FoodMicro 2026

Abstract

Milk powder contamination can occur from soil, bedding, milk equipment, or transport from the farm to the processing plant. At the dairy production site, mesophilic and thermophilic bacteria survive heat treatments and can form biofilm communities in factory equipment. This can provide a source of contamination for dairy powders. It has been reported that these microorganisms can convert nitrate to nitrite, thereby a􀆯ecting nitrite levels in final products. Thus, it is important to control these microorganisms throughout the production process to prevent spoilage and ensure product quality.

This study explores the nitrate conversion capabilities and biofilm formation of different mesophilic and thermophilic species isolated from a dairy powder site. The genomes are
sequenced to identify key genomic differences among the different bacterial species and isolates to explain phenotypic behavior.

In this work, 155 strains were isolated from different locations (n=4) at the dairy powder site during multiple sampling campaigns (n=3). Samples were obtained from the milk evaporator, the concentrate after the pre-heater and the final powder samples. The strains were identified at the species level using MALDI-TOF MS. The biofilm-forming capacity of relevant strains was evaluated by crystal violet staining, and nitrate conversion was assessed using the Griess reagent test. The genomes of 58 representative strains, selected based on phenotypic diversity, were sequenced.

Bacillus licheniformis was the most isolated microorganism in the evaporator, while in the concentrate, Geobacillus stearothermophilus was the most prevalent. In powder samples, both Geobacillus stearothermophilus and Anoxybacillus flavithermus were dominant. Among the sequenced strains, 44.8% showed complete nitrate reduction, 46.6% showed strong reduction, and 8.6% showed poor conversion to nitrite. Furthermore, differences in biofilm-forming capabilities are apparent among species and strains, with many Bacillus licheniformis strains being better biofilm producers. Five of these Bacillus licheniformis strains isolated from all locations showed high nitrate conversion and biofilm-forming capabilities. Both biofilm and nitrate conversion capabilities seem to be strain-dependent and independent of the isolation source.

Whole-genome sequencing analysis and phenotypic characterisation of spoilers are key to preventing spoilage at the dairy powder factory, enabling control of the dairy production line and
powder quality.