Writing Committee for the REMAP-CAP Investigators IMPORTANCE The evidence for benefit of convalescent plasma for critically ill patients with COVID-19 is inconclusive.OBJECTIVE To determine whether convalescent plasma would improve outcomes for critically ill adults with COVID-19. DESIGN, SETTING, AND PARTICIPANTSThe ongoing Randomized, Embedded, Multifactorial, Adaptive Platform Trial for Community-Acquired Pneumonia (REMAP-CAP) enrolled and randomized 4763 adults with suspected or confirmed COVID-19 between March 9, 2020, and January 18, 2021, within at least 1 domain; 2011 critically ill adults were randomized to open-label interventions in the immunoglobulin domain at 129 sites in 4 countries. Follow-up ended on April 19, 2021. INTERVENTIONSThe immunoglobulin domain randomized participants to receive 2 units of high-titer, ABO-compatible convalescent plasma (total volume of 550 mL ± 150 mL) within 48 hours of randomization (n = 1084) or no convalescent plasma (n = 916). MAIN OUTCOMES AND MEASURESThe primary ordinal end point was organ support-free days (days alive and free of intensive care unit-based organ support) up to day 21 (range, −1 to 21 days; patients who died were assigned -1 day). The primary analysis was an adjusted bayesian cumulative logistic model. Superiority was defined as the posterior probability of an odds ratio (OR) greater than 1 (threshold for trial conclusion of superiority >99%). Futility was defined as the posterior probability of an OR less than 1.2 (threshold for trial conclusion of futility >95%). An OR greater than 1 represented improved survival, more organ support-free days, or both. The prespecified secondary outcomes included in-hospital survival; 28-day survival; 90-day survival; respiratory support-free days; cardiovascular support-free days; progression to invasive mechanical ventilation, extracorporeal mechanical oxygenation, or death; intensive care unit length of stay; hospital length of stay; World Health Organization ordinal scale score at day 14; venous thromboembolic events at 90 days; and serious adverse events. RESULTS Among the 2011 participants who were randomized (median age, 61 [IQR, 52 to 70] years and 645/1998 [32.3%] women), 1990 (99%) completed the trial. The convalescent plasma intervention was stopped after the prespecified criterion for futility was met. The median number of organ support-free days was 0 (IQR, -1 to 16) in the convalescent plasma group and 3 (IQR, -1 to 16) in the no convalescent plasma group. The in-hospital mortality rate was 37.3% (401/1075) for the convalescent plasma group and 38.4% (347/904) for the no convalescent plasma group and the median number of days alive and free of organ support was 14 (IQR, 3 to 18) and 14 (IQR, 7 to 18), respectively. The median-adjusted OR was 0.97 (95% credible interval, 0.83 to 1.15) and the posterior probability of futility (OR <1.2) was 99.4% for the convalescent plasma group compared with the no convalescent plasma group. The treatment effects were consistent across the primary outcome and the 11...

The staphylococcal accessory regulator A (sarA) impacts the extracellular accumulation of Staphylococcus aureus virulence factors at the level of intracellular production and extracellular protease-mediated degradation. We previously used a proteomics approach that measures protein abundance of all proteoforms to demonstrate that mutation of sarA results in increased levels of extracellular proteases and assesses the impact of this on the accumulation of S. aureus exoproteins. Our previous approach was limited as it did not take into account that large, stable proteolytic products from a given protein could result in false negatives when quantified by total proteoforms. Here, our goal was to use an expanded proteomics approach utilizing a dual quantitative method for measuring abundance at both the total proteoform and full-length exoprotein levels to alleviate these false negatives and thereby provide for characterization of protease-dependent and -independent effects of sarA mutation on the S. aureus exoproteome. Proteins present in conditioned medium from overnight, stationary phase cultures of the USA300 strain LAC, an isogenic sarA mutant, and a sarA mutant unable to produce any of the known extracellular proteases (sarA/protease) were resolved using one-dimensional gel electrophoresis. Quantitative proteomic comparisons of sarA versus sarA/protease mutants identified proteins that were cleaved in a protease-dependent manner owing to mutation of sarA, and comparisons of sarA/protease mutant versus the LAC parent strain identified proteins in which abundance was altered in a sarA mutant in a protease-independent manner. Furthermore, the proteins uniquely identified by the full-length data analysis approach eliminated false negatives observed in the total proteoform analysis. This expanded approach provided for a more comprehensive analysis of the impact of mutating sarA on the S. aureus exoproteome.

Quantitative proteomics generates large datasets with increasing depth and quantitative information.

BackgroundResistant starch is a prebiotic metabolized by the gut bacteria. It has been shown to attenuate chronic kidney disease (CKD) progression in rats. Previous studies employed taxonomic analysis using 16S rRNA sequencing and untargeted metabolomics profiling. Here we expand these studies by metaproteomics, gaining new insight into the host-microbiome interaction.MethodsDifferences between cecum contents in CKD rats fed a diet containing resistant starch with those fed a diet containing digestible starch were examined by comparative metaproteomics analysis. Taxonomic information was obtained using unique protein sequences. Our methodology results in quantitative data covering both host and bacterial proteins.Results5,834 proteins were quantified, with 947 proteins originating from the host organism. Taxonomic information derived from metaproteomics data surpassed previous 16S RNA analysis, and reached species resolutions for moderately abundant taxonomic groups. In particular, the Ruminococcaceae family becomes well resolved–with butyrate producers and amylolytic species such as R. bromii clearly visible and significantly higher while fibrolytic species such as R. flavefaciens are significantly lower with resistant starch feeding. The observed changes in protein patterns are consistent with fiber-associated improvement in CKD phenotype. Several known host CKD-associated proteins and biomarkers of impaired kidney function were significantly reduced with resistant starch supplementation. Data are available via ProteomeXchange with identifier PXD008845.ConclusionsMetaproteomics analysis of cecum contents of CKD rats with and without resistant starch supplementation reveals changes within gut microbiota at unprecedented resolution, providing both functional and taxonomic information. Proteins and organisms differentially abundant with RS supplementation point toward a shift from mucin degraders to butyrate producers.