western and central africa dna barcoding meeting: abuja, nigeria dna barcoding for disease vectors...

Western and Central Africa DNA BarcodingWestern and Central Africa DNA Barcoding meeting: Abuja, Nigeria meeting: Abuja, Nigeria

DNA barcoding for disease DNA barcoding for disease vectors vectors

Daniel Adjei BoakyeDaniel Adjei BoakyeParasitology DepartmentParasitology Department

Noguchi Memorial Institute for Medical Noguchi Memorial Institute for Medical ResearchResearch


Lymphatic filariasisLymphatic filariasis

elephantiasiselephantiasis


OnchocerciasisOnchocerciasis

Health and Socio-Health and Socio-economic impacteconomic impact


LeishmaniasisLeishmaniasis


Impact of parasitic diseasesImpact of parasitic diseases

Cause immense sufferingCause immense suffering Life-long disabilitiesLife-long disabilities Impair childhood growth and developmentImpair childhood growth and development Promote poverty, impair education andPromote poverty, impair education and Economic development Economic development

• Arable land desertedArable land deserted Population affectedPopulation affected MortalityMortality


Neglected Tropical Diseases and MalariaNeglected Tropical Diseases and MalariaDiseaseDisease VBDVBD

11 Buruli ulcerBuruli ulcer ??

22 ChagasChagas **

33 CholeraCholera ****

44 DengueDengue **

55 DracunculiasisDracunculiasis **

66 Human African Human African TrypanosomiasisTrypanosomiasis

**

77 LeishmaniasisLeishmaniasis **

88 LeprosyLeprosy

99 FilariasisFilariasis **

1010 OnchocerciasisOnchocerciasis **

1111 SchistosomiasisSchistosomiasis **

1212 Soil-Transmitted Soil-Transmitted HelminthiasisHelminthiasis

1313 TrachomaTrachoma ****

1414 MalariaMalaria **


DiseasesDiseases VectorsVectors AreasAreas

ChagasChagas TriatomaTriatoma S/AmericaS/America

DengueDengue MosquitoMosquito

(Ae)(Ae)

Tropical WorldTropical World

Guinea wormGuinea worm CyclopCyclop AfricaAfrica

H/TrypanosomiasisH/Trypanosomiasis GlossinaGlossina AfricaAfrica

LeishmaniasisLeishmaniasis PhlebotomusPhlebotomus Africa, America,Africa, America,

AsiaAsia

Lymphatic filariasisLymphatic filariasis MosquitoMosquito

(An,Cx,Ae,Mn)(An,Cx,Ae,Mn)


OnchocerciasisOnchocerciasis SimuliumSimulium Africa, AmericaAfrica, America

SchistosomiasisSchistosomiasis SnailSnail Tropical WorldTropical World

MalariaMalaria MosquitoMosquito

(An)(An)


Vectors of Neglected Tropical Diseases


Importance of correct identificationImportance of correct identification

Understanding the diversity of vector Understanding the diversity of vector species and their respective vectorial species and their respective vectorial importance is necessary for any importance is necessary for any targeted control efforttargeted control effort

Studies on Vector speciesStudies on Vector species• Identification and clarification of species Identification and clarification of species

statusstatus• Biology and ecology of vectorsBiology and ecology of vectors• Vectorial importance of different speciesVectorial importance of different species


Members of the Members of the S. damnosumS. damnosum complex in West Africa complex in West Africa

Determination of important vector Determination of important vector speciesspecies

Basis for mapping limits of control Basis for mapping limits of control programmeprogramme

Only larvae can be identified with Only larvae can be identified with precisionprecision


Anopheles gambiaeAnopheles gambiae species complex species complex

Anopheles gambiae sensu lato

An. gambaie s.s

An. arabiensis

An. melas An. merus

An. quadrannulatus

An. bwambae


Taxonomic challenges within Taxonomic challenges within An. gambiaeAn. gambiae s.s s.s

Genetic discontinuitiesGenetic discontinuities• from chromosomal from chromosomal

analysis (Coluzzi analysis (Coluzzi et alet al 1985)1985)

Five chromosomal forms Five chromosomal forms in West Africa: Mopti; in West Africa: Mopti; Bamako; Bissau; Forest; Bamako; Bissau; Forest; SavannaSavanna

• in the ribosomal DNA in the ribosomal DNA region (rDNA) (Favia region (rDNA) (Favia et alet al 1997)1997)

Two molecular forms Two molecular forms described (della Torre et described (della Torre et al 2001; 2005)al 2001; 2005)


Problems associated with current Problems associated with current identificationidentification

In Mali and Burkina Faso, the M and S forms In Mali and Burkina Faso, the M and S forms correspond to the Mopti and Savanna correspond to the Mopti and Savanna chromosomal forms chromosomal forms

Molecular forms do not always correspond to the Molecular forms do not always correspond to the chromosomal forms in other countrieschromosomal forms in other countries

The two molecular forms may be a reflection of The two molecular forms may be a reflection of the presence of two different species in what is the presence of two different species in what is considered as one species – considered as one species – An. gambiaeAn. gambiae s.s. s.s.


Practical consequences for disease Practical consequences for disease control control

Susceptibility to Susceptibility to insecticidesinsecticides

Differences in disease Differences in disease transmission transmission capabilitiescapabilities

Adaptation to different Adaptation to different breeding sitesbreeding sites

There is therefore the There is therefore the need to understand need to understand whether there are 2 or whether there are 2 or only 1 speciesonly 1 species - 5 - 4 - 3 - 2 - 1 0 1 2 3 4

5

6

7

8

9

10

11

12

13

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52 74

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1 22 01 8

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1 39

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Distribution of LF in Burkina Faso, Benin, Togo and Ghana

E stim a tedp rev a len ceo f ag (% )

0

1

2

3

5

10

30

50

B u rk in a F aso

B en in

T o g oG h an a


MethodologyMethodology

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)


MethodologyMethodology

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).


Preliminary resultsPreliminary results


Preliminary resultsPreliminary results

Molecular Molecular forms of An. forms of An. gambiae s.s.gambiae s.s.


Example of phylogenetic treeExample of phylogenetic tree

TE6

TE8

TE9

TE3

TE1

TE2

39 Ankwanda melas

41 Ankwanda melas

40 Ankwanda gambiae ss M form

17 Ankwanda gambiae ss S form

35 Ankwanda gambiae ss M form

69 (nice peaks) Ada melas

23 Ada gambiae ss M form

KP7 Kpone on sea gambiae ss S form

WA33

WA44

AK130

SA2

SA4

SA3

SA1

SA10

0.02

Sudan Savanna

Coastal Savanna

Semi Deciduous Forest

Savanna


Ghana Mosquitoes’ file in the Completed Project section of the Barcode of Life Ghana Mosquitoes’ file in the Completed Project section of the Barcode of Life

website (website (http://http://www.barcodinglife.orgwww.barcodinglife.org).).


ConclusionConclusion

DNA barcode can be used to DNA barcode can be used to • Separate out closely related members of Separate out closely related members of

sibling species in vectors of diseasessibling species in vectors of diseases• Draw relationships between species Draw relationships between species

collected from wider geographical areascollected from wider geographical areas• Determine the presence of individuals Determine the presence of individuals

currently unidentified using the currently unidentified using the available techniquesavailable techniques


AcknowledgementsAcknowledgements

Funding: Funding: • JRS Biodiversity FoundationJRS Biodiversity Foundation• World Health OrganizationWorld Health Organization• Gate’s FoundationGate’s Foundation

Collaborators at the NMIMRCollaborators at the NMIMR• Prof. Michael WilsonProf. Michael Wilson• Dr Maxwell AppawuDr Maxwell Appawu• Dziedzom de SouzaDziedzom de Souza• Hilaria AmuzuHilaria Amuzu• Ignatius NdongIgnatius Ndong


Thank youThank you

western and central africa dna barcoding meeting: abuja, nigeria dna barcoding for disease vectors...

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