Salivary gland-specific P. berghei reporter lines enable rapid evaluation of tissue-specific sporozoite loads in mosquitoes.

PloS One
Chandra RamakrishnanElena A Levashina

Abstract

Malaria is a life-threatening human infectious disease transmitted by mosquitoes. Levels of the salivary gland sporozoites (sgs), the only mosquito stage infectious to a mammalian host, represent an important cumulative index of Plasmodium development within a mosquito. However, current techniques of sgs quantification are laborious and imprecise. Here, transgenic P. berghei reporter lines that produce the green fluorescent protein fused to luciferase (GFP-LUC) specifically in sgs were generated, verified and characterised. Fluorescence microscopy confirmed the sgs stage specificity of expression of the reporter gene. The luciferase activity of the reporter lines was then exploited to establish a simple and fast biochemical assay to evaluate sgs loads in whole mosquitoes. Using this assay we successfully identified differences in sgs loads in mosquitoes silenced for genes that display opposing effects on P. berghei ookinete/oocyst development. It offers a new powerful tool to study infectivity of P. berghei to the mosquito, including analysis of vector-parasite interactions and evaluation of transmission-blocking vaccines.

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Citations

Jun 28, 2013·Malaria Journal·Sreelakshmi K SreenivasamurthyT S Keshava Prasad
Sep 18, 2012·Trends in Parasitology·Robert E SindenMichael J Delves
May 16, 2015·Nature Reviews. Microbiology·Tania F de Koning-WardBrendan S Crabb
Feb 7, 2014·Malaria Journal·Henri VialKelly Chibale
Dec 17, 2014·Antimicrobial Agents and Chemotherapy·Jessey ErathAna Rodriguez

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Methods Mentioned

BETA
dissect
dissection
transgenic
fluorescence
electrophoresis
PCR
fluorescence microscopy
dissections
transfection
X-ray

Software Mentioned

ImageJ
Axiovision
MicroWin 2000

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