Naturally Fermented Milk From Northern Senegal: Bacterial Community Composition and Probiotic Enrichment With Lactobacillus rhamnosus

Abstract

A variety of foods fermented with lactic acid bacteria serve as dietary staples in many African communities; yet, their bacterial profiles are poorly characterized. The integration of health-promoting probiotics into naturally fermented milk products could make a profound impact on human health. Here, we characterize the bacterial community composition of a naturally fermented milk product (lait caillé) from northern Senegal, prepared in wooden bowls (lahals) with a bacterial biofilm to steer the fermentation process. We incorporated a probiotic starter culture containing the most documented probiotic strain Lactobacillus rhamnosus GG (generic strain name yoba 2012) into the local fermentation process. Bar-coded 16S rRNA amplicon sequencing of lait caillé samples indicated that the bacterial community of lait caillé has high species richness with over 100 bacterial genera; however, few have high abundance. In contrast to the diverse bacterial compositions of other characterized naturally fermented milk products, the composition of lait caillé predominantly consists of the lactic acid bacteria Streptococcus and Lactobacillus, resembling the bacterial composition in regular yogurt. The bacterial community composition of lait caillé varies geographically based on the presence of some genera, including Lactoccoccus, Enterococcus, Bifidobacterium, and Bacillus, but this trend is not consistent within production communities. The diversity of bacterial communities is much higher in the lahal biofilm than in the naturally fermented milk products, which is in turn greater than in commercial yogurts. Addition of a starter culture with L. rhamnosus yoba 2012 to milk in lahals led to substantial growth of this probiotic bacterium during the fermentation process. Two independent quantitative PCR-analyses specific for L. rhamnosus yoba 2012 indicated a 20- to 60-fold increase in the total number of probiotic bacteria in the first batch after inoculation. A similar increase of the probiotic was observed in a variation of lait caillé prepared with carbohydrate-rich millet granules (thiakry) added prior to fermentation. This study shows the feasibility of integrating health-promoting probiotic strains into naturally fermented foods produced in regions with a high prevalence of malnutrition.

Keywords: Lactobacillus rhamnosus GG; Lactobacillus rhamnosus yoba 2012; bacterial community; lait caillé; naturally fermented milk; probiotics.

FIGURE 1

Geographic position of project sites in northern Senegal. The sites were close to the town of Richard Toll, mostly along the south bank of the river Senegal marking the border between Senegal and Mauritania. The sites include the communities of Guilado, Medina Cheikh Mountaga, Didjiery, Pathé Badio, Keur Mbaye Peuhl, and Gaé. From each of these sites, samples of lait caillé were collected from wooden bowls (lahals) from households, except for the town of Gaé, where lait caillé samples were collected from plastic buckets in boutiques.

FIGURE 2

Stepwise production process of traditional fermented milk in northern Senegal. (A) Production process of lait caillé with cow’s milk in a lahal as practiced in households in Didjiery, Keur Mbaye Peuhl, Pathé Badio, Guilado, and Medina Cheikh Mountaga: (1) collection of milk from a cow; (2) filtration of the milk; (3) pasteurization of the milk by heating on firewood; (4) fermentation of the milk in a lahal for at least 12 h; (5) removal of fat and homogenization of the fermented milk with a wooden burgal. (B) lait caillé production process with milk powder in a plastic bucket in boutiques of the Gaé community: (1) milk powder is transferred to a plastic bucket; (2) water is added to the milk powder; (3) heated water is added to further dilute and warm the milk; (4) the milk is mixed with a small volume of lait caillé to start the fermentation (backslopping); (5) the bucket is covered with a towel and the milk is fermented for a period of 6–7 h. Written informed consent was obtained from the member of the Gaé community for the publication of these images.

FIGURE 3

Heat map of the average top 50 bacterial genera for each community. The heat map is based on normalized 16S rRNA V4 amplicon sequence data obtained from bacterial DNA isolated from 120 lait caillé samples from six communities in northern Senegal. Samples were collected from plastic buckets in boutiques from Gaé (n = 26), and from lahals in households from Didjiery (n = 10), Keur Mbaye Peuhl (n = 19), Pathé Badio (n = 19), Guilado (n = 23), and Medina Cheikh Mountaga (n = 22). The number of observations for each bacterial genus was plotted in a rainbow scale from black to blue to green to red. Saturation was set at a value of 60 (1% of the maximum).

FIGURE 4

The Streptococcus-Lactobacillus dominance. Plot of the fractions of Lactobacillus vs. the fractions of Streptococcus in the lait caillé samples. The six communities are indicated by different color and symbol coding. The dotted line indicates where f Lactobacillus +f Streptococcus = 100%. The size of the symbols indicates the fraction of Lactococcus.

FIGURE 5

Fraction of specific genera in lait caillé samples per community. Box plots for genera per production community. Only genera with a q < 5% are shown (see Table 1). (A) fraction of Lactococcus per community; (B) fraction of Enterococcus per community; (C) fraction of Bacillus per community; (D) fraction of Bifidobacterium per community.

FIGURE 6

Average microbiota composition of lahal biofilm and fermented milk. The pie charts indicate the predominant genera (>0.5%) of household lahal biofilms (10 samples), household lahal lait caillé (93 samples from 5 communities), the boutique bucket lait caillé (26 samples from Gaé), and commercial yogurts (6 samples).