Enumeration of viable counts in raw milk using the automated fluorescence microscopic method.

摘要

An automated fluorescence microscopy method developed for the measurement of viable bacterial counts was applied to the detection and enumeration of viable bacteria in milk. In the automated analysis of viable bacteria, bacterial cells were recovered from milk after treatments with EDTA and Triton X-100, stained with 6-CFDA, and the stained cells were counted with the automated microscopy method. Salmonella enteritidis expressing the enhanced green fluorescent protein (S. enteritidis-EGFP) was used to elucidate the relationship between the fluorescent bacterial counts by the automated microscopy method and the viable counts by the conventional plating method. A high correlation coefficient (r2=0.97) was obtained for the relationship between the counts by the microscopy method and those by the plating method. When S. enteritidis-EGFP was recovered from sterile milk after treatments with EDTA and Triton X-100, the correlation coefficient was r2=0.91. Viable counts by the plating method and the counts of 6-CFDA stained bacterial cells by the automated microscopy method were determined for various raw milk samples. 37 raw milk samples were measured and the correlation coefficient was r2=0.73. It is suggested that the developed automated fluorescence microscopy method could also be used to detect specific enteropathogenic bacteria by using specific antibodies labelled with fluorescent dyes..