Ecological characterisation and infection of Anophelines (Diptera: Culicidae) of the Atlantic Forest in the southeast of Brazil over a 10 year period: has the behaviour of the autochthonous malaria vector changed?

BACKGROUND In southeastern Brazil, autochthonous cases of malaria can be found near Atlantic Forest fragments. Because the transmission cycle has not been completely clarified, the behaviour of the possible vectors in those regions must be observed. A study concerning the entomological aspects and natural infection of anophelines (Diptera: Culicidae) captured in the municipalities of the mountainous region of Espírito Santo state was performed in 2004 and 2005. Similarly, between 2014 and 2015, 12 monthly collections were performed at the same area of the study mentioned above. METHODS Center for Disease Control (CDC) light traps with CO2 were set in open areas, at the edge and inside of the forest (canopy and ground), whereas Shannon traps were set on the edge. FINDINGS A total of 1,414 anophelines were collected from 13 species. Anopheles (Kerteszia) cruzii Dyar and Knab remained the most frequently captured species in the CDC traps set in the forest canopy, as well as being the vector with the highest prevalence of Plasmodium vivax/simium infection, according to molecular polymerase chain reaction techniques. CONCLUSIONS P. vivax/simium was found only in abdomens of the mosquitoes of the subgenus Nyssorhynchus, weakening the hypothesis that this subgenus also plays a role in malaria transmission in this specific region.

Ecological aspects of Pintomyia fischeri and Migonemyia migonei in municipalities with Canine Visceral Leishmaniasis, State of São Paulo, Brazil.

The objective was to study the composition of sand fly species, the seasonal activity of the dominant species and correlation with average rainfall, sex association of the dominant species with domicile habitats and analyze different landscape scenarios for the two study sites in the municipalities of Itapevi and Mogi of the Crosses. Sandflies were captured with CDC(s) traps installed in three households at each study site for a period of 24 hours, biweekly for two years. A total of 2970 specimens were collected. The dominant species were P.fischeri and M.migonei. A statistically significant difference was registered between males and females of the two species in relation to the intra and peridomicile. The seasonal activity of both species showed a weak significant positive correlation with rainfall. The species P.fischeri and M.migonei may be potential vectors of CVL in the studied sites.

DNA Barcoding of Morphologically Characterized Mosquitoes Belonging to the Genus Mansonia from the Atlantic Forest and Brazilian Savanna.

The identification of mosquito species is necessary for determining the entomological components of disease transmission. However, identification can be difficult in species that are morphologically similar. The cytochrome c oxidase subunit I (COI) DNA barcode region is considered a valuable and reliable diagnostic tool for mosquito species recognition, including those that belong to species complexes. Mansonia mosquitoes are found in forests near swampy areas. They are nocturnal and are highly attracted to light. Hematophagous adult females exhibit aggressive biting behavior and can become infected with and transmit pathogens during their feeding, including some epizootic viruses and avian malaria. In Brazil, twelve Mansonia species have been reported. In a recent study from the São Paulo Zoo in Brazil, three morphologically distinct species were collected and identified, namely: Mansonia (Mansonia) indubitans, Ma. (Man.) pseudotitillans and Ma. (Man.) titillans. However, confirmation of these species by molecular identification was unsuccessful due to a lack of COI sequences in the GenBank database. Thus, this research aimed to describe the COI DNA barcode sequences of some morphologically characterized Mansonia (Man.) species from Brazil and to determine their utility in delimiting species collected from the Atlantic Forest and Brazilian Savanna. Accordingly, we provide tools for the genetic identification of species that play a significant role in pathogen transmission in wildlife and potentially humans. We show that the delimitation of Mansonia species via five different approaches based on COI DNA sequences (BI, NJ, ASAP, bPTP and GMYC) yield basically the same groups identified by traditional taxonomy, and we provide the identification of specimens that were previously identified only up to the subgenus level. We also provide COI sequences from two Mansonia species that were not previously available in sequence databases, Ma. wilsoni and Ma. pseudotitillans, and thus contribute to the ongoing global effort to standardize DNA barcoding as a molecular means of species identification.

Assessing Diversity, Plasmodium Infection and Blood Meal Sources in Mosquitoes (Diptera: Culicidae) from a Brazilian Zoological Park with Avian Malaria Transmission.

Avian malaria parasites are widespread parasites transmitted by Culicidae insects belonging to different genera. Even though several studies have been conducted recently, there is still a lack of information about potential vectors of Plasmodium parasites, especially in Neotropical regions. Former studies with free-living and captive animals in São Paulo Zoo showed the presence of several Plasmodium and Haemoproteus species. In 2015, a pilot study was conducted at the zoo to collect mosquitoes in order to find out (i) which species of Culicidae are present in the study area, (ii) what are their blood meal sources, and (iii) to which Plasmodium species might they be potential vectors. Mosquitoes were morphologically and molecularly identified. Blood meal source and haemosporidian DNA were identified using molecular protocols. A total of 25 Culicidae species were identified, and 6 of them were positive for Plasmodium/Haemoproteus DNA. Ten mosquito species had their source of blood meal identified, which were mainly birds, including some species that were positive for haemosporidian parasites in the former study mentioned. This study allowed us to expand the list of potential vectors of avian malaria parasites and to improve our knowledge of the evolutionary and ecological relationships between the highly diverse communities of birds, parasites, and vectors present at São Paulo Zoo.

Wing Geometric Morphometrics as a Tool for the Identification of Culex Subgenus Mosquitoes of Culex (Diptera: Culicidae).

Culex is the largest subgenus within the genus Culex that includes important vectors of diseases. The correct identification of mosquitoes is critical for effective control strategies. Wing geometric morphometrics (WGM) has been used to identify mosquito species alongside traditional identification methods. Here, WGM was used for eleven Culex species from São Paulo, Brazil, and one from Esquel, Argentina. Adult mosquitoes were collected using CDC (Centers for Disease Control) traps, morphologically identified and analyzed by WGM. The canonical variate analysis (CVA) was performed and a Neighbor-joining (NJ) tree was constructed to illustrate the patterns of species segregation. A cross-validated reclassification test was also carried out. From 110 comparisons in the cross-validated reclassification test, 87 yielded values higher than 70%, with 13 comparisons yielding 100% reclassification scores. Culexquinquefasciatus yielded the highest reclassification scores among the analyzed species, corroborating with the results obtained by the CVA, in which Cx. quinquefasciatus was the most distinct species. The high values obtained at the cross-validated reclassification test and in the NJ analysis as well as the segregation observed at the CVA made it possible to distinguish among Culex species with high degrees of confidence, suggesting that WGM is a reliable tool to identify Culex species of the subgenus Culex.

[Aedes albopictus seasonality in a protected area in the city of São Paulo, Brazil]. / Sazonalidade de Aedes albopictus em área protegida na cidade de São Paulo, Brasil.

The study had the objective of assessing the seasonality of Aedes albopictus and the impact of environmental factors on breeding sites in a protected urban area. Immature individuals were collected through fluid aspiration from nine tree holes, in the Ecological Park of Tietê in the city of São Paulo, Southern Brazil, from 2001 to 2002. The index of positivity and number of individuals results were as follows: positivity x precipitation (rho=0.69, p<0.001), positivity x temperature (rho=0.35, p<0.001), number of individuals x precipitation (rho=0.29, p<0.001) and number of individuals x temperature (rho=0.13, p<0.05). These correlations suggest rainfalls had greater impact than temperature.

Studies on entomological monitoring: mosquito species frequency in riverine habitats of the Igarapava Dam, Southern Region, Brazil.

Diversity of mosquito species was evaluated in different habitats before and after the Igarapava reservoir flooding in the Grande River, Southern Cerrado of Brazil. We aimed at verifying changes in these mosquito populations in consequence of the lake formation. Four habitats were selected as sampling stations: peridomiciliary habitat, pasture, "veredas" and gallery forest patch. Bimonthly collections were made with the Shannon trap and human bait, including diurnal, crepuscular and nocturnal period of mosquito activity. The Shannon Index results from the potential vectors were compared using Student t-test. Aedes scapularis, Anopheles darlingi and An. albitarsis senso latu seasonal abundance were described with moving average and compared using chi2 test. There were changes in the mosquito frequency in the habitats, except for the "veredas" that was 13 km away from the catchment area. The altering in mosquito species seasonal abundance suggests breeding places expansion. Diversity indexes can be used to monitor changes in mosquito vector population in environments where abrupt disturbance can alter disease transmission cycles.

Ecological characterisation and infection of Anophelines (Diptera: Culicidae) of the Atlantic Forest in the southeast of Brazil over a 10 year period: has the behaviour of the autochthonous malaria vector changed?

BACKGROUND In southeastern Brazil, autochthonous cases of malaria can be found near Atlantic Forest fragments. Because the transmission cycle has not been completely clarified, the behaviour of the possible vectors in those regions must be observed. A study concerning the entomological aspects and natural infection of anophelines (Diptera: Culicidae) captured in the municipalities of the mountainous region of Espírito Santo state was performed in 2004 and 2005. Similarly, between 2014 and 2015, 12 monthly collections were performed at the same area of the study mentioned above. METHODS Center for Disease Control (CDC) light traps with CO2 were set in open areas, at the edge and inside of the forest (canopy and ground), whereas Shannon traps were set on the edge. FINDINGS A total of 1,414 anophelines were collected from 13 species. Anopheles (Kerteszia) cruzii Dyar and Knab remained the most frequently captured species in the CDC traps set in the forest canopy, as well as being the vector with the highest prevalence of Plasmodium vivax/simium infection, according to molecular polymerase chain reaction techniques. CONCLUSIONS P. vivax/simium was found only in abdomens of the mosquitoes of the subgenus Nyssorhynchus, weakening the hypothesis that this subgenus also plays a role in malaria transmission in this specific region.

[The presence of Culicidae species in medium-sized cities in the State of São Paulo, Brazil and the risk of West Nile fever and other arbovirus infection]. / Presença de culicídeos em município de porte médio do Estado de São Paulo e risco de ocorrência de febre do Nilo Ocidental e outras arboviroses.

INTRODUCTION: The objectives for this study were to measure the diversity of Culicidae species, describe their abundance and seasonal variation in São José do Rio Preto, SP, and discuss the risk of arbovirus infections. METHODS: The collection of larval and adult mosquitoes was conducted monthly from 2006 to 2007 in an urban area and four sections of forested land. In the urban area, larvae were collected from sites where oviposition by Culex mosquitoes was most likely to occur. At two of the four sites in the forested land, adult mosquitoes were collected with the use of CDC traps at night, and a Nasci aspirator was used in the daytime at the two other collection sites. RESULTS: In the urban area, 34 Culicidae species were identified out of a total sample of 8,683 specimens; of these specimens, 80.7% were Culex quinquefasciatus, 9.6% were Culex coronator, 3.2% were Aedes albopictus, and 1.1% were Ochlerotatus fluviatilis. The abundance of Cx. quinquefasciatus larvae was negatively related to rainfal. In the woods, 2,268 mosquitoes were collected, representing 10 genera and 46 species. The most abundant mosquito species were Aedeomyia squamipennis, Culex coronator, Culex (Mel.) Melanoconion section, Culex declarator, Ochlerotatus scapularis, Anopheles triannulatus, Culex bidens/interfor and Culex habilitator/pseudojhantinosoma. CONCLUSIONS: The abundance of Cx. quinquefasciatus in the urban area and the presence of other Culicidae species in urban areas and forested land point to the possibility of the transmission of West Nile virus and other arbovirus infections in São José do Rio Preto and other cities. Thus, the enacting of measures aimed at the surveillance of these arbovirus infections is essential.

Presença de culicídeos em município de porte médio do Estado de São Paulo e risco de ocorrência de febre do Nilo Ocidental e outras arboviroses / The presence of Culicidae species in medium-sized cities in the State of São Paulo, Brazil and the risk of West Nile fever and other arbovirus infection

INTRODUÇÃO: O objetivo deste estudo foi mensurar a diversidade de espécies de culicídeos, descrever sua abundância e variação sazonal em áreas urbanas e matas de São José do Rio Preto, SP, e discutir o risco de ocorrência de arboviroses. MÉTODOS: Coletas de larvas e de mosquitos adultos foram realizadas mensalmente, em 2006 e 2007, em área urbana e em quatro fragmentos de mata. No perímetro urbano, coletaram-se larvas nos sítios mais prováveis de oviposição para mosquitos do gênero Culex e nas matas foi realizada a coleta de mosquitos adultos, sendo que em duas utilizaram-se armadilhas CDC à noite e, em duas, aspirador de Nasci de dia. RESULTADOS: Na área urbana identificaram-se 34 espécies de culicídeos em um total de 8.683 exemplares; destes, 80,7% corresponderam ao Culex quinquefasciatus, 9,6% ao Culex grupo Coronator, 3,2% ao Aedes albopictus (3,2%) e 1,1% ao Ochlerotatus fluviatilis. A abundância de larvas de Cx. quinquefasciatus correlacionou-se negativamente com a chuva. Nas quatro matas, foram coletados 2.268 mosquitos distribuídos entre 10 gêneros, 46 espécies ou grupos. As mais abundantes foram Aedeomyia squamipennis, Culex. coronator, Culex (Mel.) seção Melanoconion, Culex declarator, Ochlerotatus scapularis, Anopheles triannulatus, Culex bidens/interfor e Culex habilitator/pseudojhantinosoma. CONCLUSÕES: A abundância de Cx. quinquefasciatus na área urbana e a presença de outros culicídeos nas áreas urbanas e de matas apontam para a possibilidade de transmissão do vírus do Nilo Ocidental e de outras arboviroses em São José do Rio Preto e outras cidades do Brasil, sendo fundamental o estabelecimento de medidas visando à vigilância destas arboviroses.

INTRODUCTION: The objectives for this study were to measure the diversity of Culicidae species, describe their abundance and seasonal variation in São José do Rio Preto, SP, and discuss the risk of arbovirus infections. METHODS: The collection of larval and adult mosquitoes was conducted monthly from 2006 to 2007 in an urban area and four sections of forested land. In the urban area, larvae were collected from sites where oviposition by Culex mosquitoes was most likely to occur. At two of the four sites in the forested land, adult mosquitoes were collected with the use of CDC traps at night, and a Nasci aspirator was used in the daytime at the two other collection sites. RESULTS: In the urban area, 34 Culicidae species were identified out of a total sample of 8,683 specimens; of these specimens, 80.7% were Culex quinquefasciatus, 9.6% were Culex coronator, 3.2% were Aedes albopictus, and 1.1% were Ochlerotatus fluviatilis. The abundance of Cx. quinquefasciatus larvae was negatively related to rainfal. In the woods, 2,268 mosquitoes were collected, representing 10 genera and 46 species. The most abundant mosquito species were Aedeomyia squamipennis, Culex coronator, Culex (Mel.) Melanoconion section, Culex declarator, Ochlerotatus scapularis, Anopheles triannulatus, Culex bidens/interfor and Culex habilitator/pseudojhantinosoma. CONCLUSIONS: The abundance of Cx. quinquefasciatus in the urban area and the presence of other Culicidae species in urban areas and forested land point to the possibility of the transmission of West Nile virus and other arbovirus infections in São José do Rio Preto and other cities. Thus, the enacting of measures aimed at the surveillance of these arbovirus infections is essential.

Sazonalidade de Aedes albopictus em área protegida na cidade de São Paulo, Brasil / Aedes albopictus seasonality in a protected area in the city of São Paulo, Brazil

O estudo teve por objetivo analisar a sazonalidade de Aedes albopictus e a influência de fatores ambientais em criadouros, em uma área urbana protegida. Foram realizadas coletas de indivíduos imaturos por sucção do líquido de nove ocos de árvores, no Parque Ecológico do Tietê, no Município de São Paulo, SP, de 2001 a 2002 . Os resultados do índice de positividade e do número de indivíduos foram: positividade x precipitação (ro=0,69; p<0,001); positividade x temperatura (ro=0,35; p<0,001); número de indivíduos x precipitação (ro=0,29; p<0,001) e número de indivíduos x temperatura (ro=0,13; p<0,05). As correlações sugerem que as chuvas foram mais influentes que a temperatura.

The study had the objective of assessing the seasonality of Aedes albopictus and the impact of environmental factors on breeding sites in a protected urban area. Immature individuals were collected through fluid aspiration from nine tree holes, in the Ecological Park of Tietê in the city of São Paulo, Southern Brazil, from 2001 to 2002. The index of positivity and number of individuals results were as follows: positivity x precipitation (rho=0.69, p<0.001), positivity x temperature (rho=0.35, p<0.001), number of individuals x precipitation (rho=0.29, p<0.001) and number of individuals x temperature (rho=0.13, p<0.05). These correlations suggest rainfalls had greater impact than temperature.

Studies on entomological monitoring: mosquito species frequency in riverine habitats of the Igarapava Dam, Southern Region, Brazil

Avaliou-se a diversidade de mosquitos em diferentes habitats antes e depois da formação do reservatório de Igarapava no Rio Grande, Cerrado do Sudeste do Brasil, com objetivo de verificar alteração nessas populaçäes de mosquitos. Quatro habitats foram selecionados como sítios de coleta: a) peridomicílio rural, b) pastagem, c) veredas e d) mata ciliar. Realizaram-se coletas bimestrais com isca humana e armadilha de Shannon, incluindo períodos de atividade culicídea diurna, crepuscular vespertina e noturna. Os resultados do Indice de Shannon para os potenciais vetores nos diferentes habitats foram comparados pelo teste t de Student. As abundâncias sazonais de Aedes scapularis, Anopheles darlingi e Anopheles albitarsis s.l., foram descritas com a média móvel e comparadas com o teste c2. Houve alteração nas freqüências dos mosquitos nos habitats, com exceção das veredas, que estavam afastadas 13 km da área de influência do reservatório. A modificação nas abundâncias sazonais dos potenciais vetores sugere o incremento de criadouros após a formação do reservatório. Indices de diversidade são ferramentas úteis para monitorar populaçäes de mosquitos vetores em ambientes cujas mudanças drásticas podem alterar ciclos de transmissão de doenças transmitidas por vetores.

Aspectos bioecológicos de Culex quinquefasciatus e Ochlerotatus scapularis, abrigados em habitats do Parque Ecológico do Tietê na cidade de Säo Paulo, SP / Bioecological aspects of Culex quinquefasciatus and Ochlerotatus scapularis from habitats of the Ecological Park by the river Tietê, City of Säo Paulo, SP

Objetivos. Diante da importância epidemiológica de CULEX QUINQUEFASCIATUS e OCHLEROTATUS SCAPULARIS e da conhecida dominância dessas populaçöes em áreas do Parque Ecológico do Tietê, objetivou-se estudar ao longo das estaçöes do ano aspectos bioecológicos das duas populaçöes abrigadas em diferentes habitats. Métodos. Realizaram-se coletas quinzenais nos diferentes habitats com auxílio de aspirador a bateria. O estado fisiológico e o tamanho do corpo das fêmeas das duas populaçöes foi verificada em laboratório a partir de amostras aleatórias de cada habitat. Resultados. A proporçäo entre os sexos revelou um maior predomínio de machos entre os habitats, esta diferença foi mais significante para CULEX QUINQUEFASCIATUS. OCHLEROTATUS SCAPULARIS apresentou um percentual maior de fêmeas paridas comparado a CULEX QUINQUEFASCIATUS nos diferentes habitats. Essa diferença foi significante com. A distribuiçäo de fêmeas paridas e nulíparas de CULEX QUINQUEFASCIATUS nos nos habitats do interior do parque, foi mais significante comparada à distribuiçäo de OCHLEROTATUS SCAPULARIS. As fêmeas paridas dos mosquitos foram numericamente mais expressivas ao longo da estaçäo chuvosa, nos habitats da periferia e interior do parque. As duas populaçöes apresentaram fêmeas nos estágios III de Christophers e Mer com o intestino totalmente cheio de sangue vermelho, indício de discordância gonotrófica. CULEX QUINQUEFASCIATUS foi estatisticamente maior que OCHLEROTATUS SCAPULARIS (p < 0,001), em relaçäo ao tamanho do corpo, apresentando diferenças estatística significante tanto no período de chuva quanto no de seca (H=69,9 e P< 0,001). O coeficiente de regressäo (ß1 = -14,9) e o coeficiente de correlaçäo (r= -,062) indicaram para CULEX QUINQUEFASCIATUS um efeito negativo do tamanho sobre as fêmeas paridas. A funçäo do tamanho näo se explicou em fêmeas nulípiras e de machos de CULEX QUINQUEFASCIATUS, indica a proximidade do criadouro. O tamanho do corpo de CULEX QUINQUEFASCIATUS é maior que OCHLEROTATUS SCAPULARIS. O efeito do tamanho do corpo sobre o número de fêmeas paridas foi negativo. CULEX QUINQUEFASCIATUS e OCHLEROTATUS SCASPULARIS apresentam discordância gonotrófica. As fêmeas paridas de OCHLEROTATUS SCAPULARIS estiveram em consonância com as altas temperaturas e com os picos de densidade de fêmeas. Existe diferença estatisticamente significante na distribuiçäo de fêmeas paridas e nulíparas das duas populaçöes entre os habitats estudados