Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses
Invasive mycoses pose significant challenges to immunocompromised individuals, often resulting in severe morbidity and mortality. Current diagnostic methods are hindered by delays in identification and initiation of appropriate antifungal therapy, stemming from the limitations of conventional tec...
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| Format: | Thesis |
| Language: | English |
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2024
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| Online Access: | http://eprints.usm.my/61995/1/YASMIN%20KHAIRANI%20BT%20MUHAMMAD%20ISMADI-TESIS%20P-UD002220-E.pdf |
| _version_ | 1825908575350816768 |
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| author | Ismadi, Yasmin Khairani Muhammad |
| author_facet | Ismadi, Yasmin Khairani Muhammad |
| author_sort | Ismadi, Yasmin Khairani Muhammad |
| collection | USM |
| description | Invasive mycoses pose significant challenges to immunocompromised
individuals, often resulting in severe morbidity and mortality. Current diagnostic
methods are hindered by delays in identification and initiation of appropriate
antifungal therapy, stemming from the limitations of conventional techniques. This
study undertakes a thorough investigation into fungal DNA extraction protocols and
the development of a multiplex real-time PCR assay for rapid identification of
invasive fungal pathogens. This research aims to compare fungal DNA extraction
methods and develop a multiplex real-time PCR assay capable of simultaneously
identifying multiple fungal species commonly associated with invasive mycoses.
Species-specific primer design and assay optimization target Aspergillus fumigatus,
Aspergillus terreus, Candida albicans, and Candida glabrata. Results reveal certain
extraction methods exhibiting superior sensitivity and time efficiency, with
implications for clinical use. The developed multiplex real-time PCR assay
demonstrates promising accuracy in identifying fungal pathogens. Analytical
specificity testing, which involved detecting 65 isolates including various fungal
strains alongside other reference strains, yielded a 100% identification rate for target
organisms. Performance evaluation of the multiplex PCR assay using spiked blood
samples showed high specificity and sensitivity, both at 100%, with positive
predictive value (PPV) and negative predictive value (NPV) also at 100%. Overall,
this research contributes to advancing laboratory diagnosis and management of invasive fungal infections. By addressing challenges in fungal DNA extraction and
creating a high-performance multiplex PCR assay, this study lays the groundwork
for more efficient and reliable diagnostic approaches, thereby enhancing patient care
and treatment outcomes. Further validation studies on real clinical samples are
recommended to confirm the applicability of these methodologies in the current
clinical settings. |
| first_indexed | 2025-03-06T09:56:02Z |
| format | Thesis |
| id | usm.eprints-61995 |
| institution | Universiti Sains Malaysia |
| language | English |
| last_indexed | 2025-03-06T09:56:02Z |
| publishDate | 2024 |
| record_format | dspace |
| spelling | usm.eprints-619952025-03-05T07:45:01Z http://eprints.usm.my/61995/ Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses Ismadi, Yasmin Khairani Muhammad R Medicine Invasive mycoses pose significant challenges to immunocompromised individuals, often resulting in severe morbidity and mortality. Current diagnostic methods are hindered by delays in identification and initiation of appropriate antifungal therapy, stemming from the limitations of conventional techniques. This study undertakes a thorough investigation into fungal DNA extraction protocols and the development of a multiplex real-time PCR assay for rapid identification of invasive fungal pathogens. This research aims to compare fungal DNA extraction methods and develop a multiplex real-time PCR assay capable of simultaneously identifying multiple fungal species commonly associated with invasive mycoses. Species-specific primer design and assay optimization target Aspergillus fumigatus, Aspergillus terreus, Candida albicans, and Candida glabrata. Results reveal certain extraction methods exhibiting superior sensitivity and time efficiency, with implications for clinical use. The developed multiplex real-time PCR assay demonstrates promising accuracy in identifying fungal pathogens. Analytical specificity testing, which involved detecting 65 isolates including various fungal strains alongside other reference strains, yielded a 100% identification rate for target organisms. Performance evaluation of the multiplex PCR assay using spiked blood samples showed high specificity and sensitivity, both at 100%, with positive predictive value (PPV) and negative predictive value (NPV) also at 100%. Overall, this research contributes to advancing laboratory diagnosis and management of invasive fungal infections. By addressing challenges in fungal DNA extraction and creating a high-performance multiplex PCR assay, this study lays the groundwork for more efficient and reliable diagnostic approaches, thereby enhancing patient care and treatment outcomes. Further validation studies on real clinical samples are recommended to confirm the applicability of these methodologies in the current clinical settings. 2024-08 Thesis NonPeerReviewed application/pdf en http://eprints.usm.my/61995/1/YASMIN%20KHAIRANI%20BT%20MUHAMMAD%20ISMADI-TESIS%20P-UD002220-E.pdf Ismadi, Yasmin Khairani Muhammad (2024) Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses. PhD thesis, Universiti Sains Malaysia. |
| spellingShingle | R Medicine Ismadi, Yasmin Khairani Muhammad Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title | Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title_full | Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title_fullStr | Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title_full_unstemmed | Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title_short | Development of a multiplex real-time pcr assay for detection of fungal pathogens in invasive mycoses |
| title_sort | development of a multiplex real time pcr assay for detection of fungal pathogens in invasive mycoses |
| topic | R Medicine |
| url | http://eprints.usm.my/61995/1/YASMIN%20KHAIRANI%20BT%20MUHAMMAD%20ISMADI-TESIS%20P-UD002220-E.pdf |
| work_keys_str_mv | AT ismadiyasminkhairanimuhammad developmentofamultiplexrealtimepcrassayfordetectionoffungalpathogensininvasivemycoses |