Psychological distress and alcohol use disorder in college students of color: Social support's moderating properties.

Alcohol use is prevalent among college students. Research has found that psychological distress in the form of depression, anxiety, or loneliness has been consistently associated with greater alcohol use. Because Students of Color (SoC) disproportionately experience greater psychological distress than White students, it is critical to determine buffers against psychological distress and subsequent alcohol use consequences. Previous literature found that social support can protect against the effects of psychological distress and weaken its link with alcohol use. This study aimed to determine the moderating effect of social support in the relation between psychological distress and alcohol use among SoC. College Students of Color from across the U.S. (n = 211, Mage = 27.51, SD = 9.63) were recruited via Prolific to answer an online survey through Qualtrics. Linear regression analyses showed that psychological distress variables (i.e., depression, anxiety, and loneliness) were positively associated with alcohol use. However, moderation analysis did not find any form of social support to moderate the relation between psychological distress variables and alcohol use. Future research needs to identify other protective factors against alcohol use to support SoC in their academic journey.

The genome of the colonial hydroid Hydractinia reveals that their stem cells use a toolkit of evolutionarily shared genes with all animals.

Hydractinia is a colonial marine hydroid that shows remarkable biological properties, including the capacity to regenerate its entire body throughout its lifetime, a process made possible by its adult migratory stem cells, known as i-cells. Here, we provide an in-depth characterization of the genomic structure and gene content of two Hydractinia species, Hydractinia symbiolongicarpus and Hydractinia echinata, placing them in a comparative evolutionary framework with other cnidarian genomes. We also generated and annotated a single-cell transcriptomic atlas for adult male H. symbiolongicarpus and identified cell-type markers for all major cell types, including key i-cell markers. Orthology analyses based on the markers revealed that Hydractinia's i-cells are highly enriched in genes that are widely shared amongst animals, a striking finding given that Hydractinia has a higher proportion of phylum-specific genes than any of the other 41 animals in our orthology analysis. These results indicate that Hydractinia's stem cells and early progenitor cells may use a toolkit shared with all animals, making it a promising model organism for future exploration of stem cell biology and regenerative medicine. The genomic and transcriptomic resources for Hydractinia presented here will enable further studies of their regenerative capacity, colonial morphology, and ability to distinguish self from nonself.

Innate Allorecognition in Transplantation: Ancient Mechanisms With Modern Impact.

Through the effective targeting of the adaptive immune system, solid organ transplantation became a life-saving therapy for organ failure. However, beyond 1 y of transplantation, there is little improvement in transplant outcomes. The adaptive immune response requires the activation of the innate immune system. There are no modalities for the specific targeting of the innate immune system involvement in transplant rejection. However, the recent discovery of innate allorecognition and innate immune memory presents novel targets in transplantation that will increase our understanding of organ rejection and might aid in improving transplant outcomes. In this review, we look at the latest developments in the study of innate allorecognition and innate immune memory in transplantation.

The genome of the colonial hydroid Hydractinia reveals their stem cells utilize a toolkit of evolutionarily shared genes with all animals.

Hydractinia is a colonial marine hydroid that exhibits remarkable biological properties, including the capacity to regenerate its entire body throughout its lifetime, a process made possible by its adult migratory stem cells, known as i-cells. Here, we provide an in-depth characterization of the genomic structure and gene content of two Hydractinia species, H. symbiolongicarpus and H. echinata, placing them in a comparative evolutionary framework with other cnidarian genomes. We also generated and annotated a single-cell transcriptomic atlas for adult male H. symbiolongicarpus and identified cell type markers for all major cell types, including key i-cell markers. Orthology analyses based on the markers revealed that Hydractinia's i-cells are highly enriched in genes that are widely shared amongst animals, a striking finding given that Hydractinia has a higher proportion of phylum-specific genes than any of the other 41 animals in our orthology analysis. These results indicate that Hydractinia's stem cells and early progenitor cells may use a toolkit shared with all animals, making it a promising model organism for future exploration of stem cell biology and regenerative medicine. The genomic and transcriptomic resources for Hydractinia presented here will enable further studies of their regenerative capacity, colonial morphology, and ability to distinguish self from non-self.

Coevolution of the Tlx homeobox gene with medusa development (Cnidaria: Medusozoa).

Cnidarians display a wide diversity of life cycles. Among the main cnidarian clades, only Medusozoa possesses a swimming life cycle stage called the medusa, alternating with a benthic polyp stage. The medusa stage was repeatedly lost during medusozoan evolution, notably in the most diverse medusozoan class, Hydrozoa. Here, we show that the presence of the homeobox gene Tlx in Cnidaria is correlated with the presence of the medusa stage, the gene having been lost in clades that ancestrally lack a medusa (anthozoans, endocnidozoans) and in medusozoans that secondarily lost the medusa stage. Our characterization of Tlx expression indicate an upregulation of Tlx during medusa development in three distantly related medusozoans, and spatially restricted expression patterns in developing medusae in two distantly related species, the hydrozoan Podocoryna carnea and the scyphozoan Pelagia noctiluca. These results suggest that Tlx plays a key role in medusa development and that the loss of this gene is likely linked to the repeated loss of the medusa life cycle stage in the evolution of Hydrozoa.

How Emergency Medicine Residents Have Conversations About Life-Sustaining Treatments in Critical Illness: A Qualitative Study Using Inductive Thematic Analysis.

STUDY OBJECTIVE: The inherent pressures of high-acuity, critical illness in the emergency department create a unique environment whereby acute goals-of-care discussions must be had with patients or substitute decision makers to rapidly decide between divergent treatment paths. Among university-affiliated hospitals, resident physicians are often conducting these highly consequential discussions. This study aimed to use qualitative methods to explore how emergency medicine residents make recommendations regarding life-sustaining treatments during acute goals-of-care discussions in critical illness. METHODS: Using qualitative methods, semistructured interviews were conducted with a purposive sample of emergency medicine residents in Canada from August to December 2021. Inductive thematic analysis of the interview transcripts was conducted using line-by-line coding, and key themes were identified through comparative analysis. Data collection continued until thematic saturation was reached. RESULTS: Seventeen emergency medicine residents from 9 Canadian universities were interviewed. Two factors guided residents' treatment recommendations (a duty to provide a recommendation and the balance between disease prognosis and patient values). Three factors influenced residents' comfort when making recommendations (time constraints, uncertainty, and moral distress). CONCLUSION: While conducting acute goals-of-care discussions with critically ill patients or their substitute decision makers in the emergency department, residents felt a sense of duty to provide a recommendation informed by an intersection between the patient's disease prognosis and the patient's values. Their comfort in making these recommendations was limited by time constraints, uncertainty, and moral distress. These factors are important for informing future educational strategies.

XY sex determination in a cnidarian.

BACKGROUND: Sex determination occurs across animal species, but most of our knowledge about its mechanisms comes from only a handful of bilaterian taxa. This limits our ability to infer the evolutionary history of sex determination within animals. RESULTS: In this study, we generated a linkage map of the genome of the colonial cnidarian Hydractinia symbiolongicarpus and used it to demonstrate that this species has an XX/XY sex determination system. We demonstrate that the X and Y chromosomes have pseudoautosomal and non-recombining regions. We then use the linkage map and a method based on the depth of sequencing coverage to identify genes encoded in the non-recombining region and show that many of them have male gonad-specific expression. In addition, we demonstrate that recombination rates are enhanced in the female genome and that the haploid chromosome number in Hydractinia is n = 15. CONCLUSIONS: These findings establish Hydractinia as a tractable non-bilaterian model system for the study of sex determination and the evolution of sex chromosomes.

A family of unusual immunoglobulin superfamily genes in an invertebrate histocompatibility complex.

Most colonial marine invertebrates are capable of allorecognition, the ability to distinguish between themselves and conspecifics. One long-standing question is whether invertebrate allorecognition genes are homologous to vertebrate histocompatibility genes. In the cnidarian Hydractinia symbiolongicarpus, allorecognition is controlled by at least two genes, Allorecognition 1 (Alr1) and Allorecognition 2 (Alr2), which encode highly polymorphic cell-surface proteins that serve as markers of self. Here, we show that Alr1 and Alr2 are part of a family of 41 Alr genes, all of which reside in a single genomic interval called the Allorecognition Complex (ARC). Using sensitive homology searches and highly accurate structural predictions, we demonstrate that the Alr proteins are members of the immunoglobulin superfamily (IgSF) with V-set and I-set Ig domains unlike any previously identified in animals. Specifically, their primary amino acid sequences lack many of the motifs considered diagnostic for V-set and I-set domains, yet they adopt secondary and tertiary structures nearly identical to canonical Ig domains. Thus, the V-set domain, which played a central role in the evolution of vertebrate adaptive immunity, was present in the last common ancestor of cnidarians and bilaterians. Unexpectedly, several Alr proteins also have immunoreceptor tyrosine-based activation motifs and immunoreceptor tyrosine-based inhibitory motifs in their cytoplasmic tails, suggesting they could participate in pathways homologous to those that regulate immunity in humans and flies. This work expands our definition of the IgSF with the addition of a family of unusual members, several of which play a role in invertebrate histocompatibility.

Venom system variation and the division of labor in the colonial hydrozoan Hydractinia symbiolongicarpus.

Cnidarians (jellyfish, hydroids, sea anemones, and corals) possess a unique method for venom production, maintenance, and deployment through a decentralized system composed of different types of venom-filled stinging structures called nematocysts. In many species, nematocyst types are distributed heterogeneously across functionally distinct tissues. This has led to a prediction that different nematocyst types contain specific venom components. The colonial hydrozoan, Hydractinia symbiolongicarpus, is an ideal system to study the functional distribution of nematocyst types and their venoms, given that they display a division of labor through functionally distinct polyps within the colony. Here, we characterized the composition and distribution of nematocysts (cnidome) in the different polyp types and show that the feeding polyp (gastrozooid) has a distinct cnidome compared to the reproductive (gonozooid) and predatory polyp (dactylozooid). We generated a nematocyst-specific reporter line to track nematocyst development (nematogenesis) in H. symbiolongicarpus, and were able to confirm that nematogenesis primarily occurs in the mid-region of the gastrozooid and throughout stolons (tubes of epithelia that connect the polyps in the colony). This reporter line enabled us to isolate a nematocyst-specific lineage of cells for de novo transcriptome assembly, annotate venom-like genes (VLGs) and determine differential expression (DE) across polyp types. We show that a majority of VLGs are upregulated in gastrozooids, consistent with it being the primary site of active nematogenesis. However, despite gastrozooids producing more nematocysts, we found a number of VLGs significantly upregulated in dactylozooids, suggesting that these VLGs may be important for prey-capture. Our transgenic Hydractinia reporter line provides an opportunity to explore the complex interplay between venom composition, nematocyst diversity, and ecological partitioning in a colonial hydrozoan that displays a division of labor.

The evolution and development of coloniality in hydrozoans.

Hydrozoan colonies display a variety of shapes and sizes including encrusting, upright, and pelagic forms. Phylogenetic patterns reveal a complex evolutionary history of these distinct colony forms, as well as colony loss. Within a species, phenotypic variation in colonies as a response to changing environmental cues and resources has been documented. The patterns of branching of colony specific tissue, called stolons in encrusting colonies and stalks in upright colonies, are likely under the control of signaling mechanisms whose changing expression in evolution and development are responsible for the diversity of hydrozoan colony forms. Although mechanisms of polyp development have been well studied, little research has focused on colony development and patterning. In the few studies that investigated mechanisms governing colony patterning, the Wnt signaling pathway has been implicated. The diversity of colony form, evolutionary patterns, and mechanisms of colony variation in Hydrozoa are reviewed here.

Zinc protection of fertilized eggs is an ancient feature of sexual reproduction in animals.

One of the earliest and most prevalent barriers to successful reproduction is polyspermy, or fertilization of an egg by multiple sperm. To prevent these supernumerary fertilizations, eggs have evolved multiple mechanisms. It has recently been proposed that zinc released by mammalian eggs at fertilization may block additional sperm from entering. Here, we demonstrate that eggs from amphibia and teleost fish also release zinc. Using Xenopus laevis as a model, we document that zinc reversibly blocks fertilization. Finally, we demonstrate that extracellular zinc similarly disrupts early embryonic development in eggs from diverse phyla, including Cnidaria, Echinodermata, and Chordata. Our study reveals that a fundamental strategy protecting human eggs from fertilization by multiple sperm may have evolved more than 650 million years ago.

Frizzled3 expression and colony development in hydractiniid hydrozoans.

Hydractiniid hydrozoan colonies are comprised of individual polyps connected by tube-like stolons or a sheet-like mat. Mat and stolons function to integrate the colony through continuous epithelia and shared gastrovascular cavity. Although mechanisms of hydrozoan polyp development have been well studied, little is known about the signaling processes governing the patterning of colonies. Here we investigate the Wnt receptor family Frizzled. Phylogenetic analysis reveals that hydrozoans possess four Frizzled orthologs. We find that one of these genes, Frizzled3, shows a spatially restricted expression pattern in colony-specific tissue in two hydractiniid hydrozoans, Hydractinia symbiolongicarpus and Podocoryna carnea, in a manner that corresponds to their distinct colony forms (stolonal mat in Hydractinia and free stolons in Podocoryna). Interestingly, Frizzled3 was lost in the genome of Hydra, which is a solitary polyp and thus lacks colony-specific tissue. Current evidence suggests that the Wnt signaling pathway plays a key role in the evolution of colony diversity and colony loss in Hydrozoa.

CRISPR/Cas9-mediated gene knockin in the hydroid Hydractinia symbiolongicarpus.

BACKGROUND: Hydractinia symbiolongicarpus, a colonial cnidarian, is a tractable model system for many cnidarian-specific and general biological questions. Until recently, tests of gene function in Hydractinia have relied on laborious forward genetic approaches, randomly integrated transgenes, or transient knockdown of mRNAs. RESULTS: Here, we report the use of CRISPR/Cas9 genome editing to generate targeted genomic insertions in H. symbiolonigcarpus. We used CRISPR/Cas9 to promote homologous recombination of two fluorescent reporters, eGFP and tdTomato, into the Eukaryotic elongation factor 1 alpha (Eef1a) locus. We demonstrate that the transgenes are expressed ubiquitously and are stable over two generations of breeding. We further demonstrate that CRISPR/Cas9 genome editing can be used to mark endogenous proteins with FLAG or StrepII-FLAG affinity tags to enable in vivo and ex vivo protein studies. CONCLUSIONS: This is the first account of CRISPR/Cas9 mediated knockins in Hydractinia and the first example of the germline transmission of a CRISPR/Cas9 inserted transgene in a cnidarian. The ability to precisely insert exogenous DNA into the Hydractinia genome will enable sophisticated genetic studies and further development of functional genomics tools in this understudied cnidarian model.

Fibrinogen Concentrate in the Special Operations Forces Environment.

Introduction: Hemorrhage is the most common cause of death among Special Operations Force (SOF) soldiers. Bringing remote damage control resuscitation into the far-forward combat environment is logistically challenging, as it requires blood products that generally require a robust cold chain. Alternatively, lyophilized products such as fibrinogen concentrate, which does not require thawing or blood group compatibility testing before use, might be advantageous in damage control resuscitation in the battlefield. In this report, we review the evidence for the use of fibrinogen concentrate in the Canadian SOF environment. Materials and Methods: The literature on the use of fibrinogen concentrate in the trauma setting was reviewed by Canadian Forces Services Working Group, in three separate meetings. Multiple stakeholders were consulted to obtain authoritative perspectives from subject matter experts on the use of fibrinogen concentrate in the Canadian SOF environment. We also conducted a comparison review of fibrinogen content, pathogen risk, shelf life, and methods required for use for fresh frozen plasma, cryoprecipitate, and fibrinogen concentrate relevant to their application in the far-forward combat environment. Results: Indications and a protocol for the use of fibrinogen as an adjunct to fresh whole blood were formulated based on a literature review and clinical expert opinion. Alternative strategies and other lyophilized blood products were considered before selecting fibrinogen concentrate as the lyophilized blood product of choice. Fibrinogen concentrate is an ABO-universal blood product with an excellent safety profile. Training was conducted by subject matter experts within civilian trauma centers and at military training facilities. The clinical efficacy and safety were confirmed by monitoring the use of fibrinogen concentrate in deployed combat settings. Conclusion: Fibrinogen concentrate is a useful adjunct to remote damage control resuscitation in the SOF environment. Fibrinogen concentrate was found to be robust for transport into the SOF environment and is widely accepted among SOF operators and medics.

Oligonucleotide-Mediated Genome Editing Provides Precision and Function to Engineered Nucleases and Antibiotics in Plants.

Here, we report a form of oligonucleotide-directed mutagenesis for precision genome editing in plants that uses single-stranded oligonucleotides (ssODNs) to precisely and efficiently generate genome edits at DNA strand lesions made by DNA double strand break reagents. Employing a transgene model in Arabidopsis (Arabidopsis thaliana), we obtained a high frequency of precise targeted genome edits when ssODNs were introduced into protoplasts that were pretreated with the glycopeptide antibiotic phleomycin, a nonspecific DNA double strand breaker. Simultaneous delivery of ssODN and a site-specific DNA double strand breaker, either transcription activator-like effector nucleases (TALENs) or clustered, regularly interspaced, short palindromic repeats (CRISPR/Cas9), resulted in a much greater targeted genome-editing frequency compared with treatment with DNA double strand-breaking reagents alone. Using this site-specific approach, we applied the combination of ssODN and CRISPR/Cas9 to develop an herbicide tolerance trait in flax (Linum usitatissimum) by precisely editing the 5'-ENOLPYRUVYLSHIKIMATE-3-PHOSPHATE SYNTHASE (EPSPS) genes. EPSPS edits occurred at sufficient frequency that we could regenerate whole plants from edited protoplasts without employing selection. These plants were subsequently determined to be tolerant to the herbicide glyphosate in greenhouse spray tests. Progeny (C1) of these plants showed the expected Mendelian segregation of EPSPS edits. Our findings show the enormous potential of using a genome-editing platform for precise, reliable trait development in crop plants.

Patterns of Wnt signaling in the life cycle of Podocoryna carnea and its implications for medusae evolution in Hydrozoa (Cnidaria).

Hydrozoans are known for their complex life cycles, alternating between benthic, asexually reproducing polyps and pelagic, sexually reproducing medusae. Although patterning in hydrozoan polyps has been well studied, little is known about the signaling mechanisms governing medusa development. In order to investigate the role of Wnt signaling in medusa development, we use RNA-Seq data collected from three discrete life cycle stages of Podocoryna carnea to assemble, annotate, and assess enrichment and differential expression (DE) of Wnt pathway elements in P. carnea's transcriptome. Enrichment analyses revealed a statistically significant enrichment of DE Wnt signaling transcripts in the transcriptome of P. carnea, of which, the vast majority of these were significantly up-regulated in developing and adult medusae stages. Whole mount in situ hybridization (ISH) reveals co-expression of the Wnt ligand, Wnt3, and a membrane bound Wnt receptor, frizzled3, at the distal and oral ends of the developmental axes of medusae and polyps in P. carnea. DE and ISH results presented here reveal expression of Wnt signaling components consistent with it playing a role in medusa development. Specifically, Wnt ligand expression in the oral region suggests that the Wnt pathway may play a role in medusa patterning, similar to that of polyps. Previous Wnt expression studies in hydrozoan taxa with reduced medusa have failed to detect co-expression of Wnt3 and a frizzled receptor at their truncated developmental axes, suggesting that down regulation of Wnt pathway elements may play a key role in the loss of the medusa life cycle stage in hydrozoan evolution.

Phylogenomic Analyses Support Traditional Relationships within Cnidaria.

Cnidaria, the sister group to Bilateria, is a highly diverse group of animals in terms of morphology, lifecycles, ecology, and development. How this diversity originated and evolved is not well understood because phylogenetic relationships among major cnidarian lineages are unclear, and recent studies present contrasting phylogenetic hypotheses. Here, we use transcriptome data from 15 newly-sequenced species in combination with 26 publicly available genomes and transcriptomes to assess phylogenetic relationships among major cnidarian lineages. Phylogenetic analyses using different partition schemes and models of molecular evolution, as well as topology tests for alternative phylogenetic relationships, support the monophyly of Medusozoa, Anthozoa, Octocorallia, Hydrozoa, and a clade consisting of Staurozoa, Cubozoa, and Scyphozoa. Support for the monophyly of Hexacorallia is weak due to the equivocal position of Ceriantharia. Taken together, these results further resolve deep cnidarian relationships, largely support traditional phylogenetic views on relationships, and provide a historical framework for studying the evolutionary processes involved in one of the most ancient animal radiations.

Interspecific Differential Expression Analysis of RNA-Seq Data Yields Insight into Life Cycle Variation in Hydractiniid Hydrozoans.

Hydrozoans are known for their complex life cycles, which can alternate between an asexually reproducing polyp stage and a sexually reproducing medusa stage. Most hydrozoan species, however, lack a free-living medusa stage and instead display a developmentally truncated form, called a medusoid or sporosac, which generally remains attached to the polyp. Although evolutionary transitions in medusa truncation and loss have been investigated phylogenetically, little is known about the genes involved in the development and loss of this life cycle stage. Here, we present a new workflow for evaluating differential expression (DE) between two species using short read Illumina RNA-seq data. Through interspecific DE analyses between two hydractiniid hydrozoans, Hydractinia symbiolongicarpus and Podocoryna carnea, we identified genes potentially involved in the developmental, functional, and morphological differences between the fully developed medusa of P. carnea and reduced sporosac of H. symbiolongicarpus. A total of 10,909 putative orthologs of H. symbiolongicarpus and P. carnea were identified from de novo assemblies of short read Illumina data. DE analysis revealed 938 of these are differentially expressed between P. carnea developing and adult medusa, when compared with H. symbiolongicarpus sporosacs, the majority of which have not been previously characterized in cnidarians. In addition, several genes with no corresponding ortholog in H. symbiolongicarpus were expressed in developing medusa of P. carnea. Results presented here show interspecific DE analyses of RNA-seq data to be a sensitive and reliable method for identifying genes and gene pathways potentially involved in morphological and life cycle differences between species.

Differential gene expression between functionally specialized polyps of the colonial hydrozoan Hydractinia symbiolongicarpus (Phylum Cnidaria).

BACKGROUND: A colony of the hydrozoan Hydractinia symbiolongicarpus comprises genetically identical yet morphologically distinct and functionally specialized polyp types. The main labor divisions are between feeding, reproduction and defense. In H. symbiolongicarpus, the feeding polyp (called a gastrozooid) has elongated tentacles and a mouth, which are absent in the reproductive polyp (gonozooid) and defensive polyp (dactylozooid). Instead, the dactylozooid has an extended body column with an abundance of stinging cells (nematocysts) and the gonozooid bears gonophores on its body column. Morphological differences between polyp types can be attributed to simple changes in their axial patterning during development, and it has long been hypothesized that these specialized polyps arose through evolutionary alterations in oral-aboral patterning of the ancestral gastrozooid. RESULTS: An assembly of 66,508 transcripts (>200 bp) were generated using short-read Illumina RNA-Seq libraries constructed from feeding, reproductive, and defensive polyps of H. symbiolongicarpus. Using several different annotation methods, approximately 54% of the transcripts were annotated. Differential expression analyses were conducted between these three polyp types to isolate genes that may be involved in functional, histological, and pattering differences between polyp types. Nearly 7 K transcripts were differentially expressed in a polyp-specific manner, including members of the homeodomain, myosin, toxin and BMP gene families. We report the spatial expression of a subset of these polyp-specific transcripts to validate our differential expression analyses. CONCLUSIONS: While potentially originating through simple changes in patterning, polymorphic polyps in Hydractinia are the result of differentially expressed functional, structural, and patterning genes.The differentially expressed genes identified in our study provide a starting point for future investigations of the developmental patterning and functional differences that are displayed in the different polyp types that confer a division of labor within a colony of H. symbiolongicarpus.