Culex Larvae Research Articles

PEGylation Enhances Mosquito-Larvicidal Activity of Lysinibacillus sphaericus Binary Toxin.

Toxic strains of Lysinibacillus sphaericus have been used in the field for larval control of mosquito vector diseases. The high toxicity of L. sphaericus is attributed to the binary (BinAB) toxin produced as parasporal crystalline inclusions during the early stages of sporulation. BinA and BinB, the primary components of these spore-crystals, exert high toxicity when administered together. However, instability, short half-lives, and rapid proteolytic digestion can limit their use as an effective insecticide. BinA alone displays larvicidal toxicity, in the absence of BinB, albeit with much reduced activity. Here for the first time, we demonstrate the beneficial effect of PEGylation (covalent attachment of polyethylene glycol) on mosquito-larvicidal activity of BinA. Polymer conjugation was achieved using 750 Da polyethylene glycol (PEG) at two different pH values (pH 7.2 and 8.5). Two different isoforms of the biopolymers, purified to homogeneity, were highly water-soluble and resistant to trypsin and proteinase K. The mono-PEGylated BinA isoforms also displayed preservation of the toxin structure with improved thermal stability by about 3-5 °C, as evident from thermal denaturation studies by circular dichroism and differential scanning fluorimetry. Notably, PEGylation enhanced BinA toxicity by nearly 6-fold. The PEGylated BinA isoforms alone displayed high larvicidal activity (LC50 value of ∼3.4 ng/mL) against the third instar Culex larvae, which compares favorably against LC50 reported for the combination of BinA and BinB proteins. Since BinA can be synthesized easily through recombinant technology and easily PEGylated, the conjugated biopolymers offer a promising opportunity for mosquito control programs.

Polyethylene Glycol Conjugation Enhances Mosquito-Larvicidal Activity of Lysinibacillus Sphaericus BinA Protein

Mosquitoes are known to spread human diseases like West Nile fever, dengue, malaria, zika etc. accounting for millions of deaths worldwide. Lysinibacillus sphaericus, a gram positive, spore producing commensal soil bacterium, has been used worldwide for controlling mosquito population like Culex and Anopheles, and is regarded as safe against non-target organisms. Binary toxin, composed of BinA (41.9 kDa) and BinB (51.4 kDa) component proteins, is responsible for the high larvicidal activity of several L. sphaericus strains. The two proteins exert high toxicity when administered together. BinA alone displays larvicidal activity, in the absence of BinB, albeit at reduced levels. But instability, shorter half-lives and rapid proteolytic digestion can limit their use as an effective insecticide. We for the first time demonstrate the beneficial effect of PEGylation (covalent attachment of polyethylene glycol) on mosquito-larvicidal activity of BinA protein. PEG-protein conjugates were synthesized using PEG-isocyanate polymer. The resulting bio-conjugates were purified to homogeneity by column chromatography methods. These were characterized by various biophysical methods like MALDI-TOF, DLS, DSF and CD. Two different isoforms of PEG-BinA conjugates are expected from biophysical analysis, which appear to be mono-PEGylated but may differ in the site of PEG attachment to BinA protein. The PEGylated proteins displayed preservation of protein's native structure and exhibited improved thermal stability by about 3-5 oC. The PEGylated proteins were also checked for stability against complex proteolytic environment. Regardless of the site of modification, the two isoforms showed a remarkable 7-fold improvement in the larvicidal activity of BinA protein against 3rd instar Culex larvae, compared to the unmodified protein. The PEGylation of recombinantly produced BinA can be achieved easily. It promises a judicious approach towards mosquito population control.

Biological Control of <I>Culex quinquefasciatus</I> Say (Diptera: Culicidae) Larvae

Local larvivorous fish Aphanius dispar has potential for regulating vector mosquitoes. The consumption of larvae of the mosquito Culex quinquefasciatus Say (Diptera: Culicidae) by Rasbora daniconius, Puntius ticto and Puntius conchonius was evaluated in the presence of different prey densities under simple habitat conditions. The number of mosquito larvae consumed in fresh and ditch water at three seasons was observed. Only third and fourth instars of Culex larvae were used on a particular prey density of 1 to 10 (increase 100). The maximum and minimum value of larvae eating per day in ditch and fresh water were observed to 267 to 876 and 277 to 880 respectively. The comparisons for the pair using Tukey-Kramer HSD indicated the fact that there was non-significant difference. The respective mean values in fresh and ditch water were observed 525 and 493 respectively. The analysis of water (ditch and fresh) was nonsignificant (F = 1.0116). Rasbora daniconius species of fish played a dominating role in eating the Culex larvae in ditch and fresh water.

Biological Control of <I>Culex quinquefasciatus</I> Say (Diptera: Culicidae) Larvae

Local larvivorous fish <em>Aphanius dispar</em> has potential for regulating vector mosquitoes. The consumption of larvae of the mosquito <em>Culex quinquefasciatus</em> Say (Diptera: Culicidae) by <em>Rasbora daniconius, Puntius ticto</em> and <em>Puntius conchonius</em> was evaluated in the presence of different prey densities under simple habitat conditions. The number of mosquito larvae consumed in fresh and ditch water at three seasons was observed. Only third and fourth instars of Culex larvae were used on a particular prey density of 1 to 10 (increase 100). The maximum and minimum value of larvae eating per day in ditch and fresh water were observed to 267 to 876 and 277 to 880 respectively. The comparisons for the pair using Tukey-Kramer HSD indicated the fact that there was non-significant difference. The respective mean values in fresh and ditch water were observed 525 and 493 respectively. The analysis of water (ditch and fresh) was nonsignificant (F = 1.0116). <em>Rasbora daniconius</em> species of fish played a dominating role in eating the <em>Culex</em> larvae in ditch and fresh water.

Evaluation of different formulations of IGRs against Aedes albopictus and Culex quinquefasciatus (Diptera: Culicidae)

ObjectiveTo test the relative efficacy of pyriproxyfen and methoprene on mortality, deformity, inhibition and emergence to adult stages of Culex quinquefasciatus and Aedes albopictus. MethodsSerial dilutions (0.01–0.05 mg/L) of methoprene, pyriproxyfen 0.5 water dispersible granules (WDG) and pyriproxyfen 1.0 WDG were used to assess mortality and inhibition of 3rd instar larvae of Aedes albopictus and Culex quinquefasciatus. Each concentration and control was replicated four times in completely randomized design. Data on larval mortality, growth inhibition, deformities and adult's emergence was recorded weekly. On the basis of best comparative performance, the efficacy of pyriproxyfen 1.0 WDG at 0.1 g/m3 was also tested in the field by collecting treated water samples monthly for 1–6 months after field application. Twenty five 3rd instar larvae of Aedes and Culex spp. of the same cohorts were used for bioassays and compared with larvae in control cups containing 1 L of untreated tap water. ResultsResults revealed variations in fatality of different insect growth regulators (IGRs) to the 3rd instar larvae of Culex and Aedes mosquitoes. Among the IGRs, pyriproxyfen 1.0 WDG was found best that exhibited significantly high emergence inhibition against Culex and Aedes spp. Based on the results, the IGRs were classified in terms of the tested parameters in order of pyriproxyfen 1.0 WDG > pyriproxyfen 0.5 WDG > methoprene. In case of field studies, pyriproxyfen 1.0 WDG, pool data of the entire target treated sites showed minimum adult emergence from water sampled of habitats treated with 0.1 g/m3 of pyriproxyfen 1.0 WDG. ConclusionsIt is thus concluded that IGRs can be utilized as environment friendly control measures for Culex and Aedes spp. of mosquitoes on small and large scale. This will reduce the use of conventional insecticides by the public health authorities and help in reducing selection pressure of insecticides.

Comparing efficacy of a sweep net and a dip method for collection of mosquito larvae in large bodies of water in South Africa.

In this study we tested an alternative method for collecting mosquito larvae called the sweep net catch method and compared its efficiency to that of the traditional dip method. The two methods were compared in various water bodies within Kruger National Park and Lapalala Wilderness area, South Africa. The sweep net catch method performed 5 times better in the collection of Anopheles larvae and equally as well as the dip method in the collection of Culex larvae (p =8.58 x 10 -5). Based on 15 replicates the collector's experience level did not play a significant role in the relative numbers of larvae collected using either method. This simple and effective sweep net catch method will greatly improve the mosquito larval sampling capacity in the field setting.

Larvicidal activity of extracts from Ammi visnaga Linn. (Apiaceae) seeds against Culex quinquefasciatus Say. (Diptera: Culicidae)

Efficacies of the Ammi visnaga seeds extract and a majority of substances on larval Culex quinquefasciatus mortality in various development stages including pupae were studied. The effect of exposure time on larval mortality was also studied. The effect of sublethal concentrations or short exposure times on further larval development and subsequent fecundity in adults were studied as well.Lethal doses of the extract were estimated for the 2nd, 3rd and 4th instar of C. quinquefasciatus (LC50 for 18, 23 and 180 mg L−1, respectively). The majority of furanochromenes, khellin and visnagin, were identified by analysing the extract. Khellin was significantly more effective compared to visnagin, whose LC50 was estimated at 8, 10 and 41 mg L−1 for the 2nd, 3rd and 4th instar larvae. Khellin showed very fast efficacy on mortality for the 3rd instar larvae in a concentration of 100 mg L−1. Fifty percent mortality was determined 30 min after application, a time which was considerably shorter compared to the extract (113 min) or visnagin (169 min). The effect of the application of lethal concentrations on C. quinquefasciatus larval mortality was studied. The least number of adults were hatched after application of the extract and khellin (41.8% and 37.9%, respectively), less than after visnagin application (46.7%) or in the control (94.2%). LC50 application caused lower fecundity in the hatched adults, lower hatchability of the eggs, and also very low natality, more than 77% lower for khellin compared to the control.A short exposure, corresponding to our estimated LT30, caused no significant acute toxicity in the larvae (until 24 h) for the extract or visnagin (4.3% and 11.5%, respectively); however, 18 min of action from khellin caused a 54.3% mortality rate of the larvae within 24 h.

Synthetic predator cues impair immune function and make the biological pesticide Bti more lethal for vector mosquitoes.

The control of vector mosquitoes is one of the biggest challenges facing humankind with the use of chemical pesticides often leading to environmental impact and the evolution of resistance. Although to a lesser extent, this also holds for Bacillus thuringiensis israelensis (Bti), the most widely used biological pesticide to control mosquito populations. This raises the need for the development of integrated pest management strategies that allow the reduction of Bti concentrations without loss of the mosquito control efficiency. To this end, we tested in a laboratory experiment the combined effects of larval exposure to a sublethal Bti concentration and predation risk cues on life history and physiology of larval and adult Culex pipiens mosquitoes. Besides natural predator kairomones and prey alarm cues, we also tested synthetic kairomones of Notonecta predators. Neither Bti nor predation risk cues affected mortality, yet when both stressors were combined mortality increased on average by 133% compared to the treatment with only predation risk cues. This synergistic interaction was also present when Bti was combined with synthetic kairomones. This was further reflected in changes of the composite index of population performance, which suggested lowered per capita growth rates in mosquitoes exposed to Bti but only when Bti was combined with synthetic kairomones. Furthermore, predation risk cues shortened larval development time, reduced mass at metamorphosis in males, and had an immunosuppressive effect in larval and adult mosquitoes which may affect the mosquito vector competence. We provide the first demonstration that synthetic kairomones may generate similar effects on prey as natural kairomones. The identified immunosuppressive effect of synthetic kairomones and the novel lethal synergism type between a biological pesticide and synthetic predator kairomones provide an important proof of principle illustrating the potential of this combination for integrated mosquito control and should in a next step be evaluated under more natural conditions. It may guide novel integrated pest management programs with Bti that incorporate synthetic kairomones and thereby can reduce environmental impact and evolution of resistance creating more efficient and sustainable mosquito control.

Identification of the Temperature Induced Larvicidal Efficacy of Agave angustifolia against Aedes, Culex, and Anopheles Larvae.

Synthetic insecticides are generally employed to control the mosquito population. However, their injudicious over usage and non-biodegradability are associated with many adverse effects on the environment and mosquitoes. The application of environment-friendly mosquitocidals might be an alternate to overcome these issues. In this study, we found that organic or aqueous extracts of Agave angustifolia leaves exhibited a strong larvicidal activity (LD50 28.27 μg/ml) against Aedes aegypti, Culex quinquefasciatus, and Anopheles stephensi larvae within a short exposure of 12 h. The larvicidal activity of A. angustifolia is inherited and independent of the plants vegetative growth. Interestingly, the plant larvicidal activity was observed exclusively during the summer season (April–August, when outside temperature is between 30 and 50°C) and it was significantly reduced during winter season (December–February, when the outside temperature falls to ~4°C or lower). Thus, we hypothesized that the larvicidal components of A. angustifolia might be induced by the manipulation of environmental temperature and should be resistant to the hot conditions. We found that the larvicidal activity of A. angustifolia was induced when plants were maintained at 37°C in a semi-natural environment against the controls that were growing outside in cold weather. Pre-incubation of A. angustifolia extract at 100°C for 1 h killed 60% larvae in 12 h, which gradually increased to 100% mortality after 24 h. In addition, the dry powder formulation of A. angustifolia, also displayed a strong larvicidal activity after a long shelf life. Together, these findings revealed that A. angustifolia is an excellent source of temperature induced bioactive metabolites that may assist the preparedness for vector control programs competently.

Efficacy of Two Larvasonic™ Units Against Culex Larvae and Effects on Common Aquatic Nontarget Organisms in Harris County, Texas.

The Larvasonic™ Field Arm Mobile Wetlands Unit and SD-Mini were tested for efficacy against Culex larvae, and effects on aquatic nontarget organisms (NTO). The Field Arm provided 84.61% to 100% control of caged Culex larvae out to 0.91-m distance in shallow ditches and 60.45% control of Culex larvae at 0.61-m without any effects to caged NTO. Slow ditch treatment achieved 77.35% control compared to fast treatment (20.42%), whereas 77.65% control was obtained along edges of a neglected swimming pool, compared to near the middle (23.97%). In bucket tests, the SD-Mini provided >97% control of Culex and 85.35% reduction of immature giant water bugs, which decreased slightly (83.45%) over the monitoring period, which was not significantly different from cannibalistic damselflies (62.80%), with reduction of both being significantly higher than other NTO tested. There was a small (0.37%) reduction of dragonflies (naiads), due to cannibalism. Both Larvasonic units could effectively augment conventional larvicide operations in smaller areas without causing resistance within mosquito populations or harming NTO when used properly.

Larval development of Culex quinquefasciatus in water with low to moderate.

Population growth and urbanization have increased the potential habitats, and consequently the abundance of Culex quinquefasciatus, the southern house mosquito, a vector of West Nile Virus in urban areas. Water quality is critical in larval habitat distribution and in providing microbial food resources for larvae. A mesocosm experiment was designed to demonstrate which specific components of water chemistry are conducive to larval Culex mosquitoes. Dose-response relationships between larval development and NO3 , NH4 , and PO4 concentrations in stream water were developed through this experiment to describe the isolated effects of each nutrient on pre-adult development. The emergence pattern of Culex mosquitoes was found to be strongly related to certain nutrients, and results showed that breeding sites with higher PO4 or NO3 concentrations had higher larval survival rates. High NO3 concentrations favor the development of male mosquitoes and suppress the development of female mosquitoes, but those adult females that do emerge develop faster in containers with high NO3 levels compared to the reference group. The addition of PO4 in the absence of nitrogen sources to the larval habitat slowed larval development, however, it took fewer days for larvae to reach the pupal stage in containers with combinations of NO3 and PO4 or NH4 and PO4 nutrients. Results from this study may bolster efforts to control WNV in urban landscapes by exploring water quality conditions of Culex larval habitats that produce adult mosquitoes.

Larval ecology of Anopheles coluzzii in Cape Coast, Ghana: water quality, nature of habitat and implication for larval control.

BackgroundThere is a growing interest in larval control intervention to supplement existing malaria control strategies, particularly in urban areas. However, effective implementation requires a good understanding of habitat ecology of Anopheles mosquitoes. Clean water bodies have long been reported by several studies as a preferred breeding habitat for Anopheles gambiae. Other studies have also reported the breeding of An. gambiae in polluted water bodies. However, the term clean or polluted is mostly based on visual examination and is not well defined. This study was conducted with the aim of assessing water quality in Anopheles breeding habitats and the practicability of larval control in Cape Coast, Ghana.MethodsA larval survey was conducted for 15 months in Cape Coast. In individual breeding habitats, habitat characteristics, physicochemical parameters and bacterial fauna were measured in both Anopheles positive breeding (APL) habitats and habitats colonized by only Culex species. The sibling species of An. gambiae were identified using PCR assay.ResultsAnopheles coluzzii dominated in almost all the APL habitats found in this study. The habitats had high levels of salinity and ammonium ions. However, ammonium ions were significantly higher (p = 0.001) in habitats colonized by only Culex larvae compared to APL habitats. About 47 % of the habitats that were colonized by only Culex larvae had no measurable dissolved oxygen while An. coluzzii was absent in such habitats. High concentration of faecal bacteria confirmed faecal contamination in both groups of breeding habitats.ConclusionsFrom the results, it was evident that larval stages of An. coluzzii have tolerance to high levels of salinity and organic pollution in breeding habitats. However, its level of tolerance to organic pollution is probably lower than Culex larvae. The nature of breeding habitats found in the city demonstrates the opportunistic behaviour of An. coluzzii and how its breeding requirements are so intimately intertwined with the haphazard and uncontrolled human activities in the urban area. Considering the nature of APL habitats, larval control intervention could greatly reduce Anopheles population. However, improving basic hygiene and sanitation in the city could even make larval control intervention more practical and cost effective.

Anopheles gambiae Ag55 cell line as a model for Lysinibacillus sphaericus Bin toxin action

Binary toxin (Bin) produced by Lysinibacillus sphaericus is toxic to Culex and Anopheles mosquito larvae. It has been used world-wide for control of mosquitoes that vector disease. The Bin toxin interacts with the glucosidase receptor, Cpm1, in Culex and its orthologue, Agm3, in Anopheles mosquitoes. However, the exact mechanism of its mode of action is not clearly understood. It is essential to understand mode of action of Bin toxin to circumvent the resistance that develops over generations of exposure. A suitable model cell line will facilitate investigations of the molecular action of Bin toxin. Here we report Bin toxin activity on Ag55 cell line that has been derived from an actual target, Anopheles gambiae larvae. The Bin toxin, both in pro and active forms, kills the Ag55 cells within 24h. Bin toxin internalizes in Ag55 cells and also induces vacuolation as tracked by Lysotracker dye. The dose response studies showed that 1.5nM of Bin toxin is sufficient to induce vacuolation and Ag55 cell death. Presence of α-glucosidase gene (Agm3) expression in the Ag55 cells was also confirmed. Thus, Ag55 cells constitute an appropriate model system to decipher the mode of Bin action in mosquito larvae.

First Record and Larval Habitat Description of Culex (Melanoconion) pilosus from Buenos Aires Province, Argentina.

Larvae of Culex (Melanoconion) pilosus were collected during February-April 2014 in temporary pools in "Bosques de Ezeiza," a large forested park, near Buenos Aires city, Argentina. This is the first record in Buenos Aires Province, extending the distribution of this species 380 km to the south. Regarding habitat use, Cx. (Mel.) pilosus is a generalist, although a slight association of larval abundances with pools of lower pH and higher vegetation cover was observed. The comparison of larval instars of Cx. (Mel.) pilosus with those of other genera suggests a life-history strategy similar to that of floodwater mosquitoes.

Effects of fish cues on mosquito larvae development

We investigated the effects of predator-released kairomones on life history traits of larval Culex pipiens (Linnaeus). We compared the development time and survival of sibling larvae, reared in either water conditioned by the presence of Gambusia affinis (Baird and Girard) or fishless control-water. Our results indicate that larvae developing in fish-conditioned water (FCW) pupated faster than larvae in fishless-control water. The effect of FCW on larval survival was evident only in females. Surprisingly, FCW increased female survival. In both development-time and survival, boiling the water eliminated the FCW effect, supporting our hypothesis that fish conditioning is based on kairomones. Accelerated metamorphosis in response to predator released kairomones, evident in our results, is a rarely described phenomenon. Intuitively, when exposed to predator associated signals, aquatic larvae should metamorphose earlier to escape the higher risk of predation. However, theoretical models predict this outcome only under specific conditions. Indeed, longer – rather than shorter – time to metamorphosis is usually observed in response to predation risk. We argue that the response of larval mosquitoes to predation risk is context-dependent. Shortening larval development time may not be an exceptional response, but rather represents a part of a response spectrum that depends on the level of predation risk and resource abundance.

Synthetic predator cues impair immune function and make the biological pesticideBtimore lethal for vector mosquitoes

The control of vector mosquitoes is one of the biggest challenges facing humankind with the use of chemical pesticides often leading to environmental impact and the evolution of resistance. Although to a lesser extent, this also holds for Bacillus thuringiensis israelensis (Bti), the most widely used biological pesticide to control mosquito populations. This raises the need for the development of integrated pest management strategies that allow the reduction of Bti concentrations without loss of the mosquito control efficiency. To this end, we tested in a laboratory experiment the combined effects of larval exposure to a sublethal Bti concentration and predation risk cues on life history and physiology of larval and adult Culex pipiens mosquitoes. Besides natural predator kairomones and prey alarm cues, we also tested synthetic kairomones of Notonecta predators. Neither Bti nor predation risk cues affected mortality, yet when both stressors were combined mortality increased on average by 133% compared to the treatment with only predation risk cues. This synergistic interaction was also present when Bti was combined with synthetic kairomones. This was further reflected in changes of the composite index of population performance, which suggested lowered per capita growth rates in mosquitoes exposed to Bti but only when Bti was combined with synthetic kairomones. Furthermore, predation risk cues shortened larval development time, reduced mass at metamorphosis in males, and had an immunosuppressive effect in larval and adult mosquitoes which may affect the mosquito vector competence. We provide the first demonstration that synthetic kairomones may generate similar effects on prey as natural kairomones. The identified immunosuppressive effect of synthetic kairomones and the novel lethal synergism type between a biological pesticide and synthetic predator kairomones provide an important proof of principle illustrating the potential of this combination for integrated mosquito control and should in a next step be evaluated under more natural conditions. It may guide novel integrated pest management programs with Bti that incorporate synthetic kairomones and thereby can reduce environmental impact and evolution of resistance creating more efficient and sustainable mosquito control.

Surveillance of Mosquitoes Larva at Selected Areas of Mymensingh District in Bangladesh

Mosquito borne diseases have a great impact on human and animal health throughout the world including Bangladesh. An entomological survey was conducted at the Bangladesh Agricultural University (BAU) campus and its surrounding areas from January to June, 2013 to identify the mosquito larvae and to investigate the different breeding site preferences of mosquitoes. Mosquito larvae were collected using dipper, ladle spoon, dropper and pipette from rice field, botanical garden, dairy farm, poultry farm, drains, lakes, ponds, staff quarters, residential halls and cattle sheds. A total of 1397 mosquito larvae were collected from the places. Ten species of mosquitoes under three genera were identified. The recorded genera were Anopheles (An.), Culex (Cx.) and Aedes (Ae.). The collected species were An. bengalensis, An. vagus, Cx. fuscocephala, Cx. quinquefasciatus, Cx. tritaeniorhynchus, Cx. gelidus, Cx. vishnui, Cx. whitmorei, Cx. sitiens and Ae. albopictus. Among the collected larvae, Culex was found most prevalent (64.3%), followed by Anopheles (26.7%) and Aedes were the least prevalent (9.0%). The results of this study provide evidence that drains were the most common aquatic habitat of Anopheles and Culex larvae in Boyra, Kewatkhali and Balashpur. In BAU campus, agronomy fields were found having the highest percentage (35.2%) of mosquito larvae than the other places of the campus. This study suggests that although drains and rice fields are the richest habitats, since Anopheles and Culex can breed in all available breeding habitats. This larval survey will help us to conduct future mosquito control activities and provide us useful information to schedule larvicide’s application to control the mosquito borne diseases in the study areas as well as throughout the country.DOI: http://dx.doi.org/10.3329/bjvm.v13i1.23962Bangl. J. Vet. Med. (2015). 13 (1): 79-88

Morphological and fecundity traits of Culex mosquitoes caught in gravid traps in urban and rural Berkshire, UK.

Culex pipiens s.l. is one of the primary vectors of West Nile Virus in the USA and Continental Europe. The seasonal abundance and eco-behavioural characteristics of the typical form, Cx. pipiens pipiens, make it a key putative vector in Britain. Surveillance of Culex larvae and adults is essential to detect any changes to spatial and seasonal activity or morphological traits that may increase the risk of disease transmission. Here we report the use of the modified Reiter gravid box trap, which is commonly used in the USA but scarcely used in the UK, to assess its suitability as a tool for British female Culex mosquito surveillance. Trapping was carried out at 110 sites in urban and rural gardens in Berkshire in May, July and September 2013. We tested if reproductively active adult female Culex are more abundant in urban than rural gardens and if wing characteristic traits and egg raft size are influenced by location and seasonal variations. Gravid traps were highly selective for Culex mosquitoes, on average catching significantly more per trap in urban gardens (32.4 ± 6.2) than rural gardens (19.3 ± 4.0) and more in July than in May or September. The majority of females were caught alive in a good condition. Wing lengths were measured as an indicator of size. Females flying in September were significantly smaller than females in May or July. Further non-significant differences in morphology and fecundity between urban and rural populations were found that should be explored further across the seasons.

Lysinibacillus sphaericus binary toxin induces apoptosis in susceptible Culex quinquefasciatus larvae

During sporulation, a Gram-positive bacterium Lysinibacillus sphaericus (Ls) produces the mosquito larvicidal binary toxin composed of 2 subunits, BinA and BinB. Full toxicity against Culex and Anopheles mosquito larvae is achieved when both subunits are administered together at equimolar amounts. Although cellular responses to Bin toxin have been reported in previous studies, it remains essential to extensively examine the cytopathic effects in vivo to define the underlying mechanism of larval death. In this study, 4th instar Culex quinquefasciatus larvae fed with different doses of Bin toxin were analyzed both for ultrastructural as well as biochemical effects. Typical morphological changes consistent with apoptosis were observed in mosquito larvae exposed to Bin toxin, including mitochondrial swelling, chromatin condensation, cytoplasmic vacuolization and apoptotic cell formation. Bin toxin also induced the activation of caspase-9 and caspase-3 in larval midgut cells. Our current observations thus suggest that Bin toxin triggers apoptosis via an intrinsic or mitochondrial pathway in vivo, possibly contributing to larval death.

Toxicity of seaweed-synthesized silver nanoparticles against the filariasis vector Culex quinquefasciatus and its impact on predation efficiency of the cyclopoid crustacean Mesocyclops longisetus.

Nearly 1.4 billion people in 73 countries worldwide are threatened by lymphatic filariasis, a parasitic infection that leads to a disease commonly known as elephantiasis. Filariasis is vectored by mosquitoes, with special reference to the genus Culex. The main control tool against mosquito larvae is represented by treatments with organophosphates and insect growth regulators, with negative effects on human health and the environment. Recently, green-synthesized nanoparticles have been proposed as highly effective larvicidals against mosquito vectors. In this research, we attempted a reply to the following question: do green-synthesized nanoparticles affect predation rates of copepods against mosquito larvae? We proposed a novel method of seaweed-mediated synthesis of silver nanoparticles using the frond extract of Caulerpa scalpelliformis. The toxicity of the seaweed extract and silver nanoparticles was assessed against the filarial vector Culex quinquefasciatus. Then, we evaluated the predatory efficiency of the cyclopoid crustacean Mesocyclops longisetus against larval instars of C. quinquefasciatus in a nanoparticle-contaminated water environment. Green-synthesized silver nanoparticles were characterized by UV-vis spectrum, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and X-ray diffraction (XRD). In mosquitocidal assays, the LC₅₀ values of the C. scalpelliformis extract against C. quinquefasciatus were 31.38 ppm (I), 46.49 ppm (II), 75.79 ppm (III), 102.26 ppm (IV), and 138.89 ppm (pupa), while LC₅₀ of silver nanoparticles were 3.08 ppm, (I), 3.49 ppm (II), 4.64 ppm (III), 5.86 ppm (IV), and 7.33 ppm (pupa). The predatory efficiency of the copepod M. longisetus in the control treatment was 78 and 59% against I and II instar larvae of C. quinquefasciatus. In a nanoparticle-contaminated environment, predation efficiency was 84 and 63%, respectively. Predation was higher against first instar larvae over other instars. Overall, our study showed that seaweed-synthesized silver nanoparticles can be proposed in synergy with biological control agents against Culex larvae, since their use leads to little detrimental effects against aquatic predators, such as copepods.