|Institution:||University of Helsinki|
|Full text PDF:||http://hdl.handle.net/10138/153504|
Lactobacillus rhamnosus GG is a lactic acid bacterium that is widely used as probiotic products in the dairy industry. To gain insights into the genome stability of the L. rhamnosus GG in the human gastrointestinal tract and the possible adaption mechanism under different stresses, we first examined the genotype and phenotype of the L. rhamnosus GG grown over 1000 generations under various stresses, including bile salts, osmotic stress or shearing forces. Immunoblotting analysis of L. rhamnosus grown over 1000 generations showed that the production of mucus-binding pili by L. rhamnosus GG was the most impaired when exposed to bile salts. Complementary PCR screening of 13 highly variable chromosomal regions in GG confirmed that the pilus gene cluster had been lost when exposed to bile stress over time. In vitro bile-induced genomic changes observed in GG possibly reflects the genome plasticity and stability of GG in the human intestinal tract. Still, we showed that these changes only occurred after more than 100 generations, a period of time relatively long compared to the observed time of persistence and transit of GG in the intestinal tract. Although damages and stresses may be caused by bile salts, L. rhamnosus GG still has the ability to tolerate bile salts. The resistance mechanism is still unclear but, based on previous studies, we identified one ABC transporter encoded by the gene operon called tauABC that may be involved in bile resistance. In an effort to demonstrate its function, a tauB-null mutant derivative was generated and phenotypically characterized in terms of metabolic, signaling and functional properties. The data revealed that the tauB-null mutant significantly grow slower than L. rhamnosus GG wild-type strain in the presence of ox bile extracts. Additional screenings using various bile conjugates specifically revealed that two compounds of bile salts, i.e. taurodeoxycholic and taurochenodeoxycholic acid, may be processed by the TauABC transporter, contributing at least partially to the tolerance of GG to bile salts. Overall, we showed that bile salts constitute an important stress factor for GG that causes genomic alterations, although it has bile tolerance mechanisms to bile, such as the newly-characterized tauABC operon.