Larvicidal activity of hexane extract, fatty acids, and methyl esters from Tecoma stans pericarps against Culex quinquefasciatus.

Culex quinquefasciatus (Diptera: Culicidae) is responsible for the transmission of lymphatic filariasis. The search for natural sources of new insecticides to control mosquitoes has intensified in recent years. Tecoma stans is an exotic species in Brazil, known as 'ipê de jardim'. T. stans pericarps were extracted with hexane in Soxhlet apparatus, and fatty acids (FA) and methyl esters (FAME) were obtained by transesterification reaction of hexane extract (HE). HE, FA, and FAME were evaluated against 3rd. and 4th. instar larvae of Culex quinquefasciatus. Analysis by gas chromatography-mass spectrometry (GC/MS) identified the hydrocarbon nonacosane (81.69%) as the major compound in HE, and linolenic (16.89%), linoleic (16.83%), and palmitic acids (21.00%) were predominant in FA. FA and HE, at a concentration of 250 µg/mL, promoted mortality of 81.67% and 68.66% of larvae, respectively. HE and FA obtained from T. stans pericarps have larvicidal potential for the control of C. quinquefasciatus.

Assessment of the Toxicological Effects of Pesticides and Detergent Mixtures on Zebrafish Gills: a Histological Study

Abstract Pesticides, as Temephos and Roundup, and surfactants, like Sodium Dodecyl Sulfate (SDS), when discharged into the environment, are harmful to non-target wildlife, especially fish. To evaluate the effects of those chemicals alone and their binary combinations, we assessed the histological alterations in Danio rerio gills after 15 days of exposure to Temephos (1ppm), Roundup (10ppm), SDS (2ppm), SDS plus Temephos (2ppm+1ppm), SDS plus Roundup (2ppm+10ppm) and Temephos plus Roundup (1ppm+10ppm). Hyperplasia, a regressive change, was significant in all exposed groups. The progressive change, complete fusion of lamellae, was significantly intensive in Roundup plus Temephos group. Regarding circulatory disturbances, vascular congestion was significant intensive in SDS group, while severe aneurysm was observed in Roundup group. Although xenobiotics in the blend are biologically complex, mainly for long periods, combinatorial exposure incremented only complete fusion of lamellae. Our results reinforce the rationalization of indiscriminate use of those compounds alone or in combination.

Larvicidal effect from different Annonaceae species on Culex quinquefasciatus.

The recent outbreaks of mosquito-borne diseases highlighted the pivotal importance of mosquito vector control in tropical areas worldwide. Several strategies have been developed to control vector populations and disease transmission in endemic areas. The steps to obtain natural active compounds involve the pre-selection in a biological model and subsequently evaluation on specific models. The present study reports the evaluation of 35 extracts, fractions, and essential oils obtained from five species from the Annonaceae family on Artemia salina and Culex quinquefasciatus. The A. salina results were used as a pre-screening for larvicidal test about mosquitoes. A correlation of biological activity in both bioassays was observed for the hydroethanolic extracts and their respective hexane and chloroform fractions of the leaves of Annona species, except A. nutans. The same correlation was also observed for all tested essential oils and petroleum ether extracts from Duguetia species. It was possible to limit an interval of lethality about A. salina, which has a corresponding range to the larvicidal test against the mosquito. The main components present in D. lanceolata essential oil or enriched fraction were α-selinene, aristolochene, (E)-caryophyllene, and (E)-calamenene. For D. furfuracea, the main components present of the underground parts were (E)-asarone, 2,4,5-trimethoxystyrene, spathulenol, and bicyclogermacrene for aerial parts. The A. salina test could be used as a model for the pre-screening of larvicidal activity.

Evaluation of Culex quinquefasciatus wings asymmetry after exposure of larvae to sublethal concentration of ivermectin.

The surviving insects submitted to chemical control have morphological alterations that impact on their mechanisms of resistance and their final development. Those changes are detected and measured using physical features related to symmetry, specifically named fluctuating asymmetry. This is detected when deviations from the perfect bilateral symmetry for specific morphological characteristic is influenced by genetics or environmental stress. Thus, in this paper we analyze the wing in adult of Culex quinquefasciatus (Diptera - Culicidae) after larvae exposure to ivermectin LC50. Three hundred larvae of C. quinquefasciatus were exposed to ivermectin in 1.5 µg/L (LC50) concentration during 30 min, and three hundred larvae were exposed to distilled pure water as control group. For fluctuating asymmetry, adult males and adult females were selected from each group (n = 83) from the untreated group and (n = 79) from treated group. Wings from adults of each group were mounted in glass microscope slides and coverslip in Canada's balsam and analyzed with a stereomicroscope with a video camera attached. The treatment effect on M3 + 4 was marginally significant with higher asymmetry values in the control group. The data obtained here suggest the importance of future experiments to elucidate the mechanisms associated with FA. Moreover, according to the results obtained, it may be suggested that FA is present in females in ornaments, or secondary sexual characters, as an indicator of phenotypic quality of the partners.

Toxicity of different fatty acids and methyl esters on Culex quinquefasciatus larvae.

The Culex quinquefasciatus mosquito is a vector of several diseases, and its control has been performed with synthetic insecticides, which may have human and environmental side effects. Thus, the use of new and safe molecules are important, and this study evaluated the toxicity of active substances against this mosquito. The oleic, linoleic, linolenic, palmitic and stearic acids and their respective methyl esters were tested against fourth instar C. quinquefasciatus larvae. Oleic, linoleic and linolenic acids had LC50 values of 8.58, 10.04 and 19.78 mg/L, respectively. Histological analysis showed that these three compounds caused cell instability with an increase in the number of vesicles in the fat body and in the midgut cells. Based on these results, glucose, triglyceride, and protein levels were evaluated after 1 h of acid exposure. These compounds decreased in insects treated with linoleic acid. Linolenic acid also caused a significant increase in acetylcholinesterase activity. These results show that oleic, linoleic, and linoleic acids have a lower LC50 for C. quinquefasciatus, affecting its metabolism and the morphology of midgut and fat body.

Larvicidal activity of vegetable oils and esterified compounds against Culex quinquefasciatus (Diptera: Culicidae).

Control of Culex quinquefasciatus using chemical insecticides may result in the selection of resistant mosquito strains. Thus, the use of plant-derived products has been studied as alternative for the mosquito control. Fatty acid methyl esters (FAMEs) obtained by transesterification of vegetable oils may result in compounds with larvicidal potential against C. quinquefasciatus. However, little is known about the morphological, physiological or biochemical effects that these FAMEs may have on mosquito larvae. The present study reports the effects of these FAMEs in mosquito larvae. The FAMEs were obtained by transesterification of canola, corn, sunflower, and soybean oils with acid catalysis and the determination of FAMEs composition was done by gas chromatography-mass spectrometry (GC-MS). Larvae of C. quinquefasciatus were exposed to different concentrations of the vegetable oils and FAMEs. Thereby, different FAMEs showed LC50 values ranging from 42.32 to 196.27mg/L against C. quinquefasciatus larvae. The methyl ester obtained from sunflower oil showed the lowest LC50. Histology of C. quinquefasciatus larvae exposed to LC50 of FAMEs was performed and changes in the midgut and fat body morphology were identified. Therefore, larval mortality and changes in the internal organs suggested that FAMEs might be a promising new class of larvicidalcompounds. Cytotoxicity of FAMEs compounds was assessed with the HeLa human cell line and no effect was observed.

Larvicidal activity of essential oil of Peumus boldus Molina and its ascaridole-enriched fraction against Culex quinquefasciatus.

Culex quinquefasciatus (Say, 1823), known as the domestic mosquito, is a common and abundant species throughout the world, and a cosmopolitan species. The adults of this mosquito are important in terms of public and animal health since they display adaptability to different hosts. In humans, they are responsible for the transmission of various diseases. One manner of control of this vector is the use of insecticidal or larvicidal products, which may have the drawback of toxicity to mammals and can be harmful to the environment. The present work evaluated the larvicidal potential of the essential oil (EO) and ascaridole-enriched fraction (EF4-5) obtained from the leaves of Peumus boldus Molina (boldo). The EO, obtained by steam distillation, was analyzed by GC/MS and fractionated on silica gel. EO and EF4-5, containing 31.4% and 89.5% ascaridole, respectively, were evaluated against C. quinquefasciatus at concentrations of 100, 50 and 25 µg/ml on the third and fourth instars. They showed lethal concentrations (LC50) of 82.14 and 41.85 µg/ml, respectively. Larvae treated with the EF4-5 showed morphological changes in the midgut, with cells possessing a cytoplasm that contained small vacuole-like structures, as well as a nucleus with decondensed chromatin and a cell apex with a short brush border. The cells of the fat body showed larger protein granules, which were acidophilic relative to the larvae of the control group. Moreover, the enriched fraction at a dose of 50 µg/ml showed a residual larvicidal effect according to exposure time on C. quinquefasciatus. This residual effect deserves consideration, since a long-term larvicidal product may be a useful tool for vector control.

Larvicidal activity of oils, fatty acids, and methyl esters from ripe and unripe fruit of Solanum lycocarpum (Solanaceae) against the vector Culex quinquefasciatus (Diptera: Culicidae).

INTRODUCTION: The larvicidal activity of oils, fatty acids, and methyl esters of Solanum lycocarpum fruit against Culex quinquefasciatus is unknown. METHODS: The larvicidal activity of samples of ripe and unripe fruit from S. lycocarpum was evaluated against third and fourth instar larvae of C. quinquefasciatus . RESULTS: The oils, fatty acids, and methyl esters of S. lycocarpum showed the greatest larvicidal effect (57.1-95.0%) at a concentration of 100mg/L (LC 50values between 0.70 and 27.54mg/L). CONCLUSIONS: Solanum lycocarpum fruit may be a good source of new natural products with larvicidal activity.

Dengue outbreaks in Divinopolis, south-eastern Brazil and the geographic and climatic distribution of Aedes albopictus and Aedes aegypti in 2011-2012.

OBJECTIVE: To entomologically monitor Aedes spp. and correlate the presence of these vectors with the recent epidemic of dengue in Divinopolis, Minas Gerais State, Brazil. METHODS: Ovitraps were installed at 44 points in the city, covering six urban areas, from May 2011 to May 2012. After collection, the eggs were incubated until hatching. In the 4th stage of development, the larvae were classified as Ae. aegypti or Ae. albopictus. RESULTS: In total, 25 633 Aedes spp. eggs were collected. February was the month with the highest incidence, with 5635 eggs collected and a hatching rate of 46.7%. Ae. aegypti eggs had the highest hatching rate, at 72.3%, whereas Ae. albopictus eggs had 27.7%. Climate and population density influenced the number of eggs found. Indicators of vector presence were positively correlated with the occurrence of dengue cases. CONCLUSION: These data reinforce the need for entomological studies, highlight the relevance of Ae. albopictus as a possible disease vector and demonstrate its adaptation. Ae. albopictus, most commonly found in forested areas, comprised a substantial proportion of the urban mosquito population.

Larvicidal activity of the methanol extract and fractions of the green fruits of Solanum lycocarpum (Solanaceae) against the vector Culex quinquefasciatus (Diptera: Culicidae).

INTRODUCTION: The larvicidal activity of Solanum lycocarpum against Culex quinquefasciatus is unknown. METHODS: We evaluated the larvicidal activity of extracts of the green fruits of Solanum lycocarpum against third and fourth instar larvae of C. quinquefasciatus. RESULTS: Dichloromethane and ethyl acetate fractions showed the greatest larvicidal effect at 200mg/L (83.3% and 86.7%, respectively). The methanol and dichloromethane, ethyl acetate, and hydromethanolic fractions demonstrated larvicidal effects against C. quinquefasciatus, with LC50 values of 126.24, 75.13, 83.15, and 207.05 mg/L, respectively. CONCLUSIONS: Thus, when considering new drugs with larvicidal activity from natural products, S. lycocarpum fruits may be good candidate sources.

Effects of benzodiazepine and ivermectin on Girardiatigrina (Platyhelminthes: Turbellaria) / Efeitos de benzodiazepínico e ivermectina em Girardia tigrina (Platyhelminthes: turbellaria)

Four hundred specimens of Girardia (=Dugesia) tigrina separated in groups of 5 were subjected to concentrations of 10, 25 and 50 ppm diazepam (DZP) and 1, 5 and 10 ppm of ivermectin (IVM), substances with actions associated with ionophore chlorine and GABA receptors in neuromuscular endings in various animals. The period of turbellarians exposure at different concentrations used was 3 h, for DZP while those to IVM were 30, 60, 120 minutes, 12 and 24 h for each tested concentration. In groups exposed to DZP hyperkinetic and type "C" and screw like movements were observed in all concentrations. However, at concentrations of 50 and 25 ppm, the pharynx was protruded in 100% of the specimens followed by detachment at concentration of 50 ppm and 60% at a concentration of 25 ppm. Mortality reached 100% 24 hours after exposure to a concentration of 50 ppm and 30% of planarians became degenerated at a concentration of 25 ppm. Planarians submitted to concentrations of 10 and 25 ppm remained alive and without hyperkinetic movements 48 hours after exposure. In IVM exposed groups, hyperkinetic and contractile movements were observed as well as type "C" screw like movements in all tested concentrations and time dependent. The group exposed for 24 hours showed a time dependent variation in mortality from 0 to 100%. The turbellarians were alive for 48 hours in 1 ppm during 12 hours of exposure but with 24 hours of exposure the mortality reaches 20%. The data indicate a time and concentration dependent relationship in the mode of action of these drugs.

Quatrocentos exemplares de Girardia (=Dugesia) tigrina separados em grupos de 5 foram submetidos a concentrações de 10, 25 e 50ppm de diazepam (DZP) e 1, 5 e 10ppm de ivermectina (IVM), substâncias com ações associadas ao ionóforo cloro e receptores GABA em terminações neuromusculares em vários animais. O tempo de exposição dos turbelários às diferentes concentrações utilizadas de DZP foi de 3 horas, enquanto para cada concentração de IVM os exemplares permaneceram expostos durante 30, 60, 120 minutos, 12 e 24 horas. Nos grupos expostos ao DZP foram verificados movimentos hipercinéticos tipo parafuso e tipo "C" e exposição da faringe ao exterior em todas as concentrações. Houve perda da faringe em 90% dos espécimes na concentração de 50ppm e 60% em 25ppm. Foi verificado também 100% de mortalidade após 24 horas em concentração de 50ppm e 30% de degeneração dos platyhelminthes na concentração de 25ppm. No exame realizado 48 horas após a exposição, as planárias submetidas às concentrações de 25 e 10 ppm permaneceram vivas e não mais apresentaram movimentos hipercinéticos. Nos grupos expostos à IVM, foram observados movimentos contráteis e movimentos hipercinéticos tipo parafuso e tipo "C" em todas as concentrações e foi dependente do tempo de exposição. O percentual de mortalidade variou de 0 a 100 nas primeiras 24 horas de observação também dependente do tempo de exposição. Em concentração de 1 ppm e até 12 horas de exposição não foi observada mortalidade dos animais até 48 horas após. Entretanto com 24 horas de exposição e exame 24 horas após a mortalidade chega a 20% dos exemplares persistindo até o final do período de observação. Os dados indicam uma relação dependente da concentração e do tempo no modo de ação destas substâncias.

Brine shrimp lethality test as a biological model for preliminary selection of pediculicidal components from natural source / Teste de toxicidade frente artemia como modelo preliminar de busca de substâncias pediculicidas a partir de fontes naturais

Brine shrimp lethality test as a biological model for the preliminary selection of pediculicidal components from a natural source. In order to achieve a good correlation between pediculicidal activity and brine shrimp lethality (BSL) test, several pediculicidal substances and active essential oils were tested in BSL test, with the purpose to use the latter as convenient preliminary protocol for pediculicidal activity. Benzyl benzoate, deltametrine and essential oil of Eucalyptus were purchased and clove essential oil obtained by hydrodistillation, besides essential oils, chloroform extracts from Duguetia furfuracea were also submitted to BSL test. All of them were carried out with same protocol described to pediculicidal assay found in the literature, i.e, flask tests were examined every five minutes in the first half hour and then every ten minutes until all the naupli were dead or no movements were observed (knockdown). During the BSL test, it was possible to observe the effect of a particular lethal dose or only a knockdown in the arthropod, as occurred in the test with lice. The results of the BSL test for essential oils and other active substances are essentially in agreement with those described in literature for pediculicidal activity. Extracts and essential oil obtained from aerial parts of D. furfuracea did not present activity, but the essential oil from underground stem bark was active. α-asarone has already been isolated from the underground stem and it has been previously described to possess insecticidal activity.

O teste de toxicidade frente a Artemia salina (TAS) foi utilizado como modelo biológico preliminar na busca de substâncias potencialmente pediculicidas a partir de fontes naturais. Foi encontrada uma boa correlação entre a atividade pediculicida e o TAS, várias substâncias e óleos essenciais descritos como pediculicidas foram testados sobre o microcrustáceo, com o objetivo de se obter um protocolo preliminar e apropriado para detectar aquela atividade. Benzoato de benzila, deltametrina, e óleos essenciais de eucalipto e cravo foram obtidos comercialmente e/ou por extração. Além desses, extratos e óleos essenciais de Duguetia furfuracea também foram testados frente a Artemia. Todas as amostras foram conduzidas utilizando o mesmo protocolo proposto para atividade pediculicida, ou seja, os frascos testes foram lidos a cada cinco minutos na primeira meia hora e depois a cada dez minutos até os nauplios estarem mortos ou sem movimento (knockdown). Durante o TAS foi possível observar a dose letal ou somente o knockdown na larva, como ocorrido no teste com piolho. Os resultados do TAS para os óleos essencial e os demais compostos ativos estão essencialmente de acordo com o descrito na literatura para a atividade pediculicida. Extratos das sementes e óleos essenciais das folhas e sementes D. furfuracea não apresentaram atividade, mas o óleo das cascas do caule subterrâneo foi ativo α-asarona foi isolado das cascas do caule e esta substância apresenta atividade inseticida.

[Susceptibility of Culex quinquefasciatus larvae to different insecticides]. / Suscetibilidade de larvas de Culex quinquefasciatus a diferentes inseticidas.

INTRODUCTION: This study aimed to assess the susceptibility of Culex quinquefasciatus larvae to two pyrethroids (Cypermethrin and Deltamethrin), two derivatives of Avermectin (Ivermectin and Abamectin) and an organophosphate (Temephos). METHODS: Third- and fourth-instar larvae of C. quinquefasciatus were exposed to different concentrations of insecticides (eleven repetitions) according to the World Health Organization's protocol. One hour after exposure, larvae were washed in dechlorinated tap water, transferred to plastic containers containing water without chlorine, fed and observed for periods of 24h until reaching adulthood. To determine lethal concentrations, the obtained values were submitted to regression analysis using the probit model with the Minitab 15 program. RESULTS: The highest concentration used for the LC50 from probit analysis was for Temephos. The evaluated insecticides caused more pronounced larvae mortality in the first 24h with the exception of those exposed to ivermectin. CONCLUSIONS: This study demonstrates that larvae are susceptible to all tested insecticides and that there is a need for monitoring the use of insecticides.

Suscetibilidade de larvas de Culex quinquefasciatus a diferentes inseticidas / Susceptibility of Culex quinquefasciatus larvae to different insecticides

INTRODUÇÃO: O presente estudo teve como objetivo avaliar a suscetibilidade de larvas de Culex quinquefasciatus a dois piretróides (Cipermetrina e Deltametrina), dois derivados da Avermectina (ivermectina e abamectina) e a um organofosforado (Temefós). MÉTODOS: Larvas de 3º e 4º instares de C. quinquefasciatus foram expostas a diferentes concentrações destes (onze repetições) seguindo o protocolo da Organização Mundial de Saúde. Uma hora após a exposição, as larvas foram lavadas em água desclorada, transferidas para recipientes plásticos contendo água sem cloro, alimentadas e observadas por períodos de 24h, até se transformarem em adultos. Para a determinação das concentrações letais, os valores foram submetidos à análise de regressão usando o modelo probit pelo programa Minitab 15. RESULTADOS: Diferenças entre as estimativas da CL50 e CL90 justificaram que a população de mosquitos testada apresenta heterogeneidade em resposta aos inseticidas, sendo a maior concentração utilizada para a CL50, a partir da análise de probit para o Temefós. Todos os inseticidas avaliados causaram mortalidade mais acentuada nas primeiras 24h exceto quando expostas à ivermectina. CONCLUSÕES: As larvas são suscetíveis a todos os inseticidas testados e há uma necessidade de um monitoramento dos inseticidas utilizados.

INTRODUCTION: This study aimed to assess the susceptibility of Culex quinquefasciatus larvae to two pyrethroids (Cypermethrin and Deltamethrin), two derivatives of Avermectin (Ivermectin and Abamectin) and an organophosphate (Temephos). METHODS: Third- and fourth-instar larvae of C. quinquefasciatus were exposed to different concentrations of insecticides (eleven repetitions) according to the World Health Organization's protocol. One hour after exposure, larvae were washed in dechlorinated tap water, transferred to plastic containers containing water without chlorine, fed and observed for periods of 24h until reaching adulthood. To determine lethal concentrations, the obtained values were submitted to regression analysis using the probit model with the Minitab 15 program. RESULTS: The highest concentration used for the LC50 from probit analysis was for Temephos. The evaluated insecticides caused more pronounced larvae mortality in the first 24h with the exception of those exposed to ivermectin. CONCLUSIONS: This study demonstrates that larvae are susceptible to all tested insecticides and that there is a need for monitoring the use of insecticides.

Alterations in the fat body and midgut of Culex quinquefasciatus larvae following exposure to different insecticides.

This study describes morphological alterations in the fat body and midgut of Culex quinquefasciatus larvae following exposure to different insecticides. To this end, both third and fourth instars of C. quinquefasciatus larvae were exposed for 30 and 60 min to organophosphate (50 ppb), pyrethroids (20 and 30 ppb), and avermectin derivates (1.5 and 54 ppb). Following incubation, pH measurements of the larvae gut were recorded. The fat body and midgut were also analyzed by light and transmission electron microscopy. These studies demonstrate a decrease in the pH of the larvae anterior midgut following exposure to all of the tested insecticides. Histochemical tests revealed a strong reaction for neutral lipids in the control group and a marked decrease in the group exposed to cypermethrin. Furthermore, a weak reaction with acidic lipids in larvae exposed to deltamethrin, temephos, ivermectin and abamectin was also observed. Insecticide-exposed larvae also exhibited cytoplasm granule differences, relative to control larvae. Finally, we noted a small reduction in microvilli size in the apex of digestive cells, although vesicles were found to be present. The destructive changes in the larvae were very similar regardless of the type of insecticide analyzed. These data suggest that alterations in the fat body and midgut are a common response to cellular intoxication.

Effect of ivermectin on the life cycle and larval fat body of Culex quinquefasciatus

O presente trabalho verificou o comportamento, sobrevivência e o efeito letal de 1,5 ppb de ivermectina em larvas de Culex quinquefasciatus. As possíveis alterações morfológicas no corpo gorduroso das larvas, no número médio de ovos por postura da fêmea e a duração do período larval foram avaliadas. Na sua execução foram utilizadas 601 larvas de 3o e 4o ínstar do mosquito. Após a postura, os ovos foram separados e as larvas eclodidas contadas. Algumas larvas submetidas à solução 1,5 ppb de ivermectina, foram utilizadas para o preparo de amostras para estudos histológicos. Os resultados mostraram que a ivermectina na concentração de 1,5 ppb causou paralisia nas larvas com 73,38% de mortalidade, mobilização das substâncias estocadas no corpo gorduroso larval e uma diminuição do número de posturas.

Effects of ivermectin on Culex quinquefasciatus larvae

Os efeitos da ivermectina, uma droga semi-sintetica amplamente utilizada para o tratamento de doencas parasitarias do gado, foram observados em larvas de Culex quinquefasciatus. Os efeitos toxicos e a avaliacao da mortalidade foram investigados apos 5,15 30 e 60 minutos de exposicao a 1, 5 ou 10 ppm de solucao de ivermectina. As observacoes foram realizadas 24 e 48 horas apos o inicio do experimento, e perda de mobilidade, paralisia progressiva e alta mortalidade de larvas foram registradas. Os efeitos da ivermectina observados nas larvas de mosquito estao provavelmente correlacionados com a ativacao de canais de Cloro em membranas celulares