Chikungunya virus ; genetics ; isolation & purification ; China ; Genome, Viral ; RNA, Viral ; genetics ; Sequence Analysis, RNA

OBJECTIVETo investigate the viral contamination of invasive medical instruments in dentistry and to provide health administrative institutions with surveillance data.

METHODSSterilized samples were randomly collected from the department of dentistry to detect HBV-DNA, HCV-RNA, HIV-RNA and HBsAg.

RESULTSOf the invasive medical instruments that were sterilized with 2% glutaraldehyde, one of the samples was positive for HBV-DNA, and another sample was positive for HBsAg.

CONCLUSIONThough massive virus contamination of invasive medical instruments in dentistry has been reduced to a low level, the occurrence of contamination still remains.

DNA, Viral ; analysis ; Dental Instruments ; virology ; Equipment Contamination ; HIV ; isolation & purification ; Hepacivirus ; isolation & purification ; Hepatitis B Surface Antigens ; analysis ; Hepatitis B virus ; isolation & purification ; Humans ; RNA, Viral ; analysis

OBJECTIVETo monitor and discover medically important mosquito-borne viruses circulating in Xinjiang, China.

METHODSMosquitoes were collected from Abbey Lake wetland in Bortala, in Northern Xinjiang. Viral isolates were obtained through inoculating and serial passaging into susceptible mammalian host cells (BHK-21), identified by cytopathogenic effect (CPE) observation and plague forming assay. Genetic identification of viral isolates was conducted by RT-PCR, sequencing and phylogenetic analysis.

RESULTSA virus strain which causing CPE on BHK-21 cells, was isolated from the predominant Culex modestus (36.6%) and tentatively designated as Abbey Lake virus. Information on molecular identification revealed that Abbey Lake virus belonged to Orthobunyavirus genus within Bunyaviridae. Partial sequences (651 bp and 980 bp) of viral genomic S and M segment showed that Abbey Lake virus was phylogenetically related to Germiston virus that uniquely found in South Africa with 90.6% nucleotides and 95.0% amino acids similarities in S segment. However, viral M segment displayed much variability with 78.6% nucleotides and 86.1% amino acid similarities, suggesting a new member of Orthobunyavirus genus was discovered in the area.

CONCLUSIONIn this study, Abbey Lake virus was isolated and characterized indicating its potential circulation nature of this newly-emerged mosquito-borne virus.

Animals ; China ; Culicidae ; virology ; Orthobunyavirus ; classification ; genetics ; isolation & purification ; Phylogeny ; Sequence Analysis, RNA ; Viral Proteins ; genetics

OBJECTIVE: Tick-borne encephalitis virus (TBEV) is an emerging pathogen in Europe and North Asia that causes tick-borne encephalitis (TBE). A simple, rapid method for detecting TBEV RNA is needed to control this disease. METHODS: A reverse-transcription recombinase-aided amplification (RT-RAA) assay was developed. This assay can be completed in one closed tube at 39 °C within 30 minutes. The sensitivity and specificity of RT-RAA were validated using non-infectious synthetic RNA representing a fragment of the NS5 region of the wild-type (WT) TBEV genome and the Senzhang strain. Additionally, 10 batches of tick samples were used to evaluate the performance of the RT-RAA assay. RESULTS: The analytical limit of detection of the assay was 20 copies per reaction of the TBEV synthetic transcript and 3 plaque-forming units (pfu) per reaction of TBEV titers. With the specific assay, no signal due to other arboviruses was observed. Of the 10 batches of tick samples obtained from the Changbai Mountains of China, three were TBEV-positive, which was consistent with the results of the quantitative real-time PCR assay. CONCLUSION A rapid, highly sensitive, specific, and easy-to-use method was developed for the detection of the TBEV Far-Eastern subtype.
Encephalitis Viruses, Tick-Borne ; genetics ; isolation & purification ; Nucleic Acid Amplification Techniques ; RNA, Viral ; analysis

OBJECTIVETo compare the molecular characteristics of 3 Crimean-Congo hemorrhagic fever viruses(CCHFV) isolated in Xinjiang province.

METHODSYL05035, YT05099 and LT05146 were isolated in 2005 from Hyalomma ticks and viral RNA was extracted from suckling mouse brains infected with these three strains respectively. The polymerase chain reaction(PCR) products of S segments from the 3 strains of CCHFV were directly sequenced.

RESULTSThe full-length'S RNA from the 3 strains of CCHFV all comprised 1673 nucleotides with ORF of them including 1449 nucleotides and encoding a protein which comprised 482 amino acids in a viral complementary sense. The sequences indicated that the three strains of CCHFV isolated from ticks in Xinjiang province were highly homologenic. Data from the phylogenetic analysis showed that the obtained sequences were identical. The homology between 3 strains of CCHFV was 99.5%. Their homologies compared with that of the other strains isolated from other region of Xinjiang were also high at nucleotide levels (92.7%-99.8%). The three strains which were clustered together with 7001 strain and 79121 strain (isolated from patient and rat in Xinjiang respectively) was only different by 2%-3%. The genetic difference from the prototype CCHFV Nigerian strain IBAR10200 was 13%. In comparison, the Nigerian CCHFV tick isolate was more divergent when compared with the reference China strains 66019 and with the three variants mentioned above.

CONCLUSIONThe CCHFV isolated from China comprised a group of genetically high conserved strains.

Animals ; Brain ; virology ; China ; Genes, Viral ; Hemorrhagic Fever Virus, Crimean-Congo ; genetics ; isolation & purification ; Mice ; RNA, Viral ; analysis

BACKGROUNDTo design and establish a method of multiplex PCR normalization and parallel detection of HBV and HCV.

METHODSUsing two pairs of primers, one inner and the other outer, each having a 20 bp common sequence, the authors amplified target DNA for two rounds. All products would have this common sequence. Using this common sequence as primer the authors performed further amplification. Finally, multiplex PCR was normalized to a single PCR style and eliminated multiple factors disturbance. Four kinds of nucleic acid extraction method were compared. Multiplex PCR normalization was established and optimized by using orthogonal design by analysing 6 kinds of key factors. The method was evaluated by detecting 28 samples of HBV and HCV.

RESULTSThe sensitivity, specificity, diagnostic idex and efficiency for HBsAg, HCV antibody positive patients were 83.3%, 70.0%, 153.3%, 72.2% respectively. The sensitivity, specificity, idex and efficiency for HBsAg positive patients were 78.6%, 80.0%, 158.6%, and 79.2%, respectively. The sensitivity, specificity, idex and efficiency for HCV antibody positive patients were 75.0%, 90.0%, 165.0% and 83.3% respectively.

CONCLUSIONSThe multiplex PCR normalization method may have potential applicability in parallel amplification of multiple genes of pathogens.

DNA, Viral ; analysis ; Hepacivirus ; isolation & purification ; Hepatitis B ; virology ; Hepatitis B virus ; isolation & purification ; Hepatitis C ; virology ; Humans ; Polymerase Chain Reaction ; methods ; RNA, Viral ; analysis ; Sensitivity and Specificity ; Superinfection

OBJECTIVETo study the genetic characterization of wild-type measles viruses isolated in Xinjiang in 2003 and 2004.

METHODSPeripheral blood mononuclear cells (PBMC) from 19 suspected measles cases collected between June 2003 and April 2004 were used to isolate measles virus by cocultivation with phytohemagglutinin (PHA)-stimulated cord blood mononuclear cells (CBMC). For positive samples, 676 nucleotides of the C-terminus of the nucleoprotein (N) gene of the measles virus genome were amplified by reverse transcription-polymerase chain reaction and then sequenced. These sequences were compared with those of other measles reference strains available in GenBank or measles isolates elsewhere in China using BLAST searches and phylogenetic analyses.

RESULTS6 measles virus strains were isolated with 3 strains (XJ03-26, XJ03-27, XJ03-74) from 2003 and 3 (XJ04-146, XJ04-150, XJ04-152) from 2004. The strain XJ03-26, differed from the Chinese measles vaccine strain S-191 (genotype A) by less than 1% at nucleotide level, and therefore appeared a vaccine-associated strain. The other 5 strains as XJ03-27, XJ03-74, XJ04-146, XJ04-150 and XJ04-152 were proved to be genotype H1 strains,among which XJ03-27, XJ03-74, XJ04-150 and XJ04-152, showing their nucleotide diversity were varied from 0.5% to 1.6%, when compared to the H1a reference strain China9322, and identified as H1a strains. XJ04-146 showed a nucleotide similarity of 98.7% when compared to H1b reference strain China9475, and was identified as H1b strain. Additionally, we found that there were two sets of strain (XJ03-27 and XJ04-150; XJ03-74 and XJ04-152), with almost identical nucleotide sequences, circulating in 2003 and 2004 and both having more nucleotide variability (up to 6.1%, 27 nucleotides).

CONCLUSIONGenotype H1 measles virus had been proven to have been circulated in Xinjiang, China during 2003 and 2004. H1a was the predominant epidemic strain while H1b strain stood the next.

China ; epidemiology ; Disease Outbreaks ; Genotype ; Humans ; Measles ; epidemiology ; Measles virus ; classification ; genetics ; isolation & purification ; Nucleoproteins ; genetics ; Phylogeny ; RNA, Viral ; genetics ; Sequence Analysis, RNA ; Viral Proteins ; genetics

OBJECTIVETo investigate whether HV and Ot can coexist in their host (Leptotrombidium scutellare).

METHODSCollecting the separate Leptotrombidium scutellare and the ones from mice in epidemic area. The cells of mites at larva, nymph, and adult stages were cultured and made into smear. In situ RT-PCR and PCR were used to detect and locate HV RNA and Ot DNA in the primary cultured cells.

RESULTSPositive signals of HV RNA and Ot DNA distributed mostly in epithelial cells of digestive system and ovary cells of larva and nymph. The positive rate increased by the generation of passages.

CONCLUSIONCoinfection of HV and Ot did exist in wild Leptotrombidium scutellare.

Animals ; Cells, Cultured ; DNA, Bacterial ; analysis ; Female ; Hantavirus ; isolation & purification ; Mice ; Mites ; microbiology ; virology ; Orientia tsutsugamushi ; isolation & purification ; RNA, Viral ; analysis ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate hepatitis E virus (HEV) infection among pigs in Henan province.

METHODSA total of 623 swine sera, collected from 5 districts, were divided into two groups, under 3-month of age and over 3-month of age. They were tested for HEV antigen and antibody by using ELISAs, respectively. The sera positive for HEV antigen were tested for HEV RNA with RT-PCR. The positive products of RT-PCR were cloned and sequenced.

RESULTSThe positive rates of anti-HEV antibody of the groups under 3-month and over 3-month of age were 90.27% and 92.55%, respectively, without statistical difference, while those of HEV antigen were 15.93% and 5.69%, respectively, with significant difference. The positive rates of anti-HEV antibody and HEV antigen were significantly different among different districts. HEV RNA was detectable in 5 of 47 HEV antigen positive samples. The sequence analysis showed that in 4 of 5 specimens the sequence belonged to genotype 4 while in the remaining one the sequence was genotype 1.

CONCLUSIONThe prevalence rate of HEV infection in pigs was high in Henan province and the rate differed in different districts.

Animals ; Antibodies, Viral ; analysis ; immunology ; Antigens, Viral ; analysis ; immunology ; China ; Genotype ; Hepatitis E ; epidemiology ; immunology ; veterinary ; virology ; Hepatitis E virus ; genetics ; immunology ; isolation & purification ; Phylogeny ; RNA, Viral ; analysis ; genetics ; Sequence Analysis, DNA ; Swine ; virology ; Swine Diseases ; epidemiology ; immunology ; virology

OBJECTIVETo explore the molecular characteristics and molecular variation of human rotavirus (HRV) strains and to understand the relationship between clinical characteristics and epidemiology of different HRV-VP7 and NSP4.

METHODSDouble-strand RNA of rotavirus extracted from stool samples was used as the template for reverse transcription of gene VP7, which was followed by nested PCR for VP7 typing. NSP4 genes from 22 epidemic strains of human rotavirus isolated in Kunming in 2002 and 2003 were amplified with RT-PCR. Then cDNAs were sequenced and compared with 4 human rotavirus NSP4 (Wa, KUN, AU-1, Hochi)) and 3 animal rotavirus NSP4 (EW, OSU, SA11) available in the GenBank while the epidemic strains of human rotavirus isolated in different areas of China were compared, using the Clustal-mp, DNAssist, MEGA2 software. The G serotype of VP7 was analysed by PCR.

RESULTSSerotype G1 was prevalent in 2002 while serotype G3 was the prevalent in Kumming in 2003. The NSP4 genes from 22 epidemic strains of human rotavirus isolated in Kunming in 2002 and 2003 belonged to Wa with highly conservative amino acid. Samples isolated in the same years but not in the same area shared higher homology. Symptoms associated with heavy diarrhea did not seem to be associated with NSP4 molecular variation (P > 0.05).

CONCLUSIONObvious variations of VP7 typing were seen in the same season, as well as in different areas and years. Due to the stable nature of NSP4, it seem to be a better candidate for vaccine production, than VP7.

China ; DNA, Complementary ; genetics ; DNA, Viral ; Genes, Viral ; Humans ; Polymerase Chain Reaction ; RNA, Double-Stranded ; genetics ; RNA, Viral ; RNA-Directed DNA Polymerase ; Rotavirus ; classification ; genetics ; isolation & purification ; Rotavirus Vaccines ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid ; Serotyping