140 Modern Food Microbiology
mung seeds with 98% of 750 nondisinfected and only 1.8% of surface-disinfected seeds containing a
number of mold genera including aflatoxigenic species although no aflatoxins were found.
1 On alfalfa
seeds, only 21% of 500 contained yeasts and molds.
Since sprouts are eaten without being heated or cooked, problems arise when the sprout seed
stock contains foodborne pathogens such as Salmonella or E. coli 0157:H7. Because of the excellent
nutrient state of germinating seeds as noted above, the pathogens grow very well. In an effort to control
pathogens, sprout seeds are treated in various ways to destroy these forms. The United States Food
and Drug Administration (FDA) recommends the use of 20,000 ppm calcium hypochlorite for 15 min.
to reduce pathogens, and this treatment does not significantly reduce germination efficiency or sprout
length.
Pathogens
The pathogen of greatest concern in RTU vegetables is C. botulinum and reasons for this concern are
pointed out by several studies. In one, five RTU vegetables (butternut squash, mixed salad, rutabagas,
romaine lettuce, and a stir-fry mix) were inoculated with a 10-strain cocktail—five each of proteolytic
and nonproteolytic spores.
3 The products were sealed in polystyrene trays with an oxygen transmission
rate (OTR) (see Chapter 14) of 2,100 ml and incubated at 5, 10, or 25
◦ C. All five vegetables became
toxic at some point during their storage. The time to toxin detection for nonproteolytic strains in
butternut squash was 7 days at 10
◦ C with CO 2 at 27.8%; and for proteolytics in this product, 3 days
at 25
◦ C with 64.7% CO 2 .
3 In butternut squash at 5
◦ C with an inoculum of nonproteolytic strains of
10
3 /g, toxin was detectable in 21 days. At the time of toxin detection in all samples, O 2 was <1%.
Although the packaging material was by no means of “zero” barrier quality, respiration of the products
decreased O 2 and increased CO 2 to the levels noted. It was the opinion of these investigators that the
temperature of storage of RTU vegetables of the type noted was of critical importance to their safety.
Most products were in states of detectable spoilage at the time of toxin detection.
Another study employed cabbage and lettuce inoculated with about 10
2 spores/g of a 10-strain
cocktail as above and packaged in film with an OTR of either 3,000 low OTR (LOTR) or 7,000 high
OTR (HOTR) and stored at 4, 13, or 21
◦ C for 21 or 28 days.
17 Toxin was not detected under any
conditions, and both vegetables were organoleptically spoiled before toxin could be produced. In the
cabbage stored at 21
◦ C for 10 days, the LOTR contained 69.4% and the HOTR 41.9% CO 2 , while in
lettuce at 21
◦ C after 8 days, CO 2 was 41.9% and 9.0% in LOTR and HOTR, respectively. In contrast
to the study by Austin et al.
3 where O 2 was <1%, both packaging materials allowed O 2 ranging
from 1.0% to 7.9%. In the former study, the packaging material had an OTR of 2,100, while in the
latter OTRs were 3,000 and 7,000. The more permeable film may have allowed the growth of other
organisms that interfered with C. botulinum.
A 10-strain cocktail of 7 proteolytics and 3 nonproteolytics was used in the study by Larson et al.
24 in
which five vegetables (broccoli, cabbage, carrots, lettuce, and green beans) were inoculated. Botulinal
toxin was found in all grossly spoiled broccoli stored at 21
◦ C, in one-half of those grossly spoiled at
12
◦ C, and in one-third of the grossly spoiled lettuce stored at 21
◦ C. No toxin was detected prior to
spoilage, and no toxin was found in the other three vegetables. In contrast to the two studies noted
above, the packaging material used in this study had varying OTRs ranging from 3,000 to 16,544, and
the vegetables were sealed under vacuum with vacuum pulled. Interestingly, broccoli was packaged in
material with OTRs of 13,013 to 16,544, while cabbage (which did not become toxic) was packaged
in 3,000 to 8,000 OTR materials. The broccoli packs stored at 21
◦ C for seven days contained <2%
O 2 and about 12% CO 2 , while lettuce at 21
◦ C for six days contained up to 40% CO 2 .
24 The APC of
spoiled products was in the 10
8 to > 10
9 range.
mung seeds with 98% of 750 nondisinfected and only 1.8% of surface-disinfected seeds containing a
number of mold genera including aflatoxigenic species although no aflatoxins were found.
1 On alfalfa
seeds, only 21% of 500 contained yeasts and molds.
Since sprouts are eaten without being heated or cooked, problems arise when the sprout seed
stock contains foodborne pathogens such as Salmonella or E. coli 0157:H7. Because of the excellent
nutrient state of germinating seeds as noted above, the pathogens grow very well. In an effort to control
pathogens, sprout seeds are treated in various ways to destroy these forms. The United States Food
and Drug Administration (FDA) recommends the use of 20,000 ppm calcium hypochlorite for 15 min.
to reduce pathogens, and this treatment does not significantly reduce germination efficiency or sprout
length.
Pathogens
The pathogen of greatest concern in RTU vegetables is C. botulinum and reasons for this concern are
pointed out by several studies. In one, five RTU vegetables (butternut squash, mixed salad, rutabagas,
romaine lettuce, and a stir-fry mix) were inoculated with a 10-strain cocktail—five each of proteolytic
and nonproteolytic spores.
3 The products were sealed in polystyrene trays with an oxygen transmission
rate (OTR) (see Chapter 14) of 2,100 ml and incubated at 5, 10, or 25
◦ C. All five vegetables became
toxic at some point during their storage. The time to toxin detection for nonproteolytic strains in
butternut squash was 7 days at 10
◦ C with CO 2 at 27.8%; and for proteolytics in this product, 3 days
at 25
◦ C with 64.7% CO 2 .
3 In butternut squash at 5
◦ C with an inoculum of nonproteolytic strains of
10
3 /g, toxin was detectable in 21 days. At the time of toxin detection in all samples, O 2 was <1%.
Although the packaging material was by no means of “zero” barrier quality, respiration of the products
decreased O 2 and increased CO 2 to the levels noted. It was the opinion of these investigators that the
temperature of storage of RTU vegetables of the type noted was of critical importance to their safety.
Most products were in states of detectable spoilage at the time of toxin detection.
Another study employed cabbage and lettuce inoculated with about 10
2 spores/g of a 10-strain
cocktail as above and packaged in film with an OTR of either 3,000 low OTR (LOTR) or 7,000 high
OTR (HOTR) and stored at 4, 13, or 21
◦ C for 21 or 28 days.
17 Toxin was not detected under any
conditions, and both vegetables were organoleptically spoiled before toxin could be produced. In the
cabbage stored at 21
◦ C for 10 days, the LOTR contained 69.4% and the HOTR 41.9% CO 2 , while in
lettuce at 21
◦ C after 8 days, CO 2 was 41.9% and 9.0% in LOTR and HOTR, respectively. In contrast
to the study by Austin et al.
3 where O 2 was <1%, both packaging materials allowed O 2 ranging
from 1.0% to 7.9%. In the former study, the packaging material had an OTR of 2,100, while in the
latter OTRs were 3,000 and 7,000. The more permeable film may have allowed the growth of other
organisms that interfered with C. botulinum.
A 10-strain cocktail of 7 proteolytics and 3 nonproteolytics was used in the study by Larson et al.
24 in
which five vegetables (broccoli, cabbage, carrots, lettuce, and green beans) were inoculated. Botulinal
toxin was found in all grossly spoiled broccoli stored at 21
◦ C, in one-half of those grossly spoiled at
12
◦ C, and in one-third of the grossly spoiled lettuce stored at 21
◦ C. No toxin was detected prior to
spoilage, and no toxin was found in the other three vegetables. In contrast to the two studies noted
above, the packaging material used in this study had varying OTRs ranging from 3,000 to 16,544, and
the vegetables were sealed under vacuum with vacuum pulled. Interestingly, broccoli was packaged in
material with OTRs of 13,013 to 16,544, while cabbage (which did not become toxic) was packaged
in 3,000 to 8,000 OTR materials. The broccoli packs stored at 21
◦ C for seven days contained <2%
O 2 and about 12% CO 2 , while lettuce at 21
◦ C for six days contained up to 40% CO 2 .
24 The APC of
spoiled products was in the 10
8 to > 10
9 range.