Facilitating taxonomy and phylogenetics: An informative and cost-effective protocol integrating long amplicon PCRs and third-generation sequencing.

Phylogenetic inference has become a standard technique in integrative taxonomy and systematics, as well as in biogeography and ecology. DNA barcodes are often used for phylogenetic inference, despite being strongly limited due to their low number of informative sites. Also, because current DNA barcodes are based on a fraction of a single, fast-evolving gene, they are highly unsuitable for resolving deeper phylogenetic relationships due to saturation. In recent years, methods that analyse hundreds and thousands of loci at once have improved the resolution of the Tree of Life, but these methods require resources, experience and molecular laboratories that most taxonomists do not have. This paper introduces a PCR-based protocol that produces long amplicons of both slow- and fast-evolving unlinked mitochondrial and nuclear gene regions, which can be sequenced by the affordable and portable ONT MinION platform with low infrastructure or funding requirements. As a proof of concept, we inferred a phylogeny of a sample of 63 spider species from 20 families using our proposed protocol. The results were overall consistent with the results from approaches based on hundreds and thousands of loci, while requiring just a fraction of the cost and labour of such approaches, making our protocol accessible to taxonomists worldwide.

Ectogenesis and the value of gestational ties.

Ectogenesis technology would make it possible to support the complete gestational development of a human being outside the female body. Proponents argue that this technology offers a welcome opportunity to expand reproductive options for those unable or unwilling to gestate. However, by completely bypassing pregnancy, the use of ectogenesis prevents the formation of gestational family ties. Consequently, it has faced criticism for perpetuating a patriarchal view of the family that undermines the moral significance of gestation. The concern is that the introduction of this technology might result in the loss of reproductive autonomy for those who desire to experience pregnancy, as they face pressures to opt for ectogenesis instead. Existing accounts of family values define parents' rights to rear a child, but they fail to establish a right to gestate that can protect an individual's interest in bearing a child. To provide a more comprehensive account of family values, I argue that pregnancy involves a unique quality of intimacy and can make distinct contributions to one's flourishing. Based on this premise, I defend a fundamental moral right to gestate that can help safeguard the option of pregnancy for those who desire it. In conclusion, I consider how a prospective gestator need not provide optimal conditions for fetal development in the way that ectogenesis promises in order for their choice of pregnancy to be justified.

Richness and resilience in the Pacific: DNA metabarcoding enables parallelized evaluation of biogeographic patterns.

Islands make up a large proportion of Earth's biodiversity, yet are also some of the most sensitive systems to environmental perturbation. Biogeographic theory predicts that geologic age, area, and isolation typically drive islands' diversity patterns, and thus potentially impact non-native spread and community homogenization across island systems. One limitation in testing such predictions has been the difficulty of performing comprehensive inventories of island biotas and distinguishing native from introduced taxa. Here, we use DNA metabarcoding and statistical modelling as a high throughput method to survey community-wide arthropod richness, the proportion of native and non-native species, and the incursion of non-natives into primary habitats on three archipelagos in the Pacific - the Ryukyus, the Marianas and Hawaii - which vary in age, isolation and area. Diversity patterns largely match expectations based on island biogeography theory, with the oldest and most geographically connected archipelago, the Ryukyus, showing the highest taxonomic richness and lowest proportion of introduced species. Moreover, we find evidence that forest habitats are more resilient to incursions of non-natives in the Ryukyus than in the less taxonomically rich archipelagos. Surprisingly, we do not find evidence for biotic homogenization across these three archipelagos: the assemblage of non-native species on each island is highly distinct. Our study demonstrates the potential of DNA metabarcoding to facilitate rapid estimation of biogeographic patterns, the spread of non-native species, and the resilience of ecosystems.

Collective and harmonized high throughput barcoding of insular arthropod biodiversity: Toward a Genomic Observatories Network for islands.

Current understanding of ecological and evolutionary processes underlying island biodiversity is heavily shaped by empirical data from plants and birds, although arthropods comprise the overwhelming majority of known animal species, and as such can provide key insights into processes governing biodiversity. Novel high throughput sequencing (HTS) approaches are now emerging as powerful tools to overcome limitations in the availability of arthropod biodiversity data, and hence provide insights into these processes. Here, we explored how these tools might be most effectively exploited for comprehensive and comparable inventory and monitoring of insular arthropod biodiversity. We first reviewed the strengths, limitations and potential synergies among existing approaches of high throughput barcode sequencing. We considered how this could be complemented with deep learning approaches applied to image analysis to study arthropod biodiversity. We then explored how these approaches could be implemented within the framework of an island Genomic Observatories Network (iGON) for the advancement of fundamental and applied understanding of island biodiversity. To this end, we identified seven island biology themes at the interface of ecology, evolution and conservation biology, within which collective and harmonized efforts in HTS arthropod inventory could yield significant advances in island biodiversity research.

Multiple paths toward repeated phenotypic evolution in the spiny-leg adaptive radiation (Tetragnatha; Hawai'i).

The repeated evolution of phenotypes provides clear evidence for the role of natural selection in driving evolutionary change. However, the evolutionary origin of repeated phenotypes can be difficult to disentangle as it can arise from a combination of factors such as gene flow, shared ancestral polymorphisms or mutation. Here, we investigate the presence of these evolutionary processes in the Hawaiian spiny-leg Tetragnatha adaptive radiation, which includes four microhabitat-specialists or ecomorphs, with different body pigmentation and size (Green, Large Brown, Maroon, and Small Brown). We investigated the evolutionary history of this radiation using 76 newly generated low-coverage, whole-genome resequenced samples, along with phylogenetic and population genomic tools. Considering the Green ecomorph as the ancestral state, our results suggest that the Green ecomorph likely re-evolved once, the Large Brown and Maroon ecomorphs evolved twice and the Small Brown evolved three times. We found that the evolution of the Maroon and Small Brown ecomorphs likely involved ancestral hybridization events, while the Green and Large Brown ecomorphs likely evolved through novel mutations, despite a high rate of incomplete lineage sorting in the dataset. Our findings demonstrate that the repeated evolution of ecomorphs in the Hawaiian spiny-leg Tetragnatha is influenced by multiple evolutionary processes.

STI Testing among Medicaid Enrollees Initiating PrEP for HIV Prevention in Six Southern States.

OBJECTIVES: The purpose of this study was to measure sexually transmitted infection (STI) testing among Medicaid enrollees initiating preexposure prophylaxis (PrEP) to prevent human immunodeficiency virus. Secondary data are in the form of Medicaid enrollment and claims data in six states in the US South. METHODS: Research partnerships in six states in the US South developed a distributed research network to accomplish study aims. Each state identified all first-time PrEP users in fiscal year 2017-2018 (combined N = 990) and measured the presence of STI testing for chlamydia, syphilis, and gonorrhea through 2019. Each state calculated the percentage of individuals with at least one STI test during 3-, 6-, and 12-month follow-up periods. RESULTS: The proportion of first-time PrEP users that received an STI test varied by state: 37% to 67% of all of the individuals in each state who initiated PrEP received a test within the first 6 months of PrEP treatment and 50% to 77% received a test within the first 12 months. CONCLUSIONS: Although the Centers for Disease Control and Prevention recommends STI testing at least every 6 months for PrEP users, our analysis of Medicaid data suggests that STI testing occurs less frequently than recommended in populations at elevated risk of syphilis, gonorrhea, and chlamydia.

A Review of the Molecular Landscape of Adenoid Cystic Carcinoma of the Lacrimal Gland.

Adenoid cystic carcinoma (ACC) has a worldwide incidence of three to four cases per million population. Although more cases occur in the minor and major salivary glands, it is the most common lacrimal gland malignancy. ACC has a low-grade, indolent histological appearance, but is relentlessly progressive over time and has a strong proclivity to recur and/or metastasise. Current treatment options are limited to complete surgical excision and adjuvant radiotherapy. Intra-arterial systemic therapy is a recent innovation. Recurrent/metastatic disease is common due to perineural invasion, and it is largely untreatable as it is refractory to conventional chemotherapeutic agents. Given the rarity of this tumour, the molecular mechanisms that govern disease pathogenesis are poorly understood. There is an unmet, critical need to develop effective, personalised targeted therapies for the treatment of ACC in order to reduce morbidity and mortality associated with the disease. This review details the evidence relating to the molecular underpinnings of ACC of the lacrimal gland, including the MYB-NFIB chromosomal translocations, Notch-signalling pathway aberrations, DNA damage repair gene mutations and epigenetic modifications.

Protein Tyrosine Phosphatase Non-Receptor 11 (PTPN11/Shp2) as a Driver Oncogene and a Novel Therapeutic Target in Non-Small Cell Lung Cancer (NSCLC).

PTPN11 encodes the SHP2 protein tyrosine phosphatase that activates the mitogen-activated protein kinase (MAPK) pathway upstream of KRAS and MEK. PTPN11/Shp2 somatic mutations occur frequently in Juvenile myelomonocytic leukaemia (JMML); however, the role of mutated PTPN11 in lung cancer tumourigenesis and its utility as a therapeutic target has not been fully addressed. We applied mass-spectrometry-based genotyping to DNA extracted from the tumour and matched the normal tissue of 356 NSCLC patients (98 adenocarcinomas (LUAD) and 258 squamous cell carcinomas (LUSC)). Further, PTPN11 mutation cases were identified in additional cohorts, including TCGA, Broad, and MD Anderson datasets and the COSMIC database. PTPN11 constructs harbouring PTPN11 E76A, A72D and C459S mutations were stably expressed in IL-3 dependent BaF3 cells and NSCLC cell lines (NCI-H1703, NCI-H157, NCI-H1299). The MAPK and PI3K pathway activation was evaluated using Western blotting. PTPN11/Shp2 phosphatase activity was measured in whole-cell protein lysates using an Shp2 assay kit. The Shp2 inhibitor (SHPi) was assessed both in vitro and in vivo in a PTPN11-mutated cell line for improved responses to MAPK and PI3K targeting therapies. Somatic PTPN11 hotspot mutations occurred in 4/98 (4.1%) adenocarcinomas and 7/258 (2.7%) squamous cells of 356 NSCLC patients. Additional 26 PTPN11 hotspot mutations occurred in 23 and 3 adenocarcinomas and squamous cell carcinoma, respectively, across the additional cohorts. Mutant PTPN11 significantly increased the IL-3 independent survival of Ba/F3 cells compared to wildtype PTPN11 (p < 0.0001). Ba/F3, NCI-H1703, and NCI-H157 cells expressing mutant PTPN11 exhibited increased PTPN11/Shp2 phosphatase activity and phospho-ERK1/2 levels compared to cells expressing wildtype PTPN11. The transduction of the PTPN11 inactivating mutation C459S into NSCLC cell lines led to decreased phospho-ERK, as well as decreased phospho-AKT in the PTPN11-mutated NCI-H661 cell line. NCI-H661 cells (PTPN11-mutated, KRAS-wild type) were significantly more sensitive to growth inhibition by the PI3K inhibitor copanlisib (IC50: 13.9 ± 4.7 nM) compared to NCI-H1703 (PTPN11/KRAS-wild type) cells (IC50: >10,000 nM). The SHP2 inhibitor, in combination with the PI3K targeting therapy copanlisib, showed no significant difference in tumour development in vivo; however, this significantly prevented MAPK pathway induction in vitro (p < 0.0001). PTPN11/Shp2 demonstrated the in vitro features of a driver oncogene and could potentially sensitize NSCLC cells to PI3K inhibition and inhibit MAPK pathway activation following PI3K pathway targeting.

Recurrent lacrimal gland pleomorphic adenoma.

Pleomorphic adenoma is the most common tumour of the lacrimal gland. Correct preoperative diagnosis is essential as the tumour should be removed en bloc. Incisional biopsy is contraindicated due to risk of recurrence and subsequent risk of malignancy. We identified 13 patients who were diagnosed with lacrimal gland pleomorphic adenoma in our institution over a 31-year period from 1990 to 2021. Of these, we describe four patients, three males and one female, who underwent incisional biopsy. One of these patients had a number of recurrences and required an orbital exenteration. The other two were treated with radiation to the orbit and experienced severe dry eye post-radiotherapy. All three patients are currently disease-free. The fourth patient underwent an incisional biopsy of his lacrimal gland pleomorphic adenoma (LGPA) within the last 6 months and has been referred for radiotherapy. The reasons for a lack of preoperative diagnosis of LGPA included clinical uncertainty and broad radiologic differential. None of the patients who had an en bloc resection experienced recurrence. We recommend that a fine needle aspirate (FNA) biopsy in lieu of a tissue biopsy should be performed where LGPA is included in the clinical and radiological differential diagnosis. If incisional biopsy is required, the suture track should be marked so that the entire biopsy track is removed with the specimen.

What is adaptive radiation? Many manifestations of the phenomenon in an iconic lineage of Hawaiian spiders.

Adaptive radiation provides the ideal context for identifying and testing the processes that drive evolutionary diversification. However, different adaptive radiations show a variety of different patterns, making it difficult to come up with universal rules that characterize all such systems. Diversification may occur via several mechanisms including non-adaptive divergence, adaptation to novel environments, or character displacement driven by competition. Here, we characterize the ways these different drivers contribute to present-day diversity patterns, using the exemplary adaptive radiation of Hawaiian long-jawed orbweaver (Tetragnatha) spiders. We present the most taxonomically comprehensive phylogenetic hypothesis to date for this group, using 10 molecular markers and representatives from every known species across the archipelago. Among the lineages that make up this remarkable radiation, we find evidence for multiple diversification modalities. Several clades appear to have diversified in allopatry under a narrow range of ecological conditions, highlighting the role of niche conservatism in speciation. Others have shifted into new environments and evolved traits that appear to be adaptive in those environments. Still others show evidence for character displacement by close relatives, often resulting in convergent evolution of stereotyped ecomorphs. All of the above mechanisms seem to have played a role in giving rise to the exceptional diversity of morphological, ecological and behavioral traits represented among the many species of Hawaiian Tetragnatha. Taking all these processes into account, and testing how they operate in different systems, may allow us to identify universal principles underlying adaptive radiation.

Design, Implementation, and Evolution of the Medicaid Outcomes Distributed Research Network (MODRN).

BACKGROUND: In the US, Medicaid covers over 80 million Americans. Comparing access, quality, and costs across Medicaid programs can provide policymakers with much-needed information. As each Medicaid agency collects its member data, multiple barriers prevent sharing Medicaid data between states. To address this gap, the Medicaid Outcomes Distributed Research Network (MODRN) developed a research network of states to conduct rapid multi-state analyses without sharing individual-level data across states. OBJECTIVE: To describe goals, design, implementation, and evolution of MODRN to inform other research networks. METHODS: MODRN implemented a distributed research network using a common data model, with each state analyzing its own data; developed standardized measure specifications and statistical software code to conduct analyses; and disseminated findings to state and federal Medicaid policymakers. Based on feedback on Medicaid agency priorities, MODRN first sought to inform Medicaid policy to improve opioid use disorder treatment, particularly medication treatment. RESULTS: Since its 2017 inception, MODRN created 21 opioid use disorder quality measures in 13 states. MODRN modified its common data model over time to include additional elements. Initial barriers included harmonizing utilization data from Medicaid billing codes across states and adapting statistical methods to combine state-level results. The network demonstrated its utility and addressed barriers to conducting multi-state analyses of Medicaid administrative data. CONCLUSIONS: MODRN created a new, scalable, successful model for conducting policy research while complying with federal and state regulations to protect beneficiary health information. Platforms like MODRN may prove useful for emerging health challenges to facilitate evidence-based policymaking in Medicaid programs.

The bug in a teacup-monitoring arthropod-plant associations with environmental DNA from dried plant material.

Environmental DNA analysis (eDNA) has revolutionized the field of biomonitoring in the past years. Various sources have been shown to contain eDNA of diverse organisms, for example, water, soil, gut content and plant surfaces. Here we show that dried plant material is a highly promising source for arthropod community eDNA. We designed a metabarcoding assay to enrich diverse arthropod communities while preventing amplification of plant DNA. Using this assay, we analysed various commercially produced teas and herbs. These samples recovered ecologically and taxonomically diverse arthropod communities, a total of over a thousand species in more than 20 orders, many of them specific to their host plant and its geographical origin. Atypically for eDNA, arthropod DNA in dried plants shows very high temporal stability, opening up plant archives as a source for historical arthropod eDNA. Considering these results, dried plant material appears excellently suited as a novel tool to monitor arthropods and arthropod-plant interactions, detect agricultural pests and identify the geographical origin of imported plant material. The simplicity of our approach and the ability to detect highly diverse arthropod communities from all over the world in tea bags also highlights its utility for outreach purposes and to raise awareness about biodiversity.

Endometrial Mesonephric-like Adenocarcinoma Presenting as an Ocular Lesion: A Case Report.

Endometrial mesonephric-like carcinoma (ML-CA) is a recently recognized subtype of aggressive endometrial adenocarcinoma that is morphologically and immunophenotypically similar to mesonephric carcinoma but not typically associated with mesonephric remnants. Here, we report a case of 58-yr-old female who had a past medical history of fibroids and of irregular menstrual bleeding for ~20 yr who presented with visual disturbance. On further investigation, she was found to have a large choroidal peri-papillary tumor of the right eye. A presumptive diagnosis of choroidal melanoma was made. Right eye enucleation was performed, and microscopy revealed moderately differentiated metastatic adenocarcinoma. Further work up was advised. A uterine mass was identified on imaging followed by endometrial biopsy that showed a morphologically and immunohistochemically similar tumor to that in the eye. A hysterectomy was carried out and a malignant neoplasm with varying morphologic patterns including gland formation, solid sheets of tumor cells, cribriform, glomeruloid, spindled and papillary areas was seen. The immunohistochemical profile showed diffuse strong positivity for AE1/AE3, TTF1, P16, and vimentin. CD56, GATA3, Napsin A, and CD10 were focally positive. The neoplastic cells were negative for the following markers ER, PR, WT1, calretinin, and synaptophysin. PDL-1 was negative and mismatch repair protein was proficient. An identical KRAS mutation was detected in both the uterine corpus and ocular tumors. The findings are in keeping with a uterine mesonephric-like adenocarcinoma with an ocular metastasis. An Oncomine Focus-Mutation profile, Thermo-Fisher Scientific Inc., a 60 gene oncologic panel, performed on the ocular tumor, revealed no further mutations.

A study of the immune infiltrate and patient outcomes in esophageal cancer.

OBJECTIVES: Cancer patient outcomes and selection for novel therapies are heavily influenced by the immune contexture of the tumor microenvironment. Esophageal cancer is associated with poor outcomes. In contrast to colorectal cancer, where the immunoscore is increasingly used in prognostic staging, little is known about the immune cell populations in esophageal adenocarcinoma (EAC) and esophageal squamous cell carcinoma (SCC), and their clinical significance. METHODS: Tissue microarrays were constructed from resected tumor tissue of 72 EAC patients and 23 SCC patients. Immunohistochemical staining of CD3, CD8, CD56, CD68, CD45RO, CD69, IFN-γ, IL-10, IL-4, IL-17, TGF-ß, FOXP3 and CD107a was performed. Positivity was examined in both the stromal and epithelial compartments. Statistical analysis was performed to identify differences in immune cell infiltration and functional phenotypes between cancer subtypes and tissue compartments. RESULTS: This study identified that esophageal tumors are enriched with CD45RO+ and CD8+ cells and such positivity is significantly higher in SCC compared with EAC. Furthermore, the expression of CD45RO positively correlates with that of CD8 within the tumors of both patient cohorts, suggesting a dominance of memory cytotoxic T cells. This is supported by strong positivity of degranulation marker CD107a in the stromal compartment of EAC and SCC tumors. Cytokine staining revealed a mixed pro- and anti-inflammatory profile within EAC tumors. CONCLUSIONS: Esophageal tumors are enriched with memory cytotoxic T cells. Applying these measurements to a larger cohort will ascertain the clinical utility of assessing specific lymphocyte infiltrates in EAC and SCC tumors with regards to future immunotherapy use, patient prognosis and outcomes.

Apobec1 complementation factor (A1CF) and RBM47 interact in tissue-specific regulation of C to U RNA editing in mouse intestine and liver.

Mammalian C to U RNA is mediated by APOBEC1, the catalytic deaminase, together with RNA binding cofactors (including A1CF and RBM47) whose relative physiological requirements are unresolved. Although A1CF complements APOBEC1 for in vitro RNA editing, A1cf-/- mice exhibited no change in apolipoproteinB (apoB) RNA editing, while Rbm47 mutant mice exhibited impaired intestinal RNA editing of apoB as well as other targets. Here we examined the role of A1CF and RBM47 in adult mouse liver and intestine, following deletion of either one or both gene products and also following forced (liver or intestinal) transgenic A1CF expression. There were minimal changes in hepatic and intestinal apoB RNA editing in A1cf-/- mice and no changes in either liver- or intestine-specific A1CF transgenic mice. Rbm47 liver-specific knockout (Rbm47LKO ) mice demonstrated reduced editing in a subset (11 of 20) of RNA targets, including apoB. By contrast, apoB RNA editing was virtually eliminated (<6% activity) in intestine-specific (Rbm47IKO ) mice with only five of 53 targets exhibiting C-to-U RNA editing. Double knockout of A1cf and Rbm47 in liver (ARLKO ) eliminated apoB RNA editing and reduced editing in the majority of other targets, with no changes following adenoviral APOBEC1 administration. Intestinal double knockout mice (ARIKO ) demonstrated further reduced editing (<10% activity) in four of five of the residual APOBEC1 targets identified in ARIKO mice. These data suggest that A1CF and RBM47 each function independently, yet interact in a tissue-specific manner, to regulate the activity and site selection of APOBEC1 dependent C-to-U RNA editing.

Connecting high-throughput biodiversity inventories: Opportunities for a site-based genomic framework for global integration and synthesis.

High-throughput sequencing (HTS) is increasingly being used for the characterization and monitoring of biodiversity. If applied in a structured way, across broad geographical scales, it offers the potential for a much deeper understanding of global biodiversity through the integration of massive quantities of molecular inventory data generated independently at local, regional and global scales. The universality, reliability and efficiency of HTS data can potentially facilitate the seamless linking of data among species assemblages from different sites, at different hierarchical levels of diversity, for any taxonomic group and regardless of prior taxonomic knowledge. However, collective international efforts are required to optimally exploit the potential of site-based HTS data for global integration and synthesis, efforts that at present are limited to the microbial domain. To contribute to the development of an analogous strategy for the nonmicrobial terrestrial domain, an international symposium entitled "Next Generation Biodiversity Monitoring" was held in November 2019 in Nicosia (Cyprus). The symposium brought together evolutionary geneticists, ecologists and biodiversity scientists involved in diverse regional and global initiatives using HTS as a core tool for biodiversity assessment. In this review, we summarize the consensus that emerged from the 3-day symposium. We converged on the opinion that an effective terrestrial Genomic Observatories network for global biodiversity integration and synthesis should be spatially led and strategically united under the umbrella of the metabarcoding approach. Subsequently, we outline an HTS-based strategy to collectively build an integrative framework for site-based biodiversity data generation.

Use of Medications for Treatment of Opioid Use Disorder Among US Medicaid Enrollees in 11 States, 2014-2018.

Importance: There is limited information about trends in the treatment of opioid use disorder (OUD) among Medicaid enrollees. Objective: To examine the use of medications for OUD and potential indicators of quality of care in multiple states. Design, Setting, and Participants: Exploratory serial cross-sectional study of 1 024 301 Medicaid enrollees in 11 states aged 12 through 64 years (not eligible for Medicare) with International Classification of Diseases, Ninth Revision (ICD-9 or ICD-10) codes for OUD from 2014 through 2018. Each state used generalized estimating equations to estimate associations between enrollee characteristics and outcome measure prevalence, subsequently pooled to generate global estimates using random effects meta-analyses. Exposures: Calendar year, demographic characteristics, eligibility groups, and comorbidities. Main Outcomes and Measures: Use of medications for OUD (buprenorphine, methadone, or naltrexone); potential indicators of good quality (OUD medication continuity for 180 days, behavioral health counseling, urine drug tests); potential indicators of poor quality (prescribing of opioid analgesics and benzodiazepines). Results: In 2018, 41.7% of Medicaid enrollees with OUD were aged 21 through 34 years, 51.2% were female, 76.1% were non-Hispanic White, 50.7% were eligible through Medicaid expansion, and 50.6% had other substance use disorders. Prevalence of OUD increased in these 11 states from 3.3% (290 628 of 8 737 082) in 2014 to 5.0% (527 983 of 10 585 790) in 2018. The pooled prevalence of enrollees with OUD receiving medication treatment increased from 47.8% in 2014 (range across states, 35.3% to 74.5%) to 57.1% in 2018 (range, 45.7% to 71.7%). The overall prevalence of enrollees receiving 180 days of continuous medications for OUD did not significantly change from the 2014-2015 to 2017-2018 periods (-0.01 prevalence difference, 95% CI, -0.03 to 0.02) with state variability in trend (90% prediction interval, -0.08 to 0.06). Non-Hispanic Black enrollees had lower OUD medication use than White enrollees (prevalence ratio [PR], 0.72; 95% CI, 0.64 to 0.81; P < .001; 90% prediction interval, 0.52 to 1.00). Pregnant women had higher use of OUD medications (PR, 1.18; 95% CI, 1.11-1.25; P < .001; 90% prediction interval, 1.01-1.38) and medication continuity (PR, 1.14; 95% CI, 1.10-1.17, P < .001; 90% prediction interval, 1.06-1.22) than did other eligibility groups. Conclusions and Relevance: Among US Medicaid enrollees in 11 states, the prevalence of medication use for treatment of opioid use disorder increased from 2014 through 2018. The pattern in other states requires further research.

High-throughput sequencing for community analysis: the promise of DNA barcoding to uncover diversity, relatedness, abundances and interactions in spider communities.

Large-scale studies on community ecology are highly desirable but often difficult to accomplish due to the considerable investment of time, labor and, money required to characterize richness, abundance, relatedness, and interactions. Nonetheless, such large-scale perspectives are necessary for understanding the composition, dynamics, and resilience of biological communities. Small invertebrates play a central role in ecosystems, occupying critical positions in the food web and performing a broad variety of ecological functions. However, it has been particularly difficult to adequately characterize communities of these animals because of their exceptionally high diversity and abundance. Spiders in particular fulfill key roles as both predator and prey in terrestrial food webs and are hence an important focus of ecological studies. In recent years, large-scale community analyses have benefitted tremendously from advances in DNA barcoding technology. High-throughput sequencing (HTS), particularly DNA metabarcoding, enables community-wide analyses of diversity and interactions at unprecedented scales and at a fraction of the cost that was previously possible. Here, we review the current state of the application of these technologies to the analysis of spider communities. We discuss amplicon-based DNA barcoding and metabarcoding for the analysis of community diversity and molecular gut content analysis for assessing predator-prey relationships. We also highlight applications of the third generation sequencing technology for long read and portable DNA barcoding. We then address the development of theoretical frameworks for community-level studies, and finally highlight critical gaps and future directions for DNA analysis of spider communities.

Identification and clinical impact of potentially actionable somatic oncogenic mutations in solid tumor samples.

BACKGROUND: An increasing number of anti-cancer therapeutic agents target specific mutant proteins that are expressed by many different tumor types. Successful use of these therapies is dependent on the presence or absence of somatic mutations within the patient's tumor that can confer clinical efficacy or drug resistance. METHODS: The aim of our study was to determine the type, frequency, overlap and functional proteomic effects of potentially targetable recurrent somatic hotspot mutations in 47 cancer-related genes in multiple disease sites that could be potential therapeutic targets using currently available agents or agents in clinical development. RESULTS: Using MassArray technology, of the 1300 patient tumors analysed 571 (43.9%) had at least one somatic mutation. Mutations were identified in 30 different genes. KRAS (16.5%), PIK3CA (13.6%) and BRAF (3.8%) were the most frequently mutated genes. Prostate (10.8%) had the lowest number of somatic mutations identified, while no mutations were identified in sarcoma. Ocular melanoma (90.6%), endometrial (72.4%) and colorectal (66.4%) tumors had the highest number of mutations. We noted high concordance between mutations in different parts of the tumor (94%) and matched primary and metastatic samples (90%). KRAS and BRAF mutations were mutually exclusive. Mutation co-occurrence involved mainly PIK3CA and PTPN11, and PTPN11 and APC. Reverse Phase Protein Array (RPPA) analysis demonstrated that PI3K and MAPK signalling pathways were more altered in tumors with mutations compared to wild type tumors. CONCLUSIONS: Hotspot mutational profiling is a sensitive, high-throughput approach for identifying mutations of clinical relevance to molecular based therapeutics for treatment of cancer, and could potentially be of use in identifying novel opportunities for genotype-driven clinical trials.

Impaired Chylomicron Assembly Modifies Hepatic Metabolism Through Bile Acid-Dependent and Transmissible Microbial Adaptations.

The mechanisms by which alterations in intestinal bile acid (BA) metabolism improve systemic glucose tolerance and hepatic metabolic homeostasis are incompletely understood. We examined metabolic adaptations in mice with conditional intestinal deletion of the abetalipoproteinemia (ABL) gene microsomal triglyceride transfer protein (Mttp-IKO), which blocks chylomicron assembly and impairs intestinal lipid transport. Mttp-IKO mice exhibit improved hepatic glucose metabolism and augmented insulin signaling, without weight loss. These adaptations included decreased BA excretion, increased pool size, altered BA composition, and increased fibroblast growth factor 15 production. Mttp-IKO mice absorb fructose normally but are protected against dietary fructose-induced hepatic steatosis, without weight loss or changes in energy expenditure. In addition, Mttp-IKO mice exhibit altered cecal microbial communities, both at baseline and following fructose feeding, including increased abundance of Bacteroides and Lactobacillus genera. Transplantation of cecal microbiota from chow-fed Mttp-IKO mice into antibiotic-treated wild-type recipients conferred transmissible protection against fructose-induced hepatic steatosis in association with a bloom in Akkermansia and increased Clostridium XIVa genera, whose abundance was positively correlated with fecal coprostanol and total neutral sterol excretion in recipient mice. However, antibiotic-treated Mttp-IKO mice were still protected against fructose-induced hepatic steatosis, suggesting that changes in microbiota are not required for this phenotype. Nevertheless, we found increased abundance of fecal Akkermansia from two adult ABL subjects with MTTP mutations compared to their heterozygous parents and within the range noted in six healthy control subjects. Furthermore, Akkermansia abundance across all subjects was positively correlated with fecal coprostanol excretion. Conclusion: The findings collectively suggest multiple adaptive pathways of metabolic regulation following blocked chylomicron assembly, including shifts in BA signaling and altered microbial composition that confer a transmissible phenotype.