Adaptive introgression involves the acquisition of advantageous genetic variants through hybridization, which subsequently are favored by natural selection due to their association with beneficial traits. These post-introgression adaptive alleles inherited from related species may allow the hybrid lineage to adapt to new environmental changes or exploit novel ecological niches. Here, we analyzed speciation patterns of the kleptoparasitic spider Argyrodes lanyuensis through genomic analyses and tested for genetic evidence of adaptive introgression at the Taiwan-Philippines transition zone. Using highly polymorphic SNPs, our study demonstrated that speciation occurred when the Hualien (on Taiwan Island) and Philippine (including the Orchid Island) lineages separated during the early to mid-Pleistocene. The best colonization model suggested by Approximate Bayesian Computation and Random Forests supported an inference of a bottleneck during speciation, an interpretation reinforced by observation of lower F_ST values and reduced genetic diversity of the Philippine+Orchid Island lineage. We found the highest support for the occurrence of introgression on the youngest island (Green Island) of the Taiwan-Philippines transition zone based on ABBA-BABA test. Additionally, we have identified a putative adaptive locus under balancing selection on Green Island, suggesting evolution by adaptive introgression in a newly-formed niche (or novel geographical context). Our study highlights a possible rare case of introgression at the Taiwan-Philippines transition zone under balancing selection, that could be an evolutionary response to a unique climatological zone lying between the tropical climate of the Philippines and the subtropical climate of Hualien, Taiwan.

mt-COI dataset:
We extract the mt-COI gene sequences of 2 Hualien samples and 2 Green Island samples from our RAD markers by minimap2 (Heng Li, 2018). We aligned these sequences with four previously published mt-COI haplotype sequences of Responte et al., (2021) available in Genbank (MN881069.1, MN881070.1, MN881071.1, MN881072.1) and one outgroup species Argyrodes rainbowi  (MW549752.1). In total, we aligned eight mt-COI sequences from the ingroup and one additional outgroup species using mafft (Katoh, K., & Standley, D. M., 2013). We implemented the coalescent tree model (Kingman, J. F. C., 1982) and the uncorrelated lognormal relaxed clock model (Drummond et al., 2006) as priors in BEAST. A ucld.mean of 0.0112 site-1, derived from the mitochondrial substitution rate estimates in spiders (Bidegaray-Batista & Arnedo, 2011; Kuntner et al., 2013), and used a standard deviation of ucld.stdv = 0.01. We ran the MCMC chain for 5 x 10⁸ generations with tree sampling frequency per 1 x 10⁴ generations in three replicates. Burn-in was determined using Tracer v.1.7.1 (Rambaut et al., 2018), in which the first 10 percent of the trees were discarded (Rambaut et al., 2018). The maximum clade credibility (MCC) tree was summarized in TreeAnnotator (Drummond et al., 2012) and visualized using FigTree (Rambaut, 2014).

RAD-seq dataset:

We used the Multiplex Shotgun Genotyping (MSG; Andolfatto et al., 2011) with modifications from our in-house protocol to prepare the high-throughput sequencing libraries. After size selection using Pippin Prep with a 1.5% agarose gel cassette, the size selected fragments (300-450 bp) were amplified using the Phusion High Fidelity PCR kit (NEB, USA). Then, we purified the DNA libraries using AMPure XP magnetic beads (Beckman, USA) following the manufacturer’s instructions.  We have multiplexed 130 samples and were sequenced using the Illumina Nova-seq 6000 in a sequencing facility (Genomics Co. Ltd., Taiwan). We demultiplexed Illumina raw reads using the process_radtags program from stacks manual version 2.4 (Catchen et al., 2013) to clean and trim the sequences. The combined forward and reverse reads from PEAR version 0.9.6 (Zhang et al., 2014) were aligned with the A. miniaceus reference genome using the Burrows-Wheeler Alignment (BWA) tool (Li & Durbin, 2009).