Toxin-Independent Virulence of Bacillus anthracis in Rabbits

The accepted paradigm states that anthrax is both an invasive and toxinogenic disease and that the toxins play a major role in pathogenicity. In the guinea pig (GP) model we have previously shown that deletion of all three toxin components results in a relatively moderate attenuation in virulence, indicating that B. anthracis possesses an additional toxin-independent virulence mechanism. To characterize this toxin-independent mechanism in anthrax disease, we developed a new rabbit model by intravenous injection (IV) of B. anthracis encapsulated vegetative cells, artificially creating bacteremia. Using this model we were able to demonstrate that also in rabbits, B. anthracis mutants lacking the toxins are capable of killing the host within 24 hours. This virulent trait depends on the activity of AtxA in the presence of pXO2, as, in the absence of the toxin genes, deletion of either component abolishes virulence. Furthermore, this IV virulence depends mainly on AtxA rather than the whole pXO1. A similar pattern was shown in the GP model using subcutaneous (SC) administration of spores of the mutant strains, demonstrating the generality of the phenomenon. The virulent strains showed higher bacteremia levels and more efficient tissue dissemination; however our interpretation is that tissue dissemination per se is not the main determinant of virulence whose exact nature requires further elucidation.     

The Two CcdA Proteins of Bacillus anthracis Differentially Affect Virulence Gene Expression and Sporulation

The cytochrome c maturation system influences the expression of virulence factors in Bacillus anthracis. B. anthracis carries two copies of the ccdA gene, encoding predicted thiol-disulfide oxidoreductases that contribute to cytochrome c maturation, while the closely related organism Bacillus subtilis carries only one copy of ccdA. To investigate the roles of the two ccdA gene copies in B. anthracis, strains were constructed without each ccdA gene, and one strain was constructed without both copies simultaneously. Loss of both ccdA genes results in a reduction of cytochrome c production, an increase in virulence factor expression, and a reduction in sporulation efficiency. Complementation and expression analyses indicate that ccdA2 encodes the primary CcdA in B. anthracis, active in all three pathways. While CcdA1 retains activity in cytochrome c maturation and virulence control, it has completely lost its activity in the sporulation pathway. In support of this finding, expression of ccdA1 is strongly reduced when cells are grown under sporulation-inducing conditions. When the activities of CcdA1 and CcdA2 were analyzed in B. subtilis, neither protein retained activity in cytochrome c maturation, but CcdA2 could still function in sporulation. These observations reveal the complexities of thiol-disulfide oxidoreductase function in pathways relevant to virulence and physiology.     

Comparison of Growth and Toxin Production in Two Vaccine Strains of Bacillus anthracis

Two vaccine strains of Bacillus anthracis were monitored in a 10-liter fermentor to compare growth patterns and toxin production. Under identical conditions, the Sterne strain produced all three components of anthrax toxin, whereas strain V770 produced only the protective antigen.     

The Capsule and S-Layer: Two Independent and Yet Compatible Macromolecular Structures in Bacillus anthracis

Bacillus anthracis, the etiological agent of anthrax, is a gram-positive spore-forming bacterium. Fully virulent bacilli are toxinogenic and capsulated. Two abundant surface proteins, including the major antigen, are components of the B. anthracis surface layer (S-layer). The B. anthracis paracrystalline S-layer has previously only been found in noncapsulated vegetative cells. Here we report that the S-layer proteins are also synthesized under conditions where the poly-γ-d-glutamic acid capsule is present. Structural and immunological analyses show that the capsule is exterior to and completely covers the S-layer proteins. Nevertheless, analysis of single and double S-layer protein mutants shows that the presence of these proteins is not required for normal capsulation of the bacilli. Similarly, the S-layer proteins assemble as a two-dimensional crystal, even in the presence of the capsule. Thus, both structures are compatible, and yet neither is required for the correct formation of the other.

Bacillus anthracis, a gram-positive spore-forming bacterium, is the causative agent of anthrax. This disease, to which many animals, including humans, are susceptible, involves toxemia and septicemia. In the mammalian host, B. anthracis bacilli synthesize two toxins (lethal and edema toxins) (31) and a capsule (18) encoded by two large plasmids, pXO1 and pXO2, respectively (12, 21). The capsule is composed of poly-γ-d-glutamic acid and has antiphagocytic properties (13, 31, 37). Although unusual, a similar capsule is also found on Bacillus licheniformis bacilli (9). In the absence of pXO2 or the inducer bicarbonate, the cell does not produce a capsule and the cell wall appears layered. These layers are composed of fragments displaying a highly patterned ultrastructure (10, 16). This type of cell surface is now referred to as the surface layer (S-layer).S-layers are present on the surfaces of many archaea and bacteria (for reviews, see references 29 and 30). Most are formed by noncovalent, entropy-driven assembly of a single (glyco)protein protomer on the bacterial surface, giving rise to proteinaceous paracrystalline layers. Generally, a single S-layer is present, constituting 5 to 10% of total cell protein. Its synthesis is thus presumably energy consuming for the bacterium. Numerous bacteria have S-layers, suggesting that they play important roles in the interaction between the cell and its environment. Various functions have been proposed for S-layers, including shape maintenance and molecular sieving, and they can serve in phage fixation. The S-layer may be a virulence factor, protecting pathogenic bacteria against complement killing, facilitating binding of bacteria to host molecules, or enhancing their ability to associate with macrophages (for reviews, see references 27 and 29).Some bacteria, such as cyanobacteria or Azotobacter spp., possess both a capsule and an S-layer; however, to our knowledge, their structural relationships have not been analyzed through simultaneous genetic and cytologic studies. Both of these features have been independently described for the surface of the pathogenic bacterium B. anthracis. The components of the B. anthracis S-layer are two abundant surface proteins, EA1 and Sap (6, 20). Previous analyses of the B. anthracis S-layer used plasmid-cured strains; consequently, the interaction, if any, between the capsule and the S-layer could not be studied. Temporal or environmental regulation could be such that only one or the other structure is ever present at the cell surface. However, we show that S-layer proteins are synthesized under conditions where the bacilli are capsulated. We determined the localizations of capsule and S-layer components and analyzed whether the S-layer is necessary for proper capsulation. Finally, the assembly of the S-layer proteins in a two-dimensional crystal was examined in the presence of the capsule.     

Rho与中枢神经系统轴突再生

Rho是小分子质量GTP酶Rho家族成员,在细胞的一些信号转导途径中起着分子开关的作用.Rho能通过作用于肌动蛋白骨架系统引起轴突生长锥塌陷,从而抑制轴突生长.研究表明,Nogo-A、MAG、OMgp等髓鞘源性的轴突再生抑制分子均可通过激活Rho介导的信号转导途径抑制轴突再生.     

The Glycosphingolipid Hydrolases in the Central Nervous System

Glycosphingolipids are a large group of complex lipids particularly abundant in the outer layer of the neuronal plasma membranes. Qualitative and quantitative changes in glycosphingolipids have been reported along neuronal differentiation and aging. Their half-life is short in the nervous system and their membrane composition and content are the result of a complex network of metabolic pathways involving both the de novo synthesis in the Golgi apparatus and the lysosomal catabolism. In particular, most of the enzymes of glycosphingolipid biosynthesis and catabolism have been found also at the plasma membrane level. Their action could be responsible for the fine tuning of the plasma membrane glycosphingolipid composition allowing the formation of highly specialized membrane areas, such as the synapses and the axonal growth cones. While the correlation between the changes of GSL pattern and the modulation of the expression/activity of different glycosyltransferases during the neuronal differentiation has been widely discussed, the role of the glycohydrolytic enzymes in this process is still little explored. For this reason, in the present review, we focus on the main glycolipid catabolic enzymes β-hexosaminidases, sialidases, β-galactosidases, and β-glucocerebrosidases in the process of the neuronal differentiation.     

Stereotaxic Injection of Tetanus Toxin in Rat Central Nervous System Causes Alteration in Normal Levels of Monoamines

A single intraventricular injection of tetanus toxin produced a time-dependent elevation of serotonin levels in brain and spinal cord of adult rats. This tetanus toxin-induced increase was produced in areas of high density of serotonergic innervation, such as the hypothalamus, hippocampus, and spinal cord. Little or no effect was found in the thalamus, cerebellum, and frontal cortex, areas that are poorly innervated by serotonergic terminals. The responses of catecholamines (no change in dopamine level and generalized decrease in norepinephrine) pointed to a specific action of tetanus toxin on the serotonergic system. Stereotaxic injections of tetanus toxin in dorsal or magnus raphe nuclei did not have an evident effect on biogenic amine levels in the brain and spinal cord, respectively. Because direct stereotaxic injections of the toxin in the hypothalamus or hippocampus produced significant serotonin increases in both areas, it is proposed that tetanus toxin interacts with presynaptic targets to produce serotonin accumulation; this is probably due in part to an activation of tryptophan 5-hydroxylase.     

腺苷的中枢作用

腺苷是包括中枢神经系统（ＣＮＳ）细胞外液在内的体液的正常组成成分,其正常水平为０．０３～０．３μｍｏｌ／Ｌ。ＡＴＰ合成与分解失衡的条件下明显升高,如缺血时可升高１０００倍之多。腺苷通过腺苷受体（ａｄｅｎｏｄｉｎｅｒｅｃｅｐｔｏｒ,ＡＲ）对ＣＮＳ具有多方面的生理与病理作用,被认为是ＣＮＳ的抑制性神经调质,具有神经保护作用。     

miR-124: A Promising Therapeutic Target for Central Nervous System Injuries and Diseases

Cellular and Molecular Neurobiology - Central nervous system injuries and diseases, such as ischemic stroke, spinal cord injury, neurodegenerative diseases, glioblastoma, multiple sclerosis, and...     

The genome and variation of Bacillus anthracis

The Bacillus anthracis genome reflects its close genetic ties to Bacillus cereus and Bacillus thuringiensis but has been shaped by its own unique biology and evolutionary forces. The genome is comprised of a chromosome and two large virulence plasmids, pXO1 and pXO2. The chromosome is mostly co-linear among B. anthracis strains and even with the closest near neighbor strains. An exception to this pattern has been observed in a large inversion in an attenuated strain suggesting that chromosome co-linearity is important to the natural biology of this pathogen. In general, there are few polymorphic nucleotides among B. anthracis strains reflecting the short evolutionary time since its derivation from a B. cereus-like ancestor. The exceptions to this lack of diversity are the variable number tandem repeat (VNTR) loci that exist in genic and non genic regions of the chromosome and both plasmids. Their variation is associated with high mutability that is driven by rapid insertion and deletion of the repeats within an array. A notable example is found in the vrrC locus which is homologous to known DNA translocase genes from other bacteria.     

Host–Pathogen Coevolution: The Selective Advantage of Bacillus thuringiensis Virulence and Its Cry Toxin Genes

Reciprocal coevolution between host and pathogen is widely seen as a major driver of evolution and biological innovation. Yet, to date, the underlying genetic mechanisms and associated trait functions that are unique to rapid coevolutionary change are generally unknown. We here combined experimental evolution of the bacterial biocontrol agent Bacillus thuringiensis and its nematode host Caenorhabditis elegans with large-scale phenotyping, whole genome analysis, and functional genetics to demonstrate the selective benefit of pathogen virulence and the underlying toxin genes during the adaptation process. We show that: (i) high virulence was specifically favoured during pathogen–host coevolution rather than pathogen one-sided adaptation to a nonchanging host or to an environment without host; (ii) the pathogen genotype BT-679 with known nematocidal toxin genes and high virulence specifically swept to fixation in all of the independent replicate populations under coevolution but only some under one-sided adaptation; (iii) high virulence in the BT-679-dominated populations correlated with elevated copy numbers of the plasmid containing the nematocidal toxin genes; (iv) loss of virulence in a toxin-plasmid lacking BT-679 isolate was reconstituted by genetic reintroduction or external addition of the toxins. We conclude that sustained coevolution is distinct from unidirectional selection in shaping the pathogen''s genome and life history characteristics. To our knowledge, this study is the first to characterize the pathogen genes involved in coevolutionary adaptation in an animal host–pathogen interaction system.     

外泌体及其在中枢神经系统中的功能

外泌体是一类由细胞分泌至胞外的囊泡,生物发生主要涉及细胞质膜的两次内陷、多囊泡体的形成以及外泌体的释放。外泌体具有丰富多样的内含物,包括一些标志性膜蛋白、可溶蛋白、各类RNA分子和DNA片段等。细胞可以通过分泌和接受外泌体来实现细胞间的信号交流,外泌体通过膜上携带的配体分子与其他细胞质膜表面的受体相互作用,从而激活细胞的信号转导或与受体细胞质膜发生融合释放内容物进入胞质来发挥调节功能。在中枢神经系统中,神经元及各类神经胶质细胞分泌的神经外泌体可以介导布线式的突触信号传递,但主要还是以容积传递的方式发挥类似神经调质的功能。本文详细阐述了外泌体的生物发生过程及部分重要的功能性成分,就神经外泌体在发生、内容物分选和受控释放三个方面的特性与突触囊泡进行比较,总结了神经外泌体在中枢神经系统中发挥的生理功能及其在神经退行性疾病和抑郁症发生、发展中作用的研究进展,并对外泌体在神经系统疾病早期诊断及靶向治疗方面的应用前景进行了展望。