Bovine Teat Cistern Microbiota Composition and Richness Are Associated With the Immune and Microbial Responses During Transition to Once-Daily Milking

Abstract

The relationship between microbiota and health has been widely reported in humans and animals. We established a link between teat cistern microbiota composition and bovine mastitis, an inflammatory disease often due to bacterial infections. To further decipher the relationships between teat cistern microbiota and immune and microbial responses, a switch from twice- to once-daily milking (ODM) in 31 initially healthy quarters of dairy cows was used to trigger an udder perturbation. In this study, a temporal relationship was reported between initial teat cistern microbiota composition and richness, the immune response to ODM, and mastitis development. Quarters with a low initial microbiota richness and taxonomic markers such as Bacteroidetes and Proteobacteria were associated with a higher rate of mastitis during ODM. Quarters with a higher richness and taxonomic markers such as Firmicutes, including the Lachnospiraceae family, and genera such as Bifidobacterium and Corynebacterium displayed early inflammation following transition to ODM but without developing mastitis (no infection). Short-term compositional shifts of microbiota indicates that microbiotas with a higher initial richness were more strongly altered by transition to ODM, with notably the disappearance of rare OTUs. Microbiota modifications were associated with an early innate immune system stimulation, which, in turn, may have contributed to the prevention of mastitis development.

Keywords: inflammation; mastitis; milk microbiota; milking frequency; rare species; teat microbiota.

FIGURE 1

Multi-Dimensional Scaling (MDS) and hierarchical clustering on bovine teat cistern microbiota at day 0. (A) MDS was performed based on the measurement of the UniFrac distance. Samples are indicated by points and colored with regard to groups G1 to G5. Centroid positions are indicated for each group. (B) Hierarchical clustering of samples based on the UniFrac distance using the ward linkage function. Samples are distributed into two main clusters, C1 and C2, containing 13 and 15 quarters, respectively. Cluster C1 mainly includes quarters of groups G1 and G2, and cluster C2 mainly includes quarters of groups G4 and G5. An additional cluster (C3) includes only three samples, two from group G2 and one from G3. Quarters of group G3 are distributed among the three clusters.

FIGURE 2

Alpha-diversity of teat cistern microbiota at day 0, day 3, and day 14 with regard to clusters, as illustrated by the distribution of several diversity indices: observed richness, Chao1, Shannon, Simpson, and Inverse Simpson indices.

FIGURE 3

Taxonomic representation of differentially abundant taxa between C1 (contains mainly G1 and G2 quarters) and C2 (contains mainly G4 and G5 quarters) at day 0 for Firmicutes Actinobacteria Bacteroidetes and Proteobacteria as determined by the LEfSe pipeline. Differences are represented by the color of the cluster where the taxon is more abundant (red indicates C1 and green indicates C2). See Supplementary Table S3 for a complete list of discriminant taxa.

FIGURE 4

Temporal relationship between bovine teat cistern microbiota composition and richness and the immune and microbial responses during transition to once-daily milking. Microbiota of quarters were distributed into two main clusters. Cluster C1 included microbiota with low initial richness (day 0) exerting a low colonization resistance to pathogens. C1 microbiota were hardly affected by ODM-induced perturbation (resistant microbiota), which may have resulted in a lack of immune system stimulation (green spot). The higher rate of pathogen entrance through the teat canal thus resulted in a higher rate of mastitis cases in this cluster. Cluster C2 included microbiota with higher initial richness at day 0 that may have contributed to a higher colonization resistance. These C2 microbiota were more strongly altered in response to the transition to ODM (sensitive microbiota). Changes in microbial composition and the decrease in richness may have contributed to the stimulation of the innate immune system (red spot). This might have contributed to combatting pathogens entering the mammary gland, thus preventing the development of infection.