Update on Glossina palpalis genomics at IRD/ Genoscope

and

Summarising recent results on Genetics and Morphometrics of Glossina palpalis

in West Africa

S. Ravel1, P. Wincker2,P. Solano1,3, T. de Meeüs1, JP. Dujardin1, G. Cuny1

1: IRD, MONTPELLIER, France

2: Genoscope, PARIS, France

3: CIRDES BOBO-DIOULASSO, Burkina Faso

1. Update on G. palpalis genomics at IRD/Genoscope

Dissection of midgut and salivary glands from Glossina palpalis gambiensis colony maintained at Montpellier, France

15 teneral males

+

15 males at day 2 after bloodmeal

+

15 males at day 4 after bloodmeal

45 males

15 teneral females

+

15 females at day 2 after bloodmeal

+

15 females at day 4 after bloodmeal

45 females

Midgut pool Salivary glands pool

=

Total RNA Extraction from midgut pool and salivary gland pool

Midgut pool Salivary glands pool

Nucleospin RNA II kit (clontech)

Total RNA from midgut

91 g

Total RNA from salivary glands

14 g

cDNA libraries (from midgut and from salivary glands) construction using creator smart cDNA library construction kit

(clontech)

2. Summarising recent results on genetics and morphometrics of G. palpalis in West Africa

Les glossines ou mouches tsé-tsé. Logiciel d’identification et d’enseignement. ORSTOM/CIRAD. ORSTOM editions, coll. Didactiques, 1998

Bonon, Côte d’Ivoire, 2006- D. KABA

G. p. palpalis in Côte d’IvoireBonon, Center-West CI/ 1200 mm rainfall/ Gpp/ 5 microsat loci

High and significant Fis, Wahlund effect due to spatial clustering on 2 temporal samples

Fis values estimated for both samples 1 and 2 G. palpalis palpalis before (November and January) and after BAPS partitioning of the data (clusters). 95% confidence intervals were obtained by bootstraping over loci

Explanation: Null Alleles at microsatellite loci + short allele dominance+ Wahlund effect due to spatial clustering of tsetse: 20 to 25 subpops of small size having different hosts and being differentially infected

New morphometric markers: Geometric Morphometrics

Morphometric techniques based on distances, or ratios, between anatomical points called landmarks are referred to as “traditional morphometrics”

Newer approaches based on the coordinates of these landmarks in a given system of orthogonal axes, and superimposition procedures are referred to as “geometric morphometrics”. It is now possible to work on both the size (centroid size) and the shape, this latter being likely to reflect genetic changes, tentatively independents from the size and showing good heritability (see for instance: www.mpl.ird.fr/morphometrics)

Geometric morphometrics: principle

scaling Translation rotation

Procrustes superimposition: an iterative least square adjustment of all the figures after size normalization (scaling). Procrustes superimposition algorithms translates and rotates the landmark configurations to minimize the squared differences between landmarks. Comparisons from the polygon obtained with a consensus polygon are therefore obtained.

Analysis: differences from consensus

Geometric morphometry: intraspecies analysis, G. palpalis

G. p. gambiensis in the coastal area of Guinea

Gpg/ focus of Dubreka and Loos islands/ 4000 mm rainfall/

Molecular (4 msat loci) and morphometric (11 LM on wings) analyses

UPGMA tree on genetic distances based on wing morphometry (on the left, Mahalanobis distance) and on microsatellite DNA loci (on the right, Cavalli-Sforza and Edwards distance) of the three tsetse populations.

« Data from microsatellites and from wing geometry both converged to the idea of a separation of the Loos island population from the mainland. The level of separation in terms of number of migrants per generation seems high, which conforms to the known dispersing behavior of the insect. Although occasional contacts cannot be excluded, our working hypothesis is that the Loos population of tsetse flies is a completely isolated population. »

Tsetse Elimination on the « Iles de Loos »: Available data after 2 small surveys (LTTRN mai 2005-mai 2006) and the exhaustive entomological

survey of October 2006 :

0.3 à 1 Gpg/piège/jour

10 à 100 Gpg/piège/jour

5 à 10 Gpg/piège/jour

1 à 5 Gpg/piège/jour

Pas de glossine

Résultats des enquêtes entomologiques :

638 glossines (Gpg) capturées

Densité supérieure à Kassa (10 Gpg/piège/jour) qu’à Fotoba (1 Gpg/piège/jour)

Position des pièges sentinelles pour le monitoring de la phase de suppression

Composante financement 1. Recensement exhaustif et individuel des populations des îles de Loos IRD-PNLTHA 2. Recensement et décompte des animaux domestiques d’élevage IRD-PNLTHA 3. Enquête entomologique avant lutte LTTRN- IRD- PNLTHA 4. Prospection médicale exhaustive OMS 5. Enquête Trypanosomose animaux domestiques IRD-CIRDES-DNE 6. Traitement des cochons par insecticide en « pour on » BAYER-LTTRN-SCAC-DNE 7. Eclaircissement sélectif de la végétation PNLTHA et pop locale 8. Déploiement d’écrans et de pièges imprégnés d’insecticide VESTERGAARD-LTTRN 9. Pulvérisation insecticide au sol JICA-PNLTHA 10. Suivi de l’évolution des densités (durée 2 ans minimum) IRD-LTTRN-SCAC-INCO

Elimination des tsé-tsé des îles de Loos : principales étapes

1. Phase de suppression : Etapes initiales (octobre à décembre 2006)

2. Octobre-novembre 2007 : Atelier international sur les résultats partiels et stratégies à mettre en œuvre (financement LTTRN) pour la suite

3. Phase d’élimination

4. Simultanément, discussions sur possibilités d’intervention contre autres maladies à vecteurs (paludisme, arboviroses…), en collaboration avec Ministère de la Santé- OMS- IRD- Vestergaard

DONE

G. p. gambiensis in humid savannah of Burkina FasoPopulation structuring of Glossina palpalis

gambiensis (Diptera: Glossinidae) according to landscape fragmentation in the Mouhoun river,

Burkina FasoJeremy Bouyer, Sophie Ravel, Jean-Pierre Dujardin, Thierry de Meeüs, Laurence Vial, Sophie Thévenon,

Laure Guerrini, Issa Sidibé, Philippe Solano.

In prep.

30 km

0 15

Trapping sites

1

2

3

4

■ Swamp forests

River course

Boromo

Bobo Dioulasso

Mouhoun river

Gpg/ Mouhoun river («Volta Noire»)/ area for PATTEC/ 4 pop/ ecological clines: rainfall (S-N, 1000mm) and guinean to sudanian

Msat loci (5) and wings morphometry (11 LM)

Data from microsatellites and from wing geometry both converged to the idea of a structuring of G. palpalis gambiensis along the Mouhoun river.

However, the structuring pointed out by morphometrics appeared more important than the slight (although significant) Fst value detected with microsatellites. Metric differences suggested isolation by distance (in females); genetic results did not detect it: may be due to the fact that the geographic distance is superimposed to an “ecological distance” due to a double cline in both rainfall and riverine forest ecotype, from the south to the north.

In the present situation, it is however not possible to conclude to a complete isolation of population 1, and the PATTEC campaign should consider treating all the western branch of the Mouhoun river as one population exchanging migrants.

Fis=+0.11***

Fst=+0.01*

Mean of 3538 flies/site

Les glossines ou mouches tsé-tsé. Logiciel d’identification et d’enseignement. ORSTOM/CIRAD. ORSTOM editions, coll. Didactiques, 1998