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Interpretation Criteria for Standardized Western Blot for the Predominant Species of Borrelia Burgdorferi Sensu Lato in China

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Biomedical and Environmental Sciences 2010年第5期23卷 341-349页

作者： YI JIANG XUE-XIA HOU ZHEN GENG QIN HAO KANG-LIN WAN state key laboratory for infectious diseases prevention and control National Institute of Communicable Disease control & prevention Chinese Center of Disease Control & Prevention Beijing 102206 China

Objective Western blotting （WB;immunoblotting） is a widely used tool for the serodiagnosis of Lyme borreliosis （LB）,but so far,no generally accepted criteria for its performance and interpretation have been established in *** present study was designed to determine the criteria for standardized Western blot for the predominant species of Borrelia burgdorferi sensu lato in China,in which WB was produced with strain PD91 as the representative strain attributed to predominant genospecies Borrelia garinii of Borrelia burgdorferi sensu *** Approximately 13 bands between 14 and 100 kD were differentiated for strain PD91 by using Gel-Pro analysis *** a study with 631 serum samples （taken from 127 patients with Lyme borreliosis and 504 controls）,all observed bands were *** establish criteria for a positive WB result for strain PD91,receiver operating characteristic （ROC） curves were *** The following interpretation criteria were recommended：for IgG,at least one band of P83/100,P58,P39,P30,OspC,P17,P66,and OspA;for IgM,at least one band of P83/100,P58,OspA,P30,OspC,P17 or *** addition,syphilis,leptospirosis and other related diseases should be excluded when the positive band is P41 in *** IgG criteria,the sensitivity is 73.2%,the specificity is 99.4% and Youden index is 0.726;for IgM criteria,the sensitivity is 50.6%,the specificity is 93.1% and Youden index is *** Standardization of WB assays is necessary for comparison of results from different ***,the criteria of other genospecies of Borrelia burgdorferi sensu lato should be determined in the future to complete the criteria of WB for the diagnosis of the Lyme disease in China.

关键词： Criteria Western blot Borrelia burgdorferi sensu lato

Preliminary Study on Drug Susceptibility Profile and Resistance Mechanisms to Macrolides of Clinical Isolates of Non-tuberculous Mycobacteria from China

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Biomedical and Environmental Sciences 2018年第4期31卷 290-299页

作者： LI Fu LI Gui Lian PANG Hui LIU Hai Can XIAO Tong Yang LI Shuang Jun LUO Qiao JIANG Yi WANG Rui Bai WAN Kang Lin state key laboratory for infectious diseases prevention and control Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases National Institute for Communicable Disease Control and Prevention Chinese Center for Disease Control and Prevention Immunology Department Changzhi Medical College key laboratory of laboratory Medicine Wenzhou Medical University

Objective Macrolide susceptibility and drug resistance mechanisms of clinical non-tuberculous mycobacteria（NTM） isolates were preliminarily investigated for more accurate diagnosis and treatment of the infection in China. Methods Four macrolides, including clarithromycin（CLAR）, azithromycin（AZM）, roxithromycin（ROX）, and erythromycin（ERY）, were used to test the drug susceptibility of 310 clinical NTM isolates from six provinces of China with the broth microdilution method. Two resistance mechanisms, 23 S r RNA and erm, were analyzed with nucleotide sequence analysis. Results Varied effectiveness of macrolides and species-specific resistance patterns were observed. Most Mycobacterium abscessus subsp. massiliense were susceptible and all M. fortuitum were highly resistant to macrolides. All the drugs, except for erythromycin, exhibited excellent activities against slow-growing mycobacteria, and drug resistance rates were below 22.2%. Only four highly resistant strains harbored 2,058/2,059 substitutions on rrl and none of other mutations were related to macrolide resistance. G2191 A and T2221 C on rrl were specific for the M. abscessus complex（MABC）. Seven sites, G2140 A, G2210 C, C2217 G, T2238 C, T2322 C, T2404 C, and A2406 G, were specifically carried by M. avium and M. intracellulare. Three sites, A2192 G, T2358 G, and A2636 G, were observed only in M. fortuitum and one site G2152 A was specific for M. gordonae. The genes erm（39） and erm（41） were detected in M. fortuitum and M. abscessus and inducible resistance was observed in relevant sequevar. Conclusion The susceptibility profile of macrolides against NTM was demonstrated. The well-known macrolide resistance mechanisms, 23 S r RNA and erm, failed to account for all resistant NTM isolates, and further studies are warranted to investigate macrolide resistance mechanisms in various NTM species.

关键词： Non-tuberculous mycobacteria Macrolide Drug resistance

Rapid and High-throughput Identification of Recombinant Bacteria with Mass Spectrometry Assay

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Biomedical and Environmental Sciences 2014年第4期27卷 250-258页

作者： XIAO Di TAO Xiao Xia WANG Peng LIU Guo Dong GONG Ya Nan ZHANG Hui Fang WANG Hai Bin ZHANG Jian Zhong state key laboratory for infectious Disease prevention and control National Institute for Communicable Disease Control and PreventionChinese Center for Disease Control and Prevention Collaborative Innovation Center for Diagnosis and Treatment of infectious diseases Provincial key laboratory for Plague control and prevention Yunnan Provincial Institute for Endemic Diseases Control and Prevention Beijing Municipal Center for Disease control and prevention Chaoyang District Center for Disease control and prevention Beijing

Objective To construct a rapid and high-throughput assay for identifying recombinant bacteria based on mass spectrometry. Methods Matrix assisted laser desorption ionization time-of-flight mass spectrometry （MALDI-TOF MS） techniques were used to identify 12 recombinant proteins （10 of Yersinia pestis, 1 of Campylobacter jejuni and 1 of Helicobacter pylori）. A classification model for the various phase of recombinant bacteria was established, optimized and validated, using MALDI-TOF MS-CIinProTools system. The differences in the peptide mass spectra were analyzed by using Biotyper and FIexAnalysis softwares. Results Models of GA, SNN, and QC were established. After optimizing the parameters, the GA recognition model showed good classification capabilities： RC=100%, mean CVA=98.7% （the CVA was 96.4% in phase 1, 100% in phase 2, 98.4% in phase 3, and 100% in phase 4, respectively） and PPV=95}. This model can be used to classify the bacteria and their recombinant, which only requires 3.7x103 cells for analysis. The total time needed is only 10 min from protein extraction to reporting the result for one sample. Furthermore, this assay can automatically detect and test 96 samples concurrently. A total of 48 specific peaks （9, 16, 9, and 14 for the four stages, respectively） was found in the various phase of recombinant bacteria. Conclusion MALDI-TOF MS can be used as a fast, accurate, and high-throughput method to identify recombinant bacteria, which provide a new ideas not only for recombinant bacteria but also for the identification of mutant strains and bioterrorism pathogens.

关键词： Recombinant bacteria MALDI-TOF MS ClinProTools Rapid identification Specific peaks

Distinctive Drug-resistant Mutation Profiles and Interpretations of HIV-1 Proviral DNA Revealed by Deep Sequencing in Reverse Transcriptase

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Biomedical and Environmental Sciences 2016年第4期29卷 239-247页

作者： YIN Qian Qian LI Zhen Peng ZHAO Hai PAN Dong WANG Yan XU Wei Si XING Hui FENG Yi JIANG Shi Bo SHAO Yi Ming MA Li Ying state key laboratory of infectious Disease prevention and control National Center for AIDS/STD Control and Prevention (NCAIDS) Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases Chinese Center for Disease Control and Prevention state key laboratory of infectious Disease prevention and control National Institute for Communicable Disease Control and Prevention Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases Chinese Center for Disease Control and Prevention key laboratory of Medical Molecular Virology (Ministries of Education and Health) Shanghai Medical College and Institute of Medical Microbiology Fudan University

Objective To investigate distinctive features in drug-resistant mutations （DRMs） and interpretations for reverse transcriptase inhibitors （RTIs） between proviral DNA and paired viral RNA in HIV-l-infected patients. Methods Forty-three HIV-l-infected individuals receiving first-line antiretroviral therapy were recruited to participate in a multicenter AIDS Cohort Study in Anhui and Henan Provinces in China in 2004. Drug resistance genotyping was performed by bulk sequencing and deep sequencing on the plasma and whole blood of 77 samples, respectively. Drug-resistance interpretation was compared between viral RNA and paired proviral DNA. Results Compared with bulk sequencing, deep sequencing could detect more DRMs and samples with DRMs in both viral RNA and proviral DNA. The mutations M1841 and M2301 were more prevalent in proviral DNA than in viral RNA （Fisher＇s exact test, P〈0.05）. Considering ＇majority resistant variants＇, 15 samples （19.48%） showed differences in drug resistance interpretation between viral RNA and proviral DNA, and 5 of these samples with different DRMs between proviral DNA and paired viral RNA showed a higher level of drug resistance to the first-line drugs. Considering ＇minority resistant variants＇, 22 samples （28.57%） were associated with a higher level of drug resistance to the tested RTIs for proviral DNA when compared with paired viral RNA. Conclusion Compared with viral RNA, the distinctive information of DRMs and drug resistance interpretations for proviral DNA could be obtained by deep sequencing, which could provide more detailed and precise information for drug resistance monitoring and the rational design of optimal antiretroviral therapy regimens.

关键词： HIV-1 drug-resistant mutation Drug-resistance interpretation Proviral DNA Viral RNA Deep sequencing

Investigation of the genotype III to genotype I shift in Japanese encephalitis virus and the impact on human cases

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Virologica Sinica 2015年第4期30卷 277-289页

作者： Na Han James Adams Wei Fang Si-Qing Liu Simon Rayner state key laboratory for infectious diseases prevention and control National Institute for Communicable Disease Control and Prevention Chinese Center for Disease Control and Prevention Collaborative Innovation Center for Diagnosis and Treatment of infectious diseases state key laboratory of Virology Wuhan Institute of Virology Chinese Academy of Sciences Department of Medical Genetics Oslo University Hospital

Japanese encephalitis is a mosquito borne disease and is the leading cause of viral encephalitis in the Asia-Pacific area. The causative agent, Japanese encephalitis virus(JEV) can be phylogenetically classified into five genotypes based on nucleotide sequence. In recent years, genotype I(GI) has displaced genotype III(GIII) as the dominant lineage, but the mechanisms behind this displacement event requires elucidation. In an earlier study, we compared host variation over time between the two genotypes and observed that GI appears to have evolved to achieve more efficient infection in hosts in the replication cycle, with the tradeoff of reduced infectivity in secondary hosts such as humans. To further investigate this phenomenon, we collected JEV surveillance data on human cases and, together with sequence data, and generated genotype/case profiles from seven Asia-Pacific countries and regions to characterize the GI/GIII displacement event. We found that, when comprehensive and consistent vaccination and surveillance data was available, and the GIII to GI shift occurred within a well-defined time period, there was a statistically significant drop in JEV human cases. Our findings provide further support for the argument that GI is less effective in infecting humans, who represent a dead end host. However, experimental investigation is necessary to confirm this hypothesis. The study highlights the value of alternative approaches to investigation of epidemics, as well as the importance of effective data collection for disease surveillance and control.

关键词： flavivirus Japanese encephalitis virus(JEV) bioinf

HIV-Specific IL-2^+ and/or IFN-γ^+ CD8^+ T Cell Reponses during Chronic HIV-1 Infection in Former Blood Donors

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Biomedical and Environmental Sciences 2010年第5期23卷 391-401页

作者： YAN-MENG FENG YAN-MIN WAN LIAN-XIN LIU CHAO QIU PENG-FEI MA HONG PENG YU-HUA RUAN LI-FENG HAN KUN-XUE HONG HUI XING YI-MING SHAO state key laboratory of Virology Wuhan Institute of VirologyChinese Academy of SciencesWuhan 430071HubeiChina state key laboratory for infectious diseases prevention and control Institute for Communicable Disease Control and PreventionChinese Center for Disease Control and PreventionBeijing 102206China state key laboratory for infectious diseases prevention and control National Center for AIDS/STD Control and PreventionChinese Center for Disease Control and PreventionBeijing 100050China Graduate School of the Chinese Academy of Sciences Beijing 100049China

Objective Conflicting data have been generated from previous studies to determine which kind of relationship exists between HIV-1 specific CD8 Tcell responses and HIV-1 viral load or CD4 count over the course of *** this study,153 HIV-1 infected LTNPs were enrolled to investigate the role of HIV-1 specific CD8 T-cell responses in chronic HIV-1 infection among HIV-1 infected former blood *** The patients were stratified into three groups according to CD4 count：CD4≥500 cells/μL;350 cells/μL≤CD4〈500 cells/μL;CD4〈350 cells/μ*** were isolated from the patients＇ anticoagulated blood ***-2 and IFN-γ secretions of CD 8 T cells against 17 HIV-1 consensus B full peptide pools were analyzed by using ICS *** An overall inverse correlation were observed between CD4 count and plasma viral *** no significant difference was observed during the comparisons of frequency/breadth of HIV-1 specific CD8 T cell responses,CD4 count stratification analysis showed that different correlation pattern existed in three strata：as for patients whose CD4 counts were less than 350 cells/μL,no significant correlations were identified between frequency/breadth of HIV-1 specific CD8 T cell responses and CD4 count/viral load;as for patients whose CD4 counts ranged from 350 cells /μL to 500 cells/μL,significant correlation was only observed between the response breadth of IL-2＋IFN-γ＋ CD8 T cells and CD4 count;however,as for patients whose CD4 counts were more than 500 cells/μL,direct correlations were identified between IL-2＋IFN-γ＋/IL-2＋/IFN-γ＋ CD8 T cells and viral load or CD4 *** Universal consistent inverse correlation was only indentified between CD4 count and viral *** relationship between HIV-1 specific CD8 T cell responses and CD4 count/viral load varied in different CD4 strata,which showed that better preserved CD4 T cells were correlated with better CD8 T cell functions.

关键词： HIV-1 subtype B' CD8 T cell response IFN-γ IL-2 ICS

Evaluation of Six Recombinant Proteins for Serological Diagnosis of Lyme Borreliosis in China

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Biomedical and Environmental Sciences 2016年第5期29卷 323-330页

作者： LIU Wei LIU Hui Xin ZHANG Lin HOU Xue Xia WAN Kang Lin HAO Qin state key laboratory for infectious diseases prevention and control National Institute for Communicable Disease Control and Prevention Chinese Center for Disease Control and Prevention Beijing 102206 China Collaborative Innovation Center for Diagnosis and Treatment of infectious diseases Hangzhou 310003 Zhejiang China

Objective In this study, we evaluated the diagnostic efficiency of six recombinant proteins for the serodiagnosis of Lyme borreliosis （LB） and screened out the appropriate antigens to support the production of a Chinese clinical ELISA （enzyme-linked immunosorbent assay） kit for LB. Methods Six recombinant antigens, Fla B.g, OspC B.a, OspC B.g, P39 B.g, P83 B.g, and VlsE B.a, were used for ELISA to detect serum antibodies in LB, syphilis, and healthy controls. The ELISA results were used to generate receiver operating characteristic （ROC） curves, and the sensitivity and specificity of each protein was evaluated. All recombinant proteins were evaluated and screened by using logistic regression models. Results Two IgG （VISE and OspC B.g） and two IgM （OspC B.g and OspC B.a） antigens were left by the logistic regression model screened. VIsE had the highest specificity for syphilis samples in the IgG test （87.7%, P〈0.05）. OspC B.g had the highest diagnostic value in the IgM test （AUC=0.871）. Interactive effects between OspC B.a and Fla B.g could reduce the specificity of the ELISA. Conclusion Three recombinant antigens, OspC B.g, OspC B.a, and VisE B.a, were useful for ELISAs of LB. Additionally, the interaction between OspC B.a and Fla B.g should be examined in future research.

关键词： Lyme borreliosis Recombinant proteins Serological diagnosis Logistic regression ROC

Coxsackievirus B3 Infection Triggers Autophagy through 3 Pathways of Endoplasmic Reticulum Stress

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Biomedical and Environmental Sciences 2018年第12期31卷 867-875页

作者： LUO Xiao Nuan YAO Hai Lan SONG Juan SONG Qin Qin SHI Bing Tian XIA Dong HAN Jun state key laboratory of infectious Disease prevention and control Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases National Institute for Viral Disease Control and Prevention Chinese Center for Disease Control and Prevention Molecular Immunology laboratory Capital Institute of Pediatrics

Objective Autophagy is a highly conserved intracellular degradation pathway. Many picornaviruses induce autophagy to benefit viral replication, but an understanding of how autophagy occurs remains incomplete. In this study, we explored whether coxsackievirus B3(CVB3) infection induced autophagy through endoplasmic reticulum(ER) stress. Methods In CVB3-infected HeLa cells, the specific molecules of ER stress and autophagy were detected using Western blotting, reverse transcription polymerase chain reaction(RT-PCR), and confocal microscopy. Then PKR-like ER protein kinase(PERK) inhibitor, inositol-requiring protein-1(IRE1) inhibitor, or activating transcription factor-6(ATF6) inhibitor worked on CVB3-infected cells, their effect on autophagy was assessed by Western blotting for detecting microtubule-associated protein light chain 3(LC3). Results CVB3 infection induced ER stress, and ER stress sensors PERK/eIF2α, IRE1/XBP1, and ATF6 were activated. CVB3 infection increased the accumulation of green fluorescent protein(GFP)-LC3 punctuation and induced the conversion from LC3-Ⅰ to phosphatidylethanolamine-conjugated LC3-1(LC3-Ⅱ). CVB3 infection still decreased the expression of mammalian target of rapamycin(mTOR) and p-mTOR. Inhibition of PERK, IRE1, or ATF6 significantly decreased the ratio of LC3-Ⅱ to LC3-Ⅰ in CVB3-infected HeLa cells. Conclusion CVB3 infection induced autophagy through ER stress in HeL a cells, and PERK, IRE1, and ATF6 a pathways participated in the regulation of autophagy. Our data suggested that ER stress may inhibit mTOR signaling pathway to induce autophagy during CVB3 infection.

关键词： Coxsackievirus B3(CVB3) Autophagy Endoplasmic reticulum(ER) stress Unfolded protein response(UPR)

Transforming bacterial disease surveillance and investigation using whole-genome sequence to probe the trace

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Frontiers of Medicine 2018年第1期12卷 23-33页

作者： Biao Kan Haijian Zhou Pengcheng Du Wen Zhang Xin Lu Tian Qin Jianguo Xu state key laboratory of infectious Disease prevention and control National Institute for Communicable Disease control and prevention Chinese Center for Disease Control and Prevention Beijing 102206 China Collaborative Innovation Center for Diagnosis and Treatment of infectious diseases Hangzhou 310003 China Beijing key laboratory of Emerging infectious diseases Institute of Infectious Diseases Beijing Ditan Hospital Capital Medical University Beijing 100015 China

Two decades have passed since the first bacterial whole-genome sequencing, which provides new opportunity for microbial genome. Consequently, considerable genetic diversity encoded by bacterial genomes and among the strains in the same species has been revealed. In recent years, genome sequencing techniques and bioinformatics have developed rapidly, which has resulted in transformation and expedited the application of strategy and methodology for bacterial genome comparison used in dissection of infectious disease epidemics. Bacterial whole-genome sequencing and bioinformatic computing allow genotyping to satisfy the requirements of epidemiological study in disease control. In this review, we outline the significance and summarize the roles of bacterial genome sequencing in the context of bacterial disease control and prevention. We discuss the applications of bacterial genome sequencing in outbreak detection, source tracing, transmission mode discovery, and new epidemic clone identification. Wide applications of genome sequencing and data sharing in infectious disease surveillance networks will considerably promote outbreak detection and early warning to prevent the dissemination of bacterial diseases.

关键词： genome sequencing genomic epidemiotogy bacteria surveillance infectious diseases