RESUMEN
P-glycoprotein [P-gp or the ATP-binding cassette transporter B1 (ABCB1)] is an important participant in multidrug resistance of cancer cells, yet the precise function of this arthropod transporter is unknown. The aim of this study was to determine the importance of P-gp for susceptibility to insecticides in the beet armyworm (Spodoptera exigua) using clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) gene-editing technology. We cloned an open reading frame (ORF) encoding the S. exigua P-gp protein (SeP-gp) predicted to display structural characteristics common to P-gp and other insect ABCB1 transporters. A knockout line with a frame shift deletion of four nucleotides in the SeP-gp ORF was established using the CRISPR/Cas9 gene-editing system to test its potential role in determining susceptibility to chemical insecticides or insecticidal proteins from the bacterium Bacillus thuringiensis (Bt). Results from comparative bioassays demonstrate that knockout of SeP-gp significantly increases susceptibility of S. exigua by around threefold to abamectin and emamectin benzoate (EB), but not to spinosad, chlorfenapyr, beta-cypermethrin, carbosulfan indoxacarb, chlorpyrifos, phoxim, diafenthiuron, chlorfluazuron, chlorantraniliprole or two Bt toxins (Cry1Ca and Cry1Fa). Our data support an important role for SeP-gp in susceptibility of S. exigua to abamectin and EB and imply that overexpression of SeP-gp may contribute to abamectin and EB resistance in S. exigua.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Proteínas de Insectos/genética , Resistencia a los Insecticidas/genética , Insecticidas/farmacología , Spodoptera/fisiología , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Animales , Bacillus thuringiensis/química , Toxinas de Bacillus thuringiensis , Proteínas Bacterianas/farmacología , Sistemas CRISPR-Cas , Resistencia a Múltiples Medicamentos/genética , Endotoxinas/farmacología , Técnicas de Inactivación de Genes , Proteínas Hemolisinas/farmacología , Proteínas de Insectos/metabolismo , Ivermectina/análogos & derivados , Ivermectina/farmacología , Larva/genética , Larva/crecimiento & desarrollo , Larva/fisiología , Spodoptera/genética , Spodoptera/crecimiento & desarrolloRESUMEN
The methodology combining Axisymmetric Drop Shape Analysis (ADSA) with a captive bubble (ADSA-CB) facilitates pulmonary surfactant related studies. The accuracy of ADSA-CB is crucially dependent on the quality of the bubble profile extracted from the raw image. In a previous paper, an image analysis scheme featuring a Canny edge detector and a Axisymmetric Liquid Fluid Interfaces-Smoothing (ALFI-S) algorithm was developed to process captive bubble images under a variety of conditions, including images with extensive noise and/or lack of contrast. A new version of ADSA-CB based on that image analysis scheme is developed and applied to pulmonary surfactant and pulmonary surfactant-polymer systems. The new version is found to be highly noise-resistant and well self-adjusting.