This document summarizes research on a dengue outbreak that occurred in Madeira Island from 2012-2013. Key findings include: 1) The first recorded presence of the mosquito Aedes aegypti in Madeira was in 2005. It has since rapidly dispersed in the southern part of the island. 2) From 2012-2013, there was a major dengue outbreak with over 2,000 probable cases reported across 3 municipalities. No fatalities occurred but 128 people were hospitalized. 3) Entomological studies found that Ae. aegypti in Madeira has high resistance to multiple insecticides. Genetic analysis indicated the population likely originated from a single introduction event from South America.

1. Sousa C.A.*, Seixas G., Novo M.T.,
Vacas D. , Clairouin M. , Viveiros B.
Magalhães M., Baptista M., Valente C.,
Gonçalves Y. , Gonçalves Silva J.J.,
Antunes L.C. , Araújo D., Silva A.C
Dengue outbreak in
Madeira island
* casousa@ihmt.unl.pt

2. Madeira: the garden island
• For Aedes aegypti too…

3. Aedes aegypti in Madeira Island
• 2005: First record of the mosquito– Funchal (St.Luzia)
• Rapid dispersal in the southern part of the island

4. Vector Monitoring
Since 2005-6, IHMT is participating in mosquito related activities
with local health authorities (IASAUDE)
• Identification of mosquito presence in the island.
• Bionomics studies
• Surveillance/monitoring activities
• Insecticide susceptibility bioassays (WHO)
• Genotpying of knockdown resistance (kdr) mutations
– AS-PCR and sequencing
• Population structure and history (mtDNA and microsatellites)

5. • Seasonality pattern
Aedes aegypti bionomics in
Madeira

6. • Diurnal biting activity
• Exo/endophagic species
Aedes aegypti
bionomics in Madeira
0
0.5
1
1.5
2
2.5
6h 7h 8h 9h 10h 11h 12h 13h 14h 15h 16h 17h 18h 19h 20h 21h 22h 23h 00h 1h 2h 3h 4h 5h
MeanfemaleAe.aegypti/collector
Hours
Daily female Ae. aegypti indoors biting pattern
0
0.5
1
1.5
2
2.5
6h 7h 8h 9h 10h 11h 12h 13h 14h 15h 16h 17h 18h 19h 20h 21h 22h 23h 00h 1h 2h 3h 4h 5h
MeanfemaleAe.aegypti/collector
Hours
Daily female Ae. aegypti outdoors biting pattern

7. • Man-biting rates
Aedes aegypti bionomics in
Madeira

8. Aedes aegypti bionomics in Madeira
Pictures: ECDC -2012
 Main breeding sites
– Peridomestic containers
– Abandoned houses (?!?!)

9. • Main breeding sites
– Storm drains.
Aedes aegypti bionomics
Total number of storm drains (ST) 434
% of dry ST 56,6%
% of negative ST 27,9%
% of ST with mosquito immature 15,7%
% of ST with Ae. aegypti 12,2%
% Absolute breeding index for aegypti 0,28

10. WHO insecticide resistance tests- 2009
 DDT 4.0% - 33.3% mortality
 Malathion 5.0% - 99.0% mortality (diagnostic dosage 0.8% /1h- 2012)
 Permethrin 0.75%, - 29.4% mortality
 Deltamethrin 0.05% - 65.2% mortality
Detection of 4 kdr mutations (N=35)
 Presence of only one mutation, V1016I
 In homozygosity, in an one DDT-resistant individual
 In heterozygosity, in 2 permethrin-resistant individuals

11. Origin of A. aegypti population
One (?) single
haplotype for both COI
and ND4 (N=15)
Distribution of kdr
mutation V1016I
Probable origin of
Ae. aegypti
Madeira
Venezuela
México
Senegal
BrazilPeru
EUA
Guatemala
Colonization by insecticide
resistant mosquitoes
Stream traffic of
people and goods
mtDNA analysis

12. Dengue outbreak in Madeira

13. Dengue outbreak in Madeira
• First transmission areas – St. Luzia and Nazaré

14. • Affected areas according to patients residence
Dengue outbreak in Madeira

15. • Affected areas according to patients residence
Dengue outbreak in Madeira

16. Dengue outbreak in Madeira
Number of disease cases between week 39 of 2012 and week 3 of 2013
Alert
Outbreak
Sporadic cases
Sporadic cases

17. Outbreak outline
• W39 2012 –W06 2013 : 20 weeks
• Probable cases : n= 2168
– 3 municipalities : n= 2069 cases 95 % cases
• Sex ratio (M/F) = 0.69
• 128 hospitalizations
• NO FATALITIES
Dengue outbreak in Madeira
Female
Male

18. • Incidence rate/ gender/ age group
Outbreak outline
39.3
59.9
110.1
128.3
70.6
70.9
73.5
68.5
57.3
72.1
75.9
54.8
69.2
67.8
77.9
25.5
34.7
45.6
31.9
49.4
78.4
117.6
90.8
82.4
96.3
92.0
111.1
108.2
132.7
118.9
113.3
73.0
68.0
70.8
55.1
39.9
0.0 20.0 40.0 60.0 80.0 100.0 120.0 140.0
0-4
5-9
10-14
15-19
20-24
25-29
30-34
35-39
40-44
45-49
50-54
55-59
60-64
65-69
70-74
75-79
80-84
85+
Taxa de incidência (/10.000 hab)
Grupoetário(anos)
F
M

19. Outbreak outline
.000
10.000
20.000
30.000
40.000
50.000
60.000
70.000
80.000
90.000
100.000
Febre
Mialgia
Cefaleia
Exantema
Dor retro Orbitária
Altralgia
Manifestações Hemorrágicas
Diarreia
Positivos Negativos
• Lab : 3285 samples analysed

20. • 82 dengue cases in 14 countries imported from Madeira

21. Outbreak response activities
• Epidemiological monitoring and information gathering
- Case definition for surveillance, epi-bulletin.
• Prevention and treatment
- Door-to-door campaigns and residence visits, public awareness
- Vector monitoring and control
- Specific dengue patient care management (severe and non-severe
cases)
- Blood safety: challenge at the start o the outbreak (PCR)
• Intra/inter-sectorial collaboration
• Communication plan: different media / prevention and control breeding
sites
• Internal and external evaluation

22. Epidemiological monitoring and
information gathering
Epi-bulletin

23. Madeira Dengue Surveillance System (MDSS)
Hospitals and public health centres
E- Bulletin
Private health centres
Web form
Bi-weekly extraction during the outbreak
Weekly extraction on-going
Data Management
IASAUDE,I-RAM- DSPAG/UTEIS/UES
CC DROTA
Time Person Place
Stakeholders
Regional
Nacional
International Authorities
Time-space analyse
Vector control Vector monitoring
activities

24. Intra/inter-sectorial collaboration
INSA

25. • Aedes aegypti monitoring during the outbreak
– 16 Bg-sentinel traps in the most affected areas.
– 141 ovitraps located in Aedes aegypti distribution area.
– Door-to-door larval sampling in the areas with higher
number of disease cases
Vector monitoring

26. 0
0.5
1
1.5
2
2.5
3
3.5
4
Mean number of Ae. aegypti females collected from
January 2012 to January 2013 on BG traps
Female aegypti
Bimensal work basis Daily work basis
Vector monitoring

27. 0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
0
50
100
150
200
250
Population dynamics of Aedes aegypti
Mean eggs/trap Mean Aegypti F
Vector monitoring

28. • Door-to-door activities - Educational Health
program
• Breeding sites reduction
• Measures for personal protection against aegypti bites and
virus infection
– Monitoring and elimination of breeding sites
• Infestation indexes
St. Luzia
12-11-2012
Vector monitoring (and control)

29. • Infestation indexes (2009)
Funchal
Câmara de
Lobos
St Cruz All counties
N. houses surveyed 273 125 22 420
N. of containers inspected
1681 1298 431 3410
House Index (%) 32.9 12.0 45.5 27.4
Container Index (%) 12.8 3.8 3.5 8.2
• Abandoned/closed
houses

30. • Educational campaigns for aegypti source
reduction
Vector control during Dengue
outbreak
“She
hanging
around!”
Flyers and posters
Internet update information

31. • Educational campaigns for aegypti source
reduction
Vector control during Dengue
outbreak
TV spot
RTP Madeira
2012

32. • Educational campaigns for aegypti source
reduction
– TV and Radio Programs
– 11 Radio-spots
 Information on airports and passenger
harbour (5 languages)
Vector control during Dengue
outbreak
 Information to tourists in
hotels and other
accommodation facilities

33. –Increased mosquito surveillance in airport
and harbours
–Cleaning of public artificial lakes and
ponds and dispersal of carnivorous
autochthonous fishes
–Flushing of storm drains with sea water.
Vector control during Dengue
outbreak

34. –Flushing of storm drains with sea water.
Vector control during Dengue
outbreak

35. –Application of larvicides and adulticides
at specific locations:
Vector control during Dengue
outbreak
Hospital Dr. Nélio Mendonça
Centro de saúde Bom Jesus
Escola Secundária
Francisco Franco

36. Impact of vector control measures
0
50
100
150
200
250
300
350
400
450
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
40_12 41_12 42_12 43_12 44_12 45_12 46_12 47_12 48_12 49_12 50_12 51_12 52_12 01_13 02_13 03_13 04_13 05_13 06_13 07_13 08_13 09_13
Aedes aegypti adult densities and dengue
cases
Mean Aegypti F Dengue cases

37. Post-outbreak activities
• On-going heath education programs
• Search for an efficient mosquito
adulticide
• Studies on resistance mechanisms
• Alternative strategies for vector control

38. • Highly resistant Aedes aegypti population to pyrethroids,
carbamates, organochlorines and organophosphates

39. • F1534C mutation is fixed (0.98-1.00)
• V1016I mutation is increasing (0.07-0.17)
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
2009 2012 2013
V1016I F1534C
(66) (87)
(91)(66)
(69)
(65)

40. Metabolitic resistance
E n zy m a tic p ro file o f a lfa es teras e ac tivity
P o p u la tio n s o f A e . a e g y p ti te s te d
nmol/mgptn/min
R
o
c
k
e
fe
lle
r
F
u
n
c
h
a
l
P
a
ú
l
d
o
M
a
r
0
5 0
1 0 0
1 5 0
2 0 0
E n zy m a tic p ro file o f b eta es te ra s e ac tiv ity
P o p u la tio n s o f A e . a e g y p ti te s te d
nmol/mgptn/min
R
o
c
k
e
fe
lle
r
F
u
n
c
h
a
l
P
a
ú
l
d
o
M
a
r
0
5 0
1 0 0
1 5 0
2 0 0

41. Alternative strategies for vector control
• Auto dissemination of Pyriproxifen
(juvenile hormone): DENFREE
consortium
– Pilot study in Paúl do Mar

42. Alternative strategies for vector control
• Water treatment in cemeteries with Bti (mosquito
biocide)

43. Alternative strategies for vector control
• Evaluation of salt as a control strategy for storm
drains treatment.
– Funchal county

44. Remarks & perspectives
• No autochthonous dengue cases reported in Madeira since February
2013
• Well established recently introduced Aedes aegypti population
– Distribution appears to be regressing (Central/south)
• mtDNA analysis inconclusive
– Single introduction?
– South American (Venezuela) origin?
– Human migratory flux with Brazil and Venezuela
• High levels of resistance to pyrethroid insecticide
– Environmental control (flushing storm drains w/ sea water )
– Alternative insecticides (ongoing pyriproxyfen pilot-study)
– Genetic control?

45. Obrigada