Habitat or temporal isolation: Unraveling herbivore-parasitoid speciation patterns using double digest RADseq.

Ecological speciation is often observed in phytophagous insects and their parasitoids due to divergent selection caused by host-associated or temporal differences. Most previous studies have utilized limited genetic markers or distantly related species to look for reproductive barriers of speciation. In our study, we focus on closely related species of Lygus bugs and two sister species of Peristenus parasitoid wasps. Using mitochondrial DNA COI and genomewide SNPs generated using ddRADseq, we tested for potential effects of host-associated differentiation (HAD) or temporal isolation in this system. While three species of Lygus are clearly delineated with both COI and SNPs, no evidence of HAD or temporal differentiation was detected. Two Peristenus sister species were supported by both sets of markers and separated temporally, with P. mellipes emerging early in June and attacking the first generation of Lygus, while P. howardi emerging later in August and attacking the second generation of their hosts. This is one of the few studies to examine closely related hosts and parasitoids to examine drivers of diversification. Given the results of this study, the Lygus-Peristenus system demonstrates temporal isolation as a potential barrier to reproductive isolation for parasitoids, which could indicate higher parasitoid diversity in regions of multivoltine hosts. This study also demonstrates that incorporating systematics improves studies of parasitoid speciation, particularly by obtaining accurate host records through rearing, carefully delimiting cryptic species and examining population-level differences with genomic-scale data among closely related taxa.