High-throughput environmental sequencing reveals high diversity of litter and moss associated protist communities along a gradient of drainage and tree productivity.

Although previous studies, mostly based on microscopy analyses of a few groups of protists, have suggested that protists are abundant and diverse in litter and moss habitats, the overall diversity of moss and litter associated protists remains elusive. Here, high-throughput environmental sequencing was used to characterize the diversity and community structure of litter- and moss-associated protists along a gradient of soil drainage and forest primary productivity in a temperate rainforest in British Columbia. We identified 3262 distinct protist OTUs from 36 sites. Protists were strongly structured along the landscape gradient, with a significant increase in alpha diversity from the blanket bog ecosystem to the zonal forest ecosystem. Among all investigated environmental variables, calcium content was the most strongly associated with the community composition of protists, but substrate composition, plant cover and other edaphic factors were also significantly correlated with these communities. Furthermore, a detailed phylogenetic analysis of unicellular opisthokonts identified OTUs covering most lineages, including novel OTUs branching with Discicristoidea, the sister group of Fungi, and with Filasterea, one of the closest unicellular relatives to animals. Altogether, this study provides unprecedented insight into the community composition of moss- and litter-associated protists.

Molecular phylogeny of a new gonostomatid ciliate revealing a discrepancy between interphasic and cell divisional patterns (Ciliophora, Hypotricha).

The morphology, ontogenesis, and molecular phylogeny of a new ciliate, Gonostomum koreanum n. sp., discovered in a terrestrial moss sample from South Korea, were investigated. Morphologically, it is characterized by a gonostomatid oral apparatus, two macronuclear nodules, six frontoventral rows, the two rightmost of which (frontoventral rows V and VI) extend posteriorly to near pretransverse and transverse cirri, and three dorsal kineties each with a single caudal cirrus posteriorly. The new species is easily confused with members of the genus Metagonostomum because of the long frontoterminal cirral row but differs mainly in the de novo (vs. intrakinetal) origin of anlage VI, a character found only in Gonostomum and Paragonostomum. To solve the discrepancy between the interphasic and ontogenetic patterns, we additionally performed morphological and multigene analyses on three gonostomatid species, namely Gonostomum koreanum n. sp. and its morphologically (M. gonostomoida) and ontogenetically (G. kuehnelti) most similar species. The multigene analyses show that the new species is closely related to G. kuehnelti and the core gonostomatids consists of five groups based on the origin of the frontoventral rows.

Does microhabitat influence the testate amoebae communities in the Nameri National Park, northeastern India? A tropical forest perspective.

The community structure of testate amoebae inhabiting different microhabitats (soil and tree-moss) within a tropical forest biome in Nameri National Park, northeastern India, was investigated. A total of 33 testate amoebae species belonging to 13 genera were identified. Species belonging to the class Lobosea constituted 73% of total testate amoebae density in the soil habitat, whereas the class Filosea constituted the most dominant forms (58%) in the moist tree-moss habitat. The relative abundance of species was higher in the tree-moss habitat compared to the soil habitats of the forest. Although multivariate analysis suggested a significant difference in assemblage patterns between the habitats, the turnover in species (i.e., beta diversity) was insignificant. Species accumulation curves (SAC) constructed using both parametric and non-parametric species richness estimators revealed that the asymptote of species richness was achieved by a low number of sample replicates in both habitats. The temperature and pH of the substratum on testate amoebae distribution patterns suggest the importance of additional background factors on testate amoebae community structure. Further studies involving more biotopes, seasons, and trophic interactions are recommended to document a complete record of testate amoebae diversity and their interactions with environmental gradients in the tropical forest biomes of northeastern India.

Ribosomal RNA genes challenge the monophyly of the Hyalospheniidae (Amoebozoa: Arcellinida).

To date only five partial and two complete SSU rRNA gene sequences are available for the lobose testate amoebae (Arcellinida). Consequently, the phylogenetic relationships among taxa and the definition of species are still largely dependant on morphological characters of uncertain value, which causes confusion in the phylogeny, taxonomy and the debate on cosmopolitanism of free-living protists. Here we present a SSU rRNA-based phylogeny of the Hyalospheniidae including the most common species. Similar to the filose testate amoebae of the order Euglyphida the most basal clades have a terminal aperture; the ventral position of the pseudostome appears to be a derived character. Family Hyalospheniidae appears paraphyletic and is separated into three clades: (1) Heleopera sphagni, (2) Heleopera rosea and Argynnia dentistoma and (3) the rest of the species from genera Apodera, Hyalosphenia, Porosia and Nebela. Our data support the validity of morphological characters used to define species among the Hyalospheniidae and even suggest that taxa described as varieties may deserve the rank of species (e.g. N. penardiana var. minor). Finally our results suggest that the genera Hyalosphenia and Nebela are paraphyletic, and that Porosia bigibbosa branches inside the main Nebela clade.

Contrasting diversity of testate amoebae communities in Sphagnum and brown-moss dominated patches in relation to shell counts.

Ecological studies of peatland testate amoebae are generally based on totals of 150 individuals per sample. However, the suitability of this standard has never been assessed for alkaline habitats such as spring fens. We explored the differences in testate amoeba diversity between Sphagnum and brown-moss microhabitats at a mire site with a highly diversified moss layer which reflects the small-scale heterogeneity in groundwater chemistry. Relationships between sampling efficiency and sample completeness were explored using individual-based species accumulation curves and the effort required to gain an extra species was assessed. Testate amoeba diversity differed substantially between microhabitats, with brown mosses hosting on average twice as many species and requiring greater shell totals to reach comparable sample analysis efficiency as for Sphagnum. Thus, for samples from alkaline conditions an increase in shell totals would be required and even an overall doubling up to 300 individuals might be considered for reliable community description. Our small-scale data are likely not robust enough to provide an ultimate solution for the optimization of shell totals. However, the results proved that testate amoebae communities from acidic and alkaline environments differ sharply in both species richness and composition and they might call for different methodological approaches.

Spathidium seppelti foissneri nov. subspec., Spathidium simplinucleatum nov. stat., and Dileptus americanus Kahl, 1931, one new and two poorly known soil gymnostome ciliates from soils of Slovakia.

The morphology and infraciliature of three soil gymnostome ciliates, Spathidium seppelti foissneri nov. subspec., Spathidium simplinucleatum nov. stat., and Dileptus americanus, collected from Slovakia, were investigated using live observation and protargol impregnation. S. seppelti foissneri is distinguished from its congeners by a combination of characters including: distinctly spatulate shape with convex oral bulge; 25 ellipsoidal, scattered macronuclear nodules; an average of 23 ciliary rows, 3 anteriorly differentiated to form a dorsal brush occupying 25% of the body length; and rod-shaped to slightly ellipsoidal extrusomes measuring about 5 x 0.5 microm. S. simplinucleatum is about 110 x 50 microm in vivo, characterized by solid, concave oral bulge, with the impression of doubled oral bulge in right side, macronuclear tortuous strand, usually 30 ciliary rows, 3 anteriorly differentiated to form a dorsal brush, and about 7 microm long rod-shaped extrusomes. S. simplinucleatum is highly variable in body size and length of monokinetidal bristle tail. Dileptus americanus is about 300 x 50 microm in vivo and differs from its congeners by a combination of characters including two macronuclear segments with a single micronucleus in between, two contractile vacuoles in dorsal side of trunk, two size types of rod-shaped extrusomes, a proboscis which occupies about 37% of body length, circular oral opening, and an average of 22 ciliary rows, of which up to 4 are differentiated to form a dorsal brush.

Tardigrades from Nahuel Huapi National Park (Argentina, South America) with descriptions of two new Macrobiotidae species.

In 31 samples of mosses and lichens collected in the Argentinean province of Río Negro, 657 tardigrades, 53 exuviae and 219 free-laid eggs were found. In total, 20 species were identified: Diphascon chilenense, Dip. mitrense, Echiniscus bigranulatus, Ech. corrugicaudatus, Ech. merokensis merokensis, Ech. testudo, Hebesuncus mollispinus, Hypsibius convergens, Macrobiotus cf. anderssoni, Mac. andinus, Mac. kazmierskii, Mac. patagonicus, Mesobiotus szeptyckii, Mes. pseudoblocki sp. nov., Milnesium argentinum, Mil. beatae, Mil. brachyungue, Mil. granulatum, Mopsechiniscus granulosus, Minibiotus pseudostellarus sp. nov. Of the two new species, Mesobiotus pseudoblocki sp. nov. is most similar to Mes. blocki, but it differs mainly by the lack of dentate lunules, smaller eggs and presence of reticular design on egg processes. Minibiotus pseudostellarus sp. nov. is most similar to Min. constellatus, Min. eichhorni, Min. sidereus or Min. vinciguerrae, but it differs from them by the presence of 'pseudo-star'-shaped pores in the dorsal cuticle instead of fully developed 'stars' and by other morphometric characters.

A new species of Cangshanaltica Konstantinov et al., a moss-inhabiting flea beetle from Thailand (Coleoptera: Chrysomelidae: Galerucinae: Alticini).

Moss cushions represent an interesting, but poorly understood habitat, which hosts many species of flea beetles (Coleoptera: Chrysomelidae: Galerucinae: Alticini). However, the diversity of moss-inhabiting flea beetles is not well studied, and collecting in tropical and subtropical locations that were not sampled in the past led to the discovery of many new species (Konstantinov et al. 2013). Here, a new species of a moss-inhabiting flea beetle from the genus Cangshanaltica Konstantinov et al. 2013 is described and illustrated. This genus is one of the recently described moss-inhabiting flea beetle genera and before this study, only one species was known (Konstantinov et al., 2013). This publication raises the number of flea beetle species that are known to occur in moss cushions around the world to 30, distributed among 15 genera.

Two new species of Eustigmaeus Berlese (Acari: Trombidiformes: Stigmaeidae) from Brazil, with a key to the American species.

Two new species of Eustigmaeus, E. piracicabensis Paktinat-Saeij & Bagheri sp. nov., and E. oliveirai Paktinat-Saeij & Bagheri sp. nov., are described based on females collected from soil, humus and moss in the São Paulo state, Brazil. Also, a key to the American Eustigmaeus species is provided.

A new species of Neparholaspis (Acari: Parholaspididae) from Russia, with additional information on Neparholaspis evansi Krantz, 1960.

Neparholaspis dubatolovi sp. nov. is described and illustrated from adult females and males collected from litter and moss in montane forest in north-eastern Sikhote-Alin Ridge in the Far East of Russia. Additional morphological information and illustrations of Neparholaspis evansi Krantz, 1960 are provided, based on examination of a paratype. A key to the world species of Neparholaspis is provided.

Mesobiotus philippinicus sp. nov., the first limnoterrestrial tardigrade from the Philippines.

The limnoterrestrial tardigrade fauna of the Philippines is completely unknown. In this paper, we describe the first ever limnoterrestrial water bear species from this southeast Asian country, Mesobiotus philippinicus sp. nov., found in a moss sample collected in Quezon City. Apart from morphometrics and imaging in light microscopy, we also analysed the new species under scanning electron microscope and sequenced four DNA markers differing in mutation rates, three nuclear (18S rRNA, 28S rRNA, and ITS-2) and one mitochondrial (COI). This allowed not only a detailed description but also provided barcodes to aid future species identification. The new species belongs to the harmsworthi group and is most similar to M. diffusus (Binda & Pilato, 1987), M. pseudocoronatus (Pilato et al., 2006), M. montanus (Murray, 1910) and M. mottai (Binda & Pilato, 1994), but differs from these species by whorled egg processes and dimensions of some morphometric traits. The 28S rRNA, ITS-2 and COI sequences presented in this paper are the first published DNA sequences for the genus Mesobiotus.

Bryophyte-feeding of Litoleptis (Diptera: Rhagionidae) with descriptions of new species from Japan.

Here we report the larval phytophagous habit of Litoleptis for the first time, and describe six new species of Litoleptis in Japan; L. japonica n. sp., L. kiiensis n. sp., L. niyodoensis n. sp., L. himukaensis n. sp., L. izuensis n. sp., and L. asterellaphile n. sp. All the species described here are thallus-miners of liverworts belonging to Aytoniaceae and Conocephalaceae (Marchantiopsida: Marchantiophyta). Each fly species mined thalli of only one of the following genera: Conocephalum, Reboulia, and Asterella. The descriptions of the Japanese Litoleptis species here expand the concept of this genus. The female genital morphology of Litoleptis strengthened the current placement of Litoleptis as a member of Spaniinae.

Diversity and community ecology of forest epiphyte testate amoebae from European Russia.

Testate amoebae are an abundant group of microorganisms which make a significant contribution to the diversity of protist life. Most of the world's potential habitats for testate amoebae have been barely studied and when such places are investigated they frequently reveal novel communities and species. Here we consider the testate amoeba communities associated with boreal forest epiphytes (mosses and lichens); an environment which we argue has been under-researched. We present a dataset of 165 samples from four regions of western Russia and analyse these data in relation to micro-habitat position and selected environmental data. The testate amoebae of epiphytes are abundant but dominated by ubiquitous species. We show that there are trends toward a lower species richness and test concentration with greater elevation on the trunk and in lichens compared to mosses. There are considerable differences in community composition between sampling regions. Of all measured environmental variables only moisture content showed a significant relationship with testate amoeba community structure. Our data highlight how little is known about testate amoeba communities of this habitat and call for greater research efforts, particularly in less-studied regions and biomes.

Testate amoebae (Protista) communities in Hylocomium splendens (Hedw.) B.S.G. (Bryophyta): relationships with altitude, and moss elemental chemistry.

We studied the testate amoebae in the moss Hylocomium splendens along an altitudinal gradient from 1000 to 2200 m asl. in the south-eastern Alps of Italy in relation to micro- and macro-nutrient content of moss plants. Three mountainous areas were chosen, two of them characterised by calcareous bedrock, the third by siliceous bedrock. A total of 25 testate amoebae taxa were recorded, with a mean species richness of 9.3 per sampling plot. In a canonical correspondence analysis, 63.1% of the variation in the amoebae data was explained by moss tissue chemistry, namely by C, P, Ca, Mg, Al, Fe, and Na content and a binary site variable. We interpreted this result as an indirect effect of moss chemistry on testate amoebae through an influence on prey organisms. Although two species responded to altitude, there was no overall significant relationship between testate amoebae diversity or community structure and altitude, presumably because our sampling protocol aimed at minimizing the variability due to vegetation types and soil heterogeneity. This suggests that previous evidence of altitudinal or latitudinal effects on testate amoebae diversity may at least in part be due to a sampling bias, namely differences in soil type or moss species sampled.

Relationship between testate amoeba (protist) communities and atmospheric heavy metals accumulated in Barbula indica (bryophyta) in Vietnam.

We studied the relationships between testate amoeba communities and heavy metal (Pb, Cd, Zn, Ni, Cu, Mn, and Fe) concentrations in the moss Barbula indica sampled at 29 sites in and around the city of Hanoi (Vietnam). Our first approach was to compare the heavy metal concentrations and testate amoeba variables between the city (zone 1) and the surrounding (zone 2). Mean moss concentrations of Pb, Cd, Zn, Ni, and Cu were significantly higher and testate amoeba species richness and abundance were significantly lower in zone 1 and the abundance of eight taxa differed significantly between the two zones. We then studied the correlation between heavy metals and testate amoebae. Species richness and abundance were correlated negatively to Pb concentration. Shannon H' was negatively correlated to both Pb and Cd. The abundance of several species was negatively correlated with Pb, Cd, Zn, and Ni; however, at the community level, Pb emerged as the only significant variable in a redundancy analysis. Our results suggest that testate amoebae are sensitive to and may be good bioindicators for heavy metal pollution, especially lead. Further research is needed to understand the causal relationships underlying the observed patterns.

Microhabitat selection in the simple oribatid community dwelling in epilithic moss cover (Acari: Oribatida).

The moss cover of a roof was studied as the model of a simple habitat divided into microhabitats by the members of a community of saprophagous mites. This community consisted of two species of oribatid mites: Scutovertex minutus and Trichoribates trimaculatus. They were extracted from moss onto moist paper, and subsequently, their mobility, responses to moisture fluctuation, and food selection were tested in laboratory experiments. For the nutritional biology, the microanatomy of their alimentary tract was examined according to the system of histological characteristics formulated in the laboratory of the author (type of food, digestive activity of gut walls, etc.). The paraplast sections of the mites were stained by Masson triple stain for these purposes. Moreover, the enzymological tests (chitinase and cellulase activities) were performed to detail the digestive processes. Such an approach was applied to field-sampled specimens as well as to those in the laboratory experiments. These above-mentioned data were discussed with respect to microhabitat selection, vertical and horizontal distribution, and dispersal ability of these two oribatid mites sharing this habitat. These two species differ in several characteristics from each other and these differences resulted in their different microhabitat choices and role in the habitat as a whole.