Helicobacter pylori, a Gram-negative gastric pathogen and a class I carcinogen harbors key virulence genes such as vacA, cagA, ureC, homB and oipA that contributes to gastrointestinal diseases. The study assessed the prevalence of H. pylori infection and their correlation with virulence genotypes with clinical outcomes in South Indian Tamils. Gastric antral biopsies from 500 patients were assessed for H. pylori prevalence by RUT followed by genomic DNA extraction and PCR. Molecular confirmation employed 16S rRNA, a preliminary marker, with the identification of virulent genes cagA, vacAs1, vacA m1/m2, oipA, ureC, homB are validated by sequencing. H. pylori were confirmed in 491 patients where vacA s1 (71.89%) was correlated with gastritis (OR-1.68;95%; CI: 1.11-2.53; p = 0.013) and pangastritis (OR-4.79;95%; CI: 2.02-11.34; p = 0.0004) while vacA s1/m1 and vacA s1/m2 alleles were found to be 51.53% and 14.26%, respectively. Gastritis was associated with s1/m1 (OR-2.34; 95%CI: 1.29-4.23; p = 0.004), s1/m2 (OR-2.03; 95%CI: 1.04-3.95; p = 0.03) and pangastritis (OR-2.61;95%CI: 0.97-7.02; p = 0.05) was associated with s1/m1. The cagA gene (31.36%) was correlated to gastric cancer (OR-3.45; 95%CI: 1.20-9.89; p = 0.02) and oipA gene (64.97%) was associated with PUD (OR = 7.87; 95% CI: 1.02–60.40; p = 0.04), homB (24.03%) with gastritis (OR-1.62; 95%CI: 1.01-2.59; p = 0.04) and ureC (59.67%) with no disease association. Therefore, our study provides genotypic prevalence of H. pylori infection in the South Indian Tamil population, with vacA s1 being the predominant genotype reported and significantly associated with gastritis and pangastritis. Hence, vacA s1 can serve as a potent virulent marker for gastrointestinal disease manifestations.