Development of SCAR markers for specific identification of B-biotype of Bemisia tabaci (Gennadius).

摘要

B-biotype of cotton whitefly, Bemisia tabaci (Bemisia argentifolii) due to its wide spreadability, high fecundity, polyphagous nature, fast adaptability to insecticides and being vector of many geminiviruses had emerged as a pest of quarantine significance with a negative impact on the economy of agricultural countries. Thus, monitoring of whitefly populations for B-biotype required quick and reliable identification methods. To develop B-biotype sequence-specific molecular markers, three polymorphic RAPD-DNA bands were identified in the RAPD-PCR profiles of B-biotype relative to non-B-biotype variants of B. tabaci. Based upon the sequence data of these polymorphic RAPD-DNA fragments, three pairs of sequence specific amplified region (SCAR) primers were developed for molecular identification of B-biotype through specific PCR amplification. When used in PCR amplification from different B-biotype and non-B-biotype variants, two of these SCAR primer pairs (BbF07-573_FR and BbA01-970_FR) effectively amplified a single fragment of predetermined size (480 and 938 bp, respectively) from all the B-biotype variants only (Australia, Israel and B-biotype variant-1, -2 and -3, identified in India). The third SCAR primer pair (BbF16-269_FR) through amplification of the expected 239-bp fragment similarly showed its specificity for all the other B-biotypes, but it was not specific for an Indian B-biotype-3 variant. SCAR primers were equally effective in molecular identification of B-biotype in a multiplex PCR by co-amplification of both SCAR markers in a single PCR. Analysis of 150 populations of 1351 whitefly individuals collected from nine different states of India with two SCAR primer pairs in a multiplex PCR established, whereas B-biotype was predominating type in south Indian states, this biotype has started contaminating native whitefly populations of Gujarat and Maharashtra. However, all North Indian states were free from this biotype. Molecular monitoring of whitefly populations for B-biotype with the SCAR primers through multiplex-PCR developed in this study, will help in studying the migration pattern of B-biotype with respect to the season, host crop and region for adopting appropriate control measures which were different from those found for control of non-B-biotype of whitefly and in its detection for quarantine controls.