Background: The chemical control of the mosquitoAedes aegypti, the major vector of dengue, is being seriously threatened due to the development of pyrethroid resistance. Substitutions in the 1016 and 1534 sites of the voltage gated sodium channel (AaNa_V), commonly known askdrmutations, confer the mosquito with knockdown resistance. Our aim was to evaluate the allelic composition of natural populations of BrazilianAe. aegyptiat bothkdrsites.Methods: The AaNa_VIIIS6 region was cloned and sequenced from three Brazilian populations. Additionally, individual mosquitoes from 30 populations throughout the country were genotyped for 1016 and 1534 sites, based in allele-specific PCR. For individual genotypes both sites were considered as a single locus.Results: The 350 bp sequence spanning the IIIS6 region of theAaNa_Vgene revealed the occurrence of thekdrmutation Phe1534Cys in Brazil. Concerning the individual genotyping, beyond the susceptible wild-type (Na_V^S), twokdralleles were identified: substitutions restricted to the 1534 position (Na_V^R1) or simultaneous substitutions in both 1016 and 1534 sites (Na_V^R2). A clear regional distribution pattern of these alleles was observed. The Na_V^R1kdrallele occurred in all localities, while Na_V^R2was more frequent in the Central and Southeastern localities. Locations that were sampled multiple times in the course of a decade revealed an increase in frequency of thekdrmutations, mainly the double mutant allele Na_V^R2. Recent samples also indicate that Na_V^R2is spreading towards the Northern region.Conclusions: We have found that in addition to the previously reported Val1016Ilekdrmutation, the Phe1534Cys mutation also occurs in Brazil. Allelic composition at both sites was important to elucidate the actual distribution ofkdrmutations throughout the country. Studies to determine gene flow and the fitness costs of thesekdralleles are underway and will be important to better understand the dynamics ofAe. aegyptipyrethroid resistance.

Aedes aegypti dispersion is the major reason for the increase in dengue transmission in South America. In Brazil, control of this mosquito strongly relies on the use of pyrethroids and organophosphates against adults and larvae, respectively. In consequence, many Ae. aegypti field populations are resistant to these compounds. Resistance has a significant adaptive value in the presence of insecticide treatment. However some selected mechanisms can influence important biological processes, leading to a high fitness cost in the absence of insecticide pressure. We investigated the dynamics of insecticide resistance and its potential fitness cost in five field populations and in a lineage selected for deltamethrin resistance in the laboratory, for nine generations. For all populations the life-trait parameters investigated were larval development, sex ratio, adult longevity, relative amount of ingested blood, rate of ovipositing females, size of egglaying and eggs viability. In the five natural populations, the effects on the life-trait parameters were discrete but directly proportional to resistance level. In addition, several viability parameters were strongly affected in the laboratory selected population compared to its unselected control. Our results suggest that mechanisms selected for organophosphate and pyrethroid resistance caused the accumulation of alleles with negative effects on different life-traits and corroborate the hypothesis that insecticide resistance is associated with a high fitness cost.

Distribution, abundance, feeding behaviour, host preference, parity status and human-biting and infection rates are among the medical entomological parameters evaluated when determining the vector capacity of mosquito species. To evaluate these parameters, mosquitoes must be collected using an appropriate method. Malaria is primarily transmitted by anthropophilic and synanthropic anophelines. Thus, collection methods must result in the identification of the anthropophilic species and efficiently evaluate the parameters involved in malaria transmission dynamics. Consequently, human landing catches would be the most appropriate method if not for their inherent risk. The choice of alternative anopheline collection methods, such as traps, must consider their effectiveness in reproducing the efficiency of human attraction. Collection methods lure mosquitoes by using a mixture of olfactory, visual and thermal cues. Here, we reviewed, classified and compared the efficiency of anopheline collection methods, with an emphasis on Neotropical anthropophilic species, especially Anopheles darlingi, in distinct malaria epidemiological conditions in Brazil.

Anopheles darlingi is the major malaria vector in countries located in the Amazon region. Anopheles aquasalis and Anopheles albitarsis s.l. are also proven vectors in this region. Anopheles nuneztovari s.l. and Anopheles triannulatus s.l. were found infected with Plasmodium vivax; however, their status as vectors is not yet well defined. Knowledge of susceptibility of Amazon anopheline populations to Plasmodium infection is necessary to better understand their vector capacity. Laboratory colonization of An. darlingi, the main Amazon vector, has proven to be difficult and presently An. aquasalis is the only available autonomous colony.

Culex quinquefasciatusis a successful invasive species broadly distributed in subtropical regions, including Brazil. It is an extremely annoying mosquito due to its nocturnal biting behavior, in high-density populations and it is a potential bridge between sylvatic arbovirus from birds to man in urban territories. Herein, we present a review concerning the methods of chemical control employed againstCx. quinquefasciatusin Brazil since the 1950’s and insecticide resistance data registered in the literature. As there is no specific national programme forCx. quinquefasciatuscontrol in Brazil, the selection of insecticide resistance is likely due in part to the well-designed chemical campaigns againstAedes aegyptiand the elevated employment of insecticides by households and private companies. There are very few publications about insecticide resistance inCx. quinquefasciatusfrom Brazil when compared toAe. aegypti. Nevertheless, resistance to organophosphates, carbamate, DDT, pyrethroids and biolarvicides has been registered inCx. quinquefasciatuspopulations from distinct localities of the country. Concerning physiological mechanisms selected for resistance, distinct patterns of esterases, as well as mutations in the acetylcholinesterase (ace-1) and voltage-gated sodium channel (Na_V) genes, have been identified in natural populations. Given environmental changes and socioeconomical issues in the cities, in recent years we have been experiencing an increase in the number of disease cases caused by arboviruses, which may involveCx. quinquefasciatusparticipation as a key vector. It is urgent to better understand the efficiency and susceptibility status to insecticides, as well as the genetic background of known resistant mechanisms already present inCx. quinquefasciatuspopulations for an effective and rapid chemical control when eventually required.

Arboviruses are viruses transmitted to humans and other animals by the bite of hematophagous arthropods. Infections caused by chikungunya virus (CHIKV), dengue virus (DENV), Zika virus (ZIKV), and the deadlier yellow fever virus (YFV) are current public health problems in several countries, mainly those located in tropical and subtropical regions. One of the main prevention strategies continues to be vector control, with the elimination of breeding sites and surveillance of infested areas. The use of ovitraps for Aedes mosquitos monitoring has already demonstrated promising results, and maybe be also useful for arboviral surveillance. This work aimed to detect natural vertical transmission of arboviruses in Aedes aegypti and Aedes albopictus. Mosquito egg collection was carried out using ovitraps in Itacoatiara, a mid-size city in Amazonas state, Brazil. Collected eggs were allowed to hatch and larvae were tested for CHIKV, DENV, and ZIKV RNA by RT-qPCR. A total of 2,057 specimens (1,793 Ae. aegypti and 264 Ae. albopictus), in 154 larvae pools were processed. Results showed one positive pool for CHIKV and one positive pool for ZIKV. The active ZIKV infection was further confirmed by the detection of the negative-strand viral RNA and nucleotide sequencing which confirmed the Asian genotype. The Infection Rate per 1,000 mosquitoes tested was assessed by Maximum Likelihood Estimation (MLE) with 0.45 and 0.44 for CHIKV and ZIKV, respectively, and by Minimum Infection Rate (MIR) with 0.45 for both viruses. To our knowledge, this is the first detection of ZIKV in natural vertical transmission in the Ae. aegypti, a fact that may contribute to ZIKV maintenance in nature during epidemics periods. Furthermore, our results highlight that the use of ovitraps and the molecular detection of arbovirus may contribute to health surveillance, directing the efforts to more efficient transmission blockade. The control of the vast majority of arbovirus infections relies on entomological measures to reduce mosquito infestation. Therefore, this study analyzed the use of ovitraps for arboviral surveillance in a mid-size city of the Amazonas state, Brazil. We found one larva pool infected with chikungunya virus, before the first human case confirmed in this municipality. Another pool was infected with Zika virus, demonstrating the first evidence that vertical transmission occurs in naturally infected Aedes aegypti mosquito populations.