treatment. The resistance rate for metronidazole and clarithromycin was 3.3% and 23.3%, respectively. There was no resistance for amoxicilin and levofloxacin. The rate of genotypic resistance to clarithromycin was 37.2%. The eradication rate was 97.8%.

Conclusions

The authors found a high resistance rate of H. pylori for clarithromycin in this northern portuguese pediatric center. This factor should determine a change in local current treatment, contraindicating the use of clarithromycin as a first‐line treatment for H. pylori infection in children. The high eradication rate maybe explained for the eradication treatment tailored according antimicrobial susceptibility.     

Differentiation of non-pylori Helicobacter species based on PCR–restriction fragment length polymorphism of the 23S rRNA gene

Phenotypic identification of non-pylori Helicobacter species has always been problematic and time-consuming in comparison with many other bacteria. We developed a rapid two-step identification assay based on PCR–restriction fragment length polymorphism (PCR–RFLP) analysis of the 23S rRNA gene for differentiating between non-pylori Helicobacter species. A new genus-specific primer pair based on all available complete and partial 23S rRNA sequences of Helicobacter species was designed. In silico restriction analysis of variable regions of the 23S rRNA gene suggested SmaI and HindIII endonucleases would provide a good level of differentiation. Analysis of the obtained 23S rRNA RFLP patterns divided all Helicobacter study strains into three species groups (groups A–C) and 12 unique restriction patterns. Wolinella succinogenes also gave a unique pattern. Our proposed PCR–RFLP method was found to be as a valuable tool for routine identification of non-pylori Helicobacter species from human or animal samples.     

Primary resistance of Helicobacter pylori to antimicrobial agents in Polish children

Helicobacter pylori resistance to antimicrobial agents is an important factor compromising the efficacy of treatment. Therefore the aims of our study were: to determine the prevalence of H. pylori resistance to clarithromycin, metronidazole, amoxycillin and tetracycline in children prior to eradication therapy, to compare different methods of susceptibility testing and to detect mutations responsible for clarithromycin resistance. During 1996-2000, 259 H. pylori strains were isolated from antral gastric biopsies. Susceptibility to antimicrobials was determined by the agar dilution method and the Etest. Mutations in the 23S rRNA gene associated with clarithromycin resistance were analysed by PCR-RFLP and direct sequencing. Overall, ninety-six strains (37%) were resistant to metronidazole, 50 strains (19.3%) were resistant to clarithromycin, and 20 strains (7.7%) were simultaneously resistant to both drugs. All cultured isolates were sensitive to amoxycillin and only one isolate (0.4%) was resistant to tetracycline. The agar dilution method and the Etest showed a perfect category correlation for clarithromycin and 4% discrepancies for metronidazole. Primary resistance to clarithromycin was mainly associated with an A2143G mutation in the 23S rRNA gene of H. pylori. The study highlights the high prevalence of H. pylori primary resistance to clarithromycin in Polish children, which implies a need for pretreatment susceptibility testing.     

Characteristics of phenotypic antibiotic resistance of Helicobacter pylori and its correlation with genotypic antibiotic resistance: A retrospective study in Ningxia

Background

Geographic differences exist in the antibiotic resistance patterns of Helicobacter pylori. Personalized treatment regimens based on local or individual resistance data are essential. We evaluated the current status of H. pylori resistance in Ningxia, analyzed resistance-related factors, and assessed the concordance of phenotypic and genotypic resistance.

Methods

Strains were isolated from the gastric mucosa of patients infected with H. pylori in Ningxia and relevant clinical information was collected. Phenotypic antibiotic susceptibility assays (Kirby–Bauer disk diffusion) and antibiotic resistance gene detection (Sanger sequencing) were performed.

Results

We isolated 1955 H. pylori strains. The resistance rates of H. pylori to amoxicillin, levofloxacin, clarithromycin, and metronidazole were 0.9%, 42.4%, 40.4%, and 94.2%, respectively. Only five tetracycline-resistant and one furazolidone-resistant strain were identified. Overall, 3.3% of the strains were sensitive to all six antibiotics. Multidrug-resistant strains accounted for 22.9%, of which less than 20% were from Wuzhong. Strains isolated from women and patients with nonulcerative disease had higher rates of resistance to levofloxacin and clarithromycin. Higher rates of resistance to metronidazole, levofloxacin, and clarithromycin were observed in the older age group than in the younger age group. The kappa coefficients of phenotypic resistance and genotypic resistance for levofloxacin and clarithromycin were 0.830 and 0.809, respectively, whereas the remaining antibiotics showed poor agreement.

Conclusion

H. pylori antibiotic resistance is severe in Ningxia. Therefore, furazolidone, amoxicillin, and tetracycline are better choices for the empirical therapy of H. pylori infection in this region. Host sex, age, and the presence of ulcerative diseases may affect antibiotic resistance of the bacteria. Personalized therapy based on genetic testing for levofloxacin and clarithromycin resistance may be a future direction for the eradication therapy of H. pylori infection in Ningxia.