Date of Award

2014-01-01

Degree Name

Doctor of Philosophy

Department

Biological Sciences

Advisor(s)

Siddhartha Das

Abstract

Giardia lamblia is a protozoan parasite and a major cause of the waterborne-illness, giardiasis, worldwide. Giardiasis is endemic in developing countries and is a leading cause of non-viral- and non-bacterial-associated intestinal disorders. Acute symptoms of giardiasis include diarrhea, cramps, and malabsorption. The disease is often self-limiting, although the infection can result in long-term disorders such as chronic fatigue, stunted cognitive skills, and failure to thrive even after the parasite has been cleared.

This parasite exists in two morphologic forms--infective trophozoites and transmissible, water-resistant cyst, which is passed by the fecal-oral route and is likely to be spread by contaminated drinking water. Infection can occur upon ingestion of as few as 10 cysts, and an infected individual sheds more than 109 cysts in the feces per day. Although giardiasis can effectively be treated with nitroheterocyclic compounds, such drugs can cause unpleasant side effects and ultimately lead to drug-resistant parasites. Therefore, it is necessary to identify novel targets for developing new anti-giardial agents. One of these targets could be sphingolipid (SL) pathways because the results from our laboratory indicate that SL genes and gene products play critical roles in regulating encystation (cyst formation) of Giardia. We have shown previously that SL metabolism in Giardia

The goal of my Dissertation is to establish the sphingolipidome of Giardia during encystation to provide a global perspective of the SL changes that take place during this critical stage of the parasite's life cycle and to show how Giardia can modulate these lipids. In Specific Aim 1, I used mass spectrometry to elucidate the sphingolipid profile of Giardia during encystation as well as culture media sources of SLs. In Specific Aim 2, I investigated the parasite's ability to generate ceramide from the hydrolysis of sphingomyelin (SM) via two putative sphingomyelinase (SMase) enzymes identified by mining the Giardia genome project.

Briefly, I found that the levels of ceramide and glycosylceramides increased significantly (~10 fold) during encystation followed by a decline in cysts. The level of sphingomyelin (SM), on the other hand, increased sharply in the cyst population. Analysis indicated that SM-enriched cysts are viable and undergo excystation in culture. It was also observed that Giardia expresses two active acid SMase enzymes (gASMLPD3b1 and gASMLPD3b2) with distinct pH requirements, one of which shows the characteristics of a secreted SMase. My results indicate that a dynamic metabolic conversion among various classes of SLs occurs during giardial encystation, and that this conversion could be critical for completing excystation and producing infection in the human gut.

Language

en

Provenance

Received from ProQuest

File Size

90 pages

File Format

application/pdf

Rights Holder

Trevor Thomas Duarte

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