Aedes aegypti on Madeira Island (Portugal): genetic variation of a recently introduced dengue vector

Goncalo Seixas,Paulo Eduardo Martins Ribolla,Patricia Salgueiro,Carla Alexandra Sousa,Melina Campos,Matias Reyes-Lugo,Joao Pedro Soares Da Silva Pinto,Maria Teresa Novo,Ana Clara Silva,Carine Spenassatto

doi:10.1590/0074-0276130386

Goncalo Seixas, Paulo Eduardo Martins Ribolla + Show 8 more

Open Access

https://doi.org/10.1590/0074-0276130386

Copy DOI

Abstract

The increasing population of Aedes aegypti mosquitoes on Madeira Island (Portugal) resulted in the first autochthonous dengue outbreak, which occurred in October 2012. Our study establishes the first genetic evaluation based on the mitochondrial DNA (mtDNA) genes [cytochrome oxidase subunit I (COI) and NADH dehydrogenase subunit 4 (ND4)] and knockdown resistance ( kdr ) mutations exploring the colonisation history and the genetic diversity of this insular vector population. We included mosquito populations from Brazil and Venezuela in the analysis as putative geographic sources. The Ae. aegypti population from Madeira showed extremely low mtDNA genetic variability, with a single haplotype for COI and ND4. We also detected the presence of two important kdr mutations and the quasi-fixation of one of these mutations (F1534C). These results are consistent with a unique recent founder event that occurred on the island of Ae. aegypti mosquitoes that carry kdr mutations associated with insecticide resistance. Finally, we also report the presence of the F1534C kdr mutation in the Brazil and Venezuela populations. To our knowledge, this is the first time this mutation has been found in South American Ae. aegypti mosquitoes. Given the present risk of Ae. aegypti re-invading continental Europe from Madeira and the recent dengue outbreaks on the island, this information is important to plan surveillance and control measures.

Highlights

Invasive mosquito species are a serious threat to the environment and human health
Gonçalo Seixas et al The most recent example of an epidemic driven by mosquito invasion is the dengue outbreak on Madeira Island (Portugal) that occurred in October 2012, with 1,891 cases (966 laboratory-confirmed) detected at the end of November 2012 (Sousa et al 2012)
Despite preliminary surveys suggesting a high degree of insecticide resistance (Seixas 2012, ECDC 2013), no information was yet available on the possible mechanisms underlying the resistance in the local Ae. aegypti population

Summary

MATERIALS AND METHODS

Mosquito samples and DNA extraction - Samples of Ae. aegypti were collected on Madeira Island and from two populations of Brazil and Venezuela (Figure, Table I). For the V1016I mutation, the kdr20f and kdr21r PCR primers were described in Saavedra-Rodriguez et al (2007), with the following cycling conditions: Fig. 1: map showing the geographic collection sites. PCR was performed in a 25 μL volume containing 5 μL of 1x buffer, 2 mM MgCl2, 0.4 mM dNTP, 1 μM of each primer, 1.5 U of Taq polymerase and 1 μL of DNA template diluted 1:10. PCR was performed in a 50 μL reaction volume containing 10 μL of 1x buffer, 1.5 mM MgCl2, 0.2 mM dNTP, 0.3 μM of each primer, 1 U of Taq polymerase and 2 μL of DNA template. The PCR cycling conditions included an initial denaturation step at 94oC for 2 min, followed by 35 cycles of 94oC for 30 sec, 54oC for 30 sec, 72oC for 1 min and a final extension of 72oC for 5 min. These sequences were compared with the reference Ae. aegypti gene sequences deposited in the GenBank

RESULTS

DISCUSSION

G T CGGGGCAGCTCCCT G