Venezuelan equine encephalitis (VEE) is a zoonotic disease caused by the Venezuelan equine encephalitis virus (VEEV) complex. This disease has not yet been reported in China, and it is therefore essential to establish a rapid and accurate method for detection of the virus in order to prevent and control this disease. In this study, a one-step real-time quantitative RT-PCR method was developed for the detection of the VEEV complex. A pair of specific primers and a Taqman probe were designed corresponding to a conserved region of the VEEV gene nspl, allowing the detection of all known strains of different sub- types of the virus. Using RNA synthesized by in vitro transcription as template, the sensitivity of this method was measured at 3.27 x 10(2) copies/microL. No signal was generated in response to RNA from Chikungunya virus (CHIKV), nor to RNA encoding the nsp1 fragment of Eastern equine encephalitis virus (EE-EV) or Western equine encephalitis virus (WEEV), all of which belong to the same genus as VEEV. This indicates that the method has excellent specificity. These results show that this one-step real-time quantitative RT-PCR method may provide an effective tool for the detection of VEEV in China.
China ; DNA Primers ; genetics ; Encephalitis Virus, Venezuelan Equine ; classification ; genetics ; isolation & purification ; Encephalomyelitis, Venezuelan Equine ; virology ; Humans ; RNA, Viral ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; methods

The rabies virus (RABV) is an enveloped RNA virus. It mainly damages the central nervous system and causes anencephaly in mammals and humans. There is now compelling evidence that enveloped virions released from infected cells can carry many host proteins, some of which may play an important part in viral replication. Several host proteins have been reported to be incorporated into RABV particles. However, a systematic study to reveal the proteomics of RABV particles has not been conducted. In the present study, after virus culture and purification by sucrose density gradient ultracentrifugation, a proteomics approach was used to analyze the protein composition of purified RABV particles to understand the molecular mechanisms of virus-cell interactions. Fifty host proteins, along with five virus-encoded structural proteins, were identified in purified RABV particles. These proteins could be classified into ten categories according to function: intracellular trafficking (14%), molecular chaperone (12%), cytoskeletal (24%), signal transduction (8%), transcription regulation (12%), calcium ion-binding (6%), enzyme binding (6%), metabolic process (2%), ubiquitin (2%) and other (14%). Of these, four proteins (beta-actin, p-tubulin, Cofilin, Hsc70) were validated by western blotting to be present in purified RABV particles. This novel study of the composition of host proteins in RABV particles may aid investigation of the mechanism of RABV replication.
Animals ; Humans ; Molecular Sequence Data ; Proteomics ; Rabies ; genetics ; metabolism ; virology ; Rabies virus ; chemistry ; genetics ; metabolism ; Viral Proteins ; analysis ; chemistry ; genetics ; metabolism ; Virion ; chemistry ; genetics ; metabolism