Objective To predict and screen the B-cell epitopes on nucleoprotein(NP)of human avian H5N1 virus strain.Methods As NP nucleotide sequences of strain A/GD/01/06(H5N1)were sequenced,B-cell epitopes were predicted by the analysis of the flexible regions of secondary structure for NPprotein and by screening on B-cell epitopes for NP protein with methods of Hydrophilicity Plot by KyteDoolittle,Surface Probability Plot by Emini,and Antigenic Index by Jameson-Wolf.Then a screening method was established by Hierarchical cluster,Bivariate correlate and Quartiles in SPSS13.0.and the variation of amino acids in NP protein was appraised in epitope prediction.Resuits The computer-predicted most possible epitopes for NP protein were located within or nearby its N-terminal 317-326,452-457,467-473,367-370,491-498,375-378,171-177,48-53,245-250.76-104 and so on.NP protein in A/GD/01/06(H5N1)increased a glycoprotein domain(NES368-370)by the substitution of N370S and changed the bio-features.Conclusion Stepwise prediction of the B-cell epitopes for the NP protein based on multiple parameters is helpful for the molecular-immunology and drug-screening,and the substitution of N370 S in NP of A/GD/01/06(H5N1)varied its antigenicity but didn't change the epitope size(SNEN367-370).

Objective To study the anti-viral effects of Lianhua Qingwen Capsule against influenza A virus in vitro.Methods Under different administration modes,the inhibitory effect of Lianhua Qingwen Capsule on human influenza A virus(H3N2)in vitro and its time-effect relationship were assayed by crystal violet staining assay with ribavirin as positive reference drug.Results Lianhua Qingwen Capsule showed a multi-access effect on anti-human influenza A virus,such as comprehensive inhibition effect,prevention effect on virus adsorption,inhibition on virus proliferation after adsorption and direct killing the virus effect,and the median effective concentrations(EC50)in above-mentioned effects were 0.042,0.031,0.051 and 0.050 mg/mL respectively.Among them,the prevention effect on virus adsorption was the strongest.With the expanding actuation duration of the drug,the inhibitory effect of Lianhua Qingwen Capsule against influenza A virus weakened at low concentration,but no change at high concentration(≥0.031 g/mL).Meanwhile,Lianhua Qingwen Capsule can remarkably decrease the infectivity of influenza A virus.Conclusion Lianhua Qingwen Capsule has an obvious effect against human influenza A virus in vitro.

3.00 and exposed to the solution containing 1500 mg/L peracetic acid for 30 min could completely killing the spores.Aerosol spraying with solution containing 1000 mg/L peracetic acid at a dose of 10 ml/m3 for 30 min caused over 90.0% decay rate of natural bacteria in indoor air.CONCLUSIONS Weikangshi peracetic acid has good stability and its germicidal efficacy is not affected.

Objective To investigate the genetic relationship of the Dengue virus strains isolated in Guangdong province in 2006 and 2007, and to find the sources of these virus. Methods Serum samples of the dengue fever patients from 5 cities of Guangdong province in 2006 and 4 cities in 2007 were collected. Three pairs of primers that specific for amplifying the 3 overlap fragments of E gene of Dengue virus type Ⅰ were designed. RNAs were extracted from C6/36 cells treated with patients' serum. E genes were amplified by RT-PCR, purified and then sequenced directly. To obtain the E gene complete sequences, the raw sequences were assembled and edited. Obtained E genes were compared with E genes of other Dengue virus type Ⅰ published in the GenBank, analyzed by MEGA version 4.1 software. Results In 2006, virus circulating in Guaogzhou2006(EF508203) was closest to Vietnam2006(EU482539) with 99.3% nucleotides homology, Chaozhou2006 (EF508206) and Shantou2006 (EF508207) strains were closest to Japan2004 (AB178040) and Singapore2006 (EU081280) with 99.5% nucleotides homology, while Yangjiang2006 (EF508205) and Yangjiang2001 (EF508200) were closest to each other and both with 99.5% nucleotides homology to Thailand2001 (AY732482). All 4 Dengue virus strains circulated in 2007 were closest to Singapore2005(EU081276) with 99.7% nueleotides homology. Conclusion The Dengue viruses prevalent in 2006 were from different sources while those in 2007 came from the same origin. The data also showed that there was an endemic area of Dengue virus in Guangdong province.

Objective To investigate the etiological characteristics of Streptococcus pyogenes that caused scarlet fever in different periods in Guangdong province.Methods 22 isolates from different periods were analyzed through emm typing,PCR detection for super antigen genes,antibiotic susceptibility test and pulsed-field gel electrophoresis (PFGE).Results All isolates were susceptible to cefotaxim,levofloxacin and penicillin.Streptococcus pyogenes isolated after the year 2000 were 100％ resistant to erythrocin and clindamycin,but the resistant rate for strains isolated before the year 2000 was 9.1％ (1/11).There were 3 emm types indentified from 22 isolates including emm12.0 (59.09％,13/22),emm6.0 (36.36％,8/22) and emm1.0 (4.55％,1/22),which were detected in the isolates from the year 1997 and 2011,from 1978 and 1986,and from 2008,respectively.The positive rates for speA,speB,speC,speF,speG,speH,smeZ,and ssa genes detected by PCR were 54.55％,100％,100％,100％,100％,54.55％,0％,and 86.36％ respectively.Among all strains,95.45％ of the isolates carried 6 superantigen genes simultaneously.Three clusters of 10 PFGE subtypes were identified in 22 isolates.Cluster Ⅰ consisted of all strains from 1997 and one strain from 2011.Cluster Ⅱ consisted of strains isolated from 1978 and 1986.Cluster Ⅲ consisted of nine strains from 2011 and one from 2008.Conclusion S.pyrogenes isolates in Guangdong province were susceptible to penicillin but resistant to erythrocin.emm 12.0 accounted for the majority of the three types and there was a high frequency of super antigen genes.

ObjectiveTo study the toxin genes and pandemic group distribution as well as the genetic correlation between the major serotypes( O3:K6,O1:Kut,O4:K8 ) of Vibrio parahaemolyticus (VP) isolated from the outbreaks of Guangdong province.MethodsThe tdh and trh genes,GS-PCR and orf8 gene were detected on the 62 isolates sourced from patient and seafood occurred in the 23 outbreaks during 2008-2010.44 isolates of which were analyzed on PFGE digested by Not Ⅰ enzyme.ResultsToxin genes distributions suggested that 96.8% (60/62)isolates were tdh+,trh-.Three tdh+ isolates sourced from seafood were found.Pandemic group distribution suggested that 97.2% (35/36) O3:K6,5.88% (1/17) O4:K8,66.7% (8/9)O1:Kut serotype was GS-PCR+ and/or orf8+,respectively.PFGE analysis suggested that 44 isolates were separated into 3 clusters,of which the similarity of PFGE profile was 80.5% in the pandemic group cluster constructed by 28 isolates,the similarity between pandemic group and non-pandemic group was 59.5%.Pandemic group of O3:K6,O1:Kut as well as 04:K8 isolated on some outbreaks were processing the same PFGE profiles.ConclusionThe characteristic of toxin genes of major serotypes VP isolated in the outbreaks of Guangdong province form 2008-2010 was tdh- present and trh- absent.Within the pandemic group,O3:K6 and O1:Kut were the major serotypes.In single outbreak,isolates belongs to pandemic group but with different serotype seem to be close correlations.

To explore the possibility to type the Brucella strains isolated in Guangdong province with analytical method to detect the fatty acid components and to collect the basic data of fatty acid components of Brucella strains, 29 strains of Brucella were selected for analysis on the bacterial fatty acid components and the cluster analysis on the collected data was performed with Sherlock analysis soft-ware (MIDI). It was demonstrated that the main fatty acid components of Brucella strains isolated in Guangdong province were 19∶0 cycloω8c acid, 16∶0 acid and 18∶0 acid. The content of 19∶0cycloω8c acid was highest in B.abortus, followed by B.melitensis and lowest in B.suis.-In addition, the content differences of 19∶0cycloω8c and 18∶0 acid between B. melitensis and Brucella suis were statistically significant; and that of 19∶0cycloω8c and 18∶0 acid between strains isolated in 1965 and those isolated in recent 3 years was statistically significant. It was also shown that the fatty acid components of Brucella strains were stable, but the contents of fatty acid components were different in different species.-It is evident that at certain euclidean distance, 3 species of Brucella can be differentiated in species level.

Objective To study the molecular characters of porA and porB genes which encode outer membrane proteins (OMP), and predominated clonal complex of Neisseria meningitidis isolates from Guangdong province. Methods Eighteen Neisseria meningitidis isolates from Guangdong province during year 1967 to 2007 were recovered and reconfirmed by API NH biochemical system, and serogrouped by antiserum. The characters of porA and porB gene were analyzed by DNA sequencing. The allele profiles and the sequence types (ST) were determined by multilocus sequence typing (MLST). Based on their allelic profiles, the evolution relationship was analyzed by PHYLIP software. The predominant clonal complex was determined through comparing with the information of reference strains from the PubMLST database. Results For porA gene, type 20 was more frequently in the variable region (VR) 1 and type 9 in VR2 before year 2004. However, for porB gene, type 4 was more frequently in VR Ⅰ, type7 in VR Ⅳ, type 11 in VR Ⅴ,and type 10 in VR Ⅵ, respectively. The multi-types character was presented in VR Ⅴ and VR Ⅵ after2004. VR Ⅶ and VR Ⅷ can not be found among all the isolates except for one W135 isolate in 2007. Among the seven housekeeping genes, the polymorphism of abcZ was the lowest one with 4 allele numbers, while pgm was the highest one with 13 allele numbers. The predominant clonal complex was ST-5 before 2004. The ST-4821 clone complex appeared in 2004 and caused cases every year since then. More important, highly invasive ST-11 clonal complex firstly appeared in Guangdong in 2007. Conclusion The molecular characteristic of OMP genes presents polymorphism for the Neisseria meningitidis isolates from patients in Guangdong province during 1967 to 2007. ST-5 is the predominant clonal complex before 2004 and the highly invasive clonal complex is circulating in recent 3 years. It suggests that the surveillance based on laboratory should be further enhanced.

Objective To investigate the polymorphism of Brucella melitensis biovar 3 ( B.melitensis biovar 3) strains isolated from Guangdong province .Methods PCR assays followed by agar-ose gel electrophoresis and capillary electrophoresis based on the multiple locus variable number of tandem repeats (VNTR) analysis (MLVA) were performed to analyze 43 clinic isolates of B.melitensis biovar 3 strains isolated from clinical patients in Guangdong province .Results MLVA typing showed that the simi-larities of the analyzed locus among 43 strains of Brucella ranged from 68.2%to 100%.32 genotypes identi-fied among the isolates were identical (100%similarity).27 out of 43 strains (62.8%) were single geno-types, while the other 16 strains (37.2%) belonged to 5 other genotypes with 2 to 5 strains in each of them . Conclusion B.melitensis biovar 3 isolates showed polymorphism distribution in Guangdong province as in-dicated by MLVA typing analysis .Single-genotype isolates accounted for 62.8% of all studied strains.No predominant genotype was found among all isolates .

Objective To study the epidemic and genetic characteristics of coxsackievirus A6 ( CVA6) strains isolated in Guangdong province.Methods Enterovirus strains positive for neither entero-virus A71 ( EV71) nor CVA16 were isolated from Guangdong province during 2008 to 2013 to screen CVA6 isolates by real-time PCR.The entire sequences of viral genes encoding VP1 of CVA6 positive samples were amplified and sequenced.The phylogenetic analysis was performed to analyze the full-length gene sequences encoding VP1 of CVA6 isolates and sequences downloaded from GenBank by using DNAStar6.0 and MEGA5.2 software packages.Results CVA6 strains accounted for 61.4%of the 1672 non-EV71 and non-CVA16 enterovirus strains isolated in Guangdong province during year 2008 to 2013.The positive rates were respectively 10.5%(4/38), 66.7%(34/51), 36.2% (81/224), 63.0% (182/289), 62.3% (325/522) and 73.0%(400/548) from 2008 to 2013 and the differences among different years were significant (χ2=133.79, P<0.01).The CVA6 isolates could be classified into four clusters in the phylogenetic tree, designated A, B, C and D (including D1, D2 and D3 subgenogroups) genogroups.The four clusters shared nucleotide diversity ranging from 15.5% to 23.1%.The CVA6 strains isolated in Guangdong province shared 88.7%-100.0% homologies in nucleotide and 95.7%-100.0% in amino acid.Subtype D2 strains circulated during 2008 to 2012 and subtype D3 strains circulated during 2009 to 2013.Conclusion CVA6 strains were the predominant enterovirus strains among non-EV71 and non-CVA16 enterovirus strains circula-ted in Guangdong province from year 2008 to 2013.The CVA6 isolates could be classified into A, B, C and D genogroups based on the sequence analysis of VP1 region.Subgroups D2 and D3 isolates were identified and the subgroup D3 isolates were the prevalent strains in Guangdong.