Investigating biofilm formation, enzyme activity and sporulation in varied total dissolved solids of bacillus licheniformis : a thesis presented in partial fulfilment of the requirements for the degree of Master of Food Technology at Massey University, Manawatu, New Zealand
This study investigated the sporulation, biofilm formation and enzymatic activity of isolates of Bacillus licheniformis sourced from dairy environments including butter, milk protein concentrate (MPC) and milk treatment skim module (MTSM) and from farm. Sporulation analysis with a two way ANOVA has shown significant effects of both time and TDS level on sporulation across all samples (p < 0.05). All isolates tend to show higher spore counts(especially for isolates sourced from factory) with increased TDS level. This suggests potential disruption in metabolic pathways with increased ion concentrations and prompting sporulation, even in nutrient rich environments. Biofilm was assessed at temperatures ranging from 20℃ to 60℃ using crystal violet assay. One way ANOVA indicated the temperature has a significant (p < 0.05) effect on biofilm forming for all isolates. Most isolates showed an optimal growth temperature between 30℃ to 50℃ with minimal growth at 20℃ and 60℃. Butter isolates peaked at 30℃ and 37℃, MPC isolates peaked at 50℃ which reflected potential adaptation to specific processing environments from their source. Enzymatic activity screening has shown protease activity at 37℃ after 40 hours incubation and 30℃ after 72 hours incubation. Heat sensitivity tests showed no detectable result suggesting that the enzymes were either not released to the media or concentrations were too low for detection. The findings highlight the variety of adaptive mechanisms of Bacillus licheniformis to environmental factors. Understanding these responses is essential for developing strategies to control the bacterial contamination in food products and processing. Future research is recommended to use more sensitive detection methods such as flow cytometry for spore counts, viable cell enumeration method (CFU/ml assay) for biofilm and azocasein assay for protease activity to improve accuracy.
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