Background: On the basis of molecular dating, Pleistocene glaciations have been proposed as the major driving force of biota speciation in the Palearctic and the pre-Quaternary origin of Amazonian taxa. However, the major driving factors in East Asia remain unclear. All 16 saturniine species inhabiting Taiwan with congeners of populations, subspecies, or species in East Asia constitute research objects for addressing the mode of speciation because of the repeated formation and disappearance of a landbridge from the Asian mainland to Taiwan during glacial cycles. Methods: The genetic divergences of mitochondrial cytochrome c oxidase subunit I (COI) and 16S rDNA and the nuclear 28S rDNA of the saturniine species from Taiwan and the Asian mainland were assessed to determine the monophyly of each genus and species of Saturniinae. Moreover, 519 saturniine COI sequences of 114 taxa from adjacent East and Southeast Asian populations and closely related species were retrieved from GenBank and analyzed. The differentiation timing and possible origination of the insular saturniines were elucidated based on phylogenetic relationships, haplotype networks, and lineage calibrations. Results: Approximately 90% of intraspecific COI divergence was <2%; all divergences exceeding 2% originated from comparisons between allopatric populations or subspecies. Relationship analyses revealed that multiple introductions likely occurred in insular saturniines and that some East Asian saturniines were paraphyletic as deduced by analyzing endemic insular species. Calibration dating revealed that Taiwanese endemic saturniines split from sibling Asian species 0.2–2.7 million years ago (Mya), whereas subspecific-level and population-level splitting events occurred 0.1–1.7 Mya and 0.2–1.2 Mya, respectively. Moreover, phylogenetic patterns combined with geographical distributions revealed that hill-distributed Taiwanese saturniines are closely related to those from southern China and Southeast Asia, whereas saturniines inhabiting altitudes higher than 1,500 m in Taiwan have siblings distributed in temperate Northeast Asia. Discussion: The Global DNA Barcoding Initiative was successfully applied to study the population genetic structure in species. Most Formosan saturniines are distinct and monophyletic, reflecting the vicariant barrier of the Taiwan Strait; Pleistocene glacial cycles provided opportunities for insular saturniines to experience repeated isolation from and secondary contact with the continental mainland. Each insular saturniine may have evolved with a unique differentiation timing pattern that possibly emerged in the Early, Middle, or Late Pleistocene with these patterns differing from the consistent pattern that occurred in the temperate Palearctic and tropical Amazonian regions. Moreover, multiple migrations or artificial genetic admixtures may have also occurred, as suggested by the coexistence of two divergent lineages in a few Taiwanese saturniines.

Taxonomic debates have been carrying on for decades over Formosan stag beetles, which consist of a high proportion of endemic species and subspecies featuring morphological variations associated with local adaptation. With the influence of periodical Pleistocene glaciations and the presence of several mountain ranges, the genetic differentiation and taxonomic recognition, within this medium-size island, of two endemic subspecies for each of four montane stag beetles, i.e. Lucanus ogakii, L. kanoi, Prismognathus davidis, and Neolucanus doro, has been an appealing issue. Based on monophyletic lineages and population structure, possible divergent scenarios have been proposed to clarify the subspecific status for each of the above mentioned stag beetles. Phylogenetic inferences based on COI+16S rDNA+28S rDNA of 240 Formosan lucanids have confirmed most species are monophyletic groups; and the intraspecific (2%) genetic distances of the two mitochondrial genes could be applied concordantly for taxonomic identification. On account of Bayesian-based species delimitation, geographic distribution, population structure, and sequence divergences, the subspecific status for L. ogakii, L. kanoi, and Pri. davidis are congruent with their geographic distribution in this island; and the calibration time based on the mitochondrial genes shows the subspecific split events occurred 0.7–1 million years ago. In addition, a more complicated scenario, i.e. genetic differentiation including introgression/hybridization events, might have occurred among L. ogakii, L. kanoi, and L. maculifemoratus. The geological effects of mountain hindrance accompanied by periodical glaciations could have been vital in leading to the geographical subspecific differentiation of these montane stag beetles.

Plecia longiforceps Duda (Diptera: Bibionidae) is reported for the first time from Korea. P. longiforceps has been previously known from the East Asian subtropics, south of the 33rd parallel, including southeastern China, Taiwan, and the Ryukyu Islands of Japan. An integrative taxonomic approach based on morphological examination of male genitalia and molecular analysis of mitochondrial cytochrome c oxidase subunit I gene sequences confirms the species identification. The recent outbreak of P. longiforceps in the Seoul Metropolitan Area, Korea, documented herein represents the northernmost record of this species and suggests its possible range expansion into the temperate zone. Similar to the range expansion and outbreak history of Plecia nearctica Hardy (Diptera: Bibionidae) in North America around the Gulf of Mexico in the 1960–1970s, P. longiforceps may become a new invasive pest in temperate East Asia. Here, we evaluate range expansion and invasion potential of P. longiforceps through Ensemble species distribution modeling and show that a great portion of Northeast Asia and Japan will likely become habitable for P. longiforceps in the next 50 years.

The present study demonstrates that the complicated genetic structure of the hilly lineage of the Neolucanus swinhoei complex was driven by its biological features and habitat requirements as well as hindrance by the CMR during periodical Pleistocene glaciations. The results revealed a tendency of geographical differentiation and major and sub- lineage divergences before and after the Riss glaciation, followed by stable population growth during Würm glaciation. At least four refugia were inferred for N. swinhoei during the Riss–Würm glaciations. The ancestral haplotype retention in the cytochrome oxidase subunit I (COI) gene and compensated substitution in 16S rRNA gene is a possible evolutionary scenario resulting in the inconsistent evolution pattern between COI and 16S rRNA gene coupled with the long-distance dispersal of N. swinhoei. Although the CMR did hinder the dispersal of N. swinhoei, its ancestors may have dispersed to eastern Taiwan through the northern and southern low mountains of the CMR before the Riss glaciation. Our finding suggests that the population growth in the Würm glaciation led a dispersal back to western Taiwan, which is contrast to the more common dispersal scenario from western Taiwan to eastern populations proposed in other studies.