Distinct repeat motifs at the C-terminal region of CagA of Helicobacter pylori strains isolated from diseased patients and asymptomatic individuals in West Bengal, India

Background: Infection with Helicobacter pylori strains that express CagA is associated with gastritis, peptic ulcer disease, and gastric adenocarcinoma. The biological function of CagA depends on tyrosine phosphorylation by a cellular kinase. The phosphate acceptor tyrosine moiety is present within the EPIYA motif at the C-terminal region of the protein. This region is highly polymorphic due to variations in the number of EPIYA motifs and the polymorphism found in spacer regions among EPIYA motifs. The aim of this study was to analyze the polymorphism at the C-terminal end of CagA and to evaluate its association with the clinical status of the host in West Bengal, India. Results: Seventy-seven H. pylori strains isolated from patients with various clinical statuses were used to characterize the C-ternimal polymorphic region of CagA. Our analysis showed that there is no correlation between the previously described CagA types and various disease outcomes in Indian context. Further analyses of different CagA structures revealed that the repeat units in the spacer sequences within the EPIYA motifs are actually more discrete than the previously proposed models of CagA variants. Conclusion: Our analyses suggest that EPIYA motifs as well as the spacer sequence units are present as distinct insertions and deletions, which possibly have arisen from extensive recombination events. Moreover, we have identified several new CagA types, which could not be typed by the existing systems and therefore, we have proposed a new typing system. We hypothesize that a cagA gene encoding higher number EPIYA motifs may perhaps have arisen from cagA genes that encode lesser EPIYA motifs by acquisition of DNA segments through recombination events

A study to assess the level of knowledge regarding prevention and management of acute respiratory infection among mothers of children 0-5 years in selected hospital in Siliguri

Background: In developing countries like India acute respiratory infection (ARI) contributes in child mortality upto 75% and out of 10, 7 deaths are due to ARI. The knowledge of the mothers towards the disease is a significant determinant of child’s health.Methods: A descriptive cross-sectional study included 100 mothers of children 0-5 years admitted in pediatric ward and postnatal ward in selected hospital Siliguri during the year 2022 in the month of March. Data was collected using structured interview method.Results: 20% of mothers have good knowledge in prevention and 33% had good knowledge in management of ARI.Conclusions: As the leading cause of death among children, knowledge assessment about ARI among the mothers is very important, which helps for better understanding of the intensity of the problem

Multiple infection and microdiversity among Helicobacter pylori isolates in a single host in India.

Helicobacter pylori is one of the most diverse bacterial species that chronically infects more than 70% of Indian population. Interestingly, data showing microdiversity of the H. pylori strains within a particular gastric niche remained scarce. To understand the extent of genetic diversity among H. pylori strains within a given host, 30 patients with gastro-duodenal problems were subjected to endoscopy and from each patient 10 single colonies were isolated. Characterization of each of these 10 single colonies by DNA fingerprinting as well as genotyping of several important genetic markers viz. cagA, vacA, iceA, vapD, cag PAI empty site, IS605, RFLP and two other genetic segments within cag PAI revealed that all of the 30 patients were infected with more than one strain and sometimes strains with 5 to 6 types of genetic variants. Analyses of certain genetic loci showed the microdiversity among the colonies from single patient, which may be due to the recombination events during long-term carriage of the pathogen. These results suggest that most of the patients have acquired H. pylori due to repeated exposure to this pathogen with different genetic make-up, which may increase the possibility of super infections. Genetic exchanges between these unrelated H. pylori strains may support certain H. pylori variant to grow better in a given host than the parental strain and thereby increasing the possibility for the severity of the infection

Indistinguishable cellular changes in gastric mucosa between Helicobacter pylori infected asymptomatic tribal and duodenal ulcer patients

AIM: To investigate the changing pattern of different histological parameters occurring in the stomach tissue of Helicobacter pylori (H pylori) infected tribal populations and duodenal ulcer patients among ethnic Bengalis and correlation of the genotypes of H pylori with different histological parameters. METHODS: One hundred and twelve adult individuals were enrolled into this study between 2002 and 2004. Among them, 72 had clinical features of duodenal ulcer (DU) from ethnic Bengali population and 40 were asymptomatic ethnic tribals. Endoscopic gastric biopsy samples were processed for histology, genotyping and rapid urease test. Histologically, haematoxylin and eosin staining was applied to assess the pathomorphological changes and a modified Giemsa staining was used for better detection of H pylori. For intestinal metaplasia, special stainings, i.e. Alcian blue periodic acid-Schiff and high iron diamine-Alcian blue staining, were performed. PCR was performed on bacterial DNA to characterize the presence or absence of virulence-associated genes, like cagA, and distribution of different alleles of vacA and iceA. RESULTS: Intraglandular neutrophil infiltration, a hallmark of activity of gastritis, was present in 34 (94%) of tribals (TRs) and 42 (84%) of DU individuals infected with H pylori. Lymphoid follicles and aggregates, which are important landmarks in H pylori infection, were positive amongst 15 (41%) of TRs and 20 (40%) of DU subjects. Atrophic changes were observed in 60% and 27.7%, respectively, among DU cases and tribals (P > 0.003). Metaplastic changes were detected in low numbers in both groups. Moderate to severe density distribution of H pylori in the gastric mucosa was 63% among TRs, whereas it was 62% in DU subjects. There were no significant differences in the distribution of virulence-associated genes like cagA, vacA and iceA of H pylori strains carried by these two populations. CONCLUSION: Our study showed almost similar distribution of inflammatory cells among asymptomatic tribals and DU Bengali patients. Interestingly, the tribal population are free from any clinical symptoms despite evidence of active histologic gastritis and infection with H pylori strains carrying similar virulence markers as of strains isolated from patients with DU. There was an increased cellular response, especially in terms of neutrophil infiltration, but much lower risk of developing atrophy and metaplastic changes among the tribal population

Intact cag pathogenicity island of Helicobacter pylori without disease association in Kolkata, India

Several genes including the cagA in the cag pathogenicity island (cag PAI) of Helicobacter pylori are thought to be associated with the gastroduodenal diseases and hence variation in the genetic structure of the cag PAI might be responsible for different clinical outcomes. Our study was undertaken to characterize the cag PAI of H. pylori strains from duodenal ulcer (DU) patients and asymptomatic or non-ulcer dyspepsia (NUD/AV) subjects from Kolkata, India. Strains isolated from 52 individuals (30 DU and 22 NUD/AV) were analyzed by PCR using 83 different primers for the entire cag PAI and also by dot-blot hybridization. Unlike H. pylori strains isolated from other parts of India, 82.6% of the strains used in this study had intact cag PAI, 9.6% had partially deleted cag PAI, and 7.7% of the strains lacked the entire cag PAI. Dot-blot hybridization yielded positive signals in 100% and 93.8% of PCR-negative strains for HP0522-523 and HP0532-HP0534 genes, respectively. An intact cagA promoter region was also detected in all cagA-positive strains. Furthermore, the expression of cagA mRNA was confirmed by RT-PCR for the representative strains from both DU and NUD/AV subjects indicating the active cagA promoter regions of these strains. A total of 66.7% of Kolkata strains produced a ~390-bp shorter amplicon than the standard strain 26695 for the HP0527 gene, homologue of virB10. However, sequence analyses confirmed that the deletion did not alter the reading frame of the gene, and mRNA transcripts were detected by RT-PCR analysis. The strains isolated from DU and NUD/AV express CagA protein and possess a functional type IV secretion system, as revealed by Western blot analyses. Interestingly, no significant differences in cag PAI genetic structure were found between DU and NUD/AV individuals suggesting that other bacterial virulence factors, host susceptibility, and environmental determinants also influence the disease outcome at least in certain geographical locations

Multiplex PCR Assay for Rapid Detection and Genotyping of Helicobacter pylori Directly from Biopsy Specimens

We developed and evaluated a simple, novel multiplex PCR assay for rapid detection of Helicobacter pylori infection and for the determination of vacA and cagA genotypes directly from gastric biopsy specimens. This assay did not require culturing of strains or extraction of DNA from biopsy samples. This multiplex PCR assay would be of particularly great value for laboratories in developing countries

Multiple strain colonization detected on the basis of HP0527 gene in <i>cag</i> PAI.

<p>Lanes 1 to 10, single colonies isolated from the patient; C, Control strain 26695. (a) Three types of colonies were identified in PG93. Lane 1 and 6 gave a higher amplicon than that of 26695; lanes 3, 4, 7 and 8 yielded same amplicon while lanes 2, 5, 9, 10 produced lower amplicon than that of 26695. (b) Mixed infections detected on the basis of <i>vapD</i> genetic locus PCR. M, 100 bp marker; lanes 1–10, single colonies isolated from PG137; 11, positive control (PCR225); 12, Negative control (<i>E. coli</i> DNA). All the colonies are positive for <i>vapD</i> except colony numbers 1, 6, 9 and 10.</p

Over all analysis regarding the colonization of multiple <i>H. pylori</i> strains in same host in this study population.

<p>Over all analysis regarding the colonization of multiple <i>H. pylori</i> strains in same host in this study population.</p

Combination of genotype and RAPD analysis for PG157 indicated multiple infections and microdiversity in a single host.

<p>M, 100 bp marker; lanes 1–10, single colonies isolated from PG157; P, pooled DNA. (a) RAPD patterns using primer 1281 showed two distinct patterns (lanes 1–8 and lanes 9–10) (b) RAPD patterns using primer 1283 also yielded two distinct patterns (lanes 1–8 and lanes 9–10) (c) Multiplex PCR for <i>vacA</i> alleles and <i>cagA</i> showed existence of s1m1<i>cagA</i>⁺ strains in lanes 1–8 and s2m2<i>cagA</i>⁻ strains in lanes 9–10. (d) Variant <i>cagA</i> subtypes detected on the basis of PCR for 3′ end of <i>cagA</i> using primers CAG1 and CAG2. This PCR assay showed existence of type A strains in lanes 4, 6–8 and existence of type B/D strains in lanes 1–3 and 5. Lanes 9–10, which were detected as <i>cagA</i>⁻, did not produce any amplicon and the pooled sample yielded amplicons for type A and type B/D strains.</p