Small, Acid-soluble Spore Proteins Research Articles

Functional Analysis of Stress Resistance of Bacillus cereus SCL10 Strain Based on Whole-Genome Sequencing.

A Gram-positive, rod-shaped, aerobic, motile, and spore-forming bacterium, designated SCL10, was isolated from Acaudina molpadioides exposure to Co-60 radiation. In this study, whole-genome sequencing was performed to identify the strain as Bacillus cereus and functional characterization, with a focus on stress resistance. The genome of the B. cereus SCL10 strain was sequenced and assembled, revealing a size of 4,979,182 bp and 5167 coding genes. The genes involved in biological functions were annotated by using the GO, COG, KEGG, NR, and Swiss-Prot databases. The results showed that genes related to alkyl hydroperoxide reductase (ahpC, ahpF), DNA-binding proteins from starved cells (dps), spore and biofilm formation (spoVG, spo0A, gerP), cold shock-like protein (cspC, cspE), ATP-dependent chaperone (clpB), and photolyase, small, acid-soluble spore protein (SASP) and DNA repair protein (recA, radD) could explain the stress resistance. These findings suggest that antioxidant activity, sporulation, biofilm formation, and DNA protection may be considered as the main resistance mechanisms under exposure to radiation in the B. cereus SCL10 strain.

Proteomics-based analysis of the stress response of Bacillus cereus spores under ultrasound and electrolyzed water treatment

Ultrasound is a green nonthermal technology with promising applications in microbial inactivation. Electrolyzed water has been investigated and found to have a synergistic inactivation effect of ultrasound on spores. This study used a data-independent-acquisition method to analyze the stress response of Bacillus cereus spores following ultrasound combined with electrolyzed water treatment. We identified 197 differentially expressed proteins under ultrasound combined with an electrolyzed water treatment for which the ratio in the metabolic pathway was the highest. Spores downregulated key proteins in energy metabolic and transportation pathways, in particular in phosphotransferase systems and ATP synthase under ultrasound, electrolyzed water, and combined stress. The results of this study revealed that the key proteins in intracellular metabolism decreased after ultrasound treatment, and the expression of small acid-soluble spore protein and cell wall biosynthesis protein increased. Meanwhile, DNA integration, recombination, and inversion protein and small acid-soluble spore protein were upregulated after electrolyzed water treatment. In general, the spores exhibited stress resistance under external stress. The inactivation of spores by further stress was reduced, which we called “cross-protection.”

New Thoughts on an Old Topic: Secrets of Bacterial Spore Resistance Slowly Being Revealed.

The quest for bacterial survival is exemplified by spores formed by some Firmicutes members. They turn up everywhere one looks, and their ubiquity reflects adaptations to the stresses bacteria face. Spores are impactful in public health, food safety, and biowarfare. Heat resistance is the hallmark of spores and is countered principally by a mineralized gel-like protoplast, termed the spore core, with reduced water which minimizes macromolecular movement/denaturation/aggregation. Dry heat, however, introduces mutations into spore DNA. Spores have countermeasures to extreme conditions that are multifactorial, but the fact that spore DNA is in a crystalline-like nucleoid in the spore core, likely due to DNA saturation with small acid-soluble spore proteins (SASPs), suggests that reduced macromolecular motion is also critical in spore dry heat resistance. SASPs are also central in the radiation resistance characteristic of spores, where the contributions of four spore features-SASP; Ca2+, with pyridine-2,6-dicarboxylic acid (CaDPA); photoproduct lyase; and low water content-minimize DNA damage. Notably, the spore environment steers UV photochemistry toward a product that germinated spores can repair without significant mutagenesis. This resistance extends to chemicals and macromolecules that could damage spores. Macromolecules are excluded by the spore coat which impedes the passage of moieties of ≥10 kDa. Additionally, damaging chemicals may be degraded or neutralized by coat enzymes/proteins. However, the principal protective mechanism here is the inner membrane, a compressed structure lacking lipid fluidity and presenting a barrier to the diffusion of chemicals into the spore core; SASP saturation of DNA also protects against genotoxic chemicals. Spores are also resistant to other stresses, including high pressure and abrasion. Regardless, overarching mechanisms associated with resistance seem to revolve around reduced molecular motion, a fine balance between rigidity and flexibility, and perhaps efficient repair.

Targetable nano-delivery vehicles to deliver anti-bacterial small acid-soluble spore protein (SASP) genes.

Interest in phage-based therapeutics is increasing, at least in part due to the need for new treatment options for infections caused by antibiotic-resistant bacteria. It is possible to use wild-type (WT) phages to treat bacterial infections, but it is also possible to modify WT phages to generate therapeutics with improved features. Here, we will discuss features of Phico Therapeutics' SASPject technology, which modifies phages for use as targetable nano-delivery vehicles (NDV), to introduce antibacterial Small Acid Soluble Spore Protein (SASP) genes into specific target bacteria.

Engineered Bacteriophage as a Delivery Vehicle for Antibacterial Protein, SASP.

The difficulties in developing novel classes of antibacterials is leading to a resurgence of interest in bacteriophages as therapeutic agents, and in particular engineered phages that can be optimally designed. Here, pre-clinical microbiology assessment is presented of a Staphylococcus aureus phage engineered to deliver a gene encoding an antibacterial small acid soluble spore protein (SASP) and further, rendered non-lytic to give product SASPject PT1.2. PT1.2 has been developed initially for nasal decolonisation of S. aureus, including methicillin-resistant S. aureus. Time-kill curve assays were conducted with PT1.2 against a range of staphylococcal species, and serial passaging experiments were conducted to investigate the potential for resistance to develop. SASPject PT1.2 demonstrates activity against 100% of 225 geographically diverse S. aureus isolates, exquisite specificity for S. aureus, and a rapid speed of kill. The kinetics of S. aureus/PT1.2 interaction is examined together with demonstrating that PT1.2 activity is unaffected by the presence of human serum albumin. SASPject PT1.2 shows a low propensity for resistance to develop with no consistent shift in sensitivity in S. aureus cells passaged for up to 42 days. SASPject PT1.2 shows promise as a novel first-in-class antibacterial agent and demonstrates potential for the SASPject platform.

Transcriptional response of Bacillus megaterium FDU301 to PEG200-mediated arid stress

BackgroundFor microorganisms on a paper surface, the lack of water is one of the most important stress factors. A strain of Bacillus megaterium FDU301 was isolated from plaques on a paper surface using culture medium with polyethylene glycol 200 (PEG200) to simulate an arid condition. Global transcriptomic analysis of B. megaterium FDU301 grown under normal and simulated arid conditions was performed via RNA-seq technology to identify genes involved in arid stress adaptation.ResultsThe transcriptome of B. megaterium FDU301 grown in LB medium under arid (15% PEG200 (w/w)) and normal conditions were compared. A total of 2941 genes were differentially expressed, including 1422 genes upregulated and 1519 genes downregulated under arid conditions. Oxidative stress-responsive regulatory genes perR, fur, and tipA were significantly upregulated, along with DNA protecting protein (dps), and catalase (katE). Genes related to Fe2+ uptake (feoB), sporulation stage II (spoIIB, spoIIE, spoIIGA), small acid-soluble spore protein (sspD), and biosynthesis of compatible solute ectoine (ectB, ectA) were also highly expressed to various degrees. Oxidative phosphorylation-related genes (atpB, atpE, atpF, atpH, atpA, atpG, atpD, atpC) and glycolysis-related genes (pgk, tpiA, frmA) were significantly downregulated.ConclusionThis is the first report about transcriptomic analysis of a B. megaterium to explore the mechanism of arid resistance. Major changes in transcription were seen in the arid condition simulated by PEG200 (15%), with the most important one being genes related to oxidative stress. The results showed a complex mechanism for the bacteria to adapt to arid stress.

A transcriptional response of Clostridium beijerinckii NRRL B-598 to a butanol shock

BackgroundOne of the main obstacles preventing solventogenic clostridia from achieving higher yields in biofuel production is the toxicity of produced solvents. Unfortunately, regulatory mechanisms responsible for the shock response are poorly described on the transcriptomic level. Although the strain Clostridium beijerinckii NRRL B-598, a promising butanol producer, has been studied under different conditions in the past, its transcriptional response to a shock caused by butanol in the cultivation medium remains unknown.ResultsIn this paper, we present a transcriptional response of the strain during a butanol challenge, caused by the addition of butanol to the cultivation medium at the very end of the acidogenic phase, using RNA-Seq. We resequenced and reassembled the genome sequence of the strain and prepared novel genome and gene ontology annotation to provide the most accurate results. When compared to samples under standard cultivation conditions, samples gathered during butanol shock represented a well-distinguished group. Using reference samples gathered directly before the addition of butanol, we identified genes that were differentially expressed in butanol challenge samples. We determined clusters of 293 down-regulated and 301 up-regulated genes whose expression was affected by the cultivation conditions. Enriched term “RNA binding” among down-regulated genes corresponded to the downturn of translation and the cluster contained a group of small acid-soluble spore proteins. This explained phenotype of the culture that had not sporulated. On the other hand, up-regulated genes were characterized by the term “protein binding” which corresponded to activation of heat-shock proteins that were identified within this cluster.ConclusionsWe provided an overall transcriptional response of the strain C. beijerinckii NRRL B-598 to butanol shock, supplemented by auxiliary technologies, including high-pressure liquid chromatography and flow cytometry, to capture the corresponding phenotypic response. We identified genes whose regulation was affected by the addition of butanol to the cultivation medium and inferred related molecular functions that were significantly influenced. Additionally, using high-quality genome assembly and custom-made gene ontology annotation, we demonstrated that this settled terminology, widely used for the analysis of model organisms, could also be applied to non-model organisms and for research in the field of biofuels.

Disruption of SpoIIID decreases sporulation, increases extracellular proteolytic activity and virulence in Bacillus anthracis

Endospores are important for maintenance of the B. anthracis lifecycle and necessary for its effective spread between hosts. Our experiments with B. anthracis showed that disruption of SpoIIID results in a spore formation defect, as determined by heat resistance assays and microscopic assessment. We further found complete engulfment by the ΔspoIIID mutant strain by membrane morphology staining but no synthesis of the clarity coat and exosporium by transmission electron microscopy. Reduced transcription and expression of small acid-soluble spore protein(sasP-2) and the spore development associated genes (σK, gerE and cotE) in the mother cell were found in the ΔspoIIID strain, suggesting that the spore formation defect in B. anthracis A16R is related to decreased transcription and expression of these genes. Extracellular protease and virulence enhancement in the ΔspoIIID strain may be related to the elevation of metalloproteinases (TasA and Camelysin) levels. Our findings pave the way for further research on the regulation network of sporulation, survival and virulence in these two morphological forms of B. anthracis.

Bacillus subtilis Spore Resistance to Simulated Mars Surface Conditions.

In a Mars exploration scenario, knowing if and how highly resistant Bacillus subtilis spores would survive on the Martian surface is crucial to design planetary protection measures and avoid false positives in life-detection experiments. Therefore, in this study a systematic screening was performed to determine whether B. subtilis spores could survive an average day on Mars. For that, spores from two comprehensive sets of isogenic B. subtilis mutant strains, defective in DNA protection or repair genes, were exposed to 24 h of simulated Martian atmospheric environment with or without 8 h of Martian UV radiation [M(+)UV and M(-)UV, respectively]. When exposed to M(+)UV, spore survival was dependent on: (1) core dehydration maintenance, (2) protection of DNA by α/β-type small acid soluble proteins (SASP), and (3) removal and repair of the major UV photoproduct (SP) in spore DNA. In turn, when exposed to M(-)UV, spore survival was mainly dependent on protection by the multilayered spore coat, and DNA double-strand breaks represent the main lesion accumulated. Bacillus subtilis spores were able to survive for at least a limited time in a simulated Martian environment, both with or without solar UV radiation. Moreover, M(-)UV-treated spores exhibited survival rates significantly higher than the M(+)UV-treated spores. This suggests that on a real Martian surface, radiation shielding of spores (e.g., by dust, rocks, or spacecraft surface irregularities) might significantly extend survival rates. Mutagenesis were strongly dependent on the functionality of all structural components with small acid-soluble spore proteins, coat layers and dipicolinic acid as key protectants and efficiency DNA damage removal by AP endonucleases (ExoA and Nfo), non-homologous end joining (NHEJ), mismatch repair (MMR) and error-prone translesion synthesis (TLS). Thus, future efforts should focus on: (1) determining the DNA damage in wild-type spores exposed to M(+/-)UV and (2) assessing spore survival and viability with shielding of spores via Mars regolith and other relevant materials.

Construction of Bacillus thuringiensis Simulant Strains Suitable for Environmental Release

For a surrogate bacterium to be used in outdoor studies, it is important to consider environmental and human safety and ease of detection. Recently, Bacillus thuringiensis, a popular bioinsecticide bacterium, has been gaining attention as a surrogate bacterium for use in biodefense. In this study, we constructed simulant strains of B. thuringiensis with enhanced characteristics for environmental studies. Through transposon mutagenesis, pigment genes were inserted into the chromosome, producing yellow-colored colonies for easy detection. To prevent persistence of spores in the environment, a genetic circuit was designed to produce a spore without sporulation capability. Two loxP sites were inserted, one on each side of the spo0A gene, which encodes a sporulation master regulator, and a sporulation-dependent Cre expression cassette was inserted into the chromosome. This genetic circuit successfully deleted spo0A during sporulation, producing spores that lacked the spo0A gene. In addition, two major α/β-type small acid-soluble spore protein (SASP) genes, predicted by synteny analysis, were deleted. The spores of the mutant strain showed increased UV-C sensitivity and quickly lost viability when tested in a solar simulator. When the spores of the mutant strain were administered to the lungs of BALB/c mice, cells were quickly removed from the body, suggesting enhanced in vivo safety. All strains constructed in this study contain no antibiotic resistance markers and all heterologous genes were inserted into the chromosome, which are useful features for simulants to be released into the environment.IMPORTANCEB. thuringiensis has recently been receiving increasing attention as a good spore simulant in biodefense research. However, few studies were done to properly address many important features of B. thuringiensis as a simulant in environmental studies. Since spores can persist in the environment for years after release, environmental contamination is a big problem, especially when genetically engineered strains are used. To solve these problems, we report here the development of B. thuringiensis simulant strains that are capable of forming yellow colonies for easy detection, incapable of forming spores more than once due to a genetic circuit, and lacking in two major SASP genes. The genetic circuit to produce a spore without sporulation capability, together with the deletion of SASP genes, ensures the environmental and human safety of the simulant strains developed in this study. All of these features will allow wider use of B. thuringiensis as a simulant for Bacillus anthracis in environmental release studies.

Mechanism of Sporicidal Activity for the Synergistic Combination of Peracetic Acid and Hydrogen Peroxide.

There is still great interest in controlling bacterial endospores. The use of chemical disinfectants and, notably, oxidizing agents to sterilize medical devices is increasing. With this in mind, hydrogen peroxide (H2O2) and peracetic acid (PAA) have been used in combination, but until now there has been no explanation for the observed increase in sporicidal activity. This study provides information on the mechanism of synergistic interaction of PAA and H2O2 against bacterial spores. We performed investigations of the efficacies of different combinations, including pretreatments with the two oxidizers, against wild-type spores and a range of spore mutants deficient in the spore coat or small acid-soluble spore proteins. The concentrations of the two biocides were also measured in the reaction vessels, enabling the assessment of any shift from H2O2 to PAA formation. This study confirmed the synergistic activity of the combination of H2O2 and PAA. However, we observed that the sporicidal activity of the combination is largely due to PAA and not H2O2. Furthermore, we observed that the synergistic combination was based on H2O2 compromising the spore coat, which was the main spore resistance factor, likely allowing better penetration of PAA and resulting in the increased sporicidal activity.

Small acid-soluble spore proteins of Clostridium acetobutylicum are able to protect DNA in vitro and are specifically cleaved by germination protease GPR and spore protease YyaC.

Small acid-soluble proteins (SASPs) play an important role in protection of DNA in dormant bacterial endospores against damage by heat, UV radiation or enzymic degradation. In the genome of the strict anaerobe Clostridium acetobutylicum, five genes encoding SASPs have been annotated and here a further sixth candidate is suggested. The ssp genes are expressed in parallel dependent upon Spo0A, a master regulator of sporulation. Analysis of the transcription start points revealed a σG or a σF consensus promoter upstream of each ssp gene, confirming a forespore-specific gene expression. SASPs were termed SspA (Cac2365), SspB (Cac1522), SspD (Cac1620), SspF (Cac2372), SspH (Cac1663) and Tlp (Cac1487). Here it is shown that with the exception of Tlp, every purified recombinant SASP is able to bind DNA in vitro thereby protecting it against enzymic degradation by DNase I. Moreover, SspB and SspD were specifically cleaved by the two germination-specific proteases GPR (Cac1275) and YyaC (Cac2857), which were overexpressed in Escherichia coli and activated by an autocleavage reaction. Thus, for the first time to our knowledge, GPR-like activity and SASP specificity could be demonstrated for a YyaC-like protein. Collectively, the results assign SspA, SspB, SspD, SspF and SspH of C. acetobutylicum as members of α/β-type SASPs, whereas Tlp seems to be a non-DNA-binding spore protein of unknown function. In acetic acid-extracted proteins of dormant spores of C. acetobutylicum, SspA was identified almost exclusively, indicating its dominant biological role as a major α/β-type SASP in vivo.

Resistance to and killing by the sporicidal microbicide peracetic acid.

To elucidate the mechanisms of spore resistance to and killing by the oxidizing microbicide peracetic acid (PAA). Mutants of Bacillus subtilis lacking specific spore structures were used to identify resistance properties in spores and to understand the mechanism of action of PAA. We also assessed the effect of PAA treatment on a number of spore properties including heat tolerance, membrane integrity and germination. The spore coat is essential for spore PAA resistance as spores with defective coats were greatly sensitized to PAA treatment. Small acid-soluble spore proteins apparently provide no protection against PAA. Defects in spore germination, specifically in germination via the GerB and GerK but not the GerA germination receptors, as well as leakage of internal components suggest that PAA is active at the spore inner membrane. It is therefore likely that the inner membrane is the major site of PAA's sporicidal activity. PAA treatment targets the spore membrane, with some of its activity directed specifically against the GerB and GerK germination receptors.

Differentiation of Bacillus pumilus and Bacillus safensis using MALDI-TOF-MS.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) despite being increasingly used as a method for microbial identification, still present limitations in which concerns the differentiation of closely related species. Bacillus pumillus and Bacillus safensis, are species of biotechnological and pharmaceutical significance, difficult to differentiate by conventional methodologies. In this study, using a well-characterized collection of B. pumillus and B. safensis isolates, we demonstrated the suitability of MALDI-TOF-MS combined with chemometrics to accurately and rapidly identify them. Moreover, characteristic species-specific ion masses were tentatively assigned, using UniProtKB/Swiss-Prot and UniProtKB/TrEMBL databases and primary literature. Delineation of B. pumilus (ions at m/z 5271 and 6122) and B. safensis (ions at m/z 5288, 5568 and 6413) species were supported by a congruent characteristic protein pattern. Moreover, using a chemometric approach, the score plot created by partial least square discriminant analysis (PLSDA) of mass spectra demonstrated the presence of two individualized clusters, each one enclosing isolates belonging to a species-specific spectral group. The generated pool of species-specific proteins comprised mostly ribosomal and SASPs proteins. Therefore, in B. pumilus the specific ion at m/z 5271 was associated with a small acid-soluble spore protein (SASP O) or with 50S protein L35, whereas in B. safensis specific ions at m/z 5288 and 5568 were associated with SASP J and P, respectively, and an ion at m/z 6413 with 50S protein L32. Thus, the resulting unique protein profile combined with chemometric analysis, proved to be valuable tools for B. pumilus and B. safensis discrimination, allowing their reliable, reproducible and rapid identification.

Interaction of near-infrared femtosecond laser pulses with biological materials in water

We study the effects of the interaction of 40-fs Ti-sapphire laser radiation at 800 nm with biological materials—proteins or intact Bacillus spore, dissolved or suspended in pure water, respectively. The estimated laser intensity at the target is 10 13 W/cm 2 . On the molecular level, oxidation of solvent-accessible parts of proteins has been observed even after a single femtosecond laser pulse, as demonstrated by mass spectrometry. A remarkable morphological effect of the femtosecond laser radiation is the complete disintegration of extremely refractive cells such as bacterial spores, evidenced in scanning electron micrographs. After 500 laser pulses, all suspended spores in the irradiated volume are completely destroyed, which makes them nonviable. Characteristic spore biomolecules, e.g., small acid-soluble spore proteins, are extensively oxidized after several laser pulses. In comparative studies, no effects have been observed when irradiating the same samples with 10-ns laser pulses at the same laser wavelength and fluence. We demonstrate that the laser power density (irradiance), resulting in different amounts of total deposited energy, determines the types of effects for femtosecond laser interactions with biological matter.

Resistance of Bacillus subtilis Spore DNA to Lethal Ionizing Radiation Damage Relies Primarily on Spore Core Components and DNA Repair, with Minor Effects of Oxygen Radical Detoxification

The roles of various core components, including α/β/γ-type small acid-soluble spore proteins (SASP), dipicolinic acid (DPA), core water content, and DNA repair by apurinic/apyrimidinic (AP) endonucleases or nonhomologous end joining (NHEJ), in Bacillus subtilis spore resistance to different types of ionizing radiation including X rays, protons, and high-energy charged iron ions have been studied. Spores deficient in DNA repair by NHEJ or AP endonucleases, the oxidative stress response, or protection by major α/β-type SASP, DPA, and decreased core water content were significantly more sensitive to ionizing radiation than wild-type spores, with highest sensitivity to high-energy-charged iron ions. DNA repair via NHEJ and AP endonucleases appears to be the most important mechanism for spore resistance to ionizing radiation, whereas oxygen radical detoxification via the MrgA-mediated oxidative stress response or KatX catalase activity plays only a very minor role. Synergistic radioprotective effects of α/β-type but not γ-type SASP were also identified, indicating that α/β-type SASP's binding to spore DNA is important in preventing DNA damage due to reactive oxygen species generated by ionizing radiation.

Multifactorial Resistance ofBacillus subtilisSpores to High-Energy Proton Radiation: Role of Spore Structural Components and the Homologous Recombination and Non-Homologous End Joining DNA Repair Pathways

The space environment contains high-energy charged particles (e.g., protons, neutrons, electrons, α-particles, heavy ions) emitted by the Sun and galactic sources or trapped in the radiation belts. Protons constitute the majority (87%) of high-energy charged particles. Spores of Bacillus species are one of the model systems used for astro- and radiobiological studies. In this study, spores of different Bacillus subtilis strains were used to study the effects of high energetic proton irradiation on spore survival. Spores of the wild-type B. subtilis strain [mutants deficient in the homologous recombination (HR) and non-homologous end joining (NHEJ) DNA repair pathways and mutants deficient in various spore structural components such as dipicolinic acid (DPA), α/β-type small, acid-soluble spore protein (SASP) formation, spore coats, pigmentation, or spore core water content] were irradiated as air-dried multilayers on spacecraft-qualified aluminum coupons with 218 MeV protons [with a linear energy transfer (LET) of 0.4 keV/μm] to various final doses up to 2500 Gy. Spores deficient in NHEJ- and HR-mediated DNA repair were significantly more sensitive to proton radiation than wild-type spores, indicating that both HR and NHEJ DNA repair pathways are needed for spore survival. Spores lacking DPA, α/β-type SASP, or with increased core water content were also significantly more sensitive to proton radiation, whereas the resistance of spores lacking pigmentation or spore coats was essentially identical to that of the wild-type spores. Our results indicate that α/β-type SASP, core water content, and DPA play an important role in spore resistance to high-energy proton irradiation, suggesting their essential function as radioprotectants of the spore interior.

Protective Role of Spore Structural Components in Determining Bacillus subtilis Spore Resistance to Simulated Mars Surface Conditions

Spores of wild-type and mutant Bacillus subtilis strains lacking various structural components were exposed to simulated Martian atmospheric and UV irradiation conditions. Spore survival and mutagenesis were strongly dependent on the functionality of all of the structural components, with small acid-soluble spore proteins, coat layers, and dipicolinic acid as key protectants.

Genomic determinants of sporulation in Bacilli and Clostridia: towards the minimal set of sporulation‐specific genes

Three classes of low-G+C Gram-positive bacteria (Firmicutes), Bacilli, Clostridia and Negativicutes, include numerous members that are capable of producing heat-resistant endospores. Spore-forming firmicutes include many environmentally important organisms, such as insect pathogens and cellulose-degrading industrial strains, as well as human pathogens responsible for such diseases as anthrax, botulism, gas gangrene and tetanus. In the best-studied model organism Bacillus subtilis, sporulation involves over 500 genes, many of which are conserved among other bacilli and clostridia. This work aimed to define the genomic requirements for sporulation through an analysis of the presence of sporulation genes in various firmicutes, including those with smaller genomes than B. subtilis. Cultivable spore-formers were found to have genomes larger than 2300 kb and encompass over 2150 protein-coding genes of which 60 are orthologues of genes that are apparently essential for sporulation in B. subtilis. Clostridial spore-formers lack, among others, spoIIB, sda, spoVID and safA genes and have non-orthologous displacements of spoIIQ and spoIVFA, suggesting substantial differences between bacilli and clostridia in the engulfment and spore coat formation steps. Many B. subtilis sporulation genes, particularly those encoding small acid-soluble spore proteins and spore coat proteins, were found only in the family Bacillaceae, or even in a subset of Bacillus spp. Phylogenetic profiles of sporulation genes, compiled in this work, confirm the presence of a common sporulation gene core, but also illuminate the diversity of the sporulation processes within various lineages. These profiles should help further experimental studies of uncharacterized widespread sporulation genes, which would ultimately allow delineation of the minimal set(s) of sporulation-specific genes in Bacilli and Clostridia.

Modeling of small acid-soluble spore protein (SASP) with Swiss model software

Modeling of small acid-soluble spore protein (SASP) with Swiss model software