Bacillus subtilis

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A Microbial Biorealm page on the genus Bacillus subtilis

Classification

Domain: Bacteria Division/phylum: Firmicutes Class: Bacilli Order: Bacillales Family: Bacillaceae Genus: Bacillus Species: B. safensis

Genus

Bacillus safensis

NCBI: Taxonomy

Description and significance

Bacillus safensis is a Gram-positive, spore-forming, and rod bacterium. originally isolated from a spacecraft in Florida and California.[1] Bacillus safensis could have possibly been transported to the planet Mars on spacecraft Opportunity and Spirit in 2004.[1] There are several known strains of this bacterium, all of which belong to the Firmicutes phylum of Bacteria.[1] This bacterium also belongs to the large, pervasive genus Bacillus. Bacillus safensis is an aerobic chemoheterotroph and is highly resistant to salt,[2] UV radiation, and gamma radiation.[3] Because of these features, Bacillus safensis is a powerful plant hormone producer. The bacteria is also a plant growth-promoting rhizobacteria, which enhances plant growth after root colonization.[2]

Bacillus safensis [sa.fen′sis. N.L. masc. adj. safensis arbitrarily derived from SAF (the spacecraft-assembly facility at the Jet Propulsion Laboratory, Pasadena, CA, USA), from where the organism was first isolated].

Cells are mesophilic, aerobic, chemoheterotrophic, Gram-positive, spore-forming rods that are motile by means of polar flagella. Cells are 0.5–0.7 μm in diameter and 1.0–1.2 μm in length. Growth occurs at 0–10 % (w/v) NaCl and at pH 5.6. Growth occurs at 10–50 °C (optimum, 30–37 °C) but not at 4 or 55 °C. Colonies are round, undulate, dull white, non-luminescent and have irregular margins on TSA plates incubated at 32 °C for 24 h. Oxidase, catalase, β-galactosidase, β-glucosidase, alkaline phosphatase, naphthol-AS-BI-phosphatase and esterase are produced, but H2S, indole, amylase, agarase, lecithinase, DNase, urease, leucine arylamidase, cystine arylamidase, valine arylamidase, trypsin, α-galactosidase, N-acetyl-β-glucosamidase, α-fucosidase, tryptophan deaminase, phenylalanine deaminase, arginine dihydrolase, lysine decarboxylase and ornithine decarboxylase are not. Cells do not reduce nitrate, but do hydrolyse gelatin, aesculin and RNA. Casein hydrolysis varies among strains. Voges–Proskauer test is positive. Growth occurs on agar plates supplemented with 1 % glycine and ox gall, but does not occur in 0.0001 % lysozyme broth. Negative for gas production from D-glucose. Acid is produced from D-glucose, glycerol, L-arabinose, ribose, D-xylose, galactose, fructose, mannose, inositol, mannitol, methyl α-D-mannopyranoside, methyl α-D-glucopyranoside, N-acetylglucosamine, amygdalin, arbutin, salicin, cellobiose, maltose, sucrose, trehalose, D-turanose and D-tagatose, but not from erythritol, D-arabinose, L-xylose, adonitol, methyl β-D-xylopyranoside, sorbose, rhamnose, dulcitol, sorbitol, inulin, melezitose, raffinose, starch, glycogen, xylitol, D-lyxose, D-fucose, L-fucose, D-arabitol, L-arabitol, gluconate, 2-ketogluconate or 5-ketogluconate. Reactions for lactose, melibiose and gentiobiose vary among strains. Citrate, malate, D-glucose, glycerol, L-arabinose, ribose, D-xylose, galactose, fructose, mannose, inositol, mannitol, methyl α-D-mannopyranoside, methyl α-D-glucopyranoside, N-acetylglucosamine, amygdalin, arbutin, salicin, cellobiose, maltose, sucrose, trehalose, gentiobiose, D-turanose, D-tagatose, gluconate, lactate, L-aspartate and L-glutamate are readily utilized as energy sources. Erythritol, D-arabinose, L-xylose, adonitol, methyl β-D-xylopyranoside, sorbose, dulcitol, sorbitol, inulin, lactose, melezitose, starch, glycogen, xylitol, D-lyxose, D-fucose, L-fucose, D-arabitol, L-arabitol, 2-ketogluconate, 5-ketogluconate, capric acid, adipic acid, phenylacetic acid, propionate and glycine are not utilized as energy sources. Rhamnose utilization varies among strains. The DNA G+C content is 41.0–41.4 mol%. The chain composition of the whole-cell fatty acids is primarily C15 : 0 iso, C15 : 0 anteiso, C17 : 0 iso and C17 : 0 anteiso.

Genome structure

The genome of Bacillus safensis strain FO-036b shows a GC content of 41.0-41.4 mol%.[1]

The Bacillus safensis VK genomic DNA was obtained from a 24-hr-old nutrient broth culture. Isolation of this strain was performed using a GenElute commercial DNA isolation kit, and whole-genome shotgun sequencing was carried out. Thirty-nine contigs, overlapping DNA fragments, greater in size than 200 base pairs were observed in strain VK.[2] This strain displays a G+C content of 46.1% in a circular chromosome of 3.68 Mb. 3,928 protein-coding sequences were identified, and 1,822 protein-coding sequences were appointed to one of the 457 RAST subsystems.[2] RAST, Rapid Annotation using Subsystem Technology, is a server that generates bacterial and archaeal genome annotations.[8] The genome also displays 73 tRNA genes.[2]Bacillus safensis VK genome sequence can be found in GenBank under the accession number AUPF00000000.[2] Another strain, DVL-43, can also be found in GenBank under the accession number KC156603.[3]

Cell structure and metabolism

Bacillus safensis is an aerobic, chemoheterotroph. Cell size ranges from 0.5-0.7 μm in diameter and 1.0-1.2 μm in length.[1] Bacteria are motile, and use polar flagella for locomotion. Cells are considered mesophillic, as they can grow in temperatures ranging between 10-50 °C.[1] Bacillus safensis FO-036b has an optimal temperature range from 30-37 °C, and cannot grow at 4 or 55 °C.[1] Bacillus safensis FO-036b prefers 0-10% salt, and a pH of 5.6. This strain was also found to produce spores that are resistant to hydrogen peroxide and UV radiation.[7]

Strain VK of Bacillus safensis is a salt-tolerant microbe, and can grow beyond the 0-10% salt range for the general microbial species.[2] This strain can grow in 14% NaCl, with a pH ranging from 4 to 8.[2] Strain VK also contains genes that encode for 1-aminocyclopropane-1-carboxylate deaminase enzyme.[2] This enzyme is able to generate 2-oxobutanoate and ammonia (NH3) by cleaving the precursor of plant hormone, ethylene 1-aminocyclopropane-1-carboxylate.[2] This enables the plant to tolerate salt, heavy metals, and polyaromatic hydrocarbons.[2] Because of these features, Bacillus safensis VK is a powerful plant hormone producer.

Ecology

Several isolates of the genus Bacillus are nearly identical to Bacillus pumilus. The group of isolates related to B. pumilus contains five related species: B. pumilus, B. safensis, B. stratosphericus, B. altitudinis, and B. aerophilus. These species are difficult to distinguish to due to their 99.5% similarity in their 16S rRNA gene sequence. Recently, scientists have discovered an alternate way to differentiate between these closely related species, especially B. pumilus and B. safensis.[10]

DNA gyrase is an important enzyme that introduces a negative supercoil to the DNA and is responsible for the biological processes in DNA replication and transcription.[10] DNA gyrase is made of two subunits, A and B. These subunits are denoted as gyrA and gyrB. The gyrB gene, subunit B protein, is a type II topoisomerase that is essential for DNA replication.[10] This gene is conserved among bacterial species. The rate of evolution at the molecular level deduced from gyrB related gene sequences can be determined at more accelerated rate compared to the 16S rRNA gene sequences.[10] These subunits have provided a way to phylogenetically distinguish between the diversity of species related to B. pumilus, which includes B. safensis. Strain B. safensis DSM19292 shares 90.2% gyrA sequence similarity with B. pumilus strain DSM 27.[10]

In 1952, a strain of B. pumilus was discovered in the DSMZ culture and labeled as strain DSM 354.[10] The strain was identified before B. safensis was discovered. In 2012, a gyrA sequence similarity was tested between the B. pumilus strain DSM 354 B. pumilus strain DSM 27, as well as against B. safensis strain DSM 19292.[10] Strain DSM 354 showed a 90.4% and 98% sequence similarity with B. pumilus strain DSM 27 and B. safensis strain DSM 19292, respectively.[10] These results indicated that DSM 354 may in fact be a B. safensis strain, instead of a B. pumilus strain. These results supported that gyrA sequences could be used to differentiate between closely related bacteria.[10]

Pathology

Bacillus subtilis bacteria are non-pathogenic. They can contaminate food, however, they seldom result in food poisoning. They are used on plants as a fungicide. They are also used on agricultural seeds, such as vegetable and soybean seeds, as a fungicide. The bacteria, colonized on root systems, compete with disease causing fungal organisms. Bacillus subtilis use as a fungicide fortunately does not affect humans (EMBL EBI). Some strains of Bacillus subtilis cause rots in potatoes. It grows in food that is non-acidic, and can cause ropiness in bread that is spoiled (Todar). Some strains related to Bacillus subtilis are capable of producing toxins for insects. Those strains can also be used for protecting crops as well. Bacillus thuringiensis, for example, is another bacterium in the same genus that is used for insect control (EMBL EBI).

Some Bacillus species can cause food poisoning, such as Bacillus cereus and Bacillus licheniformis. Bacillus cereus can result in two different kinds of intoxications. It can either cause nausea, vomiting, and abdominal cramps for 1-6 hours, or diarrhea and abdominal cramps for 8-16 hours. The food poisoning usually occurs from eating rice that is contaminated with Bacillus cereus (EMBL EBI).

Some Bacillus organisms can cause more severe illnesses. Bacillus anthracis, for example, causes Anthrax. It was the first bacterial organism that was known to cause disease in humans. Bacillus anthracis spores can survive for very long periods of time. Anthrax is very rare in humans, however it is more common in animals. The disease often begins with a very high fever and chest pain, and can be fatal if untreated (EMBL EBI).

Application to Biotechnology

Thirteen strains of a novel spore-forming, Gram-positive, mesophilic heterotrophic bacterium were isolated from spacecraft surfaces (Mars Odyssey Orbiter) and assembly-facility surfaces at the Jet Propulsion Laboratory in California and the Kennedy Space Center in Florida. Phylogenetic analysis of 16S rRNA gene sequences has placed these novel isolates within the genus Bacillus, the greatest sequence similarity (99.9 %) being found with Bacillus pumilus. However, these isolates share a mere 91.2 % gyrB sequence similarity with Bacillus pumilus, rendering their 16S rRNA gene-derived relatedness suspect. Furthermore, DNA–DNA hybridization showed only 54–66 % DNA relatedness between the novel isolates and strains of B. pumilus. rep-PCR fingerprinting and previously reported matrix-assisted laser desorption/ionization time-of-flight mass spectrometry protein profiling clearly distinguished these isolates from B. pumilus. Phenotypic analyses also showed some differentiation between the two genotypic groups, although the fatty acid compositions were almost identical. The polyphasic taxonomic studies revealed distinct clustering of the tested strains into two distinct species. On the basis of phenotypic characteristics and the results of phylogenetic analyses of 16S rRNA and gyrB gene sequences, repetitive element primer-PCR fingerprinting and DNA–DNA hybridization, the 13 isolates represent a novel species of the genus Bacillus, for which the name Bacillus safensis sp. nov. is proposed.

References

[1] Ara, K., et al. "Bacillus minimum genome factory: effective utilization of microbial genome information." Biotechnol. Appl. Biochem.. 2007 March; 46(Pt 3):169-78.

[2] Bandow, J.E., H. Br�tz, M. Hecker. "Bacillus subtilis Tolerance of Moderate Concentrations of Rifampin Involves the ?B-Dependent General and Multiple Stress Response". Journal of Bacteriology. 2002 January; 184(2): 459�467.

[3] Entrez Genome Project, NCBI

[4] European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL EBI).

[5] Feng, J., et al. "Effect of Fermented Soybean Meal on Intestinal Morphology and Digestive Enzyme Activities in Weaned Piglets". Digestive Diseases and Sciences. 2007 April.

[6] Graumann, P. 2007. Bacillus: Cellular and Molecular Biology. Caister Academic press. ISBN 978-1-904455-12-7.

[7] Jamil, B., et al. "Isolation of Bacillus subtilis MH-4 from Soil and its Potential of Polypeptidic Antibiotic Production". Pak J Pharm Sci. 2007 January; 20(1):26-31.

[8] Kobayashi, K., et al. "Essential Bacillus subtilis genes". Proc Natl Acad Sci U S A. 2003 April 15; 100(8): 4678�4683.

[9] Kunst, F., et al. "The complete genome sequence of the Gram-positive bacterium Bacillus subtilis". Nature. 1997 November; 390, 249-256.

[10] Liu, NJ., RJ. Dutton, K. Pogliano. "Evidence that the SpoIIIE DNA Translocase Participates in Membrane Fusion During Cytokinesis and Engulfment". Mol Microbiol 2006 February;59(4):1097-113.

[11] Marino, M., et al. "Modulation of Anaerobic Energy Metabolism of Bacillus subtilis by arfM (ywiD)". J Bacteriol. 2001 December; 183(23): 6815�6821.

[12] Morikawa, M. "Beneficial Biofilm Formation by Industrial Bacteria Bacillus subtilis and Related Species". Journal of Bioscience and Bioengineering. 2006; Vol.101, No.1, 1-8.

[13] Nakano, M.M., P. Zuber. "Anaerobic Growth of a 'Strict Aerobe' (Bacillus subtilis)". Annual Review of Microbiology. 1998 October; Vol. 52: 165-190.

[14] Perez, A.R., A. Abanes-De Mello, K. Pogliano. "SpoIIB Localizes to Active Sites of Septal Biogenesis and Spatially Regulates Septal Thinning during Engulfment in Bacillus subtilis". Journal of Bacteriology. 2000 February; 182(4): 1096�1108.

Schaechter, M., J.L. Ingraham, F.C. Neidhardt. Microbe. (ASM Press, Washington, DC, 2006).

[15] Setlow, P. "Spores of Bacillus subtilis:Their Resistance to and Killing by Radiation, Heat, and Chemicals". Journal of Applied Microbiology. 2006 September; 101(3), 514-525.

[16] The Institute for Genome Research, Comprehensive Microbial Resource (TIGR CMR).

[17] Todar, K. "Todar's Online Textbook of Bacteriology".

Edited by Margo Ucar, student of Rachel Larsen and Kit Pogliano

Edited by a student of M Glogowski at Loyola University