Discussion
Species delimitations that are informed by genetic data have undergone a
renaissance as increasingly more sophisticated sequencing technology and
analytical methods have become available. But genes are only part of
what makes a species. Species delimitations informed by ecological
characteristics – the phenotypic interactions of individuals with their
environment – have been difficult to incorporate quantitatively and are
usually relegated to natural history accounts. Our study uses a
notoriously confusing species group of butterflies to demonstrate an
analytical integration of genetic and ecological species concepts. This
approach supports species delimitations that should ultimately be more
stable and meaningful in conservation contexts and citizen science.
Evidence for historical introgression between S. hesperisand S. zerene
Our genomic analyses suggest a complex, shared evolutionary history
between S. atlantis, S. hesperis, and S. zerene . BFD*
species delimitation supported separation of northern and southernS. hesperis lineages as distinct species, and a sister
relationship between S. atlantis and northern S. hesperis ,
consistent with our SNP phylogeny (Fig. 1a). However, the relationship
of S. zerene to the other clades varied. This is interesting, as
recent molecular phylogenetic work on Speyeria generally
indicates a non-sister relationship between S. zerene andS. hesperis /S. atlantis (de Moya et al. 2017;
Campbell et al. 2017, 2019; Thompson et al. 2019). It is
likely that the recovered polyphyly of S. hesperis and the sister
relationship between southern S. hesperis and S. zerenepresented here is partly due to the omission of other Speyeriaspecies in our phylogenetic analyses, but also due to probable
introgression between S. hesperis and S. zerene, which is
more explicitly indicated by SNP-based admixture and COIhaplotype analyses (Fig. 2, Fig. 3, Table S2).
While the results of our SNP-based analyses and the lack of sequence
variation in the mitochondrial haplotype shared between S. zereneand S. hesperis support a hypothesis of introgression between
these non-sister species, they do not sufficiently clarify the direction
of gene flow between them; TreeMix and f 3 tests indicated
introgression from S. zerene into S. hesperis , however our
inference that the S. hesperis haplotype occurs throughout the
entire S. hesperis sampled range, but only in the Nevadan part of
the range of S. zerene, suggests that it originated in S.
hesperis . Expanded sampling of both species is needed to clarify the
pervasiveness of this haplotype in S. zerene and to validate its
origin.
In contrast to mtDNA, nuclear SNPs across the range of S.
hesperis do not show the same obvious reduction in genetic variability,
suggesting that a strong selective sweep leading to a severe bottleneck
event has recently caused the loss of other variable mitochondrial
haplotypes (Sonsthagen et al. 2017; Hurst & Jiggins 2005). A
candidate for facilitating such a process is Wolbachia Hertig &
Wolbach, 1924, maternally-inherited, endosymbiotic bacteria that can
facilitate the spread of particular mitochondrial haplotypes throughout
populations and species, reducing haplotype variation (Werren et
al. 2008; Kodandaramaiah et al. 2013; Ahmed et al. 2015).Wolbachia infections have been reported in severalSpeyeria species, including S. zerene (McHugh et
al. 2013), but not yet within the S. atlantis-hesperis complex
(Hamm et al. 2014). Wolbachia infection via introgression
offers a plausible and testable hypothesis that could explain the
observed haplotype sharing between Nevadan S. zerene gunderi andS. hesperis in the absence of contemporary nuclear admixture in
the sampled specimens, which may further clarify the historical
relationship between these taxa.