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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.

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, es-pecially those involving the Streptomyces genus.

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 onBacillus subtilisthat is widespread in the environment and non-pathogenic for animals and humans. Results of tests ofBacillus subtilisfor 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 ofBacillus 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 onBacillus subtilisspore germination.Bacillus subtilisstrains were found to have antagonistic effect onEscherichia coli, SalmonellaandStaphylococcus. Area of growth inhibition of the said bacteria was 15–20 mm. TestedBacillus subtilisstrains could be proposed for use as probiotics.

Background: Dictyostelium discoideum encodes both class I and II RNRs. Results: Both RNRs are expressed in D. discoideum, and depletion of class I RNR drastically reduces spore formation and viability. Conclusion: Class I RNR is central for D. discoideum growth and differentiation, whereas class II function is enigmatic. Significance: Our results underscore the evolutionary selection for this rare set of RNRs in D. discoideum., Ribonucleotide reductases (RNRs) catalyze the only pathway for de novo synthesis of deoxyribonucleotides needed for DNA replication and repair. The vast majority of eukaryotes encodes only a class I RNR, but interestingly some eukaryotes, including the social amoeba Dictyostelium discoideum, encode both a class I and a class II RNR. The amino acid sequence of the D. discoideum class I RNR is similar to other eukaryotic RNRs, whereas that of its class II RNR is most similar to the monomeric class II RNRs found in Lactobacillus spp. and a few other bacteria. Here we report the first study of RNRs in a eukaryotic organism that encodes class I and class II RNRs. Both classes of RNR genes were expressed in D. discoideum cells, although the class I transcripts were more abundant and strongly enriched during mid-development compared with the class II transcript. The quaternary structure, allosteric regulation, and properties of the diiron-oxo/radical cofactor of D. discoideum class I RNR are similar to those of the mammalian RNRs. Inhibition of D. discoideum class I RNR by hydroxyurea resulted in a 90% reduction in spore formation and decreased the germination viability of the surviving spores by 75%. Class II RNR could not compensate for class I inhibition during development, and an excess of vitamin B12 coenzyme, which is essential for class II activity, did not improve spore formation. We suggest that class I is the principal RNR during D. discoideum development and growth and is important for spore formation, possibly by providing dNTPs for mitochondrial replication.

Traditionally microorganisms were considered to be autonomous organisms that could be studied in isolation. However, over the last decades cell-to-cell communication has been found to be ubiquitous. By secreting molecular signals in the extracellular environment microorganisms can indirectly assess the cell density and respond in accordance. In one of the best-studied microorganisms, Bacillus subtilis, the differentiation processes into a number of distinct cell types have been shown to depend on cell-to-cell communication. One of these cell types is the spore. Spores are metabolically inactive cells that are highly resistant against environmental stress. The onset of sporulation is dependent on cell-to-cell communication, as well as on a number of other environmental cues. By using individual-based simulations we examine when cell-to-cell communication that is involved in the onset of sporulation can evolve. We show that it evolves when three basic premises are satisfied. First, the population of cells has to affect the nutrient conditions. Second, there should be a time-lag between the moment that a cell decides to sporulate and the moment that it turns into a mature spore. Third, there has to be environmental variation. Cell-to-cell communication is a strategy to cope with environmental variation, by allowing cells to predict future environmental conditions. As a consequence, cells can anticipate environmental stress by initiating sporulation. Furthermore, signal production could be considered a cooperative trait and therefore evolves when it is not too costly to produce signal and when there are recurrent colony bottlenecks, which facilitate assortment. Finally, we also show that cell-to-cell communication can drive ecological diversification. Different ecotypes can evolve and be maintained due to frequency-dependent selection., Author Summary Biological systems are characterized by communication; humans talk, insects produce pheromones and birds sing. Over the last decades it has been shown that even the simplest organisms on earth, the bacteria, communicate. Despite the prevalence of communication, it is often hard to explain how communicative systems evolve. In bacteria, communication results from the secretion of molecular signals that accumulate in the environment. Cells can assess the concentration of these signals, which indicate cell density, and respond in accordance. This form of cell-to-cell communication is responsible for the regulation of numerous bacterial behaviors, such as sporulation. Spores are metabolically inactive cells that are highly resistant against environmental stress. It is adaptive for a cell to sporulate when it struggles to survive. We show, via individual-based simulations, that cell-to-cell communication evolves because it allows cells to predict future environmental conditions. As a consequence, cells are capable of anticipating environmental stress by initiating sporulation before conditions are actually harmful. Furthermore, our model shows that cell-to-cell communication can even drive ecological diversification, since it facilitates the evolution of individuals that specialize on distinct ecological conditions.

Article history: Received 2 February 2010 Received in revised form 15 July 2010 Accepted 19 July 2010 Available online 27 July 2010 Keywords: Bacillus amiloliquefaciens Spore formation Anti-fungal activity Neural networks 1. Introduction Biopesticides based on natural endophytic bacteria to control plant diseases are a promising ecological alternative to chemical treatments. Bacillus species produce a wide variety of metabolites with interesting biological activities, among them iturinic lipopep- tides antibiotics (Bottone and Peluso, 2003; Cho et al., 2003; Moyne et al., 2001). The antimicrobial activity exhibited by Bacillus sp. is dependent on the culture medium composition, and different nitro- gen sources can result in the production of different antibiotics (Besson et al., 1987; Chevanet et al., 1986; Davis et al., 1999; Volpon et al., 2000). Aspartic acid is the preferred nitrogen source for the production of iturinic compounds by Bacillus subtilis (Besson et al., 1987) and Bacillus amyloliquefaciens (Caldeira et al., 2006, 2007, 2008). With increasing culture time, the nutrient content changes and adverse environmental conditions appear. Thus, incubation time is another factor influencing antibiotic production (Caldeira et al., 2008; Feio et al., 2004; Moyne et al., 2001), as the response to adverse environmental conditions can lead to activation of differ- ent mechanisms for the production of antibiotics giving a compet- itive advantage to the producer microorganism (Dieckmann et al., 2001). The link between antibiotic production and Bacillus sporulation is not fully understood. During production of lipopeptides in sub- ⇑ Corresponding author. Tel.: +351 266 745 315; fax: +351 266 745 303. E-mail address: hvicente@uevora.pt (H. Vicente). 0960-8524/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2010.07.080 abstract The combined effect of incubation time (IT) and aspartic acid concentration (AA) on the predicted bio- mass 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 con- tains experimental data of B. amyloliquefaciens cultures (AFA, BS and BC) with different incubation times (1–9 days) using aspartic acid (3–42 mM) 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 19.5 mM AA on day 9; however, maximum AFA can also be obtained with an incubation time as short as 6 days with 36.6 mM AA. Furthermore, the model results showed two distinct behaviors for AFA, depending on IT.