Structuration des populations d'Anopheles funestus et leur capacité vectrice - Conférence de la 1ère édition du Cours international « Atelier Paludisme » - COSTANTINI Carlo

1. Structuration des populations
d'Anopheles funestus et leur
capacité vectrice
Carlo Costantini
Fondazione Istituto Pasteur - Cenci Bolognetti et Dept. Santé Publique, Section Parasitologie,
Univ. Rome “La Sapienza”, Italie
Centre National de Recherche et Formation sur le Paludisme, Ouagadougou, Burkina Faso
carlo.costantini@uniroma1.it
costantini.cnlp@fasonet.bf

2. The Funestus group
funestus
parensis
sous-groupe
funestus
aruni
confusus
veneedeni
Groupe fuscivenosus
funestus
leesoni
rivulorum
rivulorum-brucei
brucei
Figure 1 : Le groupe funestus d'après GILLIES & DE M EILLON
(1968) et GILLIES & COETZEE (1987)

3. Adult bionomics
•Anthropophilic
•Endophagic
•Endophilic
•Longeve

4. Larval habitat
Large permanent water bodies, e.g.
Swamps
Lake margins
Rice fields
Ditches
Pools

5. Epidemiological relevance
•Entomological Inoculation Rates up to ~ 400 ibpy
•Extend transmission into the dry season
Population trend in Noungou (Burkina Faso) - Sampling by CDC light traps
Rainfall An. gambiae complex An. funestus
60 1000
50
mosquitoes/trap
40 100
rain (mm)
30
20 10
10
0 1
17-7 31-7 14-8 28-8 11-9 25-9 9-10 23-10 6-11 20-11

6. Chromosomal polymorphism
•No. chromosomes (2n) = 3
•No. paracentric inversions = 15
20 µ
X
a
s
t
2 4
a b a
3 5

7. Chromosomal inversions
Protection of gene linkages
from crossing over
A. Appaiamento di cromatidi in un
eterozigote per inversione
B. L’inversione non C. L’inversione
include il centromero include il centromero
Prima anafase meiotica Prima anafase meiotica

8. Methodology
•Observational studies
•Formulation of working hypothesis
•Hypothesis validation

9. Latitudinal transect

10. Sampling
Residual fauna
•Indoors
(spray-sheet catches,
manual aspiration)
•Outdoors
(pit-shelters)

11. Assortative Mating
Fis statistic <0 =0 >0

12. Hardy-Weinberg equilibrium
Village Chromosomal Arm 3
2N +/+ +/a a/a f(3a) FIS P
November 1992
Tougouri 50 0.72 0.16 0.12 0.20 0.52 0.028
Noungou 642 0.80 0.09 0.11 0.16 0.62 <0.0001
Loumbila 114 0.72 0.12 0.15 0.24 0.67 <0.0001
Lembouroudougou 78 0.90 0.03 0.08 0.09 0.85 <0.001
Kiribina 100 0.86 0.04 0.10 0.12 0.81 <0.0001
Diarabakoko 82 0.54 0.12 0.34 0.40 0.75 <0.0001
Folonzo 38 0.11 0.11 0.79 0.84 0.62 0.035
November 1994
Loumbila 48 0.58 0.13 0.29 0.35 0.74 <0.001
Kiribina 84 0.76 0.12 0.12 0.18 0.60 <0.001
Diarabakoko 132 0.38 0.11 0.52 0.57 0.79 <0.0001
Folonzo 92 0.02 0.24 0.74 0.86 0.03 ns
Costantini et al. (1999) Parassitologia 41: 595-611

13. Hardy-Weinberg equilibrium
Sample Chromosomal Arm 3
2N +/+ +/a a/a f(3a) FIS P
Noungou Peul
Endophilic 16 0.500 0.500 0.000 0.250 -0.27 ns
Exophilic 18 0.889 0.111 0.000 0.056 —
Noungou
Endophilic 1134 0.799 0.106 0.095 0.148 0.58 <0.0001
Exophilic 384 0.849 0.073 0.078 0.115 0.64 <0.0001
Loumbila
Endophilic 316 0.652 0.139 0.209 0.278 0.66 <0.0001
Exophilic 92 0.848 0.065 0.087 0.120 0.70 <0.001
Costantini et al. (1999) Parassitologia 41: 595-611

14. Chromosomal forms Hp
“Kiribina”
-> low-level polymorphism for 2a, 2s, 3a, 3b, 5a
“Folonzo”
-> high-level polymorphism for 2a, 3a, 3b, 5a
Costantini et al. (1999) Parassitologia 41: 595-611

15. Validation: new study area

16. Validation: new sampling plan
Longitudinal survey:
•Weekly (3x) indoor- and outdoor
resting collections
•3 breeding seasons (Sep to Mar)
~15,000 ovaries of which ~2,500
karyotyped

17. Longitudinal survey
Temporal pattern of inversion frequencies (bootstrapped mean ±95% confidence limits)
Inversion Inversion
0.7 0.5
0.6
0.4
0.5
0.4 0.3
0.3 0.2
0.2
0.1
0.1
0.0 0.0
Inversion Inversion
0.5 0.5
0.4 0.4
0.3 0.3
0.2 0.2
0.1 0.1
0.0 0.0

18. Hardy-Weinberg equilibrium
INDOOR samples stratified by breeding season
Season Locus Koubri Kuiti
Fis P Fis P
1999-2000 2a-s +0.303 0.00 +0.387 0.00
3a +0.699 0.00 +0.732 0.00
3b +0.466 0.00 +0.434 0.00
5a +0.232 0.01 Š0.023 1.00
2000-2001 2a-s +0.335 0.00 +0.279 0.00
3a +0.660 0.00 +0.719 0.00
3b +0.375 0.00 +0.358 0.00
5a +0.218 0.01 Š0.006 1.00
2001-2002 2a-s +0.279 0.00 +0.319 0.00
3a +0.815 0.00 +0.766 0.00
3b +0.321 0.00 +0.372 0.00
5a Š0.004 1.00 Š0.005 1.00

19. Hardy-Weinberg equilibrium
OUTDOOR samples stratified by month of collection
Arrangement 2Ra-s Arrangement 3Ra Arrangement 3Rb Arrangement 3La X2 Df P
Sample N Ho He Fis P N Ho He Fis P N Ho He Fis P N Ho He Fis P
Koubri
Sep-99 19 4 5.4 0.27 0.06 20 4 6.6 0.40 0.13 17 1 2.8 0.65 0.09 17 1 1.0 - - 14.6 6.0 0.02
Oct-99 47 6 12.2 0.51 0.00 48 3 15.4 0.81 0.00 48 5 8.2 0.40 0.04 45 1 1.0 - - 47.9 6.0 0.00
Jan-00 15 - - - - 15 3 4.3 0.31 0.33 15 2 1.9 -0.04 1.00 13 - - - - 2.2 6.0 0.69
Nov-00 16 2 3.7 0.47 0.19 18 1 5.8 0.83 0.00 18 1 5.8 0.83 0.00 15 1 1.0 - - 26.6 6.0 0.00
Oct-01 27 3 2.9 - 1.00 31 3 12.5 0.76 0.00 28 0 3.8 1.00 0.00 26 - - - - 33.3 6.0 0.00
Kuiti
Oct-99 50 8 11.0 0.28 0.02 50 7 18.9 0.63 0.00 48 6 10.6 0.44 0.02 49 2 2.0 0.0 1.0 36.8 6.0 0.00
Jan-00 13 2 1.9 -0.04 1.00 16 2 5.0 0.61 0.05 16 2 1.9 -0.03 1.00 14 - - - - 6.0 6.0 0.42
Oct-00 16 2 3.7 0.47 0.07 20 1 2.8 0.66 0.08 20 1 2.8 0.66 0.08 15 - - - - 15.7 6.0 0.02
Oct-01 13 - - - - 13 0 4.8 1.00 0.00 12 2 1.9 -0.05 1.00 8- - - - 13.4 6.0 0.01

20. Hardy-Weinberg equilibrium
Stratified by compound of collection
2
Arrangement 2Ra-s Arrangement 3Ra Arrangement 3Rb Arrangement 3La X Df P
Compound N Ho He Fis P N Ho He Fis P N Ho He Fis P N Ho He Fis P
Koubri Sep. 99
1 39 7 9.9 0.30 0.01 41 2 7.3 0.73 0.00 41 4 5.6 0.29 0.18 - - - - - 30.9 6 0.00
2 40 10 9.2 -0.09 1.00 40 2 10.3 0.81 0.00 40 2 5.6 0.65 0.01 - - - - - 22.8 6 0.07
9 42 5 6.6 0.25 0.11 43 3 8.2 0.64 0.00 43 1 4.8 0.79 0.00 - - - - - 29.0 6 0.00
Dec. 01
15 108 25 33.7 0.26 0.00 122 10 44.4 0.78 0.00 117 14 26.3 0.47 0.00 107 1 1.0 - - 110.4 6 0.00
17 120 29 42.9 0.33 0.00 134 5 45.0 0.89 0.00 129 18 28.2 0.36 0.00 114 5 8.7 0.4 0.01 161.8 8 0.00
27 70 14 24.0 0.42 0.00 82 5 26.5 0.81 0.00 76 9 13.6 0.34 0.02 73 4 3.9 0.0 1.00 77.0 8 0.00
55 62 9 12.0 0.25 0.03 65 0 5.8 1.00 0.00 62 2 2.0 -0.01 1.00 59 - - 30.8 6 0.00
Kuiti Sep. 99
1 119 21 32.7 0.36 0.00 126 12 26.5 0.55 0.00 123 12 18.4 0.35 0.00 - - - - - 66.9 6 0.00
7 52 11 15.1 0.27 0.00 53 8 13.7 0.42 0.01 52 8 7.5 -0.07 1.00 - - - - - 22.2 6 0.00
12 45 10 16.6 0.40 0.00 50 5 16.8 0.70 0.00 47 4 10.6 0.62 0.00 - - - - - 49.9 6 0.00
Dec. 01
6 102 21 34.3 0.39 0.00 109 9 44.1 0.80 0.00 108 20 28.8 0.31 0.00 104 3 6.8 0.6 0.00 115.8 8 0.00
7 79 19 26.2 0.28 0.00 81 7 28.7 0.76 0.00 78 13 16.8 0.23 0.08 74 2 1.9 0.0 1.00 61.5 8 0.00
49 80 16 31.2 0.49 0.00 81 5 26.4 0.81 0.00 79 7 16.8 0.59 0.00 73 1 1.0 - - 104.7 6 0.00
53 97 14 21.9 0.36 0.00 100 4 16.5 0.76 0.00 97 4 11.3 0.65 0.00 93 3 3.0 0.0 1.00 70.1 8 0.00

21. Linkage disequilibrium
Stratified by breeding season
Period Locus pair NS S Test number
Sep. 99 2Ra-s vs 3Ra 0% 100% 7
Period Locus pair NS S Test number 2Ra-s vs 3Rb 0% 100% 7
Season1 2Ra-s vs 3Ra 8% 92% 13 3Ra vs 3Rb 0% 100% 7
2Ra-s vs 3Rb 15% 85% 13 Dec. 01 2Ra-s vs 3Ra 0% 100% 8
2Ra-s vs 3La 80% 20% 10 2Ra-s vs 3Rb 13% 88% 8
2Ra-s vs 3La 71% 29% 7
3Ra vs 3Rb 0% 100% 13 3Ra vs 3Rb 0% 100% 8
3Ra vs 3La 90% 10% 10 3Ra vs 3La 43% 57% 7
3Rb vs 3La 90% 10% 10 3Rb vs 3La 57% 43% 7
Season2 2Ra-s vs 3Ra 0% 100% 13
2Ra-s vs 3Rb 8% 92% 12
2Ra-s vs 3La 57% 43% 7 Stratified by compound of collection
3Ra vs 3Rb 0% 100% 12
3Ra vs 3La 43% 57% 7
3Rb vs 3La 57% 43% 7
Season3 2Ra-s vs 3Ra 8% 92% 13
2Ra-s vs 3Rb 23% 77% 13
2Ra-s vs 3La 88% 13% 8
3Ra vs 3Rb 0% 100% 13
3Ra vs 3La 56% 44% 9
3Rb vs 3La 67% 33% 9

22. Test of Algorithm
Season Form Locus Koubri Kuiti
Maximum Likelihood
1999-2000 Folonzo 2a-s
Fis
+0.130
P
0.20
Fis
+0.215
P
0.01 16
Assignment Test
3a Š0.131 0.22 +0.122 0.12 15
14
3b +0.087 0.38 +0.115 0.14 13
5a +0.126 0.26 +0.077 0.36 12
Kiribina 2a-s +0.008 0.16 Š0.041 0.34 11
3a +0.068 0.30 +0.090 0.19 10
9
3b Š0.004 1.00 Š0.007 1.00
8
5a Š0.005 1.00 Š0.010 1.00 7
2000-2001 Folonzo 2a-s +0.274 0.04 +0.133 0.28 6
3a Š0.155 0.34 Š0.029 1.00 5
3b Š0.028 1.00 Š0.180 0.10 4
3
5a Š0.089 1.00 Š0.017 1.00 2
Kiribina 2a-s +0.076 0.27 Š0.014 0.45 1
3a Š0.031 1.00 Š0.024 1.00 0
3b Š0.005 1.00 Š0.004 1.00 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16
5a +0.437 0.00 Š0.003 1.00 Deviance Folonzo
2001-2002 Folonzo 2a-s +0.160 0.15 +0.168 0.05
3a +0.104 0.20 Š0.047 0.74
3b Š0.114 0.22 Š0.022 0.88
5a Š0.008 1.00 Š0.009 1.00 99.5% of 395 individuals
Kiribina 2a-s +0.094 0.14 +0.023 0.67
3a Š0.016 1.00 Š0.018 1.00 assigned according to the
3b Š0.002 1.00 Š0.004 1.00 algorithm
5a Š0.002 1.00 Š0.001 1.00

23. Temporal distribution
Folonzo Kiribina
Koubri
Kuiti Mar
Mar
Mar
Nov
Nov
Nov
Jan
Sep
Jan
Sep
Jan

24. Spatial distribution
Indoor resting samples collected over one week
• 52 sampled compounds (~ 50% of whole village)
• 812 scorable karyotyped specimens
Kriging of inversion 3a frequency
Latitude
Longitude

25. Epidemiological parameters
Sample N Chromosomal Form
Folonzo Kiribina
Frequency HBI CSP +ve Frequency HBI CSP +ve
Noungou
Endophilic 533 (528) 0.167 0.921 0.146 0.833 0.948 0.110
Exophilic 164 (159) 0.134 0.909 0.045 0.866 0.628 0.063
Pooled 697 (687) 0.159 0.919 0.126 0.841 0.872 0.099
Loumbila
Endophilic 154 (153) 0.292 0.822 0.089 0.708 0.750 0.055
Exophilic 44 (42) 0.182 0.500 0.125 0.818 0.441 0.028
Pooled 198 (195) 0.268 0.774 0.094 0.732 0.676 0.048

26. New approaches: STRs
32 microsatellite loci mapped by FISH dispersed over all chromosomes
Set of 16 located
OUTSIDE main
inversions chosen for
metapopulation analyses

27. The team
Centre National de Recherche et Formation sur le Paludisme, Burkina Faso
•N’Falé Sagnon
•Wamdaogo Moussa Guelbeogo
Istituto Superiore di Sanità, Laboratorio di Parassitologia, Italy
•Daniela Boccolini
Dept. Biological Sciences, Univ. Notre Dame, USA
•Nora Besansky
•Olga Grushko
•Igor Sharakhov
•Brandon Hackett Schemerhorn
•Andy Michel
•Matt Ingrasci