DESCRIPTION (provided by applicant): Mosquito-borne diseases impact millions of people around the globe. Malaria alone affects 40% of the world's population and kills over one million people annually, primarily young children. Unfortunately the primary control measures, particularly drugs and insecticides, are becoming less effective as resistance emerges in both the parasite and the mosquito vector respectively. Thus, it is important that we quickly develop new measures for controlling mosquito-borne diseases. One novel approach is to replace wild mosquito populations with one engineered to be refractory to key mosquito-borne parasites such as Plasmodium. While anti-malarial effector genes have been identified and successfully engineered into Anopheles mosquitoes, a mechanism for driving the effector genes through wild populations is not currently available. One possible mechanism for driving effector genes through a population is the use of transposable elements. Although several elements tested to date have proven to be ineffective drive mechanisms, transposable elements remain one of the most likely mechanisms for driving effector genes. Thus, we must continue to identify and test new transposable as they become available. Towards that goal we have identified a novel transposable element Hztransib. Hztransib represents the first intact and active Transib transposable element and was recently identified from the cotton bollworm Helicoverpa zea in the Li lab. Several characteristics of Hztransib suggest that it may have great promise as a genetic drive mechanism. This study will assess its suitability as a mechanism for driving effector genes through wild mosquito populations. Specifically, this study will first determine whether Hztransib is transpositionally active in Anopheles stephensi cell lines. An. stephensi ASE cells transfected with the Hztransib-pGEMT plasmids will be analyzed by RT-PCR analyses of the Hztransib transcripts. Transformation of DH51 E. coli cells with the enzyme-digested (two incompatible enzymes unique within the Hztransib element) plasmids recovered from the transfected An. stephensi cells and sequencing will be conducted to identify the excised smaller plasmids (which are resistant to enzyme digestion). The integration of the excised Hztransib into the An. stephensi cell genome will be verified by genome walking and sequencing. The study will then determine whether Hztransib is capable of duplicating and remobilizing within the germline of the human malaria vector An. stephensi. Hztransib helper and donor An. stephensi lines will be generated, followed by genetic crossing and screening for eye marker phenotypes to estimate the rates of post-integration excision, duplication, and remobilization. PCR and genome walking analyses of the post-integration events identified by G2 screening will also be conducted. If Hztransib is capable of replicating and remobilizing in the mosquito, it would be an important step towards implementing a population replacement strategy for malaria and other vector borne diseases. PUBLIC HEALTH RELEVANCE While anti-malarial effector genes have been identified and engineered into Anopheles mosquitoes, a mechanism for driving the effector genes through wild populations is not available. This study will assess the suitability of a novel and active transposable element Hztransib as a drive mechanism for driving the effector genes through wild mosquito populations. If successful, this project will constitute a major step toward the goal of replacing a wild vector population with a refractory one.
|Effective start/end date||6/1/09 → 5/31/11|
- National Institutes of Health: $226,500.00
- National Institutes of Health: $189,323.00
- Immunology and Microbiology(all)