GENE EXPRESSION STUDY OF COMPOUND (DEHB) TREATED LARVAE ISOLATED FROM THE Streptomyces rimosus AGAINST THE FUNCTIONAL GENES OF Culex quinquefasciatus Say (Diptera:Culicidae)

The present study is aimed at checking the effect of compound (di-2-(ethyl hexyl) phthlate) isolated from Streptomyces rimosus on the expression of the genes from mosquitoes, namely, Cytochrome P450 (CYP450), glutathione S-transferase (GST), Esterase (EST) and Beta-actin (ACTIN) using RT-PCR and qTPCR. Nucleotide sequences of GST, EST, CYP and actin genes from Cx. quinquefasciatus were extracted from NCBI database. Primers were designed in the conserved regions of mosquito nucleotide sequence using Primer3 Input web tool. Mosquitoes were treated with the compound isolated from Streptomyces rimosus. Twenty early third instar larvae of Cx. quinquefasciatus were introduced into the containers. Concentrations of 2.0 ppm of the compound were prepared using dimethyl sulfoxide (DMSO) (249 ml water and 1 ml DMSO); five replicates were maintained for treated concentration. Total RNA was extracted using RNeasy kit, according to the manufacturer’s instructions. Total RNA was converted into cDNA by using QuantiTect Reverse Transcription kit. The RT- PCR reaction was on the final volume of 20 μl. The reactions were performed under standardized qRT-PCR conditions by using gene specific primers. The results indicated that EST gene showed downregulation when compared to the standard controls. In addition GST gene showed differential expression levels and CYP 450 showed no variation in expression when compared to the controls. As per previous studies, EST and GST genes are responsible for insecticide resistance, but in our study the compound-treated larvae showed some up- and downregulation of genes in EST and GST. The present study clearly states that the isolated compound DEHP from S. rimosus may be used to disrupt the resistance development of Cx. quinquefasciatus against insecticides.