Abstract

BackgroundRapid and reliable identification of quarantine pests is essential for plant inspection services to prevent introduction of invasive species. For insects, this may be a serious problem when dealing with morphologically similar cryptic species complexes and early developmental stages that lack distinctive characters useful for taxonomic identification. DNA based barcoding could solve many of these problems. The standard barcode fragment, an approx. 650 base pairs long sequence of the 5′end of the mitochondrial cytochrome oxidase I (COI), enables differentiation of a very wide range of arthropods. However, problems remain in some taxa, such as Tephritidae, where recent genetic differentiation among some of the described species hinders accurate molecular discrimination.ResultsIn order to explore the full species discrimination potential of COI, we sequenced the barcoding region of the COI gene of a range of economically important Tephritid species and complemented these data with all GenBank and BOLD entries for the systematic group available as of January 2012. We explored the limits of species delimitation of this barcode fragment among 193 putative Tephritid species and established operational taxonomic units (OTUs), between which discrimination is reliably possible. Furthermore, to enable future development of rapid diagnostic assays based on this sequence information, we characterized all single nucleotide polymorphisms (SNPs) and established “near-minimal” sets of SNPs that differentiate among all included OTUs with at least three and four SNPs, respectively.ConclusionsWe found that although several species cannot be differentiated based on the genetic diversity observed in COI and hence form composite OTUs, 85% of all OTUs correspond to described species. Because our SNP panels are developed based on all currently available sequence information and rely on a minimal pairwise difference of three SNPs, they are highly reliable and hence represent an important resource for developing taxon-specific diagnostic assays. For selected cases, possible explanations that may cause composite OTUs are discussed.

Highlights

  • Rapid and reliable identification of quarantine pests is essential for plant inspection services to prevent introduction of invasive species

  • The main aim of the present study was to use all sequence information currently available to define operational taxonomic units (OTUs) that reflect the maximum possible level of species discrimination provided by the cytochrome oxidase I (COI) gene, and to describe nearminimal sets of single nucleotide polymorphisms (SNPs) that enable discrimination between all pairwise comparisons of these Operational taxonomic unit (OTU) with at least three or four Single nucleotide polymorphism (SNP) difference

  • We performed our script-based search with the entire data set of 202 OTUs with the threshold “Including Singletons” and we found several “near-minimal” SNP panels composed of 113 SNPs for the four and composed of 81 SNPs for the three base pair differentiation, respectively

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Summary

Introduction

Rapid and reliable identification of quarantine pests is essential for plant inspection services to prevent introduction of invasive species. This may be a serious problem when dealing with morphologically similar cryptic species complexes and early developmental stages that lack distinctive characters useful for taxonomic identification. DNA barcoding has been developed as a powerful identification tool to contribute in the prevention of introduction, establishment and spread of invasive species [3]. The four genera of greatest economic importance (Anastrepha, Bactrocera, Ceratitis and Rhagoletis) include morphologically similar sibling species, complexes of cryptic species, host races, and morphotypes [11,12,13,14,15,16,17]. Since quarantine interceptions mostly contain larvae concealed within fruit, it is important to develop tools for accurate identification of immature life stages of fruit flies

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