CHALK CULTURE METHOD FOR PRESERVING BACTERIA CULTURES AND ITS APPLICATIONS¹

Abstract

A simple and efficient chalk culture method was developed to preserve bacterial cultures in this study. The stability of Bacillus cereus, Enterobacter aerogenes ATCC 13048, Escherichia coli, Escherichia coli O157:H7 ATCC 43895, Listeria monocytogenes, Pseudomonas aeruginosa ATCC 25619, Salmonella Typhimurium and Staphylococcus aureus chalk cultures at 25C and 4C was evaluated at 0, 1, 2, 3 and 9 months on Tryptic Soy Agar and a specific agar for each organism. During drying process, B. cereus, E. coli O157:H7 and S. aureus chalk cultures showed less than 1.0 log CFU/g reduction, while E. aerogenes, E. coli, L. monocytogenes, P. aeruginosa and S. Typhimurium showed between 1 to 3 log CFU/g reduction. In the shelf‐life study of chalk cultures, B. cereus and E. coli O157:H7 were stable (< 1 log CFU/g reduction) at both 25C and 4C during the first 3 months. E. aerogenes, E. coli, L. monocytogenes, P. aeruginosa and S. aureus had better stability at 4C (<1 log CFU/g reduction) than 25C (<3 log CFU/g reduction) (P<0.05) during the first 3 months. S. Typhimurium was not stable at 25C and 4C during the first three months (P<0.05). From the third month to the ninth month, B. cereus, E. coli O157:H7, L. monocytogenes and S. Typhimurium were stable at both 4C and 25C (<1 log CFU/g reduction), while E. aerogenes, E. coli, P. aeruginosa and S. aureus were more stable at 4C than at 25C. E. coli O157:H7, L. monocytogenes, S. Typhimurium and S. aureus chalk cultures were inoculated into milk, ground beef and dry pet food, and retained constant (< 1 log CFU/mL or g reduction) after 5 days storage indicating that chalk cultures are stable in these foods. For the heat treatment (55C for 5 min) study, the counts of chalk cultures in milk and ground beef reduced approximately 2 log CFU/mL or g, and about 1 log CFU/g in pet food after heating. The difference was statistically significant compared to the unheated‐inoculated sample (P<0.05). The chalk cultures are easy to prepare, stable in storage and convenient to use for challenge studies especially for dry solid food samples.

CHALK AND FLUID INOCULATION METHODS IN STUDYING THE PERCENT RECOVERY OF CLOSTRIDIUM PERFRINGENS, ESCHERICHIA COLI AND ENTEROCOCCUS FAECALIS FROM RAW GROUND BEEF

“…A new simple method was developed by Hoffmans and Fung in 1993 called chalk culture. This method uses dried chalk powder as a medium to preserve bacteria (Liu et al. 2003).…”

“…They found that C. perfringens retained around 2 log cfu/g from an initial count of 4 log cfu/g, and all the other bacteria retained more than 4 log cfu/g from an initial count of 5–8 log cfu/g for 6 months. Furthermore, Liu et al. (2003) compared the shelf life of some bacterial chalk cultures at 25 and 4C.…”

“…Typhimurium in dried poultry feed in a validation study. Furthermore, Liu et al. (2003) studied the stability of some microorganisms' chalk cultures in food.…”

“…The hypothesis is that the microorganisms are already grown in fluid media, such as brain heart infusion (BHI), so they may have a shorter lag period in food, leading to a higher percent recovery. While the microorganisms do not grow in powder, and they do not grow in food once inoculated in powder form (Liu et al. 2003), this inoculation method should give a more precise and reliable percent recovery.…”

“…Preparation of Chalk Cultures. The procedure of Liu et al. (2003) was used to prepare chalk cultures.…”