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Postgraduate Thesis: Rapid diagnosis of isoniazid resistant mycobacterium tuberculosis by high resolution melting (HRM) assay
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TitleRapid diagnosis of isoniazid resistant mycobacterium tuberculosis by high resolution melting (HRM) assay
 
AuthorsChan, Ming-yan
陳明恩
 
Issue Date2012
 
PublisherThe University of Hong Kong (Pokfulam, Hong Kong)
 
AbstractMycobacterium tuberculosis (MTB) is a major infective agent causing human tuberculosis (TB) in the worldwide. Although tuberculosis can be treated by a six-month course of antibiotics, the prevalence of extensively drug-resistance TB (XDR-TB) made the disease becomes a global health problem. In addition to the conventional MTB detection methods, molecular methods become significant in drug resistant MTB detection which can enhance effective drug treatment. In this study, 200 MTB respiratory specimens were collected from patients with suspected tuberculosis in Tuen Mun Hospital in Hong Kong. Based on the culture method as a gold standard for MTB detection, the presence of MTB in clinical samples was determined by IS6110single tube nested real-time PCR. In addition, by using High Resolution Melting (HRM) analysis, the presence of mutant type KatG315 gene for detecting isoniazid resistant MTB was determined. Among 66 MTB culture positive samples, 10 samples had positive acid fast bacilli (AFB) smears giving the diagnostic sensitivity 15.1%. IS6110 single tube nested PCR was amplified in 51 specimens giving 77.2% MTB detection sensitivity and 97.8% specificity. Among 51 samples positive for IS6110 PCR, 66.7% showed successful amplification in subsequent KatG-HRM assay. Two samples were confirmed to be isoniazid (INH) resistance in Public Health Laboratory Centre (PHLC). However, there was only one sample showing detectable KatG315 mutation in clinical specimen by using HRM while the other was only detected in the corresponding culture isolate. From the result of this study, single tube nested real-time PCR demonstrated MTB detection in clinical samples and INH resistant strain with KatG315mutationcan be detected by HRM analysis. Early detection of mycobacteria allow earlier treatment of the patient, thus transmission of the disease can be controlled.
 
DegreeMaster of Medical Sciences
 
SubjectMycobacterium tuberculosis - Diagnosis.
 
Dept/ProgramMicrobiology
 
DC FieldValue
dc.contributor.authorChan, Ming-yan
 
dc.contributor.author陳明恩
 
dc.date.hkucongregation2012
 
dc.date.issued2012
 
dc.description.abstractMycobacterium tuberculosis (MTB) is a major infective agent causing human tuberculosis (TB) in the worldwide. Although tuberculosis can be treated by a six-month course of antibiotics, the prevalence of extensively drug-resistance TB (XDR-TB) made the disease becomes a global health problem. In addition to the conventional MTB detection methods, molecular methods become significant in drug resistant MTB detection which can enhance effective drug treatment. In this study, 200 MTB respiratory specimens were collected from patients with suspected tuberculosis in Tuen Mun Hospital in Hong Kong. Based on the culture method as a gold standard for MTB detection, the presence of MTB in clinical samples was determined by IS6110single tube nested real-time PCR. In addition, by using High Resolution Melting (HRM) analysis, the presence of mutant type KatG315 gene for detecting isoniazid resistant MTB was determined. Among 66 MTB culture positive samples, 10 samples had positive acid fast bacilli (AFB) smears giving the diagnostic sensitivity 15.1%. IS6110 single tube nested PCR was amplified in 51 specimens giving 77.2% MTB detection sensitivity and 97.8% specificity. Among 51 samples positive for IS6110 PCR, 66.7% showed successful amplification in subsequent KatG-HRM assay. Two samples were confirmed to be isoniazid (INH) resistance in Public Health Laboratory Centre (PHLC). However, there was only one sample showing detectable KatG315 mutation in clinical specimen by using HRM while the other was only detected in the corresponding culture isolate. From the result of this study, single tube nested real-time PCR demonstrated MTB detection in clinical samples and INH resistant strain with KatG315mutationcan be detected by HRM analysis. Early detection of mycobacteria allow earlier treatment of the patient, thus transmission of the disease can be controlled.
 
dc.description.naturepublished_or_final_version
 
dc.description.thesisdisciplineMicrobiology
 
dc.description.thesislevelmaster's
 
dc.description.thesisnameMaster of Medical Sciences
 
dc.identifier.hkulb4833340
 
dc.languageeng
 
dc.publisherThe University of Hong Kong (Pokfulam, Hong Kong)
 
dc.relation.ispartofHKU Theses Online (HKUTO)
 
dc.rightsThe author retains all proprietary rights, (such as patent rights) and the right to use in future works.
 
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License
 
dc.source.urihttp://hub.hku.hk/bib/B48333402
 
dc.subject.lcshMycobacterium tuberculosis - Diagnosis.
 
dc.titleRapid diagnosis of isoniazid resistant mycobacterium tuberculosis by high resolution melting (HRM) assay
 
dc.typePG_Thesis
 
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<date.issued>2012</date.issued>
<description.abstract>&#65279;Mycobacterium tuberculosis (MTB) is a major infective agent causing human tuberculosis (TB) in the worldwide. Although tuberculosis can be treated by a six-month course of antibiotics, the prevalence of extensively drug-resistance TB (XDR-TB) made the disease becomes a global health problem. In addition to the conventional MTB detection methods, molecular methods become significant in drug resistant MTB detection which can enhance effective drug treatment. 

In this study, 200 MTB respiratory specimens were collected from patients with suspected tuberculosis in Tuen Mun Hospital in Hong Kong. Based on the culture method as a gold standard for MTB detection, the presence of MTB in clinical samples was determined by IS6110single tube nested real-time PCR. In addition, by using High Resolution Melting (HRM) analysis, the presence of mutant type KatG315 gene for detecting isoniazid resistant MTB was determined.

Among 66 MTB culture positive samples, 10 samples had positive acid fast bacilli (AFB) smears giving the diagnostic sensitivity 15.1%. IS6110 single tube nested PCR was amplified in 51 specimens giving 77.2% MTB detection sensitivity and 97.8% specificity. Among 51 samples positive for IS6110 PCR, 66.7% showed successful amplification in subsequent KatG-HRM assay. Two samples were confirmed to be isoniazid (INH) resistance in Public Health Laboratory Centre (PHLC). However, there was only one sample showing detectable KatG315 mutation in clinical specimen by using HRM while the other was only detected in the corresponding culture isolate. 

From the result of this study, single tube nested real-time PCR demonstrated MTB detection in clinical samples and INH resistant strain with KatG315mutationcan be detected by HRM analysis. Early detection of mycobacteria allow earlier treatment of the patient, thus transmission of the disease can be controlled.</description.abstract>
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