Conserved and tissue-specific immune responses to biologic scaffold implantation.

Upon implantation into a patient, any biomaterial induces a cascade of immune responses that influences the outcome of that device. This cascade depends upon several factors, including the composition of the material itself and the location in which the material is implanted. There is still significant uncertainty around the role of different tissue microenvironments in the immune response to biomaterials and how that may alter downstream scaffold remodeling and integration. In this study, we present a study evaluating the immune response to decellularized extracellular matrix materials within the intraperitoneal cavity, the subcutaneous space, and in a traumatic skeletal muscle injury microenvironment. All different locations induced robust cellular recruitment, specifically of macrophages and eosinophils. The latter was most prominent in the subcutaneous space. Intraperitoneal implants uniquely recruited B cells that may alter downstream reactivity as adaptive immunity has been strongly implicated in the outcome of scaffold remodeling. These data suggest that the location of tissue implants should be taken together with the composition of the material itself when designing devices for downline therapeutics. STATEMENT OF SIGNIFICANCE: Different tissue locations have unique immune microenvironments, which can influence the immune response to biomaterial implants. By considering the specific immune profiles of the target tissue, researchers can develop implant materials that promote better integration, reduce complications, and improve the overall outcome of the implantation process.

Surgical Approach Does Not Influence Instability Risk in Revision Total Hip Arthroplasty.

BACKGROUND: Although the direct anterior (DA) approach has increased in popularity for primary total hip arthroplasty (THA), there is limited evidence regarding its use for revision THA. It is unknown whether the dislocation benefit seen in the primary setting translates to revision cases. METHODS: This retrospective review compared the dislocation rates of revision THA performed through DA versus postero-lateral (PL) approaches at a single institution (2011 to 2021). Exclusion criteria included revision for instability, ≥ 2 prior revisions, approaches other than DA or PL, and placement of dual-mobility or constrained liners. There were 182 hips in 173 patients that met the inclusion criteria. The average follow-up was 6.5 years (range, 2 to 8 years). RESULTS: There was a trend toward more both-component revisions being performed through the PL approach. There were no differences in dislocation rates between the DA revision and PL revision cohorts, which were 8.1% (5 of 72) and 7.5% (9 of 120), respectively (P = .999). Dislocation trended lower when the revision approach was discordant from the primary approach compared to cases where primary and revision had a concordant approach (4.9 versus 8.5%), but this was not statistically significant (P = .740). No significant differences were found in return to operating room, 90-day emergency department visits, or 90-day readmissions. However, the length of stay was significantly shorter in patients who had DA revisions after a primary PL procedure (P = .021). CONCLUSIONS: Dislocation rates following revision THA did not differ between the DA and PL approaches irrespective of the primary approach. Surgeons should choose their revision approach based on their experience and the specific needs of the patient.

Mixed methods assessment of the influence of demographics on medical advice of ChatGPT.

OBJECTIVES: To evaluate demographic biases in diagnostic accuracy and health advice between generative artificial intelligence (AI) (ChatGPT GPT-4) and traditional symptom checkers like WebMD. MATERIALS AND METHODS: Combination symptom and demographic vignettes were developed for 27 most common symptom complaints. Standardized prompts, written from a patient perspective, with varying demographic permutations of age, sex, and race/ethnicity were entered into ChatGPT (GPT-4) between July and August 2023. In total, 3 runs of 540 ChatGPT prompts were compared to the corresponding WebMD Symptom Checker output using a mixed-methods approach. In addition to diagnostic correctness, the associated text generated by ChatGPT was analyzed for readability (using Flesch-Kincaid Grade Level) and qualitative aspects like disclaimers and demographic tailoring. RESULTS: ChatGPT matched WebMD in 91% of diagnoses, with a 24% top diagnosis match rate. Diagnostic accuracy was not significantly different across demographic groups, including age, race/ethnicity, and sex. ChatGPT's urgent care recommendations and demographic tailoring were presented significantly more to 75-year-olds versus 25-year-olds (P < .01) but were not statistically different among race/ethnicity and sex groups. The GPT text was suitable for college students, with no significant demographic variability. DISCUSSION: The use of non-health-tailored generative AI, like ChatGPT, for simple symptom-checking functions provides comparable diagnostic accuracy to commercially available symptom checkers and does not demonstrate significant demographic bias in this setting. The text accompanying differential diagnoses, however, suggests demographic tailoring that could potentially introduce bias. CONCLUSION: These results highlight the need for continued rigorous evaluation of AI-driven medical platforms, focusing on demographic biases to ensure equitable care.

Intraoperative Surgeon-Administered Adductor Canal Block Is a Safe Alternative to Preoperative Anesthesiologist-Administered Adductor Canal Block in Primary Total Knee Arthroplasty.

BACKGROUND: The shift toward outpatient total knee arthroplasties (TKAs) has led to a demand for effective perioperative pain control methods. A surgeon-performed "low" adductor canal block ("low-ACB") technique, involving an intraoperative ACB, is gaining popularity due to its efficiency and early pain control potential. This study examined the transition from traditional preoperative anesthesiologist-performed ultrasound-guided adductor canal blocks ("high-ACB") to low-ACB, evaluating pain control, morphine consumption, first physical therapy visit gait distance, hospital length-of-stay, and complications. METHODS: There were 2,620 patients at a single institution who underwent a primary total knee arthroplasty between January 1, 2019, and December 31, 2022, and received either a low-ACB or high-ACB. Cohorts included 1,248 patients and 1,372 patients in the low-ACB and high-ACB groups, respectively. Demographics and operative times were similar. Patient characteristics and outcomes such as morphine milligram equivalents (MMEs), Visual Analog Scale pain scores, gait distance (feet), length of stay (days), and postoperative complications (30-day readmission and 30-day emergency department visit) were collected. RESULTS: The low-ACB cohort had higher pain scores over the first 24 hours (5.05 versus 4.86, P < .001) and higher MME at 6 hours (11.49 versus 8.99, P < .001), although this was not clinically significant. There was no difference in pain scores or MME at 12 or 24 hours (20.81 versus 22.07 and 44.67 versus 48.78, respectively). The low-ACB cohort showed longer gait distance at the first physical therapy visit (188.5 versus 165.1 feet, P < .001) and a shorter length of stay (0.88 versus 1.46 days, P < .01), but these were not clinically significant. There were no differences in 30-day complications. CONCLUSIONS: The low-ACB offers effective pain relief and comparable early recovery without increasing operative time or the complication rate. Low-ACB is an effective, safe, and economical alternative to high-ACB. LEVEL OF EVIDENCE: Therapeutic study, Level III (retrospective cohort study).

Trajectory of external implementation support activities across two states in the United States: A descriptive study.

Background: Reporting on strategies to advance implementation outcomes is imperative. The current study reports descriptive information about external implementation support (EIS) provided over 5 years to 13 regions in North Carolina and South Carolina scaling an evidence-based system of parenting and family supports. Regional support teams operating through the Implementation Capacity for Triple P (ICTP) projects employed core practice components (CPCs) for EIS as proposed by Aldridge et al. and further operationalized by members of The Impact Center at FPG Child Development Institute, UNC-Chapel Hill. Method: Practice activities associated with CPCs were developed and iteratively refined across the study period. ICTP regional support teams systematically tracked their use of CPCs and related activities following each substantive support interaction. Tracking included the duration of time a CPC was employed and the use of specific practice activities associated with that CPC. Data were aggregated by month of the relationship to account for differential start dates across regions. Results: From November 2016 through December 2021, ICTP support teams tracked 749 support interactions with Triple P regions in North Carolina and South Carolina. Monthly support decreased year over year, though dose varied considerably. Patterns of CPC use indicated a high dose of "foundational" and "co-design" CPCs early, followed by a blended and more diverse use thereafter, with some notable trends. Practice activities considered essential to influencing intended practice outcomes were characterized by higher rates of use. Like CPCs, practice activities were used dynamically across the study period. Conclusions: This descriptive study offers a case study for how EIS might be operationalized, tracked, and employed. Findings suggest several interpretations that might refine our understanding and use of EIS. Although the nature of this practical report precludes generalizability of findings, directions for future research and practice are discussed.

More needs to be known about how external support providers work with organizational, system, and community partners to improve their efforts to implement innovative programs and practices. Although a handful of models of external support have emerged in the literature, support strategies or activities are not often tracked systematically or prospectively. In the current study, we report on the adoption, operationalization, and use of a recently proposed model of external support across 13 regions in North Carolina and South Carolina scaling an evidence-based system of parenting and family interventions. Teams of external support providers tracked their activities using the model across more than 5 years. Results provide a description of what external support might look like across long-term support engagements, noting key patterns about dose of support and use of activities that might be responsible for influencing intended support outcomes. The study offers several findings that might refine our understanding and use of external support strategies. In addition, the study lays groundwork for examining additional research questions, such as the feasibility of support processes and whether and how support activities influence support outcomes.

Prophylactic Knee Bracing in Offensive Linemen of the National Football League: A Retrospective Analysis of Usage Trends, Player Performance, and Major Knee Injury.

Background: Offensive linemen in American football are prone to high-energy valgus forces to the knee, leading to associated injuries. Some offensive linemen in the National Football League (NFL) wear prophylactic knee braces (PKB) to prevent ligamentous injury. Purpose/Hypothesis: This purpose of the study was to compare injury rates and performance between NFL offensive linemen who wear PKB and those who do not. It was hypothesized that brace wear would be associated with fewer major knee injuries and no difference in gameplay performance. Study Design: Cohort study; Level of evidence, 3. Methods: For the 2014 through 2020 NFL seasons, offensive linemen with at least 200 game snaps per regular season were identified. Players were grouped by PKB status (bracers vs nonbracers) based on visualization of bilateral, dual-hinged metal knee braces as part of gameday uniforms on publicly available imaging databases and/or game videos. Major knee injuries, defined as those requiring the missing of games, were identified using publicly available data. Performance was assessed with Pro Football Focus grades for each season. Rates of major knee injury were compared between groups with the 2-sample Z test for proportions, and performance grades were compared with the unpaired t test. Results: For the cumulative study period, bracers demonstrated a significantly lower rate of major knee injuries than nonbracers (0.013 vs 0.049 injuries per player, respectively; P = .04). Isolated MCL injury was the most common injury for nonbracers. There was no group difference in performance for the cumulative study period or during most individual seasons. Yearly prevalence of PKB usage declined steadily from 16.3% in 2014 to 5.6% in 2020. A subgroup analysis of rookie players demonstrated an overall downtrend in usage during the study period as well. Conclusion: Results indicated that knee brace prophylaxis by NFL offensive linemen was associated with a reduced risk of major knee injury without a significant difference in performance when compared with nonbracers. Despite this, the prevalence of PKB declined over the study period.

Green Turtle Fibropapillomatosis: Tumor Morphology and Growth Rate in a Rehabilitation Setting.

Fibropapillomatosis (FP) is a neoplastic disease most often found in green turtles (Chelonia mydas). Afflicted turtles are burdened with potentially debilitating tumors concentrated externally on the soft tissues, plastron, and eyes and internally on the lungs, kidneys, and the heart. Clinical signs occur at various levels, ranging from mild disease to severe debilitation. Tumors can both progress and regress in affected turtles, with outcomes ranging from death due to the disease to complete regression. Since its official description in the scientific literature in 1938, tumor growth rates have been rarely documented. In addition, FP tumors come in two very different morphologies; yet, to our knowledge, there have been no quantified differences in growth rates between tumor types. FP tumors are often rugose in texture, with a polypoid to papillomatous morphology, and may or may not be pedunculated. In other cases, tumors are smooth, with a skin-like surface texture and little to no papillose structures. In our study, we assessed growth-rate differences between rugose and smooth tumor morphologies in a rehabilitation setting. We measured average biweekly tumor growth over time in green turtles undergoing rehabilitation at the University of Florida Whitney Laboratory Sea Turtle Hospital in St. Augustine, Florida, and compared growth between rugose and smooth tumors. Our results demonstrate that both rugose and smooth tumors follow a similar active growth progression pattern, but rugose tumors grew at significantly faster rates (p = 0.013) than smooth ones. We also documented regression across several examined tumors, ranging from -0.19% up to -10.8% average biweekly negative growth. Our study offers a first-ever assessment of differential growth between tumor morphologies and an additional diagnostic feature that may lead to a more comprehensive understanding and treatment of the disease. We support the importance of tumor morphological categorization (rugose versus smooth) being documented in future FP hospital- and field-based health assessments.

Underscoring the effect of swab type, workflow, and positive sample order on swab pooling for COVID-19 surveillance testing.

Sample pooling is a promising strategy to facilitate COVID-19 surveillance testing for a larger population in comparison to individual single testing due to resource and time constraints. Increased surveillance testing capacity will reduce the likelihood of outbreaks as the general population is returning to work, school, and other gatherings. We have analyzed the impact of three variables on the effectiveness of pooling test samples: swab type, workflow, and positive sample order. We investigated the performance of several commercially available swabs (Steripack polyester flocked, Puritan nylon flocked, Puritan foam) in comparison to a new injected molded design (Yukon). The bench-top performance of collection swab was conducted with a previously developed anterior nasal cavity tissue model, based on a silk-glycerol sponge to mimic soft tissue mechanics and saturated with a physiologically relevant synthetic nasal fluid spiked with heat-inactivated SARS-CoV-2. Overall, we demonstrated statistically significant differences in performance across the different swab types. A characterization of individual swab uptake (gravimetric analysis) and FITC microparticle release suggests that differences in absorbance and retention drive the observed differences in Ct of the pooled samples. We also proposed two distinct pooling workflows to encompass different community collection modes and analyzed the difference in resulting positive pools as an effect of workflow, swab type, and positive sample order. Overall, swab types with lower volume retention resulted in reduced false negative occurrence, also observed for collection workflows with limited incubation times. Concurrently, positive sample order did have a significant impact on pooling test outcome, particularly in the case of swab type with great volume retention. We demonstrated that the variables investigated here affect the results of pooled COVID-19 testing, and therefore should be considered while designing pooled surveillance testing.

Meta-analysis reveals between-population differences affect the link between glucocorticoids and population health.

Glucocorticoids are a popular tool for monitoring health of animal populations because they can increase with environmental stressors and can indicate chronic stress. However, individual responses to stressors create variation in the glucocorticoid-fitness relationship within populations. The inconsistency in this relationship calls into question the widespread use of glucocorticoids in conservation. We investigated the sources of variation in the glucocorticoid-fitness relationship by conducting a meta-analysis across a diverse set of species exposed to conservation-relevant stressors. We first quantified the extent to which studies inferred population health from glucocorticoids without first validating the glucocorticoid-fitness relationship in their own populations. We also tested whether population-level information like life history stage, sex and species longevity influenced the relationship between glucocorticoids and fitness. Finally, we tested for a universally consistent relationship between glucocorticoids and fitness across studies. We found more than half of peer-reviewed studies published between 2008 and 2022 inferred population health solely based on glucocorticoid levels. While life history stage explained some variation in the relationship between glucocorticoids and fitness, we found no consistent relationship between them. Much of the variation in the relationship could be the result of idiosyncratic characteristics of declining populations, such as unstable demographic structure, that coincided with large amounts of variation in glucocorticoid production. We suggest that conservation biologists capitalize on this variation in glucocorticoid production by declining populations by using the variance in glucocorticoid production as an early warning for declines in population health.

Reciprocal Associations Between Language Ability and Social Functioning Development in Pre-verbal Autistic Children.

Longitudinal research on language abilities and social functioning in young children suggests that gains in one domain affect gains in the other. However, few studies have examined inter-relations of language and social functioning jointly among young children diagnosed with autism spectrum disorder (ASD). Pre-verbal toddlers with ASD are a group of particular clinical relevance, given that greater language abilities at school entry have been associated with positive long-term adjustment in many areas, including adaptive and social functioning. Reduced attention to and engagement in social interactions among autistic toddlers who are not yet speaking may interfere with language development concurrently and over time. The present study examined reciprocal associations between language ability and social functioning over a 2-year period across three time points in a sample of 90 pre-verbal autistic toddlers using cross-lagged panel analyses conducted in MPlus. Cross-lagged panel analyses revealed significant within-timepoint synchronous correlations, within-domain autoregressive paths over time, and as hypothesized, reciprocal significance in all cross-lagged paths. For very young pre-verbal children with ASD, language ability and social functioning appear to exert concurrent and cascading developmental influences on one another. Targeting both language and social functioning simultaneously may enhance intervention efficacy for very young pre-verbal children with ASD.

Solvent Control of Chemical Identity Can Change Photodissociation into Photoisomerization.

In solution-phase chemistry, the solvent is often considered to be merely a medium that allows reacting solutes to encounter each other. In this work, however, we show that moderate locally specific solute-solvent interactions can affect not only the nature of the solute but also the types of reactive chemistry. We use quantum simulation methods to explore how solvent participation in solute chemical identity alters reactions involving the breaking of chemical bonds. In particular, we explore the photoexcitation dynamics of Na2+ dissolved in liquid tetrahydrofuran. In the gas phase, excitation of Na2+ directly leads to dissociation, but in solution, photoexcitation leads to an isomerization reaction involving rearrangement of the first-shell solvent molecules; this isomerization must go to completion before the solute can dissociate. Despite the complexity, the solution-phase reaction dynamics can be captured by a two-dimensional energy surface where one dimension involves only the isomerization of the first-shell solvent molecules.

In Vitro Nasal Tissue Model for the Validation of Nasopharyngeal and Midturbinate Swabs for SARS-CoV-2 Testing.

Large-scale population testing is a key tool to mitigate the spread of respiratory pathogens, such as the current COVID-19 pandemic, where swabs are used to collect samples in the upper airways (e.g., nasopharyngeal and midturbinate nasal cavities) for diagnostics. However, the high volume of supplies required to achieve large-scale population testing has posed unprecedented challenges for swab manufacturing and distribution, resulting in a global shortage that has heavily impacted testing capacity worldwide and prompted the development of new swabs suitable for large-scale production. Newly designed swabs require rigorous preclinical and clinical validation studies that are costly and time-consuming (i.e., months to years long); reducing the risks associated with swab validation is therefore paramount for their rapid deployment. To address these shortages, we developed a 3D-printed tissue model that mimics the nasopharyngeal and midturbinate nasal cavities, and we validated its use as a new tool to rapidly test swab performance. In addition to the nasal architecture, the tissue model mimics the soft nasal tissue with a silk-based sponge lining, and the physiological nasal fluid with asymptomatic and symptomatic viscosities of synthetic mucus. We performed several assays comparing standard flocked and injection-molded swabs. We quantified the swab pickup and release and determined the effect of viral load and mucus viscosity on swab efficacy by spiking the synthetic mucus with heat-inactivated SARS-CoV-2 virus. By molecular assay, we found that injected molded swabs performed similarly or superiorly in comparison to standard flocked swabs, and we underscored a viscosity-dependent difference in cycle threshold values between the asymptomatic and symptomatic mucuses for both swabs. To conclude, we developed an in vitro nasal tissue model that corroborated previous swab performance data from clinical studies; this model will provide to researchers a clinically relevant, reproducible, safe, and cost-effective validation tool for the rapid development of newly designed swabs.

Detection and population genomics of sea turtle species via noninvasive environmental DNA analysis of nesting beach sand tracks and oceanic water.

Elusive aquatic wildlife, such as endangered sea turtles, are difficult to monitor and conserve. As novel molecular and genetic technologies develop, it is possible to adapt and optimize them for wildlife conservation. One such technology is environmental (e)DNA - the detection of DNA shed from organisms into their surrounding environments. We developed species-specific green (Chelonia mydas) and loggerhead (Caretta caretta) sea turtle probe-based qPCR assays, which can detect and quantify sea turtle eDNA in controlled (captive tank water and sand samples) and free ranging (oceanic water samples and nesting beach sand) settings. eDNA detection complemented traditional in-water sea turtle monitoring by enabling detection even when turtles were not visually observed. Furthermore, we report that high throughput shotgun sequencing of eDNA sand samples enabled sea turtle population genetic studies and pathogen monitoring, demonstrating that noninvasive eDNA techniques are viable and efficient alternatives to biological sampling (e.g., biopsies and blood draws). Genetic information was obtained from sand many hours after nesting events, without having to observe or interact with the target individual. This greatly reduces the sampling stress experienced by nesting mothers and emerging hatchlings, and avoids sacrificing viable eggs for genetic analysis. The detection of pathogens from sand indicates significant potential for increased wildlife disease monitoring capacity and viral variant surveillance. Together, these results demonstrate the potential of eDNA approaches to ultimately help understand and conserve threatened species such as sea turtles.

The 2021 European Alliance of Associations for Rheumatology/American College of Rheumatology Points to Consider for Diagnosis and Management of Autoinflammatory Type I Interferonopathies: CANDLE/PRAAS, SAVI, and AGS.

OBJECTIVE: Autoinflammatory type I interferonopathies, chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature/proteasome-associated autoinflammatory syndrome (CANDLE/PRAAS), stimulator of interferon genes (STING)-associated vasculopathy with onset in infancy (SAVI), and Aicardi-Goutières syndrome (AGS) are rare and clinically complex immunodysregulatory diseases. With emerging knowledge of genetic causes and targeted treatments, a Task Force was charged with the development of "points to consider" to improve diagnosis, treatment, and long-term monitoring of patients with these rare diseases. METHODS: Members of a Task Force consisting of rheumatologists, neurologists, an immunologist, geneticists, patient advocates, and an allied health care professional formulated research questions for a systematic literature review. Then, based on literature, Delphi questionnaires, and consensus methodology, "points to consider" to guide patient management were developed. RESULTS: The Task Force devised consensus and evidence-based guidance of 4 overarching principles and 17 points to consider regarding the diagnosis, treatment, and long-term monitoring of patients with the autoinflammatory interferonopathies, CANDLE/PRAAS, SAVI, and AGS. CONCLUSION: These points to consider represent state-of-the-art knowledge to guide diagnostic evaluation, treatment, and management of patients with CANDLE/PRAAS, SAVI, and AGS and aim to standardize and improve care, quality of life, and disease outcomes.

Preclinical Validation of a Novel Injection-Molded Swab for the Molecular Assay Detection of SARS-CoV-2.

During the COVID-19 public health emergency, many actions have been undertaken to help ensure that patients and health care providers have timely and continued access to high-quality medical devices to respond effectively. The development and validation of new testing supplies and equipment, including collection swabs, has helped to expand the availability and capability for various diagnostic, therapeutic, and protective medical devices in high demand during the COVID-19 emergency. Here, we report the initial validation of a new injection-molded anterior nasal swab, ClearTip™, that was experimentally validated in a laboratory setting as well as in independent clinical studies in comparison to gold standard flocked swabs. We have also developed an in vitro anterior nasal tissue model which offers a novel, efficient, and clinically relevant validation tool to replicate the clinical swabbing workflow with high fidelity, while being accessible, safe, reproducible, and time- and cost-effective. ClearTip™ displayed greater inactivated virus release in the benchtop model, confirmed by its greater ability to report positive samples in a small clinical study in comparison to flocked swabs. We also quantified the detection of biological materials, as a proxy for viral material, in multi-center pre-clinical and clinical studies which showed a statistically significant difference in one study and a reduction in performance in comparison to flocked swabs. Taken together, these results emphasize the compelling benefits of non-absorbent injection-molded anterior nasal swabs for COVID-19 detection, comparable to standard flocked swabs. Injection-molded swabs, as ClearTip™, could have the potential to support future swab shortages, due to its manufacturing advantages, while offering benefits in comparison to highly absorbent swabs in terms of comfort, limited volume collection, and potential multiple usage.

The 2021 European Alliance of Associations for Rheumatology/American College of Rheumatology points to consider for diagnosis and management of autoinflammatory type I interferonopathies: CANDLE/PRAAS, SAVI and AGS.

OBJECTIVE: Autoinflammatory type I interferonopathies, chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature/proteasome-associated autoinflammatory syndrome (CANDLE/PRAAS), stimulator of interferon genes (STING)-associated vasculopathy with onset in infancy (SAVI) and Aicardi-Goutières syndrome (AGS) are rare and clinically complex immunodysregulatory diseases. With emerging knowledge of genetic causes and targeted treatments, a Task Force was charged with the development of 'points to consider' to improve diagnosis, treatment and long-term monitoring of patients with these rare diseases. METHODS: Members of a Task Force consisting of rheumatologists, neurologists, an immunologist, geneticists, patient advocates and an allied healthcare professional formulated research questions for a systematic literature review. Then, based on literature, Delphi questionnaires and consensus methodology, 'points to consider' to guide patient management were developed. RESULTS: The Task Force devised consensus and evidence-based guidance of 4 overarching principles and 17 points to consider regarding the diagnosis, treatment and long-term monitoring of patients with the autoinflammatory interferonopathies, CANDLE/PRAAS, SAVI and AGS. CONCLUSION: These points to consider represent state-of-the-art knowledge to guide diagnostic evaluation, treatment and management of patients with CANDLE/PRAAS, SAVI and AGS and aim to standardise and improve care, quality of life and disease outcomes.

RESCRIPt: Reproducible sequence taxonomy reference database management.

Nucleotide sequence and taxonomy reference databases are critical resources for widespread applications including marker-gene and metagenome sequencing for microbiome analysis, diet metabarcoding, and environmental DNA (eDNA) surveys. Reproducibly generating, managing, using, and evaluating nucleotide sequence and taxonomy reference databases creates a significant bottleneck for researchers aiming to generate custom sequence databases. Furthermore, database composition drastically influences results, and lack of standardization limits cross-study comparisons. To address these challenges, we developed RESCRIPt, a Python 3 software package and QIIME 2 plugin for reproducible generation and management of reference sequence taxonomy databases, including dedicated functions that streamline creating databases from popular sources, and functions for evaluating, comparing, and interactively exploring qualitative and quantitative characteristics across reference databases. To highlight the breadth and capabilities of RESCRIPt, we provide several examples for working with popular databases for microbiome profiling (SILVA, Greengenes, NCBI-RefSeq, GTDB), eDNA and diet metabarcoding surveys (BOLD, GenBank), as well as for genome comparison. We show that bigger is not always better, and reference databases with standardized taxonomies and those that focus on type strains have quantitative advantages, though may not be appropriate for all use cases. Most databases appear to benefit from some curation (quality filtering), though sequence clustering appears detrimental to database quality. Finally, we demonstrate the breadth and extensibility of RESCRIPt for reproducible workflows with a comparison of global hepatitis genomes. RESCRIPt provides tools to democratize the process of reference database acquisition and management, enabling researchers to reproducibly and transparently create reference materials for diverse research applications. RESCRIPt is released under a permissive BSD-3 license at https://github.com/bokulich-lab/RESCRIPt.

In Vitro Nasal Tissue Model for the Validation of Nasopharyngeal and Mid-turbinate Swabs for SARS-CoV-2 Testing.

Large-scale population testing is a key tool to mitigate the spread of respiratory pathogens, as in the current COVID-19 pandemic, where swabs are used to collect samples in the upper airways (e.g. nasopharyngeal and mid-turbinate nasal cavities) for diagnostics. However, the high volume of supplies required to achieve large-scale population testing has posed unprecedented challenges for swab manufacturing and distribution, resulting in a global shortage that has heavily impacted testing capacity world-wide and prompted the development of new swabs suitable for large-scale production. Newly designed swabs require rigorous pre-clinical and clinical validation studies that are costly and time consuming ( i . e . months to years long); reducing the risks associated with swab validation is therefore paramount for their rapid deployment. To address these shortages, we developed a 3D-printed tissue model that mimics the nasopharyngeal and mid-turbinate nasal cavities, and we validated its use as a new tool to rapidly test swab performance. In addition to the nasal architecture, the tissue model mimics the soft nasal tissue with a silk-based sponge lining, and the physiological nasal fluid with asymptomatic and symptomatic viscosities of synthetic mucus. We performed several assays comparing standard flocked and injection-molded swabs. We quantified the swab pick-up and release, and determined the effect of viral load and mucus viscosity on swab efficacy by spiking the synthetic mucus with heat-inactivated SARS-CoV-2 virus. By molecular assays, we found that injected molded swabs performed similarly or superiorly in comparison to standard flocked swabs and we underscored a viscosity-dependent difference in cycle threshold values between the asymptomatic and symptomatic mucus for both swabs. To conclude, we developed an in vitro nasal tissue model, that corroborated previous swab performance data from clinical studies, with the potential of providing researchers with a clinically relevant, reproducible, safe, and cost-effective validation tool for the rapid development of newly designed swabs.

In Vitro Nasal Tissue Model for the Validation of Nasopharyngeal and Mid-turbinate Swabs for SARS-CoV-2 Testing

Large-scale population testing is a key tool to mitigate the spread of respiratory pathogens, as in the current COVID-19 pandemic, where swabs are used to collect samples in the upper airways (e.g. nasopharyngeal and mid-turbinate nasal cavities) for diagnostics. However, the high volume of supplies required to achieve large-scale population testing has posed unprecedented challenges for swab manufacturing and distribution, resulting in a global shortage that has heavily impacted testing capacity world-wide and prompted the development of new swabs suitable for large-scale production. Newly designed swabs require rigorous pre-clinical and clinical validation studies that are costly and time consuming (i.e. months to years long); reducing the risks associated with swab validation is therefore paramount for their rapid deployment. To address these shortages, we developed a 3D-printed tissue model that mimics the nasopharyngeal and mid-turbinate nasal cavities, and we validated its use as a new tool to rapidly test swab performance. In addition to the nasal architecture, the tissue model mimics the soft nasal tissue with a silk-based sponge lining, and the physiological nasal fluid with asymptomatic and symptomatic viscosities of synthetic mucus. We performed several assays comparing standard flocked and injection-molded swabs. We quantified the swab pick-up and release, and determined the effect of viral load and mucus viscosity on swab efficacy by spiking the synthetic mucus with heat-inactivated SARS-CoV-2 virus. By molecular assays, we found that injected molded swabs performed similarly or superiorly in comparison to standard flocked swabs and we underscored a viscosity-dependent difference in cycle threshold values between the asymptomatic and symptomatic mucus for both swabs. To conclude, we developed an in vitro nasal tissue model, that corroborated previous swab performance data from clinical studies, with the potential of providing researchers with a clinically relevant, reproducible, safe, and cost-effective validation tool for the rapid development of newly designed swabs.

The KRAS-regulated kinome identifies WEE1 and ERK coinhibition as a potential therapeutic strategy in KRAS-mutant pancreatic cancer.

Oncogenic KRAS drives cancer growth by activating diverse signaling networks, not all of which have been fully delineated. We set out to establish a system-wide profile of the KRAS-regulated kinase signaling network (kinome) in KRAS-mutant pancreatic ductal adenocarcinoma (PDAC). We knocked down KRAS expression in a panel of six cell lines and then applied multiplexed inhibitor bead/MS to monitor changes in kinase activity and/or expression. We hypothesized that depletion of KRAS would result in downregulation of kinases required for KRAS-mediated transformation and in upregulation of other kinases that could potentially compensate for the deleterious consequences of the loss of KRAS. We identified 15 upregulated and 13 downregulated kinases in common across the panel of cell lines. In agreement with our hypothesis, all 15 of the upregulated kinases have established roles as cancer drivers (e.g., SRC, TGF-ß1, ILK), and pharmacological inhibition of one of these upregulated kinases, DDR1, suppressed PDAC growth. Interestingly, 11 of the 13 downregulated kinases have established driver roles in cell cycle progression, particularly in mitosis (e.g., WEE1, Aurora A, PLK1). Consistent with a crucial role for the downregulated kinases in promoting KRAS-driven proliferation, we found that pharmacological inhibition of WEE1 also suppressed PDAC growth. The unexpected paradoxical activation of ERK upon WEE1 inhibition led us to inhibit both WEE1 and ERK concurrently, which caused further potent growth suppression and enhanced apoptotic death compared with WEE1 inhibition alone. We conclude that system-wide delineation of the KRAS-regulated kinome can identify potential therapeutic targets for KRAS-mutant pancreatic cancer.