In vitro and In vivo toxicity assessment of phytofabricated ZnO nanoparticles showing bacteriostatic effect and larvicidal efficacy against Culex quinquefasciatus.

Murraya koenigii berry extract based zinc oxide nanoparticles (Mk-ZnO NPs) were synthesized by simple co-precipitation method and examined for bacteriostatic and larvicidal efficiency. Synthesized Mk-ZnO NPs were characterized by UV-Vis spectroscopy at 336 nm. X-Ray diffraction (XRD) showed crystalline nature as hexagonal. Fourier transform infrared spectroscopy (FTIR) spectrum exhibited strong peak at 3442.80 cm-1. Field emission scanning electron microscopy (FE-SEM) showed hexagonal shape of the particle. Transmission electron microscopy (TEM) measured 10-15 nm sized Mk-ZnO NPs. EDX peaks confirm 71.99% of zinc and 11.42% of oxide in Mk-ZnO NPs. Minimum inhibitory concentration (MIC) analysis reveals Mk-ZnO NPs inhibit growth of Gram positive (Staphylococcus aureus, Lysinibacillus fusiformis) and Gram negative (Proteus vulgaris, Providencia vermicola) bacteria at 40 and 50 µg mL-1 respectively. Live & dead assay confirms that Mk-ZnO NPs inhibits bacterial growth at 50 µg mL-1. Bacterial biofilm thickness significantly reduced by Mk-ZnO NPs at 50 µg mL-1. In vitro toxicity of Mk-ZnO NPs on RAW 264.7 macrophages determines 90-50% cell viability at concentrations of 10-100 µg mL-1. In vivo toxicity assay results indicate the lethal concentration of Artemia nauplii were LC50-78.73 µg mL-1 and LC90-130.03 µg mL-1. Larvicidal activity of Mk-ZnO NPs towards mosquito larvae of Culex quinquefasciatus were observed at LC50-2.1 µg mL-1 and LC90-12.1 µg mL-1. Finally the study discloses, potential bacteriostatic effect and mosquito larvae controlling capacity of Mk-ZnO NPs.

Assessment of the Suna trap for sampling mosquitoes indoors and outdoors.

BACKGROUND: Entomological monitoring is important for public health because it provides data on the distribution, abundance and host-seeking behaviour of disease vectors. Various methods for sampling mosquitoes exist, most of which are biased towards, or specifically target, certain portions of a mosquito population. This study assessed the Suna trap, an odour-baited trap for sampling host-seeking mosquitoes both indoors and outdoors. METHODS: Two separate field experiments were conducted in villages in southern Malawi. The efficiency of the Suna trap in sampling mosquitoes was compared to that of the human landing catch (HLC) indoors and outdoors and the Centers for Disease, Control and Prevention Light Trap (CDC-LT) indoors. Potential competition between two Suna traps during simultaneous use of the traps indoors and outdoors was assessed by comparing mosquito catch sizes across three treatments: one trap indoors only; one trap outdoors only; and one trap indoors and one trap outdoors used simultaneously at the same house. RESULTS: The efficiency of the Suna trap in sampling female anophelines was similar to that of HLC indoors (P = 0.271) and HLC outdoors (P = 0.125), but lower than that of CDC-LT indoors (P = 0.001). Anopheline catch sizes in the Suna trap used alone indoors were similar to indoor Suna trap catch sizes when another Suna trap was simultaneously present outdoors (P = 0.891). Similarly, catch sizes of female anophelines with the Suna trap outdoors were similar to those that were caught outdoors when another Suna trap was simultaneously present indoors (P = 0.731). CONCLUSIONS: The efficiency of the Suna trap in sampling mosquitoes was equivalent to that of the HLC. Whereas the CDC-LT was more efficient in collecting female anophelines indoors, the use of this trap outdoors is limited given the requirement of setting it next to an occupied bed net. As demonstrated in this research, outdoor collections are also essential because they provide data on the relative contribution of outdoor biting to malaria transmission. Therefore, the Suna trap could serve as an alternative to the HLC and the CDC-LT, because it does not require the use of humans as natural baits, allows standardised sampling conditions across sampling points, and can be used outdoors. Furthermore, using two Suna traps simultaneously indoors and outdoors does not interfere with the sampling efficiency of either trap, which would save a considerable amount of time, energy, and resources compared to setting the traps indoors and then outdoors in two consecutive nights.

Selection, Realized Heritability, and Fitness Cost Associated With Dimethoate Resistance in a Field Population of Culex quinquefasciatus (Diptera: Culicidae).

Mosquitoes are known to be vectors of numerous diseases leading to human morbidity and mortality at large scale in the world. Insecticide resistance has become a serious concern in controlling the insect vectors of public health importance. Dimethoate is an organophosphate insecticide used to control different insect pests including mosquitoes. Biological parameters of susceptible, unselected, and dimethoate-selected strains of Culex quinquefasciatus Say were studied in the laboratory to recognize resistance development potential and associated fitness cost. The dimethoate-selected strain showed 66.48-fold resistance to dimethoate compared with the susceptible strain after three continuous selections of generations. Realized heritability estimates of dimethoate resistance in Cx. quinquefasciatus yielded a value of 0.19. In dimethoate-selected strain, the biological traits including larval weight, survival from first instar to pupae, fecundity, number of next-generation larvae, relative fitness, net reproductive rate, intrinsic rate of natural increase, and biotic potential were significantly reduced as compared with the unselected strain. However, adult longevity, mean relative growth rate, weight of egg raft, female ratio, pupal duration, and emergence rate of the dimethoate-selected strain did not differ significantly compared with that of the unselected strain. This study provides useful information to devise retrospective management strategy for dimethoate resistance in Cx. quinquefasciatus.

Preliminary assessment of the potential role of urbanization in the distribution of carbamate and organophosphate resistant populations of Culex species in Ghana.

BACKGROUND: Besides its role as a pathogen vector, Culex species also indirectly promotes the transmission of malaria if the use of bed nets or indoor residual spraying is discontinued due to a lack of insecticide efficacy against it. A recent survey revealed widespread occurrence of pyrethroid resistance among urban populations of this mosquito in Ghana. In order to plan and implement insecticide-based resistance management strategies, this study was carried out to assess resistance status of Culex species to organophosphate and carbamate in urban areas in Ghana and the possible mechanisms involved as well as environmental factors associated with its distribution. METHODS: Mosquito larvae were sampled from various land use and ecological settings and in different seasons. In adults, susceptibility to organophosphates (fenitrothion, malathion) and carbamates (propoxur, bendiocarb) were determined. Mixed function oxidase (MFO) and α- and ß-esterase assays, as well as a PCR diagnostic assay to determine ace1 mutation were performed in individual mosquitoes. RESULTS: Culex quinquefasciatus as well as C. decens and other unidentified Culex species were found breeding in polluted water bodies in the study sites. Across all sites and seasons, carbamate induced mortality was 94.1% ± 15.4 whereas mortality caused by organophosphate was 99.5% ± 2.2. In addition, ace1 mutation and high levels of esterases were detected in some of the mosquito populations. There was a strong correlation between susceptibility status of the mosquitoes and the level of absorbance of ß-esterase (Pearson r=- 0.841, p=0.004). CONCLUSIONS: The study found low prevalence of resistance to carbamate and organophosphate insecticides among Culex species from Ghana. However, there were populations with ace1 mutations and high levels of esterases, which can confer high resistance to these classes of insecticides. Thus, it is important to monitor activities or behaviour that has the potential to select for carbamate and organophosphate resistance populations.

Assessment of residual bio-efficacy and persistence of Ipomoea cairica plant extract against Culex quinquefasciatus Say mosquito.

Specification on residual action of a possible alternative insecticide derived from plant materials is important to determine minimum interval time between applications and the environmental persistence of the biopesticides. The objective of this study is to evaluate crude acethonilic extract of Ipomoea cairica leaves for its residual and persistence effects against Culex quinquefasciatus larvae. Wild strain of Cx. quinquefasciatus larvae were used for the purpose of the study. Two test designs, replenishment of water and without replenishment of water were carried out. For the first design, a total of 10 ml of test solution containing Ip. cairica extracts was replenished daily and replaced with 10 ml of distilled water. For the second design, treatment water was maintained at 1500 ml and only evaporated water was refilled. Larval mortality was recorded at 24 hours post-treatment after each introduction period and trials were terminated when mortality rate falls below 50%. Adult emergences from survived larvae were observed and number of survivals was recorded. For the non-replenishment design, mortality rate significantly reduced to below 50% after 28 days, meanwhile for replenishment of water declined significantly after 21 days (P < 0.05). There was no adult emergence observed up to seven days for non-replenishment and first two days for replenishment of water design. The short period of residual effectiveness of crude acethonilic extract of Ip. cairica leaves with high percentage of larval mortality on the first few days, endorses fewer concerns of having excess residues in the environment which may carry the risk of insecticide resistance and environmental pollution.

Assessment of anti mosquito measures in households and resistance status of Culex species in urban areas in southern Ghana: implications for the sustainability of ITN use.

OBJECTIVE: To determine resistance status of Culex species to different class of insecticides and assess the major anti-mosquito strategies employed by urban households and their possible effects on malaria acquisition. METHODS: Structured questionnaires were randomly administered to obtain information on demographic characteristics, measures that people use to prevent mosquito bites and their perception of where mosquitoes breed in their communities. Adult susceptibility assays were also carried out using WHO discriminating dosages of four insecticides from different chemical classes against Culex species. RESULTS: Majority of respondents (54.75%) preferred the use of domestic insecticides in the form of aerosols and coils. Among households that used domestic insecticides, the use of coil was most frequent (62.9%) with a mean (95% CI) of 9.18 (8.99, 9.37) coils per week. Strong level of pyrethroid-resistance and multiple insecticide resistance in Culex species were also detected in some of the study sites. CONCLUSIONS: The excessive use of domestic insecticides and high level of resistance in Culex species observed in the study area has implications for the ITN component of the nation's malaria control program in more subtle ways. People will lose interest in the use of ITN when it fails to protect users from bites of resistant Culex species. Excessive use of domestic insecticides may also select resistance in both malaria vectors and Culex species. On this account we recommend that nuisance mosquitoes must be controlled as part of malaria control programs to improve acceptance and utilization of ITN.

Energetic cost of insecticide resistance in Culex pipiens mosquitoes.

The extensive use of insecticides to control vector populations has lead to the widespread development of different mechanisms of insecticide resistance. Mutations that confer insecticide resistance are often associated to fitness costs that prevent them from spreading to fixation. In vectors, such fitness costs include reductions in preimaginal survival, adult size, longevity, and fecundity. The most commonly invoked explanation for the nature of such pleiotropic effects of insecticide resistance is the existence of resource-based trade-offs. According to this hypothesis, insecticide resistance would deplete the energetic stores of vectors, reducing the energy available for other biological functions and generating trade-offs between insecticide resistance and key life history traits. Here we test this hypothesis by quantifying the energetic resources (lipids, glycogen, and glucose) of larvae and adult females of the mosquito Culex pipiens L. resistant to insecticides through two different mechanisms: esterase overproduction and acetylcholinesterase modification. We find that, as expected from trade-off theory, insecticide resistant mosquitoes through the overproduction of esterases contain on average 30% less energetic reserves than their susceptible counterparts. Acetylcholinesterase-modified mosquitoes, however, also showed a significant reduction in energetic resources (20% less). We suggest that, in acetylcholinesterase-modified mosquitoes, resource depletion may not be the result of resource-based trade-offs but a consequence of the hyperactivation of the nervous system. We argue that these results not only provide a mechanistic explanation for the negative pleiotropic effects of insecticide resistance on mosquito life history traits but also can have a direct effect on the development of parasites that depend on the vector's energetic reserves to fulfil their own metabolic needs.

Coffee husk waste for fermentation production of mosquitocidal bacteria.

Coffee husk waste (CHW) discarded as bio-organic waste, from coffee industries, is rich in carbohydrates. The current study emphasizes the management of solid waste from agro-industrial residues for the production of biopesticides (Bacillus sphaericus, and B. thuringiensis subsp. israelensis), to control disease transmitting mosquito vectors. An experimental culture medium was prepared by extracting the filtrates from coffee husk. A conventional culture medium (NYSM) also was prepared. The studies revealed that the quantity of mosquitocidal toxins produced from CHW is at par with NYSM. The bacteria produced in these media, were bioassayed against mosquito vectors (Culex quinquefasciatus, Anopheles stephensi, and Aedes aegypti) and it was found that the toxic effect was statistically comparable. Cost-effective analysis have revealed that, production of biopesticides from CHW is highly economical. Therefore, the utilization of CHW provides dual benefits of effective utilization of environmental waste and efficient production of mosquitocidal toxins.

Field evaluation of an insect growth regulator, pyriproxyfen 0.5% GR against Culex quinquefasciatus, the vector of Bancroftian filariasis in Pondicherry, India.

Pyriproxyfen, 0.5% granular formulation (GR), an insect growth regulator (IGR) was tested against Culex quinquefasciatus larvae and pupae in disused wells, cesspits and drains at the dosages of 0.1, 0.25, and 0.5kg(ai)/ha to determine the most appropriate field dosage. The IGR was found to be effective against C. quinquefasciatus larvae and pupae in all the larval habitats tested. In drains with slow moving water, application of pyriproxyfen 0.5% GR at 0.5kg(ai)/ha resulted in >80% emergence inhibition (EI) of adults for 4 weeks. At 0.1 and 0.25kg(ai)/ha, the EI was always less than 80%. In stagnant drains, the IGR yielded >80% EI for 1-week period at 0.1kg(ai)/ha. At 0.25 and 0.5kg(ai)/ha, the efficacy was 5-10 times higher. In cesspits, the EI was >80% for 6 weeks when pyriproxyfen 0.5% GR was applied at 0.1kg(ai)/ha, for 11 weeks at 0.25kg(ai)/ha and 9 weeks at 0.5kg(ai)/ha. In disused wells treated at the dosage 0.1kg(ai)/ha, there was >80% EI for 15 weeks and at 0.25 and 0.5kg(ai)/ha, the effective duration was about 1.6-1.8 times longer (24-28 weeks). Considering the quantum of IGR required and the cost and also for safety reasons, the low dosage 0.1kg(ai)/ha is recommended as the field dosage to be applied at weekly interval in stagnant drains, 6 weeks interval in cesspits and 15 weeks interval in disused wells. Since pyriproxyfen 0.5% GR has a relatively longer residual effect than the other IGRs the operational cost could be minimized. Pyriproxyfen 0.5% GR could be one of the choices in the chemical control strategy in Integrated Vector Control Programmes.

High Wolbachia density correlates with cost of infection for insecticide resistant Culex pipiens mosquitoes.

In the mosquito Culex pipiens, insecticide resistance genes alter many life-history traits and incur a fitness cost. Resistance to organophosphate insecticides involves two loci, with each locus coding for a different mechanism of resistance (degradation vs. insensitivity to insecticides). The density of intracellular Wolbachia bacteria has been found to be higher in resistant mosquitoes, regardless of the mechanism involved. To discriminate between costs of resistance due to resistance genes from those associated with elevated Wolbachia densities, we compared strains of mosquito sharing the same genetic background but differing in their resistance alleles and Wolbachia infection status. Life-history traits measured included strength of insecticide resistance, larval mortality, adult female size, fecundity, predation avoidance, mating competition, and strength of cytoplasmic incompatibility (CI). We found that: (1) when Wolbachia are removed, insecticide resistance genes still affect some life-history traits; (2) Wolbachia are capable of modifying the cost of resistance; (3) the cost of Wolbachia infections increases with their density; (4) different interactions occurred depending on the resistance alleles involved; and (5) high densities of Wolbachia do not increase the strength of CI or maternal transmission efficiency relative to low Wolbachia densities. Insecticide resistance genes generated variation in the costs of Wolbachia infections and provided an interesting opportunity to study how these costs evolve, a process generally operating when Wolbachia colonizes a new host.

Assessment of two natural marine toxins (Microcystis aeruginosa and Parasicyonis actinostoloides) for the control of some medical and agriculture insects with reference to the action on mice.

The present work describes the insecticidal potentiality from Microcystis aeruginosa (cyanobacteria) and Parasicyonis actinostoloides (Sea anemone) water extracts against 3rd larval instar of medical insects (Musca domestica and Culex pipiens) and 4th larval instar of agriculture insects (Spodoptera littoralis and Agrotis ipsilon. The LD50s of M. aeruginosa extract were 1.94, 2.33, 7.59 and 9.10 mg/ml. for M. domestica, C. pipiens, S. littoralis and A. ipsilon). respectively. While LD50s of P. actinostoloides extract were 19.28, 24.7, 27.3 and (29.4) mg/ml. for M. domestica, C. pipiens, S. littoralis, and A. ipsilon respectively. The aqueous extracts of M. aeruginosa and P. actinostoloides had no acute or chronic marked effect on mice as serum acetyl cholinesterase and gave more or less nearly the same level of AchE activity at the end of decapitation periods.