A temperate pseudoscorpion lineage in Taiwan: species delimitation and evolutionary history

Abstract

Integrative taxonomy facilitates determining species boundaries in morphologically conserved taxa with limited dispersal abilities, such as litter-dwelling pseudoscorpions in the genus Allochthonius (family Pseudotyrannochthiniidae). This genus is common in East Asia but taxonomically challenging due to its cryptic morphology and variable diagnostic characters. We present a multi-locus phylogeny of Allochthonius in Japan, Korea, and Taiwan, including 55 newly-acquired specimens from 11 locations across Taiwan. We find that Allochthonius in Taiwan is monophyletic and comprises multiple undescribed species. Consolidating morphology with molecular species delimitation based on three tree-based or distance-based methods, we recognize seven new species with replicable morphological and molecular differences: A. agyaqensis, A. aquaeductensis, A. brunus, A. kiunnensis, A. nebulosus, A. nubicolus, and A. tamanensis. Phylogenetic analyses and divergence time estimations suggest that Taiwanese Allochthonius separated from its Korea-Japan relatives ca. 15 million years ago, and that cooling climates during the Late Miocene and subsequent warming and rapid uplift of the Central Mountain Range during Pliocene likely promoted diversification. Spatial mapping suggests that litter-dwelling Allochthonius species living in tropical and subtropical montane regions may have a sky-island distribution with high endemism and naturally small distribution ranges.

Key words: False scorpions, Molecular phylogeny, New species, Species delineation, Taxonomy

Methodology and Implementation Steps

Specimens were collected from the leaf litter of mid- to high-altitude forests in Taiwan. Pseudoscorpions were extracted from the leaf litter using Berlese funnels. All specimens were preserved in 95% ethanol. Prior to DNA extraction, two legs were removed from each specimen for molecular analyses. Genomic DNA was extracted using the QIAGEN DNA Micro Kit, and fragments of COI, H3, and ITS2 were amplified.

1. Molecular species delimitation
   Phylogenetic analysis: IQ-TREE with 1,000 ultrafast bootstrap replicates
   Haplotype network: PopArt (TCS network)
   Species delimitation analyses: ASAP, mPTP, and bPTP
   p-distance: MEGAX
2. Morphological examination
   After estimating the number of putative species based on the molecular analyses, morphological characters were examined to identify diagnostic features that were congruent with the phylogenetic relationships.

Innovation and Cross-Disciplinary Collaboration

• This study contributes substantially to our understanding of Taiwanese pseudoscorpion diversity and advances the taxonomy of the group.
• It represents the first integrative taxonomic study of this genus to incorporate molecular data and reconstruct its phylogenetic relationships.
• It is also the first biogeographic study focusing on East Asian pseudoscorpions, whereas previous studies have primarily been restricted to Europe and Australia.
• Furthermore, this is the first study to comprehensively examine morphological variation within the genus, allowing us to identify a set of more reliable diagnostic characters for species identification.

Expected Results and Contributions

By integrating morphological and molecular evidence, we delimited species boundaries within Allochthonius, described seven new species, and identified diagnostic morphological characters suitable for species delimitation in the genus. Diversification of this genus in Taiwan was likely driven by late Miocene cooling followed by the uplift of the Central Mountain Range. Future climate warming and habitat loss may threaten these narrowly endemic species.