Molecular Identification of Ten Economically Important Fruit Flies (Diptera: Tephritidae) of Bangladesh by Using PCR-RFLP

Abstract

Proper identification of insect pests at species level is considered as the first step towards implementing successful control strategies against their dispersal. In this context, molecular techniques are best used to support morphological identification. PCR-RFLP is a rapid tool for differentiating tephritid fruit fly pests. In the present study, an initiative was taken to identify ten economically important dacine fruit flies using a PCR amplification technique to successfully establish RFLP patterns of the Internal Transcribed Spacer 1 (ITS1) region. The specific primer pair baITS1f/ baITS1r was used to amplify ∼520-840 base pair long fragment of the ITS1 region. The amplified fragments were varied in size among species and able to differentiate at genus level from Bactrocera to Zeugodacus. But interspecies variations are not clearly distinguished by ITS1 banding profile alone. Due to the ambiguity in band size, they were then subjected to RFLP digestion with two restriction endonuclease enzymes (HhaI and Sau3aI). The restriction enzymes have different cutting sites and thus displayed different banding patterns among species. Analysis of restriction fragments of the ITS1 is able to distinguish six of the ten species successfully. In addition, two morphologically similar species Bactrocera dorsalis (Hendel) and Bactrocera carambolae (Drew & Hancock) were differentiated by restriction fragments of ITS1. The outcome of this study will enhance early detection and easy monitoring of these quarantine pest species at the port of entry.