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This study aims to evaluate the biofilm formation of clinical isolates of H. pylori and its impact on antibiotic eradication. Methods: The thickness, morphology, and structure of biofilms derived from nine H. pylori strains were examined using confocal laser scanning microscopy, scanning electron microscopy, and transmission electron microscopy. Subsequently, the susceptibility of both planktonic and biofilm bacteria was assessed through the determination of minimum inhibitory concentration and minimum biofilm eradication concentration for amoxicillin, clarithromycin, levofloxacin, and tetracycline. Results: The results revealed varying biofilm thicknesses and densities among the strains, characterised by the presence of numerous filaments intertwining and connecting bacterial cells. Additionally, several cases exhibited susceptibility based on MIC measurements but resistance according to MBEC measurements, with MBEC indicating a higher resistance rate. Pearson Correlation analysis demonstrated a positive correlation between biofilm thickness and MBEC results (0 < r < 1), notably significant for amoxicillin (r = 0.801, P = 0.009) and tetracycline (r = 0.696, P = 0.037). Conclusion: Different strains of H. pylori exhibit variations in their capacity to release outer membrane vesicles (OMVs) and form biofilms. Biofilm formation can influence the effectiveness of amoxicillin and tetracycline in eradicating susceptible bacterial strains. [ABSTRACT FROM AUTHOR]

Background: Helicobacter pylori infections are generally acquired during childhood and affect half of the global population, but its transmission route remains unclear. It is reported that H. pylori can be internalized into Candida, but more evidence is needed for the internalization of H. pylori in human gastrointestinal Candida and vaginal Candida. Methods: Candida was isolated from vaginal discharge and gastric mucosa biopsies. We PCR-amplified and sequenced H. pylori-specific genes from Candida genomic DNA. Using optical and immunofluorescence microscopy, we identified and observed bacteria-like bodies (BLBs) in Candida isolates and subcultures. Intracellular H. pylori antigen were detected by immunofluorescence using Fluorescein isothiocyanate (FITC)-labeled anti-H. pylori IgG antibodies. Urease activity in H. pylori internalized by Candida was detected by inoculating with urea-based Sabouraud dextrose agar, which changed the agar color from yellow to pink, indicating urease activity. Results: A total of 59 vaginal Candida and two gastric Candida strains were isolated from vaginal discharge and gastric mucosa. Twenty-three isolates were positive for H. pylori 16S rDNA, 12 were positive for cagA and 21 were positive for ureA. The BLBs could be observed in Candida cells, which were positive for H. pylori 16S rDNA, and were viable determined by the LIVE/DEAD BacLight Bacterial Viability kit. Fluorescein isothiocyanate (FITC)-conjugated antibodies could be reacted specifically with H. pylori antigen inside Candida cells by immunofluorescence. Finally, H. pylori-positive Candida remained positive for H. pylori 16S rDNA even after ten subcultures. Urease activity of H. pylori internalized by Candida was positive. Conclusion: In the form of BLBs, H. pylori can internalize into gastric Candida and even vaginal Candida, which might have great significance in its transmission and pathogenicity. [ABSTRACT FROM AUTHOR]

Helicobacter pylori are transmissible from person to person and among family members. Mother‐to‐child transmission is the main intrafamilial route of H. pylori transmission. However, how it transmits from mother to child is still being determined. Vaginal yeast often transmits to neonates during delivery. Therefore, H. pylori hosted in yeast might follow the same transmission route. This study aimed to detect intracellular H. pylori in vaginal and fecal yeasts isolates and explore the role of yeast in H. pylori transmission. Yeast was isolated from the mothers' vaginal discharge and neonates' feces and identified by internal transcribed spacer (ITS) sequencing. H. pylori 16S rRNA and antigen were detected in yeast isolates by polymerase chain reaction and direct immunofluorescence assay. Genetic relationships of Candida strains isolated from seven mothers and their corresponding neonates were determined by random amplified polymorphic DNA (RAPD) fingerprinting and ITS alignment. The Candida isolates from four mother–neonate pairs had identical RAPD patterns and highly homologous ITS sequences. The current study showed H. pylori could be sheltered within yeast colonizing the vagina, and fecal yeast from neonates is genetically related to the vaginal yeast from their mothers. Thus, vaginal yeast presents a potential reservoir of H. pylori and plays a vital role in the transmission from mother to neonate. Take‐away: Helicobacter pylori is sheltered in both vaginal and fecal yeasts.Vaginal yeast is a potential reservoir for H. pylori.Vaginal yeast may facilitate H. pylori transmission from mother to neonate. [ABSTRACT FROM AUTHOR]

Purpose: Chronic Helicobacter pylori infection causes peptic ulcers in a subpopulation of individuals and is a risk factor for the development of gastric cancer. Multiple infections and heteroresistant H. pylori contribute to poor treatment efficacy. Here, we investigated the extent of genetic diversity among H. pylori strains within a given host and its influence on the results of antibiotic (metronidazole, levofloxacin, clarithromycin, amoxicillin, and tetracycline) susceptibility testing. Materials and Methods: Gastric mucosa biopsy samples were obtained from patients with gastric disorders, including 48 H. pylori positive patients, who were never previously treated for H. pylori infection. Five potential H. pylori colonies isolated from each sample were subcultured for enrichment. Enriched H. pylori colonies were identified through Gram staining and assays for urease, oxidase, and catalase. For each H. pylori monoclonal colony, the antibiotic susceptibility was assessed, genomic DNA was sequenced, and the cytotoxin-associated gene A (cagA) genotype was verified. Co-infection with multiple H. pylori strains was determined using random amplified polymorphic DNA (RAPD)-polymerase chain reaction (PCR). Results: Thirteen gastric mucosa biopsy samples were positive for H. pylori. Five monoclonal strains isolated from each of these 13 patients were identified as H. pylori. RAPD-PCR indicated that intra-patient monoclonal strains of H. pylori in 10 of the 13 samples exhibited heterogeneity. Among the 13 patients, intra-patient monoclonal strains isolated from 4 patients had identical cagA genotype, whereas intra-patient monoclonal strains isolated from the other 9 patients harbored more than one cagA genotype. The antibiotic susceptibility of five intra-patient monoclonal strains from seven patients was inconsistent. Conclusion: The existence of heterogeneous H. pylori strains with resistance to different drugs and virulence were common within the gastric mucosa of an individual patient. [ABSTRACT FROM AUTHOR]

Background: The multidrug resistance of Helicobacter pylori is becoming an increasingly serious issue. It is therefore necessary to study the mechanism of multidrug resistance of H pylori. We have previously identified that the HP0939, HP0497, and HP0471 transporters affect the efflux of drugs from H pylori. As efflux pumps participate in bacterial multidrug resistance and biofilm formation, we hypothesized that these transporters could be involved in the multidrug resistance and biofilm formation of H pylori. Materials and Methods: We therefore constructed three knockout strains, Δhp0939, Δhp0497, and Δhp0471, and three high‐expression strains, Hp0939he, Hp0497he, and Hp0471he, using the wild‐type (WT) 26 695 strain of H pylori as the template. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of wild strains, knockout strains, and high‐expression strains to amoxicillin, metronidazole, and other antibiotics were measured. The efflux capacity of high‐expression strains and wild strains was compared by Hoechst 33 342 accumulation assay. Results: Determination of the MIC and MBC of the antibiotics revealed that the knockout strains were more sensitive to antibiotics, while the high‐expression strains were less sensitive to antibiotics, compared to the WT. The ability of the high‐expression strains to efflux drugs was significantly higher than that of the WT. We also induced H pylori to form biofilms, and observed that the knockout strains could barely form biofilms and were more sensitive to several antibiotics, compared to the WT. The mRNA expression of hp0939, hp0497, and hp0471 in the clinically sensitive and multidrug‐resistant strains was determined, and it was found that these genes were highly expressed in the multidrug‐resistant strains that were isolated from the clinics. Conclusions: In this study, we found three transporters involved in intrinsic multidrug resistance of H pylori. [ABSTRACT FROM AUTHOR]

Biofilm formation is one of the most important factors causing drug resistance of Helicobacter pylori. Therefore, it is necessary to explore the mechanism underlying the biofilm formation and its eradication methods. The outer membrane proteins (OMPs) play important roles in the formation of bacterial biofilms and are considered the essential targets for new drug discovery. Natural products play significant roles in anti-bacterial and anti-biofilm functions. This study explored the key OMPs involved in the biofilm formation of H. pylori and the natural products that target these OMPs. Transcriptome sequencing, gene knockout, and electrophoretic mobility shift assay (EMSA) were performed to reveal that OMP6 was involved in the biofilm formation of H. pylori , which was regulated by non-phosphorylated ArsR. Molecular docking suggested that aloe-emodin (AE) could target OMP6 and destroy the biofilms of H. pylori. Further exploration of its mechanism found that AE could also inhibit the expression of omp6 mRNA by binding to its regulator ArsR. In summary, we have discovered a novel molecular mechanism regulating the biofilm formation of H. pylori and identified a natural product against H. pylori biofilms, providing potential clues for clinical treatment of H. pylori. [ABSTRACT FROM AUTHOR]

The present study aimed to investigate the virulence diversity of Helicobacter pylori (H. pylori) in major ethnic groups residing in Guizhou province, China, and its association with clinical outcomes. Gastric mucosal biopsies were collected from the pylorus of patients with gastrointestinal disorders. H. pylori was identified by colonial morphology, Gram staining, a urease test and H. pylori-specific 16S rRNA gene fragment PCR amplification. DNA was extracted from pure culture and used for virulence gene analysis. The cytotoxin associated gene A (cagA), vacuolating cytotoxin A (vacA) and induced by contact with epithelium gene A (iceA) genes were analyzed by polymerase chain reaction analysis. The cagA gene was further analyzed through sequencing of the C-terminal region containing EPIYA motifs, and phylogenetic analysis of the cagA C-terminal variable region was performed using MEGA 6.0 software. In the present study, 73 H. pylori strains were isolated from clinical samples. cagA genotypes were detected in all strains, namely cagA-AB, -ABC, -ABD and -BD genotypes were found in five (6.85%), three (4.11%), 63 (86.30%) and two (2.74%) isolates, respectively. Phylogenetic analysis showed that there was a clustering association between the cagA-AB and cagA-ABC genotypes, and between the cagA-ABD and cagA-BD genotypes. In terms of the frequency of the four EPIYA or EPIYA-like motifs, the most predominant was EPIYA (92.92%), followed by EPIYT (3.77%), ESIYA (2.83%) and ESIYT (0.47%). The predominant vacA genotype was s1c/m2 (65.75%), and the predominant iceA genotype was iceA1 (79.45%). There were no associations between the H. pylori cagA, vacA or iceA genotypes and clinical outcomes. No significant difference was found in the distribution of these genotypes according to the age, ethnicity or location of residence of patients. In conclusion, H. pylori isolated from patients in Guizhou region, China, showed a unique genotype, which was mainly East Asia-type cagA (ABD), vacA s1c/m2 genotype or iceA1-postiive. These results provide important information on the distribution of H. pylori virulence genotypes in Guizhou province, China. [ABSTRACT FROM AUTHOR]