Jenny ScottOffice of Food Safety

Center for Food Safety and Applied NutritionU.S. Food & Drug Administration

Bacillus spp. that Can Act as Spoilage Organisms and Pathogens:

Bacillus cereus, Bacillus licheniformis and Others

Characteristics of Bacillus spp.

• Sporeformers• Aerobic or facultatively anaerobic• Psychrotrophic thermophilic

– primarily mesophilic• Few produce gas during growth

Bacillus spp. as Food Spoilage Organisms

• Dairy – common – survive pasteurization that kills competitors

• Egg products – less common– Spoilage due to post-processing contamination with

psychrotrophic Gram-negative bacteria• Fresh vegetables – important in spoilage

– Carrots, cucumbers, onions, potatoes, tomatoes• Bakery products – rope spoilage

Canned Food Spoilage• Bacillus spp. are common when processes are

improperly applied• Common cause of thermophilic spoilage

– flat sour spoilage (e.g., B. stearothermophilus)

• Common recontaminant (post-process contamination)• Some species can raise the pH

– Bacillus licheniformis– Concern that this could allow germination, growth and toxin

production by Clostridium botulinum

pH Elevating Bacillus licheniformis• In laboratory studies, several strains shown to grow at

pH as low as 4.2 (but not 4.0)• Several strains shown to elevate the pH in tomato puree

from 4.4 to 5.0-5.5 in one week at 30ºC• Occasionally pH exceeded 4.6 after 7 days without

visible signs of microbial growth. • Inoculated packs – surviving spores raised pH in jars in

which the seals were broken, but not when they were intact

Montville and Sapers, 1981

Metabiotic Effect of Bacillus licheniformis

• In laboratory studies, tubes of media (pH 4.4) co-inoculated with 100 B. licheniformis and 100 C. botulinum spores and incubated at 30ºC, 8 days:– Developed a pH gradient, e.g., pH 8.2 surface, pH 6.2

in zone of anaerobic growth– Contained botulinum toxin

Montville 1982

pH Elevating Bacillus licheniformis• Risk of botulism low but not zero

– the presence and survival of both C. botulinum and B. licheniformis spores

– a poor jar seal – permissive post-process temperature – failure to heat prior to consumption– an unobservant consumer

• Low probability event can be expected eventually• May depend on food type

Montville and Sapers, 1981; Montville, 1982

Other pH-elevating strains• Anderson, 1984

– B. coagulans strain grew in heat-processed tomato juice

– Elevated juice pH from 4.5 to 5.07 in 6 days at 35ºC• Dujaili and Anderson, 1991

– Isolated 24 strains of Bacillus spp. from soil or vegetable materials that elevated pH of tomato juice

– Morphologically and physiologically different from B. coagulans and B. licheniformis

FDA Position on pH Elevating Strains ofBacillus spp.

• FDA draft guidance on acidified foods:– thermal process for an acidified food with a pH in the

range of 4.0 – 4.6 is developed to ensure that the spores of acid-tolerant spoilage microorganisms (e.g., B. licheniformis) are destroyed.

– D200F = 4.5 min, z=27ºF

Bacillus spp. and Foodborne Illness

• Bacillus cereus is recognized as the species that most commonly causes foodborne illness

• CDC estimates 63,400 B. cereus foodborne illness cases annually in the U.S.

Bacillus cereus Group

• B. anthracis• B. cereus• B. mycoides• B. pseudomycoides• B. thuringiensis• B. weihenstephanensis

B. cereus Foodborne Illness• Diarrheal

– Enterotoxins produced by growth of the organism in the intestine

• Emetic (vomiting)– Ingestion of pre-formed emetic toxin cereulide– New cereulide strains of B. cereus emerging– Recent detection of cereulide-producing strains of B.

weihenstephanensis• Both can be experienced concurrently

Symptoms of Illness• Diarrheal

– Abdominal pain, diarrhea, occasionally nausea and vomiting 8-16 h after ingestion

– Recovery 12-14 h• Emetic

– Nausea, vomiting, abdominal cramping 1-5 h after ingestion

– Recovery 6-24 h

Severe Illness Reported• Outbreak in Norway from stew containing 104 –

105 B. cereus per serving affected 17 people– 3 hospitalized, one for 3 weeks

• 3 reported cases of fatal B. cereus food poisoning in journal reports between 1963 and 1977

Severe Illness Reported• 1997 report in NEJM of fatal fulminant liver failure in a

17-year old boy after consumption of spaghetti with homemade pesto. – Prepared four days earlier and refrigerated; on several

occasions it had been left at room temperature for one or more hours before being reheated in a pan

– B. cereus strain produced both enterotoxin and emetic toxin – enterotoxin and emetic toxin were likely to have caused the

gastrointestinal symptoms, but the emetic toxin was responsible for the liver failure

Mahler et al. 1997

Severe Illness Reported• 2005 report in J. Clinical Microbiol. of a fatal

case due to liver failure in a 7-year old girl after consumption of pasta salad. – Prepared Friday, taken to picnic Saturday; leftovers

stored in fridge (temperature 14ºC) until Monday evening supper

– Multiple B. cereus strains detected in multiple food samples, with 107 – 108 CFU/g in pasta salad

– Same PFGE from pasta salad, vomit of deceased girl and pasta bowl

Dierick et al. 2005

Psychrotrophic B. cereus• Psychrotrophic strains of B. cereus have been reported.• Despite a high worldwide occurrence of B. cereus in

milk, surprisingly few reports on food poisoning have been reported.

• The incidence of cytotoxigenic B. cereus is high in milk and cream, but the significance of the toxins is not known.

• There is limited epidemiologic evidence that growth of B. cereus in a food held at refrigerated temperatures is a significant cause of foodborne illness.

Other Bacillus spp. Causing Foodborne Illness

• B. thuringiensis – occasionally reported responsible for outbreaks of foodborne illness– Laboratory studies – presence of enterotoxin genes and toxicity

of culture supernatants• B. licheniformis outbreaks have been associated with

cooked meats and vegetables.– Toxic agent with physicochemical properties similar to cereulide

but different biological activity identified.• B. subtilis, B. pumilus – cytotoxicity has occasionally

been demonstrated

Other Bacillus spp. that May Be Potential Foodborne Pathogens

• Enterotoxin gene targets present by PCR– B. amyloliquefaciens, B. circulans, B. lentimorbis, B.

pasteurii (Phelps and McKillip. 2002)• Found to produce diarrheal enterotoxin in infant

milk formula– B. licheniformis, B. circulans, B. megaterium (Rowan

et al., 2001)

Likelihood that Other Bacillus spp. are Etiologic Agents

• Etiologic agent is not identified in ~30% of all foodborne outbreaks and 14% of cases in U.S. each year

• Sporadic cases of foodborne illness - likely to include illnesses from organisms yet to be identified as etiologic agents

• Bacillus spp. may be responsible for some of these illnesses

Thank you!