Evaluation of DNA barcoding and identification of new haplomorphs in Canadian deerflies and horseflies

Abstract

This paper reports the first tests of the suitability of the standardized mitochondrial cytochrome c oxidase subunit I (COI) barcoding system for the identification of Canadian deerflies and horseflies. Two additional mitochondrial molecular markers were used to determine whether unambiguous species recognition in tabanids can be achieved. Our 332 Canadian tabanid samples yielded 650 sequences from five genera and 42 species. Standard COI barcodes demonstrated a strong A + T bias (mean 68.1%), especially at third codon positions (mean 93.0%). Our preliminary test of this system showed that the standard COI barcode worked well for Canadian Tabanidae: the target DNA can be easily recovered from small amounts of insect tissue and aligned for all tabanid taxa. Each tabanid species possessed distinctive sets of COI haplotypes which discriminated well among species. Average conspecific Kimura two-parameter (K2P) divergence (0.49%) was 12 times lower than the average divergence within species. Both the neighbour-joining and the Bayesian methods produced trees with identical monophyletic species groups. Two species, Chrysops dawsoni Philip and Chrysops montanus Osten Sacken (Diptera: Tabanidae), showed relatively deep intraspecific sequence divergences (∼ 10 times the average) for all three mitochondrial gene regions analysed. We suggest provisional differentiation of Ch. montanus into two haplotypes, namely, Ch. montanus haplomorph 1 and Ch. montanus haplomorph 2, both defined by their molecular sequences and by newly discovered differences in structural features near their ocelli.

Identification through DNA barcoding of Tabanidae (Diptera) vectors of surra disease in India

“…DNA barcode dataset of this study depicted biasness towards A + T content (68%) richness which was also supported by the A or T termination of the triplet codon in the current dataset. This is in accord with earlier studies on Tabanidae (Cywinska et al, 2010). A + T codon use biasness seems to be consistent in class Insecta mitochondrial genes (Hoy, 2006;Lunt et al, 1996).…”

“…A + T codon use biasness seems to be consistent in class Insecta mitochondrial genes (Hoy, 2006;Lunt et al, 1996). The G + C content in the current dataset was found to be (32%) in the range of values from an earlier study on tabanids (Cywinska et al, 2010) and the class Insecta as a whole (Clare et al, 2008).…”

“…A further fifty two sequences for these four genera from a current study (Cywinska et al, 2010) were added to the dataset for NJ tree construction. A total of ninety eight sequences were available in the final dataset for further analysis.…”

“…Forty six DNA sequences generated in this study were aligned against representative sequence of tabanids from early study (Cywinska et al, 2010) from Canada as retrieved from BOLD (Barcode of Life Data System) under the project Tabanidae of Canada (MHTAB) (http://www.boldsystems.org/index.php/ MAS Management RecordList). These sequence were manually screened and edited with MEGA6.0 (Tamura et al, 2013).…”

“…However, similar studies on tabanids are largely wanting. There are a few molecular studies related to higher level phylogenetic relationships in this family (Wiegmann et al, 2000(Wiegmann et al, , 2003Winterton et al, 2007 Yeates andWiegmann, 1999), but the utility of DNA barcoding has been evaluated for species discrimination only for the Canadian tabanid fauna (Cywinska et al, 2010). However, effectiveness of DNA barcoding in the discrimination of closely related pestiferous species of Tabanidae has not previously been attempted.…”