C. botulinum is frequently isolated from human and animal gastrointestinal tracts. Approximately 64% of samples of cow feces contain C. botulinum; 73% of which are non-proteolytic type B and only less than 5% are type E or F (Dahlenborg et al., 2003). According to Dahlenborg et al. (2001), C. botulinum type B is isolated from 62% of pig feces. Such a frequent occurrence of C. botulinum in feces of farm animals leads to meat contamination with its spores. Prior to the introduction of modern methods of meat processing, meat products had been the most frequently implicated source of botulism outbreaks. The Latin word botulus means sausage, hence C. botulinum was named after the meat products that were suspected to be its main source. However, spores may penetrate soil and contaminate plants via animal feces. Since the 1950s, vegetable products, surprisingly, have been the most important vehicles of C. botulinum in the U.S. The 'vegetable cases' - caused only by type A (61.1%) and type B (57.4%) - make up the majority of all reported botulism cases. Fruit and spices are also involved, but less frequently. It is significant that majority of cases (65.1%) are usually linked to home-made products.
Fish and seafood are generally responsible for infections caused by C. botulinum type E (Centers for Disease Control and Prevention, 1998). Many verified cases of botulism type E have been reported in Japan (166 cases and 58 deaths between 1951 and 1960). In 2003, C. botulinum type E was involved in the outbreak in western Alaska linked to consumption of a beached whale (Anonymous, 2003). Many outbreaks were also associated with a Japanese izuschi dish containing fermented raw fish, vegetables, and cooked and malted rice (okji). In Canada, Alaska, or Scandinavia, botulism is caused by consumption of fish and fermented meat dishes, very often prepared as traditional native dishes (Kotev et al., 1987; Knubley et al., 1995).
Uneviscerated salted mullet fish was a source of the outbreak (type E) in Egypt (Weber et al., 1993). The level of carbohydrates in such products is usually too low for lactic acid fermentation. If the pH of the product is not low enough, it will not protect against the outgrowth of C. botulinum.
Cucumbers and cabbage fermented by lactobacilli are popular dishes in the central and eastern Europe. Sauerkraut is frequently consumed by low-income communities, especially in winter. Surprisingly, no cases of botulism have been linked to consumption of such products. This observation may be explained based on the results of studies carried out by Braconnier et al. (2003). The authors analyzed germination of spores of C. botulinum type A and B, as well as changes in spore counts, in mushroom, broccoli, and potato purees. The addition of mixtures containing L-cysteine, L-alanine, and sodium lactate to vegetable puree induced an increase in germination, which suggests lack of these components in vegetable puree. Most probably, a low pH, lack of particular amino acids, and potential bacteriocin activity of lactobacilli in fermented vegetable products prevent spore germination.
Commercial products usually do not pose health threats to their consumers. However, botulism cases acquired after consumption of commercially prepared canned foods have been reported. In the U.S., 62 outbreaks occurred in the years 1899 to 1973 (Lynt et al., 1975). Only 7% of outbreaks reported between 1950 and 1996 were linked to commercially processed foods (Centers for Disease Control and Prevention, 1998). The implicated foodstuffs included chopped garlic in soy oil stored in glass bottles at room temperature (Louis et al., 1988), sliced roasted eggplant in oil, yogurt with hazelnuts, stuffed lotus rhizome, bottled caviar, and canned peanuts (Chou et al., 1988; D'Argenio et al., 1995).
Canned mushrooms were involved in many cases of botulism. It is suggested that Agaricus bisporus may induce C. botulinum spore germination due to oxygen consumption (Sugiyama and Yang, 1975).
Some cases are also connected with service establishments and caused mainly by improper temperature of food storage (Dodds, 1990).
Food stored in a proper way and in proper conditions is not a vehicle of C. botulinum. Unlike non-proteolytic strains, proteolytic strains will not grow in refrigeration temperatures. The number of spores in meat and poultry is rather low, much higher numbers are observed in fish. If stored at 3 to 5°C, vacuum-packed, not very sour meat products usually remain safe for consumers up to 21 days. Botulin toxin was not detected in raw rockfish fillets or red snapper homogenates after being stored for 21 days at 4°C. None of 1074 samples of commercially packed fresh fish stored for 12 days at 12°C contained botulin toxin (Lilly and Kautter, 1990).
The outgrowth of C. botulinum requires a suitable medium, temperature, atmosphere, pH, Eh potential, and water activity. Toxin is usually only produced in optimal or close-to-optimal conditions. Nutrient demands of C. botulinum are complex, and include amino acids, B vitamins, and minerals. In broth, non-proteolytic strains of type B and F grow and produce toxin at 4°C, but in crab meat the outgrowth and toxin production occurs solely at 26°C (Alberto et al., 2003).
It is believed that C. botulinum does not grow at pH lower than 4.5, yet botulin toxins were detected in sour home-canned foods. In Poland, toxins are sometimes detected in home-canned vinegar-preserved mushrooms. This may be explained by the presence of Aspergillus (Odlaug and Pflug, 1979) or other moulds. The outgrowth of moulds increases pH and enables growth of Clostridium. The growth of C. botulinum and toxin production depend also on the acidifying agent. The addition of even slight amounts of NaCl (3 to 4%) increases the susceptibility of Clostridium to pH - growth is inhibited at pH higher than 4.5.
A low water activity (aw), which for C. botulinum ranges from 0.94 (type B) to 0.97 (type E), enables the outgrowth and toxin production in packaged foods and is typical for foodborne pathogens. The actual effect of aw will depend on the strain properties and the type of agent regulating water activity.
The presence of other microorganisms is also very important for the outgrowth of C. botulinum. Botulin toxins in foods are usually inactivated by heat treatments. However, in non-processed food products, the presence of microflora significantly influences the outgrowth of clostridia. The outgrowth and toxin production is promoted by yeasts at pH 4.0. Yeasts produce a factor that enables the outgrowth of Clostridium at lower pH. A synergistic effect of clostridia and lactic acid bacilli (LAB) was also observed. However, LABs may also inhibit the outgrowth of C. botulinum due to bacteriocin activity. Such properties are revealed by Lactobacillus, Lactococcus, Streptococcus and Pediococcus (Rodgers et al., 2003). The outgrowth of C. botulinum may be inhibited by the presence of other Clostridium spp., e.g., C. sporogenes, and C. perfringens (Smith,1975). A similar activity was observed in case of Peanibacillus and Bacillus (Girardin et al., 2002). This is a commonly observed phenomenon of interspecies and intraspecies competition.
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