Growth-related variations in theBacillus cereus secretome

Institut National de la Recherche Agronomique (INRA), Génétique microbienne et Environnement, Guyancourt, France.
PROTEOMICS (Impact Factor: 3.81). 05/2007; 7(10):1719-28. DOI: 10.1002/pmic.200600502
Source: PubMed


Using 2-DE, transcriptional gene fusions and cell cytotoxicity assays, we followed changes in the Bacillus cereus strain ATCC14579 secretome, gene expression and culture supernatant cytotoxicity from the end of the vegetative phase up to 5 h after entry into the stationary phase. The concentration of each of the 22 proteins in the culture supernatant was determined at various times. In addition, the stability of the proteins was studied. Fifteen of these proteins, including 14 members of the virulence regulon PlcR, were known or predicted to be secreted. All of the secreted proteins reached a maximum concentration during early stationary phase, but there were significant differences in the kinetics of their concentrations. The time courses of protein concentrations were in agreement with gene expression data, except for cytotoxin CytK, which was unstable, and for the metalloprotease InhA1. Supernatant cytoxicity also peaked in early stationary phase, and the kinetics of cytotoxicity paralleled the time course of concentration of the PlcR-controlled toxin, CytK. Our concomitant study of the time course of protein concentrations, gene expression and supernatant cytotoxicity reveals that the pathogenic potential of B. cereus peaks during the transition state. It also suggests that there is diversity in the regulation of gene expression within the PlcR regulon.

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Available from: Stéphane Perchat, Aug 28, 2014
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    • "In the early stationary phase, B. cereus produces several extracellular compounds (degradative enzymes, cytotoxic factors and cell surface proteins) that might contribute to virulence [8], [9], [10], [11], [12]. However, the mechanisms leading to the various pathologies of B. cereus are not completely elucidated. "
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    ABSTRACT: Bacillus cereus is a Gram-positive spore-forming bacterium causing food poisoning and serious opportunistic infections. These infections are characterized by bacterial accumulation despite the recruitment of phagocytic cells. We have previously shown that B. cereus Haemolysin II (HlyII) induces macrophage cell death by apoptosis. In this work, we investigated the regulation of the hlyII gene. We show that HlyIIR, the negative regulator of hlyII expression in B. cereus, is especially active during the early bacterial growth phase. We demonstrate that glucose 6P directly binds to HlyIIR and enhances its activity at a post-transcriptional level. Glucose 6P activates HlyIIR, increasing its capacity to bind to its DNA-box located upstream of the hlyII gene, inhibiting its expression. Thus, hlyII expression is modulated by the availability of glucose. As HlyII induces haemocyte and macrophage death, two cell types that play a role in the sequestration of nutrients upon infection, HlyII may induce host cell death to allow the bacteria to gain access to carbon sources that are essential components for bacterial growth.
    PLoS ONE 02/2013; 8(2):e55085. DOI:10.1371/journal.pone.0055085 · 3.23 Impact Factor
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    • "Assim, os autores sugerem que a metaloprotease InhA2 é essencial para proporcionar um efeito sinergístico dos esporos de B. thuringiensis sobre a toxicidade da proteína Cry1C contra G. mellonella após a infecção por via oral. Uma terceira proteína, InhA3, 72% idêntica a InhA1, também é encontrado no sobrenadante da cultura de B. cereus, mas apenas em quantidades residuais (Gilois et al. 2007). Sua função e regulação ainda não foram descritas. "
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    ABSTRACT: e-ISSN 1983-0572 Publicação do Projeto Entomologistas do Brasil Distribuído através da Creative Commons Licence v3.0 (BY-NC-ND) Resumo. As proteínas Cry produzidas pela bactéria entomopatogênica Bacillus thuringiensis Berliner são bem conhecidas devido a alta citotoxicidade que exibem a uma variedade de insetos-alvo. O modo de ação destas proteínas é específico e torna os produtos à base de B. thuringiensis os mais amplamente utilizados em programas de controle biológico de pragas na agricultura e de importantes vetores de doenças humanas. Contudo, embora as proteínas Cry sejam os fatores de virulência inseto-específico mais conhecidos, linhagens de B. thuringiensis apresentam também uma ampla gama de fatores de virulência, os quais permitem à bactéria atingir a hemolinfa e colonizar eficientemente o inseto hospedeiro. Dentre estes fatores, destacam-se as proteínas Vip, Cyt, enterotoxinas, hemolisinas, fosfolipases, proteases, enzimas de degradação, além das recentemente descritas parasporinas. Essa revisão aborda a ação desses fatores de virulência, bem como a caracterização e o controle da expressão de seus genes. Adicionalmente, são discutidos aspectos relacionados ao nicho ecológico da bactéria com ênfase nas características envolvidas com a biossegurança da utilização dos produtos à base de B. thuringiensis para o controle biológico de insetos-alvo. Abstract. The Cry proteins produced by the entomopathogenic bacterium Bacillus thuringiensis Berliner are widely known due to its high toxicity against a variety of insects. The mode of action of these proteins is specific and becomes B. thuringiensis-based products the most used in biological control programs of insect pests in agriculture and of important human disease vectors. However, while the Cry proteins are the best-known insect-specific virulence factor, strains of B. thuringiensis show also a wide range of other virulence factors, which allow the bacteria to achieve the hemolymph and colonize efficiently the insect host. Among these factors, we highlight the Vip proteins, Cyt, enterotoxins, hemolysins, phospholipases, proteases and enzymes of degradation, in addition to the recently described parasporin. This review explores the action of these virulence factors, as well as, the characterization and control of expression of their genes. Additionally, we discuss aspects related to the ecological niche of the bacteria with emphasis on the characteristics involved in the biosafety of the use of B. thuringiensis-based products for biological control of target insects.
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    • "In addition, the supernatants of B. cepacia cultures obtained at different time-points of growth were initially demonstrated to exert cytotoxic effects on the mammalian A549 human lung epithelial cells. The cytotoxic effects apparently increased with time, suggesting that this might be due to the increasing amounts and/or types of cytotoxic proteins that accumulated in the culture supernatant during the growth cycle [28]. The variation in cytotoxic potency of supernatants collected at the different growth phases, particularly the mid-logarithmic and early-stationary phases, suggests differences in the pathogenic potentials of B. cepacia to infect and cause disease in man. "
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    ABSTRACT: Burkholderia cepacia is a Gram-negative pathogen that causes serious respiratory infections in immunocompromised patients and individuals with cystic fibrosis. This bacterium is known to release extracellular proteins that may be involved in virulence. In the present study, B. cepacia grown to mid-logarithmic and early-stationary phases were investigated on their ability to invade and survive intracellularly in A549 lung epithelial cells in order to discern the fate of these bacteria in the pathogenesis of B. cepacia lung infections in in vitro condition. The early-stationary phase B. cepacia was demonstrated to be more invasive than mid-logarithmic phase. In addition, culture supernatants of B. cepacia obtained from these phases of growth were also demonstrated to cause different cytotoxic potency on the A549 human lung epithelial cells. Profiling of the supernatants using the gel-based proteomics approach identified 43 proteins that were commonly released in both the growth phases and 40 proteins newly-released at the early-stationary phase. The latter proteins may account for the higher cytotoxic activity of the early-stationary culture supernatant compared to that obtained at the mid-logarithmic phase. Among the newly-released proteins in the early-stationary phase supernatant were flagellar hook-associated domain protein (FliD), flagellar hook-associated protein (FlgK), TonB-dependent siderophore (Fiu), Elongation factor G (FusA), phosphoglycerate kinase (Pgk) and sulfatase (AslA) which are known for their virulence. Differences in the ability of B. cepacia to invade and survive intracellularly inside the epithelial cells at different phases of growth may improve our understanding of the varied disease progressions associated with B. cepacia infections. In addition, the identified culture supernatant proteins may be used as targets for the development of new strategies to control B. cepacia infection using agents that can block their release.
    PLoS ONE 10/2011; 6(10):e26518. DOI:10.1371/journal.pone.0026518 · 3.23 Impact Factor
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