Carbonate-sensitive phytotransferrin controls high-affinity iron uptake in diatoms.

In vast areas of the ocean, the scarcity of iron controls the growth and productivity of phytoplankton. Although most dissolved iron in the marine environment is complexed with organic molecules, picomolar amounts of labile inorganic iron species (labile iron) are maintained within the euphotic zone and serve as an important source of iron for eukaryotic phytoplankton and particularly for diatoms. Genome-enabled studies of labile iron utilization by diatoms have previously revealed novel iron-responsive transcripts, including the ferric iron-concentrating protein ISIP2A, but the mechanism behind the acquisition of picomolar labile iron remains unknown. Here we show that ISIP2A is a phytotransferrin that independently and convergently evolved carbonate ion-coordinated ferric iron binding. Deletion of ISIP2A disrupts high-affinity iron uptake in the diatom Phaeodactylum tricornutum, and uptake is restored by complementation with human transferrin. ISIP2A is internalized by endocytosis, and manipulation of the seawater carbonic acid system reveals a second-order dependence on the concentrations of labile iron and carbonate ions. In P. tricornutum, the synergistic interaction of labile iron and carbonate ions occurs at environmentally relevant concentrations, revealing that carbonate availability co-limits iron uptake. Phytotransferrin sequences have a broad taxonomic distribution and are abundant in marine environmental genomic datasets, suggesting that acidification-driven declines in the concentration of seawater carbonate ions will have a negative effect on this globally important eukaryotic iron acquisition mechanism.

Genomic Confirmation of Borrelia garinii, United States.

Lyme disease is a multisystem disorder primarily caused by Borrelia burgdorferi sensu lato. However, B. garinii, which has been identified on islands off the coast of Newfoundland and Labrador, Canada, is a cause of Lyme disease in Eurasia. We report isolation and whole-genome nucleotide sequencing of a B. garinii isolate from a cotton mouse (Peromyscus gossypinus) in South Carolina, USA. We identified a second B. garinii isolate from the same repository. Phylogenetic analysis does not associate these isolates with the previously described isolates of B. garinii from Canada.

Reduction-dependent siderophore assimilation in a model pennate diatom.

Iron uptake by diatoms is a biochemical process with global biogeochemical implications. In large regions of the surface ocean diatoms are both responsible for the majority of primary production and frequently experiencing iron limitation of growth. The strategies used by these phytoplankton to extract iron from seawater constrain carbon flux into higher trophic levels and sequestration into sediments. In this study we use reverse genetic techniques to target putative iron-acquisition genes in the model pennate diatom Phaeodactylum tricornutum We describe components of a reduction-dependent siderophore acquisition pathway that relies on a bacterial-derived receptor protein and provides a viable alternative to inorganic iron uptake under certain conditions. This form of iron uptake entails a close association between diatoms and siderophore-producing organisms during low-iron conditions. Homologs of these proteins are found distributed across diatom lineages, suggesting the significance of siderophore utilization by diatoms in the marine environment. Evaluation of specific proteins enables us to confirm independent iron-acquisition pathways in diatoms and characterize their preferred substrates. These findings refine our mechanistic understanding of the multiple iron-uptake systems used by diatoms and help us better predict the influence of iron speciation on taxa-specific iron bioavailability.

Highly flexible metabolism of the marine euglenozoan protist Diplonema papillatum.

BACKGROUND: The phylum Euglenozoa is a group of flagellated protists comprising the diplonemids, euglenids, symbiontids, and kinetoplastids. The diplonemids are highly abundant and speciose, and recent tools have rendered the best studied representative, Diplonema papillatum, genetically tractable. However, despite the high diversity of diplonemids, their lifestyles, ecological functions, and even primary energy source are mostly unknown. RESULTS: We designed a metabolic map of D. papillatum cellular bioenergetic pathways based on the alterations of transcriptomic, proteomic, and metabolomic profiles obtained from cells grown under different conditions. Comparative analysis in the nutrient-rich and nutrient-poor media, as well as the absence and presence of oxygen, revealed its capacity for extensive metabolic reprogramming that occurs predominantly on the proteomic rather than the transcriptomic level. D. papillatum is equipped with fundamental metabolic routes such as glycolysis, gluconeogenesis, TCA cycle, pentose phosphate pathway, respiratory complexes, ß-oxidation, and synthesis of fatty acids. Gluconeogenesis is uniquely dominant over glycolysis under all surveyed conditions, while the TCA cycle represents an eclectic combination of standard and unusual enzymes. CONCLUSIONS: The identification of conventional anaerobic enzymes reflects the ability of this protist to survive in low-oxygen environments. Furthermore, its metabolism quickly reacts to restricted carbon availability, suggesting a high metabolic flexibility of diplonemids, which is further reflected in cell morphology and motility, correlating well with their extreme ecological valence.

Complex Evolution of Insect Insulin Receptors and Homologous Decoy Receptors, and Functional Significance of Their Multiplicity.

Evidence accumulates that the functional plasticity of insulin and insulin-like growth factor signaling in insects could spring, among others, from the multiplicity of insulin receptors (InRs). Their multiple variants may be implemented in the control of insect polyphenism, such as wing or caste polyphenism. Here, we present a comprehensive phylogenetic analysis of insect InR sequences in 118 species from 23 orders and investigate the role of three InRs identified in the linden bug, Pyrrhocoris apterus, in wing polymorphism control. We identified two gene clusters (Clusters I and II) resulting from an ancestral duplication in a late ancestor of winged insects, which remained conserved in most lineages, only in some of them being subject to further duplications or losses. One remarkable yet neglected feature of InR evolution is the loss of the tyrosine kinase catalytic domain, giving rise to decoys of InR in both clusters. Within the Cluster I, we confirmed the presence of the secreted decoy of insulin receptor in all studied Muscomorpha. More importantly, we described a new tyrosine kinase-less gene (DR2) in the Cluster II, conserved in apical Holometabola for ∼300 My. We differentially silenced the three P. apterus InRs and confirmed their participation in wing polymorphism control. We observed a pattern of Cluster I and Cluster II InRs impact on wing development, which differed from that postulated in planthoppers, suggesting an independent establishment of insulin/insulin-like growth factor signaling control over wing development, leading to idiosyncrasies in the co-option of multiple InRs in polyphenism control in different taxa.

Olisthodiscus represents a new class of Ochrophyta.

The phylogenetic diversity of Ochrophyta, a diverse and ecologically important radiation of algae, is still incompletely understood even at the level of the principal lineages. One taxon that has eluded simple classification is the marine flagellate genus Olisthodiscus. We investigated Olisthodiscus luteus K-0444 and documented its morphological and genetic differences from the NIES-15 strain, which we described as Olisthodiscus tomasii sp. nov. Phylogenetic analyses of combined 18S and 28S rRNA sequences confirmed that Olisthodiscus constitutes a separate, deep, ochrophyte lineage, but its position could not be resolved. To overcome this problem, we sequenced the plastid genome of O. luteus K-0444 and used the new data in multigene phylogenetic analyses, which suggested that Olisthodiscus is a sister lineage of the class Pinguiophyceae within a broader clade additionally including Chrysophyceae, Synchromophyceae, and Eustigmatophyceae. Surprisingly, the Olisthodiscus plastid genome contained three genes, ycf80, cysT, and cysW, inherited from the rhodophyte ancestor of the ochrophyte plastid yet lost from all other ochrophyte groups studied so far. Combined with nuclear genes for CysA and Sbp proteins, Olisthodiscus is the only known ochrophyte possessing a plastidial sulfate transporter SulT. In addition, the finding of a cemA gene in the Olisthodiscus plastid genome and an updated phylogenetic analysis ruled out the previously proposed hypothesis invoking horizontal cemA transfer from a green algal plastid into Synurales. Altogether, Olisthodiscus clearly represents a novel phylogenetically distinct ochrophyte lineage, which we have proposed as a new class, Olisthodiscophyceae.

Evolution of metabolic capabilities and molecular features of diplonemids, kinetoplastids, and euglenids.

BACKGROUND: The Euglenozoa are a protist group with an especially rich history of evolutionary diversity. They include diplonemids, representing arguably the most species-rich clade of marine planktonic eukaryotes; trypanosomatids, which are notorious parasites of medical and veterinary importance; and free-living euglenids. These different lifestyles, and particularly the transition from free-living to parasitic, likely require different metabolic capabilities. We carried out a comparative genomic analysis across euglenozoan diversity to see how changing repertoires of enzymes and structural features correspond to major changes in lifestyles. RESULTS: We find a gradual loss of genes encoding enzymes in the evolution of kinetoplastids, rather than a sudden decrease in metabolic capabilities corresponding to the origin of parasitism, while diplonemids and euglenids maintain more metabolic versatility. Distinctive characteristics of molecular machines such as kinetochores and the pre-replication complex that were previously considered specific to parasitic kinetoplastids were also identified in their free-living relatives. Therefore, we argue that they represent an ancestral rather than a derived state, as thought until the present. We also found evidence of ancient redundancy in systems such as NADPH-dependent thiol-redox. Only the genus Euglena possesses the combination of trypanothione-, glutathione-, and thioredoxin-based systems supposedly present in the euglenozoan common ancestor, while other representatives of the phylum have lost one or two of these systems. Lastly, we identified convergent losses of specific metabolic capabilities between free-living kinetoplastids and ciliates. Although this observation requires further examination, it suggests that certain eukaryotic lineages are predisposed to such convergent losses of key enzymes or whole pathways. CONCLUSIONS: The loss of metabolic capabilities might not be associated with the switch to parasitic lifestyle in kinetoplastids, and the presence of a highly divergent (or unconventional) kinetochore machinery might not be restricted to this protist group. The data derived from the transcriptomes of free-living early branching prokinetoplastids suggests that the pre-replication complex of Trypanosomatidae is a highly divergent version of the conventional machinery. Our findings shed light on trends in the evolution of metabolism in protists in general and open multiple avenues for future research.

Environmental determinants of the distribution of planktonic diplonemids and kinetoplastids in the oceans.

We analysed a widely used barcode, the V9 region of the 18S rRNA gene, to study the effect of environmental conditions on the distribution of two related heterotrophic protistan lineages in marine plankton, kinetoplastids and diplonemids. We relied on a major published dataset (Tara Oceans) where samples from the mesopelagic zone were available from just 32 of 123 locations, and both groups are most abundant in this zone. To close sampling gaps and obtain more information from the deeper ocean, we collected 57 new samples targeting especially the mesopelagic zone. We sampled in three geographic regions: the Arctic, two depth transects in the Adriatic Sea, and the anoxic Cariaco Basin. In agreement with previous studies, both protist groups are most abundant and diverse in the mesopelagic zone. In addition to that, we found that their abundance, richness, and community structure also depend on geography, oxygen concentration, salinity, temperature, and other environmental variables reflecting the abundance of algae and nutrients. Both groups studied here demonstrated similar patterns, although some differences were also observed. Kinetoplastids and diplonemids prefer tropical regions and nutrient-rich conditions and avoid high oxygen concentration, high salinity, and high density of algae.

Neobodonids are dominant kinetoplastids in the global ocean.

Kinetoplastid flagellates comprise basal mostly free-living bodonids and derived obligatory parasitic trypanosomatids, which belong to the best-studied protists. Due to their omnipresence in aquatic environments and soil, the bodonids are of ecological significance. Here, we present the first global survey of marine kinetoplastids and compare it with the strikingly different patterns of abundance and diversity in their sister clade, the diplonemids. Based on analysis of 18S rDNA V9 ribotypes obtained from 124 sites sampled during the Tara Oceans expedition, our results show generally low to moderate abundance and diversity of planktonic kinetoplastids. Although we have identified all major kinetoplastid lineages, 98% of kinetoplastid reads are represented by neobodonids, namely specimens of the Neobodo and Rhynchomonas genera, which make up 59% and 18% of all reads, respectively. Most kinetoplastids have small cell size (0.8-5 µm) and tend to be more abundant in the mesopelagic as compared to the euphotic zone. Some of the most abundant operational taxonomic units have distinct geographical distributions, and three novel putatively parasitic neobodonids were identified, along with their potential hosts.

Evolution and metabolic significance of the urea cycle in photosynthetic diatoms.

Diatoms dominate the biomass of phytoplankton in nutrient-rich conditions and form the basis of some of the world's most productive marine food webs. The diatom nuclear genome contains genes with bacterial and plastid origins as well as genes of the secondary endosymbiotic host (the exosymbiont), yet little is known about the relative contribution of each gene group to diatom metabolism. Here we show that the exosymbiont-derived ornithine-urea cycle, which is similar to that of metazoans but is absent in green algae and plants, facilitates rapid recovery from prolonged nitrogen limitation. RNA-interference-mediated knockdown of a mitochondrial carbamoyl phosphate synthase impairs the response of nitrogen-limited diatoms to nitrogen addition. Metabolomic analyses indicate that intermediates in the ornithine-urea cycle are particularly depleted and that both the tricarboxylic acid cycle and the glutamine synthetase/glutamate synthase cycles are linked directly with the ornithine-urea cycle. Several other depleted metabolites are generated from ornithine-urea cycle intermediates by the products of genes laterally acquired from bacteria. This metabolic coupling of bacterial- and exosymbiont-derived proteins seems to be fundamental to diatom physiology because the compounds affected include the major diatom osmolyte proline and the precursors for long-chain polyamines required for silica precipitation during cell wall formation. So far, the ornithine-urea cycle is only known for its essential role in the removal of fixed nitrogen in metazoans. In diatoms, this cycle serves as a distribution and repackaging hub for inorganic carbon and nitrogen and contributes significantly to the metabolic response of diatoms to episodic nitrogen availability. The diatom ornithine-urea cycle therefore represents a key pathway for anaplerotic carbon fixation into nitrogenous compounds that are essential for diatom growth and for the contribution of diatoms to marine productivity.

The protist cultural renaissance.

Protists are key players in the biosphere. Here, we provide a perspective on integrating protist culturing with omics approaches, imaging, and high-throughput single-cell manipulation strategies, concluding with actions required for a successful return of the golden age of protist culturing.

A common red algal origin of the apicomplexan, dinoflagellate, and heterokont plastids.

The discovery of a nonphotosynthetic plastid in malaria and other apicomplexan parasites has sparked a contentious debate about its evolutionary origin. Molecular data have led to conflicting conclusions supporting either its green algal origin or red algal origin, perhaps in common with the plastid of related dinoflagellates. This distinction is critical to our understanding of apicomplexan evolution and the evolutionary history of endosymbiosis and photosynthesis; however, the two plastids are nearly impossible to compare due to their nonoverlapping information content. Here we describe the complete plastid genome sequences and plastid-associated data from two independent photosynthetic lineages represented by Chromera velia and an undescribed alga CCMP3155 that we show are closely related to apicomplexans. These plastids contain a suite of features retained in either apicomplexan (four plastid membranes, the ribosomal superoperon, conserved gene order) or dinoflagellate plastids (form II Rubisco acquired by horizontal transfer, transcript polyuridylylation, thylakoids stacked in triplets) and encode a full collective complement of their reduced gene sets. Together with whole plastid genome phylogenies, these characteristics provide multiple lines of evidence that the extant plastids of apicomplexans and dinoflagellates were inherited by linear descent from a common red algal endosymbiont. Our phylogenetic analyses also support their close relationship to plastids of heterokont algae, indicating they all derive from the same endosymbiosis. Altogether, these findings support a relatively simple path of linear descent for the evolution of photosynthesis in a large proportion of algae and emphasize plastid loss in several lineages (e.g., ciliates, Cryptosporidium, and Phytophthora).

Intragenomic diversity of the V9 hypervariable domain in eukaryotes has little effect on metabarcoding.

Metabarcoding revolutionized our understanding of diversity and ecology of microorganisms in different habitats. However, it is also associated with several inherent biases, one of which is associated with intragenomic diversity of a molecular barcode. Here, we compare intragenomic variability of the V9 region of the 18S rRNA gene in 19 eukaryotic phyla abundant in marine plankton. The level of intragenomic variability is comparable across all the phyla, and in most genomes and transcriptomes one V9 sequence and one OTU is predominant. However, most of the variability observed at the barcode level is probably caused by sequencing errors and is mitigated by using a denoising tool, DADA2. The SWARM algorithm commonly used in metabarcoding studies is not optimal for collapsing genuine and erroneous sequences into a single OTU, leading to an overestimation of diversity in metabarcoding data. For an unknown reason, SWARM inflates diversity of eupelagonemids more than that of other eukaryotes.

Ultrastructure and 3D reconstruction of a diplonemid protist (Diplonemea) and its novel membranous organelle.

IMPORTANCE: The knowledge of cell biology of a eukaryotic group is essential for correct interpretation of ecological and molecular data. Although diplonemid protists are one of the most species-rich lineages of marine eukaryotes, only very fragmentary information is available about the cellular architecture of this taxonomically diverse group. Here, a large serial block-face scanning electron microscopy data set complemented with light and fluorescence microscopy allowed the first detailed three-dimensional reconstruction of a diplonemid species. We describe numerous previously unknown peculiarities of the cellular architecture and cell division characteristic for diplonemid flagellates, and illustrate the obtained results with multiple three-dimensional models, comprehensible for non-specialists in protist ultrastructure.

Water masses shape pico-nano eukaryotic communities of the Weddell Sea.

Polar oceans belong to the most productive and rapidly changing environments, yet our understanding of this fragile ecosystem remains limited. Here we present an analysis of a unique set of DNA metabarcoding samples from the western Weddell Sea sampled throughout the whole water column and across five water masses with different characteristics and different origin. We focus on factors affecting the distribution of planktonic pico-nano eukaryotes and observe an ecological succession of eukaryotic communities as the water masses move away from the surface and as oxygen becomes depleted with time. At the beginning of this succession, in the photic zone, algae, bacteriovores, and predators of small eukaryotes dominate the community, while another community develops as the water sinks deeper, mostly composed of parasitoids (syndinians), mesoplankton predators (radiolarians), and diplonemids. The strongly correlated distribution of syndinians and diplonemids along the depth and oxygen gradients suggests their close ecological link and moves us closer to understanding the biological role of the latter group in the ocean ecosystem.

Circadian rhythms and circadian clock gene homologs of complex alga Chromera velia.

Most organisms on Earth are affected by periodic changes in their environment. The circadian clock is an endogenous device that synchronizes behavior, physiology, or biochemical processes to an approximately 24-hour cycle, allowing organisms to anticipate the periodic changes of day and night. Although circadian clocks are widespread in organisms, the actual molecular components differ remarkably among the clocks of plants, animals, fungi, and prokaryotes. Chromera velia is the closest known photosynthetic relative of apicomplexan parasites. Formation of its motile stage, zoospores, has been described as associated with the light part of the day. We examined the effects on the periodic release of the zoospores under different light conditions and investigated the influence of the spectral composition on zoosporogenesis. We performed a genomic search for homologs of known circadian clock genes. Our results demonstrate the presence of an almost 24-hour free-running cycle of zoosporogenesis. We also identified the blue light spectra as the essential compound for zoosporogenesis. Further, we developed a new and effective method for zoospore separation from the culture and estimated the average motility speed and lifespan of the C. velia zoospores. Our genomic search identified six cryptochrome-like genes, two genes possibly related to Arabidopsis thaliana CCA/LHY, whereas no homolog of an animal, cyanobacterial, or fungal circadian clock gene was found. Our results suggest that C. velia has a functional circadian clock, probably based mainly on a yet undefined mechanism.

Short-term acidification promotes diverse iron acquisition and conservation mechanisms in upwelling-associated phytoplankton.

Coastal upwelling regions are among the most productive marine ecosystems but may be threatened by amplified ocean acidification. Increased acidification is hypothesized to reduce iron bioavailability for phytoplankton thereby expanding iron limitation and impacting primary production. Here we show from community to molecular levels that phytoplankton in an upwelling region respond to short-term acidification exposure with iron uptake pathways and strategies that reduce cellular iron demand. A combined physiological and multi-omics approach was applied to trace metal clean incubations that introduced 1200 ppm CO2 for up to four days. Although variable, molecular-level responses indicate a prioritization of iron uptake pathways that are less hindered by acidification and reductions in iron utilization. Growth, nutrient uptake, and community compositions remained largely unaffected suggesting that these mechanisms may confer short-term resistance to acidification; however, we speculate that cellular iron demand is only temporarily satisfied, and longer-term acidification exposure without increased iron inputs may result in increased iron stress.

Colpodella spp.-like parasite infection in woman, China.

The phylum Apicomplexa comprises intracellular protozoa that include many human pathogens. Their nearest relatives are chromerids and colpodellids. We report a case of a Babesia spp.-like relapsing infection caused by a newly described microorganism related to the Apicomplexa. This case is highly suggestive of a previously undescribed type of colpodellid that infects vertebrates.

Molecular identification and genotyping of Microsporidia in selected hosts.

The work is described by microscopic analysis, the serological analysis (IFAT) and the molecular analysis of isolates from clinical samples (blood, faeces and urine) from ten domestic rabbits (Oryctolagus cuniculus), breed Malický, four New Zealand domestic rabbits, 11 sows of breed Slo0076akian Improved White and 15 clinically healthy laboratory BALB/c mice. The aim of the study was to validate the suitability of species-unspecific primer pairs 530F and 580R for genotype determination of the Microsporidia strain and species-specific primer pairs ECUNF and ECUNR, SINTF and SINTR and EBIER1 and EBIEF1 for the determination of E ncephalitozoon cuniculi, Encephalitozoon intestinalis and Enterocytozoon bieneusi species for diagnostic purposes. Sequences of animals were compared with those from the GenBank database. In rabbits, two murine genotypes II and four canine genotypes III were identified. Genotype II was identified in mice. The Encephalitozoon intestinalis identified in the sample from swine showed no genetic heterogeneity.

Deep Learning Analysis of Polish Electronic Health Records for Diagnosis Prediction in Patients with Cardiovascular Diseases.

Electronic health records naturally contain most of the medical information in the form of doctor's notes as unstructured or semi-structured texts. Current deep learning text analysis approaches allow researchers to reveal the inner semantics of text information and even identify hidden consequences that can offer extra decision support to doctors. In the presented article, we offer a new automated analysis of Polish summary texts of patient hospitalizations. The presented models were found to be able to predict the final diagnosis with almost 70% accuracy based just on the patient's medical history (only 132 words on average), with possible accuracy increases when adding further sentences from hospitalization results; even one sentence was found to improve the results by 4%, and the best accuracy of 78% was achieved with five extra sentences. In addition to detailed descriptions of the data and methodology, we present an evaluation of the analysis using more than 50,000 Polish cardiology patient texts and dive into a detailed error analysis of the approach. The results indicate that the deep analysis of just the medical history summary can suggest the direction of diagnosis with a high probability that can be further increased just by supplementing the records with further examination results.