360 Modern Food Microbiology
3. cured with NaCl or NO 2 ,
4. contain high levels of nonpathogens (for raw meat, poultry, and the like),
5. maintained in frozen state,
6. maintained at 40
◦ F (4.4
◦ C) or below, and
7. have a definitive self-life (e.g., not to exceed 10 days).
Since this organism is of greatest concern in such products, a number of studies have been conducted
relative to its behavior under MAP conditions.
The question of the organoleptic state of MAP fish products at the time of botulinal toxin production
has been addressed by a number of researchers, among whom are Garcia et al.
27 With an inoculum
of 13 nonproteolytic types B, E, and F spores at levels of 10
1 –10
4 , 50 g-samples of salmon fillets
were stored at varying temperatures and tested for toxin under vacuum, 100% CO 2 , and 70% CO 2 +
30% air. Overall, toxin detection coincided with spoilage at 30
◦ C but preceded spoilage at 8
◦ C and
12
◦ C, and followed spoilage at 4
◦ C.
27 Regarding start-up time to toxin detection, at 30
◦ C the fillets
were toxic after 1 day, after 2 days at 16
◦ C, 6 days at 12
◦ C, 6–12 days at 8
◦ C, and no toxin at 4
◦ C
in 60 days. Only type B toxin was detected. In another study, channel catfish was inoculated with a
mixture of four strains of type E at a level of three or four spores per gram and stored in 80:20 CO 2 :N 2
in O 2 -barrier bags at 4
◦ C and 10
◦ C.
9 Those stored at 10
◦ C all contained toxin by day 6. Those at
4
◦ C contained toxin on day 9 in overwrapped packages (O 2 permeable) but not until day 18 in those
stored under MAP. These investigators found that toxin detection and spoilage coincided at 10
◦ C,
while at 4
◦ C spoilage preceded toxin detection.
9 When raw beef was inoculated with types A and B
spores and stored for up to 15 days at 25
◦ C, toxin was first detected after 6 days, always accompanied
by significant organoleptic changes, indicating that the vacuum-packaged toxic samples would be
rejected before consumption.
45 In a later study, sliced raw potatoes were stored in O 2 -impermeable
bags with 30% N 2 + 70% CO 2 and incubated at 22
◦ C.
85 The untreated potatoes became toxic in 4
to 5 days while those treated with NaHSO 3 were toxic after 4 days, but appeared to be acceptable
through day 7. Type A toxin appeared earlier than type B. Potato salad was the vehicle food for two
cases of botulism in Colorado.
7 This product was temperature-abused in the home, and type A toxin
was found in leftovers.
Five vegetables (lettuce, cabbage, broccoli, carrots, and green beans) were used in a study assessing
the botulism hazard of MAP. They were inoculated with a spore cocktail of ten types A, B, and E (7
proteolytics, 3 nonproteolytics) and stored in bags with OTRs that varied between 3,000 and 16,544
ml at 4
◦ C, 12
◦ C, or 21
◦ C.
59 No toxin was found in any at 4
◦ C for up to 50 days, and no toxin was
found in cabbage, carrots, or green beans. Toxin was detected in all broccoli at 21
◦ C, in one-half of the
broccoli held at 12
◦ C, and in one-third of the lettuce held at 21
◦ C.
59 Both toxigenic vegetables were
grossly spoiled. These investigators concluded that toxin production does not precede gross spoilage.
In a study of MAP cabbage, an inoculum of 7 type A and 7 proteolytic type B strains was added
to shredded cabbage stored in high-barrier bags containing 70% CO 2 + 30% N 2 and incubated at
22–25
◦ C.
86 The inoculum size was 100–200 spores per gram. Only type A strains grew and produced
toxin as early as day 4 while the cabbage was still organoleptically acceptable. No toxin was detected
on day 3, and the product was organoleptically unacceptable by day 7.
86 The type B strains did not
produce toxin even with an inoculum of 14,000 spores per gram. In a survey of 1,118 commercially
available one-pound packages of precut MAP vegetables in the United States, only 4 contained type
A spores—1 each of shredded cabbage, chopped green pepper, and an Italian salad mix; another salad
mix contained types A and B.
62
Fresh Italian pasta was inoculated with types A, B, and F spores at a level of 1.2 × 10
2 spores/g
and stored at 12
◦ C and 20
◦ C for up to 50 days in an atmosphere of 15% CO 2 + 83% N 2 + 2% O 2 .
18
No toxin was detected in any tortelli that was stored at 12
◦ C but at 20
◦ C, toxin was detected in the
3. cured with NaCl or NO 2 ,
4. contain high levels of nonpathogens (for raw meat, poultry, and the like),
5. maintained in frozen state,
6. maintained at 40
◦ F (4.4
◦ C) or below, and
7. have a definitive self-life (e.g., not to exceed 10 days).
Since this organism is of greatest concern in such products, a number of studies have been conducted
relative to its behavior under MAP conditions.
The question of the organoleptic state of MAP fish products at the time of botulinal toxin production
has been addressed by a number of researchers, among whom are Garcia et al.
27 With an inoculum
of 13 nonproteolytic types B, E, and F spores at levels of 10
1 –10
4 , 50 g-samples of salmon fillets
were stored at varying temperatures and tested for toxin under vacuum, 100% CO 2 , and 70% CO 2 +
30% air. Overall, toxin detection coincided with spoilage at 30
◦ C but preceded spoilage at 8
◦ C and
12
◦ C, and followed spoilage at 4
◦ C.
27 Regarding start-up time to toxin detection, at 30
◦ C the fillets
were toxic after 1 day, after 2 days at 16
◦ C, 6 days at 12
◦ C, 6–12 days at 8
◦ C, and no toxin at 4
◦ C
in 60 days. Only type B toxin was detected. In another study, channel catfish was inoculated with a
mixture of four strains of type E at a level of three or four spores per gram and stored in 80:20 CO 2 :N 2
in O 2 -barrier bags at 4
◦ C and 10
◦ C.
9 Those stored at 10
◦ C all contained toxin by day 6. Those at
4
◦ C contained toxin on day 9 in overwrapped packages (O 2 permeable) but not until day 18 in those
stored under MAP. These investigators found that toxin detection and spoilage coincided at 10
◦ C,
while at 4
◦ C spoilage preceded toxin detection.
9 When raw beef was inoculated with types A and B
spores and stored for up to 15 days at 25
◦ C, toxin was first detected after 6 days, always accompanied
by significant organoleptic changes, indicating that the vacuum-packaged toxic samples would be
rejected before consumption.
45 In a later study, sliced raw potatoes were stored in O 2 -impermeable
bags with 30% N 2 + 70% CO 2 and incubated at 22
◦ C.
85 The untreated potatoes became toxic in 4
to 5 days while those treated with NaHSO 3 were toxic after 4 days, but appeared to be acceptable
through day 7. Type A toxin appeared earlier than type B. Potato salad was the vehicle food for two
cases of botulism in Colorado.
7 This product was temperature-abused in the home, and type A toxin
was found in leftovers.
Five vegetables (lettuce, cabbage, broccoli, carrots, and green beans) were used in a study assessing
the botulism hazard of MAP. They were inoculated with a spore cocktail of ten types A, B, and E (7
proteolytics, 3 nonproteolytics) and stored in bags with OTRs that varied between 3,000 and 16,544
ml at 4
◦ C, 12
◦ C, or 21
◦ C.
59 No toxin was found in any at 4
◦ C for up to 50 days, and no toxin was
found in cabbage, carrots, or green beans. Toxin was detected in all broccoli at 21
◦ C, in one-half of the
broccoli held at 12
◦ C, and in one-third of the lettuce held at 21
◦ C.
59 Both toxigenic vegetables were
grossly spoiled. These investigators concluded that toxin production does not precede gross spoilage.
In a study of MAP cabbage, an inoculum of 7 type A and 7 proteolytic type B strains was added
to shredded cabbage stored in high-barrier bags containing 70% CO 2 + 30% N 2 and incubated at
22–25
◦ C.
86 The inoculum size was 100–200 spores per gram. Only type A strains grew and produced
toxin as early as day 4 while the cabbage was still organoleptically acceptable. No toxin was detected
on day 3, and the product was organoleptically unacceptable by day 7.
86 The type B strains did not
produce toxin even with an inoculum of 14,000 spores per gram. In a survey of 1,118 commercially
available one-pound packages of precut MAP vegetables in the United States, only 4 contained type
A spores—1 each of shredded cabbage, chopped green pepper, and an Italian salad mix; another salad
mix contained types A and B.
62
Fresh Italian pasta was inoculated with types A, B, and F spores at a level of 1.2 × 10
2 spores/g
and stored at 12
◦ C and 20
◦ C for up to 50 days in an atmosphere of 15% CO 2 + 83% N 2 + 2% O 2 .
18
No toxin was detected in any tortelli that was stored at 12
◦ C but at 20
◦ C, toxin was detected in the