A transcriptomic approach in elucidating helicobacter pylori pathogenesis and occurrence in human and periplaneta / Noor Masyitah Jumahat

Helicobacter pylori (H. pylori) is a Gram-negative spiral-shaped bacterium that colonises the human stomach. H. pylori infection constitutes an established risk factor in the development of gastritis, peptic ulcer, and gastric cancer. One of the most critical cofactors in H. pylori infection is the host immune response. However, the underlying mechanism involved remains poorly understood. Furthermore, the exact route of transmission of H. pylori is still unclear and not well documented. Contaminated food caused by cockroaches may contribute to the transmission of H. pylori infection. Therefore, the objectives of this study are to investigate the pathogenesis of H. pylori infection occurring in human and cockroaches, the variation of the virulence factor between strains isolated from human and cockroaches and to elucidate immune response in human gastric cancer (AGS) and mice model. In this study, a total of one hundred gastric biopsies from patients undergoing endoscopy at Universiti Teknologi MARA (UiTM) Sungai Buloh and cockroaches around the eateries area in Sungai Buloh were collected. The bacterial isolates were subjected to bacteria identification and followed by genotyping using polymerase chain reaction (PCR). Differential expression of virulence genes among both this group of strains was investigated by inoculating the isolated strains into AGS cells. The expression was then measured by using quantitative real-time PCR (qRT-PCR). Simultaneously, the expression of host immune response upon infection with different H. pylori strains was conducted using RT2 Profiler PCR array. In the in vivo study, mice comprising of six groups, including a control group of uninfected mice were used. The experimental groups were orogastrically infected with different H. pylori strains on days 0, 3, and 5, respectively. The mice were euthanised, and stomachs were harvested after post-infection for microbiology, histology, and molecular analysis. The expression of inflammatory cytokines was conducted using quantitative real-time PCR (qRT-PCR). A total of five H. pylori isolates (H. pylori S3, S5, S7, S33, S38)from gastric biopsies and ten H. pylori isolates (H. pylori C3, C6, C7, C8, C12, C23, C32, C33, C38, C40) from cockroaches were obtained in study. The cagA gene presence in 80% of the isolates. The expression of sabA, babA, ureA, and flaA gene was elevated and showed the highest expression in H. pylori cagA+ strain after interaction with the host cells. Nine gene encoding toll-like receptors (TLRs) were significantly elevated in H. pylori-infected cells, especially the TLR9 gene that showed the highest expression in the infected cells with H. pylori cagA+ strain. The regulation of TLRs activates the signalling of NFκβ gene expression. Moreover, H. pylori-infected cells regulate strong Th1 and Th17 immune responses. Th cells secrete higher expression of pro-inflammatory cytokines, including IL1β, IL6, IL8, IL12, IL17α, IL18, IL23, and TNFα in the infected cells. Lower expression of the anti-inflammatory cytokine, IL10 in the infected cells downregulates the JAK-STAT signaling pathway. The level of pro-inflammatory cytokines also elevated in the H. pylori-infected mice. The comprehensive data on the differentially expressed gene was successfully generated, and the host immune response involved in the diverse clinical outcome of H. pylori infection. The comparative study has also revealed that H. pylori strains isolated from humans and cockroaches demonstrated a comparable immune response gene expression profiling. This data could provide new insight into the pathogenesis of H. pylori infection in humans and establish the potential role of cockroaches in the transmission of H. pylori infection.