Rapid identification of Bactrocera zonata (Dip.: Tephritidae) using TaqMan real-time PCR assay

Abstract

Tephritid fruit flies are ranked as one of the most damaging groups of insect pests. Morphological identification of fruit flies is mainly performed on adults due to the lack of adequate identification keys for immature stages. The peach fruit fly, Bactrocera zonata (Saunders), infests some of the principal commercial fruits and vegetables. It is, therefore important to avert its global dispersal, particularly by accurately identifying this species at ports of entry. In this study, a TaqMan real-time polymerase chain reaction (PCR) was developed for the accurate identification and sensitive detection of the peach fruit fly. A novel set of primers and probe were designed to specifically identify the mitochondrial cytochrome oxidase I (COI) gene. All specimens of peach fruit fly (including various life stages) were detected, and no cross reactivity with other tested tephritids were observed. Since this assay performed equally well with crushed insects and purified DNA, we note added efficiency by eliminating DNA extraction step. Considering the speed, specificity as well as sensitivity of the assay, Taqman real-time PCR can be used as a swift and specific method for pest species at ports of entry.

Development of a loop-mediated isothermal amplification (LAMP) assay for the identification of the invasive wood borer Aromia bungii (Coleoptera: Cerambycidae) from frass

“…Ten samples in triplicate, diluted to a final concentration of 5 ng/µL, were tested in two separate series for repeatability. The reproducibility of each protocol was assessed in the same way as carried out for the repeatability by comparing the data of two series of samples by two different operators on different days (Dhami et al 2016 ; Koohkanzade et al 2018 ).…”

Identification of the Red-Necked Longhorn Beetle Aromia bungii (Faldermann, 1835) (Coleoptera: Cerambycidae) with Real-Time PCR on Frass

“…To estimate the repeatability, we tested ten samples in triplicate in two separate runs and calculated the % CV for each sample. To estimate reproducibility, the data of the two runs were compared [37,38].…”

Molecular Identification of Anoplophora glabripennis (Coleoptera: Cerambycidae) From Frass by Loop-Mediated Isothermal Amplification

“…For the repeatability test, 10 samples diluted to 5 ng/µl, run in triplicate and in two separate series, were used. The mean values, SD, and % CV were calculated for each sample and for each series of samples (Dhami et al 2016, Koohkanzade et al 2018. The reproducibility for each protocol was tested in the same way as the repeatability test but with two different operators, on different days, and comparing the data of the two sample series.…”