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Virulence Determinants (virulence + determinant)
Kinds of Virulence Determinants Selected AbstractsApplication of proton nuclear magnetic resonance spectroscopy to the study of Cryptococcus and cryptococcosisFEMS YEAST RESEARCH, Issue 4 2006Tania C. Sorrell Abstract Proton nuclear magnetic resonance spectroscopy is a nondestructive technique that identifies chemicals in solution and in living cells. It has been used in cryptococcal research to identify the primary structure of capsular glucuronoxylomannans, link cellular apoptosis susceptibility (CAS) genes to positioning of residues on the mannose backbone of glucuronoxylomannan, and verify that the cryptococcal virulence determinant, phospholipase B, is elaborated in vivo. Promising clinical applications include speciation (Cryptococcus neoformans and Cryptococcus gattii), with preliminary evidence that varieties neoformans and grubii can also be distinguished, non-invasive diagnosis of cerebral cryptococcomas, and, in cases of meningitis, monitoring therapeutic response by analysis of cerebrospinal fluid. [source] The Versatility of Helicobacter pylori CagA Effector Protein Functions: The Master Key HypothesisHELICOBACTER, Issue 3 2010Steffen Backert Abstract Several bacterial pathogens inject virulence proteins into host target cells that are substrates of eukaryotic tyrosine kinases. One of the key examples is the Helicobacter pylori CagA effector protein which is translocated by a type-IV secretion system. Injected CagA becomes tyrosine-phosphorylated on EPIYA sequence motifs by Src and Abl family kinases. CagA then binds to and activates/inactivates multiple signaling proteins in a phosphorylation-dependent and phosphorylation-independent manner. A recent proteomic screen systematically identified eukaryotic binding partners of the EPIYA phosphorylation sites of CagA and similar sites in other bacterial effectors by high-resolution mass spectrometry. Individual phosphorylation sites recruited a surprisingly high number of interaction partners suggesting that each phosphorylation site can interfere with many downstream pathways. We now count 20 reported cellular binding partners of CagA, which represents the highest quantitiy among all yet known virulence-associated effector proteins in the microbial world. This complexity generates a highly remarkable and puzzling scenario. In addition, the first crystal structure of CagA provided us with new information on the function of this important virulence determinant. Here we review the recent advances in characterizing the multiple binding signaling activities of CagA. Injected CagA can act as a ,master key' that evolved the ability to highjack multiple host cell signalling cascades, which include the induction of membrane dynamics, actin-cytoskeletal rearrangements and the disruption of cell-to-cell junctions as well as proliferative, pro-inflammatory and anti-apoptotic nuclear responses. The discovery that different pathogens use this common strategy to subvert host cell functions suggests that more examples will emerge soon. [source] The Maurer's cleft protein MAHRP1 is essential for trafficking of PfEMP1 to the surface of Plasmodium falciparum -infected erythrocytesMOLECULAR MICROBIOLOGY, Issue 5 2008Cornelia Spycher Summary During the intra-erythrocytic development of Plasmodium falciparum, the parasite modifies the host cell surface by exporting proteins that interact with or insert into the erythrocyte membrane. These proteins include the principal mediator of cytoadherence, P. falciparum erythrocyte membrane protein 1 (PfEMP1). To implement these changes, the parasite establishes a protein-trafficking system beyond its confines. Membrane-bound structures called Maurer's clefts are intermediate trafficking compartments for proteins destined for the host cell membrane. We disrupted the gene for the membrane-associated histidine-rich protein 1 (MAHRP1). MAHRP1 is not essential for parasite viability or Maurer's cleft formation; however, in its absence, these organelles become disorganized in permeabilized cells. Maurer's cleft-resident proteins and transit cargo are exported normally in the absence of MAHRP1; however, the virulence determinant, PfEMP1, accumulates within the parasite, is depleted from the Maurer's clefts and is not presented at the red blood cell surface. Complementation of the mutant parasites with mahrp1 led to the reappearance of PfEMP1 on the infected red blood cell surface, and binding studies show that PfEMP1-mediated binding to CD36 is restored. These data suggest an important role of MAHRP1 in the translocation of PfEMP1 from the parasite to the host cell membrane. [source] Identification of Candida albicans genes induced during thrush offers insight into pathogenesisMOLECULAR MICROBIOLOGY, Issue 5 2003Shaoji Cheng Summary Candida albicans causes a wide spectrum of diseases, ranging from mucocutaneous infections like oral thrush to disseminated candidiasis. Screening for C. albicans genes expressed within infected hosts might advance understanding of candidal pathogenesis, but is impractical using existing techniques. In this study, we used an antibody-based strategy to identify C. albicans genes expressed during thrush. We adsorbed sera from HIV-infected patients with thrush against candidal cells grown in vitro and screened a C. albicans genomic expression library. We identified 10 genes encoding immunogenic antigens and used reverse transcription-polymerase chain reaction to verify that they were induced within thrush pseudomembranes recovered from a patient. The in vivo induced genes are involved in diverse functions, including regulation of yeast-hyphal morphogenesis, adhesion to host cells, nutrient uptake, phospholipid biosynthesis and amino acid catabolism. Four genes encode known virulence determinants (HWP1, CST20, CPP1 and RBF1). Another gene, LPD1, for which a role in candidal pathogenesis is unknown, encodes a protein homologous to a bacterial virulence determinant. Most importantly, disruption of CaNOT5, a newly identified gene, conferred defects in morphogenesis, decreased adherence to human buccal epithelial cells and attenuated mortality during murine disseminated candidiasis, proving that our strategy can identify genes encoding novel virulence determinants. [source] Microplitis demolitor bracovirus inhibits phagocytosis by hemocytes from Pseudoplusia includens,ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 3 2006Michael R. Strand Abstract The braconid wasp Microplitis demolitor carries Microplitis demolitor bracovirus (MdBV) and parasitizes the larval stage of several noctuid moths. A key function of MdBV in parasitism is suppression of the host's cellular immune response. Prior studies in the host Pseudoplusia includens indicated that MdBV blocks encapsulation by preventing two types of hemocytes, plasmatocytes and granulocytes, from adhering to foreign targets. The other main immune response mediated by insect hemocytes is phagocytosis. The goal of this study was to determine which hemocyte types were phagocytic in P. includens and to assess whether MdBV infection affects this defense response. Using the bacterium Escherichia coli and inert polystyrene beads as targets, our results indicated that the professional phagocyte in P. includens is granulocytes. The phagocytic responses of granulocytes were very similar to those of High Five cells that prior studies have suggested are a granulocyte-like cell line. MdBV infection dose-dependently disrupted phagocytosis in both cell types by inhibiting adhesion of targets to the cell surface. The MdBV glc1.8 gene encodes a cell surface glycoprotein that had previously been implicated in disruption of adhesion and encapsulation responses by immune cells. Knockdown of glc1.8 expression by RNA interference (RNAi) during the current study rescued the ability of MdBV-infected High Five cells to phagocytize targets. Collectively, these results indicate that glc1.8 is a key virulence determinant in disruption of both adhesion and phagocytosis by insect immune cells. Arch. Insect Biochem. Physiol. 61:134,145, 2006. © 2006 Wiley-Liss, Inc. [source] Critical determinants of the interactions of capsule-expressing Neisseria meningitidis with host cells: the role of receptor density in increased cellular targeting via the outer membrane Opa proteinsCELLULAR MICROBIOLOGY, Issue 10 2005Christopher J. Bradley Summary Neisseria meningitidis capsule is an important virulence determinant required for survival in the blood but is reportedly involved in inhibiting cellular interactions mediated by meningococcal outer membrane adhesins. However, evidence from our previous studies suggested that target receptor density on host cells may determine whether or not capsulate bacteria can adhere via outer membrane proteins such as Opa. To confirm this and evaluate the impact of capsulation on bacterial interactions, we used Opa+ and Opa, derivatives of capsulate and acapsulate meningococcal isolates and transfected cell lines expressing CEACAM1, a receptor targeted by Opa proteins. To assess the extent and rate of cell association, subpopulations of stably transfected Chinese hamster ovary cells with different receptor levels were derived. A quantitative correlation of CEACAM1 levels and Opa-dependent binding of both capsulate and acapsulate bacteria was demonstrated, which was accelerated at high receptor densities. However, it appears that invasion by Opa+ capsulate bacteria only occurs when a threshold level of CEACAM density has been reached. Target cells expressing high levels of CEACAM1 (MFI c. 400) bound threefold more, but internalized 20-fold more Opa+ capsulate bacteria than those with intermediate expression (MFI c. 100). No overall selection of acapsulate phenotype was observed in the internalized population. These observations confirm that capsule may not be an adequate barrier for cellular interactions and demonstrate the role of a host factor that may determine capsulate bacterial invasion potential. Upregulation of CEACAMs, which can occur in response to inflammatory cytokines, could lead to translocation of a small number of fully capsulate bacteria across mucosal epithelium into the bloodstream sufficient to cause a rapid onset of disseminated disease. Thus the data also suggest a novel rationale for the epidemiological observations that individuals with prior infectious/inflammatory conditions carry a high risk of invasive meningococcal disease. [source] Effects of clinical isolates of Pseudomonas aeruginosa on Staphylococcus epidermidis biofilm formationFEMS IMMUNOLOGY & MEDICAL MICROBIOLOGY, Issue 3 2010Maria Pihl Abstract Pseudomonas aeruginosa is often found in chronic infections, including cystic fibrosis lung infections and those related to chronic wounds and venous ulcers. At the latter sites, P. aeruginosa can be isolated together with Staphylococcus epidermidis, and we have therefore explored the effect of clinical isolates and laboratory strains of P. aeruginosa strains on colonization by S. epidermidis in dual-species biofilms. Biofilm formation was assayed using 16S rRNA FISH and confocal laser scanning microscopy. Among the six P. aeruginosa strains tested, one particular strain, denoted 14:2, exerted a significant inhibitory effect, and even after 6 h, S. epidermidis levels in dual-species biofilms were reduced by >85% compared with those without P. aeruginosa. Interestingly, strain 14:2 was found to be negative for classical virulence determinants including pyocyanin, elastase and alkaline protease. Therefore, we suggest that less virulent phenotypes of P. aeruginosa, which may develop over time in chronic infections, could counteract colonization by S. epidermidis, ensuring persistence and dominance by P. aeruginosa in the host micro-habitat. Further studies are required to explain the inhibitory effect on S. epidermidis, although extracellular polysaccharides produced by P. aeruginosa might play a role in this phenomenon. [source] From soil to gut: Bacillus cereus and its food poisoning toxinsFEMS MICROBIOLOGY REVIEWS, Issue 4 2008Lotte P. Stenfors Arnesen Abstract Bacillus cereus is widespread in nature and frequently isolated from soil and growing plants, but it is also well adapted for growth in the intestinal tract of insects and mammals. From these habitats it is easily spread to foods, where it may cause an emetic or a diarrhoeal type of food-associated illness that is becoming increasingly important in the industrialized world. The emetic disease is a food intoxication caused by cereulide, a small ring-formed dodecadepsipeptide. Similar to the virulence determinants that distinguish Bacillus thuringiensis and Bacillus anthracis from B. cereus, the genetic determinants of cereulide are plasmid-borne. The diarrhoeal syndrome of B. cereus is an infection caused by vegetative cells, ingested as viable cells or spores, thought to produce protein enterotoxins in the small intestine. Three pore-forming cytotoxins have been associated with diarrhoeal disease: haemolysin BL (Hbl), nonhaemolytic enterotoxin (Nhe) and cytotoxin K. Hbl and Nhe are homologous three-component toxins, which appear to be related to the monooligomeric toxin cytolysin A found in Escherichia coli. This review will focus on the toxins associated with foodborne diseases frequently caused by B. cereus. The disease characteristics are described, and recent findings regarding the associated toxins are discussed, as well as the present knowledge on virulence regulation. [source] Regulation of virulence determinants in Staphylococcus aureus: complexity and applicationsFEMS MICROBIOLOGY REVIEWS, Issue 2 2004Stéphane Bronner Abstract The virulence of Staphylococcus aureus is essentially determined by cell wall associated proteins and secreted toxins that are regulated and expressed according to growth phases and/or growth conditions. Gene expression is regulated by specific and sensitive mechanisms, most of which act at the transcriptional level. Regulatory factors constitute numerous complex networks, driving specific interactions with target gene promoters. These factors are largely regulated by two-component regulatory systems, such as the agr, saeRS, srrAB, arlSR and lytRS systems. These systems are sensitive to environmental signals and consist of a sensor histidine kinase and a response regulator protein. DNA-binding proteins, such as SarA and the recently identified SarA homologues (SarR, Rot, SarS, SarT, SarU), also regulate virulence factor expression. These homologues might be intermediates in the regulatory networks. The multiple pathways generated by these factors allow the bacterium to adapt to environmental conditions rapidly and specifically, and to develop infection. Precise knowledge of these regulatory mechanisms and how they control virulence factor expression would open up new perspectives for antimicrobial chemotherapy using key inhibitors of these systems. [source] Phage-selected lipopolysaccharide mutants of Pectobacterium atrosepticum exhibit different impacts on virulenceJOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2010T.J. Evans Abstract Aims:, To positively select Pectobacterium atrosepticum (Pa) mutants with cell surface defects and to assess the impact of these mutations on phytopathogenesis. Methods and Results:, Several phages were isolated from treated sewage effluent and were found to require bacterial lipopolysaccharide (LPS) for infection. Two strains with distinct mutations in LPS were obtained by transposon mutagenesis. Along with a third LPS mutant, these strains were characterized with respect to various virulence-associated phenotypes, including growth rate, motility and exoenzyme production, demonstrating that LPS mutations are pleiotropic. Two of the strains were deficient in the synthesis of the O-antigen portion of LPS, and both were less virulent than the wild type. A waaJ mutant, which has severe defects in LPS biosynthesis, was dramatically impaired in potato tuber rot assays. The infectivity of these novel phages on 32 additional strains of Pa was tested, showing that most Pa isolates were sensitive to the LPS-dependent phages. Conclusions:, Native LPS is crucial for optimal growth, survival and virulence of Pa in vivo, but simultaneously renders such strains susceptible to phage infection. Significance and Impact of the Study:, This work demonstrates the power of phages to select and identify the virulence determinants on the bacterial surface, and as potential biocontrol agents for Pa infections. [source] Listeria monocytogenes in spontaneous abortions in humans and its detection by multiplex PCRJOURNAL OF APPLIED MICROBIOLOGY, Issue 5 2007S. Kaur Abstract Aim: To assess the extent of Listeria monocytogenes in causation of human spontaneous abortions by isolation methods and PCR analysis for the presence of virulence-associated genes. Methods and Results: A total of 305 samples comprising blood, urine, placental bits, faecal and vaginal swabs were collected from 61 patients with spontaneous abortions. Listeria spp. were isolated from 10 samples collected from nine (14·8%) patients. Confirmation of these isolates was based on biochemical tests, haemolysis on blood agar, CAMP test, phosphatidylinositol-specific phospholipase C (PI-PLC) assay followed by in vivo pathogenicity tests and multiplex PCR to detect virulence-associated genes (prfA, plcA, hlyA, actA and iap). Three isolates were confirmed as L. monocytogenes. Of these, two isolates turned out to be pathogenic and found to posses all five genes. However, the remaining two haemolytic L. monocytogenes isolates lacking the plcA gene and activity in the PI-PLC assay were found to be nonpathogenic by in vivo tests. Conclusions: The occurrence of pathogenic L. monocytogenes in cases of spontaneous abortions was 3·3%. It seems that the plcA gene and its expression have an important role as essential virulence determinants in pathogenic Listeria spp. Significance and Impact of the Study: The recovery of pathogenic L. monocytogenes isolates from cases of spontaneous abortion indicates the significance of listeric infection in pregnant women. [source] Safety and Functional Aspects of Preselected Enterococci for Probiotic Use in Iberian Dry-Fermented SausagesJOURNAL OF FOOD SCIENCE, Issue 7 2009Santiago Ruiz-Moyano ABSTRACT:, The purpose of this study was to investigate enterococci for potential probiotic use in Iberian dry-fermented sausages. A total of 15 strains isolated from Iberian dry-fermented sausages, human feces, and pig feces were evaluated for their safety and functional characteristics including biogenic amine (BA) production, antibiotic susceptibility, hemolysis, virulence determinants, cell adhesion, and antimicrobial activity against foodborne pathogens. The strain,Enterococcus faecium,SE906 was able to establish itself on the intestinal epithelium, inhibiting such pathogenic bacteria as,Listeria monocytogenes in vitro. This strain was also considered safe to be used for its low aminogenic potential, and its antibiotic resistance pattern and virulence determinants, being identified as a potential probiotic meat starter culture suitable for manufacture of dry-fermented Iberian sausages. [source] Genotypic characterization of hospital Enterococcus faecalis strains using multiple-locus variable-number tandem-repeat analysisLETTERS IN APPLIED MICROBIOLOGY, Issue 1 2009E. Wa, ecka Abstract Aims:, The level of genetic diversity and relationships between the specific genotypes and the distribution of virulence determinants among Enterococcus faecalis strains isolated from patients hospitalized in different wards of two hospitals were investigated. Methods and Results:, Fifty-six clinical strains of E. faecalis, isolated from patients hospitalized in the period of 1999,2004 in several wards in Wroc,aw (Poland), were analysed by multiple-locus variable-number tandem-repeat analysis (MLVA). Analysis of seven genomic loci identified 40 novel genotypes among the analysed E. faecalis strains, with two major genomic groups, designated I and II, distinguished at a cut-off of 35%. With a similarity cut-off of 85·7%, the genotypes could be combined into 12 clusters (C1,C12), containing at least two isolates. The remaining 18 MLVA types were represented by a single isolate. Conclusions:, Based on the data obtained by MLVA, it was found that (i) many E. faecalis isolates recovered from patients from the wards whose location allowed the potential transmission of micro-organisms, belonged to closely related MLVA types and (ii) possible relationships between specific E. faecalis genotype and the virulence factors lipase, haemolysin and esp gene can exist. Significance and Impact of the Study:, Our study confirms that MLVA is a suitable method for the epidemiological study of E. faecalis and for the first time shows possible relationships between specific genotypes and such virulence determinants, i.e. lipase, haemolysin and esp gene. [source] Persistence of Shiga toxin-producing Escherichia coli O26 in cow slurryLETTERS IN APPLIED MICROBIOLOGY, Issue 1 2007B. Fremaux Abstract Aims: The main objective of this study was to evaluate the growth and survival of Shiga toxin-producing Escherichia coli (STEC) O26 in cow slurry; this serogroup is regarded as an important cause of STEC-associated diseases. Methods and Results: Four STEC were examined by polymerase chain reaction (PCR) to determine whether they harbour key virulence determinants and also by pulsed-field gel electrophoresis (PFGE) to obtain overview fingerprints of their genomes. They were transformed with the pGFPuv plasmid and were separately inoculated at a level of 106 CFU ml,1 in 15 l of cow slurry. All STEC O26 strains could be detected for at least 3 months in cow slurry without any genetic changes. The moisture content of the slurry decreased over time to reach a final value of 75% while the pH increased from 8·5 to 9·5 units during the last 50 days. Conclusion: STEC O26 strains were able to survive in cow slurry for an extended period. Significance and Impact of the Study: Long-term storage of waste slurry should be required to reduce the pathogen load and to limit environmental contamination by STEC O26. [source] Identification and characterization of enterococci from bryndza cheeseLETTERS IN APPLIED MICROBIOLOGY, Issue 6 2006D. Jurkovi Abstract Aims:, To identify enterococci isolated from sheep milk cheese , bryndza, and to compare differences in the composition of enterococcal microflora affected by the season, and to evaluate the potential presence of vancomycin resistance and virulence determinants. Methods and Results:, Bacterial strains were isolated during analysis of bryndza cheese and identified on the genus and species level by phenotypic methods and with commercial biochemical sets. The identification of the species, Enterococcus faecium, Ent. durans and Ent. faecalis, was confirmed by PCR using species-specific primers for ddl genes. PCR was also used for assessment of presence of vanA and vanB genes and virulence determinants gelE, agg and cytolysin genes namely: cylLL, cylLS, cylM, cylB and cylA. Among 308 Enterococcus sp. strains, 177 isolates were proved to be Ent. faecium, 59 to be Ent. durans and 41 to be Ent. faecalis. Vancomycin resistance genes vanA and vanB were not detected. Agar plate testing confirmed their absence. Gene gelE, however, was found in 20 Ent. faecalis isolates, but only 13 of them showed gelatinase-positive phenotype. Seven isolates had five cytolysin genes, but none of the isolates exhibited a positive haemolytic phenotype. Four isolates possessed the agg gene. The prevalence of Ent. faecium species was highest in samples from the winter season harvest. Conclusions:,Ent. faecium is the dominant enterococcal species in bryndza cheese and the most prevalent in the winter season product. None of the Enterococcus sp. strains was proved to have vanA or vanB genes and the vancomycin resistance. Significance and Impact of the Study:, To our knowledge, this is the first report of enterococcal microflora in bryndza cheese and its evaluation for the presence of vanA and vanB genes as well as virulence determinants. [source] Structural analysis of a novel anionic polysaccharide from Porphyromonas gingivalis strain W50 related to Arg-gingipain glycansMOLECULAR MICROBIOLOGY, Issue 3 2005Nikolay Paramonov Summary The Arg-gingipains (RgpsA and B) of Porphyromonas gingivalis are a family of extracellular cysteine proteases and are important virulence determinants of this periodontal bacterium. A monoclonal antibody, MAb1B5, which recognizes an epitope on glycosylated monomeric RgpAs also cross-reacts with a cell-surface polysaccharide of P. gingivalis W50 suggesting that the maturation pathway of the Arg-gingipains may be linked to the biosynthesis of a surface carbohydrate. We report the purification and structural characterization of the cross-reacting anionic polysaccharide (APS), which is distinct from both the lipopolysaccharide and serotype capsule polysaccharide of P. gingivalis W50. The structure of APS was determined by 1D and 2D NMR spectroscopy and methylation analysis, which showed it to be a phosphorylated branched mannan. The backbone is built up of ,-1,6-linked mannose residues and the side-chains contain ,-1,2-linked mannose oligosaccharides of different lengths (one to two sugar residues) attached to the backbone via 1,2-linkage. One of the side-chains in the repeating unit contains Man,1-2Man,1-phosphate linked via phosphorus to a backbone mannose at position 2. De- O -phosphorylation of APS abolished cross-reactivity suggesting that Man,1-2Man,1-phosphate fragment forms part of the epitope recognized by MAb1B5. This phosphorylated branched mannan represents a novel polysaccharide that is immunologically related to the post-translational additions of Arg-gingipains. [source] MicroReview: Envelope stress responses and Gram-negative bacterial pathogenesisMOLECULAR MICROBIOLOGY, Issue 5 2005Tracy L. Raivio Summary The ,E, Cpx and Bae envelope stress responses of Escherichia coli are involved in the maintenance, adaptation and protection of the bacterial envelope in response to a variety of stressors. Recent studies indicate that the Cpx and ,E stress responses exist in many Gram-negative bacterial pathogens. The envelope is of particular importance to these organisms because most virulence determinants reside in, or must transit through, this cellular compartment. The Cpx system has been implicated in expression of pili, type IV secretion systems and key virulence regulators, while the ,E pathway has been shown to be critical for protection from oxidative stress and intracellular survival. Homologues of the ,E, and Cpx-regulated protease DegP are essential for full virulence in numerous pathogens, and, like ,E, DegP appears to confer resistance to oxidative stress and intracellular survival capacity. Some pathogens contain multiple homologues of the Cpx-regulated, disulphide bond catalyst DsbA protein, which has been demonstrated to play roles in the expression of secreted virulence determinants, type III secretion systems and pili. This review highlights recent studies that indicate roles for the ,E, Cpx and Bae envelope stress responses in Gram-negative bacterial pathogenesis. [source] Identification of Candida albicans genes induced during thrush offers insight into pathogenesisMOLECULAR MICROBIOLOGY, Issue 5 2003Shaoji Cheng Summary Candida albicans causes a wide spectrum of diseases, ranging from mucocutaneous infections like oral thrush to disseminated candidiasis. Screening for C. albicans genes expressed within infected hosts might advance understanding of candidal pathogenesis, but is impractical using existing techniques. In this study, we used an antibody-based strategy to identify C. albicans genes expressed during thrush. We adsorbed sera from HIV-infected patients with thrush against candidal cells grown in vitro and screened a C. albicans genomic expression library. We identified 10 genes encoding immunogenic antigens and used reverse transcription-polymerase chain reaction to verify that they were induced within thrush pseudomembranes recovered from a patient. The in vivo induced genes are involved in diverse functions, including regulation of yeast-hyphal morphogenesis, adhesion to host cells, nutrient uptake, phospholipid biosynthesis and amino acid catabolism. Four genes encode known virulence determinants (HWP1, CST20, CPP1 and RBF1). Another gene, LPD1, for which a role in candidal pathogenesis is unknown, encodes a protein homologous to a bacterial virulence determinant. Most importantly, disruption of CaNOT5, a newly identified gene, conferred defects in morphogenesis, decreased adherence to human buccal epithelial cells and attenuated mortality during murine disseminated candidiasis, proving that our strategy can identify genes encoding novel virulence determinants. [source] Agrobacterium type IV secretion is a two-step process in which export substrates associate with the virulence protein VirJ in the periplasmMOLECULAR MICROBIOLOGY, Issue 5 2002Mario Pantoja Summary Type IV secretion systems are virulence determinants in many bacteria and share extensive homology with many conjugal transfer systems. Although type IV systems and their homologues have been studied widely, the mechanism by which substrates are secreted remains unclear. In Agrobacterium, we show that type IV secretion substrates that lack signal peptides form a soluble complex in the periplasm with the virulence protein VirJ. Additionally, these proteins co-precipitate with constituents of the type IV transporter: the VirB pilus and the VirD4 protein. Our findings suggest that the substrate proteins localized to the periplasm may associate with the pilus in a manner that is mediated by VirJ, and suggest a two-step process for type IV secretion in Agrobacterium. Our analyses of protein,protein interactions in a variety of mutant backgrounds indicate that substrates are probably secreted independently of one another. [source] Comparative analysis of virulence determinants and mass spectral profiles of Finnish and Lithuanian endodontic Enterococcus faecalis isolatesMOLECULAR ORAL MICROBIOLOGY, Issue 2 2007A. Reynaud af Geijersstam Introduction:, Putative virulence factors of Enterococcus faecalis have been proposed by several workers and, by analogy, these have been linked to strains of endodontic origin. However, their distribution within the cell population is unknown. In the present study, isolates were taken from the dental root canals of two defined human populations, Lithuanian and Finnish, and examined for a range of virulence properties. In addition, surface-associated molecules and intracellular proteins were compared using matrix-assisted laser desorption-ionization/mass spectrometry (MALDI-TOF-MS) and ProteinChipTM capture/MS (SELDI-TOF-MS), respectively. Methods:, Twenty-three Lithuanian and 35 Finnish dental root canal isolates were included. The esp, gelE, ace and efaA genes were detected by polymerase chain reaction, and cytolysin and gelatinase phenotypes were determined by hydrolysis of horse blood agar and gelatine agar, respectively. Protein extracts and surface-associated molecules of whole cells were analysed by SELDI-TOF-MS and MALDI-TOF-MS, respectively. Results:, Presence of esp (n = 15), cytolysin (n = 9), ace (n = 55) and efaA (n = 58) was not statistically different in the two samples, whereas gelE and gelatinase production was detected more frequently in the Finnish material (chi-squared, P < 0.01). Analysis of protein profiles by SELDI-TOF-MS showed clustering of cytolysin-producing strains, whereas MALDI-TOF-MS generated profiles that clustered according to the samples' origin and, furthermore, to atypical quinupristin,dalfopristin susceptibility. Conclusion:, A high prevalence of virulence factors was demonstrated in both population types. SELDI-TOF-MS and MALDI-TOF-MS proved useful in distinguishing between different E. faecalis phenotypes and they may be useful technologies for elucidating the eco-distribution of E. faecalis in humans. [source] Role of group A Streptococcus HtrA in the maturation of SpeB proteasePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 24 2007Jason N. Cole Abstract The serine protease high-temperature requirement A (HtrA) (DegP) of the human pathogen Streptococcus pyogenes (group A Streptococcus; GAS) is localized to the ExPortal secretory microdomain and is reportedly essential for the maturation of cysteine protease streptococcal pyrogenic exotoxin B (SpeB). Here, we utilize HSC5 (M5 serotype) and the in-frame isogenic mutant HSC5,htrA to determine whether HtrA contributes to the maturation of other GAS virulence determinants. Mutanolysin cell wall extracts and secreted proteins were arrayed by 2-DE and identified by MALDI-TOF PMF analysis. HSC5,htrA had elevated levels of cell wall-associated M protein, whilst the supernatant had higher concentrations of M protein fragments and a reduced amount of mature SpeB protease, compared to wild-type (WT). Western blot analysis and protease assays revealed a delay in the maturation of SpeB in the HSC5,htrA supernatant. HtrA was unable to directly process SpeB zymogen (proSpeB) to the active form in vitro. We therefore conclude that HtrA plays an indirect role in the maturation of cysteine protease SpeB. [source] The N-terminal region of Pseudomonas type III effector AvrPtoB elicits Pto-dependent immunity and has two distinct virulence determinantsTHE PLANT JOURNAL, Issue 4 2007Fangming Xiao Summary Resistance to bacterial speck disease in tomato is activated by the physical interaction of the host Pto kinase with either of the sequence-dissimilar type III effector proteins AvrPto or AvrPtoB (HopAB2) from Pseudomonas syringae pv. tomato. Pto-mediated immunity requires Prf, a protein with a nucleotide-binding site and leucine-rich repeats. The N-terminal 307 amino acids of AvrPtoB were previously reported to interact with the Pto kinase, and we show here that this region (AvrPtoB1-307) is sufficient for eliciting Pto/Prf-dependent immunity against P. s. pv. tomato. AvrPtoB1-307 was also found to be sufficient for a virulence activity that enhances ethylene production and increases growth of P. s. pv. tomato and severity of speck disease on susceptible tomato lines lacking either Pto or Prf. Moreover, we found that residues 308,387 of AvrPtoB are required for the previously reported ability of AvrPtoB to suppress pathogen-associated molecular patterns-induced basal defenses in Arabidopsis. Thus, the N-terminal region of AvrPtoB has two structurally distinct domains involved in different virulence-promoting mechanisms. Random and targeted mutagenesis identified five tightly clustered residues in AvrPtoB1-307 that are required for interaction with Pto and for elicitation of immunity to P. s. pv. tomato. Mutation of one of the five clustered residues abolished the ethylene-associated virulence activity of AvrPtoB1-307. However, individual mutations of the other four residues, despite abolishing interaction with Pto and avirulence activity, had no effect on AvrPtoB1-307 virulence activity. None of these mutations affected the basal defense-suppressing activity of AvrPtoB1-387. Based on sequence alignments, estimates of helical propensity, and the previously reported structure of AvrPto, we hypothesize that the Pto-interacting domains of AvrPto and AvrPtoB1-307 have structural similarity. Together, these data support a model in which AvrPtoB1-307 promotes ethylene-associated virulence by interaction not with Pto but with another unknown host protein. [source] Infectious bovine keratoconjunctivitis vaccine developmentAUSTRALIAN VETERINARY JOURNAL, Issue 8 2005CS McCONNEL Infectious bovine keratoconjunctivitis is a common and highly contagious ocular disease affecting cattle worldwide. The tremendous economic losses attributable to this disease warrant continued investigation into methods of prevention. Multiple virulence factors have been linked to the primary aetiologic agent, Moraxella bovis. Efforts to develop an efficacious vaccine have primarily focused upon the use of surface pili or cytolysin to stimulate host immunity; however, M bovis possesses other virulence determinants that include proteases, fibrinolysins, phospholipases and other cell surface components such as outer membrane proteins. These potentially conserved antigens provide additional possibilities for vaccine development. Examination of appropriate antigen presentation is necessary to attain an adequate immune response. Further, the potential for antigenic diversity as well as epitope conversion requires continuous epidemiological surveillance of isolates recovered from outbreaks. Current work targeting conserved immunogens provides hope for efficacious vaccines that when used in tandem with proper management may control, if not prevent, infectious bovine keratoconjunctivitis. [source] Expression and crystallization of SeDsbA, SeDsbL and SeSrgA from Salmonella enterica serovar TyphimuriumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010R. Jarrott Pathogens require protein-folding enzymes to produce functional virulence determinants. These foldases include the Dsb family of proteins, which catalyze oxidative folding in bacteria. Bacterial disulfide catalytic processes have been well characterized in Escherichia coli K-12 and these mechanisms have been extrapolated to other organisms. However, recent research indicates that the K-12 complement of Dsb proteins is not common to all bacteria. Importantly, many pathogenic bacteria have an extended arsenal of Dsb catalysts that is linked to their virulence. To help to elucidate the process of oxidative folding in pathogens containing a wide repertoire of Dsb proteins, Salmonella enterica serovar Typhimurium has been focused on. This Gram-negative bacterium contains three DsbA proteins: SeDsbA, SeDsbL and SeSrgA. Here, the expression, purification, crystallization and preliminary diffraction analysis of these three proteins are reported. SeDsbA, SeDsbL and SeSrgA crystals diffracted to resolution limits of 1.55, 1.57 and 2.6,Å and belonged to space groups P21, P21212 and C2, respectively. [source] Crystallization and preliminary crystallographic analysis of the bacterial capsule assembly-regulating tyrosine phosphatases Wzb of Escherichia coli and Cps4B of Streptococcus pneumoniaeACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2009Hexian Huang Bacterial tyrosine kinases and their cognate phosphatases are key players in the regulation of capsule assembly and thus are important virulence determinants of these bacteria. Examples of the kinase/phosphatase pairing are found in Gram-negative bacteria such as Escherichia coli (Wzc and Wzb) and in Gram-positive bacteria such as Streptococcus pneumoniae (CpsCD and CpsB). Although Wzb and Cps4B are both predicted to dephosphorylate the C-terminal tyrosine cluster of their cognate tyrosine kinase, they appear on the basis of protein sequence to belong to quite different enzyme classes. Recombinant purified proteins Cps4B of S. pneumoniae TIGR4 and Wzb of E. coli K-30 have been crystallized. Wzb crystals belonged to space-group family P3x21 and diffracted to 2.7,Å resolution. Crystal form I of Cps4B belonged to space-group family P4x212 and diffracted to 2.8,Å resolution; crystal form II belonged to space group P212121 and diffracted to 1.9,Å resolution. [source] Intracellular biology and virulence determinants of Francisella tularensis revealed by transcriptional profiling inside macrophagesCELLULAR MICROBIOLOGY, Issue 7 2009Tara D. Wehrly Summary The highly infectious bacterium Francisella tularensis is a facultative intracellular pathogen, whose virulence requires proliferation inside host cells, including macrophages. Here we have performed a global transcriptional profiling of the highly virulent F. tularensis ssp. tularensis Schu S4 strain during its intracellular cycle within primary murine macrophages, to characterize its intracellular biology and identify pathogenic determinants based on their intracellular expression profiles. Phagocytosed bacteria rapidly responded to their intracellular environment and subsequently altered their transcriptional profile. Differential gene expression profiles were revealed that correlated with specific intracellular locale of the bacteria. Upregulation of general and oxidative stress response genes was a hallmark of the early phagosomal and late endosomal stages, while induction of transport and metabolic genes characterized the cytosolic replication stage. Expression of the Francisella Pathogenicity Island (FPI) genes, which are required for intracellular proliferation, increased during the intracellular cycle. Similarly, 27 chromosomal loci encoding putative hypothetical, secreted, outer membrane proteins or transcriptional regulators were identified as upregulated. Among these, deletion of FTT0383, FTT0369c or FTT1676 abolished the ability of Schu S4 to survive or proliferate intracellularly and cause lethality in mice, therefore identifying novel determinants of Francisella virulence from their intracellular expression profile. [source] Plant models for animal pathogenesisCELLULAR MICROBIOLOGY, Issue 3 2005B. Prithiviraj Summary Several bacteria that are pathogenic to animals also infect plants. Mechanistic studies have proven that some human/animal pathogenic bacteria employ a similar subset of virulence determinants to elicit disease in animals, invertebrates and plants. Therefore, the results of plant infection studies are relevant to animal pathogenesis. This discovery has resulted in the development of convenient, cost-effective, and reliable plant infection models to study the molecular basis of infection by animal pathogens. Plant infection models provide a number of advantages in the study of animal pathogenesis. Using a plant model, mutations in animal pathogenic bacteria can easily be screened for putative virulence factors, a process which if done using existing animal infection models would be time-consuming and tedious. High-throughput screening of plants also provides the potential for unravelling the mechanisms by which plants resist animal pathogenic bacteria, and provides a means to discover novel therapeutic agents such as antibiotics and anti-infective compounds. In this review, we describe the developing technique of using plants as a model system to study Pseudomonas aeruginosa, Enterococcus faecalis and Staphylococcus aureus pathogenesis, and discuss ways to use this new technology against disease warfare and other types of bioterrorism. [source] | ||||||||||||||||||||||||||||