Mating in the Closest Living Relatives of Animals Is Induced by a Bacterial Chondroitinase.

We serendipitously discovered that the marine bacterium Vibrio fischeri induces sexual reproduction in one of the closest living relatives of animals, the choanoflagellate Salpingoeca rosetta. Although bacteria influence everything from nutrition and metabolism to cell biology and development in eukaryotes, bacterial regulation of eukaryotic mating was unexpected. Here, we show that a single V. fischeri protein, the previously uncharacterized EroS, fully recapitulates the aphrodisiac-like activity of live V. fischeri. EroS is a chondroitin lyase; although its substrate, chondroitin sulfate, was previously thought to be an animal synapomorphy, we demonstrate that S. rosetta produces chondroitin sulfate and thus extend the ancestry of this important glycosaminoglycan to the premetazoan era. Finally, we show that V. fischeri, purified EroS, and other bacterial chondroitin lyases induce S. rosetta mating at environmentally relevant concentrations, suggesting that bacteria likely regulate choanoflagellate mating in nature.

The promise and pitfalls of synteny in phylogenomics.

Reconstructing the tree of life remains a central goal in biology. Early methods, which relied on small numbers of morphological or genetic characters, often yielded conflicting evolutionary histories, undermining confidence in the results. Investigations based on phylogenomics, which use hundreds to thousands of loci for phylogenetic inquiry, have provided a clearer picture of life's history, but certain branches remain problematic. To resolve difficult nodes on the tree of life, 2 recent studies tested the utility of synteny, the conserved collinearity of orthologous genetic loci in 2 or more organisms, for phylogenetics. Synteny exhibits compelling phylogenomic potential while also raising new challenges. This Essay identifies and discusses specific opportunities and challenges that bear on the value of synteny data and other rare genomic changes for phylogenomic studies. Synteny-based analyses of highly contiguous genome assemblies mark a new chapter in the phylogenomic era and the quest to reconstruct the tree of life.

Doctors', Patients' and Physician Associates' Perceptions of the Physician Associate Role in the Emergency Department.

INTRODUCTION: The Emergency Department (ED) has seen increased patient attendance and difficulty meeting demands. New healthcare professions such as Physician Associates (PAs) are being utilised to complement the existing medical workforce. Despite the growth of their professions in the United Kingdom, little evidence is available about the perceptions of their roles. OBJECTIVE: This study aims to provide evidence of doctors', PAs' and patients' perceptions of the PA role in the UK ED. METHODS: A mixed methods approach consisted of the following: 1. An online exploratory survey of ED doctors at one English ED over 1 month (February-March 2022). 2. Post consultation semi-structured patient questionnaires over 2 weeks (April 2022). 3. Semi-structured virtual interviews with ED consultants across the four regions of the United Kingdom (3 months in 2022). 4. Semi-structured virtual interviews with ED PAs across the four regions of the United Kingdom (3 months in 2022). The analysis methods that were used included frequency counts and percentages from closed questions, and hybrid thematic analysis of free text and interview transcripts. RESULTS: Four ED consultants and four ED PAs across the United Kingdom were interviewed. Twenty-eight ED doctors participated in the online survey. Fifty-seven patients completed the post consultation questionnaire. Four main themes (PAs being fit for purpose; patient recognition of PAs, PAs providing continuity of care, and future PAs and regulation) were deduced as per the General Medical Council, Good Medical Practice domains (knowledge, skills and development; patients, partnership and communication; colleagues, culture and safety; and trust and professionalism). Other subthemes were induced via hybrid thematic analysis. In this study, doctors and patients had mixed comments about the role of PAs. Most of them were positive as doctor participants perceived PAs to be knowledgeable, highly skilled, with mostly good communication skills, team players, providing continuity of care and overall being fit for purpose. However, some doctor participants commented negatively about PAs for providing little quality healthcare and being inexperienced. There was a desire for career progression among the PA participants and a need to work to their full potential. Although the clinicians of this study displayed a clear understanding of the PA role in the ED, a high frequency of surveyed patients mistook PAs for doctors. It was suggested that future PAs could complete a postqualification programme in emergency medicine, combine roles, be paid on an alternative scale and be formally regulated. CONCLUSION: In this study, mixed views were expressed by ED consultants, ED junior doctors and patients regarding the role of the PA in the ED. Stakeholders can use the information presented to develop a better understanding of the perceptions of the PA role within the UK ED. PATIENT OR PUBLIC CONTRIBUTION: The Patient and Public Involvement and Engagement (PPIE) group, led by Healthwatch, made significant contributions to the study's design by providing valuable feedback on the information sheets and consent forms utilised. The patients' responses helped guide the study's direction and shape its future work. As part of the dissemination activities, the study findings was shared with both the PPIE team and Healthwatch media production team.

Biophysical principles of choanoflagellate self-organization.

Inspired by the patterns of multicellularity in choanoflagellates, the closest living relatives of animals, we quantify the biophysical processes underlying the morphogenesis of rosette colonies in the choanoflagellate Salpingoeca rosetta We find that rosettes reproducibly transition from an early stage of 2-dimensional (2D) growth to a later stage of 3D growth, despite the underlying variability of the cell lineages. Our perturbative experiments demonstrate the fundamental importance of a basally secreted extracellular matrix (ECM) for rosette morphogenesis and show that the interaction of the ECM with cells in the colony physically constrains the packing of proliferating cells and, thus, controls colony shape. Simulations of a biophysically inspired model that accounts for the size and shape of the individual cells, the fraction of ECM, and its stiffness relative to that of the cells suffices to explain our observations and yields a morphospace consistent with observations across a range of multicellular choanoflagellate colonies. Overall, our biophysical perspective on rosette development complements previous genetic perspectives and, thus, helps illuminate the interplay between cell biology and physics in regulating morphogenesis.

Structural and Functional Analysis of Bacterial Sulfonosphingolipids and Rosette-Inducing Factor 2 (RIF-2) by Mass Spectrometry-Guided Isolation and Total Synthesis.

We have analyzed the abundance of bacterial sulfonosphingolipids, including rosette-inducing factors (RIFs), in seven bacterial prey strains by using high-resolution tandem mass spectrometry (HRMS2 ) and molecular networking (MN) within the Global Natural Product Social Molecular Networking (GNPS) web platform. Six sulfonosphingolipids resembling RIFs were isolated and their structures were elucidated based on comparative MS and NMR studies. Here, we also report the first total synthesis of two RIF-2 diastereomers and one congener in 15 and eight synthetic steps, respectively. For the total synthesis of RIF-2 congeners, we employed a decarboxylative cross-coupling reaction to synthesize the necessary branched α-hydroxy fatty acids, and the Garner-aldehyde approach to generate the capnine base carrying three stereogenic centers. Bioactivity studies in the choanoflagellate Salpingoeca rosetta revealed that the rosette inducing activity of RIFs is inhibited dose dependently by the co-occurring sulfonosphingolipid sulfobacins D and F and that activity of RIFs is specific for isolates obtained from Algoriphagus.

The architecture of cell differentiation in choanoflagellates and sponge choanocytes.

Although collar cells are conserved across animals and their closest relatives, the choanoflagellates, little is known about their ancestry, their subcellular architecture, or how they differentiate. The choanoflagellate Salpingoeca rosetta expresses genes necessary for animal development and can alternate between unicellular and multicellular states, making it a powerful model for investigating the origin of animal multicellularity and mechanisms underlying cell differentiation. To compare the subcellular architecture of solitary collar cells in S. rosetta with that of multicellular 'rosette' colonies and collar cells in sponges, we reconstructed entire cells in 3D through transmission electron microscopy on serial ultrathin sections. Structural analysis of our 3D reconstructions revealed important differences between single and colonial choanoflagellate cells, with colonial cells exhibiting a more amoeboid morphology consistent with higher levels of macropinocytotic activity. Comparison of multiple reconstructed rosette colonies highlighted the variable nature of cell sizes, cell-cell contact networks, and colony arrangement. Importantly, we uncovered the presence of elongated cells in some rosette colonies that likely represent a distinct and differentiated cell type, pointing toward spatial cell differentiation. Intercellular bridges within choanoflagellate colonies displayed a variety of morphologies and connected some but not all neighbouring cells. Reconstruction of sponge choanocytes revealed ultrastructural commonalities but also differences in major organelle composition in comparison to choanoflagellates. Together, our comparative reconstructions uncover the architecture of cell differentiation in choanoflagellates and sponge choanocytes and constitute an important step in reconstructing the cell biology of the last common ancestor of animals.

Total Synthesis and Functional Evaluation of IORs, Sulfonolipid-based Inhibitors of Cell Differentiation in Salpingoeca rosetta.

The choanoflagellate Salpingoeca rosetta is an important model system to study the evolution of multicellularity. In this study we developed a new, modular, and scalable synthesis of sulfonolipid IOR-1A (six steps, 27 % overall yield), which acts as bacterial inhibitor of rosette formation in S. rosetta. The synthesis features a decarboxylative cross-coupling reaction of a sulfonic acid-containing tartaric acid derivative with alkyl zinc reagents. Synthesis of 15 modified IOR-1A derivatives, including fluorescent and photoaffinity-based probes, allowed quantification of IOR-1A, localization studies within S. rosetta cells, and evaluation of structure-activity relations. In a proof of concept study, an inhibitory bifunctional probe was employed in proteomic profiling studies, which allowed to deduce binding partners in bacteria and S. rosetta. These results showcase the power of synthetic chemistry to decipher the biochemical basis of cell differentiation processes within S. rosetta.

The genomic and cellular foundations of animal origins.

The first animals arose more than six hundred million years ago, yet they left little impression in the fossil record. Nonetheless, the cell biology and genome composition of the first animal, the Urmetazoan, can be reconstructed through the study of phylogenetically relevant living organisms. Comparisons among animals and their unicellular and colonial relatives reveal that the Urmetazoan likely possessed a layer of epithelium-like collar cells, preyed on bacteria, reproduced by sperm and egg, and developed through cell division, cell differentiation, and invagination. Although many genes involved in development, body patterning, immunity, and cell-type specification evolved in the animal stem lineage or after animal origins, several gene families critical for cell adhesion, signaling, and gene regulation predate the origin of animals. The ancestral functions of these and other genes may eventually be revealed through studies of gene and genome function in early-branching animals and their closest non-animal relatives.

Synergistic Cues from Diverse Bacteria Enhance Multicellular Development in a Choanoflagellate.

Bacteria regulate the life histories of diverse eukaryotes, but relatively little is known about how eukaryotes interpret and respond to multiple bacterial cues encountered simultaneously. To explore how a eukaryote might respond to a combination of bioactive molecules from multiple bacteria, we treated the choanoflagellate Salpingoeca rosetta with two sets of bacterial cues, one that induces mating and another that induces multicellular development. We found that simultaneous exposure to both sets of cues enhanced multicellular development in S. rosetta, eliciting both larger multicellular colonies and an increase in the number of colonies. Thus, rather than conveying conflicting sets of information, these distinct bacterial cues synergize to augment multicellular development. This study demonstrates how a eukaryote can integrate and modulate its response to cues from diverse bacteria, underscoring the potential impact of complex microbial communities on eukaryotic life histories.IMPORTANCE Eukaryotic biology is profoundly influenced by interactions with diverse environmental and host-associated bacteria. However, it is not well understood how eukaryotes interpret multiple bacterial cues encountered simultaneously. This question has been challenging to address because of the complexity of many eukaryotic model systems and their associated bacterial communities. Here, we studied a close relative of animals, the choanoflagellate Salpingoeca rosetta, to explore how eukaryotes respond to diverse bacterial cues. We found that a bacterial chondroitinase that induces mating on its own can also synergize with bacterial lipids that induce multicellular "rosette" development. When encountered together, these cues enhance rosette development, resulting in both the formation of larger rosettes and an increase in the number of rosettes compared to rosette development in the absence of the chondroitinase. These findings highlight how synergistic interactions among bacterial cues can influence the biology of eukaryotes.

Bacterial lipids activate, synergize, and inhibit a developmental switch in choanoflagellates.

In choanoflagellates, the closest living relatives of animals, multicellular rosette development is regulated by environmental bacteria. The simplicity of this evolutionarily relevant interaction provides an opportunity to identify the molecules and regulatory logic underpinning bacterial regulation of development. We find that the rosette-inducing bacterium Algoriphagus machipongonensis produces three structurally divergent classes of bioactive lipids that, together, activate, enhance, and inhibit rosette development in the choanoflagellate Salpingoeca rosetta. One class of molecules, the lysophosphatidylethanolamines (LPEs), elicits no response on its own but synergizes with activating sulfonolipid rosette-inducing factors (RIFs) to recapitulate the full bioactivity of live Algoriphagus. LPEs, although ubiquitous in bacteria and eukaryotes, have not previously been implicated in the regulation of a host-microbe interaction. This study reveals that multiple bacterially produced lipids converge to activate, enhance, and inhibit multicellular development in a choanoflagellate.

The Evolution of Silicon Transport in Eukaryotes.

Biosilicification (the formation of biological structures from silica) occurs in diverse eukaryotic lineages, plays a major role in global biogeochemical cycles, and has significant biotechnological applications. Silicon (Si) uptake is crucial for biosilicification, yet the evolutionary history of the transporters involved remains poorly known. Recent evidence suggests that the SIT family of Si transporters, initially identified in diatoms, may be widely distributed, with an extended family of related transporters (SIT-Ls) present in some nonsilicified organisms. Here, we identify SITs and SIT-Ls in a range of eukaryotes, including major silicified lineages (radiolarians and chrysophytes) and also bacterial SIT-Ls. Our evidence suggests that the symmetrical 10-transmembrane-domain SIT structure has independently evolved multiple times via duplication and fusion of 5-transmembrane-domain SIT-Ls. We also identify a second gene family, similar to the active Si transporter Lsi2, that is broadly distributed amongst siliceous and nonsiliceous eukaryotes. Our analyses resolve a distinct group of Lsi2-like genes, including plant and diatom Si-responsive genes, and sequences unique to siliceous sponges and choanoflagellates. The SIT/SIT-L and Lsi2 transporter families likely contribute to biosilicification in diverse lineages, indicating an ancient role for Si transport in eukaryotes. We propose that these Si transporters may have arisen initially to prevent Si toxicity in the high Si Precambrian oceans, with subsequent biologically induced reductions in Si concentrations of Phanerozoic seas leading to widespread losses of SIT, SIT-L, and Lsi2-like genes in diverse lineages. Thus, the origin and diversification of two independent Si transporter families both drove and were driven by ancient ocean Si levels.

Age-related hyperkyphosis: update of its potential causes and clinical impacts-narrative review.

The present study aims to qualitatively review the contributing factors and health implications of age-related hyperkyphosis. We conducted a narrative review of observational and cohort studies describing the risk factors and epidemiology of hyperkyphosis from 1955 to 2016 using the following key words: kyphosis, hyperkyphosis, posture, age-related hyperkyphosis, kyphotic posture, aetiology and causes. This review included 77 studies. Approximately 60-70 % of the most severe hyperkyphosis cases have no evidence of underlying vertebral compression fractures. Other proposed factors contributing to hyperkyphosis are degenerative disc disease, weakness of back extensor muscles and genetic predisposition. Strength and endurance of back extensor muscles are very important for maintaining normal postural alignment. Recent evidence suggests that age-related hyperkyphosis is not equivalent to spinal osteoporosis. Due to the negative impact of hyperkyphosis on physical function, quality of life and mortality rates, physicians should focus not only on osteoporosis, but also on age-related postural changes. More research about the relationship between spinal morphology and modifiable factors, especially the structural and functional parameters of trunk muscles, could further illuminate our understanding and treatment options for hyperkyphosis.

The Reliability of Standing Sagittal Measurements of Spinal Curvature and Range of Motion in Older Women With and Without Hyperkyphosis Using a Skin-Surface Device.

OBJECTIVE: The purpose of this study was to investigate the intrarater reliability of a skin-surface instrument (Spinal Mouse, Idiag, Voletswil, Switzerland) in measuring standing sagittal curvature and global mobility of the spine in older women with and without hyperkyphosis. METHODS: Measurements were made in 19 women with hyperkyphosis (thoracic kyphosis angle ≥50°), mean age 67 ± 5 years, and 14 women without hyperkyphosis (thoracic kyphosis angle <50°), mean age 63 ± 6 years. Sagittal thoracic and lumbar curvature and mobility of the spine were assessed with the Spinal Mouse during neutral standing, full spinal flexion, and full spinal extension. Tests were performed by the same examiner on 2 days with a 72-hour interval. The intrarater reliability of the measurements was analyzed using the intraclass correlation coefficient, standard error of measurement and minimal detectable change. RESULTS: Intraclass correlation coefficients ranged from 0.89 to 0.99 in both groups. The standard errors of measurement ranged from 1.02° to 2.06° in the hyperkyphosis group and from 1.15° to 2.22° in the normal group. The minimal detectable change ranged from 2.85° to 5.73° in the hyperkyphosis group and from 3.20° to 6.17° in the normal group. CONCLUSIONS: Our results indicated that the Spinal Mouse has excellent intrarater reliability for the measurement of sagittal thoracic and lumbar curvature and mobility of the spine in older women.

Isolation and Synthesis of a Bacterially Produced Inhibitor of Rosette Development in Choanoflagellates.

The choanoflagellate Salpingoeca rosetta is a microbial marine eukaryote that can switch between unicellular and multicellular states. As one of the closest living relatives of animals, this organism has become a model for understanding how multicellularity evolved in the animal lineage. Previously our laboratories isolated and synthesized a bacterially produced sulfonolipid that induces S. rosetta to form multicellular "rosettes." In this study, we report the identification of a bacterially produced inhibitor of rosettes (IOR-1) as well as the total synthesis of this molecule and all of its stereoisomers. Our results confirm the previously noted specificity and potency of rosette-modulating molecules, expand our understanding of the complex chemical ecology between choanoflagellates and rosette-inducing bacteria, and provide a synthetic probe template for conducting further mechanistic studies on the emergence of multicellularity.

Animals in a bacterial world, a new imperative for the life sciences.

In the last two decades, the widespread application of genetic and genomic approaches has revealed a bacterial world astonishing in its ubiquity and diversity. This review examines how a growing knowledge of the vast range of animal-bacterial interactions, whether in shared ecosystems or intimate symbioses, is fundamentally altering our understanding of animal biology. Specifically, we highlight recent technological and intellectual advances that have changed our thinking about five questions: how have bacteria facilitated the origin and evolution of animals; how do animals and bacteria affect each other's genomes; how does normal animal development depend on bacterial partners; how is homeostasis maintained between animals and their symbionts; and how can ecological approaches deepen our understanding of the multiple levels of animal-bacterial interaction. As answers to these fundamental questions emerge, all biologists will be challenged to broaden their appreciation of these interactions and to include investigations of the relationships between and among bacteria and their animal partners as we seek a better understanding of the natural world.

Evolutionary insights into premetazoan functions of the neuronal protein homer.

Reconstructing the evolution and ancestral functions of synaptic proteins promises to shed light on how neurons first evolved. The postsynaptic density (PSD) protein Homer scaffolds membrane receptors and regulates Ca(2+) signaling in diverse metazoan cell types (including neurons and muscle cells), yet its ancestry and core functions are poorly understood. We find that the protein domain organization and essential biochemical properties of metazoan Homer proteins, including their ability to tetramerize, are conserved in the choanoflagellate Salpingoeca rosetta, one of the closest living relatives of metazoans. Unlike in neurons, Homer localizes to the nucleoplasm in S. rosetta and interacts directly with Flotillin, a protein more commonly associated with cell membranes. Surprisingly, we found that the Homer/Flotillin interaction and its localization to the nucleus are conserved in metazoan astrocytes. These findings suggest that Homer originally interacted with Flotillin in the nucleus of the last common ancestor of metazoans and choanoflagellates and was later co-opted to function as a membrane receptor scaffold in the PSD.

Modern disposable hydrogel contact lens removal has a minimal effect on intraocular pressure.

PURPOSE: To determine whether the removal of modern disposable hydrogel contact lenses may influence intraocular pressure (IOP) measurement, and if so, how long the effect may last. METHODS: Twenty-five healthy experienced contact lens wearers aged 19-25 inserted their lenses at least 30 min prior to the study. Each participant was asked to remove a contact lens from one eye (control eye, selected at random) upon commencement of the study, and then to remove the lens in the other eye (lens-wearing eye) after a 30 min washout period. IOP was measured immediately in both eyes using non-contact tonometry, then repeated every 3 min for 12 min. To determine the change in IOP due to lens removal, the IOP in the lens-wearing eye was compared to the control eye using paired t-tests at each time point. RESULTS: The IOP was significantly higher in the lens-wearing eye immediately following lens removal (0.7 ± 1.0 mmHg, t(24) = 3.46, p < 0.01), but was within baseline values at 3 min (0.2 ± 1.0 mmHg, t(24) = 0.84, p = 0.41), 6 min (0.3 ± 1.1 mmHg, t(24) = 1.39, p = 0.18), 9 min (0.3 ± 1.2 mmHg, t(24) = 1.14, p = 0.27) and 12 min (-0.1 ± 0.9 mmHg, t(24) = -0.49, p = 0.63, paired t-test). CONCLUSIONS: There was a slight statistically significant increase in IOP following contact lens removal, with a maximum duration of 3 min. Given the small magnitude of the change in IOP, and its transient nature, there appears to be no clinical reason to delay IOP measurements following the removal of modern disposable hydrogel contact lenses.

Origin of metazoan cadherin diversity and the antiquity of the classical cadherin/ß-catenin complex.

The evolution of cadherins, which are essential for metazoan multicellularity and restricted to metazoans and their closest relatives, has special relevance for understanding metazoan origins. To reconstruct the ancestry and evolution of cadherin gene families, we analyzed the genomes of the choanoflagellate Salpingoeca rosetta, the unicellular outgroup of choanoflagellates and metazoans Capsaspora owczarzaki, and a draft genome assembly from the homoscleromorph sponge Oscarella carmela. Our finding of a cadherin gene in C. owczarzaki reveals that cadherins predate the divergence of the C. owczarzaki, choanoflagellate, and metazoan lineages. Data from these analyses also suggest that the last common ancestor of metazoans and choanoflagellates contained representatives of at least three cadherin families, lefftyrin, coherin, and hedgling. Additionally, we find that an O. carmela classical cadherin has predicted structural features that, in bilaterian classical cadherins, facilitate binding to the cytoplasmic protein ß-catenin and, thereby, promote cadherin-mediated cell adhesion. In contrast with premetazoan cadherin families (i.e., those conserved between choanoflagellates and metazoans), the later appearance of classical cadherins coincides with metazoan origins.