2.6. Quantification of surviving and damaged cells
The number of surviving bacteria/sporeswas counted at 0,1, 3, 7,
14, and 28 days. In all, 108 tubes were used to measure the initial
populations (six bacterial species, six rice wines, and three replications),
and 1080 tubes were used to measure the survival rates
(six bacterial species, five different storage times, two different
storage temperatures, six rice wines, and three replications). If no
survivors were detected, no further experiments were performed.
Briefly, each sample (1 ml) was diluted 10-fold in 0.85% sterile saline.
The diluents (100 mL) were then spread-plated onto each of the
selective media described in Section 2.4 and incubated at the
required temperature. For the lower detection limit, 200 mL of undiluted
sample was spread-plated on five plates (1 ml of undiluted
sample in total; detection limit ¼ 1 CFU/ml). After incubation,
colonies growing on the selective agar plates were counted. To
determine the number of damaged cells (viable but unable to grow
on the selective media), the remaining sample (about 8e9 ml) was
enriched by adding it to 80 ml of selective broth (see Section 2.4).
After incubation at 5 C or 22 C for the optimal time, the enriched
samples were then streak-plated onto selective agar plates as