"Candidatus Intestinibacterium parameciiphilum"-member of the "Candidatus Paracaedibacteraceae" family (Alphaproteobacteria, Holosporales) inhabiting the ciliated protist Paramecium.

Protists frequently host diverse bacterial symbionts, in particular those affiliated with the order Holosporales (Alphaproteobacteria). All characterised members of this bacterial lineage have been retrieved in obligate association with a wide range of eukaryotes, especially multiple protist lineages (e.g. amoebozoans, ciliates, cercozoans, euglenids, and nucleariids), as well as some metazoans (especially arthropods and related ecdysozoans). While the genus Paramecium and other ciliates have been deeply investigated for the presence of symbionts, known members of the family "Candidatus Paracaedibacteraceae" (Holosporales) are currently underrepresented in such hosts. Herein, we report the description of "Candidatus Intestinibacterium parameciiphilum" within the family "Candidatus Paracaedibacteraceae", inhabiting the cytoplasm of Paramecium biaurelia. This novel bacterium is almost twice as big as its relative "Candidatus Intestinibacterium nucleariae" from the opisthokont Nuclearia and does not present a surrounding halo. Based on phylogenetic analyses of 16S rRNA gene sequences, we identified six further potential species-level lineages within the genus. Based on the provenance of the respective samples, we investigated the environmental distribution of the representatives of "Candidatus Intestinibacterium" species. Obtained results are consistent with an obligate endosymbiotic lifestyle, with protists, in particular freshwater ones, as hosts. Thus, available data suggest that association with freshwater protists could be the ancestral condition for the members of the "Candidatus Intestinibacterium" genus.

Morphology, ultrastructure, genomics, and phylogeny of Euplotes vanleeuwenhoeki sp. nov. and its ultra-reduced endosymbiont "Candidatus Pinguicoccus supinus" sp. nov.

Taxonomy is the science of defining and naming groups of biological organisms based on shared characteristics and, more recently, on evolutionary relationships. With the birth of novel genomics/bioinformatics techniques and the increasing interest in microbiome studies, a further advance of taxonomic discipline appears not only possible but highly desirable. The present work proposes a new approach to modern taxonomy, consisting in the inclusion of novel descriptors in the organism characterization: (1) the presence of associated microorganisms (e.g.: symbionts, microbiome), (2) the mitochondrial genome of the host, (3) the symbiont genome. This approach aims to provide a deeper comprehension of the evolutionary/ecological dimensions of organisms since their very first description. Particularly interesting, are those complexes formed by the host plus associated microorganisms, that in the present study we refer to as "holobionts". We illustrate this approach through the description of the ciliate Euplotes vanleeuwenhoeki sp. nov. and its bacterial endosymbiont "Candidatus Pinguicoccus supinus" gen. nov., sp. nov. The endosymbiont possesses an extremely reduced genome (~ 163 kbp); intriguingly, this suggests a high integration between host and symbiont.

"Candidatus Trichorickettsia mobilis", a Rickettsiales bacterium, can be transiently transferred from the unicellular eukaryote Paramecium to the planarian Dugesia japonica.

Most of the microorganisms responsible for vector-borne diseases (VBD) have hematophagous arthropods as vector/reservoir. Recently, many new species of microorganisms phylogenetically related to agents of VBD were found in a variety of aquatic eukaryotic hosts; in particular, numerous new bacterial species related to the genus Rickettsia (Alphaproteobacteria, Rickettsiales) were discovered in protist ciliates and other unicellular eukaryotes. Although their pathogenicity for humans and terrestrial animals is not known, several indirect indications exist that these bacteria might act as etiological agents of possible VBD of aquatic organisms, with protists as vectors. In the present study, a novel strain of the Rickettsia-Like Organism (RLO) endosymbiont "Candidatus (Ca.) Trichorickettsia mobilis" was identified in the macronucleus of the ciliate Paramecium multimicronucleatum. We performed transfection experiments of this RLO to planarians (Dugesia japonica) per os. Indeed, the latter is a widely used model system for studying bacteria pathogenic to humans and other Metazoa. In transfection experiments, homogenized paramecia were added to food of antibiotic-treated planarians. Treated and non-treated (i.e. control) planarians were investigated at day 1, 3, and 7 after feeding for endosymbiont presence by means of PCR and ultrastructural analyses. Obtained results were fully concordant and suggest that this RLO endosymbiont can be transiently transferred from ciliates to metazoans, being detected up to day 7 in treated planarians' enterocytes. Our findings might offer insights into the potential role of ciliates or other protists as putative vectors for diseases caused by Rickettsiales or other RLOs and occurring in fish farms or in the wild.

Role of bacterivorous organisms on fungal-based systems for natural tannin degradation.

Tannery wastewater presents high concentrations of organic load and pollutant recalcitrant molecules (e.g. tannins), which reduce the efficiency of biological treatment processes. Recent studies showed that several fungal species and strains are effective in the degradation of tannins. However, high bacterial load can negatively affect fungal growth, reducing system stability and degradation performances. The aim of the present study was to evaluate the effects of the introduction of bacterivorous grazers (ciliates and/or rotifers) in batch scale experiments using fungi to remove Tara tannin, i.e. to check the potential synergistic effect between fungi and bacterivorous grazers in the degradation of recalcitrant compounds. In this context, the ciliated grazers Paramecium calkinsi, Tetrahymena sp., Pseudovorticella sp., and the rotifer Lecane inermis, preliminary selected according to their ability to grow in a solution prepared with Tara tannin, were separately tested. Activated sludge, including a complex mixture of native grazers, was used as experimental control. The following parameters were monitored: bacterial load, number of grazers/mL and Soluble Chemical Oxygen Demand (SCOD). Colony Forming Unit (CFU)/grazers ratio was also calculated. Particular attention was paid to: i) bacterial load reduction and ii) enhancement of recalcitrant compounds degradation, and we observed that in all experimental conditions where grazers occurred bacterial load was significantly reduced and the system achieved a higher SCOD removal in a shorter time. Our findings provide useful insights for the stabilization of fungal-based systems in non-sterile conditions.

Diversity and environmental distribution of the cosmopolitan endosymbiont "Candidatus Megaira".

Members of the order Rickettsiales are often found in association with ciliated protists. An interesting case is the bacterial endosymbiont "Candidatus Megaira", which is phylogenetically closely related to the pathogen Rickettsia. "Candidatus Megaira" was first described as an intracellular bacterium in several ciliate species. Since then it has been found in association with diverse evolutionary distantly-related hosts, among them other unicellular eukaryotes, and also algae, and metazoa, such as cnidarians. We provide the characterization of several new strains of the type species "Candidatus Megaira polyxenophila", and the multidisciplinary description of a novel species, "Candidatus Megaira venefica", presenting peculiar features, which highlight the diversity and variability of these widespread bacterial endosymbionts. Screening of the 16S rRNA gene short amplicon database and phylogenetic analysis of 16S rRNA gene hypervariable regions revealed the presence of further hidden lineages, and provided hints on the possibility that these bacteria may be horizontally transmitted among aquatic protists and metazoa. The phylogenetic reconstruction supports the existence of at least five different separate species-level clades of "Candidatus Megaira", and we designed a set of specific probes allowing easy recognition of the four major clades of the genus.

Symbiont replacement between bacteria of different classes reveals additional layers of complexity in the evolution of symbiosis in the ciliate Euplotes.

Symbiosis is a diverse and complex phenomenon requiring diverse model systems. The obligate relationship between a monophyletic group of Euplotes species ("clade B") and the betaproteobacteria Polynucleobacter and "Candidatus Protistobacter" is among the best-studied in ciliates, and provides a framework to investigate symbiont replacements. Several other Euplotes-bacteria relationships exist but are less understood, such as the co-dependent symbiosis between Euplotes magnicirratus (which belongs to "clade A") and the alphaproteobacterium "Candidatus Devosia euplotis". Here we describe a new Devosia inhabiting the cytoplasm of a strain of Euplotes harpa, a clade B species that usually depends on Polynucleobacter for survival. The novel bacterial species, "Candidatus Devosia symbiotica", is closely related to the symbiont of E. magnicirratus, casting a different light on the history of bacteria colonizing ciliates of this genus. The two Devosia species may have become symbionts independently or as the result of a symbiont exchange between hosts, in either case replacing a previous essential bacterium in E. harpa. Alternatively, both may be remnants of an ancient symbiotic relationship between Euplotes and Devosia, in which case Polynucleobacter and "Ca. Protistobacter" are recent invaders. Either way, symbiont replacement between bacteria belonging to different classes must be evoked to explain this fascinating system.

Strengths and Biases of High-Throughput Sequencing Data in the Characterization of Freshwater Ciliate Microbiomes.

Molecular surveys of eukaryotic microbial communities employing high-throughput sequencing (HTS) techniques are rapidly supplanting traditional morphological approaches due to their larger data output and reduced bench work time. Here, we directly compare morphological and Illumina data obtained from the same samples, in an effort to characterize ciliate faunas from sediments in freshwater environments. We show how in silico processing affects the final outcome of our HTS analysis, providing evidence that quality filtering protocols strongly impact the number of predicted taxa, but not downstream conclusions such as biogeography patterns. We determine the abundance distribution of ciliates, showing that a small fraction of abundant taxa dominates read counts. At the same time, we advance reasons to believe that biases affecting HTS abundances may be significant enough to blur part of the underlying biological picture. We confirmed that the HTS approach detects many more taxa than morphological inspections, and highlight how the difference varies among taxonomic groups. Finally, we hypothesize that the two datasets actually correspond to different conceptions of "diversity," and consequently that neither is entirely superior to the other when investigating environmental protists.

"Candidatus Gortzia shahrazadis", a Novel Endosymbiont of Paramecium multimicronucleatum and a Revision of the Biogeographical Distribution of Holospora-Like Bacteria.

Holospora spp. and "Candidatus Gortzia infectiva", known as Holospora-like bacteria (HLB), are commonly found as nuclear endosymbionts of ciliates, especially the Paramecium genus. HLB are related by phylogenetic relationships, morphological features, and life-cycles, which involve two alternating morphotypes: reproductive and infectious forms (RF, IF). In this paper we describe a novel species belonging to the "Ca. Gortzia" genus, detected in P. multimicronucleatum, a ciliate for which infection by an HLB has not been reported, discovered in India. This novel endosymbiont shows unusual and surprising features with respect to other HLB, such as large variations in IF morphology and the occasional ability to reproduce in the host cytoplasm. We propose the name of "Candidatus Gortzia shahrazadis" for this novel HLB. Moreover, we report two additional species of HLB from Indian Paramecium populations: "Ca. Gortzia infectiva" (from P. jenningsi), and H. obtusa (from P. caudatum); the latter is the first record of Holospora from a tropical country. Although tropical, we retrieved H. obtusa at an elevation of 706 m corresponding to a moderate climate not unlike conditions where Holospora are normally found, suggesting the genus Holospora does exist in tropical countries, but restricted to higher elevations.

Ciliate communities and hidden biodiversity in freshwater biotopes of the Pistoia province (Tuscany, Italy).

Ciliates are essential components of aquatic environments, playing a pivotal role in microbial loops. Thus, the composition and dynamics of ciliate communities have been subjected to intense studying. Morphological methods have been traditionally employed, until the development of next-generation sequencing recently allowed to explore the topic with exclusively molecular techniques. However, the results of the two approaches are hardly comparable, and the pictures they offer can be quite different. This may be due, among other reasons, to two factors: (1) morphological descriptions may miss a large portion of "hidden biodiversity" (including rare species and resistance forms) that is detected instead by molecular methods; (2) identification errors may arise due to difficulties in recognizing microbial taxa without in-depth analyses. In this survey of freshwater systems of the Pistoia province (Tuscany, Italy) we address both issues, trying to quantify the hidden diversity through prolonged observations of differentially treated sample aliquots, combining morphological identification with Sanger sequencing. We provide the first insights into the ciliate fauna of this area presenting results that are suitable for future comparisons thanks to their multidisciplinary origin, and supply the first molecular data on well-known taxa such as Linostomella and Disematostoma.

A House for Two--Double Bacterial Infection in Euplotes woodruffi Sq1 (Ciliophora, Euplotia) Sampled in Southeastern Brazil.

Several ciliated protists form symbiotic associations with a diversity of microorganisms, leading to drastic impact on their ecology and evolution. In this work, two Euplotes spp. sampled in Rio de Janeiro, Brazil, were identified based on morphological and molecular features as Euplotes woodruffi strain Sq1 and E. encysticus strain Sq2 and investigated for the presence of endosymbionts. While E. woodruffi Sq1 stably hosts two bacterial populations, namely Polynucleobacter necessarius (Betaproteobacteria) and a new member of the family "Candidatus Midichloriaceae" (Alphaproteobacteria, Rickettsiales), here described as "Candidatus Bandiella woodruffii," branching with a broad host range bacterial group found in association with cnidarians, sponges, euglenoids, and some arthropods; in E. encysticus Sq2 no symbiotic bacterium could be detected. The dispersion ability of this novel bacterium was tested by co-incubating E. woodruffi Sq1 with three different ciliate species. Among the tested strains "Ca. B. woodruffii" could only be detected in association with E. encysticus Sq2 with a prevalence of 20 % after 1 week and 40 % after 2 weeks, maintaining this level for up to 6 months. Nevertheless, this apparent in vitro association was abolished when E. woodruffi Sq1 donor was removed from the microcosm, suggesting that this bacterium has the capacity for at least a short-term survival outside its natural host and the aptitude to ephemerally interact with other organisms. Together, these findings strongly suggest the need for more detailed investigations to evaluate the host range for "Ca. B. woodruffii" and any possible pathogenic effect of this bacterium on other organisms including humans.

Nitrifying biomass characterization and monitoring during bioaugmentation in a membrane bioreactor.

A membrane bioreactor (MBR), fed with domestic wastewater, was bioaugmented with nitrifying biomass selected in a side-stream MBR fed with a synthetic high nitrogen-loaded influent. Microbial communities evolution was monitored and comparatively analysed through an extensive bio-molecular investigation (16S rRNA gene library construction and terminal-restriction fragment length polymorphism techniques) followed by statistical analyses. As expected, a highly specialized nitrifying biomass was selected in the side-stream reactor fed with high-strength ammonia synthetic wastewater. The bioaugmentation process caused an increase of nitrifying bacteria of the genera Nitrosomonas (up to more than 30%) and Nitrobacter in the inoculated MBR reactor. The overall structure of the microbial community changed in the mainstream MBR as a result of bioaugmentation. The effect of bioaugmentation in the shift of the microbial community was also verified through statistical analysis.

Focusing on genera to improve species identification: revised systematics of the ciliate Spirostomum.

Although many papers dealing with the description of new ciliate taxa are published each year, species taxonomy and identification in most groups of the phylum Ciliophora remain confused. This is largely due to a scarcity of surveys on the systematics of immediately higher levels (genera and families) providing data for old and new species together. Spirostomum is a common and distinctive inhabitant of fresh- and brackish water environments, including artificial and eutrophic ones, and is a good model for applied ecology and symbiosis research. Despite this, only 3 of the numerous species are commonly cited, and no studies have yet confirmed their monophyly, with the consequence that reproducibility of the results may be flawed. In this paper we present morphological and molecular data for 30 Spirostomum populations representing 6 different morphospecies, some of which were collected in previously unreported countries. We performed a detailed revision of Spirostomum systematics combining literature surveys, new data on hundreds of organisms and statistical and phylogenetic analyses; our results provide insights on the evolution, ecology and distribution of known morphospecies and a novel one: Spirostomum subtilis sp. n. We also offer tools for quick species identification.

Free-living ciliates as potential reservoirs for eukaryotic parasites: occurrence of a trypanosomatid in the macronucleus of Euplotes encysticus.

BACKGROUND: Flagellates of the family Trypanosomatidae are obligate endoparasites, which can be found in various hosts. Several genera infect insects and occur as monoxenous parasites especially in representatives of Diptera and Hemiptera. These trypanosomatid flagellates probably share the worldwide distribution of their hosts, which are often infested by large numbers of endoparasites. Traditionally, their taxonomy was based on morphology, host origin, and life cycle. Here we report the characterization of a trypanosomatid infection detected in a protozoan, a ciliate collected from a polluted freshwater pond in a suburb of New Delhi (India). METHODS: Live observations and morphological studies applying light, fluorescence and transmission electron microscopy were conducted. Molecular analyses of host and parasite were performed and used for phylogenetic reconstructions and species (host) or genus level (parasite) identification. RESULTS: Although the morphological characteristics were not revealing, a high similarity of the trypanosomatids 18S rRNA gene sequence to Herpetomonas ztiplika and Herpetomonas trimorpha (Kinetoplastida, Trypanosomatidae), both parasites of biting midges (Culicoides kibunensis and Culicoides truncorum, respectively) allowed the assignment to this genus. The majority of the host population displayed a heavy infection that significantly affected the shape of the host macronucleus, which was the main site of parasite localization. In addition, the growth rate of host cultures, identified as Euplotes encysticus according to cell morphology and 18S rRNA gene sequence, was severely impacted by the infection. CONCLUSIONS: The host-parasite system described here represents a recent example of free-living protists acting as environmental reservoirs for parasitic eukaryotic microorganisms.

Bacterial communities in polluted seabed sediments: a molecular biology assay in Leghorn Harbor.

Seabed sediments of commercial ports are often characterized by high pollution levels. Differences in number and distribution of bacteria in such areas can be related to distribution of pollutants in the port and to sediment conditions. In this study, the bacterial communities of five sites from Leghorn Harbor seabed were characterized, and the main bacterial groups were identified. T-RFLP was used for all samples; two 16S rRNA libraries and in silico digestion of clones were used to identify fingerprint profiles. Library data, phylogenetic analysis, and T-RFLP coupled with in silico digestion of the obtained sequences evidenced the dominance of Proteobacteria and the high percentage of Bacteroidetes in all sites. The approach highlighted similar bacterial communities between samples coming from the five sites, suggesting a modest differentiation among bacterial communities of different harbor seabed sediments and hence the capacity of bacterial communities to adapt to different levels and types of pollution.

Rediscovering the genus Lyticum, multiflagellated symbionts of the order Rickettsiales.

Among the bacterial symbionts harbored by the model organism Paramecium, many still lack a recent investigation that includes a molecular characterization. The genus Lyticum consists of two species of large-sized bacteria displaying numerous flagella, despite their inability to move inside their hosts' cytoplasm. We present a multidisciplinary redescription of both species, using the deposited type strains as well as newly collected material. On the basis of 16S rRNA gene sequences, we assigned Lyticum to the order Rickettsiales, that is intensely studied because of its pathogenic representatives and its position as the extant group most closely related to the mitochondrial ancestor. We provide conclusive proofs that at least some Rickettsiales possess actual flagella, a feature that has been recently predicted from genomic data but never confirmed. We give support to the hypothesis that the mitochondrial ancestor could have been flagellated, and provide the basis for further studies on these ciliate endosymbionts.

'Candidatus Megaira polyxenophila' gen. nov., sp. nov.: considerations on evolutionary history, host range and shift of early divergent rickettsiae.

"Neglected Rickettsiaceae" (i.e. those harboured by non-hematophagous eukaryotic hosts) display greater phylogenetic variability and more widespread dispersal than pathogenic ones; yet, the knowledge about their actual host range and host shift mechanism is scarce. The present work reports the characterization following the full-cycle rRNA approach (SSU rRNA sequence, specific in situ hybridization, and ultrastructure) of a novel rickettsial bacterium, herewith proposed as 'Candidatus Megaira polyxenophila' gen. nov., sp. nov. We found it in association with four different free-living ciliates (Diophrys oligothrix, Euplotes octocarinatus, Paramecium caudatum, and Spirostomum sp., all belonging to Alveolata, Ciliophora); furthermore it was recently observed as intracellular occurring in Carteria cerasiformis and Pleodorina japonica (Chlorophyceae, Chlorophyta). Phylogenetic analyses demonstrated the belonging of the candidate new genus to the family Rickettsiaceae (Alphaproteobacteria, Rickettsiales) as a sister group of the genus Rickettsia. In situ observations revealed the ability of the candidate new species to colonize either nuclear or cytoplasmic compartments, depending on the host organism. The presence of the same bacterial species within different, evolutionary distant, hosts indicates that 'Candidatus Megaira polyxenophila' recently underwent several distinct host shifts, thus suggesting the existence of horizontal transmission pathways. We consider these findings as indicative of an unexpected spread of rickettsial infections in aquatic communities, possibly by means of trophic interactions, and hence propose a new interpretation of the origin and phylogenetic diversification of rickettsial bacteria.

Morphology, ultrastructure, and molecular phylogeny of the ciliate Sonderia vorax with insights into the systematics of order Plagiopylida.

BACKGROUND: Ciliates of the family Sonderiidae are common members of the eukaryotic communities in various anoxic environments. They host both ecto- and endosymbiotic prokaryotes (the latter associated with hydrogenosomes) and possess peculiar morpho-ultrastructural features, whose functions and homologies are not known. Their phylogenetic relationships with other ciliates are not completely resolved and the available literature, especially concerning electron microscopy and molecular studies, is quite scarce. RESULTS: Sonderia vorax Kahl, 1928 is redescribed from an oxygen-deficient, brackish-water pond along the Ligurian Sea coastlines of Italy. Data on morphology, morphometry, and ultrastructure are reported. S. vorax is ovoid-ellipsoid in shape, dorsoventrally flattened, 130 x 69 µm (mean in vivo); it shows an almost spherical macronucleus, and one relatively large micronucleus. The ventral kinetom has a "secant system" including fronto-ventral and fronto-lateral kineties. A distinctive layer of bacteria laying between kineties covers the ciliate surface. Two types of extrusomes and hydrogenosomes-endosymbiotic bacteria assemblages are present in the cytoplasm. The phylogeny based on 18S rRNA gene sequences places S. vorax among Plagiopylida; Sonderiidae clusters with Plagiopylidae, although lower-level relationships remain uncertain. The studied population is fixed as neotype and the ciliate is established as type species of the genus, currently lacking. CONCLUSIONS: This is the first description of a representative of Sonderiidae performed with both morphological and molecular data. To sum up, many previous hypotheses on this interesting, poorly known taxon are confirmed but confusion and contradictory data are as well highlighted.

Characterization and comparison of bacterial communities selected in conventional activated sludge and membrane bioreactor pilot plants: a focus on Nitrospira and Planctomycetes bacterial phyla.

A pilot-scale membrane bioreactor (MBR) and a conventional activated sludge system (CAS) were in parallel operated to investigate the impact of the separation technology on the structure and functionality of the selected microbial community. Microbial communities as well as nitrogen removal efficiency of the biomass were characterized. Kinetics and microbial community structure turned out to be duly correlated. The impact of the separation technology on selective conditions and, in particular, the higher variability of solid separation efficiency in CAS with respect to MBR pilot plant possibly represented the main factor influencing the selection of bacterial communities. Concerning nitrifiers, bacteria of the genus Nitrospira were predominant in the MBR. This was in accordance with kinetics of nitrite-oxidizing bacteria that suggested the presence of k-strategists, while r-strategists were selected in the CAS plant, possibly because of the presence of transient higher concentrations of nitrite (in the range of 0.05-0.18 and of 0.05-4.4 mg [Formula: see text]-N L(-1) in the MBR and CAS effluents, respectively). An unexpectedly high presence of bacteria belonging to two specific phylogenetic clades of Planctomycetes was found in both reactors.

"Candidatus Defluviella procrastinata" and "Candidatus Cyrtobacter zanobii", two novel ciliate endosymbionts belonging to the "Midichloria clade".

The "Midichloria clade" is a recently discovered but well-established evolutionary lineage clustering inside the order Rickettsiales (Alphaproteobacteria). Not much is known about the biology of these organisms. The best characterized ones are endocellular symbionts of very different eukaryotic hosts, ranging from arthropods to protists. "Candidatus Midichloria mitochondrii", the most studied organism of the group, is an interesting object of study because of its unique capability to infect metazoans' mitochondria and the presence of flagellar genes in its genome. With this work, we aim at increasing the knowledge on the biodiversity and phylogeny of the "Midichloria group". We characterized according to the "full cycle rRNA approach" two novel endosymbionts of ciliated protozoa, i.e. Paramecium nephridiatum and Euplotes aediculatus. According to the nomenclatural rules for uncultivated prokaryotes, we established the novel taxa "Candidatus Defluviella procrastinata" and "Candidatus Cyrtobacter zanobii" for the two bacterial symbionts. Our phylogenetic analysis based on 16S rRNA gene sequences confirms that the evolutionary histories of "Midichloria clade" representatives and of their hosts are very different. This suggests that the symbiotic processes arose many times independently, perhaps through ways of transmission still not described in Rickettsiales.

Morphological, ultrastructural, and molecular characterization of Euplotidium rosati n. sp. (Ciliophora, Euplotida) from Guam.

We combined morphological (i.e. live, stained, scanning, and transmission electron microscopy) with morphometric and molecular analysis to describe a ciliate species collected from shallow reefs in Guam, grown, and maintained in our laboratory. The species was recognized as a member of Euplotidium, and compared with established species of the genus: Euplotidium itoi Ito 1958; Euplotidium psammophilus (Vacelet 1961) Borror 1972; Euplotidium arenarium Magagnini and Nobili 1964; Euplotidium helgae Hartwig 1980; Euplotidium prosaltans Tuffrau 1985, and Euplotidium smalli Lei, Choi and Xu, 2002. To obtain more elements to compare the species, new morphometric data and additional SSU rRNA gene sequences of E. itoi and of E. arenarium are reported. On the basis of this comparison, we established the new species Euplotidium rosati that has a cirral pattern composed of 12 frontoventral and six transverse cirri, and lacks the left marginal cirrus. Euplotidium rosati harbors on its dorsal surface epixenosomes, the peculiar extrusive symbionts described in other Euplotidium species. The whole body of our observations together with the analysis of the data available in the literature leads us to propose a redefinition of the genus. The results may also be useful to clarify the tangled relationship between Euplotidium and Gastrocirrhus.