High prevalence of cagA positive Vac A s1m1 Helicobacter pylori strains isolated from patients suffering from various gastroduodenal diseases in Guwahati, Assam, India.

AIM AND BACKGROUND: Genotyping of H. pylori strains was identified on formalin-fixed paraffin wax-embedded tissue (FFPE) sections and correlated with severity grades of gastric mucosal pathologies in biopsies from upper gastrointestinal (UGI) tract from Guwahati, Assam. MATERIALS AND METHODS: In total, 168 gastric biopsies collected from patients with UGI symptoms underwent histological evaluation as per the updated Sydney system. RESULT: H. pylori-like organisms were identified with Warthin and Starry stain, and virulent genes were amplified using polymerase chain reaction (PCR) from genomic DNA extracted from FFPE sections by using QIAamp® DNA FFPE Tissue Kit. Histological examination identified H. pylori-like organisms in 100 biopsies, of which 96 were urease + ve. The prevalence of H. pylori infection was high in age groups 71-80 (88.8%) as compared to other age groups, and it was higher in females (78.9%) when compared to males. The prevalence of virulent genes in biopsies was 88.5% cagA and vacA s1m1, 31.2% iceA1, 32.2% iceA2, and 85.2% babA2. The histological parameters mononuclear cell infiltrate (P = 0.04) and atrophy (P = 0.03), showed statistically significant association with iceA2 and intestinal metaplasia with cagA (P = 0.01) vacAs1m1 (P = 0.01) and babA (P = 0.02) genotypes. Gastric erosion due to H. pylori infection and atrophy showed a significant association. A high bacterial density score was seen with the virulent genotypes. CONCLUSION: Our work reports for the first time a high prevalence (88.5%) of H. pylori cagA vacA s1m1 genotype in Guwahati, Assam. Association of gastric atrophy and intestinal metaplasia was seen with virulent genotypes. Results show the effectiveness of the FFPE kit for DNA extraction in remote areas where transportation and storage of biopsies are otherwise difficult.

Altered expression of endosomal Toll-like receptors and HBeAg seropositivity may act synergistically towards the vertical transmission of HBV.

PROBLEM: Hepatitis B is one of the leading causes of mortality in India. Despite the mass vaccination programme, the burden of the infection is still increasing due to its vertical transmission. Asymptomatic nature of hepatitis B virus (HBV) infection owing to immune tolerance among pregnant women is a major issue in this regard. METHOD OF STUDY: As such, this study aims to investigate the potential role of altered Toll-like receptor (TLR) expression (TLR-3, 7 and 9) along with peripheral blood HBeAg status in attaining differential cord blood (CB) HBV DNA status. RESULT: Expression analysis reveals an overall downregulation of expression with mean ± SD value 1.14 ± 1.05, 0.86 ± 0.5 and 0.71 ± 0.4 (TLR 3, 7 and 9, respectively) upon comparison with healthy women. Further stratification based on CB HBV DNA status; the downregulation of expression was found to be significantly (p < .05) associated with positive CB HBV DNA status apart from peripheral HBeAg status. One hundred percent HBeAg positive parturiting women exhibit positive CB HBV DNA. Pearson's correlation analysis reveals a positive correlation between CB HBV DNA status and altered TLR expression, HBeAg status and mother HBV DNA status and as such can be associated with the potential risk of HBV vertical transmission. CONCLUSION: This study suggests that the downregulation of TLR 3, 7 and 9 may be a risk factor for potential vertical transmission of HBV.

Human Leukocyte Antigen DQ (HLA-DQ) genotypes and haplotypes and their association with phenotype in patients with celiac disease in India.

BACKGROUND AND AIM: Human Leukocyte Antigen DQ (HLA-DQ) genotypes play a permissive role in the genesis of celiac disease (CeD). In this case-control study, we used next-generation sequencing to determine HLA-DQA1 and ~DQB1 genotypes and haplotypes associated with CeD in Indian patients. METHODS: HLA-DQA1 and ~DQB1 loci were amplified, using long-range polymerase chain reaction (PCR), from DNA of 259 patients with symptomatic CeD (160 typical and 99 atypical), 45 asymptomatic CeD, 96 potential CeD, and 300 healthy adults. Amplicons were fragmented and sequenced on the Illumina platform, and alleles and haplotypes were assigned by matching against the HLA-international ImMunoGeneTics (IMGT) database. RESULTS: HLA-DQA1*05:01 (odds ratio [OR] 8.39, 95% confidence interval [CI] 5.64-12.47) and HLA-DQB1*02:01 (OR 8.59, 95% CI 5.75-12.83) were the genotypes that showed a risk association with symptomatic CeD. Among the haplotypes, HLA-DQA1*05:01 ~ HLA-DQB1*02:01 (OR 8.56, 95% CI 5.67-13.19) showed a strong risk association with symptomatic CeD. When comparing symptomatic CeD with subclinical forms (asymptomatic and potential) CeD, HLA-DQA1*05:01 ~ HLA-DQB1*02:01 (OR 2.34, 95% CI 1.61-3.43) was significantly associated with risk of symptomatic disease. The strength of association between the HLA-DQA1*05:01 ~ HLA-DQB1*02:01 haplotype and the CeD phenotype showed a gradient in the order typical > atypical > asymptomatic > potential CeD. Genotypes consistent with expression of HLA DQ2 and/or 8 were noted in 128 (80%) typical, 73 atypical (74%), 27 (60%) asymptomatic, and 52 (54%) potential CeD participants. CONCLUSION: HLA-DQA1*05:01 ~ HLA-DQB1*02:01 (haplotype DQ2.5) showed a very strong risk association with symptomatic CeD in Indian patients. The strength of association showed a gradient of increase from potential to typical CeD, coinciding with a phenotypic change in the celiac iceberg.

In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity.

As the COVID-19 infection continues to ravage the world, the advent of an efficient as well as the economization of the existing RT-PCR based detection assay essentially can become a blessing in these testing times and significantly help in the management of the pandemic. This study demonstrated an innovative and rapid corroboration of COVID-19 test based on innovative multiplex PCR. An assessment of optimal PCR conditions to simultaneously amplify the SARS-CoV-2 genes E, S and RdRp has been made by fast-conventional and HRM coupled multiplex real-time PCR using the same sets of primers. All variables of practical value were studied by amplifying known target-sequences from ten-fold dilutions of archived positive samples of COVID-19 disease. The multiplexing with newly designed E, S and RdRp primers have shown an efficient amplification of the target region of SARS-CoV-2. A distinct amplification was observed in 37 min using thermal cycler while it took 96 min in HRM coupled real time detection using SYBR green over a wide range of template concentrations. Our findings revealed decent concordance with other commercially available detection kits. This fast HRM coupled multiplex real-time PCR with SYBR green approach offers rapid and sensitive detection of SARS-CoV-2 in a cost-effective manner apart from the added advantage of primer compatibility for use in conventional multiplex PCR. The highly reproducible novel approach can propel extended applicability for developing sustainable commercial product besides providing relief to a resource limited setting.

Geographically distinct North-East Indian Helicobacter pylori strains are highly sensitive to clarithromycin but are levofloxacin resistant.

Purpose: Helicobacter pylori causes various gastro-intestinal diseases. Antibiotic resistance to commonly used antibiotics for the treatment of H. pylori infection is the major cause for treatment failure. The aim of this study is to determine the antimicrobial susceptibility pattern for clarithromycin and levofloxacin and find the evolutionary relationship of the partial sequence of 23S rRNA and gyraseA gene of H. pylori by phylogenetic analysis. Materials and Methods: A total of 46 H. pylori strains were tested for clarithromycin and levofloxacin susceptibility pattern and phylogenetic tree were reconstructed by PhyML software. Results: In this study, we observed that only 6.5% of North-East Indian H. pylori strains were resistant for clarithromycin showing mutation at A2143G and T2182C positions of 23S rRNA gene. Resistance for levofloxacin was observed in 89.1% of the H. pylori strains showing mutations at asparagine to lysine at 87 and aspartic acid to glycine/tyrosine/asparagine at 91 positions of gyraseA gene. The phylogenetic tree of the partial sequence of 23S rRNA and gyraseA gene depicts that the North-East Indian strains falls in different cluster when compared to other countries. Conclusions: Resistance for clarithromycin was less in North-East Indian strains but high for levofloxacin indicating that first-line therapy may be best and effective for eradication of H. pylori in this region. This study is the first report that showed antibiotic susceptibility pattern for clarithromycin and levofloxacin by mutation analysis. By partial sequencing of 23s rRNA and gyraseA gene, we found that North-East Indian strains are geographically distinct.

Prevalence of Adult Celiac Disease in India: Regional Variations and Associations.

OBJECTIVES: Although celiac disease (CeD) affects 1% of people in the northern part of India, it is believed to be uncommon in the southern and northeastern parts because of significant differences in dietary pattern and ethnicity. We estimated the prevalence of CeD in these three populations. In a subset, we also investigated differences in the prevalence of HLA-DQ 2/8 allelotype and dietary grain consumption. METHODS: A total of 23,331 healthy adults were sampled from three regions of India-northern (n=6207), northeastern (n=8149), and southern (n=8973)-and screened for CeD using IgA anti-tissue transglutaminase antibody. Positive tests were reconfirmed using a second ELISA. CeD was diagnosed if the second test was positive and these participants were further investigated. A subsample of participants was tested for HLA-DQ2/-DQ8 and underwent detailed dietary evaluation. RESULTS: Age-adjusted prevalence of celiac autoantibodies was 1.23% in northern, 0.87% in northeastern, and 0.10% in southern India (P<0.0001). Prevalence of CeD and latent CeD, respectively, was 8.53/1,000 and 3.70/1,000 in northern, 4.66/1,000 and 3.92/1,000 in northeastern, and 0.11/1,000 and 1.22/1,000 in the southern part. The population prevalence of genes determining HLA-DQ2 and/or -DQ8 expression was 38.1% in northern, 31.4% in northeastern, and 36.4% in southern India. Mean daily wheat intake was highest in northern (455 g) compared with northeastern (37 g) or southern part (25 g), whereas daily rice intake showed an inverse pattern. CONCLUSIONS: CeD and latent CeD were most prevalent in northern India and were the least in southern India. The prevalence correlated with wheat intake and did not reflect differences in the genetic background.