The global population stru cture of Lacticaseibacillus rhamnosus and its application to an investigation of a rare case of infective endocarditis

Abstract

Background: Lacticaseibacillus (formerly Lactobacillus) rhamnosus is widely used in probiotics or food supplements to promote microbiome health and may also be part of the normal microbiota of the human gastrointestinal tract. However, it rarely also causes invasive or severe infections in patients. It has been postulated that these infections may originate from probiotics or from endogenous commensal reservoirs. In this report, we examine the population structure of Lacticaseibacillus rhamnosus and investigate the utility of using bacterial genomics to identify the source of invasive Lacticaseibacillus infections.

Methods: Core genome phylogenetic analysis was performed on 602 L. rhamnosus genome sequences from the National Center for Biotechnology public database. This information was then used along with newly generated sequences of L. rhamnosus isolates from yogurt to investigate a fatal case of L. rhamnosus endocarditis.

Results: Phylogenetic analysis demonstrated substantial genetic overlap of L. rhamnosus isolates cultured from food, probiotics, infected patients, and colonized individuals. This was applied to a patient who had both consumed yogurt and developed L. rhamnosus endocarditis to attempt to identify the source of his infection. The sequence of the isolate from the patient's bloodstream differed at only one nucleotide position from one of the yogurt isolates. Both isolates belonged to a clade, identified here as clade YC, composed of mostly gastrointestinal isolates from healthy individuals, some of which also differed by only a single nucleotide change from the patient's isolate.

Conclusions: As illustrated by this case, whole genome sequencing may be insufficient to reliably determine the source of invasive infections caused by L. rhamnosus.

Fig 1. Whole genome phylogenetic analysis of Lacticaseibacillus rhamnosus.

A) Maximum likelihood phylogenetic tree, midpoint-rooted, from whole genome alignment of L. rhamnosus assemblies and sequence reads from the NCBI database (n = 602) to the reference genome sequence of strain ATCC 11443 (orange star). Taxa highlighted with colored squares indicate major clades consisting of 20 or more sequences. Sequences of the case patient’s bloodstream isolate (“Patient”–red star) and representative yogurt isolates (“Yogurt A, C, D”–green, blue, purple stars) are also included. B) Maximum likelihood phylogenetic tree of clade YC isolates, including case patient’s bloodstream isolate and yogurt C isolates. Turquoise stars indicate “Hansen 1968” isolate sequences. In both panels, rings from inside to outside indicate, in order: 1. isolation source, 2. body site (compartment) of isolation for human sourced isolates, 3. continent of isolation, and 4. year of isolation. Taxon labels in bold indicate sequences for which only genome assemblies were available. Scale bars indicate genetic distance.

Fig 2. Demographic characteristics of Lacticaseibacillus rhamnosus sequences among major phylogenetic clades.

Distributions of isolation source (A), isolation site or compartment for human-derived isolates (B), continent of isolation (C), and year of isolation (D) among isolates belonging to each of the five major clades and among the complete sequence collection (“All”). Bar heights indicate the percent of the total sequences in each group.

Fig 3. Maximum likelihood whole-genome phylogenetic tree of Lacticaseibacillus rhamnosus sequences from a patient with bloodstream infection and sampled yogurt brands.

Sequencing reads were aligned to the genome sequence of strain ATCC 11443 and the maximum likelihood phylogenetic tree was determined from the whole genome sequence alignment. For yogurt isolates, letters (A, C, D) indicate separate yogurt brands sampled and numbers (1–6) indicate separate colonies cultured from each yogurt brand. Scale bar indicates genetic distance.