Bacillus subtilis

Original article: “Bacillus subtilis” by Giovanna Spinosa and updated by Gennaro Velotto

Characteristics

Bacillus subtilis, also known as hay or pasture bacillus, is a Gram-positive bacterium belonging to the genus Bacillus. Its genome has 4,214,810 base pairs and contains 4,100 genes useful for encoding specific proteins.

This bacterium, commonly present in the soil, is not a human pathogen. In fact, even though it is capable of degrading, contaminating, and modifying foods, it rarely causes food poisoning. Moreover, we are talking about a microorganism remarkably susceptible to genetic manipulation and not dangerous: it is widely adopted as a model for laboratory studies, in particular on the sporulation process (simplified way to follow cell differentiation).

Phylogeny

Colony morphology

B. subtilis is a rod-shaped, flagellated obligate aerobic microorganism (Fig. 1). It can form a hard protective structure, a protective endospore, allowing the organism to tolerate extreme environmental conditions. These structures will be described below (Spore section).

In cultures with blood agar (Fig. 2), they form colonies as large as 2/4 mm that appear smooth, rough, or mucousy with edges that may be extended into the medium or wavy.

Identification methods

The cells of these organisms are less than 1 μm wide, have sporangium without swelling, and the spores are ellipsoidal in shape (Fig. 3). They are usually mesophilic for temperature and neutrophilic for growth pH, although they are often tolerant to higher pH levels.

Spores

In specific conditions, not optimal for growth, Bacillus subtilis can be in the form of a spore (otherwise called endospore) living in a quiescent phase. In particular, spores are produced by the bacterium by a specific process called sporulation that is activated when other survival strategies cannot be exploited. Therefore, sporulation in B. subtilis is an adaptive response to stress that involves the development of biological structures essential for the preservation of the species and the long-term conservation of genetic material.

Endospores are specific structures produced by prokaryotic organisms in response to environmental stressors that create unsuitable living conditions. Several factors can result in this phenomenon, such as suboptimal temperatures and low water or nutrients. The production of endospores occurs through a complex system of proteins responsible for the reception and transmission of environmental signals within the cell while, at the same time, the arrest of vegetative growth or cell division occurs.

The spores are released by breaking the cell wall and remain in a quiescent phase waiting to develop, through the process of germination, in the presence of new favorable conditions. The formation of mature endospores occurs in 7-8 hours. Endospores are structures able to protect the inner nucleus thanks to a specific morphological organization (Fig. 4) and the presence of particular pigments: in case of exposure to radiations (e.g. UV rays), in fact, various types of pigments can be activated, useful to absorb harmful radiations and limit the damages associated with the formation of photoproducts that will be subsequently repaired. 

The formidable resistance of spores to specific stresses: UV light and temperature variation

Bacillus subtilis spores contain small acid-soluble proteins (SASP proteins) and they represent 20% of the total proteins present in spores. These proteins are localized at the core level and are essential to defend the genetic material from possible environmental damage. There are various types of SASP proteins and among them, the “multiple SASP” of alpha/beta-type have been shown to confer resistance to UV rays, heat, peroxides, and other sporicidal treatments.

Some studies, based on the study of mutant strains in which these proteins were not functional, have shown their action during exposure to UV radiation in different wavelengths. In addition, some temperature-related research has shown that sporulation at high temperatures can affect the water content of the spore nucleus, giving the forming spore a much higher heat-wet resistance. At the same time, it has been verified that the SASPs protein content is identical in every type of spore, regardless of the conditions under which it was formed. Therefore, all B. subtilis spores possess the same unique and extraordinary specific defense mechanism.

General defense in B. subtilis

The General Stress Response (GSP) involves the intervention of a specific factor defined as Sigma B to which is related the transcription of specific genes involved in the stress response. This type of response takes the specific name of “Sig.B-GSR” and gives to the microorganism a great resistance in stress conditions such as exposure to high or low temperatures, ethanol, salt, nitric oxide and vancomycin or stress situations in which the bacterium is looking for glucose, phosphate and/or oxygen.

The activation of sigma factor B is strongly regulated by the mechanism of “partner switching” in which phosphorylation and dephosphorylation processes are observed to be associated with the intervention of specific proteins such as RsbV (positive regulator) and RsbW (negative regulator). At the spore stage, however, the microorganism can activate other types of response. 

Disease cases and course

In the few cases of food poisoning by B. subtilis, the symptoms described are similar to the poisoning generated by Bacillus cereus bacteria. Among the most striking are: febrile state, diarrhea, nausea, and general sickness.

Therapy

The prognosis is always positive, and the treatment is purely symptomatic. Many patients do not need any medication since the pathogens and toxins are easily eradicated in a few hours.

References

• https://it.wikipedia.org/wiki/Bacillus_subtilis
• https://www.microbiologiaitalia.it/batteriologia/le-super-spore-di-bacillus-subtilis-una-vita-interminabile/
• https://academic.oup.com/femsec/article/13/3/159/655609
• Kunst, F., et al. The complete genome sequence of the Gram-positive bacterium Bacillus subtilis. Nature 390, 249–256 (1997). https://doi.org/10.1038/36786
• The Intestinal Life Cycle of Bacillus subtilis and Close Relatives. Nguyen K., et al. Journal of Bacteriology Mar 2006, 188 (7) 2692-2700; DOI: 10.1128/JB.188.7.2692-2700.2006
• Bacillus subtilis sporulation: regulation of gene expression and control of morphogenesis. J Errington. Microbiology and Molecular Biology Reviews Mar 1993, 57 (1) 1-33; 0146-0749/93/010001-33$02.00/0
• General Stress Response. CHESTER W. PRICE. Bacillus subtilis and Its Closest Relatives: from Genes to Cells. Ed. by A. L. Sonenshein et al., 2002 ASM Press, Washington, D.C.
• SigB-Dependent General Stress Response in Bacillus subtilis and Related Gram-Positive Bacteria. Michael Hecker et al. Annu. Rev. Microbiol. 2007. 61:215–36. DOI: 10.1146/annurev.micro.61.080706.093445
• Roles of small, acid-soluble spore proteins and core water content in survival of Bacillus subtilis spores exposed to environmental solar UV radiation. Moeller R et al.  Appl Environ Microbiol. 2009 Aug; 75(16):5202-8. doi: 10.1128/AEM.00789-09. Epub 2009 Jun 19. PMID: 19542328; PMCID: PMC2725452
• Analysis of the properties of spores of Bacillus subtilis prepared at different temperatures E. Melly et al. Journal of Applied Microbiology 2002, 92, 1105–1115. https://doi.org/10.1046/j.1365-2672.2002.01644.x
• Bacillus subtilis Spore Resistance to Simulated Mars Surface Conditions. Cortesao et al. Front. Microbiol., 26 February 2019. https://doi.org/10.3389/fmicb.2019.00333