Genetic Approaches for Malaria Control. Marcelo Jacobs-Lorena, PhD
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1 This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike License. Your use of this material constitutes acceptance of that license and the conditions of use of materials on this site. Copyright 2006, The Johns Hopkins University and Marcelo Jacobs-Lorena. All rights reserved. Use of these materials permitted only in accordance with license rights granted. Materials provided AS IS ; no representations or warranties provided. User assumes all responsibility for use, and all liability related thereto, and must independently review all materials for accuracy and efficacy. May contain materials owned by others. User is responsible for obtaining permissions for use from third parties as needed.
2 Genetic Approaches for Malaria Control Marcelo Jacobs-Lorena, PhD
3 The Malaria Problem At risk: >2,000,000,000 in 102 countries Infected: >270,000,000 Clinically ill: >110,000,000 cases each year Deaths: >2,000,000 each year 90% in Africa Mainly children 5-yr-old or younger
4 From CDC web site
5 Possible Weapons to Fight Malaria 1. Drugs that kill the parasite in humans 2. Insecticides that kill the mosquito vector 3. Vaccines
6 1. Drugs Plasmodium falciparum is now largely resistant to chloroquine, which used to be the most effective drug.
7 2. Insecticides: Mosquitoes Quickly Acquire Resistance Before During After Biologic niche intact Mosquitoes return
8 Mosquito Breeding Photo: Marcelo Jacobs-Lorena
9 Photo: Marcelo Jacobs-Lorena Mosquito Breeding
10 Mosquito Breeding Photo: Marcelo Jacobs-Lorena
11 3. Vaccine does not exist (yet). It does not exist (yet)
12 The mosquito is an obligatory vector (transmission by transfusion has no epidemiological significance) Interference with parasite development will result in decreased transmission
13 Anopheles gambiae CDC
14 NEW APPROACH: GENETIC MODIFICATION OF MOSQUITOES 1) GENETICALLY ENGINEER MOSQUITOES TO MAKE THEM RESISTANT TO PLASMODIUM 2) INTRODUCE THE REFRACTORY GENE(S) INTO MOSQUITO POPULATIONS LESS TRANSMISSION LESS MALARIA
15 Plasmodium development in the mosquito Ghosh et al. (2003) Trends in Parasitology 19: Used with Permission.
16 The use of a PHAGE DISPLAY LIBRARY to search for peptides with affinity to mosquito salivary gland and midgut epithelia
17
18 Salivary gland Midgut lumen Inject Feeder Dissect, wash Dissect gut, open into a sheet, wash Elute Elute Repeat selection 3 times Repeat selection 3 times Enriched phage population Enriched phage population From Ghosh et al. Proc Nat Acad Sci USA 2001;98: Copyright 2001 National Academy of Sciences, USA. Used with permission.
19 SM1 PEPTIDE I F A R Q I Q S BIOTIN P C C N
20 SALIVARY GLANDS INCUBATED WITH BIOTINYLATED PEPTIDES psm1 FITC DAPI unrelated peptide From Ghosh et al. Proc Nat Acad Sci USA 2001;98: Copyright 2001 National Academy of Sciences, USA. Used with permission.
21 THE SM1 PEPTIDE BINDS TO THE MIDGUT LUMEN BUT NOT TO THE OUTSIDE Open midgut sheet FITC DIC optics Closed (intact) midgut From Ghosh et al. Proc Nat Acad Sci USA 2001;98: Copyright 2001 National Academy of Sciences, USA. Used with permission.
22 SM1 Construct for Mosquito Germ Line Transformation Gut-specific & blood-inducible carboxypeptidase promoter [SM1] 4 Secretion signal HA1 epitope
23 An. stephensi - piggybac (3xP3-eGFP) eye-specific promoter Transgenic WT Transgenic WT Transgenic Larvae Adults Images courtesy of Jacobs-Lorena
24 Inhibition of oocyst formation in CP-[SM1] 4 transgenic mosquitoes Average of 9 experiments: Oocysts/ gut Non-trangenic 93 CP-[SM1] 4 18 Inhibition 82% (range: 69% - 95%) Ito et al. (2002) Nature 417:
25 Challenge: Driving Genes into Mosquito Populations What drive mechanisms are being considered? 1. Transposable elements 2. Wolbachia 3. Meiotic drive
26 An Alternative Introduce genes into bacteria of the mosquito midgut, instead of into the mosquito itself
27 - Feed E.coli (, or ) to An. stephensi mosquitoes - After 24 h feed a P. berghei-infected infected blood meal - After 15 d count oocysts per gut (n > 100 mosquitoes) 300 Mean oocysts/midgut * * OmpA SM1-1 SM1-2 * p < E. coli OmpA SM1
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