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serodiagnosis of severe leptospirosis: evaluation of elisa based on the recombinant ompl1 or lipl41 antigens of leptospira interrogans serovar autumnalis.elisa based on the recombinant ompl1 and lipl41 antigens of leptospira interrogans serovar autumnalis strain n2 have been developed, for the serodiagnosis of the severe pulmonary form of leptospirosis on the endemic andaman islands. the recombinant ompl1 and lipl41 were produced using escherichia coli expression systems and then purified before each was evaluated in an igm-elisa. the sera tested came from 224 patients who had the severe form of leptospirosis and pulmonary pneumonitis as major sy ...200819000387
putative outer membrane proteins of leptospira interrogans stimulate human umbilical vein endothelial cells (huvecs) and express during infection.cell adhesion molecules (cams) are surface receptors present in eukaryotic cells that mediate cell-cell or cell-extracellular matrix interactions. vascular endothelium stimulation in vitro that lead to the upregulation of cams was reported for the pathogenic spirochaetes, including rlic10365 of leptospira interrogans. in this study, we report the cloning of lic10507, lic10508, lic10509 genes of l. interrogans using escherichia coli as a host system. the rational for selecting these sequences is ...200818778767
evaluation of leptospiral recombinant antigens mpl17 and mpl21 for serological diagnosis of leptospirosis by enzyme-linked immunosorbent assays.leptospirosis is a zoonosis of multisystem involvement caused by pathogenic strains of the genus leptospira. in the last few years, intensive studies aimed at the development of a vaccine have provided important knowledge about the nature of the immunological mechanisms of the host. the purpose of this study was to analyze the immune responses to two recombinant proteins, mpl17 and mpl21 (encoded by the genes lic10765 and lic13131, respectively) of leptospira interrogans serovar copenhageni in i ...200818799647
synthetic tetra-acylated derivatives of lipid a from porphyromonas gingivalis are antagonists of human tlr4.tetra-acylated lipid as derived from porphyromonas gingivalis lps have been synthesized using a key disaccharide intermediate functionalized with levulinate (lev), allyloxycarbonate (alloc) and anomeric dimethylthexylsilyl (tds) as orthogonal protecting groups and 9-fluorenylmethoxycarbamate (fmoc) and azido as amino protecting groups. furthermore, an efficient cross-metathesis has been employed for the preparation of the unusual branched r-(3)-hydroxy-13-methyltetradecanic acid and (r)-3-hexade ...200818802645
the genome of rhodobacter sphaeroides strain 2.4.1 encodes functional cobinamide salvaging systems of archaeal and bacterial origins.bacteria and archaea use distinct pathways for salvaging exogenous cobinamide (cbi), a precursor of adenosylcobalamin (coenzyme b(12)). the bacterial pathway depends on a bifunctional enzyme with kinase and guanylyltransferase activities (cobp in aerobic adenosylcobalamin synthesizers) to convert adenosylcobinamide (adocbi) to adocbi-guanosine diphosphate (adocbi-gdp) via an adocbi-phosphate intermediate. archaea lack cobp, and use a different strategy for the synthesis of adocbi-gdp. archaea cl ...200818808385
targeted mutagenesis in pathogenic leptospira species: disruption of the ligb gene does not affect virulence in animal models of leptospirosis.the pathogenic mechanisms of leptospira interrogans, the causal agent of leptospirosis, remain largely unknown. this is mainly due to the lack of tools for genetically manipulating pathogenic leptospira species. thus, homologous recombination between introduced dna and the corresponding chromosomal locus has never been demonstrated for this pathogen. leptospiral immunoglobulin-like repeat (lig) proteins were previously identified as putative leptospira virulence factors. in this study, a ligb mu ...200818809657
an insect symbiosis is influenced by bacterium-specific polymorphisms in outer-membrane protein a.beneficial bacterial symbioses are ubiquitous in nature. however, the functional and molecular basis of host tolerance to resident symbiotic microbes, in contrast to resistance to closely related bacteria that are recognized as foreign, remain largely unknown. we used the tsetse fly (glossina morsitans), which depends on symbiotic flora for fecundity and has limited exposure to foreign microbes, to investigate the tolerance phenomenon exhibited during symbiosis. we examined the potential role of ...200818815366
recombinant multiepitope protein for diagnosis of leptospirosis.leptospirosis is an emerging infectious disease and is considered to be the most widespread zoonotic disease in the world. it can be misdiagnosed because manifestations of this febrile disease vary from mild flu-like symptoms to severe illness involving vital organs such as the liver and lungs. therefore, accurate diagnosis for differentiation of leptospirosis from other pyrogenic infections prevailing in the same locality is imperative for proper treatment. here, we report a customized recombin ...200818827193
viral and bacterial serology of six free-ranging bearded seals erignathus barbatus.serum or heparinized plasma samples were obtained from 3 male (2 adult and 1 weaned calf) and 3 adult female free-ranging bearded seals erignathus barbatus in may of 1994, 1995, or 1996. blood samples were obtained from animals taken in subsistence hunts near st. lawrence island, alaska and screened for antibodies to a suite of bacteria and viruses potentially pathogenic for pinnipeds and/or humans. no samples had detectable antibodies to brucella spp., phocine distemper virus, influenza a virus ...200818828565
[optimization of prokaryotic expression conditions of leptospira interrogans trigeminy genus-specific protein antigen based on surface response analysis].lipoproteins lipl32 and lipl21 and transmembrane protein ompl1 have been confirmed as the superficial genus-specific antigens of leptospira interrogans, which can be used as antigens for developing a universal genetic engineering vaccine.200818837366
[application of multilocus variable-number tandem-repeat analysis for gene typing and epidemiology of leptospira interrogans]. 200818844004
diagnosis of leptospirosis by recombinant antigen based single serum dilution elisa.leptospirosis, a zoonosis with a worldwide distribution is an acute febrile illness caused by spirochaetes of the pathogenic leptospira interrogans. microscopic agglutination test (mat), the reference method for diagnosis was successively done to evaluate the modified elisa which was developed with the recombinant lipl32 antigen for the detection of anti-leptospiral antibodies in human serum samples.200819001681
human leptospira interrogans serogroup icterohaemorrhagiae infection (weil's disease) acquired from pet rats. 200819011247
[advance in research of pathogenic mechanism and novel genetic engineering vaccines of leptospira interrogans and bacterial drug resistance]. 200819084949
[upregulation of fasl/fas expression and fasl/fas-associated apoptosis in j774a.1 cells induced by leptospira interrogans].to determine the involvement of fasl/fas pathway in apoptosis of j774a.1 cells induced by leptospira interrogans.200819084951
[j774a.1 cell apoptosis induced by leptospira interrogans and effects of caspase-3, -6 activation on apoptosis].to investigate the apoptosis of j774a.1 cells induced by leptospira interrogans and the effect of caspase-3, -6 activation on the apoptosis.200819084952
[sequences and expression pattern of mce gene in leptospira interrogans of different serogroups].to determine the frequency of mce gene in leptospira interrogans, and to investigate the gene transcription levels of l. interrogans before and after infecting cells.200819084953
[association of flir gene in leptospira interrogans with adhesion and pathogenicity to host cells].to investigate the pathogenicity of leptospira interrogans flir gene to j774a.1 cells.200819084954
[adhering effect of leptospira interrongan to major extracellular matrix molecules].to determine the adhering ability of leptospira interrogans to extracellular matrix molecules (ecm) of host cells and its diversity.200819084955
[prediction and immunologic identification of antigenic epitopes in genus-specific outer membrane protein lipl41 of leptospira interrogans].to predict and screen the efficient antigenic epitopes in genus-specific envelope protein lipl41 of leptospira interrogans and to determine the immunoreactive diversity of lipl41s from different genotypes.200819084956
[establishment and application of elisas based on rlipl32/1-lipl21-ompl1/2 fusion antigen of leptospira interrogans].to establish elisas based on rlipl32/1-lipl21-ompl1/2 fusion antigen of leptospira interrogans for detecting specific igg and igm in serum of patients with leptospirosis.200819084957
[prokaryotic expression of trigeminy artificial fusion gene of leptospira interrogans and the immunogenicity of its products].to construct lipl32/1-lipl21-ompl1/2 fusion gene of leptospira interrogans and its prokaryotic expression system, and to identify the immunogenicity of its products.200819084958
characterization of the ompl1 gene of pathogenic leptospira species in china and cross-immunogenicity of the ompl1 protein.the usefulness of available vaccine and serological tests for leptospirosis is limited by the low cross-reactivity of antigens from numerous serovars of pathogenic leptospira spp. identification of genus-specific protein antigens (gp-ag) of leptospira would be important for development of universal vaccines and serodiagnostic methods. ompl1, a transmembrane porin of pathogenic leptospires, was identified as a possible gp-ag, but its sequence diversity and immune cross-reactivity among different ...200819087358
directed evolution of methanococcus jannaschii citramalate synthase for biosynthesis of 1-propanol and 1-butanol by escherichia coli.biofuels synthesized from renewable resources are of increasing interest because of global energy and environmental problems. we have previously demonstrated production of higher alcohols from escherichia coli using a 2-keto acid-based pathway. here, we took advantage of the growth phenotype associated with 2-keto acid deficiency to construct a hyperproducer of 1-propanol and 1-butanol by evolving citramalate synthase (cima) from methanococcus jannaschii. this new pathway, which directly convert ...200818952866
protection of guinea pigs against leptospira interrogans serovar lai by lipl21 dna vaccine.in this study, the full lipl21 gene fragment encoding outer membrane protein lipl21 was cloned from l. interrogans serovar lai and inserted into eukaryotic expression vector pcdna3.1(+). the guinea pigs were immunized with pcdna3.1(+)-lipl21, pcdna3.1(+) or pbs. six weeks after the second immunization, the splenocytes were isolated to detect their proliferative ability by lymphocyte transformation experiments. in addition, microscopic agglutination test was used for quantitative detection of spe ...200818954563
predictors of lethality in severe leptospirosis in urban brazil.to ascertain prognostic factors associated with fatal outcomes in severe leptospirosis, a retrospective case-control study was done using population-based surveillance data. centralized death certificate reporting of leptospirosis mortality was combined with details of patients' hospitalizations, which were obtained from hospitals representing all sectors of são paulo city. among identified leptospirosis cases, 89 lethal cases and 281 survivor cases were analyzed. predictors of death included ag ...200819052303
overcoming function annotation errors in the gram-positive pathogen streptococcus suis by a proteomics-driven approach.annotation of protein-coding genes is a key step in sequencing projects. protein functions are mainly assigned on the basis of the amino acid sequence alone by searching of homologous proteins. however, fully automated annotation processes often lead to wrong prediction of protein functions, and therefore time-intensive manual curation is often essential. here we describe a fast and reliable way to correct function annotation in sequencing projects, focusing on surface proteomes. we use a proteo ...200819061494
toll-like receptors and cytokines as surrogate biomarkers for evaluating vaginal immune response following microbicide administration.topical microbicides are intended for frequent use by women in reproductive age. hence, it is essential to evaluate their impact on mucosal immune function in the vagina. in the present study, we evaluated nisin, a naturally occurring antimicrobial peptide (amp), for its efficacy as an intravaginal microbicide. its effect on the vaginal immune function was determined by localizing toll-like receptors (tlrs-3, 9) and cytokines (il-4, 6 , 10 and tnf-alpha) in the rabbit cervicovaginal epithelium f ...200819125187
rickettsia phylogenomics: unwinding the intricacies of obligate intracellular life.completed genome sequences are rapidly increasing for rickettsia, obligate intracellular alpha-proteobacteria responsible for various human diseases, including epidemic typhus and rocky mountain spotted fever. in light of phylogeny, the establishment of orthologous groups (ogs) of open reading frames (orfs) will distinguish the core rickettsial genes and other group specific genes (class 1 ogs or c1ogs) from those distributed indiscriminately throughout the rickettsial tree (class 2 og or c2ogs) ...200819194535
differential response of cytokines induced by leptospira interrogans, serogroup pomona, serovar pomona, in mouse and human cell lines.leptospira interrogans, the causative agent of leptospirosis, is an important zoonotic bacterium. the mechanisms and roles of cytokine induction in both humans and animals remain unclear. therefore, the ifn-gamma, il-6, il-10 and il-12 levels were measured by enzyme-linked immunosorbent assay (elisa) in human thp-1 and mouse raw 264.7 monocyte cell lines following stimulation with heat-killed leptospira interrogans serogroup pomona serovar pomona, l. biflexa, e. coli or salmonella group b. the p ...200819317342
expression and characterization of recombinant leptospiral outer membrane protein lipl32 from leptospira interrogans serovar autumnalis.leptospira interrogans serovar autumnalis, a causative agent of leptospirosis in thailand, was isolated from a patient for dna extraction and amplification of lipl32 gene by polymerase chain reaction (pcr). the 782 bp pcr product was obtained, which was inserted into pae plasmid with polyhistidine (his6 tag) to construct pae-lipl32. this recombinant plasmid was transfected into e. coli bl21 (de3). his6-lipl32 was purified by ni-nta affinity chromatography. the recombinant protein was used as ant ...200919323048
leptospira seroprevalence and associations between seropositivity, clinical disease and host factors in horses.a cross-sectional study was carried out to determine the seroprevalence of different serovars of leptospira spp. and their association with clinical disease and host factors in swedish horses.200919331656
leptospira noguchii and human and animal leptospirosis, southern brazil. 200919331754
molecular basis of the inhibitor selectivity and insights into the feedback inhibition mechanism of citramalate synthase from leptospira interrogans.licms (leptospira interrogans citramalate synthase) catalyses the first reaction of the isoleucine biosynthesis pathway in l. interrogans, the pathogen of leptospirosis. the catalytic reaction is regulated through feedback inhibition by its end product isoleucine. to understand the molecular basis of the high selectivity of the inhibitor and the mechanism of feedback inhibition, we determined the crystal structure of licmsc (c-terminal regulatory domain of licms) in complex with isoleucine, and ...200919351325
use of quantitative real-time pcr for studying the dissemination of leptospira interrogans in the guinea pig infection model of leptospirosis.the dynamics of leptospirosis infection have been poorly studied. the purpose of this study was to determine the ld(50), rate of bacterial dissemination, histopathology and antibody responses against leptospira following inoculation with the highly virulent leptospira interrogans fiocruz l1-130 strain in a guinea pig model of leptospirosis. three routes of infection (intraperitoneal, conjunctival and subcutaneous inoculation) were used to establish disease in guinea pigs. the size and kinetics o ...200919369528
characterization of two homologous disulfide bond systems involved in virulence factor biogenesis in uropathogenic escherichia coli cft073.disulfide bond (dsb) formation is catalyzed by disulfide bond proteins and is critical for the proper folding and functioning of secreted and membrane-associated bacterial proteins. uropathogenic escherichia coli (upec) strains possess two paralogous disulfide bond systems: the well-characterized dsbab system and the recently described dsbli system. in the dsbab system, the highly oxidizing dsba protein introduces disulfide bonds into unfolded polypeptides by donating its redox-active disulfide ...200919376849
multiple-locus variable-number tandem-repeat analysis for clonal identification of vibrio parahaemolyticus isolates by using capillary electrophoresis.epidemics of vibrio parahaemolyticus in chile have occurred since 1998. direct genome restriction enzyme analysis (dgrea) using conventional gel electrophoresis permitted discrimination of different v. parahaemolyticus isolates obtained from these outbreaks and showed that this species consists of a highly diverse population. a multiple-locus variable-number tandem-repeat (vntr) analysis (mlva) approach was developed and applied to 22 clinical and 91 environmental v. parahaemolyticus isolates fr ...200919376898
detection of pathogenic leptospira spp. through taqman polymerase chain reaction targeting the lipl32 gene.rapid diagnosis of leptospirosis, through culture and/or serology, can be difficult without proper expertise and is often delayed because of the length of time required to obtain results. in this study, we developed a real-time polymerase chain reaction (pcr) assay using a taqman probe targeting lipl32, which is present only in pathogenic leptospira spp. using leptospira interrogans serovar icterohaemorrhagiae dna, the lower limit of detection was found to be 20 genomic equivalents/reaction with ...200919395218
6-pyruvoyltetrahydropterin synthase paralogs replace the folate synthesis enzyme dihydroneopterin aldolase in diverse bacteria.dihydroneopterin aldolase (folb) catalyzes conversion of dihydroneopterin to 6-hydroxymethyldihydropterin (hmdhp) in the classical folate biosynthesis pathway. however, folb genes are missing from the genomes of certain bacteria from the phyla chloroflexi, acidobacteria, firmicutes, planctomycetes, and spirochaetes. almost all of these folb-deficient genomes contain an unusual paralog of the tetrahydrobiopterin synthesis enzyme 6-pyruvoyltetrahydropterin synthase (ptps) in which a glutamate resi ...200919395485
serologic survey for selected infectious diseases in raccoons (procyon lotor) in indiana, usa.the objective of this study was to characterize the antibody prevalence of important livestock and domestic animal pathogens in raccoons (procyon lotor) trapped and sampled in 39 forest patches in north-central indiana, usa, between 2004 and 2005. a total of 459 serum samples were tested for antibodies to leptospira serovars, 512 for canine distemper virus (cdv) antibodies, and 340 for antibodies to porcine reproductive and respiratory syndrome virus (prrsv). in total, 47, 16, and 0% of the samp ...200919395767
borrelia burgdorferi reva antigen binds host fibronectin.borrelia burgdorferi, the lyme disease-causing spirochete, can persistently infect its vertebrate hosts for years. b. burgdorferi is often found associated with host connective tissue, where it interacts with components of the extracellular matrix, including fibronectin. some years ago, a borrelial surface protein, named bbk32, was identified as a fibronectin-binding protein. however, b. burgdorferi bbk32 mutants are still able to bind fibronectin, indicating that the spirochete possesses additi ...200919398540
mutational analysis of escherichia coli sigma28 and its target promoters reveals recognition of a composite -10 region, comprised of an 'extended -10' motif and a core -10 element.sigma28 controls the expression of flagella-related genes and is the most widely distributed alternative sigma factor, present in motile gram-positive and gram-negative bacteria. the distinguishing feature of sigma28 promoters is a long -10 region (gccgataa). despite the fact that the upstream gc is highly conserved, previous studies have not indicated a functional role for this motif. here we examine the functional relevance of the gccg motif and determine which residues in sigma28 participate ...200919400790
reclassification of leptospira meyeri serovar perameles to leptospira interrogans serovar perameles through serological and molecular analysis: evidence of a need for changes to current procedures in leptospira taxonomy.it has been recognized that there is heterogeneity among leptospira isolates in culture collections worldwide, causing confounding results for researchers utilizing these organisms; one such culture is leptospira meyeri serovar perameles. the serovar reference strain bandicoot 343 was previously identified to the species level by dna-dna hybridization; however, subsequent published studies demonstrated results that contradicted the initial speciation. in this study, initial serological testing w ...200919406819
pulmonary disease in hamsters infected with leptospira interrogans: histopathologic findings and cytokine mrna expressions.our aim was to evaluate the pulmonary changes induced by leptospira interrogans infection in hamsters, and the gene expression of endogenous mediators in lung fragments during 28 days of observation. the animals were euthanized on days 4, 7, 14, 21, and 28 post-inoculation. histopathologic lung analysis showed hemorrhage, pneumonia, alveolar congestion, and infiltrated cellular areas, with increasing severity until day 21 post-inoculation. tumor necrosis factor (tnf)-alpha mrna expression enhanc ...200919407133
wild canids as sentinels of ecological health: a conservation medicine perspective.abstract : the extinction of species across the globe is accelerating, directly or indirectly due to human activities. biological impoverishment, habitat fragmentation, climate change, increasing toxification, and the rapid global movement of people and other living organisms have worked synergistically to diminish ecosystem function. this has resulted in unprecedented levels of disease emergence, driven by human-induced environmental degradation, which poses a threat to the survival and health ...200919426446
intact flagellar motor of borrelia burgdorferi revealed by cryo-electron tomography: evidence for stator ring curvature and rotor/c-ring assembly flexion.the bacterial flagellar motor is a remarkable nanomachine that provides motility through flagellar rotation. prior structural studies have revealed the stunning complexity of the purified rotor and c-ring assemblies from flagellar motors. in this study, we used high-throughput cryo-electron tomography and image analysis of intact borrelia burgdorferi to produce a three-dimensional (3-d) model of the in situ flagellar motor without imposing rotational symmetry. structural details of b. burgdorfer ...200919429612
acute disseminated encephalomyelitis after leptospirosis.we report a case of acute disseminated encephalomyelitis after infection by the spirochete leptospira interrogans and review the few cases of acute disseminated encephalomyelitis and central nervous system disorders described in literature. the high prevalence of leptospirosis in developing countries and the possibility of acute disseminated encephalomyelitis, per se, highlights the importance of complete investigation for diagnosis and early treatment, leading to a better prognosis with reducti ...200919433285
limited transcriptional responses of rickettsia rickettsii exposed to environmental stimuli.rickettsiae are strict obligate intracellular pathogens that alternate between arthropod and mammalian hosts in a zoonotic cycle. typically, pathogenic bacteria that cycle between environmental sources and mammalian hosts adapt to the respective environments by coordinately regulating gene expression such that genes essential for survival and virulence are expressed only upon infection of mammals. temperature is a common environmental signal for upregulation of virulence gene expression although ...200919440298
structural basis for the sugar nucleotide and acyl-chain selectivity of leptospira interrogans lpxa.the first step of lipid a biosynthesis is catalyzed by lpxa in escherichia coli (eclpxa), an acyltransferase selective for udp-glcnac and r-3-hydroxymyristoyl-acyl carrier protein (acp). leptospira interrogans lpxa (lilpxa) is extremely selective for r-3-hydroxylauroyl-acp and an analogue of udp-glcnac, designated udp-glcnac3n, in which nh(2) replaces the glcnac 3-oh group. eclpxa does not discriminate between udp-glcnac and udp-glcnac3n; however, e. coli does not make udp-glcnac3n. with lilpxa, ...200919456129
chemokines expression during leptospira interrogans serovar copenhageni infection in resistant balb/c and susceptible c3h/hej mice.the role of innate immune responses in protection against leptospirosis remains unclear. we examined the expression of the chemokines ccl2/je (mcp-1), ccl3/mip-1 alpha (mip-1 alpha) and cxcl1/kc (il-8) regarding resistance and susceptibility to leptospirosis in experimental mice models balb/c and c3h/hej, respectively. a virulent strain of leptospira interrogans serovar copenhageni was used in this study. twenty-five animals of each mouse strain of c3h/hej and balb/c, were infected intraperitone ...200919460427
repeated domains of leptospira immunoglobulin-like proteins interact with elastin and tropoelastin.leptospira spp., the causative agents of leptospirosis, adhere to components of the extracellular matrix, a pivotal role for colonization of host tissues during infection. previously, we and others have shown that leptospira immunoglobulin-like proteins (lig) of leptospira spp. bind to fibronectin, laminin, collagen, and fibrinogen. in this study, we report that leptospira can be immobilized by human tropoelastin (hte) or elastin from different tissues, including lung, skin, and blood vessels, a ...200919473986
severe human infection with rickettsia felis associated with hepatitis in yucatan, mexico.rickettsia felis infection usually is a mild-to-moderate illness characterized by general signs and symptoms. generally, patients do not require hospitalization. however, we detected 2 severe infections with r. felis. our findings support the importance of r. felis infection as a potentially severe illness in humans.200919481502
the elastic basis for the shape of borrelia burgdorferi.the mechanisms that determine bacterial shape are in many ways poorly understood. a prime example is the lyme disease spirochete, borrelia burgdorferi (b. burgdorferi), which mechanically couples its motility organelles, helical flagella, to its rod-shaped cell body, producing a striking flat-wave morphology. a mathematical model is developed here that accounts for the elastic coupling of the flagella to the cell cylinder and shows that the flat-wave morphology is in fact a natural consequence o ...200919486665
soluble st2 levels are associated with bleeding in patients with severe leptospirosis.severe leptospirosis features bleeding and multi-organ failure, leading to shock and death. currently it is assumed that both exaggerated inflammation and immune suppression contribute to mortality in sepsis. indeed, several proinflammatory cytokines are reported to be induced during leptospirosis. toll-like receptors, which play an important role in the initiation of an innate immune response, are inhibited by negative regulators including the membrane-bound st2 (mst2) receptor. soluble st2 (ss ...200919488407
proteome analysis of leptospira interrogans virulent strain.leptospirosis is a worldwide zoonotic infection of human and veterinary concern. caused by pathogenic spirochetes of the genus leptospira, the disease presents greater incidence in tropical and subtropical regions. the identification of proteins that could be involved in the bacteria host interactions may facilitate the search for immune protective antigens. we report the proteomic analysis of leptospira interrogans serovar pomona virulent strain lpf cultured from kidney and liver of infected ha ...200919590580
proteome-wide cellular protein concentrations of the human pathogen leptospira interrogans.mass-spectrometry-based methods for relative proteome quantification have broadly affected life science research. however, important research directions, particularly those involving mathematical modelling and simulation of biological processes, also critically depend on absolutely quantitative data--that is, knowledge of the concentration of the expressed proteins as a function of cellular state. until now, absolute protein concentration measurements of a considerable fraction of the proteome ( ...200919606093
leptospirosis research: fast, easy and reliable enumeration of mobile leptospires.leptospirosis caused by leptospira interrogans is the most widespread zoonosis and a major public health problem worldwide. based on light-scattering and absorption, quantification of leptospires using uv-vis spectroscopy was used as an indirect counting technique by measuring the optical density and comparing this to automated direct counting using a counting chamber in combination with imaging and analyzing software. two serovars, bangkok and copenhagenii, from log-phase growth were used for t ...200919621128
bayesian inference for within-herd prevalence of leptospira interrogans serovar hardjo using bulk milk antibody testing.leptospirosis is the most widespread zoonosis throughout the world and human mortality from severe disease forms is high even when optimal treatment is provided. leptospirosis is also one of the most common causes of reproductive losses in cattle worldwide and is associated with significant economic costs to the dairy farming industry. herds are tested for exposure to the causal organism either through serum testing of individual animals or through testing bulk milk samples. using serum results ...200919628639
tlr4- and tlr2-mediated b cell responses control the clearance of the bacterial pathogen, leptospira interrogans.leptospirosis is a widespread zoonosis caused by pathogenic leptospira interrogans that are transmitted by asymptomatic infected rodents. leptospiral lipoproteins and lps have been shown to stimulate murine cells via tlrs 2 and 4. host defense mechanisms remain obscure, although tlr4 has been shown to be involved in clearing leptospira. in this study, we show that double (tlr2 and tlr4) knockout (dko) mice rapidly died from severe hepatic and renal failure following leptospira inoculation. strik ...200919635914
a serological survey of leptospirosis in cattle of rural communities in the province of kwazulu-natal, south africa.a serological survey of leptospirosis in cattle originating from rural communities of the province of kwazulu-natal (kzn) in south africa was carried out between march 2001 and december 2003. the survey was designed as a 2-stage survey, using the local diptank as the primary sampling point. in total, 2021 animals from 379 diptanks in 33 magisterial districts were sampled and tested with the microscopic agglutination test (mat). the apparent prevalence at district level was adjusted for clusterin ...200919653519
crystal structure and acyl chain selectivity of escherichia coli lpxd, the n-acyltransferase of lipid a biosynthesis.lpxd catalyzes the third step of lipid a biosynthesis, the r-3-hydroxyacyl-acp-dependent n-acylation of udp-3-o-(acyl)-alpha-d-glucosamine, and is a target for new antibiotic development. here we report the 2.6 a crystal structure of the escherichia coli lpxd homotrimer (eclpxd). as is the case in chlamydia trachomatis lpxd (ctlxpd), each eclpxd chain consists of an n-terminal uridine-binding region, a left-handed parallel beta-helix (lbetah), and a c-terminal alpha-helical domain. the backbones ...200919655786
global proteome analysis of leptospira interrogans.comparative global proteome analyses were performed on leptospira interrogans serovar copenhageni grown under conventional in vitro conditions and those mimicking in vivo conditions (iron limitation and serum presence). proteomic analyses were conducted using itraq and lc-esi-tandem mass spectrometry complemented with two-dimensional gel electrophoresis and maldi-tof mass spectrometry. a total of 563 proteins were identified in this study. altered expression of 65 proteins, including upregulatio ...200919663501
innovations in host and microbial sialic acid biosynthesis revealed by phylogenomic prediction of nonulosonic acid structure.sialic acids (sias) are nonulosonic acid (nulo) sugars prominently displayed on vertebrate cells and occasionally mimicked by bacterial pathogens using homologous biosynthetic pathways. it has been suggested that sias were an animal innovation and later emerged in pathogens by convergent evolution or horizontal gene transfer. to better illuminate the evolutionary processes underlying the phenomenon of sia molecular mimicry, we performed phylogenomic analyses of biosynthetic pathways for sias and ...200919666579
an imprint method for detecting leptospires in the hamster model of vaccine-mediated immunity for leptospirosis.in determining the efficacy of new vaccine candidates for leptospirosis, the primary end point is death and an important secondary end point is sterilizing immunity. however, evaluation of this end point is often hampered by the time-consuming demands and complexity of methods such as culture isolation (ci). in this study, we evaluated the use of an imprint (or touch preparation) method (im) in detecting the presence of leptospires in tissues of hamsters infected with leptospira interrogans sero ...200919679685
wild boars as sources for infectious diseases in livestock and humans.wild boars (sus scrofa) are indigenous in many countries in the world. these free-living swine are known reservoirs for a number of viruses, bacteria and parasites that are transmissible to domestic animals and humans. changes of human habitation to suburban areas, increased use of lands for agricultural purposes, increased hunting activities and consumption of wild boar meat have increased the chances of exposure of wild boars to domestic animals and humans. wild boars can act as reservoirs for ...200919687039
immunoproteomic analysis of outer membrane proteins and extracellular proteins of actinobacillus pleuropneumoniae jl03 serotype 3.actinobacillus pleuropneumoniae is the causative agent of porcine contagious pleuropneumonia, a highly contagious respiratory infection in pigs, and all the 15 serotypes are able to cause disease. current vaccines including subunit vaccines could not provide satisfactory protection against a. pleuropneumoniae. in this study, the immunoproteomic approach was applied to the analysis of extracellular and outer membrane proteins of a. pleuropneumoniae jl03 serotype 3 for the identification of novel ...200919695095
a novel fibronectin type iii module binding motif identified on c-terminus of leptospira immunoglobulin-like protein, ligb.infection by pathogenic strains of leptospira hinges on the pathogen's ability to adhere to host cells via extracellular matrix such as fibronectin (fn). previously, the immunoglobulin-like domains of leptospira lig proteins were recognized as adhesins binding to n-terminal domain (ntd) and gelatin binding domain (gbd) of fn. in this study, we identified another fn-binding motif on the c-terminus of the leptospira adhesin ligb (ligbctv), residues 1708-1712 containing sequence lipad with a beta-s ...200919699715
immune response and prophylactic efficacy of smegmosomes in a hamster model of leptospirosis.leptospirosis is an important zoonotic disease worldwide. subunit vaccines are an attractive intervention strategy against this disease, but potent, non-toxic adjuvants are necessary components to any effective vaccine. among various adjuvant candidates, liposomes have garnered recent attention for their capacity as carriers of vaccines. in the present study we prepared novel liposomes using total polar lipids from the nonpathogenic bacterium, mycobacterium smegmatis (designated smegmosomes). th ...200919715780
disruption of the bmei0066 gene attenuates the virulence of brucella melitensis and decreases its stress tolerance.brucella melitensis is a facultative intracellular pathogen. an operon composed of bmei0066, which encodes a two-component response regulator cenr, and bmei0067, which encodes a camp-dependent protein kinase regulatory subunit, has been predicted to exist in many bacterial species. however, little is known about the function of this operon. in order to characterize this operon and assess its role in virulence, we constructed a marked deletion mutant of bmei0066. the mutant was less able to withs ...200919742243
development and validation of a real-time pcr for detection of pathogenic leptospira species in clinical materials.available serological diagnostics do not allow the confirmation of clinically suspected leptospirosis at the early acute phase of illness. several conventional and real-time pcrs for the early diagnosis of leptospirosis have been described but these have been incompletely evaluated. we developed a sybr green-based real-time pcr targeting secy and validated it according to international guidelines. to determine the analytical specificity, dna from 56 leptospira strains belonging to pathogenic, no ...200919763264
variable nucleotide tandem-repeat analysis revealing a unique group of leptospira interrogans serovar pomona isolates associated with california sea lions.leptospira interrogans serovar pomona isolates were compared by variable nucleotide tandem-repeat typing. most cattle isolates grouped together, while isolates from pigs and wildlife were distributed across several groups. significantly, california sea lion isolates formed a unique group, providing evidence that these animals are maintenance hosts of serovar pomona.200919204099
low-density macroarray for rapid detection and identification of crimean-congo hemorrhagic fever virus.crimean-congo hemorrhagic fever (cchf) is a tick-borne viral zoonosis which occurs throughout africa, eastern europe, and asia and results in an approximately 30% fatality rate. a reverse transcription-pcr assay including a competitive internal control was developed on the basis of the most up-to-date genome information. biotinylated amplification products were hybridized to dna macroarrays on the surfaces of polymer supports, and hybridization events were visualized by incubation with a strepta ...200919225100
structure and activity of human mitochondrial peptide deformylase, a novel cancer target.peptide deformylase proteins (pdfs) participate in the n-terminal methionine excision pathway of newly synthesized peptides. we show that the human pdf (hspdf) can deformylate its putative substrates derived from mitochondrial dna-encoded proteins. the first structural model of a mammalian pdf (1.7 a), hspdf, shows a dimer with conserved topology of the catalytic residues and fold as non-mammalian pdfs. the hspdf c-terminus topology and the presence of a helical loop (h2 and h3), however, shape ...200919236878
are white-spot lesions in kidneys in sheep associated with leptospirosis?to determine the association between white-spot lesions in kidneys and serological and cultural prevalence of leptospirosis in sheep, and to evaluate the diagnostic value of these lesions in individual sheep and lines of sheep at slaughter as indicators of past or current episodes of leptospirosis.200919252540
crystallization and preliminary x-ray analysis of lipl32 from leptospira interrogans serovar copenhageni.lipl32 is a major surface protein that is expressed during infection by pathogenic leptospira. here, the crystallization of recombinant lipl32(21-272), which corresponds to the mature lipl32 protein minus its n-terminal lipid-anchored cysteine residue, is described. selenomethionine-labelled lipl32(21-272) crystals diffracted to 2.25 a resolution at a synchrotron source. the space group was p3(1)21 or p3(2)21 and the unit-cell parameters were a = b = 126.7, c = 96.0 a.200919255491
genome sequence of the pathogenic intestinal spirochete brachyspira hyodysenteriae reveals adaptations to its lifestyle in the porcine large intestine.brachyspira hyodysenteriae is an anaerobic intestinal spirochete that colonizes the large intestine of pigs and causes swine dysentery, a disease of significant economic importance. the genome sequence of b. hyodysenteriae strain wa1 was determined, making it the first representative of the genus brachyspira to be sequenced, and the seventeenth spirochete genome to be reported. the genome consisted of a circular 3,000,694 base pair (bp) chromosome, and a 35,940 bp circular plasmid that has not p ...200919262690
rapid and accurate diagnosis of human intestinal spirochetosis by fluorescence in situ hybridization.human intestinal spirochetosis (his) is associated with overgrowth of the large intestine by spirochetes of the genus brachyspira. the microbiological diagnosis of his is hampered by the fastidious nature and slow growth of brachyspira spp. in clinical practice, his is diagnosed histopathologically, and a significant portion of cases may be missed. fluorescence in situ hybridization (fish) is a molecular method that allows the visualization and identification of single bacteria within tissue sec ...200919279178
infection and stillbirth.infection may cause stillbirth by several mechanisms, including direct infection, placental damage, and severe maternal illness. various organisms have been associated with stillbirth, including many bacteria, viruses, and protozoa. in developed countries, between 10% and 25% of stillbirths may be caused by an infection, whereas in developing countries, which have much higher stillbirth rates, the contribution of infection is much greater. in developed countries, ascending bacterial infection, b ...200919285457
serum activity of platelet-activating factor acetylhydrolase is a potential clinical marker for leptospirosis pulmonary hemorrhage.pulmonary hemorrhage has been recognized as a major, often lethal, manifestation of severe leptospirosis albeit the pathogenesis remains unclear. the leptospira interrogans virulent serogroup icterohaemorrhagiae serovar lai encodes a protein (la2144), which exhibited the platelet-activating factor acetylhydrolase (paf-ah) activity in vitro similar to that of human serum with respect to its substrate affinity and specificity and thus designated l-paf-ah. on the other hand, the primary amino acid ...200919145243
onset and duration of protective immunity against clinical disease and renal carriage in dogs provided by a bi-valent inactivated leptospirosis vaccine.protection against clinical disease and prevention of the renal carrier state remain the key objectives of vaccination against leptospirosis in the dog. in the present paper, groups of dogs were vaccinated twice with a commercial bacterin (eurican l) containing leptospira interrogans serovars icterohaemorrhagiae and canicola and challenged with heterologous representatives of both serovars at 2 weeks (onset of immunity) or 14 months (duration of immunity) after the second vaccination. control do ...200919179023
expression of major histocompatibility complex class ii antigens in porcine leptospiral nephritis.class ii major histocompatibility complex (mhcii) is required for the presentation of antigens to cd4 helper t cells. during nephritis, not only primary antigen presenting cells such as histiocytes and lymphocytes, but also cytokine-stimulated tubular epithelial cells express mhcii. leptospirosis in fattening pigs is characterized by several degrees of nephritis, from absence of lesions to severe multifocal tubulo-interstitial inflammation. renal tissue from 20 8-month-old pigs with spontaneous ...200919179617
geographical dissemination of leptospira interrogans serovar pomona during seasonal migration of california sea lions.leptospirosis is one of the most widespread bacterial zoonoses in the world and affects most mammalian species. although leptospirosis is well documented and characterized in terrestrial species, less information is available regarding the distribution and impact of leptospirosis in marine mammals. additionally, the role of animal migrations on the geographical spread of leptospirosis has not been reported. periodic epizootic outbreaks of acute leptospirosis among california sea lions (zalophus ...200919186009
genome sequence of desulfobacterium autotrophicum hrm2, a marine sulfate reducer oxidizing organic carbon completely to carbon dioxide.sulfate-reducing bacteria (srb) belonging to the metabolically versatile desulfobacteriaceae are abundant in marine sediments and contribute to the global carbon cycle by complete oxidation of organic compounds. desulfobacterium autotrophicum hrm2 is the first member of this ecophysiologically important group with a now available genome sequence. with 5.6 megabasepairs (mbp) the genome of db. autotrophicum hrm2 is about 2 mbp larger than the sequenced genomes of other sulfate reducers (srb). a h ...200919187283
novel activities of glycolytic enzymes in bacillus subtilis: interactions with essential proteins involved in mrna processing.glycolysis is one of the most important metabolic pathways in heterotrophic organisms. several genes encoding glycolytic enzymes are essential in many bacteria even under conditions when neither glycolytic nor gluconeogenic activities are required. in this study, a screening for in vivo interaction partners of glycolytic enzymes of the soil bacterium bacillus subtilis was used to provide a rationale for essentiality of glycolytic enzymes. glycolytic enzymes proved to be in close contact with sev ...200919193632
immunogenicity and protective efficacy of recombinant leptospira immunoglobulin-like protein b (rligb) in a hamster challenge model.leptospiral immunoglobulin-like protein (ligb) was truncated into conserved (ligbcon) and variable (varb1, varb2) fragments and expressed as gst/his-tag fusion proteins. four-week-old hamsters were immunized with equal amounts of each fragment individually or combined in alum adjuvant at days 0 and 21 and subsequently challenged three weeks after the booster with 2.5 ld(50) live virulent leptospira interrogans serovar pomona. our results demonstrate that immunization with ligb produced strong hu ...200919070678
leptospirosis in wild and domestic carnivores in natural areas in andalusia, spain.leptospirosis is a zoonosis that affects humans, domestic animals, and wildlife. carnivores are at the top of the feeding chain, thus being exposed to pathogens through their preys. from june 2004 to april 2007, we analyzed for evidences of contact with 14 serovars of leptospira interrogans sensu lato serum (analyzed by indirect microscopic agglutination test) and urine or kidney samples (analyzed by microscopic observation, immunostaining and culture) collected from 201 wild and domestic carniv ...200918973450
association of high orientia tsutsugamushi dna loads with disease of greater severity in adults with scrub typhus.orientia tsutsugamushi, the cause of scrub typhus, is a major pathogen in the asia-pacific region. the severity of infection ranges from mild features to multiorgan failure and death. the aim of this prospective study was to define the o. tsutsugamushi loads in the blood samples of patients with scrub typhus on the day of hospital admission and to determine whether this was associated with disease severity. quantitation was performed using a real-time pcr assay targeting the 16s rrna gene of o. ...200919091812
association of high orientia tsutsugamushi dna loads with disease of greater severity in adults with scrub typhus.orientia tsutsugamushi, the cause of scrub typhus, is a major pathogen in the asia-pacific region. the severity of infection ranges from mild features to multiorgan failure and death. the aim of this prospective study was to define the o. tsutsugamushi loads in the blood samples of patients with scrub typhus on the day of hospital admission and to determine whether this was associated with disease severity. quantitation was performed using a real-time pcr assay targeting the 16s rrna gene of o. ...200919091812
a simple and rapid nested polymerase chain reaction-restriction fragment length polymorphism technique for differentiation of pathogenic and nonpathogenic leptospira spp.a rapid and specific nested polymerase chain reaction-restriction fragment length polymorphism (pcr-rflp) has been developed to detect and differentiate pathogenic and nonpathogenic leptospira spp. leptospiral genomic dna was extracted from suspected human sera using an improved method of standard phenol-chloroform, and specific primers have been used to amplify 16s ribosomal rna from all pathogenic and nonpathogenic leptospira spp. the pcr products of all nonpathogenic species were digested wit ...200919097839
major surface protein lipl32 is not required for either acute or chronic infection with leptospira interrogans.leptospira interrogans is responsible for leptospirosis, a zoonosis of worldwide distribution. lipl32 is the major outer membrane protein of pathogenic leptospires, accounting for up to 75% of total outer membrane protein. in recent times lipl32 has become the focus of intense study because of its surface location, dominance in the host immune response, and conservation among pathogenic species. in this study, an lipl32 mutant was constructed in l. interrogans using transposon mutagenesis. the l ...200919103763
leptospira interrogans requires heme oxygenase for disease pathogenesis.we recently characterised the leptospira interrogans heme oxygenase (hemo) gene and showed that hemo was required for growth with hemoglobin as the sole iron source. here we investigated the role of hemo in pathogenesis. hamsters inoculated with the hemo mutant showed 83% survival, compared with 33% for a control mutant (intergenic transposon insertion). lung pathology was consistent with survival data, showing that hemo contributes significantly to pathogenesis and heme is a major in vivo iron ...200919114124
haem acquisition is facilitated by a novel receptor hma and required by uropathogenic escherichia coli for kidney infection.iron acquisition, mediated by specific outer membrane receptors, is critical for colonization of the urinary tract by uropathogenic escherichia coli (upec). the role of specific iron sources in vivo, however, remains largely unknown. in this study, we identified a 79 kda haem receptor, haemacquisition protein hma, and established that it functions independently of chua to mediate haemin uptake by upec strain cft073. we demonstrated that expression of hma promotes tonb-dependent haemin utilizatio ...200919019144
haem acquisition is facilitated by a novel receptor hma and required by uropathogenic escherichia coli for kidney infection.iron acquisition, mediated by specific outer membrane receptors, is critical for colonization of the urinary tract by uropathogenic escherichia coli (upec). the role of specific iron sources in vivo, however, remains largely unknown. in this study, we identified a 79 kda haem receptor, haemacquisition protein hma, and established that it functions independently of chua to mediate haemin uptake by upec strain cft073. we demonstrated that expression of hma promotes tonb-dependent haemin utilizatio ...200919019144
leptospira immunoglobulin-like protein a variable region (ligavar) incorporated in liposomes and plga microspheres produces a robust immune response correlating to protective immunity.subunit vaccines are attractive as an intervention strategy against leptospirosis, an important zoonotic disease afflicting both humans and livestock. however, the success of subunit vaccines has been hampered by weak or short-term immunity and unavailability of nontoxic, potent adjuvants. in the present study, the variable region of recombinant leptospira immunoglobulin like protein a (ligavar) incorporated into conventional liposomes and plga microspheres produced robust immune responses that ...200919022317
transcription and genetic analyses of a putative n-acetylmuramyl-l-alanine amidase in borrelia burgdorferi.in this study, a putative n-acetylmuramyl-l-alanine amidase gene (bb0666) was identified in the genome of the lyme disease spirochete borrelia burgdorferi. this protein shares c. 30% identity with its counterparts from other bacteria. reverse transcriptase-pcr analysis showed that bb0666 along with two other genes (bb0665 and bb0667) are cotranscribed with the motility and chemotaxis genes. this newly identified operon is termed as pami. sequence and primer extension analyses showed that pami wa ...200919025570
transcription and genetic analyses of a putative n-acetylmuramyl-l-alanine amidase in borrelia burgdorferi.in this study, a putative n-acetylmuramyl-l-alanine amidase gene (bb0666) was identified in the genome of the lyme disease spirochete borrelia burgdorferi. this protein shares c. 30% identity with its counterparts from other bacteria. reverse transcriptase-pcr analysis showed that bb0666 along with two other genes (bb0665 and bb0667) are cotranscribed with the motility and chemotaxis genes. this newly identified operon is termed as pami. sequence and primer extension analyses showed that pami wa ...200919025570
genetic diversity of the leptospiral immunoglobulin-like (lig) genes in pathogenic leptospira spp.recent serologic, immunoprotection, and pathogenesis studies identified the lig proteins as key virulence determinants in interactions of leptospiral pathogens with the mammalian host. we examined the sequence variation and recombination patterns of liga, ligb, and ligc among 10 pathogenic strains from five leptospira species. all strains were found to have intact ligb genes and genetic drift accounting for most of the ligb genetic diversity observed. the liga gene was found exclusively in l. in ...200919028604
leptospira interrogans induces apoptosis in macrophages via caspase-8- and caspase-3-dependent pathways.apoptosis of host cells plays an important role in modulating the pathogenesis of many infectious diseases. it has been reported that leptospira interrogans, the causal agent of leptospirosis, induces apoptosis in macrophages and hepatocytes. however, the molecular mechanisms responsible for host cell death remained largely unknown. here we demonstrate that l. interrogans induced apoptosis in a macrophage-like cell line, j774a.1, and primary murine macrophages in a time- and dose-dependent manne ...200919029301
genome-wide transposon mutagenesis in pathogenic leptospira species.leptospira interrogans is the most common cause of leptospirosis in humans and animals. genetic analysis of l. interrogans has been severely hindered by a lack of tools for genetic manipulation. recently we developed the mariner-based transposon himar1 to generate the first defined mutants in l. interrogans. in this study, a total of 929 independent transposon mutants were obtained and the location of insertion determined. of these mutants, 721 were located in the protein coding regions of 551 d ...200919047402
hprk regulates succinate-mediated catabolite repression in the gram-negative symbiont sinorhizobium meliloti.the hprk kinase/phosphatase is a common component of the phosphotransferase system (pts) of gram-positive bacteria and regulates catabolite repression through phosphorylation/dephosphorylation of its substrate, the pts protein hpr, at a conserved serine residue. phosphorylation of hpr by hprk also affects additional phosphorylation of hpr by the pts enzyme ei at a conserved histidine residue. sinorhizobium meliloti can live as symbionts inside legume root nodules or as free-living organisms and ...200918931135
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