Eastern Africa Barcode Workshop, Oct DNA Barcoding - Parasites and Vectors Dan Masiga Molecular Biology and Biotechnology Department

Eastern Africa Barcode Workshop, Oct Insects

Eastern Africa Barcode Workshop, Oct Vector-transmitted pathogens Viruses Rickettsia Bacteria Protozoa helminths

Eastern Africa Barcode Workshop, Oct TDR Disease Focus 1.African trypanosomiasis 2.Chagas disease 3.Leishmaniasis 4.Dengue 5.Lymphatic filariasis 6.Malaria 7.Onchocerciasis 8.Schistosomiasis 9.Tuberculosis 10.Leprosy UNICEF/UNDP/World Bank/WHO - TDR TDR Disease Focus 1.African trypanosomiasis 2.Chagas disease 3.Leishmaniasis TDR Disease Focus 1.African trypanosomiasis 2.Chagas disease 3.Leishmaniasis 4.Dengue 5.Lymphatic filariasis 6.Malaria 7.Onchocerciasis 4.Dengue 5.Lymphatic filariasis 6.Malaria

Eastern Africa Barcode Workshop, Oct Distribution of human-infective trypanosomes causing sleeping sickness Animal-infective trypanosomes Are found throughout sub-saharan Africa, with the exception of the Southern tip.

Eastern Africa Barcode Workshop, Oct Well fed tsetse Barcoding tsetse flies and trypanosomes T. brucei in blood smear tsetse fly feeding

Eastern Africa Barcode Workshop, Oct Tsetse flies and trypanosomes There are about 31 species of tsetse flies, the only members of genus Glossina, are recognized. About 10 tsetse-transmitted trypanosome species and sub-species are known; probably >100 other species. Beyond Africa, T. evansi and T. vivax are transmitted mechanically (without parasite development in the vector)

Eastern Africa Barcode Workshop, Oct 18-19

Loci that have been used Tsetse:ITS 1 Trypanosomes:Satellite DNA; small-subunit ribosomal RNA (ssu rRNA) gene Barcoding tsetse and trypanosomes

Eastern Africa Barcode Workshop, Oct Identification of trypanosomes based on satellite DNA Satellite DNA is defined as “a portion of DNA in eukaryotes whose density differs from that of the majority of DNA and that consists of short, repeating sequences of nucleotide pairs (typically bp long).

Eastern Africa Barcode Workshop, Oct Trypanosome phylogeny (and ID) using ssu rDNA (18S rDNA locus)

Eastern Africa Barcode Workshop, Oct Current phylogeny is based on 18S rDNA phylogeny is based on 18S rDNA (taken from Hamilton et al., 2004)

Eastern Africa Barcode Workshop, Oct Can COI be used for barcoding trypanosomatids?

Eastern Africa Barcode Workshop, Oct mitochondrion glycosomes Inside a trypanosome

Eastern Africa Barcode Workshop, Oct Kinetoplast DNA (mitochondrial DNA) identical maxicircles (20-40 kb) encode 18 protein encoding (12 non- functional) genes, 2 rRNAs and a few guide (g) RNAs heterogeneous minicircles encode (over 900 different) small guide RNAs. Maxi- and minicircles are interlocked in a giant catenate. Dyskinetoplastic cells lack minicircles, maxicircles or both, e.g. T. evansi. Cannot be transmitted by Tsetse. EM of the periphery of an isolated kDNA network Network of minicircles condensed network Prof. Christine Clayton

Eastern Africa Barcode Workshop, Oct Some benefits of using COI A single locus of accessible size. Resolve classification of Glossina at sub-generic level (3 or 4 have been proposed: fusca, palpalis, austeni & morsitans). A more accurate indication of relationships and distances, perhaps providing explaining some key phenotypes, e.g. transmission efficiencies.

Eastern Africa Barcode Workshop, Oct Sandflies, vectors of Leishmaniasis Species of Leishmania L. major L. aethiopica L. infantum L. donovani L. tropica L. mexicana complex L. guyanensis L. naiffi L. braziliensis L. tarentolae E. scheideri Lutzomyia spp. (New World ) Phlebotomus spp. (Old World) Lutzomyia longipalpis lapinha TDR image Lib ID

Eastern Africa Barcode Workshop, Oct Insects

Eastern Africa Barcode Workshop, Oct Barcoding and epidemiology/disease control

Eastern Africa Barcode Workshop, Oct Global view of endemic regions for leishmaniasis With such a wide distribution, you can start small, but must network to advance.

Eastern Africa Barcode Workshop, Oct NHGRI Awards $13M for Next Generation Sequencing Technologies [October 4, 2006] NEW YORK (GenomeWeb News) – The National Human Genome Research Institute has awarded eleven grants worth a total of more than $13 million to develop technologies that will lower the cost of DNA sequencing. The agency has a near-term goal of lowering the costs of sequencing a mammalian-sized genome to $100,000, and to eventually cut the cost of whole- genome sequencing to $1,000 or less. Lowering the cost will make genome sequencing possible as part of routine medical care, NHGRI said. Nine grants fund researchers developing technologies that will make the $1,000 genome possible and two grants for researchers working on the $100,000 mammalian genome: