Cytotoxin Gene Research Articles

Genomic insights into the antimicrobial resistance and virulence of Helicobacter pylori isolates from gastritis patients in Pereira, Colombia

BackgroundHelicobacter pylori infects the stomach and/or small intestines in more than half of the human population. Infection with H. pylori is the most common cause of chronic gastritis, which can lead to more severe gastroduodenal pathologies such as peptic ulcer, mucosa-associated lymphoid tissue lymphoma, and gastric cancer. H. pylori infection is particularly concerning in Colombia in South America, where > 80% of the population is estimated to be infected with H. pylori and the rate of stomach cancer is one of the highest in the continent.ResultsWe compared the antimicrobial susceptibility profiles and short-read genome sequences of five H. pylori isolates obtained from patients diagnosed with gastritis of varying severity (chronic gastritis, antral erosive gastritis, superficial gastritis) in Pereira, Colombia sampled in 2015. Antimicrobial susceptibility tests revealed the isolates to be resistant to at least one of the five antimicrobials tested: four isolates were resistant to metronidazole, two to clarithromycin, two to levofloxacin, and one to rifampin. All isolates were susceptible to tetracycline and amoxicillin. Comparative genome analyses revealed the presence of genes associated with efflux pump, restriction modification systems, phages and insertion sequences, and virulence genes including the cytotoxin genes cagA and vacA. The five genomes represent three novel sequence types. In the context of the Colombian and global populations, the five H. pylori isolates from Pereira were phylogenetically distant to each other but were closely related to other lineages circulating in the country.ConclusionsH. pylori from gastritis of different severity varied in their antimicrobial susceptibility profiles and genome content. This knowledge will be useful in implementing appropriate eradication treatment regimens for specific types of gastritis. Understanding the genetic and phenotypic heterogeneity in H. pylori across the geographical landscape is critical in informing health policies for effective disease prevention and management that is most effective at local and country-wide scales. This is especially important in Colombia and other South American countries that are poorly represented in global genomic surveillance studies of bacterial pathogens.

Analyses of the association between Helicobacter pylori antibody titre and pathogenicity before and after eradication: results of the Kyushu and Okinawa population study, a retrospective observational cohort study

ObjectivesTo assess the utility of Helicobacter pylori antibody testing, we evaluated the correlation between the H. pylori antibody titre and H. pylori-associated pathogenicity and the changes in antibody titre after...

Distribution and molecular analysis of Subtilase cytotoxin gene (subAB) variants in Shiga toxin-producing Escherichia coli (STEC) isolated from different sources in Iran.

Subtilase exhibits strong cytotoxicity that was first described in O113:H21 strain in Australia as a plasmid- encoded cytotoxin (subAB1). Subsequently, chromosomal variants including subAB2-1, subAB2-2, and subAB2-3 were described. We aimed to investigate the presence of subAB genes in a collection of Shiga toxin-producing Escherichia coli (STEC) strains (n=101) isolated from different sources in Iran. A collection of 101 archived STEC strains isolated from cattle (n=50), goats (n=25), sheep (n=15), wild captive animals (n=8: persian fallow deer, n=3; caspian pony, n=1; Macaca mulatta, n=4), and humans (n=3) during 2007-2016 were analyzed for the detection of different genes encoding the Subtilase variants, plasmidic and chromosomal virulence genes, phylogroups and serogroups. Overall, 57 isolates (56.4%) carried at least one variant of subAB. Most strains from small ruminants including 93% of sheep and 96% of caprine isolates carried at least one chromosomally encoded variant (subAB-2-1 and/or subAb2-2). In contrast, 12 cattle isolates (24%) only harbored the plasmid encoded variant (subAB1). STEC strains from other sources, including deer, pony and humans were positive for subAB-2-1 and/or subAb2-2. Our results reveal the presence of potentially pathogenic genotypes among locus of enterocyte effacement (LEE)-negative isolates, and some host specificity related to Subtilase variants and other virulence markers that may aid in source tracking of STEC during outbreak investigations.

Helicobacter pylori Cytotoxin-Associated Gene A (cagA) and Vacuolating Cytotoxin Gene A (vacA) Genotypes in Gastrointestinal Patients From Central Thailand.

Introduction The development of diseases associated with Helicobacter pylori(H. pylori) infection is closely linked to its virulence genes, which vary by geographic region. This study aimed to determine the prevalence of H. pylori cytotoxin-associated gene A (cagA) and vacuolating cytotoxin gene A (vacA) genes and their genotypes in patients with gastrointestinal diseases. Methods Patients diagnosed with gastrointestinal disease based on endoscopic findings were recruited for the study. Gastric biopsies were collected to screen for H. pylori infection using polymerase chain reaction (PCR). Subsequently, infected samples were tested forcagAand vacA genes, and their genotypes were analyzed by sequencing. Results Among 250 cases, 56% (140/250) exhibited gastrointestinal diseases. Of these cases, 32.1% (45/140) were infected with H. pylori. Regarding gene detection, 40 (88.9%)samples were positive for cagA, while all samples were positive for vacA. For cagA, the Western type with the ABC pattern was the most prominent. There was a statistically significant association between cagA genotypes and clinical outcomes, with the Western type being more prevalent in gastritis patients. For vacA, there was a high prevalence of the s1 and i1, while the m1 and m2 showed similar prevalence. In our combined analysis, the dominant vacA genotype combinations were s1m1i1 (46.7%). There were no statistical differences between the vacA genotypesand clinical outcomes (P > 0.05). Conclusion This study revealed a high prevalence of H. pylori cagA and vacA genes, but there were variations in their genotypes. A correlation was observed between the Western-type cagA and gastritis; however, no association was found between vacA genotypes and clinical outcomes.

Detection of some cytotoxin genes in local isolate of Oscillatoria (Cyanophyta)

Detection of some cytotoxin genes in local isolate of Oscillatoria (Cyanophyta)

Unveiling the Silent Intruder: H. pylori's Hidden Link to Ischemic Heart Disease.

Cardiovascular disease is the leading cause of death. In addition to the well-known risk factors associated with cardiovascular disease, such as age, diabetes mellitus, smoking, hypertension, and obesity, there has been a growing concern regarding cardiac complications stemming from the Gram-negative bacteria Helicobacter pylori. While H. pylori is most commonly associated with chronic gastritis, peptic ulcer disease, gastric adenocarcinoma, and gastric lymphoma, it has also been implicated in extra gastric manifestations, encompassing cardiac, neurologic, ocular, and dermatologic issues. Key virulent factors for coronary artery disease include the vacuolating cytotoxin gene A and the cytotoxin-associated gene A. The most likely pathogenic mechanism of the relationship between H. pylori and coronary artery disease is initiating a chronic inflammatory process associated with infection and the modifications of classic risk factors. These alterations lead to the creation of prothrombotic and procoagulant environments. Here, we review the cardiac manifestations of H. pylori and the underlying pathophysiological mechanisms.

Mechanistic research: Selenium regulates virulence factors, reducing adhesion ability and inflammatory damage of Helicobacter pylori.

The pathogenicity of Helicobacter pylori is dependent on factors including the environment and the host. Although selenium is closely related to pathogenicity as an environmental factor, the specific correlation between them remains unclear. To investigate how selenium acts on virulence factors and reduces their toxicity. H. pylori strains were induced by sodium selenite. The expression of cytotoxin-associated protein A (CagA) and vacuolating cytotoxin gene A (VacA) was determined by quantitative PCR and Western blotting. Transcriptomics was used to analyze CagA, CagM, CagE, Cag1, Cag3, and CagT. C57BL/6A mice were infected with the attenuated strains subjected to sodium selenite induction, and H. pylori colonization, inflammatory reactions, and the cell adhesion ability of H. pylori were assessed. CagA and VacA expression was upregulated at first and then downregulated in the H. pylori strains after sodium selenite treatment. Their expression was significantly and steadily downregulated after the 5th cycle (10 d). Transcriptome analysis revealed that sodium selenite altered the levels affect H. pylori virulence factors such as CagA, CagM, CagE, Cag1, Cag3, and CagT. Of these factors, CagM and CagE expression was continuously downregulated and further downregulated after 2 h of induction with sodium selenite. Moreover, CagT expression was upregulated before the 3rd cycle (6 d) and significantly downregulated after the 5th cycle. Cag1 and Cag3 expression was upregulated and downregulated, respectively, but no significant change was observed by the 5th cycle. C57BL/6A mice were infected with the attenuated strains subjected to sodium selenite induction. The extent of H. pylori colonization in the stomach increased; however, sodium selenite also induced a mild inflammatory reaction in the gastric mucosa of H. pylori-infected mice, and the cell adhesion ability of H. pylori was significantly weakened. These results demonstrate that H. pylori displayed virulence attenuation after the 10th d of sodium selenite treatment. Sodium selenite is a low toxicity compound with strong stability that can reduce the cell adhesion ability of H. pylori, thus mitigating the inflammatory damage to the gastric mucosa.

De novo whole transcriptome analysis of Aeromonas hydrophila isolated from the gut of an infected Labeo rohita.

Aeromonas hydrophila is a major generalist bacterial pathogen causing severe infections and mortalities in aquatic animals. Its genome, which was the first to be sequenced from the Aeromonas genus, may serve as a model for studying pathogenic mechanisms. To explore the pathogen-host fitness mechanism of bacterium, a comprehensive comparative transcriptome ecotype analysis of A. hydrophila isolated from the gut of Labeo rohita during infection was performed. Special characteristics in gene expression, gene ontology terms and expression of pathogenesis-associated genes, including genes encoding secreted proteins, candidate effectors, hydrolases, and proteins involved in secondary metabolite production were revealed. Among the database, 6,533 were gene ontology (GO) annotated, while 1,480 were not allocated in any GO terms. Investigation on GO illustrated that the articulated genes were improved with molecular function, cellular components, and biological processes. Further bioinformatics analysis identified the outer membrane protein genes (ompA, ompts, ompw, omp38, and omp48), cytotoxin, amylase, and lipase genes. Overall, this work allowed to designate, for the first time, a global view on the pathogenicity of Aeromonas hydrophila during infection. Furthermore, the study provides information on the fitness of A. hydrophila, a severe pathogen with a wide host range.

Association between the presence and genotype of Helicobacterpylori and periodontitis.

Whether Helicobacter pylori (H. pylori) infection is associated with periodontitis has been contested for decades. The relationship between H. pylori genotypes and periodontitis has not been clarified either. The present study provides a novel perspective to better understand the role of H. pylori in the pathogenesis of periodontitis. A total of 53 volunteers were recruited and divided into 3 groups in this cross-sectional study, namely the periodontally healthy group (15 participants), the stage I/II periodontitis group (20 participants) and the stage III/IV periodontitis group (18 participants). DNA from the subgingival plaque of all participants was extracted and PCR was performed using specific primers for the urease C gene and cytotoxin-associated gene A (cagA)/vacuolating cytotoxin gene A (vacA) to detect the presence and genotype of H. pylori. A χ2 test and one-way ANOVA were performed on the data. There was no significant difference in sex, age or body mass index between the groups. The detection rate of H. pylori was 39.62% in the total population and increased with the deepening of probing depth and clinical attachment loss. There were significant differences in the detection rate of H. pylori among the three groups, with 13.33, 40.00 and 61.11% in the periodontally healthy, stage I/II periodontitis and stage III/IV periodontitis groups, respectively (χ2=8.760, P<0.001). The cagA-/vacAs2m2 genotype was most commonly detected in the periodontally healthy group (100%). In the periodontitis group, cagA+/vacAs1m2 was the most commonly detected genotype in the stage I/II periodontitis group (37.5%) and cagA+/vacAs1m1 in the stage III/IV periodontitis group (36.3%). The results of the present study suggest that the detection rates and genotypes of H. pylori in the subgingival plaque are associated with the status of periodontitis. cagA+/vacAs1m1 and cagA+/vacAs1m2 may be considered virulence markers of periodontitis. However, given the small sample size and lack of correlation analysis of the study, further larger scale and high-quality clinical trials are required to confirm these findings.

Molecular identification, antimicrobial resistance and virulence gene profiling of Staphylococcus spp. associated with bovine sub-clinical mastitis in Bangladesh

This study was conducted to investigate the diversity and antimicrobial resistance profiling of Staphylococcus species causing sub-clinical mastitis (SCM) in dairy herds in Bangladesh as well as putative risk factors associated with the infections. Individual quarter milk samples were collected from a total of 284 lactating cows from 30 dairy farms were screened by means of California mastitis test; 178 (62.7%) of them had at least of quarter affected by SCM. After conventional microbiological isolation procedures, PCR tests were used for Staphylococcus species identification and detection of antimicrobial resistance and virulence genes. S. chromogenes (65.7%) was the most predominant species followed by, S. epidermidis (20.2%), S. haemolyticus (19.1%), S. aureus (15.7%), and S. sciuri (5.6%). High levels of antimicrobial resistance to ampicillin and amoxicillin/clavulanic acid were observed in S. aureus (82.1% and 75%) and S. sciuri (80% and 70%), while resistance to cefepime was markedly higher in S. chromogenes (95.7%), S. haemolyticus (94.1%), and S. epidermidis (97.2%). Multidrug resistance isolates were identified in all five species. The mecA gene was detected in S. aureus (32.1%) and S. chromogenes (5.98%). In addition, 20% S. sciuri and 17.7% S. haemolyticus carried the cytotoxin (pvl) gene, while 14.3% S. aureus harbored the toxic shock syndrome toxin (tst) gene. Multivariable logistic regression analysis identified “Old aged” (OR [CI]: 3.5 [1–12.4]); “Early stage of lactation” (OR [CI]: 3.4 [1.2–9.7]) and, “Firm udder condition” (OR [CI]: 4.2 [1.2–14.6]) as risk factors associated with SCM caused by S. aureus, S. chromogenes, and S. haemolyticus, respectively. Moreover, “Use of antimicrobials” (OR [CI]: 10.4 [3.4–32.1] and “History of previous clinical mastitis” (OR [CI]: 4.9 [1.2–19.7] for the carriage of methicillin-resistant Staphylococcus spp.

Is Helicobacter pylori infection associated with pancreatic cancer? A systematic review and meta-analysis of observational studies.

Recent observational studies have investigated the association between Helicobacter pylori (H. pylori) infection and pancreatic cancer with conflicting data. Therefore, we conducted a systematic review and meta-analysis to assess the potential association. This is a systematic review and meta-analysis. We searched three databases (PubMed, Embase, and Web of Science) from inception to 30 August 2022. The summary results as odds ratio (OR) or hazard ratio (HR) with 95% confidence interval (CI) were pooled by generic inverse variance method based on random-effects model. A total of 20 observational studies involving 67,718 participants were included in the meta-analysis. Meta-analysis of data from 12 case-control studies and 5 nested case-control studies showed that there was no significant association between H. pylori infection and the risk of pancreatic cancer (OR = 1.20, 95% CI = 0.95-1.51, p = 0.13). Similarly, we also did not find significant association between cytotoxin-associated gene A (CagA) positive strains, CagA negative strains, vacuolating cytotoxin gene A (VacA) positive strains H. pylori infection, and the risk of pancreatic cancer. Meta-analysis of data from three cohort studies showed that H. pylori infection was not significantly associated with an increased risk of incident pancreatic cancer (HR = 1.26, 95% CI = 0.65-2.42, p = 0.50). We found insufficient evidence to support the proposed association between H. pylori infection and increased risk of pancreatic cancer. To better understand any association, future evidence from large, well-designed, high-quality prospective cohort studies that accounts for diverse ethnic populations, certain H. pylori strains, and confounding factors would be useful to settle this controversy.

The relationship between Helicobacter pylori and pancreatic cancer: a meta-analysis.

BackgroundThe relationship between Helicobacter pylori (H. pylori, HP) infection and pancreatic cancer would be investigated in this article.MethodsAll cohort studies and case-control studies about H. pylori infection and pancreatic cancer up to October 2021 were searched in the databases of PubMed, Embase and Cochrane. The combined odds ratio (OR) and 95% confidence interval (CI) were calculated by R 4.1.0 software. Funnel plot and Egger test were used to evaluate publication bias.ResultsA total of 17 studies which included 8 case-control studies, 5 nested case-control studies, and 4 cohort studies were included in this study, and the results of this article have confirmed that the H. pylori infection was significantly correlated with the occurrence of pancreatic cancer (OR =1.30, 95% CI: 1.02–1.64), especially in economically underdeveloped areas (OR =2.10, 95% CI: 1.44–3.05). However, negative results were obtained in the relationship between CagA + H. pylori and pancreatic cancer. Similarly, we also did not find an association between vacuolating cytotoxin gene A-positive strains (VacA-positive H. pylori) and pancreatic cancer. The heterogeneity of this study was significant. Through a sensitivity analysis by the leave-one-out method, we found the results remained unchanged on the whole but the correlation between H. pylori infection and the occurrence of pancreatic cancer in the Asian population was significant. The tests for funnel plot asymmetry indicated that there might be obvious publication bias in this study. After carrying out the Egger test, we proved the existence of the publication bias in this study, which could have a certain impact on the results.DiscussionBased on the currently available data, we confirm that H. pylori infection can increase the incidence of pancreatic cancer in general. CagA/VacA-positive H. pylori infection is not associated with the incidence of pancreatic cancer. H. pylori infection is significantly associated with the incidence of pancreatic cancer in economically underdeveloped areas, while the relationship between H. pylori infection and the incidence of pancreatic cancer in the Asian population is uncertain. In addition, more high-quality studies are needed to be included to confirm this conclusion.

Association of PVL Gene in MSSA and MRSA Strains among Diabetic Ulcer Patients from Odisha, India.

Staphylococcus aureus has emerged as an important pathogen among diabetic foot ulcers in patients with diabetes. Infections with S. aureus in diabetic ulcers need surveillance of resistant microbial profile to provide the basis for empirical therapy for the reduction of lower extremities amputation. Panton valentine leucocidin (PVL) is considered as one of the major virulence gene of S. aureus which is responsible for destruction of white blood cells and tissue necrosis. This pore forming cytotoxin gene is carried out by both methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains. The present study described the prevalence of PVL gene in MSSA and MRSA strains isolated from diabetic ulcer patients treated during November, 2019 to January, 2021 from a tertiary care hospital, Odisha. Infected tissue and blood samples from these patients were collected aseptically and sub-cultured using different media and standard techniques. The isolated genomic DNA of MSSA and MRSA strains were subjected to PCR assay for the detection of PVL gene. Two hundred ten S. aureus out of 402 diabetic ulcer patients were isolated having 59.52% MSSA and 40.47% MRSA strains. Wagner's grade III and grade IV ulcers were most prevalent in these ulcer patients. The prevalence of PVL gene in MSSA strains was more in comparison to MRSA strains. Forty five resistance patterns were observed from the antibiogram profiles of S. aureus. The present study highlighted that PVL gene could not be a marker for the detection of MRSA and MSSA strains in diabetic ulcer patients.

Exploring the potential of Cinnamomum zeylanicum oil against drug resistant Helicobacter pylori-producing cytotoxic genes.

Thirty-one of sixty dyspeptic patients tested positive for Helicobacter pylori colonization in this study, as determined by histopathology and 16S rRNA. The cytotoxin-associated gene A (cagA) and vacuolating cytotoxin A (vacA) genes were found in 67.7 and 93.5% of H. pylori patients, respectively. The cagA gene was found to be associated with 100% of patients with duodenal erosion and ulceration identified via endoscopy examination. In addition, 86.7% of patients with cancerous and precancerous lesions, glandular atrophy, and intestinal metaplasia identified via histopathology examination. The vacA s1m1 mutation was associated with more severe forms of gastric erosion and ulceration, as well as the presence of precancerous and cancerous lesions. Eighteen (64.3%) of the twenty-eight isolates were classified as multi-drug resistant (MDR) or pan-drug resistant (PDR) H. pylori. Due to a resurgence of interest in alternative therapies derived from plants as a result of H. pylori resistance to the majority of commonly used antibiotics, the inhibitory activity of five essential oils extracted from some commonly used medicinal plants was evaluated in vitro against drug-resistant H. pylori clinical isolates. Cinnamomum zeylanicum essential oil demonstrated the highest anti-H. pylori activity when compared to the other essential oils tested. Cinnamaldehyde was the most abundant compound in C. zeylanicum (65.91%). The toxicological evaluation established the safety of C. zeylanicum oil for human use. As a result, C. zeylanicum essential oil may represent a novel antibacterial agent capable of combating drug-resistant H. pylori carrying cytotoxin genes.

Distribution of virulence factors, antimicrobial resistance genes and phylogenetic relatedness among Shiga toxin-producing Escherichia coli serogroup O91 from human infections

Shiga toxin-producing Escherichia coli (STEC) belonging to the serogroup O91 are among the most common non-O157 STEC serogroups associated with human illness in Europe. This study aimed to analyse the virulence factors, antimicrobial resistance genes and phylogenetic relatedness among 48 clinical STEC O91 isolates collected during 2003–2019 in Switzerland. The isolates were subjected to whole genome sequencing using short-read sequencing technologies and a subset of isolates additionally to long-read sequencing. They belonged to O91:H10 (n=6), O91:H14 (n=40), and O91:H21 (n=2). Multilocus sequence typing showed that the O91:H10 isolates all belonged to sequence type (ST)641, while the O91:H14 isolates were assigned to ST33, ST9700, or were non-typeable. Both O91:H21 isolates belonged to ST442. Shiga toxin gene stx1a was the most common Shiga toxin gene subtype among the isolates, followed by stx2b, stx2d and stx2a. All isolates were LEE-negative and carried one or two copies of the IrgA adhesin gene iha. In a subset of long-read sequenced isolates, modules of the Locus of Adhesion and Autoaggregation pathogenicity island (LAA-PAI) carrying iha and other genes such as hes, lesP or agn43 were identified. A large proportion of STEC O91:H14 carried the subtilase cytotoxin gene subA, colicin genes (cba, cea, cib and cma) or microcin genes (mcmA, mchB, mchC and mchF). STEC O91:H14 were further distinguished from STEC O91:H10/H21 by one or more virulence factors found in extraintestinal pathogenic E. coli (ExPEC), including hlyF, iucC/iutA, kpsE and traT. The hlyF gene was identified on a novel mosaic plasmid that was unrelated to hlyF+ plasmids described previously in STEC. Core genome phylogenetic analysis revealed that STEC O91:H10 and STEC O91:H21 were clonally conserved, whereas STEC O91:H14 were clonally diverse. Among three STEC O91:H14 isolates, a number of resistance genes were identified, including genes that mediate resistance to aminoglycosides (aadA, aadA2, aadA9, aadA23, aph(3'')-Ib and aph(6)-Id), chloramphenicol (cmlA), sulphonamides (sul2 and sul3), and trimethoprim (drfA12). Our data contribute to understanding the genetic diversity and differing levels of virulence potential within the STEC O91 serogroup.

The crosstalk between H. pylori virulence factors and the PD1:PD-L1 immune checkpoint inhibitors in progression to gastric cancer

BackgroundThe progression to gastric cancer has been linked to chronic infection with Helicobacter pylori (H. pylori). Immune checkpoint inhibitors (programmed cell death -1, PD-1; programmed cell death –ligand 1, PD-L1) have a role in cancer immune escape. The relationship between H. pylori virulence factors with PD-1, PD-L1 T helper 1 (Th1), T helper 17 (Th17), and regulatory T cell (Treg) response genes, has not been thoroughly investigated in the development of gastric cancer. Therefore, we evaluated how H. pylori virulence factors influence the expression levels of immune-related genes in the development of gastric immunopathology. MethodsA total of 92 gastric tissues of normal controls and patients with gastritis, gastric ulcer, and gastric cancer were examined for the expression of immune-checkpoint inhibitor genes (PD-1 PD-L1), Th1 (interferon- γ, IFN-γ), Th17 (interleukin- 17, IL-17, Retinoic-acid-receptor- related orphan nuclear receptor gamma t, RORγ-t), and Treg (Forkhead box P3, FOXP3) response genes with quantitative real-time PCR (qRT-PCR). Furthermore, correlation of H. pylori virulence factors’ (cytotoxin-associated gene A, cagA; vacuolating cytotoxin gene A, vacA (s1,s2,m1,m2); blood group antigen-binding adhesin gene A, babA, duodenal ulcer promoting gene A, dupA; the putative neuraminyllactose-binding hemagglutinin homolog, hpaA; neutrophil-activating protein A napA; outer inflammatory protein A, oipA; urease A, ureA; and urease B, ureB) genotypes with a degree of inflammation and density of H. pylori were investigated. Next, the relationship between H. pylori virulence factors and immune-checkpoint inhibitor genes, and T-cell response genes was evaluated. Eventually, a decision tree model was developed to determine the clinical outcome of patients using expression data. ResultsThe intensity of PD-1 and PD-L1 mRNA expression was increased significantly in gastric tissue of patients with gastric ulcer (PD-1: 2.3 fold, p=0.01; PD-L1: 2.1 fold, p=0.004), and gastric cancer (PD-1: 2 fold, p= 0.04; PD-L1: 1.8 fold, p=0.05) compared with control subjects. Also, PD-1: PD-L1 expression was significantly higher in patients with gastritis, who were infected with a marked density of H. pylori compared with its mildly infected counterparts. Furthermore, a novel negative correlation was found between PD-1 (r= -0.43) and PD-L1 (r= -0.42) with FOXP3 in patients with gastritis. CagA-positive H. pylori strain's negative association with PD-L1 expression (r=-0.34) was detected in patients with gastritis. Interestingly, PD-1 mRNA expression correlated positively with vacA s2/m2, in gastritis (r=0.43) and ulcer (r=0.43) patients. Furthermore, PD-1: PDL1 expression negatively correlated with vacA m1/m2 (r=-0.43 for PD-1; r=-0.38 for PD-L1) in gastritis patients. Moreover, an inverse correlation of PDL1 was present with vacA m1 (r=0.52) and vacA s1/m1 (r=0.46) versus vacA m2 (r=-0.44) and vacA m1 (r=0.52) and vacA s1/m2 (r=-0.14) in ulcer patients, respectively. Also, a correlation of vacA m2 (r=-0.47) and vacA s1/s2 (r= 0.45) with PD-1 was detected in ulcer patients. In addition, a novel negative correlation between FOXP3 mRNA levels and napA was shown in patients with gastritis and ulcer (r=-0.59). Finally, a computer-based model that was developed showed that knowing the expression levels of PD-L1, RORγ-t, and vacA s1/m2 would be useful to detect the clinical outcome of a patient. ConclusionOur results suggested that PD-1:PD-L1 immune checkpoint inhibitors were increased in gastric pre-cancerous lesions that progress to gastric cancer. Herein, we report the relationship between H. pylori virulence factors and expression of host immune checkpoint inhibitors for diagnostic prediction of gastric malignancies using computer-based models.

Genotyping of Helicobacter pylori Virulence Genes cagA and vacA: Regional and National Study.

Helicobacter pylori (H. pylori) plays a crucial role in the pathogenesis of gastritis, peptic ulcer, and gastric cancer. The presence of pathogenicity islands (PAI) genes contributes to the pathogenesis of many gastrointestinal disorders. Cytotoxin-associated gene A (cagA) and vacuolating cytotoxin gene (vacA) are the most known virulence genes in H. pylori. So, our aim was to study H. pylori virulence genes' role in gastric disorders pathogenesis. Our study included 150 adult patients who suffered dyspeptic symptoms and were referred to the GIT endoscopy unit. Gastric biopsies were attained for rapid urease test (RUT) and histopathological examination, and multiplex PCR technique for detection of virulence genes was performed. It was found that 100 specimens were (RUT) positive, of which sixty samples (60%) were PCR positive for H. pylori ureC gene. The vacA and cagA genes were identified in 61.6% and 53% of H. pylori strains, respectively. Only 5 cases were vacA-positive and cagA-negative. The most virulent vacA s1 allele existed in 56.6% of cases. Out of the 60 H. pylori strains, 66% had at least one virulence gene and 34% did not show any virulence gene. H. pylori infection showed significant increase with age. H. pylori are prevalent amid dyspeptic patients in our region. The main genotype combinations were vacA+/cagA+ of s1m1 genotype and they were frequently associated with peptic ulcer diseases, gastritis, and gastroesophageal reflux disease.

Gastroprotective effect of phytoncide extract from Pinus koraiensis pinecone in Helicobacter pylori infection

For centuries, herbs have been used by traditional therapists around the world to treat gastrointestinal tract disorders, such as gastritis. We hypothesized that the anti-Helicobacter pylori properties of phytoncide, which is extracted from pinecone waste, would facilitate use as a natural gastroprotective product to treat gastrointestinal tract disorders. Thus, we investigated in vitro antibacterial efficacy against H. pylori by agar diffusion assay. To determine the gastroprotective properties of phytoncide, we conducted hematoxylin and eosin staining, performed assays for the detection of the cytotoxin gene, and evaluated pro-inflammatory cytokine expression in H. pylori-infected C57BL/6 mice. Phytoncide significantly inhibited the survival of H. pylori in the gastrointestinal system of C57BL/6 mice. Reduction of gastric severity in H. pylori-infected mice was associated with reductions in the expression levels of pro-inflammatory cytokines in the gastric mucosa, and of the cytotoxin CagA gene in phytoncide treated groups (P < 0.05 and P < 0.01). In conclusion, phytoncide significantly inhibited the growth of H. pylori in gastro tissue, possibly due to the abundant α-pinene present in the phytoncide as detected by HPLC analysis. Further studies are needed to validate our findings, but we suggest that phytoncide has the potential to be used as a natural ingredient in anti-H. pylori products.

Whole-genome sequence analysis of Vibrio cholerae from three outbreaks in Uganda, 2014 - 2016

Background: Cholera remains a serious public health problem in Uganda and Africa. The aim of this study was to provide the complete array of antimicrobial resistance genes, integrative and conjugative elements, virulence genes, pathogenicity islands, plasmids, and insertion sequences in the strains. In addition, this study also aimed to provide a single nucleotide polymorphism (SNP) based phylogenetic analysis of the strains. Methods: In the analysis, both Linux and web-based bioinformatics approaches were used to analyze the study sequences. Databases used included; FastQC, MultiQC, Snippy, PANTHER, PATRIC, Unicycler, ISFinder, Center for Genomic Epidemiology pipelines (i.e. MLST, PlasmidFinder, MyDbFinder, and ResFinder), MashTree and IcyTree. Results: The 10 sequenced strains of Vibrio cholerae were found to carry virulence-associated genes including MakA, ctxA, ctxB, carA, carB, trpB, clpB, ace, toxR, zot, rtxA, ompW, ompR, gmhA, fur, hlyA, and rstR. Also identified were: genes of the Type VI secretion system including vasA-L, vgrG-2, vgrG-3, vipA/mglA, and vipB/mglB; alsD (VC1589), involved in the synthesis of 2,3-butanediol; alsR, involved in the acetate-responsive LysR-type regulation; makA, the flagella-mediated cytotoxin gene; Type VI pilus genes including tcpA-F, tcpH-J, tcpN, tcpP-T, and icmF/vasK; adherence genes acfA-D and IlpA; and quorum sensing system genes luxS and cqsA. Pathogenicity islands identified comprised of VSP-1 and VSP-2, as well as VPI-1 and VPI-2. In addition, strA and B, APH(3'')-I, APH(3'')-Ib, APH(6)-Id, APH(6)-Ic, murA, pare, dfrA1, floR, catB, and catB9 were among the antimicrobial resistance genes found in the sequences. Analysis for SNPs shared among the sequences showed that the sequenced strains shared 218 SNPs and of these, 98 SNPs were missense. Gene enrichment analysis of these SNPs showed enrichment in genes that mediate transmembrane-signaling receptor activity, peptidyl-prolyl cis-trans isomerase activity, and phosphor-relay response regulator activity. Conclusions: This study applied bioinformatics approaches to provide comprehensive genomic analysis of V. cholerae genomes obtained from Uganda.

The association between iceA and vacA allelic gene variations of Helicobacter pylori strains and gastric disorders

Helicobacter pylori is a causative agent of gastric disorders and associated with gastric adenocarcinoma. Different kind of gene products such as induced by contact with epithelium (iceA), vacuolating cytotoxin (vacA), and cytotoxin-associated gene A (cagA) confers the ability to this bacterium to destroy gastric cells and can lead to gastric cancer. The aim of the present study was to evaluate the frequency and association of iceA and vacA allelic genes variations of H. pylori strains and gastric disorders. This cross-sectional study included 90 nonduplicated H. pylori isolates from gastric biopsy of 300 patients in the north of Iran. DNA was extracted from all isolated strains and polymerase chain reaction (PCR) was used to determine the prevalence of iceA and vacA genes allelic variants. In overall, from 90 samples of H. pylori iceA, vacA s1/s2, vacA m1 and vacA m2 genes were detected in 68.9%, 33.3%, 53.3%, and 37.8% of patients, respectively. Regard to the distribution of the virulence genes, the co-occurrence of iceA/vacA m1 (20%) was the most prevalent genotype followed by iceA (14.4%) and iceA/vacA m2 (10%). In summary, as a first reports from Guilan province in the north of Iran we showed the association of iceA and vacA allelic variations in different types of gastric disorders. In this regard, we found a significant association between the presence of iceA/vacA m1 genotype and gastric diseases.