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.
Beier, Kevin T; Mundell, Nathan A; Pan, Y Albert; Cepko, Constance L
Viruses have been used as transsynaptic tracers, allowing one to map the inputs and outputs of neuronal populations, due to their ability to replicate in neurons and transmit in vivo only across synaptically connected cells. To date, their use has been largely restricted to mammals. In order to explore the use of such viruses in an expanded host range, we tested the transsynaptic tracing ability of recombinant vesicular stomatitis virus (rVSV) vectors in a variety of organisms. Successful infection and gene expression were achieved in a wide range of organisms, including vertebrate and invertebrate model organisms. Moreover, rVSV enabled transsynaptic tracing of neural circuitry in predictable directions dictated by the viral envelope glycoprotein (G), derived from either VSV or rabies virus (RABV). Anterograde and retrograde labeling, from initial infection and/or viral replication and transmission, was observed in Old and New World monkeys, seahorses, jellyfish, zebrafish, chickens, and mice. These vectors are widely applicable for gene delivery, afferent tract tracing, and/or directional connectivity mapping. Here, we detail the use of these vectors and provide protocols for propagating virus, changing the surface glycoprotein, and infecting multiple organisms using several injection strategies. Copyright © 2016 John Wiley & Sons, Inc.
Marlena M. Westcott
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.
Ma, Yuanmei; Duan, Yue; Wei, Yongwei; Liang, Xueya; Niewiesk, Stefan; Oglesbee, Michael
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
Vesicular Stomatitis (VS) is caused by the Vesicular Stomatitis Virus (VSV), a negative single stranded RNA arthropod-borne virus member of the Family Rhabdoviridae. The virion is composed of the host derived plasma membrane, the envelope, and an internal ribonucleoprotein core. The envelope contain...
Regules, Jason A; Beigel, John H; Paolino, Kristopher M; Voell, Jocelyn; Castellano, Amy R; Hu, Zonghui; Muñoz, Paula; Moon, James E; Ruck, Richard C; Bennett, Jason W; Twomey, Patrick S; Gutiérrez, Ramiro L; Remich, Shon A; Hack, Holly R; Wisniewski, Meagan L; Josleyn, Matthew D; Kwilas, Steven A; Van Deusen, Nicole; Mbaya, Olivier Tshiani; Zhou, Yan; Stanley, Daphne A; Jing, Wang; Smith, Kirsten S; Shi, Meng; Ledgerwood, Julie E; Graham, Barney S; Sullivan, Nancy J; Jagodzinski, Linda L; Peel, Sheila A; Alimonti, Judie B; Hooper, Jay W; Silvera, Peter M; Martin, Brian K; Monath, Thomas P; Ramsey, W Jay; Link, Charles J; Lane, H Clifford; Michael, Nelson L; Davey, Richard T; Thomas, Stephen J
The worst Ebola virus disease (EVD) outbreak in history has resulted in more than 28,000 cases and 11,000 deaths. We present the final results of two phase 1 trials of an attenuated, replication-competent, recombinant vesicular stomatitis virus (rVSV)-based vaccine candidate designed to prevent EVD. We conducted two phase 1, placebo-controlled, double-blind, dose-escalation trials of an rVSV-based vaccine candidate expressing the glycoprotein of a Zaire strain of Ebola virus (ZEBOV). A total of 39 adults at each site (78 participants in all) were consecutively enrolled into groups of 13. At each site, volunteers received one of three doses of the rVSV-ZEBOV vaccine (3 million plaque-forming units [PFU], 20 million PFU, or 100 million PFU) or placebo. Volunteers at one of the sites received a second dose at day 28. Safety and immunogenicity were assessed. The most common adverse events were injection-site pain, fatigue, myalgia, and headache. Transient rVSV viremia was noted in all the vaccine recipients after dose 1. The rates of adverse events and viremia were lower after the second dose than after the first dose. By day 28, all the vaccine recipients had seroconversion as assessed by an enzyme-linked immunosorbent assay (ELISA) against the glycoprotein of the ZEBOV-Kikwit strain. At day 28, geometric mean titers of antibodies against ZEBOV glycoprotein were higher in the groups that received 20 million PFU or 100 million PFU than in the group that received 3 million PFU, as assessed by ELISA and by pseudovirion neutralization assay. A second dose at 28 days after dose 1 significantly increased antibody titers at day 56, but the effect was diminished at 6 months. This Ebola vaccine candidate elicited anti-Ebola antibody responses. After vaccination, rVSV viremia occurred frequently but was transient. These results support further evaluation of the vaccine dose of 20 million PFU for preexposure prophylaxis and suggest that a second dose may boost antibody responses
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.
Bishnoi, Suman; Tiwari, Ritudhwaj; Gupta, Sharad; Byrareddy, Siddappa N; Nayak, Debasis
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.
Metsikkö, K.; van Meer, G.; Simons, K.
Enveloped virus glycoproteins exhibit membrane fusion activity. We have analysed whether the G protein of vesicular stomatitis virus, reconstituted into liposomes, is able to fuse nucleated cells in a pH-dependent fashion. Proteoliposomes produced by octylglucoside dialysis did not exhibit cell
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
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.
Tripp, Ralph A.; Tompkins, S. Mark
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
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
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.
Mundell, Nathan A; Beier, Kevin T; Pan, Y Albert; Lapan, Sylvain W; Göz Aytürk, Didem; Berezovskii, Vladimir K; Wark, Abigail R; Drokhlyansky, Eugene; Bielecki, Jan; Born, Richard T; Schier, Alexander F; Cepko, Constance L
Current limitations in technology have prevented an extensive analysis of the connections among neurons, particularly within nonmammalian organisms. We developed a transsynaptic viral tracer originally for use in mice, and then tested its utility in a broader range of organisms. By engineering the vesicular stomatitis virus (VSV) to encode a fluorophore and either the rabies virus glycoprotein (RABV-G) or its own glycoprotein (VSV-G), we created viruses that can transsynaptically label neuronal circuits in either the retrograde or anterograde direction, respectively. The vectors were investigated for their utility as polysynaptic tracers of chicken and zebrafish visual pathways. They showed patterns of connectivity consistent with previously characterized visual system connections, and revealed several potentially novel connections. Further, these vectors were shown to infect neurons in several other vertebrates, including Old and New World monkeys, seahorses, axolotls, and Xenopus. They were also shown to infect two invertebrates, Drosophila melanogaster, and the box jellyfish, Tripedalia cystophora, a species previously intractable for gene transfer, although no clear evidence of transsynaptic spread was observed in these species. These vectors provide a starting point for transsynaptic tracing in most vertebrates, and are also excellent candidates for gene transfer in organisms that have been refractory to other methods. © 2015 Wiley Periodicals, Inc.
Mead, Daniel G; Howerth, Elizabeth W; Murphy, Molly D; Gray, Elmer W; Noblet, Raymond; Stallknecht, David E
The transmission routes of Vesicular stomatitis New Jersey virus (VSNJV), a causative agent of vesicular stomatitis, an Office International des Epizooties List-A disease, are not completely understood. Epidemiological and entomological studies conducted during the sporadic epidemics in the western United States have identified potential virus transmission routes involving insect vectors and animal-to-animal contact. In the present study we experimentally tested the previously proposed transmission routes which were primarily based on field observations. Results obtained provide strong evidence for the following: (1) hematophagous insects acquire VSNJV by unconventional routes while blood feeding on livestock, (2) clinical course of VSNJV infection in livestock following transmission by an infected insect is related to insect bite site, (3) infection of livestock via insect bite can result in multiple transmission possibilities, including animal-to-animal contact. Taken together, these data significantly add to our understanding of the transmission routes of a causative agent of one of the oldest known infectious diseases of livestock, for which the details have remained largely unknown despite decades of research.
Drolet, Barbara S; Stuart, Melissa A; Derner, Justin D
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.
Kalvodova, Lucie; Sampaio, Julio L; Cordo, Sandra
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...
Chad E Mire
Full Text Available Ebola virus (EBOV causes severe and often fatal hemorrhagic fever in humans and nonhuman primates (NHPs. Currently, there are no licensed vaccines or therapeutics for human use. Recombinant vesicular stomatitis virus (rVSV-based vaccine vectors, which encode an EBOV glycoprotein in place of the VSV glycoprotein, have shown 100% efficacy against homologous Sudan ebolavirus (SEBOV or Zaire ebolavirus (ZEBOV challenge in NHPs. In addition, a single injection of a blend of three rVSV vectors completely protected NHPs against challenge with SEBOV, ZEBOV, the former Côte d'Ivoire ebolavirus, and Marburg virus. However, recent studies suggest that complete protection against the newly discovered Bundibugyo ebolavirus (BEBOV using several different heterologous filovirus vaccines is more difficult and presents a new challenge. As BEBOV caused nearly 50% mortality in a recent outbreak any filovirus vaccine advanced for human use must be able to protect against this new species. Here, we evaluated several different strategies against BEBOV using rVSV-based vaccines. Groups of cynomolgus macaques were vaccinated with a single injection of a homologous BEBOV vaccine, a single injection of a blended heterologous vaccine (SEBOV/ZEBOV, or a prime-boost using heterologous SEBOV and ZEBOV vectors. Animals were challenged with BEBOV 29-36 days after initial vaccination. Macaques vaccinated with the homologous BEBOV vaccine or the prime-boost showed no overt signs of illness and survived challenge. In contrast, animals vaccinated with the heterologous blended vaccine and unvaccinated control animals developed severe clinical symptoms consistent with BEBOV infection with 2 of 3 animals in each group succumbing. These data show that complete protection against BEBOV will likely require incorporation of BEBOV glycoprotein into the vaccine or employment of a prime-boost regimen. Fortunately, our results demonstrate that heterologous rVSV-based filovirus vaccine
Thomas W Geisbert
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.
von Rhein, Christine; Weidner, Tatjana; Henß, Lisa; Martin, Judith; Weber, Christopher; Sliva, Katja; Schnierle, Barbara S
Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes chikungunya fever and has infected millions of people mainly in developing countries. The associated disease is characterized by rash, high fever, and severe arthritis that can persist for years. CHIKV has adapted to Aedes albopictus, which also inhabits temperate regions including Europe and the United States of America. CHIKV has recently caused large outbreaks in Latin America. No treatment or licensed CHIKV vaccine exists. Traditional medicines are known to have anti-viral effects; therefore, we examined whether curcumin or Boswellia serrata gum resin extract have antiviral activity against CHIKV. Both compounds blocked entry of CHIKV Env-pseudotyped lentiviral vectors and inhibited CHIKV infection in vitro. In addition, vesicular stomatitis virus vector particles and viral infections were also inhibited to the same extent, indicating a broad antiviral activity. Although the bioavailability of these compounds is rather poor, they might be used as a lead structure to develop more effective antiviral drugs or might be used topically to prevent CHIKV spread in the skin after mosquito bites. Copyright © 2015 Elsevier B.V. All rights reserved.
Pérez De León, Adalberto A; O'Toole, Donal; Tabachnick, Walter J
Intrathoracically inoculated Culicoides sonorensis Wirth & Jones were capable of transmitting vesicular stomatitis New Jersey virus (family Rhabdoviridae, genus Vesiculovirus, VSNJV) during blood feeding on the abdomen of six guinea pigs. None of the guinea pigs infected in this manner developed clinical signs of vesicular stomatitis despite seroconversion for VSNJV. Guinea pigs infected by intradermal inoculations of VSNJV in the abdomen also failed to develop clinical signs of vesicular stomatitis. Three guinea pigs given intradermal inoculations of VSNJV in the foot pad developed lesions typical of vesicular stomatitis. Transmission by the bite of C. sonorensis may have facilitated guinea pig infection with VSNJV because a single infected C. sonorensis caused seroconversion and all guinea pigs infected by insect bite seroconverted compared with 50% of the guinea pigs infected by intradermal inoculation with a higher titer VSNJV inoculum. The role of C. sonorensis in the transmission of VSNJV is discussed.
Jensen, Helle; Andresen, Lars; Nielsen, Jens
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 ...
Shinde, Prashant V; Xu, Haifeng C; Maney, Sathish Kumar
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...... stomatitis virus infection, phagocytes produce tumor necrosis factor (TNF) which signals via TNFR1 and promote "enforced virus replication" in CD169(+) macrophages. Consequently, lack of TNF or TNFR1 resulted in defective immune activation and VSV clearance....
Buonocore, Linda; Blight, Keril J.; Rice, Charles M.; Rose, John K.
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...
Ramsauer, Katrin; Tangy, Frédéric
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 firstname.lastname@example.org.
Dunigan, D.D.; Lucas-Lenard, J.M.
When mouse L-cells are infected with vesicular stomatitis virus, there is a decrease in the rate of protein synthesis ranging from 20 to 85% of that in mock-infected cells. Vesicular stomatitis virus, irradiated with increasing doses of UV light, eventually loses this capacity to inhibit protein synthesis. The UV inactivation curve was biphasic, suggesting that transcription of two regions of the viral genome is necessary for the virus to become inactivated in this capacity. The first transcription produced corresponded to about 373 nucleotides, and the second corresponded to about 42 nucleotides. Inhibition of transcription of the larger product by irradiating the virus with low doses of UV light left a residual inhibition of protein synthesis consisting of approximately 60 to 65% of the total inhibition. This residual inhibition could be obviated by irradiating the virus with a UV dose of greater than 20,000 ergs/mm 2 and was thus considered to represent the effect of the smaller transcription product. In the R1 mutant of another author, the inhibition of transcription of the larger product sufficed to restore protein synthesis to the mock-infected level, suggesting that the smaller transcription product is nonfunctional with respect to protein synthesis inhibition. Extracts from cells infected with virus irradiated with low doses of UV light showed a protein synthesis capacity quite similar to that of their in vivo counterparts, indicating that these extracts closely reflect the in vivo effects of virus infection
Full Text Available MeCab user dictionary for science technology term vesicular stomatitis virus 名詞 一般 ...* * * * 水疱性口内炎ウイルス スイホウセイコウナイエンウイルス スイホーセイコーナイエンウイルス Thesaurus2015 200906056003651861 C LS07 UNKNOWN_2 vesicular stomatitis virus
Gumpper, Ryan H.; Li, Weike; Castañeda, Carlos H.; Scuderi, M. José; Bashkin, James K.; Luo, Ming; Dutch, Rebecca Ellis
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.
van der Sluijs, Mirjam Tineke Willemijn; de Smit, Abraham J; Moormann, Rob J M
Bluetongue is an economically important disease of ruminants. The causative agent, Bluetongue virus (BTV), is mainly transmitted by insect vectors. This review focuses on vector-free BTV transmission, and its epizootic and economic consequences. Vector-free transmission can either be vertical, from dam to fetus, or horizontal via direct contract. For several BTV-serotypes, vertical (transplacental) transmission has been described, resulting in severe congenital malformations. Transplacental transmission had been mainly associated with live vaccine strains. Yet, the European BTV-8 strain demonstrated a high incidence of transplacental transmission in natural circumstances. The relevance of transplacental transmission for the epizootiology is considered limited, especially in enzootic areas. However, transplacental transmission can have a substantial economic impact due to the loss of progeny. Inactivated vaccines have demonstrated to prevent transplacental transmission. Vector-free horizontal transmission has also been demonstrated. Since direct horizontal transmission requires close contact of animals, it is considered only relevant for within-farm spreading of BTV. The genetic determinants which enable vector-free transmission are present in virus strains circulating in the field. More research into the genetic changes which enable vector-free transmission is essential to better evaluate the risks associated with outbreaks of new BTV serotypes and to design more appropriate control measures.
Abe, Akihiro; Miyanohara, Atsushi; Friedmann, Theodore
Exposure of Lipofectin-DNA complexes to the partially purified G glycoprotein of the vesicular stomatitis virus envelope (VSV-G) results in loss of serum-mediated inhibition and in enhanced efficiency of gene transfer. Sucrose density gradient sedimentation analysis indicated that the VSV-G associates physically with the DNA-lipid complex to produce a VSV-G liposome. The ability to incorporate surrogate viral or cellular envelope components such as VSV-G into liposomes may allow more-efficient and possibly targeted gene delivery by lipofection, both in vitro and in vivo. PMID:9621082
Adler, R; Banerjee, A K
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.
Rose, J K; Shafferman, A
Bacterial plasmids that directed expression of the vesicular stomatitis virus glycoprotein (G-protein) gene under control of the tryptophan operon regulatory region were constructed. A plasmid directing the synthesis of a G-protein-like protein (containing the NH2-terminal segment of seven amino acids encoded by the trpE gene fused to the complete G-protein sequence lacking only its NH2-terminal methionine) could be transformed into trpR+ (repressed) but not into trpR- (derepressed) cells. Th...
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.
Moeschler, Sarah; Locher, Samira; Conzelmann, Karl-Klaus; Krämer, Beate; Zimmer, Gert
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.
Marzi, Andrea; Hanley, Patrick W; Haddock, Elaine; Martellaro, Cynthia; Kobinger, Gary; Feldmann, Heinz
The Ebola virus (EBOV) epidemic in West Africa increased the focus on vaccine development against this hemorrhagic fever-causing pathogen, and as a consequence human clinical trials for a few selected platforms were accelerated. One of these vaccines is vesicular stomatitis virus (VSV)-EBOV, also known as rVSV-ZEBOV, a fast-acting vaccine against EBOV and so far the only vaccine with reported efficacy against EBOV infections in humans in phase III clinical trials. In this study, we analyzed the potential of VSV-EBOV for postexposure treatment of rhesus macaques infected with EBOV-Makona. We treated groups of animals with 1 dose of VSV-EBOV either in a single injection at 1 or 24 hours after EBOV exposure or with 2 injections, half the dose at each time point; 1 control group received the same dose of the VSV-based Marburg virus vaccine at both time points; another group remained untreated. Although all untreated animals succumbed to EBOV infection, 33%-67% of the animals in each treatment group survived the infection, including the group treated with the VSV-based Marburg virus vaccine. This result suggests that protection from postexposure vaccination may be antigen unspecific and due rather to an early activation of the innate immune system. In conclusion, VSV-EBOV remains a potent and fast-acting prophylactic vaccine but demonstrates only limited efficacy in postexposure treatment. Published by Oxford University Press for the Infectious Diseases Society of America 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.
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
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
Ivec, Martin; Botic, Tanja; Koren, Srecko
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....
Logan, Nicola; Dundon, William G; Diallo, Adama; Baron, Michael D; James Nyarobi, M; Cleaveland, Sarah; Keyyu, Julius; Fyumagwa, Robert; Hosie, Margaret J; Willett, Brian J
The measurement of virus-specific neutralising antibodies represents the "gold-standard" for diagnostic serology. For animal morbilliviruses, such as peste des petits ruminants (PPRV) or rinderpest virus (RPV), live virus-based neutralisation tests require high-level biocontainment to prevent the accidental escape of the infectious agents. In this study, we describe the adaptation of a replication-defective vesicular stomatitis virus (VSVΔG) based pseudotyping system for the measurement of neutralising antibodies against animal morbilliviruses. By expressing the haemagglutinin (H) and fusion (F) proteins of PPRV on VSVΔG pseudotypes bearing a luciferase marker gene, neutralising antibody titres could be measured rapidly and with high sensitivity. Serological responses against the four distinct lineages of PPRV could be measured simultaneously and cross-neutralising responses against other morbilliviruses compared. Using this approach, we observed that titres of neutralising antibodies induced by vaccination with live attenuated PPRV were lower than those induced by wild type virus infection and the level of cross-lineage neutralisation varied between vaccinates. By comparing neutralising responses from animals infected with either PPRV or RPV, we found that responses were highest against the homologous virus, indicating that retrospective analyses of serum samples could be used to confirm the nature of the original pathogen to which an animal had been exposed. Accordingly, when screening sera from domestic livestock and wild ruminants in Tanzania, we detected evidence of cross-species infection with PPRV, canine distemper virus (CDV) and a RPV-related bovine morbillivirus, suggesting that exposure to animal morbilliviruses may be more widespread than indicated previously using existing diagnostic techniques. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.
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
Nelson Nogueira Reis
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.
Whitt, M.A.; Chong, L.; Rose, J.K.
The authors have used transient expression of the wild-type vesicular stomatitis virus (VSV) glycoprotein (G protein) from cloned cDNA to rescue a temperature-sensitive G protein mutant of VSV in cells at the nonpermissive temperature. Using cDNAs encoding G proteins with deletions in the normal 29-amino-acid cytoplasmic domain, they determined that the presence of either the membrane-proximal 9 amino acids or the membrane-distal 12 amino acids was sufficient for rescue of the temperature-sensitive mutant. G proteins with cytoplasmic domains derived from other cellular or viral G proteins did not rescue the mutant, nor did G proteins with one or three amino acids of the normal cytoplasmic domain. Rescue correlated directly with the ability of the G proteins to be incorporated into virus particles. This was shown by analysis of radiolabeled particles separated on sucrose gradients as well as by electron microscopy of rescued virus after immunogold labeling. Quantitation of surface expression showed that all of the mutated G proteins were expressed less efficiently on the cell surface than was wild-type G protein. However, they were able to correct for differences in rescue efficiency resulting from differences in the level of surface expression by reducing wild-type G protein expression to levels equivalent to those observed for the mutated G proteins. The results provide evidence that at least a portion of the cytoplasmic domain is required for efficient assembly of the VSV G protein into virions during virus budding
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
...). 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...
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.
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
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.
Vinícius Leobet Lunkes
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.
Lin, Shi-jia; Yu, Qin-mei; Meng, Wen-tong; Wen, Yan-jun; Chen, Li-juan; Niu, Ting
To explore antitumor effects of plasmid pcDNA3. 1-MP encoding matrix protein of vesicular stomatitis virus (VSV) complexed with cationic liposome (DOTAP:CHOL) in mice with EL4 lymphoma. C57BL/6 mouse model with EL4 lymphoma was established. Sixty mice bearing EL4 lymphoma were divided randomly into five groups including Lip-MP, Lip-pVAX, Lip, ADM and NS groups, which were intravenously injected with liposome-pcDNA 3. 1-MP complex, liposome-pVAX complex, empty liposome, Adriamycin and normal saline respectively every three days. Tumor volumes and survival time were monitored. Microvessel density and tumor proliferative index in tumor tissues were determined by CD31, Ki-67 immunohistochemistry staining, meanwhile the tumor apoptosis index was measured by TUNEL method. From 6 days after treatments on, the tumor volume in Lip-MP group was much smaller than that in Lip-pVAX, Lip and NS group (P EL4 tumor cells in vivo (P EL4 lymphoma, which may be related to the induction of tumor cell apoptosis, inhibition of tumor angiogenesis, and suppression of tumor cell proliferation.
Fabrice Le Boeuf
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.
Perez de Leon, Adalberto A; Tabachnick, Walter J
Laboratory-reared Culicoides sonorensis Wirth & Jones were infected with vesicular stomatitis virus serotype New Jersey (family Rhabdoviridae, genus Vesiculovirus, VSNJV) through intrathoracic inoculation. After 10-d incubation at 25 degrees C, these insects were allowed to blood feed on four steers. Two other steers were exposed to VSNJV through intralingual inoculation with 10(8) tissue culture infective dose50 VSNJV. All six steers became seropositive for VSNJV. The results demonstrate the ability of C. sonorensis to transmit VSNJV to livestock. Only the animals intralingually inoculated with VSNJV showed clinical signs in the form of vesicles at the site of inoculation. Uninfected C. sonorensis allowed to feed on the exposed animals did not become infected with VSNJV. Animals infected by C. sonorensis showed a slower antibody response compared with intralingually inoculated animals. This is probably because of different amounts of virus received via insect transmission and syringe inoculation. A significant difference was found in the serum acute-phase protein alpha-1-acid glycoprotein in animals that received VSNJV through C. sonorensis transmission. These animals had previously been exposed to insect attack in the field compared with intralingually inoculated animals and C. sonorensis-infected animals that had been protected from insect attack. The failure to observe clinical signs of vesicular stomatitis through transmission of VSNJV by C. sonorensis may explain widespread subclinical infections during vesicular stomatitis epidemics.
Evans, Michelle V; Dallas, Tad A; Han, Barbara A; Murdock, Courtney C; Drake, John M
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
Martin-Acebes, Miguel A.; Gonzalez-Magaldi, Monica; Rosas, Maria F.; Borrego, Belen; Brocchi, Emiliana; Armas-Portela, Rosario; Sobrino, Francisco
The intracellular distribution of swine vesicular disease virus (SVDV) proteins and the induced reorganization of endomembranes in IBRS-2 cells were analyzed. Fluorescence to new SVDV capsids appeared first upon infection, concentrated in perinuclear circular structures and colocalized to dsRNA. As in foot-and-mouth disease virus (FMDV)-infected cells, a vesicular pattern was predominantly found in later stages of SVDV capsid morphogenesis that colocalized with those of non-structural proteins 2C, 2BC and 3A. These results suggest that assembly of capsid proteins is associated to the replication complex. Confocal microscopy showed a decreased fluorescence to ER markers (calreticulin and protein disulfide isomerase), and disorganization of cis-Golgi gp74 and trans-Golgi caveolin-1 markers in SVDV- and FMDV-, but not in vesicular stomatitis virus (VSV)-infected cells. Electron microscopy of SVDV-infected cells at an early stage of infection revealed fragmented ER cisternae with expanded lumen and accumulation of large Golgi vesicles, suggesting alterations of vesicle traffic through Golgi compartments. At this early stage, FMDV induced different patterns of ER fragmentation and Golgi alterations. At later stages of SVDV cytopathology, cells showed a completely vacuolated cytoplasm containing vesicles of different sizes. Cell treatment with brefeldin A, which disrupts the Golgi complex, reduced SVDV (∼ 5 log) and VSV (∼ 4 log) titers, but did not affect FMDV growth. Thus, three viruses, which share target tissues and clinical signs in natural hosts, induce different intracellular effects in cultured cells
Michael Karl Melzer
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.
Wei, Y; Wang, S; Wang, X
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.
Belliure, B.; Janssen, A.; Maris, P.C.; Peters, D.; Sabelis, M.W.
Plants infected with pathogens often attract the pathogens' vectors, but it is not clear if this is advantageous to the vectors. We therefore quantified the direct and indirect (through the host plant) effects of a pathogen on its vector. A positive direct effect of the plant-pathogenic Tomato
Rommelfanger, D M; Offord, C P; Dev, J; Bajzer, Z; Vile, R G; Dingli, D
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.
Erlwein, O; McClure, M O
Foamy viruses, distantly related to the major subfamily of Retroviruses, Orthoretroviruses that include oncoviruses (for example, murine leukemia virus (MLV)) and lentiviruses (human immunodeficiency virus (HIV)), are endemic in mammalian species, but not in human populations. Humans infected by accidental or occupational exposure remain well. The virus is not transmitted to others, nor is it associated with any disease. These features added to its broad host range, efficient transduction of progenitor cells and an integration profile less likely to induce insertional mutagenesis, make these viruses attractive as vectors. Long-term reversal of disease phenotype in dogs with the genetic defect, leukocyte adhesion deficiency, by foamy virus vector therapy strengthens the case for their clinical exploitation.
Knop, David R; Harrell, Heather
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.
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.
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
Folimonov, Alexey S.; Folimonova, Svetlana Y.; Bar-Joseph, Moshe; Dawson, William O.
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
Fraefel, Cornel; Marconi, Peggy; Epstein, Alberto L
Herpes simplex virus type 1 (HSV-1) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153 kbp double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes (1) the two approaches most commonly used to prepare recombinant vectors through homologous recombination, either in eukaryotic cells or in bacteria, and (2) the two methodologies currently used to generate helper-free amplicon vectors, either using a bacterial artificial chromosome (BAC)-based approach or a Cre/loxP site-specific recombination strategy.
Emerson, S.U.; Schubert, M.
Recombinant DNA techniques were used to delete regions of a cDNA clone of the phosphoprotein NS gene of vesicular stomatitis virus. The complete NS gene and four mutant genes containing internal or terminal deletions were inserted into a modified pGem4 vector under the transcriptional control of the page T7 promoter. Run-off transcripts were synthesized and translated in vitro to provide [ 35 S]methionine-labeled complete NS or deletion mutant NS proteins. Immune coprecipitation assays involving these proteins were developed to map the regions of the NS protein responsible for binding to the structural viral nucleocapsid protein N and the catalytic RNA polymerase protein L. The data indicate the NS protein is a bivalent protein consisting of two discrete functional domains. Contrary to previous suggestions, the negatively charged amino-terminal half of NS protein binds to L protein, while the carboxyl-terminal half of NS protein binds to both soluble recombinant nucleocapsid protein N and viral ribonucleocapsid template
Rasmussen, Thomas Bruun; Uttenthal, Åse; Fernandez, Jovita
In order to establish a rapid and reliable system for the detection of vesicular stomatitis virus (VSV), we developed a quantitative reverse transcription-PCR assay for the detection, quantification, and differentiation of the major serotypes, VSV Indiana and VSV New Jersey, using a closed......-tube multiplex format. The detection system is based on the recently invented primer-probe energy transfer (PriProET) system. A region of the gene encoding the RNA-dependent RNA polymerase was amplified by using VSV-specific primers in the presence of two serotype-specific fluorescent probes. By incorporating...... probes. The limits of detection ware found to be less than 10 50% tissue culture infective doses/ml for both serotypes. The diagnostic value of the new method was tested with clinical materials from experimentally infected pigs, and it is concluded that the method is a powerful tool for the rapid...
Chen, Gong; Pan, Huipeng; Xie, Wen; Wang, Shaoli; Wu, Qingjun; Fang, Yong; Shi, Xiaobin; Zhang, Youjun
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.
Brittany L. Dodson
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.
Olagnier, David; Lababidi, Rassin R; Hadj, Samar Bel; Sze, Alexandre; Liu, Yiliu; Naidu, Sharadha Dayalan; Ferrari, Matteo; Jiang, Yuan; Chiang, Cindy; Beljanski, Vladimir; Goulet, Marie-Line; Knatko, Elena V; Dinkova-Kostova, Albena T; Hiscott, John; Lin, Rongtuan
Oncolytic viruses (OVs) offer a promising therapeutic approach to treat multiple types of cancer. In this study, we show that the manipulation of the antioxidant network via transcription factor Nrf2 augments vesicular stomatitis virus Δ51 (VSVΔ51) replication and sensitizes cancer cells to viral oncolysis. Activation of Nrf2 signaling by the antioxidant compound sulforaphane (SFN) leads to enhanced VSVΔ51 spread in OV-resistant cancer cells and improves the therapeutic outcome in different murine syngeneic and xenograft tumor models. Chemoresistant A549 lung cancer cells that display constitutive dominant hyperactivation of Nrf2 signaling are particularly vulnerable to VSVΔ51 oncolysis. Mechanistically, enhanced Nrf2 signaling stimulated viral replication in cancer cells and disrupted the type I IFN response via increased autophagy. This study reveals a previously unappreciated role for Nrf2 in the regulation of autophagy and the innate antiviral response that complements the therapeutic potential of VSV-directed oncolysis against multiple types of OV-resistant or chemoresistant cancer. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.
Dong, Rui; Zheng, Hui; Tian, Kun; Yau, Shek-Chung; Mao, Weiguang; Yu, Wenping; Yin, Changchuan; Yu, Chenglong; He, Rong Lucy; Yang, Jie; Yau, Stephen St
We construct a virus database called VirusDB (http://yaulab.math.tsinghua.edu.cn/VirusDB/) and an online inquiry system to serve people who are interested in viral classification and prediction. The database stores all viral genomes, their corresponding natural vectors, and the classification information of the single/multiple-segmented viral reference sequences downloaded from National Center for Biotechnology Information. The online inquiry system serves the purpose of computing natural vectors and their distances based on submitted genomes, providing an online interface for accessing and using the database for viral classification and prediction, and back-end processes for automatic and manual updating of database content to synchronize with GenBank. Submitted genomes data in FASTA format will be carried out and the prediction results with 5 closest neighbors and their classifications will be returned by email. Considering the one-to-one correspondence between sequence and natural vector, time efficiency, and high accuracy, natural vector is a significant advance compared with alignment methods, which makes VirusDB a useful database in further research.
LeBlanc, Amy K; Naik, Shruthi; Galyon, Gina D; Jenks, Nathan; Steele, Mike; Peng, Kah-Whye; Federspiel, Mark J; Donnell, Robert; Russell, Stephen J
VSV-IFNβ-NIS is a novel recombinant oncolytic vesicular stomatitis virus (VSV) with documented efficacy and safety in preclinical murine models of cancer. To facilitate clinical translation of this promising oncolytic therapy in patients with disseminated cancer, we are utilizing a comparative oncology approach to gather data describing the safety and efficacy of systemic VSV-IFNβ-NIS administration in dogs with naturally occurring cancer. In support of this, we executed a dose-escalation study in purpose-bred dogs to determine the maximum tolerated dose (MTD) of systemic VSV-hIFNβ-NIS, characterize the adverse event profile, and describe routes and duration of viral shedding in healthy, immune-competent dogs. The data indicate that an intravenous dose of 10(10) TCID50 is well tolerated in dogs. Expected adverse events were mild to moderate fever, self-limiting nausea and vomiting, lymphopenia, and oral mucosal lesions. Unexpected adverse events included prolongation of partial thromboplastin time, development of bacterial urinary tract infection, and scrotal dermatitis, and in one dog receiving 10(11) TCID50 (10 × the MTD), the development of severe hepatotoxicity and symptoms of shock leading to euthanasia. Viral shedding data indicate that detectable viral genome in blood diminishes rapidly with anti-VSV neutralizing antibodies detectable in blood as early as day 5 postintravenous virus administration. While low levels of viral genome copies were detectable in plasma, urine, and buccal swabs of dogs treated at the MTD, no infectious virus was detectable in plasma, urine, or buccal swabs at any of the doses tested. These studies confirm that VSV can be safely administered systemically in dogs, justifying the use of oncolytic VSV as a novel therapy for the treatment of canine cancer.
Mei, Yu; Zhang, Chunquan; Kernodle, Bliss M; Hill, John H; Whitham, Steven A
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.
Mei, Yu; Kernodle, Bliss M.; Hill, John H.
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
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.
Telmadarraiy, Zakkyeh; Chinikar, Sadegh; Vatandoost, Hassan; Faghihi, Faezeh; Hosseini-Chegeni, Asadollah
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. PMID:26623426
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.
Zhou, Y; Wen, F; Zhang, P; Tang, R; Li, Q
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.
Ogino, Minako; Ogino, Tomoaki
The GDP polyribonucleotidyltransferase (PRNTase) domain of the multifunctional L protein of rhabdoviruses, such as vesicular stomatitis virus (VSV) and rabies virus, catalyzes the transfer of 5'-phospho-RNA (pRNA) from 5'-triphospho-RNA (pppRNA) to GDP via a covalent enzyme-pRNA intermediate to generate a 5'-cap structure (GpppA). Here, using an improved oligo-RNA capping assay with the VSV L protein, we showed that the Michaelis constants for GDP and pppAACAG (VSV mRNA-start sequence) are 0.03 and 0.4 μM, respectively. A competition assay between GDP and GDP analogues in the GpppA formation and pRNA transfer assay using GDP analogues as pRNA acceptors indicated that the PRNTase domain recognizes the C-2-amino group, but not the C-6-oxo group, N-1-hydrogen, or N-7-nitrogen, of GDP for the cap formation. 2,6-Diaminopurine-riboside (DAP), 7-deazaguanosine (7-deaza-G), and 7-methylguanosine (m 7 G) diphosphates efficiently accepted pRNA, resulting in the formation of DAPpppA, 7-deaza-GpppA, and m 7 GpppA (cap 0), respectively. Furthermore, either the 2'- or 3'-hydroxyl group of GDP was found to be required for efficient pRNA transfer. A 5'-diphosphate form of antiviral ribavirin weakly inhibited the GpppA formation but did not act as a pRNA acceptor. These results indicate that the PRNTase domain has a unique guanosine-binding mode different from that of eukaryotic mRNA capping enzyme, guanylyltransferase. IMPORTANCE mRNAs of nonsegmented negative-strand (NNS) RNA viruses, such as VSV, possess a fully methylated cap structure, which is required for mRNA stability, efficient translation, and evasion of antiviral innate immunity in host cells. GDP polyribonucleotidyltransferase (PRNTase) is an unconventional mRNA capping enzyme of NNS RNA viruses that is distinct from the eukaryotic mRNA capping enzyme, guanylyltransferase. In this study, we studied the pRNA acceptor specificity of VSV PRNTase using various GDP analogues and identified chemical groups of GDP as
ABSTRACT The GDP polyribonucleotidyltransferase (PRNTase) domain of the multifunctional L protein of rhabdoviruses, such as vesicular stomatitis virus (VSV) and rabies virus, catalyzes the transfer of 5′-phospho-RNA (pRNA) from 5′-triphospho-RNA (pppRNA) to GDP via a covalent enzyme-pRNA intermediate to generate a 5′-cap structure (GpppA). Here, using an improved oligo-RNA capping assay with the VSV L protein, we showed that the Michaelis constants for GDP and pppAACAG (VSV mRNA-start sequence) are 0.03 and 0.4 μM, respectively. A competition assay between GDP and GDP analogues in the GpppA formation and pRNA transfer assay using GDP analogues as pRNA acceptors indicated that the PRNTase domain recognizes the C-2-amino group, but not the C-6-oxo group, N-1-hydrogen, or N-7-nitrogen, of GDP for the cap formation. 2,6-Diaminopurine-riboside (DAP), 7-deazaguanosine (7-deaza-G), and 7-methylguanosine (m7G) diphosphates efficiently accepted pRNA, resulting in the formation of DAPpppA, 7-deaza-GpppA, and m7GpppA (cap 0), respectively. Furthermore, either the 2′- or 3′-hydroxyl group of GDP was found to be required for efficient pRNA transfer. A 5′-diphosphate form of antiviral ribavirin weakly inhibited the GpppA formation but did not act as a pRNA acceptor. These results indicate that the PRNTase domain has a unique guanosine-binding mode different from that of eukaryotic mRNA capping enzyme, guanylyltransferase. IMPORTANCE mRNAs of nonsegmented negative-strand (NNS) RNA viruses, such as VSV, possess a fully methylated cap structure, which is required for mRNA stability, efficient translation, and evasion of antiviral innate immunity in host cells. GDP polyribonucleotidyltransferase (PRNTase) is an unconventional mRNA capping enzyme of NNS RNA viruses that is distinct from the eukaryotic mRNA capping enzyme, guanylyltransferase. In this study, we studied the pRNA acceptor specificity of VSV PRNTase using various GDP analogues and identified chemical groups
Matsushita, T; Elliger, S; Elliger, C; Podsakoff, G; Villarreal, L; Kurtzman, G J; Iwaki, Y; Colosi, P
The purpose of this work was to develop an efficient method for the production of adeno-associated virus (AAV) vectors in the absence of helper virus. The adenovirus regions that mediate AAV vector replication were identified and assembled into a helper plasmid. These included the VA, E2A and E4 regions. When this helper plasmid was cotransfected into 293 cells, along with plasmids encoding the AAV vector, and rep and cap genes, AAV vector was produced as efficiently as when using adenovirus infection as a source of help. CMV-driven constructs expressing the E4orf6 and the 72-M(r), E2A proteins were able to functionally replace the E4 and E2A regions, respectively. Therefore the minimum set of genes required to produce AAV helper activity equivalent to that provided by adenovirus infection consists of, or is a subset of, the following genes: the E4orf6 gene, the 72-M(r), E2A protein gene, the VA RNA genes and the E1 region. AAV vector preparations made with adenovirus and by the helper virus-free method were essentially indistinguishable with respect to particle density, particle to infectivity ratio, capsimer ratio and efficiency of muscle transduction in vivo. Only AAV vector preparations made by the helper virus-free method were not reactive with anti-adenovirus sera.
Full Text Available Abstract Background Sepsis remains the leading cause of death in critically ill patients. One of the primary organs affected by sepsis is the lung, presenting as the Acute Respiratory Distress Syndrome (ARDS. Organ damage in sepsis involves an alteration in gene expression, making gene transfer a potential therapeutic modality. This work examines the feasibility of applying simian virus 40 (SV40 vectors for pulmonary gene therapy. Methods Sepsis-induced ARDS was established by cecal ligation double puncture (2CLP. SV40 vectors carrying the luciferase reporter gene (SV/luc were administered intratracheally immediately after sepsis induction. Sham operated (SO as well as 2CLP rats given intratracheal PBS or adenovirus expressing luciferase served as controls. Luc transduction was evaluated by in vivo light detection, immunoassay and luciferase mRNA detection by RT-PCR in tissue harvested from septic rats. Vector abundance and distribution into alveolar cells was evaluated using immunostaining for the SV40 VP1 capsid protein as well as by double staining for VP1 and for the surfactant protein C (proSP-C. Immunostaining for T-lymphocytes was used to evaluate the cellular immune response induced by the vector. Results Luc expression measured by in vivo light detection correlated with immunoassay from lung tissue harvested from the same rats. Moreover, our results showed vector presence in type II alveolar cells. The vector did not induce significant cellular immune response. Conclusion In the present study we have demonstrated efficient uptake and expression of an SV40 vector in the lungs of animals with sepsis-induced ARDS. These vectors appear to be capable of in vivo transduction of alveolar type II cells and may thus become a future therapeutic tool.
Bergman, Ira; Whitaker-Dowling, Patricia; Gao Yanhua; Griffin, Judith A.; Watkins, Simon C.
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
Hruby, D E
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
He, Xiao-Chan; Xu, Hong-Xing; Zhou, Xiao-Jun; Zheng, Xu-Song; Sun, Yu-Jian; Yang, Ya-Jun; Tian, Jun-Ce; Lü, Zhong-Xian
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.
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.
Lévy, Camille; Amirache, Fouzia; Girard-Gagnepain, Anais; Frecha, Cecilia; Roman-Rodríguez, Francisco J; Bernadin, Ornellie; Costa, Caroline; Nègre, Didier; Gutierrez-Guerrero, Alejandra; Vranckx, Lenard S; Clerc, Isabelle; Taylor, Naomi; Thielecke, Lars; Cornils, Kerstin; Bueren, Juan A; Rio, Paula; Gijsbers, Rik; Cosset, François-Loïc; Verhoeyen, Els
Hematopoietic stem cell (HSC)-based gene therapy trials are now moving toward the use of lentiviral vectors (LVs) with success. However, one challenge in the field remains: efficient transduction of HSCs without compromising their stem cell potential. Here we showed that measles virus glycoprotein-displaying LVs (hemagglutinin and fusion protein LVs [H/F-LVs]) were capable of transducing 100% of early-acting cytokine-stimulated human CD34 + (hCD34 + ) progenitor cells upon a single application. Strikingly, these H/F-LVs also allowed transduction of up to 70% of nonstimulated quiescent hCD34 + cells, whereas conventional vesicular stomatitis virus G (VSV-G)-LVs reached 5% at the most with H/F-LV entry occurring exclusively through the CD46 complement receptor. Importantly, reconstitution of NOD/SCIDγc -/- (NSG) mice with H/F-LV transduced prestimulated or resting hCD34 + cells confirmed these high transduction levels in all myeloid and lymphoid lineages. Remarkably, for resting CD34 + cells, secondary recipients exhibited increasing transduction levels of up to 100%, emphasizing that H/F-LVs efficiently gene-marked HSCs in the resting state. Because H/F-LVs promoted ex vivo gene modification of minimally manipulated CD34 + progenitors that maintained stemness, we assessed their applicability in Fanconi anemia, a bone marrow (BM) failure with chromosomal fragility. Notably, only H/F-LVs efficiently gene-corrected minimally stimulated hCD34 + cells in unfractionated BM from these patients. These H/F-LVs improved HSC gene delivery in the absence of cytokine stimulation while maintaining their stem cell potential. Thus, H/F-LVs will facilitate future clinical applications requiring HSC gene modification, including BM failure syndromes, for which treatment has been very challenging up to now.
In the United States, Culicoides midges vector arboviruses of economic importance such as Bluetongue Virus and Epizootic Hemorrhagic Disease Virus. A limited number of studies have demonstrated the complexities of midge-virus interactions, including dynamic changes in virus titer and prevalence over...
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.
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
Full Text Available Spuma- or foamy viruses (FV, endemic in most non-human primates, cats, cattle and horses, comprise a special type of retrovirus that has developed a replication strategy combining features of both retroviruses and hepadnaviruses. Unique features of FVs include an apparent apathogenicity in natural hosts as well as zoonotically infected humans, a reverse transcription of the packaged viral RNA genome late during viral replication resulting in an infectious DNA genome in released FV particles and a special particle release strategy depending capsid and glycoprotein coexpression and specific interaction between both components. In addition, particular features with respect to the integration profile into the host genomic DNA discriminate FV from orthoretroviruses. It appears that some inherent properties of FV vectors set them favorably apart from orthoretroviral vectors and ask for additional basic research on the viruses as well as on the application in Gene Therapy. This review will summarize the current knowledge of FV biology and the development as a gene transfer system.
Full Text Available Zika virus (ZIKV has emerged as a new global health threat. Since its first discovery in Zika forest in Uganda, this virus has been isolated from several mosquito species, including Aedes aegypti and Aedes albopictus. The geographical distribution of these mosquito species across tropical and subtropical regions has led to several outbreaks, including the recent pandemic in Brazil, followed by the Pacific islands and other areas of North and South America. This has gained attention of the scientific community to elucidate the epidemiology and transmission of ZIKV. Despite its strong attention on clinical aspects for healthcare professionals, the relationships between ZIKV and its principal vectors, A. aegypti and A. albopictus, have not gained substantial interest in the scientific research community. As such, this review aims to summarize the current knowledge on ZIKV tropism and some important mechanisms which may be employed by the virus for effective strategies on viral survival in mosquitoes. In addition, this review identifies the areas of research that should be placed attention to, for which to be exploited for novel mosquito control strategies.
Tham, Hong-Wai; Balasubramaniam, Vinod; Ooi, Man K.; Chew, Miaw-Fang
Zika virus (ZIKV) has emerged as a new global health threat. Since its first discovery in Zika forest in Uganda, this virus has been isolated from several mosquito species, including Aedes aegypti and Aedes albopictus. The geographical distribution of these mosquito species across tropical and subtropical regions has led to several outbreaks, including the recent pandemic in Brazil, followed by the Pacific islands and other areas of North and South America. This has gained attention of the scientific community to elucidate the epidemiology and transmission of ZIKV. Despite its strong attention on clinical aspects for healthcare professionals, the relationships between ZIKV and its principal vectors, A. aegypti and A. albopictus, have not gained substantial interest in the scientific research community. As such, this review aims to summarize the current knowledge on ZIKV tropism and some important mechanisms which may be employed by the virus for effective strategies on viral survival in mosquitoes. In addition, this review identifies the areas of research that should be placed attention to, for which to be exploited for novel mosquito control strategies. PMID:29875751
Velazquez-Salinas, Lauro; Pauszek, Steven J; Zarate, Selene; Basurto-Alcantara, Francisco J; Verdugo-Rodriguez, Antonio; Perez, Andres M; Rodriguez, Luis L
We analyzed the phylogenetic and time-space relationships (phylodynamics) of 181 isolates of vesicular stomatitis New Jersey virus (VSNJV) causing disease in Mexico and the United States (US) from 2005 through 2012. We detail the emergence of a genetic lineage in southern Mexico causing outbreaks in central Mexico spreading into northern Mexico and eventually into the US. That emerging lineage showed higher nucleotide sequence identity (99.5%) than that observed for multiple lineages circulating concurrently in southern Mexico (96.8%). Additionally, we identified 58 isolates from Mexico that, unlike previous isolates from Mexico, grouped with northern Central America clade II viruses. This study provides the first direct evidence for the emergence and northward migration of a specific VSNJV genetic lineage from endemic areas in Mexico causing VS outbreaks in the US. In addition we document the emergence of a Central American VSNJV genetic lineage moving northward and causing outbreaks in central Mexico. © 2013 Published by Elsevier Inc.
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
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.
Yang, Lijuan; Sanchez, Anthony; Ward, Jerrold M; Murphy, Brian R; Collins, Peter L; Bukreyev, Alexander
Ebola virus (EBOV) causes outbreaks of a highly lethal hemorrhagic fever in humans. The virus can be transmitted by direct contact as well as by aerosol and is considered a potential bioweapon. Because direct immunization of the respiratory tract should be particularly effective against infection of mucosal surfaces, we previously developed an intranasal vaccine based on replication-competent human parainfluenza virus type 3 (HPIV3) expressing EBOV glycoprotein GP (HPIV3/EboGP) and showed that it is immunogenic and protective against a high dose parenteral EBOV challenge. However, because the adult human population has considerable immunity to HPIV3, which is a common human pathogen, replication and immunogenicity of the vaccine in this population might be greatly restricted. Indeed, in the present study, replication of the vaccine in the respiratory tract of HPIV3-immune guinea pigs was found to be restricted to undetectable levels. This restriction appeared to be based on both neutralizing antibodies and cellular or other components of the immunity to HPIV3. Surprisingly, even though replication of HPIV3/EboGP was highly restricted in HPIV3-immune animals, it induced a high level of EBOV-specific antibodies that nearly equaled that obtained in HPIV3-naive animals. We also show that the previously demonstrated presence of functional GP in the vector particle was not associated with increased replication in the respiratory tract nor with spread beyond the respiratory tract of HPIV3-naive guinea pigs, indicating that expression and functional incorporation of the attachment/penetration glycoprotein of this systemic virus did not mediate a change in tissue tropism.
Dingli, David; Peng, K.-W.; Harvey, Mary E.; Vongpunsawad, Sompong; Bergert, Elizabeth R.; Kyle, Robert A.; Cattaneo, Roberto; Morris, John C.; Russell, Stephen J.
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
Full Text Available Many viral pathogens are persistently transmitted by insect vectors and cause agricultural or health problems. Generally, an insect vector can use autophagy as an intrinsic antiviral defense mechanism against viral infection. Whether viruses can evolve to exploit autophagy to promote their transmission by insect vectors is still unknown. Here, we show that the autophagic process is triggered by the persistent replication of a plant reovirus, rice gall dwarf virus (RGDV in cultured leafhopper vector cells and in intact insects, as demonstrated by the appearance of obvious virus-containing double-membrane autophagosomes, conversion of ATG8-I to ATG8-II and increased level of autophagic flux. Such virus-containing autophagosomes seem able to mediate nonlytic viral release from cultured cells or facilitate viral spread in the leafhopper intestine. Applying the autophagy inhibitor 3-methyladenine or silencing the expression of Atg5 significantly decrease viral spread in vitro and in vivo, whereas applying the autophagy inducer rapamycin or silencing the expression of Torc1 facilitate such viral spread. Furthermore, we find that activation of autophagy facilitates efficient viral transmission, whereas inhibiting autophagy blocks viral transmission by its insect vector. Together, these results indicate a plant virus can induce the formation of autophagosomes for carrying virions, thus facilitating viral spread and transmission by its insect vector. We believe that such a role for virus-induced autophagy is common for vector-borne persistent viruses during their transmission by insect vectors.
Nasar, Farooq; Haddow, Andrew D; Tesh, Robert B; Weaver, Scott C
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
Alajez Nehad M
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.
Ferlin, Anna; Raux, Hélène; Baquero, Eduard; Lepault, Jean; Gaudin, Yves
Vesicular stomatitis virus (VSV; the prototype rhabdovirus) fusion is triggered at low pH and mediated by glycoprotein G, which undergoes a low-pH-induced structural transition. A unique feature of rhabdovirus G is that its conformational change is reversible. This allows G to recover its native prefusion state at the viral surface after its transport through the acidic Golgi compartments. The crystal structures of G pre- and postfusion states have been elucidated, leading to the identification of several acidic amino acid residues, clustered in the postfusion trimer, as potential pH-sensitive switches controlling the transition back toward the prefusion state. We mutated these residues and produced a panel of single and double mutants whose fusion properties, conformational change characteristics, and ability to pseudotype a virus lacking the glycoprotein gene were assayed. Some of these mutations were also introduced in the genome of recombinant viruses which were further characterized. We show that D268, located in the segment consisting of residues 264 to 273, which refolds into postfusion helix F during G structural transition, is the major pH sensor while D274, D395, and D393 have additional contributions. Furthermore, a single passage of recombinant virus bearing the mutation D268L (which was demonstrated to stabilize the G postfusion state) resulted in a pseudorevertant with a compensatory second mutation, L271P. This revealed that the propensity of the segment of residues 264 to 273 to refold into helix F has to be finely tuned since either an increase (mutation D268L alone) or a decrease (mutation L271P alone) of this propensity is detrimental to the virus. Vesicular stomatitis virus enters cells via endocytosis. Endosome acidification induces a structural transition of its unique glycoprotein (G), which mediates fusion between viral and endosomal membranes. G conformational change is reversible upon increases in pH. This allows G to recover its native
Richards, Stephanie L; Lord, Cynthia C; Pesko, Kendra N; Tabachnick, Walter J
Interactions between environmental and biological factors affect the vector competence of Culex pipiens quinquefasciatus for West Nile virus. Three age cohorts from two Cx. p. quinquefasciatus colonies were fed blood containing a low- or high-virus dose, and each group was held at two different extrinsic incubation temperatures (EIT) for 13 days. The colonies differed in the way that they responded to the effects of the environment on vector competence. The effects of mosquito age on aspects of vector competence were dependent on the EIT and dose, and they changed depending on the colony. Complex interactions must be considered in laboratory studies of vector competence, because the extent of the genetic and environmental variation controlling vector competence in nature is largely unknown. Differences in the environmental (EIT and dose) and biological (mosquito age and colony) effects from previous studies of Cx. p. quinquefasciatus vector competence for St. Louis encephalitis virus are discussed.
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.
Single-Injection Trivalent Filovirus 428 Vaccine: Proof of Concept Study in Outbred Guinea Pigs . J Infect Dis. 429 29. Murin, C. D., M. L. Fusco, Z...Jahrling, and J. F. Smith. 2000. Recombinant RNA replicons derived from attenuated 442 Venezuelan equine encephalitis virus protect guinea pigs and...platform, 65 including ease of production and characterization, absence of virus replication concerns and the 66 robust immune stimulatory activity
Pérez Jiménez, Eva; Larraga, Vicente; Esteban, Mariano
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,...
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 ...
Shiau, Ai-Li; Liu, Pu-Ste; Wu, Chao-Liang
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.
Liu, Zhuanzhuan; Zhou, Tengfei; Lai, Zetian; Zhang, Zhenhong; Jia, Zhirong; Zhou, Guofa; Williams, Tricia; Xu, Jiabao; Gu, Jinbao; Zhou, Xiaohong; Lin, Lifeng; Yan, Guiyun
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
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
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.
Carbonaro Sarracino, Denise; Tarantal, Alice F; Lee, C Chang I.; Martinez, Michele; Jin, Xiangyang; Wang, Xiaoyan; Hardee, Cinnamon L; Geiger, Sabine; Kahl, Christoph A; Kohn, Donald B
Systemic delivery of a lentiviral vector carrying a therapeutic gene represents a new treatment for monogenic disease. Previously, we have shown that transfer of the adenosine deaminase (ADA) cDNA in vivo rescues the lethal phenotype and reconstitutes immune function in ADA-deficient mice. In order to translate this approach to ADA-deficient severe combined immune deficiency patients, neonatal ADA-deficient mice and newborn rhesus monkeys were treated with species-matched and mismatched vectors and pseudotypes. We compared gene delivery by the HIV-1-based vector to murine γ-retroviral vectors pseudotyped with vesicular stomatitis virus-glycoprotein or murine retroviral envelopes in ADA-deficient mice. The vesicular stomatitis virus-glycoprotein pseudotyped lentiviral vectors had the highest titer and resulted in the highest vector copy number in multiple tissues, particularly liver and lung. In monkeys, HIV-1 or simian immunodeficiency virus vectors resulted in similar biodistribution in most tissues including bone marrow, spleen, liver, and lung. Simian immunodeficiency virus pseudotyped with the gibbon ape leukemia virus envelope produced 10- to 30-fold lower titers than the vesicular stomatitis virus-glycoprotein pseudotype, but had a similar tissue biodistribution and similar copy number in blood cells. The relative copy numbers achieved in mice and monkeys were similar when adjusted to the administered dose per kg. These results suggest that this approach can be scaled-up to clinical levels for treatment of ADA-deficient severe combined immune deficiency subjects with suboptimal hematopoietic stem cell transplantation options. PMID:24925206
Carbonaro Sarracino, Denise; Tarantal, Alice F; Lee, C Chang I; Martinez, Michele; Jin, Xiangyang; Wang, Xiaoyan; Hardee, Cinnamon L; Geiger, Sabine; Kahl, Christoph A; Kohn, Donald B
Systemic delivery of a lentiviral vector carrying a therapeutic gene represents a new treatment for monogenic disease. Previously, we have shown that transfer of the adenosine deaminase (ADA) cDNA in vivo rescues the lethal phenotype and reconstitutes immune function in ADA-deficient mice. In order to translate this approach to ADA-deficient severe combined immune deficiency patients, neonatal ADA-deficient mice and newborn rhesus monkeys were treated with species-matched and mismatched vectors and pseudotypes. We compared gene delivery by the HIV-1-based vector to murine γ-retroviral vectors pseudotyped with vesicular stomatitis virus-glycoprotein or murine retroviral envelopes in ADA-deficient mice. The vesicular stomatitis virus-glycoprotein pseudotyped lentiviral vectors had the highest titer and resulted in the highest vector copy number in multiple tissues, particularly liver and lung. In monkeys, HIV-1 or simian immunodeficiency virus vectors resulted in similar biodistribution in most tissues including bone marrow, spleen, liver, and lung. Simian immunodeficiency virus pseudotyped with the gibbon ape leukemia virus envelope produced 10- to 30-fold lower titers than the vesicular stomatitis virus-glycoprotein pseudotype, but had a similar tissue biodistribution and similar copy number in blood cells. The relative copy numbers achieved in mice and monkeys were similar when adjusted to the administered dose per kg. These results suggest that this approach can be scaled-up to clinical levels for treatment of ADA-deficient severe combined immune deficiency subjects with suboptimal hematopoietic stem cell transplantation options.
He, Wen-Bo; Li, Jie; Liu, Shu-Sheng
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.
Machida, Mayumi; Ambrozewicz, Marta A.; Breving, Kimberly; Wellman, Laurie L.; Yang, Linghui; Ciavarra, Richard P.; Sanford, Larry D.
Intranasal application of vesicular stomatitis virus (VSV) produces a well-characterized model of viral encephalitis in mice. Within one day post-infection (PI), VSV travels to the olfactory bulb and, over the course of 7 days, it infects regions and tracts extending into the brainstem followed by clearance and recovery in most mice by PI day 14 (PI 14). Infectious diseases are commonly accompanied by excessive sleepiness; thus, sleep is considered a component of the acute phase response to infection. In this project, we studied the relationship between sleep and VSV infection using C57BL/6 (B6) and BALB/c mice. Mice were implanted with transmitters for recording EEG, activity and temperature by telemetry. After uninterrupted baseline recordings were collected for 2 days, each animal was infected intranasally with a single low dose of VSV (5 × 104 PFU). Sleep was recorded for 15 consecutive days and analyzed on PI 0, 1, 3, 5, 7, 10, and 14. Compared to baseline, amounts of non-rapid eye movement sleep (NREM) were increased in B6 mice during the dark period of PI 1–5, whereas rapid eye movement sleep (REM) was significantly reduced during the light periods of PI 0–14. In contrast, BALB/c mice showed significantly fewer changes in NREM and REM. These data demonstrate sleep architecture is differentially altered in these mouse strains and suggests that, in B6 mice, VSV can alter sleep before virus progresses into brain regions that control sleep. PMID:24055862
Johnson, J E; Rodgers, W; Rose, J K
Previous studies showed that the HIV-1 envelope (Env) protein was not incorporated into vesicular stomatitis virus (VSV) virions unless its cytoplasmic tail was replaced with that of the VSV glycoprotein (G). To determine whether the G tail provided a positive incorporation signal for Env, or if sequences in the Env tail prevented incorporation, we generated mutants of Env with its 150-amino-acid tail shortened to 29, 10, or 3 amino acids (Envtr mutants). Cells infected with VSV recombinants expressing these proteins or an Env-G tail hybrid showed similar amounts of Env protein at the surface. The Env-G tail hybrid or the Envtr3 mutant were incorporated at the highest levels into budding VSV virions. In contrast, the Envtr29 or Envtr10 mutants were incorporated poorly. These results defined a signal preventing incorporation within the 10 membrane-proximal amino acids of the Env tail. Confocal microscopy revealed that this signal functioned by causing localization of human immunodeficiency virus type 1 Env to plasma membrane domains distinct from the VSV budding sites, where VSV proteins were concentrated. Copyright 1998 Academic Press.
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.
Lou, Fangfei; Neetoo, Hudaa; Li, Junan; Chen, Haiqiang; Li, Jianrong
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.
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 clinica...
Japanese encephalitis virus (JEV) is a virus of the Flavivirus genus that may result in encephalitis in human hosts. This vector-borne zoonosis occurs in Eastern and Southeastern Asia and an intentional or inadvertent introduction into the United States (US) will have major public health and economi...
Insect pests and disease infestations are the primary constraints in rice (Oryza sativa) production .... Asia. Of all the rice diseases, the one caused by the rice yellow mottle virus (RYMV), first reported ..... yellow mottle virus in Central Africa.
Recombinant pox viruses have been generated for vaccination against heterologous pathogens. Amongst these, the following are notable examples. (i) The engineering of the Copenhagen strain of vaccinia virus to express the rabies virus glycoprotein. When applied in baits, this recombinant has been shown to vaccinate the red fox in Europe and raccoons in the United States, stemming the spread of rabies virus infection in the wild. (ii) A fowlpox-based recombinant expressing the Newcastle disease...
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
Brown, Erica L.; Lyles, Douglas S.
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
Kanakala, Surapathrudu; Ghanim, Murad
Insects and other arthropods are the most important vectors of plant pathogens. The majority of plant pathogens are disseminated by arthropod vectors such as aphids, beetles, leafhoppers, planthoppers, thrips and whiteflies. Transmission of plant pathogens and the challenges in managing insect vectors due to insecticide resistance are factors that contribute to major food losses in agriculture. RNA interference (RNAi) was recently suggested as a promising strategy for controlling insect pests...
Blagrove, Marcus S C; Caminade, Cyril; Waldmann, Elisabeth; Sutton, Elizabeth R; Wardeh, Maya; Baylis, Matthew
Mosquito-borne viruses have been estimated to cause over 100 million cases of human disease annually. Many methodologies have been developed to help identify areas most at risk from transmission of these viruses. However, generally, these methodologies focus predominantly on the effects of climate on either the vectors or the pathogens they spread, and do not consider the dynamic interaction between the optimal conditions for both vector and virus. Here, we use a new approach that considers the complex interplay between the optimal temperature for virus transmission, and the optimal climate for the mosquito vectors. Using published geolocated data we identified temperature and rainfall ranges in which a number of mosquito vectors have been observed to co-occur with West Nile virus, dengue virus or chikungunya virus. We then investigated whether the optimal climate for co-occurrence of vector and virus varies between "warmer" and "cooler" adapted vectors for the same virus. We found that different mosquito vectors co-occur with the same virus at different temperatures, despite significant overlap in vector temperature ranges. Specifically, we found that co-occurrence correlates with the optimal climatic conditions for the respective vector; cooler-adapted mosquitoes tend to co-occur with the same virus in cooler conditions than their warmer-adapted counterparts. We conclude that mosquitoes appear to be most able to transmit virus in the mosquitoes' optimal climate range, and hypothesise that this may be due to proportionally over-extended vector longevity, and other increased fitness attributes, within this optimal range. These results suggest that the threat posed by vector-competent mosquito species indigenous to temperate regions may have been underestimated, whilst the threat arising from invasive tropical vectors moving to cooler temperate regions may be overestimated.
Marconi, Peggy; Fraefel, Cornel; Epstein, Alberto L
Herpes simplex virus type 1 (HSV-1 ) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153-kilobase pair (kbp) double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes the approach most commonly used to prepare recombinant HSV-1 vectors through homologous recombination, either in eukaryotic cells or in bacteria.
Yao Yi; Ghosh, Kakoli; Epand, Raquel F.; Epand, Richard M.; Ghosh, Hara P.
The fusogenic envelope glycoprotein G of the rhabdovirus vesicular stomatitis virus (VSV) induces membrane fusion at acidic pH. At acidic pH the G protein undergoes a major structural reorganization leading to the fusogenic conformation. However, unlike other viral fusion proteins, the low-pH-induced conformational change of VSV G is completely reversible. As well, the presence of an α-helical coiled-coil motif required for fusion by a number of viral and cellular fusion proteins was not predicted in VSV G protein by using a number of algorithms. Results of pH dependence of the thermal stability of G protein as determined by intrinsic Trp fluorescence and circular dichroism (CD) spectroscopy show that the G protein is equally stable at neutral or acidic pH. Destabilization of G structure at neutral pH with either heat or urea did not induce membrane fusion or conformational change(s) leading to membrane fusion. Taken together, these data suggest that the mechanism of VSV G-induced fusion is distinct from the fusion mechanism of fusion proteins that involve a coiled-coil motif
Dutto, M; Bertero, M; Petrosillo, N; Pombi, M; Otranto, D
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.
Tomato spotted wilt virus (TSWV), member of the genus Tospovirus within the family Bunyaviridae, ranks among the top ten of economically most important plant viruses. Tospoviruses cause significant yield losses in agricultural crops such as tomato,
Podsakoff, G; Wong, K K; Chatterjee, S
Gene transfer vectors based on adeno-associated virus (AAV) are emerging as highly promising for use in human gene therapy by virtue of their characteristics of wide host range, high transduction efficiencies, and lack of cytopathogenicity. To better define the biology of AAV-mediated gene transfer, we tested the ability of an AAV vector to efficiently introduce transgenes into nonproliferating cell populations. Cells were induced into a nonproliferative state by treatment with the DNA synthesis inhibitors fluorodeoxyuridine and aphidicolin or by contact inhibition induced by confluence and serum starvation. Cells in logarithmic growth or DNA synthesis arrest were transduced with vCWR:beta gal, an AAV-based vector encoding beta-galactosidase under Rous sarcoma virus long terminal repeat promoter control. Under each condition tested, vCWR:beta Gal expression in nondividing cells was at least equivalent to that in actively proliferating cells, suggesting that mechanisms for virus attachment, nuclear transport, virion uncoating, and perhaps some limited second-strand synthesis of AAV vectors were present in nondividing cells. Southern hybridization analysis of vector sequences from cells transduced while in DNA synthetic arrest and expanded after release of the block confirmed ultimate integration of the vector genome into cellular chromosomal DNA. These findings may provide the basis for the use of AAV-based vectors for gene transfer into quiescent cell populations such as totipotent hematopoietic stem cells.
Bialosuknia, Sean M.; Zink, Steven D.; Brecher, Matthew; Ehrbar, Dylan J.; Morrissette, Madeline N.; Kramer, Laura D.
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
Ciota, Alexander T; Bialosuknia, Sean M; Zink, Steven D; Brecher, Matthew; Ehrbar, Dylan J; Morrissette, Madeline N; Kramer, Laura D
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 log 10 PFU/mL; minimum infective dose was 4.2 log 10 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.
Full Text Available Insects and other arthropods are the most important vectors of plant pathogens. The majority of plant pathogens are disseminated by arthropod vectors such as aphids, beetles, leafhoppers, planthoppers, thrips and whiteflies. Transmission of plant pathogens and the challenges in managing insect vectors due to insecticide resistance are factors that contribute to major food losses in agriculture. RNA interference (RNAi was recently suggested as a promising strategy for controlling insect pests, including those that serve as important vectors for plant pathogens. The last decade has witnessed a dramatic increase in the functional analysis of insect genes, especially those whose silencing results in mortality or interference with pathogen transmission. The identification of such candidates poses a major challenge for increasing the role of RNAi in pest control. Another challenge is to understand the RNAi machinery in insect cells and whether components that were identified in other organisms are also present in insect. This review will focus on summarizing success cases in which RNAi was used for silencing genes in insect vector for plant pathogens, and will be particularly helpful for vector biologists.
Uchida, Hiroaki; Hamada, Hirofumi; Nakano, Kenji; Kwon, Heechung; Tahara, Hideaki; Cohen, Justus B; Glorioso, Joseph C
Oncolytic virotherapy is a novel therapeutic modality for malignant diseases that exploits selective viral replication in cancer cells. Herpes simplex virus (HSV) is a promising agent for oncolytic virotherapy due to its broad cell tropism and the identification of mutations that favor its replication in tumor over normal cells. However, these attenuating mutations also tend to limit the potency of current oncolytic HSV vectors that have entered clinical studies. As an alternative, vector retargeting to novel entry receptors has the potential to achieve tumor specificity at the stage of virus entry, eliminating the need for replication-attenuating mutations. Here, we summarize the molecular mechanism of HSV entry and recent advances in the development of fully retargeted HSV vectors for oncolytic virotherapy. Retargeted HSV vectors offer an attractive platform for the creation of a new generation of oncolytic HSV with improved efficacy and specificity. Copyright© Bentham Science Publishers; For any queries, please email at email@example.com.
et al. 1996; Lee, Groebner et al. 2006; Jones, Feldmann et al. 2005). However, NHPs display disease characteristics such as clinical disease, and...2003; Jones, Feldmann et al. 2005; Wang, Schmaljohn et al. 2006; Lee, Groebner et al. 2006). Among the successful vaccine platforms evaluated in...primary influenza infection and helps to prevent reinfection." J Immunol 175(9): 5827-38. 152 Lee, J. S., J. L. Groebner , et al. (2006). "Multiagent
Podsakoff, G; Wong, K K; Chatterjee, S
Gene transfer vectors based on adeno-associated virus (AAV) are emerging as highly promising for use in human gene therapy by virtue of their characteristics of wide host range, high transduction efficiencies, and lack of cytopathogenicity. To better define the biology of AAV-mediated gene transfer, we tested the ability of an AAV vector to efficiently introduce transgenes into nonproliferating cell populations. Cells were induced into a nonproliferative state by treatment with the DNA synthe...
Reiss, Carol Shoshkes
James M Miller1, Sarah McNulty Bidula1,5, Troels Mygind Jensen1,6, Carol Shoshkes Reiss1,2,3,41Department of Biology, New York University, New York, NY, USA; 2Center for Neural Science, NYU; 3NYU Cancer Institute; 4Departments of Microbiology, NYU School of Medicine and Mt Sinai School of Medicine, New York, NY, USA; 5Present address Graduate Program, University of Pittsburgh School of Medicine, Pittsburgh, PA,USA 6Present address: Univercity of Copenhagen, Copenhagen, DenmarkAbstract: Viruse...
Sotomayor-Bonilla, Jesús; Abella-Medrano, Carlos Antonio; Chaves, Andrea; Álvarez-Mendizábal, Paulina; Rico-Chávez, Óscar; Ibáñez-Bernal, Sergio; Rostal, Melinda K; Ojeda-Flores, Rafael; Barbachano-Guerrero, Arturo; Gutiérrez-Espeleta, Gustavo; Aguirre, A Alonso; Daszak, Peter; Suzán, Gerardo
Arboviruses are important zoonotic agents with complex transmission cycles and are not well understood because they may involve many vectors and hosts. We studied sympatric wild mammals and hematophagous mosquitoes having the potential to act as hosts and vectors in two areas of southern Mexico. Mosquitoes, bats, and rodents were captured in Calakmul (Campeche) and Montes Azules (Chiapas), between November 2010 and August 2011. Spleen samples from 146 bats and 14 rodents were tested for molecular evidence of Venezuelan equine encephalitis virus (VEEV), eastern equine encephalitis virus (EEEV), western equine encephalitis virus (WEEV), and West Nile virus (WNV) using PCR protocols. Bat ( Artibeus lituratus , Carollia sowelli , Glossophaga soricina , and Sturnira parvidens) and rodent ( Sigmodon hispidus and Oryzomys alfaroi ) species were positive for VEEV. No individuals were positive for WNV, EEEV, or WEEV. A total of 1,298 mosquitoes were collected at the same sites, and five of the mosquito species collected were known VEEV vectors (Aedes fulvus, Mansonia indubitans, Psorophora ferox, Psorophora cilipes, and Psorophora confinnis). This survey simultaneously presents the first molecular evidence, to our knowledge, of VEEV in bats and rodents from southern Mexico and the identification of potential sympatric vectors. Studies investigating sympatric nonhuman hosts, vectors, and arboviruses must be expanded to determine arboviral dynamics in complex systems in which outbreaks of emerging and reemerging zoonoses are continuously occurring.
Yu, Jing; Qi, Mengchun; Deng, Jiupeng; Liu, Gang; Chen, Huaiqing
This experiment was designed to construct mouse Smad6 recombinant RNA interference vectors and determine their interference effects on bone marrow mesenchymal stem cells (BMSCs). Three recombinant Smad6 RNA interference vectors were constructed by molecular clone techniques with a lenti-virus vector expressing green fluorescent protein (GFP), and the correctness of recombinant vectors was verified by DNA sequencing. Mouse BMSCs were used for transfection experiments and BMP-2 was in use for osteogenic induction of MSCs. The transfection efficiency of recombinant vectors was examined by Laser confocal scanning microscope and the interference effect of recombinant vectors on Smad6 gene expression was determined by real-time RT-PCR and Western blot, respectively. Three Smad6 recombinant RNA interference vectors were successfully constructed and their correctness was proved by DNA sequencing. After transfection, GFPs were effectively expressed in MSCs and all of three recombinant vectors gained high transfection efficiency (> 95%). Both real-time PCR and Western blot examination indicated that among three recombinant vectors, No. 2 Svector had the best interference effect and the interference effect was nearly 91% at protein level. In conclusion, Mouse recombinant Smad6 RNA interference (RNAi) vector was successfully constructed and it provided an effective tool for further studies on BMP signal pathways.
Nakai, Hiroyuki; Fuess, Sally; Storm, Theresa A.; Muramatsu, Shin-ichi; Nara, Yuko; Kay, Mark A.
Recombinant adeno-associated virus (rAAV) vectors can mediate long-term stable transduction in various target tissues. However, with rAAV serotype 2 (rAAV2) vectors, liver transduction is confined to only a small portion of hepatocytes even after administration of extremely high vector doses. In order to investigate whether rAAV vectors of other serotypes exhibit similar restricted liver transduction, we performed a dose-response study by injecting mice with β-galactosidase-expressing rAAV1 and rAAV8 vectors via the portal vein. The rAAV1 vector showed a blunted dose-response similar to that of rAAV2 at high doses, while the rAAV8 vector dose-response remained unchanged at any dose and ultimately could transduce all the hepatocytes at a dose of 7.2 × 1012 vector genomes/mouse without toxicity. This indicates that all hepatocytes have the ability to process incoming single-stranded vector genomes into duplex DNA. A single tail vein injection of the rAAV8 vector was as efficient as portal vein injection at any dose. In addition, intravascular administration of the rAAV8 vector at a high dose transduced all the skeletal muscles throughout the body, including the diaphragm, the entire cardiac muscle, and substantial numbers of cells in the pancreas, smooth muscles, and brain. Thus, rAAV8 is a robust vector for gene transfer to the liver and provides a promising research tool for delivering genes to various target organs. In addition, the rAAV8 vector may offer a potential therapeutic agent for various diseases affecting nonhepatic tissues, but great caution is required for vector spillover and tight control of tissue-specific gene expression. PMID:15596817
Andersen, C; Jensen, T; Nansen, A
experiments using B cell- and T cell-deficient recipients revealed that no protection could be obtained in the absence of B cells, whereas treatment with virus-specific immune (IgG) serum controlled viral spreading to the central nervous system (CNS), but did not necessarily accomplish virus elimination......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...... the capacity to secrete both IFN-gamma and perforin completely resisted disease. Similar results were obtained using IL-4 knockout mice, indicating that neither cell-mediated nor T(h)2-dependent effector systems were required. In contrast, mice deficient in expression of CD40 ligand were more susceptible than...
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.
Rivera, C; Gámez, R
The enzyme-linked immunosorbent assay (ELISA) was used to demonstrate the increase in titer of maize rayado fino virus (MRFV) in its leafhopper vector, Dalbulus maidis. Viral antigen concentration attained a maximum in the body of the insect 25 days after virus acquisition and decreased thereafter. Substantial differences in concentration were observed among viruliferous leafhoppers. MRFV was serially passed through 5 successive leafhopper populations. The results provide further evidence of multiplication of MRFV in D. maidis.
Díaz Desani, Beatriz M.; Biurrun, R.; Moreno, Aránzazu; Nebreda, Miguel; Fereres, Alberto
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...
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.
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.
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.
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. PMID:26941044
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.
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
Lawrence, M S; Ho, D Y; Dash, R; Sapolsky, R M
We have generated herpes simplex virus (HSV) vectors vIE1GT and v alpha 4GT bearing the GLUT-1 isoform of the rat brain glucose transporter (GT) under the control of the human cytomegalovirus ie1 and HSV alpha 4 promoters, respectively. We previously reported that such vectors enhance glucose uptake in hippocampal cultures and the hippocampus. In this study we demonstrate that such vectors can maintain neuronal metabolism and reduce the extent of neuron loss in cultures after a period of hypo...
Valles, S.M.; Porter, S.D.
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
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
Schultz Ronald D
Full Text Available Abstract Background Potato virus X has been developed into an expression vector for plants. It is widely used to express foreign genes. In molecular manipulation, the foreign genes need to be sub-cloned into the vector. The constructed plasmid needs to be amplified. Usually, during amplification stage, the foreign genes are not expressed. However, if the foreign gene is expressed, the construction work could be interrupted. Two different viral genes were sub-cloned into the vector, but only one foreign gene was successfully sub-cloned. The other foreign gene, canine parvovirus type 2 (CPV-2 VP1 could not be sub-cloned into the vector and amplified without mutation (frame shift mutation. Results A cryptic promoter in the PVX vector was discovered with RT-PCR. The promoter activity was studied with Northern blots and Real-time RT-PCR. Conclusion It is important to recognize the homologous promoter sequences in the vector when a virus is developed as an expression vector. During the plasmid amplification stage, an unexpected expression of the CPV-2 VP1 gene (not in the target plants, but in E. coli can interrupt the downstream work.
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 ...
Doumayrou, Juliette; Sheber, Melissa; Bonning, Bryony C; Miller, W Allen
Understanding the molecular mechanisms involved in plant virus-vector interactions is essential for the development of effective control measures for aphid-vectored epidemic plant diseases. The coat proteins (CP) are the main component of the viral capsids, and they are implicated in practically every stage of the viral infection cycle. Pea enation mosaic virus 1 (PEMV1, Enamovirus , Luteoviridae ) and Pea enation mosaic virus 2 (PEMV2, Umbravirus , Tombusviridae ) are two RNA viruses in an obligate symbiosis causing the pea enation mosaic disease. Sixteen mutant viruses were generated with mutations in different domains of the CP to evaluate the role of specific amino acids in viral replication, virion assembly, long-distance movement in Pisum sativum , and aphid transmission. Twelve mutant viruses were unable to assemble but were able to replicate in inoculated leaves, move long-distance, and express the CP in newly infected leaves. Four mutant viruses produced virions, but three were not transmissible by the pea aphid, Acyrthosiphon pisum . Three-dimensional modeling of the PEMV CP, combined with biological assays for virion assembly and aphid transmission, allowed for a model of the assembly of PEMV coat protein subunits.
Duch, M.; Carrasco, M.L.; Jespersen, T.
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...
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...
We examined the chemical composition of garlic and asafoetida essential oils and their individual and combined toxicity against larvae of two West Nile virus vectors, Culex pipiens pipiens and Cx. restuans. The effect of the two essential oils on egg hatch was also examined. Ten and twelve compounds...
Haller, Aurelia A.; Miller, Tessa; Mitiku, Misrach; Coelingh, Kathleen
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...
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.
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.
Full Text Available Understanding the molecular mechanisms involved in plant virus–vector interactions is essential for the development of effective control measures for aphid-vectored epidemic plant diseases. The coat proteins (CP are the main component of the viral capsids, and they are implicated in practically every stage of the viral infection cycle. Pea enation mosaic virus 1 (PEMV1, Enamovirus, Luteoviridae and Pea enation mosaic virus 2 (PEMV2, Umbravirus, Tombusviridae are two RNA viruses in an obligate symbiosis causing the pea enation mosaic disease. Sixteen mutant viruses were generated with mutations in different domains of the CP to evaluate the role of specific amino acids in viral replication, virion assembly, long-distance movement in Pisum sativum, and aphid transmission. Twelve mutant viruses were unable to assemble but were able to replicate in inoculated leaves, move long-distance, and express the CP in newly infected leaves. Four mutant viruses produced virions, but three were not transmissible by the pea aphid, Acyrthosiphon pisum. Three-dimensional modeling of the PEMV CP, combined with biological assays for virion assembly and aphid transmission, allowed for a model of the assembly of PEMV coat protein subunits.
Sakkas, Hercules; Economou, Vangelis; Papadopoulou, Chrissanthy
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.
Alfred O. Ochieng
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.
Liu, Wenwen; Gray, Stewart; Huo, Yan; Li, Li; Wei, Taiyun; Wang, Xifeng
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.
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.
Trawinski, Patricia R; Mackay, D Scott
The rapid spread of West Nile virus (WNv) in North America is a major public health concern. Culex pipiens-restuans is the principle mosquito vector of WNv in the northeastern United States while Aedes vexans is an important bridge vector of the virus in this region. Vector mosquito abundance is directly dependent on physical environmental factors that provide mosquito habitats. The objective of this research is to determine landscape elements that explain the population abundance and distribution of WNv vector mosquitoes using stepwise linear regression. We developed a novel approach for examining a large set of landscape variables based on a land use and land cover classification by selecting variables in stages to minimize multicollinearity. We also investigated the distance at which landscape elements influence abundance of vector populations using buffer distances of 200, 400, and 1000 m. Results show landscape effects have a significant impact on Cx. pipiens-estuans population distribution while the effects of landscape features are less important for prediction of Ae. vexans population distributions. Cx. pipiens-restuans population abundance is positively correlated with human population density, housing unit density, and urban land use and land cover classes and negatively correlated with age of dwellings and amount of forested land.
Zhou Qi; Schneider, Irene C.; Gallet, Manuela; Kneissl, Sabrina; Buchholz, Christian J.
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.
Diaz, James H
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 Â© 2016 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.
Boeriis, Morten; van Leeuwen, Theo
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......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...
Gentle, J. N., Jr.; Kahn, A.; Pierce, S. A.; Wang, S.; Wade, C.; Moran, S.
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.
Suwanmanee, San; Luplertlop, Natthanej
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.
Ruder Mark G
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
Ruder, Mark G; Howerth, Elizabeth W; Stallknecht, David E; Allison, Andrew B; Carter, Deborah L; Drolet, Barbara S; Klement, Eyal; Mead, Daniel G
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. 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 ≥ 10(2.7) median tissue culture infective doses (TCID(50))/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. From 4-16 dpf, 45% (156/350) of midges that fed on WTD with high titer viremia (>10(7) TCID(50)/ml) were virus isolation-positive, and starting from 10-16 dpf, 32% (35/109) of these virus isolation-positive midges were potentially competent (≥ 10(2.7) TCID(50)/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. 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 that North America has a susceptible ruminant and
Full Text Available Mosquito feeding behaviour determines the degree of vector-host contact and may have a serious impact on the risk of West Nile virus (WNV epidemics. Feeding behaviour also interacts with other biotic and abiotic factors that affect virus amplification and transmission.We identified the origin of blood meals in five mosquito species from three different wetlands in SW Spain. All mosquito species analysed fed with different frequencies on birds, mammals and reptiles. Both 'mosquito species' and 'locality' explained a similar amount of variance in the occurrence of avian blood meals. However, 'season of year' was the main factor explaining the presence of human blood meals. The differences in diet resulted in a marked spatial heterogeneity in the estimated WNV transmission risk. Culex perexiguus, Cx. modestus and Cx. pipiens were the main mosquito species involved in WNV enzootic circulation since they feed mainly on birds, were abundant in a number of localities and had high vector competence. Cx. perexiguus may also be important for WNV transmission to horses, as are Cx. pipiens and Cx. theileri in transmission to humans. Estimates of the WNV transmission risk based on mosquito diet, abundance and vector competence matched the results of previous WNV monitoring programs in the area. Our sensitivity analyses suggested that mosquito diet, followed by mosquito abundance and vector competence, are all relevant factors in understanding virus amplification and transmission risk in the studied wild ecosystems. At some of the studied localities, the risk of enzootic circulation of WNV was relatively high, even if the risk of transmission to humans and horses was less.Our results describe for first time the role of five WNV candidate vectors in SW Spain. Interspecific and local differences in mosquito diet composition has an important effect on the potential transmission risk of WNV to birds, horses and humans.
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
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.
Strive, T; Hardy, C M; Wright, J; Reubel, G H
Canine Herpesvirus (CHV) is being developed as a virus vector for the vaccination of European red foxes. However, initial studies using recombinant CHV vaccines in foxes revealed viral attenuation and lack of antibody response to inserted foreign antigens. These findings were attributed both to inactivation of the thymidine kinase (TK) gene and excess foreign genetic material in the recombinant viral genome. In this study, we report an improved CHV-bacterial artificial chromosome (BAC) vector system designed to overcome attenuation in foxes. A non-essential region was identified in the CHV genome as an alternative insertion site for foreign genes. Replacement of a guanine/cytosine (GC)-rich intergenic region between UL21 and UL22 of CHV with a marker gene did not change growth behaviour in vitro, showing that this region is not essential for virus growth in cell culture. We subsequently produced a CHV-BAC vector with an intact TK gene in which the bacterial genes and the antigen expression cassette were inserted into this GC-rich locus. Unlike earlier constructs, the new CHV-BAC allowed self-excision of the bacterial genes via homologous recombination after transfection of BACs into cell culture. The BAC-CHV system was used to produce a recombinant virus that constitutively expressed porcine zona pellucida subunit C protein between the UL21 and UL22 genes of CHV. Complete self-excision of the bacterial genes from CHV was achieved within one round of replication whilst retaining antigen gene expression.
Grigg, Patricia; Titong, Allison; Jones, Leslie A; Yilma, Tilahun D; Verardi, Paulo H
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.
Fujino, Kan; Yamamoto, Yusuke; Daito, Takuji; Makino, Akiko; Honda, Tomoyuki; Tomonaga, Keizo
Borna disease virus (BoDV), a prototype of mammalian bornavirus, is a non-segmented, negative strand RNA virus that often causes severe neurological disorders in infected animals, including horses and sheep. Unique among animal RNA viruses, BoDV transcribes and replicates non-cytopathically in the cell nucleus, leading to establishment of long-lasting persistent infection. This striking feature of BoDV indicates its potential as an RNA virus vector system. It has previously been demonstrated by our team that recombinant BoDV (rBoDV) lacking an envelope glycoprotein (G) gene develops persistent infections in transduced cells without loss of the viral genome. In this study, a novel non-transmissive rBoDV, rBoDV ΔMG, which lacks both matrix (M) and G genes in the genome, is reported. rBoDV-ΔMG expressing green fluorescence protein (GFP), rBoDV ΔMG-GFP, was efficiently generated in Vero/MG cells stably expressing both BoDV M and G proteins. Infection with rBoDV ΔMG-GFP was persistently maintained in the parent Vero cells without propagation within cell culture. The optimal ratio of M and G for efficient viral particle production by transient transfection of M and G expression plasmids into cells persistently infected with rBoDV ΔMG-GFP was also demonstrated. These findings indicate that the rBoDV ΔMG-based BoDV vector may provide an extremely safe virus vector system and could be a novel strategy for investigating the function of M and G proteins and the host range of bornaviruses. © 2017 The Societies and John Wiley & Sons Australia, Ltd.
Thompson, Sarah J.; Pearce, John; Ramey, Andy M.
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.
Lai, Huafang; He, Junyun; Engle, Michael; Diamond, Michael S.; Chen, Qiang
Summary Pharmaceutical protein production in plants has been greatly promoted by the development of viral-based vectors and transient expression systems. Tobacco and related Nicotiana species are currently the most common host plants for generation of plant-made pharmaceutical proteins (PMPs). Downstream processing of target PMPs from these plants, however, is hindered by potential technical and regulatory difficulties due to the presence of high levels of phenolics and toxic alkaloids. Here, we explored the use of lettuce, which grows quickly yet produces low levels of secondary metabolites, and viral vector-based transient expression systems to develop a robust PMP production platform. Our results showed that a geminiviral replicon system based on the bean yellow dwarf virus permits high-level expression in lettuce of virus-like particles (VLP) derived from the Norwalk virus capsid protein and therapeutic monoclonal antibodies (mAbs) against Ebola and West Nile viruses. These vaccine and therapeutic candidates can be readily purified from lettuce leaves with scalable processing methods while fully retaining functional activity. Furthermore, this study also demonstrated the feasibility of using commercially produced lettuce for high-level PMP production. This allows our production system to have access to unlimited quantities of inexpensive plant material for large-scale production. These results establish a new production platform for biological pharmaceutical agents that is effective, safe, low-cost, and amenable to large-scale manufacturing. PMID:21883868
Aponte-Ubillus, Juan Jose; Barajas, Daniel; Peltier, Joseph; Bardliving, Cameron; Shamlou, Parviz; Gold, Daniel
Recombinant adeno-associated virus (rAAV) vectors are increasingly popular tools for gene therapy applications. Their non-pathogenic status, low inflammatory potential, availability of viral serotypes with different tissue tropisms, and prospective long-lasting gene expression are important attributes that make rAAVs safe and efficient therapeutic options. Over the last three decades, several groups have engineered recombinant AAV-producing platforms, yielding high titers of transducing vector particles. Current specific productivity yields from different platforms range from 10 3 to 10 5 vector genomes (vg) per cell, and there is an ongoing effort to improve vector yields in order to satisfy high product demands required for clinical trials and future commercialization.Crucial aspects of vector production include the molecular design of the rAAV-producing host cell line along with the design of AAV genes, promoters, and regulatory elements. Appropriately, configuring and balancing the expression of these elements not only contributes toward high productivity, it also improves process robustness and product quality. In this mini-review, the rational design of rAAV-producing expression systems is discussed, with special attention to molecular strategies that contribute to high-yielding, biomanufacturing-amenable rAAV production processes. Details on molecular optimization from four rAAV expression systems are covered: adenovirus, herpesvirus, and baculovirus complementation systems, as well as a recently explored yeast expression system.
Fisher-Adams, G; Wong, K K; Podsakoff, G; Forman, S J; Chatterjee, S
Gene transfer vectors based on adeno-associated virus (AAV) appear promising because of their high transduction frequencies regardless of cell cycle status and ability to integrate into chromosomal DNA. We tested AAV-mediated gene transfer into a panel of human bone marrow or umbilical cord-derived CD34+ hematopoietic progenitor cells, using vectors encoding several transgenes under the control of viral and cellular promoters. Gene transfer was evaluated by (1) chromosomal integration of vector sequences and (2) analysis of transgene expression. Southern hybridization and fluorescence in situ hybridization analysis of transduced CD34 genomic DNA showed the presence of integrated vector sequences in chromosomal DNA in a portion of transduced cells and showed that integrated vector sequences were replicated along with cellular DNA during mitosis. Transgene expression in transduced CD34 cells in suspension cultures and in myeloid colonies differentiating in vitro from transduced CD34 cells approximated that predicted by the multiplicity of transduction. This was true in CD34 cells from different donors, regardless of the transgene or selective pressure. Comparisons of CD34 cell transduction either before or after cytokine stimulation showed similar gene transfer frequencies. Our findings suggest that AAV transduction of CD34+ hematopoietic progenitor cells is efficient, can lead to stable integration in a population of transduced cells, and may therefore provide the basis for safe and efficient ex vivo gene therapy of the hematopoietic system.
Virus-induced gene silencing (VIGS) is an efficient and rapid method to identify plant gene functions. One of the most widely used VIGS vectors is Tobacco rattle virus (TRV) which has been used successfully for RNA interference (RNAi) in N. benthamiana and tomato. We previously modified a TRV VIGS v...
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...
Wu, Zhijian; Wu, Xiaobing; Cao, Hui; Dong, Xiaoyan; Wang, Hong; Hou, Yunde
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.
Brown, Nolan; Song, Liujiang; Kollu, Nageswara R; Hirsch, Matthew L
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
Strains of Lentinula edodes suppress growth of phytopathogenic fungi and inhibit Alagoas serotype of vesicular stomatitis virus Linhagens de Lentinula edodes inibem fungos fitopatogênicos e o vírus da estomatite vesicular, sorotipo Alagoas
Selma H. Sasaki
Full Text Available Four Lentinula edodes strains (Le10, 46, K2, Assai were assessed for their antagonistic effect on four filamentous fungus species of agricultural importance (Helminthosporium euphorbiae, Helminthosporium sp, Fusarium solani and Phomopsis sojae and on Alagoas serotype of Vesicular Stomatitis Virus (VSA. The L. edodes strains studied had variable effects on the filamentous fungi and on VSA. The K2 and Le10 strains were antagonistic on the fungi assessed and the 46 and K2 strains were efficient on the Vesicular Stomatitis Virus. The results widened the list of beneficial effects of L. edodes on the control and prevention of animal pathogenic virus and filamentous fungi.Quatro linhagens de Lentinula edodes (Le10, 46, K2, ASSAI foram avaliadas quanto ao seu efeito inibitório sobre quatro espécies de fungos filamentosos de importância agrícola (Helminthosporium euphorbiae, Helminthosporium sp., Fusarium solani, Phomopsis sojae e sobre o sorotipo Alagoas vírus da estomatite vesicular (VSA. Foi observado que as linhagens de L. edodes estudadas apresentaram variabilidade quanto ao seu efeito, tanto sobre os fungos filamentosos quanto sobre o vírus VSA. As linhagens K2 e Le10 apresentaram-se antagônicas sobre os fungos e as linhagens 46 e K2 foram eficientes na inibição do vírus VSA. Os resultados obtidos permitem ampliar a lista de efeitos benéficos de algumas linhagens de L. edodes no controle e prevenção de vírus patogênicos animais e de fungos filamentosos.
Full Text Available Japanese encephalitis virus (JEV is among the major threats to public health in Asia. For disease control and prevention, the efficient production of safe and effective vaccines against JEV is in urgent need. In this study, we produced a plant-made JEV vaccine candidate using a chimeric virus particle (CVP strategy based on bamboo mosaic virus (BaMV for epitope presentation. The chimeric virus, designated BJ2A, was constructed by fusing JEV envelope protein domain III (EDIII at the N-terminus of BaMV coat protein, with an insertion of the foot-and-mouth disease virus 2A peptide to facilitate the production of both unfused and epitope-presenting for efficient assembly of the CVP vaccine candidate. The strategy allowed stable maintenance of the fusion construct over long-term serial passages in plants. Immuno-electron microscopy examination and immunization assays revealed that BJ2A is able to present the EDIII epitope on the surface of the CVPs, which stimulated effective neutralizing antibodies against JEV infection in mice. This study demonstrates the efficient production of an effective CVP vaccine candidate against JEV in plants by the BaMV-based epitope presentation system.
Mélissanne de Wispelaere
Full Text Available Japanese encephalitis virus (JEV is the causative agent of Japanese encephalitis, the leading cause of viral encephalitis in Asia. JEV transmission cycle involves mosquitoes and vertebrate hosts. The detection of JEV RNA in a pool of Culex pipiens caught in 2010 in Italy raised the concern of a putative emergence of the virus in Europe. We aimed to study the vector competence of European mosquito populations, such as Cx. pipiens and Aedes albopictus for JEV genotypes 3 and 5.After oral feeding on an infectious blood meal, mosquitoes were dissected at various times post-virus exposure. We found that the peak for JEV infection and transmission was between 11 and 13 days post-virus exposure. We observed a faster dissemination of both JEV genotypes in Ae. albopictus mosquitoes, when compared with Cx. pipiens mosquitoes. We also dissected salivary glands and collected saliva from infected mosquitoes and showed that Ae. albopictus mosquitoes transmitted JEV earlier than Cx. pipiens. The virus collected from Ae. albopictus and Cx. pipiens saliva was competent at causing pathogenesis in a mouse model for JEV infection. Using this model, we found that mosquito saliva or salivary glands did not enhance the severity of the disease.In this study, we demonstrated that European populations of Ae. albopictus and Cx. pipiens were efficient vectors for JEV transmission. Susceptible vertebrate species that develop high viremia are an obligatory part of the JEV transmission cycle. This study highlights the need to investigate the susceptibility of potential JEV reservoir hosts in Europe, notably amongst swine populations and local water birds.
Maria R. Mendoza
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.
Mosquitoes (Diptera: Culicidae) represent a key threat for millions of humans and ani-mals 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 hemi-sphere, 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 nano-particles, 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.
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.
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.
Xiao, Ping-Jie; Mitchell, Angela M.; Huang, Lu; Li, Chengwen; Samulski, R. Jude
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
Oliveira, Ana R S; Cohnstaedt, Lee W; Strathe, Erin; Hernández, Luciana Etcheverry; McVey, D Scott; Piaggio, José; Cernicchiaro, Natalia
Japanese encephalitis (JE) is a zoonosis in Southeast Asia vectored by mosquitoes infected with the Japanese encephalitis virus (JEV). Japanese encephalitis is considered an emerging exotic infectious disease with potential for introduction in currently JEV-free countries. Pigs and ardeid birds are reservoir hosts and play a major role on the transmission dynamics of the disease. The objective of the study was to quantitatively summarize the proportion of JEV infection in vectors and vertebrate hosts from data pertaining to observational studies obtained in a systematic review of the literature on vector and host competence for JEV, using meta-analyses. Data gathered in this study pertained to three outcomes: proportion of JEV infection in vectors, proportion of JEV infection in vertebrate hosts, and minimum infection rate (MIR) in vectors. Random-effects subgroup meta-analysis models were fitted by species (mosquito or vertebrate host species) to estimate pooled summary measures, as well as to compute the variance between studies. Meta-regression models were fitted to assess the association between different predictors and the outcomes of interest and to identify sources of heterogeneity among studies. Predictors included in all models were mosquito/vertebrate host species, diagnostic methods, mosquito capture methods, season, country/region, age category, and number of mosquitos per pool. Mosquito species, diagnostic method, country, and capture method represented important sources of heterogeneity associated with the proportion of JEV infection; host species and region were considered sources of heterogeneity associated with the proportion of JEV infection in hosts; and diagnostic and mosquito capture methods were deemed important contributors of heterogeneity for the MIR outcome. Our findings provide reference pooled summary estimates of vector competence for JEV for some mosquito species, as well as of sources of variability for these outcomes. Moreover, this
Duan, Zhiqiang; Xu, Houqiang; Ji, Xinqin; Zhao, Jiafu
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.
Vite, Charles H; Passini, Marco A; Haskins, Mark E; Wolfe, John H
Adeno-associated virus (AAV) vectors are capable of delivering a therapeutic gene to the mouse brain that can result in long-term and widespread protein production. However, the human infant brain is more than 1000 times larger than the mouse brain, which will make the treatment of global neurometabolic disorders in children more difficult. In this study, we evaluated the ability of three AAV serotypes (1,2, and 5) to transduce cells in the cat brain as a model of a large mammalian brain. The human lysosomal enzyme beta-glucuronidase (GUSB) was used as a reporter gene, because it can be distinguished from feline GUSB by heat stability. The vectors were injected into the cerebral cortex, caudate nucleus, thalamus, corona radiata, internal capsule, and centrum semiovale of 8-week-old cats. The brains were evaluated for gene expression using in situ hybridization and enzyme histochemistry 10 weeks after surgery. The AAV2 vector was capable of transducing cells in the gray matter, while the AAV1 vector resulted in greater transduction of the gray matter than AAV2 as well as transduction of the white matter. AAV5 did not result in detectable transduction in the cat brain.
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.
Kenney, Joan L; Anishchenko, Michael; Hermance, Meghan; Romo, Hannah; Chen, Ching-I; Thangamani, Saravanan; Brault, Aaron C
The Flavivirus genus comprises a diverse group of viruses that utilize a wide range of vertebrate hosts and arthropod vectors. The genus includes viruses that are transmitted solely by mosquitoes or vertebrate hosts as well as viruses that alternate transmission between mosquitoes or ticks and vertebrates. Nevertheless, the viral genetic determinants that dictate these unique flaviviral host and vector specificities have been poorly characterized. In this report, a cDNA clone of a flavivirus that is transmitted between ticks and vertebrates (Powassan lineage II, deer tick virus [DTV]) was generated and chimeric viruses between the mosquito/vertebrate flavivirus, West Nile virus (WNV), were constructed. These chimeric viruses expressed the prM and E genes of either WNV or DTV in the heterologous nonstructural (NS) backbone. Recombinant chimeric viruses rescued from cDNAs were characterized for their capacity to grow in vertebrate and arthropod (mosquito and tick) cells as well as for in vivo vector competence in mosquitoes and ticks. Results demonstrated that the NS elements were insufficient to impart the complete mosquito or tick growth phenotypes of parental viruses; however, these NS genetic elements did contribute to a 100- and 100,000-fold increase in viral growth in vitro in tick and mosquito cells, respectively. Mosquito competence was observed only with parental WNV, while infection and transmission potential by ticks were observed with both DTV and WNV-prME/DTV chimeric viruses. These data indicate that NS genetic elements play a significant, but not exclusive, role for vector usage of mosquito- and tick-borne flaviviruses.
Michael R. Strand
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.
Deardorff, Eleanor R.; Estrada-Franco, Jose G.; Freier, Jerome E.; Navarro-Lopez, Roberto; Da Rosa, Amelia Travassos; Tesh, Robert B.; Weaver, Scott C.
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
Kim, Shin-Hee; Samal, Siba K
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.
Rodrigo Villar, Gema
L' ENGROGUIMENT DE LES CUCURBITÀCIES: DIAGNÓSTIC I MICROSCOPIA DE LES RELACIONS VIRUS-PLANTA I VIRUS-VECTOR La malaltia de l'engroguiment de les cucurbitàcies, de gran importància, en els cultius del sud-est espanyol, està produïda per dos Closteroviridae transmesos per mosques blanques, el virus del fals engroguiment de la remolatxa (BPYV) i el virus de l'engroguiment enanitzant de les cucurbitàcies (CYSDV). BPYV és transmès específicament i de forma semipersistent per Trialeurodes vaporario...
Eliasson, Dubravka Grdic; Helgeby, Anja; Schön, Karin; Nygren, Caroline; El-Bakkouri, Karim; Fiers, Walter; Saelens, Xavier; Lövgren, Karin Bengtsson; Nyström, Ida; Lycke, Nils Y
Here we demonstrate that by using non-toxic fractions of saponin combined with CTA1-DD we can achieve a safe and above all highly efficacious mucosal adjuvant vector. We optimized the construction, tested the requirements for function and evaluated proof-of-concept in an influenza A virus challenge model. We demonstrated that the CTA1-3M2e-DD/ISCOMS vector provided 100% protection against mortality and greatly reduced morbidity in the mouse model. The immunogenicity of the vector was superior to other vaccine formulations using the ISCOM or CTA1-DD adjuvants alone. The versatility of the vector was best exemplified by the many options to insert, incorporate or admix vaccine antigens with the vector. Furthermore, the CTA1-3M2e-DD/ISCOMS could be kept 1 year at 4°C or as a freeze-dried powder without affecting immunogenicity or adjuvanticity of the vector. Strong serum IgG and mucosal IgA responses were elicited and CD4 T cell responses were greatly enhanced after intranasal administration of the combined vector. Together these findings hold promise for the combined vector as a mucosal vaccine against influenza virus infections including pandemic influenza. The CTA1-DD/ISCOMS technology represents a breakthrough in mucosal vaccine vector design which successfully combines immunomodulation and targeting in a safe and stable particulate formation. Copyright © 2011 Elsevier Ltd. All rights reserved.
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
Conway, J E; Rhys, C M; Zolotukhin, I; Zolotukhin, S; Muzyczka, N; Hayward, G S; Byrne, B J
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.
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.
Xuan, X; Tuchiya, K; Sato, I; Nishikawa, Y; Onoderaz, Y; Takashima, Y; Yamamoto, A; Katsumata, A; Iwata, A; Ueda, S; Mikami, T; Otsuka, H
In order to evaluate whether canine herpesvirus (CHV) could be used as a live vector for the expression of heterologous immunogenes, we constructed a recombinant canine herpesvirus (CHV) expressing glycoprotein (G protein) of rabies virus (RV). The gene of G protein was inserted within the thymidine kinase gene of CHV YP11mu strain under the control of the human cytomegalovirus immediate early promoter. The G protein expressed by the recombinant CHV was processed and transported to the cell surface as in RV infected cells, and showed the same biological activities such as low pH dependent cell fusion and hemadsorption. The antigenic authenticity of the recombinant G protein was confirmed by a panel of monoclonal antibodies specific for G protein. Dogs inoculated intransally with the recombinant CHV produced higher titres of virus neutralizing antibodies against RV than those inoculated with a commercial, inactivated rabies vaccine. These results suggest that the CHV recombinant expressing G protein can be used as a vaccine to control canine rabies and that CHV may be useful as a vector to develop live recombinant against other infectious diseases in dogs.
Gale, P; Brouwer, A; Ramnial, V; Kelly, L; Kosmider, R; Fooks, A R; Snary, E L
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.
Igarashi, Aki; Yamagata, Kousuke; Sugai, Tomokazu; Takahashi, Yukari; Sugawara, Emiko; Tamura, Akihiro; Yaegashi, Hajime; Yamagishi, Noriko; Takahashi, Tsubasa; Isogai, Masamichi; Takahashi, Hideki; Yoshikawa, Nobuyuki
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.
Lei, Wenbin; Liu, Danfeng; Li, Pei; Hou, Maolin
Performance of insect vectors can be influenced by the viruses they transmit, either directly by infection of the vectors or indirectly via infection of the host plants. Southern rice black-streaked dwarf virus (SRBSDV) is a propagative virus transmitted by the white-backed planthopper, Sogatella furcifera (Hovath). To elucidate the influence of SRBSDV on the performance of white-backed planthopper, life parameters of viruliferous and nonviruliferous white-backed planthopper fed rice seedlings infected or noninfected with SRBSDV were measured using a factorial design. Regardless of the infection status of the rice plant host, viruliferous white-backed planthopper nymphs took longer to develop from nymph to adult than did nonviruliferous nymphs. Viruliferous white-backed planthopper females deposited fewer eggs than nonviruliferous females and both viruliferous and nonviruliferous white-backed planthopper females laid fewer eggs on infected than on noninfected plants. Longevity of white-backed planthopper females was also affected by the infection status of the rice plant and white-backed planthopper. Nonviruliferous white-backed planthopper females that fed on infected rice plants lived longer than the other three treatment groups. These results indicate that the performance of white-backed planthopper is affected by SRBSDV either directly (by infection of white-backed planthopper) or indirectly (by infection of rice plant). The extended development of viruliferous nymphs and the prolonged life span of nonviruliferous adults on infected plants may increase their likelihood of transmitting virus, which would increase virus spread. © 2014 Entomological Society of America.
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
Full Text Available Viral vectors have been available in various fields such as medical and biological research or gene therapy applications. Targeting vectors pseudotyped with distinct viral envelope proteins that influence cell tropism and transfection efficiency is a useful tool not only for examining entry mechanisms or cell tropisms but also for vaccine vector development. Vesicular stomatitis virus (VSV is an excellent candidate for development as a pseudotype vector. A recombinant VSV lacking its own envelope (G gene has been used to produce a pseudotype or recombinant VSV possessing the envelope proteins of heterologous viruses. These viruses possess a reporter gene instead of a VSV G gene in their genome, and therefore it is easy to evaluate their infectivity in the study of viral entry, including identification of viral receptors. Furthermore, advantage can be taken of a property of the pseudotype VSV, which is competence for single-round infection, in handling many different viruses that are either difficult to amplify in cultured cells or animals or that require specialized containment facilities. Here we describe procedures for producing pseudotype or recombinant VSVs and a few of the more prominent examples from among envelope viruses, such as hepatitis C virus, Japanese encephalitis virus, baculovirus, and hemorrhagic fever viruses.
Malachy I. Okeke
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.
Martin, Kathleen M; Barandoc-Alviar, Karen; Schneweis, Derek J; Stewart, Catherine L; Rotenberg, Dorith; Whitfield, Anna E
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.
Casimiro, Danilo R.; Chen, Ling; Fu, Tong-Ming; Evans, Robert K.; Caulfield, Michael J.; Davies, Mary-Ellen; Tang, Aimin; Chen, Minchun; Huang, Lingyi; Harris, Virginia; Freed, Daniel C.; Wilson, Keith A.; Dubey, Sheri; Zhu, De-Min; Nawrocki, Denise
Cellular immune responses, particularly those associated with CD3+ CD8+ cytotoxic T lymphocytes (CTL), play a primary role in controlling viral infection, including persistent infection with human immunodeficiency virus type 1 (HIV-1). Accordingly, recent HIV-1 vaccine research efforts have focused on establishing the optimal means of eliciting such antiviral CTL immune responses. We evaluated several DNA vaccine formulations, a modified vaccinia virus Ankara vector, and a replication-defecti...
Richards, Stephanie L; Lord, Cynthia C; Pesko, Kendra; Tabachnick, Walter J
Complex interactions between environmental and biological factors influence the susceptibility of Culex pipiens quinquefasciatus to St. Louis encephalitis virus and could affect the epidemiology of virus transmission. Similar interactions could have epidemiologic implications for other vector-virus systems. We conducted an experiment to examine four such factors in combination: mosquito age, extrinsic incubation temperature (EIT), virus dose, and colony. The proportion of mosquitoes with body infections or disseminated infections varied between colonies, and was dependant on age, EIT, and dose. We also show that the probability of a body or leg infection interacted in complex ways between colonies, ages, EITs, and doses. The complex interactive effects of environmental and biological factors must be taken into account for studies of vector competence and epidemiology, especially when laboratory studies are used to generalize to natural transmission dynamics where the extent of variation is largely unknown.
Liu, Danfeng; Li, Pei; Han, Yongqiang; Lei, Wenbin; Hou, Maolin
Southern rice black-streaked dwarf virus (SRBSDV) is a novel virus transmitted by white-backed planthopper Sogatella furcifera (Hováth) (Hemiptera: Delphacidae). Due to low virus transmission efficiency by the planthopper, researchers are frequently confronted with shortage of viruliferous vectors or infected rice plants, especially in winter and the following spring. To find new ways to maintain virus-infected materials, viral rice plants were stored at -80°C for 45 or 140 d and evaluated as virus sources in virus transmission by the vector. SRBSDV virions were not degraded during storage at -80°C as indicated by reverse transcription-polymerase chain reaction and reverse transcription real-time PCR detection. The planthopper nymphs fed on the infected thawed plants for 48 h survived at about 40% and showed positive detection of SRBSDV, but they lost the virus after feeding for another 20 d (the circulative transmission period) on noninfected plants. Transmission electron microscope images indicated broken capsid of virions in infected thawed leaves in contrast to integrity capsid of virions in infected fresh leaves. These results show that low temperature storage of SRBSDV-infected rice plants cannot sustain virus transmission by white-backed planthopper. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: firstname.lastname@example.org.
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
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.
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.
Chen, Yuting; Cassone, Bryan J.; Bai, Xiaodong; Redinbaugh, Margaret G.; Michel, Andrew P.
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. PMID:22808205
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.
Schirtzinger Erin E
Full Text Available Abstract Background Competitive displacement of a weakly virulent pathogen strain by a more virulent strain is one route to disease emergence. However the mechanisms by which pathogens compete for access to hosts are poorly understood. Among vector-borne pathogens, variation in the ability to infect vectors may effect displacement. The current study focused on competitive displacement in dengue virus serotype 3 (DENV3, a mosquito-borne pathogen of humans. In Sri Lanka in the 1980's, a native DENV3 strain associated with relatively mild dengue disease was displaced by an invasive DENV3 strain associated with the most severe disease manifestations, dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS, resulting in an outbreak of DHF/DSS. Here we tested the hypothesis that differences between the invasive and native strain in their infectivity for Aedes aegypti mosquitoes, the primary vector of DENV, contributed to the competitive success of the invasive strain Results To be transmitted by a mosquito, DENV must infect and replicate in the midgut, disseminate into the hemocoel, infect the salivary glands, and be released into the saliva. The ability of the native and invasive DENV3 strains to complete the first three steps of this process in Aedes aegypti mosquitoes was measured in vivo. The invasive strain infected a similar proportion of mosquitoes as the native strain but replicated to significantly higher titers in the midgut and disseminated with significantly greater efficiency than the native strain. In contrast, the native and invasive strain showed no significant difference in replication in cultured mosquito, monkey or human cells. Conclusion The invasive DENV3 strain infects and disseminates in Ae. aegypti more efficiently than the displaced native DENV3 strain, suggesting that the invasive strain is transmitted more efficiently. Replication in cultured cells did not adequately characterize the known phenotypic differences between
Kevin A. Caillouët
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.
Ghanim, Murad; Brumin, Marina; Popovski, Smadar
A simple, rapid, inexpensive method for the localization of virus transcripts in plant and insect vector tissues is reported here. The method based on fluorescent in situ hybridization using short DNA oligonucleotides complementary to an RNA segment representing a virus transcript in the infected plant or insect vector. The DNA probe harbors a fluorescent molecule at its 5' or 3' ends. The protocol: simple fixation, hybridization, minimal washing and confocal microscopy, provides a highly specific signal. The reliability of the protocol was tested by localizing two phloem-limited plant virus transcripts in infected plants and insect tissues: Tomato yellow leaf curl virus (TYLCV) (Begomovirus: Geminiviridae), exclusively transmitted by the whitefly Bemisia tabaci (Gennadius) in a circulative non-propagative manner, and Potato leafroll virus (Polerovirus: Luteoviridae), similarly transmitted by the aphid Myzus persicae (Sulzer). Transcripts for both viruses were localized specifically to the phloem sieve elements of infected plants, while negative controls showed no signal. TYLCV transcripts were also localized to the digestive tract of B. tabaci, confirming TYLCV route of transmission. Compared to previous methods for localizing virus transcripts in plant and insect tissues that include complex steps for in-vitro probe preparation or antibody raising, tissue fixation, block preparation, sectioning and hybridization, the method described below provides very reliable, convincing, background-free results with much less time, effort and cost.
Meador, Lydia R. [Ira A. Fulton School of Engineering, Arizona State University, Tempe, AZ (United States); Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); Kessans, Sarah A. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Kilbourne, Jacquelyn; Kibler, Karen V. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); Pantaleo, Giuseppe [Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois, University of Lausanne, Lausanne (Switzerland); Swiss Vaccine Research Institute, Lausanne (Switzerland); Roderiguez, Mariano Esteban [Department of Molecular and Cellular Biology, Centro Nacional de Biotecnologia – CSIC, Madrid (Spain); Blattman, Joseph N. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Jacobs, Bertram L., E-mail: email@example.com [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Mor, Tsafrir S., E-mail: firstname.lastname@example.org [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States)
Showing modest efficacy, the RV144 HIV-1 vaccine clinical trial utilized a non-replicating canarypox viral vector and a soluble gp120 protein boost. Here we built upon the RV144 strategy by developing a novel combination of a replicating, but highly-attenuated Vaccinia virus vector, NYVAC-KC, and plant-produced HIV-1 virus-like particles (VLPs). Both components contained the full-length Gag and a membrane anchored truncated gp41 presenting the membrane proximal external region with its conserved broadly neutralizing epitopes in the pre-fusion conformation. We tested different prime/boost combinations of these components in mice and showed that the group primed with NYVAC-KC and boosted with both the viral vectors and plant-produced VLPs have the most robust Gag-specific CD8 T cell responses, at 12.7% of CD8 T cells expressing IFN-γ in response to stimulation with five Gag epitopes. The same immunization group elicited the best systemic and mucosal antibody responses to Gag and dgp41 with a bias towards IgG1. - Highlights: • We devised a prime/boost anti HIV-1 vaccination strategy modeled after RV144. • We used plant-derived virus-like particles (VLPs) consisting of Gag and dgp41. • We used attenuated, replicating vaccinia virus vectors expressing the same antigens. • The immunogens elicited strong cellular and humoral immune responses.
Tabachnick, W J
Seven Colorado populations of the bluetongue virus vector Culicoides varipennis (Coquillett) were analyzed for genetic variation at 19-21 isozyme loci. Permanent populations, which overwinter as larvae, showed little temporal genetic change at 19 loci. PGD and MDH showed seasonal changes in gene frequencies, attributable to selection at two permanent populations. Two temporary populations showed low heterozygosity compared with permanent populations. Independent estimates of gene flow, calculated using FST and the private allele method, were Nm* = 2.15 and 6.95, respectively. Colorado C. variipennis permanent populations showed high levels of gene flow which prevented significant genetic differentiation due to genetic drift. Temporary populations showed significant gene frequency differences from nearby permanent populations due to the "founder effect" associated with chance colonization.
Hartemink, N.; Purse, B.V.; Meiswinkel, R.; Brown, H.E.; Koeijer, de A.A.; Elbers, A.R.W.; Boender, G.J.; Rogers, D.J.; Heesterbeek, J.A.P.
Geographical maps indicating the value of the basic reproduction number, R0, can be used to identify areas of higher risk for an outbreak after an introduction. We develop a methodology to create R0 maps for vector-borne diseases, using bluetongue virus as a case study. This method provides a tool
Miesbach, Wolfgang; Meijer, Karina; Coppens, Michiel
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 expre...
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 ...
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
Lubicz, Jeanmarie Verchot; Rush, Charles M; Payton, Mark; Colberg, Terry
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.
Richards, Stephanie L; Anderson, Sheri L; Lord, Cynthia C; Tabachnick, Walter J
Culex nigripalpus Theobald is a primary vector of St. Louis encephalitis virus in the southeastern United States. Cx. nigripalpus females were fed blood containing a low (4.0 +/- 0.01 log10 plaque-forming unit equivalents (PFUeq) /ml) or high (4.7 +/- 0.1 log10 PFUeq/ml) St. Louis encephalitis virus dose and maintained at extrinsic incubation temperatures (EIT) of 25 or 28 degrees C for 12 d. Vector competence was measured via quantitative real-time reverse transcriptase polymerase chain reaction to estimate PFUeq using rates of infection, dissemination, and transmission. There were no differences in infection rates between the two EITs at either dose. The low dose had higher infection rates at both EITs. Dissemination rates were significantly higher at 28 degrees C compared with 25 degrees C at both doses. Virus transmission was observed (<7%) only at 28 degrees C for both doses. The virus titer in body tissues was greater at 28 degrees C compared with 25 degrees C at both doses. The difference between the EITs was greater at the low dose, resulting in a higher titer for the low dose than the high dose at 28 degrees C. Virus titers in leg tissues were greater in mosquitoes fed the high versus low dose, but were not influenced by EIT. Further investigations using a variety of environmental and biological factors would be useful in exploring the complexity of vector competence.
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
Huang, Shuohao; Kawabe, Yoshinori; Ito, Akira; Kamihira, Masamichi
Highlights: ► Adeno-associated virus (AAV) is capable of targeted integration in human cells. ► Integrase-defective retroviral vector (IDRV) enables a circular DNA delivery. ► A targeted integration system of IDRV DNA using the AAV integration mechanism. ► 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.
Ba, Y; Diallo, D; Dia, I; Diallo, M
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
Ding, Xin Shun; Schneider, William L; Chaluvadi, Srinivasa Rao; Mian, M A Rouf; Nelson, Richard S
Virus-induced gene silencing (VIGS) is used to analyze gene function in dicotyledonous plants but less so in monocotyledonous plants (particularly rice and corn), partially due to the limited number of virus expression vectors available. Here, we report the cloning and modification for VIGS of a virus from Festuca arundinacea Schreb. (tall fescue) that caused systemic mosaic symptoms on barley, rice, and a specific cultivar of maize (Va35) under greenhouse conditions. Through sequencing, the virus was determined to be a strain of Brome mosaic virus (BMV). The virus was named F-BMV (F for Festuca), and genetic determinants that controlled the systemic infection of rice were mapped to RNAs 1 and 2 of the tripartite genome. cDNA from RNA 3 of the Russian strain of BMV (R-BMV) was modified to accept inserts from foreign genes. Coinoculation of RNAs 1 and 2 from F-BMV and RNA 3 from R-BMV expressing a portion of a plant gene to leaves of barley, rice, and maize plants resulted in visual silencing-like phenotypes. The visual phenotypes were correlated with decreased target host transcript levels in the corresponding leaves. The VIGS visual phenotype varied from maintained during silencing of actin 1 transcript expression to transient with incomplete penetration through affected tissue during silencing of phytoene desaturase expression. F-BMV RNA 3 was modified to allow greater accumulation of virus while minimizing virus pathogenicity. The modified vector C-BMV(A/G) (C for chimeric) was shown to be useful for VIGS. These BMV vectors will be useful for analysis of gene function in rice and maize for which no VIGS system is reported.
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
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
Philip Samuel, P; Arunachalam, N; Hiriyan, J; Tyagi, B K
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.
Van doninck, J.; Peters, J.; De Baets, B.; Ducheyne, E.; Verhoest, N. E. C.
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.
Iatrou, K; Meidinger, R G
A pair of silkmoth chorion chromosomal genes, HcA.12-HcB.12, was inserted into a baculovirus transfer vector, pBmp2, derived from the nuclear polyhedrosis virus of Bombyx mori. This vector, which permits the insertion of foreign genetic material in the vicinity of a mutationally inactivated polyhedrin gene, was used to acquire the corresponding recombinant virus. Injection of mutant silkmoth pupae that lack all Hc chorion genes with the recombinant virus resulted in the infection of all internal organs including follicular tissue. Analysis of RNA from infected tissues has demonstrated that the two chorion genes present in the viral genome are correctly transcribed under the control of their own promoter in follicular cells, the tissue in which chorion genes are normally expressed. The chorion primary transcripts are also correctly processed in the infected follicular cells and yield mature mRNAs indistinguishable from authentic chorion mRNAs present in wild-type follicles. These results demonstrate that recombinant nuclear polyhedrosis viruses can be used as transducing vectors for introducing genetic material of host origin into the cells of the organism and that the transduced genes are transiently expressed in a tissue-specific manner under the control of their resident regulatory sequences. Thus we show the in vivo expression of cloned genes under cellular promoter control in an insect other than Drosophila melanogaster. The approach should be applicable to all insect systems that are subject to nuclear polyhedrosis virus infection. Images PMID:2187186
Drolet, Barbara S.; Stuart, Melissa A.; Derner, Justin D.
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
Calvez, Elodie; Guillaumot, Laurent; Girault, Dominique; Richard, Vaea; O'Connor, Olivia; Paoaafaite, Tuterarii; Teurlai, Magali; Pocquet, Nicolas; Cao-Lormeau, Van-Mai; Dupont-Rouzeyrol, Myrielle
Dengue virus (DENV) is the arbovirus with the highest incidence in New Caledonia and in the South Pacific region. In 2012-2014, a major DENV-1 outbreak occurred in New Caledonia. The only known vector of DENV in New Caledonia is Aedes aegypti but no study has yet evaluated the competence of New Caledonia Ae. aegypti populations to transmit DENV. This study compared the ability of field-collected Ae. aegypti from different locations in New Caledonia to transmit the DENV-1 responsible for the 2012-2014 outbreak. This study also aimed to compare the New Caledonia results with the vector competence of Ae. aegypti from French Polynesia as these two French countries have close links, including arbovirus circulation. Three wild Ae. aegypti populations were collected in New Caledonia and one in French Polynesia. Female mosquitoes were orally exposed to DENV-1 (10 6 FFU/ml). Mosquito bodies (thorax and abdomen), heads and saliva were analyzed to measure infection, dissemination, transmission rates and transmission efficiency, at 7, 14 and 21 days post-infection (dpi), respectively. DENV-1 infection rates were heterogeneous, but dissemination rates were high and homogenous among the three Ae. aegypti populations from New Caledonia. Despite this high DENV-1 dissemination rate, the transmission rate, and therefore the transmission efficiency, observed were low. Aedes aegypti population from New Caledonia was less susceptible to infection and had lower ability to transmit DENV-1 than Ae. aegypti populations from French Polynesia. This study suggests that even if susceptible to infection, the New Caledonian Ae. aegypti populations were moderately competent vectors for DENV-1 strain from the 2012-2014 outbreak. These results strongly suggest that other factors might have contributed to the spread of this DENV-1 strain in New Caledonia and in the Pacific region.
Volak, Adrienn; LeRoy, Stanley G; Natasan, Jeya Shree; Park, David J; Cheah, Pike See; Maus, Andreas; Fitzpatrick, Zachary; Hudry, Eloise; Pinkham, Kelsey; Gandhi, Sheetal; Hyman, Bradley T; Mu, Dakai; GuhaSarkar, Dwijit; Stemmer-Rachamimov, Anat O; Sena-Esteves, Miguel; Badr, Christian E; Maguire, Casey A
The malignant primary brain tumor, glioblastoma (GBM) is generally incurable. New approaches are desperately needed. Adeno-associated virus (AAV) vector-mediated delivery of anti-tumor transgenes is a promising strategy, however direct injection leads to focal transgene spread in tumor and rapid tumor division dilutes out the extra-chromosomal AAV genome, limiting duration of transgene expression. Intravenous (IV) injection gives widespread distribution of AAV in normal brain, however poor transgene expression in tumor, and high expression in non-target cells which may lead to ineffective therapy and high toxicity, respectively. Delivery of transgenes encoding secreted, anti-tumor proteins to tumor stromal cells may provide a more stable and localized reservoir of therapy as they are more differentiated than fast-dividing tumor cells. Reactive astrocytes and tumor-associated macrophage/microglia (TAMs) are stromal cells that comprise a large portion of the tumor mass and are associated with tumorigenesis. In mouse models of GBM, we used IV delivery of exosome-associated AAV vectors driving green fluorescent protein expression by specific promoters (NF-κB-responsive promoter and a truncated glial fibrillary acidic protein promoter), to obtain targeted transduction of TAMs and reactive astrocytes, respectively, while avoiding transgene expression in the periphery. We used our approach to express the potent, yet toxic anti-tumor cytokine, interferon beta, in tumor stroma of a mouse model of GBM, and achieved a modest, yet significant enhancement in survival compared to controls. Noninvasive genetic modification of tumor microenvironment represents a promising approach for therapy against cancers. Additionally, the vectors described here may facilitate basic research in the study of tumor stromal cells in situ.
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.
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
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
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.
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
Sheila B Agha
Full Text Available In April, 2004, chikungunya virus (CHIKV re-emerged in Kenya and eventually spread to the islands in the Indian Ocean basin, South-East Asia, and the Americas. The virus, which is often associated with high levels of viremia in humans, is mostly transmitted by the urban vector, Aedes aegypti. The expansion of CHIKV presents a public health challenge both locally and internationally. In this study, we investigated the ability of Ae. aegypti mosquitoes from three distinct cities in Kenya; Mombasa (outbreak prone, Kisumu, and Nairobi (no documented outbreak to transmit CHIKV.Aedes aegypti mosquito populations were exposed to different doses of CHIKV (105.6-7.5 plaque-forming units[PFU]/ml in an infectious blood meal. Transmission was ascertained by collecting and testing saliva samples from individual mosquitoes at 5, 7, 9, and 14 days post exposure. Infection and dissemination were estimated by testing body and legs, respectively, for individual mosquitoes at selected days post exposure. Tissue culture assays were used to determine the presence of infectious viral particles in the body, leg, and saliva samples. The number of days post exposure had no effect on infection, dissemination, or transmission rates, but these rates increased with an increase in exposure dose in all three populations. Although the rates were highest in Ae. aegypti from Mombasa at titers ≥106.9 PFU/ml, the differences observed were not statistically significant (χ2 ≤ 1.04, DF = 1, P ≥ 0.31. Overall, about 71% of the infected mosquitoes developed a disseminated infection, of which 21% successfully transmitted the virus into a capillary tube, giving an estimated transmission rate of about 10% for mosquitoes that ingested ≥106.9 PFU/ml of CHIKV. All three populations of Ae. aegypti were infectious as early as 5-7 days post exposure. On average, viral dissemination only occurred when body titers were ≥104 PFU/ml in all populations.Populations of Ae. aegypti from
Agha, Sheila B; Chepkorir, Edith; Mulwa, Francis; Tigoi, Caroline; Arum, Samwel; Guarido, Milehna M; Ambala, Peris; Chelangat, Betty; Lutomiah, Joel; Tchouassi, David P; Turell, Michael J; Sang, Rosemary
In April, 2004, chikungunya virus (CHIKV) re-emerged in Kenya and eventually spread to the islands in the Indian Ocean basin, South-East Asia, and the Americas. The virus, which is often associated with high levels of viremia in humans, is mostly transmitted by the urban vector, Aedes aegypti. The expansion of CHIKV presents a public health challenge both locally and internationally. In this study, we investigated the ability of Ae. aegypti mosquitoes from three distinct cities in Kenya; Mombasa (outbreak prone), Kisumu, and Nairobi (no documented outbreak) to transmit CHIKV. Aedes aegypti mosquito populations were exposed to different doses of CHIKV (105.6-7.5 plaque-forming units[PFU]/ml) in an infectious blood meal. Transmission was ascertained by collecting and testing saliva samples from individual mosquitoes at 5, 7, 9, and 14 days post exposure. Infection and dissemination were estimated by testing body and legs, respectively, for individual mosquitoes at selected days post exposure. Tissue culture assays were used to determine the presence of infectious viral particles in the body, leg, and saliva samples. The number of days post exposure had no effect on infection, dissemination, or transmission rates, but these rates increased with an increase in exposure dose in all three populations. Although the rates were highest in Ae. aegypti from Mombasa at titers ≥106.9 PFU/ml, the differences observed were not statistically significant (χ2 ≤ 1.04, DF = 1, P ≥ 0.31). Overall, about 71% of the infected mosquitoes developed a disseminated infection, of which 21% successfully transmitted the virus into a capillary tube, giving an estimated transmission rate of about 10% for mosquitoes that ingested ≥106.9 PFU/ml of CHIKV. All three populations of Ae. aegypti were infectious as early as 5-7 days post exposure. On average, viral dissemination only occurred when body titers were ≥104 PFU/ml in all populations. Populations of Ae. aegypti from Mombasa, Nairobi
Zhong, Li; Li, Baozheng; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Cooper, Mario; Herzog, Roland W.; Zolotukhin, Irene; Warrington, Kenneth H.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun
Recombinant adeno-associated virus 2 (AAV2) vectors are in use in several Phase I/II clinical trials, but relatively large vector doses are needed to achieve therapeutic benefits. Large vector doses also trigger an immune response as a significant fraction of the vectors fails to traffic efficiently to the nucleus and is targeted for degradation by the host cell proteasome machinery. We have reported that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively...
Crowder, David W; Dykstra, Elizabeth A; Brauner, Jo Marie; Duffy, Anne; Reed, Caitlin; Martin, Emily; Peterson, Wade; Carrière, Yves; Dutilleul, Pierre; Owen, Jeb P
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.
Tabachnick, W J
Vesicular stomatitis (VS) epizootics in the Western United States have caused substantial economic losses to U.S. livestock industries in 1995, 1997, and 1998. The role of arthropods in transmitting VS to U.S. livestock is unclear. In particular, the impact of arthropod salivary gland factors in VS infections in livestock needs study. Pharmacological effects of arthropod salivary gland factors on animals are reviewed. The potential effects of arthropod saliva on the transmission and spread of VS virus to livestock in the Western U.S. is presented with emphasis on the biting midge, Culicoides sonorensis. Information is discussed with attention to vector potential of C. sonorensis, and its use as a model for evaluating insect salivary gland pharmacology on livestock response to VS.
Perez de Leon, A A; Ribeiro, J M; Tabachnick, W J; Valenzuela, J G
Several species of Culicoides biting midges are important pests and vectors of pathogens affecting humans and other animals. Bluetongue is the most economically important arthropod-borne animal disease in the United States. Culicoides variipennis is the primary North American vector of the bluetongue viruses. A reddish halo surrounding a petechial hemorrhage was noticed at the site of C. variipennis blood feeding in previously unexposed sheep and rabbits. Salivary gland extracts of nonblood-fed C. variipennis injected intradermally into sheep and rabbits induced cutaneous vasodilation in the form of erythema. A local, dose-dependent erythema, without edema or pruritus, was noted 30 min after injection. Erythema was inapparent with salivary gland extracts obtained after blood feeding. This observation suggested that the vasodilatory activity was inoculated into the host skin at the feeding site. The vasodilatory activity was insoluble in ethanol and destroyed by trypsin or chymotrypsin, which indicated that vasodilation was due to a protein. The association of cutaneous vasodilation with a salivary protein was corroborated by reversed-phase, high-performance liquid chromatography (HPLC). Fractionation of salivary gland extracts by molecular sieving HPLC resulted in maximal vasodilatory activity that coeluted with a protein having a relative molecular weight (MWr) of 22.45 kD. The C. variipennis vasodilator appears to be biologically active at the nanogram level. This vasodilator likely assists C. variipennis during feeding by increasing blood flow from host superficial blood vessels surrounding the bite site. The identification of a salivary vasodilator in C. variipennis may have implications for the transmission of Culicoides-borne pathogens and in the development of dermatitis resulting from the sensitization of humans and animals to Culicoides salivary antigens.
Roilides, E.; Munson, P.J.; Levine, A.S.; Dixon, K.
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
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.
Patel, M; Carritt, K; Lane, J; Jayappa, H; Stahl, M; Bourgeois, M
Four vaccines for feline leukemia virus (FeLV) are available in the United States. This study's purpose was to compare the efficacy of Nobivac feline 2-FeLV (an inactivated, adjuvanted whole-virus vaccine) and PureVax recombinant FeLV (a live, canarypox virus-vectored vaccine) following FeLV challenge. Cats were vaccinated at 9 and 12 weeks with Nobivac feline 2-FeLV (group A, n = 11) or PureVax recombinant FeLV (group B, n = 10). Group C (n = 11) comprised unvaccinated controls. At 3 months postvaccination, cats were immunosuppressed and challenged with FeLV-A/61E. The outcomes measured were persistent antigenemia at 12 weeks postchallenge (PC) and proviral DNA and viral RNA at 3 to 9 weeks PC. Persistent antigenemia was observed in 0 of 11 cats in group A, 5 of 10 cats in group B, and 10 of 11 cats in group C. Group A was significantly protected compared to those in groups B (P 0.063). The preventable fraction was 100% for group A and 45% for group B. At 9 weeks PC, proviral DNA and viral RNA were detected 1 of 11 cats in group A, 6 of 10 cats in group B, and 9 of 11 cats in group C. Nucleic acid loads were significantly lower in group A than in group C (P feline 2-FeLV-vaccinated cats were fully protected against persistent antigenemia and had significantly smaller amounts of proviral DNA and plasma viral RNA loads than PureVax recombinant FeLV-vaccinated cats and unvaccinated controls. Copyright © 2015, Patel et al.
Kali D Saxton-Shaw
Full Text Available O'nyong nyong virus (ONNV and Chikungunya virus (CHIKV are two closely related alphaviruses with very different infection patterns in the mosquito, Anopheles gambiae. ONNV is the only alphavirus transmitted by anopheline mosquitoes, but specific molecular determinants of infection of this unique vector specificity remain unidentified. Fifteen distinct chimeric viruses were constructed to evaluate both structural and non-structural regions of the genome and infection patterns were determined through artificial infectious feeds in An. gambiae with each of these chimeras. Only one region, non-structural protein 3 (nsP3, was sufficient to up-regulate infection to rates similar to those seen with parental ONNV. When ONNV non-structural protein 3 (nsP3 replaced nsP3 from CHIKV virus in one of the chimeric viruses, infection rates in An. gambiae went from 0% to 63.5%. No other single gene or viral region addition was able to restore infection rates. Thus, we have shown that a non-structural genome element involved in viral replication is a major element involved in ONNV's unique vector specificity.
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.
Geller, A.I.; Keyomarsi, K.; Bryan, J.; Pardee, A.B.
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
Benelli, Giovanni; Mehlhorn, Heinz
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
Wiegand, Marian Alexander; Gori-Savellini, Gianni; Gandolfo, Claudia; Papa, Guido; Kaufmann, Christine; Felder, Eva; Ginori, Alessandro; Disanto, Maria Giulia; Spina, Donatella; Cusi, Maria Grazia
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
Rey-Jurado, Emma; Soto, Jorge; Gálvez, Nicolás; Kalergis, Alexis M
The human Respiratory Syncytial Virus (hRSV) causes lower respiratory tract infections including pneumonia and bronchiolitis. Such infections also cause a large number of hospitalizations and affects mainly newborns, young children and the elderly worldwide. Symptoms associated with hRSV infection are due to an exacerbated immune response characterized by low levels of IFN-γ, recruitment of neutrophils and eosinophils to the site of infection and lung damage. Although hRSV is a major health problem, no vaccines are currently available. Different immunization approaches have been developed to achieve a vaccine that activates the immune system, without triggering an unbalanced inflammation. These approaches include live attenuated vaccine, DNA or proteins technologies, and the use of vectors to express proteins of the virus. In this review, we discuss the host immune response to hRSV and the immunological mechanisms underlying an effective and safe BCG vectored vaccine against hRSV.
This report describes the procedures used to clone a 673 base pair gene fragment of the major nucleocapsid protein gene of Newcastle disease virus into a viral vector molecule for the purpose of maintaining a stable, long-term, renewable source of this target sequence for gene probe studies. The gene fragment was prepared by reverse-transcription polymerase chain reaction of Newcastle disease virus RNA and was cloned into the viral DNA vector Ml3mp18 RF to produce a recombinant DNA molecule. The cloned fragment was shown to be present in the recombinant clones based on (i) clonal selection on indicator plates; (ii) restriction enzyme analysis; (iii) gene probe analysis and (iv) nested PCR amplification.
Schmidtmann, E T; Tabachnick, W J; Hunt, G J; Thompson, L H; Hurd, H S
Entomologic and epizootic data are reviewed concerning the potential for transmission of vesicular stomatitis (VS) virus by insects, including field data from case-positive premises in New Mexico and Colorado during the 1995 outbreak of the New Jersey serotype (VSNJ). As with previous outbreaks of VSNJ in the western United States, the 1995 epizootic illustrated that risk of exposure is seasonal, increasing during warm weather and decreasing with onset of cool weather; virus activity spread from south to north along river valleys of the southwestern and Rocky Mountain states; clinical disease was detected most commonly in horses, but also occurred in cattle and 1 llama; and most infections were subclinical. Overall, 367 case-positive premises were identified during the 1995 outbreak, with foci of virus activity along the Rio Grande River south of Albuquerque, NM, in southwestern Colorado, and along the Colorado River near Grand Junction, CO. The establishment of a 16-km (10-mile) radius zone of restricted animal movement around confirmed positive premises, along with imposition of state and international embargoes, created economic hardship for livestock owners and producers. The importance of defining the role of blood-feeding insects as biological vectors of VSNJ virus relative to risk factors that promote high levels of insect transmission, such as the presence of livestock along western river valleys, blood feeding activity, and frequent transport of animals for recreational purposes, is emphasized as a basis for developing effective disease management.
Ammar, El-Desouky; Hogenhout, Saskia A
To investigate the dissemination route of Maize mosaic virus (MMV, Rhabdoviridae) in its planthopper vector Peregrinus maidis (Delphacidae, Hemiptera), temporal and spatial distribution of MMV was studied by immunofluorescence confocal laser scanning microscopy following 1-week acquisition feeding of planthoppers on infected plants. MMV was detected 1-week post first access to diseased plants (padp) in the midgut and anterior diverticulum, 2-week padp in the esophagus, nerves, nerve ganglia and visceral muscles, and 3-week padp in hemocytes, tracheae, salivary glands and other tissues. MMV is neurotropic in P. maidis; infection was more extensive in the nervous system compared to other tissues. A significantly higher proportion of planthoppers had infected midguts (28.1%) compared to those with infected salivary glands (20.4%) or to those that transmitted MMV (15.7%), suggesting the occurrence of midgut and salivary gland barriers to MMV transmission in P. maidis. In this planthopper, the esophagus and anterior diverticulum are located between the compound ganglionic mass and the salivary glands. We postulate that MMV may overcome transmission barriers in P. maidis by proceeding from the midgut to the anterior diverticulum and esophagus, and from these to the salivary glands via the nervous system: a neurotropic route similar to that of some vertebrate-infecting rhabdoviruses.
See, Raymond H; Petric, Martin; Lawrence, David J; Mok, Catherine P Y; Rowe, Thomas; Zitzow, Lois A; Karunakaran, Karuna P; Voss, Thomas G; Brunham, Robert C; Gauldie, Jack; Finlay, B Brett; Roper, Rachel L
Although the 2003 severe acute respiratory syndrome (SARS) outbreak was controlled, repeated transmission of SARS coronavirus (CoV) over several years makes the development of a SARS vaccine desirable. We performed a comparative evaluation of two SARS vaccines for their ability to protect against live SARS-CoV intranasal challenge in ferrets. Both the whole killed SARS-CoV vaccine (with and without alum) and adenovirus-based vectors encoding the nucleocapsid (N) and spike (S) protein induced neutralizing antibody responses and reduced viral replication and shedding in the upper respiratory tract and progression of virus to the lower respiratory tract. The vaccines also diminished haemorrhage in the thymus and reduced the severity and extent of pneumonia and damage to lung epithelium. However, despite high neutralizing antibody titres, protection was incomplete for all vaccine preparations and administration routes. Our data suggest that a combination of vaccine strategies may be required for effective protection from this pathogen. The ferret may be a good model for SARS-CoV infection because it is the only model that replicates the fever seen in human patients, as well as replicating other SARS disease features including infection by the respiratory route, clinical signs, viral replication in upper and lower respiratory tract and lung damage.
White, JD; Thesier, DM; Swain, JBD; Katz, MG; Tomasulo, C; Henderson, A; Wang, L; Yarnall, C; Fargnoli, A; Sumaroka, M; Isidro, A; Petrov, M; Holt, D; Nolen-Walston, R; Koch, WJ; Stedman, HH; Rabinowitz, J; Bridges, CR
We use a novel technique that allows for closed recirculation of vector genomes in the cardiac circulation using cardiopulmonary bypass, referred to here as molecular cardiac surgery with recirculating delivery (MCARD). We demonstrate that this platform technology is highly efficient in isolating the heart from the systemic circulation in vivo. Using MCARD, we compare the relative efficacy of single-stranded (ss) adeno-associated virus (AAV)6, ssAAV9 and self-complimentary (sc)AAV6-encoding enhanced green fluorescent protein, driven by the constitutive cytomegalovirus promoter to transduce the ovine myocardium in situ. MCARD allows for the unprecedented delivery of up to 48 green fluorescent protein genome copies per cell globally in the sheep left ventricular (LV) myocardium. We demonstrate that scAAV6-mediated MCARD delivery results in global, cardiac-specific LV gene expression in the ovine heart and provides for considerably more robust and cardiac-specific gene delivery than other available delivery techniques such as intramuscular injection or intracoronary injection; thus, representing a potential, clinically translatable platform for heart failure gene therapy. PMID:21228882
Full Text Available Abstract Background Ebola virus (EBOV is a Category A pathogen that is a member of Filoviridae family that causes hemorrhagic fever in humans and non-human primates. Unpredictable and devastating outbreaks of disease have recently occurred in Africa and current immunoprophylaxis and therapies are limited. The main limitation of working with pathogens like EBOV is the need for costly containment. To potentiate further and wider opportunity for EBOV prophylactics and therapies development, innovative approaches are necessary. Methods In the present study, an antigen delivery platform based on a recombinant bovine herpesvirus 4 (BoHV-4, delivering a synthetic EBOV glycoprotein (GP gene sequence, BoHV-4-syEBOVgD106ΔTK, was generated. Results EBOV GP was abundantly expressed by BoHV-4-syEBOVgD106ΔTK transduced cells without decreasing viral replication. BoHV-4-syEBOVgD106ΔTK immunized goats produced high titers of anti-EBOV GP antibodies and conferred a long lasting (up to 6 months, detectable antibody response. Furthermore, no evidence of BoHV-4-syEBOVgD106ΔTK viremia and secondary localization was detected in any of the immunized animals. Conclusions The BoHV-4-based vector approach described here, represents: an alternative antigen delivery system for vaccination and a proof of principle study for anti-EBOV antibodies generation in goats for potential immunotherapy applications.
Transcriptomic profiling of diverse Aedes aegypti strains reveals increased basal-level immune activation in dengue virus-refractory populations and identifies novel virus-vector molecular interactions.
Full Text Available Genetic variation among Aedes aegypti populations can greatly influence their vector competence for human pathogens such as the dengue virus (DENV. While intra-species transcriptome differences remain relatively unstudied when compared to coding sequence polymorphisms, they also affect numerous aspects of mosquito biology. Comparative molecular profiling of mosquito strain transcriptomes can therefore provide valuable insight into the regulation of vector competence. We established a panel of A. aegypti strains with varying levels of susceptibility to DENV, comprising both laboratory-maintained strains and field-derived colonies collected from geographically distinct dengue-endemic regions spanning South America, the Caribbean, and Southeast Asia. A comparative genome-wide gene expression microarray-based analysis revealed higher basal levels of numerous immunity-related gene transcripts in DENV-refractory mosquito strains than in susceptible strains, and RNA interference assays further showed different degrees of immune pathway contribution to refractoriness in different strains. By correlating transcript abundance patterns with DENV susceptibility across our panel, we also identified new candidate modulators of DENV infection in the mosquito, and we provide functional evidence for two potential DENV host factors and one potential restriction factor. Our comparative transcriptome dataset thus not only provides valuable information about immune gene regulation and usage in natural refractoriness of mosquito populations to dengue virus but also allows us to identify new molecular interactions between the virus and its mosquito vector.
Han, Zongchao; Zhong, Li; Maina, Njeri; Hu, Zhongbo; Li, Xiaomiao; Chouthai, Nitin S; Bischof, Daniela; Weigel-Van Aken, Kirsten A; Slayton, William B; Yoder, Mervin C; Srivastava, Arun
We previously reported that among single-stranded adeno-associated virus (ssAAV) vectors, serotypes 1 through 5, ssAAV1 is the most efficient in transducing murine hematopoietic stem cells (HSCs), but viral second-strand DNA synthesis remains a rate-limiting step. Subsequently, using double-stranded, self-complementary AAV (scAAV) vectors, serotypes 7 through 10, we observed that scAAV7 vectors also transduce murine HSCs efficiently. In the present study, we used scAAV1 and scAAV7 shuttle vectors to transduce HSCs in a murine bone marrow serial transplant model in vivo, which allowed examination of the AAV proviral integration pattern in the mouse genome, as well as recovery and nucleotide sequence analyses of AAV-HSC DNA junction fragments. The proviral genomes were stably integrated, and integration sites were localized to different mouse chromosomes. None of the integration sites was found to be in a transcribed gene, or near a cellular oncogene. None of the animals, monitored for up to 1 year, exhibited pathological abnormalities. Thus, AAV proviral integration-induced risk of oncogenesis was not found in our study, which provides functional confirmation of stable transduction of self-renewing multipotential HSCs by scAAV vectors as well as promise for the use of these vectors in the potential treatment of disorders of the hematopoietic system.
Mweya, Clement N.; Kimera, Sharadhuli I.; Mellau, Lesakit S. B.; Mboera, Leonard E. G.
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...
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
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.
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
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.
Hall-Mendelin, Sonja; Pyke, Alyssa T; Moore, Peter R; Mackay, Ian M; McMahon, Jamie L; Ritchie, Scott A; Taylor, Carmel T; Moore, Frederick A J; van den Hurk, Andrew F
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.
Rees, Robert C; McArdle, Stephanie; Mian, Shahid; Li, Geng; Ahmad, Murrium; Parkinson, Richard; Ali, Selman A
Disabled infectious single cycle-herpes simplex viruses (DISC-HSV) have been shown to be safe for use in humans and may be considered efficacious as vectors for immunogene therapy in cancer. Preclinical studies show that DISC-HSV is an efficient delivery system for cytokine genes and antigens. DISC-HSV infects a high proportion of cells, resulting in rapid gene expression for at least 72 h. The DISC-HSV-mGM-CSF vector, when inoculated into tumors, induces tumor regression in a high percentage of animals, concomitant with establishing a cytotoxic T-cell response, which is MHC class I restricted and directed against peptides of known tumor antigens. The inherent properties of DISC-HSV makes it a suitable vector for consideration in human immunogene therapy trials.
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.
Venter, G J; Paweska, J T; Van Dijk, A A; Mellor, P S; Tabachnick, W J
The susceptibility of field-collected Culicoides bolitinos to infection by oral ingestion of bluetongue virus serotypes 1, 3 and 4 (BLU 1, 3 and 4) was compared with that of field-collected C. imicola and laboratory reared C. variipennis sonorensis. The concentration of the virus per millilitre of bloodmeal was 10(5.0) and 10(6.0)TCID50 for BLU 4 and 10(7.2)TCID50 for BLU 1 and 3. Of 4927 C. bolitinos and 9585 C. imicola fed, 386 and 287 individual midges survived 10 days extrinsic incubation, respectively. Midges were assayed for the presence of virus using a microtitration assay on BHK-21 cells and/or an antigen capture ELISA. Infection prevalences for the different serotypes as determined by virus isolation ranged from 22.7 to 82.0% in C. bolitinos and from 1.9 to 9.8% in C. imicola; infection prevalences were highest for BLU 1, and lowest for BLU 4 in both species. The mean log10 TCID50 titre of the three BLU viruses per single fly was higher in C. bolitinos than in C. imicola. The results suggested that C. bolitinos populations are capable vectors of the BLU viruses in South Africa. A high correlation was found between virus isolation and ELISA results for the detection of BLU 1, and less for BLU 4; the ELISA failed to detect the presence of BLU 3 in infected flies. The C. v. sonorensis colonies had a significantly lower susceptibility to infection with BLU 1, 3 and 4 than C. bolitinos and C. imicola. However, since infection prevalence of C. v. sonorensis was determined only by ELISA, this finding may merely reflect the insensitivity of this assay at low virus titres, compared to virus isolation.
Hansen, Lars M.; Nielsen, Steen L.
Aphids are major vectors of plant viruses. Potato virus Y (PVY) is the most important aphid-transmitted virus affecting potato crops in Denmark. Because of a changed seed potato growing strategy, the seed potato area in Denmark is changing from regions with a low average temperature to regions...... with a higher average temperature. This means that the aphids may infest the potato crops earlier and the population development of the aphids may be faster, and consequently PVY may more easily become epidemic in seed potato crops. With a view to reducing the spread of PVY a 3-year experiment was carried out...... with a combination of mineral oil and insecticides. In 2005 and 2007 when a very high number of aphids were present, nearly all plants were infected with PVY. In 2006 with a lower number of aphids a smaller proportion of the plants were infected, and a tendency to a lower PVY incidence in mineral-oil treated plots...
Zimmer, Jean-Yves; Losson, Bertrand; Saegerman, Claude; Haubruge, Eric; Francis, Frédéric
Several species of Culicoides (Diptera: Ceratopogonidae) biting midges are biological vectors of bluetongue virus (BTV) and, as recently discovered, Schmallenberg virus (SBV) in northern Europe. Since their recent emergence in this part of the continent, these diseases that affect domestic and wild ruminants have caused considerable economic losses to the sheep and cattle industries. The substrates that are suitable for larval development of the main vector species are still relatively unknow...
Yu, Lu; Shi, Jing; Cao, Lianlian; Zhang, Guoping; Wang, Wenli; Hu, Deyu; Song, Baoan
Southern rice black-streaked dwarf virus (SRBSDV) has spread from the south of China to the north of Vietnam in the past few years, and has severely influenced rice production. However, previous study of traditional SRBSDV transmission method by the natural virus vector, the white-backed planthopper (WBPH, Sogatella furcifera), in the laboratory, researchers are frequently confronted with lack of enough viral samples due to the limited life span of infected vectors and rice plants and low virus acquisition and inoculation efficiency by the vector. Meanwhile, traditional mechanical inoculation of virus only apply to dicotyledon because of the higher content of lignin in the leaves of the monocot. Therefore, establishing an efficient and persistent-transmitting model, with a shorter virus transmission time and a higher virus transmission efficiency, for screening novel anti-SRBSDV drugs is an urgent need. In this study, we firstly reported a novel method for transmitting SRBSDV in rice using the bud-cutting method. The transmission efficiency of SRBSDV in rice was investigated via the polymerase chain reaction (PCR) method and the replication of SRBSDV in rice was also investigated via the proteomics analysis. Rice infected with SRBSDV using the bud-cutting method exhibited similar symptoms to those infected by the WBPH, and the transmission efficiency (>80.00%), which was determined using the PCR method, and the virus transmission time (30 min) were superior to those achieved that transmitted by the WBPH. Proteomics analysis confirmed that SRBSDV P1, P2, P3, P4, P5-1, P5-2, P6, P8, P9-1, P9-2, and P10 proteins were present in infected rice seedlings infected via the bud-cutting method. The results showed that SRBSDV could be successfully transmitted via the bud-cutting method and plants infected SRBSDV exhibited the symptoms were similar to those transmitted by the WBPH. Therefore, the use of the bud-cutting method to generate a cheap, efficient, reliable supply of
Ge, Jinying; Wang, Xijun; Tian, Meijie; Gao, Yuwei; Wen, Zhiyuan; Yu, Guimei; Zhou, Weiwei; Zu, Shulong; Bu, Zhigao
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.
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.
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
Lord, Cynthia C; Tabachnick, Walter J
Nonsystemic transmission, where a pathogen is transmitted between infected and uninfected vectors without the vertebrate host becoming viremic, may provide an explanation for transmission in systems where the vertebrate hosts have been difficult to identify. This transmission pathway had been previously demonstrated for tick-borne viruses and bacteria, but the recent demonstration for Simulium and vesicular stomatitis virus is the first for a blood-feeding insect. The epidemiology of vesicular stomatitis viruses has been difficult to understand, and nonsystemic transmission may be important. We use mathematical formulations of the basic reproduction number, R(0), to compare systemic and nonsystemic transmission. The absence of a latent period before host infectiousness in nonsystemic transmission may allow a more rapid increase in prevalence in the biting flies early in the development of a new outbreak. Aggregation of flies between hosts and at favored feeding sites on hosts will be important, but further data on nonsystemic transmission as a function of space and time are required to fully assess this pathway. The data needed to compare the two pathways and their relative roles in virus epidemiology are discussed.
Paiboonsukwong, Kittiphong; Ohbayashi, Fumi; Shiiba, Haruka; Aizawa, Emi; Yamashita, Takayuki; Mitani, Kohnosuke
Adeno-associated virus (AAV) vectors have been shown to correct a variety of mutations in human cells by homologous recombination (HR) at high rates, which can overcome insertional mutagenesis and transgene silencing, two of the major hurdles in conventional gene addition therapy of inherited diseases. We examined an ability of AAV vectors to repair a mutation in human hematopoietic cells by HR. We infected a human B-lymphoblastoid cell line (BCL) derived from a normal subject with an AAV, which disrupts the hypoxanthine phosphoribosyl transferase1 (HPRT1) locus, to measure the frequency of AAV-mediated HR in BCL cells. We subsequently constructed an AAV vector encoding the normal sequences from the Fanconi anemia group A (FANCA) locus to correct a mutation in the gene in BCL derived from a FANCA patient. Under optimal conditions, approximately 50% of BCL cells were transduced with an AAV serotype 2 (AAV-2) vector. In FANCA BCL cells, up to 0.016% of infected cells were gene-corrected by HR. AAV-mediated restoration of normal genotypic and phenotypic characteristics in FANCA-mutant cells was confirmed at the DNA, protein and functional levels. The results obtained in the present study indicate that AAV vectors may be applicable for gene correction therapy of inherited hematopoietic disorders.
Roberts, H C; Elbers, A R W; Conraths, F J; Holsteg, M; Hoereth-Boentgen, D; Gethmann, J; van Schaik, G
Surveillance for new emerging animal diseases from a European perspective is complicated by the non-harmonised approach across Member States for data capture, recording livestock populations and case definitions. In the summer of 2011, a new vector-borne Orthobunyavirus emerged in Northern Europe and for the first time, a coordinated approach to horizon scanning, risk communication, data and diagnostic test sharing allowed EU Member States to develop early predictions of the disease, its impact and risk management options. There are many different systems in place across the EU for syndromic and scanning surveillance and the differences in these systems have presented epidemiologists and risk assessors with concerns about their combined use in early identification of an emerging disease. The emergence of a new disease always will raise challenging issues around lack of capability and lack of knowledge; however, Schmallenberg virus (SBV) gave veterinary authorities an additional complex problem: the infection caused few clinical signs in adult animals, with no indication of the possible source and little evidence about its spread or means of transmission. This paper documents the different systems in place in some of the countries (Germany and the Netherlands) which detected disease initially and predicted its spread (to the UK) and how information sharing helped to inform early warning and risk assessment for Member States. Microarray technology was used to identify SBV as a new pathogen and data from the automated cattle milking systems coupled with farmer-derived data on reporting non-specific clinical signs gave the first indications of a widespread issue while the UK used meteorological modelling to map disease incursion. The coordinating role of both EFSA and the European Commission were vital as are the opportunities presented by web-based publishing for disseminating information to industry and the public. The future of detecting emerging disease looks more
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.
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
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
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.
Guillermo León M
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
da Moura, Aires Januário Fernandes; de Melo Santos, Maria Alice Varjal; Oliveira, Claudia Maria Fontes; Guedes, Duschinka Ribeiro Duarte; de Carvalho-Leandro, Danilo; da Cruz Brito, Maria Lidia; Rocha, Hélio Daniel Ribeiro; Gómez, Lara Ferrero; Ayres, Constância Flávia Junqueira
Dengue is an arboviral disease caused by dengue virus (DENV), whose main vectors are the mosquitoes Aedes aegypti and Aedes albopictus. A. aegypti is the only DENV vector in Cape Verde, an African country that suffered its first outbreak of dengue in 2009. However, little is known about the variation in the level of vector competence of this mosquito population to the different DENV serotypes. This study aimed to evaluate the vector competence of A. aegypti from the island of Santiago, Cape Verde, to four DENV serotypes and to detect DENV vertical transmission. Mosquitoes were fed on blood containing DENV serotypes and were dissected at 7, 14 and 21 days post-infection (dpi) to detect the virus in the midgut, head and salivary glands (SG) using RT-PCR. Additionally, the number of copies of viral RNA present in the SG was determined by qRT-PCR. Furthermore, eggs were collected in the field and adult mosquitoes obtained were analyzed by RT-PCR and the platelia dengue NS1 antigen kit to detect transovarial transmission. High rates of SG infection were observed for DENV-2 and DENV-3 whereas for DENV-1, viral RNA was only detected in the midgut and head. DENV-4 did not spread to the head or SG, maintaining the infection only in the midgut. The number of viral RNA copies in the SG did not vary significantly between DENV-2 and DENV-3 or among the different periods of incubation and the various titers of DENV tested. With respect to DENV surveillance in mosquitoes obtained from the eggs collected in the field, no samples were positive. Although no DENV positive samples were collected from the field in 2014, it is important to highlight that the A. aegypti population from Santiago Islands exhibited different degrees of susceptibility to DENV serotypes. This population showed a high vector competence for DENV-2 and DENV-3 strains and a low susceptibility to DENV-1 and DENV-4. Viral RNA copies in the SG remained constant for at least 21 dpi, which may enhance the vector
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
Florêncio Figueiredo Cavalcanti Neto
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
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
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.
Full Text Available Gene transfer through retrograde axonal transport of viral vectors offers a substantial advantage for analyzing roles of specific neuronal pathways or cell types forming complex neural networks. This genetic approach may also be useful in gene therapy trials by enabling delivery of transgenes into a target brain region distant from the injection site of the vectors. Pseudotyping of a lentiviral vector based on human immunodeficiency virus type 1 (HIV-1 with various fusion envelope glycoproteins composed of different combinations of rabies virus glycoprotein (RV-G and vesicular stomatitis virus glycoprotein (VSV-G enhances the efficiency of retrograde gene transfer in both rodent and nonhuman primate brains. The most recently developed lentiviral vector is a pseudotype with fusion glycoprotein type E (FuG-E, which demonstrates highly efficient retrograde gene transfer in the brain. The FuG-E–pseudotyped vector permits powerful experimental strategies for more precisely investigating the mechanisms underlying various brain functions. It also contributes to the development of new gene therapy approaches for neurodegenerative disorders, such as Parkinson’s disease, by delivering genes required for survival and protection into specific neuronal populations. In this review article, we report the properties of the FuG-E–pseudotyped vector, and we describe the application of the vector to neural circuit analysis and the potential use of the FuG-E vector in gene therapy for Parkinson’s disease.
Wu, Bing-Jie; Chow, Wah Soon; Liu, Yu-Jun; Shi, Lei; Jiang, Chuang-Dao
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.
Zhao Weihong; Zhong Li; Wu Jianqing; Chen Linyuan; Qing Keyun; Weigel-Kelley, Kirsten A.; Larsen, Steven H.; Shou Weinian; Warrington, Kenneth H.; Srivastava, Arun
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
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.
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
Xet Mull, A.M.
The purpose of this investigation was to characterize the yeast forms symbionts of Togo soles Orizicolus (YLSTo), through its morphologic description and locating in situ through microscopy of light, electronic microscopy of transmission and of sweeping and immuno microscopy. Likewise, molecular tests were carried out to classify phylogenetically the symbionts, utilizing partial sequences of the ribosomal DNA 18S. This study will permit to determine, the existence or not of interactions among the YLSTo and the insect vector, in the future. The paper of that interaction in the mechanism of trans ovarial transmission of the virus and the search of alternatives for the control of the disease in the rice. (S. Grainger) [es
Græsbøll, Kaare; Bødker, Rene; Enøe, Claes
Bluetongue is a disease of ruminants which reached Denmark in 2007. We present a process-based stochastic simulation model of vector-borne diseases, where host animals are not confined to a central geographic farm coordinate, but can be distributed onto pasture areas. Furthermore vectors fly freely...
Nebreda, M; Moreno, A; Pérez, N; Palacios, I; Seco-Fernández, V; Fereres, A
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.
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
Molaei, Goudarz; Armstrong, Philip M; Abadam, Charles F; Akaratovic, Karen I; Kiser, Jay P; Andreadis, Theodore G
Eastern equine encephalitis virus (EEEV) causes a highly pathogenic mosquito-borne zoonosis that is responsible for sporadic outbreaks of severe illness in humans and equines in the eastern USA. Culiseta (Cs.) melanura is the primary vector of EEEV in most geographic regions but its feeding patterns on specific avian and mammalian hosts are largely unknown in the mid-Atlantic region. The objectives of our study were to: 1) identify avian hosts of Cs. melanura and evaluate their potential role in enzootic amplification of EEEV, 2) assess spatial and temporal patterns of virus activity during a season of intense virus transmission, and 3) investigate the potential role of Cs. melanura in epidemic/epizootic transmission of EEEV to humans and equines. Accordingly, we collected mosquitoes at 55 sites in Suffolk, Virginia in 2013, and identified the source of blood meals in engorged mosquitoes by nucleotide sequencing PCR products of the mitochondrial cytochrome b gene. We also examined field-collected mosquitoes for evidence of infection with EEEV using Vector Test, cell culture, and PCR. Analysis of 188 engorged Cs. melanura sampled from April through October 2013 indicated that 95.2%, 4.3%, and 0.5% obtained blood meals from avian, mammalian, and reptilian hosts, respectively. American Robin was the most frequently identified host for Cs. melanura (42.6% of blood meals) followed by Northern Cardinal (16.0%), European Starling (11.2%), Carolina Wren (4.3%), and Common Grackle (4.3%). EEEV was detected in 106 mosquito pools of Cs. melanura, and the number of virus positive pools peaked in late July with 22 positive pools and a Maximum Likelihood Estimation (MLE) infection rate of 4.46 per 1,000 mosquitoes. Our findings highlight the importance of Cs. melanura as a regional EEEV vector based on frequent feeding on virus-competent bird species. A small proportion of blood meals acquired from mammalian hosts suggests the possibility that this species may occasionally
Killiny, N; Harper, S J; Alfaress, S; El Mohtar, C; Dawson, W O
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
Stable flies, which are notorious pests of cattle and other livestock, were suspected of transmitting West Nile virus (WNV) among American white pelicans at the Medicine Lake Wildlife Refuge in northeastern Montana in 2006-2007. However the ability of stable flies to transmit the virus was unknown. ...
Kamencayová, M; Košík, I; Hunková, J; Subr, Z W
PB1-F2 protein of influenza A virus (IAV) was cloned in a plum pox virus (PPV) genome-based vector and attempts to express it in biolistically transfected Nicotiana benthamiana plants were performed. The vector-insert construct replicated in infected plants properly and was stable during repeated passage by mechanical inoculation, as demonstrated by disease symptoms and immunoblot detection of PPV capsid protein, while PB1-F2-specific band was more faint. We showed that it was due its low solubility. Modification of sample preparation (denaturation/solubilization preceding the centrifugation of cell debris) led to substantial signal enhancement. Maximal level of PB1-F2 expression in plants was observed 12 days post inoculation (dpi). Only 1% SDS properly solubilized the protein, other detergents were much less efficient. Solubilization with 8M urea released approximately 50% of PB1-F2 from the plant tissues, thus the treatment with this removable chaotropic agent may be a good starting point for the purification of the protein for eventual functional studies in the future.
Voronin, Yegor A.; Pathak, Vinay K.
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
Richards, Stephanie L; Anderson, Sheri L; Lord, Cynthia C; Tabachnick, Walter J
Female Culex nigripalpus were fed blood containing a low dose (6.3±0.01 logs plaque-forming units (PFU)/mL) or high dose (7.3±0.1 logs PFU/mL) of West Nile virus (WNV) and maintained at 28°C for incubation periods (IPs) of 6 or 12 days. Vector competence was measured using rates of infection (% with WNV-positive bodies), dissemination (% infected with WNV-positive legs), and transmission (% infected with WNV-positive saliva). Infection rates were not influenced by dose or IP. Dissemination rates were significantly higher at the high dose, and this was dependent on IP. Despite 100% infection and 90% dissemination in the most permissive treatment of high dose and 12 days, only 11% transmission was observed. Virus titers in body and leg tissues were significantly lower at the low dose and the titers were not influenced by IP. We show that not all mosquitoes with infections and/or disseminated infections transmit WNV under the conditions of this test. Therefore, characterizing the transmission ability of a vector population using infection or dissemination as indicators of transmission may provide inaccurate information. The complex relationships between infection, dissemination, and transmission must be evaluated under a variety of biological and environmental conditions to begin to assess the epidemiological risk of natural mosquito populations.
Cheng, Yang-Jian; Liang, Ji-Xuan; Li, Qing-Ge
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.
van Kuppeveld, Frank J M; de Jong, Arjan; Dijkman, Henri B P M; Andino, Raul; Melchers, Willem J G
Development of human cervical carcinomas is associated with infection by certain human papillomavirus (HPV) types. Thus, protection against HPV infection through vaccination may prevent development of cervical cancer. The purpose of this study was to investigate the possibility of using a poliovirus recombinant vector to induce immunity against HPV. A poliovirus recombinant was constructed which contained the complete coding sequence of the HPV 16 major capsid protein L1, between the P1 and P2 region of the poliovirus polyprotein. A replication-competent virus was obtained after transfection of the recombinant RNA into tissue culture cells. Electron microscopically examination of cells infected with the poliovirus-HPV L1 recombinant indicated that HPV 16 L1 self-assembles into virus-like particles. To investigate the immunological response in vivo, susceptible transgenic mice carrying the poliovirus receptor were infected with the recombinant poliovirus. In all mice a modest but consistent immune response against HPV 16 was observed. Based on these results, the potential for picornavirus-derived vectors in vaccine development against HPV infection is discussed.
de Haro, Luis A; Dumón, Analía D; Mattio, María F; Argüello Caro, Evangelina Beatriz; Llauger, Gabriela; Zavallo, Diego; Blanc, Hervé; Mongelli, Vanesa C; Truol, Graciela; Saleh, María-Carla; Asurmendi, Sebastián; Del Vas, Mariana
Plant reoviruses are able to multiply in gramineae plants and delphacid vectors encountering different defense strategies with unique features. This study aims to comparatively assess alterations of small RNA (sRNA) populations in both hosts upon virus infection. For this purpose, we characterized the sRNA profiles of wheat and planthopper vectors infected by Mal de Río Cuarto virus (MRCV, Fijivirus, Reoviridae ) and quantified virus genome segments by quantitative reverse transcription PCR We provide evidence that plant and insect silencing machineries differentially recognize the viral genome, thus giving rise to distinct profiles of virus-derived small interfering RNAs (vsiRNAs). In plants, most of the virus genome segments were targeted preferentially within their upstream sequences and vsiRNAs mapped with higher density to the smaller genome segments than to the medium or larger ones. This tendency, however, was not observed in insects. In both hosts, vsiRNAs were equally derived from sense and antisense RNA strands and the differences in vsiRNAs accumulation did not correlate with mRNAs accumulation. We also established that the piwi-interacting RNA (piRNA) pathway was active in the delphacid vector but, contrary to what is observed in virus-infected mosquitoes, virus-specific piRNAs were not detected. This work contributes to the understanding of the silencing response in insect and plant hosts.
Joanne, Sylvia; Vythilingam, Indra; Teoh, Boon-Teong; Leong, Cherng-Shii; Tan, Kim-Kee; Wong, Meng-Li; Yugavathy, Nava; AbuBakar, Sazaly
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.
Mahmood, Farida; Chiles, Robert E; Fang, Ying; Green, Emily N; Reisen, William K
The vector competence of Culex tarsalis Coquillett for the BFS 1703 strain of western equine encephalomyelitis virus (WEEV) changed significantly as a function of time after infection, mosquito genotype, and infectious virus dose. After ingesting a high virus dose (5 log10 plaque-forming units [PFU]/0.1 ml), female of the susceptible high virus producer (HVP) strain rapidly amplified the virus, developed a disseminated infection, and efficiently transmitted WEEV by 4 days postinfection (dpi). The quantity of virus expectorated peaked at 4 dpi (mean 3.4 log10 PFU), and the percentage of females transmitting per os peaked at 7 dpi (80%); both measures of transmission subsequently decreased to low levels throughout the remainder of infected life. HVP females imbibing a low virus dose (3 log10 PFU/0.1 ml) were infected less frequently and took longer to amplify virus to levels recorded for the high virus dose group and did not transmit virus efficiently, thereby indicating midgut infection and escape barriers were dose and time dependent. These data emphasized the importance of elevated avian viremias in Cx. tarsalis vector competence. Females from the WEEV-resistant (WR) strain and two wild-type strains from Kern and Riverside counties were significantly less susceptible to infection at both high and low doses than was the HVP strain. Overall, females with a high virus titer more frequently had a disseminated infection, but there did not seem to be a distinct threshold demarcating this relationship. In marked contrast, all infected females transmitting virus had body titers >4.3 log10 PFU, and most had titers >4.8 log10 PFU. These data indicated that not all females with a disseminated infection transmitted virus because of the presence of one or more salivary gland barriers.
Mwaengo, D; Lorenzo, G; Iglesias, J; Warigia, M; Sang, R; Bishop, R P; Brun, A
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.
Filone, Claire Marie; Heise, Mark; Doms, Robert W.; Bertolotti-Ciarlet, Andrea
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
Galal Fatma H
Full Text Available Abstract Background Rift Valley fever (RVF is an acute febrile arthropod-borne viral disease of man and animals caused by a member of the Phlebovirus genus, one of the five genera in the family Bunyaviridae. RVF virus (RVFV is transmitted between animals and human by mosquitoes, particularly those belonging to the Culex, Anopheles and Aedes genera. Methods Experiments were designed during RVF outbreak, 2007 in Sudan to provide an answer about many raised questions about the estimated role of vector in RVFV epidemiology. During this study, adult and immature mosquito species were collected from Khartoum and White Nile states, identified and species abundance was calculated. All samples were frozen individually for further virus detection. Total RNA was extracted from individual insects and RVF virus was detected from Culex, Anopheles and Aedes species using RT-PCR. In addition, data were collected about human cases up to November 24th, 2007 to asses the situation of the disease in affected states. Furthermore, a historical background of the RVF outbreaks was discussed in relation to global climatic anomalies and incriminated vector species. Results A total of 978 mosquitoes, belonging to 3 genera and 7 species, were collected during Sudan outbreak, 2007. Anopheles gambiae arabiensis was the most frequent species (80.7% in White Nile state. Meanwhile, Cx. pipiens complex was the most abundant species (91.2% in Khartoum state. RT-PCR was used and successfully amplified 551 bp within the M segment of the tripartite negative-sense single stranded RNA genome of RVFV. The virus was detected in female, male and larval stages of Culex and Anopheles species. The most affected human age interval was 15-29 years old followed by ≥ 45 years old, 30-44 years old, and then 5-14 years old. Regarding to the profession, housewives followed by farmers, students, shepherd, workers and the free were more vulnerable to the infection. Furthermore, connection between
Casteel, Clare L; De Alwis, Manori; Bak, Aurélie; Dong, Haili; Whitham, Steven A; Jander, Georg
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.
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.
Batallán, Gonzalo P; Estallo, Elizabet L; Flores, Fernando S; Sartor, Paolo; Contigiani, Marta S; Almirón, Walter R
In Argentina the St. Louis Encephalitis virus (SLEV) is an endemic and widely distributed pathogen transmitted by the cosmopolitan mosquito Culex quinquefasciatus. During two outbreaks in Córdoba city, in 2005 and 2010, Culex interfor was also found infected, but its role as vector of SLEV is poorly known. This mosquito species is distributed from central Argentina to southern Brazil. The primary aim of this study was to analyze the population dynamic of Cx. interfor and Cx. quinquefasciatus in three different environments (urban, suburban and non-urban) in relation to remotely sensed environmental data for vegetation (NDVI and NDWI) and temperature (brightness temperature). Cx. quinquefasciatus and Cx. interfor were found at the three sampled sites, being both the most abundant Culex species, with peaks in early and midsummer. Temporal distribution patterns of both mosquito species were highly correlated in a non-urban area of high SLEV risk transmission. Cx. quinquefasciatus and Cx. interfor were associated with the most urbanized site and the non-urban environment, respectively; high significant correlations were detected between vegetation indices and abundance of both mosquito species confirming these associations. These data provide a foundation for building density maps of these two SLEV mosquito vectors using remotely sensed data to help inform vector control programs. Copyright © 2015 Elsevier B.V. All rights reserved.
Campbell, Lindsay P; Alexander, Alana M
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: email@example.com.
Geisbert, Joan B; Shedlock, Devon J; Xu, Ling; Lamoreaux, Laurie; Custers, Jerome H. H. V; Popernack, Paul M; Yang, Zhi-Yong; Pau, Maria G; Roederer, Mario; Koup, Richard A; Goudsmit, Jaap; Jahrling, Peter B; Nabel, Gary J
Background Ebola virus causes a hemorrhagic fever syndrome that is associated with high mortality in humans. In the absence of effective therapies for Ebola virus infection, the development of a vaccine becomes an important strategy to contain outbreaks. Immunization with DNA and/or replication-defective adenoviral vectors (rAd) encoding the Ebola glycoprotein (GP) and nucleoprotein (NP) has been previously shown to confer specific protective immunity in nonhuman primates. GP can exert cytopathic effects on transfected cells in vitro, and multiple GP forms have been identified in nature, raising the question of which would be optimal for a human vaccine. Methods and Findings To address this question, we have explored the efficacy of mutant GPs from multiple Ebola virus strains with reduced in vitro cytopathicity and analyzed their protective effects in the primate challenge model, with or without NP. Deletion of the GP transmembrane domain eliminated in vitro cytopathicity but reduced its protective efficacy by at least one order of magnitude. In contrast, a point mutation was identified that abolished this cytopathicity but retained immunogenicity and conferred immune protection in the absence of NP. The minimal effective rAd dose was established at 1010 particles, two logs lower than that used previously. Conclusions Expression of specific GPs alone vectored by rAd are sufficient to confer protection against lethal challenge in a relevant nonhuman primate model. Elimination of NP from the vaccine and dose reductions to 1010 rAd particles do not diminish protection and simplify the vaccine, providing the basis for selection of a human vaccine candidate. PMID:16683867
Nancy J Sullivan
Full Text Available Ebola virus causes a hemorrhagic fever syndrome that is associated with high mortality in humans. In the absence of effective therapies for Ebola virus infection, the development of a vaccine becomes an important strategy to contain outbreaks. Immunization with DNA and/or replication-defective adenoviral vectors (rAd encoding the Ebola glycoprotein (GP and nucleoprotein (NP has been previously shown to confer specific protective immunity in nonhuman primates. GP can exert cytopathic effects on transfected cells in vitro, and multiple GP forms have been identified in nature, raising the question of which would be optimal for a human vaccine.To address this question, we have explored the efficacy of mutant GPs from multiple Ebola virus strains with reduced in vitro cytopathicity and analyzed their protective effects in the primate challenge model, with or without NP. Deletion of the GP transmembrane domain eliminated in vitro cytopathicity but reduced its protective efficacy by at least one order of magnitude. In contrast, a point mutation was identified that abolished this cytopathicity but retained immunogenicity and conferred immune protection in the absence of NP. The minimal effective rAd dose was established at 10(10 particles, two logs lower than that used previously.Expression of specific GPs alone vectored by rAd are sufficient to confer protection against lethal challenge in a relevant nonhuman primate model. Elimination of NP from the vaccine and dose reductions to 10(10 rAd particles do not diminish protection and simplify the vaccine, providing the basis for selection of a human vaccine candidate.
Ruder, M G; Stallknecht, D E; Allison, A B; Mead, D G; Carter, D L; Howerth, E W
Epizootic hemorrhagic disease viruses (EHDVs) are orbiviruses transmitted by Culicoides biting midges to domestic and wild ruminants. EHDV-1 and EHDV-2 are endemic in the United States, where epizootic hemorrhagic disease is the most significant viral disease of white-tailed deer (WTD;Odocoileus virginianus) and reports of epizootic hemorrhagic disease in cattle are increasing. In 2006, a reassortant EHDV-6 was isolated from dead WTD in Indiana and has been detected each subsequent year over a wide geographic region. Since EHDV-6 is not a historically endemic serotype in the United States, it is important to understand infection outcome in potential hosts. Specifically, we aimed to evaluate the pathogenicity of the virus in 2 primary US ruminant hosts (WTD and cattle) and the susceptibility of a confirmed US vector (Culicoides sonorensis). Five WTD and 4 cattle were inoculated with >10(6)TCID50EHDV-6 by intradermal and subcutaneous injection. All 5 WTD exhibited moderate to severe disease, and 3 died. Viremia was first detected 3 to 5 days postinfection (dpi) with surviving animals seroconverting by 10 dpi. Two of 4 inoculated cattle had detectable viremia, 5 to 10 dpi and 7 to 24 dpi, respectively. No clinical, hematologic, or pathologic abnormalities were observed. Antibodies were detected by 10 dpi in 3 of 4 cows.C. sonorensis were fed on WTD blood spiked with EHDV-6 and held for 4 to 14 days postfeeding at 25°C. From 4 to 14 days postfeeding, 19 of 171 midges were virus isolation positive and 6 of 171 had ≥10(2.7)TCID50EHDV-6. Although outcomes varied, these studies demonstrate the susceptibility of ruminant and vector hosts in the United States for this recently emerged EHDV serotype. © The Author(s) 2015.
Kassem, Mona A; Juarez, Miguel; Gómez, Pedro; Mengual, Carmen M; Sempere, Raquel N; Plaza, María; Elena, Santiago F; Moreno, Aranzazu; Fereres, Alberto; Aranda, Miguel A
The genetic variability of a Cucurbit aphid-borne yellows virus (CABYV) (genus Polerovirus, family Luteoviridae) population was evaluated by determining the nucleotide sequences of two genomic regions of CABYV isolates collected in open-field melon and squash crops during three consecutive years in Murcia (southeastern Spain). A phylogenetic analysis showed the existence of two major clades. The sequences did not cluster according to host, year, or locality of collection, and nucleotide similarities among isolates were 97 to 100 and 94 to 97% within and between clades, respectively. The ratio of nonsynonymous to synonymous nucleotide substitutions reflected that all open reading frames have been under purifying selection. Estimates of the population's genetic diversity were of the same magnitude as those previously reported for other plant virus populations sampled at larger spatial and temporal scales, suggesting either the presence of CABYV in the surveyed area long before it was first described, multiple introductions, or a particularly rapid diversification. We also determined the full-length sequences of three isolates, identifying the occurrence and location of recombination events along the CABYV genome. Furthermore, our field surveys indicated that Aphis gossypii was the major vector species of CABYV and the most abundant aphid species colonizing melon fields in the Murcia (Spain) region. Our surveys also suggested the importance of the weed species Ecballium elaterium as an alternative host and potential virus reservoir.
Mardulyn, Patrick; Goffredo, Maria; Conte, Annamaria; Hendrickx, Guy; Meiswinkel, Rudolf; Balenghien, Thomas; Sghaier, Soufien; Lohr, Youssef; Gilbert, Marius
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.
Bricault, Christine A.; Perry, Keith L.
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
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)
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.
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.
van Dodewaard, Caitlin A M; Richards, Stephanie L; Harris, Jonathan W
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: firstname.lastname@example.org.
Ito, Akira; Takahashi, Tetsuya; Kameyama, Yujiro; Kawabe, Yoshinori; Kamihira, Masamichi
For tissue engineering purposes, retroviral vectors represent an efficient method of delivering exogenous genes such as growth factors to injured tissues because gene-transduced cells can produce stable and constant levels of the gene product. However, retroviral vector technology suffers from low yields. In the present study, we used magnetite nanoparticles and magnetic force to concentrate the retroviral vectors to enhance the transduction efficiency and to enable their magnetic manipulation. Magnetite nanoparticles modified with cationic liposomes were added to a solution containing a retroviral vector pseudotyped with vesicular stomatitis virus glycoprotein. The magnetic particles that captured the viral vectors were collected using a magnetic force and seeded into mouse neuroblastoma Neuro2a cells. The viral titer was up to 55 times greater (up to 3 x 10(8) infectious units/mL). Additionally, the magnetically labeled retroviral vectors can be directed to the desired regions for infection by applying magnetic fields, and micro-patterns of gene-transduced cell regions could be created on a cellular monolayer using micro-patterned magnetic concentrators. These results suggest that this technique provides a promising approach to capturing and concentrating viral vectors, thus achieving high transduction efficiency and the ability to deliver genes to a specific injured site by applying a magnetic field.
Schlecht-Louf, Géraldine; Mangeney, Marianne; El-Garch, Hanane; Lacombe, Valérie; Poulet, Hervé; Heidmann, Thierry
We previously delineated a highly conserved immunosuppressive (IS) domain within murine and primate retroviral envelope proteins that is critical for virus propagation in vivo. The envelope-mediated immunosuppression was assessed by the ability of the proteins, when expressed by allogeneic tumor cells normally rejected by engrafted mice, to allow these cells to escape, at least transiently, immune rejection. Using this approach, we identified key residues whose mutation (i) specifically abolishes immunosuppressive activity without affecting the "mechanical" function of the envelope protein and (ii) significantly enhances humoral and cellular immune responses elicited against the virus. The objective of this work was to study the immunosuppressive activity of the envelope protein (p15E) of feline leukemia virus (FeLV) and evaluate the effect of its abolition on the efficacy of a vaccine against FeLV. Here we demonstrate that the FeLV envelope protein is immunosuppressive in vivo and that this immunosuppressive activity can be "switched off" by targeted mutation of a specific amino acid. As a result of the introduction of the mutated envelope sequence into a previously well characterized canarypox virus-vectored vaccine (ALVAC-FeLV), the frequency of vaccine-induced FeLV-specific gamma interferon (IFN-γ)-producing cells was increased, whereas conversely, the frequency of vaccine-induced FeLV-specific interleukin-10 (IL-10)-producing cells was reduced. This shift in the IFN-γ/IL-10 response was associated with a higher efficacy of ALVAC-FeLV against FeLV infection. This study demonstrates that FeLV p15E is immunosuppressive in vivo, that the immunosuppressive domain of p15E can modulate the FeLV-specific immune response, and that the efficacy of FeLV vaccines can be enhanced by inhibiting the immunosuppressive activity of the IS domain through an appropriate mutation.
The widespread epidemic of Zika virus infection in South and Central America and the Caribbean in 2015, along with the increased incidence of microcephaly in fetuses born to mothers infected with Zika virus and the potential for worldwide spread, indicate the need to review the current literature regarding vectors, reservoirs, and amplification hosts. The virus has been isolated in Africa in mosquitoes of the genera Aedes, Anopheles, and Mansonia, and in Southeast Asia and the Pacific area in mosquitoes of the genus Aedes. Aedes albopictus has invaded several countries in Central Africa and all Mediterranean countries, and continues to spread throughout Central and Northern Europe. The wide distribution of the virus in animal hosts and vectors favors the emergence of recombinants. The virus has been isolated in monkeys, and antibodies have been detected in domestic sheep, goats, horses, cows, ducks, rodents, bats, orangutans, and carabaos. It is a public health imperative to define the domestic and wild animal reservoirs, amplification hosts, and vector capacity of the genera Aedes, Anopheles, and Mansonia. These variables will define the geographic distribution of Zika virus along with the indicated timing and scale of the environmental public health interventions worldwide. Copyright © 2016 The Author. Published by Elsevier Ltd.. All rights reserved.
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
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
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
Full Text Available 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 10(7.8 and 10(8.5 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.
Zhong, Li; Li, Baozheng; Mah, Cathryn S.; Govindasamy, Lakshmanan; Agbandje-McKenna, Mavis; Cooper, Mario; Herzog, Roland W.; Zolotukhin, Irene; Warrington, Kenneth H.; Weigel-Van Aken, Kirsten A.; Hobbs, Jacqueline A.; Zolotukhin, Sergei; Muzyczka, Nicholas; Srivastava, Arun
Recombinant adeno-associated virus 2 (AAV2) vectors are in use in several Phase I/II clinical trials, but relatively large vector doses are needed to achieve therapeutic benefits. Large vector doses also trigger an immune response as a significant fraction of the vectors fails to traffic efficiently to the nucleus and is targeted for degradation by the host cell proteasome machinery. We have reported that epidermal growth factor receptor protein tyrosine kinase (EGFR-PTK) signaling negatively affects transduction by AAV2 vectors by impairing nuclear transport of the vectors. We have also observed that EGFR-PTK can phosphorylate AAV2 capsids at tyrosine residues. Tyrosine-phosphorylated AAV2 vectors enter cells efficiently but fail to transduce effectively, in part because of ubiquitination of AAV capsids followed by proteasome-mediated degradation. We reasoned that mutations of the surface-exposed tyrosine residues might allow the vectors to evade phosphorylation and subsequent ubiquitination and, thus, prevent proteasome-mediated degradation. Here, we document that site-directed mutagenesis of surface-exposed tyrosine residues leads to production of vectors that transduce HeLa cells ≈10-fold more efficiently in vitro and murine hepatocytes nearly 30-fold more efficiently in vivo at a log lower vector dose. Therapeutic levels of human Factor IX (F.IX) are also produced at an ≈10-fold reduced vector dose. The increased transduction efficiency of tyrosine-mutant vectors is due to lack of capsid ubiquitination and improved intracellular trafficking to the nucleus. These studies have led to the development of AAV vectors that are capable of high-efficiency transduction at lower doses, which has important implications in their use in human gene therapy. PMID:18511559
Cui, Hongguang; Wang, Aiming
RNA silencing is a powerful technology for molecular characterization of gene functions in plants. A commonly used approach to the induction of RNA silencing is through genetic transformation. A potent alternative is to use a modified viral vector for virus-induced gene silencing (VIGS) to degrade RNA molecules sharing similar nucleotide sequence. Unfortunately, genomic studies in many allogamous woody perennials such as peach are severely hindered because they have a long juvenile period and are recalcitrant to genetic transformation. Here, we report the development of a viral vector derived from Prunus necrotic ringspot virus (PNRSV), a widespread fruit tree virus that is endemic in all Prunus fruit production countries and regions in the world. We show that the modified PNRSV vector, harbouring the sense-orientated target gene sequence of 100-200 bp in length in genomic RNA3, could efficiently trigger the silencing of a transgene or an endogenous gene in the model plant Nicotiana benthamiana. We further demonstrate that the PNRSV-based vector could be manipulated to silence endogenous genes in peach such as eukaryotic translation initiation factor 4E isoform (eIF(iso)4E), a host factor of many potyviruses including Plum pox virus (PPV). Moreover, the eIF(iso)4E-knocked down peach plants were resistant to PPV. This work opens a potential avenue for the control of virus diseases in perennial trees via viral vector-mediated silencing of host factors, and the PNRSV vector may serve as a powerful molecular tool for functional genomic studies of Prunus fruit trees. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.
Tchouassi, David P.; Bastos, Armanda D. S.; Sole, Catherine L.; Diallo, Mawlouth; Lutomiah, Joel; Mutisya, James; Mulwa, Francis; Borgemeister, Christian; Sang, Rosemary; Torto, Baldwyn
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
David P Tchouassi
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.
Young, Katherine I; Mundis, Stephanie; Widen, Steven G; Wood, Thomas G; Tesh, Robert B; Cardosa, Jane; Vasilakis, Nikos; Perera, David; Hanley, Kathryn A
Mosquito-borne dengue virus (DENV) is maintained in a sylvatic, enzootic cycle of transmission between canopy-dwelling non-human primates and Aedes mosquitoes in Borneo. Sylvatic DENV can spill over into humans living in proximity to forest foci of transmission, in some cases resulting in severe dengue disease. The most likely vectors of such spillover (bridge vectors) in Borneo are Ae. albopictus and Ae. niveus. Borneo is currently experiencing extensive forest clearance. To gauge the effect of this change in forest cover on the likelihood of sylvatic DENV spillover, it is first necessary to characterize the distribution of bridge vectors in different land cover types. In the current study, we hypothesized that Ae. niveus and Ae. albopictus would show significantly different distributions in different land cover types; specifically, we predicted that Ae. niveus would be most abundant in forests whereas Ae. albopictus would have a more even distribution in the landscape. Mosquitoes were collected from a total of 15 sites using gravid traps and a backpack aspirator around Kampong Puruh Karu, Sarawak, Malaysian Borneo, where sylvatic DENV spillover has been documented. A total of 2447 mosquitoes comprising 10 genera and 4 species of Aedes, were collected over the three years, 2013, 2014 and 2016, in the three major land cover types in the area, homestead, agriculture and forest. Mosquitoes were identified morphologically, pooled by species and gender, homogenized, and subject to DNA barcoding of each Aedes species and to arbovirus screening. As predicted, Ae. niveus was found almost exclusively in forests whereas Ae. albopictus was collected in all land cover types. Aedes albopictus was significantly (P = 0.04) more abundant in agricultural fields than forests. Sylvatic DENV was not detected in any Aedes mosquito pools, however genomes of 14 viruses were detected using next generation sequencing. Land cover type affects the abundance and distribution of the most
Belliure, B.; Janssen, A.; Sabelis, M.W.
Herbivores can profit from vectoring plant pathogens because the induced defence of plants against pathogens sometimes interferes with the induced defence of plants against herbivores. Plants can also defend themselves indirectly by the action of the natural enemies of the herbivores. It is unknown
1972-01-01In 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 canalThe most anterior part of
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.
Environmental temperature has been shown to affect the ability of mosquitoes to transmit numerous arboviruses and for Rift Valley fever virus (RVFV) in particular. We evaluated the effect of incubation temperatures ranging from 14-26ºC on infection, dissemination, and transmission rates for Culex ta...
van Mölken, Tamara; Caluwe, Hannie de; Hordijk, Cornelis A.
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 ...
Lommen, S.T.E.; Conijn, C.G.M.; Lemmers, M.E.C.; Pham, K.T.K.; Kock, de M.J.D.
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
Bichaud, Laurence; Souris, Marc; Mary, Charles; Ninove, Laëtitia; Thirion, Laurence; Piarroux, Raphaël P; Piarroux, Renaud; De Lamballerie, Xavier; Charrel, Rémi N
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
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
Riezebos-Brilman, Annelies; Regts, Joke; Chen, Margaret; Wilschut, Jan; Daemen, Toos
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
Janke, M.; Peeters, B.P.H.; Leeuw, de O.S.; Moormann, R.J.M.; Arnold, A.; Fournier, P.; Schirrmacher, V.
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
Conclusions: It is a public health imperative to define the domestic and wild animal reservoirs, amplification hosts, and vector capacity of the genera Aedes, Anopheles, and Mansonia. These variables will define the geographic distribution of Zika virus along with the indicated timing and scale of the environmental public health interventions worldwide.
Bahrami, Shervin; Jespersen, Thomas; Pedersen, Finn Skou
The envelope protein of retroviruses is responsible for viral entry into host cells. Here, we describe a mutational library approach to dissect functional domains of the envelope protein involving a retroviral vector, which expresses both the envelope protein of Akv murine leukemia virus (MLV) an...
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...
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.
Chao, Chun-Nun; Yang, Yu-Hsuan; Wu, Mu-Sheng; Chou, Ming-Chieh; Fang, Chiung-Yao; Lin, Mien-Chun; Tai, Chien-Kuo; Shen, Cheng-Huang; Chen, Pei-Lain; Chang, Deching; Wang, Meilin
Glioblastoma multiforme (GBM), the most common malignant brain tumor, has a short period of survival even with recent multimodality treatment. The neurotropic JC polyomavirus (JCPyV) infects glial cells and oligodendrocytes and causes fatal progressive multifocal leukoencephalopathy in patients with AIDS. In this study, a possible gene therapy strategy for GBM using JCPyV virus-like particles (VLPs) as a gene delivery vector was investigated. We found that JCPyV VLPs were able to deliver the GFP reporter gene into tumor cells (U87-MG) for expression. In an orthotopic xenograft model, nude mice implanted with U87 cells expressing the near-infrared fluorescent protein and then treated by intratumoral injection of JCPyV VLPs carrying the thymidine kinase suicide gene, combined with ganciclovir administration, exhibited significantly prolonged survival and less tumor fluorescence during the experiment compared with controls. Furthermore, JCPyV VLPs were able to protect and deliver a suicide gene to distal subcutaneously implanted U87 cells in nude mice via blood circulation and inhibit tumor growth. These findings show that metastatic brain tumors can be targeted by JCPyV VLPs carrying a therapeutic gene, thus demonstrating the potential of JCPyV VLPs to serve as a gene therapy vector for the far highly treatment-refractory GBM.
Thomas, Stephen J.; Aldstadt, Jared; Jarman, Richard G.; Buddhari, Darunee; Yoon, In-Kyu; Richardson, Jason H.; Ponlawat, Alongkot; Iamsirithaworn, Sopon; Scott, Thomas W.; Rothman, Alan L.; Gibbons, Robert V.; Lambrechts, Louis; Endy, Timothy P.
Dengue is of public health importance in tropical and sub-tropical regions. Dengue virus (DENV) transmission dynamics was studied in Kamphaeng Phet Province, Thailand, using an enhanced spatiotemporal surveillance of 93 hospitalized subjects with confirmed dengue (initiates) and associated cluster individuals (associates) with entomologic sampling. A total of 438 associates were enrolled from 208 houses with household members with a history of fever, located within a 200-m radius of an initiate case. Of 409 associates, 86 (21%) had laboratory-confirmed DENV infection. A total of 63 (1.8%) of the 3,565 mosquitoes collected were dengue polymerase chain reaction positive (PCR+). There was a significant relationship between spatial proximity to the initiate case and likelihood of detecting DENV from associate cases and Aedes mosquitoes. The viral detection rate from human hosts and mosquito vectors in this study was higher than previously observed by the study team in the same geographic area using different methodologies. We propose that the sampling strategy used in this study could support surveillance of DENV transmission and vector interactions. PMID:25986580
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
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
Full Text Available 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
Ahani, Roshank; Roohvand, Farzin; Cohan, Reza Ahangari; Etemadzadeh, Mohammad Hossein; Mohajel, Nasir; Behdani, Mahdi; Shahosseini, Zahra; Madani, Navid; Azadmanesh, Kayhan
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.
Anderson, Sheri L; Richards, Stephanie L; Tabachnick, Walter J; Smartt, Chelsea T
Culex pipiens quinquefasciatus were fed blood containing either 7.0 +/- 0.1 logs plaque-forming units (pfu)/ml (high dose) or 5.9 +/- 0.1 logs pfu/ml (low dose) of West Nile virus and held at extrinsic incubation temperatures (EIT) of 28 degrees C or 25 degrees C. Approximately 20 mosquitoes per dose were collected after incubation periods (IP) of 4, 6, 8, and 12 days postinfection (dpi). Infection rates were influenced by EIT and virus dose but not by IP. Body titer was significantly higher for mosquitoes fed the high dose and held at 28 degrees C at the later IPs (6, 8, and 12 dpi). However, leg titer was significantly higher for mosquitoes at the later IPs but did not differ between EITs or doses. Because infection rates varied with EIT and dose, there is likely a midgut infection barrier influenced by these factors that is not influenced by IP. Dissemination rates were influenced by all 3 factors consistent with the presence of a midgut escape barrier. Dissemination rate, body titer, and leg titer were dependent on IP, indicating the need to investigate multiple time points in vector competence studies to elucidate critical events in infection and dissemination.
Xu, Hong-Xing; He, Xiao-Chan; Zheng, Xu-Song; Yang, Ya-Jun; Lu, Zhong-Xian
Rice black streak dwarf virus (RBSDV) is transmitted by the small brown planthopper (SBPH), Laodelphax striatellus (Fallen). Non-vector rice brown planthopper (BPH), Nilaparvata lugens (Stål), shares the same host rice plants with SBPH in paddy fields. The changes in nutritional composition of rice plants infected by RBSDV and the ecological fitness of BPH feeding on the infected plants were studied under both artificial climate chamber and field conditions. Contents of 16 detected amino acids and soluble sugar in RBSDV infected rice plants were higher than those in the healthy ones. On the diseased plants BPH had significantly higher nymphal survival rates, nymphal duration of the males, weight of the female adults, as well as egg hatchability compared to BPH being fed on healthy plants. However, there was no obvious difference in female nymph duration, longevity and fecundity. Defense enzymes (superoxidase dismutase, SOD and catalase, CAT) and detoxifying enzymes (carboxylesterase, CAE and glutathione S-transferase, GST) in BPH adults fed on diseased plants had markedly higher activities. The results indicate rice plants infected by RBSDV improved the ecological fitness of the brown planthopper, a serious pest but not a transmitter of the RBSDV virus. © 2013 Institute of Zoology, Chinese Academy of Sciences.
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.
Full Text Available Abstract Background We investigated the interplay between complement and antibodies upon priming with single-cycle replicating viral vectors (SCIV encoding SIV antigens combined with Adeno5-SIV or SCIV pseudotyped with murine leukemia virus envelope boosting strategies. The vaccine was applied via spray-immunization to the tonsils of rhesus macaques and compared with systemic regimens. Results Independent of the application regimen or route, viral loads were significantly reduced after challenge with SIVmac239 (p Conclusion The heterologous prime-boost strategy with replication-deficient viral vectors administered exclusively via the tonsils did not induce any neutralizing antibodies before challenge. However, after challenge, comparable SIV-specific humoral immune responses were observed in all vaccinated animals. Immunization with single cycle immunodeficiency viruses mounts humoral immune responses comparable to live-attenuated immunodeficiency virus vaccines.
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.
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.
López-Lastra, M; Gabus, C; Darlix, J L
The murine leukemia virus (MLV)-related type C viruses constitute a major class of retroviruses that includes numerous endogenous and exogenous mammalian viruses and the related avian spleen necrosis virus (SNV). The MLV-related viruses possess a long and multifunctional 5' untranslated leader involved in key steps of the viral life cycle--splicing, translation, RNA dimerization, encapsidation, and reverse transcription. Recent studies have shown that the 5' leader of Friend murine leukemia virus and Moloney murine leukemia virus can direct cap independent translation of gag precursor proteins (Berlioz et al., 1995; Vagner et al., 1995b). These data, together with structural homology studies (Koning et al., 1992), prompted us to undertake a search for new internal ribosome entry segment (IRES) of retroviral origin. Here we describe an IRES element within the 5' leader of avian reticuloendotheliosis virus type A (REV-A) genomic RNA. Data show that the REV-A 5' IRES element maps downstream of the packaging/dimerization (E/DLS) sequence (Watanabe and Temin, 1982; Darlix et al., 1992) and the minimal IRES sequence appears to be within a 129 nt fragment (nucleotides 452-580) of the 5' leader, immediately upstream of the gag AUG codon. The REV-A IRES has been successfully utilized in the construction of novel high titer MLV-based retroviral vectors, containing one or more IRES elements of retroviral origin. These retroviral constructs, which represent a starting point for the design of novel vectors suitable for gene therapy, are also of interest as a model system of internal translation initiation and its possible regulation during development, cancer, or virus infection.
Carmo-Sousa, Michele; Moreno, Aranzazu; Garzo, Elisa; Fereres, Alberto
Plant viruses are known to modify the behaviour of their insect vectors, both directly and indirectly, generally adapting to each type of virus-vector relationship in a way that enhances transmission efficiency. Here, we report results of three different studies showing how a virus transmitted in a non-persistent (NP) manner (Cucumber mosaic virus; CMV, Cucumovirus) can induce changes in its host plant, cucumber (Cucumis sativus cv. Marumba) that modifies the behaviour of its aphid vector (Aphis gossypii Glover; Hemiptera: Aphididae) in a way that enhances virus transmission and spread non-viruliferous aphids changed their alighting, settling and probing behaviour activities over time when exposed to CMV-infected and mock-inoculated cucumber plants. Aphids exhibited no preference to migrate from CMV-infected to mock-inoculated plants at short time intervals (1, 10 and 30 min after release), but showed a clear shift in preference to migrate from CMV-infected to mock-inoculated plants 60 min after release. Our free-choice preference assays showed that A. gossypii alates preferred CMV-infected over mock-inoculated plants at an early stage (30 min), but this behaviour was reverted at a later stage and aphids preferred to settle and reproduce on mock-inoculated plants. The electrical penetration graph (EPG) technique revealed a sharp change in aphid probing behaviour over time when exposed to CMV-infected plants. At the beginning (first 15 min) aphid vectors dramatically increased the number of short superficial probes and intracellular punctures when exposed to CMV-infected plants. At a later stage (second hour of recording) aphids diminished their feeding on CMV-infected plants as indicated by much less time spent in phloem salivation and ingestion (E1 and E2). This particular probing behaviour including an early increase in the number of short superficial probes and intracellular punctures followed by a phloem feeding deterrence is known to enhance the transmission
Torrent, C; Gabus, C; Darlix, J L
Retroviral genomes consist of two identical RNA molecules associated at their 5' ends by the dimer linkage structure located in the packaging element (Psi or E) necessary for RNA dimerization in vitro and packaging in vivo. In murine leukemia virus (MLV)-derived vectors designed for gene transfer, the Psi + sequence of 600 nucleotides directs the packaging of recombinant RNAs into MLV virions produced by helper cells. By using in vitro RNA dimerization as a screening system, a sequence of rat VL30 RNA located next to the 5' end of the Harvey mouse sarcoma virus genome and as small as 67 nucleotides was found to form stable dimeric RNA. In addition, a purine-rich sequence located at the 5' end of this VL30 RNA seems to be critical for RNA dimerization. When this VL30 element was extended by 107 nucleotides at its 3' end and inserted into an MLV-derived vector lacking MLV Psi +, it directed the efficient encapsidation of recombinant RNAs into MLV virions. Because this VL30 packaging signal is smaller and more efficient in packaging recombinant RNAs than the MLV Psi + and does not contain gag or glyco-gag coding sequences, its use in MLV-derived vectors should render even more unlikely recombinations which could generate replication-competent viruses. Therefore, utilization of the rat VL30 packaging sequence should improve the biological safety of MLV vectors for human gene transfer.
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.
Becker, Pablo D; Hervouet, Catherine; Mason, Gavin M; Kwon, Sung-Yun; Klavinskis, Linda S
A simple dissolvable microneedle array (MA) platform has emerged as a promising technology for vaccine delivery, due to needle-free injection with a formulation that preserves the immunogenicity of live viral vectored vaccines dried in the MA matrix. While recent studies have focused largely on design parameters optimized to induce primary CD8(+) T cell responses, the hallmark of a vaccine is synonymous with engendering long-lasting memory. Here, we address the capacity of dried MA vaccination to programme phenotypic markers indicative of effector/memory CD8(+) T cell subsets and also responsiveness to recall antigen benchmarked against conventional intradermal (ID) injection. We show that despite a slightly lower frequency of dividing T cell receptor transgenic CD8(+) T cells in secondary lymphoid tissue at an early time point, the absolute number of CD8(+) T cells expressing an effector memory (CD62L(-)CD127(+)) and central memory (CD62L(+)CD127(+)) phenotype during peak expansion were comparable after MA and ID vaccination with a recombinant human adenovirus type 5 vector (AdHu5) encoding HIV-1 gag. Similarly, both vaccination routes generated CD8(+) memory T cell subsets detected in draining LNs for at least two years post-vaccination capable of responding to secondary antigen. These data suggest that CD8(+) T cell effector/memory generation and long-term memory is largely unaffected by physical differences in vaccine delivery to the skin via dried MA or ID suspension. Copyright © 2015 Elsevier Ltd. All rights reserved.
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.
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
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.
Lee, John S; Groebner, Jennifer L; Hadjipanayis, Angela G; Negley, Diane L; Schmaljohn, Alan L; Welkos, Susan L; Smith, Leonard A; Smith, Jonathan F
The development of multiagent vaccines offers the advantage of eliciting protection against multiple diseases with minimal inoculations over a shorter time span. We report here the results of using formulations of individual Venezuelan equine encephalitis (VEE) virus replicon-vectored vaccines against a bacterial disease, anthrax; a viral disease, Marburg fever; and against a toxin-mediated disease, botulism. The individual VEE replicon particles (VRP) expressed mature 83-kDa protective antigen (MAT-PA) from Bacillus anthracis, the glycoprotein (GP) from Marburg virus (MBGV), or the H(C) fragment from botulinum neurotoxin (BoNT H(C)). CBA/J mice inoculated with a mixture of VRP expressing BoNT H(C) serotype C (BoNT/C H(C)) and MAT-PA were 80% protected from a B. anthracis (Sterne strain) challenge and then 100% protected from a sequential BoNT/C challenge. Swiss mice inoculated with individual VRP or with mixtures of VRP vaccines expressing BoNT H(C) serotype A (BoNT/A H(C)), MAT-PA, and MBGV-GP produced antibody responses specific to the corresponding replicon-expressed protein. Combination of the different VRP vaccines did not diminish the antibody responses measured for Swiss mice inoculated with formulations of two or three VRP vaccines as compared to mice that received only one VRP vaccine. Swiss mice inoculated with VRP expressing BoNT/A H(C) alone or in combination with VRP expressing MAT-PA and MBGV GP, were completely protected from a BoNT/A challenge. These studies demonstrate the utility of combining individual VRP vaccines into multiagent formulations for eliciting protective immune responses to various types of diseases.
Rasmussen, Lasse Dam; Kristensen, Birgit; Kirkeby, Carsten
, 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...
Lin, Yan-Shih; Medlyn, Belinda E.; Duursma, Remko A.
, 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...
Kotsakiozi, Panayiota; Gloria-Soria, Andrea; Caccone, Adalgisa; Evans, Benjamin; Schama, Renata; Martins, Ademir Jesus; Powell, Jeffrey R
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.
Full Text Available 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.
Raj K. Singh
Full Text Available 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
Mweya, Clement N; Kimera, Sharadhuli I; Mellau, Lesakit S B; Mboera, Leonard E G
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.
Clement N. Mweya
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.
Kant, Ravi; Dasgupta, Indranil
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.
Full Text Available Stephanie L Richards, Avian V White, Jo Anne G Balanay Department of Health Education and Promotion, College of Health and Human Performance, East Carolina University, Greenville, NC, USA Abstract: Chikungunya, dengue, and Zika viruses (CHIKV, family Togaviridae, genus Alphavirus; DENV and ZIKV, family Flaviviridae, genus Flavivirus are arboviruses that cause human epidemics. Due to the lack of vaccines for many mosquito-borne diseases, there is a need for mosquito control. In the US and other regions, residual barrier insecticide sprays are applied to foliage where female mosquitoes rest and/or sugar feed between blood meals. Residual sprays are an important control method for anthropogenic day-active sylvan mosquitoes such as Aedes albopictus (vector of CHIKV, DENV, and ZIKV that are difficult to control using ultralow-volume sprays applied only at dusk or dawn when these mosquitoes are not active. In this exploratory study, we analyzed the extent to which ingestion of a sublethal dose of the active ingredient bifenthrin affected vector competence (i.e., infection, dissemination, and transmission of Ae. albopictus for DENV and ZIKV. Two incubation periods (IPs; 7 and 14 d were tested at 28°C for insecticide-fed and sugar-fed mosquitoes. We show that mosquitoes that were fed bifenthrin (0.128 µg/mL mixed with sucrose solution exhibited significantly lower DENV infection rates and body titers after a 14-d IP. During the 7-d IP, one mosquito (sugar group transmitted ZIKV. During the 14-d IP, 100% of mosquitoes showed body and leg ZIKV infections, and one mosquito (sugar+bifenthrin group transmitted ZIKV. This is a preliminary communication, and more studies will be required to further investigate these findings. We expect the findings of this small-scale study to spur larger-scale investigations of the impacts of insecticides on mechanisms regulating vector competence, and exposure to other active ingredients, and aid in development of new
Gardner, Allison M; Allan, Brian F; Frisbie, Lauren A; Muturi, Ephantus J
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
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.
Hsu, Cynthia L; Hoepting, Christine A; Fuchs, Marc; Shelton, Anthony M; Nault, Brian A
Onion thrips, Thrips tabaci (Lindeman) (Thysanoptera: Thripidae), can reduce onion bulb yield and transmit iris yellow spot virus (IYSV) (Bunyaviridae: Tospovirus), which can cause additional yield losses. In New York, onions are planted using seeds and imported transplants. IYSV is not seed transmitted, but infected transplants have been found in other U.S. states. Transplants are also larger than seeded onions early in the season, and thrips, some of which may be viruliferous, may preferentially colonize larger plants. Limited information is available on the temporal dynamics of IYSV and its vector in onion fields. In 2007 and 2008, T. tabaci and IYSV levels were monitored in six seeded and six transplanted fields. We found significantly more thrips in transplanted fields early in the season, but by the end of the season seeded fields had higher levels of IYSV. The percentage of sample sites with IYSV-infected plants remained low (fields. The densities of adult and larval thrips in August and September were better predictors of final IYSV levels than early season thrips densities. For 2007 and 2008, the time onions were harvested may have been more important in determining IYSV levels than whether the onions were seeded or transplanted. Viruliferous thrips emigrating from harvested onion fields into nonharvested ones may be increasing the primary spread of IYSV in late-harvested onions. Managing T. tabaci populations before harvest, and manipulating the spatial arrangement of fields based on harvest date could mitigate the spread of IYSV.
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.
Muturi, Ephantus J; Ramirez, Jose L; Zilkowski, Bruce; Flor-Weiler, Lina B; Rooney, Alejandro P
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. PMID:29718505
Clement Nyamunura Mweya
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.
Mweya, Clement Nyamunura; Kimera, Sharadhuli Iddi; Kija, John Bukombe; Mboera, Leonard E G
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.
Hajeri, Subhas; Killiny, Nabil; El-Mohtar, Choaa; Dawson, William O; Gowda, Siddarame
A transient expression vector based on Citrus tristeza virus (CTV) is unusually stable. Because of its stability it is being considered for use in the field to control Huanglongbing (HLB), which is caused by Candidatus Liberibacter asiaticus (CLas) and vectored by Asian citrus psyllid, Diaphorina citri. In the absence of effective control strategies for CLas, emphasis has been on control of D. citri. Coincident cohabitation in phloem tissue by CLas, D. citri and CTV was exploited to develop a novel method to mitigate HLB through RNA interference (RNAi). Since CTV has three RNA silencing suppressors, it was not known if CTV-based vector could induce RNAi in citrus. Yet, expression of sequences targeting citrus phytoene desaturase gene by CTV-RNAi resulted in photo-bleaching phenotype. CTV-RNAi vector, engineered with truncated abnormal wing disc (Awd) gene of D. citri, induced altered Awd expression when silencing triggers ingested by feeding D. citri nymphs. Decreased Awd in nymphs resulted in malformed-wing phenotype in adults and increased adult mortality. This impaired ability of D. citri to fly would potentially limit the successful vectoring of CLas bacteria between citrus trees in the grove. CTV-RNAi vector would be relevant for fast-track screening of candidate sequences for RNAi-mediated pest control. Copyright © 2014. Published by Elsevier B.V.
Coke, Rob L; Backues, Kay A; Hoover, John P; Saliki, Jeremiah T; Ritchey, Jerry W; West, Gary D
Fennec foxes (Vulpes zerda) and meerkats (Suricata suricatta) are considered to be susceptible to canine distemper virus (CDV) infection. Although no definitive clinical cases of natural CDV infections have been reported, mortalities due to CDV have been suspected and are reported in other closely related species. A commercially available monovalent, live, canarypox-vectored CDV vaccine induced neutralizing antibody titers that were maintained for at least a year in both fennec foxes and meerkats.
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.
Tsai, Cheng-Hui; Chen, Tien-Huang; Lin, Cheo; Shu, Pei-Yun; Su, Chien-Ling; Teng, Hwa-Jen
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.
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
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
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.
Noris, Emanuela; Miozzi, Laura
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.
Lokhandwala, Shehnaz; Waghela, Suryakant D; Bray, Jocelyn; Martin, Cameron L; Sangewar, Neha; Charendoff, Chloe; Shetti, Rashmi; Ashley, Clay; Chen, Chang-Hsin; Berghman, Luc R; Mwangi, Duncan; Dominowski, Paul J; Foss, Dennis L; Rai, Sharath; Vora, Shaunak; Gabbert, Lindsay; Burrage, Thomas G; Brake, David; Neilan, John; Mwangi, Waithaka
The African swine fever virus (ASFV) causes a fatal hemorrhagic disease in domestic swine, and at present no treatment or vaccine is available. Natural and gene-deleted, live attenuated strains protect against closely related virulent strains; however, they are yet to be deployed and evaluated in the field to rule out chronic persistence and a potential for reversion to virulence. Previous studies suggest that antibodies play a role in protection, but induction of cytotoxic T lymphocytes (CTLs) could be the key to complete protection. Hence, generation of an efficacious subunit vaccine depends on identification of CTL targets along with a suitable delivery method that will elicit effector CTLs capable of eliminating ASFV-infected host cells and confer long-term protection. To this end, we evaluated the safety and immunogenicity of an adenovirus-vectored ASFV (Ad-ASFV) multiantigen cocktail formulated in two different adjuvants and at two immunizing doses in swine. Immunization with the cocktail rapidly induced unprecedented ASFV antigen-specific antibody and cellular immune responses against all of the antigens. The robust antibody responses underwent rapid isotype switching within 1 week postpriming, steadily increased over a 2-month period, and underwent rapid recall upon boost. Importantly, the primed antibodies strongly recognized the parental ASFV (Georgia 2007/1) by indirect fluorescence antibody (IFA) assay and Western blotting. Significant antigen-specific gamma interferon-positive (IFN-γ + ) responses were detected postpriming and postboosting. Furthermore, this study is the first to demonstrate induction of ASFV antigen-specific CTL responses in commercial swine using Ad-ASFV multiantigens. The relevance of the induced immune responses in regard to protection needs to be evaluated in a challenge study. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
Segoli, Michal; Hoffmann, Ary A; Lloyd, Jane; Omodei, Gavin J; Ritchie, Scott A
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.
Murugan, Kadarkarai; Wei, Jiang; Alsalhi, Mohamad Saleh; Nicoletti, Marcello; Paulpandi, Manickam; Samidoss, Christina Mary; Dinesh, Devakumar; Chandramohan, Balamurugan; Paneerselvam, Chellasamy; Subramaniam, Jayapal; Vadivalagan, Chithravel; Wei, Hui; Amuthavalli, Pandiyan; Jaganathan, Anitha; Devanesan, Sandhanasamy; Higuchi, Akon; Kumar, Suresh; Aziz, Al Thabiani; Nataraj, Devaraj; Vaseeharan, Baskaralingam; Canale, Angelo; Benelli, Giovanni
A main challenge in parasitology is the development of reliable tools to prevent or treat mosquito-borne diseases. We investigated the toxicity of magnetic nanoparticles (MNP) produced by Magnetospirillum gryphiswaldense (strain MSR-1) on chloroquine-resistant (CQ-r) and sensitive (CQ-s) Plasmodium falciparum, dengue virus (DEN-2), and two of their main vectors, Anopheles stephensi and Aedes aegypti, respectively. MNP were studied by Fourier-transform infrared spectroscopy and transmission electron microscopy. They were toxic to larvae and pupae of An. stephensi, LC 50 ranged from 2.563 ppm (1st instar larva) to 6.430 ppm (pupa), and Ae. aegypti, LC 50 ranged from 3.231 ppm (1st instar larva) to 7.545 ppm (pupa). MNP IC 50 on P. falciparum were 83.32 μg ml -1 (CQ-s) and 87.47 μg ml -1 (CQ-r). However, the in vivo efficacy of MNP on Plasmodium berghei was low if compared to CQ-based treatments. Moderate cytotoxicity was detected on Vero cells post-treatment with MNP doses lower than 4 μg ml -1 . MNP evaluated at 2-8 μg ml -1 inhibited DEN-2 replication inhibiting the expression of the envelope (E) protein. In conclusion, our findings represent the first report about the use of MNP in medical and veterinary entomology, proposing them as suitable materials to develop reliable tools to combat mosquito-borne diseases.
Kröber, Wenzel; Plath, Isa; Heklau, Heike; Bruelheide, Helge
Leaf functional traits are important because they reflect physiological functions, such as transpiration and carbon assimilation. In particular, morphological leaf traits have the potential to summarize plants strategies in terms of water use efficiency, growth pattern and nutrient use. The leaf economics spectrum (LES) is a recognized framework in functional plant ecology and reflects a gradient of increasing specific leaf area (SLA), leaf nitrogen, phosphorus and cation content, and decreasing leaf dry matter content (LDMC) and carbon nitrogen ratio (CN). The LES describes different strategies ranging from that of short-lived leaves with high photosynthetic capacity per leaf mass to long-lived leaves with low mass-based carbon assimilation rates. However, traits that are not included in the LES might provide additional information on the species' physiology, such as those related to stomatal control. Protocols are presented for a wide range of leaf functional traits, including traits of the LES, but also traits that are independent of the LES. In particular, a new method is introduced that relates the plants' regulatory behavior in stomatal conductance to vapor pressure deficit. The resulting parameters of stomatal regulation can then be compared to the LES and other plant functional traits. The results show that functional leaf traits of the LES were also valid predictors for the parameters of stomatal regulation. For example, leaf carbon concentration was positively related to the vapor pressure deficit (vpd) at the point of inflection and the maximum of the conductance-vpd curve. However, traits that are not included in the LES added information in explaining parameters of stomatal control: the vpd at the point of inflection of the conductance-vpd curve was lower for species with higher stomatal density and higher stomatal index. Overall, stomata and vein traits were more powerful predictors for explaining stomatal regulation than traits used in the LES.
Introduction of a prosthesis into the oral cavity results in profound alterations of the environmental conditions as the prosthesis and the underlying mucosa become colonized with oral microorganisms, including Candida spp. This may lead to denture stomatitis, a non-specific inflammatory reaction against microbial antigens, toxins and enzymes produced by the colonizing microorganisms. The role of Candida in the etiology of denture stomatitis is indicated by an increased number of yeasts on th...
Göertz, Giel P.; Vogels, Chantal B.F.; Geertsema, Corinne; Koenraadt, Constantianus J.M.; Pijlman, Gorben P.
Background: Zika virus (ZIKV) and chikungunya virus (CHIKV) are highly pathogenic arthropod-borne viruses that are currently a serious health burden in the Americas, and elsewhere in the world. ZIKV and CHIKV co-circulate in the same geographical regions and are mainly transmitted by Aedes
Mardulyn, P.; Goffredo, M.; Conte, A.; Hendrickx, G.; Meiswinkel, R.; Balenghien, T.; Sghaier, S.; Lohr, Y.; Gilbert, M.
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
Morin, Cory; Monaghan, Andrew; Quattrochi, Dale; Crosson, William; Hayden, Mary; Ernst, Kacey
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.
Mikkelsen, J G; Lund, Anders Henrik; Duch, M
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-imp....... 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.......-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...... harbouring the supposed RNA dimer-forming cis -elements and (ii) that copackaged retroviral RNAs can recombine despite pronounced sequence dissimilarity at the cross-over site(s) and within parts of the genome involved in RNA dimerization, encapsidation and strand transferring during reverse transcription...
Kate McElroy Horne
Full Text Available The Bunyaviridae family is comprised of more than 350 viruses, of which many within the Hantavirus, Orthobunyavirus, Nairovirus, Tospovirus, and Phlebovirus genera are significant human or agricultural pathogens. The viruses within the Orthobunyavirus, Nairovirus, and Phlebovirus genera are transmitted by hematophagous arthropods, such as mosquitoes, midges, flies, and ticks, and their associated arthropods not only serve as vectors but also as virus reservoirs in many cases. This review presents an overview of several important emerging or re-emerging bunyaviruses and describes what is known about bunyavirus-vector interactions based on epidemiological, ultrastructural, and genetic studies of members of this virus family.
McAdam, Scott A M; Brodribb, Timothy J
Motivated by studies suggesting that the stomata of ferns and lycophytes do not conform to the standard active abscisic acid (ABA) -mediated stomatal control model, we examined stomatal behavior in a conifer species (Metasequoia glyptostroboides) that is phylogenetically midway between the fern and angiosperm clades. Similar to ferns, daytime stomatal closure in response to moderate water stress seemed to be a passive hydraulic process in M. glyptostroboides immediately alleviated by rehydrating excised shoots. Only after prolonged exposure to more extreme water stress did active ABA-mediated stomatal closure become important, because foliar ABA production was triggered after leaf turgor loss. The influence of foliar ABA on stomatal conductance and stomatal aperture was highly predictable and additive with the passive hydraulic influence. M. glyptostroboides thus occupies a stomatal behavior type intermediate between the passively controlled ferns and the characteristic ABA-dependent stomatal closure described in angiosperm herbs. These results highlight the importance of considering phylogeny as a major determinant of stomatal behavior.
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.
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
Xu, Hongxing; He, Xiaochan; Zheng, Xusong; Yang, Yajun; Tian, Junce; Lu, Zhongxian
Southern rice black-streaked dwarf virus (SRBSDV) is a recently discovered member of the genus Fijivirus and it is transmitted by the rice whitebacked planthopper (WBPH), Sogatella furcifera (Horváth). It was found that SRBSDV infected vectors might contribute negatively to the WBPH population, although the longer nymphal period might benefit viral acquisition, transmission and increase infection rate. The interaction between SRBSDV and its vector need to be further explored to gain better understanding of the dispersal of WBPH and the spread of virus disease, in particular the feeding and reproduction behavior of viruliferous WBPH. Newly hatched nymphs of WBPH were fed on healthy rice plant after feeding on SRBSDV-infected rice plants for 2 h, and newly emerged adults were numbered and tested. Feeding behaviors of WBPH adults were monitored electronically within a Faraday cage using a Giga-4 DC EPG amplifier. The newly emerged adults were paired, and the fecundity and egg hatchability were investigated. WBPH was molecularly identified for SRBSDV when they dead. According to the identification results, data on viruliferous and non-viruliferous WBPH were collected and analyzed. Feeding behavior of viruliferous WBPH was different from those of non-viruliferous WBPH. Frequency of phloem sap ingestion of viruliferous WBPH increased significantly, however the total feeding duration did not increase markedly. When both WBPH parents were infected with SRBSDV, their fecundity and hatchability of the eggs produced were significant lower than those of normal WBPH parents. However, if only one of the parents was viruliferous, fecundity and egg hatchability were only slightly affected. Viruliferous WBPH fed on the phloem more frequently than non-viruliferous WBPH and can thus contribute to virus transmission. When both vector parents are viruliferous fecundity and hatchability of the eggs were significantly reduced. However when only one of the parents WBPH was viruliferous
Hernandez Alfaro, Myriam
Tagosodes orizicolus (Homoptera: Delphacidae), is a monophagous insect of the rice and virus vector of the white leaf (RHBV). It is distributed in America Central, El Caribe, part of the America del Sur and in Costa Rica it is in all the producing zones of rice. The genetic variability was analyzed by means of RAPD-PCR of individuals from three populations of Costa Rica: Parrita, Guanacaste (Liberia) y San Carlos (Santa Clara), that they are found separated geographically. The technique consisted of amplifying regions at random of the genome of these insects utilizing five primers. A total of 72 polymorphic bands were obtained, that upon being analyzed statistically by means of the multivariate analysis program of numerical taxonomy could show a clear genetic distancing among said populations. The genetic distance observed in the molecular analysis can be explained for the climatic and/or geographical isolation of the populations or by the incident of Wolbachia, riquettsia that induces cytoplasmic sterility in insects. These symbionts are transmitted of generation in generation, of the mother to their offspring and they cause reproductive alterations as cytoplasmic incompatibility, parthenogenesis and feminization. The presence of Wolbachia was determined by means of transmission electronic microscopy being observed in the greasy and muscular weave of the abdomen of T. orizicolus. Rickettsias present pleomorphic morphology and form small groups, that are characterized for the presence of electrondense material semidetached to the cell wall with an interior electronlucent. Its size in transverse cuts ranged between 520 nm of length X 470 nm of width. In addition, its presence was detected by means of the amplification by PCR of the genomic DNA of the insects; a specific primer for the DNA ribosomal 16S of Wolbachia was utilized for it. A 86% of insects of the San Carlos population were positive, a 96% was determined for Guanacaste, a 37% for Parrita and a 100% for
Full Text Available With the aim to develop an efficient and cost-effective approach to control malaria, we have generated porcine parvovirus-like particles (PPV-VLPs carrying the CD8(+ T cell epitope (SYVPSAEQI of the circumsporozoite (CS protein from Plasmodium yoelii fused to the PPV VP2 capsid protein (PPV-PYCS, and tested in prime/boost protocols with poxvirus vectors for efficacy in a rodent malaria model. As a proof-of concept, we have characterized the anti-CS CD8(+ T cell response elicited by these hybrid PPV-VLPs in BALB/c mice after immunizations with the protein PPV-PYCS administered alone or in combination with recombinant vaccinia virus (VACV vectors from the Western Reserve (WR and modified virus Ankara (MVA strains expressing the entire P. yoelii CS protein. The results of different immunization protocols showed that the combination of PPV-PYCS prime/poxvirus boost was highly immunogenic, inducing specific CD8+ T cell responses to CS resulting in 95% reduction in liver stage parasites two days following sporozoite challenge. In contrast, neither the administration of PPV-PYCS alone nor the immunization with the vectors given in the order poxvirus/VLPs was as effective. The immune profile induced by VLPs/MVA boost was associated with polyfunctional and effector memory CD8+ T cell responses. These findings highlight the use of recombinant parvovirus PPV-PYCS particles as priming agents and poxvirus vectors, like MVA, as booster to enhance specific CD8+ T cell responses to Plasmodium antigens and to control infection. These observations are relevant in the design of T cell-inducing vaccines against malaria.
Rodríguez, Dolores; González-Aseguinolaza, Gloria; Rodríguez, Juan R; Vijayan, Aneesh; Gherardi, Magdalena; Rueda, Paloma; Casal, J Ignacio; Esteban, Mariano
With the aim to develop an efficient and cost-effective approach to control malaria, we have generated porcine parvovirus-like particles (PPV-VLPs) carrying the CD8(+) T cell epitope (SYVPSAEQI) of the circumsporozoite (CS) protein from Plasmodium yoelii fused to the PPV VP2 capsid protein (PPV-PYCS), and tested in prime/boost protocols with poxvirus vectors for efficacy in a rodent malaria model. As a proof-of concept, we have characterized the anti-CS CD8(+) T cell response elicited by these hybrid PPV-VLPs in BALB/c mice after immunizations with the protein PPV-PYCS administered alone or in combination with recombinant vaccinia virus (VACV) vectors from the Western Reserve (WR) and modified virus Ankara (MVA) strains expressing the entire P. yoelii CS protein. The results of different immunization protocols showed that the combination of PPV-PYCS prime/poxvirus boost was highly immunogenic, inducing specific CD8+ T cell responses to CS resulting in 95% reduction in liver stage parasites two days following sporozoite challenge. In contrast, neither the administration of PPV-PYCS alone nor the immunization with the vectors given in the order poxvirus/VLPs was as effective. The immune profile induced by VLPs/MVA boost was associated with polyfunctional and effector memory CD8+ T cell responses. These findings highlight the use of recombinant parvovirus PPV-PYCS particles as priming agents and poxvirus vectors, like MVA, as booster to enhance specific CD8+ T cell responses to Plasmodium antigens and to control infection. These observations are relevant in the design of T cell-inducing vaccines against malaria.
Heeney, Jonathan L.; Mooij, Petra; Nájera García, José Luis; Jiménez, Victoria; Esteban, Mariano; Gómez, Carmen E.
Vectores Recombinantes basados en el Virus Modificado de Ankara (MVA) como Vacunas Preventivas y Terapéuticas contra el SIDA. Losvirus recombinantes de la invención contienen secuencias que se encuentran insertadas en el mismo sitio de inserción del MVA y permiten la expresión simultánea de varios antígenos, una proteína Env del VIH-I consistente en una proteína gpl20 carente de secuencias correspondientes a la proteína gp41, y una proteína quiméricade fusión de Gag, Pol y Nef. Son virus esta...
Jespersen, T.; Duch, M.; Carrasco, M.L.
of spliced env mRNA for the SL3-3 derived vector relative to the Akv derived vectors, seemingly contributing to its low replication capacity. The EGFP expressing Akv-MLV was genetically stable for multiple rounds of infection; marker-cassette deletion revertants appeared after several replication rounds...
Grosse, Stefanie; Penaud-Budloo, Magalie; Herrmann, Anne-Kathrin; Börner, Kathleen; Fakhiri, Julia; Laketa, Vibor; Krämer, Chiara; Wiedtke, Ellen; Gunkel, Manuel; Ménard, Lucie; Ayuso, Eduard; Grimm, Dirk
The discovery that adeno-associated virus 2 (AAV2) encodes an eighth protein, called assembly-activating protein (AAP), transformed our understanding of wild-type AAV biology. Concurrently, it raised questions about the role of AAP during production of recombinant vectors based on natural or molecularly engineered AAV capsids. Here, we show that AAP is indeed essential for generation of functional recombinant AAV2 vectors in both mammalian and insect cell-based vector production systems. Surprisingly, we observed that AAV2 capsid proteins VP1 to -3 are unstable in the absence of AAP2, likely due to rapid proteasomal degradation. Inhibition of the proteasome led to an increase of intracellular VP1 to -3 but neither triggered assembly of functional capsids nor promoted nuclear localization of the capsid proteins. Together, this underscores the crucial and unique role of AAP in the AAV life cycle, where it rapidly chaperones capsid assembly, thus preventing degradation of free capsid proteins. An expanded analysis comprising nine alternative AAV serotypes (1, 3 to 9, and rh10) showed that vector production always depends on the presence of AAP, with the exceptions of AAV4 and AAV5, which exhibited AAP-independent, albeit low-level, particle assembly. Interestingly, AAPs from all 10 serotypes could cross-complement AAP-depleted helper plasmids during vector production, despite there being distinct intracellular AAP localization patterns. These were most pronounced for AAP4 and AAP5, congruent with their inability to rescue an AAV2/AAP2 knockout. We conclude that AAP is key for assembly of genuine capsids from at least 10 different AAV serotypes, which has implications for vectors derived from wild-type or synthetic AAV capsids. IMPORTANCE Assembly of adeno-associated virus 2 (AAV2) is regulated by the assembly-activating protein (AAP), whose open reading frame overlaps with that of the viral capsid proteins. As the majority of evidence was obtained using virus
Fever versus Fever: the role of host and vector susceptibility and interspecific competition in shaping the current and future distributions of the sylvatic cycles of dengue virus and yellow fever virus
Hanley, Kathryn A.; Monath, Thomas P.; Weaver, Scott C.; Rossi, Shannan L.; Richman, Rebecca L.; Vasilakis, Nikos
Two different species of flaviviruses, dengue virus (DENV) and yellow fever virus (YFV), that originated in sylvatic cycles maintained in non-human primates and forest-dwelling mosquitoes have emerged repeatedly into sustained human-to-human transmission by Aedes aegypti mosquitoes. Sylvatic cycles of both viruses remain active, and where the two viruses overlap in West Africa they utilize similar suites of monkeys and Aedes mosquitoes. These extensive similarities render the differences in the biogeography and epidemiology of the two viruses all the more striking. First, the sylvatic cycle of YFV originated in Africa and was introduced into the New World, probably as a result of the slave trade, but is absent in Asia; in contrast, sylvatic DENV likely originated in Asia and has spread to Africa but not to the New World. Second, while sylvatic YFV can emerge into extensive urban outbreaks in humans, these invariably die out, whereas four different types of DENV have established human transmission cycles that are ecologically and evolutionarily distinct from their sylvatic ancestors. Finally, transmission of YFV among humans has been documented only in Africa and the Americas, whereas DENV is transmitted among humans across most of the range of competent Aedes vectors, which in the last decade has included every continent save Antarctica. This review summarizes current understanding of sylvatic transmission cycles of YFV and DENV, considers possible explanations for their disjunct distributions, and speculates on the potential consequences of future establishment of a sylvatic cycle of DENV in the Americas. PMID:23523817
Alternanthera mosaic virus (AltMV; genus Potexvirus) is distinguished from the type member of the genus, Potato virus X by features of viral movement and variation within triple gene block protein 1 (TGB1). AltMV TGB1 variants TGB1L88 and TGB1P88 confer strong and weak silencing suppression, respect...
Japanese Encephalitis virus (JEV) is a mosquito-borne arbovirus that causes endemic and epidemic encephalitis in Eastern and Southeastern Asia. Swine and wading birds serve as reservoirs for the virus, which can be transmitted to humans via mosquitos. Currently, there is no specific treatment availa...
Sullivan, Nancy J.; Geisbert, Thomas W.; Geisbert, Joan B.; Shedlock, Devon J.; Xu, Ling; Lamoreaux, Laurie; Custers, Jerome H. H. V.; Popernack, Paul M.; Yang, Zhi-Yong; Pau, Maria G.; Roederer, Mario; Koup, Richard A.; Goudsmit, Jaap; Jahrling, Peter B.; Nabel, Gary J.
BACKGROUND: Ebola virus causes a hemorrhagic fever syndrome that is associated with high mortality in humans. In the absence of effective therapies for Ebola virus infection, the development of a vaccine becomes an important strategy to contain outbreaks. Immunization with DNA and/or
Full Text Available Persistent viral vector-mediated transgene expression in the airways requires delivery to cells with progenitor capacity and avoidance of immune responses. Previously, we observed that GP64-pseudotyped feline immunodeficiency virus (FIV-mediated gene transfer was more efficient in the nasal airways than the large airways of the murine lung. We hypothesized that in vivo gene transfer was limited by immunological and physiological barriers in the murine intrapulmonary airways. Here, we systematically investigate multiple potential barriers to lentiviral gene transfer in the airways of mice. We show that GP64-FIV vector transduced primary cultures of well-differentiated murine nasal epithelia with greater efficiency than primary cultures of murine tracheal epithelia. We further demonstrate that neutrophils, type I interferon (IFN responses, as well as T and B lymphocytes are not the major factors limiting the transduction of murine conducting airways. In addition, we observed better transduction of GP64-pseudotyped vesicular stomatitis virus (VSV in the nasal epithelia compared with the intrapulmonary airways in mice. VSVG glycoprotein pseudotyped VSV transduced intrapulmonary epithelia with similar efficiency as nasal epithelia. Our results suggest that the differential transduction efficiency of nasal versus intrapulmonary airways by FIV vector is not a result of immunological barriers or surface area, but rather differential expression of cellular factors specific for FIV vector transduction.
Tarek S. El-Madany
Full Text Available Ozone is a key trace gas in the troposphere; because it is a greenhouse gas, it is very reactive, and it is potentially toxic to humans, fauna, and vegetation. The main sink processes for ozone are chemical reactions and the turbulent deposition flux to the earth’s surface. The deposition process itself is rather complex: The interactions between co-varying drivers such as the tropospheric ozone concentration, turbulence, and chemical reactions are not well understood. In the case of ozone deposition to vegetation, another aspect that must be studied is the role of stomatal regulation for a wide range of conditions. Therefore, we measured turbulent deposition fluxes of ozone with the eddy covariance technique during the peak of the growing season in 2014 over a managed, rewetted peatland in NW Germany. The deposition flux was large during the day (up to −15 nmol m−2 s−1 and relatively small during the night (between −1 and −2 nmol m−2 s−1. Flux partitioning by applying the surface resistance analogy and further analysis showed that the stomatal uptake was smaller than non-stomatal deposition. The correction of stomatal conductance with the gross primary production (GPP improved the estimation of day- and nighttime stomatal deposition fluxes. Statistical analysis confirmed that the friction velocity (u* was the single most important driver of non-stomatal ozone deposition and that relationships with other environmental drivers are not linear and highly variable. Further research is needed to develop a better process understanding of non-stomatal ozone deposition, to quantify the role of surface deposition to the ozone budget of the atmospheric boundary layer, and to estimate uncertainties associated with the partitioning of ozone deposition into stomatal and non-stomatal fluxes.
Marcela Cristina Correa de Freitas
Full Text Available OBJECTIVE: Nowadays recombinant factor VIII is produced in murine cells including in Chinese hamster ovary (CHO and baby hamster kidney cells (BHK. Previous studies, using the murine leukemia virus-derived retroviral vector pMFG-FVIII-P140K, modified two recombinant human cell lines, HepG2 and Hek293 to produce recombinant factor VIII. In order to characterize these cells, the present study aimed to analyze the integration pattern of retroviral vector pMFG-FVIII-P140K.METHODS: This study used ligation-mediated polymerase chain reaction to locate the site of viral vector integration by sequencing polymerase chain reaction products. The sequences were compared to genomic databases to characterize respective clones.RESULTS: The retroviral vector presented different and non-random profiles of integration between cells lines. A preference of integration for chromosomes 19, 17 and 11 was observed for HepG2FVIIIdB/P140K and chromosome 9 for Hek293FVIIIdB/P140K. In genomic regions such as CpG islands and transcription factor binding sites, there was no difference in the integration profiles for both cell lines. Integration in intronic regions of encoding protein genes (RefSeq genes was also observed in both cell lines. Twenty percent of integrations occurred at fragile sites in the genome of the HepG2 cell line and 17% in Hek293.CONCLUSION: The results suggest that the cell type can affect the profile of chromosomal integration of the retroviral vector used; these differences may interfere in the level of expression of recombinant proteins.
Rapid diagnosis of vesicular stomatitis virus in Ecuador by the use of polymerase chain reaction Diagnóstico rápido do vírus da estomatite vesicular no Equador mediante o uso da reação em cadeia da polimerase
Lya Madureira Sepúlveda
Full Text Available Vesicular Stomatitis (VS is a viral disease that has a great impact in animal health, as infected animals present marked decrease in meat and milk production. Its presence is a limiting factor for international animal trade. Besides the damage in the livestock productivity, such disease assumes an important role in animal health programs since it is clinically indistinguishable from Foot-and-Mouth Disease. The diagnosis of the VS has been made, mainly, through Complement Fixation, ELISA and Virus Neutralization tests, assays that allow not only for viral detection but also for differentiation of the two serotypes described for Vesicular Stomatitis Virus (VSV: New Jersey (NJ and Indiana (Ind. In this work, a molecular diagnostic approach, the polymerase chain reaction performed after reverse transcription (RT - PCR, based on the specific partial amplification of NS gene of VSV was used, as an alternative method for the detection of the virus. A total of 10 VSV reference samples and 12 specimens collected from animals with clinical signs of vesicular disease obtained from field episodes in Ecuador were tested. The method allowed for the specific partial amplification of the region coding for protein P, both for VSV serotypes New Jersey (642 bp and Indiana 1 (614 bp. The results were compatible with data obtained by Complement Fixation test and the identity of the amplified products was confirmed by nucleotide sequencing.A Estomatite Vesicular (EV é uma enfermidade viral de grande impacto na saúde animal. O animal enfermo apresenta queda na produtividade em rebanho de carne e na produção leiteira, sendo um fator limitante para o comércio internacional de animais. Além dos danos à produtividade essa enfermidade assume importante papel nos programas de saúde animal por ser indistinguível clinicamente da Febre Aftosa. As técnicas para o diagnóstico da EV são, principalmente, a Fixação de Complemento, a ELISA e a Virusneutraliza
Cottingham, Matthew G; van Maurik, Andre; Zago, Manola; Newton, Angela T; Anderson, Richard J; Howard, M Keith; Schneider, Jörg; Skinner, Michael A
The FP9 strain of F has been described as a more immunogenic recombinant vaccine vector than the Webster FPV-M (FPW) strain (R. J. Anderson et al., J. Immunol. 172:3094-3100, 2004). This study expands the comparison to include two separate recombinant antigens and multiple, rather than single, independent viral clones derived from the two strains. Dual-poxvirus heterologous prime-boost vaccination regimens using individual clones of recombinant FP9 or FPW in combination with recombinant modified V Ankara expressing the same antigen were evaluated for their ability to elicit T-cell responses against recombinant antigens from Plasmodium berghei (circumsporozoite protein) or human immunodeficiency virus type 1 (a Gag-Pol-Nef fusion protein). Gamma interferon enzyme-linked immunospot assay and fluorescence-activated cell sorting assays of the responses to specific epitopes confirmed the approximately twofold-greater cellular immunogenicity of FP9 compared to FPW, when given as the priming or boosting immunization. Equality of transgene expression in mouse cells infected with the two strains in vitro was verified by Western blotting. Directed partial sequence analysis and PCR analysis of FPW and comparison to available whole-genome sequences revealed that many loci that are mutated in the highly attenuated and culture-adapted FP9 strain are wild type in FPW, including the seven multikilobase deletions. These "passage-specific" alterations are hypothesized to be involved in determining the immunogenicity of fowlpox virus as a recombinant vaccine vector.
STOMATAL REGULATION AND ROOT GROWTH ... maintenance of high plant water potential in common bean under stress was the function of stomatal regulation and/or root ... disadvantage since it will reduce CO2 fixation and hence may ...
Ghobadloo, Shahrokh M.; Balcerzak, Anna K.; Gargaun, Ana; Muharemagic, Darija; Mironov, Gleb G.; Capicciotti, Chantelle J.; Briard, Jennie G.; Ben, Robert N.; Berezovski, Maxim V.
The inability of vaccines to retain sufficient thermostability has been an obstacle to global vaccination programs. To address this major limitation, we utilized carbohydrate-based ice recrystallization inhibitors (IRIs) to eliminate the cold chain and stabilize the potency of Vaccinia virus (VV), Vesicular Stomatitis virus (VSV) and Herpes virus-1 (HSV-1). The impact of these IRIs was tested on the potency of the viral vectors using a plaque forming unit assay following room temperature storage, cryopreservation with successive freeze-thaw cycles and lyophilization. Viral potency after storage with all three conditions demonstrated that N-octyl-gluconamide (NOGlc) recovered the infectivity of shelf stored VV, 5.6 Log10 PFU mL-1 during 40 days, and HSV-1, 2.7 Log10 PFU mL-1 during 9 days. Carbon-linked antifreeze glycoprotein analogue ornithine-glycine-glycine-galactose (OGG-Gal) increases the recovery of VV and VSV more than 1 Log10 PFU mL-1 after 10 freeze-thaw cycles. In VSV, cryostorage with OGG-Gal maintains high infectivity and reduces temperature-induced aggregation of viral particles by 2 times that of the control. In total, OGG-Gal and NOGlc preserve virus potency during cryostorage. Remarkably, NOGlc has potential to eliminate the cold chain and permit room temperature storage of viral vectors.
Jiao, Liya; Wang, Lihong; Zhou, Qing; Huang, Xiaohua
Bisphenol A (BPA) is an emerging environmental endocrine disruptor that has toxic effects on plants growth. Photosynthesis supplies the substances and energy required for plant growth, and regulated by stomatal and non-stomatal factors. Therefore, in this study, to reveal how BPA affects photosynthesis in soybean seedlings (Glycine max L.) from the perspective of stomatal and non-stomatal factors, the stomatal factors (stomatal conductance and behaviours) and non-stomatal factors (Hill reaction, apparent quantum efficiency, Rubisco activity, carboxylation efficiency, the maximum Rubisco carboxylation velocity, ribulose-1,5-bisphospate regeneration capacities mediated by maximum electron transport rates, and triose phosphate utilization rate) were investigated using a portable photosynthesis system. Moreover, the pollution of BPA in the environment was simulated. The results indicate that low-dose BPA enhanced net photosynthetic rate (P n ) primarily by promoting stomatal factors, resulting in increased relative growth rates and accelerated soybean seedling growth. High-dose BPA decreases the P n by simultaneously inhibiting stomatal and non-stomatal factors, and this inhibition decreases the relative growth rates further reducing soybean seedling growth. Following the withdrawal of BPA, all of the indices were restored to varying degrees. In conclusion, low-dose BPA increased the P n by promoting stomatal factors while high-dose BPA decreased the P n by simultaneously inhibiting stomatal and non-stomatal factors. These findings provide a model (or, hypothesis) for the effects of BPA on plant photosynthesis. Copyright © 2017 Elsevier Inc. All rights reserved.
Walton, T E; Webb, P A; Kramer, W L; Smith, G C; Davis, T; Holbrook, F R; Moore, C G; Schiefer, T J; Jones, R H; Janney, G C
An epizootic of vesicular stomatitis (VS) caused by the New Jersey serotype of VS virus affected livestock and humans in 14 western states in 1982-1983. Epidemiological observations were made on at least 10% of the cattle in 4 dairy herds that were located in the vicinity of Grand Junction, Colorado. High rates of neutralizing antibody to the New Jersey serotype were seen in all cattle regardless of whether livestock in the dairy had clinical VS or a decrease in mild production. Antibody titers remained high in these cattle for as long as 2 years after the epizootic. No virus isolations were made from 32 humans with clinical signs compatible with viral disease. Entomological information was obtained during the epizootic from 23 premises in northwestern Colorado. Insect collections yielded 4 isolates from Culicoides spp. midges, 2 from C. variipennis, and 1 each from C. stellifer and C. (Selfia) spp. This is the first report of VS virus isolations from field-collected Culicoides.
Kanao, Megumi; Kanda, Hirotsugu; Huang, Wan; Liu, Shue; Yi, Hyun; Candiotti, Keith A; Lubarsky, David A; Levitt, Roy C; Hao, Shuanglin
Human immunodeficiency virus (HIV)-related painful sensory neuropathies primarily consist of the HIV infection-related distal sensory polyneuropathy and antiretroviral toxic neuropathies. Pharmacotherapy provides only partial relief of pain in patients with HIV/acquired immune deficiency syndrome because little is known about the exact neuropathological mechanisms for HIV-associated neuropathic pain (NP). Hypofunction of γ-aminobutyric acid (GABA) GABAergic inhibitory mechanisms has been reported after peripheral nerve injury. In this study, we tested the hypothesis that HIV gp120 combined with antiretroviral therapy reduces spinal GABAergic inhibitory tone and that restoration of GABAergic inhibitory tone will reduce HIV-related NP in a rat model. The application of recombinant HIV-1 envelope protein gp120 into the sciatic nerve plus systemic ddC (one antiretroviral drug) induced mechanical allodynia. The hind paws of rats were inoculated with replication-defective herpes simplex virus (HSV) vectors genetically encoding gad1 gene to express glutamic acid decarboxylase 67 (GAD67), an enzyme that catalyzes the decarboxylation of glutamate to GABA. Mechanical threshold was tested using von Frey filaments before and after treatments with the vectors. The expression of GAD67 in both the lumbar spinal cord and the L4-5 dorsal root ganglia was examined using western blots. The expression of mitochondrial superoxide in the spinal dorsal horn was examined using MitoSox imaging. The immunoreactivity of spinal GABA, pCREB, and pC/EBPβ was tested using immunohistochemistry. In the gp120 with ddC-induced neuropathic pain model, GAD67 expression mediated by the HSV vector caused an elevation of mechanical threshold that was apparent on day 3 after vector inoculation. The antiallodynic effect of the single HSV vector inoculation expressing GAD67 lasted >28 days. The area under the time-effect curves in the HSV vector expressing GAD67 was increased compared with that in the
Zheng, Wenwen; Huang, Wan; Liu, Shue; Levitt, Roy C; Candiotti, Keith A; Lubarsky, David A; Hao, Shuanglin
HIV-associated sensory neuropathy affects over 50% of HIV patients and is a common peripheral nerve complication of HIV infection and highly active antiretroviral therapy (HAART). Evidence shows that painful HIV sensory neuropathy is influenced by neuroinflammatory events that include the proinflammatory molecules, MAP Kinase, tumor necrosis factor-α (TNFα), stromal cell-derived factor 1-α (SDF1α), and C-X-C chemokine receptor type 4 (CXCR4). However, the exact mechanisms of painful HIV sensory neuropathy are not known, which hinders our ability to develop effective treatments. In this study, we investigated whether inhibition of proinflammatory factors reduces the HIV-associated neuropathic pain state. Neuropathic pain was induced by peripheral HIV coat protein gp120 combined with 2',3'-dideoxycytidine (ddC, one of the nucleoside reverse transcriptase inhibitors (NRTIs)). Mechanical threshold was tested using von Frey filament fibers. Non-replicating herpes simplex virus (HSV) vectors expressing interleukin 10 (IL10) were inoculated into the hindpaws of rats. The expression of TNFα, SDF1α, and CXCR4 in the lumbar spinal cord and L4/5 dorsal root ganglia (DRG) was examined using western blots. IL-10 expression mediated by the HSV vectors resulted in a significant elevation of mechanical threshold. The anti-allodynic effect of IL-10 expression mediated by the HSV vectors lasted more than 3 weeks. The area under the effect-time curves (AUC) in mechanical threshold in rats inoculated with the HSV vectors expressing IL-10, was increased compared with the control vectors, indicating antinociceptive effect of the IL-10 vectors. The HSV vectors expressing IL-10 also concomitantly reversed the upregulation of p-p38, TNFα, SDF1α, and CXCR4 induced by gp120 in the lumbar spinal dorsal horn and/or the DRG at 2 and/or 4 weeks. The blocking of the signaling of these proinflammatory molecules is able to reduce HIV-related neuropathic pain, which provide a novel
Full Text Available Abstract Background Foot-and-mouth disease virus (FMDV causes an economically important and highly contagious disease of cloven-hoofed animals. RNAi triggered by small RNA molecules, including siRNAs and miRNAs, offers a new approach for controlling viral infections. There is no report available for FMDV inhibition by vector-delivered miRNA, although miRNA is believed to have more potential than siRNA. In this study, the inhibitory effects of vector-delivered miRNAs targeting the 3D gene on FMDV replication were examined. Results Four pairs of oligonucleotides encoding 3D-specific miRNA of FMDV were designed and selected for construction of miRNA expression plasmids. In the reporter assays, two of four miRNA expression plasmids were able to significantly silence the expression of 3D-GFP fusion proteins from the reporter plasmid, p3D-GFP, which was cotransfected with each miRNA expression plasmid. After detecting the silencing effects of the reporter genes, the inhibitory effects of FMDV replication were determined in the miRNA expression plasmid-transfected and FMDV-infected cells. Virus titration and real-time RT-PCR assays showed that the p3D715-miR and p3D983-miR plasmids were able to potently inhibit the replication of FMDV when BHK-21 cells were infected with FMDV. Conclusion Our results indicated that vector-delivered miRNAs targeting the 3D gene efficiently inhibits FMDV replication in vitro. This finding provides evidence that miRNAs could be used as a potential tool against FMDV infection.
Melanson, Vanessa R; Jochim, Ryan; Yarnell, Michael; Ferlez, Karen Bingham; Shashikumar, Soumya; Richardson, Jason H
Vector-borne pathogen surveillance programmes typically rely on the collection of large numbers of potential vectors followed by screening protocols focused on detecting pathogens in the arthropods. These processes are laborious, time consuming, expensive, and require screening of large numbers of samples. To streamline the surveillance process, increase sample throughput, and improve cost-effectiveness, a method to detect dengue virus and malaria parasites (Plasmodium falciparum) by leveraging the sugar-feeding behaviour of mosquitoes and their habit of expectorating infectious agents in their saliva during feeding was investigated in this study. Dengue virus 2 (DENV-2) infected female Aedes aegypti mosquitoes and P. falciparum infected female Anopheles stephensi mosquitoes were allowed to feed on honey coated Flinders Technical Associates -FTA® cards dyed with blue food colouring. The feeding resulted in deposition of saliva containing either DENV-2 particles or P. falciparum sporozoites onto the FTA card. Nucleic acid was extracted from each card and the appropriate real-time PCR (qPCR) assay was run to detect the pathogen of interest. As little as one plaque forming unit (PFU) of DENV-2 and as few as 60 P. falciparum parasites deposited on FTA cards from infected mosquitoes were detected via qPCR. Hence, their use to collect mosquito saliva for pathogen detection is a relevant technique for vector surveillance. This study provides laboratory confirmation that FTA cards can be used to capture and stabilize expectorated DENV-2 particles and P. falciparum sporozoites from infectious, sugar-feeding mosquitoes in very low numbers. Thus, the FTA card-based mosquito saliva capture method offers promise to overcome current limitations and revolutionize traditional mosquito-based pathogen surveillance programmes. Field testing and further method development are required to optimize this strategy.
Full Text Available Wolbachia is a maternally transmitted endosymbiotic bacterium that is estimated to infect up to 65% of insect species. The ability of Wolbachia to both induce pathogen interference and spread into mosquito vector populations makes it possible to develop Wolbachia as a biological control agent for vector-borne disease control. Although Wolbachia induces resistance to dengue virus (DENV, filarial worms, and Plasmodium in mosquitoes, species like Aedes polynesiensis and Aedes albopictus, which carry native Wolbachia infections, are able to transmit dengue and filariasis. In a previous study, the native wPolA in Ae. polynesiensis was replaced with wAlbB from Ae. albopictus, and resulted in the generation of the transinfected "MTB" strain with low susceptibility for filarial worms. In this study, we compare the dynamics of DENV serotype 2 (DENV-2 within the wild type "APM" strain and the MTB strain of Ae. polynesiensis by measuring viral infection in the mosquito whole body, midgut, head, and saliva at different time points post infection. The results show that wAlbB can induce a strong resistance to DENV-2 in the MTB mosquito. Evidence also supports that this resistance is related to a dramatic increase in Wolbachia density in the MTB's somatic tissues, including the midgut and salivary gland. Our results suggests that replacement of a native Wolbachia with a novel infection could serve as a strategy for developing a Wolbachia-based approach to target naturally infected insects for vector-borne disease control.
Sallam, Mohamed F; Michaels, Sarah R; Riegel, Claudia; Pereira, Roberto M; Zipperer, Wayne; Lockaby, B Graeme; Koehler, Philip G
The consistent sporadic transmission of West Nile Virus (WNV) in the city of New Orleans justifies the need for distribution risk maps highlighting human risk of mosquito bites. We modeled the influence of biophysical and socioeconomic metrics on the spatio-temporal distributions of presence/vector-host contact (VHC) ratios of WNV vector, Culex quinquefasciatus , within their flight range . Biophysical and socioeconomic data were extracted within 5-km buffer radii around sampling localities of gravid female Culex quinquefasciatus . The spatio-temporal correlations between VHC data and 33 variables, including climate, land use-land cover (LULC), socioeconomic, and land surface terrain were analyzed using stepwise linear regression models (RM). Using MaxEnt, we developed a distribution model using the correlated predicting variables. Only 12 factors showed significant correlations with spatial distribution of VHC ratios ( R ² = 81.62, p < 0.01). Non-forested wetland (NFWL), tree density (TD) and residential-urban (RU) settings demonstrated the strongest relationship. The VHC ratios showed monthly environmental resilience in terms of number and type of influential factors. The highest prediction power of RU and other urban and built up land (OUBL), was demonstrated during May-August. This association was positively correlated with the onset of the mosquito WNV infection rate during June. These findings were confirmed by the Jackknife analysis in MaxEnt and independently collected field validation points. The spatial and temporal correlations of VHC ratios and their response to the predicting variables are discussed.
This study describes the stomatal response occurring during water stress and subsequent recovery of three Eucalyptus grandis clonal hybrids. The aim was to investigate the degree to which stomatal conductance (gs) and stomatal density differ between the clonal hybrids across seasons and in response to water stress.
Lin, Yan-Shih; Medlyn, Belinda E.; Duursma, Remko A.; Prentice, I. Colin; Wang, Han; Baig, Sofia; Eamus, Derek; de Dios, Victor Resco; Mitchell, Patrick; Ellsworth, David S.; de Beeck, Maarten Op; Wallin, Göran; Uddling, Johan; Tarvainen, Lasse; Linderson, Maj-Lena; Cernusak, Lucas A.; Nippert, Jesse B.; Ocheltree, Troy W.; Tissue, David T.; Martin-Stpaul, Nicolas K.; Rogers, Alistair; Warren, Jeff M.; de Angelis, Paolo; Hikosaka, Kouki; Han, Qingmin; Onoda, Yusuke; Gimeno, Teresa E.; Barton, Craig V. M.; Bennie, Jonathan; Bonal, Damien; Bosc, Alexandre; Löw, Markus; Macinins-Ng, Cate; Rey, Ana; Rowland, Lucy; Setterfield, Samantha A.; Tausz-Posch, Sabine; Zaragoza-Castells, Joana; Broadmeadow, Mark S. J.; Drake, John E.; Freeman, Michael; Ghannoum, Oula; Hutley, Lindsay B.; Kelly, Jeff W.; Kikuzawa, Kihachiro; Kolari, Pasi; Koyama, Kohei; Limousin, Jean-Marc; Meir, Patrick; Lola da Costa, Antonio C.; Mikkelsen, Teis N.; Salinas, Norma; Sun, Wei; Wingate, Lisa
Stomatal conductance (gs) is a key land-surface attribute as it links transpiration, the dominant component of global land evapotranspiration, and photosynthesis, the driving force of the global carbon cycle. Despite the pivotal role of gs in predictions of global water and carbon cycle changes, a global-scale 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 differs among PFTs according to their marginal carbon cost of water use, as predicted by the theory underpinning the optimal stomatal model and the leaf and wood economics spectrum. We also demonstrate a global relationship with climate. These findings provide a robust theoretical framework for understanding and predicting the behaviour of gs across biomes and across PFTs that can be applied to regional, continental and global-scale modelling of ecosystem productivity, energy balance and ecohydrological processes in a future changing climate.
Jiang, Xiaohong; Dalebout, Tim J; Bredenbeek, Peter J; Carrion, Ricardo; Brasky, Kathleen; Patterson, Jean; Goicochea, Marco; Bryant, Joseph; Salvato, Maria S; Lukashevich, Igor S
Yellow Fever (YF) and Lassa Fever (LF) are two prevalent hemorrhagic fevers co-circulating in West Africa and responsible for thousands of deaths annually. The YF vaccine 17D has been used as a vector for the Lassa virus glycoprotein precursor (LASV-GPC) or their subunits, GP1 (attachment glycoprotein) and GP2 (fusion glycoprotein). Cloning shorter inserts, LASV-GP1 and -GP2, between YF17D E and NS1 genes enhanced genetic stability of recombinant viruses, YF17D/LASV-GP1 and -GP2, in comparison with YF17D/LASV-GPC recombinant. The recombinant viruses were replication competent and properly processed YF proteins and LASV GP antigens in infected cells. YF17D/LASV-GP1 and -GP2 induced specific CD8+ T cell responses in mice and protected strain 13 guinea pigs against fatal LF. Unlike immunization with live attenuated reassortant vaccine ML29, immunization with YF17D/LASV-GP1 and -GP2 did not provide sterilizing immunity. This study demonstrates the feasibility of YF17D-based vaccine to control LF in West Africa. Copyright Â© 2010 Elsevier Ltd. All rights reserved.
Jiang, Xiaohong; Dalebout, Tim J.; Bredenbeek, Peter J.; Carrion, Ricardo; Brasky, Kathleen; Patterson, Jean; Goicochea, Marco; Bryant, Joseph; Salvato, Maria S.; Lukashevich, Igor S.
Yellow Fever (YF) and Lassa Fever (LF) are two prevalent hemorrhagic fevers co-circulating in West Africa and responsible for thousands of deaths annually. The YF vaccine 17D has been used as a vector for the Lassa virus glycoprotein precursor (LASV-GPC) or their subunits, GP1 (attachment glycoprotein) and GP2 (fusion glycoprotein). Cloning shorter inserts, LASV GP1 and GP2, between YF17D E and NS1 genes enhanced genetic stability of recombinant viruses, YF17D/LASV-GP1 and –GP2, in comparison with YF17D/LASV-GPC recombinant. The recombinant viruses were replication competent and properly processed YF and LASV GP proteins in infected cells. YF17D/LASV-GP1&GP2 induced specific CD8+ T cell responses in mice and protected strain 13 guinea pigs against fatal LF. Unlike immunization with live attenuated reassortant vaccine ML29, immunization with YF17D/LASV-GP1&GP2 did not provide sterilizing immunity. This study demonstrates the feasibility of YF17D-based vaccine to control LF in West Africa. PMID:21145373
Andret-Link, Peggy; Schmitt-Keichinger, Corinne; Demangeat, Gerard; Komar, Veronique; Fuchs, Marc
The viral determinants involved in the specific transmission of Grapevine fanleaf virus (GFLV) by its nematode vector Xiphinema index are located within the 513 C-terminal residues of the RNA2-encoded polyprotein, that is, the 9 C-terminal amino acids of the movement protein (2B MP ) and contiguous 504 amino acids of the coat protein (2C CP ) [Virology 291 (2001) 161]. To further delineate the viral determinants responsible for the specific spread, the four amino acids that are different within the 9 C-terminal 2B MP residues between GFLV and Arabis mosaic virus (ArMV), another nepovirus which is transmitted by Xiphinema diversicaudatum but not by X. index, were subjected to mutational analysis. Of the recombinant viruses derived from transcripts of GFLV RNA1 and RNA2 mutants that systemically infected herbaceous host plants, all with the 2C CP of GFLV were transmitted by X. index unlike none with the 2C CP of ArMV, regardless of the mutations within the 2B MP C-terminus. These results demonstrate that the coat protein is the sole viral determinant for the specific spread of GFLV by X. index
Wu, Xiao-Xin; Yao, Hang-Ping; Wu, Nan-Ping; Gao, Hai-Nv; Wu, Hai-Bo; Jin, Chang-Zhong; Lu, Xiang-Yun; Xie, Tian-Shen; Li, Lan-Juan
Ebolaviruses are highly infectious pathogens that cause lethal Ebola virus disease (EVD) in humans and non-human primates (NHPs). Due to their high pathogenicity and transmissibility, as well as the potential to be misused as a bioterrorism agent, ebolaviruses would threaten the health of global populations if not controlled. In this review, we describe the origin and structure of ebolaviruses and the development of vaccines from the beginning of the 1980s, including conventional ebolavirus vaccines, DNA vaccines, Ebola virus-like particles (VLPs), vaccinia virus-based vaccines, Venezuelan equine encephalitis virus (VEEV)-like replicon particles, Kunjin virus-based vaccine, recombinant Zaire Ebolavirusx2206;VP30, recombinant cytomegalovirus (CMV)-based vaccines, recombinant rabies virus (RABV)-based vaccines, recombinant paramyxovirus-based vaccines, adenovirus-based vaccines and vesicular stomatitis virus (VSV)-based vaccines. No licensed vaccine or specific treatment is currently available to counteract ebolavirus infection, although DNA plasmids and several viral vector approaches have been evaluated as promising vaccine platforms. These vaccine candidates have been confirmed to be successful in protecting NHPs against lethal infection. Moreover, these vaccine candidates were successfully advanced to clinical trials. The present review provides an update of the current research on Ebola vaccines, with the aim of providing an overview on current prospects in the fight against EVD. © 2015 The Author(s) Published by S. Karger AG, Basel.
Full Text Available Ebolaviruses are highly infectious pathogens that cause lethal Ebola virus disease (EVD in humans and non-human primates (NHPs. Due to their high pathogenicity and transmissibility, as well as the potential to be misused as a bioterrorism agent, ebolaviruses would threaten the health of global populations if not controlled. In this review, we describe the origin and structure of ebolaviruses and the development of vaccines from the beginning of the 1980s, including conventional ebolavirus vaccines, DNA vaccines, Ebola virus-like particles (VLPs, vaccinia virus-based vaccines, Venezuelan equine encephalitis virus (VEEV-like replicon particles, Kunjin virus-based vaccine, recombinant Zaire Ebolavirus∆VP30, recombinant cytomegalovirus (CMV-based vaccines, recombinant rabies virus (RABV-based vaccines, recombinant paramyxovirus-based vaccines, adenovirus-based vaccines and vesicular stomatitis virus (VSV-based vaccines. No licensed vaccine or specific treatment is currently available to counteract ebolavirus infection, although DNA plasmids and several viral vector approaches have been evaluated as promising vaccine platforms. These vaccine candidates have been confirmed to be successful in protecting NHPs against lethal infection. Moreover, these vaccine candidates were successfully advanced to clinical trials. The present review provides an update of the current research on Ebola vaccines, with the aim of providing an overview on current prospects in the fight against EVD.
Tratschin, J.D.; West, M.H.P.; Sandbank, T.; Carter, B.J.
The authors have used the defective human parvovirus adeno-associated virus (AAV) as a novel eurocaryotic vector (parvector) for the expression of a foreign gene in human cells. The recombinant, pAV2, contains the AAV genome in a pBR322-derived bacterial plasmid. When pAV2 is transfected into human cells together with helper adenovirus particles, the AAV genome is rescued from the recombinant plasmid and replicated to produce infectious AAV particles at high efficiency. To create a vector, we inserted a procaryotic sequence coding for chloramphenicol acetyltransferase (CAT) into derivatives of pAV2 following either of the AAV promoters p/sub 40/ (pAVHiCAT) and p/sub 19/ (pAVBcCAT). When transfected into human 293 cells or HeLa cells, pAVHiCAT expressed CAT activity in the absence of adenovirus. In the presence of adenovirus, this vector produced increased amounts of CAT activity and the recombinant AAV-CAT genome was replicated. In 293 cells, pAVBcCAT expressed a similar amount of CAT activity in the absence or presence of adenovirus and the recombinant AAV-CAT genome was not replicated. In HeLa cells, pAVBcCAT expressed low levels of CAT activity, but this level was elevated by coinfection with adenovirus particles or by cotransfection with a plasmid which expressed the adenovirus early region 1A (E1A) product. The E1A product is a transcriptional activator and is expressed in 293 cells. Thus, expression from two AAV promoters is differentially regulated: expression from p/sub 19/ is increased by E1A, whereas p/sub 40/ yields high levels of constitutive expression in the absence of E1A. Both AAV vectors were packaged into AAV particles by complementation with wild-type AAV and yielded CAT activity when subsequently infected into cells in the presence of adenovirus.
Lee, John S; Hadjipanayis, Angela G; Parker, Michael D
.... DNA vectors, live-attenuated viruses and bacteria, recombinant proteins combined with adjuvant, and viral- or bacterial-vectored vaccines have been developed as countermeasures against many potential...
Diseño y construcción de vectores de transferencia para la obtención de virus vaccinia Ankara modificado (MVA recombinantes Design and construction of transfer vectors in order to obtain recombinant modified vaccinia virus Ankara (MVA
M. F. Ferrer
Full Text Available El virus vaccinia Ankara modificado (MVA constituye un buen candidato para el desarrollo de vectores virales de expresión no replicativos porque no replica en la mayoría de las células de mamíferos. Para la producción de MVA recombinantes es fundamental disponer de vectores de transferencia que, por recombinación homóloga con el genoma viral, permitan introducir los genes de interés en regiones no esenciales para la replicación in vitro. En este trabajo se diseñaron y obtuvieron los vectores de transferencia denominados VT-MHA y VT-MTK que portan las regiones correspondientes a las posiciones 1-303 y 608-948 del gen MVA165R y 1-244 y 325-534 del gen MVA086R, respectivamente, las que flanquean un sitio de clonado múltiple para la inserción de los genes foráneos. En dichos vectores se clonaron los casetes para la expresión de los genes lac Z o uid A, y la actividad de las enzimas marcadoras b-galactosidasa y b-glucuronidasa se confirmó in situ. Además, utilizando el vector denominado VT-MTK-GUS, se obtuvieron y aislaron MVA recombinantes puros que portan y expresan el gen uid A. Los resultados obtenidos constituyen las herramientas básicas para establecer la metodología de obtención de MVA recombinantes, con el propósito de desarrollar localmente vectores virales no replicativos candidatos a vacunas.Modified Vaccinia virus Ankara (MVA constitutes a good candidate for the development of non-replicative expression viral vectors because it does not replicate in most of mammalian cells. It is essential, for the production of recombinant MVA, the availability of transfer vectors which allow the introduction of desired genes into non-essential regions for in vitro viral replication, by homologous recombination with the viral genome. In the present work, the transfer vectors named VT-MHA and VT-MTK were designed and obtained. They carried genomic regions corresponding to 1- 303 and 608-948 positions of the MVA165R gene and 1-244 and
Hunt, G J; Tabachnick, W J
Equipment and procedures are described for biosafety level 3 (BL-3) containment work with small, zoophilic arthropods. BL-3 classified pathogens always must be manipulated in biological safety cabinets. Procedures, including physical barriers and handling methods, that prevent the escape of potentially virus-infected insects are discussed, and the use of a monitoring system for insect security is explained. The inability to recover escaped minute, flying insects poses a major difference from similar work with larger insects, such as mosquitoes. Methods were developed for the safe and secure handling of Culicoides variipennis sonorensis Wirth & Jones infected with exotic bluetongue viruses during BL-3 containment.
Full Text Available Gene-based vaccination strategies, specifically viral vectors encoding vaccine immunogens are effective at priming strong immune responses. Mucosal routes offer practical advantages for vaccination by ease of needle-free administration, and immunogen delivery at readily accessible oral/nasal sites to efficiently induce immunity at distant gut and genital tissues. However, since mucosal tissues are inherently tolerant for induction of immune responses, incorporation of adjuvants for optimal mucosal vaccination strategies is important. We report here the effectiveness of alpha-galactosylceramide (α-GalCer, a synthetic glycolipid agonist of natural killer T (NKT cells, as an adjuvant for enhancing immunogenicity of vaccine antigens delivered using viral vectors by mucosal routes in murine and nonhuman primate models. Significant improvement in adaptive immune responses in systemic and mucosal tissues was observed by including α-GalCer adjuvant for intranasal immunization of mice with vesicular stomatitis virus vector encoding the model antigen ovalbumin and adenoviral vectors expressing HIV env and Gag antigens. Activation of NKT cells in systemic and mucosal tissues along with significant increases in adaptive immune responses were observed in rhesus macaques immunized by intranasal and sublingual routes with protein or adenovirus vectored antigens when combined with α-GalCer adjuvant. These results support the utility of α-GalCer adjuvant for enhancing immunogenicity of mucosal vaccines delivered using viral vectors.
Pope, K. O.; Sheffner, E. J.; Linthicum, K. J.; Bailey, C. L.; Logan, T. M.; Kasischke, E. S.; Birney, K.; Njogu, A. R.; Roberts, C. R.
Rift Valley Fever (RVF) is a mosquito-borne virus that affects livestock and humans in Africa. Landsat TM data are shown to be effective in identifying dambos, intermittently flooded areas that are potential mosquite breeding sites, in an area north of Nairobi, Kenya. Positive results were obtained from a limited test of flood detection in dambos with airborne high resolution L, C, and X band multipolarization SAR imagery. L and C bands were effective in detecting flooded dambos, but LHH was by far the best channel for discrimination between flooded and nonflooded sites in both sedge and short-grass environments. This study demonstrates the feasibility of a combined passive and active remote sensing program for monitoring the location and condition of RVF vector habitats, thus making future control of the disease more promising.
Over 300 viruses are transmitted by the whitefly, Bemisia tabaci, with 90% of them belonging to the genus, Begomovirus. Begomoviruses are exclusively transmitted by whiteflies to a range of agriculture crops, resulting in billions of dollars lost annually, while jeopardizing food security worldwide....
Wheat streak mosaic virus (WSMV; genus Tritimovirus; family Potyviridae), is transmitted by the wheat curl mite (Aceria tosichella Keifer). The requirement of coat protein (CP) for WSMV transmission by the wheat curl mite was examined using a series of viable deletion and point mutations. Mite trans...
Full Text Available Abstract Background: Newcastle disease virus (NDV is one of the major pathogens in poultry and vaccination is intended to control the disease, as an effective solution, yet. Fusion protein (F on surface of NDV, has a fundamental role in virus pathogenicity and can induce protective immunity, alone. With this background, here our aim was to construct a baculovirus derived recombinant bacmid shuttle vector (encoding F-protein in order to express it in insect cell line. Materials and Methods: In this experimental study, at first complete F gene from avirulent strain La Sota of NDV was amplified by RT-PCR to produce F cDNA. The amplicon was cloned into T/A cloning vector and afterwards into pFastBac Dual donor plasmid. After the verification of cloning process by two methods, PCR and enzymatic digestion analysis, the accuracy of F gene sequence was confirmed by sequencing. Finally, F-containing recombinant bacmid was subsequently generated in DH10Bac cell and the construct production was confirmed by a special PCR panel, using F specific primers and M13 universal primers. Results: Analysis of confirmatory tests showed that the recombinant bacmid, expressing of F-protein gene in correct sequence and framework, has been constructed successfully. Conclusion: The product of this F-containing recombinant bacmid, in addition to its independent application in the induction of protective immunity, can be used with the other individual recombinant baculoviruses, expressing HN and NP genes to produce NDV-VLPs in insect cell line.
The full-length sequence of a new isolate of Apple chlorotic leaf spot virus (ACLSV) from Korea was divergent, but most closely related to the Japanese isolate A4, at 84% nucleotide identity. The full-length cDNA of the Korean isolate of ACLSV was cloned into a binary vector downstream of the bacter...
Andrew F van den Hurk
Full Text Available Incidence of disease due to dengue (DENV, chikungunya (CHIKV and yellow fever (YFV viruses is increasing in many parts of the world. The viruses are primarily transmitted by Aedes aegypti, a highly domesticated mosquito species that is notoriously difficult to control. When transinfected into Ae. aegypti, the intracellular bacterium Wolbachia has recently been shown to inhibit replication of DENVs, CHIKV, malaria parasites and filarial nematodes, providing a potentially powerful biocontrol strategy for human pathogens. Because the extent of pathogen reduction can be influenced by the strain of bacterium, we examined whether the wMel strain of Wolbachia influenced CHIKV and YFV infection in Ae. aegypti. Following exposure to viremic blood meals, CHIKV infection and dissemination rates were significantly reduced in mosquitoes with the wMel strain of Wolbachia compared to Wolbachia-uninfected controls. However, similar rates of infection and dissemination were observed in wMel infected and non-infected Ae. aegypti when intrathoracic inoculation was used to deliver virus. YFV infection, dissemination and replication were similar in wMel-infected and control mosquitoes following intrathoracic inoculations. In contrast, mosquitoes with the wMelPop strain of Wolbachia showed at least a 10(4 times reduction in YFV RNA copies compared to controls. The extent of reduction in virus infection depended on Wolbachia strain, titer and strain of the virus, and mode of exposure. Although originally proposed for dengue biocontrol, our results indicate a Wolbachia-based strategy also holds considerable promise for YFV and CHIKV suppression.
Wang, JianYing; Zhang, Tong; Lu, Yi; Zhou, GuangYa; Chen, Qin; Niu, Bing
Vesicular stomatitis (VS) is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends. American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression. For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity), SP (specificity) and ACC (prediction accuracy) values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively. This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast.
Lu, Yi; Zhou, GuangYa; Chen, Qin
Background Vesicular stomatitis (VS) is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends. Methods American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression. Results For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity), SP (specificity) and ACC (prediction accuracy) values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively. Conclusion This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast. PMID:29385198
Full Text Available Vesicular stomatitis (VS is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends.American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression.For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity, SP (specificity and ACC (prediction accuracy values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively.This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast.
Clement N Mweya
Full Text Available Rift Valley Fever (RVF is weather dependent arboviral infection of livestock and humans. Population dynamics of mosquito vectors is associated with disease epidemics. In our study, we use daily temperature and rainfall as model inputs to simulate dynamics of mosquito vectors population in relation to disease epidemics.Time-varying distributed delays (TVDD and multi-way functional response equations were implemented to simulate mosquito vectors and hosts developmental stages and to establish interactions between stages and phases of mosquito vectors in relation to vertebrate hosts for infection introduction in compartmental phases. An open-source modelling platforms, Universal Simulator and Qt integrated development environment were used to develop models in C++ programming language. Developed models include source codes for mosquito fecundity, host fecundity, water level, mosquito infection, host infection, interactions, and egg time. Extensible Markup Language (XML files were used as recipes to integrate source codes in Qt creator with Universal Simulator plug-in. We observed that Floodwater Aedines and Culicine population continued to fluctuate with temperature and water level over simulation period while controlled by availability of host for blood feeding. Infection in the system was introduced by floodwater Aedines. Culicines pick infection from infected host once to amplify disease epidemic. Simulated mosquito population show sudden unusual increase between December 1997 and January 1998 a similar period when RVF outbreak occurred in Ngorongoro district.Findings presented here provide new opportunities for weather-driven RVF epidemic simulation modelling. This is an ideal approach for understanding disease transmission dynamics towards epidemics prediction, prevention and control. This approach can be used as an alternative source for generation of calibrated RVF epidemics data in different settings.
Mweya, Clement N; Holst, Niels; Mboera, Leonard E G; Kimera, Sharadhuli I
Rift Valley Fever (RVF) is weather dependent arboviral infection of livestock and humans. Population dynamics of mosquito vectors is associated with disease epidemics. In our study, we use daily temperature and rainfall as model inputs to simulate dynamics of mosquito vectors population in relation to disease epidemics. Time-varying distributed delays (TVDD) and multi-way functional response equations were implemented to simulate mosquito vectors and hosts developmental stages and to establish interactions between stages and phases of mosquito vectors in relation to vertebrate hosts for infection introduction in compartmental phases. An open-source modelling platforms, Universal Simulator and Qt integrated development environment were used to develop models in C++ programming language. Developed models include source codes for mosquito fecundity, host fecundity, water level, mosquito infection, host infection, interactions, and egg time. Extensible Markup Language (XML) files were used as recipes to integrate source codes in Qt creator with Universal Simulator plug-in. We observed that Floodwater Aedines and Culicine population continued to fluctuate with temperature and water level over simulation period while controlled by availability of host for blood feeding. Infection in the system was introduced by floodwater Aedines. Culicines pick infection from infected host once to amplify disease epidemic. Simulated mosquito population show sudden unusual increase between December 1997 and January 1998 a similar period when RVF outbreak occurred in Ngorongoro district. Findings presented here provide new opportunities for weather-driven RVF epidemic simulation modelling. This is an ideal approach for understanding disease transmission dynamics towards epidemics prediction, prevention and control. This approach can be used as an alternative source for generation of calibrated RVF epidemics data in different settings.
Brustolin, M; Talavera, S; Santamaría, C; Rivas, R; Pujol, N; Aranda, C; Marquès, E; Valle, M; Verdún, M; Pagès, N; Busquets, N
The emerging disease West Nile fever is caused by West Nile virus (WNV), one of the most widespread arboviruses. This study represents the first test of the vectorial competence of European Culex pipiens Linnaeus 1758 and Stegomyia albopicta (= Aedes albopictus) (both: Diptera: Culicidae) populations for lineage 1 and 2 WNV isolated in Europe. Culex pipiens and S. albopicta populations were susceptible to WNV infection, had disseminated infection, and were capable of transmitting both WNV lineages. This is the first WNV competence assay to maintain mosquito specimens under environmental conditions mimicking the field (day/night) conditions associated with the period of maximum expected WNV activity. The importance of environmental conditions is discussed and the issue of how previous experiments conducted in fixed high temperatures may have overestimated WNV vector competence results with respect to natural environmental conditions is analysed. The information presented should be useful to policymakers and public health authorities for establishing effective WNV surveillance and vector control programmes. This would improve preparedness to prevent future outbreaks. © 2016 The Authors. Medical and Veterinary Entomology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.
Tsukahara, Tomonori; Agawa, Hideyuki; Matsumoto, Sayori; Matsuda, Mizuho; Ueno, Shuichi; Yamashita, Yuki; Yamada, Koichiro; Tanaka, Nobuyuki; Kojima, Katsuhiko; Takeshita, Toshikazu
Genomic analysis of integration will be important in evaluating the safety of human gene therapy with retroviral vectors. Here, we investigated MLV vector integration sites in human T-cells, since they are amenable to gene transfer studies, and have been used therapeutically in clinical trials. We mapped 340 MLV vector integration sites in the infected human T-cell clones we established. The data showed that MLV preferred integration near the transcription start sites (±5 kb), near CpG islands (±1 kb), and within the first intron of RefSeq genes. We also identified MLV integration hot spots that contained three or more integrations within a 100 kb region. RT-PCR revealed that mRNA-levels of T-cell clones that contained MLV integrations near transcription start sites or introns were dysregulated compared to the uninfected cells. These studies help define the profile of MLV integration in T-cells and the risks associated with MLV-based gene therapy
Osório, Hugo Costa; Zé-Zé, Líbia; Alves, Maria João
The host blood-feeding patterns of mosquito vectors affects the likelihood of human exposure to zoonotic pathogens, including West Nile Virus (family Flaviviridae, genus Flavivirus, WNV). In Portugal, data are unavailable regarding the blood-feeding habits of common mosquito species, including Culex pipiens L., considered the primary vector of WNV to humans. The sources of bloodmeals in 203 blood-fed mosquitoes of nine species collected from June 2007 to November 2010 in 34 Portuguese counties were analyzed by sequencing cytochrome-b partial fragments. Cx. pipiens was the most common species collected and successfully analyzed (n = 135/78). In addition, blood-fed females of the following species were analyzed: Ochlerotatus caspius Pallas (n = 20), Culex theileri Theobald (n = 16), Anopheles maculipennis s.l. Meigen (n = 10), Culiseta longiareolata Macquart (n = 7), Aedes aegypti L. (n = 6), Culex perexiguus Theobald (n = 3), Culiseta annulata Schrank (n = 3), and Ochlerotatus detritus Haliday (n = 3). The Cx. pipiens mosquitoes fed predominantly on birds (n = 55/78, 70.5%), with a high diversity of avian species used as hosts, although human blood was identified in 18 specimens (18/78, 23.1%). No significant differences were found between the host-feeding patterns of blood-fed Cx. pipiens collected in residential and nonresidential habitats. The occurrence of human derived blood meals and the presence of a mix avian-human bloodmeal accordingly suggest this species as a potential vector of WNV. Therefore, in Portugal, Cx. pipiens may play a role both in the avian-to-avian enzootic WNV cycle and in the avian-to-mammal transmission. In this context, the identity of Cx. pipiens (considering the forms molestus and pipiens) and the potential consequence on feeding behavior and WNV transmission are discussed.
Al-Nbaheen, M.; Pourzand, C.; Tyrrell, R.M.
One way of targeting gene expression in vivo is to control transcription using a tissue-specific regulatory system. Tissue specific promoters or enhancers are in use in transgenic animals and could be utilized in medical for gene therapy. At present the usual method for selection of a tissue-specific promoter is to identify a gene, which is expressed at unusually high level in the target tissue, and then to use the promoter for this gene to drive expression of another therapeutic gene in the target tissue. This approach is logical but does not always lead to high levels of gene expression. A second approach is to investigate the scope for discovery of synthetic specific promoters using a target tissue. The objective of the work described in this paper was to use both approach to design plasmid DNA expression vectors that would carry liver-specific promoter/enhancer linked to reporter gene (i.e. luciferase). Then transfect these vectors to both liver-derived and non-liver cell lines. This is followed by evaluation of the liver-specificity of each construct by measuring the basal level expression of the reporter gene (i.e. luciferase activity) in both cell lines. Hepatocyte nuclear factor-4 (HNF-4) is liver-enriched transcription factor used to design new synthetic enhancers by inserting a tandem array of 1', 3' or 5' repeats of the HNF-4 binding site upstream of the SV40 promoter linked to the luciferase reporter gene within an Epstein-Barr virus (EBV)-based vector, p 706. The results of transfection revealed that unexpectedly the HNF-4 binding sites in these constructs act as a repressor rather than enhancer of the liver-specific expression of the luciferase gene. (author)
Hemphill, D D; McIlwraith, C W; Slayden, R A; Samulski, R J; Goodrich, L R
IGF-I is one of several anabolic factors being investigated for the treatment of osteoarthritis (OA). Due to the short biological half-life, extended administration is required for more robust cartilage healing. Here we create a self-complimentary adeno-associated virus (AAV) gene therapy vector utilizing the transgene for IGF-I. Various biochemical assays were performed to investigate the cellular response to scAAVIGF-I treatment vs an scAAVGFP positive transduction control and a negative for transduction control culture. RNA-sequencing analysis was also performed to establish a differential regulation profile of scAAVIGF-I transduced chondrocytes. Biochemical analyses indicated an average media IGF-I concentration of 608 ng/ml in the scAAVIGF-I transduced chondrocytes. This increase in IGF-I led to increased expression of collagen type II and aggrecan and increased protein concentrations of cellular collagen type II and media glycosaminoglycan vs both controls. RNA-seq revealed a global regulatory pattern consisting of 113 differentially regulated GO categories including those for chondrocyte and cartilage development and regulation of apoptosis. This research substantiates that scAAVIGF-I gene therapy vector increased production of IGF-I to clinically relevant levels with a biological response by chondrocytes conducive to increased cartilage healing. The RNA-seq further established a set of differentially expressed genes and gene ontologies induced by the scAAVIGF-I vector while controlling for AAV infection. This dataset provides a static representation of the cellular transcriptome that, while only consisting of one time point, will allow for further gene expression analyses to compare additional cartilage healing therapeutics or a transient cellular response. Copyright © 2015. Published by Elsevier Ltd.
Chimeric avian paramyxovirus-based vector immunization against highly pathogenic avian influenza followed by conventional Newcastle disease vaccination eliminates lack of protection from virulent ND virus
Full Text Available Recently, we described a chimeric, hemagglutinin of highly pathogenic avian influenza virus (HPAIV H5 expressing Newcastle disease virus (NDV-based vector vaccine (chNDVFHNPMV8H5 in which NDV envelope glycoproteins were replaced by those of avian paramyxovirus-8 (APMV-8. This chimeric vaccine induced solid protection against lethal HPAIV H5N1 even in chickens with maternal antibodies against NDV (MDA+. However, due to the absence of the major NDV immunogens it failed to induce protection against Newcastle disease (ND. Here, we report on protection of MDA+ chickens against HPAI H5N1 and ND, by vaccination with chNDVFHNPMV8H5 either on day 1 or day seven after hatch, and subsequent immunization with live attenuated NDV seven days later. Vaccination was well tolerated and three weeks after immunization, challenge infections with highly pathogenic NDV as well as HPAIV H5N1 were carried out. All animals remained healthy without exhibiting any clinical signs, whereas non-vaccinated animals showed morbidity and mortality. Therefore, vaccination with chNDVFHNPMV8H5 can be followed by NDV vaccination to protect chickens from HPAIV as well as NDV, indicating that the antibody response against chNDVFHNPMV8H5 does not interfere with live ND vaccination.
Liu, Liang; Cooper, Tamara; Eldi, Preethi; Garcia-Valtanen, Pablo; Diener, Kerrilyn R; Howley, Paul M; Hayball, John D
Recombinant vaccinia viruses (rVACVs) are promising antigen-delivery systems for vaccine development that are also useful as research tools. Two common methods for selection during construction of rVACV clones are (i) co-insertion of drug resistance or reporter protein genes, which requires the use of additional selection drugs or detection methods, and (ii) dominant host-range selection. The latter uses VACV variants rendered replication-incompetent in host cell lines by the deletion of host-range genes. Replicative ability is restored by co-insertion of the host-range genes, providing for dominant selection of the recombinant viruses. Here, we describe a new method for the construction of rVACVs using the cowpox CP77 protein and unmodified VACV as the starting material. Our selection system will expand the range of tools available for positive selection of rVACV during vector construction, and it is substantially more high-fidelity than approaches based on selection for drug resistance.
Full Text Available Influenza (flu pandemics have exhibited a great threat to human health throughout history. With the emergence of drug-resistant strains of influenza A virus (IAV, it is necessary to look for new agents for treatment and transmission prevention of the flu. Defensins are small (2–6 kDa cationic peptides known for their broad-spectrum antimicrobial activity. Beta-defensins (β-defensins are mainly produced by barrier epithelial cells and play an important role in attacking microbe invasion by epithelium. In this study, we focused on the anti-influenza A virus activity of mouse β-defensin 1 (mBD1 and β defensin-3 (mBD3 by synthesizing their fusion peptide with standard recombinant methods. The eukaryotic expression vectors pcDNA3.1(+/mBD1-mBD3 were constructed successfully by overlap-PCR and transfected into Madin-Darby canine kidney (MDCK cells. The MDCK cells transfected by pcDNA3.1(+/mBD1-mBD3 were obtained by G418 screening, and the mBD1-mBD3 stable expression pattern was confirmed in MDCK cells by RT-PCR and immunofluorescence assay. The acquired stable transfected MDCK cells were infected with IAV (A/PR/8/34, H1N1, 0.1 MOI subsequently and the virus titers in cell culture supernatants were analyzed by TCID50 72 h later. The TCID50 titer of the experimental group was clearly lower than that of the control group (p < 0.001. Furthermore, BALB/C mice were injected with liposome-encapsulated pcDNA3.1(+/mBD1-mBD3 through muscle and then challenged with the A/PR/8/34 virus. Results showed the survival rate of 100% and lung index inhibitory rate of 32.6% in pcDNA3.1(+/mBD1-mBD3group; the TCID50 titer of lung homogenates was clearly lower than that of the control group (p < 0.001. This study demonstrates that mBD1-mBD3 expressed by the recombinant plasmid pcDNA3.1(+/mBD1-mBD3 could inhibit influenza A virus replication both in vitro and in vivo. These observations suggested that the recombinant mBD1-mBD3 might be developed into an agent for
Fischer, Dominik; Thomas, Stephanie M; Suk, Jonathan E; Sudre, Bertrand; Hess, Andrea; Tjaden, Nils B; Beierkuhnlein, Carl; Semenza, Jan C
Chikungunya was, from the European perspective, considered to be a travel-related tropical mosquito-borne disease prior to the first European outbreak in Northern Italy in 2007. This was followed by cases of autochthonous transmission reported in South-eastern France in 2010. Both events occurred after the introduction, establishment and expansion of the Chikungunya-competent and highly invasive disease vector Aedes albopictus (Asian tiger mosquito) in Europe. In order to assess whether these outbreaks are indicative of the beginning of a trend or one-off events, there is a need to further examine the factors driving the potential transmission of Chikungunya in Europe. The climatic suitability, both now and in the future, is an essential starting point for such an analysis. The climatic suitability for Chikungunya outbreaks was determined by using bioclimatic factors that influence, both vector and, pathogen. Climatic suitability for the European distribution of the vector Aedes albopictus was based upon previous correlative environmental niche models. Climatic risk classes were derived by combining climatic suitability for the vector with known temperature requirements for pathogen transmission, obtained from outbreak regions. In addition, the longest potential intra-annual season for Chikungunya transmission was estimated for regions with expected vector occurrences.In order to analyse spatio-temporal trends for risk exposure and season of transmission in Europe, climate change impacts are projected for three time-frames (2011-2040, 2041-2070 and 2071-2100) and two climate scenarios (A1B and B1) from the Intergovernmental Panel on Climate Change (IPCC). These climatic projections are based on regional climate model COSMO-CLM, which builds on the global model ECHAM5. European areas with current and future climatic suitability of Chikungunya transmission are identified. An increase in risk is projected for Western Europe (e.g. France and Benelux-States) in the