Reverse Transcription-PCR-Enzyme-Linked Immunosorbent Assay for Rapid Detection and Differentiation of Alphavirus Infections

Wang, Eryu; Paessler, Slobodan; Aguilar, Patricia V.; Carrara, Anne-Sophie; Ni, Haolin; Greene, Ivorlyne P.; Weaver, Scott C.

doi:10.1128/jcm.00175-06

Cited by 33 publications

(28 citation statements)

References 21 publications

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“…Examination of these mutants revealed that L526 and L528 were required for function of the NES, while V521 was not necessary. Sequence analysis of the identified NES across 10 representative alphaviruses revealed that the presence of a hydrophobic residue at VEE position 526 is conserved and that the leucine residue at VEE position 528 is highly conserved (7,13,27,30,37,54,57,62,64).…”

Section: Comparison Of the Nlss Of Sfv And Veementioning

confidence: 99%

Nuclear Import and Export of Venezuelan Equine Encephalitis Virus Nonstructural Protein 2

Montgomery

Johnston

2007

J Virol

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Many RNA viruses, which replicate predominantly in the cytoplasm, have nuclear components that contribute to their life cycle or pathogenesis. We investigated the intracellular localization of the multifunctional nonstructural protein 2 (nsP2) in mammalian cells infected with Venezuelan equine encephalitis virus (VEE), an important, naturally emerging zoonotic alphavirus. VEE nsP2 localizes to both the cytoplasm and the nucleus of mammalian cells in the context of infection and also when expressed alone. Through the analysis of a series of enhanced green fluorescent protein fusions, a segment of nsP2 that completely localizes to the nucleus of mammalian cells was identified. Within this region, mutation of the putative nuclear localization signal (NLS) PGKMV diminished, but did not obliterate, the ability of the protein to localize to the nucleus, suggesting that this sequence contributes to the nuclear localization of VEE nsP2. Furthermore, VEE nsP2 specifically interacted with the nuclear import protein karyopherin-␣1 but not with karyopherin-␣2, -3, or -4, suggesting that karyopherin-␣1 transports nsP2 to the nucleus during infection. Additionally, a novel nuclear export signal (NES) was identified, which included residues L526 and L528 of VEE nsP2. Leptomycin B treatment resulted in nuclear accumulation of nsP2, demonstrating that nuclear export of nsP2 is mediated via the CRM1 nuclear export pathway. Disruption of either the NLS or the NES in nsP2 compromised essential viral functions. Taken together, these results establish the bidirectional transport of nsP2 across the nuclear membrane, suggesting that a critical function of nsP2 during infection involves its shuttling between the cytoplasm and the nucleus.Venezuelan equine encephalitis virus (VEE), a New World member of the genus Alphavirus in the Togaviridae family, is a pathogen that is responsible for significant disease in equines and can cause a wide range of human symptoms, from inapparent infection to acute encephalitis. In nature, VEE is transmitted between susceptible hosts through a mosquito vector and is responsible for periodic outbreaks of widespread human and equine disease throughout the Americas.The alphaviruses are enveloped viruses with an 11.5-kb genome of single-stranded message-sense RNA. The viral genome, which resembles cellular messages because it contains 5Ј-and 3Ј-untranslated regions, a 5Ј-terminal methylguanylate cap, and a 3Ј-terminal polyadenylate tail, encodes four nonstructural proteins (nsP1 through 4) and three mature structural proteins (capsid, E2, and E1). Upon infection, this RNA genome is directly translated in the cell to produce the nonstructural polyproteins P1234 and P123, depending on whether there is readthrough at the opal stop codon present between nsP3 and nsP4 in most alphaviruses (10, 32). The polyprotein is processed by nsP2, which contains a protease domain in its carboxy terminus, to form the four individual nonstructural proteins, nsP1 to -4 (22). Early during infection, negative-sense viral RNA is sy...

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Section: Comparison Of the Nlss Of Sfv And Veementioning

confidence: 99%

Nuclear Import and Export of Venezuelan Equine Encephalitis Virus Nonstructural Protein 2

Montgomery

Johnston

2007

J Virol

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“…La mayoría de los aislamientos del virus Mayaro en mosquitos se obtuvo a partir de Haemagogus recolectados en Brasil durante períodos interepidémicos, lo cual sugiere que podrían estar desempeñando un importante papel en el mantenimiento del virus en su ciclo enzoótico (33) (figura 3). Por otra parte, en muchas localidades donde abundan estas especies se ha encontrado alta seroprevalencia de anticuerpos contra virus Mayaro en monos de vida silvestre, que son la fuente alimentaria predilecta de los mosquitos Haemagogus; en los humanos, existe mayor seroprevalencia contra el virus Mayaro en la población de trabajadores agrícolas y exploradores (10)(11)(12)(13)33,39), patrón similar al que se presenta con el virus de la fiebre amarilla.…”

Section: Ciclo De Transmisión Del Virus Mayaro Artrópodos Vectoresunclassified

“…Un diagnóstico clínico del mal de Mayaro con base en la sintomatología inicial, es confuso por la existencia de cuadros clínicos similares, incluso con otros grupos antigénicos como el dengue y Oropuche, por lo que es de gran importancia emplear métodos diagnósticos inmunológicos y moleculares eficientes (11,13). Los métodos de laboratorio utilizados son el aislamiento viral mediante la inoculación de cultivos celulares en ratones experimentales con posterior aplicación (13,33,77,78).…”

Section: Brotes En Humanos Sintomatología Y Diagnósticounclassified

“…Los métodos de laboratorio utilizados son el aislamiento viral mediante la inoculación de cultivos celulares en ratones experimentales con posterior aplicación (13,33,77,78). Entre los marcadores moleculares utilizados se encuentran secuencias parciales del gen nsP1, de la región no codificante 3´(RNC), E1, E2 y E3 (2,10-13).…”

Section: Brotes En Humanos Sintomatología Y Diagnósticounclassified

“…El virus Mayaro, cuyo nombre se debe a la localidad donde fue aislado por primera vez, en Trinidad y Tobago en 1954, pertenece a la familia Togaviridae, género Alfavirus, el cual incluye a otros 29 virus, entre ellos los agentes causales de la encefalitis equina del este, la encefalitis equina venezolana, la encefalitis equina del oeste y la fiebre Chikungunya (1,2,13). El virus Mayaro es un arbovirus (Arthropod Borne Virus, dado que intervienen artrópodos en su ciclo de transmisión como vectores o transmisores) que se mantiene en la naturaleza en un ciclo que involucra principalmente a primates no humanos de vida silvestre y a mosquitos de la familia Culicidae (Diptera: Nematocera) de hábito selvático, en especial, especies pertenecientes al género Haemagogus.…”

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Virus Mayaro: un arbovirus reemergente en Venezuela y Latinoamérica

Múñoz

Navarro

2012

biomedica

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El virus Mayaro produce una enfermedad de síntomas inespecíficos y subletales, frecuentemente confundida con dengue, con síntomas de artralgias que pueden generar incapacidad laboral. Los brotes han sido esporádicos y localizados en la región selvática de la Panamazonia, posterior a su primer aislamiento en 1954 (Trinidad y Tobago). La información en la literatura científica es escasa, diversa y dispersa. El virus Mayaro es un alfavirus filogenéticamente relacionado con el complejo del virus Semliki Forest (SFV); con el UNA, son los únicos virus de dicho complejo aislados en el Nuevo mundo. Está conformado por ARN de cadena simple de carga positiva, con longitud de 12 kb con la región 42S y 26S, que codifican para proteínas no estructurales y estructurales. El virus Mayaro presenta gran adaptabilidad para la infección en vertebrados y especificidad hacia la familia Culicidae (mosquitos) como vectores u hospedadores invertebrados. Los factores de riesgo están asociados a zonas boscosas de la región septentrional de Suramérica y en temporada lluviosa. Se conocen dos genotipos, L (Belterra, Brasil) y D (distribución panamazónica). El ciclo enzoótico es similar al de la fiebre amarilla, que involucra a mosquitos del género Haemagogus y, como reservorios, a monos, sin descartar la participación de vectores secundarios y otros hospedadores que pudieran participar en la diseminación del virus. El humano puede presentar viremia elevada y se ha demostrado la transmisión experimental en Aedes aegypti, Ae. scapularis y Ae. albopictus, constituyendo un riesgo para la salud pública en centros urbanos y zonas rurales cercanas a focos de virus Mayaro.Palabras clave: Aedes, alfavirus, ecosistema amazónico, arbovirus, artralgia, filogenia, Culicidae, virus del bosque Semliki, Togaviridae Mayaro: A re-emerging Arbovirus in Venezuela and Latin AmericaMayaro virus produces nonspecific, sublethal disease symptoms, often confused with dengue, but with symptoms of arthalgias that can cause incapacitating disability. Outbreaks have been localized and sporadic in the Pan-Amazonia forest since its first isolation in 1954 (Trinidad and Tobago). The literature available is scarce, diverse and dispersed. Mayaro virus is an alphavirus, phylogenetically related to the Semliki forest antigenic complex. UNA and Mayaro viruses are the only viruses of this complex that have been isolated in the New World. Mayaro consists of single-stranded RNA of positive charge, length of 12 kb, subdivided into genomic and subgenomic regions, which encode nonstructural and structural proteins respectively. Mayaro shows a great plasticity in vertebrate host infection, whereas high specificity in the family Culicidae (mosquitoes). Risk factors of infection are associated with forest areas of northern South America and the rainy season. Two genotypes of MAYV have been identified, L (Belterra, Brazil) and D (widely distributed in the Pan-Amazonia). The enzootic cycle is similar to the jungle cycle of yellow fever, which involves Haemagogus mosquitoes and monkeys as ...

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