具有致病性的寄生蠕蟲感染在全球范圍內引起嚴重的健康問題并造成巨大的經濟損失。因此,能夠及時、準確地診斷蠕蟲感染是控制病原傳播的關鍵策略。以核酸檢測為基礎的診斷方法,主要包括聚合酶鏈式反應、定量qPCR、環(huán)介導等溫擴增技術和重組酶聚合酶鏈式擴增等,均是能夠有效檢測蠕蟲感染的方法。這些檢測方法具有準確性、及時性和靈敏度高等優(yōu)點。由血吸蟲感染引起的血吸蟲病是一個被忽視的熱帶性疾病,在全球范圍內引起重大的公共衛(wèi)生問題。血吸蟲病的早期診斷和檢測對該病的防控至關重要。傳統的診斷方法主要包括檢測糞便中的蟲卵,或者通過免疫學方法檢測血吸蟲循環(huán)抗原。但是這些方法對血吸蟲病感染早期的診斷敏感性低。循環(huán)性DNA（cfDNA）是游離于宿主體液循環(huán)中的細胞外DNA片段,它被視為一種快速、有效的檢測和診斷病原體的方法。本研究中,我們探索了不同的實驗動物（兔和小鼠）在感染血吸蟲后線粒體循環(huán)DNA的診斷效果。首先,我們評估了三種不同的循環(huán)性DNA提取試劑盒對循環(huán)性DNA的分離效果,結果表明,QIAamp循環(huán)核酸試劑盒在提取循環(huán)性DNA的效果最好。隨后,我們對日本血吸蟲線粒體DNA的一些分子進行了分析,并鑒定出了線粒體... 

【文章來源】：中國農業(yè)科學院北京市

【文章頁數】：63 頁

【學位級別】：博士

【文章目錄】：
摘要
Abstract
英文縮略表
CHAPTER 1
    1.1 Introduction
    1.2 Distribution of Schistosomes
    1.3 Life Cycle of schistosoma
    1.4 Pathogenesis of Schistosomiasis
    1.5 Cell-Free DNA and parasitic Infections
    1.6 Source of cf DNA and Its Application in Diagnosis
    1.7 Detection of schistosoma in the Circulatory System
    1.8 Problem and solution
CHAPTER 2 RECENT ADVANCES IN NUCLEIC ACID-BASED METHODS FOR DETECTION OF HELMINTH INFECTIONS AND THE PERSPECTIVE OF BIOSENSORS FOR FUTURE DEVELOPMENT
    2.2 Introduction
    2.3 Nucleic acid based methods for detection of helminth infections
        2.3.1 PCR
        2.3.2 Multiplex PCR
        2.3.3 LAMP
        2.3.4 Recombinase polymerase amplification(RPA)
    2.4 Biosensors for nucleic acid biomarker detection
        2.4.1 Electrochemical detection techniques
        2.4.2 Optical detection techniques
    2.5 Advantages and limitations of nucleic acid-based methods for detection of helminth infections
    2.6 Conclusions
CHAPTER 3 CIRCULATING CELL FREE MITOCHONDRIAL DNA FRAGMENTS ARE INFECTION
    3.2 Introduction
    3.3 Material and Methods
        3.3.1 Establishment of S.japonicum infection
        3.3.2 Praziquantel(PZQ)treatment of infected mice and blood samples collection
        3.3.3 Isolation of eggs from liver homogenate of infected mice and DNA extraction from eggs and adult S. japonicum
        3.3.4 cf DNA extraction from infected host serum
        3.3.5 Primer design and PCR amplification
        3.3.6 Gene cloning in p MD19 T vector and standard curve construction by qRT-PCR
        3.3.7 Statistical analyses
    3.4 Results
        3.4.1 Cf DNA extraction kits comparison
        3.4.2 Reference Gene of S.japonicum detected in infected rabbit serum
        3.4.3 Detection of Schistosoma japonicum cf DNA by PCR assay
        3.4.4 Detection of S.japonicum cf DNA in different week of post infection
        3.4.5 Detection of S.japonicum cf DNA in different number of cercariae of infection
        3.4.6 Efficacy evaluation after chemotherapy
        3.4.7 Characterization and copy number of target fragments
    3.5 Disscussion
    3.6 Conclusion
CHAPTER 4 DETECTION OF CIRCULATING CELL-FREE NUCLEAR DNA TO DIAGNOSE SCHISTOSOMA JAPONICUM INFECTION
    4.2 Introduction
    4.3 Material and Methods
        4.3.1 Ethics statement
        4.3.2 Establishment of S.japonicum infection and collection of samples
        4.3.3 Extraction of eggs from liver homogenate of infected mice and DNA extraction from eggs and adult S. japonicum infected host serum
        4.3.4 PCR amplification
        4.3.5 Gene cloning and standard curve construction by qRT-PCR for copy number detection
    4.4 Results
    4.5 Disscussion
    4.6 Conclusion
References
Acknowledgement
Curriculum Vitae



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