A DNA barcode library for katydids, cave crickets, and leaf-rolling crickets (Tettigoniidae, Rhaphidophoridae and Gryllacrididae) from Zhejiang Province, China

Abstract

Barcode libraries are generally assembled with two main objectives in mind: specimen identification and species discovery/delimitation. In this study, the standard COI barcode region was sequenced from 681 specimens belonging to katydids (Tettigoniidae), cave crickets (Rhaphidophoridae), and leaf-rolling crickets (Gryllacrididae) from Zhejiang Province, China. Of these, four COI-5P sequences were excluded from subsequent analyses because they were likely NUMTs (nuclear mitochondrial pseudogenes). The final dataset consisted of 677 barcode sequences representing 90 putative species-level taxa. Automated cluster delineation using the Barcode of Life Data System (BOLD) revealed 118 BINs (Barcodes Index Numbers). Among these 90 species-level taxa, 68 corresponded with morphospecies, while the remaining 22 were identified based on reverse taxonomy using BIN assignment. Thirteen of these morphospecies were represented by a single barcode (so-called singletons), and each of 19 morphospecies were split into more than one BIN. The consensus delimitation scheme yielded 55 Molecular Operational Taxonomic Units (MOTUs). Only four morphospecies (I _max > DNN) failed to be recovered as monophyletic clades (i.e., Elimaeaterminalis, Phyllomimusklapperichi, Sinochloraszechwanensis and Xizicushowardi), so it is speculated that these may be species complexes. Therefore, the diversity of katydids, cave crickets, and leaf-rolling crickets in Zhejiang Province is probably slightly higher than what current taxonomy would suggest.

Keywords: Barcode Index Number; Ensifera; Orthoptera; cryptic species; species delimitation.

Figure 1.

Sampling coverage of katydids, cave crickets, and leaf-rolling crickets in Zhejiang Province, China.

Figure 2.

BIN discordance and the problems of interpreting potential NUMTs A–E represent the five cases of BIN discordance and individuals in red font represent individuals with potential NUMTs.

Figure 3.

Scatter plot of maximum intraspecific distance and distance to nearest neighbour (NN). The four species to the right of the line represent a large intraspecific genetic distance.

Figure 4.

Examples of species split into more than one BIN A–D represent Sinochloraszechwanensis split into 2 BINs, Phyllomimusklapperichi split into 3 BINs, Elimaeaterminalis split into 2 BINs, and Xizicushowardi split into 3 BINs.

Figure 5.

BOLD TaxonID Tree based on K2P distances and species delimitation results based on COI-5P sequences. The four barcode sequences marked by red are highly likely NUMTs and excluded from the species delimitation analyses. The MOTUs created by each delimitation algorithm are represented as squares on the right. The number within the rectangles indicates the number of MOTUs; no number indicates a single MOTU.