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.