Disentangling the rhizosphere community through stable isotope informed genome-resolved metagenomics and assembled metatranscriptomes

The functioning, health, and productivity of soil is intimately tied to the complex network of interactions in the rhizosphere. Because of this, the rhizosphere has been rigorously studied for over a century, but due to technical limitations many aspects of soil biology have been overlooked. In order to better understand rhizosphere functioning, my work has focused on the less explored organisms and interactions in microbial communities, this includes unculturable bacteria along with viruses and eukaryotes. Only by considering soil biology more holistically can we better understand the functioning of this enigmatic yet critical ecosystem. Knowledge about these interactions could direct how we think about plant-microbe relationships, soil carbon stabilization and the roles of understudied organisms in biogeochemical cycling.The transformation of plant photosynthate into soil organic carbon and its recycling to CO2 by soil microorganisms is one of the central components of the terrestrial carbon cycle. There are currently large knowledge gaps related to which soil-associated microorganisms take up plant carbon in the rhizosphere and the fate of that carbon. Additionally, understanding about obligate symbionts such as members of the Candidate Phyla Radiation (CPR) in soil is severely limited, both from the perspective of their genomic potential and their interactions with the greater soil community. We conducted an experiment in which common wild oats (Avena fatua) were grown in a 13CO2 atmosphere and the rhizosphere and non-rhizosphere soil was sampled for genomic analyses. Density gradient centrifugation of DNA extracted from soil samples enabled distinction of microbes that did and did not incorporate the 13C into their DNA. A 1.45-Mbp genome of a Saccharibacteria (TM7) was identified and, despite the microbial complexity of rhizosphere soil, curated to completion. The genome lacks many biosynthetic pathways, including genes required to synthesize DNA de novo. Rather, it acquires externally derived nucleic acids for DNA and RNA synthesis. Given this, we conclude that rhizosphere-associated Saccharibacteria recycle DNA from bacteria that live off plant exudates and/or phage that acquired 13C because they preyed upon these bacteria and/or directly from the labeled plant DNA. Isotopic labeling indicates that the population was replicating during the 6-week period of plant growth. Interestingly, the genome is ~ 30% larger than other complete Saccharibacteria genomes from non-soil environments, largely due to more genes for complex carbon utilization and amino acid metabolism. Given the ability to degrade cellulose, hemicellulose, pectin, starch, and 1,3-β-glucan, we predict that this Saccharibacteria generates energy by fermentation of soil necromass and plant root exudates to acetate and lactate. The genome also encodes a linear electron transport chain featuring a terminal oxidase, suggesting that this Saccharibacteria may respire aerobically. The genome encodes a hydrolase that could breakdown salicylic acid, a plant defense signaling molecule, and genes to interconvert a variety of isoprenoids, including the plant hormone zeatin. We propose that isotopically labeled CO2 is incorporated into plant-derived carbon and then into the DNA of rhizosphere organisms capable of nucleotide synthesis, and the nucleotides are recycled into Saccharibacterial genomes. We collected paired rhizosphere and non-rhizosphere soil at six and nine weeks of plant growth and extracted DNA that was separated by density gradient centrifugation. The separate fractions were sequenced, assembled, and binned to generate 55 unique microbial genomes that were >70% complete. Evidence for close interaction between bacteria, micro-eukaryotes and plant roots includes the ability to modulate plant signaling hormones, abundant plant pathogenicity factors and production of cyanide and insecticidal toxins. We reconstructed eukaryotic 18S rRNA sequences and identified micro-eukaryotic bacterivores and fungi in the rhizosphere soil. In addition, we reconstructed two complete genomes for phage that were among the most highly 13C-enriched entities in our study. CRISPR locus targeting connected a phage to a Burkholderiales host predicted to be a plant pathogen and a possible plant growth promoting Catenulispora may serve as the host for another phage. Thus, 13C could be tracked from the atmosphere into plant roots, soil and through the rhizosphere food web. Viruses impact nearly all organisms on Earth, with ripples of influence in agriculture, health and biogeochemical processes. We previously investigated DNA phage, however, very little is known about RNA viruses in an environmental context, and even less is known about their diversity and ecology in the most complex microbial system, soil. Here, we assembled 48 individual metatranscriptomes from four habitats within a soil sampled over a 22-day time series: rhizosphere alone, detritosphere alone, a combination of the two, and unamended soil (four time points and three biological replicates per time point). We resolved the RNA viral community, uncovering a high diversity of viral sequences. We also investigated possible host organisms by analyzing metatranscriptome marker gene content. Based on viral phylogeny, much of the diversity was Narnaviridae that parasitize fungi or Leviviridae that infect Proteobacteria. Both host and viral communities appear to be highly dynamic, and rapidly diverged depending on experimental conditions. The viral communities were structured based on the presence of litter, while putative hosts appeared to be impacted by both the presence of litter and roots. A clear time signature from Leviviridae and their hosts indicated that viruses were replicating. With this time-resolved analysis, we show that RNA viruses are diverse, abundant and active in soil. Their replication causes host cell death, mobilizing carbon in a process that represents a largely overlooked component of carbon cycling in soil.By combining state of the art techniques, stable isotope probing, genome-resolved metagenomics and assembled metatranscriptomics, we advanced knowledge about the interplay between understudied players in the rhizosphere and provided some clues for the fate of plant derived carbon in the soil microbial ecosystem. The use of genome resolved metagenomics is the only current way to determine the lifestyle of uncultured microbes. Complete genomes are still difficult to reconstruct however, they contain extensive of information, both regarding the presence and the absence of capabilities. Stable isotope probing allowed us to follow plant fixed carbon into the microbial community and in several cases across multiple trophic levels. This study demonstrates the power of stable isotope-informed genome-resolved metagenomics to resolve aspects of the complex rhizosphere food web. The approach will find broad application for study of other soils and different ecosystems

Noncompete Agreements in the U.S. Labor Force

Using nationally representative survey data on 11,505 labor force participants, we examine the use and implementation of noncompete agreements and the employee outcomes associated with these provisions. Approximately 18 percent of labor force participants are bound by noncompetes, with 38 percent having agreed to at least one in the past. Noncompetes are more likely to be found in high-skill, high-paying jobs, but they are also common in low-skill, low-paying jobs and in states where noncompetes are unenforceable. Only 10 percent of employees negotiate over their noncompetes, and about one-third of employees are presented with noncompetes after having already accepted job offers. Early-notice noncompetes are associated with better employee outcomes, while employees who agree to late-notice noncompetes are comparatively worse off. Regardless of noncompete timing, however, wages are relatively lower where noncompetes are easier to enforce. We discuss these findings in light of competing theories of the economic value of noncompetes

Comparison of Tissue Injury from Focused Ultrasonic Propulsion of Kidney Stones Versus Extracorporeal Shock Wave Lithotripsy

Purpose Focused ultrasonic propulsion is a new non-invasive technique designed to move kidney stones and stone fragments out of the urinary collecting system. However, the extent of tissue injury associated with this technique is not known. As such, we quantitated the amount of tissue injury produced by focused ultrasonic propulsion under simulated clinical treatment conditions, and under conditions of higher power or continuous duty cycles, and compared those results to SWL injury. Materials and Methods A human calcium oxalate monohydrate stone and/or nickel beads were implanted (with ureteroscopy) into 3 kidneys of live pigs (45–55 kg) and repositioned using focused ultrasonic propulsion. Additional pig kidneys were exposed to SWL level pulse intensities or continuous ultrasound exposure of 10 minutes duration (ultrasound probe either transcutaneous or on the kidney). These kidneys were compared to 6 kidneys treated with an unmodified Dornier HM3 Lithotripter (2400 shocks, 120 SWs/min and 24 kV). Histological analysis was performed to assess the volume of hemorrhagic tissue injury created by each technique (% functional renal volume, FRV). Results SWL produced a lesion of 1.56±0.45% FRV. Ultrasonic propulsion produced no detectable lesion with the simulated clinical treatment. A lesion of 0.46±0.37% FRV or 1.15±0.49% FRV could be produced if excessive treatment parameters were used while the ultrasound probe was placed on the kidney. Conclusions Focused ultrasonic propulsion produced no detectable morphological injury to the renal parenchyma when using clinical treatment parameters and produced injury comparable in size to SWL when using excessive treatment parameters

Breed-Specific Hematological Phenotypes in the Dog: A Natural Resource for the Genetic Dissection of Hematological Parameters in a Mammalian Species

Remarkably little has been published on hematological phenotypes of the domestic dog, the most polymorphic species on the planet. Information on the signalment and complete blood cell count of all dogs with normal red and white blood cell parameters judged by existing reference intervals was extracted from a veterinary database. Normal hematological profiles were available for 6046 dogs, 5447 of which also had machine platelet concentrations within the reference interval. Seventy-five pure breeds plus a mixed breed control group were represented by 10 or more dogs. All measured parameters except mean corpuscular hemoglobin concentration (MCHC) varied with age. Concentrations of white blood cells (WBCs), neutrophils, monocytes, lymphocytes, eosinophils and platelets, but not red blood cell parameters, all varied with sex. Neutering status had an impact on hemoglobin concentration, mean corpuscular hemoglobin (MCH), MCHC, and concentrations of WBCs, neutrophils, monocytes, lymphocytes and platelets. Principal component analysis of hematological data revealed 37 pure breeds with distinctive phenotypes. Furthermore, all hematological parameters except MCHC showed significant differences between specific individual breeds and the mixed breed group. Twenty-nine breeds had distinctive phenotypes when assessed in this way, of which 19 had already been identified by principal component analysis. Tentative breed-specific reference intervals were generated for breeds with a distinctive phenotype identified by comparative analysis. This study represents the first large-scale analysis of hematological phenotypes in the dog and underlines the important potential of this species in the elucidation of genetic determinants of hematological traits, triangulating phenotype, breed and genetic predisposition

The James Webb Space Telescope Mission

Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least $4m$. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the $6.5m$ James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 years, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space Telescope Overview, 29 pages, 4 figure

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries