Determining the types of Bacillus endospores in whey protein concentrate and nonfat dried milk powders


S. Jha
S. Anand

Document Type


Publication Date



American Dairy Science Association


Journal of Dairy Science




Suppl. 1






endospore, mesophile, thermophile


Several spore-forming bacterial species are commonly encountered in dairy powders. These are known to produce thermally resistant endo- spores. Determining the types of endospores formed by these bacteria could thus be useful in developing control strategies to improve their microbial quality and further application in product development. In the current study, samples of whey protein concentrate (WPC) and nonfat dried milk powder (NFDM) were analyzed for the presence of different types of Bacillus endospores, based on their thermal activation treat- ments. Twenty-five samples of each of WPC and NFDM were procured from commercial dairy plants. The 11 g of each sample was reconstituted in 99mL of phosphate buffer saline. A 20 mL portion of the respective reconstituted sample was heated at 80°C/12min for regular spores (SP), 100°C/30min for high-heat resistant spores (HHRS), and 106°C/30min for specially thermoresistant spores (STS). After the heat treatment, the samples were cooled to room temperature, and the desired serial dilutions were plated on the tryptic soy agar. The plates were incubated for 24 h at 37°C for mesophiles, and 55°C for thermophiles. The experiments were performed in duplicates and the means were compared using ANOVA. Several spore-forming species were identified based on colony morphology followed by MALDI-TOF. In WPC the samples containing the mesophilic spores were found to be 64% SP, 20% HHRS, and 16% STS. The samples containing thermophilic spores were 73% SP, 14% HHRS, and 13% STS. Similarly, in NFDM the samples containing the mesophilic spores were 95% SP and 5% HHRS. The samples containing thermophilic spores were 60% SP and 40% HHRS. This distribution reveals the higher incidence of SP in the dairy powders that could play a vital role as the powders are intended for reconstitution before use. MALDI-TOF analysis revealed Bacillus licheniformis and Bacillus cereus as the predominant species in WPC and NFDM, respectively. Some other prominent Bacillus species identified were B. pumilus, B. subtilis, and Geobacillus stearothermophilus.

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