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Sample records for host-cell infection potential

  1. Ureaplasma parvum infection alters filamin a dynamics in host cells

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    Brown Mary B

    2011-04-01

    Full Text Available Abstract Background Ureaplasmas are among the most common bacteria isolated from the human urogenital tract. Ureaplasmas can produce asymptomatic infections or disease characterized by an exaggerated inflammatory response. Most investigations have focused on elucidating the pathogenic potential of Ureaplasma species, but little attention has been paid to understanding the mechanisms by which these organisms are capable of establishing asymptomatic infection. Methods We employed differential proteome profiling of bladder tissues from rats experimentally infected with U. parvum in order to identify host cell processes perturbed by colonization with the microbe. Tissues were grouped into four categories: sham inoculated controls, animals that spontaneously cleared infection, asymptomatic urinary tract infection (UTI, and complicated UTI. One protein that was perturbed by infection (filamin A was used to further elucidate the mechanism of U. parvum-induced disruption in human benign prostate cells (BPH-1. BPH-1 cells were evaluated by confocal microscopy, immunoblotting and ELISA. Results Bladder tissue from animals actively colonized with U. parvum displayed significant alterations in actin binding proteins (profilin 1, vinculin, α actinin, and filamin A that regulate both actin polymerization and cell cytoskeletal function pertaining to focal adhesion formation and signal transduction (Fisher's exact test, P U. parvum perturbed the regulation of filamin A. Specifically, infected BPH-1 cells exhibited a significant increase in filamin A phosphorylated at serine2152 (P ≤ 0.01, which correlated with impaired proteolysis of the protein and its normal intracellular distribution. Conclusion Filamin A dynamics were perturbed in both models of infection. Phosphorylation of filamin A occurs in response to various cell signaling cascades that regulate cell motility, differentiation, apoptosis and inflammation. Thus, this phenomenon may be a useful

  2. Microsporidia infection impacts the host cell's cycle and reduces host cell apoptosis

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    Higes, Mariano; Sagastume, Soledad; Juarranz, Ángeles; Dias-Almeida, Joyce; Budge, Giles E.; Meana, Aránzazu; Boonham, Neil

    2017-01-01

    Intracellular parasites can alter the cellular machinery of host cells to create a safe haven for their survival. In this regard, microsporidia are obligate intracellular fungal parasites with extremely reduced genomes and hence, they are strongly dependent on their host for energy and resources. To date, there are few studies into host cell manipulation by microsporidia, most of which have focused on morphological aspects. The microsporidia Nosema apis and Nosema ceranae are worldwide parasites of honey bees, infecting their ventricular epithelial cells. In this work, quantitative gene expression and histology were studied to investigate how these two parasites manipulate their host’s cells at the molecular level. Both these microsporidia provoke infection-induced regulation of genes involved in apoptosis and the cell cycle. The up-regulation of buffy (which encodes a pro-survival protein) and BIRC5 (belonging to the Inhibitor Apoptosis protein family) was observed after infection, shedding light on the pathways that these pathogens use to inhibit host cell apoptosis. Curiously, different routes related to cell cycle were modified after infection by each microsporidia. In the case of N. apis, cyclin B1, dacapo and E2F2 were up-regulated, whereas only cyclin E was up-regulated by N. ceranae, in both cases promoting the G1/S phase transition. This is the first report describing molecular pathways related to parasite-host interactions that are probably intended to ensure the parasite’s survival within the cell. PMID:28152065

  3. Host Cell Restriction Factors that Limit Influenza A Infection

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    Fernando Villalón-Letelier

    2017-12-01

    Full Text Available Viral infection of different cell types induces a unique spectrum of host defence genes, including interferon-stimulated genes (ISGs and genes encoding other proteins with antiviral potential. Although hundreds of ISGs have been described, the vast majority have not been functionally characterised. Cellular proteins with putative antiviral activity (hereafter referred to as “restriction factors” can target various steps in the virus life-cycle. In the context of influenza virus infection, restriction factors have been described that target virus entry, genomic replication, translation and virus release. Genome wide analyses, in combination with ectopic overexpression and/or gene silencing studies, have accelerated the identification of restriction factors that are active against influenza and other viruses, as well as providing important insights regarding mechanisms of antiviral activity. Herein, we review current knowledge regarding restriction factors that mediate anti-influenza virus activity and consider the viral countermeasures that are known to limit their impact. Moreover, we consider the strengths and limitations of experimental approaches to study restriction factors, discrepancies between in vitro and in vivo studies, and the potential to exploit restriction factors to limit disease caused by influenza and other respiratory viruses.

  4. Coxiella burnetii Nine Mile II proteins modulate gene expression of monocytic host cells during infection

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    Shaw Edward I

    2010-09-01

    Full Text Available Abstract Background Coxiella burnetii is an intracellular bacterial pathogen that causes acute and chronic disease in humans. Bacterial replication occurs within enlarged parasitophorous vacuoles (PV of eukaryotic cells, the biogenesis and maintenance of which is dependent on C. burnetii protein synthesis. These observations suggest that C. burnetii actively subverts host cell processes, however little is known about the cellular biology mechanisms manipulated by the pathogen during infection. Here, we examined host cell gene expression changes specifically induced by C. burnetii proteins during infection. Results We have identified 36 host cell genes that are specifically regulated when de novo C. burnetii protein synthesis occurs during infection using comparative microarray analysis. Two parallel sets of infected and uninfected THP-1 cells were grown for 48 h followed by the addition of chloramphenicol (CAM to 10 μg/ml in one set. Total RNA was harvested at 72 hpi from all conditions, and microarrays performed using Phalanx Human OneArray™ slides. A total of 784 (mock treated and 901 (CAM treated THP-1 genes were up or down regulated ≥2 fold in the C. burnetii infected vs. uninfected cell sets, respectively. Comparisons between the complementary data sets (using >0 fold, eliminated the common gene expression changes. A stringent comparison (≥2 fold between the separate microarrays revealed 36 host cell genes modulated by C. burnetii protein synthesis. Ontological analysis of these genes identified the innate immune response, cell death and proliferation, vesicle trafficking and development, lipid homeostasis, and cytoskeletal organization as predominant cellular functions modulated by C. burnetii protein synthesis. Conclusions Collectively, these data indicate that C. burnetii proteins actively regulate the expression of specific host cell genes and pathways. This is in addition to host cell genes that respond to the presence of the

  5. Differential proteome analysis of chikungunya virus infection on host cells.

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    Christina Li-Ping Thio

    Full Text Available BACKGROUND: Chikungunya virus (CHIKV is an emerging mosquito-borne alphavirus that has caused multiple unprecedented and re-emerging outbreaks in both tropical and temperate countries. Despite ongoing research efforts, the underlying factors involved in facilitating CHIKV replication during early infection remains ill-characterized. The present study serves to identify host proteins modulated in response to early CHIKV infection using a proteomics approach. METHODOLOGY AND PRINCIPAL FINDINGS: The whole cell proteome profiles of CHIKV-infected and mock control WRL-68 cells were compared and analyzed using two-dimensional gel electrophoresis (2-DGE. Fifty-three spots were found to be differentially modulated and 50 were successfully identified by MALDI-TOF/TOF. Eight were significantly up-regulated and 42 were down-regulated. The mRNA expressions of 15 genes were also found to correlate with the corresponding protein expression. STRING network analysis identified several biological processes to be affected, including mRNA processing, translation, energy production and cellular metabolism, ubiquitin-proteasome pathway (UPP and cell cycle regulation. CONCLUSION/SIGNIFICANCE: This study constitutes a first attempt to investigate alteration of the host cellular proteome during early CHIKV infection. Our proteomics data showed that during early infection, CHIKV affected the expression of proteins that are involved in mRNA processing, host metabolic machinery, UPP, and cyclin-dependent kinase 1 (CDK1 regulation (in favour of virus survival, replication and transmission. While results from this study complement the proteomics results obtained from previous late host response studies, functional characterization of these proteins is warranted to reinforce our understanding of their roles during early CHIKV infection in humans.

  6. SPOC1-mediated antiviral host cell response is antagonized early in human adenovirus type 5 infection

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    Schreiner, Sabrina; Kinkley, Sarah; Bürck, Carolin

    2013-01-01

    , and playing a role in DNA damage response. SPOC1 co-localized with viral replication centers in the host cell nucleus, interacted with Ad DNA, and repressed viral gene expression at the transcriptional level. We discovered that this SPOC1-mediated restriction imposed upon Ad growth is relieved by its...... viruses (HSV-1, HSV-2, HIV-1, and HCV) also depleted SPOC1 in infected cells. Our findings provide a general model for how pathogenic human viruses antagonize intrinsic SPOC1-mediated antiviral responses in their host cells. A better understanding of viral entry and early restrictive functions in host...

  7. Coliphage 186 replication is delayed when the host cell is UV irradiated before infection

    International Nuclear Information System (INIS)

    Hooper, I.; Woods, W.H.; Egan, B.

    1981-01-01

    In contrast to results with injections by lambda and P2, the latent period for infection by coliphage 186 is extended when the host cell is UV irradiated before infection. We find that 186 replication is significantly delayed in such a cell, even though the phage itself has not been irradiated. In contrast, replication of the closely related phage P2 under the same conditions is not affected

  8. Selective alterations of the host cell architecture upon infection with parvovirus minute virus of mice

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    Nueesch, Juerg P.F.; Lachmann, Sylvie; Rommelaere, Jean

    2005-01-01

    During a productive infection, the prototype strain of parvovirus minute virus of mice (MVMp) induces dramatic morphological alterations to the fibroblast host cell A9, resulting in cell lysis and progeny virus release. In order to understand the mechanisms underlying these changes, we characterized the fate of various cytoskeletal filaments and investigated the nuclear/cytoplasmic compartmentalization of infected cells. While most pronounced effects could be seen on micro- and intermediate filaments, manifest in dramatic rearrangements and degradation of filamentous (F-)actin and vimentin structures, only little impact could be seen on microtubules or the nuclear envelope during the entire monitored time of infection. To further analyze the disruption of the cytoskeletal structures, we investigated the viral impact on selective regulatory pathways. Thereby, we found a correlation between microtubule stability and MVM-induced phosphorylation of α/β tubulin. In contrast, disassembly of actin filaments late in infection could be traced back to the disregulation of two F-actin associated proteins gelsolin and Wiscott-Aldrich Syndrome Protein (WASP). Thereby, an increase in the amount of gelsolin, an F-actin severing protein was observed during infection, accounting for the disruption of stress fibers upon infection. Concomitantly, the actin polymerization activity also diminished due to a loss of WASP, the activator protein of the actin polymerization machinery the Arp2/3 complex. No effects could be seen in amount and distribution of other F-actin regulatory factors such as cortactin, cofilin, and profilin. In summary, the selective attack of MVM towards distinct host cell cytoskeletal structures argues for a regulatory feature during infection, rather than a collapse of the host cell as a mere side effect of virus production

  9. Mycobacterium-Host Cell Relationships in Granulomatous Lesions in a Mouse Model of Latent Tuberculous Infection

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    Elena Ufimtseva

    2015-01-01

    Full Text Available Tuberculosis (TB is a dangerous infectious disease characterized by a tight interplay between mycobacteria and host cells in granulomatous lesions (granulomas during the latent, asymptomatic stage of infection. Mycobacterium-host cell relationships were analyzed in granulomas obtained from various organs of BALB/c mice with chronic TB infection caused by in vivo exposure to the Bacillus Calmette-Guérin (BCG vaccine. Acid-fast BCG-mycobacteria were found to be morphologically and functionally heterogeneous (in size, shape, and replication rates in colonies in granuloma macrophages, dendritic cells, and multinucleate Langhans giant cells. Cord formation by BCG-mycobacteria in granuloma cells has been observed. Granuloma macrophages retained their ability to ingest damaged lymphocytes and thrombocytes in the phagosomes; however, their ability to destroy BCG-mycobacteria contained in these cells was compromised. No colocalization of BCG-mycobacteria and the LysoTracker dye was observed in the mouse cells. Various relationships between granuloma cells and BCG-mycobacteria were observed in different mice belonging to the same line. Several mice totally eliminated mycobacterial infection. Granulomas in the other mice had mycobacteria actively replicating in cells of different types and forming cords, which is an indicator of mycobacterial virulence and, probably, a marker of the activation of tuberculous infection in animals.

  10. Mycobacterium-Host Cell Relationships in Granulomatous Lesions in a Mouse Model of Latent Tuberculous Infection

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    2015-01-01

    Tuberculosis (TB) is a dangerous infectious disease characterized by a tight interplay between mycobacteria and host cells in granulomatous lesions (granulomas) during the latent, asymptomatic stage of infection. Mycobacterium-host cell relationships were analyzed in granulomas obtained from various organs of BALB/c mice with chronic TB infection caused by in vivo exposure to the Bacillus Calmette-Guérin (BCG) vaccine. Acid-fast BCG-mycobacteria were found to be morphologically and functionally heterogeneous (in size, shape, and replication rates in colonies) in granuloma macrophages, dendritic cells, and multinucleate Langhans giant cells. Cord formation by BCG-mycobacteria in granuloma cells has been observed. Granuloma macrophages retained their ability to ingest damaged lymphocytes and thrombocytes in the phagosomes; however, their ability to destroy BCG-mycobacteria contained in these cells was compromised. No colocalization of BCG-mycobacteria and the LysoTracker dye was observed in the mouse cells. Various relationships between granuloma cells and BCG-mycobacteria were observed in different mice belonging to the same line. Several mice totally eliminated mycobacterial infection. Granulomas in the other mice had mycobacteria actively replicating in cells of different types and forming cords, which is an indicator of mycobacterial virulence and, probably, a marker of the activation of tuberculous infection in animals. PMID:26064970

  11. Recognizing the SINEs of Infection: Regulation of Retrotransposon Expression and Modulation of Host Cell Processes

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    William Dunker

    2017-12-01

    Full Text Available Short interspersed elements (SINEs are a family of retrotransposons evolutionarily derived from cellular RNA polymerase III transcripts. Over evolutionary time, SINEs have expanded throughout the human genome and today comprise ~11% of total chromosomal DNA. While generally transcriptionally silent in healthy somatic cells, SINE expression increases during a variety of types of stresses, including DNA virus infection. The relevance of SINE expression to viral infection was largely unexplored, however, recent years have seen great progress towards defining the impact of SINE expression on viral replication and host gene expression. Here we review the origin and diversity of SINE elements and their transcriptional control, with an emphasis on how their expression impacts host cell biology during viral infection.

  12. Transcriptional profiling of the host cell response to feline immunodeficiency virus infection.

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    Ertl, Reinhard; Klein, Dieter

    2014-03-19

    Feline immunodeficiency virus (FIV) is a widespread pathogen of the domestic cat and an important animal model for human immunodeficiency virus (HIV) research. In contrast to HIV, only limited information is available on the transcriptional host cell response to FIV infections. This study aims to identify FIV-induced gene expression changes in feline T-cells during the early phase of the infection. Illumina RNA-sequencing (RNA-seq) was used identify differentially expressed genes (DEGs) at 24 h after FIV infection. After removal of low-quality reads, the remaining sequencing data were mapped against the cat genome and the numbers of mapping reads were counted for each gene. Regulated genes were identified through the comparison of FIV and mock-infected data sets. After statistical analysis and the removal of genes with insufficient coverage, we detected a total of 69 significantly DEGs (44 up- and 25 down-regulated genes) upon FIV infection. The results obtained by RNA-seq were validated by reverse transcription qPCR analysis for 10 genes. Out of the most distinct DEGs identified in this study, several genes are already known to interact with HIV in humans, indicating comparable effects of both viruses on the host cell gene expression and furthermore, highlighting the importance of FIV as a model system for HIV. In addition, a set of new genes not previously linked to virus infections could be identified. The provided list of virus-induced genes may represent useful information for future studies focusing on the molecular mechanisms of virus-host interactions in FIV pathogenesis.

  13. Membrane rafts: a potential gateway for bacterial entry into host cells.

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    Hartlova, Anetta; Cerveny, Lukas; Hubalek, Martin; Krocova, Zuzana; Stulik, Jiri

    2010-04-01

    Pathogenic bacteria have developed various mechanisms to evade host immune defense systems. Invasion of pathogenic bacteria requires interaction of the pathogen with host receptors, followed by activation of signal transduction pathways and rearrangement of the cytoskeleton to facilitate bacterial entry. Numerous bacteria exploit specialized plasma membrane microdomains, commonly called membrane rafts, which are rich in cholesterol, sphingolipids and a special set of signaling molecules which allow entry to host cells and establishment of a protected niche within the host. This review focuses on the current understanding of the raft hypothesis and the means by which pathogenic bacteria subvert membrane microdomains to promote infection.

  14. Anaplasma phagocytophilum Manipulates Host Cell Apoptosis by Different Mechanisms to Establish Infection

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    Pilar Alberdi

    2016-07-01

    Full Text Available Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human and animal granulocytic anaplasmosis and tick-borne fever of ruminants. This obligate intracellular bacterium evolved to use common strategies to establish infection in both vertebrate hosts and tick vectors. Herein, we discuss the different strategies used by the pathogen to modulate cell apoptosis and establish infection in host cells. In vertebrate neutrophils and human promyelocytic cells HL-60, both pro-apoptotic and anti-apoptotic factors have been reported. Tissue-specific differences in tick response to infection and differential regulation of apoptosis pathways have been observed in adult female midguts and salivary glands in response to infection with A. phagocytophilum. In tick midguts, pathogen inhibits apoptosis through the Janus kinase/signal transducers and activators of transcription (JAK/STAT pathway, while in salivary glands, the intrinsic apoptosis pathways is inhibited but tick cells respond with the activation of the extrinsic apoptosis pathway. In Ixodes scapularis ISE6 cells, bacterial infection down-regulates mitochondrial porin and manipulates protein processing in the endoplasmic reticulum and cell glucose metabolism to inhibit apoptosis and facilitate infection, whereas in IRE/CTVM20 tick cells, inhibition of apoptosis appears to be regulated by lower caspase levels. These results suggest that A. phagocytophilum uses different mechanisms to inhibit apoptosis for infection of both vertebrate and invertebrate hosts.

  15. Fierce competition between Toxoplasma and Chlamydia for host cell structures in dually infected cells.

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    Romano, Julia D; de Beaumont, Catherine; Carrasco, Jose A; Ehrenman, Karen; Bavoil, Patrik M; Coppens, Isabelle

    2013-02-01

    The prokaryote Chlamydia trachomatis and the protozoan Toxoplasma gondii, two obligate intracellular pathogens of humans, have evolved a similar modus operandi to colonize their host cell and salvage nutrients from organelles. In order to gain fundamental knowledge on the pathogenicity of these microorganisms, we have established a cell culture model whereby single fibroblasts are coinfected by C. trachomatis and T. gondii. We previously reported that the two pathogens compete for the same nutrient pools in coinfected cells and that Toxoplasma holds a significant competitive advantage over Chlamydia. Here we have expanded our coinfection studies by examining the respective abilities of Chlamydia and Toxoplasma to co-opt the host cytoskeleton and recruit organelles. We demonstrate that the two pathogen-containing vacuoles migrate independently to the host perinuclear region and rearrange the host microtubular network around each vacuole. However, Toxoplasma outcompetes Chlamydia to the host microtubule-organizing center to the detriment of the bacterium, which then shifts to a stress-induced persistent state. Solely in cells preinfected with Chlamydia, the centrosomes become associated with the chlamydial inclusion, while the Toxoplasma parasitophorous vacuole displays growth defects. Both pathogens fragment the host Golgi apparatus and recruit Golgi elements to retrieve sphingolipids. This study demonstrates that the productive infection by both Chlamydia and Toxoplasma depends on the capability of each pathogen to successfully adhere to a finely tuned developmental program that aims to remodel the host cell for the pathogen's benefit. In particular, this investigation emphasizes the essentiality of host organelle interception by intravacuolar pathogens to facilitate access to nutrients.

  16. Mycobacterium tuberculosis nuoG is a virulence gene that inhibits apoptosis of infected host cells.

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    Kamalakannan Velmurugan

    2007-07-01

    Full Text Available The survival and persistence of Mycobacterium tuberculosis depends on its capacity to manipulate multiple host defense pathways, including the ability to actively inhibit the death by apoptosis of infected host cells. The genetic basis for this anti-apoptotic activity and its implication for mycobacterial virulence have not been demonstrated or elucidated. Using a novel gain-of-function genetic screen, we demonstrated that inhibition of infection-induced apoptosis of macrophages is controlled by multiple genetic loci in M. tuberculosis. Characterization of one of these loci in detail revealed that the anti-apoptosis activity was attributable to the type I NADH-dehydrogenase of M. tuberculosis, and was mainly due to the subunit of this multicomponent complex encoded by the nuoG gene. Expression of M. tuberculosis nuoG in nonpathogenic mycobacteria endowed them with the ability to inhibit apoptosis of infected human or mouse macrophages, and increased their virulence in a SCID mouse model. Conversely, deletion of nuoG in M. tuberculosis ablated its ability to inhibit macrophage apoptosis and significantly reduced its virulence in mice. These results identify a key component of the genetic basis for an important virulence trait of M. tuberculosis and support a direct causal relationship between virulence of pathogenic mycobacteria and their ability to inhibit macrophage apoptosis.

  17. Synthetic immunostimulatory glycans interference with host cell apoptosis upon of Toxoplasma gondii infection, in vitro

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    S.H. Eassa

    2017-06-01

    Full Text Available Toxoplasmosis is a protozoan infection of humans and animals caused by Toxoplasma gondii, and it’s continuous public health and food safety issue. The tachyzoites (Tg of T. gondii are the most important stage, as they come in direct contact with immune cells such as a macrophage. Tg can modulate and prevent apoptosis of immune cells while promoting survival of the pathogen. Infections caused by Tg can be eradicated if immune cells could stimulate apoptosis and kill pathogens upon exposure. Apoptosis is characterized by the release of mediators, namely Caspases (Cas. New means are required for inducing apoptosis and enhance immunity in the infected host cell to control toxoplasmosis. The present study investigated whether Synthetic Immuno-stimulatory Glycans (SIGs influence Cas and Nitric oxide (NO release and led to Tg damage. Galβ1-3Gal-PAA-fluor (SIG1, Fucα1-4GlcNAcβ-PAA-fluor (SIG2 and GlcNAcβ1-3GalNAcα-PAA-fluor (SIG3 constituted samples studied principally. Murine macrophage had been exposed to the Tg then the SIGs effects on Cas and NO production were determined after 20 hours of pathogen phagocytosis. Here we report that the SIGs had potent in vitro activity against T. gondii; SIG2 was more effective than SIG1 and SIG3, representative by SIG2 treated infected macrophages can induced infected macrophages to release Cas1, 3, and 9. Maximum production of NO by infected macrophages was noticed following the expoxure to all SIGs. Therefore the present study provided the method for the selection of SIGs ligands bearing immunostimulatory factor and apoptotic stimuli properties.

  18. Anaplasma phagocytophilum MSP4 and HSP70 Proteins Are Involved in Interactions with Host Cells during Pathogen Infection

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    Marinela Contreras

    2017-07-01

    Full Text Available Anaplasma phagocytophilum transmembrane and surface proteins play a role during infection and multiplication in host neutrophils and tick vector cells. Recently, A. phagocytophilum Major surface protein 4 (MSP4 and Heat shock protein 70 (HSP70 were shown to be localized on the bacterial membrane, with a possible role during pathogen infection in ticks. In this study, we hypothesized that A. phagocytophilum MSP4 and HSP70 have similar functions in tick-pathogen and host-pathogen interactions. To address this hypothesis, herein we characterized the role of these bacterial proteins in interaction and infection of vertebrate host cells. The results showed that A. phagocytophilum MSP4 and HSP70 are involved in host-pathogen interactions, with a role for HSP70 during pathogen infection. The analysis of the potential protective capacity of MSP4 and MSP4-HSP70 antigens in immunized sheep showed that MSP4-HSP70 was only partially protective against pathogen infection. This limited protection may be associated with several factors, including the recognition of non-protective epitopes by IgG in immunized lambs. Nevertheless, these antigens may be combined with other candidate protective antigens for the development of vaccines for the control of human and animal granulocytic anaplasmosis. Focusing on the characterization of host protective immune mechanisms and protein-protein interactions at the host-pathogen interface may lead to the discovery and design of new effective protective antigens.

  19. Internalization of components of the host cell plasma membrane during infection by Trypanosoma cruzi

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    Carvalho TMU

    1999-01-01

    Full Text Available Epimastigote and trypomastigote forms of Trypanosoma cruzi attach to the macrophage surface and are internalized with the formation of a membrane bounded vacuole, known as the parasitophorous vacuole (PV. In order to determine if components of the host cell membrane are internalized during formation of the PV we labeled the macrophage surface with fluorescent probes for proteins, lipids and sialic acid residues and then allowed the labeled cells to interact with the parasites. The interaction process was interrupted after 1 hr at 37ºC and the distribution of the probes analyzed by confocal laser scanning microscopy. During attachment of the parasites to the macrophage surface an intense labeling of the attachment regions was observed. Subsequently labeling of the membrane lining the parasitophorous vacuole containing epimastigote and trypomastigote forms was seen. Labeling was not uniform, with regions of intense and light or no labeling. The results obtained show that host cell membrane lipids, proteins and sialoglycoconjugates contribute to the formation of the membrane lining the PV containing epimastigote and trypomastigote T. cruzi forms. Lysosomes of the host cell may participate in the process of PV membrane formation.

  20. The Influence of Virus Infection on the Extracellular pH of the Host Cell Detected on Cell Membrane.

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    Liu, Hengjun; Maruyama, Hisataka; Masuda, Taisuke; Honda, Ayae; Arai, Fumihito

    2016-01-01

    Influenza virus infection can result in changes in the cellular ion levels at 2-3 h post-infection. More H(+) is produced by glycolysis, and the viral M2 proton channel also plays a role in the capture and release of H(+) during both viral entry and egress. Then the cells might regulate the intracellular pH by increasing the export of H(+) from the intracellular compartment. Increased H(+) export could lead indirectly to increased extracellular acidity. To detect changes in extracellular pH of both virus-infected and uninfected cells, pH sensors were synthesized using polystyrene beads (ϕ1 μm) containing Rhodamine B and Fluorescein isothiocyanate (FITC). The fluorescence intensity of FITC can respond to both pH and temperature. So Rhodamine B was also introduced in the sensor for temperature compensation. Then the pH can be measured after temperature compensation. The sensor was adhered to cell membrane for extracellular pH measurement. The results showed that the multiplication of influenza virus in host cell decreased extracellular pH of the host cell by 0.5-0.6 in 4 h after the virus bound to the cell membrane, compared to that in uninfected cells. Immunostaining revealed the presence of viral PB1 protein in the nucleus of virus-bound cells that exhibited extracellular pH changes, but no PB1 protein are detected in virus-unbound cells where the extracellular pH remained constant.

  1. The influence of virus infection on the extracellular pH of the host cell detected on cell membrane

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    Hengjun Liu

    2016-08-01

    Full Text Available Influenza virus infection can result in changes in the cellular ion levels at 2–3 hours post-infection. More H+ is produced by glycolysis, and the viral M2 proton channel also plays a role in the capture and release of H+ during both viral entry and egress. Then the cells might regulate the intracellular pH by increasing the export of H+ from the intracellular compartment. Increased H+ export could lead indirectly to increased extracellular acidity. To detect changes in extracellular pH of both virus-infected and uninfected cells, pH sensors were synthesized using polystyrene beads (1μm containing Rhodamine B and Fluorescein isothiocyanate (FITC. The fluorescence intensity of FITC can respond to both pH and temperature. So Rhodamine B was also introduced in the sensor for temperature compensation. Then the pH can be measured after temperature compensation. The sensor was adhered to cell membrane for extracellular pH measurement. The results showed that the multiplication of influenza virus in host cell decreased extracellular pH of the host cell by 0.5–0.6 in 4 hours after the virus bound to the cell membrane, compared to that in uninfected cells. Immunostaining revealed the presence of viral PB1 subunits in the nucleus of virus-bound cells that exhibited extracellular pH changes, but no PB1 subunits are detected in virus-unbound cells where the extracellular pH remained constant.

  2. The host cell sulfonation pathway contributes to retroviral infection at a step coincident with provirus establishment.

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    James W Bruce

    2008-11-01

    Full Text Available The early steps of retrovirus replication leading up to provirus establishment are highly dependent on cellular processes and represent a time when the virus is particularly vulnerable to antivirals and host defense mechanisms. However, the roles played by cellular factors are only partially understood. To identify cellular processes that participate in these critical steps, we employed a high volume screening of insertionally mutagenized somatic cells using a murine leukemia virus (MLV vector. This approach identified a role for 3'-phosphoadenosine 5'-phosphosulfate synthase 1 (PAPSS1, one of two enzymes that synthesize PAPS, the high energy sulfate donor used in all sulfonation reactions catalyzed by cellular sulfotransferases. The role of the cellular sulfonation pathway was confirmed using chemical inhibitors of PAPS synthases and cellular sulfotransferases. The requirement for sulfonation was mapped to a stage during or shortly after MLV provirus establishment and influenced subsequent gene expression from the viral long terminal repeat (LTR promoter. Infection of cells by an HIV vector was also shown to be highly dependent on the cellular sulfonation pathway. These studies have uncovered a heretofore unknown regulatory step of retroviral replication, have defined a new biological function for sulfonation in nuclear gene expression, and provide a potentially valuable new target for HIV/AIDS therapy.

  3. Rhodiolae Kirliowii Radix et Rhizoma and Crataegus pinnatifida Fructus Extracts Effectively Inhibit BK Virus and JC Virus Infection of Host Cells

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    San-Yuan Chen

    2017-01-01

    Full Text Available The human polyomaviruses BK (BKPyV and JC (JCPyV are ubiquitous pathogens long associated with severe disease in immunocompromised individuals. BKPyV causes polyomavirus-associated nephropathy and hemorrhagic cystitis, whereas JCPyV is the causative agent of the fatal demyelinating disease progressive multifocal leukoencephalopathy. No effective therapies targeting these viruses are currently available. The goal of this study was to identify Chinese medicinal herbs with antiviral activity against BKPyV and JCPyV. We screened extracts of Chinese medicinal herbs for the ability to inhibit hemagglutination by BKPyV and JCPyV virus-like particles (VLPs and the ability to inhibit BKPyV and JCPyV binding and infection of host cells. Two of the 40 herbal extracts screened, Rhodiolae Kirliowii Radix et Rhizoma and Crataegus pinnatifida Fructus, had hemagglutination inhibition activity on BKPyV and JCPyV VLPs and further inhibited infection of the cells by BKPyV and JCPyV, as evidenced by reduced expression of viral proteins in BKPyV-infected and JCPyV-infected cells after treatment with Rhodiolae Kirliowii Radix et Rhizoma or Crataegus pinnatifida Fructus extract. The results in this work show that both Rhodiolae Kirliowii Radix et Rhizoma and Crataegus pinnatifida Fructus may be sources of potential antiviral compounds for treating BKPyV and JCPyV infections.

  4. Host Cell Responses to Persistent Mycoplasmas - Different Stages in Infection of HeLa Cells with Mycoplasma hominis

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    Hopfe, Miriam; Deenen, René; Degrandi, Daniel; Köhrer, Karl; Henrich, Birgit

    2013-01-01

    Mycoplasma hominis is a facultative human pathogen primarily associated with bacterial vaginosis and pelvic inflammatory disease, but it is also able to spread to other sites, leading to arthritis or, in neonates, meningitis. With a minimal set of 537 annotated genes, M. hominis is the second smallest self-replicating mycoplasma and thus an ideal model organism for studying the effects of an infectious agent on its host more closely. M. hominis adherence, colonisation and invasion of HeLa cells were characterised in a time-course study using scanning electron microscopy, confocal microscopy and microarray-based analysis of the HeLa cell transcriptome. At 4 h post infection, cytoadherence of M. hominis to the HeLa cell surface was accompanied by differential regulation of 723 host genes (>2 fold change in expression). Genes associated with immune responses and signal transduction pathways were mainly affected and components involved in cell-cycle regulation, growth and death were highly upregulated. At 48 h post infection, when mycoplasma invasion started, 1588 host genes were differentially expressed and expression of genes for lysosome-specific proteins associated with bacterial lysis was detected. In a chronically infected HeLa cell line (2 weeks), the proportion of intracellular mycoplasmas reached a maximum of 10% and M. hominis-filled protrusions of the host cell membrane were seen by confocal microscopy, suggesting exocytotic dissemination. Of the 1972 regulated host genes, components of the ECM-receptor interaction pathway and phagosome-related integrins were markedly increased. The immune response was quite different to that at the beginning of infection, with a prominent induction of IL1B gene expression, affecting pathways of MAPK signalling, and genes connected with cytokine-cytokine interactions and apoptosis. These data show for the first time the complex, time-dependent reaction of the host directed at mycoplasmal clearance and the counter measures of

  5. Temperature shift and host cell contact up-regulate sporozoite expression of Plasmodium falciparum genes involved in hepatocyte infection.

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    Anthony Siau

    Full Text Available Plasmodium sporozoites are deposited in the skin by Anopheles mosquitoes. They then find their way to the liver, where they specifically invade hepatocytes in which they develop to yield merozoites infective to red blood cells. Relatively little is known of the molecular interactions during these initial obligatory phases of the infection. Recent data suggested that many of the inoculated sporozoites invade hepatocytes an hour or more after the infective bite. We hypothesised that this pre-invasive period in the mammalian host prepares sporozoites for successful hepatocyte infection. Therefore, the genes whose expression becomes modified prior to hepatocyte invasion would be those likely to code for proteins implicated in the subsequent events of invasion and development. We have used P. falciparum sporozoites and their natural host cells, primary human hepatocytes, in in vitro co-culture system as a model for the pre-invasive period. We first established that under co-culture conditions, sporozoites maintain infectivity for an hour or more, in contrast to a drastic loss in infectivity when hepatocytes were not included. Thus, a differential transcriptome of salivary gland sporozoites versus sporozoites co-cultured with hepatocytes was established using a pan-genomic P. falciparum microarray. The expression of 532 genes was found to have been up-regulated following co-culture. A fifth of these genes had no orthologues in the genomes of Plasmodium species used in rodent models of malaria. Quantitative RT-PCR analysis of a selection of 21 genes confirmed the reliability of the microarray data. Time-course analysis further indicated two patterns of up-regulation following sporozoite co-culture, one transient and the other sustained, suggesting roles in hepatocyte invasion and liver stage development, respectively. This was supported by functional studies of four hitherto uncharacterized proteins of which two were shown to be sporozoite surface

  6. Direct identification of the Meloidogyne incognita secretome reveals proteins with host cell reprogramming potential.

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    Stéphane Bellafiore

    2008-10-01

    Full Text Available The root knot nematode, Meloidogyne incognita, is an obligate parasite that causes significant damage to a broad range of host plants. Infection is associated with secretion of proteins surrounded by proliferating cells. Many parasites are known to secrete effectors that interfere with plant innate immunity, enabling infection to occur; they can also release pathogen-associated molecular patterns (PAMPs, e.g., flagellin that trigger basal immunity through the nematode stylet into the plant cell. This leads to suppression of innate immunity and reprogramming of plant cells to form a feeding structure containing multinucleate giant cells. Effectors have generally been discovered using genetics or bioinformatics, but M. incognita is non-sexual and its genome sequence has not yet been reported. To partially overcome these limitations, we have used mass spectrometry to directly identify 486 proteins secreted by M. incognita. These proteins contain at least segmental sequence identity to those found in our 3 reference databases (published nematode proteins; unpublished M. incognita ESTs; published plant proteins. Several secreted proteins are homologous to plant proteins, which they may mimic, and they contain domains that suggest known effector functions (e.g., regulating the plant cell cycle or growth. Others have regulatory domains that could reprogram cells. Using in situ hybridization we observed that most secreted proteins were produced by the subventral glands, but we found that phasmids also secreted proteins. We annotated the functions of the secreted proteins and classified them according to roles they may play in the development of root knot disease. Our results show that parasite secretomes can be partially characterized without cognate genomic DNA sequence. We observed that the M. incognita secretome overlaps the reported secretome of mammalian parasitic nematodes (e.g., Brugia malayi, suggesting a common parasitic behavior and a possible

  7. Differences between Mycobacterium-Host Cell Relationships in Latent Tuberculous Infection of Mice Ex Vivo and Mycobacterial Infection of Mouse Cells In Vitro

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    Elena Ufimtseva

    2016-01-01

    Full Text Available The search for factors that account for the reproduction and survival of mycobacteria, including vaccine strains, in host cells is the priority for studies on tuberculosis. A comparison of BCG-mycobacterial loads in granuloma cells obtained from bone marrow and spleens of mice with latent tuberculous infection and cells from mouse bone marrow and peritoneal macrophage cultures infected with the BCG vaccine in vitro has demonstrated that granuloma macrophages each normally contained a single BCG-Mycobacterium, while those acutely infected in vitro had increased mycobacterial loads and death rates. Mouse granuloma cells were observed to produce the IFNγ, IL-1α, GM-CSF, CD1d, CD25, CD31, СD35, and S100 proteins. None of these activation markers were found in mouse cell cultures infected in vitro or in intact macrophages. Lack of colocalization of lipoarabinomannan-labeled BCG-mycobacteria with the lysosomotropic LysoTracker dye in activated granuloma macrophages suggests that these macrophages were unable to destroy BCG-mycobacteria. However, activated mouse granuloma macrophages could control mycobacterial reproduction in cells both in vivo and in ex vivo culture. By contrast, a considerable increase in the number of BCG-mycobacteria was observed in mouse bone marrow and peritoneal macrophages after BCG infection in vitro, when no expression of the activation-related molecules was detected in these cells.

  8. The structural proteins of epidemic and historical strains of Zika virus differ in their ability to initiate viral infection in human host cells.

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    Bos, Sandra; Viranaicken, Wildriss; Turpin, Jonathan; El-Kalamouni, Chaker; Roche, Marjolaine; Krejbich-Trotot, Pascale; Desprès, Philippe; Gadea, Gilles

    2018-03-01

    Mosquito-borne Zika virus (ZIKV) recently emerged in South Pacific islands and Americas where large epidemics were documented. In the present study, we investigated the contribution of the structural proteins C, prM and E in the permissiveness of human host cells to epidemic strains of ZIKV. To this end, we evaluated the capacity of the epidemic strain BeH819015 to infect epithelial A549 and neuronal SH-SY5Y cells in comparison to the African historical MR766 strain. For that purpose, we generated a molecular clone of BeH819015 and a chimeric clone of MR766 which contains the BeH819015 structural protein region. We showed that ZIKV containing BeH819015 structural proteins was much less efficient in cell-attachment leading to a reduced susceptibility of A549 and SH-SY5Y cells to viral infection. Our data illustrate a previously underrated role for C, prM, and E in ZIKV epidemic strain ability to initiate viral infection in human host cells. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  9. Ebola virus host cell entry.

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    Sakurai, Yasuteru

    2015-01-01

    Ebola virus is an enveloped virus with filamentous structure and causes a severe hemorrhagic fever in human and nonhuman primates. Host cell entry is the first essential step in the viral life cycle, which has been extensively studied as one of the therapeutic targets. A virus factor of cell entry is a surface glycoprotein (GP), which is an only essential viral protein in the step, as well as the unique particle structure. The virus also interacts with a lot of host factors to successfully enter host cells. Ebola virus at first binds to cell surface proteins and internalizes into cells, followed by trafficking through endosomal vesicles to intracellular acidic compartments. There, host proteases process GPs, which can interact with an intracellular receptor. Then, under an appropriate circumstance, viral and endosomal membranes are fused, which is enhanced by major structural changes of GPs, to complete host cell entry. Recently the basic research of Ebola virus infection mechanism has markedly progressed, largely contributed by identification of host factors and detailed structural analyses of GPs. This article highlights the mechanism of Ebola virus host cell entry, including recent findings.

  10. Relationship between VacA Toxin and Host Cell Autophagy in Helicobacter pylori Infection of the Human Stomach: A Few Answers, Many Questions

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    Vittorio Ricci

    2016-07-01

    Full Text Available Helicobacter pylori is a Gram-negative bacterium that colonizes the stomach of about half the global population and represents the greatest risk factor for gastric malignancy. The relevance of H. pylori for gastric cancer development is equivalent to that of tobacco smoking for lung cancer. VacA toxin seems to play a pivotal role in the overall strategy of H. pylori towards achieving persistent gastric colonization. This strategy appears to involve the modulation of host cell autophagy. After an overview of autophagy and its role in infection and carcinogenesis, I critically review current knowledge about the action of VacA on host cell autophagy during H. pylori infection of the human stomach. Although VacA is a key player in modulation of H. pylori-induced autophagy, a few discrepancies in the data are also evident and many questions remain to be answered. We are thus still far from a definitive understanding of the molecular mechanisms through which VacA affects autophagy and the consequences of this toxin action on the overall pathogenic activity of H. pylori.

  11. Unfolding Role of a Danger Molecule Adenosine Signaling in Modulation of Microbial Infection and Host Cell Response

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    Jaden S. Lee

    2018-01-01

    Full Text Available Ectonucleotidases CD39 and CD73, specific nucleotide metabolizing enzymes located on the surface of the host, can convert a pro-inflammatory environment driven by a danger molecule extracellular-ATP to an adenosine-mediated anti-inflammatory milieu. Accordingly, CD39/CD73 signaling has been strongly implicated in modulating the intensity, duration, and composition of purinergic danger signals delivered to host. Recent studies have eluted potential roles for CD39 and CD73 in selective triggering of a variety of host immune cells and molecules in the presence of pathogenic microorganisms or microbial virulence molecules. Growing evidence also suggests that CD39 and CD73 present complimentary, but likely differential, actions against pathogens to shape the course and severity of microbial infection as well as the associated immune response. Similarly, adenosine receptors A2A and A2B have been proposed to be major immunomodulators of adenosine signaling during chronic inflammatory conditions induced by opportunistic pathogens, such as oral colonizer Porphyromonas gingivalis. Therefore, we here review the recent studies that demonstrate how complex network of molecules in the extracellular adenosine signaling machinery and their interactions can reshape immune responses and may also be targeted by opportunistic pathogens to establish successful colonization in human mucosal tissues and modulate the host immune response.

  12. Distinct temporal changes in host cell lncRNA expression during the course of an adenovirus infection

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    Zhao, Hongxing, E-mail: Hongxing.Zhao@igp.uu.se [The Beijer Laboratory, Dept. of Immunology, Genetics and Pathology, Uppsala University, S-751 85 Uppsala (Sweden); Chen, Maoshan [Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria 3086 (Australia); Lind, Sara Bergström [Department of Chemistry-BMC, Analytical Chemistry, Science for Life Laboratory, Uppsala University, Box 599, SE-751 24 Uppsala (Sweden); Pettersson, Ulf [The Beijer Laboratory, Dept. of Immunology, Genetics and Pathology, Uppsala University, S-751 85 Uppsala (Sweden)

    2016-05-15

    The deregulation of cellular long non-coding RNA (lncRNA) expression during a human adenovirus infection was studied by deep sequencing. Expression of lncRNAs increased substantially following the progression of the infection. Among 645 significantly expressed lncRNAs, the expression of 398 was changed more than 2-fold. More than 80% of them were up-regulated and 80% of them were detected during the late phase. Based on the genomic locations of the deregulated lncRNAs in relation to known mRNAs and miRNAs, they were predicted to be involved in growth, structure, apoptosis and wound healing in the early phase, cell proliferation in the intermediate phase and protein synthesis, modification and transport in the late phase. The most significant functions of cellular RNA-binding proteins, previously shown to interact with the deregulated lncRNAs identified here, are involved in RNA splicing, nuclear export and translation events. We hypothesize that adenoviruses exploit the lncRNA network to optimize their reproduction. - Highlights: • The expression of 398 lncRNAs showed a distinct temporal pattern during Ad2 infection. • 80% of the deregulated lncRNAs were up-regulated during the late phase of infection. • The deregulated lncRNAs potentiallyinteract with 33 cellular RNA binding proteins. • These RBPs are involved in RNA splicing, nuclear export and translation. • Adenovirus exploits the cellular lncRNA network to optimize its replication.

  13. Structural Basis of the Interaction of a Trypanosoma cruzi Surface Molecule Implicated in Oral Infection with Host Cells and Gastric Mucin

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    Cortez, Cristian; Yoshida, Nobuko; Bahia, Diana; Sobreira, Tiago J.P.

    2012-01-01

    Host cell invasion and dissemination within the host are hallmarks of virulence for many pathogenic microorganisms. As concerns Trypanosoma cruzi, which causes Chagas disease, the insect vector-derived metacyclic trypomastigotes (MT) initiate infection by invading host cells, and later blood trypomastigotes disseminate to diverse organs and tissues. Studies with MT generated in vitro and tissue culture-derived trypomastigotes (TCT), as counterparts of insect-borne and bloodstream parasites, have implicated members of the gp85/trans-sialidase superfamily, MT gp82 and TCT Tc85-11, in cell invasion and interaction with host factors. Here we analyzed the gp82 structure/function characteristics and compared them with those previously reported for Tc85-11. One of the gp82 sequences identified as a cell binding site consisted of an α-helix, which connects the N-terminal β-propeller domain to the C-terminal β-sandwich domain where the second binding site is nested. In the gp82 structure model, both sites were exposed at the surface. Unlike gp82, the Tc85-11 cell adhesion sites are located in the N-terminal β-propeller region. The gp82 sequence corresponding to the epitope for a monoclonal antibody that inhibits MT entry into target cells was exposed on the surface, upstream and contiguous to the α-helix. Located downstream and close to the α-helix was the gp82 gastric mucin binding site, which plays a central role in oral T. cruzi infection. The sequences equivalent to Tc85-11 laminin-binding sites, which have been associated with the parasite ability to overcome extracellular matrices and basal laminae, was poorly conserved in gp82, compatible with its reduced capacity to bind laminin. Our study indicates that gp82 is structurally suited for MT to initiate infection by the oral route, whereas Tc85-11, with its affinity for laminin, would facilitate the parasite dissemination through diverse organs and tissues. PMID:22860068

  14. Enhanced mutagenesis of UV-irradiated simian virus 40 occurs in mitomycin C-treated host cells only at a low multiplicity of infection

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    Sarasin, A.; Benoit, A.

    1986-01-01

    Treatment of monkey kidney cells with mitomycin C (MMC) 24 h prior to infection with UV-irradiated simian virus 40 (SV40) enhanced both virus survival and virus mutagenesis. The use of SV40 as a biological probe has been taken as an easy method to analyse SOS response of mammalian cells to the stress caused by DNA damage or inhibition of DNA replication. The mutation assay we used was based on the reversion from a temperature-sensitive phenotype (tsA58 mutant) to a wild-type phenotype. The optimal conditions for producing enhanced survival and mutagenesis in the virus progeny were determined with regard to the multiplicity of infection (MOI). Results showed that the level of enhanced mutagenesis observed for UV-irradiated virus grown in MMC-treated cells was an inverse function of the MOI, while enhanced survival was observed at nearly the same level regardless of the MOI. For the unirradiated virus, almost no increase in the mutation of virus progeny issued from MMC-treated cells was observed, while a small amount of enhanced virus survival was obtained. These results show that enhanced virus mutagenesis and enhanced virus survival can be dissociated under some experimental conditions. Enhanced virus mutagenesis, analogous to the error-prone replication of phages in SOS-induced bacteria, was observed, at least for SV40, only when DNA of both virus and host cells was damaged and when infection occurred with a small number of viral particles. We therefore hypothesize that an error-prone replication mode of UV-damaged templates is observed in induced monkey kidney cells

  15. In vitro studies of Rickettsia-host cell interactions: Confocal laser scanning microscopy of Rickettsia helvetica-infected eukaryotic cell lines.

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    Speck, Stephanie; Kern, Tanja; Aistleitner, Karin; Dilcher, Meik; Dobler, Gerhard; Essbauer, Sandra

    2018-02-01

    Rickettsia (R.) helvetica is the most prevalent rickettsia found in Ixodes ricinus ticks in Germany. Several studies reported antibodies against R. helvetica up to 12.5% in humans investigated, however, fulminant clinical cases are rare indicating a rather low pathogenicity compared to other rickettsiae. We investigated growth characteristics of R. helvetica isolate AS819 in two different eukaryotic cell lines with focus on ultra-structural changes of host cells during infection determined by confocal laser scanning microscopy. Further investigations included partially sequencing of rickA, sca4 and sca2 genes, which have been reported to encode proteins involved in cell-to-cell spread and virulence in some rickettsiae. R. helvetica grew constantly but slowly in both cell lines used. Confocal laser scanning microscopy revealed that the dissemination of R. helvetica AS819 in both cell lines was rather mediated by cell break-down and bacterial release than cell-to-cell spread. The cytoskeleton of both investigated eukaryotic cell lines was not altered. R. helvetica possesses rickA, but its expression is not sufficient to promote actin-based motility as demonstrated by confocal laser scanning microscopy. Hypothetical Sca2 and Sca4 proteins were deduced from nucleotide gene sequences but the predicted amino acid sequences were disrupted or truncated compared to other rickettsiae most likely resulting in non-functional proteins. Taken together, these results might give a first hint to the underlying causes of the reduced virulence and pathogenicity of R. helvetica.

  16. Genetic reprogramming of host cells by bacterial pathogens.

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    Tran Van Nhieu, Guy; Arbibe, Laurence

    2009-10-29

    During the course of infection, pathogens often induce changes in gene expression in host cells and these changes can be long lasting and global or transient and of limited amplitude. Defining how, when, and why bacterial pathogens reprogram host cells represents an exciting challenge that opens up the opportunity to grasp the essence of pathogenesis and its molecular details.

  17. The Coxiella Burnetii type IVB secretion system (T4BSS) component DotA is released/secreted during infection of host cells and during in vitro growth in a T4BSS-dependent manner.

    Science.gov (United States)

    Luedtke, Brandon E; Mahapatra, Saugata; Lutter, Erika I; Shaw, Edward I

    2017-06-01

    Coxiella burnetii is a Gram-negative intracellular pathogen and is the causative agent of the zoonotic disease Q fever. To cause disease, C. burnetii requires a functional type IVB secretion system (T4BSS) to transfer effector proteins required for the establishment and maintenance of a membrane-bound parasitophorous vacuole (PV) and further modulation of host cell process. However, it is not clear how the T4BSS interacts with the PV membrane since neither a secretion pilus nor an extracellular pore forming apparatus has not been described. To address this, we used the acidified citrate cysteine medium (ACCM) along with cell culture infection and immunological techniques to identify the cellular and extracellular localization of T4BSS components. Interestingly, we found that DotA and IcmX were secreted/released in a T4BSS-dependent manner into the ACCM. Analysis of C. burnetii-infected cell lines revealed that DotA colocalized with the host cell marker CD63 (LAMP3) at the PV membrane. In the absence of bacterial protein synthesis, DotA also became depleted from the PV membrane. These data are the first to identify the release/secretion of C. burnetii T4BSS components during axenic growth and the interaction of a T4BSS component with the PV membrane during infection of host cells. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  18. Human subtilase SKI-1/S1P is a master regulator of the HCV Lifecycle and a potential host cell target for developing indirect-acting antiviral agents.

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    Andrea D Olmstead

    2012-01-01

    Full Text Available HCV infection is a major risk factor for liver cancer and liver transplantation worldwide. Overstimulation of host lipid metabolism in the liver by HCV-encoded proteins during viral infection creates a favorable environment for virus propagation and pathogenesis. In this study, we hypothesize that targeting cellular enzymes acting as master regulators of lipid homeostasis could represent a powerful approach to developing a novel class of broad-spectrum antivirals against infection associated with human Flaviviridae viruses such as hepatitis C virus (HCV, whose assembly and pathogenesis depend on interaction with lipid droplets (LDs. One such master regulator of cholesterol metabolic pathways is the host subtilisin/kexin-isozyme-1 (SKI-1--or site-1 protease (S1P. SKI-1/S1P plays a critical role in the proteolytic activation of sterol regulatory element binding proteins (SREBPs, which control expression of the key enzymes of cholesterol and fatty-acid biosynthesis. Here we report the development of a SKI-1/S1P-specific protein-based inhibitor and its application to blocking the SREBP signaling cascade. We demonstrate that SKI-1/S1P inhibition effectively blocks HCV from establishing infection in hepatoma cells. The inhibitory mechanism is associated with a dramatic reduction in the abundance of neutral lipids, LDs, and the LD marker: adipose differentiation-related protein (ADRP/perilipin 2. Reduction of LD formation inhibits virus assembly from infected cells. Importantly, we confirm that SKI-1/S1P is a key host factor for HCV infection by using a specific active, site-directed, small-molecule inhibitor of SKI-1/S1P: PF-429242. Our studies identify SKI-1/S1P as both a novel regulator of the HCV lifecycle and as a potential host-directed therapeutic target against HCV infection and liver steatosis. With identification of an increasing number of human viruses that use host LDs for infection, our results suggest that SKI-1/S1P inhibitors may allow

  19. Enforcing host cell polarity: an apicomplexan parasite strategy towards dissemination.

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    Baumgartner, Martin

    2011-08-01

    The propagation of apicomplexan parasites through transmitting vectors is dependent on effective dissemination of parasites inside the mammalian host. Intracellular Toxoplasma and Theileria parasites face the challenge that their spread inside the host depends in part on the motile capacities of their host cells. In response, these parasites influence the efficiency of dissemination by altering adhesive and/or motile properties of their host cells. Theileria parasites do so by targeting signalling pathways that control host cell actin dynamics. The resulting enforced polar host cell morphology facilitates motility and invasiveness, by establishing focal adhesion and invasion structures at the leading edge of the infected cell. This parasite strategy highlights mechanisms of motility regulation that are also likely relevant for immune or cancer cell motility. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. Simultaneous transcriptional profiling of bacteria and their host cells.

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    Michael S Humphrys

    Full Text Available We developed an RNA-Seq-based method to simultaneously capture prokaryotic and eukaryotic expression profiles of cells infected with intracellular bacteria. As proof of principle, this method was applied to Chlamydia trachomatis-infected epithelial cell monolayers in vitro, successfully obtaining transcriptomes of both C. trachomatis and the host cells at 1 and 24 hours post-infection. Chlamydiae are obligate intracellular bacterial pathogens that cause a range of mammalian diseases. In humans chlamydiae are responsible for the most common sexually transmitted bacterial infections and trachoma (infectious blindness. Disease arises by adverse host inflammatory reactions that induce tissue damage & scarring. However, little is known about the mechanisms underlying these outcomes. Chlamydia are genetically intractable as replication outside of the host cell is not yet possible and there are no practical tools for routine genetic manipulation, making genome-scale approaches critical. The early timeframe of infection is poorly understood and the host transcriptional response to chlamydial infection is not well defined. Our simultaneous RNA-Seq method was applied to a simplified in vitro model of chlamydial infection. We discovered a possible chlamydial strategy for early iron acquisition, putative immune dampening effects of chlamydial infection on the host cell, and present a hypothesis for Chlamydia-induced fibrotic scarring through runaway positive feedback loops. In general, simultaneous RNA-Seq helps to reveal the complex interplay between invading bacterial pathogens and their host mammalian cells and is immediately applicable to any bacteria/host cell interaction.

  1. Salmonella modulation of host cell gene expression promotes its intracellular growth.

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    Sebastian Hannemann

    Full Text Available Salmonella Typhimurium has evolved a complex functional interface with its host cell largely determined by two type III secretion systems (T3SS, which through the delivery of bacterial effector proteins modulate a variety of cellular processes. We show here that Salmonella Typhimurium infection of epithelial cells results in a profound transcriptional reprogramming that changes over time. This response is triggered by Salmonella T3SS effector proteins, which stimulate unique signal transduction pathways leading to STAT3 activation. We found that the Salmonella-stimulated changes in host cell gene expression are required for the formation of its specialized vesicular compartment that is permissive for its intracellular replication. This study uncovers a cell-autonomous process required for Salmonella pathogenesis potentially opening up new avenues for the development of anti-infective strategies that target relevant host pathways.

  2. Salmonella modulation of host cell gene expression promotes its intracellular growth.

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    Hannemann, Sebastian; Gao, Beile; Galán, Jorge E

    2013-01-01

    Salmonella Typhimurium has evolved a complex functional interface with its host cell largely determined by two type III secretion systems (T3SS), which through the delivery of bacterial effector proteins modulate a variety of cellular processes. We show here that Salmonella Typhimurium infection of epithelial cells results in a profound transcriptional reprogramming that changes over time. This response is triggered by Salmonella T3SS effector proteins, which stimulate unique signal transduction pathways leading to STAT3 activation. We found that the Salmonella-stimulated changes in host cell gene expression are required for the formation of its specialized vesicular compartment that is permissive for its intracellular replication. This study uncovers a cell-autonomous process required for Salmonella pathogenesis potentially opening up new avenues for the development of anti-infective strategies that target relevant host pathways.

  3. The influenza fingerprints: NS1 and M1 proteins contribute to specific host cell ultrastructure signatures upon infection by different influenza A viruses

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    Terrier, Olivier; Moules, Vincent; Carron, Coralie; Cartet, Gaeelle [Equipe VirCell, Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Frobert, Emilie [Laboratoire de Virologie, Centre de Biologie et de Pathologie Est, Hospices Civils de Lyon, 59 boulevard Pinel, F-69677 Bron Cedex, Lyon (France); Yver, Matthieu; Traversier, Aurelien [Equipe VirCell, Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Wolff, Thorsten [Division of Influenza/Respiratory Viruses, Robert Koch Institute, Nordufer 20, D-13353 Berlin (Germany); Riteau, Beatrice [Laboratoire de Virologie et Pathologie Humaine, VirPath EMR 4610, Universite de Lyon, Universite Claude Bernard Lyon 1, Hospices Civils de Lyon, Faculte de medecine RTH Laennec, rue Guillaume Paradin, F-69008 Lyon (France); Naffakh, Nadia [Institut Pasteur, Unite de Genetique Moleculaire des Virus Respiratoires, URA CNRS 3015, EA302 Universite Paris Diderot, Paris (France); and others

    2012-10-10

    Influenza A are nuclear replicating viruses which hijack host machineries in order to achieve optimal infection. Numerous functional virus-host interactions have now been characterized, but little information has been gathered concerning their link to the virally induced remodeling of the host cellular architecture. In this study, we infected cells with several human and avian influenza viruses and we have analyzed their ultrastructural modifications by using electron and confocal microscopy. We discovered that infections lead to a major and systematic disruption of nucleoli and the formation of a large number of diverse viral structures showing specificity that depended on the subtype origin and genomic composition of viruses. We identified NS1 and M1 proteins as the main actors in the remodeling of the host ultra-structure and our results suggest that each influenza A virus strain could be associated with a specific cellular fingerprint, possibly correlated to the functional properties of their viral components.

  4. The influenza fingerprints: NS1 and M1 proteins contribute to specific host cell ultrastructure signatures upon infection by different influenza A viruses

    International Nuclear Information System (INIS)

    Terrier, Olivier; Moules, Vincent; Carron, Coralie; Cartet, Gaëlle; Frobert, Emilie; Yver, Matthieu; Traversier, Aurelien; Wolff, Thorsten; Riteau, Beatrice; Naffakh, Nadia

    2012-01-01

    Influenza A are nuclear replicating viruses which hijack host machineries in order to achieve optimal infection. Numerous functional virus–host interactions have now been characterized, but little information has been gathered concerning their link to the virally induced remodeling of the host cellular architecture. In this study, we infected cells with several human and avian influenza viruses and we have analyzed their ultrastructural modifications by using electron and confocal microscopy. We discovered that infections lead to a major and systematic disruption of nucleoli and the formation of a large number of diverse viral structures showing specificity that depended on the subtype origin and genomic composition of viruses. We identified NS1 and M1 proteins as the main actors in the remodeling of the host ultra-structure and our results suggest that each influenza A virus strain could be associated with a specific cellular fingerprint, possibly correlated to the functional properties of their viral components.

  5. Re-localization of Cellular Protein SRp20 during Poliovirus Infection: Bridging a Viral IRES to the Host Cell Translation Apparatus

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    Fitzgerald, Kerry D.; Semler, Bert L.

    2011-01-01

    Poliovirus IRES-mediated translation requires the functions of certain canonical as well as non-canonical factors for the recruitment of ribosomes to the viral RNA. The interaction of cellular proteins PCBP2 and SRp20 in extracts from poliovirus-infected cells has been previously described, and these two proteins were shown to function synergistically in viral translation. To further define the mechanism of ribosome recruitment for the initiation of poliovirus IRES-dependent translation, we focused on the role of the interaction between cellular proteins PCBP2 and SRp20. Work described here demonstrates that SRp20 dramatically re-localizes from the nucleus to the cytoplasm of poliovirus-infected neuroblastoma cells during the course of infection. Importantly, SRp20 partially co-localizes with PCBP2 in the cytoplasm of infected cells, corroborating our previous in vitro interaction data. In addition, the data presented implicate the presence of these two proteins in viral translation initiation complexes. We show that in extracts from poliovirus-infected cells, SRp20 is associated with PCBP2 bound to poliovirus RNA, indicating that this interaction occurs on the viral RNA. Finally, we generated a mutated version of SRp20 lacking the RNA recognition motif (SRp20ΔRRM) and found that this protein is localized similar to the full length SRp20, and also partially co-localizes with PCBP2 during poliovirus infection. Expression of this mutated version of SRp20 results in a ∼100 fold decrease in virus yield for poliovirus when compared to expression of wild type SRp20, possibly via a dominant negative effect. Taken together, these results are consistent with a model in which SRp20 interacts with PCBP2 bound to the viral RNA, and this interaction functions to recruit ribosomes to the viral RNA in a direct or indirect manner, with the participation of additional protein-protein or protein-RNA interactions. PMID:21779168

  6. Alteration of protein levels during influenza virus H1N1 infection in host cells: a proteomic survey of host and virus reveals differential dynamics.

    Directory of Open Access Journals (Sweden)

    Susann Kummer

    Full Text Available We studied the dynamics of the proteome of influenza virus A/PR/8/34 (H1N1 infected Madin-Darby canine kidney cells up to 12 hours post infection by mass spectrometry based quantitative proteomics using the approach of stable isotope labeling by amino acids in cell culture (SILAC. We identified 1311 cell proteins and, apart from the proton channel M2, all major virus proteins. Based on their abundance two groups of virus proteins could be distinguished being in line with the function of the proteins in genesis and formation of new virions. Further, the data indicate a correlation between the amount of proteins synthesized and their previously determined copy number inside the viral particle. We employed bioinformatic approaches such as functional clustering, gene ontology, and pathway (KEGG enrichment tests to uncover co-regulated cellular protein sets, assigned the individual subsets to their biological function, and determined their interrelation within the progression of viral infection. For the first time we are able to describe dynamic changes of the cellular and, of note, the viral proteome in a time dependent manner simultaneously. Through cluster analysis, time dependent patterns of protein abundances revealed highly dynamic up- and/or down-regulation processes. Taken together our study provides strong evidence that virus infection has a major impact on the cell status at the protein level.

  7. N-Terminomics TAILS Identifies Host Cell Substrates of Poliovirus and Coxsackievirus B3 3C Proteinases That Modulate Virus Infection

    Science.gov (United States)

    Jagdeo, Julienne M.; Dufour, Antoine; Klein, Theo; Solis, Nestor; Kleifeld, Oded; Kizhakkedathu, Jayachandran; Luo, Honglin; Overall, Christopher M.

    2018-01-01

    ABSTRACT Enteroviruses encode proteinases that are essential for processing of the translated viral polyprotein. In addition, viral proteinases also target host proteins to manipulate cellular processes and evade innate antiviral responses to promote replication and infection. Although some host protein substrates of enterovirus proteinases have been identified, the full repertoire of targets remains unknown. We used a novel quantitative in vitro proteomics-based approach, termed terminal amine isotopic labeling of substrates (TAILS), to identify with high confidence 72 and 34 new host protein targets of poliovirus and coxsackievirus B3 (CVB3) 3C proteinases (3Cpros) in HeLa cell and cardiomyocyte HL-1 cell lysates, respectively. We validated a subset of candidate substrates that are targets of poliovirus 3Cpro in vitro including three common protein targets, phosphoribosylformylglycinamidine synthetase (PFAS), hnRNP K, and hnRNP M, of both proteinases. 3Cpro-targeted substrates were also cleaved in virus-infected cells but not noncleavable mutant proteins designed from the TAILS-identified cleavage sites. Knockdown of TAILS-identified target proteins modulated infection both negatively and positively, suggesting that cleavage by 3Cpro promotes infection. Indeed, expression of a cleavage-resistant mutant form of the endoplasmic reticulum (ER)-Golgi vesicle-tethering protein p115 decreased viral replication and yield. As the first comprehensive study to identify and validate functional enterovirus 3Cpro substrates in vivo, we conclude that N-terminomics by TAILS is an effective strategy to identify host targets of viral proteinases in a nonbiased manner. IMPORTANCE Enteroviruses are positive-strand RNA viruses that encode proteases that cleave the viral polyprotein into the individual mature viral proteins. In addition, viral proteases target host proteins in order to modulate cellular pathways and block antiviral responses in order to facilitate virus infection

  8. Bartonella entry mechanisms into mammalian host cells.

    Science.gov (United States)

    Eicher, Simone C; Dehio, Christoph

    2012-08-01

    The Gram-negative genus Bartonella comprises arthropod-borne pathogens that typically infect mammals in a host-specific manner. Bartonella bacilliformis and Bartonella quintana are human-specific pathogens, while several zoonotic bartonellae specific for diverse animal hosts infect humans as an incidental host. Clinical manifestations of Bartonella infections range from mild symptoms to life-threatening disease. Following transmission by blood-sucking arthropods or traumatic contact with infected animals, bartonellae display sequential tropisms towards endothelial and possibly other nucleated cells and erythrocytes, the latter in a host-specific manner. Attachment to the extracellular matrix (ECM) and to nucleated cells is mediated by surface-exposed bacterial adhesins, in particular trimeric autotransporter adhesins (TAAs). The subsequent engulfment of the pathogen into a vacuolar structure follows a unique series of events whereby the pathogen avoids the endolysosomal compartments. For Bartonella henselae and assumingly most other species, the infection process is aided at different steps by Bartonella effector proteins (Beps). They are injected into host cells through the type IV secretion system (T4SS) VirB/D4 and subvert host cellular functions to favour pathogen uptake. Bacterial binding to erythrocytes is mediated by Trw, another T4SS, in a strictly host-specific manner, followed by pathogen-forced uptake involving the IalB invasin and subsequent replication and persistence within a membrane-bound intra-erythrocytic compartment. © 2012 Blackwell Publishing Ltd.

  9. Hepatitis C virus host cell interactions uncovered

    DEFF Research Database (Denmark)

    Gottwein, Judith; Bukh, Jens

    2007-01-01

      Insights into virus-host cell interactions as uncovered by Randall et al. (1) in a recent issue of PNAS further our understanding of the hepatitis C virus (HCV) life cycle, persistence, and pathogenesis and might lead to the identification of new therapeutic targets. HCV persistently infects 180...... million individuals worldwide, causing chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. The only approved treatment, combination therapy with IFN- and ribavirin, targets cellular pathways (2); however, a sustained virologic response is achieved only in approximately half of the patients...... treated. Therefore, there is a pressing need for the identification of novel drugs against hepatitis C. Although most research focuses on the development of HCV-specific antivirals, such as protease and polymerase inhibitors (3), cellular targets could be pursued and might allow the development of broad...

  10. How pathogens use linear motifs to perturb host cell networks

    KAUST Repository

    Via, Allegra; Uyar, Bora; Brun, Christine; Zanzoni, Andreas

    2015-01-01

    Molecular mimicry is one of the powerful stratagems that pathogens employ to colonise their hosts and take advantage of host cell functions to guarantee their replication and dissemination. In particular, several viruses have evolved the ability to interact with host cell components through protein short linear motifs (SLiMs) that mimic host SLiMs, thus facilitating their internalisation and the manipulation of a wide range of cellular networks. Here we present convincing evidence from the literature that motif mimicry also represents an effective, widespread hijacking strategy in prokaryotic and eukaryotic parasites. Further insights into host motif mimicry would be of great help in the elucidation of the molecular mechanisms behind host cell invasion and the development of anti-infective therapeutic strategies.

  11. RNA interference silences Microplitis demolitor bracovirus genes and implicates glc1.8 in disruption of adhesion in infected host cells

    International Nuclear Information System (INIS)

    Beck, Markus; Strand, Michael R.

    2003-01-01

    The family Polydnaviridae consists of ds-DNA viruses that are symbiotically associated with certain parasitoid wasps. PDVs are transmitted vertically but also are injected by wasps into hosts where they cause several physiological alterations including immunosuppression. The PDV genes responsible for mediating immunosuppression and other host alterations remain poorly characterized in large measure because viral mutants cannot be produced to study gene function. Here we report the use of RNA interference (RNAi) to specifically silence the glc1.8 and egf1.0 genes from Microplitis demolitor bracovirus (MdBV) in High Five cells derived from the lepidopteran Trichoplusia ni. Dose-response studies indicated that MdBV infects High Five cells and blocks the ability of these cells to adhere to culture plates. This response was very similar to what occurs in two classes of hemocytes, granular cells, and plasmatocytes, after infection by MdBV. Screening of monoclonal antibody (mAb) markers that distinguish different classes of lepidopteran hemocytes indicated that High Five cells cross-react with three mAbs that recognize granular cells from T. ni. Double-stranded RNA (dsRNA) complementary to glc1.8 specifically silenced glc1.8 expression and rescued the adhesive phenotype of High Five cells. Reciprocally, dsRNA complementary to egf1.0 silenced egf1.0 expression but had no effect on adhesion. The simplicity and potency of RNAi could be extremely useful for analysis of other PDV genes

  12. The nuclear protein Sam68 is cleaved by the FMDV 3C protease redistributing Sam68 to the cytoplasm during FMDV infection of host cells

    International Nuclear Information System (INIS)

    Lawrence, Paul; Schafer, Elizabeth A.; Rieder, Elizabeth

    2012-01-01

    Picornavirus infection can lead to disruption of nuclear pore traffic, shut-off of cell translation machinery, and cleavage of proteins involved in cellular signal transduction and the innate response to infection. Here, we demonstrated that the FMDV 3C pro induced the cleavage of nuclear RNA-binding protein Sam68 C-terminus containing the nuclear localization sequence (NLS). Consequently, it stimulated the redistribution of Sam68 to the cytoplasm. The siRNA knockdown of Sam68 resulted in a 1000-fold reduction in viral titers, which prompted us to study the effect of Sam68 on FMDV post-entry events. Interestingly, Sam68 interacts with the internal ribosomal entry site within the 5′ non-translated region of the FMDV genome, and Sam68 knockdown decreased FMDV IRES-driven activity in vitro suggesting that it could modulate translation of the viral genome. The results uncover a novel role for Sam68 in the context of picornaviruses and the proteolysis of a new cellular target of the FMDV 3C pro .

  13. Transcriptome and microRNome of Theileria annulata Host Cells

    KAUST Repository

    Rchiad, Zineb

    2016-06-01

    Tropical Theileriosis is a parasitic disease of calves with a profound economic impact caused by Theileria annulata, an apicomplexan parasite of the genus Theileria. Transmitted by Hyalomma ticks, T. annulata infects and transforms bovine lymphocytes and macrophages into a cancer-like phenotype characterized by all six hallmarks of cancer. In the current study we investigate the transcriptional landscape of T. annulata-infected lymphocytes to define genes and miRNAs regulated by host cell transformation using next generation sequencing. We also define genes and miRNAs differentially expressed as a result of the attenuation of a T.annulata-infected macrophage cell line used as a vaccine. By comparing the transcriptional landscape of one attenuated and two transformed cell lines we identify four genes that we propose as key factors in transformation and virulence of the T. annulata host cells. We also identify miR- 126-5p as a key regulator of infected cells proliferation, adhesion, survival and invasiveness. In addition to the host cell trascriptome we studied T. annulata transcriptome and identified the role of ROS and TGF-β2 in controlling parasite gene expression. Moreover, we have used the deep parasite ssRNA-seq data to refine the available T. annulata annotation. Taken together, this study provides the full list of host cell’s genes and miRNAs transcriptionally perturbed after infection with T. annulata and after attenuation and describes genes and miRNAs never identified before as players in this type of host cell transformation. Moreover, this study provides the first database for the transcriptome of T. annulata and its host cells using next generation sequencing.

  14. Nuclear entry of poliovirus protease-polymerase precursor 3CD: implications for host cell transcription shut-off

    International Nuclear Information System (INIS)

    Sharma, Rakhi; Raychaudhuri, Santanu; Dasgupta, Asim

    2004-01-01

    Host cell transcription mediated by all three RNA polymerases is rapidly inhibited after infection of mammalian cells with poliovirus (PV). Both genetic and biochemical studies have shown that the virus-encoded protease 3C cleaves the TATA-binding protein and other transcription factors at glutamine-glycine sites and is directly responsible for host cell transcription shut-off. PV replicates in the cytoplasm of infected cells. To shut-off host cell transcription, 3C or a precursor of 3C must enter the nucleus of infected cells. Although the 3C protease itself lacks a nuclear localization signal (NLS), amino acid sequence examination of 3D identified a potential single basic type NLS, KKKRD, spanning amino acids 125-129 within this polypeptide. Thus, a plausible scenario is that 3C enters the nucleus in the form of its precursor, 3CD, which then generates 3C by auto-proteolysis ultimately leading to cleavage of transcription factors in the nucleus. Using transient transfection of enhanced green fluorescent protein (EGFP) fusion polypeptides, we demonstrate here that both 3CD and 3D are capable of entering the nucleus in PV-infected cells. However, both polypeptides remain in the cytoplasm in uninfected HeLa cells. Mutagenesis of the NLS sequence in 3D prevents nuclear entry of 3D and 3CD in PV-infected cells. We also demonstrate that 3CD can be detected in the nuclear fraction from PV-infected HeLa cells as early as 2 h postinfection. Significant amount of 3CD is found associated with the nuclear fraction by 3-4 h of infection. Taken together, these results suggest that both the 3D NLS and PV infection are required for the entry of 3CD into the nucleus and that this may constitute a means by which viral protease 3C is delivered into the nucleus leading to host cell transcription shut-off

  15. Host cell responses to dengue virus infection

    NARCIS (Netherlands)

    Diosa Toro, Mayra

    2017-01-01

    Dengue (ook wel knokkelkoorts) is de meest voorkomende virale infectieziekte dat wordt overgedragen door muggen in de wereld met naar schatting 390 miljoen infecties per jaar. Ondanks de grote klinische impact en economische schade van het dengue virus is er nog steeds geen behandeling beschikbaar.

  16. Trichomonas vaginalis exosomes deliver cargo to host cells and mediate host∶parasite interactions.

    Directory of Open Access Journals (Sweden)

    Olivia Twu

    Full Text Available Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogential tract where it remains extracellular and adheres to epithelial cells. Infections range from asymptomatic to highly inflammatory, depending on the host and the parasite strain. Here, we use a combination of methodologies including cell fractionation, immunofluorescence and electron microscopy, RNA, proteomic and cytokine analyses and cell adherence assays to examine pathogenic properties of T. vaginalis. We have found that T.vaginalis produces and secretes microvesicles with physical and biochemical properties similar to mammalian exosomes. The parasite-derived exosomes are characterized by the presence of RNA and core, conserved exosomal proteins as well as parasite-specific proteins. We demonstrate that T. vaginalis exosomes fuse with and deliver their contents to host cells and modulate host cell immune responses. Moreover, exosomes from highly adherent parasite strains increase the adherence of poorly adherent parasites to vaginal and prostate epithelial cells. In contrast, exosomes from poorly adherent strains had no measurable effect on parasite adherence. Exosomes from parasite strains that preferentially bind prostate cells increased binding of parasites to these cells relative to vaginal cells. In addition to establishing that parasite exosomes act to modulate host∶parasite interactions, these studies are the first to reveal a potential role for exosomes in promoting parasite∶parasite communication and host cell colonization.

  17. Phage adsorption and lytic propagation in Lactobacillus plantarum: Could host cell starvation affect them?

    OpenAIRE

    Briggiler Marc?, Mari?ngeles; Reinheimer, Jorge; Quiberoni, Andrea

    2015-01-01

    Background Bacteriophages constitute a great threat to the activity of lactic acid bacteria used in industrial processes. Several factors can influence the infection cycle of bacteriophages. That is the case of the physiological state of host cells, which could produce inhibition or delay of the phage infection process. In the present work, the influence of Lactobacillus plantarum host cell starvation on phage B1 adsorption and propagation was investigated. Result First, cell growth kinetics ...

  18. Host cell reactivation in mammalian cells

    International Nuclear Information System (INIS)

    Lytle, C.D.; Benane, S.G.; Stafford, J.E.

    1976-01-01

    The survival of UV-irradiated herpes simplex virus was determined in cultured Potoroo (a marsupial) and human cells under lighting conditions which promoted photereactivation. Photoreactivation was readily demonstrated for herpes virus in two lines of Potoroo cells with dose reduction factors of 0.7 to 0.8 for ovary cells and 0.5 to 0.7 for kidney cells. Light from Blacklite (near UV) lamps was more effective than from Daylight (mostly visible) lamps, suggesting that near UV radiation was more effecient for photoreactivation in Potoroo cells. The quantitative and qualitative aspects of this photoreactivation were similar to those reported for a similar virus infecting chick embryo cells. UV-survival curves of herpes virus in Potoroo cells indicated a high level of 'dark' host cell reactivation. No photoreactivation was found for UV-irradiated vaccinia virus in Potoroo cells. A similar photoreactivation study was done using special control lighting (lambda>600 nm) and human cells with normal repair and with cells deficient in excision repair (XP). No photoreactivation was found for UV-irradiated herpes virus in either human cell with either Blacklite or Daylight lamps as the sources of photoreactivating light. This result contrasts with a report of photoreactivation for a herpes virus in the same XP cells using incandescent lamps. (author)

  19. Inhibition of host cell protein synthesis by UV-inactivated poliovirus

    International Nuclear Information System (INIS)

    Helentjaris, T.; Ehrenfeld, E.

    1977-01-01

    The ability of poliovirus that was irradiated with UV light at energies up to 2,160 ergs/mm 2 to subsequently inhibit host cell protein synthesis was measured. The inactivation of the host cell shutoff function followed one-hit kinetics. Increasing irradiation did not affect the rate of inhibition until the multiplicity of infection after irradiation was reduced to approximately 1 PFU/cell. At higher functional multiplicities, the rate was unchanged, but an increasing lag before the onset of inhibition was observed with increasing irradiation. The energy levels required to inactivate virus-induced inhibition of host cell protein synthesis suggest that damage to virus RNA rather than to virus capsid proteins is responsible for the loss of function. When the inactivation of host cell shutoff was compared with the inactivation of other viral functions by UV irradiation, it correlated exactly with the loss of infectivity but not with other viral functions measured. Guanidine treatment, which prevents detectable viral RNA and protein synthesis, completely inhibited host cell shutoff by low multiplicities of unirradiated virus infection but not higher multiplicities. When a high multiplicity of virus was first reduced to a low titer by irradiation, host cell shutoff was still evident in the presence of guanidine. The results demonstrate that the complete inhibition of host cell protein synthesis can be accomplished by one infectious viral genome per cell

  20. A T4SS Effector Targets Host Cell Alpha-Enolase Contributing to Brucella abortus Intracellular Lifestyle.

    Science.gov (United States)

    Marchesini, María I; Morrone Seijo, Susana M; Guaimas, Francisco F; Comerci, Diego J

    2016-01-01

    Brucella abortus , the causative agent of bovine brucellosis, invades and replicates within cells inside a membrane-bound compartment known as the Brucella containing vacuole (BCV). After trafficking along the endocytic and secretory pathways, BCVs mature into endoplasmic reticulum-derived compartments permissive for bacterial replication. Brucella Type IV Secretion System (VirB) is a major virulence factor essential for the biogenesis of the replicative organelle. Upon infection, Brucella uses the VirB system to translocate effector proteins from the BCV into the host cell cytoplasm. Although the functions of many translocated proteins remain unknown, some of them have been demonstrated to modulate host cell signaling pathways to favor intracellular survival and replication. BPE123 (BAB2_0123) is a B. abortus VirB-translocated effector protein recently identified by our group whose function is yet unknown. In an attempt to identify host cell proteins interacting with BPE123, a pull-down assay was performed and human alpha-enolase (ENO-1) was identified by LC/MS-MS as a potential interaction partner of BPE123. These results were confirmed by immunoprecipitation assays. In bone-marrow derived macrophages infected with B. abortus , ENO-1 associates to BCVs in a BPE123-dependent manner, indicating that interaction with translocated BPE123 is also occurring during the intracellular phase of the bacterium. Furthermore, ENO-1 depletion by siRNA impaired B. abortus intracellular replication in HeLa cells, confirming a role for α-enolase during the infection process. Indeed, ENO-1 activity levels were enhanced upon B. abortus infection of THP-1 macrophagic cells, and this activation is highly dependent on BPE123. Taken together, these results suggest that interaction between BPE123 and host cell ENO-1 contributes to the intracellular lifestyle of B. abortus .

  1. Fungal invasion of normally non-phagocytic host cells.

    Directory of Open Access Journals (Sweden)

    Scott G Filler

    2006-12-01

    Full Text Available Many fungi that cause invasive disease invade host epithelial cells during mucosal and respiratory infection, and subsequently invade endothelial cells during hematogenous infection. Most fungi invade these normally non-phagocytic host cells by inducing their own uptake. Candida albicans hyphae interact with endothelial cells in vitro by binding to N-cadherin on the endothelial cell surface. This binding induces rearrangement of endothelial cell microfilaments, which results in the endocytosis of the organism. The capsule of Cryptococcus neoformans is composed of glucuronoxylomannan, which binds specifically to brain endothelial cells, and appears to mediate both adherence and induction of endocytosis. The mechanisms by which other fungal pathogens induce their own uptake are largely unknown. Some angioinvasive fungi, such as Aspergillus species and the Zygomycetes, invade endothelial cells from the abluminal surface during the initiation of invasive disease, and subsequently invade the luminal surface of endothelial cells during hematogenous dissemination. Invasion of normally non-phagocytic host cells has different consequences, depending on the type of invading fungus. Aspergillus fumigatus blocks apoptosis of pulmonary epithelial cells, whereas Paracoccidioides brasiliensis induces apoptosis of epithelial cells. This review summarizes the mechanisms by which diverse fungal pathogens invade normally non-phagocytic host cells and discusses gaps in our knowledge that provide opportunities for future research.

  2. Host cell tropism mediated by Australian bat lyssavirus envelope glycoproteins.

    Science.gov (United States)

    Weir, Dawn L; Smith, Ina L; Bossart, Katharine N; Wang, Lin-Fa; Broder, Christopher C

    2013-09-01

    Australian bat lyssavirus (ABLV) is a rhabdovirus of the lyssavirus genus capable of causing fatal rabies-like encephalitis in humans. There are two variants of ABLV, one circulating in pteropid fruit bats and another in insectivorous bats. Three fatal human cases of ABLV infection have been reported with the third case in 2013. Importantly, two equine cases also arose in 2013; the first occurrence of ABLV in a species other than bats or humans. We examined the host cell entry of ABLV, characterizing its tropism and exploring its cross-species transmission potential using maxGFP-encoding recombinant vesicular stomatitis viruses that express ABLV G glycoproteins. Results indicate that the ABLV receptor(s) is conserved but not ubiquitous among mammalian cell lines and that the two ABLV variants can utilize alternate receptors for entry. Proposed rabies virus receptors were not sufficient to permit ABLV entry into resistant cells, suggesting that ABLV utilizes an unknown alternative receptor(s). Published by Elsevier Inc.

  3. Contribution of MS-based proteomics to the understanding of Herpes Simplex Virus type 1 interaction with host cells

    Directory of Open Access Journals (Sweden)

    Enrique eSantamaría

    2012-03-01

    Full Text Available Like other DNA viruses, Herpes Simplex Virus type 1 (HSV-1 replicates and proliferates in host cells continuously modulating the host molecular environment. Following a sophisticated temporal expression pattern, HSV-1 encodes at least 89 multifunctional proteins that interplay with and modify the host cell proteome. During the last decade, advances in mass spectrometry applications coupled to the development of proteomic separation methods have allowed to partially monitor the impact of HSV-1 infection in human cells. In this review, we discuss the current use of different proteome fractionation strategies to define HSV-1 targets on two major application areas: i viral protein interactomics to decipher viral protein interactions in host cells and ii differential quantitative proteomics to analyse the virally induced changes in the cellular proteome. Moreover, we will also discuss the potential application of high throughput proteomic approaches to study global proteome dynamics and also post-translational modifications in HSV-1-infected cells, what will greatly improved our molecular knowledge of HSV-1 infection.

  4. A Carbohydrate Moiety of Secreted Stage-Specific Glycoprotein 4 Participates in Host Cell Invasion by Trypanosoma cruzi Extracellular Amastigotes

    Directory of Open Access Journals (Sweden)

    Pilar T. V. Florentino

    2018-04-01

    Full Text Available Trypanosoma cruzi is the etiologic agent of Chagas’ disease. It is known that amastigotes derived from trypomastigotes in the extracellular milieu are infective in vitro and in vivo. Extracellular amastigotes (EAs have a stage-specific surface antigen called Ssp-4, a GPI-anchored glycoprotein that is secreted by the parasites. By immunoprecipitation with the Ssp-4-specific monoclonal antibodies (mAb 2C2 and 1D9, we isolated the glycoprotein from EAs. By mass spectrometry, we identified the core protein of Ssp-4 and evaluated mRNA expression and the presence of Ssp-4 carbohydrate epitopes recognized by mAb1D9. We demonstrated that the carbohydrate epitope recognized by mAb1D9 could promote host cell invasion by EAs. Although infectious EAs express lower amounts of Ssp-4 compared with less-infectious EAs (at the mRNA and protein levels, it is the glycosylation of Ssp-4 (identified by mAb1D9 staining only in infectious strains and recognized by galectin-3 on host cells that is the determinant of EA invasion of host cells. Furthermore, Ssp-4 is secreted by EAs, either free or associated with parasite vesicles, and can participate in host-cell interactions. The results presented here describe the possible role of a carbohydrate moiety of T. cruzi surface glycoproteins in host cell invasion by EA forms, highlighting the potential of these moieties as therapeutic and vaccine targets for the treatment of Chagas’ disease.

  5. A Carbohydrate Moiety of Secreted Stage-Specific Glycoprotein 4 Participates in Host Cell Invasion by Trypanosoma cruzi Extracellular Amastigotes

    Science.gov (United States)

    Florentino, Pilar T. V.; Real, Fernando; Orikaza, Cristina M.; da Cunha, Julia P. C.; Vitorino, Francisca N. L.; Cordero, Esteban M.; Sobreira, Tiago J. P.; Mortara, Renato A.

    2018-01-01

    Trypanosoma cruzi is the etiologic agent of Chagas’ disease. It is known that amastigotes derived from trypomastigotes in the extracellular milieu are infective in vitro and in vivo. Extracellular amastigotes (EAs) have a stage-specific surface antigen called Ssp-4, a GPI-anchored glycoprotein that is secreted by the parasites. By immunoprecipitation with the Ssp-4-specific monoclonal antibodies (mAb) 2C2 and 1D9, we isolated the glycoprotein from EAs. By mass spectrometry, we identified the core protein of Ssp-4 and evaluated mRNA expression and the presence of Ssp-4 carbohydrate epitopes recognized by mAb1D9. We demonstrated that the carbohydrate epitope recognized by mAb1D9 could promote host cell invasion by EAs. Although infectious EAs express lower amounts of Ssp-4 compared with less-infectious EAs (at the mRNA and protein levels), it is the glycosylation of Ssp-4 (identified by mAb1D9 staining only in infectious strains and recognized by galectin-3 on host cells) that is the determinant of EA invasion of host cells. Furthermore, Ssp-4 is secreted by EAs, either free or associated with parasite vesicles, and can participate in host-cell interactions. The results presented here describe the possible role of a carbohydrate moiety of T. cruzi surface glycoproteins in host cell invasion by EA forms, highlighting the potential of these moieties as therapeutic and vaccine targets for the treatment of Chagas’ disease. PMID:29692765

  6. A MAM7 peptide-based inhibitor of Staphylococcus aureus adhesion does not interfere with in vitro host cell function.

    Directory of Open Access Journals (Sweden)

    Catherine Alice Hawley

    Full Text Available Adhesion inhibitors that block the attachment of pathogens to host tissues may be used synergistically with or as an alternative to antibiotics. The wide-spread bacterial adhesin Multivalent Adhesion Molecule (MAM 7 has recently emerged as a candidate molecule for a broad-spectrum adhesion inhibitor which may be used to prevent bacterial colonization of wounds. Here we have tested if the antibacterial properties of a MAM-based inhibitor could be used to competitively inhibit adhesion of methicillin-resistant Staphylococcus aureus (MRSA to host cells. Additionally, we analyzed its effect on host cellular functions linked to the host receptor fibronectin, such as migration, adhesion and matrix formation in vitro, to evaluate potential side effects prior to advancing our studies to in vivo infection models. As controls, we used inhibitors based on well-characterized bacterial adhesin-derived peptides from F1 and FnBPA, which are known to affect host cellular functions. Inhibitors based on F1 or FnBPA blocked MRSA attachment but at the same time abrogated important cellular functions. A MAM7-based inhibitor did not interfere with host cell function while showing good efficacy against MRSA adhesion in a tissue culture model. These observations provide a possible candidate for a bacterial adhesion inhibitor that does not cause adverse effects on host cells while preventing bacterial infection.

  7. Putative alternative polyadenylation (APA) events in the early interaction of Salmonella enterica Typhimurium and human host cells.

    Science.gov (United States)

    Afonso-Grunz, Fabian

    2015-12-01

    The immune response of epithelial cells upon infection is mediated by changing activity levels of a variety of proteins along with changes in mRNA, and also ncRNA abundance. Alternative polyadenylation (APA) represents a mechanism that diversifies gene expression similar to alternative splicing. T-cell activation, neuronal activity, development and several human diseases including viral infections involve APA, but at present it remains unclear if this mechanism is also implicated in the response to bacterial infections. Our recently published study of interacting Salmonella enterica Typhimurium and human host cells includes genome-wide expression profiles of human epithelial cells prior and subsequent to infection with the invasive pathogen. The generated dataset (GEO accession number: GSE61730) covers several points of time post infection, and one of these interaction stages was additionally profiled with MACE-based dual 3'Seq, which allows for identification of polyadenylation (PA) sites. The present study features the polyadenylation landscape in early interacting cells based on this data, and provides a comparison of the identified PA sites with those of a corresponding 3P-Seq dataset of non-interacting cells. Differential PA site usage of FTL , PRDX1 and VAPA results in transcription of mRNA isoforms with distinct sets of miRNA and protein binding sites that influence processing, localization, stability, and translation of the respective mRNA. APA of these candidate genes consequently harbors the potential to modulate the host cell response to bacterial infection.

  8. Putative alternative polyadenylation (APA events in the early interaction of Salmonella enterica Typhimurium and human host cells

    Directory of Open Access Journals (Sweden)

    Fabian Afonso-Grunz

    2015-12-01

    Full Text Available The immune response of epithelial cells upon infection is mediated by changing activity levels of a variety of proteins along with changes in mRNA, and also ncRNA abundance. Alternative polyadenylation (APA represents a mechanism that diversifies gene expression similar to alternative splicing. T-cell activation, neuronal activity, development and several human diseases including viral infections involve APA, but at present it remains unclear if this mechanism is also implicated in the response to bacterial infections. Our recently published study of interacting Salmonella enterica Typhimurium and human host cells includes genome-wide expression profiles of human epithelial cells prior and subsequent to infection with the invasive pathogen. The generated dataset (GEO accession number: GSE61730 covers several points of time post infection, and one of these interaction stages was additionally profiled with MACE-based dual 3'Seq, which allows for identification of polyadenylation (PA sites. The present study features the polyadenylation landscape in early interacting cells based on this data, and provides a comparison of the identified PA sites with those of a corresponding 3P-Seq dataset of non-interacting cells. Differential PA site usage of FTL, PRDX1 and VAPA results in transcription of mRNA isoforms with distinct sets of miRNA and protein binding sites that influence processing, localization, stability, and translation of the respective mRNA. APA of these candidate genes consequently harbors the potential to modulate the host cell response to bacterial infection.

  9. The effect of Bulgarian propolis against Trypanosoma cruzi and during its interaction with host cells

    Directory of Open Access Journals (Sweden)

    Andréia Pires Dantas

    2006-03-01

    Full Text Available Propolis has shown activity against pathogenic microorganisms that cause diseases in humans and animals. The ethanol (Et-Blg and acetone (Ket-Blg extracts from a Bulgarian propolis, with known chemical compositions, presented similar activity against tissue culture-derived amastigotes. The treatment of Trypanosoma cruzi-infected skeletal muscle cells with Et-Blg led to a decrease of infection and of the intracellular proliferation of amastigotes, while damage to the host cell was observed only at concentration 12.5 times higher than those affecting the parasite. Ultrastructural analysis of the effect of both extracts in epimastigotes revealed that the main targets were the mitochondrion and reservosomes. Et-Blg also affected the mitochondrion-kinetoplast complex in trypomastigotes, offering a potential target for chemotherapeutic agents.

  10. Molecular model of a type III secretion system needle: Implications for host-cell sensing.

    Science.gov (United States)

    Deane, Janet E; Roversi, Pietro; Cordes, Frank S; Johnson, Steven; Kenjale, Roma; Daniell, Sarah; Booy, Frank; Picking, William D; Picking, Wendy L; Blocker, Ariel J; Lea, Susan M

    2006-08-15

    Type III secretion systems are essential virulence determinants for many Gram-negative bacterial pathogens. The type III secretion system consists of cytoplasmic, transmembrane, and extracellular domains. The extracellular domain is a hollow needle protruding above the bacterial surface and is held within a basal body that traverses both bacterial membranes. Effector proteins are translocated, via this external needle, directly into host cells, where they subvert normal cell functions to aid infection. Physical contact with host cells initiates secretion and leads to formation of a pore, thought to be contiguous with the needle channel, in the host-cell membrane. Here, we report the crystal structure of the Shigella flexneri needle subunit MxiH and a complete model for the needle assembly built into our three-dimensional EM reconstruction. The model, combined with mutagenesis data, reveals that signaling of host-cell contact is relayed through the needle via intersubunit contacts and suggests a mode of binding for a tip complex.

  11. ARF6, PI3-kinase and host cell actin cytoskeleton in Toxoplasma gondii cell invasion

    International Nuclear Information System (INIS)

    Vieira da Silva, Claudio; Alves da Silva, Erika; Costa Cruz, Mario; Chavrier, Philippe; Arruda Mortara, Renato

    2009-01-01

    Toxoplasma gondii infects a variety of different cell types in a range of different hosts. Host cell invasion by T. gondii occurs by active penetration of the host cell, a process previously described as independent of host actin polymerization. Also, the parasitophorous vacuole has been shown to resist fusion with endocytic and exocytic pathways of the host cell. ADP-ribosylation factor-6 (ARF6) belongs to the ARF family of small GTP-binding proteins. ARF6 regulates membrane trafficking and actin cytoskeleton rearrangements at the plasma membrane. Here, we have observed that ARF6 is recruited to the parasitophorous vacuole of tachyzoites of T. gondii RH strain and it also plays an important role in the parasite cell invasion with activation of PI3-kinase and recruitment of PIP 2 and PIP 3 to the parasitophorous vacuole of invading parasites. Moreover, it was verified that maintenance of host cell actin cytoskeleton integrity is important to parasite invasion.

  12. Trichomonas vaginalis and Tritrichomonas foetus: interaction with fibroblasts and muscle cells - new insights into parasite-mediated host cell cytotoxicity

    Directory of Open Access Journals (Sweden)

    Ricardo Chaves Vilela

    2012-09-01

    Full Text Available Trichomonas vaginalis and Tritrichomonas foetus are parasitic, flagellated protists that inhabit the urogenital tract of humans and bovines, respectively. T. vaginalis causes the most prevalent non-viral sexually transmitted disease worldwide and has been associated with an increased risk for human immunodeficiency virus-1 infection in humans. Infections by T. foetus cause significant losses to the beef industry worldwide due to infertility and spontaneous abortion in cows. Several studies have shown a close association between trichomonads and the epithelium of the urogenital tract. However, little is known concerning the interaction of trichomonads with cells from deeper tissues, such as fibroblasts and muscle cells. Published parasite-host cell interaction studies have reported contradictory results regarding the ability of T. foetus and T. vaginalis to interact with and damage cells of different tissues. In this study, parasite-host cell interactions were examined by culturing primary human fibroblasts obtained from abdominal biopsies performed during plastic surgeries with trichomonads. In addition, mouse 3T3 fibroblasts, primary chick embryo myogenic cells and L6 muscle cells were also used as models of target cells. The parasite-host cell cultures were processed for scanning and transmission electron microscopy and were tested for cell viability and cell death. JC-1 staining, which measures mitochondrial membrane potential, was used to determine whether the parasites induced target cell damage. Terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling staining was used as an indicator of chromatin damage. The colorimetric crystal violet assay was performed to ana-lyse the cytotoxicity induced by the parasite. The results showed that T. foetus and T. vaginalis adhered to and were cytotoxic to both fibroblasts and muscle cells, indicating that trichomonas infection of the connective and muscle tissues is likely to occur; such

  13. The chlamydial periplasmic stress response serine protease cHtrA is secreted into host cell cytosol

    Directory of Open Access Journals (Sweden)

    Flores Rhonda

    2011-04-01

    Full Text Available Abstract Background The periplasmic High Temperature Requirement protein A (HtrA plays important roles in bacterial protein folding and stress responses. However, the role of chlamydial HtrA (cHtrA in chlamydial pathogenesis is not clear. Results The cHtrA was detected both inside and outside the chlamydial inclusions. The detection was specific since both polyclonal and monoclonal anti-cHtrA antibodies revealed similar intracellular labeling patterns that were only removed by absorption with cHtrA but not control fusion proteins. In a Western blot assay, the anti-cHtrA antibodies detected the endogenous cHtrA in Chlamydia-infected cells without cross-reacting with any other chlamydial or host cell antigens. Fractionation of the infected cells revealed cHtrA in the host cell cytosol fraction. The periplasmic cHtrA protein appeared to be actively secreted into host cell cytosol since no other chlamydial periplasmic proteins were detected in the host cell cytoplasm. Most chlamydial species secreted cHtrA into host cell cytosol and the secretion was not inhibitable by a type III secretion inhibitor. Conclusion Since it is hypothesized that chlamydial organisms possess a proteolysis strategy to manipulate host cell signaling pathways, secretion of the serine protease cHtrA into host cell cytosol suggests that the periplasmic cHtrA may also play an important role in chlamydial interactions with host cells.

  14. Insights into Host Cell Modulation and Induction of New Cells by the Corn Smut Ustilago maydis

    Directory of Open Access Journals (Sweden)

    Amey Redkar

    2017-05-01

    Full Text Available Many filamentous fungal pathogens induce drastic modulation of host cells causing abnormal infectious structures such as galls, or tumors that arise as a result of re-programming in the original developmental cell fate of a colonized host cell. Developmental consequences occur predominantly with biotrophic phytopathogens. This suggests that these host structures result as an outcome of efficient defense suppression and intimate fungal–host interaction to suit the pathogen’s needs for completion of its infection cycle. This mini-review mainly summarizes host cell re-programming that occurs in the Ustilago maydis – maize interaction, in which the pathogen deploys cell-type specific effector proteins with varying activities. The fungus senses the physiological status and identity of colonized host cells and re-directs the endogenous developmental program of its host. The disturbance of host cell physiology and cell fate leads to novel cell shapes, increased cell size, and/or the number of host cells. We particularly highlight the strategies of U. maydis to induce physiologically varied host organs to form the characteristic tumors in both vegetative and floral parts of maize.

  15. Staphylococcus aureus produces membrane-derived vesicles that induce host cell death.

    Directory of Open Access Journals (Sweden)

    Mamata Gurung

    Full Text Available Gram-negative bacteria produce outer membrane vesicles that play a role in the delivery of virulence factors to host cells. However, little is known about the membrane-derived vesicles (MVs produced by gram-positive bacteria. The present study examined the production of MVs from Staphylococcus aureus and investigated the delivery of MVs to host cells and subsequent cytotoxicity. Four S. aureus strains tested, two type strains and two clinical isolates, produced spherical nanovesicles during in vitro culture. MVs were also produced during in vivo infection of a clinical S. aureus isolate in a mouse pneumonia model. Proteomic analysis showed that 143 different proteins were identified in the S. aureus-derived MVs. S. aureus MVs were interacted with the plasma membrane of host cells via a cholesterol-rich membrane microdomain and then delivered their component protein A to host cells within 30 min. Intact S. aureus MVs induced apoptosis of HEp-2 cells in a dose-dependent manner, whereas lysed MVs neither delivered their component into the cytosol of host cells nor induced cytotoxicity. In conclusion, this study is the first report that S. aureus MVs are an important vehicle for delivery of bacterial effector molecules to host cells.

  16. Easy labeling of proliferative phase and sporogonic phase of microsporidia Nosema bombycis in host cells.

    Directory of Open Access Journals (Sweden)

    Jie Chen

    Full Text Available Microsporidia are eukaryotic, unicellular parasites that have been studied for more than 150 years. These organisms are extraordinary in their ability to invade a wide range of hosts including vertebrates and invertebrates, such as human and commercially important animals. A lack of appropriate labeling methods has limited the research of the cell cycle and protein locations in intracellular stages. In this report, an easy fluorescent labeling method has been developed to mark the proliferative and sporogonic phases of microsporidia Nosema bombycis in host cells. Based on the presence of chitin, Calcofluor White M2R was used to label the sporogonic phase, while β-tubulin antibody coupled with fluorescence secondary antibody were used to label the proliferative phase by immunofluorescence. This method is simple, efficient and can be used on both infected cells and tissue slices, providing a great potential application in microsporidia research.

  17. Orientia tsutsugamushi ankyrin repeat-containing protein family members are Type 1 secretion system substrates that traffic to the host cell endoplasmic reticulum.

    Science.gov (United States)

    VieBrock, Lauren; Evans, Sean M; Beyer, Andrea R; Larson, Charles L; Beare, Paul A; Ge, Hong; Singh, Smita; Rodino, Kyle G; Heinzen, Robert A; Richards, Allen L; Carlyon, Jason A

    2014-01-01

    Scrub typhus is an understudied, potentially fatal infection that threatens one billion persons in the Asia-Pacific region. How the causative obligate intracellular bacterium, Orientia tsutsugamushi, facilitates its intracellular survival and pathogenesis is poorly understood. Many intracellular bacterial pathogens utilize the Type 1 (T1SS) or Type 4 secretion system (T4SS) to translocate ankyrin repeat-containing proteins (Anks) that traffic to distinct subcellular locations and modulate host cell processes. The O. tsutsugamushi genome encodes one of the largest known bacterial Ank repertoires plus T1SS and T4SS components. Whether these potential virulence factors are expressed during infection, how the Anks are potentially secreted, and to where they localize in the host cell are not known. We determined that O. tsutsugamushi transcriptionally expresses 20 unique ank genes as well as genes for both T1SS and T4SS during infection of mammalian host cells. Examination of the Anks' C-termini revealed that the majority of them resemble T1SS substrates. Escherichia coli expressing a functional T1SS was able to secrete chimeric hemolysin proteins bearing the C-termini of 19 of 20 O. tsutsugamushi Anks in an HlyBD-dependent manner. Thus, O. tsutsugamushi Anks C-termini are T1SS-compatible. Conversely, Coxiella burnetii could not secrete heterologously expressed Anks in a T4SS-dependent manner. Analysis of the subcellular distribution patterns of 20 ectopically expressed Anks revealed that, while 6 remained cytosolic or trafficked to the nucleus, 14 localized to, and in some cases, altered the morphology of the endoplasmic reticulum. This study identifies O. tsutsugamushi Anks as T1SS substrates and indicates that many display a tropism for the host cell secretory pathway.

  18. Staphylococcus aureus ?-Toxin-Dependent Induction of Host Cell Death by Membrane-Derived Vesicles

    OpenAIRE

    Thay, Bernard; Wai, Sun Nyunt; Oscarsson, Jan

    2013-01-01

    Staphylococcus aureus causes a wide spectrum of infections in humans, ranging from superficial cutaneous infections, infections in the circum-oral region, to life-threatening bacteremia. It was recently demonstrated that Gram-positive organisms such as S. aureus liberate membrane-derived vesicles (MVs), which analogously to outer membrane vesicles (OMVs) of Gram-negative bacteria can play a role in delivering virulence factors to host cells. In the present study we have shown that cholesterol...

  19. Effects of host cell sterol composition upon internalization of Yersinia pseudotuberculosis and clustered β1 integrin.

    Science.gov (United States)

    Kim, JiHyun; Fukuto, Hana S; Brown, Deborah A; Bliska, James B; London, Erwin

    2018-01-26

    Yersinia pseudotuberculosis is a foodborne pathogenic bacterium that causes acute gastrointestinal illness, but its mechanisms of infection are incompletely described. We examined how host cell sterol composition affected Y. pseudotuberculosis uptake. To do this, we depleted or substituted cholesterol in human MDA-MB-231 epithelial cells with various alternative sterols. Decreasing host cell cholesterol significantly reduced pathogen internalization. When host cell cholesterol was substituted with various sterols, only desmosterol and 7-dehydrocholesterol supported internalization. This specificity was not due to sterol dependence of bacterial attachment to host cells, which was similar with all sterols studied. Because a key step in Y. pseudotuberculosis internalization is interaction of the bacterial adhesins invasin and YadA with host cell β1 integrin, we compared the sterol dependence of wildtype Y. pseudotuberculosis internalization with that of Δ inv , Δ yadA , and Δ inv Δ yadA mutant strains. YadA deletion decreased bacterial adherence to host cells, whereas invasin deletion had no effect. Nevertheless, host cell sterol substitution had a similar effect on internalization of these bacterial deletion strains as on the wildtype bacteria. The Δ inv Δ yadA double mutant adhered least to cells and so was not significantly internalized. The sterol structure dependence of Y. pseudotuberculosis internalization differed from that of endocytosis, as monitored using antibody-clustered β1 integrin and previous studies on other proteins, which had a more permissive sterol dependence. This study suggests that agents could be designed to interfere with internalization of Yersinia without disturbing endocytosis. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Complexities in human herpesvirus-6A and -6B binding to host cells

    DEFF Research Database (Denmark)

    Pedersen, Simon Metz; Höllsberg, Per

    2006-01-01

    Human herpesvirus-6A and -6B uses the cellular receptor CD46 for fusion and infection of the host cell. The viral glycoprotein complex gH-gL from HHV-6A binds to the short consensus repeat 2 and 3 in CD46. Although all the major isoforms of CD46 bind the virus, certain isoforms may have higher...

  1. Differences in infectivity between endosymbiotic Chlorella variabilis cultivated outside host Paramecium bursaria for 50 years and those immediately isolated from host cells after one year of reendosymbiosis

    Directory of Open Access Journals (Sweden)

    Y. Kodama

    2016-01-01

    Full Text Available Chlorella variabilis strain NC64A is an intracellular photobiont of the ciliate Paramecium bursaria. NC64A was isolated from P. bursaria nearly 50 years ago and was thereafter cultivated outside the host. This study was undertaken to detect changes in its infectivity to P. bursaria and its auxotrophy for growth outside the host induced during long-term cultivation. NC64A can grow in Modified Bold's Basal Medium but not in C medium, whereas another symbiotic Chlorella variabilis strain, 1N, that was recently isolated from the host grew in C medium but not in Modified Bold's Basal Medium. With regards infectivity, NC64A in the logarithmic phase of growth showed low infectivity to alga-removed P. bursaria cells, whereas those in the early stationary phase showed high infectivity of about 30%. Those in the decay phase of growth showed no infectivity. Results show that NC64A has infectivity, but the infection rate depends on their culture age in the growth curve. Furthermore, NC64A that had been re-infected to P. bursaria for more than one year and isolated from the host showed a nearly 100% infection rate, which indicates that NC64A can recover its infectivity by re-infection to P. bursaria.

  2. Porphyromonas gingivalis as a Model Organism for Assessing Interaction of Anaerobic Bacteria with Host Cells.

    Science.gov (United States)

    Wunsch, Christopher M; Lewis, Janina P

    2015-12-17

    Anaerobic bacteria far outnumber aerobes in many human niches such as the gut, mouth, and vagina. Furthermore, anaerobic infections are common and frequently of indigenous origin. The ability of some anaerobic pathogens to invade human cells gives them adaptive measures to escape innate immunity as well as to modulate host cell behavior. However, ensuring that the anaerobic bacteria are live during experimental investigation of the events may pose challenges. Porphyromonas gingivalis, a Gram-negative anaerobe, is capable of invading a variety of eukaryotic non-phagocytic cells. This article outlines how to successfully culture and assess the ability of P. gingivalis to invade human umbilical vein endothelial cells (HUVECs). Two protocols were developed: one to measure bacteria that can successfully invade and survive within the host, and the other to visualize bacteria interacting with host cells. These techniques necessitate the use of an anaerobic chamber to supply P. gingivalis with an anaerobic environment for optimal growth. The first protocol is based on the antibiotic protection assay, which is largely used to study the invasion of host cells by bacteria. However, the antibiotic protection assay is limited; only intracellular bacteria that are culturable following antibiotic treatment and host cell lysis are measured. To assess all bacteria interacting with host cells, both live and dead, we developed a protocol that uses fluorescent microscopy to examine host-pathogen interaction. Bacteria are fluorescently labeled with 2',7'-Bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM) and used to infect eukaryotic cells under anaerobic conditions. Following fixing with paraformaldehyde and permeabilization with 0.2% Triton X-100, host cells are labeled with TRITC phalloidin and DAPI to label the cell cytoskeleton and nucleus, respectively. Multiple images taken at different focal points (Z-stack) are obtained for temporal

  3. Perturbation of host-cell membrane is a primary mechanism of HIV cytopathology.

    Science.gov (United States)

    Cloyd, M W; Lynn, W S

    1991-04-01

    Cytopathic viruses injure cells by a number of different mechanisms. The mechanism by which HIV-1 injures T cells was studied by temporally examining host-cell macromolecular syntheses, stages of the cell cycle, and membrane permeability following acute infection. T cells cytopathically infected at an m.o.i. of 1-5 grew normally for 24-72 hr, depending on the cell line, followed by the first manifestation of cell injury, slowing of cell division. At that time significant amounts of unintegrated HIV DNA and p24 core protein became detectable, and acridine orange flow cytometric cell cycle studies demonstrated the presence of fewer cells in the G2/M stage of the cell cycle. There was no change in the frequency of cells in the S-stage, and metabolic pulsing with radioactive precursors demonstrated that host-cell DNA, RNA, and protein syntheses were normal at that time and normal up to the time cells started to die (approximately 24 hr later), when all three decreased. Cellular lipid synthesis, however, was perturbed when cell multiplication slowed, with phospholipid synthesis reduced and neutral lipid synthesis enhanced. Permeability of the host-cell membrane to small molecules, such as Ca2+ and sucrose, was slightly enhanced early postinfection, and by the time of slowing of cell division, host membrane permeability was greatly increased to both Ca2+ and sucrose (Stokes radius 5.2 A) but not to inulin (Stokes radium 20 A). These changes in host-cell membrane permeability and phospholipid synthesis were not observed in acutely infected H9 cells, which are not susceptible to HIV cytopathology. Thus, HIV-1 appeared to predominantly injure T cells by perturbing host-cell membrane permeability and lipid synthesis, which is similar to the cytopathic mechanisms of paramyxoviruses.

  4. Cellular Aspects of Shigella Pathogenesis: Focus on the Manipulation of Host Cell Processes.

    Science.gov (United States)

    Killackey, Samuel A; Sorbara, Matthew T; Girardin, Stephen E

    2016-01-01

    Shigella is a Gram-negative bacterium that is responsible for shigellosis. Over the years, the study of Shigella has provided a greater understanding of how the host responds to bacterial infection, and how bacteria have evolved to effectively counter the host defenses. In this review, we provide an update on some of the most recent advances in our understanding of pivotal processes associated with Shigella infection, including the invasion into host cells, the metabolic changes that occur within the bacterium and the infected cell, cell-to-cell spread mechanisms, autophagy and membrane trafficking, inflammatory signaling and cell death. This recent progress sheds a new light into the mechanisms underlying Shigella pathogenesis, and also more generally provides deeper understanding of the complex interplay between host cells and bacterial pathogens in general.

  5. Host cells and methods for production of isobutanol

    Science.gov (United States)

    Anthony, Larry Cameron; He, Hongxian; Huang, Lixuan Lisa; Okeefe, Daniel P.; Kruckeberg, Arthur Leo; Li, Yougen; Maggio-Hall, Lori; McElvain, Jessica; Nelson, Mark J.; Patnaik, Ranjan; Rothman, Steven Cary

    2017-10-17

    Provided herein are recombinant yeast host cells and methods for their use for production of isobutanol. Yeast host cells provided comprise an isobutanol biosynthetic pathway and at least one of reduced or eliminated aldehyde dehydrogenase activity, reduced or eliminated acetolactate reductase activity; or a heterologous polynucleotide encoding a polypeptide having ketol-acid reductoisomerase activity.

  6. Methods for production of proteins in host cells

    Science.gov (United States)

    Donnelly, Mark; Joachimiak, Andrzej

    2004-01-13

    The present invention provides methods for the production of proteins, particularly toxic proteins, in host cells. The invention provides methods which use a fusion protein comprising a chaperonin binding domain in host cells induced or regulated to have increased levels of chaperonin which binds the chaperonin binding domain.

  7. Peptide Nucleic Acid Knockdown and Intra-host Cell Complementation of Ehrlichia Type IV Secretion System Effector

    Directory of Open Access Journals (Sweden)

    Pratibha Sharma

    2017-06-01

    Full Text Available Survival of Ehrlichia chaffeensis depends on obligatory intracellular infection. One of the barriers to E. chaffeensis research progress has been the inability, using conventional techniques, to generate knock-out mutants for genes essential for intracellular infection. This study examined the use of Peptide Nucleic Acids (PNAs technology to interrupt type IV secretion system (T4SS effector protein expression in E. chaffeensis followed by intracellular complementation of the effector to determine its requirement for infection. Successful E. chaffeensis infection depends on the E. chaffeensis-specific T4SS protein effector, ehrlichial translocated factor-1 (Etf-1, which induces Rab5-regulated autophagy to provide host cytosolic nutrients required for E. chaffeensis proliferation. Etf-1 is also imported by host cell mitochondria where it inhibits host cell apoptosis to prolong its infection. We designed a PNA specific to Etf-1 and showed that the PNA bound to the target region of single-stranded Etf-1 RNA using a competitive binding assay. Electroporation of E. chaffeensis with this PNA significantly reduced Etf-1 mRNA and protein, and the bacteria's ability to induce host cell autophagy and infect host cells. Etf-1 PNA-mediated inhibition of ehrlichial Etf-1 expression and E. chaffeensis infection could be intracellularly trans-complemented by ectopic expression of Etf-1-GFP in host cells. These data affirmed the critical role of bacterial T4SS effector in host cell autophagy and E. chaffeensis infection, and demonstrated the use of PNA to analyze the gene functions of obligate intracellular bacteria.

  8. Infection potential of vegetative incompatible Ganoderma boninense ...

    African Journals Online (AJOL)

    LiP), manganese peroxidase (MnP) and laccase to degrade the lignin composition in plant cell walls. The present study aimed to evaluate the infection potential of vegetative incompatible isolates of G. boninense in causing Basal Stem Rot ...

  9. Adjustment of host cells for accommodation of symbiotic bacteria: vacuole defunctionalization, HOPS suppression, and TIP1g retargeting in Medicago

    NARCIS (Netherlands)

    Gavrin, A.Y.; Kaiser, B.N.; Geiger, D.; Tyerman, S.D.; Wen, Z.; Bisseling, T.; Fedorova, E.E.

    2014-01-01

    In legume–rhizobia symbioses, the bacteria in infected cells are enclosed in a plant membrane, forming organelle-like compartments called symbiosomes. Symbiosomes remain as individual units and avoid fusion with lytic vacuoles of host cells. We observed changes in the vacuole volume of infected

  10. Host cell reactivation and UV-enhanced reactivation in synchronized mammalian cells

    International Nuclear Information System (INIS)

    Lytle, C.D.; Schmidt, B.J.

    1981-01-01

    Does host cell reactivation (HCR) or UV-enhanced reactivation (UVER) of UV-irradiated Herpes simplex virus (UV-HSV) vary during the host mammalian cell cycle. The answer could be useful for interpreting UVER and or the two-component nature of the UV-HSV survival curve. Procedures were developed for infection of mitotically-synchronized CV-l monkey kidney cells. All virus survival curves determined at different cell cycle stages had two components with similar D 0 's and intercepts of the second components. Thus, no single stage of the host cell cycle was responsible for the second component of the virus survival curve. When the cells were UV-irradiated immediately prior to infection, enhanced survival of UV-HSV occurred for cell irradiation and virus infection initiated during late G 1 early S phase or late S early G 2 phase but not during early G 1 phase. For infection delayed by 24 h after cell irradiation, UVER was found at all investigated times. These results indicate that: (1) HCR is similar at all stages of the host cell cycle: and (2) the ''induction'' of UVER is not as rapid for cell-irradiation in early G 1 phase. This latter observation may be one reason why normal, contact-inhibited cells do not express UVER as rapidly as faster growing, less contact-inhibited cells. (author)

  11. Dual analysis of the murine cytomegalovirus and host cell transcriptomes reveal new aspects of the virus-host cell interface.

    Directory of Open Access Journals (Sweden)

    Vanda Juranic Lisnic

    Full Text Available Major gaps in our knowledge of pathogen genes and how these gene products interact with host gene products to cause disease represent a major obstacle to progress in vaccine and antiviral drug development for the herpesviruses. To begin to bridge these gaps, we conducted a dual analysis of Murine Cytomegalovirus (MCMV and host cell transcriptomes during lytic infection. We analyzed the MCMV transcriptome during lytic infection using both classical cDNA cloning and sequencing of viral transcripts and next generation sequencing of transcripts (RNA-Seq. We also investigated the host transcriptome using RNA-Seq combined with differential gene expression analysis, biological pathway analysis, and gene ontology analysis. We identify numerous novel spliced and unspliced transcripts of MCMV. Unexpectedly, the most abundantly transcribed viral genes are of unknown function. We found that the most abundant viral transcript, recently identified as a noncoding RNA regulating cellular microRNAs, also codes for a novel protein. To our knowledge, this is the first viral transcript that functions both as a noncoding RNA and an mRNA. We also report that lytic infection elicits a profound cellular response in fibroblasts. Highly upregulated and induced host genes included those involved in inflammation and immunity, but also many unexpected transcription factors and host genes related to development and differentiation. Many top downregulated and repressed genes are associated with functions whose roles in infection are obscure, including host long intergenic noncoding RNAs, antisense RNAs or small nucleolar RNAs. Correspondingly, many differentially expressed genes cluster in biological pathways that may shed new light on cytomegalovirus pathogenesis. Together, these findings provide new insights into the molecular warfare at the virus-host interface and suggest new areas of research to advance the understanding and treatment of cytomegalovirus

  12. Host cells and methods for producing isoprenyl alkanoates

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Taek Soon; Fortman, Jeffrey L.; Keasling, Jay D.

    2015-12-01

    The invention provides for a method of producing an isoprenyl alkanoate in a genetically modified host cell. In one embodiment, the method comprises culturing a genetically modified host cell which expresses an enzyme capable of catalyzing the esterification of an isoprenol and a straight-chain fatty acid, such as an alcohol acetyltransferase (AAT), wax ester synthase/diacylglycerol acyltransferase (WS/DGAT) or lipase, under a suitable condition so that the isoprenyl alkanoate is produced.

  13. Epigenetic silencing of host cell defense genes enhances intracellular survival of the rickettsial pathogen Anaplasma phagocytophilum.

    Directory of Open Access Journals (Sweden)

    Jose C Garcia-Garcia

    2009-06-01

    Full Text Available Intracellular bacteria have evolved mechanisms that promote survival within hostile host environments, often resulting in functional dysregulation and disease. Using the Anaplasma phagocytophilum-infected granulocyte model, we establish a link between host chromatin modifications, defense gene transcription and intracellular bacterial infection. Infection of THP-1 cells with A. phagocytophilum led to silencing of host defense gene expression. Histone deacetylase 1 (HDAC1 expression, activity and binding to the defense gene promoters significantly increased during infection, which resulted in decreased histone H3 acetylation in infected cells. HDAC1 overexpression enhanced infection, whereas pharmacologic and siRNA HDAC1 inhibition significantly decreased bacterial load. HDAC2 does not seem to be involved, since HDAC2 silencing by siRNA had no effect on A. phagocytophilum intracellular propagation. These data indicate that HDAC up-regulation and epigenetic silencing of host cell defense genes is required for A. phagocytophilum infection. Bacterial epigenetic regulation of host cell gene transcription could be a general mechanism that enhances intracellular pathogen survival while altering cell function and promoting disease.

  14. Knowledge to Predict Pathogens: Legionella pneumophila Lifecycle Critical Review Part I Uptake into Host Cells

    Directory of Open Access Journals (Sweden)

    Alexis L. Mraz

    2018-01-01

    Full Text Available Legionella pneumophila (L. pneumophila is an infectious disease agent of increasing concern due to its ability to cause Legionnaires’ Disease, a severe community pneumonia, and the difficulty in controlling it within water systems. L. pneumophila thrives within the biofilm of premise plumbing systems, utilizing protozoan hosts for protection from disinfectants and other environmental stressors. While there is a great deal of information regarding how L. pneumophila interacts with protozoa and human macrophages (host for human infection, the ability to use this data in a model to attempt to predict a concentration of L. pneumophila in a water system is not known. The lifecycle of L. pneumophila within host cells involves three processes: uptake, growth, and egression from the host cell. The complexity of these three processes would risk conflation of the concepts; therefore, this review details the available information regarding how L. pneumophila invades host cells (uptake within the context of data needed to model this process, while a second review will focus on growth and egression. The overall intent of both reviews is to detail how the steps in L. pneumophila’s lifecycle in drinking water systems affect human infectivity, as opposed to detailing just its growth and persistence in drinking water systems.

  15. RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.

    Directory of Open Access Journals (Sweden)

    Julia F Pielage

    2008-03-01

    Full Text Available Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of approximately 80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa-induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections.

  16. Host cell subversion by Toxoplasma GRA16, an exported dense granule protein that targets the host cell nucleus and alters gene expression.

    Science.gov (United States)

    Bougdour, Alexandre; Durandau, Eric; Brenier-Pinchart, Marie-Pierre; Ortet, Philippe; Barakat, Mohamed; Kieffer, Sylvie; Curt-Varesano, Aurélie; Curt-Bertini, Rose-Laurence; Bastien, Olivier; Coute, Yohann; Pelloux, Hervé; Hakimi, Mohamed-Ali

    2013-04-17

    After invading host cells, Toxoplasma gondii multiplies within a parasitophorous vacuole (PV) that is maintained by parasite proteins secreted from organelles called dense granules. Most dense granule proteins remain within the PV, and few are known to access the host cell cytosol. We identify GRA16 as a dense granule protein that is exported through the PV membrane and reaches the host cell nucleus, where it positively modulates genes involved in cell-cycle progression and the p53 tumor suppressor pathway. GRA16 binds two host enzymes, the deubiquitinase HAUSP and PP2A phosphatase, which exert several functions, including regulation of p53 and the cell cycle. GRA16 alters p53 levels in a HAUSP-dependent manner and induces nuclear translocation of the PP2A holoenzyme. Additionally, certain GRA16-deficient strains exhibit attenuated virulence, indicating the importance of these host alterations in pathogenesis. Therefore, GRA16 represents a potentially emerging subfamily of exported dense granule proteins that modulate host function. Copyright © 2013 Elsevier Inc. All rights reserved.

  17. Multiple factors and processes involved in host cell killing by bacteriophage Mu: characterization and mapping.

    Science.gov (United States)

    Waggoner, B T; Marrs, C F; Howe, M M; Pato, M L

    1984-07-15

    The regions of bacteriophage Mu involved in host cell killing were determined by infection of a lambda-immune host with 12 lambda pMu-transducing phages carrying different amounts of Mu DNA beginning at the left end. Infecting lambda pMu phages containing 5.0 (+/- 0.2) kb or less of the left end of Mu DNA did not kill the lambda-immune host, whereas lambda pMu containing 5.1 kb did kill, thus locating the right end of the kil gene between approximately 5.0 and 5.1 kb. For the Kil+ phages the extent of killing increased as the multiplicity of infection (m.o.i.) increased. In addition, killing was also affected by the presence of at least two other regions of Mu DNA: one, located between 5.1 and 5.8 kb, decreased the extent of killing; the other, located between 6.3 and 7.9 kb, greatly increased host cell killing. Killing was also assayed after lambda pMu infection of a lambda-immune host carrying a mini-Mu deleted for most of the B gene and the middle region of Mu DNA. Complementation of mini-Mu replication by infecting B+ lambda pMu phages resulted in killing of the lambda-immune, mini-Mu-containing host, regardless of the presence or absence of the Mu kil gene. The extent of host cell killing increased as the m.o.i. of the infecting lambda pMu increased, and was further enhanced by both the presence of the kil gene and the region located between 6.3 and 7.9 kb. These distinct processes of kil-mediated killing in the absence of replication and non-kil-mediated killing in the presence of replication were also observed after induction of replication-deficient and kil mutant prophages, respectively.

  18. Nanomimics of host cell membranes block invasion and expose invasive malaria parasites.

    Science.gov (United States)

    Najer, Adrian; Wu, Dalin; Bieri, Andrej; Brand, Françoise; Palivan, Cornelia G; Beck, Hans-Peter; Meier, Wolfgang

    2014-12-23

    The fight against most infectious diseases, including malaria, is often hampered by the emergence of drug resistance and lack or limited efficacies of vaccines. Therefore, new drugs, vaccines, or other strategies to control these diseases are needed. Here, we present an innovative nanotechnological strategy in which the nanostructure itself represents the active substance with no necessity to release compounds to attain therapeutic effect and which might act in a drug- and vaccine-like dual function. Invasion of Plasmodium falciparum parasites into red blood cells was selected as a biological model for the initial validation of this approach. Stable nanomimics-polymersomes presenting receptors required for parasite attachment to host cells-were designed to efficiently interrupt the life cycle of the parasite by inhibiting invasion. A simple way to build nanomimics without postformation modifications was established. First, a block copolymer of the receptor with a hydrophobic polymer was synthesized and then mixed with a polymersome-forming block copolymer. The resulting nanomimics bound parasite-derived ligands involved in the initial attachment to host cells and they efficiently blocked reinvasion of malaria parasites after their egress from host cells in vitro. They exhibited efficacies of more than 2 orders of magnitude higher than the soluble form of the receptor, which can be explained by multivalent interactions of several receptors on one nanomimic with multiple ligands on the infective parasite. In the future, our strategy might offer interesting treatment options for severe malaria or a way to modulate the immune response.

  19. Molecular mechanisms of Porphyromonas gingivalis-host cell interaction on periodontal diseases

    Directory of Open Access Journals (Sweden)

    Masaaki Nakayama

    2017-11-01

    Full Text Available Porphyromonas gingivalis (P. gingivalis is a major oral pathogen and associated with periodontal diseases including periodontitis and alveolar bone loss. In this review, we indicate that two virulence factors, which are hemoglobin receptor protein (HbR and cysteine proteases “gingipains”, expressed by P. gingivalis have novel functions on the pathogenicity of P. gingivalis. P. gingivalis produces three types of gingipains and concomitantly several adhesin domains. Among the adhesin domains, hemoglobin receptor protein (HbR, also called HGP15, has the function of induction of interleukin-8 (IL-8 expression in human gingival epithelial cells, indicating the possibility that HbR is associated with P. gingivalis-induced periodontal inflammation. On bacteria-host cells contact, P. gingivalis induces cellular signaling alteration in host cells. Phosphatidylinositol 3-kinase (PI3K and Akt are well known to play a pivotal role in various cellular physiological functions including cell survival and glucose metabolism in mammalian cells. Recently, we demonstrated that gingipains attenuate the activity of PI3K and Akt, which might have a causal influence on periodontal diseases by chronic infection to the host cells from the speculation of molecular analysis. In this review, we discuss new molecular and biological characterization of the virulence factors from P. gingivalis.

  20. Complexities in human herpesvirus-6A and -6B binding to host cells

    International Nuclear Information System (INIS)

    Pedersen, Simon Metz; Hoellsberg, Per

    2006-01-01

    Human herpesvirus-6A and -6B uses the cellular receptor CD46 for fusion and infection of the host cell. The viral glycoprotein complex gH-gL from HHV-6A binds to the short consensus repeat 2 and 3 in CD46. Although all the major isoforms of CD46 bind the virus, certain isoforms may have higher affinity than others for the virus. Within recent years, elucidation of the viral complex has identified additional HHV-6A and -6B specific glycoproteins. Thus, gH-gL associates with a gQ1-gQ2 dimer to form a heterotetrameric complex. In addition, a novel complex consisting of gH-gL-gO has been described that does not bind CD46. Accumulating evidence suggests that an additional HHV-6A and -6B receptor exists. The previous simple picture of HHV-6A/B-host cell contact therefore includes more layers of complexities on both the viral and the host cell side of the interaction

  1. Host cells and methods for producing diacid compounds

    Energy Technology Data Exchange (ETDEWEB)

    Steen, Eric J.; Fortman, Jeffrey L.; Dietrich, Jeffrey A.; Keasling, Jay D.

    2018-04-24

    The present invention provides for a method of producing one or more fatty acid derived dicarboxylic acids in a genetically modified host cell which does not naturally produce the one or more derived fatty acid derived dicarboxylic acids. The invention provides for the biosynthesis of dicarboxylic acid ranging in length from C3 to C26. The host cell can be further modified to increase fatty acid production or export of the desired fatty acid derived compound, and/or decrease fatty acid storage or metabolism.

  2. Potential use of rapamycin in HIV infection

    DEFF Research Database (Denmark)

    Donia, Marco; McCubrey, James A; Bendtzen, Klaus

    2010-01-01

    The strong need for the development of alternative anti-HIV agents is primarily due to the emergence of strain-resistant viruses, the need for sustained adherence to complex treatment regimens and the toxicity of currently used antiviral drugs. This review analyzes proof of concept studies...... indicating that the immunomodulatory drug rapamycin (RAPA) possesses anti-HIV properties both in vitro and in vivo that qualifies it as a potential new anti-HIV drug. It represents a literature review of published studies that evaluated the in vitro and in vivo activity of RAPA in HIV. RAPA represses HIV-1...... replication in vitro through different mechanisms including, but not limited, to down regulation of CCR5. In addition RAPA synergistically enhances the anti-HIV activity of entry inhibitors such as vicriviroc, aplaviroc and enfuvirtide in vitro. RAPA also inhibits HIV-1 infection in human peripheral blood...

  3. Potential use of rapamycin in HIV infection

    DEFF Research Database (Denmark)

    Donia, Marco; McCubrey, James A; Bendtzen, Klaus

    2010-01-01

    The strong need for the development of alternative anti-HIV agents is primarily due to the emergence of strain-resistant viruses, the need for sustained adherence to complex treatment regimens and the toxicity of currently used antiviral drugs. This review analyzes proof of concept studies......, the evidence presented in this review suggests that RAPA may be a useful drug that should be evaluated for the prevention and treatment of HIV-1 infection....... indicating that the immunomodulatory drug rapamycin (RAPA) possesses anti-HIV properties both in vitro and in vivo that qualifies it as a potential new anti-HIV drug. It represents a literature review of published studies that evaluated the in vitro and in vivo activity of RAPA in HIV. RAPA represses HIV-1...

  4. Inhibition of host cell translation elongation by Legionella pneumophila blocks the host cell unfolded protein response.

    Science.gov (United States)

    Hempstead, Andrew D; Isberg, Ralph R

    2015-12-08

    Cells of the innate immune system recognize bacterial pathogens by detecting common microbial patterns as well as pathogen-specific activities. One system that responds to these stimuli is the IRE1 branch of the unfolded protein response (UPR), a sensor of endoplasmic reticulum (ER) stress. Activation of IRE1, in the context of Toll-like receptor (TLR) signaling, induces strong proinflammatory cytokine induction. We show here that Legionella pneumophila, an intravacuolar pathogen that replicates in an ER-associated compartment, blocks activation of the IRE1 pathway despite presenting pathogen products that stimulate this response. L. pneumophila TLR ligands induced the splicing of mRNA encoding XBP1s, the main target of IRE1 activity. L. pneumophila was able to inhibit both chemical and bacterial induction of XBP1 splicing via bacterial translocated proteins that interfere with host protein translation. A strain lacking five translocated translation elongation inhibitors was unable to block XBP1 splicing, but this could be rescued by expression of a single such inhibitor, consistent with limitation of the response by translation elongation inhibitors. Chemical inhibition of translation elongation blocked pattern recognition receptor-mediated XBP1 splicing, mimicking the effects of the bacterial translation inhibitors. In contrast, host cell-promoted inhibition of translation initiation in response to the pathogen was ineffective in blocking XBP1 splicing, demonstrating the need for the elongation inhibitors for protection from the UPR. The inhibition of host translation elongation may be a common strategy used by pathogens to limit the innate immune response by interfering with signaling via the UPR.

  5. Melaleuca alternifolia Concentrate Inhibits in Vitro Entry of Influenza Virus into Host Cells

    Directory of Open Access Journals (Sweden)

    Lifang Jiang

    2013-08-01

    Full Text Available Influenza virus causes high morbidity among the infected population annually and occasionally the spread of pandemics. Melaleuca alternifolia Concentrate (MAC is an essential oil derived from a native Australian tea tree. Our aim was to investigate whether MAC has any in vitro inhibitory effect on influenza virus infection and what mechanism does the MAC use to fight the virus infection. In this study, the antiviral activity of MAC was examined by its inhibition of cytopathic effects. In silico prediction was performed to evaluate the interaction between MAC and the viral haemagglutinin. We found that when the influenza virus was incubated with 0.010% MAC for one hour, no cytopathic effect on MDCK cells was found after the virus infection and no immunofluorescence signal was detected in the host cells. Electron microscopy showed that the virus treated with MAC retained its structural integrity. By computational simulations, we found that terpinen-4-ol, which is the major bioactive component of MAC, could combine with the membrane fusion site of haemagglutinin. Thus, we proved that MAC could prevent influenza virus from entering the host cells by disturbing the normal viral membrane fusion procedure.

  6. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    International Nuclear Information System (INIS)

    Liu, Qinghao; Qin, Yixian; Zhou, Lei; Kou, Qiuwen; Guo, Xin; Ge, Xinna; Yang, Hanchun; Hu, Hongbo

    2012-01-01

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/β-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome–lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  7. Autophagy sustains the replication of porcine reproductive and respiratory virus in host cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Qinghao; Qin, Yixian; Zhou, Lei; Kou, Qiuwen; Guo, Xin; Ge, Xinna [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Yang, Hanchun, E-mail: yanghanchun1@cau.edu.cn [Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine and State Key Laboratory of Agribiotechnology, China Agricultural University, Beijing (China); Hu, Hongbo, E-mail: hongbo@cau.edu.cn [College of Food Science and Nutritional Engineering, China Agricultural University, Beijing (China)

    2012-08-01

    In this study, we confirmed the autophagy induced by porcine reproductive and respiratory syndrome virus (PRRSV) in permissive cells and investigated the role of autophagy in the replication of PRRSV. We first demonstrated that PRRSV infection significantly results in the increased double-membrane vesicles, the accumulation of LC3 fluorescence puncta, and the raised ratio of LC3-II/{beta}-actin, in MARC-145 cells. Then we discovered that induction of autophagy by rapamycin significantly enhances the viral titers of PRRSV, while inhibition of autophagy by 3-MA and silencing of LC3 gene by siRNA reduces the yield of PRRSV. The results showed functional autolysosomes can be formed after PRRSV infection and the autophagosome-lysosome-fusion inhibitor decreases the virus titers. We also examined the induction of autophagy by PRRSV infection in pulmonary alveolar macrophages. These findings indicate that autophagy induced by PRRSV infection plays a role in sustaining the replication of PRRSV in host cells.

  8. On the lack of host-cell reactivation of UV-irradiated phage T5

    International Nuclear Information System (INIS)

    Chiang, T.; Harm, W.

    1976-01-01

    Previously reported experiments have shown that host-cell reactivation (HCR) of UV-irradiated phage T1 in excision-repair proficient Escherichia coli cells is inhibited by superinfection with phage T5. Theoretical considerations have led to predictions concerning the dependence of repair inhibition on the multiplicity of superinfecting T5 phage and on the UV fluence to which they were exposed. These predictions have been supported by experimental results described in this paper. The fluence dependence permitted calculation of the relative UV sensitivity of the gene function responsible for repair inhibition; it was found to be about 2.3% that of the plaque-forming ability of phage T5. The T5-inhibitable step in excision repair occurs early in the infective cycle of T1. Furthermore, experiments involving the presence of 400 μg/ml chloramphenicol showed that HCR inhibition of T1 is caused by a protein produced after the FST segment of T5 (i.e. the first 8% of the T5 genome) has entered the host cell. A previously described minor T1 recovery process, occuring in both excision-repair-proficient and -deficient host-cells, is inhibited by T5 infection due to a different substance, which is most likely associated with the 'second-step-transfer' region of T5 DNA (involving the remainder of the genome). Superinfection with T4v 1 phage resulted in HCR inhibition of T1, resembling that observed after T5 superinfection. The discussion of these results suggests that inhibition of the bacterial excision repair system by T5 or T4 infection occurs at the level of UV-endonucleolytic incision, and that lack of HCR both in T-even phages and in T5 can be explained in the same manner

  9. Helicobacter pylori Disrupts Host Cell Membranes, Initiating a Repair Response and Cell Proliferation

    Directory of Open Access Journals (Sweden)

    Hsueh-Fen Juan

    2012-08-01

    Full Text Available Helicobacter pylori (H. pylori, the human stomach pathogen, lives on the inner surface of the stomach and causes chronic gastritis, peptic ulcer, and gastric cancer. Plasma membrane repair response is a matter of life and death for human cells against physical and biological damage. We here test the hypothesis that H. pylori also causes plasma membrane disruption injury, and that not only a membrane repair response but also a cell proliferation response are thereby activated. Vacuolating cytotoxin A (VacA and cytotoxin-associated gene A (CagA have been considered to be major H. pylori virulence factors. Gastric cancer cells were infected with H. pylori wild type (vacA+/cagA+, single mutant (ΔvacA or ΔcagA or double mutant (ΔvacA/ΔcagA strains and plasma membrane disruption events and consequent activation of membrane repair components monitored. H. pylori disrupts the host cell plasma membrane, allowing localized dye and extracellular Ca2+ influx. Ca2+-triggered members of the annexin family, A1 and A4, translocate, in response to injury, to the plasma membrane, and cell surface expression of an exocytotic maker of repair, LAMP-2, increases. Additional forms of plasma membrane disruption, unrelated to H. pylori exposure, also promote host cell proliferation. We propose that H. pylori activation of a plasma membrane repair is pro-proliferative. This study might therefore provide new insight into potential mechanisms of H. pylori-induced gastric carcinogenesis.

  10. Profiling antibody responses to infections by Chlamydia abortus enables identification of potential virulence factors and candidates for serodiagnosis.

    Science.gov (United States)

    Forsbach-Birk, Vera; Foddis, Corinna; Simnacher, Ulrike; Wilkat, Max; Longbottom, David; Walder, Gernot; Benesch, Christiane; Ganter, Martin; Sachse, Konrad; Essig, Andreas

    2013-01-01

    Enzootic abortion of ewes (EAE) due to infection with the obligate intracellular pathogen Chlamydia (C.) abortus is an important zoonosis leading to considerable economic loss to agriculture worldwide. The pathogen can be transmitted to humans and may lead to serious infection in pregnant women. Knowledge about epidemiology, clinical course and transmission to humans is hampered by the lack of reliable diagnostic tools. Immunoreactive proteins, which are expressed in infected animals and humans, may serve as novel candidates for diagnostic marker proteins and represent putative virulence factors. In order to broaden the spectrum of immunogenic C. abortus proteins we applied 2D immunoblot analysis and screening of an expression library using human and animal sera. We have identified 48 immunoreactive proteins representing potential diagnostic markers and also putative virulence factors, such as CAB080 (homologue of the "macrophage infectivity potentiator", MIP), CAB167 (homologue of the "translocated actin recruitment protein", TARP), CAB712 (homologue of the "chlamydial protease-like activity factor", CPAF), CAB776 (homologue of the "Polymorphic membrane protein D", PmpD), and the "hypothetical proteins" CAB063, CAB408 and CAB821, which are predicted to be type III secreted. We selected two putative virulence factors for further characterization, i.e. CAB080 (cMIP) and CAB063, and studied their expression profiles at transcript and protein levels. Analysis of the subcellular localization of both proteins throughout the developmental cycle revealed CAB063 being the first C. abortus protein shown to be translocated to the host cell nucleus.

  11. Profiling antibody responses to infections by Chlamydia abortus enables identification of potential virulence factors and candidates for serodiagnosis.

    Directory of Open Access Journals (Sweden)

    Vera Forsbach-Birk

    Full Text Available Enzootic abortion of ewes (EAE due to infection with the obligate intracellular pathogen Chlamydia (C. abortus is an important zoonosis leading to considerable economic loss to agriculture worldwide. The pathogen can be transmitted to humans and may lead to serious infection in pregnant women. Knowledge about epidemiology, clinical course and transmission to humans is hampered by the lack of reliable diagnostic tools. Immunoreactive proteins, which are expressed in infected animals and humans, may serve as novel candidates for diagnostic marker proteins and represent putative virulence factors. In order to broaden the spectrum of immunogenic C. abortus proteins we applied 2D immunoblot analysis and screening of an expression library using human and animal sera. We have identified 48 immunoreactive proteins representing potential diagnostic markers and also putative virulence factors, such as CAB080 (homologue of the "macrophage infectivity potentiator", MIP, CAB167 (homologue of the "translocated actin recruitment protein", TARP, CAB712 (homologue of the "chlamydial protease-like activity factor", CPAF, CAB776 (homologue of the "Polymorphic membrane protein D", PmpD, and the "hypothetical proteins" CAB063, CAB408 and CAB821, which are predicted to be type III secreted. We selected two putative virulence factors for further characterization, i.e. CAB080 (cMIP and CAB063, and studied their expression profiles at transcript and protein levels. Analysis of the subcellular localization of both proteins throughout the developmental cycle revealed CAB063 being the first C. abortus protein shown to be translocated to the host cell nucleus.

  12. Genome-wide analysis of gene expression and protein secretion of Babesia canis during virulent infection identifies potential pathogenicity factors.

    Science.gov (United States)

    Eichenberger, Ramon M; Ramakrishnan, Chandra; Russo, Giancarlo; Deplazes, Peter; Hehl, Adrian B

    2017-06-13

    Infections of dogs with virulent strains of Babesia canis are characterized by rapid onset and high mortality, comparable to complicated human malaria. As in other apicomplexan parasites, most Babesia virulence factors responsible for survival and pathogenicity are secreted to the host cell surface and beyond where they remodel and biochemically modify the infected cell interacting with host proteins in a very specific manner. Here, we investigated factors secreted by B. canis during acute infections in dogs and report on in silico predictions and experimental analysis of the parasite's exportome. As a backdrop, we generated a fully annotated B. canis genome sequence of a virulent Hungarian field isolate (strain BcH-CHIPZ) underpinned by extensive genome-wide RNA-seq analysis. We find evidence for conserved factors in apicomplexan hemoparasites involved in immune-evasion (e.g. VESA-protein family), proteins secreted across the iRBC membrane into the host bloodstream (e.g. SA- and Bc28 protein families), potential moonlighting proteins (e.g. profilin and histones), and uncharacterized antigens present during acute crisis in dogs. The combined data provides a first predicted and partially validated set of potential virulence factors exported during fatal infections, which can be exploited for urgently needed innovative intervention strategies aimed at facilitating diagnosis and management of canine babesiosis.

  13. Shigella reroutes host cell central metabolism to obtain high-flux nutrient supply for vigorous intracellular growth.

    Science.gov (United States)

    Kentner, David; Martano, Giuseppe; Callon, Morgane; Chiquet, Petra; Brodmann, Maj; Burton, Olga; Wahlander, Asa; Nanni, Paolo; Delmotte, Nathanaël; Grossmann, Jonas; Limenitakis, Julien; Schlapbach, Ralph; Kiefer, Patrick; Vorholt, Julia A; Hiller, Sebastian; Bumann, Dirk

    2014-07-08

    Shigella flexneri proliferate in infected human epithelial cells at exceptionally high rates. This vigorous growth has important consequences for rapid progression to life-threatening bloody diarrhea, but the underlying metabolic mechanisms remain poorly understood. Here, we used metabolomics, proteomics, and genetic experiments to determine host and Shigella metabolism during infection in a cell culture model. The data suggest that infected host cells maintain largely normal fluxes through glycolytic pathways, but the entire output of these pathways is captured by Shigella, most likely in the form of pyruvate. This striking strategy provides Shigella with an abundant favorable energy source, while preserving host cell ATP generation, energy charge maintenance, and survival, despite ongoing vigorous exploitation. Shigella uses a simple three-step pathway to metabolize pyruvate at high rates with acetate as an excreted waste product. The crucial role of this pathway for Shigella intracellular growth suggests targets for antimicrobial chemotherapy of this devastating disease.

  14. Staphylococcus aureus α-toxin-dependent induction of host cell death by membrane-derived vesicles.

    Directory of Open Access Journals (Sweden)

    Bernard Thay

    Full Text Available Staphylococcus aureus causes a wide spectrum of infections in humans, ranging from superficial cutaneous infections, infections in the circum-oral region, to life-threatening bacteremia. It was recently demonstrated that Gram-positive organisms such as S. aureus liberate membrane-derived vesicles (MVs, which analogously to outer membrane vesicles (OMVs of Gram-negative bacteria can play a role in delivering virulence factors to host cells. In the present study we have shown that cholesterol-dependent fusion of S. aureus MVs with the plasma membrane represents a route for delivery of a key virulence factor, α-toxin (α-hemolysin; Hla to human cells. Most S. aureus strains produce this 33-kDa pore-forming protein, which can lyse a wide range of human cells, and induce apoptosis in T-lymphocytes. Our results revealed a tight association of biologically active α-toxin with membrane-derived vesicles isolated from S. aureus strain 8325-4. Concomitantly, α-toxin contributed to HeLa cell cytotoxicity of MVs, and was the main vesicle-associated protein responsible for erythrocyte lysis. In contrast, MVs obtained from an isogenic hla mutant were significantly attenuated with regards to both causing lysis of erythrocytes and death of HeLa cells. This is to our knowledge the first recognition of an S. aureus MV-associated factor contributing to host cell cytotoxicity.

  15. The Pla Protease of Yersinia pestis Degrades Fas Ligand to Manipulate Host Cell Death and Inflammation

    Science.gov (United States)

    Caulfield, Adam J.; Walker, Margaret E.; Gielda, Lindsay M.; Lathem, Wyndham W.

    2014-01-01

    SUMMARY Pneumonic plague is a deadly respiratory disease caused by Yersinia pestis. The bacterial protease Pla contributes to disease progression and manipulation of host immunity, but the mechanisms by which this occurs are largely unknown. Here we show that Pla degrades the apoptotic signaling molecule Fas ligand (FasL) to prevent host cell apoptosis and inflammation. Wild-type Y. pestis, but not a Pla mutant (Δpla), degrades FasL, which results in decreased downstream caspase-3/7 activation and reduced apoptosis. Similarly, lungs of mice challenged with wild-type Y. pestis show reduced levels of FasL and activated caspase-3/7 compared to Δpla infection. Consistent with a role for FasL in regulating immune responses, Δpla infection results in aberrant pro-inflammatory cytokine levels. The loss of FasL or inhibition of caspase activity alters host inflammatory responses and enables enhanced Y. pestis outgrowth in the lungs. Thus, by degrading FasL, Y. pestis manipulates host cell death pathways to facilitate infection. PMID:24721571

  16. Cytokines and the anorexia of infection: potential mechanisms and treatments.

    Science.gov (United States)

    McCarthy, D O

    2000-04-01

    Anorexia during infection is thought to be mediated by immunoregulatory cytokines such as interleukins 1 and 6 and tumor necrosis factor. This article reviews the potential mechanisms of action by which these cytokines are thought to suppress food intake during infection and examines the proposition that blocking of cytokine activity might be one approach to improving food intake of the infected host.

  17. Transfusion transmissible viral infections among potential blood ...

    African Journals Online (AJOL)

    effective approach for prevention and control of transfusion-transmissible infections (TTIs). Also, it has been documented that sub-standard test kits are mostly used in resource limited settings for transfusion related diagnosis. However, the role of ...

  18. Metal binding proteins, recombinant host cells and methods

    Science.gov (United States)

    Summers, Anne O.; Caguiat, Jonathan J.

    2004-06-15

    The present disclosure provides artificial heavy metal binding proteins termed chelons by the inventors. These chelons bind cadmium and/or mercuric ions with relatively high affinity. Also disclosed are coding sequences, recombinant DNA molecules and recombinant host cells comprising those recombinant DNA molecules for expression of the chelon proteins. In the recombinant host cells or transgenic plants, the chelons can be used to bind heavy metals taken up from contaminated soil, groundwater or irrigation water and to concentrate and sequester those ions. Recombinant enteric bacteria can be used within the gastrointestinal tracts of animals or humans exposed to toxic metal ions such as mercury and/or cadmium, where the chelon recombinantly expressed in chosen in accordance with the ion to be rededicated. Alternatively, the chelons can be immobilized to solid supports to bind and concentrate heavy metals from a contaminated aqueous medium including biological fluids.

  19. Cryptosporidia: Epicellular parasites embraced by the host cell membrane

    Czech Academy of Sciences Publication Activity Database

    Valigurová, A.; Jirků, Miloslav; Koudela, Břetislav; Gelnar, M.; Modrý, David; Šlapeta, J.

    2008-01-01

    Roč. 38, 8/9 (2008), s. 913-922 ISSN 0020-7519 R&D Projects: GA ČR GD524/03/H133; GA ČR GA524/05/0992; GA MŠk LC522 Institutional research plan: CEZ:AV0Z60220518 Keywords : Cryptosporidium * host cell invasion * epicellular * parasitophorous sac * ultrastructure Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 3.752, year: 2008

  20. Mycobacterial secretion systems ESX-1 and ESX-5 play distinct roles in host cell death and inflammasome activation

    KAUST Repository

    Abdallah, Abdallah

    2011-09-28

    During infection of humans and animals, pathogenic mycobacteria manipulate the host cell causing severe diseases such as tuberculosis and leprosy. To understand the basis of mycobacterial pathogenicity, it is crucial to identify the molecular virulence mechanisms. In this study, we address the contribution of ESX-1 and ESX-5 - two homologous type VII secretion systems of mycobacteria that secrete distinct sets of immune modulators - during the macrophage infection cycle. Using wild-type, ESX-1- and ESX-5-deficient mycobacterial strains, we demonstrate that these secretion systems differentially affect subcellular localization and macrophage cell responses. We show that in contrast to ESX-1, the effector proteins secreted by ESX-5 are not required for the translocation of Mycobacterium tuberculosis or Mycobacterium marinum to the cytosol of host cells. However, the M. marinum ESX-5 mutant does not induce inflammasome activation and IL-1b activation. The ESX-5 system also induces a caspase-independent cell death after translocation has taken place. Importantly, by means of inhibitory agents and small interfering RNA experiments, we reveal that cathepsin B is involved in both the induction of cell death and inflammasome activation upon infection with wild-type mycobacteria. These results reveal distinct roles for two different type VII secretion systems during infection and shed light on how virulent mycobacteria manipulate the host cell in various ways to replicate and spread. Copyright © 2011 by The American Association of Immunologists, Inc.

  1. Adenovirus E4ORF1-induced MYC activation promotes host cell anabolic glucose metabolism and virus replication.

    Science.gov (United States)

    Thai, Minh; Graham, Nicholas A; Braas, Daniel; Nehil, Michael; Komisopoulou, Evangelia; Kurdistani, Siavash K; McCormick, Frank; Graeber, Thomas G; Christofk, Heather R

    2014-04-01

    Virus infections trigger metabolic changes in host cells that support the bioenergetic and biosynthetic demands of viral replication. Although recent studies have characterized virus-induced changes in host cell metabolism (Munger et al., 2008; Terry et al., 2012), the molecular mechanisms by which viruses reprogram cellular metabolism have remained elusive. Here, we show that the gene product of adenovirus E4ORF1 is necessary for adenovirus-induced upregulation of host cell glucose metabolism and sufficient to promote enhanced glycolysis in cultured epithelial cells by activation of MYC. E4ORF1 localizes to the nucleus, binds to MYC, and enhances MYC binding to glycolytic target genes, resulting in elevated expression of specific glycolytic enzymes. E4ORF1 activation of MYC promotes increased nucleotide biosynthesis from glucose intermediates and enables optimal adenovirus replication in primary lung epithelial cells. Our findings show how a viral protein exploits host cell machinery to reprogram cellular metabolism and promote optimal progeny virion generation. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Increased ATP generation in the host cell is required for efficient vaccinia virus production

    Directory of Open Access Journals (Sweden)

    Hsu Che-Fang

    2009-09-01

    Full Text Available Abstract To search for cellular genes up-regulated by vaccinia virus (VV infection, differential display-reverse transcription-polymerase chain reaction (ddRT-PCR assays were used to examine the expression of mRNAs from mock-infected and VV-infected HeLa cells. Two mitochondrial genes for proteins that are part of the electron transport chain that generates ATP, ND4 and CO II, were up-regulated after VV infection. Up-regulation of ND4 level by VV infection was confirmed by Western blotting analysis. Up-regulation of ND4 was reduced by the MAPK inhibitor, apigenin, which has been demonstrated elsewhere to inhibit VV replication. The induction of ND4 expression occurred after viral DNA replication since ara C, an inhibitor of poxviral DNA replication, could block this induction. ATP production was increased in the host cells after VV infection. Moreover, 4.5 μM oligomycin, an inhibitor of ATP production, reduced the ATP level 13 hr after virus infection to that of mock-infected cells and inhibited viral protein expression and virus production, suggesting that increased ATP production is required for efficient VV production. Our results further suggest that induction of ND4 expression is through a Bcl-2 independent pathway.

  3. Early intranuclear replication of African swine fever virus genome modifies the landscape of the host cell nucleus.

    Science.gov (United States)

    Simões, Margarida; Martins, Carlos; Ferreira, Fernando

    2015-12-02

    Although African swine fever virus (ASFV) replicates in viral cytoplasmic factories, the presence of viral DNA within the host cell nucleus has been previously reported to be essential for productive infection. Herein, we described, for the first time, the intranuclear distribution patterns of viral DNA replication events, preceding those that occur in the cytoplasmic compartment. Using BrdU pulse-labelling experiments, newly synthesized ASFV genomes were exclusively detected inside the host cell nucleus at the early phase of infection, both in swine monocyte-derived macrophages (MDMs) and Vero cells. From 8hpi onwards, BrdU labelling was only observed in ASFV cytoplasmic factories. Our results also show that ASFV specifically activates the Ataxia Telangiectasia Mutated Rad-3 related (ATR) pathway in ASFV-infected swine MDMs from the early phase of infection, most probably because ASFV genome is recognized as foreign DNA. Morphological changes of promyelocytic leukaemia nuclear bodies (PML-NBs), nuclear speckles and Cajal bodies were also found in ASFV-infected swine MDMs, strongly suggesting the viral modulation of cellular antiviral responses and cellular transcription, respectively. As described for other viral infections, the nuclear reorganization that takes place during ASFV infection may also provide an environment that favours its intranuclear replication events. Altogether, our results contribute for a better understanding of ASFV replication strategies, starting with an essential intranuclear DNA replication phase which induces host nucleus changes towards a successful viral infection. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Phospholipase D promotes Arcanobacterium haemolyticum adhesion via lipid raft remodeling and host cell death following bacterial invasion

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    Carlson Petteri

    2010-10-01

    Full Text Available Abstract Background Arcanobacterium haemolyticum is an emerging bacterial pathogen, causing pharyngitis and more invasive infections. This organism expresses an unusual phospholipase D (PLD, which we propose promotes bacterial pathogenesis through its action on host cell membranes. The pld gene is found on a genomic region of reduced %G + C, suggesting recent horizontal acquisition. Results Recombinant PLD rearranged HeLa cell lipid rafts in a dose-dependent manner and this was inhibited by cholesterol sequestration. PLD also promoted host cell adhesion, as a pld mutant had a 60.3% reduction in its ability to adhere to HeLa cells as compared to the wild type. Conversely, the pld mutant appeared to invade HeLa cells approximately two-fold more efficiently as the wild type. This finding was attributable to a significant loss of host cell viability following secretion of PLD from intracellular bacteria. As determined by viability assay, only 15.6% and 82.3% of HeLa cells remained viable following invasion by the wild type or pld mutant, respectively, as compared to untreated HeLa cells. Transmission electron microscopy of HeLa cells inoculated with A. haemolyticum strains revealed that the pld mutant was contained within intracellular vacuoles, as compared to the wild type, which escaped the vacuole. Wild type-infected HeLa cells also displayed the hallmarks of necrosis. Similarly inoculated HeLa cells displayed no signs of apoptosis, as measured by induction of caspase 3/7, 8 or 9 activities. Conclusions These data indicate that PLD enhances bacterial adhesion and promotes host cell necrosis following invasion, and therefore, may be important in the disease pathogenesis of A. haemolyticum infections.

  5. Transcriptional Profiling of Immune-Related Genes in Leishmania infantum-Infected Mice: Identification of Potential Biomarkers of Infection and Progression of Disease

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    Eduardo Ontoria

    2018-06-01

    Full Text Available Leishmania spp. is a protozoan parasite that affects millions of people around the world. At present, there is no effective vaccine to prevent leishmaniases in humans. A major limitation in vaccine development is the lack of precise understanding of the particular immunological mechanisms that allow parasite survival in the host. The parasite-host cell interaction induces dramatic changes in transcriptome patterns in both organisms, therefore, a detailed analysis of gene expression in infected tissues will contribute to the evaluation of drug and vaccine candidates, the identification of potential biomarkers, and the understanding of the immunological pathways that lead to protection or progression of disease. In this large-scale analysis, differential expression of 112 immune-related genes has been analyzed using high-throughput qPCR in spleens of infected and naïve Balb/c mice at four different time points. This analysis revealed that early response against Leishmania infection is characterized by the upregulation of Th1 markers and M1-macrophage activation molecules such as Ifng, Stat1, Cxcl9, Cxcl10, Ccr5, Cxcr3, Xcl1, and Ccl3. This activation doesn't protect spleen from infection, since parasitic burden rises along time. This marked difference in gene expression between infected and control mice disappears during intermediate stages of infection, probably related to the strong anti-inflammatory and immunosuppresory signals that are activated early upon infection (Ctla4 or remain activated throughout the experiment (Il18bp. The overexpression of these Th1/M1 markers is restored later in the chronic phase (8 wpi, suggesting the generation of a classical “protective response” against leishmaniasis. Nonetheless, the parasitic burden rockets at this timepoint. This apparent contradiction can be explained by the generation of a regulatory immune response characterized by overexpression of Ifng, Tnfa, Il10, and downregulation Il4 that

  6. Bystander Host Cell Killing Effects of Clostridium perfringens Enterotoxin

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    Archana Shrestha

    2016-12-01

    Full Text Available Clostridium perfringens enterotoxin (CPE binds to claudin receptors, e.g., claudin-4, and then forms a pore that triggers cell death. Pure cultures of host cells that do not express claudin receptors, e.g., fibroblasts, are unaffected by pathophysiologically relevant CPE concentrations in vitro. However, both CPE-insensitive and CPE-sensitive host cells are present in vivo. Therefore, this study tested whether CPE treatment might affect fibroblasts when cocultured with CPE-sensitive claudin-4 fibroblast transfectants or Caco-2 cells. Under these conditions, immunofluorescence microscopy detected increased death of fibroblasts. This cytotoxic effect involved release of a toxic factor from the dying CPE-sensitive cells, since it could be reproduced using culture supernatants from CPE-treated sensitive cells. Supernatants from CPE-treated sensitive cells, particularly Caco-2 cells, were found to contain high levels of membrane vesicles, often containing a CPE species. However, most cytotoxic activity remained in those supernatants even after membrane vesicle depletion, and CPE was not detected in fibroblasts treated with supernatants from CPE-treated sensitive cells. Instead, characterization studies suggest that a major cytotoxic factor present in supernatants from CPE-treated sensitive cells may be a 10- to 30-kDa host serine protease or require the action of that host serine protease. Induction of caspase-3-mediated apoptosis was found to be important for triggering release of the cytotoxic factor(s from CPE-treated sensitive host cells. Furthermore, the cytotoxic factor(s in these supernatants was shown to induce a caspase-3-mediated killing of fibroblasts. This bystander killing effect due to release of cytotoxic factors from CPE-treated sensitive cells could contribute to CPE-mediated disease.

  7. Sexual Transmission of XMRV: A Potential Infection Route

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    Prachi Sharma

    2011-01-01

    Full Text Available Although XMRV dissemination in humans is a matter of debate, the prostate of select patients seem to harbor XMRV, which raises questions about its potential route of transmission. We established a model of infection in rhesus macaques inoculated with XMRV. In spite of the intravenous inoculation, all infected macaques exhibited readily detectable XMRV signal in the reproductive tract of all 4 males and 1 female during both acute and chronic infection stages. XMRV showed explosive growth in the acini of prostate during acute but not chronic infection. In seminal vesicles, epididymis, and testes, XMRV protein production was detected throughout infection in interstitial or epithelial cells. In the female monkey, epithelial cells in the cervix and vagina were also positive for XMRV gag. The ready detection of XMRV in the reproductive tract of male and female macaques infected intravenously suggests the potential for sexual transmission for XMRV.

  8. Adhesion to the host cell surface is sufficient to mediate Listeria monocytogenes entry into epithelial cells

    Science.gov (United States)

    Ortega, Fabian E.; Rengarajan, Michelle; Chavez, Natalie; Radhakrishnan, Prathima; Gloerich, Martijn; Bianchini, Julie; Siemers, Kathleen; Luckett, William S.; Lauer, Peter; Nelson, W. James; Theriot, Julie A.

    2017-01-01

    The intestinal epithelium is the first physiological barrier breached by the Gram-positive facultative pathogen Listeria monocytogenes during an in vivo infection. Listeria monocytogenes binds to the epithelial host cell receptor E-cadherin, which mediates a physical link between the bacterium and filamentous actin (F-actin). However, the importance of anchoring the bacterium to F-actin through E-cadherin for bacterial invasion has not been tested directly in epithelial cells. Here we demonstrate that depleting αE-catenin, which indirectly links E-cadherin to F-actin, did not decrease L. monocytogenes invasion of epithelial cells in tissue culture. Instead, invasion increased due to increased bacterial adhesion to epithelial monolayers with compromised cell–cell junctions. Furthermore, expression of a mutant E-cadherin lacking the intracellular domain was sufficient for efficient L. monocytogenes invasion of epithelial cells. Importantly, direct biotin-mediated binding of bacteria to surface lipids in the plasma membrane of host epithelial cells was sufficient for uptake. Our results indicate that the only requirement for L. monocytogenes invasion of epithelial cells is adhesion to the host cell surface, and that E-cadherin–mediated coupling of the bacterium to F-actin is not required. PMID:28877987

  9. Sexual Transmission of XMRV: A Potential Infection Route

    OpenAIRE

    Sharma, Prachi; Rogers, Kenneth A.; Suppiah, Suganthi; Molinaro, Ross J.; Onlamoon, Nattawat; Hackett, John; Schochetman, Gerald; Klein, Eric A.; Silverman, Robert H.; Villinger, Fran?ois

    2011-01-01

    Although XMRV dissemination in humans is a matter of debate, the prostate of select patients seem to harbor XMRV, which raises questions about its potential route of transmission. We established a model of infection in rhesus macaques inoculated with XMRV. In spite of the intravenous inoculation, all infected macaques exhibited readily detectable XMRV signal in the reproductive tract of all 4 males and 1 female during both acute and chronic infection stages. XMRV showed explosive growth in th...

  10. Early host cell targets of Yersinia pestis during primary pneumonic plague.

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    Roger D Pechous

    Full Text Available Inhalation of Yersinia pestis causes primary pneumonic plague, a highly lethal syndrome with mortality rates approaching 100%. Pneumonic plague progression is biphasic, with an initial pre-inflammatory phase facilitating bacterial growth in the absence of host inflammation, followed by a pro-inflammatory phase marked by extensive neutrophil influx, an inflammatory cytokine storm, and severe tissue destruction. Using a FRET-based probe to quantitate injection of effector proteins by the Y. pestis type III secretion system, we show that these bacteria target alveolar macrophages early during infection of mice, followed by a switch in host cell preference to neutrophils. We also demonstrate that neutrophil influx is unable to limit bacterial growth in the lung and is ultimately responsible for the severe inflammation during the lethal pro-inflammatory phase.

  11. Host Cell Plasma Membrane Phosphatidylserine Regulates the Assembly and Budding of Ebola Virus.

    Science.gov (United States)

    Adu-Gyamfi, Emmanuel; Johnson, Kristen A; Fraser, Mark E; Scott, Jordan L; Soni, Smita P; Jones, Keaton R; Digman, Michelle A; Gratton, Enrico; Tessier, Charles R; Stahelin, Robert V

    2015-09-01

    Lipid-enveloped viruses replicate and bud from the host cell where they acquire their lipid coat. Ebola virus, which buds from the plasma membrane of the host cell, causes viral hemorrhagic fever and has a high fatality rate. To date, little has been known about how budding and egress of Ebola virus are mediated at the plasma membrane. We have found that the lipid phosphatidylserine (PS) regulates the assembly of Ebola virus matrix protein VP40. VP40 binds PS-containing membranes with nanomolar affinity, and binding of PS regulates VP40 localization and oligomerization on the plasma membrane inner leaflet. Further, alteration of PS levels in mammalian cells inhibits assembly and egress of VP40. Notably, interactions of VP40 with the plasma membrane induced exposure of PS on the outer leaflet of the plasma membrane at sites of egress, whereas PS is typically found only on the inner leaflet. Taking the data together, we present a model accounting for the role of plasma membrane PS in assembly of Ebola virus-like particles. The lipid-enveloped Ebola virus causes severe infection with a high mortality rate and currently lacks FDA-approved therapeutics or vaccines. Ebola virus harbors just seven genes in its genome, and there is a critical requirement for acquisition of its lipid envelope from the plasma membrane of the human cell that it infects during the replication process. There is, however, a dearth of information available on the required contents of this envelope for egress and subsequent attachment and entry. Here we demonstrate that plasma membrane phosphatidylserine is critical for Ebola virus budding from the host cell plasma membrane. This report, to our knowledge, is the first to highlight the role of lipids in human cell membranes in the Ebola virus replication cycle and draws a clear link between selective binding and transport of a lipid across the membrane of the human cell and use of that lipid for subsequent viral entry. Copyright © 2015, American

  12. Legionella Effector AnkX Disrupts Host Cell Endocytic Recycling in a Phosphocholination-Dependent Manner

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    Samual C. Allgood

    2017-09-01

    Full Text Available The facultative intracellular bacterium Legionella pneumophila proliferates within amoebae and human alveolar macrophages, and it is the causative agent of Legionnaires' disease, a life-threatening pneumonia. Within host cells, L. pneumophila establishes a replicative haven by delivering numerous effector proteins into the host cytosol, many of which target membrane trafficking by manipulating the function of Rab GTPases. The Legionella effector AnkX is a phosphocholine transferase that covalently modifies host Rab1 and Rab35. However, a detailed understanding of the biological consequence of Rab GTPase phosphocholination remains elusive. Here, we broaden the understanding of AnkX function by presenting three lines of evidence that it interferes with host endocytic recycling. First, using immunogold transmission electron microscopy, we determined that GFP-tagged AnkX ectopically produced in mammalian cells localizes at the plasma membrane and tubular membrane compartments, sites consistent with targeting the endocytic recycling pathway. Furthermore, the C-terminal region of AnkX was responsible for association with the plasma membrane, and we determined that this region was also able to bind the phosphoinositide lipids PI(3P and PI(4P in vitro. Second, we observed that mCherry-AnkX co-localized with Rab35, a regulator of recycling endocytosis and with major histocompatibility class I protein (MHC-I, a key immunoregulatory protein whose recycling from and back to the plasma membrane is Rab35-dependent. Third, we report that during infection of macrophages, AnkX is responsible for the disruption of endocytic recycling of transferrin, and AnkX's phosphocholination activity is critical for this function. These results support the hypothesis that AnkX targets endocytic recycling during host cell infection. Finally, we have demonstrated that the phosphocholination activity of AnkX is also critical for inhibiting fusion of the Legionella

  13. On the entry of an emerging arbovirus into host cells: Mayaro virus takes the highway to the cytoplasm through fusion with early endosomes and caveolae-derived vesicles

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    Carlos A.M. Carvalho

    2017-04-01

    Full Text Available Mayaro virus (MAYV is an emergent sylvatic alphavirus in South America, related to sporadic outbreaks of a chikungunya-like human febrile illness accompanied by severe arthralgia. Despite its high potential for urban emergence, MAYV is still an obscure virus with scarce information about its infection cycle, including the corresponding early events. Even for prototypical alphaviruses, the cell entry mechanism still has some rough edges to trim: although clathrin-mediated endocytosis is quoted as the putative route, alternative paths as distinct as direct virus genome injection through the cell plasma membrane seems to be possible. Our aim was to clarify crucial details on the entry route exploited by MAYV to gain access into the host cell. Tracking the virus since its first contact with the surface of Vero cells by fluorescence microscopy, we show that its entry occurs by a fast endocytic process and relies on fusion with acidic endosomal compartments. Moreover, blocking clathrin-mediated endocytosis or depleting cholesterol from the cell membrane leads to a strong inhibition of viral infection, as assessed by plaque assays. Following this clue, we found that early endosomes and caveolae-derived vesicles are both implicated as target membranes for MAYV fusion. Our findings unravel the very first events that culminate in a productive infection by MAYV and shed light on potential targets for a rational antiviral therapy, besides providing a better comprehension of the entry routes exploited by alphaviruses to get into the cell.

  14. Lipid exchange between Borrelia burgdorferi and host cells.

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    Jameson T Crowley

    2013-01-01

    Full Text Available Borrelia burgdorferi, the agent of Lyme disease, has cholesterol and cholesterol-glycolipids that are essential for bacterial fitness, are antigenic, and could be important in mediating interactions with cells of the eukaryotic host. We show that the spirochetes can acquire cholesterol from plasma membranes of epithelial cells. In addition, through fluorescent and confocal microscopy combined with biochemical approaches, we demonstrated that B. burgdorferi labeled with the fluorescent cholesterol analog BODIPY-cholesterol or (3H-labeled cholesterol transfer both cholesterol and cholesterol-glycolipids to HeLa cells. The transfer occurs through two different mechanisms, by direct contact between the bacteria and eukaryotic cell and/or through release of outer membrane vesicles. Thus, two-way lipid exchange between spirochetes and host cells can occur. This lipid exchange could be an important process that contributes to the pathogenesis of Lyme disease.

  15. Comparative Analysis of Host Cell Entry of Ebola Virus From Sierra Leone, 2014, and Zaire, 1976.

    Science.gov (United States)

    Hofmann-Winkler, Heike; Gnirß, Kerstin; Wrensch, Florian; Pöhlmann, Stefan

    2015-10-01

    The ongoing Ebola virus (EBOV) disease (EVD) epidemic in Western Africa is the largest EVD outbreak recorded to date and requires the rapid development and deployment of antiviral measures. The viral glycoprotein (GP) facilitates host cell entry and, jointly with cellular interaction partners, constitutes a potential target for antiviral intervention. However, it is unknown whether the GPs of the currently and previously circulating EBOVs use the same mechanisms for cellular entry and are thus susceptible to inhibition by the same antivirals and cellular defenses. Here, we show that the GPs of the EBOVs circulating in 1976 and 2014 transduce the same spectrum of target cells, use the same cellular factors for host cell entry, and are comparably susceptible to blockade by antiviral interferon-induced transmembrane proteins and neutralizing antibody KZ52. Thus, the viruses responsible for the ongoing EVD epidemic should be fully susceptible to established antiviral strategies targeting GP and cellular entry factors. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America.

  16. Potential Cellular Signatures of Viral Infections in Human Hematopoietic Cells

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    J. Mikovits

    2001-01-01

    Full Text Available Expression profiling of cellular genes was performed using a 10,000 cDNA human gene array in order to identify expression changes following chronic infection of human hematopoietic cells with Kapsosi’s Sarcoma -associated Virus (KSHV also known as Human Herpesvirus 8 (HHV8 and Human T cell leukemia virus-1 (HTLV-1. We performed cell-free {\\it in vitro} infection of primary bone marrow derived CD34+ cells using semi-purified HHV8 and a mature IL-2 dependent T cell line, KIT 225, using highly concentrated viral stocks prepared from an infectious molecular clone of HTLV-1. Thirty days post infection, mRNA was isolated from infected cultures and uninfected controls and submitted for microarray analysis. More than 400 genes were differentially expressed more than two-fold following HHV8 infection of primary bone marrow derived CD34+ cells. Of these 400, interferon regulatory factor 4 (IRF4, cyclin B2, TBP-associated factor, eukaryotic elongation factor and pim 2 were up-regulated more than 3.5 fold. In contrast, less than 100 genes were differentially expressed more than two-fold following chronic infection of a mature T cell line with HTLV-1. Of these, only cdc7 was up-regulated more than 3.5 fold. These data may provide insight into cellular signatures of infection useful for diagnosis of infection as well as potential targets for therapeutic intervention.

  17. Profiling Antibody Responses to Infections by Chlamydia abortus Enables Identification of Potential Virulence Factors and Candidates for Serodiagnosis

    Science.gov (United States)

    Forsbach-Birk, Vera; Foddis, Corinna; Simnacher, Ulrike; Wilkat, Max; Longbottom, David; Walder, Gernot; Benesch, Christiane; Ganter, Martin; Sachse, Konrad; Essig, Andreas

    2013-01-01

    Enzootic abortion of ewes (EAE) due to infection with the obligate intracellular pathogen Chlamydia (C.) abortus is an important zoonosis leading to considerable economic loss to agriculture worldwide. The pathogen can be transmitted to humans and may lead to serious infection in pregnant women. Knowledge about epidemiology, clinical course and transmission to humans is hampered by the lack of reliable diagnostic tools. Immunoreactive proteins, which are expressed in infected animals and humans, may serve as novel candidates for diagnostic marker proteins and represent putative virulence factors. In order to broaden the spectrum of immunogenic C. abortus proteins we applied 2D immunoblot analysis and screening of an expression library using human and animal sera. We have identified 48 immunoreactive proteins representing potential diagnostic markers and also putative virulence factors, such as CAB080 (homologue of the “macrophage infectivity potentiator”, MIP), CAB167 (homologue of the “translocated actin recruitment protein”, TARP), CAB712 (homologue of the “chlamydial protease-like activity factor”, CPAF), CAB776 (homologue of the “Polymorphic membrane protein D”, PmpD), and the “hypothetical proteins” CAB063, CAB408 and CAB821, which are predicted to be type III secreted. We selected two putative virulence factors for further characterization, i.e. CAB080 (cMIP) and CAB063, and studied their expression profiles at transcript and protein levels. Analysis of the subcellular localization of both proteins throughout the developmental cycle revealed CAB063 being the first C. abortus protein shown to be translocated to the host cell nucleus. PMID:24260366

  18. Host cell capable of producing enzymes useful for degradation of lignocellulosic material

    Energy Technology Data Exchange (ETDEWEB)

    Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schoonneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

    2017-08-22

    The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

  19. Host cell capable of producing enzymes useful for degradation of lignocellulosic material

    Science.gov (United States)

    Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schooneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

    2015-08-18

    The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

  20. Leishmania (L.) mexicana Infected Bats in Mexico: Novel Potential Reservoirs

    Science.gov (United States)

    Berzunza-Cruz, Miriam; Rodríguez-Moreno, Ángel; Gutiérrez-Granados, Gabriel; González-Salazar, Constantino; Stephens, Christopher R.; Hidalgo-Mihart, Mircea; Marina, Carlos F.; Rebollar-Téllez, Eduardo A.; Bailón-Martínez, Dulce; Balcells, Cristina Domingo; Ibarra-Cerdeña, Carlos N.; Sánchez-Cordero, Víctor; Becker, Ingeborg

    2015-01-01

    Leishmania (Leishmania) mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L.) mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L.) mexicana DNA. We found that 41 bats (9.77%), belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus), and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L.) mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L.) mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology. PMID:25629729

  1. Leishmania (L. mexicana infected bats in Mexico: novel potential reservoirs.

    Directory of Open Access Journals (Sweden)

    Miriam Berzunza-Cruz

    2015-01-01

    Full Text Available Leishmania (Leishmania mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L. mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L. mexicana DNA. We found that 41 bats (9.77%, belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus, and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L. mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L. mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology.

  2. Involvement of Cryptosporidium parvum Cdg7_FLc_1000 RNA in the Attenuation of Intestinal Epithelial Cell Migration via Trans-Suppression of Host Cell SMPD3.

    Science.gov (United States)

    Ming, Zhenping; Gong, Ai-Yu; Wang, Yang; Zhang, Xin-Tian; Li, Min; Mathy, Nicholas W; Strauss-Soukup, Juliane K; Chen, Xian-Ming

    2017-12-27

    Intestinal infection by Cryptosporidium parvum causes inhibition of epithelial turnover, but underlying mechanisms are unclear. Previous studies demonstrate that a panel of parasite RNA transcripts of low protein-coding potential are delivered into infected epithelial cells. Using in vitro and in vivo models of intestinal cryptosporidiosis, we report here that host delivery of parasite Cdg7_FLc_1000 RNA results in inhibition of epithelial cell migration through suppression of the gene encoding sphingomyelinase 3 (SMPD3). Delivery of Cdg7_FLc_1000 into infected cells promotes the histone methyltransferase G9a-mediated H3K9 methylation in the SMPD3 locus. The DNA-binding transcriptional repressor, PR domain zinc finger protein 1, is required for the assembly of Cdg7_FLc_1000 into the G9a complex and associated with the enrichment of H3K9 methylation at the gene locus. Pathologically, nuclear transfer of Cryptosporidium parvum Cdg7_FLc_1000 RNA is involved in the attenuation of intestinal epithelial cell migration via trans-suppression of host cell SMPD3. © The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

  3. A subset of herpes simplex virus replication genes induces DNA amplification within the host cell genome

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    Heilbronn, R.; zur Hausen, H. (Deutsches Krebsforschungszentrum, Heidelberg (West Germany))

    1989-09-01

    Herpes simplex virus (HSV) induces DNA amplification of target genes within the host cell chromosome. To characterize the HSV genes that mediate the amplification effect, combinations of cloned DNA fragments covering the entire HSV genome were transiently transfected into simian virus 40 (SV40)-transformed hamster cells. This led to amplification of the integrated SV40 DNA sequences to a degree comparable to that observed after transfection of intact virion DNA. Transfection of combinations of subclones and of human cytomegalovirus immediate-early promoter-driven expression constructs for individual open reading frames led to the identification of sic HSV genes which together were necessary and sufficient for the induction of DNA amplification: UL30 (DNA polymerase), UL29 (major DNA-binding protein), UL5, UL8, UL42, and UL52. All of these genes encode proteins necessary for HSV DNA replication. However, an additional gene coding for an HSV origin-binding protein (UL9) was required for origin-dependent HSV DNA replication but was dispensable for SV40 DNA amplification. The results show that a subset of HSV replication genes is sufficient for the induction of DNA amplification. This opens the possibility that HSV expresses functions sufficient for DNA amplification but separate from those responsible for lytic viral growth. HSV infection may thereby induce DNA amplification within the host cell genome without killing the host by lytic viral growth. This may lead to persistence of a cell with a new genetic phenotype, which would have implications for the pathogenicity of the virus in vivo.

  4. A Trichomonas vaginalis Rhomboid Protease and Its Substrate Modulate Parasite Attachment and Cytolysis of Host Cells

    Science.gov (United States)

    Riestra, Angelica M.; Gandhi, Shiv; Sweredoski, Michael J.; Moradian, Annie; Hess, Sonja; Urban, Sinisa; Johnson, Patricia J.

    2015-01-01

    Trichomonas vaginalis is an extracellular eukaryotic parasite that causes the most common, non-viral sexually transmitted infection worldwide. Although disease burden is high, molecular mechanisms underlying T. vaginalis pathogenesis are poorly understood. Here, we identify a family of putative T. vaginalis rhomboid proteases and demonstrate catalytic activity for two, TvROM1 and TvROM3, using a heterologous cell cleavage assay. The two T. vaginalis intramembrane serine proteases display different subcellular localization and substrate specificities. TvROM1 is a cell surface membrane protein and cleaves atypical model rhomboid protease substrates, whereas TvROM3 appears to localize to the Golgi apparatus and recognizes a typical model substrate. To identify TvROM substrates, we interrogated the T. vaginalis surface proteome using both quantitative proteomic and bioinformatic approaches. Of the nine candidates identified, TVAG_166850 and TVAG_280090 were shown to be cleaved by TvROM1. Comparison of amino acid residues surrounding the predicted cleavage sites of TvROM1 substrates revealed a preference for small amino acids in the predicted transmembrane domain. Over-expression of TvROM1 increased attachment to and cytolysis of host ectocervical cells. Similarly, mutations that block the cleavage of a TvROM1 substrate lead to its accumulation on the cell surface and increased parasite adherence to host cells. Together, these data indicate a role for TvROM1 and its substrate(s) in modulating attachment to and lysis of host cells, which are key processes in T. vaginalis pathogenesis. PMID:26684303

  5. Identification of host cell proteins which interact with herpes simplex virus type 1 tegument protein pUL37.

    Science.gov (United States)

    Kelly, Barbara J; Diefenbach, Eve; Fraefel, Cornel; Diefenbach, Russell J

    2012-01-20

    The herpes simplex virus type 1 (HSV-1) structural tegument protein pUL37, which is conserved across the Herpesviridae family, is known to be essential for secondary envelopment during the egress of viral particles. To shed light on additional roles of pUL37 during viral replication a yeast two-hybrid screen of a human brain cDNA library was undertaken. This screen identified ten host cell proteins as potential pUL37 interactors. One of the interactors, serine threonine kinase TAOK3, was subsequently confirmed to interact with pUL37 using an in vitro pulldown assay. Such host cell/pUL37 interactions provide further insights into the multifunctional role of this herpesviral tegument protein. Copyright © 2011 Elsevier Inc. All rights reserved.

  6. Brucella abortus choloylglycine hydrolase affects cell envelope composition and host cell internalization.

    Directory of Open Access Journals (Sweden)

    María Inés Marchesini

    Full Text Available Choloylglycine hydrolase (CGH, E.C. 3.5.1.24 is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization.

  7. Equine arteritis virus is delivered to an acidic compartment of host cells via clathrin-dependent endocytosis

    International Nuclear Information System (INIS)

    Nitschke, Matthias; Korte, Thomas; Tielesch, Claudia; Ter-Avetisyan, Gohar; Tuennemann, Gisela; Cardoso, M. Cristina; Veit, Michael; Herrmann, Andreas

    2008-01-01

    Equine arteritis virus (EAV) is an enveloped, positive-stranded RNA virus belonging to the family Arteriviridae. Infection by EAV requires the release of the viral genome by fusion with the respective target membrane of the host cell. We have investigated the entry pathway of EAV into Baby Hamster Kindey cells (BHK). Infection of cells assessed by the plaque reduction assay was strongly inhibited by substances which interfere with clathrin-dependent endocytosis and by lysosomotropic compounds. Furthermore, infection of BHK cells was suppressed when clathrin-dependent endocytosis was inhibited by expression of antisense RNA of the clathrin-heavy chain before infection. These results strongly suggest that EAV is taken up via clathrin-dependent endocytosis and is delivered to acidic endosomal compartments

  8. Characterizing the transmission potential of zoonotic infections from minor outbreaks.

    Directory of Open Access Journals (Sweden)

    Adam J Kucharski

    2015-04-01

    Full Text Available The transmission potential of a novel infection depends on both the inherent transmissibility of a pathogen, and the level of susceptibility in the host population. However, distinguishing between these pathogen- and population-specific properties typically requires detailed serological studies, which are rarely available in the early stages of an outbreak. Using a simple transmission model that incorporates age-stratified social mixing patterns, we present a novel method for characterizing the transmission potential of subcritical infections, which have effective reproduction number R<1, from readily available data on the size of outbreaks. We show that the model can identify the extent to which outbreaks are driven by inherent pathogen transmissibility and pre-existing population immunity, and can generate unbiased estimates of the effective reproduction number. Applying the method to real-life infections, we obtained accurate estimates for the degree of age-specific immunity against monkeypox, influenza A(H5N1 and A(H7N9, and refined existing estimates of the reproduction number. Our results also suggest minimal pre-existing immunity to MERS-CoV in humans. The approach we describe can therefore provide crucial information about novel infections before serological surveys and other detailed analyses are available. The methods would also be applicable to data stratified by factors such as profession or location, which would make it possible to measure the transmission potential of emerging infections in a wide range of settings.

  9. Leptospirosis risk around a potential source of infection

    Science.gov (United States)

    Loaiza-Echeverry, Erica; Hincapié-Palacio, Doracelly; Ochoa Acosta, Jesús; Ospina Giraldo, Juan

    2015-05-01

    Leptospirosis is a bacterial zoonosis with world distribution and multiform clinical spectrum in men and animals. The etiology of this disease is the pathogenic species of Leptospira, which cause diverse manifestations of the disease, from mild to serious, such as the Weil disease and the lung hemorrhagic syndrome with lethal proportions of 10% - 50%. This is an emerging problem of urban health due to the growth of marginal neighborhoods without basic sanitary conditions and an increased number of rodents. The presence of rodents and the probability of having contact with their urine determine the likelihood for humans to get infected. In this paper, we simulate the spatial distribution of risk infection of human leptospirosis according to the proximity to rodent burrows considered as potential source of infection. The Bessel function K0 with an r distance from the potential point source, and the scale parameter α in meters was used. Simulation inputs were published data of leptospirosis incidence rate (range of 5 to 79 x 10 000), and a distance of 100 to 5000 meters from the source of infection. We obtained an adequate adjustment between the function and the simulated data. The risk of infection increases with the proximity of the potential source. This estimation can become a guide to propose effective measures of control and prevention.

  10. Screening for potential anti-infective agents towards Burkholderia pseudomallei infection

    Science.gov (United States)

    Eng, Su Anne; Nathan, Sheila

    2014-09-01

    The established treatment for melioidosis is antibiotic therapy. However, a constant threat to this form of treatment is resistance development of the causative agent, Burkholderia pseudomallei, towards antibiotics. One option to circumvent this threat of antibiotic resistance is to search for new alternative anti-infectives which target the host innate immune system and/or bacterial virulence. In this study, 29 synthetic compounds were evaluated for their potential to increase the lifespan of an infected host. The nematode Caenorhabditis elegans was adopted as the infection model as its innate immune pathways are homologous to humans. Screens were performed in a liquid-based survival assay containing infected worms exposed to individual compounds and survival of untreated and compound-treated worms were compared. A primary screen identified nine synthetic compounds that extended the lifespan of B. pseudomallei-infected worms. Subsequently, a disc diffusion test was performed on these selected compounds to delineate compounds into those that enhanced the survival of worms via antimicrobial activity i.e. reducing the number of infecting bacteria, or into those that did not target pathogen viability. Out of the nine hits selected, two demonstrated antimicrobial effects on B. pseudomallei. Therefore, the findings from this study suggest that the other seven identified compounds are potential anti-infectives which could protect a host against B. pseudomallei infection without developing the risk of drug resistance.

  11. Rift Valley fever potential mosquito vectors and their infection status ...

    African Journals Online (AJOL)

    Background: Rift Valley fever (RVF) is a mosquito-borne viral zoonotic disease. Rift Valley fever virus (RVFV) has been isolated from more than 40 species of mosquitoes from eight genera. This study was conducted to determine the abundance of potential mosquito vectors and their RVFV infection status in Ngorongoro ...

  12. Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing.

    Directory of Open Access Journals (Sweden)

    Katrin Bomans

    Full Text Available Biotherapeutics are often produced in non-human host cells like Escherichia coli, yeast, and various mammalian cell lines. A major focus of any therapeutic protein purification process is to reduce host cell proteins to an acceptable low level. In this study, various E. coli host cell proteins were identified at different purifications steps by HPLC fractionation, SDS-PAGE analysis, and tryptic peptide mapping combined with online liquid chromatography mass spectrometry (LC-MS. However, no host cell proteins could be verified by direct LC-MS analysis of final drug substance material. In contrast, the application of affinity enrichment chromatography prior to comprehensive LC-MS was adequate to identify several low abundant host cell proteins at the final drug substance level. Bacterial alkaline phosphatase (BAP was identified as being the most abundant host cell protein at several purification steps. Thus, we firstly established two different assays for enzymatic and immunological BAP monitoring using the cobas® technology. By using this strategy we were able to demonstrate an almost complete removal of BAP enzymatic activity by the established therapeutic protein purification process. In summary, the impact of fermentation, purification, and formulation conditions on host cell protein removal and biological activity can be conducted by monitoring process-specific host cell proteins in a GMP-compatible and high-throughput (> 1000 samples/day manner.

  13. Dengue virus-induced regulation of the host cell translational machinery

    Directory of Open Access Journals (Sweden)

    C.S.A. Villas-Bôas

    2009-11-01

    Full Text Available Dengue virus (DV-induced changes in the host cell protein synthesis machinery are not well understood. We investigated the transcriptional changes related to initiation of protein synthesis. The human hepatoma cell line, HepG2, was infected with DV serotype 2 for 1 h at a multiplicity of infection of one. RNA was extracted after 6, 24 and 48 h. Microarray results showed that 36.5% of the translation factors related to initiation of protein synthesis had significant differential expression (Z-score ≥ ±2.0. Confirmation was obtained by quantitative real-time reverse transcription-PCR. Of the genes involved in the activation of mRNA for cap-dependent translation (eIF4 factors, eIF4A, eIF4G1 and eIF4B were up-regulated while the negative regulator of translation eIF4E-BP3 was down-regulated. This activation was transient since at 24 h post-infection levels were not significantly different from control cells. However, at 48 h post-infection, eIF4A, eIF4E, eIF4G1, eIF4G3, eIF4B, and eIF4E-BP3 were down-regulated, suggesting that cap-dependent translation could be inhibited during the progression of infection. To test this hypothesis, phosphorylation of p70S6K and 4E-BP1, which induce cap-dependent protein synthesis, was assayed. Both proteins remained phosphorylated when assayed at 6 h after infection, while infection induced dephosphorylation of p70S6K and 4E-BP1 at 24 and 48 h of infection, respectively. Taken together, these results provide biological evidence suggesting that in HepG2 cells DV sustains activation of the cap-dependent machinery at early stages of infection, but progression of infection switches protein synthesis to a cap-independent process.

  14. Roles of Chaperone/Usher Pathways of Yersinia pestis in a Murine Model of Plague and Adhesion to Host Cells

    Science.gov (United States)

    Hatkoff, Matthew; Runco, Lisa M.; Pujol, Celine; Jayatilaka, Indralatha; Furie, Martha B.; Bliska, James B.

    2012-01-01

    Yersinia pestis and many other Gram-negative pathogenic bacteria use the chaperone/usher (CU) pathway to assemble virulence-associated surface fibers termed pili or fimbriae. Y. pestis has two well-characterized CU pathways: the caf genes coding for the F1 capsule and the psa genes coding for the pH 6 antigen. The Y. pestis genome contains additional CU pathways that are capable of assembling pilus fibers, but the roles of these pathways in the pathogenesis of plague are not understood. We constructed deletion mutations in the usher genes for six of the additional Y. pestis CU pathways. The wild-type (WT) and usher deletion strains were compared in the murine bubonic (subcutaneous) and pneumonic (intranasal) plague infection models. Y. pestis strains containing deletions in CU pathways y0348-0352, y1858-1862, and y1869-1873 were attenuated for virulence compared to the WT strain by the intranasal, but not subcutaneous, routes of infection, suggesting specific roles for these pathways during pneumonic plague. We examined binding of the Y. pestis WT and usher deletion strains to A549 human lung epithelial cells, HEp-2 human cervical epithelial cells, and primary human and murine macrophages. Y. pestis CU pathways y0348-0352 and y1858-1862 were found to contribute to adhesion to all host cells tested, whereas pathway y1869-1873 was specific for binding to macrophages. The correlation between the virulence attenuation and host cell binding phenotypes of the usher deletion mutants identifies three of the additional CU pathways of Y. pestis as mediating interactions with host cells that are important for the pathogenesis of plague. PMID:22851745

  15. Development of hyper osmotic resistant CHO host cells for enhanced antibody production.

    Science.gov (United States)

    Kamachi, Yasuharu; Omasa, Takeshi

    2018-04-01

    Cell culture platform processes are generally employed to shorten the duration of new product development. A fed-batch process with continuous feeding is a conventional platform process for monoclonal antibody production using Chinese hamster ovary (CHO) cells. To establish a simplified platform process, the feeding method can be changed from continuous feed to bolus feed. However, this change induces a rapid increase of osmolality by the bolus addition of nutrients. The increased osmolality suppresses cell culture growth, and the final product concentration is decreased. In this study, osmotic resistant CHO host cells were developed to attain a high product concentration. To establish hyper osmotic resistant CHO host cells, CHO-S host cells were passaged long-term in a hyper osmotic basal medium. There were marked differences in cell growth of the original and established host cells under iso- (328 mOsm/kg) or hyper-osmolality (over 450 mOsm/kg) conditions. Cell growth of the original CHO host cells was markedly decreased by the induction of osmotic stress, whereas cell growth of the hyper osmotic resistant CHO host cells was not affected. The maximum viable cell concentration of hyper osmotic resistant CHO host cells was 132% of CHO-S host cells after the induction of osmotic stress. Moreover, the hyper osmotic resistant characteristic of established CHO host cells was maintained even after seven passages in iso-osmolality basal medium. The use of hyper osmotic resistance CHO host cells to create a monoclonal antibody production cell line might be a new approach to increase final antibody concentrations with a fed-batch process. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  16. Ebola virus. Two-pore channels control Ebola virus host cell entry and are drug targets for disease treatment.

    Science.gov (United States)

    Sakurai, Yasuteru; Kolokoltsov, Andrey A; Chen, Cheng-Chang; Tidwell, Michael W; Bauta, William E; Klugbauer, Norbert; Grimm, Christian; Wahl-Schott, Christian; Biel, Martin; Davey, Robert A

    2015-02-27

    Ebola virus causes sporadic outbreaks of lethal hemorrhagic fever in humans, but there is no currently approved therapy. Cells take up Ebola virus by macropinocytosis, followed by trafficking through endosomal vesicles. However, few factors controlling endosomal virus movement are known. Here we find that Ebola virus entry into host cells requires the endosomal calcium channels called two-pore channels (TPCs). Disrupting TPC function by gene knockout, small interfering RNAs, or small-molecule inhibitors halted virus trafficking and prevented infection. Tetrandrine, the most potent small molecule that we tested, inhibited infection of human macrophages, the primary target of Ebola virus in vivo, and also showed therapeutic efficacy in mice. Therefore, TPC proteins play a key role in Ebola virus infection and may be effective targets for antiviral therapy. Copyright © 2015, American Association for the Advancement of Science.

  17. Dendrimers as Potential Therapeutic Tools in HIV Inhibition

    Directory of Open Access Journals (Sweden)

    Xiangbo Li

    2013-07-01

    Full Text Available The present treatments for HIV transfection include chemical agents and gene therapies. Although many chemical drugs, peptides and genes have been developed for HIV inhibition, a variety of non-ignorable drawbacks limited the efficiency of these materials. In this review, we discuss the application of dendrimers as both therapeutic agents and non-viral vectors of chemical agents and genes for HIV treatment. On the one hand, dendrimers with functional end groups combine with the gp120 of HIV and CD4 molecule of host cell to suppress the attachment of HIV to the host cell. Some of the dendrimers are capable of intruding into the cell and interfere with the later stages of HIV replication as well. On the other hand, dendrimers are also able to transfer chemical drugs and genes into the host cells, which conspicuously increase the anti-HIV activity of these materials. Dendrimers as therapeutic tools provide a potential treatment for HIV infection.

  18. Dengue Virus Selectively Annexes Endoplasmic Reticulum-Associated Translation Machinery as a Strategy for Co-opting Host Cell Protein Synthesis.

    Science.gov (United States)

    Reid, David W; Campos, Rafael K; Child, Jessica R; Zheng, Tianli; Chan, Kitti Wing Ki; Bradrick, Shelton S; Vasudevan, Subhash G; Garcia-Blanco, Mariano A; Nicchitta, Christopher V

    2018-04-01

    A primary question in dengue virus (DENV) biology is the molecular strategy for recruitment of host cell protein synthesis machinery. Here, we combined cell fractionation, ribosome profiling, and transcriptome sequencing (RNA-seq) to investigate the subcellular organization of viral genome translation and replication as well as host cell translation and its response to DENV infection. We report that throughout the viral life cycle, DENV plus- and minus-strand RNAs were highly partitioned to the endoplasmic reticulum (ER), identifying the ER as the primary site of DENV translation. DENV infection was accompanied by an ER compartment-specific remodeling of translation, where ER translation capacity was subverted from host transcripts to DENV plus-strand RNA, particularly at late stages of infection. Remarkably, translation levels and patterns in the cytosol compartment were only modestly affected throughout the experimental time course of infection. Comparisons of ribosome footprinting densities of the DENV plus-strand RNA and host mRNAs indicated that DENV plus-strand RNA was only sparsely loaded with ribosomes. Combined, these observations suggest a mechanism where ER-localized translation and translational control mechanisms, likely cis encoded, are used to repurpose the ER for DENV virion production. Consistent with this view, we found ER-linked cellular stress response pathways commonly associated with viral infection, namely, the interferon response and unfolded protein response, to be only modestly activated during DENV infection. These data support a model where DENV reprograms the ER protein synthesis and processing environment to promote viral survival and replication while minimizing the activation of antiviral and proteostatic stress response pathways. IMPORTANCE DENV, a prominent human health threat with no broadly effective or specific treatment, depends on host cell translation machinery for viral replication, immune evasion, and virion biogenesis. The

  19. Scientists discover how deadly fungal microbes enter host cells

    OpenAIRE

    Whyte, Barry James

    2010-01-01

    A research team led by scientists at the Virginia Bioinformatics Institute at Virginia Tech has discovered a fundamental entry mechanism that allows dangerous fungal microbes to infect plants and cause disease.

  20. Purification of infectious human herpesvirus 6A virions and association of host cell proteins

    Directory of Open Access Journals (Sweden)

    Garoff Henrik

    2007-10-01

    Full Text Available Abstract Background Viruses that are incorporating host cell proteins might trigger autoimmune diseases. It is therefore of interest to identify possible host proteins associated with viruses, especially for enveloped viruses that have been suggested to play a role in autoimmune diseases, like human herpesvirus 6A (HHV-6A in multiple sclerosis (MS. Results We have established a method for rapid and morphology preserving purification of HHV-6A virions, which in combination with parallel analyses with background control material released from mock-infected cells facilitates qualitative and quantitative investigations of the protein content of HHV-6A virions. In our iodixanol gradient purified preparation, we detected high levels of viral DNA by real-time PCR and viral proteins by metabolic labelling, silver staining and western blots. In contrast, the background level of cellular contamination was low in the purified samples as demonstrated by the silver staining and metabolic labelling analyses. Western blot analyses showed that the cellular complement protein CD46, the receptor for HHV-6A, is associated with the purified and infectious virions. Also, the cellular proteins clathrin, ezrin and Tsg101 are associated with intact HHV-6A virions. Conclusion Cellular proteins are associated with HHV-6A virions. The relevance of the association in disease and especially in autoimmunity will be further investigated.

  1. Mycobacterium leprae–host-cell interactions and genetic determinants in leprosy: an overview

    Science.gov (United States)

    Pinheiro, Roberta Olmo; de Souza Salles, Jorgenilce; Sarno, Euzenir Nunes; Sampaio, Elizabeth Pereira

    2011-01-01

    Leprosy, also known as Hansen’s disease, is a chronic infectious disease caused by Mycobacterium leprae in which susceptibility to the mycobacteria and its clinical manifestations are attributed to the host immune response. Even though leprosy prevalence has decreased dramatically, the high number of new cases indicates active transmission. Owing to its singular features, M. leprae infection is an attractive model for investigating the regulation of human immune responses to pathogen-induced disease. Leprosy is one of the most common causes of nontraumatic peripheral neuropathy worldwide. The proportion of patients with disabilities is affected by the type of leprosy and delay in diagnosis. This article briefly reviews the clinical features as well as the immunopathological mechanisms related to the establishment of the different polar forms of leprosy, the mechanisms related to M. leprae–host cell interactions and prophylaxis and diagnosis of this complex disease. Host genetic factors are summarized and the impact of the development of interventions that prevent, reverse or limit leprosy-related nerve impairments are discussed. PMID:21366421

  2. [Stethoscope or staphyloscope?: Potential vector in nosocomial infections].

    Science.gov (United States)

    Zúniga, Andrés; Mañalich, Jaime; Cortés, Rosario

    2016-02-01

    Healthcare-associated infections (HCAI) are a problem worldwide. In our country, the estimated incidence of HCAI is 70,000 per year. This results in an increase in the average length of hospital stay by 10 days per patient, an estimated annual cost of US $ 70 million and an overstay of 700 thousand bed days a year. For over 30 years stethoscopes have been considered as potential HCAI vectors, since pathogens like methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus strains adhere and colonize them. These organisms can be transmitted between patients if the instruments are not sanitized. Several studies conclude that disinfecting the stethoscope with isopropyl alcohol eliminates up to 99% of bacteria. Simple, economic measures such as implementation of guidelines for stethoscope disinfection are a clear opportunity for preventing infections.

  3. Group 2 coronaviruses prevent immediate early interferon induction by protection of viral RNA from host cell recognition

    International Nuclear Information System (INIS)

    Versteeg, Gijs A.; Bredenbeek, Peter J.; Worm, Sjoerd H.E. van den; Spaan, Willy J.M.

    2007-01-01

    Many viruses encode antagonists to prevent interferon (IFN) induction. Infection of fibroblasts with the murine hepatitis coronavirus (MHV) and SARS-coronavirus (SARS-CoV) did not result in nuclear translocation of interferon-regulatory factor 3 (IRF3), a key transcription factor involved in IFN induction, and induction of IFN mRNA transcription. Furthermore, MHV and SARS-CoV infection could not prevent IFN induction by poly (I:C) or Sendai virus, suggesting that these CoVs do not inactivate IRF3-mediated transcription regulation, but apparently prevent detection of replicative RNA by cellular sensory molecules. Our data indicate that shielding of viral RNA to host cell sensors might be the main general mechanism for coronaviruses to prevent IFN induction

  4. Members of a novel protein family containing microneme adhesive repeat domains act as sialic acid-binding lectins during host cell invasion by apicomplexan parasites.

    Science.gov (United States)

    Friedrich, Nikolas; Santos, Joana M; Liu, Yan; Palma, Angelina S; Leon, Ester; Saouros, Savvas; Kiso, Makoto; Blackman, Michael J; Matthews, Stephen; Feizi, Ten; Soldati-Favre, Dominique

    2010-01-15

    Numerous intracellular pathogens exploit cell surface glycoconjugates for host cell recognition and entry. Unlike bacteria and viruses, Toxoplasma gondii and other parasites of the phylum Apicomplexa actively invade host cells, and this process critically depends on adhesins (microneme proteins) released onto the parasite surface from intracellular organelles called micronemes (MIC). The microneme adhesive repeat (MAR) domain of T. gondii MIC1 (TgMIC1) recognizes sialic acid (Sia), a key determinant on the host cell surface for invasion by this pathogen. By complementation and invasion assays, we demonstrate that TgMIC1 is one important player in Sia-dependent invasion and that another novel Sia-binding lectin, designated TgMIC13, is also involved. Using BLAST searches, we identify a family of MAR-containing proteins in enteroparasitic coccidians, a subclass of apicomplexans, including T. gondii, suggesting that all these parasites exploit sialylated glycoconjugates on host cells as determinants for enteric invasion. Furthermore, this protein family might provide a basis for the broad host cell range observed for coccidians that form tissue cysts during chronic infection. Carbohydrate microarray analyses, corroborated by structural considerations, show that TgMIC13, TgMIC1, and its homologue Neospora caninum MIC1 (NcMIC1) share a preference for alpha2-3- over alpha2-6-linked sialyl-N-acetyllactosamine sequences. However, the three lectins also display differences in binding preferences. Intense binding of TgMIC13 to alpha2-9-linked disialyl sequence reported on embryonal cells and relatively strong binding to 4-O-acetylated-Sia found on gut epithelium and binding of NcMIC1 to 6'sulfo-sialyl Lewis(x) might have implications for tissue tropism.

  5. [Animals as a potential source of human fungal infections].

    Science.gov (United States)

    Dworecka-Kaszak, Bozena

    2008-01-01

    Changing environment is a reason, that many saprotrophic fungi became opportunists and in the end also maybe a pathogenic. Host specific adaptation is not so strong among fungi, so there are many common fungal pathogens for people and for animals. Animals suffering from dermatomycosis are well recognize as source of human superficial mycoses. Breeding of different exotic animals such as parrots, various Reptiles and Amphibians, miniature Rodents and keeping them as a pets in the peoples houses, have become more and more popular in the recent years. This article is shortly presenting which animals maybe a potential source of fungal infections for humans. Looking for the other mycoses as systemic mycoses, especially candidiasis or aspergilosis there are no data, which allow excluding sick animals as a source of infection for human, even if those deep mycoses have endogenic reactivation mechanism. Immunocompromised people are in high-risk group when they take care of animals. Another important source of potentially pathogenic, mostly air-born fungi may be animal use in experimental laboratory work. During the experiments is possible that laboratory workers maybe hurt and these animals and their environment, food and house boxes could be the possible source of microorganisms, pathogenic for humans or other animals. Unusual way to inoculate these potentially pathogens into the skin of laboratory personnel may cause granulomatous, local lesions on their hands.

  6. Infection Unit Density as an Index of Infection Potential of Arbuscular Mycorrhizal Fungi.

    Science.gov (United States)

    Ohtomo, Ryo; Kobae, Yoshihiro; Morimoto, Sho; Oka, Norikuni

    2018-03-29

    The effective use of arbuscular mycorrhizal (AM) fungal function to promote host plant phosphate uptake in agricultural practice requires the accurate quantitative evaluation of AM fungal infection potential in field soil or AM fungal inoculation material. The number of infection units (IUs), intraradical fungal structures derived from single root entries formed after a short cultivation period, may reflect the number of propagules in soil when pot soil is completely permeated by the host root. However, the original IU method, in which all AM propagules in a pot are counted, requires the fine tuning of plant growing conditions and is considered to be laborious. The objective of the present study was to test whether IU density, not the total count of IU, but the number of IUs per unit root length, reflects the density of AM fungal propagules in soil. IU density assessed after 12 d of host plant cultivation and 3,3'-diaminobenzidine (DAB) staining showed a stronger linear correlation with propagule density than the mean infection percentage (MIP). In addition, IU density was affected less by the host plant species than MIP. We suggest that IU density provides a more rapid and reliable quantitation of the propagule density of AM fungi than MIP or the original IU method. Thus, IU density may be a more robust index of AM fungal infection potential for research and practical applications.

  7. Transmission potential of Zika virus infection in the South Pacific.

    Science.gov (United States)

    Nishiura, Hiroshi; Kinoshita, Ryo; Mizumoto, Kenji; Yasuda, Yohei; Nah, Kyeongah

    2016-04-01

    Zika virus has spread internationally through countries in the South Pacific and Americas. The present study aimed to estimate the basic reproduction number, R0, of Zika virus infection as a measurement of the transmission potential, reanalyzing past epidemic data from the South Pacific. Incidence data from two epidemics, one on Yap Island, Federal State of Micronesia in 2007 and the other in French Polynesia in 2013-2014, were reanalyzed. R0 of Zika virus infection was estimated from the early exponential growth rate of these two epidemics. The maximum likelihood estimate (MLE) of R0 for the Yap Island epidemic was in the order of 4.3-5.8 with broad uncertainty bounds due to the small sample size of confirmed and probable cases. The MLE of R0 for French Polynesia based on syndromic data ranged from 1.8 to 2.0 with narrow uncertainty bounds. The transmissibility of Zika virus infection appears to be comparable to those of dengue and chikungunya viruses. Considering that Aedes species are a shared vector, this finding indicates that Zika virus replication within the vector is perhaps comparable to dengue and chikungunya. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  8. Surface-layer protein A (SlpA is a major contributor to host-cell adherence of Clostridium difficile.

    Directory of Open Access Journals (Sweden)

    Michelle M Merrigan

    Full Text Available Clostridium difficile is a leading cause of antibiotic-associated diarrhea, and a significant etiologic agent of healthcare-associated infections. The mechanisms of attachment and host colonization of C. difficile are not well defined. We hypothesize that non-toxin bacterial factors, especially those facilitating the interaction of C. difficile with the host gut, contribute to the initiation of C. difficile infection. In this work, we optimized a completely anaerobic, quantitative, epithelial-cell adherence assay for vegetative C. difficile cells, determined adherence proficiency under multiple conditions, and investigated C. difficile surface protein variation via immunological and DNA sequencing approaches focused on Surface-Layer Protein A (SlpA. In total, thirty-six epidemic-associated and non-epidemic associated C. difficile clinical isolates were tested in this study, and displayed intra- and inter-clade differences in attachment that were unrelated to toxin production. SlpA was a major contributor to bacterial adherence, and individual subunits of the protein (varying in sequence between strains mediated host-cell attachment to different extents. Pre-treatment of host cells with crude or purified SlpA subunits, or incubation of vegetative bacteria with anti-SlpA antisera significantly reduced C. difficile attachment. SlpA-mediated adherence-interference correlated with the attachment efficiency of the strain from which the protein was derived, with maximal blockage observed when SlpA was derived from highly adherent strains. In addition, SlpA-containing preparations from a non-toxigenic strain effectively blocked adherence of a phylogenetically distant, epidemic-associated strain, and vice-versa. Taken together, these results suggest that SlpA plays a major role in C. difficile infection, and that it may represent an attractive target for interventions aimed at abrogating gut colonization by this pathogen.

  9. Whole genome transcription profiling of Anaplasma phagocytophilum in human and tick host cells by tiling array analysis

    Directory of Open Access Journals (Sweden)

    Chavez Adela

    2008-07-01

    Full Text Available Abstract Background Anaplasma phagocytophilum (Ap is an obligate intracellular bacterium and the agent of human granulocytic anaplasmosis, an emerging tick-borne disease. Ap alternately infects ticks and mammals and a variety of cell types within each. Understanding the biology behind such versatile cellular parasitism may be derived through the use of tiling microarrays to establish high resolution, genome-wide transcription profiles of the organism as it infects cell lines representative of its life cycle (tick; ISE6 and pathogenesis (human; HL-60 and HMEC-1. Results Detailed, host cell specific transcriptional behavior was revealed. There was extensive differential Ap gene transcription between the tick (ISE6 and the human (HL-60 and HMEC-1 cell lines, with far fewer differentially transcribed genes between the human cell lines, and all disproportionately represented by membrane or surface proteins. There were Ap genes exclusively transcribed in each cell line, apparent human- and tick-specific operons and paralogs, and anti-sense transcripts that suggest novel expression regulation processes. Seven virB2 paralogs (of the bacterial type IV secretion system showed human or tick cell dependent transcription. Previously unrecognized genes and coding sequences were identified, as were the expressed p44/msp2 (major surface proteins paralogs (of 114 total, through elevated signal produced to the unique hypervariable region of each – 2/114 in HL-60, 3/114 in HMEC-1, and none in ISE6. Conclusion Using these methods, whole genome transcription profiles can likely be generated for Ap, as well as other obligate intracellular organisms, in any host cells and for all stages of the cell infection process. Visual representation of comprehensive transcription data alongside an annotated map of the genome renders complex transcription into discernable patterns.

  10. Comparative proteome analysis reveals conserved and specific adaptation patterns of Staphylococcus aureus after internalization by different types of human non-professional phagocytic host cells

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    Kristin eSurmann

    2014-08-01

    Full Text Available Staphylococcus aureus is a human pathogen that can cause a wide range of diseases. Although formerly regarded as extracellular pathogen, it has been shown that S. aureus can also be internalized by host cells and persist within these cells. In the present study, we comparatively analyzed survival and physiological adaptation of S. aureus HG001 after internalization by two human lung epithelial cell lines (S9 and A549, and human embryonic kidney cells (HEK 293. Combining enrichment of bacteria from host-pathogen assays by cell sorting and quantitation of the pathogen´s proteome by mass spectrometry we characterized S. aureus adaptation during the initial phase between 2.5 h and 6.5 h post-infection. Starting with about 2x106 bacteria, roughly 1,450 S. aureus proteins, including virulence factors and metabolic enzymes were identified by spectral comparison and classical database searches. Most of the bacterial adaptation reactions, such as decreases in levels of ribosomal proteins and metabolic enzymes or increases in amounts of proteins involved in arginine and lysine biosynthesis, coding for terminal oxidases and stress responsive genes or activation of the sigma factor SigB were observed after internalization into any of the three cell lines studied. However, differences were noted in central carbon metabolism including regulation of fermentation and threonine degradation. Since these differences coincided with different intracellular growth behavior, complementary profiling of the metabolome of the different non-infected host cell types was performed. This revealed similar levels of intracellular glucose but host cell specific differences in the amounts of amino acids such as glycine, threonine or glutamate. With this comparative study we provide an impression of the common and specific features of the adaptation of S. aureus HG001 to specific host cell environments as a starting point for follow-up studies with different strain isolates and

  11. Trans-suppression of defense DEFB1 gene in intestinal epithelial cells following Cryptosporidium parvum infection is associated with host delivery of parasite Cdg7_FLc_1000 RNA.

    Science.gov (United States)

    Ming, Zhenping; Gong, Ai-Yu; Wang, Yang; Zhang, Xin-Tian; Li, Min; Dolata, Courtney E; Chen, Xian-Ming

    2018-03-01

    To counteract host immunity, Cryptosporidium parvum has evolved multiple strategies to suppress host antimicrobial defense. One such strategy is to reduce the production of the antimicrobial peptide beta-defensin 1 (DEFB1) by host epithelial cells but the underlying mechanisms remain unclear. Recent studies demonstrate that a panel of parasite RNA transcripts of low protein-coding potential are delivered into infected host cells and may modulate host gene transcription. Using in vitro models of intestinal cryptosporidiosis, in this study, we analyzed the expression profile of host beta-defensin genes in host cells following infection. We found that C. parvum infection caused a significant downregulation of the DEFB1 gene. Interestingly, downregulation of DEFB1 gene was associated with host delivery of Cdg7_FLc_1000 RNA transcript, a C. parvum RNA that has previously demonstrated to be delivered into the nuclei of infected host cells. Knockdown of Cdg7_FLc_1000 in host cells could attenuate the trans-suppression of host DEFB1 gene and decreased the parasite burden. Therefore, our data suggest that trans-suppression of DEFB1 gene in intestinal epithelial cells following C. parvum infection involves host delivery of parasite Cdg7_FLc_1000 RNA, a process that may be relevant to the epithelial defense evasion by C. parvum at the early stage of infection.

  12. The functions of the variable lipoprotein family of Mycoplasma hyorhinis in adherence to host cells.

    Science.gov (United States)

    Xiong, Qiyan; Wang, Jia; Ji, Yan; Ni, Bo; Zhang, Bixiong; Ma, Qinghong; Wei, Yanna; Xiao, Shaobo; Feng, Zhixin; Liu, Maojun; Shao, Guoqing

    2016-04-15

    Mycoplasma hyorhinis (M. hyorhinis) is a swine pathogen that is associated with various human cancers and contamination in cell cultures. However, no studies on the adhesion molecules of this pathogen have yet been reported. The variable lipoprotein (Vlp) family is an important surface component of M. hyorhinis. Herein, we performed several experiments to identify the function of the Vlp family in adherence to host cells. Seven recombinant Vlp (rVlp) proteins were expressed in Escherichia coli and purified by affinity chromatography. The potential role of rVlp adherence to pig kidney (PK-15) and swine tracheal epithelial (STEC) cells was then studied by indirect immunofluorescence assay and microtiter plate adherence assay. Adhesion of M. hyorhinis to PK-15 and STEC cells was specifically inhibited by the addition of a cocktail of rVlp proteins. The rVlp protein mixture was shown to bind to both PK-15 and STEC cells. The binding increased in a dose-dependent manner and could be blocked by antisera against the rVlp proteins. Most of the rVlp proteins could bind individually to both PK-15 and STEC cells except for rVlpD and rVlpF, which bound only to STEC cells. Because Vlp members vary in size among different strains and generations, they may vary in their cytoadhesion capabilities in various strains. In summary, the present results indicate that the Vlp family functions as adhesins of M. hyorhinis. Copyright © 2016. Published by Elsevier B.V.

  13. Host cell proteins in biotechnology-derived products: A risk assessment framework.

    Science.gov (United States)

    de Zafra, Christina L Zuch; Quarmby, Valerie; Francissen, Kathleen; Vanderlaan, Martin; Zhu-Shimoni, Judith

    2015-11-01

    To manufacture biotechnology products, mammalian or bacterial cells are engineered for the production of recombinant therapeutic human proteins including monoclonal antibodies. Host cells synthesize an entire repertoire of proteins which are essential for their own function and survival. Biotechnology manufacturing processes are designed to produce recombinant therapeutics with a very high degree of purity. While there is typically a low residual level of host cell protein in the final drug product, under some circumstances a host cell protein(s) may copurify with the therapeutic protein and, if it is not detected and removed, it may become an unintended component of the final product. The purpose of this article is to enumerate and discuss factors to be considered in an assessment of risk of residual host cell protein(s) detected and identified in the drug product. The consideration of these factors and their relative ranking will lead to an overall risk assessment that informs decision-making around how to control the levels of host cell proteins. © 2015 Wiley Periodicals, Inc.

  14. Apicomplexans pulling the strings: manipulation of the host cell cytoskeleton dynamics.

    Science.gov (United States)

    Cardoso, Rita; Soares, Helena; Hemphill, Andrew; Leitão, Alexandre

    2016-07-01

    Invasive stages of apicomplexan parasites require a host cell to survive, proliferate and advance to the next life cycle stage. Once invasion is achieved, apicomplexans interact closely with the host cell cytoskeleton, but in many cases the different species have evolved distinct mechanisms and pathways to modulate the structural organization of cytoskeletal filaments. The host cell cytoskeleton is a complex network, largely, but not exclusively, composed of microtubules, actin microfilaments and intermediate filaments, all of which are modulated by associated proteins, and it is involved in diverse functions including maintenance of cell morphology and mechanical support, migration, signal transduction, nutrient uptake, membrane and organelle trafficking and cell division. The ability of apicomplexans to modulate the cytoskeleton to their own advantage is clearly beneficial. We here review different aspects of the interactions of apicomplexans with the three main cytoskeletal filament types, provide information on the currently known parasite effector proteins and respective host cell targets involved, and how these interactions modulate the host cell physiology. Some of these findings could provide novel targets that could be exploited for the development of preventive and/or therapeutic strategies.

  15. Replication and virus-induced transcriptome of HAdV-5 in normal host cells versus cancer cells--differences of relevance for adenoviral oncolysis.

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    Dominik E Dorer

    Full Text Available Adenoviruses (Ads, especially HAdV-5, have been genetically equipped with tumor-restricted replication potential to enable applications in oncolytic cancer therapy. Such oncolytic adenoviruses have been well tolerated in cancer patients, but their anti-tumor efficacy needs to be enhanced. In this regard, it should be considered that cancer cells, dependent on their tissue of origin, can differ substantially from the normal host cells to which Ads are adapted by complex virus-host interactions. Consequently, viral replication efficiency, a key determinant of oncolytic activity, might be suboptimal in cancer cells. Therefore, we have analyzed both the replication kinetics of HAdV-5 and the virus-induced transcriptome in human bronchial epithelial cells (HBEC in comparison to cancer cells. This is the first report on genome-wide expression profiling of Ads in their native host cells. We found that E1A expression and onset of viral genome replication are most rapid in HBEC and considerably delayed in melanoma cells. In squamous cell lung carcinoma cells, we observed intermediate HAdV-5 replication kinetics. Infectious particle production, viral spread and lytic activity of HAdV-5 were attenuated in melanoma cells versus HBEC. Expression profiling at the onset of viral genome replication revealed that HAdV-5 induced the strongest changes in the cellular transcriptome in HBEC, followed by lung cancer and melanoma cells. We identified prominent regulation of genes involved in cell cycle and DNA metabolism, replication and packaging in HBEC, which is in accord with the necessity to induce S phase for viral replication. Strikingly, in melanoma cells HAdV-5 triggered opposing regulation of said genes and, in contrast to lung cancer cells, no weak S phase induction was detected when using the E2F promoter as reporter. Our results provide a rationale for improving oncolytic adenoviruses either by adaptation of viral infection to target tumor cells or by

  16. Global impact of Salmonella type III secretion effector SteA on host cells

    International Nuclear Information System (INIS)

    Cardenal-Muñoz, Elena; Gutiérrez, Gabriel; Ramos-Morales, Francisco

    2014-01-01

    Highlights: • We analyzed HeLa cells transcriptome in response to Salmonella SteA. • Significant differential expression was detected for 58 human genes. • They are involved in ECM organization and regulation of some signaling pathways. • Cell death, cell adhesion and cell migration were decreased in SteA-expressing cells. • These results contribute to understand the role of SteA during infections. - Abstract: Salmonella enterica is a Gram-negative bacterium that causes gastroenteritis, bacteremia and typhoid fever in several animal species including humans. Its virulence is greatly dependent on two type III secretion systems, encoded in pathogenicity islands 1 and 2. These systems translocate proteins called effectors into eukaryotic host cell. Effectors interfere with host signal transduction pathways to allow the internalization of pathogens and their survival and proliferation inside vacuoles. SteA is one of the few Salmonella effectors that are substrates of both type III secretion systems. Here, we used gene arrays and bioinformatics analysis to study the genetic response of human epithelial cells to SteA. We found that constitutive synthesis of SteA in HeLa cells leads to induction of genes related to extracellular matrix organization and regulation of cell proliferation and serine/threonine kinase signaling pathways. SteA also causes repression of genes related to immune processes and regulation of purine nucleotide synthesis and pathway-restricted SMAD protein phosphorylation. In addition, a cell biology approach revealed that epithelial cells expressing steA show altered cell morphology, and decreased cytotoxicity, cell–cell adhesion and migration

  17. Global impact of Salmonella type III secretion effector SteA on host cells

    Energy Technology Data Exchange (ETDEWEB)

    Cardenal-Muñoz, Elena; Gutiérrez, Gabriel; Ramos-Morales, Francisco

    2014-07-11

    Highlights: • We analyzed HeLa cells transcriptome in response to Salmonella SteA. • Significant differential expression was detected for 58 human genes. • They are involved in ECM organization and regulation of some signaling pathways. • Cell death, cell adhesion and cell migration were decreased in SteA-expressing cells. • These results contribute to understand the role of SteA during infections. - Abstract: Salmonella enterica is a Gram-negative bacterium that causes gastroenteritis, bacteremia and typhoid fever in several animal species including humans. Its virulence is greatly dependent on two type III secretion systems, encoded in pathogenicity islands 1 and 2. These systems translocate proteins called effectors into eukaryotic host cell. Effectors interfere with host signal transduction pathways to allow the internalization of pathogens and their survival and proliferation inside vacuoles. SteA is one of the few Salmonella effectors that are substrates of both type III secretion systems. Here, we used gene arrays and bioinformatics analysis to study the genetic response of human epithelial cells to SteA. We found that constitutive synthesis of SteA in HeLa cells leads to induction of genes related to extracellular matrix organization and regulation of cell proliferation and serine/threonine kinase signaling pathways. SteA also causes repression of genes related to immune processes and regulation of purine nucleotide synthesis and pathway-restricted SMAD protein phosphorylation. In addition, a cell biology approach revealed that epithelial cells expressing steA show altered cell morphology, and decreased cytotoxicity, cell–cell adhesion and migration.

  18. Co-ordinate regulation of distinct host cell signalling pathways by multifunctional enteropathogenic Escherichia coli effector molecules.

    Science.gov (United States)

    Kenny, Brendan; Ellis, Sarah; Leard, Alan D; Warawa, Jonathan; Mellor, Harry; Jepson, Mark A

    2002-05-01

    Enteropathogenic Escherichia coli (EPEC) is a major cause of paediatric diarrhoea and a model for the family of attaching and effacing (A/E) pathogens. A/E pathogens encode a type III secretion system to transfer effector proteins into host cells. The EPEC Tir effector protein acts as a receptor for the bacterial surface protein intimin and is involved in the formation of Cdc42-independent, actin-rich pedestal structures beneath the adhered bacteria. In this paper, we demonstrate that EPEC binding to HeLa cells also induces Tir-independent, cytoskeletal rearrangement evidenced by the early, transient formation of filopodia-like structures at sites of infection. Filopodia formation is dependent on expression of the EPEC Map effector molecule - a protein that targets mitochondria and induces their dysfunction. We show that Map-induced filopodia formation is independent of mitochondrial targeting and is abolished by cellular expression of the Cdc42 inhibitory WASP-CRIB domain, demonstrating that Map has at least two distinct functions in host cells. The transient nature of the filopodia is related to an ability of EPEC to downregulate Map-induced cell signalling that, like pedestal formation, was dependent on both Tir and intimin proteins. The ability of Tir to downregulate filopodia was impaired by disrupting a putative GTPase-activating protein (GAP) motif, suggesting that Tir may possess such a function, with its interaction with intimin triggering this activity. Furthermore, we also found that Map-induced cell signalling inhibits pedestal formation, revealing that the cellular effects of Tir and Map must be co-ordinately regulated during infection. Possible implications of the multifunctional nature of EPEC effector molecules in pathogenesis are discussed.

  19. The Arf GTPase-activating protein family is exploited by Salmonella enterica serovar Typhimurium to invade nonphagocytic host cells.

    Science.gov (United States)

    Davidson, Anthony C; Humphreys, Daniel; Brooks, Andrew B E; Hume, Peter J; Koronakis, Vassilis

    2015-02-10

    To establish intracellular infections, Salmonella bacteria trigger host cell membrane ruffling and invasion by subverting cellular Arf guanine nucleotide exchange factors (GEFs) that activate Arf1 and Arf6 GTPases by promoting GTP binding. A family of cellular Arf GTPase-activating proteins (GAPs) can downregulate Arf signaling by stimulating GTP hydrolysis, but whether they do this during infection is unknown. Here, we uncovered a remarkable role for distinct Arf GAP family members in Salmonella invasion. The Arf6 GAPs ACAP1 and ADAP1 and the Arf1 GAP ASAP1 localized at Salmonella-induced ruffles, which was not the case for the plasma membrane-localized Arf6 GAPs ARAP3 and GIT1 or the Golgi-associated Arf1 GAP1. Surprisingly, we found that loss of ACAP1, ADAP1, or ASAP1 impaired Salmonella invasion, revealing that GAPs cannot be considered mere terminators of cytoskeleton remodeling. Salmonella invasion was restored in Arf GAP-depleted cells by expressing fast-cycling Arf derivatives, demonstrating that Arf GTP/GDP cycles facilitate Salmonella invasion. Consistent with this view, both constitutively active and dominant-negative Arf derivatives that cannot undergo GTP/GDP cycles inhibited invasion. Furthermore, we demonstrated that Arf GEFs and GAPs colocalize at invading Salmonella and collaborate to drive Arf1-dependent pathogen invasion. This study revealed that Salmonella bacteria exploit a remarkable interplay between Arf GEFs and GAPs to direct cycles of Arf GTPase activation and inactivation. These cycles drive Salmonella cytoskeleton remodeling and enable intracellular infections. To initiate infections, the Salmonella bacterial pathogen remodels the mammalian actin cytoskeleton and invades host cells by subverting host Arf GEFs that activate Arf1 and Arf6 GTPases. Cellular Arf GAPs deactivate Arf GTPases and negatively regulate cell processes, but whether they target Arfs during infection is unknown. Here, we uncovered an important role for the Arf GAP

  20. Host cell reactivation of uv- and X-ray-damaged herpes simplex virus by Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines

    International Nuclear Information System (INIS)

    Henderson, E.E.; Long, W.K.

    1981-01-01

    The efficacy of using an infected centers assay, employing herpes simplex virus-infected, Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) as components, to study host cell reactivation has been explored. Herpes simplex virus type 1 (HSV-1) was shown through the infected centers assay to have detectable but varying ability to lytically infect LCLs established from chromosomal breakage syndromes or closely related genetic disorders. The rate of HSV inactivation by ultraviolet (uv) irradiation was faster in LCLs established from Cockaynes's syndrome than in normal LCLs, and faster still in LCLs established from xeroderma pigmentosum. These results indicate that Cockayne's syndrome, while having what appears to be quantitatively normal levels of uv-induced DNA repair replication, shows decreased ability to host cell reactivated uv-damaged HSV. In direct contrast, X-irradiated HSV showed identical survival when assayed on normal LCLs or LCLs established from ataxia telangiectasia showing increased sensitivity to X irradiation as measured by colony formation. Through the infected centers assay, it has also been possible to demonstrate low levels of multiplicity reactivation of mutagen-damaged HSV in permanently proliferating LCLs

  1. Potential for transmission of infections in networks of cattle farms

    Directory of Open Access Journals (Sweden)

    V.V. Volkova

    2010-09-01

    Full Text Available The aim of this analysis is to evaluate how generic properties of networks of livestock farms connected by movements of cattle impact on the potential for spread of infectious diseases. We focus on endemic diseases with long infectious periods in affected cattle, such as bovine tuberculosis. Livestock farm networks provide a rare example of large but fully specified directed contact networks, allowing investigations into how properties of such networks impact the potential for spread of infections within them. Here we quantify the latter in terms of the basic reproduction number, R0, and partition the contributions to R0 from first order moments (mean contact rates and second order moments (variances and covariances of contact rates of the farm contact matrices. We find that the second order properties make a substantial contribution to the magnitude of R0, similarly to that reported for other populations. Importantly, however, we find that the magnitude of these effects depends on exactly how the contacts between farms are defined or weighted. We note that the second order properties of a directed contact network may vary through time even with little change in the mean contact rates or in overall connectedness of the network. Keywords: Basic reproduction number, Infectious disease, Heterogeneity, 20–80 rule, Contact network, Bovine tuberculosis

  2. The Vibrio parahaemolyticus Type III Secretion Systems manipulate host cell MAPK for critical steps in pathogenesis.

    LENUS (Irish Health Repository)

    Matlawska-Wasowska, Ksenia

    2010-12-01

    Vibrio parahaemolyticus is a food-borne pathogen causing inflammation of the gastrointestinal epithelium. Pathogenic strains of this bacterium possess two Type III Secretion Systems (TTSS) that deliver effector proteins into host cells. In order to better understand human host cell responses to V. parahaemolyticus, the modulation of Mitogen Activated Protein Kinase (MAPK) activation in epithelial cells by an O3:K6 clinical isolate, RIMD2210633, was investigated. The importance of MAPK activation for the ability of the bacterium to be cytotoxic and to induce secretion of Interleukin-8 (IL-8) was determined.

  3. A Polymorphism within the Internal Fusion Loop of the Ebola Virus Glycoprotein Modulates Host Cell Entry.

    Science.gov (United States)

    Hoffmann, Markus; Crone, Lisa; Dietzel, Erik; Paijo, Jennifer; González-Hernández, Mariana; Nehlmeier, Inga; Kalinke, Ulrich; Becker, Stephan; Pöhlmann, Stefan

    2017-05-01

    The large scale of the Ebola virus disease (EVD) outbreak in West Africa in 2013-2016 raised the question whether the host cell interactions of the responsible Ebola virus (EBOV) strain differed from those of other ebolaviruses. We previously reported that the glycoprotein (GP) of the virus circulating in West Africa in 2014 (EBOV2014) exhibited reduced ability to mediate entry into two nonhuman primate (NHP)-derived cell lines relative to the GP of EBOV1976. Here, we investigated the molecular determinants underlying the differential entry efficiency. We found that EBOV2014-GP-driven entry into diverse NHP-derived cell lines, as well as human monocyte-derived macrophages and dendritic cells, was reduced compared to EBOV1976-GP, although entry into most human- and all bat-derived cell lines tested was comparable. Moreover, EBOV2014 replication in NHP but not human cells was diminished relative to EBOV1976, suggesting that reduced cell entry translated into reduced viral spread. Mutagenic analysis of EBOV2014-GP and EBOV1976-GP revealed that an amino acid polymorphism in the receptor-binding domain, A82V, modulated entry efficiency in a cell line-independent manner and did not account for the reduced EBOV2014-GP-driven entry into NHP cells. In contrast, polymorphism T544I, located in the internal fusion loop in the GP2 subunit, was found to be responsible for the entry phenotype. These results suggest that position 544 is an important determinant of EBOV infectivity for both NHP and certain human target cells. IMPORTANCE The Ebola virus disease outbreak in West Africa in 2013 entailed more than 10,000 deaths. The scale of the outbreak and its dramatic impact on human health raised the question whether the responsible virus was particularly adept at infecting human cells. Our study shows that an amino acid exchange, A82V, that was acquired during the epidemic and that was not observed in previously circulating viruses, increases viral entry into diverse target cells

  4. Which Way In? The RalF Arf-GEF Orchestrates Rickettsia Host Cell Invasion

    Science.gov (United States)

    Rennoll-Bankert, Kristen E.; Rahman, M. Sayeedur; Gillespie, Joseph J.; Guillotte, Mark L.; Kaur, Simran J.; Lehman, Stephanie S.; Beier-Sexton, Magda; Azad, Abdu F.

    2015-01-01

    Bacterial Sec7-domain-containing proteins (RalF) are known only from species of Legionella and Rickettsia, which have facultative and obligate intracellular lifestyles, respectively. L. pneumophila RalF, a type IV secretion system (T4SS) effector, is a guanine nucleotide exchange factor (GEF) of ADP-ribosylation factors (Arfs), activating and recruiting host Arf1 to the Legionella-containing vacuole. In contrast, previous in vitro studies showed R. prowazekii (Typhus Group) RalF is a functional Arf-GEF that localizes to the host plasma membrane and interacts with the actin cytoskeleton via a unique C-terminal domain. As RalF is differentially encoded across Rickettsia species (e.g., pseudogenized in all Spotted Fever Group species), it may function in lineage-specific biology and pathogenicity. Herein, we demonstrate RalF of R. typhi (Typhus Group) interacts with the Rickettsia T4SS coupling protein (RvhD4) via its proximal C-terminal sequence. RalF is expressed early during infection, with its inactivation via antibody blocking significantly reducing R. typhi host cell invasion. For R. typhi and R. felis (Transitional Group), RalF ectopic expression revealed subcellular localization with the host plasma membrane and actin cytoskeleton. Remarkably, R. bellii (Ancestral Group) RalF showed perinuclear localization reminiscent of ectopically expressed Legionella RalF, for which it shares several structural features. For R. typhi, RalF co-localization with Arf6 and PI(4,5)P2 at entry foci on the host plasma membrane was determined to be critical for invasion. Thus, we propose recruitment of PI(4,5)P2 at entry foci, mediated by RalF activation of Arf6, initiates actin remodeling and ultimately facilitates bacterial invasion. Collectively, our characterization of RalF as an invasin suggests that, despite carrying a similar Arf-GEF unknown from other bacteria, different intracellular lifestyles across Rickettsia and Legionella species have driven divergent roles for Ral

  5. The potential for emerging therapeutic options for Clostridium difficile infection

    Science.gov (United States)

    Mathur, Harsh; Rea, Mary C; Cotter, Paul D; Ross, R Paul; Hill, Colin

    2014-01-01

    Clostridium difficile is mainly a nosocomial pathogen and is a significant cause of antibiotic-associated diarrhea. It is also implicated in the majority of cases of pseudomembranous colitis. Recently, advancements in next generation sequencing technology (NGS) have highlighted the extent of damage to the gut microbiota caused by broad-spectrum antibiotics, often resulting in C. difficile infection (CDI). Currently the treatment of choice for CDI involves the use of metronidazole and vancomycin. However, recurrence and relapse of CDI, even after rounds of metronidazole/vancomycin administration is a problem that must be addressed. The efficacy of alternative antibiotics such as fidaxomicin, rifaximin, nitazoxanide, ramoplanin and tigecycline, as well as faecal microbiota transplantation has been assessed and some have yielded positive outcomes against C. difficile. Some bacteriocins have also shown promising effects against C. difficile in recent years. In light of this, the potential for emerging treatment options and efficacy of anti-C. difficile vaccines are discussed in this review. PMID:25564777

  6. Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

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    Bárbara Hissa

    Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

  7. Proteomic analysis of the action of the Mycobacterium ulcerans toxin mycolactone: targeting host cells cytoskeleton and collagen.

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    José B Gama

    2014-08-01

    Full Text Available Buruli ulcer (BU is a neglected tropical disease caused by Mycobacterium ulcerans. The tissue damage characteristic of BU lesions is known to be driven by the secretion of the potent lipidic exotoxin mycolactone. However, the molecular action of mycolactone on host cell biology mediating cytopathogenesis is not fully understood. Here we applied two-dimensional electrophoresis (2-DE to identify the mechanisms of mycolactone's cellular action in the L929 mouse fibroblast proteome. This revealed 20 changed spots corresponding to 18 proteins which were clustered mainly into cytoskeleton-related proteins (Dync1i2, Cfl1, Crmp2, Actg1, Stmn1 and collagen biosynthesis enzymes (Plod1, Plod3, P4ha1. In line with cytoskeleton conformational disarrangements that are observed by immunofluorescence, we found several regulators and constituents of both actin- and tubulin-cytoskeleton affected upon exposure to the toxin, providing a novel molecular basis for the effect of mycolactone. Consistent with these cytoskeleton-related alterations, accumulation of autophagosomes as well as an increased protein ubiquitination were observed in mycolactone-treated cells. In vivo analyses in a BU mouse model revealed mycolactone-dependent structural changes in collagen upon infection with M. ulcerans, associated with the reduction of dermal collagen content, which is in line with our proteomic finding of mycolactone-induced down-regulation of several collagen biosynthesis enzymes. Our results unveil the mechanisms of mycolactone-induced molecular cytopathogenesis on exposed host cells, with the toxin compromising cell structure and homeostasis by inducing cytoskeleton alterations, as well as disrupting tissue structure, by impairing the extracellular matrix biosynthesis.

  8. Proteomic analysis of the action of the Mycobacterium ulcerans toxin mycolactone: targeting host cells cytoskeleton and collagen.

    Science.gov (United States)

    Gama, José B; Ohlmeier, Steffen; Martins, Teresa G; Fraga, Alexandra G; Sampaio-Marques, Belém; Carvalho, Maria A; Proença, Fernanda; Silva, Manuel T; Pedrosa, Jorge; Ludovico, Paula

    2014-08-01

    Buruli ulcer (BU) is a neglected tropical disease caused by Mycobacterium ulcerans. The tissue damage characteristic of BU lesions is known to be driven by the secretion of the potent lipidic exotoxin mycolactone. However, the molecular action of mycolactone on host cell biology mediating cytopathogenesis is not fully understood. Here we applied two-dimensional electrophoresis (2-DE) to identify the mechanisms of mycolactone's cellular action in the L929 mouse fibroblast proteome. This revealed 20 changed spots corresponding to 18 proteins which were clustered mainly into cytoskeleton-related proteins (Dync1i2, Cfl1, Crmp2, Actg1, Stmn1) and collagen biosynthesis enzymes (Plod1, Plod3, P4ha1). In line with cytoskeleton conformational disarrangements that are observed by immunofluorescence, we found several regulators and constituents of both actin- and tubulin-cytoskeleton affected upon exposure to the toxin, providing a novel molecular basis for the effect of mycolactone. Consistent with these cytoskeleton-related alterations, accumulation of autophagosomes as well as an increased protein ubiquitination were observed in mycolactone-treated cells. In vivo analyses in a BU mouse model revealed mycolactone-dependent structural changes in collagen upon infection with M. ulcerans, associated with the reduction of dermal collagen content, which is in line with our proteomic finding of mycolactone-induced down-regulation of several collagen biosynthesis enzymes. Our results unveil the mechanisms of mycolactone-induced molecular cytopathogenesis on exposed host cells, with the toxin compromising cell structure and homeostasis by inducing cytoskeleton alterations, as well as disrupting tissue structure, by impairing the extracellular matrix biosynthesis.

  9. SH3 domain-mediated recruitment of host cell amphiphysins by alphavirus nsP3 promotes viral RNA replication.

    Directory of Open Access Journals (Sweden)

    Maarit Neuvonen

    2011-11-01

    Full Text Available Among the four non-structural proteins of alphaviruses the function of nsP3 is the least well understood. NsP3 is a component of the viral replication complex, and composed of a conserved aminoterminal macro domain implicated in viral RNA synthesis, and a poorly conserved carboxyterminal region. Despite the lack of overall homology we noted a carboxyterminal proline-rich sequence motif shared by many alphaviral nsP3 proteins, and found it to serve as a preferred target site for the Src-homology 3 (SH3 domains of amphiphysin-1 and -2. Nsp3 proteins of Semliki Forest (SFV, Sindbis (SINV, and Chikungunya viruses all showed avid and SH3-dependent binding to amphiphysins. Upon alphavirus infection the intracellular distribution of amphiphysin was dramatically altered and colocalized with nsP3. Mutations in nsP3 disrupting the amphiphysin SH3 binding motif as well as RNAi-mediated silencing of amphiphysin-2 expression resulted in impaired viral RNA replication in HeLa cells infected with SINV or SFV. Infection of Balb/c mice with SFV carrying an SH3 binding-defective nsP3 was associated with significantly decreased mortality. These data establish SH3 domain-mediated binding of nsP3 with amphiphysin as an important host cell interaction promoting alphavirus replication.

  10. Lipids in host-pathogen interactions: pathogens exploit the complexity of the host cell lipidome.

    Science.gov (United States)

    van der Meer-Janssen, Ynske P M; van Galen, Josse; Batenburg, Joseph J; Helms, J Bernd

    2010-01-01

    Lipids were long believed to have a structural role in biomembranes and a role in energy storage utilizing cellular lipid droplets and plasma lipoproteins. Research over the last decades has identified an additional role of lipids in cellular signaling, membrane microdomain organization and dynamics, and membrane trafficking. These properties make lipids an attractive target for pathogens to modulate host cell processes in order to allow their survival and replication. In this review we will summarize the often ingenious strategies of pathogens to modify the lipid homeostasis of host cells, allowing them to divert cellular processes. To this end pathogens take full advantage of the complexity of the lipidome. The examples are categorized in generalized and emerging principles describing the involvement of lipids in host-pathogen interactions. Several pathogens are described that simultaneously induce multiple changes in the host cell signaling and trafficking mechanisms. Elucidation of these pathogen-induced changes may have important implications for drug development. The emergence of high-throughput lipidomic techniques will allow the description of changes of the host cell lipidome at the level of individual molecular lipid species and the identification of lipid biomarkers.

  11. Identification of a Peptide-Pheromone that Enhances Listeria monocytogenes Escape from Host Cell Vacuoles

    Science.gov (United States)

    Xayarath, Bobbi; Alonzo, Francis; Freitag, Nancy E.

    2015-01-01

    Listeria monocytogenes is a Gram-positive facultative intracellular bacterial pathogen that invades mammalian cells and escapes from membrane-bound vacuoles to replicate within the host cell cytosol. Gene products required for intracellular bacterial growth and bacterial spread to adjacent cells are regulated by a transcriptional activator known as PrfA. PrfA becomes activated following L. monocytogenes entry into host cells, however the signal that stimulates PrfA activation has not yet been defined. Here we provide evidence for L. monocytogenes secretion of a small peptide pheromone, pPplA, which enhances the escape of L. monocytogenes from host cell vacuoles and may facilitate PrfA activation. The pPplA pheromone is generated via the proteolytic processing of the PplA lipoprotein secretion signal peptide. While the PplA lipoprotein is dispensable for pathogenesis, bacteria lacking the pPplA pheromone are significantly attenuated for virulence in mice and have a reduced efficiency of bacterial escape from the vacuoles of nonprofessional phagocytic cells. Mutational activation of PrfA restores virulence and eliminates the need for pPplA-dependent signaling. Experimental evidence suggests that the pPplA peptide may help signal to L. monocytogenes its presence within the confines of the host cell vacuole, stimulating the expression of gene products that contribute to vacuole escape and facilitating PrfA activation to promote bacterial growth within the cytosol. PMID:25822753

  12. Mild hypothermic culture conditions affect residual host cell protein composition post-Protein A chromatography.

    Science.gov (United States)

    Goey, Cher Hui; Bell, David; Kontoravdi, Cleo

    2018-04-01

    Host cell proteins (HCPs) are endogenous impurities, and their proteolytic and binding properties can compromise the integrity, and, hence, the stability and efficacy of recombinant therapeutic proteins such as monoclonal antibodies (mAbs). Nonetheless, purification of mAbs currently presents a challenge because they often co-elute with certain HCP species during the capture step of protein A affinity chromatography. A Quality-by-Design (QbD) strategy to overcome this challenge involves identifying residual HCPs and tracing their source to the harvested cell culture fluid (HCCF) and the corresponding cell culture operating parameters. Then, problematic HCPs in HCCF may be reduced by cell engineering or culture process optimization. Here, we present experimental results linking cell culture temperature and post-protein A residual HCP profile. We had previously reported that Chinese hamster ovary cell cultures conducted at standard physiological temperature and with a shift to mild hypothermia on day 5 produced HCCF of comparable product titer and HCP concentration, but with considerably different HCP composition. In this study, we show that differences in HCP variety at harvest cascaded to downstream purification where different residual HCPs were present in the two sets of samples post-protein A purification. To detect low-abundant residual HCPs, we designed a looping liquid chromatography-mass spectrometry method with continuous expansion of a preferred, exclude, and targeted peptide list. Mild hypothermic cultures produced 20% more residual HCP species, especially cell membrane proteins, distinct from the control. Critically, we identified that half of the potentially immunogenic residual HCP species were different between the two sets of samples.

  13. Variation among Staphylococcus aureus membrane vesicle proteomes affects cytotoxicity of host cells.

    Science.gov (United States)

    Jeon, Hyejin; Oh, Man Hwan; Jun, So Hyun; Kim, Seung Il; Choi, Chi Won; Kwon, Hyo Il; Na, Seok Hyeon; Kim, Yoo Jeong; Nicholas, Asiimwe; Selasi, Gati Noble; Lee, Je Chul

    2016-04-01

    Staphylococcus aureus secretes membrane-derived vesicles (MVs), which can deliver virulence factors to host cells and induce cytopathology. However, the cytopathology of host cells induced by MVs derived from different S. aureus strains has not yet been characterized. In the present study, the cytotoxic activity of MVs from different S. aureus isolates on host cells was compared and the proteomes of S. aureus MVs were analyzed. The MVs purified from S. aureus M060 isolated from a patient with staphylococcal scalded skin syndrome showed higher cytotoxic activity toward host cells than that shown by MVs from three other clinical S. aureus isolates. S. aureus M060 MVs induced HEp-2 cell apoptosis in a dose-dependent manner, but the cytotoxic activity of MVs was completely abolished by treatment with proteinase K. In a proteomic analysis, the MVs from three S. aureus isolates not only carry 25 common proteins, but also carry ≥60 strain-specific proteins. All S. aureus MVs contained δ-hemolysin (Hld), γ-hemolysin, leukocidin D, and exfoliative toxin C, but exfoliative toxin A (ETA) was specifically identified in S. aureus M060 MVs. ETA was delivered to HEp-2 cells via S. aureus MVs. Both rETA and rHld induced cytotoxicity in HEp-2 cells. In conclusion, MVs from clinical S. aureus isolates differ with respect to cytotoxic activity in host cells, and these differences may result from differences in the MV proteomes. Further proteogenomic analysis or mutagenesis of specific genes is necessary to identify cytotoxic factors in S. aureus MVs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. MYR1-Dependent Effectors Are the Major Drivers of a Host Cell's Early Response to Toxoplasma, Including Counteracting MYR1-Independent Effects.

    Science.gov (United States)

    Naor, Adit; Panas, Michael W; Marino, Nicole; Coffey, Michael J; Tonkin, Christopher J; Boothroyd, John C

    2018-04-03

    The obligate intracellular parasite Toxoplasma gondii controls its host cell from within the parasitophorous vacuole (PV) by using a number of diverse effector proteins, a subset of which require the aspartyl protease 5 enzyme (ASP5) and/or the recently discovered MYR1 protein to cross the PV membrane. To examine the impact these effectors have in the context of the entirety of the host response to Toxoplasma , we used RNA-Seq to analyze the transcriptome expression profiles of human foreskin fibroblasts infected with wild-type RH (RH-WT), RHΔ myr1 , and RHΔ asp5 tachyzoites. Interestingly, the majority of the differentially regulated genes responding to Toxoplasma infection are MYR1 dependent. A subset of MYR1 responses were ASP5 independent, and MYR1 function did not require ASP5 cleavage, suggesting the export of some effectors requires only MYR1. Gene set enrichment analysis of MYR1-dependent host responses suggests an upregulation of E2F transcription factors and the cell cycle and a downregulation related to interferon signaling, among numerous others. Most surprisingly, "hidden" responses arising in RHΔ myr1 - but not RH-WT-infected host cells indicate counterbalancing actions of MYR1-dependent and -independent activities. The host genes and gene sets revealed here to be MYR1 dependent provide new insight into the parasite's ability to co-opt host cell functions. IMPORTANCE Toxoplasma gondii is unique in its ability to successfully invade and replicate in a broad range of host species and cells within those hosts. The complex interplay of effector proteins exported by Toxoplasma is key to its success in co-opting the host cell to create a favorable replicative niche. Here we show that a majority of the transcriptomic effects in tachyzoite-infected cells depend on the activity of a novel translocation system involving MYR1 and that the effectors delivered by this system are part of an intricate interplay of activators and suppressors. Removal of all MYR1

  15. Impact of 4 Lactobacillus plantarum capsular polysaccharide clusters on surface glycan composition and host cell signaling

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    Remus Daniela M

    2012-11-01

    -J produced decreased levels of surface polysaccharides, whereas the molar mass and the composition of polysaccharides was not affected by these cluster mutations. In the quadruple mutant, the amount of surface polysaccharides was strongly reduced. The impact of the cps cluster mutations on toll-like receptor (TLR-mediated human nuclear factor (NF-κB activation in host cells was evaluated using a TLR2 reporter cell line. In comparison to a L. plantarum wild-type derivative, TLR2 activation remained unaffected by the Δcps1A-I and Δcps3A-J mutants but appeared slightly increased after stimulation with the Δcps2A-J and Δcps4A-J mutants, while the Δcps1A-3J and Δcps1A-3J, Δcps4A-J mutants elicited the strongest responses and clearly displayed enhanced TLR2 signaling. Conclusions Our study reveals that modulation of surface glycan characteristics in L. plantarum highlights the role of these molecules in shielding of cell envelope embedded host receptor ligands. Although the apparently complete cps clusters (cps2A-J and cps4A-J contributed individually to this shielding, the removal of all cps clusters led to the strongest signaling enhancement. Our findings provide new insights into cell surface glycan biosynthesis in L. plantarum, which bears relevance in the context of host-cell signaling by probiotic bacteria.

  16. Established and potential risk factors for clostridum difficile infection

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    Vaishnavi C

    2009-01-01

    Full Text Available Clostridium difficile is the aetiological agent for almost all cases of pseudo membranous colitis and 15-25% of antibiotic associated diarrhoea. In recent years, C. difficile associated disease (CDAD has been increasing in frequency and severity due to the emergence of virulent strains. Severe cases of toxic mega colon may be associated with mortality rates of 24-38%. The prevalence of CDAD is global and the incidence varies considerably from place to place. In the initial stages of its discovery, C. difficile infection was regarded mainly as an outcome of antibiotic intake and not as a life threatening disease. Intervention by man has produced conditions making C. difficile a significant cause of morbidity and mortality. The recent outbreak of CDAD in Quebec has sent the alarm bells ringing. Apart from a threefold increase in the incidence of CDAD, clinicians have also reported a higher number of cases involving toxic mega colon, colectomy or death. Among all the risk factors, inclusive of the host and the environmental factors, antibiotics are the most important ones. Surgical patients comprise 55-75% of all patients with CDAD due to the fact that perioperative prophylaxis requires the use of antibiotics. However, other drugs such as immunosuppressants and proton pump inhibitors are also important risk factors. Thus CDAD is a growing nosocomial and public health challenge. Additionally, the recognition of community acquired CDAD signals the presence of several risk factors. In this review, the established and potential risk factors of CDAD, along with the epidemiology, diagnostic modalities, management and preventive measures of the disease have been elaborated.

  17. Interactions of Prototype Foamy Virus Capsids with Host Cell Polo-Like Kinases Are Important for Efficient Viral DNA Integration.

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    Irena Zurnic

    2016-08-01

    Full Text Available Unlike for other retroviruses, only a few host cell factors that aid the replication of foamy viruses (FVs via interaction with viral structural components are known. Using a yeast-two-hybrid (Y2H screen with prototype FV (PFV Gag protein as bait we identified human polo-like kinase 2 (hPLK2, a member of cell cycle regulatory kinases, as a new interactor of PFV capsids. Further Y2H studies confirmed interaction of PFV Gag with several PLKs of both human and rat origin. A consensus Ser-Thr/Ser-Pro (S-T/S-P motif in Gag, which is conserved among primate FVs and phosphorylated in PFV virions, was essential for recognition by PLKs. In the case of rat PLK2, functional kinase and polo-box domains were required for interaction with PFV Gag. Fluorescently-tagged PFV Gag, through its chromatin tethering function, selectively relocalized ectopically expressed eGFP-tagged PLK proteins to mitotic chromosomes in a Gag STP motif-dependent manner, confirming a specific and dominant nature of the Gag-PLK interaction in mammalian cells. The functional relevance of the Gag-PLK interaction was examined in the context of replication-competent FVs and single-round PFV vectors. Although STP motif mutated viruses displayed wild type (wt particle release, RNA packaging and intra-particle reverse transcription, their replication capacity was decreased 3-fold in single-cycle infections, and up to 20-fold in spreading infections over an extended time period. Strikingly similar defects were observed when cells infected with single-round wt Gag PFV vectors were treated with a pan PLK inhibitor. Analysis of entry kinetics of the mutant viruses indicated a post-fusion defect resulting in delayed and reduced integration, which was accompanied with an enhanced preference to integrate into heterochromatin. We conclude that interaction between PFV Gag and cellular PLK proteins is important for early replication steps of PFV within host cells.

  18. Reduced host cell invasiveness and oxidative stress tolerance in double and triple csp gene family deletion mutants of Listeria monocytogenes.

    Science.gov (United States)

    Loepfe, Chantal; Raimann, Eveline; Stephan, Roger; Tasara, Taurai

    2010-07-01

    The cold shock protein (Csp) family comprises small, highly conserved proteins that bind nucleic acids to modulate various bacterial gene expressions. In addition to cold adaptation functions, this group of proteins is thought to facilitate various cellular processes to promote normal growth and stress adaptation responses. Three proteins making up the Listeria monocytogenes Csp family (CspA, CspB, and CspD) promote both cold and osmotic stress adaptation functions in this bacterium. The contribution of these three Csps in the host cell invasion processes of L. monocytogenes was investigated based on human Caco-2 and murine macrophage in vitro cell infection models. The DeltacspB, DeltacspD, DeltacspAB, DeltacspAD, DeltacspBD, and DeltacspABD strains were all significantly impaired in Caco-2 cell invasion compared with the wild-type strain, whereas in the murine macrophage infection assay only, the double (DeltacspBD) and triple (DeltacspABD) csp mutants were also significantly impaired in cell invasion compared with the wild-type strain. The DeltacspBD and DeltacspABD mutants displayed the most severely impaired invasion phenotypes. The invasion ability of these two mutant strains was also further analyzed using cold-stress-exposed organisms. In both cell infection models a significant reduction in invasiveness was observed after cold stress exposure of Listeria organisms. The negative impact of cold stress on subsequent cell invasion ability was, however, more severe in cold-sensitive csp mutants (DeltacspBD and DeltacspABD) compared with the wild type. The impaired macrophage invasion and intracellular growth of DeltacspBD and DeltacspABD also led us to examine oxidative stress resistance capacity in these two mutant strains. Both strains also displayed higher oxidative stress sensitivity relative to the wild-type strain. Our data indicate that besides cold and osmotic stress adaptation roles, Csp family proteins also promote efficient host cell invasion and

  19. Detection of Sleeping Beauty transposition in the genome of host cells by non-radioactive Southern blot analysis

    Energy Technology Data Exchange (ETDEWEB)

    Aravalli, Rajagopal N., E-mail: aravalli@umn.edu [Department of Radiology, University of Minnesota Medical School, MMC 292, 420 Delaware Street SE, Minneapolis, MN 55455 (United States); Park, Chang W. [Department of Medicine, University of Minnesota Medical School, MMC 36, 420 Delaware Street SE, Minneapolis, MN 55455 (United States); Steer, Clifford J., E-mail: steer001@umn.edu [Department of Medicine, University of Minnesota Medical School, MMC 36, 420 Delaware Street SE, Minneapolis, MN 55455 (United States); Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455 (United States)

    2016-08-26

    The Sleeping Beauty transposon (SB-Tn) system is being used widely as a DNA vector for the delivery of therapeutic transgenes, as well as a tool for the insertional mutagenesis in animal models. In order to accurately assess the insertional potential and properties related to the integration of SB it is essential to determine the copy number of SB-Tn in the host genome. Recently developed SB100X transposase has demonstrated an integration rate that was much higher than the original SB10 and that of other versions of hyperactive SB transposases, such as HSB3 or HSB17. In this study, we have constructed a series of SB vectors carrying either a DsRed or a human β-globin transgene that was encompassed by cHS4 insulator elements, and containing the SB100X transposase gene outside the SB-Tn unit within the same vector in cis configuration. These SB-Tn constructs were introduced into the K-562 erythroid cell line, and their presence in the genomes of host cells was analyzed by Southern blot analysis using non-radioactive probes. Many copies of SB-Tn insertions were detected in host cells regardless of transgene sequences or the presence of cHS4 insulator elements. Interestingly, the size difference of 2.4 kb between insulated SB and non-insulated controls did not reflect the proportional difference in copy numbers of inserted SB-Tns. We then attempted methylation-sensitive Southern blots to assess the potential influence of cHS4 insulator elements on the epigenetic modification of SB-Tn. Our results indicated that SB100X was able to integrate at multiple sites with the number of SB-Tn copies larger than 6 kb in size. In addition, the non-radioactive Southern blot protocols developed here will be useful to detect integrated SB-Tn copies in any mammalian cell type.

  20. Detection of Sleeping Beauty transposition in the genome of host cells by non-radioactive Southern blot analysis

    International Nuclear Information System (INIS)

    Aravalli, Rajagopal N.; Park, Chang W.; Steer, Clifford J.

    2016-01-01

    The Sleeping Beauty transposon (SB-Tn) system is being used widely as a DNA vector for the delivery of therapeutic transgenes, as well as a tool for the insertional mutagenesis in animal models. In order to accurately assess the insertional potential and properties related to the integration of SB it is essential to determine the copy number of SB-Tn in the host genome. Recently developed SB100X transposase has demonstrated an integration rate that was much higher than the original SB10 and that of other versions of hyperactive SB transposases, such as HSB3 or HSB17. In this study, we have constructed a series of SB vectors carrying either a DsRed or a human β-globin transgene that was encompassed by cHS4 insulator elements, and containing the SB100X transposase gene outside the SB-Tn unit within the same vector in cis configuration. These SB-Tn constructs were introduced into the K-562 erythroid cell line, and their presence in the genomes of host cells was analyzed by Southern blot analysis using non-radioactive probes. Many copies of SB-Tn insertions were detected in host cells regardless of transgene sequences or the presence of cHS4 insulator elements. Interestingly, the size difference of 2.4 kb between insulated SB and non-insulated controls did not reflect the proportional difference in copy numbers of inserted SB-Tns. We then attempted methylation-sensitive Southern blots to assess the potential influence of cHS4 insulator elements on the epigenetic modification of SB-Tn. Our results indicated that SB100X was able to integrate at multiple sites with the number of SB-Tn copies larger than 6 kb in size. In addition, the non-radioactive Southern blot protocols developed here will be useful to detect integrated SB-Tn copies in any mammalian cell type.

  1. Impact of sub-inhibitory antibiotics on fibronectin-mediated host cell adhesion and invasion by Staphylococcus aureus

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    Rasigade Jean

    2011-12-01

    Full Text Available Abstract Background Staphylococcus aureus is a well-armed pathogen prevalent in severe infections such as endocarditis and osteomyelitis. Fibronectin-binding proteins A and B, encoded by fnbA/B, are major pathogenesis determinants in these infections through their involvement in S. aureus adhesion to and invasion of host cells. Sub-minimum inhibitory concentrations (sub-MICs of antibiotics, frequently occurring in vivo because of impaired drug diffusion at the infection site, can alter S. aureus phenotype. We therefore investigated their impact on S. aureus fibronectin-mediated adhesiveness and invasiveness. Methods After in vitro challenge of S. aureus 8325-4 and clinical isolates with sub-MICs of major anti-staphylococcal agents, we explored fnbA/B transcription levels, bacterial adhesiveness to immobilised human fibronectin and human osteoblasts in culture, and bacterial invasion of human osteoblasts. Results Oxacillin, moxifloxacin and linezolid led to the development of a hyper-adhesive phenotype in the fibronectin adhesion assay that was consistent with an increase in fnbA/B transcription. Conversely, rifampin treatment decreased fibronectin binding in all strains tested without affecting fnbA/B transcription. Gentamicin and vancomycin had no impact on fibronectin binding or fnbA/B transcription levels. Only oxacillin-treated S. aureus displayed a significantly increased adhesion to cultured osteoblasts, but its invasiveness did not differ from that of untreated controls. Conclusion Our findings demonstrate that several antibiotics at sub-MICs modulate fibronectin binding in S. aureus in a drug-specific fashion. However, hyper- and hypo- adhesive phenotypes observed in controlled in vitro conditions were not fully confirmed in whole cell infection assays. The relevance of adhesion modulation during in vivo infections is thus still uncertain and requires further investigations.

  2. Human Immunodeficiency Virus Type 1 Nef protein modulates the lipid composition of virions and host cell membrane microdomains

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    Geyer Matthias

    2007-10-01

    Full Text Available Abstract Background The Nef protein of Human Immunodeficiency Viruses optimizes viral spread in the infected host by manipulating cellular transport and signal transduction machineries. Nef also boosts the infectivity of HIV particles by an unknown mechanism. Recent studies suggested a correlation between the association of Nef with lipid raft microdomains and its positive effects on virion infectivity. Furthermore, the lipidome analysis of HIV-1 particles revealed a marked enrichment of classical raft lipids and thus identified HIV-1 virions as an example for naturally occurring membrane microdomains. Since Nef modulates the protein composition and function of membrane microdomains we tested here if Nef also has the propensity to alter microdomain lipid composition. Results Quantitative mass spectrometric lipidome analysis of highly purified HIV-1 particles revealed that the presence of Nef during virus production from T lymphocytes enforced their raft character via a significant reduction of polyunsaturated phosphatidylcholine species and a specific enrichment of sphingomyelin. In contrast, Nef did not significantly affect virion levels of phosphoglycerolipids or cholesterol. The observed alterations in virion lipid composition were insufficient to mediate Nef's effect on particle infectivity and Nef augmented virion infectivity independently of whether virus entry was targeted to or excluded from membrane microdomains. However, altered lipid compositions similar to those observed in virions were also detected in detergent-resistant membrane preparations of virus producing cells. Conclusion Nef alters not only the proteome but also the lipid composition of host cell microdomains. This novel activity represents a previously unrecognized mechanism by which Nef could manipulate HIV-1 target cells to facilitate virus propagation in vivo.

  3. Prebiotic Oligosaccharides Potentiate Host Protective Responses against L. Monocytogenes Infection

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    Poyin Chen

    2017-12-01

    Full Text Available Prebiotic oligosaccharides are used to modulate enteric pathogens and reduce pathogen shedding. The interactions with prebiotics that alter Listeria monocytogenes infection are not yet clearly delineated. L. monocytogenes cellular invasion requires a concerted manipulation of host epithelial cell membrane receptors to initiate internalization and infection often via receptor glycosylation. Bacterial interactions with host glycans are intimately involved in modulating cellular responses through signaling cascades at the membrane and in intracellular compartments. Characterizing the mechanisms underpinning these modulations is essential for predictive use of dietary prebiotics to diminish pathogen association. We demonstrated that human milk oligosaccharide (HMO pretreatment of colonic epithelial cells (Caco-2 led to a 50% decrease in Listeria association, while Biomos pretreatment increased host association by 150%. L. monocytogenes-induced gene expression changes due to oligosaccharide pretreatment revealed global alterations in host signaling pathways that resulted in differential subcellular localization of L. monocytogenes during early infection. Ultimately, HMO pretreatment led to bacterial clearance in Caco-2 cells via induction of the unfolded protein response and eIF2 signaling, while Biomos pretreatment resulted in the induction of host autophagy and L. monocytogenes vacuolar escape earlier in the infection progression. This study demonstrates the capacity of prebiotic oligosaccharides to minimize infection through induction of host-intrinsic protective responses.

  4. Infection prevention workforce: Potential benefits to educational diversity.

    Science.gov (United States)

    Reese, Sara M; Gilmartin, Heather M

    2017-06-01

    Nurses have historically occupied the infection preventionist (IP) role. As the knowledge and skills needed to advance the field expand, professionals from public health and the laboratory sciences have become IPs. Our study describes the characteristics of current IPs and assesses for relationships between background, certification, experience, and type of work performed. The data were drawn from an existing dataset collected in the conduct of the Association for Professionals in Infection Control and Epidemiology (APIC) MegaSurvey. Descriptive statistics were computed. Associations were calculated using χ 2 or Cochran-Mantel-Haenszel tests. Characteristics of IPs were stratified by work-related activities to allow for comparisons between groups. Of the 13,050 active APIC members, 4,079 participated in the survey (31% response rate). The primary job activity for nurses (97.9%; n = 2,434) was preventing and controlling the transmission of infectious agents or health care-associated infections, for laboratory scientists (97.5%; n = 307) it was the interpretation of surveillance data, and for public health professionals (96.1%; n = 136) it was management and communication: feedback. Infection control departments would benefit from hiring IPs with diverse education and training to address the expanding roles and responsibilities of IPs. This may facilitate the implementation of novel and innovative processes that will impact patient care. Copyright © 2017 Association for Professionals in Infection Control and Epidemiology, Inc. All rights reserved.

  5. Strategies for targeting tetraspanin proteins: potential therapeutic applications in microbial infections.

    Science.gov (United States)

    Hassuna, Noha; Monk, Peter N; Moseley, Gregory W; Partridge, Lynda J

    2009-01-01

    possibilities of targeting tetraspanins for the treatment of infectious disease, using specific antibodies, recombinant EC2 domains, small-molecule mimetics, and small interfering RNA. Such therapies, directed at host-cell molecules, may provide alternative options for combating fast-mutating or newly emerging pathogens, where conventional approaches face difficulties.

  6. Current Animal Models of Postoperative Spine Infection and Potential Future Advances

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    Alexandra eStavrakis

    2015-05-01

    Full Text Available Implant related infection following spine surgery is a devastating complication for patients and can potentially lead to significant neurological compromise, disability, morbidity, and even mortality. This paper provides an overview of the existing animal models of postoperative spine infection and highlights the strengths and weaknesses of each model. In addition there is discussion regarding potential modifications to these animal models to better evaluate preventative and treatment strategies for this challenging complication. Current models are effective in simulating surgical procedures but fail to evaluate infection longitudinally using multiple techniques. Potential future modifications to these models include using advanced imaging technologies to evaluate infection, use of bioluminescent bacterial species, and testing of novel treatment strategies against multiple bacterial strains. There is potential to establish a postoperative spine infection model using smaller animals, such as mice, as these would be a more cost-effective screening tool for potential therapeutic interventions.

  7. Predicting the subcellular localization of viral proteins within a mammalian host cell

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    Thomas DY

    2006-04-01

    Full Text Available Abstract Background The bioinformatic prediction of protein subcellular localization has been extensively studied for prokaryotic and eukaryotic organisms. However, this is not the case for viruses whose proteins are often involved in extensive interactions at various subcellular localizations with host proteins. Results Here, we investigate the extent of utilization of human cellular localization mechanisms by viral proteins and we demonstrate that appropriate eukaryotic subcellular localization predictors can be used to predict viral protein localization within the host cell. Conclusion Such predictions provide a method to rapidly annotate viral proteomes with subcellular localization information. They are likely to have widespread applications both in the study of the functions of viral proteins in the host cell and in the design of antiviral drugs.

  8. Cycle Inhibiting Factors (Cifs: Cyclomodulins That Usurp the Ubiquitin-Dependent Degradation Pathway of Host Cells

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    Eric Oswald

    2011-03-01

    Full Text Available Cycle inhibiting factors (Cifs are type III secreted effectors produced by diverse pathogenic bacteria. Cifs are “cyclomodulins” that inhibit the eukaryotic host cell cycle and also hijack other key cellular processes such as those controlling the actin network and apoptosis. This review summarizes current knowledge on Cif since its first characterization in enteropathogenic Escherichia coli, the identification of several xenologues in distant pathogenic bacteria, to its structure elucidation and the recent deciphering of its mode of action. Cif impairs the host ubiquitin proteasome system through deamidation of ubiquitin or the ubiquitin-like protein NEDD8 that regulates Cullin-Ring-ubiquitin Ligase (CRL complexes. The hijacking of the ubiquitin-dependent degradation pathway of host cells results in the modulation of various cellular functions such as epithelium renewal, apoptosis and immune response. Cif is therefore a powerful weapon in the continuous arm race that characterizes host-bacteria interactions.

  9. Viral infections as potential triggers of type 1 diabetes

    NARCIS (Netherlands)

    van der Werf, Nienke; Kroese, Frans G. M.; Rozing, Jan; Hillebrands, Jan-Luuk

    During the last decades, the incidence of type 1 diabetes (T1D) has increased significantly, reaching percentages of 3% annually worldwide. This increase suggests that besides genetical factors environmental perturbations (including viral infections) are also involved in the pathogenesis of T1D. T1D

  10. Transmission potential of Zika virus infection in the South Pacific

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    Hiroshi Nishiura

    2016-04-01

    Conclusions: The transmissibility of Zika virus infection appears to be comparable to those of dengue and chikungunya viruses. Considering that Aedes species are a shared vector, this finding indicates that Zika virus replication within the vector is perhaps comparable to dengue and chikungunya.

  11. Industrial production of clotting factors: Challenges of expression, and choice of host cells.

    Science.gov (United States)

    Kumar, Sampath R

    2015-07-01

    The development of recombinant forms of blood coagulation factors as safer alternatives to plasma derived factors marked a major advance in the treatment of common coagulation disorders. These are complex proteins, mostly enzymes or co-enzymes, involving multiple post-translational modifications, and therefore are difficult to express. This article reviews the nature of the expression challenges for the industrial production of these factors, vis-à-vis the translational and post-translational bottlenecks, as well as the choice of host cell lines for high-fidelity production. For achieving high productivities of vitamin K dependent proteins, which include factors II (prothrombin), VII, IX and X, and protein C, host cell limitation of γ-glutamyl carboxylation is a major bottleneck. Despite progress in addressing this, involvement of yet unidentified protein(s) impedes a complete cell engineering solution. Human factor VIII expresses at very low levels due to limitations at several steps in the protein secretion pathway. Protein and cell engineering, vector improvement and alternate host cells promise improvement in the productivity. Production of Von Willebrand factor is constrained by its large size, complex structure, and the need for extensive glycosylation and disulfide-bonded oligomerization. All the licensed therapeutic factors are produced in CHO, BHK or HEK293 cells. While HEK293 is a recent adoption, BHK cells appear to be disfavored. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Manipulation of the Host Cell Membrane during Plasmodium Liver Stage Egress

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    Paul-Christian Burda

    2017-04-01

    Full Text Available A crucial step in the life cycle of Plasmodium parasites is the transition from the liver stage to the blood stage. Hepatocyte-derived merozoites reach the blood vessels of the liver inside host cell-derived vesicles called merosomes. The molecular basis of merosome formation is only partially understood. Here we show that Plasmodium berghei liver stage merozoites, upon rupture of the parasitophorous vacuole membrane, destabilize the host cell membrane (HCM and induce separation of the host cell actin cytoskeleton from the HCM. At the same time, the phospholipid and protein composition of the HCM appears to be substantially altered. This includes the loss of a phosphatidylinositol 4,5-bisphosphate (PIP2 reporter and the PIP2-dependent actin-plasma membrane linker ezrin from the HCM. Furthermore, transmembrane domain-containing proteins and palmitoylated and myristoylated proteins, as well as glycosylphosphatidylinositol-anchored proteins, lose their HCM localization. Collectively, these findings provide an explanation of HCM destabilization during Plasmodium liver stage egress and thereby contribute to our understanding of the molecular mechanisms that lead to merosome formation.

  13. Effect of endocytosis inhibitors on Coxiella burnetii interaction with host cells

    International Nuclear Information System (INIS)

    Tujulin, E.; Macellaro, A.; Norlander, L.; Liliehoeoek, B.

    1998-01-01

    The obligate intracellular rickettsia Coxiella burnetii has previously been reported to reach the intra-vacuolar compartment of host cells by phagocytosis. With the aim to further examine the mechanisms of C. burnetii internalisation, macrophage monolayers were treated with well characterised inhibitors of endocytosis. The treatment with two general inhibitors, colchicine and methylamine, resulted in a pronounced dose-dependent decrease of radiolabelled phase II rickettsiae retained from the intracellular fraction. A third inhibitor used, amiloride, has been reported to reduce effectively clathrin-independent pinocytic pathways. The internalisation of C. burnetii was shown to be substantially reduced also by amiloride and the effect was dependent on its concentration. The passive role of C. burnetii in the internalisation was verified by using heat-killed C. burnetii. Host cells treated with either of the three inhibitors (amiloride, colchicine and methylamine) showed a similar reduction of intracellular C. burnetii after exposure to killed as weal as live organisms. The data presented indicate that different endocytic mechanisms, pinocytosis as well as phagocytosis, may mediate the uptake of C. burnetii by a host cell. Key words: Coxiella burnetii; internalisation; endocytosis (authors)

  14. An in silico pipeline to filter the Toxoplasma gondii proteome for proteins that could traffic to the host cell nucleus and influence host cell epigenetic regulation.

    Science.gov (United States)

    Syn, Genevieve; Blackwell, Jenefer M; Jamieson, Sarra E; Francis, Richard W

    2018-01-01

    Toxoplasma gondii uses epigenetic mechanisms to regulate both endogenous and host cell gene expression. To identify genes with putative epigenetic functions, we developed an in silico pipeline to interrogate the T. gondii proteome of 8313 proteins. Step 1 employs PredictNLS and NucPred to identify genes predicted to target eukaryotic nuclei. Step 2 uses GOLink to identify proteins of epigenetic function based on Gene Ontology terms. This resulted in 611 putative nuclear localised proteins with predicted epigenetic functions. Step 3 filtered for secretory proteins using SignalP, SecretomeP, and experimental data. This identified 57 of the 611 putative epigenetic proteins as likely to be secreted. The pipeline is freely available online, uses open access tools and software with user-friendly Perl scripts to automate and manage the results, and is readily adaptable to undertake any such in silico search for genes contributing to particular functions.

  15. Potentiation of zinc stress caused by parastic infection of snails

    Energy Technology Data Exchange (ETDEWEB)

    Guth, D.J. (Univ. of Michigan, Flint); Blankespoor, H.D.; Cairns, J. Jr.

    1977-09-08

    The purpose of this experiment was to determine the effects of parasitism (Schistosomatium douthitti Price and Trichobilharzia sp.) on the tolerance of snails Lymnaea stagnalis (L.) to acutely lethal concentrations of zinc. Significant reduction in tolerance occurred for snails with patent infections at 24 and 75 ppM of Zn/sup + +/. At two selected prepatent levels of parasite development, significant differences occurred at the higher concentration only.

  16. Prevalence of Hookworm infection and Strongyloidiasis in Cats and Potential Risk Factor of Human Diseases

    Science.gov (United States)

    Sedionoto, Blego; Anamnart, Witthaya

    2018-02-01

    Hookworm infection and Stronyloidiasis are public health problem in the worldwide which both of them could infective in human by penetrated on skin and they have potential risk from Gastrointestinal zoonotic helminths of pets, including cats. We investigated the prevalence soil transmitted helminths infection in human and cats used modified Formal-Ether Concentration and agar plate culture. Fecal samples of 23 cats and human from Naitung and Subua Villages (area study 1), and fecal samples of 15 cats and 17 humans from Thasala Beach villages (area study 2) were collected. Result of study in area study 1 showed prevalence of infection in human was not hookworm and strongyloidiasis but 10% humans have infected Ascaris and Tricuris, and in cats have infected by hookworm 75.2% and S. strercoralis 8.5%, toxocara 13%, spirometra 13% and overall prevalence 82.5%. In area study 2 showed in human has infected by Trichuris 100% and S. stercoralis 29.4% and in cats have infected by hookworm 100% and S. strercoralis 40%, toxocora 20%, and spirometra 20%. Helminth infection found in both humans in two areas study are S. strercoralis. Hookworms were the most common helminth in cats but did not connection with infection in human, while S. strercoralis was helminth infection in cats which has potential zoonotic disease to human.

  17. Rift Valley fever potential mosquito vectors and their infection status ...

    African Journals Online (AJOL)

    important entomological features for the identification of potential high risk areas for RVF occurrence, which can provide guidance in the design of appropriate prevention and control measures. The findings of this study have shown that the abundance and diversity of potential. RVF mosquito vectors vary between the study ...

  18. Potential influence of plant chemicals on infectivity of Batrachochytrium dendrobatidis.

    Science.gov (United States)

    Davidson, Elizabeth W; Larsen, Andrew; Meins Palmer, Crystal

    2012-11-08

    We explored whether extracts of trees frequently found associated with amphibian habitats in Australia and Arizona, USA, may be inhibitory to the fungal pathogen Batrachochytrium dendrobatidis (Bd), which has been associated with global amphibian declines. We used salamanders Ambystoma tigrinum as the model system. Salamanders acquired significantly lower loads of Bd when exposed on leaves and extracts from the river red gum Eucalyptus camaldulensis, and loads were also low in some animals exposed on extracts of 2 oak species, Quercus emoryi and Q. turbinella. Some previously infected salamanders had their pathogen loads reduced, and some were fully cured, by placing them in leaf extracts, although some animals also self cured when housed in water alone. A significant number of animals cured of Bd infections 6 mo earlier were found to be resistant to reinfection. These results suggest that plants associated with amphibian habitats should be taken into consideration when explaining the prevalence of Bd in these habitats and that some amphibians may acquire resistance to the fungus if previously cured.

  19. Synthesis of 3-O-sulfonated heparan sulfate octasaccharides that inhibit the herpes simplex virus type 1 host-cell interaction

    Science.gov (United States)

    Hu, Yu-Peng; Lin, Shu-Yi; Huang, Cheng-Yen; Zulueta, Medel Manuel L.; Liu, Jing-Yuan; Chang, Wen; Hung, Shang-Cheng

    2011-07-01

    Cell surface carbohydrates play significant roles in a number of biologically important processes. Heparan sulfate, for instance, is a ubiquitously distributed polysulfated polysaccharide that is involved, among other things, in the initial step of herpes simplex virus type 1 (HSV-1) infection. The virus interacts with cell-surface heparan sulfate to facilitate host-cell attachment and entry. 3-O-Sulfonated heparan sulfate has been found to function as an HSV-1 entry receptor. Achieving a complete understanding of these interactions requires the chemical synthesis of such oligosaccharides, but this remains challenging. Here, we present a convenient approach for the synthesis of two irregular 3-O-sulfonated heparan sulfate octasaccharides, making use of a key disaccharide intermediate to acquire different building blocks for the oligosaccharide chain assembly. Despite substantial structural differences, the prepared 3-O-sulfonated sugars blocked viral infection in a dosage-dependent manner with remarkable similarity to one another.

  20. Comparative studies of host-cell reactivation, cellular capacity and enhanced reactivation of herpes simplex virus in normal, xeroderma pigmentosum and Cockayne syndrome fibroblasts

    International Nuclear Information System (INIS)

    Ryan, D.K.G.; Rainbow, A.J.; McMaster Univ., Hamilton, Ontario

    1986-01-01

    Host-cell reactivation (HCR) of UV-irradiated herpes simplex virus type 2 (HSV-2), capacity of UV-irradiated cells to support HSV-2 plaque formation and UV-enhanced reactivation (UVER) of UV-irradiated HSV-2 were examined in fibroblasts from 4 patients with Cockayne syndrome (CS), 5 with xeroderma pigmentosum and 5 normals. The results indicate that delayed capacity for HSV-2 plaque formation is a more sensitive assay than HCR in the detection of cellular DNA-repair deficiency for XP and CS. For the examination of UVER, fibroblasts were irradiated with various UV doses and subsequently infected with either unirradiated or UV-irradiated HSV and scored for plaque formation 2 days later. UVER expression was maximum when the delay between UV-irradiation of the cells and HSV infection was 48 h. (Auth.)

  1. Molecular Characterization of a Novel Family of Trypanosoma cruzi Surface Membrane Proteins (TcSMP) Involved in Mammalian Host Cell Invasion.

    Science.gov (United States)

    Martins, Nadini Oliveira; Souza, Renata Torres de; Cordero, Esteban Mauricio; Maldonado, Danielle Cortez; Cortez, Cristian; Marini, Marjorie Mendes; Ferreira, Eden Ramalho; Bayer-Santos, Ethel; Almeida, Igor Correia de; Yoshida, Nobuko; Silveira, José Franco da

    2015-11-01

    TcSMP may be additive to that triggered by the major surface molecule gp82, further increasing the host cell responses required for infection.

  2. Trans-suppression of host CDH3 and LOXL4 genes during Cryptosporidium parvum infection involves nuclear delivery of parasite Cdg7_FLc_1000 RNA.

    Science.gov (United States)

    Ming, Zhenping; Gong, Ai-Yu; Wang, Yang; Zhang, Xin-Tian; Li, Min; Li, Yao; Pang, Jing; Dong, Stephanie; Strauss-Soukup, Juliane K; Chen, Xian-Ming

    2018-05-01

    Intestinal infection by Cryptosporidium parvum causes significant alterations in the gene expression profile in host epithelial cells. Previous studies demonstrate that a panel of parasite RNA transcripts of low protein-coding potential are delivered into infected host cells and may modulate host gene transcription. Using in vitro models of human intestinal cryptosporidiosis, we report here that trans-suppression of the cadherin 3 (CDH3) and lysyl oxidase like 4 (LOXL4) genes in human intestinal epithelial cells following C. parvum infection involves host delivery of the Cdg7_FLc_1000 RNA, a C. parvum RNA that has been previously demonstrated to be delivered into the nuclei of infected host cells. Downregulation of CDH3 and LOXL4 genes was detected in host epithelial cells following C. parvum infection or in cells expressing the parasite Cdg7_FLc_1000 RNA. Knockdown of Cdg7_FLc_1000 attenuated the trans-suppression of CDH3 and LOXL4 genes in host cells induced by infection. Interestingly, Cdg7_FLc_1000 was detected to be recruited to the promoter regions of both CDH3 and LOXL4 gene loci in host cells following C. parvum infection. Host delivery of Cdg7_FLc_1000 promoted the PH domain zinc finger protein 1 (PRDM1)-mediated H3K9 methylation associated with trans-suppression in the CDH3 gene locus, but not the LOXL4 gene. Therefore, our data suggest that host delivery of Cdg7_FLc_1000 causes CDH3 trans-suppression in human intestinal epithelial cells following C. parvum infection through PRDM1-mediated H3K9 methylation in the CDH3 gene locus, whereas Cdg7_FLc_1000 induces trans-suppression of the host LOXL4 gene through H3K9/H3K27 methylation-independent mechanisms. Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

  3. Opposing regulation of PROX1 by interleukin-3 receptor and NOTCH directs differential host cell fate reprogramming by Kaposi sarcoma herpes virus.

    Directory of Open Access Journals (Sweden)

    Jaehyuk Yoo

    Full Text Available Lymphatic endothelial cells (LECs are differentiated from blood vascular endothelial cells (BECs during embryogenesis and this physiological cell fate specification is controlled by PROX1, the master regulator for lymphatic development. When Kaposi sarcoma herpes virus (KSHV infects host cells, it activates the otherwise silenced embryonic endothelial differentiation program and reprograms their cell fates. Interestingly, previous studies demonstrated that KSHV drives BECs to acquire a partial lymphatic phenotype by upregulating PROX1 (forward reprogramming, but stimulates LECs to regain some BEC-signature genes by downregulating PROX1 (reverse reprogramming. Despite the significance of this KSHV-induced bidirectional cell fate reprogramming in KS pathogenesis, its underlying molecular mechanism remains undefined. Here, we report that IL3 receptor alpha (IL3Rα and NOTCH play integral roles in the host cell type-specific regulation of PROX1 by KSHV. In BECs, KSHV upregulates IL3Rα and phosphorylates STAT5, which binds and activates the PROX1 promoter. In LECs, however, PROX1 was rather downregulated by KSHV-induced NOTCH signal via HEY1, which binds and represses the PROX1 promoter. Moreover, PROX1 was found to be required to maintain HEY1 expression in LECs, establishing a reciprocal regulation between PROX1 and HEY1. Upon co-activation of IL3Rα and NOTCH, PROX1 was upregulated in BECs, but downregulated in LECs. Together, our study provides the molecular mechanism underlying the cell type-specific endothelial fate reprogramming by KSHV.

  4. Sialoglycoconjugates in Trypanosoma cruzi-host cell interaction: possible biological model - a review

    Directory of Open Access Journals (Sweden)

    Alane Beatriz Vermelho

    1994-03-01

    Full Text Available A number of glycoconjugates, including glycolipids and glycoproteins, participate in the process of host-cell invasion by Trypanosoma cruzi and one of the most important carbohydrates involved on this interaction is sialic acid. It is known that parasite trans-sialidase participates with sialic acid in a coordinated fashion in the initial stages of invasion. Given the importance of these sialogycoconjugates, this review sets out various possible biological models for the interaction between the parasite and mammalian cells that possess a sialylated receptor/ligand system.

  5. Evaluation of maternal urinary tract infection as a potential risk factor for neonatal urinary tract infection.

    OpenAIRE

    Nasrin Khalesi; Nastaran Khosravi; Ali Jalali; Leila Amini

    2014-01-01

    Objective To assess the relationship between maternal UTI during pregnancy and neonatal UTI. Materials and methods This cross-sectional study included eighty neonates referred to Ali-e-Asghar Hospital, Tehran, Iran, in 2011. The participants were divided into the study (with diagnosis of UTI; n = 40) and the control (without this type of infection; n = 40) groups. The mothers were asked about the history of UTI during pregnancy, and if the response was positive, the trimester in which UTI had...

  6. Natural Products for the Treatment of Chlamydiaceae Infections

    Directory of Open Access Journals (Sweden)

    Mika A. Brown

    2016-10-01

    Full Text Available Due to the global prevalence of Chlamydiae, exploring studies of diverse antichlamydial compounds is important in the development of effective treatment strategies and global infectious disease management. Chlamydiaceae is the most widely known bacterial family of the Chlamydiae order. Among the species in the family Chlamydiaceae, Chlamydia trachomatis and Chlamydia pneumoniae cause common human diseases, while Chlamydia abortus, Chlamydia psittaci, and Chlamydia suis represent zoonotic threats or are endemic in human food sources. Although chlamydial infections are currently manageable in human populations, chlamydial infections in livestock are endemic and there is significant difficulty achieving effective treatment. To combat the spread of Chlamydiaceae in humans and other hosts, improved methods for treatment and prevention of infection are needed. There exist various studies exploring the potential of natural products for developing new antichlamydial treatment modalities. Polyphenolic compounds can inhibit chlamydial growth by membrane disruption, reestablishment of host cell apoptosis, or improving host immune system detection. Fatty acids, monoglycerides, and lipids can disrupt the cell membranes of infective chlamydial elementary bodies (EBs. Peptides can disrupt the cell membranes of chlamydial EBs, and transferrins can inhibit chlamydial EBs from attachment to and permeation through the membranes of host cells. Cellular metabolites and probiotic bacteria can inhibit chlamydial infection by modulating host immune responses and directly inhibiting chlamydial growth. Finally, early stage clinical trials indicate that polyherbal formulations can be effective in treating chlamydial infections. Herein, we review an important body of literature in the field of antichlamydial research.

  7. Host cell virus entry mediated by Australian bat lyssavirus G envelope glycoprotein occurs through a clathrin-mediated endocytic pathway that requires actin and Rab5.

    Science.gov (United States)

    Weir, Dawn L; Laing, Eric D; Smith, Ina L; Wang, Lin-Fa; Broder, Christopher C

    2014-02-27

    Australian bat lyssavirus (ABLV), a rhabdovirus of the genus Lyssavirus which circulates in both pteropid fruit bats and insectivorous bats in mainland Australia, has caused three fatal human infections, the most recent in February 2013, manifested as acute neurological disease indistinguishable from clinical rabies. Rhabdoviruses infect host cells through receptor-mediated endocytosis and subsequent pH-dependent fusion mediated by their single envelope glycoprotein (G), but the specific host factors and pathways involved in ABLV entry have not been determined. ABLV internalization into HEK293T cells was examined using maxGFP-encoding recombinant vesicular stomatitis viruses (rVSV) that express ABLV G glycoproteins. A combination of chemical and molecular approaches was used to investigate the contribution of different endocytic pathways to ABLV entry. Dominant negative Rab GTPases were used to identify the endosomal compartment utilized by ABLV to gain entry into the host cell cytosol. Here we show that ABLV G-mediated entry into HEK293T cells was significantly inhibited by the dynamin-specific inhibitor dynasore, chlorpromazine, a drug that blocks clathrin-mediated endocytosis, and the actin depolymerizing drug latrunculin B. Over expression of dominant negative mutants of Eps15 and Rab5 also significantly reduced ABLV G-mediated entry into HEK293T cells. Chemical inhibitors of caveolae-dependent endocytosis and macropinocytosis and dominant negative mutants of Rab7 and Rab11 had no effect on ABLV entry. The predominant pathway utilized by ABLV for internalization into HEK293T cells is clathrin-and actin-dependent. The requirement of Rab5 for productive infection indicates that ABLV G-mediated fusion occurs within the early endosome compartment.

  8. Capture of cell culture-derived influenza virus by lectins: strain independent, but host cell dependent.

    Science.gov (United States)

    Opitz, Lars; Zimmermann, Anke; Lehmann, Sylvia; Genzel, Yvonne; Lübben, Holger; Reichl, Udo; Wolff, Michael W

    2008-12-01

    Strategies to control influenza outbreaks are focused mainly on prophylactic vaccination. Human influenza vaccines are trivalent blends of different virus subtypes. Therefore and due to frequent antigenic drifts, strain independent manufacturing processes are required for vaccine production. This study verifies the strain independency of a capture method based on Euonymus europaeus lectin-affinity chromatography (EEL-AC) for downstream processing of influenza viruses under various culture conditions propagated in MDCK cells. A comprehensive lectin binding screening was conducted for two influenza virus types from the season 2007/2008 (A/Wisconsin/67/2005, B/Malaysia/2506/2004) including a comparison of virus-lectin interaction by surface plasmon resonance technology. EEL-AC resulted in a reproducible high product recovery rate and a high degree of contaminant removal in the case of both MDCK cell-derived influenza virus types demonstrating clearly the general applicability of EEL-AC. In addition, host cell dependency of EEL-AC was studied with two industrial relevant cell lines: Vero and MDCK cells. However, the choice of the host cell lines is known to lead to different product glycosylation profiles. Hence, altered lectin specificities have been observed between the two cell lines, requiring process adaptations between different influenza vaccine production systems.

  9. Interaction of KSHV with Host Cell Surface Receptors and Cell Entry

    Directory of Open Access Journals (Sweden)

    Mohanan Valiya Veettil

    2014-10-01

    Full Text Available Virus entry is a complex process characterized by a sequence of events. Since the discovery of KSHV in 1994, tremendous progress has been made in our understanding of KSHV entry into its in vitro target cells. KSHV entry is a complex multistep process involving viral envelope glycoproteins and several cell surface molecules that is utilized by KSHV for its attachment and entry. KSHV has a broad cell tropism and the attachment and receptor engagement on target cells have an important role in determining the cell type-specific mode of entry. KSHV utilizes heparan sulfate, integrins and EphrinA2 molecules as receptors which results in the activation of host cell pre-existing signal pathways that facilitate the subsequent cascade of events resulting in the rapid entry of virus particles, trafficking towards the nucleus followed by viral and host gene expression. KSHV enters human fibroblast cells by dynamin dependant clathrin mediated endocytosis and by dynamin independent macropinocytosis in dermal endothelial cells. Once internalized into endosomes, fusion of the viral envelope with the endosomal membranes in an acidification dependent manner results in the release of capsids which subsequently reaches the nuclear pore vicinity leading to the delivery of viral DNA into the nucleus. In this review, we discuss the principal mechanisms that enable KSHV to interact with the host cell surface receptors as well as the mechanisms that are required to modulate cell signaling machinery for a successful entry.

  10. The Pseudomonas aeruginosa lectin LecA triggers host cell signalling by glycosphingolipid-dependent phosphorylation of the adaptor protein CrkII.

    Science.gov (United States)

    Zheng, Shuangshuang; Eierhoff, Thorsten; Aigal, Sahaja; Brandel, Annette; Thuenauer, Roland; de Bentzmann, Sophie; Imberty, Anne; Römer, Winfried

    2017-07-01

    The human pathogen Pseudomonas aeruginosa induces phosphorylation of the adaptor protein CrkII by activating the non-receptor tyrosine kinase Abl to promote its uptake into host cells. So far, specific factors of P. aeruginosa, which induce Abl/CrkII signalling, are entirely unknown. In this research, we employed human lung epithelial cells H1299, Chinese hamster ovary cells and P. aeruginosa wild type strain PAO1 to study the invasion process of P. aeruginosa into host cells by using microbiological, biochemical and cell biological approaches such as Western Blot, immunofluorescence microscopy and flow cytometry. Here, we demonstrate that the host glycosphingolipid globotriaosylceramide, also termed Gb3, represents a signalling receptor for the P. aeruginosa lectin LecA to induce CrkII phosphorylation at tyrosine 221. Alterations in Gb3 expression and LecA function correlate with CrkII phosphorylation. Interestingly, phosphorylation of CrkII Y221 occurs independently of Abl kinase. We further show that Src family kinases transduce the signal induced by LecA binding to Gb3, leading to Crk Y221 phosphorylation. In summary, we identified LecA as a bacterial factor, which utilizes a so far unrecognized mechanism for phospho-CrkII Y221 induction by binding to the host glycosphingolipid receptor Gb3. The LecA/Gb3 interaction highlights the potential of glycolipids to mediate signalling processes across the plasma membrane and should be further elucidated to gain deeper insights into this non-canonical mechanism of activating host cell processes. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Evaluation of maternal urinary tract infection as a potential risk factor for neonatal urinary tract infection.

    Science.gov (United States)

    Khalesi, Nasrin; Khosravi, Nastaran; Jalali, Ali; Amini, Leila

    2014-06-01

    To assess the relationship between maternal UTI during pregnancy and neonatal UTI. This cross-sectional study included eighty neonates referred to Ali-e-Asghar Hospital, Tehran, Iran, in 2011. The participants were divided into the study (with diagnosis of UTI; n = 40) and the control (without this type of infection; n = 40) groups. The mothers were asked about the history of UTI during pregnancy, and if the response was positive, the trimester in which UTI had occurred. Urinalysis and urine culture were carried out for all neonates. Overall, 14.9% of neonates had mothers with a positive history of UTI during their pregnancy (4.4%, 6.1%, and 4.4% during the 1(st), 2(nd), and 3(rd) trimesters, respectively). A significant relationship was detected between the occurrence of UTI in neonates and maternal UTI, so that the overall prevalence of UTI among neonates of affected mothers was significantly higher than that observed among non-infected mothers (30.0% versus 6.8%; p = 0.001). Maternal UTI resulted in 5.9-fold increased risk of neonatal UTI. In UTI group, the most common bacterial etiologies of UTI were Escherichia coli (65.9%), followed by Klebsiella (14.6%) and Staphylococci (9.8%). Our findings confirmed the association between the history of UTI in mother and occurrence of UTI in neonate, emphasizing to pay more attention for assessing and managing UTI in neonates in order to reduce the related complications.

  12. Lymphocytes and macrophages are infected by Theileria equi, but T cells and B cells are not required to establish infection in vivo.

    Directory of Open Access Journals (Sweden)

    Joshua D Ramsay

    Full Text Available Theileria equi has a biphasic life cycle in horses, with a period of intraleukocyte development followed by patent erythrocytic parasitemia that causes acute and sometimes fatal hemolytic disease. Unlike Theileria spp. that infect cattle (Theileria parva and Theileria annulata, the intraleukocyte stage (schizont of Theileria equi does not cause uncontrolled host cell proliferation or other significant pathology. Nevertheless, schizont-infected leukocytes are of interest because of their potential to alter host cell function and because immune responses directed against this stage could halt infection and prevent disease. Based on cellular morphology, Theileria equi has been reported to infect lymphocytes in vivo and in vitro, but the specific phenotype of schizont-infected cells has yet to be defined. To resolve this knowledge gap in Theileria equi pathogenesis, peripheral blood mononuclear cells were infected in vitro and the phenotype of infected cells determined using flow cytometry and immunofluorescence microscopy. These experiments demonstrated that the host cell range of Theileria equi was broader than initially reported and included B lymphocytes, T lymphocytes and monocyte/macrophages. To determine if B and T lymphocytes were required to establish infection in vivo, horses affected with severe combined immunodeficiency (SCID, which lack functional B and T lymphocytes, were inoculated with Theileria equi sporozoites. SCID horses developed patent erythrocytic parasitemia, indicating that B and T lymphocytes are not necessary to complete the Theileria equi life cycle in vivo. These findings suggest that the factors mediating Theileria equi leukocyte invasion and intracytoplasmic differentiation are common to several leukocyte subsets and are less restricted than for Theileria annulata and Theileria parva. These data will greatly facilitate future investigation into the relationships between Theileria equi leukocyte tropism and pathogenesis

  13. Evaluation of maternal urinary tract infection as a potential risk factor for neonatal urinary tract infection.

    Directory of Open Access Journals (Sweden)

    Nasrin Khalesi

    2014-06-01

    Full Text Available To assess the relationship between maternal UTI during pregnancy and neonatal UTI.This cross-sectional study included eighty neonates referred to Ali-e-Asghar Hospital, Tehran, Iran, in 2011. The participants were divided into the study (with diagnosis of UTI; n = 40 and the control (without this type of infection; n = 40 groups. The mothers were asked about the history of UTI during pregnancy, and if the response was positive, the trimester in which UTI had occurred. Urinalysis and urine culture were carried out for all neonates.Overall, 14.9% of neonates had mothers with a positive history of UTI during their pregnancy (4.4%, 6.1%, and 4.4% during the 1(st, 2(nd, and 3(rd trimesters, respectively. A significant relationship was detected between the occurrence of UTI in neonates and maternal UTI, so that the overall prevalence of UTI among neonates of affected mothers was significantly higher than that observed among non-infected mothers (30.0% versus 6.8%; p = 0.001. Maternal UTI resulted in 5.9-fold increased risk of neonatal UTI. In UTI group, the most common bacterial etiologies of UTI were Escherichia coli (65.9%, followed by Klebsiella (14.6% and Staphylococci (9.8%.Our findings confirmed the association between the history of UTI in mother and occurrence of UTI in neonate, emphasizing to pay more attention for assessing and managing UTI in neonates in order to reduce the related complications.

  14. Potential Role of Carvedilol in the Cardiac Immune Response Induced by Experimental Infection with Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Aline Luciano Horta

    2017-01-01

    Full Text Available Trypanosoma cruzi causes a cardiac infection characterized by an inflammatory imbalance that could become the inciting factor of the illness. To this end, we evaluated the role of carvedilol, a beta-blocker with potential immunomodulatory properties, on the immune response in C57BL/6 mice infected with VL-10 strain of T. cruzi in the acute phase. Animals (n=40 were grouped: (i not infected, (ii infected, (iii infected + carvedilol, and (iv not infected + carvedilol. We analyzed parameters related to parasitemia, plasma levels of TNF, IL-10, and CCL2, and cardiac histopathology after the administration of carvedilol for 30 days. We did not observe differences in the maximum peaks of parasitemia in the day of their detection among the groups. The plasma TNF was elevated at 60 days of infection in mice treated or not with carvedilol. However, we observed a decreased CCL2 level and increased IL-10 levels in those infected animals treated with carvedilol, which impacted the reduction of the inflammatory infiltration in cardiac tissue. For this experimental model, carvedilol therapy was not able to alter the levels of circulating parasites but modulates the pattern of CCL2 and IL-10 mediators when the VL10 strain of T. cruzi was used in C57BL6 mice.

  15. Neisseria meningitidis differentially controls host cell motility through PilC1 and PilC2 components of type IV Pili.

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    Philippe C Morand

    Full Text Available Neisseria meningitidis is a strictly human pathogen that has two facets since asymptomatic carriage can unpredictably turn into fulminant forms of infection. Meningococcal pathogenesis relies on the ability of the bacteria to break host epithelial or endothelial cellular barriers. Highly restrictive, yet poorly understood, mechanisms allow meningococcal adhesion to cells of only human origin. Adhesion of encapsulated and virulent meningococci to human cells relies on the expression of bacterial type four pili (T4P that trigger intense host cell signalling. Among the components of the meningococcal T4P, the concomitantly expressed PilC1 and PilC2 proteins regulate pili exposure at the bacterial surface, and until now, PilC1 was believed to be specifically responsible for T4P-mediated meningococcal adhesion to human cells. Contrary to previous reports, we show that, like PilC1, the meningococcal PilC2 component is capable of mediating adhesion to human ME180 epithelial cells, with cortical plaque formation and F-actin condensation. However, PilC1 and PilC2 promote different effects on infected cells. Cellular tracking analysis revealed that PilC1-expressing meningococci caused a severe reduction in the motility of infected cells, which was not the case when cells were infected with PilC2-expressing strains. The amount of both total and phosphorylated forms of EGFR was dramatically reduced in cells upon PilC1-mediated infection. In contrast, PilC2-mediated infection did not notably affect the EGFR pathway, and these specificities were shared among unrelated meningococcal strains. These results suggest that meningococci have evolved a highly discriminative tool for differential adhesion in specific microenvironments where different cell types are present. Moreover, the fine-tuning of cellular control through the combined action of two concomitantly expressed, but distinctly regulated, T4P-associated variants of the same molecule (i.e. PilC1 and Pil

  16. Immunoregulatory Effects Triggered by Lactic Acid Bacteria Exopolysaccharides: New Insights into Molecular Interactions with Host Cells

    Directory of Open Access Journals (Sweden)

    Jonathan Laiño

    2016-08-01

    Full Text Available Researchers have demonstrated that lactic acid bacteria (LAB with immunomodulatory capabilities (immunobiotics exert their beneficial effects through several molecules, including cell wall, peptidoglycan, and exopolysaccharides (EPS, that are able to interact with specific host cell receptors. EPS from LAB show a wide heterogeneity in its composition, meaning that biological properties depend on the strain and. therefore, only a part of the mechanism of action has been elucidated for these molecules. In this review, we summarize the current knowledge of the health-promoting actions of EPS from LAB with special focus on their immunoregulatory actions. In addition, we describe our studies using porcine intestinal epithelial cells (PIE cells as a model to evaluate the molecular interactions of EPS from two immunobiotic LAB strains and the host cells. Our studies showed that EPS from immunobiotic LAB have anti-inflammatory capacities in PIE cells since they are able to reduce the production of inflammatory cytokines in cells challenged with the Toll-like receptor (TLR-4-agonist lipopolysaccharide. The effects of EPS were dependent on TLR2, TLR4, and negative regulators of TLR signaling. We also reported that the radioprotective 105 (RP105/MD1 complex, a member of the TLR family, is partially involved in the immunoregulatory effects of the EPS from LAB. Our work described, for the first time, that LAB and their EPS reduce inflammation in intestinal epithelial cells in a RP105/MD1-dependent manner. A continuing challenge for the future is to reveal more effector-receptor relationships in immunobiotic-host interactions that contribute to the beneficial effects of these bacteria on mucosal immune homeostasis. A detailed molecular understanding should lead to a more rational use of immunobiotics in general, and their EPS in particular, as efficient prevention and therapies for specific immune-related disorders in humans and animals.

  17. [Redox-potential of blood and consistence of energoproviding defence system in cytomegalovirus infection in pregnancy].

    Science.gov (United States)

    Dzhikiia, I V; Rizhvadze, M A; Dzhangidze, M A

    2006-05-01

    We have studied the relationship between the hypoxic change and mitochondrial redox-potential disturbances in the mechanism of pheto-placental insufficiency in pregnancy with cytomegalovirus infection (CMV), detected by the positive anti-CMV-IgG titer and more then 4-fold increase of low avid anti-CMV-IgG. It was shown, that chronic CMV infection induces production of active forms of oxygen, peroxidation of structures and concurrently damage of mitochondria with essential decrease of ATP level. Results of the study have shown the important diagnostic value of estimation of hypoxic-oxidative damage induced by CMV infection. The results also revealed important relationships between the activity of the CMV infection and intensity of mitochondrial damage. On the basis of our investigations we suggest the additional diagnostic test (the determination of citozol NADH dependent isocitratdehydrogenaze activity) to evaluate the depth of CMV induced metabolic disturbances.

  18. Bcl-2 and Bcl-xL overexpression inhibits cytochrome c release, activation of multiple caspases, and virus release following coxsackievirus B3 infection

    International Nuclear Information System (INIS)

    Carthy, Christopher M.; Yanagawa, Bobby; Luo Honglin; Granville, David J.; Yang, Decheng; Cheung, Paul; Cheung, Caroline; Esfandiarei, Mitra; Rudin, Charles M.; Thompson, Craig B.; Hunt, David W.C.; McManus, Bruce M.

    2003-01-01

    Coxsackievirus B3, a cytopathic virus in the family Picornaviridae, induces degenerative changes in host cell morphology. Here we demonstrate cytochrome c release and caspases-2, -3, -6, -7, -8, and -9 processing. Enforced Bcl-2 and Bcl-xL expression markedly reduced release of cytochrome c, presentation of the mitochondrial epitope 7A6, and depressed caspase activation following infection. In comparison, cell death using TRAIL ligand caused caspase-8 processing prior to cytochrome c release and executioner caspases and cell death was only partially rescued by Bcl-2 and Bcl-xL overexpression. Disruption of the mitochondrial inner membrane potential following CVB3 infection was not inhibited by zVAD.fmk treatment. Bcl-2 or Bcl-xL overexpression or zVAD.fmk treatment delayed the loss of host cell viability and decreased progeny virus release following infection. Our data suggest that mitochondrial release of cytochrome c may be an important early event in caspase activation in CVB3 infection, and, as such, may contribute to the loss of host-cell viability and progeny virus release

  19. Plant RNA binding proteins for control of RNA virus infection

    Directory of Open Access Journals (Sweden)

    Sung Un eHuh

    2013-12-01

    Full Text Available Plant RNA viruses have effective strategies to infect host plants through either direct or indirect interactions with various host proteins, thus suppressing the host immune system. When plant RNA viruses enter host cells exposed RNAs of viruses are recognized by the host immune system through processes such as siRNA-dependent silencing. Interestingly, some host RNA binding proteins have been involved in the inhibition of RNA virus replication, movement, and translation through RNA-specific binding. Host plants intensively use RNA binding proteins for defense against viral infections in nature. In this mini review, we will summarize the function of some host RNA binding proteins which act in a sequence-specific binding manner to the infecting virus RNA. It is important to understand how plants effectively suppresses RNA virus infections via RNA binding proteins, and this defense system can be potentially developed as a synthetic virus defense strategy for use in crop engineering.

  20. A new hypothesis of pathogenesis based on the divorce between mitochondria and their host cells: possible relevance for Alzheimer's disease.

    Science.gov (United States)

    Agnati, L F; Guidolin, D; Baluska, F; Leo, G; Barlow, P W; Carone, C; Genedani, S

    2010-06-01

    On the basis of not only the endosymbiotic theory of eukaryotic cell organization and evolution but also of observations of transcellular communication via Tunneling NanoTubes (TNTs), the hypothesis is put forward that when mitochondria, which were once independently living prokaryote-like organisms, are subjected to detrimental genetic, toxic, or environmental conditions, including age-related endogenous factors, they can regress towards their original independent state. At that point, they can become potentially pathogenic intruders within their eukaryotic host cell. Because of the protoplasmic disequilibrium caused by an altered, or mutated, mitochondral population, certain host cells with a minimal capacity for self-renewal, such as dopaminergic neurons, risk a loss of function and degenerate. It is also proposed that altered mitochondria, as well as their mutated mtDNA, can migrate, via TNTs, into adjacent cells. In this way, neurodegenerative states are propagated between cells (glia and/or neurons) of the Central Nervous System (CNS) and that this leads to conditions such as Alzheimer's and Parkinson's disease. This proposal finds indirect support from observations on rotenone-poisoned glioblastoma cells which have been co-cultured with non-poisoned cells. Immunocytochemical techniques revealed that mitochondria, moving along the TNTs, migrated from the poisoned cells towards the healthy cells. It has also been demonstrated by means of immunocytochemistry that, in glioblastoma cell cultures, Amyloid Precursor Protein (APP) is present in TNTs, hence it may migrate from one cell to neighbouring cells. This datum may be of high relevance for a better understanding of Alzheimer's Disease (AD) since molecular, cellular, and animal model studies have revealed that the formation of amyloid beta (Abeta) and other derivatives of the APP are key pathogenic factors in AD, causing mitochondrial dysfunction, free radical generation, oxidative damage, and inflammation

  1. Mycobacterial secretion systems ESX-1 and ESX-5 play distinct roles in host cell death and inflammasome activation

    KAUST Repository

    Abdallah, Abdallah; Bestebroer, Jovanka; Savage, Nigel D L; De Punder, Karin; Van Zon, Maaike; Wilson, Louis D.; Korbee, Cees J.; Van Der Sar, Astrid M.; Ottenhoff, Tom Hm M; Van Der Wel, Nicole N.; Bitter, Wilbert M.; Peters, Peter J.

    2011-01-01

    for the translocation of Mycobacterium tuberculosis or Mycobacterium marinum to the cytosol of host cells. However, the M. marinum ESX-5 mutant does not induce inflammasome activation and IL-1b activation. The ESX-5 system also induces a caspase-independent cell death

  2. Expression of human CD81 differently affects host cell susceptibility to malaria sporozoites depending on the Plasmodium species.

    NARCIS (Netherlands)

    Silvie, O.; Greco, C.; Franetich, J.F.; Dubart-Kupperschmitt, A.; Hannoun, L.; Gemert, G.J.A. van; Sauerwein, R.W.; Levy, S.; Boucheix, C.; Rubinstein, E.; Mazier, D.

    2006-01-01

    Plasmodium sporozoites can enter host cells by two distinct pathways, either through disruption of the plasma membrane followed by parasite transmigration through cells, or by formation of a parasitophorous vacuole (PV) where the parasite further differentiates into a replicative exo-erythrocytic

  3. Yersinia pseudotuberculosis Spatially Controls Activation and Misregulation of Host Cell Rac1.

    Directory of Open Access Journals (Sweden)

    2005-10-01

    Full Text Available Yersinia pseudotuberculosis binds host cells and modulates the mammalian Rac1 guanosine triphosphatase (GTPase at two levels. Activation of Rac1 results from integrin receptor engagement, while misregulation is promoted by translocation of YopE and YopT proteins into target cells. Little is known regarding how these various factors interplay to control Rac1 dynamics. To investigate these competing processes, the localization of Rac1 activation was imaged microscopically using fluorescence resonance energy transfer. In the absence of translocated effectors, bacteria induced activation of the GTPase at the site of bacterial binding. In contrast, the entire cellular pool of Rac1 was inactivated shortly after translocation of YopE RhoGAP. Inactivation required membrane localization of Rac1. The translocated protease YopT had very different effects on Rac1. This protein, which removes the membrane localization site of Rac1, did not inactivate Rac1, but promoted entry of cleaved activated Rac1 molecules into the host cell nucleus, allowing Rac1 to localize with nuclear guanosine nucleotide exchange factors. As was true for YopE, membrane-associated Rac1 was the target for YopT, indicating that the two translocated effectors may compete for the same pool of target protein. Consistent with the observation that YopE inactivation requires membrane localization of Rac1, the presence of YopT in the cell interfered with the action of the YopE RhoGAP. As a result, interaction of target cells with a strain that produces both YopT and YopE resulted in two spatially distinct pools of Rac1: an inactive cytoplasmic pool and an activated nuclear pool. These studies demonstrate that competition between bacterial virulence factors for access to host substrates is controlled by the spatial arrangement of a target protein. In turn, the combined effects of translocated bacterial proteins are to generate pools of a single signaling molecule with distinct localization and

  4. Potential management of resistant microbial infections with a novel non-antibiotic

    DEFF Research Database (Denmark)

    Dutta, Noton Kumar; Annadurai, Subramanian; Mazumdar, Kaushiki

    2007-01-01

    Diclofenac sodium (Dc), an anti-inflammatory agent, has remarkable inhibitory action both against drug-sensitive and drug-resistant clinical isolates of various Gram-positive and Gram-negative bacteria. The aim of this study was to determine the ability of Dc to protect mice from a virulent...... Salmonella infection. Dc injected at 1.5 microg/g and 3.0 microg/g mouse body weight significantly protected animals from the lethality of Salmonella infection. As was the case for the in vitro interaction, Dc in combination with streptomycin was even more effective. The non-antibiotic drug Dc has potential...... for the management of problematic antibiotic-resistant bacterial infections....

  5. Bay laurel (Laurus nobilis) as potential antiviral treatment in naturally BQCV infected honeybees.

    Science.gov (United States)

    Aurori, Adriana C; Bobiş, Otilia; Dezmirean, Daniel S; Mărghitaş, Liviu A; Erler, Silvio

    2016-08-15

    Viral diseases are one of the multiple factors associated with honeybee colony losses. Apart from their innate immune system, including the RNAi machinery, honeybees can use secondary plant metabolites to reduce or fully cure pathogen infections. Here, we tested the antiviral potential of Laurus nobilis leaf ethanolic extracts on forager honeybees naturally infected with BQCV (Black queen cell virus). Total viral loads were reduced even at the lowest concentration tested (1mg/ml). Higher extract concentrations (≥5mg/ml) significantly reduced virus replication. Measuring vitellogenin gene expression as an indicator for transcript homeostasis revealed constant RNA levels before and after treatment, suggesting that its expression was not impacted by the L. nobilis treatment. In conclusion, plant secondary metabolites can reduce virus loads and virus replication in naturally infected honeybees. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Potential strategies for the eradication of multidrug-resistant Gram-negative bacterial infections.

    Science.gov (United States)

    Huwaitat, Rawan; McCloskey, Alice P; Gilmore, Brendan F; Laverty, Garry

    2016-07-01

    Antimicrobial resistance is one of the leading threats to society. The increasing burden of multidrug-resistant Gram-negative infection is particularly concerning as such bacteria are demonstrating resistance to nearly all currently licensed therapies. Various strategies have been hypothesized to treat multidrug-resistant Gram-negative infections including: targeting the Gram-negative outer membrane; neutralization of lipopolysaccharide; inhibition of bacterial efflux pumps and prevention of protein folding. Silver and silver nanoparticles, fusogenic liposomes and nanotubes are potential strategies for extending the activity of licensed, Gram-positive selective, antibiotics to Gram-negatives. This may serve as a strategy to fill the current void in pharmaceutical development in the short term. This review outlines the most promising strategies that could be implemented to solve the threat of multidrug-resistant Gram-negative infections.

  7. Toroidal surface complexes of bacteriophage φ12 are responsible for host-cell attachment

    International Nuclear Information System (INIS)

    Leo-Macias, Alejandra; Katz, Garrett; Wei Hui; Alimova, Alexandra; Katz, A.; Rice, William J.; Diaz-Avalos, Ruben; Hu Guobin; Stokes, David L.; Gottlieb, Paul

    2011-01-01

    Cryo-electron tomography and subtomogram averaging are utilized to determine that the bacteriophage φ12, a member of the Cystoviridae family, contains surface complexes that are toroidal in shape, are composed of six globular domains with six-fold symmetry, and have a discrete density connecting them to the virus membrane-envelope surface. The lack of this kind of spike in a reassortant of φ12 demonstrates that the gene for the hexameric spike is located in φ12's medium length genome segment, likely to the P3 open reading frames which are the proteins involved in viral-host cell attachment. Based on this and on protein mass estimates derived from the obtained averaged structure, it is suggested that each of the globular domains is most likely composed of a total of four copies of P3a and/or P3c proteins. Our findings may have implications in the study of the evolution of the cystovirus species in regard to their host specificity. - Research Highlights: → Subtomogram averaging reveals enhanced detail of a φ12 cystovirus surface protein complex. → The surface protein complex has a toroidal shape and six-fold symmetry. → It is encoded by the medium-size genome segment. → The proteins of the surface complex most likely are one copy of P3a and three copies of P3c.

  8. Impact of host cell variation on the neutralization of HIV-1 in vitro.

    Science.gov (United States)

    Polonis, Victoria R; Schuitemaker, Hanneke; Bunnik, Evelien M; Brown, Bruce K; Scarlatti, Gabriella

    2009-09-01

    In this review we present current advances in our understanding of HIV-1 neutralization assays that employ primary cell types, as compared with those that utilize cell lines and the newer, more standardized pseudovirus assays. A commentary on the challenges of standardizing in-vitro neutralization assays using primary cells is included. The data from reporter cell line neutralization assays may agree with results observed in primary cells; however, exceptions have recently been reported. Multiple variables exist in primary cell assays using peripheral blood mononuclear cells from HIV-seronegative donors; in-vitro neutralization titers can vary significantly based on the donor cells used for assay targets and for virus propagation. Thus, more research is required to achieve validated primary cell neutralization assays. HIV-vaccine-induced antibody performance in the current neutralization assays may function as a 'gatekeeper' for HIV-1 subunit vaccine advancement. Development of standardized platforms for reproducible measurement of in-vitro neutralization is therefore a high priority. Given the considerable variation in results obtained from some widely applied HIV neutralization platforms, parallel evaluation of new antibodies using different host cells for assay targets, as well as virus propagation, is recommended until immune correlates of protection are identified.

  9. Host cell proteins in biologics development: Identification, quantitation and risk assessment.

    Science.gov (United States)

    Wang, Xing; Hunter, Alan K; Mozier, Ned M

    2009-06-15

    Host cell proteins (HCPs) are those produced or encoded by the organisms and unrelated to the intended recombinant product. Some are necessary for growth, survival, and normal cellular processing whereas others may be non-essential, simply carried along as baggage. Like the recombinant product, HCPs may also be modified by the host with a number of post-translational modifications. Regardless of the utility, or lack thereof, HCPs are undesirable in the final drug substance. Though commonly present in small quantities (parts per million expressed as nanograms per milligrams of the intended recombinant protein) much effort and cost is expended by industry to remove them. The purpose of this review is to summarize what is of relevance in regards to the biology, the impact of genomics and proteomics on HCP evaluation, the regulatory expectations, analytical approaches, and various methodologies to remove HCPs with bioprocessing. Historical data, bioinformatics approaches and industrial case study examples are provided. Finally, a proposal for a risk assessment tool is provided which brings these facets together and proposes a means for manufacturers to classify and organize a control strategy leading to meaningful product specifications. 2009 Wiley Periodicals, Inc.

  10. Exploitation of the host cell ubiquitin machinery by microbial effector proteins.

    Science.gov (United States)

    Lin, Yi-Han; Machner, Matthias P

    2017-06-15

    Pathogenic bacteria are in a constant battle for survival with their host. In order to gain a competitive edge, they employ a variety of sophisticated strategies that allow them to modify conserved host cell processes in ways that favor bacterial survival and growth. Ubiquitylation, the covalent attachment of the small modifier ubiquitin to target proteins, is such a pathway. Ubiquitylation profoundly alters the fate of a myriad of cellular proteins by inducing changes in their stability or function, subcellular localization or interaction with other proteins. Given the importance of ubiquitylation in cell development, protein homeostasis and innate immunity, it is not surprising that this post-translational modification is exploited by a variety of effector proteins from microbial pathogens. Here, we highlight recent advances in our understanding of the many ways microbes take advantage of host ubiquitylation, along with some surprising deviations from the canonical theme. The lessons learned from the in-depth analyses of these host-pathogen interactions provide a fresh perspective on an ancient post-translational modification that we thought was well understood.This article is part of a Minifocus on Ubiquitin Regulation and Function. For further reading, please see related articles: 'Mechanisms of regulation and diversification of deubiquitylating enzyme function' by Pawel Leznicki and Yogesh Kulathu ( J. Cell Sci. 130 , 1997-2006). 'Cell scientist to watch - Mads Gyrd-Hansen' ( J. Cell Sci. 130 , 1981-1983). © 2017. Published by The Company of Biologists Ltd.

  11. Apa is a trimeric autotransporter adhesin of Actinobacillus pleuropneumoniae responsible for autoagglutination and host cell adherence.

    Science.gov (United States)

    Xiao, Longwen; Zhou, Liang; Sun, Changjiang; Feng, Xin; Du, ChongTao; Gao, Yu; Ji, Qun; Yang, Shuxin; Wang, Yu; Han, Wenyu; Langford, P R; Lei, Liancheng

    2012-10-01

    Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia, and adherence to host cells is a key step in the pathogenic process. Although trimeric autotransporter adhesins (TAAs) were identified in many pathogenic bacteria in recent years, none in A. pleuropneumoniae have been characterized. In this study, we identified a TAA from A. pleuropneumoniae, Apa, and characterized the contribution of its amino acid residues to the adhesion process. Sequence analysis of the C-terminal amino acid residues of Apa revealed the presence of a putative translocator domain and six conserved HsfBD1-like or HsfBD2-like binding domains. Western blot analysis revealed that the 126 C-terminal amino acids of Apa could form trimeric molecules. By confocal laser scanning microscopy, one of these six domains (ApaBD3) was determined to mediate adherence to epithelial cells. Adherence assays and adherence inhibition assays using a recombinant E. coli- ApaBD3 strain which expressed ApaBD3 on the surface of E. coli confirmed that this domain was responsible for the adhesion activity. Moreover, cellular enzyme-linked immunosorbent assays demonstrated that ApaBD3 mediated high-level adherence to epithelial cell lines. Intriguingly, autoagglutination was observed with the E. coli- ApaBD3 strain, and this phenomenon was dependent upon the association of the expressed ApaBD3 with the C-terminal translocator domain. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Capillary zone electrophoresis-tandem mass spectrometry detects low concentration host cell impurities in monoclonal antibodies

    Science.gov (United States)

    Zhu, Guijie; Sun, Liangliang; Heidbrink-Thompson, Jennifer; Kuntumalla, Srilatha; Lin, Hung-yu; Larkin, Christopher J.; McGivney, James B.; Dovichi, Norman J.

    2016-01-01

    We have evaluated capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) for detection of trace amounts of host cell protein impurities in recombinant therapeutics. Compared to previously published procedures, we have optimized the buffer pH used in the formation of a pH junction to increase injection volume. We also prepared a five-point calibration curve by spiking twelve standard proteins into a solution of a human monoclonal antibody. A custom CZE-MS/MS system was used to analyze the tryptic digest of this mixture without depletion of the antibody. CZE generated a ~70 min separation window (~90 min total analysis duration) and ~300 peak capacity. We also analyzed the sample using ultra-performance liquid chromatography (UPLC)-MS/MS. CZE-MS/MS generated ~five times higher base peak intensity and more peptide identifications for low-level spiked proteins. Both methods detected all proteins spiked at the ~100 ppm level with respect to the antibody. PMID:26530276

  13. Host cell reactivation by fibroblasts from patients with pigmentary degeneration of the retina

    International Nuclear Information System (INIS)

    Lytle, C.D.; Tarone, R.E.; Barrett, S.F.; Robbins, J.H.; Wirtschafter, J.D.; Dupuy, J.-M.

    1983-01-01

    Cockayne syndrome (CS) is an autosomal recessive disease characterized by numerous clinical abnormalities including acute sun sensitivity and primary pigmentary degeneration of the retina. Cultured fibroblasts from CS patients are hypersensitive to ultraviolet radiation. Host cell reactivation of irradiated virus was studied in CS and in other diseases with retinal degeneration to evaluate repair. The survival of UV-irradiated Herpes simplex virus type 1 was determined in fibroblast lines from four normal donors, two patients with CS, one with both xeroderma pigmentosum (XP) and CS, and from several other patients with (Usher syndrome, olivopontocerebellar atrophy, retinitis pigmentosa) and without (XP, ataxia telangiectasia) primary pigmentary degeneration of the retina. The viral survival curves in all cell lines showed two components: a very sensitive initial component followed by an exponential, less sensitive component. The exponential component had greater sensitivity than normal in the case of the CS patients, the patient with both XP and CS, and the XP patient. It was proposed that patients with CS have defective repair of DNA which may be the cause of their retinal degeneration. (author)

  14. Host cell reactivation by fibroblasts from patients with pigmentary degeneration of the retina

    Energy Technology Data Exchange (ETDEWEB)

    Lytle, C.D. (Food and Drug Administration, Rockville, MD (USA)); Tarone, R.E.; Barrett, S.F.; Robbins, J.H. (National Cancer Inst., Bethesda, MD (USA)); Wirtschafter, J.D. (Minnesota Univ., Minneapolis (USA). Hospitals); Dupuy, J.M. (Quebec Univ., Laval-des-Rapides (Canada). Inst. Armand-Frappier)

    1983-05-01

    Cockayne syndrome (CS) is an autosomal recessive disease characterized by numerous clinical abnormalities including acute sun sensitivity and primary pigmentary degeneration of the retina. Cultured fibroblasts from CS patients are hypersensitive to ultraviolet radiation. Host cell reactivation of irradiated virus was studied in CS and in other diseases with retinal degeneration to evaluate repair. The survival of UV-irradiated Herpes simplex virus type 1 was determined in fibroblast lines from four normal donors, two patients with CS, one with both xeroderma pigmentosum (XP) and CS, and from several other patients with (Usher syndrome, olivopontocerebellar atrophy, retinitis pigmentosa) and without (XP, ataxia telangiectasia) primary pigmentary degeneration of the retina. The viral survival curves in all cell lines showed two components: a very sensitive initial component followed by an exponential, less sensitive component. The exponential component had greater sensitivity than normal in the case of the CS patients, the patient with both XP and CS, and the XP patient. It was proposed that patients with CS have defective repair of DNA which may be the cause of their retinal degeneration.

  15. Identification of Host Cell Factors Associated with Astrovirus Replication in Caco-2 Cells.

    Science.gov (United States)

    Murillo, Andrea; Vera-Estrella, Rosario; Barkla, Bronwyn J; Méndez, Ernesto; Arias, Carlos F

    2015-10-01

    Astroviruses are small, nonenveloped viruses with a single-stranded positive-sense RNA genome causing acute gastroenteritis in children and immunocompromised patients. Since positive-sense RNA viruses have frequently been found to replicate in association with membranous structures, in this work we characterized the replication of the human astrovirus serotype 8 strain Yuc8 in Caco-2 cells, using density gradient centrifugation and free-flow zonal electrophoresis (FFZE) to fractionate cellular membranes. Structural and nonstructural viral proteins, positive- and negative-sense viral RNA, and infectious virus particles were found to be associated with a distinct population of membranes separated by FFZE. The cellular proteins associated with this membrane population in infected and mock-infected cells were identified by tandem mass spectrometry. The results indicated that membranes derived from multiple cell organelles were present in the population. Gene ontology and protein-protein interaction network analysis showed that groups of proteins with roles in fatty acid synthesis and ATP biosynthesis were highly enriched in the fractions of this population in infected cells. Based on this information, we investigated by RNA interference the role that some of the identified proteins might have in the replication cycle of the virus. Silencing of the expression of genes involved in cholesterol (DHCR7, CYP51A1) and fatty acid (FASN) synthesis, phosphatidylinositol (PI4KIIIβ) and inositol phosphate (ITPR3) metabolism, and RNA helicase activity (DDX23) significantly decreased the amounts of Yuc8 genomic and antigenomic RNA, synthesis of the structural protein VP90, and virus yield. These results strongly suggest that astrovirus RNA replication and particle assembly take place in association with modified membranes potentially derived from multiple cell organelles. Astroviruses are common etiological agents of acute gastroenteritis in children and immunocompromised patients

  16. Identification of Host Cell Factors Associated with Astrovirus Replication in Caco-2 Cells

    Science.gov (United States)

    Murillo, Andrea; Vera-Estrella, Rosario; Barkla, Bronwyn J.; Méndez, Ernesto

    2015-01-01

    ABSTRACT Astroviruses are small, nonenveloped viruses with a single-stranded positive-sense RNA genome causing acute gastroenteritis in children and immunocompromised patients. Since positive-sense RNA viruses have frequently been found to replicate in association with membranous structures, in this work we characterized the replication of the human astrovirus serotype 8 strain Yuc8 in Caco-2 cells, using density gradient centrifugation and free-flow zonal electrophoresis (FFZE) to fractionate cellular membranes. Structural and nonstructural viral proteins, positive- and negative-sense viral RNA, and infectious virus particles were found to be associated with a distinct population of membranes separated by FFZE. The cellular proteins associated with this membrane population in infected and mock-infected cells were identified by tandem mass spectrometry. The results indicated that membranes derived from multiple cell organelles were present in the population. Gene ontology and protein-protein interaction network analysis showed that groups of proteins with roles in fatty acid synthesis and ATP biosynthesis were highly enriched in the fractions of this population in infected cells. Based on this information, we investigated by RNA interference the role that some of the identified proteins might have in the replication cycle of the virus. Silencing of the expression of genes involved in cholesterol (DHCR7, CYP51A1) and fatty acid (FASN) synthesis, phosphatidylinositol (PI4KIIIβ) and inositol phosphate (ITPR3) metabolism, and RNA helicase activity (DDX23) significantly decreased the amounts of Yuc8 genomic and antigenomic RNA, synthesis of the structural protein VP90, and virus yield. These results strongly suggest that astrovirus RNA replication and particle assembly take place in association with modified membranes potentially derived from multiple cell organelles. IMPORTANCE Astroviruses are common etiological agents of acute gastroenteritis in children and

  17. Characterization of in vitro phenotypes of Burkholderia pseudomallei and Burkholderia mallei strains potentially associated with persistent infection in mice.

    Science.gov (United States)

    Bernhards, R C; Cote, C K; Amemiya, K; Waag, D M; Klimko, C P; Worsham, P L; Welkos, S L

    2017-03-01

    Burkholderia pseudomallei (Bp) and Burkholderia mallei (Bm), the agents of melioidosis and glanders, respectively, are Tier 1 biothreats. They infect humans and animals, causing disease ranging from acute and fatal to protracted and chronic. Chronic infections are especially challenging to treat, and the identification of in vitro phenotypic markers which signal progression from acute to persistent infection would be extremely valuable. First, a phenotyping strategy was developed employing colony morphotyping, chemical sensitivity testing, macrophage infection, and lipopolysaccharide fingerprint analyses to distinguish Burkholderia strains. Then mouse spleen isolates collected 3-180 days after infection were characterized phenotypically. Isolates from long-term infections often exhibited increased colony morphology differences and altered patterns of antimicrobial sensitivity and macrophage infection. Some of the Bp and Bm persistent infection isolates clearly displayed enhanced virulence in mice. Future studies will evaluate the potential role and significance of these phenotypic markers in signaling the establishment of a chronic infection.

  18. Animal herpesviruses and their zoonotic potential for cross-species infection

    Directory of Open Access Journals (Sweden)

    Grzegorz Woźniakowski

    2015-05-01

    Full Text Available Herpesviruses of humans and animals cause severe diseases that influence not only the health and epidemiological status but are also economically important in the context of food production. The members of Herpesviridae are host specific agents that also share many properties that potentially make them capable of crossing the species barriers. The objective of presented review paper was to summarize the relationship between herpesviruses of animals and humans and their zoonotic potential. In humans, the most epidemiologically important herpesviruses are represented by Human herepesvirus-1 and Human herpesvirus-2, which are commonly known as herpes simplex virus type 1 and 2, varicella-zooster virus (VZV, Epstein-Barr virus (EBV, Kaposi’s Sarcoma-associated herpesvirus (KSHV, cytomegalovirus (CMV, as well as Human herpesviruses: HHV-6A, HHV-6B, and HHV-7. However, in terms of the potential to cross the species barrier, there are a few herpesviruses, including B virus disease (CeHV-1, Marek’s disease virus (MDV, Equid herpesvirus-1 (EHV-1 or pseudorabies virus (PRV, which are potentially able to infect different hosts. To summarize, in advantageous conditions the host specific herpesviruses may pose a threat for public health but also may exert a negative impact on the economical aspects of animal production. The most probable of these are zoonotic infections caused by B virus disease; however, close contact between infected animal hosts and humans may lead to transmission and replication of other Herpesviridae members.

  19. The potential of photo-deposited silver coatings on Foley catheters to prevent urinary tract infections

    International Nuclear Information System (INIS)

    Cooper, Ian Richard; Pollini, Mauro; Paladini, Federica

    2016-01-01

    Catheter-associated urinary tract infection (CAUTI) represents one of the most common causes of morbidity and mortality. The resistance demonstrated by many microorganisms to conventional antibiotic therapies and the increasing health-care costs have recently encouraged the definition of alternative preventive strategies, which can have a positive effect in the management of infections. Antimicrobial urinary catheters have been developed through the photo-chemical deposition of silver coatings on the external and luminal surfaces. The substrates are exposed to ultraviolet radiation after impregnation into a silver-based solution, thus inducing the in situ synthesis of silver particles. The effect of the surface treatment on the material was investigated through scanning electron microscopy (SEM) and silver ion release measurements. The ability of microorganisms commonly associated with urinary tract infections was investigated in terms of bacterial viability, proliferation and biofilm development, using Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis as target organisms. The silver coatings demonstrated good distribution of silver particles to the substrate, and proved an effective antibacterial capability in simulated biological conditions. The low values of silver ion release demonstrated the optimum adhesion of the coating. The results indicated a good potential of silver-based antimicrobial materials for prevention of catheter-associated urinary tract infection. - Highlights: • Silver nanocoatings were deposited on urinary catheters. • Both luminal and outer surface were successfully treated. • The treated devices demonstrated were effective against different microorganisms. • The antibacterial potential of the devices was assessed.

  20. The potential of photo-deposited silver coatings on Foley catheters to prevent urinary tract infections

    Energy Technology Data Exchange (ETDEWEB)

    Cooper, Ian Richard [School of Pharmacy & Biomolecular Sciences, University of Brighton, Lewes Road, Brighton BN2 4GJ (United Kingdom); Pollini, Mauro, E-mail: mauro.pollini@unisalento.it [Department of Engineering for Innovation, University of Salento, Via Monteroni, 73100 Lecce (Italy); Silvertech Ltd, Via Monteroni, 73100 Lecce (Italy); Paladini, Federica [Department of Engineering for Innovation, University of Salento, Via Monteroni, 73100 Lecce (Italy)

    2016-12-01

    Catheter-associated urinary tract infection (CAUTI) represents one of the most common causes of morbidity and mortality. The resistance demonstrated by many microorganisms to conventional antibiotic therapies and the increasing health-care costs have recently encouraged the definition of alternative preventive strategies, which can have a positive effect in the management of infections. Antimicrobial urinary catheters have been developed through the photo-chemical deposition of silver coatings on the external and luminal surfaces. The substrates are exposed to ultraviolet radiation after impregnation into a silver-based solution, thus inducing the in situ synthesis of silver particles. The effect of the surface treatment on the material was investigated through scanning electron microscopy (SEM) and silver ion release measurements. The ability of microorganisms commonly associated with urinary tract infections was investigated in terms of bacterial viability, proliferation and biofilm development, using Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis as target organisms. The silver coatings demonstrated good distribution of silver particles to the substrate, and proved an effective antibacterial capability in simulated biological conditions. The low values of silver ion release demonstrated the optimum adhesion of the coating. The results indicated a good potential of silver-based antimicrobial materials for prevention of catheter-associated urinary tract infection. - Highlights: • Silver nanocoatings were deposited on urinary catheters. • Both luminal and outer surface were successfully treated. • The treated devices demonstrated were effective against different microorganisms. • The antibacterial potential of the devices was assessed.

  1. Potential Cost-effectiveness of Early Identification of Hospital-acquired Infection in Critically Ill Patients.

    Science.gov (United States)

    Tsalik, Ephraim L; Li, Yanhong; Hudson, Lori L; Chu, Vivian H; Himmel, Tiffany; Limkakeng, Alex T; Katz, Jason N; Glickman, Seth W; McClain, Micah T; Welty-Wolf, Karen E; Fowler, Vance G; Ginsburg, Geoffrey S; Woods, Christopher W; Reed, Shelby D

    2016-03-01

    Limitations in methods for the rapid diagnosis of hospital-acquired infections often delay initiation of effective antimicrobial therapy. New diagnostic approaches offer potential clinical and cost-related improvements in the management of these infections. We developed a decision modeling framework to assess the potential cost-effectiveness of a rapid biomarker assay to identify hospital-acquired infection in high-risk patients earlier than standard diagnostic testing. The framework includes parameters representing rates of infection, rates of delayed appropriate therapy, and impact of delayed therapy on mortality, along with assumptions about diagnostic test characteristics and their impact on delayed therapy and length of stay. Parameter estimates were based on contemporary, published studies and supplemented with data from a four-site, observational, clinical study. Extensive sensitivity analyses were performed. The base-case analysis assumed 17.6% of ventilated patients and 11.2% of nonventilated patients develop hospital-acquired infection and that 28.7% of patients with hospital-acquired infection experience delays in appropriate antibiotic therapy with standard care. We assumed this percentage decreased by 50% (to 14.4%) among patients with true-positive results and increased by 50% (to 43.1%) among patients with false-negative results using a hypothetical biomarker assay. Cost of testing was set at $110/d. In the base-case analysis, among ventilated patients, daily diagnostic testing starting on admission reduced inpatient mortality from 12.3 to 11.9% and increased mean costs by $1,640 per patient, resulting in an incremental cost-effectiveness ratio of $21,389 per life-year saved. Among nonventilated patients, inpatient mortality decreased from 7.3 to 7.1% and costs increased by $1,381 with diagnostic testing. The resulting incremental cost-effectiveness ratio was $42,325 per life-year saved. Threshold analyses revealed the probabilities of developing

  2. Prevalence and Potential Risk Factors for Bartonella Infection in Tunisian Stray Dogs.

    Science.gov (United States)

    Belkhiria, Jaber; Chomel, Bruno B; Ben Hamida, Taoufik; Kasten, Rickie W; Stuckey, Matthew J; Fleischman, Drew A; Christopher, Mary M; Boulouis, Henri-Jean; Farver, Thomas B

    2017-06-01

    Bartonellae are blood-borne and vector-transmitted pathogens, some are zoonotic, which have been reported in several Mediterranean countries. Transmission from dogs to humans is suspected, but has not been clearly demonstrated. Our objectives were to determine the seroprevalence of Bartonella henselae, Bartonella vinsonii subsp. berkhoffii, Bartonella clarridgeiae, and Bartonella bovis (as a proxy for Candidatus Bartonella merieuxii) in stray dogs from Tunisia, identify the Bartonella species infecting the dogs and evaluate potential risk factors for canine infection. Blood samples were collected between January and November 2013 from 149 dogs in 10 Tunisian governorates covering several climatic zones. Dog-specific and geographic variables were analyzed as potential risk factors for Bartonella spp. seropositivity and PCR-positivity. DNA was extracted from the blood of all dogs and tested by PCR for Bartonella, targeting the ftsZ and rpoB genes. Partial sequencing was performed on PCR-positive dogs. Twenty-nine dogs (19.5%, 95% confidence interval: 14-27.4) were seropositive for one or more Bartonella species, including 17 (11.4%) for B. vinsonii subsp. berkhoffii, 14 (9.4%) for B. henselae, 13 (8.4%) for B. clarridgeiae, and 7 (4.7%) for B. bovis. Statistical analysis revealed a few potential risk factors, mainly dog's age and breed, latitude and average winter temperature. Twenty-two (14.8%) dogs, including 8 of the 29 seropositive dogs, were PCR-positive for Bartonella based on the ftsZ gene, with 18 (81.8%) of these 22 dogs also positive for the rpoB gene. Partial sequencing showed that all PCR-positive dogs were infected with Candidatus B. merieuxii. Dogs from arid regions and regions with cold average winter temperatures were less likely to be PCR-positive than dogs from other climatic zones. The widespread presence of Bartonella spp. infection in Tunisian dogs suggests a role for stray dogs as potential reservoirs of Bartonella species in Tunisia.

  3. The Potential Role of Direct and Indirect Contacts on Infection Spread in Dairy Farm Networks.

    Directory of Open Access Journals (Sweden)

    Gianluigi Rossi

    2017-01-01

    Full Text Available Animals' exchanges are considered the most effective route of between-farm infectious disease transmission. However, despite being often overlooked, the infection spread due to contaminated equipment, vehicles, or personnel proved to be important for several livestock epidemics. This study investigated the role of indirect contacts in a potential infection spread in the dairy farm network of the Province of Parma (Northern Italy. We built between-farm contact networks using data on cattle exchange (direct contacts, and on-farm visits by veterinarians (indirect contacts. We compared the features of the contact structures by using measures on static and temporal networks. We assessed the disease spreading potential of the direct and indirect network structures in the farm system by using data on the infection state of farms by paratuberculosis. Direct and indirect networks showed non-trivial differences with respect to connectivity, contact distribution, and super-spreaders identification. Furthermore, our analyses on paratuberculosis data suggested that the contributions of direct and indirect contacts on diseases spread are apparent at different spatial scales. Our results highlighted the potential role of indirect contacts in between-farm disease spread and underlined the need for a deeper understanding of these contacts to develop better strategies for prevention of livestock epidemics.

  4. A virulence-associated filamentous bacteriophage of Neisseria meningitidis increases host-cell colonisation.

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    Emmanuelle Bille

    2017-07-01

    Full Text Available Neisseria meningitidis is a commensal of human nasopharynx. In some circumstances, this bacteria can invade the bloodstream and, after crossing the blood brain barrier, the meninges. A filamentous phage, designated MDAΦ for Meningococcal Disease Associated, has been associated with invasive disease. In this work we show that the prophage is not associated with a higher virulence during the bloodstream phase of the disease. However, looking at the interaction of N. meningitidis with epithelial cells, a step essential for colonization of the nasopharynx, we demonstrate that the presence of the prophage, via the production of viruses, increases colonization of encapsulated meningococci onto monolayers of epithelial cells. The analysis of the biomass covering the epithelial cells revealed that meningococci are bound to the apical surface of host cells by few layers of heavily piliated bacteria, whereas, in the upper layers, bacteria are non-piliated but surrounded by phage particles which (i form bundles of filaments, and/or (ii are in some places associated with bacteria. The latter are likely to correspond to growing bacteriophages during their extrusion through the outer membrane. These data suggest that, as the biomass increases, the loss of piliation in the upper layers of the biomass does not allow type IV pilus bacterial aggregation, but is compensated by a large production of phage particles that promote bacterial aggregation via the formation of bundles of phage filaments linked to the bacterial cell walls. We propose that MDAΦ by increasing bacterial colonization in the mucosa at the site-of-entry, increase the occurrence of diseases.

  5. Exploring the hidden potential of fosfomycin for the fight against severe Gram-negative infections

    Directory of Open Access Journals (Sweden)

    P V Saiprasad

    2016-01-01

    Full Text Available Gram-negative resistance is a serious global crisis putting the world on the cusp of 'pre-antibiotic era'. This serious crisis has been catalysed by the rapid increase in carbapenem-resistant Enterobacteriaceae (CRE. Spurge in colistin usage to combat CRE infections leads to the reports of (colistin and carbapenem resistant enterobacteriaceae CCRE (resistance to colistin in isolates of CRE infections further jeopardising our last defence. The antibacterial apocalypse imposed by global resistance crisis requires urgent alternative therapeutic options. Interest in the use of fosfomycin renewed recently for serious systemic infections caused by multidrug-resistant Enterobacteriaceae. This review aimed at analysing the recent evidence on intravenous fosfomycin to explore its hidden potential, especially when fosfomycin disodium is going to be available in India. Although a number of promising evidence are coming up for fosfomycin, there are still areas where more work is required to establish intravenous fosfomycin as the last resort antibacterial for severe Gram-negative infections.

  6. Recurrent paediatric pinworm infection of the vagina as a potential reservoir for Enterobius vermicularis.

    Science.gov (United States)

    Kashyap, B; Samantray, J C; Kumar, S; Jhamb, R; Singh, A K; Kaur, I R

    2014-09-01

    Enterobius vermicularis infection remains one of the most common parasitic infections, particularly prevalent in children. Enterobiasis, although not usually dangerous, may cause significant morbidity. Elimination of the parasite from a family or an institution often poses problems, either because of an incomplete cure or re-infection. While there have been limited reports of ectopic enterobiasis throughout the world, ours is probably one of the rarest reports of recurrent vaginal E. vermicularis infection in the absence of any gastrointestinal symptoms despite complete treatment. A 4-year-old girl presented with recurrent episodes of vulval itching on 3-4 occasions over 2 years. There was no pruritis ani nor urinary/gastrointestinal complaints. The vulva was inflamed with 4-5 living worms, 6-7 mm in length, emerging from the anterior vaginal fornix, but with no vaginal discharge. Direct microscopic examination of vaginal swabs revealed adult worms of Enterobius but no eggs. Repeated stool samples from the patient, parents and a sibling were negative. The patient was treated orally with 100 mg of mebendazole for 3 days followed by two more courses at 3-week intervals over a period of 3 months. Recurrent vaginal enterobiasis despite complete treatment and in the absence of any gastrointestinal involvement suggests that the vagina is a potential reservoir for E. vermicularis, which supports the theory of rare ectopic enterobiasis through the ascending pathway of the female genital tract.

  7. Effects of actonomycin D and ultraviolet irradiation on multiplication of brome mosaic virus in host and non-host cells

    International Nuclear Information System (INIS)

    Maekawa, K.; Furusawa, I.; Okuno, T.

    1981-01-01

    The modes of multiplication of brome mosaic virus (BMV) were compared in protoplasts isolated from host and non-host plants. BMV actively multiplied in the leaves and isolated mesophyll protoplasts of barley, a host of BMV. BMV multiplication in barley protoplasts was inhibited by addition of actinomycin D immediately after inoculation or by u.v. irradiation of the protoplasts before inoculation. In contrast, although BMV could not multiply in leaves of radish and turnip (non-hosts for BMV) it multiplied at a low level in protoplasts isolated from these two plant species. Moreover, u.v. irradiation, or the addition of actinomycin D, enhanced multiplication of BMV in radish and turnip protoplasts. These results suggest that (i) in the host cells replication of BMV is dependent on cellular metabolism of nucleic acid and protein, and (ii) in the non-host cells a substance(s) inhibitory to replication of BMV is synthesized. (author)

  8. Tracing Conidial Fate and Measuring Host Cell Antifungal Activity Using a Reporter of Microbial Viability in the Lung

    Directory of Open Access Journals (Sweden)

    Anupam Jhingran

    2012-12-01

    Full Text Available Fluorescence can be harnessed to monitor microbial fate and to investigate functional outcomes of individual microbial cell-host cell encounters at portals of entry in native tissue environments. We illustrate this concept by introducing fluorescent Aspergillus reporter (FLARE conidia that simultaneously report phagocytic uptake and fungal viability during cellular interactions with the murine respiratory innate immune system. Our studies using FLARE conidia reveal stepwise and cell-type-specific requirements for CARD9 and Syk, transducers of C-type lectin receptor and integrin signals, in neutrophil recruitment, conidial uptake, and conidial killing in the lung. By achieving single-event resolution in defined leukocyte populations, the FLARE method enables host cell profiling on the basis of pathogen uptake and killing and may be extended to other pathogens in diverse model host organisms to query molecular, cellular, and pharmacologic mechanisms that shape host-microbe interactions.

  9. Hijacked then lost in translation: the plight of the recombinant host cell in membrane protein structural biology projects.

    Science.gov (United States)

    Bill, Roslyn M; von der Haar, Tobias

    2015-06-01

    Membrane protein structural biology is critically dependent upon the supply of high-quality protein. Over the last few years, the value of crystallising biochemically characterised, recombinant targets that incorporate stabilising mutations has been established. Nonetheless, obtaining sufficient yields of many recombinant membrane proteins is still a major challenge. Solutions are now emerging based on an improved understanding of recombinant host cells; as a 'cell factory' each cell is tasked with managing limited resources to simultaneously balance its own growth demands with those imposed by an expression plasmid. This review examines emerging insights into the role of translation and protein folding in defining high-yielding recombinant membrane protein production in a range of host cells. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  10. Roles of the Picornaviral 3C Proteinase in the Viral Life Cycle and Host Cells

    Directory of Open Access Journals (Sweden)

    Di Sun

    2016-03-01

    Full Text Available The Picornaviridae family comprises a large group of non-enveloped viruses that have a major impact on human and veterinary health. The viral genome contains one open reading frame encoding a single polyprotein that can be processed by viral proteinases. The crucial 3C proteinases (3Cpros of picornaviruses share similar spatial structures and it is becoming apparent that 3Cpro plays a significant role in the viral life cycle and virus host interaction. Importantly, the proteinase and RNA-binding activity of 3Cpro are involved in viral polyprotein processing and the initiation of viral RNA synthesis. In addition, 3Cpro can induce the cleavage of certain cellular factors required for transcription, translation and nucleocytoplasmic trafficking to modulate cell physiology for viral replication. Due to interactions between 3Cpro and these essential factors, 3Cpro is also involved in viral pathogenesis to support efficient infection. Furthermore, based on the structural conservation, the development of irreversible inhibitors and discovery of non-covalent inhibitors for 3Cpro are ongoing and a better understanding of the roles played by 3Cpro may provide insights into the development of potential antiviral treatments. In this review, the current knowledge regarding the structural features, multiple functions in the viral life cycle, pathogen host interaction, and development of antiviral compounds for 3Cpro is summarized.

  11. Als3 is a Candida albicans invasin that binds to cadherins and induces endocytosis by host cells.

    Directory of Open Access Journals (Sweden)

    Quynh T Phan

    2007-03-01

    Full Text Available Candida albicans is the most common cause of hematogenously disseminated and oropharyngeal candidiasis. Both of these diseases are characterized by fungal invasion of host cells. Previously, we have found that C. albicans hyphae invade endothelial cells and oral epithelial cells in vitro by inducing their own endocytosis. Therefore, we set out to identify the fungal surface protein and host cell receptors that mediate this process. We found that the C. albicans Als3 is required for the organism to be endocytosed by human umbilical vein endothelial cells and two different human oral epithelial lines. Affinity purification experiments with wild-type and an als3delta/als3delta mutant strain of C. albicans demonstrated that Als3 was required for C. albicans to bind to multiple host cell surface proteins, including N-cadherin on endothelial cells and E-cadherin on oral epithelial cells. Furthermore, latex beads coated with the recombinant N-terminal portion of Als3 were endocytosed by Chinese hamster ovary cells expressing human N-cadherin or E-cadherin, whereas control beads coated with bovine serum albumin were not. Molecular modeling of the interactions of the N-terminal region of Als3 with the ectodomains of N-cadherin and E-cadherin indicated that the binding parameters of Als3 to either cadherin are similar to those of cadherin-cadherin binding. Therefore, Als3 is a fungal invasin that mimics host cell cadherins and induces endocytosis by binding to N-cadherin on endothelial cells and E-cadherin on oral epithelial cells. These results uncover the first known fungal invasin and provide evidence that C. albicans Als3 is a molecular mimic of human cadherins.

  12. Actin-binding protein regulation by microRNAs as a novel microbial strategy to modulate phagocytosis by host cells: the case of N-Wasp and miR-142-3p.

    Science.gov (United States)

    Bettencourt, Paulo; Marion, Sabrina; Pires, David; Santos, Leonor F; Lastrucci, Claire; Carmo, Nuno; Blake, Jonathon; Benes, Vladimir; Griffiths, Gareth; Neyrolles, Olivier; Lugo-Villarino, Geanncarlo; Anes, Elsa

    2013-01-01

    Mycobacterium tuberculosis (Mtb) is a successful intracellular pathogen that thrives in macrophages (Mφs). There is a need to better understand how Mtb alters cellular processes like phagolysosome biogenesis, a classical determinant of its pathogenesis. A central feature of this bacteria's strategy is the manipulation of Mφ actin. Here, we examined the role of microRNAs (miRNAs) as a potential mechanism in the regulation of actin-mediated events leading to phagocytosis in the context of mycobacteria infection. Given that non-virulent Mycobacterium smegmatis also controls actin filament assembly to prolong its intracellular survival inside host cells, we performed a global transcriptomic analysis to assess the modulation of miRNAs upon M. smegmatis infection of the murine Mφ cell line, J774A.1. This approach identified miR-142-3p as a key candidate to be involved in the regulation of actin dynamics required in phagocytosis. We unequivocally demonstrate that miR-142-3p targets N-Wasp, an actin-binding protein required during microbial challenge. A gain-of-function approach for miR-142-3p revealed a down-regulation of N-Wasp expression accompanied by a decrease of mycobacteria intake, while a loss-of-function approach yielded the reciprocal increase of the phagocytosis process. Equally important, we show Mtb induces the early expression of miR-142-3p and partially down-regulates N-Wasp protein levels in both the murine J774A.1 cell line and primary human Mφs. As proof of principle, the partial siRNA-mediated knock down of N-Wasp resulted in a decrease of Mtb intake by human Mφs, reflected in lower levels of colony-forming units (CFU) counts over time. We therefore propose the modulation of miRNAs as a novel strategy in mycobacterial infection to control factors involved in actin filament assembly and other early events of phagolysosome biogenesis.

  13. Ultrasound scan as a potential source of nosocomial and cross-infection: a literature review

    Directory of Open Access Journals (Sweden)

    André Hadyme Miyague

    2015-10-01

    Full Text Available AbstractThe authors review the main concepts regarding the importance of cleaning/disinfection of ultrasonography probes, aiming a better comprehension by practitioners and thus enabling strategies to establish a safe practice without compromising the quality of the examination and the operator productivity. In the context of biosafety, it is imperative to assume that contact with blood or body fluids represents a potential source of infection. Thus, in order to implement cleaning/disinfection practice, it is necessary to understand the principles of infection control, to consider the cost/benefit ratio of the measures to be implemented, and most importantly, to comprehend that such measures will not only benefit the health professional and the patient, but the society as a whole.

  14. Potentially Preventable Hospitalizations and the Burden of Healthcare-Associated Infections

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    Andrea L. Lorden

    2017-08-01

    Full Text Available Background: An estimated 4% of hospital admissions acquired healthcare-associated infections (HAIs and accounted for $9.8 (USD billion in direct cost during 2011. In 2010, nearly 140 000 of the 3.5 million potentially preventable hospitalizations (PPHs may have acquired an HAI. There is a knowledge gap regarding the co-occurrence of these events. Aims: To estimate the period occurrences and likelihood of acquiring an HAI for the PPH population. Methods: Retrospective, cross-sectional study using logistic regression analysis of 2011 Texas Inpatient Discharge Public Use Data File including 2.6 million admissions from 576 acute care hospitals. Agency for Healthcare Research and Quality Prevention Quality Indicator software identified PPH, and existing administrative data identification methodologies were refined for Clostridium difficile infection, central line–associated bloodstream infection, catheter-associated urinary tract infection, and ventilator-associated pneumonia. Odds of acquiring HAIs when admitted with PPH were adjusted for demographic, health status, hospital, and community characteristics. Findings: We identified 272 923 PPH, 14 219 HAI, and 986 admissions with PPH and HAI. Odds of acquiring an HAI for diabetic patients admitted for lower extremity amputation demonstrated significantly increased odds ratio of 2.9 (95% confidence interval: 2.16-3.91 for Clostridium difficile infection. Other PPH patients had lower odds of acquiring HAI compared to non-PPH patients, and results were frequently significant. Conclusions: Clinical implications include increased risk of HAI among diabetic patients admitted for lower extremity amputation. Methodological implications include identification of rare events for inpatient subpopulations and the need for improved codification of HAIs to improve cost and policy analyses regarding allocation of resources toward clinical improvements.

  15. A translocated effector required for Bartonella dissemination from derma to blood safeguards migratory host cells from damage by co-translocated effectors.

    Science.gov (United States)

    Okujava, Rusudan; Guye, Patrick; Lu, Yun-Yueh; Mistl, Claudia; Polus, Florine; Vayssier-Taussat, Muriel; Halin, Cornelia; Rolink, Antonius G; Dehio, Christoph

    2014-06-01

    Numerous bacterial pathogens secrete multiple effectors to modulate host cellular functions. These effectors may interfere with each other to efficiently control the infection process. Bartonellae are Gram-negative, facultative intracellular bacteria using a VirB type IV secretion system to translocate a cocktail of Bartonella effector proteins (Beps) into host cells. Based on in vitro infection models we demonstrate here that BepE protects infected migratory cells from injurious effects triggered by BepC and is required for in vivo dissemination of bacteria from the dermal site of inoculation to blood. Human endothelial cells (HUVECs) infected with a ΔbepE mutant of B. henselae (Bhe) displayed a cell fragmentation phenotype resulting from Bep-dependent disturbance of rear edge detachment during migration. A ΔbepCE mutant did not show cell fragmentation, indicating that BepC is critical for triggering this deleterious phenotype. Complementation of ΔbepE with BepEBhe or its homologues from other Bartonella species abolished cell fragmentation. This cyto-protective activity is confined to the C-terminal Bartonella intracellular delivery (BID) domain of BepEBhe (BID2.EBhe). Ectopic expression of BID2.EBhe impeded the disruption of actin stress fibers by Rho Inhibitor 1, indicating that BepE restores normal cell migration via the RhoA signaling pathway, a major regulator of rear edge retraction. An intradermal (i.d.) model for B. tribocorum (Btr) infection in the rat reservoir host mimicking the natural route of infection by blood sucking arthropods allowed demonstrating a vital role for BepE in bacterial dissemination from derma to blood. While the Btr mutant ΔbepDE was abacteremic following i.d. inoculation, complementation with BepEBtr, BepEBhe or BIDs.EBhe restored bacteremia. Given that we observed a similar protective effect of BepEBhe on infected bone marrow-derived dendritic cells migrating through a monolayer of lymphatic endothelial cells we propose that

  16. A translocated effector required for Bartonella dissemination from derma to blood safeguards migratory host cells from damage by co-translocated effectors.

    Directory of Open Access Journals (Sweden)

    Rusudan Okujava

    2014-06-01

    Full Text Available Numerous bacterial pathogens secrete multiple effectors to modulate host cellular functions. These effectors may interfere with each other to efficiently control the infection process. Bartonellae are Gram-negative, facultative intracellular bacteria using a VirB type IV secretion system to translocate a cocktail of Bartonella effector proteins (Beps into host cells. Based on in vitro infection models we demonstrate here that BepE protects infected migratory cells from injurious effects triggered by BepC and is required for in vivo dissemination of bacteria from the dermal site of inoculation to blood. Human endothelial cells (HUVECs infected with a ΔbepE mutant of B. henselae (Bhe displayed a cell fragmentation phenotype resulting from Bep-dependent disturbance of rear edge detachment during migration. A ΔbepCE mutant did not show cell fragmentation, indicating that BepC is critical for triggering this deleterious phenotype. Complementation of ΔbepE with BepEBhe or its homologues from other Bartonella species abolished cell fragmentation. This cyto-protective activity is confined to the C-terminal Bartonella intracellular delivery (BID domain of BepEBhe (BID2.EBhe. Ectopic expression of BID2.EBhe impeded the disruption of actin stress fibers by Rho Inhibitor 1, indicating that BepE restores normal cell migration via the RhoA signaling pathway, a major regulator of rear edge retraction. An intradermal (i.d. model for B. tribocorum (Btr infection in the rat reservoir host mimicking the natural route of infection by blood sucking arthropods allowed demonstrating a vital role for BepE in bacterial dissemination from derma to blood. While the Btr mutant ΔbepDE was abacteremic following i.d. inoculation, complementation with BepEBtr, BepEBhe or BIDs.EBhe restored bacteremia. Given that we observed a similar protective effect of BepEBhe on infected bone marrow-derived dendritic cells migrating through a monolayer of lymphatic endothelial cells we

  17. Coronavirus infection of polarized epithelial cells

    NARCIS (Netherlands)

    Rossen, J W; Horzinek, M C; Rottier, P J

    1995-01-01

    Epithelial cells are the first host cells to be infected by incoming c oronaviruses. Recent observations in vitro show that coronaviruses are released from a specific side of these polarized cells, and this polarized release might be important for the spread of the infection in vivo. Mechanisms for

  18. A modified fluorimetric host cell reactivation assay to determine the repair capacity of primary keratinocytes, melanocytes and fibroblasts

    Directory of Open Access Journals (Sweden)

    Gebhard Daniel

    2010-06-01

    Full Text Available Abstract Background The Host Cell Reactivation Assay (HCRA is widely used to identify circumstances and substances affecting the repair capacity of cells, however, it is restricted by the transfection procedure used and the sensitivity of the detection method. Primary skin cells are particularly difficult to transfect, and therefore sensitive methods are needed to detect any variations due to the cell-type or inter-individual differences or changes induced by diverse substances. A sensitive and repeatable method to detect the repair capacity of skin cells would be useful in two different aspects: On the one hand, to identify substances influencing the repair capacity in a positive manner (these substances could be promising ingredients for cosmetic products and on the other hand, to exclude the negative effects of substances on the repair capacity (this could serve as one step further towards replacing or at least reducing animal testing. Results In this paper, we present a rapid and sensitive assay to determine the repair capacity of primary keratinocytes, melanocytes and fibroblasts based on two wave-length Green Fluorescent Protein (GFP and DsRed reporter technology in order to test different substances and their potential to influence the DNA repair capacity. For the detection of plasmid restoration, we used FACS technology, which, in comparison to luminometer technology, is highly sensitive and allows single cell based analysis. The usefulness of this assay and studying the repair capacity is demonstrated by the evidence that DNA repair is repressed by Cyclosporin A in fibroblasts. Conclusions The methodology described in this paper determines the DNA repair capacity in different types of human skin cells. The described transfection protocol is suitable for the transfection of melanocytes, keratinocytes and fibroblasts, reaching efficacies suitable for the detection of the restored plasmids by FACS technology. Therefore the repair capacity

  19. Infection and transmission of Rift Valley fever viruses lacking the NSs and/or NSm genes in mosquitoes: potential role for NSm in mosquito infection.

    Science.gov (United States)

    Crabtree, Mary B; Kent Crockett, Rebekah J; Bird, Brian H; Nichol, Stuart T; Erickson, Bobbie Rae; Biggerstaff, Brad J; Horiuchi, Kalanthe; Miller, Barry R

    2012-01-01

    Rift Valley fever virus is an arthropod-borne human and animal pathogen responsible for large outbreaks of acute and febrile illness throughout Africa and the Arabian Peninsula. Reverse genetics technology has been used to develop deletion mutants of the virus that lack the NSs and/or NSm virulence genes and have been shown to be stable, immunogenic and protective against Rift Valley fever virus infection in animals. We assessed the potential for these deletion mutant viruses to infect and be transmitted by Aedes mosquitoes, which are the principal vectors for maintenance of the virus in nature and emergence of virus initiating disease outbreaks, and by Culex mosquitoes which are important amplification vectors. Aedes aegypti and Culex quinquefasciatus mosquitoes were fed bloodmeals containing the deletion mutant viruses. Two weeks post-exposure mosquitoes were assayed for infection, dissemination, and transmission. In Ae. aegypti, infection and transmission rates of the NSs deletion virus were similar to wild type virus while dissemination rates were significantly reduced. Infection and dissemination rates for the NSm deletion virus were lower compared to wild type. Virus lacking both NSs and NSm failed to infect Ae. aegypti. In Cx. quinquefasciatus, infection rates for viruses lacking NSm or both NSs and NSm were lower than for wild type virus. In both species, deletion of NSm or both NSs and NSm reduced the infection and transmission potential of the virus. Deletion of both NSs and NSm resulted in the highest level of attenuation of virus replication. Deletion of NSm alone was sufficient to nearly abolish infection in Aedes aegypti mosquitoes, indicating an important role for this protein. The double deleted viruses represent an ideal vaccine profile in terms of environmental containment due to lack of ability to efficiently infect and be transmitted by mosquitoes.

  20. Infection and transmission of Rift Valley fever viruses lacking the NSs and/or NSm genes in mosquitoes: potential role for NSm in mosquito infection.

    Directory of Open Access Journals (Sweden)

    Mary B Crabtree

    Full Text Available BACKGROUND: Rift Valley fever virus is an arthropod-borne human and animal pathogen responsible for large outbreaks of acute and febrile illness throughout Africa and the Arabian Peninsula. Reverse genetics technology has been used to develop deletion mutants of the virus that lack the NSs and/or NSm virulence genes and have been shown to be stable, immunogenic and protective against Rift Valley fever virus infection in animals. We assessed the potential for these deletion mutant viruses to infect and be transmitted by Aedes mosquitoes, which are the principal vectors for maintenance of the virus in nature and emergence of virus initiating disease outbreaks, and by Culex mosquitoes which are important amplification vectors. METHODOLOGY AND PRINCIPAL FINDINGS: Aedes aegypti and Culex quinquefasciatus mosquitoes were fed bloodmeals containing the deletion mutant viruses. Two weeks post-exposure mosquitoes were assayed for infection, dissemination, and transmission. In Ae. aegypti, infection and transmission rates of the NSs deletion virus were similar to wild type virus while dissemination rates were significantly reduced. Infection and dissemination rates for the NSm deletion virus were lower compared to wild type. Virus lacking both NSs and NSm failed to infect Ae. aegypti. In Cx. quinquefasciatus, infection rates for viruses lacking NSm or both NSs and NSm were lower than for wild type virus. CONCLUSIONS/SIGNIFICANCE: In both species, deletion of NSm or both NSs and NSm reduced the infection and transmission potential of the virus. Deletion of both NSs and NSm resulted in the highest level of attenuation of virus replication. Deletion of NSm alone was sufficient to nearly abolish infection in Aedes aegypti mosquitoes, indicating an important role for this protein. The double deleted viruses represent an ideal vaccine profile in terms of environmental containment due to lack of ability to efficiently infect and be transmitted by mosquitoes.

  1. Role of therapeutic drug monitoring in pulmonary infections : use and potential for expanded use of dried blood spot samples

    NARCIS (Netherlands)

    Hofman, Susan; Bolhuis, Mathieu S.; Koster, Remco A.; Akkerman, Onno W.; van Assen, Sander; Stove, Christophe; Alffenaar, Jan-Willem C.

    Respiratory tract infections are among the most common infections in men. We reviewed literature to document their pharmacological treatments, and the extent to which therapeutic drug monitoring (TDM) is needed during treatment. We subsequently examined potential use of dried blood spots as sample

  2. Synovial calprotectin: a potential biomarker to exclude a prosthetic joint infection.

    Science.gov (United States)

    Wouthuyzen-Bakker, M; Ploegmakers, J J W; Kampinga, G A; Wagenmakers-Huizenga, L; Jutte, P C; Muller Kobold, A C

    2017-05-01

    Recently, several synovial biomarkers have been introduced into the algorithm for the diagnosis of a prosthetic joint infection (PJI). Alpha defensin is a promising biomarker, with a high sensitivity and specificity, but it is expensive. Calprotectin is a protein that is present in the cytoplasm of neutrophils, is released upon neutrophil activation and exhibits anti-microbial activity. Our aim, in this study, was to determine the diagnostic potential of synovial calprotectin in the diagnosis of a PJI. In this pilot study, we prospectively collected synovial fluid from the hip, knee, shoulder and elbow of 19 patients with a proven PJI and from a control group of 42 patients who underwent revision surgery without a PJI. PJI was diagnosed according to the current diagnostic criteria of the Musculoskeletal Infection Society. Synovial fluid was centrifuged and the supernatant was used to measure the level of calprotectin after applying a lateral flow immunoassay. The median synovial calprotectin level was 991 mg/L (interquartile range (IQR) 154 to 1787) in those with a PJI and 11 mg/L (IQR 3 to 29) in the control group (p infection. With a lateral flow immunoassay, a relatively rapid quantitative diagnosis can be made. The measurement is cheap and is easy to use. Cite this article: Bone Joint J 2017;99-B:660-5. ©2017 The British Editorial Society of Bone & Joint Surgery.

  3. Adipose Tissue-Derived Mesenchymal Stem Cells as a New Host Cell in Latent Leishmaniasis

    Science.gov (United States)

    Allahverdiyev, Adil M.; Bagirova, Melahat; Elcicek, Serhat; Koc, Rabia Cakir; Baydar, Serap Yesilkir; Findikli, Necati; Oztel, Olga N.

    2011-01-01

    Some protozoan infections such as Toxoplasma, Cryptosporidium, and Plasmodium can be transmitted through stem cell transplantations. To our knowledge, so far, there is no study about transmission of Leishmania parasites in stem cell transplantation and interactions between parasites and stem cells in vitro. Therefore, the aim of this study was to investigate the interaction between different species of Leishmania parasites and adipose tissue-derived mesenchymal stem cells (ADMSCs). ADMSCs have been isolated, cultured, characterized, and infected with different species of Leishmania parasites (L. donovani, L. major, L. tropica, and L. infantum). Infectivity was examined by Giemsa staining, microculture, and polymerase chain reaction methods. As a result, infectivity of ADMSCs by Leishmania parasites has been determined for the first time in this study. According to our findings, it is very important that donors are screened for Leishmania parasites before stem cell transplantations in regions where leishmaniasis is endemic. PMID:21896818

  4. Surface localization of glucosylceramide during Cryptococcus neoformans infection allows targeting as a potential antifungal.

    Directory of Open Access Journals (Sweden)

    Ryan Rhome

    Full Text Available Cryptococcus neoformans (Cn is a significant human pathogen that, despite current treatments, continues to have a high morbidity rate especially in sub-Saharan Africa. The need for more tolerable and specific therapies has been clearly shown. In the search for novel drug targets, the gene for glucosylceramide synthase (GCS1 was deleted in Cn, resulting in a strain (Δgcs1 that does not produce glucosylceramide (GlcCer and is avirulent in mouse models of infection. To understand the biology behind the connection between virulence and GlcCer, the production and localization of GlcCer must be characterized in conditions that are prohibitive to the growth of Δgcs1 (neutral pH and high CO(2. These prohibitive conditions are physiologically similar to those found in the extracellular spaces of the lung during infection. Here, using immunofluorescence, we have shown that GlcCer localization to the cell surface is significantly increased during growth in these conditions and during infection. We further seek to exploit this localization by treatment with Cerezyme (Cz, a recombinant enzyme that metabolizes GlcCer, as a potential treatment for Cn. Cz treatment was found to reduce the amount of GlcCer in vitro, in cultures, and in Cn cells inhabiting the mouse lung. Treatment with Cz induced a membrane integrity defect in wild type Cn cells similar to Δgcs1. Cz treatment also reduced the in vitro growth of Cn in a dose and condition dependent manner. Finally, Cz treatment was shown to have a protective effect on survival in mice infected with Cn. Taken together, these studies have established the legitimacy of targeting the GlcCer and other related sphingolipid systems in the development of novel therapeutics.

  5. Clinical definition of respiratory viral infections in young children and potential bronchiolitis misclassification.

    Science.gov (United States)

    Megalaa, Rosemary; Perez, Geovanny F; Kilaikode-Cheruveettara, Sasikumar; Kotwal, Nidhi; Rodriguez-Martinez, Carlos E; Nino, Gustavo

    2018-01-01

    Viral respiratory infections are often grouped as a single respiratory syndrome named 'viral bronchiolitis', independently of the viral etiology or individual risk factors. Clinical trials and guidelines have used a more stringent definition of viral bronchiolitis, including only the first episode of wheezing in children less than 12 months of age without concomitant respiratory comorbidities. There is increasing evidence suggesting that this definition is not being followed by pediatric care providers, but it is unclear to what extent viral respiratory infections are currently misclassified as viral bronchiolitis using standard definitions. We conducted a retrospective analysis of hospitalized young children (≤3 years) due to viral respiratory infections. Bronchiolitis was defined as the first wheezing episode less than 12 months of age. Demographic variables and comorbidities were obtained by electronic medical record review. The study comprised a total of 513 hospitalizations (n=453). Viral bronchiolitis was diagnosed in 144 admissions (28.1%). Notably, we identified that the majority of children diagnosed with bronchiolitis (63%) were misclassified as they had prior episodes of wheezing. Many children with bronchiolitis misclassification had significant comorbidities, including prematurity (51%), neuromuscular conditions (9.8%), and congenital heart disease (9.8%). Misclassification of bronchiolitis is a common problem that may lead to inappropriate management of viral respiratory infections in young children. A comprehensive approach that takes into consideration viral etiology and individual risk factors may lead to a more accurate clinical assessment of this condition and would potentially prevent bronchiolitis misclassification. © American Federation for Medical Research (unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  6. Host-cell reactivation of UV-irradiated and chemically-treated herpes simplex virus-1 by xeroderma pigmentosum, xp heterozygotes and normal skin fibroblasts

    International Nuclear Information System (INIS)

    Selsky, C.A.

    1978-01-01

    The host-cell reactivation of UV-irradiated and N-acetoxy-2-acetylamino-fluorene-treated herpes simplex virus type 1 strain MP was studied in normal and xeroderma pigmentosum human skin fibroblasts. Virus treated with either agent demonstrated lower survival in XP cells from complementation groups A, B, C and D than in normal fibroblasts. The relative reactivation ability of XP cells from the different genetic complementation groups was found to be the same for both irradiated and chemically treated virus. In addition, the inactivation kinetics for virus treated with either agent in the XP variant were comparable to that seen in normal skin fibroblasts. The addition of 2 or 4 mmoles caffeine to the post-infection assay medium had no effect on the inactivation kinetics of virus treated by either agent in the XP variant or in XP cells from the different genetic complementation groups. Treatment of the virus with nitrogen mustard resulted in equivalent survival in normal and XP genetic complementation group D cells. No apparent defect was observed in the ability of XP heterozygous skin fibroblasts to repair virus damaged with up to 100 μg N-acetoxy-2-acetylaminofluorene per ml. These findings indicate that the repair of UV-irradiated and N-acetoxy-2-acetylaminofluorene-treated virus is accomplished by the same pathway or different pathways sharing a common intermediate step and that the excision defect of XP cells plays little if any role in the reactivation of nitrogen mustard treated virus. (Auth.)

  7. Anaplasma phagocytophilum MSP4 and HSP70 Proteins Are Involved in Interactions with Host Cells during Pathogen Infection

    Czech Academy of Sciences Publication Activity Database

    Contreras, M.; Alberdi, P.; Mateos-Hernández, L.; Fernández de Mera, I.G.; García-Pérez, A. L.; Vancová, Marie; Villar, M.; Ayllón, N.; Cabezas Cruz, Alejandro; Valdés, James J.; Stuen, S.; Gortazar, C.; de la Fuente, J.

    2017-01-01

    Roč. 7, JUL 5 (2017), č. článku 307. ISSN 2235-2988 EU Projects: European Commission(XE) 278976 - ANTIGONE Institutional support: RVO:60077344 Keywords : anaplasmosis * immunology * hl60 * tick * vaccine * sheep * Anaplasma phagocytophilum Subject RIV: EC - Immunology OBOR OECD: Immunology Impact factor: 4.300, year: 2016

  8. Universal newborn screening for congenital CMV infection: what is the evidence of potential benefit?

    Science.gov (United States)

    Cannon, Michael J; Griffiths, Paul D; Aston, Van; Rawlinson, William D

    2014-09-01

    Congenital CMV infection is a leading cause of childhood disability. Many children born with congenital CMV infection are asymptomatic or have nonspecific symptoms and therefore are typically not diagnosed. A strategy of newborn CMV screening could allow for early detection and intervention to improve clinical outcomes. Interventions might include antiviral drugs or nonpharmaceutical therapies such as speech-language therapy or cochlear implants. Using published data from developed countries, we analyzed existing evidence of potential benefit that could result from newborn CMV screening. We first estimated the numbers of children with the most important CMV-related disabilities (i.e. hearing loss, cognitive deficit, and vision impairment), including the age at which the disabilities occur. Then, for each of the disabilities, we examined the existing evidence for the effectiveness of various interventions. We concluded that there is good evidence of potential benefit from nonpharmaceutical interventions for children with delayed hearing loss that occurs by 9 months of age. Similarly, we concluded that there is fair evidence of potential benefit from antiviral therapy for children with hearing loss at birth and from nonpharmaceutical interventions for children with delayed hearing loss occurring between 9 and 24 months of age and for children with CMV-related cognitive deficits. We found poor evidence of potential benefit for children with delayed hearing loss occurring after 24 months of age and for children with vision impairment. Overall, we estimated that in the United States, several thousand children with congenital CMV could benefit each year from newborn CMV screening, early detection, and interventions. Copyright © 2014 John Wiley & Sons, Ltd.

  9. Universal newborn screening for congenital CMV infection: what is the evidence of potential benefit?†

    Science.gov (United States)

    Cannon, Michael J.; Griffiths, Paul D.; Aston, Van; Rawlinson, William D.

    2015-01-01

    SUMMARY Congenital CMV infection is a leading cause of childhood disability. Many children born with congenital CMV infection are asymptomatic or have nonspecific symptoms and therefore are typically not diagnosed. A strategy of newborn CMV screening could allow for early detection and intervention to improve clinical outcomes. Interventions might include antiviral drugs or nonpharmaceutical therapies such as speech-language therapy or cochlear implants. Using published data from developed countries, we analyzed existing evidence of potential benefit that could result from newborn CMV screening. We first estimated the numbers of children with the most important CMV-related disabilities (i.e. hearing loss, cognitive deficit, and vision impairment), including the age at which the disabilities occur. Then, for each of the disabilities, we examined the existing evidence for the effectiveness of various interventions. We concluded that there is good evidence of potential benefit from nonpharmaceutical interventions for children with delayed hearing loss that occurs by 9 months of age. Similarly, we concluded that there is fair evidence of potential benefit from antiviral therapy for children with hearing loss at birth and from nonpharmaceutical interventions for children with delayed hearing loss occurring between 9 and 24 months of age and for children with CMV-related cognitive deficits. We found poor evidence of potential benefit for children with delayed hearing loss occurring after 24 months of age and for children with vision impairment. Overall, we estimated that in the United States, several thousand children with congenital CMV could benefit each year from newborn CMV screening, early detection, and interventions. Copyright © 2014 John Wiley & Sons, Ltd. PMID:24760655

  10. Unique physiology of host-parasite interactions in microsporidia infections.

    Science.gov (United States)

    Williams, Bryony A P

    2009-11-01

    Microsporidia are intracellular parasites of all major animal lineages and have a described diversity of over 1200 species and an actual diversity that is estimated to be much higher. They are important pathogens of mammals, and are now one of the most common infections among immunocompromised humans. Although related to fungi, microsporidia are atypical in genomic biology, cell structure and infection mechanism. Host cell infection involves the rapid expulsion of a polar tube from a dormant spore to pierce the host cell membrane and allow the direct transfer of the spore contents into the host cell cytoplasm. This intimate relationship between parasite and host is unique. It allows the microsporidia to be highly exploitative of the host cell environment and cause such diverse effects as the induction of hypertrophied cells to harbour prolific spore development, host sex ratio distortion and host cell organelle and microtubule reorganization. Genome sequencing has revealed that microsporidia have achieved this high level of parasite sophistication with radically reduced proteomes and with many typical eukaryotic pathways pared-down to what appear to be minimal functional units. These traits make microsporidia intriguing model systems for understanding the extremes of reductive parasite evolution and host cell manipulation.

  11. Transport pathways in the malaria-infected erythrocyte: characterization and their use as potential targets for chemotherapy

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    Hagai Ginsburg

    1994-01-01

    Full Text Available The intraerythrocytic malarial parasite is involved in an extremely intensive anabolic activity while it resides in its metabolically quiescent host cell. The necessary fast uptake of nutrients and the discharge of waste product, are guaranteed by parasite-induced alterations of the constitutive transporters of the host cell and the production of new parallel pathways. The membrane of the host cell thus becomes permeable to phospholipids, purine bases and nucleosides, small non-electrolytes, anions and cations. When the new pathways are quantitatively unimportant, classical inhibitors of native transporters can be used to inhibit parasite growth. Several compounds were found to effectively inhibit the new pathways and consequently, parasite growth. The pathways have also been used to introduce cytotoxic agents. The parasitophorous membrane consists of channels which are highly permeable to small solutes and display no ion selectivity. Transport of some cations and anions across the parasite membrane is rapid and insensitive to classical inhibitors, and in some cases it is mediated by specific antiporters which respond to their respective inhibitors. Macromolecules have been shown to reach the parasitophorous space through a duct contiguous with the host cell membrane, and subsequently to be endocytosed at the parasite membrane. The simultaneous presence of the parasitophorous membrane channels and the duct, however, is incompatible with experimental evidences. No specific inhibitors were found as yet that would efficiently inhibit transport through the channels or the duct.

  12. Characteristics and quantities of HIV host cells in human genital tract secretions.

    Science.gov (United States)

    Politch, Joseph A; Marathe, Jai; Anderson, Deborah J

    2014-12-15

    Human immunodeficiency virus (HIV)-infected leukocytes have been detected in genital secretions from HIV-infected men and women and may play an important role in the sexual transmission of HIV. However, they have been largely overlooked in studies on mechanisms of HIV transmission and in the design and testing of HIV vaccine and microbicide candidates. This article describes the characteristics and quantities of leukocytes in male and female genital secretions under various conditions and also reviews evidence for the involvement of HIV-infected cells in both horizontal and vertical cell-associated HIV transmission. Additional research is needed in this area to better target HIV prevention strategies. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. The potential role of stethoscopes as a source of nosocomial infection

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    Neetu Gupta

    2014-01-01

    Full Text Available Background: Infectious diseases can be transmitted in various ways . Indirect transmission or vehicle-borne transmission of infection can occur through various agents like instruments, utensils, water, or food. In hospital settings, transmission of infection through contaminated medical devices is always a possibility. Stethoscope is the symbol of health professionals. However, the diaphragm and bells of the stethoscopes have been shown to harbor pathogenic and nonpathogenic microorganisms. Aims: 1 To determine the degree of contamination of stethoscopes used by clinicians and nurses. 2 To identify various microbes from such contaminated stethoscopes and to assess their role as potential pathogens. 3 To determine the effectiveness of 70% ethanol as a disinfecting agent. Materials and Methods: Stethoscopes of 50 healthcare workers from different departments were included in the study. Bacteriological cultures of the samples were done on blood agar and MacConkey agar plates. Organisms were identified by conventional phenotypic methods. Antibiotic sensitivity test (ABST of the microorganisms was performed by Kirby-Bauer disk diffusion method. Results: Average number of colonies on each stethoscope was 30 . Pathogenic microorganisms including Staphylococcus aureus, Acinetobacter species, Citrobacter species, Pseudomonas stutzeri, Bacillus species, and Aspergillus fumigatus were identified. Among the potential pathogenic organisms, coagulase-negative staphylococci were isolated. Numbers of colonies were highest among the stethoscopes sampled from surgery ward and Medical Intensive Care Unit MICU. Conclusion: Stethoscopes harbor potential pathogens capable of surviving on their surfaces. Despite their best intentions, health professionals can act as a vector for transmission of disease. Disinfecting procedures may be necessary between consecutive patients. Attention to simple preventive strategies can reduce the disease transmission rate.

  14. Polymeric immunoglobulin receptor-mediated invasion of Streptococcus pneumoniae into host cells requires a coordinate signaling of SRC family of protein-tyrosine kinases, ERK, and c-Jun N-terminal kinase.

    Science.gov (United States)

    Agarwal, Vaibhav; Asmat, Tauseef M; Dierdorf, Nina I; Hauck, Christof R; Hammerschmidt, Sven

    2010-11-12

    Streptococcus pneumoniae are commensals of the human nasopharynx with the capacity to invade mucosal respiratory cells. PspC, a pneumococcal surface protein, interacts with the human polymeric immunoglobulin receptor (pIgR) to promote bacterial adherence to and invasion into epithelial cells. Internalization of pneumococci requires the coordinated action of actin cytoskeleton rearrangements and the retrograde machinery of pIgR. Here, we demonstrate the involvement of Src protein-tyrosine kinases (PTKs), focal adhesion kinase (FAK), extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK) but not p38 mitogen-activated protein kinases (MAPK) in pneumococcal invasion via pIgR. Pharmacological inhibitors of PTKs and MAPKs and genetic interference with Src PTK and FAK functions caused a significant reduction of pIgR-mediated pneumococcal invasion but did not influence bacterial adhesion to host cells. Furthermore, pneumococcal ingestion by host cells induces activation of ERK1/2 and JNK. In agreement with activated JNK, its target molecule and DNA-binding protein c-Jun was phosphorylated. We also show that functionally active Src PTK is essential for activation of ERK1/2 upon pneumococcal infections. In conclusion, these data illustrate the importance of a coordinated signaling between Src PTKs, ERK1/2, and JNK during PspC-pIgR-mediated uptake of pneumococci by host epithelial cells.

  15. The guanylhydrazone CNI-1493: an inhibitor with dual activity against malaria-inhibition of host cell pro-inflammatory cytokine release and parasitic deoxyhypusine synthase.

    Science.gov (United States)

    Specht, Sabine; Sarite, Salem Ramadan; Hauber, Ilona; Hauber, Joachim; Görbig, Ulf F; Meier, Chris; Bevec, Dorian; Hoerauf, Achim; Kaiser, Annette

    2008-05-01

    Malaria is still a major cause of death in the tropics. There is an urgent need for new anti-malarial drugs because drug-resistant plasmodia frequently occur. Over recent years, we elucidated the biosynthesis of hypusine, a novel amino acid contained in eukaryotic initiation factor 5A (eIF-5A) in Plasmodium. Hypusine biosynthesis involves catalysis of deoxyhypusine synthase (DHS) in the first step of post-translational modification. In a screen for new inhibitors of purified plasmodium DHS, CNI-1493, a novel selective pro-inflammatory cytokine inhibitor used in clinical phase II for the treatment of Crohn's disease, inhibited the enzyme of the parasite 3-fold at a concentration of 2 microM. In vitro experiments with 200 microM CNI-1493 in Plasmodium-infected erythrocytes, which lack nuclei and DHS protein, showed a parasite clearance within 2 days. This can presumably be attributed to an anti-proliferating effect because of the inhibition of DHS by the parasite. The determined IC50 of CNI-1493 was 135.79 microM after 72 h. In vivo application of this substance in Plasmodium berghei ANKA-infected C57BL/6 mice significantly reduced parasitemia after dosage of 1 mg/kg or 4 mg/kg/body weight and prevented death of mice with cerebral malaria. This effect was paralleled by a decrease in serum TNF levels of the mice. We suggest that the new mechanism of CNI-1493 is caused by a decrease in modified eIF-5A biosynthesis with a downstream effect on the TNF synthesis of the host. From the current data, we consider CNI-1493 to be a promising drug for anti-malarial therapy because of its combined action, i.e., the decrease in eIF-5A biosynthesis of the parasite and host cell TNF biosynthesis.

  16. Antimicrobial Peptides and Their Therapeutic Potential for Bacterial Skin Infections and Wounds

    Science.gov (United States)

    Pfalzgraff, Anja; Brandenburg, Klaus; Weindl, Günther

    2018-01-01

    Alarming data about increasing resistance to conventional antibiotics are reported, while at the same time the development of new antibiotics is stagnating. Skin and soft tissue infections (SSTIs) are mainly caused by the so called ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) which belong to the most recalcitrant bacteria and are resistant to almost all common antibiotics. S. aureus and P. aeruginosa are the most frequent pathogens isolated from chronic wounds and increasing resistance to topical antibiotics has become a major issue. Therefore, new treatment options are urgently needed. In recent years, research focused on the development of synthetic antimicrobial peptides (AMPs) with lower toxicity and improved activity compared to their endogenous counterparts. AMPs appear to be promising therapeutic options for the treatment of SSTIs and wounds as they show a broad spectrum of antimicrobial activity, low resistance rates and display pivotal immunomodulatory as well as wound healing promoting activities such as induction of cell migration and proliferation and angiogenesis. In this review, we evaluate the potential of AMPs for the treatment of bacterial SSTIs and wounds and provide an overview of the mechanisms of actions of AMPs that contribute to combat skin infections and to improve wound healing. Bacteria growing in biofilms are more resistant to conventional antibiotics than their planktonic counterparts due to limited biofilm penetration and distinct metabolic and physiological functions, and often result in chronification of infections and wounds. Thus, we further discuss the feasibility of AMPs as anti-biofilm agents. Finally, we highlight perspectives for future therapies and which issues remain to bring AMPs successfully to the market. PMID:29643807

  17. The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk-based management for their control.

    Science.gov (United States)

    Bracewell, Daniel G; Francis, Richard; Smales, C Mark

    2015-09-01

    The use of biological systems to synthesize complex therapeutic products has been a remarkable success. However, during product development, great attention must be devoted to defining acceptable levels of impurities that derive from that biological system, heading this list are host cell proteins (HCPs). Recent advances in proteomic analytics have shown how diverse this class of impurities is; as such knowledge and capability grows inevitable questions have arisen about how thorough current approaches to measuring HCPs are. The fundamental issue is how to adequately measure (and in turn monitor and control) such a large number of protein species (potentially thousands of components) to ensure safe and efficacious products. A rather elegant solution is to use an immunoassay (enzyme-linked immunosorbent assay [ELISA]) based on polyclonal antibodies raised to the host cell (biological system) used to synthesize a particular therapeutic product. However, the measurement is entirely dependent on the antibody serum used, which dictates the sensitivity of the assay and the degree of coverage of the HCP spectrum. It provides one summed analog value for HCP amount; a positive if all HCP components can be considered equal, a negative in the more likely event one associates greater risk with certain components of the HCP proteome. In a thorough risk-based approach, one would wish to be able to account for this. These issues have led to the investigation of orthogonal analytical methods; most prominently mass spectrometry. These techniques can potentially both identify and quantify HCPs. The ability to measure and monitor thousands of proteins proportionally increases the amount of data acquired. Significant benefits exist if the information can be used to determine critical HCPs and thereby create an improved basis for risk management. We describe a nascent approach to risk assessment of HCPs based upon such data, drawing attention to timeliness in relation to biosimilar

  18. The future of host cell protein (HCP) identification during process development and manufacturing linked to a risk‐based management for their control

    Science.gov (United States)

    Francis, Richard; Smales, C. Mark

    2015-01-01

    ABSTRACT The use of biological systems to synthesize complex therapeutic products has been a remarkable success. However, during product development, great attention must be devoted to defining acceptable levels of impurities that derive from that biological system, heading this list are host cell proteins (HCPs). Recent advances in proteomic analytics have shown how diverse this class of impurities is; as such knowledge and capability grows inevitable questions have arisen about how thorough current approaches to measuring HCPs are. The fundamental issue is how to adequately measure (and in turn monitor and control) such a large number of protein species (potentially thousands of components) to ensure safe and efficacious products. A rather elegant solution is to use an immunoassay (enzyme‐linked immunosorbent assay [ELISA]) based on polyclonal antibodies raised to the host cell (biological system) used to synthesize a particular therapeutic product. However, the measurement is entirely dependent on the antibody serum used, which dictates the sensitivity of the assay and the degree of coverage of the HCP spectrum. It provides one summed analog value for HCP amount; a positive if all HCP components can be considered equal, a negative in the more likely event one associates greater risk with certain components of the HCP proteome. In a thorough risk‐based approach, one would wish to be able to account for this. These issues have led to the investigation of orthogonal analytical methods; most prominently mass spectrometry. These techniques can potentially both identify and quantify HCPs. The ability to measure and monitor thousands of proteins proportionally increases the amount of data acquired. Significant benefits exist if the information can be used to determine critical HCPs and thereby create an improved basis for risk management. We describe a nascent approach to risk assessment of HCPs based upon such data, drawing attention to timeliness in relation to

  19. Astrocyte Apoptosis and HIV Replication Are Modulated in Host Cells Coinfected with Trypanosoma cruzi

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    Javier M. Urquiza

    2017-08-01

    Full Text Available The protozoan Trypanosoma cruzi is the etiological agent of Chagas disease. In immunosuppressed individuals, as it occurs in the coinfection with human immunodeficiency virus (HIV, the central nervous system may be affected. In this regard, reactivation of Chagas disease is severe and often lethal, and it accounts for meningoencephalitis. Astrocytes play a crucial role in the environment maintenance of healthy neurons; however, they can host HIV and T. cruzi. In this report, human astrocytes were infected in vitro with both genetically modified-pathogens to express alternative fluorophore. As evidenced by fluorescence microscopy and flow cytometry, HIV and T. cruzi coexist in the same astrocyte, likely favoring reciprocal interactions. In this context, lower rates of cell death were observed in both T. cruzi monoinfected-astrocytes and HIV-T. cruzi coinfection in comparison with those infected only with HIV. The level of HIV replication is significantly diminished under T. cruzi coinfection, but without affecting the infectivity of the HIV progeny. This interference with viral replication appears to be related to the T. cruzi multiplication rate or its increased intracellular presence but does not require their intracellular cohabitation or infected cell-to-cell contact. Among several Th1/Th2/Th17 profile-related cytokines, only IL-6 was overexpressed in HIV-T. cruzi coinfection exhibiting its cytoprotective role. This study demonstrates that T. cruzi and HIV are able to coinfect astrocytes thus altering viral replication and apoptosis.

  20. Rotavirus replication is correlated with S/G2 interphase arrest of the host cell cycle.

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    Selene Glück

    Full Text Available In infected cells rotavirus (RV replicates in viroplasms, cytosolic structures that require a stabilized microtubule (MT network for their assembly, maintenance of the structure and perinuclear localization. Therefore, we hypothesized that RV could interfere with the MT-breakdown that takes place in mitosis during cell division. Using synchronized RV-permissive cells, we show that RV infection arrests the cell cycle in S/G2 phase, thus favoring replication by improving viroplasms formation, viral protein translation, and viral assembly. The arrest in S/G2 phase is independent of the host or viral strain and relies on active RV replication. RV infection causes cyclin B1 down-regulation, consistent with blocking entry into mitosis. With the aid of chemical inhibitors, the cytoskeleton network was linked to specific signaling pathways of the RV-induced cell cycle arrest. We found that upon RV infection Eg5 kinesin was delocalized from the pericentriolar region to the viroplasms. We used a MA104-Fucci system to identify three RV proteins (NSP3, NSP5, and VP2 involved in cell cycle arrest in the S-phase. Our data indicate that there is a strong correlation between the cell cycle arrest and RV replication.

  1. Potential risk of norovirus infection due to the consumption of "ready to eat" food.

    Science.gov (United States)

    Serracca, Laura; Rossini, Irene; Battistini, Roberta; Goria, Maria; Sant, Serena; De Montis, Gabriella; Ercolini, Carlo

    2012-09-01

    In this study, we investigated the presence of enteric viruses such as norovirus (NoV), hepatitis A virus (HAV), hepatitis E virus (HEV), and adenovirus (HAdV), in vegetables available on the Italian markets. For this aim, 110 national and international "ready to eat" samples were collected and analyzed by biomolecular tests and positive samples were confirmed by sequencing. All samples (100 %) were negative for HAV, HEV, and HAdV, while 13.6 % (15/110) were positive for NoV. Actually there is not a formal surveillance system for NoV infections in Italy but we clearly demonstrated a potential risk associated with the consumption of "ready to eat" vegetables. This study confirmed for the first time in Italy the presence of norovirus in semi-dried tomatoes by PCR technique.

  2. Host, family and community proxies for infections potentially associated with leukaemia

    International Nuclear Information System (INIS)

    Law, G. R.

    2008-01-01

    Three hypotheses have proposed the involvement of infections in the aetiology of childhood leukaemia, suggesting either a specific leukemogenic infection or a series of common infections that lead to a dys-regulation of the immune system. Much of the evidence for the link with infections has been based on epidemiological observations, often using proxy measures of infection. Proxy measures include population mixing, parental occupation, age distribution of incidence, spatial and space-time clustering of cases, birth order and day care during infancy. This paper discusses the proxies used and examines to what extent a commonly used proxy measure, birth order, is a fair representation of either specific infections or general infectious load. It is clear that although leukaemia, and other diseases, may be linked with infections, one needs to (1) measure specific and general infections with more accuracy and (2) understand how proxy measures relate to real infections in the population. (authors)

  3. Activation of Natural Killer cells during microbial infections

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    Amir eHorowitz

    2012-01-01

    Full Text Available Natural killer (NK cells are large granular lymphocytes that express a diverse array of germline encoded inhibitory and activating receptors for MHC Class I and Class I-like molecules, classical co-stimulatory ligands and cytokines. The ability of NK cells to be very rapidly activated by inflammatory cytokines, to secrete effector cytokines and to kill infected or stressed host cells, suggests that they may be among the very early responders during infection. Recent studies have also identified a small number of pathogen-derived ligands that can bind to NK cell surface receptors and directly induce their activation. Here we review recent studies that have begun to elucidate the various pathways by which viral, bacterial and parasite pathogens activate NK cells. We also consider two emerging themes of NK cell-pathogen interactions, namely their contribution to adaptive immune responses and their potential to take on regulatory and immunomodulatory functions.

  4. Diagnostic accuracy of clinical and blood examination for sepsis in potentially infected neonates

    Directory of Open Access Journals (Sweden)

    Ari Mulyani

    2006-10-01

    Full Text Available Background Neonatal sepsis remains a diagnostic challenge due to its nonspesific symptoms and signs. Blood culture as the gold standard is still a problem because it takes time, is expensive, and not every health facility is able to perionn. Objective To evaluate the diagnostic accuracy of clinical symptoms, hematologic findings, and C-reactive protein (CRP in neonatal sepsis. Methods Samples were taken from potentially infected neonates admitted to the Matemal-Perinatal Unit of Sardjito Hospital, between December 1st, 2000 and March 31st, 2001 using at least one of the criteria: prematurity, very low birth weight infants, matemal pyrexia during delivery, premature membrane rupture, or thick, cloudy amniotic fluid. Clinical symptoms, total leukocyte, neutrophil, platelet count, CRP, and blood culture as the gold standard were examined. Results Among 99 neonates enrolled, the sensitivity, specificity, positive and negative predictive value of clinical symptoms were 79.3%, 75.7%, 57.5%, and 89.9%, respectively; leukopenia/leukocytosis were 27.6%, 85.7%, 44.4%, and 74.1%; neutropenia! neutrophilia were 41.4%, 71.4%, 37.5%, and 74.6%; thrombocytopenia were 79.3%, 51.8%, 40.4%, and 85.7%; positive CRP were 58.6%,78.6%,53.1%, and 82.1%. Parallel tests increased the sensitivity up to 89.7%. Specificity, positive and negative predictive value, and likelihood ratio were 44.3%, 40%, 91.2%, and 1.6, respectively. Serial tests increased the specificity up to 88.6%. Sensitivity, positive and negative predictive value, and likelihood ratio were 58.6%, 68%, 83.8%, and 5.1, respectively. Conclusion Clinical sepsis, thrombocytopenia, and CRP are sufficiently accurate as diagnostic tests for sepsis in potentially infected neonates. Parallel tests will increase the sensitivity, while serial tests increase the specificity.

  5. Identification of Sphingomyelinase on the Surface of Chlamydia pneumoniae: Possible Role in the Entry into Its Host Cells

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    Tuula A. Peñate Medina

    2014-01-01

    Full Text Available We have recently suggested a novel mechanism, autoendocytosis, for the entry of certain microbes into their hosts, with a key role played by the sphingomyelinase-catalyzed topical conversion of sphingomyelin to ceramide, the differences in the biophysical properties of these two lipids providing the driving force. The only requirement for such microbes to utilize this mechanism is that they should have a catalytically active SMase on their outer surface while the target cells should expose sphingomyelin in the external leaflet of their plasma membrane. In pursuit of possible microbial candidates, which could utilize this putative mechanism, we conducted a sequence similarity search for SMase. Because of the intriguing cellular and biochemical characteristics of the poorly understood entry of Chlamydia into its host cells these microbes were of particular interest. SMase activity was measured in vitro from isolated C. pneumoniae elementary bodies (EB and in the lysate from E. coli cells transfected with a plasmid expressing CPn0300 protein having sequence similarity to SMase. Finally, pretreatment of host cells with exogenous SMase resulting in loss plasma membrane sphingomyelin attenuated attachment of EB.

  6. Critical importance of the de novo pyrimidine biosynthesis pathway for Trypanosoma cruzi growth in the mammalian host cell cytoplasm

    International Nuclear Information System (INIS)

    Hashimoto, Muneaki; Morales, Jorge; Fukai, Yoshihisa; Suzuki, Shigeo; Takamiya, Shinzaburo; Tsubouchi, Akiko; Inoue, Syou; Inoue, Masayuki; Kita, Kiyoshi; Harada, Shigeharu; Tanaka, Akiko; Aoki, Takashi; Nara, Takeshi

    2012-01-01

    Highlights: ► We established Trypanosoma cruzi lacking the gene for carbamoyl phosphate synthetase II. ► Disruption of the cpsII gene significantly reduced the growth of epimastigotes. ► In particular, the CPSII-null mutant severely retarded intracellular growth. ► The de novo pyrimidine pathway is critical for the parasite growth in the host cell. -- Abstract: The intracellular parasitic protist Trypanosoma cruzi is the causative agent of Chagas disease in Latin America. In general, pyrimidine nucleotides are supplied by both de novo biosynthesis and salvage pathways. While epimastigotes—an insect form—possess both activities, amastigotes—an intracellular replicating form of T. cruzi—are unable to mediate the uptake of pyrimidine. However, the requirement of de novo pyrimidine biosynthesis for parasite growth and survival has not yet been elucidated. Carbamoyl-phosphate synthetase II (CPSII) is the first and rate-limiting enzyme of the de novo biosynthetic pathway, and increased CPSII activity is associated with the rapid proliferation of tumor cells. In the present study, we showed that disruption of the T. cruzicpsII gene significantly reduced parasite growth. In particular, the growth of amastigotes lacking the cpsII gene was severely suppressed. Thus, the de novo pyrimidine pathway is important for proliferation of T. cruzi in the host cell cytoplasm and represents a promising target for chemotherapy against Chagas disease.

  7. Mutation of a Conserved Nuclear Export Sequence in Chikungunya Virus Capsid Protein Disrupts Host Cell Nuclear Import.

    Science.gov (United States)

    Jacobs, Susan C; Taylor, Adam; Herrero, Lara J; Mahalingam, Suresh; Fazakerley, John K

    2017-10-20

    Transmitted by mosquitoes; chikungunya virus (CHIKV) is responsible for frequent outbreaks of arthritic disease in humans. CHIKV is an arthritogenic alphavirus of the Togaviridae family. Capsid protein, a structural protein encoded by the CHIKV RNA genome, is able to translocate to the host cell nucleus. In encephalitic alphaviruses nuclear translocation induces host cell shut off; however, the role of capsid protein nuclear localisation in arthritogenic alphaviruses remains unclear. Using replicon systems, we investigated a nuclear export sequence (NES) in the N-terminal region of capsid protein; analogous to that found in encephalitic alphavirus capsid but uncharacterised in CHIKV. The chromosomal maintenance 1 (CRM1) export adaptor protein mediated CHIKV capsid protein export from the nucleus and a region within the N-terminal part of CHIKV capsid protein was required for active nuclear targeting. In contrast to encephalitic alphaviruses, CHIKV capsid protein did not inhibit host nuclear import; however, mutating the NES of capsid protein (∆NES) blocked host protein access to the nucleus. Interactions between capsid protein and the nucleus warrant further investigation.

  8. Frequent occurrence of nonprimate hepacivirus infections in Thoroughbred breeding horses - A cross-sectional study for the occurrence of infections and potential risk factors.

    Science.gov (United States)

    Reichert, Claudia; Campe, Amely; Walter, Stephanie; Pfaender, Stephanie; Welsch, Kathrin; Ruddat, Inga; Sieme, Harald; Feige, Karsten; Steinmann, Eike; Cavalleri, Jessika M V

    2017-05-01

    Recently, several new hepaciviruses have been identified of which the nonprimate hepacivirus (NPHV) - the closest relative to hepatitis C virus (HCV) discovered to date - is highly prevalent in horses. However, potential risk factors for the transmission of NPHV among horses remain still unknown. Therefore, the objective of this study was to investigate the occurrence of NPHV infections in Thoroughbreds in northern and western Germany and to identify potential risk factors associated with NPHV infections. Using a cross-sectional study design, a total of 733 serum samples from Thoroughbred broodmares and stallions from northern and western Germany were analyzed for the presence of anti-NPHV nonstructural protein 3 (NS3) antibodies and NPHV RNA using the luciferase immunoprecipitation system (LIPS) and a quantitative real-time PCR, respectively. Information regarding signalment, stud farm, breeding history and international transportation history of each horse were collected and evaluated. A frequent occurrence of NPHV was found in the study population with 453 seropositive horses (61.8%) and 134 horses (18.3%) carrying NPHV RNA. Furthermore, statistical analysis revealed that the probability of being infected decreased for horses with a transportation history with increasing age by 20% each year. For horses that stayed in Germany no association between age and infection could be observed. In conclusion, the high occurrence of NPHV infections in breeding Thoroughbreds suggests circulating NPHV infections, endemic herds or persistent shedding in these animals and revealed the association of age and international transportation as risk factor for NPHV infections. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Infection,

    Science.gov (United States)

    1980-10-16

    characteristic in severe gram-negative sepsis. Hypertriglyceridemia results from an increase in hepatic synthesis in combination with diminished activity of...induced stress, and tissue repair (1). The magnitude and type of nutritional losses caused by an infection reflect both the severity and duration of an... several functional forms of nutrient loss must be anticipated. Functional losses are defined as the within-body losses of nutrients due to infection

  10. Staphylococcus aureus-induced G2/M phase transition delay in host epithelial cells increases bacterial infective efficiency.

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    Ludmila Alekseeva

    Full Text Available Staphylococcus aureus is a highly versatile, opportunistic pathogen and the etiological agent of a wide range of infections in humans and warm-blooded animals. The epithelial surface is its principal site of colonization and infection. In this work, we investigated the cytopathic effect of S. aureus strains from human and animal origins and their ability to affect the host cell cycle in human HeLa and bovine MAC-T epithelial cell lines. S. aureus invasion slowed down cell proliferation and induced a cytopathic effect, resulting in the enlargement of host cells. A dramatic decrease in the number of mitotic cells was observed in the infected cultures. Flow cytometry analysis revealed an S. aureus-induced delay in the G2/M phase transition in synchronous HeLa cells. This delay required the presence of live S. aureus since the addition of the heat-killed bacteria did not alter the cell cycle. The results of Western blot experiments showed that the G2/M transition delay was associated with the accumulation of inactive cyclin-dependent kinase Cdk1, a key inducer of mitosis entry, and with the accumulation of unphosphorylated histone H3, which was correlated with a reduction of the mitotic cell number. Analysis of S. aureus proliferation in asynchronous, G1- and G2-phase-enriched HeLa cells showed that the G2 phase was preferential for bacterial infective efficiency, suggesting that the G2 phase delay may be used by S. aureus for propagation within the host. Taken together, our results divulge the potential of S. aureus in the subversion of key cellular processes such as cell cycle progression, and shed light on the biological significance of S. aureus-induced host cell cycle alteration.

  11. Postgenomic approach to identify novel Mycobacterium leprae antigens with potential to improve immunodiagnosis of infection

    NARCIS (Netherlands)

    Geluk, Annemieke; Klein, Michèl R.; Franken, Kees L. M. C.; van Meijgaarden, Krista E.; Wieles, Brigitte; Pereira, Kelly Cristina; Bührer-Sékula, Samira; Klatser, Paul R.; Brennan, Patrick J.; Spencer, John S.; Williams, Diana L.; Pessolani, Maria C. V.; Sampaio, Elizabeth P.; Ottenhoff, Tom H. M.

    2005-01-01

    Early detection of Mycobacterium leprae infection is considered an important component of strategies aiming at reducing transmission of infection, but currently available diagnostic tools often lack sufficient sensitivity and specificity to reach this goal. Recent comparative genomics have revealed

  12. Investigating Potential Effects of Dengue Virus Infection and Pre-exposure to DEET on Aedes aegypti Behaviors

    Science.gov (United States)

    2016-02-05

    Investigating Potential Effects of Dengue Virus Infection and Pre-exposure to DEET on Aedes aegypti Behaviors by Victor A...exposure to DEET on Aedes aegypti Behaviors" Name of Candidate: Victor Sugiharto Doctor of Philosophy Degree February 5, 2016 DISSERTATION AND...Infection and Pre-exposure to DEET on Aedes aegypti Behaviors" is appropriately acknowledged and. beyond brief excerpts , is with the permission of

  13. Proteome profile of swine testicular cells infected with porcine transmissible gastroenteritis coronavirus.

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    Ruili Ma

    Full Text Available The interactions occurring between a virus and a host cell during a viral infection are complex. The purpose of this paper was to analyze altered cellular protein levels in porcine transmissible gastroenteritis coronavirus (TGEV-infected swine testicular (ST cells in order to determine potential virus-host interactions. A proteomic approach using isobaric tags for relative and absolute quantitation (iTRAQ-coupled two-dimensional liquid chromatography-tandem mass spectrometry identification was conducted on the TGEV-infected ST cells. The results showed that the 4-plex iTRAQ-based quantitative approach identified 4,112 proteins, 146 of which showed significant changes in expression 48 h after infection. At 64 h post infection, 219 of these proteins showed significant change, further indicating that a larger number of proteomic changes appear to occur during the later stages of infection. Gene ontology analysis of the altered proteins showed enrichment in multiple biological processes, including cell adhesion, response to stress, generation of precursor metabolites and energy, cell motility, protein complex assembly, growth, developmental maturation, immune system process, extracellular matrix organization, locomotion, cell-cell signaling, neurological system process, and cell junction organization. Changes in the expression levels of transforming growth factor beta 1 (TGF-β1, caspase-8, and heat shock protein 90 alpha (HSP90α were also verified by western blot analysis. To our knowledge, this study is the first time the response profile of ST host cells following TGEV infection has been analyzed using iTRAQ technology, and our description of the late proteomic changes that are occurring after the time of vigorous viral production are novel. Therefore, this study provides a solid foundation for further investigation, and will likely help us to better understand the mechanisms of TGEV infection and pathogenesis.

  14. Biological Activities of Uric Acid in Infection Due to Enteropathogenic and Shiga-Toxigenic Escherichia coli

    Science.gov (United States)

    Broome, Jacqueline E.; Lis, Agnieszka

    2016-01-01

    In previous work, we identified xanthine oxidase (XO) as an important enzyme in the interaction between the host and enteropathogenic Escherichia coli (EPEC) and Shiga-toxigenic E. coli (STEC). Many of the biological effects of XO were due to the hydrogen peroxide produced by the enzyme. We wondered, however, if uric acid generated by XO also had biological effects in the gastrointestinal tract. Uric acid triggered inflammatory responses in the gut, including increased submucosal edema and release of extracellular DNA from host cells. While uric acid alone was unable to trigger a chloride secretory response in intestinal monolayers, it did potentiate the secretory response to cyclic AMP agonists. Uric acid crystals were formed in vivo in the lumen of the gut in response to EPEC and STEC infections. While trying to visualize uric acid crystals formed during EPEC and STEC infections, we noticed that uric acid crystals became enmeshed in the neutrophilic extracellular traps (NETs) produced from host cells in response to bacteria in cultured cell systems and in the intestine in vivo. Uric acid levels in the gut lumen increased in response to exogenous DNA, and these increases were enhanced by the actions of DNase I. Interestingly, addition of DNase I reduced the numbers of EPEC bacteria recovered after a 20-h infection and protected against EPEC-induced histologic damage. PMID:26787720

  15. Does Preoperative Antimicrobial Prophylaxis Influence the Diagnostic Potential of Periprosthetic Tissues in Hip or Knee Infections?

    Science.gov (United States)

    Bedenčič, Klemen; Kavčič, Martina; Faganeli, Nataša; Mihalič, Rene; Mavčič, Blaž; Dolenc, Jožica; Bajc, Zlatka; Trebše, Rihard

    2016-01-01

    Undiagnosed low-grade prosthetic joint infections (PJI) are recognized as an important reason for early failure of presumably aseptic revisions. Preoperatively administered antimicrobial prophylaxis reduces the incidence of PJI but it may reduce the sensitivity of microbiologic periprosthetic tissue cultures and consequently increase the incidence of undiagnosed septic prosthetic joint failures, which can lead to catastrophic serial revisions. We wished to determine whether administration of preoperative antibiotics decreases the likelihood of diagnosing PJI in patients undergoing revision hip or knee arthroplasty in whom infection is suspected. We prospectively enrolled and evaluated 40 patients (29 with THAs and 11 with TKAs) who met the following inclusion criteria: older than 18 years, with suspected PJI of unknown cause, undergoing surgical revision. After arthrotomy, three tissue samples were obtained for microbiologic analysis and diagnosis, and antimicrobial prophylaxis (cefazolin 2 g intravenously) then was administered. Later during the procedure, but before débridement and irrigation, the second set of three tissue samples was obtained from the same surgical area and was cultured. Tissue concentration of prophylactic antibiotic was verified with the second set of samples. A positive culture result was defined as one or more positive cultures (growth on agar at or before 14 days). We then compared the yield on the microbiologic cultures obtained before administration of antibiotics with the yield on the cultures obtained after antibiotics were administered. An a priori analysis was performed; with the numbers available, we had 98% power to detect a difference in diagnostic sensitivity of 33%. With the numbers available, we found no difference in the likelihood that an infection would be diagnosed between the samples obtained before and after administration of antimicrobial prophylaxis (odds ratio [OR] for positive microbial culture = 0.99; 95% CI, 0

  16. Eicosanoids and Respiratory Viral Infection: Coordinators of Inflammation and Potential Therapeutic Targets

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    Mary K. McCarthy

    2012-01-01

    Full Text Available Viruses are frequent causes of respiratory infection, and viral respiratory infections are significant causes of hospitalization, morbidity, and sometimes mortality in a variety of patient populations. Lung inflammation induced by infection with common respiratory pathogens such as influenza and respiratory syncytial virus is accompanied by increased lung production of prostaglandins and leukotrienes, lipid mediators with a wide range of effects on host immune function. Deficiency or pharmacologic inhibition of prostaglandin and leukotriene production often results in a dampened inflammatory response to acute infection with a respiratory virus. These mediators may, therefore, serve as appealing therapeutic targets for disease caused by respiratory viral infection.

  17. Inhibition of influenza virus replication by targeting broad host cell pathways.

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    Isabelle Marois

    Full Text Available Antivirals that are currently used to treat influenza virus infections target components of the virus which can mutate rapidly. Consequently, there has been an increase in the number of resistant strains to one or many antivirals in recent years. Here we compared the antiviral effects of lysosomotropic alkalinizing agents (LAAs and calcium modulators (CMs, which interfere with crucial events in the influenza virus replication cycle, against avian, swine, and human viruses of different subtypes in MDCK cells. We observed that treatment with LAAs, CMs, or a combination of both, significantly inhibited viral replication. Moreover, the drugs were effective even when they were administered 8 h after infection. Finally, analysis of the expression of viral acidic polymerase (PA revealed that both drugs classes interfered with early events in the viral replication cycle. This study demonstrates that targeting broad host cellular pathways can be an efficient strategy to inhibit influenza replication. Furthermore, it provides an interesting avenue for drug development where resistance by the virus might be reduced since the virus is not targeted directly.

  18. A Lipopeptide Facilitate Induction of Mycobacterium leprae Killing in Host Cells

    Science.gov (United States)

    Maeda, Yumi; Tamura, Toshiki; Fukutomi, Yasuo; Mukai, Tetsu; Kai, Masanori; Makino, Masahiko

    2011-01-01

    Little is known of the direct microbicidal activity of T cells in leprosy, so a lipopeptide consisting of the N-terminal 13 amino acids lipopeptide (LipoK) of a 33-kD lipoprotein of Mycobacterium leprae, was synthesized. LipoK activated M. leprae infected human dendritic cells (DCs) to induce the production of IL-12. These activated DCs stimulated autologous CD4+ or CD8+ T cells towards type 1 immune response by inducing interferon-gamma secretion. T cell proliferation was also evident from the CFSE labeling of target CD4+ or CD8+ T cells. The direct microbicidal activity of T cells in the control of M. leprae multiplication is not well understood. The present study showed significant production of granulysin, granzyme B and perforin from these activated CD4+ and CD8+ T cells when stimulated with LipoK activated, M. leprae infected DCs. Assessment of the viability of M. leprae in DCs indicated LipoK mediated T cell-dependent killing of M. leprae. Remarkably, granulysin as well as granzyme B could directly kill M. leprae in vitro. Our results provide evidence that LipoK could facilitate M. leprae killing through the production of effector molecules granulysin and granzyme B in T cells. PMID:22132248

  19. Flavivirus infection from mosquitoes in vitro reveals cell entry at the plasma membrane

    International Nuclear Information System (INIS)

    Vancini, Ricardo; Kramer, Laura D.; Ribeiro, Mariana; Hernandez, Raquel; Brown, Dennis

    2013-01-01

    Dengue and West Nile viruses are enveloped RNA viruses that belong to genus Flavivirus (family Flaviviridae) and are considered important mosquito-borne viral pathogenic agents worldwide. A potential target for intervention strategies is the virus cell entry mechanism. Previous studies of flavivirus entry have focused on the effects of biochemical and molecular inhibitors on viral entry leading to controversial conclusions suggesting that the process is dependent upon endocytosis and low pH mediated membrane fusion. In this study we analyzed the early events in the infection process by means of electron microscopy and immuno-gold labeling of viral particles during cell entry, and used as a new approach for infecting cells with viruses obtained directly from mosquitoes. The results show that Dengue and West Nile viruses may infect cells by a mechanism that involves direct penetration of the host cell plasma membrane as proposed for alphaviruses.

  20. Chlamydia infection across host species boundaries promotes distinct sets of transcribed anti-apoptotic factors.

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    Joshua eMessinger

    2015-12-01

    Full Text Available Chlamydiae, obligate intracellular bacteria, cause significant human and veterinary associated diseases. Having emerged an estimated 700-million years ago, these bacteria have twice adapted to humans as a host species, causing sexually transmitted infection (C. trachomatis and respiratory associated disease (C. pneumoniae. The principle mechanism of host cell defense against these intracellular bacteria is the induction of cell death via apoptosis. However, in the arms race of co-evolution, Chlamydiae have developed mechanisms to promote cell viability and inhibit cell death. Herein we examine the impact of Chlamydiae infection across multiple host species on transcription of anti-apoptotic genes. We found mostly distinct patterns of gene expression (Mcl1 and cIAPs elicited by each pathogen-host pair indicating Chlamydiae infection across host species boundaries does not induce a universally shared host response. Understanding species specific host-pathogen interactions is paramount to deciphering how potential pathogens become emerging diseases.

  1. Flavivirus infection from mosquitoes in vitro reveals cell entry at the plasma membrane

    Energy Technology Data Exchange (ETDEWEB)

    Vancini, Ricardo [Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC (United States); Kramer, Laura D. [Wadsworth Center, New York State Department of Health, and School of Public Health, State University of New York at Albany, Albany, NY (United States); Ribeiro, Mariana; Hernandez, Raquel [Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC (United States); Brown, Dennis, E-mail: dennis_brown@ncsu.edu [Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC (United States)

    2013-01-20

    Dengue and West Nile viruses are enveloped RNA viruses that belong to genus Flavivirus (family Flaviviridae) and are considered important mosquito-borne viral pathogenic agents worldwide. A potential target for intervention strategies is the virus cell entry mechanism. Previous studies of flavivirus entry have focused on the effects of biochemical and molecular inhibitors on viral entry leading to controversial conclusions suggesting that the process is dependent upon endocytosis and low pH mediated membrane fusion. In this study we analyzed the early events in the infection process by means of electron microscopy and immuno-gold labeling of viral particles during cell entry, and used as a new approach for infecting cells with viruses obtained directly from mosquitoes. The results show that Dengue and West Nile viruses may infect cells by a mechanism that involves direct penetration of the host cell plasma membrane as proposed for alphaviruses.

  2. [Distribution diversity of integrins and calcium channels on major human and mouse host cells of Leptospira species].

    Science.gov (United States)

    Li, Cheng-xue; Zhao, Xin; Qian, Jing; Yan, Jie

    2012-07-01

    To determine the distribution of integrins and calcium channels on major human and mouse host cells of Leptospira species. The expression of β1, β2 and β3 integrins was detected with immunofluorescence assay on the surface of human monocyte line THP-1, mouse mononuclear-macrophage-like cell line J774A.1, human vascular endothelial cell line HUVEC, mouse vascular endothelial cell EOMA, human hepatocyte line L-02, mouse hepatocyte line Hepa1-6, human renal tubular epithelial cell line HEK-293, mouse glomerular membrane epithelial cell line SV40-MES13, mouse collagen blast line NIH/3T3, human and mouse platelets. The distribution of voltage gate control calcium channels Cav3.1, Cav3.2, Cav3.3 and Cav2.3, and receptor gate calcium channels P(2)X(1), P(2)2X(2), P(2)X(3), P(2)X(4), P(2)X(5), P(2)X(6) and P(2)X(7) were determined with Western blot assay. β1 integrin proteins were positively expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, L-02, Hepa1-6 and HEK-239 cells as well as human and mouse platelets. β2 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, and NIH/3T3 cells. β3 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, Hepa1-6, HEK-239 and NIH/3T3 cells as well as human and mouse platelets. P(2)X(1) receptor gate calcium channel was expressed on the membrane surface of human and mouse platelets, while P(2)X(5) receptor gate calcium channel was expressed on the membrane surface of J774A.1, THP-1, L-02, Hepa1-6, HEK-239 and HUVEC cells. However, the other calcium channels were not detected on the tested cell lines or platelets. There is a large distribution diversity of integrins and calcium channel proteins on the major human and mouse host cells of Leptospira species, which may be associated with the differences of leptospira-induced injury in different host cells.

  3. Campylobacter jejuni type VI secretion system: roles in adaptation to deoxycholic acid, host cell adherence, invasion, and in vivo colonization.

    Science.gov (United States)

    Lertpiriyapong, Kvin; Gamazon, Eric R; Feng, Yan; Park, Danny S; Pang, Jassia; Botka, Georgina; Graffam, Michelle E; Ge, Zhongming; Fox, James G

    2012-01-01

    The recently identified type VI secretion system (T6SS) of proteobacteria has been shown to promote pathogenicity, competitive advantage over competing microorganisms, and adaptation to environmental perturbation. By detailed phenotypic characterization of loss-of-function mutants, in silico, in vitro and in vivo analyses, we provide evidence that the enteric pathogen, Campylobacter jejuni, possesses a functional T6SS and that the secretion system exerts pleiotropic effects on two crucial processes--survival in a bile salt, deoxycholic acid (DCA), and host cell adherence and invasion. The expression of T6SS during initial exposure to the upper range of physiological levels of DCA (0.075%-0.2%) was detrimental to C. jejuni proliferation, whereas down-regulation or inactivation of T6SS enabled C. jejuni to resist this effect. The C. jejuni multidrug efflux transporter gene, cmeA, was significantly up-regulated during the initial exposure to DCA in the wild type C. jejuni relative to the T6SS-deficient strains, suggesting that inhibition of proliferation is the consequence of T6SS-mediated DCA influx. A sequential modulation of the efflux transporter activity and the T6SS represents, in part, an adaptive mechanism for C. jejuni to overcome this inhibitory effect, thereby ensuring its survival. C. jejuni T6SS plays important roles in host cell adhesion and invasion as T6SS inactivation resulted in a reduction of adherence to and invasion of in vitro cell lines, while over-expression of a hemolysin co-regulated protein, which encodes a secreted T6SS component, greatly enhanced these processes. When inoculated into B6.129P2-IL-10(tm1Cgn) mice, the T6SS-deficient C. jejuni strains did not effectively establish persistent colonization, indicating that T6SS contributes to colonization in vivo. Taken together, our data demonstrate the importance of bacterial T6SS in host cell adhesion, invasion, colonization and, for the first time to our knowledge, adaptation to DCA

  4. Campylobacter jejuni type VI secretion system: roles in adaptation to deoxycholic acid, host cell adherence, invasion, and in vivo colonization.

    Directory of Open Access Journals (Sweden)

    Kvin Lertpiriyapong

    Full Text Available The recently identified type VI secretion system (T6SS of proteobacteria has been shown to promote pathogenicity, competitive advantage over competing microorganisms, and adaptation to environmental perturbation. By detailed phenotypic characterization of loss-of-function mutants, in silico, in vitro and in vivo analyses, we provide evidence that the enteric pathogen, Campylobacter jejuni, possesses a functional T6SS and that the secretion system exerts pleiotropic effects on two crucial processes--survival in a bile salt, deoxycholic acid (DCA, and host cell adherence and invasion. The expression of T6SS during initial exposure to the upper range of physiological levels of DCA (0.075%-0.2% was detrimental to C. jejuni proliferation, whereas down-regulation or inactivation of T6SS enabled C. jejuni to resist this effect. The C. jejuni multidrug efflux transporter gene, cmeA, was significantly up-regulated during the initial exposure to DCA in the wild type C. jejuni relative to the T6SS-deficient strains, suggesting that inhibition of proliferation is the consequence of T6SS-mediated DCA influx. A sequential modulation of the efflux transporter activity and the T6SS represents, in part, an adaptive mechanism for C. jejuni to overcome this inhibitory effect, thereby ensuring its survival. C. jejuni T6SS plays important roles in host cell adhesion and invasion as T6SS inactivation resulted in a reduction of adherence to and invasion of in vitro cell lines, while over-expression of a hemolysin co-regulated protein, which encodes a secreted T6SS component, greatly enhanced these processes. When inoculated into B6.129P2-IL-10(tm1Cgn mice, the T6SS-deficient C. jejuni strains did not effectively establish persistent colonization, indicating that T6SS contributes to colonization in vivo. Taken together, our data demonstrate the importance of bacterial T6SS in host cell adhesion, invasion, colonization and, for the first time to our knowledge

  5. Host cell entry of powdery mildew is correlated with endosomal transport of antagonistically acting VvPEN1 and VvMLO to the papilla.

    Science.gov (United States)

    Feechan, A; Jermakow, A M; Ivancevic, A; Godfrey, D; Pak, H; Panstruga, R; Dry, I B

    2013-10-01

    Challenge by a nonadapted powdery mildew fungal pathogen leads to the formation of a local cell-wall apposition (papilla) beneath the point of attempted penetration. Several plasma membrane (PM) proteins with opposing roles in powdery mildew infection, including Arabidopsis thaliana PENETRATION1 (PEN1) and barley (Hordeum vulgare) MILDEW RESISTANCE LOCUS O (MLO), are localized to the site of powdery mildew attack. PEN1 contributes to penetration resistance to nonadapted powdery mildews, whereas MLO is a susceptibility factor required by adapted powdery mildew pathogens for host cell entry. Our previous studies have demonstrated that the vesicle and endosomal trafficking inhibitors, brefeldin A and wortmannin, have opposite effects on the penetration rates of adapted and nonadapted powdery mildews on grapevine. These findings prompted us to study the pathogen-induced intracellular trafficking of grapevine variants of MLO and PEN1. We first identified grapevine (Vitis vinifera) VvPEN1 and VvMLO orthologs that rescue Arabidopsis Atpen1 and Atmlo2 mlo6 mlo12 null mutants, respectively. By using endomembrane trafficking inhibitors in combination with fluorescence microscopy, we demonstrate that VvMLO3/VvMLO4 and VvPEN1 are co-trafficked together from the PM to the site of powdery mildew challenge. This focal accumulation of VvMLO3/VvMLO4 and VvPEN1 to the site of attack seems to be required for their opposing functions during powdery mildew attack, because their subcellular localization is correlated with the outcome of attempted powdery mildew penetration.

  6. The Xanthomonas campestris type III effector XopJ targets the host cell proteasome to suppress salicylic-acid mediated plant defence.

    Directory of Open Access Journals (Sweden)

    Suayib Üstün

    Full Text Available The phytopathogenic bacterium Xanthomonas campestris pv. vesicatoria (Xcv requires type III effector proteins (T3Es for virulence. After translocation into the host cell, T3Es are thought to interact with components of host immunity to suppress defence responses. XopJ is a T3E protein from Xcv that interferes with plant immune responses; however, its host cellular target is unknown. Here we show that XopJ interacts with the proteasomal subunit RPT6 in yeast and in planta to inhibit proteasome activity. A C235A mutation within the catalytic triad of XopJ as well as a G2A exchange within the N-terminal myristoylation motif abolishes the ability of XopJ to inhibit the proteasome. Xcv ΔxopJ mutants are impaired in growth and display accelerated symptom development including tissue necrosis on susceptible pepper leaves. Application of the proteasome inhibitor MG132 restored the ability of the Xcv ΔxopJ to attenuate the development of leaf necrosis. The XopJ dependent delay of tissue degeneration correlates with reduced levels of salicylic acid (SA and changes in defence- and senescence-associated gene expression. Necrosis upon infection with Xcv ΔxopJ was greatly reduced in pepper plants with reduced expression of NPR1, a central regulator of SA responses, demonstrating the involvement of SA-signalling in the development of XopJ dependent phenotypes. Our results suggest that XopJ-mediated inhibition of the proteasome interferes with SA-dependent defence response to attenuate onset of necrosis and to alter host transcription. A central role of the proteasome in plant defence is discussed.

  7. Host cell killing by the West Nile Virus NS2B-NS3 proteolytic complex: NS3 alone is sufficient to recruit caspase-8-based apoptotic pathway

    International Nuclear Information System (INIS)

    Ramanathan, Mathura P.; Chambers, Jerome A.; Pankhong, Panyupa; Chattergoon, Michael; Attatippaholkun, Watcharee; Dang, Kesen; Shah, Neelima; Weiner, David B.

    2006-01-01

    The West Nile Virus (WNV) non-structural proteins 2B and 3 (NS2B-NS3) constitute the proteolytic complex that mediates the cleavage and processing of the viral polyprotein. NS3 recruits NS2B and NS5 proteins to direct protease and replication activities. In an effort to investigate the biology of the viral protease, we cloned cDNA encoding the NS2B-NS3 proteolytic complex from brain tissue of a WNV-infected dead crow, collected from the Lower Merion area (Merion strain). Expression of the NS2B-NS3 gene cassette induced apoptosis within 48 h of transfection. Electron microscopic analysis of NS2B-NS3-transfected cells revealed ultra-structural changes that are typical of apoptotic cells including membrane blebbing, nuclear disintegration and cytoplasmic vacuolations. The role of NS3 or NS2B in contributing to host cell apoptosis was examined. NS3 alone triggers the apoptotic pathways involving caspases-8 and -3. Experimental results from the use of caspase-specific inhibitors and caspase-8 siRNA demonstrated that the activation of caspase-8 was essential to initiate apoptotic signaling in NS3-expressing cells. Downstream of caspase-3 activation, we observed nuclear membrane ruptures and cleavage of the DNA-repair enzyme, PARP in NS3-expressing cells. Nuclear herniations due to NS3 expression were absent in the cells treated with a caspase-3 inhibitor. Expression of protease and helicase domains themselves was sufficient to trigger apoptosis generating insight into the apoptotic pathways triggered by NS3 from WNV

  8. Establishment of human papillomavirus infection requires cell cycle progression.

    Directory of Open Access Journals (Sweden)

    Dohun Pyeon

    2009-02-01

    Full Text Available Human papillomaviruses (HPVs are DNA viruses associated with major human cancers. As such there is a strong interest in developing new means, such as vaccines and microbicides, to prevent HPV infections. Developing the latter requires a better understanding of the infectious life cycle of HPVs. The HPV infectious life cycle is closely linked to the differentiation state of the stratified epithelium it infects, with progeny virus only made in the terminally differentiating suprabasal compartment. It has long been recognized that HPV must first establish its infection within the basal layer of stratified epithelium, but why this is the case has not been understood. In part this restriction might reflect specificity of expression of entry receptors. However, this hypothesis could not fully explain the differentiation restriction of HPV infection, since many cell types can be infected with HPVs in monolayer cell culture. Here, we used chemical biology approaches to reveal that cell cycle progression through mitosis is critical for HPV infection. Using infectious HPV16 particles containing the intact viral genome, G1-synchronized human keratinocytes as hosts, and early viral gene expression as a readout for infection, we learned that the recipient cell must enter M phase (mitosis for HPV infection to take place. Late M phase inhibitors had no effect on infection, whereas G1, S, G2, and early M phase cell cycle inhibitors efficiently prevented infection. We conclude that host cells need to pass through early prophase for successful onset of transcription of the HPV encapsidated genes. These findings provide one reason why HPVs initially establish infections in the basal compartment of stratified epithelia. Only this compartment of the epithelium contains cells progressing through the cell cycle, and therefore it is only in these cells that HPVs can establish their infection. By defining a major condition for cell susceptibility to HPV infection, these

  9. Potential health impacts of heavy metals on HIV-infected population in USA.

    Directory of Open Access Journals (Sweden)

    Xiaohui Xu

    Full Text Available PURPOSE: Noninfectious comorbidities such as cardiovascular diseases have become increasingly prevalent and occur earlier in life in persons with HIV infection. Despite the emerging body of literature linking environmental exposures to chronic disease outcomes in the general population, the impacts of environmental exposures have received little attention in HIV-infected population. The aim of this study is to investigate whether individuals living with HIV have elevated prevalence of heavy metals compared to non-HIV infected individuals in United States. METHODS: We used the National Health and Nutrition Examination Survey (NHANES 2003-2010 to compare exposures to heavy metals including cadmium, lead, and total mercury in HIV infected and non-HIV infected subjects. RESULTS: In this cross-sectional study, we found that HIV-infected individuals had higher concentrations of all heavy metals than the non-HIV infected group. In a multivariate linear regression model, HIV status was significantly associated with increased blood cadmium (p=0.03 after adjusting for age, sex, race, education, poverty income ratio, and smoking. However, HIV status was not statistically associated with lead or mercury levels after adjusting for the same covariates. CONCLUSIONS: Our findings suggest that HIV-infected patients might be significantly more exposed to cadmium compared to non-HIV infected individuals which could contribute to higher prevalence of chronic diseases among HIV-infected subjects. Further research is warranted to identify sources of exposure and to understand more about specific health outcomes.

  10. A Lipidomics Approach in the Characterization of Zika-Infected Mosquito Cells: Potential Targets for Breaking the Transmission Cycle.

    Directory of Open Access Journals (Sweden)

    Carlos Fernando Odir Rodrigues Melo

    Full Text Available Recent outbreaks of Zika virus in Oceania and Latin America, accompanied by unexpected clinical complications, made this infection a global public health concern. This virus has tropism to neural tissue, leading to microcephaly in newborns in a significant proportion of infected mothers. The clinical relevance of this infection, the difficulty to perform accurate diagnosis and the small amount of data in literature indicate the necessity of studies on Zika infection in order to characterize new biomarkers of this infection and to establish new targets for viral control in vertebrates and invertebrate vectors. Thus, this study aims at establishing a lipidomics profile of infected mosquito cells compared to a control group to define potential targets for viral control in mosquitoes. Thirteen lipids were elected as specific markers for Zika virus infection (Brazilian strain, which were identified as putatively linked to the intracellular mechanism of viral replication and/or cell recognition. Our findings bring biochemical information that may translate into useful targets for breaking the transmission cycle.

  11. Antimycobacterial drug discovery using Mycobacteria-infected amoebae identifies anti-infectives and new molecular targets.

    Science.gov (United States)

    Trofimov, Valentin; Kicka, Sébastien; Mucaria, Sabrina; Hanna, Nabil; Ramon-Olayo, Fernando; Del Peral, Laura Vela-Gonzalez; Lelièvre, Joël; Ballell, Lluís; Scapozza, Leonardo; Besra, Gurdyal S; Cox, Jonathan A G; Soldati, Thierry

    2018-03-02

    Tuberculosis remains a serious threat to human health world-wide, and improved efficiency of medical treatment requires a better understanding of the pathogenesis and the discovery of new drugs. In the present study, we performed a whole-cell based screen in order to complete the characterization of 168 compounds from the GlaxoSmithKline TB-set. We have established and utilized novel previously unexplored host-model systems to characterize the GSK compounds, i.e. the amoeboid organisms D. discoideum and A. castellanii, as well as a microglial phagocytic cell line, BV2. We infected these host cells with Mycobacterium marinum to monitor and characterize the anti-infective activity of the compounds with quantitative fluorescence measurements and high-content microscopy. In summary, 88.1% of the compounds were confirmed as antibiotics against M. marinum, 11.3% and 4.8% displayed strong anti-infective activity in, respectively, the mammalian and protozoan infection models. Additionally, in the two systems, 13-14% of the compounds displayed pro-infective activity. Our studies underline the relevance of using evolutionarily distant pathogen and host models in order to reveal conserved mechanisms of virulence and defence, respectively, which are potential "universal" targets for intervention. Subsequent mechanism of action studies based on generation of over-expresser M. bovis BCG strains, generation of spontaneous resistant mutants and whole genome sequencing revealed four new molecular targets, including FbpA, MurC, MmpL3 and GlpK.

  12. Host-cell reactivation of ultraviolet-irradiated SV 40 DNA in five complementation groups of xeroderma pigmentosum

    International Nuclear Information System (INIS)

    Abrahams, P.J.; Eb, A.J. van der

    1976-01-01

    Host-cell reactivation of UV-irradiated double-stranded SV40 DNA was studied in BSC-1 monkey cells, normal human cells, heterozygous Xeroderma pigmentosum xp cells, representative cell strains of the five complemention groups of XP and in XP 'variant' cells. The following percentages of survival of the plaque-forming ability of double-stranded SV40 DNA were found in XP cells compared with the value found in normal monkey and human cells: groupA, 13%; group B, 30%; group C, 18%; group D, 14%; group E, 59%; and in the heterozygous XP cells almost 100%. The survival in XP 'variant' cells was 66%. The survival of single-stranded SV40 DNA in BSC-1 cells was much lower than that of double-stranded SV40 DNA in XP cells of complementation group A, which possibly indicates that some repair of UV damage occurs even in XP cells of group A

  13. Nasal Acai Polysaccharides Potentiate Innate Immunity to Protect against Pulmonary Francisella tularensis and Burkholderia pseudomallei Infections

    OpenAIRE

    Skyberg, Jerod A.; Rollins, MaryClare F.; Holderness, Jeff S.; Marlenee, Nicole L.; Schepetkin, Igor A.; Goodyear, Andrew; Dow, Steven W.; Jutila, Mark A.; Pascual, David W.

    2012-01-01

    Pulmonary Francisella tularensis and Burkholderia pseudomallei infections are highly lethal in untreated patients, and current antibiotic regimens are not always effective. Activating the innate immune system provides an alternative means of treating infection and can also complement antibiotic therapies. Several natural agonists were screened for their ability to enhance host resistance to infection, and polysaccharides derived from the Acai berry (Acai PS) were found to have potent abilitie...

  14. Assessing Potential Environmental Contamination by Baylisascaris procyonis Eggs from Infected Raccoons in Southern Texas.

    Science.gov (United States)

    Ogdee, Jacob L; Henke, Scott E; Wester, David B; Fedynich, Alan M

    2017-03-01

    Baylisascaris procyonis is a large ascarid of raccoons (Procyon lotor) and is a zoonotic threat. We documented the potential rate a raccoon population can contaminate their environment with B. procyonis eggs. We estimated the population size of raccoons using a 9 × 7 trapping grid of Havahart traps, identified locations of raccoon scats through systematic searches, and enumerated the distance B. procyonis eggs passively travel from site of origin upon scat decay. During an 8-week capture period, the raccoon population was estimated to be 19.6 ± 1.3 raccoons within the 63-ha study area (1 raccoon/3.2 ha). There were 781 defecation sites, of which 744 (95.3%) were isolated sites and 37 (4.7%) were latrine sites. Fifty-three (6.8%) defecation sites occurred in areas associated with human structures (commensal zone). Of the noncommensal sites, 9 (1.2%) and 719 (98.8%) sites were identified as latrine sites and isolated scats, respectively. More latrine sites were located within the commensal zone (p contaminate 0.03 ± 0.01 ha/year with B. procyonis eggs. Our findings indicate that B. procyonis represents a substantial risk to humans in areas where infected raccoons and humans co-occur.

  15. NLF20: an antimicrobial peptide with therapeutic potential against invasive Pseudomonas aeruginosa infection.

    Science.gov (United States)

    Papareddy, Praveen; Kasetty, Gopinath; Kalle, Martina; Bhongir, Ravi K V; Mörgelin, Matthias; Schmidtchen, Artur; Malmsten, Martin

    2016-01-01

    Increasing resistance to antibiotics makes antimicrobial peptides interesting as novel therapeutics. Here, we report on studies of the peptide NLF20 (NLFRKLTHRLFRRNFGYTLR), corresponding to an epitope of the D helix of heparin cofactor II (HCII), a plasma protein mediating bacterial clearance. Peptide effects were evaluated by a combination of in vitro and in vivo methods, including antibacterial, anti-inflammatory and cytotoxicity assays, fluorescence and electron microscopy, and experimental models of endotoxin shock and Pseudomonas aeruginosa sepsis. The results showed that NLF20 displayed potent antimicrobial effects against the Gram-negative bacteria Escherichia coli and P. aeruginosa, the Gram-positive Bacillus subtilis and Staphylococcus aureus and the fungi Candida albicans and Candida parapsilosis. Importantly, this antimicrobial effect was retained in human blood, particularly for P. aeruginosa. Fluorescence and electron microscopy studies showed that the peptide exerted membrane-breaking effects. In an animal model of P. aeruginosa sepsis, NLF20 reduced bacterial levels, resulting in improved survival. Reduced mortality was also observed in experimental animal models of endotoxin shock, which was paralleled with modulated IFN-γ, IL-10 and coagulation responses. Together, these results indicate that functional epitopes of HCII may have therapeutic potential against bacterial infection. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  16. Molecular characterization of Trypanosoma cruzi SAP proteins with host-cell lysosome exocytosis-inducing activity required for parasite invasion.

    Science.gov (United States)

    Zanforlin, Tamiris; Bayer-Santos, Ethel; Cortez, Cristian; Almeida, Igor C; Yoshida, Nobuko; da Silveira, José Franco

    2013-01-01

    To invade target cells, Trypanosoma cruzi metacyclic forms engage distinct sets of surface and secreted molecules that interact with host components. Serine-, alanine-, and proline-rich proteins (SAP) comprise a multigene family constituted of molecules with a high serine, alanine and proline residue content. SAP proteins have a central domain (SAP-CD) responsible for interaction with and invasion of mammalian cells by metacyclic forms. Using a 513 bp sequence from SAP-CD in blastn analysis, we identified 39 full-length SAP genes in the genome of T. cruzi. Although most of these genes were mapped in the T. cruzi in silico chromosome TcChr41, several SAP sequences were spread out across the genome. The level of SAP transcripts was twice as high in metacyclic forms as in epimastigotes. Monoclonal (MAb-SAP) and polyclonal (anti-SAP) antibodies produced against the recombinant protein SAP-CD were used to investigate the expression and localization of SAP proteins. MAb-SAP reacted with a 55 kDa SAP protein released by epimastigotes and metacyclic forms and with distinct sets of SAP variants expressed in amastigotes and tissue culture-derived trypomastigotes (TCTs). Anti-SAP antibodies reacted with components located in the anterior region of epimastigotes and between the nucleus and the kinetoplast in metacyclic trypomastigotes. In contrast, anti-SAP recognized surface components of amastigotes and TCTs, suggesting that SAP proteins are directed to different cellular compartments. Ten SAP peptides were identified by mass spectrometry in vesicle and soluble-protein fractions obtained from parasite conditioned medium. Using overlapping sequences from SAP-CD, we identified a 54-aa peptide (SAP-CE) that was able to induce host-cell lysosome exocytosis and inhibit parasite internalization by 52%. This study provides novel information about the genomic organization, expression and cellular localization of SAP proteins and proposes a triggering role for extracellular SAP

  17. Yeast casein kinase 2 governs morphology, biofilm formation, cell wall integrity, and host cell damage of Candida albicans.

    Science.gov (United States)

    Jung, Sook-In; Rodriguez, Natalie; Irrizary, Jihyun; Liboro, Karl; Bogarin, Thania; Macias, Marlene; Eivers, Edward; Porter, Edith; Filler, Scott G; Park, Hyunsook

    2017-01-01

    The regulatory networks governing morphogenesis of a pleomorphic fungus, Candida albicans are extremely complex and remain to be completely elucidated. This study investigated the function of C. albicans yeast casein kinase 2 (CaYck2p). The yck2Δ/yck2Δ strain displayed constitutive pseudohyphae in both yeast and hyphal growth conditions, and formed enhanced biofilm under non-biofilm inducing condition. This finding was further supported by gene expression analysis of the yck2Δ/yck2Δ strain which showed significant upregulation of UME6, a key transcriptional regulator of hyphal transition and biofilm formation, and cell wall protein genes ALS3, HWP1, and SUN41, all of which are associated with morphogenesis and biofilm architecture. The yck2Δ/yck2Δ strain was hypersensitive to cell wall damaging agents and had increased compensatory chitin deposition in the cell wall accompanied by an upregulation of the expression of the chitin synthase genes, CHS2, CHS3, and CHS8. Absence of CaYck2p also affected fungal-host interaction; the yck2Δ/yck2Δ strain had significantly reduced ability to damage host cells. However, the yck2Δ/yck2Δ strain had wild-type susceptibility to cyclosporine and FK506, suggesting that CaYck2p functions independently from the Ca+/calcineurin pathway. Thus, in C. albicans, Yck2p is a multifunctional kinase that governs morphogenesis, biofilm formation, cell wall integrity, and host cell interactions.

  18. RNAi screen reveals host cell kinases specifically involved in Listeria monocytogenes spread from cell to cell.

    Directory of Open Access Journals (Sweden)

    Ryan Chong

    Full Text Available Intracellular bacterial pathogens, such as Listeria monocytogenes and Rickettsia conorii display actin-based motility in the cytosol of infected cells and spread from cell to cell through the formation of membrane protrusions at the cell cortex. Whereas the mechanisms supporting cytosolic actin-based motility are fairly well understood, it is unclear whether specific host factors may be required for supporting the formation and resolution of membrane protrusions. To address this gap in knowledge, we have developed high-throughput fluorescence microscopy and computer-assisted image analysis procedures to quantify pathogen spread in human epithelial cells. We used the approach to screen a siRNA library covering the human kinome and identified 7 candidate kinases whose depletion led to severe spreading defects in cells infected with L. monocytogenes. We conducted systematic validation procedures with redundant silencing reagents and confirmed the involvement of the serine/threonine kinases, CSNK1A1 and CSNK2B. We conducted secondary assays showing that, in contrast with the situation observed in CSNK2B-depleted cells, L. monocytogenes formed wild-type cytosolic tails and displayed wild-type actin-based motility in the cytosol of CSNK1A1-depleted cells. Furthermore, we developed a protrusion formation assay and showed that the spreading defect observed in CSNK1A1-depleted cells correlated with the formation of protrusion that did not resolve into double-membrane vacuoles. Moreover, we developed sending and receiving cell-specific RNAi procedures and showed that CSNK1A was required in the sending cells, but was dispensable in the receiving cells, for protrusion resolution. Finally, we showed that the observed defects were specific to Listeria monocytogenes, as Rickettsia conorii displayed wild-type cell-to-cell spread in CSNK1A1- and CSNK2B-depleted cells. We conclude that, in addition to the specific host factors supporting cytosolic actin

  19. A role for host cell exocytosis in InlB-mediated internalisation of Listeria monocytogenes.

    Science.gov (United States)

    Van Ngo, Hoan; Bhalla, Manmeet; Chen, Da-Yuan; Ireton, Keith

    2017-11-01

    The bacterial surface protein InlB mediates internalisation of Listeria monocytogenes into human cells through interaction with the host receptor tyrosine kinase, Met. InlB-mediated entry requires localised polymerisation of the host actin cytoskeleton. Apart from actin polymerisation, roles for other host processes in Listeria entry are unknown. Here, we demonstrate that exocytosis in the human cell promotes InlB-dependent internalisation. Using a probe consisting of VAMP3 with an exofacial green fluorescent protein tag, focal exocytosis was detected during InlB-mediated entry. Exocytosis was dependent on Met tyrosine kinase activity and the GTPase RalA. Depletion of SNARE proteins by small interfering RNA demonstrated an important role for exocytosis in Listeria internalisation. Depletion of SNARE proteins failed to affect actin filaments during internalisation, suggesting that actin polymerisation and exocytosis are separable host responses. SNARE proteins were required for delivery of the human GTPase Dynamin 2, which promotes InlB-mediated entry. Our results identify exocytosis as a novel host process exploited by Listeria for infection. © 2017 John Wiley & Sons Ltd.

  20. Isolating E.Coli Bacteriophage from Raw Sewage and Determining its Selectivity to the Host Cell

    Directory of Open Access Journals (Sweden)

    SM Imeni

    2016-05-01

    Full Text Available Introduction: Bacteriophages are viruses that infect and destroy prokaryote cells, specifically the bacteria. They act too selective, so as each bacteriophage affects only on specific type of bacteria. Due to their specific features, bacteriophages can be used as an appropriate substitute for antibiotics in infectious diseases treatment. Therefore, this study aimed to isolate E. coli-specific bacteriophage from raw sewage. Methods: Eight samples of raw sewage, each containing approximately 50 ml of raw sewage with 10 minute gap, were prepared from Zargandeh wastewater treatment plant, Tehran, Iran. The sewages were mixed with Brain-heart infusion medium (BHI as a liquid culture medium in order to let the microorganisms grow. Incubation, purification and determination of bacteria were followed repeatedly to isolate the bacteriophage. Then it was tested on E.coli (ATCC 25922, Enterococcus faecalis (ATCC 19433, Staphylococcus aureus (ATCC 2392, and Yersinia enterocolitica (ATCC 9610 in order to determine the bacteriophage selectivity. Results: The E.coli bacteriophages were successfully isolated from all the eight samples, that were completely able to lyse and destroy E.coli bacterial cells, though no effect was observed on other types of bacteria. Conclusion: The study findings revealed that bacteriophages act selectively. Considering the raise of antibiotic resistance in the world, bacteriophages can serve as a good substitute for antibiotics in treating infectious diseases.

  1. Bacteriophages-potential for application in wastewater treatment processes

    International Nuclear Information System (INIS)

    Withey, S.; Cartmell, E.; Avery, L.M.; Stephenson, T.

    2005-01-01

    Bacteriophages are viruses that infect and lyse bacteria. Interest in the ability of phages to control bacterial populations has extended from medical applications into the fields of agriculture, aquaculture and the food industry. Here, the potential application of phage techniques in wastewater treatment systems to improve effluent and sludge emissions into the environment is discussed. Phage-mediated bacterial mortality has the potential to influence treatment performance by controlling the abundance of key functional groups. Phage treatments have the potential to control environmental wastewater process problems such as: foaming in activated sludge plants; sludge dewaterability and digestibility; pathogenic bacteria; and to reduce competition between nuisance bacteria and functionally important microbial populations. Successful application of phage therapy to wastewater treatment does though require a fuller understanding of wastewater microbial community dynamics and interactions. Strategies to counter host specificity and host cell resistance must also be developed, as should safety considerations regarding pathogen emergence through transduction

  2. CXCL10/IP10 is a novel potential in vitro marker of TB infection

    Directory of Open Access Journals (Sweden)

    Ilaria Sauzullo

    2009-06-01

    Full Text Available Introduction IFN-γ is a pivotal cytokine in the immune response to Myc. tuberculosis, infact this is the key cytokine produced in response to antigens specific following tuberculosis exposure causing either active or latent tuberculosis (TB and this observation forms the basis of interferon gamma release assay (IGRA, but there are alternative or additional cytokines and chemokines that could be used to improve detection of Myc. tuberculosis infection.The aim of this study was to evaluate the diagnostic utility of chemokine CXCL10/IP-10 as biomarker of active TB and to compare the results with classical QuantiFERON-Gold assay . Methods CXCL10/IP-10 and IFN-γ responses to stimulation with ESAT-6 and CFP-10 were evaluated in 21 patients with active tuberculosis and in 6 healthy unexposed subjects with no history of TB or TB contact were used as controls healthy controls. QuantiFERON-TB Gold (QFT-G, Cellestis was used for the measurement of IFN-γ levels; CXCL10/IP-10 was detected by ELISA (R&D Systems . Results Of the 21 TB patients included, 11 had a QFT-G positive and 10 had negative QFT-G results.All QFT-G positive patients had increased levels of CXCL10/IP-10 (median, pg/ml in both ESAT-6 and CFP-10 stimulated samples patients compared to healthy controls (1807 and 1111 vs 251 and 188 of controls, respectively (p<0.001 for both. The patients with active TB and QFT-G negative exhibited higher concentrations of CXCL10/IP-10 following antigen stimulation (837 pg/ml for ESAT-6;1674 pg/ml for CFP-10 (p<0.001. Conclusion Our study showed that in all patients with active TB, the CXCL10/IP-10 is expressed in higher amounts than IFN-γ following Myc. tuberculosis antigen-specific stimulation, and CXCL10/IP-10 appeared to be even more sensitive than QuantiFERON TB-Gold in TB patients with negative IFN-γ response. The measurement of chemokine CXCL10/IP-10, although not specific for tuberculosis, may have potential as an alternative or additional marker

  3. Atomic evidence that modification of H-bonds established with amino acids critical for host-cell binding induces sterile immunity against malaria

    International Nuclear Information System (INIS)

    Patarroyo, Manuel E.; Cifuentes, Gladys; Pirajan, Camilo; Moreno-Vranich, Armando; Vanegas, Magnolia

    2010-01-01

    Based on the 3D X-ray crystallographic structures of relevant proteins of the malaria parasite involved in invasion to host cells and 3D NMR structures of High Activity Binding Peptides (HABPs) and their respective analogues, it was found that HABPs are rendered into highly immunogenic and sterile immunity inducers in the Aotus experimental model by modifying those amino acids that establish H-bonds with other HABPs or binding to host's cells. This finding adds striking and novel physicochemical principles, at the atomic level, for a logical and rational vaccine development methodology against infectious disease, among them malaria.

  4. Acute respiratory tract infections: a potential trigger for the acute coronary syndrome

    NARCIS (Netherlands)

    Harskamp, Ralf E.; van Ginkel, Margreet W.

    2008-01-01

    Clinical studies suggest that acute respiratory tract infection (ARTI) may be a risk factor for the acute coronary syndrome (ACS). ARTI is associated with an increased risk for ACS up to 2 weeks prior to a cardiac event. The mechanism that may underlie this association is unclear. Infections are

  5. The potential beneficial role of faecal microbiota transplantation in diseases other than Clostridium difficile infection

    NARCIS (Netherlands)

    Singh, R.; Nieuwdorp, M.; ten Berge, I. J. M.; Bemelman, F. J.; Geerlings, S. E.

    2014-01-01

    This review gives an outline of the indications for faecal microbiota transplantation (FMT) for diseases other than Clostridium difficile (C. difficile) infection. The remarkable efficacy of FMT against C. difficile infection has already been demonstrated. The use of FMT for other diseases, such as

  6. The ins and outs of phosphosignalling in Plasmodium: Parasite regulation and host cell manipulation.

    Science.gov (United States)

    Carvalho, Teresa Gil; Morahan, Belinda; John von Freyend, Simona; Boeuf, Philippe; Grau, Georges; Garcia-Bustos, Jose; Doerig, Christian

    2016-07-01

    Signal transduction and kinomics have been rapidly expanding areas of investigation within the malaria research field. Here, we provide an overview of phosphosignalling pathways that operate in all stages of the Plasmodium life cycle. We review signalling pathways in the parasite itself, in the cells it invades, and in other cells of the vertebrate host with which it interacts. We also discuss the potential of these pathways as novel targets for antimalarial intervention. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. DNA repair in human fibroblasts, as reflected by host-cell reactivation of a transfected UV-irradiated luciferase gene, is not related to donor age

    International Nuclear Information System (INIS)

    Merkle, Thomas J.; O'Brien, Katherine; Brooks, Philip J.; Tarone, Robert E.; Robbins, Jay H.

    2004-01-01

    The effect of donor age on the ability of mammalian cells to repair ultraviolet (UV)-induced DNA damage has been studied using several approaches, most recently via assays that measure the host-cell reactivation (HCR) of UV-irradiated reporter gene-containing plasmid vectors following their transfection into cells. Plasmid HCR assays indirectly quantify a cell line's ability to perform nucleotide excision repair (NER) by measuring the enzyme activity of the repaired reporter gene, e.g., chloramphenical acetyltransferase (cat) or luciferase (luc), and are useful in studies investigating whether increasing age may be a risk factor for the deficient repair of potentially cancer-causing, sunlight-induced, DNA lesions in skin cells. In our study, we quantified the DNA repair ability of cultured, nontransformed, human skin fibroblast lines through their HCR of a transfected UV-C-irradiated plasmid containing luc. HCR was measured at various times after transfection in five lines from normal donors of ages 21-96 years, and from one donor who had xeroderma pigmentosum (XP). The normal lines displayed increasing HCR at successive post-transfection time points and showed no significant correlation between HCR and donor age. The XP-A line, known to be markedly deficient in NER of UV-induced DNA damage, showed minimal evidence of HCR compared to the normal lines. To further assess potential variation in HCR with donor age, fibroblast lines from five old donors, ages 84-94 years, were compared with lines from five young donors, ages 17-26 years. While significant differences in HCR were found between some lines, no significant difference was found between the young and old age groups (P=0.44). Our study provides no indication that the higher incidence of skin cancer observed with increasing age is due to an age-related decrease in the ability to repair UV-induced DNA damage

  8. DNA repair in human fibroblasts, as reflected by host-cell reactivation of a transfected UV-irradiated luciferase gene, is not related to donor age

    Energy Technology Data Exchange (ETDEWEB)

    Merkle, Thomas J.; O' Brien, Katherine; Brooks, Philip J.; Tarone, Robert E.; Robbins, Jay H

    2004-10-04

    The effect of donor age on the ability of mammalian cells to repair ultraviolet (UV)-induced DNA damage has been studied using several approaches, most recently via assays that measure the host-cell reactivation (HCR) of UV-irradiated reporter gene-containing plasmid vectors following their transfection into cells. Plasmid HCR assays indirectly quantify a cell line's ability to perform nucleotide excision repair (NER) by measuring the enzyme activity of the repaired reporter gene, e.g., chloramphenical acetyltransferase (cat) or luciferase (luc), and are useful in studies investigating whether increasing age may be a risk factor for the deficient repair of potentially cancer-causing, sunlight-induced, DNA lesions in skin cells. In our study, we quantified the DNA repair ability of cultured, nontransformed, human skin fibroblast lines through their HCR of a transfected UV-C-irradiated plasmid containing luc. HCR was measured at various times after transfection in five lines from normal donors of ages 21-96 years, and from one donor who had xeroderma pigmentosum (XP). The normal lines displayed increasing HCR at successive post-transfection time points and showed no significant correlation between HCR and donor age. The XP-A line, known to be markedly deficient in NER of UV-induced DNA damage, showed minimal evidence of HCR compared to the normal lines. To further assess potential variation in HCR with donor age, fibroblast lines from five old donors, ages 84-94 years, were compared with lines from five young donors, ages 17-26 years. While significant differences in HCR were found between some lines, no significant difference was found between the young and old age groups (P=0.44). Our study provides no indication that the higher incidence of skin cancer observed with increasing age is due to an age-related decrease in the ability to repair UV-induced DNA damage.

  9. Infection, Reproduction Potential, and Root Galling by Root-knot Nematode Species and Concomitant Populations on Peanut and Tobacco

    Science.gov (United States)

    Hirunsalee, Anan; Barker, K. R.; Beute, M. K.

    1995-01-01

    Single populations of Meloidogyne arenaria races 1 (MA1) and 2 (MA2) and M. hapla (MH), and mixed populations of MA1 + MA2 and MA1 + MH with four inoculum levels of eggs were tested on peanut cv. 'Florigiant' and M. incognita-resistant tobacco cv. 'McNair 373' in a greenhouse experiment. Root infection, female development, and reproduction of MA2 on peanut and MA1 on resistant tobacco were limited at 2 and 6 weeks. MA1, MH, and MA1 + MH on peanut had similar root infection (total parasitic forms per root unit) at both 2 and 6 weeks, and similar female development and reproduction potentials at 6 weeks. MA2 tended to depress root infection, female development, and reproduction of MA1 on peanut. MH had little effect on MA1 on this crop. On tobacco, MA2 population had greater incidence of root infection than did MH at 2 weeks. The two nematode species had similar development in roots at 6 weeks. All of these processes were restricted when either MA2 or MH was present together with MA1. As initial inoculum level of parasitically fit populations increased, relative infection ratio on both peanut and tobacco, and reproduction factor on peanut decreased. Populations that had high infection incidence and reproduction rates induced greater root galling than did other populations. Root galling was suppressed in the presence of antagonistic response between nematode populations. PMID:19277277

  10. Existing and potential infection risk zones of yellow fever worldwide: a modelling analysis.

    Science.gov (United States)

    Shearer, Freya M; Longbottom, Joshua; Browne, Annie J; Pigott, David M; Brady, Oliver J; Kraemer, Moritz U G; Marinho, Fatima; Yactayo, Sergio; de Araújo, Valdelaine E M; da Nóbrega, Aglaêr A; Fullman, Nancy; Ray, Sarah E; Mosser, Jonathan F; Stanaway, Jeffrey D; Lim, Stephen S; Reiner, Robert C; Moyes, Catherine L; Hay, Simon I; Golding, Nick

    2018-03-01

    Yellow fever cases are under-reported and the exact distribution of the disease is unknown. An effective vaccine is available but more information is needed about which populations within risk zones should be targeted to implement interventions. Substantial outbreaks of yellow fever in Angola, Democratic Republic of the Congo, and Brazil, coupled with the global expansion of the range of its main urban vector, Aedes aegypti, suggest that yellow fever has the propensity to spread further internationally. The aim of this study was to estimate the disease's contemporary distribution and potential for spread into new areas to help inform optimal control and prevention strategies. We assembled 1155 geographical records of yellow fever virus infection in people from 1970 to 2016. We used a Poisson point process boosted regression tree model that explicitly incorporated environmental and biological explanatory covariates, vaccination coverage, and spatial variability in disease reporting rates to predict the relative risk of apparent yellow fever virus infection at a 5 × 5 km resolution across all risk zones (47 countries across the Americas and Africa). We also used the fitted model to predict the receptivity of areas outside at-risk zones to the introduction or reintroduction of yellow fever transmission. By use of previously published estimates of annual national case numbers, we used the model to map subnational variation in incidence of yellow fever across at-risk countries and to estimate the number of cases averted by vaccination worldwide. Substantial international and subnational spatial variation exists in relative risk and incidence of yellow fever as well as varied success of vaccination in reducing incidence in several high-risk regions, including Brazil, Cameroon, and Togo. Areas with the highest predicted average annual case numbers include large parts of Nigeria, the Democratic Republic of the Congo, and South Sudan, where vaccination coverage in 2016

  11. Existing and potential infection risk zones of yellow fever worldwide: a modelling analysis

    Directory of Open Access Journals (Sweden)

    Freya M Shearer, BSc

    2018-03-01

    Full Text Available Summary: Background: Yellow fever cases are under-reported and the exact distribution of the disease is unknown. An effective vaccine is available but more information is needed about which populations within risk zones should be targeted to implement interventions. Substantial outbreaks of yellow fever in Angola, Democratic Republic of the Congo, and Brazil, coupled with the global expansion of the range of its main urban vector, Aedes aegypti, suggest that yellow fever has the propensity to spread further internationally. The aim of this study was to estimate the disease's contemporary distribution and potential for spread into new areas to help inform optimal control and prevention strategies. Methods: We assembled 1155 geographical records of yellow fever virus infection in people from 1970 to 2016. We used a Poisson point process boosted regression tree model that explicitly incorporated environmental and biological explanatory covariates, vaccination coverage, and spatial variability in disease reporting rates to predict the relative risk of apparent yellow fever virus infection at a 5 × 5 km resolution across all risk zones (47 countries across the Americas and Africa. We also used the fitted model to predict the receptivity of areas outside at-risk zones to the introduction or reintroduction of yellow fever transmission. By use of previously published estimates of annual national case numbers, we used the model to map subnational variation in incidence of yellow fever across at-risk countries and to estimate the number of cases averted by vaccination worldwide. Findings: Substantial international and subnational spatial variation exists in relative risk and incidence of yellow fever as well as varied success of vaccination in reducing incidence in several high-risk regions, including Brazil, Cameroon, and Togo. Areas with the highest predicted average annual case numbers include large parts of Nigeria, the Democratic Republic of the

  12. Antibacterial and Antifungal Potential of Himalayan Medicinal Plants for Treating Wound Infections

    International Nuclear Information System (INIS)

    Habiba, U.; Ahmad, M.; Shinwari, S.; Sultana, S.; Zafar, M.; Shinwari, Z. K.

    2016-01-01

    Many bacterial and fungal strains are involved in wound infectious diseases as most of these strains become resistant to the most commonly used synthetic drugs in Himalayan region. Plant based natural products seem to be an alternative to this problem. The aim of this investigation was to evaluate the In vitro antibacterial and antifungal activities of 30 medicinal plants used in folk recipes by Himalayan people to treat wound infections against multi-drug resistant pathogens. In total of six medically important Myco-bacterial strains Streptococcus pyogenes, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, Candida albicans and Aspergillus niger were tested against methanolic plant extracts at 5 mg/ml concentration using agar disc well diffusion method to determined Minimum inhibitory concentrations (MICs). The plant extracts showed varied levels of MICs against test microorganisms. The strongest antibacterial activity was reported in methanolic extract of Cynadon dactylon (L.) Pers. against Klebsiella pneumoniae with 20.67±1.36 mm MICs, while Candida albicans was considered to be the most resistant pathogen with MICs 9.6±0.57 mm. The findings were compared with results obtained using standard antibiotics, aminooxanilic, ciprofloxacin, cefotaxime, fluconazole and itraconazole at conc. 5mg/ ml. The results provide an evidence of folk medicinal uses of plants among the Himalayan communities to treat wounds. Further research needs to be carried out to identify the active molecules and evaluate the in vivo antibacterial and antifungal activities as well as toxicity level with clinical trials to use full potential of these plants for drug discovery development to control wounds globally. (author)

  13. Estimating potential infection transmission routes in hospital wards using wearable proximity sensors.

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    Philippe Vanhems

    Full Text Available BACKGROUND: Contacts between patients, patients and health care workers (HCWs and among HCWs represent one of the important routes of transmission of hospital-acquired infections (HAI. A detailed description and quantification of contacts in hospitals provides key information for HAIs epidemiology and for the design and validation of control measures. METHODS AND FINDINGS: We used wearable sensors to detect close-range interactions ("contacts" between individuals in the geriatric unit of a university hospital. Contact events were measured with a spatial resolution of about 1.5 meters and a temporal resolution of 20 seconds. The study included 46 HCWs and 29 patients and lasted for 4 days and 4 nights. 14,037 contacts were recorded overall, 94.1% of which during daytime. The number and duration of contacts varied between mornings, afternoons and nights, and contact matrices describing the mixing patterns between HCW and patients were built for each time period. Contact patterns were qualitatively similar from one day to the next. 38% of the contacts occurred between pairs of HCWs and 6 HCWs accounted for 42% of all the contacts including at least one patient, suggesting a population of individuals who could potentially act as super-spreaders. CONCLUSIONS: Wearable sensors represent a novel tool for the measurement of contact patterns in hospitals. The collected data can provide information on important aspects that impact the spreading patterns of infectious diseases, such as the strong heterogeneity of contact numbers and durations across individuals, the variability in the number of contacts during a day, and the fraction of repeated contacts across days. This variability is however associated with a marked statistical stability of contact and mixing patterns across days. Our results highlight the need for such measurement efforts in order to correctly inform mathematical models of HAIs and use them to inform the design and evaluation of

  14. Evaluating the potential of cubosomal nanoparticles for oral delivery of amphotericin B in treating fungal infection

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    Yang Z

    2014-01-01

    Full Text Available Zhiwen Yang,1,3 Meiwan Chen,2 Muhua Yang,1 Jian Chen,1 Weijun Fang,1 Ping Xu11Department of Pharmacy, Songjiang Hospital Affiliated The First People's Hospital, Shanghai Jiao Tong University, Shanghai, 2State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macau, 3Shanghai Songjiang Hospital Affiliated Nanjing Medical University, Nanjing, People's Republic of ChinaAbstract: The oral administration of amphotericin B (AmB has a major drawback of poor bioavailability. The aim of this study was to investigate the potential of glyceryl monoolein (GMO cubosomes as lipid nanocarriers to improve the oral efficacy of AmB. Antifungal efficacy was determined in vivo in rats after oral administration, to investigate its therapeutic use. The human colon adenocarcinoma cell line (Caco-2 was used in vitro to evaluate transport across a model of the intestinal barrier. In vivo antifungal results showed that AmB, loaded in GMO cubosomes, could significantly enhance oral efficacy, compared against Fungizone®, and that during a 2 day course of dosage 10 mg/kg the drug reached effective therapeutic concentrations in renal tissue for treating fungal infections. In the Caco-2 transport studies, GMO cubosomes resulted in a significantly larger amount of AmB being transported into Caco-2 cells, via both clathrin- and caveolae-mediated endocytosis, but not macropinocytosis. These results suggest that GMO cubosomes, as lipid nanovectors, could facilitate the oral delivery of AmB.Keywords: glyceryl monoolein cubosomes, oral delivery, amphotericin B, antifungal activity, absorption mechanism

  15. Comparative proteomics analysis of apoptotic Spodoptera frugiperda cells during p35 knockout Autographa californica multiple nucleopolyhedrovirus infection.

    Science.gov (United States)

    Yu, Qian; Xiong, Youhua; Liu, Jianliang; Wang, Qin; Qiu, Yuanxin; Wen, Dongling

    2016-06-01

    Infection with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) mutants lacking a functional p35 gene can induce host cell apoptosis, which provides the possibility to use the potential of these viruses in the biological control of pest insects. Nonetheless, the proteomics or the protein changes of Spodoptera frugiperda (Sf9) cells infected with p35 knockout AcMNPV have not yet been studied. To further improve the use of AcMNPV, we set out to analyze the protein composition and protein changes of Sf9 cells of different infection stages by isobaric tag for relative and absolute quantification (iTRAQ) techniques. A total of 4004 sf9 proteins were identified by iTRAQ. After comparation of the significantly expressed 483 proteins from p35koAcMNPV-infected Sf9 cells and the significantly expressed 413 proteins from wtAcMNPV-infected Sf9 cells, we found that 226 proteins were specific to p35koAcMNPV-infected Sf9 cells. The 226 proteins were categorized according to GO classification for insects and were categorized into: biological processes, molecular functions and cellular components. Of interest, the most up-regulated proteins related to Epstein-Barr virus infection, RNA transport, Calcium signaling pathway, cGMP-PKG signaling pathway, oxidative phosphorylation and N-Glycan biosynthesis. Determination of the protein changes in p35 knockout AcMNPV-infected Sf9 cells would facilitate the better use of this virus-host cell interaction in pest insect control and other related fields. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Bioactive activities of natural products against herpesvirus infection.

    Science.gov (United States)

    Son, Myoungki; Lee, Minjung; Sung, Gi-Ho; Lee, Taeho; Shin, Yu Su; Cho, Hyosun; Lieberman, Paul M; Kang, Hyojeung

    2013-10-01

    More than 90% of adults have been infected with at least one human herpesvirus, which establish long-term latent infection for the life of the host. While anti-viral drugs exist that limit herpesvirus replication, many of these are ineffective against latent infection. Moreover, drug-resistant strains of herpesvirus emerge following chemotherapeutic treatment. For example, resistance to acyclovir and related nucleoside analogues can occur when mutations arise in either HSV thymidine kinase or DNA polymerases. Thus, there exists an unmet medical need to develop new anti-herpesvirus agents with different mechanisms of action. In this Review, we discuss the promise of anti-herpetic substances derived from natural products including extracts and pure compounds from potential herbal medicines. One example is Glycyrrhizic acid isolated from licorice that shows promising antiviral activity towards human gammaherpesviruses. Secondly, we discuss anti-herpetic mechanisms utilized by several natural products in molecular level. While nucleoside analogues inhibit replicating herpesviruses in lytic replication, some natural products can disrupt the herpesvirus latent infection in the host cell. In addition, natural products can stimulate immune responses against herpesviral infection. These findings suggest that natural products could be one of the best choices for development of new treatments for latent herpesvirus infection, and may provide synergistic anti-viral activity when supplemented with nucleoside analogues. Therefore, it is important to identify which natural products are more efficacious anti-herpetic agents, and to understand the molecular mechanism in detail for further advance in the anti-viral therapies.

  17. Potential of RNA aptamers in the prevention of HIV-1 subtype C infections

    CSIR Research Space (South Africa)

    London, GM

    2014-10-01

    Full Text Available Compounds that have been used to prevent human immunodeficiency virus type-I (HIV-1) infections include synthetic chemicals, plant extras and monoclonal antibodies. Although most of these compounds have potent antiviral activity, they often fail...

  18. Fungal infection intensity and zoospore output of Atelopus zeteki, a potential acute chytrid supershedder.

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    Graziella V Direnzo

    Full Text Available Amphibians vary in their response to infection by the amphibian-killing chytrid fungus, Batrachochytrium dendrobatidis (Bd. Highly susceptible species are the first to decline and/or disappear once Bd arrives at a site. These competent hosts likely facilitate Bd proliferation because of ineffective innate and/or acquired immune defenses. We show that Atelopus zeteki, a highly susceptible species that has undergone substantial population declines throughout its range, rapidly and exponentially increases skin Bd infection intensity, achieving intensities that are several orders of magnitude greater than most other species reported. We experimentally infected individuals that were never exposed to Bd (n = 5 or previously exposed to an attenuated Bd strain (JEL427-P39; n = 3. Within seven days post-inoculation, the average Bd infection intensity was 18,213 zoospores (SE: 9,010; range: 0 to 66,928. Both average Bd infection intensity and zoospore output (i.e., the number of zoospores released per minute by an infected individual increased exponentially until time of death (t50 = 7.018, p<0.001, t46 = 3.164, p = 0.001, respectively. Mean Bd infection intensity and zoospore output at death were 4,334,422 zoospores (SE: 1,236,431 and 23.55 zoospores per minute (SE: 22.78, respectively, with as many as 9,584,158 zoospores on a single individual. The daily percent increases in Bd infection intensity and zoospore output were 35.4% (SE: 0.05 and 13.1% (SE: 0.04, respectively. We also found that Bd infection intensity and zoospore output were positively correlated (t43 = 3.926, p<0.001. All animals died between 22 and 33 days post-inoculation (mean: 28.88; SE: 1.58. Prior Bd infection had no effect on survival, Bd infection intensity, or zoospore output. We conclude that A. zeteki, a highly susceptible amphibian species, may be an acute supershedder. Our results can inform epidemiological models to estimate Bd outbreak probability, especially as they relate

  19. Universal newborn screening for congenital CMV infection: what is the evidence of potential benefit?†

    OpenAIRE

    Cannon, Michael J.; Griffiths, Paul D.; Aston, Van; Rawlinson, William D.

    2014-01-01

    Congenital CMV infection is a leading cause of childhood disability. Many children born with congenital CMV infection are asymptomatic or have nonspecific symptoms and therefore are typically not diagnosed. A strategy of newborn CMV screening could allow for early detection and intervention to improve clinical outcomes. Interventions might include antiviral drugs or nonpharmaceutical therapies such as speech-language therapy or cochlear implants. Using published data from developed countries,...

  20. Serum sphingomyelin has potential to reflect hepatic injury in chronic hepatitis B virus infection

    OpenAIRE

    Su-Jun Zheng; Feng Qu; Jun-Feng Li; Jing Zhao; Jing-Yun Zhang; Mei Liu; Feng Ren; Yu Chen; Jin-Lan Zhang; Zhong-Ping Duan

    2015-01-01

    Objective: To explore the relation between serum sphingolipids and hepatic injury in chronic HBV infection. Methods: A cohort of participants including 48 healthy persons, 103 chronic HBV-infected patients containing chronic hepatitis B (CHB) and HBV-related cirrhosis were included. High performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was performed to detect serum sphingolipids. The serological indicators were detected and quantified. The valid liver biop...

  1. Preclinical evaluation of two 68Ga-siderophores as potential radiopharmaceuticals for Aspergillus fumigatus infection imaging

    International Nuclear Information System (INIS)

    Petrik, Milos; Franssen, Gerben M.; Laverman, Peter; Haas, Hubertus; Schrettl, Markus; Hoertnagl, Caroline; Lass-Floerl, Cornelia; Helbok, Anna; Decristoforo, Clemens

    2012-01-01

    Invasive pulmonary aspergillosis is mainly caused by Aspergillus fumigatus, and is one of the major causes of morbidity and mortality in immunocompromised patients. The mortality associated with invasive pulmonary aspergillosis remains high, mainly due to the difficulties and limitations in diagnosis. We have shown that siderophores can be labelled with 68 Ga and can be used for PET imaging of A. fumigatus infection in rats. Here we report on the further evaluation of the most promising 68 Ga-siderophore candidates, triacetylfusarinine (TAFC) and ferrioxamine E (FOXE). Siderophores were labelled with 68 Ga using acetate buffer. Log P, protein binding and stability values were determined. Uptake by A. fumigatus was studied in vitro in cultures with high and low iron loads. In vivo biodistribution was determined in normal mice and an infection model was established using neutropenic rats inoculated with A. fumigatus. Static and dynamic μPET imaging was performed and correlated with CT images, and lung infection was evaluated ex vivo. 68 Ga-siderophores were labelled with high radiochemical purity and specific activity. 68 Ga-TAFC and 68 Ga-FOXE showed high uptake by A. fumigatus in iron-deficient cultures. In normal mice, 68 Ga-TAFC and 68 Ga-FOXE showed rapid renal excretion with high metabolic stability. In the rat infection model focal lung uptake was detected by μPET with both compounds and increased with severity of the infection, correlating with abnormal CT images. 68 Ga-TAFC and 68 Ga-FOXE displayed excellent in vitro stability and high uptake by A. fumigatus. Both compounds showed excellent pharmacokinetics, highly selective accumulation in infected lung tissue and good correlation with severity of disease in a rat infection model, which makes them promising agents for A. fumigatus infection imaging. (orig.)

  2. Parasites of the flounder Platichthys flesus (L.) from the German Bight, North Sea, and their potential use in ecosystem monitoring. A. Infection characteristics of potential indicator species

    Science.gov (United States)

    Schmidt, V.; Zander, S.; Körting, W.; Steinhagen, D.

    2003-10-01

    As part of integrated biological-effect monitoring, the parasite fauna of the flounder Platichthys flesus (L.) was investigated at five locations in the German Bight, with a view to using parasite species as bio-indicators. Over a period of 6 years, parasites from 30 different taxa were identified, but only 7 taxa of the parasite community occurred regularly at all locations and in sufficient abundance that they could be considered as potential indicator species. These species were the ciliophoran Trichodina spp., the copepods Acanthochondria cornuta, Lepeophtheirus pectoralis and Lernaeocera branchialis, the helminths Zoogonoides viviparus and Cucullanus heterochrous and metacercaria of an unidentified digenean species. Infection characteristics of these parasites are presented, with a comparison of the results from individual sampling periods and those of the long-term data set. Natural influences on the infection levels, such as temporal variations, habitat conditions and host-related factors, were evaluated. All of these parasite species showed significant differences in their infection levels between the Elbe estuary, as the most polluted site, and the less polluted coastal and marine locations: Helgoland, Outer Eider estuary and Spiekeroog, especially in the long-term data set. Gradual differences between the Elbe, the Outer Eider and Helgoland, which were not detected in individual sampling periods, also became evident in the pooled-data set. These were found in the prevalence of Trichodina spp., A. cornuta, Z. viviparus and C. heterochrous. Although salinity is considered as the most important natural factor, influencing the distribution pattern of the majority of the potential indicator species, infection levels of most of these species differed between locations with similar salinity conditions. Infection levels corresponded to a contamination gradient (Elbe > Inner Eider, Outer Eider > Helgoland) established across the locations. Seasonal variation in

  3. Pertussis Toxin Exploits Host Cell Signaling Pathways Induced by Meningitis-Causing E. coli K1-RS218 and Enhances Adherence of Monocytic THP-1 Cells to Human Cerebral Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Laura Julia Starost

    2016-10-01

    Full Text Available Pertussis toxin (PTx, the major virulence factor of the whooping cough-causing bacterial pathogen Bordetella pertussis, permeabilizes the blood–brain barrier (BBB in vitro and in vivo. Breaking barriers might promote translocation of meningitis-causing bacteria across the BBB, thereby facilitating infection. PTx activates several host cell signaling pathways exploited by the neonatal meningitis-causing Escherichia coli K1-RS218 for invasion and translocation across the BBB. Here, we investigated whether PTx and E. coli K1-RS218 exert similar effects on MAPK p38, NF-κB activation and transcription of downstream targets in human cerebral endothelial TY10 cells using qRT-PCR, Western blotting, and ELISA in combination with specific inhibitors. PTx and E. coli K1-RS218 activate MAPK p38, but only E. coli K1-RS218 activates the NF-κB pathway. mRNA and protein levels of p38 and NF-κB downstream targets including IL-6, IL-8, CxCL-1, CxCL-2 and ICAM-1 were increased. The p38 specific inhibitor SB203590 blocked PTx-enhanced activity, whereas E. coli K1-RS218’s effects were inhibited by the NF-κB inhibitor Bay 11-7082. Further, we found that PTx enhances the adherence of human monocytic THP-1 cells to human cerebral endothelial TY10 cells, thereby contributing to enhanced translocation. These modulations of host cell signaling pathways by PTx and meningitis-causing E. coli support their contributions to pathogen and monocytic THP-1 cells translocation across the BBB.

  4. Immunomodulatory role for membrane vesicles released by THP-1 macrophages and respiratory pathogens during macrophage infection.

    Science.gov (United States)

    Volgers, Charlotte; Benedikter, Birke J; Grauls, Gert E; Savelkoul, Paul H M; Stassen, Frank R M

    2017-11-13

    During infection, inflammation is partially driven by the release of mediators which facilitate intercellular communication. Amongst these mediators are small membrane vesicles (MVs) that can be released by both host cells and Gram-negative and -positive bacteria. Bacterial membrane vesicles are known to exert immuno-modulatory and -stimulatory actions. Moreover, it has been proposed that host cell-derived vesicles, released during infection, also have immunostimulatory properties. In this study, we assessed the release and activity of host cell-derived and bacterial MVs during the first hours following infection of THP-1 macrophages with the common respiratory pathogens non-typeable Haemophilus influenzae, Moraxella catarrhalis, Streptococcus pneumoniae, and Pseudomonas aeruginosa. Using a combination of flow cytometry, tunable resistive pulse sensing (TRPS)-based analysis and electron microscopy, we demonstrated that the release of MVs occurs by both host cells and bacteria during infection. MVs released during infection and bacterial culture were found to induce a strong pro-inflammatory response by naive THP-1 macrophages. Yet, these MVs were also found to induce tolerance of host cells to secondary immunogenic stimuli and to enhance bacterial adherence and the number of intracellular bacteria. Bacterial MVs may play a dual role during infection, as they can both trigger and dampen immune responses thereby contributing to immune defence and bacterial survival.

  5. The civRT operon is important for Campylobacter jejuni strain 81-176 host cell interactions through regulation of the formate dehydrogenase operon

    Science.gov (United States)

    C. jejuni colonizes the intestinal mucosa, and the severity of disease in different strains is correlated with host cell interaction and invasion. A microarray screen to identify genes differentially regulated during C. jejuni interaction with tissue culture cells revealed the up-regulation of a two...

  6. Human gastric mucins differently regulate Helicobacter pylori proliferation, gene expression and interactions with host cells.

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    Emma C Skoog

    Full Text Available Helicobacter pylori colonizes the mucus niche of the gastric mucosa and is a risk factor for gastritis, ulcers and cancer. The main components of the mucus layer are heavily glycosylated mucins, to which H. pylori can adhere. Mucin glycosylation differs between individuals and changes during disease. Here we have examined the H. pylori response to purified mucins from a range of tumor and normal human gastric tissue samples. Our results demonstrate that mucins from different individuals differ in how they modulate both proliferation and gene expression of H. pylori. The mucin effect on proliferation varied significantly between samples, and ranged from stimulatory to inhibitory, depending on the type of mucins and the ability of the mucins to bind to H. pylori. Tumor-derived mucins and mucins from the surface mucosa had potential to stimulate proliferation, while gland-derived mucins tended to inhibit proliferation and mucins from healthy uninfected individuals showed little effect. Artificial glycoconjugates containing H. pylori ligands also modulated H. pylori proliferation, albeit to a lesser degree than human mucins. Expression of genes important for the pathogenicity of H. pylori (babA, sabA, cagA, flaA and ureA appeared co-regulated in response to mucins. The addition of mucins to co-cultures of H. pylori and gastric epithelial cells protected the viability of the cells and modulated the cytokine production in a manner that differed between individuals, was partially dependent of adhesion of H. pylori to the gastric cells, but also revealed that other mucin factors in addition to adhesion are important for H. pylori-induced host signaling. The combined data reveal host-specific effects on proliferation, gene expression and virulence of H. pylori due to the gastric mucin environment, demonstrating a dynamic interplay between the bacterium and its host.

  7. High Shedding Potential and Significant Individual Heterogeneity in Naturally-Infected Alpine ibex (Capra ibex With Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Sébastien Lambert

    2018-05-01

    Full Text Available Wildlife reservoirs of infectious diseases raise major management issues. In Europe, brucellosis has been eradicated in domestic ruminants from most countries and wild ruminants have not been considered important reservoirs so far. However, a high prevalence of Brucella melitensis infection has been recently identified in a French population of Alpine ibex (Capra ibex, after the emergence of brucellosis was confirmed in a dairy cattle farm and two human cases. This situation raised the need to identify the factors driving the persistence of Brucella infection at high prevalence levels in this ibex population. In the present paper, we studied the shedding pattern of B. melitensis in ibex from Bargy Massif, French Alps. Bacteriological examinations (1–15 tissues/samples per individual were performed on 88 seropositive, supposedly infected and euthanized individuals. Among them, 51 (58% showed at least one positive culture, including 45 ibex with at least one Brucella isolation from a urogenital sample or a lymph node in the pelvic area (active infection in organs in the pelvic area. Among these 45 ibex, 26 (30% of the total number of necropsied animals showed at least one positive culture for a urogenital organ and were considered as being at risk of shedding the bacteria at the time of capture. We observed significant heterogeneity between sex-and-age classes: seropositive females were most at risk to excrete Brucella before the age of 5 years, possibly corresponding to abortion during the first pregnancy following infection such as reported in the domestic ruminants. The high shedding potential observed in young females may have contributed to the self-sustained maintenance of infection in this population, whereas males are supposed to play a role of transmission between spatial units through venereal transmission during mating. This heterogeneity in the shedding potential of seropositive individuals should be considered in the future to

  8. Fiber mediated receptor masking in non-infected bystander cells restricts adenovirus cell killing effect but promotes adenovirus host co-existence.

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    Johan Rebetz

    Full Text Available The basic concept of conditionally replicating adenoviruses (CRAD as oncolytic agents is that progenies generated from each round of infection will disperse, infect and kill new cancer cells. However, CRAD has only inhibited, but not eradicated tumor growth in xenograft tumor therapy, and CRAD therapy has had only marginal clinical benefit to cancer patients. Here, we found that CRAD propagation and cancer cell survival co-existed for long periods of time when infection was initiated at low multiplicity of infection (MOI, and cancer cell killing was inefficient and slow compared to the assumed cell killing effect upon infection at high MOI. Excessive production of fiber molecules from initial CRAD infection of only 1 to 2% cancer cells and their release prior to the viral particle itself caused a tropism-specific receptor masking in both infected and non-infected bystander cells. Consequently, the non-infected bystander cells were inefficiently bound and infected by CRAD progenies. Further, fiber overproduction with concomitant restriction of adenovirus spread was observed in xenograft cancer therapy models. Besides the CAR-binding Ad4, Ad5, and Ad37, infection with CD46-binding Ad35 and Ad11 also caused receptor masking. Fiber overproduction and its resulting receptor masking thus play a key role in limiting CRAD functionality, but potentially promote adenovirus and host cell co-existence. These findings also give important clues for understanding mechanisms underlying the natural infection course of various adenoviruses.

  9. iNKT Cells and Their potential Lipid Ligands during Viral Infection

    Directory of Open Access Journals (Sweden)

    Anunya eOpasawatchai

    2015-07-01

    Full Text Available Invariant natural killer T (iNKT cells are a unique population of lipid reactive CD1d restricted innate-like T lymphocytes. Despite being a minor population, they serve as an early source of cytokines and promote immunological crosstalk thus bridging innate and adaptive immunity. Diseases ranging from allergy, autoimmunity, and cancer as well as infectious diseases, including viral infection, have been reported to be influenced by iNKT cells. However, it remains unclear how iNKT cells are activated during viral infection, as virus derived lipid antigens have not been reported. Cytokines may activate iNKT cells during infections from influenza and murine cytomegalovirus (MCMV, although CD1d dependent activation is evident in other viral infections. Several viruses, such as dengue virus (DENV, induce CD1d upregulation which correlates with iNKT cell activation. In contrast, Herpes simplex virus type 1 (HSV-1, Human immunodeficiency virus (HIV, Epstein-Barr virus (EBV and Human papiloma virus (HPV promote CD1d downregulation as a strategy to evade iNKT cell recognition. These observations suggest the participation of a CD1d-dependent process in the activation of iNKT cells in response to viral infection. Endogenous lipid ligands, including phospholipids as well as glycosphingolipids, such as glucosylceramide have been proposed to mediate iNKT cell activation. Pro-inflammatory signals produced during viral infection may stimulate iNKT cells through enhanced CD1d dependent endogenous lipid presentation. Furthermore, viral infection may alter lipid composition and inhibit endogenous lipid degradation. Recent advances in this field are reviewed.

  10. Potential Association of IL1B Polymorphism With Iron Deficiency Risk in Childhood Helicobacter pylori Infection.

    Science.gov (United States)

    Chen, Szu-Ta; Ni, Yen-Hsuan; Liu, Shing-Hwa

    2018-02-01

    Helicobacter pylori infection occurs predominantly in childhood. Host immune response gene polymorphism is reported to affect the susceptibility to H pylori infection and the outcome of H pylori-related gastric cancer. Not all H pylori-infected patients, however, exhibit iron deficiency (ID). The relationship between host genetic polymorphisms and ID mediated by H pylori infection is not well understood. Subjects (n = 644) from the general population of age 10 to 18 years were divided into 2 groups based on serology testing for anti-H pylori IgG: seropositive study group; and seronegative control group. Five single nucleotide polymorphisms (SNPs) in IL1B (rs1143627 and rs16944), IL8 (rs4073), IL10 (rs1800896), and ABO (rs505922), were genotyped and the iron status of the 2 groups was compared. The seroprevalence rate for H pylori was 10.7% in this study. Infected subjects were significantly older and had lower serum iron levels than uninfected subjects (P = 0.0195 and 0.0059, respectively). Multivariate analysis revealed a significantly higher frequency of the T allele of rs505922 (odds ratio [OR] = 6.128; P < 0.001) and lower frequency of the T allele of rs1143627 (OR = 0.846; P = 0.014) in seropositive subjects. Among 59 seropositive subjects, the T allele frequency of rs1143627 was significantly higher in those with ID (OR = 3.156; P = 0.043), compared with those without ID. ABO (rs505922) and IL1B (rs1143627) may affect H pylori infection susceptibility, and IL1B (rs1143627) may also influence ID risk in infected children.

  11. Radiosynthesis and biodistribution of 99mTcN-Garenoxacin dithiocarbamate complex a potential infection imaging agent

    International Nuclear Information System (INIS)

    Syed Qaiser Shah; Aakif Ullah Khan; Muhammad Rafiullah Khan

    2011-01-01

    Garenoxacin (GXN) was modified to its dithiocarbamate followed by radiolabeling with technetium-99m ( 99m Tc) through [ 99m Tc-N] 2+ core. The suitability of the 99m TcN-Garenoxacin dithiocarbamate (GXND) complex as a potential multiresistant Staphylococcus aureus (MDRSA) and penicillin-resistant Streptococci (PRSC) infection radiotracer was assessed in artificially infected rats (AFRT). The radiolabeled complex was investigated for its radiochemical purity (RCP), permanence in serum using HPLC and TLC methods. In vitro binding with MDRSA and PRSC was performed at 37 deg C. The 99m TcN-GXND showed maximum RCP of 98.00 ± 0.22% and remained more than 90% stable up to 4 h. The 99m TcN-GXND showed saturated in vitro binding with living MDRSA and PRSC, respectively. The complex showed normal biodistribution in healthy rats (HRT), however in AFRT, seven fold uptakes was observed in infected muscle as compared to inflamed and normal muscles. Based on the high RCP, stability in serum, better in vitro binding with bacteria, biodistribution behavior and the target to non-target (infected to inflamed muscle) ratio, we recommend the 99m TcN-GXND complex for in vivo investigation of MDRSA and PRSC infection in human. (author)

  12. Epilepsy after cerebral infection: review of the literature and the potential for surgery.

    Science.gov (United States)

    Ramantani, Georgia; Holthausen, Hans

    2017-06-01

    The risk of unprovoked seizures in population-based cohorts of cerebral infection survivors is 7-8% in developed countries, rising to considerably higher rates in resource-poor countries. The main risk factors for epilepsy after cerebral infection, besides acute seizures, are infection-associated brain lesions and status epilepticus during the acute phase. Despite the high prevalence of pharmacoresistant epilepsies after cerebral infections, especially in patients with MRI-identifiable lesions, only a small minority undergoes epilepsy surgery. However, excellent surgical candidates are particularly those with a history of meningitis or encephalitis in early childhood, hippocampal sclerosis on MRI, as well as a history, seizure semiology, and EEG-findings compatible with the diagnosis of a mesial temporal lobe epilepsy syndrome. More challenging are patients with neocortical/extratemporal lobe epilepsies post cerebral infection. Finally, patients with a severe hemispheric injury with contralateral hemiparesis are candidates for hemispherectomy/hemispherotomy. This review attempts to shed some light on this frequent cause of symptomatic focal epilepsy, with an emphasis on the chances offered by epilepsy surgery.

  13. Construction and expression of an anti-VEGFR2 Nanobody-Fc fusionbody in NS0 host cell.

    Science.gov (United States)

    Qasemi, Maryam; Behdani, Mahdi; Shokrgozar, Mohammad Ali; Molla-Kazemiha, Vahid; Mohseni-Kuchesfahani, Homa; Habibi-Anbouhi, Mahdi

    2016-07-01

    Angiogenesis is the formation of new blood vessels which is involved in migration, growth and differentiation of endothelial cells. This process regularly occurs during growth and development in children however, in adults is usually part of a disease process such as cancer. The vascular endothelial growth factor (VEGF) is a vital player in the vascular development and angiogenesis in physiological and pathological processes. Camelid's immune system has unique antibodies which are composed of only a heavy chain homodimer and the variable domain (VHH, Nanobody). Nanobodies are small, around 15 kDa and stable. In this study, we engineered and constructed a new Nanobody-Fc fusion protein (fusionbody) composed of an anti-VEGFR2 Nanobody and an Fc fragment of human IgG1 antibody. The recombinant vector was transfected into NS0 host cells. Stable producer clones were developed and the recombinant fusionbody was expressed and purified. Functional assay showed the anti-VEGFR2 fusionbody could bind to VEGFR2 on cell surface via VHH part and could mediate killing the targeted cells through direct cell death and complement-dependent cytotoxicity (CDC). Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Full automation and validation of a flexible ELISA platform for host cell protein and protein A impurity detection in biopharmaceuticals.

    Science.gov (United States)

    Rey, Guillaume; Wendeler, Markus W

    2012-11-01

    Monitoring host cell protein (HCP) and protein A impurities is important to ensure successful development of recombinant antibody drugs. Here, we report the full automation and validation of an ELISA platform on a robotic system that allows the detection of Chinese hamster ovary (CHO) HCPs and residual protein A of in-process control samples and final drug substance. The ELISA setup is designed to serve three main goals: high sample throughput, high quality of results, and sample handling flexibility. The processing of analysis requests, determination of optimal sample dilutions, and calculation of impurity content is performed automatically by a spreadsheet. Up to 48 samples in three unspiked and spiked dilutions each are processed within 24 h. The dilution of each sample is individually prepared based on the drug concentration and the expected impurity content. Adaptable dilution protocols allow the analysis of sample dilutions ranging from 1:2 to 1:2×10(7). The validity of results is assessed by automatic testing for dilutional linearity and spike recovery for each sample. This automated impurity ELISA facilitates multi-project process development, is easily adaptable to other impurity ELISA formats, and increases analytical capacity by combining flexible sample handling with high data quality. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. Genetic recombination of Herpes simplex virus, the role of the host cell and UV-irradiation of the virus

    International Nuclear Information System (INIS)

    Dasgupta, U.B.; Summers, W.C.; Yale Univ., New Haven, CT; Yale Univ., New Haven, CT

    1980-01-01

    Recombination frequencies for two sets of genetic markers of Herpes simplex virus were determined in various host cells with and without ultraviolet irradiation of the virus. UV irradiation increased the recombination frequency in all the cell types studied in direct proportion to the unrepaired lethal damage. In human skin fibroblasts derived from a patient with xeroderma pigmentosum (XP) of complementation group A, a given dose of UV stimulated recombination more than that in fibroblasts from normal individuals. On the other hand, UV stimulation of HSV recombination was slightly less than normal in fibroblasts derived from a patient with a variant form XP and from an ataxia telangiectasia patient. Caffeine, an agent known to inhibit repair of UV damage, reduced recombination in most of the cell types studied but did not suppress the UV-induced increase in recombination. These findings suggest that for virus DNA with the same number of unrepaired UV-lesions, each of the tested cell types promoted HSV-recombination to an equivalent extent. (orig.) [de

  16. Evaluation of infection imaging potential of 131I-labeled imidazolium salt

    International Nuclear Information System (INIS)

    Ayca Tuncel; Fatma Yurt; Osman Yilmaz; Ismail Oeztuerk

    2018-01-01

    Effective antimicrobial compounds are necessary due to increased resistance of antibiotics against microorganisms causing infectious diseases. In this study, imidazolium-TFSI salt [ITFSI: octyl-bis(3-methylimidazolium)-di(bis(trifluoromethane)sulfonimide)] was labeled with 131 I with high efficiency. In vitro uptake experiments of 131 I-ITFSI showed high uptake in gram-positive Staphylococcus aureus bacteria. 131 I-ITFSI was also evaluated for comparison between bacterial infection and sterile inflammation by in vivo studies. The biodistribution results revealed that 131 I-ITFSI might be used as a nuclear imaging agent for detection of bacterial infection. (author)

  17. Assessment of the potential risk of infection associated with Clostridium difficile from porcine xenografts

    Czech Academy of Sciences Publication Activity Database

    Bakri, M.M.; Sutherland, A.D.; Brown, D.J.; Veselý, Pavel; Crossan, C.; Scobie, L.

    2009-01-01

    Roč. 16, č. 6 (2009), s. 472-476 ISSN 0908-665X Grant - others:EC(XE) LSHB-CT-2006-037377 Institutional research plan: CEZ:AV0Z50520514 Keywords : C. difficile * hospital acquired infection * xenotransplant * zoonoses Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.711, year: 2009

  18. Infection resistance of degradable versus non-degradable biomaterials : An assessment of the potential mechanisms

    NARCIS (Netherlands)

    Daghighi, Seyedmojtaba; Sjollema, Jelmer; van der Mei, Henny C.; Busscher, Henk J.; Rochford, Edward T. J.

    Extended life expectancy and medical development has led to an increased reliance on biomaterial implants and devices to support or restore human anatomy and function. However, the presence of an implanted biomaterial results in an increased susceptibility to infection. Due to the severity of the

  19. Mining RNA-seq data for infections and contaminations.

    Directory of Open Access Journals (Sweden)

    Thomas Bonfert

    Full Text Available RNA sequencing (RNA-seq provides novel opportunities for transcriptomic studies at nucleotide resolution, including transcriptomics of viruses or microbes infecting a cell. However, standard approaches for mapping the resulting sequencing reads generally ignore alternative sources of expression other than the host cell and are little equipped to address the problems arising from redundancies and gaps among sequenced microbe and virus genomes. We show that screening of sequencing reads for contaminations and infections can be performed easily using ContextMap, our recently developed mapping software. Based on mapping-derived statistics, mapping confidence, similarities and misidentifications (e.g. due to missing genome sequences of species/strains can be assessed. Performance of our approach is evaluated on three real-life sequencing data sets and compared to state-of-the-art metagenomics tools. In particular, ContextMap vastly outperformed GASiC and GRAMMy in terms of runtime. In contrast to MEGAN4, it was capable of providing individual read mappings to species and resolving non-unique mappings, thus allowing the identification of misalignments caused by sequence similarities between genomes and missing genome sequences. Our study illustrates the importance and potentials of routinely mining RNA-seq experiments for infections or contaminations by microbes and viruses. By using ContextMap, gene expression of infecting agents can be analyzed and novel insights in infection processes and tumorigenesis can be obtained.

  20. Seroprevalence, genotypic distribution and potential risk factors of hepatitis B and C virus infections among adults in Siem Reap, Cambodia.

    Science.gov (United States)

    Yamada, Hiroko; Fujimoto, Mayumi; Svay, Somana; Lim, Olline; Hok, Sirany; Goto, Noboru; Ohisa, Masayuki; Akita, Tomoyuki; Matsuo, Junko; Do, Son Huy; Katayama, Keiko; Miyakawa, Yuzo; Tanaka, Junko

    2015-04-01

    We investigated hepatitis B virus (HBV) and hepatitis C virus (HCV) infections among adults in Siem Reap, Cambodia, to consider the prevention strategy in cooperation with the Ministry of Health in Cambodia. Serological tests for determining HBV and HCV infections and questionnaires were performed from 2010 to 2012 among the general population in the province of Siem Reap. Multivariate logistic regression analysis was conducted to clarify the factors related to HBV and HCV infections. There were 483 participants, comprising 194 men and 289 women (age range, 18-89 years). The prevalence of hepatitis B surface antigen was not very high at 4.6%, while anti-hepatitis B core (anti-HBc) was high at 38.5%. All HBV DNA samples were classified as genotype C. Anti-HBc showed the trend that the older the age, the higher the positive rate (P = 0.0002). The prevalence of HCV RNA and anti-HCV were 2.3% and 5.8%, respectively. HCV RNA was detected in 39.3% of anti-HCV positive samples and most of them were classified as genotype 6 (54.5%) and 1 (27.3%). Remarkably, in multivariate logistic regression analysis, history of operation and blood transfusion were significantly associated with the positivity for HBV infection and HCV RNA, respectively. Our results showed that operation and blood transfusion were potential risk factors for HBV and HCV infection, respectively, and supposed that horizontal HBV transmission may be frequent in adults in Cambodia. Hence, for reducing HBV and HCV infections, it is necessary to improve the safety of blood and medical treatment. © 2014 The Japan Society of Hepatology.

  1. Radiosynthesis and biological evaluation of 99mTcN-sitafloxacin dithiocarbamate as a potential radiotracer for Staphylococcus aureus infection

    International Nuclear Information System (INIS)

    Syed Qaiser Shah; Aakif Ullah Khan; Muhammad Rafiullah Khan

    2011-01-01

    Sitafloxacin dithocarbamate (SFDE) was synthesized, radiolabeled with technetium-99m ( 99m Tc) using [ 99m Tc-N] 2+ core and evaluated its biological efficacy as a potential radiotracer for Staphylococcus aureus (S. aureus) infection in artificially infected rats (AIRT) and rabbits (AIRB). The radiochemical stability of the 99m Tc labeled SFDE ( 99m TcN-SFDE) in saline and serum was determined by radio-HPLC and TLC methods, respectively. After, 1 min of reconstitution the value of radiochemical purity (RCP) was 99.00 ± 0.20% and was remained more than 90% unwavering even after 240 min of the radiolabeling. The 99m TcN-SFDE complex showed similar radiochemical permanence behavior in serum at 37 deg C. The complex showed almost six fold higher specific in vitro binding with living than heat killed S. aureus. Biodistribution behavior was evaluated in S. aureus AIRT and whole body imaging (WBI) in AIRB, respectively. Seven fold up take was observed in infected muscle of the AIRT as compared to inflamed and normal muscles. The disappearance of activity from blood and appearance in urinary system indicated normal route of excretion of the complex. Scintigraphically, it was confirmed that the labeled SFDE was higher accumulated in the infected muscle higher than in inflamed and normal muscle. The high radiochemical stability in saline and serum, specific in vitro binding with S. aureus, precise in vivo distribution in S. aureus AIRT and targeted WBI in AIRB confirmed the possibility of the 99m TcN-SFDE complex as a potential and promising S. aureus infection radiotracer. (author)

  2. [Aetiology and outcomes of potentially serious infections in febrile infants less than 3 months old].

    Science.gov (United States)

    de la Torre, Mercedes; de Lucas, Nieves; Velasco, Roberto; Gómez, Borja; Mintegi, Santiago

    2017-07-01

    Recent studies have shown changes in the aetiology of serious bacterial infections in febrile infants ≤ 90 days of age. The aim of this study was to describe the current microbiology and outcomes of these infections in Spain. Sub-analysis of a prospective multicentre study focusing on febrile infants of less than 91 days of life, admitted between October 2011 and September 2013 to Emergency Departments of 19 Spanish hospitals, members of the Spanish Paediatric Emergency Research Group of the Spanish Society of Paediatric Emergencies (RISeuP/SPERG). The analysis included 3,401 febrile infants ≤90 days of age with fever without source. There were 896 positive cultures: 766 urine (85.5%), 100 blood (11.2%), 18 cerebrospinal fluid (2%), 10 stool, and 2 umbilical cultures. Among the 3,401 infants included, 784 (23%) were diagnosed with a serious bacterial infection, and 107 of them (3.1%) with an invasive infection. E. coli was the most common pathogen isolated from urine (628; 82%), blood (46; 46%), and cerebrospinal fluid cultures (7; 38.9%), followed by S. agalactiae that was isolated from 24 (24%) blood cultures and 3 (16.7%) cerebrospinal fluid cultures. There were only 2 L. monocytogenes infections. Four children died, and seven had severe complications. Among infants ≤ 90 days of age with fever without source, E. coli was the most common pathogen isolated from urine, blood, and cerebrospinal fluid cultures. Copyright © 2016 Asociación Española de Pediatría. Publicado por Elsevier España, S.L.U. All rights reserved.

  3. Experimental Infection of Rhodnius prolixus (Hemiptera, Triatominae with Mycobacterium leprae Indicates Potential for Leprosy Transmission.

    Directory of Open Access Journals (Sweden)

    Arthur da Silva Neumann

    Full Text Available Leprosy is a chronic dermato-neurological disease caused by infection with Mycobacterium leprae. In 2013 almost 200,000 new cases of leprosy were detected around the world. Since the first symptoms take from years to decades to appear, the total number of asymptomatic patients is impossible to predict. Although leprosy is one of the oldest records of human disease, the mechanisms involved with its transmission and epidemiology are still not completely understood. In the present work, we experimentally investigated the hypothesis that the mosquitoes Aedes aegypti and Culex quinquefasciatus and the hemiptera Rhodnius prolixus act as leprosy vectors. By means of real-time PCR quantification of M. leprae 16SrRNA, we found that M. leprae remained viable inside the digestive tract of Rhodnius prolixus for 20 days after oral infection. In contrast, in the gut of both mosquito species tested, we were not able to detect M. leprae RNA after a similar period of time. Inside the kissing bug Rhodnius prolixus digestive tract, M. leprae was initially restricted to the anterior midgut, but gradually moved towards the hindgut, in a time course reminiscent of the life cycle of Trypanosoma cruzi, a well-known pathogen transmitted by this insect. The maintenance of M. leprae infectivity inside the digestive tract of this kissing bug is further supported by successful mice footpad inoculation with feces collected 20 days after infection. We conclude that Rhodnius prolixus defecate infective M. leprae, justifying the evaluation of the presence of M. leprae among sylvatic and domestic kissing bugs in countries endemic for leprosy.

  4. Experimental Infection of Rhodnius prolixus (Hemiptera, Triatominae) with Mycobacterium leprae Indicates Potential for Leprosy Transmission.

    Science.gov (United States)

    Neumann, Arthur da Silva; Dias, Felipe de Almeida; Ferreira, Jéssica da Silva; Fontes, Amanda Nogueira Brum; Rosa, Patricia Sammarco; Macedo, Rafael Enrique; Oliveira, José Henrique; Teixeira, Raquel Lima de Figueiredo; Pessolani, Maria Cristina Vidal; Moraes, Milton Ozório; Suffys, Philip Noel; Oliveira, Pedro L; Sorgine, Marcos Henrique Ferreira; Lara, Flavio Alves

    2016-01-01

    Leprosy is a chronic dermato-neurological disease caused by infection with Mycobacterium leprae. In 2013 almost 200,000 new cases of leprosy were detected around the world. Since the first symptoms take from years to decades to appear, the total number of asymptomatic patients is impossible to predict. Although leprosy is one of the oldest records of human disease, the mechanisms involved with its transmission and epidemiology are still not completely understood. In the present work, we experimentally investigated the hypothesis that the mosquitoes Aedes aegypti and Culex quinquefasciatus and the hemiptera Rhodnius prolixus act as leprosy vectors. By means of real-time PCR quantification of M. leprae 16SrRNA, we found that M. leprae remained viable inside the digestive tract of Rhodnius prolixus for 20 days after oral infection. In contrast, in the gut of both mosquito species tested, we were not able to detect M. leprae RNA after a similar period of time. Inside the kissing bug Rhodnius prolixus digestive tract, M. leprae was initially restricted to the anterior midgut, but gradually moved towards the hindgut, in a time course reminiscent of the life cycle of Trypanosoma cruzi, a well-known pathogen transmitted by this insect. The maintenance of M. leprae infectivity inside the digestive tract of this kissing bug is further supported by successful mice footpad inoculation with feces collected 20 days after infection. We conclude that Rhodnius prolixus defecate infective M. leprae, justifying the evaluation of the presence of M. leprae among sylvatic and domestic kissing bugs in countries endemic for leprosy.

  5. Peptide inhibition of human cytomegalovirus infection

    Directory of Open Access Journals (Sweden)

    Morris Cindy A

    2011-02-01

    Full Text Available Abstract Background Human cytomegalovirus (HCMV is the most prevalent congenital viral infection in the United States and Europe causing significant morbidity and mortality to both mother and child. HCMV is also an opportunistic pathogen in immunocompromised individuals, including human immunodeficiency virus (HIV- infected patients with AIDS, and solid organ and allogeneic stem cell transplantation recipients. Current treatments for HCMV-associated diseases are insufficient due to the emergence of drug-induced resistance and cytotoxicity, necessitating novel approaches to limit HCMV infection. The aim of this study was to develop therapeutic peptides targeting glycoprotein B (gB, a major glycoprotein of HCMV that is highly conserved across the Herpesviridae family, that specifically inhibit fusion of the viral envelope with the host cell membrane preventing HCMV entry and infection. Results Using the Wimley-White Interfacial Hydrophobicity Scale (WWIHS, several regions within gB were identified that display a high potential to interact with lipid bilayers of cell membranes and hydrophobic surfaces within proteins. The ability of synthetic peptides analogous to WWIHS-positive sequences of HCMV gB to inhibit viral infectivity was evaluated. Human foreskin fibroblasts (HFF were infected with the Towne-GFP strain of HCMV (0.5 MOI, preincubated with peptides at a range of concentrations (78 nm to 100 μM, and GFP-positive cells were visualized 48 hours post-infection by fluorescence microscopy and analyzed quantitatively by flow cytometry. Peptides that inhibited HCMV infection demonstrated different inhibitory concentration curves indicating that each peptide possesses distinct biophysical properties. Peptide 174-200 showed 80% inhibition of viral infection at a concentration of 100 μM, and 51% and 62% inhibition at concentrations of 5 μM and 2.5 μM, respectively. Peptide 233-263 inhibited infection by 97% and 92% at concentrations of 100

  6. Infection-Induced Thrombin Production: A Potential Novel Mechanism for Preterm Premature Rupture of Membranes (PPROM).

    Science.gov (United States)

    Feng, Liping; Allen, Terrence K; Marinello, William P; Murtha, Amy P

    2018-04-13

    Preterm premature rupture of membranes (PPROM) is a leading contributor to maternal and neonatal morbidity and mortality. Epidemiologic and experimental studies have demonstrated that thrombin causes fetal membrane weakening and subsequently PPROM. Although blood is suspected as the likely source of thrombin in fetal membranes and amniotic fluid of patients with PPROM, this has not been proven. Ureaplasma Parvum (U. parvum) is emerging as a pathogen involved in prematurity, including PPROM, but until now, prothrombin production directly induced by bacteria in fetal membranes has not been described. This study was designed to investigate whether U. parvum exposure can induce prothrombin production in fetal membranes cells. Primary fetal membrane cells (amnion epithelial, chorion trophoblast, and decidua stromal) or full-thickness fetal membrane tissue explants from elective, term, uncomplicated cesarean deliveries were harvested. Cells or tissue explants were infected with live U. parvum (1 x 10 5 , 1 x 10 6 , or 1 x 10 7 colony forming units (cfu)/ml) or lipopolysaccharide (Escherichia coli J5, L-5014, Sigma, 100 ng/ml or 1000 ng/ml) for 24 hours. Tissue explants were fixed for immunohistochemistry staining of thrombin/prothrombin. Fetal membrane cells were fixed for confocal immunofluorescent staining of the biomarkers of fetal membrane cell types and thrombin/prothrombin. Protein and mRNA were harvested from the cells and tissue explants for Western blot or qRT-PCR to quantify thrombin/prothrombin protein or mRNA production, respectively. Data are presented as mean values ± standard errors of mean. Data were analyzed using one-way ANOVA with post hoc Dunnett's test. Prothrombin production and localization was confirmed by Western blot and immunostainings in all primary fetal membrane cells and tissue explants. Immunofluorescence observations revealed a perinuclear localization of prothrombin in amnion epithelial cells. Localization of prothrombin in chorion and

  7. Oligopeptide Targeting Sortase A as Potential Anti-infective Therapy for Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Jianfeng Wang

    2018-02-01

    Full Text Available Sortase A (SrtA-catalyzed anchorage of surface proteins in most Gram-positive bacteria is indispensable for their virulence, suggesting that this transpeptidase is a promising target for antivirulence therapy. Here, an oligopeptide, LPRDA, was identified as an effective inhibitor of SrtA via virtual screening based on the LPXTG substrate sequence, and it was found to inhibit SrtA activity in vitro and in vivo (IC50 = 10.61 μM by competitively occupying the active site of SrtA. Further, the oligopeptide treatment had no anti-Staphylococcus aureus activity, but it provided protection against S. aureus-induced mastitis in a mouse model. These findings indicate that the oligopeptide could be used as an effective anti-infective agent for the treatment of infection caused by S. aureus or other Gram-positive bacteria via the targeting of SrtA.

  8. Respiratory infections and pneumonia: potential benefits of switching from smoking to vaping.

    Science.gov (United States)

    Campagna, Davide; Amaradio, Maria Domenica; Sands, Mark F; Polosa, Riccardo

    2016-01-01

    Abstaining from tobacco smoking is likely to lower the risk of respiratory infections and pneumonia. Unfortunately, quitting smoking is not easy. Electronic cigarettes (ECs) are emerging as an attractive long-term alternative nicotine source to conventional cigarettes and are being adopted by smokers who wish to reduce or quit cigarette consumption. Also, given that the propylene glycol in EC aerosols is a potent bactericidal agent, switching from smoking to regular vaping is likely to produce additional lung health benefits. Here, we critically address some of the concerns arising from regular EC use in relation to lung health, including respiratory infections and pneumonia. In conclusion, smokers who quit by switching to regular ECs use can reduce risk and reverse harm from tobacco smoking. Innovation in the e-vapour category is likely not only to further minimise residual health risks, but also to maximise health benefits.

  9. Ultraviolet C Light for Acinetobacter baumannii Wound Infections in Mice: Potential Use for Battlefield Wound Decontamination?

    Science.gov (United States)

    2012-01-01

    al. Fiber optic microneedles for transdermal light delivery : ex vivo porcine skin penetration experiments. J Biomech Eng. 2010;132:091014. 24...critically ill patients often with limited treatment options because of drug resistance.1Y3 Since the beginning of the Middle East conflicts, the outbreak...of A. baumannii infections has been a threat to the health of wounded US soldiers.4Y7 The primary measures used in the military evacuation system to

  10. Acid suppression therapy does not predispose to Clostridium difficile infection: the case of the potential bias.

    Directory of Open Access Journals (Sweden)

    Lena Novack

    Full Text Available OBJECTIVE: An adverse effect of acid-suppression medications on the occurrence of Clostridium difficile infection (CDI has been a common finding of many, but not all studies. We hypothesized that association between acid-suppression medications and CDI is due to the residual confounding in comparison between patients with infection to those without, predominantly from non-tested and less sick subjects. We aimed to evaluate the effect of acid suppression therapy on incidence of CDI by comparing patients with CDI to two control groups: not tested patients and patients suspected of having CDI, but with a negative test. METHODS: We conducted a case-control study of adult patients hospitalized in internal medicine department of tertiary teaching hospital between 2005-2010 for at least three days. Controls from each of two groups (negative for CDI and non-tested were individually matched (1:1 to cases by primary diagnosis, Charlson comorbidity index, year of hospitalization and gender. Primary outcomes were diagnoses of International Classification of Diseases (ICD-9-coded CDI occurring 72 hours or more after admission. RESULTS: Patients with CDI were similar to controls with a negative test, while controls without CDI testing had lower clinical severity. In multivariable analysis, treatment by acid suppression medications was associated with CDI compared to those who were not tested (OR = 1.88, p-value = 0.032. Conversely, use of acid suppression medications in those who tested negative for the infection was not associated with CDI risk as compared to the cases (OR = 0.66; p = 0.059. CONCLUSIONS: These findings suggest that the reported epidemiologic associations between use of acid suppression medications and CDI risk may be spurious. The control group choice has an important impact on the results. Clinical differences between the patients with CDI and those not tested and not suspected of having the infection may explain the different conclusions

  11. Cytotoxic potential of decidual NK cells and CD8+ T cells awakened by infections.

    Science.gov (United States)

    Crespo, Ângela C; van der Zwan, Anita; Ramalho-Santos, João; Strominger, Jack L; Tilburgs, Tamara

    2017-02-01

    To establish a healthy pregnancy the maternal immune system must tolerate fetal allo-antigens, yet remain competent to respond to infections. The ability of decidual NK cells (dNK) to promote migration of fetal extravillous trophoblasts (EVT) and placental growth as well as the capacity of EVT to promote immune tolerance are topics of high interest and extensive research. However, the problem of how dNK and decidual CD8+ T cells (CD8+ dT) provide immunity to infections of the placenta and the mechanisms that regulate their cytolytic function has thus far largely been ignored. Fetal EVT are the most invasive cells of the placenta and directly interact with maternal decidual immune cells at this maternal-fetal interface. Besides the expression of non-polymorphic HLA-E and HLA-G molecules that are associated with immune tolerance, EVT also express highly polymorphic HLA-C molecules that can serve as targets for maternal dNK and CD8+ dT responses. HLA-C expression by EVT has a dual role as the main molecule to which immune tolerance needs to be established and as the only molecule that can present pathogen-derived peptides and provide protective immunity when EVT are infected. The focus of this review is to address the regulation of cytotoxicity of dNK and CD8+ dT, which is essential for maternal-fetal immune tolerance as well as recent evidence that both cell types can provide immunity to infections at the maternal-fetal interface. A particular emphasis is given to the role of HLA-C expressed by EVT and its capacity to elicit dNK and CD8+ dT responses. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  12. Ageratum enation virus—A Begomovirus of Weeds with the Potential to Infect Crops

    Directory of Open Access Journals (Sweden)

    Muhammad Tahir

    2015-02-01

    Full Text Available Samples of two Ageratum conyzoides, one Sonchus oleraceus and one turnip (Brassica rapa var. rapa exhibiting virus-like symptoms were collected from Pakistan and Nepal. Full-length begomovirus clones were obtained from the four plant samples and betasatellite clones from three of these. The begomovirus sequences were shown to be isolates of Ageratum enation virus (AEV with greater than 89.1% nucleotide sequence identity to the 26 AEV sequences available in the databases. The three betasatellite sequences were shown to be isolates of Ageratum yellow leaf curl betasatellite (AYLCB with greater than 90% identity to the 18 AYLCB sequences available in the databases. The AEV sequences were shown to fall into two distinct strains, for which the names Nepal (consisting of isolates from Nepal, India, and Pakistan—including the isolates identified here and India (isolates occurring only in India strains are proposed. For the clones obtained from two AEV isolates, with their AYLCB, infectivity was shown by Agrobacterium-mediated inoculation to Nicotiana benthamiana, N. tabacum, Solanum lycopersicon and A. conyzoides. N. benthamiana plants infected with AEV alone or betasatellite alone showed no symptoms. N. benthamiana plants infected with AEV with its associated betasatellite showed leaf curl symptoms. The findings show that AEV is predominantly a virus of weeds that has the capacity to infect crops. AYLCB appears to be the common partner betasatellite of AEV and is associated with diseases with a range of very different symptoms in the same plant species. The inability to satisfy Koch’s postulates with the cloned components of isolate SOL in A. conyzoides suggests that the etiology may be more complex than a single virus with a single betasatellite.

  13. Ageratum enation virus—A Begomovirus of Weeds with the Potential to Infect Crops

    Science.gov (United States)

    Tahir, Muhammad; Amin, Imran; Haider, Muhammad Saleem; Mansoor, Shahid; Briddon, Rob W.

    2015-01-01

    Samples of two Ageratum conyzoides, one Sonchus oleraceus and one turnip (Brassica rapa var. rapa) exhibiting virus-like symptoms were collected from Pakistan and Nepal. Full-length begomovirus clones were obtained from the four plant samples and betasatellite clones from three of these. The begomovirus sequences were shown to be isolates of Ageratum enation virus (AEV) with greater than 89.1% nucleotide sequence identity to the 26 AEV sequences available in the databases. The three betasatellite sequences were shown to be isolates of Ageratum yellow leaf curl betasatellite (AYLCB) with greater than 90% identity to the 18 AYLCB sequences available in the databases. The AEV sequences were shown to fall into two distinct strains, for which the names Nepal (consisting of isolates from Nepal, India, and Pakistan—including the isolates identified here) and India (isolates occurring only in India) strains are proposed. For the clones obtained from two AEV isolates, with their AYLCB, infectivity was shown by Agrobacterium-mediated inoculation to Nicotiana benthamiana, N. tabacum, Solanum lycopersicon and A. conyzoides. N. benthamiana plants infected with AEV alone or betasatellite alone showed no symptoms. N. benthamiana plants infected with AEV with its associated betasatellite showed leaf curl symptoms. The findings show that AEV is predominantly a virus of weeds that has the capacity to infect crops. AYLCB appears to be the common partner betasatellite of AEV and is associated with diseases with a range of very different symptoms in the same plant species. The inability to satisfy Koch’s postulates with the cloned components of isolate SOL in A. conyzoides suggests that the etiology may be more complex than a single virus with a single betasatellite. PMID:25674770

  14. Ageratum enation virus-a begomovirus of weeds with the potential to infect crops.

    Science.gov (United States)

    Tahir, Muhammad; Amin, Imran; Haider, Muhammad Saleem; Mansoor, Shahid; Briddon, Rob W

    2015-02-10

    Samples of two Ageratum conyzoides, one Sonchus oleraceus and one turnip (Brassica rapa var. rapa) exhibiting virus-like symptoms were collected from Pakistan and Nepal. Full-length begomovirus clones were obtained from the four plant samples and betasatellite clones from three of these. The begomovirus sequences were shown to be isolates of Ageratum enation virus (AEV) with greater than 89.1% nucleotide sequence identity to the 26 AEV sequences available in the databases. The three betasatellite sequences were shown to be isolates of Ageratum yellow leaf curl betasatellite (AYLCB) with greater than 90% identity to the 18 AYLCB sequences available in the databases. The AEV sequences were shown to fall into two distinct strains, for which the names Nepal (consisting of isolates from Nepal, India, and Pakistan-including the isolates identified here) and India (isolates occurring only in India) strains are proposed. For the clones obtained from two AEV isolates, with their AYLCB, infectivity was shown by Agrobacterium-mediated inoculation to Nicotiana benthamiana, N. tabacum, Solanum lycopersicon and A. conyzoides. N. benthamiana plants infected with AEV alone or betasatellite alone showed no symptoms. N. benthamiana plants infected with AEV with its associated betasatellite showed leaf curl symptoms. The findings show that AEV is predominantly a virus of weeds that has the capacity to infect crops. AYLCB appears to be the common partner betasatellite of AEV and is associated with diseases with a range of very different symptoms in the same plant species. The inability to satisfy Koch's postulates with the cloned components of isolate SOL in A. conyzoides suggests that the etiology may be more complex than a single virus with a single betasatellite.

  15. Vitamin D deficiency: A potential risk factor for Clostridium difficile infection

    OpenAIRE

    Youssef, Dima; Grant, William B; Peiris, Alan N

    2012-01-01

    Dima Youssef,1 William B Grant,2 Alan N Peiris3,41Department of Internal Medicine, Division of Infectious Diseases, 2Sunlight, Nutrition and Health Research Center, San Francisco, CA USA; 3Department of Medicine, Mountain Home VAMC, 4Department of Medicine, East Tennessee State University, Johnson City, Tennessee, USAIn the July 3, 2012 issue of the journal of Risk Management and Healthcare Policy, Martinez et al present a nice review on Clostridium difficile (C. difficile) infections.1 The d...

  16. Cerebrospinal Fluid Proteomics Reveals Potential Pathogenic Changes in the Brains of SIV-infected Monkeys

    OpenAIRE

    Pendyala, Gurudutt; Trauger, Sunia A.; Kalisiak, Ewa; Ellis, Ronald J.; Siuzdak, Gary; Fox, Howard S.

    2009-01-01

    The HIV-1-associated neurocognitive disorder occurs in approximately one-third of infected individuals. It has persisted in the current era of anti-retroviral therapy, and its study is complicated by the lack of biomarkers for this condition. Since the cerebrospinal fluid is the most proximal biofluid to the site of pathology, we studied the cerebrospinal fluid in a nonhuman primate model for HIV-1-associated neurocognitive disorder. Here we present a simple and efficient liquid chromatograph...

  17. Diel infection of cyanobacteria by cyanophages

    Directory of Open Access Journals (Sweden)

    Tianchi eNi

    2016-01-01

    Full Text Available Cyanobacteria exhibit biological rhythms as an adaptation to the daily light-dark (diel cycle. Light is also crucial for bacteriophages (cyanophages that infect cyanobacteria. As the first step of infection, the adsorption of some cyanophages to their host cells is light-dependent. Moreover, cyanophage replication is affected by light intensity and possibly the host cell cycle. Photosynthesis and carbon metabolism genes have been found in cyanophage genomes. With these genes, cyanophages may affect the host metabolic rhythm. Field studies suggest that cyanophage infection of cyanobacteria in aquatic environments is synchronized directly or indirectly to the light-dark cycle. These discoveries are beginning to reveal how the daily light-dark cycle shapes the interaction of cyanophages and cyanobacteria, which eventually influences matter and energy transformation in aquatic environments.

  18. Susceptibility of pregnant women to toxoplasma infection--potential benefits for newborn screening.

    LENUS (Irish Health Repository)

    Ferguson, W

    2008-08-20

    Congenital toxoplasmosis (CT) arises as a result of new acquisition of Toxoplasma infection by a susceptible woman during pregnancy. Early detection of CT through neonatal screening programmes could optimize management and improve infant outcome. This study sought to estimate the prevalence of Toxoplasma susceptibility in pregnant women. As detection of Toxoplasma antibodies in neonatal blood reflects maternal exposure history, maternal antibody seroprevalence was determined using anonymized residual blood from newborn screening cards. A total of 20,252 cards were tested in 1 year. 4,991 (24.6%) cards tested positive for Toxoplasma antibody. Results were stratified by county. Toxoplasma antibody seroprevalence rates of 25% indicated that Toxoplasma infection is common in Ireland and that up to 75% of women remain susceptible to primary infection during pregnancy. This study aimed to a) determine the seroprevalence of Toxoplasma antibody in pregnant women, and hence b) estimate the risk for acquisition of primary toxoplasmosis in pregnancy in order to support an application to fund a pilot newborn screening programme.

  19. Tourniquets and exsanguinators: a potential source of infection in the orthopedic operating theater?

    LENUS (Irish Health Repository)

    Brennan, Stephen A

    2009-04-01

    BACKGROUND AND PURPOSE: Fomites are increasingly being recognised as a source of hospital-acquired infection. We have therefore assessed tourniquets and exsanguinators for the presence of bacterial pathogens in 1 elective and 2 trauma orthopedic hospitals. MATERIAL AND METHODS: Swabs were taken prior to and after decontaminating these devices with 1 of 3 different cleaning modalities. These were then assessed for colony counts and organisms identified. RESULTS: Bacteria commonly implicated in surgical site infections such as coagulase-negative staphylococci, Staphylococcus aureus and Proteus spp. were prevalent. We also found a resistant strain of Acinetobacter and Candida. Exsanguinators were the most heavily contaminated devices, and colony counts in the trauma hospitals were up to 400% higher than in the elective hospital. Alcohol- and non-alcohol-based sterile wipes were both highly effective in decontaminating the devices. INTERPRETATION: Infectious organisms reside on the tourniquets and exsanguinators presently used in the orthopedic theater. These fomites may possibly be a source of surgical site infection. We have demonstrated a simple and effective means of decontaminating these devices between cases.

  20. Methylated Host Cell Gene Promoters and Human Papillomavirus Type 16 and 18 Predicting Cervical Lesions and Cancer.

    Directory of Open Access Journals (Sweden)

    Nina Milutin Gašperov

    Full Text Available Change in the host and/or human papillomavirus (HPV DNA methylation profile is probably one of the main factors responsible for the malignant progression of cervical lesions to cancer. To investigate those changes we studied 173 cervical samples with different grades of cervical lesion, from normal to cervical cancer. The methylation status of nine cellular gene promoters, CCNA1, CDH1, C13ORF18, DAPK1, HIC1, RARβ2, hTERT1, hTERT2 and TWIST1, was investigated by Methylation Specific Polymerase Chain Reaction (MSP. The methylation of HPV18 L1-gene was also investigated by MSP, while the methylated cytosines within four regions, L1, 5'LCR, enhancer, and promoter of the HPV16 genome covering 19 CpG sites were evaluated by bisulfite sequencing. Statistically significant methylation biomarkers distinguishing between cervical precursor lesions from normal cervix were primarily C13ORF18 and secondly CCNA1, and those distinguishing cervical cancer from normal or cervical precursor lesions were CCNA1, C13ORF18, hTERT1, hTERT2 and TWIST1. In addition, the methylation analysis of individual CpG sites of the HPV16 genome in different sample groups, notably the 7455 and 7694 sites, proved to be more important than the overall methylation frequency. The majority of HPV18 positive samples contained both methylated and unmethylated L1 gene, and samples with L1-gene methylated forms alone had better prognosis when correlated with the host cell gene promoters' methylation profiles. In conclusion, both cellular and viral methylation biomarkers should be used for monitoring cervical lesion progression to prevent invasive cervical cancer.

  1. Interaction of MYC with host cell factor-1 is mediated by the evolutionarily conserved Myc box IV motif.

    Science.gov (United States)

    Thomas, L R; Foshage, A M; Weissmiller, A M; Popay, T M; Grieb, B C; Qualls, S J; Ng, V; Carboneau, B; Lorey, S; Eischen, C M; Tansey, W P

    2016-07-07

    The MYC family of oncogenes encodes a set of three related transcription factors that are overexpressed in many human tumors and contribute to the cancer-related deaths of more than 70,000 Americans every year. MYC proteins drive tumorigenesis by interacting with co-factors that enable them to regulate the expression of thousands of genes linked to cell growth, proliferation, metabolism and genome stability. One effective way to identify critical co-factors required for MYC function has been to focus on sequence motifs within MYC that are conserved throughout evolution, on the assumption that their conservation is driven by protein-protein interactions that are vital for MYC activity. In addition to their DNA-binding domains, MYC proteins carry five regions of high sequence conservation known as Myc boxes (Mb). To date, four of the Mb motifs (MbI, MbII, MbIIIa and MbIIIb) have had a molecular function assigned to them, but the precise role of the remaining Mb, MbIV, and the reason for its preservation in vertebrate Myc proteins, is unknown. Here, we show that MbIV is required for the association of MYC with the abundant transcriptional coregulator host cell factor-1 (HCF-1). We show that the invariant core of MbIV resembles the tetrapeptide HCF-binding motif (HBM) found in many HCF-interaction partners, and demonstrate that MYC interacts with HCF-1 in a manner indistinguishable from the prototypical HBM-containing protein VP16. Finally, we show that rationalized point mutations in MYC that disrupt interaction with HCF-1 attenuate the ability of MYC to drive tumorigenesis in mice. Together, these data expose a molecular function for MbIV and indicate that HCF-1 is an important co-factor for MYC.

  2. Importance of CD200 expression by tumor or host cells to regulation of immunotherapy in a mouse breast cancer model.

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    Anna Curry

    Full Text Available Cell-surface CD200 expression by mouse EMT6 breast tumor cells increased primary tumor growth and metastasis to the draining lymph nodes (DLN in normal (WT BALB/c female recipients, while lack of CD200R1 expression in a CD200R1-/- host negated this effect. Silencing CD200 expression in EMT6siCD200 tumor cells also reduced their ability to grow and metastasize in WT animals. The cellular mechanisms responsible for these effects have not been studied in detail. We report characterization of tumor infiltrating (TILs and draining lymph node (DLN cells in WT and CD200-/- BALB/c mice, receiving WT tumor cells, or EMT6 lacking CD200 expression (EMT6siCD200 cells. Our data show an important correlation with augmented CD8+ cytotoxic T cells and resistance to tumor growth in mice lacking exposure (on either host cells or tumor to the immunoregulatory molecule CD200. Confirmation of the importance of such CD8+ cells came from monitoring tumor growth and characterization of the TILs and DLN cells in WT mice challenged with EMT6 and EMT6siCD200 tumors and treated with CD8 and CD4 depleting antibodies. Finally, we have assessed the mechanisms(s whereby addition of metformin as an augmenting chemotherapeutic agent in CD200-/- animals given EMT6 tumors and treated with a previously established immunotherapy regime can increase host resistance. Our data support the hypothesis that increased autophagy in the presence of metformin increases CD8+ responses and tumor resistance, an effect attenuated by the autophagy inhibitor verteporfin.

  3. New Parameters to Quantitatively Express the Invasiveness of Bacterial Strains from Implant-Related Orthopaedic Infections into Osteoblast Cells

    Directory of Open Access Journals (Sweden)

    Davide Campoccia

    2018-04-01

    Full Text Available Complete eradication of bacterial infections is often a challenging task, especially in presence of prosthetic devices. Invasion of non-phagocytic host cells appears to be a critical mechanism of microbial persistence in host tissues. Hidden within host cells, bacteria elude host defences and antibiotic treatments that are intracellularly inactive. The intracellular invasiveness of bacteria is generally measured by conventional gentamicin protection assays. The efficiency of invasion, however, markedly differs across bacterial species and adjustments to the titre of the microbial inocula used in the assays are often needed to enumerate intracellular bacteria. Such changes affect the standardisation of the method and hamper a direct comparison of bacteria on a same scale. This study aims at investigating the precise relation between inoculum, in terms of multiplicity of infection (MOI, and internalised bacteria. The investigation included nine Staphylococcus aureus, seven Staphylococcus epidermidis, five Staphylococcus lugdunensis and two Enterococcus faecalis clinical strains, which are co-cultured with MG63 human osteoblasts. Unprecedented insights are offered on the relations existing between MOI, number of internalised bacteria and per cent of internalised bacteria. New parameters are identified that are of potential use for qualifying the efficiency of internalization and compare the behaviour of bacterial strains.

  4. New Parameters to Quantitatively Express the Invasiveness of Bacterial Strains from Implant-Related Orthopaedic Infections into Osteoblast Cells.

    Science.gov (United States)

    Campoccia, Davide; Montanaro, Lucio; Ravaioli, Stefano; Cangini, Ilaria; Testoni, Francesca; Visai, Livia; Arciola, Carla Renata

    2018-04-03

    Complete eradication of bacterial infections is often a challenging task, especially in presence of prosthetic devices. Invasion of non-phagocytic host cells appears to be a critical mechanism of microbial persistence in host tissues. Hidden within host cells, bacteria elude host defences and antibiotic treatments that are intracellularly inactive. The intracellular invasiveness of bacteria is generally measured by conventional gentamicin protection assays. The efficiency of invasion, however, markedly differs across bacterial species and adjustments to the titre of the microbial inocula used in the assays are often needed to enumerate intracellular bacteria. Such changes affect the standardisation of the method and hamper a direct comparison of bacteria on a same scale. This study aims at investigating the precise relation between inoculum, in terms of multiplicity of infection (MOI), and internalised bacteria. The investigation included nine Staphylococcus aureus , seven Staphylococcus epidermidis , five Staphylococcus lugdunensis and two Enterococcus faecalis clinical strains, which are co-cultured with MG63 human osteoblasts. Unprecedented insights are offered on the relations existing between MOI, number of internalised bacteria and per cent of internalised bacteria. New parameters are identified that are of potential use for qualifying the efficiency of internalization and compare the behaviour of bacterial strains.

  5. Ichthyophonus-infected walleye pollock Theragra chalcogramma (Pallas) in the eastern Bering Sea: a potential reservoir of infections in the North Pacific.

    Science.gov (United States)

    White, V C; Morado, J F; Friedman, C S

    2014-07-01

    In 2003, the Alaska walleye pollock industry reported product quality issues attributed to an unspecified parasite in fish muscle. Using molecular and histological methods, we identified the parasite in Bering Sea pollock as Ichthyophonus. Infected pollock were identified throughout the study area, and prevalence was greater in adults than in juveniles. This study not only provides the first documented report of Ichthyophonus in any fish species captured in the Bering Sea, but also reveals that the parasite has been present in this region for nearly 20 years and is not a recent introduction. Sequence analysis of 18S rDNA from Ichthyophonus in pollock revealed that consensus sequences were identical to published parasite sequences from Pacific herring and Yukon River Chinook salmon. Results from this study suggest potential for Ichthyophonus exposures from infected pollock via two trophic pathways; feeding on whole fish as prey and scavenging on industry-discharged offal. Considering the notable Ichthyophonus levels in pollock, the low host specificity of the parasite and the role of this host as a central prey item in the Bering Sea, pollock likely serve as a key Ichthyophonus reservoir for other susceptible hosts in the North Pacific. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  6. Children’s Caregivers and Public Playgrounds: Potential Reservoirs of Infection of Hand-foot-and-mouth Disease

    Science.gov (United States)

    Li, Pengyuan; Li, Tao; Gu, Qiuyun; Chen, Xiaomin; Li, Jiahui; Chen, Xiashi; Chen, Yan; Zhang, Danwei; Gao, Rong; He, Zhenjian; Zhu, Xun; Zhang, Wangjian; Hao, Yuantao; Zhang, Dingmei

    2016-11-01

    Hand-foot-and-mouth disease (HFMD) is a common infectious disease, which has led to millions of clinical cases and hundreds of deaths every year in China. This study aimed to exploring the effects on HFMD transmission of children’s caregivers and public area, as well as trying to locate the potential reservoirs of infections in primary cases. Total children’s 257 samples (98 children’s caregivers and 159 environmental samples) were tested for the presence of universal enterovirus, enterovirus 71, coxsackie virus A6 and A16 by real-time fluorescence quantitative polymerase chain reaction (qPCR). 5.84% (15/257, 95% confidence interval [CI]: 2.98%, 8.70%) of total samples had positive results of enterovirus. The enterovirus positive rates of children’s caregiver samples and environmental samples were respectively 7.14% (7/98, 95% CI: 2.04%, 12.24%), and 5.03% (8/159, 95% CI: 1.63%, 8.43%); 7.61% (7/92, 95% CI: 2.21%, 13.01%) of wiping samples from playgrounds and 1.49% (1/67, 95% CI: 0, 7.00%) of air samples in indoor market places had positive result of enterovirus. High positive rates of enterovirus in children’s caregivers and from playgrounds indicated that they would be potential reservoirs of HFMD infection, as children might be infected via contacting with asymptomatic-infected individuals or exposure of contaminated surface of public facilities.

  7. Bacteria Facilitate Enteric Virus Co-infection of Mammalian Cells and Promote Genetic Recombination.

    Science.gov (United States)

    Erickson, Andrea K; Jesudhasan, Palmy R; Mayer, Melinda J; Narbad, Arjan; Winter, Sebastian E; Pfeiffer, Julie K

    2018-01-10

    RNA viruses exist in genetically diverse populations due to high levels of mutations, many of which reduce viral fitness. Interestingly, intestinal bacteria can promote infection of several mammalian enteric RNA viruses, but the mechanisms and consequences are unclear. We screened a panel of 41 bacterial strains as a platform to determine how different bacteria impact infection of poliovirus, a model enteric virus. Most bacterial strains, including those extracted from cecal contents of mice, bound poliovirus, with each bacterium binding multiple virions. Certain bacterial strains increased viral co-infection of mammalian cells even at a low virus-to-host cell ratio. Bacteria-mediated viral co-infection correlated with bacterial adherence to cells. Importantly, bacterial strains that induced viral co-infection facilitated genetic recombination between two different viruses, thereby removing deleterious mutations and restoring viral fitness. Thus, bacteria-virus interactions may increase viral fitness through viral recombination at initial sites of infection, potentially limiting abortive infections. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. The degree of microbiome complexity influences the epithelial response to infection

    Directory of Open Access Journals (Sweden)

    Baker Henry V

    2009-08-01

    Full Text Available Abstract Background The human microflora is known to be extremely complex, yet most pathogenesis research is conducted in mono-species models of infection. Consequently, it remains unclear whether the level of complexity of a host's indigenous flora can affect the virulence potential of pathogenic species. Furthermore, it remains unclear whether the colonization by commensal species affects a host cell's response to pathogenic species beyond the direct physical saturation of surface receptors, the sequestration of nutrients, the modulation of the physico-chemical environment in the oral cavity, or the production of bacteriocins. Using oral epithelial cells as a model, we hypothesized that the virulence of pathogenic species may vary depending on the complexity of the flora that interacts with host cells. Results This is the first report that determines the global epithelial transcriptional response to co-culture with defined complex microbiota. In our model, human immortalized gingival keratinocytes (HIGK were infected with mono- and mixed cultures of commensal and pathogenic species. The global transcriptional response of infected cells was validated and confirmed phenotypically. In our model, commensal species were able to modulate the expression of host genes with a broad diversity of physiological functions and antagonize the effect of pathogenic species at the cellular level. Unexpectedly, the inhibitory effect of commensal species was not correlated with its ability to inhibit adhesion or invasion by pathogenic species. Conclusion Studying the global transcriptome of epithelial cells to single and complex microbial challenges offers clues towards a better understanding of how bacteria-bacteria interactions and bacteria-host interactions impact the overall host response. This work provides evidence that the degree of complexity of a mixed microbiota does influence the transcriptional response to infection of host epithelial cells, and

  9. Toxoplasma gondii Infection Is Associated with Mitochondrial Dysfunction in-Vitro

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    Genevieve Syn

    2017-12-01

    Full Text Available Upon invasion of host cells, the ubiquitous pathogen Toxoplasma gondii manipulates several host processes, including re-organization of host organelles, to create a replicative niche. Host mitochondrial association to T. gondii parasitophorous vacuoles is rapid and has roles in modulating host immune responses. Here gene expression profiling of T. gondii infected cells reveals enrichment of genes involved in oxidative phosphorylation (OXPHOS and mitochondrial dysfunction 6 h post-infection. We identified 11 hub genes (HIF-1α, CASP8, FN1, POU5F1, CD44, ISG15, HNRNPA1, MDM2, RPL35, VHL, and NUPR1 and 10 predicted upstream regulators, including 4 endogenous regulators RICTOR, KDM5A, RB1, and D-glucose. We characterized a number of mitochondrial parameters in T. gondii infected human foreskin fibroblast cells over a 36 h time-course. In addition to the usual rapid recruitment and apparent enlargement of mitochondria around the parasitophorous vacuole we observed fragmented host mitochondria in infected cells, not linked to cellular apoptosis, from 24 h post-infection. An increase in mitochondrial superoxide levels in T. gondii infected cells was observed that required active parasite invasion and peaked at 30 h post-infection. Measurement of OXPHOS proteins showed decreased expression of Complex IV in infected cells at 24 h post-infection, followed by decreased expression of Complexes I and II at 36 h post-infection. No change occurred in Complex V. No difference in host mitochondrial membrane potential between infected and mock-infected cells was observed at any time. Our results show perturbation of host mitochondrial function following T. gondii infection that likely impacts on pathogenesis of disease.

  10. Export of a Toxoplasma gondii rhoptry neck protein complex at the host cell membrane to form the moving junction during invasion.

    Directory of Open Access Journals (Sweden)

    Sébastien Besteiro

    2009-02-01

    Full Text Available One of the most conserved features of the invasion process in Apicomplexa parasites is the formation of a moving junction (MJ between the apex of the parasite and the host cell membrane that moves along the parasite and serves as support to propel it inside the host cell. The MJ was, up to a recent period, completely unknown at the molecular level. Recently, proteins originated from two distinct post-Golgi specialised secretory organelles, the micronemes (for AMA1 and the neck of the rhoptries (for RON2/RON4/RON5 proteins, have been shown to form a complex. AMA1 and RON4 in particular, have been localised to the MJ during invasion. Using biochemical approaches, we have identified RON8 as an additional member of the complex. We also demonstrated that all RON proteins are present at the MJ during invasion. Using metabolic labelling and immunoprecipitation, we showed that RON2 and AMA1 were able to interact in the absence of the other members. We also discovered that all MJ proteins are subjected to proteolytic maturation during trafficking to their respective organelles and that they could associate as non-mature forms in vitro. Finally, whereas AMA1 has previously been shown to be inserted into the parasite membrane upon secretion, we demonstrated, using differential permeabilization and loading of RON-specific antibodies into the host cell, that the RON complex is targeted to the host cell membrane, where RON4/5/8 remain associated with the cytoplasmic face. Globally, these results point toward a model of MJ organization where the parasite would be secreting and inserting interacting components on either side of the MJ, both at the host and at its own plasma membranes.

  11. Ebola Virus Infection Modelling and Identifiability Problems

    Directory of Open Access Journals (Sweden)

    Van-Kinh eNguyen

    2015-04-01

    Full Text Available The recent outbreaks of Ebola virus (EBOV infections have underlined the impact of the virus as a major threat for human health. Due to the high biosafety classification of EBOV (level 4, basic research is very limited. Therefore, the development of new avenues of thinking to advance quantitative comprehension of the virus and its interaction with the host cells is urgently neededto tackle this lethal disease. Mathematical modelling of the EBOV dynamics can be instrumental to interpret Ebola infection kinetics on quantitative grounds. To the best of our knowledge, a mathematical modelling approach to unravel the interaction between EBOV and the host cells isstill missing. In this paper, a mathematical model based on differential equations is used to represent the basic interactions between EBOV and wild-type Vero cells in vitro. Parameter sets that represent infectivity of pathogens are estimated for EBOV infection and compared with influenza virus infection kinetics. The average infecting time of wild-type Vero cells in EBOV is slower than in influenza infection. Simulation results suggest that the slow infecting time of EBOV could be compensated by its efficient replication. This study reveals several identifiability problems and what kind of experiments are necessary to advance the quantification of EBOV infection. A first mathematical approach of EBOV dynamics and the estimation of standard parametersin viral infections kinetics is the key contribution of this work, paving the way for future modelling work on EBOV infection.

  12. Innate Immunity to Leishmania Infection: Within Phagocytes

    Directory of Open Access Journals (Sweden)

    Marcela Freitas Lopes

    2014-01-01

    Full Text Available Infection by Leishmania takes place in the context of inflammation and tissue repair. Besides tissue resident macrophages, inflammatory macrophages and neutrophils are recruited to the infection site and serve both as host cells and as effectors against infection. Recent studies suggest additional important roles for monocytes and dendritic cells. This paper addresses recent experimental findings regarding the regulation of Leishmania major infection by these major phagocyte populations. In addition, the role of IL-4 on dendritic cells and monocytes is discussed.

  13. An evaluation of the infection control potential of a UV clinical podiatry unit.

    Science.gov (United States)

    Humphreys, Paul N; Davies, Chris S; Rout, Simon

    2014-02-28

    Infection control is a key issue in podiatry as it is in all forms of clinical practice. Airborne contamination may be particularly important in podiatry due to the generation of particulates during treatment. Consequently, technologies that prevent contamination in podiatry settings may have a useful role. The aims of this investigation were twofold, firstly to determine the ability of a UV cabinet to protect instruments from airborne contamination and secondly to determine its ability to remove microbes from contaminated surfaces and instruments. A UV instrument cabinet was installed in a University podiatry suite. Impact samplers and standard microbiological techniques were used to determine the nature and extent of microbial airborne contamination. Sterile filters were used to determine the ability of the UV cabinet to protect exposed surfaces. Artificially contaminated instruments were used to determine the ability of the cabinet to remove microbial contamination. Airborne bacterial contamination was dominated by Gram positive cocci including Staphylococcus aureus. Airborne fungal levels were much lower than those observed for bacteria. The UV cabinet significantly reduced (p podiatry settings due to the presence of S. aureus. The use of a UV instrument cabinet can reduce the risk of contamination by airborne microbes. The UV cabinet tested was unable to decontaminate instruments and as such could pose an infection risk if misused.

  14. The potential danger of eating wild lettuce: a brief review of human rat lungworm infection.

    Science.gov (United States)

    Ewers, Evan C; Anisowicz, Sarah K

    2014-11-01

    Angiostrongylus cantonensis, the causative agent of human rat lungworm disease, is the most common cause of eosinophilic meningitis worldwide and is endemic throughout Asia Pacific. It is acquired through the consumption of infected freshwater mollusks or contaminated produce. Human angiostrongyliasis is usually a self-limited disease presenting with headache and various neurologic sequelae varying from cranial nerve palsies to radiculitis and/or paresthesias. Fatal cases are rare, and manifest as fulminant meningomyeloencephalitis. The diagnosis is made through the use of clinical history, exam, and laboratory data including peripheral blood counts, cerebrospinal fluid (CSF) examinations, and serologic or molecular diagnostic techniques. Medical therapy is largely focused on symptomatic relief, and includes analgesics, lumbar puncture, and corticosteroids. In resource-limited settings, prevention is key, and the use of analgesics can provide symptomatic relief after infection. Efforts to increase disease awareness have been made in endemic areas, as evidenced by the recent Rat Lungworm Disease Scientific Workshop which was held in Honolulu in 2011. The proceedings of the workshop were published in a supplement to this journal (Hawaii J Med Public Health. Jun 2013;72(6):Supp 2). However, wilderness medicine and travel medicine specialists must also be aware of the disease, how it is contracted, its presentation, and treatment options should they encounter a patient who is in or has returned from an endemic area. This brief review highlights eosinophilic meningitis caused by A. cantonensis, including an example case, an overview of its clinical presentation, treatment options, and prevention.

  15. Antimicrobial Properties of Mesenchymal Stem Cells: Therapeutic Potential for Cystic Fibrosis Infection, and Treatment

    Directory of Open Access Journals (Sweden)

    Morgan T. Sutton

    2016-01-01

    Full Text Available Cystic fibrosis (CF is a genetic disease in which the battle between pulmonary infection and inflammation becomes the major cause of morbidity and mortality. We have previously shown that human MSCs (hMSCs decrease inflammation and infection in the in vivo murine model of CF. The studies in this paper focus on the specificity of the hMSC antimicrobial effectiveness using Pseudomonas aeruginosa (gram negative bacteria and Staphylococcus aureus (gram positive bacteria. Our studies show that hMSCs secrete bioactive molecules which are antimicrobial in vitro against Pseudomonas aeruginosa, Staphylococcus aureus, and Streptococcus pneumonia, impacting the rate of bacterial growth and transition into colony forming units regardless of the pathogen. Further, we show that the hMSCs have the capacity to enhance antibiotic sensitivity, improving the capacity to kill bacteria. We present data which suggests that the antimicrobial effectiveness is associated with the capacity to slow bacterial growth and the ability of the hMSCs to secrete the antimicrobial peptide LL-37. Lastly, our studies demonstrate that the tissue origin of the hMSCs (bone marrow or adipose tissue derived, the presence of functional cystic fibrosis transmembrane conductance regulator (CFTR: human, Cftr: mouse activity, and response to effector cytokines can impact both hMSC phenotype and antimicrobial potency and efficacy. These studies demonstrate, the unique capacity of the hMSCs to manage different pathogens and the significance of their phenotype in both the antimicrobial and antibiotic enhancing activities.

  16. Effects of minimal exposures to atmospheric pressure plasma on the activity of Salmonella Typhimurium: Deactivation of bacterial motility and suppression of host-cell invasion.

    Science.gov (United States)

    Park, Jin-Sung; Kim, Kijung; Han, Je-Hyun; Gweon, Bomi; Ko, Ung Hyun; Yoo, Suk Jae; Choe, Wonho; Shin, Jennifer H

    2016-09-01

    Atmospheric pressure plasma (APP) has been shown effective in sterilization by reducing the number of viable microbes during surface cleaning, food processing, or human tissue treatment. For safe conduct, the majority of previous research focused on complete abolition of microbes, which may require severe treatments. Our aim is to investigate the minimal treatment conditions necessary for effective inactivation of bacteria in such a manner that the APP treated bacteria would not be able to harm the host cells. For this, we ought to identify the objective criteria to make the bacteria dysfunctional. We choose the motile properties and the host-cell invasion capability as two measures to quantify the pathogenic state of bacteria. In this paper, we investigated how the APP treatment in a minimal dosage affects the activity of Salmonella Typhimurium. At 100 W and 15 kHz for 20 s, the APP treatment effectively suppressed active "run and tumble" type motility and induced formation of abnormally long structures. With 20 s exposure, the bacterial cells failed to cause pyroptosis in the host cells with >90% survival after 12 h of co-incubation. Our results suggest novel measures to evaluate the functional pathogenic state for identifying safe APP treatment conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Sampling the light-organ microenvironment of Euprymna scolopes: description of a population of host cells in association with the bacterial symbiont Vibrio fischeri.

    Science.gov (United States)

    Nyholm, S V; McFall-Ngai, M J

    1998-10-01

    The symbiosis between the squid Euprymna scolopes and the luminous bacterium Vibrio fischeri has a pronounced diel rhythm, one component of which is the venting of the contents of the light organ into the surrounding seawater each day at dawn. In this study, we explored the use of this behavior to sample the microenvironment of the light-organ crypts. Intact crypt contents, which emerge from the lateral pores of the organ as a thick paste-like exudate, were collected from anesthetized host animals that had been exposed to a light cue. Microscopy revealed that the expelled material is composed of a conspicuous population of host cells in association with the bacterial symbionts, all of which are embedded in a dense acellular matrix that strongly resembles the bacteria-based biofilms described in other systems. Assays of the viability of expelled crypt cells revealed no dead bacterial symbionts and a mixture of live and dead host cells. Analyses of the ultrastructure, biochemistry, and phagocytic activity of a subset of the host cell population suggested that some of these cells are macrophage-like molluscan hemocytes.

  18. Directed evolution and targeted mutagenesis to murinize Listeria monocytogenes internalin A for enhanced infectivity in the murine oral infection model.

    LENUS (Irish Health Repository)

    Monk, Ian R

    2010-01-01

    Internalin A (InlA) is a critical virulence factor which mediates the initiation of Listeria monocytogenes infection by the oral route in permissive hosts. The interaction of InlA with the host cell ligand E-cadherin efficiently stimulates L. monocytogenes entry into human enterocytes, but has only a limited interaction with murine cells.

  19. Potential place of 99mTc-DMSA scintigraphy in the management of children with urinary tract infection

    International Nuclear Information System (INIS)

    Piepsz, A.

    2010-01-01

    Cortical 99m Tc DMSA scintigraphy is accepted as a highly sensitive technique for the detection of regional lesions. It reflects accurately the histological changes and the interobserver reproducibility in reporting is high. Potential technical pitfalls should be recognized, such as the normal variants and the difficulty to differentiate acute lesions from permanent ones, or acquired lesions from congenital ones. Although DMSA scintgraphy seems to play a minor role in the traditional approach of urinary tract infection, recent studies suggest that this examination might influence the treatment of the acute phase, the indication of chemoprophylaxis and of micturating cystography, as well as the duration of follow-up. (orig.)

  20. Identification of bovine leukemia virus tax function associated with host cell transcription, signaling, stress response and immune response pathway by microarray-based gene expression analysis

    Directory of Open Access Journals (Sweden)

    Arainga Mariluz

    2012-03-01

    Full Text Available Abstract Background Bovine leukemia virus (BLV is associated with enzootic bovine leukosis and is closely related to human T-cell leukemia virus type I. The Tax protein of BLV is a transcriptional activator of viral replication and a key contributor to oncogenic potential. We previously identified interesting mutant forms of Tax with elevated (TaxD247G or reduced (TaxS240P transactivation effects on BLV replication and propagation. However, the effects of these mutations on functions other than transcriptional activation are unknown. In this study, to identify genes that play a role in the cascade of signal events regulated by wild-type and mutant Tax proteins, we used a large-scale host cell gene-profiling approach. Results Using a microarray containing approximately 18,400 human mRNA transcripts, we found several alterations after the expression of Tax proteins in genes involved in many cellular functions such as transcription, signal transduction, cell growth, apoptosis, stress response, and immune response, indicating that Tax protein has multiple biological effects on various cellular environments. We also found that TaxD247G strongly regulated more genes involved in transcription, signal transduction, and cell growth functions, contrary to TaxS240P, which regulated fewer genes. In addition, the expression of genes related to stress response significantly increased in the presence of TaxS240P as compared to wild-type Tax and TaxD247G. By contrast, the largest group of downregulated genes was related to immune response, and the majority of these genes belonged to the interferon family. However, no significant difference in the expression level of downregulated genes was observed among the Tax proteins. Finally, the expression of important cellular factors obtained from the human microarray results were validated at the RNA and protein levels by real-time quantitative reverse transcription-polymerase chain reaction and western blotting

  1. Inhibition of Wnt Signaling Pathways Impairs Chlamydia trachomatis Infection in Endometrial Epithelial Cells.

    Science.gov (United States)

    Kintner, Jennifer; Moore, Cheryl G; Whittimore, Judy D; Butler, Megan; Hall, Jennifer V

    2017-01-01

    Chlamydia trachomatis infections represent the predominant cause of bacterial sexually transmitted infections. As an obligate intracellular bacterium, C. trachomatis is dependent on the host cell for survival, propagation, and transmission. Thus, factors that affect the host cell, including nutrition, cell cycle, and environmental signals, have the potential to impact chlamydial development. Previous studies have demonstrated that activation of Wnt/β-catenin signaling benefits C. trachomatis infections in fallopian tube epithelia. In cervical epithelial cells chlamydiae sequester β-catenin within the inclusion. These data indicate that chlamydiae interact with the Wnt signaling pathway in both the upper and lower female genital tract (FGT). However, hormonal activation of canonical and non-canonical Wnt signaling pathways is an essential component of cyclic remodeling in another prominent area of the FGT, the endometrium. Given this information, we hypothesized that Wnt signaling would impact chlamydial infection in endometrial epithelial cells. To investigate this hypothesis, we analyzed the effect of Wnt inhibition on chlamydial inclusion development and elementary body (EB) production in two endometrial cell lines, Ishikawa (IK) and Hec-1B, in nonpolarized cell culture and in a polarized endometrial epithelial (IK)/stromal (SHT-290) cell co-culture model. Inhibition of Wnt by the small molecule inhibitor (IWP2) significantly decreased inclusion size in IK and IK/SHT-290 cultures ( p Wnt inhibition caused chlamydiae to become aberrant in morphology. EB formation was also impaired in IK, Hec-1B and IK/SHT-290 cultures regardless of whether Wnt inhibition occurred throughout, in the middle (24 hpi) or late (36 hpi) during the development cycle. Overall, these data lead us to conclude that Wnt signaling in the endometrium is a key host pathway for the proper development of C. trachomatis .

  2. Preliminary study of Malaysian fruit bats species diversity in Lenggong Livestock Breeding Center, Perak: Potential risk of spill over infection

    Directory of Open Access Journals (Sweden)

    Muhammed Mikail

    2017-11-01

    Full Text Available Aim: Farms that are neighboring wildlife sanctuaries are at risk of spillover infection from wildlife, and the objective of this research is to examine the species diversity of Malaysian fruit bats in livestock farm in determining the possible risk of spill over infection to livestock. Materials and Methods: Fifty individual fruit bats were captured using six mists net, from May to July 2017. The nets were set at dusk (1830 h as bats emerge for foraging and monitored at every 30-min intervals throughout the night until dawn when they returned to the roost. The nets were closed for the day until next night, and captured bats were identified to species levels. Results: All the captured bats were mega chiropterans, and Cynopterus brachyotis was the highest captured species, representing 40% of the total capture. Shannon-Weiner index is 2.80, and Simpson index is 0.2. Our result suggests that there is a degree of species dominance with low diversity in Lenggong Livestock Breeding Center. Conclusion: We concluded that fruit bats are indeed, encroaching livestock areas and the species identified could be a potential source of infection to susceptible livestock. Hence, an active surveillance should be embarked on farms that border wildlife sanctuaries.

  3. Identifying Potential Areas of Human Zika Infection in the City of Los Angeles, California by Use of Remote Sensing Imagery

    Science.gov (United States)

    Lee, J.

    2017-12-01

    As of April 2017, California is the third most prevalent state on the United States for Zika Infection and Southern California has an ever growing population of Aedes mosquitos. Zika is a disease which poses a significant risk to humans and other mammals due to its effects on pregnancy. This emerging disease is highly contagious due to its spread of infection primarily by Aedes aegypti mosquitos. Aedes mosquitos are able to breed in small rain collecting containers which allow the species to persevere in urban and semi urban environments. We hope to identify potential areas with risk of human infection within Los Angeles and its surrounding areas. This study integrates remote sensing, GIS, statistical, and environmental techniques to study favorable habitats for this particular species of mosquitos and their larvae. The study of the geographic and landscape factors which promote the larvae development allow for the disease spread to be analyzed and modeled. There are several goals in the development of this study. These include the coordination of statistical data with local epidemiology departments, identify workflows to improve efficiency, create models which can be utilized for disease prevention, and identify geographic risk factors for the spread of Zika.

  4. IMPY, a potential {beta}-amyloid imaging probe for detection of prion deposits in scrapie-infected mice

    Energy Technology Data Exchange (ETDEWEB)

    Song, P.-J. [INSERM, U619, F-37000 Tours (France); Universite Francois-Rabelais, F-37000 Tours (France); IFR135, F-37000 Tours (France); Bernard, Serge [IFR135, F-37000 Tours (France); INRA, UR1282, IASP, 37380 Nouzilly (France)], E-mail: bernard@tours.inra.fr; Sarradin, Pierre [INRA, UR1282, IASP, 37380 Nouzilly (France); Vergote, Jackie [INSERM, U619, F-37000 Tours (France); Universite Francois-Rabelais, F-37000 Tours (France); IFR135, F-37000 Tours (France); Barc, Celine [INRA, UR1282, IASP, 37380 Nouzilly (France); Chalon, Sylvie [INSERM, U619, F-37000 Tours (France); Universite Francois-Rabelais, F-37000 Tours (France); IFR135, F-37000 Tours (France); Kung, M.-P.; Kung, Hank F. [Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Department of Pharmacology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Guilloteau, Denis [INSERM, U619, F-37000 Tours (France); Universite Francois-Rabelais, F-37000 Tours (France); IFR135, F-37000 Tours (France)

    2008-02-15

    Introduction: A potential single-photon emission computed tomography imaging agent for labeling of A{beta} plaques of Alzheimer's disease, IMPY (2-(4'-dimethylaminophenyl)-6-iodo-imidazo[1,2-a]pyridine), would be effective in detection of prion amyloid deposits in transmissible spongiform encephalopathies (TSEs). Methods: In vitro autoradiographic studies were carried out with [{sup 125}I]IMPY on brain sections from scrapie-infected mice and age-matched controls. Competition study was performed to evaluate the prion deposit binding specificity with nonradioactive IMPY. Results: Binding of [{sup 125}I]IMPY was observed in infected brain sections, while on age-matched control brain sections, there was no or very low labeling. Prion deposit binding was confirmed by histoblots with prion protein-specific monoclonal antibody 2D6. In the presence of nonradioactive IMPY, the binding of [{sup 125}I]IMPY was significantly inhibited in all regions studied. Conclusions: These findings indicate that IMPY can detect the prion deposits in vitro in scrapie-infected mice. Labeled with {sup 123}I, this ligand may be useful to quantitate prion deposit burdens in TSEs by in vivo imaging.

  5. IMPY, a potential β-amyloid imaging probe for detection of prion deposits in scrapie-infected mice

    International Nuclear Information System (INIS)

    Song, P.-J.; Bernard, Serge; Sarradin, Pierre; Vergote, Jackie; Barc, Celine; Chalon, Sylvie; Kung, M.-P.; Kung, Hank F.; Guilloteau, Denis

    2008-01-01

    Introduction: A potential single-photon emission computed tomography imaging agent for labeling of Aβ plaques of Alzheimer's disease, IMPY (2-(4'-dimethylaminophenyl)-6-iodo-imidazo[1,2-a]pyridine), would be effective in detection of prion amyloid deposits in transmissible spongiform encephalopathies (TSEs). Methods: In vitro autoradiographic studies were carried out with [ 125 I]IMPY on brain sections from scrapie-infected mice and age-matched controls. Competition study was performed to evaluate the prion deposit binding specificity with nonradioactive IMPY. Results: Binding of [ 125 I]IMPY was observed in infected brain sections, while on age-matched control brain sections, there was no or very low labeling. Prion deposit binding was confirmed by histoblots with prion protein-specific monoclonal antibody 2D6. In the presence of nonradioactive IMPY, the binding of [ 125 I]IMPY was significantly inhibited in all regions studied. Conclusions: These findings indicate that IMPY can detect the prion deposits in vitro in scrapie-infected mice. Labeled with 123 I, this ligand may be useful to quantitate prion deposit burdens in TSEs by in vivo imaging

  6. Expression and potential roles of IL-33/ST2 in the immune regulation during Clonorchis sinensis infection.

    Science.gov (United States)

    Yu, Qian; Li, Xiang-Yang; Cheng, Xiao-Dan; Shen, Li-Ping; Fang, Fan; Zhang, Bo; Hua, Hui; Yan, Chao; Tang, Ren-Xian; Zheng, Kui-Yang

    2016-06-01

    During clonorchiasis, immune responses of hosts are responsible for the removal of the worms and also are involved in the progress of the pathological damage caused by Clonorchis sinensis. Interleukin-33 (IL-33), a recently described cytokine signaling through the ST2 receptor, has emerged as a potent inducer to bile duct proliferation and fibrosis; however, little is known of this signaling in the pathogen-caused periductal inflammation and fibrosis. In the present study, using immunohistochemistry, real-time PCR, enzyme-linked immunosorbent assay (ELISA), and flow cytometry, we studied the expression of IL-33/ST2 during C. sinensis infection, as well as their potential roles in C. sinensis-induced host immune responses. The results showed that a higher level of IL-33 was detected in the sera of patients of clonorchiasis (n = 45), compared with in those of healthy donors (n = 16). Similarly, in FVB mice experimentally infected with C. sinensis, a higher level of IL-33 was detected at latent stage both in the serum and in the liver, as well as the up-regulated expression of ST2 receptor on the inflammatory cells, especially on CD4(+) T cells in the liver of infected mice. Our results, for the first time, indicated that the increased IL-33/ST2 may be involved in the regulation of immunopathology induced by C. sinensis.

  7. Clomiphene and Its Isomers Block Ebola Virus Particle Entry and Infection with Similar Potency: Potential Therapeutic Implications.

    Science.gov (United States)

    Nelson, Elizabeth A; Barnes, Alyson B; Wiehle, Ronald D; Fontenot, Gregory K; Hoenen, Thomas; White, Judith M

    2016-08-02

    The 2014 outbreak of Ebola virus (EBOV) in Western Africa highlighted the need for anti-EBOV therapeutics. Clomiphene is a U.S. Food and Drug Administration (FDA)-approved drug that blocks EBOV entry and infection in cells and significantly protects EBOV-challenged mice. As provided, clomiphene is, approximately, a 60:40 mixture of two stereoisomers, enclomiphene and zuclomiphene. The pharmacokinetic properties of the two isomers vary, but both accumulate in the eye and male reproductive tract, tissues in which EBOV can persist. Here we compared the ability of clomiphene and its isomers to inhibit EBOV using viral-like particle (VLP) entry and transcription/replication-competent VLP (trVLP) assays. Clomiphene and its isomers inhibited the entry and infection of VLPs and trVLPs with similar potencies. This was demonstrated with VLPs bearing the glycoproteins from three filoviruses (EBOV Mayinga, EBOV Makona, and Marburg virus) and in two cell lines (293T/17 and Vero E6). Visual problems have been noted in EBOV survivors, and viral RNA has been isolated from semen up to nine months post-infection. Since the clomiphene isomers accumulate in these affected tissues, clomiphene or one of its isomers warrants consideration as an anti-EBOV agent, for example, to potentially help ameliorate symptoms in EBOV survivors.

  8. Clomiphene and Its Isomers Block Ebola Virus Particle Entry and Infection with Similar Potency: Potential Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Elizabeth A. Nelson

    2016-08-01

    Full Text Available The 2014 outbreak of Ebola virus (EBOV in Western Africa highlighted the need for anti-EBOV therapeutics. Clomiphene is a U.S. Food and Drug Administration (FDA-approved drug that blocks EBOV entry and infection in cells and significantly protects EBOV-challenged mice. As provided, clomiphene is, approximately, a 60:40 mixture of two stereoisomers, enclomiphene and zuclomiphene. The pharmacokinetic properties of the two isomers vary, but both accumulate in the eye and male reproductive tract, tissues in which EBOV can persist. Here we compared the ability of clomiphene and its isomers to inhibit EBOV using viral-like particle (VLP entry and transcription/replication-competent VLP (trVLP assays. Clomiphene and its isomers inhibited the entry and infection of VLPs and trVLPs with similar potencies. This was demonstrated with VLPs bearing the glycoproteins from three filoviruses (EBOV Mayinga, EBOV Makona, and Marburg virus and in two cell lines (293T/17 and Vero E6. Visual problems have been noted in EBOV survivors, and viral RNA has been isolated from semen up to nine months post-infection. Since the clomiphene isomers accumulate in these affected tissues, clomiphene or one of its isomers warrants consideration as an anti-EBOV agent, for example, to potentially help ameliorate symptoms in EBOV survivors.

  9. Potential Immunological Biomarkers for Detection of Mycobacterium tuberculosis Infection in a Setting Where M. tuberculosis Is Endemic, Ethiopia.

    Science.gov (United States)

    Teklu, Takele; Kwon, Keehwan; Wondale, Biniam; HaileMariam, Milkessa; Zewude, Aboma; Medhin, Girmay; Legesse, Mengistu; Pieper, Rembert; Ameni, Gobena

    2018-04-01

    Accurate diagnosis and early treatment of tuberculosis (TB) and latent TB infection (LTBI) are vital to prevent and control TB. The lack of specific biomarkers hinders these efforts. This study's purpose was to screen immunological markers that discriminate Mycobacterium tuberculosis infection outcomes in a setting where it is endemic, Ethiopia. Whole blood from 90 participants was stimulated using the ESAT-6/CFP-10 antigen cocktail. The interferon gamma (IFN-γ)-based QuantiFERON diagnostic test was used to distinguish between LTBI and uninfected control cases. Forty cytokines/chemokines were detected from antigen-stimulated plasma supernatants (SPSs) and unstimulated plasma samples (UPSs) using human cytokine/chemokine antibody microarrays. Statistical tests allowed us to identify potential biomarkers that distinguish the TB, LTBI, and healthy control groups. As expected, the levels of IFN-γ in SPSs returned a high area under the receiver operating characteristic curve (AUC) value comparing healthy controls and LTBI cases (Z = 0.911; P SPSs of TB-infected compared to healthy controls ( P SPSs and UPSs, with P values of 0.013 and 0.012, respectively, in active TB versus LTBI cases and 0.001 and 0.002, respectively, in active TB versus healthy controls. These results encourage biomarker verification studies for IL-17 and RANTES. Combinations of these cytokines may complement IFN-γ measurements to diagnose LTBI and distinguish active TB from LTBI cases. Copyright © 2018 American Society for Microbiology.

  10. Vitamin D deficiency: A potential risk factor for Clostridium difficile infection

    Directory of Open Access Journals (Sweden)

    Youssef D

    2012-10-01

    Full Text Available Dima Youssef,1 William B Grant,2 Alan N Peiris3,41Department of Internal Medicine, Division of Infectious Diseases, 2Sunlight, Nutrition and Health Research Center, San Francisco, CA USA; 3Department of Medicine, Mountain Home VAMC, 4Department of Medicine, East Tennessee State University, Johnson City, Tennessee, USAIn the July 3, 2012 issue of the journal of Risk Management and Healthcare Policy, Martinez et al present a nice review on Clostridium difficile (C. difficile infections.1 The different manifestations of this challenging disease along with the high cost and burden on the health care system were discussed. While the authors did an admirable job in discussing traditional risk factors, they do not mention vitamin D deficiency.View original paper by Martinez and colleagues.

  11. Taurine modulates neutrophil function but potentiates uropathogenic E. coli infection in the murine bladder.

    LENUS (Irish Health Repository)

    Condron, Claire

    2010-08-01

    Eradication of a urinary tract infection (UTI) appears to be related to a number of innate host defence mechanisms and their interactions with invading bacteria. Recurrent UTIs (rUTIs) pose a difficult problem in that these bacteria use both host and bacterial factors to evade elimination. Neutrophil bactericidal function is depressed, both systemically and in urine, in patients with a history of recurrent UTI. Taurine is a semi-essential amino acid and is successful in preserving neutrophil bactericidal function in urine. Taurine may preserve neutrophil function at the urothelium and thus aid UTI resolution. Adult female (6 weeks old) C57Bl\\/6 mice were randomised into three groups: a saline gavage only control group, a saline gavage + E. coli group, and a taurine gavage + E. coli group [21 g\\/70 kg taurine in 0.9% normal saline (N\\/S) for 5 days]. Whilst taurine gavage pre-treatment resulted in increased serum neutrophils respiratory burst activity, at the urothelial-endothelial interface it caused higher colony forming units in the urine and a higher incidence of E. coli invasion in the bladder wall with no evidence of increased bladder wall neutrophils infiltration on MPO assay of histological assessment. Histologically there was also evidence of reduced bladder inflammation and urothelial cell apoptosis. In conclusion, taurine effectively increases neutrophils activity but given its anti-inflammatory properties, at the expense of decreased urothelial-endothelial activation thus preventing clearance of active E. coli infection in the bladder. Despite the negative results, this study demonstrates the importance of modulating interactions at the urothelial interface.

  12. The Pathogenic Potential of Proteus mirabilis Is Enhanced by Other Uropathogens during Polymicrobial Urinary Tract Infection.

    Science.gov (United States)

    Armbruster, Chelsie E; Smith, Sara N; Johnson, Alexandra O; DeOrnellas, Valerie; Eaton, Kathryn A; Yep, Alejandra; Mody, Lona; Wu, Weisheng; Mobley, Harry L T

    2017-02-01

    Urinary catheter use is prevalent in health care settings, and polymicrobial colonization by urease-positive organisms, such as Proteus mirabilis and Providencia stuartii, commonly occurs with long-term catheterization. We previously demonstrated that coinfection with P. mirabilis and P. stuartii increased overall urease activity in vitro and disease severity in a model of urinary tract infection (UTI). In this study, we expanded these findings to a murine model of catheter-associated UTI (CAUTI), delineated the contribution of enhanced urease activity to coinfection pathogenesis, and screened for enhanced urease activity with other common CAUTI pathogens. In the UTI model, mice coinfected with the two species exhibited higher urine pH values, urolithiasis, bacteremia, and more pronounced tissue damage and inflammation compared to the findings for mice infected with a single species, despite having a similar bacterial burden within the urinary tract. The presence of P. stuartii, regardless of urease production by this organism, was sufficient to enhance P. mirabilis urease activity and increase disease severity, and enhanced urease activity was the predominant factor driving tissue damage and the dissemination of both organisms to the bloodstream during coinfection. These findings were largely recapitulated in the CAUTI model. Other uropathogens also enhanced P. mirabilis urease activity in vitro, including recent clinical isolates of Escherichia coli, Enterococcus faecalis, Klebsiella pneumoniae, and Pseudomonas aeruginosa We therefore conclude that the underlying mechanism of enhanced urease activity may represent a widespread target for limiting the detrimental consequences of polymicrobial catheter colonization, particularly by P. mirabilis and other urease-positive bacteria. Copyright © 2017 American Society for Microbiology.

  13. Immunomodulating microRNAs of mycobacterial infections.

    Science.gov (United States)

    Bettencourt, Paulo; Pires, David; Anes, Elsa

    2016-03-01

    MicroRNAs are a class of small non-coding RNAs that have emerged as key regulators of gene expression at the post-transcriptional level by sequence-specific binding to target mRNAs. Some microRNAs block translation, while others promote mRNA degradation, leading to a reduction in protein availability. A single miRNA can potentially regulate the expression of multiple genes and their encoded proteins. Therefore, miRNAs can influence molecular signalling pathways and regulate many biological processes in health and disease. Upon infection, host cells rapidly change their transcriptional programs, including miRNA expression, as a response against the invading microorganism. Not surprisingly, pathogens can also alter the host miRNA profile to their own benefit, which is of major importance to scientists addressing high morbidity and mortality infectious diseases such as tuberculosis. In this review, we present recent findings on the miRNAs regulation of the host response against mycobacterial infections, providing new insights into host-pathogen interactions. Understanding these findings and its implications could reveal new opportunities for designing better diagnostic tools, therapies and more effective vaccines. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Proteomic analysis of the herpes simplex virus 1 virion protein 16 transactivator protein in infected cells.

    Science.gov (United States)

    Suk, Hyung; Knipe, David M

    2015-06-01

    The herpes simplex virus 1 virion protein 16 (VP16) tegument protein forms a transactivation complex with the cellular proteins host cell factor 1 (HCF-1) and octamer-binding transcription factor 1 (Oct-1) upon entry into the host cell. VP16 has also been shown to interact with a number of virion tegument proteins and viral glycoprotein H to promote viral assembly, but no comprehensive study of the VP16 proteome has been performed at early times postinfection. We therefore performed a proteomic analysis of VP16-interacting proteins at 3 h postinfection. We confirmed the interaction of VP16 with HCF-1 and a large number of cellular Mediator complex proteins, but most surprisingly, we found that the major viral protein associating with VP16 is the infected cell protein 4 (ICP4) immediate-early (IE) transactivator protein. These results raise the potential for a new function for VP16 in associating with the IE ICP4 and playing a role in transactivation of early and late gene expression, in addition to its well-documented function in transactivation of IE gene expression. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Role of Small RNAs in Trypanosomatid Infections

    Science.gov (United States)

    Linhares-Lacerda, Leandra; Morrot, Alexandre

    2016-01-01

    Trypanosomatid parasites survive and replicate in the host by using mechanisms that aim to establish a successful infection and ensure parasite survival. Evidence points to microRNAs as new players in the host-parasite interplay. MicroRNAs are small non-coding RNAs that control proteins levels via post-transcriptional gene down-regulation, either within the cells where they were produced or in other cells via intercellular transfer. These microRNAs can be modulated in host cells during infection and are among the growing group of small regulatory RNAs, for which many classes have been described, including the transfer RNA-derived small RNAs. Parasites can either manipulate microRNAs to evade host-driven damage and/or transfer small RNAs to host cells. In this mini-review, we present evidence for the involvement of small RNAs, such as microRNAs, in trypanosomatid infections which lack RNA interference. We highlight both microRNA profile alterations in host cells during those infections and the horizontal transfer of small RNAs and proteins from parasites to the host by membrane-derived extracellular vesicles in a cell communication mechanism. PMID:27065454

  16. Small cationic antimicrobial peptidomimetics: emerging candidate for the development of potential anti-infective agents.

    Science.gov (United States)

    Lohan, Sandeep; Bisht, Gopal Singh

    2013-01-01

    Rapid increase in the emergence and spread of microbes resistant to conventionally used antibiotics has become a major threat to global health care. Antimicrobial peptides (AMPs) are considered as a potential source of novel antibiotics because of their numerous advantages such as broad-spectrum activity, lower tendency to induce resistance, immunomodulatory response and unique mode of action. However, AMPs have several drawbacks such as; susceptibility to protease degradation, toxicity and high costs of manufacturing. Therefore, extensive research efforts are underway to explore the therapeutic potential of these fascinating natural compounds. This review highlights the potential of small cationic antimicrobial peptidomimetics (SCAMPs; M.W. ≅ 700 Da) as new generation antibiotics. In particular, we focused on recently identified small active pharmacophore from bulky templates of native AMPs, β-peptides, and lipopeptides. In addition, various design strategies recently undertaken to improve the physicochemical properties (proteolytic stability & plasma protein binding) of small cationic peptides have also been discussed.

  17. The multifunctional LigB adhesin binds homeostatic proteins with potential roles in cutaneous infection by pathogenic Leptospira interrogans.

    Directory of Open Access Journals (Sweden)

    Henry A Choy

    Full Text Available Leptospirosis is a potentially fatal zoonotic disease in humans and animals caused by pathogenic spirochetes, such as Leptospira interrogans. The mode of transmission is commonly limited to the exposure of mucous membrane or damaged skin to water contaminated by leptospires shed in the urine of carriers, such as rats. Infection occurs during seasonal flooding of impoverished tropical urban habitats with large rat populations, but also during recreational activity in open water, suggesting it is very efficient. LigA and LigB are surface localized proteins in pathogenic Leptospira strains with properties that could facilitate the infection of damaged skin. Their expression is rapidly induced by the increase in osmolarity encountered by leptospires upon transition from water to host. In addition, the immunoglobulin-like repeats of the Lig proteins bind proteins that mediate attachment to host tissue, such as fibronectin, fibrinogen, collagens, laminin, and elastin, some of which are important in cutaneous wound healing and repair. Hemostasis is critical in a fresh injury, where fibrinogen from damaged vasculature mediates coagulation. We show that fibrinogen binding by recombinant LigB inhibits fibrin formation, which could aid leptospiral entry into the circulation, dissemination, and further infection by impairing healing. LigB also binds fibroblast fibronectin and type III collagen, two proteins prevalent in wound repair, thus potentially enhancing leptospiral adhesion to skin openings. LigA or LigB expression by transformation of a nonpathogenic saprophyte, L. biflexa, enhances bacterial adhesion to fibrinogen. Our results suggest that by binding homeostatic proteins found in cutaneous wounds, LigB could facilitate leptospirosis transmission. Both fibronectin and fibrinogen binding have been mapped to an overlapping domain in LigB comprising repeats 9-11, with repeat 11 possibly enhancing binding by a conformational effect. Leptospirosis

  18. Strongyloides stercoralis age-1: a potential regulator of infective larval development in a parasitic nematode.

    Directory of Open Access Journals (Sweden)

    Jonathan D Stoltzfus

    Full Text Available Infective third-stage larvae (L3i of the human parasite Strongyloides stercoralis share many morphological, developmental, and behavioral attributes with Caenorhabditis elegans dauer larvae. The 'dauer hypothesis' predicts that the same molecular genetic mechanisms control both dauer larval development in C. elegans and L3i morphogenesis in S. stercoralis. In C. elegans, the phosphatidylinositol-3 (PI3 kinase catalytic subunit AGE-1 functions in the insulin/IGF-1 signaling (IIS pathway to regulate formation of dauer larvae. Here we identify and characterize Ss-age-1, the S. stercoralis homolog of the gene encoding C. elegans AGE-1. Our analysis of the Ss-age-1 genomic region revealed three exons encoding a predicted protein of 1,209 amino acids, which clustered with C. elegans AGE-1 in phylogenetic analysis. We examined temporal patterns of expression in the S. stercoralis life cycle by reverse transcription quantitative PCR and observed low levels of Ss-age-1 transcripts in all stages. To compare anatomical patterns of expression between the two species, we used Ss-age-1 or Ce-age-1 promoter::enhanced green fluorescent protein reporter constructs expressed in transgenic animals for each species. We observed conservation of expression in amphidial neurons, which play a critical role in developmental regulation of both dauer larvae and L3i. Application of the PI3 kinase inhibitor LY294002 suppressed L3i in vitro activation in a dose-dependent fashion, with 100 µM resulting in a 90% decrease (odds ratio: 0.10, 95% confidence interval: 0.08-0.13 in the odds of resumption of feeding for treated L3i in comparison to the control. Together, these data support the hypothesis that Ss-age-1 regulates the development of S. stercoralis L3i via an IIS pathway in a manner similar to that observed in C. elegans dauer larvae. Understanding the mechanisms by which infective larvae are formed and activated may lead to novel control measures and treatments for

  19. The potential of methylethylpiridinol in treatment of bacterial infections caused by Klebsiella pneumoniae (experimental study

    Directory of Open Access Journals (Sweden)

    V. M. Brykhanov

    2016-01-01

    Full Text Available Aim. Investigated the activity of methylethylpiridinol (6-methyl-2-ethyl-pyridin-3-ol hydrochloride in the comprehensive treatment of the experimental bacterial infection caused by Klebsiella pneumoniae.Materials and methods. The study was conducted on clinical isolates of Klebsiella pneumoniae. At the first stage of the study (in vitro studied the effect of methylethylpiridinol in concentrations 0,25–4 mM on the growth of the strain and the activity of the sublethal concentrations of antibiotics – gentamicin, ciprofloxacin, tetracycline, ceftazidime. In the second stage of the study (in vivo in rats Wistar simulated bacterial peritonitis by intraperitoneal injection of a suspension of Klebsiella pneumoniae and investigated the effect of methylethylpiridinol (80 mg/kg on the effectiveness of antibiotic therapy with gentamicin (30 mg/kg, ciprofloxacin (50 mg/kg, ceftazidime (120 mg/kg or tetracycline (80 mg/kg. The animal blood plasma was determined ceruloplasmin concentration (marker of the intensity of infectious-inflammatory process and thiobarbiturate-jet products, erythrocytes – the concentration of reduced glutathione, catalase and glutathione peroxidase.Results. It is found that a methylethylpiridinol inhibits the development of periodic bacterial cultures, but exhibits a pronounced antagonism with respect to gentamicin. Antioxidant slightly increases the activity of ciprofloxacin and tetracycline. The bacteriostatic effect of antioxidant reduces the action of ceftazidime in vitro. In conditions of chemotherapy by using of gentamicin and ciprofloxacin additional injection of methylethylpiridinol leads to the preservation of ceruloplasmin level to the level of non-treated animals without showing the antioxidant effect. Ceftazidime exhibits antioxidant effect, reduces the introduction of methylethylpiridinol. The antioxidant properties of methylethylpiridinol did not appear in the application of

  20. Influence of Stress Connected with Moving to a New Farm on Potentially MAP-Infected Mouflons.

    Science.gov (United States)

    Pribylova-Dziedzinska, Radka; Slana, Iva; Lamka, Jiri; Pavlik, Ivo

    2014-01-01

    There is no European legislation concerning paratuberculosis that requires that imported animals be kept in quarantine and commonly they are directly released into areas with other animals. In this study, detection of latent infection of paratuberculosis in healthy mouflons previously diagnosed as paratuberculosis-free, but originating from a real time quantitative PCR- (qPCR-) positive herd, occurred after their transport to a new farm. During a twelve-day quarantine period, all mouflons irregularly shed Mycobacterium avium subsp. paratuberculosis (MAP) in faeces, and in a small number of cases also in milk. After the animals were released from quarantine, MAP was detected for a further two days, after which, testing was negative, except in one case. Therefore, the stress connected with transport, novel environment, dietary change, or limited area with high density of animals might have contributed to the induction of paratuberculosis and the shedding of MAP from the animals, previously diagnosed as MAP-negative. According to these results, the keeping of imported animals in quarantine and their examination for MAP presence not only before the transport but also afterwards should be recommended. The designation of a particular area of a farm as a quarantine enclosure could help to mitigate the impact of stress caused by a confined space with a high density of animals.

  1. Are healthcare workers' mobile phones a potential source of nosocomial infections? Review of the literature.

    Science.gov (United States)

    Ulger, Fatma; Dilek, Ahmet; Esen, Saban; Sunbul, Mustafa; Leblebicioglu, Hakan

    2015-10-29

    Mobile communication devices help accelerate in-hospital flow of medical information, information sharing and querying, and contribute to communications in the event of emergencies through their application and access to wireless media technology. Healthcare-associated infections remain a leading and high-cost problem of global health systems despite improvements in modern therapies. The objective of this article was to review different studies on the relationship between mobile phones (MPs) and bacterial cross-contamination and report common findings. Thirty-nine studies published between 2005 and 2013 were reviewed. Of these, 19 (48.7%) identified coagulase-negative staphylococci (CoNS), and 26 (66.7%) identified Staphylococcus aureus; frequency of growth varied. The use of MPs by healthcare workers increases the risk of repetitive cyclic contamination between the hands and face (e.g., nose, ears, and lips), and differences in personal hygiene and behaviors can further contribute to the risks. MPs are rarely cleaned after handling. They may transmit microorganisms, including multiple resistant strains, after contact with patients, and can be a source of bacterial cross-contamination. To prevent bacterial contamination of MPs, hand-washing guidelines must be followed and technical standards for prevention strategies should be developed.

  2. Assessing the potential spread and maintenance of foot-and-mouth disease virus infection in wild ungulates: general principles and application to a specific scenario in Thrace

    DEFF Research Database (Denmark)

    Dhollander, S.; Belsham, Graham; Lange, M.

    2016-01-01

    Foot-and-mouth disease (FMD), due to infection with serotype O virus, occurred in wild boar and within eleven outbreaks in domestic livestock in the south-east of Bulgaria, Thrace region, in 2011. Hence, the issue of the potential for the spread and maintenance of FMD virus (FMDV) infection...... in a population of wild ungulates became important. This assessment focused on the spread and maintenance of FMDV infection within a hypothetical wild boar and deer population in an environment, which is characterized by a climate transitional between Mediterranean and continental and variable wildlife population...... densities. The assessment was based on three aspects: (i) a systematic review of the literature focusing on experimental infection studies to identify the parameters describing the duration of FMDV infection in deer and wild boar, as well as observational studies assessing the occurrence of FMDV infection...

  3. A novel, broad-spectrum inhibitor of enterovirus replication that targets host cell factor phosphatidylinositol 4-kinase IIIβ

    NARCIS (Netherlands)

    van der Schaar, H.M.; Leyssen, Pieter; Thibaut, H.J.; de Palma, Armando; van der Linden, Lonneke; Lanke, Kjerstin H.W.; Lacroix, Céline; Verbeken, Erik; Conrath, Katja; Macleod, Angus M; Mitchell, Dale R; Palmer, Nicholas J; van de Poël, Hervé; Andrews, Martin; Neyts, Johan; van Kuppeveld, F.J.M.

    2013-01-01

    Despite their high clinical and socioeconomic impacts, there is currently no approved antiviral therapy for the prophylaxis or treatment of enterovirus infections. Here we report on a novel inhibitor of enterovirus replication, compound 1,

  4. Mobile Phones as a Potential Vehicle of Infection in a Hospital Setting.

    Science.gov (United States)

    Chao Foong, Yi; Green, Mark; Zargari, Ahmad; Siddique, Romana; Tan, Vanessa; Brain, Terry; Ogden, Kathryn

    2015-01-01

    The objective of this article is to investigate the potential role of mobile phones as a reservoir for bacterial colonization and the risk factors for bacterial colonization in a hospital setting. We screened 226 staff members at a regional Australian hospital (146 doctors and 80 medical students) between January 2013 and March 2014. The main outcomes of interest were the types of microorganisms and the amount of contamination of the mobile phones. This study found a high level of bacterial contamination (n = 168/226, 74%) on the mobile phones of staff members in a tertiary hospital, with similar organisms isolated from the staff member's dominant hand and mobile phones. While most of the isolated organisms were normal skin flora, a small percentage were potentially pathogenic (n = 12/226, 5%). Being a junior medical staff was found to be a risk factor for heavy microbial growth (OR 4.00, 95% CI 1.54, 10.37). Only 31% (70/226) of our participants reported cleaning their phones routinely, and only 21% (47/226) reported using alcohol containing wipes on their phones. This study demonstrates that mobile phones are potentially vehicles for pathogenic bacteria in a hospital setting. Only a minority of our participants reported cleaning their phones routinely. Disinfection guidelines utilizing alcohol wipes should be developed and implemented.

  5. A Pathogenic Potential of Acinetobacter baumannii-Derived Membrane Vesicles

    Directory of Open Access Journals (Sweden)

    Jong Suk Jin

    2011-12-01

    Full Text Available Acinetobacter baumannii secretes outer membrane vesicles (OMVs. A. baumannii OMVs deliver many virulence factors to host cells and then induce cytotoxicity and innate immune response. OMVs secreted from bacteria contribute directly to host pathology during A. baumannii infection.

  6. Methylene Blue-Loaded Dissolving Microneedles: Potential Use in Photodynamic Antimicrobial Chemotherapy of Infected Wounds

    Directory of Open Access Journals (Sweden)

    Ester Caffarel-Salvador

    2015-09-01

    Full Text Available Photodynamic therapy involves delivery of a photosensitising drug that is activated by light of a specific wavelength, resulting in generation of highly reactive radicals. This activated species can cause destruction of targeted cells. Application of this process for treatment of microbial infections has been termed “photodynamic antimicrobial chemotherapy” (PACT. In the treatment of chronic wounds, the delivery of photosensitising agents is often impeded by the presence of a thick hyperkeratotic/necrotic tissue layer, reducing their therapeutic efficacy. Microneedles (MNs are an emerging drug delivery technology that have been demonstrated to successfully penetrate the outer layers of the skin, whilst minimising damage to skin barrier function. Delivering photosensitising drugs using this platform has been demonstrated to have several advantages over conventional photodynamic therapy, such as, painless application, reduced erythema, enhanced cosmetic results and improved intradermal delivery. The aim of this study was to physically characterise dissolving MNs loaded with the photosensitising agent, methylene blue and assess their photodynamic antimicrobial activity. Dissolving MNs were fabricated from aqueous blends of Gantrez® AN-139 co-polymer containing varying loadings of methylene blue. A height reduction of 29.8% was observed for MNs prepared from blends containing 0.5% w/w methylene blue following application of a total force of 70.56 N/array. A previously validated insertion test was used to assess the effect of drug loading on MN insertion into a wound model. Staphylococcus aureus, Escherichia coli and Candida albicans biofilms were incubated with various methylene blue concentrations within the range delivered by MNs in vitro (0.1–2.5 mg/mL and either irradiated at 635 nm using a Paterson Lamp or subjected to a dark period. Microbial susceptibility to PACT was determined by assessing the total viable count. Kill rates of >96

  7. PRODUCTIVE POTENTIAL OF THE CHERRY TOMATO GENOTYPE GROUP BEFORE INFECTION BY Alternaria tomatophila

    Directory of Open Access Journals (Sweden)

    HUGO CESAR RODRIGUES MOREIRA CATÃO

    2017-01-01

    Full Text Available Early blight (caused by Alternaria tomatophila is a major disease of tomato with no resistant cultivars. Thus, it is necessary to identify sources of resistance and productive genotypes for the development of new cultivars. Therefore, this study aimed to evaluate the productive potential of cherry tomato genotypes grown in the summer / fall, the severity of early blight on leaves and the incidence of disease in fruits. The treatments consisted of Carolina tomato genotypes, Cereja Vermelho, CH 152 and CLN1561A. The experimental design consisted of randomized blocks with six replications, and the experimental plot had 16 plants. The following characteristics were evaluated: area under the disease progress curve (AUDPC, average number of microinjuries on the fruits (MF, average number of fruits per bunch (NFC, average number of bunches per plant (NCP, average number of fruits per plant (NFP, average yield, number of fruits with incidence of early blight per plant (NFI and the severity of early blight in leaves (%. The cherry tomato genotype CH152 showed tolerance to early blight with a smaller area under the disease progress curve, lower severity and fruits with incidence of A. tomatophila were not observed in this genotype. The CH152 had the highest number of fruits per bunch, greater number of bunches per plant, higher number of fruits per plant and higher productivity. This line has great potential of being integrated into breeding programs.

  8. Banana infecting fungus, Fusarium musae, is also an opportunistic human pathogen: are bananas potential carriers and source of fusariosis?

    Science.gov (United States)

    Triest, David; Stubbe, Dirk; De Cremer, Koen; Piérard, Denis; Detandt, Monique; Hendrickx, Marijke

    2015-01-01

    During re-identification of Fusarium strains in the BCCM™/IHEM fungal collection by multilocus sequence-analysis we observed that five strains, previously identified as Fusarium verticillioides, were Fusarium musae, a species described in 2011 from banana fruits. Four strains were isolated from blood samples or biopsies of immune-suppressed patients and one was isolated from the clinical environment, all originating from different hospitals in Belgium or France, 2001-2008. The F. musae identity of our isolates was confirmed by phylogenetic analysis using reference sequences of type material. Absence of the gene cluster necessary for fumonisin biosynthesis, characteristic to F. musae, was also the case for our isolates. In vitro antifungal susceptibility testing revealed no important differences in their susceptibility compared to clinical F. verticillioides strains and terbinafine was the most effective drug. Additional clini