Brucellosis is a widespread zoonotic disease primarily transmitted by consumption of unpasteurized dairy products derived from cow's or sheep's and goat's milk contaminated with Brucella abortus or Brucella melitensis. Both Brucella species are viable in milk for several days, yet, the impact of milk on the metabolic activity and propagation of Brucella is unknown.

Whole genome sequence analyses of B. abortus identified a panel of proteases, peptidases and peptide transporters that may contribute to the degradation and utilization of milk proteins. We demonstrated that B. abortus uses milk efficiently as a substrate for growth: It proliferates in commercial cow's milk with an increase in colony counts of several magnitudes over 72 hours. To examine the proteolytic activity of B. abortus, the peptide changes of commercial cow's milk upon cultivation with B. abortus were examined by LC-MS analysis.

Since casein proteins are the major components of milk, we investigated the impact of casein, its derived peptides and free amino acids on the growth of B. abortus. Bacteria were cultivated in minimal medium supplemented with casein, tryptic-digested casein or casein hydrolysate as sole energy sources. While, casein peptides and free amino acids from casein hydrolysate promoted Brucella growth, uncleaved casein could not be catabolized by B. abortus.

We hypothesized that milk bacteria produce the casein-derived products utilized by Brucella during their growth in milk. We therefore cultivated bacteria from fresh, unpasteurized cow's milk and identified isolates by MALDI-TOF MS and NGS analyses. Among the bacterial species identified were Staphylococcus equorum and S. sciuri. The latter isolate exhibited pronounced proteolytic activity when grown on milk agar plates. We incubated the milk bacteria in minimal medium containing casein to produce spent media. B. abortus growth was promoted in the spent medium derived from the proteolytically active S. sciuri. This observation suggests that B. abortus takes advantage of the casein-derived amino acids and peptides as well as other metabolic products being generated through the catabolic activity of S. sciuri in cow's milk.