Morphometrics, molecular ecology and multivariate environmental niche define the evolutionary history of the Western Rattlesnake (Crotalus viridis) complex

Species are the currency of biodiversity and an accurate recognition of their status is a scientific necessity, particularly given the onset of the Anthropocene (the most recent biodiversity crisis). Yet, concept-based species approaches are contentious whereas those more deterministic work against the fluidity of speciation itself. This conceptual gap can be bridged through a comprehensive assessment of the nine subspecies comprising the historically enigmatic Western Rattlesnake (Crotalus viridis) complex, one that employs three disparate datasets. First, mitochondrial DNA (mtDNA) sequence data were used to derive a Bayesian phylogenetic hypothesis that revealed two well- supported lineages, each with subspecies as distinct clades. Second, morphological data relating to head shape were analyzed using Geometric Morphometric (GM) methodology to again reveal two distinct lineages, each composed of subspecies that differ significantly in shape, yet with confounding factors that obscure evolutionary relationships. Finally, GIS-based macroecological variables gathered from museum specimens again demonstrated significant subspecific niches, suggesting the potential for ecological speciation within the complex. The three datasets (molecular, morphological, and ecological) were coalesced using supertree methodology to derive a single hypothesis that supported two distinct lineages but with obscured subspecific relationships. They were also utilized in crosshair classification tests that quantified ‘historical’ and ‘recent’ non-exchangeability among lineages and subspecies (i.e., if these entities were distinct amongst themselves and worthy of taxonomic recognition). In this regard, sufficient genetic, ecological and morphological non-exchangeability exists between lineages and among subspecies to warrant species designations for six of 9 Western Rattlesnake subspecies, with the remaining three retained at subspecific status. These efforts thus represent a comprehensive and contemporary perspective of Western Rattlesnake biodiversity, and shed light on a group that has proven elusive across two centuries of scientific inquiry