Diagnostic Tool for the Identification of Bactrocera dorsalis (Hendel) (Diptera: Tephritidae) Using Real-Time PCR

The accurate identification of the Oriental fruit fly, Bactrocera dorsalis, is complicated by its similarities to other species and taxonomic uncertainties. This represents a significant threat to fruit crops as it is already present in Europe, and this is a cause for great concern. To expedite identification, a reliable method using a unique technical approach was developed. The initial phase involved collecting specimens from the population present in Italy to create a large and representative sample, enabling us to optimize the method. This method has demonstrated high sensitivity and accuracy in detecting small amounts of B. dorsalis DNA. It now serves as a valuable tool for routine diagnostics, facilitating efficient pest management and detection. Given the recent infestations in Italy, this diagnostic protocol is crucial for monitoring and preventing the passive spread of B. dorsalis in Europe. Accurate identification of Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), commonly known as the Oriental fruit fly, is a significant challenge due to the morphological convergence and taxonomic uncertainties of species belonging to the same genus. This highly polyphagous species poses a significant threat to fruit crops. With its potential establishment in Europe becoming a growing concern, there is an urgent need for rapid and efficient diagnostic methods. The study presented here introduces a diagnostic protocol based on real-time PCR using a TaqMan probe for the early and reproducible identification of B. dorsalis. Specimens representing the genetic diversity of the Italian population were collected and analyzed. Specific primers and probe were designed based on the conserved regions and an in silico analysis confirmed their specificity. The assay conditions were optimized, and analytical sensitivity, specificity, repeatability, and reproducibility were evaluated. The protocol showed high sensitivity and specificity, accurately detecting low DNA concentrations of B. dorsalis. This standardized method provides a reliable tool for routine diagnostics, enhancing the accuracy and efficiency of identifying the Oriental fruit fly at all stages of its development, thereby facilitating effective pest management measures. The development of this diagnostic protocol is crucial for monitoring and supporting efforts to prevent the passive spread of B. dorsalis in Europe, particularly in light of the recent active infestations detected in Italy.