Specific Signatures of the Gut Microbiota and Increased Levels of Butyrate in Children Treated with Fermented Cow's Milk Containing Heat-Killed Lactobacillus paracasei CBA L74

Abstract

We recently demonstrated that cow's milk fermented with the probiotic Lactobacillus paracasei CBA L74 (FM-CBAL74) reduces the incidence of respiratory and gastrointestinal tract infections in young children attending school. This effect apparently derives from a complex regulation of non-immune and immune protective mechanisms. We investigated whether FM-CBAL74 could regulate gut microbiota composition and butyrate production. We randomly selected 20 healthy children (12 to 48 months) from the previous randomized controlled trial, before (t0) and after 3 months (t3) of dietary treatment with FM-CBAL74 (FM) or placebo (PL). Fecal microbiota was profiled using 16S rRNA gene amplicon sequencing, and the fecal butyrate concentration was also measured. Microbial alpha and beta diversities were not significantly different between groups prior to treatment. FM-CBAL74 but not PL treatment increased the relative abundance of Lactobacillus Individual Blautia, Roseburia, and Faecalibacterium oligotypes were associated with FM-CBAL74 treatment and demonstrated correlative associations with immune biomarkers. Accordingly, PICRUSt analysis predicted an increase in the proportion of genes involved in butyrate production pathways, consistent with an increase in fecal butyrate observed only in the FM group. Dietary supplementation with FM-CBAL74 induces specific signatures in gut microbiota composition and stimulates butyrate production. These effects are associated with changes in innate and acquired immunity.IMPORTANCE The use of a fermented milk product containing the heat-killed probiotic strain Lactobacillus paracasei CBAL74 induces changes in the gut microbiota, promoting the development of butyrate producers. These changes in the gut microbiota composition correlate with increased levels of innate and acquired immunity biomarkers.

Keywords: fecal butyrate; gut microbiota; immune system.

FIG 1

Oscillospira and Faecalibacterium levels. Box plots show the abundance of Oscillospira and Faecalibacterium in the studied population at baseline (t0) and after 3 months of treatment (t3) with fermented milk (MILK) and placebo (PL). Boxes represent the interquartile range (IQR) between the first and third quartiles, and the line inside represents the median (second quartile). Whiskers denote the lowest and highest values within 1.5 × IQR from the first and third quartiles, respectively. Asterisks indicate a significant difference as obtained by a pairwise Wilcoxon test (P < 0.05).

FIG 2

Lachnospiraceae and Ruminococcaceae abundance correlates with innate and acquired immunity. A heat plot shows the Spearman correlations between genera belonging to Lachnospiraceae and Ruminococcaceae and the levels of immunity biomarkers. Rows and columns are clustered by Euclidean distance and Ward linkage hierarchical clustering. The intensity of the colors represents the degree of association, as measured by Spearman correlations. Only genera occurring in at least 20% of the samples were included. Asterisks indicate a significant correlation after Benjamini-Hochberg correction.

FIG 3

Fecal butyrate concentration. Box plots show the abundance of fecal butyrate in the study population at baseline (t0) and after 3 months of treatment (t3) with fermented milk (MILK) and placebo (PL). Asterisks indicate a significant difference. as obtained by pairwise Wilcoxon test (P < 0.05). See the legend to Fig. 1 for a description of the box plots.