1. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria DNA barcoding for disease vectors Daniel Adjei Boakye Parasitology Department Noguchi Memorial Institute for Medical Research

2. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Lymphatic filariasis elephantiasis elephantiasis

3. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Onchocerciasis Health and Socio- economic impact Health and Socio- economic impact

4. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Leishmaniasis

5. Impact of parasitic diseases Cause immense suffering Cause immense suffering Life-long disabilities Life-long disabilities Impair childhood growth and development Impair childhood growth and development Promote poverty, impair education and Promote poverty, impair education and Economic development Economic development Arable land desertedArable land deserted Population affected Population affected Mortality Mortality

6. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Neglected Tropical Diseases and Malaria VBDDisease ? Buruli ulcer 1 *Chagas2 **Cholera3 *Dengue4 *Dracunculiasis5 * Human African Trypanosomiasis 6 *Leishmaniasis7 Leprosy8 *Filariasis9 *Onchocerciasis10 *Schistosomiasis11 Soil-Transmitted Helminthiasis 12 **Trachoma13 *Malaria14

7. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria DiseasesVectorsAreas ChagasTriatomaS/America DengueMosquito(Ae) Tropical World Guinea worm CyclopAfrica H/TrypanosomiasisGlossinaAfrica LeishmaniasisPhlebotomus Africa, America, Asia Lymphatic filariasis Mosquito(An,Cx,Ae,Mn) Tropical World OnchocerciasisSimulium Africa, America SchistosomiasisSnail Tropical World MalariaMosquito(An) Vectors of Neglected Tropical Diseases

8. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Importance of correct identification Understanding the diversity of vector species and their respective vectorial importance is necessary for any targeted control effort Understanding the diversity of vector species and their respective vectorial importance is necessary for any targeted control effort Studies on Vector species Studies on Vector species Identification and clarification of species statusIdentification and clarification of species status Biology and ecology of vectorsBiology and ecology of vectors Vectorial importance of different speciesVectorial importance of different species

9. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Members of the S. damnosum complex in West Africa Determination of important vector species Determination of important vector species Basis for mapping limits of control programme Basis for mapping limits of control programme Only larvae can be identified with precision Only larvae can be identified with precision

10. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Anopheles gambiae species complex Anopheles gambiae sensu lato An. gambaie s.s An. arabiensis An. melasAn. merus An. quadrannulatus An. bwambae

11. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Taxonomic challenges within An. gambiae s.s Genetic discontinuities Genetic discontinuities from chromosomal analysis (Coluzzi et al 1985) Five chromosomal forms in West Africa: Mopti; Bamako; Bissau; Forest; Savanna in the ribosomal DNA region (rDNA) (Favia et al 1997) Two molecular forms described (della Torre et al 2001; 2005)

12. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Problems associated with current identification In Mali and Burkina Faso, the M and S forms correspond to the Mopti and Savanna chromosomal forms In Mali and Burkina Faso, the M and S forms correspond to the Mopti and Savanna chromosomal forms Molecular forms do not always correspond to the chromosomal forms in other countries Molecular forms do not always correspond to the chromosomal forms in other countries The two molecular forms may be a reflection of the presence of two different species in what is considered as one species – An. gambiae s.s. The two molecular forms may be a reflection of the presence of two different species in what is considered as one species – An. gambiae s.s.

13. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Practical consequences for disease control Susceptibility to insecticides Susceptibility to insecticides Differences in disease transmission capabilities Differences in disease transmission capabilities Adaptation to different breeding sites Adaptation to different breeding sites There is therefore the need to understand whether there are 2 or only 1 species There is therefore the need to understand whether there are 2 or only 1 species

14. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Methodology Multiplex PCR for species identification PCR – RFLP for molecular form identification Morphological identification - Gillies and Coetzee (1987) Intergenic spacer of the rDNA region of the X-chromosome Scott et al. (1993), Favia et al. (1997) and Fanello et al (2002)

15. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Methodology PCR of CO1 fragment & Sequencing: (Applied Biosystems 377 automated DNA sequencer) Sequence alignment & comparisons: The sequences obtained were aligned using BioEdit (Hall, 1999) BLAST database search for closely related sequences in the Genebank database. Analysis for species relationships (Kimura 2-parameter (K2P) model; Kimura, 1980: Neighbour-Joining (NJ) tree; Saitou & Nei, 1987) were conducted in Mega 4 (Tamura et al., 2007).

16. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Preliminary results

17. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Preliminary results Molecular forms of An. gambiae s.s. Molecular forms of An. gambiae s.s.

18. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Example of phylogenetic tree

19. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Ghana Mosquitoes’ file in the Completed Project section of the Barcode of Life website (http://www.barcodinglife.org). http://www.barcodinglife.org

20. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Conclusion DNA barcode can be used to DNA barcode can be used to Separate out closely related members of sibling species in vectors of diseasesSeparate out closely related members of sibling species in vectors of diseases Draw relationships between species collected from wider geographical areasDraw relationships between species collected from wider geographical areas Determine the presence of individuals currently unidentified using the available techniquesDetermine the presence of individuals currently unidentified using the available techniques

21. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Acknowledgements Funding: Funding: JRS Biodiversity FoundationJRS Biodiversity Foundation World Health OrganizationWorld Health Organization Gate’s FoundationGate’s Foundation Collaborators at the NMIMR Collaborators at the NMIMR Prof. Michael WilsonProf. Michael Wilson Dr Maxwell AppawuDr Maxwell Appawu Dziedzom de SouzaDziedzom de Souza Hilaria AmuzuHilaria Amuzu Ignatius NdongIgnatius Ndong

22. Western and Central Africa DNA Barcoding meeting: Abuja, Nigeria Thank you