Improved Recovery of Stressed Bifidobacterium from Water and Frozen Yogurt

A roll-tube repair-detection procedure was developed to enumerate injured and noninjured cells of Bifidobacterium species from water and food samples. This procedure combined the Virginia Polytechnic Institute and State University's anaerobic roll-tube procedure and the repair-detection technique for detecting stressed cells. Mara and Oragui's human bifid sorbitol agar medium was modified for use in the roll-tube procedure by replacing the indicator bromocresol purple with phenyl red (0.027 g/l), adding 0.0006 g of methylene blue per 1, increasing the agar content to 25 g/l and adjusting the pH of the medium to 7.1 ± 0.1. The repair-detection roll-tube technique was shown to recover Bifidobacterium cells significantly (P < 0.01) better than pour plates, using the same medium incubated in anaerobe jars, even when a repair-detection system was used. Most repair in the roll tubes occurred in the first hour. B. adolescentis had a poor survival rate after 96 hours in water. Glucose was substituted for sorbitol in the medium used for enumeration of B. longum added to frozen yogurt, because this organism cannot utilize sorbitol. This medium, when used as part of a repair detection system, significantly (P < 0.01) recovered more cells than anaerobic pour plate techniques and was able to separate Bifidobacterium species and Lactobacillus acidophilus by colony morphology. Bifidobacterium cells were 1mm or larger, round and yellow, while the L. acidophilus colonies were so small (< 1/4 mm) their detection and enumeration was difficult.