Cold-tolerant microorganisms causing spoilage of vacuum-packed beef under time-temperature abuse determined by culture and qPCR

Vacuum packaging and storage conditions at chilled temperatures are commonly used in order to prolong the shelf life of meat. Under these conditions and time-temperature abuse, cold-tolerant (facultatively) anaerobic spoilage microorganisms can continue growing. This study investigated growth of six relevant spoilage micro-organisms in vacuum-packed beef (n = 12, 72 subsamples, stored at 10 degrees C for 28 days) using culture and qPCR methods. Correspondingly, six qPCRs were newly developed/modified (for total bacteria, lactic acid bacteria (LAB), Enterobacterales, total fungi, Kazachstania psychrophila, and cold-tolerant Clostridium spp.). Besides mi-crobial quantification, four spoilage appearances of meat (gas production, spoilage odor, % drip loss, and meat color) were observed. Results obtained from culture and qPCR show that total bacteria, LAB, and Enter-obacterales reached their stationary phase at day 7 when spoilage parameters such as gas production were statistically increased and a deviation of odor was detected. Fastidious cold-tolerant Clostridium spp. and K. psychrophila could be detected from day 7. Based on microbiological and sensory analysis results, the maximum shelf life of vacuum-packed beef stored at 10 degrees C is 7 days. The developed qPCR has the potential to be used as an alternative method to culturing for determination of microbial growth.