Prevalent Ciliate Symbiosis on Copepods: High Genetic Diversity and Wide Distribution Detected Using Small Subunit Ribosomal RNA Gene

Toward understanding the genetic diversity and distribution of copepod-associated symbiotic ciliates and the evolutionary relationships with their hosts in the marine environment, we developed a small subunit ribosomal RNA gene (18S rDNA)-based molecular method and investigated the genetic diversity and genotype distribution of the symbiotic ciliates on copepods. Of the 10 copepod species representing six families collected from six locations of Pacific and Atlantic Oceans, 9 were found to harbor ciliate symbionts. Phylogenetic analysis of the 391 ciliate 18S rDNA sequences obtained revealed seven groups (ribogroups), six (containing 99% of all the sequences) belonging to subclass Apostomatida, the other clustered with peritrich ciliate Vorticella gracilis. Among the Apostomatida groups, Group III were essentially identical to Vampyrophrya pelagica, and the other five groups represented the undocumented ciliates that were close to Vampyrophrya/Gymnodinioides/Hyalophysa. Group VI ciliates were found in all copepod species but one (Calanus sinicus), and were most abundant among all ciliate sequences obtained, indicating that they are the dominant symbiotic ciliates universally associated with copepods. In contrast, some ciliate sequences were found only in some of the copepods examined, suggesting the host selectivity and geographic differentiation of ciliates, which requires further verification by more extensive sampling. Our results reveal the wide occurrence and high genetic diversity of symbiotic ciliates on marine copepods and highlight the need to systematically investigate the host- and geography-based genetic differentiation and ecological roles of these ciliates globally.     

Role of anaerobic ciliates in planktonic food webs: abundance, feeding, and impact on bacteria in the field

We studied the dynamics of two populations of anaerobic ciliates, Plagiopyla sp. and Metopus sp., and of their potential prey, heterotrophic and phototrophic purple bacteria, in Lake Cisó throughout a 1-year cycle. The abundance of both ciliates was very low (less than 2 individuals per ml). During mixing, Plagiopyla ciliates exhibited high clearance rates (about 100 nl ciliate h), its integrated abundance increased with a net doubling time of 47 days, and its potential doubling times, as calculated from the number of bacteria consumed, ranged between 5 and 8 days. During stratification, the activity of Plagiopyla ciliates was reduced and the population decreased; this was related to the higher amounts of sulfide present. The impact of predation by the Plagiopyla population on bacterioplankton was found to be insignificant, less than 0.1% of bacterial biomass consumed per day. Thus, anaerobic ciliates cannot control the bacterioplankton in Lake Cisó because of both the low abundance over the period studied and the low feeding rates during certain periods. A review of available field studies suggests that this conclusion can be extrapolated to most other anoxic systems.     

西藏三种苔藓栖生纤毛虫的形态和系统发育分析

本研究利用活体观察和蛋白银染色方法对采自西藏日喀则和那曲的3种苔藓栖生纤毛虫,鬃异源棘尾虫(Tetmemena pustulata)、棘毛虫未定种(Sterkiella sp.)和殖口虫未定种(Gonostomum sp.)进行了形态学研究,描述了活体、核器以及纤毛图式等形态学特征.同时,测定了这3种纤毛虫的SSU r...     

Composition,diversity and distribution of microbenthos across the intertidal zones of Ryazhkov Island (the White Sea)

The composition and distribution of the main unicellular eukaryotic groups (diatom algae, ciliates, dinoflagellates (DF), other phototrophic (PF) and heterotrophic flagellates (HF)) were investigated in sandy sediments at five stations allocated across the tidal sheltered beach of the White Sea. Overall, 75 diatoms, 98 ciliates, 16 DF, 3 PF and 34 HF species were identified; some are new records for the White Sea. Common species for each group are illustrated. Diatoms and ciliates showed high alpha-diversity (species richness per sample), whereas flagellates were characterized by high beta-diversity (species turnover across the intertidal flat). Each group demonstrated its own spatial pattern that was best matched with its own subset of abiotic variables, reflecting group-specific responses to environmental gradients. Species richness increased from the upper intertidal zone seaward for ciliates but decreased for HF, whereas autotrophs showed a relatively uniform pattern with a slight peak at the mid-intertidal zone. Across the littoral zone, all groups showed distinct compositional changes; however, the position of the boundary between “upper” and “lower” intertidal communities varied among groups. Most of the species found at Ryazhkov Island are known from many other regions worldwide, indicating a wide geographic distribution of microbial eukaryotic species.     

八种纤毛虫减数分裂基因的分子进化研究

减数分裂是真核生物适应性进化的重要机制,以8种纤毛虫作为实验对象,通过生物信息学方法对其14个减数分裂基因进行了鉴定及分子进化研究。结果表明:(1)不同的纤毛虫种类存在一些特异性的减数分裂基因的丢失与复制现象;(2)减数分裂相关基因在纤毛虫中很保守;(3)纤毛虫减数分裂重要的同源重组过程是在真核生物中不常见的Ⅱ型。本研究表明,纤毛虫减数分裂可能代表了真核生物较原始的减数分裂方式,在进化的过程中很保守,为研究真核生物减数分裂起源与进化提供了重要线索。     

Laboratory experiments on the impact of biofilms on the plankton of a large river

1. Benthic organisms can have a strong effect on the plankton in rivers, although normally only members of the macrofauna are considered as important consumers. In the present study we conducted experiments on four different dates (in December, March, June and September) to assess the potential role of periphytic heterotrophic flagellates (HF), ciliates and rotifers in the control of potamoplankton (bacteria, algae, HF and ciliates). 2. Natural periphyton was established on the walls of circular flow channels by exposing them to river water (River Rhine, Germany). The experimental channels (with periphyton) and control channels (without periphyton) were filled with riverine water and the increase rates of planktonic bacteria, algae, HF and oligotrich ciliates were determined for the two treatments. 3. The abundance of periphytic ciliates and rotifers at the beginning of the four experiments showed large differences with low values in December and March, and high values in June and September. Dominant potential consumers of plankton were the heterotrich ciliate Stentor sp. and bdelloid rotifers. 4. The rates of increase of planktonic algae, HF and ciliates were significantly smaller in the presence of periphyton compared with those in their absence. Significant interactions between the treatment (with and without periphyton) and the time of experiment were found for the planktonic HF and algae, indicating that the impact of the periphyton varied temporarily. The planktonic groups responded differently to the periphyton with the planktonic HF showing the highest loss rate. Significant differences were also found among the loss rates of different HF groups and different diatom size classes. 5. These laboratory experiments demonstrate that periphytic ciliates and rotifers are potentially important consumers of different planktonic groups. The quantitative impact of periphyton on plankton with respect to the selective feeding needs further attention.     

Phylogeny of the Rumen Ciliates Entodinium, Epidinium and Polyplastron (Litostomatea: Entodiniomorphida) Inferred from Small Subunit Ribosomal RNA Sequences

ABSTRACT. Three complete 18S ribosomal RNA gene sequences from the rumen ciliates, Entodinium coudatum (1,639 bp), Epidinium caudarum (1,638 bp), and Polyplastron multivesiculatum (1,640 bp) were determined and confrimed in the opposite direction. Trees produced using maximum parsimony and distance-matrix methods (lest squares and neighbour-joining). with strong bootstrap support, depict the rumen ciliates as a monophyletic group, Entodinium caudatum is the earliest branching rumen ciliate. However, Entodiniwn simplex does not pair with En. caudatum , but rather with Polyplastron multivesiculatum. Signature sequences for these rumen ciliates reveal that the published SSrRNA gene sequence from En. simplex is in fact a Polyoplastron species. The free-living haptorian ciliates, Loxophyllum, Homalozoon and Spathidium (Subclass Hoptoria), are monophyletic and are the sister group to the rumen cilates. The litostomes (class Litostomatea), consisting of the haptorians and the rumen ciliates, are also a monophyletic group.     

Trophic interactions within the microbial food web in a tropical floodplain lake (Laguna Bufeos, Bolivia)

Whether the primary role of bacterioplankton is to act as "remineralizers" of nutrients or as direct nutritional source for higher trophic levels will depend on factors controlling their production and abundance. In tropical lakes, low nutrient concentration is probably the main factor limiting bacterial growth, while grazing by microzooplankton is generally assumed to be the main loss factor for bacteria. Bottom-up and top-down regulation of microbial abundance was studied in six nutrient limitation and dilution gradient-size fractionation in situ experiments. Bacteria, heterotrophic nanoflagellates (HNF), ciliates and rotifers showed relatively low densities. Predation losses of HNF and ciliates accounted for a major part of their daily production, suggesting a top-down regulation of protistan populations by rotifers. Phosphorus was found to be strongly limiting for bacterial growth, whereas no response to enrichment with Nitrogen or DOC was detected. HNF were the major grazers on bacteria (g-0.43 d(-1)), the grazing coefficient increased when ciliates were added (g- 0.80 d(-1)) but decreased when rotifers were added (g- 0.23 d(-1)) probably due to nutrient recycling or top-down control of HNF and ciliates by rotifers.     

Philasterides dicentrarchi, a histophagous ciliate causing scuticociliatosis in olive flounder, Philasterides dicentrarchi--histopathology investigations

Philasterides dicentrarchi is a histophagous scuticociliate infecting Korean olive flounder farms in Jeju Island, South Korea, where it causes significant economic losses. However, the route of entry of these parasites in olive flounder is currently unknown. In the present study, we attempted experimental infection with different doses (concentrations) of P. dicentrarchi by intraperitoneal (IP) injection; maximum cumulative mortality of 95% was recorded on the administration of 2.1×10(8) ciliates/ml. In small size group (3 cm length, 24±2 gwt) the ciliates were found in connective tissues of muscle, fins, and nervous tissue while in large size group (5 cm length, 47±3 gwt), the ciliates were predominantly occurred with associated symptoms of liquefaction in the central nervous system. The major clinico-pathological manifestations were loss of scales, appearance of bleached spots that coalesced to form brownish patches, hemorrhagic, and severe dermal necrotic lesion. In addition there was degeneration of muscle fibers, necrosis of the hepatic parenchyma, and severe edema of the intestinal walls, extensive fouling, necrotic degeneration and hyperplasia in the gill bronchial epithelium. In this study, many ciliates were found in the blood vessel, brain, dorsal fins, muscle, kidney, and vertebra of infected flounder. The histopathological investigations are of significant importance in view of possible routes of entry into the host and doses of ciliates that trigger fast infection for potential utility of drugs as a strategy for the control of P. dicentrarchi in farmed olive flounder.     

Do microbes have macroecology? Large‐scale patterns in the diversity and distribution of marine benthic ciliates

Aim The biogeography and global distribution of protists has long been disputed, with two primary, opposing views. To test these two sets of views in greater detail, we have compiled the available data for marine benthic ciliates and assessed the general patterns of their diversity and distribution compared with Metazoa. Location World‐wide. Methods A comprehensive database (1342 species, over 350 sources) was used to analyse the diversity, distribution, species occurrences and range size distribution of free‐living ciliates that inhabit marine sediments in 17 geographical regions. Results Twenty‐five per cent of the species have been found in a single region only, whereas 18% are widespread (they occur in more than half the regions covering both hemispheres). Only 5–7% of regional faunas are endemic, which is much lower than for macroorganisms. Regional diversity depends neither on total area nor on coastline length and does not show any obvious latitudinal trends, but correlates highly with the investigation effort expended in a region and (negatively) with the average salinity. A comparison of species composition reveals distinctions between the Arctic Area (the White, Barents and Kara seas), Laurasian Area (north Atlantic, north Pacific and European seas), Gondwanian Area (Southern Ocean) and the Antarctic. No clear geographical correlations are found for faunistic composition at the genus or family levels. There is the tendency to narrow the latitudinal ranges for species found at high latitudes (reversal of Rapoport's rule). Main conclusions Undersampling and data insufficiency are the key factors affecting the observed diversity and distribution of microorganisms. Nevertheless, marine benthic ciliates demonstrate certain patterns that generally agree with the ‘moderate endemicity’ model ( Foissner, 2004, 2008 ), but consistently contradict the regularities commonly observed for multicellular taxa. Thus, ciliates do have a biogeography, but their macroecological patterns may be different in some respects from that of macroorganisms.     

春季和秋季东海陆架区浮游纤毛虫的丰度和生物量

于2011年春季(5月)和秋季(11月)在东海陆架区进行浮游纤毛虫丰度和生物量的调查.春季和秋季纤毛虫的平均丰度分别为(614±861)和(934±809) ind·L^-1,平均生物量分别为(1.70±3.91)和(0.93±0.99) μg C·L^-1.表层纤毛虫丰度和生物量的高值区春季主要分布在近岸及远岸海区,秋季主要分布在远岸海区.春季纤毛虫的丰度和生物量在水体上层较高;秋季纤毛虫主要分布在水体上层,有时在水体底层也会出现丰度和生物量的高值.春季无壳纤毛虫群落的粒级较大,秋季较小.砂壳纤毛虫占纤毛虫丰度的平均比例春季和秋季分别为(26.9±34.3)%和(44.9±25.2)%.两个季节共鉴定出砂壳纤毛虫27属52种,春季丰度较大的种为原始筒壳虫、橄榄领细壳虫及筒状拟铃虫,秋季丰度较大的种为原始筒壳虫、小领细壳虫及矮小拟铃虫.纤毛虫丰度与温度、叶绿素a(Chl a)浓度呈显著正相关.砂壳纤毛虫丰度与盐度呈显著负相关,群落结构变化与温度显著相关.     

Monsoonal and lunar variability in microzooplankton abundance and community structure in the Terusan mangrove creek (Malaysia)

This study documents the monsoonal and lunar effects on species composition and abundance of microzooplankton in a tropical estuary. We investigated microzooplankton abundance in relation to the various environmental and biotic parameters, sampled in the Matang mangrove (Malaysia) from April 2013 to February 2014. A total of 39 microzooplankton taxa comprising four major groups, i.e. loricate ciliates (37.72%), aloricate ciliates (29.46%), dinoflagellates (24.33%) and meroplanktonic nauplii (8.49%) were identified. The loricate ciliates were the most diverse group with 31 taxa recorded. Four major species of loricate ciliates were identified, i.e. Tintinnopsis beroidea, Tintinnopsis rotundata, Stenosemella avellana and Tintinnidium primitivum, while Strombidiidae and Strobilidiidae dominated the aloricate ciliates. Although small loricate ciliates were ubiquitous, redundancy analysis shows marked shifts in microzooplankton community structure, from one that was dominated by loricate ciliates during the drier SW monsoon, to aloricate ciliates at the onset of the wet NE monsoon, and then to dinoflagellates towards the end of the drier NE monsoon period. These shifts were associated with rainfall, dissolved inorganic nutrients, salinity, temperature and microbial food abundance. There was no clear lunar effect on abundance of microzooplankton except for Favella ehrenbergii and copepod nauplii, which were more abundant during neap than spring tides.     

Effects of temperature, sulfide, and food abundance on growth and feeding of anaerobic ciliates

The trophic role of ciliates in anaerobic food webs has not been assessed experimentally. In order to obtain basic information necessary to interpret field situations, we studied the effects of temperature, sulfide concentration, and food abundance on the growth and feeding activities of two anaerobic ciliates, Plagiopyla nasuta and Metopus es. The growth rate of P. nasuta increased with temperature from 8 to 18 degrees C (Q(10) = 2.0) and remained constant in the range between 18 and 24 degrees C (0.22 day). Sulfide concentrations of between 0 and 1 mM did not affect the feeding activities, but concentrations greater than 2 mM were inhibitory. The functional response of P. nasuta feeding on fluorescently labeled heterotrophic and phototrophic bacteria was investigated. In both cases, the parameters of the functional response were almost identical when expressed in terms of biovolume: the maximal uptake rate (U(m)) was 1,800 mum ciliate h and the half-saturation constant for ingestion (k) was 1.5 x 10 mum ml. The functional response of M. es feeding on heterotrophic bacteria was found to be similar to that of P. nasuta. These ciliates needed high bacterial abundances in order to maintain their growth (k of about 4 x 10 bacteria ml), implying that they will frequently be food limited in planktonic environments. Both the maximal uptake rates and the maximal clearance rates were comparable to those of aerobic ciliates. By combining the growth and feeding data, we estimated gross growth efficiencies of 12 and 13% for P. nasuta and M. es, respectively. These results indicate that the feeding rates of anaerobic ciliates are similar to those of aerobic ciliates. Their slower growth must, therefore, be due to the lower gross growth efficiency (likely due to anaerobic metabolism).     

Microbial genetic diversity and ciliate community structure along an environmental gradient in coastal soil

We investigated the microbial genetic diversity and ciliate community in coastal soil from five sites with an environmental gradient using denaturing gradient gel electrophoresis (DGGE), gene sequencing and the Ludox–QPS method. The analyses of both the 16S ribosomal RNA (rRNA) gene and 18S rRNA gene DGGE resulted in equal or even a higher number of bands found in the samples taken from the high-salinity sites IV and V than in those taken from the low-to-moderate-salinity sites I–III. Cluster analysis of both DGGE profiles classified the five sites into three main groups (sites I, II and III and IV and V), which corresponded well to the analysis of environmental factors. There were 13 species observed at site I, three species at site II and nine species at site III, while no active ciliates were observed at the high-salinity sites IV and V. By contrast, the ciliate-specific DGGE revealed a higher number of bands in the samples taken from the high-salinity soil. Furthermore, gene sequencing suggested that the ciliates in the high-salinity soil comprised forms originating not only from soil but also from marine environments. The data indicate that saline soil may maintain a high diversity of ciliates and soil salinity might be the most influential factor regulating the community structure of ciliates.     

The Impact of UV Radiation on Paramecium Populations from Alpine Lakes

Paramecium populations from a clear and a glacier‐fed turbid alpine lake were exposed to solar simulated ultraviolet (UVR) and photosynthetically active radiation (PAR) at 8 and 15 °C. The ciliates were tested for DNA damage (comet assay), behavioral changes, and mortality after UVR + PAR exposure. High DNA damage levels (~58% tail DNA) and abnormal swimming behavior were observed, although no significant changes in cell numbers were found irrespective of the lake origin (clear, turbid), and temperatures. We conclude that environmental stressors such as UVR and their effects may influence the adaptation of ciliates living in alpine lakes.     

Intestinal ciliate composition found in the feces of racing horses from Izmir, Turkey

Species composition and distribution of intestinal ciliates were investigated in the feces from 15 racing horses living near Izmir, Turkey. Thirty-seven species belonging to 21 genera were identified. Although no new species were observed, this is the first report on intestinal ciliates in racing horses living in Turkey. The mean number of ciliates was 26.4 ± 13.9 × 10(4) cells ml(-1) of feces and the mean number of ciliate species per host was 18.8 ± 7.1. No ciliates were observed in one horse. Bundleia and Polymorphella were found to be the two dominant genera, occurring in high proportions. In contrast, Didesmis and Prorodonopsis were only observed at a low frequency. Bundleia nana, Blepharoconus hemiciliatus, Paraisotrichopsis composita, Prorodonopsis coli and Spirodinium equi were newly recorded from Turkey.     

Can the surface immobilization antigens of Philasterides dicentrarchi (Ciliophora: Scuticociliatida) be used as target antigens to develop vaccines in cultured fish?

In order to test whether immobilization antigens (i-antigens) of Philasterides dicentrarchi could be suitable antigenic targets against scuticociliatosis, polyclonal olive flounder (Paralichthys olivaceus) sera were raised against P. dicentrarchi by immunization with lysates of ciliates grown using chinook salmon epithelial (CHSE) cells, and the ability of the immune sera to kill the ciliates via classical complement pathway was analyzed in relation to agglutination activity. The immune sera showed clear agglutination activity against the CHSE-cultured ciliates. However, the agglutinated ciliates were not killed but escaped from the agglutinated mass within a few hours. Ciliates isolated from fish artificially infected with the same population of CHSE-cultured ciliates were not agglutinated by the immune sera even at the lowest dilution. In antibody-dependent complement-mediated killing (ADCK), the immune sera completely killed the CHSE-cultured ciliates at relatively higher serum dilutions (showing low or no agglutination activity). However, CHSE-cultured ciliates were not killed completely at lower immune serum dilutions (showing high agglutination activity). In contrast to CHSE-cultured ciliates, the ciliates isolated from infected fish were killed at lower dilutions of the immune sera in spite of no agglutination response. Considering the presence of various i-antigen types, ability to change i-antigen type in response to corresponding antibody, and relatively low ADCK activity at high agglutination titer, i-antigens of P. dicentrarchi may not be good targets for subunit vaccine development. To develop subunit vaccines against scuticociliatosis, other surface antigens expressed constitutively or expressed specifically under the infection state for survival of the ciliates in the host fish might be more favorable to elicit protective antibodies than the surface i-antigens.     

Intestinal ciliate composition found in the feces of the Turk rahvan horse Equus caballus, Linnaeus 1758

Species composition and distribution of large intestinal ciliates were investigated in the feces from 15 Turk rahvan horses, living in the vicinity of Izmir, Turkey. Twenty-two ciliate genera consisting of 36 species were identified. This is the first report on intestinal ciliates in Turk rahvan horses and no previously unknown species were observed. The mean number of ciliates was 14.2 ± 13.9 × 10⁴ cells ml⁻¹ of feces and the mean number of ciliate species per host was 9.9 ± 7.1. No ciliates were observed in 2 horses. Bundleia and Blepharocorys were considered to be the major genera since these ciliates were constantly found in high proportions. In contrast, Paraisotricha, Didesmis and Gassovskiella were only observed at low frequencies. The ciliates found in this survey had almost the same characteristics as those described in previous reports, suggesting that there was no significant geographic variation in the intestinal ciliate fauna of equids.     

Benthic–pelagic coupling: a comparison of the community structure of benthic and planktonic heterotrophic protists in shallow inlets of the southern Baltic

1. The taxonomic composition, abundance and biomass of heterotrophic protists (ciliates, heterotrophic flagellates (HF), rhizopods and actinopods) in the sediment and water column of shallow inlets of the Southern Baltic was studied under a variety of environmental conditions during 1996–1997. A shallow, highly eutrophic station and a deeper, less eutrophic station were compared.
2. Community biomass ranged from 0.12 to 0.34 μg C cm?3 in the water column and from 1.5 to 105 μg C cm?3 in the sediment. Heterotrophic protists dominated zooplankton biomass at both stations (73% and 84% mean contribution), while they were of minor importance within the zoobenthos. Expressed per unit area, benthic biomass contributed a significant part (44% and 49%) to the total heterotrophic protistan community at both stations.
3. Although the methodology for counting ciliates and HF was focussed on a high taxonomic resolution, the results reveal some general trends in the distribution of heterotrophic protists: protozooplankton biomass was dominated by flagellates (80% mean biomass contribution) at the shallow station and by ciliates (73% mean biomass contribution) at the deep station. In the benthos at both stations, ciliates were the dominant protozoans, followed by the hitherto little‐studied rhizopods (25% and 35% mean biomass contribution) and flagellates.
4. The degree of benthic–pelagic coupling differed between taxonomic groups. Benthic and pelagic communities of ciliates showed little taxonomic overlap. In contrast, many heterotrophic flagellate species were found both in the benthos and in the pelagic. These benthic–pelagic species contributed significantly to the biomass of HF in the water column. The planktonic rhizopod community consisted of a subset of those species found in the benthos.
5. The abundance of benthic and pelagic protists was positively correlated at the shallow station, but taxonomic data indicate that the direct exchange between benthic and pelagic communities was only partly responsible.     

Evolution of anaerobic ciliates from the gastrointestinal tract: phylogenetic analysis of the ribosomal repeat from Nyctotherus ovalis and its relatives

The 18S and 5.8S rDNA genes and the internal transcribed spacers ITS-1 and ITS-2 of ciliates living in the hindgut of frogs, millipedes, and cockroaches were analyzed in order to study the evolution of intestinal protists. All ciliates studied here belong to the genus Nycrotherus. Phylogenetic analysis revealed that these ciliates from a monophyletic group that includes the distantly related anaerobic free-living heterotrichous ciliates Metopus palaeformis and Metopus contortus. The intestinal ciliates from the different vertebrate and invertebrate hosts are clearly divergent at the level of their rDNA repeats. This argues for the antiquity of the associations and a predominantly vertical transmission. This mode of transmission seems to be controlled primarily by the behavior of the host. The different degrees of divergence between ciliates living in different strains of one and the same cockroach species most likely reflect the different geographical origins of the hosts. In addition, host switches must have occurred during the evolution of cockroaches, since identical ciliates were found only in distantly related hosts. These phenomena prevent the reconstruction of potential cospeciation events.