Orientador : Prof. Dr. Mário Antônio Navarro da Silva / Coorientador : Dr. Ademir de Jesus Martins Junior / Tese (doutorado) - Universidade Federal do Paraná, Setor de Ciências Biológicas, Programa de Pós-Graduação em Ciências Biológicas (Entomologia). Defesa: Curitiba, 17/11/2016 / Inclui referências: f. 81-98 / Área de concentração : Entomologia / Resumo: O canal de sódio voltagem-dependente (NaV) é fundamental para a sinalização elétrica no sistema nervoso e é o sítio-alvo para Piretróides (PY) e DDT. A resistência knockdown (kdr), causada por alterações no NaV, é portanto, um dos mecanismos que confere resistência a estes inseticidas. Pelo menos 12 mutações não sinônimas foram relatadas em nove códons diferentes no gene do NaV em Aedes aegypti, a maioria delas nos segmentos IIS6 e IIIS6. A mutação kdr Phe1534Cys no segmento IIIS6 é a mais prevalente em todo o mundo, e também foi encontrada em populações asiáticas de Aedes albopictus resistente aos PY. Aqui foi investigada a ocorrência, frequência e distribuição de mutações kdr presentes nos segmentos IIS6 e IIIS6 do NaV em populações naturais de A. aegypti da Colômbia e Brasil e de A. albopictus do Brasil. O DNA de 13 populações de A. aegypti da Colômbia (quatro) e Brasil (nove) e nove de A.albopictus do Brasil foram individualmente extraídos e agrupados por Estado (A.aegypti) e localidade (A. albopictus), amplificados, clonados e sequenciados para os correspondentes segmentos IIS6 e IIIS6 do NaV. Adicionalmente, os indivíduos de cada espécie, para cada localidade foram individualmente genotipados via PCR alelo específica (AS-PCR) para se obter as frequências genotípicas e alélicas para os sítios kdr 1016 e 1534 para A. aegypti, e 1534 para A. albopictus. A área urbana de Foz do Iguaçu foi dividida por estratos, de forma que as frequências genotípicas e alélicas foram então analisadas separadamente e, em seguida, como uma unidade. Nas populações de A. aegypti de ambos os países foram detectados três polimorfismos não sinônimos no NaV (Ile1011Met, Val1016Ile no segmento IIS6 e Phe1534Cys no IIIS6). Já em A. albopictus apenas a mutação Phe1534Cys foi evidenciada em populações brasileiras dos Estados de Paraná (PR) e Rondônia (RO). Substituições restritas ao sítio 1534 (NaV R1) e substituições simultâneas em ambos os sítios 1016 e 1534 (NaV R2) foram observadas em todas as populações de A. aegypti da Colômbia e Brasil. O alelo kdr NaV R1 foi detectado em todas as localidades da Colômbia e Brasil. No Brasil, o alelo kdr NaV R2 foi mais frequente nas localidades da região Sul e Centro-Oeste. Em Foz do Iguaçu, o alelo kdr NaV R2 estava distribuido nos estratos em frequências ?55%, com exceção do estrato 1. Por sua parte, em A. albopictus, o alelo 1534Cyskdr variou entre 3% (Marilena/PR e Porto Velho/RO) e 10% (Foz do Iguaçu/PR). Esta é a primeira evidencia de ocorrência e distribuição de uma mutação kdr em A. albopictus para América do Sul. Os resultados aqui apresentados sugerem a necessidade de um uso integrado apropriado de inseticidas na Colômbia e no Brasil, assim como a inclusão de A. albopictus no monitoramento da resistência aos inseticidas, a fim de manter os compostos químicos como uma ferramenta de controle de vetores quando necessário. Palavras-chave: chikungunya, dengue, piretróides, controle vetorial, canal de sódio voltagem-dependente, Zika. / Abstract: The voltage-gated sodium channel (NaV) is critical for electrical signaling in the nervous system and is the target site for Pyrethroids (PY) and DDT. The knockdown resistance (kdr) caused by changes in NaV, is one of the mechanisms that confers resistance to those insecticides. At least 12 non-synonymous mutations were reported at nine different codons in the NaV gene in Aedes aegypti, most of them in the segments IIS6 and IIIS6. The kdr mutation Phe1534Cys in IIIS6 NaV segment is the most prevalent throughout the world, and has also been found in Asian Aedes albopictus populations resistant to PY. Here, we studied the occurrence, frequency and distribution of kdr mutations present in the IIS6 and IIIS6 NaV segments in natural populations of A.aegypti from Colombia and Brazil and A. albopictus from Brazil. The DNA from 13 A. aegypti populations from Colombia (four) and Brazil (nine) and nine from A. albopictus from Brazil were individually extracted and grouped by State (A. aegypti) and location (A. albopictus), amplified, cloned and sequenced to the corresponding IIS6 and IIIS6 NaV segment. In addition, individuals from each location for each species were individually genotyped using PCR specific allele (AS-PCR) for obtaining the genotypic and allelic frequencies for the kdr sites 1016 and 1534 for A. aegypti and 1534 for A.albopictus. The urban area of Foz do Iguaçu was divided by strata, and genotypic and allelic frequencies were analyzed, first by strata and then as a single unit. In A. aegyptipopulations from both countries three non-synonymous polymorphisms (Ile1011Met, Val1016Ile in IIS6 and Phe1534Cys in IIIS6 NaV segment) in the NaV gene were detected. However, in A. albopictus, only the Phe1534Cys kdr mutation was evidenced in Brazilian populations' from the states of Paraná (PR) and Rondônia (RO). Restricted substitutions to 1534 site (NaV R1) and simultaneous substitutions at both 1016 and 1534 sites in (NaV R2) were observed in all A. aegypti populations from Colombia and Brazil. The NaV R1 kdr allele was detected in all locations from Colombia and Brazil. In Brazil, the NaV R2 kdr allele was more frequent in the localities from the South and Central West. In Foz do Iguaçu, the NaV R2 kdr allele was spread in all strata in frequencies ?55%, except for the stratum 1. On the other hand, in A. albopictus, the allele 1534Cyskdr ranged from 3% (Marilena/PR and Porto Velho/RO) 10% (Foz do Iguaçu/PR). This is the first evidence of occurrence and distribution of kdr mutation in A. albopictus from South America. The results presented here suggest the need for an appropriate integrated use of insecticides in Colombia and Brazil, as well as the inclusion of A. albopictus monitoring to insecticides resistance in order to maintain the chemical compounds as an efficient vector control tool when necessary. Keywords: chikungunya, dengue, pyrethroid resistance, vector control, voltage-gated sodium channel, Zika.

Made available in DSpace on 2014-06-12T18:06:20Z (GMT). No. of bitstreams: 2 arquivo6297_1.pdf: 1280422 bytes, checksum: 6dbf4fac7abd5986311944cbb4ed97fd (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2005 / Aedes aegypti e Aedes albopictus são espécies invasivas, considerada importante do ponto de vista médico, porque eles são vetores de arboviroses muitos. Uma pesquisa de doze meses de variação sazonal na freqüência de alelos em populações destas espécies foi realizado para verificar as possíveis alterações na estrutura genética e sua relação com as condições ambientais. Padrões de diversidade genética foram examinadas usando Random Amplified Polymorphic DNA e haplótipos de Citochrome c oxidase subunidade II do mtDNA. Amostras de Ae. aegypti e Ae. albopictus foram coletados entre dezembro de 2003 a novembro de 2004 em duas áreas do Recife. Evidência de diferenciação genética significativa foi detectada ao longo do ano para ambas as espécies, sem perdas de diversidade genética dentro das populações (heterozigosidade e polimorfismo). Amostras de Ae. albopictus em áreas silvestres e urbanas mostrou fluxo gênico intensiva (Nm = 4.2 e 22), durante dois momentos distintos. Os dados obtidos podem ser muito úteis para melhorar as estratégias de controle de vetores

Mycobacterium ulcerans is an acid-fast bacillus that is the causative agent of Buruli ulcer, a necrotizing skin disease. The transmission route for M. ulcerans is unknown, but many insects have been posited as part of the web, including Belostomatids, Naucorids, and Culicids. Aedes albopictus was selected for use in a set of experiments where the first-generation larvae were inoculated with M. ulcerans, and mosquitoes were reared throughout the third generation to interrogate presence and quantity of the bacteria. Using qPCR, second and third generations displayed positivity (22% and 5.6% respectively). 16S V4 sequencing was used to obtain microbiota for all life stages as well as environmental samples, and many relationships between generations, life stages, and treatments displayed statistical significance in alpha diversity, beta diversity, and relative abundance of microbiomes. This study opens multiple avenues of further investigation into the transmission web of Buruli ulcer.

Phytophagy (i.e., feeding on plant-derived materials) is an essential component of mosquito biology. Yet, it has been historically neglected as most research effort has been concentrated on host-seeking behavior and pathogen transmission. As mosquitoes are the deadliest animals on earth and because challenges, such as the rise of insecticide resistance, arise, there is an urgent need for developing effective and ecologically friendly disease vector control strategies. It is therefore important to deepen our understanding of mosquito phytophagy and, consequently, its potential to develop novel vector control methods. Particular major disease vectors are Ae. aegypti and Ae. albopictus, which are spreading rapidly through the US, in part due to climate change. Herein, we first examine the effect of temperature on Ae. aegypti sugar-feeding behavior as well as overall locomotive activity and survival, using total carbohydrate assays and actometer experiments. An optimum temperature range for mosquito activity is proposed and discussed in the context of global warming. We then observe the tentative benefit provided by city-planted ornamental flowers to Ae. aegypti and Ae. albopictus living in heavily-populated, urban areas. Mosquito sugar-feeding activity and, subsequently, sugar consumption were tested for eleven commonly-planted ornamentals. Additionally, scents were collected from the headspace of each ornamental, and volatile composition was analyzed and discussed as potential cues that could mediate mosquito-plant interactions. / Master of Science in Life Sciences / Sugar-feeding is an important, but generally neglected, aspect of mosquito biology, affecting, for example, their survival, metabolism, and ability to lay eggs. While females need blood to mature their eggs, males feed exclusively on sugar, further highlighting the importance of this food source. Mosquitoes use several cues to locate flowers to feed on, including the plant scent. Aedes aegypti and Aedes albopictus are two urban species that are invasive to the US and are the vectors of several deadly pathogens including dengue, Zika and chikungunya. It is thus to study any aspect of their biology that could lead to the development of new tools to limit their propagation. Because of the nutrients provided to mosquitoes by flowering plant species, considering the dynamic ecological relationship between human, plant, and mosquito in urban, heavily populated areas is critical. Additionally, how temperature is mediating each of these interactions is important to understand and keep in consideration. Here, we first examine the effect of temperature on sugar-feeding, activity and survival in Ae. aegypti, and discuss our results in the context of potential changes in temperature caused by climate change. We also provide some insights on the role that ornamental flower species play in urban areas in the ability of these mosquito species to thrive, by attracting mosquitoes to areas where they will have access to a high number of human blood meals.

Recently, the number of human cases of La Crosse encephalitis (LACE), an illness caused by mosquito-borne La Crosse (LAC) virus, has increased in southwestern Virginia, resulting in a need for better understanding of the virus cycle and the biology of its vectors in the region. This project examined the spatial and temporal distributions of the primary vector of LAC virus, Ochlerotatus triseriatus, and a potential secondary vector, Aedes albopictus. Ovitrapping surveys were conducted in 1998 and 1999 to determine distributions and oviposition habitat preferences of the two species in southwestern Virginia. For virus assay, adult mosquitoes were collected at a tire dump and a human case site during 1998 and 1999, and ovitrap samples were taken from a human case site in 2000. In a separate study, a landcover map of Wise County was created by supervised classification of Landsat Enhanced Thematic Mapper imagery, and maps indicating posterior probabilities of high mosquito abundance were created by combining ovitrap survey-derived, landcover-based prior and conditional probabilities for high and low mosquito abundance using remote sensing techniques and Bayesian decision-making rules. Both Oc. triseriatus and Ae. albopictus were collected from all ovitrap sites surveyed in Wise, Scott, and Lee Counties during 1998. Numbers of Oc. triseriatus remained high from late June through late August, while Ae. albopictus numbers increased gradually through June and July, reaching a peak in late August and declining thereafter. Overall, Oc. triseriatus accounted for 90.1% of eggs collected during this period, and Ae. albopictus made up the remaining 9.9%. Abundance of the two species differed among the sites, and in Wise County, relative Ae. albopictus abundance was highest in sites with traps placed in open residential areas. Lowest numbers of both species were found in densely forested areas. Ovitrapping at a human LACE case site during 1998 and 1999 revealed that Aedes albopictus was well-established and overwintering in the area. An oviposition comparison between yard and adjacent forest at the Duncan Gap human LACE case site in 1999 showed that Ae. albopictus preferentially oviposited in the yard surrounding the home over adjacent forested areas, but Oc. triseriatus showed no preference. LAC virus was isolated from 1 larval and 1 adult collection of Oc. triseriatus females from the Duncan Gap human case site, indicating the occurrence of transovarial transmission at this site. The supervised landcover classification for Wise County yielded a landcover map with an overall accuracy of 98% based on comparison of output classification with user-defined ground truth data. Posterior probability maps for Oc. triseriatus and Ae. albopictus abundance reflected seasonal and spatial fluctuations in mosquito abundance with an accuracy of 55-79% for Oc. triseriatus (Kappa=0.00-0.53) and 70-94% for Ae. albopictus (Kappa=0.00-0.49) when model output was compared with results of an ovitrapping survey. Other accuracy measures were also considered, and suggestions were offered for improvement of the model. / Master of Science

Doenças como a dengue, chikungunya, Zika e febre amarela urbana são causadas por patógenos transmitidos pela picada de fêmeas infectadas de Ae. aegypti e Ae. albopictus. Fatores bióticos, como a competição na fase larval, e abióticos, como a variação da temperatura, provocam alterações no ciclo de desenvolvimento dessas espécies. O presente estudo avaliou os efeitos da competição intraespecífica e da temperatura na atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus. Os objetivos do trabalho foram: comparar o tamanho das fêmeas de Ae. aegypti e Ae. albopictus oriundas de criadouros com baixa e alta competição intraespecífica; avaliar o efeito da competição intraespecífica na atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus, sob regime de 12 horas de claro e 12 horas de escuro, à 25ºC e analisar o padrão da atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus, sob regime de 12 horas de claro e 12 horas de escuro, sob diferentes temperaturas (20ºC, 25ºC e 30ºC). As colônias dos mosquitos utilizados nos experimentos são provenientes da FIOCRUZ-RJ. Para o experimento de competição, larvas de ambas as espécies foram criadas sob condições de alta (100 larvas) e baixa (20 larvas) densidades. Para isso, foram utilizadas 14 bacias preenchidas com 500 ml de água e 0,2 g de ração para peixes. Após a competição, as fêmeas oriundas de cada densidade foram transferidas para incubadoras para a realização da análise da atividade locomotora. Após a atividade, verificou-se o tamanho dos adultos oriundos de cada densidade larvária, por meio da medição das asas de cada fêmea. Para a análise da atividade locomotora sob diferentes temperaturas, as fêmeas utilizadas foram desenvolvidas em ambiente com quantidade de ovos similares. Foi utilizado o mesmo procedimento para análise da atividade do experimento anterior, porém, cada experimento ocorreu sob diferentes temperaturas: 20°C, 25°C e 30°C. Para a análise dos dados obtidos foram utilizados os testes T e ANOVA multifatorial. Para o experimento de competição, fêmeas de ambas as espécies oriundas de competição de baixa densidade foram maiores do que as oriundas de desenvolvimento sob alta densidade larvária. A atividade das fêmeas de Ae. aegypti e Ae. albopictus de ambas as densidades foi muito semelhante, sugerindo que fêmeas menores tem atividade equivalente à de fêmeas maiores. Já no experimento que avaliou a atividade das fêmeas sob faixas térmicas distintas, notou-se que, em Ae. aegypti, houve diferença significativa na atividade total sob 20°C quando comparadas a 25°C e 30°C, demonstrando que a temperatura mais baixa foi limitante para a atividade desta espécie. Houve diferenças em algumas médias avaliadas na atividade de Ae. albopictus, porém, esta espécie pareceu ter sido menos influenciada com a variação da temperatura. Análises estatísticas confirmaram que as fêmeas de Ae. aegypti respondem de forma distinta das fêmeas de Ae. albopictus quando as temperaturas foram modificadas. O presente estudo ajuda a esclarecer alguns aspectos relacionados ao comportamento dos mosquitos Ae. aegypti e Ae. albopictus, que são considerados de grande importância para a saúde pública. / Diseases such as dengue, chikungunya, Zika and urban yellow fever are caused by pathogens transmitted by the bites of infected females of Ae. aegypti and Ae. albopictus. Biotics factors essentially competition on larval stages, and abiotics factors, for instance, temperature variation, occasion adjustments on development cycle of these species. The present study evaluated the effects of intraspecific competition and temperature on locomotor activity of Ae. aegypti and Ae. albopictus. The aims of this study were: to compare the size of females Ae. aegypti and Ae. albopictus from breeding sites with low and high intraspecific competition; to evaluate the effect of intraspecific competition on locomotor activity of virgin females of Ae. aegypti and Ae. albopictus, under regime of 12 hours of light and 12 hours of dark at 25°C and to analyze the pattern of the locomotor activity of virgins females of Ae. aegypti and Ae. albopictus, under regime of 12 hours of light and 12 hours of dark, under different temperatures (20ºC, 25ºC e 30ºC). The mosquitoes colonies used on experiment are from FIOCRUZ-RJ. For the competition experiment, larvae from both species were raised under conditions of high (100 larvae) and low (20 larvae) densities. For this 14 containers filled with 500 ml of water and 0,2 g of fish food were used. After the competition, the females from each density were transferred to incubators to perform locomotor activity analysis. After the activity, the size of adult from each larval density, were verified by wings measurement of each female. For the analysis of the locomotor activity under different temperatures, the females used were developed in an environment with similar amount of eggs. The same procedure was used to the analyze the activity of the previous experiment, however, each experiment happened under different temperatures: 20°C, 25°C e 30°C. For the analysis of the obtained data were applied T tests and ANOVA multivariate. For the competition experiment, females of both species from low density of competition were longer than females from competition under high larval density. The activity of females of Ae. aegypti and Ae. albopictus from both densities was very similar, suggesting that smaller females have equivalent activity to that of larger females. In the experiment that evaluated the activity of females under different thermal bands, it was noticed that in Ae. aegypti there was significant difference in total activity under 20°C when compared with 25°C and 30°C degrees, demonstrating the lower temperature was limiting to activity for this species. There were differences in some averages evaluated in Ae. albopictus activity, however, this species seemed to have been less influenced with the temperature variation. Statistical analysis confirmed that females of Ae. aegypti respond differently than females of Ae. albopictus when the temperatures were modified. The present study helps to clarify some aspects about the behavior of mosquitoes Ae. aegypti and Ae. albopictus which are considered of great concern for public health.

Doenças como a dengue, chikungunya, Zika e febre amarela urbana são causadas por patógenos transmitidos pela picada de fêmeas infectadas de Ae. aegypti e Ae. albopictus. Fatores bióticos, como a competição na fase larval, e abióticos, como a variação da temperatura, provocam alterações no ciclo de desenvolvimento dessas espécies. O presente estudo avaliou os efeitos da competição intraespecífica e da temperatura na atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus. Os objetivos do trabalho foram: comparar o tamanho das fêmeas de Ae. aegypti e Ae. albopictus oriundas de criadouros com baixa e alta competição intraespecífica; avaliar o efeito da competição intraespecífica na atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus, sob regime de 12 horas de claro e 12 horas de escuro, à 25ºC e analisar o padrão da atividade locomotora de fêmeas virgens de Ae. aegypti e Ae. albopictus, sob regime de 12 horas de claro e 12 horas de escuro, sob diferentes temperaturas (20ºC, 25ºC e 30ºC). As colônias dos mosquitos utilizados nos experimentos são provenientes da FIOCRUZ-RJ. Para o experimento de competição, larvas de ambas as espécies foram criadas sob condições de alta (100 larvas) e baixa (20 larvas) densidades. Para isso, foram utilizadas 14 bacias preenchidas com 500 ml de água e 0,2 g de ração para peixes. Após a competição, as fêmeas oriundas de cada densidade foram transferidas para incubadoras para a realização da análise da atividade locomotora. Após a atividade, verificou-se o tamanho dos adultos oriundos de cada densidade larvária, por meio da medição das asas de cada fêmea. Para a análise da atividade locomotora sob diferentes temperaturas, as fêmeas utilizadas foram desenvolvidas em ambiente com quantidade de ovos similares. Foi utilizado o mesmo procedimento para análise da atividade do experimento anterior, porém, cada experimento ocorreu sob diferentes temperaturas: 20°C, 25°C e 30°C. Para a análise dos dados obtidos foram utilizados os testes T e ANOVA multifatorial. Para o experimento de competição, fêmeas de ambas as espécies oriundas de competição de baixa densidade foram maiores do que as oriundas de desenvolvimento sob alta densidade larvária. A atividade das fêmeas de Ae. aegypti e Ae. albopictus de ambas as densidades foi muito semelhante, sugerindo que fêmeas menores tem atividade equivalente à de fêmeas maiores. Já no experimento que avaliou a atividade das fêmeas sob faixas térmicas distintas, notou-se que, em Ae. aegypti, houve diferença significativa na atividade total sob 20°C quando comparadas a 25°C e 30°C, demonstrando que a temperatura mais baixa foi limitante para a atividade desta espécie. Houve diferenças em algumas médias avaliadas na atividade de Ae. albopictus, porém, esta espécie pareceu ter sido menos influenciada com a variação da temperatura. Análises estatísticas confirmaram que as fêmeas de Ae. aegypti respondem de forma distinta das fêmeas de Ae. albopictus quando as temperaturas foram modificadas. O presente estudo ajuda a esclarecer alguns aspectos relacionados ao comportamento dos mosquitos Ae. aegypti e Ae. albopictus, que são considerados de grande importância para a saúde pública. / Diseases such as dengue, chikungunya, Zika and urban yellow fever are caused by pathogens transmitted by the bites of infected females of Ae. aegypti and Ae. albopictus. Biotics factors essentially competition on larval stages, and abiotics factors, for instance, temperature variation, occasion adjustments on development cycle of these species. The present study evaluated the effects of intraspecific competition and temperature on locomotor activity of Ae. aegypti and Ae. albopictus. The aims of this study were: to compare the size of females Ae. aegypti and Ae. albopictus from breeding sites with low and high intraspecific competition; to evaluate the effect of intraspecific competition on locomotor activity of virgin females of Ae. aegypti and Ae. albopictus, under regime of 12 hours of light and 12 hours of dark at 25°C and to analyze the pattern of the locomotor activity of virgins females of Ae. aegypti and Ae. albopictus, under regime of 12 hours of light and 12 hours of dark, under different temperatures (20ºC, 25ºC e 30ºC). The mosquitoes colonies used on experiment are from FIOCRUZ-RJ. For the competition experiment, larvae from both species were raised under conditions of high (100 larvae) and low (20 larvae) densities. For this 14 containers filled with 500 ml of water and 0,2 g of fish food were used. After the competition, the females from each density were transferred to incubators to perform locomotor activity analysis. After the activity, the size of adult from each larval density, were verified by wings measurement of each female. For the analysis of the locomotor activity under different temperatures, the females used were developed in an environment with similar amount of eggs. The same procedure was used to the analyze the activity of the previous experiment, however, each experiment happened under different temperatures: 20°C, 25°C e 30°C. For the analysis of the obtained data were applied T tests and ANOVA multivariate. For the competition experiment, females of both species from low density of competition were longer than females from competition under high larval density. The activity of females of Ae. aegypti and Ae. albopictus from both densities was very similar, suggesting that smaller females have equivalent activity to that of larger females. In the experiment that evaluated the activity of females under different thermal bands, it was noticed that in Ae. aegypti there was significant difference in total activity under 20°C when compared with 25°C and 30°C degrees, demonstrating the lower temperature was limiting to activity for this species. There were differences in some averages evaluated in Ae. albopictus activity, however, this species seemed to have been less influenced with the temperature variation. Statistical analysis confirmed that females of Ae. aegypti respond differently than females of Ae. albopictus when the temperatures were modified. The present study helps to clarify some aspects about the behavior of mosquitoes Ae. aegypti and Ae. albopictus which are considered of great concern for public health.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / O Aedes albopictus à vetor de diversas arboviroses e utiliza os criadouros naturais como local de reproduÃÃo. O objetivo deste estudo foi descrever os determinantes da infestaÃÃo pelo Ae. albopictus em Ãreas verdes na cidade de Fortaleza. Trata-se de estudo descritivo exploratÃrio realizado em quatro Ãreas com alta concentraÃÃo de Ãrvores e presenÃa de criadouros naturais. Foram caracterizados os aspectos ambientais e fÃsicos dos criadouros, a dinÃmica populacional e a possibilidade de criadouros naturais servirem como fonte mantenedora de ovos viÃveis na estaÃÃo nÃo chuvosa. O estudo foi composto por duas etapas ao longo do ano de 2007. A primeira etapa realizada na estaÃÃo chuvosa, com coleta de toda a Ãgua existente dentro dos criadouros, naturais ou artificiais. A segunda etapa foi realizada na estaÃÃo nÃo chuvosa, quando foi coletado o material interno dos criadouros naturais. As caracterÃsticas dos criadouros foram classificadas de acordo com a literatura e relacionadas com a presenÃa de formas imaturas de Ae. albopictus. Na estaÃÃo chuvosa, dos 62 criadouros existentes 49 (79%) eram naturais e destes 25 (51%) estavam infestados pelo Ae. albopictus. Dentre as Ãrvores com criadouros positivos as do gÃnero Terminalia, AcÃcia, Bambusa e Magnifera se destacaram. O Ae. albopictus foi habitante solitÃrio em 24 (96%) dos criadouros naturais e a maior densidade de larvas se deu nos criadouros de menor volume e mais prÃximos do solo. Nove (37,5%) criadouros se mantiveram infestados nas duas estaÃÃes e outros cinco apenas na estaÃÃo chuvosa (14; 30,4%; IC 95%: 16,62 â 44,25). Conclui-se que os criadouros naturais, em Ãrvores de vÃrios gÃneros, oferecem condiÃÃes adequadas para a manutenÃÃo de formas imaturas de Ae. albopictus. Neste sentido pode haver sua propagaÃÃo na estaÃÃo chuvosa seguinte na Ãrea urbana de Fortaleza. Esse cenÃrio amplia os desafios para o controle do dengue e de outros processos infecciosos transmitidos por este vetor. / The Aedes albopictus mosquito is a vector of several different arboviruses and uses natural breeding sites for its reproduction. The objective of this study was to describe the determinants of Ae. albopictus infestation in verdant areas of Fortaleza, Cearà (Brazil). It is a descriptive, exploratory study conducted in four areas of the city with a high density of vegetation and the presence of natural breeding sites. The environmental, populational and physical aspects of these areas were detailed as well as the possibility that they serve as a source of protection for viable eggs in the dry season. The study was composed of two parts conducted throughout 2007. The first was undertaken during the rainy season with water samples taken from natural and artificial breeding sites. The second took place in the dry season and samples were collected only from within the natural breeding sites. These areas were classified in accordance with the literature and correlated with the presence of immature forms of Ae. albopictus. During the rainy period, of the 62 breeding areas in existence, 49 (79%) were natural and of these, 25 (51%) were infested with Ae. albopictus. Among the trees that tested positively in these areas, the genera Terminalia, Acacia, Bambusa and Magnifera stood out, while in the artificial breeding sites tires, opened coconuts and small plastic containers were most common. Ae. albopictus was the lone inhabitant of 24 (96%) of the natural breeding sites and the greatest density of larvae was found in areas of lesser vegetation and closer to the soil, whereas tires held the higher density within the artificial areas. Nine (37.5%) natural breeding sites continued to be infested in the two seasons and five others only in the rainy season (14; 30.4%; CI 95%: 16.62 â 44.25). In general terms, there was infestation of 51.0% of natural breeding only in the rainy season, 30.4% in only the dry season and 63.8% if considered in totality. One can conclude that trees of several genera in natural breeding areas offer adequate conditions for the development of immature forms of Ae. albopictus at several stages. In this sense, there may be expanded propagation in the next rainy season in the urban areas of Fortaleza. This scenario amplifies the challenges inherent in the control of dengue fever and other infectious diseases transmitted by this vector.

Made available in DSpace on 2015-04-22T22:14:01Z (GMT). No. of bitstreams: 1 Ilia Gilmara Carvalho.pdf: 530450 bytes, checksum: 50f84ac4bf26a69a84cf2a4b2a031978 (MD5) Previous issue date: 2008-12-31 / Fundação de Amparo à Pesquisa do Estado do Amazonas / The study had for objective the detection and typing of the dengue virus, in the vectors Aedes aegypti and Aedes albopictus through the Polimerase Chain Reaction (PCR) using specific primers. During the period of December of 2005 to December of 2006 8984 mosquitoes were collected, of these 827 they were Aedes, being 819 Ae. aegypti (414 females and 405 males) and 8 Ae. albopictus (7 females and 1 male), in 46 neighborhoods of the city of Manaus including all of the geographical areas of the city. The females were grouped in pools from 1 to 10 mosquitoes for microtubes, in agreement with the gender, it dates from collection, neighborhood and stocked to -70C in the same day of the collection, totaling 143 pools (138 of Ae. aegypti and 5 of Ae. albopictus). Those pools were macerated in PBS containing albumin, and a bracket of the softened was removed for inoculation in cell culture, and other bracket was used to accomplish the extraction of the viral RNA. Once extracted, RNA was submitted the reaction of reverse transcription following by the chain reaction of the polimerase (RT-PCR). The positive samples in RT-PCR were submitted the reaction of semi nested PCR and the products of this reaction were resolved in agarose gel to 1,5% with etídio bromide. Of the 143 analyzed pools, 114 (111 of Ae. aegypti and 3 of Ae. albopictus) they were positive for DENV 3, just a pool was shown positive for two sorotipos, DENV 1 and DENV 3. The prevalence pools Ae. aegypti infected with DENV 3 in the city of Manaus was of 53%, being larger in the areas Center-west (70%), South (60%) and West (53%), and smaller in the area East (23%). This is the first report of the detection of DENV 3 in field-caught Ae. albopictus naturally infected in Brazil. The monitoring of the circulation viral in mosquitoes with the use of the technique of RT-PCR allows the previous knowledge of the levels of spread viral in certain areas contributing to determine the time and place to apply the prevention measures and control. / O estudo teve por objetivo a detecção e tipagem do vírus dengue, nos vetores Aedes aegypti e Aedes albopictus por meio da Reação em Cadeia da Polimerase (PCR) utilizando primers específicos. Durante o período de Dezembro de 2005 a Dezembro de 2006 foram coletados 8984 mosquitos, destes 827 eram Aedes, sendo 819 Ae. aegypti (414 fêmeas e 405 machos) e 8 Ae. albopictus (7 fêmeas e 1 macho), em 46 bairros da cidade de Manaus abrangendo todas as zonas geográficas da cidade. As fêmeas de Ae. aegypti e Ae. albopictus foram agrupadas em pools de 1 a 10 mosquitos por microtubos, de acordo com o gênero, data de coleta, bairro e estocado a -700C no mesmo dia da coleta, totalizando 143 pools (138 de Ae. aegypti e 5 de Ae. albopictus). Esses pools foram macerados em PBS contendo albumina, e uma alíquota do macerado foi retirada para inoculação em cultura de célula, e outra alíquota foi utilizada para realizar a extração do RNA viral. Uma vez extraído, o RNA foi submetido a reação de transcrição reversa seguida da reação em cadeia da polimerase (RT-PCR). As amostras positivas na RT-PCR foram submetidas a reação de Semi Nested PCR e os produtos desta reação foram resolvidos em gel de agarose a 1,5% corado com brometo de etídio. Dos 143 pools analisados, 114 (111 de Ae. aegypti e 3 de Ae. albopictus) foram positivos para DENV 3, apenas um pool mostrou-se positivo para dois sorotipos, DENV 1 e DENV 3. A prevalência de mosquitos Ae. aegypti infectados com DENV 3 na cidade de Manaus foi de 80%. Em todas as zonas a prevalência foi maior que 53%, sendo maior nas zonas Centro-oeste (70%), Sul (60%) e Oeste (53%), e menor na zona Leste (23%). Este foi o primeiro achado no Brasil da identificação do DENV 3 em amostras de Ae. albopictus , coletados em campo na forma adulta. O monitoramento da circulação viral em mosquitos com o uso da técnica de RT-PCR permite o conhecimento prévio dos níveis de disseminação viral em determinadas áreas contribuindo para determinar a época e local para aplicar as medidas de prevenção e controle.

Le moustique tigre Aedes albopictus, pur produit de la mondialisation, s’est installé en Europe dans les années 1970s, et en région PACA en 2004, grâce au transport intercontinental de ses œufs, capables de résister à la dessiccation et d’effectuer une diapause. Ce moustique est capable de transmettre 27 virus comme la dengue, zika et chikungunya. Le premier objectif du présent travail a été l’étude des aspects biologique d’Ae. albopictus dans son milieu naturel dans une région où il s’est implanté depuis plus de 10 ans, à Nice au sud-est de la France. Nous avons utilisé différentes méthodes de captures : les pièges pour adultes BGs ®, les pièges pondoirs avec une infusion de foin pour la récolte des œufs ainsi que la capture sur Homme, afin d’apprécier la densité de ce moustique, ses fluctuations spatiotemporelles et les taux de parturité des femelles. Les deux années de l’étude de la dynamique saisonnière ont montrés un début d’activité des moustiques au mois d’avril atteignant un pique d’activité en Juillet-Août et diminuant par la suite jusqu'à ce que l'activité a cessé au mois de novembre. Nous avons enregistré des taux de parturité élevées qui se sont stabilisés entre 0,52 et 0,71 entre le mois de juin et octobre respectivement. Ces taux de parturités indiquent un taux de survie élevé qui pourrait être encore supérieur à cause du comportement de skip oviposition des femelles Ae. albopictus. De plus nous avons mis en évidence un taux important de femelles gravides (jusqu’à 44 %) capturées sur Homme indiquant une prise de repas multiples durant un même cycle gonotrophique. L'objectif principal de ce travail a été d'évaluer l'efficacité des pulvérisations spatiales d’Ultra Bas Volume (UBV) d’insecticides, la principale méthode utilisée dans le cas de foyers urbains. La population locale d’Ae. albopictus a montré une forte sensibilité à la deltaméthrine (le seul insecticide homologué pour lutter contre les moustiques adultes en France) bien que les mâles ont été beaucoup plus sensible que les femelles. Une comparaison des résultats par application topique et par les tests en tube OMS a confirmé que cette différence a été due à la plus petite taille des mâles par rapport aux femelles. Durant six essais sur le terrain, on n’a obtenu aucun impact significatif des traitements UBV de deltaméthrine sur les femelles, bien qu'il y ait eu une réduction significative des mâles. En revanche, le traitement par thermonébulisation autour d’un petit groupe de maisons a permis une quasi-élimination (95%) des moustiques mâles et femelles. Ces résultats apportent un élément de réponse sur les stratégies à mener en santé publique. Enfin, l'efficacité de plusieurs formulations de peintures insecticides issues de la technologie de microencapsulation ont été testée dans des conditions de laboratoire et de terrain. Les résultats ont été encourageants et les tests ont démontré que les peintures à base d’organophosphorés ont une rémanence de 2 ans. Le but ultime était d'utiliser des pièges pondoirs traités avec ces peintures insecticides comme une méthode de contrôle: en théorie, cette méthode vise l’exploitation du comportement de skip oviposition qui oblige les femelles à déposer leurs œufs sur plusieurs gîtes différents augmentant la probabilité pour que celles-ci rencontrent un piège pondoir traité avec un insecticide. Dans un petit essai sur le terrain, cependant, aucun impact sur la population sauvage n’a été observé. Nous discutons de cette méthode ainsi que de son intérêt face à la menace toujours grandissante liée à la propagation continuelle d’Ae. albopictus à travers le monde. / The Asian Tiger mosquito, Aedes albopictus, first appeared in Europe in the 1970s,almost certainly a result ofintercontinental transport of its dessication-resistant eggs. In the laboratory,the species is capable of transmitting 27 viruses including Yellow Fever, dengue, chikungunya and zika, although in the field it is not considered an efficient vector because it is not host-specific.The first objective of our studies was to monitor aspects of the biology of the species in Nice (southeastern France), where it has been established for more than 10 years. Infusion-baited ovitraps, B-G® adult traps and human landing captures were used to reveal seasonal population fluctuations, parous rates and other parameters. In two complete years of study, adult mosquitoes began to appear in April, reached a plateau in July-August and declined thereafter until activity ceased in November. Parous rates were stable from June to October (0,52 to 0,71%) indicating a balance between recruitment (eclosion) and mortality. During this period, daily survival probability was high although this may be an underestimate if the duration of the gonotrophic cycle is extended by skip oviposition. Up to 44% of females captured by landing capture were gravid, confirmation of multiple blood-meals per gonotrophic cycle.The main focus of the work was an assessment of the efficacy of Ultra-low Volume (ULV) space sprays, the principal method used in the event of urban outbreaks. The local population proved highly susceptible to deltamethrin (the only insecticide approved for adult mosquito control in France) although femaleswere significantly less susceptible than males. A comparison of results by topical application and by the WHO tube test confirmed this difference is a function of size. In six separate field trials,there was no evidence of any impact of ULVdeltamethrin on females although there was a significant reduction of males. By contrast, outdoor treatment of a small cluster of homes with a portable thermal fogger gave virtual elimination (ca. 95%) of both sexes.These results are clearly of public health significance.Lastly, the efficacy of several slow-release formulations of microencapsulated insecticide in paint was tested in laboratory and semi-field conditions. Results were encouraging and tests demonstrated that the paints remained effective for 24 months on plastic surface. The ultimate goal was to use paint-treated ovitraps as a control method: in theory, skip ovipositon should ensure a high probability that ovipositing females will make contact with a treated trap on their rounds of suitable oviposition sites. In a small field trial, however, no impact on the wild population was observed. This may have been because too few treated traps were deployed; further studies along these lines could prove worthwhile.