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.     

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.     

Population Structure in the Roundtail Chub (Gila robusta Complex) of the Gila River Basin as Determined by Microsatellites: Evolutionary and Conservation Implications

Ten microsatellite loci were characterized for 34 locations from roundtail chub (Gila robusta complex) to better resolve patterns of genetic variation among local populations in the lower Colorado River basin. This group has had a complex taxonomic history and previous molecular analyses failed to identify species diagnostic molecular markers. Our results supported previous molecular studies based on allozymes and DNA sequences, which found that most genetic variance was explained by differences among local populations. Samples from most localities were so divergent species-level diagnostic markers were not found. Some geographic samples were discordant with current taxonomy due to admixture or misidentification; therefore, additional morphological studies are necessary. Differences in spatial genetic structure were consistent with differences in connectivity of stream habitats, with the typically mainstem species, G. robusta, exhibiting greater genetic connectedness within the Gila River drainage. No species exhibited strong isolation by distance over the entire stream network, but the two species typically found in headwaters, G. nigra and G. intermedia, exhibited greater than expected genetic similarity between geographically proximate populations, and usually clustered with individuals from the same geographic location and/or sub-basin. These results highlight the significance of microevolutionary processes and importance of maintaining local populations to maximize evolutionary potential for this complex. Augmentation stocking as a conservation management strategy should only occur under extreme circumstances, and potential source populations should be geographically proximate stocks of the same species, especially for the headwater forms.     

Genetic and morphological characterization of the freshwater mussel clubshell species complex (Pleurobema clava and Pleurobema oviforme) to inform conservation planning

The shell morphologies of the freshwater mussel species Pleurobema clava (federally endangered) and Pleurobema oviforme (species of concern) are similar, causing considerable taxonomic confusion between the two species over the last 100 years. While P. clava was historically widespread throughout the Ohio River basin and tributaries to the lower Laurentian Great Lakes, P. oviforme was confined to the Tennessee and the upper Cumberland River basins. We used two mitochondrial DNA (mtDNA) genes, 13 novel nuclear DNA microsatellite markers, and shell morphometrics to help resolve this taxonomic confusion. Evidence for a single species was apparent in phylogenetic analyses of each mtDNA gene, revealing monophyletic relationships with minimal differentiation and shared haplotypes. Analyses of microsatellites showed significant genetic structuring, with four main genetic clusters detected, respectively, in the upper Ohio River basin, the lower Ohio River and Great Lakes, and upper Tennessee River basin, and a fourth genetic cluster, which included geographically intermediate populations in the Ohio and Tennessee river basins. While principal components analysis (PCA) of morphometric variables (i.e., length, height, width, and weight) showed significant differences in shell shape, only 3% of the variance in shell shape was explained by nominal species. Using Linear Discriminant and Random Forest (RF) analyses, correct classification rates for the two species'' shell forms were 65.5% and 83.2%, respectively. Random Forest classification rates for some populations were higher; for example, for North Fork Holston (HOLS), it was >90%. While nuclear DNA and shell morphology indicate that the HOLS population is strongly differentiated, perhaps indicative of cryptic biodiversity, we consider the presence of a single widespread species the most likely biological scenario for many of the investigated populations based on our mtDNA dataset. However, additional sampling of P. oviforme populations at nuclear loci is needed to corroborate this finding.     

Two new species of Yushania (Poaceae: Bambusoideae) from South China,with a taxonomic revision of related species

Two new species of Yushania (Poaceae, Bambusoideae, Arundinarieae) are described and illustrated from Hunan, China. Yushania longshanensis D.Z. Li & X.Y. Ye is distinguished from related species (Y. confusa, Y. angustifolia and Y. pachyclada) by having a thinner culm (0.2–0.3 cm in diameter), glabrous sheath scar, no oral setae, a large glabrous leaf blade (10–20 × 0.9–1.3 cm) and 3–4 pairs of secondary veins. Yushania stoloniforma D.Z. Li & X.Y. Ye has a distinctive scrambling habit, which differs from its putative close allies. Both of these two new species have a solitary branch at the basal nodes and can be assigned to Yushania sect. Yushania based on morphological features. Additionally, we treated Yushania gigantea T.P. Yi & L. Yang as a new synonym of Y. elevata T.P. Yi and renamed Y. microphylla T.P. Yi & L. Yang as Y. weiningensis D.Z. Li & X.Y. Ye.     

First record of harpacticoid copepods from Lake Tahoe,United States: two new species of Attheyella (Harpacticoida,Canthocamptidae)

Benthic harpacticoids were collected for the first time at Lake Tahoe, California-Nevada, United States. Two species were identified as members of the genus Attheyella Brady, 1880. The genus Attheyella comprises about 150 species within six subgenera, but only twelve species have previously been reported from North American freshwater habitats. The two new species of Attheyella described here have a 3-segmented endopod on P1 and 2-segmented P2–P4 endopods, the distal segment of exopod of P2–P4 has three outer spines, and the P5 has five setae on the exopod and six setae on the baseoendopod. Attheyella (Attheyella) tahoensis sp. n. most closely resembles Attheyella (Attheyella) idahoensis (Marsh, 1903) from Idaho, Montana, and Alaska (United States) and Attheyella (Attheyella) namkungi Kim, Soh & Lee, 2005 from Gosu Cave in South Korea. They differ mainly by the number of setae on the distal endopodal segment of P2–P4. In addition, intraspecific variation has been observed on the caudal rami. Attheyella (Neomrazekiella) tessiae sp. n. is characterized by the extension of P5 baseoendopod, 2-segmented endopod of female P2–P3, and naked third seta of male P5 exopod. The two new species are likely endemic to Lake Tahoe, an isolated alpine lake within the Great Basin watershed in the western United States.     

A new highly specialized cave harvestman from Brazil and the first blind species of the genus: Iandumoema smeagol sp. n. (Arachnida,Opiliones, Gonyleptidae)

A new species of troglobitic harvestman, Iandumoema smeagol sp. n., is described from Toca do Geraldo, Monjolos municipality, Minas Gerais state, Brazil. Iandumoema smeagol sp. n. is distinguished from the other two species of the genus by four exclusive characteristics – dorsal scutum areas with conspicuous tubercles, enlarged retrolateral spiniform tubercle on the distal third of femur IV, eyes absent and the penial ventral process slender and of approximately the same length of the stylus. The species is the most highly modified in the genus and its distribution is restricted only to caves in that particular area of Minas Gerais state. The type locality is not inside a legally protected area, and there are anthropogenic impacts in its surroundings. Therefore, Iandumoema smeagol sp. n. is vulnerable and it must be considered in future conservation projects.     

Sylphella puccoon gen. n., sp. n. and two additional new species of aquatic oligochaetes (Lumbriculidae,Clitellata) from poorly-known lotic habitats in North Carolina (USA)

Three new species of Lumbriculidae were collected from floodplain seeps and small streams in southeastern North America. Some of these habitats are naturally acidic. Sylphella puccoon gen. n., sp. n. has prosoporous male ducts in X–XI, and spermathecae in XII–XIII. Muscular, spherical atrial ampullae and acuminate penial sheaths distinguish this monotypic new genus from other lumbriculid genera having similar arrangements of reproductive organs. Cookidrilus pocosinus sp. n. resembles its two subterranean, Palearctic congeners in the arrangement of reproductive organs, but is easily distinguished by the position of the spermathecal pores in front of the chaetae in X–XIII. Stylodrilus coreyi sp. n. differs from congeners having simple-pointed chaetae and elongate atria primarily by the structure of the male duct and the large clusters of prostate cells. Streams and wetlands of Southeastern USA have a remarkably high diversity of endemic lumbriculids, and these poorly-known invertebrates should be considered in conservation efforts.     

Annotated checklist of the leech species diversity in the Maloe More Strait of Lake Baikal,Russia

In this paper, the very first checklist of the freshwater leeches of Maloe More Strait, a special part of Lake Baikal, is presented. It includes 14 free-living and parasitic species, of which four species belong to endemic Baikal genera – two species from Baicalobdella and one species each from Baicaloclepsis and Codonobdella. The checklist highlights six potentially new morphological species recorded for the first time in the area. The exact systematic position is stated for all leech species. Each species from the list is provided with information on taxonomic synonymy, data on its geographic distribution, and ecological characteristics. New species records are additionally provided with brief morphological characteristics and photos of their external morphology.     

Genetic architecture and genomic patterns of gene flow between hybridizing species of Picea

Hybrid zones provide an opportunity to study the effects of selection and gene flow in natural settings. We employed nuclear microsatellites (single sequence repeat (SSR)) and candidate gene single-nucleotide polymorphism markers (SNPs) to characterize the genetic architecture and patterns of interspecific gene flow in the Picea glauca × P. engelmannii hybrid zone across a broad latitudinal (40–60 degrees) and elevational (350–3500 m) range in western North America. Our results revealed a wide and complex hybrid zone with broad ancestry levels and low interspecific heterozygosity, shaped by asymmetric advanced-generation introgression, and low reproductive barriers between parental species. The clinal variation based on geographic variables, lack of concordance in clines among loci and the width of the hybrid zone points towards the maintenance of species integrity through environmental selection. Congruency between geographic and genomic clines suggests that loci with narrow clines are under strong selection, favoring either one parental species (directional selection) or their hybrids (overdominance) as a result of strong associations with climatic variables such as precipitation as snow and mean annual temperature. Cline movement due to past demographic events (evidenced by allelic richness and heterozygosity shifts from the average cline center) may explain the asymmetry in introgression and predominance of P. engelmannii found in this study. These results provide insights into the genetic architecture and fine-scale patterns of admixture, and identify loci that may be involved in reproductive barriers between the species.     

Two new Megalothorax species of the minimus group (Collembola,Neelidae)

Two new Megalothorax species, Megalothorax potapovi sp. n. from the Russian Far East and Megalothorax sanguineus sp. n. from the French Pyrénées are described. The two new species have a set of morphological characters (including a smooth mucro) that places them among the minimus group sensu Schneider and D’Haese (2013). Megalothorax potapovi characteristics include dorsal protuberance on forehead, peculiar chaetotaxy of antenna III and strong lanceolate chaetae on body. Megalothorax sanguineus characteristics include strong red pigmentation, large network of integumentary channels on head and elongated apex of the two postero-distal spines of dens. The DNA barcodes ( cytochrome oxidase subunit I–COI) of the two species are also provided and analyzed among a broader sampling of the genus in order to support further their specific status. A special focus is given to the labral morphological characteristics. Pseudopores-like elements are reported for the first time in the genus. Positions of the τ-chaetae near the dorsal sensory field of thorax II are compared for several species of the genus.     

Genetic variation of the major histocompatibility complex (MHC class II β gene) in the threatened Gila trout, <Emphasis Type="Italic">Oncorhynchus gilae gilae</Emphasis>

Gila trout (Oncorhynchus gilae gilae) was federally protected in 1973 because of severe declines in abundance and geographic range size. At present, four relict genetic lineages of the species remain in mountain streams of New Mexico and Arizona, USA. Management actions aimed at species recovery, including hatchery production and restocking of formerly occupied streams, have been guided by information from non-functional genetic markers. In this study, we investigated genetic variation at exon 2 of the major histocompatibility complex (MHC) class II β gene that is involved in pathogen resistance and thus presumably under natural selection. Phylogenetic analysis revealed trans-species polymorphism and a significantly high ratio of non-synonymous to synonymous amino acid changes consistent with the action of historical balancing selection that maintained diversity at this locus in the past. However, Gila trout exhibited low allelic diversity (five alleles from 142 individuals assayed) compared to some other salmonid fishes, and populations that originated exclusively from hatcheries possessed three or fewer MHC alleles. Comparative analysis of genetic variation at MHC and six presumably neutrally evolving microsatellite loci revealed that genetic drift cannot be rejected as a primary force governing evolution of MHC in contemporary populations of Gila trout. Maintenance of diversity at MHC will require careful implementation of hatchery breeding protocols and continued protection of wild populations to prevent loss of allelic diversity due to drift.     

Morphometric and meristic diversity of the species Scolopendra cingulata Latreille, 1829 (Chilopoda: Scolopendridae) in the Mediterranean region

Scolopendra cingulata Latreille, 1829 is the commonest scolopendromorph species in the Mediterranean area. However, its morphological and geographic variation has not been examined so far, and therefore robust hypotheses about the factors that have shaped that variability are lacking. We examined, using multivariate methods for 19 morphometric and meristic characters, the morphological variation of that species using a comprehensive sample of 503 adult specimens from 130 restricted geographic localities. The localities were distributed in three major geographic areas (Balkans, Asia Minor-Middle East, and Italy) in order to discern biological entities and to estimate the morphological relationships between populations and geographic regions. Results showed significant differences between the three geographic groups. Characters such as the distance between the paramedian sulci of the 7th and 8th tergite, the number of antennal segments, the number of spines on the dorsal side of 21st prefemur and number of spines on the 21st prefemoral process significantly discriminated populations of S. cingulata along a west-east geographic gradient. Both eastern (Balkan) and easternmost populations (Asia Minor-Middle East) showed higher mean values of antennal articles and spines of 21st prefemur than the Italian populations. No significant morphological variation was discovered between the sexes of S. cingulata apart from cephalic width and distance between the two paramedian sulci of the 7th and 8th tergite. Based on certain morphometric differences among the three major geographic regions, we suggest that S. cingulata originated in the east and colonized south European area from two different geographic routes. It is suggested that west-east morphological trend of S. cingulata is related either to the prominent palaeogeographic events of the area or to the competition with S. oraniensis Lucas, 1846 in the western Mediterranean.     

A new species of Noblella (Amphibia,Anura, Craugastoridae) from the humid montane forests of Cusco,Peru

A new species of Noblella is described from the humid montane forest of the Región Cusco in Peru. Specimens were collected at 2330–2370 m elevation in Madre Selva, near Santa Ana, in the province of La Convención. The new species is readily distinguished from all other species of Noblella by having a broad, irregularly shaped, white mark on black background on chest and belly. The new species further differs from known Peruvian species of Noblella by the combination of the following characters: tympanic membrane absent, small tubercles on the upper eyelid and on dorsum, tarsal tubercles or folds absent, tips of digits not expanded, no circumferential grooves on digits, dark brown facial mask and lateral band extending from the tip of the snout to the inguinal region. The new species has a snout-to-vent length of 15.6 mm in one adult male and 17.6 mm in one adult female. Like other recently described species in the genus, this new Noblella inhabits high-elevation forests in the Andes and likely has a restricted geographic distribution.     

A Multi-Year Longitudinal Study of Water Quality Parameters in Four Salmon-Bearing and Recreational Streams on Mount Hood,Oregon

Four streams–Clear Fork, Lost Creek, Camp Creek and Still Creek–in northwestern Oregon’s Sandy River Basin were monitored for temperature, dissolved oxygen levels, and fecal bacterial concentrations in a multi-year analysis examining stream health for recreational users and anchor habitat for Pacific Salmon. Temperatures were recorded using micro –T temperature loggers at 15 locations, during 22 July - 5 September 2006, 2 July - 4 September 2007, 20 June - 7 September 2008, 23 June - 9 September 2009, and 2 July –9 September 2010. The Seven-Day Average Maximum water temperature (7-DAM) of 13°C was used as a reference value for the biological limit governing suitable salmonid spawning and egg incubation conditions. The maximum 7-DAM temperatures occurred on different dates and all streams neared or exceeded the 13°C standard at least once each summer. Dissolved oxygen levels were measured at weekly or longer intervals in 2006, 2007, 2008, and 2009. Dissolved oxygen levels fell below the 9.0 ppm standard for Clear Fork on almost half the sampling dates in 2006, 2007, and 2009. Concentrations of the bacterial genus Enterococcus were measured as an indicator of fecal contamination. Samples were collected at 15 sites along the four streams. Weekly samples were collected during a 9 week period from July - September 2007, an 11 week period from June - September 2008, and an 11 week period from June - September 2009. Enterococcus counts exceeded the federal recommended national criterion value of 61 colony forming units (CFU) per 100 mL every year in Camp Creek and occasionally elsewhere, with exceedances trending towards late summer.     

Importance of demographic history for phylogeographic inference on the arctic–alpine plant Phyllodoce caerulea in East Asia

Arctic–alpine plants have enormous ranges in the Northern Hemisphere. Phylogeographic studies have provided insights into their glacial survival as well as their postglacial colonization history. However, our understanding of the population dynamics of disjunct alpine populations in temperate regions remains limited. During Pleistocene cold periods, alpine populations of arctic–alpine species in East Asia were either connected to an ice-free Beringia refugium or they persisted with prolonged isolation after their establishment. To estimate which of these scenarios is more likely, we elucidated the genetic structure of Phyllodoce caerulea (Ericaceae) in Beringia and northern Japan, East Asia. Sequence variation in multiple nuclear loci revealed that P. caerulea can be distinguished into northern and southern groups. A demographic analysis demonstrated that the north–south divergence did not predate the last glacial period and detected introgression from Phyllodoce aleutica, relative widely distributed in East Asia, exclusively into the southern group. Therefore, although there has been genetic divergence between northern Japan and Beringia in P. caerulea, the divergence is unlikely to have resulted from their prolonged geographic separation throughout several cycles of glacial and interglacial periods. Instead, our study suggests that the introgression contributed to the genetic divergence of P. caerulea and that the range of P. caerulea was plausibly connected between northern Japan and Beringia during the last glacial period. Overall, our study not only provides a biogeographic insight into alpine populations of arctic–alpine plants in East Asia but also emphasizes the importance of careful interpretation of genetic structure for inferring phylogeographic history.     

Vicariance and Its Impact on the Molecular Ecology of a Chinese Ranid Frog Species-Complex (Odorrana schmackeri,Ranidae)

Paleogeological events and Pleistocene climatic fluctuations have had profound influences on the genetic patterns and phylogeographic structure of species in southern China. In this study, we investigated the population genetic structure and Phylogeography of the Odorrana schmackeri species complex, mountain stream-dwelling odorous frogs, endemic to southern China. We obtained mitochondrial sequences (1,151bp) of the complete ND2 gene and two flanking tRNAs of 511 individuals from 25 sites for phylogeographic analyses. Phylogenetic reconstruction revealed seven divergent evolutionary lineages, with mean pairwise (K2P) sequence distances from 7.8% to 21.1%, except for a closer ND2 distance (3.4%). The complex geological history of southern China drove matrilineal divergence in the O. schmackeri species complex into highly structured geographical units. The first divergence between lineage A+B and other lineages (C-G) had likely been influenced by the uplift of coastal mountains of Southeast China during the Mio-Pliocene period. The subsequent divergences between the lineages C-G may have followed the formation of the Three Gorges and the intensification of the East Asian summer monsoon during the late Pliocene and early Pleistocene. Demographic analyses indicated that major lineages A and C have been experienced recent population expansion (c. 0.045–0.245 Ma) from multiple refugia prior to the Last Glacial Maximum (LGM). Molecular analysis suggest that these seven lineages may represent seven different species, three described species and four cryptic species and should at least be separated into seven management units corresponding to these seven geographic lineages for conservation.     

Mesozoic origin and ‘out-of-India’ radiation of ricefishes (Adrianichthyidae)

The Indian subcontinent has an origin geologically different from Eurasia, but many terrestrial animal and plant species on it have congeneric or sister species in other parts of Asia, especially in the Southeast. This faunal and floral similarity between India and Southeast Asia is explained by either of the two biogeographic scenarios, ‘into-India’ or ‘out-of-India’. Phylogenies based on complete mitochondrial genomes and five nuclear genes were undertaken for ricefishes (Adrianichthyidae) to examine which of these two biogeographic scenarios fits better. We found that Oryzias setnai, the only adrianichthyid distributed in and endemic to the Western Ghats, a mountain range running parallel to the western coast of the Indian subcontinent, is sister to all other adrianichthyids from eastern India and Southeast–East Asia. Divergence time estimates and ancestral area reconstructions reveal that this western Indian species diverged in the late Mesozoic during the northward drift of the Indian subcontinent. These findings indicate that adrianichthyids dispersed eastward ‘out-of-India’ after the collision of the Indian subcontinent with Eurasia, and subsequently diversified in Southeast–East Asia. A review of geographic distributions of ‘out-of-India’ taxa reveals that they may have largely fuelled or modified the biodiversity of Eurasia.     

Microsatellite loci in the Lahontan tui chub,Gila bicolor obesa,and their utilization in other chub species

Primers were developed for 17 microsatellite loci from an enriched (GATA)_n library from DNA from Gila bicolor obesa. These primers were tested for cross‐species amplification in Gila bicolor snyderi, Gila orcutti and Gila coerulea.