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The ongoing SARS-Cov-2 pandemic had many drastic effects upon society beyond the illness and death it caused. Pandemic mitigation measures disrupted and altered behaviors related to social mobility, significantly increasing the time spent at home compared to the pre-pandemic period. Further, it was well documented that social anxiety and stress increased at a population level. Early in the pandemic there was speculation in the popular media that reporting of paranormal phenomena (e.g., UFOs, ghosts, etc.) increased due to factors associated with the pandemic. Past research on UFO/UAP reporting has theorized that increases are triggered by social factors, and so the pandemic provided a natural experiment to test these claims. To measure UFO reports we utilized two public databases of UFO reports for sightings in the United States, provided by the National UFO Reporting Center and the Mutual UFO Network. To estimate the impact of the pandemic we utilized two measures, one for social mobility and one for pandemic/disease severity. Google Community Mobility Reports provided a metric of social mobility for people who use Google Maps on their cellular telephone (i.e., amount of time spent at work compared to home), which we aggregated to a state level to estimate time spent at home. Second, we used new weekly SARS-CoV-2 cases and deaths, both absolute counts and per capita, which can be considered to be an indirect measure of anxiety and stress. We find that UFO reports did increase in 2020 compared to 2019 (p

Smallholder farms in the semi-arid and sub-humid tropics are particularly vulnerable to increased climate variability. Indigenous agrisystems that have co-evolved with climate variability may have developed resilience strategies. In the Southwest Ethiopian Highlands, agrisystems are dominated by the multipurpose perennial staple enset ( Ensete ventricosum ), which has flexible harvest timing, high yield, long storage, and putative drought tolerance, earning it the name 'tree against hunger'. We tested three hypotheses using crop production area and climate data. First, that enset production area is greatest in the most drought-prone locations. Second, that farmers respond to drought events by increasing enset production area. And third, that drought encourages shifts in agrisystem composition more widely towards perennial or storable crops. We found that regions with a higher severe drought frequency are associated with significantly higher proportion of enset production. Similarly, the Standardised Precipitation Evapotranspiration Index of the previous three years is significantly negatively correlated with enset production time series, suggesting that prior drier conditions led farmers to increase the area under enset production. Regarding other crops, storage crops roots and tubers were also preferentially selected after long-term drought over annual crops, indicating their capacity for longer-term resilience. Promoting the production of crops such as perennials, which have more extensive and established root systems, may be a strategy to ensure food security during drought or climate variability. These results indicate the potential of farmer's resilience strategies to improve food security in a changing climate.

Peripheral nerve injury results in loss of motor and sensory function distal to the nerve injury and is often permanent in nerve gaps longer than 5 cm. Autologous nerve grafts (nerve autografts) utilize patients’ own nerve tissue from another part of their body to repair the defect and are the gold standard in care. However, there is a limited autologous tissue supply, size mismatch between donor nerve and injured nerve, and morbidity at the site of nerve donation. Decellularized cadaveric nerve tissue alleviates some of these limitations and has demonstrated success clinically. We previously developed an alternative apoptosis-assisted decellularization process for nerve tissue. This new process may result in an ideal scaffold for peripheral nerve regeneration by gently removing cells and antigens while preserving delicate topographical cues. In addition, the apoptosis-assisted process requires less active processing time and is inexpensive. This study examines the utility of apoptosis-decellularized peripheral nerve scaffolds compared to detergent-decellularized peripheral nerve scaffolds and isograft controls in a rat nerve gap model. Results indicate that, at 8 weeks post-injury, apoptosis-decellularized peripheral nerve scaffolds perform similarly to detergent-decellularized and isograft controls in both functional (muscle weight recovery, gait analysis) and histological measures (neurofilament staining, macrophage infiltration). These new apoptosis-decellularized scaffolds hold great promise to provide a less expensive scaffold for nerve injury repair, with the potential to improve nerve regeneration and functional outcomes compared to current detergent-decellularized scaffolds.

The Physician Fee Schedule was updated in 2021 by the Centers for Medicare and Medicaid Services. A case study on Tetralogy of Fallot, the most common cyanotic congenital heart defect, is presented with guidance in billing the office visit to reflect the current guidelines.

BackgroundCombatting the tuberculosis (TB) epidemic caused byMycobacterium tuberculosis(Mtb) necessitates a better understanding of the factors contributing to patient clinical outcomes and transmission. While host and environmental factors have been evaluated, the impact ofMtbgenetic background and phenotypic diversity is underexplored. Previous work has made associations betweenMtbgenetic lineages and some clinical and epidemiological features, but the bacterial traits underlying these connections are largely unknown.MethodsWe developed a high-throughput functional genomics platform for defining genotype-phenotype relationships across a panel ofMtbclinical isolates. These phenotypic fitness profiles function as intermediate traits which can be linked toMtbgenetic variants and associated with clinical and epidemiological outcomes. We applied this approach to a collection of 158Mtbstrains from a study ofMtbtransmission in Ho Chi Minh City, Vietnam.Mtbstrains were genetically tagged in multiplicate, which allowed us to pool the strains and assessin vitrocompetitive fitness using deep sequencing across a set of 14 host-relevant antibiotic and metabolic conditions. Phylogenetic and monogenic associations with these intermediate traits were identified and then associated with clinical outcomes.FindingsMtbclinical strains have a broad range of growth and drug response dynamics that can be clustered by their phylogenetic relationships. We identified novel monogenic associations withMtbfitness in various metabolic and antibiotic conditions. Among these, we find that mutations inRv1339, a phosphodiesterase, which were identified through their association with slow growth in glycerol, are further associated with treatment failure. We also identify a previously uncharacterized subclade of Lineage 1 strains (L1.1.1.1) that is phenotypically distinguished by slow growth under most antibiotic and metabolic stress conditionsin vitro. This clade is associated with cavitary disease, treatment failure, and demonstrates increased transmission potential.InterpretationHigh-throughput phenogenotyping of Mtb clinical strains enabled bacterial intermediate trait identification that can provide a mechanistic link betweenMtbgenetic variation and patient clinical outcomes.Mtbstrains associated with cavitary disease, treatment failure, and transmission potential display intermediate phenotypes distinguished by slow growth under various antibiotic and metabolic conditions. These data suggest that Mtb growth regulation is an adaptive advantage for host bacterial success in human populations, in at least some circumstances. These data further suggest markers for the underlying bacterial processes that govern these clinical outcomes.FundingNational Institutes of Allergy and Infectious Diseases: P01 AI132130 (SS, SMF); P01 AI143575 (XW, SMF); U19 AI142793 (QL, SMF); 5T32AI132120-03 (SS); 5T32AI132120-04 (SS); 5T32AI049928-17 (SS) Wellcome Trust Fellowship in Public Health and Tropical Medicine: 097124/Z/11/Z (NTTT) National Health and Medical Research Council (NHMRC)/A*STAR joint call: APP1056689 (SJD) The funding sources had no involvement in study methodology, data collection, analysis, and interpretation nor in the writing or submission of the manuscript.Research in contextEvidence before this studyWe used different combinations of the words mycobacterium tuberculosis, tuberculosis, clinical strains, intermediate phenotypes, genetic barcoding, phenogenomics, cavitary disease, treatment failure, and transmission to search the PubMed database for all studies published up until January 20th, 2022. We only considered English language publications, which biases our search. Previous work linkingMtbdeterminants to clinical or epidemiological data has made associations between bacterial lineage, or less frequently, genetic polymorphisms toin vitroorin vivomodels of pathogenesis, transmission, and clinical outcomes such as cavitary disease, treatment failure, delayed culture conversion, and severity. Many of these studies focus on the global pandemic Lineage 2 and Lineage 4Mtbstrains due in part to a deletion in a polyketide synthase implicated in host-pathogen interactions. There are a number ofMtbGWAS studies that have led to novel genetic determinants ofin vitrodrug resistance and tolerance. PreviousMtbGWAS analyses with clinical outcomes did not experimentally test any predicted phenotypes of the clinical strains. Published laboratory-based studies ofMtbclinical strains involve relatively small numbers of strains, do not identify the genetic basis of relevant phenotypes, or link findings to the corresponding clinical outcomes. There are two recent studies of other pathogens that describe phenogenomic analyses. One study of 331M. abscessusclinical strains performed one-by-one phenotyping to identify bacterial features associated with clearance of infection and another details a competition experiment utilizing three barcodedPlasmodium falciparumclinical isolates to assay antimalarial fitness and resistance.Added value of this studyWe developed a functional genomics platform to perform high-throughput phenotyping ofMtbclinical strains. We then used these phenotypes as intermediate traits to identify novel bacterial genetic features associated with clinical outcomes. We leveraged this platform with a sample of 158Mtbclinical strains from a cross sectional study ofMtbtransmission in Ho Chi Minh City, Vietnam. To enable high-throughput phenotyping of large numbers ofMtbclinical isolates, we applied a DNA barcoding approach that has not been previously utilized for the high-throughput analysis ofMtbclinical strains. This approach allowed us to perform pooled competitive fitness assays, tracking strain fitness using deep sequencing. We measured the replicative fitness of the clinical strains in multiplicate under 14 metabolic and antibiotic stress condition. To our knowledge, this is the largest phenotypic screen ofMtbclinical isolates to date. We performed bacterial GWAS to delineate theMtbgenetic variants associated with each fitness phenotype, identifying monogenic associations with several conditions. We then definedMtbphenotypic and genetic features associated with clinical outcomes. We find that a subclade ofMtbstrains, defined by variants largely involved in fatty acid metabolic pathways, share a universal slow growth phenotype that is associated with cavitary disease, treatment failure and increased transmission potential in Vietnam. We also find that mutations inRv1339, a poorly characterized phosphodiesterase, also associate with slow growthin vitroand with treatment failure in patients.Implications of all the available evidencePhenogenomic profiling demonstrates thatMtbstrains exhibit distinct growth characteristics under metabolic and antibiotic stress conditions. These fitness profiles can serve as intermediate traits for GWAS and association with clinical outcomes. Intermediate phenotyping allows us to examine potential processes by which bacterial strain differences contribute to clinical outcomes. Our study identifies clinical strains with slow growth phenotypes underin vitromodels of antibiotic and host-like metabolic conditions that are associated with adverse clinical outcomes. It is possible that the bacterial intermediate phenotypes we identified are directly related to the mechanisms of these outcomes, or they may serve as markers for the causal yet unidentified bacterial determinants. Via the intermediate phenotyping, we also discovered a surprising diversity inMtbresponses to the new anti-mycobacterial drugs that target central metabolic processes, which will be important in considering roll-out of these new agents. Our study and others that have identifiedMtbdeterminants of TB clinical and epidemiological phenotypes should inform efforts to improve diagnostics and drug regimen design.

Cell-derived matrices are useful tools for studying the extracellular matrix (ECM) of different cell types and testing the effects on cell migration or wound repair. These matrices typically are generated using extended culture with ascorbic acid to boost ECM production. Applying this technique to cancer cell cultures could advance the study of cancer ECM and its effects on recruitment and training of the tumor microenvironment, but ascorbic acid is potently cytotoxic to cancer cells. Macromolecular crowding agents can also be added to increase matrix deposition based on the excluded volume principle. We report the use of macromolecular crowding (MMC) alone as an effective strategy to generate brain cancer cell-derived matrices for downstream analyses and cell migration studies. We cultured the mouse glioblastoma cell line GL261 for 1 week in the presence of three previously-reported MMC agents (carrageenan, Ficoll 70/400, and hyaluronic acid). We measured the resulting deposition of collagens and sulfated glycosaminoglycans using quantitative assays, as well as other matrix components by immunostaining. Both carrageenan and Ficoll promoted significantly more accumulation of total collagen content, sulfated glycosaminoglycan content, and fibronectin staining. Only Ficoll, however, also demonstrated a significant increase in collagen I staining. The results were more variable in 3D spheroid culture. We focused on Ficoll MMC matrices, which were isolated using the small molecule Raptinal to induce cancer cell apoptosis and matrix decellularization. The cancer cell-derived matrix promoted significantly faster migration of human astrocytes in a scratch wound assay, which may be explained by focal adhesion morphology and an increase in cellular metabolic activity. Ultimately, these data show MMC culture is a useful technique to generate cancer cell-derived matrices and study the effects on stromal cell migration related to wound repair.

The mechanisms and regulation of RNA degradation in mycobacteria have been subject to increased interest following the identification of interplay between RNA metabolism and drug resistance. Mycobacteria encode multiple ribonucleases that are predicted to participate in mRNA degradation and/or processing of stable RNAs. RNase E is an endoribonuclease hypothesized to play a major role in mRNA degradation due to its essentiality in mycobacteria and its role in mRNA degradation in gram- negative bacteria. Here, we defined the impact of RNase E on mRNA degradation rates transcriptome- wide in the non-pathogenic modelMycolicibacterium smegmatis. RNase E played a rate-limiting role in the degradation of at least 89% of protein-coding genes, with leadered transcripts generally being more affected by RNase E repression than leaderless transcripts. There was an apparent global slowing of transcription in response to knockdown of RNase E, suggesting thatM. smegmatisregulates transcription in responses to changes in mRNA degradation. This compensation was incomplete, as the abundance of most transcripts increased upon RNase E knockdown. We assessed the sequence preferences for cleavage by RNase E transcriptome-wide in bothM. smegmatisandM. tuberculosis, and found a consistent bias for cleavage in C-rich regions. Purified RNase E had a clear preference for cleavage immediately upstream of cytidines, distinct from the sequence preferences of RNase E in gram-negatives. We furthermore report a high-resolution map of mRNA cleavage sites inM. tuberculosis, which occur primarily within the RNase E-preferred sequence context, confirming RNase E as a broad contributor toM. tuberculosistranscriptome structure.