Analysis of the insecticide resistance mechanism in Anopheles culicifacies sensu lato from a malaria-endemic state in India.

BACKGROUND: Understanding the dynamics and mechanisms of insecticide resistance in malaria vectors is crucial for vector control activities. The present study investigates the level of insecticide resistance in Anopheles culicifacies and explores the role of two main mechanisms in conferring resistance target site insensitivity and metabolic resistance. METHODS: A. culicifacies mosquitoes were collected and the voltage-gated sodium channel (VGSC) gene was amplified and sequenced to analyse the knockdown resistance (kdr) mutations. Further, a non-experimental homology model was generated to investigate the effect of kdr mutations on the conformation of protein. Metabolic resistance was determined using bioassay-based resistant and susceptible mosquitoes and the expression levels of the genes CYP6Z1 and GSTe2 were compared between the two groups. RESULTS: Sequence analysis of the VGSC gene revealed the presence of L1014F (n=48 [17%]), L1014S and V1010L (n=5 [1.7%]) mutations in the study area. In gene expression studies, a significant upregulation of CYP6Z1 in deltamethrin-resistant (fold change 243.62; p=0.02) mosquitoes and that of GSTe2 in dichlorodiphenyltrichloroethane (fold change 403.45; p=0.01) and alpha-cypemethrin resistant (fold change 217.51; p=0.0005) mosquitoes was observed. CONCLUSIONS: The study revealed that expression of the genes (CYP6Z1 and GSTe2) conferring metabolic resistance play a key role in insecticide resistance in A. culicifacies populations in central India. However, mutations L101F, L10104S and V10101L also have a role to some extent in spreading resistance. GeneBank accession numbers: MW559058, MW559059 and MW559060 Cover Image: Workflow of Chimera-Modeller interface. In the top window of Chimera's multi-align viewer the sequence alignment of VGSC proteins of human (pdb id_6AGF), cockroach (pdb id 5XOM) and A. culicifacies (ACT176122.1) is shown. The dialog box in the middle is of the comparative modelling tool of Modeller. The A. culicifacies sequence is designated as the target while human and cockroach sequences are templates. Upon selection of the template sequences in the dialog box, the structures of the respective proteins are displayed in the Chimera window. As the run is completed, the results are displayed in the form of a list of models with their scores in a table.

Population Dynamics and Insecticide Susceptibility of Anopheles culicifacies in Malaria Endemic Districts of Chhattisgarh, India.

A study was undertaken in the villages of Korea and Bastar district (Chhattisgarh) during the years 2012-2015 to investigate the bionomics of malaria vectors and the prevalence of their sibling species complexes. Entomological surveys carried out every month included indoor resting collections, pyrethrum spray catches, light trap catches, and insecticide susceptibility status of Anopheles culicifacies using World Health Organization (WHO) methods. Anopheles culicifacies and Anopheles fluviatilis species were assayed by polymerase chain reaction (PCR) for the detection of malaria parasite, and sibling species were identified using PCR and DNA sequencing. A total of 13,186 samples of Anopheles comprising 15 species from Bastar and 16 from Korea were collected. An. Culicifacies was recorded as the most dominant species and also the only active vector at both sites. This species was found to be resistant to dichlorodiphenyltrichloroethane (DDT) and Malathion, showing signs of emerging resistance against pyrethroids. Among the sibling species of An. culicifacies, the group BCE was found in maximum numbers, while sibling species T of the An. fluviatilis was recorded to be dominant among its complex. The study provides a comprehensive view of the vector bionomics in the highly malarious regions of India that may have importance in developing vector control strategies.