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New approaches to fluorescence-based diagnostics for human African trypanosomiasis

Giordani, Federica (2011) New approaches to fluorescence-based diagnostics for human African trypanosomiasis. PhD thesis, University of Glasgow.

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Abstract

In the absence of any vaccine, prophylactic drug and effective vector control, the fight against human African trypanosomiais (HAT) is based on the the combination of active case-finding and consequent drug treatment of identified positive cases. Unfortunately, low sensitivity and specificity of current diagnostic techniques often result in misdiagnosis, leaving infected patients without cure or exposing them to inappropriate chemotherapy protocols, which use dangerous and expensive drugs. The development of more efficient, simple, cheap and field-robust diagnostic tests is, therefore, urgently needed. In the field, direct observation by light microscopy of trypanosomes in human fluids (blood, lymph node aspirate, cerebrospinal fluid) is considered the ideal way of confirming HAT infection. However, in practice this approach is problematic, especially for the Gambian form of the disease, where patients may present with very low parasitaemia. Detection limits of parasitological techniques can be improved by adding a preliminary step of sample concentration, although this further increases the laboriousness of HAT diagnostic algorithm. Recent advances in fluorescence microscopy could be exploited to facilitate trypanosome detection. The introduction and implementation of fluorescence microscopy in HAT endemic countries would offer the advantages of an increased overall sensitivity of microscopical examination and a more rapid screening of the specimen. In contrast to traditional, expensive and fragile fluorescence microscopes, new LED-illuminated instruments are relatively cheap, very efficient and portable, lending themselves to utilisation in poorly equipped rural settings. In order to design a new diagnostic tool that exploits LED technology, however, selective and reliable fluorescent markers to label trypanosomes in human fluids are needed. The development of new tools to assist in the diagnosis of African trypanosomiasis by use of LED fluorescence microscopy was the overall objective of this project. The work was mainly focused on testing various fluorescent compounds for their ability to selectively stain trypanosomes. Fluorophores were otained from commercial and academic sources, or else directly synthesised during the project. An important requirement evaluated was the compounds’ compatibility with the currently available SMR LED Cytoscience fluorescence microscope, developed and kindly provided by our collaborator Prof. D. Jones (Philipps University, Marburg). The utility of a UV LED-driven microscope in performing the arsenical drug resistance test was also assessed. This assay, developed in our laboratory to detect trypanosome strains resistant to arsenical and diamidine compounds, could represent a useful tool for chemotherapeutic decision making in the field, where resistance to arsenical drugs is a rising problem.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Human African trypanosomiasis, diagnosis, fluorescence microscopy, light-emitting diodes, fluorophores
Subjects: Q Science > QR Microbiology
R Medicine > RB Pathology
Colleges/Schools: College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Supervisor's Name: Michael, Prof. Barrett and Andrew, Dr. Sutherland
Date of Award: 2011
Depositing User: Ms Federica Giordani
Unique ID: glathesis:2011-2454
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 23 Mar 2011
Last Modified: 10 Dec 2012 13:55
URI: http://theses.gla.ac.uk/id/eprint/2454

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