Abstract: Bacillus cereus continues to be a growing concern for a multitude of manufacturers within the food industry. Its ability to survive in a variety of environments, and its ability to resist and mutate against standard methods of sterilization and cleaning induces large concern for all producers . This paper is a review outlining not only the physiological characteristics of b. cereus and its ability to cause illness and survive in a multitude of environments, but also how these mechanisms continue to affect the food industry and how to control their highly abrasive nature.
Bacillus cereus is a spore-forming, Gram positive, motile, facultative anaerobic bacterial rod that has imposed tremendous threat due to its ability to germinate and thrive in both volatile and nonvolatile environments (Buttone, 2010). The morphology and phenotype in which B. cereus exhibits is greatly dependent on the environment in which it is cultured. For example, when isolated and streaked on a broth culture, B. cereus is straight or slightly curved with a slender rod shape (Marrollo, 2016). Furthermore, this broth indicates a viable cell living in optimal conditions such as body fluids and other nutrient rich environments. However, when isolated and streaked on an Agar plate, its morphology consists of more uniform, rod-shapes as well as containing an oval spore within its center (Marrollo, 2016). This provides evidence towards its ability to develop a secondary defense against its external environment in the chance that conditions become less than desirable. A variable also affecting its phenotypic variation is temperature. B. cereus can viably withstand temperatures ranging from as low as 10C and as high as 50C (Marrollo, 2016).
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Beyond its observable characteristics, B. cereus contains genotypic functions that help prolong life when conditions decrease in sustainability. For example, the production of biofilm facilitates in further protecting the bacteria from antimicrobials and common sanitation procedures (Galie et al., 2018). However, in circumstances that completely inhibit the growth of vegetative cells, B. cereus can protect its DNA and become dormant by producing endospores until external conditions become more favorable/sustainable (Hoover and Markland, 2016).
Although microbiologists have been implementing ways to prevent and control the spread of B. cereus for decades, it is estimated that on average, over 60,000 illnesses, and up to 20 hospitalizations each year are caused by the intoxication of B. cereus (Markland and Hoover, 2016). As technology advances and population grows, the demand for larger output in a shorter amount of time imposes great risk for the growth and contamination of B. cereus. Furthermore, this review analyzes B. cereus’s resistance to the food industry as well as current research pertaining to the pathogenic characteristic of B. cereus to further convey the importance of both preventing and controlling its germination.
2. Bacillus cereus Resistancetothe Food Industry
Understanding the morphology and mechanisms of B. cereus further exemplifies the wide range of environments where the foodborne pathogen can colonize (Marrollo, 2016). B. cereus can germinate in various types of soils, plants, and decaying organic matter (Markland and Hoover, 2016). Due to its motility, it can also survive in both fresh and marine aquatic environments (Marrollo, 2016; Markland and Hoover, 2016). Along with its ability to germinate in a variety of temperaments, its ability to form endospores and biofilm increases the duration of which B. cereus can live within these ecosystems and more specifically, the food industry.
2.1 Biofilm Production
Biofilms are complex in nature and can contain a single organism or multiple organisms immersed in an extracellular matrix that facilitates not only adhesion to surfaces but protection from toxic compounds that may hinder bacterial growth (Coughlan et al., 2016; Galie et al., 2018). The extracellular matrix consists of proteins, lipids, cellulose, and DNA which form a nutrient rich film (Coughlan et al., 2016; Galie et al., 2018). Many bacteria form biofilm as a defense mechanism. More specifically, foodborne bacteria such as Bacillus cereus.
The formation of B. cereus biofilm enables a nutrient rich environment allowing other bacterial species to colonize within the biofilm (Galie et al., 2018). Areas of which are easily permissible by B. cereus biofilm include factory stainless steel, pipes, and conveyer belts (Wijman et al., 2007). B. cereus is also known to have flagella movement both at the air-liquid interface of the biofilm as well as the surface submerged beneath the biofilm (Galie et al., 2018). In the food industry, this motility allows the bacteria to travel long distances along pipelines increasing the probability for contamination over a wide-spread area (Galie et al., 2018). While there are many strains of B. cereus that alter genotypic variations and health concerns, some require phospholipids and surfactants as a nutrient rich medium to colonize . This imposes a high risk on equipment in dairy factories and can cause cross-contamination, lower shelf life, and furthermore an increase health risk associated with the product . While other pathogens can be removed by the flow of water, milk, and other nutrient inhibiting substances, biofilm created by B. cereus can create a sustainable environment for a multitude of organisms to grow and help protect them from instability within their external environment (Marchand et al., 2012).
2.2 Endospore Production
Bacterial endospores are dormant forms of capsulation that many organisms use to resist unsustainable environments (Markland and Hoover, 2016). Spores can withstand the most extreme environment conditions including high pressure, extremes of pH, disinfectants, substantial heat, and many other conditions that induce intense stress to a spore-forming organism (Markland and Hoover, 2016). The main method of extinguishing endospores today is by forcing germination by extreme heat shock and sterilizing the vegetative cell, which is less resistant to disinfection and sterilization (Markland and Hoover, 2016). However, not all spores germinate at the same rate or by the same methods (Ghosh and Setlow, 2009).
B. cereus produces a spore that is extremely resistant to the external environment (Ghosh and Setlow, 2009). Known as a superdormant spore, it requires greater efforts to force germination (Ghosh and Setlow, 2009). In an experiment conducted by Ghosh and Setlow, they developed a method to isolate spores based on differing rates of germination through what they term, buoyant density configuration (2009). This process consists of numerous cycles of heat shock causing normal spores to germinate and separate from spores that germinate at slower rates and require higher heat activation (Ghosh and Setlow, 2009)
3. Pathogenic Characteristics of Vegetative Bacillus cereus Cells
The dormancy of endospores does not impose immediate threat to a producer. However, if conditions allow them to germinate, B. cereus can create substantial risk to the environment around it. In cases of poor sanitation practices, B. cereus can cause substantial damage to producers and consumers. B. cereus can withstand great resistance to biological control . Once B. cereus germinates into a viable cell, it can produce harmful toxins that cause disease and intoxication to consumers (Carretto et al., 2016). There are two types of food poisoning caused by B. cereus, emetic and diarrheal (Carretto et al., 2016).
2.1 Emetic Food Poisoning
Emetic food poisoning is caused by a single toxin known as cereulide (Agata et al., 2002). Cereulide is resistant to acidic environments and substantial heat (Carretto et al., 2016). The production of this toxin forms during the beginning of the stationary phase of growth and retains production between the temperatures of 12-37C (Carretto et al., 2016). Due to its formation only during the stationary phase of production, it requires a high number of vegetative cells to impose threat (Markland and Hoover, 2016; Carretto, 2016). Its high resistance to the external environment and extremely low pH makes it evasive from deactivation by human enzymes (Carretto, 2016). The cereulide toxin is considered intoxicating, meaning it requires ingestion to enter the bloodstream, and can lead to nausea and emesis lasting, at the most, 24 hours (Markland and Hoover; 2016).
2.2 Diarrheal Food Poisoning
The diarrheal form of food poisoning is more complex in that multiple enterotoxins and degenerative enzymes cause intoxication (Markland and Hoover, 2016; Carretto et al., 2016). While cereulide is produced during the stationary phase, enterotoxins are produced during the exponential phase (Markland and Hoover, 2016). This means that B. cereus does not require a high number of vegetative cells to produce these toxins, rather if it is colonizing, these toxins are released. Fortunately, enterotoxins are less heat resistant than cereulide meaning they can be destroyed by heating or cooking (Markland and Hoover, 2016). As far as symptoms, the disease consists of diarrhea, abdominal pain, and occasionally nausea lasting up to 48 hours (Markland and Hoover, 2016).
3. Methods to Prevent Bacillus cereus Germination
Although B. cereus directly effects the consumer, producers are indirectly effected by the cost and liability B. cereus can cause because of poor sterilization practices. Good Manufacturing Practices and Hazard Analysis Critical Control Point schemes are resourced to food processing facilities to guarantee that food quality and safety is given conscious awareness and high priority (Sharma and Anand, 2002; Alvarez-Ordinez et al., 2016). As the food industry expands, companies are becoming more aware of how dangerous and prevalent B. cereus contamination can be, and how important good sterilization practices are for not only the prevention but control of foodborne pathogens such as B. cereus.
Upstream and downstream processes deal with the growth and development of product before it reaches the consumer. It is the producer’s responsibility to prevent the germination and presence of spores before products reach the consumer. B. cereus in the form of spores can contaminate the agriculture development of food before reaching packaging facilities (Markland and Hoover, 2016). Its ability to grow in soil and plants imposes risk and cross-contamination in the presence of poor washing and sterilization practices. Raw materials that are at high risk include rice, milk, grains, potatoes, vegetables, and low-nutrient foods (Markland and Hoover, 2016). Although spores are resistant to a variety of sterilization methods, the use of effective surface cleaning and washing techniques as well as controlled packaging environments can prevent the germination of spores (Markland and Hoover, 2016). Spores can only germinate when conditions allow them to, therefore the absence of organic matter as well as unsustainable temperatures lower the probability that viable cells will form (Markland and Hoover, 2016). Storing products at low temperatures is a method in preventing the germination of spores as well (Marrollo, 2016). However, it is important to have an awareness of the food pathogens that put certain products at risk. Foods that are susceptible to B. cereus must be stored in temperatures below normal regulation (Markland and Hoover, 2016). Psychotolerant spores can germinate in temperatures below 15C (Markland and Hoover, 2016).
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