A new species and range extensions for three other species of pebblesnails (Lithoglyphidae, Fluminicola) from the upper Klamath basin, California-Oregon.

This is the fifth in a recent series of papers on the poorly known western North American pebblesnail genus Fluminicola (Caenogastropoda, Lithoglyphidae). Herein we clarify the taxonomic status of the currently undescribed pebblesnail fauna in the upper Klamath River drainage (UKL) based on morphologic evidence, and mitochondrial DNA sequence data from 58 UKL collection localities. We describe one new species (F.klamathensis) from eight UKL localities which is differentiated by mtDNA sequences and unique penial morphology, and document range extensions to the UKL for three species from closely proximal drainages (F.fresti, F.modoci, F.multifarius). Fluminicolafresti was found at a single locality along the western edge of upper Klamath Lake. Fluminicolamodoci and F.multifarius are widely distributed in the UKL; both species exhibit marked morphologic variation yet are relatively little differentiated genetically in this basin.

Two new species of Fluminicola (Caenogastropoda, Lithoglyphidae) from southwest Oregon, USA, and a range extension for F. multifarius.

We describe two new species of pebblesnails (Lithoglyphidae: Fluminicola) from southwest Oregon based on morphologic and mitochondrial DNA (COI, cytB) evidence. Fluminicola umpquaensissp. n., which had been traditionally identified as F. virens prior to the recent restriction of the latter to the lower Columbia River drainage, lives in lotic habitats in the Umpqua River basin. This species is readily distinguished from closely related F. gustafsoni and F. virens by shell and anatomical characters, and by its mtDNA sequences (divergence >3.6% for both genes). Fluminicola frestisp. n. ranges among lotic habitats in the North Umpqua River basin, and in the upper Rogue River drainage north of Little Butte Creek. This species differs from other congeners by >9.1% for both genes and is distinguished from closely similar and geographically proximal F. multifarius by several anatomical characters. Additionally, new records are provided for F. multifarius from the upper Rogue River basin south of Little Butte Creek, which extend the geographic range of this species about 80 km northward from the Sacramento River headwater region. This continues a recent series of taxonomic papers on the poorly known and little studied pebblesnail fauna of the vast Pacific watershed from northern California to southern British Columbia.

Three new species of western California springsnails previously confused with Pyrgulopsis stearnsiana (Caenogastropoda, Hydrobiidae).

We describe three new, allopatric species of springsnails (genus Pyrgulopsis) from western California (Pyrgulopsis lindae, Pyrgulopsis ojaiensis, Pyrgulopsis torrida) that were previously identified as Pyrgulopsis stearnsiana. The new species are differentiated from Pyrgulopsis stearnsiana and each other both by mtCOI sequences (3.9-9.9%) and details of penial morphology. We also provide a phylogeny with increased sampling which confirms a previous finding that Pyrgulopsis stearnsiana sensu stricto is paraphyletic relative to two other California species (Pyrgulopsis diablensis, Pyrgulopsis giulianii). Our molecular and morphological evidence suggests that Pyrgulopsis stearnsiana paraphyly is an artifact of conservative taxonomy, however additional studies utilizing rapidly evolving genetic markers will be needed to confidently tease apart the cryptic diversity in this widely ranging springsnail. The new species described herein are narrowly distributed and vulnerable to anthropogenic stressors. The single known population of Pyrgulopsis torrida may have become extirpated between 2000 and 2015.

Taxonomic status of the Columbia duskysnail (Truncatelloidea, Amnicolidae, Colligyrus).

Undescribed freshwater snails (Amnicolidae: Colligyrus) from the Mount Hood region (northwestern United States) identified as a new species (commonly known as the Columbia duskysnail) in grey literature have been provided federal protection under the "survey and manage" provisions of the Northwest Forest Plan and have been placed on conservation watch lists. However, there are no published studies of the identity of these snails aside from a molecular phylogenetic analysis which delineated a close relationship between the single sampled population and Colligyrusgreggi, which is distributed more than 750 km to the east of the Mount Hood area. Here we examine the taxonomic status of the Columbia duskysnail based on additional molecular sampling of mitochondrial DNA sequences (COI) and morphological evidence. We found that the Columbia duskysnail is not a monophyletic group and forms a strongly supported clade with Colligyrusgreggi. The COI divergence between these broadly disjunct groups (2.1%) was somewhat larger than that within Colligyrusgreggi (1.0%) but considerably less than that among the three currently recognized species of Colligyrus (8.7-12.1%). Additionally we found that the Columbia duskysnail and Colligyrusgreggi cannot be consistently differentiated by previously reported diagnostic characters (size and shape of shell spire, pigmentation of body and penis) and are closely similar in other aspects of morphology. Based on these results we conclude that the Columbia duskysnail is conspecific with Colligyrusgreggi.

Assembly of a micro-hotspot of caenogastropod endemism in the southern Nevada desert, with a description of a new species of Tryonia (Truncatelloidea, Cochliopidae).

Newly obtained and previously published sequences of the cytochrome c oxidase subunit I (COI) gene were analyzed to examine the biogeographic assembly of the caenogastropod fauna (belonging to the families Assimineidae, Cochliopidae, and Hydrobiidae) of an isolated spring along the lower Colorado River in southern Nevada (Blue Point Spring). Based on available COI clock calibrations, the three lineages that comprise this fauna are 2.78-1.42 million years old, which is roughly coeval or slightly younger than the age of Blue Point Spring (inferred from local fossil spring deposits). Two of the lineages-endemic Pyrgulopsiscoloradensis and Assimineaaff.infima-are most closely related to snails in the Death Valley area (well to the west) and likely colonized Blue Point Spring by transport on birds. A single haplotype was detected in both of these snails, suggesting that they may have only recently colonized Blue Point Spring. The third lineage-endemic Tryoniainfernalis, newly described herein based on morphological and molecular evidence-is most closely related to a geographically proximal species in a lower Colorado River tributary (Tryoniaclathrata); the split between these taxa may be the product of vicariance (severance of a prior drainage connection) or a separate jump dispersal event. The considerable genetic diversity in Tryoniainfernalis (three haplotypes differing by 0.6% mean sequence divergence) suggests a possibly lengthy history of local differentiation. Our findings also identify Blue Point Spring as a new micro-hotspot of groundwater-dependent biodiversity in Nevada and will assist ongoing efforts to protect and conserve these imperiled ecosystems.

Taxonomic revision of the Pyrgulopsis gilae (Caenogastropoda, Hydrobiidae) species complex, with descriptions of two new species from the Gila River basin, New Mexico.

We describe two new species of springsnails (genus Pyrgulopsis) for populations from the middle Fork and upper East Fork of the Gila River Basin (New Mexico) that had been previously identified as P. gilae. We also restrict P. gilae to its originally circumscribed geographic range which consists of a short reach of the East Fork Gila River and a single spring along the Gila River (below the East Fork confluence). These three species form genetically distinct lineages that differ from each other by 3.9-6.3% for mtCOI and 3.7-8.7% for mtNDI (the latter data were newly obtained for this study), and are diagnosable by shell and penial characters. Collectively the three species form a strongly supported clade that is distinguished from other congeners by the unique presence of two glandular strips on the dorsal surface of the penial filament. These findings suggest that the conservation status of P. gilae, which was recently removed from the list of candidates for listing as endangered or threatened by the United States Fish and Wildlife Service, should be revisited and that the two new species may also merit protective measures given their narrow geographic ranges.

Systematics of a widely distributed western North American springsnail, Pyrgulopsis micrococcus (Caenogastropoda, Hydrobiidae), with descriptions of three new congeners.

We describe three new species of springsnails (genus Pyrgulopsis) from the Amargosa River basin, California and Nevada (P. licina sp. n., P. perforata sp. n., P. sanchezi sp. n.), each of which was previously considered to be part of P. micrococcus. We also restrict P. micrococcus to its type locality area (Oasis Valley) and redefine a regional congener, P. turbatrix, to include populations from the central Death Valley region and San Bernardino Mountains that had been previously identified as P. micrococcus. The five species treated herein form genetically distinct lineages that differ from each other by 4.2-12.6% for mtCOI and 5.2-13.6% for mtNDI (based on previously published and newly obtained data), and are diagnosable by shell and/or penial characters. The new molecular data presented herein confirm sympatry of P. licina and P. sanchezi in Ash Meadows (consistent with morphological evidence) and delineate an additional lineage of P. micrococcus (in the broad sense) that we do not treat taxonomically owing to the paucity of morphological material. Conservation measures are needed to ensure the long term persistence of populations of P. micrococcus and a genetically differentiated lineage of P. sanchezi which live in disturbed habitats on private lands.

Pushing short DNA fragments to the limit: Phylogenetic relationships of 'hydrobioid' gastropods (Caenogastropoda: Rissooidea).

Although phylogenetic studies are increasingly utilizing multi-locus datasets, a review of GenBank data for the Gastropoda indicates a strong bias towards a few short gene fragments (most commonly COI, LSU rRNA, and SSU rRNA). This is particularly the case for the Rissooidea, one of the largest and most taxonomically difficult gastropod superfamilies. Here we analyze fragments of these three genes from 90 species to determine whether they can well resolve higher relationships within this superfamily, whether structurally aligned sequence datasets increase phylogenetic signal, and whether the inclusion of highly variable regions introduces noise. We also used the resulting phylogenetic data in combination with morphological/anatomical evidence to re-evaluate the taxonomic status of 'hydrobioid' family-level groups. Our results indicate that all three of the alignment strategies that were used resulted in phylogenies having similar signal levels. However, there was a slight advantage to using structural alignment for inferring family-level relationships. Moreover, the set of 'standard' gastropod genes supported recognition of many previously recognized families and provides new insight into the systematics of several problematic groups. However, some family-group taxa were unresolved and the relationships among families were also poorly supported, suggesting a need for more extensive sampling and inclusion of additional genes.

The endemic gastropod fauna of Lake Titicaca: correlation between molecular evolution and hydrographic history.

Lake Titicaca, situated in the Altiplano high plateau, is the only ancient lake in South America. This 2- to 3-My-old (where My is million years) water body has had a complex history that included at least five major hydrological phases during the Pleistocene. It is generally assumed that these physical events helped shape the evolutionary history of the lake's biota. Herein, we study an endemic species assemblage in Lake Titicaca, composed of members of the microgastropod genus Heleobia, to determine whether the lake has functioned as a reservoir of relic species or the site of local diversification, to evaluate congruence of the regional paleohydrology and the evolutionary history of this assemblage, and to assess whether the geographic distributions of endemic lineages are hierarchical. Our phylogenetic analyses indicate that the Titicaca/Altiplano Heleobia fauna (together with few extralimital taxa) forms a species flock. A molecular clock analysis suggests that the most recent common ancestor (MRCAs) of the Altiplano taxa evolved 0.53 (0.28-0.80) My ago and the MRCAs of the Altiplano taxa and their extralimital sister group 0.92 (0.46-1.52) My ago. The endemic species of Lake Titicaca are younger than the lake itself, implying primarily intralacustrine speciation. Moreover, the timing of evolutionary branching events and the ages of two precursors of Lake Titicaca, lakes Cabana and Ballivián, is congruent. Although Lake Titicaca appears to have been the principal site of speciation for the regional Heleobia fauna, the contemporary spatial patterns of endemism have been masked by immigration and/or emigration events of local riverine taxa, which we attribute to the unstable hydrographic history of the Altiplano. Thus, a hierarchical distribution of endemism is not evident, but instead there is a single genetic break between two regional clades. We also discuss our findings in relation to studies of other regional biota and suggest that salinity tolerance was the most likely limiting factor in the evolution of Altiplano species flocks.

Redescription of Marstonia comalensis (Pilsbry & Ferriss, 1906), a poorly known and possibly threatened freshwater gastropod from the Edwards Plateau region (Texas).

Marstonia comalensis, a poorly known nymphophiline gastropod (originally described from Comal Creek, Texas) that has often been confused with Cincinnatia integra, is re-described and the generic placement of this species, which was recently allocated to Marstonia based on unpublished evidence, is confirmed by anatomical study. Marstonia comalensis is a large congener having an ovate-conic, openly umbilicate shell and penis having a short filament and oblique, squarish lobe bearing a narrow gland along its distal edge. It is well differentiated morphologically from congeners having similar shells and penes and is also genetically divergent relative to those congeners that have been sequenced (mtCOI divergence 3.0-8.5%). A Bayesian analysis of a small COI dataset resolved Marstonia comalensis in a poorly supported sub-clade together with Marstonia hershleri, Marstonia lustrica and Marstonia pachyta. The predominantly new records presented herein indicate that Marstonia comalensis was historically distributed in the upper portions of the Brazos, Colorado, Guadalupe and Nueces River basins, south-central Texas. The species has been live collected at only 12 localities and only two of these have been re-visited since 1993. These data suggest that the conservation status of this snail, which has a critically imperiled (G1) NatureServe ranking and was recently proposed for federal listing, needs to be re-assessed.

Molecular systematics and radiation of western North American nymphophiline gastropods.

Three mitochondrial DNA (mtDNA) markers were used to infer the phylogenetic relationships of the morphologically diverse, species rich, and poorly understood western North American aquatic gastropod genus Pyrgulopsis (Hydrobiidae: Nymphophilinae). Sequences were obtained from 62 of 124 currently recognized species of Pyrgulopsis and representatives of four related genera of North American nymphophilines. Separate and combined analyses of the mtDNA datasets recovered a well supported clade composed of Pyrgulopsis and two other North American nymphophiline genera (Floridobia, Nymphophilus) consistent with the results of a prior study based on a single gene and with anatomical evidence suggesting that these taxa form a monophyletic group. Phylogenetic relationships among lineages of Pyrgulopsis were little resolved in our analyses and provided no obvious basis for splitting this large genus into multiple genera. The little differentiated Mexican genus Nymphophilus was consistently placed within Pyrgulopsis in our trees and is formally synonymized with it herein. Pyrgulopsis was also depicted as paraphyletic with respect to Floridobia in some of our trees while in others the latter was sister to a Nymphophilus + Pyrgulopsis clade. Based on these equivocal results and the morphological and geographical divergence of eastern North American Floridobia relative to Pyrgulopsis, we recommend that the former be maintained as a separate genus. The short, weakly supported branches within Pyrgulopsis and the noncongruence between our molecular phylogenetic hypotheses and geographical groupings of species are attributed to an early rapid diversification of the genus, perhaps triggered by the complex changes in western topography which occurred during the late Tertiary. Our results also indicate that penial morphologies used to define species groups of Pyrgulopsis have been subject to striking convergence throughout the West, suggesting another compelling facet of the radiation of these snails.

A molecular phylogeny of aquatic gastropods provides a new perspective on biogeographic history of the Snake River Region.

Mitochondrial DNA sequences of aquatic gastropods of the subgenus Pyrgulopsis (Natricola) were analyzed to test a commonly accepted hypothesis concerning the early history of the Snake River in the northwestern US. Distributions of Natricola and other regional biota were previously used to infer that the Snake River flowed to the Pacific through southeastern Oregon and northern California during the Neogene prior to its capture by the Columbia River in the late Pliocene (2 Ma). A molecular phylogeny based on partial sequences of COI and NDI (1149 bp) indicates that the Natricola clade is restricted to the modern Snake-Columbia River Basin and the Oregon Lakes region whereas northern California populations previously assigned to this subgenus belong to other lineages. The Natricola clade is not deeply subdivided into Oregon Lakes and Snake River Basin units consistent with late Pliocene fragmentation of the hypothesized paleodrainage, but instead is shallowly structured and contains multiple transitions among these two geographic areas. The strongly supported sister relationship between Natricola and a species from northwest Nevada (P. imperialis) is consistent with a recent proposal that the ancestral Snake River did not flow through southeast Oregon but instead flowed south to the Humboldt River. Within the context of this hypothesis, the multiple transitions between the Snake River Basin and the Oregon Lakes region that occurred within Natricola may be attributed to a late Pleistocene connection between these areas that was unrelated to the early course of the Snake River.

Mitochondrial DNA sequences reveal extensive cryptic diversity within a western American springsnail.

We analysed cytochrome c oxidase subunit I and NADH dehydrogenase subunit I sequence variation among 29 populations of a widely ranging southwestern springsnail (Pyrgulopsis micrococcus) and 18 regional congeners. Cladistic analyses of these sequences depict P. micrococcus as a polyphyletic composite of five well-supported clades. Sequence divergences among these clades and subclades imply the possible occurrence of as many as seven or eight cryptic species in addition to P. micrococcus. Our finding that P. micrococcus contains multiple, genetically distinct and geographically restricted lineages suggests that diversification within this highly speciose aquatic genus has been structured in large part by the operation of terrestrial barriers to gene flow. However, these sequence data also indicate that recent dispersal among hydrographically separated areas has occurred within one of these lineages, which we attribute to passive transport on migratory waterbirds.