Genomic Stability and Phenotypic Characteristics of Industrially Produced Lacticaseibacillus rhamnosus GG in a Yogurt Matrix.

Lacticaseibacillus rhamnosus GG is a widely marketed probiotic with well-documented probiotic properties. Previously, deletion of the mucus-adhesive spaCBA-srtC1 genes in dairy isolates was reported. In this study, we examined the genome preservation of industrially produced L. rhamnosus GG (DSM 33156) cofermented in yogurts. In total, DNA of 66 samples, including 60 isolates, was sequenced. Population samples and 59 isolates exhibited an intact genome. One isolate exhibited loss of spaCBA-srtC1. In addition, we examined phenotypes related to the probiotic properties of L. rhamnosus GG either from frozen pellets or cofermented in yogurt. L. rhamnosus GG from frozen pellets induced a response in intestinal barrier function in vitro, in contrast to frozen pellets of the starter culture. Yogurt matrix, containing only the starter culture, induced a response, but cofermentation with L. rhamnosus GG induced a higher response. Conversely, only the starter culture stimulated cytokine secretion in dendritic cells, and it was observed that the addition of L. rhamnosus GG to the starter culture reduced the response. We conclude that the L. rhamnosus GG genome is preserved in yogurt and that common in vitro probiotic effects of L. rhamnosus GG are observed when examined in the yogurt matrix. IMPORTANCELacticaseibacillus rhamnosus GG is a well-documented probiotic strain recognized for its high acid and bile tolerance and properties of adhesion to enterocytes and mucus. The strain exhibits SpaCBA pili, which have been demonstrated to play an important role in adhesion and therefore are relevant for persistence in the gastrointestinal tract. Recently we demonstrated that the genome and phenotypes of L. rhamnosus GG are preserved throughout an industrial production pipeline. However, as gene deletions in L. rhamnosus GG were previously reported for isolates from dairy products, a key question on the genomic stability of L. rhamnosus GG in a yogurt matrix remained. The aim of this study was to analyze genome stability and phenotypic characteristics of L. rhamnosus GG in yogurt. We found that the genome of L. rhamnosus GG is well conserved when the organism is cofermented in yogurt. Some phenotypic characteristics are consistent in all product matrixes, while other characteristics are modulated.

Lactobacillus rhamnosus GG Genomic and Phenotypic Stability in an Industrial Production Process.

Lactobacillus rhamnosus GG is one of the most widely marketed and studied probiotic strains. In L. rhamnosus GG, the spaCBA-srtC1 gene cluster encodes pili, which are important for some of the probiotic properties of the strain. A previous study showed that the DNA sequence of the spaCBA-srtC1 gene cluster was not present in some L. rhamnosus GG variants isolated from liquid dairy products. To examine the stability of the L. rhamnosus GG genome in an industrial production process, we sequenced the genome of samples of L. rhamnosus GG (DSM 33156) collected at specific steps of the industrial production process, including the culture collection stock, intermediate fermentations, and final freeze-dried products. We found that the L. rhamnosus GG genome sequence was unchanged throughout the production process. Consequently, the spaCBA-srtC1 gene locus was intact and fully conserved in all 31 samples examined. In addition, different production batches of L. rhamnosus GG exhibited consistent phenotypes, including the presence of pili in final freeze-dried products, and consistent characteristics in in vitro assays of probiotic properties. Our data show that L. rhamnosus GG is highly stable in this industrial production process.IMPORTANCELactobacillus rhamnosus GG is one of the best-studied probiotic strains. One of the well-characterized features of the strain is the pili encoded by the spaCBA-srtC1 gene cluster. These pili are involved in persistence in the gastrointestinal tract and are important for the probiotic properties of L. rhamnosus GG. Previous studies demonstrated that the L. rhamnosus GG genome can be unstable under certain conditions and can lose the spaCBA-srtC1 gene cluster. Since in vitro studies have shown that the loss of the spaCBA-srtC1 gene cluster decreases certain L. rhamnosus GG probiotic properties, we assessed both the genomic stability and phenotypic properties of L. rhamnosus GG throughout an industrial production process. We found that neither genomic nor phenotypic changes occurred in the samples. Therefore, we demonstrate that L. rhamnosus GG retains the spaCBA-srtC1 cluster and exhibits excellent genomic and phenotypic stability in the specific industrial process examined here.