Experimental Evaluation of Host Adaptation of Lactobacillus reuteri to Different Vertebrate Species

Abstract

The species has diversified into host-specific lineages, implying a long-term association with different vertebrates. Strains from rodent lineages show specific adaptations to mice, but the processes underlying the evolution of in other hosts remain unknown. We administered three standardized inocula composed of strains from different host-confined lineages to mice, pigs, chickens, and humans. The ecological performance of each strain in the gastrointestinal tract of each host was determined by typing random colonies recovered from fecal samples collected over five consecutive days postadministration. Results revealed that rodent strains were predominant in mice, confirming previous findings of host adaptation. In chickens, poultry strains of the lineage VI (poultry VI) and human isolates from the same lineage (human VI) were recovered at the highest and second highest rates, respectively. Interestingly, human VI strains were virtually undetected in human feces. These findings, together with ancestral state reconstructions, indicate poultry VI and human VI strains share an evolutionary history with chickens. Genomic analysis revealed that poultry VI strains possess a large and variable accessory genome, whereas human VI strains display low genetic diversity and possess genes encoding antibiotic resistance and capsular polysaccharide synthesis, which might have allowed temporal colonization of humans. Experiments in pigs and humans did not provide evidence of host adaptation of to these hosts. Overall, our findings demonstrate host adaptation of to rodents and chickens, supporting a joint evolution of this bacterial species with several vertebrate hosts, although questions remain about its natural history in humans and pigs. Gut microbes are often hypothesized to have coevolved with their vertebrate hosts. However, the evidence is sparse and the evolutionary mechanisms have not been identified. We developed and applied an experimental approach to determine host adaptation of to different hosts. Our findings confirmed adaptation to rodents and provided evidence of adaptation to poultry, suggesting that evolved via natural selection in different hosts. By complementing phylogenetic analyses with experimental evidence, this study provides novel information about the mechanisms driving host-microbe coevolution with vertebrates and serve as a basis to inform the application of as a probiotic for different host species.

Coadministration of the Campylobacter jejuni N-Glycan-Based Vaccine with Probiotics Improves Vaccine Performance in Broiler Chickens

“…To select a strain of L. reuteri, we performed an experiment to determine the strain with the highest ecological fitness among a list of poultry isolates. Lactobacillusfree SPF leghorn chickens were given an inoculum containing nine L. reuteri strains of poultry origin by gavage, as previously described (39). As shown in Fig.…”

“…Briefly, individual strains were grown in MRS medium (Difco) supplemented with 0.5% fructose and 1% maltose at 37°C for 16 h. Prior to gavage, cells from individual cultures were harvested by centrifugation (3,000 ϫ g for 10 min), washed twice with sterile PBS (pH 7.0), and mixed in equal numbers in the same buffer to generate the inocula. Lactobacillus-free SPF leghorn chickens, reared as described previously (39), received a 300-l PBS suspension containing 1 ϫ 10 6 cells of each L. reuteri strain by gavage. Cloacal swabs were taken from the chickens before gavage at day 0 and for 5 days after gavage and cultured by dilution plating on LRIM (39).…”

“…Nine L. reuteri strains were used for the in vivo probiotic selection experiment (Table S1). Strains were originally isolated from either chickens or turkeys and belonged to a phylogenetic lineage specific to poultry (39,80). Each of the strains carried a distinct leuS allele to allow differentiation via colony typing (Table S1).…”

“…Thus, L. reuteri probiotic strain CSF8 was also selected by using an experimental design validated for comparison of the ecological competitiveness of L. reuteri strains in chickens in vivo (39). The fact that L. reuteri CSF8 outcompeted all other L. reuteri strains that were also of poultry origin suggests that this strain has evolved specific adaptations to be competitive and metabolically active under the ecological conditions expected in the chicken intestine, and this strain was therefore selected as another probiotic strain for use in the vaccine experiment.…”

“…Each of the strains carried a distinct leuS allele to allow differentiation via colony typing (Table S1). In vivo competition experiments were conducted according to methods described previously by Duar et al (39), with some modifications. Briefly, individual strains were grown in MRS medium (Difco) supplemented with 0.5% fructose and 1% maltose at 37°C for 16 h. Prior to gavage, cells from individual cultures were harvested by centrifugation (3,000 ϫ g for 10 min), washed twice with sterile PBS (pH 7.0), and mixed in equal numbers in the same buffer to generate the inocula.…”