Microbiota and free amino acid dynamics during aging of Cougar Gold cheese: insights into the puffy defects

This study investigated microbial succession and free amino acid (FAA) dynamics in Cougar Gold Cheddar cheese over 18 months of commercial cold storage, with a focus on identifying microbial signatures associated with puffing spoilage. Fourteen proteinogenic FAAs and ornithine were monitored over time, revealing a steady increase in total FAA concentration, particularly flavor-active amino acids such as isoleucine, leucine, phenylalanine, and valine. A notable surge occurred after 9 months, suggesting enhanced proteolytic activity during late-stage ripening. Amplicon-based 16S rRNA sequencing revealed distinct microbial succession. Lactococcus dominated the early stages (0-3 months), followed by a marked increase in Lactobacillus, particularly Lb. helveticus, by 6 months, which remained dominant through 12 months before declining in the final stages. Paenibacillus wasatchensis was detected at later stages of aging. Cheese from puffed cans exhibited significantly higher alpha diversity and distinct microbial communities compared to standard-aged samples. Puffed samples were enriched in Pa. wasatchensis and several heterofermentative non-starter lactic acid bacteria, including Lb. curvatus, Lb. sakei, and Lb. graminis, accompanied by a reduction in Lb. helveticus. While overall FAA profiles were largely maintained, puffed samples showed modest reductions in aspartate, glycine, and ornithine, suggesting altered microbial metabolism. These findings provide valuable insights into the interplay between microbiota, proteolysis, and potential spoilage in aged cheese.