Clostridium botulinum: An overview and the dangers of neurotoxicity and Botulism
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Overview
By Emily Buckwalter
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What is Clostridium botulinum?
Clostridium botulinum is a large Gram positive, motile, spore-forming, rod-shaped bacteria that ranges from 4-6 μm by 0.9-1.2 μm in length and width respectively [10]. C. botulinum is a member of the family Clostridiaceae and of the order Clostridiales, under the branch of bacterial organisms known as firmicutes [1]. This bacterial species is commonly known for its ability to produce a dangerous neurotoxin, botulinum (BoNT) [1]. Even the smallest amount of exposure to this neurotoxin causes the onset of foodborne botulism, as it has been ranked one of the six highest risk threat agents for bioterrorism by the Centers for Disease Control and prevention (CDC) [9]. Botulism is a lethal and potentially fatal disease with sudden onset that ultimately results in extreme muscle weakness and paralysis [5]. There are other recognized types of botulism caused by C. botulinum, such as infant and wound botulism, but foodborne botulism is most commonly associated with this disease [6].
Given that this bacteria produces a lethal neurotoxin, it is classified as a pathogenic bacteria. C. botulinum has also been divided into four distinct phenotypes, groups I-IV, all of which produce one of seven (A-G) types of BoNT [3]. Group I phenotypes are proteolytic, meaning that they are able to break down protein, specifically by lysing native proteins, whereas Group II phenotypes are non-proteolytic [4]. Group I and II are specific to infecting humans as their host organism. Group III and IV strains are not as well defined in their capabilities because they are produced in either animal only or neither animal or human diseases [1]. The seven types of BoTN follow in a similar fashion, where only types A, B, E and F target human hosts and for that reason are present in both Groups I and II phenotypes [8].
Clostridium botulinum is composed of chromosomal DNA that is 3,886,916 base pairs long. The known genome of this bacterial species has been found to contain approximately 3,650 genes. However, the plasmid of C. botulinum that integrates into host DNA is around 16,344 base pairs long, encoding for a total of 19 genes [13]. The genes encoding BoNT have been suggested to be located in a cluster on the main chromosomal DNA or on the plasmid, however further research needs to be conducted to pinpoint an exact location [14]. While neurotoxin production is one of the widely studied functions of C. botulinum, most of the bacterial genome is responsible for encoding for proteases, enzymes that are used for the metabolism of proteins. In a study conducted by Sebaihia et al. 2007, C. botulinum was found to have very little newly acquired DNA, which tells us that the genome is very stable but also that there is not an extremely long relationship between the host organism and the bacteria, further emphasizing the role of the produced neurotoxin BoNT. Additionally this suggests that C. botulinum is well-adapted to its environment and can sufficiently carry out its own protection and metabolism without gene transfer to select for more optimal pathways [13].
Clostridium botulinum is of great importance to the food industry because it is one of many bacterial species that is commonly associated with the canning of foods, such as vegetables, that stock supermarket shelves [1]. C. botulinum is found just about everywhere in the form of spores, a stage in the reproductive cycle of spore-forming bacterial species [1]. These spores can easily become trapped in the packaging process in food production plants, and without proper heat and pressure to kill off these microbial spores, the bacteria will continue to grow and reproduce [2]. Clostridium botulinum are also well known to be found in the soil thanks largely in part to the excretion of bacterial spores from human and animal intestines [7]. Once these bacteria are in the soil, they can still pose a threat by infecting the roots of plants and ultimately crops that are consumed by humans and other animals. C. botulinum is also found in high abundance in aquatic environments, specifically sediments of oceans and lakes [8]. Researchers have been able to distinguish between the different types of BoTN found in these locations, which makes it easy to identify which phenotype of C. botulinum exists in that environment [1]. For example, type E has been found in high concentrations in the ocean, linking it to cases of botulism involving contaminated fish, whereas type A and B have been linked to foods associated with soil contamination [8].
Life Cycle
Clostridium botulinum grows and reproduces via the process of endospore formation and germination [1]. Most bacteria experience vegetative growth, whereby the cell divides in a 1, 2, 4… fashion and continues to multiply and divide [2]. However in the case of spore-forming bacteria, given specific stimuli typically from the environment, the bacteria can pause vegetation growth and start germination [2]. Since C. botulinum is able to form spores, this not only provides the cell with a means of protection but it also promotes the production of BoTN during the process of germination [11].
It is also known that spores can be activated based on environmental signals such as specific pH concentrations or temperatures, which is important especially in the food industry for means of food preservation [8]. Clostridium botulinum grows best under acidic conditions, specifically around a pH of 5 [10]. Each of the two major groups (Group I and Group II) that target humans as hosts have their own optimal temperature for growth at 35℃ and 28℃ respectively [8]. However, it is interesting to note that the Group II phenotype of C. botulinum, with BoTN types B, E and F, is able to generate neurotoxin at 3-4℃, which is standard refrigeration temperature [8]. This is dangerous especially for food production and safety. Therefore, it is important that any materials associated with food processing are properly sterilized, using extreme heat and pressure [2].
By selecting a temperature that is well out of the growth temperature for C. botulinum, along with pressure, this will effectively ensure that almost every microbe has been killed so that they cannot grow once packaged and stored with food. This is one of the reasons why home-canned food is related to many recent outbreaks of botulism [8]. Home canned foods typically can not ensure the same level of safety as commercial canner’s who are using equipment that produces extreme heat and pressure to kill any spores of C. botulinum that may be contaminating the packaging of the product [8]. Some spores are able to resist the cooking and boiling process of at home canning processes [8].
The danger surrounding C. botulinum presents itself under temperatures ranging from 40℉ to 120℉, anaerobic and low pH environments such as in a can, where these heat resistant spores can then convert and mature into growing cells [12]. Previous research has found that each C. botulinum group demonstrates different levels of heat resistance, with Group I spores being the most heat resistant, Group II being the least heat resistant [13]. The spores of Clostridium botulinum have been able to achieve heat resistance through the structure of the spore formed [13].
Clostridium botulinum is an obligate anaerobe, meaning that it does not function under the presence of oxygen, and therefore must reside in low oxygen environments to prevent the toxicity of oxygen to the cells [1]. In order to metabolize sugars to produce energy for the bacterial cell, Clostridium botulinum primarily will use the anaerobic process of fermentation to produce energy. However it has been identified that some species of C. botulinum are able to use both fermentation pathways, while still being equipped with a glycolysis pathway [13]. The fermentation pathway consists largely of a series of oxidation-reduction reactions that are coupled and require electron transport via specific carriers such as NADH [2]. C. botulinum has been found to ferment specific amino acids including: glycine, proline, phenylalanine and leucine, all of which are nonpolar amino acids [13]. However one of the largest means of energy production for the bacterial species comes from the breakdown of chitin, a polysaccharide. The bacteria accomplish this via five different encoded enzymes, but also with the help of proteases which are able to cleave some of the large polysaccharide into more manageable, smaller molecules for metabolic pathways [14]. Chitin is most commonly used by this bacterial species due to its relative presence in the environment. Chitin is most commonly found in the exoskeletons of arthropods and insects as well as in the cell walls of fungi, both of which are prevalent in marine and soil environments where C. botulinum live. Additionally, chitin can be a source of supplementary carbon and nitrogen for the bacteria [14]. Every point of information REQUIRES CITATION using the citation tool shown above.
Lyme Disease- Overview
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Metabolism and Energetics
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Dangers of Neurotoxicity
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Historical Outbreaks
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Current Research
Conclusion
References
Authored for BIOL 238 Microbiology, taught by Joan Slonczewski, 2022, Kenyon College
