CONICET | Buscador de Institutos y Recursos Humanos

Helicobacter pylori BIOFILM FORMATION AND EXPRESSION OF luxS and omp18 GENES Biofilms are communities of bacteria able to survive in unfavorable conditions where they often choose a sessile biofilm lifestyle as a strategy to overcome environmental stress. The expression of luxS gene represents an indicator of biofilm production, in which bacteria migrate and adhere to surfaces forming micro-colonies. H. pylori have the capacity to attach to different abiotic surfaces and the production of biofilm may be an important step for growth and survival during transmission. The intercellular communication, quorum sensing, is encoded in H. pylori by the luxS gene that is essential for colonization of the human stomach. Bacterial outer membrane proteins are important for ion transport, bacterial virulence and adherence. In H. pylori omp18 is a peptidoglycan-associated lipoprotein precursor that is involved in the adherence to gastric cells. Therefore, it is possible that the luxS and omp18 genes can be involved in biofilm formation. We analyzed the luxS and omp18 gene expression in H. pylori during biofilm formation in glass and polystyrene surfaces. H. pylori, NCTC11638 reference strain and HP796 clarithromicyn and metronidazole resistant strain, were grown in Mueller-Hinton broth supplemented with: a) 5% fetal calf serum (reference) and b) 0.5% alternative supplement of microbial origin. The cultures were incubated under microaerophylic conditions for 196 h at 37ºC. The viability of adhered H. pylori bacteria to abiotic surfaces was determined by plate counting and fluorescence microscopy using the LIVE/DEAD BacLight viability kit. The luxS and omp18 genes together with the 16SRNA, as housekeeping gene, were analyzed. For RNA extraction, the confluent growth on the two abiotic surfaces was treated with TRIzol reagent. The results showed that both strains have the capacity of biofilm formation in the surfaces assayed. The resistant strain showed a higher ability of biofilm formation (p≤0.005), maintaining the viability for 196 h. In relation to bacteria viability, differences of 2 to 3 logarithmic units were detected in the total and viable counts (p≤0.05) at 196h, regardless of nutritional conditions. Morphological changes were observed by fluorescence microscopy, spiral to cocoid forms and live and dead bacteria were clearly scored in samples processed at 96 h and 196 h. Both strains showed a higher expression of luxS and omp18 genes in bacteria present in the biofilm. The study found no statistical differences in gene expression for conditions assayed. The increase of luxS and omp18 gene expression and the changes in morphology and viability observed during biofilm formation in both surfaces and nutritional conditions, could indicate that H. pylori prefers a community-based surface-bound lifestyle, regulating gene expression to optimize the survival in hostile environments.