Stuart C. Willis

The Tree of Life and a New Classification of Bony Fishes

The tree of life of fishes is in a state of flux because we still lack a comprehensive phylogeny that includes all major groups. The situation is most critical for a large clade of spiny-finned fishes, traditionally referred to as percomorphs, whose uncertain relationships have plagued ichthyologists for over a century. Most of what we know about the higher-level relationships among fish lineages has been based on morphology, but rapid influx of molecular studies is changing many established systematic concepts. We report a comprehensive molecular phylogeny for bony fishes that includes representatives of all major lineages. DNA sequence data for 21 molecular markers (one mitochondrial and 20 nuclear genes) were collected for 1410 bony fish taxa, plus four tetrapod species and two chondrichthyan outgroups (total 1416 terminals). Bony fish diversity is represented by 1093 genera, 369 families, and all traditionally recognized orders. The maximum likelihood tree provides unprecedented resolution and high bootstrap support for most backbone nodes, defining for the first time a global phylogeny of fishes. The general structure of the tree is in agreement with expectations from previous morphological and molecular studies, but significant new clades arise. Most interestingly, the high degree of uncertainty among percomorphs is now resolved into nine well-supported supraordinal groups. The order Perciformes, considered by many a polyphyletic taxonomic waste basket, is defined for the first time as a monophyletic group in the global phylogeny. A new classification that reflects our phylogenetic hypothesis is proposed to facilitate communication about the newly found structure of the tree of life of fishes. Finally, the molecular phylogeny is calibrated using 60 fossil constraints to produce a comprehensive time tree. The new time-calibrated phylogeny will provide the basis for and stimulate new comparative studies to better understand the evolution of the amazing diversity of fishes.

The Casiquiare river acts as a corridor between the Amazonas and Orinoco river basins: biogeographic analysis of the genus Cichla

The Casiquiare River is a unique biogeographic corridor between the Orinoco and Amazonas basins. We investigated the importance of this connection for Neotropical fishes using peacock cichlids (Cichla spp.) as a model system. We tested whether the Casiquiare provides a conduit for gene flow between contemporary populations, and investigated the origin of biogeographic distributions that span the Casiquiare. Using sequences from the mitochondrial control region of three focal species (C. temensis, C. monoculus, and C. orinocensis) whose distributions include the Amazonas, Orinoco, and Casiquiare, we constructed maximum likelihood phylograms of haplotypes and analyzed the populations under an isolation‐with‐migration coalescent model. Our analyses suggest that populations of all three species have experienced some degree of gene flow via the Casiquiare. We also generated a mitochondrial genealogy of all Cichla species using >2000 bp and performed a dispersal‐vicariance analysis (DIVA) to reconstruct the historical biogeography of the genus. This analysis, when combined with the intraspecific results, supports two instances of dispersal from the Amazonas to the Orinoco. Thus, our results support the idea that the Casiquiare connection is important across temporal scales, facilitating both gene flow and the dispersal and range expansion of species.

Simultaneous delimitation of species and quantification of interspecific hybridization in Amazonian peacock cichlids (genus cichla) using multi-locus data.

BackgroundIntrogression likely plays a significant role in evolution, but understanding the extent and consequences of this process requires a clear identification of species boundaries in each focal group. The delimitation of species, however, is a contentious endeavor. This is true not only because of the inadequacy of current tools to identify species lineages, but also because of the inherent ambiguity between natural populations and species paradigms. The result has been a debate about the supremacy of various species concepts and criteria. Here, we utilized multiple separate sources of molecular data, mtDNA, nuclear sequences, and microsatellites, to delimit species under a polytypic species concept (PTSC) and estimate the frequency and genomic extent of introgression in a Neotropical genus of cichlid fishes (Cichla). We compared our inferences of species boundaries and introgression under this paradigm to those when species are identified under a diagnostic species concept (DSC).ResultsWe find that, based on extensive molecular data and an inclusive species concept, 8 separate biological entities should be recognized rather than the 15 described species of Cichla. Under the PTSC, fewer individuals are expected to exhibit hybrid ancestry than under the DSC (~2% vs. ~12%), but a similar number of the species exhibit introgression from at least one other species (75% vs. 60%). Under either species concept, the phylogenetic breadth of introgression in this group is notable, with both sister species and species from different major mtDNA clades exhibiting introgression.ConclusionsIntrogression was observed to be a widespread phenomenon for delimited species in this group. While several instances of introgressive hybridization were observed in anthropogenically altered habitats, most were found in undisturbed natural habitats, suggesting that introgression is a natural but ephemeral part of the evolution of many tropical species. Nevertheless, even transient introgression may facilitate an increase in genetic diversity or transfer of adaptive mutations that have important consequences in the evolution of tropical biodiversity.

Molecular systematics of the neotropical shovelnose catfish genus Pseudoplatystoma Bleeker 1862 based on nuclear and mtDNA markers

Pseudoplatystoma is a commercially important genus of Neotropical migratory catfishes widely distributed in all major river basins of South America. Historically, only three species were recognized, but a recent revision proposed eight putative morphospecies for the genus. A molecular study based on mitochondria DNA (mtDNA) provided support for recognition of only some of the species and raised questions about species boundaries in this group. We present a more encompassing analysis based on mtDNA (cytochrome b, 818bp) and nuclear DNA-based phylogenies (Rag1 intron 1, 664bp and S7 intron 1, 635bp) for a more extensive sampling (279 individuals from 42 localities) of all putative species in all major river basins. Patterns generated by individual gene genealogies and a multispecies coalescent analysis provided evidence to suggest recognition of only four distinct species in this genus: Pseudoplatystoma magdaleniatum, Pseudoplatystoma corruscans, Pseudoplatystoma tigrimun (sensu lato) and Pseudoplatystoma fasciatum (sensu lato). The species phylogeny places P. magdaleniatum as the sister group to all the other species in the genus, but the relationships among P. fasciatum s.l, P. tigrimum s.l., and P. corruscans could not be resolved with confidence.

TESTING MITOCHONDRIAL CAPTURE AND DEEP COALESCENCE IN AMAZONIAN CICHLID FISHES (CICHLIDAE: CICHLA)

Hybridization and introgression have important consequences in evolution, such as increasing the genetic diversity and adaptive potential of a species. One of their most conspicuous footprints is discordance among gene trees or between genes and phenotypes. However, most studies that report introgression fail to disprove the null hypothesis that genetic incongruence may result from stochastic sorting of ancestral allelic polymorphisms. In the case of ancient introgression, these two processes may be especially difficult to distinguish topologically, but they make different predictions about the patterns of coalescence among loci. Here we apply three methods, molecular dating, multispecies coalescent models, and gene tree simulation under coalescence, to compare these two hypotheses that explain the polyphyletic mtDNA of the butterfly peacock bass, Cichla orinocensis. In comparison with a species tree based on 20 unlinked nuclear loci, we determined that mtDNA divergences were too recent to be explained by ancestral polymorphism. Similarly, coalescent species tree branches were significantly shorter when putative introgressed mtDNA was incorporated, and simulations showed the mtDNA topology to be unlikely under lineage sorting only. We conclude that introgression approximately 1.5 million years ago resulted in capture by C. orinocensis of an mtDNA lineage ancestral to the modern subspecies C. oc. monoculus.

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 April 2012 - 31 May 2012: PERMANENT GENETIC RESOURCES NOTE

This article documents the addition of 123 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Brenthis ino, Cichla orinocensis, Cichla temensis, Epinephelus striatus, Gobio gobio, Liocarcinus depurator, Macrolophus pygmaeus, Monilinia vaccinii‐corymbosi, Pelochelys cantorii, Philotrypesis josephi, Romanogobio vladykovi, Takydromus luyeanus and Takydromus viridipunctatus. These loci were cross‐tested on the following species: Cichla intermedia, Cichla ocellaris, Cichla pinima, Epinephelus acanthistius, Gobio carpathicus, Gobio obtusirostris, Gobio sp. 1, Gobio volgensis, Macrolophus costalis, Macrolophus melanotoma, Macrolophus pygmaeus, Romanogobio albipinnatus, Romanogobio banaticus, Romanogobio belingi, Romanogobio kesslerii, Romanogobio parvus, Romanogobio pentatrichus, Romanogobio uranoscopus, Takydromus formosanus, Takydromus hsuehshanesis and Takydromus stejnegeri.

Species-level phylogeny of ‘Satan’s perches’ based on discordant gene trees (Teleostei: Cichlidae: Satanoperca Günther 1862)

Neotropical rivers are home to the largest assemblage of freshwater fishes, but little is known about the phylogeny of these fishes at the species level using multi-locus molecular markers. Here, we present a phylogeny for all known species of the genus Satanoperca, a widespread group of Neotropical cichlid fishes, based on analysis of six unlinked genetic loci. To test nominal and proposed species limits for this group, we surveyed mtDNA sequence variation among 320 individuals representing all know species. Most nominal species were supported by this approach but we determined that populations in the Xingu, Tapajós, and Araguaia+Paraná Rivers are likely undescribed species, while S. jurupari and S. mapiritensis did not show clear genetic distinction. To infer a phylogeny of these putative species, we conducted maximum likelihood and Bayesian non-clock and relaxed clock analyses of concatenated data from three genes (one mitochondrial, two nuclear). We also used a multi-species coalescent model to estimate a species tree from six unlinked loci (one mitochondrial, five nuclear). The topologies obtained were congruent with other results, but showed only minimal to moderate support for some nodes, suggesting that more loci will be needed to satisfactorily estimate the distribution of coalescent histories within Satanoperca. We determined that this variation results from topological discordance among separate gene trees, likely due to differential sorting of ancestral polymorphisms.

Population genetics of the speckled peacock bass (Cichla temensis), South America’s most important inland sport fishery

Abstract The Neotropics harbor the world’s most diverse freshwater fish fauna, with many of these species supporting major commercial, subsistence, or sport fisheries. Knowledge of population genetic structure is available for very few Neotropical fishes, thereby restricting management. To address this need, we examined population genetic variation in mtDNA control region sequences and twelve microsatellite loci in the speckled or barred peacock bass, Cichla temensis. Moderate and statistically significant genetic divergence among localities indicates that migration is low in this species, implying that populations inhabiting tributaries or even smaller spatial units should constitute management units. Analysis of molecular variance of mtDNA sequences identified six areas with largely exclusive haplotype clades, and a seventh area of high admixture, but major drainage basins harbored non-monophyletic haplotype groups. On the other hand, molecular variation in the microsatellite data was best explained by drainage basin and, subsequently, by the seven areas. Populations in these seven areas could be considered evolutionarily significant units (ESUs), and, therefore, we tested hypotheses explaining the discordant signal of mtDNA and microsatellite data using approximate Bayesian computation. This analysis indicated that the divergence of mtDNA clades preceded the divergence of contemporary ESUs across basins, with subsequent lineage sorting among ESUs due to reduced gene flow. Available genetic and ecological information indicates that C. temensis populations of major tributary rivers should be managed as separate stocks that likely are adapted to local environmental conditions.

Multi-locus species tree for the Amazonian peacock basses (Cichlidae: Cichla): Emergent phylogenetic signal despite limited nuclear variation

The inference of phylogenies of closely related species is obstructed by phenomena such as porous species boundaries and deep coalescence, and is often exacerbated by low levels of nucleotide variation among most loci surveyed in phylogenetic studies. We investigated the utility of twenty-one nuclear loci that had a range of 5-40 (median of 14) variable sites per locus to estimate the phylogeny of the genus Cichla, a group of 15 Neotropical cichlid fishes that began to diverge in the early to mid Miocene. We found that under a concatenated approach, the least variable loci, while contributing less to the overall phylogenetic signal (posterior node support), nevertheless provided information that increased support for the final tree. Moreover, this was not a result of misdirection by mutational noise, as the inference from all data was far superior to those from reduced datasets (those with more variable loci) in terms of the relative precision of posterior tree space. Phylogenetic methods that allowed each locus to have a separate genealogy, including Bayesian concordance analysis and a multispecies coalescent model, provided phylogenies that were also compatible with the concatenated tree in terms of the eight recently delimited species of Cichla, albeit with somewhat diminished support for some branches. In contrast, described species that still regularly exchange genes showed unstable relationships among analyses: not a surprising result from analyses that assume that gene tree heterogeneity results from incomplete lineage sorting and not gene flow. Importantly, we also observed that the confidence intervals for node ages in the coalescent analyses were quite wide, and likely susceptible to influence of the prior on node density (e.g. birth-death).