A method to determine unique lipids in mosquitoes using High Performance Thin Layer Chromatography coupled to Matrix Assisted Laser Desorption lonisation Fourier Transform Mass Spectrometry (TLC-MALDI-FTMS)
- Authors
- Sarabia L?pez, Lenin Daniel
- Format
- MasterThesis
- Status
- publishedVersion
- Description
Plasmodium is the etiological agent of malaria which causes more than 3569 million clinical cases and around 0.86 million deaths per year. The lack of effective treatments and the spread of resistance against currently used drugs encourages the need to identify new targets and approaches to control this disease. The malaria parasite comprises differing stages in two hosts, namely, the vertebrate and the protozoan. Several studies and models have been developed for the mammalian stage, whilst just a handful have been performed in the protozoan host. The mosquito (protozoan) stage constitutes the major population bottleneck to the parasite, as parasites have an inefficient passage to the salivary glands. Therefore, a better understanding of the parasite-mosquito development may elucidate new targets for intervention. In this work, I have attempted to address the hypothesis that the parasite has a variety of specific lipids and that it must synthesize these lipids de novo or scavenge lipids from the mosquito host. Therefore, this study aimed to develop methods to measure the changes in the lipid profile in the mosquito midguts during infection so that they could be very important to understanding parasite biology in the mosquito host. This study reveals that it is possible to use an approach based on TLC coupled to MALDIIMS to analyze lipids from whole extracts by TLC and map them back in vivo using MALDI-IMS. Results showed that for TLC separation of mosquito lipids extracts, the use of a highly basic eluant favored the separation of different lipid classes. Coupling of TLC to MALDI-IMS allowed to identify several lipid classes off of the TLC plate coated with DHB and produced a significant amount of valuable data. Also, sectioning of whole frozen-mosquitoes for MALDI-IMS was achieved by embedding them between boiled egg-white and freezing them over liquid nitrogen. Mosquito cross-sections analyzed by MALDI-IMS allowed to map back to the tissue some of the lipids found in TLC-MALDI-MS part of the study. Lipid species such as glycerophosphoserine (PS(41:7)), glycerophosphoinositol (PI(35:2)) and sphingoid base (LysoSM(d17:1)) were localized in the midgut region of the mosquito. These findings allowed to verify the changes that occur in the lipid profile of mosquitoes as a result of the Plasmodium infection and development in the midgut. This study has advanced our understanding of Plasmodium development in the midgut and showed that imaging mass spectrometry techniques, such as MALDI, have proven to be a powerful and useful tool in the analysis of changes in the biological system formed between parasite and mosquito.
- Publication Year
- 2014
- Language
- eng
- Topic
- BIOTECHNOLOGY
METABOLOMICS
MALARIA
PLASMODIUM
- Repository
- Repositorio SENESCYT
- Rights
- openAccess
- License