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Objective: To investigate the etiology of severe hand-foot-mouth disease (HFMD) in children in Henan province., Methods: A total of 244 HFMD cases admitted to a hospital in Zhengzhou from April to June of 2014 were recruited for research sampling, Real-time RT-PCR, virus isolation, VP1 sequencing and alignment methods were used to test the enterovirus-related etiology. SPSS 17.0 was used in performing statistical analysis., Results: There were 109 severe and 135 mild cases among all the 244 HFMD cases. The number of enterovirus positive stool samples was 229, with positive rate as 93.85%. EV71, Cox A16 and Cox A10 made up 83.84%, 5.68% and 8.30% of the enterovirus etiologicy, strains, respectively. EV71 infection caused 8 HFMD cases with heart-lung failure and 2 death, Cox A10 infection led to 1 HFMD case with heart-lung failure and death. There were statistically differences seen regarding the enterovirus infection rates between severe and the mild HFMD cases (χ(2)=5.312,P=0.021). Statistically significant difference was seen in the constituent ratio of EV71, Cox A16 and the others by Fisher' s exact test (P=0.048). There was statistically significant difference seen between the cardiorespiratory failure rate and the fatality rate by EV71 and Cox A10 infection (χ(2)=0.051,P=0.821; χ(2)=2.198,P=0.138). Cox A10 strains idenfied in Henan in 2014 belonged to genotype 6. The rates on homology of nucleotide and amino acid among the Cox A10 strains in Henan in 2014 were 94.3%-99.7% and 96.3%-100.0% respectively., Conclusions: EV71 still remained the most common pathogen that causing severe HFMD in children, with the increasing Cox A10 percentage in the pathogens spectrum of HFMD infection. Cox A10 strains in Henan in 2014 belonged to genotype 6. Genotype 6 Cox A10 had appeared and widely distributed in Henan for long time, but not yet variated or reconstructed. Cox A10 infection could lead to cardio-respiratory failure thus called for the monitoring program on non-EV71 and non-Cox A16 enterovirus, especially Cox A10 to be strenthened.

Objective: To detect and analyze the distribution of virulence factors, antimicrobial resistance and pulsed field gel electrophoresis (PFGE) patterns of Shigella sonnei, isolated in Henan province from 2011 to 2014., Methods: Samples of diarrhea patients were collected and isolated with SS selective culture medium in 37 ℃ for 18-24 hours. All strains were identified under the Kligler iron agar/motility-indol-urea biochemical action and API20E biochemical system. Serological typing and prepared DNA template were carried out with thermal cracking method and multiplex PCR, to detect the virulence genes of Shigella sonnei. According to the molecular typing method and K-B drug susceptibility testing method published by the international PulseNet bacterial infectious disease monitoring network and USA clinical and Laboratory Standards Institute, antimicrobial susceptibility tests and PFGE molecular characteristics of these positive strains isolated from sentinel hospitals patients stool samples were analyzed., Results: Among the 98 strains of Sonnei type Ⅰ and 118 strains of Sonnei type Ⅱ, all the strains carried carry different virulence genes including SHET-1B, SHET-2, ial, ipaH genes, with 4 kinds of virulence gene combination types. All the 216 strains of Shigella sonnei belonged to the multi-drug resistant strains, including 34 isolates resistant to 2-4 kinds of antibiotics(15.7%), 147 isolates to 5-8 kinds of antibiotics (68.1%), 24 to 9-10 kinds of antibiotics (11.1%), 7 to 11 kinds of antibiotics (3.2%), and 4 to 13 kinds of antibiotics (1.9%). A total of 100 strains of Shigella sonnei were divided into 31 molecular patterns, digested by XbaⅠ and PFGE. Each pattern contained 1-13 strains with similarities ranged from 68.6%-100.0%., Conclusions: All the Shigella sonnei strains carried virulence pathogenic factors, presenting serious status on drug resistance. PFGE fingerprinting patterns showed high polymorphism and dominant characteristics. PFGE patterns of partial strains and corresponding antidrug spectrum presented certain relevance and with same aggregation relationship.

Objective: To investigate the antibiotic resistance and pulsed field gel electrophoresis (PFGE) patterns of clinical isolates of Salmonella (S.) typhi and S. paratyphi in Henan province during 2009-2011., Methods: According to molecular typing and Salmonella K-B drug susceptibility test method published by international PulseNet bacterial infectious disease monitoring network and USA Clinical and Laboratory Standards Institute (CLSI), the drug susceptibility and PFGE molecule characteristics of 78 S. typhi and S. paratyphi strains isolated from sentinel hospitals in Henan were analyzed., Results: The 78 strains of S. typhi and S. paratyphi were resistant to 13 kinds of antibiotics, in which 62 were multidrug resistant (79.5%), 4 were resistant to 2-3 kinds of antibiotics (5.1%), 41 were resistant to 5-8 kinds of antibiotics (52.6%), 14 were resistant to 9-10 kinds of antibiotics (17.9%), 3 were resistant to 11-12 kinds of antibiotics (3.8%). The resistant rate to cephalosporins, quinolones and other 3 kinds of antibiotic showed an increase trends. Seventy two strains of S. typhi and S. paratyphi could be divided 14 molecular patterns by digestion with XbaⅠ and PFGE, each pattern contains 1-47 strains which shared the similarity of 66.03%-100.00%., Conclusions: The drug resistance of clinical isolates of S. typhi and S. paratyphi was serious in Henan. The PFGE patterns showed diversity, but the predominant patterns could be still found. The PFGE patterns of some strains were associated with their drug resistance.

Objective: To diagnose imported dengue fever case from Henan province, and to sequence and analyze the characteristics of whole genome sequence, and to explore the possible viral origin source., Methods: A suspected dengue fever case was reported in Yuzhou city, Henan province. The patient returned from foshan, Guangdong province on September 19, 2014, after the epidemiological investigation and serum specimen collected, which dengue fever case was diagnosed in the laboratory, then it was inoculated on Vero cells. Whole genome sequence was amplified by several pairs primers and characterized using biologic software., Results: The imported case was diagnosed as dengue virus 1 serotype infection. Dengue 1 strain was isolated using Vero cells successfully. Whole genome was 10,670 nt, which belonged to dengue virus 1 serotype V genotype and didn't found any recombination event. The phylogenetic analysis demonstrated that the strain was closed to Indian starins isolated in 2008-2011, and the homology of nucleotide sequence was between 98.2%-99.4%., Conclusion: It was the first time to discover imported dengue 1 serotype case in Henan province. However, according to the patient has been to Guangdong province before onset, it inferred that the Indian strain had been imported to Guangdong province before this case in Henan province.

Objective: To compare the survival effect of initiation of antiretroviral treatment on AIDS patients at different stages so as to explore the best time of initiation with the treatment., Methods: Information regarding the dates of AIDS patient initially receiving the anti-retroviral therapy during 2007-2012 was collected from the Chinese HIV/AIDS Integrated Control System. According to the level of baseline immunology, all the participants were divided into earlier treatment group (baseline CD4(+) T cell counts between 350/µl and 500/µl) and conventional treatment group (baseline CD4(+) T cell counts ≤350/µl). Data was analyzed under Survival and Review methods., Results: A total number of 16 282 cases were selected. The mortality of conventional treatment group was obviously higher than the earlier treatment group (5.78/100 person year vs. 1.64/100 person year), and the median survival time was lower than the earlier treatment group (2.07 year vs. 3.15 year). The cumulative survival rate of the 6-year conventional treatment group was lower than the earlier treatment group (77.39% vs. 92.10%, χ(2) = 156.00, P < 0.01). By means of multi-variable analysis, we found that factors as age, gender, marital status, route of transmission, schedule on initial therapy, number of symptoms at the baseline etc. in the conventional treatment group were associated with survival time of patients after the ART initiation (P < 0.05), while gender, schedule for initial therapy, number of symptoms at baseline etc. in the earlier treatment group, were associated with survival time of patients after the initiation of ART (P < 0.05)., Conclusion: Earlier initiation of antiretroviral treatment on AIDS patients could improve the survival rate and prolong the survival time.

Objective: To clone and express the recombinant capsid protein VP2 of enterovirus type 71 (EV71) and to identify the immune activity of expressed protein in order to build a basis for the investigation work of vaccine and diagnostic antigen., Methods: VP2 gene of EV71 was amplified by PCR, and then was cut by restriction enzyme and inserted into expression vector pMAL-c2X. The positive recombinants were transferred into E.coli TB1, the genetically engineered bacteria including pMAL-c2X-VP2 plasmids were induced by isopropyl thiogalactoside ( IPTG) , and the expression products were analyzed by SDS-PAGE and western blotting method. EV71 IgM antibody detection method by ELISA was set up, and the sensitivity and specificity of this method was assessed; 60 neutralizing antibody positive serum samples from hand foot and mouth disease (HFMD) patients were determined, of which 52 samples were positive and 8 samples were negative; a total of 88 acute phase serum samples of HFMD patients diagnosed in clinical were also detected., Results: VP2 gene of 762 bp was obtained by PCR, the gene segment inserted into the recombinant vector was identified using restriction enzyme digestion. The recombinant vector could express a specific about 71 500 fusion protein in E.coli by SDS-PAGE. The purified recombinant protein of EV71-VP2 can react with the serum of HFMD patients to produce a specific band by western blotting. The sensitivity and specificity of ELISA was 87% and 83%, respectively. Of the 88 acute phase serum samples from children with HFMD, 48 samples (55%) were positive by the ELISA assay., Conclusions: VP2 gene of EV71 has been cloned and a prokaryotic high expression system for VP2 gene was successfully constructed in the present study. The recombination EV71-VP2 has well antigenicity, which could be useful for developing diagnose reagent or vaccine of EV71.