Search

The phylum Ciliophora is one of the most broadly studied protozoan lineages. The era of molecular investigation has brought forth a major ongoing debate: is the subclass Peritrichia Stein, 1859 monophyletic? Numerous analyses mostly using the small subunit (SSU) rRNA gene have failed to recover the Mobilida and Sessilida, the two peritrich orders, as sister clades. Here we have sequenced five peritrich species - three sessilids and two mobilids. We constructed a supermatrix of 158 genes and 44,696 characters for 24 ciliate species, and as outgroup taxa, nine species from the Apicomplexa and four from the Dinophyceae. Our analyses using both maximum likelihood and Bayesian methods recover a monophyletic class Oligohymenophorea and two robust clades within it. The first clade is a monophyletic Peritrichia with the orders Sessilida and Mobilida maximally supported as sister clades. The second oligohymenophorean clade includes species of the subclasses Scuticociliatia and Hymenostomatia, which are sister clades. Our analyses resolve a long-standing debate in ciliate molecular phylogenetics and provide support for the classical view that the morphological features of the two peritrich orders Mobilida and Sessilida arose by descent from the same common ancestor and are not the result of convergence.

Background Persons with schizophrenia often come in contact with the criminal justice system (CJS). This analysis of subjects with schizophrenia and a history of contact with the CJS estimated and compared mean cumulative function (MCF) of treatment failure events when treated with paliperidone palmitate (PP) or oral antipsychotics (OAs). All events identified during the full study period of the Paliperidone Palmitate Research in Demonstrating Effectiveness (PRIDE) trial were evaluated. Methods Subjects were randomly assigned to flexibly dosed, monthly, injectable PP (78–234 mg) or daily OA in a 15-month prospective, open-label, multicenter US study (May 5, 2010–December 9, 2013). Subjects could continue participation after a treatment failure event. Multiple treatment failures in individual subjects were analyzed as recurrent events. Analyses estimated MCF of treatment failure events and MCF of institutionalizations (arrests, incarcerations, or psychiatric hospitalizations) during the 15-month study period. Results The ITT population included 226 (PP) and 218 (OA) subjects, of whom 41.2% and 40.4%, respectively, completed 15 months of follow-up. The MCF of treatment failures and institutionalizations differed significantly in favor of PP compared with OA (P = 0.007 and P = 0.005, respectively). Overall, TEAEs were reported by 86.3% of subjects in the PP group and 81.7% in the OA group. Conclusions This pragmatic analysis suggests that, compared with OA, PP is not only more effective in delaying median time to treatment failure, but it also reduces the number of treatment failures and institutionalizations per person-year follow-up. Clinical trials registration: Clinicaltrials.gov identifier: NCT01157351

DNA replication is a ubiquitous and conserved cellular process. However, regulation of DNA replication is only understood in a small fraction of organisms that poorly represent the diversity of genetic systems in nature. Here we used computational and experimental approaches to examine the function and evolution of one such system, the replication band (RB) in spirotrich ciliates, which is a localized, motile hub that traverses the macronucleus while replicating DNA. We show that the RB can take unique forms in different species, from polar bands to a "replication envelope," where replication initiates at the nuclear periphery before advancing inward. Furthermore, we identify genes involved in cellular transport, including calcium transporters and cytoskeletal regulators, that are associated with the RB and may be involved in its function and translocation. These findings highlight the evolution and diversity of DNA replication systems and provide insights into the regulation of nuclear organization and processes.

We report here the draft genome sequence of a strain of Tenacibaculum discolor (Bacteroidetes) that was isolated from the river-ocean interface at Trunk River in Falmouth, Massachusetts. The isolation and genomic sequencing were performed during the 2016 and 2018 Microbial Diversity summer programs at the Marine Biological Laboratory in Woods Hole, Massachusetts.

116 pages, 1 figure, 4 tables, 3 appendices, This revision of the classification of eukaryotes follows that of Adl et al., 2012 [J. Euk. Microbiol. 59(5)] and retains an emphasis on protists. Changes since have improved the resolution of many nodes in phylogenetic analyses. For some clades even families are being clearly resolved. As we had predicted, environmental sampling in the intervening years has massively increased the genetic information at hand. Consequently, we have discovered novel clades, exciting new genera and uncovered a massive species level diversity beyond the morphological species descriptions. Several clades known from environmental samples only have now found their home. Sampling soils, deeper marine waters and the deep sea will continue to fill us with surprises. The main changes in this revision are the confirmation that eukaryotes form at least two domains, the loss of monophyly in the Excavata, robust support for the Haptista and Cryptista. We provide suggested primer sets for DNA sequences from environmental samples that are effective for each clade. We have provided a guide to trophic functional guilds in an appendix, to facilitate the interpretation of environmental samples, and a standardized taxonomic guide for East Asian users, Research support was provided as follows: SMA by NSERC 249889‐2007; DB by NERC NE/H009426/1 and NE/H000887/1; MWB by NSF 1456054; FB by a Fellowship from Science for Life Laboratory and VR/2017‐04563; PC by EU‐Horizon 2020 research and innovation program through the SponGES project 679849 [...]; IC by CSF 18‐18699S; BE by RCN TaxMArc 268286/GMR; LG by ANR HAPAR (ANR‐14‐CE02‐0007); VH MK JL by ERDF; MEYS with ERC 771592 CZ 1.05/1.1.00/02.0109 BIOCEV; SK by RSF 16‐14‐10302; MK by CSF GA18‐28103S; CEL by NSF 1541510 and NIH‐AI124092; EL by CAM: 2017‐T1/AMB‐5210; and by grant 2017‐T1/AMB‐5210 from the program >Atracción de talentos> from the Consejería de Educación, Juventud y Deporte, Comunidad de Madrid; JL by ERC CZ LL1601 and OPVVV 16_019/0000759; MP by NSF DEB‐1455611; DJR by the Beatriu de Pinós postdoctoral programme of the Government of Catalonia's Secretariat for Universities and Research of the Ministry of Economy and Knowledge; CLS by the intramural research program of the National Library of Medicine, National Institutes of Health; AS by RSF 17‐14‐01391 and RFBR 16‐04‐01454 NY by NSF DEB 1557102; VZ by RFBR 16‐34‐60102 mol‐a‐dk; UniEuk and EukRef by the Gordon and Betty Moore Foundation

We report here the draft genome sequence of a strain of Tenacibaculum discolor (Bacteroidetes) that was isolated from the river-ocean interface at Trunk River in Falmouth, Massachusetts. The isolation and genomic sequencing were performed during the 2016 and 2018 Microbial Diversity summer programs at the Marine Biological Laboratory in Woods Hole, Massachusetts.

Traditional approaches (e.g., neurobehavior, neuropathology) can detect alterations in apical endpoints indicative of developmental neurotoxicity (DNT). However, there is an increasing desire to understand mode-of-action (MOA) for DNT effects; thus, this short communication describes initial work on a neuronal differentiation assay. Basically, our laboratory used the human NT2/D1 cell line to develop an assay to evaluate toxicants for effects on all-trans retinoic acid (RA)-induced neuronal differentiation. Based on literature reports, we selected a neuronal protein, neuronal class III β-tubulin (β3-tubulin), as a marker of differentiation. For this assay, cultured RA-treated NT2 cells were trypsinized to individual cells, methanol fixed, and labeled with a β3-tubulin specific monoclonal antibody (TUJ1). Characterization studies using 100,000 cells/sample showed that NT2 cells had appreciable expression of β3-tubulin starting around day 7 of the differentiation process with a peak expression noted around day 12. Methylmercury, 22(R)-hydroxycholesterol, N-(4-hydroxyphenol)retinamide (4HPR), and 9-cis retinoic acid were selected as initial test compounds. Of these, only 9-cis RA, which is known to affect the RA pathway, was positive for specific impacts on differentiation. These results demonstrate the feasibility of using a flow cytometry method targeting specific cellular biomarkers for evaluating effects on neuronal differentiation. Additional assays are needed to detect compounds targeting other (non-RA) neuronal differentiation pathways. Ultimately, a battery of in vitro assays would be needed to evaluate the potential MOAs involved in altered neuronal differentiation.

Undergraduates learn that gene editing in diverse organisms is now possible. How targeted manipulation of genes and genomes is utilized in basic science and biomedicine to address biological questions is challenging for undergraduates to conceptualize. Thus, we developed a lab experience that would allow students to be actively engaged in the full process of design, implementation of a gene editing strategy, and interpretation of results within an 8-week lab period of a Genetics course. The laboratory experience combines two transformative biotechnology tools; the utilization of green fluorescent protein (GFP) as a diagnostic marker of gene expression and the fundamentals and specificity of Clustered Regularly Interspaced Short Palindromic Repeats-cas9 (CRISPR-cas9) gene editing in bacterial cells. The students designed and constructed plasmids that express single guide RNA targeted to GFP, expressed the sgRNA and cas9 in bacteria cells, and successfully deactivated GFP gene expression in the bacterial cells with their designed CRISPR-cas9 tools. Student assessment revealed most students achieved student learning objectives. We conclude this lab experience is an effective and accessible method for engaging students in the scientific practices, knowledge and challenges revolving targeted CRISPR-cas9 gene manipulation. © 2019 International Union of Biochemistry and Molecular Biology, 47(2): 145-155, 2019.

This study investigated the effect of a previously uncharacterized species of ciliated protozoan, Tetrahymena glochidiophila, on the viability of glochidia from three species of freshwater mussel (Lampsilis siliquoidea, Lampsilis fasciola, and Lampsilis cardium). Over the course of 72 h, the viability of glochidia exposed to T. glochidiophila declined by60% while the decline in the viability of uninfected glochidia was10%. The density of T. glochidiophila increased1000-fold during the experiment in treatments with infected glochidia. Lampsilis cardium glochidia were also either exposed to gill rinsate or gill contents from infected gravid female L. siliquoidea for the purpose of elucidating the location of the greatest density of ciliates within infected mussels. Glochidia exposed to gill contents declined significantly (p 0.05) more than glochidia exposed to gill rinsate. Finally, a clone of ciliates was isolated from infected glochidia and cultured on bacterized medium. The clonal culture was then used to expose uninfected glochidia for the purpose of further confirming a parasitic relationship between glochidia and T. glochidiophila. The viability of glochidia exposed to T. glochidiophila from the clonal culture declined significantly relative to uninfected glochidia but not to the extent of glochidia exposed to ciliates from the gills of infected L. siliquoidea.

A ciliate protozoan was discovered whose presence coincided with a rapid decrease in the viability (i.e. ability to close valves) of glochidia of the freshwater mussel Lampsilis siliquoidea. Microscopic examination showed it to be a histophagous tetrahymenine ciliate. Small subunit (SSU) rRNA and cytochrome c oxidase subunit 1 (cox1) barcode sequences from cultured cells showed that it belongs to the same new species isolated from water samples as a free-living ciliate. Phylogenetic analyses place this new ciliate in the same clade with the macrostome species Tetrahymena paravorax, and we propose the name T. glochidiophila n. sp. for this new species. The phylogeny provides further support for the hypothesis that histophagy was a life history trait of the ancestor of Tetrahymena.

This case is based on an actual inventory accounting dilemma for a fish processing company located in Washington State. Rex Seafood, Incorporated (Rex) and its wholly owned subsidiary, Juneau Supply Company, purchase, process, and sell fish to restaurants and other retail outlets in the state of Washington and surrounding inland states. Rex has an agreement with Condor Enterprises to transport fish purchased from Juneau Supply in Alaska to Rex's facility in Washington for processing. The central issue of the case involves appropriate accounting and reporting for intercompany sales: How should the sales of inventory from Juneau Supply to Condor and the purchases by Rex from Condor be recorded and reported? Using the FASB Accounting Standards Codification, students must determine whether Rex's arrangement with Condor is a product financing arrangement, whether the purchases and sales of inventory should be considered a single exchange transaction, and whether the events should be considered intercompany transactions. In addition, students are asked to consider recording and disclosure implications of recommended solutions. This case is suitable for use in undergraduate Intermediate and graduate Financial Accounting classes.

Background/ContextIn response to a shortage of highly qualified teachers, states and school districts have enacted a number of policy innovations over the last two decades. While researchers have evaluated many of these reforms, little is known about the initial effectiveness of individuals with prior teaching assistant experience. We review existing literature related to teaching assistants and utilize administrative data to examine a number of characteristics of individuals who transition from work as teaching assistants to jobs as regular classroom teachers.Study PurposeIndividuals who become classroom teachers after having prior experience in schools working as teaching assistants may perform differently than other teachers due to additional exposure to classroom practices. The purpose of this study is to identify differences in the characteristics, effectiveness, and persistence of teachers who work as teaching assistants prior to entering the classroom as regular classroom teachers.PopulationWhile the population of focus varies somewhat across research questions, we generally focus on teachers with less than five years of experience working in elementary and middle school grades in all North Carolina public schools from 2005–2006 to 2009–2010. Additional data on teaching assistants employed from 2000–2001 to 2009–2010 in North Carolina public schools allows identification of individuals who had prior experience as teaching assistants in the state.Research DesignUsing administrative data records, we utilize quantitative descriptions of teacher characteristics and persistence to identify differences between teachers who entered the classroom with prior teaching assistant experience and other groups of teachers. For effectiveness comparisons, we estimate value-added models to compare the effectiveness of teachers in the identified groups of interest.Findings/ResultsOverall, we find that teachers who began as teaching assistants are (1) older, more racially diverse, lower-scoring on licensure exams, and more likely to enter the profession alternatively; (2) more effective in elementary-grades math and reading; and (3) more likely to remain as classroom teachers in North Carolina public schools.Conclusions/RecommendationsThese findings indicate that former teaching assistants represent a quality and highly persistent labor source and call for continued research to understand how additional exposure to classrooms benefits early-career teacher performance.

Aim Long-acting injectable antipsychotics (APs) are not well studied in recent-onset schizophrenia. This exploratory analysis of a study designed to reflect real-world schizophrenia, as defined by patients, interventions and outcomes, compared relative treatment effect between once-monthly paliperidone palmitate (PP) and daily oral APs in patients with recent-onset or chronic illness Methods This randomized, open-label, event monitoring board–blinded study compared treatment response in subjects with schizophrenia and a history of criminal justice system involvement following treatment with PP or oral APs for 15 months (ClinicalTrials.gov identifier, NCT01157351). Event-free probabilities were estimated using Kaplan–Meier method; hazard ratios (HRs) were estimated using Cox proportional hazard models. This subgroup analysis analysed data by disease duration (≤5 (recent-onset) or >5 years (chronic illness) since first psychiatric diagnosis). Results Seventy-seven subjects met the criteria for recent-onset illness; 365 for chronic illness. HRs (95% CI) for treatment failure for oral APs versus PP were 1.73 (0.87–3.45; P = 0.121) for recent-onset and 1.37 (1.02–1.85; P = 0.039) for chronic illness. Most common adverse events for PP versus oral APs were injection site pain (recent-onset, 26% vs. 0%; chronic, 17% vs. 0%), increased weight (14% vs. 6%; 12% vs. 6%), akathisia (14% vs. 9%; 10% vs. 7%), insomnia (12% vs. 17%; 18% vs. 10%) and anxiety (12% vs. 6%; 10% vs. 8%). Conclusions Although neither pre-planned nor adequately powered, the estimated HRs suggest that the relative advantage of PP over oral APs for reducing the risk for treatment failure may be greater in patients with recent-onset schizophrenia than in those with more chronic illness.

Aim Long‐acting injectable antipsychotics (APs) are not well studied in recent‐onset schizophrenia. This exploratory analysis of a study designed to reflect real‐world schizophrenia, as defined by patients, interventions and outcomes, compared relative treatment effect between once‐monthly paliperidone palmitate (PP) and daily oral APs in patients with recent‐onset or chronic illness Methods This randomized, open‐label, event monitoring board–blinded study compared treatment response in subjects with schizophrenia and a history of criminal justice system involvement following treatment with PP or oral APs for 15 months (ClinicalTrials.gov identifier, NCT01157351). Event‐free probabilities were estimated using Kaplan–Meier method; hazard ratios (HRs) were estimated using Cox proportional hazard models. This subgroup analysis analysed data by disease duration (≤5 (recent‐onset) or >5 years (chronic illness) since first psychiatric diagnosis). Results Seventy‐seven subjects met the criteria for recent‐onset illness; 365 for chronic illness. HRs (95% CI) for treatment failure for oral APs versus PP were 1.73 (0.87–3.45; P = 0.121) for recent‐onset and 1.37 (1.02–1.85; P = 0.039) for chronic illness. Most common adverse events for PP versus oral APs were injection site pain (recent‐onset, 26% vs. 0%; chronic, 17% vs. 0%), increased weight (14% vs. 6%; 12% vs. 6%), akathisia (14% vs. 9%; 10% vs. 7%), insomnia (12% vs. 17%; 18% vs. 10%) and anxiety (12% vs. 6%; 10% vs. 8%). Conclusions Although neither pre‐planned nor adequately powered, the estimated HRs suggest that the relative advantage of PP over oral APs for reducing the risk for treatment failure may be greater in patients with recent‐onset schizophrenia than in those with more chronic illness.

A food blog and a catering business provide an issue-rich context in which students examine tax issues involving individuals, a partnership, and a single owner LLC. The case illustrates how advances in technology and social media occur without the issuance of directly applicable tax guidance, thus introducing students to uncertainty in interpreting the tax law and providing professional advice. The goals of the case are to increase students' (1) abilities to identify tax issues for individuals and small home-based businesses, (2) tax research proficiency, (3) written communication skills, (4) technical knowledge, and (5) experience in tax planning. The case helps students develop four of the core competencies expressed in the AICPA's (2012) CPA Vision Project, including improving students' abilities to interpret converging information, think strategically and critically, communicate effectively, and adeptly use technology. Student responses to a posttest questionnaire indicate that the case helped develop their issue identification, tax research, and writing skills.

OBJECTIVE The Paliperidone Palmitate Research in Demonstrating Effectiveness (PRIDE) study compared the effects of once-monthly paliperidone palmitate with daily oral antipsychotics on treatment failure in adults with schizophrenia. METHOD The PRIDE study is a 15-month, randomized, multicenter study (May 5, 2010, to December 9, 2013) of adult subjects with a DSM-IV diagnosis of schizophrenia and a history of incarceration. Subjects were randomly assigned to once-monthly paliperidone palmitate injections or daily oral antipsychotics (randomly assigned from 7 acceptable, prespecified oral antipsychotics) for 15 months. The primary end point was time to first treatment failure, defined as arrest/incarceration; psychiatric hospitalization; suicide; treatment discontinuation or supplementation due to inadequate efficacy, safety, or tolerability; or increased psychiatric services to prevent hospitalization. Time to first treatment failure was determined by a blinded event-monitoring board and analyzed with the Kaplan-Meier method. RESULTS In this study, 450 patients were randomly assigned, and 444 were included in the intent-to-treat population. Paliperidone palmitate was associated with significant delay in time to first treatment failure versus oral antipsychotics (hazard ratio, 1.43; 95% CI, 1.09-1.88; log rank P = .011). Observed treatment failure rates over 15 months were 39.8% and 53.7%, respectively. Arrest/incarceration and psychiatric hospitalization were the most common reasons for treatment failure in the paliperidone palmitate and oral antipsychotic groups (21.2% vs 29.4% and 8.0% vs 11.9%, respectively). The 5 most common treatment-emergent adverse events for the paliperidone palmitate treatment group were injection site pain (18.6% of subjects), insomnia (16.8%), weight increased (11.9%), akathisia (11.1%), and anxiety (10.6%). CONCLUSIONS In a trial designed to reflect real-world management of schizophrenia, once-monthly paliperidone palmitate demonstrated superiority compared to oral antipsychotics in delaying time to treatment failure. TRIAL REGISTRATION Clinicaltrials.gov Identifier: NCT01157351.

The phylogenetic relationships of the ciliate subclass Peritrichia, composed of the orders Mobilida and Sessilida, have recently come under debate as morphological and molecular analyses have struck contrasting conclusions as to the monophyly of the group. We provide additional molecular data to assess the monophyly of the Peritrichia by sequencing the small subunit ribosomal RNA genes of two symbiotic peritrichs, Urceolaria korschelti and Scyphidia ubiquita, found inhabiting the mantle cavity of limpets. Although phylogenetic analyses indicated a nonmonophyletic Peritrichia, approximately unbiased tests revealed that the monophyletic hypothesis could not be rejected. With regard to the Mobilida, our analysis showed divergence within the family Trichodinidae related to host taxa-a molluscan clade and a fish clade. For the Sessilida, the family Scyphidiidae was sister to the Astylozoidae. In our sampling of U. korschelti and S. ubiquita, both species showed significant genetic divergence among geographically isolated, yet morphologically indistinguishable populations. We hypothesize that cryptic speciation has produced these morphologically identical species and argue that more extensive genomic analyses are required to fully assess the monophyly, biogeography, and ultimately biodiversity of the peritrichs.

Background Observational studies of switching from branded to generic formulations of the same drug substance often lack appropriate comparators for the subjects who switched. Three generic formulations were deemed equivalent to Concerta: an authorized generic (AG) identical except for external packaging, and two other generics (EG). Objective Compare the incidence of a combined endpoint (switching back to Concerta, changing the use of immediate release methylphenidate (MPH), stopping all long-acting methylphenidate, or starting a new medication) among people switched from Concerta to the AG versus the EG. Methods Cohort study from the Truven CCAE database of people aged 6 to 65 diagnosed with ADHD, treated with Concerta, and switched to the EG or to the AG formulation. Results In the EG arm 24.6% and in the AG arm 19.7% of subjects switched back to Concerta. The proportion of subjects meeting the combined endpoint was 39.5% in the EG arm, 32.9% in the AG arm, a crude risk ratio of 1.20 (95% CI 0.94, 1.54). After adjustment by propensity score stratification, the adjusted odds ratio (OR) was 1.23 (95% CI 0.90, 1.70). In an unplanned analysis using a different method of adjustment, the adjusted OR was 1.00 (95% CI 0.69, 1.44). Discussion This study did not detect a difference between the proportion of people who met the study endpoint in the two study arms, i.e. between those who switched to a generic formulation that was identical to Concerta except for external packaging and those who switched to the comparison generics. The high incidence of the combined endpoint in the AG arm demonstrates the need for an appropriate comparator in studies of this type. Trial registration ClinicalTrials.gov NCT02730572

The class Nassophorea includes the microthoracids and nassulids, which share morphological similarities in their somatic kinetids and cytopharyngeal baskets. The monophyly of this clade has been challenged by small subunit rRNA gene sequences and multi-gene analyses that do not provide strong support. To provide a more robust test of the monophyly of the Nassophorea, phylogenomic analyses were based on 124 genes derived from the single-cell transcriptomes of the microthoracid Pseudomicrothorax dubius and the nassulid Furgasonia blochmanni. The nassulid Nassula sorex from the Culture Centre for Algae and Protozoa was also included, but this isolate was discovered to have been misidentified. We first redescribe, using light and scanning electron microscopical techniques, this "N. sorex" as a new species of Nassula, Nassula variabilis n. sp., characterized by its highly variable nassulid frange. We have sequenced the single-cell transcriptomes to obtain data for phylogenomic analyses. These gave robust support for the Nassophorea, which are sister to a clade of Colpodea species. If our topology truly represents the order of divergence of taxa, a cytopharyngeal basket with microtubular nematodesmata and with Y and Z microtubular ribbons was likely an ancestral feature, at least of the Phyllopharyngea, Colpodea, Nassophorea, and Oligohymenophorea.

SummaryIn the endocytic pathway of animals, two related complexes, called CORVET (Class C Core Vacuole/Endosome Transport) and HOPS (Homotypic fusion and protein sorting), act as both tethers and fusion factors for early and late endosomes, respectively. Mutations in CORVET or HOPS lead to trafficking defects and contribute to human disease including immune dysfunction. HOPS and CORVET are conserved throughout eukaryotes but remarkably, in the ciliateTetrahymena thermophila,the HOPS-specific subunits are absent while CORVET-specific subunits have proliferated. VPS8 (Vacuolar Protein Sorting), a CORVET subunit, expanded to 6 paralogs inTetrahymena. This expansion correlated with loss of HOPS within a ciliate subgroup including the Oligohymenophorea, which containsTetrahymena. As uncovered via forward genetics, a singleVPS8paralog inTetrahymena(VPS8A) is required to synthesize prominent secretory granules called mucocysts. More specifically,∆vps8acells fail to deliver a subset of cargo proteins to developing mucocysts, instead accumulating that cargo in vesicles also bearing the mucocyst sorting receptor, Sor4p. Surprisingly, although this transport step relies on CORVET, it does not appear to involve early endosomes. Instead,Vps8aassociates with the late endosomal/lysosomal marker Rab7, indicating target specificity switching occurred in CORVET subunits during the evolution of ciliates. Mucocysts belong to a markedly diverse and understudied class of protist secretory organelles called extrusomes. Our results underscore that biogenesis of mucocysts depends on endolysosomal trafficking, revealing parallels with invasive organelles in apicomplexan parasites and suggesting that a wide array of secretory adaptations in protists, like in animals, depend on mechanisms related to lysosome biogenesis.AbbreviationsLRO(Lysosome-related organelle)HOPS(homotypic fusion and protein sorting complex)CORVET(Class C core Vacuole/Endosome Transport)VPS(vacuolar protein sorting)GRL(granule lattice)GRT(granule tip)Igr(Induced upon granule regeneration)SNARE(Soluble NSF attachment protein receptor)LECA(last eukaryotic common ancestor)

International audience; Microscopic and phylogenetic analyses were performed on endocommensal astome ciliates retrieved from the middle intestine of a marine cirratulid polychaete, Cirriformia tentaculata, collected in the bay of Roscoff (English Channel, Northwest French coast) and on the Southwest English coast. Three morphotypes of the astome genus Durchoniella were identified, two corresponding to described species (the type species Durchoniella brasili (Léger and Duboscq, 1904) De Puytorac, 1954 and Durchoniella legeriduboscqui De Puytorac, 1954) while a third morphotype remains undescribed. Their small subunit (SSU) rRNA gene sequences showed at least 97.2% identity and phylogenetic analyses grouped them at the base of the subclass Scuticociliatia (Oligohymenophorea), as a sister lineage to all astomes from terrestrial oligochaete annelids. Ultrastructural examination by transmission electron microscopy and fluorescence in situ hybridization analyses revealed the presence of endocytoplasmic cocci and rod-shaped bacteria surrounded by a very thin membrane. These endocytoplasmic bacteria may play a role in the association between endocommensal astome ciliates and cirratulid polychaetes inhabiting in anoxic coastal sediments.

Summary In the endocytic pathway of animals, two related complexes, called CORVET (class C core vacuole/endosome transport) and HOPS (homotypic fusion and protein sorting), act as both tethers and fusion factors for early and late endosomes, respectively. Mutations in CORVET or HOPS lead to trafficking defects and contribute to human disease, including immune dysfunction. HOPS and CORVET are conserved throughout eukaryotes, but remarkably, in the ciliate Tetrahymena thermophila , the HOPS-specific subunits are absent, while CORVET-specific subunits have proliferated. VPS8 (vacuolar protein sorting), a CORVET subunit, expanded to 6 paralogs in Tetrahymena . This expansion correlated with loss of HOPS within a ciliate subgroup, including the Oligohymenophorea, which contains Tetrahymena . As uncovered via forward genetics, a single VPS8 paralog in Tetrahymena ( VPS8A ) is required to synthesize prominent secretory granules called mucocysts. More specifically, Δvps8a cells fail to deliver a subset of cargo proteins to developing mucocysts, instead accumulating that cargo in vesicles also bearing the mucocyst-sorting receptor Sor4p. Surprisingly, although this transport step relies on CORVET, it does not appear to involve early endosomes. Instead, Vps8a associates with the late endosomal/lysosomal marker Rab7, indicating that target specificity switching occurred in CORVET subunits during the evolution of ciliates. Mucocysts belong to a markedly diverse and understudied class of protist secretory organelles called extrusomes. Our results underscore that biogenesis of mucocysts depends on endolysosomal trafficking, revealing parallels with invasive organelles in apicomplexan parasites and suggesting that a wide array of secretory adaptations in protists, like in animals, depend on mechanisms related to lysosome biogenesis.

Experiments were conducted on an aqueous growth medium containing cultures of Escherichia coli ( E. coli ) XL1-Blue, to investigate, in a single experiment, the effect of two types of dynamic mechanical loading on cellular integrity. A bespoke shock tube was used to subject separate portions of a planktonic bacterial culture to two different loading sequences: (i) shock compression followed by cavitation, and (ii) shock compression followed by spray. The apparatus allows the generation of an adjustable loading shock wave of magnitude up to 300 MPa in a sterile laboratory environment. Cultures of E. coli were tested with this apparatus and the spread-plate technique was used to measure the survivability after mechanical loading. The loading sequence (ii) gave higher mortality than (i), suggesting that the bacteria are more vulnerable to shear deformation and cavitation than to hydrostatic compression. We present the results of preliminary experiments and suggestions for further experimental work; we discuss the potential applications of this technique to sterilize large volumes of fluid samples.

Ciliates within the order Mobilida comprise a group of ectosymbionts that form ecologically and economically important symbioses with aquatic organisms. However, despite their significance, the factors driving mobilid diversification are poorly understood. To address this, we sought to examine the genetic diversity of two mobilid families, the trichodinids and urceolariids, over geographical areas and from different hosts. To this end we collected and provide morphological and molecular characterizations of three species of Trichodina: Trichodina domerguei and Trichodina tenuidens from three-spined sticklebacks, and Trichodina unionis from snails. We also sequenced the small subunit rRNA gene from Urceolaria korschelti collected from its type host at the type locality of Helgoland, Germany. The Trichodina from sticklebacks show intra-specific genetic variation that is not related to geographic distribution. Furthermore, we find evidence of fish to mollusc host transitions within the Trichodinidae and suggest that the most recent common ancestor of the Trichodina was a symbiont of vertebrates. Lastly we confirm that U. korschelti constitutes a cryptic species complex, which in turn justifies the description of Urceolaria parakorschelti sp. nov. These data suggest that not only host transitions, but allopatric and or genetically-driven sympatric speciation play a role in facilitating the diversification of mobilid ciliates.

Background Important work has been carried out adapting cognitive behavioural therapy for people with intellectual disabilities. However, there is a lack of alternative psychological therapies available for people with intellectual disabilities and emotional difficulties. Behavioural activation for depression is less reliant on verbal communication and focuses on increasing purposeful activity and reducing avoidance. Method This feasibility study involved the development and piloting of an adapted manual of behavioural activation for people with intellectual disabilities. The intervention consisted of 10–12 sessions and a key adaptation was that the therapist worked with the clients alongside a significant other in their life, either a paid carer or family member. Baseline, post-intervention (3 months after entering the study) and 6-month quantitative follow-up data were obtained. Primary outcome data were gathered, concerning depressive symptoms, participants' levels of activity and general well-being. Results Twenty-three adults with intellectual disabilities with symptoms of depression were recruited from specialist health services. In terms of acceptability, the behavioural activation intervention was well received and only two individuals dropped out, with a further two lost to follow-up. The main measures of depression appeared to be sensitive to change. Pre- to post-intervention data showed a significant reduction in self-report of depressive symptoms with a strong effect size (r = 0.78), that was maintained at follow-up (r = 0.86). Positive change was also obtained for informant reports of depressive symptoms from pre- to post-intervention, with a strong effect size (r = 0.7). Once again, this positive change was maintained at follow-up (r = 0.72). Conclusions The study suggested that behavioural activation may be a feasible and worthwhile approach to tackling depression in people with intellectual disabilities. However, a randomised controlled trial would be required to establish its effectiveness, with more sensitive measurement of change in activity.

All parasitoid apostome ciliates infecting krill in the northeastern Pacific are currently assigned to the genus Pseudocollinia. Each krill specimen is apparently infected by only 1 Pseudocollinia species. We describe Pseudocollinia similis sp. nov., discovered infecting the krill Thysanoessa spinifera off Oregon, USA. Its protomite-tomite stage resembles that of P. beringensis, which infects T. inermis (type host species), T. longipes, and T. raschii females in the Bering Sea. These ciliates have similar numbers of somatic kineties (18-21 vs. 16-20) and typically have 3 oral kineties. Furthermore, these 2 apostomes are sister species on gene trees based on sequences of small subunit rRNA (0.06% difference) and cytochrome c oxidase subunit 1 (cox1; 30% difference). P. brintoni and P. oregonensis are closely related as a separate group from P. similis and P. beringensis. The similar tree topologies based on the cox1 sequences of 21 host krill individuals representing 6 krill species (Euphausia pacifica, Nyctiphanes simplex, T. inermis, T. longipes, T. raschii, and T. spinifera) and the apostomes isolated from these krill suggest host-parasitoid codiversification. However, this hypothesis was statistically rejected by an approximately unbiased test in which the host tree topology was used to model parasitoid evolution (p ≤ 0.05).

As a cellular organelle, the cilium contains a unique protein composition [1, 2]. Entry of both membrane [3–5] and cytosolic components [6–8] is tightly regulated by gating mechanisms at the cilium base, however, the mechanistic details of ciliary gating are largely unknown. We previously proposed that entry of cytosolic components is regulated by mechanisms similar to those of nuclear transport and is dependent on nucleoporins (NUPs) which comprise a ciliary pore complex (CPC) [6, 9]. To investigate ciliary gating mechanisms, we developed a system to clog the pore by inhibiting NUP function via forced dimerization. We targeted NUP62, a component of the central channel of the nuclear pore complex (NPC) [10], for forced dimerization by tagging it with the homodimerizing Fv domain. As proof of principle, we show that forced dimerization of NUP62-Fv attenuated active transport of bovine serum albumin into the nuclear compartment and of the kinesin-2 motor KIF17 into the ciliary compartment. Using the pore clogging technique, we find that forced dimerization of NUP62 attenuated the gated entry of cytosolic proteins but did not affect entry of membrane proteins or diffusional entry of small cytosolic proteins. We propose a model in which active transport of cytosolic proteins into both nuclear and ciliary compartments requires functional NUPs of the central pore whereas lateral entry of membrane proteins utilizes a different mechanism that is likely specific to each organelle’s limiting membrane.

Fatigue is reflected in a broad array of symptoms that can be particularly impactful in patients with central nervous system (CNS) disorders. Fatigue symptoms can present as core components of the underlying CNS disorder, as part of co-morbid medical conditions, as secondary to psychosocial stressors or physical or mental exertion, and/or secondary to the medications used to treat the disorders. The complex, multi-factorial etiology of fatigue symptoms can obscure the assessment and complicate the treatment intervention for the underlying primary CNS disorder. In this narrative clinical review, we focus on identification, assessment, and adjunctive pharmacological treatment of fatigue symptoms across five distinct CNS disorders. Given the potential impact of fatigue on function and quality of life, it is important to address this symptom as part of a comprehensive evaluation.

A 15-year-old male with transposition of the great arteries presented with exertional chest pain. He was found to have a circumflex coronary artery from the neo-pulmonary artery that had not been transferred during his arterial switch operation. The circumflex coronary artery, fed through collaterals from a re-implanted single coronary artery, resulted in coronary steal. This report describes a management pathway to treat this rare anomaly.