Purpose: The goal of this study was to enumerate and identify bacterial spores in bulk tank raw milk.
Methods: Bulk tank raw milk samples and corresponding surveys examining farm management practices (n = 198) were collected from 33 farms, sampled every other month for 1 year. Raw milk samples were spore pasteurized (80°C for 12 min) to enumerate psychrotolerant, mesophilic, and thermophilic spore-forming bacteria, and PCR amplification of the rpoB gene was used for isolate identification. Bacillus cereus Group isolates were screened for i) the presence of toxin encoding genes, and ii) for hemolysis on sheep’s blood agar.
Results: Overall, psychrotolerant, mesophilic, and thermophilic spores were detected (> 10 spores/ml) in 1% (2/198), 74% (147/198) and 58% (115/198) of samples, respectively. Different management practices were associated with mesophilic and thermophilic spore levels, with the exception of herd size. Bacillus licheniformis was the most frequently isolated organism, representing 48% (313/654) of all isolates. Genes encoding the non-hemolytic enterotoxin and enteric FM toxins were present in all B. cereus Group allelic types (n = 10), while genes encoding the hemolysin BL toxin were absent for 5 out of 10 of the B. cereus Group allelic types, as determined by PCR screening. All representative B. cereus Group strains, except one, were hemolytic on sheep’s blood agar.
Significance: The results presented here provide relevant baseline spore counts in raw milk, and associations between management practices and spore counts that could be used to reduce spore contamination of raw milk for enhanced quality and safety.