Exploring Iodide and Hydrogen Sulfide as ROS Scavengers to Delay Acute Rejection in MHC-Defined Vascularized Composite Allografts.

Vascularized composite allografts (VCA) face ischemic challenges due to their limited availability. Reperfusion following ischemia triggers oxidative stress and immune reactions, and scavenger molecules could mitigate ischemia-reperfusion injuries and, therefore, immune rejection. We compared two scavengers in a myocutaneous flap VCA model. In total, 18 myocutaneous flap transplants were performed in Major histocompatibility complex (MHC)-defined miniature swine. In the MATCH group (n = 9), donors and recipients had minor antigen mismatch, while the animals were fully mismatched in the MISMATCH group (n = 9). Grafts were pretreated with saline, sodium iodide (NaI), or hydrogen sulfide (H2S), stored at 4 °C for 3 h, and then transplanted. Flaps were monitored until clinical rejection without immunosuppression. In the MATCH group, flap survival did not significantly differ between the saline and hydrogen sulfide treatments (p = 0.483) but was reduced with the sodium iodide treatment (p = 0.007). In the MISMATCH group, survival was similar between the saline and hydrogen sulfide treatments (p = 0.483) but decreased with the sodium iodide treatment (p = 0.007). Rhabdomyolysis markers showed lower but non-significant levels in the experimental subgroups for both the MATCH and MISMATCH animals. This study provides insightful data for the field of antioxidant-based approaches in VCA and transplantation.

Reference genome of Townsend's big-eared bat, Corynorhinus townsendii.

Townsend's big-eared bat, Corynorhinus townsendii, is a cave- and mine-roosting species found largely in western North America. Considered a species of conservation concern throughout much of its range, protection efforts would greatly benefit from understanding patterns of population structure, genetic diversity, and local adaptation. To facilitate such research, we present the first de novo genome assembly of C. townsendii as part of the California Conservation Genomics Project (CCGP). Pacific Biosciences HiFi long reads and Omni-C chromatin-proximity sequencing technologies were used to produce a de novo genome assembly, consistent with the standard CCGP reference genome protocol. This assembly comprises 391 scaffolds spanning 2.1 Gb, represented by a scaffold N50 of 174.6 Mb, a contig N50 of 23.4 Mb, and a benchmarking universal single-copy ortholog (BUSCO) completeness score of 96.6%. This high-quality genome will be a key tool for informed conservation and management of this vulnerable species in California and across its range.

The Oxford Needle Experience (ONE) scale: a UK-based and US-based online mixed-methods psychometric development and validation study of an instrument to assess needle fear, attitudes and expectations in the general public.

OBJECTIVES: To develop and validate the Oxford Needle Experience (ONE) scale, an instrument to assess needle fear, attitudes and expectations in the general population. DESIGN: Cross-sectional validation study. SETTING: Internet-based with participants in the UK and USA. PARTICIPANTS: UK and US representative samples stratified by age, sex, and ethnicity using the Prolific Academic platform. MAIN OUTCOME MEASURES: Exploratory factor analysis with categorical variables and a polychoric correlation matrix followed by promax oblique rotation on the UK sample for the ONE scale. Confirmatory factor analysis (CFA) with a Satorra-Bentler scaled test statistic evaluating the root mean squared error of approximation (RMSEA), standardised root mean squared residual (SRMR) and comparative fit index (CFI) on the US sample. Reliability as internal consistency using McDonald's omega. Convergent validity using the Pearson correlation coefficient. Predictive and discriminant validity using logistic regression ORs of association (OR). RESULTS: The population included 1000 respondents, 500 in the UK and 500 in the USA. Minimum average partial correlation and a scree plot suggested four factors should be retained: injection hesitancy, blood-related hesitancy, recalled negative experiences and perceived benefits, yielding a 19-question scale. On CFA, the RMSEA was 0.070 (90% CI, 0.064 to 0.077), SRMR 0.053 and CFI 0.925. McDonald's omega was 0.92 and 0.93 in the UK and US samples, respectively. Convergent validity with the four-item Oxford Coronavirus Explanations, Attitudes and Narratives Survey (OCEANS) needle fear scale demonstrated a strong correlation (r=0.83). Predictive validity with a single-question COVID-19 vaccination status question demonstrated a strong association, OR (95% CI) 0.97 (0.96 to 0.98), p<0.0001 in the US sample. Discriminant validity with a question regarding the importance of controlling what enters the body confirmed the ONE score does not predict this unrelated outcome, OR 1.00 (0.99, 1.01), p=0.996 in the US sample. CONCLUSIONS: The ONE scale is a reliable and valid multidimensional scale that may be useful in predicting vaccine hesitancy, designing public health interventions to improve vaccine uptake and exploring alternatives to needles for medical procedures.

Efforts to enhance reproducibility in a human performance research project.

Background: Ensuring the validity of results from funded programs is a critical concern for agencies that sponsor biological research. In recent years, the open science movement has sought to promote reproducibility by encouraging sharing not only of finished manuscripts but also of data and code supporting their findings. While these innovations have lent support to third-party efforts to replicate calculations underlying key results in the scientific literature, fields of inquiry where privacy considerations or other sensitivities preclude the broad distribution of raw data or analysis may require a more targeted approach to promote the quality of research output. Methods: We describe efforts oriented toward this goal that were implemented in one human performance research program, Measuring Biological Aptitude, organized by the Defense Advanced Research Project Agency's Biological Technologies Office. Our team implemented a four-pronged independent verification and validation (IV&V) strategy including 1) a centralized data storage and exchange platform, 2) quality assurance and quality control (QA/QC) of data collection, 3) test and evaluation of performer models, and 4) an archival software and data repository. Results: Our IV&V plan was carried out with assistance from both the funding agency and participating teams of researchers. QA/QC of data acquisition aided in process improvement and the flagging of experimental errors. Holdout validation set tests provided an independent gauge of model performance. Conclusions: In circumstances that do not support a fully open approach to scientific criticism, standing up independent teams to cross-check and validate the results generated by primary investigators can be an important tool to promote reproducibility of results.

Predicting habitat suitability for Townsend's big-eared bats across California in relation to climate change.

Effective management decisions depend on knowledge of species distribution and habitat use. Maps generated from species distribution models are important in predicting previously unknown occurrences of protected species. However, if populations are seasonally dynamic or locally adapted, failing to consider population level differences could lead to erroneous determinations of occurrence probability and ineffective management. The study goal was to model the distribution of a species of special concern, Townsend's big-eared bats (Corynorhinus townsendii), in California. We incorporate seasonal and spatial differences to estimate the distribution under current and future climate conditions. We built species distribution models using all records from statewide roost surveys and by subsetting data to seasonal colonies, representing different phenological stages, and to Environmental Protection Agency Level III Ecoregions to understand how environmental needs vary based on these factors. We projected species' distribution for 2061-2080 in response to low and high emissions scenarios and calculated the expected range shifts. The estimated distribution differed between the combined (full dataset) and phenologically explicit models, while ecoregion-specific models were largely congruent with the combined model. Across the majority of models, precipitation was the most important variable predicting the presence of C. townsendii roosts. Under future climate scenarios, distribution of C. townsendii is expected to contract throughout the state, however suitable areas will expand within some ecoregions. Comparison of phenologically explicit models with combined models indicates the combined models better predict the extent of the known range of C. townsendii in California. However, life-history-explicit models aid in understanding of different environmental needs and distribution of their major phenological stages. Differences between ecoregion-specific and statewide predictions of habitat contractions highlight the need to consider regional variation when forecasting species' responses to climate change. These models can aid in directing seasonally explicit surveys and predicting regions most vulnerable under future climate conditions.

Evaluation of co-circulating pathogens and microbiome from COVID-19 infections.

Co-infections or secondary infections with SARS-CoV-2 have the potential to affect disease severity and morbidity. Additionally, the potential influence of the nasal microbiome on COVID-19 illness is not well understood. In this study, we analyzed 203 residual samples, originally submitted for SARS-CoV-2 testing, for the presence of viral, bacterial, and fungal pathogens and non-pathogens using a comprehensive microarray technology, the Lawrence Livermore Microbial Detection Array (LLMDA). Eighty-seven percent of the samples were nasopharyngeal samples, and 23% of the samples were oral, nasal and oral pharyngeal swabs. We conducted bioinformatics analyses to examine differences in microbial populations of these samples, as a proxy for the nasal and oral microbiome, from SARS-CoV-2 positive and negative specimens. We found 91% concordance with the LLMDA relative to a diagnostic RT-qPCR assay for detection of SARS-CoV-2. Sixteen percent of all the samples (32/203) revealed the presence of an opportunistic bacterial or frank viral pathogen with the potential to cause co-infections. The two most detected bacteria, Streptococcus pyogenes and Streptococcus pneumoniae, were present in both SARS-CoV-2 positive and negative samples. Human metapneumovirus was the most prevalent viral pathogen in the SARS-CoV-2 negative samples. Sequence analysis of 16S rRNA was also conducted to evaluate bacterial diversity and confirm LLMDA results.

Metagenomic features of bioburden serve as outcome indicators in combat extremity wounds.

Battlefield injury management requires specialized care, and wound infection is a frequent complication. Challenges related to characterizing relevant pathogens further complicates treatment. Applying metagenomics to wounds offers a comprehensive path toward assessing microbial genomic fingerprints and could indicate prognostic variables for future decision support tools. Wound specimens from combat-injured U.S. service members, obtained during surgical debridements before delayed wound closure, were subjected to whole metagenome analysis and targeted enrichment of antimicrobial resistance genes. Results did not indicate a singular, common microbial metagenomic profile for wound failure, instead reflecting a complex microenvironment with varying bioburden diversity across outcomes. Genus-level Pseudomonas detection was associated with wound failure at all surgeries. A logistic regression model was fit to the presence and absence of antimicrobial resistance classes to assess associations with nosocomial pathogens. A. baumannii detection was associated with detection of genomic signatures for resistance to trimethoprim, aminoglycosides, bacitracin, and polymyxin. Machine learning classifiers were applied to identify wound and microbial variables associated with outcome. Feature importance rankings averaged across models indicated the variables with the largest effects on predicting wound outcome, including an increase in P. putida sequence reads. These results describe the microbial genomic determinants in combat wound bioburden and demonstrate metagenomic investigation as a comprehensive tool for providing information toward aiding treatment of combat-related injuries.

Monitoring GAPDH activity and inhibition with cysteine-reactive chemical probes.

Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a central enzyme in glycolysis that regulates the Warburg effect in cancer cells. In addition to its role in metabolism, GAPDH is also implicated in diverse cellular processes, including transcription and apoptosis. Dysregulated GAPDH activity is associated with a variety of pathologies, and GAPDH inhibitors have demonstrated therapeutic potential as anticancer and immunomodulatory agents. Given the critical role of GAPDH in pathophysiology, it is important to have access to tools that enable rapid monitoring of GAPDH activity and inhibition within a complex biological system. Here, we report an electrophilic peptide-based probe, SEC1, which covalently modifies the active-site cysteine, C152, of GAPDH to directly report on GAPDH activity within a proteome. We demonstrate the utility of SEC1 to assess changes in GAPDH activity in response to oncogenic transformation, reactive oxygen species (ROS) and small-molecule GAPDH inhibitors, including Koningic acid (KA). We then further evaluated KA, to determine the detailed mechanism of inhibition. Our mechanistic studies confirm that KA is a highly effective irreversible inhibitor of GAPDH, which acts through a NAD+-uncompetitive and G3P-competitive mechanism. Proteome-wide evaluation of the cysteine targets of KA demonstrated high selectivity for the active-site cysteine of GAPDH over other reactive cysteines within the proteome. Lastly, the therapeutic potential of KA was investigated in an autoimmune model, where treatment with KA resulted in decreased cytokine production by Th1 effector cells. Together, these studies describe methods to evaluate GAPDH activity and inhibition within a proteome, and report on the high potency and selectivity of KA as an irreversible inhibitor of GAPDH.

Biomodifying the 'natural': from Adaptive Regulation to Adaptive Societal Governance.

Biomodifying technologies-such as gene editing, induced pluripotent stem cells, and bioprinting-are being developed for a wide range of applications, from pest control to lab-grown meat. In medicine, regulators have responded to the challenge of evaluating modified 'natural' material as a therapeutic 'product' by introducing more flexible assessment schemes. Attempts have also been made to engage stakeholders across the globe on the acceptable parameters for these technologies, particularly in the case of gene editing. Regulatory flexibility and stakeholder engagement are important, but a broader perspective is also needed to respond to the potential disruption of biomodification. Our case-study technologies problematize basic ideas-such as 'nature', 'product', and 'donation'-that underpin the legal categories used to regulate biotechnology. Where such foundational concepts are rendered uncertain, a socially responsive and sustainable solution would involve exploring evolutions in these concepts across different societies. We suggest that the global observatory model is a good starting point for this 'Adaptive Societal Governance' approach, in which a self-organizing network of scholars and interested parties could carry out the multi-modal (meta)analyses needed to understand societal constructions of ideas inherent to our understanding of 'life'.

Microbial Tracking-2, a metagenomics analysis of bacteria and fungi onboard the International Space Station.

BACKGROUND: The International Space Station (ISS) is a unique and complex built environment with the ISS surface microbiome originating from crew and cargo or from life support recirculation in an almost entirely closed system. The Microbial Tracking 1 (MT-1) project was the first ISS environmental surface study to report on the metagenome profiles without using whole-genome amplification. The study surveyed the microbial communities from eight surfaces over a 14-month period. The Microbial Tracking 2 (MT-2) project aimed to continue the work of MT-1, sampling an additional four flights from the same locations, over another 14 months. METHODS: Eight surfaces across the ISS were sampled with sterile wipes and processed upon return to Earth. DNA extracted from the processed samples (and controls) were treated with propidium monoazide (PMA) to detect intact/viable cells or left untreated and to detect the total DNA population (free DNA/compromised cells/intact cells/viable cells). DNA extracted from PMA-treated and untreated samples were analyzed using shotgun metagenomics. Samples were cultured for bacteria and fungi to supplement the above results. RESULTS: Staphylococcus sp. and Malassezia sp. were the most represented bacterial and fungal species, respectively, on the ISS. Overall, the ISS surface microbiome was dominated by organisms associated with the human skin. Multi-dimensional scaling and differential abundance analysis showed significant temporal changes in the microbial population but no spatial differences. The ISS antimicrobial resistance gene profiles were however more stable over time, with no differences over the 5-year span of the MT-1 and MT-2 studies. Twenty-nine antimicrobial resistance genes were detected across all samples, with macrolide/lincosamide/streptogramin resistance being the most widespread. Metagenomic assembled genomes were reconstructed from the dataset, resulting in 82 MAGs. Functional assessment of the collective MAGs showed a propensity for amino acid utilization over carbohydrate metabolism. Co-occurrence analyses showed strong associations between bacterial and fungal genera. Culture analysis showed the microbial load to be on average 3.0 × 105 cfu/m2 CONCLUSIONS: Utilizing various metagenomics analyses and culture methods, we provided a comprehensive analysis of the ISS surface microbiome, showing microbial burden, bacterial and fungal species prevalence, changes in the microbiome, and resistome over time and space, as well as the functional capabilities and microbial interactions of this unique built microbiome. Data from this study may help to inform policies for future space missions to ensure an ISS surface microbiome that promotes astronaut health and spacecraft integrity. Video Abstract.