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.     

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.     

Genetic and morphologic variation of the Pecos assiminea,an endangered mollusk of the Rio Grande region,United States and Mexico (Caenogastropoda: Rissooidea: Assimineidae)

Assiminea pecos is an endangered species of amphibious gastropod that occupies four widely separated portions of the Rio Grande region in the southwestern United States (Pecos River basin) and northeastern Mexico (Cuatro Cienegas basin). Our statistical and discriminant function analyses of shell variation among the disjunct populations of this species indicate that Mexican specimens differ in their morphometry from those of the United States and can be diagnosed by several characters. We also analyzed variation in the mitochondrial genome by sequencing 658 bp of mitochondrial COI from populations of A. pecos, representatives of the other three North American species of Assiminea, and several outgroups. Our results indicated substantial divergence of the Mexican population of A. pecos, which was consistently depicted as a monophyletic unit nested within or sister to the shallowly structured group comprised of American members of this species. Consistent with our findings, we describe the Mexican population as a new species, which is provisionally placed in the large, worldwide genus Assiminea pending further study of the phylogentic relationships of the North American assimineids. Our molecular data suggest that the Rio Grande region assimineids, which are among the few inland members of the otherwise estuarine subfamily Assimineinae, diverged from coastal progenitors in the late Miocene, with subsequent Pleistocene vicariance of Mexican and American species perhaps associated with development of the modern, lower course of the Rio Grande. Handling editor: K. Martens     

Phylogenetic relationships of assimineid gastropods of the Death Valley–lower Colorado River region: relicts of a late Neogene marine incursion?

Aim A small fauna of amphibious snails (genus Assiminea Fleming, 1828) living in association with highly mineralized springs in the Death Valley–lower Colorado River region (DVLCR) is thought to be a relict of the Bouse Embayment, a putative late Miocene–early Pliocene transgression of the ancestral Gulf of California along the lower Colorado River valley. We analysed the phylogenetic relationships of this fauna using mtDNA sequence data (1171 bp) to determine whether, as would be consistent with this hypothesis, it forms a substantially divergent unit sister to marine coastal congeners. Location South‐western Great Basin and lower Colorado River region, USA. Methods Two genes [mitochondrial cytochrome c oxidase subunit I (COI) and the mitochondrial 16S ribosomal RNA gene] were sequenced for 10 populations of DVLCR assimineas (Assiminea infima Berry, 1947 ; Assiminea sp.). We also sequenced an undescribed population from a spring in the Colorado River delta; western North American Pacific Coastal Assiminea californica (Tryon, 1865); the three other congeners that live on the continent; and three Old World assimineids (outgroups). Phylogenies based on the combined data set were obtained using Bayesian methods, and divergence times were estimated using a COI molecular clock for related gastropods. Results Composite haplotypes of the DVLCR assimineas, together with that observed in the Colorado River delta population, formed a weakly supported clade that was sister to a clade composed of populations of North American Pacific and Atlantic coastal species. The genetic distance between members of these two clades was 3.46 ± 0.47% for COI and 1.69 ± 0.38% for 16S. The former clade was composed of five subunits that differed from each other by 1.29–2.84% (COI) and 0.52–1.98% (16S) sequence divergence. Main conclusions Application of the COI clock suggests that progenitors of the DVLCR fauna diverged from coastal ancestors 2.13–1.89 Ma (late Pliocene), several million years after the Bouse Embayment would have been terminated by the establishment of the lower (freshwater) Colorado River. This finding, together with shallow genetic structuring of several DVLCR lineages that are widely distributed across the topographically complex regional landscape, suggests that the Assiminea fauna of this inland area was more likely to have been founded by coastal colonists transported on water birds than through a direct connection with the sea.     

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.     

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.     

Genetic variation in the Desert Springsnail (Tryonia porrecta): implications for reproductive mode and dispersal

Allozymes and mitochondrial cytochrome c oxidase subunit I (mtCOI) sequences were analysed to determine whether populations of the western North American gastropod Tryonia porrecta (from California, Nevada, Utah, and northwest Mexico) are strongly differentiated in accordance with traditional interpretation of regional fauna as ancient relicts inhabiting isolated fragments of late Tertiary palaeodrainages. These data were also used to assess whether this species, for which males have not been recorded, is a rare example of a molluscan parthenogen. Both data sets strongly supported monophyly of T. porrecta populations. Five of the nine sampled populations consisted of a single monoallelic allozyme genotype while the others contained two to 10 distinct genotypes. Allozymic data for genetically diverse Utah populations provided evidence of clonal and sexual reproduction. mtCOI haplotypes of T. porrecta formed two subgroups which differed by 1.99-2.60%. The common haplotype was found in seven populations with rare haplotypes observed in single populations. Based on these results and an available mtCOI molecular clock for related hydrobiid snails, T. porrecta is interpreted as a primarily parthenogenetic species that undergoes occasional sexual reproduction and has accumulated substantial diversity following its mid-Pliocene to mid-Pleistocene origin. Our results also suggest that the distribution of present-day populations of these gill-breathing snails did not result from fragmentation of an ancient, well-integrated drainage but instead reflects overland colonization of habitats which only recently became available following desiccation of late Quaternary pluvial lakes.     

Molecular evidence for cryptic species in a narrowly endemic western North American springsnail (Pyrgulopsis gilae)

Pyrgulopsis gilae is a small springsnail that is narrowly distributed along the forks of the upper Gila River and currently being managed as a threatened and sensitive species by the State of New Mexico and United States Forest Service, respectively. A previous phylogeographic study of this species based on mitochondrial COI sequences delineated substantial divergence between several populations along the lower and upper reaches of the East Fork Gila River. The present study surveyed COI variation among a larger number of populations across the entire geographic range of P. gilae. Three haplotype groupings were delineated that were congruently resolved as clades by a Bayesian analysis. One of the clades was composed of populations along the lower East Fork and mainstem Gila River and corresponds to P. gilae as originally circumscribed. The other two clades were composed of populations along the Middle Fork and upper East Fork Gila River that were recently referred to P. gilae. These three geographically isolated clades do not share any haplotypes, have significant F_ST values, and are differentiated from each other by 3.9–6.3 % sequence divergence. Based on this evidence we suggest that the clades represent distinct species and should be managed as separate conservation units pending taxonomic revision of P. gilae. This study provides additional evidence that geographically disjunct subunits of Pyrgulopsis species often represent distinct monophyletic lineages that may warrant formal taxonomic recognition, and thus underscores the importance of fine-scale conservation genetics studies of these imperiled organisms.     

Discovery of introduced and cryptogenic cochliopid gastropods in the San Francisco Estuary, California

We report the first discovery in the San Francisco Estuary (‘Estuary’)of two cochliopid gastropods, Littoridinops monroensis and Tryoniaporrecta. These identifications were based on morphologicalcriteria and supported by analysis of mitochondrial DNA (cytochromec oxidase subunit I, COI) sequence data. We also report thefirst discovery of males in parthenogenetic T. porrecta. Thenew records represent large range extensions for both of theseNorth American species, which were previously recorded frompredominantly brackish habitats along the western Atlantic-GulfCoast (L. monroensis) and thermal springs in the Great Basinand lower Colorado River region (T. porrecta). The COI haplotypeobserved in Estuary specimens of L. monroensis differed fromthose detected in two western Atlantic populations by only 1–3 bp,suggesting recent divergence which is not consistent with theseparation of these two areas by imposing terrestrial barrierssince at least the Pliocene. We suggest that L. monroensis wasrecently introduced to the Estuary by transoceanic shipping,adding to the large exotic biota that has invaded this highlydisturbed ecosystem. The COI haplotype observed in Estuary specimensof T. porrecta is closely similar to haplotypes detected inNevada and Utah populations and highly divergent relative tothe single haplotype observed in other California populations.The implications of these findings for the status of T. porrectain the Estuary are unclear, because the native range of thisparthenogen has not been established and its scattered distributionin the West may be attributable to natural dispersal acrossland (on birds) and/or anthropogenic spread. Although we suggesttreating T. porrecta as cryptogenic in the Estuary, a nativestatus may be suggested by independent (subfossil) evidencethat this snail was locally present prior to establishment ofthe area as a major centre of human population and commercein the 1850s. (Received 13 April 2007; accepted 25 June 2007)     

Origin and diversification of the soldier meadow springsnails (Hydrobiidae: Pyrgulopsis), a species flock in the northwestern Great Basin, United States

The large, western North American hydrobiid gastropod genusPyrgulopsis (commonly known as springsnails) includes a seriesof locally endemic faunas that are thought to be species flocks.Although these assemblages are of interest from the evolutionary,biogeographic and conservation perspectives, their monophylyand phylogenetic relationships have yet to be rigorously evaluated.Here we present a molecular phylogeny (based on mitochondrialsequence data) of a putative flock of four thermal spring-dwellingspringsnails that is distributed in many sites in Soldier Meadowand a single locality in Bog Hot Valley (northwestern Nevada).Our analyses support monophyly of this assemblage (‘SoldierMeadow clade’) and a close relationship with other regionalspecies and suggest that the invasion of thermal habitats bythese springsnails occurred independently of other such radiationswithin the genus. The divergence of the Soldier Meadow claderelative to its sister group is substantial (6.79–10.36%for COI, 10.35–15.88% for NDI), suggesting a split inthe early Pliocene, based on the application of a COI clockfor Pyrgulopsis. The splits within the Soldier Meadow cladeinto three main subunits also appear to be old events, basedon their 5.78–8.54% COI divergence relative to each other.These findings are consistent with a long history of springsnailevolution in Soldier Meadow, which is intriguing given thatthis basin was flooded by Lake Lahontan during periods of theearly and middle Pleistocene. We suggest that progenitors ofthe contemporary fauna survived in high elevation springs thatmay have been present in the basin during these pluvial periodsand subsequently colonized contemporary habitats following thetermination of the extreme Lake Lahontan highstands. We speculatethat the broadly disjunct population (of P. militaris) in BogHot Valley, which is consistently nested within the SoldierMeadow clade in our phylogenetic analyses, is either a vicariantrelict of a spring zone that may have once extended betweenthese two areas; or was founded by a past ‘jump’dispersal event from Soldier Meadow. Phylogeographic structureof springsnail populations in Soldier Meadow bears the strongstamp of geologically recent, allopatric diversification, perhapsreflecting the short time that basin floor habitats have beenoccupied. We describe a new species (P. varneri) for a seriesof recently discovered populations that are monophyletic, substantiallydivergent and morphologically distinctive. Additional studieswill be necessary to confidently assess the taxonomic statusof morphologically distinctive P. limaria and P. umbilicata,which are shown herein to be little divergent genetically; anda recently discovered minute springsnail that is morphologicallydivergent yet closely similar genetically to P. notidicola. (Received 20 December 2006; accepted 22 February 2007)