Assessing species boundaries using multilocus species delimitation in a morphologically conserved group of neotropical freshwater fishes, the Poecilia sphenops species complex (Poeciliidae).

M Florencia Breitman,Fernando Alda,Jerald B Johnson,Eldredge Bermingham,Justin C Bagley,Eric P Van Den Berghe,Michael A Russello

doi:10.1371/journal.pone.0121139

Abstract

Accurately delimiting species is fundamentally important for understanding species diversity and distributions and devising effective strategies to conserve biodiversity. However, species delimitation is problematic in many taxa, including ‘non-adaptive radiations’ containing morphologically cryptic lineages. Fortunately, coalescent-based species delimitation methods hold promise for objectively estimating species limits in such radiations, using multilocus genetic data. Using coalescent-based approaches, we delimit species and infer evolutionary relationships in a morphologically conserved group of Central American freshwater fishes, the Poecilia sphenops species complex. Phylogenetic analyses of multiple genetic markers (sequences of two mitochondrial DNA genes and five nuclear loci) from 10/15 species and genetic lineages recognized in the group support the P. sphenops species complex as monophyletic with respect to outgroups, with eight mitochondrial ‘major-lineages’ diverged by ≥2% pairwise genetic distances. From general mixed Yule-coalescent models, we discovered (conservatively) 10 species within our concatenated mitochondrial DNA dataset, 9 of which were strongly supported by subsequent multilocus Bayesian species delimitation and species tree analyses. Results suggested species-level diversity is underestimated or overestimated by at least ~15% in different lineages in the complex. Nonparametric statistics and coalescent simulations indicate genealogical discordance among our gene tree results has mainly derived from interspecific hybridization in the nuclear genome. However, mitochondrial DNA show little evidence for introgression, and our species delimitation results appear robust to effects of this process. Overall, our findings support the utility of combining multiple lines of genetic evidence and broad phylogeographical sampling to discover and validate species using coalescent-based methods. Our study also highlights the importance of testing for hybridization versus incomplete lineage sorting, which aids inference of not only species limits but also evolutionary processes influencing genetic diversity.

Highlights

Species are widely used as fundamental units of analysis in biogeography, ecology, and evolutionary biology [1,2,3]
An outstanding majority of tests (91.4%) recovered no evidence for recombination in any of the nuclear loci analyzed (S1 Appendix): six tests of each of five loci in RDP3 inferred a total of only three recombination signals, and coalescent simulations showed no evidence of recombination based on recombination events (RM) values (P > 0.05; S1 Appendix)
The position of the Tipitapa lineage was poorly resolved by mitochondrial DNA (mtDNA), and its sister relationship to all other P. sphenops species complex lineages received marginal support, yet given its genetic distinctiveness we refer to this P. mexicana-like lineage as a ‘candidate species’, P. sp

Summary

Introduction

Species are widely used as fundamental units of analysis in biogeography, ecology, and evolutionary biology [1,2,3]. Species delimitation, the practice of determining species boundaries and discovering new species, is of fundamental importance for understanding species diversity and distributions, and devising effective strategies to conserve biodiversity [5,6,7]. Inaccurately classifying individuals or populations to species could result in erroneous inferences in any analysis requiring a priori designation of species limits, such as comparative analyses of diversification [8,9], or misallocation of conservation resources and loss of species (e.g. under the U.S Endangered Species Act of 1973; [5]). In light of practical difficulties presented by applying alternative operational criteria, there is a growing consensus that multiple perspectives from different datatypes or analyses are necessary to accurately delimit species, through ‘integrative taxonomy’, e.g. uniting classical morphology, phylogenetics, and ecological data and modeling [15,16,17]

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