Equine epizootics of Venezuelan equine encephalitis (VEE) occurred in the southern Mexican states of Chiapas in 1993 and Oaxaca in 1996. To assess the impact of continuing circulation of VEE virus (VEEV) on human and animal populations, serologic and viral isolation studies were conducted in 2000 to 2001 in Chiapas State. Human serosurveys and risk analyses indicated that long-term endemic transmission of VEEV occurred among villages with seroprevalence levels of 18% to 75% and that medical personnel had a high risk for VEEV exposure. Seroprevalence in wild animals suggested cotton rats as possible reservoir hosts in the region. Virus isolations from sentinel animals and genetic characterizations of these strains indicated continuing circulation of a subtype IE genotype, which was isolated from equines during the recent VEE outbreaks. These data indicate long-term enzootic and endemic VEEV circulation in the region and continued risk for disease in equines and humans. (+info)
Venezuelan equine encephalitis virus in the guinea pig model: evidence for epizootic virulence determinants outside the E2 envelope glycoprotein gene.
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Epizootic strains of Venezuelan equine encephalitis virus (VEEV) cause epidemics by exploiting equines as highly efficient amplification hosts for mosquito transmission. Although phylogenetic studies indicate that epizootic VEEV strains emerge via mutation from enzootic progenitors that are incapable of efficient equine amplification, the molecular mechanism(s) involved remain enigmatic. The convergent evolution of E2 envelope glycoprotein mutations suggests that they are critical to VEEV emergence, but little is known about the role of non-envelope genes. We used the guinea pig, the small animal model that best predicts the ability to generate equine viremia, to assess the role of envelope versus other mutations in the epizootic phenotype. Using reciprocal chimeric viruses generated by swapping the envelope genes of closely related epizootic IC and enzootic ID strains, infections of guinea pigs demonstrated that envelope and non-envelope genes and sequences both contributed to virulence. However, early replication in lymphoid tissues appeared to be primarily envelope dependent. (+info)
Heterologous gene expression by infectious and replicon vectors derived from tick-borne encephalitis virus and direct comparison of this flavivirus system with an alphavirus replicon.
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The flavivirus tick-borne encephaltis virus (TBEV) was established as a vector system for heterologous gene expression. The variable region of the genomic 3' non-coding region was replaced by an expression cassette consisting of the reporter gene enhanced green fluorescent protein (EGFP) under the translational control of an internal ribosomal entry site element, both in the context of an infectious virus genome and of a replicon lacking the genes of the surface proteins prM/M and E. The expression level and the stability of expression were measured by fluorescence-activated cell-sorting analysis and compared to an established alphavirus replicon vector derived from Venezuelan equine encephaltis virus (VEEV), expressing EGFP under the control of its natural subgenomic promoter. On the first day, the alphavirus replicon exhibited an approximately 180-fold higher expression level than the flavivirus replicon, but this difference decreased to about 20- and 10-fold on days 2 and 3, respectively. Four to six days post-transfection, foreign gene expression by the VEEV replicon vanished almost completely, due to extensive cell killing. In contrast, in the case of the TBEV replicon, the percentage of positive cells and the amount of EGFP expression exhibited only a moderate decline over a time period of almost 4 weeks. The infectious TBEV vector expressed less EGFP than the TBEV replicon at all times. Significant expression from the infectious vector was maintained for four cell-culture passages. The results indicate that the VEEV vector is superior with respect to achieving high expression levels, but the TBEV system may be advantageous for applications that require a moderate, but more enduring, gene expression. (+info)
Duration of infectivity and RNA of Venezuelan equine encephalitis, West Nile, and yellow fever viruses dried on filter paper and maintained at room temperature.
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Samples of laboratory propagated Venezuelan equine encephalitis (VEE), West Nile (WN), and yellow fever (YF) viruses were blotted onto filter paper discs, air-dried, and stored at room temperature. At regular intervals over a 90-day period, the dried virus samples were eluted, tested for infectivity by culture and titration in Vero cells, and examined for viral RNA by a reverse transcriptase-polymerase chain reaction. The VEE, WN, and YF viral RNA was detected throughout the 90-day period in all samples examined. Infectious VEE virus could be recovered for up to 40 days; WN and YF viruses were cultured in Vero cells for up to 60 and 90 days, respectively. The results of this study demonstrate that viral nucleic acids and infectious virus can be recovered from arbovirus samples air-dried on filter paper and stored at room temperature for a month or more after collection. This procedure offers a simple and inexpensive method for collecting arbovirus field specimens and transporting them to diagnostic laboratories. (+info)
Venezuelan equine encephalitis virus infection of spiny rats.
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Enzootic strains of Venezuelan equine encephalitis virus (VEEV) circulate in forested habitats of Mexico, Central, and South America, and spiny rats (Proechimys spp.) are believed to be the principal reservoir hosts in several foci. To better understand the host-pathogen interactions and resistance to disease characteristic of many reservoir hosts, we performed experimental infections of F1 progeny from Proechimys chrysaeolus collected at a Colombian enzootic VEEV focus using sympatric and allopatric virus strains. All animals became viremic with a mean peak titer of 3.3 log10 PFU/mL, and all seroconverted with antibody titers from 1:20 to 1:640, which persisted up to 15 months. No signs of disease were observed, including after intracerebral injections. The lack of detectable disease and limited histopathologic lesions in these animals contrast dramatically with the severe disease and histopathologic findings observed in other laboratory rodents and humans, and support their role as reservoir hosts with a long-term coevolutionary relationship to VEEV. (+info)
Characterization of human immunodeficiency virus Gag-specific gamma interferon-expressing cells following protective mucosal immunization with alphavirus replicon particles.
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A safe, replication-defective viral vector that can induce mucosal and systemic immune responses and confer protection against many infectious pathogens, such as human immunodeficiency virus type 1 (HIV-1), may be an ideal vaccine platform. Accordingly, we have generated and tested alphavirus replicon particles encoding HIV-1 Gag from Sindbis virus (SIN-Gag) and Venezuelan equine encephalitis virus (VEE-Gag), as well as chimeras between the two (VEE/SIN-Gag). Following intramuscular (i.m.), intranasal (i.n.), or intravaginal (IVAG) immunization with VEE/SIN-Gag and an IVAG challenge with vaccinia virus encoding HIV Gag (VV-Gag), a larger number of Gag-specific CD8+ intracellular gamma interferon-expressing cells (iIFNEC) were detected in iliac lymph nodes (ILN), which drain the vaginal/uterine mucosa (VUM), than were observed after immunizations with SIN-Gag. Moreover, a single i.n. or IVAG immunization with VEE/SIN-Gag induced a larger number of cells expressing HIV Gag in ILN, and immunizations with VEE/SIN-Gag through any route induced better protective responses than immunizations with SIN-Gag. In VUM, a larger percentage of iIFNEC expressed alpha4beta7 or alpha(Ebeta)7 integrin than expressed CD62L integrin. However, in spleens (SP), a larger percentage of iIFNEC expressed alpha4beta7 or CD62L than expressed alpha(Ebeta)7. Moreover, a larger percentage of iIFNEC expressed the chemokine receptor CCR5 in VUM and ILN than in SP. These results demonstrate a better induction of cellular and protective responses following immunizations with VEE/SIN-Gag than that following immunizations with SIN-Gag and also indicate a differential expression of homing and chemokine receptors on iIFNEC in mucosal effector and inductive sites versus systemic lymphoid tissues. (+info)
Noncytopathic replication of Venezuelan equine encephalitis virus and eastern equine encephalitis virus replicons in Mammalian cells.
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Venezuelan equine encephalitis (VEE) and eastern equine encephalitis (EEE) viruses are important, naturally emerging zoonotic viruses. They are significant human and equine pathogens which still pose a serious public health threat. Both VEE and EEE cause chronic infection in mosquitoes and persistent or chronic infection in mosquito-derived cell lines. In contrast, vertebrate hosts infected with either virus develop an acute infection with high-titer viremia and encephalitis, followed by host death or virus clearance by the immune system. Accordingly, EEE and VEE infection in vertebrate cell lines is highly cytopathic. To further understand the pathogenesis of alphaviruses on molecular and cellular levels, we designed EEE- and VEE-based replicons and investigated their replication and their ability to generate cytopathic effect (CPE) and to interfere with other viral infections. VEE and EEE replicons appeared to be less cytopathic than Sindbis virus-based constructs that we designed in our previous research and readily established persistent replication in BHK-21 cells. VEE replicons required additional mutations in the 5' untranslated region and nsP2 or nsP3 genes to further reduce cytopathicity and to become capable of persisting in cells with no defects in alpha/beta interferon production or signaling. The results indicated that alphaviruses strongly differ in virus-host cell interactions, and the ability to cause CPE in tissue culture does not necessarily correlate with pathogenesis and strongly depends on the sequence of viral nonstructural proteins. (+info)
A novel, rapid assay for detection and differentiation of serotype-specific antibodies to Venezuelan equine encephalitis complex alphaviruses.
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An epitope-blocking enzyme-linked immunosorbent assay was developed for the rapid differentiation of serologic responses to enzootic variety IE and ID versus epizootic variety IAB and IC strains of Venezuelan equine encephalitis (VEE) virus. Two monoclonal antibodies that differentially recognize epizootic versus enzootic VEE virus epitopes were used to measure the serotype-specific blocking abilities of antibodies in sera of naturally infected humans, equines, and bovines, as well as in experimentally infected equines. The assay is simple, species-independent, rapid, and sensitive, and will improve surveillance for VEE emergence. It could also be used to determine the epidemic potential of a VEE virus following an intentional introduction for bioterrorism. (+info)