ABSTRACT
To study on the molecular evolution of Coxsackie virus A16 (CVA16)isolated from clinical speci-mens of Hand, foot and mouth Disease( HFMD) patients in Inner Mongolia in 2010. A total of 921 clinical specimens including stools, throat swabs and vesicle fluids were collected from 888 HFMD patients in out-patient service in Inner Mongolia and viral isolation was then performed, the positive viral isolates were identified by using the real-time PCR method detecting CVA16. A total of 50 CVA16 isolates were selected from the patients presenting mild symptoms, severe symptoms and the death patients randomly, and the VP1 coding regions of representative CVA16 isolates were amplified and sequenced. Finally the phylogenetic tree was constructed among the VP1 coding regions of the different genotypes and subgenotypes of CVA16 strains. Eighty two viruses were isolated form 921 clinical specimens, the positive rate was 8. 90%, of which 3 viruses were isolated from severe cases and 1 viruses was from death cases. The nucleotide acid of 50 representative CVA16 strains in Inner Mongolia were closed to CVA16 strains isolated from mainland China since 1998, especially from Beijing in 2009 and from Henan in 2010, the identity were 96. 18% approximately 98. 88% and 94. 94a approximately 98. 76%, respectively. There was a little difference in the nucleotide acid between the CVA16 strains from Inner Mongolia in 2010 and in 2007, the identity were 91. 68% approximately 96. 52% The phylogenetic tree showed that all CVA16 strains clustered within Bla and B1b evolution branch of B1 genotype. There was slight difference in the nucleotide and the amino acid in VP1 region among the 50 Inner Mongolia CVA16 strains, the identity were 89. 99% approximately 100% and 98. 31% approximately 100%, respectively, indicating that these strains belonged to many different viral transmission chains. The CVA16 strains circulated in Inner Mongolia in 2010 were all belong to B1a and B1b evolution branch of B1 genotype, and the two evolutionary branchs of Coxsackie virus A16 were co-evolved and co-prevailed in Inner Mongolia Autonomous Region.
Subject(s)
Adolescent , Adult , Animals , Child , Child, Preschool , Female , Humans , Infant , Male , Young Adult , Capsid Proteins , Genetics , Cell Line, Tumor , Chlorocebus aethiops , China , Epidemiology , Coxsackievirus Infections , Epidemiology , Mortality , Virology , Enterovirus , Classification , Genetics , Evolution, Molecular , Feces , Virology , Genotype , Hand, Foot and Mouth Disease , Epidemiology , Mortality , Virology , Phylogeny , RNA, Viral , Genetics , Sequence Analysis, DNA , Vero CellsABSTRACT
Strain TB-Chen is a group A rotavirus (RV) isolated from a Chinese infant suffering from gastroenteritis in hospital. The NSP5 and NSP6 of strain TB-Chen are encoded by the 10th gene segment (816bp in whole length) of the viral genome. The results obtained in this study showed that the NSP5 was encoded in the first open-reading-frame (ORF) of the gene segment (from 22bp to 624bp), and NSP6 was encoded in the second ORF (from 80bp to 355bp). The NSP5 protein consisted of 200 amino acid residues with a putative molecular mass of 21.9 kD, and a putative isoelectric point of 7.86. The NSP6 protein consisted of 92 amino acids with a putative molecular mass of 11 kD, and a putative isoelectric point of 9.65. This study further analyzed phylogenetic relationship of the NSP5/NSP6 ORF nucleotide sequence. The results showed that the NSP5s of group A rotavirus could be at least classified into 7 genotypes (H1-H7), the NSP6s could be at least classified into 8 genotypes (hl-h8); the genotypes of the NSP5 and NSP6 derived from strain TB-Chen was classified as H2 and h2. This was the first report on the genotype classification of the NSP6 of group A RVs, and it was proposed English letter "h" to represent genotype of the NSP6, e. g. strain 69M classified as H7h7, strains Wa and KU classified as H1h8.
Subject(s)
Humans , Evolution, Molecular , Gastroenteritis , Virology , Genotype , Open Reading Frames , Genetics , Phylogeny , Reverse Transcriptase Polymerase Chain Reaction , Rotavirus , Classification , Genetics , Viral Nonstructural Proteins , GeneticsABSTRACT
Little is known about the non-structural protein 6(NSP6)of rotavirus.This report describes expression of the NSP6 of a group A human rotavirus strain TB-Chen in bacteria,and its immunological properties and cellular distribution.The results showed that the recombinant NSP6(rNSP6)was expressed in high efficiency without any other proteins fused(possesses about 34.2% of total bacterial proteins).rNSP6 elicited mono-specific antibodies in immunized guinea pigs and the antibodies could react with the rNSP6 itself and the viral NSP6 proteins synthesized in SA11-or Wa-infected MA104 cells in Western blot and immunofluorescence assay.The NSP6 distributed evenly in the cytoplasm mainly around the nucleus of virus-infected cells,no viroplasm-like gathering observed;The top amount of NSP6 synthesized in SA11-infected cells or Wa-infected cells could be detected at 12h after infection.This is the first report about the high expression of entire NSP6(without any other proteins fused)in prokaryotic expression system and detection of NSP6 synthesis in virus infected cells by immunofluorescence assay.The results are important to understand the structure,biological properties and further application of the NSP6.
