The functional proteome landscape of Escherichia coli.

Recent developments in high-throughput reverse genetics1,2 have revolutionized our ability to map gene function and interactions3-6. The power of these approaches depends on their ability to identify functionally associated genes, which elicit similar phenotypic changes across several perturbations (chemical, environmental or genetic) when knocked out7-9. However, owing to the large number of perturbations, these approaches have been limited to growth or morphological readouts10. Here we use a high-content biochemical readout, thermal proteome profiling11, to measure the proteome-wide protein abundance and thermal stability in response to 121 genetic perturbations in Escherichia coli. We show that thermal stability, and therefore the state and interactions of essential proteins, is commonly modulated, raising the possibility of studying a protein group that is particularly inaccessible to genetics. We find that functionally associated proteins have coordinated changes in abundance and thermal stability across perturbations, owing to their co-regulation and physical interactions (with proteins, metabolites or cofactors). Finally, we provide mechanistic insights into previously determined growth phenotypes12 that go beyond the deleted gene. These data represent a rich resource for inferring protein functions and interactions.

Assessing LSAM's ability to account for changes in organ donation and transplant center behavior.

The Liver Simulated Allocation Model (LSAM) is used to evaluate proposed organ allocation policies. Although LSAM has been shown to predict the directionality of changes in transplants and nonused organs, the magnitude is often overestimated. One reason is that policymakers and researchers using LSAM assume static levels of organ donation and center behavior because of challenges with predicting future behavior. We sought to assess the ability of LSAM to account for changes in organ donation and organ acceptance behavior using LSAM 2019. We ran 1-year simulations with the default model and then ran simulations changing donor arrival rates (ie, organ donation) and center acceptance behavior. Changing the donor arrival rate was associated with a progressive simulated increase in transplants, with corresponding simulated decreases in waitlist deaths. Changing parameters related to organ acceptance was associated with important changes in transplants, nonused organs, and waitlist deaths in the expected direction in data simulations, although to a much lesser degree than changing the donor arrival rate. Increasing the donor arrival rate was associated with a marked decrease in the travel distance of donor livers in simulations. In conclusion, we demonstrate that LSAM can account for changes in organ donation and organ acceptance in a manner aligned with historical precedent that can inform future policy analyses. As Scientific Registry of Transplant Recipients develops new simulation programs, the importance of considering changes in donation and center practice is critical to accurately estimate the impact of new allocation policies.

Liver transplantation for biliary cysts: perioperative and long-term outcomes.

BACKGROUND: Biliary cysts (BC) is a rare indication for orthotopic liver transplantation (OLT). METHODS: We queried the UNOS dataset to identify patients who underwent OLT for Caroli's disease (CD) and choledochal cysts (CC). All patients with BC (CD + CC) were compared to a cohort of patients transplanted for other indications. Patients with CC were also compared to those with CD. Cox proportional hazard model was performed to assess predictors of graft and patient survival. RESULTS: 261 patients underwent OLT for BC. Patients with BC had better pre-operative liver function compared to those transplanted for other indications. 5-year graft and patient survival were 72% and 81%, respectively, similar to those transplanted for other indications after matching. Patients with CC were younger and had increased preoperative cholestasis compared to those with CD. Donor age, race, and gender were predictors of poor graft and patient survival in patients transplanted for CC. CONCLUSIONS: Patients with BC have similar outcomes to those transplanted for other indications and more frequently require MELD score exception. In patients transplanted for choledochal cysts, female gender, donor age, and African-American race were independent predictors of poor survival. Pediatric patients transplanted for Caroli's disease had better survival compared to adults.

COVID-19 and transplantation-Data censoring.

During the early wave of the COVID-19 pandemic, the Scientific Registry of Transplant Recipients (SRTR) designated a "black out" period between March 12, 2020, and June 12, 2020, for transplant outcomes reporting. We discuss the implications and potential bias it has introduced as it may selectively favor the outcomes for certain regions and harm other regions due to varied effects of different waves of COVID-19 infections across the United States.

Predictors of Liver Failure in Non-Cirrhotic Patients Undergoing Hepatectomy.

BACKGROUND: Post-hepatectomy liver failure (PHLF) is associated with high mortality following liver resection. There have been limited studies evaluating predictors of PHLF and clinically significant PHLF in non-cirrhotic patients. METHODS: This was a retrospective cohort study using the National Surgical Quality Improvement Program database (NSQIP) to evaluate 8,093 non-cirrhotic patients undergoing hepatectomy from 2014 to 2018. Primary endpoints were PHLF and clinically significant PHLF (PHLF grade B or C). RESULTS: Among all patients, 4.74% (n = 383) developed PHLF and 2.5% clinically significant PHLF (n = 203). The overall 30-day mortality was 1.35% (n = 109), 11.5% (n = 44) in patients with PHLF, and 19.2% in those with clinically significant PHLF. Factors associated with PHLF were: metastatic liver disease (OR = 1.84, CI = 1.14-2.98), trisectionectomy (OR = 3.71, CI = 2.59-5.32), right total lobectomy (OR = 4.17, CI = 3.06-5.68), transfusions (OR = 1.99, CI = 1.52-2.62), organ/space SSI (OR = 2.84, CI = 2.02-3.98), post-operative pneumonia (OR = 2.43, CI = 1.57-3.76), sepsis (OR = 2.27, CI = 1.47-3.51), and septic shock (OR = 5.67, CI = 3.43-9.36). Patients who developed PHLF or clinically significant PHLF had 2-threefold increased risk of perioperative mortality. Post-hepatectomy renal failure (OR = 8.47, CI = 3.96-18.1), older age (OR = 1.04, CI = 1.014-1.063), male sex (OR = 1.83, CI = 1.07-3.14), sepsis (OR = 2.96, CI = 1.22-7.2), and septic shock (OR = 3.92, CI = 1.61-9.58) were independently associated with 30-mortality in patients with clinically significant PHLF. CONCLUSION: PHLF in non-cirrhotic patients increased the risk of perioperative mortality and is associated with the extent of hepatectomy and infectious complications. Careful evaluation of the liver remnant, antibiotic prophylaxis, nutritional assessment, and timely management of post-operative infections could decrease major morbidity and mortality following hepatectomy.

Results of a data-driven performance improvement initiative in organ donation.

New metrics for organ procurement organization (OPO) performance utilize National Center for Health Statistics data to measure cause, age, and location consistent (CALC) deaths. We used this denominator to identify opportunities for improved donor conversion at one OPO, Indiana Donor Network (INOP). We sought to determine whether such analyses are immediately actionable for quality improvement (QI) initiatives directed at increased donor conversion. CALC-based assessment of INOP's performance revealed an opportunity to improve conversion of older donors. Following the QI initiative, INOP donor yield rose by 44%, while organs transplanted rose by 29%. These changes tolerated temporary disruption around the COVID-19 pandemic. Improved donor yield was primarily seen in older groups identified by CALC-based methods. Process changes in resource allocation and monitoring were associated with a 57% increase in the number of potential donors approached in the QI period and a subsequent rise in the number of potential donor referrals, suggesting positive feedback at area hospitals. Post-intervention, INOP's projected donation performance rose from 51st to 18th among all OPOs. OPOs can use CALC death data to accurately assess donor conversion by categories including age and race/ethnicity. These data can be used in real time to inform OPO-level processes to maximize donor recovery.

Perioperative Challenges in Patients Transplanted with Livers from Extreme Obese Donors.

The combination of rising rates of obesity and the shortage of deceased donor livers have forced the consideration of marginal liver donors in terms of body mass index (BMI) for liver transplantation (LT). To date, there are still conflicting data on the impact of donor obesity on post-LT outcomes. We analyzed all patients undergoing LT alone in the United States (US) from October 2005 through December 2019 using the United Network of Organ Sharing (UNOS) data set. We categorized donor BMI >40 kg/m2 as extremely obese (EO). Primary endpoints included 30-day perioperative mortality and early graft loss (EGL) within 7 days. A subgroup analysis was performed for the EO donor group to assess how macrovesicular steatosis (MaS) >30% affects 30-day mortality and EGL within 7 days. A total of 72,616 patients underwent LT during the study period. The 30-day perioperative mortality was significantly higher in the EO donor group (P = 0.02). On multivariate analysis, recipients undergoing LT with EO donors had a 38% higher 30-day mortality risk (odds ratio [OR], 1.38; 95% confidence interval [CI], 1.21-1.69) and 53% increased risk of EGL (OR, 1.53; 95% CI, 1.22-1.90). MaS >30% was independently associated with a 2-fold increased risk of 30-day mortality (P = 0.003) and 3.5-fold increased risk of EGL within 7 days (P < 0.001). The impact of MaS >30% in EGL was 2-fold for all patients transplanted during the study period compared with 3.5-fold in the EO donor group. There is an increased risk of EGL and 30-day perioperative mortality in recipients transplanted with EO donors. Future studies are warranted in morbid and super obese donors to assess the possible effect of obesity-related proinflammatory factors in EGL.

Alcohol-related cirrhosis is associated with high coronary artery calcium scores in patients undergoing evaluation for orthotopic liver transplantation.

BACKGROUND AND AIMS: Coronary artery disease is a major cause of morbidity and mortality in liver transplant patients. Coronary artery calcium (CAC) score has been used to evaluate the risk of CAD in non-cirrhotic patients. However, its significance in cirrhotic patients is unknown. This study aimed to identify factors associated with elevated CAC scores in patients with end-stage liver disease undergoing liver transplant evaluation. METHODS: We retrospectively reviewed all patients who underwent liver transplantation evaluation and had coronary CT scan between January 2015 and December 2018. Patients with prior history of CAD were excluded. CAC score was calculated based on the method described by Agatston. RESULTS: Sixty-two patients were included. 37.1% had alcohol-related liver disease and 27.4% had NASH cirrhosis. Mean CAC score was 261.1 ± SD, 463.84. Alcohol-related liver disease, male gender, and hypertension were significantly associated with CAC score >100 and only alcohol-related liver disease was associated with CAC score >300. In logistic regression, patients with alcohol-related liver disease had more than sixfold increase in risk of having CAC scores >100 and 300 (OR 6.14, and 6.70, respectively). CONCLUSION: Alcohol-related liver disease, male gender, and hypertension were significantly associated with an increased CAC score >100. However, alcohol-related liver disease was the only factor associated with CAC score >300.

Hepatic Steatosis is Associated with an Increased Risk of Postoperative Infections and Perioperative Transfusion Requirements in Patients Undergoing Hepatectomy.

BACKGROUND: To determine the impact of hepatic steatosis on perioperative outcomes of patients undergoing hepatectomy. METHODS: We analyzed all hepatectomy patients with normal and fatty liver texture, between 2014 and 2018 using NSQIP. Main endpoints included perioperative transfusions (within 72 h) and infectious complications. RESULTS: A total of 8,237 patients underwent hepatectomy during the study period. The overall rate of fatty liver texture (FLG) was 31% (2,557). Operative duration was significantly longer; inflow occlusion was more common (Pringle maneuver), and the need of transfusions was significantly higher in the FLG compared to the normal liver group (NLG) (p = < 0.001). On multivariate analysis, patients in the FLG had increased risk of developing infectious complications (OR 1.22 [95%IC 1.05-1.41]) and transfusion requirements within 72 h after hepatectomy (OR 1.43 [95% CI 1.24-1.63]). CONCLUSIONS: Hepatic steatosis is an independent risk factor for the development of infectious complications and increased perioperative transfusion requirements in patients undergoing hepatectomy. Those requiring transfusions within 72 h had also an increased risk of infections after hepatectomy.

Development and Evaluation of a Quantitative Fluorescent Lateral Flow Immunoassay for Cystatin-C, a Renal Dysfunction Biomarker.

The diagnosis, prognosis, and control of chronic kidney disease rely on an understanding of the glomerular filtration rate (GFR). The renal clearance of the cystatin-C is closely associated with the GFR. Cystatin-C is a more suitable GFR marker than the commonly used creatinine. General techniques for cystatin-C calculation, such as particle-enhanced turbidimetric and nephelometric assay, are time-consuming and tedious. Here, we propose a rapid, quantitative immunoassay for the detection of cystatin-C. A fluorescence-based lateral-flow kit was developed in a sandwich format by using a monoclonal antibody. A Linear calibration was obtained over the clinical diagnostic range of 0.023-32 µg/mL and the limit of detection (LOD) was 0.023 µg/mL and the limit of quantification (LOQ) was 0.029 µg/mL. Average recoveries from spiked urine samples ranged from 96-100% and the coefficient of variation was less than 4% for both intra and inter-day assays with excellent repeatability. With the comparison with an ELISA kit, the developed kit is highly sensitive, performs well over the detection range, provides repeatable results in a short time, and can easily be used at point-of-care (POC), making it an ideal candidate for rapid testing in early detection, community screening for renal function disorders.

Rejecting bias: The case against race adjustment for OPO performance in communities of color.

In December of 2019, the Centers for Medicare and Medicaid Services (CMS) put out a notice of proposed rule-making for 42 CFR Part 486, specifically the section that covers the organ procurement organization (OPO) Conditions for Coverage. Most crucially, the proposed rule included two new OPO performance metrics using objective, standardized data from the Centers for Disease Control and Prevention (CDC). These new metrics would employ a denominator that included inpatient deaths from certain causes that could lead to organ donation, rather than the current unverifiable eligible death metric. Although there has been near-uniform support for replacing the eligible death denominator with CDC data, a source of contention is CMS's proposal not to adjust risk for race in their OPO outcome. Nonetheless, there have been calls for race and ethnicity to be included as risk-adjusted variables in the CMS donation metric. Herein, we lay out an argument as to why inclusion of race and ethnicity as risk adjustment variables in an OPO performance metric is not only statistically suspect but also will hide the inequities that are detrimental to optimal system performance and assurance that all patients have timely access to donation.

Utilization of deceased donors during a pandemic: argument against using SARS-CoV-2-positive donors.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly become an unprecedented pandemic that has impacted society, disrupted hospital functions, strained health care resources, and impacted the lives of transplant professionals. Despite this, organ failure and the need for transplant continues throughout the United States. Considering the perpetual scarcity of deceased donor organs, Kates et al present a viewpoint that advocates for the utilization of coronavirus disease 2019 (COVID-19)-positive donors in selected cases. We present a review of the current literature that details the potential negative consequences of COVID-19-positive donors. The factors we consider include (1) the risk of blood transmission of SARS-CoV-2, (2) involvement of donor organs, (3) lack of effective therapies, (4) exposure of health care and recovery teams, (5) disease transmission and propagation, and (6) hospital resource utilization. While we acknowledge that transplant fulfills the mission of saving lives, it is imperative to consider the consequences not only to our recipients but also to the community and to health care workers, particularly in the absence of effective preventative or curative therapies. For these reasons, we believe the evidence and risks show that COVID-19 infection should continue to remain a contraindication for donation, as has been the initial response of donation and transplant societies.

The COVID-19 pandemic: A community approach.

An unprecedented global pandemic caused by a novel coronavirus, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has quickly overwhelmed the health care systems worldwide. While there is an absence of consensus among the community in how to manage solid organ transplant recipients and donors, a platform provided by the American Society of Transplantation online community "Outstanding Questions in Transplantation," hosted a collaborative multicenter, multinational discussions to share knowledge in a rapidly evolving global situation. Here, we present a summary of the discussion in addition to the latest published literature.

Importance of incorporating standardized, verifiable, objective metrics of organ procurement organization performance into discussions about organ allocation.

Identifying and supporting specific organ procurement organizations (OPOs) with the greatest opportunity to increase donation rates could significantly increase the number of organs available for transplant. Accomplishing this is complicated by current Scientific Registry of Transplant Recipients/Centers for Medicare & Medicaid Services metrics of donation rates and OPO performance that rely on eligible deaths. These data are self-reported and unverifiable and have been shown to underestimate potential organ donors. We examine the limitations of current OPO performance/donation metrics to inform discussions related to strategies to increase donation. We propose changing to a simple, verifiable, and uniformly applied donation metric. This would allow the transplant community to (1) better understand inherent differences in donor availability based on geography and (2) identify underperforming areas that would benefit from systems improvement agreements to increase donation rates.

Patterns of geographic variability in mortality and eligible deaths between organ procurement organizations.

Eligible deaths are currently used as the denominator of the donor conversion ratio to mitigate the effect of varying mortality patterns in the populations served by different organ procurement organizations (OPOs). Eligible death is an OPO-reported metric rather than a product of formal epidemiological analysis, however, and may be confounded with OPO performance. Using Scientific Registry of Transplant Recipients and Centers for Disease Control and Prevention data, patterns of mortality and eligible deaths within each OPO were analyzed with the use of formal geostatistical analysis to determine whether eligible deaths truly reflect the geographic patterns they are intended to mitigate. There was a 2.1-fold difference in mortality between the OPOs with the highest and lowest rates, with significant positive spatial autocorrelation evident in mortality rates (Moran I = .110; P < .001), meaning geographically proximate OPOs tended to have similar mortality rates. The eligible death ratio demonstrated greater variability, with a 4.5-fold difference between the OPOs with the highest and lowest rates. Contrary to the pattern of mortality rates, the geographic distribution of eligible deaths among OPOs was random (Moran I = -.002; P = .410). This finding suggests geographic patterns do not play a significant role in eligible deaths, thus questioning its continuing use in OPO performance comparisons.

A paper microfluidics-based fluorescent lateral flow immunoassay for point-of-care diagnostics of non-communicable diseases.

In the emergency diagnosis of patients, acute myocardial infarction (AMI) is always time-consuming to diagnose, and the process requires multiple laboratory procedures, expensive equipment and skilled workers. Herein, we developed an easy-to-use, low-cost and portable fluorescent lateral flow immunoassay based on paper microfluidics for the point-of-care diagnostics of non-communicable diseases. The fluorescent lateral flow immunoassay can produce results in less than 10 minutes, and the limit of detection (LOD) is 0.019 ng ml-1. The slope was linear from 0 to 100 ng ml-1; the equation is y = 0.0342e2.1181x and R2 = 0.9618, which are distinctive features that ensure maximum amplification of the signal and recording of quantitative values by an analyser. The detection sensitivity showed an exceptional increase to 0.01 ng ml-1. Compared with conventional bioassay readers, our analyser shows some advantages to easily, clearly and effectively read data. The present point-of-care test for cardiac troponin I decreases the turnaround time and has a high coefficient of variation even at lower concentrations of troponin. So, the development of lateral flow assay-based point-of-care assays with higher analytical performance for real world samples can decrease the rule-out time for AMI in emergency departments and other fields.

Effect of operative duration on infectious complications and mortality following hepatectomy.

BACKGROUND: To study mortality and infectious complications (IC) risk relative to operative duration in a large and contemporary cohort of patients undergoing hepatectomy. METHODS: A retrospective cohort study of 21,443 patients from the National Surgical Quality Improvement Program dataset of patients who underwent liver resection from 2012 to 2016. RESULTS: Patients undergoing hepatectomy during the study period (N = 21,443) had a mean operative duration of 243.5 min of which 16.6% (3533) developed at least one IC. The overall 30-day mortality was 1.6%. A significant increase in mortality and IC was demonstrated from 3 h of operating time (OR: 1.99 and OR: 1.94, respectively), peaking at 8 h (OR: 7.15 and OR: 6.37, respectively). Pneumonia, sepsis/septic shock, and SSI presented high prevalence and were linked to significant mortality. After case-matching, elective hepatectomy was associated with a 4-fold increased risk of infectious complications. CONCLUSIONS: Operative duration was associated with a linear increased risk of mortality and IC after hepatectomy. The most critical determinants of IC were ASA class, COPD, CHF, and type of hepatectomy.

Long term outcomes of patients transplanted for hepatocellular carcinoma with human immunodeficiency virus infection.

BACKGROUND: We aimed to study outcomes in HIV + patients with HCC in the US following Liver Transplantation (LT) using the UNOS dataset. METHODS: The database was queried from 2003 to 2016 for patients undergoing LT with HCC, HIV+, and HCC/HIV+. RESULTS: Out of 17,397 LT performed for HCC during the study period, 113 were transplanted for HCC with HIV infection (91 isolated livers). Patients transplanted for HCC/HIV+ were younger (55.54 ± 5.89 vs 58.80 ± 7.37, p < 0.001), had lower total bilirubin (1.20 vs 1.60, p = 0.042) significantly lower BMI (25.35 ± 4.43 vs 28.39 ± 5.17, p < 0.001) and were more likely to be co-infected with HBV (25.3% vs 8.2% p < 0.001) than those transplanted for HCC alone. HCC/HIV + patients were found to have a 3.8 fold increased risk of peri-operative mortality at 90 days after matching. HCC/HIV + recipients had 54% decreased long-term survival within the HCC cohort. Our initial analysis of overall graft and patient survival found significant differences between HCC/HIV and HCC/HIV + recipients. However, these variances were lost after case-matching. Recurrence and disease free survival were similar in HCC alone vs HCC/HIV + recipients. CONCLUSIONS: Our analysis suggests that excellent outcomes can be achieved in selected patients with HCC/HIV+.

A randomized, phase 2 study of ASP0113, a DNA-based vaccine, for the prevention of CMV in CMV-seronegative kidney transplant recipients receiving a kidney from a CMV-seropositive donor.

Cytomegalovirus (CMV) is a latent infection in most infected individuals, but can be pathogenic in immunocompromised kidney transplant recipients. ASP0113 is a DNA-based vaccine for the prevention of CMV-related mortality and end-organ disease in transplant recipients. The efficacy, safety, and immunogenicity of ASP0113 was assessed in a phase 2, double-blind, placebo-controlled study in CMV-seronegative kidney transplant recipients receiving a kidney from a CMV-seropositive donor. Transplant recipients were randomized (1:1) to receive 5 doses of ASP0113 (5 mg; n = 75) or placebo (n = 74) on Days 30/60/90/120/180 posttransplant, and they received prophylactic valganciclovir/ganciclovir 10-100 days posttransplant. The primary endpoint was the proportion of transplant recipients with CMV viremia ≥1000 IU/mL from Day 100 through to 1 year after the first study vaccine injection. There was no statistically significant difference in the primary endpoint between the ASP0113 and placebo groups (odds ratio 0.79, 95% confidence interval 0.43-1.47; P = .307). There were similar numbers of transplant recipients with treatment-emergent adverse events between groups; however, more transplant recipients reported injection site pain in the ASP0113 group compared with placebo. ASP0113 did not demonstrate efficacy in the prevention of CMV viremia in this CMV-seronegative kidney transplant population, but demonstrated a safety profile similar to placebo. ClinicalTrials.gov registration number: NCT01974206.

Implementation of the Treat All Policy Among Persons with HIV Infection Enrolled in Care But Not on Antiretroviral Therapy - India, May 2017-June 2018.

Since September 2015, the World Health Organization has recommended antiretroviral therapy (ART) for all persons with human immunodeficiency virus (HIV) infection, regardless of clinical stage or CD4 count (1). This Treat All policy was based on evidence that ART initiation early in HIV infection as opposed to waiting for the CD4 count to decline to certain levels (e.g., <500 cells/mm3, per previous guidelines), was associated with reduced morbidity, mortality, and HIV transmission (2-4). Further, approximately half of persons enrolled in non-ART care that included monitoring for HIV disease progression (i.e., in pre-ART care) were lost to follow-up before becoming ART-eligible (5). India, the country with the third largest number of persons with HIV infection in the world (2.1 million), adopted the Treat All policy on April 28, 2017. This report describes implementation of Treat All during May 2017-June 2018, by India's National AIDS Control Organization (NACO) and partners, by facilitating ART initiation among persons previously in pre-ART care at 46 ART centers supported by the U.S. President's Emergency Plan for AIDS Relief (PEPFAR)* in six districts in the states of Maharashtra and Andhra Pradesh. Partners supported these 46 ART centers in identifying and attempting to contact persons who were enrolled in pre-ART care during January 2014-April 2017, and educating those reached about Treat All. ART center-based records were used to monitor implementation indicators, including ART initiation. A total of 9,898 (39.6%) of 25,007 persons previously enrolled in pre-ART care initiated ART; among these 9,898 persons, 6,315 (63.8%) initiated ART after being reached during May 2017-June 2018, including 1,635 (16.5%) who had been lost to follow-up before ART initiation. NACO scaled up efforts nationwide to build ART centers' capacity to implement Treat All. Active tracking and tracing of persons with HIV infection enrolled in care but not on ART, combined with education about the benefits of early HIV treatment, can facilitate ART initiation.