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Engineering G-Protein Coupled Receptor-Based Living Yeast Diagnostics for Infectious DiseasesRios, Davida Marie January 2023 (has links)
Diagnostics serve as the frontline defense for the containment and mitigation of infectious diseases. The emerging synthetic biology field established numerous useful applications of engineered biological systems and networks that led to the development of living biosensors. Significant effort has been made to develop G-coupled protein receptor (GPCR)-based yeast biosensors for applications in drug discovery, environmental monitoring, and clinical diagnostics of small molecules and fungal pathogens. Of the living biosensor chassis, yeast-based biosensors offer exceptional advantages over other in vitro diagnostics, such as long-term storage in a reagent-free and dried dormant state, an engineered colorimetric readout for yes/no result interpretation, and high scalability through industrial fermentation. These advantages could be the next innovation as a low-cost, low-tech, and reliable diagnostic alternative in point-of-care and at-home contexts.
Chapter 1 provides background information related to synthetic biology, living biosensors, direction evolution, and point-of-care diagnostics. Chapter 2 covers the development of engineered living yeast as a diagnostic tool for viral infections by tailoring the biosensing recognition element to sense any amino acid-based biomarker of choice via directed evolution. Chapter 3 describes the development of living yeast biosensor for the detection of the pathogenic fungus, Aspergillus fumigatus, in clinical supernatants and patient samples. Chapter 4 describes the progression of a living yeast biosensor for the multi-drug resistant pathogenic fungus, Candida auris, and its detection in clinical culture supernatants and samples.
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Neutrophil CD64 and monocyte HLA-DR cell surface markers for diagnosis of early-onset neonatal infection. / CUHK electronic theses & dissertations collectionJanuary 2005 (has links)
A total of 338 infants with suspected clinical sepsis were investigated, 115 of whom were found to be clinically infected. Twenty-one healthy term neonates were recruited as control subjects. The expression of CD64 on neutrophils in infected infants was significantly elevated at both 0 h and 24 h, compared with those of noninfected infants or controls (both p < 0.0005). The calculated optimal cutoff value for CD64 was 6136 antibody-phycoerythrin molecules bound/cell. CD64 has a very high sensitivity (96%) and NPV (97%) at 24 h. The use of CRP in combination with CD64 as predictive markers only marginally enhanced the sensitivity and NPV (97% and 98%, respectively). There was no statistical difference in the expression of monocyte HLA-DR among infected, noninfected, and control subjects. As a result, the optimal cutoff value for HLA-DR could not be determined. The technology of flow cytometry has potential applications for use in the diagnosis of neonatal sepsis because the measurement is quantitative, requiring only a minimal amount of whole blood and a short duration (within 3 h) for the provision of results. (Abstract shortened by UMI.) / Term newborns in whom infection was suspected when they were <72 h of age were recruited into the study. The expressions of CD64 on neutrophils and HLA-DR on monocytes were measured by flow cytometry at 0 h (the time of sepsis evaluation) and 24 h after the onset of presentation. A full sepsis screen, including complete blood count, serial C-reactive protein (CRP), blood culture, cerebrospinal fluid culture, and chest radiograph were performed. The demographic and clinical data were documented. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of neutrophil CD64, monocyte HLA-DR and the combination of markers for predicting neonatal sepsis were determined. / This prospective study aimed to evaluate the diagnostic utilities of two cell surface markers, neutrophil CD64 and monocyte HLA-DR, for the identification of early-onset clinical infection and pneumonia in term infants. The optimal cutoff value of each marker was defined according to the Receiver Operating Characteristic curve so that it could be used as a reference with which future studies can be compared. / Li Geng. / "May 2005." / Adviser: Pak Cheung Ng. / Source: Dissertation Abstracts International, Volume: 67-01, Section: B, page: 0174. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (p. 129-150). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.
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