Efeito da tunicamicina sobre a replicação do vírus Mayaro em células de Aedes albopictus

Tunicamycin (TM), an inhibiting substance of bacteria, fungi and yeast proliferation, is an antibiotic isolated from Streptomyces lysosuperficus. This antibiotic inhibits the formation of N-acetylglucosamine, thereby it greatly alters the glycosilation of proteins. In this study, we found that the presence of TM causes a drastic inhibition of Mayaro virus replication in Aedes albopictus cells. Even at low concentration (0,05mg/ml), the treatment of the cells for 24 hours reduced in 50% the production of the viral infectious particles. At higher concentrations, the antibiotic reduced progressively the virus replication, resulting in 98,5% inhibition after a 48 hour treatment. The polyacrylamide gel electrophoresis analysis of [35S]-methionine-labelled proteins showed that TM at 0,05mg/ml, 0,1mg/ml e 0,15mg/ml concentration does not affect the synthesis of Mayaro virus polypeptides. Only at higher concentration (0,5mg/ml), the modification of the synthesis and of the electrophoretic mobility of viral proteins were noticed. These results suggest that, at low concentration, TM may affect Mayaro virus replication in Aedes albopictus cells through other mechanisms.Tunicamicina (TM) é um antibiótico isolado de Streptomyces lysosuperficus, caracterizado inicialmente como uma substância com atividade inibidora da proliferação de bactérias, fungos e leveduras. Este antibiótico possui um efeito bloqueador da formação de N -acetilglicosamina, alterando de maneira significativa o processo de glicosilação das proteínas. Neste trabalho, verificamos que, em células de Aedes albopictus, a presença de TM acarreta uma drástica inibição da replicação do vírus Mayaro. Mesmo em baixas concentrações de TM (0,05mg/ml), o tratamento das células durante 24 horas produz uma inibição de 50% na produção de partículas virais infecciosas. Em concentrações mais elevadas, este anti-biótico diminuiu progressivamente a replicação do vírus Mayaro, atingindo valores de 98,5% de inibição, após 48h de tratamento. A análise por eletroforese em gel de poliacrilamida das proteínas virais previamente marcadas com metionina-[35S] revela que a TM nas concentrações de 0,05 mg/ml, 0,1 mg/ml e 0,15 mg/ml não afeta a síntese dos polipeptídeos do vírus Mayaro. Apenas em concentrações mais elevadas (0,5mg/ml) observam-se alterações na síntese e na mobilidade eletroforética das glicoproteínas virais. Estes dados sugerem que, em baixas concentrações, a TM pode afetar a replicação do vírus Mayaro em células de Aedes albopictus através de outros mecanismos

Molecular Characterization and Phylogenetic Study of Coxsackievirus A24v Causing Outbreaks of Acute Hemorrhagic Conjunctivitis (AHC) in Brazil

Coxsackievirus A24 variant (CA24v) is the most prevalent viral pathogen associated with acute hemorrhagic conjunctivitis (AHC) outbreaks. Sixteen years after its first outbreak in Brazil, this agent reemerged in 2003 in Brazil, spread to nearly all states and caused outbreaks until 2005. In 2009, a new outbreak occurred in the northeast region of the country. In this study, we performed a viral isolation in cell culture and characterized clinical samples collected from patients presenting symptoms during the outbreak of 2005 in Vitória, Espírito Santo State (ES) and the outbreak of 2009 in Recife, Pernambuco State (PE). We also performed a phylogenetic analysis of worldwide strains and all meaningful Brazilian isolates since 2003.Sterile cotton swabs were used to collect eye discharges, and all 210 clinical samples were used to inoculate cell cultures. Cytopathic effects in HEp-2 cells were seen in 58 of 180 (32%) samples from Vitória and 3 of 30 (10%) samples from Recife. Phylogenetic analysis based on a fragment of the VP1 and 3C gene revealed that the CA24v causing outbreaks in Brazil during the years 2003, 2004 and 2005 evolved from Asian isolates that had caused the South Korean outbreak of AHC during the summer of 2002. However, the 2009 outbreak of AHC in Pernambuco was originated from the reintroduction of a new CA24v strain that was circulating during 2007 in Asia, where CA24v outbreaks has been continuously reported since 1970.This study is the first phylogenetic analysis of AHC outbreaks caused by CA24v in Brazil. The results showed that Asian strains of CA24v were responsible for the outbreaks since 1987 and were independently introduced to Brazil in 2003 and 2009. Phylogenetic analysis of complete VP1 gene is a useful tool for studying the epidemiology of enteroviruses associated with outbreaks

Prostaglandina A1 inibe a replicação do vírus Sindbis em células de rim de macaco e em células de mosquito

The present study reports the effect of prostaglandin A1 (PGA1) on the replication of Sindbis virus in monkey kidney and mosquito cells. In PGA1 treated cells we observed a severe reduction of virus yield. In both cells lines the highest nontoxic concentration of PGA1 (10 ìg/mL) decreased virus replication, dose dependently, by more than 90%. SDS-PAGE analysis of [35S] methionine labeled proteins showed that viral proteins (E1/E2 and C) were normally synthesized in PGA1 treated Vero cells, and induction of stress proteins (HSP70 and HSP90 ) was detected in uninfected and infected cells. In Vero cells the inhibition of virus replication was accompanied by a decrease in [3H] glucosamine incorporation into the virus glycoproteins.Neste estudo nós avaliamos o efeito da prostaglandina A1 (PGA1) na replicação do vírus Síndbis em células de macaco e em células de mosquito. Nas células tratadas com PGA1 nós observamos uma redução significativa na produção de partículas virais infecciosas. Em ambas as linhagens celulares tratadas com concentrações não-tóxicas de PGA1 (1-10 µg/ml) foi observado uma redução da replicação viral, de forma dose dependente, chegando a 90% na maior dose utilizada (10 µg/ml). A análise das proteínas virais e celulares marcadas com [35S]-metionina em SDS-PAGE mostrou que as proteínas virais (E1, E2 e C) foram normalmente sintetizas em células Vero tratadas com PGA1. Além disso, observamos também a indução das proteínas de estresse (HSP70 e HSP90) nas células Vero infectadas ou não infectadas. Em células Vero foi observado uma diminuição na incorporação de [3H] glicosamina nas glicoproteínas virais

Origin and number of CA24v isolates from Brazilian AHC outbreaks that were used for phylogenetic study.

<p>*Samples received previously isolated in cell culture.</p

Oligonucleotide primer sequences for VP1 and 3C amplification of CA24v.

<p>*Nucleotide position according CA24v reference strain EH24/70.</p

Environmental Monitoring for Enteroviruses in Maputo, Mozambique—2018

Due to the possibility of wild poliovirus importation from endemic regions and the high circulation of vaccine-derived poliovirus type 2 in the African region, Mozambique implemented a surveillance program to monitor the circulation of enteroviruses in the environment. From January to November 2018, a period that immediately preceded the cVDPV outbreak in Africa, 63 wastewater samples were collected from different areas in Maputo city. A total of 25 samples (39.7%) were positive based on cell culture isolation. Non-polio enteroviruses were found in 24 samples (24/25; 96%), whereas 1 Sabin-related poliovirus was isolated. Neither wild nor vaccine-derived poliovirus was detected. High circulation of EVB species was detected. Environmental surveillance in the One Health approach, if effectively applied as support to acute flaccid paralysis, can be a powerful aid to the public health system to monitor poliovirus besides non-polio enteroviruses in polio-free areas

Geographic distribution of CA24v outbreaks in Brazil, 2003–2009.

<p>Geographic distribution of CA24v outbreaks in Brazil, 2003–2009.</p

Phylogenetic analysis based on a 473-nucleotide region of the VP1 gene.

<p>This phylogenetic tree was constructed by the neighbor joining method using Kimura-two parameter model from MEGA4 software, with 1000 replicates. Thirty-seven sequences were compared to those strains reported from others countries in previous years available in GenBank. Only bootstrap values >70% are shown at the node. Geometric shapes indicate the years of the Brazilian isolates.</p

A RT-PCR method for selective amplification and phenotypic characterization of all three serotypes of Sabin-related polioviruses from viral mixtures

Outbreaks caused by vaccine-derived polioviruses are challenging the final eradication of paralytic poliomyelitis. Therefore, the surveillance of the acute flaccid paralysis cases based on poliovirus isolation and characterization remains an essential activity. Due to the use of trivalent oral poliovirus vaccine (OPV), mixtures containing more than one serotype of Sabin-related polioviruses are frequently isolated from clinical samples. Because each poliovirus isolate needs to be individually analyzed, we designed polymerase chain reaction primers that can selectively distinguish and amplify a genomic segment of the three Sabin-related poliovirus serotypes present in mixtures, thus, optimizing the diagnosis and providing prompt information to support epidemiologic actions