Evaluation of indigenous plants’ extracts for mosquitocidal activity against different stages of Culex quinquefasciatus say (Diptera: Culicidae)

Being vector of West Nile Virus and falariasis the control of Culex quinquefasciatus is likely to be essential. Synthetic insecticide treatment is looking most effective for vectors mosquito control. However, these products are toxic to the environment and non-target organisms. Consequently, ecofriendly control of vectors mosquito is needed. In this regard botanical insecticide is looking more fruitful. Therefore, the present research aimed to investigate the effectiveness of methanolic extract and various fractions, including, n-hexane, ethyl-acetate, chloroform, and aqueous fraction, obtained from methanolic extract of Ailanthus altissima, Artemisia scoparia, and Justicia adhatoda using separating funnel against larval, pupal, and adult stages of Culex quinquefasciatus. The larvae and pupae of Culex quinquefasciatus were exposed to various concentrations (31.25-1000 ppm) of methanolic extract and its fractions for 24 hours of exposure period. For knock-down bioassay (filter paper impregnation bioassay) different concentration of the methanolic extract and its various fractions (i.e. 0.0625, 0.125, 0.25, 0.5 and 1mg/mL) were applied for 1 hour exposure period. The results were statistically analysed using standard deviation, probit analysis, and linear regression. The R2 values of larvae, pupae, and adult range from 0.4 to 0.99. The values of LC50 (concentration causing 50% mortality) for late 3rd instar larvae after 24 hours exposure period range from 93-1856.7 ppm, while LC90 values range from 424 -7635.5ppm. The values of LC50for pupae range form 1326.7-6818.4ppm and and values of LC90 range from 3667.3-17427.9ppm, respectively. The KDT50 range from 0.30 to 2.8% and KDT90 values range from1.2 to 110.8%, respectively. In conclusion, Justicia adhatoda may be effective for controlling populations of vector mosquito.

Mosquitocidal activities of Chenopodium botrys whole plant n-hexane extract against Culex quinquefasciatus

This research aimed to investigate various mosquitocidal activities of Chenopodium botrys whole- plant n-hexane extract against Culex quinquefasciatus. The extract showed remarkable larvicidal, pupicidal, adulticidal, oviposition deterrent and adult emergence inhibitory activities against Cx. quinquefasciatus. During the larvicidal and pupicidal activities, the 24-hour lethal concentration (LC50) of extract against 2nd instar larvae, 4th instar larvae and pupae were 324.6, 495.6 and 950.8 ppm, respectively. During the CDC (Centers for Disease Control and Prevention) bottle bioassay for adulticidal activity, the median knockdown times (KDT50) at 1.25% concentration was 123.4 minutes. During the filter paper impregnation bioassay for adulticidal activity, the KDT50 value at 0.138 mg/cm2 concentration was 48.6 minutes. The extract was fractionated into 14 fractions through silica gel column chromatography which were then combined into six fractions on the basis of similar retention factor (Rf) value. These fractions were screened for adulticidal activity by applying CDC bottle bioassay. The fraction obtained through 60:40 to 50:50% n-hexanes-chloroform mobile phase with 0.5 Rf value showed 100% adulticidal activity at 0.2% concentration. During oviposition deterrent activity, the highest concentration (1000 ppm) showed 71.3 ± 4.4% effective repellence and 0.6 ± 0.1 oviposition activity index. During adult emergence inhibition activity, the median emergence inhibition (EI50) value was 312.3 ppm. From the outcome of the present investigation, it is concluded that the n-hexane extract of C. botrys whole- plant possesses strong larvicidal, pupicidal, adulticidal, oviposition deterrent and adult emergence inhibitory activities against Cx. quinquefasciatus.

Larvicidal Activity of Leaf Extract From Mauritiella armata (Aceraceae) on Aedes aegypti and Culex quinquefasciatus (Culicidae)

The mosquitoes Aedes aegypti and Culex quinquefasciatus (Diptera: Culicidae) are important vectors of several arboviruses, and are relevant public health problems. Conventional control, using chemical larvicides have selected resistant Culicidae populations and caused negative effects on the environment and human health. However, the use of plant extracts has represented a sustainable alternative for insect control. Popularly known as Xiriri, Mauritiella armata (Mart.) Burret (Aceraceae) is an abundant palm tree in Vereda ecosystems and has economic and social importance. In this study, the aim was to evaluate the larvicidal activity of the aqueous extract (AE) leaves of this plant on Ae. aegypti and Cx. quinquefasciatus larvae. The mortalities of larvae were analyzed after treatment with four concentrations of the extract, comparing with a negative control using mineral water. The AE promoted 100% efficacy against Ae. aegypti larvae at 7.9 mg/mL. The lethal concentration to promote 90% mortality of Cx. quinquefasciatus larvae was estimated at 30.57 mg/mL. After chromatographic analyses, flavonoids, catechin and carbohydrates were detected. AE from M. armata leaves presented high larvicidal activity against Ae. aegypti and Cx. quinquefasciatus, and represents a promising alternative to be used in vector control.

Occurrence of L1014F and L1014S mutations in insecticide resistant Culex quinquefasciatus from filariasis endemic districts of West Bengal, India

Introduction Lymphatic filariasis causes long term morbidity and hampers the socio-economic status. Apart from the available treatments and medication, control of vector population Culex quinquefasciatus Say through the use of chemical insecticides is a widely applied strategy. However, the unrestrained application of these insecticides over many decades has led to resistance development in the vectors. Methods In order to determine the insecticide susceptibility/resistance status of Cx. quinquefasciatus from two filariasis endemic districts of West Bengal, India, wild mosquito populations were collected and assayed against six different insecticides and presence of L1014F; L1014S kdr mutations in the voltage-gated sodium channel gene was also screened along with the use of synergists to evaluate the role of major detoxifying enzymes in resistance development. Results The collected mosquito populations showed severe resistance to insecticides and the two synergists used–PBO (piperonyl butoxide) and TPP (triphenyl phosphate), were unable to restore the susceptibility status of the vector thereupon pointing towards a minor role of metabolic enzymes. kdr mutations were present in the studied populations in varying percent with higher L1014F frequency indicating its association with the observed resistance to pyrethroids and DDT. This study reports L1014S mutation in Cx. quinquefasciatus for the first time.

The Central Role of Multiple P450 Genes and Their Co-factor CPR in the Development of Permethrin Resistance in the Mosquito Culex quinquefasciatus

Mosquitoes’ increasing resistance to insecticides is becoming a major threat for control efforts worldwide. Multiple P450 genes that are up-regulated in permethrin resistant strains of Culex quinquefasciatus have been linked to the development of resistance. In the current study, we characterized the function of six P450 genes, CYP6P14, CYP6BZ2, CYP9J33, CYP9J34, CYP9J40, and CYP9J45, that are overexpressed in the permethrin resistant Culex mosquitoes and showed their capability in metabolism of permethrin. These six P450 genes can convert 3-phenoxybenzoic alcohol (PBCHO) to a less toxic product, 3-phenoxybenzoic acid (PBCOOH), indicating that these P450s play an important role in permethrin degradation pathways. Although we know multiple P450 genes are over-expressed in permethrin resistant Culex mosquitoes, it remains to be seen whether cytochrome P450-reductase (CPR) gene that are co-overexpressed with P450 genes in permethrin resistant mosquitoes do indeed serve as a resistance mechanism. An in-depth investigation of the expression of CPR gene in resistant mosquitoes was conducted in permethrin resistant mosquitoes. The finding of CPR gene overexpression in permethrin resistant mosquitoes suggested the importance of co-overexpression of multiple P450 genes with their obligatory electron donor CPR in the complex detoxification system, boosting the metabolism of permethrin and hence the development of permethrin resistance in Cx. quinquefasciatus.

Pyrethroid Susceptibility in Culex quinquefasciatus Say. (Diptera: Culicidae) Populations from Delta State, Niger-Delta Region, Nigeria

The development of insecticide resistance in different species of mosquitoes to Pyrethroids is a major challenge for vector-borne diseases transmitted by mosquitoes. Failure of Pyrethroids in control of mosquitoes would impact negatively on the gains recorded in control of mosquito-borne diseases in previous years. In anticipation of a country-wide deployment of Pyrethroid-treated nets for control of mosquito-borne diseases in Nigeria, this study assessed susceptibility of Culex quinquefasciatus Say. (Diptera: Culicidae) to Pyrethroids in Owhelogbo, Ejeme and Oria-Abraka communities in Delta State, Niger-Delta, Nigeria. Three to five day old Cx. quinquefasciatus were exposed to Deltamethrin (0.05%), Permethrin (0.75%), and Alphacypermethrin (0.05%) using World Health Organization bioassay method. Polymerase chain reaction (PCR) was employed in characterization of species and knockdown mutation. Results revealed that Cx. quinquefasciatus were generally susceptible (98-100%) to Deltamethrin, Permethrin, and Alphacypermethrin in the three communities with the exception of Owhelogbo where resistance to Deltamethrin (97%) was suspected. Knockdown time to Deltamethrin (11.51, 11.23, and 12.68 min), Permethrin (28.75, 13.26, and 14.49 min), and Alphacypermethrin (15.07, 12.50, and 13.03 min) were considerably low for Owhelogbo, Ejeme, and Oria-Abraka Cx. quinquefasciatus populations, respectively. Species identification result showed that all amplified samples were Cx. quinquefasciatus; however, no kdr allele was found in the three populations. Deployment of pyrethroid-treated nets for control of mosquito-borne diseases in Niger-Delta region of Nigeria is capable of reducing burden of diseases transmitted by Cx. quinquefasciatus as well as addressing nuisance value of the vector; however, caution must be entertained so as not to increase selection pressure thereby aiding resistance development.

Deltamethrin interacts with Culex quinquefasciatus odorant-binding protein: a novel potential resistance mechanism

Background Odorant-binding proteins (OBPs) play important roles in many physiological processes of mosquitoes. Previous high-throughput sequencing studies have revealed that some OBPs of Culex quinquefasciatus might be involved in the development of resistance to insecticides. Methods Based on the results of sequencing analyses, the OBP28 gene was selected for evaluation in this study. Three laboratory strains of Cx. quinquefasciatus [susceptible strain (SS), deltamethrin-resistant strain 1 (HN) and deltamethrin-resistant strain 2 (RR)] were first examined by using the Centers for Disease Control and Prevention bottle bioassay, after which the expression level of the OBP28 gene in the susceptible and deltamethrin-resistant strains was determined by real-time quantitative polymerase chain reaction. The OBP28 gene in deltamethrin-resistant strain RR was silenced using RNA interference technology. The expression level of OBP28 and the resistance level were tested in the silenced strain and control strain after microinjection of double-stranded RNA for a 48-h interference period. Four field-collected strains (henceforth ‘field strains’) of Cx. quinquefasciatus were also examined for their resistance to deltamethrin and levels of OBP28 expression. Finally, a correlation analysis between deltamethrin resistance and gene expression was carried out for all seven strains, i.e. the four field strains and the three laboratory strains. Results In the bioassay, the mortality of SS, HN and RR was 100%, 21.33% and 1.67%, respectively. The relative expression levels of OBP28 in strains HN and RR were 6.30- and 6.86-fold higher, respectively, than that of strain SS. After silencing of the OBP28 gene, the mortality of strain RR was 72.20% and that of the control strain 26.32%. The mortality of strain RR increased significantly after interference compared to that of the control strain. There was a negative correlation between OBP28 gene expression and mortality in adult mosquitoes after exposure to deltamethrin. Conclusions To our knowledge, this study shows for the first time a correlation between the expression of a gene coding for OBP and insecticide resistance in mosquitoes. The potential resistance mechanism that was elucidated provides a new target gene for the surveillance of resistance in mosquitoes. Graphical abstract