Larvicide and oviposition deterrent effects of fruit and leaf extracts from Melia azedarach L. on Aedes aegypti (L.) (Diptera: Culicidae)

Aedes aegypti (L.) (Diptera: Culicidae), the main urban vector of dengue, has developed resistance to various insecticides, making its control increasingly difficult. We explored the effects of Argentine Melia azedarach L. (Meliaceae) fruit and senescent leaf extracts on Ae. aegypti larval development and survival, by rearing cohorts of first instar mosquitoes in water with different extract concentrations. We also analysed oviposition deterrent activity in choice tests with extract-treated ovitraps. The leaf extract showed a strong larvicide activity, with all larvae dying before pupation, and significantly delayed development time. It strongly inhibited oviposition by Ae. aegypti females. The fruit extract showed much weaker effects. This first report of highly effective larvicidal, growth regulating and oviposition deterrent activity of a senescent leaf extract of M. azedarach against Ae. aegypti, suggests that such extract could represent a promising tool in the management of this mosquito pest.     

Sterculia guttata seeds extractives--an effective mosquito larvicide

The larvicidal activity of ethanol, chloroform and hexane soxhlet extracts obtained from S. guttata seeds was investigated against the IVth instar larvae of Dengue fever vector, Aedes aegypti and filarial vector, Culex quinquefasciatus. All extracts including fractions of ethanol extract exhibited 100% larval kill within 24 hr exposure period at 500 ppm concentration. Fraction A1 of ethanol was found to be most promising; its LC50 was 21.552 and 35.520 ppm against C. quinquefasciatus and A. aegypti respectively. Naturally occurring S. guttata seed derived fractions merit further study as potential mosquito larval control agents or lead compounds.     

Is there a relationship between N‐acetyl‐β‐d‐glucosaminidase activity of Metarhizium anisopliae (Metschn.) Sorokin (Hyphomycetes) isolates from peridomestic areas in Central Brazil and larvicidal effect on Aedes aegypti (L.) (Diptera,Culicidae)?

Abstract:  The larvicidal effect of Metarhizium anisopliae (Metschn.) Sorokin (Hyphomycetes) isolated in peridomestic areas in Central Brazil was tested in Aedes aegypti (L.) (Dipt., Culicidae), which is worldwide the primary vector for the viruses that cause human dengue and yellow fever. Highest susceptibility of larvae was found after application of suspended ungerminated conidia. However, conidia, which were found on the larval cuticle and in the gut did not germinate in live or dead larvae. Mortality dropped when testing germinating conidia or supernatants, which originated from chitin-amended minimal medium (MM) inoculated conidia and cultures up to 72 h. Paralysis of larval movement was observed a few hours after application, especially of untreated conidia. Isolates showed a high variability of total protein production and N -acetyl- β - d -glucosaminidase activity after 48 and 72 h incubation in MM. No relationship between enzyme levels and insecticidal activity could be detected. The results indicate that toxic compounds emitted by ungerminated conidia on the cuticle or in the gut are involved in the activity of M. anisopliae against A. aegypti larvae.     

Larvicidal activity of the essential oil from Lippia sidoides Cham. Against Aedes aegypti linn

The aim of this work was to study the larvicidal activity of Lippia sidoides essential oil against Aedes aegypti larvae. The essential oil and its hydrolate (saturated solution of essential oil in water) were obtained by vapor extraction and their chemical composition determined by GL-chromatography coupled to mass spectroscopy. Bioassays were run with the essential oil, pure and diluted hydrolate and with their main constituents thymol and carvacrol. The results obtained showed that L. sidoides essential oil and its hydrolate have larvicidal action against the mosquito A. aegypti, causing an almost instantaneous mortality. Thymol, an alkylated phenol derivative and one of the major components of L. sidoides essential oil, was identified as the active principle responsible for the larvicidal action, causing 100% larval mortality at the lowest tested concentration of 0.017% (w/v). These results suggest that the essential oil of L. sidoides is promising as larvicide against A. aegypti and could be useful in the search of newer, more selective, and biodegradable larvicidal natural compounds to be used in official combat programs and at home.     

Larvicidal and chemosterilant activity of the acetone fraction of petroleum ether extract from Argemone mexicana L seed

The acetone fraction of the petroleum ether extract of seeds from Argemone mexicana L. exhibited larvicidal and growth inhibiting activity against the second instar larvae of Aedes aegypti (Linn). This activity occurred at higher concentrations (200, 100, 50 and 25 ppm). Chemosterilant activity, including reduction in blood meal utilization (27.70%), reduction in fecundity (19.00%), formation of larval-pupal intermediates, formation of pupal-adult intermediates, adult mortality and sterility of first generation eggs (100%), occurred at low concentration (10 ppm).     

Two unusual rotenoid derivatives, 7a-O-methyl-12a-hydroxydeguelol and spiro-13-homo-13-oxaelliptone, from the seeds of Derris trifoliata

The crude methanol extract of the seeds of Derris trifoliata showed potent and dose dependent larvicidal activity against the 2nd instar larvae of Aedes aegypti. From this extract two unusual rotenoid derivatives, a rotenoloid (named 7a-O-methyl-12a-hydroxydeguelol) and a spirohomooxarotenoid (named spiro-13-homo-13-oxaelliptone), were isolated and characterised. In addition a rare natural chromanone (6,7-dimethoxy-4-chromanone) and the known rotenoids rotenone, tephrosin and dehydrodeguelin were identified. The structures were assigned on the basis of spectroscopic evidence. The larvicidal activity of the crude extract is mainly due to rotenone.     

Mosquito larvicidal activity of aqueous extracts of long pepper (Piper retrofractum vahl) from Thailand.

Aqueous extracts of nine medicinal plants were bioassayed against larvae of Culex quinquefasciatus Say and Aedes aegypt (L.). Among these plants, the long pepper, Piper retrofractum Vahl (Piperaceae), showed the highest level of activity against mosquito larvae. To gain more information on larvicidal activity of P. retrofractum, fresh fruits of this plant were extracted in water and the extracts made into powder and bioassayed against 3rd and 4th instar larvae of Cx. quinquefasciatus and Ae. aegypti in the laboratory. Extracts of unripe (001/3) and ripe (002/3 and 001/4) fruits showed different levels of activity against Cx. quinquefasciatus larvae. Extracts 001/3 and 002/3 were equi-toxic to a Bacillus sphaericus resistant and susceptible strains, both from Thailand. The ripe fruit extract 002/3 was somewhat more active against Ae. aegypti than Cx. quinquefasciatus. Another ripe fruit extract (001/4) was much more toxic to both mosquito species. Diluted solutions of the solid extract (002/3) in distilled water lost their larvicidal activity upon aging. Loss of activity at 25 degrees C was greater than that stored at 4 degrees C, and greater in water than in acetone solution.     

Derris (Lonchocarpus) urucu (Leguminosae) extract modifies the peritrophic matrix structure of Aedes aegypti (Diptera:Culicidae)

Aqueous suspension of ethanol extracts of Derris (Lonchocarpus) urucu (Leguminosae), collected in the state of Amazonas, Brazil, were tested for larvicidal activity against the mosquito Aedes aegypti (Diptera:Culicidae). The aim of this study was to observe the alterations of peritrophic matrix in Ae. aegypti larvae treated with an aqueous suspension of D. urucu extract. Different concentrations of D. urucu root extract were tested against fourth instar larvae. One hundred percent mortality was observed at 150 microg/ml (LC(50) 17.6 microg/ml) 24 h following treatment. In response to D. urucu feeding, larvae excreted a large amount of amorphous feces, while control larvae did not produce feces during the assay period. Ultrastructural studies showed tha larvae fed with 150 microg/ml of D. urucu extract for 4 h have an imperfect peritrophic matrix and extensive damage of the midgut epithelium. Data indicate a protective role for the peritrophic matrix. The structural modification of the peritrophic matrix is intrinsically associated with larval mortality.     

市售长叶松油和锡兰肉桂油对登革热媒蚊埃及伊蚊的灭幼效果(英文)

松油和桂皮油由于具有芳香性气味, 因而成为良好的成虫驱避剂, 但是关于它们对蚊虫的杀幼虫作用研究不多。为揭示市售的长叶松Pinus longifolia油和锡兰肉桂Cinnamomum zeylanicum油对来源于印度德里的埃及伊蚊Aedes aegypti 4龄幼虫的毒杀潜力, 我们进行了室内研究, 以幼虫死亡率及行为改变和形态改变等指标评估其杀幼虫潜力。结果表明： 两种油对埃及伊蚊4龄幼虫均具有毒杀作用, 且松油的灭幼效果优于桂皮油。松油的LC₅₀和LC₇₀ 值分别为0.33093 mg/L 和0.54476 mg/L, 而桂皮油的LC₅₀和LC₇₀ 值分别为0.63159 mg/L和0.77736。进一步观察发现, LC₉₀剂量下桂皮油的杀幼虫潜力强于松油, 其LC₉₀为1.11879 mg/L, 而松油的LC₉₀为 1.04915 mg/L。在处理的幼虫中观察到行为改变, 如兴奋、 坐立不安、 颤抖、 痉挛然后瘫痪, 说明这两种油可能对其神经肌肉系统产生了影响。显微观察处理幼虫的形态改变发现, 与对照相比, 大多数器官的外观正常, 只是肛鳃略为内收缩而引起结构畸形, 提示肛腮可能是这两种油的作用位点, 腮的功能异常引起了幼虫死亡。这两种油品可开发用作防治蚊虫的新型杀幼虫药剂。     

Biochemical and cytoimmunological evidence for the control of Aedes aegypti larval trypsin with Aea-TMOF

Trypsin and chymotrypsin-like enzymes were detected in the gut of Aedes aegypti in the four larval instar and pupal developmental stages. Although overall the amount of trypsin synthesized in the larval gut was 2-fold higher than chymotrypsin, both enzymes are important in food digestion. Feeding Aea-Trypsin Modulating Oostatic Factor (TMOF) to Ae. aegypti and Culex quinquefasciatus larvae inhibited trypsin biosynthesis in the larval gut, stunted larval growth and development, and caused mortality. Aea-TMOF induced mortality in Ae. aegypti, Cx. quinquefasciatus, Culex nigripalpus, Anopheles quadrimaculatus, and Aedes taeniorhynchus larvae, indicating that many mosquito species have a TMOF-like hormone. The differences in potency of TMOF on different mosquito species suggest that analogues in other species are similar but may differ in amino acid sequence or are transported differently through the gut. Feeding of 29 different Aea-TMOF analogues to mosquito larvae indicated that full biological activity of the hormone is achieved with the tetrapeptide YDPA. Using cytoimmunochemical analysis, intrinsic TMOF was localized to ganglia of the central nervous system in larvae and male and female Ae. aegypti adults. The subesophageal, thoracic, and abdominal ganglia of both larval and adult mosquitoes contained immunoreactive cells. Immunoreactive cells were absent in the corpus cardiacum of newly molted 4th instar larvae but were found in late 4th instar larvae. In both males and females, the intrinsic neurosecretory cells of the corpus cardiacum were filled with densely stained immunoreactive material. These results indicate that TMOF-immunoreactive material is synthesized in sugar-fed male and female adults and larvae by the central nervous system cells.     

Larvicidal activity of tectoquinone isolated from red heartwood-type Cryptomeria japonica against two mosquito species

Mosquito larvicidal activities of methanolic extracts from different plant parts of red heartwood-type Cryptomeria japonica D. Don against the fourth-instar larvae of Aedes aegypti and Aedes albopictus were examined. Results of mosquito larvicidal tests demonstrated that the n-hexane fraction of C. japonica sapwood methanolic extract had an excellent inhibitory effect against the larvae of A. aegypti and A. albopictus and its LC50 values were 2.4 and 3.3 microg/ml, respectively, in 24h. Following the bioactivity-guided fractionation procedure, the active constituent isolated from C. japonica sapwood was characterized as tectoquinone by spectroscopic analyses. The LC50 values of tectoquinone against A. aegypti and A. albopictus in 24h were 3.3 and 5.4 microg/ml, respectively. In addition, comparisons of mosquito larvicidal activity of anthraquinone congeners demonstrated that anthraquinone skeleton with a methyl group at C-2 position, such as tectoquinone, exhibited the strongest mosquito larvicidal activity. Results of this study show that the methanolic extract of C. japonica sapwood may be considered as a potent source and tectoquinone as a new natural mosquito larvicidal agent.     

Larvicidal activity of saponins from Balanites aegyptiaca callus against Aedes aegypti mosquito

Seeking an alternative approach for producing a larvicidal product from Balanites aegyptiaca plants, callus was produced from in vitro cultures of root explants and its larvicidal activity against Aedes aegypti mosquito larvae was evaluated. Concentrations of 0, 50, 100, 500, 1000, and 1500 ppm of saponins from the root-derived callus of B. aegyptiaca were used to determine larvicidal effects and consequent effect on adult emergence. A dose-dependent effect was observed. In a chronic mortality assessment (after 7 days of exposure), concentrations of 500 ppm or greater killed 100% of the test larvae population. Fifty parts per million showed no difference in larval mortality compared to the control (0 ppm); however, this concentration allowed one-fourth of the adult emergence of the control treatment. These results suggest that saponins from in vitro cultures of the root explant of B. aegyptiaca can be used as a larvicidal agent against A. aegypti larvae.     

Biocontrol evaluation of extracts and a major component,clusianone, from Clusia fluminensis Planch. & Triana against Aedes aegypti

Studies evaluated the effects of hexanic extracts from the fruits and flowers ofClusia fluminensis and the main component of the flower extract, a purified benzophenone (clusianone), against Aedes aegypti. The treatment of larvae with the crude fruit or flower extracts from C. fluminensis did not affect the survival ofAe. aegypti (50 mg/L), however, the flower extracts significantly delayed development of Ae. aegypti. In contrast, the clusianone (50 mg/L) isolate from the flower extract, representing 54.85% of this sample composition, showed a highly significant inhibition of survival, killing 93.3% of the larvae and completely blocking development of Ae. aegypti. The results showed, for the first time, high activity of clusianone against Ae. aegypti that both killed and inhibited mosquito development. Therefore, clusianone has potential for development as a biopesticide for controlling insect vectors of tropical diseases. Future work will elucidate the mode of action of clusianone isolated from C. fluminensis.     

Cytotoxicity of a cloned Bacillus thuringiensis subsp. israelensis CryIVB toxin to an Aedes aegypti cell line

A cloned CryIVB toxin was purified from a cured strain of Bacillus thuringiensis (BT) containing the cryIVB gene on the recombinant plasmid Cam135. Solubilized protoxin was treated with Aedes gut extract or trypsin for varying times and tested for toxicity in vitro on three dipteran and one lepidopteran cell line. Treatment with the Aedes extract but not trypsin, produced an active toxin which lysed only Aedes aegypti cells out of those tested. This activation was time-dependent reaching a maximum after 6 h. Both the Aedes extract-treated and trypsin-treated toxin killed A. aegypti larvae, but this toxicity declined rapidly with increasing time of exposure to the proteolytic preparations.     

Meliternatin: a feeding deterrent and larvicidal polyoxygenated flavone from Melicope subunifoliolata

We screened more than 60 Malaysian plants against two species of insects and found that Melicope subunifoliolata (Stapf) T.G. Hartley (Rutaceae) showed strong feeding deterrent activity against Sitophilus zeamais Motsch. (Curculionidae) and very good larvicidal activity against Aedes aegypti L. (Diptera). One anti-insect compound, meliternatin (3,5-dimethoxy-3',4',6,7-bismethylendioxyflavone) (6) and six other minor polyoxygenated flavones were isolated from M. subunifoliolata.     

Digestion of Bacillus thuringiensis var. israelensis spores by larvae of Aedes aegypti.

The larvicidal activity of Bacillus thuringiensis var. israelensis against mosquitoes and the blackfly is included in parasporal crystalline bodies which are produced during sporulation. Following ingestion, the crystals are solubilized in the larval midgut and induce death within a short time; the spores germinate in the dead larvae and complete a growth cycle. The fate of the spores in surviving live larvae was elucidated by using a nonlarvicidal B. thuringiensis var. israelensis mutant. When introduced as the only food source, spores of this mutant support development to the adult stage of newly hatched Aedes aegypti larvae at a rate directly related to spore concentration. The conclusion that spores of B. thuringiensis var. israelensis are digested in the larval gut was substantiated by following the incorporation of [35S]methionine-labeled spores into larval tissues.     

Specificity of Bacillus thuringiensis var. colmeri insecticidal delta-endotoxin is determined by differential proteolytic processing of the protoxin by larval gut proteases

The native crystal delta-endotoxin produced by Bacillus thuringiensis var. colmeri, serotype 21, is toxic to both lepidopteran (Pieris brassicae) and dipteran (Aedes aegypti) larvae. Solubilization of the crystal delta-endotoxin in alkaline reducing conditions and activation with trypsin and gut extracts from susceptible insects yielded a preparation whose toxicity could be assayed in vitro against a range of insect cell lines. After activation with Aedes aegypti gut extract the preparation was toxic to all of the mosquito cell lines but only one lepidopteran line (Spodoptera frugiperda), whereas an activated preparation produced by treatment with P. brassicae gut enzymes or trypsin was toxic only to lepidopteran cell lines. These in vitro results were paralleled by the results of in vivo bioassays. Gel electrophoretic analysis of the products of these different activation regimes suggested that a 130-kDa protoxin in the native crystal is converted to a 55-kDa lepidopteran-specific toxin by trypsin or P. brassicae enzymes and to a 52-kDa dipteran toxin by A. aegypti enzymes. Two-step activation of the 130-kDa protoxin by successive treatment with trypsin and A. aegypti enzymes further suggested that the 52-kDa dipteran toxin is derived from the 55-kDa lepidopteran toxin by enzymes specific to the mosquito gut. Confirmation of this suggestion was obtained by peptide mapping of these two polypeptides. The native crystal 130 kDa delta-endotoxin and the two insect-specific toxins all cross-reacted with antiserum to B. thuringiensis var. kurstaki P1 lepidopteran toxin. Preincubation of the two activated colmeri toxins with P1 antiserum neutralized their cytotoxicity to both lepidopteran and dipteran cell lines.     

Comparison of development of Bacillus thuringiensis subsp. israelensis and Bacillus sphaericus in mosquito larvae

Immunofluorescent staining was used with thin sections of paraffin-embedded specimens to detect the development of Bacillus thuringiensis var. israelensis and Bacillus sphaericus in the gut of mosquito larvae. The third- and fourth-instar larvae of Aedes aegypti, Anopheles maculatus, and Culex quinquefasciatus were fed either vegetative cells or spores of the bacteria. Spore germination, multiplication, and sporulation were studied in the larvae of each species. The spores of B. thuringiensis var. israelensis and B. sphaericus strain 2297 could germinate and cells could sporulate in the larval body. The vegetative cells of B. sphaericus strain 810428 were also able to produce spores in the mosquito larval gut, but the germination of spores could not be detected in the larvae. Multiplication of all bacterial species was observed after the larvae died. Growth of the bacteria in distilled water containing crude extracts of larvae made from each species was compared with that in synthetic medium (nutrient broth). They could produce spores and toxins in all the media used and the toxins had larvicidal activity against the target mosquitos Ae. aegypti, An. maculatus, and C. quinquefasciatus.     

Bruchid (Coleoptera: Bruchidae) ovicidal phenylbutanoid from Zingiber purpureum

The larvicidal activity of the dichloromethane extract of Zingiber purpureum Roscoe (Zingiberaceae) rhizome against the second instar of Aedes aegypti (L.) (Diptera: Culicidae) is shown to be due to 4-(3',4'-dimethoxyphenyl)buta-1,3-diene. The diene also showed ovicidal activity against the bruchid Callosobruchus maculatus (F.) (Coleoptera: Bruchidae). Most of the eggs laid by bruchids on treated cowpea seeds were transparent, and very few of them contained developing embryos. The few larvae produced from these embryos were unable to penetrate the seed coat and enter the seed. Similar effects were seen when adults were exposed to the compound and then placed on untreated cowpea seeds, suggesting that a new type of maternally mediated ovicidal effect was involved. Coated and impregnated granular formulations of the extract were evaluated for use in the control of bruchid infestation of stored cowpea seeds. Coated granules showed activity similar to that of the crude extract but were found to lose activity rapidly. Impregnated granules were found to be less active than the crude extract.     

Inhibition of gut proteases and development of dengue vector, Aedes aegypti by Allium sativum protease inhibitor

The paper describes the bio efficacy of a protease inhibitor; isolated from Allium sativum ‘garlic’ (ASPI); against Aedes aegypti mosquito, a well-known transmitter of dengue and Chikungunya. The purification of protease inhibitor from Allium sativum ‘garlic’ (ASPI) was carried out by ammonium sulfate precipitation followed by Fast Protein Liquid Chromatography using akta DEAE-Cellulose column. The protein fraction demonstrating trypsin inhibitory activity was further evaluated for its insecticidal activity using gut protease inhibition assay and larvicidal assay. ASPI is an inhibitor of porcine trypsin (IC₅₀ of 650.726?μg/mL) and has molecular weight of ~15?kDa determined by SDS PAGE similar to other inhibitors of the Kunitz-type family (14–26?kDa). ASPI demonstrated 50% reduced activity of Ae. aegypti midgut proteases and showed a dose-dependent acute toxicity on Ae. aegypti 3rd instars exhibiting LC₅₀ value of ~50.827?μg/mL. After ten days of larval exposure ASPI resulted in a 24-h delay of larval development and ~72% mortality at 61.5?μg/mL. These results suggest that ASPI may serve as potent insecticidal agent and hence opens a new gateway in the field of phyto-remediation.