title: Polymorphisms, Chromosomal Rearrangements, and Mutator Phenotype Development during Experimental Evolution of Lactobacillus rhamnosus GG.
abstract: Lactobacillus rhamnosus GG is a lactic acid bacterium widely marketed by the food industry. Its genomic analysis led to the identification of a gene cluster encoding mucus-binding SpaCBA pili, which is located in a genomic island enriched in insertion sequence (IS) elements. In the present study, we analyzed by genome-wide resequencing the genomic integrity of L. rhamnosus GG in four distinct evolutionary experiments conducted for approximately 1,000 generations under conditions of no stress or salt, bile, and repetitive-shearing stress. Under both stress-free and salt-induced stress conditions, the GG population (excluding the mutator lineage in the stress-free series [see below]) accumulated only a few single nucleotide polymorphisms (SNPs) and no frequent chromosomal rearrangements. In contrast, in the presence of bile salts or repetitive shearing stress, some IS elements were found to be activated, resulting in the deletion of large chromosomal segments that include the spaCBA-srtC1 pilus gene cluster. Remarkably, a high number of SNPs were found in three strains obtained after 900 generations of stress-free growth. Detailed analysis showed that these three strains derived from a founder mutant with an altered DNA polymerase subunit that resulted in a mutator phenotype. The present work confirms the stability of the pilus production phenotype in L. rhamnosus GG under stress-free conditions, highlights the possible evolutionary scenarios that may occur when this probiotic strain is extensively cultured, and identifies external factors that affect the chromosomal integrity of GG. The results provide mechanistic insights into the stability of GG in regard to its extensive use in probiotic and other functional food products.
  
  
  
  Lactobacillus rhamnosus GG is a widely marketed probiotic strain that has been used in numerous clinical studies to assess its health-promoting properties. Hence, the stability of the probiotic functions of L. rhamnosus GG is of importance, and here we studied the impact of external stresses on the genomic integrity of L. rhamnosus GG. We studied three different stresses that are relevant for understanding its robustness and integrity under both ex vivo conditions, i.e., industrial manufacturing conditions, and in vivo conditions, i.e., intestinal tract-associated stress. Overall, our findings contribute to predicting the genomic stability of L. rhamnosus GG and its ecological performance.
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: François P
authors.initials: FP
authors.lastname: Douillard
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland Laboratory of Microbiology, Wageningen University, Wageningen, the Netherlands.
authors.forename: Angela
authors.initials: A
authors.lastname: Ribbera
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: Kun
authors.initials: K
authors.lastname: Xiao
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: Jarmo
authors.initials: J
authors.lastname: Ritari
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: Pia
authors.initials: P
authors.lastname: Rasinkangas
authors.affiliation: Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
authors.forename: Lars
authors.initials: L
authors.lastname: Paulin
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: Airi
authors.initials: A
authors.lastname: Palva
authors.affiliation: Laboratory of Microbiology, Wageningen University, Wageningen, the Netherlands Key Laboratory of Functional Dairy, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China.
authors.forename: Yanling
authors.initials: Y
authors.lastname: Hao
authors.affiliation: Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland Laboratory of Microbiology, Wageningen University, Wageningen, the Netherlands RPU Immunobiology, Department of Bacteriology and Immunology, Haartman Institute, University of Helsinki, Helsinki, Finland willem.devos@wur.nl.
authors.forename: Willem M
authors.initials: WM
authors.lastname: de Vos
mesh_terms: D004251:DNA Transposable Elements; D015321:Gene Rearrangement; D042822:Genomic Instability; D052201:Lacticaseibacillus rhamnosus / Q000235:genetics* / Q000254:growth & development*; D009154:Mutation; D010641:Phenotype; D011110:Polymorphism, Genetic; D019936:Probiotics; D011995:Recombination, Genetic
chemical_list: D004251:DNA Transposable Elements