KASP Genotyping as a Molecular Tool for Diagnosis of Cassava-Colonizing Bemisia tabaci

Abstract

<i>Bemisia tabaci</i> is a cryptic species complex that requires the use of molecular tools for identification. The most widely used approach for achieving this is the partial sequencing of the mitochondrial DNA cytochrome oxidase I gene (<i>COI</i>). A more reliable single nucleotide polymorphism (SNP)-based genotyping approach, using Nextera restriction-site-associated DNA (NextRAD) sequencing, has demonstrated the existence of six major haplogroups of <i>B. tabaci</i> on cassava in Africa. However, NextRAD sequencing is costly and time-consuming. We, therefore, developed a cheaper and more rapid diagnostic using the Kompetitive Allele-Specific PCR (KASP) approach. Seven sets of primers were designed to distinguish the six <i>B. tabaci</i> haplogroups based on the NextRAD data. Out of the 152 whitefly samples that were tested using these primer sets, 151 (99.3%) produced genotyping results consistent with NextRAD. The KASP assay was designed using NextRAD data on whiteflies from cassava in 18 countries across sub-Saharan Africa. This assay can, therefore, be routinely used to rapidly diagnose cassava <i>B. tabaci</i> by laboratories that are researching or monitoring this pest in Africa. This is the first study to develop an SNP-based assay to distinguish <i>B. tabaci</i> whiteflies on cassava in Africa, and the first application of the KASP technique for insect identification.