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

Abstract

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.

Half of the European fruit fly species barcoded (Diptera, Tephritidae); a feasibility test for molecular identification

“…The threshold value in BCM is of strong influence on the results, as already noted by Virgilio et al (2012). The success-rates have been calculated for a range of arbitrary threshold values between the largest observed distance and 0.00 (Figure 5).…”

Description of new Ceratitis species (Diptera: Tephritidae) from Africa, or how morphological and DNA data are complementary in discovering unknown species and matching sexes

“…Female terminalia were dissected, cleared in potassium hydroxide solution and temporarily mounted in glycerinegelatine for drawings made by camera lucida. a set of Dna sequences were produced for part of the mitochondrial Cytochrome c oxidase subunit i (Coi) and deposited in the Barcode of life Data Systems (BolD, http://www.boldsystems.org) as reference DNA barcodes for the molecular identification of the new species (see Virgilio et al 2012 for laboratory protocols and discussion). Kimura's two parameter genetic distances (Kimura 1980) between the newly generated Ceratitis Dna barcodes and a selection of 367 reference Dna barcodes deposited in BolD (corresponding to 51 species from all six Ceratitis subgenera) were visualized in a neighbor joining tree.…”

DNA barcodes for bio-surveillance: regulated and economically important arthropod plant pests

“…For example, 92.2% of 3514 species of European beetles were assigned to a distinct BIN that coincided with a known morphological species, while most of the other species were assigned to two or three BINs, suggesting they represent cryptic species complexes (Hendrich et al 2015). Work on Diptera has similarly validated the effectiveness of DNA barcoding for the identification of species in this order, including some of the most important agricultural pests, such as fruit flies (Virgilio et al 2012) and leafminers (Amin et al 2014). Another study that examined 1849 species of Canadian Hemiptera assigned these species to 1867 BINs with high correspondence between species and BINs, but also revealed 27 species with high divergences suggestive of cryptic species complexes (Gwiazdowski et al 2015).…”