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

In this review, we mainly focus on zoonotic encephalitides caused by arthropod-borne viruses (arboviruses) of the families Flaviviridae (genus Flavivirus) and Togaviridae (genus Alphavirus) that are important in both humans and domestic animals. Specifically, we will focus on alphaviruses (Eastern equine encephalitis virus, Western equine encephalitis virus, Venezuelan equine encephalitis virus) and flaviviruses (Japanese encephalitis virus and West Nile virus). Most of these viruses were originally found in tropical regions such as Africa and South America or in some regions in Asia. However, they have dispersed widely and currently cause diseases around the world. Global warming, increasing urbanization and population size in tropical regions, faster transportation and rapid spread of arthropod vectors contribute in continuous spreading of arboviruses into new geographic areas causing reemerging or resurging diseases. Most of the reemerging arboviruses also have emerged as zoonotic disease agents and created major public health issues and disease epidemics.
Africa ; Alphavirus* ; Animals, Domestic ; Arboviruses* ; Arthropod Vectors ; Asia ; Encephalitis ; Encephalitis Virus, Venezuelan Equine ; Encephalitis Virus, Western Equine ; Encephalitis Viruses ; Encephalomyelitis, Equine ; Epidemiology* ; Flaviviridae ; Flavivirus* ; Global Warming ; Humans ; Population Density ; Public Health ; South America ; Togaviridae ; Transportation ; Urbanization ; Zoonoses

OBJECTIVETo develop a rapid, cost effective RT-PCR method for the mass scale diagnosis of such diseases at the viremia stage to find out the actual disease burden in that area.

METHODSFor this purpose, cases with the history of only short febrile illness were considered. Thus 157 samples with the history of dengue/chikungunya like illness and only 58 samples with a history of acute encephalitis syndrome (AES) were selected.

RESULTSOut of 157 samples, 42 and 74 were detected as dengue and chikungunya, respectively and out of 58 AES cases only 23 could be detected as Japanese encephalitis by this RT-PCR method.

CONCLUSIONSThis cost effective RT-PCR method can detect the total positive cases that remain undetected by ELISA method. Moreover, this method is capable to detect the viral RNA from patients' sera even after the appearance of IgM antibody at one fifth costs as compared with the other commercially available kits.

Antibodies, Viral ; blood ; Arbovirus Infections ; diagnosis ; virology ; Arboviruses ; genetics ; Chikungunya Fever ; diagnosis ; virology ; Dengue ; diagnosis ; virology ; Encephalitis Virus, Japanese ; genetics ; Encephalitis, Japanese ; diagnosis ; virology ; Fever ; diagnosis ; virology ; Humans ; Immunoglobulin M ; blood ; Mass Screening ; RNA, Viral ; blood ; Reverse Transcriptase Polymerase Chain Reaction ; economics ; methods ; Sensitivity and Specificity ; Viremia ; diagnosis ; virology

Child ; China ; epidemiology ; Encephalitis Virus, Japanese ; Encephalitis, Viral ; epidemiology ; prevention & control ; Female ; Humans ; Japanese Encephalitis Vaccines ; immunology ; Male ; Vaccination

Japanese encephalitis is a potentially lethal disease of the central nervous system caused by infection with Japanese encephalitis virus (JEV). JEV is the most common cause of encephalitis over a large part of eastern Asia. To establish and characterize in vivo model to study the Japanese encephalitis, the immunohistochemical localization of JEV and the histopathological finding were investigated in the brains of young adult mice infected with JEV by intraperitoneal inoculation. JEV was localized to neurons in discrete regions of the brain. Histopathological finding showed typical pattern of acute viral encephalitis, such as inflammatory cell infiltration in brain parenchyme and perivascular cuffs of mononuclear cells. These results suggest that this in vivo system can be used to study the mechanism of virus entry into the brain, cell specific tropism, and pathophysiology in Japanese encephalitis.
Animals ; Asian Continental Ancestry Group* ; Brain* ; Central Nervous System ; Encephalitis ; Encephalitis Virus, Japanese ; Encephalitis, Japanese* ; Encephalitis, Viral ; Far East ; Humans ; Immunohistochemistry ; Mice* ; Neurons ; Tropism ; Virus Internalization ; Young Adult

Japanese encephalitis has been the leading cause of epidemic viral encephalitis in Korea. In 2010, 26 patients with Japanese encephalitis were documented in Korea. Patients older than 50 years accounted for 54% of this cohort, and none of the patients was younger than 10 years. The number of documented cases of Japanese encephalitis increased markedly in 2010 compared to the annual reported cases during the previous 10 years.
Asian Continental Ancestry Group ; Cohort Studies ; Encephalitis, Japanese ; Encephalitis, Viral ; Humans ; Korea

OBJECTIVETo detect the relations between incidence rate of the epidemical encephalitis B and related factors, to provide a simple, valid and practical new method for forecasting encephalitis B eipdemics.

METHODSConnection number between the incidence rate of encephalitis B and the historical forecast factors was computed, before ranking the first, second and the third principal factor, to remove the factor with the smallest value in the light of the connection number before comparing the newest value of forecast factors with the same kind of history while the most nearly value becoming the forecasting factor value and to establish a forecasting equation according to the factor value and the consistent degree of the incidence rate of encephalitis B at that time. Finally, to put into the new factor value to get this forecast value under this equation. Assuming that there are n' (n' >or= 2) forecast factors, this time forecast value can then be directly obtained from the average of these estimate values.

RESULTSUsing above forecast method to forecast the incidence rate of encephalitis B at certain place and year, the predicting value is very much close to the actual incidence rate. Difference between the predicting value forecasted by the above-mentioned method and the actual incidence rate is only 0.0264/100 000 with an accurate rate of 97.94%.

CONCLUSIONThis principal factor analysis forecast method based on connection number in forecasting the incidence rate of encephalitis B prevention is acceptable.

China ; epidemiology ; Encephalitis Virus, Japanese ; Encephalitis, Viral ; epidemiology ; virology ; Factor Analysis, Statistical ; Forecasting ; Humans ; Incidence

OBJECTIVETo investigate the molecular basis of pathogenicity of Japanese encephalitis virus (JEV) by sequencing of complete nucleotide sequence and analyze the characteristics of full-length genome of genotype I Japanese encephalitis virus strains (GZ56) which was isolated from the first cerebrospinal fluid (CSF) of Japanese encephalitis patients.

METHODSThe complete nucleotide sequence was obtained by RT-PCR and sequencing was performed directly. Bioinformatics was used to analyze the nucleic acid data, deduced amino acid sequence and phylogenetic trees.

RESULTSThe result of sequence analysis showed that the genome of GZ56 strains had 10 965 nucleotides, which coded for a 3432-amino acid polyprotein. Phyolngenetic analysis based on full-length genome showed that GZ56 strains and M-28 strains which were the first isolated from mosquitoes in Yunnan in 1977 were in the same evolutionary branch. GZ56 strains belongs to genotype I of Japanese encephalitis virus, the homology of genome ranged from 96.2% to 98.6% in nucleotide and from 98.2% to 99.7% in amino acid sequences respectively when compared with selected genotype I of JEV strains in GenBank. There were 11 amino acid divergences in E protein when compared with the JEV inactivated P3 strain but they are not the key virulence sites. However, there were 14 amino acid divergences in E protein when compared with the JEV live attenuated vaccine SA14-14-2 strain and 8 amino acid divergences were the key virulence sites.

CONCLUSIONThis study indicated that the full length of genome GZ56 strains had no ignificant change. It can be hypothesized from genomic level that the currently available JEV vaccines(inactivated and live attenuated) can protect against GZ56 strains infection, meanwhile, the JEV live attenuated vaccine (SA14-14-2) formulation conferred higher levels of protection.

Computational Biology ; Encephalitis Virus, Japanese ; classification ; genetics ; isolation & purification ; Encephalitis, Japanese ; virology ; Enzyme-Linked Immunosorbent Assay ; Genome, Viral ; Genotype ; Japanese Encephalitis Vaccines ; immunology ; Phylogeny ; Sequence Analysis, DNA

OBJECTIVESequence and analysis the complete nucleotide of the Japanese encephalitis virus (JEV) newly strain SD08-10, isolated in 2008 in Shandong, China in order to understand the characterization of the virus.

METHODSOverlapping primers were designed according to the full-length genomes in GenBank. RT-PCR was used to amplify the fragments and the full-length genome was obtained by sequencing and splicing. Using the computer software to analysis the nucleic acid data, deduced amino acid sequence and phylogenetic tree, including Clustal X (1.8), DNASTAR, GENEDOC (3.2).

RESULTSThe result of sequence analysis shows that the genome of SD08-10 strain was 10 965 nucleotides long. An open reading frame from 96 to 10 392 including 10 296 nucleotides is capable of coding for a 3432 amino acid polyprotein. Compared with the live attenuated vaccine strain SA-14-14-2 in China, there was 1253 nucleotide difference and 82 amino acid divergence. Comparison of the complete genome sequences with 59 different JEV isolates showed a 0.7%-18.9% nucleotide sequence divergence among them, which resulted in 0.1%-5.2% amino acid sequence divergence. Phylogenetic analysis of full-length genome showed that the SD08-10 strain was belonging to genotype I.

CONCLUSIONAnalysis based on the complete genome sequences of different JEV isolates showed that the SD08-10 strain isolated in 2008 in Shandong was belonging to genotype I and close to SH17M-07 isolated in 2007 in China.

China ; Encephalitis Virus, Japanese ; classification ; genetics ; isolation & purification ; Encephalitis, Japanese ; virology ; Genome, Viral ; Molecular Sequence Data ; Phylogeny

The Japanese encephalitis virus (JEV), a member of the Flaviviridae family and Flavivirus genus, is transmitted by mosquitoes. JEV, of which some 35,000 cases are recorded every year, is a positive RNA virus. Two types of JEV vaccines have been developed to prevent the onset of encephalitis in humans, namely formalin-inactivated and liveattenuated vaccines. JEV inactivated vaccines are usually made using the Nakayama-NIH or Beijing-1 strains of the JEV virus. In this study, the immunological response to the Nakayama-NIH and Beijing-1 strains was analyzed as part of the effort to compile basic data which could lead to the selection of a suitable vaccine strain. To this end, the virus titer of Beijing-1 was found to be two-fold higher than that of Nakayama-NIH by plaque assay. Moreover, Beijing-1-induced neutralizing antibodies showed a higher level of titers when confronted by Korean JEV isolates than Nakayama-NIH-induced neutralizing antibodies (1:320 vs. 1:160, respectively). However, as a minimum ratio of 1:10 neutralizing antibody titers are required to protect against JEV infection, both strains in effect exhibited a sufficient level of neutralizing antibody titers. What's more, Beijing-1 was found to induce a somewhat higher cytotoxic T lymphocyte (CTL) response than Nakayama-NIH. Taken together, this can be taken to mean that Beijing-1 may in fact be a more effective vaccine candidate strain when it comes to inducing a high level of protective immunity against JEV infection.
Antibodies, Neutralizing* ; Asian Continental Ancestry Group* ; Culicidae ; Encephalitis ; Encephalitis Virus, Japanese* ; Encephalitis, Japanese* ; Flaviviridae ; Flavivirus ; Humans ; Lymphocytes* ; RNA Viruses ; Vaccines ; Vaccines, Inactivated ; Viral Load