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Identifier

000412591

Title

New rapid one-step PCR diagnostic assay for Plasmodium falciparum infective mosquitoes

Alternative Title

Νέα διαγνωστική μέθοδος για την ανίχνευση του παρασίτου Plasmodium σε μολυσματικά κουνούπια

Author

Κεφή, Μαρία Α.

Abstract

An essential component of malaria vector control programmes is the detection of Plasmodium falciparum within its mosquito hosts, particularly in the salivary gland where the infective sporozoites reside. Several protocols have been developed for this purpose; however they require dissection of mosquito specimens prior to analysis, in order to detect the parasite in the salivary gland. Here, a novel one step RT qPCR TaqMan diagnostic assay was developed specifically for mosquitoes with infective Plasmodium falciparum sporozoites in the salivary glands. The assay is based on detection of the sporozoite-specific Pfslarp and Pfplp1 gene transcripts. These two transcripts were chosen based on bioinformatics analysis and experimentally verified to be specifically expressed in the salivary gland sporozoite stage of the parasite but not in other mosquito parasite stages. The proof of principle and the performance of the assay were demonstrated using RNA later preserved mosquito samples. Tests of analytical sensitivity showed the novel TaqMan assay to be 100% accurate, although its performance in the field needs to be further demonstrated. This method has no requirement for head/thorax dissection and post-PCR processing and thus is simpler and more rapid to perform in individual mosquitoes or mosquito pools. Importantly it can be used in single or multiplex formats also targeting additional markers expressed in different tissues, such as detoxification enzymes associated with insecticide resistance.

Language

English

Subject

Malaria

Διάγνωση

Ελονοσία

Πλασμώδιο

Issue date

2017-11-22

Collection

School/Department--School of Sciences and Engineering--Department of Biology--Post-graduate theses

Type of Work--Post-graduate theses