This research firstly reported the molecular analysis of ECHO25 (Entric Cytopathic Human Orphanviruses Type 25). To clarify molecular characteristics of the ECHO25 virus isolates in Henan Province and its relationship with the rest of world's isolates,the complete VP1 sequences of the 4 isolates in Henan were successfully amplified by RT-PCR and were compared with other ECHO25 isolates available from GenBank. Compared with the prototype strain JV-4, the nucleotide sequence identity was 79.2%-80.1%, and the amino sequence identity was 89.0%-92.4, the nucleotide sequence identity among the 4 strains isolates in Henan Province was 93.0%-99.0%, the amino sequence identity was 92.4%-97.5%. HN-01 and HN-26 strains had the highest level of homology, the nucleotide homology was 99.0%; All the 4 strains belonged to the B1 genotype.
China ; epidemiology ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Enterovirus Infections ; epidemiology ; virology ; Humans ; Molecular Sequence Data ; Phylogeny

This report presents an overview of human enterovirus B species in Henan Province. A total of 14 isolates of HEV-B species isolated under HFMD surveillance network during the six months in 2010 were examined. Based on molecular typing results targeting VP1 region, 14 isolates were classified into 6 serotypes of HEV-B. Furthermore, comparison of these 14 isolates with reference strains and strains in mainland China was conducted. The phylogenetic analysis revealed that E25, E11 and E6 showed homology with those from Shandong Province which adjoins Henan Province. E1 and E13 showed homology with those from Yunnan Province, and E30 showed homology with Henan strain isolated in 2008. Cocirculation of two lineages of echovirus 6 was observed.
China ; epidemiology ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Enterovirus Infections ; epidemiology ; virology ; Feces ; virology ; Humans ; Molecular Sequence Data ; Phylogeny

We wished to analyze the genetic characterization of echovirus 11 (Echo11) from samples of environmental sewage in Shandong Province (China). The VP1 coding region was typed as the strains were amplified. Phylogenetic analyses on the VP1 sequences from these isolates, strains isolated from AFP cases in the period 1994-2010 and others published in GenBank were conducted. From 2011 to 2012, 94 Echo11 strains were isolated from samples of environmental sewage in Jinan and Linyi City in Shandong Province. Numbers of Echo11 were seasonal and reached peaks in the summer and autumn in both cities; A- mong these isolates, nucleotide (nt) identities were 89.5%-100.0% whereas amino acid (aa) identities were 95.4%-100.0%. The nt and aa identities were 76.6%-79.7% and 90.4%-92.5% between those strains and the prototype (Gregory) strain of Echo11, respectively. All isolates from Shandong Province were the A genotype and the strains evolved very rapidly, which suggested that several transmission chains was co-circulating. We described the temporal fluctuation and genetic characterization of Echo11 isolates from surveillance of environmental sewage in Shandong Province, thereby providing important information for exploring the dynamic change and genetic variation of circulating human enteroviruses in this Province in China.
China ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Phylogeny ; Sewage ; virology

<b>OBJECTIVEb>To analyze the genetic characterization of the complete genome from a human coxsackievirus B3 strain A103/KM/09 isolated in Yunnan province, 2009.

<b>METHODSb>By using RT-PCR, all the eight fragments which containing about 1000 nucleotides and covering full viral genome, were sequenced. By using Mega 5.05,Geneious, RDP 3 and SimPlot 3.5.1 software, sequences were aligned with other enterovirus reference sequences. Phylogenetic and recombination analysis were also carried out.

<b>RESULTSb>The A103/KM/09 isolate genome showed 7389 nucleotides in length , encoding for 2185 amino acids. In the complete genome, the homology of nucleotide and amino acid among the seven coxsackievirus B3 isolates were 81.0%-88.0% and 95.7%-98.0%, respectively. There appeared 81.0% and 95.7% homology when compared with that of Nancy prototype strain. Results from the Phylogenetic analysis showed that the coxsackievirus B3 formed five distinct clades, I-V. Nucleotide divergence rates between clades were 16.2%-24.3% . The A103/KM/09 strain belonged to clade V. Clade V was further divided into four sub-clades,A-D. The nucleotide divergence between sub-clades was 4.3%-11.4%. Putative recombinant event for A103/ KM/09 was detected.

<b>CONCLUSIONb>All coxsackievirus B3 isolates could be divided into five clades, with A103/KM/09 strain belonged to Clade V-D. Evolution of coxsackievirus B3 had occurred in China.

Base Sequence ; Child, Preschool ; China ; epidemiology ; Encephalitis, Viral ; epidemiology ; virology ; Enterovirus B, Human ; genetics ; isolation & purification ; Enterovirus Infections ; epidemiology ; virology ; Genome, Viral ; Humans ; Male ; Phylogeny ; Viral Proteins ; genetics

<b>OBJECTIVEb>To characterize the complete genome sequence of coxsackievirus B5 (CVB5)A210/KM/09 strain which was isolated from Yunnan, China, 2009.

<b>METHODSb>Eight overlapping clones covering the whole viral genome (excluding the poly-A tail)were obtained by RT-PCR and sequenced, with their nucleotide and amino acid sequences compared with other known CVB5 strains.

<b>RESULTSb>The genome of the CVB5 A210/KM/09 strain had 7 372 nucleotides in length, and containing a 742-nt non-translated region (NTR) at the 5' end and a 98-nt NTR at the 3' end. The entire open reading frame contained 6 555 nt, encoding a 2 185-aa polyprotein. In the coding region, there appeared no nucleotide deletion or insertion, but some changes of amino acid seemed unique. Based on the complete genome sequence alignments, CVB5 isolate A210/KM/09 strain showed the highest nucleotide (92.5%) and amino acid (97.3%) identities to the CVB5/CC10/10. It also shared nucleotide (80.1%-92.5%) and amino acid (95.0%-97.3%) homology with other CVB5 strains: 17Y, 19CSF, 20CSF, 1954/85/US, 2000/CSF/KOR, 03001N, CoxB5/Henan/2010, VB5/SD/09 and Faulkner. Blast between genome fragments, A210/KM/09 showed similarity on nucleotide (80.1%-92.5%) and amino acid (95.0%-97.3%) identities with other CVB5 strains. The phylogenetic tree, constructed on the complete VP1 regions, indicated that CVB5 could be divided into genotype A, B, C and D. while Genotype C and D could be further divided into C1-C4 and D1-D4 subgenotypes.

<b>CONCLUSIONb>A210/KM/09 and other CVB5 predominant strains isolated in China belonged to CVB5 subgenotype C4.

Child, Preschool ; China ; epidemiology ; Encephalitis, Viral ; epidemiology ; virology ; Enterovirus B, Human ; genetics ; isolation & purification ; Female ; Genome, Viral ; Genotype ; Humans

<b>OBJECTIVEb>To analyze the complete genome sequences of two coxsackievirus B5 (CVB5) isolated in Henan province, 2011.

<b>METHODSb>Specimens were collected from viral encephalitis patients and followed by viral isolation on them. RNA were extracted from positive isolates and the amplified products were sequenced. The full-length genomes of them were acquired by assembling the fragments, using DNAStar 5.01 software while phylogenetic analysis were performed with Mega 5.05 and other software.

<b>RESULTSb>The genomes RNA of 03001N and 17Y showed 7408 bp and 7404 bp long, and the 5'- and 3'-untranslated regions were 747 bp, 743 bp and 103 bp, 103 bp, respectively. BLAST analysis of these two isolates, based on the complete genome, showed 97% identity, with both of them having the highest similarity(98%, 99%)to the CVB5 strain isolated from Henan in 2010 rather than other CVB5 strains. Coding regions of both isolates were 6558 bp, code for a polyprotein of 2185 amino acids (aa) and both of them showed 99% amino acid identity. Phylogenetic tree in VP1 region showed that the two isolates belonged to the same clade with other strains isolated from all over the country in the past years, except for some CVB5 strains isolated from Henan and Shandong province in 2009 that formed the other cluster.

<b>CONCLUSIONb>It seemed that more than one group of CVB5 were circulating in Henan province and these two isolates appeared the main epidemic strains circulating in the past years.

Base Sequence ; China ; epidemiology ; Encephalitis, Viral ; epidemiology ; virology ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Genes, Viral ; Humans ; Phylogeny

<b>OBJECTIVEb>To investigate the etiology of the outbreak of viral encephalitis in Jinan area in 2003.

<b>METHODSb>Virus-specific nucleic acid fragments were amplified by random PCR and RT-PCR using specific primers to enterovirus. After sequencing, the gene sequence was handled by the program BLAST for homologous analysis and the software Clustal W 1.82 for multiple sequence alignment to identify the etiology and its genotype.

<b>RESULTSb>Five strains were isolated from clinical specimens. A gene fragment for one strain was acquired using random PCR, which was highly homologous to enterovirus. Then, the 5' non-translated region and partial VP1 region were amplified and sequenced. The five isolated strains were all identified as Coxsackievirus B5, and what was more, they were most homologous to the strain isolated during the outbreak of aseptic meningitis and encephalitis in Zhejiang province from 2002 to 2004.

<b>CONCLUSIONb>Coxsackievirus B5 is closely associated with the outbreak of viral encephalitis in Jinan area in 2003. It is an important etiology but other viruses may also played a role which remains to be clarified.

China ; epidemiology ; Disease Outbreaks ; Encephalitis, Viral ; virology ; Enterovirus B, Human ; genetics ; isolation & purification ; Humans ; Polymerase Chain Reaction

<b>OBJECTIVEb>To develop a method for detection of coxsackie B virus type 1-6 by RT-PCR.

<b>METHODSb>A pair of primers were designed to amplify all types of coxsackie B virus 1-6 efficiently. The PCR product was hybridized in micro-wells in which 6 type specific oligonucleotide probes had been coated respectively, colorimetric detection was performed to discriminate the types of coxsackie B virus.

<b>RESULTSb>This method was shown to be concordant with the IgM ELISA, 71.7% of anti-coxsackie B positive cases could be detected by RT-PCR.

<b>CONCLUSIONb>The RT-PCR method can type coxsackie B virus efficiently and provides a tool for clinical diagnosis and epidemiological investigation.

DNA Primers ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Enterovirus Infections ; diagnosis ; virology ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunoglobulin M ; blood ; Reverse Transcriptase Polymerase Chain Reaction ; methods

An outbreak of hand foot and mouth disease occurred in Shang dong, China in 2009. Almost 20% of patient's swabs was positive for Coxsackie virus B5 (CVB5) identified by RT-PCR and sequencing. It was suggested that CVB5 may be another important pathogen for HFMD. Fifteen pairs of overlapping primers were designed and the genome sequence was sequenced. The genome of CVB5 was 7 399 nt in length, coding for 2 185aa. The genome displayed 80.6%-85.3% nucleotide sequence identity and 96.1%-96.9% amino acid sequence identity with another three CVB5 respectively. Phylogenetic analysis showed that different segment of genome underwent a distinct evolutionary and selective pressure. Simplot analysis displayed no evident recombination between genome of CVB5 and other HEV B viruses. The complete and characterized genome of CVB5/09 provides further insight into the genetics of CVB5 and other HEV B viruses, aiding in the surveillance and control of HFMD.
Base Sequence ; China ; Coxsackievirus Infections ; virology ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Genome, Viral ; Humans ; Molecular Sequence Data ; Phylogeny ; Viral Proteins ; genetics

<b>OBJECTIVEb>To determine the partial sequence of virus strains causing an aseptic meningitis outbreak in northern part of Jiangsu province in 2003 and to compare them with the same serotype strains isolated in other countries to better understand its genetic characteristics and hereditary trend of development.

<b>METHODSb>Virus RNA was amplified using two sets of specific enteroviral 3' half of VP1 primers 012/011 and 040/011. Polymerase chain reaction (PCR) products were purified and sequenced. BLAST program was then used to perform on nucleotide and amino acid pairwise-alignment with all available sequences in NCBI database. Phylogenetic trees were drawed to compare with other enteroviral sequences using PHYLIP software.

<b>RESULTSb>Under BLAST program, three sequences we submitted to GenBank were identically inferred as echovirus type 30, which had been identified by neutralization test in previous study. Phylogenetic analysis demonstrated that strains isolated from this outbreak were aggregated into a cluster, and the closest relationships with them were those isolated in 1999 and 2000. This phenomenon indicated that Echo30 from this outbreak was different from other strains in different epidemic area.

<b>CONCLUSIONb>3' half of VP1 sequence could be used to quickly identify the serotype of isolated enterovirus. Strains isolated from this outbreak had the similar hereditary developing trend comparing with Echo30 strains isolated from other countries.

Base Sequence ; China ; epidemiology ; DNA, Viral ; genetics ; Disease Outbreaks ; Enterovirus B, Human ; genetics ; isolation & purification ; Female ; Humans ; Male ; Meningitis, Aseptic ; epidemiology ; virology ; Molecular Sequence Data ; Sequence Analysis, DNA