Aedes aegypti, the ‘yellow fever mosquito’, is the primary vector to humans of dengue and yellow fever flaviviruses (DENV, YFV), and is a known vector of the chikungunya alphavirus (CV). Because vaccines are not yet available for DENV or CV or are inadequately distributed in developing countries (YFV), management of Ae. aegypti remains the primary option to prevent and control outbreaks of the diseases caused by these arboviruses. Permethrin is one of the most widely used active ingredients in insecticides for suppression of adult Ae. aegypti. In 2007, we documented a replacement mutation in codon 1,016 of the voltage-gated sodium channel gene (para) of Ae. aegypti that encodes an isoleucine rather than a valine and confers resistance to permethrin. Ile1,016 segregates as a recessive allele conferring knockdown resistance to homozygous mosquitoes at 5–10 µg of permethrin in bottle bioassays. A total of 81 field collections containing 3,951 Ae. aegypti were made throughout México from 1996 to 2009. These mosquitoes were analyzed for the frequency of the Ile1,016 mutation using a melting-curve PCR assay. Dramatic increases in frequencies of Ile1,016 were recorded from the late 1990's to 2006–2009 in several states including Nuevo León in the north, Veracruz on the central Atlantic coast, and Yucatán, Quintana Roo and Chiapas in the south. From 1996 to 2000, the overall frequency of Ile1,016 was 0.04% (95% confidence interval (CI95) = 0.12%; n = 1,359 mosquitoes examined). The earliest detection of Ile1,016 was in Nuevo Laredo on the U.S. border in 1997. By 2003–2004 the overall frequency of Ile1,016 had increased ∼100-fold to 2.7% (±0.80% CI95; n = 808). When checked again in 2006, the frequency had increased slightly to 3.9% (±1.15% CI95; n = 473). This was followed in 2007–2009 by a sudden jump in Ile1,016 frequency to 33.2% (±1.99% CI95; n = 1,074 mosquitoes). There was spatial heterogeneity in Ile1,016 frequencies among 2007–2008 collections, which ranged from 45.7% (±2.00% CI95) in the state of Veracruz to 51.2% (±4.36% CI95) in the Yucatán peninsula and 14.5% (±2.23% CI95) in and around Tapachula in the state of Chiapas. Spatial heterogeneity was also evident at smaller geographic scales. For example within the city of Chetumal, Quintana Roo, Ile1,016 frequencies varied from 38.3%–88.3%. A linear regression analysis based on seven collections from 2007 revealed that the frequency of Ile1,016 homozygotes accurately predicted knockdown rate for mosquitoes exposed to permethrin in a bioassay (R² = 0.98). We have recorded a dramatic increase in the frequency of the Ile1,016 mutation in the voltage-gated sodium channel gene of Ae. aegypti in México from 1996 to 2009. This may be related to heavy use of permethrin-based insecticides in mosquito control programs. Spatial heterogeneity in Ile1,016 frequencies in 2007 and 2008 collections may reflect differences in selection pressure or in the initial frequency of Ile1,016. The rapid recent increase in Ile1,016 is predicted by a simple model of positive directional selection on a recessive allele. Unfortunately this model also predicts rapid fixation of Ile1,016 unless there is negative fitness associated with Ile1,016 in the absence of permethrin. If so, then spatial refugia of susceptible Ae. aegypti or rotational schedules of different classes of adulticides could be established to slow or prevent fixation of Ile1,016. Pyrethroid insecticides prolong the opening of voltage-dependent sodium channels in insect nerves to produce instant paralysis and “knock-down.” Many insects have evolved knock-down resistance through nonsynonymous mutations that reduce pyrethroid binding in the channels. In 2006 we discovered one such mutation in the arbovirus mosquito vector Aedes aegypti, called Ile1,016, that confers very high knockdown resistance to the pyrethroid insecticide permethrin in mosquitoes homozygous for this mutation. We examined collections of Ae. aegypti from México during 1996–2009 and found that the overall Ile1,016 frequency increased from <0.1% in 1996–2000, to 2%–5% in 2003–2006, to 38.3%–88.3% in 2007–2009 depending upon collection location. We also demonstrate a strong linear relationship between the frequency of Ile1,016 homozygotes and knockdown rate in bioassays and speculate that widespread use of permethrin-based insecticides in México may be impacting the frequency of Ile1,016. Such a rapid increase is predicted by a simple model of positive directional selection acting on a recessive allele. Unfortunately this model also predicts rapid fixation of Ile1,016 unless there is a negative fitness associated with Ile1,016 in the absence of permethrin and if insecticidal pressure can be reduced.

The mosquito Aedes aegypti is the main vector of Dengue and Yellow Fever flaviviruses. The organophosphate insecticide temephos is a larvicide that is used globally to control Ae. aegypti populations; many of which have in turn evolved resistance. Target site alteration in the acetylcholine esterase of this species has not being identified. Instead, we tracked changes in transcription of metabolic detoxification genes using the Ae. aegypti ‘Detox Chip’ microarray during five generations of temephos selection. We selected for temephos resistance in three replicates in each of six collections, five from Mexico, and one from Peru. The response to selection was tracked in terms of lethal concentrations. Uniform upregulation was seen in the epsilon class glutathione‐S‐transferase (eGST) genes in strains from Mexico prior to laboratory selection, while eGSTs in the Iquitos Peru strain became upregulated after five generations of temephos selection. While expression of many carboxyl/cholinesterase esterase (CCE) genes increased with selection, no single esterase was consistently upregulated and this same pattern was noted in the cytochrome P₄₅₀ monooxygenase (CYP) genes and in other genes involved in reduction or oxidation of xenobiotics. Bioassays using glutathione‐S‐transferase (GST), CCE and CYP inhibitors suggest that various CCEs instead of GSTs are the main metabolic mechanism conferring resistance to temephos. We show that temephos‐selected strains show no cross resistance to permethrin and that genes associated with temephos selection are largely independent of those selected with permethrin in a previous study.

The mosquito Aedes aegypti is the major vector of the four serotypes of dengue virus (DENV1–4). Previous studies have shown that Ae. aegypti in Mexico have a high effective migration rate and that gene flow occurs among populations that are up to 150 km apart. Since 2000, pyrethroids have been widely used for suppression of Ae. aegypti in cities in Mexico. In Yucatan State in particular, pyrethroids have been applied in and around dengue case households creating an opportunity for local selection and evolution of resistance. Herein, we test for evidence of local adaptation by comparing patterns of variation among 27 Ae. aegypti collections at 13 single nucleotide polymorphisms (SNPs): two in the voltage-gated sodium channel gene para known to confer knockdown resistance, three in detoxification genes previously associated with pyrethroid resistance, and eight in putatively neutral loci. The SNPs in para varied greatly in frequency among collections, whereas SNPs at the remaining 11 loci showed little variation supporting previous evidence for extensive local gene flow. Among Ae. aegypti in Yucatan State, Mexico, local adaptation to pyrethroids appears to offset the homogenizing effects of gene flow.

Background: Aedes aegyptiis arguably the most studied of all mosquito species in the laboratory and is the primary vector of both Dengue and Yellow Fever flaviviruses in the field. A large number of transcriptional studies have been made in the species and these usually report transcript quantities observed at a certain age or stage of development. However, circadian oscillation is an important characteristic of gene expression in many animals and plants, modulating both their physiology and behavior. Circadian gene expression in mosquito species has been previously reported but for only a few genes directly involved in the function of the molecular clock.Results: Herein we analyze the transcription profiles of 21,494 messenger RNAs using anAe. aegyptiAgilent^®microarray. Transcripts were quantified in adult female heads at 24 hours and then again at 72 hours and eight subsequent time points spaced four hours apart. We document circadian rhythms in multiple molecular pathways essential for growth, development, immune response, detoxification/pesticide resistance. Circadian rhythms were also noted in ribosomal protein genes used for normalization in reverse transcribed PCR (RT-PCR) to determine transcript abundance. We report pervasive oscillations and intricate synchronization patterns relevant to all known biological pathways.Conclusion: These results argue strongly that transcriptional analyses either need to be made over time periods rather than confining analyses to a single time point or development stage or exceptional care needs to be made to synchronize all mosquitoes to be analyzed and compared among treatment groups.

The majority of the Yucatán State, México, presents subtropical climate that is suitable for many species of mosquitoes that are known to be vectors of diseases, including those from the genera Aedes and Culex. The objective of this study is to identify the geographic distribution of five species from these two genera and estimate the human population at risk of coming in contact with them. We compiled distributional data for Aedes aegypti (L.), Aedes (Howardina) cozumelensis (Diaz Najera), Culex coronator Dyar and Knab, Culex quinquefasciatus Say, and Culex thriambus Dyar from several entomological studies in Yucatán between March 2010 and September 2014. Based on these data, we constructed ecological niche models to predict the spatial distribution of each species using the MaxEnt algorithm. Our models identified areas with suitable environments for Ae. aegypti in most of Yucatán. A similar percentage of urban (97.1%) and rural (96.5%) populations were contained in areas of highest suitability for Ae. aegypti, and no spatial pattern was found (Moran’s I = 0.33, P = 0.38); however, we found an association of abundance of immature forms of this species with annual mean temperature (r = 0.19, P ≤ 0.001) and annual precipitation (r = 0.21, P ≤ 0.001). Aedes cozumelensis is also distributed in most areas of the Yucatán State; Cx. quinquefasciatus, Cx. coronator, and Cx. thriambus are restricted to the northwest. The information generated in this study can inform decision-making to address control measures in priority areas with presence of these vectors.

Chikungunya virus (CHIKV) was isolated from 12 febrile humans in Yucatan, Mexico, in 2015. One patient was co-infected with dengue virus type 1. Two additional CHIKV isolates were obtained from Aedes aegypti mosquitoes collected in the homes of patients. Phylogenetic analysis showed that the CHIKV isolates belong to the Asian lineage.