Development and characterization of microsatellite markers for the endangered northern riffleshell mussel Epioblasma torulosa rangiana (Bivalvia: Unionidae)

Primers for six polymorphic microsatellite loci were developed for the endangered northern riffleshell Epioblasma torulosa rangiana from the Sydenham River, Ontario, Canada. These loci along with an additional nine microsatellite loci, developed from other unionoid species, available in the literature, were tested and characterized on individuals from two populations in the Allegheny River; one population in French Creek, PA, USA, and one population from the Sydenham River. Allelic diversity ranged from two to 28 alleles per locus and averaged 13.7 per locus. These primers are being used in a rangewide study on the conservation genetics of remaining extant populations of the northern riffleshell.     

Evaluation of potential health risks to Eastern Elliptio (Elliptio complanata) (Mollusca: Bivalvia: Unionida: Unionidae) and implications for sympatric endangered freshwater mussel species

Conservation efforts for freshwater musselspecies require identification and evaluationof potential health risks to populations. Sampling large numbers of individual mussels,however, could damage the small, fragile,extant populations of imperiled mussel species. In this study, a small sample size was designedto reduce lethal sampling, costs, andenvironmental disruption, while still allowing95% confidence in detecting health risks of25% or greater prevalence in the population. Health assessments were conducted on twentyspecimens of the Eastern Elliptio, Elliptio complanata, collected from two NorthCarolina sites as part of a survey to evaluatepotential disease threats to mussels in theregion. Bacteriological sampling of thegastrointestinal tracts yielded 18 aerobicbacterial species, of which Aeromonashydrophila (55.0%), Enterobacter spp.(40.0%), and Bacillus spp.(30.0%) were predominant. Histologicallesions of internal organs included mild tomoderate digestive gland atrophy andinflammation in one mussel, and mild tomoderate parasitism in several individuals. Adistinct difference in parasite prevalence wasevident between infections in E.complanata from the two collection sites. Thetrematode metacercaria of Homalometronarmatum and what appears to be three gillciliate species, the most abundant beingtentatively identified as the scyphidiidperitrich Mantoscyphidia sp., were foundin mussels from one site only. This studydemonstrates a comprehensive diagnosticapproach incorporating multiple modalities toassess the health status of mussel populations,while minimizing the sample size required toobtain valuable information. Furthermore, thisstudy provides baseline health data of E.complanata at two sites in south-central NorthCarolina and suggests the potential usefulnessof E. complanata as an environmentalbioindicator of health risks to sympatricthreatened freshwater mussel populations.     

Sediment particles as a cause of nacre staining in the freshwater mussel,Amblema plicata (Say) (Bivalvia: Unionidae)

Brown stains sometimes appear in the inner shell layers (nacre) of freshwater mussels. An electron microprobe was used to analyze the stained nacre of the unionid Amblema plicata (Say, 1817) from selected localities on the Mississippi River in the vicinity of LaCrosse and Prairie du Chien, Wisconsin.Several elements such as Na, Mg, Al, Si, P, S, Cl, K, and Fe are more highly concentrated in stained than in unstained nacre. Concentrations of these elements relative to Ca were found to vary significantly among the localities from which the specimens were obtained. Ratios have significantly higher variances downstream of the confluence of the Yellow and Mississippi Rivers, downstream of a barge fleeting area, near the town of Marquette, Iowa, near the site of sewage effluent for Prairie du Chien, Wisconsin and downstream of a scrap metal yard near LaCrosse, Wisconsin in contrast to control localities.Suspended silt, the result of runoff and river activity (barge traffic, dredging, pleasure boating) may be the stimulus for stain formation. Clay minerals adsorb accessory elements and in turbid water are trapped within the pallial space of A. plicata. The mussel secretes an organic-rich, periostracum-like layer over the entrapped sediment, and later reverts to prismatic and finally nacreous shell deposition. Some of the elements found in the stain could directly disturb Ca metabolism by competing with Ca for binding sites in shell aragonite.     

Integrative taxonomy reveals a new species of freshwater mussel,Potamilus streckersoni sp. nov. (Bivalvia: Unionidae): implications for conservation and management

Inaccurate systematics confound our ability to determine evolutionary processes that have led to the diversification of many taxa. The North American freshwater mussel tribe Lampsilini is one of the better-studied groups in Unionidae, however, many supraspecific relationships between lampsiline genera remain unresolved. Two genera previously hypothesized to be non-monophyletic that have been largely overlooked are Leptodea and Potamilus. We set out to resolve supraspecific relationships in Lampsilini and test the monophyly of Leptodea and Potamilus by integrating molecular, morphological, and life history data. Our molecular matrix consisted of four loci: cytochrome c oxidase subunit 1 (CO1), NADH dehydrogenase subunit 1 (ND1), internal transcribed spacer 1 (ITS1), and 28S ribosomal RNA. Secondly, we performed both traditional and Fourier shape morphometric analyses to evaluate morphological differences and finally, we compared our results with available life history data. Molecular data supported the paraphyly of both Leptodea and Potamilus, but nodal support was insufficient to make any conclusions regarding generic-level assignments at this time. In contrast, inference from our integrative taxonomic assessment depicts significant support for the recognition of a new species, Potamilus streckersoni sp. nov., the Brazos Heelsplitter. Our data show clear separation of three taxonomic entities in the P. ohiensis species complex: P. amphichaenus, P. ohiensis, and P. streckersoni sp. nov.; all molecularly, geographically, and morphologically diagnosable. Our findings have profound implications for unionid taxonomy and will aid stakeholders in establishing effective conservation and management strategies.http://www.zoobank.org/urn:lsid:zoobank.org:pub:502647C0-418B-4CC4-85A8-BD89FC3F674F     

Mitochondrial genome of the cockscomb pearl mussel Cristaria plicata (Bivalvia, Unionoida, Unionidae)

The complete mitochondrial genome sequence of the cockscomb pearl mussel Cristaria plicata, which is an endangered species in South Korea, was sequenced. The circle genome (15,708 bp in size) consists of 13 protein-coding genes, 2 rRNA genes, and 22 tRNA genes. There were 26 noncoding regions (NCs) found throughout the mitogenome of C. plicata, ranging in size from 2 to 327 bp, and the two largest NC regions, NC286 and NC326, were found between ND5 and tRNA(Gln) (286 bp) and between tRNA(Glu) and ND2 (326 bp), respectively. The 13 mitochondrial protein-coding genes of a female individual of C. plicata collected from Korea (15,708 bp) were compared to those of the Chinese individual (15,712 bp) published before. The result showed that ND3 is the most conserved with 100% nucleotide similarity, and each of the other protein-coding genes has ca. 99%, respectively. The two largest NCs among 26 NCs have totally 98% nucleotide similarity between Korean and Chinese ones.     

Distribution, abundance, and roles of freshwater clams (Bivalvia, Unionidae) in the freshwater tidal Hudson River

1. An extensive series of PONAR grabs was used to determine the distribution and abundance of unionid clams in the freshwater tidal Hudson River. 2. The five species of unionids collected were distributed very unevenly within the river. Mean river-wide density and biomass of unionids were 8.0m^?2 and 6.2 g DM m^?2 (shell-free)., respectively. 3. The environmental variables that we measured (water depth, distance from shore, sediment granulometry and organic content, presence or absence of macrophytes, and the chlorophyll a and particulate organic matter content of the water) explained little of the variation in abundance of unionids. 4. The distributions of the various species of clams did not differ significantly with respect to the environmental variables measured. 5. We estimate that unionids filter a significant amount of water (0.14m³ m^?2 day^?1, on average) in the Hudson River estuary, roughly equivalent in magnitude to downstream flushing. 6. We project that unionids will serve as a major substratum for the settlement of the zebra mussel (Dreissena polymorpha), which is now invading the estuary. We emphasize that unionids may play important non-trophic roles in large river ecosystems.     

Acquired resistance of bluegill sunfish Lepomis macrochirus to glochidia larvae of the freshwater mussel Utterbackia imbecillis (Bivalvia: Unionidae) after multiple infections

The effects of multiple infections on the host-parasite relationship between bluegill sunfish (Lepomis macrochirus) and parasitic glochidial larvae of the freshwater mussel Utterbackia imbecillis were examined. Na?ve, young-of-the-year bluegills were infected with glochidia and placed in individual observation chambers. Each day, water was drained from each chamber and the numbers of dead glochidia, live glochidia, partially metamorphosed glochidia, and fully metamorphosed juvenile mussels were counted. The same fishes were infected a total of 4 times. After 2 infections, the fish began to exhibit evidence of acquired resistance to glochidia. During the third and fourth infections, this resistance was clearly evidenced by the marked increase in the percentage of dead and live glochidia shed during the first 5 days of the infection and by the significant decrease in the success of metamorphosis. The total number of glochidia that successfully attached to the fish decreased significantly during the fourth infection relative to the first. The number of larvae attached to the host fish was positively correlated with the size of the fish during the first infection but was negatively correlated during all subsequent infections. Variance to mean ratios indicated that larvae were aggregated among host fishes during the infections. This study has important implications in propagation and conservation efforts of this endangered group of organisms.     

Unravelling the systematics of Nodularia (Bivalvia,Unionidae) species from eastern Russia

Conservation efforts have been hindered by data deficient conservation status assessments, especially due to taxonomic problems. This is especially true for many eastern Russian species of freshwater mussels, where distinct classification systems have complicated their delimitation and identification. Nodularia is a widespread eastern Asian freshwater mussel genus, present from Vietnam in the south to the Magadan region in eastern Russia in the north. The number of recognized species in the genus Nodularia in eastern Russia has been inflated over the last several decades due to the use of a typological species concept, the so-called 'Comparatory Method'. This method uses a single diagnostic character for species delimitation, i.e., the arc of maximal convexity of the shell's outline. Using this classification system, 10 species were recognized for far eastern Russia under the genus Nodularia Conrad, 1853, divided into three subgenera: Nodularia s. str., Amurunio and Magadaninaia. Since it is not supported by any other classification methods, the current comparatory classification is rejected by many Russian and international scientists, who only recognize a single species for that region, i.e., Nodularia douglasiae. Therefore, the aim of the present study is to clarify the taxonomy and systematics of the Nodularia genus in far eastern Russia and adjacent territories, using a multiple dataset approach that combines distribution with detailed conchological and anatomical analyses with morphometry and COI barcoding molecular techniques. All analyses performed in this study support the existence of a single Nodularia species in eastern Russia, i.e., N. douglasiae.     

Molecular phylogeny,taxonomy, and distribution of French Unio species (Bivalvia,Unionidae)

A plethora of unionid names was established in the nineteenth century by the “Nouvelle Ecole”. Although naiad morphological plasticity is well documented, the currently recognized fauna, with 17 species and subspecies included in the French checklist for the Unio genus, is still based upon morphological characters only. Insights have been provided from molecular data elsewhere in Europe and North Africa, but the French fauna remains unstudied. We present a molecular phylogeny of the Unio genus in France based on COI, 16S and 28S genes; taking up all available data in Europe plus 273 specimens collected in all main French drainages. The results show that there are either three valid species in France, with U. pictorum and U. mancus synonymized, or five, with the subspecies U. crassus courtillieri elevated to species level. Subspecies were generally not recovered, which questions the evolutionary units tacitly implied by subspecific names. Although sampling topotypes is the most reliable way to evaluate the status of a nominal subspecies, major human-induced changes in aquatic hydrosystems challenge the method. Nevertheless, operational taxonomy has to rely on ground-truthed data and we propose to reduce the actual number of valid taxa in France to the seven observed operational taxonomic units.     

Evaluation of conservation status, distribution, and reproductive characteristics of an endemic Gulf Coast freshwater mussel, Lampsilis australis (Bivalvia: Unionidae)

The southern sandshell, Lampsilis australisSimpson, 1900, is a rare freshwater mussel endemic to the Escambia, Yellow, andChoctawhatchee river drainages in Alabama and Florida. Laboratory observationsconfirmed L. australis as a superconglutinate producer.Superconglutinate and glochidial morphology are similar to those of othersuperconglutinate producers. The current distribution of L.australis, as determined from recent surveys (1995–2000) andfield notes (1990–2000), is compared with its historical distribution asdefined by museum collections and field notes made between 1900 and 1989. Thecurrent distribution of L. australis is reduced by76% compared to its historical distribution, although L. australis waslocated at new sites in the recent surveys. Relative abundance of L.australis was low at all but two sites. Based on reproductivestrategy, range reductions, low relative abundance, and potential threats withinthe Escambia, Yellow, and Choctawhatchee river drainages, we believeL. australis should be considered for protection under theEndangered Species Act.     

Conservation genetics of critically endangered Crepidiastrum grandicollum (Asteraceae) and two closely related woody species of the Bonin Islands,Japan

Crepidiastrum grandicollum is a critically endangered insular endemic herb found only on two oceanic islands of the Bonin Islands in Japan, namely Chichijima Island and Anijima Island. It is explicitly threatened by herbivory pressure from introduced animals. In 2009, a conservation program for C. grandicollum was begun to ensure its future persistence. To provide further information for conservation planning, we investigated the genetic diversity of C. grandicollum using 13 novel microsatellite markers in 55 individuals from four wild populations and an ex situ living collection. Two closely related woody species were also included for interspecific comparison: seven individuals of Crepidiastrum ameristophyllum and 13 of Crepidiastrum linguifolium. The 13 markers were applicable to all three species and identified 129 alleles in total. We found a clear genetic differentiation between C. grandicollum from Anijima Island and Chichijima Island. Crepidiastrum grandicollum also had low expected heterozygosity and allelic richness in populations compared to the two closely related species. Reconstructed divergence history suggested that differentiation between the islands had occurred several thousand generations ago. We suggest separate conservation units for C. grandicollum on Anijima Island and Chichijima Island given the clear (and putatively historical) genetic differentiation, which may result in speciation in the future.

     

Shape variation in the Australian freshwater mussel Alathyria jacksoni Iredale (Bivalvia,Hyriidae)

Different forms of the hydriid mussel Alathyria jacksoni occur in riverine habitats of the Murray-Darling river system, south-eastern Australia. Electrophoretic data for three enzymes (glucose phosphate isomerase, malate dehydrogenase and malic enzyme) support the assumption that these are morphological variants of one species. Shells from the Murray in South Australia are elongate-oval, with the dorsal margin extended posteriorly as a blade or wing, whereas shells from Victoria and New South Wales have a dorsal arch and a postero-ventral inflection. The variation appears to be associated with water velocity. The arched shells have comparatively large, strong adductor muscles; the posterior adductors are elongate and both anterior and posterior adductors are displaced ventrally. These internal modifications are reflected in changing shell shape as size increases. The changes apparently enhance the mussel's ability to maintain an anchorage in a strong current, but lessen the effectiveness of its valve seal. An incidental observation is that the related species A. condola Iredale, known from the Murrumbidgee and Lachlan rivers, may also occur in the middle reaches of the Murray.     

Extensive allozyme monomorphism in a threatened species of freshwater mussel, Margaritifera hembeli Conrad (Bivalvia: Margaritiferidae)

A threatened but under-studied component offreshwater biodiversity in North America is thenative freshwater mussels (Bivalvia:Unionoidea: Margaritiferidae and Unionidae). Genetic data suggest that these mussel speciesgenerally exhibit levels of variability similarto other invertebrates. We surveyed allozymevariation in the Louisiana Pearlshell, Margaritifera hembeli (Margaritiferidae), athreatened freshwater mussel. Five examinedpopulations are monomorphic for 25 allozymeloci, the first report of a native freshwatermussel species with extensive allozymemonomorphism. Low genetic diversity appears tobe characteristic of margaritiferids, as anANOVA indicated that mussels of the familyMargaritiferidae have significantly lowerlevels of heterozygosity than the mussels ofthe family Unionidae. Margaritiferids havestrong habitat preferences and modification ofhabitat leads to rapid loss of populations.Although bottlenecks are known to cause lowgenetic variability, margaritiferids mayexhibit meta-population structure withextinction/re-colonization dynamics leading tolow genetic variability. Margaritiferidsgenerally exhibit a patchy distribution with acolonization rate that is approximately twicethat of extinction. Tests of themetapopulation hypothesis will requireadditional allozyme population genetic data aswell as hypervariable microsatellite loci.     

Population size and structure of three mussel species (Bivalvia: Unionidae) in a northeastern German river with special regard to influences of environmental factors

Unionid mussels are of great ecological importance in running and standing waters, however their populations declined continually during the 20th century. In order to collect more data on the situation of these mussels in running waters, the populations of Anodonta anatina, Anodonta cygnea, and Unio tumidus (Unionidae) were investigated in the northeastern German river Ryck from 1996 to 1998. At three sampling stations along the river, the mussel stocks as well as sediment and water properties were analyzed. In this river abundances of up to 138.7 indiv./m² were detected. The average population densities varied from 10.9 to 34.9 indiv./m². More critically however, the age structures showed significant signs of irregular reproductive success and overaging. Consequently, the mussel stocks are going to decline, and furthermore, U.tumidus should be listed as an endangered species in the Ryck river. Negative influences on bivalves can be expected from sporadically occurring low oxygen and high ion concentrations in the water. But after all, the conditions within the sediments seem to be decisive for the survival of juvenile mussels and thus for the age structure of the populations.     

Complete mtDNA sequence of the North American freshwater mussel,Lampsilis ornata (Unionidae): an examination of the evolution and phylogenetic utility of mitochondrial genome organization in Bivalvia (Mollusca)

Molluscs in general, and bivalves in particular, exhibit an extraordinary degree of mitochondrial gene order variation when compared with other metazoans. Two factors inhibiting our understanding the evolution of gene rearrangement in bivalves are inadequate taxonomic sampling and failure to examine gene order in a phylogenetic framework. Here, we report the first complete nucleotide sequence (16,060 bp) of the mitochondrial (mt) genome of a North American freshwater bivalve, Lampsilis ornata (Mollusca: Paleoheterodonta: Unionidae). Gene order and mt genome content is examined in a comparative phylogenetic framework for Lampsilis and five other bivalves, representing five families. Mitochondrial genome content is shown to vary by gene duplication and loss among taxa and between male and female mitotypes within a species. Although mt gene arrangement is highly variable among bivalves, when optimized on an independently derived phylogenetic hypothesis, it allows for the reconstruction of ancestral gene order states and indicates the potential phylogenetic utility of the data. However, the interpretation of reconstructed ancestral gene order states must take in to account both the accuracy of the phylogenetic estimation and the probability of character state change across the topology, such as the presence/absence of atp8 in bivalve lineages. We discuss what role, if any, doubly uniparental inheritance (DUI) and recombination between sexual mitotypes may play in influencing gene rearrangement of the mt genome in some bivalve lineages.     

Population genetics and phylogeography of freshwater mussels in North America, Elliptio dilatata and Actinonaias ligamentina (Bivalvia: Unionidae)

Extrinsic and intrinsic forces combined shape the population structure of every species differently. Freshwater mussels are obligate parasites to a host fish during a juvenile stage (glochidia). Elliptio dilatata (ED) and Actinonaias ligamentina (AL) are co-occurring freshwater mussel taxa with similar North American distribution and share some potential host fish. Using mitochondrial DNA, we determined the genotypes of 190 + individuals from collection sites in at least two tributaries in the Lake Erie and Ohio River watersheds, along with the Ouachita and Strawberry rivers in the southeast. Both species had followed a stepping-stone model of dispersal, with greater pairwise genetic structure among collection sites of ED. Also, phylogeographical analysis for ED found significant geographical structuring of haplotype diversity. Overall, within-population variation increased significantly from north to south, with low genetic diversity in the Strawberry River. We calculated significant among-population structure for both species (ED: Phi(ST) = 0.62, P < 0.001; AL: Phi(ST) = 0.16, P < 0.001). Genetic analysis identified the Ouachita River as an area of significant reproductive isolation for both species. Results for AL indicated dispersal into northern areas from two genetically distinct glacial refugia, where results for ED indicated dispersal followed by low gene flow in northern areas. The conservation strategies for mussels that co-occur in the same 'bed' could be species specific. Species such as ED have management units on the population scale, where AL has a more homogeneous genetic structure across its range.     

The intraspecific taxonomic structure and identification of closely related species of ixodid ticks (Ixodidae)

Two variants of intraspecific morphological differentiation of 7 palearctic species of Ixodidae and respectively two variants of their intraspecific taxonomic pattern have been revealed on the basis of the study of geographic variation of each species. Three species--Ixodes pavlovskyi Pom., 1946, Haemaphysalis erinacei Pavesi, 1884, Hyalomma asiaticum Sch. et Schl., 1929, are subdivided into subspecies, and four ones--I. ricinus (L., 1758), I. persulcatus Sch., 1930, I. crenulatus Koch, 1844, Dermacentor marginatus (Sulzer, 1776),--into morphotypes by the degree of distinction of the whole complex of active stages in ontogenesis. Acknowledging the conventional character of the categories of subspecies and morphotype their criteria have been formulated for ixodid ticks. Paleogeographic and ecological grounds are given. Some data of geographic variation of immature stages favour the identification of closely related species in concrete localities.     

Reach- and catchment-scale determinants of the distribution of freshwater mussels (Bivalvia: Unionidae) in south-eastern Michigan, U.S.A.

1. We investigated the diversity and distribution of freshwater mussels at 40 sites in an agricultural catchment, the River Raisin in south‐eastern Michigan, to relate mussel assemblages and individual taxa to reach and catchment‐scale variables. Unionids were surveyed by timed searches in 100‐m reaches, and in‐stream and riparian habitat were quantified as well as flow, water chemistry and channel morphology. Land use/cover and surficial geology were determined for site subcatchments and riparian buffers. 2. Some 21 mussel species were found overall; richness ranged from 0 to 12 living species per site. From the upper to middle to lower catchment, the number of individuals, number of species, Shannon–Weaver diversity and relative abundance of intolerant unionids all declined significantly. 3. Four groupings based on overall mussel diversity and abundance were significantly related to reach‐scale habitat variables. The richest mussel assemblages were associated with sites with higher overall habitat quality, greater flow stability, less fine substratum, and lower specific conductance. 4. Stepwise multiple regressions revealed that the distribution and abundance of the total mussel assemblage, as well as the most common species, could be predicted from a combination of reach‐ and catchment‐scale variables (R² = 0.63 for total mussels, R² = 0.51–0.86 for individual species). 5. Flow stability, substratum composition and overall reach habitat quality were the most commonly identified reach‐scale variables, and measures of surficial geology were the most effective catchment‐scale variables. The spatial pattern of geology is likely to be responsible for the diversity gradient from the upper to the lower catchment. 6. Prior studies, attempting to explain mussel distributions from local habitat features alone, have found relatively weak relationships. By employing a combination of reach‐ and catchment‐scale habitat variables, this study was able to account for a substantial amount of the spatial variability in mussel distributions.