Applications of ecological niche modeling for species delimitation: a review and empirical evaluation using day geckos (Phelsuma) from Madagascar

Although the systematic utility of ecological niche modeling is generally well known (e.g., concerning the recognition and discovery of areas of endemism for biogeographic analyses), there has been little discussion of applications concerning species delimitation, and to date, no empirical evaluation has been conducted. However, ecological niche modeling can provide compelling evidence for allopatry between populations, and can also detect divergent ecological niches between candidate species. Here we present results for two taxonomically problematic groups of Phelsuma day geckos from Madagascar, where we integrate ecological niche modeling with mitochondrial DNA and morphological data to evaluate species limits. Despite relatively modest levels of genetic and morphological divergence, for both species groups we find divergent ecological niches between closely related species and parapatric ecological niche models. Niche models based on the new species limits provide a better fit to the known distribution than models based upon the combined (lumped) species limits. Based on these results, we elevate three subspecies of Phelsuma madagascariensis to species rank and describe a new species of Phelsuma from the P. dubia species group. Our phylogeny continues to support a major endemic radiation of Phelsuma in Madagascar, with dispersals to Pemba Island and the Mascarene Islands. We conclude that ecological niche modeling offers great potential for species delimitation, especially for taxonomic groups exhibiting low vagility and localized endemism and for groups with more poorly known distributions. In particular, niche modeling should be especially sensitive for detecting recent parapatric speciation driven by ecological divergence, when the environmental gradients driving speciation are represented within the ecological niche models.     

Integrative species delimitation reveals cryptic diversity in the southern Appalachian Antrodiaetus unicolor (Araneae: Antrodiaetidae) species complex

Although species delimitation can be highly contentious, the development of reliable methods to accurately ascertain species boundaries is an imperative step in cataloguing and describing Earth's quickly disappearing biodiversity. Spider species delimitation remains largely based on morphological characters; however, many mygalomorph spider populations are morphologically indistinguishable from each other yet have considerable molecular divergence. The focus of our study, the Antrodiaetus unicolor species complex containing two sympatric species, exhibits this pattern of relative morphological stasis with considerable genetic divergence across its distribution. A past study using two molecular markers, COI and 28S, revealed that A. unicolor is paraphyletic with respect to A. microunicolor. To better investigate species boundaries in the complex, we implement the cohesion species concept and use multiple lines of evidence for testing genetic exchangeability and ecological interchangeability. Our integrative approach includes extensively sampling homologous loci across the genome using a RADseq approach (3RAD), assessing population structure across their geographic range using multiple genetic clustering analyses that include structure , principal components analysis and a recently developed unsupervised machine learning approach (Variational Autoencoder). We evaluate ecological similarity by using large‐scale ecological data for niche‐based distribution modelling. Based on our analyses, we conclude that this complex has at least one additional species as well as confirm species delimitations based on previous less comprehensive approaches. Our study demonstrates the efficacy of genomic‐scale data for recognizing cryptic species, suggesting that species delimitation with one data type, whether one mitochondrial gene or morphology, may underestimate true species diversity in morphologically homogenous taxa with low vagility.     

Investigating niche and lineage diversification in widely distributed taxa: phylogeography and ecological niche modeling of the Peromyscus maniculatus species group

The relationship between lineage formation and variation in the ecological niche is a fundamental evolutionary question. Two prevailing hypotheses reflect this relationship: niche conservatism and niche divergence. Niche conservatism predicts a pattern where sister taxa will occupy similar niche spaces; whereas niche divergence predicts that sister taxa will occupy different niche spaces. Widely distributed species often show distinct phylogeographic structure, but little research has been conducted on how the environment may be related to these phylogenetic patterns. We investigated the relationship between lineage divergence and environmental space for the closely related species Peromyscus maniculatus and P. polionotus utilizing phylogenetic techniques and ecological niche modeling (ENM). We estimated the phylogenetic relationship among individuals based on complete cytochrome b sequences that represent individuals from a majority of the species ranges. Niche spaces that lineages occupy were estimated by using 12 environmental layers. Differences in niche space were tested using multivariate statistics based on location data, and ENMs were employed using maximum entropy algorithms. Two similarity indices estimated significant divergence in environmental space based on the ENM. Six geographically structured lineages were identified within P. maniculatus. Nested within P. maniculatus we found that P. polionotus recently diverged from a clade occupying central and western United States. We estimated that the majority of the genetic lineages occupy distinct environmental niches, which supports a pattern of niche divergence. Two sister taxa showed niche divergence and represent different ecomorphs, suggesting morphological, genetic and ecological divergence between the two lineages. Two other sister taxa were observed in the same environmental space based on multivariate statistics, suggesting niche conservatism. Overall our results indicate that a widely distributed species may exhibit both niche conservatism and niche divergence, and that most lineages seem to occupy distinct environmental niches.     

Delimitation despite discordance: Evaluating the species limits of a confounding species complex in the face of mitonuclear discordance

The delimitation of species is an essential pursuit of biology, and proper taxonomies are crucial for the assessment and conservation management of organismal diversity. However, delimiting species can be hindered by a number of factors including highly conserved morphologies (e.g., cryptic species), differences in criteria of species concepts, lineages being in the early stages of the speciation or divergence process, and discordance between gene topologies (e.g., mitonuclear discordance). Here we use a taxonomically confounded species complex of toads in Central America that exhibits extensive mitonuclear discordance to test delimitation hypotheses. Our investigation integrates mitochondrial sequences, nuclear SNPs, morphology, and macroecological data to determine which taxonomy best explains the divergence and evolutionary relationships among these toads. We found that a three species taxonomy following the distributions of the nuclear SNP haplotypes offers the best explanation of the species in this complex based off of the integrated data types. Due to the taxonomic instability of this group, we also discuss conservation concerns in the face of improper taxonomic delimitation. Our study provides an empirical and integrative hypothesis testing framework to assess species delimitation hypotheses in the face of cryptic morphology and mitonuclear discordance and highlights the importance that a stable taxonomy has over conservation‐related actions.     

Exploring the Effect of Asymmetric Mitochondrial DNA Introgression on Estimating Niche Divergence in Morphologically Cryptic Species

If potential morphologically cryptic species, identified based on differentiated mitochondrial DNA, express ecological divergence, this increases support for their treatment as distinct species. However, mitochondrial DNA introgression hampers the correct estimation of ecological divergence. We test the hypothesis that estimated niche divergence differs when considering nuclear DNA composition or mitochondrial DNA type as representing the true species range. We use empirical data of two crested newt species (Amphibia: Triturus) which possess introgressed mitochondrial DNA from a third species in part of their ranges. We analyze the data in environmental space by determining Fisher distances in a principal component analysis and in geographical space by determining geographical overlap of species distribution models. We find that under mtDNA guidance in one of the two study cases niche divergence is overestimated, whereas in the other it is underestimated. In the light of our results we discuss the role of estimated niche divergence in species delineation.     

The Role of Integrative Taxonomy in the Conservation Management of Cryptic Species: The Taxonomic Status of Endangered Earless Dragons (Agamidae: Tympanocryptis) in the Grasslands of Queensland,Australia

Molecular phylogenetics is increasingly highlighting the prevalence of cryptic species, where morphologically similar organisms have long independent evolutionary histories. When such cryptic species are known to be declining in numbers and are at risk of extinction due to a range of threatening processes, the disjunction between molecular systematics research and conservation policy becomes a significant problem. We investigate the taxonomic status of Tympanocryptis populations in Queensland, which have previously been assigned to T. tetraporophora, using three species delimitation approaches. The taxonomic uncertainties in this species-group are of particular importance in the Darling Downs Earless Dragon (T. cf. tetraporophora), which is ranked as an endangered ‘species’ of high priority for conservation by the Queensland Department of Environment and Heritage Protection. We undertook a morphological study, integrated with a comprehensive genetic study and species delimitation analyses, to investigate the species status of populations in the region. Phylogenetic analyses of two gene regions (mtDNA: ND2; nuclear: RAG1) revealed high levels of genetic divergence between populations, indicating isolation over long evolutionary time frames, and strongly supporting two independent evolutionary lineages in southeastern Queensland, from the Darling Downs, and a third in the Gulf Region of northern Queensland. Of the three species delimitation protocols used, we found integrative taxonomy the most applicable to this cryptic species complex. Our study demonstrates the utility of integrative taxonomy as a species delimitation approach in cryptic complexes of species with conservation significance, where limited numbers of specimens are available.     

Genomic data reveal two distinct species from the widespread alpine ginger Roscoea tibetica Batalin (Zingiberaceae)

Species delimitation is a key foundation for exploring biodiversity. However, the existence of continuous phenotypic variation in widespread species challenges accurate species delimitation based on classical taxonomy. In this study, we investigated the cryptic diversity of a widespread herb (Roscoea tibetica Batalin) in a biodiversity hotspot (the Hengduan Mountains, China) using genotyping by sequencing, examining morphological traits, developing species distribution models, and simulating demographic history. Phylogenomic reconstruction, principal component analysis, and genetic structure inferences indicated that previously reported R. tibetica comprised two monophyletic lineages with a deep divergence. Several morphological diagnostic characteristics were discovered from field and common garden that corresponded to these independent evolutionary lineages. Species distribution models illustrated significant ecological divergence between both lineages. All evidence strongly supported that R. tibetica, as described in previous taxonomy, actually comprises two distinct species. Model test of gene flow and effective population size changes in fastsimcoal2, and a negative Tajima's D-value suggested that recent contact likely occurred between the two lineages. Our results proposed that cryptic diversity in previously reported R. tibetica was possibly associated with phenotypic plasticity in heterogeneous environments and morphological convergence in similar habitats. This study suggests that caution should be exercised when attempting to gain biological insight into species with large-scale morphological variation, and species delimitation should be done in advance.     

Coalescent species delimitation of a Sumatran parachuting frog

We conduct species delimitation of the widespread parachuting frog species Rhacophorus catamitus using samples from across the island of Sumatra, Indonesia. We use mitochondrial, genomic and morphological data, and find that R. catamitus is composed of three lineages corresponding to northern, central and southern lineages. Mitochondrial and genomic data show admixture or incomplete lineage sorting between the central and southern lineages, but deep divergence from the northern lineage. Coalescent species delimitation supports a three species model for this complex, and we recommend that the northern lineage be described as a new species. Our study highlights the power of coalescent species delimitation in an integrative framework for identifying unrecognised diversity in understudied tropical species complexes. We also emphasise the evolutionary importance of northern Sumatra, a region that harboured montane refugia during Pliocene–Pleistocene climate change, but has also been heavily affected by volcanic activity.     

Lineage diversification in a widespread species: roles for niche divergence and conservatism in the common kingsnake, Lampropeltis getula

Niche conservatism and niche divergence are both important ecological mechanisms associated with promoting allopatric speciation across geographical barriers. However, the potential for variable responses in widely distributed organisms has not been fully investigated. For allopatric sister lineages, three patterns for the interaction of ecological niche preference and geographical barriers are possible: (i) niche conservatism at a physical barrier; (ii) niche divergence at a physical barrier; and (iii) niche divergence in the absence of a physical barrier. We test for the presence of these patterns in a transcontinentally distributed snake species, the common kingsnake ( Lampropeltis getula ), to determine the relative frequency of niche conservatism or divergence in a single species complex inhabiting multiple distinct ecoregions. We infer the phylogeographic structure of the kingsnake using a range-wide data set sampled for the mitochondrial gene cytochrome b . We use coalescent simulation methods to test for the presence of structured lineage formation vs. fragmentation of a widespread ancestor. Finally, we use statistical techniques for creating and evaluating ecological niche models to test for conservatism of ecological niche preferences. Significant geographical structure is present in the kingsnake, for which coalescent tests indicate structured population division. Surprisingly, we find evidence for all three patterns of conservatism and divergence. This suggests that ecological niche preferences may be labile on recent phylogenetic timescales, and that lineage formation in widespread species can result from an interaction between inertial tendencies of niche conservatism and natural selection on populations in ecologically divergent habitats.     

Lack of congruence of genetic and niche divergence in Podarcis hispanicus complex

Niche divergence among closely related lineages can be informative on the ecological and evolutionary processes involved in differentiation, particularly in the case of cryptic species complexes. Here we compared phylogenetic relationships and niche similarity between pairs of lineages included in the Podarcis hispanicus complex to examine patterns of niche divergence and its role in the organization of current diversity patterns, as allopatric, parapatric, and sympatric lineages occur in the Western Mediterranean Basin. First, we used ecological niche models to characterize the realized climatic niche of each Podarcis hispanicus complex lineage based on topographic and climatic variables, to identify important variables, and to test for niche conservatism or divergence between pairs of lineages. Variables related to precipitation generally exhibited the highest contribution to niche models, highlighting the importance of rainfall levels in shaping distributions of Podarcis wall lizards. We found that most forms have significant differences in realized climatic niches that do not follow the pattern of mitochondrial divergence. These results lend support to the hypothesis that genetic divergence across Podarcis hispanicus complex most likely occurred in allopatric conditions, mostly with significant niche divergence. Competition after secondary contact is also suggested by the common occurrence of niche overlap between lineages that exhibit strictly parapatric distribution. The almost continuous distribution of Podarcis lizards in the study area appears to be a result of a combination of complementary suitable niches and competition, which seem two important mechanisms limiting geographic distributions and restricting the existence of extensive contact zones.     

Corresponding Mitochondrial DNA and Niche Divergence for Crested Newt Candidate Species

Genetic divergence of mitochondrial DNA does not necessarily correspond to reproductive isolation. However, if mitochondrial DNA lineages occupy separate segments of environmental space, this supports the notion of their evolutionary independence. We explore niche differentiation among three candidate species of crested newt (characterized by distinct mitochondrial DNA lineages) and interpret the results in the light of differences observed for recognized crested newt species. We quantify niche differences among all crested newt (candidate) species and test hypotheses regarding niche evolution, employing two ordination techniques (PCA-env and ENFA). Niche equivalency is rejected: all (candidate) species are found to occupy significantly different segments of environmental space. Furthermore, niche overlap values for the three candidate species are not significantly higher than those for the recognized species. As the three candidate crested newt species are, not only in terms of mitochondrial DNA genetic divergence, but also ecologically speaking, as diverged as the recognized crested newt species, our findings are in line with the hypothesis that they represent cryptic species. We address potential pitfalls of our methodology.     

The importance (or lack thereof) of niche divergence to the maintenance of a northern species complex: the case of the long‐toed salamander (Ambystoma macrodactylum Baird)

The relative importance of ecological vs. nonecological factors for the origin and maintenance of species is an open question in evolutionary biology. Young lineages – such as the distinct genetic groups that make up the ranges of many northern species – represent an opportunity to study the importance of ecological divergence during the early stages of diversification. Yet, few studies have examined the extent of niche divergence between lineages in previously glaciated regions and the role of ecology in maintaining the contact zones between them. In this study, we used tests of niche overlap in combination with ecological niche models to explore the extent of niche divergence between lineages of the long‐toed salamander (Ambystoma macrodactylum Baird) species complex and to determine whether contact zones correspond to (divergent) niche limits. We found limited evidence for niche divergence between the different long‐toed salamander lineages, substantial overlap in the predicted distribution of suitable climatic space for all lineages and range limits that are independent of niche limits. These results raise questions as to the importance of ecological divergence to the development of this widespread species complex and highlight the potential for non‐ecological factors to play a more important role in the maintenance of northern taxa.     

Genetic and ecological data reveal species boundaries between viviparous and oviparous lizard lineages

Identification of cryptic species is an essential aim for conservation biologists to avoid premature extinctions of ‘unrecognized'' species. Integrating different types of data can undoubtedly aid in resolving the issue of species delimitation. We studied here two lineages of the common lizard Zootoca vivipara that display different reproductive mode (the viviparous Z. v. vivipara and the oviparous Z. v. carniolica) and that overlap their distributional ranges in the European Alps. With the purpose of delimiting species'' boundaries, we analyzed their ecological, genetic and natural history features. More than 300 samples were collected and analyzed at cytochrome b and 11 microsatellites loci for investigating genetic variation, population structure, individual relatedness and evolutionary histories of the two lineages. Additionally, we compared their ecological niches using eight ecological variables. Genetic data showed contrasting patterns of genetic structure between the two lineages, different demographic dynamics and no hybridization events. Also strong ecological differences (such as temperature) emerged between the two lineages, and niche overlap was limited. Taken together, these results indicate that Z. v. vivipara and Z. v. carniolica should be recognized as two separate species, and particular conservation consideration should be given to the oviparous lineage that tends to live in areas threatened by increasing impact of human activities. However, recent and rapid climate warming might determine an increasing risk for the persistence of the viviparous lineage, being adapted to cold environments.     

Glacial cycles drive rapid divergence of cryptic field vole species

Understanding the factors that contribute to the generation of reproductively isolated forms is a fundamental goal of evolutionary biology. Cryptic species are an especially interesting challenge to study in this context since they lack obvious morphological differentiation that provides clues to adaptive divergence that may drive reproductive isolation. Geographical isolation in refugial areas during glacial cycling is known to be important for generating genetically divergent populations, but its role in the origination of new species is still not fully understood and likely to be situation dependent. We combine analysis of 35,434 single‐nucleotide polymorphisms (SNPs) with environmental niche modeling (ENM) to investigate genomic and ecological divergence in three cryptic species formerly classified as the field vole (Microtus agrestis). The SNPs demonstrate high genomic divergence (pairwise F_ST values of 0.45–0.72) and little evidence of gene flow among the three field vole cryptic species, and we argue that genetic drift may have been a particularly important mechanism for divergence in the group. The ENM reveals three areas as potential glacial refugia for the cryptic species and differing climatic niches, although with spatial overlap between species pairs. This evidence underscores the role that glacial cycling has in promoting genetic differentiation and reproductive isolation by subdivision into disjunct distributions at glacial maxima in areas relatively close to ice sheets. Future investigation of the intrinsic barriers to gene flow between the field vole cryptic species is required to fully assess the mechanisms that contribute to reproductive isolation. In addition, the Portuguese field vole (M. rozianus) shows a high inbreeding coefficient and a restricted climatic niche, and warrants investigation into its conservation status.     

DNA-based species delimitation in algae

Given the problems of species delimitation in algae using morphology or sexual compatibility, molecular data are becoming the standard for delimiting species and testing their traditional boundaries. The idea that species are separately evolving metapopulation lineages, along with theoretical progress in phylogenetic and population genetic analyses, has led to the development of new methods of species delimitation. We review these recent developments in DNA-based species delimitation methods, and discuss how they have changed and continue to change our understanding of algal species boundaries. Although single-locus approaches have proven effective for a first rapid and large-scale assessment of species diversity, species delimitation based on single gene trees falls short due to gene tree–species tree incongruence, caused by confounding processes like incomplete lineage sorting, trans-species polymorphism, hybridization and introgression. Data from unlinked loci and multi-species coalescent methods, which combine principles from phylogenetics and population genetics, may now be able to account for these complicating factors. Several of these methods also provide statistical support regarding species boundaries, which is important because speciation is a process and therefore uncertainty about precise species boundaries is inevitable in recently diverged lineages.     

The influence of locus number and information content on species delimitation: an empirical test case in an endangered Mexican salamander

Perhaps the most important recent advance in species delimitation has been the development of model‐based approaches to objectively diagnose species diversity from genetic data. Additionally, the growing accessibility of next‐generation sequence data sets provides powerful insights into genome‐wide patterns of divergence during speciation. However, applying complex models to large data sets is time‐consuming and computationally costly, requiring careful consideration of the influence of both individual and population sampling, as well as the number and informativeness of loci on species delimitation conclusions. Here, we investigated how locus number and information content affect species delimitation results for an endangered Mexican salamander species, Ambystoma ordinarium. We compared results for an eight‐locus, 137‐individual data set and an 89‐locus, seven‐individual data set. For both data sets, we used species discovery methods to define delimitation models and species validation methods to rigorously test these hypotheses. We also used integrated demographic model selection tools to choose among delimitation models, while accounting for gene flow. Our results indicate that while cryptic lineages may be delimited with relatively few loci, sampling larger numbers of loci may be required to ensure that enough informative loci are available to accurately identify and validate shallow‐scale divergences. These analyses highlight the importance of striking a balance between dense sampling of loci and individuals, particularly in shallowly diverged lineages. They also suggest the presence of a currently unrecognized, endangered species in the western part of A. ordinarium's range.     

Cryptic lineages of a common alpine mayfly show strong life‐history divergence

Understanding ecological divergence of morphologically similar but genetically distinct species – previously considered as a single morphospecies – is of key importance in evolutionary ecology and conservation biology. Despite their morphological similarity, cryptic species may have evolved distinct adaptations. If such ecological divergence is unaccounted for, any predictions about their responses to environmental change and biodiversity loss may be biased. We used spatio‐temporally replicated field surveys of larval cohort structure and population genetic analyses (using nuclear microsatellite markers) to test for life‐history divergence between two cryptic lineages of the alpine mayfly Baetis alpinus in the Swiss Alps. We found that the more widespread and abundant cryptic lineage represents a ‘generalist’ with at least two cohorts per year, whereas the less abundant lineage is restricted to higher elevations and represents a ‘specialist’ with a single cohort per year. Importantly, our results indicate partial temporal segregation in reproductive periods between these lineages, potentially facilitating local coexistence and reproductive isolation. Taken together, our findings emphasize the need for a taxonomic revision: widespread and apparently generalist morphospecies can hide cryptic lineages with much narrower ecological niches and distribution ranges.     

Evolutionary and demographic history of the Californian scrub white oak species complex: an integrative approach

Understanding the factors promoting species formation is a major task in evolutionary research. Here, we employ an integrative approach to study the evolutionary history of the Californian scrub white oak species complex (genus Quercus). To infer the relative importance of geographical isolation and ecological divergence in driving the speciation process, we (i) analysed inter‐ and intraspecific patterns of genetic differentiation and employed an approximate Bayesian computation (ABC) framework to evaluate different plausible scenarios of species divergence. In a second step, we (ii) linked the inferred divergence pathways with current and past species distribution models (SDMs) and (iii) tested for niche differentiation and phylogenetic niche conservatism across taxa. ABC analyses showed that the most plausible scenario is the one considering the divergence of two main lineages followed by a more recent pulse of speciation. Genotypic data in conjunction with SDMs and niche differentiation analyses support that different factors (geography vs. environment) and modes of speciation (parapatry, allopatry and maybe sympatry) have played a role in the divergence process within this complex. We found no significant relationship between genetic differentiation and niche overlap, which probably reflects niche lability and/or that multiple factors, have contributed to speciation. Our study shows that different mechanisms can drive divergence even among closely related taxa representing early stages of species formation and exemplifies the importance of adopting integrative approaches to get a better understanding of the speciation process.     

Environmental niche divergence between genetically distant lineages of an endangered water beetle

Historically, there has been considerable disagreement between researchers about the criteria used to discriminate among species. Decisions based on traditional morphological and genetic data alone can be potentially problematic, especially if the hypotheses are contradictory. Today, taxonomy is integrating new methods from different disciplines that study species' limits and evolution; this diverse range of evidence aids researchers in the recognition of species. Differences in niche characteristics could become a new and useful criterion in helping to decide the status of conflicting taxonomical entities. Ochthebius glaber (family Hydraenidae) is an endangered water beetle typical from southeast Iberian hypersaline streams that shows three clear discrete genetic units within its distribution range. However, there is no evidence to date that these lineages of O. glaber exhibit any adaptive morphological or ecological divergence. Using a modelling approach directed to generate niche representation from distributional data, we found a significant environmental niche divergence for allopatric lineages of O. glaber that followed an aridity gradient. Although we can not conclude firmly at present that the separate populations of O. glaber studied represent separate, reproductively isolated species, the present study complements and supports previous phylogeographic analyses through the inclusion of measures of another form of evolutionary change; in this case, ecological diversification. Despite the existence of some methodological limitations, also discussed in the present study, we emphasize the importance of recent conceptual advances that allow taxonomy to improve species delimitation practices through the integration of theory and methods from disciplines that study the origin and evolution of species. © 2011 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 103 , 891–903.