The Gulf pipefish Syngnathus scovelli has emerged as an important species for studying sexual selection, development, and physiology. Comparative evolutionary genomics research involving fishes from Syngnathidae depends on having a high-quality genome assembly and annotation. However, the first S. scovelli genome assembled using short-read sequences and a smaller RNA-sequence dataset has limited contiguity and a relatively poor annotation. Here, using PacBio long-read high-fidelity sequences and a proximity ligation library, we generate an improved assembly to obtain 22 chromosome-level scaffolds. Compared to the first assembly, the gaps in the improved assembly are smaller, the N75 is larger, and our genome is ~95% BUSCO complete. Using a large body of RNA-Seq reads from different tissue types and NCBI's Eukaryotic Annotation Pipeline, we discovered 28,162 genes, of which 8,061 are non-coding genes. Our new genome assembly and annotation are tagged as a RefSeq genome by NCBI and provide enhanced resources for research work involving S. scovelli..

Gynogenetic embryos - those inheriting only maternal DNA - can be experimentally created by fertilizing eggs with radiation-treated sperm containing inactivated paternal chromosomes. Diploidy in the zygotes can be maintained through prevention of the second meiosis or restored by preventing the first mitosis after the maternal chromosome complement has been replicated. These gynogenetic organisms are useful in many fields including aquaculture, evolutionary biology and genomics. Although gynogenetic organisms have been created in numerous species, the completeness of uni-parental inheritance has often been assumed rather than thoroughly quantified across the genome. Instead, when tests of uni-parental inheritance occur, they typically rely on well-studied genetically determined phenotypes that represent a very small sub-set of the genome. Only assessing small genomic regions for paternal inheritance leaves the question of whether some paternal contributions to offspring might still have occurred. In this study, the authors quantify the efficacy of creating gynogenetic diploid three-spined stickleback fish (Gasterosteus aculeatus). To this end, the authors mirrored previous assessments of paternal contribution using well-studied genetically determined phenotypes including sex and genetically dominant morphological traits but expanded on previous studies using dense restriction site-associated DNA sequencing (RAD-seq) markers in parents and offspring to assess paternal inheritance genome-wide. In the gynogenetic diploids, the authors found no male genotypes underlying their phenotypes of interest - sex and dominant phenotypic traits. Using genome-wide assessments of paternal contribution, nevertheless, the authors found evidence of a small, yet potentially important, amount of paternally "leaked" genetic material. The application of this genome-wide approach identifies the need for more widespread assessment of paternal contributions to gynogenetic animals and promises benefits for many aspects of aquaculture, evolutionary biology and genomics.

Abstract The Gulf pipefish Syngnathus scovelli has emerged as an important species in the study of sexual selection, development, and physiology, among other topics. The fish family Syngnathidae, which includes pipefishes, seahorses, and seadragons, has become an increasingly attractive target for comparative research in ecological and evolutionary genomics. These endeavors depend on having a high-quality genome assembly and annotation. However, the first version of the S. scovelli genome assembly was generated by short-read sequencing and annotated using a small set of RNA-sequence data, resulting in limited contiguity and a relatively poor annotation. Here, we present an improved genome assembly and an enhanced annotation, resulting in a new official gene set for S. scovelli . By using PacBio long-read high-fidelity (Hi-Fi) sequences and a proximity ligation (Hi-C) library, we fill small gaps and join the contigs to obtain 22 chromosome-level scaffolds. Compared to the previously published genome, the gaps in our novel genome assembly are smaller, the N75 is much larger (13.3 Mb), and this new genome is around 95% BUSCO complete. The precision of the gene models in the NCBI’s eukaryotic annotation pipeline was enhanced by using a large body of RNA-Seq reads from different tissue types, leading to the discovery of 28,162 genes, of which 8,061 were non-coding genes. This new genome assembly and the annotation are tagged as a RefSeq genome by NCBI and thus provide substantially enhanced genomic resources for future research involving S. scovelli .

ABSTRACT Variation among host-associated microbiomes is well documented across species, populations, and individuals. While numerous factors can contribute to this variation, understanding the influence of host genetic differences on microbial variation is particularly important for predicting co-evolutionary dynamics between hosts and their microbiota. Functional understanding of host genetic and microbial covariation is also of biomedical relevance, for example, providing insights into why some humans are more susceptible to chronic disorders like inflammatory bowel diseases. Unfortunately, disentangling the relative contribution to microbiome variation of host genetics from the environment has been difficult, particularly in humans where confounding environmental effects cannot be completely controlled experimentally. While isogenic laboratory models can be used in controlled environments, the effects on microbiomes of induced large-effect mutations may not recapitulate those of genetic variation observed in nature. Few studies have tested for the direct influence of natural host genetic variation on microbiome differences within a highly controlled environment in which hosts interact freely. To fill this gap, we performed a common garden experiment using families of genetically divergent populations of threespine stickleback fish—an outbred model organism commonly used for determining the genetic basis of complex traits in the context of natural genetic variation. Using germ-free derivation of divergent lines and hybrids between them in this experimental framework, we detected a clear, positive association between stickleback genetic dissimilarity and microbiome dissimilarity. With RAD-seq data, we identified regions of the genome that contributed most significantly to this relationship, providing insight into the genomic architecture of gut microbiome variation. IMPORTANCE A major focus of host-microbe research is to understand how genetic differences, of various magnitudes, among hosts translate to differences in their microbiomes. This has been challenging for animal hosts, including humans, because it is difficult to control environmental variables tightly enough to isolate direct genetic effects on the microbiome. Our work in stickleback fish is a significant contribution because our experimental approach allowed strict control over environmental factors, including standardization of the microbiome from the earliest stage of development and unrestricted co-housing of fish in a truly common environment. Furthermore, we measured host genetic variation over 2,000 regions of the stickleback genome, comparing this information and microbiome composition data among fish from very similar and very different genetic backgrounds. Our findings highlight how differences in the host genome influence microbiome diversity and make a case for future manipulative microbiome experiments that use host systems with naturally occurring genetic variation.

Key Points Question Can a culturally informed community-based outreach intervention increase Latinx participation at SARS-CoV-2 testing events? Findings In this cluster randomized trial of 33 SARS-CoV-2 testing sites, the community health promoters intervention was associated with 3.84 times more Latinx individuals tested per event than control sites, and the intervention was associated with testing a greater proportion of the Latinx populace per event. Meaning The reduction of health disparities experienced by individuals identifying as Latinx during the COVID-19 pandemic may be supported by culturally informed outreach strategies.

Seadragons are a remarkable lineage of teleost fishes in the family Syngnathidae, renowned for having evolved male pregnancy. Comprising three known species, seadragons are widely recognized and admired for their fantastical body forms and coloration, and their specific habitat requirements have made them flagship representatives for marine conservation and natural history interests. Until recently, a gap has been the lack of significant genomic resources for seadragons. We have produced gene-annotated, chromosome-scale genome models for the leafy and weedy seadragon to advance investigations of evolutionary innovation and elaboration of morphological traits in seadragons as well as their pipefish and seahorse relatives. We identified several interesting features specific to seadragon genomes, including divergent noncoding regions near a developmental gene important for integumentary outgrowth, a high genome-wide density of repetitive DNA, and recent expansions of transposable elements and a vesicular trafficking gene family. Surprisingly, comparative analyses leveraging the seadragon genomes and additional syngnathid and outgroup genomes revealed striking, syngnathid-specific losses in the family of fibroblast growth factors (FGFs), which likely involve reorganization of highly conserved gene regulatory networks in ways that have not previously been documented in natural populations. The resources presented here serve as important tools for future evolutionary studies of developmental processes in syngnathids and hold value for conservation of the extravagant seadragons and their relatives.

Model organism research is essential to understand disease mechanisms. However, laboratory-induced genetic models can lack genetic variation and often fail to mimic the spectrum of disease severity. Evolutionary mutant models (EMMs) are species with evolved phenotypes that mimic human disease. EMMs complement traditional laboratory models by providing unique avenues to study gene-by-environment interactions, modular mutations in noncoding regions, and their evolved compensations. EMMs have improved our understanding of complex diseases, including cancer, diabetes, and aging, and illuminated mechanisms in many organs. Rapid advancements of sequencing and genome-editing technologies have catapulted the utility of EMMs, particularly in fish. Fish are the most diverse group of vertebrates, exhibiting a kaleidoscope of specialized phenotypes, many that would be pathogenic in humans but are adaptive in the species' specialized habitat. Importantly, evolved compensations can suggest avenues for novel disease therapies. This review summarizes current research using fish EMMs to advance our understanding of human disease.

Multicellular organisms interact with resident microbes in important ways, and a better understanding of host-microbe interactions is aided by tools such as high-throughput 16S sequencing. However, rigorous evaluation of the veracity of these tools in a different context from which they were developed has often lagged behind. Our goal was to perform one such critical test by examining how variation in tissue preparation and DNA isolation could affect inferences about gut microbiome variation between two genetically divergent lines of threespine stickleback fish maintained in the same laboratory environment. Using careful experimental design and intensive sampling of individuals, we addressed technical and biological sources of variation in 16S-based estimates of microbial diversity. After employing a two-tiered bead beating approach that comprised tissue homogenization followed by microbial lysis in subsamples, we found an extremely minor effect of DNA isolation protocol relative to among-host microbial diversity differences. Abundance estimates for rare operational taxonomic units (OTUs), however, showed much lower reproducibility. Gut microbiome composition was highly variable across fish-even among cohoused siblings-relative to technical replicates, but a subtle effect of host genotype (stickleback line) was nevertheless detected for some microbial taxa. IMPORTANCE Our findings demonstrate the importance of appropriately quantifying biological and technical variance components when attempting to understand major influences on high-throughput microbiome data. Our focus was on understanding among-host (biological) variance in community metrics and its magnitude in relation to within-host (technical) variance, because meaningful comparisons among individuals are necessary in addressing major questions in host-microbe ecology and evolution, such as heritability of the microbiome. Our study design and insights should provide a useful example for others desiring to quantify microbiome variation at biological levels in the face of various technical factors in a variety of systems.

Mounting evidence shows that humans respond heterogeneously to diseases and treatments due to their genetic variation, leading to the growth of the field of personalized medicine. Research on model organisms is vital to seek scientific understandings of disease in this broader context, but current research in nonhuman disease models struggles to demonstrate the genetic variability of humans. One disease that exemplifies this issue is Hermansky-Pudlak Syndrome (HPS). HPS is a genetic syndrome resulting from mutations in one of ten HPS genes involved in endosome trafficking, which can result in oculocutaneous albinism, blood diathesis, pulmonary fibrosis, vision impairment, and granulomatous enterocolitis phenotypically indistinguishable from Crohn’s Disease. Currently no disease models for HPS exist that accurately demonstrate the variability of the human disease. Here we present an outbred mutant model of threespine stickleback (Gasterosteus aculeatus) fish exhibiting symptoms consistent with HPS. Using genetic mapping approaches we narrowed this mutation to a region of the genome containing hps4the causative disease gene of Hermansky-Pudlak Syndrome Type 4. Using histological staining and imaging we are confirming the presence of each symptom comprising the syndrome and hope to develop the first outbred system to tackle this disease. Ultimately, the application of outbred medical models in diverse organisms will be important for the advancement of personalized medicine, greatly enhancing our understanding of how genetic variation affects disease states.

Host selection is often required to foster beneficial microbial symbionts and suppress deleterious pathogens. In animals, the host immune system is at the center of this relationship. Failed host immune system-microbial interactions can result in a persistent inflammatory response in which the immune system indiscriminately attacks resident microbes, and at times the host cells themselves, leading to diseases such as Ulcerative Colitis, Crohn’s Disease, and Psoriasis. Host genetic variation has been linked to both microbiome diversity and to severity of such inflammatory disease states in humans. However, the microbiome and inflammatory states manifest as quantitative traits, which encompass many genes interacting with one another and the environment. The mechanistic relationships among all of these interacting components are still not clear. Developing natural genetic models of host-microbe interactions is therefore fundamental to understanding the complex genetics of these and other diseases. Threespine stickleback ( Gasterosteus aculeatus ) fish are a tractable model for attacking this problem because of abundant population-level genetic and phenotypic variation in the gut inflammatory response. Previous work in our laboratory identified genetically divergent stickleback populations exhibiting differences in intestinal neutrophil activity. We took advantage of this diversity to genetically map variation in an emblematic element of gut inflammation – intestinal neutrophil recruitment – using an F2-intercross mapping framework. We identified three regions of the genome associated with increased intestinal inflammation containing several promising candidate genes. Within these regions we found candidates in the Coagulation/Complement System, NFkB and MAPK pathways along with several genes associated with neurodegenerative diseases commonly accompanying intestinal inflammation as a secondary symptom. These findings highlight the utility of using naturally genetically diverse ‘evolutionary mutant models’ such as threespine stickleback to better understand interactions among host genetic diversity and microbiome variation in health and disease states.

Species such as threespine stickleback (Gasterosteus aculeatus) that inhabit divergent selective environments and that have diversified on different time scales can be of value for understanding evolutionary processes. Here we synthesize high-resolution genotypic and phenotypic data to explore a largely unstudied distribution of threespine stickleback populations living in oceanic and freshwater habitats along coastal and inland regions of Oregon. Many inland aquatic habitats of Oregon remained unglaciated during the last ice age, meaning that some extant Oregon lake and river stickleback may have descended from freshwater populations established long before more well-studied, post-glacial freshwater populations. To address the degree of congruence between genetic and phenotypic divergence, we directly compared Oregon stickleback to much younger (post-glacial) Alaskan populations. We found phenotypic variation in Oregon stickleback to be primarily partitioned between oceanic and freshwater habitats, as has been documented in other stickleback systems. However, the main axis of genetic divergence was between coastal and inland regions regardless of habitat type. Furthermore, when comparing patterns between Oregon and Alaska we found similar levels of phenotypic divergence, but much greater genetic divergence among Oregon's populations. The Oregon stickleback system therefore appears well suited for future studies linking genotypic and phenotypic change, further extending the utility of this small fish to provide general insights into evolutionary processes.

Much of animal-associated microbiome research has been conducted in species for which little is known of their natural ecology and evolution. Microbiome studies that combine population genetic, environment, and geographic data for wild organisms can be very informative, especially in situations where host genetic variation and the environment both influence microbiome variation. The few studies that have related population genetic and microbiome variation in wild populations have been constrained by observation-based kinship data or incomplete genomic information. Here we integrate population genomic and microbiome analyses in wild threespine stickleback fish distributed throughout western Oregon, USA. We found that gut microbiome diversity and composition partitioned more among than within wild host populations and was better explained by host population genetic divergence than by environment and geography. We also identified gut microbial taxa that were most differentially abundant across environments and across genetically divergent populations. Our findings highlight the benefits of studies that investigate host-associated microbiomes in wild organisms.

Selection, via host immunity, is often required to foster beneficial microbial symbionts and suppress deleterious pathogens. In animals, the host immune system is at the center of this relationship. Failed host immune system-microbial interactions can result in a persistent inflammatory response in which the immune system indiscriminately attacks resident microbes, and at times the host cells themselves, leading to diseases such as Ulcerative Colitis, Crohn’s Disease, and Psoriasis. Host genetic variation has been linked to both microbiome diversity and to severity of such inflammatory disease states in humans. However, the microbiome and inflammatory states manifest as quantitative traits, which encompass many genes interacting with one another and the environment. The mechanistic relationships among all of these interacting components are still not clear. Developing natural genetic models of host-microbe interactions is therefore fundamental to understanding the complex genetics of these and other diseases. Threespine stickleback (Gasterosteus aculeatus) fish are a tractable model for attacking this problem because of abundant population-level genetic and phenotypic variation in the gut inflammatory response. Previous work in our laboratory identified genetically divergent stickleback populations exhibiting differences in intestinal neutrophil activity. We took advantage of this diversity to genetically map variation in an emblematic element of gut inflammation - intestinal neutrophil recruitment - using an F2-intercross mapping framework. We identified two regions of the genome associated with increased intestinal inflammation containing several promising candidate genes. Within these regions we found candidates in the Coagulation/Complement System, NFkB and MAPK pathways along with several genes associated with intestinal diseases and neurological diseases commonly accompanying intestinal inflammation as a secondary symptom. These findings highlight the utility of using naturally genetically diverse ‘evolutionary mutant models’ such as threespine stickleback to better understand interactions among host genetic diversity and microbiome variation in health and disease states.

Comparing ontogenetic patterns within a well-described evolutionary context aids in inferring mechanisms of change, including heterochronies or deletion of developmental pathways. Because selection acts on phenotypes throughout ontogeny, any within-taxon developmental variation has implications for evolvability. We compare ontogenetic order and timing of locomotion and defensive traits in three populations of threespine stickleback that have evolutionarily divergent adult forms. This analysis adds to the growing understanding of developmental genetic mechanisms of adaptive change in this evolutionary model species by delineating when chondrogenesis and osteogenesis in two derived populations begin to deviate from the developmental pattern in their immediate ancestors. We found that differences in adult defensive morphologies arise through abolished or delayed initiation of these traits rather than via an overall heterochronic shift, that intra-population ontogenetic variation is increased for some derived traits, and that altered armor developmental timing differentiates the derived populations from each other despite parallels in adult lateral plate armor phenotypes. We found that changes in ossified elements of the pelvic armor are linked to delayed and incomplete development of an early-forming pelvic cartilage, and that this disruption likely presages the variable pelvic vestiges documented in many derived populations.

Animal hosts must co-exist with beneficial microbes while simultaneously being able to mount rapid, non-specific, innate immune responses to pathogenic microbes. How this balance is achieved is not fully understood, and disruption of this relationship can lead to disease. Excessive inflammatory responses to resident microbes are characteristic of certain gastrointestinal pathologies such as inflammatory bowel disease (IBD). The immune dysregulation of IBD has complex genetic underpinnings that cannot be fully recapitulated with single-gene-knockout models. A deeper understanding of the genetic regulation of innate immune responses to resident microbes requires the ability to measure immune responses in the presence and absence of the microbiota using vertebrate models with complex genetic variation. Here, we describe a new gnotobiotic vertebrate model to explore the natural genetic variation that contributes to differences in innate immune responses to microbiota. Threespine stickleback, Gasterosteus aculeatus, has been used to study the developmental genetics of complex traits during the repeated evolution from ancestral oceanic to derived freshwater forms. We established methods to rear germ-free stickleback larvae and gnotobiotic animals monoassociated with single bacterial isolates. We characterized the innate immune response of these fish to resident gut microbes by quantifying the neutrophil cells in conventionally reared monoassociated or germ-free stickleback from both oceanic and freshwater populations grown in a common intermediate salinity environment. We found that oceanic and freshwater fish in the wild and in the laboratory share many intestinal microbial community members. However, oceanic fish mount a strong immune response to residential microbiota, whereas freshwater fish frequently do not. A strong innate immune response was uniformly observed across oceanic families, but this response varied among families of freshwater fish. The gnotobiotic stickleback model that we have developed therefore provides a platform for future studies mapping the natural genetic basis of the variation in immune response to microbes.

Understanding how genetic variation is partitioned across genomes within and among populations is a fundamental problem in ecological and evolutionary genetics. To address this problem, we studied the threespine stickleback fish, which has repeatedly undergone parallel phenotypic and genetic differentiation when oceanic fish have invaded freshwater habitats. While significant evolutionary genetic research has been performed using stickleback from geographic regions that have been deglaciated in the last 20 000 years, less research has focused on freshwater populations that predate the last glacial maximum. We performed restriction‐site associated DNA ‐sequencing ( RAD ‐seq) based population genomic analyses on stickleback from across O regon, which was not glaciated during the last maximum. We sampled stickleback from coastal, Willamette Basin and central O regon sites, analysed their genetic diversity using RAD ‐seq, performed structure analyses, reconstructed their phylogeographic history and tested the hypothesis of recent stickleback introduction into central O regon, where incidence of this species was only recently documented. Our results showed a clear phylogeographic break between coastal and inland populations, with oceanic populations exhibiting the lowest levels of divergence from one another. Willamette Basin and central Oregon populations formed a clade of closely related populations, a finding consistent with a recent introduction of stickleback into central Oregon. Finally, genome‐wide analysis of genetic diversity (π) and correlations of alleles within individuals in subpopulations ( F IS ) supported a role for introgressive hybridization in coastal populations and a recent expansion in central Oregon. Our results exhibit the power of next‐generation sequencing genomic approaches such as RAD ‐seq to identify both historical population structure and recent colonization history.

Evolution of similar phenotypes in independent populations is often taken as evidence of adaptation to the same fitness optimum. However, the genetic architecture of traits might cause evolution to proceed more often toward particular phenotypes, and less often toward others, independently of the adaptive value of the traits. Freshwater populations of Alaskan threespine stickleback have repeatedly evolved the same distinctive opercle shape after divergence from an oceanic ancestor. Here we demonstrate that this pattern of parallel evolution is widespread, distinguishing oceanic and freshwater populations across the Pacific Coast of North America and Iceland. We test whether this parallel evolution reflects genetic bias by estimating the additive genetic variance-covariance matrix (G) of opercle shape in an Alaskan oceanic (putative ancestral) population. We find significant additive genetic variance for opercle shape and that G has the potential to be biasing, because of the existence of regions of phenotypic space with low additive genetic variation. However, evolution did not occur along major eigenvectors of G, rather it occurred repeatedly in the same directions of high evolvability. We conclude that the parallel opercle evolution is most likely due to selection during adaptation to freshwater habitats, rather than due to biasing effects of opercle genetic architecture.