Prebiotic utilisation provides Lactiplantibacillus plantarum a competitive advantage in vitro, but is not reflected by an increased intestinal fitness

Abstract

Synbiotics combine the concepts of probiotics and prebiotics to synergistically enhance the health-associated effects of both components. Previously, we have shown that the intestinal persistence of inulin-utilizing L. plantarum Lp900 is significantly increased in rats fed an inulin-supplemented, high-calcium diet. Here we employed a competitive population dynamics approach to demonstrate that inulin and GOS can selectively enrich L. plantarum strains that utilize these substrates for growth during in vitro cultivation, but that such enrichment did not occur during intestinal transit in rats fed a GOS or inulin-supplemented diet. The intestinal persistence of all L. plantarum strains increased irrespective of their prebiotic utilization phenotype, which was dependent on the calcium level of the diet. Analysis of fecal microbiota and intestinal persistence decline rates indicated that prebiotic utilization capacity did not selectively stimulate intestinal persistence in prebiotic supplemented diets. Moreover, microbiota and organic acid profile analyses indicate that the prebiotic utilizing probiotic strains are vastly outcompeted by the endogenous prebiotic-utilizing microbiota, and that the collective enhanced persistence of all L. plantarum strains is most likely explained by their well-established tolerance to organic acids.

Keywords: Lactiplantibacillus plantarum; competition; galacto-oligosaccharides; intestinal persistence; inulin; prebiotics; probiotics.

Figure 1.

Time-resolved persistence and relative abundance of intragastrically administered L. plantarum-R strains in adult male rats. (a) L. plantarum-R strain panel legend (Figure 1(a)) with growth phenotypes. A plus (+) symbol indicates that capacity of a strain to utilise >DP3 inulin or DP3–4 GOS, while a minus (-) symbol indicates the lack of this capacity (see supplemental Figures S1–2).^, Strains with a (-) phenotype fail to utilise higher DP compounds but are still capable of utilising most mono- and di-saccharides present in either prebiotic substrate. (b) Persistence of administered L. plantarum-R strain cocktail in rats on Hca (circles, open), HcaGOS (black circles, closed) and HcaInu (grey circles, closed) diets after a single gavage with approximately 6 × 10⁹ CFU of L. plantarum-R on day 0. All CFU counts are log-transformed and expressed as mean ± SEM (n = 8 per diet group). #, the GOS-supplemented group is significantly different from the Hca control group (#: p < .05, ##: p < .01). *, the inulin-supplemented group is significantly different from the Hca control group control group (*: p < .05). (c) L. plantarum-R mixture was grown on ½ MRS-C supplemented with either 0.5% (w/v) GOS or inulin and passaged 1:500 after 24 hours of incubation for 8 days. Stacked area charts represent the relative abundance of L. plantarum strain-specific 339-IR-340 sequences amplified from isolated DNA from mixed bacterial cultures collected at different time points corresponding to estimated generations (see figure SF3 in the supplementary material). (d) Stacked area charts with relative abundance of L. plantarum strain-specific 339-IR-340 sequences amplified from isolated faecal DNA collected at different time points after L. plantarum-R gavage in rats on the different diets. Results are expressed as average of relative abundance, as fraction of the whole.

Figure 2.

Exploring the relation between persistence, microbiota and fecal organic acid levels. RDA with the endogenous microbiota (genera) as response variables and the explanatory variable of diet: Hca (yellow squares), Lca (yellow circles), HcaGOS (blue squares), LcaGOS (blue circles) and HcaInu (red squares) with fecal organic lactic (LA), succinic (SA), acetic (AA), propionic (PA) and butyric (BA) acid levels, total acid levels and the persistence rate (i.e., inverse of linear decline rate) as supplementary variables. Blue arrows correspond to selected genera associated with the dietary regimes.

Figure 3.

Relative abundance of L. plantarum-R compared to endogenous Bifidobacterium, Faecalibaculum abundance post-gavage. Relative abundance of Bifidobacterium (green), Faecalibaculum (orange), and the L. plantarum-R cocktail (yellow) after (a) one day post-gavage and (b) three days post-gavage. Asterisks (*) indicate significance after a two-sided Mann-Whitney t test. *: p < .05.