Insecticidal Activity of Bacillus laterosporus

The Bacillus laterosporus strains 921 and 615 were shown to have toxicity for larvae of the mosquitoes Aedes aegypti, Anopheles stephensi, and Culex pipiens. The larvicidal activity of B. laterosporus was associated with spores and crystalline inclusions. Purified B. laterosporus 615 crystals were highly toxic for Aedes aegypti and Anopheles stephensi.     

Comparison of water ingestion between black flies (Simuliidae) and a mosquito (Aedes aegypti; Culicidae) using radiolabelled glucose

Ingestion of water by larvae of the black flies Simulium verecundum, Prosimulium mixtum/fuscum and Cnephnia dacotensis and by larvae of the mosquito Aedes aegypti was studied with radiolabelled glucose as a tracer. Extremely low ( 200 dpm/3 h) radiolabel uptake occurred in the black flies species, while Aedes aegypti, a species that is known to drink, had high radiolabel counts (> 6000 dpm/3 h). Radiolabel uptake by S. verecundum did not change for larvae fed in absence of current or with plugged mouthparts versus normal larvae. Radiolabel uptake by Aedes aegypti larvae with plugged mouthparts was < 3% of radiolabel uptake in normal larvae. Aedes aegypti was able to ingest the soluble glucose. Simulium verecundum, Prosimulium mixtum/fuscum and Cnephia dacotensis were unable to ingest the soluble sugar. The rationale for this difference is in the different filter-feeding mechanism of Culicidae and Simuliidae.     

Organophosphate Resistance in Aedes aegypti: Study from Dengue Hemorrhagic Fever Endemic Subdistrict in Riau,Indonesia

Background:Dengue hemorrhagic fever (DHF) is a significant health problem. The high number of cases requires preventions, including controlling the dengue vector, Aedes aegypti mosquito. One of the control methods is the use of insecticides containing organophosphate. This study aims to detect organophosphate resistance in Aedes aegypti from DHF endemic subdistrict, Riau, Indonesia by a sensitivity test of temephos and 5% malathion and measuring the activity of non-specific alpha and beta esterase enzymes.Methods:This observational study determined Aedes aegypti resistance from larvae to adult in one DHF endemic subdistrict in Riau, Indonesia. The bioassay was used for temephos sensitivity of Aedes aegypti larvae. The LC99 value was analyzed using probit and compared with the diagnostic value from WHO. The WHO susceptibility test was conducted to determine 5% malathion resistance from adult mosquitoes. The mortality of less than 90% was declared as resistant. Measurement of alpha and beta esterase levels used Lee''s microplate assay technique based on visual identification and absorbance value (AV).Results:The results showed that Aedes aegypti were resistant to temephos. It also showed that adult mosquitoes were resistant to 5% malathion. Based on the alpha esterase activity test, it was found that most of the mosquitoes showed very sensitive meanwhile, based on the beta esterase activity test, most of the mosquitoes were moderate resistance.Conclusion:This study suggests that Aedes aegypti population from DHF endemic subdistrict in Riau, Indonesia are indicated to develop resistance to organophosphate.Key Words: Aedes aegypti, Dengue Hemorrhagic fever, Organophosphate, Resistance     

Purification of the toxic protein from Bacillus thuringiensis serotype 10 isolate demonstrating a preferential larvicidal activity to the mosquito

The protein demonstrating larvicidal activity to the mosquito Aedes aegypti was purified from the alkali extract of the spore-parasporal inclusion complex of the isolate, 73-E-10-2, belonging to Bacillus thuringiensis serotype 10. By Sepharose CL-4B gel filtration and DEAE-cellulose column chromatography, a toxic protein was obtained, and its homogeneity was confirmed by Sephadex G-150 gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of the toxic protein was 67,000, when estimated by SDS-PAGE. The LC₅₀ of the toxic protein against 4-day-old larvae of A. aegypti was 16.8 μg/ml. There was no serological relationship between the toxic protein from the isolate 73-E-10-2 and that (M_r 67,000) from the type strain of B. thuringiensis subsp. israelensis.     

Involvement of male accessory gland substance in the fertility of mosquitoes.

The male accessory gland substance is shown to be involved in the fertility of eggs for the first time in Aedes aegypti and Culex pipiens fatigans. Surgical removal of the paired accessory glands of males did not impair their mating ability. However, the eggs laid by the females after mating with operated males were found to be sterile. This condition could be reversed if the accessory gland substance was later received by the females. Evidence suggests that the accessory gland substance is essential for fertilization.     

Plasmodium gallinaceum: development in Aedes aegypti maintained on various carbohydrate diets

Fructose, galactose, glucose, maltose, melibiose, and trehalose were evaluated for their nutritional and survival values for Aedes aegypti. The development of the exogenous stages of Plasmodium gallinaceum was evaluated by counting and averaging the number of oocysts developing on the midguts of Aedes aegypti.The nutritional and survival value for A. aegypti was greatest on glucose, sucrose, and fructose and lowest on galactose and melibiose. Mosquitoes maintained on fructose produced the greatest numbers of oocysts. Only two other sugars, galactose and melibiose produced more oocysts than their respective controls. Glucose and maltose, both of which had high nutritive and survival value for A. aegypti were less efficient than the control (sucrose) for the development of P. gallinaceum.     

Nationwide Investigation of the Pyrethroid Susceptibility of Mosquito Larvae Collected from Used Tires in Vietnam

Pyrethroid resistance is envisioned to be a major problem for the vector control program since, at present, there are no suitable chemical substitutes for pyrethroids. Cross-resistance to knockdown agents, which are mainly used in mosquito coils and related products as spatial repellents, is the most serious concern. Since cross-resistance is a global phenomenon, we have started to monitor the distribution of mosquito resistance to pyrethroids. The first pilot study was carried out in Vietnam. We periodically drove along the national road from the north end to the Mekong Delta in Vietnam and collected mosquito larvae from used tires. Simplified susceptibility tests were performed using the fourth instar larvae of Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus. Compared with the other species, Ae. aegypti demonstrated the most prominent reduction in susceptibility. For Ae. aegypti, significant increases in the susceptibility indices with a decrease in the latitude of collection points were observed, indicating that the susceptibility of Ae. aegypti against d-allethrin was lower in the southern part, including mountainous areas, as compared to that in the northern part of Vietnam. There was a significant correlation between the susceptibility indices in Ae. aegypti and the sum of annual pyrethroid use for malaria control (1998–2002). This might explain that the use of pyrethroids as residual treatment inside houses and pyrethroid-impregnated bed nets for malaria control is attributable to low pyrethroid susceptibility in Ae. aegypti. Such insecticide treatment appeared to have been intensively administered in the interior and along the periphery of human habitation areas where, incidentally, the breeding and resting sites of Ae. aegypti are located. This might account for the strong selection pressure toward Ae. aegypti and not Ae. albopictus.     

The effect of matrone on oviposition in the mosquito, Aedes Aegypti

Male Aedes aegypti produce a substance made or stored in their accessory glands, which when passed on to females via mating, will prevent subsequent insemination. In addition to becoming refractory with regard to further insemination, mated females also oviposit far more eggs than virgins. The male accessory gland substance, called matrone, has been highly purified and when injected into virgin females, prevents any initial insemination, and will also stimulate oviposition.     

Insecticidal activity of Bacillus laterosporus

Strains of Bacillus laterosporus demonstrated pathogenic activity for second-instar larvae of the mosquito, Culex quinquefasciatus, but failed to demonstrate detectable pathogenicity against larvae of the cabbage looper, Trichoplusia ni. Of 29 strains of the bacterium screened, 16 displayed pathogenicity for mosquito larvae. One of the most pathogenic strains, NRS 590, also demonstrated pathogenic activity for larvae of the mosquito, Aedes aegypti, and for larvae of the black fly, Simulium vittatum. The pathogenicity for Culex larvae was associated with the cell mass rather than with the culture supernatant. A suspension of ultraviolet irradiation-killed cells demonstrated no loss in pathogenic activity, an indication that the pathogenicity is toxin mediated. The toxic substance produced by NRS 590 was found to be resistant to heating at 96°C for 10 min. The toxin was not associated with the heat-resistant, bacterial endospore or with the associated paraspore since a suspension consisting primarily of spores was not toxic to mosquito larvae. Toxic activity in stationary phase cells of NRS 590 was associated with the cell's particulate fraction rather than with the soluble fraction.     

Contribution of S-Layer Proteins to the Mosquitocidal Activity of Lysinibacillus sphaericus

Lysinibacillus sphaericus strains belonging the antigenic group H5a5b produce spores with larvicidal activity against larvae of Culex mosquitoes. C7, a new isolated strain, which presents similar biochemical characteristics and Bin toxins in their spores as the reference strain 2362, was, however, more active against larvae of Culex mosquitoes. The contribution of the surface layer protein (S-layer) to this behaviour was envisaged since this envelope protein has been implicated in the pathogenicity of several bacilli, and we had previously reported its association to spores. Microscopic observation by immunofluorescence detection with anti S-layer antibody in the spores confirms their attachment. S-layers and BinA and BinB toxins formed high molecular weight multimers in spores as shown by SDS-PAGE and western blot detection. Purified S-layer from both L. sphaericus C7 and 2362 strain cultures was by itself toxic against Culex sp larvae, however, that from C7 strain was also toxic against Aedes aegypti. Synergistic effect between purified S-layer and spore-crystal preparations was observed against Culex sp. and Aedes aegypti larvae. This effect was more evident with the C7 strain. In silico analyses of the S-layer sequence suggest the presence of chitin-binding and hemolytic domains. Both biochemical characteristics were detected for both S-layers strains that must justify their contribution to pathogenicity.     

Rôle of male accessory gland substance in the regulation of blood intake by mosquitoes

The presence of male accessory gland substance during mating is shown to increase the blood intake in the adult female Aedes aegypti and Culex pipiens fatigans. Females mated with males whose accessory glands were surgically removed took significantly less blood than females mated with normal males. However, females mated with males whose seminal vesicles were surgically removed took as much blood as the controls.     

Plant species from Brazilian Caatinga: a control alternative for Aedes aegypti

The mosquito Aedes aegypti is the main vector for the virus dengue, chikungunya and Zika. For its control, it is essential to search for natural products with insecticidal effects. The climatic singularity of Caatinga, an exclusive Brazilian biome, aids the survival of plants that produce secondary metabolites, which could be toxic to insects. Therefore, this review discusses the insecticidal potential of Caatinga plants on Aedes aegypti mosquitoes. The meta-analysis was performed using Review Manager Software 5.4.1®. Several studies have demonstrated the insecticidal efficacy of Caatinga plants on the egg, larvae, pupae and adult phases of Ae. aegypti, with a predominance of the plant activity in the larval stage. The leaves were the most utilized part of the plant. The essential oils from Caatinga plants were significantly active against Ae. aegypti (RR = 0.21, 95 % CI = 0.07 – 0.68, p = 0.009). The most promising botanical genera as an insecticide are: Abarema, Myracrodruon, Croton, Lippia and Syagrus. Among chemical compounds from these insecticidal plants has been identified and isolated flavonoids and fatty acids. Therefore, the Caatinga plant is a promising plant that contain bioactive compounds that are useful in the control of vector insects. This could contribute to the characterisation and valorisation of flora of this biome, as well as the production of environmentally friendly insecticides with specific action on target insects.     

Cyt1A from Bacillus thuringiensis Synergizes Activity of Bacillus sphaericus against Aedes aegypti (Diptera: Culicidae)

Bacillus sphaericus is a mosquitocidal bacterium recently developed as a commercial larvicide that is used worldwide to control pestiferous and vector mosquitoes. Whereas B. sphaericus is highly active against larvae of Culex and Anopheles mosquitoes, it is virtually nontoxic to Aedes aegypti, an important vector species. In the present study, we evaluated the capacity of the cytolytic protein Cyt1A from Bacillus thuringiensis subsp. israelensis to enhance the toxicity of B. sphaericus toward A. aegypti. Various combinations of these two materials were evaluated, and all were highly toxic. A ratio of 10:1 of B. sphaericus to Cyt1A was 3,600-fold more toxic to A. aegypti than B. sphaericus alone. Statistical analysis showed this high activity was due to synergism between the Cyt1A toxin and B. sphaericus. These results suggest that Cyt1A could be useful in expanding the host range of B. sphaericus.     

Different Repellents for Aedes aegypti against Blood-Feeding and Oviposition

Methyl N,N-dimethyl anthranilate (MDA), ethyl anthranilate (EA) and butyl anthranilate (BA) were previously shown to repel Aedes aegypti mosquitoes from landing on human skin. However, the effect of these compounds on the orientation of flying mosquitoes in a choice situation and their effect on mosquito oviposition are not yet known. Here, we used a modified Y-tube olfactometer to test the effect of these compounds on the orientation of Aedes aegypti flying towards skin odor (human fingers), and we tested their effect on Aedes aegypti oviposition choice in a cage assay. In both behavioral situations we compared the effect to the well-documented repellent N,N-diethyl-meta-toluamide (DEET). MDA, EA, and DEET inhibited Aedes aegypti from flying towards skin odor while BA had no such effect. Conversely, MDA had no effect on oviposition while EA, BA, and DEET deterred oviposition, with the strongest effect observed for BA. Thus, we confirm that EA and DEET are generally repellent, while MDA is repellent only in a host-seeking context, and BA is deterrent only in an oviposition context. These compounds appear of potential use in mosquito control programs.     

Natural arbovirus infection rate and detectability of indoor female Aedes aegypti from Mérida,Yucatán,Mexico

Arbovirus infection in Aedes aegypti has historically been quantified from a sample of the adult population by pooling collected mosquitoes to increase detectability. However, there is a significant knowledge gap about the magnitude of natural arbovirus infection within areas of active transmission, as well as the sensitivity of detection of such an approach. We used indoor Ae. aegypti sequential sampling with Prokopack aspirators to collect all mosquitoes inside 200 houses with suspected active ABV transmission from the city of Mérida, Mexico, and tested all collected specimens by RT-PCR to quantify: a) the absolute arbovirus infection rate in individually tested Ae. aegypti females; b) the sensitivity of using Prokopack aspirators in detecting ABV-infected mosquitoes; and c) the sensitivity of entomological inoculation rate (EIR) and vectorial capacity (VC), two measures ABV transmission potential, to different estimates of indoor Ae. aegypti abundance. The total number of Ae. aegypti (total catch, the sum of all Ae. aegypti across all collection intervals) as well as the number on the first 10-min of collection (sample, equivalent to a routine adult aspiration session) were calculated. We individually tested by RT-PCR 2,161 Aedes aegypti females and found that 7.7% of them were positive to any ABV. Most infections were CHIKV (77.7%), followed by DENV (11.4%) and ZIKV (9.0%). The distribution of infected Aedes aegypti was overdispersed; 33% houses contributed 81% of the infected mosquitoes. A significant association between ABV infection and Ae. aegypti total catch indoors was found (binomial GLMM, Odds Ratio > 1). A 10-min indoor Prokopack collection led to a low sensitivity of detecting ABV infection (16.3% for detecting infected mosquitoes and 23.4% for detecting infected houses). When averaged across all infested houses, mean EIR ranged between 0.04 and 0.06 infective bites per person per day, and mean VC was 0.6 infectious vectors generated from a population feeding on a single infected host per house/day. Both measures were significantly and positively associated with Ae. aegypti total catch indoors. Our findings provide evidence that the accurate estimation and quantification of arbovirus infection rate and transmission risk is a function of the sampling effort, the local abundance of Aedes aegypti and the intensity of arbovirus circulation.     

Toxicity of Bacillus thuringiensis toward Aedes aegypti larvae.

Among six strains of Bacillus thuringiensis and five other species of Bacillus, only two strains of B. thuringiensis, strains HD-1 and BA-068, were toxic to Aedes aegypti larvae within 24 hr. The LC₅₀s were 5.6 × 10⁴ and 2.4 × 10⁵ spores/ml for strains HD-1 and BA-068, respectively. The toxic factor(s) was heat sensitive and γ ray resistant and preliminary evidences indicated that it was associated with the crystalline body of B. thuringiensis.     

Defense reactions of mosquitoes to filarial worms: comparative studies on the response of three different mosquitoes to inoculated Brugia pahangi and Dirofilaria immitis microfilariae

The melanization response of Aedes trivittatus and the Rockefeller (RKF) and black-eyed Liverpool (LVP) strains of Aedes aegypti against intrathoracically inoculated Dirofilaria immitis and Brugia pahangi microfilariae (mff) was investigated. All mff of either species were melanized in A. trivittatus following Day 2 postinoculation, and the response of this species was significantly more rapid and effective than either strain of A. aegypti. The refractory RKF strain had a significantly greater response against both D. immitis and B. pahangi than the highly susceptible LVP strain, but data suggest that the increased responsiveness was due to a physiologic incompatibility in RKF A. aegypti, thereby resulting in a greater mortality and subsequent melanization of inoculated mff. Inoculation of large numbers of mff overloaded the defense capabilities of A. aegypti (LVP), but not those of A. trivittatus. The melanization response against D. immitis mff was effectively reduced for up to 4 days in A. aegypti (LVP), but for only 1 day in A. trivittatus, when mosquitoes were maintained on a 0.3 m sucrose diet containing from 0.1 to 1.0% (w/v) phenylthiourea.     

Chromosomal differentiation in two species of Aedes and their hybrids revealed by giemsa C-banding

Giemsa C-banding patterns in two species of mosquitoes, Aedes aegypti and Aedes mascarensis, their hybrids and backcross progeny revealed differences in the sex chromosome pair. In A. aegypti, the female determining or the m chromosome in both males and females shows a conspicuous band in the centromere region and another band in one arm. The male determining or the M chromosome is devoid of any bands. Progeny of crosses involving A. aegypti females and A. mascarensis males showed interesting albeit unexpected results. The intercalary band was suppressed in both sons and daughters. When such F₁ sons were backcrossed to A. aegypti females, a proportion of males developed into intersexes. These intersex progeny differed from the normal males in terms of their banding pattern. In the reciprocal cross (A. mascarensis female × A. aegypti male), the F₁ and the backcross progeny yielded the expected C-banding patterns. The implications of the reversible expression of the intercalary band on the A. aegypti m chromosome and its relevance to genetic regulation are discussed.     

Neutral lipids of Aedes aegypti and Aedes albopictus cells cultured in vitro.

Aedes aegypti feeding on chickens infected with Plasmodium gallinaceum take less blood and lay fewer eggs than those feeding on uninfected hosts. Both activities show an inverse correlation with the degree of parasitemia. Mosquitoes feeding on infected chickens ingest blood in amounts directly proportional to the length of time spent on the hosts, whereas there is no relationship between host contact and blood meal size for mosquitoes feeding on uninfected hosts. Feeding and probing choice experiments demonstrate that infected chickens are less attractive to Aedes aegypti than uninfected chickens.     

Production of Cry11A and Cry11Ba Toxins in Bacillus sphaericus Confers Toxicity towards Aedes aegypti and Resistant Culex Populations

Cry11A from Bacillus thuringiensis subsp. israelensis and Cry11Ba from Bacillus thuringiensis subsp. jegathesan were introduced, separately and in combination, into the chromosome of Bacillus sphaericus 2297 by in vivo recombination. Two loci on the B. sphaericus chromosome were chosen as target sites for recombination: the binary toxin locus and the gene encoding the 36-kDa protease that may be responsible for the cleavage of the Mtx protein. Disruption of the protease gene did not increase the larvicidal activity of the recombinant strain against Aedes aegypti and Culex pipiens. Synthesis of the Cry11A and Cry11Ba toxins made the recombinant strains toxic to A. aegypti larvae to which the parental strain was not toxic. The strain containing Cry11Ba was more toxic than strains containing the added Cry11A or both Cry11A and Cry11Ba. The production of the two toxins together with the binary toxin did not significantly increase the toxicity of the recombinant strain to susceptible C. pipiens larvae. However, the production of Cry11A and/or Cry11Ba partially overcame the resistance of C. pipiens SPHAE and Culex quinquefasciatus GeoR to B. sphaericus strain 2297.