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Sample records for stomatitis virus vectors

  1. Recombinant vesicular stomatitis virus vaccine vectors expressing filovirus glycoproteins lack neurovirulence in nonhuman primates.

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    Chad E Mire

    Full Text Available The filoviruses, Marburg virus and Ebola virus, cause severe hemorrhagic fever with high mortality in humans and nonhuman primates. Among the most promising filovirus vaccines under development is a system based on recombinant vesicular stomatitis virus (rVSV that expresses an individual filovirus glycoprotein (GP in place of the VSV glycoprotein (G. The main concern with all replication-competent vaccines, including the rVSV filovirus GP vectors, is their safety. To address this concern, we performed a neurovirulence study using 21 cynomolgus macaques where the vaccines were administered intrathalamically. Seven animals received a rVSV vector expressing the Zaire ebolavirus (ZEBOV GP; seven animals received a rVSV vector expressing the Lake Victoria marburgvirus (MARV GP; three animals received rVSV-wild type (wt vector, and four animals received vehicle control. Two of three animals given rVSV-wt showed severe neurological symptoms whereas animals receiving vehicle control, rVSV-ZEBOV-GP, or rVSV-MARV-GP did not develop these symptoms. Histological analysis revealed major lesions in neural tissues of all three rVSV-wt animals; however, no significant lesions were observed in any animals from the filovirus vaccine or vehicle control groups. These data strongly suggest that rVSV filovirus GP vaccine vectors lack the neurovirulence properties associated with the rVSV-wt parent vector and support their further development as a vaccine platform for human use.

  2. Grasshoppers (Orthoptera: Acrididae) could serve as reservoirs and vectors of vesicular stomatitis virus.

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    Nunamaker, Richard A; Lockwood, Jeffrey A; Stith, Charles E; Campbell, Corey L; Schell, Scott P; Drolet, Barbara S; Wilson, William C; White, David M; Letchworth, Geoffrey J

    2003-11-01

    Vesicular stomatitis (VS) is an economically devastating disease of livestock in the Americas. Despite strong circumstantial evidence for the role of arthropods in epizootics, no hematophagous vector explains the field evidence. Based on the spatiotemporal association of grasshopper outbreaks and VS epizootics, we investigated the potential role of these insects as vectors and reservoirs of the disease. The critical steps in the grasshopper-bovine transmission cycle were demonstrated, including 1) 62% of grasshoppers [Melanoplus sanguinipes (F.)] fed vesicular stomatitis virus (VSV) from cell culture became infected, with titers reaching 40,000 times the inoculative dose; 2) 40% of grasshoppers that cannibalized VSV-infected grasshopper cadavers became infected, amplifying virus up to 1,000-fold; 3) one of three cattle consuming VSV-infected grasshopper cadavers contracted typical VS and shed virus in saliva; and 4) 15% of grasshoppers became infected when fed saliva from this infected cow. The ecological conditions and biological processes necessary for these transmissions to occur are present throughout much of the Americas. Field studies will be required to show these findings are relevant to the natural epidemiology of VSV.

  3. Immunogenicity in African Green Monkeys of M Protein Mutant Vesicular Stomatitis Virus Vectors and Contribution of Vector-Encoded Flagellin

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    Marlena M. Westcott

    2018-03-01

    Full Text Available Recombinant vesicular stomatitis virus (VSV is a promising platform for vaccine development. M51R VSV, an attenuated, M protein mutant strain, is an effective inducer of Type I interferon and dendritic cell (DC maturation, which are desirable properties to exploit for vaccine design. We have previously evaluated M51R VSV (M51R and M51R VSV that produces flagellin (M51R-F as vaccine vectors using murine models, and found that flagellin enhanced DC activation and VSV-specific antibody production after low-dose vaccination. In this report, the immunogenicity of M51R vectors and the adjuvant effect of virus-produced flagellin were evaluated in nonhuman primates following high-dose (108 pfu and low-dose (105 pfu vaccination. A single intramuscular vaccination of African green monkeys with M51R or M51R-F induced VSV-specific, dose-dependent humoral immune responses. Flagellin induced a significant increase in antibody production (IgM, IgG and neutralizing antibody at the low vaccination dose. A VSV-specific cellular response was detected at 6 weeks post-vaccination, but was neither dose-dependent nor enhanced by flagellin; similar numbers of VSV-specific, IFNγ-producing cells were detected in lymph node and spleen of all animals. These results indicate that virus-directed, intracellular flagellin production may improve VSV-based vaccines encoding heterologous antigens by lowering the dose required to achieve humoral immunity.

  4. Vaccination with Vesicular Stomatitis Virus-Vectored Chimeric Hemagglutinins Protects Mice against Divergent Influenza Virus Challenge Strains.

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    Ryder, Alex B; Nachbagauer, Raffael; Buonocore, Linda; Palese, Peter; Krammer, Florian; Rose, John K

    2015-12-16

    Seasonal influenza virus infections continue to cause significant disease each year, and there is a constant threat of the emergence of reassortant influenza strains causing a new pandemic. Available influenza vaccines are variably effective each season, are of limited scope at protecting against viruses that have undergone significant antigenic drift, and offer low protection against newly emergent pandemic strains. "Universal" influenza vaccine strategies that focus on the development of humoral immunity directed against the stalk domains of the viral hemagglutinin (HA) show promise for protecting against diverse influenza viruses. Here, we describe such a strategy that utilizes vesicular stomatitis virus (VSV) as a vector for chimeric hemagglutinin (cHA) antigens. This vaccination strategy is effective at generating HA stalk-specific, broadly cross-reactive serum antibodies by both intramuscular and intranasal routes of vaccination. We show that prime-boost vaccination strategies provide protection against both lethal homologous and heterosubtypic influenza challenge and that protection is significantly improved with intranasal vaccine administration. Additionally, we show that vaccination with VSV-cHAs generates greater stalk-specific and cross-reactive serum antibodies than does vaccination with VSV-vectored full-length HAs, confirming that cHA-based vaccination strategies are superior at generating stalk-specific humoral immunity. VSV-vectored influenza vaccines that express chimeric hemagglutinin antigens offer a novel means for protecting against widely diverging influenza viruses. Universal influenza vaccination strategies should be capable of protecting against a wide array of influenza viruses, and we have developed such an approach utilizing a single viral vector system. The potent antibody responses that these vaccines generate are shown to protect mice against lethal influenza challenges with highly divergent viruses. Notably, intranasal vaccination

  5. Emergency postexposure vaccination with vesicular stomatitis virus-vectored Ebola vaccine after needlestick.

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    Lai, Lilin; Davey, Richard; Beck, Allison; Xu, Yongxian; Suffredini, Anthony F; Palmore, Tara; Kabbani, Sarah; Rogers, Susan; Kobinger, Gary; Alimonti, Judie; Link, Charles J; Rubinson, Lewis; Ströher, Ute; Wolcott, Mark; Dorman, William; Uyeki, Timothy M; Feldmann, Heinz; Lane, H Clifford; Mulligan, Mark J

    Safe and effective vaccines and drugs are needed for the prevention and treatment of Ebola virus disease, including following a potentially high-risk exposure such as a needlestick. To assess response to postexposure vaccination in a health care worker who was exposed to the Ebola virus. Case report of a physician who experienced a needlestick while working in an Ebola treatment unit in Sierra Leone on September 26, 2014. Medical evacuation to the United States was rapidly initiated. Given the concern about potentially lethal Ebola virus disease, the patient was offered, and provided his consent for, postexposure vaccination with an experimental vaccine available through an emergency Investigational New Drug application. He was vaccinated on September 28, 2014. The vaccine used was VSVΔG-ZEBOV, a replicating, attenuated, recombinant vesicular stomatitis virus (serotype Indiana) whose surface glycoprotein gene was replaced by the Zaire Ebola virus glycoprotein gene. This vaccine has entered a clinical trial for the prevention of Ebola in West Africa. The vaccine was administered 43 hours after the needlestick occurred. Fever and moderate to severe symptoms developed 12 hours after vaccination and diminished over 3 to 4 days. The real-time reverse transcription polymerase chain reaction results were transiently positive for vesicular stomatitis virus nucleoprotein gene and Ebola virus glycoprotein gene (both included in the vaccine) but consistently negative for Ebola virus nucleoprotein gene (not in the vaccine). Early postvaccination cytokine secretion and T lymphocyte and plasmablast activation were detected. Subsequently, Ebola virus glycoprotein-specific antibodies and T cells became detectable, but antibodies against Ebola viral matrix protein 40 (not in the vaccine) were not detected. It is unknown if VSVΔG-ZEBOV is safe or effective for postexposure vaccination in humans who have experienced a high-risk occupational exposure to the Ebola virus, such as a

  6. Attenuation of Recombinant Vesicular Stomatitis Virus-Human Immunodeficiency Virus Type 1 Vaccine Vectors by Gene Translocations and G Gene Truncation Reduces Neurovirulence and Enhances Immunogenicity in Mice▿

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    Cooper, David; Wright, Kevin J.; Calderon, Priscilla C.; Guo, Min; Nasar, Farooq; Johnson, J. Erik; Coleman, John W.; Lee, Margaret; Kotash, Cheryl; Yurgelonis, Irene; Natuk, Robert J.; Hendry, R. Michael; Udem, Stephen A.; Clarke, David K.

    2008-01-01

    Recombinant vesicular stomatitis virus (rVSV) has shown great potential as a new viral vector for vaccination. However, the prototypic rVSV vector described previously was found to be insufficiently attenuated for clinical evaluation when assessed for neurovirulence in nonhuman primates. Here, we describe the attenuation, neurovirulence, and immunogenicity of rVSV vectors expressing human immunodeficiency virus type 1 Gag. These rVSV vectors were attenuated by combinations of the following manipulations: N gene translocations (N4), G gene truncations (CT1 or CT9), noncytopathic M gene mutations (Mncp), and positioning of the gag gene into the first position of the viral genome (gag1). The resulting N4CT1-gag1, N4CT9-gag1, and MncpCT1-gag1 vectors demonstrated dramatically reduced neurovirulence in mice following direct intracranial inoculation. Surprisingly, in spite of a very high level of attenuation, the N4CT1-gag1 and N4CT9-gag1 vectors generated robust Gag-specific immune responses following intramuscular immunization that were equivalent to or greater than immune responses generated by the more virulent prototypic vectors. MncpCT1-gag1 also induced Gag-specific immune responses following intramuscular immunization that were equivalent to immune responses generated by the prototypic rVSV vector. Placement of the gag gene in the first position of the VSV genome was associated with increased in vitro expression of Gag protein, in vivo expression of Gag mRNA, and enhanced immunogenicity of the vector. These findings demonstrate that through directed manipulation of the rVSV genome, vectors that have reduced neurovirulence and enhanced immunogenicity can be made. PMID:17942549

  7. Heat Shock Protein 70 Enhances Mucosal Immunity against Human Norovirus When Coexpressed from a Vesicular Stomatitis Virus Vector

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    Ma, Yuanmei; Duan, Yue; Wei, Yongwei; Liang, Xueya; Niewiesk, Stefan; Oglesbee, Michael

    2014-01-01

    ABSTRACT Human norovirus (NoV) accounts for 95% of nonbacterial gastroenteritis worldwide. Currently, there is no vaccine available to combat human NoV as it is not cultivable and lacks a small-animal model. Recently, we demonstrated that recombinant vesicular stomatitis virus (rVSV) expressing human NoV capsid protein (rVSV-VP1) induced strong immunities in mice (Y. Ma and J. Li, J. Virol. 85:2942–2952, 2011). To further improve the safety and efficacy of the vaccine candidate, heat shock protein 70 (HSP70) was inserted into the rVSV-VP1 backbone vector. A second construct was generated in which the firefly luciferase (Luc) gene was inserted in place of HSP70 as a control for the double insertion. The resultant recombinant viruses (rVSV-HSP70-VP1 and rVSV-Luc-VP1) were significantly more attenuated in cell culture and viral spread in mice than rVSV-VP1. At the inoculation dose of 1.0 × 106 PFU, rVSV-HSP70-VP1 triggered significantly higher vaginal IgA than rVSV-VP1 and significantly higher fecal and vaginal IgA responses than rVSV-Luc-VP1, although serum IgG and T cell responses were similar. At the inoculation dose of 5.0 × 106 PFU, rVSV-HSP70-VP1 stimulated significantly higher T cell, fecal, and vaginal IgA responses than rVSV-VP1. Fecal and vaginal IgA responses were also significantly increased when combined vaccination of rVSV-VP1 and rVSV-HSP70 was used. Collectively, these data indicate that (i) insertion of an additional gene (HSP70 or Luc) into the rVSV-VP1 backbone further attenuates the VSV-based vaccine in vitro and in vivo, thus improving the safety of the vaccine candidate, and (ii) HSP70 enhances the human NoV-specific mucosal and T cell immunities triggered by a VSV-based human NoV vaccine. IMPORTANCE Human norovirus (NoV) is responsible for more than 95% of acute nonbacterial gastroenteritis worldwide. Currently, there is no vaccine for this virus. Development of a live attenuated vaccine for human NoV has not been possible because it is

  8. Understanding and altering cell tropism of vesicular stomatitis virus

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    Hastie, Eric; Cataldi, Marcela; Marriott, Ian; Grdzelishvili, Valery Z.

    2013-01-01

    Vesicular stomatitis virus (VSV) is a prototypic nonsegmented negative-strand RNA virus. VSV’s broad cell tropism makes it a popular model virus for many basic research applications. In addition, a lack of preexisting human immunity against VSV, inherent oncotropism and other features make VSV a widely used platform for vaccine and oncolytic vectors. However, VSV’s neurotropism that can result in viral encephalitis in experimental animals needs to be addressed for the use of the virus as a sa...

  9. A novel, live-attenuated vesicular stomatitis virus vector displaying conformationally intact, functional HIV-1 envelope trimers that elicits potent cellular and humoral responses in mice.

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    Svetlana Rabinovich

    Full Text Available Though vaccination with live-attenuated SIV provides the greatest protection from progressive disease caused by SIV challenge in rhesus macaques, attenuated HIV presents safety concerns as a vaccine; therefore, live viral vectors carrying HIV immunogens must be considered. We have designed a replication-competent vesicular stomatitis virus (VSV displaying immunogenic HIV-1 Env trimers and attenuating quantities of the native surface glycoprotein (G. The clade B Env immunogen is an Env-VSV G hybrid (EnvG in which the transmembrane and cytoplasmic tail regions are derived from G. Relocation of the G gene to the 5'terminus of the genome and insertion of EnvG into the natural G position induced a ∼1 log reduction in surface G, significant growth attenuation compared to wild-type, and incorporation of abundant EnvG. Western blot analysis indicated that ∼75% of incorporated EnvG was a mature proteolytically processed form. Flow cytometry showed that surface EnvG bound various conformationally- and trimer-specific antibodies (Abs, and in-vitro growth assays on CD4+CCR5+ cells demonstrated EnvG functionality. Neither intranasal (IN or intramuscular (IM administration in mice induced any observable pathology and all regimens tested generated potent Env-specific ELISA titers of 10(4-10(5, with an IM VSV prime/IN VSV boost regimen eliciting the highest binding and neutralizing Ab titers. Significant quantities of Env-specific CD4+ T cells were also detected, which were augmented as much as 70-fold by priming with IM electroporated plasmids encoding EnvG and IL-12. These data suggest that our novel vector can achieve balanced safety and immunogenicity and should be considered as an HIV vaccine platform.

  10. Evaluation of the Protective Efficacy of Recombinant Vesicular Stomatitis Virus Vectors Against Marburg Hemorrhagic Fever in Nonhuman Primate Models

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    2007-01-19

    thought to range from four to ten days and the collection of clinical manifestations varies in severity depending on viral virulence , routes of exposure...number of viral infections including rabies (Rupprecht, Hanlon et al. 2004), hepatitis B (Yu, Cheung et al. 2004), smallpox (Massoudi, Barker et al...2003; Mortimer P 2003), and herpes simplex virus 2 (Stanberry L 2004). For example, rabies is responsible for a large extent of morbidity and

  11. Immunogenicity of viral vector, prime-boost SIV vaccine regimens in infant rhesus macaques: attenuated vesicular stomatitis virus (VSV) and modified vaccinia Ankara (MVA) recombinant SIV vaccines compared to live-attenuated SIV.

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    Van Rompay, Koen K A; Abel, Kristina; Earl, Patricia; Kozlowski, Pamela A; Easlick, Juliet; Moore, Joseph; Buonocore-Buzzelli, Linda; Schmidt, Kimberli A; Wilson, Robert L; Simon, Ian; Moss, Bernard; Rose, Nina; Rose, John; Marthas, Marta L

    2010-02-10

    In a previously developed infant macaque model mimicking HIV infection by breast-feeding, we demonstrated that intramuscular immunization with recombinant poxvirus vaccines expressing simian immunodeficiency virus (SIV) structural proteins provided partial protection against infection following oral inoculation with virulent SIV. In an attempt to further increase systemic but also local antiviral immune responses at the site of viral entry, we tested the immunogenicity of different orally administered, replicating vaccines. One group of newborn macaques received an oral prime immunization with a recombinant vesicular stomatitis virus expressing SIVmac239 gag, pol and env (VSV-SIVgpe), followed 2 weeks later by an intramuscular boost immunization with MVA-SIV. Another group received two immunizations with live-attenuated SIVmac1A11, administered each time both orally and intravenously. Control animals received mock immunizations or non-SIV VSV and MVA control vectors. Analysis of SIV-specific immune responses in blood and lymphoid tissues at 4 weeks of age demonstrated that both vaccine regimens induced systemic antibody responses and both systemic and local cell-mediated immune responses. The safety and immunogenicity of the VSV-SIVgpe+MVA-SIV immunization regimen described in this report provide the scientific incentive to explore the efficacy of this vaccine regimen against virulent SIV exposure in the infant macaque model. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

  12. Oncotargeting by Vesicular Stomatitis Virus (VSV: Advances in Cancer Therapy

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    Suman Bishnoi

    2018-02-01

    Full Text Available Modern oncotherapy approaches are based on inducing controlled apoptosis in tumor cells. Although a number of apoptosis-induction approaches are available, site-specific delivery of therapeutic agents still remain the biggest hurdle in achieving the desired cancer treatment benefit. Additionally, systemic treatment-induced toxicity remains a major limiting factor in chemotherapy. To specifically address drug-accessibility and chemotherapy side effects, oncolytic virotherapy (OV has emerged as a novel cancer treatment alternative. In OV, recombinant viruses with higher replication capacity and stronger lytic properties are being considered for tumor cell-targeting and subsequent cell lysing. Successful application of OVs lies in achieving strict tumor-specific tropism called oncotropism, which is contingent upon the biophysical interactions of tumor cell surface receptors with viral receptors and subsequent replication of oncolytic viruses in cancer cells. In this direction, few viral vector platforms have been developed and some of these have entered pre-clinical/clinical trials. Among these, the Vesicular stomatitis virus (VSV-based platform shows high promise, as it is not pathogenic to humans. Further, modern molecular biology techniques such as reverse genetics tools have favorably advanced this field by creating efficient recombinant VSVs for OV; some have entered into clinical trials. In this review, we discuss the current status of VSV based oncotherapy, challenges, and future perspectives regarding its therapeutic applications in the cancer treatment.

  13. Current good manufacturing practice production of an oncolytic recombinant vesicular stomatitis viral vector for cancer treatment.

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    Ausubel, L J; Meseck, M; Derecho, I; Lopez, P; Knoblauch, C; McMahon, R; Anderson, J; Dunphy, N; Quezada, V; Khan, R; Huang, P; Dang, W; Luo, M; Hsu, D; Woo, S L C; Couture, L

    2011-04-01

    Vesicular stomatitis virus (VSV) is an oncolytic virus currently being investigated as a promising tool to treat cancer because of its ability to selectively replicate in cancer cells. To enhance the oncolytic property of the nonpathologic laboratory strain of VSV, we generated a recombinant vector [rVSV(MΔ51)-M3] expressing murine gammaherpesvirus M3, a secreted viral chemokine-binding protein that binds to a broad range of mammalian chemokines with high affinity. As previously reported, when rVSV(MΔ51)-M3 was used in an orthotopic model of hepatocellular carcinoma (HCC) in rats, it suppressed inflammatory cell migration to the virus-infected tumor site, which allowed for enhanced intratumoral virus replication leading to increased tumor necrosis and substantially prolonged survival. These encouraging results led to the development of this vector for clinical translation in patients with HCC. However, a scalable current Good Manufacturing Practice (cGMP)-compliant manufacturing process has not been described for this vector. To produce the quantities of high-titer virus required for clinical trials, a process that is amenable to GMP manufacturing and scale-up was developed. We describe here a large-scale (50-liter) vector production process capable of achieving crude titers on the order of 10(9) plaque-forming units (PFU)/ml under cGMP. This process was used to generate a master virus seed stock and a clinical lot of the clinical trial agent under cGMP with an infectious viral titer of approximately 2 × 10(10) PFU/ml (total yield, 1 × 10(13) PFU). The lot has passed all U.S. Food and Drug Administration-mandated release testing and will be used in a phase 1 clinical translational trial in patients with advanced HCC.

  14. Virus-Vectored Influenza Virus Vaccines

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    Tripp, Ralph A.; Tompkins, S. Mark

    2014-01-01

    Despite the availability of an inactivated vaccine that has been licensed for >50 years, the influenza virus continues to cause morbidity and mortality worldwide. Constant evolution of circulating influenza virus strains and the emergence of new strains diminishes the effectiveness of annual vaccines that rely on a match with circulating influenza strains. Thus, there is a continued need for new, efficacious vaccines conferring cross-clade protection to avoid the need for biannual reformulation of seasonal influenza vaccines. Recombinant virus-vectored vaccines are an appealing alternative to classical inactivated vaccines because virus vectors enable native expression of influenza antigens, even from virulent influenza viruses, while expressed in the context of the vector that can improve immunogenicity. In addition, a vectored vaccine often enables delivery of the vaccine to sites of inductive immunity such as the respiratory tract enabling protection from influenza virus infection. Moreover, the ability to readily manipulate virus vectors to produce novel influenza vaccines may provide the quickest path toward a universal vaccine protecting against all influenza viruses. This review will discuss experimental virus-vectored vaccines for use in humans, comparing them to licensed vaccines and the hurdles faced for licensure of these next-generation influenza virus vaccines. PMID:25105278

  15. Leucine-rich repeat-containing G protein-coupled receptor 4 facilitates vesicular stomatitis virus infection by binding vesicular stomatitis virus glycoprotein.

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    Zhang, Na; Huang, Hongjun; Tan, Binghe; Wei, Yinglei; Xiong, Qingqing; Yan, Yan; Hou, Lili; Wu, Nannan; Siwko, Stefan; Cimarelli, Andrea; Xu, Jianrong; Han, Honghui; Qian, Min; Liu, Mingyao; Du, Bing

    2017-10-06

    Vesicular stomatitis virus (VSV) and rabies and Chandipura viruses belong to the Rhabdovirus family. VSV is a common laboratory virus to study viral evolution and host immune responses to viral infection, and recombinant VSV-based vectors have been widely used for viral oncolysis, vaccination, and gene therapy. Although the tropism of VSV is broad, and its envelope glycoprotein G is often used for pseudotyping other viruses, the host cellular components involved in VSV infection remain unclear. Here, we demonstrate that the host protein leucine-rich repeat-containing G protein-coupled receptor 4 (Lgr4) is essential for VSV and VSV-G pseudotyped lentivirus (VSVG-LV) to infect susceptible cells. Accordingly, Lgr4-deficient mice had dramatically decreased VSV levels in the olfactory bulb. Furthermore, Lgr4 knockdown in RAW 264.7 cells also significantly suppressed VSV infection, and Lgr4 overexpression in RAW 264.7 cells enhanced VSV infection. Interestingly, only VSV infection relied on Lgr4, whereas infections with Newcastle disease virus, influenza A virus (A/WSN/33), and herpes simplex virus were unaffected by Lgr4 status. Of note, assays of virus entry, cell ELISA, immunoprecipitation, and surface plasmon resonance indicated that VSV bound susceptible cells via the Lgr4 extracellular domain. Pretreating cells with an Lgr4 antibody, soluble LGR4 extracellular domain, or R-spondin 1 blocked VSV infection by competitively inhibiting VSV binding to Lgr4. Taken together, the identification of Lgr4 as a VSV-specific host factor provides important insights into understanding VSV entry and its pathogenesis and lays the foundation for VSV-based gene therapy and viral oncolytic therapeutics. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Protective efficacy of a recombinant Newcastle disease virus expressing glycoprotein of vesicular stomatitis virus in mice

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    Zhang, Minmin; Ge, Jinying; Li, Xiaofang; Chen, Weiye; Wang, Xijun; Wen, Zhiyuan; Bu, Zhigao

    2016-01-01

    Background Vesicular stomatitis virus (VSV) causes severe losses to the animal husbandry industry. In this study, a recombinant Newcastle disease virus (NDV) expressing the glycoprotein (G) of VSV (rL-VSV-G) was constructed and its pathogenicity and immune protective efficacy in mouse were evaluated. Results In pathogenicity evaluation test, the analysis of the viral distribution in mouse organs and body weight change showed that rL-VSV-G was safe in mice. In immune protection assay, the reco...

  17. Infection of Melanoplus sanguinipes grasshoppers following ingestion of rangeland plant species harboring vesicular stomatitis virus.

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    Drolet, Barbara S; Stuart, Melissa A; Derner, Justin D

    2009-05-01

    Knowledge of the many mechanisms of vesicular stomatitis virus (VSV) transmission is critical for understanding of the epidemiology of sporadic disease outbreaks in the western United States. Migratory grasshoppers [Melanoplus sanguinipes (Fabricius)] have been implicated as reservoirs and mechanical vectors of VSV. The grasshopper-cattle-grasshopper transmission cycle is based on the assumptions that (i) virus shed from clinically infected animals would contaminate pasture plants and remain infectious on plant surfaces and (ii) grasshoppers would become infected by eating the virus-contaminated plants. Our objectives were to determine the stability of VSV on common plant species of U.S. Northern Plains rangelands and to assess the potential of these plant species as a source of virus for grasshoppers. Fourteen plant species were exposed to VSV and assayed for infectious virus over time (0 to 24 h). The frequency of viable virus recovery at 24 h postexposure was as high as 73%. The two most common plant species in Northern Plains rangelands (western wheatgrass [Pascopyrum smithii] and needle and thread [Hesperostipa comata]) were fed to groups of grasshoppers. At 3 weeks postfeeding, the grasshopper infection rate was 44 to 50%. Exposure of VSV to a commonly used grasshopper pesticide resulted in complete viral inactivation. This is the first report demonstrating the stability of VSV on rangeland plant surfaces, and it suggests that a significant window of opportunity exists for grasshoppers to ingest VSV from contaminated plants. The use of grasshopper pesticides on pastures would decrease the incidence of a virus-amplifying mechanical vector and might also decontaminate pastures, thereby decreasing the inter- and intraherd spread of VSV.

  18. Pathogenesis of experimental vesicular stomatitis virus (New Jersey serotype) infection in the deer mouse (Peromyscus maniculatus).

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    Cornish, T E; Stallknecht, D E; Brown, C C; Seal, B S; Howerth, E W

    2001-07-01

    The pathogenesis of vesicular stomatitis virus (VSV) infection has not been investigated previously in native New World rodents that may have a role in the epidemiology of the disease. In the present study, 45 juvenile and 80 adult deer mice (Peromyscus maniculatus) were inoculated intranasally with VSV New Jersey serotype (VSV-NJ) and examined sequentially over a 7-day period. Virus was detected by means of immunohistochemistry and in situ hybridization in all tissues containing histologic lesions. Viral antigen and mRNA were observed initially in olfactory epithelium neurons, followed by olfactory bulbs and more caudal olfactory pathways in the brain. Virus also was detected throughout the ventricular system in the brain and central canal of the spinal cord. These results support both viral retrograde transneuronal transport and viral spread within the ventricular system. Other tissues containing viral antigen included airway epithelium and macrophages in the lungs, cardiac myocytes, and macrophages in cervical lymph nodes. In a second experiment, 15 adult, 20 juvenile, and 16 nestling deer mice were inoculated intradermally with VSV-NJ. Adults were refractory to infection by this route; however, nestlings and juveniles developed disseminated central nervous system infections. Viral antigen also was detected in cardiac myocytes and lymph node macrophages in these animals. Viremia was detected by virus isolation in 35/72 (49%) intranasally inoculated juvenile and adult mice and in 17/36 (47%) intradermally inoculated nestlings and juveniles from day 1 to day 3 postinoculation. The documentation of viremia in these animals suggests that they may have a role in the epidemiology of vector-borne vesicular stomatitis.

  19. Vesicular stomatitis virus-based ebola vaccine is well-tolerated and protects immunocompromised nonhuman primates.

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    Thomas W Geisbert

    2008-11-01

    Full Text Available Ebola virus (EBOV is a significant human pathogen that presents a public health concern as an emerging/re-emerging virus and as a potential biological weapon. Substantial progress has been made over the last decade in developing candidate preventive vaccines that can protect nonhuman primates against EBOV. Among these prospects, a vaccine based on recombinant vesicular stomatitis virus (VSV is particularly robust, as it can also confer protection when administered as a postexposure treatment. A concern that has been raised regarding the replication-competent VSV vectors that express EBOV glycoproteins is how these vectors would be tolerated by individuals with altered or compromised immune systems such as patients infected with HIV. This is especially important as all EBOV outbreaks to date have occurred in areas of Central and Western Africa with high HIV incidence rates in the population. In order to address this concern, we evaluated the safety of the recombinant VSV vector expressing the Zaire ebolavirus glycoprotein (VSVDeltaG/ZEBOVGP in six rhesus macaques infected with simian-human immunodeficiency virus (SHIV. All six animals showed no evidence of illness associated with the VSVDeltaG/ZEBOVGP vaccine, suggesting that this vaccine may be safe in immunocompromised populations. While one goal of the study was to evaluate the safety of the candidate vaccine platform, it was also of interest to determine if altered immune status would affect vaccine efficacy. The vaccine protected 4 of 6 SHIV-infected macaques from death following ZEBOV challenge. Evaluation of CD4+ T cells in all animals showed that the animals that succumbed to lethal ZEBOV challenge had the lowest CD4+ counts, suggesting that CD4+ T cells may play a role in mediating protection against ZEBOV.

  20. [Mechanism of resistance of a clone isolated from L cells chronically infected with the virus of vesicular stomatitis].

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    Sovetova, G P; Zhdanov, V M

    1976-01-01

    The results of the study on antiviral immunity acquired by L cells to vesicular stomatitis virus are presented. We failed to detect the presence of the indicator virus in the resistant culture by means of virological, electron microscopic or cytochemical methods. Molecular hybrization experiments demonstrated the lack in the nuclear DNA of sequences homologous to vesicular stomatitis virus RNA.

  1. In Vitro and In Vivo Attenuation of Vesicular Stomatitis Virus (VSV by Phosphoprotein Deletion.

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    Phonphimon Wongthida

    Full Text Available Vesicular stomatitis virus (VSV is highly immunogenic and able to stimulate both innate and adaptive immune responses. However, its ability to induce adverse effects has held back the use of VSV as a potential vaccine vector. In this study we developed VSV-ΔP, a safe yet potent replication-defective recombinant VSV in which the phosphoprotein (P gene was deleted. VSV-ΔP replicated only in supporting cells expressing P (BHK-P cells and at levels more than 2 logs lower than VSV. In vivo studies indicated that the moderate replication of VSV-ΔP in vitro was associated with the attenuation of this virus in the mouse model, whereas mice intracranially injected with VSV succumbed to neurotoxicity. Furthermore, we constructed VSV and VSV-ΔP expressing a variety of antigens including hemagglutinin-neuraminidase (HN from Newcastle disease virus (NDV, hemagglutinin (HA from either a 2009 H1N1 pandemic influenza virus (pdm/09 or the avian H7N9. VSV and VSV-ΔP incorporated the foreign antigens on their surface resulting in induction of robust neutralizing antibody, serum IgG, and hemagglutination inhibition (HAI titers against their corresponding viruses. These results indicated that VSV with P gene deletion was attenuated in vitro and in vivo, and possibly expressed the foreign antigen on its surface. Therefore, the P gene-deletion strategy may offer a potentially useful and safer approach for attenuating negative-sense RNA viruses which use phosphoprotein as a cofactor for viral replication.

  2. Cellular proteins associated with the interior and exterior of vesicular stomatitis virus virions.

    Directory of Open Access Journals (Sweden)

    Megan Moerdyk-Schauwecker

    Full Text Available Virus particles (virions often contain not only virus-encoded but also host-encoded proteins. Some of these host proteins are enclosed within the virion structure, while others, in the case of enveloped viruses, are embedded in the host-derived membrane. While many of these host protein incorporations are likely accidental, some may play a role in virus infectivity, replication and/or immunoreactivity in the next host. Host protein incorporations may be especially important in therapeutic applications where large numbers of virus particles are administered. Vesicular stomatitis virus (VSV is the prototypic rhabdovirus and a candidate vaccine, gene therapy and oncolytic vector. Using mass spectrometry, we previously examined cell type dependent host protein content of VSV virions using intact ("whole" virions purified from three cell lines originating from different species. Here we aimed to determine the localization of host proteins within the VSV virions by analyzing: i whole VSV virions; and ii whole VSV virions treated with Proteinase K to remove all proteins outside the viral envelope. A total of 257 proteins were identified, with 181 identified in whole virions and 183 identified in Proteinase K treated virions. Most of these proteins have not been previously shown to be associated with VSV. Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization. Using western blotting, the presence of several host proteins, including some not previously shown in association with VSV (such as Yes1, Prl1 and Ddx3y, was confirmed and their relative quantities in various virion fractions determined. Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.

  3. Mechanical transmission of vesicular stomatitis New Jersey virus by Simulium vittatum (Diptera: Simuliidae) to domestic swine (Sus scrofa).

    Science.gov (United States)

    Smith, Paul F; Howerth, Elizabeth W; Carter, Deborah; Gray, Elmer W; Noblet, Raymond; Mead, Daniel G

    2009-11-01

    Biting flies have been suggested as mechanical vectors of vesicular stomatitis New Jersey Virus (family Rhabdoviridae, genus Vesiculovirus, VSNJV) in livestock populations during epidemic outbreaks in the western United States. We conducted a proof-of-concept study to determine whether biting flies could mechanically transmit VSNJV to livestock by using a black fly, Simulium vittatum Zetterstedt (Diptera: Simuliidae), domestic swine, Sus scrofa L., model. Black flies mechanically transmitted VSNJV to a naive host after interrupted feeding on a vesicular lesion on a previously infected host. Transmission resulted in clinical disease in the naïve host. This is the first demonstration of mechanical transmission of VSNJV to livestock by insects.

  4. Stimulation of Vesicular Stomatitis Virus in vitro RNA Synthesis by Microtubule-Associated Proteins

    Science.gov (United States)

    Hill, Virginia M.; Harmon, Shirley A.; Summers, Donald F.

    1986-08-01

    Microtubule-associated proteins purified from bovine brains stimulated the in vitro transcription and replication reactions of vesicular stomatitis virus. The products of these reactions were intact messenger or genome-sized RNA species. A preparation from HeLa cells containing tubulin and microtubule-associated proteins also stimulated vesicular stomatitis virus transcription in vitro. This observation is in accord with previous studies, which suggested that a host cell factor was involved with the function of the vesicular stomatitis virus RNA polymerase, and others that indicated that several animal viruses displayed an association with host cell cytoskeletal elements during their replication cycles. We show evidence in this report of a host cell protein that seems to have a functional role in interacting with the virion polymerase.

  5. Retro-transduction by virus pseudotyped with glycoprotein of vesicular stomatitis virus

    International Nuclear Information System (INIS)

    Ohishi, Masahisa; Shioda, Tatsuo; Sakuragi, Jun-ichi

    2007-01-01

    A virus pseudotyped with glycoprotein of vesicular stomatitis virus (VSV-G) can enter various cell types at a relatively high titer. We observed that the amount of viral antigen from VSV-G pseudotyped human immunodeficiency virus type 1 (HIV-1) producing cells was much higher than that from their non-pseudotyped counterparts. This enhanced viral antigen production was not observed when we used HIV-1 pol mutant, viral enzyme inhibitors, HIV Env protein, or VSV-G fusion defective mutants. The transfection experiment using GFP-expressing virus showed time-dependent expansion of GFP-positive cells and viral DNA integration. These results suggested that the increase in viral antigen yield was caused by the release of a progeny virus following retro-transduction by the pseudotyped virus of the cells within the transfected cell culture. The infectivity as well as the amount of VSV-G on virus particles per unit of viral antigen was significantly different before and after the onset of the yield enhancement. This suggests that results of infection assays of the virus pseudotyped with VSV-G may be affected by the occurrence of such enhancement. This means that, while pseudotyping with VSV-G is a simple and effective method, this procedure should be carefully considered when the virus is produced for infectivity assays

  6. The lipidomes of vesicular stomatitis virus, semliki forest virus, and the host plasma membrane analyzed by quantitative shotgun mass spectrometry

    DEFF Research Database (Denmark)

    Kalvodova, Lucie; Sampaio, Julio L; Cordo, Sandra

    2009-01-01

    Although enveloped virus assembly in the host cell is a crucial step in the virus life cycle, it remains poorly understood. One issue is how viruses include lipids in their membranes during budding from infected host cells. To analyze this issue, we took advantage of the fact that baby hamster...... kidney cells can be infected by two different viruses, namely, vesicular stomatitis virus and Semliki Forest virus, from the Rhabdoviridae and Togaviridae families, respectively. We purified the host plasma membrane and the two different viruses after exit from the host cells and analyzed the lipid...

  7. Protective efficacy of a recombinant Newcastle disease virus expressing glycoprotein of vesicular stomatitis virus in mice.

    Science.gov (United States)

    Zhang, Minmin; Ge, Jinying; Li, Xiaofang; Chen, Weiye; Wang, Xijun; Wen, Zhiyuan; Bu, Zhigao

    2016-02-24

    Vesicular stomatitis virus (VSV) causes severe losses to the animal husbandry industry. In this study, a recombinant Newcastle disease virus (NDV) expressing the glycoprotein (G) of VSV (rL-VSV-G) was constructed and its pathogenicity and immune protective efficacy in mouse were evaluated. In pathogenicity evaluation test, the analysis of the viral distribution in mouse organs and body weight change showed that rL-VSV-G was safe in mice. In immune protection assay, the recombinant rL-VSV-G triggered a high titer of neutralizing antibodies against VSV. After challenge, the wild-type (wt) VSV viral load in mouse organs was lower in rL-VSV-G group than that in rLaSota groups. wt VSV was not detected in the blood, liver, or kidneys of mice, whereas it was found in these tissues in control groups. The mice body weight had no significant change after challenge in the rL-VSV-G group. Additionally, suckling mice produced from female mice immunized with rL-VSV-G were partially protected from wt VSV challenge. These results demonstrated that rL-VSV-G may be a suitable candidate vaccine against vesicular stomatitis (VS).

  8. Vesicular Stomatitis Virus Infection Promotes Immune Evasion by Preventing NKG2D-Ligand Surface Expression

    DEFF Research Database (Denmark)

    Jensen, Helle; Andresen, Lars; Nielsen, Jens

    2011-01-01

    Vesicular stomatitis virus (VSV) has recently gained attention for its oncolytic ability in cancer treatment. Initially, we hypothesized that VSV infection could increase immune recognition of cancer cells through induction of the immune stimulatory NKG2D-ligands. Here we show that VSV infection ...

  9. Vesicular stomatitis virus with the rabies virus glycoprotein directs retrograde transsynaptic transport among neurons in vivo

    Directory of Open Access Journals (Sweden)

    Kevin T Beier

    2013-02-01

    Full Text Available Defining the connections among neurons is critical to our understanding of the structure and function of the nervous system. Recombinant viruses engineered to transmit across synapses provide a powerful approach for the dissection of neuronal circuitry in vivo. We recently demonstrated that recombinant vesicular stomatitis virus (VSV can be endowed with anterograde or retrograde synaptic tracing ability by providing the virus with different glycoproteins. Here we extend the characterization of the transmission and gene expression of VSV with the rabies virus glycoprotein (RABV-G, and provide examples of its activity relative to the anterograde tracer form of rVSV. rVSV with RABV-G was found to drive strong expression of transgenes and to spread rapidly from neuron to neuron in only a retrograde manner. Depending upon how the RABV-G was delivered, VSV served as a polysynaptic or monosynaptic tracer, or was able to define projections through axonal uptake and retrograde transport. In animals co-infected with rVSV in its anterograde form, rVSV with RABV-G could be used to begin to characterize the similarities and differences in connections to a given area. rVSV with RABV-G provides a flexible, rapid, and versatile tracing tool that complements the previously described VSV-based anterograde transsynaptic tracer.

  10. Vesicular stomatitis virus with the rabies virus glycoprotein directs retrograde transsynaptic transport among neurons in vivo.

    Science.gov (United States)

    Beier, Kevin T; Saunders, Arpiar B; Oldenburg, Ian A; Sabatini, Bernardo L; Cepko, Constance L

    2013-01-01

    Defining the connections among neurons is critical to our understanding of the structure and function of the nervous system. Recombinant viruses engineered to transmit across synapses provide a powerful approach for the dissection of neuronal circuitry in vivo. We recently demonstrated that recombinant vesicular stomatitis virus (VSV) can be endowed with anterograde or retrograde transsynaptic tracing ability by providing the virus with different glycoproteins. Here we extend the characterization of the transmission and gene expression of recombinant VSV (rVSV) with the rabies virus glycoprotein (RABV-G), and provide examples of its activity relative to the anterograde transsynaptic tracer form of rVSV. rVSV with RABV-G was found to drive strong expression of transgenes and to spread rapidly from neuron to neuron in only a retrograde manner. Depending upon how the RABV-G was delivered, VSV served as a polysynaptic or monosynaptic tracer, or was able to define projections through axonal uptake and retrograde transport. In animals co-infected with rVSV in its anterograde form, rVSV with RABV-G could be used to begin to characterize the similarities and differences in connections to different areas. rVSV with RABV-G provides a flexible, rapid, and versatile tracing tool that complements the previously described VSV-based anterograde transsynaptic tracer.

  11. Chikungunya Virus Vaccines: Viral Vector-Based Approaches.

    Science.gov (United States)

    Ramsauer, Katrin; Tangy, Frédéric

    2016-12-15

    In 2013, a major chikungunya virus (CHIKV) epidemic reached the Americas. In the past 2 years, >1.7 million people have been infected. In light of the current epidemic, with millions of people in North and South America at risk, efforts to rapidly develop effective vaccines have increased. Here, we focus on CHIKV vaccines that use viral-vector technologies. This group of vaccine candidates shares an ability to potently induce humoral and cellular immune responses by use of highly attenuated and safe vaccine backbones. So far, well-described vectors such as modified vaccinia virus Ankara, complex adenovirus, vesicular stomatitis virus, alphavirus-based chimeras, and measles vaccine Schwarz strain (MV/Schw) have been described as potential vaccines. We summarize here the recent data on these experimental vaccines, with a focus on the preclinical and clinical activities on the MV/Schw-based candidate, which is the first CHIKV-vectored vaccine that has completed a clinical trial. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  12. Host predilection and transmissibility of vesicular stomatitis New Jersey virus strains in domestic cattle (Bos taurus and swine (Sus scrofa

    Directory of Open Access Journals (Sweden)

    Smith Paul F

    2012-10-01

    Full Text Available Abstract Background Epidemiologic data collected during epidemics in the western United States combined with limited experimental studies involving swine and cattle suggest that host predilection of epidemic vesicular stomatitis New Jersey virus (VSNJV strains results in variations in clinical response, extent and duration of virus shedding and transmissibility following infection in different hosts. Laboratory challenge of livestock with heterologous VSNJV strains to investigate potential viral predilections for these hosts has not been thoroughly investigated. In separate trials, homologous VSNJV strains (NJ82COB and NJ82AZB, and heterologous strains (NJ06WYE and NJOSF [Ossabaw Island, sand fly] were inoculated into cattle via infected black fly bite. NJ82AZB and NJ06WYE were similarly inoculated into swine. Results Clinical scores among viruses infecting cattle were significantly different and indicated that infection with a homologous virus resulted in more severe clinical presentation and greater extent and duration of viral shedding. No differences in clinical severity or extent and duration of viral shedding were detected in swine. Conclusions Differences in clinical presentation and extent and duration of viral shedding may have direct impacts on viral spread during epidemics. Viral transmission via animal-to-animal contact and insect vectored transmission are likely to occur at higher rates when affected animals are presenting severe clinical signs and shedding high concentrations of virus. More virulent viral strains resulting in more severe disease in livestock hosts are expected to spread more rapidly and greater distances during epidemics than those causing mild or inapparent signs.

  13. Host predilection and transmissibility of vesicular stomatitis New Jersey virus strains in domestic cattle (Bos taurus) and swine (Sus scrofa).

    Science.gov (United States)

    Smith, Paul F; Howerth, Elizabeth W; Carter, Deborah; Gray, Elmer W; Noblet, Raymond; Berghaus, Roy D; Stallknecht, David E; Mead, Daniel G

    2012-10-03

    Epidemiologic data collected during epidemics in the western United States combined with limited experimental studies involving swine and cattle suggest that host predilection of epidemic vesicular stomatitis New Jersey virus (VSNJV) strains results in variations in clinical response, extent and duration of virus shedding and transmissibility following infection in different hosts. Laboratory challenge of livestock with heterologous VSNJV strains to investigate potential viral predilections for these hosts has not been thoroughly investigated. In separate trials, homologous VSNJV strains (NJ82COB and NJ82AZB), and heterologous strains (NJ06WYE and NJOSF [Ossabaw Island, sand fly]) were inoculated into cattle via infected black fly bite. NJ82AZB and NJ06WYE were similarly inoculated into swine. Clinical scores among viruses infecting cattle were significantly different and indicated that infection with a homologous virus resulted in more severe clinical presentation and greater extent and duration of viral shedding. No differences in clinical severity or extent and duration of viral shedding were detected in swine. Differences in clinical presentation and extent and duration of viral shedding may have direct impacts on viral spread during epidemics. Viral transmission via animal-to-animal contact and insect vectored transmission are likely to occur at higher rates when affected animals are presenting severe clinical signs and shedding high concentrations of virus. More virulent viral strains resulting in more severe disease in livestock hosts are expected to spread more rapidly and greater distances during epidemics than those causing mild or inapparent signs.

  14. Characterization of Vesicular Stomatitis Virus Recombinants That Express and Incorporate High Levels of Hepatitis C Virus Glycoproteins

    OpenAIRE

    Buonocore, Linda; Blight, Keril J.; Rice, Charles M.; Rose, John K.

    2002-01-01

    We generated recombinant vesicular stomatitis viruses (VSV) expressing genes encoding hybrid proteins consisting of the extracellular domains of hepatitis C virus (HCV) glycoproteins fused at different positions to the transmembrane and cytoplasmic domains of the VSV G glycoprotein (E1G and E2G). We show that these chimeric proteins are transported to the cell surface and incorporated into VSV virions efficiently. We also generated VSV recombinants in which the gene encoding the VSV G protein...

  15. Vesicular Stomatitis Virus Pseudotyped with Ebola Virus Glycoprotein Serves as a Highly Protective, Non-infectious Vaccine Against Ebola Virus Challenge

    Science.gov (United States)

    2016-07-01

    1 Vesicular stomatitis virus pseudotyped with Ebola virus glycoprotein serves as 1 a highly protective, non-infectious vaccine against Ebola virus...Iowa City, IA 52242 USA 16 1 319 335 8021 17 wendy-maury@uiowa.edu 18 19 Running title: Pseudovirion vaccine against Ebola virus 20...measures to combat the associated disease. Since, no vaccines or antivirals are currently FDA 24 approved, we sought to assess protection conferred by an

  16. A Polyamide Inhibits Replication of Vesicular Stomatitis Virus by Targeting RNA in the Nucleocapsid

    Energy Technology Data Exchange (ETDEWEB)

    Gumpper, Ryan H.; Li, Weike; Castañeda, Carlos H.; Scuderi, M. José; Bashkin, James K.; Luo, Ming; Dutch, Rebecca Ellis

    2018-02-07

    Polyamides have been shown to bind double-stranded DNA by complementing the curvature of the minor groove and forming various hydrogen bonds with DNA. Several polyamide molecules have been found to have potent antiviral activities against papillomavirus, a double-stranded DNA virus. By analogy, we reason that polyamides may also interact with the structured RNA bound in the nucleocapsid of a negative-strand RNA virus. Vesicular stomatitis virus (VSV) was selected as a prototype virus to test this possibility since its genomic RNA encapsidated in the nucleocapsid forms a structure resembling one strand of an A-form RNA duplex. One polyamide molecule, UMSL1011, was found to inhibit infection of VSV. To confirm that the polyamide targeted the nucleocapsid, a nucleocapsid-like particle (NLP) was incubated with UMSL1011. The encapsidated RNA in the polyamide-treated NLP was protected from thermo-release and digestion by RNase A. UMSL1011 also inhibits viral RNA synthesis in the intracellular activity assay for the viral RNA-dependent RNA polymerase. The crystal structure revealed that UMSL1011 binds the structured RNA in the nucleocapsid. The conclusion of our studies is that the RNA in the nucleocapsid is a viable antiviral target of polyamides. Since the RNA structure in the nucleocapsid is similar in all negative-strand RNA viruses, polyamides may be optimized to target the specific RNA genome of a negative-strand RNA virus, such as respiratory syncytial virus and Ebola virus.

    IMPORTANCENegative-strand RNA viruses (NSVs) include several life-threatening pathogens, such as rabies virus, respiratory syncytial virus, and Ebola virus. There are no effective antiviral drugs against these viruses. Polyamides offer an exceptional opportunity because they may be optimized to target each NSV. Our studies on vesicular stomatitis virus, an NSV, demonstrated that a polyamide molecule could specifically target the viral RNA in the nucleocapsid and inhibit

  17. Analysis of the RNA species isolated from defective particles of vesicular stomatitis virus.

    Science.gov (United States)

    Adler, R; Banerjee, A K

    1976-10-01

    Serial high multiplicity passage of a cloned stock of vesicular stomatitis virus was found to generate defective interfering particles containing three size classes of RNA, with sedimentaiton coefficients of 31 S, 23 S and 19 S. The 31 S and 23 S RNA species were found to be complementary to both the 12 to 18 S and 31 S size classes of VSV mRNAs. The 19 S class of RNA was found to be partially base-paired. All three RNA species were found to contain ppAp at their 5' termini.

  18. Quantification of Lyssavirus-Neutralizing Antibodies Using Vesicular Stomatitis Virus Pseudotype Particles

    Directory of Open Access Journals (Sweden)

    Sarah Moeschler

    2016-09-01

    Full Text Available Rabies is a highly fatal zoonotic disease which is primarily caused by rabies virus (RABV although other members of the genus Lyssavirus can cause rabies as well. As yet, 14 serologically and genetically diverse lyssaviruses have been identified, mostly in bats. To assess the quality of rabies vaccines and immunoglobulin preparations, virus neutralization tests with live RABV are performed in accordance with enhanced biosafety standards. In the present work, a novel neutralization test is presented which takes advantage of a modified vesicular stomatitis virus (VSV from which the glycoprotein G gene has been deleted and replaced by reporter genes. This single-cycle virus was trans-complemented with RABV envelope glycoprotein. Neutralization of this pseudotype virus with RABV reference serum or immune sera from vaccinated mice showed a strong correlation with the rapid fluorescent focus inhibition test (RFFIT. Importantly, pseudotype viruses containing the envelope glycoproteins of other lyssaviruses were neutralized by reference serum to a significantly lesser extent or were not neutralized at all. Taken together, a pseudotype virus system has been successfully developed which allows the safe, fast, and sensitive detection of neutralizing antibodies directed against different lyssaviruses.

  19. Recombinant vesicular stomatitis virus-based dengue-2 vaccine candidate induces humoral response and protects mice against lethal infection.

    Science.gov (United States)

    Lauretti, Flavio; Chattopadhyay, Anasuya; de Oliveira França, Rafael Freitas; Castro-Jorge, Luiza; Rose, John; Fonseca, Benedito A L da

    2016-09-01

    Dengue is the most important arbovirus disease throughout the world and it is responsible for more than 500,000 dengue hemorrhagic cases and 22,000 deaths every year. One vaccine was recently licensed for human use in Brazil, Mexico and Philippines and although at least seven candidates have been in clinical trials the results of the most developed CYD vaccine have demonstrated immunization problems, such as uneven protection and interference between serotypes. We constructed a vaccine candidate based on vesicular stomatitis virus (VSV) expression of pre-membrane (prM) and envelope (E) proteins of dengue-2 virus (DENV-2) and tested it in mice to evaluate immunogenicity and protection against DENV-2 infection. VSV has been successfully used as vaccine vectors for several viruses to induce strong humoral and cellular immune responses. The VSV-DENV-2 recombinant was constructed by inserting the DENV-2 structural proteins into a VSV plasmid DNA for recombinant VSV-DENV-2 recovery. Infectious recombinant VSV viruses were plaque purified and prM and E expression were confirmed by immunofluorescence and radiolabeling of proteins of infected cells. Forty Balb/C mice were inoculated through subcutaneous (s.c.) route with VSV-DENV-2 vaccine in a two doses schedule 15 d apart and 29 d after first inoculation, sera were collected and the mice were challenged with 50 lethal doses (LD50) of a neurovirulent DENV-2. The VSV-DENV-2 induced anti-DENV-2 antibodies and protected animals in the challenge experiment comparable to DENV-2 immunization control group. We conclude that VSV is a promising platform to test as a DENV vaccine and perhaps against others Flaviviridae.

  20. Immunogenicity and efficacy of immunodeficiency virus-like particles pseudotyped with the G protein of vesicular stomatitis virus

    International Nuclear Information System (INIS)

    Kuate, Seraphin; Stahl-Hennig, Christiane; Stoiber, Heribert; Nchinda, Godwin; Floto, Anja; Franz, Monika; Sauermann, Ulrike; Bredl, Simon; Deml, Ludwig; Ignatius, Ralf; Norley, Steve; Racz, Paul; Tenner-Racz, Klara; Steinman, Ralph M.; Wagner, Ralf; Uberla, Klaus

    2006-01-01

    Vaccination with exogenous antigens such as recombinant viral proteins, immunodeficiency virus-derived whole inactivated virus particles, or virus-like particles (VLP) has generally failed to provide sufficient protection in animal models for AIDS. Pseudotyping VLPs with the vesicular stomatitis virus G protein (VSV-G), which is known to mediate entry into dendritic cells, might allow more efficient stimulation of immune responses. Therefore, we pseudotyped noninfectious immunodeficiency virus-like particles with VSV-G and carried out a preliminary screen of their immunogenicity and vaccination efficacy. Incorporation of VSV-G into HIV-1 VLPs led to hundred-fold higher antibody titers to HIV-1 Gag and enhancement of T cell responses in mice. Repeated vaccination of rhesus monkeys for 65 weeks with VSV-G pseudotyped simian immunodeficiency virus (SIV)-like particles (VLP[G]) provided initial evidence for efficient suppression of viral load after mucosal challenge with the SIVmac239 virus. Challenge of monkeys after a 28 week vaccination regimen with VLP[G] led to a reduction in peak viremia, but persistent suppression of viral load was not achieved. Due to limitations in the number of animals available for this study, improved efficacy of VSV-G pseudotyped VLPs in nonhuman primates could not be demonstrated. However, mouse experiments revealed that pseudotyping of VLPs with fusion-competent VSV-G clearly improves their immunogenicity. Additional strategies, particularly adjuvants, should be considered to provide greater protection against a challenge with pathogenic immunodeficiency virus

  1. Viruses vector control proposal: genus Aedes emphasis

    Directory of Open Access Journals (Sweden)

    Nelson Nogueira Reis

    2017-07-01

    Full Text Available The dengue fever is a major public health problem in the world. In Brazil, in 2015, there were 1,534,932 cases, being 20,320 cases of severe form, and 811 deaths related to this disease. The distribution of Aedes aegypti, the vector, is extensive. Recently, Zika and Chikungunya viruses had arisen, sharing the same vector as dengue and became a huge public health issue. Without specific treatment, it is urgently required as an effective vector control. This article is focused on reviewing vector control strategies, their effectiveness, viability and economical impact. Among all, the Sterile Insect Technique is highlighted as the best option to be adopted in Brazil, once it is largely effectively used in the USA and Mexico for plagues related to agribusiness.

  2. Interactions of macrophages with probiotic bacteria lead to increased antiviral response against vesicular stomatitis virus

    DEFF Research Database (Denmark)

    Ivec, Martin; Botic, Tanja; Koren, Srecko

    2007-01-01

    and by producing chemokines and immunoregulatory cytokines that enable the adaptive immune response to recognize infected cells and perform antiviral effector functions. Probiotics, as a part of the normal gut intestinal flora, are important in supporting a functional yet balanced immune system. Improving our...... understanding of their role in the activation of macrophages and their stimulation of proinflammatory cytokine production in early viral infection was the main goal of this study. Our in vitro model study showed that probiotic bacteria, either from the species Lactobacillus or Bifidobacteria have the ability...... dehydrogenases activity could be implied as the first indicator of potential inhibitory effects of the probiotics on virus replication. The interactions between probiotic bacteria, macrophages and vesicular stomatitis virus (VSV), markedly depended on the bacterial strain studied....

  3. In vivo Biodistribution of a Highly Attenuated Recombinant Vesicular Stomatitis Virus Expressing HIV-1 Gag Following Intramuscular, Intranasal, or Intravenous Inoculation

    Science.gov (United States)

    Johnson, J. Erik; Coleman, John W.; Kalyan, Narender K.; Calderon, Priscilla; Wright, Kevin J.; Obregon, Jennifer; Ogin-Wilson, Eleanor; Natuk, Robert J.; Clarke, David K.; Udem, Stephen A.; Cooper, David; Hendry, R. Michael

    2009-01-01

    Recombinant vesicular stomatitis viruses (rVSVs) are being developed as potential HIV-1 vaccine candidates. To characterize the in vivo replication and dissemination of rVSV vectors in mice, high doses of a highly attenuated vector expressing HIV-1 Gag, rVSVIN- N4CT9-Gag1, and a prototypic reference virus, rVSVIN-HIVGag5, were delivered intramuscularly (IM), intranasally (IN), or intravenously (IV). We used quantitative, real-time RT-PCR (Q-PCR) and standard plaque assays to measure the temporal dissemination of these viruses to various tissues. Following IM inoculation, both viruses were detected primarily at the injection site as well as in draining lymph nodes; neither virus induced significant weight loss, pathologic signs, or evidence of neuroinvasion. In contrast, following IN inoculation, the prototypic virus was detected in all tissues tested and caused significant weight loss leading to death. IN administration of rVSVIN- N4CT9-Gag1 resulted in detection in numerous tissues (brain, lung, nasal turbinates, and lymph nodes) albeit in significantly reduced levels, which caused little or no weight loss nor any mortality. Following IV inoculation, both prototypic and attenuated viruses were detected by Q-PCR in all tissues tested. In contrast to the prototype, rVSVIN-N4CT9-Gag1 viral loads were significantly lower in all organs tested, and no infectious virus was detected in the brain following IV inoculation, despite the presence of viral RNA. These studies demonstrated significant differences in the biodistribution patterns of and the associated pathogenicity engendered by the prototypic and attenuated vectors in a highly susceptible host. PMID:19428903

  4. Trojan horse lymphocytes: a vesicular stomatitis virus-specific T-cell clone lyses target cells by carrying virus.

    OpenAIRE

    Hom, R C; Soman, G; Finberg, R

    1989-01-01

    We have isolated a vesicular stomatitis virus (VSV)-specific CD4+ CD8- murine T-cell clone. This clone proliferates only in response to VSV and lyses infected tumor cells bearing class II major histocompatibility antigens in short-term chromium release assays. In addition, the cell has VSV antigens on its surface and is capable of killing uninfected tumor cells without major histocompatibility antigen restriction in a 2-day assay. This latter cytolytic activity is eliminated by anti-VSV antib...

  5. Acute reactogenicity after intramuscular immunization with recombinant vesicular stomatitis virus is linked to production of IL-1β.

    Directory of Open Access Journals (Sweden)

    Kathleen Athearn

    Full Text Available Vaccines based on live viruses are attractive because they are immunogenic, cost-effective, and can be delivered by multiple routes. However, live virus vaccines also cause reactogenic side effects such as fever, myalgia, and injection site pain that have reduced their acceptance in the clinic. Several recent studies have linked vaccine-induced reactogenic side effects to production of the pro-inflammatory cytokine interleukin-1β (IL-1β in humans. Our objective was therefore to determine whether IL-1β contributed to pathology after immunization with recombinant vesicular stomatitis virus (rVSV vaccine vectors, and if so, to identify strategies by which IL-1β mediated pathology might be reduced without compromising immunogenicity. We found that an rVSV vaccine induced local and systemic production of IL-1β in vivo, and that accumulation of IL-1β correlated with acute pathology after rVSV immunization. rVSV-induced pathology was reduced in mice deficient in the IL-1 receptor Type I, but the IL-1R-/- mice were fully protected from lethal rechallenge with a high dose of VSV. This result demonstrated that IL-1 contributed to reactogenicity of the rVSV, but was dispensable for induction of protective immunity. The amount of IL-1β detected in mice deficient in either caspase-1 or the inflammasome adaptor molecule ASC after rVSV immunization was not significantly different than that produced by wild type animals, and caspase-1-/- and ASC-/- mice were only partially protected from rVSV-induced pathology. Those data support the idea that some of the IL-1β expressed in vivo in response to VSV may be activated by a caspase-1 and ASC-independent mechanism. Together these results suggest that rVSV vectors engineered to suppress the induction of IL-1β, or signaling through the IL-1R would be less reactogenic in vivo, but would retain their immunogenicity and protective capacity. Such rVSV would be highly desirable as either vaccine vectors or

  6. Tumor Necrosis Factor-Mediated Survival of CD169+ Cells Promotes Immune Activation during Vesicular Stomatitis Virus Infection

    DEFF Research Database (Denmark)

    Shinde, Prashant V; Xu, Haifeng C; Maney, Sathish Kumar

    2018-01-01

    Innate immune activation is essential to mount an effective antiviral response and to prime adaptive immunity. Although a crucial role of CD169(+) cells during vesicular stomatitis virus (VSV) infections is increasingly recognized, factors regulating CD169(+) cells during viral infections remain ...

  7. Plant Virus Expression Vector Development: New Perspectives

    Directory of Open Access Journals (Sweden)

    Kathleen Hefferon

    2014-01-01

    Full Text Available Plant made biologics have elicited much attention over recent years for their potential in assisting those in developing countries who have poor access to modern medicine. Additional applications such as the stockpiling of vaccines against pandemic infectious diseases or potential biological warfare agents are also under investigation. Plant virus expression vectors represent a technology that enables high levels of pharmaceutical proteins to be produced in a very short period of time. Recent advances in research and development have brought about the generation of superior virus expression systems which can be readily delivered to the host plant in a manner that is both efficient and cost effective. This review presents recent innovations in plant virus expression systems and their uses for producing biologics from plants.

  8. Trojan horse lymphocytes: a vesicular stomatitis virus-specific T-cell clone lyses target cells by carrying virus.

    Science.gov (United States)

    Hom, R C; Soman, G; Finberg, R

    1989-10-01

    We have isolated a vesicular stomatitis virus (VSV)-specific CD4+ CD8- murine T-cell clone. This clone proliferates only in response to VSV and lyses infected tumor cells bearing class II major histocompatibility antigens in short-term chromium release assays. In addition, the cell has VSV antigens on its surface and is capable of killing uninfected tumor cells without major histocompatibility antigen restriction in a 2-day assay. This latter cytolytic activity is eliminated by anti-VSV antibody, indicating that its lytic activity is provided by the virus. [35S]methionine labeling and immunoprecipitation experiments demonstrated that viral protein translation is initiated after incubation of the clone with a tumor target cell, defining this as the mechanism of its cytolytic activity.

  9. Vesicular Stomatitis Virus Variants Selectively Infect and Kill Human Melanomas but Not Normal Melanocytes

    Science.gov (United States)

    Wollmann, Guido; Davis, John N.; Bosenberg, Marcus W.

    2013-01-01

    Metastatic malignant melanoma remains one of the most therapeutically challenging forms of cancer. Here we test replication-competent vesicular stomatitis viruses (VSV) on 19 primary human melanoma samples and compare these infections with those of normal human melanocyte control cells. Even at a low viral concentration, we found a strong susceptibility to viral oncolysis in over 70% of melanomas. In contrast, melanocytes displayed strong resistance to virus infection and showed complete protection by interferon. Several recombinant VSVs were compared, and all infected and killed most melanomas with differences in the time course with increasing rates of melanoma infection, as follows: VSV-CT9-M51 melanoma xenografts in SCID mice after tail vein virus application. Sequence analysis of mutations in the melanomas used revealed that BRAF but not NRAS gene mutation status was predictive for enhanced susceptibility to infection. In mouse melanoma models with specific induced gene mutations including mutations of the Braf, Pten, and Cdkn2a genes, viral infection correlated with the extent of malignant transformation. Similar to human melanocytes, mouse melanocytes resisted VSV-rp30 infection. This study confirms the general susceptibility of the majority of human melanoma types for VSV-mediated oncolysis. PMID:23552414

  10. Preimplantation bovine embryos: Pathobiology of Haemophilus somnus exposure and resistance mechanisms to vesicular stomatitis virus

    International Nuclear Information System (INIS)

    Thomson, M.S.

    1988-01-01

    Preimplantation bovine embryos were exposed in vitro to H. somnus to determine if the bacteria would adhere to zona pellucida-intact (ZP-I) embryos or adhere to or infect ZP-free embryos. The effect of H. somnus on embryonic development in vitro was also investigated. Electrophoretic comparisons of outer membrane proteins of H. somnus revealed 2 major protein bands common to 10 H. somnus isolates. A monoclonal antibody produced against the outer membrane proteins reacted to one of the major protein bands. The sensitivity of a nucleic acid probe for detection of vesicular stomatitis virus (VSV) was validated in cells in culture and used to determine if the synthetic double-stranded complex of polyriboinosinic and polyribocytidylic acids (poly I:C) would induce viral resistance in cultured bovine embryos. Two 32 P-nick translated probes of high specific activity prepared from plasmids containing nucleic acid sequences of VSV virus were employed for viral mRNA detection in the tissue culture cells using a DNA-hybridization dot-blot technique. Using one of the probes, the technique was applied to detect differences in viral replication between four groups of bovine embryos (nonexposed, exposed to VSV virus, poly I:C-treated, and poly I:C-treated and exposed to VSV). The nucleic acid probe was sufficiently sensitive to detect differences in quantities of VSV mRNA among embryo treatment groups, resulting in the demonstration that resistance to viral infection was induced in day 9 bovine embryos

  11. Live Attenuated Recombinant Vaccine Protects Nonhuman Primates Against Ebola and Marburg Viruses

    National Research Council Canada - National Science Library

    Jones, Steven M; Feldmann, Heinz; Stroher, Ute; Geisbert, Joan B; Fernando, Lisa; Grolla, Allen; Klenk, Hans-Dieter; Sullivan, Nancy J; Volchkov, Viktor E; Fritz, Elizabeth A; Daddario, Kathleen M; Hensley, Lisa E; Jahrling, Peter B; Geisbert, Thomas W

    2005-01-01

    ...). Here, we developed replication-competent vaccines against EBOV and MARV based on attenuated recombinant vesicular stomatitis virus vectors expressing either the EBOV glycoprotein or MARV glycoprotein...

  12. Different effect of proteasome inhibition on vesicular stomatitis virus and poliovirus replication.

    Directory of Open Access Journals (Sweden)

    Nickolay Neznanov

    2008-04-01

    Full Text Available Proteasome activity is an important part of viral replication. In this study, we examined the effect of proteasome inhibitors on the replication of vesicular stomatitis virus (VSV and poliovirus. We found that the proteasome inhibitors significantly suppressed VSV protein synthesis, virus accumulation, and protected infected cells from toxic effect of VSV replication. In contrast, poliovirus replication was delayed, but not diminished in the presence of the proteasome inhibitors MG132 and Bortezomib. We also found that inhibition of proteasomes stimulated stress-related processes, such as accumulation of chaperone hsp70, phosphorylation of eIF2alpha, and overall inhibition of translation. VSV replication was sensitive to this stress with significant decline in replication process. Poliovirus growth was less sensitive with only delay in replication. Inhibition of proteasome activity suppressed cellular and VSV protein synthesis, but did not reduce poliovirus protein synthesis. Protein kinase GCN2 supported the ability of proteasome inhibitors to attenuate general translation and to suppress VSV replication. We propose that different mechanisms of translational initiation by VSV and poliovirus determine their sensitivity to stress induced by the inhibition of proteasomes. To our knowledge, this is the first study that connects the effect of stress induced by proteasome inhibition with the efficiency of viral infection.

  13. Long-Term Single-Dose Efficacy of a Vesicular Stomatitis Virus-Based Andes Virus Vaccine in Syrian Hamsters

    Directory of Open Access Journals (Sweden)

    Joseph Prescott

    2014-01-01

    Full Text Available Andes virus (ANDV is highly pathogenic in humans and is the primary etiologic agent of hantavirus cardiopulmonary syndrome (HCPS in South America. Case-fatality rates are as high as 50% and there are no approved vaccines or specific therapies for infection. Our laboratory has recently developed a replication-competent recombinant vesicular stomatitis virus (VSV-based vaccine that expressed the glycoproteins of Andes virus in place of the native VSV glycoprotein (G. This vaccine is highly efficacious in the Syrian hamster model of HCPS when given 28 days before challenge with ANDV, or when given around the time of challenge (peri-exposure, and even protects when administered post-exposure. Herein, we sought to test the durability of the immune response to a single dose of this vaccine in Syrian hamsters. This vaccine was efficacious in hamsters challenged intranasally with ANDV 6 months after vaccination (p = 0.025, but animals were not significantly protected following 1 year of vaccination (p = 0.090. The decrease in protection correlated with a reduction of measurable neutralizing antibody responses, and suggests that a more robust vaccination schedule might be required to provide long-term immunity.

  14. An effective AIDS vaccine based on live attenuated vesicular stomatitis virus recombinants.

    Science.gov (United States)

    Rose, N F; Marx, P A; Luckay, A; Nixon, D F; Moretto, W J; Donahoe, S M; Montefiori, D; Roberts, A; Buonocore, L; Rose, J K

    2001-09-07

    We developed an AIDS vaccine based on attenuated VSV vectors expressing env and gag genes and tested it in rhesus monkeys. Boosting was accomplished using vectors with glycoproteins from different VSV serotypes. Animals were challenged with a pathogenic AIDS virus (SHIV89.6P). Control monkeys showed a severe loss of CD4+ T cells and high viral loads, and 7/8 progressed to AIDS with an average time of 148 days. All seven vaccinees were initially infected with SHIV89.6P but have remained healthy for up to 14 months after challenge with low or undetectable viral loads. Protection from AIDS was highly significant (p = 0.001). VSV vectors are promising candidates for human AIDS vaccine trials because they propagate to high titers and can be delivered without injection.

  15. Antibodies against vesicular stomatitis virus in horses from southern, midwestern and northeastern Brazilian States

    Directory of Open Access Journals (Sweden)

    Vinícius Leobet Lunkes

    2016-08-01

    Full Text Available ABSTRACT: Vesicular stomatitis virus (VSV is the agent of a vesicular disease that affects many animal species and may be clinically confounded with foot-and-mouth disease in ruminant and swine. Horses are especially susceptible to VSV and may serve as sentinels for virus circulation. The present study investigated the presence of neutralizing antibodies against VSV Indiana III (VSIV-3 in serum samples of 3,626 horses from six states in three Brazilian regions: Southern (RS, n = 1,011, Midwest (GO/DF, n = 1,767 and Northeast (PB, PE, RN and CE, n = 848 collected between 2013 and 2014. Neutralizing antibodies against VSIV-3 (titers ≥40 were detected in 641 samples (positivity of 17.7%; CI95%:16.5-19.0%, being 317 samples from CE (87.3%; CI95%: 83.4-90.5 %; 109 from RN (65.7%; CI95%: 57.8 -72.7%; 124 from PB (45.4%; CI95%: 39.4-51.5%; 78 from GO/DF (4.4%; CI95%: 3.5-5.5% and nine samples of RS (0.9%; CI95%: 0.4-1.7%. Several samples from the Northeast and Midwest harbored high neutralizing titers, indicating a recent exposure to the virus. In contrast, samples from RS had low titers, possibly due to a past remote exposure. Several positive samples presented neutralizing activity against other VSV serotypes (Indiana I and New Jersey, yet in lower titers, indicating the specificity of the response to VSIV-3. These results demonstrated a relatively recent circulation of VSIV-3 in northeastern Brazilian States, confirming clinical findings and demonstrating the sanitary importance of this infection.

  16. The transcriptional form of the phosphoprotein of vesicular stomatitis virus is a trimer: structure and stability.

    Science.gov (United States)

    Gao, Y; Greenfield, N J; Cleverley, D Z; Lenard, J

    1996-11-19

    The phosphoprotein (P) of vesicular stomatitis virus was previously shown to assemble into a homomultimer upon phosphorylation by casein kinase II. It thus acquired transcriptional activity, including the ability to bind to the other two transcriptional components, the polymerase L and the N-RNA template. This multimer has now been found to be a trimer using a His-tag dilution method. Trimer stability was assessed using a variation of this method, by measuring the rate of exchange of monomers between preformed tagged and untagged trimers at different values of pH and ionic strength. Exchange rates increased with increasing ionic strength and were similar at pH 6, 8, and 10, but the trimer was completely dissociated at pH 4. This suggests that the trimer is stabilized by electrostatic interactions, probably involving carboxylate and guanidino groups. Addition of viral L protein stabilized the P trimers, completely preventing subunit exchange under transcription conditions. The association constants (Kass) for trimerization of partially active D and A substitution mutants were also determined by His-tag dilution and found to correlate well with transcriptional activity, further confirming that the active species is the trimer. Circular dichroism spectra were identical for phosphorylated and unphosphorylated wild-type P protein and for D and A mutants known to be predominantly trimeric and monomeric, respectively.

  17. Vesicular stomatitis virus infection promotes immune evasion by preventing NKG2D-ligand surface expression.

    Science.gov (United States)

    Jensen, Helle; Andresen, Lars; Nielsen, Jens; Christensen, Jan Pravsgaard; Skov, Søren

    2011-01-01

    Vesicular stomatitis virus (VSV) has recently gained attention for its oncolytic ability in cancer treatment. Initially, we hypothesized that VSV infection could increase immune recognition of cancer cells through induction of the immune stimulatory NKG2D-ligands. Here we show that VSV infection leads to a robust induction of MICA mRNA expression, however the subsequent surface expression is potently hindered. Thus, VSV lines up with human cytomegalovirus (HCMV) and adenovirus, which actively subvert the immune system by negatively affecting NKG2D-ligand surface expression. VSV infection caused an active suppression of NKG2D-ligand surface expression, affecting both endogenous and histone deacetylase (HDAC)-inhibitor induced MICA, MICB and ULBP-2 expression. The classical immune escape mechanism of VSV (i.e., the M protein blockade of nucleocytoplasmic mRNA transport) was not involved, as the VSV mutant strain, VSV(ΔM51), which possess a defective M protein, prevented MICA surface expression similarly to wild-type VSV. The VSV mediated down modulation of NKG2D-ligand expression did not involve apoptosis. Constitutive expression of MICA bypassed the escape mechanism, suggesting that VSV affect NKG2D-ligand expression at an early post-transcriptional level. Our results show that VSV possess an escape mechanism, which could affect the immune recognition of VSV infected cancer cells. This may also have implications for immune recognition of cancer cells after combined treatment with VSV and chemotherapeutic drugs.

  18. Intrinsic Innate Immunity Fails To Control Herpes Simplex Virus and Vesicular Stomatitis Virus Replication in Sensory Neurons and Fibroblasts

    Science.gov (United States)

    Rosato, Pamela C.

    2014-01-01

    ABSTRACT Herpes simplex virus 1 (HSV-1) establishes lifelong latent infections in the sensory neurons of the trigeminal ganglia (TG), wherein it retains the capacity to reactivate. The interferon (IFN)-driven antiviral response is critical for the control of HSV-1 acute replication. We therefore sought to further investigate this response in TG neurons cultured from adult mice deficient in a variety of IFN signaling components. Parallel experiments were also performed in fibroblasts isolated concurrently. We showed that HSV-1 replication was comparable in wild-type (WT) and IFN signaling-deficient neurons and fibroblasts. Unexpectedly, a similar pattern was observed for the IFN-sensitive vesicular stomatitis virus (VSV). Despite these findings, TG neurons responded to IFN-β pretreatment with STAT1 nuclear localization and restricted replication of both VSV and an HSV-1 strain deficient in γ34.5, while wild-type HSV-1 replication was unaffected. This was in contrast to fibroblasts in which all viruses were restricted by the addition of IFN-β. Taken together, these data show that adult TG neurons can mount an effective antiviral response only if provided with an exogenous source of IFN-β, and HSV-1 combats this response through γ34.5. These results further our understanding of the antiviral response of neurons and highlight the importance of paracrine IFN-β signaling in establishing an antiviral state. IMPORTANCE Herpes simplex virus 1 (HSV-1) is a ubiquitous virus that establishes a lifelong latent infection in neurons. Reactivation from latency can cause cold sores, blindness, and death from encephalitis. Humans with deficiencies in innate immunity have significant problems controlling HSV infections. In this study, we therefore sought to elucidate the role of neuronal innate immunity in the control of viral infection. Using neurons isolated from mice, we found that the intrinsic capacity of neurons to restrict virus replication was unaffected by the presence

  19. Reovirus FAST Protein Enhances Vesicular Stomatitis Virus Oncolytic Virotherapy in Primary and Metastatic Tumor Models

    Directory of Open Access Journals (Sweden)

    Fabrice Le Boeuf

    2017-09-01

    Full Text Available The reovirus fusion-associated small transmembrane (FAST proteins are the smallest known viral fusogens (∼100–150 amino acids and efficiently induce cell-cell fusion and syncytium formation in multiple cell types. Syncytium formation enhances cell-cell virus transmission and may also induce immunogenic cell death, a form of apoptosis that stimulates immune recognition of tumor cells. These properties suggest that FAST proteins might serve to enhance oncolytic virotherapy. The oncolytic activity of recombinant VSVΔM51 (an interferon-sensitive vesicular stomatitis virus [VSV] mutant encoding the p14 FAST protein (VSV-p14 was compared with a similar construct encoding GFP (VSV-GFP in cell culture and syngeneic BALB/c tumor models. Compared with VSV-GFP, VSV-p14 exhibited increased oncolytic activity against MCF-7 and 4T1 breast cancer spheroids in culture and reduced primary 4T1 breast tumor growth in vivo. VSV-p14 prolonged survival in both primary and metastatic 4T1 breast cancer models, and in a CT26 metastatic colon cancer model. As with VSV-GFP, VSV-p14 preferentially replicated in vivo in tumors and was cleared rapidly from other sites. Furthermore, VSV-p14 increased the numbers of activated splenic CD4, CD8, natural killer (NK, and natural killer T (NKT cells, and increased the number of activated CD4 and CD8 cells in tumors. FAST proteins may therefore provide a multi-pronged approach to improving oncolytic virotherapy via syncytium formation and enhanced immune stimulation.

  20. Data-driven identification of potential Zika virus vectors

    Science.gov (United States)

    Evans, Michelle V; Dallas, Tad A; Han, Barbara A; Murdock, Courtney C; Drake, John M

    2017-01-01

    Zika is an emerging virus whose rapid spread is of great public health concern. Knowledge about transmission remains incomplete, especially concerning potential transmission in geographic areas in which it has not yet been introduced. To identify unknown vectors of Zika, we developed a data-driven model linking vector species and the Zika virus via vector-virus trait combinations that confer a propensity toward associations in an ecological network connecting flaviviruses and their mosquito vectors. Our model predicts that thirty-five species may be able to transmit the virus, seven of which are found in the continental United States, including Culex quinquefasciatus and Cx. pipiens. We suggest that empirical studies prioritize these species to confirm predictions of vector competence, enabling the correct identification of populations at risk for transmission within the United States. DOI: http://dx.doi.org/10.7554/eLife.22053.001 PMID:28244371

  1. Vectors expressing chimeric Japanese encephalitis dengue 2 viruses.

    Science.gov (United States)

    Wei, Y; Wang, S; Wang, X

    2014-01-01

    Vectors based on self-replicating RNAs (replicons) of flaviviruses are becoming powerful tool for expression of heterologous genes in mammalian cells and development of novel antiviral and anticancer vaccines. We constructed two vectors expressing chimeric viruses consisting of attenuated SA14-14-2 strain of Japanese encephalitis virus (JEV) in which the PrM/M-E genes were replaced fully or partially with those of dengue 2 virus (DENV-2). These vectors, named pJED2 and pJED2-1770 were transfected to BHK-21 cells and produced chimeric viruses JED2V and JED2-1770V, respectively. The chimeric viruses could be passaged in C6/36 but not BHK-21 cells. The chimeric viruses produced in C6/36 cells CPE 4-5 days after infection and RT-PCR, sequencing, immunofluorescence assay (IFA) and Western blot analysis confirmed the chimeric nature of produced viruses. The immunogenicity of chimeric viruses in mice was proved by detecting DENV-2 E protein-specific serum IgG antibodies with neutralization titer of 10. Successful preparation of infectious clones of chimeric JEV-DENV-2 viruses showed that JEV-based expression vectors are fully functional.

  2. Recoding of the vesicular stomatitis virus L gene by computer-aided design provides a live, attenuated vaccine candidate.

    Science.gov (United States)

    Wang, Bingyin; Yang, Chen; Tekes, Gergely; Mueller, Steffen; Paul, Aniko; Whelan, Sean P J; Wimmer, Eckard

    2015-03-31

    Codon pair bias (CPB), which has been observed in all organisms, is a neglected genomic phenomenon that affects gene expression. CPB results from synonymous codons that are paired more or less frequently in ORFeomes regardless of codon bias. The effect of an individual codon pair change is usually small, but when it is amplified by large-scale genome recoding, strikingly altered biological phenotypes are observed. The utility of codon pair bias in the development of live attenuated vaccines was recently demonstrated by recodings of poliovirus (a positive-strand RNA virus) and influenza virus (a negative-strand segmented RNA virus). Here, the L gene of vesicular stomatitis virus (VSV), a nonsegmented negative-sense RNA virus, was partially recoded based on codon pair bias. Totals of 858 and 623 silent mutations were introduced into a 5'-terminal segment of the viral L gene (designated L1) to create sequences containing either overrepresented or underrepresented codon pairs, designated L1(sdmax) and L1(min), respectively. Analysis revealed that recombinant VSV containing the L1(min) sequence could not be recovered, whereas the virus with the sdmax sequence showed a modest level of attenuation in cell culture. More strikingly, in mice the L1(sdmax) virus was almost as immunogenic as the parental strain but highly attenuated. Taken together, these results open a new road to attain a balance between VSV virulence and immunogenicity, which could serve as an example for the attenuation of other negative-strand, nonsegmented RNA viruses. Vesicular stomatitis virus (VSV) is the prototypic rhabdovirus in the order Mononegavirales. A wide range of human pathogens belong to this family. Using a unique computer algorithm and large-scale genome synthesis, we attempted to develop a live attenuated vaccine strain for VSV, which could be used as an antigen delivery platform for humans. Recombinant VSVs with distinct codon pair biases were rationally designed, constructed, and

  3. Improvement and optimization of a multiplex real-time reverse transcription polymerase chain reaction assay for the detection and typing of Vesicular stomatitis virus.

    Science.gov (United States)

    Hole, Kate; Velazquez-Salinas, Lauro; Velazques-Salinas, Lauro; Clavijo, Alfonso

    2010-05-01

    An improvement to a previously reported real-time reverse transcription polymerase chain reaction (real-time RT-PCR) assay for the detection of Vesicular stomatitis virus (VSV) is described. Results indicate that the new assay is capable of detecting a panel of genetically representative strains of VSV present in North, Central, and South America. The assay is specific for VSV and allows for simultaneous differentiation between Vesicular stomatitis Indiana virus and Vesicular stomatitis New Jersey virus. This real-time RT-PCR is able to detect current circulating strains of VSV and can be used for rapid diagnosis of VSV and differentiation of VSV from other vesicular diseases, such as foot-and-mouth disease.

  4. Bioreactor production of recombinant herpes simplex virus vectors.

    Science.gov (United States)

    Knop, David R; Harrell, Heather

    2007-01-01

    Serotypical application of herpes simplex virus (HSV) vectors to gene therapy (type 1) and prophylactic vaccines (types 1 and 2) has garnered substantial clinical interest recently. HSV vectors and amplicons have also been employed as helper virus constructs for manufacture of the dependovirus adeno-associated virus (AAV). Large quantities of infectious HSV stocks are requisite for these therapeutic applications, requiring a scalable vector manufacturing and processing platform comprised of unit operations which accommodate the fragility of HSV. In this study, production of a replication deficient rHSV-1 vector bearing the rep and cap genes of AAV-2 (denoted rHSV-rep2/cap2) was investigated. Adaptation of rHSV production from T225 flasks to a packed bed, fed-batch bioreactor permitted an 1100-fold increment in total vector production without a decrease in specific vector yield (pfu/cell). The fed-batch bioreactor system afforded a rHSV-rep2/cap2 vector recovery of 2.8 x 10(12) pfu. The recovered vector was concentrated by tangential flow filtration (TFF), permitting vector stocks to be formulated at greater than 1.5 x 10(9) pfu/mL.

  5. Evolutionary dynamics of dengue virus populations within the mosquito vector.

    Science.gov (United States)

    Lambrechts, Louis; Lequime, Sebastian

    2016-12-01

    To date, dengue virus evolution has mainly been addressed by studies conducted at the between-host level. Like other pathogens with high mutation rate and rapid replication, dengue viruses also evolve during the course of an infection. Over the last few years, the advent of deep-sequencing technologies has facilitated studies of dengue virus populations at the within-host level. Here, we review recent advances on the evolutionary dynamics of dengue virus populations within their mosquito vector. We discuss how identifying the evolutionary forces acting on dengue virus populations within the mosquito can shed light on the processes underlying vector-virus interactions and the evolution of epidemiologically relevant traits. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Vaccinia virus as an expression vector.

    Science.gov (United States)

    Talavera, A; Rodriguez, J M

    1992-01-01

    Vaccinia virus (Vv) is a member of the genus Orthopoxvirus, one of seven genera included in the family Poxviridae. Most of these viruses infect vertebrates (Orthopoxvirus, Avipoxvirus, Capripoxvirus, Leporipoxvirus, Suipoxvirus, and Parapoxvirus), but one genus, Entomopoxvirus, infects insects. It is interesting to note that the Fibroma and Mixoma viruses of the leporipoxvirus genus cause tumors in their hosts (rabbits), these being the only tumorigenic viruses in the family (1,2).

  7. Digital sensing and sizing of vesicular stomatitis virus pseudotypes in complex media: a model for Ebola and Marburg detection.

    Science.gov (United States)

    Daaboul, George G; Lopez, Carlos A; Chinnala, Jyothsna; Goldberg, Bennett B; Connor, John H; Ünlü, M Selim

    2014-06-24

    Rapid, sensitive, and direct label-free capture and characterization of nanoparticles from complex media such as blood or serum will broadly impact medicine and the life sciences. We demonstrate identification of virus particles in complex samples for replication-competent wild-type vesicular stomatitis virus (VSV), defective VSV, and Ebola- and Marburg-pseudotyped VSV with high sensitivity and specificity. Size discrimination of the imaged nanoparticles (virions) allows differentiation between modified viruses having different genome lengths and facilitates a reduction in the counting of nonspecifically bound particles to achieve a limit-of-detection (LOD) of 5 × 10(3) pfu/mL for the Ebola and Marburg VSV pseudotypes. We demonstrate the simultaneous detection of multiple viruses in a single sample (composed of serum or whole blood) for screening applications and uncompromised detection capabilities in samples contaminated with high levels of bacteria. By employing affinity-based capture, size discrimination, and a "digital" detection scheme to count single virus particles, we show that a robust and sensitive virus/nanoparticle sensing assay can be established for targets in complex samples. The nanoparticle microscopy system is termed the Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) and is capable of high-throughput and rapid sizing of large numbers of biological nanoparticles on an antibody microarray for research and diagnostic applications.

  8. Oncolytic Vesicular Stomatitis Virus as a Viro-Immunotherapy: Defeating Cancer with a “Hammer” and “Anvil”

    Directory of Open Access Journals (Sweden)

    Michael Karl Melzer

    2017-02-01

    Full Text Available Oncolytic viruses have gained much attention in recent years, due, not only to their ability to selectively replicate in and lyse tumor cells, but to their potential to stimulate antitumor immune responses directed against the tumor. Vesicular stomatitis virus (VSV, a negative-strand RNA virus, is under intense development as an oncolytic virus due to a variety of favorable properties, including its rapid replication kinetics, inherent tumor specificity, and its potential to elicit a broad range of immunomodulatory responses to break immune tolerance in the tumor microenvironment. Based on this powerful platform, a multitude of strategies have been applied to further improve the immune-stimulating potential of VSV and synergize these responses with the direct oncolytic effect. These strategies include: 1. modification of endogenous virus genes to stimulate interferon induction; 2. virus-mediated expression of cytokines or immune-stimulatory molecules to enhance anti-tumor immune responses; 3. vaccination approaches to stimulate adaptive immune responses against a tumor antigen; 4. combination with adoptive immune cell therapy for potentially synergistic therapeutic responses. A summary of these approaches will be presented in this review.

  9. A stable RNA virus-based vector for citrus trees

    International Nuclear Information System (INIS)

    Folimonov, Alexey S.; Folimonova, Svetlana Y.; Bar-Joseph, Moshe; Dawson, William O.

    2007-01-01

    Virus-based vectors are important tools in plant molecular biology and plant genomics. A number of vectors based on viruses that infect herbaceous plants are in use for expression or silencing of genes in plants as well as screening unknown sequences for function. Yet there is a need for useful virus-based vectors for woody plants, which demand much greater stability because of the longer time required for systemic infection and analysis. We examined several strategies to develop a Citrus tristeza virus (CTV)-based vector for transient expression of foreign genes in citrus trees using a green fluorescent protein (GFP) as a reporter. These strategies included substitution of the p13 open reading frame (ORF) by the ORF of GFP, construction of a self-processing fusion of GFP in-frame with the major coat protein (CP), or expression of the GFP ORF as an extra gene from a subgenomic (sg) mRNA controlled either by a duplicated CTV CP sgRNA controller element (CE) or an introduced heterologous CE of Beet yellows virus. Engineered vector constructs were examined for replication, encapsidation, GFP expression during multiple passages in protoplasts, and for their ability to infect, move, express GFP, and be maintained in citrus plants. The most successful vectors based on the 'add-a-gene' strategy have been unusually stable, continuing to produce GFP fluorescence after more than 4 years in citrus trees

  10. Unique Safety Issues Associated with Virus Vectored Vaccines: Potential for and Theoretical Consequences of Recombination with Wild Type Virus Strains

    Science.gov (United States)

    Condit, Richard C.; Williamson, Anna-Lise; Sheets, Rebecca; Seligman, Stephen J.; Monath, Thomas P.; Excler, Jean-Louis; Gurwith, Marc; Bok, Karin; Robertson, James S.; Kim, Denny; Hendry, Michael; Singh, Vidisha; Mac, Lisa M.; Chen, Robert T.

    2016-01-01

    In 2003 and 2013, the World Health Organization convened informal consultations on characterization and quality aspects of vaccines based on live virus vectors. In the resulting reports, one of several issues raised for future study was the potential for recombination of virus-vectored vaccines with wild type pathogenic virus strains. This paper presents an assessment of this issue formulated by the Brighton Collaboration. To provide an appropriate context for understanding the potential for recombination of virus-vectored vaccines, we review briefly the current status of virus vectored vaccines, mechanisms of recombination between viruses, experience with recombination involving live attenuated vaccines in the field, and concerns raised previously in the literature regarding recombination of virus-vectored vaccines with wild type virus strains. We then present a discussion of the major variables that could influence recombination between a virus-vectored vaccine and circulating wild type virus and the consequences of such recombination, including intrinsic recombination properties of the parent virus used as a vector; sequence relatedness of vector and wild virus; virus host range, pathogenesis and transmission; replication competency of vector in target host; mechanism of vector attenuation; additional factors potentially affecting virulence; and circulation of multiple recombinant vectors in the same target population. Finally, we present some guiding principles for vector design and testing intended to anticipate and mitigate the potential for and consequences of recombination of virus-vectored vaccines with wild type pathogenic virus strains. PMID:27346303

  11. Cooperation of B cells and T cells is required for survival of mice infected with vesicular stomatitis virus

    DEFF Research Database (Denmark)

    Thomsen, Allan Randrup; Nansen, A; Andersen, C

    1997-01-01

    To define the role of T cells and B cells in resistance to vesicular stomatitis virus (VSV) infection, knockout mice with different specific immune defects on an identical background were infected i.v. and the outcome of infection was compared; in this way a more complete picture of the relative....... In contrast, B cell-deficient mice were highly susceptible even to low doses of virus and mortality could be prevented by transfer of naive B cells prior to challenge as well as by immune serum given after challenge. Analysis of MHC class I- and class II-deficient mice revealed that CD8+ T cells could exert...... importance of various host defence mechanisms could be obtained. Compared to T and B cell-deficient SCID mice which all succumbed from encephalitis within 5-9 days of infection, T cell-deficient nude mice generally lived longer, but within a period of approximately 1 month after challenge all died...

  12. Large Animal Models for Foamy Virus Vector Gene Therapy

    Directory of Open Access Journals (Sweden)

    Peter A. Horn

    2012-12-01

    Full Text Available Foamy virus (FV vectors have shown great promise for hematopoietic stem cell (HSC gene therapy. Their ability to efficiently deliver transgenes to multi-lineage long-term repopulating cells in large animal models suggests they will be effective for several human hematopoietic diseases. Here, we review FV vector studies in large animal models, including the use of FV vectors with the mutant O6-methylguanine-DNA methyltransferase, MGMTP140K to increase the number of genetically modified cells after transplantation. In these studies, FV vectors have mediated efficient gene transfer to polyclonal repopulating cells using short ex vivo transduction protocols designed to minimize the negative effects of ex vivo culture on stem cell engraftment. In this regard, FV vectors appear superior to gammaretroviral vectors, which require longer ex vivo culture to effect efficient transduction. FV vectors have also compared favorably with lentiviral vectors when directly compared in the dog model. FV vectors have corrected leukocyte adhesion deficiency and pyruvate kinase deficiency in the dog large animal model. FV vectors also appear safer than gammaretroviral vectors based on a reduced frequency of integrants near promoters and also near proto-oncogenes in canine repopulating cells. Together, these studies suggest that FV vectors should be highly effective for several human hematopoietic diseases, including those that will require relatively high percentages of gene-modified cells to achieve clinical benefit.

  13. Dynamics of melanoma tumor therapy with vesicular stomatitis virus: explaining the variability in outcomes using mathematical modeling.

    Science.gov (United States)

    Rommelfanger, D M; Offord, C P; Dev, J; Bajzer, Z; Vile, R G; Dingli, D

    2012-05-01

    Tumor selective, replication competent viruses are being tested for cancer gene therapy. This approach introduces a new therapeutic paradigm due to potential replication of the therapeutic agent and induction of a tumor-specific immune response. However, the experimental outcomes are quite variable, even when studies utilize highly inbred strains of mice and the same cell line and virus. Recognizing that virotherapy is an exercise in population dynamics, we utilize mathematical modeling to understand the variable outcomes observed when B16ova malignant melanoma tumors are treated with vesicular stomatitis virus in syngeneic, fully immunocompetent mice. We show how variability in the initial tumor size and the actual amount of virus delivered to the tumor have critical roles on the outcome of therapy. Virotherapy works best when tumors are small, and a robust innate immune response can lead to superior tumor control. Strategies that reduce tumor burden without suppressing the immune response and methods that maximize the amount of virus delivered to the tumor should optimize tumor control in this model system.

  14. Virus infection of a weed increases vector attraction to and vector fitness on the weed

    Science.gov (United States)

    Chen, Gong; Pan, Huipeng; Xie, Wen; Wang, Shaoli; Wu, Qingjun; Fang, Yong; Shi, Xiaobin; Zhang, Youjun

    2013-01-01

    Weeds are important in the ecology of field crops, and when crops are harvested, weeds often become the main hosts for plant viruses and their insect vectors. Few studies, however, have examined the relationships between plant viruses, vectors, and weeds. Here, we investigated how infection of the weed Datura stramonium L. by tomato yellow leaf curl virus (TYLCV) affects the host preference and performance of the TYLCV vector, Bemisia tabaci (Gennadius) Q. The results of a choice experiment indicated that B. tabaci Q preferentially settled and oviposited on TYLCV-infected plants rather than on healthy plants. In addition, B. tabaci Q performed better on TYLCV-infected plants than on healthy plants. These results demonstrate that TYLCV is indirectly mutualistic to B. tabaci Q. The mutually beneficial interaction between TYLCV and B. tabaci Q may help explain the concurrent outbreaks of TYLCV and B. tabaci Q in China. PMID:23872717

  15. Vector competence of Anopheles and Culex mosquitoes for Zika virus

    Directory of Open Access Journals (Sweden)

    Brittany L. Dodson

    2017-03-01

    Full Text Available Zika virus is a newly emergent mosquito-borne flavivirus that has caused recent large outbreaks in the new world, leading to dramatic increases in serious disease pathology including Guillain-Barre syndrome, newborn microcephaly, and infant brain damage. Although Aedes mosquitoes are thought to be the primary mosquito species driving infection, the virus has been isolated from dozens of mosquito species, including Culex and Anopheles species, and we lack a thorough understanding of which mosquito species to target for vector control. We exposed Anopheles gambiae, Anopheles stephensi, and Culex quinquefasciatus mosquitoes to blood meals supplemented with two Zika virus strains. Mosquito bodies, legs, and saliva were collected five, seven, and 14 days post blood meal and tested for infectious virus by plaque assay. Regardless of titer, virus strain, or timepoint, Anopheles gambiae, Anopheles stephensi, and Culex quinquefasciatus mosquitoes were refractory to Zika virus infection. We conclude that Anopheles gambiae, Anopheles stephensi, and Culex quinquefasciatus mosquitoes likely do not contribute significantly to Zika virus transmission to humans. However, future studies should continue to explore the potential for other novel potential vectors to transmit the virus.

  16. Potential for biological control of rice yellow mottle virus vectors ...

    African Journals Online (AJOL)

    Insect pests and disease infestations are the primary constraints in rice (Oryza sativa) production systems in Africa and Asia. ... Unfortunately, 2002 - 2004, two sampling methods were combined to assess the population of insects vectors of rice yellow mottle virus (RYMV) in the three major irrigated rice ecosystems in ...

  17. A Foxtail mosaic virus Vector for Virus-Induced Gene Silencing in Maize1[OPEN

    Science.gov (United States)

    Mei, Yu; Kernodle, Bliss M.; Hill, John H.

    2016-01-01

    Plant viruses have been widely used as vectors for foreign gene expression and virus-induced gene silencing (VIGS). A limited number of viruses have been developed into viral vectors for the purposes of gene expression or VIGS in monocotyledonous plants, and among these, the tripartite viruses Brome mosaic virus and Cucumber mosaic virus have been shown to induce VIGS in maize (Zea mays). We describe here a new DNA-based VIGS system derived from Foxtail mosaic virus (FoMV), a monopartite virus that is able to establish systemic infection and silencing of endogenous maize genes homologous to gene fragments inserted into the FoMV genome. To demonstrate VIGS applications of this FoMV vector system, four genes, phytoene desaturase (functions in carotenoid biosynthesis), lesion mimic22 (encodes a key enzyme of the porphyrin pathway), iojap (functions in plastid development), and brown midrib3 (caffeic acid O-methyltransferase), were silenced and characterized in the sweet corn line Golden × Bantam. Furthermore, we demonstrate that the FoMV infectious clone establishes systemic infection in maize inbred lines, sorghum (Sorghum bicolor), and green foxtail (Setaria viridis), indicating the potential wide applications of this viral vector system for functional genomics studies in maize and other monocots. PMID:27208311

  18. A Foxtail mosaic virus Vector for Virus-Induced Gene Silencing in Maize.

    Science.gov (United States)

    Mei, Yu; Zhang, Chunquan; Kernodle, Bliss M; Hill, John H; Whitham, Steven A

    2016-06-01

    Plant viruses have been widely used as vectors for foreign gene expression and virus-induced gene silencing (VIGS). A limited number of viruses have been developed into viral vectors for the purposes of gene expression or VIGS in monocotyledonous plants, and among these, the tripartite viruses Brome mosaic virus and Cucumber mosaic virus have been shown to induce VIGS in maize (Zea mays). We describe here a new DNA-based VIGS system derived from Foxtail mosaic virus (FoMV), a monopartite virus that is able to establish systemic infection and silencing of endogenous maize genes homologous to gene fragments inserted into the FoMV genome. To demonstrate VIGS applications of this FoMV vector system, four genes, phytoene desaturase (functions in carotenoid biosynthesis), lesion mimic22 (encodes a key enzyme of the porphyrin pathway), iojap (functions in plastid development), and brown midrib3 (caffeic acid O-methyltransferase), were silenced and characterized in the sweet corn line Golden × Bantam. Furthermore, we demonstrate that the FoMV infectious clone establishes systemic infection in maize inbred lines, sorghum (Sorghum bicolor), and green foxtail (Setaria viridis), indicating the potential wide applications of this viral vector system for functional genomics studies in maize and other monocots. © 2016 American Society of Plant Biologists. All Rights Reserved.

  19. Vectors of Crimean Congo Hemorrhagic Fever Virus in Iran

    Directory of Open Access Journals (Sweden)

    Zakkyeh Telmadarraiy

    2015-10-01

    Full Text Available Background: Ticks are important vectors and reservoirs of Crimean Congo Hemorrhagic Fever (CCHF virus. Human beings may be infected whenever the normal life cycle of the infected ticks on non- human vertebrate hosts is interrupted by the undesirable presence of humans in the cycle. A total of 26 species of Argasid and Ixodid ticks have been recorded in Iran; including nine Hyalomma, two Rhipicephalus, two Dermacentor, five Haemaphysalis, two Boophilus, one Ixodes and two Argas as well as three Ornithodoros species as blood sucking ectoparasites of livestock and poultries. The present paper reviews tick vectors of CCHF virus in Iran, focusing on the role of ticks in different provinces of Iran using reverse transcription polymerase chain reaction (RT-PCR assay.Methods: During ten years study, 1054 tick specimens; including two species of Argasidae and 17 species of Ixodidae were examined for their infection to CCHF virus genome. The output of all studies as well as related publications were discussed in the current paper.Results: The results show that Rhipicephalus sanguineus, Hyalomma marginatum, H. anatolicum, H. asiaticum and H. dromedarii were known as the most frequent species which were positive for CCHF virus.Conclusion: The status of ticks which were positive for CCHF virus revealed that unlike the most common idea that Hyalomma species are the most important vectors of CCHF virus, other ticks including Rhipicephalus,Haemaphysalis and Dermacentor can be reservoir of this virus; thus, considering geographical distribution, type of host and environmental conditions, different tick control measurements should be carried out in areas with high incidence of CCHF disease.

  20. Nonreciprocal Pseudotyping: Murine Leukemia Virus Proteins Cannot Efficiently Package Spleen Necrosis Virus-Based Vector RNA

    OpenAIRE

    Certo, Jeanine L.; Shook, Betsy F.; Yin, Philip D.; Snider, John T.; Hu, Wei-Shau

    1998-01-01

    It has been documented that spleen necrosis virus (SNV) can package murine leukemia virus (MLV) RNA efficiently and propagate MLV vectors to the same titers as it propagates SNV-based vectors. Although the SNV packaging signal (E) and MLV packaging signal (Ψ) have little sequence homology, similar double-hairpin RNA structures were predicted and supported by experimental evidence. To test whether SNV RNA can be packaged by MLV proteins, we modified an SNV vector to be expressed in an MLV-base...

  1. Virusoverdracht en vectorbestrijding in pootaardappelgewassen = Virus transmission and vector control in seed potatoes (Solanum tuberosum L.)

    NARCIS (Netherlands)

    Wolf, van der J.P.M.

    1961-01-01

    The extensive literature concerning leafroll virus, virus Y and their vector Myzus persicae Sulz. was surveyed. The relations between virus, aphid and plant were discussed. In virus spread, endogenous infection (spread of virus within the field whereby virus diseased plants act as sources of

  2. In Vivo Replication and Pathogenesis of Vesicular Stomatitis Virus Recombinant M40 Containing Ebola Virus L-Domain Sequences

    Directory of Open Access Journals (Sweden)

    Takashi Irie

    2012-01-01

    Full Text Available The M40 VSV recombinant was engineered to contain overlapping PTAP and PPxY L-domain motifs and flanking residues from the VP40 protein of Ebola virus. Replication of M40 in cell culture is virtually indistinguishable from that of control viruses. However, the presence of the Ebola PTAP motif in the M40 recombinant enabled this virus to interact with and recruit host Tsg101, which was packaged into M40 virions. In this brief report, we compared replication and the pathogenic profiles of M40 and the parental virus M51R in mice to determine whether the presence of the Ebola L-domains and flanking residues altered in vivo characteristics of the virus. Overall, the in vivo characteristics of M40 were similar to those of the parental M51R virus, indicating that the Ebola sequences did not alter pathogenesis of VSV in this small animal model of infection.

  3. Vaccinia virus vectors: new strategies for producing recombinant vaccines.

    Science.gov (United States)

    Hruby, D E

    1990-01-01

    The development and continued refinement of techniques for the efficient insertion and expression of heterologous DNA sequences from within the genomic context of infectious vaccinia virus recombinants are among the most promising current approaches towards effective immunoprophylaxis against a variety of protozoan, viral, and bacterial human pathogens. Because of its medical relevance, this area is the subject of intense research interest and has evolved rapidly during the past several years. This review (i) provides an updated overview of the technology that exists for assembling recombinant vaccinia virus strains, (ii) discusses the advantages and disadvantages of these approaches, (iii) outlines the areas of outgoing research directed towards overcoming the limitations of current techniques, and (iv) provides some insight (i.e., speculation) about probable future refinements in the use of vaccinia virus as a vector. PMID:2187593

  4. Matrix protein of vesicular stomatitis virus: a potent inhibitor of vascular endothelial growth factor and malignant ascites formation.

    Science.gov (United States)

    Zhou, Y; Wen, F; Zhang, P; Tang, R; Li, Q

    2013-03-01

    Malignant ascites is common in various types of cancers and is difficult to manage. Vascular endothelial growth factor (VEGF) has a pivotal role in malignant ascites. The matrix protein of vesicular stomatitis virus (VSVMP) has been shown to inhibit host gene expression and induce the apoptosis of cancer cells. The present study was designed to determine whether VSVMP suppresses the formation of ascites in ascites-producing peritoneal carcinomatosis. BALB/c female mice, 6-8 weeks old, bearing peritoneal tumors of H22 or MethA cells received an intraperitoneal administration of 50 μg VSVMP/250 μg liposome complexes, 50 μg empty plasmid/250 μg liposome complexes or 0.9% NaCl solution, respectively, every 2 days for 3 weeks. Administration of VSVMP resulted in a significant inhibition in ascites formation, improvement in health condition and prolonged survival of the treated mice. Decreased peritoneum osmolarity and reduced tumor vascularity coincided with dramatic reductions in the VEGF level in ascites fluid and plasma. Examination of floating tumor cells collected from the peritoneal wash revealed an apparently increased number of apoptotic cells and profound downregulation of VEGF mRNA in the VSVMP-treated mice. Our data indicate for the first time that in BALB/c mice bearing H22 or MethA cell peritoneal tumors, VSVMP may inhibit VEGF production and suppress angiogenesis, consequently abolishing ascites formation.

  5. [Effects of plant viruses on vector and non-vector herbivorous arthropods and their natural enemies: a mini review].

    Science.gov (United States)

    He, Xiao-Chan; Xu, Hong-Xing; Zhou, Xiao-Jun; Zheng, Xu-Song; Sun, Yu-Jian; Yang, Ya-Jun; Tian, Jun-Ce; Lü, Zhong-Xian

    2014-05-01

    Plant viruses transmitted by arthropods, as an important biotic factor, may not only directly affect the yield and quality of host plants, and development, physiological characteristics and ecological performances of their vector arthropods, but also directly or indirectly affect the non-vector herbivorous arthropods and their natural enemies in the same ecosystem, thereby causing influences to the whole agro-ecosystem. This paper reviewed the progress on the effects of plant viruses on herbivorous arthropods, including vector and non-vector, and their natural enemies, and on their ecological mechanisms to provide a reference for optimizing the management of vector and non-vector arthropod populations and sustainable control of plant viruses in agro-ecosystem.

  6. Status and prospects of plant virus control through interference with vector transmission

    NARCIS (Netherlands)

    Bragard, C.; Caciagli, P.; Lemaire, O.; Lopez-Moya, J.J.; MacFarlane, S.; Peters, D.; Susi, P.; Torrance, L.

    2013-01-01

    Most plant viruses rely on vector organisms for their plant-to-plant spread. Although there are many different natural vectors, few plant virus–vector systems have been well studied. This review describes our current understanding of virus transmission by aphids, thrips, whiteflies, leafhoppers,

  7. Vesicular stomatitis virus expressing a chimeric Sindbis glycoprotein containing an Fc antibody binding domain targets to Her2/neu overexpressing breast cancer cells

    International Nuclear Information System (INIS)

    Bergman, Ira; Whitaker-Dowling, Patricia; Gao Yanhua; Griffin, Judith A.; Watkins, Simon C.

    2003-01-01

    Vesicular stomatitis virus (VSV) is a candidate for development for cancer therapy. It is an oncolytic virus that is safe in humans. Recombinant virus can be made directly from plasmid components. We attempted to create a virus that targeted specifically to breast cancer cells. Nonreplicating and replicating pseudotype VSV were created whose only surface glycoprotein (gp) was a Sindbis gp, called Sindbis-ZZ, modified to severely reduce its native binding function and to contain the Fc-binding domain of Staphylococcus aureus protein A. When titered on Her2/neu overexpressing SKBR3 human breast cancer cells, pseudotype VSV coated with Sindbis-ZZ had 5 /ml. This work demonstrates the ability to easily create, directly from plasmid components, an oncolytic replicating VSV with a restricted host cell range

  8. A spatio-temporal analysis of matrix protein and nucleocapsid trafficking during vesicular stomatitis virus uncoating.

    Directory of Open Access Journals (Sweden)

    Chad E Mire

    2010-07-01

    Full Text Available To study VSV entry and the fate of incoming matrix (M protein during virus uncoating we used recombinant viruses encoding M proteins with a C-terminal tetracysteine tag that could be fluorescently labeled using biarsenical (Lumio compounds. We found that uncoating occurs early in the endocytic pathway and is inhibited by expression of dominant-negative (DN Rab5, but is not inhibited by DN-Rab7 or DN-Rab11. Uncoating, as defined by the separation of nucleocapsids from M protein, occurred between 15 and 20 minutes post-entry and did not require microtubules or an intact actin cytoskeleton. Unexpectedly, the bulk of M protein remained associated with endosomal membranes after uncoating and was eventually trafficked to recycling endosomes. Another small, but significant fraction of M distributed to nuclear pore complexes, which was also not dependent on microtubules or polymerized actin. Quantification of fluorescence from high-resolution confocal micrographs indicated that after membrane fusion, M protein diffuses across the endosomal membrane with a concomitant increase in fluorescence from the Lumio label which occurred soon after the release of RNPs into the cytoplasm. These data support a new model for VSV uncoating in which RNPs are released from M which remains bound to the endosomal membrane rather than the dissociation of M protein from RNPs after release of the complex into the cytoplasm following membrane fusion.

  9. Use of genetically modified viruses and genetically engineered virus-vector vaccines: environmental effects.

    Science.gov (United States)

    Chan, Vivian S W

    2006-11-01

    Despite major therapeutic advances, infectious diseases remain highly problematic. Recent advancements in technology in producing DNA-based vaccines, together with the growing knowledge of the immune system, have provided new insights into the identification of the epitopes needed to target the development of highly targeted vaccines. Genetically modified (GM) viruses and genetically engineered virus-vector vaccines possess significant unpredictability and a number of inherent harmful potential hazards. For all these vaccines, safety assessment concerning unintended and unwanted side effects with regard to targeted vaccinees has always been the main focus. Important questions concerning effects on nontargeted individuals within the same species or other species remain unknown. Horizontal transfer of genes, though lacking supportive experimental or epidemiological investigations, is well established. New hybrid virus progenies resulting from genetic recombination between genetically engineered vaccine viruses and their naturally occurring relatives may possess totally unpredictable characteristics with regard to host preferences and disease-causing potentials. Furthermore, when genetically modified or engineered virus particles break down in the environment, their nuclei acids are released. Appropriate risk management is the key to minimizing any potential risks to humans and environment resulting from the use of these GM vaccines. There is inadequate knowledge to define either the probability of unintended events or the consequences of genetic modifications. The objective of this article is to highlight the limitations in environmental risk assessment and raise awareness of the potential risks involving the use of genetically modified viruses and genetically engineered virus-vector vaccines.

  10. Adeno-Associated Virus Vectors (AAV Expressing Phenylalanine Hydroxylase (PAH

    Directory of Open Access Journals (Sweden)

    Ayşegül Akbay Yarpuzlu

    2009-06-01

    Full Text Available Recent articles have appeared in the literature reporting use of adeno-associated virus vectors (AAV expressing phenylalanine hydroxylase in animal trials and suggesting its use in treatment of phenylketonuria (PKU as a form of gene therapy However, agents used in gene therapy to deliver genes are not site-specific and DNA is may be put in the wrong place, causing damage to the organism. The adverse immunogenicity of AAVs also needs to be reconsidered. This letter is written to discuss present unreadiness for Phase 1 clinical trials of gene therapy of PKU. Turk Jem 2009; 13: 18-9

  11. Satellite panicum mosaic virus coat protein enhances the performance of plant virus gene vectors.

    Science.gov (United States)

    Everett, Anthany L; Scholthof, Herman B; Scholthof, Karen-Beth G

    2010-01-05

    The coat protein of satellite panicum mosaic virus (SPCP) is known to effectively protect its cognate RNA from deleterious events, and here, we tested its stabilizing potential for heterologous virus-based gene vectors in planta. In support of this, a Potato virus X (PVX) vector carrying the SPMV capsid protein (PVX-SPCP) gene was stable for at least three serial systemic passages through Nicotiana benthamiana. To test the effect of SPCP in trans, PVX-SPCP was co-inoculated onto N. benthamiana together with a Tomato bushy stunt virus (TBSV) vector carrying a green fluorescent protein (GFP) gene that normally does not support systemic GFP expression. In contrast, co-inoculation of TBSV-GFP plus PVX-SPCP resulted in GFP accumulation and concomitant green fluorescent spots in upper, non-inoculated leaves in a temperature-responsive manner. These results suggest that the multifaceted SPMV CP has intriguing effects on virus-host interactions that surface in heterologous systems.

  12. Complexes of vesicular stomatitis virus matrix protein with host Rae1 and Nup98 involved in inhibition of host transcription.

    Science.gov (United States)

    Rajani, Karishma R; Pettit Kneller, Elizabeth L; McKenzie, Margie O; Horita, David A; Chou, Jeff W; Lyles, Douglas S

    2012-09-01

    Vesicular stomatitis virus (VSV) suppresses antiviral responses in infected cells by inhibiting host gene expression at multiple levels, including transcription, nuclear cytoplasmic transport, and translation. The inhibition of host gene expression is due to the activity of the viral matrix (M) protein. Previous studies have shown that M protein interacts with host proteins Rae1 and Nup98 that have been implicated in regulating nuclear-cytoplasmic transport. However, Rae1 function is not essential for host mRNA transport, raising the question of how interaction of a viral protein with a host protein that is not essential for gene expression causes a global inhibition at multiple levels. We tested the hypothesis that there may be multiple M protein-Rae1 complexes involved in inhibiting host gene expression at multiple levels. Using size exclusion chromatography and sedimentation velocity analysis, it was determined that Rae1 exists in high, intermediate, and low molecular weight complexes. The intermediate molecular weight complexes containing Nup98 interacted most efficiently with M protein. The low molecular weight form also interacted with M protein in cells that overexpress Rae1 or cells in which Nup98 expression was silenced. Silencing Rae1 expression had little if any effect on nuclear accumulation of host mRNA in VSV-infected cells, nor did it affect VSV's ability to inhibit host translation. Instead, silencing Rae1 expression reduced the ability of VSV to inhibit host transcription. M protein interacted efficiently with Rae1-Nup98 complexes associated with the chromatin fraction of host nuclei, consistent with an effect on host transcription. These results support the idea that M protein-Rae1 complexes serve as platforms to promote the interaction of M protein with other factors involved in host transcription. They also support the idea that Rae1-Nup98 complexes play a previously under-appreciated role in regulation of transcription.

  13. Complexes of vesicular stomatitis virus matrix protein with host Rae1 and Nup98 involved in inhibition of host transcription.

    Directory of Open Access Journals (Sweden)

    Karishma R Rajani

    2012-09-01

    Full Text Available Vesicular stomatitis virus (VSV suppresses antiviral responses in infected cells by inhibiting host gene expression at multiple levels, including transcription, nuclear cytoplasmic transport, and translation. The inhibition of host gene expression is due to the activity of the viral matrix (M protein. Previous studies have shown that M protein interacts with host proteins Rae1 and Nup98 that have been implicated in regulating nuclear-cytoplasmic transport. However, Rae1 function is not essential for host mRNA transport, raising the question of how interaction of a viral protein with a host protein that is not essential for gene expression causes a global inhibition at multiple levels. We tested the hypothesis that there may be multiple M protein-Rae1 complexes involved in inhibiting host gene expression at multiple levels. Using size exclusion chromatography and sedimentation velocity analysis, it was determined that Rae1 exists in high, intermediate, and low molecular weight complexes. The intermediate molecular weight complexes containing Nup98 interacted most efficiently with M protein. The low molecular weight form also interacted with M protein in cells that overexpress Rae1 or cells in which Nup98 expression was silenced. Silencing Rae1 expression had little if any effect on nuclear accumulation of host mRNA in VSV-infected cells, nor did it affect VSV's ability to inhibit host translation. Instead, silencing Rae1 expression reduced the ability of VSV to inhibit host transcription. M protein interacted efficiently with Rae1-Nup98 complexes associated with the chromatin fraction of host nuclei, consistent with an effect on host transcription. These results support the idea that M protein-Rae1 complexes serve as platforms to promote the interaction of M protein with other factors involved in host transcription. They also support the idea that Rae1-Nup98 complexes play a previously under-appreciated role in regulation of transcription.

  14. Plasma membrane phosphatidylinositol 4,5 bisphosphate is required for internalization of foot-and-mouth disease virus and vesicular stomatitis virus.

    Directory of Open Access Journals (Sweden)

    Angela Vázquez-Calvo

    Full Text Available Phosphatidylinositol-4,5-bisphosphate, PI(4,5P(2, is a phospholipid which plays important roles in clathrin-mediated endocytosis. To investigate the possible role of this lipid on viral entry, two viruses important for animal health were selected: the enveloped vesicular stomatitis virus (VSV - which uses a well characterized clathrin mediated endocytic route - and two different variants of the non-enveloped foot-and-mouth disease virus (FMDV with distinct receptor specificities. The expression of a dominant negative dynamin, a PI(4,5P(2 effector protein, inhibited the internalization and infection of VSV and both FMDV isolates. Depletion of PI(4,5P(2 from plasma membrane using ionomycin or an inducible system, and inhibition of its de novo synthesis with 1-butanol revealed that VSV as well as FMDV C-S8c1, which uses integrins as receptor, displayed a high dependence on PI(4,5P(2 for internalization. Expression of a kinase dead mutant (KD of phosphatidylinositol-4-phosphate-5-kinase Iα (PIP5K-Iα, an enzyme responsible for PI(4,5P(2 synthesis that regulates clathrin-dependent endocytosis, also impaired entry and infection of VSV and FMDV C-S8c1. Interestingly FMDV MARLS variant that uses receptors other than integrins for cell entry was less sensitive to PI(4,5P(2 depletion, and was not inhibited by the expression of the KD PIP5K-Iα mutant suggesting the involvement of endocytic routes other than the clathrin-mediated on its entry. These results highlight the role of PI(4,5P(2 and PIP5K-Iα on clathrin-mediated viral entry.

  15. Enhancers Are Major Targets for Murine Leukemia Virus Vector Integration

    Science.gov (United States)

    De Ravin, Suk See; Su, Ling; Theobald, Narda; Choi, Uimook; Macpherson, Janet L.; Poidinger, Michael; Symonds, Geoff; Pond, Susan M.; Ferris, Andrea L.; Hughes, Stephen H.

    2014-01-01

    ABSTRACT Retroviral vectors have been used in successful gene therapies. However, in some patients, insertional mutagenesis led to leukemia or myelodysplasia. Both the strong promoter/enhancer elements in the long terminal repeats (LTRs) of murine leukemia virus (MLV)-based vectors and the vector-specific integration site preferences played an important role in these adverse clinical events. MLV integration is known to prefer regions in or near transcription start sites (TSS). Recently, BET family proteins were shown to be the major cellular proteins responsible for targeting MLV integration. Although MLV integration sites are significantly enriched at TSS, only a small fraction of the MLV integration sites (integration map of more than one million integration sites from CD34+ hematopoietic stem cells transduced with a clinically relevant MLV-based vector. The integration sites form ∼60,000 tight clusters. These clusters comprise ∼1.9% of the genome. The vast majority (87%) of the integration sites are located within histone H3K4me1 islands, a hallmark of enhancers. The majority of these clusters also have H3K27ac histone modifications, which mark active enhancers. The enhancers of some oncogenes, including LMO2, are highly preferred targets for integration without in vivo selection. IMPORTANCE We show that active enhancer regions are the major targets for MLV integration; this means that MLV preferentially integrates in regions that are favorable for viral gene expression in a variety of cell types. The results provide insights for MLV integration target site selection and also explain the high risk of insertional mutagenesis that is associated with gene therapy trials using MLV vectors. PMID:24501411

  16. Potato virus X and Tobacco mosaic virus-based vectors compatible with the Gateway-TM cloning system

    NARCIS (Netherlands)

    Lacorte, C.C.; Ribeiro, S.G.; Lohuis, H.; Goldbach, R.W.; Prins, M.W.

    2010-01-01

    Virus-based expression vectors are important tools for high-level production of foreign proteins and for gene function analysis through virus induced gene silencing. To exploit further their advantages as fast, high yield replicons, a set of vectors was produced by converting and adapting Potato

  17. Enhancing the Oncolytic Activity of CD133-Targeted Measles Virus: Receptor Extension or Chimerism with Vesicular Stomatitis Virus Are Most Effective

    Directory of Open Access Journals (Sweden)

    Dina Kleinlützum

    2017-06-01

    Full Text Available Therapy resistance and tumor recurrence are often linked to a small refractory and highly tumorigenic subpopulation of neoplastic cells, known as cancer stem cells (CSCs. A putative marker of CSCs is CD133 (prominin-1. We have previously described a CD133-targeted oncolytic measles virus (MV-CD133 as a promising approach to specifically eliminate CD133-positive tumor cells. Selectivity was introduced at the level of cell entry by an engineered MV hemagglutinin (H. The H protein was blinded for its native receptors and displayed a CD133-specific single-chain antibody fragment (scFv as targeting domain. Interestingly, MV-CD133 was more active in killing CD133-positive tumors than the unmodified MV-NSe despite being highly selective for its target cells. To further enhance the antitumoral activity of MV-CD133, we here pursued arming technologies, receptor extension, and chimeras between MV-CD133 and vesicular stomatitis virus (VSV. All newly generated viruses including VSV-CD133 were highly selective in eliminating CD133-positive cells. MV-CD46/CD133 killed in addition CD133-negative cells being positive for the MV receptors. In an orthotopic glioma model, MV-CD46/CD133 and MVSCD-CD133, which encodes the super cytosine deaminase, were most effective. Notably, VSV-CD133 caused fatal neurotoxicity in this tumor model. Use of CD133 as receptor could be excluded as being causative. In a subcutaneous tumor model of hepatocellular cancer, VSV-CD133 revealed the most potent oncolytic activity and also significantly prolonged survival of the mice when injected intravenously. Compared to MV-CD133, VSV-CD133 infected a more than 104-fold larger area of the tumor within the same time period. Our data not only suggest new concepts and approaches toward enhancing the oncolytic activity of CD133-targeted oncolytic viruses but also raise awareness about careful toxicity testing of novel virus types.

  18. Characterization of Vesicular Stomatitis Virus Pseudotypes Bearing Essential Entry Glycoproteins gB, gD, gH, and gL of Herpes Simplex Virus 1.

    Science.gov (United States)

    Rogalin, Henry B; Heldwein, Ekaterina E

    2016-11-15

    Herpes simplex viruses (HSVs) are unusual in that unlike most enveloped viruses, they require at least four entry glycoproteins, gB, gD, gH, and gL, for entry into target cells in addition to a cellular receptor for gD. The dissection of the herpes simplex virus 1 (HSV-1) entry mechanism is complicated by the presence of more than a dozen proteins on the viral envelope. To investigate HSV-1 entry requirements in a simplified system, we generated vesicular stomatitis virus (VSV) virions pseudotyped with HSV-1 essential entry glycoproteins gB, gD, gH, and gL but lacking the native VSV fusogen G. These virions, referred to here as VSVΔG-BHLD virions, infected a cell line expressing a gD receptor, demonstrating for the first time that the four essential entry glycoproteins of HSV-1 are not only required but also sufficient for cell entry. To our knowledge, this is the first time the VSV pseudotyping system has been successfully extended beyond two proteins. Entry of pseudotyped virions required a gD receptor and was inhibited by HSV-1 specific anti-gB or anti-gH/gL neutralizing antibodies, which suggests that membrane fusion during the entry of the pseudotyped virions shares common requirements with the membrane fusion involved in HSV-1 entry and HSV-1-mediated syncytium formation. The HSV pseudotyping system established in this study presents a novel tool for systematic exploration of the HSV entry and membrane fusion mechanisms. Herpes simplex viruses (HSVs) are human pathogens that can cause cold sores, genital herpes, and blindness. No vaccines or preventatives are available. HSV entry into cells-a prerequisite for a successful infection-is a complex process that involves multiple viral and host proteins and occurs by different routes. Detailed mechanistic knowledge of the HSV entry is important for understanding its pathogenesis and would benefit antiviral and vaccine development, yet the presence of more than a dozen proteins on the viral envelope complicates

  19. Virus-based transient expression vectors for woody crops: a new frontier for vector design and use.

    Science.gov (United States)

    Dawson, William O; Folimonova, Svetlana Y

    2013-01-01

    Virus-based expression vectors are commonplace tools for the production of proteins or the induction of RNA silencing in herbaceous plants. This review considers a completely different set of uses for viral vectors in perennial fruit and nut crops, which can be productive for periods of up to 100 years. Viral vectors could be used in the field to modify existing plants. Furthermore, with continually emerging pathogens and pests, viral vectors could express genes to protect the plants or even to treat plants after they become infected. As technologies develop during the life span of these crops, viral vectors can be used for adding new genes as an alternative to pushing up the crop and replanting with transgenic plants. Another value of virus-based vectors is that they add nothing permanently to the environment. This requires that effective and stable viral vectors be developed for specific crops from endemic viruses. Studies using viruses from perennial hosts suggest that these objectives could be accomplished.

  20. Use of Insecticide-Treated House Screens to Reduce Infestations of Dengue Virus Vectors, Mexico

    Science.gov (United States)

    Manrique-Saide, Pablo; Che-Mendoza, Azael; Barrera-Perez, Mario; Guillermo-May, Guillermo; Herrera-Bojorquez, Josue; Dzul-Manzanilla, Felipe; Gutierrez-Castro, Cipriano; Lenhart, Audrey; Vazquez-Prokopec, Gonzalo; Sommerfeld, Johannes; McCall, Philip J.; Kroeger, Axel

    2015-01-01

    Dengue prevention efforts rely on control of virus vectors. We investigated use of insecticide-treated screens permanently affixed to windows and doors in Mexico and found that the screens significantly reduced infestations of Aedes aegypti mosquitoes in treated houses. Our findings demonstrate the value of this method for dengue virus vector control. PMID:25625483

  1. Use of insecticide-treated house screens to reduce infestations of dengue virus vectors, Mexico.

    Science.gov (United States)

    Manrique-Saide, Pablo; Che-Mendoza, Azael; Barrera-Perez, Mario; Guillermo-May, Guillermo; Herrera-Bojorquez, Josue; Dzul-Manzanilla, Felipe; Gutierrez-Castro, Cipriano; Lenhart, Audrey; Vazquez-Prokopec, Gonzalo; Sommerfeld, Johannes; McCall, Philip J; Kroeger, Axel; Arredondo-Jimenez, Juan I

    2015-02-01

    Dengue prevention efforts rely on control of virus vectors. We investigated use of insecticide-treated screens permanently affixed to windows and doors in Mexico and found that the screens significantly reduced infestations of Aedes aegypti mosquitoes in treated houses. Our findings demonstrate the value of this method for dengue virus vector control.

  2. The Role of Bacterial Chaperones in the Circulative Transmission of Plant Viruses by Insect Vectors

    Directory of Open Access Journals (Sweden)

    Murad Ghanim

    2013-06-01

    Full Text Available Persistent circulative transmission of plant viruses involves complex interactions between the transmitted virus and its insect vector. Several studies have shown that insect vector proteins are involved in the passage and the transmission of the virus. Interestingly, proteins expressed by bacterial endosymbionts that reside in the insect vector, were also shown to influence the transmission of these viruses. Thus far, the transmission of two plant viruses that belong to different virus genera was shown to be facilitated by a bacterial chaperone protein called GroEL. This protein was shown to be implicated in the transmission of Potato leafroll virus (PLRV by the green peach aphid Myzus persicae, and the transmission of Tomato yellow leaf curl virus (TYLCV by the sweetpotato whitefly Bemisia tabaci. These tri-trophic levels of interactions and their possible evolutionary implications are reviewed.

  3. Interaction of measles virus vectors with Auger electron emitting radioisotopes

    International Nuclear Information System (INIS)

    Dingli, David; Peng, K.-W.; Harvey, Mary E.; Vongpunsawad, Sompong; Bergert, Elizabeth R.; Kyle, Robert A.; Cattaneo, Roberto; Morris, John C.; Russell, Stephen J.

    2005-01-01

    A recombinant measles virus (MV) expressing the sodium iodide symporter (NIS) is being considered for therapy of advanced multiple myeloma. Auger electrons selectively damage cells in which the isotope decays. We hypothesized that the Auger electron emitting isotope 125 I can be used to control viral proliferation. MV was engineered to express both carcinoembryonic antigen and NIS (MV-NICE). Cells were infected with MV-NICE and exposed to 125 I with appropriate controls. MV-NICE replication in vitro is inhibited by the selective uptake of 125 I by cells expressing NIS. Auger electron damage is partly mediated by free radicals and abrogated by glutathione. In myeloma xenografts, control of MV-NICE with 125 I was not possible under the conditions of the experiment. MV-NICE does not replicate faster in the presence of radiation. Auger electron emitting isotopes effectively stop propagation of MV vectors expressing NIS in vitro. Additional work is necessary to translate these observations in vivo

  4. Highly efficient virus-induced gene silencing in apple and soybean by apple latent spherical virus vector and biolistic inoculation.

    Science.gov (United States)

    Yamagishi, Noriko; Yoshikawa, Nobuyuki

    2013-01-01

    Virus-induced gene silencing (VIGS) is an effective tool for the analysis of the gene function in plants within a short time. However, in woody fruit tree like apple, some of Solanum crops, and soybean, it is generally difficult to inoculate virus vector by conventional inoculation methods. Here, we show efficient VIGS in apple and soybean by Apple latent spherical virus (ALSV) vector and biolistic inoculation. The plants inoculated with ALSV vectors by particle bombardment showed uniform silenced phenotypes of target genes within 2-3 weeks post inoculation.

  5. TMV-Gate vectors: Gateway compatible tobacco mosaic virus based expression vectors for functional analysis of proteins

    Science.gov (United States)

    Kagale, Sateesh; Uzuhashi, Shihomi; Wigness, Merek; Bender, Tricia; Yang, Wen; Borhan, M. Hossein; Rozwadowski, Kevin

    2012-01-01

    Plant viral expression vectors are advantageous for high-throughput functional characterization studies of genes due to their capability for rapid, high-level transient expression of proteins. We have constructed a series of tobacco mosaic virus (TMV) based vectors that are compatible with Gateway technology to enable rapid assembly of expression constructs and exploitation of ORFeome collections. In addition to the potential of producing recombinant protein at grams per kilogram FW of leaf tissue, these vectors facilitate either N- or C-terminal fusions to a broad series of epitope tag(s) and fluorescent proteins. We demonstrate the utility of these vectors in affinity purification, immunodetection and subcellular localisation studies. We also apply the vectors to characterize protein-protein interactions and demonstrate their utility in screening plant pathogen effectors. Given its broad utility in defining protein properties, this vector series will serve as a useful resource to expedite gene characterization efforts. PMID:23166857

  6. Virus-Induced Gene Silencing in Maize with a Foxtail mosaic virus Vector.

    Science.gov (United States)

    Mei, Yu; Whitham, Steven A

    2018-01-01

    Virus-induced gene silencing (VIGS) is a powerful technology for rapidly and transiently knocking down the expression of plant genes to study their functions. A VIGS vector for maize derived from Foxtail mosaic virus (FoMV), a positive-sense single-stranded RNA virus, was recently developed. A cloning site created near the 3' end of the FoMV genome enables insertion of 200-400 nucleotide fragments of maize genes targeted for silencing. The recombinant FoMV clones are inoculated into leaves of maize seedlings by biolistic particle delivery, and silencing is typically observed within 2 weeks after inoculation. This chapter provides a protocol for constructing FoMV VIGS clones and inoculating them into maize seedlings.

  7. Field Evaluation of a Multiplex Real-Time Reverse Transcription Polymerase Chain Reaction Assay for Detection of Vesicular Stomatitis Virus

    Science.gov (United States)

    Sporadic outbreaks of vesicular stomatitis (VS) in the United States result in significant economic losses for the US livestock industries because VS is an OIE reportable disease and also clinically mimics foot-and-mouth disease. Rapid and accurate differentiation of these two diseases is critical ...

  8. Eilat virus displays a narrow mosquito vector range.

    Science.gov (United States)

    Nasar, Farooq; Haddow, Andrew D; Tesh, Robert B; Weaver, Scott C

    2014-12-17

    Most alphaviruses are arthropod-borne and utilize mosquitoes as vectors for transmission to susceptible vertebrate hosts. This ability to infect both mosquitoes and vertebrates is essential for maintenance of most alphaviruses in nature. A recently characterized alphavirus, Eilat virus (EILV), isolated from a pool of Anopheles coustani s.I. is unable to replicate in vertebrate cell lines. The EILV host range restriction occurs at both attachment/entry as well as genomic RNA replication levels. Here we investigated the mosquito vector range of EILV in species encompassing three genera that are responsible for maintenance of other alphaviruses in nature. Susceptibility studies were performed in four mosquito species: Aedes albopictus, A. aegypti, Anopheles gambiae, and Culex quinquefasciatus via intrathoracic and oral routes utilizing EILV and EILV expressing red fluorescent protein (-eRFP) clones. EILV-eRFP was injected at 10(7) PFU/mL to visualize replication in various mosquito organs at 7 days post-infection. Mosquitoes were also injected with EILV at 10(4)-10(1) PFU/mosquito and virus replication was measured via plaque assays at day 7 post-infection. Lastly, mosquitoes were provided bloodmeals containing EILV-eRFP at doses of 10(9), 10(7), 10(5) PFU/mL, and infection and dissemination rates were determined at 14 days post-infection. All four species were susceptible via the intrathoracic route; however, replication was 10-100 fold less than typical for most alphaviruses, and infection was limited to midgut-associated muscle tissue and salivary glands. A. albopictus was refractory to oral infection, while A. gambiae and C. quinquefasciatus were susceptible only at 10(9) PFU/mL dose. In contrast, A. aegypti was susceptible at both 10(9) and 10(7) PFU/mL doses, with body infection rates of 78% and 63%, and dissemination rates of 26% and 8%, respectively. The exclusion of vertebrates in its maintenance cycle may have facilitated the adaptation of EILV to a single

  9. Tubular structure induced by a plant virus facilitates viral spread in its vector insect.

    Directory of Open Access Journals (Sweden)

    Qian Chen

    Full Text Available Rice dwarf virus (RDV replicates in and is transmitted by a leafhopper vector in a persistent-propagative manner. Previous cytopathologic and genetic data revealed that tubular structures, constructed by the nonstructural viral protein Pns10, contain viral particles and are directly involved in the intercellular spread of RDV among cultured leafhopper cells. Here, we demonstrated that RDV exploited these virus-containing tubules to move along actin-based microvilli of the epithelial cells and muscle fibers of visceral muscle tissues in the alimentary canal, facilitating the spread of virus in the body of its insect vector leafhoppers. In cultured leafhopper cells, the knockdown of Pns10 expression due to RNA interference (RNAi induced by synthesized dsRNA from Pns10 gene strongly inhibited tubule formation and prevented the spread of virus among insect vector cells. RNAi induced after ingestion of dsRNA from Pns10 gene strongly inhibited formation of tubules, preventing intercellular spread and transmission of the virus by the leafhopper. All these results, for the first time, show that a persistent-propagative virus exploits virus-containing tubules composed of a nonstructural viral protein to traffic along actin-based cellular protrusions, facilitating the intercellular spread of the virus in the vector insect. The RNAi strategy and the insect vector cell culture provide useful tools to investigate the molecular mechanisms enabling efficient transmission of persistent-propagative plant viruses by vector insects.

  10. Varroa destructor, a potential vector of Israeli Acute Paralysis Virus in honey bees, Apis mellifera

    Science.gov (United States)

    Although the role of the parasitic mite, Varroa destructor, as a vector in transmission of viruses between honey bees is well established, no study has shown that it can similarly transmit Israeli Acute Paralysis Virus (IAPV), a virus that was found to be associated with Colony Collapse Disorder (CC...

  11. The Function of Herpes Simplex Virus Genes: A Primer for Genetic Engineering of Novel Vectors

    Science.gov (United States)

    Roizman, Bernard

    1996-10-01

    Herpes simplex virus vectors are being developed for delivery and expression of human genes to the central nervous system, selective destruction of cancer cells, and as carriers for genes encoding antigens that induce protective immunity against infectious agents. Vectors constructed to meet these objectives must differ from wild-type virus with respect to host range, reactivation from latency, and expression of viral genes. The vectors currently being developed are (i) helper free amplicons, (ii) replication defective viruses, and (iii) genetically engineered replication competent viruses with restricted host range. Whereas the former two types of vectors require stable, continuous cell lines expressing viral genes for their replication, the replication competent viruses will replicate on approved primary human cell strains.

  12. Efficient Strategy to Generate a Vectored Duck Enteritis Virus Delivering Envelope of Duck Tembusu Virus

    Directory of Open Access Journals (Sweden)

    Zhong Zou

    2014-06-01

    Full Text Available Duck Tembusu virus (DTMUV is a recently emerging pathogenic flavivirus that has resulted in a huge economic loss in the duck industry. However, no vaccine is currently available to control this pathogen. Consequently, a practical strategy to construct a vaccine against this pathogen should be determined. In this study, duck enteritis virus (DEV was examined as a candidate vaccine vector to deliver the envelope (E of DTMUV. A modified mini-F vector was inserted into the SORF3 and US2 gene junctions of the attenuated DEV vaccine strain C-KCE genome to generate an infectious bacterial artificial chromosome (BAC of C-KCE (vBAC-C-KCE. The envelope (E gene of DTMUV was inserted into the C-KCE genome through the mating-assisted genetically integrated cloning (MAGIC strategy, resulting in the recombinant vector, pBAC-C-KCE-E. A bivalent vaccine C-KCE-E was generated by eliminating the BAC backbone. Immunofluorescence and western blot analysis results indicated that the E proteins were vigorously expressed in C-KCE-E-infected chicken embryo fibroblasts (CEFs. Duck experiments demonstrated that the insertion of the E gene did not alter the protective efficacy of C-KCE. Moreover, C-KCE-E-immunized ducks induced neutralization antibodies against DTMUV. These results demonstrated, for the first time, that recombinant C-KCE-E can serve as a potential bivalent vaccine against DEV and DTMUV.

  13. Vectores recombinantes basados en el virus Vaccinia modificado de Ankara (MVA) como vacunas contra la leishmaniasis

    OpenAIRE

    Pérez Jiménez, Eva; Larraga, Vicente; Esteban, Mariano

    2005-01-01

    Vectores recombinantes basados en el virus vaccinia modificado de Ankara (MVA) como vacunas contra la leishmaniasis. Los vectores de la invención contienen secuencias codificantes de la proteína LACK, preferentemente insertadas en el locus de hemaglutinina del virus y bajo el control de un promotor que permite su expresión a lo largo del ciclo de infección del virus. Son vectores seguros, estables, que dan lugar a una potente respuesta inmune que confiere protección frente a la leishmaniasis,...

  14. Enhanced vesicular stomatitis virus (VSVΔ51 targeting of head and neck cancer in combination with radiation therapy or ZD6126 vascular disrupting agent

    Directory of Open Access Journals (Sweden)

    Alajez Nehad M

    2012-06-01

    Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC is the 5th most common cancer worldwide. Locally advanced HNSCC are treated with either radiation or chemo-radiotherapy, but still associated with high mortality rate, underscoring the need to develop novel therapies. Oncolytic viruses have been garnering increasing interest as anti-cancer agents due to their preferential killing of transformed cells. In this study, we evaluated the therapeutic potential of mutant vesicular stomatitis virus (VSVΔ51 against the human hypopharyngeal FaDu tumour model in vitro and in vivo. Results Our data demonstrated high toxicity of the virus against FaDu cells in vitro, which was associated with induction of apoptosis. In vivo, systemic injection of 1 × 109 pfu had minimal effect on tumour growth; however, when combined with two doses of ionizing radiation (IR; 5 Gy each or a single injection of the vascular disrupting agent (ZD6126, the virus exhibited profound suppression of tumour growth, which translated to a prolonged survival in the treated mice. Concordantly, VSVΔ51 combined with ZD6126 led to a significant increase in viral replication in these tumours. Conclusions Our data suggest that the combinations of VSVΔ51 with either IR or ZD6126 are potentially novel therapeutic opportunities for HNSCC.

  15. Novel strategy for generation and titration of recombinant adeno-associated virus vectors.

    Science.gov (United States)

    Shiau, Ai-Li; Liu, Pu-Ste; Wu, Chao-Liang

    2005-01-01

    Recombinant adeno-associated virus (rAAV) vectors have many advantages for gene therapeutic applications compared with other vector systems. Several methods that use plasmids or helper viruses have been reported for the generation of rAAV vectors. Unfortunately, the preparation of large-scale rAAV stocks is labor-intensive. Moreover, the biological titration of rAAV is still difficult, which may limit its preclinical and clinical applications. For this study, we developed a novel strategy to generate and biologically titrate rAAV vectors. A recombinant pseudorabies virus (PrV) with defects in its gD, gE, and thymidine kinase genes was engineered to express the AAV rep and cap genes, yielding PS virus, which served as a packaging and helper virus for the generation of rAAV vectors. PS virus was useful not only for generating high-titer rAAV vectors by cotransfection with an rAAV vector plasmid, but also for amplifying rAAV stocks. Notably, the biological titration of rAAV vectors was also feasible when cells were coinfected with rAAV and PS virus. Based on this strategy, we produced an rAAV that expresses prothymosin alpha (ProT). Expression of the ProT protein in vitro and in vivo mediated by rAAV/ProT gene transfer was detected by immunohistochemistry and a bioassay. Taken together, our results demonstrate that the PrV vector-based system is useful for generating rAAV vectors carrying various transgenes.

  16. An Improved Brome mosaic virus Silencing Vector: Greater Insert Stability and More Extensive VIGS1[OPEN

    Science.gov (United States)

    2018-01-01

    Virus-induced gene silencing (VIGS) is used extensively for gene function studies in plants. VIGS is inexpensive and rapid compared with silencing conducted through stable transformation, but many virus-silencing vectors, especially in grasses, induce only transient silencing phenotypes. A major reason for transient phenotypes is the instability of the foreign gene fragment (insert) in the vector during VIGS. Here, we report the development of a Brome mosaic virus (BMV)-based vector that better maintains inserts through modification of the original BMV vector RNA sequence. Modification of the BMV RNA3 sequence yielded a vector, BMVCP5, that better maintained phytoene desaturase and heat shock protein70-1 (HSP70-1) inserts in Nicotiana benthamiana and maize (Zea mays). Longer maintenance of inserts was correlated with greater target gene silencing and more extensive visible silencing phenotypes displaying greater tissue penetration and involving more leaves. The modified vector accumulated similarly to the original vector in N. benthamiana after agroinfiltration, thus maintaining a high titer of virus in this intermediate host used to produce virus inoculum for grass hosts. For HSP70, silencing one family member led to a large increase in the expression of another family member, an increase likely related to the target gene knockdown and not a general effect of virus infection. The cause of the increased insert stability in the modified vector is discussed in relationship to its recombination and accumulation potential. The modified vector will improve functional genomic studies in grasses, and the conceptual methods used to improve the vector may be applied to other VIGS vectors. PMID:29127260

  17. An ImprovedBrome mosaic virusSilencing Vector: Greater Insert Stability and More Extensive VIGS.

    Science.gov (United States)

    Ding, Xin Shun; Mannas, Stephen W; Bishop, Bethany A; Rao, Xiaolan; Lecoultre, Mitchell; Kwon, Soonil; Nelson, Richard S

    2018-01-01

    Virus-induced gene silencing (VIGS) is used extensively for gene function studies in plants. VIGS is inexpensive and rapid compared with silencing conducted through stable transformation, but many virus-silencing vectors, especially in grasses, induce only transient silencing phenotypes. A major reason for transient phenotypes is the instability of the foreign gene fragment (insert) in the vector during VIGS. Here, we report the development of a Brome mosaic virus (BMV)-based vector that better maintains inserts through modification of the original BMV vector RNA sequence. Modification of the BMV RNA3 sequence yielded a vector, BMVCP5, that better maintained phytoene desaturase and heat shock protein70-1 ( HSP70-1 ) inserts in Nicotiana benthamiana and maize ( Zea mays ). Longer maintenance of inserts was correlated with greater target gene silencing and more extensive visible silencing phenotypes displaying greater tissue penetration and involving more leaves. The modified vector accumulated similarly to the original vector in N. benthamiana after agroinfiltration, thus maintaining a high titer of virus in this intermediate host used to produce virus inoculum for grass hosts. For HSP70 , silencing one family member led to a large increase in the expression of another family member, an increase likely related to the target gene knockdown and not a general effect of virus infection. The cause of the increased insert stability in the modified vector is discussed in relationship to its recombination and accumulation potential. The modified vector will improve functional genomic studies in grasses, and the conceptual methods used to improve the vector may be applied to other VIGS vectors. © 2018 American Society of Plant Biologists. All Rights Reserved.

  18. Virus-induced gene silencing in diverse maize lines using the Brome Mosaic virus-based silencing vector

    Science.gov (United States)

    Virus-induced gene silencing (VIGS) is a widely used tool for gene function studies in many plant species, though its use in monocots has been limited. Using a Brome mosaic virus (BMV) vector designed to silence the maize phytoene desaturase gene, a genetically diverse set of maize inbred lines was ...

  19. Competence of Aedes aegypti, Ae. albopictus, and Culex quinquefasciatus Mosquitoes as Zika Virus Vectors, China.

    Science.gov (United States)

    Liu, Zhuanzhuan; Zhou, Tengfei; Lai, Zetian; Zhang, Zhenhong; Jia, Zhirong; Zhou, Guofa; Williams, Tricia; Xu, Jiabao; Gu, Jinbao; Zhou, Xiaohong; Lin, Lifeng; Yan, Guiyun; Chen, Xiao-Guang

    2017-07-01

    In China, the prevention and control of Zika virus disease has been a public health threat since the first imported case was reported in February 2016. To determine the vector competence of potential vector mosquito species, we experimentally infected Aedes aegypti, Ae. albopictus, and Culex quinquefasciatus mosquitoes and determined infection rates, dissemination rates, and transmission rates. We found the highest vector competence for the imported Zika virus in Ae. aegypti mosquitoes, some susceptibility of Ae. albopictus mosquitoes, but no transmission ability for Cx. quinquefasciatus mosquitoes. Considering that, in China, Ae. albopictus mosquitoes are widely distributed but Ae. aegypti mosquito distribution is limited, Ae. albopictus mosquitoes are a potential primary vector for Zika virus and should be targeted in vector control strategies.

  20. Competence of Aedes aegypti, Ae. albopictus, and Culex quinquefasciatus Mosquitoes as Zika Virus Vectors, China

    Science.gov (United States)

    Liu, Zhuanzhuan; Zhou, Tengfei; Lai, Zetian; Zhang, Zhenhong; Jia, Zhirong; Zhou, Guofa; Williams, Tricia; Xu, Jiabao; Gu, Jinbao; Zhou, Xiaohong; Lin, Lifeng; Yan, Guiyun

    2017-01-01

    In China, the prevention and control of Zika virus disease has been a public health threat since the first imported case was reported in February 2016. To determine the vector competence of potential vector mosquito species, we experimentally infected Aedes aegypti, Ae. albopictus, and Culex quinquefasciatus mosquitoes and determined infection rates, dissemination rates, and transmission rates. We found the highest vector competence for the imported Zika virus in Ae. aegypti mosquitoes, some susceptibility of Ae. albopictus mosquitoes, but no transmission ability for Cx. quinquefasciatus mosquitoes. Considering that, in China, Ae. albopictus mosquitoes are widely distributed but Ae. aegypti mosquito distribution is limited, Ae. albopictus mosquitoes are a potential primary vector for Zika virus and should be targeted in vector control strategies. PMID:28430562

  1. Differential profiles of direct and indirect modification of vector feeding behaviour by a plant virus

    Science.gov (United States)

    He, Wen-Bo; Li, Jie; Liu, Shu-Sheng

    2015-01-01

    Plant viruses interact with their insect vectors directly and indirectly via host plants, and this tripartite interaction may produce fitness benefits to both the vectors and the viruses. Our previous studies show that the Middle East-Asia Minor 1 (MEAM1) species of the whitefly Bemisia tabaci complex improved its performance on tobacco plants infected by the Tomato yellow leaf curl China virus (TYLCCNV), which it transmits, although virus infection of the whitefly per se reduced its performance. Here, we use electrical penetration graph recording to investigate the direct and indirect effects of TYLCCNV on the feeding behaviour of MEAM1. When feeding on either cotton, a non-host of TYLCCNV, or uninfected tobacco, a host of TYLCCNV, virus-infection of the whiteflies impeded their feeding. Interestingly, when viruliferous whiteflies fed on virus-infected tobacco, their feeding activities were no longer negatively affected; instead, the virus promoted whitefly behaviour related to rapid and effective sap ingestion. Our findings show differential profiles of direct and indirect modification of vector feeding behaviour by a plant virus, and help to unravel the behavioural mechanisms underlying a mutualistic relationship between an insect vector and a plant virus that also has features reminiscent of an insect pathogen. PMID:25567524

  2. Vector-Borne Transmission Imposes a Severe Bottleneck on an RNA Virus Population

    OpenAIRE

    Forrester, Naomi L.; Guerbois, Mathilde; Seymour, Robert L.; Spratt, Heidi; Weaver, Scott C.

    2012-01-01

    RNA viruses typically occur in genetically diverse populations due to their error-prone genome replication. Genetic diversity is thought to be important in allowing RNA viruses to explore sequence space, facilitating adaptation to changing environments and hosts. Some arboviruses that infect both a mosquito vector and a mammalian host are known to experience population bottlenecks in their vectors, which may constrain their genetic diversity and could potentially lead to extinction events via...

  3. Replication of Tomato Yellow Leaf Curl Virus in Its Whitefly Vector, Bemisia tabaci.

    Science.gov (United States)

    Pakkianathan, Britto Cathrin; Kontsedalov, Svetlana; Lebedev, Galina; Mahadav, Assaf; Zeidan, Muhammad; Czosnek, Henryk; Ghanim, Murad

    2015-10-01

    Tomato yellow leaf curl virus (TYLCV) is a begomovirus transmitted exclusively by the whitefly Bemisia tabaci in a persistent, circulative manner. Replication of TYLCV in its vector remains controversial, and thus far, the virus has been considered to be nonpropagative. Following 8 h of acquisition on TYLCV-infected tomato plants or purified virions and then transfer to non-TYLCV-host cotton plants, the amounts of virus inside whitefly adults significantly increased (>2-fold) during the first few days and then continuously decreased, as measured by the amounts of genes on both virus DNA strands. Reported alterations in insect immune and defense responses upon virus retention led us to hypothesize a role for the immune response in suppressing virus replication. After virus acquisition, stress conditions were imposed on whiteflies, and the levels of three viral gene sequences were measured over time. When whiteflies were exposed to TYLCV and treatment with two different pesticides, the virus levels continuously increased. Upon exposure to heat stress, the virus levels gradually decreased, without any initial accumulation. Switching of whiteflies between pesticide, heat stress, and control treatments caused fluctuating increases and decreases in virus levels. Fluorescence in situ hybridization analysis confirmed these results and showed virus signals inside midgut epithelial cell nuclei. Combining the pesticide and heat treatments with virus acquisition had significant effects on fecundity. Altogether, our results demonstrate for the first time that a single-stranded DNA plant virus can replicate in its hemipteran vector. Plant viruses in agricultural crops are of great concern worldwide. Many of them are transmitted from infected to healthy plants by insects. Persistently transmitted viruses often have a complex association with their vectors; however, most are believed not to replicate within these vectors. Such replication is important, as it contributes to the

  4. Parainfluenza virus 5-vectored vaccines against human and animal infectious diseases.

    Science.gov (United States)

    Chen, Zhenhai

    2018-03-01

    Parainfluenza virus 5 (PIV5), known as canine parainfluenza virus in the veterinary field, is a negative-sense, nonsegmented, single-stranded RNA virus belonging to the Paramyxoviridae family. Parainfluenza virus 5 is an excellent viral vector and has been used as a live vaccine for kennel cough for many years in dogs without any safety concern. It can grow to high titers in many cell types, and its genome is stable even in the presence of foreign gene insertions. So far, PIV5 has been used to develop vaccines against influenza virus, respiratory syncytial virus, rabies virus, and Mycobacterium tuberculosis, demonstrating its ability to elicit robust and protective immune responses in preclinical animal models. Parainfluenza virus 5-based vaccines can be administered intranasally, intramuscularly, or orally. Interestingly, prior exposure of PIV5 does not prevent a PIV5-vectored vaccine from generating robust immunity, indicating that the vector can be used more than once. Here, these encouraging results are reviewed together along with discussion of the desirable advantages of the PIV5 vaccine vector to aid future vaccine design and to accelerate progression of PIV5-based vaccines into clinical trials. Copyright © 2018 John Wiley & Sons, Ltd.

  5. Lack of correlation between virus barosensitivity and the presence of a viral envelope during inactivation of human rotavirus, vesicular stomatitis virus, and avian metapneumovirus by high-pressure processing.

    Science.gov (United States)

    Lou, Fangfei; Neetoo, Hudaa; Li, Junan; Chen, Haiqiang; Li, Jianrong

    2011-12-01

    High-pressure processing (HPP) is a nonthermal technology that has been shown to effectively inactivate a wide range of microorganisms. However, the effectiveness of HPP on inactivation of viruses is relatively less well understood. We systematically investigated the effects of intrinsic (pH) and processing (pressure, time, and temperature) parameters on the pressure inactivation of a nonenveloped virus (human rotavirus [HRV]) and two enveloped viruses (vesicular stomatitis virus [VSV] and avian metapneumovirus [aMPV]). We demonstrated that HPP can efficiently inactivate all tested viruses under optimal conditions, although the pressure susceptibilities and the roles of temperature and pH substantially varied among these viruses regardless of the presence of a viral envelope. We found that VSV was much more stable than most food-borne viruses, whereas aMPV was highly susceptible to HPP. When viruses were held for 2 min under 350 MPa at 4°C, 1.1-log, 3.9-log, and 5.0-log virus reductions were achieved for VSV, HRV, and aMPV, respectively. Both VSV and aMPV were more susceptible to HPP at higher temperature and lower pH. In contrast, HRV was more easily inactivated at higher pH, although temperature did not have a significant impact on inactivation. Furthermore, we demonstrated that the damage of virion structure by disruption of the viral envelope and/or capsid is the primary mechanism underlying HPP-induced viral inactivation. In addition, VSV glycoprotein remained antigenic although VSV was completely inactivated. Taken together, our findings suggest that HPP is a promising technology to eliminate viral contaminants in high-risk foods, water, and other fomites.

  6. Ebola virus and arthropods: a literature review and entomological consideration on the vector role.

    Science.gov (United States)

    Dutto, M; Bertero, M; Petrosillo, N; Pombi, M; Otranto, D

    2016-10-01

    Ebola virus is a pathogen responsible for a severe disease that affects humans and several animal species. To date, the natural reservoir of this virus is not known with certainty, although it is believed that fruit bats (Chiroptera: Pteropodidae) play an important role in maintaining the virus in nature. Although information on viral transmission from animals to humans is not clear, the role of arthropods has come under suspicion. In this article, we review the potential role of arthropods in spreading Ebola virus, acting as mechanical or biological vectors.

  7. A novel method for analysis of membrane microdomains: vesicular stomatitis virus glycoprotein microdomains change in size during infection, and those outside of budding sites resemble sites of virus budding

    International Nuclear Information System (INIS)

    Brown, Erica L.; Lyles, Douglas S.

    2003-01-01

    Membrane proteins, including viral envelope glycoproteins, may be organized into areas of locally high concentration, commonly referred to as membrane microdomains. Some viruses bud from detergent-resistant microdomains referred to as lipid rafts. However, vesicular stomatitis virus (VSV) serves as a prototype for viruses that bud from areas of plasma membrane that are not detergent resistant. We developed a new analytical method for immunoelectron microscopy data to determine whether the VSV envelope glycoprotein (G protein) is organized into plasma membrane microdomains. This method was used to quantify the distribution of the G protein in microdomains in areas of plasma membrane that did not contain budding sites. These microdomains were compared to budding virus envelopes to address the question of whether G protein-containing microdomains were formed only at the sites of budding. At early times postinfection, most of the G protein was organized into membrane microdomains outside of virus budding sites that were approximately 100-150 nm, with smaller amounts distributed into larger microdomains. In contrast to early times postinfection, the increased level of G protein in the host plasma membrane at later times postinfection led to distribution of G protein among membrane microdomains of a wider variety of sizes, rather than a higher G protein concentration in the 100- to 150-nm microdomains. VSV budding occurred in G protein-containing microdomains with a range of sizes, some of which were smaller than the virus envelope. These microdomains extended in size to a maximum of 300-400 nm from the tip of the budding virion. The data support a model for virus assembly in which G protein organizes into membrane microdomains that resemble virus envelopes prior to formation of budding sites, and these microdomains serve as the sites of assembly of internal virion components

  8. A recombinant vesicular stomatitis virus encoding CCR5-tropic HIV-1 receptors targets HIV-1-infected cells and controls HIV-1 infection.

    Science.gov (United States)

    Okuma, Kazu; Fukagawa, Koji; Kohma, Takuya; Takahama, Youichi; Hamaguchi, Yukio; Ito, Mamoru; Tanaka, Yuetsu; Buonocore, Linda; Rose, John K; Hamaguchi, Isao

    Anti-retroviral therapy is useful to treat human immunodeficiency virus type 1 (HIV-1)-infected individuals, but has some major problems, such as the generation of multidrug-resistant viruses. To develop a novel supplemental or alternative therapeutic for CCR5-tropic (R5) HIV-1 infection, we generated a recombinant vesicular stomatitis virus (rVSV) in which the gene encoding its envelope glycoprotein (G) was replaced with the genes encoding R5 HIV-1 receptors (human CD4 and CCR5), designated VSVΔG-CC5. Our present data demonstrate that this rVSV specifically infects cells that are transiently expressing R5 HIV-1 envelope glycoproteins, but does not infect those expressing CXCR4-tropic HIV-1 envelope glycoproteins. Notably, after a CD4 + CCR5 + T cell line or primary cells initially infected with R5 HIV-1 were inoculated with G-complemented VSVΔG-CC5, the rVSV significantly reduced the number of HIV-1-infected cells, probably through direct targeting of the rVSV and VSV-mediated cytolysis and/or through syncytium formation- or cell-cell fusion-dependent killing, and markedly inhibited HIV-1 production. Furthermore, G-complemented VSVΔG-CC5 also efficiently inhibited HIV-1 infection in R5 HIV-1-infected humanized immunodeficient mice. Taken together, our findings indicate that a cytolytic rVSV that targets and eliminates R5 HIV-1-infected cells potentially has therapeutic value for controlling R5 HIV-1 infection. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  9. Effects of Zika Virus Strain and Aedes Mosquito Species on Vector Competence

    Science.gov (United States)

    Bialosuknia, Sean M.; Zink, Steven D.; Brecher, Matthew; Ehrbar, Dylan J.; Morrissette, Madeline N.; Kramer, Laura D.

    2017-01-01

    In the Western Hemisphere, Zika virus is thought to be transmitted primarily by Aedes aegypti mosquitoes. To determine the extent to which Ae. albopictus mosquitoes from the United States are capable of transmitting Zika virus and the influence of virus dose, virus strain, and mosquito species on vector competence, we evaluated multiple doses of representative Zika virus strains in Ae. aegypti and Ae. albopictus mosquitoes. Virus preparation (fresh vs. frozen) significantly affected virus infectivity in mosquitoes. We calculated 50% infectious doses to be 6.1–7.5 log10 PFU/mL; minimum infective dose was 4.2 log10 PFU/mL. Ae. albopictus mosquitoes were more susceptible to infection than Ae. aegypti mosquitoes, but transmission efficiency was higher for Ae. aegypti mosquitoes, indicating a transmission barrier in Ae. albopictus mosquitoes. Results suggest that, although Zika virus transmission is relatively inefficient overall and dependent on virus strain and mosquito species, Ae. albopictus mosquitoes could become major vectors in the Americas. PMID:28430564

  10. Plant virus expression vectors set the stage as production platforms for biopharmaceutical proteins.

    Science.gov (United States)

    Hefferon, Kathleen Laura

    2012-11-10

    Transgenic plants present enormous potential as a cost-effective and safe platform for large-scale production of vaccines and other therapeutic proteins. A number of different technologies are under development for the production of pharmaceutical proteins from plant tissues. One method used to express high levels of protein in plants involves the employment of plant virus expression vectors. Plant virus vectors have been designed to carry vaccine epitopes as well as full therapeutic proteins such as monoclonal antibodies in plant tissue both safely and effectively. Biopharmaceuticals such as these offer enormous potential on many levels, from providing relief to those who have little access to modern medicine, to playing an active role in the battle against cancer. This review describes the current design and status of plant virus expression vectors used as production platforms for biopharmaceutical proteins. Copyright © 2012 Elsevier Inc. All rights reserved.

  11. Foamy virus vectors expressing anti-HIV transgenes efficiently block HIV-1 replication.

    Science.gov (United States)

    Taylor, Jason A; Vojtech, Lucia; Bahner, Ingrid; Kohn, Donald B; Laer, Dorothee Von; Russell, David W; Richard, Robert E

    2008-01-01

    Gene therapy has the potential to control human immunodeficiency virus (HIV) in patients who do not respond to traditional antiviral therapy. In this study, we tested foamy virus (FV) vectors expressing three anti-HIV transgenes, both individually and in a combination vector. The transgenes tested in this study are RevM10, a dominant negative version of the viral rev protein, Sh1, a short hairpin RNA directed against a conserved overlapping sequence of tat and rev, and membrane-associated C46 (maC46), a membrane-attached peptide that blocks HIV cell entry. FV vectors efficiently transduce hematopoietic stem cells and, unlike lentivirus (LV) vectors, do not share viral proteins with HIV. The titers of the FV vectors described in this study were not affected by anti-HIV transgenes. On a direct comparison of FV vectors expressing the individual transgenes, entry inhibition using the maC46 transgene was found to be the most effective at blocking HIV replication. A clinically relevant FV vector expressing three anti-HIV transgenes effectively blocked HIV infection in primary macrophages derived from transduced, peripheral blood CD34-selected cells and in a cell line used for propagating HIV, CEMx174. These results suggest that there are potential benefits of using FV vectors in HIV gene therapy.

  12. Gene therapy model of X-linked severe combined immunodeficiency using a modified foamy virus vector.

    Science.gov (United States)

    Horino, Satoshi; Uchiyama, Toru; So, Takanori; Nagashima, Hiroyuki; Sun, Shu-Lan; Sato, Miki; Asao, Atsuko; Haji, Yoichi; Sasahara, Yoji; Candotti, Fabio; Tsuchiya, Shigeru; Kure, Shigeo; Sugamura, Kazuo; Ishii, Naoto

    2013-01-01

    X-linked severe combined immunodeficiency (SCID-X1) is an inherited genetic immunodeficiency associated with mutations in the common cytokine receptor γ chain (γc) gene, and characterized by a complete defect of T and natural killer (NK) cells. Gene therapy for SCID-X1 using conventional retroviral (RV) vectors carrying the γc gene results in the successful reconstitution of T cell immunity. However, the high incidence of vector-mediated T cell leukemia, caused by vector insertion near or within cancer-related genes has been a serious problem. In this study, we established a gene therapy model of mouse SCID-X1 using a modified foamy virus (FV) vector expressing human γc. Analysis of vector integration in a human T cell line demonstrated that the FV vector integration sites were significantly less likely to be located within or near transcriptional start sites than RV vector integration sites. To evaluate the therapeutic efficacy, bone marrow cells from γc-knockout (γc-KO) mice were infected with the FV vector and transplanted into γc-KO mice. Transplantation of the FV-treated cells resulted in the successful reconstitution of functionally active T and B cells. These data suggest that FV vectors can be effective and may be safer than conventional RV vectors for gene therapy for SCID-X1.

  13. Gene therapy model of X-linked severe combined immunodeficiency using a modified foamy virus vector.

    Directory of Open Access Journals (Sweden)

    Satoshi Horino

    Full Text Available X-linked severe combined immunodeficiency (SCID-X1 is an inherited genetic immunodeficiency associated with mutations in the common cytokine receptor γ chain (γc gene, and characterized by a complete defect of T and natural killer (NK cells. Gene therapy for SCID-X1 using conventional retroviral (RV vectors carrying the γc gene results in the successful reconstitution of T cell immunity. However, the high incidence of vector-mediated T cell leukemia, caused by vector insertion near or within cancer-related genes has been a serious problem. In this study, we established a gene therapy model of mouse SCID-X1 using a modified foamy virus (FV vector expressing human γc. Analysis of vector integration in a human T cell line demonstrated that the FV vector integration sites were significantly less likely to be located within or near transcriptional start sites than RV vector integration sites. To evaluate the therapeutic efficacy, bone marrow cells from γc-knockout (γc-KO mice were infected with the FV vector and transplanted into γc-KO mice. Transplantation of the FV-treated cells resulted in the successful reconstitution of functionally active T and B cells. These data suggest that FV vectors can be effective and may be safer than conventional RV vectors for gene therapy for SCID-X1.

  14. Vector competence of Brazilian Aedes aegypti and Ae. albopictus for a Brazilian yellow fever virus isolate.

    Science.gov (United States)

    Johnson, Barbara W; Chambers, Trudy V; Crabtree, Mary B; Filippis, Ana M B; Vilarinhos, Paulo T R; Resende, Marcelo C; Macoris, Maria de Lourdes G; Miller, Barry R

    2002-01-01

    Because the potential urban yellow fever (YF) mosquito vectors Aedes aegypti and Ae. albopictus are at historical highs in Brazil, both in terms of density and geographical range, we assessed the risk of an urban YF epidemic in Brazil. We evaluated and confirmed in a laboratory setting the vector competence of Brazilian Ae. aegypti for a currently circulating strain of YF virus, and investigated the potential for Brazilian Ae. albopictus to transmit YF.

  15. Genetic specificity and potential for local adaptation between dengue viruses and mosquito vectors

    Directory of Open Access Journals (Sweden)

    Richardson Jason H

    2009-07-01

    Full Text Available Abstract Background Several observations support the hypothesis that vector-driven selection plays an important role in shaping dengue virus (DENV genetic diversity. Clustering of DENV genetic diversity at a particular location may reflect underlying genetic structure of vector populations, which combined with specific vector genotype × virus genotype (G × G interactions may promote adaptation of viral lineages to local mosquito vector genotypes. Although spatial structure of vector polymorphism at neutral genetic loci is well-documented, existence of G × G interactions between mosquito and virus genotypes has not been formally demonstrated in natural populations. Here we measure G × G interactions in a system representative of a natural situation in Thailand by challenging three isofemale families from field-derived Aedes aegypti with three contemporaneous low-passage isolates of DENV-1. Results Among indices of vector competence examined, the proportion of mosquitoes with a midgut infection, viral RNA concentration in the body, and quantity of virus disseminated to the head/legs (but not the proportion of infected mosquitoes with a disseminated infection strongly depended on the specific combinations of isofemale families and viral isolates, demonstrating significant G × G interactions. Conclusion Evidence for genetic specificity of interactions in our simple experimental design indicates that vector competence of Ae. aegypti for DENV is likely governed to a large extent by G × G interactions in genetically diverse, natural populations. This result challenges the general relevance of conclusions from laboratory systems that consist of a single combination of mosquito and DENV genotypes. Combined with earlier evidence for fine-scale genetic structure of natural Ae. aegypti populations, our finding indicates that the necessary conditions for local DENV adaptation to mosquito vectors are met.

  16. Evaluating the feeding preferences of West Nile virus mosquito vectors using bird-baited traps

    OpenAIRE

    Victoriano Llopis, Isis; Tomassone, Laura; Grego, Elena; Serrano, Emmanuel; Mosca, Andrea; Vaschetti, Gabriella; Andrade, Daniela; Rossi, Luca

    2016-01-01

    Background The total contact rates (TCRs) between mosquito vectors and their potential hosts have a serious impact on disease transmission dynamics. Culex pipiens (sensu stricto) (s.s.) is considered the main vector of the West Nile Virus (WNV) in Europe and birds are the reservoir hosts. The results of our previous study showed that WNV seroreactors are significantly more prevalent among raptors compared to a range of other wild avian groups. The current study aims to assess the role of bird...

  17. Vector-borne transmission imposes a severe bottleneck on an RNA virus population.

    Science.gov (United States)

    Forrester, Naomi L; Guerbois, Mathilde; Seymour, Robert L; Spratt, Heidi; Weaver, Scott C

    2012-09-01

    RNA viruses typically occur in genetically diverse populations due to their error-prone genome replication. Genetic diversity is thought to be important in allowing RNA viruses to explore sequence space, facilitating adaptation to changing environments and hosts. Some arboviruses that infect both a mosquito vector and a mammalian host are known to experience population bottlenecks in their vectors, which may constrain their genetic diversity and could potentially lead to extinction events via Muller's ratchet. To examine this potential challenge of bottlenecks for arbovirus perpetuation, we studied Venezuelan equine encephalitis virus (VEEV) enzootic subtype IE and its natural vector Culex (Melanoconion) taeniopus, as an example of a virus-vector interaction with a long evolutionary history. Using a mixture of marked VEEV clones to infect C. taeniopus and real-time RT-PCR to track these clones during mosquito infection and dissemination, we observed severe bottleneck events that resulted in a significant drop in the number of clones present. At higher initial doses, the midgut was readily infected and there was a severe bottleneck at the midgut escape. Following a lower initial dose, the major bottleneck occurred at initial midgut infection. A second, less severe bottleneck was identified at the salivary gland infection stage following intrathoracic inoculation. Our results suggest that VEEV consistently encounters bottlenecks during infection, dissemination and transmission by its natural enzootic vector. The potential impacts of these bottlenecks on viral fitness and transmission, and the viral mechanisms that prevent genetic drift leading to extinction, deserve further study.

  18. Immunogenic Subviral Particles Displaying Domain III of Dengue 2 Envelope Protein Vectored by Measles Virus

    Directory of Open Access Journals (Sweden)

    Indira S. Harahap-Carrillo

    2015-07-01

    Full Text Available Vaccines against dengue virus (DV are commercially nonexistent. A subunit vaccination strategy may be of value, especially if a safe viral vector acts as biologically active adjuvant. In this paper, we focus on an immunoglobulin-like, independently folded domain III (DIII from DV 2 envelope protein (E, which contains epitopes that elicits highly specific neutralizing antibodies. We modified the hepatitis B small surface antigen (HBsAg, S in order to display DV 2 DIII on a virus-like particle (VLP, thus generating the hybrid antigen DIII-S. Two varieties of measles virus (MV vectors were developed to express DIII-S. The first expresses the hybrid antigen from an additional transcription unit (ATU and the second additionally expresses HBsAg from a separate ATU. We found that this second MV vectoring the hybrid VLPs displaying DIII-S on an unmodified HBsAg scaffold were immunogenic in MV-susceptible mice (HuCD46Ge-IFNarko, eliciting robust neutralizing responses (averages against MV (1:1280 NT90, hepatitis B virus (787 mIU/mL, and DV2 (1:160 NT50 in all of the tested animals. Conversely, the MV vector expressing only DIII-S induced immunity against MV alone. In summary, DV2 neutralizing responses can be generated by displaying E DIII on a scaffold of HBsAg-based VLPs, vectored by MV.

  19. High-efficiency protein expression in plants from agroinfection-compatible Tobacco mosaic virus expression vectors

    Directory of Open Access Journals (Sweden)

    Lindbo John A

    2007-08-01

    Full Text Available Abstract Background Plants are increasingly being examined as alternative recombinant protein expression systems. Recombinant protein expression levels in plants from Tobacco mosaic virus (TMV-based vectors are much higher than those possible from plant promoters. However the common TMV expression vectors are costly, and at times technically challenging, to work with. Therefore it was a goal to develop TMV expression vectors that express high levels of recombinant protein and are easier, more reliable, and more cost-effective to use. Results We have constructed a Cauliflower mosaic virus (CaMV 35S promoter-driven TMV expression vector that can be delivered as a T-DNA to plant cells by Agrobacterium tumefaciens. Co-introduction (by agroinfiltration of this T-DNA along with a 35S promoter driven gene for the RNA silencing suppressor P19, from Tomato bushy stunt virus (TBSV resulted in essentially complete infection of the infiltrated plant tissue with the TMV vector by 4 days post infiltration (DPI. The TMV vector produced between 600 and 1200 micrograms of recombinant protein per gram of infiltrated tissue by 6 DPI. Similar levels of recombinant protein were detected in systemically infected plant tissue 10–14 DPI. These expression levels were 10 to 25 times higher than the most efficient 35S promoter driven transient expression systems described to date. Conclusion These modifications to the TMV-based expression vector system have made TMV vectors an easier, more reliable and more cost-effective way to produce recombinant proteins in plants. These improvements should facilitate the production of recombinant proteins in plants for both research and product development purposes. The vector should be especially useful in high-throughput experiments.

  20. Aedes (Stegomyia) albopictus (Skuse): a potential vector of Zika virus in Singapore.

    Science.gov (United States)

    Wong, Pei-Sze Jeslyn; Li, Mei-zhi Irene; Chong, Chee-Seng; Ng, Lee-Ching; Tan, Cheong-Huat

    2013-01-01

    Zika virus (ZIKV) is a little known arbovirus until it caused a major outbreak in the Pacific Island of Yap in 2007. Although the virus has a wide geographic distribution, most of the known vectors are sylvatic Aedes mosquitoes from Africa where the virus was first isolated. Presently, Ae. aegypti is the only known vector to transmit the virus outside the African continent, though Ae. albopictus has long been a suspected vector. Currently, Ae. albopictus has been shown capable of transmitting more than 20 arboviruses and its notoriety as an important vector came to light during the recent chikungunya pandemic. The vulnerability of Singapore to emerging infectious arboviruses has stimulated our interest to determine the competence of local Ae. albopictus to transmit ZIKV. To determine the competence of Ae. albopictus to ZIKV, we orally infected local mosquito strains to a Ugandan strain virus. Fully engorged mosquitoes were maintained in an environmental chamber set at 29°C and 80-85%RH. Twelve mosquitoes were then sampled daily from day one to seven and on day 10 and 14 post infection (pi). Zika virus titre in the midgut and salivary glands of each mosquito were determined using tissue culture infectious dose50 assay, while transmissibility of the virus was determined by detecting viral antigen in the mosquito saliva by qRT-PCR. High dissemination and transmission rate of ZIKV were observed. By day 7-pi, all mosquitoes have disseminated infection and 73% of these mosquitoes have ZIKV in their saliva. By day 10-pi, all mosquitoes were potentially infectious. The study highlighted the potential of Ae. albopictus to transmit ZIKV and the possibility that the virus could be established locally. Nonetheless, the threat of ZIKV can be mitigated by existing dengue and chikungunya control program being implemented in Singapore.

  1. Aedes (Stegomyia albopictus (Skuse: a potential vector of Zika virus in Singapore.

    Directory of Open Access Journals (Sweden)

    Pei-Sze Jeslyn Wong

    Full Text Available Zika virus (ZIKV is a little known arbovirus until it caused a major outbreak in the Pacific Island of Yap in 2007. Although the virus has a wide geographic distribution, most of the known vectors are sylvatic Aedes mosquitoes from Africa where the virus was first isolated. Presently, Ae. aegypti is the only known vector to transmit the virus outside the African continent, though Ae. albopictus has long been a suspected vector. Currently, Ae. albopictus has been shown capable of transmitting more than 20 arboviruses and its notoriety as an important vector came to light during the recent chikungunya pandemic. The vulnerability of Singapore to emerging infectious arboviruses has stimulated our interest to determine the competence of local Ae. albopictus to transmit ZIKV.To determine the competence of Ae. albopictus to ZIKV, we orally infected local mosquito strains to a Ugandan strain virus. Fully engorged mosquitoes were maintained in an environmental chamber set at 29°C and 80-85%RH. Twelve mosquitoes were then sampled daily from day one to seven and on day 10 and 14 post infection (pi. Zika virus titre in the midgut and salivary glands of each mosquito were determined using tissue culture infectious dose50 assay, while transmissibility of the virus was determined by detecting viral antigen in the mosquito saliva by qRT-PCR. High dissemination and transmission rate of ZIKV were observed. By day 7-pi, all mosquitoes have disseminated infection and 73% of these mosquitoes have ZIKV in their saliva. By day 10-pi, all mosquitoes were potentially infectious.The study highlighted the potential of Ae. albopictus to transmit ZIKV and the possibility that the virus could be established locally. Nonetheless, the threat of ZIKV can be mitigated by existing dengue and chikungunya control program being implemented in Singapore.

  2. Impact of Ultraviolet-Blocking Plastic Films on Insect Vectors of Virus Diseases Infesting Crisp Lettuce

    OpenAIRE

    Díaz Desani, Beatriz M.; Biurrun, R.; Moreno, Aránzazu; Nebreda, Miguel; Fereres, Alberto

    2006-01-01

    Ultraviolet (UV)-absorbing plastic films are being used as a photoselective barrier to control insect vectors and associated virus diseases in different horticultural crops. A 2-year experiment was carried out in northeastern Spain (Navarra) to evaluate the impact of a UV-blocking film (AD-IR AV) on the population density of insect pests and the spread of insect-transmitted virus diseases associated with head lettuce [Lactuca sativa (L.)]. Results showed that the UV-absorbing plastic film did...

  3. Ability of herpes simplex virus vectors to boost immune responses to DNA vectors and to protect against challenge by simian immunodeficiency virus

    International Nuclear Information System (INIS)

    Kaur, Amitinder; Sanford, Hannah B.; Garry, Deirdre; Lang, Sabine; Klumpp, Sherry A.; Watanabe, Daisuke; Bronson, Roderick T.; Lifson, Jeffrey D.; Rosati, Margherita; Pavlakis, George N.; Felber, Barbara K.; Knipe, David M.; Desrosiers, Ronald C.

    2007-01-01

    The immunogenicity and protective capacity of replication-defective herpes simplex virus (HSV) vector-based vaccines were examined in rhesus macaques. Three macaques were inoculated with recombinant HSV vectors expressing Gag, Env, and a Tat-Rev-Nef fusion protein of simian immunodeficiency virus (SIV). Three other macaques were primed with recombinant DNA vectors expressing Gag, Env, and a Pol-Tat-Nef-Vif fusion protein prior to boosting with the HSV vectors. Robust anti-Gag and anti-Env cellular responses were detected in all six macaques. Following intravenous challenge with wild-type, cloned SIV239, peak and 12-week plasma viremia levels were significantly lower in vaccinated compared to control macaques. Plasma SIV RNA in vaccinated macaques was inversely correlated with anti-Rev ELISPOT responses on the day of challenge (P value < 0.05), anti-Tat ELISPOT responses at 2 weeks post challenge (P value < 0.05) and peak neutralizing antibody titers pre-challenge (P value 0.06). These findings support continued study of recombinant herpesviruses as a vaccine approach for AIDS

  4. [Rapid selection of recombinant orf virus expression vectors using green fluorescent protein].

    Science.gov (United States)

    Zhang, Jiachun; Guo, Xianfeng; Zhang, Min; Wu, Feifan; Peng, Yongzheng

    2016-01-01

    To construct a universal, highly attenuated orf virus expression vector for exogenous genes using green fluorescent protein (GFP) as the reporter gene. The flanking regions of the ORFV132 of orf virus DNA were amplified by PCR to construct the shuttle plasmid pSPV-132LF-EGFP-132RF. The shuttle plasmid was transfected into OFTu cells and GFP was incorporated into orf virus IA82Delta 121 by homologous recombination. The recombinant IA82Delta121-V was selected by green fluorescent signal. The deletion gene was identified by PCR and sequencing. The effects of ORFV132 knockout were evaluated by virus titration and by observing the proliferation of the infected vascular endothelial cells in vitro. The recombinant orf virus IA82Delta121-V was obtained successfully and quickly, and the deletion of ORFV132 did not affect the replication of the virus in vitro but reduced its virulence. Green fluorescent protein is a selectable marker for rapid, convenient and stable selection of the recombinant viruses. Highly attenuated recombinant orf virus IA82Delta121-V can serve as a new expression vector for exogenous genes.

  5. Virus infection mediates the effects of elevated CO2 on plants and vectors

    Science.gov (United States)

    Trębicki, Piotr; Vandegeer, Rebecca K.; Bosque-Pérez, Nilsa A.; Powell, Kevin S.; Dader, Beatriz; Freeman, Angela J.; Yen, Alan L.; Fitzgerald, Glenn J.; Luck, Jo E.

    2016-03-01

    Atmospheric carbon dioxide (CO2) concentration has increased significantly and is projected to double by 2100. To increase current food production levels, understanding how pests and diseases respond to future climate driven by increasing CO2 is imperative. We investigated the effects of elevated CO2 (eCO2) on the interactions among wheat (cv. Yitpi), Barley yellow dwarf virus and an important pest and virus vector, the bird cherry-oat aphid (Rhopalosiphum padi), by examining aphid life history, feeding behavior and plant physiology and biochemistry. Our results showed for the first time that virus infection can mediate effects of eCO2 on plants and pathogen vectors. Changes in plant N concentration influenced aphid life history and behavior, and N concentration was affected by virus infection under eCO2. We observed a reduction in aphid population size and increased feeding damage on noninfected plants under eCO2 but no changes to population and feeding on virus-infected plants irrespective of CO2 treatment. We expect potentially lower future aphid populations on noninfected plants but no change or increased aphid populations on virus-infected plants therefore subsequent virus spread. Our findings underscore the complexity of interactions between plants, insects and viruses under future climate with implications for plant disease epidemiology and crop production.

  6. CD4(+) T cell-mediated protection against a lethal outcome of systemic infection with vesicular stomatitis virus requires CD40 ligand expression, but not IFN-gamma or IL-4

    DEFF Research Database (Denmark)

    Andersen, C; Jensen, T; Nansen, A

    1999-01-01

    . Taken together, these results underscore that B cells are essential in preventing early infection of the CNS, but T cells are required for long-term survival. CD4(+) T cells are most efficient in this context and the key function is to provide cognate help to B cells. However, if CD4(+) cell function......To investigate the mechanism(s) whereby T cells protect against a lethal outcome of systemic infection with vesicular stomatitis virus, mice with targeted defects in genes central to T cell function were tested for resistance to i.v. infection with this virus. Our results show that mice lacking...

  7. Pseudacteon decapitating flies: Potential vectors of a fire ant virus?

    International Nuclear Information System (INIS)

    Valles, S.M.; Porter, S.D.

    2007-01-01

    Solenopsis invicta virus (SINV-1) is a positive-stranded RNA virus recently found to infect all stages of the red imported fire ant, Solenopsis invicta (Valles et al. 2004; Valles and Strong 2005). SINV-1 and a second genotype have been tentatively assigned to the Dicistroviridae (Mayo 2002). Infected individuals or colonies did not exhibit any immediate, discernible symptoms in the field. However, under stress from introduction into the laboratory, brood death was often observed among infected colonies, ultimately leading to the death of the entire colony (Valles et al. 2004). These characteristics are consistent with other insect-infecting positive-stranded RNA viruses. They often persist as inapparent, asymptomatic infections that, under certain conditions, induce replication within the host, resulting in observable symptoms and often death (Christian and Scotti 1998; Fernandez et al. 2002). The SINV infection rate among colonies was reported to be around 25% in Gainesville, Florida (Valles et al. 2004; Valles and Strong 2005). SINV vertical and horizontal transmission were inferred based on RT-PCR detection of virus genome in eggs and successful colony to colony transfer under lab conditions (Valles et al. 2004). However, the exact mechanisms by which the virus is spread from nest to nest in the field are unknown. Our results indicate that SINV does not replicate within Pseudacteon decapitating flies that parasitize S. invicta. Flies appeared to develop normally from SINV-infected S. invicta workers. Mechanical transmission of SINV to uninfected ants by oviposition appears unlikely

  8. Construction of PVX virus-expression vector to express enterotoxin ...

    African Journals Online (AJOL)

    Potato X potyvirus (PVX)-based vector has been comprehensively applied in transient expression system. In order to produce the heterologous proteins more quickly and stably, the ClaI and NotI enzyme sites were introduced into the Enterotoxin fusion gene LTB-ST by polymerase chain reaction (PCR) and the LTB-ST ...

  9. potential for biological control of rice yellow mottle virus vectors

    African Journals Online (AJOL)

    Administrator

    irrigated fields, samplings occurred between December and April. Rearing using dead insects was conducted simultaneously in the laboratory to identify the parasitoid insect species. From samples obtained at different sites: (i) the dominant structure of the RYMV insect vectors was analysed according to the rice phenology; ...

  10. Recombinant influenza viruses as delivery vectors for hepatis B virus epitopes.

    Science.gov (United States)

    Song, Jae-Min; Lee, Kwang-Hee; Seong, Baik-Lin

    2012-07-01

    Neuraminidase (NA) of influenza virus contains stalk region that shows a great deal of variability in both amino acid sequence and length. In this paper, we investigated generation of recombinant influenza viruses that had hepatitis B virus (HBV) B cell epitopes in the NA stalk region as a dual vaccine candidate. We used the WSH-HK reassortant helper virus for rescue of recombinant influenza virus containing HBV epitopes and reverse genetic protocol based on the use of micrococcal nuclease-treated virus cores for reconstitution of ribonucleoproteins. We successfully generated a chimeric influenza viruses which contained 22 amino acid peptides in the stalk region derived from the surface and pre-surface protein HBV. The growth kinetics of the recombinant viruses was investigated after infection of Madin-Darby canine kidney (MDCK) and Madin-Darby bovine kidney (MDBK) cells and the rIV-BVPreS virus showed higher titer than other viruses in MDCK cells. We also confirmed the presence of HBV epitopes in the chimeric viruses by enzyme-linked immunosorbent assay (ELISA) using anti-HBV polyclonal antibody. When the ratio of recombinant virus verse wild type virus was calculated by ELISA, recombinant viruses exhibited 2 fold higher values than the wild type virus. These results suggest that chimeric influenza virus which contained foreign antigens can be used as dual vaccine against both HBV and influenza viruses.

  11. Vector and Serologic Survey for Crimean-Congo Hemorrhagic Fever Virus in Poland.

    Science.gov (United States)

    Bażanów, Barbara A; Pacoń, Jarosław; Gadzała, Łukasz; Frącka, Agnieszka; Welz, Mirosław; Paweska, Janusz

    2017-07-01

    In contrast to animals, Crimean-Congo hemorrhagic fever (CCHF) causes a severe disease in humans with a high mortality rate. The etiological agent, CCHF virus (CCHFV), can be transmitted by argasid and ixodid ticks, but arachnids of the genus Hyalomma, followed by Rhipicephalus and Dermacentor serve as the major vectors of this virus. The goal of the study was to assess the epidemiological situation of CCHFV infection in cattle in south-east Poland, and survey for potential tick vector species. A total of 592 bovine blood samples from animals located in the southernmost region in Poland were tested by IgG sandwich enzyme-linked immunosorbent assay. Ticks (n = 993) from south-east Poland were collected from dogs, cats, cattle, and horses and tested by RT-PCR. All 592 serum samples were negative for IgG antibodies to CCHFV. Of the ticks collected, 125 were Dermacentor reticulatus and 868 represented Ixodes ricinus, both species are regarded as potential vectors of CCHFV. All tick samples were negative for the presence of CCHFV. Considering the zoonotic nature, public health importance, and the virus increasing spread, it was prudent to assess the seroprevalence of CCHFV in the south-east area of Poland, bordering with CCHFV endemic areas. It seems unlikely that CCHFV infection will suddenly spread in Poland, but considering the multiple possibilities of the virus introduction, serosurveys and vector biosurveillance should be conducted at regular intervals.

  12. Fowl adenovirus serotype 9 vectored vaccine for protection of avian influenza virus

    Science.gov (United States)

    A fowl adenovirus serotype 9, a non-pathogenic large double stranded DNA virus, was developed as a viral vector to express influenza genes as a potential vaccine. Two separate constructs were developed that expressed either the hemagglutinin gene of A/Chicken/Jalisco/2012 (H7) or A/ Chicken/Iowa/20...

  13. Comparison of transmission efficiency of different isolates of Potato virus Y among three aphid vectors

    Science.gov (United States)

    Potato virus Y (PVY) strains are transmitted by different aphid species in a non-persistent, non-circulative manner. Green peach aphid (GPA, Myzus persicae Sulzer; Aphididae, Macrosiphini) is the most efficient vector in laboratory studies, but potato aphid (PA, Macrosiphum euphorbiae Thomas; Aphidi...

  14. Transient expression of heterologous model gene in plants using Potato virusX-based vector

    Czech Academy of Sciences Publication Activity Database

    Čeřovská, Noemi; Pečenková, Tamara; Moravec, Tomáš; Velemínský, Jiří

    2004-01-01

    Roč. 79, č. 2 (2004), s. 147-152 ISSN 0167-6857 R&D Projects: GA ČR GA310/00/0381 Institutional research plan: CEZ:AV0Z5038910 Keywords : plant virus * based vector * transient expression Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.028, year: 2004

  15. Recombinant Fluorescent Rabies Virus Vectors for Tracing Neurons and Synaptic Connections.

    Science.gov (United States)

    Hagendorf, Nadin; Conzelmann, Karl-Klaus

    2015-12-02

    Recombinant rabies virus (RV) vectors expressing fluorescent proteins allow staining of neurons from many mammalian species and enable the study of neuron morphology. Because viral spread occurs only between neurons that have synaptic connections, these vectors also permit transsynaptic tracing. A recently established system for restriction of transsynaptic tracing to a single transsynaptic jump, dubbed monosynaptic tracing, uses glycoprotein gene-defective, pseudotyped RV. This allows infection of defined cells and transient complementation with the glycoprotein in situ to support a single step of transsynaptic crossing to presynaptic cells. Here, we introduce protocols describing the production of RV vectors, including the recovery of recombinant RV from complementary DNA (cDNA) and virus pseudotyping in vitro. This allows retrograde staining of neurons projecting to the inoculation site. © 2015 Cold Spring Harbor Laboratory Press.

  16. Vector-virus mutualism accelerates population increase of an invasive whitefly.

    Science.gov (United States)

    Jiu, Min; Zhou, Xue-Ping; Tong, Lin; Xu, Jing; Yang, Xiao; Wan, Fang-Hao; Liu, Shu-Sheng

    2007-01-31

    The relationships between plant viruses, their herbivore vectors and host plants can be beneficial, neutral, or antagonistic, depending on the species involved. This variation in relationships may affect the process of biological invasion and the displacement of indigenous species by invaders when the invasive and indigenous organisms occur with niche overlap but differ in the interactions. The notorious invasive B biotype of the whitefly complex Bemisia tabaci entered China in the late 1990s and is now the predominant or only biotype in many regions of the country. Tobacco curly shoot virus (TbCSV) and Tomato yellow leaf curl China virus (TYLCCNV) are two whitefly-transmitted begomoviruses that have become widespread recently in south China. We compared the performance of the invasive B and indigenous ZHJ1 whitefly biotypes on healthy, TbCSV-infected and TYLCCNV-infected tobacco plants. Compared to its performance on healthy plants, the invasive B biotype increased its fecundity and longevity by 12 and 6 fold when feeding on TbCSV-infected plants, and by 18 and 7 fold when feeding on TYLCCNV-infected plants. Population density of the B biotype on TbCSV- and TYLCCNV-infected plants reached 2 and 13 times that on healthy plants respectively in 56 days. In contrast, the indigenous ZHJ1 performed similarly on healthy and virus-infected plants. Virus-infection status of the whiteflies per se of both biotypes showed limited effects on performance of vectors on cotton, a nonhost plant of the viruses. The indirect mutualism between the B biotype whitefly and these viruses via their host plants, and the apparent lack of such mutualism for the indigenous whitefly, may contribute to the ability of the B whitefly biotype to invade, the displacement of indigenous whiteflies, and the disease pandemics of the viruses associated with this vector.

  17. Vector-virus mutualism accelerates population increase of an invasive whitefly.

    Directory of Open Access Journals (Sweden)

    Min Jiu

    2007-01-01

    Full Text Available The relationships between plant viruses, their herbivore vectors and host plants can be beneficial, neutral, or antagonistic, depending on the species involved. This variation in relationships may affect the process of biological invasion and the displacement of indigenous species by invaders when the invasive and indigenous organisms occur with niche overlap but differ in the interactions. The notorious invasive B biotype of the whitefly complex Bemisia tabaci entered China in the late 1990s and is now the predominant or only biotype in many regions of the country. Tobacco curly shoot virus (TbCSV and Tomato yellow leaf curl China virus (TYLCCNV are two whitefly-transmitted begomoviruses that have become widespread recently in south China. We compared the performance of the invasive B and indigenous ZHJ1 whitefly biotypes on healthy, TbCSV-infected and TYLCCNV-infected tobacco plants. Compared to its performance on healthy plants, the invasive B biotype increased its fecundity and longevity by 12 and 6 fold when feeding on TbCSV-infected plants, and by 18 and 7 fold when feeding on TYLCCNV-infected plants. Population density of the B biotype on TbCSV- and TYLCCNV-infected plants reached 2 and 13 times that on healthy plants respectively in 56 days. In contrast, the indigenous ZHJ1 performed similarly on healthy and virus-infected plants. Virus-infection status of the whiteflies per se of both biotypes showed limited effects on performance of vectors on cotton, a nonhost plant of the viruses. The indirect mutualism between the B biotype whitefly and these viruses via their host plants, and the apparent lack of such mutualism for the indigenous whitefly, may contribute to the ability of the B whitefly biotype to invade, the displacement of indigenous whiteflies, and the disease pandemics of the viruses associated with this vector.

  18. Expression of the Surface Glycoproteins of Human Parainfluenza Virus Type 3 by Bovine Parainfluenza Virus Type 3, a Novel Attenuated Virus Vaccine Vector

    OpenAIRE

    Haller, Aurelia A.; Miller, Tessa; Mitiku, Misrach; Coelingh, Kathleen

    2000-01-01

    Bovine parainfluenza virus type 3 (bPIV3) is being evaluated as an intranasal vaccine for protection against human PIV3 (hPIV3). In young infants, the bPIV3 vaccine appears to be infectious, attenuated, immunogenic, and genetically stable, which are desirable characteristics for an RNA virus vector. To test the potential of the bPIV3 vaccine strain as a vector, an infectious DNA clone of bPIV3 was assembled and recombinant bPIV3 (r-bPIV3) was rescued. r-bPIV3 displayed a temperature-sensitive...

  19. Vectors

    DEFF Research Database (Denmark)

    Boeriis, Morten; van Leeuwen, Theo

    2017-01-01

    This article revisits the concept of vectors, which, in Kress and van Leeuwen’s Reading Images (2006), plays a crucial role in distinguishing between ‘narrative’, action-oriented processes and ‘conceptual’, state-oriented processes. The use of this concept in image analysis has usually focused...... on the most salient vectors, and this works well, but many images contain a plethora of vectors, which makes their structure quite different from the linguistic transitivity structures with which Kress and van Leeuwen have compared ‘narrative’ images. It can also be asked whether facial expression vectors...... should be taken into account in discussing ‘reactions’, which Kress and van Leeuwen link only to eyeline vectors. Finally, the question can be raised as to whether actions are always realized by vectors. Drawing on a re-reading of Rudolf Arnheim’s account of vectors, these issues are outlined...

  20. Meta-Analyses of Japanese Encephalitis Virus Infection, Dissemination, and Transmission Rates in Vectors.

    Science.gov (United States)

    Oliveira, Ana R S; Cohnstaedt, Lee W; Strathe, Erin; Etcheverry, Luciana; McVey, D Scott; Piaggio, José; Cernicchiaro, Natalia

    2018-03-01

    The objective of this work was to summarize and quantify Japanese encephalitis virus (JEV) infection, dissemination, and transmission rates in mosquitoes, using a meta-analysis approach. Data were obtained from experimental studies, gathered by means of a systematic review of the literature. Random-effects subgroup meta-analysis models by mosquito species were fitted to estimate pooled estimates and to calculate the variance between studies for three outcomes of interest: JEV infection, dissemination, and transmission rates in mosquitoes. To identify sources of heterogeneity among studies and to assess the association between different predictors (mosquito species, virus administration route, incubation period, and diagnostic method) with the outcome JEV infection rate in vectors, we fitted univariable meta-regression models. Mosquito species and administration route represented the main sources of heterogeneity associated with JEV infection rate in vectors. This study provided summary effect size estimates to be used as reference for other investigators when assessing transmission efficiency of vectors and explored sources of variability for JEV infection rates in vectors. Because transmission efficiency, as part of vector competence assessment, is an important parameter when studying the relative contribution of vectors to JEV transmission, our findings contribute to further our knowledge, potentially moving us toward more informed and targeted actions to prevent and control JEV in both affected and susceptible regions worldwide.

  1. Safety of inoculation of bovine parainfluenza virus 3 as potential vaccine vector in pigs.

    Science.gov (United States)

    Wang, Feng-Xue; Liu, Ying; Zhu, Hong-Wei; Liu, Xing; Yang, Yong; Sun, Na; Cheng, Shi-Peng; Wen, Yong-Jun

    2015-06-01

    Bovine parainfluenza virus 3 (BPIV3) is one of the most important respiratory pathogens in cattle. One BPIV3, named NM09, was isolated from cattle suffering from severe respiratory diseases in 2009. BPIV3 is a potential recombinant vaccine vector. To investigate whether NM09 can infect pigs and determine BPIV3 defense in these animals, BPIV3 antibody-free pigs were inoculated intramuscularly with the BPIV3 NM09 strain in a continuous passage. Clinical signs were observed each day after inoculation. Viral nucleic acid was detected in nasal and anal secretions. Results showed that virus-inoculated pigs displayed few observable clinical signs related to respiratory diseases. The antibody was identified, but the virus could not be detected in the second continuous passage in pigs. Thus, BPIV3 is a potential vaccine vector for genetic engineering.

  2. Tomato yellow leaf curl virus: No evidence for replication in the insect vector Bemisia tabaci.

    Science.gov (United States)

    Sánchez-Campos, Sonia; Rodríguez-Negrete, Edgar A; Cruzado, Lucía; Grande-Pérez, Ana; Bejarano, Eduardo R; Navas-Castillo, Jesús; Moriones, Enrique

    2016-08-01

    Begomovirus ssDNA plant virus (family Geminiviridae) replication within the Bemisia tabaci vector is controversial. Transovarial transmission, alteration to whitefly biology, or detection of viral transcripts in the vector are proposed as indirect evidence of replication of tomato yellow leaf curl virus (TYLCV). Recently, contrasting direct evidence has been reported regarding the capacity of TYLCV to replicate within individuals of B. tabaci based on quantitave PCR approaches. Time-course experiments to quantify complementary and virion sense viral nucleic acid accumulation within B. tabaci using a recently implemented two step qPCR procedure revealed that viral DNA quantities did not increase for time points up to 96 hours after acquisition of the virus. Our findings do not support a recent report claiming TYLCV replication in individuals of B. tabaci.

  3. Evidence for Culicoides obsoletus group as vector for Schmallenberg virus in Denmark

    DEFF Research Database (Denmark)

    Rasmussen, Lasse Dam; Kristensen, Birgit; Kirkeby, Carsten

    the Bunyaviridae family and is closely related to Shamonda and Akabane viruses. These viruses are transmitted by insect vectors (including biting midges (Culicoides sp.) and mosquitoes). To determine whether these insects may act as vectors for SBV, biting midges (Culicoides spp.) caught in October 2011......, in the south-west of Denmark (close to the German border), were sorted into pools and tested for the presence of Schmallenberg virus RNA by RT-qPCR. From 18 pools of 5 midges from the C. obsoletus group, 2 pools were both found positive in two separate assays, targeting the L- and S- segments of the SBV RNA....... However, 4 pools of C. punctatus s.str were negative. The sequence of 80bp (excluding the primer sequences) from the amplicons (ca. 145bp) was identical to that published for the expected region of the SBV L-segment. The levels of SBV RNA detected in the biting midges were much higher than could...

  4. Glycoprotein is enough for sindbis virus-derived DNA vector to express heterogenous genes

    Directory of Open Access Journals (Sweden)

    Fu Juanjuan

    2011-07-01

    Full Text Available Abstract To investigate the necessity and potential application of structural genes for expressing heterogenous genes from Sindbis virus-derived vector, the DNA-based expression vector pVaXJ was constructed by placing the recombinant genome of sindbis-like virus XJ-160 under the control of the human cytomegalovirus (CMV promoter of the plasmid pVAX1, in which viral structural genes were replaced by a polylinker cassette to allow for insertion of heterologous genes. The defect helper plasmids pVaE or pVaC were developed by cloning the gene of glycoprotein E3E26KE1 or capsid protein of XJ-160 virus into pVAX1, respectively. The report gene cassette pVaXJ-EGFP or pV-Gluc expressing enhanced green fluorescence protein (EGFP or Gaussia luciferase (G.luc were constructed by cloning EGFP or G.luc gene into pVaXJ. EGFP or G.luc was expressed in the BHK-21 cells co-transfected with report gene cassettes and pVaE at levels that were comparable to those produced by report gene cassettes, pVaC and pVaE and were much higher than the levels produced by report gene cassette and pVaC, suggesting that glycoprotein is enough for Sindbis virus-derived DNA vector to express heterogenous genes in host cells. The method of gene expression from Sindbis virus-based DNA vector only co-transfected with envelop E gene increase the conveniency and the utility of alphavirus-based vector systems in general.

  5. Cross linkage studies with the membranes of the vesicular stomatitis virus using radioactive 4-acido and 5-acido palmitic acid

    International Nuclear Information System (INIS)

    Verfondern, M.

    1983-01-01

    In the study described here the spatial arrangement of lipids and proteins in the VS virus was investigated on the basis of the covalent cross linkage technique. The formation of such cross linkages is brought about by the action of photosensitive acidosubstituted lipids, which permit acido functions to be introduced into a membrane in a previously defined position. Subsequently, photolysis helps to trigger the generation of radioactive nitrenes that react with the proteins and lipids in their immediate vicinity in a direct and non-selective way. The findings revealed by this study have raised questions as to the possibility of lipid-protein and lipid-lipid interactions, which is also discussed. (orig./MG) [de

  6. Vectors

    DEFF Research Database (Denmark)

    Boeriis, Morten; van Leeuwen, Theo

    2017-01-01

    This article revisits the concept of vectors, which, in Kress and van Leeuwen’s Reading Images (2006), plays a crucial role in distinguishing between ‘narrative’, action-oriented processes and ‘conceptual’, state-oriented processes. The use of this concept in image analysis has usually focused...... should be taken into account in discussing ‘reactions’, which Kress and van Leeuwen link only to eyeline vectors. Finally, the question can be raised as to whether actions are always realized by vectors. Drawing on a re-reading of Rudolf Arnheim’s account of vectors, these issues are outlined...

  7. Use of a Virus Gene Silencing Vector for Maize Functional Genomics Research.

    Science.gov (United States)

    Zhou, Tao; Liu, Xuedong; Fan, Zaifeng

    2018-01-01

    Virus-induced gene silencing (VIGS) is a genetic technology that exploits the RNA-mediated defense against virus. The method has great potential for plant reverse genetics as it could knock down gene expression in a rapid way, which is triggered by a replicating viral genome engineered to carry a fragment of host gene to be silenced. A number of efficient VIGS vectors are available for dicots, such as for model plant Nicotiana benthamiana; however, only a few of VIGS vectors for monocotyledonous cereal crops. Here, we describe the method for the use of a newly developed VIGS vector based on a maize-infecting Cucumber mosaic virus (CMV) strain ZMBJ-CMV for maize. The RNA2 of ZMBJ-CMV was modified as a vector pCMV201-2b N81 having multiple cloning sites for the insert of 100-300 bp fragment of target gene. Using a method of vascular puncture inoculation of maize seeds with crude sap prepared from Agrobacterium-infiltrated N. benthamiana leaves, silencing of target genes could be obtained in 4 weeks.

  8. Zika virus infection: Past and present of another emerging vector-borne disease.

    Science.gov (United States)

    Sakkas, Hercules; Economou, Vangelis; Papadopoulou, Chrissanthy

    2016-01-01

    Zika virus infection is an emerging mosquito-borne disease, first identified in Uganda in 1947. It is caused by the Zika arbovirus, and transmitted by the bites of infected mosquitoes of the genus Aedes. For almost half a century, the Zika virus was reported as the causative agent of sporadic human infections. In 2007, the Zika virus emerged outside Asia and Africa causing an epidemic on the Island of Yap in Micronesia. The manifestation of the newly acquired human infection varies from asymptomatic to self-limiting acute febrile illness with symptoms and clinical features similar to those caused by the Dengue virus ('Dengue-like syndrome'). The real-time PCR and serological methods have been successfully applied for the diagnosis of the disease. The treatment is symptomatic, since there is no specific antiviral treatment or a vaccine. During the recent outbreaks in French Polynesia and Brazil, incidents of Guillain-Barrι syndrome and microcephaly were associated with Zika virus infection, giving rise to fears of further global spread of the virus. Prevention and vector control strategies have to be urgently implemented by national health authorities in order to contain future outbreaks in vulnerable populations. This review summarizes the existing information on Zika virus characteristics, pathogenesis and epidemiology, the available methods for the diagnosis of Zika virus infection and recent approaches for prevention and control.

  9. Ecological niche modelling of Rift Valley fever virus vectors in Baringo, Kenya

    Directory of Open Access Journals (Sweden)

    Alfred O. Ochieng

    2016-11-01

    Full Text Available Background: Rift Valley fever (RVF is a vector-borne zoonotic disease that has an impact on human health and animal productivity. Here, we explore the use of vector presence modelling to predict the distribution of RVF vector species under climate change scenario to demonstrate the potential for geographic spread of Rift Valley fever virus (RVFV. Objectives: To evaluate the effect of climate change on RVF vector distribution in Baringo County, Kenya, with an aim of developing a risk map for spatial prediction of RVF outbreaks. Methodology: The study used data on vector presence and ecological niche modelling (MaxEnt algorithm to predict the effect of climatic change on habitat suitability and the spatial distribution of RVF vectors in Baringo County. Data on species occurrence were obtained from longitudinal sampling of adult mosquitoes and larvae in the study area. We used present (2000 and future (2050 Bioclim climate databases to model the vector distribution. Results: Model results predicted potential suitable areas with high success rates for Culex quinquefasciatus, Culex univitattus, Mansonia africana, and Mansonia uniformis. Under the present climatic conditions, the lowlands were found to be highly suitable for all the species. Future climatic conditions indicate an increase in the spatial distribution of Cx. quinquefasciatus and M. africana. Model performance was statistically significant. Conclusion: Soil types, precipitation in the driest quarter, precipitation seasonality, and isothermality showed the highest predictive potential for the four species.

  10. Protective efficacy of a virus-vectored multi-component vaccine against porcine reproductive and respiratory syndrome virus, porcine circovirus type 2 and swine influenza virus.

    Science.gov (United States)

    Tian, Debin; Sooryanarain, Harini; Matzinger, Shannon R; Gauger, Phil C; Karuppannan, Anbu K; Elankumaran, Subbiah; Opriessnig, Tanja; Meng, Xiang-Jin

    2017-12-01

    Porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2) and swine influenza virus (SIV) are three of the most economically important swine pathogens, causing immense economic losses to the global swine industry. Monovalent commercial vaccines against each of the three viruses are routinely used in pig farms worldwide. A trivalent vaccine against all three pathogens would greatly simplify the vaccination programme and reduce the financial burden to the swine industry. In this study, by using an attenuated strain of PRRSV (strain DS722) as a live virus vector, we generated a multi-component vaccine virus, DS722-SIV-PCV2, which expresses the protective antigens from SIV and PCV2. The DS722-SIV-PCV2 trivalent vaccine virus replicates well, and expresses PCV2 capsid and SIV HA proteins in vitro. A subsequent vaccination and challenge study in 48 pigs revealed that the DS722-SIV-PCV2-vaccinated pigs had significantly reduced lung lesions and viral RNA loads when challenged with PRRSV. Upon challenge with PCV2, the vaccinated pigs had partially reduced lymphoid lesions and viral DNA loads, and when challenged with SIV the vaccinated pigs had significantly reduced acute respiratory sign scores. The results from this study demonstrate the potential of DS722-SIV-PCV2 as a candidate trivalent vaccine, and also shed light on exploring PRRSV as a potential live virus vaccine vector.

  11. Vector Competence of American Mosquitoes for Three Strains of Zika Virus.

    Directory of Open Access Journals (Sweden)

    James Weger-Lucarelli

    2016-10-01

    Full Text Available In 2015, Zika virus (ZIKV; Flaviviridae; Flavivirus emerged in the Americas, causing millions of infections in dozens of countries. The rapid spread of the virus and the association with disease outcomes such as Guillain-Barré syndrome and microcephaly make understanding transmission dynamics essential. Currently, there are no reports of vector competence (VC of American mosquitoes for ZIKV isolates from the Americas. Further, it is not clear whether ZIKV strains from other genetic lineages can be transmitted by American Aedes aegypti populations, and whether the scope of the current epidemic is in part facilitated by viral factors such as enhanced replicative fitness or increased vector competence. Therefore, we characterized replication of three ZIKV strains, one from each of the three phylogenetic clades in several cell lines and assessed their abilities to be transmitted by Ae. aegypti mosquitoes. Additionally, laboratory colonies of different Culex spp. were infected with an American outbreak strain of ZIKV to assess VC. Replication rates were variable and depended on virus strain, cell line and MOI. African strains used in this study outcompeted the American strain in vitro in both mammalian and mosquito cell culture. West and East African strains of ZIKV tested here were more efficiently transmitted by Ae. aegypti from Mexico than was the currently circulating American strain of the Asian lineage. Long-established laboratory colonies of Culex mosquitoes were not efficient ZIKV vectors. These data demonstrate the capacity for additional ZIKV strains to infect and replicate in American Aedes mosquitoes and suggest that neither enhanced virus replicative fitness nor virus adaptation to local vector mosquitoes seems likely to explain the extent and intensity of ZIKV transmission in the Americas.

  12. Proteomic Analysis of Interaction between a Plant Virus and Its Vector Insect Reveals New Functions of Hemipteran Cuticular Protein.

    Science.gov (United States)

    Liu, Wenwen; Gray, Stewart; Huo, Yan; Li, Li; Wei, Taiyun; Wang, Xifeng

    2015-08-01

    Numerous viruses can be transmitted by their corresponding vector insects; however, the molecular mechanisms enabling virus transmission by vector insects have been poorly understood, especially the identity of vector components interacting with the virus. Here, we used the yeast two-hybrid system to study proteomic interactions of a plant virus (Rice stripe virus, RSV, genus Tenuivirus) with its vector insect, small brown planthopper (Laodelphax striatellus). Sixty-six proteins of L. striatellus that interacted with the nucleocapsid protein (pc3) of RSV were identified. A virus-insect interaction network, constructed for pc3 and 29 protein homologs of Drosophila melanogaster, suggested that nine proteins might directly interact with pc3. Of the 66 proteins, five (atlasin, a novel cuticular protein, jagunal, NAC domain protein, and vitellogenin) were most likely to be involved in viral movement, replication, and transovarial transmission. This work also provides evidence that the novel cuticular protein, CPR1, from L. striatellus is essential for RSV transmission by its vector insect. CPR1 binds the nucleocapsid protein (pc3) of RSV both in vivo and in vitro and colocalizes with RSV in the hemocytes of L. striatellus. Knockdown of CPR1 transcription using RNA interference resulted in a decrease in the concentration of RSV in the hemolymph, salivary glands and in viral transmission efficiency. These data suggest that CPR1 binds RSV in the insect and stabilizes the viral concentration in the hemolymph, perhaps to protect the virus or to help move the virus to the salivary tissues. Our studies provide direct experimental evidence that viruses can use existing vector proteins to aid their survival in the hemolymph. Identifying these putative vector molecules should lead to a better understanding of the interactions between viruses and vector insects. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Adeno-associated virus vector transduction of vascular smooth muscle cells in vivo.

    Science.gov (United States)

    Richter, M; Iwata, A; Nyhuis, J; Nitta, Y; Miller, A D; Halbert, C L; Allen, M D

    2000-04-27

    Adeno-associated virus (AAV) vectors might offer solutions for restenosis and angiogenesis by transducing nondividing cells and providing long-term gene expression. We investigated the feasibility of vascular cell transduction by AAV vectors in an in vivo rabbit carotid artery model. Time course of gene expression, inflammatory reaction to the vector, and effects of varying viral titer, exposure time, and intraluminal pressures on gene expression were examined. Recombinant AAV vectors with an Rous sarcoma virus promoter and alkaline phosphatase reporter gene were injected intraluminally into transiently isolated carotid segments. Following transduction, gene expression increased significantly over 14 days and then remained stable to 28 days, the last time point examined. Medial vascular smooth muscle cells were the main cell type transduced even with an intact endothelial layer. Increasing the viral titer and intraluminal pressure both enhanced transduction efficiency to achieve a mean of 34 +/- 7% of the subintimal layer of smooth muscle cells expressing gene product. A mild inflammatory reaction, composed of T cells with only rare macrophages, with minimal intimal thickening was demonstrated in 40% of transduced vessels; inflammatory cells were not detected in sham-operated control arteries. These findings demonstrate that AAV is a promising vector for intravascular applications in coronary and peripheral vascular diseases.

  14. A high throughput barley stripe mosaic virus vector for virus induced gene silencing in monocots and dicots.

    Directory of Open Access Journals (Sweden)

    Cheng Yuan

    Full Text Available Barley stripe mosaic virus (BSMV is a single-stranded RNA virus with three genome components designated alpha, beta, and gamma. BSMV vectors have previously been shown to be efficient virus induced gene silencing (VIGS vehicles in barley and wheat and have provided important information about host genes functioning during pathogenesis as well as various aspects of genes functioning in development. To permit more effective use of BSMV VIGS for functional genomics experiments, we have developed an Agrobacterium delivery system for BSMV and have coupled this with a ligation independent cloning (LIC strategy to mediate efficient cloning of host genes. Infiltrated Nicotiana benthamiana leaves provided excellent sources of virus for secondary BSMV infections and VIGS in cereals. The Agro/LIC BSMV VIGS vectors were able to function in high efficiency down regulation of phytoene desaturase (PDS, magnesium chelatase subunit H (ChlH, and plastid transketolase (TK gene silencing in N. benthamiana and in the monocots, wheat, barley, and the model grass, Brachypodium distachyon. Suppression of an Arabidopsis orthologue cloned from wheat (TaPMR5 also interfered with wheat powdery mildew (Blumeria graminis f. sp. tritici infections in a manner similar to that of the A. thaliana PMR5 loss-of-function allele. These results imply that the PMR5 gene has maintained similar functions across monocot and dicot families. Our BSMV VIGS system provides substantial advantages in expense, cloning efficiency, ease of manipulation and ability to apply VIGS for high throughput genomics studies.

  15. Resting lymphocyte transduction with measles virus glycoprotein pseudotyped lentiviral vectors relies on CD46 and SLAM

    International Nuclear Information System (INIS)

    Zhou Qi; Schneider, Irene C.; Gallet, Manuela; Kneissl, Sabrina; Buchholz, Christian J.

    2011-01-01

    The measles virus (MV) glycoproteins hemagglutinin (H) and fusion (F) were recently shown to mediate transduction of resting lymphocytes by lentiviral vectors. MV vaccine strains use CD46 or signaling lymphocyte activation molecule (SLAM) as receptor for cell entry. A panel of H protein mutants derived from vaccine strain or wild-type MVs that lost or gained CD46 or SLAM receptor usage were investigated for their ability to mediate gene transfer into unstimulated T lymphocytes. The results demonstrate that CD46 is sufficient for efficient vector particle association with unstimulated lymphocytes. For stable gene transfer into these cells, however, both MV receptors were found to be essential.

  16. Preparing the United States for Zika Virus: Pre-emptive Vector Control and Personal Protection.

    Science.gov (United States)

    Diaz, James H

    2016-12-01

    Discovered in 1947 in a monkey in the Zika forest of Uganda, Zika virus was dismissed as a cause of a mild illness that was confined to Africa and Southeast Asia and transmitted by Aedes mosquitoes. In 2007, Zika virus appeared outside of its endemic borders in an outbreak on the South Pacific Island of Yap. In 2013, Zika virus was associated with a major neurological complication, Guillain-Barré syndrome, in a larger outbreak in the French Polynesian Islands. From the South Pacific, Zika invaded Brazil in 2015 and caused another severe neurological complication, fetal microcephaly. The mosquito-borne transmission of Zika virus can be propagated by sexual transmission and, possibly, by blood transfusions, close personal contacts, and organ transplants, like other flaviviruses. Since these combined mechanisms of infectious disease transmission could result in catastrophic incidences of severe neurological diseases in adults and children, the public should know what to expect from Zika virus, how to prevent infection, and what the most likely failures in preventive measures will be. With federal research funding stalled, a Zika vaccine is far away. The only national strategies to prepare the United States for Zika virus invasion now are effective vector control measures and personal protection from mosquito bites. In addition to a basic knowledge of Aedes mosquito vectors and their biting behaviors, an understanding of simple household vector control measures, and the selection of the best chemical and physical mosquito repellents will be required to repel the Zika threat. Copyright © 2016 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.

  17. Wolbachia Occurrence in Planthopper (Hemiptera: Delphacidae) Vectors of Cereal Viruses in Argentina.

    Science.gov (United States)

    Mattio, M F; Argüello Caro, E B; Rodriguero, M S; Dumón, A D; Alemandri, V M; Truol, G

    2015-08-01

    Maize (Zea mays L.) and wheat (Triticum aestivum L.) are the most important cereal crops for the Argentinean economy and are affected by several diseases. Different planthopper species transmit causal agents of some of those diseases, including Mal de Río Cuarto virus, barley yellow striate mosaic virus, and the recently proposed maize yellow striate virus. Many planthopper species are sap feeders and therefore are expected to host bacteria providing essential nutrients lacking in the diet. Previous studies have evidenced that some of these bacterial symbionts are involved in the virus transmission. Wolbachia is a group of obligate intracellular bacteria infecting numerous arthropod species and causing reproductive alterations in their hosts. These bacteria have been detected in planthopper species, considered rice pests in various regions of the world. To date, Wolbachia infection status of planthopper species of Argentina is unknown. Amplification by PCR and sequencing of 16S rDNA, wsp- and ftsZ-specific genes demonstrated Wolbachia infection in Caenodelphax teapae (Fowler), Delphacodes kuscheli Fennah, Pyrophagus tigrinus Remes Lenicov & Varela, Tagosodes orizicolus (Muir), and Toya propinqua (Fieber). This is the first report of Wolbachia in delphacid vectors of viruses affecting maize and wheat. An understanding of the bacterial diversity harbored by these insect vectors could lead to new options for future management of diseases of economically important crops in a developing country. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  18. Data fusion and machine learning to identify threat vectors for the Zika virus and classify vulnerability

    Science.gov (United States)

    Gentle, J. N., Jr.; Kahn, A.; Pierce, S. A.; Wang, S.; Wade, C.; Moran, S.

    2016-12-01

    With the continued spread of the zika virus in the United States in both Florida and Virginia, increased public awareness, prevention and targeted prediction is necessary to effectively mitigate further infection and propagation of the virus throughout the human population. The goal of this project is to utilize publicly accessible data and HPC resources coupled with machine learning algorithms to identify potential threat vectors for the spread of the zika virus in Texas, the United States and globally by correlating available zika case data collected from incident reports in medical databases (e.g., CDC, Florida Department of Health) with known bodies of water in various earth science databases (e.g., USGS NAQWA Data, NASA ASTER Data, TWDB Data) and by using known mosquito population centers as a proxy for trends in population distribution (e.g., WHO, European CDC, Texas Data) while correlating historical trends in the spread of other mosquito borne diseases (e.g., chikungunya, malaria, dengue, yellow fever, west nile, etc.). The resulting analysis should refine the identification of the specific threat vectors for the spread of the virus which will correspondingly increase the effectiveness of the limited resources allocated towards combating the disease through better strategic implementation of defense measures. The minimal outcome of this research is a better understanding of the factors involved in the spread of the zika virus, with the greater potential to save additional lives through more effective resource utilization and public outreach.

  19. Dengue and Zika viruses: lessons learned from the similarities between these Aedes mosquito-vectored arboviruses.

    Science.gov (United States)

    Suwanmanee, San; Luplertlop, Natthanej

    2017-02-01

    The currently spreading arbovirus epidemic is having a severe impact on human health worldwide. The two most common flaviviruses, dengue virus (DENV) and Zika virus (ZIKV), are transmitted through the same viral vector, Aedes spp. mosquitoes. Since the discovery of DENV in 1943, this virus has been reported to cause around 390 million human infections per year, approximately 500,000 of which require hospitalization and over 20,000 of which are lethal. The present DENV epidemic is primarily concentrated in Southeast Asia. ZIKV, which was discovered in 1952, is another important arthropod-borne flavivirus. The neurotropic role of ZIKV has been reported in infected newborns with microcephaly and in adults with Guillain-Barre syndrome. Despite DENV and ZIKV sharing the same viral vector, their complex pathogenic natures are poorly understood, and the infections they cause do not have specific treatments or effective vaccines. Therefore, this review will describe what is currently known about the clinical characteristics, pathogenesis mechanisms, and transmission of these two viruses. Better understanding of the interrelationships between DENV and ZIKV will provide a useful perspective for developing an effective strategy for controlling both viruses in the future.

  20. Vector competence of Culicoides sonorensis (Diptera: Ceratopogonidae to epizootic hemorrhagic disease virus serotype 7

    Directory of Open Access Journals (Sweden)

    Ruder Mark G

    2012-10-01

    Full Text Available Abstract Background Culicoides sonorensis (Diptera: Ceratopogonidae is a vector of epizootic hemorrhagic disease virus (EHDV serotypes 1 and 2 in North America, where these viruses are well-known pathogens of white-tailed deer (WTD and other wild ruminants. Although historically rare, reports of clinical EHDV infection in cattle have increased in some parts of the world over the past decade. In 2006, an EHDV-7 epizootic in cattle resulted in economic loss for the Israeli dairy industry. White-tailed deer are susceptible to EHDV-7 infection and disease; however, this serotype is exotic to the US and the susceptibility of C. sonorensis to this cattle-virulent EHDV is not known. The objective of the study was to determine if C. sonorensis is susceptible to EHDV-7 infection and is a competent vector. Methods To evaluate the susceptibility of C. sonorensis, midges were fed on EHDV-7 infected WTD, held at 22 ± 1°C, and processed individually for virus isolation and titration on 4–16 days post feeding (dpf. Midges with a virus titer of ≥102.7 median tissue culture infective doses (TCID50/midge were considered potentially competent. To determine if infected C. sonorensis were capable of transmitting EHDV-7 to a host, a susceptible WTD was then fed on by a group of 14–16 dpf midges. Results From 4–16 dpf, 45% (156/350 of midges that fed on WTD with high titer viremia (>107 TCID50/ml were virus isolation-positive, and starting from 10–16 dpf, 32% (35/109 of these virus isolation-positive midges were potentially competent (≥102.7 TCID50/midge. Midges that fed on infected deer transmitted the virus to a susceptible WTD at 14–16 dpf. The WTD developed viremia and severe clinical disease. Conclusion This study demonstrates that C. sonorensis is susceptible to EHDV-7 infection and can transmit the virus to susceptible WTD, thus, C. sonorensis should be considered a potential vector of EHDV-7. Together with previous work, this study demonstrates

  1. Self-Complementary Adeno-Associated Virus Vectors Improve Transduction Efficiency of Corneal Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Anja K Gruenert

    Full Text Available Transplantation of a donor cornea to restore vision is the most frequently performed transplantation in the world. Corneal endothelial cells (CEC are crucial for the outcome of a graft as they maintain corneal transparency and avoid graft failure due to corneal opaqueness. Given the characteristic of being a monolayer and in direct contact with culture medium during cultivation in eye banks, CEC are specifically suitable for gene therapeutic approaches prior to transplantation. Recombinant adeno-associated virus 2 (rAAV2 vectors represent a promising tool for gene therapy of CEC. However, high vector titers are needed to achieve sufficient gene expression. One of the rate-limiting steps for transgene expression is the conversion of single-stranded (ss- DNA vector genome into double-stranded (ds- DNA. This step can be bypassed by using self-complementary (sc- AAV2 vectors. Aim of this study was to compare for the first time transduction efficiencies of ss- and scAAV2 vectors in CEC. For this purpose AAV2 vectors containing enhanced green fluorescent protein (GFP as transgene were used. Both in CEC and in donor corneas, transduction with scAAV2 resulted in significantly higher transgene expression compared to ssAAV2. The difference in transduction efficiency decreased with increasing vector titer. In most cases, only half the vector titer of scAAV2 was required for equal or higher gene expression rates than those of ssAAV2. In human donor corneas, GFP expression was 64.7±11.3% (scAAV and 38.0±8.6% (ssAAV (p<0.001, respectively. Furthermore, transduced cells maintained their viability and showed regular morphology. Working together with regulatory authorities, a translation of AAV2 vector-mediated gene therapy to achieve a temporary protection of corneal allografts during cultivation and transplantation could therefore become more realistic.

  2. Expression of IMP1 Enhances Production of Murine Leukemia Virus Vector by Facilitating Viral Genomic RNA Packaging

    OpenAIRE

    Mai, Yun; Gao, Guangxia

    2010-01-01

    Murine leukemia virus (MLV)-based retroviral vector is widely used for gene transfer. Efficient packaging of the genomic RNA is critical for production of high-titer virus. Here, we report that expression of the insulin-like growth factor II mRNA binding protein 1 (IMP1) enhanced the production of infectious MLV vector. Overexpression of IMP1 increased the stability of viral genomic RNA in virus producer cells and packaging of the RNA into progeny virus in a dose-dependent manner. Downregulat...

  3. Immunogenicity of ORFV-based vectors expressing the rabies virus glycoprotein in livestock species.

    Science.gov (United States)

    Martins, Mathias; Joshi, Lok R; Rodrigues, Fernando S; Anziliero, Deniz; Frandoloso, Rafael; Kutish, Gerald F; Rock, Daniel L; Weiblen, Rudi; Flores, Eduardo F; Diel, Diego G

    2017-11-01

    The parapoxvirus Orf virus (ORFV) encodes several immunomodulatory proteins (IMPs) that modulate host-innate and pro-inflammatory responses and has been proposed as a vaccine delivery vector for use in animal species. Here we describe the construction and characterization of two recombinant ORFV vectors expressing the rabies virus (RABV) glycoprotein (G). The RABV-G gene was inserted in the ORFV024 or ORFV121 gene loci, which encode for IMPs that are unique to parapoxviruses and inhibit activation of the NF-κB signaling pathway. The immunogenicity of the resultant recombinant viruses (ORFV ∆024 RABV-G or ORFV ∆121 RABV-G, respectively) was evaluated in pigs and cattle. Immunization of the target species with ORFV ∆024 RABV-G and ORFV ∆121 RABV-G elicited robust neutralizing antibody responses against RABV. Notably, neutralizing antibody titers induced in ORFV ∆121 RABV-G-immunized pigs and cattle were significantly higher than those detected in ORFV ∆024 RABV-G-immunized animals, indicating a higher immunogenicity of ORFV Δ121 -based vectors in these animal species. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Safety mechanism assisted by the repressor of tetracycline (SMART) vaccinia virus vectors for vaccines and therapeutics.

    Science.gov (United States)

    Grigg, Patricia; Titong, Allison; Jones, Leslie A; Yilma, Tilahun D; Verardi, Paulo H

    2013-09-17

    Replication-competent viruses, such as Vaccinia virus (VACV), are powerful tools for the development of oncolytic viral therapies and elicit superior immune responses when used as vaccine and immunotherapeutic vectors. However, severe complications from uncontrolled viral replication can occur, particularly in immunocompromised individuals or in those with other predisposing conditions. VACVs constitutively expressing interferon-γ (IFN-γ) replicate in cell culture indistinguishably from control viruses; however, they replicate in vivo to low or undetectable levels, and are rapidly cleared even in immunodeficient animals. In an effort to develop safe and highly effective replication-competent VACV vectors, we established a system to inducibly express IFN-γ. Our SMART (safety mechanism assisted by the repressor of tetracycline) vectors are designed to express the tetracycline repressor under a constitutive VACV promoter and IFN-γ under engineered tetracycline-inducible promoters. Immunodeficient SCID mice inoculated with VACVs not expressing IFN-γ demonstrated severe weight loss, whereas those given VACVs expressing IFN-γ under constitutive VACV promoters showed no signs of infection. Most importantly, mice inoculated with a VACV expressing the IFN-γ gene under an inducible promoter remained healthy in the presence of doxycycline, but exhibited severe weight loss in the absence of doxycycline. In this study, we developed a safety mechanism for VACV based on the conditional expression of IFN-γ under a tightly controlled tetracycline-inducible VACV promoter for use in vaccines and oncolytic cancer therapies.

  5. Evolution of codon usage in Zika virus genomes is host and vector specific

    Science.gov (United States)

    Butt, Azeem Mehmood; Nasrullah, Izza; Qamar, Raheel; Tong, Yigang

    2016-01-01

    The codon usage patterns of viruses reflect the evolutionary changes that allow them to optimize their survival and adapt their fitness to the external environment and, most importantly, their hosts. Here we report the genotype-specific codon usage patterns of Zika virus (ZIKV) strains from the current and previous outbreaks. Several genotype-specific and common codon usage traits were noted in the ZIKV coding sequences, indicating their independent evolutionary origins from a common ancestor. The overall influence of natural selection was more profound than that of mutation pressure, acting on a specific set of viral genes in the Asian-genotype ZIKV strains from the recent outbreak. An interplay between codon adaptation and deoptimization may have allowed the virus to adapt to multiple host and vectors and is reported for the first time in ZIKV genomes. Combining our codon analysis with geographical data on Aedes populations in the Americas suggested that ZIKV has evolved host- and vector-specific codon usage patterns to maintain successful replication and transmission chains within multiple hosts and vectors. PMID:27729643

  6. Vectors, hosts, and control measures for Zika virus in the Americas

    Science.gov (United States)

    Thompson, Sarah J.; Pearce, John; Ramey, Andy M.

    2017-01-01

    We examine Zika virus (ZIKV) from an ecological perspective and with a focus on the Americas. We assess (1) the role of wildlife in ZIKV disease ecology, (2) how mosquito behavior and biology influence disease dynamics, and (3) how nontarget species and ecosystems may be impacted by vector control programs. Our review suggests that free-ranging, non-human primates may be involved in ZIKV transmission in the Old World; however, other wildlife species likely play a limited role in maintaining or transmitting ZIKV. In the Americas, a zoonotic cycle has not yet been definitively established. Understanding behaviors and habitat tolerances of Aedes aegypti and Aedes albopictus, two ZIKV competent vectors in the Americas, will allow more accurate modeling of disease spread and facilitate targeted and effective control efforts. Vector control efforts may have direct and indirect impacts to wildlife, particularly invertebrate feeding species; however, strategies could be implemented to limit detrimental ecological effects.

  7. Generation of a Replication-Competent, Propagation-Deficient Virus Vector Based on the Transmissible Gastroenteritis Coronavirus Genome

    OpenAIRE

    Ortego, Javier; Escors, David; Laude, Hubert; Enjuanes, Luis

    2002-01-01

    Replication-competent propagation-deficient virus vectors based on the transmissible gastroenteritis coronavirus (TGEV) genome that are deficient in the essential E gene have been developed by complementation within E+ packaging cell lines. Cell lines expressing the TGEV E protein were established using the noncytopathic Sindbis virus replicon pSINrep21. In addition, cell lines stably expressing the E gene under the CMV promoter have been developed. The Sindbis replicon vector and the ectopic...

  8. Preventive and curative effects of Apple latent spherical virus vectors harboring part of the target virus genome against potyvirus and cucumovirus infections.

    Science.gov (United States)

    Tamura, Akihiro; Kato, Takahiro; Taki, Ayano; Sone, Mikako; Satoh, Nozomi; Yamagishi, Noriko; Takahashi, Tsubasa; Ryo, Bo-Song; Natsuaki, Tomohide; Yoshikawa, Nobuyuki

    2013-11-01

    Apple latent spherical virus (ALSV)-based vectors experimentally infect a broad range of plant species without causing symptoms and can effectively induce stable virus-induced gene silencing in plants. Here, we show that pre-infection of ALSV vectors harboring part of a target viral genome (we called ALSV vector vaccines here) inhibits the multiplication and spread of the corresponding challenge viruses [Bean yellow mosaic virus, Zucchini yellow mosaic virus (ZYMV), and Cucumber mosaic virus (CMV)] by a homology-dependent resistance. Further, the plants pre-infected with an ALSV vector having genome sequences of both ZYMV and CMV were protected against double inoculation of ZYMV and CMV. More interestingly, a curative effect of an ALSV vector vaccine could also be expected in ZYMV-infected cucumber plants, because the symptoms subsided on subsequent inoculation with an ALSV vector vaccine. This may be due to the invasion of ALSV, but not ZYMV, in the shoot apical meristem of cucumber. © 2013 Elsevier Inc. All rights reserved.

  9. Expression of the Surface Glycoproteins of Human Parainfluenza Virus Type 3 by Bovine Parainfluenza Virus Type 3, a Novel Attenuated Virus Vaccine Vector

    Science.gov (United States)

    Haller, Aurelia A.; Miller, Tessa; Mitiku, Misrach; Coelingh, Kathleen

    2000-01-01

    Bovine parainfluenza virus type 3 (bPIV3) is being evaluated as an intranasal vaccine for protection against human PIV3 (hPIV3). In young infants, the bPIV3 vaccine appears to be infectious, attenuated, immunogenic, and genetically stable, which are desirable characteristics for an RNA virus vector. To test the potential of the bPIV3 vaccine strain as a vector, an infectious DNA clone of bPIV3 was assembled and recombinant bPIV3 (r-bPIV3) was rescued. r-bPIV3 displayed a temperature-sensitive phenotype for growth in tissue culture at 39°C and was attenuated in the lungs of Syrian golden hamsters. In order to test whether r-bPIV3 could serve as a vector, the fusion and hemagglutinin-neuraminidase genes of bPIV3 were replaced with those of hPIV3. The resulting bovine/human PIV3 was temperature sensitive for growth in Vero cells at 37°C. The replication of bovine/human PIV3 was also restricted in the lungs of hamsters, albeit not as severely as was observed for r-bPIV3. Despite the attenuation phenotypes observed for r-bPIV3 and bovine/human PIV3, both of these viruses protected hamsters completely upon challenge with hPIV3. In summary, bPIV3 was shown to function as a virus vector that may be especially suitable for vaccination of infants and children against PIV3 and other viruses. PMID:11090161

  10. Rana grylio virus as a vector for foreign gene expression in fish cells.

    Science.gov (United States)

    He, Li-Bo; Ke, Fei; Zhang, Qi-Ya

    2012-01-01

    In the present study, Rana grylio virus (RGV, an iridovirus) thymidine kinase (TK) gene and viral envelope protein 53R gene were chosen as targets for foreign gene insertion. ΔTK-RGV and Δ53R-RGV, two recombinant RGV, expressing enhanced green fluorescence protein (EGFP) were constructed and analyzed in Epithelioma papulosum cyprinid (EPC) cells. The EGFP gene which fused to the virus major capsid protein (MCP) promoter p50 was inserted into TK and 53R gene loci of RGV, respectively. Cells infected with these two recombinant viruses not only displayed plaques, but also emitted strong green fluorescence under fluorescence microscope, providing a simple method for selection and purification of recombinant viruses. ΔTK-RGV was purified by seven successive rounds of plaque isolation and could be stably propagated in EPC cells. All of the plaques produced by the purified recombinant virus emitted green fluorescence. However, Δ53R-RGV was hard to be purified even through twenty rounds of plaque isolation. The purified recombinant virus ΔTK-RGV was verified by PCR analysis and Western blotting. These results showed EGFP was expressed in ΔTK-RGV infected cells. Furthermore, one-step growth curves and electron microscopy revealed that infection with recombinant ΔTK-RGV and wild-type RGV are similar. Therefore, RGV was demonstrated could be as a viral vector for foreign gene expression in fish cells. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

  11. Vector competence of northern and southern European Culex pipiens pipiens mosquitoes for West Nile virus across a gradient of temperatures

    NARCIS (Netherlands)

    Vogels, C.B.F.; Göertz, G.P.; Pijlman, G.P.; Koenraadt, C.J.M.

    2017-01-01

    In Europe, West Nile virus (WNV) outbreaks have been limited to southern and central European countries. However, competent mosquito vectors and susceptible bird hosts are present in northern Europe. Differences in temperature and vector competence of mosquito populations may explain the absence

  12. Swinepox virus vector-based vaccines: attenuation and biosafety assessments following subcutaneous prick inoculation.

    Science.gov (United States)

    Yuan, Xiaomin; Lin, Huixing; Li, Bin; He, Kongwang; Fan, Hongjie

    2018-02-07

    Swinepox virus (SPV) has several advantages as a potential clinical vector for a live vector vaccine. In this study, to obtain a safer and more efficient SPV vector, three SPV mutants, Δ003, Δ010, and ΔTK were successfully constructed. A virus replication experiment showed that these SPV mutants had lower replication abilities compared to wtSPV in 10 different host-derived cell lines. Animal experiments with mouse and rabbit models demonstrate that these three mutants and wtSPV did not cause any clinical signs of dermatitis. No fatalities were observed during a peritoneal challenge assay with these mutants and wtSPV in a mouse model. Additionally, the three mutants and wtSPV were not infectious at 60 h after vaccination in rabbit models. Furthermore, we evaluated biosafety, immunogenicity and effectiveness of the three mutants in 65 1-month-old piglets. The results show that there were no clinical signs of dermatitis in the Δ003 and ΔTK vaccination groups. However, mild signs were observed in the Δ010 vaccination groups when virus titres were high, and apparent clinical signs were observed at the sites of inoculation. Samples from all experimental pig groups were assessed by qPCR, and no SPV genomic DNA was found in five organs, faeces or blood. This suggests that the infectious abilities of wtSPV and the SPV mutants were poor and limited. In summary, this study indicates that two mutants of SPV, Δ003 and ΔTK, may be promising candidates for an attenuated viral vector in veterinary medicine.

  13. Adeno-Associated Virus Vectors and Stem Cells: Friends or Foes?

    Science.gov (United States)

    Brown, Nolan; Song, Liujiang; Kollu, Nageswara R; Hirsch, Matthew L

    2017-06-01

    The infusion of healthy stem cells into a patient-termed "stem-cell therapy"-has shown great promise for the treatment of genetic and non-genetic diseases, including mucopolysaccharidosis type 1, Parkinson's disease, multiple sclerosis, numerous immunodeficiency disorders, and aplastic anemia. Stem cells for cell therapy can be collected from the patient (autologous) or collected from another "healthy" individual (allogeneic). The use of allogenic stem cells is accompanied with the potentially fatal risk that the transplanted donor T cells will reject the patient's cells-a process termed "graft-versus-host disease." Therefore, the use of autologous stem cells is preferred, at least from the immunological perspective. However, an obvious drawback is that inherently as "self," they contain the disease mutation. As such, autologous cells for use in cell therapies often require genetic "correction" (i.e., gene addition or editing) prior to cell infusion and therefore the requirement for some form of nucleic acid delivery, which sets the stage for the AAV controversy discussed herein. Despite being the most clinically applied gene delivery context to date, unlike other more concerning integrating and non-integrating vectors such as retroviruses and adenovirus, those based on adeno-associated virus (AAV) have not been employed in the clinic. Furthermore, published data regarding AAV vector transduction of stem cells are inconsistent in regards to vector transduction efficiency, while the pendulum swings far in the other direction with demonstrations of AAV vector-induced toxicity in undifferentiated cells. The variation present in the literature examining the transduction efficiency of AAV vectors in stem cells may be due to numerous factors, including inconsistencies in stem-cell collection, cell culture, vector preparation, and/or transduction conditions. This review summarizes the controversy surrounding AAV vector transduction of stem cells, hopefully setting the

  14. A novel and highly efficient production system for recombinant adeno-associated virus vector.

    Science.gov (United States)

    Wu, Zhijian; Wu, Xiaobing; Cao, Hui; Dong, Xiaoyan; Wang, Hong; Hou, Yunde

    2002-02-01

    Recombinant adeno-associated virus (rAAV) has proven to be a promising gene delivery vector for human gene therapy. However, its application has been limited by difficulty in obtaining enough quantities of high-titer vector stocks. In this paper, a novel and highly efficient production system for rAAV is described. A recombinant herpes simplex virus type 1 (rHSV-1) designated HSV1-rc/DeltaUL2, which expressed adeno-associated virus type2 (AAV-2) Rep and Cap proteins, was constructed previously. The data confirmed that its functions were to support rAAV replication and packaging, and the generated rAAV was infectious. Meanwhile, an rAAV proviral cell line designated BHK/SG2, which carried the green fluorescent protein (GFP) gene expression cassette, was established by transfecting BHK-21 cells with rAAV vector plasmid pSNAV-2-GFP. Infecting BHK/SG2 with HSV1-rc/DeltaUL2 at an MOI of 0.1 resulted in the optimal yields of rAAV, reaching 250 transducing unit (TU) or 4.28x10(4) particles per cell. Therefore, compared with the conventional transfection method, the yield of rAAV using this "one proviral cell line, one helper virus" strategy was increased by two orders of magnitude. Large-scale production of rAAV can be easily achieved using this strategy and might meet the demands for clinical trials of rAAV-mediated gene therapy.

  15. Quantification of vector and host competence and abundance for Japanese Encephalitis Virus: a systematic review of the literature.

    Science.gov (United States)

    Japanese encephalitis (JE) is a vector-borne disease caused by the Japanese encephalitis virus (JEV) that affects humans in Eastern and Southeastern Asia. Although it could be prevented by a vaccine, JE has no treatment and the inadvertent introduction of the virus into JEV-free countries, such as t...

  16. Improving Dengue Virus Capture Rates in Humans and Vectors in Kamphaeng Phet Province, Thailand, Using an Enhanced Spatiotemporal Surveillance Strategy

    Science.gov (United States)

    2015-05-18

    THOMAS AND OTHERS ENHANCED SURVEILLANCE FOR DENGUE Improving Dengue Virus Capture Rates in Humans and Vectors in Kamphaeng Phet Province...of Medical Sciences, Bangkok, Thailand. Abstract. Dengue is of public health importance in tropical and sub-tropical regions. Dengue virus (DENV...with confirmed dengue (initiates) and associated cluster individuals (associates) with entomologic sampling. A total of 438 associates were enrolled

  17. Host Plants Indirectly Influence Plant Virus Transmission by Altering Gut Cysteine Protease Activity of Aphid Vectors*

    Science.gov (United States)

    Pinheiro, Patricia V.; Ghanim, Murad; Rebelo, Ana Rita; Santos, Rogerio S.; Orsburn, Benjamin C.; Gray, Stewart

    2017-01-01

    The green peach aphid, Myzus persicae, is a vector of the Potato leafroll virus (PLRV, Luteoviridae), transmitted exclusively by aphids in a circulative manner. PLRV transmission efficiency was significantly reduced when a clonal lineage of M. persicae was reared on turnip as compared with the weed physalis, and this was a transient effect caused by a host-switch response. A trend of higher PLRV titer in physalis-reared aphids as compared with turnip-reared aphids was observed at 24 h and 72 h after virus acquisition. The major difference in the proteomes of these aphids was the up-regulation of predicted lysosomal enzymes, in particular the cysteine protease cathepsin B (cathB), in aphids reared on turnip. The aphid midgut is the site of PLRV acquisition, and cathB and PLRV localization were starkly different in midguts of the aphids reared on the two host plants. In viruliferous aphids that were reared on turnip, there was near complete colocalization of cathB and PLRV at the cell membranes, which was not observed in physalis-reared aphids. Chemical inhibition of cathB restored the ability of aphids reared on turnip to transmit PLRV in a dose-dependent manner, showing that the increased activity of cathB and other cysteine proteases at the cell membrane indirectly decreased virus transmission by aphids. Understanding how the host plant influences virus transmission by aphids is critical for growers to manage the spread of virus among field crops. PMID:27932519

  18. Targeting Sindbis virus-based vectors to Fc receptor-positive cell types

    International Nuclear Information System (INIS)

    Klimstra, William B.; Williams, Jacqueline C.; Ryman, Kate D.; Heidner, Hans W.

    2005-01-01

    Some viruses display enhanced infection for Fc receptor (FcR)-positive cell types when complexed with virus-specific immunoglobulin (Ig). This process has been termed antibody-dependent enhancement of viral infection (ADE). We reasoned that the mechanism of ADE could be exploited and adapted to target alphavirus-based vectors to FcR-positive cell types. Towards this goal, recombinant Sindbis viruses were constructed that express 1 to 4 immunoglobulin-binding domains of protein L (PpL) as N-terminal extensions of the E2 glycoprotein. PpL is a bacterial protein that binds the variable region of antibody kappa light chains from a range of mammalian species. The recombinant viruses incorporated PpL/E2 fusion proteins into the virion structure and recapitulated the species-specific Ig-binding phenotypes of native PpL. Virions reacted with non-immune serum or purified IgG displayed enhanced binding and ADE for several species-matched FcR-positive murine and human cell lines. ADE required virus expression of a functional PpL Ig-binding domain, and appeared to be FcγR-mediated. Specifically, ADE did not occur with FcγR-negative cells, did not require active complement proteins, and did not occur on FcγR-positive murine cell lines when virions were bound by murine IgG-derived F(ab') 2 fragments

  19. Expression of Separate Proteins in the Same Plant Leaves and Cells Using Two Independent Virus-Based Gene Vectors

    Directory of Open Access Journals (Sweden)

    Maria R. Mendoza

    2017-11-01

    Full Text Available Plant viral vectors enable the expression of proteins at high levels in a relatively short time. For many purposes (e.g., cell biological interaction studies it may be desirable to express more than one protein in a single cell but that is often not feasible when using a single virus vector. Such a co-expression strategy requires the simultaneous delivery by two compatible and non-competitive viruses that can co-exist to each express a separate protein. Here, we report on the use of two agro-launchable coat-protein gene substitution GFP-expressing virus vector systems based on Tomato bushy stunt virus (TBSV referred to as TG, and Tobacco mosaic virus (TMV annotated as TRBO-G. TG expressed GFP in Nicotiana benthamiana, tomato, lettuce and cowpea, whereas expression from TRBO-G was detected only in the first two species. Upon co-infiltration of the two vectors co-expression was monitored by: molecular detection of the two slightly differently sized GFPs, suppressor-complementation assays, and using TG in combination with TRBO-RFP. All the results revealed that in N. benthamiana and tomato the TBSV and TMV vectors accumulated and expressed proteins in the same plants, the same leaves, and in the same cells. Therefore, co-expression by these two vectors provides a platform for fast and high level expression of proteins to study their cell biology or other properties.

  20. Transcriptome of the plant virus vector Graminella nigrifrons, and the molecular interactions of maize fine streak rhabdovirus transmission.

    Science.gov (United States)

    Chen, Yuting; Cassone, Bryan J; Bai, Xiaodong; Redinbaugh, Margaret G; Michel, Andrew P

    2012-01-01

    Leafhoppers (HEmiptera: Cicadellidae) are plant-phloem feeders that are known for their ability to vector plant pathogens. The black-faced leafhopper (Graminella nigrifrons) has been identified as the only known vector for the Maize fine streak virus (MFSV), an emerging plant pathogen in the Rhabdoviridae. Within G. nigrifrons populations, individuals can be experimentally separated into three classes based on their capacity for viral transmission: transmitters, acquirers and non-acquirers. Understanding the molecular interactions between vector and virus can reveal important insights in virus immune defense and vector transmission. RNA sequencing (RNA-Seq) was performed to characterize the transcriptome of G. nigrifrons. A total of 38,240 ESTs of a minimum 100 bp were generated from two separate cDNA libraries consisting of virus transmitters and acquirers. More than 60% of known D. melanogaster, A. gambiae, T. castaneum immune response genes mapped to our G. nigrifrons EST database. Real time quantitative PCR (RT-qPCR) showed significant down-regulation of three genes for peptidoglycan recognition proteins (PGRP - SB1, SD, and LC) in G. nigrifrons transmitters versus control leafhoppers. Our study is the first to characterize the transcriptome of a leafhopper vector species. Significant sequence similarity in immune defense genes existed between G. nigrifrons and other well characterized insects. The down-regulation of PGRPs in MFSV transmitters suggested a possible role in rhabdovirus transmission. The results provide a framework for future studies aimed at elucidating the molecular mechanisms of plant virus vector competence.

  1. Regional and seasonal response of a West Nile virus vector to climate change.

    Science.gov (United States)

    Morin, Cory W; Comrie, Andrew C

    2013-09-24

    Climate change will affect the abundance and seasonality of West Nile virus (WNV) vectors, altering the risk of virus transmission to humans. Using downscaled general circulation model output, we calculate a WNV vector's response to climate change across the southern United States using process-based modeling. In the eastern United States, Culex quinquefasciatus response to projected climate change displays a latitudinal and elevational gradient. Projected summer population depressions as a result of increased immature mortality and habitat drying are most severe in the south and almost absent further north; extended spring and fall survival is ubiquitous. Much of California also exhibits a bimodal pattern. Projected onset of mosquito season is delayed in the southwestern United States because of extremely dry and hot spring and summers; however, increased temperature and late summer and fall rains extend the mosquito season. These results are unique in being a broad-scale calculation of the projected impacts of climate change on a WNV vector. The results show that, despite projected widespread future warming, the future seasonal response of C. quinquefasciatus populations across the southern United States will not be homogeneous, and will depend on specific combinations of local and regional conditions.

  2. Spread of Zika virus: The key role of mosquito vector control

    Directory of Open Access Journals (Sweden)

    Giovanni Benelli

    2016-06-01

    Full Text Available Mosquitoes (Diptera: Culicidae represent a key threat for millions of humans and animals worldwide, since they act as vectors for important parasites and pathogens, including malaria, filariasis and a wide number of arboviruses. The recent outbreaks of Zika virus infections occurring in South America, Central America, and the Caribbean, represent the most recent four arrivals of important arboviruses in the western hemisphere, over the last 20 years, namely dengue, West Nile virus, and chikungunya. Since there are no specific treatments for Zika virus and the other arboviruses mentioned above, it should be highlighted that the eco-friendly and effective control of mosquito vectors is of pivotal importance. Besides radiation, transgenic and symbiont-based mosquito control approaches, an effective option may be the employ of biological control agents of mosquito young instars, in presence of ultra-low quantities of green-synthesized nanoparticles, which magnify their predation efficiency. Furthermore, behaviour-based control tools relying on the employ of swarming behaviour manipulation (i.e. the “lure and kill” approach, pheromone traps, sound traps need further research attention. In particular, detailed basic information on the physical and chemical cues routing mosquito swarming and mating dynamics is urgently required.

  3. Adenovirus vectors lacking virus-associated RNA expression enhance shRNA activity to suppress hepatitis C virus replication

    Science.gov (United States)

    Pei, Zheng; Shi, Guoli; Kondo, Saki; Ito, Masahiko; Maekawa, Aya; Suzuki, Mariko; Saito, Izumu; Suzuki, Tetsuro; Kanegae, Yumi

    2013-12-01

    First-generation adenovirus vectors (FG AdVs) expressing short-hairpin RNA (shRNA) effectively downregulate the expressions of target genes. However, this vector, in fact, expresses not only the transgene product, but also virus-associated RNAs (VA RNAs) that disturb cellular RNAi machinery. We have established a production method for VA-deleted AdVs lacking expression of VA RNAs. Here, we showed that the highest shRNA activity was obtained when the shRNA was inserted not at the popularly used E1 site, but at the E4 site. We then compared the activities of shRNAs against hepatitis C virus (HCV) expressed from VA-deleted AdVs or conventional AdVs. The VA-deleted AdVs inhibited HCV production much more efficiently. Therefore, VA-deleted AdVs were more effective than the currently used AdVs for shRNA downregulation, probably because of the lack of competition between VA RNAs and the shRNAs. These VA-deleted AdVs might enable more effective gene therapies for chronic hepatitis C.

  4. Repeated Delivery of Adeno-Associated Virus Vectors to the Rabbit Airway

    Science.gov (United States)

    Beck, Suzanne E.; Jones, Lori A.; Chesnut, Kye; Walsh, Scott M.; Reynolds, Thomas C.; Carter, Barrie J.; Askin, Frederic B.; Flotte, Terence R.; Guggino, William B.

    1999-01-01

    Efficient local expression from recombinant adeno-associated virus (rAAV)-cystic fibrosis (CF) transmembrane conductance regulator (CFTR) vectors has been observed in the airways of rabbits and monkeys for up to 6 months following a single bronchoscopic delivery. However, it is likely that repeated administrations of rAAV vectors will be necessary for sustained correction of the CF defect in the airways. The current study was designed to test the feasibility of repeated airway delivery of rAAV vectors in the rabbit lung. After two doses of rAAV-CFTR to the airways, rabbits generated high titers of serum anti-AAV neutralizing antibodies. Rabbits then received a third dose of a rAAV vector containing the green fluorescent protein (GFP) reporter gene packaged in either AAV serotype 2 (AAV2) or serotype 3 (AAV3) capsids. Each dose consisted of 1 ml containing 5 × 109 DNase-resistant particles of rAAV vector, having no detectable replication-competent AAV or adenovirus. Three weeks later, GFP expression was observed in airway epithelial cells despite high anti-AAV neutralizing titers at the time of delivery. There was no significant difference in the efficiency of DNA transfer or expression between the rAAV3 and rAAV2 groups. No significant inflammatory responses to either repeated airway exposure to rAAV2-CFTR vectors or to GFP expression were observed. These experiments demonstrate that serum anti-AAV neutralizing antibody titers do not predict airway neutralization in vivo and that repeated airway delivery rAAV allows for safe and effective gene transfer. PMID:10516053

  5. Infectious bursal disease virus as a replication-incompetent viral vector expressing green fluorescent protein.

    Science.gov (United States)

    Mosley, Yung-Yi C; Wu, Ching Ching; Lin, Tsang Long

    2017-01-01

    Infectious bursal disease virus (IBDV) has been established as a replication-competent viral vector capable of carrying an epitope at multiple loci in the genome. To enhance the safety and increase the insertion capacity of IBDV as a vector, a replication-incompetent IBDV vector was developed in the present study. The feasibility of replacing one of the viral gene loci, including pvp2, vp3, vp1, or the polyprotein vp243, with the sequence of green fluorescent protein (GFP) was explored. A method combining TCID 50 and immunoperoxidase monolayer assay (IPMA) determined the most feasible locus for gene replacement to be pvp2. The genomic segment containing gfp at the pvp2 locus was able to be encapsidated into IBDV particles. Furthermore, the expression of GFP in GFP-IBDV infected cells was confirmed by Western blotting and GFP-IBDV particles showed similar morphology and size to that of wildtype IBDV by electron microscopy. By providing the deleted protein in trans in a packaging cell line (pVP2-DF1), replication-incompetent GFP-IBDV particles were successfully plaque-quantified. The gfp sequence from the plaque-forming GFP-IBDV in pVP2-DF1 was confirmed by RT-PCR and sequencing. To our knowledge, GFP-IBDV developed in the present study is the first replication-incompetent IBDV vector which expresses a foreign protein in infected cells without the capability to produce viral progeny. Additionally, such replication-incompetent IBDV vectors could serve as bivalent vaccine vectors for conferring protection against infections with IBDV and other economically important, or zoonotic, avian pathogens.

  6. A marker-free system for highly efficient construction of vaccinia virus vectors using CRISPR Cas9

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    Ming Yuan

    Full Text Available The current method for creation of vaccinia virus (VACV vectors involves using a selection and purification marker, however inclusion of a gene without therapeutic value in the resulting vector is not desirable for clinical use. The Cre-LoxP system has been used to make marker-free Poxviruses, but the efficiency was very low. To obtain a marker-free VACV vector, we developed marker gene excision systems to modify the thymidine kinase (TK region and N1L regions using Cre-Loxp and Flp-FRET systems respectively. CRISPR-Cas9 system significantly resulted in a high efficiency (∼90% in generation of marker gene-positive TK-mutant VACV vector. The marker gene (RFP could be excised from the recombinant virus using Cre recombinase. To make a marker-free VV vector with double gene deletions targeting the TK and N1L gene, we constructed a donor repair vector targeting the N1L gene, which can carry a therapeutic gene and the marker (RFP that could be excised from the recombinant virus using Flp recombinase. The marker-free system developed here can be used to efficiently construct VACV vectors armed with any therapeutic genes in the TK region or N1L region without marker genes. Our marker-free system platform has significant potential for development of new marker-free VACV vectors for clinical application.

  7. Exploring the Diversity of Plant DNA Viruses and Their Satellites Using Vector-Enabled Metagenomics on Whiteflies

    Science.gov (United States)

    Ng, Terry Fei Fan; Duffy, Siobain; Polston, Jane E.; Bixby, Elise; Vallad, Gary E.; Breitbart, Mya

    2011-01-01

    Current knowledge of plant virus diversity is biased towards agents of visible and economically important diseases. Less is known about viruses that have not caused major diseases in crops, or viruses from native vegetation, which are a reservoir of biodiversity that can contribute to viral emergence. Discovery of these plant viruses is hindered by the traditional approach of sampling individual symptomatic plants. Since many damaging plant viruses are transmitted by insect vectors, we have developed “vector-enabled metagenomics” (VEM) to investigate the diversity of plant viruses. VEM involves sampling of insect vectors (in this case, whiteflies) from plants, followed by purification of viral particles and metagenomic sequencing. The VEM approach exploits the natural ability of highly mobile adult whiteflies to integrate viruses from many plants over time and space, and leverages the capability of metagenomics for discovering novel viruses. This study utilized VEM to describe the DNA viral community from whiteflies (Bemisia tabaci) collected from two important agricultural regions in Florida, USA. VEM successfully characterized the active and abundant viruses that produce disease symptoms in crops, as well as the less abundant viruses infecting adjacent native vegetation. PCR assays designed from the metagenomic sequences enabled the complete sequencing of four novel begomovirus genome components, as well as the first discovery of plant virus satellites in North America. One of the novel begomoviruses was subsequently identified in symptomatic Chenopodium ambrosiodes from the same field site, validating VEM as an effective method for proactive monitoring of plant viruses without a priori knowledge of the pathogens. This study demonstrates the power of VEM for describing the circulating viral community in a given region, which will enhance our understanding of plant viral diversity, and facilitate emerging plant virus surveillance and management of viral

  8. Recombinant Newcastle disease virus-vectored vaccines against human and animal infectious diseases.

    Science.gov (United States)

    Duan, Zhiqiang; Xu, Houqiang; Ji, Xinqin; Zhao, Jiafu

    2015-01-01

    Recent advances in recombinant genetic engineering techniques have brought forward a leap in designing new vaccines in modern medicine. One attractive strategy is the application of reverse genetics technology to make recombinant Newcastle disease virus (rNDV) deliver protective antigens of pathogens. In recent years, numerous studies have demonstrated that rNDV-vectored vaccines can induce quicker and better humoral and mucosal immune responses than conventional vaccines and are protective against pathogen challenges. With deeper understanding of NDV molecular biology, it is feasible to develop gene-modified rNDV vaccines accompanied by good safety, high efficacy, low toxicity and better immunogenicity. This review summarizes the development of reverse genetics technology in using NDV as a promising vaccine vector to design new vaccines for human and animal use.

  9. Disruption of Microtubules Post-Virus Entry Enhances Adeno-Associated Virus Vector Transduction

    Science.gov (United States)

    Xiao, Ping-Jie; Mitchell, Angela M.; Huang, Lu; Li, Chengwen; Samulski, R. Jude

    2016-01-01

    Perinuclear retention of viral particles is a poorly understood phenomenon observed during many virus infections. In this study, we investigated whether perinuclear accumulation acts as a barrier to limit recombinant adeno-associated virus (rAAV) transduction. After nocodazole treatment to disrupt microtubules at microtubule-organization center (MT-MTOC) after virus entry, we observed higher rAAV transduction. To elucidate the role of MT-MTOC in rAAV infection and study its underlying mechanisms, we demonstrated that rAAV's perinuclear localization was retained by MT-MTOC with fluorescent analysis, and enhanced rAAV transduction from MT-MTOC disruption was dependent on the rAAV capsid's nuclear import signals. Interestingly, after knocking down RhoA or inhibiting its downstream effectors (ROCK and Actin), MT-MTOC disruption failed to increase rAAV transduction or nuclear entry. These data suggest that enhancement of rAAV transduction is the result of increased trafficking to the nucleus via the RhoA-ROCK-Actin pathway. Ten-fold higher rAAV transduction was also observed by disrupting MT-MTOC in brain, liver, and tumor in vivo. In summary, this study indicates that virus perinuclear accumulation at MT-MTOC is a barrier-limiting parameter for effective rAAV transduction and defines a novel defense mechanism by which host cells restrain viral invasion. PMID:26942476

  10. An efficient virus-induced gene silencing vector for maize functional genomics research.

    Science.gov (United States)

    Wang, Rong; Yang, Xinxin; Wang, Nian; Liu, Xuedong; Nelson, Richard S; Li, Weimin; Fan, Zaifeng; Zhou, Tao

    2016-04-01

    Maize is a major crop whose rich genetic diversity provides an advanced resource for genetic research. However, a tool for rapid transient gene function analysis in maize that may be utilized in most maize cultivars has been lacking, resulting in reliance on time-consuming stable transformation and mutation studies to obtain answers. We developed an efficient virus-induced gene silencing (VIGS) vector for maize based on a naturally maize-infecting cucumber mosaic virus (CMV) strain, ZMBJ-CMV. An infectious clone of ZMBJ-CMV was constructed, and a vascular puncture inoculation method utilizing Agrobacterium was optimized to improve its utility for CMV infection of maize. ZMBJ-CMV was then modified to function as a VIGS vector. The ZMBJ-CMV vector induced mild to moderate symptoms in many maize lines, making it useful for gene function studies in critically important maize cultivars, such as the sequenced reference inbred line B73. Using this CMV VIGS system, expression of two endogenous genes, ZmPDS and ZmIspH, was found to be decreased by 75% and 78%, respectively, compared with non-silenced tissue. Inserts with lengths of 100-300 bp produced the most complete transcriptional and visual silencing phenotypes. Moreover, genes related to autophagy, ZmATG3 and ZmATG8a, were also silenced, and it was found that they function in leaf starch degradation. These results indicate that our ZMBJ-CMV VIGS vector provides a tool for rapid and efficient gene function studies in maize. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

  11. Vector Competence of Aedes aegypti and Aedes polynesiensis Populations from French Polynesia for Chikungunya Virus.

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    Vaea Richard

    2016-05-01

    Full Text Available From October 2014 to March 2015, French Polynesia experienced for the first time a chikungunya outbreak. Two Aedes mosquitoes may have contributed to chikungunya virus (CHIKV transmission in French Polynesia: the worldwide distributed Ae. aegypti and the Polynesian islands-endemic Ae. polynesiensis mosquito.To investigate the vector competence of French Polynesian populations of Ae. aegypti and Ae. polynesiensis for CHIKV, mosquitoes were exposed per os at viral titers of 7 logs tissue culture infectious dose 50%. At 2, 6, 9, 14 and 21 days post-infection (dpi, saliva was collected from each mosquito and inoculated onto C6/36 mosquito cells to check for the presence of CHIKV infectious particles. Legs and body (thorax and abdomen of each mosquito were also collected at the different dpi and submitted separately to viral RNA extraction and CHIKV real-time RT-PCR.CHIKV infection rate, dissemination and transmission efficiencies ranged from 7-90%, 18-78% and 5-53% respectively for Ae. aegypti and from 39-41%, 3-17% and 0-14% respectively for Ae. polynesiensis, depending on the dpi. Infectious saliva was found as early as 2 dpi for Ae. aegypti and from 6 dpi for Ae. polynesiensis. Our laboratory results confirm that the French Polynesian population of Ae. aegypti is highly competent for CHIKV and they provide clear evidence for Ae. polynesiensis to act as an efficient CHIKV vector.As supported by our findings, the presence of two CHIKV competent vectors in French Polynesia certainly contributed to enabling this virus to quickly disseminate from the urban/peri-urban areas colonized by Ae. aegypti to the most remote atolls where Ae. polynesiensis is predominating. Ae. polynesiensis was probably involved in the recent chikungunya outbreaks in Samoa and the Cook Islands. Moreover, this vector may contribute to the risk for CHIKV to emerge in other Polynesian islands like Fiji, and more particularly Wallis where there is no Ae. aegypti.

  12. The Immatures of Culicoides trilineatus (Diptera: Ceratopogonidae) Potential Vector of the Bluetongue Virus.

    Science.gov (United States)

    Diaz, F; Mangudo, C; Spinelli, G R; Gleiser, R M; Ronderos, M M

    2018-03-05

    The fourth instar larva and pupa of Culicoides trilineatus Fox (Diptera, Ceratopogonidae), a species considered as potential vector of the bluetongue virus in Central and South America, are described, illustrated, and photomicrographed for the first time by using binocular, phase-contrast, and scanning electron microscopy. The immatures were collected by using a siphon bottle in tree holes in Salta Province, Argentina, transported to the laboratory, and there reared to the adult's emergence. They are compared with the immatures of Culicoides debilipalpis Lutz (Diptera, Ceratopogonidae), another Neotropical species that develops in tree holes. Details on larval biology and habitat are given.

  13. Polydnaviruses of Parasitic Wasps: Domestication of Viruses To Act as Gene Delivery Vectors

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    Michael R. Strand

    2012-01-01

    Full Text Available Symbiosis is a common phenomenon in which associated organisms can cooperate in ways that increase their ability to survive, reproduce, or utilize hostile environments. Here, we discuss polydnavirus symbionts of parasitic wasps. These viruses are novel in two ways: (1 they have become non-autonomous domesticated entities that cannot replicate outside of wasps; and (2 they function as a delivery vector of genes that ensure successful parasitism of host insects that wasps parasitize. In this review we discuss how these novelties may have arisen, which genes are potentially involved, and what the consequences have been for genome evolution.

  14. Candidate Vectors and Rodent Hosts of Venezuelan Equine Encephalitis Virus, Chiapas, 2006–2007

    Science.gov (United States)

    Deardorff, Eleanor R.; Estrada-Franco, Jose G.; Freier, Jerome E.; Navarro-Lopez, Roberto; Da Rosa, Amelia Travassos; Tesh, Robert B.; Weaver, Scott C.

    2011-01-01

    Enzootic Venezuelan equine encephalitis virus (VEEV) has been known to occur in Mexico since the 1960s. The first natural equine epizootic was recognized in Chiapas in 1993 and since then, numerous studies have characterized the etiologic strains, including reverse genetic studies that incriminated a specific mutation that enhanced infection of epizootic mosquito vectors. The aim of this study was to determine the mosquito and rodent species involved in enzootic maintenance of subtype IE VEEV in coastal Chiapas. A longitudinal study was conducted over a year to discern which species and habitats could be associated with VEEV circulation. Antibody was rarely detected in mammals and virus was not isolated from mosquitoes. Additionally, Culex (Melanoconion) taeniopus populations were found to be spatially related to high levels of human and bovine seroprevalence. These mosquito populations were concentrated in areas that appear to represent foci of stable, enzootic VEEV circulation. PMID:22144461

  15. Heterologous prime-boost immunization of Newcastle disease virus vectored vaccines protected broiler chickens against highly pathogenic avian influenza and Newcastle disease viruses.

    Science.gov (United States)

    Kim, Shin-Hee; Samal, Siba K

    2017-07-24

    Avian Influenza virus (AIV) is an important pathogen for both human and animal health. There is a great need to develop a safe and effective vaccine for AI infections in the field. Live-attenuated Newcastle disease virus (NDV) vectored AI vaccines have shown to be effective, but preexisting antibodies to the vaccine vector can affect the protective efficacy of the vaccine in the field. To improve the efficacy of AI vaccine, we generated a novel vectored vaccine by using a chimeric NDV vector that is serologically distant from NDV. In this study, the protective efficacy of our vaccines was evaluated by using H5N1 highly pathogenic avian influenza virus (HPAIV) strain A/Vietnam/1203/2004, a prototype strain for vaccine development. The vaccine viruses were three chimeric NDVs expressing the hemagglutinin (HA) protein in combination with the neuraminidase (NA) protein, matrix 1 protein, or nonstructural 1 protein. Comparison of their protective efficacy between a single and prime-boost immunizations indicated that prime immunization of 1-day-old SPF chicks with our vaccine viruses followed by boosting with the conventional NDV vector strain LaSota expressing the HA protein provided complete protection of chickens against mortality, clinical signs and virus shedding. Further verification of our heterologous prime-boost immunization using commercial broiler chickens suggested that a sequential immunization of chickens with chimeric NDV vector expressing the HA and NA proteins following the boost with NDV vector expressing the HA protein can be a promising strategy for the field vaccination against HPAIVs and against highly virulent NDVs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Replication-competent infectious hepatitis B virus vectors carrying substantially sized transgenes by redesigned viral polymerase translation.

    Science.gov (United States)

    Wang, Zihua; Wu, Li; Cheng, Xin; Liu, Shizhu; Li, Baosheng; Li, Haijun; Kang, Fubiao; Wang, Junping; Xia, Huan; Ping, Caiyan; Nassal, Michael; Sun, Dianxing

    2013-01-01

    Viral vectors are engineered virus variants able to deliver nonviral genetic information into cells, usually by the same routes as the parental viruses. For several virus families, replication-competent vectors carrying reporter genes have become invaluable tools for easy and quantitative monitoring of replication and infection, and thus also for identifying antivirals and virus susceptible cells. For hepatitis B virus (HBV), a small enveloped DNA virus causing B-type hepatitis, such vectors are not available because insertions into its tiny 3.2 kb genome almost inevitably affect essential replication elements. HBV replicates by reverse transcription of the pregenomic (pg) RNA which is also required as bicistronic mRNA for the capsid (core) protein and the reverse transcriptase (Pol); their open reading frames (ORFs) overlap by some 150 basepairs. Translation of the downstream Pol ORF does not involve a conventional internal ribosome entry site (IRES). We reasoned that duplicating the overlap region and providing artificial IRES control for translation of both Pol and an in-between inserted transgene might yield a functional tricistronic pgRNA, without interfering with envelope protein expression. As IRESs we used a 22 nucleotide element termed Rbm3 IRES to minimize genome size increase. Model plasmids confirmed its activity even in tricistronic arrangements. Analogous plasmids for complete HBV genomes carrying 399 bp and 720 bp transgenes for blasticidin resistance (BsdR) and humanized Renilla green fluorescent protein (hrGFP) produced core and envelope proteins like wild-type HBV; while the hrGFP vector replicated poorly, the BsdR vector generated around 40% as much replicative DNA as wild-type HBV. Both vectors, however, formed enveloped virions which were infectious for HBV-susceptible HepaRG cells. Because numerous reporter and effector genes with sizes of around 500 bp or less are available, the new HBV vectors should become highly useful tools to better

  17. Replication-competent infectious hepatitis B virus vectors carrying substantially sized transgenes by redesigned viral polymerase translation.

    Directory of Open Access Journals (Sweden)

    Zihua Wang

    Full Text Available Viral vectors are engineered virus variants able to deliver nonviral genetic information into cells, usually by the same routes as the parental viruses. For several virus families, replication-competent vectors carrying reporter genes have become invaluable tools for easy and quantitative monitoring of replication and infection, and thus also for identifying antivirals and virus susceptible cells. For hepatitis B virus (HBV, a small enveloped DNA virus causing B-type hepatitis, such vectors are not available because insertions into its tiny 3.2 kb genome almost inevitably affect essential replication elements. HBV replicates by reverse transcription of the pregenomic (pg RNA which is also required as bicistronic mRNA for the capsid (core protein and the reverse transcriptase (Pol; their open reading frames (ORFs overlap by some 150 basepairs. Translation of the downstream Pol ORF does not involve a conventional internal ribosome entry site (IRES. We reasoned that duplicating the overlap region and providing artificial IRES control for translation of both Pol and an in-between inserted transgene might yield a functional tricistronic pgRNA, without interfering with envelope protein expression. As IRESs we used a 22 nucleotide element termed Rbm3 IRES to minimize genome size increase. Model plasmids confirmed its activity even in tricistronic arrangements. Analogous plasmids for complete HBV genomes carrying 399 bp and 720 bp transgenes for blasticidin resistance (BsdR and humanized Renilla green fluorescent protein (hrGFP produced core and envelope proteins like wild-type HBV; while the hrGFP vector replicated poorly, the BsdR vector generated around 40% as much replicative DNA as wild-type HBV. Both vectors, however, formed enveloped virions which were infectious for HBV-susceptible HepaRG cells. Because numerous reporter and effector genes with sizes of around 500 bp or less are available, the new HBV vectors should become highly useful tools to

  18. Geminiviral vectors based on bean yellow dwarf virus for production of vaccine antigens and monoclonal antibodies in plants.

    Science.gov (United States)

    Chen, Qiang; He, Junyun; Phoolcharoen, Waranyoo; Mason, Hugh S

    2011-03-01

    Expression of recombinant vaccine antigens and monoclonal antibodies using plant viral vectors has developed extensively during the past several years. The approach benefits from high yields of recombinant protein obtained within days after transient delivery of viral vectors to leaves of Nicotiana benthamiana, a tobacco relative. Modified viral genomes of both RNA and DNA viruses have been created. Geminiviruses such as bean yellow dwarf virus (BeYDV) have a small, single stranded DNA genome that replicates in the nucleus of an infected plant cell, using the cellular DNA synthesis apparatus and a virus-encoded replication initiator protein (Rep). BeYDV-derived expression vectors contain deletions of the viral genes encoding coat and movement proteins and insertion of an expression cassette for a protein of interest. Delivery of the geminiviral vector to leaf cells via Agrobacterium-mediated delivery produces very high levels of recombinant DNA that can act as a transcription template, yielding high levels of mRNA for the protein of interest. Several vaccine antigens, including Norwalk virus capsid protein and hepatitis B core antigen, were expressed using the BeYDV vector at levels up to 1 mg per g of leaf mass. BeYDV replicons can be stacked in the same vector molecule by linking them in tandem, which enables production of multi-subunit proteins like monoclonal antibody (mAb) heavy and light chains. The protective mAb 6D8 against Ebola virus was produced at 0.5 mg per g of leaf mass. Multi-replicon vectors could be conveniently used to produce protein complexes, e.g. virus-like particles that require two or more subunits.

  19. Vector competence of Aedes albopictus from Houston, Texas, for dengue serotypes 1 to 4, yellow fever and Ross River viruses.

    Science.gov (United States)

    Mitchell, C J; Miller, B R; Gubler, D J

    1987-09-01

    A combination of virus infection and transmission experiments showed that a Houston, Texas strain of Aedes albopictus is a competent vector for dengue (DEN), yellow fever (YF) and Ross River (RR) viruses. However, at 14 days incubation, DEN virus infection rates in a Puerto Rican strain of Aedes aegypti were significantly higher for each of the four DEN serotypes, except DEN-1, than in Houston Ae. albopictus fed simultaneously on the same virus suspensions. The degree of correlation between disseminated DEN infection rates in Houston Ae. albopictus and transmission to an in vitro system ranged from 42 to 88% for the four DEN serotypes. No significant difference was noted in YF virus infection rates or transmission rates in the two mosquito species fed on the same virus suspensions and incubated for the same time period. Also, RR virus infection and transmission rates in Houston and Hawaiian strains of Ae. albopictus were generally comparable.

  20. Inhibiting avian influenza virus shedding using a novel RNAi antiviral vector technology: proof of concept in an avian cell model.

    Science.gov (United States)

    Linke, Lyndsey M; Wilusz, Jeffrey; Pabilonia, Kristy L; Fruehauf, Johannes; Magnuson, Roberta; Olea-Popelka, Francisco; Triantis, Joni; Landolt, Gabriele; Salman, Mo

    2016-03-01

    Influenza A viruses pose significant health and economic threats to humans and animals. Outbreaks of avian influenza virus (AIV) are a liability to the poultry industry and increase the risk for transmission to humans. There are limitations to using the AIV vaccine in poultry, creating barriers to controlling outbreaks and a need for alternative effective control measures. Application of RNA interference (RNAi) techniques hold potential; however, the delivery of RNAi-mediating agents is a well-known obstacle to harnessing its clinical application. We introduce a novel antiviral approach using bacterial vectors that target avian mucosal epithelial cells and deliver (small interfering RNA) siRNAs against two AIV genes, nucleoprotein (NP) and polymerase acidic protein (PA). Using a red fluorescent reporter, we first demonstrated vector delivery and intracellular expression in avian epithelial cells. Subsequently, we demonstrated significant reductions in AIV shedding when applying these anti-AIV vectors prophylactically. These antiviral vectors provided up to a 10,000-fold reduction in viral titers shed, demonstrating in vitro proof-of-concept for using these novel anti-AIV vectors to inhibit AIV shedding. Our results indicate this siRNA vector technology could represent a scalable and clinically applicable antiviral technology for avian and human influenza and a prototype for RNAi-based vectors against other viruses.

  1. Improved Production Efficiency of Virus-Like Particles by the Baculovirus Expression Vector System.

    Directory of Open Access Journals (Sweden)

    Javier López-Vidal

    Full Text Available Vaccines based on virus-like particles (VLPs have proven effective in humans and animals. In this regard, the baculovirus expression vector system (BEVS is one of the technologies of choice to generate such highly immunogenic vaccines. The extended use of these vaccines for human and animal populations is constrained because of high production costs, therefore a significant improvement in productivity is crucial to ensure their commercial viability. Here we describe the use of the previously described baculovirus expression cassette, called TB, to model the production of two VLP-forming vaccine antigens in insect cells. Capsid proteins from porcine circovirus type 2 (PCV2 Cap and from the calicivirus that causes rabbit hemorrhagic disease (RHDV VP60 were expressed in insect cells using baculoviruses genetically engineered with the TB expression cassette. Productivity was compared to that obtained using standard counterpart vectors expressing the same proteins under the control of the polyhedrin promoter. Our results demonstrate that the use of the TB expression cassette increased the production yields of these vaccine antigens by around 300% with respect to the standard vectors. The recombinant proteins produced by TB-modified vectors were fully functional, forming VLPs identical in size and shape to those generated by the standard baculoviruses, as determined by electron microscopy analysis. The use of the TB expression cassette implies a simple modification of the baculovirus vectors that significantly improves the cost efficiency of VLP-based vaccine production, thereby facilitating the commercial viability and broad application of these vaccines for human and animal health.

  2. Preclinical evaluation of a recombinant adeno-associated virus vector expressing human alpha-1 antitrypsin made using a recombinant herpes simplex virus production method.

    Science.gov (United States)

    Chulay, Jeffrey D; Ye, Guo-Jie; Thomas, Darby L; Knop, David R; Benson, Janet M; Hutt, Julie A; Wang, Gensheng; Humphries, Margaret; Flotte, Terence R

    2011-02-01

    Recombinant adeno-associated virus (rAAV) vectors offer promise for gene therapy of alpha-1 antitrypsin (AAT) deficiency. A toxicology study in mice evaluated intramuscular injection of an rAAV vector expressing human AAT (rAAV-CB-hAAT) produced using a herpes simplex virus (HSV) complementation system or a plasmid transfection (TFX) method at doses of 3 × 10(11) vg (1.2 × 10(13) vg/kg) for both vectors and 2 × 10(12) vg (8 × 10(13) vg/kg) for the HSV-produced vector. The HSV-produced vector had favorable in vitro characteristics in terms of purity, efficiency of transduction, and hAAT expression. There were no significant differences in clinical findings or hematology and clinical chemistry values between test article and control groups and no gross pathology findings. Histopathological examination demonstrated minimal to mild changes in skeletal muscle at the injection site, consisting of focal chronic interstitial inflammation and muscle degeneration, regeneration, and vacuolization, in vector-injected animals. At the 3 × 10(11) vg dose, serum hAAT levels were higher with the HSV-produced vector than with the TFX-produced vector. With the higher dose of HSV-produced vector, the increase in serum hAAT levels was dose-proportional in females and greater than dose-proportional in males. Vector copy numbers in blood were highest 24 hr after dosing and declined thereafter, with no detectable copies present 90 days after dosing. Antibodies to hAAT were detected in almost all vector-treated animals, and antibodies to HSV were detected in most animals that received the highest vector dose. These results support continued development of rAAV-CB-hAAT for treatment of AAT deficiency.

  3. High-titer recombinant adeno-associated virus production utilizing a recombinant herpes simplex virus type I vector expressing AAV-2 Rep and Cap.

    Science.gov (United States)

    Conway, J E; Rhys, C M; Zolotukhin, I; Zolotukhin, S; Muzyczka, N; Hayward, G S; Byrne, B J

    1999-06-01

    Recombinant adeno-associated virus type 2 (rAAV) vectors have recently been used to achieve long-term, high level transduction in vivo. Further development of rAAV vectors for clinical use requires significant technological improvements in large-scale vector production. In order to facilitate the production of rAAV vectors, a recombinant herpes simplex virus type I vector (rHSV-1) which does not produce ICP27, has been engineered to express the AAV-2 rep and cap genes. The optimal dose of this vector, d27.1-rc, for AAV production has been determined and results in a yield of 380 expression units (EU) of AAV-GFP produced from 293 cells following transfection with AAV-GFP plasmid DNA. In addition, d27.1-rc was also efficient at producing rAAV from cell lines that have an integrated AAV-GFP provirus. Up to 480 EU/cell of AAV-GFP could be produced from the cell line GFP-92, a proviral, 293 derived cell line. Effective amplification of rAAV vectors introduced into 293 cells by infection was also demonstrated. Passage of rAAV with d27. 1-rc results in up to 200-fold amplification of AAV-GFP with each passage after coinfection of the vectors. Efficient, large-scale production (>109 cells) of AAV-GFP from a proviral cell line was also achieved and these stocks were free of replication-competent AAV. The described rHSV-1 vector provides a novel, simple and flexible way to introduce the AAV-2 rep and cap genes and helper virus functions required to produce high-titer rAAV preparations from any rAAV proviral construct. The efficiency and potential for scalable delivery of d27.1-rc to producer cell cultures should facilitate the production of sufficient quantities of rAAV vectors for clinical application.

  4. Vector Competence of French Polynesian Aedes aegypti and Aedes polynesiensis for Zika Virus.

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    Vaea Richard

    2016-09-01

    Full Text Available In 2013-2014, French Polynesia experienced for the first time a Zika outbreak. Two Aedes mosquitoes may have contributed to Zika virus (ZIKV transmission in French Polynesia: the worldwide distributed Ae. aegypti and the Polynesian islands-endemic Ae. polynesiensis mosquito.To evaluate their vector competence for ZIKV, mosquitoes were infected per os at viral titers of 7 logs tissue culture infectious dose 50%. At several days post-infection (dpi, saliva was collected from each mosquito and inoculated onto C6/36 mosquito cells to check for the presence of ZIKV infectious particles. Legs and body of each mosquito were also collected and submitted separately to RNA extraction and ZIKV RT-PCR. In Ae. aegypti the infection rate was high as early as 6 dpi and the dissemination efficiency get substantial from 9 dpi while the both rates remained quite low in Ae. polynesiensis. The transmission efficiency was poor in Ae. aegypti until 14 dpi and no infectious saliva was found in Ae. polynesiensis at the time points studied.In our experimental conditions, the late ability of the French Polynesian Ae. aegypti to transmit ZIKV added by the poor competence of Ae. polynesiensis for this virus suggest the possible contribution of another vector for the propagation of ZIKV during the outbreak, in particular in remote islands where Ae. polynesiensis is predominating.

  5. Apple latent spherical virus vectors for reliable and effective virus-induced gene silencing among a broad range of plants including tobacco, tomato, Arabidopsis thaliana, cucurbits, and legumes

    International Nuclear Information System (INIS)

    Igarashi, Aki; Yamagata, Kousuke; Sugai, Tomokazu; Takahashi, Yukari; Sugawara, Emiko; Tamura, Akihiro; Yaegashi, Hajime; Yamagishi, Noriko; Takahashi, Tsubasa; Isogai, Masamichi; Takahashi, Hideki; Yoshikawa, Nobuyuki

    2009-01-01

    Apple latent spherical virus (ALSV) vectors were evaluated for virus-induced gene silencing (VIGS) of endogenous genes among a broad range of plant species. ALSV vectors carrying partial sequences of a subunit of magnesium chelatase (SU) and phytoene desaturase (PDS) genes induced highly uniform knockout phenotypes typical of SU and PDS inhibition on model plants such as tobacco and Arabidopsis thaliana, and economically important crops such as tomato, legume, and cucurbit species. The silencing phenotypes persisted throughout plant growth in these plants. In addition, ALSV vectors could be successfully used to silence a meristem gene, proliferating cell nuclear antigen and disease resistant N gene in tobacco and RCY1 gene in A. thaliana. As ALSV infects most host plants symptomlessly and effectively induces stable VIGS for long periods, the ALSV vector is a valuable tool to determine the functions of interested genes among a broad range of plant species.

  6. Assessing the impact of climate change on vector-borne viruses in the EU through the elicitation of expert opinion.

    Science.gov (United States)

    Gale, P; Brouwer, A; Ramnial, V; Kelly, L; Kosmider, R; Fooks, A R; Snary, E L

    2010-02-01

    Expert opinion was elicited to undertake a qualitative risk assessment to estimate the current and future risks to the European Union (EU) from five vector-borne viruses listed by the World Organization for Animal Health. It was predicted that climate change will increase the risk of incursions of African horse sickness virus (AHSV), Crimean-Congo haemorrhagic fever virus (CCHFV) and Rift Valley fever virus (RVFV) into the EU from other parts of the world, with African swine fever virus (ASFV) and West Nile virus (WNV) being less affected. Currently the predicted risks of incursion were lowest for RVFV and highest for ASFV. Risks of incursion were considered for six routes of entry (namely vectors, livestock, meat products, wildlife, pets and people). Climate change was predicted to increase the risk of incursion from entry of vectors for all five viruses to some degree, the strongest effects being predicted for AHSV, CCHFV and WNV. This work will facilitate identification of appropriate risk management options in relation to adaptations to climate change.

  7. Vector status of Aedes species determines geographical risk of autochthonous Zika virus establishment.

    Directory of Open Access Journals (Sweden)

    Lauren Gardner

    2017-03-01

    Full Text Available The 2015-16 Zika virus pandemic originating in Latin America led to predictions of a catastrophic global spread of the disease. Since the current outbreak began in Brazil in May 2015 local transmission of Zika has been reported in over 60 countries and territories, with over 750 thousand confirmed and suspected cases. As a result of its range expansion attention has focused on possible modes of transmission, of which the arthropod vector-based disease spread cycle involving Aedes species is believed to be the most important. Additional causes of concern are the emerging new links between Zika disease and Guillain-Barre Syndrome (GBS, and a once rare congenital disease, microcephaly.Like dengue and chikungunya, the geographic establishment of Zika is thought to be limited by the occurrence of its principal vector mosquito species, Ae. aegypti and, possibly, Ae. albopictus. While Ae. albopictus populations are more widely established than those of Ae. aegypti, the relative competence of these species as a Zika vector is unknown. The analysis reported here presents a global risk model that considers the role of each vector species independently, and quantifies the potential spreading risk of Zika into new regions. Six scenarios are evaluated which vary in the weight assigned to Ae. albopictus as a possible spreading vector. The scenarios are bounded by the extreme assumptions that spread is driven by air travel and Ae. aegypti presence alone and spread driven equally by both species. For each scenario destination cities at highest risk of Zika outbreaks are prioritized, as are source cities in affected regions. Finally, intercontinental air travel routes that pose the highest risk for Zika spread are also ranked. The results are compared between scenarios.Results from the analysis reveal that if Ae. aegypti is the only competent Zika vector, then risk is geographically limited; in North America mainly to Florida and Texas. However, if Ae

  8. Vector status of Aedes species determines geographical risk of autochthonous Zika virus establishment.

    Science.gov (United States)

    Gardner, Lauren; Chen, Nan; Sarkar, Sahotra

    2017-03-01

    The 2015-16 Zika virus pandemic originating in Latin America led to predictions of a catastrophic global spread of the disease. Since the current outbreak began in Brazil in May 2015 local transmission of Zika has been reported in over 60 countries and territories, with over 750 thousand confirmed and suspected cases. As a result of its range expansion attention has focused on possible modes of transmission, of which the arthropod vector-based disease spread cycle involving Aedes species is believed to be the most important. Additional causes of concern are the emerging new links between Zika disease and Guillain-Barre Syndrome (GBS), and a once rare congenital disease, microcephaly. Like dengue and chikungunya, the geographic establishment of Zika is thought to be limited by the occurrence of its principal vector mosquito species, Ae. aegypti and, possibly, Ae. albopictus. While Ae. albopictus populations are more widely established than those of Ae. aegypti, the relative competence of these species as a Zika vector is unknown. The analysis reported here presents a global risk model that considers the role of each vector species independently, and quantifies the potential spreading risk of Zika into new regions. Six scenarios are evaluated which vary in the weight assigned to Ae. albopictus as a possible spreading vector. The scenarios are bounded by the extreme assumptions that spread is driven by air travel and Ae. aegypti presence alone and spread driven equally by both species. For each scenario destination cities at highest risk of Zika outbreaks are prioritized, as are source cities in affected regions. Finally, intercontinental air travel routes that pose the highest risk for Zika spread are also ranked. The results are compared between scenarios. Results from the analysis reveal that if Ae. aegypti is the only competent Zika vector, then risk is geographically limited; in North America mainly to Florida and Texas. However, if Ae. albopictus proves to be a

  9. Host Plants Indirectly Influence Plant Virus Transmission by Altering Gut Cysteine Protease Activity of Aphid Vectors.

    Science.gov (United States)

    Pinheiro, Patricia V; Ghanim, Murad; Alexander, Mariko; Rebelo, Ana Rita; Santos, Rogerio S; Orsburn, Benjamin C; Gray, Stewart; Cilia, Michelle

    2017-04-01

    The green peach aphid, Myzus persicae , is a vector of the Potato leafroll virus (PLRV, Luteoviridae), transmitted exclusively by aphids in a circulative manner. PLRV transmission efficiency was significantly reduced when a clonal lineage of M. persicae was reared on turnip as compared with the weed physalis, and this was a transient effect caused by a host-switch response. A trend of higher PLRV titer in physalis-reared aphids as compared with turnip-reared aphids was observed at 24 h and 72 h after virus acquisition. The major difference in the proteomes of these aphids was the up-regulation of predicted lysosomal enzymes, in particular the cysteine protease cathepsin B (cathB), in aphids reared on turnip. The aphid midgut is the site of PLRV acquisition, and cathB and PLRV localization were starkly different in midguts of the aphids reared on the two host plants. In viruliferous aphids that were reared on turnip, there was near complete colocalization of cathB and PLRV at the cell membranes, which was not observed in physalis-reared aphids. Chemical inhibition of cathB restored the ability of aphids reared on turnip to transmit PLRV in a dose-dependent manner, showing that the increased activity of cathB and other cysteine proteases at the cell membrane indirectly decreased virus transmission by aphids. Understanding how the host plant influences virus transmission by aphids is critical for growers to manage the spread of virus among field crops. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. West Nile virus surveillance in Europe: moving towards an integrated animal-human-vector approach.

    Science.gov (United States)

    Gossner, Céline M; Marrama, Laurence; Carson, Marianne; Allerberger, Franz; Calistri, Paolo; Dilaveris, Dimitrios; Lecollinet, Sylvie; Morgan, Dilys; Nowotny, Norbert; Paty, Marie-Claire; Pervanidou, Danai; Rizzo, Caterina; Roberts, Helen; Schmoll, Friedrich; Van Bortel, Wim; Gervelmeyer, Andrea

    2017-05-04

    This article uses the experience of five European countries to review the integrated approaches (human, animal and vector) for surveillance and monitoring of West Nile virus (WNV) at national and European levels. The epidemiological situation of West Nile fever in Europe is heterogeneous. No model of surveillance and monitoring fits all, hence this article merely encourages countries to implement the integrated approach that meets their needs. Integration of surveillance and monitoring activities conducted by the public health authorities, the animal health authorities and the authorities in charge of vector surveillance and control should improve efficiency and save resources by implementing targeted measures. The creation of a formal interagency working group is identified as a crucial step towards integration. Blood safety is a key incentive for public health authorities to allocate sufficient resources for WNV surveillance, while the facts that an effective vaccine is available for horses and that most infected animals remain asymptomatic make the disease a lesser priority for animal health authorities. The examples described here can support other European countries wishing to strengthen their WNV surveillance or preparedness, and also serve as a model for surveillance and monitoring of other (vector-borne) zoonotic infections. This article is copyright of The Authors, 2017.

  11. West Nile virus vector competency of Culex quinquefasciatus mosquitoes in the Galapagos Islands.

    Science.gov (United States)

    Eastwood, Gillian; Kramer, Laura D; Goodman, Simon J; Cunningham, Andrew A

    2011-09-01

    The mosquito-transmitted pathogen West Nile virus (WNV) is not yet present in the Galápagos Archipelago of Ecuador. However, concern exists for fragile endemic island fauna after population decreases in several North American bird species and pathology in certain reptiles. We examined WNV vector competency of a Galápagos strain of mosquito (Culex quinquefasciatus Say). Field specimens were tested for their capacity to transmit the WN02-1956 strain of WNV after incubation at 27°C or 30°C. Rates of infection, dissemination, and transmission all increased with days post-exposure to WNV, and the highest rates were observed at 28 days. Infection rates peaked at 59% and transmission rates peaked at 44% (of mosquitoes tested). Vector efficiency increased after day 14. Rates of infection but not of transmission were significantly influence by temperature. No vertical transmission was detectable. We demonstrate that Galápagos Cx. quinquefasciatus are competent WNV vectors, and therefore should be considered an animal and public health risk for the islands and controlled wherever possible.

  12. Hazard Characterization of Modified Vaccinia Virus Ankara Vector: What Are the Knowledge Gaps?

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    Malachy I. Okeke

    2017-10-01

    Full Text Available Modified vaccinia virus Ankara (MVA is the vector of choice for human and veterinary applications due to its strong safety profile and immunogenicity in vivo. The use of MVA and MVA-vectored vaccines against human and animal diseases must comply with regulatory requirements as they pertain to environmental risk assessment, particularly the characterization of potential adverse effects to humans, animals and the environment. MVA and recombinant MVA are widely believed to pose low or negligible risk to ecosystem health. However, key aspects of MVA biology require further research in order to provide data needed to evaluate the potential risks that may occur due to the use of MVA and MVA-vectored vaccines. The purpose of this paper is to identify knowledge gaps in the biology of MVA and recombinant MVA that are of relevance to its hazard characterization and discuss ongoing and future experiments aimed at providing data necessary to fill in the knowledge gaps. In addition, we presented arguments for the inclusion of uncertainty analysis and experimental investigation of verifiable worst-case scenarios in the environmental risk assessment of MVA and recombinant MVA. These will contribute to improved risk assessment of MVA and recombinant MVA vaccines.

  13. Promotion of Flowering by Apple Latent Spherical Virus Vector and Virus Elimination at High Temperature Allow Accelerated Breeding of Apple and Pear

    OpenAIRE

    Yamagishi, Norioko; Li, Chunjiang; Yoshikawa, Nobuyuki

    2016-01-01

    Plant viral vectors are superior tools for genetic manipulation, allowing rapid induction or suppression of expression of a target gene in plants. This is a particularly effective technology for use in breeding fruit trees, which are difficult to manipulate using recombinant DNA technologies. We reported previously that if apple seed embryos (cotyledons) are infected with an Apple latent spherical virus (ALSV) vector (ALSV-AtFT/MdTFL1) concurrently expressing the Arabidopsis thaliana florigen...

  14. Transcriptomic response of the insect vector, Peregrinus maidis, to Maize mosaic rhabdovirus and identification of conserved responses to propagative viruses in hopper vectors.

    Science.gov (United States)

    Martin, Kathleen M; Barandoc-Alviar, Karen; Schneweis, Derek J; Stewart, Catherine L; Rotenberg, Dorith; Whitfield, Anna E

    2017-09-01

    Maize mosaic virus (MMV) is a plant-pathogenic rhabdovirus that is transmitted by the corn planthopper, Peregrinus maidis, in a propagative manner. P. maidis supports long-term MMV infections with no negative effects on insect performance. To elucidate whole-body transcriptome responses to virus infection, RNA-Seq was used to examine differential gene expression of virus-infected adult insects, and libraries were prepared from replicated groups of virus-exposed insects and non-exposed insects. From the 68,003 de novo-assembled transcripts, 144 were differentially-expressed (DE) during viral infection with comparable numbers up- and down-regulated. DE transcripts with similarity to genes associated with transposable elements (i.e., RNA-directed DNA polymerases) were enriched and may represent a mechanisim for modulating virus infection. Comparison of the P. maidis DE transcripts to published propagative virus-responsive transcript databases for two other hopper vectors revealed that 16% of the DE transcripts were shared across the three systems and may represent conserved responses to propagative viruses. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. P and M gene junction is the optimal insertion site in Newcastle disease virus vaccine vector for foreign gene expression.

    Science.gov (United States)

    Zhao, Wei; Zhang, Zhenyu; Zsak, Laszlo; Yu, Qingzhong

    2015-01-01

    Newcastle disease virus (NDV) has been developed as a vector for vaccine and gene therapy purposes. However, the optimal insertion site for foreign gene expression remained to be determined. In the present study, we inserted the green fluorescence protein (GFP) gene into five different intergenic regions of the enterotropic NDV VG/GA vaccine strain using reverse genetics technology. The rescued recombinant viruses retained lentogenic pathotype and displayed delayed growth dynamics, particularly when the GFP gene was inserted between the NP and P genes of the virus. The GFP mRNA level was most abundant when the gene was inserted closer to the 3' end and gradually decreased as the gene was inserted closer to the 5' end. Measurement of the GFP fluorescence intensity in recombinant virus-infected cells demonstrated that the non-coding region between the P and M genes is the optimal insertion site for foreign gene expression in the VG/GA vaccine vector.

  16. Biology and management of Bemisia whitefly vectors of cassava virus pandemics in Africa.

    Science.gov (United States)

    Legg, James P; Shirima, Rudolph; Tajebe, Lensa S; Guastella, Devid; Boniface, Simon; Jeremiah, Simon; Nsami, Elibariki; Chikoti, Patrick; Rapisarda, Carmelo

    2014-10-01

    Cassava mosaic disease and cassava brown streak disease are caused by viruses transmitted by Bemisia tabaci and affect approximately half of all cassava plants in Africa, resulting in annual production losses of more than $US 1 billion. A historical and current bias towards virus rather than vector control means that these diseases continue to spread, and high Bemisia populations threaten future virus spread even if the extant strains and species are controlled. Progress has been made in parts of Africa in replicating some of the successes of integrated Bemisia control programmes in the south-western United States. However, these management efforts, which utilise chemical insecticides that conserve the Bemisia natural enemy fauna, are only suitable for commercial agriculture, which presently excludes most cassava cultivation in Africa. Initiatives to strengthen the control of B. tabaci on cassava in Africa need to be aware of this limitation, and to focus primarily on control methods that are cheap, effective, sustainable and readily disseminated, such as host-plant resistance and biological control. A framework based on the application of force multipliers is proposed as a means of prioritising elements of future Bemisia control strategies for cassava in Africa. © 2014 Society of Chemical Industry.

  17. The effect of laboratory colonization on the vector-pathogen interactions of Egyptian Culex pipiens and Rift Valley fever virus.

    Science.gov (United States)

    Gargan, T P; Bailey, C L; Higbee, G A; Gad, A; El Said, S

    1983-09-01

    Field and laboratory findings implicated Culex pipiens as a vector of Rift Valley fever (RVF) virus during the 1977-1978 epizootics/epidemics in Egypt. This study evaluated changes in infection and transmission rates, and viral titers in F1 through F16 generation Cx. pipiens mosquitoes orally infected with RVF virus. Infection and transmission rates of RVF virus by this species changed significantly during the colonization process. However, the ultimate viral titers of either the transmitting or the infected nontransmitting mosquitoes were not affected by the colonization process. Following ingestion of virus, Cx. pipiens could be separated into three distinct subpopulations, an uninfected group and two types of infected mosquitoes--transmitters and nontransmitters. Transmitters contained significantly more virus (approximately 100-fold) than nontransmitters. These results demonstrated that not every infected female mosquito should be considered a competent vector, even if the species (population) is known to be a primary vector. Transmission was also accomplished by probing mosquitoes which were unsuccessful in obtaining a blood meal. These data document the long-held suspicion that vector competence studies based upon laboratory-colonized specimens may not represent the field situation.

  18. The effects of monitoring the abundance and species composition of aphids as virus vectors on seed potato production in Serbia

    Directory of Open Access Journals (Sweden)

    Drago Milošević

    2014-03-01

    Full Text Available Aphids are the most important vectors of potato viruses during the crop’s growing season. The most widespread and damaging viruses, the potato virus Y and potato leaf roll virus, are transmitted by aphids in non-persistent and persistent manner, respectively. The two viruses cause the greatest concern of potato producers and a great constraint to seed potato production in Serbia, the region and across the world. Potato virus Y is particularly harmful, given its distribution and spreading rate. Seed potato production systems under well-managed conditions involve a series of virus control measures, including the monitoring of outbreaks of winged aphids, their abundance and species composition, in order to forecast virosis, i.e. potential plant and tuber infection periods. Monitoring the aphid vectors of potato viruses enables determination of optimum dates for haulm destruction when higher than normal numbers of winged aphids as vectors of economically harmful diseases have been observed. Haulm destruction in a potato crop reduces the risk of plant infection and virus translocation from the aboveground parts to tubers, thus keeping the proportion of infected tubers within tolerance limits allowed for certain categories of seed potatoes. This practice has positive effects if used in combination with other viral disease control measures; otherwise, it becomes ineffective. This paper provides an integral analysis of the effects and role of monitoring outbreaks of aphids, their abundance and species composition in timing haulm growth termination to prevent plant infection, virus translocation and tuber infestation in potato crops in Serbia and the wider region.

  19. Spatio-temporal patterns of distribution of West Nile virus vectors in eastern Piedmont Region, Italy

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    Bisanzio Donal

    2011-12-01

    Full Text Available Abstract Background West Nile Virus (WNV transmission in Italy was first reported in 1998 as an equine outbreak near the swamps of Padule di Fucecchio, Tuscany. No other cases were identified during the following decade until 2008, when horse and human outbreaks were reported in Emilia Romagna, North Italy. Since then, WNV outbreaks have occurred annually, spreading from their initial northern foci throughout the country. Following the outbreak in 1998 the Italian public health authority defined a surveillance plan to detect WNV circulation in birds, horses and mosquitoes. By applying spatial statistical analysis (spatial point pattern analysis and models (Bayesian GLMM models to a longitudinal dataset on the abundance of the three putative WNV vectors [Ochlerotatus caspius (Pallas 1771, Culex pipiens (Linnaeus 1758 and Culex modestus (Ficalbi 1890] in eastern Piedmont, we quantified their abundance and distribution in space and time and generated prediction maps outlining the areas with the highest vector productivity and potential for WNV introduction and amplification. Results The highest abundance and significant spatial clusters of Oc. caspius and Cx. modestus were in proximity to rice fields, and for Cx. pipiens, in proximity to highly populated urban areas. The GLMM model showed the importance of weather conditions and environmental factors in predicting mosquito abundance. Distance from the preferential breeding sites and elevation were negatively associated with the number of collected mosquitoes. The Normalized Difference Vegetation Index (NDVI was positively correlated with mosquito abundance in rice fields (Oc. caspius and Cx. modestus. Based on the best models, we developed prediction maps for the year 2010 outlining the areas where high abundance of vectors could favour the introduction and amplification of WNV. Conclusions Our findings provide useful information for surveillance activities aiming to identify locations where the

  20. Buckwheat stomatal traits under aluminium toxicity

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    Oleksandr E. Smirnov

    2014-04-01

    Full Text Available Aluminium influence on some stomatal parameters of common buckwheat (Fagopyrum esculentum Moench. was studied. Significant changes in stomatal density, stomatal index and stomatal shape coefficient under aluminium treatment were revealed. Stomatal closure and no difference in total stomatal potential conductance index of treatment plants were suggested as aluminium resistance characteristics.

  1. On the Seasonal Occurrence and Abundance of the Zika Virus Vector Mosquito Aedes Aegypti in the Contiguous United States

    OpenAIRE

    Monaghan, Andrew J.; Morin, Cory W.; Steinhoff, Daniel F.; Wilhelmi, Olga; Hayden, Mary; Quattrochi, Dale A.; Reiskind, Michael; Lloyd, Alun L.; Smith, Kirk; Schmidt, Chris A.; Scalf, Paige E.; Ernst, Kacey

    2016-01-01

    Introduction: An ongoing Zika virus pandemic in Latin America and the Caribbean has raised concerns that travel-related introduction of Zika virus could initiate local transmission in the United States (U.S.) by its primary vector, the mosquito Aedes aegypti. Methods: We employed meteorologically driven models for 2006-2015 to simulate the potential seasonal abundance of adult Aedes aegypti for fifty cities within or near the margins of its known U.S. range. Mosquito abundance results were an...

  2. Apparently nonspecific enzyme elevations after portal vein delivery of recombinant adeno-associated virus serotype 2 vector in hepatitis C virus-infected chimpanzees.

    Science.gov (United States)

    Flotte, Terence R; Goetzmann, Jason; Caridi, James; Paolillo, Joseph; Conlon, Thomas J; Potter, Mark; Mueller, Christian; Byrne, Barry J

    2008-07-01

    Hepatic gene transfer is envisioned as a substitute for protein replacement therapies, many of which are derived from blood products. Thus, the target populations may have a high prevalence of blood-borne pathogens, such as hepatitis C virus (HCV). We sought to determine whether the safety of recombinant adeno-associated virus serotype 2 (rAAV2) would be altered by preexisting HCV infection. Doses of approximately 1 x 10(13) vector genomes of an rAAV2-chimpanzee alpha(1)-antitrypsin (rAAV2-cAAT) vector were injected into the portal vein of each of three HCV genome-positive (HCV+) chimpanzees and three HCV-negative (HCV-) controls. Acute safety studies were performed up to 90 days after vector administration, along with analyses of the peripheral blood and liver tissue for rAAV2-cAAT genomes. Vector genome copy numbers in blood and liver tissue were similar in both groups. All animals demonstrated increases in liver and muscle enzyme levels after the pretreatment liver biopsy (5 days before vector injection) and after the vector injection. However, HCV+ animals demonstrated a substantially greater rise in aspartate aminotransferase, alanine aminotransferase, and creatinine phosphokinase values than HCV- animals. Histopathology demonstrated abnormal lipid accumulation (steatosis) in the hepatocytes of HCV+ animals, both before and after vector injection. These data indicate an increased susceptibility to subclinical liver toxicity from portal vein injection of rAAV2 in the presence of HCV infection.

  3. Vector Contact Rates on Eastern Bluebird Nestlings Do Not Indicate West Nile Virus Transmission in Henrico County, Virginia, USA

    Directory of Open Access Journals (Sweden)

    Kevin A. Caillouët

    2013-11-01

    Full Text Available Sensitive indicators of spatial and temporal variation in vector-host contact rates are critical to understanding the transmission and eventual prevention of arboviruses such as West Nile virus (WNV. Monitoring vector contact rates on particularly susceptible and perhaps more exposed avian nestlings may provide an advanced indication of local WNV amplification. To test this hypothesis we monitored WNV infection and vector contact rates among nestlings occupying nest boxes (primarily Eastern bluebirds; Sialia sialis, Turdidae across Henrico County, Virginia, USA, from May to August 2012. Observed host-seeking rates were temporally variable and associated with absolute vector and host abundances. Despite substantial effort to monitor WNV among nestlings and mosquitoes, we did not detect the presence of WNV in these populations. Generally low vector-nestling host contact rates combined with the negative WNV infection data suggest that monitoring transmission parameters among nestling Eastern bluebirds in Henrico County, Virginia, USA may not be a sensitive indicator of WNV activity.

  4. The feasibility of rabies virus-vectored immunocontraception in a mouse model

    Directory of Open Access Journals (Sweden)

    Xianfu Wu

    2014-01-01

    Full Text Available Immunocontraceptive vaccines may be an alternative to surgical sterilization. Dual rabies vaccination and dog population management is a helpful tool for rabies prevention. A synthetic gonadotropin-releasing hormone (GnRH peptide coupled to a carrier protein or T cell epitope is efficacious in inducing immunocontraception in a variety of mammals. However, virus-vectored GnRH recombinant vaccines have advantages over the conjugation method. In a previous in vitro study, we were able to insert a GnRH-coding sequence into the rabies virus (RABV glycoprotein (G gene, and the recombinant viruses grew to high titers in cells. Here, we further focused on the RABV G in accepting various copy numbers of GnRH. We demonstrated although RABV G protein with up to 4 copies of GnRH was well expressed, the recombinant virus was recovered only when 2 copies of GnRH (20 amino acids were incorporated into the G, indicating a possible insertion limit in making a full infectious clone. The investigation provides insight into the utility of RABV G as a carrier for small peptides and its suitability for vaccine studies. Following our previous study, we selected ERAg3p/2GnRH and tested the construct in mice. The vaccine induced ⩾80% infertility after three doses without any adjuvant, in live (8 of 10 mice infertility or inactivated (13 of 14 mice infertility formulations; while the pregnancy rate was 100% (10 of 10 mice in the controls. This initial success of immunocontraception in mice is promising, and we are now optimizing the vaccine formulation by using adjuvants and exploring novel delivery methods to minimize the dosage.

  5. Transcriptome of the plant virus vector Graminella nigrifrons, and the molecular interactions of maize fine streak rhabdovirus transmission.

    Directory of Open Access Journals (Sweden)

    Yuting Chen

    Full Text Available BACKGROUND: Leafhoppers (HEmiptera: Cicadellidae are plant-phloem feeders that are known for their ability to vector plant pathogens. The black-faced leafhopper (Graminella nigrifrons has been identified as the only known vector for the Maize fine streak virus (MFSV, an emerging plant pathogen in the Rhabdoviridae. Within G. nigrifrons populations, individuals can be experimentally separated into three classes based on their capacity for viral transmission: transmitters, acquirers and non-acquirers. Understanding the molecular interactions between vector and virus can reveal important insights in virus immune defense and vector transmission. RESULTS: RNA sequencing (RNA-Seq was performed to characterize the transcriptome of G. nigrifrons. A total of 38,240 ESTs of a minimum 100 bp were generated from two separate cDNA libraries consisting of virus transmitters and acquirers. More than 60% of known D. melanogaster, A. gambiae, T. castaneum immune response genes mapped to our G. nigrifrons EST database. Real time quantitative PCR (RT-qPCR showed significant down-regulation of three genes for peptidoglycan recognition proteins (PGRP - SB1, SD, and LC in G. nigrifrons transmitters versus control leafhoppers. CONCLUSIONS: Our study is the first to characterize the transcriptome of a leafhopper vector species. Significant sequence similarity in immune defense genes existed between G. nigrifrons and other well characterized insects. The down-regulation of PGRPs in MFSV transmitters suggested a possible role in rhabdovirus transmission. The results provide a framework for future studies aimed at elucidating the molecular mechanisms of plant virus vector competence.

  6. Use of a Recombinant Gamma-2 Herpesvirus Vaccine Vector against Dengue Virus in Rhesus Monkeys.

    Science.gov (United States)

    Bischof, Georg F; Magnani, Diogo M; Ricciardi, Michael; Shin, Young C; Domingues, Aline; Bailey, Varian K; Gonzalez-Nieto, Lucas; Rakasz, Eva G; Watkins, David I; Desrosiers, Ronald C

    2017-08-15

    Research on vaccine approaches that can provide long-term protection against dengue virus infection is needed. Here we describe the construction, immunogenicity, and preliminary information on the protective capacity of recombinant, replication-competent rhesus monkey rhadinovirus (RRV), a persisting herpesvirus. One RRV construct expressed nonstructural protein 5 (NS5), while a second recombinant expressed a soluble variant of the E protein (E85) of dengue virus 2 (DENV2). Four rhesus macaques received a single vaccination with a mixture of both recombinant RRVs and were subsequently challenged 19 weeks later with 1 × 10 5 PFU of DENV2. During the vaccine phase, plasma of all vaccinated monkeys showed neutralizing activity against DENV2. Cellular immune responses against NS5 were also elicited, as evidenced by major histocompatibility complex class I (MHC-I) tetramer staining in the one vaccinated monkey that was Mamu-A*01 positive. Unlike two of two unvaccinated controls, two of the four vaccinated monkeys showed no detectable viral RNA sequences in plasma after challenge. One of these two monkeys also showed no anamnestic increases in antibody levels following challenge and thus appeared to be protected against the acquisition of DENV2 following high-dose challenge. Continued study will be needed to evaluate the performance of herpesviral and other persisting vectors for achieving long-term protection against dengue virus infection. IMPORTANCE Continuing studies of vaccine approaches against dengue virus (DENV) infection are warranted, particularly ones that may provide long-term immunity against all four serotypes. Here we investigated whether recombinant rhesus monkey rhadinovirus (RRV) could be used as a vaccine against DENV2 infection in rhesus monkeys. Upon vaccination, all animals generated antibodies capable of neutralizing DENV2. Two of four vaccinated monkeys showed no detectable viral RNA after subsequent high-dose DENV2 challenge at 19 weeks

  7. Control of aphid-vectored and thrips-borne virus spread in lily, tulip, iris and dahlia by sprays of mineral oil, polydimethylsiloxane and pyrethroid insecticide in the field

    NARCIS (Netherlands)

    Asjes, J.; Blom-Barnhoorn, G.J.

    2001-01-01

    In this study control of spread by insect vectors of non-persistent Lily symptomless virus and Lily mottle virus in lily, Tulip breaking virus in tulip, Iris mild mosaic virus, Narcissus latent virus and Iris severe mosaic virus in bulbous iris, and semi-persistent Dahlia mosaic virus and persistent

  8. Adenovirus vectored vaccines against influenza a virus do not result in vaccine associated enhanced respiratory disease following heterologous challenge in contrast to whole inactivated virus vaccine

    Science.gov (United States)

    Heterologous influenza A virus (IAV) challenge following vaccination with an intramuscular (IM) whole inactivated vaccine (WIV) can result in vaccine-associated enhanced respiratory disease (VAERD). The objective of this study was to use an adenovirus (Ad5) vector vaccine platform that expressed IAV...

  9. Rise and Fall of Vector Infectivity During Sequential Strain Displacements by Mosquito-Borne Dengue Virus

    Science.gov (United States)

    Andrade, Christy C.; Young, Katherine I.; Johnson, William L.; Villa, Maria E.; Buraczyk, Cynthia A.; Messer, William B.; Hanley, Kathryn A.

    2016-01-01

    Each of the four serotypes of mosquito-borne dengue virus (DENV-1-4) comprises multiple, genetically-distinct strains. Competitive displacement between strains within a serotype is a common feature of DENV epidemiology and can trigger outbreaks of dengue disease. We investigated the mechanisms underlying two sequential displacements by DENV-3 strains in Sri Lanka that each coincided with abrupt increases in dengue hemorrhagic fever (DHF) incidence. First, the post-DHF strain, displaced the pre-DHF strain in the 1980s. We have previously shown that post-DHF is more infectious than pre-DHF for the major DENV vector, Aedes aegypti. Then, the ultra-DHF strain evolved in situ from post-DHF and displaced its ancestor in the 2000s. We predicted that ultra-DHF would be more infectious for Ae. aegypti than post-DHF but found that ultra-DHF infected a significantly lower percentage of mosquitoes than post-DHF. We therefore hypothesized that ultra-DHF had effected displacement by disseminating in Ae. aegypti more rapidly than post-DHF, but this was not borne out by a timecourse of mosquito infection. To elucidate the mechanisms that shape these virus-vector interactions, we tested the impact of RNA interference, the principal mosquito defense against DENV, on replication of each of the three DENV strains. Replication of all strains was similar in mosquito cells with dysfunctional RNAi, but in cells with functional RNAi, replication of pre-DHF was significantly suppressed relative to the other two strains. Thus differences in susceptibility to RNAi may account for the differences in mosquito infectivity between pre-DHF and post-DHF, but other mechanisms underlie the difference between post-DHF and ultra-DHF. PMID:27500505

  10. [Feeding pattern of Rift Valley Fever virus vectors in Senegal. Implications in the disease epidemiology].

    Science.gov (United States)

    Ba, Y; Diallo, D; Dia, I; Diallo, M

    2006-10-01

    During the rainy season 2003, an entomological survey was undertaken in the Sahelian bioclimatic zone of the Ferlo area in northern Senegal, in order to evaluate the degree of interaction between Rift valley fever (RVF) virus vectors and domestic animals and to determine the role of natural vertebrate hosts in the transmission and maintenance cycle. The study of vector-host contact was carried out under bed net traps using man, cow, sheep, chicken as bait whereas the RVFV vectors-vertebrate host interactions were studied through the analysis by an ELISA technique of the origin of the blood meals from naturally engorged females collected by aspiration. Blood meals sources were determined using a set of eight antibodies. Overall, the different known RVFV vectors (Culex poicilipes, Aedes vexans and Aedes ochraceus) were opportunistic although the bovine-baited net was, as far the more effective trap with 53.6% of collected mosquitoes. It was followed by the sheep-baited net (16.7%), man-baited net (12.6%) and chicken-baited net (11.6%). The more effectiveness of the bovine-baited net confirms the degree of implication of this host in RVF epidemiology. The study of vector-hosts interactions in nature showed that among the 1,112 mosquito blood meals tested, 701 were identified of which 693 were from Aedes vexans. The percentage of non-reacting blood meal was 36.7% whereas 16.9 % of the blood meals were taken at least on two vertebrate hosts. Overall, 53.2% of the blood meals from Ae. vexans were taken on equine, 18.6% on bovines, 7.1% on sheep and 0.6% on human. No blood meal was taken on rodent. The greatest diversity was observed in August. These host feedings patterns show that although equine is known to play a minor role in RVF epidemiology a thorough attention should be made to this host with regard to the percentage of blood meals taken in this host. The low percentage of blood meals taken on human could probably explain the low human infection rate observed up

  11. The role of environmental, virological and vector interactions in dictating biological transmission of arthropod-borne viruses by mosquitoes.

    Science.gov (United States)

    Kenney, Joan L; Brault, Aaron C

    2014-01-01

    Arthropod-borne viruses (arboviruses) are transmitted between vertebrate hosts and arthropod vectors. An inherently complex interaction among virus, vector, and the environment determines successful transmission of the virus. Once believed to be "flying syringes," recent advances in the field have demonstrated that mosquito genetics, microbiota, salivary components, and mosquito innate immune responses all play important roles in modulating arbovirus transmissibility. The literature on the interaction among virus, mosquito, and environment has expanded dramatically in the preceding decade and the utilization of next-generation sequencing and transgenic vector methodologies assuredly will increase the pace of knowledge acquisition in this field. This chapter outlines the interplay among the three factors in both direct physical and biochemical manners as well as indirectly through superinfection barriers and altered induction of innate immune responses in mosquito vectors. The culmination of the aforementioned interactions and the arms race between the mosquito innate immune response and the capacity of arboviruses to antagonize such a response ultimately results in the subjugation of mosquito cells for viral replication and subsequent transmission. © 2014 Elsevier Inc. All rights reserved.

  12. Apple latent spherical virus vector-induced flowering for shortening the juvenile phase in Japanese gentian and lisianthus plants.

    Science.gov (United States)

    Fekih, Rym; Yamagishi, Noriko; Yoshikawa, Nobuyuki

    2016-07-01

    Infection by apple latent spherical virus (ALSV) vectors that promote the expression of Arabidopsis thaliana FLOWERING LOCUS T ( AtFT ) or Gentiana triflora GtFT s accelerates flowering in gentian and lisianthus plants. Apple latent spherical virus (ALSV) has isometric virus particles (25 nm in diameter) that contain two ssRNA species (RNA1 and RNA2) and three capsid proteins (Vp25, Vp20, and Vp24). ALSV vectors are used for foreign gene expression and virus-induced gene silencing in a broad range of plant species. Here, we report the infection by ALSV vectors that express FLOWERING LOCUS T (AtFT) from Arabidopsis thaliana or its homolog GtFT1 from Gentiana triflora in three gentian cultivars ('Iwate Yume Aoi' [early flowering], 'Iwate' [medium flowering], and 'Alta' [late flowering]), and two lisianthus cultivars ('Newlination Pink ver. 2' and 'Torukogikyou daburu mikkusu') promotes flowering within 90 days post-inoculation using particle bombardment. Additionally, seedlings from the progeny of the early-flowering plants were tested by tissue blot hybridization, and the results showed that ALSV was not transmitted to the next generation. The promotion of flowering in the family Gentianaceae by ALSV vectors shortened the juvenile phase from 1-3 years to 3-5 months, and thus, it could be considered as a new plant breeding technique in ornamental gentian and lisianthus plants.

  13. Hepatitis virus protein X-Phenylalanine Hydroxylase fusion proteins identified in PKU mice treated with AAV-WPRE vectors

    Science.gov (United States)

    Utilizing the Pahenu2 mouse model for phenylketonuria (PKU), we developed an improved expression vector containing the Woodchuck Hepatitis Virus post-transcriptional regulatory element inserted into a rAAV-mPAH construct (rAAV-mPAH-WPRE) for treatment of PKU. Following portal vein delivery of these ...

  14. Gene therapy with adeno-associated virus vector 5-human factor IX in adults with hemophilia B

    DEFF Research Database (Denmark)

    Miesbach, Wolfgang; Meijer, Karina; Coppens, Michiel

    2018-01-01

    Hemophilia B gene therapy aims to ameliorate bleeding risk and provide endogenous factor IX (FIX) activity/synthesis through a single treatment, eliminating the requirement for FIX concentrate. AMT-060 combines an adeno-associated virus-5 (AAV5) vector with a liver-specific promoter driving...

  15. Beet necrotic yellow vein virus accumulates inside resting spores and zoosporangia of its vector Polymyxa betae BNYVV infects P. betae

    Directory of Open Access Journals (Sweden)

    Payton Mark

    2007-04-01

    Full Text Available Abstract Background Plasmodiophorids and chytrids are zoosporic parasites of algae and land plant and are distributed worldwide. There are 35 species belonging to the order Plasmodiophorales and three species, Polymyxa betae, P. graminis, and Spongospora subterranea, are plant viral vectors. Plasmodiophorid transmitted viruses are positive strand RNA viruses belonging to five genera. Beet necrotic yellow vein virus (BNYVV and its vector, P. betae, are the causal agents for rhizomania. Results Evidence of BNYVV replication and movement proteins associating with P. betae resting spores was initially obtained using immunofluorescence labeling and well characterized antisera to each of the BNYVV proteins. Root cross sections were further examined using immunogold labeling and electron microscopy. BNYVV proteins translated from each of the four genomic and subgenomic RNAs accumulate inside P. betae resting spores and zoospores. Statistical analysis was used to determine if immunolabelling detected viral proteins in specific subcellular domains and at a level greater than in control samples. Conclusion Virus-like particles were detected in zoosporangia. Association of BNYVV replication and movement proteins with sporangial and sporogenic stages of P. betae suggest that BNYVV resides inside its vector during more than one life cycle stage. These data suggest that P. betae might be a host as well as a vector for BNYVV

  16. Beet necrotic yellow vein virus accumulates inside resting spores and zoosporangia of its vector Polymyxa betae BNYVV infects P. betae.

    Science.gov (United States)

    Lubicz, Jeanmarie Verchot; Rush, Charles M; Payton, Mark; Colberg, Terry

    2007-04-05

    Plasmodiophorids and chytrids are zoosporic parasites of algae and land plant and are distributed worldwide. There are 35 species belonging to the order Plasmodiophorales and three species, Polymyxa betae, P. graminis, and Spongospora subterranea, are plant viral vectors. Plasmodiophorid transmitted viruses are positive strand RNA viruses belonging to five genera. Beet necrotic yellow vein virus (BNYVV) and its vector, P. betae, are the causal agents for rhizomania. Evidence of BNYVV replication and movement proteins associating with P. betae resting spores was initially obtained using immunofluorescence labeling and well characterized antisera to each of the BNYVV proteins. Root cross sections were further examined using immunogold labeling and electron microscopy. BNYVV proteins translated from each of the four genomic and subgenomic RNAs accumulate inside P. betae resting spores and zoospores. Statistical analysis was used to determine if immunolabelling detected viral proteins in specific subcellular domains and at a level greater than in control samples. Virus-like particles were detected in zoosporangia. Association of BNYVV replication and movement proteins with sporangial and sporogenic stages of P. betae suggest that BNYVV resides inside its vector during more than one life cycle stage. These data suggest that P. betae might be a host as well as a vector for BNYVV.

  17. Effects of insertion sites in a Newcastle disease virus vector on foreign gene expression through an internal ribosomal entry site

    Science.gov (United States)

    Newcastle disease virus (NDV), avian paramyxovirus type 1, has been developed as a vector to express foreign genes for vaccine and gene therapy purposes. The foreign genes are usually inserted into a non-coding region of the NDV genome as an independent transcription unit (ITU), which potentially a...

  18. Development of a Newcastle disease virus vector expressing a foreign gene through an internal ribosomal entry site

    Science.gov (United States)

    Newcastle disease virus (NDV) has been developed as a vector to express foreign genes for vaccine and gene therapy purposes. The foreign genes are usually inserted into a non-coding region of the NDV genome as an independent transcription unit (ITU). Based on the well-accepted “stop-start” transcr...

  19. A nuclear localization signal in the matrix of spleen necrosis virus (SNV) does not allow efficient gene transfer into quiescent cells with SNV-derived vectors.

    Science.gov (United States)

    Caron, Marie-Christine; Caruso, Manuel

    2005-08-01

    A major limitation in gene therapy for vectors derived from Moloney murine leukemia virus (MLV) is that they only deliver genes into dividing cells. In this study, a careful comparison of spleen necrosis virus (SNV)-derived vectors with MLV and human immunodeficiency virus (HIV)-1 retroviral vectors indicated that SNV vectors can deliver genes 4-fold more efficiently than MLV vectors into aphidicolin-arrested cells, although at a 25-fold lower efficiency than HIV-1-derived vectors. Furthermore, the addition of a NLS in the SNV matrix (MA) that mimics the one located in HIV-1 MA did not increase the ability of SNV vectors to transfer genes into arrested cells. Also, we found that the RD114 envelope was able to pseudotype SNV viral particles in a very efficient manner.

  20. Adeno-associated virus Rep-mediated targeting of integrase-defective retroviral vector DNA circles into human chromosome 19

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Shuohao [Graduate School of Systems Life Sciences, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Kawabe, Yoshinori; Ito, Akira [Department of Chemical Engineering, Faculty of Engineering, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Kamihira, Masamichi, E-mail: kamihira@chem-eng.kyushu-u.ac.jp [Graduate School of Systems Life Sciences, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Department of Chemical Engineering, Faculty of Engineering, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Adeno-associated virus (AAV) is capable of targeted integration in human cells. Black-Right-Pointing-Pointer Integrase-defective retroviral vector (IDRV) enables a circular DNA delivery. Black-Right-Pointing-Pointer A targeted integration system of IDRV DNA using the AAV integration mechanism. Black-Right-Pointing-Pointer Targeted IDRV integration ameliorates the safety concerns for retroviral vectors. -- Abstract: Retroviral vectors have been employed in clinical trials for gene therapy owing to their relative large packaging capacity, alterable cell tropism, and chromosomal integration for stable transgene expression. However, uncontrollable integrations of transgenes are likely to cause safety issues, such as insertional mutagenesis. A targeted transgene integration system for retroviral vectors, therefore, is a straightforward way to address the insertional mutagenesis issue. Adeno-associated virus (AAV) is the only known virus capable of targeted integration in human cells. In the presence of AAV Rep proteins, plasmids possessing the p5 integration efficiency element (p5IEE) can be integrated into the AAV integration site (AAVS1) in the human genome. In this report, we describe a system that can target the circular DNA derived from non-integrating retroviral vectors to the AAVS1 site by utilizing the Rep/p5IEE integration mechanism. Our results showed that after G418 selection 30% of collected clones had retroviral DNA targeted at the AAVS1 site.

  1. Using remote sensing and machine learning for the spatial modelling of a bluetongue virus vector

    Science.gov (United States)

    Van doninck, J.; Peters, J.; De Baets, B.; Ducheyne, E.; Verhoest, N. E. C.

    2012-04-01

    Bluetongue is a viral vector-borne disease transmitted between hosts, mostly cattle and small ruminants, by some species of Culicoides midges. Within the Mediterranean basin, C. imicola is the main vector of the bluetongue virus. The spatial distribution of this species is limited by a number of environmental factors, including temperature, soil properties and land cover. The identification of zones at risk of bluetongue outbreaks thus requires detailed information on these environmental factors, as well as appropriate epidemiological modelling techniques. We here give an overview of the environmental factors assumed to be constraining the spatial distribution of C. imicola, as identified in different studies. Subsequently, remote sensing products that can be used as proxies for these environmental constraints are presented. Remote sensing data are then used together with species occurrence data from the Spanish Bluetongue National Surveillance Programme to calibrate a supervised learning model, based on Random Forests, to model the probability of occurrence of the C. imicola midge. The model will then be applied for a pixel-based prediction over the Iberian peninsula using remote sensing products for habitat characterization.

  2. La Crosse Virus Field Detection and Vector Competence of Culex Mosquitoes.

    Science.gov (United States)

    Harris, M Camille; Yang, Fan; Jackson, Dorian M; Dotseth, Eric J; Paulson, Sally L; Hawley, Dana M

    2015-09-01

    La Crosse virus (LACV), a leading cause of arboviral pediatric encephalitis in the United States, is emerging in Appalachia. Here, we report field and laboratory evidence that suggest LACV may be using Culex mosquitoes as additional vectors in this region. This bunyavirus was detected by reverse-transcriptase polymerase chain reaction in two pools of Culex mosquitoes in southwestern Virginia and in six pools in West Virginia. To assess vector competence, we offered LACV blood meals to field-collected Culex restuans Theobald, Cx. pipiens L., and Aedes triseriatus (Say). Both Culex species were susceptible to infection. LACV-positive salivary expectorate, indicative of the ability to transmit, was detected in a small proportion of Cx. restuans (9%) and Cx. pipiens (4%) compared with Ae. triseriatus (40%). In a companion study of Cx. restuans only, we found that adults derived from nutritionally stressed larvae were significantly more likely to disseminate and transmit LACV. Our results indicate a potential role of Culex spp. in LACV dynamics that should be explored further in endemic areas. © The American Society of Tropical Medicine and Hygiene.

  3. Major QTLs Control Resistance to Rice Hoja Blanca Virus and Its Vector Tagosodes orizicolus

    Science.gov (United States)

    Romero, Luz E.; Lozano, Ivan; Garavito, Andrea; Carabali, Silvio J.; Triana, Monica; Villareal, Natalia; Reyes, Luis; Duque, Myriam C.; Martinez, César P.; Calvert, Lee; Lorieux, Mathias

    2013-01-01

    Rice hoja blanca (white leaf) disease can cause severe yield losses in rice in the Americas. The disease is caused by the rice hoja blanca virus (RHBV), which is transmitted by the planthopper vector Tagosodes orizicolus. Because classical breeding schemes for this disease rely on expensive, time-consuming screenings, there is a need for alternatives such as marker-aided selection. The varieties Fedearroz 2000 and Fedearroz 50, which are resistant to RHBV and to the feeding damage caused by T. orizicolus, were crossed with the susceptible line WC366 to produce segregating F2:3 populations. The F3 families were scored for their resistance level to RHBV and T. orizicolus. The F2:3 lines of both crosses were genotyped using microsatellite markers. One major QTL on the short arm of chromosome 4 was identified for resistance to RHBV in the two populations. Two major QTL on chromosomes 5 and 7 were identified for resistance to T. orizicolus in the Fd2000 × WC366 and Fd50 × WC366 crosses, respectively. This comparative study using two distinct rice populations allowed for a better understanding of how the resistance to RHBV and its vector are controlled genetically. Simple marker-aided breeding schemes based on QTL information can be designed to improve rice germplasm to reduce losses caused by this important disease. PMID:24240781

  4. Host feeding pattern of Japanese encephalitis virus vector mosquitoes (Diptera: Culicidae) from Kuttanadu, Kerala, India.

    Science.gov (United States)

    Philip Samuel, P; Arunachalam, N; Hiriyan, J; Tyagi, B K

    2008-09-01

    Identification of blood meals of vector mosquitoes is an important tool in the epidemiological investigations of vector-borne diseases. The blood meals of three mosquito species involved in the transmission of Japanese encephalitis virus (JEV) from the Kuttanadu area, Kerala, were determined using the agarose gel diffusion technique. A total of 4959 blood smears belonging to Culex (Culex) tritaeniorhynchus Giles (3273), Cx. (Culex) gelidus Theobald (64), Mansonia (Mnd.) indiana Edwards (735) ,and Ma. (Mnd.) uniformis (Theobald) (887) were tested. Cx. tritaeniorhynchus had predominantly fed on bovids (46.4%), and a good proportion (29%) had fed on more than one host. Cx. tritaeniorhynchus was highly zoophagic, and human feeding accounted for only 1.5% of those individuals successfully tested. Cx. gelidus showed bovid feeding at 36% and pig feeding at 12.5%. The test results showed 42.3% Ma. indiana and 12.2% Ma. uniformis had fed on humans. Multiple feeding was observed in Ma. indiana and Ma. uniformis, and most of the double feedings were from bovids and ovids (7.9 and 20.1%, respectively). Pig feeding accounted for 4.8% of the feedings by Cx. tritaeniorhynchus, 5.3% of Ma. indiana, and 6.4% of Ma. uniformis. This study is significant because of the role played by these mosquitoes in the transmission of JEV in the Kuttanadu area of Kerala, India.

  5. The yellow fever 17D vaccine virus: molecular basis of viral attenuation and its use as an expression vector

    Directory of Open Access Journals (Sweden)

    Galler R.

    1997-01-01

    Full Text Available The yellow fever (YF virus is the prototype flavivirus. The use of molecular techniques has unraveled the basic mechanisms of viral genome structure and expression. Recent trends in flavivirus research include the use of infectious clone technology with which it is possible to recover virus from cloned cDNA. Using this technique, mutations can be introduced at any point of the viral genome and their resulting effect on virus phenotype can be assessed. This approach has opened new possibilities to study several biological viral features with special emphasis on the issue of virulence/attenuation of the YF virus. The feasibility of using YF virus 17D vaccine strain, for which infectious cDNA is available, as a vector for the expression of heterologous antigens is reviewed

  6. Study of entomophatogenic fungus to control vector insect of citrus tristeza virus on citrus

    Directory of Open Access Journals (Sweden)

    Dwiastuti M.E.

    2017-08-01

    Full Text Available Citrus Tristeza Virus (CTV disease is a silent killer, which threatens to decrease productivity, quality and even death of citrus plants and the erosion of genetic resources. Spreading in the field very quickly by the intermediate insect vector pest, aphid (Toxoptera citricida, T. Aurantii and A. Gosypii. The microbes studied for potential biopesticide candidates are: Beauveria bassiana and Hirsutella citriformis, and Metarhizium anisopliae (Metch Sorokin previously reported to control Diaphorina citri pests resulting effectiveness of > 25% and was able to suppress yield loss up to 10%. The objectives of the study examined the effectiveness of entomopathogen in controlling the pest of CTV vector, Toxoptera citricida, in the laboratory and screen house, to findout the physiological, biochemical and molecular physiology of entomopathogen. The results showed that the best entomopathogen suspension concentration was B.bassiana 106 followed by H. citriformis 106 and M. anisopliae 106. Entomopatogen B. bassiana and H. citriformis effectively controled the CTV vector pest in the laboratory. In the semi-field experiments at the screen house, the most effective result was H.citriformis 106 and the combination of H.citriformis 106 + B.bassiana 106, killing up to 50% and 100% on day 7th H.citriformis had the most physiological character, was able to develop optimally at a temperature of 20-400C and humidity between 60-80%. The biochemical character of the entomopathogenic fungus B.bassiana contained cellulase enzyme and phosphate solvent and IAA hormone, at most compared to the others. H.citriformis had not been found to contain enzymes and hormones. The molecular biochemical characterization of entomopathogenic fungi using FS1 and NS2 primers more clearly distinguished isolates and entomopathogenic species.

  7. Complex adenovirus-vectored vaccine protects guinea pigs from three strains of Marburg virus challenges

    International Nuclear Information System (INIS)

    Wang Danher; Hevey, Michael; Juompan, Laure Y.; Trubey, Charles M.; Raja, Nicholas U.; Deitz, Stephen B.; Woraratanadharm, Jan; Luo Min; Yu Hong; Swain, Benjamin M.; Moore, Kevin M.; Dong, John Y.

    2006-01-01

    The Marburg virus (MARV), an African filovirus closely related to the Ebola virus, causes a deadly hemorrhagic fever in humans, with up to 90% mortality. Currently, treatment of disease is only supportive, and no vaccines are available to prevent spread of MARV infections. In order to address this need, we have developed and characterized a novel recombinant vaccine that utilizes a single complex adenovirus-vectored vaccine (cAdVax) to overexpress a MARV glycoprotein (GP) fusion protein derived from the Musoke and Ci67 strains of MARV. Vaccination with the cAdVaxM(fus) vaccine led to efficient production of MARV-specific antibodies in both mice and guinea pigs. Significantly, guinea pigs vaccinated with at least 5 x 10 7 pfu of cAdVaxM(fus) vaccine were 100% protected against lethal challenges by the Musoke, Ci67 and Ravn strains of MARV, making it a vaccine with trivalent protective efficacy. Therefore, the cAdVaxM(fus) vaccine serves as a promising vaccine candidate to prevent and contain multi-strain infections by MARV

  8. Mucosal immunization with integrase-defective lentiviral vectors protects against influenza virus challenge in mice.

    Directory of Open Access Journals (Sweden)

    Judith M Fontana

    Full Text Available Recent reports highlight the potential for integrase-defective lentiviral vectors (IDLV to be developed as vaccines due to their ability to elicit cell-mediated and humoral immune responses after intramuscular administration. Differently from their integrase-competent counterpart, whose utility for vaccine development is limited by the potential for insertional mutagenesis, IDLV possess a mutation in their integrase gene that prevents genomic integration. Instead, they are maintained as episomal DNA circles that retain the ability to stably express functional proteins. Despite their favorable profile, it is unknown whether IDLV elicit immune responses after intranasal administration, a route that could be advantageous in the case of infection with a respiratory agent. Using influenza as a model, we constructed IDLV expressing the influenza virus nucleoprotein (IDLV-NP, and tested their ability to generate NP-specific immune responses and protect from challenge in vivo. We found that administration of IDLV-NP elicited NP-specific T cell and antibody responses in BALB/c mice. Importantly, IDLV-NP was protective against homologous and heterosubtypic influenza virus challenge only when given by the intranasal route. This is the first report demonstrating that IDLV can induce protective immunity after intranasal administration, and suggests that IDLV may represent a promising vaccine platform against infectious agents.

  9. The Dengue Virus Mosquito Vector Aedes aegypti at High Elevation in México

    Science.gov (United States)

    Lozano-Fuentes, Saul; Hayden, Mary H.; Welsh-Rodriguez, Carlos; Ochoa-Martinez, Carolina; Tapia-Santos, Berenice; Kobylinski, Kevin C.; Uejio, Christopher K.; Zielinski-Gutierrez, Emily; Monache, Luca Delle; Monaghan, Andrew J.; Steinhoff, Daniel F.; Eisen, Lars

    2012-01-01

    México has cities (e.g., México City and Puebla City) located at elevations > 2,000 m and above the elevation ceiling below which local climates allow the dengue virus mosquito vector Aedes aegypti to proliferate. Climate warming could raise this ceiling and place high-elevation cities at risk for dengue virus transmission. To assess the elevation ceiling for Ae. aegypti and determine the potential for using weather/climate parameters to predict mosquito abundance, we surveyed 12 communities along an elevation/climate gradient from Veracruz City (sea level) to Puebla City (∼2,100 m). Ae. aegypti was commonly encountered up to 1,700 m and present but rare from 1,700 to 2,130 m. This finding extends the known elevation range in México by > 300 m. Mosquito abundance was correlated with weather parameters, including temperature indices. Potential larval development sites were abundant in Puebla City and other high-elevation communities, suggesting that Ae. aegypti could proliferate should the climate become warmer. PMID:22987656

  10. The dengue virus mosquito vector Aedes aegypti at high elevation in Mexico.

    Science.gov (United States)

    Lozano-Fuentes, Saul; Hayden, Mary H; Welsh-Rodriguez, Carlos; Ochoa-Martinez, Carolina; Tapia-Santos, Berenice; Kobylinski, Kevin C; Uejio, Christopher K; Zielinski-Gutierrez, Emily; Monache, Luca Delle; Monaghan, Andrew J; Steinhoff, Daniel F; Eisen, Lars

    2012-11-01

    México has cities (e.g., México City and Puebla City) located at elevations > 2,000 m and above the elevation ceiling below which local climates allow the dengue virus mosquito vector Aedes aegypti to proliferate. Climate warming could raise this ceiling and place high-elevation cities at risk for dengue virus transmission. To assess the elevation ceiling for Ae. aegypti and determine the potential for using weather/climate parameters to predict mosquito abundance, we surveyed 12 communities along an elevation/climate gradient from Veracruz City (sea level) to Puebla City (∼2,100 m). Ae. aegypti was commonly encountered up to 1,700 m and present but rare from 1,700 to 2,130 m. This finding extends the known elevation range in México by > 300 m. Mosquito abundance was correlated with weather parameters, including temperature indices. Potential larval development sites were abundant in Puebla City and other high-elevation communities, suggesting that Ae. aegypti could proliferate should the climate become warmer.

  11. Development of an equine-tropic replication-competent lentivirus assay for equine infectious anemia virus-based lentiviral vectors.

    Science.gov (United States)

    Farley, Daniel C; Bannister, Richard; Leroux-Carlucci, Marie A; Evans, Nerys E; Miskin, James E; Mitrophanous, Kyriacos A

    2012-10-01

    The release of lentiviral vectors for clinical use requires the testing of vector material, production cells, and, if applicable, ex vivo-transduced cells for the presence of replication-competent lentivirus (RCL). Vectors derived from the nonprimate lentivirus equine infectious anemia virus (EIAV) have been directly administered to patients in several clinical trials, with no toxicity observed to date. Because EIAV does not replicate in human cells, and because putative RCLs derived from vector components within human vector production cells would most likely be human cell-tropic, we previously developed an RCL assay using amphotropic murine leukemia virus (MLV) as a surrogate positive control and human cells as RCL amplification/indicator cells. Here we report an additional RCL assay that tests for the presence of theoretical "equine-tropic" RCLs. This approach provides further assurance of safety by detecting putative RCLs with an equine cell-specific tropism that might not be efficiently amplified by the human cell-based RCL assay. We tested the ability of accessory gene-deficient EIAV mutant viruses to replicate in a highly permissive equine cell line to direct our choice of a suitable EIAV-derived positive control. In addition, we report for the first time the mathematical rationale for use of the Poisson distribution to calculate minimal infectious dose of positive control virus and for use in monitoring assay positive/spike control failures in accumulating data sets. No RCLs have been detected in Good Manufacturing Practice (GMP)-compliant RCL assays to date, further demonstrating that RCL formation is highly unlikely in contemporary minimal lentiviral vector systems.

  12. Virus-induced down-regulation of GmERA1A and GmERA1B genes enhances the stomatal response to abscisic acid and drought resistance in soybean.

    Directory of Open Access Journals (Sweden)

    Takuya Ogata

    Full Text Available Drought is a major threat to global soybean production. The limited transformation potential and polyploid nature of soybean have hindered functional analysis of soybean genes. Previous research has implicated farnesylation in the plant's response to abscisic acid (ABA and drought tolerance. We therefore used virus-induced gene silencing (VIGS to evaluate farnesyltransferase genes, GmERA1A and GmERA1B (Glycine max Enhanced Response to ABA1-A and -B, as potential targets for increasing drought resistance in soybean. Apple latent spherical virus (ALSV-mediated GmERA1-down-regulated soybean leaves displayed an enhanced stomatal response to ABA and reduced water loss and wilting under dehydration conditions, suggesting that GmERA1A and GmERA1B negatively regulate ABA signaling in soybean guard cells. The findings provide evidence that the ALSV-VIGS system, which bypasses the need to generate transgenic plants, is a useful tool for analyzing gene function using only a single down-regulated leaf. Thus, the ALSV-VIGS system could constitute part of a next-generation molecular breeding pipeline to accelerate drought resistance breeding in soybean.

  13. DIPHTHERITIC STOMATITIS IN YELLOW-EYED PENGUINS (MEGADYPTES ANTIPODES) IN NEW ZEALAND.

    Science.gov (United States)

    Alley, Maurice R; Suepaul, Rod B; McKinlay, Bruce; Young, Melanie J; Wang, Jianning; Morgan, Kerri J; Hunter, Stuart A; Gartrell, Brett D

    2017-01-01

    Diphtheritic stomatitis is a seasonal disease that has been recognized as a syndrome in Yellow-eyed Penguin ( Megadyptes antipodes ) chicks in New Zealand for >10 yr. It was present in about 50% of 234 chicks examined since 2002 and is characterized by a thick serocellular exudate in the oral cavity of 1-4-wk-old chicks. The syndrome includes inanition, weight loss, and death in many affected birds. Microscopically, the lesions varied in severity. Most affected chicks had severe, locally extensive, ulcerative stomatitis with large amounts of exudate containing numerous bacteria; a smaller number had mild focal lesions with smaller amounts of exudate and bacteria. Although Corynebacterium amycolatum has been consistently isolated from the oral lesions, it was also present in the oral cavity of 34% of normal adult penguins and their chicks and is not known to possess diphtheritic toxins. A primary viral pathogen was therefore suspected, and intracytoplasmic inclusion bodies were occasionally seen in oral mucosal epithelial cells. No herpesvirus DNA was detected with PCR. Avipoxvirus DNA and an unidentified virus-like agent were detected in some early oral lesions, but could not be confirmed in subsequent testing. Electron microscopy on early affected epithelium with intracytoplasmic inclusion bodies was unrewarding. Our findings raise the possibility that the disease is caused by an unknown primary virus infection followed by secondary Corynebacterium invasion, but this requires confirmation. The means of transmission has not been established but insect vectors are suspected.

  14. Development of a reverse transcription loop-mediated isothermal amplification assay for the detection of vesicular stomatitis New Jersey virus: use of rapid molecular assay to differentiate between vesicular disease viruses

    Science.gov (United States)

    Vesicular stomatitis (VS) is endemic in Central America and northern regions of South America. Sporadic outbreaks of VS can occur in cattle and pigs where the clinical presentation can be similar to foot-and-mouth disease (FMD). There is therefore a pressing need for rapid, sensitive and specific d...

  15. Preclinical characterization of a recombinant adeno-associated virus type 1-pseudotyped vector demonstrates dose-dependent injection site inflammation and dissemination of vector genomes to distant sites.

    Science.gov (United States)

    Flotte, Terence R; Conlon, Thomas J; Poirier, Amy; Campbell-Thompson, Martha; Byrne, Barry J

    2007-03-01

    To translate the potential advantages of recombinant adeno-associated virus type 1 (rAAV1) vectors into a clinical application for muscle-directed gene therapy for alpha1 -antitrypsin (AAT) deficiency, we performed safety studies in 170 C57BL/6 mice and 26 New Zealand White rabbits. A mouse toxicology study included 8 cohorts of 10 mice each (5 per sex). Mice were killed either 21 or 90 days after intramuscular injection of doses ranging up to 1x10(13)vector genomes (VG), equivalent to 4 x 10(14)VG/kg. A mouse biodistribution study was performed in 5 cohorts of 10 mice, receiving intramuscular injections at the same doses; as well as in a lower dose cohort (3 x 10(8) VG; equivalent to 1.2 x 10(10)VG/kg); and in 4 other cohorts (excluding the vehicle control) injected with identical doses intravenously. Finally, biodistribution was examined in rabbits, with serial collection of blood and semen, as well as terminal tissue collection. Two significant findings were present, both of which were dose dependent. First, inflammatory cell infiltrates were detected at the site of injection 21, 60, or 90 days after intramuscular injection of 1 x 10(13)VG. This was not associated with loss of transgene expression. Second, vector DNA sequences were detected in most animals, levels being highest with the highest doses and earliest time points. Vector DNA was also present in liver, spleen, kidneys, and a number of other organs, including the gonads of animals receiving the highest dose. Likewise, vector DNA was present in the semen of male rabbits at higher doses. The copy number of vector DNA in the blood and semen declined over time throughout the study. These two dose-dependent findings have served to guide to the design of a phase 1 human trial of rAAV1-AAT.

  16. [Construction and identification of recombinant adeno-associated virus vector harboring fusion gene NT4-Apoptin-HA2-TAT].

    Science.gov (United States)

    Wang, Jian-Sheng; Zhang, Ming-Xin; Liu, De-Chun; Duan, Xiao-Yi; Zhou, Su-Na; Zhang, Guang-Jian; Yang, Guang-Xiao; Wang, Quan-Ying

    2008-08-01

    To construct a recombinant adeno-associated virus vector harboring fusion gene NT4-Apoptin-HA2-TAT, laying a foundation for gene therapy research of malignant tumors. The Apoptin and HA2-TAT gene were inserted in pUC19/NT4 vector after digested with restriction enzyme. The fusion gene of NT4-Apoptin-HA2-TAT was sub-cloned into the shuttle plasmid of adeno-associated virus; the products were co-transferred into HEK-293 cell line with helper plasmid pAAV/Ad and adeno-plasmid pFG140.The recombinant adeno-associated virus was produced by homologous recombination of above 3 plasmids in HEK-293 cells and its titer was measured by quantitative dot blot hybridization. The effect of AAV-NT4-Apoptin -HA2-TAT on HepG2 cell line was measured by a colorimetric 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay. The NT4-Apoptin-HA2-TAT was confirmed by restriction enzyme digestion and DNA sequencing. High titer of recombinant adeno-associated virus was obtained by homologous recombination in HEK-293 cells (3.14 x 10(15) pfu/L). AAV-NT4-Apoptin-HA2-TAT had strong deduce apoptosis effect on HepG2 cells. Compared with Adeno-associated mock virus and in normal cell line NIH3T3, Aden-associated virus NT4-Apoptin-HA2-TAT significantly decreased the survival rate of HepG2 cells. The recombinant adeno-associated virus vector encoding gene NT4-Apoptin-HA2-TAT has been successfully constructed in this experiment by molecular cloning and in vitro recombination techniques, laying a foundation for further research of gene therapy of cancer.

  17. Developing the framework for a risk map for mite vectored viruses in wheat resulting from pre-harvest hail damage

    Science.gov (United States)

    Nguy-Robertson, A. L.; Stilwell, A.; Zygielbaum, A. I.; McMechan, J.; Hein, G.; Wegulo, S.; Smith, T.

    2014-12-01

    Climate change is expected to adversely influence the weather and may increase the severity of meteorological events, including thunderstorms capable of producing crop damaging hail. Hail events occurring on pre-harvest wheat fields are known to be a significant contributor to outbreaks of virus diseases such as Wheat streak mosaic virus, Triticum mosaic virus , and Wheat mosaic virus, in the subsequent year. These viruses are vectored by the wheat curl mite (WCM) and volunteer wheat from pre-harvest hail events provides WCMs with a green bridge between harvest in the summer and planting in the fall. Controlling volunteer wheat through herbicide application and tillage has been shown to reduce WSMV transmission as long as it is conducted before emergence of the winter wheat crop. The goal of this study was to develop a risk product for WCM-vectored viruses based on pre-harvest hail damage. Traditionally, vegetation indices (VI; i.e. normalized difference vegetation index) are used to remotely identify hail damage in satellite imagery. However, during the highest risk period for volunteer wheat resulting from pre-harvest hail damage, the wheat crop is senescing and dramatically reducing its 'greenness', decreasing its sensitivity to changes in many VIs. Ground-based hyperspectral remote sensing over mechanically hailed sites indicates that reflectance from hailed wheat increases in the visible/near infrared spectrum. This information can be incorporated into a model using satellite imagery (LandSat and MODIS) and a 1 x 1 km resolution map of estimated maximal hail size produced by NOAA. The risk maps produced from these inputs will enable farmers and stakeholders to more effectively manage WCM-vectored viruses of wheat.

  18. Infection of Melanoplus sanguinipes Grasshoppers following Ingestion of Rangeland Plant Species Harboring Vesicular Stomatitis Virus▿

    Science.gov (United States)

    Drolet, Barbara S.; Stuart, Melissa A.; Derner, Justin D.

    2009-01-01

    Knowledge of the many mechanisms of vesicular stomatitis virus (VSV) transmission is critical for understanding of the epidemiology of sporadic disease outbreaks in the western United States. Migratory grasshoppers [Melanoplus sanguinipes (Fabricius)] have been implicated as reservoirs and mechanical vectors of VSV. The grasshopper-cattle-grasshopper transmission cycle is based on the assumptions that (i) virus shed from clinically infected animals would contaminate pasture plants and remain infectious on plant surfaces and (ii) grasshoppers would become infected by eating the virus-contaminated plants. Our objectives were to determine the stability of VSV on common plant species of U.S. Northern Plains rangelands and to assess the potential of these plant species as a source of virus for grasshoppers. Fourteen plant species were exposed to VSV and assayed for infectious virus over time (0 to 24 h). The frequency of viable virus recovery at 24 h postexposure was as high as 73%. The two most common plant species in Northern Plains rangelands (western wheatgrass [Pascopyrum smithii] and needle and thread [Hesperostipa comata]) were fed to groups of grasshoppers. At 3 weeks postfeeding, the grasshopper infection rate was 44 to 50%. Exposure of VSV to a commonly used grasshopper pesticide resulted in complete viral inactivation. This is the first report demonstrating the stability of VSV on rangeland plant surfaces, and it suggests that a significant window of opportunity exists for grasshoppers to ingest VSV from contaminated plants. The use of grasshopper pesticides on pastures would decrease the incidence of a virus-amplifying mechanical vector and might also decontaminate pastures, thereby decreasing the inter- and intraherd spread of VSV. PMID:19286779

  19. West nile virus prevalence across landscapes is mediated by local effects of agriculture on vector and host communities.

    Directory of Open Access Journals (Sweden)

    David W Crowder

    Full Text Available Arthropod-borne viruses (arboviruses threaten the health of humans, livestock, and wildlife. West Nile virus (WNV, the world's most widespread arbovirus, invaded the United States in 1999 and rapidly spread across the county. Although the ecology of vectors and hosts are key determinants of WNV prevalence across landscapes, the factors shaping local vector and host populations remain unclear. Here, we used spatially-explicit models to evaluate how three land-use types (orchards, vegetable/forage crops, natural and two climatic variables (temperature, precipitation influence the prevalence of WNV infections and vector/host distributions at landscape and local spatial scales. Across landscapes, we show that orchard habitats were associated with greater prevalence of WNV infections in reservoirs (birds and incidental hosts (horses, while increased precipitation was associated with fewer infections. At local scales, orchard habitats increased the prevalence of WNV infections in vectors (mosquitoes and the abundance of mosquitoes and two key reservoir species, the American robin and the house sparrow. Thus, orchard habitats benefitted WNV vectors and reservoir hosts locally, creating focal points for the transmission of WNV at landscape scales in the presence of suitable climatic conditions.

  20. The distribution of potential West Nile virus vectors, Culex pipiens pipiens and Culex pipiens quinquefasciatus (Diptera: Culicidae), in Mexico City

    OpenAIRE

    Diaz-Badillo, Alvaro; Bolling, Bethany G; Perez-Ramirez, Gerardo; Moore, Chester G; Martinez-Munoz, Jorge P; Padilla-Viveros, America A; Camacho-Nuez, Minerva; Diaz-Perez, Alfonso; Beaty, Barry J; de Lourdes Munoz, Maria

    2011-01-01

    Abstract Background Culex spp. mosquitoes are considered to be the most important vectors of West Nile virus (WNV) detected in at least 34 species of mosquitoes in the United States. In North America, Culex pipiens pipiens, Culex pipiens quinquefasciatus, and Culex tarsalis are all competent vectors of WNV, which is considered to be enzootic in the United States and has also been detected in equines and birds in many states of Mexico and in humans in Nuevo Leon. There is potential for WNV to ...

  1. Transcriptomics of differential vector competence: West Nile virus infection in two populations of Culex pipiens quinquefasciatus linked to ovary development.

    Science.gov (United States)

    Shin, Dongyoung; Civana, Ayse; Acevedo, Carolina; Smartt, Chelsea T

    2014-06-22

    Understanding mechanisms that contribute to viral dissemination in mosquito vectors will contribute to our ability to interfere with the transmission of viral pathogens that impact public health. The expression of genes in two Culex pipiens quinquefasciatus populations from Florida with known differences in vector competence to West Nile virus (WNV) were compared using high throughput sequencing. A total of 15,176 transcripts were combined for comparison of expression differences between the two populations and 118 transcripts were differentially expressed (p Culex populations in Florida. Both populations control energy allocations to reproduction as a response to WNV. This result provides novel insight into the defense mechanism used by Culex spp. mosquitoes against WNV.

  2. Use of a simian virus 40-based shuttle vector to analyze enhanced mutagenesis in mitomycin C-treated monkey cells

    International Nuclear Information System (INIS)

    Roilides, E.; Munson, P.J.; Levine, A.S.; Dixon, K.

    1988-01-01

    When monkey cells were treated with mitomycin C 24 h before transfection with UV-irradiated pZ189 (a simian virus 40-based shuttle vector), there was a twofold increase in the frequency of mutations in the supF gene of the vector. These results suggest the existence of an enhancible mutagenesis pathway in mammalian cells. However, DNA sequence analysis of the SupF- mutants suggested no dramatic changes in the mechanisms of mutagenesis due to mitomycin C treatment of the cells

  3. Dynamics of epizootic hemorrhagic disease virus infection within the vector, Culicoides sonorensis (Diptera: Ceratopogonidae.

    Directory of Open Access Journals (Sweden)

    Mary K Mills

    Full Text Available Culicoides sonorensis biting midges are confirmed vectors of epizootic hemorrhagic disease virus (EHDV, which causes mortality in white-tailed deer and ruminant populations. Currently, of the seven EHDV serotypes, only 1, 2, and 6 are detected in the USA, and very few studies have focused on the infection time course of these serotypes within the midge. The objective of this current research was to characterize EHDV-2 infection within the midge by measuring infection prevalence, virus dissemination, and viral load over the course of infection. Midges were fed a blood meal containing 106.9 PFU/ml EHDV-2, collected every 12 h from 0-2 days post feeding (dpf and daily from 3-10 dpf, and cohorts of 20 C. sonorensis were processed using techniques that assessed EHDV infection and dissemination. Cytopathic effect assays and quantitative (qPCR were used to determine infection prevalence, revealing a 50% infection rate by 10 dpf using both methods. Using immunohistochemistry, EHDV-2 infection was detectable at 5 dpf, and shown to disseminate from the midgut to other tissues, including fat body, eyes, and salivary glands by 5 dpf. Stain intensity increased from 5-8 dpf, indicating replication of EHDV-2 in secondary infection sites after dissemination. This finding is also supported by trends in viral load over time as determined by plaque assays and qPCR. An increase in titer between 4-5 dpf correlated with viral replication in the midgut as seen with staining at day 5, while the subsequent gradual increase in viral load from 8-10 dpf suggested viral replication in midges with disseminated infection. Overall, the data presented herein suggest that EHDV-2 disseminates via the hemolymph to secondary infection sites throughout the midge and demonstrate a high potential for transmission at five days at 25°C after an infective blood-meal.

  4. On the Seasonal Occurrence and Abundance of the Zika Virus Vector Mosquito Aedes Aegypti in the Contiguous United States.

    Science.gov (United States)

    Monaghan, Andrew J; Morin, Cory W; Steinhoff, Daniel F; Wilhelmi, Olga; Hayden, Mary; Quattrochi, Dale A; Reiskind, Michael; Lloyd, Alun L; Smith, Kirk; Schmidt, Chris A; Scalf, Paige E; Ernst, Kacey

    2016-03-16

    An ongoing Zika virus pandemic in Latin America and the Caribbean has raised concerns that travel-related introduction of Zika virus could initiate local transmission in the United States (U.S.) by its primary vector, the mosquito Aedes aegypti. We employed meteorologically driven models for 2006-2015 to simulate the potential seasonal abundance of adult Aedes aegypti for fifty cities within or near the margins of its known U.S. range. Mosquito abundance results were analyzed alongside travel and socioeconomic factors that are proxies of viral introduction and vulnerability to human-vector contact.     Meteorological conditions are largely unsuitable for Aedes aegypti over the U.S. during winter months (December-March), except in southern Florida and south Texas where comparatively warm conditions can sustain low-to-moderate potential mosquito abundance. Meteorological conditions are suitable for Aedes aegypti across all fifty cities during peak summer months (July-September), though the mosquito has not been documented in all cities. Simulations indicate the highest mosquito abundance occurs in the Southeast and south Texas where locally acquired cases of Aedes-transmitted viruses have been reported previously. Cities in southern Florida and south Texas are at the nexus of high seasonal suitability for Aedes aegypti and strong potential for travel-related virus introduction. Higher poverty rates in cities along the U.S.-Mexico border may correlate with factors that increase human exposure to Aedes aegypti.     Our results can inform baseline risk for local Zika virus transmission in the U.S. and the optimal timing of vector control activities, and underscore the need for enhanced surveillance for Aedes mosquitoes and Aedes-transmitted viruses.

  5. Evaluating the feeding preferences of West Nile virus mosquito vectors using bird-baited traps.

    Science.gov (United States)

    Victoriano Llopis, Isis; Tomassone, Laura; Grego, Elena; Serrano, Emmanuel; Mosca, Andrea; Vaschetti, Gabriella; Andrade, Daniela; Rossi, Luca

    2016-08-31

    The total contact rates (TCRs) between mosquito vectors and their potential hosts have a serious impact on disease transmission dynamics. Culex pipiens (sensu stricto) (s.s.) is considered the main vector of the West Nile Virus (WNV) in Europe and birds are the reservoir hosts. The results of our previous study showed that WNV seroreactors are significantly more prevalent among raptors compared to a range of other wild avian groups. The current study aims to assess the role of bird type (raptor vs others) and bird size on mosquito feeding preferences in a free-choice experiment using bird-baited traps. From July to September 2014, a battery of six bird-baited traps was operated in twelve mosquito capture sessions. Eight bird species, belonging to five different orders, including raptors, were used. After each session, the trapped mosquitoes were collected and identified using standard keys. Two sets of independent generalized linear mixed models (GLMM) were used to assess mosquito vector feeding preferences (MFp) among different bird species and types. A total of 304 mosquitoes belonging to seven taxa were collected, C. pipiens being by far the most abundant (84.2 % of the total mosquito catch). Most C. pipiens were engorged (83.59 %). The selected model showed that 25.6 % of the observed variability of MFp is explained by the interaction between bird size and bird type, with C. pipiens preferring to feed on large birds, especially raptors. The proportion of engorged mosquitoes was 1.9-fold higher in large (22.88 %; range 0-42 %) than in medium-sized raptors (11.71 %; range 0-33 %), and was nearly the same in medium-sized (9.08 %; range 0-26 %) and large (8.5 %; 6-24 %) non-raptor species. Culex pipiens showed an obvious preference for large raptors, which concurs with the higher seroprevalence to WNV in our previous study. The appreciable feeding by C. pipiens on large raptors makes them useful alternative sentinels to poultry for WNV surveillance. Thus

  6. A Semipersistent Plant Virus Differentially Manipulates Feeding Behaviors of Different Sexes and Biotypes of Its Whitefly Vector.

    Science.gov (United States)

    Lu, Shaohua; Li, Jingjing; Wang, Xueli; Song, Danyang; Bai, Rune; Shi, Yan; Gu, Qinsheng; Kuo, Yen-Wen; Falk, Bryce W; Yan, Fengming

    2017-01-13

    It is known that plant viruses can change the performance of their vectors. However, there have been no reports on whether or how a semipersistent plant virus manipulates the feeding behaviors of its whitefly vectors. Cucurbit chlorotic yellows virus (CCYV) (genus Crinivirus , family Closteroviridae ) is an emergent plant virus in many Asian countries and is transmitted specifically by B and Q biotypes of tobacco whitefly, Bemisia tabaci (Gennadius), in a semipersistent manner. In the present study, we used electrical penetration graph (EPG) technique to investigate the effect of CCYV on the feeding behaviors of B. tabaci . The results showed that CCYV altered feeding behaviors of both biotypes and sexes of B. tabaci with different degrees. CCYV had stronger effects on feeding behaviors of Q biotype than those of B biotype, by increasing duration of phloem salivation and sap ingestion, and could differentially manipulate feeding behaviors of males and females in both biotype whiteflies, with more phloem ingestion in Q biotype males and more non-phloem probing in B biotype males than their respective females. With regard to feeding behaviors related to virus transmission, these results indicated that, when carrying CCYV, B. tabaci Q biotype plays more roles than B biotype, and males make greater contribution than females.

  7. Hepatorenal correction in murine glycogen storage disease type I with a double-stranded adeno-associated virus vector.

    LENUS (Irish Health Repository)

    Luo, Xiaoyan

    2011-11-01

    Glycogen storage disease type Ia (GSD-Ia) is caused by the deficiency of glucose-6-phosphatase (G6Pase). Long-term complications of GSD-Ia include life-threatening hypoglycemia and proteinuria progressing to renal failure. A double-stranded (ds) adeno-associated virus serotype 2 (AAV2) vector encoding human G6Pase was pseudotyped with four serotypes, AAV2, AAV7, AAV8, and AAV9, and we evaluated efficacy in 12-day-old G6pase (-\\/-) mice. Hypoglycemia during fasting (plasma glucose <100 mg\\/dl) was prevented for >6 months by the dsAAV2\\/7, dsAAV2\\/8, and dsAAV2\\/9 vectors. Prolonged fasting for 8 hours revealed normalization of blood glucose following dsAAV2\\/9 vector administration at the higher dose. The glycogen content of kidney was reduced by >65% with both the dsAAV2\\/7 and dsAAV2\\/9 vectors, and renal glycogen content was stably reduced between 7 and 12 months of age for the dsAAV2\\/9 vector-treated mice. Every vector-treated group had significantly reduced glycogen content in the liver, in comparison with untreated G6pase (-\\/-) mice. G6Pase was expressed in many renal epithelial cells of with the dsAAV2\\/9 vector for up to 12 months. Albuminuria and renal fibrosis were reduced by the dsAAV2\\/9 vector. Hepatorenal correction in G6pase (-\\/-) mice demonstrates the potential of AAV vectors for the correction of inherited diseases of metabolism.

  8. Declining malaria, rising of dengue and Zika virus: insights for mosquito vector control.

    Science.gov (United States)

    Benelli, Giovanni; Mehlhorn, Heinz

    2016-05-01

    The fight against mosquito-borne diseases is a challenge of huge public health importance. To our mind, 2015 was an extraordinary year for malaria control, due to three hot news: the Nobel Prize to Youyou Tu for the discovery of artemisinin, the development of the first vaccine against Plasmodium falciparum malaria [i.e. RTS,S/AS01 (RTS,S)], and the fall of malaria infection rates worldwide, with special reference to sub-Saharan Africa. However, there are major challenges that still deserve attention, in order to boost malaria prevention and control. Indeed, parasite strains resistant to artemisinin have been detected, and RTS,S vaccine does not offer protection against Plasmodium vivax malaria, which predominates in many countries outside of Africa. Furthermore, the recent outbreaks of Zika virus infections, occurring in South America, Central America and the Caribbean, represent the most recent of four arrivals of important arboviruses in the Western Hemisphere, over the last 20 years. Zika virus follows dengue (which slyly arrived in the hemisphere over decades and became more aggressive in the 1990s), West Nile virus (emerged in 1999) and chikungunya (emerged in 2013). Notably, there are no specific treatments for these arboviruses. The emerging scenario highlights that the effective and eco-friendly control of mosquito vectors, with special reference to highly invasive species such as Aedes aegypti and Aedes albopictus, is crucial. The concrete potential of screening plant species as sources of metabolites for parasitological purposes is worthy of attention, as elucidated by the Y. Tu's example. Notably, plant-borne molecules are often effective at few parts per million against Aedes, Ochlerotatus, Anopheles and Culex young instars, can be used for the rapid synthesis of mosquitocidal nanoformulations and even employed to prepare cheap repellents with low human toxicity. In addition, behaviour-based control tools relying to the employ of sound traps and the

  9. An efficient deletion mutant packaging system for defective herpes simplex virus vectors: Potential applications to human gene therapy and neuronal physiology

    International Nuclear Information System (INIS)

    Geller, A.I.; Keyomarsi, K.; Bryan, J.; Pardee, A.B.

    1990-01-01

    The authors have previously described a defective herpes simplex virus (HSV-1) vector system that permits that introduction of virtually any gene into nonmitotic cells. pHSVlac, the prototype vector, stably expresses Escherichia coli β-galactosidase from a constitutive promoter in many human cell lines, in cultured rat neurons from throughout the nervous system, and in cells in the adult rat brain. HSV-1 vectors expressing other genes may prove useful for studying neuronal physiology or performing human gene therapy for neurological diseases, such as Parkinson disease or brain tumors. A HSV-1 temperature-sensitive (ts) mutant, ts K, has been used as helper virus; ts mutants revert to wild type. In contrast, HSV-1 deletion mutants essentially cannot revert to wild type; therefore, use of a deletion mutant as helper virus might permit human gene therapy with HSV-1 vectors. They now report an efficient packaging system for HSV-1 VECTORS USING A DELETION MUTANT, d30EBA, as helper virus; virus is grown on the complementing cell line M64A. pHSVlac virus prepared using the deletion mutant packaging system stably expresses β-galactosidase in cultured rat sympathetic neurons and glia. Both D30EBA and ts K contain a mutation in the IE3 gene of HSV-1 strain 17 and have the same phenotype; therefore, changing the helper virus from ts K to D30EBA does not alter the host range or other properties of the HSV-1 vector system

  10. A Respiratory Syncytial Virus Vaccine Vectored by a Stable Chimeric and Replication-Deficient Sendai Virus Protects Mice without Inducing Enhanced Disease.

    Science.gov (United States)

    Wiegand, Marian Alexander; Gori-Savellini, Gianni; Gandolfo, Claudia; Papa, Guido; Kaufmann, Christine; Felder, Eva; Ginori, Alessandro; Disanto, Maria Giulia; Spina, Donatella; Cusi, Maria Grazia

    2017-05-15

    Respiratory syncytial virus (RSV) is a major cause of severe respiratory infections in children and elderly people, and no marketed vaccine exists. In this study, we generated and analyzed a subunit vaccine against RSV based on a novel genome replication-deficient Sendai virus (SeV) vector. We inserted the RSV F protein, known to be a genetically stable antigen, into our vector in a specific way to optimize the vaccine features. By exchanging the ectodomain of the SeV F protein for its counterpart from RSV, we created a chimeric vectored vaccine that contains the RSV F protein as an essential structural component. In this way, the antigen is actively expressed on the surfaces of vaccine particles in its prefusion conformation, and as recently reported for other vectored vaccines, the occurrence of silencing mutations of the transgene in the vaccine genome can be prevented. In addition, its active gene expression contributes to further stimulation of the immune response. In order to understand the best route of immunization, we compared vaccine efficacies after intranasal (i.n.) or intramuscular (i.m.) immunization of BALB/c mice. Via both routes, substantial RSV-specific immune responses were induced, consisting of serum IgG and neutralizing antibodies, as well as cytotoxic T cells. Moreover, i.n. immunization was also able to stimulate specific mucosal IgA in the upper and lower respiratory tract. In virus challenge experiments, animals were protected against RSV infection after both i.n. and i.m. immunization without inducing vaccine-enhanced disease. Above all, the replication-deficient SeV appeared to be safe and well tolerated. IMPORTANCE Respiratory syncytial virus (RSV) is a major cause of respiratory diseases in young children and elderly people worldwide. There is a great demand for a licensed vaccine. Promising existing vaccine approaches based on live-attenuated vaccines or viral vectors have suffered from unforeseen drawbacks related to immunogenicity

  11. Recombinant Adeno-Associated Virus Vector Genomes Take the Form of Long-Lived, Transcriptionally Competent Episomes in Human Muscle

    Science.gov (United States)

    Schnepp, Bruce C.; Chulay, Jeffrey D.; Ye, Guo-Jie; Flotte, Terence R.; Trapnell, Bruce C.; Johnson, Philip R.

    2016-01-01

    Gene augmentation therapy as a strategy to treat alpha-1 antitrypsin (AAT) deficiency has reached phase 2 clinical testing in humans. Sustained serum levels of AAT have been observed beyond one year after intramuscular administration of a recombinant adeno-associated virus (rAAV) vector expressing the AAT gene. In this study, sequential muscle biopsies obtained at 3 and 12 months after vector injection were examined for the presence of rAAV vector genomes. Each biopsy sample contained readily detectable vector DNA, the majority of which existed as double-stranded supercoiled and open circular episomes. Episomes persisted through 12 months, although at slightly lower levels than observed at 3 months. There was a clear dose response when comparing the low- and mid-vector-dose groups to the high-dose group. The highest absolute copy numbers were found in a high-dose subject, and serum AAT levels at 12 months confirmed that the high-dose group also had the highest sustained serum AAT levels. Sequence analysis revealed that the vast majority of episomes contained double-D inverted terminal repeats ranging from fully intact to severely deleted. Molecular clones of vector genomes derived directly from the biopsies were transcriptionally active, potentially identifying them as the source of serum AAT in the trial subjects. PMID:26650966

  12. Inter-epidemic abundance and distribution of potential mosquito vectors for Rift Valley fever virus in Ngorongoro district, Tanzania

    OpenAIRE

    Mweya, Clement N.; Kimera, Sharadhuli I.; Mellau, Lesakit S. B.; Mboera, Leonard E. G.

    2015-01-01

    Background: Rift Valley fever (RVF) is a mosquito-borne viral zoonosis that primarily affects ruminants but also has the capacity to infect humans. Objective: To determine the abundance and distribution of mosquito vectors in relation to their potential role in the virus transmission and maintenance in disease epidemic areas of Ngorongoro district in northern Tanzania. Methods: A cross-sectional entomological investigation was carried out before the suspected RVF outbreak in October 2012. Mos...

  13. Culicoides (Diptera: Ceratopogonidae) midges, the vectors of African horse sickness virus--a host/vector contact study in the Niayes area of Senegal.

    Science.gov (United States)

    Fall, Moussa; Diarra, Maryam; Fall, Assane G; Balenghien, Thomas; Seck, Momar T; Bouyer, Jérémy; Garros, Claire; Gimonneau, Geoffrey; Allène, Xavier; Mall, Iba; Delécolle, Jean-Claude; Rakotoarivony, Ignace; Bakhoum, Mame T; Dusom, Ange M; Ndao, Massouka; Konaté, Lassana; Faye, Ousmane; Baldet, Thierry

    2015-01-21

    African horse sickness (AHS) is an equine disease endemic to Senegal. The African horse sickness virus (AHSV) is transmitted to the mammalian hosts by midges of the Culicoides Latreille genus. During the last epizootic outbreak of AHS in Senegal in 2007, 1,169 horses died from this disease entailing an estimated cost of 1.4 million euros. In spite of the serious animal health and economic implications of AHS, very little is known about determinants involved in transmission such as contact between horses and the Culicoides species suspected of being its vectors. The monthly variation in host/vector contact was determined in the Niayes area, Senegal, an area which was severely affected by the 2007 outbreak of AHS. A horse-baited trap and two suction light traps (OVI type) were set up at each of five sites for three consecutive nights every month for one year. Of 254,338 Culicoides midges collected 209,543 (82.4%) were female and 44,795 (17.6%) male. Nineteen of the 41 species collected were new distribution records for Senegal. This increased the number of described Culicoides species found in Senegal to 53. Only 19 species, of the 41 species found in light trap, were collected in the horse-baited trap (23,669 specimens) largely dominated by Culicoides oxystoma (22,300 specimens, i.e. 94.2%) followed by Culicoides imicola (482 specimens, i.e. 2.0%) and Culicoides kingi (446 specimens, i.e. 1.9%). Culicoides oxystoma should be considered as a potential vector of AHSV in the Niayes area of Senegal due to its abundance on horses and its role in the transmission of other Culicoides-borne viruses.

  14. Assessment of Local Mosquito Species Incriminates Aedes aegypti as the Potential Vector of Zika Virus in Australia.

    Directory of Open Access Journals (Sweden)

    Sonja Hall-Mendelin

    2016-09-01

    Full Text Available Within the last 10 years Zika virus (ZIKV has caused unprecedented epidemics of human disease in the nations and territories of the western Pacific and South America, and continues to escalate in both endemic and non-endemic regions. We evaluated the vector competence of Australian mosquitoes for ZIKV to assess their potential role in virus transmission.Mosquitoes were exposed to infectious blood meals containing the prototype African ZIKV strain. After 14 days incubation at 28°C and high relative humidity, infection, dissemination and transmission rates were assessed. Infection in Culex annulirostris and Cx. sitiens could not be detected. 8% of Cx. quinquefasciatus were infected, but the virus did not disseminate in this species. Despite having infection rates > 50%, Aedes notoscriptus and Ae. vigilax did not transmit ZIKV. In contrast, Ae. aegypti had infection and transmission rates of 57% and 27%, respectively. In susceptibility trials, the virus dose required to infect 50% (ID50 of Ae. aegypti was106.4 tissue culture infectious dose50 (TCID50/mL. Additionally, a threshold viral load within the mosquito of at least 105.1 TCID50 equivalents/mL had to be reached before virus transmission occurred.We confirmed Ae. aegypti to be the most likely mosquito vector of ZIKV in Australia, although the restricted distribution of this species will limit the receptive zone to northern Queensland where this species occurs. Importantly, the role in ZIKV transmission of Culex and other Aedes spp. tested will be negligible. Despite being the implicated vector, the relatively high ID50 and need for a high titer disseminated infection in Ae. aegypti suggest that high mosquito population densities will be required to facilitate epidemic ZIKV transmission among the currently immunologically naïve human population in Australia.

  15. Migration of rice planthoppers and their vectored re-emerging and novel rice viruses in East Asia

    Directory of Open Access Journals (Sweden)

    Akira eOtuka

    2013-10-01

    Full Text Available This review examines recent studies of the migration of three rice planthoppers, Laodelphax striatellus, Sogatella furcifera, and Nilaparvata lugens, in East Asia. Laodelphax striatellus has recently broken out in Jiangsu province, eastern China. The population density in the province started to increase in the early 2000s and peaked in 2004. In 2005, Rice stripe virus (RSV viruliferous rate of L. striatellus peaked at 31.3%. Since then, rice stripe disease spread severely across the whole province. Due to the migration of the RSV vectors, the rice stripe disease spread to neighboring countries Japan and Korea. An overseas migration of L. striatellus that occurred in 2008 was analyzed, when a slow-moving cold vortex, a type of low pressure system, reached western Japan from Jiangsu, carrying the insects into Japan. Subsequently the rice stripe diseases struck these areas in Japan severely. In Korea, similar situations occurred in 2009, 2011, and 2012. Their migration sources were also estimated to be in Jiangsu by backward trajectory analysis. Rice black-streaked dwarf virus, whose vector is L. striatellus, has recently re-emerged in eastern China, and the evidence for overseas migrations of the virus, just like the RSV’s migrations, has been given. A method of predicting the overseas migration of L. striatellus has been developed by Japanese, Chinese, and Korean institutes. An evaluation of the prediction showed that this method properly predicted migration events that occurred in East Asia from 2008 to 2011. Southern rice black-streaked dwarf virus (SRBSDV was first found in Guangdong province. Its vector is S. furcifera. An outbreak of SRBSDV occurred in southern China in 2009 and spread to Vietnam the same year. This disease and virus were also found in Japan in 2010. The epidemic triggered many migration studies to investigate concrete spring-summer migration routes in China, and the addition of migration sources for early arrivals in

  16. Characterization of rice black-streaked dwarf virus- and rice stripe virus-derived siRNAs in singly and doubly infected insect vector Laodelphax striatellus.

    Directory of Open Access Journals (Sweden)

    Junmin Li

    Full Text Available Replication of RNA viruses in insect cells triggers an antiviral defense that is mediated by RNA interference (RNAi which generates viral-derived small interfering RNAs (siRNAs. However, it is not known whether an antiviral RNAi response is also induced in insects by reoviruses, whose double-stranded RNA genome replication is thought to occur within core particles. Deep sequencing of small RNAs showed that when the small brown planthopper (Laodelphax striatellus was infected by Rice black-streaked dwarf virus (RBSDV (Reoviridae; Fijivirus, more viral-derived siRNAs accumulated than when the vector insect was infected by Rice stripe virus (RSV, a negative single-stranded RNA virus. RBSDV siRNAs were predominantly 21 and 22 nucleotides long and there were almost equal numbers of positive and negative sense. RBSDV siRNAs were frequently generated from hotspots in the 5'- and 3'-terminal regions of viral genome segments but these hotspots were not associated with any predicted RNA secondary structures. Under laboratory condition, L. striatellus can be infected simultaneously with RBSDV and RSV. Double infection enhanced the accumulation of particular genome segments but not viral coat protein of RBSDV and correlated with an increase in the abundance of siRNAs derived from RBSDV. The results of this study suggest that reovirus replication in its insect vector potentially induces an RNAi-mediated antiviral response.

  17. Characterization of rice black-streaked dwarf virus- and rice stripe virus-derived siRNAs in singly and doubly infected insect vector Laodelphax striatellus.

    Science.gov (United States)

    Li, Junmin; Andika, Ida Bagus; Shen, Jiangfeng; Lv, Yuanda; Ji, Yongqiang; Sun, Liying; Chen, Jianping

    2013-01-01

    Replication of RNA viruses in insect cells triggers an antiviral defense that is mediated by RNA interference (RNAi) which generates viral-derived small interfering RNAs (siRNAs). However, it is not known whether an antiviral RNAi response is also induced in insects by reoviruses, whose double-stranded RNA genome replication is thought to occur within core particles. Deep sequencing of small RNAs showed that when the small brown planthopper (Laodelphax striatellus) was infected by Rice black-streaked dwarf virus (RBSDV) (Reoviridae; Fijivirus), more viral-derived siRNAs accumulated than when the vector insect was infected by Rice stripe virus (RSV), a negative single-stranded RNA virus. RBSDV siRNAs were predominantly 21 and 22 nucleotides long and there were almost equal numbers of positive and negative sense. RBSDV siRNAs were frequently generated from hotspots in the 5'- and 3'-terminal regions of viral genome segments but these hotspots were not associated with any predicted RNA secondary structures. Under laboratory condition, L. striatellus can be infected simultaneously with RBSDV and RSV. Double infection enhanced the accumulation of particular genome segments but not viral coat protein of RBSDV and correlated with an increase in the abundance of siRNAs derived from RBSDV. The results of this study suggest that reovirus replication in its insect vector potentially induces an RNAi-mediated antiviral response.

  18. Vector competence of Culex neavei and Culex quinquefasciatus (Diptera: Culicidae) from Senegal for lineages 1, 2, Koutango and a putative new lineage of West Nile virus.

    Science.gov (United States)

    Fall, Gamou; Diallo, Mawlouth; Loucoubar, Cheikh; Faye, Ousmane; Sall, Amadou Alpha

    2014-04-01

    West Nile virus (WN virus) is one of the most widespread arbovirus and exhibits a great genetic diversity with 8 lineages, at least 4 (1, 2, Koutango, and putative new) are present in Africa. In West Africa, Culex neavei and Culex quinquefasciatus are considered as potential vectors for WN virus transmission in sylvatic or urban context. We analyzed the vector competence of these Culex species from Senegal for African lineages and envelope proteins sequences of viral strains used. We showed that lineage 1 is transmitted by both Culex mosquitoes, whereas the putative new lineage 8 is transmitted only by Cx. neavei. Our findings suggest that genetic variability can affect vector competence and depend on mosquito. However, when considering the infective life rate, the mosquito population seems to be inefficient for WN virus transmission in the field and could explain the low impact of WN virus in Africa.

  19. Recombinant Newcastle disease viral vector expressing hemagglutinin or fusion of canine distemper virus is safe and immunogenic in minks.

    Science.gov (United States)

    Ge, Jinying; Wang, Xijun; Tian, Meijie; Gao, Yuwei; Wen, Zhiyuan; Yu, Guimei; Zhou, Weiwei; Zu, Shulong; Bu, Zhigao

    2015-05-15

    Canine Distemper Virus (CDV) infects many carnivores and cause several high-mortality disease outbreaks. The current CDV live vaccine cannot be safely used in some exotic species, such as mink and ferret. Here, we generated recombinant lentogenic Newcastle disease virus (NDV) LaSota expressing either envelope glycoproyein, heamagglutinine (H) or fusion protein (F), named as rLa-CDVH and rLa-CDVF, respectively. The feasibility of these recombinant NDVs to serve as live virus-vectored CD vaccine was evaluated in minks. rLa-CDVH induced significant neutralization antibodies (NA) to CDV and provided solid protection against virulent CDV challenge. On the contrast, rLa-CDVF induced much lower NA to CDV and fail to protected mink from virulent CDV challenge. Results suggest that recombinant NDV expressing CDV H is safe and efficient candidate vaccine against CDV in mink, and maybe other host species. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Simulating spread of Bluetongue Virus by flying vectors between hosts on pasture

    DEFF Research Database (Denmark)

    Græsbøll, Kaare; Bødker, Rene; Enøe, Claes

    2012-01-01

    and display search behavior to locate areas with hosts. We also include wind spread of vectors, host movements, and vector seasonality. Results show that temperature and seasonality of vectors determines the period in which an incursion of Bluetongue may lead to epidemic spread in Denmark. Within this period...

  1. Vector choice determines immunogenicity and potency of genetic vaccines against Angola Marburg virus in nonhuman primates

    NARCIS (Netherlands)

    Geisbert, Thomas W.; Bailey, Michael; Geisbert, Joan B.; Asiedu, Clement; Roederer, Mario; Grazia-Pau, Maria; Custers, Jerome; Jahrling, Peter; Goudsmit, Jaap; Koup, Richard; Sullivan, Nancy J.

    2010-01-01

    The immunogenicity and durability of genetic vaccines are influenced by the composition of gene inserts and choice of delivery vector. DNA vectors are a promising vaccine approach showing efficacy when combined in prime-boost regimens with recombinant protein or viral vectors, but they have shown

  2. Vector competence of the stable fly (Diptera: Muscidae) for West Nile virus.

    Science.gov (United States)

    Doyle, Michael S; Swope, Bethany N; Hogsette, Jerome A; Burkhalter, Kristen L; Savage, Harry M; Nasci, Roger S

    2011-05-01

    In 2006-2007, stable flies, Stomoxys calcitrans (L.) (Diptera: Muscidae), were suspected of being enzootic vectors of West Nile virus (family Flaviviridae, genus Flavivirus, WNV) during a die-off of American white pelicans (Pelecanus erythrorhynchos Gmelin) (Pelecanidae) in Montana, USA. WNV-positive stable flies were observed feeding en masse on incapacitated, WNV-positive pelicans, arousing suspicions that the flies could have been involved in WNV transmission among pelicans, and perhaps to livestock and humans. We assessed biological transmission by infecting stable flies intrathoracically with WNV and testing them at 2-d intervals over 20 d. Infectious WNV was detected in fly bodies in decreasing amounts over time for only the first 6 d postinfection, an indication that WNV did not replicate within fly tissues and that stable flies cannot biologically transmit WNV. We assessed mechanical transmission using a novel technique. Specifically, we fed WNV-infected blood to individual flies by using a cotton swab (i.e., artificial donor), and at intervals of 1 min-24 h, we allowed flies to refeed on a different swab saturated with WNV-negative blood (i.e., artificial recipient). Flies mechanically transmitted viable WNV from donor to recipient swabs for up to 6 h postinfection, with the majority of the transmission events occurring within the first hour. Flies mechanically transmitted WNV RNA to recipient swabs for up to 24 h, mostly within the first 6 h. Given its predilection to feed multiple times when disturbed, these findings support the possibility that the stable fly could mechanically transmit WNV.

  3. Herpes simplex virus type 1-based amplicon vectors for fundamental research in neurosciences and gene therapy of neurological diseases.

    Science.gov (United States)

    Jerusalinsky, Diana; Baez, María Verónica; Epstein, Alberto Luis

    2012-01-01

    Somatic manipulation of the nervous system without the involvement of the germinal line appears as a powerful counterpart of the transgenic strategy. The use of viral vectors to produce specific, transient and localized knockout, knockdown, ectopic expression or overexpression of a gene, leads to the possibility of analyzing both in vitro and in vivo molecular basis of neural function. In this approach, viral particles engineered to carry transgenic sequences are delivered into discrete brain regions, to transduce cells that will express the transgenic products. Amplicons are replication-incompetent helper-dependent vectors derived from herpes simplex virus type 1 (HSV-1), with several advantages that potentiate their use in neurosciences: (1) minimal toxicity: amplicons do not encode any virus proteins, are neither toxic for the infected cells nor pathogenic for the inoculated animals and elicit low levels of adaptive immune responses; (2) extensive transgene capacity to carry up to 150-kb of foreign DNA; i.e., entire genes with regulatory sequences could be delivered; (3) widespread cellular tropism: amplicons can experimentally infect several cell types including glial cells, though naturally the virus infects mainly neurons and epithelial cells; (4) since the viral genome does not integrate into cellular chromosomes there is low probability to induce insertional mutagenesis. Recent investigations on gene transfer into the brain using these vectors, have focused on gene therapy of inherited genetic diseases affecting the nervous system, such as ataxias, or on neurodegenerative disorders using experimental models of Parkinson's or Alzheimer's disease. Another group of studies used amplicons to investigate complex neural functions such as neuroplasticity, anxiety, learning and memory. In this short review, we summarize recent data supporting the potential of HSV-1 based amplicon vector model for gene delivery and modulation of gene expression in primary cultures

  4. Sustained secretion of human alpha-1-antitrypsin from murine muscle transduced with adeno-associated virus vectors

    Science.gov (United States)

    Song, Sihong; Morgan, Michael; Ellis, Tamir; Poirier, Amy; Chesnut, Kye; Wang, Jianming; Brantly, Mark; Muzyczka, Nicholas; Byrne, Barry J.; Atkinson, Mark; Flotte, Terence R.

    1998-01-01

    Recombinant adeno-associated virus (AAV) vectors have been used to transduce murine skeletal muscle as a platform for secretion of therapeutic proteins. The utility of this approach for treating alpha-1-antitrypsin (AAT) deficiency was tested in murine myocytes in vitro and in vivo. AAV vectors expressing the human AAT gene from either the cytomegalovirus (CMV) promoter (AAV-C-AT) or the human elongation factor 1-α promoter (AAV-E-AT) were examined. In vitro in C2C12 murine myoblasts, the expression levels in transient transfections were similar between the two vectors. One month after transduction, however, the human elongation factor 1 promoter mediated 10-fold higher stable human AAT expression than the CMV promoter. In vivo transduction was performed by injecting doses of up to 1.4 × 1013 particles into skeletal muscles of several mouse strains (C57BL/6, BALB/c, and SCID). In vivo, the CMV vector mediated higher levels of expression, with sustained serum levels over 800 μg/ml in SCID and over 400 μg/ml in C57BL/6 mice. These serum concentrations are 100,000-fold higher than those previously observed with AAV vectors in muscle and are at levels which would be therapeutic if achieved in humans. High level expression was delayed for several weeks but was sustained for over 15 wk. Immune responses were dependent upon the mouse strain and the vector dosage. These data suggest that recombinant AAV vector transduction of skeletal muscle could provide a means for replacing AAT or other essential serum proteins but that immune responses may be elicited under certain conditions. PMID:9826709

  5. Adeno-associated Virus Vectors Efficiently Transduce Mouse and Rabbit Sensory Neurons Coinfected with Herpes Simplex Virus 1 following Peripheral Inoculation.

    Science.gov (United States)

    Watson, Zachary L; Ertel, Monica K; Lewin, Alfred S; Tuli, Sonal S; Schultz, Gregory S; Neumann, Donna M; Bloom, David C

    2016-09-01

    Following infection of epithelial tissues, herpes simplex virus 1 (HSV-1) virions travel via axonal transport to sensory ganglia and establish a lifelong latent infection within neurons. Recent studies have revealed that, following intraganglionic or intrathecal injection, recombinant adeno-associated virus (rAAV) vectors can also infect sensory neurons and are capable of stable, long-term transgene expression. We sought to determine if application of rAAV to peripheral nerve termini at the epithelial surface would allow rAAV to traffic to sensory ganglia in a manner similar to that seen with HSV. We hypothesized that footpad or ocular inoculation with rAAV8 would result in transduction of dorsal root ganglia (DRG) or trigeminal ganglia (TG), respectively. To test this, we inoculated the footpads of mice with various amounts of rAAV as well as rAAV capsid mutants. We demonstrated that this method of inoculation can achieve a transduction rate of >90% of the sensory neurons in the DRG that innervate the footpad. Similarly, we showed that corneal inoculation with rAAV vectors in the rabbit efficiently transduced >70% of the TG neurons in the optic tract. Finally, we demonstrated that coinfection of mouse footpads or rabbit eyes with rAAV vectors and HSV-1 resulted in colocalization in nearly all of the HSV-1-positive neurons. These results suggest that rAAV is a useful tool for the study of HSV-1 infection and may provide a means to deliver therapeutic cargos for the treatment of HSV infections or of dysfunctions of sensory ganglia. Adeno-associated virus (AAV) has been shown to transduce dorsal root ganglion sensory neurons following direct intraganglionic sciatic nerve injection and intraperitoneal and intravenous injection as well as intrathecal injection. We sought to determine if rAAV vectors would be delivered to the same sensory neurons that herpes simplex virus (HSV-1) infects when applied peripherally at an epithelial surface that had been treated to expose

  6. Virus-independent and common transcriptome responses of leafhopper vectors feeding on maize infected with semi-persistently and persistent propagatively transmitted viruses.

    Science.gov (United States)

    Cassone, Bryan J; Wijeratne, Saranga; Michel, Andrew P; Stewart, Lucy R; Chen, Yuting; Yan, Pearlly; Redinbaugh, Margaret G

    2014-02-14

    innate immune system in G. nigrifrons fed on a foregut-borne virus (MCDV) that does not infect leafhoppers was less expected. The changes in transcript accumulation that occur independent of the mode of pathogen transmission could be key for identifying insect factors that disrupt vector-mediated plant virus transmission.

  7. Implicated vectors and spread of grapevine red blotch-associated virus in Oregon vineyards

    Science.gov (United States)

    Grapevine viruses have detrimental consequences for wine grape production, as is known for Grapevine leafroll -associated viruses (GLRaVs) and Grapevine red blotch -associated virus (GRBaV). From 2013-2016, vineyards in three wine grape production regions of Oregon were surveyed for the presence of ...

  8. Thrips developmental stage-specific transcriptome response to tomato spotted wilt virus during the virus infection cycle in Frankliniella occidentalis, the primary vector.

    Science.gov (United States)

    Schneweis, Derek J; Whitfield, Anna E; Rotenberg, Dorith

    2017-01-01

    Tomato spotted wilt virus (TSWV) is transmitted by Frankliniella occidentalis in a circulative-propagative manner. Little is known about thrips vector response to TSWV during the infection process from larval acquisition to adult inoculation of plants. Whole-body transcriptome response to virus infection was determined for first-instar larval, pre-pupal and adult thrips using RNA-Seq. TSWV responsive genes were identified using preliminary sequence of a draft genome of F. occidentalis as a reference and three developmental-stage transcriptomes were assembled. Processes and functions associated with host defense, insect cuticle structure and development, metabolism and transport were perturbed by TSWV infection as inferred by ontologies of responsive genes. The repertoire of genes responsive to TSWV varied between developmental stages, possibly reflecting the link between thrips development and the virus dissemination route in the vector. This study provides the foundation for exploration of tissue-specific expression in response to TSWV and functional analysis of thrips gene function. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Current status of the Citrus leprosis virus (CiLV -C and its vector Brevipalpus phoenicis (Geijskes

    Directory of Open Access Journals (Sweden)

    Guillermo León M

    2012-08-01

    Full Text Available The Citrus leprosis virus CiLV-C is a quarantine disease of economic importance. Over the past 15 years, this disease has spread to several countries of Central and South America. Colombia has about 45,000 hectares of citrus planted with an annual production of 750,000 tonnes. The CiLV-C has only been detected in the departments of Meta, Casanare and recently Tolima. Meta has 4,300 hectares representing 10% of the national cultivated area, and Casanare, where CiLV-C appeared in 2004, has no more than 500 ha planted with citrus. The presence of the Citrus leprosis virus in Colombia could affect the international market for citrus, other crops and ornamental plants with the United States and other countries without the disease. The false spider mite Brevipalpus phoenicis (Geijskes (Acari: Tenuipalpidae is the main vector of the CiLV-C. Disease management is based on control programs of the vector and diminishing host plants. Chemical mite control is expensive, wasteful and generates resistance to different acaricides. This paper provides basic information on CiLV-C and its vector, advances in diagnosis and methods to control the disease and prevention of its spread

  10. Neonatal gene therapy of glycogen storage disease type Ia using a feline immunodeficiency virus-based vector.

    Science.gov (United States)

    Grinshpun, Albert; Condiotti, Reba; Waddington, Simon N; Peer, Michael; Zeig, Eli; Peretz, Sima; Simerzin, Alina; Chou, Janice; Pann, Chi-Jiunn; Giladi, Hilla; Galun, Eithan

    2010-09-01

    Glycogen storage disease type Ia (GSD-Ia), also known as von Gierke disease, is caused by a deficiency of glucose-6-phosphatase-alpha (G6Pase), a key enzyme in glucose homeostasis. From birth, affected individuals cannot maintain normal blood glucose levels and suffer from a variety of metabolic disorders, leading to life-threatening complications. Gene therapy has been proposed as a possible option for treatment of this illness. Vectors have been constructed from feline immunodeficiency virus (FIV), a nonprimate lentivirus, because the wild-type virus does not cause disease in humans. Previously, we have shown that these vectors are capable of integrating stably into hepatocyte cell lines and adult murine livers and lead to long-term transgene expression. In the current work, we have assessed the ability to attenuate disease symptoms in a murine model of GSD-Ia. Single administration of FIV vectors containing the human G6Pase gene to G6Pase-alpha(-/-) mice did not change the biochemical and pathological phenotype. However, a double neonatal administration protocol led to normalized blood glucose levels, significantly extended survival, improved body weight, and decreased accumulation of liver glycogen associated with the disease. This approach shows a promising paradigm for treating GSD-Ia patients early in life thereby avoiding long-term consequences.

  11. Identification of potential vectors of and detection of antibodies against Rift Valley fever virus in livestock during interepizootic periods.

    Science.gov (United States)

    Rostal, Melinda K; Evans, Alina L; Sang, Rosemary; Gikundi, Solomon; Wakhule, Lilian; Munyua, Peninah; Macharia, Joseph; Feikin, Daniel R; Breiman, Robert F; Njenga, M Kariuki

    2010-05-01

    To evaluate the prevalence of Rift Valley fever virus (RVFV) antibodies in livestock and presence of competent mosquito vectors of RVFV during an interepizootic period (IEP) in Kenya. 208 sheep and 84 goats ranging in age from 4 months to 15 years, from 2 breeding herds. Blood specimens were collected from the sheep and goats during the 1999-2006 IEP in Rift Valley Province, and serum was harvested. Serum specimens were tested for IgG and IgM antibodies against RVFV by use of an ELISA. In addition, 7,134 mosquitoes were trapped in Naivasha, Nairobi, and Northeastern Province, and speciation was performed. No animals were seropositive for IgM against RVFV. Of the animals born after the 1997-1998 epizootic, 18% (34/188) of sheep were seropositive for IgG against RVFV, compared with 3% (2/75) of goats. Seventy percent (8,144/11,678) of the mosquitoes collected were of the Culex subgenera; 18% (2,102/11,678) were Aedes spp. Detection of IgG in the sera of sheep and goats born after the 1997-1998 epizootic and before the 2006 epizootic indicated that virus activity existed during the IEP. Detection of Aedes mosquitoes, which are competent vectors of RVFV, suggested that a cryptic vector-to-vertebrate cycle may exist during IEPs.

  12. Mutational library analysis of selected amino acids in the receptor binding domain of envelope of Akv murine leukemia virus by conditionally replication competent bicistronic vectors

    DEFF Research Database (Denmark)

    Bahrami, Shervin; Jespersen, Thomas; Pedersen, Finn Skou

    2003-01-01

    envelope expression. This vector functions as a replication competent mini-virus in a culture of NIH 3T3 derived semi-packaging cells that express the viral Gag and Pol proteins. Titers comparable to those of wild type virus were achieved by this system. To test this vector system, we created a random...... mutational library of Arg 85 and Asp 86 in the first variable region of Akv envelope protein. Homologous amino acids to Asp 86 in Moloney and Friend murine leukemia viruses are thought to be directly involved in receptor binding. Subsequent selection of mutants capable of infecting murine NIH 3T3 cells...

  13. Properties of a herpes simplex virus multiple immediate-early gene-deleted recombinant as a vaccine vector

    International Nuclear Information System (INIS)

    Watanabe, Daisuke; Brockman, Mark A.; Ndung'u, Thumbi; Mathews, Lydia; Lucas, William T.; Murphy, Cynthia G.; Felber, Barbara K.; Pavlakis, George N.; Deluca, Neal A.; Knipe, David M.

    2007-01-01

    Herpes simplex virus (HSV) recombinants induce durable immune responses in rhesus macaques and mice and have induced partial protection in rhesus macaques against mucosal challenge with virulent simian immunodeficiency virus (SIV). In this study, we evaluated the properties of a new generation HSV vaccine vector, an HSV-1 multiple immediate-early (IE) gene deletion mutant virus, d106, which contains deletions in the ICP4, ICP27, ICP22, and ICP47 genes. Because several of the HSV IE genes have been implicated in immune evasion, inactivation of the genes encoding these proteins was expected to result in enhanced immunogenicity. The d106 virus expresses few HSV gene products and shows minimal cytopathic effect in cultured cells. When d106 was inoculated into mice, viral DNA accumulated at high levels in draining lymph nodes, consistent with an ability to transduce dendritic cells and activate their maturation and movement to lymph nodes. A d106 recombinant expressing Escherichia coli β-galactosidase induced durable β-gal-specific IgG and CD8 + T cell responses in naive and HSV-immune mice. Finally, d106-based recombinants have been constructed that express simian immunodeficiency virus (SIV) gag, env, or a rev-tat-nef fusion protein for several days in cultured cells. Thus, d106 shows many of the properties desirable in a vaccine vector: limited expression of HSV gene products and cytopathogenicity, high level expression of transgenes, ability to induce durable immune responses, and an ability to transduce dendritic cells and induce their maturation and migration to lymph nodes

  14. An adenovirus vectored mucosal adjuvant augments protection of mice immunized intranasally with an adenovirus-vectored foot-and-mouth disease virus subunit vaccine.

    Science.gov (United States)

    Alejo, Diana M; Moraes, Mauro P; Liao, Xiaofen; Dias, Camila C; Tulman, Edan R; Diaz-San Segundo, Fayna; Rood, Debra; Grubman, Marvin J; Silbart, Lawrence K

    2013-04-26

    Foot-and-mouth disease virus (FMDV) is a highly contagious pathogen that causes severe morbidity and economic losses to the livestock industry in many countries. The oral and respiratory mucosae are the main ports of entry of FMDV, so the stimulation of local immunity in these tissues may help prevent initial infection and viral spread. E. coli heat-labile enterotoxin (LT) has been described as one of the few molecules that have adjuvant activity at mucosal surfaces. The objective of this study was to evaluate the efficacy of replication-defective adenovirus 5 (Ad5) vectors encoding either of two LT-based mucosal adjuvants, LTB or LTR72. These vectored adjuvants were delivered intranasally to mice concurrent with an Ad5-FMDV vaccine (Ad5-A24) to assess their ability to augment mucosal and systemic humoral immune responses to Ad5-A24 and protection against FMDV. Mice receiving Ad5-A24 plus Ad5-LTR72 had higher levels of mucosal and systemic neutralizing antibodies than those receiving Ad5-A24 alone or Ad5-A24 plus Ad5-LTB. The vaccine plus Ad5-LTR72 group also demonstrated 100% survival after intradermal challenge with a lethal dose of homologous FMDV serotype A24. These results suggest that Ad5-LTR72 could be used as an important tool to enhance mucosal and systemic immunity against FMDV and potentially other pathogens with a common route of entry. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Rift Valley fever virus and European mosquitoes: vector competence of Culex pipiens and Stegomyia albopicta (= Aedes albopictus).

    Science.gov (United States)

    Brustolin, M; Talavera, S; Nuñez, A; Santamaría, C; Rivas, R; Pujol, N; Valle, M; Verdún, M; Brun, A; Pagès, N; Busquets, N

    2017-12-01

    Rift Valley fever (RVF) is a mosquito-borne disease caused by the Rift Valley fever virus (RVFV). Rift Valley fever affects a large number of species, including human, and has severe impact on public health and the economy, especially in African countries. The present study examined the vector competence of three different European mosquito species, Culex pipiens (Linnaeus, 1758) form molestus (Diptera: Culicidae), Culex pipiens hybrid form and Stegomyia albopicta (= Aedes albopictus) (Skuse, 1894) (Diptera: Culicidae). Mosquitoes were artificially fed with blood containing RVFV. Infection, disseminated infection and transmission efficiency were evaluated. This is the first study to assess the transmission efficiency of European mosquito species using a virulent RVFV strain. The virus disseminated in Cx. pipiens hybrid form and in S. albopicta. Moreover, infectious viral particles were isolated from saliva of both species, showing their RVFV transmission capacity. The presence of competent Cx. pipiens and S. albopicta in Spain indicates that an autochthonous outbreak of RVF may occur if the virus is introduced. These findings provide information that will help health authorities to set up efficient entomological surveillance and RVFV vector control programmes. © 2017 The Authors. Medical and Veterinary Entomology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.

  16. Stomatal closure by ultraviolet radiation

    International Nuclear Information System (INIS)

    Negash, L.; Björn, L.O.

    1986-01-01

    The effect of ultraviolet radiation (UV) (255–325 nm) on stomatal closure was investigated on tef [Eragrostis tef (Zucc) Trotter] in the presence of white light (ca 50 ·mol m −2 s −1 ). The action spectrum showed that UV (ca 2 ·mol m −2 s −1 , half band width about 10 nm) of 285 nm or shorter wavelengths was very efficient in causing stomatal closure. The effectiveness decreased sharply towards longer wavelengths. Radiation of 313 nm or longer wavelengths was practically without effect. Increasing UV intensity increased stomatal resistance. When stronger white light (5 to 9 times stronger than the one used during irradiation) was administered, stomates re-opened rapidly irrespective of whether the UV was on or off, although a subsequent gradual closing tendency was observed when the UV was on. (author)

  17. Transduction of Photoreceptors With Equine Infectious Anemia Virus Lentiviral Vectors: Safety and Biodistribution of StarGen for Stargardt Disease

    Science.gov (United States)

    Binley, Katie; Widdowson, Peter; Loader, Julie; Kelleher, Michelle; Iqball, Sharifah; Ferrige, Georgina; de Belin, Jackie; Carlucci, Marie; Angell-Manning, Diana; Hurst, Felicity; Ellis, Scott; Miskin, James; Fernandes, Alcides; Wong, Paul; Allikmets, Rando; Bergstrom, Christopher; Aaberg, Thomas; Yan, Jiong; Kong, Jian; Gouras, Peter; Prefontaine, Annick; Vezina, Mark; Bussieres, Martin; Naylor, Stuart; Mitrophanous, Kyriacos A.

    2013-01-01

    Purpose. StarGen is an equine infectious anemia virus (EIAV)-based lentiviral vector that expresses the photoreceptor-specific adenosine triphosphate (ATP)-binding cassette transporter (ABCA4) protein that is mutated in Stargardt disease (STGD1), a juvenile macular dystrophy. EIAV vectors are able to efficiently transduce rod and cone photoreceptors in addition to retinal pigment epithelium in the adult macaque and rabbit retina following subretinal delivery. The safety and biodistribution of StarGen following subretinal delivery in macaques and rabbits was assessed. Methods. Regular ophthalmic examinations, IOP measurements, ERG responses, and histopathology were carried out in both species to compare control and vector-treated eyes. Tissue and fluid samples were obtained to evaluate the persistence, biodistribution, and shedding of the vector following subretinal delivery. Results. Ophthalmic examinations revealed a slightly higher level of inflammation in StarGen compared with control treated eyes in both species. However, inflammation was transient and no overt toxicity was observed in StarGen treated eyes and there were no abnormal clinical findings. There was no StarGen-associated rise in IOP or abnormal ERG response in either rabbits or macaques. Histopathologic examination of the eyes did not reveal any detrimental changes resulting from subretinal administration of StarGen. Although antibodies to StarGen vector components were detected in rabbit but not macaque serum, this immunologic response did not result in any long-term toxicity. Biodistribution analysis demonstrated that the StarGen vector was restricted to the ocular compartment. Conclusions. In summary, these studies demonstrate StarGen to be well tolerated and localized following subretinal administration. PMID:23620430

  18. Role of cellular FKBP52 protein in intracellular trafficking of recombinant adeno-associated virus 2 vectors

    International Nuclear Information System (INIS)

    Zhao Weihong; Zhong Li; Wu Jianqing; Chen Linyuan; Qing Keyun; Weigel-Kelley, Kirsten A.; Larsen, Steven H.; Shou Weinian; Warrington, Kenneth H.; Srivastava, Arun

    2006-01-01

    We have reported that tyrosine-phosphorylated forms of a cellular protein, FKBP52, inhibit the second-strand DNA synthesis of adeno-associated virus 2 (AAV), leading to inefficient transgene expression from recombinant AAV vectors. To further explore the role of FKBP52 in AAV-mediated transduction, we established murine embryo fibroblasts (MEFs) cultures from FKBP52 wild-type (WT), heterozygous (HE), and knockout (KO) mice. Conventional AAV vectors failed to transduce WT MEFs efficiently, and the transduction efficiency was not significantly increased in HE or KO MEFs. AAV vectors failed to traffic efficiently to the nucleus in these cells. Treatment with hydroxyurea (HU) increased the transduction efficiency of conventional AAV vectors by ∼25-fold in WT MEFs, but only by ∼4-fold in KO MEFs. The use of self-complementary AAV (scAAV) vectors, which bypass the requirement of viral second-strand DNA synthesis, revealed that HU treatment increased the transduction efficiency ∼23-fold in WT MEFs, but only ∼4-fold in KO MEFs, indicating that the lack of HU treatment-mediated increase in KO MEFs was not due to failure of AAV to undergo viral second-strand DNA synthesis. Following HU treatment, ∼59% of AAV genomes were present in the nuclear fraction from WT MEFs, but only ∼28% in KO MEFs, indicating that the pathway by which HU treatment mediates nuclear transport of AAV was impaired in KO MEFs. When KO MEFs were stably transfected with an FKBP52 expression plasmid, HU treatment-mediated increase in the transduction efficiency was restored in these cells, which correlated directly with improved intracellular trafficking. Intact AAV particles were also shown to interact with FKBP52 as well as with dynein, a known cellular protein involved in AAV trafficking. These studies suggest that FKBP52, being a cellular chaperone protein, facilitates intracellular trafficking of AAV, which has implications in the optimal use of recombinant AAV vectors in human gene

  19. Ecological Fitness of Non-vector Planthopper Sogatella furcifera on Rice Plants Infected with Rice Black Streaked Dwarf Virus

    Directory of Open Access Journals (Sweden)

    Xiao-chan HE

    2012-12-01

    Full Text Available We evaluated the effects of rice black streak dwarf virus (RBSDV-infested rice plants on the ecological parameters and its relevant defensive and detoxification enzymes of white-backed planthopper (WBPH in laboratory for exploring the relationship between RBSDV and the non-vector planthopper. The results showed that nymph survival rate, female adult weight and fecundity, and egg hatchability of WBPH fed on RBSDV-infested rice plants did not markedly differ from those on healthy plants, whereas the female adult longevity and egg duration significantly shortened on diseased plants. Furthermore, significantly higher activities of defensive enzymes (dismutase, catalase and peroxidase and detoxification enzymes (acetylcholinesterase, carboxylesterase and glutathione S-transferase were found in WBPH adults fed on infected plants. Results implied that infestation by RBSDV increased the ecological fitness of non-vector planthopper population.

  20. Synchrony of sylvatic dengue isolations: a multi-host, multi-vector SIR model of dengue virus transmission in Senegal.

    Directory of Open Access Journals (Sweden)

    Benjamin M Althouse

    Full Text Available Isolations of sylvatic dengue-2 virus from mosquitoes, humans and non-human primates in Senegal show synchronized multi-annual dynamics over the past 50 years. Host demography has been shown to directly affect the period between epidemics in other pathogen systems, therefore, one might expect unsynchronized multi-annual cycles occurring in hosts with dramatically different birth rates and life spans. However, in Senegal, we observe a single synchronized eight-year cycle across all vector species, suggesting synchronized dynamics in all vertebrate hosts. In the current study, we aim to explore two specific hypotheses: 1 primates with different demographics will experience outbreaks of dengue at different periodicities when observed as isolated systems, and that coupling of these subsystems through mosquito biting will act to synchronize incidence; and 2 the eight-year periodicity of isolations observed across multiple primate species is the result of long-term cycling in population immunity in the host populations. To test these hypotheses, we develop a multi-host, multi-vector Susceptible, Infected, Removed (SIR model to explore the effects of coupling multiple host-vector systems of dengue virus transmission through cross-species biting rates. We find that under small amounts of coupling, incidence in the host species synchronize. Long-period multi-annual dynamics are observed only when prevalence in troughs reaches vanishingly small levels (< 10(-10, suggesting that these dynamics are inconsistent with sustained transmission in this setting, but are consistent with local dengue virus extinctions followed by reintroductions. Inclusion of a constant introduction of infectious individuals into the system causes the multi-annual periods to shrink, while the effects of coupling remain the same. Inclusion of a stochastic rate of introduction allows for multi-annual periods at a cost of reduced synchrony. Thus, we conclude that the eight-year period

  1. Assessing the tobacco-rattle-virus-based vectors system as an efficient gene silencing technique in Datura stramonium (Solanaceae).

    Science.gov (United States)

    Eftekhariyan Ghamsari, Mohammad Reza; Karimi, Farah; Mousavi Gargari, Seyed Latif; Hosseini Tafreshi, Seyed Ali; Salami, Seyed Alireza

    2014-12-01

    Datura stramonium is a well-known medicinal plant, which is important for its alkaloids. There are intrinsic limitations for the natural production of alkaloids in plants; metabolic engineering methods can be effectively used to conquer these limitations. In order for this the genes involved in corresponding pathways need to be studied. Virus-Induced Gene Silencing is known as a functional genomics technique to knock-down expression of endogenous genes. In this study, we silenced phytoene desaturase as a marker gene in D. stramonium in a heterologous and homologous manner by tobacco-rattle-virus-based VIGS vectors. Recombinant TRV vector containing pds gene from D. stramonium (pTRV2-Dspds) was constructed and injected into seedlings. The plants injected with pTRV2-Dspds showed photobleaching 2 weeks after infiltration. Spectrophotometric analysis demonstrated that the amount of chlorophylls and carotenoids in leaves of the bleached plants decreased considerably compared to that of the control plants. Semi-Quantitative RT-PCR results also confirmed that the expression of pds gene in the silenced plants was significantly reduced in comparison with the control plants. The results showed that the viral vector was able to influence the levels of total alkaloid content in D. stramonium. Our results illustrated that TRV-based VIGS vectors are able to induce effective and reliable functional gene silencing in D. stramonium as an alternative tool for studying the genes of interest in this plant, such as the targeted genes in tropane alkaloid biosynthetic pathway. The present work is the first report of establishing VIGS as an efficient method for transient silencing of any gene of interest in D. stramonium.

  2. Complementation between avirulent Newcastle disease virus and a fusion protein gene expressed from a retrovirus vector: requirements for membrane fusion.

    OpenAIRE

    Morrison, T; McQuain, C; McGinnes, L

    1991-01-01

    The cDNA derived from the fusion gene of the virulent AV strain of Newcastle disease virus (NDV) was expressed in chicken embryo cells by using a retrovirus vector. The fusion protein expressed in this system was transported to the cell surface and was efficiently cleaved into the disulfide-linked F1-F2 form found in infectious virions. The cells expressing the fusion gene grew normally and could be passaged many times. Monolayers of these cells would plaque, in the absence of trypsin, avirul...

  3. Human Immunodeficiency Virus Type 1 Cloning Vectors for Antiretroviral Resistance Testing

    OpenAIRE

    Martinez-Picado, Javier; Sutton, Lorraine; De Pasquale, Maria Pia; Savara, Anu V.; D’Aquila, Richard T.

    1999-01-01

    Better detection of minority human immunodeficiency virus type 1 (HIV-1) populations containing gene mutations may improve the usefulness of antiretroviral resistance testing for clinical management. Molecular cloning of HIV-1 PCR products which might improve minority detection can be slow and difficult, and commercially available recombinant virus assays test drug susceptibility of virus pools. We describe novel plasmids and simple methods for rapid cloning of HIV-1 PCR products from patient...

  4. Promotion of flowering by Apple latent spherical virus vector and virus elimination at high temperature allow accelerated breeding of apple and pear

    Directory of Open Access Journals (Sweden)

    Noriko eYamagishi

    2016-02-01

    Full Text Available Plant viral vectors are superior tools for genetic manipulation, allowing rapid induction or suppression of expression of a target gene in plants. This is a particularly effective technology for use in breeding fruit trees, which are difficult to manipulate using recombinant DNA technologies. We reported previously that if apple seed embryos (cotyledons are infected with an Apple latent spherical virus (ALSV vector (ALSV-AtFT/MdTFL1 concurrently expressing the Arabidopsis thaliana florigen (AtFT gene and suppressing the expression of the apple MdTFL1-1 gene, the period prior to initial flowering (generally lasts 5–12 years will be reduced to about two months. In this study, we examined whether or not ALSV vector technology can be used to promote flowering in pear, which undergoes a very long juvenile period (germination to flowering similar to that of apple. The MdTFL1 sequence in ALSV-AtFT/MdTFL1 was replaced with a portion of the pear PcTFL1-1 gene. The resulting virus (ALSV-AtFT/PcTFL1 and ALSV-AtFT/MdTFL1 were used individually for inoculation to pear cotyledons immediately after germination in two inoculation groups. Those inoculated with ALSV-AtFT/MdTFL1 and ALSV-AtFT/PcTFL1 then initiated flower bud formation starting one to three months after inoculation, and subsequently exhibited continuous flowering and fruition by pollination. Conversely, Japanese pear exhibited extremely low systemic infection rates when inoculated with ALSV-AtFT/MdTFL1, and failed to exhibit any induction of flowering. We also developed a simple method for eliminating ALSV vectors from infected plants. An evaluation of the method for eliminating the ALSV vectors from infected apple and pear seedlings revealed that a four-week high-temperature (37˚C incubation of ALSV-infected apples and pears disabled the movement of ALSV to new growing tissues. This demonstrates that only high-temperature treatment can easily eliminate ALSV from infected fruit trees. A method

  5. Promotion of Flowering by Apple Latent Spherical Virus Vector and Virus Elimination at High Temperature Allow Accelerated Breeding of Apple and Pear.

    Science.gov (United States)

    Yamagishi, Norioko; Li, Chunjiang; Yoshikawa, Nobuyuki

    2016-01-01

    Plant viral vectors are superior tools for genetic manipulation, allowing rapid induction or suppression of expression of a target gene in plants. This is a particularly effective technology for use in breeding fruit trees, which are difficult to manipulate using recombinant DNA technologies. We reported previously that if apple seed embryos (cotyledons) are infected with an Apple latent spherical virus (ALSV) vector (ALSV-AtFT/MdTFL1) concurrently expressing the Arabidopsis thaliana florigen (AtFT) gene and suppressing the expression of the apple MdTFL1-1 gene, the period prior to initial flowering (generally lasts 5-12 years) will be reduced to about 2 months. In this study, we examined whether or not ALSV vector technology can be used to promote flowering in pear, which undergoes a very long juvenile period (germination to flowering) similar to that of apple. The MdTFL1 sequence in ALSV-AtFT/MdTFL1 was replaced with a portion of the pear PcTFL1-1 gene. The resulting virus (ALSV-AtFT/PcTFL1) and ALSV-AtFT/MdTFL1 were used individually for inoculation to pear cotyledons immediately after germination in two inoculation groups. Those inoculated with ALSV-AtFT/MdTFL1 and ALSV-AtFT/PcTFL1 then initiated flower bud formation starting one to 3 months after inoculation, and subsequently exhibited continuous flowering and fruition by pollination. Conversely, Japanese pear exhibited extremely low systemic infection rates when inoculated with ALSV-AtFT/MdTFL1, and failed to exhibit any induction of flowering. We also developed a simple method for eliminating ALSV vectors from infected plants. An evaluation of the method for eliminating the ALSV vectors from infected apple and pear seedlings revealed that a 4-week high-temperature (37°C) incubation of ALSV-infected apples and pears disabled the movement of ALSV to new growing tissues. This demonstrates that only high-temperature treatment can easily eliminate ALSV from infected fruit trees. A method combining the promotion

  6. Experimental and Natural Infections of Goats with Severe Fever with Thrombocytopenia Syndrome Virus: Evidence for Ticks as Viral Vector.

    Directory of Open Access Journals (Sweden)

    Yongjun Jiao

    Full Text Available Severe fever with thrombocytopenia syndrome virus (SFTSV, the causative agent for the fatal life-threatening infectious disease, severe fever with thrombocytopenia syndrome (SFTS, was first identified in the central and eastern regions of China. Although the viral RNA was detected in free-living and parasitic ticks, the vector for SFTSV remains unsettled.Firstly, an experimental infection study in goats was conducted in a bio-safety level-2 (BSL-2 facility to investigate virus transmission between animals. The results showed that infected animals did not shed virus to the outside through respiratory or digestive tract route, and the control animals did not get infected. Then, a natural infection study was carried out in the SFTSV endemic region. A cohort of naïve goats was used as sentinel animals in the study site. A variety of daily samples including goat sera, ticks and mosquitoes were collected for viral RNA and antibody (from serum only detection, and virus isolation. We detected viral RNA from free-living and parasitic ticks rather than mosquitoes, and from goats after ticks' infestation. We also observed sero-conversion in all members of the animal cohort subsequently. The S segment sequences of the two recovered viral isolates from one infected goat and its parasitic ticks showed a 100% homology at the nucleic acid level.In our natural infection study, close contact between goats does not appear to transmit SFTSV, however, the naïve animals were infected after ticks' infestation and two viral isolates derived from an infected goat and its parasitic ticks shared 100% of sequence identity. These data demonstrate that the etiologic agent for goat cohort's natural infection comes from environmental factors. Of these, ticks, especially the predominant species Haemaphysalis longicornis, probably act as vector for this pathogen. The findings in this study may help local health authorities formulate and focus preventive measures to contain

  7. Experimental and Natural Infections of Goats with Severe Fever with Thrombocytopenia Syndrome Virus: Evidence for Ticks as Viral Vector.

    Science.gov (United States)

    Jiao, Yongjun; Qi, Xian; Liu, Dapeng; Zeng, Xiaoyan; Han, Yewu; Guo, Xiling; Shi, Zhiyang; Wang, Hua; Zhou, Minghao

    2015-01-01

    Severe fever with thrombocytopenia syndrome virus (SFTSV), the causative agent for the fatal life-threatening infectious disease, severe fever with thrombocytopenia syndrome (SFTS), was first identified in the central and eastern regions of China. Although the viral RNA was detected in free-living and parasitic ticks, the vector for SFTSV remains unsettled. Firstly, an experimental infection study in goats was conducted in a bio-safety level-2 (BSL-2) facility to investigate virus transmission between animals. The results showed that infected animals did not shed virus to the outside through respiratory or digestive tract route, and the control animals did not get infected. Then, a natural infection study was carried out in the SFTSV endemic region. A cohort of naïve goats was used as sentinel animals in the study site. A variety of daily samples including goat sera, ticks and mosquitoes were collected for viral RNA and antibody (from serum only) detection, and virus isolation. We detected viral RNA from free-living and parasitic ticks rather than mosquitoes, and from goats after ticks' infestation. We also observed sero-conversion in all members of the animal cohort subsequently. The S segment sequences of the two recovered viral isolates from one infected goat and its parasitic ticks showed a 100% homology at the nucleic acid level. In our natural infection study, close contact between goats does not appear to transmit SFTSV, however, the naïve animals were infected after ticks' infestation and two viral isolates derived from an infected goat and its parasitic ticks shared 100% of sequence identity. These data demonstrate that the etiologic agent for goat cohort's natural infection comes from environmental factors. Of these, ticks, especially the predominant species Haemaphysalis longicornis, probably act as vector for this pathogen. The findings in this study may help local health authorities formulate and focus preventive measures to contain this infection.

  8. Activity of aphids associated with lettuce and broccoli in Spain and their efficiency as vectors of Lettuce mosaic virus.

    Science.gov (United States)

    Nebreda, M; Moreno, A; Pérez, N; Palacios, I; Seco-Fernández, V; Fereres, A

    2004-03-01

    This research sought to identify the aphid virus vector species associated with lettuce and broccoli crops in Spain, and to determine their population dynamics and ability to transmit Lettuce mosaic virus (LMV). Green tile traps and Moericke yellow water-pan traps were used to monitor aphid flights during the spring and autumn growing seasons of 2001. Aphid species feeding on lettuce were counted weekly. The transmission efficiencies of LMV were determined for the aphid species caught most frequently. The Moericke traps generally caught more aphid species than the tile trap, but the latter was the most suitable to estimate flight activity of species involved in virus spread. Spring aphid catches indicated that the main aphid species landing on lettuce in the regions of Madrid and Murcia was Hyperomyzus lactucae, but Brachycaudus helichrysi was also abundant in both regions. In broccoli in the Navarra region, the most abundant species in spring were Aphis fabae, B. helichrysi and H. lactucae. In autumn-sown crops, the main species landing on lettuce in the Madrid region were Hyadaphis coriandri and Aphis spiraecola. In Murcia, A. spiraecola and Myzus persicae were the most abundant, while in Navarra, Therioaphis trifolii, and various Aphis spp. were the most numerous landing on broccoli. The main aphid species colonising lettuce was Nasonovia ribisnigri, but other less abundant colonising species were Aulacorthum solani and Macrosiphum euphorbiae. The most efficient vectors of LMV were M. persicae, Aphis gossypii and M. euphorbiae, while A. fabae and H. lactucae transmitted with low efficiency, and Rhopalosiphum padi and N. ribisnigri did not transmit. Occurrence of LMV epidemics in central Spain in relation to aphid flights and the role of weeds as virus reservoirs is discussed.

  9. Migration of rice planthoppers and their vectored re-emerging and novel rice viruses in East Asia.

    Science.gov (United States)

    Otuka, Akira

    2013-10-28

    This review examines recent studies of the migration of three rice planthoppers, Laodelphax striatellus, Sogatella furcifera, and Nilaparvata lugens, in East Asia. Laodelphax striatellus has recently broken out in Jiangsu province, eastern China. The population density in the province started to increase in the early 2000s and peaked in 2004. In 2005, Rice stripe virus (RSV) viruliferous rate of L. striatellus peaked at 31.3%. Since then, rice stripe disease spread severely across the whole province. Due to the migration of the RSV vectors, the rice stripe disease spread to neighboring countries Japan and Korea. An overseas migration of L. striatellus that occurred in 2008 was analyzed, when a slow-moving cold vortex, a type of low pressure system, reached western Japan from Jiangsu, carrying the insects into Japan. Subsequently the rice stripe diseases struck these areas in Japan severely. In Korea, similar situations occurred in 2009, 2011, and 2012. Their migration sources were also estimated to be in Jiangsu by backward trajectory analysis. Rice black-streaked dwarf virus, whose vector is L. striatellus, has recently re-emerged in eastern China, and the evidence for overseas migrations of the virus, just like the RSV's migrations, has been given. A method of predicting the overseas migration of L. striatellus has been developed by Japanese, Chinese, and Korean institutes. An evaluation of the prediction showed that this method properly predicted migration events that occurred in East Asia from 2008 to 2011. Southern rice black-streaked dwarf virus (SRBSDV) was first found in Guangdong province. Its vector is S. furcifera. An outbreak of SRBSDV occurred in southern China in 2009 and spread to Vietnam the same year. This disease and virus were also found in Japan in 2010. The epidemic triggered many migration studies to investigate concrete spring-summer migration routes in China, and the addition of migration sources for early arrivals in Guangdong and Guangxi

  10. Identification of the subgenomic promoter of the coat protein gene of cucumber fruit mottle mosaic virus and development of a heterologous expression vector.

    Science.gov (United States)

    Rhee, Sun-Ju; Jang, Yoon Jeong; Lee, Gung Pyo

    2016-06-01

    Heterologous gene expression using plant virus vectors enables research on host-virus interactions and the production of useful proteins, but the host range of plant viruses limits the practical applications of such vectors. Here, we aimed to develop a viral vector based on cucumber fruit mottle mosaic virus (CFMMV), a member of the genus Tobamovirus, whose members infect cucurbits. The subgenomic promoter (SGP) in the coat protein (CP) gene, which was used to drive heterologous expression, was mapped by analyzing deletion mutants from a CaMV 35S promoter-driven infectious CFMMV clone. The region from nucleotides (nt) -55 to +160 relative to the start codon of the open reading frame (ORF) of CP was found to be a fully active promoter, and the region from nt -55 to +100 was identified as the active core promoter. Based on these SGPs, we constructed a cloning site in the CFMMV vector and successfully expressed enhanced green fluorescent protein (EGFP) in Nicotiana benthamiana and watermelon (Citrullus lanatus). Co-inoculation with the P19 suppressor increased EGFP expression and viral replication by blocking degradation of the viral genome. Our CFMMV vector will be useful as an expression vector in cucurbits.

  11. Minor Coat and Heat Shock Proteins Are Involved in the Binding of Citrus Tristeza Virus to the Foregut of Its Aphid Vector, Toxoptera citricida.

    Science.gov (United States)

    Killiny, N; Harper, S J; Alfaress, S; El Mohtar, C; Dawson, W O

    2016-11-01

    Vector transmission is a critical stage in the viral life cycle, yet for most plant viruses how they interact with their vector is unknown or is explained by analogy with previously described relatives. Here we examined the mechanism underlying the transmission of citrus tristeza virus (CTV) by its aphid vector, Toxoptera citricida, with the objective of identifying what virus-encoded proteins it uses to interact with the vector. Using fluorescently labeled virions, we demonstrated that CTV binds specifically to the lining of the cibarium of the aphid. Through in vitro competitive binding assays between fluorescent virions and free viral proteins, we determined that the minor coat protein is involved in vector interaction. We also found that the presence of two heat shock-like proteins, p61 and p65, reduces virion binding in vitro Additionally, treating the dissected mouthparts with proteases did not affect the binding of CTV virions. In contrast, chitinase treatment reduced CTV binding to the foregut. Finally, competition with glucose, N-acetyl-β-d-glucosamine, chitobiose, and chitotriose reduced the binding. These findings together suggest that CTV binds to the sugar moieties of the cuticular surface of the aphid cibarium, and the binding involves the concerted activity of three virus-encoded proteins. Limited information is known about the specific interactions between citrus tristeza virus and its aphid vectors. These interactions are important for the process of successful transmission. In this study, we localized the CTV retention site as the cibarium of the aphid foregut. Moreover, we demonstrated that the nature of these interactions is protein-carbohydrate binding. The viral proteins, including the minor coat protein and two heat shock proteins, bind to sugar moieties on the surface of the foregut. These findings will help in understanding the transmission mechanism of CTV by the aphid vector and may help in developing control strategies which interfere

  12. The successful induction of T-cell and antibody responses by a recombinant measles virus-vectored tetravalent dengue vaccine provides partial protection against dengue-2 infection.

    Science.gov (United States)

    Hu, Hui-Mei; Chen, Hsin-Wei; Hsiao, Yu-Ju; Wu, Szu-Hsien; Chung, Han-Hsuan; Hsieh, Chun-Hsiang; Chong, Pele; Leng, Chih-Hsiang; Pan, Chien-Hsiung

    2016-07-02

    Dengue has a major impact on global public health, and the use of dengue vaccine is very limited. In this study, we evaluated the immunogenicity and protective efficacy of a dengue vaccine made from a recombinant measles virus (MV) that expresses envelope protein domain III (ED3) of dengue-1 to 4. Following immunization with the MV-vectored dengue vaccine, mice developed specific interferon-gamma and antibody responses against dengue virus and MV. Neutralizing antibodies against MV and dengue viruses were also induced, and protective levels of FRNT50 ≥ 10 to 4 serotypes of dengue viruses were detected in the MV-vectored dengue vaccine-immunized mice. In addition, specific interferon-gamma and antibody responses to dengue viruses were still induced by the MV-vectored dengue vaccine in mice that were pre-infected with MV. This finding suggests that the pre-existing immunity to MV did not block the initiation of immune responses. By contrast, mice that were pre-infected with dengue-3 exhibited no effect in terms of their antibody responses to MV and dengue viruses, but a dominant dengue-3-specific T-cell response was observed. After injection with dengue-2, a detectable but significantly lower viremia and a higher titer of anti-dengue-2 neutralizing antibodies were observed in MV-vectored dengue vaccine-immunized mice versus the vector control, suggesting that an anamnestic antibody response that provided partial protection against dengue-2 was elicited. Our results with regard to T-cell responses and the effect of pre-immunity to MV or dengue viruses provide clues for the future applications of an MV-vectored dengue vaccine.

  13. Vesicular stomatitis virus (indiana 2 serotype as experimental model to study acute encephalitis – morphological features Vírus da estomatite vesicular (sorotipo indiana 2 como modelo experimental para o estudo de encefalite aguda – aspectos morfológicos

    Directory of Open Access Journals (Sweden)

    Florêncio Figueiredo Cavalcanti Neto

    2003-10-01

    Full Text Available The Vesicular Stomatitis Virus (VSV is a Vesiculovirus of the Rhabdoviridae family that infects mammals and causes vesicular lesions similar to those of foot-and-mouth disease. VSV experimental encephalitis can be induced in rodents and the symptoms are similar to those observed in rabies. However, the lesions observed in the animals´ encephalon are different. Inclusion bodies are not observed. There is necrosis, particularly in the region of the olfactory bulb, and, in some cases, ventriculitis. It was observed that the time pattern of VSV dissemination and the morphological aspects of the lesions are similar to those described in literature. The virus seems to be disseminated through the brain ventricles, being multiplied in the ependyma cells and in the neurons, besides using retrograde and anterograde transport. It was noticed that, due to the facility of virus manipulation, this experimental model has been used in innumerable research studies in several fields. If, on the one hand there are plenty of reports on the infection pathogenesis, on the other hand there are many gaps involving, for instance, aspects about virus transmission, recovery of infected animals and participation of glial cells in the acute as well as in the recovery phases.   O vírus da estomatite vesicular (VEV é um Vesiculovírus da família Rhabdoviridae que infecta mamíferos e causa lesões vesiculares semelhantes às observadas na febre aftosa. A encefalite experimental pode ser induzida em roedores e os sintomas são semelhantes aos observados na raiva; entretanto, as lesões observadas no encéfalo dos animais são diferentes. Corpúsculos de inclusão não são observados, há necrose especialmente da região do bulbo olfatório e em alguns casos, ventriculite. Observamos que o padrão temporal de disseminação do VEV e os aspectos morfológicos das lesões são similares aos descritos na literatura. O vírus parece se disseminar através dos ventr

  14. Low titer lentiviral transgenesis in rodents with simian immundeficiency virus vector.

    Science.gov (United States)

    Bender, Balázs; Hoffmann, Orsolya Ivett; Negre, Didier; Kvell, Krisztián; Bősze, Zsuzsanna; Hiripi, László

    2013-09-01

    Efficient production of transgenic animals using low-titer lentiviral constructs remains challenging. Here we demonstrate that microinjection of simian immundeficiency virus-derived lentiviral constructs can produce transgenic mice and rats with high efficiency even when using low-titer virus preparations.

  15. Expression of heterologous genes from an IRES translational cassette in replication competent murine leukemia virus vectors

    DEFF Research Database (Denmark)

    Jespersen, Thomas; Duch, Mogens R.; Carrasco, M L

    1999-01-01

    We describe replication competent retroviruses capable of expressing heterologous genes during multiple rounds of infection. An internal ribosome entry site (IRES) from encephalomyocarditis virus was inserted in the U3 region of Akv- and SL3-3-murine leukemia viruses (MLV) to direct translation...

  16. Effects of stomatal development on stomatal conductance and on stomatal limitation of photosynthesis in Syringa oblata and Euonymus japonicus Thunb.

    Science.gov (United States)

    Wu, Bing-Jie; Chow, Wah Soon; Liu, Yu-Jun; Shi, Lei; Jiang, Chuang-Dao

    2014-12-01

    During leaf development, the increase in stomatal conductance cannot meet photosynthetic demand for CO2, thus leading to stomatal limitation of photosynthesis (Ls). Considering the crucial influences of stomatal development on stomatal conductance, we speculated whether stomatal development limits photosynthesis to some extent. To test this hypothesis, stomatal development, stomatal conductance and photosynthesis were carefully studied in both Syringa oblata (normal greening species) and Euonymus japonicus Thunb (delayed greening species). Our results show that the size of stomata increased gradually with leaf expansion, resulting in increased stomatal conductance up to the time of full leaf expansion. During this process, photosynthesis also increased steadily. Compared to that in S. oblata, the development of chloroplasts in E. japonicus Thunb was obviously delayed, leading to a delay in the improvement of photosynthetic capacity. Further analysis revealed that before full leaf expansion, stomatal limitation increased rapidly in both S. oblata and E. japonicus Thunb; after full leaf expansion, stomatal limitation continually increased in E. japonicus Thunb. Accordingly, we suggested that the enhancement of photosynthetic capacity is the main factor leading to stomatal limitation during leaf development but that stomatal development can alleviate stomatal limitation with the increase of photosynthesis by controlling gas exchange. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  17. Ecological niche modeling of mosquito vectors of West Nile virus in St. John's County, Florida, USA.

    Science.gov (United States)

    Sallam, Mohamed F; Xue, Rui-De; Pereira, Roberto M; Koehler, Phillip G

    2016-06-29

    The lack of available vaccines and consistent sporadic transmission of WNV justify the need for mosquito vector control and prediction of their geographic distribution. However, the distribution of WNV transmission is dependent on the mosquito vector and the ecological requirements, which vary from one place to another. Presence/density data of two WNV mosquito vectors, Culex nigripalpus and Cx. quinquefasciatus, was extracted within 5 km buffer zones around seropositive records of sentinel chickens in order to delineate their predicting variables and model the habitat suitability of probable infective mosquito using MaxEnt software. Different correlations between density data of the extracted mosquito vectors and 27 climate, land use-land cover, and land surface terrain variables were analyzed using linear regression analysis. Accordingly, the correlated predicting variables were used in building up habitat suitability model for the occurrence records of both mosquito vectors using MaxEnt. The density of both WNV mosquito vectors showed variation in their ecological requirements. Eight predicting variables, out of 27, had significant influence on density of Cx. nigripalpus. Precipitation of driest months was shown to be the best predicting variable for the density of this vector (R (2) = 41.70). Whereas, two variables were proven to predict the distribution of Cx. quinquefasciatus density. Vegetation showed the maximum predicting gain to the density of this mosquito vector (R (2) = 15.74), where nestling birds, in particular exotics, are found. Moreover, Jackknife analysis in MaxEnt demonstrated that urbanization and vegetation data layers significantly contribute in predicting habitat suitability of Cx. nigripalpus and Cx. quinquefasciatus occurrence, respectively, which justifies the contribution of the former in urban and the latter in epizootic transmission cycles of WNV. In addition, habitat suitability risk maps were produced for both vectors in

  18. [Construction of RNA-containing virus-like nanoparticles expression vector with cysteine residues on surface and fluorescent decoration].

    Science.gov (United States)

    Cheng, Yang-Jian; Liang, Ji-Xuan; Li, Qing-Ge

    2005-08-01

    Site-directed mutagenesis was performed at the codon 15 of the MS2 bacteriophage coat protein gene,which had been cloned to the virus-like particles expression vector containing non-self RNA fragment. The produced expression vector,termed pARSC, was transformed to E. coli DH5alpha. The positive clones were selected and proliferated. The harvested cells were treated with sonication and the supernatant was then subjected to linear sucrose density gradients centrifugation (15% to 60%) at 32000 r/min for 4 h at 4 degrees C. The virus-like particles, VLP-Cy, were collected at 35% sucrose density. The particles were examined by transmission electron microscopy and the spherical viral particles of approximately 27 nm in diameter were found. The thiolated VLP-Cy was then chemically modified with fluorescein -5'-maleimide. The covalent fluorescent labeling was confirmed by absorption analysis, SDS-PAGE and MALDI-TOF mass spectroscopy. This is the first report of preparation of RNA-containing natural fluorescent nanoparticles. The study highlight the versatility of MS2 bacteriophage capsids as building blocks for functional nanomaterials construction for a variety of application purposes.

  19. [Development of transgenic maize with anti-rough dwarf virus artificial miRNA vector and their disease resistance].

    Science.gov (United States)

    Xuan, Ning; Zhao, Chuanzhi; Peng, Zhenying; Chen, Gao; Bian, Fei; Lian, Mingzheng; Liu, Guoxia; Wang, Xingjun; Bi, Yuping

    2015-09-01

    Maize is one of the most important food crops. Rice black-streaked dwarf virus is a maize rough dwarf disease pathogen. The occurrence and transmission of maize rough dwarf disease brings great damage to maize production. The technology of using artificial miRNA to build antiviral plant has been proven effective in a variety of plants. However, such trials in maize have not been reported. We designed primers based on the sequence of maize zea-miR159a precursor and sequence of function protein genes and silencing RBSDV coding genes in RBSDV genome. We constructed amiRNA (artificial miRNA) gene for silencing RBSDV coding gene and gene silencing suppressor. We constructed pCAMBIA3301-121-amiRNA plant expression vector for transforming maize inbred lines Z31 by using agrobacterium mediated method. After molecular analysis of transgenic maize, homozygous lines with high miRNA expression were selected by molecular detection for a subsequent natural infection experiment. We studied the severity of maize rough dwarf disease according to a grading standard (grade 0 to 4). The experiment results showed that the disease resistance of transgenic homozygous maize with the anti-rough dwarf virus amiRNA vector was better than that of wild type. Among the transgenic maize, S6-miR159 transgenic maize had high disease resistance. It is feasible to create new maize variety by the use of artificial miRNA.

  20. Frequent dual initiation of reverse transcription in murine leukemia virus-based vectors containing two primer-binding sites

    International Nuclear Information System (INIS)

    Voronin, Yegor A.; Pathak, Vinay K.

    2003-01-01

    Retroviruses package two copies of viral RNA into each virion. Although each RNA contains a primer-binding site for initiation of DNA synthesis, it is unknown whether reverse transcription is initiated on both RNAs. To determine whether a single virion is capable of initiating reverse transcription more than once, we constructed a murine leukemia virus-based vector containing a second primer-binding site (PBS) derived from spleen necrosis virus and inserted the green fluorescent protein gene (GFP) between the two PBSs. Initiation of reverse transcription at either PBS results in a provirus that expresses GFP. However, initiation at both PBSs can result in the deletion of GFP, which can be detected by flow cytometry and Southern blotting analysis. Approximately 22-29% of the proviruses formed deleted the GFP in a single replication cycle, indicating the minimum proportion of virions that initiated reverse transcription on both PBSs. These results show that a significant proportion of MLV-based vectors containing two PBSs have the capacity to initiate reverse transcription more than once

  1. Vector competence of Malaysian Aedes albopictus with and without Wolbachia to four dengue virus serotypes.

    Science.gov (United States)

    Joanne, Sylvia; Vythilingam, Indra; Teoh, Boon-Teong; Leong, Cherng-Shii; Tan, Kim-Kee; Wong, Meng-Li; Yugavathy, Nava; AbuBakar, Sazaly

    2017-09-01

    To determine the susceptibility status of Aedes albopictus with and without Wolbachia to the four dengue virus serotypes. Two newly colonised colonies of Ae. albopictus from the wild were used for the study. One colony was naturally infected with Wolbachia while in the other Wolbachia was removed by tetracycline treatment. Both colonies were orally infected with dengue virus-infected fresh blood meal. Dengue virus load was measured using quantitative RT-PCR at four-time intervals in the salivary glands, midguts and ovaries. Wolbachia did not significantly affect Malaysian Ae. albopictus dengue infection or the dissemination rate for all four dengue virus serotypes. Malaysian Ae. albopictus had the highest replication kinetics for DENV-1 and the highest salivary gland and midgut infection rate for DENV-4. Wolbachia, which naturally exists in Malaysian Ae. albopictus, does not significantly affect dengue virus replication. Malaysian Ae. albopictus is susceptible to dengue virus infections and capable of transmitting dengue virus, especially DENV-1 and DENV-4. Removal of Wolbachia from Malaysian Ae. albopictus would not reduce their susceptibility status. © 2017 John Wiley & Sons Ltd.

  2. Development of a new vector using Soybean yellow common mosaic virus for gene function study or heterologous protein expression in soybeans.

    Science.gov (United States)

    Lim, Seungmo; Nam, Moon; Kim, Kil Hyun; Lee, Su-Heon; Moon, Jung-Kyung; Lim, Hyoun-Sub; Choung, Myoung-Gun; Kim, Sang-Mok; Moon, Jae Sun

    2016-02-01

    A new vector using Soybean yellow common mosaic virus (SYCMV) was constructed for gene function study or heterologous protein expression in soybeans. The in vitro transcript with a 5' cap analog m7GpppG from an SYCMV full-length infectious vector driven by a T7 promoter infected soybeans (pSYCMVT7-full). The symptoms observed in the soybeans infected with either the sap from SYCMV-infected leaves or pSYCMVT7-full were indistinguishable, suggesting that the vector exhibits equivalent biological activity as the virus itself. To utilize the vector further, a DNA-based vector driven by the Cauliflower mosaic virus (CaMV) 35S promoter was constructed. The complete sequence of the SYCMV genome was inserted into a binary vector flanked by a CaMV 35S promoter at the 5' terminus of the SYCMV genome and a cis-cleaving ribozyme sequence followed by a nopaline synthase terminator at the 3' terminus of the SYCMV genome (pSYCMV-full). The SYCMV-derived vector was tested for use as a virus-induced gene silencing (VIGS) vector for the functional analysis of soybean genes. VIGS constructs containing either a fragment of the Phytoene desaturase (PDS) gene (pSYCMV-PDS1) or a fragment of the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RbcS) gene (pSYCMV-RbcS2) were constructed. Plants infiltrated with each vector using the Agrobacterium-mediated inoculation method exhibited distinct symptoms, such as photo-bleaching in plants infiltrated with pSYCMV-PDS1 and yellow or pale green coloring in plants infiltrated with pSYCMV-RbcS2. In addition, down-regulation of the transcripts of the two target genes was confirmed via northern blot analysis. Particle bombardment and direct plasmid DNA rubbing were also confirmed as alternative inoculation methods. To determine if the SYCMV vector can be used for the expression of heterologous proteins in soybean plants, the vector encoding amino acids 135-160 of VP1 of Foot-and-mouth disease virus (FMDV) serotype O1 Campos (O1C

  3. Development and characterization of a Rift Valley fever virus cell-cell fusion assay using alphavirus replicon vectors

    International Nuclear Information System (INIS)

    Filone, Claire Marie; Heise, Mark; Doms, Robert W.; Bertolotti-Ciarlet, Andrea

    2006-01-01

    Rift Valley fever virus (RVFV), a member of the Phlebovirus genus in the Bunyaviridae family, is transmitted by mosquitoes and infects both humans and domestic animals, particularly cattle and sheep. Since primary RVFV strains must be handled in BSL-3+ or BSL-4 facilities, a RVFV cell-cell fusion assay will facilitate the investigation of RVFV glycoprotein function under BSL-2 conditions. As for other members of the Bunyaviridae family, RVFV glycoproteins are targeted to the Golgi, where the virus buds, and are not efficiently delivered to the cell surface. However, overexpression of RVFV glycoproteins using an alphavirus replicon vector resulted in the expression of the glycoproteins on the surface of multiple cell types. Brief treatment of RVFV glycoprotein expressing cells with mildly acidic media (pH 6.2 and below) resulted in rapid and efficient syncytia formation, which we quantified by β-galactosidase α-complementation. Fusion was observed with several cell types, suggesting that the receptor(s) for RVFV is widely expressed or that this acid-dependent virus does not require a specific receptor to mediate cell-cell fusion. Fusion occurred over a broad temperature range, as expected for a virus with both mosquito and mammalian hosts. In contrast to cell fusion mediated by the VSV-G glycoprotein, RVFV glycoprotein-dependent cell fusion could be prevented by treating target cells with trypsin, indicating that one or more proteins (or protein-associated carbohydrate) on the host cell surface are needed to support membrane fusion. The cell-cell fusion assay reported here will make it possible to study the membrane fusion activity of RVFV glycoproteins in a high-throughput format and to screen small molecule inhibitors for the ability to block virus-specific membrane fusion

  4. Detection and identification of Rift Valley fever virus in mosquito vectors by quantitative real-time PCR.

    Science.gov (United States)

    Mwaengo, D; Lorenzo, G; Iglesias, J; Warigia, M; Sang, R; Bishop, R P; Brun, A

    2012-10-01

    Diagnostic methods allowing for rapid identification of pathogens are crucial for controlling and preventing dissemination after disease outbreaks as well as for use in surveillance programs. For arboviruses, detection of the presence of virus in their arthropod hosts is important for monitoring of viral activity and quantitative information is useful for modeling of transmission dynamics. In this study, molecular detection of Rift Valley fever virus (RVFV) in mosquito samples from the 2006 to 2007 East African outbreaks was performed using quantitative real-time PCR assay (qRT-PCR). Specific RVFV sequence-based primer/fluorogenic (TaqMan) probe sets were derived from the L and S RNA segments of the virus. Both primer-probe L and S segment-based combinations detected genomic RVFV sequences, with generally comparable levels of sensitivity. Viral loads from three mosquito species, Aedes mcintoshi, Aedes ochraceus and Mansonia uniformis were estimated and significant differences of between 5- and 1000-fold were detected between Ae. mcintoshi and M. uniformis using both the L and S primer-probe-based assays. The genetic relationships of the viral sequences in mosquito samples were established by partial M segment sequencing and assigned to the two previously described viral lineages defined by analysis of livestock isolates obtained during the 2006-2007 outbreak, confirming that similar viruses were present in both the vector and mammalian host. The data confirms the utility of qRT-PCR for identification and initial quantification of virus in mosquito samples during RVFV outbreaks. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. Evaluation of nonspreading Rift Valley fever virus as a vaccine vector using influenza virus hemagglutinin as a model antigen

    NARCIS (Netherlands)

    Oreshkova, N; Cornelissen, L A H M; de Haan, C A M; Moormann, R J M; Kortekaas, J

    2014-01-01

    Virus replicon particles are capable of infection, genome replication and gene expression, but are unable to produce progeny virions, rendering their use inherently safe. By virtue of this unique combination of features, replicon particles hold great promise for vaccine applications. We previously

  6. New type of Sendai virus vector provides transgene-free iPS cells derived from chimpanzee blood.

    Directory of Open Access Journals (Sweden)

    Yasumitsu Fujie

    Full Text Available Induced pluripotent stem cells (iPSCs are potentially valuable cell sources for disease models and future therapeutic applications; however, inefficient generation and the presence of integrated transgenes remain as problems limiting their current use. Here, we developed a new Sendai virus vector, TS12KOS, which has improved efficiency, does not integrate into the cellular DNA, and can be easily eliminated. TS12KOS carries KLF4, OCT3/4, and SOX2 in a single vector and can easily generate iPSCs from human blood cells. Using TS12KOS, we established iPSC lines from chimpanzee blood, and used DNA array analysis to show that the global gene-expression pattern of chimpanzee iPSCs is similar to those of human embryonic stem cell and iPSC lines. These results demonstrated that our new vector is useful for generating iPSCs from the blood cells of both human and chimpanzee. In addition, the chimpanzee iPSCs are expected to facilitate unique studies into human physiology and disease.

  7. Modeled response of the West Nile virus vector Culex quinquefasciatus to changing climate using the dynamic mosquito simulation model.

    Science.gov (United States)

    Morin, Cory W; Comrie, Andrew C

    2010-09-01

    Climate can strongly influence the population dynamics of disease vectors and is consequently a key component of disease ecology. Future climate change and variability may alter the location and seasonality of many disease vectors, possibly increasing the risk of disease transmission to humans. The mosquito species Culex quinquefasciatus is a concern across the southern United States because of its role as a West Nile virus vector and its affinity for urban environments. Using established relationships between atmospheric variables (temperature and precipitation) and mosquito development, we have created the Dynamic Mosquito Simulation Model (DyMSiM) to simulate Cx. quinquefasciatus population dynamics. The model is driven with climate data and validated against mosquito count data from Pasco County, Florida and Coachella Valley, California. Using 1-week and 2-week filters, mosquito trap data are reproduced well by the model (P climate projection data generated by the National Center for Atmospheric Research CCSM3 general circulation model, we applied temperature and precipitation offsets to the climate data at each location to evaluate mosquito population sensitivity to possible future climate conditions. We found that temperature and precipitation shifts act interdependently to cause remarkable changes in modeled mosquito population dynamics. Impacts include a summer population decline from drying in California due to loss of immature mosquito habitats, and in Florida a decrease in late-season mosquito populations due to drier late summer conditions.

  8. Investigations of barley stripe mosaic virus as a gene silencing vector in barley roots and in Brachypodium distachyon and oat

    Directory of Open Access Journals (Sweden)

    Nilsson Lena

    2010-11-01

    Full Text Available Abstract Background Gene silencing vectors based on Barley stripe mosaic virus (BSMV are used extensively in cereals to study gene function, but nearly all studies have been limited to genes expressed in leaves of barley and wheat. However since many important aspects of plant biology are based on root-expressed genes we wanted to explore the potential of BSMV for silencing genes in root tissues. Furthermore, the newly completed genome sequence of the emerging cereal model species Brachypodium distachyon as well as the increasing amount of EST sequence information available for oat (Avena species have created a need for tools to study gene function in these species. Results Here we demonstrate the successful BSMV-mediated virus induced gene silencing (VIGS of three different genes in barley roots, i.e. the barley homologues of the IPS1, PHR1, and PHO2 genes known to participate in Pi uptake and reallocation in Arabidopsis. Attempts to silence two other genes, the Pi transporter gene HvPht1;1 and the endo-β-1,4-glucanase gene HvCel1, in barley roots were unsuccessful, probably due to instability of the plant gene inserts in the viral vector. In B. distachyon leaves, significant silencing of the PHYTOENE DESATURASE (BdPDS gene was obtained as shown by photobleaching as well as quantitative RT-PCR analysis. On the other hand, only very limited silencing of the oat AsPDS gene was observed in both hexaploid (A. sativa and diploid (A. strigosa oat. Finally, two modifications of the BSMV vector are presented, allowing ligation-free cloning of DNA fragments into the BSMV-γ component. Conclusions Our results show that BSMV can be used as a vector for gene silencing in barley roots and in B. distachyon leaves and possibly roots, opening up possibilities for using VIGS to study cereal root biology and to exploit the wealth of genome information in the new cereal model plant B. distachyon. On the other hand, the silencing induced by BSMV in oat seemed too

  9. Disruption of Ethylene Responses by Turnip mosaic virus Mediates Suppression of Plant Defense against the Green Peach Aphid Vector.

    Science.gov (United States)

    Casteel, Clare L; De Alwis, Manori; Bak, Aurélie; Dong, Haili; Whitham, Steven A; Jander, Georg

    2015-09-01

    Plants employ diverse responses mediated by phytohormones to defend themselves against pathogens and herbivores. Adapted pathogens and herbivores often manipulate these responses to their benefit. Previously, we demonstrated that Turnip mosaic virus (TuMV) infection suppresses callose deposition, an important plant defense induced in response to feeding by its aphid vector, the green peach aphid (Myzus persicae), and increases aphid fecundity compared with uninfected control plants. Further, we determined that production of a single TuMV protein, Nuclear Inclusion a-Protease (NIa-Pro) domain, was responsible for changes in host plant physiology and increased green peach aphid reproduction. To characterize the underlying molecular mechanisms of this phenomenon, we examined the role of three phytohormone signaling pathways, jasmonic acid, salicylic acid, and ethylene (ET), in TuMV-infected Arabidopsis (Arabidopsis thaliana), with or without aphid herbivory. Experiments with Arabidopsis mutants ethylene insensitive2 and ethylene response1, and chemical inhibitors of ET synthesis and perception (aminoethoxyvinyl-glycine and 1-methylcyclopropene, respectively), show that the ET signaling pathway is required for TuMV-mediated suppression of Arabidopsis resistance to the green peach aphid. Additionally, transgenic expression of NIa-Pro in Arabidopsis alters ET responses and suppresses aphid-induced callose formation in an ET-dependent manner. Thus, disruption of ET responses in plants is an additional function of NIa-Pro, a highly conserved potyvirus protein. Virus-induced changes in ET responses may mediate vector-plant interactions more broadly and thus represent a conserved mechanism for increasing transmission by insect vectors across generations. © 2015 American Society of Plant Biologists. All Rights Reserved.

  10. Recombinant viruses obtained from co-infection in vitro with a live vaccinia-vectored influenza vaccine and a naturally occurring cowpox virus display different plaque phenotypes and loss of the transgene.

    Science.gov (United States)

    Hansen, Hilde; Okeke, Malachy Ifeanyi; Nilssen, Oivind; Traavik, Terje

    2004-12-09

    Some poxviruses are very attractive as transgenic vaccine vectors for humans, domestic animals and wildlife. Poxviridae family members circulate in different ecosystems and parts of the world, providing a pool of possible recombination partners for released or escaped genetically modified poxviruses. We performed in vitro double infections with a vaccinia virus strain Ankara (MVA) vectored influenza vaccine and a cowpox virus isolate from Norway, isolated hybrids, and further analyzed three hybrid viruses with different plaque phenotypes. One of the hybrids was genetically unstable, and during adaptation to new host cells its MVA derived influenza gene was deleted at a high frequency. This is significant in a risk assessment context, since the transgene would be the only logical tag for monitoring unwanted spread and non-target effects of a vaccine virus. Putative recombination events involving genetically modified and naturally occurring viruses should be included in health and environmental risk assessments.

  11. Landscape Genetics of Aedes mcintoshi (Diptera: Culicidae), an Important Vector of Rift Valley Fever Virus in Northeastern Kenya.

    Science.gov (United States)

    Campbell, Lindsay P; Alexander, Alana M

    2017-09-01

    Rift Valley fever virus (RVFV) is a vector-borne, zoonotic disease that affects humans, wild ungulates, and domesticated livestock in Africa and the Arabian Peninsula. Rift Valley fever virus exhibits interepizootic and epizootic phases, the latter defined by widespread virus occurrence in domesticated livestock. Kenya appears to be particularly vulnerable to epizootics, with 11 outbreaks occurring between 1951 and 2007. The mosquito species Aedes mcintoshi (subgenus Neomelaniconion) is an important primary vector for RVFV in Kenya. Here, we investigate associations between genetic diversity and differentiation of one regional subclade of Ae. mcintoshi in Northeastern Kenya with environmental variables, using a multivariate statistical approach. Using CO1 (cytochrome oxidase subunit 1) sequence data deposited in GenBank, we found no evidence of isolation by distance contributing to genetic differentiation across the study area. However, we did find significant CO1 subpopulation structure and associations with recent mean precipitation values. In addition, variation in genetic diversity across our seven sample sites was associated with both precipitation and percentage clay in the soil. The large number of haplotypes found in this data set indicates that a great deal of diversity remains unsampled in this region. Additional sampling across a larger geographic area, combined with next-generation sequencing approaches that better characterize the genome, would provide a more robust assessment of genetic diversity and differentiation. Further understanding of the genetic structure of Ae. mcintoshi could provide useful information regarding the potential for RVFV to spread across East African landscapes. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  12. Dengue fever virus in Pakistan: effects of seasonal pattern and temperature change on distribution of vector and virus.

    Science.gov (United States)

    Bostan, Nazish; Javed, Sundus; Nabgha-E-Amen; Eqani, Syed Ali Musstjab Akber Shah; Tahir, Faheem; Bokhari, Habib

    2017-01-01

    Dengue fever is regarded as one of the most prominent emerging arboviral infections in Pakistan since its first epidemic almost 2 decades ago. Interplay between potential vectors, susceptible host, and lax environmental conditions may promote the infection, leading to an epidemic. These factors may indeed have played a major role in the spread of the disease in the country, which was limited to Karachi till 2006. With recent natural disasters such as the earthquake in 2005 and flooding in 2010, 2011 and 2012, numbers of vector-borne diseases and outbreaks including dengue fever are on the rise in Pakistan. Therefore, it is a major concern for health sector workers and of utmost importance to have some understanding of the factors affecting disease outbreak for better risk assessment in the region. In the following report we review the climatic as well as host- and vector-associated factors involved in the outbreak of dengue epidemics in Pakistan and highlight high-risk zones in the country. Copyright © 2016 John Wiley & Sons, Ltd.

  13. Development of tobacco ringspot virus-based vectors for foreign gene expression and virus-induced gene silencing in a variety of plants.

    Science.gov (United States)

    Zhao, Fumei; Lim, Seungmo; Igori, Davaajargal; Yoo, Ran Hee; Kwon, Suk-Yoon; Moon, Jae Sun

    2016-05-01

    We report here the development of tobacco ringspot virus (TRSV)-based vectors for the transient expression of foreign genes and for the analysis of endogenous gene function in plants using virus-induced gene silencing. The jellyfish green fluorescent protein (GFP) gene was inserted between the TRSV movement protein (MP) and coat protein (CP) regions, resulting in high in-frame expression of the RNA2-encoded viral polyprotein. GFP was released from the polyprotein via an N-terminal homologous MP-CP cleavage site and a C-terminal foot-and-mouth disease virus (FMDV) 2 A catalytic peptide in Nicotiana benthamiana. The VIGS target gene was introduced in the sense and antisense orientations into a SnaBI site, which was created by mutating the sequence following the CP stop codon. VIGS of phytoene desaturase (PDS) in N. benthamiana, Arabidopsis ecotype Col-0, cucurbits and legumes led to obvious photo-bleaching phenotypes. A significant reduction in PDS mRNA levels in silenced plants was confirmed by semi-quantitative RT-PCR. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Tomato spotted wilt virus infection improves host suitability for its vector Frankliniella occidentalis

    NARCIS (Netherlands)

    Maris, P.C.; Joosten, N.N.; Goldbach, R.W.; Peters, D.

    2004-01-01

    The effect of Tomato spotted wilt virus (TSWV) infection on plant attractiveness for the western flower thrips (Frankliniella occidentalis) was studied. Significantly more thrips were recovered on infected than were recovered on noninfected pepper (Capsicum annuum) plants in different preference

  15. Climate change and the spread of vector-borne diseases: using approximate Bayesian computation to compare invasion scenarios for the bluetongue virus vector Culicoides imicola in Italy.

    Science.gov (United States)

    Mardulyn, Patrick; Goffredo, Maria; Conte, Annamaria; Hendrickx, Guy; Meiswinkel, Rudolf; Balenghien, Thomas; Sghaier, Soufien; Lohr, Youssef; Gilbert, Marius

    2013-05-01

    Bluetongue (BT) is a commonly cited example of a disease with a distribution believed to have recently expanded in response to global warming. The BT virus is transmitted to ruminants by biting midges of the genus Culicoides, and it has been hypothesized that the emergence of BT in Mediterranean Europe during the last two decades is a consequence of the recent colonization of the region by Culicoides imicola and linked to climate change. To better understand the mechanism responsible for the northward spread of BT, we tested the hypothesis of a recent colonization of Italy by C. imicola, by obtaining samples from more than 60 localities across Italy, Corsica, Southern France, and Northern Africa (the hypothesized source point for the recent invasion of C. imicola), and by genotyping them with 10 newly identified microsatellite loci. The patterns of genetic variation within and among the sampled populations were characterized and used in a rigorous approximate Bayesian computation framework to compare three competing historical hypotheses related to the arrival and establishment of C. imicola in Italy. The hypothesis of an ancient presence of the insect vector was strongly favoured by this analysis, with an associated P ≥ 99%, suggesting that causes other than the northward range expansion of C. imicola may have supported the emergence of BT in southern Europe. Overall, this study illustrates the potential of molecular genetic markers for exploring the assumed link between climate change and the spread of diseases. © 2013 Blackwell Publishing Ltd.

  16. Defining Challenges and Proposing Solutions for Control of the Virus VectorAedes aegypti

    OpenAIRE

    Gérardin, Patrick; Barau, Georges; Michault, Alain; Bintner, Marc; Randrianaivo, Hanitra; Choker, Ghassan; Lenglet, Yann; Touret, Yasmina; Bouveret, Anne; Grivard, Philippe; Roux, Karin Le; Blanc, Séverine; Schuffenecker, Isabelle; Couderc, Thérèse; Arenzana-Seisdedos, Fernando

    2008-01-01

    Editors' Summary Background. Chikungunya virus, an emerging infectious agent that is transmitted by day-biting mosquitoes, was first isolated from a patient in Tanzania in the early 1950s. Since then, major outbreaks of chikungunya fever have occurred throughout sub-Saharan Africa and in Southeast Asia, India, and the Western Pacific, usually at intervals of about 7–8 years. The virus causes fever, rash, severe joint and muscle pains, and sometimes arthritis (joint inflammation). These sympto...

  17. Alteration of intersubunit acid–base pair interactions at the quasi-threefold axis of symmetry of Cucumber mosaic virus disrupts aphid vector transmission

    Energy Technology Data Exchange (ETDEWEB)

    Bricault, Christine A. [Department of Plant Pathology and Plant-Microbe Biology, 334 Plant Science Building, Cornell University, Ithaca, NY 14850 (United States); Perry, Keith L., E-mail: KLP3@cornell.edu [Department of Plant Pathology and Plant-Microbe Biology, 334 Plant Science Building, Cornell University, Ithaca, NY 14850 (United States)

    2013-06-05

    In the atomic model of Cucumber mosaic virus (CMV), six amino acid residues form stabilizing salt bridges between subunits of the asymmetric unit at the quasi-threefold axis of symmetry. To evaluate the effects of these positions on virion stability and aphid vector transmissibility, six charged amino acid residues were individually mutated to alanine. All of the six engineered viruses were viable and exhibited near wild type levels of virion stability in the presence of urea. Aphid vector transmissibility was nearly or completely eliminated in the case of four of the mutants; two mutants demonstrated intermediate aphid transmissibility. For the majority of the engineered mutants, second-site mutations were observed following aphid transmission and/or mechanical passaging, and one restored transmission rates to that of the wild type. CMV capsids tolerate disruption of acid–base pairing interactions at the quasi-threefold axis of symmetry, but these interactions are essential for maintaining aphid vector transmissibility. - Highlights: ► Amino acids between structural subunits of Cucumber mosaic virus affect vector transmission. ► Mutant structural stability was retained, while aphid vector transmissibility was disrupted. ► Spontaneous, second-site mutations restored aphid vector transmissibility.

  18. Alteration of intersubunit acid–base pair interactions at the quasi-threefold axis of symmetry of Cucumber mosaic virus disrupts aphid vector transmission

    International Nuclear Information System (INIS)

    Bricault, Christine A.; Perry, Keith L.

    2013-01-01

    In the atomic model of Cucumber mosaic virus (CMV), six amino acid residues form stabilizing salt bridges between subunits of the asymmetric unit at the quasi-threefold axis of symmetry. To evaluate the effects of these positions on virion stability and aphid vector transmissibility, six charged amino acid residues were individually mutated to alanine. All of the six engineered viruses were viable and exhibited near wild type levels of virion stability in the presence of urea. Aphid vector transmissibility was nearly or completely eliminated in the case of four of the mutants; two mutants demonstrated intermediate aphid transmissibility. For the majority of the engineered mutants, second-site mutations were observed following aphid transmission and/or mechanical passaging, and one restored transmission rates to that of the wild type. CMV capsids tolerate disruption of acid–base pairing interactions at the quasi-threefold axis of symmetry, but these interactions are essential for maintaining aphid vector transmissibility. - Highlights: ► Amino acids between structural subunits of Cucumber mosaic virus affect vector transmission. ► Mutant structural stability was retained, while aphid vector transmissibility was disrupted. ► Spontaneous, second-site mutations restored aphid vector transmissibility

  19. Effects of Blood Coagulate Removal Method on Aedes albopictus (Diptera: Culicidae) Life Table Characteristics and Vector Competence for Dengue Virus.

    Science.gov (United States)

    van Dodewaard, Caitlin A M; Richards, Stephanie L; Harris, Jonathan W

    2016-01-01

    Commercially available blood can be used as an alternative to live animals to maintain mosquito colonies and deliver infectious bloodmeals during research studies. We analyzed the extent to which two methods for blood coagulate removal (defibrination or addition of sodium citrate) affected life table characteristics (i.e., fecundity, fertility, hatch rate, and adult survival) and vector competence (infection, dissemination, and transmission) of Aedes albopictus (Skuse) for dengue virus (DENV). Two types of bovine blood were tested at two extrinsic incubation temperatures (27 or 30°C) for DENV-infected and uninfected mosquitoes. Fully engorged mosquitoes were transferred to individual cages containing an oviposition cup and a substrate. Eggs (fecundity) and hatched larvae (fertility) were counted. At 14 and 21 d post feeding on a DENV-infected bloodmeal, 15 mosquitoes were sampled from each group, and vector competence was analyzed (bodies [infection], legs [dissemination], and saliva [transmission]). Differences in life table characteristics and vector competence were analyzed for mosquitoes fed blood processed using different methods for removal of coagulates. The method for removal of coagulates significantly impacted fecundity, fertility, and hatch time in the uninfected group, but not DENV-infected group. Infected mosquitoes showed significantly higher fecundity and faster hatch time than uninfected mosquitoes. We show no significant differences in infection or dissemination rates between groups; however, horizontal transmission rate was significantly higher in mosquitoes fed DENV-infected citrated compared with defibrinated blood. We expect the findings of this study to inform research using artificial blood delivery methods to assess vector competence. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  20. Culex quinquefasciatus (Diptera: Culicidae as a potential West Nile virus vector in Tucson, Arizona: Blood meal analysis indicates feeding on both humans and birds

    Directory of Open Access Journals (Sweden)

    Margaret Zinser

    2004-06-01

    Full Text Available Most reports from the United States suggest Culex quinquefasciatus mosquitoes feed minimally on humans. Given the abundance of C. quinquefasciatus in residential Tucson and parts of metropolitan Phoenix, and the arrival of West Nile virus to this area, discovering the blood meal hosts of the local population is important. Using a sandwich ELISA technique, the local C. quinquefasciatus were found to feed on both humans and birds. This suggests they should be considered potential West Nile virus vectors.

  1. Culex quinquefasciatus (Diptera: Culicidae) as a potential West Nile virus vector in Tucson, Arizona: Blood meal analysis indicates feeding on both humans and birds

    Science.gov (United States)

    Zinser, Margaret; Ramberg, Frank; Willott, Elizabeth

    2004-01-01

    Most reports from the United States suggest Culex quinquefasciatus mosquitoes feed minimally on humans. Given the abundance of C. quinquefasciatus in residential Tucson and parts of metropolitan Phoenix, and the arrival of West Nile virus to this area, discovering the blood meal hosts of the local population is important. Using a sandwich ELISA technique, the local C. quinquefasciatus were found to feed on both humans and birds. This suggests they should be considered potential West Nile virus vectors. PMID:15861236

  2. Multitracer Positron Emission Tomographic Imaging of Exogenous Gene Expression Mediated by a Universal Herpes Simplex Virus 1 Amplicon Vector

    Directory of Open Access Journals (Sweden)

    Christiane Kummer

    2007-05-01

    Full Text Available To develop efficient and safe gene therapy approaches, the herpes simplex virus type 1 thymidine kinase gene (HSV-1-tk has been shown to function as a marker gene for the direct noninvasive in vivo localization of thymidine kinase (TK expression by positron emission tomography (PET using radiolabeled nucleoside analogues as specific TK substrates. Moreover, the gene encoding dopamine type 2 receptor (d2r could be used as a PET marker gene using specific radiolabeled receptor binding compounds. Here we describe the quantitative colocalization of d2r and HSV-1-tk gene expression mediated from a universal HSV-1 amplicon vector in a subcutaneous human Gli36dEGFR glioma model by PET. The HSV-1 amplicon vector was constructed using a bicistronic gene cassette to contain (1 the d2r80A mutant, which is able to bind its ligand racloprid but unable to activate downstream signal transduction pathways, and (2 the tk39 mutant with enhanced enzymatic activity toward guanosine analogues fused to the green fluorescent protein gene (tk39gfp serving as a marker gene in cell culture. After infection of human Gli36dEGFR glioma cells with the HSV-d2r80AIREStk39gfp (HSV-DITG amplicon vector in cell culture, D2 receptor expression and its targeting to the cell surface were determined by Western blotting and immunolabeling. Vector application in vivo served for quantitative colocalization of d2r80A- and tk39gfp-derived PET signals employing the specific D2 receptor binding compound [11C]racloprid and the specific TK39 substrate 9-(4-[18F]fluoro-3-hydroxymethylbutylguanine. Our results demonstrate that for the range of gene expression studied in vivo, both enzymatic and receptor binding assays give comparable quantitative information on the level of vector-mediated gene expression in vivo. The d2r80A in combination with a specific binding compound passing the intact blood-brain barrier might be an alternative marker gene for the noninvasive assessment of vector

  3. The distribution of potential West Nile virus vectors, Culex pipiens pipiens and Culex pipiens quinquefasciatus (Diptera: Culicidae), in Mexico City

    Science.gov (United States)

    2011-01-01

    Background Culex spp. mosquitoes are considered to be the most important vectors of West Nile virus (WNV) detected in at least 34 species of mosquitoes in the United States. In North America, Culex pipiens pipiens, Culex pipiens quinquefasciatus, and Culex tarsalis are all competent vectors of WNV, which is considered to be enzootic in the United States and has also been detected in equines and birds in many states of Mexico and in humans in Nuevo Leon. There is potential for WNV to be introduced into Mexico City by various means including infected mosquitoes on airplanes, migrating birds, ground transportation and infected humans. Little is known of the geographic distribution of Culex pipiens complex mosquitoes and hybrids in Mexico City. Culex pipiens pipiens preferentially feed on avian hosts; Culex pipiens quinquefasciatus have historically been considered to prefer mammalian hosts; and hybrids of these two species could theoretically serve as bridge vectors to transmit WNV from avian hosts to humans and other mammalian hosts. In order to address the potential of WNV being introduced into Mexico City, we have determined the identity and spatial distribution of Culex pipiens complex mosquitoes and their hybrids. Results Mosquito larvae collected from 103 sites throughout Mexico City during 2004-2005 were identified as Culex, Culiseta or Ochlerotatus by morphological analysis. Within the genus Culex, specimens were further identified as Culex tarsalis or as belonging to the Culex pipiens complex. Members of the Culex pipiens complex were separated by measuring the ratio of the dorsal and ventral arms (DV/D ratio) of the male genitalia and also by using diagnostic primers designed for the Ace.2 gene. Culex pipiens quinquefasciatus was the most abundant form collected. Conclusions Important WNV vectors species, Cx. p. pipiens, Cx. p. quinquefasciatus and Cx. tarsalis, are all present in Mexico City. Hybrids of Cx. p. pipiens and Cx. p. quinquefasciatus were also

  4. The distribution of potential West Nile virus vectors, Culex pipiens pipiens and Culex pipiens quinquefasciatus (Diptera: Culicidae), in Mexico City.

    Science.gov (United States)

    Diaz-Badillo, Alvaro; Bolling, Bethany G; Perez-Ramirez, Gerardo; Moore, Chester G; Martinez-Munoz, Jorge P; Padilla-Viveros, America A; Camacho-Nuez, Minerva; Diaz-Perez, Alfonso; Beaty, Barry J; Munoz, Maria de Lourdes

    2011-05-09

    Culex spp. mosquitoes are considered to be the most important vectors of West Nile virus (WNV) detected in at least 34 species of mosquitoes in the United States. In North America, Culex pipiens pipiens, Culex pipiens quinquefasciatus, and Culex tarsalis are all competent vectors of WNV, which is considered to be enzootic in the United States and has also been detected in equines and birds in many states of Mexico and in humans in Nuevo Leon. There is potential for WNV to be introduced into Mexico City by various means including infected mosquitoes on airplanes, migrating birds, ground transportation and infected humans. Little is known of the geographic distribution of Culex pipiens complex mosquitoes and hybrids in Mexico City. Culex pipiens pipiens preferentially feed on avian hosts; Culex pipiens quinquefasciatus have historically been considered to prefer mammalian hosts; and hybrids of these two species could theoretically serve as bridge vectors to transmit WNV from avian hosts to humans and other mammalian hosts. In order to address the potential of WNV being introduced into Mexico City, we have determined the identity and spatial distribution of Culex pipiens complex mosquitoes and their hybrids. Mosquito larvae collected from 103 sites throughout Mexico City during 2004-2005 were identified as Culex, Culiseta or Ochlerotatus by morphological analysis. Within the genus Culex, specimens were further identified as Culex tarsalis or as belonging to the Culex pipiens complex. Members of the Culex pipiens complex were separated by measuring the ratio of the dorsal and ventral arms (DV/D ratio) of the male genitalia and also by using diagnostic primers designed for the Ace.2 gene. Culex pipiens quinquefasciatus was the most abundant form collected. Important WNV vectors species, Cx. p. pipiens, Cx. p. quinquefasciatus and Cx. tarsalis, are all present in Mexico City. Hybrids of Cx. p. pipiens and Cx. p. quinquefasciatus were also collected and identified. The

  5. The distribution of potential West Nile virus vectors, Culex pipiens pipiens and Culex pipiens quinquefasciatus (Diptera: Culicidae, in Mexico City

    Directory of Open Access Journals (Sweden)

    Diaz-Perez Alfonso

    2011-05-01

    Full Text Available Abstract Background Culex spp. mosquitoes are considered to be the most important vectors of West Nile virus (WNV detected in at least 34 species of mosquitoes in the United States. In North America, Culex pipiens pipiens, Culex pipiens quinquefasciatus, and Culex tarsalis are all competent vectors of WNV, which is considered to be enzootic in the United States and has also been detected in equines and birds in many states of Mexico and in humans in Nuevo Leon. There is potential for WNV to be introduced into Mexico City by various means including infected mosquitoes on airplanes, migrating birds, ground transportation and infected humans. Little is known of the geographic distribution of Culex pipiens complex mosquitoes and hybrids in Mexico City. Culex pipiens pipiens preferentially feed on avian hosts; Culex pipiens quinquefasciatus have historically been considered to prefer mammalian hosts; and hybrids of these two species could theoretically serve as bridge vectors to transmit WNV from avian hosts to humans and other mammalian hosts. In order to address the potential of WNV being introduced into Mexico City, we have determined the identity and spatial distribution of Culex pipiens complex mosquitoes and their hybrids. Results Mosquito larvae collected from 103 sites throughout Mexico City during 2004-2005 were identified as Culex, Culiseta or Ochlerotatus by morphological analysis. Within the genus Culex, specimens were further identified as Culex tarsalis or as belonging to the Culex pipiens complex. Members of the Culex pipiens complex were separated by measuring the ratio of the dorsal and ventral arms (DV/D ratio of the male genitalia and also by using diagnostic primers designed for the Ace.2 gene. Culex pipiens quinquefasciatus was the most abundant form collected. Conclusions Important WNV vectors species, Cx. p. pipiens, Cx. p. quinquefasciatus and Cx. tarsalis, are all present in Mexico City. Hybrids of Cx. p. pipiens and Cx. p

  6. Disease dynamics of honeybees with Varroa destructor as parasite and virus vector.

    Science.gov (United States)

    Kang, Yun; Blanco, Krystal; Davis, Talia; Wang, Ying; DeGrandi-Hoffman, Gloria

    2016-05-01

    The worldwide decline in honeybee colonies during the past 50 years has often been linked to the spread of the parasitic mite Varroa destructor and its interaction with certain honeybee viruses carried by Varroa mites. In this paper, we propose a honeybee-mite-virus model that incorporates (1) parasitic interactions between honeybees and the Varroa mites; (2) five virus transmission terms between honeybees and mites at different stages of Varroa mites: from honeybees to honeybees, from adult honeybees to the phoretic mites, from brood to the reproductive mites, from the reproductive mites to brood, and from adult honeybees to the phoretic mites; and (3) Allee effects in the honeybee population generated by its internal organization such as division of labor. We provide completed local and global analysis for the full system and its subsystems. Our analytical and numerical results allow us have a better understanding of the synergistic effects of parasitism and virus infections on honeybee population dynamics and its persistence. Interesting findings from our work include: (a) due to Allee effects experienced by the honeybee population, initial conditions are essential for the survival of the colony. (b) Low adult honeybees to brood ratios have destabilizing effects on the system which generate fluctuating dynamics that lead to a catastrophic event where both honeybees and mites suddenly become extinct. This catastrophic event could be potentially linked to Colony Collapse Disorder (CCD) of honeybee colonies. (c) Virus infections may have stabilizing effects on the system, and parasitic mites could make disease more persistent. Our model illustrates how the synergy between the parasitic mites and virus infections consequently generates rich dynamics including multiple attractors where all species can coexist or go extinct depending on initial conditions. Our findings may provide important insights on honeybee viruses and parasites and how to best control them

  7. Rapid Construction of Complex Plant RNA Virus Infectious cDNA Clones for Agroinfection Using a Yeast-E. coli-Agrobacterium Shuttle Vector.

    Science.gov (United States)

    Sun, Kai; Zhao, Danyang; Liu, Yong; Huang, Changjun; Zhang, Wei; Li, Zhenghe

    2017-11-07

    The availability of infectious full-length clone is indispensable for reverse genetics studies of virus biology, pathology and construction of viral vectors. However, for RNA viruses with large genome sizes or those exhibiting inherent cloning difficulties, procedure to generate biologically active circular DNA (cDNA) clones can be time-consuming or technically challenging. Here we have constructed a yeast- Escherichia coli - Agrobacterium shuttle vector that enables highly efficient homologous recombination in yeast for assembly of Agrobacterium compatible plant virus clones. Using this vector, we show that infectious cDNA clones of a plant negative-stranded RNA virus, sonchus yellow net rhabdovirus, can be rapidly assembled. In addition, one-step assembly of infectious clones of potato virus Y in yeast, either with or without intron, was readily achieved from as many as eight overlapping DNA fragments. More importantly, the recovered yeast plasmids can be transformed directly into Agrobacterium for inoculation, thereby obviating the E. coli cloning steps and associated toxicity issues. This method is rapid, highly efficient and cost-effective and should be readily applicable to a broad range of plant viruses.

  8. Cloning of replication-incompetent herpes simplex viruses as bacterial artificial chromosomes to facilitate development of vectors for gene delivery into differentiated neurons.

    Science.gov (United States)

    Schmeisser, Falko; Weir, Jerry P

    2006-01-01

    We have previously described the adaptation of a tetracycline-regulated system of gene expression for herpes simplex virus (HSV) vectors and demonstrated that such a system was capable of inducible foreign gene expression in irreversibly differentiated neurons. These studies suggested that such gene delivery vectors would be especially useful for studying the neuron in vitro. Here, we describe the cloning of a replication-incompetent HSV vector as a bacterial artificial chromosome (BAC) to facilitate vector construction. Using prokaryotic genetic techniques for allele replacement, we demonstrate the ease of manipulation of the BAC-containing vector, including the construction of vector mutations for which there is no simple phenotypic selection. Such constructions include the insertion of a tetracycline-regulated gene cassette into the UL41 gene for regulated gene expression and the mutation of the UL48 gene to reduce vector toxicity. In addition, HSV vectors cloned as BACs can be sequentially modified to make multiple changes to the vector platform. Finally, using the BAC system, we constructed an HSV vector that expressed an inducible human superoxide dismutase-1 (SOD1) gene for delivery into differentiated human NT-neurons (cells of the human embryonal carcinoma cell line NT2, which differentiate irreversibly into postmitotic neuron-like cells after treatment with retinoic acid). The results indicated that there is appreciable expression of SOD1 from this HSV vector in the presence of doxycycline and that vector-expressed SOD1 interacts with endogenous SOD1. Thus, the BAC system provides a practicable platform for construction and manipulation of HSV vectors that are suitable for gene delivery into postmitotic neurons in vitro.

  9. Population genetics of two key mosquito vectors of Rift Valley Fever virus reveals new insights into the changing disease outbreak patterns in Kenya.

    Directory of Open Access Journals (Sweden)

    David P Tchouassi

    2014-12-01

    Full Text Available Rift Valley fever (RVF outbreaks in Kenya have increased in frequency and range to include northeastern Kenya where viruses are increasingly being isolated from known (Aedes mcintoshi and newly-associated (Ae. ochraceus vectors. The factors contributing to these changing outbreak patterns are unclear and the population genetic structure of key vectors and/or specific virus-vector associations, in particular, are under-studied. By conducting mitochondrial and nuclear DNA analyses on >220 Kenyan specimens of Ae. mcintoshi and Ae. ochraceus, we uncovered high levels of vector complexity which may partly explain the disease outbreak pattern. Results indicate that Ae. mcintoshi consists of a species complex with one of the member species being unique to the newly-established RVF outbreak-prone northeastern region of Kenya, whereas Ae. ochraceus is a homogeneous population that appears to be undergoing expansion. Characterization of specimens from a RVF-prone site in Senegal, where Ae. ochraceus is a primary vector, revealed direct genetic links between the two Ae. ochraceus populations from both countries. Our data strongly suggest that unlike Ae. mcintoshi, Ae. ochraceus appears to be a relatively recent, single 'introduction' into Kenya. These results, together with increasing isolations from this vector, indicate that Ae. ochraceus will likely be of greater epidemiological importance in future RVF outbreaks in Kenya. Furthermore, the overall vector complexity calls into question the feasibility of mosquito population control approaches reliant on genetic modification.

  10. Population Genetics of Two Key Mosquito Vectors of Rift Valley Fever Virus Reveals New Insights into the Changing Disease Outbreak Patterns in Kenya

    Science.gov (United States)

    Tchouassi, David P.; Bastos, Armanda D. S.; Sole, Catherine L.; Diallo, Mawlouth; Lutomiah, Joel; Mutisya, James; Mulwa, Francis; Borgemeister, Christian; Sang, Rosemary; Torto, Baldwyn

    2014-01-01

    Rift Valley fever (RVF) outbreaks in Kenya have increased in frequency and range to include northeastern Kenya where viruses are increasingly being isolated from known (Aedes mcintoshi) and newly-associated (Ae. ochraceus) vectors. The factors contributing to these changing outbreak patterns are unclear and the population genetic structure of key vectors and/or specific virus-vector associations, in particular, are under-studied. By conducting mitochondrial and nuclear DNA analyses on >220 Kenyan specimens of Ae. mcintoshi and Ae. ochraceus, we uncovered high levels of vector complexity which may partly explain the disease outbreak pattern. Results indicate that Ae. mcintoshi consists of a species complex with one of the member species being unique to the newly-established RVF outbreak-prone northeastern region of Kenya, whereas Ae. ochraceus is a homogeneous population that appears to be undergoing expansion. Characterization of specimens from a RVF-prone site in Senegal, where Ae. ochraceus is a primary vector, revealed direct genetic links between the two Ae. ochraceus populations from both countries. Our data strongly suggest that unlike Ae. mcintoshi, Ae. ochraceus appears to be a relatively recent, single 'introduction' into Kenya. These results, together with increasing isolations from this vector, indicate that Ae. ochraceus will likely be of greater epidemiological importance in future RVF outbreaks in Kenya. Furthermore, the overall vector complexity calls into question the feasibility of mosquito population control approaches reliant on genetic modification. PMID:25474018

  11. The site of potato leafroll virus multiplication in its vector, Myzus persicae : an anatomical study

    NARCIS (Netherlands)

    Ponsen, M.B.

    1972-01-01

    In search of the site of PLRV multiplication in its vector a detailed study was made of the anatomy of the aphid, Myzus persicae SULZ. The findings are summarized in the following lines:

    Alimentary canal

    The most anterior part of

  12. Culex pipiens, an Experimental Efficient Vector of West Nile and Rift Valley Fever Viruses in the Maghreb Region

    Science.gov (United States)

    Amraoui, Fadila; Krida, Ghazi; Bouattour, Ali; Rhim, Adel; Daaboub, Jabeur; Harrat, Zoubir; Boubidi, Said-Chawki; Tijane, Mhamed; Sarih, Mhammed; Failloux, Anna-Bella

    2012-01-01

    West Nile fever (WNF) and Rift Valley fever (RVF) are emerging diseases causing epidemics outside their natural range of distribution. West Nile virus (WNV) circulates widely and harmlessly in the old world among birds as amplifying hosts, and horses and humans as accidental dead-end hosts. Rift Valley fever virus (RVFV) re-emerges periodically in Africa causing massive outbreaks. In the Maghreb, eco-climatic and entomologic conditions are favourable for WNV and RVFV emergence. Both viruses are transmitted by mosquitoes belonging to the Culex pipiens complex. We evaluated the ability of different populations of Cx. pipiens from North Africa to transmit WNV and the avirulent RVFV Clone 13 strain. Mosquitoes collected in Algeria, Morocco, and Tunisia during the summer 2010 were experimentally infected with WNV and RVFV Clone 13 strain at titers of 107.8 and 108.5 plaque forming units/mL, respectively. Disseminated infection and transmission rates were estimated 14–21 days following the exposure to the infectious blood-meal. We show that 14 days after exposure to WNV, all mosquito st developed a high disseminated infection and were able to excrete infectious saliva. However, only 69.2% of mosquito strains developed a disseminated infection with RVFV Clone 13 strain, and among them, 77.8% were able to deliver virus through saliva. Thus, Cx. pipiens from the Maghreb are efficient experimental vectors to transmit WNV and to a lesser extent, RVFV Clone 13 strain. The epidemiologic importance of our findings should be considered in the light of other parameters related to mosquito ecology and biology. PMID:22693557

  13. Virus infection decreases the attractiveness of white clover plants for a non-vectoring herbivore

    DEFF Research Database (Denmark)

    van Mölken, Tamara; Caluwe, Hannie de; Hordijk, Cornelis A.

    2012-01-01

    Plant pathogens and insect herbivores are prone to share hosts under natural conditions. Consequently, pathogen-induced changes in the host plant can affect herbivory, and vice versa. Even though plant viruses are ubiquitous in the field, little is known about plant-mediated interactions between...... feeding on plant growth, (2) the attractiveness of white clover for adult fungus gnat females, and (3) the volatile emission of white clover plants. We observed only marginal effects of WClMV infection on the interaction between fungus gnat larvae and white clover. However, adult fungus gnat females....... This suggests that virus infections may contribute to protecting their hosts by decreasing herbivore infestation rates. Consequently, it is conceivable that viruses play a more beneficial role in plant-herbivore interactions than generally thought....

  14. Investigations of barley stripe mosaic virus as a gene silencing vector in barley roots and in Brachypodium distachyon and oat

    DEFF Research Database (Denmark)

    Pacak, Andrzej; Geisler, Katrin; Jørgensen, Bodil

    2010-01-01

    -expressed genes we wanted to explore the potential of BSMV for silencing genes in root tissues. Furthermore, the newly completed genome sequence of the emerging cereal model species Brachypodium distachyon as well as the increasing amount of EST sequence information available for oat (Avena species) have created...... a need for tools to study gene function in these species. Results Here we demonstrate the successful BSMV-mediated virus induced gene silencing (VIGS) of three different genes in barley roots, i.e. the barley homologues of the IPS1, PHR1, and PHO2 genes known to participate in Pi uptake and reallocation...... the wealth of genome information in the new cereal model plant B. distachyon. On the other hand, the silencing induced by BSMV in oat seemed too weak to be of practical use. The new BSMV vectors modified for ligation-free cloning will allow rapid insertion of plant gene fragments for future experiments....

  15. An Engineered Virus Library as a Resource for the Spectrum-wide Exploration of Virus and Vector Diversity

    Directory of Open Access Journals (Sweden)

    Wenli Zhang

    2017-05-01

    Full Text Available Adenoviruses (Ads are large human-pathogenic double-stranded DNA (dsDNA viruses presenting an enormous natural diversity associated with a broad variety of diseases. However, only a small fraction of adenoviruses has been explored in basic virology and biomedical research, highlighting the need to develop robust and adaptable methodologies and resources. We developed a method for high-throughput direct cloning and engineering of adenoviral genomes from different sources utilizing advanced linear-linear homologous recombination (LLHR and linear-circular homologous recombination (LCHR. We describe 34 cloned adenoviral genomes originating from clinical samples, which were characterized by next-generation sequencing (NGS. We anticipate that this recombineering strategy and the engineered adenovirus library will provide an approach to study basic and clinical virology. High-throughput screening (HTS of the reporter-tagged Ad library in a panel of cell lines including osteosarcoma disease-specific cell lines revealed alternative virus types with enhanced transduction and oncolysis efficiencies. This highlights the usefulness of this resource.

  16. Transgene stability for three replication-competent murine leukemia virus vectors

    DEFF Research Database (Denmark)

    Duch, Mogens R.; Carrasco, Maria L; Jespersen, Thomas

    2004-01-01

    cassette consisting of an internal ribosome entry site followed by the enhanced green fluorescent protein coding sequence inserted in different configurations into murine leukemia virus genomes. In two of the constructs, the insert was located in the upstream part of the U3 region while in the third...

  17. Vectoring gypsy moth nuclear polyhedrosis virus by Apanteles melanoscelus (Hym.:Braconidae)

    Science.gov (United States)

    B. Raimo; R.C. Reardon; J.D. Podgwaite

    1977-01-01

    Gypsy moth Lymantria dispar L. larvae were exposed to Apanteles melanoscelus (Ratzeburg) females contaminated with nuclear polyhedrosis virus. Three methods of contamination (ovipositor, total body surface, and exposure to infected hosts) and two exposure periods (2 and 24 hours) were tested. A significantly greater incidence of...

  18. Efficiency of non-vector methods of Cassava brown streak virus ...

    African Journals Online (AJOL)

    Several etiological and epidemiological studies have been undertaken to determine the disease causal agent and the mechanism of spread of Cassava brown streak disease (CBSD). Until recently, two distinct potyviruses have been reported to cause the disease. These are Cassava brown streak virus (originated in ...

  19. Mites as vector of Tulip Virus X in stored tulip bulbs

    NARCIS (Netherlands)

    Lommen, S.T.E.; Conijn, C.G.M.; Lemmers, M.E.C.; Pham, K.T.K.; Kock, de M.J.D.

    2012-01-01

    Tulip virus X (TVX) is a Potexvirus causing economic losses in tulip. Potexviruses are generally transmitted by mechanical contact and, indeed, several mechanical transmission pathways for TVX have been identified during tulip bulb production. However, TVX transmission does also seem to occur during

  20. Vector Transmission Alone Fails to Explain the Potato Yellow Vein Virus Epidemic among Potato Crops in Colombia.

    Science.gov (United States)

    Cuadros, Diego F; Hernandez, Anngie; Torres, Maria F; Torres, Diana M; Branscum, Adam J; Rincon, Diego F

    2017-01-01

    The potato yellow vein disease, caused by the potato yellow vein virus (PYVV), is a limiting potato disease in northern South America. The virus can be transmitted either by the greenhouse whitefly (GWF), Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae), or through vegetative propagules, such as infected tubers. Recently, GWF populations have been spotlighted as one of the main drivers of PYVV re-emergence, and consequently, PYVV management has been predominantly directed toward vector control, which is heavily based on insecticide use. However, the drivers of the PYVV outbreaks as well as the contribution of GWF populations on the spread of PYVV among potato crops are still not completely understood. This study aims to assess the role of the GWF as a driver of the PYVV epidemic in the potato-producing areas in Colombia, one of the countries more severely affected by the PYVV epidemic, and whose geography allows the study of the spatial association between the vector and the disease epidemic across a wide altitude range. The geographical clusters where the PYVV epidemic is concentrated, as well as those of farms affected by the GWF were identified using a novel spatial epidemiology approach. The influence of altitude range on the association between PYVV and T. vaporarioum was also assessed. We found a relatively poor spatial association between PYVV epidemic and the presence of the GWF, especially at altitudes above 3,000 m above mean sea level. Furthermore, GWF populations could only explain a small fraction of the extent of the PYVV epidemic in Colombia. Movement of infected seed tubers might be the main mechanism of dispersion, and could be a key driver for the PYVV infection among potato crops. Agricultural policies focused on improving quality of seed tubers and their appropriate distribution could be the most efficient control intervention against PYVV dispersion.

  1. Vector Transmission Alone Fails to Explain the Potato Yellow Vein Virus Epidemic among Potato Crops in Colombia

    Directory of Open Access Journals (Sweden)

    Diego F. Cuadros

    2017-09-01

    Full Text Available The potato yellow vein disease, caused by the potato yellow vein virus (PYVV, is a limiting potato disease in northern South America. The virus can be transmitted either by the greenhouse whitefly (GWF, Trialeurodes vaporariorum (Westwood (Hemiptera: Aleyrodidae, or through vegetative propagules, such as infected tubers. Recently, GWF populations have been spotlighted as one of the main drivers of PYVV re-emergence, and consequently, PYVV management has been predominantly directed toward vector control, which is heavily based on insecticide use. However, the drivers of the PYVV outbreaks as well as the contribution of GWF populations on the spread of PYVV among potato crops are still not completely understood. This study aims to assess the role of the GWF as a driver of the PYVV epidemic in the potato-producing areas in Colombia, one of the countries more severely affected by the PYVV epidemic, and whose geography allows the study of the spatial association between the vector and the disease epidemic across a wide altitude range. The geographical clusters where the PYVV epidemic is concentrated, as well as those of farms affected by the GWF were identified using a novel spatial epidemiology approach. The influence of altitude range on the association between PYVV and T. vaporarioum was also assessed. We found a relatively poor spatial association between PYVV epidemic and the presence of the GWF, especially at altitudes above 3,000 m above mean sea level. Furthermore, GWF populations could only explain a small fraction of the extent of the PYVV epidemic in Colombia. Movement of infected seed tubers might be the main mechanism of dispersion, and could be a key driver for the PYVV infection among potato crops. Agricultural policies focused on improving quality of seed tubers and their appropriate distribution could be the most efficient control intervention against PYVV dispersion.

  2. Epidemiologic relationship between Toscana virus infection and Leishmania infantum due to common exposure to Phlebotomus perniciosus sandfly vector.

    Science.gov (United States)

    Bichaud, Laurence; Souris, Marc; Mary, Charles; Ninove, Laëtitia; Thirion, Laurence; Piarroux, Raphaël P; Piarroux, Renaud; De Lamballerie, Xavier; Charrel, Rémi N

    2011-09-01

    Sand flies are recognised vectors of parasites in the genus Leishmania and a number of arthropod-borne viruses, in particular viruses within the genus Phlebovirus, family Bunyaviridae. In southern France, Toscana phlebovirus (TOSV) is recognized as a prominent cause of summer meningitis. Since Leishmania and TOSV have a common vector (Phlebotomus perniciosus), an epidemiologic link has been assumed for a long time. However, there is no scientific evidence of such a link between human leishmaniosis and phleboviral infections. To identify a possible link, we investigated the presence and distribution of antibodies against these two microorganisms (i) in individuals and (ii) at a spatial level in the city of Marseille (south-eastern France). Five hundred sera were selected randomly in the biobank of the Department of Parasitology of the Public Hospitals of Marseille. All sera were previously tested for IgG against Leishmania by Western Blotting, and TOSV IgG were detected by indirect immunofluorescence. The seropositivity rates were 21.4% for TOSV and 28% for Leishmania. Statistical analysis demonstrated that seropositivity for one pathogen was significantly associated with seropositivity to the other pathogen. This result provided the first robust evidence for the existence of an epidemiological relationship between Leishmania infantum and TOSV. Addresses of tested patients were geolocalized and integrated into Geographical Information System software, in order to test spatial relationship between the two pathogens. Spatial analysis did not allow to identify (i) specific patterns for the spatial distribution of positive serological results for TOSV or Leishmania, and (ii) a spatial relationship between Leishmania and TOSV positive serological results. This may reflect the fact that the sample studied was not powerful enough to demonstrate either a spatial clustering or co-location, i.e. that the actual risk exposure area is smaller than the mean of distance between

  3. Epidemiologic relationship between Toscana virus infection and Leishmania infantum due to common exposure to Phlebotomus perniciosus sandfly vector.

    Directory of Open Access Journals (Sweden)

    Laurence Bichaud

    2011-09-01

    Full Text Available Sand flies are recognised vectors of parasites in the genus Leishmania and a number of arthropod-borne viruses, in particular viruses within the genus Phlebovirus, family Bunyaviridae. In southern France, Toscana phlebovirus (TOSV is recognized as a prominent cause of summer meningitis. Since Leishmania and TOSV have a common vector (Phlebotomus perniciosus, an epidemiologic link has been assumed for a long time. However, there is no scientific evidence of such a link between human leishmaniosis and phleboviral infections. To identify a possible link, we investigated the presence and distribution of antibodies against these two microorganisms (i in individuals and (ii at a spatial level in the city of Marseille (south-eastern France. Five hundred sera were selected randomly in the biobank of the Department of Parasitology of the Public Hospitals of Marseille. All sera were previously tested for IgG against Leishmania by Western Blotting, and TOSV IgG were detected by indirect immunofluorescence. The seropositivity rates were 21.4% for TOSV and 28% for Leishmania. Statistical analysis demonstrated that seropositivity for one pathogen was significantly associated with seropositivity to the other pathogen. This result provided the first robust evidence for the existence of an epidemiological relationship between Leishmania infantum and TOSV. Addresses of tested patients were geolocalized and integrated into Geographical Information System software, in order to test spatial relationship between the two pathogens. Spatial analysis did not allow to identify (i specific patterns for the spatial distribution of positive serological results for TOSV or Leishmania, and (ii a spatial relationship between Leishmania and TOSV positive serological results. This may reflect the fact that the sample studied was not powerful enough to demonstrate either a spatial clustering or co-location, i.e. that the actual risk exposure area is smaller than the mean of

  4. Recombinant Newcastle disease virus (NDV) with inserted gene coding for GM-CSF as a new vector for cancer immunogene therapy

    NARCIS (Netherlands)

    Janke, M.; Peeters, B.P.H.; Leeuw, de O.S.; Moormann, R.J.M.; Arnold, A.; Fournier, P.; Schirrmacher, V.

    2007-01-01

    This is the first report describing recombinant (rec) Newcastle disease virus (NDV) as vector for gene therapy of cancer. The gene encoding granulocyte/macrophage colony-stimulating factor (GM-CSF) was inserted as an additional transcription unit at two different positions into the NDV genome. The

  5. Have we found an optimal insertion site in a Newcastle disease virus vector to express a foreign gene for vaccine and gene therapy purposes?

    Science.gov (United States)

    Using reverse genetics technology, many strains of Newcastle disease virus (NDV) have been developed as vectors to express foreign genes for vaccine and gene therapy purposes. The foreign gene is usually inserted into a non-coding region of the NDV genome as an independent transcription unit. Eval...

  6. Replication-defective recombinant Semliki Forest virus encoding GM-CSF as a vector system for rapid and facile generation of autologous human tumor cell vaccines

    NARCIS (Netherlands)

    Withoff, S; Glazenburg, KL; van Veen, ML; Kraak, MMJ; Hospers, GAP; Storkel, S; de Vries, EGE; Wischut, J; Daemen, T

    2001-01-01

    This paper describes the production of recombinant Semliki Forest virus encoding murine or human granulocyte-macrophage colony-stimulating factor (GM-CSF) and the capacity of these vectors to transduce murine and human tumor cells ex vivo. High-titer stocks (up to 3 x 10(9) particles/ml) of

  7. Augmentation of alphavirus vector-induced human papilloma virus-specific immune and anti-tumour responses by co-expression of interleukin-12

    NARCIS (Netherlands)

    Riezebos-Brilman, Annelies; Regts, Joke; Chen, Margaret; Wilschut, Jan; Daemen, Toos

    2009-01-01

    To enhance the efficacy of a therapeutic immunisition strategy against human papillomavirus-induced cervical cancer we evaluated the adjuvant effect of interleukin-12 (IL12) expressed by a Semliki Forest virus vector (SFV) in mice. Depending on the dose and schedule. SFV-IL12 Stimulated

  8. Multiple efficacy studies of an adenovirus-vectored foot-and-mouth disease virus serotype A24 subunit vaccine in cattle using direct homologous challenge

    Science.gov (United States)

    The safety and efficacy of an experimental, replication-deficient, human adenovirus-vectored foot-and-mouth disease virus (FMDV) serotype A24 Cruzeiro capsid-based subunit vaccine (AdtA24) was examined in eight independent cattle studies. AdtA24 non-adjuvanted vaccine was administered intramuscularl...

  9. Efficient Transduction of Vascular Endothelial Cells with Recombinant Adeno-Associated Virus Serotype 1 and 5 Vectors

    Science.gov (United States)

    CHEN, SIFENG; KAPTURCZAK, MATTHIAS; LOILER, SCOTT A.; ZOLOTUKHIN, SERGEI; GLUSHAKOVA, OLENA Y.; MADSEN, KIRSTEN M.; SAMULSKI, RICHARD J.; HAUSWIRTH, WILLIAM W.; CAMPBELL-THOMPSON, MARTHA; BERNS, KENNETH I.; FLOTTE, TERENCE R.; ATKINSON, MARK A.; TISHER, C. CRAIG

    2006-01-01

    Recombinant adeno-associated virus (rAAV) has become an attractive tool for gene therapy because of its ability to transduce both dividing and nondividing cells, elicit a limited immune response, and the capacity for imparting long-term transgene expression. Previous studies have utilized rAAV serotype 2 predominantly and found that transduction of vascular cells is relatively inefficient. The purpose of the present study was to evaluate the transduction efficiency of rAAV serotypes 1 through 5 in human and rat aortic endothelial cells (HAEC and RAEC). rAAV vectors with AAV2 inverted terminal repeats containing the human α1-antitrypsin (hAAT) gene were transcapsidated using helper plasmids to provide viral capsids for the AAV1 through 5 serotypes. True type rAAV2 and 5 vectors encoding β-galactosidase or green fluorescence protein were also studied. Infection with rAAV1 resulted in the most efficient transduction in both HAEC and RAEC compared to other serotypes (p < 0.001) at 7 days posttransduction. Interestingly, expression was increased in cells transduced with rAAV5 to levels surpassing rAAV1 by day 14 and 21. Transduction with rAAV1 was completely inhibited by removal of sialic acid with sialidase, while heparin had no effect. These studies are the first demonstration that sialic acid residues are required for rAAV1 transduction in endothelial cells. Transduction of rat aortic segments ex vivo and in vivo demonstrated significant transgene expression in endothelial and smooth muscle cells with rAAV1 and 5 serotype vectors, in comparison to rAAV2. These results suggest the unique potential of rAAV1 and rAAV5-based vectors for vascular-targeted gene-based therapeutic strategies. OVERVIEW SUMMARY Gene delivery to the vasculature has significant potential as a therapeutic strategy for several cardiovascular disorders including atherosclerosis, hypertension, angiogenesis, and chronic vascular rejection of transplanted organs. However, limited advances have been

  10. Adeno-associated virus vector-mediated IL-10 gene delivery prevents type 1 diabetes in NOD mice

    Science.gov (United States)

    Goudy, Kevin; Song, Sihong; Wasserfall, Clive; Zhang, Y. Clare; Kapturczak, Matthias; Muir, Andrew; Powers, Matthew; Scott-Jorgensen, Marda; Campbell-Thompson, Martha; Crawford, James M.; Ellis, Tamir M.; Flotte, Terence R.; Atkinson, Mark A.

    2001-01-01

    The development of spontaneous autoimmune diabetes in nonobese diabetic (NOD) mice provides for their use as a model of human type 1 diabetes. To test the feasibility of muscle-directed gene therapy to prevent type 1 diabetes, we developed recombinant adeno-associated virus (rAAV) vectors containing murine cDNAs for immunomodulatory cytokines IL-4 or IL-10. Skeletal muscle transduction of female NOD mice with IL-10, but not IL-4, completely abrogated diabetes. rAAV-IL-10 transduction attenuated the production of insulin autoantibodies, quantitatively reduced pancreatic insulitis, maintained islet insulin content, and altered splenocyte cytokine responses to mitogenic stimulation. The beneficial effects were host specific, as adoptive transfer of splenocytes from rAAV IL-10-treated animals rapidly imparted diabetes in naive hosts, and the cells contained no protective immunomodulatory capacity, as defined through adoptive cotransfer analyses. These results indicate the utility for rAAV, a vector with advantages for therapeutic gene delivery, to transfer immunoregulatory cytokines capable of preventing type 1 diabetes. In addition, these studies provide foundational support for the concept of using immunoregulatory agents delivered by rAAV to modulate a variety of disorders associated with deleterious immune responses, including allergic reactions, transplantation rejection, immunodeficiencies, and autoimmune disorders. PMID:11717448

  11. Apple Latent Spherical Virus Vector as Vaccine for the Prevention and Treatment of Mosaic Diseases in Pea, Broad Bean, and Eustoma Plants by Bean Yellow Mosaic Virus

    Directory of Open Access Journals (Sweden)

    Nozomi Satoh

    2014-11-01

    Full Text Available We investigated the protective effects of a viral vector based on an Apple latent spherical virus (ALSV harboring a segment of the Bean yellow mosaic virus (BYMV genome against mosaic diseases in pea, broad bean, and eustoma plants caused by BYMV infection. In pea plants pre-inoculated with the ALSV vaccine and challenge inoculated with BYMV expressing green fluorescence protein, BYMV multiplication occurred in inoculated leaves, but was markedly inhibited in the upper leaves. No mosaic symptoms due to BYMV infection were observed in the challenged plants pre-inoculated with the ALSV vaccine. Simultaneous inoculation with the ALSV vaccine and BYMV also prevented mosaic symptoms in broad bean and eustoma plants, and BYMV accumulation was strongly inhibited in the upper leaves of plants treated with the ALSV vaccine. Pea and eustoma plants were pre-inoculated with BYMV followed by inoculation with the ALSV vaccine to investigate the curative effects of the ALSV vaccine. In both plant species, recovery from mosaic symptoms was observed in upper leaves and BYMV accumulation was inhibited in leaves developing post-ALSV vaccination. These results show that ALSV vaccination not only prevents mosaic diseases in pea, broad bean, and eustoma, but that it is also effective in curing these diseases.

  12. Modeled response of the West Nile virus vector Culex quinquefasciatus to changing climate using the dynamic mosquito simulation model

    Science.gov (United States)

    Morin, Cory W.; Comrie, Andrew C.

    2010-09-01

    Climate can strongly influence the population dynamics of disease vectors and is consequently a key component of disease ecology. Future climate change and variability may alter the location and seasonality of many disease vectors, possibly increasing the risk of disease transmission to humans. The mosquito species Culex quinquefasciatus is a concern across the southern United States because of its role as a West Nile virus vector and its affinity for urban environments. Using established relationships between atmospheric variables (temperature and precipitation) and mosquito development, we have created the Dynamic Mosquito Simulation Model (DyMSiM) to simulate Cx. quinquefasciatus population dynamics. The model is driven with climate data and validated against mosquito count data from Pasco County, Florida and Coachella Valley, California. Using 1-week and 2-week filters, mosquito trap data are reproduced well by the model ( P < 0.0001). Dry environments in southern California produce different mosquito population trends than moist locations in Florida. Florida and California mosquito populations are generally temperature-limited in winter. In California, locations are water-limited through much of the year. Using future climate projection data generated by the National Center for Atmospheric Research CCSM3 general circulation model, we applied temperature and precipitation offsets to the climate data at each location to evaluate mosquito population sensitivity to possible future climate conditions. We found that temperature and precipitation shifts act interdependently to cause remarkable changes in modeled mosquito population dynamics. Impacts include a summer population decline from drying in California due to loss of immature mosquito habitats, and in Florida a decrease in late-season mosquito populations due to drier late summer conditions.

  13. Forced recombination of psi-modified murine leukaemia virus-based vectors with murine leukaemia-like and VL30 murine endogenous retroviruses

    DEFF Research Database (Denmark)

    Mikkelsen, J G; Lund, Anders Henrik; Duch, M

    1999-01-01

    -impaired Akv-MLV-derived vectors, we here examine putative genetic interactions between vector RNAs and copackaged endogenous retroviral RNAs of the murine leukaemia virus (MLV) and VL30 retroelement families. We show (i) that MLV recombination is not blocked by nonhomology within the 5' untranslated region....... We note that recombination-based rescue of primer binding site knock-out retroviral vectors may constitute a sensitive assay to register putative genetic interactions involving endogenous retroviral RNAs present in cells of various species.......Co-encapsidation of retroviral RNAs into virus particles allows for the generation of recombinant proviruses through events of template switching during reverse transcription. By use of a forced recombination system based on recombinational rescue of replication- defective primer binding site...

  14. Activation of the cellular unfolded protein response by recombinant adeno-associated virus vectors.

    Directory of Open Access Journals (Sweden)

    Balaji Balakrishnan

    Full Text Available The unfolded protein response (UPR is a stress-induced cyto-protective mechanism elicited towards an influx of large amount of proteins in the endoplasmic reticulum (ER. In the present study, we evaluated if AAV manipulates the UPR pathways during its infection. We first examined the role of the three major UPR axes, namely, endoribonuclease inositol-requiring enzyme-1 (IRE1α, activating transcription factor 6 (ATF6 and PKR-like ER kinase (PERK in AAV infected cells. Total RNA from mock or AAV infected HeLa cells were used to determine the levels of 8 different ER-stress responsive transcripts from these pathways. We observed a significant up-regulation of IRE1α (up to 11 fold and PERK (up to 8 fold genes 12-48 hours after infection with self-complementary (scAAV2 but less prominent with single-stranded (ssAAV2 vectors. Further studies demonstrated that scAAV1 and scAAV6 also induce cellular UPR in vitro, with AAV1 vectors activating the PERK pathway (3 fold while AAV6 vectors induced a significant increase on all the three major UPR pathways [6-16 fold]. These data suggest that the type and strength of UPR activation is dependent on the viral capsid. We then examined if transient inhibition of UPR pathways by RNA interference has an effect on AAV transduction. siRNA mediated silencing of PERK and IRE1α had a modest effect on AAV2 and AAV6 mediated gene expression (∼1.5-2 fold in vitro. Furthermore, hepatic gene transfer of scAAV2 vectors in vivo, strongly elevated IRE1α and PERK pathways (2 and 3.5 fold, respectively. However, when animals were pre-treated with a pharmacological UPR inhibitor (metformin during scAAV2 gene transfer, the UPR signalling and its subsequent inflammatory response was attenuated concomitant to a modest 2.8 fold increase in transgene expression. Collectively, these data suggest that AAV vectors activate the cellular UPR pathways and their selective inhibition may be beneficial during AAV mediated gene transfer.

  15. Tomato Infection by Whitefly-Transmitted Circulative and Non-Circulative Viruses Induce Contrasting Changes in Plant Volatiles and Vector Behaviour.

    Science.gov (United States)

    Fereres, Alberto; Peñaflor, Maria Fernanda G V; Favaro, Carla F; Azevedo, Kamila E X; Landi, Carolina H; Maluta, Nathalie K P; Bento, José Mauricio S; Lopes, Joao R S

    2016-08-11

    Virus infection frequently modifies plant phenotypes, leading to changes in behaviour and performance of their insect vectors in a way that transmission is enhanced, although this may not always be the case. Here, we investigated Bemisia tabaci response to tomato plants infected by Tomato chlorosis virus (ToCV), a non-circulative-transmitted crinivirus, and Tomato severe rugose virus (ToSRV), a circulative-transmitted begomovirus. Moreover, we examined the role of visual and olfactory cues in host plant selection by both viruliferous and non-viruliferous B. tabaci. Visual cues alone were assessed as targets for whitefly landing by placing leaves underneath a Plexiglas plate. A dual-choice arena was used to assess whitefly response to virus-infected and mock-inoculated tomato leaves under light and dark conditions. Thereafter, we tested the whitefly response to volatiles using an active air-flow Y-tube olfactometer, and chemically characterized the blends using gas chromatography coupled to mass spectrometry. Visual stimuli tests showed that whiteflies, irrespective of their infectious status, always preferred to land on virus-infected rather than on mock-inoculated leaves. Furthermore, whiteflies had no preference for either virus-infected or mock-inoculated leaves under dark conditions, but preferred virus-infected leaves in the presence of light. ToSRV-infection promoted a sharp decline in the concentration of some tomato volatiles, while an increase in the emission of some terpenes after ToCV infection was found. ToSRV-viruliferous whiteflies preferred volatiles emitted from mock-inoculated plants, a conducive behaviour to enhance virus spread, while volatiles from ToCV-infected plants were avoided by non-viruliferous whiteflies, a behaviour that is likely detrimental to the secondary spread of the virus. In conclusion, the circulative persistent begomovirus, ToSRV, seems to have evolved together with its vector B. tabaci to optimise its own spread. However

  16. Insecticidal Effects on the Spatial Progression of Tomato Yellow Leaf Curl Virus and Movement of Its Whitefly Vector in Tomato.

    Science.gov (United States)

    Dempsey, M; Riley, D G; Srinivasan, R

    2017-06-01

    Commercial management of whitefly-transmitted Tomato yellow leaf curl virus (TYLCV) typically relies on insecticide control of whitefly vectors as a first line of defense. We quantified this effect in crop tunnel studies, with validation in a tomato field setting. Tomato yellow leaf curl virus-infected and Bemisia tabaci (Gennadius)-infested source plants were planted at the beginning of tunneled rows to serve as inoculum source, so that movement of whiteflies and TYLCV symptoms could be tracked down the length of the tunnel over time. Tunnel study results showed that proximity to the source plant was a more important factor than insecticide treatments. Insecticide-treated tomato transplants did tend to suppress whitefly incidence and slowed TYLCV movement in comparison with the untreated check; however, tomato plants planted closer to the source plant had higher incidence of whiteflies and TYLCV infection, regardless of treatment. In a large tomato plot study with a controlled inoculum source, insecticide treatments significantly reduced the spread of TYLCV. When uninhibited by insecticide treatment, 80% of the TYLCV spread was restricted to <15 m from the source plant (<11 m in the validation study), with insecticide treatment generally reducing the distance and magnitude of this spread. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  17. Sindbis Virus-Pseudotyped Lentiviral Vectors Carrying VEGFR2-Specific Nanobody for Potential Transductional Targeting of Tumor Vasculature.

    Science.gov (United States)

    Ahani, Roshank; Roohvand, Farzin; Cohan, Reza Ahangari; Etemadzadeh, Mohammad Hossein; Mohajel, Nasir; Behdani, Mahdi; Shahosseini, Zahra; Madani, Navid; Azadmanesh, Kayhan

    2016-11-01

    Introduction of selectivity/specificity into viral-based gene delivery systems, such as lentiviral vectors (LVs), is crucial in their systemic administration for cancer gene therapy. The pivotal role of tumor-associated endothelial cells (TAECs) in tumor angiogenesis and overexpression of vascular endothelial growth factor receptor-2 (VEGFR2 or KDR) in TAECs makes them a potent target in cancer treatment. Herein, we report the development of VEGFR2-targeted LVs pseudotyped with chimeric sindbis virus E2 glycoprotein (cSVE2s). For this purpose, either sequence of a VEGFR2-specific nanobody or its natural ligand (VEGF 121 ) was inserted into the binding site of sindbis virus E2 glycoprotein. In silico modeling data suggested that the inserted targeting motifs were exposed in the context of cSVE2s. Western blot analysis of LVs indicated the incorporation of cSVE2s into viral particles. Capture ELISA demonstrated the specificity/functionality of the incorporated cSVE2s. Transduction of 293/KDR (expressing VEGFR2) or 293T cells (negative control) by constructed LVs followed by fluorescent microscopy and flow cytometric analyses indicated selective transduction of 293/KDR cells (30 %) by both targeting motifs compared to 293T control cells (1-2 %). These results implied similar targeting properties of VEGFR2-specific nanobody compared to the VEGF 121 and indicated the potential for transductional targeting of tumor vasculature by the nanobody displaying LVs.

  18. Mouse Mammary Tumor Virus Promoter-Containing Retroviral Promoter Conversion Vectors for Gene-Directed Enzyme Prodrug Therapy are Functional in Vitro and in Vivo

    Directory of Open Access Journals (Sweden)

    Reinhard Klein

    2008-01-01

    Full Text Available Gene directed-enzyme prodrug therapy (GDEPT is an approach for sensitization of tumor cells to an enzymatically activated, otherwise nontoxic, prodrug. Cytochrome P450 2B1 (CYP2B1 metabolizes the prodrugs cyclophosphamide (CPA and ifosfamide (IFA to produce the cytotoxic substances phosphoramide mustard and isophosphoramide mustard as well as the byproduct acrolein. We have constructed a retroviral promoter conversion (ProCon vector for breast cancer GDEPT. The vector allows expression of CYP2B1 from the mouse mammary tumor virus (MMTV promoter known to be active in the mammary glands of transgenic animals. It is anticipated to be used for the generation of encapsulated viral vector producing cells which, when placed inside or close to a tumor, will act as suppliers of the therapeutic CYP2B1 protein as well as of the therapeutic vector itself. The generated vector was effectively packaged by virus producing cells and allowed the production of high levels of enzymatically active CYP2B1 in infected cells which sensitized them to killing upon treatment with both IFA and CPA. Determination of the respective IC50 values demonstrated that the effective IFA dose was reduced by sixteen folds. Infection efficiencies in vivo were determined using a reporter gene-bearing vector in a mammary cancer cell-derived xenograft tumor mouse model.

  19. Generation of retroviral particles for the spleen necrosis virus (SNV)-based vector system and their use in transduction of various cell types.

    Science.gov (United States)

    Parveen, Zahida; Mukhtar, Muhammad; Pomerantz, Roger J

    2010-06-01

    Genetically engineered retroviruses are widely used for gene delivery into human cells. A number of investigators have studied spleen necrosis virus (SNV) as a vehicle for gene delivery. Vectors developed from SNV and its closely associated avian reticuloendotheliosis virus strain A (REV-A) can be used for gene transfer into a variety of cells, including primary hematopoietic cells and human brain and post-mitotic neuronal cells that are difficult to transduce with other vector systems. SNV-based vector systems have the advantage of being quite safe, because wild-type SNV is unable to infect human cells and has less preference for integration into transcriptionally active sites or genes. However, the generation of retroviral vectors requires cotransfection of more than one plasmid into a packaging cell line, which is a tedious process. The development of stable packaging cell lines expressing envelope (Env) proteins and the structural proteins Gag-Pol will enhance mass production of retroviral vectors for future gene therapy experiments both in vitro and in vivo. This protocol describes the generation of retroviral particles for the SNV-based vector system. These particles can then be used for transduction of various cell types; as an example, a technique for transduction of post-mitotic neurons is also presented.

  20. Transmission Biology of Rice Stripe Mosaic Virus by an Efficient Insect Vector Recilia dorsalis (Hemiptera: Cicadellidae

    Directory of Open Access Journals (Sweden)

    Xin Yang

    2017-12-01

    Full Text Available Rice stripe mosaic virus (RSMV is a newly discovered species of cytorhabdovirus infecting rice plants that is transmitted by the leafhopper Recilia dorsalis. In this study, the transmission characteristics of RSMV by R. dorsalis were investigated. Under suitable growth conditions for R. dorsalis, the RSMV acquisition rate reached 71.9% in the second-generation population raised on RSMV-infected rice plants. The minimum acquisition and inoculation access periods of R. dorsalis were 3 and 30 min, respectively. The minimum and maximum latent transmission periods of RSMV in R. dorsalis were 6 and 18 d, respectively, and some R. dorsalis intermittently transmitted RSMV at 2–6 d intervals. Our findings revealed that the virus can replicate in the leafhopper body, but is likely not transovarially transmitted to offspring. These transmission characteristics will help guide the formulation of RSMV prevention and control strategies.

  1. Effect of Temperature on the Vector Efficiency of Aedes aegypti for Dengue 2 Virus

    Science.gov (United States)

    1986-06-26

    50% endpoints, em- ploying LLC-MK2 cells. Mosquitoes The Ae. aegypti adults were F, progeny from eggs oviposited by adults collected as larvae in...a low socioeconomic sector of Bangkok. Eggs were hatched, and larvae were reared to adults, according to standard procedures, at 25°C and at 70% to...R. McN., and Burke, D. S., 1982. Evaluation of Toxorhynchites splendens (Diptera: Culici- dae) as a bioassay host for dengue viruses. J. Med

  2. Productive Infection of Human Peripheral Blood Mononuclear Cells by Feline Immunodeficiency Virus: Implications for Vector Development

    OpenAIRE

    Johnston, James; Power, Christopher

    1999-01-01

    Feline immunodeficiency virus (FIV) is a lentivirus causing immune suppression and neurological disease in cats. Like primate lentiviruses, FIV utilizes the chemokine receptor CXCR4 for infection. In addition, FIV gene expression has been demonstrated in immortalized human cell lines. To investigate the extent and mechanism by which FIV infected primary and immortalized human cell lines, we compared the infectivity of two FIV strains, V1CSF and Petaluma, after cell-free infection. FIV genome ...

  3. Allergic contact stomatitis from colophony.

    Science.gov (United States)

    Sharma, Paul R

    2006-09-01

    Colophony is an ubiquitous contact sensitizer which may be present in dental materials, such as periodontal dressings, impression materials, cements, fix adhesives and varnishes. Exposure to a sensitizer in a hypersensitive person may initiate an allergic contact dermatitis/stomatitis. This usually occurs after direct skin/mucosa contact with the sensitizer. This paper reports the case of a colophony hypersensitive male who developed contact stomatitis after dental treatment with a colophony-containing product. Sensitizing colophony is present in Duraphat 2.26%F varnish, a fluoride varnish used all over the world. A case of hypersensitivity to Duraphat 2.26%F varnish is presented in a patient who, at the initial visit, indicated only an allergy to sticking plasters.

  4. Full-genome characterisation of Orungo, Lebombo and Changuinola viruses provides evidence for co-evolution of orbiviruses with their arthropod vectors.

    Directory of Open Access Journals (Sweden)

    Fauziah Mohd Jaafar

    Full Text Available The complete genomes of Orungo virus (ORUV, Lebombo virus (LEBV and Changuinola virus (CGLV were sequenced, confirming that they each encode 11 distinct proteins (VP1-VP7 and NS1-NS4. Phylogenetic analyses of cell-attachment protein 'outer-capsid protein 1' (OC1, show that orbiviruses fall into three large groups, identified as: VP2(OC1, in which OC1 is the 2nd largest protein, including the Culicoides transmitted orbiviruses; VP3(OC1, which includes the mosquito transmitted orbiviruses; and VP4(OC1 which includes the tick transmitted viruses. Differences in the size of OC1 between these groups, places the T2 'subcore-shell protein' as the third largest protein 'VP3(T2' in the first of these groups, but the second largest protein 'VP3(T2' in the other two groups. ORUV, LEBV and CGLV all group with the Culicoides-borne VP2(OC1/VP3(T2 viruses. The G+C content of the ORUV, LEBV and CGLV genomes is also similar to that of the Culicoides-borne, rather than the mosquito-borne, or tick borne orbiviruses. These data suggest that ORUV and LEBV are Culicoides- rather than mosquito-borne. Multiple isolations of CGLV from sand flies suggest that they are its primary vector. OC1 of the insect-borne orbiviruses is approximately twice the size of the equivalent protein of the tick borne viruses. Together with internal sequence similarities, this suggests its origin by duplication (concatermerisation of a smaller OC1 from an ancestral tick-borne orbivirus. Phylogenetic comparisons showing linear relationships between the dates of evolutionary-separation of their vector species, and genetic-distances between tick-, mosquito- or Culicoides-borne virus-groups, provide evidence for co-evolution of the orbiviruses with their arthropod vectors.

  5. Progress with Recombinant Adeno-Associated Virus Vectors for Gene Therapy of Alpha-1 Antitrypsin Deficiency.

    Science.gov (United States)

    Gruntman, Alisha M; Flotte, Terence R

    2015-06-01

    The pathway to a clinical gene therapy product often involves many changes of course and strategy before obtaining successful results. Here we outline the methodologies, both clinical and preclinical, that went into developing a gene therapy approach to the treatment of alpha-1 antitrypsin deficiency lung disease using muscle-targeted recombinant adeno-associated virus. From initial gene construct development in mouse models through multiple rounds of safety and biodistribution studies in rodents, rabbits, and nonhuman primates to ultimate human trials, this review seeks to provide insight into what clinical translation entails and could thereby inform the process for future investigators.

  6. Comparative analysis of simian immunodeficiency virus gag-specific effector and memory CD8+ T cells induced by different adenovirus vectors.

    Science.gov (United States)

    Tan, Wendy G; Jin, Hyun-Tak; West, Erin E; Penaloza-MacMaster, Pablo; Wieland, Andreas; Zilliox, Michael J; McElrath, M Juliana; Barouch, Dan H; Ahmed, Rafi

    2013-02-01

    Adenovirus (Ad) vectors are widely used as experimental vaccines against several infectious diseases, but the magnitude, phenotype, and functionality of CD8(+) T cell responses induced by different adenovirus serotypes have not been compared. To address this question, we have analyzed simian immunodeficiency virus Gag-specific CD8(+) T cell responses in mice following vaccination with Ad5, Ad26, and Ad35. Our results show that although Ad5 is more immunogenic than Ad26 and Ad35, the phenotype, function, and recall potential of memory CD8(+) T cells elicited by these vectors are substantially different. Ad26 and Ad35 vectors generated CD8(+) T cells that display the phenotype and function of long-lived memory T cells, whereas Ad5 vector-elicited CD8(+) T cells are of a more terminally differentiated phenotype. In addition, hepatic memory CD8(+) T cells elicited by Ad26 and Ad35 mounted more robust recall proliferation following secondary challenge than those induced by Ad5. Furthermore, the boosting potential was higher following priming with alternative-serotype Ad vectors than with Ad5 vectors in heterologous prime-boost regimens. Anamnestic CD8(+) T cell responses were further enhanced when the duration between priming and boosting was extended from 30 to 60 days. Our results demonstrate that heterologous prime-boost vaccine regimens with alternative-serotype Ad vectors elicited more functional memory CD8(+) T cells than any of the regimens containing Ad5. In summary, these results suggest that alternative-serotype Ad vectors will prove useful as candidates for vaccine development against human immunodeficiency virus type 1 and other pathogens and also emphasize the importance of a longer rest period between prime and boost for generating optimal CD8(+) T cell immunity.

  7. Personal clothing as a potential vector of respiratory virus transmission in childcare settings.

    Science.gov (United States)

    Gralton, Jan; McLaws, Mary-Louise; Rawlinson, William D

    2015-06-01

    Previous investigations of fomite transmission have focused on the presence of pathogens on inanimate objects in clinical settings. There has been limited investigation of fomite transmission in non-clinical pediatric settings where there is a high prevalence of respiratory virus infections. Over a 5 week period, this study investigated whether the personal clothing of teachers working in childcare centers was contaminated with viral RNA, and potentially could mediate virus transmission. Matched morning and evening clothing and nasal samples were collected for 313 teacher work days (TWDs). Human rhinoviruses (hRV) RNA were detected from samples using real-time PCR. Human rhinovirus RNA was detected in clothing samples on 16 TWDs and in nasal samples on 32 TWDs. There were no TWDs when teachers provided both positive nasal and clothing samples and only three TWDs when hRV persisted on clothing for the entire day. The detection of hRV RNA was significantly predicted by self-recognition of symptomatic illness by the teacher 2 days prior to detection. These findings suggest that teachers' personal clothing in childcare settings is unlikely to facilitate the transmission of hRV. © 2015 Wiley Periodicals, Inc.

  8. Adenovirus vector-based multi-epitope vaccine provides partial protection against H5, H7, and H9 avian influenza viruses

    Science.gov (United States)

    Hassan, Ahmed O.; Amen, Omar; Sayedahmed, Ekramy E.; Vemula, Sai V.; Amoah, Samuel; York, Ian; Gangappa, Shivaprakash; Sambhara, Suryaprakash; Mittal, Suresh K.

    2017-01-01

    The emergence of H5, H7, and H9 avian influenza virus subtypes in humans reveals their pandemic potential. Although human-to-human transmission has been limited, the genetic reassortment of the avian and human/porcine influenza viruses or mutations in some of the genes resulting in virus replication in the upper respiratory tract of humans could generate novel pandemic influenza viruses. Current vaccines do not provide cross protection against antigenically distinct strains of the H5, H7, and H9 influenza viruses. Therefore, newer vaccine approaches are needed to overcome these potential threats. We developed an egg-independent, adenovirus vector-based, multi-epitope (ME) vaccine approach using the relatively conserved immunogenic domains of the H5N1 influenza virus [M2 ectodomain (M2e), hemagglutinin (HA) fusion domain (HFD), T-cell epitope of nucleoprotein (TNP). and HA α-helix domain (HαD)]. Our ME vaccine induced humoral and cell-mediated immune responses and caused a significant reduction in the viral loads in the lungs of vaccinated mice that were challenged with antigenically distinct H5, H7, or H9 avian influenza viruses. These results suggest that our ME vaccine approach provided broad protection against the avian influenza viruses. Further improvement of this vaccine will lead to a pre-pandemic vaccine that may lower morbidity, hinder transmission, and prevent mortality in a pandemic situation before a strain-matched vaccine becomes available. PMID:29023601

  9. Inflammation and Immune Response of Intra-Articular Serotype 2 Adeno-Associated Virus or Adenovirus Vectors in a Large Animal Model

    Directory of Open Access Journals (Sweden)

    Akikazu Ishihara

    2012-01-01

    Full Text Available Intra-articular gene therapy has potential for the treatment of osteoarthritis and rheumatoid arthritis. To quantify in vitro relative gene transduction, equine chondrocytes and synovial cells were treated with adenovirus vectors (Ad, serotype 2 adeno-associated virus vectors (rAAV2, or self-complementary (sc AAV2 vectors carrying green fluorescent protein (GFP. Using 6 horses, bilateral metacarpophalangeal joints were injected with Ad, rAAV2, or scAAV2 vectors carrying GFP genes to assess the in vivo joint inflammation and neutralizing antibody (NAb titer in serum and joint fluid. In vitro, the greater transduction efficiency and sustained gene expression were achieved by scAAV2 compared to rAAV2 in equine chondrocytes and synovial cells. In vivo, AAV2 demonstrated less joint inflammation than Ad, but similar NAb titer. The scAAV2 vectors can induce superior gene transduction than rAAV2 in articular cells, and both rAAV2 and scAAV2 vectors were showed to be safer for intra-articular use than Ad vectors.

  10. The Tomato spotted wilt virus genome is processed differentially in its plant host Arachis hypogaea and its thrips vector Frankliniella fusca

    Directory of Open Access Journals (Sweden)

    Stephen John Fletcher

    2016-09-01

    Full Text Available Thrips-transmitted tospoviruses are economically important viruses affecting a wide range of field and horticultural crops worldwide. Tomato spotted wilt virus is the type member of the Tospovirus genus with a broad host range of more than 900 plant species. Interactions between these viruses and their plant hosts and insect vectors via RNA interference pathways are likely a key determinant of pathogenicity. The current investigation, for the first time, compares biogenesis of small RNAs between the plant host and insect vector in the presence or absence of TSWV. Unique viral small interfering RNA (vsiRNA profiles are evident for Arachis hypogaea (peanut and Frankliniella fusca (thrips vector following infection with TSWV. Differences between vsiRNA profiles for these plant and insect species, such as the relative abundance of 21 nt and 22 nt vsiRNAs and locations of alignment hotspots, reflect the diverse siRNA biosynthesis pathways of their respective kingdoms. The presence of unique vsiRNAs in F. fusca samples indicates that vsiRNA generation takes place within the thrips, and not solely through uptake via feeding on vsiRNAs produced in infected A. hypogaea. The study also shows key vsiRNA profile differences for TSWV among plant families, which are evident in the case of A. hypogaea, a legume, and members of Solanaceae (S. lycopersicum and N. benthamiana. Distinctively, overall small RNA biogenesis in A. hypogaea is markedly affected with an absence of the 24 nt small RNAs in TSWV-infected plants, possibly leading to wide-spread molecular and phenotypic perturbations specific to this species. These findings add significant information on the host-virus-vector interaction in terms of RNAi pathways and may lead to better crop and vector specific control strategies.

  11. THE EFFECT OF AGRO-ECOLOGICAL ZONES ON THE INCIDENCE AND DISTRIBUTION OF APHID VECTORS OF PEPPER VEINAL MOTTLE VIRUS, ON CULTIVATED PEPPER (CAPSICUM ANNUUM L. IN NIGERIA

    Directory of Open Access Journals (Sweden)

    A.A. FAJINMI

    2011-11-01

    Full Text Available The distribution of aphid vectors of Pepper veinal mottle virus (PVMV was studied on cultivated pepper between 2003 and 2005 in the major pepper producing areas of the six agro-ecological zones in Nigeria. The aphids were isolated, identified and their transmission ability determined. Population of aphid vectors increased progressively in all the agro-ecological zones from March at the onset of raining season reaching a peak in August and then declining from September at the on set of the dry season. The Humid forest and Derived Savanna agro-ecological zones recorded highest mean population of aphids / leaf and types of aphid’s species compared with other agro-ecological zones. Six species of aphid, Myzus persicae (Sulzer, Aphis gossipii (Glover, Aphis craccivora (Koch, Aphis spiraecola (Patch, Aphis fabae (Blanchard, and Rhopalosiphum maidis (Fitch, were identified on the pepper fields surveyed in all the agro-ecological zones. There was no significant difference in the occurrence of M. persicae, A. gossipii, A. craccivora, and R. maidis while A. spiraecola and A. fabae that varied in occurrence in all the zones. Ability to transmit PVMV to a healthy pepper plants varied in all the identified aphid species but R. maidis was not able to transmit PVMV from infected to a healthy pepper plant. A more sustainable approach to controlling pepper viruses is by targeting the aphid vectors and preventing the vectors from reaching the crops because the aphid vectors which are virus specific are the major means of transmitting virus to healthy plant.

  12. [Construction and identification of recombinant adeno-associated virus vector harboring fusion gene NT4-Ant-Shepherdin[79-87

    Science.gov (United States)

    Tang, Xiao-Jiang; Ping, Bao-Hua; Pan, Cheng-En; Yang, Guang-Xiao; Wang, Quan-Ying

    2008-12-01

    To construct a recombinant adeno-associated virus vector harboring fusion gene NT4-Ant-Shepherdin[79-87] and investigate Survivin as a anticancer therapeutic target by use of Shepherdin[79-87]. The gene of Ant-Shepherdin[79-87] was obtained by PCR and T-vector method. After inserted in PBV220-NT4 vector and digested with restricted enzyme, The fusion gene of NT4-Ant-Shepherdin[79-87] was sub-cloned into the shuttle plasmid of adeno-associated virus; the products were co-transferred into HEK-293 cell line with helper plasmid pAAV-Ad and adeno-plasmid pFG140. The recombinant adeno-associated virus was produced by homologous recombination of above 3 plasmids in HEK-293 cells and its titer was measured by Dot-blot hybridization. The effect of rAAV-NT4-Ant-Shepherdin[79-87] on A549 cell line was measured by a colorimetric 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay. DNA sequencing results verified that the sequence of Ant-Shepherdin[79-87] was consistent with that we had designed. After transformed E.coli DH5alpha, a fragment of 321 bp was confirmed. High titer of recombinant adeno-associated virus was obtained by homologous recombination in HEK-293 cell lines (3.4x10(13)pfu/L). rAAV-NT4-Ant-Shepherdin[79-87] had strong induce apoptosis effect on A549 cells. The recombinant adeno-associated virus vector encoding fusion gene NT4-Ant-Shepherdin[79-87] is successfully constructed in this experiment by molecular cloning and in vitro recombination techniques, which provided the basis of further research of Survivin for cancer gene therapy.

  13. Membrane-bound SIV envelope trimers are immunogenic in ferrets after intranasal vaccination with a replication-competent canine distemper virus vector.

    Science.gov (United States)

    Zhang, Xinsheng; Wallace, Olivia; Wright, Kevin J; Backer, Martin; Coleman, John W; Koehnke, Rebecca; Frenk, Esther; Domi, Arban; Chiuchiolo, Maria J; DeStefano, Joanne; Narpala, Sandeep; Powell, Rebecca; Morrow, Gavin; Boggiano, Cesar; Zamb, Timothy J; Richter King, C; Parks, Christopher L

    2013-11-01

    We are investigating canine distemper virus (CDV) as a vaccine vector for the delivery of HIV envelope (Env) that closely resembles the native trimeric spike. We selected CDV because it will promote vaccine delivery to lymphoid tissues, and because human exposure is infrequent, reducing potential effects of pre-existing immunity. Using SIV Env as a model, we tested a number of vector and gene insert designs. Vectors containing a gene inserted between the CDV H and L genes, which encoded Env lacking most of its cytoplasmic tail, propagated efficiently in Vero cells, expressed the immunogen on the cell surface, and incorporated the SIV glycoprotein into progeny virus particles. When ferrets were vaccinated intranasally, there were no signs of distress, vector replication was observed in the gut-associated lymphoid tissues, and the animals produced anti-SIV Env antibodies. These data show that live CDV-SIV Env vectors can safely induce anti-Env immune responses following intranasal vaccination. © 2013 Elsevier Inc. All rights reserved.

  14. Transfection of Infectious RNA and DNA/RNA Layered Vectors of Semliki Forest Virus by the Cell-Penetrating Peptide Based Reagent PepFect6

    Science.gov (United States)

    Pärn, Kalle; Viru, Liane; Lehto, Taavi; Oskolkov, Nikita; Langel, Ülo; Merits, Andres

    2013-01-01

    Viral vectors have a wide variety of applications ranging from fundamental studies of viruses to therapeutics. Recombinant viral vectors are usually constructed using methods of reverse genetics to obtain the genetic material of the viral vector. The physicochemical properties of DNA and RNA make them unable to access cells by themselves, and they require assistance to achieve intracellular delivery. Non-viral delivery vectors can be used for this purpose if they enable efficient intracellular delivery without interfering with the viral life cycle. In this report, we utilize Semliki Forest virus (genus alphavirus) based RNA and DNA vectors to study the transfection efficiency of the non-viral cell-penetrating peptide-based delivery vector PepFect6 in comparison with that of the cationic liposome-based Lipofectamine 2000, and assess their impact on viral replication. The optimal conditions for transfection were determined for both reagents. These results demonstrate, for the first time, the ability of PepFect6 to transport large (13-19 kbp) constructs across the cell membrane. Curiously, DNA molecules delivered using the PepFect6 reagent were found to be transported to the cell nucleus approximately 1.5 hours later than DNA molecules delivered using the Lipofectamine 2000 reagent. Finally, although both PepFect6 and Lipofectamine 2000 reagents can be used for alphavirus research, PepFect6 is preferred because it does not induce changes in the normal cellular phenotype and it does not affect the normal replication-infection cycle of viruses in previously transfected cells. PMID:23861978

  15. Novel Strategy to Control Transgene Expression Mediated by a Sendai Virus-Based Vector Using a Nonstructural C Protein and Endogenous MicroRNAs.

    Directory of Open Access Journals (Sweden)

    Masayuki Sano

    Full Text Available Tissue-specific control of gene expression is an invaluable tool for studying various biological processes and medical applications. Efficient regulatory systems have been utilized to control transgene expression in various types of DNA viral or integrating viral vectors. However, existing regulatory systems are difficult to transfer into negative-strand RNA virus vector platforms because of significant differences in their transcriptional machineries. In this study, we developed a novel strategy for regulating transgene expression mediated by a cytoplasmic RNA vector based on a replication-defective and persistent Sendai virus (SeVdp. Because of the capacity of Sendai virus (SeV nonstructural C proteins to specifically inhibit viral RNA synthesis, overexpression of C protein significantly reduced transgene expression mediated by SeVdp vectors. We found that SeV C overexpression concomitantly reduced SeVdp mRNA levels and genomic RNA synthesis. To control C expression, target sequences for an endogenous microRNA were incorporated into the 3' untranslated region of the C genes. Incorporation of target sequences for miR-21 into the SeVdp vector restored transgene expression in HeLa cells by decreasing C expression. Furthermore, the SeVdp vector containing target sequences for let-7a enabled cell-specific control of transgene expression in human fibroblasts and induced pluripotent stem cells. Our findings demonstrate that SeV C can be used as an effective regulator for controlling transgene expression. This strategy will contribute to efficient and less toxic SeVdp-mediated gene transfer in various biological applications.

  16. Protective Efficacy in Sheep of Adenovirus-Vectored Vaccines against Bluetongue Virus Is Associated with Specific T Cell Responses

    Science.gov (United States)

    Martín, Verónica; Pascual, Elena; Avia, Miguel; Peña, Lourdes; Valcárcel, Félix; Sevilla, Noemí

    2015-01-01

    Bluetongue virus (BTV) is an economically important Orbivirus of the Reoviridae family that causes a hemorrhagic disease in ruminants. Its control has been achieved by inactivated-vaccines that have proven to protect against homologous BTV challenge although unable to induce long-term immunity. Therefore, a more efficient control strategy needs to be developed. Recombinant adenovirus vectors are lead vaccine candidates for protection of several diseases, mainly because of their potency to induce potent T cell immunity. Here we report the induction of humoral and T-cell mediated responses able to protect animals against BTV challenge by recombinant replication-defective human adenovirus serotype 5 (Ad5) expressing either VP7, VP2 or NS3 BTV proteins. First we used the IFNAR(-/-) mouse model system to establish a proof of principle, and afterwards we assayed the protective efficacy in sheep, the natural host of BTV. Mice were completely protected against BTV challenge, developing humoral and BTV-specific CD8+- and CD4+-T cell responses by vaccination with the different rAd5. Sheep vaccinated with Ad5-BTV-VP2 and Ad5-BTV-VP7 or only with Ad5-BTV-VP7 and challenged with BTV showed mild disease symptoms and reduced viremia. This partial protection was achieved in the absence of neutralizing antibodies but strong BTV-specific CD8+ T cell responses in those sheep vaccinated with Ad5-BTV-VP7. These data indicate that rAd5 is a suitable vaccine vector to induce T cell immunity during BTV vaccination and provide new data regarding the relevance of T cell responses in protection during BTV infection. PMID:26619062

  17. Tracking the return of Aedes aegypti to Brazil, the major vector of the dengue, chikungunya and Zika viruses.

    Science.gov (United States)

    Kotsakiozi, Panayiota; Gloria-Soria, Andrea; Caccone, Adalgisa; Evans, Benjamin; Schama, Renata; Martins, Ademir Jesus; Powell, Jeffrey R

    2017-07-01

    Aedes aegypti, commonly known as "the yellow fever mosquito", is of great medical concern today primarily as the major vector of dengue, chikungunya and Zika viruses, although yellow fever remains a serious health concern in some regions. The history of Ae. aegypti in Brazil is of particular interest because the country was subjected to a well-documented eradication program during 1940s-1950s. After cessation of the campaign, the mosquito quickly re-established in the early 1970s with several dengue outbreaks reported during the last 30 years. Brazil can be considered the country suffering the most from the yellow fever mosquito, given the high number of dengue, chikungunya and Zika cases reported in the country, after having once been declared "free of Ae. aegypti". We used 12 microsatellite markers to infer the genetic structure of Brazilian Ae. aegypti populations, genetic variability, genetic affinities with neighboring geographic areas, and the timing of their arrival and spread. This enabled us to reconstruct their recent history and evaluate whether the reappearance in Brazil was the result of re-invasion from neighboring non-eradicated areas or re-emergence from local refugia surviving the eradication program. Our results indicate a genetic break separating the northern and southern Brazilian Ae. aegypti populations, with further genetic differentiation within each cluster, especially in southern Brazil. Based on our results, re-invasions from non-eradicated regions are the most likely scenario for the reappearance of Ae. aegypti in Brazil. While populations in the northern cluster are likely to have descended from Venezuela populations as early as the 1970s, southern populations seem to have derived more recently from northern Brazilian areas. Possible entry points are also revealed within both southern and northern clusters that could inform strategies to control and monitor this important arbovirus vector.

  18. Flip-Flop HSV-BAC: bacterial artificial chromosome based system for rapid generation of recombinant herpes simplex virus vectors using two independent site-specific recombinases

    Directory of Open Access Journals (Sweden)

    Todo Tomoki

    2006-09-01

    Full Text Available Abstract Background Oncolytic herpes simplex virus (HSV vectors that specifically replicate in and kill tumor cells sparing normal cells are a promising cancer therapy. Traditionally, recombinant HSV vectors have been generated through homologous recombination between the HSV genome and a recombination plasmid, which usually requires laborious screening or selection and can take several months. Recent advances in bacterial artificial chromosome (BAC technology have enabled cloning of the whole HSV genome as a BAC plasmid and subsequent manipulation in E. coli. Thus, we sought a method to generate recombinant oncolytic HSV vectors more easily and quickly using BAC technology. Results We have developed an HSV-BAC system, termed the Flip-Flop HSV-BAC system, for the rapid generation of oncolytic HSV vectors. This system has the following features: (i two site-specific recombinases, Cre and FLPe, are used sequentially to integrate desired sequences and to excise the BAC sequences, respectively; and (ii the size of the HSV-BAC-insert genome exceeds the packaging limit of HSV so only correctly recombined virus grows efficiently. We applied this to the construction of an HSV-BAC plasmid that can be used for the generation of transcriptionally-targeted HSV vectors. BAC sequences were recombined into the UL39 gene of HSV ICP4-deletion mutant d120 to generate M24-BAC virus, from which HSV-BAC plasmid pM24-BAC was isolated. An ICP4 expression cassette driven by an exogenous promoter was re-introduced to pM24-BAC by Cre-mediated recombination and nearly pure preparations of recombinant virus were obtained typically in two weeks. Insertion of the ICP4 coding sequence alone did not restore viral replication and was only minimally better than an ICP4-null construct, whereas insertion of a CMVIE promoter-ICP4 transgene (bM24-CMV efficiently drove viral replication. The levels of bM24-CMV replication in tumor cells varied considerably compared to hrR3 (UL39

  19. Inter-epidemic abundance and distribution of potential mosquito vectors for Rift Valley fever virus in Ngorongoro district, Tanzania.

    Science.gov (United States)

    Mweya, Clement N; Kimera, Sharadhuli I; Mellau, Lesakit S B; Mboera, Leonard E G

    2015-01-01

    Rift Valley fever (RVF) is a mosquito-borne viral zoonosis that primarily affects ruminants but also has the capacity to infect humans. To determine the abundance and distribution of mosquito vectors in relation to their potential role in the virus transmission and maintenance in disease epidemic areas of Ngorongoro district in northern Tanzania. A cross-sectional entomological investigation was carried out before the suspected RVF outbreak in October 2012. Mosquitoes were sampled both outdoors and indoors using the Centre for Disease Control (CDC) light traps and Mosquito Magnets baited with attractants. Outdoor traps were placed in proximity with breeding sites and under canopy in banana plantations close to the sleeping places of animals. A total of 1,823 mosquitoes were collected, of which 87% (N=1,588) were Culex pipiens complex, 12% (N=226) Aedes aegypti, and 0.5% (N=9) Anopheles species. About two-thirds (67%; N=1,095) of C. pipiens complex and nearly 100% (N=225) of A. aegypti were trapped outdoors using Mosquito Magnets. All Anopheles species were trapped indoors using CDC light traps. There were variations in abundance of C. pipiens complex and A. aegypti among different ecological and vegetation habitats. Over three quarters (78%) of C. pipiens complex and most (85%) of the A. aegypti were trapped in banana and maize farms. Both C. pipiens complex and A. aegypti were more abundant in proximity with cattle and in semi-arid thorn bushes and lower Afro-montane. The highest number of mosquitoes was recorded in villages that were most affected during the RVF epidemic of 2007. Of the tested 150 pools of C. pipiens complex and 45 pools of A. aegypti, none was infected with RVF virus. These results provide insights into unique habitat characterisation relating to mosquito abundances and distribution in RVF epidemic-prone areas of Ngorongoro district in northern Tanzania.

  20. Inter-epidemic abundance and distribution of potential mosquito vectors for Rift Valley fever virus in Ngorongoro district, Tanzania

    Directory of Open Access Journals (Sweden)

    Clement N. Mweya

    2015-01-01

    Full Text Available Background: Rift Valley fever (RVF is a mosquito-borne viral zoonosis that primarily affects ruminants but also has the capacity to infect humans. Objective: To determine the abundance and distribution of mosquito vectors in relation to their potential role in the virus transmission and maintenance in disease epidemic areas of Ngorongoro district in northern Tanzania. Methods: A cross-sectional entomological investigation was carried out before the suspected RVF outbreak in October 2012. Mosquitoes were sampled both outdoors and indoors using the Centre for Disease Control (CDC light traps and Mosquito Magnets baited with attractants. Outdoor traps were placed in proximity with breeding sites and under canopy in banana plantations close to the sleeping places of animals. Results: A total of 1,823 mosquitoes were collected, of which 87% (N=1,588 were Culex pipiens complex, 12% (N=226 Aedes aegypti, and 0.5% (N=9 Anopheles species. About two-thirds (67%; N=1,095 of C. pipiens complex and nearly 100% (N=225 of A. aegypti were trapped outdoors using Mosquito Magnets. All Anopheles species were trapped indoors using CDC light traps. There were variations in abundance of C. pipiens complex and A. aegypti among different ecological and vegetation habitats. Over three quarters (78% of C. pipiens complex and most (85% of the A. aegypti were trapped in banana and maize farms. Both C. pipiens complex and A. aegypti were more abundant in proximity with cattle and in semi-arid thorn bushes and lower Afro-montane. The highest number of mosquitoes was recorded in villages that were most affected during the RVF epidemic of 2007. Of the tested 150 pools of C. pipiens complex and 45 pools of A. aegypti, none was infected with RVF virus. Conclusions: These results provide insights into unique habitat characterisation relating to mosquito abundances and distribution in RVF epidemic-prone areas of Ngorongoro district in northern Tanzania.

  1. Prevention and Control Strategies to Counter Zika Virus, a Special Focus on Intervention Approaches against Vector Mosquitoes-Current Updates.

    Science.gov (United States)

    Singh, Raj K; Dhama, Kuldeep; Khandia, Rekha; Munjal, Ashok; Karthik, Kumaragurubaran; Tiwari, Ruchi; Chakraborty, Sandip; Malik, Yashpal S; Bueno-Marí, Rubén

    2018-01-01

    Zika virus (ZIKV) is the most recent intruder that acquired the status of global threat creating panic and frightening situation to public owing to its rapid spread, attaining higher virulence and causing complex clinical manifestations including microcephaly in newborns and Guillain Barré Syndrome. Alike other flaviviruses, the principal mode of ZIKV transmission is by mosquitoes. Advances in research have provided reliable diagnostics for detecting ZIKV infection, while several drug/therapeutic targets and vaccine candidates have been identified recently. Despite these progresses, currently there is neither any effective drug nor any vaccine available against ZIKV. Under such circumstances and to tackle the problem at large, control measures of which mosquito population control need to be strengthened following appropriate mechanical, chemical, biological and genetic control measures. Apart from this, several other known modes of ZIKV transmission which have gained importance in recent past such as intrauterine, sexual intercourse, and blood-borne spread need to be checked and kept under control by adopting appropriate precautions and utmost care during sexual intercourse, blood transfusion and organ transplantation. The virus inactivation by pasteurization, detergents, chemicals, and filtration can effectively reduce viral load in plasma-derived medicinal products. Added to this, strengthening of the surveillance and monitoring of ZIKV as well as avoiding travel to Zika infected areas would aid in keeping viral infection under check. Here, we discuss the salient advances in the prevention and control strategies to combat ZIKV with a focus on highlighting various intervention approaches against the vector mosquitoes of this viral pathogen along with presenting an overview regarding human intervention measures to counter other modes of ZIKV transmission and spread. Additionally, owing to the success of vaccines for a number of infections globally, a separate

  2. Phenotyping of VIGS-mediated gene silencing in rice using a vector derived from a DNA virus.

    Science.gov (United States)

    Kant, Ravi; Dasgupta, Indranil

    2017-07-01

    Target genes in rice can be optimally silenced if inserted in antisense or hairpin orientation in the RTBV-derived VIGS vector and plants grown at 28 °C and 80% humidity after inoculation. Virus induced gene silencing (VIGS) is a method used to transiently silence genes in dicot as well as monocot plants. For the important monocot species rice, the Rice tungro bacilliform virus (RTBV)-derived VIGS system (RTBV-VIGS), which uses agroinoculation to initiate silencing, has not been standardized for optimal use. Here, using RTBV-VIGS, three sets of conditions were tested to achieve optimal silencing of the rice marker gene phytoene desaturase (pds). The effect of orientation of the insert in the RTBV-VIGS plasmid (sense, antisense and hairpin) on the silencing of the target gene was then evaluated using rice magnesium chelatase subunit H (chlH). Finally, the rice Xa21 gene, conferring resistance against bacterial leaf blight disease (BLB) was silenced using RTBV-VIGS system. In each case, real-time PCR-based assessment indicated approximately 40-80% fall in the accumulation levels of the transcripts of pds, chlH and Xa21. In the case of pds, the appearance of white streaks in the emerging leaves, and for chlH, chlorophyll levels and F v /F m ratio were assessed as phenotypes for silencing. For Xa21, the resistance levels to BLB were assessed by measuring the lesion length and the percent diseased areas of leaves, following challenge inoculation with Xanthomonas oryzae. In each case, the RTBV-MVIGS system gave rise to a discernible phenotype indicating the silencing of the respective target gene using condition III (temperature 28 °C, humidity 80% and 1 mM MES and 20 µM acetosyringone in secondary agrobacterium culture), which revealed the robustness of this gene silencing system for rice.

  3. Highly efficient retrograde gene transfer into motor neurons by a lentiviral vector pseudotyped with fusion glycoprotein.

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    Miyabi Hirano

    Full Text Available The development of gene therapy techniques to introduce transgenes that promote neuronal survival and protection provides effective therapeutic approaches for neurological and neurodegenerative diseases. Intramuscular injection of adenoviral and adeno-associated viral vectors, as well as lentiviral vectors pseudotyped with rabies virus glycoprotein (RV-G, permits gene delivery into motor neurons in animal models for motor neuron diseases. Recently, we developed a vector with highly efficient retrograde gene transfer (HiRet by pseudotyping a human immunodeficiency virus type 1 (HIV-1-based vector with fusion glycoprotein B type (FuG-B or a variant of FuG-B (FuG-B2, in which the cytoplasmic domain of RV-G was replaced by the corresponding part of vesicular stomatitis virus glycoprotein (VSV-G. We have also developed another vector showing neuron-specific retrograde gene transfer (NeuRet with fusion glycoprotein C type, in which the short C-terminal segment of the extracellular domain and transmembrane/cytoplasmic domains of RV-G was substituted with the corresponding regions of VSV-G. These two vectors afford the high efficiency of retrograde gene transfer into different neuronal populations in the brain. Here we investigated the efficiency of the HiRet (with FuG-B2 and NeuRet vectors for retrograde gene transfer into motor neurons in the spinal cord and hindbrain in mice after intramuscular injection and compared it with the efficiency of the RV-G pseudotype of the HIV-1-based vector. The main highlight of our results is that the HiRet vector shows the most efficient retrograde gene transfer into both spinal cord and hindbrain motor neurons, offering its promising use as a gene therapeutic approach for the treatment of motor neuron diseases.

  4. Asymmetric effects of native and exotic invasive shrubs on ecology of the West Nile virus vector Culex pipiens (Diptera: Culicidae).

    Science.gov (United States)

    Gardner, Allison M; Allan, Brian F; Frisbie, Lauren A; Muturi, Ephantus J

    2015-06-16

    Exotic invasive plants alter the structure and function of native ecosystems and may influence the distribution and abundance of arthropod disease vectors by modifying habitat quality. This study investigated how invasive plants alter the ecology of Culex pipiens, an important vector of West Nile virus (WNV) in northeastern and midwestern regions of the United States. Field and laboratory experiments were conducted to test the hypothesis that three native leaf species (Rubus allegheniensis, blackberry; Sambucus canadensis, elderberry; and Amelanchier laevis, serviceberry), and three exotic invasive leaf species (Lonicera maackii, Amur honeysuckle; Elaeagnus umbellata, autumn olive; and Rosa multiflora, multiflora rose) alter Cx. pipiens oviposition site selection, emergence rates, development time, and adult body size. The relative abundance of seven bacterial phyla in infusions of the six leaf species also was determined using quantitative real-time polymerase chain reaction to test the hypothesis that variation in emergence, development, and oviposition site selection is correlated to differences in the diversity and abundance of bacteria associated with different leaf species, important determinants of nutrient quality and availability for mosquito larvae. Leaf detritus from invasive honeysuckle and autumn olive yielded significantly higher adult emergence rates compared to detritus from the remaining leaf species and honeysuckle alleviated the negative effects of intraspecific competition on adult emergence. Conversely, leaves of native blackberry acted as an ecological trap, generating high oviposition but low emergence rates. Variation in bacterial flora associated with different leaf species may explain this asymmetrical production of mosquitoes: emergence rates and oviposition rates were positively correlated to bacterial abundance and diversity, respectively. We conclude that the displacement of native understory plant species by certain invasive shrubs

  5. A preferred region for recombinational patch repair in the 5' untranslated region of primer binding site-impaired murine leukemia virus vectors

    DEFF Research Database (Denmark)

    Mikkelsen, J G; Lund, Anders Henrik; Kristensen, K D

    1996-01-01

    Transduction of primer binding site-impaired Akv murine leukemia virus-based retroviral vectors from the murine packaging cell lines psi-2 and omega E was studied. The efficiency of transduction of the neo marker of all mutated constructs was found to decrease by 5 to 6 orders of magnitude compared......, suggesting the involvement of a specific endogenous virus-like sequence in patch repair rescue of the primer binding site mutants. The putative recombination partner RNA was found in virions from psi-2 cells as detected by analysis of glutamine tRNA-initiated cDNA and by sequence analysis of regions...

  6. Toxin delivery by the coat protein of an aphid-vectored plant virus provides plant resistance to aphids.

    Science.gov (United States)

    Bonning, Bryony C; Pal, Narinder; Liu, Sijun; Wang, Zhaohui; Sivakumar, S; Dixon, Philip M; King, Glenn F; Miller, W Allen

    2014-01-01

    The sap-sucking insects (order Hemiptera), including aphids, planthoppers, whiteflies and stink bugs, present one of the greatest challenges for pest management in global agriculture. Insect neurotoxins offer an alternative to chemical insecticides for controlling these pests, but require delivery into the insect hemocoel. Here we use the coat protein of a luteovirus, an aphid-vectored plant virus, to deliver a spider-derived, insect-specific toxin that acts within the hemocoel. The luteovirid coat protein is sufficient for delivery of fused proteins into the hemocoel of pea aphids, Acyrthosiphon pisum, without virion assembly. We show that when four aphid pest species-A. pisum, Rhopalosiphum padi, Aphis glycines and Myzus persicae-feed on a recombinant coat protein-toxin fusion, either in an experimental membrane sachet or in transgenic Arabidopsis plants, they experience significant mortality. Aphids fed on these fusion proteins showed signs of neurotoxin-induced paralysis. Luteovirid coat protein-insect neurotoxin fusions represent a promising strategy for transgenic control of aphids and potentially other hemipteran pests.

  7. Enhancement of Antituberculosis Immunity in a Humanized Model System by a Novel Virus-Vectored Respiratory Mucosal Vaccine.

    Science.gov (United States)

    Yao, Yushi; Lai, Rocky; Afkhami, Sam; Haddadi, Siamak; Zganiacz, Anna; Vahedi, Fatemeh; Ashkar, Ali A; Kaushic, Charu; Jeyanathan, Mangalakumari; Xing, Zhou

    2017-07-01

    The translation of preclinically promising novel tuberculosis vaccines to ultimate human applications has been challenged by the lack of animal models with an immune system equivalent to the human immune system in its genetic diversity and level of susceptibility to tuberculosis. We have developed a humanized mice (Hu-mice) tuberculosis model system to investigate the clinical relevance of a novel virus-vectored (VV) tuberculosis vaccine administered via respiratory mucosal or parenteral route. We find that VV vaccine activates T cells in Hu-mice as it does in human vaccinees. The respiratory mucosal route for delivery of VV vaccine in Hu-mice, but not the parenteral route, significantly reduces the humanlike lung tuberculosis outcomes in a human T-cell-dependent manner. Our results suggest that the Hu-mouse can be used to predict the protective efficacy of novel tuberculosis vaccines/strategies before they proceed to large, expensive human trials. This new vaccine testing system will facilitate the global pace of clinical tuberculosis vaccine development. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

  8. Long-term sex-biased correction of circulating propionic acidemia disease markers by adeno-associated virus vectors.

    Science.gov (United States)

    Guenzel, Adam J; Collard, Renata; Kraus, Jan P; Matern, Dietrich; Barry, Michael A

    2015-03-01

    Propionic academia (PA) occurs because of mutations in the PCCA or PCCB genes encoding the two subunits of propionyl-CoA carboxylase, a pivotal enzyme in the breakdown of certain amino acids and odd-chain fatty acids. There is no cure for PA, but dietary protein restriction and liver transplantation can attenuate its symptoms. We show here that a single intravenous injection of adeno-associated virus 2/8 (AAV8) or AAVrh10 expressing PCCA into PA hypomorphic mice decreased systemic propionylcarnitine and methyl citrate for up to 1.5 years. However, long-term phenotypic correction was always better in male mice. AAV-mediated PCCA expression was similar in most tissues in males and females at early time points and differed only in the liver. Over 1.5 years, luciferase and PCCA expression remained elevated in cardiac tissue for both sexes. In contrast, transgene expression in the liver and skeletal muscles of female, but not male, mice waned—suggesting that these tissues were major sinks for systemic phenotypic correction. These data indicate that single systemic intravenous therapy by AAV vectors can mediate long-term phenotype correction for PA. However, tissue-specific loss of expression in females reduces efficacy when compared with males. Whether similar sex-biased AAV effects occur in human gene therapy remains to be determined.

  9. Anthropogenic and meteorological factors influence vector abundance and prevalence of bluetongue virus infection of dairy cattle in California.

    Science.gov (United States)

    Mayo, Christie E; Gardner, Ian A; Mullens, Bradley A; Barker, Christopher M; Gerry, Alec C; Guthrie, Alan J; MacLachlan, N James

    2012-03-23

    Bluetongue is an economically important arboviral disease of ruminants that is transmitted by hematophagous Culicoides midges. In light of dramatic recent changes in the global distribution of bluetongue virus (BTV), the goals of this study were to re-evaluate the prevalence of BTV infection of cattle and abundance of Culicoides midges on individual dairy farms in California. A serosurvey of adult dairy cattle confirmed that BTV infection is prevalent throughout much of the state, although the coastal northwestern region remains free of infection and prevalence varies markedly among farms in the remainder of the state. Intensive sampling for one year of 4 farms in the northern Central Valley of California showed that the abundance of Culicoides midges was markedly different and coincided with the prevalence of BTV infection of sentinel cattle on each farm. Mean maximum and minimum temperatures and other meteorological parameters were similar on all 4 farms, thus we speculate that particular management practices were responsible for both the increased midge abundance and prevalence of BTV infection of cattle at individual farms. Specifically, it is concluded that variation in vector abundance at individual farms most likely is the result of waste-water lagoon and irrigation management practices, leading to higher BTV infection rates among livestock held on farms with more waste-water lagoons and greater acreage of land for waste-water irrigation. Copyright © 2011 Elsevier B.V. All rights reserved.

  10. Ovicidal and Larvicidal Effects of Garlic and Asafoetida Essential Oils Against West Nile Virus Vectors

    Science.gov (United States)

    Muturi, Ephantus J; Ramirez, Jose L; Zilkowski, Bruce; Flor-Weiler, Lina B; Rooney, Alejandro P

    2018-01-01

    Abstract We examined the chemical composition of garlic and asafoetida essential oils and their individual and combined toxicity against larvae of Culex pipiens Linnaeus and Culex restuans Theobald (Diptera: Culicidae). The effect of the two essential oils on egg hatch was also examined. Ten and 12 compounds, respectively, were identified in garlic and asafoetida essential oils. Allyl disulfide (49.13%) and diallyl trisulfide (31.08%) were the most abundant compounds in garlic essential oil accounting for 80.2% of the total oil. In contrast, (E)-sec-butyl propenyl disulfide (30.03%), (Z)-sec-butyl propenyl disulfide (24.32%), and disulfide, methyl 1-(methylthio)propyl (21.87%) were the most abundant compounds in asafoetida essential oil. Allyl disulfide accounted for 7.38% of the total oil in asafoetida essential oil and was one of only three compounds found in both oils. For both mosquito species, garlic essential oil was more toxic than asafoetida essential oil with Cx. restuans (LC50: garlic = 2.7 ppm; asafoetida = 10.1 ppm) being more sensitive than Cx. pipiens (LC50: garlic = 7.5 ppm; asafoetida = 13.5 ppm). When combined, the two essential oils had antagonistic effects. The majority of Culex egg rafts exposed to garlic (73.1%) or asafoetida (55.8%) essential oils failed to hatch and larvae of the few that did hatch mostly died as first instars. Allyl disulfide exhibited strong ovicidal and larvicidal activity suggesting its important contribution to the overall toxicity of the two essential oils. Thus, garlic and asafoetida essential oils are potent mosquito ovicides and larvicides but if used jointly, they could undermine vector control programs.

  11. Predicting distribution of Aedes aegypti and Culex pipiens complex, potential vectors of Rift Valley fever virus in relation to disease epidemics in East Africa

    Directory of Open Access Journals (Sweden)

    Clement Nyamunura Mweya

    2013-10-01

    Full Text Available Background: The East African region has experienced several Rift Valley fever (RVF outbreaks since the 1930s. The objective of this study was to identify distributions of potential disease vectors in relation to disease epidemics. Understanding disease vector potential distributions is a major concern for disease transmission dynamics. Methods: Diverse ecological niche modelling techniques have been developed for this purpose: we present a maximum entropy (Maxent approach for estimating distributions of potential RVF vectors in un-sampled areas in East Africa. We modelled the distribution of two species of mosquitoes (Aedes aegypti and Culex pipiens complex responsible for potential maintenance and amplification of the virus, respectively. Predicted distributions of environmentally suitable areas in East Africa were based on the presence-only occurrence data derived from our entomological study in Ngorongoro District in northern Tanzania. Results: Our model predicted potential suitable areas with high success rates of 90.9% for A. aegypti and 91.6% for C. pipiens complex. Model performance was statistically significantly better than random for both species. Most suitable sites for the two vectors were predicted in central and northwestern Tanzania with previous disease epidemics. Other important risk areas include western Lake Victoria, northern parts of Lake Malawi, and the Rift Valley region of Kenya. Conclusion: Findings from this study show distributions of vectors had biological and epidemiological significance in relation to disease outbreak hotspots, and hence provide guidance for the selection of sampling areas for RVF vectors during inter-epidemic periods.

  12. Predicting distribution of Aedes aegypti and Culex pipiens complex, potential vectors of Rift Valley fever virus in relation to disease epidemics in East Africa.

    Science.gov (United States)

    Mweya, Clement Nyamunura; Kimera, Sharadhuli Iddi; Kija, John Bukombe; Mboera, Leonard E G

    2013-01-01

    The East African region has experienced several Rift Valley fever (RVF) outbreaks since the 1930s. The objective of this study was to identify distributions of potential disease vectors in relation to disease epidemics. Understanding disease vector potential distributions is a major concern for disease transmission dynamics. DIVERSE ECOLOGICAL NICHE MODELLING TECHNIQUES HAVE BEEN DEVELOPED FOR THIS PURPOSE: we present a maximum entropy (Maxent) approach for estimating distributions of potential RVF vectors in un-sampled areas in East Africa. We modelled the distribution of two species of mosquitoes (Aedes aegypti and Culex pipiens complex) responsible for potential maintenance and amplification of the virus, respectively. Predicted distributions of environmentally suitable areas in East Africa were based on the presence-only occurrence data derived from our entomological study in Ngorongoro District in northern Tanzania. Our model predicted potential suitable areas with high success rates of 90.9% for A. aegypti and 91.6% for C. pipiens complex. Model performance was statistically significantly better than random for both species. Most suitable sites for the two vectors were predicted in central and northwestern Tanzania with previous disease epidemics. Other important risk areas include western Lake Victoria, northern parts of Lake Malawi, and the Rift Valley region of Kenya. Findings from this study show distributions of vectors had biological and epidemiological significance in relation to disease outbreak hotspots, and hence provide guidance for the selection of sampling areas for RVF vectors during inter-epidemic periods.

  13. Coordination of stomatal physiological behavior and morphology with carbon dioxide determines stomatal control.

    Science.gov (United States)

    Haworth, Matthew; Killi, Dilek; Materassi, Alessandro; Raschi, Antonio

    2015-05-01

    Stomatal control is determined by the ability to alter stomatal aperture and/or the number of stomata on the surface of new leaves in response to growth conditions. The development of stomatal control mechanisms to the concentration of CO₂within the atmosphere ([CO₂]) is fundamental to our understanding of plant evolutionary history and the prediction of gas exchange responses to future [CO₂]. In a controlled environment, fern and angiosperm species were grown in atmospheres of ambient (400 ppm) and elevated (2000 ppm) [CO₂]. Physiological stomatal behavior was compared with the stomatal morphological response to [CO₂]. An increase in [CO₂] or darkness induced physiological stomatal responses ranging from reductions (active) to no change (passive) in stomatal conductance. Those species with passive stomatal behavior exhibited pronounced reductions of stomatal density in new foliage when grown in elevated [CO₂], whereas species with active stomata showed little morphological response to [CO₂]. Analysis of the physiological and morphological stomatal responses of a wider range of species suggests that patterns of stomatal control to [CO₂] do not follow a phylogenetic pattern associated with plant evolution. Selective pressures may have driven the development of divergent stomatal control strategies to increased [CO₂]. Those species that are able to actively regulate guard cell turgor are more likely to respond to [CO₂] through a change in stomatal aperture than stomatal number. We propose a model of stomatal control strategies in response to [CO₂] characterized by a trade-off between short-term physiological behavior and longer-term morphological response. © 2015 Botanical Society of America, Inc.

  14. Caligus rogercresseyi: posible vector en la transmisión horizontal del virus de la anemia infecciosa del salmón (ISAv

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    Karin Oelckers

    2015-05-01

    Full Text Available Chile el año 2007 se convirtió en el segundo país productor de salmónidos a nivel mundial. Al año siguiente la industria salmonera nacional comenzó a experimentar una severa crisis sanitaria producida por el virus causante de la anemia infecciosa del salmón. Este virus se presentó por primera vez en Noruega (1984, luego en Canadá, Escocia, Islas Faroe, Estados Unidos y Chile (2007. La anemia infecciosa del salmón (ISA, es una enfermedad altamente contagiosa entre los peces, producida por un virus de la familia Orthomyxoviridae. La especie más vulnerable a este virus es el salmón del Atlántico (Salmo salar. La plaga parasitaria producida por el piojo de mar, Caligus rogercresseyi, copépodo ectoparásito, ha ido en aumento lo que favorece el contagio de enfermedades bacterianas y virales. De todas las especies cultivadas en Chile, el salmón del Atlántico, S. salar es una de las especies más susceptibles de ser infestadas por C. rogercresseyi. Durante el 2006, la industria presentó un aumento significativo en las tasas de infestación por Caligus; luego en el 2007, aparecieron brotes del virus ISA. En Noruega, se ha demostrado que el piojo de mar, Lepeophtherius salmonis puede tener un rol como vector en la transmisión del virus ISA, por lo que el objetivo de este trabajo fue determinar si C. rogercresseyi es un vector de transmisión del virus ISA en el salmón del Atlántico, cultivado en el sur de Chile.

  15. Transmissible gastroenteritis virus (TGEV)-based vectors with engineered murine tropism express the rotavirus VP7 protein and immunize mice against rotavirus.

    Science.gov (United States)

    Ribes, Juan Manuel; Ortego, Javier; Ceriani, Juan; Montava, Rebeca; Enjuanes, Luis; Buesa, Javier

    2011-02-05

    A coronavirus vector based on the genome of the porcine transmissible gastroenteritis virus (TGEV) expressing the rotavirus VP7 protein was constructed to immunize and protect against rotavirus infections in a murine model. The tropism of this TGEV-derived vector was modified by replacing the spike S protein with the homologous protein from mouse hepatitis virus (MHV). The rotavirus gene encoding the VP7 protein was cloned into the coronavirus cDNA. BALB/c and STAT1-deficient mice were inoculated with the recombinant viral vector rTGEV(S-MHV)-VP7, which replicates in the intestine and spreads to other organs such as liver, spleen and lungs. TGEV-specific antibodies were detected in all the inoculated BALB/c mice, while rotavirus-specific antibodies were found only after immunization by the intraperitoneal route. Partial protection against rotavirus-induced diarrhea was achieved in suckling BALB/c mice born to dams immunized with the recombinant virus expressing VP7 when they were orally challenged with the homotypic rotavirus strain. Copyright © 2010 Elsevier Inc. All rights reserved.

  16. The impact of temperature and Wolbachia infection on vector competence of potential dengue vectors Aedes aegypti and Aedes albopictus in the transmission of dengue virus serotype 1 in southern Taiwan.

    Science.gov (United States)

    Tsai, Cheng-Hui; Chen, Tien-Huang; Lin, Cheo; Shu, Pei-Yun; Su, Chien-Ling; Teng, Hwa-Jen

    2017-11-07

    We evaluated the impact of temperature and Wolbachia infection on vector competence of the local Aedes aegypti and Ae. albopictus populations of southern Taiwan in the laboratory. After oral infection with dengue serotype 1 virus (DENV-1), female mosquitoes were incubated at temperatures of 10, 16, 22, 28 and 34 °C. Subsequently, salivary gland, head, and thorax-abdomen samples were analyzed for their virus titer at 0, 5, 10, 15, 20, 25 and 30 days post-infection (dpi) by real-time RT-PCR. The results showed that Ae. aegypti survived significantly longer and that dengue viral genome levels in the thorax-abdomen (10 3.25 ± 0.53 -10 4.09 ± 0.71 PFU equivalents/ml) and salivary gland samples (10 2.67 ± 0.33 -10 3.89 ± 0.58 PFU equivalents/ml) were significantly higher at high temperature (28-34 °C). The survival of Ae. albopictus was significantly better at 16 or 28 °C, but the virus titers from thorax-abdomen (10 0.70 -10 2.39 ± 1.31 PFU equivalents/ml) and salivary gland samples (10 0.12 ± 0.05 -10 1.51 ± 0.31 PFU equivalents/ml) were significantly higher at 22-28 °C. Within viable temperature ranges, the viruses were detectable after 10 dpi in salivary glands and head tissues in Ae. aegypti and after 5-10 dpi in Ae. albopictus. Vector competence was measured in Ae. albopictus with and without Wolbachia at 28 °C. Wolbachia-infected mosquitoes survived significantly better and carried lower virus titers than Wolbachia-free mosquitoes. Wolbachia coinfections (92.8-97.2%) with wAlbA and wAlbB strains were commonly found in a wild population of Ae. albopictus. In southern Taiwan, Ae. aegypti is the main vector of dengue and Ae. albopictus has a non-significant role in the transmission of dengue virus due to the high prevalence of Wolbachia infection in the local mosquito population of southern Taiwan.

  17. The impact of temperature and Wolbachia infection on vector competence of potential dengue vectors Aedes aegypti and Aedes albopictus in the transmission of dengue virus serotype 1 in southern Taiwan

    Directory of Open Access Journals (Sweden)

    Cheng-Hui Tsai

    2017-11-01

    Full Text Available Abstract Background We evaluated the impact of temperature and Wolbachia infection on vector competence of the local Aedes aegypti and Ae. albopictus populations of southern Taiwan in the laboratory. Results After oral infection with dengue serotype 1 virus (DENV-1, female mosquitoes were incubated at temperatures of 10, 16, 22, 28 and 34 °C. Subsequently, salivary gland, head, and thorax-abdomen samples were analyzed for their virus titer at 0, 5, 10, 15, 20, 25 and 30 days post-infection (dpi by real-time RT-PCR. The results showed that Ae. aegypti survived significantly longer and that dengue viral genome levels in the thorax-abdomen (103.25 ± 0.53–104.09 ± 0.71 PFU equivalents/ml and salivary gland samples (102.67 ± 0.33–103.89 ± 0.58 PFU equivalents/ml were significantly higher at high temperature (28–34 °C. The survival of Ae. albopictus was significantly better at 16 or 28 °C, but the virus titers from thorax-abdomen (100.70–102.39 ± 1.31 PFU equivalents/ml and salivary gland samples (100.12 ± 0.05–101.51 ± 0.31 PFU equivalents/ml were significantly higher at 22–28 °C. Within viable temperature ranges, the viruses were detectable after 10 dpi in salivary glands and head tissues in Ae. aegypti and after 5–10 dpi in Ae. albopictus. Vector competence was measured in Ae. albopictus with and without Wolbachia at 28 °C. Wolbachia-infected mosquitoes survived significantly better and carried lower virus titers than Wolbachia-free mosquitoes. Wolbachia coinfections (92.8–97.2% with wAlbA and wAlbB strains were commonly found in a wild population of Ae. albopictus. Conclusions In southern Taiwan, Ae. aegypti is the main vector of dengue and Ae. albopictus has a non-significant role in the transmission of dengue virus due to the high prevalence of Wolbachia infection in the local mosquito population of southern Taiwan.

  18. Optimal stomatal behaviour around the world

    DEFF Research Database (Denmark)

    Lin, Yan-Shih; Medlyn, Belinda E.; Duursma, Remko A.

    2015-01-01

    , a globalscale database and an associated globally applicable model of gs that allow predictions of stomatal behaviour are lacking. Here,we present a database of globally distributed gs obtained in the field for a wide range of plant functional types (PFTs) and biomes. We find that stomatal behaviour diers among...

  19. Morphological changes in different populations of bladder afferent neurons detected by herpes simplex virus (HSV) vectors with cell-type-specific promoters in mice with spinal cord injury.

    Science.gov (United States)

    Shimizu, Nobutaka; Doyal, Mark F; Goins, William F; Kadekawa, Katsumi; Wada, Naoki; Kanai, Anthony J; de Groat, William C; Hirayama, Akihide; Uemura, Hirotsugu; Glorioso, Joseph C; Yoshimura, Naoki

    2017-11-19

    Functional and morphological changes in C-fiber bladder afferent pathways are reportedly involved in neurogenic detrusor overactivity (NDO) after spinal cord injury (SCI). This study examined the morphological changes in different populations of bladder afferent neurons after SCI using replication-defective herpes simplex virus (HSV) vectors encoding the mCherry reporter driven by neuronal cell-type-specific promoters. Spinal intact (SI) and SCI mice were injected into the bladder wall with HSV mCherry vectors driven by the cytomegalovirus (CMV) promoter, CGRP promoter, TRPV1 promoter or neurofilament 200 (NF200) promoter. Two weeks after vector inoculation into the bladder wall, L1 and L6 dorsal root ganglia (DRG) were removed bilaterally for immunofluorescent staining using anti-mCherry antibody. The number of CMV promoter vector-labeled neurons was not altered after SCI. The number of CGRP and TRPV1 promoter vector-labeled neurons was significantly increased whereas the number of NF200 vector-labeled neurons was decreased in L6 DRG after SCI. The median size of CGRP promoter-labeled C-fiber neurons was increased from 247.0 in SI mice to 271.3μm 2 in SCI mice whereas the median cell size of TRPV1 promoter vector-labeled neurons was decreased from 245.2 in SI mice to 216.5μm 2 in SCI mice. CGRP and TRPV1 mRNA levels of laser-captured bladder afferent neurons labeled with Fast Blue were significantly increased in SCI mice compared to SI mice. Thus, using a novel HSV vector-mediated neuronal labeling technique, we found that SCI induces expansion of the CGRP- and TRPV1-expressing C-fiber cell population, which could contribute to C-fiber afferent hyperexcitability and NDO after SCI. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

  20. Assessing the potential impacts of a changing climate on the distribution of a rabies virus vector.

    Directory of Open Access Journals (Sweden)

    Mark A Hayes

    Full Text Available Common vampire bats (Desmodus rotundus occur throughout much of South America to northern México. Vampire bats have not been documented in recent history in the United States, but have been documented within about 50 km of the U.S. state of Texas. Vampire bats feed regularly on the blood of mammals and can transmit rabies virus to native species and livestock, causing impacts on the health of prey. Thus cattle producers, wildlife management agencies, and other stakeholders have expressed concerns about whether vampire bats might spread into the southern United States. On the other hand, concerns about vampire-borne rabies can also result in wanton destruction at bat roosts in areas occupied by vampire bats, but also in areas not known to be occupied by this species. This can in turn negatively affect some bat roosts, populations, and species that are of conservation concern, including vampire bats. To better understand the current and possible future distribution of vampire bats in North America and help mitigate future cattle management problems, we used 7,094 vampire bat occurrence records from North America and species distribution modeling (SDM to map the potential distribution of vampire bats in North America under current and future climate change scenarios. We analysed and mapped the potential distribution of this species using 5 approaches to species distribution modeling: logistic regression, multivariate adaptive regression splines, boosted regression trees, random forest, and maximum entropy. We then projected these models into 17 "worst-case" future climate scenarios for year 2070 to generate hypotheses about how the vampire bat distribution in North America might change in the future. Of the variables used in this analysis, minimum temperature of the coldest month had the highest variable importance using all 5 SDM approaches. These results suggest two potential near-future routes of vampire bat dispersal into the U.S., one via

  1. Real-time PCR protocols for the quantification of the begomovirus tomato yellow leaf curl Sardinia virus in tomato plants and in its insect vector.

    Science.gov (United States)

    Noris, Emanuela; Miozzi, Laura

    2015-01-01

    Tomato yellow leaf curl Sardinia virus (TYLCSV) (Geminiviridae) is an important pathogen, transmitted by the whitefly Bemisia tabaci, that severely affects the tomato production in the Mediterranean basin. Here, we describe real-time PCR protocols suitable for relative and absolute quantification of TYLCSV in tomato plants and in whitefly extracts. Using primers and probe specifically designed for TYLCSV, the protocols for relative quantification allow to compare the amount of TYLCSV present in different plant or whitefly samples, normalized to the amount of DNA present in each sample using endogenous tomato or Bemisia genes as internal references. The absolute quantification protocol allows to calculate the number of genomic units of TYLCSV over the genomic units of the plant host (tomato), with a sensitivity of as few as ten viral genome copies per sample. The described protocols are potentially suitable for several applications, such as plant breeding for resistance, analysis of virus replication, and virus-vector interaction studies.

  2. Population Genetic Structure and Potential Incursion Pathways of the Bluetongue Virus Vector Culicoides brevitarsis (Diptera: Ceratopogonidae) in Australia

    Science.gov (United States)

    Tay, W. T.; Kerr, P. J.; Jermiin, L. S.

    2016-01-01

    Culicoides brevitarsis is a vector of the bluetongue virus (BTV), which infects sheep and cattle. It is an invasive species in Australia with an assumed Asian/South East Asian origin. Using one mitochondrial marker (i.e., part of the cytochrome oxidase subunit I gene) and six nuclear markers, we inferred population genetic structure and possible incursion pathways for Australian C. brevitarsis. Nine mitochondrial haplotypes, with low nucleotide sequence diversity (0.0–0.7%) among these, were identified in a sample of 70 individuals from seven sites. Both sets of markers revealed a homogeneous population structure, albeit with evidence of isolation by distance and two genetically distinct clusters distributed along a north-to-south cline. No evidence of a cryptic species complex was found. The geographical distribution of the mitochondrial haplotypes is consistent with at least two incursion pathways into Australia since the arrival of suitable livestock hosts. By contrast, 15 mitochondrial haplotypes, with up to four times greater nucleotide sequence diversity (0.0–2.9%) among these, were identified in a sample of 16 individuals of the endemic C. marksi (sampled from a site in South Australia and another in New South Wales). A phylogenetic tree inferred using the mitochondrial marker revealed that the Australian and Japanese samples of C. brevitarsis are as evolutionarily different from one another as some of the other Australian species (e.g., C. marksi, C. henryi, C. pallidothorax) are. The phylogenetic tree placed four of the species endemic to Australia (C. pallidothorax, C. bundyensis, C. marksi, C. henryi) in a clade, with a fifth such species (C. bunrooensis) sharing a common ancestor with that clade and a clade comprising two Japanese species (C. verbosus, C. kibunensis). PMID:26771743

  3. The effect of virus-blocking Wolbachia on male competitiveness of the dengue vector mosquito, Aedes aegypti.

    Science.gov (United States)

    Segoli, Michal; Hoffmann, Ary A; Lloyd, Jane; Omodei, Gavin J; Ritchie, Scott A

    2014-12-01

    The bacterial endosymbiont Wolbachia blocks the transmission of dengue virus by its vector mosquito Aedes aegypti, and is currently being evaluated for control of dengue outbreaks. Wolbachia induces cytoplasmic incompatibility (CI) that results in the developmental failure of offspring in the cross between Wolbachia-infected males and uninfected females. This increases the relative success of infected females in the population, thereby enhancing the spread of the beneficial bacterium. However, Wolbachia spread via CI will only be feasible if infected males are sufficiently competitive in obtaining a mate under field conditions. We tested the effect of Wolbachia on the competitiveness of A. aegypti males under semi-field conditions. In a series of experiments we exposed uninfected females to Wolbachia-infected and uninfected males simultaneously. We scored the competitiveness of infected males according to the proportion of females producing non-viable eggs due to incompatibility. We found that infected males were equally successful to uninfected males in securing a mate within experimental tents and semi-field cages. This was true for males infected by the benign wMel Wolbachia strain, but also for males infected by the virulent wMelPop (popcorn) strain. By manipulating male size we found that larger males had a higher success than smaller underfed males in the semi-field cages, regardless of their infection status. The results indicate that Wolbachia infection does not reduce the competitiveness of A. aegypti males. Moreover, the body size effect suggests a potential advantage for lab-reared Wolbachia-males during a field release episode, due to their better nutrition and larger size. This may promote Wolbachia spread via CI in wild mosquito populations and underscores its potential use for disease control.

  4. Winter Activity and Aboveground Hybridization Between the Two Biotypes of the West Nile Virus Vector Culex pipiens.

    Science.gov (United States)

    Vogels, Chantal B F; van de Peppel, Lennart J J; van Vliet, Arnold J H; Westenberg, Marcel; Ibañez-Justicia, Adolfo; Stroo, Arjan; Buijs, Jan A; Visser, Tessa M; Koenraadt, Constantianus J M

    2015-10-01

    Culex (Cx.) pipiens mosquitoes are important vectors of West Nile virus (WNV). In Europe, the species Cx. pipiens consists of two biotypes, pipiens and molestus, which are morphologically identical, but differ in behavior. Typical behavior of the molestus biotype is the ability to remain active during winter, whereas the pipiens biotype enters diapause. The current paradigm is that the two biotypes occur sympatrically in southern Europe, but occur in isolated above- and belowground populations in northern Europe. In northern Europe, hybridization between biotypes is considered to be low because of the barrier that exists between typical habitats. Data on the occurrence of the biotypes and hybrids in northern Europe, however, are scarce, because identification to the level of biotype is often not performed. Our objective was to clarify the distribution of the Cx. pipiens biotypes and to determine hybridization rates in The Netherlands. Cx. pipiens mosquitoes were collected using three different approaches. First, traps were deployed randomly throughout The Netherlands during the summers of 2011 and 2012 (active surveillance). Second, using a web-based reporting platform and media campaign, Dutch citizens were asked to send dead mosquitoes to our laboratory during the winter and summer of 2014 (passive surveillance). Third, larvae and adults were collected during the summer of 2014 from aboveground locations in Amsterdam to identify molestus larval habitats. Real-time PCR was used for identification to the level of biotype. We found that biotype molestus and hybrids were feeding indoors during winter and summer in The Netherlands and that hybridization rates ranged between 6% and 15%. Larval habitats of biotype molestus were found to occur aboveground. The high percentage of hybridization has implications for assessing the risk of WNV transmission, because hybrids are thought to have ideal characteristics for bridging WNV between birds and humans.

  5. Phase I trial of intramuscular injection of a recombinant adeno-associated virus serotype 2 alphal-antitrypsin (AAT) vector in AAT-deficient adults.

    Science.gov (United States)

    Brantly, Mark L; Spencer, L Terry; Humphries, Margaret; Conlon, Thomas J; Spencer, Carolyn T; Poirier, Amy; Garlington, Wendy; Baker, Dawn; Song, Sihong; Berns, Kenneth I; Muzyczka, Nicholas; Snyder, Richard O; Byrne, Barry J; Flotte, Terence R

    2006-12-01

    A phase I trial of intramuscular injection of a recombinant adeno-associated virus serotype 2 (rAAV2) alpha1-antitrypsin (AAT) vector was performed in 12 AAT-deficient adults, 10 of whom were male. All subjects were either homozygous for the most common AAT mutation (a missense mutation designated PI*Z) or compound heterozygous for PI*Z and another mutation known to cause disease. There were four dose cohorts, ranging from 2.1 x 10(12) vector genomes (VG) to 6.9 x 10(13) VG, with three subjects per cohort. Subjects were injected sequentially in a dose-escalating fashion with a minimum of 14 days between patients. Subjects who had been receiving AAT protein replacement discontinued that therapy 28 days before vector administration. There were no vector-related serious adverse events in any of the 12 participants. Vector DNA sequences were detected in the blood between 1 and 3 days after injection in nearly all patients receiving doses of 6.9 x 10(12) VG or higher. Anti-AAV2 capsid antibodies were present and rose after vector injection, but no other immune responses were detected. One subject who had not been receiving protein replacement exhibited low-level expression of wild-type M-AAT in the serum (82 nM), which was detectable 30 days after receiving an injection of 2.1 x 10(13) VG. Unfortunately, residual but declining M-AAT levels from the washout of the protein replacement elevated background levels sufficiently to obscure any possible vector expression in that range in most of the other individuals in the higher dose cohorts.

  6. Origins and Evolution of Stomatal Development.

    Science.gov (United States)

    Chater, Caspar C C; Caine, Robert S; Fleming, Andrew J; Gray, Julie E

    2017-06-01

    The fossil record suggests stomata-like pores were present on the surfaces of land plants over 400 million years ago. Whether stomata arose once or whether they arose independently across newly evolving land plant lineages has long been a matter of debate. In Arabidopsis, a genetic toolbox has been identified that tightly controls stomatal development and patterning. This includes the basic helix-loop-helix (bHLH) transcription factors SPEECHLESS ( SPCH ), MUTE , FAMA , and ICE/SCREAMs ( SCRMs ), which promote stomatal formation. These factors are regulated via a signaling cascade, which includes mobile EPIDERMAL PATTERNING FACTOR ( EPF ) peptides to enforce stomatal spacing. Mosses and hornworts, the most ancient extant lineages to possess stomata, possess orthologs of these Arabidopsis ( Arabidopsis thaliana ) stomatal toolbox genes, and manipulation in the model bryophyte Physcomitrella patens has shown that the bHLH and EPF components are also required for moss stomatal development and patterning. This supports an ancient and tightly conserved genetic origin of stomata. Here, we review recent discoveries and, by interrogating newly available plant genomes, we advance the story of stomatal development and patterning across land plant evolution. Furthermore, we identify potential orthologs of the key toolbox genes in a hornwort, further supporting a single ancient genetic origin of stomata in the ancestor to all stomatous land plants. © 2017 American Society of Plant Biologists. All Rights Reserved.

  7. Interferon signaling in Peromyscus leucopus confers a potent and specific restriction to vector-borne flaviviruses.

    Directory of Open Access Journals (Sweden)

    Adaeze O Izuogu

    Full Text Available Tick-borne flaviviruses (TBFVs, including Powassan virus and tick-borne encephalitis virus cause encephalitis or hemorrhagic fevers in humans with case-fatality rates ranging from 1-30%. Despite severe disease in humans, TBFV infection of natural rodent hosts has little noticeable effect. Currently, the basis for resistance to disease is not known. We hypothesize that the coevolution of flaviviruses with their respective hosts has shaped the evolution of potent antiviral factors that suppress virus replication and protect the host from lethal infection. In the current study, we compared virus infection between reservoir host cells and related susceptible species. Infection of primary fibroblasts from the white-footed mouse (Peromyscus leucopus, a representative host with a panel of vector-borne flaviviruses showed up to a 10,000-fold reduction in virus titer compared to control Mus musculus cells. Replication of vesicular stomatitis virus was equivalent in P. leucopus and M. musculus cells suggesting that restriction was flavivirus-specific. Step-wise comparison of the virus infection cycle revealed a significant block to viral RNA replication, but not virus entry, in P. leucopus cells. To understand the role of the type I interferon (IFN response in virus restriction, we knocked down signal transducer and activator of transcription 1 (STAT1 or the type I IFN receptor (IFNAR1 by RNA interference. Loss of IFNAR1 or STAT1 significantly relieved the block in virus replication in P. leucopus cells. The major IFN antagonist encoded by TBFV, nonstructural protein 5, was functional in P. leucopus cells, thus ruling out ineffective viral antagonism of the host IFN response. Collectively, this work demonstrates that the IFN response of P. leucopus imparts a strong and virus-specific barrier to flavivirus replication. Future identification of the IFN-stimulated genes responsible for virus restriction specifically in P. leucopus will yield mechanistic

  8. Climate change and the spread of vector-borne diseases: using approximate Bayesian computation to compare invasion scenarios for the bluetongue virus vector Culicoides imicola in Italy

    NARCIS (Netherlands)

    Mardulyn, P.; Goffredo, M.; Conte, A.; Hendrickx, G.; Meiswinkel, R.; Balenghien, T.; Sghaier, S.; Lohr, Y.; Gilbert, M.

    2013-01-01

    Bluetongue (BT) is a commonly cited example of a disease with a distribution believed to have recently expanded in response to global warming. The BT virus is transmitted to ruminants by biting midges of the genus Culicoides, and it has been hypothesized that the emergence of BT in Mediterranean

  9. An adenovirus vectored mucosal adjuvant augments protection of mice immunized intranasally with an adenovirus-vectored foot-and-mouth disease virus subunit vaccine

    Science.gov (United States)

    Foot-and-mouth disease virus (FMDV) is a highly contagious pathogen that causes severe morbidity and economic losses to the livestock industry in many countries. The oral and respiratory mucosae are the main ports of entry of FMDV, so the stimulation of local immunity in these tissues may help preve...

  10. An alphavirus vector-based tetravalent dengue vaccine induces a rapid and protective immune response in macaques that differs qualitatively from immunity induced by live virus infection.

    Science.gov (United States)

    White, Laura J; Sariol, Carlos A; Mattocks, Melissa D; Wahala M P B, Wahala; Yingsiwaphat, Vorraphun; Collier, Martha L; Whitley, Jill; Mikkelsen, Rochelle; Rodriguez, Idia V; Martinez, Melween I; de Silva, Aravinda; Johnston, Robert E

    2013-03-01

    Despite many years of research, a dengue vaccine is not available, and the more advanced live attenuated vaccine candidate in clinical trials requires multiple immunizations with long interdose periods and provides low protective efficacy. Here, we report important contributions to the development of a second-generation dengue vaccine. First, we demonstrate that a nonpropagating vaccine vector based on Venezuelan equine encephalitis virus replicon particles (VRP) expressing two configurations of dengue virus E antigen (subviral particles [prME] and soluble E dimers [E85]) successfully immunized and protected macaques against dengue virus, while antivector antibodies did not interfere with a booster immunization. Second, compared to prME-VRP, E85-VRP induced neutralizing antibodies faster, to higher titers, and with improved protective efficacy. Third, this study is the first to map antigenic domains and specificities targeted by vaccination versus natural infection, revealing that, unlike prME-VRP and live virus, E85-VRP induced only serotype-specific antibodies, which predominantly targeted EDIII, suggesting a protective mechanism different from that induced by live virus and possibly live attenuated vaccines. Fourth, a tetravalent E85-VRP dengue vaccine induced a simultaneous and protective response to all 4 serotypes after 2 doses given 6 weeks apart. Balanced responses and protection in macaques provided further support for exploring the immunogenicity and safety of this vaccine candidate in humans.

  11. An Alphavirus Vector-Based Tetravalent Dengue Vaccine Induces a Rapid and Protective Immune Response in Macaques That Differs Qualitatively from Immunity Induced by Live Virus Infection

    Science.gov (United States)

    Sariol, Carlos A.; Mattocks, Melissa D.; Wahala M. P. B., Wahala; Yingsiwaphat, Vorraphun; Collier, Martha L.; Whitley, Jill; Mikkelsen, Rochelle; Rodriguez, Idia V.; Martinez, Melween I.; de Silva, Aravinda; Johnston, Robert E.

    2013-01-01

    Despite many years of research, a dengue vaccine is not available, and the more advanced live attenuated vaccine candidate in clinical trials requires multiple immunizations with long interdose periods and provides low protective efficacy. Here, we report important contributions to the development of a second-generation dengue vaccine. First, we demonstrate that a nonpropagating vaccine vector based on Venezuelan equine encephalitis virus replicon particles (VRP) expressing two configurations of dengue virus E antigen (subviral particles [prME] and soluble E dimers [E85]) successfully immunized and protected macaques against dengue virus, while antivector antibodies did not interfere with a booster immunization. Second, compared to prME-VRP, E85-VRP induced neutralizing antibodies faster, to higher titers, and with improved protective efficacy. Third, this study is the first to map antigenic domains and specificities targeted by vaccination versus natural infection, revealing that, unlike prME-VRP and live virus, E85-VRP induced only serotype-specific antibodies, which predominantly targeted EDIII, suggesting a protective mechanism different from that induced by live virus and possibly live attenuated vaccines. Fourth, a tetravalent E85-VRP dengue vaccine induced a simultaneous and protective response to all 4 serotypes after 2 doses given 6 weeks apart. Balanced responses and protection in macaques provided further support for exploring the immunogenicity and safety of this vaccine candidate in humans. PMID:23302884

  12. Climate Change Influences on the Global Potential Distribution of the Mosquito Culex quinquefasciatus, Vector of West Nile Virus and Lymphatic Filariasis.

    Directory of Open Access Journals (Sweden)

    Abdallah M Samy

    Full Text Available Rapid emergence of most vector-borne diseases (VBDs may be associated with range expansion of vector populations. Culex quinquefasciatus Say 1823 is a potential vector of West Nile virus, Saint Louis encephalitis virus, and lymphatic filariasis. We estimated the potential distribution of Cx. quinquefasciatus under both current and future climate conditions. The present potential distribution of Cx. quinquefasciatus showed high suitability across low-latitude parts of the world, reflecting the current distribution of the species. Suitable conditions were identified also in narrow zones of North Africa and Western Europe. Model transfers to future conditions showed a potential distribution similar to that under present-day conditions, although with higher suitability in southern Australia. Highest stability with changing climate was between 30°S and 30°N. The areas present high agreement among diverse climate models as regards distributional potential in the future, but differed in anticipating potential for distribution in North and Central Africa, southern Asia, central USA, and southeastern Europe. Highest disparity in model predictions across representative concentration pathways (RCPs was in Saudi Arabia and Europe. The model predictions allow anticipation of changing distributional potential of the species in coming decades.

  13. Climate Change Influences on the Global Potential Distribution of the Mosquito Culex quinquefasciatus, Vector of West Nile Virus and Lymphatic Filariasis.

    Science.gov (United States)

    Samy, Abdallah M; Elaagip, Arwa H; Kenawy, Mohamed A; Ayres, Constância F J; Peterson, A Townsend; Soliman, Doaa E

    2016-01-01

    Rapid emergence of most vector-borne diseases (VBDs) may be associated with range expansion of vector populations. Culex quinquefasciatus Say 1823 is a potential vector of West Nile virus, Saint Louis encephalitis virus, and lymphatic filariasis. We estimated the potential distribution of Cx. quinquefasciatus under both current and future climate conditions. The present potential distribution of Cx. quinquefasciatus showed high suitability across low-latitude parts of the world, reflecting the current distribution of the species. Suitable conditions were identified also in narrow zones of North Africa and Western Europe. Model transfers to future conditions showed a potential distribution similar to that under present-day conditions, although with higher suitability in southern Australia. Highest stability with changing climate was between 30°S and 30°N. The areas present high agreement among diverse climate models as regards distributional potential in the future, but differed in anticipating potential for distribution in North and Central Africa, southern Asia, central USA, and southeastern Europe. Highest disparity in model predictions across representative concentration pathways (RCPs) was in Saudi Arabia and Europe. The model predictions allow anticipation of changing distributional potential of the species in coming decades.

  14. Assessing Disparities of Dengue Virus Transmission Risk across the US-Mexican Border Using a Climate Driven Vector-Epidemiological Model

    Science.gov (United States)

    Morin, Cory; Monaghan, Andrew; Quattrochi, Dale; Crosson, William; Hayden, Mary; Ernst, Kacey

    2015-01-01

    Dengue fever is a mosquito-borne viral disease reemerging throughout much of the tropical Americas. Dengue virus transmission is explicitly influenced by climate and the environment through its primary vector, Aedes aegypti. Temperature regulates Ae. aegypti development, survival, and replication rates as well as the incubation period of the virus within the mosquito. Precipitation provides water for many of the preferred breeding habitats of the mosquito, including buckets, old tires, and other places water can collect. Although transmission regularly occurs along the border region in Mexico, dengue virus transmission in bordering Arizona has not occurred. Using NASA's TRMM (Tropical Rainfall Measuring Mission) satellite for precipitation input and Daymet for temperature and supplemental precipitation input, we modeled dengue transmission along a US-Mexico transect using a dynamic dengue transmission model that includes interacting vector ecology and epidemiological components. Model runs were performed for 5 cities in Sonora, Mexico and southern Arizona. Employing a Monte Carlo approach, we performed ensembles of several thousands of model simulations in order to resolve the model uncertainty arising from using different combinations of parameter values that are not well known. For cities with reported dengue case data, the top model simulations that best reproduced dengue case numbers were retained and their parameter values were extracted for comparison. These parameter values were used to run simulations in areas where dengue virus transmission does not occur or where dengue fever case data was unavailable. Additional model runs were performed to reveal how changes in climate or parameter values could alter transmission risk along the transect. The relative influence of climate variability and model parameters on dengue virus transmission is assessed to help public health workers prepare location specific infection prevention strategies.

  15. [Creation of DNA vaccine vector based on codon-optimized gene of rabies virus glycoprotein (G protein) with consensus amino acid sequence].

    Science.gov (United States)

    Starodubova, E S; Kuzmenko, Y V; Latanova, A A; Preobrazhenskaya, O V; Karpov, V L

    2016-01-01

    An optimized design of the rabies virus glycoprotein (G protein) for use within DNA vaccines has been suggested. The design represents a territorially adapted antigen constructed taking into account glycoprotein amino acid sequences of the rabies viruses registered in the Russian Federation and the vaccine Vnukovo-32 strain. Based on the created consensus amino acid sequence, the nucleotide codon-optimized sequence of this modified glycoprotein was obtained and cloned into the pVAX1 plasmid (a vector of the last generation used in the creation of DNA vaccines). A twofold increase in this gene expression compared to the expression of the Vnukovo-32 strain viral glycoprotein gene in a similar vector was registered in the transfected cell culture. It has been demonstrated that the accumulation of modified G protein exceeds the number of the control protein synthesized using the plasmid with the Vnukovo-32 strain viral glycoprotein gene by 20 times. Thus, the obtained modified rabies virus glycoprotein can be considered to be a promising DNA vaccine antigen.

  16. Expression of NS3/NS4A Proteins of Hepatitis C Virus in Huh7 Cells Following Engineering Its Eukaryotic Expression Vector.

    Science.gov (United States)

    Behzadi, Mohammad Amin; Alborzi, Abdolvahab; Pouladfar, Gholamreza; Dianatpour, Mehdi; Ziyaeyan, Mazyar

    2015-11-01

    Although the development of novel therapeutic regimens to combat hepatitis C virus (HCV) infection have been speeded up with successful results, no efficient vaccines exist yet. This study aimed to construct a eukaryotic expression vector encoding nonstructural proteins, NS3/NS4A, of HCV genotype 3a, and evaluate its expression on Huh7 cell surface. The NS3/NS4A sequence was isolated from a patient with HCV-3a chronic infection, cloned into intermediate vector pTZ57R/T, and then used for engineering a mammalian expression vector, pDisplay, to direct the respective protein to the secretory pathway and anchor it to the plasma membrane. The expression of the protein in Huh7 cell, which was transiently transfected with the vector using Lipofectamine, was determined by immunocytochemical staining assay with fluorescein isothiocyanate (FITC)-conjugated antibodies to the HA/myc tags located besides the fusion fragment. The results showed that the fragment was successfully amplified and cloned into a eukaryotic expression vector. Sequencing and enzyme digestion analysis confirmed the cloned gene completion and its correct position in the pDisply-NS3/NS4A plasmid. Immunocytochemical staining revealed that the target protein was expressed as a membrane-anchored protein in the Huh7 cells. This study can serve as a fundamental experiment for the construction of a NS3/NS4A eukaryotic expression vector and its expression in mammalian cells. Further research is underway to evaluate the fragment immunogenicity in lab animal models.

  17. Long-Term Efficacy Following Readministration of an Adeno-Associated Virus Vector in Dogs with Glycogen Storage Disease Type Ia

    Science.gov (United States)

    Demaster, Amanda; Luo, Xiaoyan; Curtis, Sarah; Williams, Kyha D.; Landau, Dustin J.; Drake, Elizabeth J.; Kozink, Daniel M.; Bird, Andrew; Crane, Bayley; Sun, Francis; Pinto, Carlos R.; Brown, Talmage T.; Kemper, Alex R.

    2012-01-01

    Abstract Glycogen storage disease type Ia (GSD-Ia) is the inherited deficiency of glucose-6-phosphatase (G6Pase), primarily found in liver and kidney, which causes life-threatening hypoglycemia. Dogs with GSD-Ia were treated with double-stranded adeno-associated virus (AAV) vectors encoding human G6Pase. Administration of an AAV9 pseudotyped (AAV2/9) vector to seven consecutive GSD-Ia neonates prevented hypoglycemia during fasting for up to 8 hr; however, efficacy eventually waned between 2 and 30 months of age, and readministration of a new pseudotype was eventually required to maintain control of hypoglycemia. Three of these dogs succumbed to acute hypoglycemia between 7 and 9 weeks of age; however, this demise could have been prevented by earlier readministration an AAV vector, as demonstrated by successful prevention of mortality of three dogs treated earlier in life. Over the course of this study, six out of nine dogs survived after readministration of an AAV vector. Of these, each dog required readministration on average every 9 months. However, two were not retreated until >34 months of age, while one with preexisting antibodies was re-treated three times in 10 months. Glycogen content was normalized in the liver following vector administration, and G6Pase activity was increased in the liver of vector-treated dogs in comparison with GSD-Ia dogs that received only with dietary treatment. G6Pase activity reached approximately 40% of normal in two female dogs following AAV2/9 vector administration. Elevated aspartate transaminase in absence of inflammation indicated that hepatocellular turnover in the liver might drive the loss of vector genomes. Survival was prolonged for up to 60 months in dogs treated by readministration, and all dogs treated by readministration continue to thrive despite the demonstrated risk for recurrent hypoglycemia and mortality from waning efficacy of the AAV2/9 vector. These preclinical data support the further translation of AAV

  18. Seasonal abundance and molecular identification of West Nile virus vectors, Culex pipens and Culex quinquefasciatus (diptera: culicidae) in Abeokuta, South-West, Nigeria.

    Science.gov (United States)

    Motayo, Babatunde Olanrewaju; Onoja, Bernard Aneibe; Faneye, Adedayo Omotayo; Adeniji, Johnson Adekunle

    2016-03-01

    West Nile virus (WNV) infection, is an arbovirus infection with high morbidity and mortality, the vector responsible for both human and animal transmission is Culex pipens complex. To determine the species distribution and seasonal abundance of Culex pipens and Culex quinquefasciatus mosquitoes in Abeokuta, Nigeria. Mosquitoes belonging to the Culex pipens complex were captured in three different locations located within Abeokuta Metropolis between March 2012 and January 2013. Individual species were identified using morphometric methods. Amplification of the Ace2 gene by PCR confirmed morphormetric identification of the mosquitoes. A total of 751 mosquitoes were captured. Culex quinquefaciatus recorded the highest distribution of vectors with 56.6% and Culex pipens 43.4% (P > 0.05). Idi aba community recorded the highest distribution of mosquito vectors with 42.9% (n=322) and Culex quinqueaciatus was more abundantly distributed with 183 mosquitoes. Aro community recorded 32% (n=240) of captured mosquitoes with Culex quinquefaciatus having a higher level of abundance and lastly Kemta with a distribution of 25.1% (n=189). Results from this study show that potential vectors of WNV abound within Abeokuta, putting residents at high risk of West Nile infection. We advocate for introduction of routine testing of WNV in Abeokuta and Nigeria.