Riverine flow rate drives widespread convergence in the shell morphology of imperiled freshwater mussels.

Frequent and strong morphological convergence suggests that determinism tends to supersede historical contingencies in evolutionary radiations. For many lineages living within the water column of rivers and streams, hydrodynamic forces drive widespread morphological convergence. Living below the sediment-water interface may release organisms from these hydrodynamic pressures, permitting a broad array of morphologies, and thus less convergence. However, we show here that the semi-infaunal freshwater mussels have environmentally determined convergence in shell morphology. Using 3D morphometric data from 715 individuals among 164 Nearctic species, we find that species occurring in rivers with high flow rates have evolved traits that resist dislodgement from their burrowed position in the streambed: thicker shells for their body size, with the thickest sector of the shell being the most deeply buried. Species occurring in low flow environments have evolved thinner and more uniformly thickened shells, corresponding to an alternative adaptation to dislodgement: increased burrowing efficiency. Within species, individuals also show increased shell thickness for their body size at higher flow rates, suggesting that ecophenotypy may, in part, be an important mechanism for establishing populations in new environments and thus evolutionary divergence in this highly imperiledinvertebrate group.

A trait dataset for freshwater mussels of the United States of America.

The United States of America has a diverse collection of freshwater mussels comprising 301 species distributed among 59 genera and two families (Margaritiferidae and Unionidae), each having a unique suite of traits. Mussels are among the most imperilled animals and are critical components of their ecosystems, and successful management, conservation and research requires a cohesive and widely accessible data source. Although trait-based analysis for mussels has increased, only a small proportion of traits reflecting mussel diversity in this region has been collated. Decentralized and non-standardized trait information impedes large-scale analysis. Assembling trait data in a synthetic dataset enables comparison across species and lineages and identification of data gaps. We collated data from the primary literature, books, state and federal reports, theses and dissertations, and museum collections into a centralized dataset covering information on taxonomy, morphology, reproductive ecology and life history, fish hosts, habitats, thermal tolerance, geographic distribution, available genetic information, and conservation status. By collating these traits, we aid researchers in assessing variation in mussel traits and modelling ecosystem change.

A novel assembly pipeline and functional annotations for targeted sequencing: A case study on the globally threatened Margaritiferidae (Bivalvia: Unionida).

The proliferation of genomic sequencing approaches has significantly impacted the field of phylogenetics. Target capture approaches provide a cost-effective, fast and easily applied strategy for phylogenetic inference of non-model organisms. However, several existing target capture processing pipelines are incapable of incorporating whole genome sequencing (WGS). Here, we develop a new pipeline for capture and de novo assembly of the targeted regions using whole genome re-sequencing reads. This new pipeline captured targeted loci accurately, and given its unbiased nature, can be used with any target capture probe set. Moreover, due to its low computational demand, this new pipeline may be ideal for users with limited resources and when high-coverage sequencing outputs are required. We demonstrate the utility of our approach by incorporating WGS data into the first comprehensive phylogenomic reconstruction of the freshwater mussel family Margaritiferidae. We also provide a catalogue of well-curated functional annotations of these previously uncharacterized freshwater mussel-specific target regions, representing a complementary tool for scrutinizing phylogenetic inferences while expanding future applications of the probe set.

A tale of two paths: The evolution of mitochondrial recombination in bivalves with doubly uniparental inheritance.

In most animals, mitochondrial DNA is strictly maternally inherited and non-recombining. One exception to this pattern is called doubly uniparental inheritance (DUI), a phenomenon involving the independent transmission of female and male mitochondrial genomes. DUI is known only from the molluskan class Bivalvia. The phylogenetic distribution of male-transmitted mitochondrial DNA (M mtDNA) in bivalves is consistent with several evolutionary scenarios, including multiple independent gains, losses, and varying degrees of recombination with female-transmitted mitochondrial DNA (F mtDNA). In this study, we use phylogenetic methods to test M mtDNA origination hypotheses and infer the prevalence of mitochondrial recombination in bivalves with DUI. Phylogenetic modeling using site concordance factors supported a single origin of M mtDNA in bivalves coupled with recombination acting over long evolutionary timescales. Ongoing mitochondrial recombination is present in Mytilida and Venerida, which results in a pattern of concerted evolution of F mtDNA and M mtDNA. Mitochondrial recombination could be favored to offset the deleterious effects of asexual inheritance and maintain mitonuclear compatibility across tissues. Cardiida and Unionida have gone without recent recombination, possibly due to an extension of the COX2 gene in male mitochondrial DNA. The loss of recombination could be connected to the role of M mtDNA in sex determination or sexual development. Our results support that recombination events may occur throughout the mitochondrial genomes of DUI species. Future investigations may reveal more complex patterns of inheritance of recombinants, which could explain the retention of signal for a single origination of M mtDNA in protein-coding genes.

New freshwater mussels from two Southeast Asian genera Bineurus and Thaiconcha (Pseudodontini, Gonideinae, Unionidae).

The Mekong and Chao Phraya rivers harbor a species-rich freshwater mussel assemblage containing a large radiation of the Pseudodontini species. Members of the genera Bineurus Simpson 1900 and Thaiconcha Bolotov et al., 2020 primarily inhabit small and medium-sized tributaries of these rivers. Here, we present an integrative taxonomic review of these genus-level clades. We show that Bineurus contains four species: B. mouhotii (Lea, 1863), B. exilis (Morelet, 1866) stat. rev., B. anodontinum (Rochebrune, 1882) stat. rev., and B. loeiensis sp. nov. In its turn, Thaiconcha comprises three species: T. callifera (Martens, 1860), T. munelliptica sp. nov., and T. thaiensis sp. nov. Two species, Pseudodon ovalis Morlet, 1889 and P. thomsoni Morlet, 1884, are considered here as questionable taxa. These findings further highlight that Southeast Asia represents a significant evolutionary hotspot of freshwater mussels, which requires further international collaborative research and conservation efforts.

Evolutionary history drives aspects of stoichiometric niche variation and functional effects within a guild.

Functional traits are characteristics of an organism that represents how it interacts with its environment and can influence the structure and function of ecosystems. Ecological stoichiometry provides a framework to understand ecosystem structure and function by modeling the coupled flow of elements (e.g. carbon [C], nitrogen [N], phosphorus [P]) between consumers and their environment. Animals tend to be homeostatic in their nutrient requirements and preferentially sequester the element in shortest supply relative to demand, and release relatively more of the element in excess. Tissue stoichiometry is an important functional trait that allows for predictions among the elemental composition of animals, their diet, and their waste products, with important effects on the cycling and availability of nutrients in ecosystems. Here, we examined the tissue stoichiometric niches (C:N:P) and nutrient recycling stoichiometries (N:P) of several filter-feeding freshwater mussels in the subfamily Ambleminae. Despite occupying the same functional-feeding group and being restricted to a single subfamily-level radiation, we found that species occupied distinct stoichiometric niches and that these niches varied, in part, as a function of their evolutionary history. The relationship between phylogenetic divergence and functional divergence suggests that evolutionary processes may be shaping niche complementarity and resource partitioning. Tissue and excretion stoichiometry were negatively correlated as predicted by stoichiometric theory. When scaled to the community, higher species richness and phylogenetic diversity resulted in greater functional evenness and reduced functional dispersion. Filter-feeding bivalves are an ecologically important guild in freshwater ecosystems globally, and our study provides a more nuanced view of the stoichiometric niches and ecological functions performed by this phylogenetically and ecologically diverse assemblage.

Comparative phylogenomics reveal complex evolution of life history strategies in a clade of bivalves with parasitic larvae (Bivalvia: Unionoida: Ambleminae).

Freshwater mussels are a species-rich group with biodiversity patterns strongly shaped by a life history strategy that includes an obligate parasitic larval stage. In this study, we set out to reconstruct the life history evolution and systematics in a clade of freshwater mussels adapted to parasitizing a molluscivorous host fish. Anchored hybrid enrichment and ancestral character reconstruction revealed a complex pattern of life history evolution with host switching and multiple instances of convergence, including reduction in size of larvae, increased fecundity, and growth during encapsulation. Our phylogenomic analyses also recovered non-monophyly of taxa exhibiting multiple traits used as the basis for previous taxonomic hypotheses. Taxa with axe-head shaped glochidia were resolved as paraphyletic, but our results strongly suggest the complex morphology is an adaptation to reduce larval size, with reduction in size further accentuated in taxa previously assigned to Leptodea. To more accurately reflect the evolutionary history of this group, we make multiple systematic changes, including the description of a new genus, Atlanticoncha gen. nov., and the synonymy of the genus Leptodea under Potamilus. Our findings contribute to the growing body of literature showing that cladistic hypotheses based solely on morphological characters, including larval morphology, can be flawed in freshwater mussels.

A new genus and two new species of freshwater mussels (Unionidae) from western Indochina.

The systematics of Oriental freshwater mussels (Bivalvia: Unionidae) is poorly known. Here, we present an integrative revision of the genus Trapezoideus Simpson, 1900 to further understanding of freshwater mussel diversity in the region. We demonstrate that Trapezoideus as currently circumscribed is non-monophyletic, with its former species belonging to six other genera, one of which is new to science and described here. We recognize Trapezoideus as a monotypic genus, comprised of the type species, T. foliaceus. Trapezoideus comptus, T. misellus, T. pallegoixi, and T. peninsularis are transferred to the genus Contradens, T. subclathratus is moved to Indonaia, and T. theca is transferred to Lamellidens. Trapezoideus prashadi is found to be a junior synonym of Arcidopsis footei. Trapezoideus dallianus, T. nesemanni, T. panhai, T. peguensis, and two species new to science are placed in Yaukthwa gen. nov. This genus appears to be endemic of the Western Indochina Subregion. The two new species, Yaukthwa paiensis sp. nov. and Y. inlenensis sp. nov., are both endemic to the Salween River basin. Our results highlight that Southeast Asia is a species-rich freshwater mussel diversity hotspot with numerous local endemic species, which are in need of special conservation efforts.

Unioverse: A phylogenomic resource for reconstructing the evolution of freshwater mussels (Bivalvia, Unionoida).

Freshwater mussels (order Unionoida) are a diverse radiation of parasitic bivalves that require temporary larval encystment on vertebrate hosts to complete metamorphosis to free-living juveniles. The freshwater mussel-fish symbiosis represents a useful relationship for understanding eco-evolutionary dynamics in freshwater ecosystems but the practicality of this promising model system is undermined by the absence of a stable freshwater mussel phylogeny. Inadequate character sampling is the primary analytical impediment obfuscating a coherent phylogeny of freshwater mussels, specifically the lack of nuclear molecular markers appropriate for reconstructing supraspecific relationships and testing macroevolutionary hypotheses. The objective of this study is to develop a phylogenomic resource, specifically an anchored hybrid enrichment probe set, capable of capturing hundreds of molecular markers from taxa distributed across the entirety of freshwater mussel biodiversity. Our freshwater mussel specific anchored hybrid enrichment probe set, called Unioverse, successfully captures hundreds of nuclear protein-coding loci from all major lineages of the Unionoida and will facilitate more data-rich and taxonomically inclusive reconstructions of freshwater mussel evolution. We demonstrate the utility of this resource at three disparate evolutionary scales by estimating a backbone phylogeny of the Bivalvia with a focus on the Unionoida, reconstructing the subfamily-level relationships of the Unionidae, and recovering the systematic position of the phylogenetically unstable genus Plectomerus.

Integrative taxonomy resolves taxonomic uncertainty for freshwater mussels being considered for protection under the U.S. Endangered Species Act.

Objectively delimiting species boundaries remains an important challenge in systematics and becomes urgent when unresolved taxonomy complicates conservation and recovery efforts. We examined species boundaries in the imperiled freshwater mussel genus Cyclonaias (Bivalvia: Unionidae) using morphometrics, molecular phylogenetics, and multispecies coalescent models to help guide pending conservation assessments and legislative decisions. Congruence across multiple lines of evidence indicated that current taxonomy overestimates diversity in the C. pustulosa species complex. The only genetically and morphologically diagnosable species in the C. pustulosa species complex were C. pustulosa and C. succissa and we consider C. aurea, C. houstonensis, C. mortoni, and C. refulgens to be synonyms of C. pustulosa. In contrast, all three species in the C. nodulata complex (C. necki, C. nodulata, and C. petrina) were genetically, geographically, and morphologically diagnosable. Our findings have important conservation and management implications, as three nominal species (C. aurea, C. houstonensis, and C. petrina) are being considered for protection under the Endangered Species Act.

A new genus and tribe of freshwater mussel (Unionidae) from Southeast Asia.

The freshwater mussel genus Oxynaia Haas, 1911 is thought to be comprised of two geographically disjunct and morphologically variable species groups but the monophyly of this taxon has yet to be tested in any modern cladistic sense. This generic hypothesis has important systematic and biogeographic implications as Oxynaia is the type genus of the currently recognized tribe Oxynaiini (Parreysiinae) and is one of the few genera thought to cross several biogeographically important barriers in Southeast Asia. Morphological and molecular data clearly demonstrate that Oxynaia is not monophyletic, and the type species and its allies (O. jourdyi group) belong to the Unioninae, and more specifically as members of the genus Nodularia Conrad, 1853. Therefore, neither Oxynaia syn. nov. nor Oxynaiini Starobogatov, 1970 are applicable to the Parreysiinae and in the absence of an available name, Indochinella gen. nov. and Indochinellini trib. nov. are described. Several combinations are proposed as follows: Indochinella pugio (Benson, 1862) gen. et comb. nov., Nodularia jourdyi (Morlet, 1886) comb. res., N. gladiator (Ancey, 1881) comb. res., N. diespiter (Mabille, 1887) comb. res. and N. micheloti (Morlet, 1886) comb. res. Finally, we provide an updated freshwater biogeographic division of Southeast Asia.

Molecular and morphological data to facilitate future research on freshwater mussels (Bivalvia: Unionidae: Anodontinae).

This data article presents the multi-locus DNA alignments, morphometric data, and details on specimens examined to resolve the evolutionary history of Anodontoides and Strophitus, primarily generic placement and species boundaries. We sequenced 3 loci to create our molecular matrix: cytochrome c oxidase subunit I, NADH dehydrogenase subunit I, and the nuclear-encoded ribosomal internal transcribed spacer I. Aligned sequences were used in phylogenetic analyses and to identify diagnostic nucleotides for Strophitus pascagoulaensis, Strophitus radiatus, Strophitus sp. cf. pascagoulaensis, and Strophitus williamsi. Linear morphometrics (i.e. maximum height, length, and width) were also implemented to further evaluate species boundaries within Strophitus. For further details and experimental findings, please refer to the article published in Molecular Phylogenetics and Evolution (Smith et al., 2018) [1].

Molecular and morphological data reveal non-monophyly and speciation in imperiled freshwater mussels (Anodontoides and Strophitus).

Accurate taxonomic placement is vital to conservation efforts considering many intrinsic biological characteristics of understudied species are inferred from closely related taxa. The rayed creekshell, Anodontoides radiatus (Conrad, 1834), exists in the Gulf of Mexico drainages from western Florida to Louisiana and has been petitioned for listing under the Endangered Species Act. We set out to resolve the evolutionary history of A. radiatus, primarily generic placement and species boundaries, using phylogenetic, morphometric, and geographic information. Our molecular matrix contained 3 loci: cytochrome c oxidase subunit I, NADH dehydrogenase subunit I, and the nuclear-encoded ribosomal internal transcribed spacer I. We employed maximum likelihood and Bayesian inference to estimate a phylogeny and test the monophyly of Anodontoides and Strophitus. We implemented two coalescent-based species delimitation models to test seven species models and evaluate species boundaries within A. radiatus. Concomitant to molecular data, we also employed linear morphometrics and geographic information to further evaluate species boundaries. Molecular and morphological evidence supports the inclusion of A. radiatus in the genus Strophitus, and we resurrect the binomial Strophitus radiatus to reflect their shared common ancestry. We also found strong support for polyphyly in Strophitus and advocate the resurrection of the genus Pseudodontoideus to represent 'Strophitus' connasaugaensis and 'Strophitus' subvexus. Strophitus radiatus exists in six well-supported clades that were distinguished as evolutionary independent lineages using Bayesian inference, maximum likelihood, and coalescent-based species delimitation models. Our integrative approach found evidence for as many as 4 evolutionary divergent clades within S. radiatus. Therefore, we formally describe two new species from the S. radiatus species complex (Strophitus williamsi and Strophitus pascagoulaensis) and recognize the potential for a third putative species (Strophitus sp. cf. pascagoulaensis). Our findings aid stakeholders in establishing conservation and management strategies for the members of Anodontoides, Strophitus, and Pseudodontoideus.

Revision of the horseface loaches (Cobitidae, Acantopsis), with descriptions of three new species from Southeast Asia.

Acantopsis (Cobitidae) is revised based on analysis of morphological and molecular data. Four of the six available names, A. dialuzona, A. spectabilis, A. octoactinotos, and A. thiemmedhi, are valid, and three new species, A. rungthipae, A. dinema, and A. ioa, are described. All species are described morphologically, distributions are mapped, and relationships are discussed for those for which molecular data (CO1, RAG1) are available. Labial barbels, color pattern, and meristic counts are the most diagnostic features. Although the long snout of Acantopsis is perhaps the most emblematic attribute of the genus, its relative length increases with growth, reducing its taxonomic value. Species can be difficult to identify on the basis of color pattern alone, as habitat and preservation methods appear to strongly influence the color pattern. Despite interspecific overlap of some highly variable traits, each species has a unique set of morphological characteristics that remain observable even when the color pattern is obscured, and some species are restricted to single drainages, greatly simplifying identification. The phylogenetic analyses revealed high molecular divergence between even the most morphologically similar species, with mean uncorrected CO1 p-distances between species ranging from 12.1-15.4%. Species of Acantopsis exhibit significant genetic structuring consistent with recognized freshwater ecoregions. Acanthopsis lachnostoma Rutter 1897, from Swatow, China, is not assignable to Acantopsis.

Phylogeny of the most species-rich freshwater bivalve family (Bivalvia: Unionida: Unionidae): Defining modern subfamilies and tribes.

Freshwater mussels of the order Unionida are key elements of freshwater habitats and are responsible for important ecological functions and services. Unfortunately, these bivalves are among the most threatened freshwater taxa in the world. However, conservation planning and management are hindered by taxonomic problems and a lack of detailed ecological data. This highlights the urgent need for advances in the areas of systematics and evolutionary relationships within the Unionida. This study presents the most comprehensive phylogeny to date of the larger Unionida family, i.e., the Unionidae. The phylogeny is based on a combined dataset of 1032bp (COI+28S) of 70 species in 46 genera, with 7 of this genera being sequenced for the first time. The resulting phylogeny divided the Unionidae into 6 supported subfamilies and 18 tribes, three of which are here named for the first time (i.e., Chamberlainiini nomen novum, Cristariini nomen novum and Lanceolariini nomen novum). Molecular analyses were complemented by investigations of selected morphological, anatomical and behavioral characters used in traditional phylogenetic studies. No single morphological, anatomical or behavioral character was diagnostic at the subfamily level and few were useful at the tribe level. However, within subfamilies, many tribes can be recognized based on a subset of these characters. The geographical distribution of each of the subfamilies and tribes is also presented. The present study provides important advances in the systematics of these extraordinary taxa with implications for future ecological and conservation studies.

Molecular phylogenetic analysis supports a Gondwanan origin of the Hyriidae (Mollusca: Bivalvia: Unionida) and the paraphyly of Australasian taxa.

The freshwater mussel family Hyriidae (Mollusca: Bivalvia: Unionida) has a disjunct trans-Pacific distribution in Australasia and South America. Previous phylogenetic analyses have estimated the evolutionary relationships of the family and the major infra-familial taxa (Velesunioninae and Hyriinae: Hyridellini in Australia; Hyriinae: Hyriini, Castaliini, and Rhipidodontini in South America), but taxon and character sampling have been too incomplete to support a predictive classification or allow testing of biogeographical hypotheses. We sampled 30 freshwater mussel individuals representing the aforementioned hyriid taxa, as well as outgroup species representing the five other freshwater mussel families and their marine sister group (order Trigoniida). Our ingroup included representatives of all Australian genera. Phylogenetic relationships were estimated from three gene fragments (nuclear 28S, COI and 16S mtDNA) using maximum parsimony, maximum likelihood, and Bayesian inference, and we applied a Bayesian relaxed clock model calibrated with fossil dates to estimate node ages. Our analyses found good support for monophyly of the Hyriidae and the subfamilies and tribes, as well as the paraphyly of the Australasian taxa (Velesunioninae, (Hyridellini, (Rhipidodontini, (Castaliini, Hyriini)))). The Hyriidae was recovered as sister to a clade comprised of all other Recent freshwater mussel families. Our molecular date estimation supported Cretaceous origins of the major hyriid clades, pre-dating the Tertiary isolation of South America from Antarctica/Australia. We hypothesize that early diversification of the Hyriidae was driven by terrestrial barriers on Gondwana rather than marine barriers following disintegration of the super-continent.