Molecular Delimitation And Integrative Taxonomy Of Gampsocleis From China

The genus Gampsocleis Fieber,1852 belongs to the Orthoptera,Tettigoniidae.So far,the genus includes 17 species in the world and 11 species from China.Most species descriptions for this genus were published earlier and described simply,bringing about difficulties for morphological species delimitation.Based on the mitochondrial COI(DNA barcode standard sequences)and CytB gene sequences,this study used five single-locus species delimitation methods such as BIN,ABGD,jMOTU,GMYC and bPTP to delimit species boundaries and interspecific relationships,integrating with morphological characterisation and the acoustic.In this study,a total of 536 specimens of 6 species in Gampsocleis were initially identified as G.gratiosa,G.sedakovii,G.ussuriensis,G.sinensis,G.carinata and G.glabra.3 specimens of U.inflatus were selected as the outgroup.The results are as follows:1.536 COI and 487 CytB sequences were sequenced,and 264 and 237 haplotypes were obtained after repeated removal for subsequent analysis.The COI and CytB sequences of 3 U.inflatus specimens were obtained,respectively,3 and 2 haplotypes were obtained.2.The Neighbor-joining(NJ)tree under the Kimura 2-parameter(K2P)model and five species delimitation methods supported the taxonomic status of G.glabra.3.Multiple methods could not effectively separate G.ussuriensis from G.sinensis.After detailed examination of the specimens,there were not significant differences in morphological characteristics.G.sinensis was compared with the holotype in detail to determine that the G.sinensis specimens were identified correctly.Therefore,the G.ussuriensis specimens in this study is the wrong identification of G.sinensis.It is speculated that G.ussuriensis from northern China should be G.sinensis.4 The NJ trees constructed by COI and CytB data divided G.sedakovii into two clusters.Among them,2 clusters were composed large monophyly based on COI data;based CytB,there was a sister group that G.sedakovii sedakovii clustered with G.sinensis.Examinating the specimens of the two clusters,there were some differences in titillators and cerci of male and subgenital plate of female.However,based on the previous morphological classification(tegmen length,the ratio between tegmen length and pronotum),the two clusters could not be completely separated.This study supported the division of G.sedakovii sedakovii and G.sedakovii obscura,but the morphological basis for the division still need to be further explored.5.Different species delimitation methods divided G.carinata into 1–5 OTUs.Based on the NJ trees constructed by COI,respectively,specimens come from Gansu and Shanxi form two monophyly clusters;while based on CytB,there was no monophyly cluster of the specimens which come from Shanxi.Examining the specimens,there was no stable differences in morphological characterisation.The acoustic analysis differed in the duration of the syllables and impulse.6.Based on multiple methods of COI and CytB data,G.gratiosa was separated from Shandong and other geographical regions.Examining the specimens,there were some significant differences in the titillators and cerci.The acoustic analysis differed in the duration of the syllables and impulse.Therefore,this study suggested that the taxonomic status of G.fletcheri should be recovered.This study found that short-winged insects with weaker diffusion ability are more susceptible to habitat fragmentation.It is difficult to effectively delimite based on DNA molecular delimitation methods only.Integrating multiple data for species definition is necessary.Therefore,this study integrated three data and combined with geographical distribution.Gampsocleis from China were finally divided into 6 species(subspecies),namely: G.sinensis,G.sedakovii sedakovii,G.sedakovii obscura,G.carinata,G.gratiosa and G.fletcheri.