Identifying Insects with Incomplete DNA Barcode Libraries, African Fruit Flies (Diptera: Tephritidae) as a Test Case

Royal Museum for Central Africa, Tervuren, Belgium.
PLoS ONE (Impact Factor: 3.23). 02/2012; 7(2):e31581. DOI: 10.1371/journal.pone.0031581
Source: PubMed


We propose a general working strategy to deal with incomplete reference libraries in the DNA barcoding identification of species. Considering that (1) queries with a large genetic distance with their best DNA barcode match are more likely to be misidentified and (2) imposing a distance threshold profitably reduces identification errors, we modelled relationships between identification performances and distance thresholds in four DNA barcode libraries of Diptera (n = 4270), Lepidoptera (n = 7577), Hymenoptera (n = 2067) and Tephritidae (n = 602 DNA barcodes). In all cases, more restrictive distance thresholds produced a gradual increase in the proportion of true negatives, a gradual decrease of false positives and more abrupt variations in the proportions of true positives and false negatives. More restrictive distance thresholds improved precision, yet negatively affected accuracy due to the higher proportions of queries discarded (viz. having a distance query-best match above the threshold). Using a simple linear regression we calculated an ad hoc distance threshold for the tephritid library producing an estimated relative identification error <0.05. According to the expectations, when we used this threshold for the identification of 188 independently collected tephritids, less than 5% of queries with a distance query-best match below the threshold were misidentified. Ad hoc thresholds can be calculated for each particular reference library of DNA barcodes and should be used as cut-off mark defining whether we can proceed identifying the query with a known estimated error probability (e.g. 5%) or whether we should discard the query and consider alternative/complementary identification methods.


Available from: Massimiliano Virgilio
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    • "In these conditions, minibarcodes have proven to be very successful (Sundquist et al. 2007), so we identified potential minibarcodes for feather mites and explored their efficacy. 2 The use of thresholds to differentiate species has been repeatedly discussed in the DNA barcoding literature, finding that no single threshold is optimal for all species (Puillandre et al. 2012; Virgilio et al. 2012; Collins & Cruickshank 2013). Moreover, the accuracy of a threshold-based approach critically depends upon the level of overlap between intra-and interspecific variation across a phylogeny (Meyer & Paulay 2005). "
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    ABSTRACT: Feather mites (Astigmata: Analgoidea, Pterolichoidea) are among the most abundantand commonly occurring bird ectosymbionts. Basic questions on the ecology and evolution of feather mites remain unanswered because feather mite species identification is often only possible for adult males and it is laborious even for specialised taxonomists, thus precluding large-scale identifications. Here, we tested DNA barcoding as a useful molecular tool to identify feather mites from passerine birds. 361 specimens of 72 species of feather mites from 68 species of European passerine birds from Russia and Spain were barcoded. The accuracy of barcoding and mini-barcoding was tested. Moreover, threshold choice (a controversial issue in barcoding studies) was also explored in a new way, by calculating through simulations the effect of sampling effort (in species number and species composition) on threshold calculations. We found one 200 bp mini-barcode region that showed the same accuracy as the full-length barcode (602 bp) and was surrounded by conserved regions potentially useful for group-specific degenerate primers. Species identification accuracy was perfect (100%) but decreased when singletons or species of the Proctophyllodes pinnatus group were included. In fact, barcoding confirmed previous taxonomic issues within the Proctophyllodes pinnatus group. Following an integrative taxonomy approach, we compared our barcode study with previous taxonomic knowledge on feather mites, discovering three new putative cryptic species and validating three previous morphologically different (but still undescribed) new species.
    Molecular Ecology Resources 02/2015; 15(5). DOI:10.1111/1755-0998.12384 · 3.71 Impact Factor
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    • "DNA barcoding has been proposed as a relatively rapid and effective tool for the identification of fruit flies (Armstrong and Ball 2005). Yet, despite the availability of relatively large reference libraries of DNA barcodes for tephritid fruit flies, this method is still not routinely used for identification mainly due to shortcomings such as the difficulty of resolving important species complexes (Smit et al. 2013; Virgilio et al. 2010) and the incompleteness of reference libraries (Virgilio et al. 2012). "

    ZooKeys 07/2014; 428(428):97-108. DOI:10.3897/zookeys.428.7366 · 0.93 Impact Factor
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    • "First, species delimitation simply based on genetic divergence is difficult to convey and interpret in a " universally acceptable " species concept (Krishnamurthy and Francis 2012). Second, large intraspecific distances and low interspecific distances among closely related species may pose a major problem, and even the use of refined criteria such as best close match (Meier et al. 2006) or ad hoc thresholds (Virgilio et al. 2012) might not solve this issue. Therefore, in datasets where intra-and interspecific divergences largely overlap, using distance-based thresholds alone may not work. "

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