An Integrative Approach Using Phylogenomics and High-Resolution X-Ray Computed Tomography for Species Delimitation in Cryptic Taxa

Abstract

Morphologically cryptic taxa have proved to be a long-standing challenge for taxonomists. Lineages that show strong genomic structuring across the landscape but are phenotypically similar pose a conundrum, with traditional morphological analyses of these cryptic lineages struggling to keep up with species delimitation advances. Micro X-ray computed tomography (CT) combined with geometric morphometric analyses provides a promising avenue for identification of morphologically cryptic taxa, given its ability to detect subtle differences in anatomical structures. However, this approach has yet to be used in combination with genomic data in a comparative analytical framework to distinguish cryptic taxa. We present an integrative approach incorporating genomic and geometric morphometric evidence to assess the species delimitation of grassland earless dragons (Tympanocryptis spp.) in north-eastern Australia. Using mitochondrial and nuclear genes (ND2 and RAG1, respectively), along with $>$8500 SNPs (nuclear single nucleotide polymorphisms), we assess the evolutionary independence of target lineages and several closely related species. We then integrate phylogenomic data with osteological cranial variation between lineages using landmark-based analyses of three-dimensional CT models. High levels of genomic differentiation between the three target lineages were uncovered, also supported by significant osteological differences. By incorporating multiple lines of evidence, we provide strong support for three undescribed cryptic lineages of Tympanocryptis in north-eastern Australia that warrant taxonomic review. Our approach demonstrates the successful application of CT with integrative taxonomic approaches for cryptic species delimitation, which is broadly applicable across vertebrates containing morphologically similar yet genetically distinct lineages. Additionally, we provide a review of recent integrative taxonomic approaches for cryptic species delimitation and an assessment of how our approach can value-add to taxonomic research.

Biogeographic barriers, Pleistocene refugia, and climatic gradients in the southeastern Nearctic drive diversification in cornsnakes (Pantherophis guttatus complex)

“…Previous analyses of morphological characters have shown that scale counts are not discreet but instead continuous between the currently named taxa (Burbrink, ), whether this is the case for the eastern and western populations of P. guttatus is not yet known. Morphological variation among these recently evolved taxa might be subtle (e.g., differences in head shape variation; Ruane, ) and may be detected using high‐resolution morphological data from specimen‐based studies (Chaplin, Sumner, Hipsley, & Melville, ).…”

“…Morphological variation among these recently evolved taxa might be subtle (e.g., differences in head shape variation; Ruane, 2015) and may be detected using high-resolution morphological data from specimen-based studies (Chaplin, Sumner, Hipsley, & Melville, 2019).…”

‘Barcode fishing’ for archival DNA from historical type material overcomes taxonomic hurdles, enabling the description of a new frog species

“…Intrinsic hindrances, which we define as properties of the species themselves that complicate taxonomy, such as morphologically cryptic species, can be overcome by diversification of data types in species delimitation; in most cases, diagnostic features of cryptic species can be found with the right methods and datasets (e.g. micro-CT 2,3 ). Extrinsic hindrances, which typically concern nomenclatural issues but can also relate, for example, to the lack of availability of specimen material, are less often discussed, but can be equally difficult, or even more difficult, to overcome.…”

High-throughput microCT scanning of small specimens: preparation, packing, parameters and post-processing

“…Common biodiversity metrics such as species identity, richness, and evenness are inferable from μCT data, e.g. 27 , 28 , especially for cryptic taxa and species complexes with few observable differences 29 – 31 . Therefore, optimization of the scanning process for high resolution 3D images of multiple specimens would not only contribute to our understanding of how biodiversity responds to global change, but also to the value of museum collections and researchers’ abilities to access them digitally.…”