ABSTRACT Aedes (Stegomyia) aegypti is an important vector of dengue, yellow fever, chikungunya and Zika virus. It is well known that resistance monitoring and genetic diversity data help designing the vector control programs. This study aimed to evaluate resistance to pyrethroids (PYs) through the frequency of kdr mutations Val1016IIe and F1534C, and the genetic variation of the mitochondrial gene ND4 in six natural populations of A. aegypti from Paraná - Brazil. Adults were obtained from eggs collected from Alvorada do Sul, Marilena, Maringá, Nova Londrina, Paranavaí and São Carlos do Ivaí. From these adults, 345 were used to identify the 1016 and 1534 sites, and 120 were used to perform the ND4 gene analysis. The studied populations from Paraná showed PYs resistance, low gene flow and genetic diversity. Additionally, a relationship was observed among the haplotypes of populations from the Amazon and Southeastern Brazil, Peru, Mexico, and North America.

ABSTRACT This contribution endeavored to investigate the genetic structure and gene flow of the flood mosquito, Aedes vexans (Meigen, 1830). Using partial sequences of the mitochondrial COI gene, available from BOLD Systems and GenBank, the Haplotypic (Hd) and nucleotide (π) gene diversity, genetic structuring and gene flow of A. vexans at the global, continental, and country levels were calculated. In total, 1,184 sequences were obtained, distributed among America (88.60%; represented by EUA and Canada), Europe (7.35%), Asia (3.89%), and Africa (0.17%). From these, 395 haplotypes (H) without presence of pseudogenes (NUMTs) were detected. The cluster analyses grouped the haplotypes into six clades. Clade I includes haplotypes from countries in America and Europe, while clades II and III include haplotypes exclusively from Asia and Europe; clade IV grouped only one haplotype from Africa and clade V grouped haplotypes from America and Africa. The global Hd and π were 0.92 and 0.01, respectively. In addition, there is evidence of genetic structuring among continents (7.07%), countries (1.62%), and within countries (91.30%; FST = 0.08, p < 0.05) and no isolation by distance was detected (r = 0.003, p > 0.05). The genetic diversity of A. vexans was found to be greater in North America than in other continents. Although this provisional conclusion might be influenced by a sample bias, since 88.60% of the sequences are from America, is also plausible to consider that America corresponds to the ancestral distribution area of the flood mosquito. This hypothesis needs further testing, using a more comprehensive sample from other continents. Additionally, the six clusters found and their geographical distribution do not support previous proposals of splitting the genus into three subspecies confined to certain geographical areas.

ABSTRACT In Colombia Aedes aegypti is present in 80% of the country up to 2,300 m; however, little is known of its genetic relations within a country context and, hence, within a broader context, for example, America. The aforementioned, herein, analyzed the gene flow within a context of the Americas, its directionality and genetic diversity of the mitochondrial lineages in the A. aegypti populations for Colombia. This called for the use of the sequences for A. aegypti available of the mitochondrial ND4 gene in the GenBank for Colombia and the American continent. No presence was found of nuclear mitochondrial pseudogenes (NUMTs) for Colombia. It is estimated that in Colombia the gene flow of the A. aegypti populations is occurring from the southeast and northeast toward the center of the country. In comparison with the mitochondrial sequences for America, the vector’s haplotypes in Colombia suggest connections between the populations of mosquitoes from the south with those from the north of the continent. The gene flow model at continental scale suggests bidirectional connections between the populations from the north of the continent with those from the south, while at South American scale it proposes the gene flow in all the directions with respect to the Colombian. The Colombian A. aegypti vector monitoring and control strategies must pay special attention to the vector’s points of entry into Colombia related with Peru, Venezuela, Brazil, Mexico, and North America to avoid the entry of populations with characteristics like resistance to insecticides or vector competition.

RESUMEN En Colombia, Aedes aegypti es resistente a la mayoría de insecticidas utilizados. Debido al lento desarrollo de resistencia a Bacillus thuringiensis var. israelensis (Bti), así como su alta especificidad e inocuidad ambiental, el uso de este larvicida, se torna una alternativa en el manejo de este vector. En este trabajo, se evaluó experimentalmente y describió por medio de un modelo matemático, la dinámica del control de poblaciones naturales de A. aegypti, utilizando Bti. Se determinó el perfil de susceptibilidad, a través de bioensayos dosis-respuesta con larvas colectadas de Armenia (Quindío, Colombia). Adicionalmente, con los datos de mortalidad de las localidades analizadas, un nuevo análisis fue realizado, para estimar el perfil de susceptibilidad del municipio de Armenia. Los datos de mortalidad fueron utilizados para calcular las concentraciones letales 50 y 95. Con éstas, se realizaron simulaciones hipotéticas del comportamiento del vector, obtenidas a partir de un modelo matemático, que describe la dinámica poblacional, usando aplicaciones sucesivas de Bti y diferentes intervalos de tiempo. Los bioensayos dosis-respuesta indican que las poblaciones del vector analizadas son susceptibles al Bti, por presentar una respuesta biológica similar a la obtenida en la cepa de referencia Rockefeller. Las simulaciones aplicando un control periódico sostenido, sugieren que el Bti es efectivo para controlar el vector; sin embargo, su eficiencia a largo plazo depende de la relación entre concentración letal y frecuencia de aplicación. Se concluye que la aplicación sostenida de Bti constituye una alternativa viable para el control de poblaciones de A. aegypti, a largo plazo.

ABSTRACT In Colombia, Aedes aegypti is resistant to most used insecticides. Due to the slow development of resistance to Bacillus thuringiensis var. israelensis (Bti) as well as its high specificity and environmental safety, the use of this larvicide becomes an alternative in the management of this vector. In this work, we evaluated experimentally and describe by a mathematical model the dyamics of control of natural populations of A. aegypti using Bti. The susceptibility profile to Bti was determined through dose-response bioassays with larvae collected in Armenia (Quindío, Colombia). In addition, in order to estimate the susceptibility profile, an analysis was carried out using the mortality data obtained from the four localities analyzed. The mortality data were used to estimate the lethal concentrations (LC50 and 95) from each locality. Using these LC, hypothetical simulations of vector behavior were carried out, obtained from a mathematical model that describes the population dynamics, using successive applications of Bti at different time intervals. The dose-response bioassays indicate tha the analyzed vector populations are susceptible to Bti since they present a biological response similar to the one obtained from A. aegypti Rockefeller strain. Computer simulations using sustained periodic inspection indicate Bti is effective for the vector control. Nevertheless, its long-term efficiency depends on relation between the LC and the frequency of application. It is concluded that the sustained application of Bti represents a long-term viable alternative for the control of A. aegypti populations.

Resumen El dengue, chikunguña y Zika, todas transmitidas por Aedes aegypti, son enfermedades que afectan ampliamente la población mundial. La evaluación de extractos vegetales permite el establecimiento de productos eficientes para el control de éste mosquito. Este trabajo evaluó la actividad larvicida en A. aegypti de 23 especies de la familia Asteraceae, y su composición fitoquímica preliminar. El material vegetal utilizado se recolectó en el Departamento del Quindío, Colombia. Con este material se prepararon los extractos vegetales etanólicos utilizados en la caracterización fitoquímica y en los bioensayos. Para cada extracto se realizó un bioensayo dosis-respuesta con larvas provenientes del municipio de Armenia (Quindío, Colombia) siguiendo el protocolo de la OMS. Estos indican que, después de 48 h, los extractos de Jaegeria hirta (694,8% ± 149,9), Austroeupatorium inulaefolium (753,3% ± 198,8) y Heliopsis oppsitifolia (764,4% ± 170,0) requieren menor concentración para matar el 95% de las larvas. Adicionalmente, se construyó un modelo matemático que describe el comportamiento de las poblaciones, con el fin de evaluar diferentes estrategias de control con los extractos; las simulaciones obtenidas a partir de la solución numérica del sistema permiten concluir que la aplicación de extractos de estas plantas constituye una herramienta viable para el control de A. aegypti. Por su parte, la marcha fitoquímica preliminar de las 23 especies muestra la presencia de taninos, quinonas, flavonoides, esteroles, cumarinas y alcaloides. Se concluye que J. hirta, A. inulaefoliu y H. oppsitifolia ameritan ser estudiadas en profundidad, dado su potencial larvicida para el control de A. aegypti.

Abstract Dengue, chikungunya and Zika, all transmitted by Aedes aegypti, are diseases that affect the human world population. The evaluation of raw plant extracts allows the establishment of efficient products for the control of populations of this mosquito. This work evaluated the larvicidal activity of 23 species of the family Asteraceae, and its preliminary phytochemical composition, to control populations of A. aegypti. The vegetable material was collected in the department of Quindío, Colombia. We prepared the plant extracts used in the phytochemical characterization and bioassays with the material collected. For each extract, a dose-response bioassay was performed with larvae from the municipality of Armenia (Quindío, Colombia) following the WHO protocol. These assays indicated that after 48h the extracts of Jaegeria hirta (694.8% ± 149.9), Austroeupatorium inulaefolium (753.3% ± 198.8) and Heliopsis oppsitifolia (764.4% ± 170.0) require less concentration to kill 95% of larvae. Additionally, a mathematical model that describes the behavior of the mosquito populations was constructed, in order to evaluate different control strategies using the extracts; the simulations obtained from the numerical solution of the system allow us to conclude that the application of extracts from these plants constitutes a viable tool for A. aegypti control. On the other hand, the preliminary phytochemical analysis of the 23 plant species shows the presence of tannins, quinones, falvonoides, sterols, coumarins and alkaloids. It is concluded then, that J. hirta, A. inulaefoliu and H. oppsitifolia deserve to be further studied, given their larvicidal potential for the control of A. aegypti.

RESUMEN Introducción: Aedes (Stegomyia) albopictus (Skuse, 1894) es un vector para los virus del dengue y chicunguña en la naturaleza, junto con cerca de 24 arbovirus en condiciones de laboratorio. El conocimiento de la diversidad genética de los insectos vectores es fundamental para el control eficaz y la eliminación de enfermedades transmitidas por estos. Objetivo: Aquí se determinó el escenario actual de la diversidad genética en poblaciones naturales de A. albopictus a través de una revisión sistemática. Metodología: Se pudieron establecer los primeros registros y distribución de las poblaciones de A. albopictus en el mundo, así como su diversidad genética, estructura genética poblacional y marcadores moleculares utilizados para determinar su diversidad genética. Resultados: A. albopictus se distribuye en todo el mundo con poblaciones genéticamente estructuradas y baja diversidad; Sin embargo, el 89,5% de la diversidad genética conocida se basa en el uso de RFLP, aloenzimas, isoenzimas y marcadores moleculares mitocondriales que presentan problemas significativos según la literatura. Una vez obtenidos los resultados, se realizó un análisis crítico y se detectaron deficiencias existentes. Conclusión: El conocimiento actual de la diversidad genética de A. albopictus se basa en marcadores genéticos que presentan problemas significativos reportados en la literatura; Por lo tanto, los programas de control de vectores dirigidos a las poblaciones de A. albopictus pueden verse comprometidos.

ABSTRACT Introduction: Aedes (Stegomyia) albopictus (Skuse, 1894) is a vector for dengue and chikungunya viruses in the field, along with around 24 additional arboviruses under laboratory conditions. Knowledge of the genetic diversity of insect vectors is critical for the effective control and elimination of vector-borne diseases. Objective: We determined the current scenario of the genetic diversity in natural populations of A. albopictus through a systematic review. Methodology: It was possible to establish the first reports and distribution of A. albopictus populations in the world, as well as its genetic diversity, population genetic structure and molecular markers used to determine its genetic diversity. Results: A. albopictus is distributed worldwide with genetically structured populations and low diversity; however, 89.5% of the genetic diversity known is based on the use of RFLP, allozymes, isozymes, and mtDNA molecular markers that exhibit significant problems according to the literature. After the results were obtained, a critical analysis was carried out and existing shortcomings were detected. Conclusion: The current knowledge of genetic diversity of A. albopictus is based on genetic markers that exhibit significant problems reported in the literature; therefore, vector control programs targeting A. albopictus populations, may be compromised.

ABSTRACT After a dengue outbreak, the knowledge on the extent, distribution and mechanisms of insecticide resistance is essential for successful insecticide-based dengue control interventions. Therefore, we evaluated the potential changes to insecticide resistance in natural Aedes aegypti populations to Organophosphates (OP) and Pyrethroids (PY) after chemical vector control interventions. After a Dengue outbreak in 2010, A. aegypti mosquitoes from the urban area of Jacarezinho (Paraná, Brazil) were collected in 2011 and 2012. Insecticide resistance to OP Temephos was assessed in 2011 and 2012 by dose–response bioassays adopting WHO-based protocols. Additionally, in both sampling, PY resistance was also investigated by the Val1016Ile mutation genotyping. In 2011, a random collection of mosquitoes was carried out; while in 2012, the urban area was divided into four regions where mosquitoes were sampled randomly. Bioassays conducted with larvae in 2011 (82 ± 10%; RR95 = 3.6) and 2012 (95 ± 3%; RR95 = 2.5) indicated an incipient altered susceptibility to Temephos. On the other hand, the Val1016IIe mutation analysis in 2011, presented frequencies of the 1016Ilekdr allele equal to 80%. Nevertheless, in 2012, when the urban area of Jacarezinho was analyzed as a single unit, the frequency of the mutant allele was 70%. Additionally, the distribution analysis of the Val1016Ile mutation in 2012 showed the mutant allele frequencies ≥60% in all regions. These outcomes indicated the necessity of developing alternative strategies such as insecticide rotations for delaying the evolution of resistance.

Mosquito control prevails as the most efficient method to protect humans from the dengue virus, despite recent efforts to find a vaccine for this disease. We evaluated insecticide resistance and genetic variability in natural populations of Aedes aegypti (Linnaeus, 1762) from Colombia. This is the first Colombian study examining kdr mutations and population structure. Bioassays with larvae of three mosquito populations (Armenia, Calarcá and Montenegro) were performed according to the World Health Organization (WHO) guidelines, using Temephos. For the analysis of the Val1016Ile mutation and genetic diversity, we sampled recently-emerged adults from four mosquito populations (Armenia, Calarcá, Montenegro and Barcelona). Following the WHO protocol, bioassays implemented with larvae showed resistance to Temephos in mosquito populations from Armenia (77% ± 2) and Calarcá (62% ± 14), and an incipient altered susceptibility at Montenegro (88% ± 8). The RR95 of mosquito populations ranged from 3.7 (Montenegro) to 6.0 (Calarca). The Val1016Ile mutation analysis of 107 genotyped samples indicates that 94% of the specimens were homozygous for the wild allele (1016Val) and 6% were heterozygous (Val1016Ile). The 1016Ile allele was not found in Barcelona. Genetic variability analysis found three mitochondrial lineages with low genetic diversity and gene flow. In comparison with haplotypes from the American continent, those from this study suggest connections with Mexican and North American populations. These results confirm that a continuous monitoring and managing program of A. aegypti resistance in the state of Quindío is required.

Objective Evaluating the larvicidal activity of two ether factions from Asteraceae (the aster, daisy or sunflower family, i.e. Heli opsisoppositifolia (L.) Druce (oxeye, sunflower-like) and Jaegeria hirta (Lag.) Less (weed-like)) on Aedes (Stegomyia) aegypti (L.) final third instar or initial fourth instar larvae near the town of Armenia in the Quindío Department in Colombia. Methods H. oppositifolia and J. hirta plants were collected and submitted to phytochemical analysis. Ether fractions were prepared form both species to assess 11 concentrations for determining LC50, LC90 and LC95 lethal concentrations after 24 and 48h. The LC50, LC90 and LC95 results were used to create a mathematical model for describing lethal population-concentration dynamics. Results Phytochemical analysis identified tannins, flavonoids, quinones, cardiac glycosides, sterols, lactones, terpenes, courmarins and alkaloids in H. oppositifolia and J. hirta. LC after 48h regarding J. hirta (LC50 24, LC90 70 and LC95 93ppm) was lower than those for H. oppositifolia (LC50 39, LC90 77 and LC95 94 ppm). A factorial ANOVA test confirmed this trend: 66 %, F=18.5 and p<0.05 for J. hirta and 34 %, F=18.5 and p<0.05 for H. oppositifolia. The mathematical simulation model suggested that using LC50 every 15 days and LC90 and LC95 every 30 days from either of these species led to the same response compared to using LC90 and LC95 every 30 days or LC50 every 15 days. Conclusion Both species had a larvicidal effect. However, J. Hirta turned out to be more promising as an eventual bioinsecticide for controlling A. Aegypti immature states.

Objetivo Evaluar la actividad larvicida de dos fracciones etéreas de Heli opsisoppositifolia (L.) Drucey Jaegeria hirta (Lag.) Less en larvas de Aedes (Stegomyia) aegypti (L.) de 3-4 ínstar, en el municipio de Armenia, Quindio. Métodos Se realizó la colecta y análisis fitoquímico preliminar de dos especies vegetales, H. oppositifolia y J. hirta. Fueron preparadas fracciones etéreas de las dos especies. Con estas fracciones, fueron evaluadas 11 concentraciones para determinar las concentraciones letales CL50, CL90 y CL95 después de 24 y 48h. Con los resultados de las CL50,CL90 y CL95, se simuló y construyó un modelo que describe la dinámica población-concentración letal. Resultados La marcha fitoquímica preliminar permitió caracterizar en H. oppositifolia y J. hirta la presencia de: Taninos, flavonoides, quinonas, glicósidos cardiotónicos, esteroles, lactonas, terpenos, coumarinas y alcaloides. Las CL, 48 después para J. hirta (CL5024 CL9070 y CL9593 ppm) fueron menores que H. oppositifolia (CL5039, CL9077y CL9594 ppm). El ANOVA factorial confirmó esta tendencia, J. hirta (66 %, F=18.5, p<0.05) y H. oppositifolia (34 %, F=18.5, p<0.05). La simulación matemática sugiere que la aplicación cada 15 días de la CL50, y cada 30 días de las CL90 y CL95 de cualquiera de las dos especies, tienen la misma respuesta que la utilización de las CL90 y CL95 cada 30 días o la CL50 cada 15 días. Conclusión Ambas especies poseen efecto larvicida. Sin embargo, J. hirta se mostró más promisoria como futuro bioinsecticida para el control de estados inmaduros de Ae. aegypti.

Objetivo Evaluar la actividad larvicida de dos fracciones etéreas de Heli opsisoppositifolia (L.) Drucey Jaegeria hirta (Lag.) Less en larvas de Aedes (Stegomyia) aegypti (L.) de 3-4 ínstar, en el municipio de Armenia, Quindio. Métodos Se realizó la colecta y análisis fitoquímico preliminar de dos especies vegetales, H. oppositifolia y J. hirta. Fueron preparadas fracciones etéreas de las dos especies. Con estas fracciones, fueron evaluadas 11 concentraciones para determinar las concentraciones letales CL50, CL90 y CL95 después de 24 y 48h. Con los resultados de las CL50,CL90 y CL95, se simuló y construyó un modelo que describe la dinámica población-concentración letal. Resultados La marcha fitoquímica preliminar permitió caracterizar en H. oppositifolia y J. hirta la presencia de: Taninos, flavonoides, quinonas, glicósidos cardiotónicos, esteroles, lactonas, terpenos, coumarinas y alcaloides. Las CL, 48 después para J. hirta (CL5024 CL9070 y CL9593 ppm) fueron menores que H. oppositifolia (CL5039, CL9077y CL9594 ppm). El ANOVA factorial confirmó esta tendencia, J. hirta (66 %, F=18.5, p<0.05) y H. oppositifolia (34 %, F=18.5, p<0.05). La simulación matemática sugiere que la aplicación cada 15 días de la CL50, y cada 30 días de las CL90 y CL95 de cualquiera de las dos especies, tienen la misma respuesta que la utilización de las CL90 y CL95 cada 30 días o la CL50 cada 15 días. Conclusión Ambas especies poseen efecto larvicida. Sin embargo, J. hirta se mostró más promisoria como futuro bioinsecticida para el control de estados inmaduros de Ae. aegypti.

Objective Evaluating the larvicidal activity of two ether factions from Asteraceae (the aster, daisy or sunflower family, i.e. Heli opsisoppositifolia (L.) Druce (oxeye, sunflower-like) and Jaegeria hirta (Lag.) Less (weed-like)) on Aedes (Stegomyia) aegypti (L.) final third instar or initial fourth instar larvae near the town of Armenia in the Quindío Department in Colombia. Methods H. oppositifolia and J. hirta plants were collected and submitted to phytochemical analysis. Ether fractions were prepared form both species to assess 11 concentrations for determining LC50, LC90 and LC95 lethal concentrations after 24 and 48h. The LC50, LC90 and LC95 results were used to create a mathematical model for describing lethal population-concentration dynamics. Results Phytochemical analysis identified tannins, flavonoids, quinones, cardiac glycosides, sterols, lactones, terpenes, courmarins and alkaloids in H. oppositifolia and J. hirta. LC after 48h regarding J. hirta (LC50 24, LC90 70 and LC95 93ppm) was lower than those for H. oppositifolia (LC50 39, LC90 77 and LC95 94 ppm). A factorial ANOVA test confirmed this trend: 66 %, F=18.5 and p<0.05 for J. hirta and 34 %, F=18.5 and p<0.05 for H. oppositifolia. The mathematical simulation model suggested that using LC50 every 15 days and LC90 and LC95 every 30 days from either of these species led to the same response compared to using LC90 and LC95 every 30 days or LC50 every 15 days. Conclusion Both species had a larvicidal effect. However, J. Hirta turned out to be more promising as an eventual bioinsecticide for controlling A. Aegypti immature states.