Plasmid-encoded autolysin in Bacillus anthracis: modular structure and catalytic properties.

A Bacillus anthracis virulence plasmid-encoded peptidoglycan hydrolase (AmiA) with sequence similarity to N-acetylmuramoyl-L-alanine amidases hydrolyzes peptidoglycan independently of cell wall binding. Residues H341, E355, H415, and E486 are absolutely required for catalysis. Many AmiA paralogs are fused to different sorting signals, suggesting that these modular proteins result from domain shuffling.     

Secreted Bacillus anthracis proteases target the host fibrinolytic system

The fibrinolytic system is often the target for pathogenic bacteria, resulting in increased fibrinolysis, bacterial dissemination, and inflammation. The purpose of this study was to explore whether proteases NprB and InhA secreted by Bacillus anthracis could activate the host's fibrinolytic system. NprB efficiently activated human pro-urokinase plasminogen activator (pro-uPA), a key protein in the fibrinolytic cascade. Conversely, InhA had little effect on pro-uPA. Plasminogen activator inhibitors (PAI)-1, 2 and the uPA receptor were also targets for NprB in vitro. InhA efficiently degraded the thrombin-activatable fibrinolysis inhibitor (TAFI) in vitro. Mice infected with B. anthracis showed a significant decrease in blood TAFI levels. In another mouse experiment, animals infected with isogenic inhA deletion mutants restored TAFI levels, while the levels in the parent strain decreased. We propose that NprB and InhA may contribute to the activation of the fibrinolytic system in anthrax infection.     

Serum albumin disrupts Cryptococcus neoformans and Bacillus anthracis extracellular vesicles

For both pathogenic fungi and bacteria, extracellular vesicles have been shown to contain many microbial components associated with virulence, suggesting a role in pathogenesis. However, there are many unresolved issues regarding vesicle synthesis and stability, including the fact that vesicular packaging for extracellular factors involved in virulence must also have a mechanism for vesicle unloading. Consequently, we studied the kinetics of vesicle production and stability using [1-(14) C] palmitic acid metabolic labelling and dynamic light scattering techniques. Cryptococcus neoformans vesicles were produced throughout all stages of fungal culture growth and they were stable once isolated. Density gradient analysis revealed that only a portion of the vesicle population carried cryptococcal polysaccharide, implying heterogeneity in vesicular cargo. Vesicle incubation with macrophages resulted in rapid vesicle instability, a phenomenon that was ultimately associated with serum albumin. Additionally, albumin, along with mouse serum and murine immunoglobulin destabilized Bacillus anthracis vesicles, but the effect was not observed with ovalbumin or keyhole limpet haemocyanin, demonstrating that this phenomenon is neither host-, microbe- nor protein-specific. Our findings strongly suggest that cryptococcal vesicles are short-lived in vivo and vesicle destabilization is mediated by albumin. The ability of albumin to promote vesicular offload through destabilization indicates a new activity for this abundant serum protein.     

Bovine Bacillus anthracis in Cameroon

Bovine Bacillus anthracis isolates from Cameroon were genetically characterized. They showed a strong homogeneity, and they belong, together with strains from Chad, to cluster Aβ, which appears to be predominant in western Africa. However, one strain that belongs to a newly defined clade (D) and cluster (D1) is penicillin resistant and shows certain phenotypes typical of Bacillus cereus.     

Construction of a Bacillus subtilis double mutant deficient in extracellular alkaline and neutral proteases

A mutant strain of Bacillus subtilis carrying lesions in the structural genes for extracellular neutral (nprE) and serine (aprA) proteases was constructed by the gene conversion technique. This mutant had less than 4% of the extracellular protease activity of the wild type and sporulated normally, indicating that neither of these sporulation-associated proteases is essential for development.     

Molecular diversity in Bacillus anthracis

Molecular typing of Bacillus anthracis has been extremely difficult due to the lack of polymorphic DNA markers. We have identified nine novel variable number tandemly repeated loci from previously known amplified fragment length polymorphism markers or from the DNA sequence. In combination with the previously known vrrA locus, these markers provide discrimination power to genetically characterize B. anthracis isolates. The variable number tandem repeat (VNTR) loci are found in both gene coding (genic) and non-coding (non-genic) regions. The genic differences are 'in frame' and result in additions or deletion of amino acids to the predicted proteins. Due the rarity of molecular differences, the VNTR changes represent a significant portion of the genetic variation found within B. anthracis. This variation could represent an important adaptive mechanism. Marker similarity and differences among diverse isolates have identified seven major diversity groups that may represent the only world-wide B. anthracis clones. The lineages reconstructed using these data may reflect the dispersal and evolution of this pathogen.     

Production of Bacillus anthracis protective antigen is dependent on the extracellular chaperone,PrsA

Protective antigen (PA) is a component of the Bacillus anthracis lethal and edema toxins and the basis of the current anthrax vaccine. In its heptameric form, PA targets host cells and internalizes the enzymatically active components of the toxins, namely lethal and edema factors. PA and other toxin components are secreted from B. anthracis using the Sec-dependent secretion pathway. This requires them to be translocated across the cytoplasmic membrane in an unfolded state and then to be folded into their native configurations on the trans side of the membrane, prior to their release from the environment of the cell wall. In this study we show that recombinant PA (rPA) requires the extracellular chaperone PrsA for efficient folding when produced in the heterologous host, B. subtilis; increasing the concentration of PrsA leads to an increase in rPA production. To determine the likelihood of PrsA being required for PA production in its native host, we have analyzed the B. anthracis genome sequence for the presence of genes encoding homologues of B. subtilis PrsA. We identified three putative B. anthracis PrsA proteins (PrsAA, PrsAB, and PrsAC) that are able to complement the activity of B. subtilis PrsA with respect to cell viability and rPA secretion, as well as that of AmyQ, a protein previously shown to be PrsA-dependent.     

炭疽杆菌致病性研究进展

炭疽杆菌是人类历史上第一个被发现的病原菌。炭疽杆菌的研究在近几年取得了较大进展 ,特别是本年度其基因组序列测定已完成并向全世界公布 ,进一步深化了对炭疽杆菌的研究。炭疽杆菌致病性的研究一直是炭疽杆菌研究的重点 ,近年来此方面的研究取得了很多新进展 ,从基因组、致病物质及致病机制 3个方面对此作一个简单的介绍。     

The Bacillus anthracis spore

In response to starvation, Bacillus anthracis can form a specialized cell type called the spore, which is the infectious particle for the disease anthrax. The spore is largely metabolically inactive and can resist a wide range of stresses found in nature. In spite of its dormancy, the spore can sense the presence of nutrient and rapidly return to vegetative growth. These properties help the spore to persist for long periods of time in the environment, survive host defenses after entering the body, and cause disease when the correct location in the host is reached. The anatomy of the spore is unique among bacteria, being comprised of a series of specialized concentric shells, each of which provides specific critical functions. Surrounding the spore core (which houses the chromosome) is a peptidoglycan layer important for spore dormancy, a protein shell that resists a variety of toxic molecules, and finally an exterior protein and glycoprotein layer that, among other functions, mediates interactions with surfaces, including those encountered by the spore within the host. Detailed molecular analysis of these shells has shed considerable light on how each layer determines specific spore properties. Future work, especially on the outermost spore layer, is likely to advance therapeutics, methods for spore decontamination and other critical biodefense technologies.