Enhancing the Biopreservative Effect of Non-Starter Lactic Acid Bacteria (NSLAB) by Soluble Fiber During Cheese Ripening

This thesis investigated the microbial and physicochemical changes during Cheddar cheese ripening, with a specific focus on the role of non-starter lactic acid bacteria (NSLAB) and the influence of soluble fiber (inulin) on microbial population and controlling spoilage due to spores. In the first phase (Chapter 3), cheese samples were produced using pasteurized standardized milk with four treatments: control, inoculated with aerobic spores of Bacillus licheniformis T1 (BL), inoculated with aerobic spores of Clostridium tyrobutyricum T2 (CT) spore-formers, and a mixture of both, T3 (BL+CT). Over a six-month ripening period, NSLAB populations, particularly Lactobacillus rhamnosus and Lactobacillus paracasei, increased steadily, reaching 8.5 Log₁₀ CFU/g after 3 months and remained above 8.5 Log₁₀ CFU/g through month six across all treatments, indicating that spore inoculation did not impact NSLAB proliferation. Spore counts in spore-inoculated cheese samples increased significantly, reaching up to 2.94 Log₁₀ CFU/g, with visible late-blowing defects observed after five months in samples containing Cl. Tyrobutyricum T2 (CT) or mixed spores T3 (BL+CT), while in control cheese, spore count reached only up to 1.5 Log₁₀ CFU/g with no signs of visual spoilage. These results indicate that although NSLAB dominate during cheese ripening, their natural abundance alone is insufficient to prevent spoilage when high levels of bacterial spores are present, while in cheeses without spore inoculation, the naturally occurring NSLAB were able to suppress spores, resulting in no visible spoilage under standard ripening conditions. In the second phase (Chapter 4), inulin was incorporated into cheese samples at the time of manufacture to evaluate its prebiotic effect on NSLAB proliferation and its potential to control spoilage. Cheese samples were manufactured with four treatments: control with soluble fiber T1 (C SF), inoculated with aerobic spores of Bacillus licheniformis and soluble fiber T2 (BL SF), inoculated with anaerobic spores of Clostridium tyrobutyricum and soluble fiber T3 (CT SF), and a mixture of both spores and soluble fiber T4 (BL+CT SF). Soluble fiber (inulin) enriched cheese samples showed a significant increase in NSLAB counts, reaching an average count of 10.6 Log₁₀ CFU/g after three months and maintained at the same level after six months across all treatments, with L. rhamnosus and L. paracasei dominating the microbial community across all treatments. Spore counts remained below 0.9 Log₁₀ CFU/g throughout the ripening period in all fiber-enriched cheese samples irrespective of spores inoculation, with no late-blowing defects or slits/holes observed in any treatment. This effective suppression of spore proliferation is likely attributed to the enhanced growth and activity of NSLAB, particularly L. rhamnosus and L. paracasei, promoted by the addition of soluble fiber (inulin). The elevated NSLAB populations likely exerted competitive exclusion, produced antimicrobial compounds, and depleted key nutrients, thereby limiting the metabolic activity and germination potential of spores during ripening. Physicochemical parameters, including pH (4.9 to 5.1), protein, fat, and moisture content, remained within acceptable ranges, confirming that soluble fiber incorporation at a 1% level did not adversely affect cheese composition.