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BACKGROUND: Bacillus velezensis 83 is a biocontrol agent and plant growth promoting bacteria that produces antifungal compounds (surfactin and bacilomycin D), plant growth stimulation metabolites such as acetoin and butanediol and the polymer poly‐gamma‐glutamic acid (γ‐PGA), which is essential for bacterial plant colonization. Bacillus sporulation is an important trait in order to obtain long shelf‐life formulations for biocontrol purposes. γ‐PGA production and sporulation are closed‐related processes controlled by glucose availability and modulated by quorum sensing. High spore production (and its recovery) in B. velezensis 83 cultures is limited by the high viscosity oxygen‐limited broths resulting from γ‐PGA accumulation. This work studied the influence of acidic growing conditions and glucose specific uptake rate on growth, quorum sensing‐related metabolites production and sporulation of B. velezensis 83. RESULTS: In batch cultures developed at pH = 5, no bacterial sporulation was detected and quorum sensing related metabolites as γ‐PGA (not detected) and surfactin and bacillomycin were significantly decreased. Instead, acetoin and butanediol were the main products. In contrast, in continuous cultures developed at pH = 5, spores were detected at D = 0.06 h−1. The results indicate that sporulation at pH = 5 is not inhibited by affectations on quorum sensing signals but could be due to a low availability/transport of energy sources (i.e., glucose or acetoin/butanediol) or an affectation on Rap‐Phr system. CONCLUSION: The findings in this study allow to establish a two‐stage process for Bacillus velezensis 83 spore production: bacterial growth at pH = 5 and sporulation at pH = 6.8. © 2023 Society of Chemical Industry (SCI). [ABSTRACT FROM AUTHOR]

Introduction. The study of the ability of Bacillus anthracis strains with different phenotypic properties to spore germination, reproduction and sporulation on a medium based on an aqueous soil extract can help assess the significance of these processes in the formation and maintenance of soil anthrax foci. Aim. The analysis of individual characteristics of the development of a vegetative culture of anthrax pathogen strains with different phenotypes in a soil medium model. Materials and methods. On a group of anthrax microbe strains with different plasmid composition and virulence, the possibility of spore germination, reproduction of bacilli and, at least in some of them, productive spore formation on the soil medium was studied. Results. Three variants of culture development of B. anthracis strains were identified: 1 spores remain intact, not germinating; 2 after germination of spores, bacilli are formed, which multiply with different intensity, passing into involutional forms without spore formation; 3 the passage of a complete physiological cycle "sporebacillusspore". The presence of 2% blood in the soil environment contributed to the germination of spores and reproduction of the culture, but inhibited the process of sporulation during the observation period of 3 days. No correlation was found between a certain phenotype of the studied strains of B. anthracis and the ability to germinate and vegetate on soil media. Conclusion. The data obtained that only 17% of CFU gives rise to the formation of colonies on the soil medium suggest the heterogeneity of the properties of the population of the studied strains. Isolation of such cultures and their further detailed study will make it possible to identify the most significant complexes of biological properties for the realization of a complete physiological cycle under soil-simulating conditions.

Clostridium saccharoperbutylacetonicum strain N1-4 (ATCC13564) is a butanol-producing strain suitable for application to butanol production from cellulosic materials by co-culture with cellulolytic and thermophilic species, such as Hungateiclostridium thermocellum (synonym: Clostridium thermocellum). The optimal temperature for butanol production by strain N1-4 is 30 °C, and the strain is sensitive to a high culture temperature of 37 °C. Given that spore formation is observed at high frequency when strain N1-4 is cultivated at 37 °C, we assumed in a previous study that the initiation of sporulation is related to a decrease in butanol production. Therefore, to investigate the relationship between butanol production and spore formation, we generated strain N1-4 isolates in which genes related to spore formation were disrupted. The sporulation-related gene disruptants of spo0A , sigE , sigG , and sigK lost the ability to produce heat-resistant spores, irrespective of the culture temperature. Among the gene disruptants produced, only the spo0A disruptant lost butanol-producing ability when cultivated at 30 °C. Interestingly, the sigE disruptant maintained butanol productivity similar to that observed at 30 °C, even when cultivated at 37 °C. In addition, the sigE disruptant successfully produced butanol from Avicel cellulose by co-culture with H. thermocellum at a fermentation temperature of 37 °C. [ABSTRACT FROM AUTHOR]

Clostridioides difficile (C. difficile) is a Gram-positive, spore-forming, toxin-producing, obligate anaerobic bacterium. C. difficile infection (CDI) is the leading cause of healthcare-associated infective diarrhoea. The infection is mediated by the spore, a metabolically inactive form of C. difficile. The spore coat acts as a physical barrier to defend against chemical insults from hosts and natural environments. The composition of spore coat has already been revealed; therefore, the interactive networks of spore coat proteins and the dynamic process of coat assembly are the keys to design strategies to control and cure CDI. This review gives a brief discussion of the signal processing and transcriptional regulation of C. difficile sporulation initiation. Following the discussion, the spore formation is also introduced. Finally, this review mainly focuses on the spore coat assembly, a poorly understood process in C. difficile, and important proteins that have been studied.

Botrytis cinerea is the pathogen of many crops gray mold disease and its conidia is the major carrier for disease epidemic. In this work, the carbon utilization and conidia formation of B. cinerea were analyzed by phenomic technology. In addition, the carbon sources for conidia production of this pathogen under five different fungicides (carbendazim, propiconazole, pyrimethanil, iprodione and prochloraz) were also tested. These results showed that about 92 carbon sources could be metabolized by B. cinerea, including carbohydrates, amino acids, and others. Among them, about 35 carbon sources such as amygdalin and L-arabinose could be metabolized to produce conidia. And about 57 carbon sources (tween 80, D-arabinose, glucuronic acid, etc.) could be metabolized by B. cinerea, but could not promote the conidia formation. When treated by 1 mg/L of carbendazim and 5 mg/L of prochloraz, and at both 1 and 10 mg/L of propiconazole, the carbon sources for conidia production of B. cinerea declined greatly. While treated for 0.08 and 1 mg/L of pyrimethanil or 0.1 and 5 mg/L of iprodione, they showed no inhibition. The results mentioned above revealed the carbon sources for conidia production of B. cinerea under chemical pressures of fungicides, which give us some constructive suggestions for the fungicide selection in gray mold management. [ABSTRACT FROM AUTHOR]

Introduction. The study of the protein composition of the causative agent of anthrax — Bacillus anthracis, allows you to identify both general species and individual characteristics of strains that differ in phenotypic properties, manifested mainly in the vegetative form and which are important for virulence, immunogenicity and the ability to adapt to different vegetation conditions.Purpose of the work. In the group of anthrax microbe strains having different plasmid composition and virulence, different methods of extraction of the total proteome from vegetative bacillus cells have been tested.Results. In the course of the work, it was shown that the rate of spore formation varies significantly between individual strains of the anthrax microbe and can have a significant impact on the efficiency of extraction and the composition of the protein complex. Preliminary treatment with lysozyme, which affects the cell membrane, promotes a more complete lysis of cells, and ultramicrocentrifuge filtration provides complete specific sterility of the obtained samples.Conclusion. A culture preparation scheme was developed for B. anthracis, which allows one to obtain a culture in the vegetative phase of the life cycle and to efficiently extract proteins in combination with reliable disinfection of samples.

Streptomyces griseus S4-7, a well-characterized keystone taxon among strawberry microbial communities, shows exceptional disease-preventing ability. The whole-genome sequence, functional genes, and bioactive secondary metabolites of the strain have been described in previous studies. However, proteomics studies of not only the S4-7 strain, but also the Streptomyces genus as a whole, remain limited to date. Therefore, in the present study, we created a proteomics reference map for S. griseus S4-7. Additionally, analysis of differentially expressed proteins was performed against a wblE2 mutant, which was deficient in spore chain development and did not express an antifungal activity-regulatory transcription factor. We believe that our data provide a foundation for further in-depth studies of functional keystone taxa of the phytobiome and elucidation of the mechanisms underlying plant-microbe interactions, especially those involving the Streptomyces genus.

Ulva prolifera, the main causative species of green tide, has drawn much attention. Spore reproduction was one of the main reproduction strategies that could be induced by changing environmental factors, but the mechanism of spore formation remains obscure. Here, we cultured U. prolifera (segments) under the optimal sporulation condition, and four states in total from vegetative to reproductive were defined. Additionally, the chlorophyll fluorescence and transcriptome analysis were determined at these four states. The results showed that: (1) Compared with state I, the photosynthetic capacity (Fv/Fm, Fv′/Fm′, rETRmax) and chlorophyll content (Chl a, Chl b, carotenoids) were enhanced in state II, whereas it decreased in state III and IV (the spore formation period); (2) a total of 41,058 unigenes were expressed during the spore formation process; (3) compared with state I, the genes related with photosynthesis, terpenoid backbone biosynthesis, and carotenoid biosynthesis were significantly upregulated in states II, III, and IV whereas glycolysis was downregulated in state I; (4) some genes of the transcription factors families, such as the C3H family, may be one of the key factors that regulate genes in the spore formation; (5) 574 of the differentially expressed genes (DEGs) associated with flagella biosynthesis were annotated according to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes Orthology, including 20 unigenes encoding intraflagellar transport proteins (IFTs) which had not been reported in previous transcriptome analysis in U. prolifera. This study provides a new perspective of spore formation at the gene transcriptional level, although the detailed transcription regulatory spore formation network remains to be unveiled.

We present the case of a 19-year-old man with an open fracture of the tibia and fibula secondary to an accident with an all-terrain vehicle. He underwent operative excisional irrigation, debridement, and fixation on the day of injury. His course was complicated by nonunion of the tibia fracture. Infection is a common factor in fracture nonunion, even in patients who receive appropriate surgical and antimicrobial management. Paenibacillus turicensis , an organism adapted to survive in the environment via spore formation, was responsible for nonunion in our patient. A brief discussion of this unusual organism, fracture nonunion, and the role of infection in etiology of nonunion follows. [ABSTRACT FROM AUTHOR]

A decrease in therapeutic effect of some live lacto- and bifdobacteria-based drugs for veterinary use has been observed for the last 20 years that urges scientists to search for new microorganisms possessing probiotic properties. Many studies in this feld are focused on Bacillus subtilis that is widespread in the environment and non-pathogenic for animals and humans. Results of tests of Bacillus subtilis for its biological properties and antagonistic activity aimed at optimization of methodical approaches for detection of strain with the highest antagonistic effect on some opportunistic microorganisms and their further use as probiotics are described. Cultural morphological and biochemical characteristics of the tested strains conformed to the species characteristics of Bacillus subtilis. Tested strains were nonpathogenic for white mice. Tests showed that spore biomass could be prepared both in liquid and on solid nutrient media. Methodically, spore biomass preparation in liquid nutrient medium is preferable. The tests showed that spores emerged from anabiosis non-uniformly and it depended on original seed spore storage period. Spore cultures stored less than one year emerged from anabiosis more quickly. It was found that the spores formed more readily when the cultures were aerated with oxygen as well as that lag-phase culture medium had a stimulating effect on Bacillus subtilis spore germination. Bacillus subtilis strains were found to have antagonistic effect on Escherichia coli, Salmonella and Staphylococcus. Area of growth inhibition of the said bacteria was 15–20 mm. Tested Bacillus subtilis strains could be proposed for use as probiotics.

Geobacillus species are important contaminants in the dairy industry, and their presence is often considered an indicator of poor plant hygiene with the potential to cause spoilage. They can form heat-resistant spores that adhere to surfaces of processing equipment and germinate to form biofilms. Therefore, strategies aimed toward preventing or controlling biofilm formation in the dairy industry are desirable. In this study, we demonstrated that the preferred temperature for biofilm and spore formation among Geobacillus stearothermophilus A1, D1, P3, and ATCC 12980 was 65°C. Increasing the total dissolved milk solid concentration to 20% (wt/vol) caused an apparent delay in the onset of biofilm and spore formation to detectable concentrations among all the strains at 55°C. Compared to the onset time of the biofilm formation of A1 in 10% (wt/vol) reconstituted skim milk, addition of milk protein (whey protein and sodium caseinate) caused an apparent delay in the onset of biofilm formation to detectable concentrations by an average of 10 h at 55°C. This study proposes that temperature and total dissolved solid concentration have a cumulative effect on biofilm and spore formation by G. stearothermophilus A1, D1, P3, and ATCC 12980. In addition, the findings from this study may indicate that preconditioning of stainless steel surfaces with adsorbed milk proteins may delay the onset of biofilm and spore formation by thermophilic bacteria during milk powder manufacture. [ABSTRACT FROM AUTHOR]

Fungal sporulation is affected by many environmental factors, for example, we previously observed that embedding of a hydrophobic polymer net in an agar plate medium significantly accelerates spore formation of some fungi. Here, it was found that the fungal spore formation depended on the surface hydrophobicity of cultivation vessels used for the plate cultivation. In a polypropylene (PP) vessel, six fungal strains produced spores of 1.5 to 514.8 times of those growing in a glass vessel. The contact of vegetative hyphae on the surface of the vessels might trigger the fungal spore formation. Moreover, the spore formation was synergistically accelerated by the reduction of nitrogen source content in an agar plate medium and by the contact to hydrophobic polymers. The synergistic effect depended on the surface area of the hydrophobic polymer. Thus, the combination of the reduction of nitrogen source and the embedding of hydrophobic polymer is expected as a novel and effective procedure for production of fungal spores which are useful for the inoculum in fermentation industry and biocontrol agent in agriculture. [Display omitted] • The higher the hydrophobicity of culture vessel, the more the spore formation. • Spore production in polypropylene vessel reached 1.5 to 514.8 times of glass one. • N-source reduction and hydrophobicity synergistically accelerated spore formation. • The synergistic effect depended on the surface area of hydrophobic polymer. [ABSTRACT FROM AUTHOR]

Many factors determine the resistance properties of a Bacillus spore to heat, chemical and physical processing, including thick proteinaceous coats, peptidoglycan cortex and low water content, high levels of dipicolinic acid (DPA), and divalent cations in the spore core. Recently, attention has been focused on non-thermal inactivation methods based on high pressure, ultrasonic, high voltage electric fields and cold plasmas for inactivating Bacillus spores associated with deterioration in quality and safety. The important chemical sporicides are glutaraldehyde, chorine-releasing agents, peroxygens, and ethylene oxide. Some food-grade antimicrobial agents exhibit sporostatic and sporicidal activities, such as protamine, polylysine, sodium lactate, essential oils. Surfactants with hydrophilic and hydrophobic properties have been reported to have inactivation activity against spores. The combined treatment of physical and chemical treatment such as heating, UHP (ultra high pressure), PEF (pulsed electric field), UV (ultraviolet), IPL (intense pulsed light) and natural antimicrobial agents can act synergistically and effectively to kill Bacillus spores in the food industry. [ABSTRACT FROM AUTHOR]

Many peptidoglycan-deficient bacteria such as the Mycoplasmatales are known host-associated lineages, lacking the environmental resistance mechanisms and metabolic capabilities necessary for a free-living lifestyle. Several peptidoglycan-deficient and non-sporulating orders of interest are thought to be descended from Gram-positive sporulating Bacilli through reductive evolution. Here we annotate 2650 genomes belonging to the class Bacilli , according to the Genome Taxonomy Database, to predict the peptidoglycan and sporulation phenotypes of three novel orders, RFN20 , RF39 and ML615J-28 , known only through environmental sequence surveys. These lineages are interspersed between peptidoglycan-deficient non-sporulating orders including the Mycoplasmatales and Acholeplasmatales , and more typical Gram-positive orders such as the Erysipelotrichales and Staphylococcales . We use the extant genotypes to perform ancestral state reconstructions. The novel orders are predicted to have small genomes with minimal metabolic capabilities and to comprise a mix of peptidoglycan-deficient and/or non-sporulating species. In contrast to expectations based on cultured representatives, the order Erysipelotrichales lacks many of the genes involved in peptidoglycan and endospore formation. The reconstructed evolutionary history of these traits suggests multiple independent whole-genome reductions and loss of phenotype via intermediate transition states that continue into the present. We suggest that the evolutionary history of the reduced-genome lineages within the class Bacilli is one driven by multiple independent transitions to host-associated lifestyles, with the degree of reduction in environmental resistance and metabolic capabilities correlated with degree of host association.

Aims: Equol is a nonsteroidal oestrogen of the isoflavone class. We investigated the antibacterial ability of equol with respect to the growth rate, toxin production and spore‐forming abilities of Clostridioides difficile BI/027/NAP1. Methods and Results: Isoflavones, or female hormones, were added to bacterial culture, which was grown at 35°C. The absorbance of the culture was measured at various time points for evaluating the growth inhibition. The toxin levels in the media and morphological changes were also assessed. To evaluate the influence of equol on the sporulation of C. difficile, cells were collected at various time points from the equol‐supplemented culture and the number of spores was counted. Our results show that equol inhibits bacterial growth in a concentration‐dependent manner. However, it does not inhibit the production of toxin by C. difficile. Other isoflavones and female hormones did not inhibit the C. difficile growth. At the 14th day, approximately 600 spores were present in the control medium and only six were seen in the equol‐containing medium. Conclusion: Our results suggest that equol may directly inhibit the C. difficile growth in a concentration‐dependent manner and spore formation. Significance and Impact of the Study: This is the first report on the antimicrobial ability of equol. [ABSTRACT FROM AUTHOR]

Anthrax toxins and capsules, which are encoded by genes located on pXO1 and pXO2, respectively, are major virulence factors of Bacillus anthracis. Our previous studies demonstrated that exposure to high-temperatures is unable to abolish the pXO1 plasmid of the Pasteur II strain, but the growth of the strain was obviously slower than that of the Sterne strain and wild-type virulent strain. To elucidate a potential regulatory mechanism of slowing growth, we employed comparative genome and bioinformatic analysis and revealed a unique SNP (G to T) at the 143135 bp position in pXO1 that is possibly involved in the mediation of growth of Pasteur II. However, the T to G mutation in groR did not result in any change of the amino acid sequence. A predominant nucleotide G existed at the 143135 bp in pXO1 of 100 wild-type B. anthracis isolates and 9 isolates documented in GenBank, whereas T replaced G in pXO1 of the Pasteur II strain. Further analysis indicate that the SNP is located in a gene between 143042 and 143173 bp, and that it encodes a small protein of 43 amino acids and is termed as a growth regulator (GroR). Site-directed mutagenesis and gene deletion demonstrates that groR regulates the growth and spore formation of B. anthracis. Our results indicate that the pXO1 plasmid is involved in the regulation of growth and spore formation in B. anthracis.

Botulinum neurotoxin (BoNT), produced by neurotoxigenic clostridia, is the most potent biological toxin known and the causative agent of the paralytic disease botulism. The nutritional, environmental, and genetic regulation of BoNT synthesis, activation, stability, and toxin complex (TC) formation is not well studied. Previous studies indicated that growth and BoNT formation were affected by arginine and glucose in Clostridium botulinum types A and B. In the present study, C. botulinum ATCC 3502 was grown in toxin production medium (TPM) with different levels of arginine and glucose and of three products of arginine metabolism, citrulline, proline, and ornithine. Cultures were analyzed for growth (optical density at 600 nm [OD600]), spore formation, and BoNT and TC formation by Western blotting and immunoprecipitation and for BoNT activity by mouse bioassay. A high level of arginine (20 g/liter) repressed BoNT production approximately 1,000-fold, enhanced growth, slowed lysis, and reduced endospore production by greater than 1,000-fold. Similar effects on toxin production were seen with equivalent levels of citrulline but not ornithine or proline. In TPM lacking glucose, levels of formation of BoNT/A1 and TC were significantly decreased, and extracellular BoNT and TC proteins were partially inactivated after the first day of culture. An understanding of the regulation of C. botulinum growth and BoNT and TC formation should be valuable in defining requirements for BoNT formation in foods and clinical samples, improving the quality of BoNT for pharmaceutical preparations, and elucidating the biological functions of BoNTs for the bacterium. IMPORTANCE Botulinum neurotoxin (BoNT) is a major food safety and bioterrorism concern and is also an important pharmaceutical, and yet the regulation of its synthesis, activation, and stability in culture media, foods, and clinical samples is not well understood. This paper provides insights into the effects of critical nutrients on growth, lysis, spore formation, BoNT and TC production, and stability of BoNTs of C. botulinum. We show that for C. botulinum ATCC 3502 cultured in a complex medium, a high level of arginine repressed BoNT expression by ca. 1,000-fold and also strongly reduced sporulation. Arginine stimulated growth and compensated for a lack of glucose. BoNT and toxin complex proteins were partially inactivated in a complex medium lacking glucose. This work should aid in optimizing BoNT production for pharmaceutical uses, and furthermore, an understanding of the nutritional regulation of growth and BoNT formation may provide insights into growth and BoNT formation in foods and clinical samples and into the enigmatic function of BoNTs in nature. [ABSTRACT FROM AUTHOR]

The two-component phosphorelay system is the most prevalent mechanism for sensing and transducing environmental signals in bacteria. Spore formation, which relies on the two-component phosphorelay system, enables the long-term survival of the glacial bacterium Paenisporosarcina sp. TG-14 in the extreme cold environment. Spo0A is a key response regulator of the phosphorelay system in the early stage of spore formation. The protein is composed of a regulatory N-terminal phospho-receiver domain and a DNA-binding C-terminal activator domain. We solved the three-dimensional structure of the unphosphorylated (inactive) form of the receiver domain of Spo0A ( PaSpo0A-R) from Paenisporosarcina sp. TG-14. A structural comparison with phosphorylated (active form) Spo0A from Bacillus stearothermophilus ( BsSpo0A) showed minor notable differences. A molecular dynamics study of a model of the active form and the crystal structures revealed significant differences in the α4 helix and the preceding loop region where phosphorylation occurs. Although an oligomerization study of PaSpo0A-R by analytical ultracentrifugation (AUC) has shown that the protein is in a monomeric state in solution, both crosslinking and crystal-packing analyses indicate the possibility of weak dimer formation by a previously undocumented mechanism. Collectively, these observations provide insight into the mechanism of phosphorylation-dependent activation unique to Spo0A. [ABSTRACT FROM AUTHOR]

We have developed butanol-producing consolidated bioprocessing from cellulosic substrates through coculture of cellulolytic clostridia and butanol-producing Clostridium saccharoperbutylacetonicum strain N1-4. However, the butanol fermentation by strain N1-4 (which has an optimal growth temperature of 30°C) is sensitive to the higher cultivation temperature of 37°C; the nature of this deleterious effect remains unclear. Comparison of the intracellular metabolites of strain N1-4 cultivated at 30°C and 37°C revealed decreased levels of multiple primary metabolites (notably including nucleic acids and cofactors) during growth at the higher temperature. Supplementation of the culture medium with 250 mg/liter adenine enhanced both cell growth (with the optical density at 600 nm increasing from 4.3 to 10.2) and butanol production (increasing from 3.9 g/liter to 9.6 g/liter) at 37°C, compared to those obtained without adenine supplementation, such that the supplemented 37°C culture exhibited growth and butanol production approaching those observed at 30°C in the absence of adenine supplementation. These improved properties were based on the maintenance of cell viability. We further showed that adenine supplementation enhanced cell viability during growth at 37°C by maintaining ATP levels and inhibiting spore formation. This work represents the first demonstration (to our knowledge) of the importance of adenine-related metabolism for clostridial butanol production, suggesting a new means of enhancing target pathways based on metabolite levels. IMPORTANCE Metabolomic analysis revealed decreased levels of multiple primary metabolites during growth at 37°C, compared to 30°C, in C. saccharoperbutylacetonicum strain N1-4. We found that adenine supplementation restored the cell growth and butanol production of strain N1-4 at 37°C. The effects of adenine supplementation reflected the maintenance of cell viability originating from the maintenance of ATP levels and the inhibition of spore formation. Thus, our metabolomic analysis identified the depleted metabolites that were required to maintain cell viability. Our strategy, which is expected to be applicable to a wide range of organisms, permits the identification of the limiting metabolic pathway, which can serve as a new target for molecular breeding. The other novel finding of this work is that adenine supplementation inhibits clostridial spore formation. The mechanism linking spore formation and metabolomic status in butanol-producing clostridia is expected to be the focus of further research. [ABSTRACT FROM AUTHOR]

Clostridium thermocellum can rapidly solubilize cellulose and produces ethanol as an end product of its metabolism. As such, it is a candidate for bioethanol production from plant matter. In this study, we developed an inducible expression system for C. thermocellum based on its native celC operon. We enhanced expression over the native operon structure by placing the repressor gene, glyR3, immediately after the celC promoter, and expressing the target gene after glyR3. Upon the addition of the inducer substrate, laminaribiose, an approximately 40-fold increase in gene expression was obtained using the test gene spo0A. Furthermore, induction of the sporulation histidine kinase, clo1313_ 1942, increased sporulation frequency by approximately 10,000-fold relative to an uninduced control. We have also shown that the laminaribiose (β1-3-linked carbon source) utilization pathway is not catabolite repressed by cellobiose, a β1-4-linked carbon source frequently used for C. thermocellum cultivation in laboratory conditions. Selective expression of target genes has the potential to inform metabolic engineering strategies as well as increase fundamental understanding of C. thermocellum biology. [ABSTRACT FROM AUTHOR]

Background: Dipicolinic acid (DPA) comprising 5-15% of the spore dry weight is important for spore stability and resistance properties. In some Gram-positive bacteria such as Clostridium perfringens, an electron transfer flavoprotein, EtfA, acts in an incorporative way in DPA formation. Objectives: The aim of this study was to explore if the fixA and fixB genes which are electron transfer flavoproteins could incorporate in DPA formation in Mycobacterium marinum CCUG 20988. Materials and Methods: In this descriptive-analytic study, the fixA and fixB genes were PCR-amplified and ligated between the Nde I and Hind III sites of pET19b vector with ampicillin resistance gene in frame with an N-terminal T7-promotor and a C-terminal of 6-histidin tag and transformed into BL21-Codon Plus-RIL Escherichia coli competent cells. For over expression of fixA and fixB genes, SDS-polyacrylamide gel electrophoresis was used. Also the direct interaction between dihydrodipicolinate acid (DHDPA) synthase and fixA and fixB proteins for DPA formation was investigated. Results: Detectable DPA formation has been identified upon isopropyl-β-D-thiogalactopyranoside (IPTG) induction using fixA and fixB genes together. The amount of dipicolinic was about 32.8 and 49.2 µg/mL acids in LB and M9 media, respectively. Conclusions: It can be concluded that both fixA and fixB genes as electron transfer flavoproteins may incorporate in dipicolinic acid formation in M. marinum. We assume that if sporulation turns out to be a common mechanism used by mycobacteria in response to environmental conditions, it might be one of the means by which it attains dormancy within the host. [ABSTRACT FROM AUTHOR]

Clostridioides difficile ( C. difficile ) is a Gram-positive, spore-forming, toxin-producing, obligate anaerobic bacterium. C. difficile infection (CDI) is the leading cause of healthcare-associated infective diarrhoea. The infection is mediated by the spore, a metabolically inactive form of C. difficile . The spore coat acts as a physical barrier to defend against chemical insults from hosts and natural environments. The composition of spore coat has already been revealed; therefore, the interactive networks of spore coat proteins and the dynamic process of coat assembly are the keys to design strategies to control and cure CDI. This review gives a brief discussion of the signal processing and transcriptional regulation of C. difficile sporulation initiation. Following the discussion, the spore formation is also introduced. Finally, this review mainly focuses on the spore coat assembly, a poorly understood process in C. difficile , and important proteins that have been studied.

With more than 400 species, the diatom genus Chaetoceros Ehrenberg is one of the most species-rich genera of marine planktonic diatoms. Although many Chaetoceros taxa are considered to be cosmopolitan, research has mainly focused on polar and temperate areas. Studies on material from tropical waters suggested that a high and undescribed species diversity of Chaetoceros occur here. On the basis of plankton samples collected in Daya Bay, South China Sea, a new species, C. rotosporus sp. nov., is described. Vegetative cells and resting spores were examined by light microscopy as well as transmission and scanning electron microscopy. The most variable part of the nuclear large-subunit ribosomal DNA was sequenced for information on phylogenetic relationships. Characteristic traits of C. rotosporus were a 90° rotation of the resting spore during spore maturation and a nearly rectangular silica wing that extended around the girdle of the immature resting spore. Neither character has been observed in previously described Chaetoceros species. Other distinguishing characters were the very large apertures and the single chloroplast, which formed a shallow bowl with four arms within each cell and appeared H-shaped in broad girdle view. Chaetoceros rotosporus sp. nov. belongs to the subgenus Hyalochaete Gran, but it could not be placed in any of the traditional sections of Chaetoceros, indicating a need for revision of Chaetoceros using a combination of morphological and molecular data [ABSTRACT FROM AUTHOR]

Several reports state that three architectural units, including integration host factor, leucyl aminopeptidase (PepA), and purine regulator, are involved in transcriptional process with RNA polymerase in Escherichia coli. Similarly, Streptomyces species possess the same structural units. We previously identified a protein, Streptomyces integration host factor (sIHF), involved in antibiotic production and sporulation. Subsequently, the function of PepA (SCO2179) was examined in detail. PepA is highly conserved among various Streptomyces spp., but it has not yet been characterized in Streptomyces coelicolor. While it is annotated as a putative leucyl aminopeptidase because it contains a peptidase M17 superfamily domain, this protein did not exhibit leucyl aminopeptidase activity. SCO2179 deletion mutant showed increased actinorhodin production and sporulation, as well as more distinct physiological differences, particularly when cultured on N-acetylglucosamine (GlcNAc) minimal media. The results of two-dimensional gel analysis and reverse transcription PCR showed that the SCO2179 deletion increased protein and mRNA levels of ftsZ, ssgA, and actinorhodin (ACT)-related genes such as actII- ORF4, resulting in increased actinorhodin production and spore formation in minimal media containing GlcNAc. [ABSTRACT FROM AUTHOR]

Abstract: Diaphanous-related formins (DRFs) drive the nucleation and elongation of linear actin filaments downstream of Rho GTPase signalling pathways. Dictyostelium formin C (ForC) resembles a DRF, except that it lacks a genuine formin homology domain 1 (FH1), raising the questions whether or not ForC can nucleate and elongate actin filaments. We found that a recombinant ForC-FH2 fragment does not nucleate actin polymerization, but moderately decreases the rate of spontaneous actin assembly and disassembly, although the barbed-end elongation rate in the presence of the formin was not markedly changed. However, the protein bound to and crosslinked actin filaments into loose bundles of mixed polarity. Furthermore, ForC is an important regulator of morphogenesis since ForC-null cells are severely impaired in development resulting in the formation of aberrant fruiting bodies. Immunoblotting revealed that ForC is absent during growth, but becomes detectable at the onset of early aggregation when cells chemotactically stream together to form a multicellular organism, and peaks around the culmination stage. Fluorescence microscopy of cells ectopically expressing a GFP-tagged, N-terminal ForC fragment showed its prominent accumulation in the leading edge, suggesting that ForC may play a role in cell migration. In agreement with its expression profile, no defects were observed in random migration of vegetative mutant cells. Notably, chemotaxis of starved cells towards a source of cAMP was severely impaired as opposed to control. This was, however, largely due to a marked developmental delay of the mutant, as evidenced by the expression profile of the early developmental marker csA. In line with this, chemotaxis was almost restored to wild type levels after prolonged starvation. Finally, we observed a complete failure of phototaxis due to abolished slug formation and a massive reduction of spores consistent with forC promoter-driven expression of β-galactosidase in prespore cells. Together, these findings demonstrate ForC to be critically involved in signalling of the cytoskeleton during various stages of development. [Copyright &y& Elsevier]

The review deals with the phenomenon of allolysis, i.e., lysis of a part of a bacterial population induced by a group of epigenetically differentiated cells of the same species or phylotype. Allolysis is best studied in two species of gram-positive bacteria, Streptococcus pneumoniae and Bacillus subtilis. In S. pneumoniae, allolysis is associated with the onset of the competence stage, while in B. subtilis it is associated with transition to the stage of spore formation. The mechanisms of allolysis are considered, as well as its possible role in the populational and symbiotic relationships of bacterial cells. The relation between allolysis ant the programmed death of a part of the cells within a bacterial population (apoptosis) is discussed. [ABSTRACT FROM AUTHOR]

Abstract: The combined effect of incubation time (IT) and aspartic acid concentration (AA) on the predicted biomass concentration (BC), Bacillus sporulation (BS) and anti-fungal activity of compounds (AFA) produced by Bacillus amyloliquefaciens CCMI 1051, was studied using Artificial Neural Networks (ANNs). The values predicted by ANN were in good agreement with experimental results, and were better than those obtained when using Response Surface Methodology. The database used to train and validate ANNs contains experimental data of B. amyloliquefaciens cultures (AFA, BS and BC) with different incubation times (1–9days) using aspartic acid (3–42mM) as nitrogen source. After the training and validation stages, the 2–7-6–3 neural network results showed that maximum AFA can be achieved with 19.5mM AA on day 9; however, maximum AFA can also be obtained with an incubation time as short as 6days with 36.6mM AA. Furthermore, the model results showed two distinct behaviors for AFA, depending on IT. [ABSTRACT FROM AUTHOR]

Molecular genetic characteristics and capacity for sporulation under different levels of temperature and humidity were compared for three saprotrophic and four clinical strains of A. sydowii. Analysis of the ITS and D1/D2 loci of these A. sydowii strains revealed two clades, each including both the clinical and saprotrophic strains. The differences in sporulation in the saprotrophic and clinical strains of the potentially pathogenic microscopic fungus A. sydowii under different environmental conditions were demonstrated. In the clinical A. sydowii strains, the level of spore formation was generally higher, especially at humidity levels of 0.90 and 0.95 a and 20-25°C. The level of spore formation for the clinical strains inoculated into sterile soil was several times higher than for the saprotrophic ones. On the contrary, nonsterile soils (sod-podzolic and urban soils) exhibited a fungistatic effect against A. sydowii populations. [ABSTRACT FROM AUTHOR]

Meiosis is a specialised form of the cell cycle that gives rise to haploid gametes. In Schizosaccharomyces pombe, the products of meiosis are four spores, which are formed by encapsulation of the four meiosis II nuclei within the cytoplasm of the zygote produced by fusion of the mating cells. The S. pombe spindle pole body is remodelled during meiosis II and membrane vesicles are then recruited there to form the forespore membrane, which encapsulates the haploid nucleus to form a prespore. Spore wall material is then deposited, giving rise to the mature spore. The septation initiation network is required to coordinate cytokinesis and mitosis in the vegetative cycle and for spore formation in the meiotic cycle. We have investigated the role of the SIN regulator dma1p in meiosis; we find that although both meiotic divisions occur in the absence of dma1p, asci frequently contain fewer than four spores, which are larger than in wild-type meiosis. Our data indicate that dma1p acts in parallel to the leading-edge proteins and septins to assure proper formation for the forespore membrane. Dma1p also contributes to the temporal regulation of the abundance of the meiosis-specific SIN component mug27p. [ABSTRACT FROM AUTHOR]

The aim of the present experiment was to investigate the influence of silver nanoparticles on Fusarium culmorum (W.G. Smith) Sacc. (FC) spores. The silver nanoparticles were produced by the high-voltage arc discharge method. To test the effect of silver nanoparticles on FC spores, 3 parameters were tested. One of these parameters was the vegetative mycelial growth in 2 experiments. The first involved the growth of FC spores on potato dextrose agar (PDA) medium after contact with 0.12-10 ppm of silver nanoparticles, and the second the growth of spores after contact with 0.12-2.5 ppm solutions of silver, but with culturing on 3 types of media (PDA, nutrient-poor PDA, and agar) instead. The next parameter was the formation of spores after the mycelia were cultured. The last parameter was spore germination in a 2.5 ppm solution of silver nanoparticles. A significant reduction in mycelial growth was observed for spores incubated with silver nanoparticles. This relationship was dependent on the incubation time and type of growth medium, but did not depend significantly on the concentration of silver nanoparticles up to 2.5 ppm. The sporulation test showed that, relative to control samples, the number of spores formed by mycelia increased in the culture after contact with silver nanoparticles, especially on the nutrient-poor PDA medium. The 24 h incubation of FC spores with a 2.5 ppm solution of silver nanoparticles greatly reduced the number of germinating fragments and sprout length relative to the control. Le but de ces expériences était d’examiner l’influence de nanoparticules d’argent sur des spores de Fusarium culmorum (W.G. Smith) (FC). Les nanoparticules d’argent ont été produites par la méthode de décharge en arc à haut voltage. Afin de tester l’effet des nanoparticules d’argent sur les spores de FC, 3 paramètres ont été examinés. Un de ces paramètres consistait à mesurer la croissance du mycélium végétatif lors de deux expériences: premièrement, à partir des spores mises en contact avec 0,12 à 10 ppm de nanoparticules d’argent dans du milieu PDA, et deuxièmement, à partir des spores mises en contact avec 0,12 à 10 ppm d’argent en solution, dans trois types de milieux de culture (PDA, PDA appauvri et gélose). Le paramètre suivant consistait à examiner la formation de spores obtenues après la culture du mycélium, et le dernier, à examiner la germination des spores dans une solution contenant 2,5 ppm de particules d’Ag. Une réduction significative de la croissance du mycélium a été observée à partir des spores incubées en présence de nanoparticules d’argent. Cette relation dépendait du temps d’incubation et du type de milieu de croissance, mais ne dépendait pas significativement de la concentration de nanoparticules d’argent jusqu’à 2,5 ppm. Le test de sporulation a montré que le nombre de spores formées par le mycélium en culture augmentait après un contact avec les nanoparticules d’argent comparativement au milieu contrôle, notamment dans le milieu PDA appauvri. L’incubation des spores de FC pendant 24 h dans une solution contenant 2,5 ppm de nanoparticules d’argent réduisait grandement le nombre et la longueur des pousses germinatives, comparativement au contrôle. [ABSTRACT FROM AUTHOR]

Aims: The aim of this study was to evaluate the influence of environmental conditions on the antifungal activity of the Bacillus sp. CCMI 1053 cultures. Methods and Results: The electrospray ionization mass spectra (ESI-MS) analysis was used to detect the active peptides produced by Bacillus amyloliquefaciens CCMI 1051 cultures in a glucose-containing medium to which four different nitrogen sources were added. The cultures produced different patterns of Bacillus sporulation and distinct antifungal activity of the cell-free culture broths. Conclusions: The highest sporulation obtained corresponds to higher antifungal activity when it is formed after 3 days of microbial growth. The antifungal activity against Trichoderma harzianum CCMI 783 is more influenced by the concentration on the nitrogen source than the culture time of incubation. The association of nitrogen concentration and the time of incubation is particularly relevant in the expression of the antifungal activity. Significance and Impact of the Study: The present findings allow the reduction of the use of chemical pesticides and to limit some plant diseases. The association of the nitrogen source and the time of incubation is a novelty, which would improve the production of secondary metabolites. Both economical and environmental benefits arise from the study. [ABSTRACT FROM AUTHOR]

Abstract: Dictyostelium has 55 genes encoding seven-transmembrane G-protein-coupled receptors (GPCR) that belong to five of the six GPCR families. GrlA is one of the 17 family 3 GPCRs in Dictyostelium all of which resemble GABAB receptors from higher eukaryotes. GrlA is a 90-kDa protein present on the plasma membrane and on membranes of the ER. It has a large extracellular domain with homology to bacterial periplasmic proteins. The GrlA message is present throughout development and shows increased levels during the post aggregation stages. Inactivation of the grlA gene does not severely affect the growth phase, however, it leads to a delay in the development at the post aggregation stage. GrlA deficient strains show an altered DIF-1 response specific to the prestalk-specific ecmA and ecmB gene, reduced car2 and pkaC transcript levels and form a reduced number of spores. Germination of the spores was as in wild type. Transcriptional profiling supported the defect in the sporulation pathway as a large number of genes involved in the biogenesis and organization of the extracellular matrix and the sporulation process were significantly downregulated in the mutant. [Copyright &y& Elsevier]

A survey was undertaken at a whole milk powder manufacturing plant to determine the origin and the rate of spore formation of thermophilic bacteria. Spores were generally not detected (< 10 cfu/mL) in either the pasteurization process or the pasteurized milk directed into the powder plant. The predominant sites of spore formation were the preheater plate heat exchanger and the evaporator. Spores began to be detected approximately 9 h into an 18-h manufacturing run. Spore isolates were identified as Anoxybacillus flavithermus and Geobacillus species. A. flavithermus predominated in the preheat section, whereas a mix of both organisms was present in the later manufacturing stages. [ABSTRACT FROM AUTHOR]

The combined use of the contemporary vector systems, the bacterial artificial chromosome (BAC) vector and the Bacillus subtilis genome (BGM) vector, makes possible the handling of giant-length DNA (above 100kb). Our newly constructed BGM vector efficiently integrated DNA prepared in the BAC vector. A BAC library comprised of 18 independent clones prepared from mitochondrial DNA (mtDNA) of Arabidopsis thaliana was converted to a parallel BGM library using the new BGM vector. The effectiveness of the combined use of the vector systems was confirmed by the stable recovery of all 18 DNAs as BAC clones from the respective BGM clones. We show that DNA in BGM was stably preserved at room temperature after spore formation of the host B.subtilis. Rapid and stable shuttling between Escherichiacoli and the B.subtilis host, combined with spore-mediated DNA storage, may facilitate the long-term and low-cost preservation and the transportation of DNA resources. [Copyright &y& Elsevier]

Five strains of sulfate-reducing bacteria were isolated from the highest positive dilutions of a most probable number (MPN) series supplemented with lactate and inoculated with sediments from the oligotrophic Lake Stechlin. The isolates were endospore-forming and were motile by means of laterally inserted flagella. They stained Gram-negative and containedb-type cytochromes. CO difference spectra indicated the presence of P582 as a sulfite reductase. Phylogenetic analyses of the 16S rDNA sequences revealed that the isolates were very closely affiliated with the genusSporomusa. However, sulfate and amorphous Fe(OH)3, but not sulfite, elemental sulfur, MnO2, or nitrate were used as terminal electron acceptors. Homoacetogenic growth was found with H2/CO2 gas mixture, formate, methanol, ethanol, and methoxylated aromatic compounds. The strains grew autotrophically with H2 plus CO2 in the presence or absence of sulfate. Formate, butyrate, several alcohols, organic acids, carbohydrates, some amino acids, choline, and betaine were also utilized as substrates. The growth yield with lactate and sulfate as substrate was 7.0 g dry mass/mol lactate and thus two times higher than in sulfate-free fermenting cultures. All isolates were able to grow in a temperature range of 4-37°C. Physiologically and by the presence of a Gram-negative cell wall, the new isolates resemble knownDesulfosporosinusspecies. However, phylogenetically they are affiliated with the Gram-negative genusSporomusabelonging to theSelenomonassubgroup of the Firmicutes. Therefore, the new isolates reveal a new phylogenetic lineage of sulfate-reducing bacteria. A new genus and species,Desulfosporomusa polytropagen. nov., sp. nov. is proposed. [ABSTRACT FROM AUTHOR]

Fungi employ different carbohydrate uptake systems to adapt to certain environmental conditions and to different carbon source concentrations. The hydrolysis of polymeric carbohydrates and the subsequent uptake of monomeric forms may also play a role in development. Aspergillus nidulans accumulates cell wall components during vegetative growth and degrades them during sexual development. We have identified the hxtA (high affinity hexose transporter) gene in a differential library, which was enriched for sexual-specific genes. The hxtA gene is disrupted by 6 introns and predicted to encode a 531 amino acid protein with high similarity to major facilitator superfamily members including the high affinity hexose transporter Gtt1 from Trichoderma harzianum. A. nidulans HxtA contains the 12 predicted transmembrane domains characteristic for this family. Deletion of hxtA did not impair growth of A. nidulans on a variety of carbon sources nor did it inhibit sexual development suggesting redundant sugar uptake systems. We found at least 17 putative hexose transporters in the genome of A. nidulans. Despite the high similarity of HxtA to fungal high affinity glucose transporters, the hxtA gene did not restore growth on glucose of a Saccharomyces cerevisiae mutant, in which all hexose transporters were deleted. Northern blot analysis revealed that the A. nidulans hxtA gene was repressed under high glucose conditions and expressed in vegetative hyphae upon carbon starvation and during sexual development. We found hxtA(p)::sgfp expression in developing cleistothecia specifically in ascogenous hyphae and propose that HxtA is a high affinity glucose transporter involved in sugar metabolism during sexual development. [Copyright &y& Elsevier]

Anaerobic enrichment cultures, with erythritol as substrate, resulted in the isolation of a strain with properties not yet found in an existing genus in this combination. The strain, FKBS1, was strictly anaerobic, stained gram-negative and formed spores. Cells were small motile vibrios with flagella inserted at the concave side of the cell. Spores were located terminally and caused only slight swelling of the cells if compared to related spore-forming genera. FKBS1 fermented fructose, mannitol, sorbitol, xylitol and erythritol to propionic acid, acetic acid, CO2 and small amounts of H2 to balance the difference in the oxidation-reduction value between substrate and cell mass. The 16S rDNA sequence revealed relationship to the Sporomusa-Pectinatus-Selenomonas group. However, the phylogenetic distance to any of its members was too great to allow it to be placed in one of the existing genera. Morphologically the strain resembled Sporomusa, which, however, performs an acetogenic type of fermentation. The propionic-acid-forming genera of the group are either not spore-formers or, in the case of Dendrosporobacter quercicolus (syn. Clostridium quercicolum), morphologically different. It is therefore proposed to classify strain FKBS1 as a new genus and species, Propionispora vibrioides. [ABSTRACT FROM AUTHOR]