Environmental and Nutritional Requirements of Lactobacillus casei
Lactobacillus species have attracted considerable attention during the past few years due to the health benefits to the gastrointestinal tract when they are added as probiotics to food products such as yogurt and other fermented milk drinks (Broadbent, Larsen, Deibel, & Steele, 2010). In general, Lactobacillus species exhibit very complex nutritional and environmental requirements for optimal growth. Particularly, Lactobacillus casei is a rod shaped, aciduric (prefers acidic environments), facultative (capable of living in aerobic or anaerobic conditions) and mesophilic (temperature range between 15-40°C) lactic acid bacterium. These characteristics allow its isolation from different environments including fermented milk, meat or vegetable products, and the oral, gastrointestinal, and reproductive tracts of mammals. This just proves that this microorganism has a very versatile and easily adaptable metabolism (Broadbent et al., 2012). Moreover, recent studies have demonstrated that Lactobacillus casei strains exhibit a hypervariable region in their genome that help them survive in various habitats and under different environmental conditions (Broadbent et al., 2012). This adaptability has been attributed to horizontal gene transfer from other bacterial species and particularly from close relatives of the Lactobacilli family. In terms of amino acid biosynthesis, Lactobacillus casei is equipped with all the enzymes to produce all amino acids with the exception of valine, leucine and isoleucine. Also, it can use a system of proteolytic enzymes to extract amino acids from the environment (Cai, Rodriguez, Zhang, Broadbent, & Steele, 2007; Cai, Thompson, Budinich, Broadbent, & Steele, 2009). The system is composed by an extracellular proteinase called lactoceptin, a transport system to pass peptides along the cell membrane, and intracellular peptidases to process the incorporated oligopeptides. The carbohydrate metabolism is quite varied: Lactobacillus casei is capable of using ribose, galactose, fructose, sucrose, mannitol, mannose, N-acetyl glucosamine, cellobiose, maltose, trehalose, turanose, salicin, melezitose and inulin (Cai et al., 2007, 2009). During growth, Lactobacillus casei produces important amounts of lactic acid from hexose sugars using the Embden-Meyerhof pathway and phosphoketolase pathway (Cai et al., 2007, 2009). Additionally, Lactobacillus casei can survive on pentoses by using the pentose phosphate pathway. This has been exploited at the industrial level for the production of fermented foods. Lactic acid is transported by Lactobacillus casei across the cell membrane through a symporter in the form of lactate ion. Upon accumulation on the exterior medium, the lactate molecule protonates and the pH start to drop. This protonated form can return to the cytoplasm by simple diffusion due to its higher membrane solubility and the pH gradient with respect to the cell interior, which is maintained at a much more basic level by Lactobacillus casei. This buffering capacity is what renders Lactobacillus casei its exceptional capacity to tolerate extremely acidic environments (Broadbent et al., 2010). This attribute is called inducible acid tolerance response (ATR) and is associated with specific protein expression and physiological changes that include altered membrane lipid content, malolactic fermentation and intracellular accumulation of histidine (Broadbent et al., 2010).
Environmental and Nutritional Requirements of Clostridium botulinum
Clostridial microorganisms are capable of producing some of the most potent neurotoxins. These substances are lethal and unfortunately have found application in the field of biological warfare (Mitchell, Tewatia, & Meaden, 2007). Particularly, Clostridium botulinum species produce botulinum neurotoxins, which can attack the nervous system causing a major blockage in the production of acetylcholine, a chemical responsible for mediating nerve impulse transmission. As a result, blurred vision, and difficulty in swallowing and speaking will be experienced. Clostridium botulinum is a rod-shaped, gram positive, obligate anaerobe (requires complete absence of oxygen), mesophilic, and spore forming (dormant state to assure survival under adverse conditions) microorganism. Clostridium botulinum can be found in acidified substrates or media (pH 4.8 to 7) and for this reason is commonly isolated from poorly canned or refrigerated foodstuff (Mitchell et al., 2007; Sebaihia et al., 2007). Fish, corn, beets, spinach, asparagus, chili peppers, sautéed onions and potatoes, apricots, pears and peaches are particularly susceptible to lower their pH as storage time passes. Accordingly, they serve as an ideal media for Clostridium botulinum proliferation Clostridium botulinum has a set of proteases that are secreted to act upon substrates available in the environment. Some of the proteases include alpha-clostripain, several thermolysin metalloproteases, two hemolysins and two peptidases (Sebaihia et al., 2007). The obtained peptides and amino acids upon protease action are incorporated by the microorganism through the numerous specialized transporters. Clostridium botulinum is also able to ferment amino acids such as glycine and proline by their incorporation into specialized fermentation pathways (Sebaihia et al., 2007). Besides amino acids, Clostridium botulinum is fully equipped to survive on various complex polysaccharides and sugars (Sebaihia et al., 2007). For instance, Clostridium botulinum is capable of secreting five chitinases, enzymes with the ability of degrading chitin, a very abundant linear polymer of N-acetyl-glucosamine (Sebaihia et al., 2007). Clostridium botulinum also expressed an enzyme called beta-amylase that is responsible for removing maltose molecules from the nonreducing ends of the starch polymers. For simple sugars and sugar derivative-phosphorylation, Clostridium botulinum counts on 15 phosphoenolpyruvate (PEP)-dependent phosphotransferase systems (PTS) (Sebaihia et al., 2007). It has been reported to conduct the complete glycolysis pathway and some reactions of the Krebs Cycle Additionally, it has the ability to produce either acids such as acetate and butyrate or solvents such as butanol and ethanol (Sebaihia et al., 2007).
Both Lactobacillus casei and Clostridium botulinum are rod-shaped, mesophilic microorganisms, and exhibit very complex metabolisms that are still subject of research studies. While Lactobacillus casei can be very useful in improving human health by increasing the nutritional value of food, Clostridium botulinum produces a neurotoxin capable of killing a person almost instantaneously. Even though both organisms prefer to thrive in acidic media, Lactobacillus casei has unique adaptations to tolerate extremely low pH values. They both have the ability to metabolize proteins and have sophisticated transporter systems to incorporate the products of these activities. They both have the enzymatic system to process sugars and in the case of Clostridium botulinum, even complex polysaccharides. This flexibility has conferred both organisms tremendous adaptability to different niches and conditions. It is believed that Lactobacillus casei obtained this ample genetic pool via horizontal gene transfer, however, there is no evidence, thus far, that this was the case for Clostridium botulinum.
Broadbent, J. R., Larsen, R. L., Deibel, V., & Steele, J. L. (2010). Physiological and Transcriptional Response of Lactobacillus casei ATCC 334 to Acid Stress. Journal of Bacteriology, 192(9), 2445–2458. doi:10.1128/JB.01618-09
Broadbent, J. R., Neeno-Eckwall, E. C., Stahl, B., Tandee, K., Cai, H., Morovic, W., Steele, J. L. (2012). Analysis of the Lactobacillus casei supragenome and its influence in species evolution and lifestyle adaptation. Bmc Genomics, 13. doi:10.1186/1471-2164-13-533
Cai, H., Rodriguez, B. T., Zhang, W., Broadbent, J. R., & Steele, J. L. (2007). Genotypic and phenotypic characterization of Lactobacillus casei strains isolated from different ecological niches suggests frequent recombination and niche specificity. Microbiology-Sgm, 153, 2655–2665. doi:10.1099/mic.0.2007/006452-0
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Mitchell, W. J., Tewatia, P., & Meaden, P. G. (2007). Genomic analysis of the phosphotransferase system in Clostridium botulinum. Journal of Molecular Microbiology and Biotechnology, 12(1-2), 33–42. doi:10.1159/000096457
Sebaihia, M., Peck, M. W., Minton, N. P., Thomson, N. R., Holden, M. T. G., Mitchell, W. J., Parkhill, J. (2007). Genome sequence of a proteolytic (Group I) Clostridium botulinum strain Hall A and comparative analysis of the clostridial genomes. Genome Research, 17(7), 1082–1092. doi:10.1101/gr.6282807