Advancing COVID-19 diagnostics:rapid detection of intact SARS-CoV-2 using viability RT-PCR assay

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19). Commonly used methods for both clinical diagnosis of SARS-CoV-2 infection and management of infected patients involve the detection of viral RNA, but the presence of infectious virus particles is unknown. Viability PCR (v-PCR) uses a photoreactive dye to bind non-infectious RNA, ideally resulting in the detection of RNA only from intact virions. This study aimed to develop and validate a rapid v-PCR assay for distinguishing intact and compromised SARS-CoV-2. Propidium monoazide (PMAxx) was used as a photoreactive dye. Mixtures with decreasing percentages of intact SARS-CoV-2 (from 100% to 0%) were prepared from SARS-CoV-2 virus stock and a clinical sample. Each sample was divided into a PMAxx-treated part and a non-PMAxx-treated part. Reverse transcription-PCR (RT-PCR) using an in-house developed SARS-CoV-2 viability assay was then applied to both sample sets. The difference in intact SARS-CoV-2 was determined by subtracting the cycle threshold (Ct) value of the PMAxx-treated sample from the non-PMAxx-treated sample. Mixtures with decreasing concentrations of intact SARS-CoV-2 showed increasingly lower delta Ct values as the percentage of intact SARS-CoV-2 decreased, as expected. This relationship was observed in both high and low viral load samples prepared from cultured SARS-CoV-2 virus stock, as well as for a clinical sample prepared directly from a SARS-CoV-2 positive nasopharyngeal swab. In this study, a rapid v-PCR assay has been validated that can distinguish intact from compromised SARS-CoV-2. The presence of intact virus particles, as determined by v-PCR, may indicate SARS-CoV-2 infectiousness.</p

Variables associated with in-hospital and postdischarge outcomes after postcardiotomy extracorporeal membrane oxygenation:Netherlands Heart Registration Cohort

Objectives: Extracorporeal membrane oxygenation (ECMO) for postcardiotomy cardiogenic shock has been increasingly used without concomitant mortality reduction. This study aims to investigate determinants of in-hospital and postdischarge mortality in patients requiring postcardiotomy ECMO in the Netherlands. Methods: The Netherlands Heart Registration collects nationwide prospective data from cardiac surgery units. Adults receiving intraoperative or postoperative ECMO included in the register from January 2013 to December 2019 were studied. Survival status was established through the national Personal Records Database. Multivariable logistic regression analyses were used to investigate determinants of in-hospital (3 models) and 12-month postdischarge mortality (4 models). Each model was developed to target specific time points during a patient's clinical course. Results: Overall, 406 patients (67.2% men, median age, 66.0 years [interquartile range, 55.0-72.0 years]) were included. In-hospital mortality was 51.7%, with death occurring in a median of 5 days (interquartile range, 2-14 days) after surgery. Hospital survivors (n = 196) experienced considerable rates of pulmonary infections, respiratory failure, arrhythmias, and deep sternal wound infections during a hospitalization of median 29 days (interquartile range, 17-51 days). Older age (odds ratio [OR], 1.02; 95% CI, 1.0-1.04) and preoperative higher body mass index (OR, 1.08; 95% CI, 1.02-1.14) were associated with in-hospital death. Within 12 months after discharge, 35.1% of hospital survivors (n = 63) died. Postoperative renal failure (OR, 2.3; 95% CI, 1.6-4.9), respiratory failure (OR, 3.6; 95% CI, 1.3-9.9), and re-thoracotomy (OR, 2.9; 95% CI, 1.3-6.5) were associated with 12-month postdischarge mortality. Conclusions: In-hospital and postdischarge mortality after postcardiotomy ECMO in adults remains high in the Netherlands. ECMO support in patients with higher age and body mass index, which drive associations with higher in-hospital mortality, should be carefully considered. Further observations suggest that prevention of re-thoracotomies, renal failure, and respiratory failure are targets that may improve postdischarge outcomes

New clinical prediction model for early recognition of sepsis in adult primary care patients:a prospective diagnostic cohort study of development and external validation

Background Recognising patients who need immediate hospital treatment for sepsis while simultaneously limiting unnecessary referrals is challenging for GPs.Aim To develop and validate a sepsis prediction model for adult patients in primary care.Design and setting This was a prospective cohort study in four out-of-hours primary care services in the Netherlands, conducted between June 2018 and March 2020.Method Adult patients who were acutely ill and received home visits were included. A total of nine clinical variables were selected as candidate predictors, next to the biomarkers C-reactive protein, procalcitonin, and lactate. The primary endpoint was sepsis within 72 hours of inclusion, as established by an expert panel. Multivariable logistic regression with backwards selection was used to design an optimal model with continuous clinical variables. The added value of the biomarkers was evaluated. Subsequently, a simple model using single cut-off points of continuous variables was developed and externally validated in two emergency department populations.Results A total of 357 patients were included with a median age of 80 years (interquartile range 71–86), of which 151 (42%) were diagnosed with sepsis. A model based on a simple count of one point for each of six variables (aged &gt;65 years; temperature &gt;38°C; systolic blood pressure ≤110 mmHg; heart rate &gt;110/min; saturation ≤95%; and altered mental status) had good discrimination and calibration (C-statistic of 0.80 [95% confidence interval = 0.75 to 0.84]; Brier score 0.175). Biomarkers did not improve the performance of the model and were therefore not included. The model was robust during external validation.Conclusion Based on this study’s GP out-of-hours population, a simple model can accurately predict sepsis in acutely ill adult patients using readily available clinical parameters

Neurologic complications in patients receiving aortic versus subclavian versus femoral arterial cannulation for post-cardiotomy extracorporeal life support:results of the PELS observational multicenter study

BACKGROUND: Cerebral perfusion may change depending on arterial cannulation site and may affect the incidence of neurologic adverse events in post-cardiotomy extracorporeal life support (ECLS). The current study compares patients' neurologic outcomes with three commonly used arterial cannulation strategies (aortic vs. subclavian/axillary vs. femoral artery) to evaluate if each ECLS configuration is associated with different rates of neurologic complications. METHODS: This retrospective, multicenter (34 centers), observational study included adults requiring post-cardiotomy ECLS between January 2000 and December 2020 present in the Post-Cardiotomy Extracorporeal Life Support (PELS) Study database. Patients with Aortic, Subclavian/Axillary and Femoral cannulation were compared on the incidence of a composite neurological end-point (ischemic stroke, cerebral hemorrhage, brain edema). Secondary outcomes were overall in-hospital mortality, neurologic complications as cause of in-hospital death, and post-operative minor neurologic complications (seizures). Association between cannulation and neurological outcomes were investigated through linear mixed-effects models. RESULTS: This study included 1897 patients comprising 26.5% Aortic (n = 503), 20.9% Subclavian/Axillary (n = 397) and 52.6% Femoral (n = 997) cannulations. The Subclavian/Axillary group featured a more frequent history of hypertension, smoking, diabetes, previous myocardial infarction, dialysis, peripheral artery disease and previous stroke. Neuro-monitoring was used infrequently in all groups. Major neurologic complications were more frequent in Subclavian/Axillary (Aortic: n = 79, 15.8%; Subclavian/Axillary: n = 78, 19.6%; Femoral: n = 118, 11.9%; p &lt; 0.001) also after mixed-effects model adjustment (OR 1.53 [95% CI 1.02-2.31], p = 0.041). Seizures were more common in Subclavian/Axillary (n = 13, 3.4%) than Aortic (n = 9, 1.8%) and Femoral cannulation (n = 12, 1.3%, p = 0.036). In-hospital mortality was higher after Aortic cannulation (Aortic: n = 344, 68.4%, Subclavian/Axillary: n = 223, 56.2%, Femoral: n = 587, 58.9%, p &lt; 0.001), as shown by Kaplan-Meier curves. Anyhow, neurologic cause of death (Aortic: n = 12, 3.9%, Subclavian/Axillary: n = 14, 6.6%, Femoral: n = 28, 5.0%, p = 0.433) was similar. CONCLUSIONS:In this analysis of the PELS Study, Subclavian/Axillary cannulation was associated with higher rates of major neurologic complications and seizures. In-hospital mortality was higher after Aortic cannulation, despite no significant differences in incidence of neurological cause of death in these patients. These results encourage vigilance for neurologic complications and neuromonitoring use in patients on ECLS, especially with Subclavian/Axillary cannulation.</p

Characteristics and Outcomes of Prolonged Venoarterial Extracorporeal Membrane Oxygenation after Cardiac Surgery:The Post-Cardiotomy Extracorporeal Life Support (PELS-1) Cohort Study

OBJECTIVES: Most post-cardiotomy (PC) extracorporeal membrane oxygenation (ECMO) runs last less than 7 days. Studies on the outcomes of longer runs have provided conflicting results. This study investigates patient characteristics and short- and long-term outcomes in relation to PC ECMO duration, with a focus on prolonged (&gt; 7 d) ECMO. DESIGN: Retrospective observational cohort study. SETTING: Thirty-four centers from 16 countries between January 2000 and December 2020. PATIENTS: Adults requiring post PC ECMO between 2000 and 2020. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Characteristics, in-hospital, and post-discharge outcomes were compared among patients categorized by ECMO duration. Survivors and nonsurvivors were compared in the subgroup of patients with ECMO duration greater than 7 days. The primary outcome was in-hospital mortality. Two thousand twenty-one patients were included who required PC ECMO for 0-3 days (n = 649 [32.1%]), 4-7 days (n = 776 [38.3%]), 8-10 days (n = 263 [13.0%]), and greater than 10 days (n = 333 [16.5%]). There were no major differences in the investigated preoperative and procedural characteristics among ECMO duration groups. However, the longer ECMO duration category was associated with multiple complications including bleeding, acute kidney injury, arrhythmias, and sepsis. Hospital mortality followed a U-shape curve, with lowest mortality in patients with ECMO duration of 4-7 days (n = 394, 50.8%) and highest in patients with greater than 10 days ECMO support (n = 242, 72.7%). There was no significant difference in post-discharge survival between ECMO duration groups. In patients with ECMO duration greater than 7 days, age, comorbidities, valvular diseases, and complex procedures were associated with nonsurvival. CONCLUSIONS: Nearly 30% of PC ECMO patients were supported for greater than 7 days. In-hospital mortality increased after 7 days of support, especially in patients undergoing valvular and complex surgery, or who had complications, although the long-term post-discharge prognosis was comparable to PC ECMO patients with shorter support duration.</p

Systemic inflammation down-regulates glyoxalase-1 expression: an experimental study in healthy males

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The CoLab score is associated with SARS-CoV-2 viral load during admission in individuals admitted to the intensive care unit:the CoLaIC cohort study

ObjectivesThe present study examines the temporal association between the changes in SARS-CoV-2 viral load during infection and whether the CoLab-score can facilitate de-isolation.MethodsNasal swabs and blood samples were collected from ICU-admitted SARS-CoV-2 positive patients at Maastricht UMC+ from March 25, 2020 to October 1, 2021. The CoLab-score was calculated based on 10 blood parameters and age and can range from −43 to 6. Three mixed effects analyses compared patient categories based on initial PCR Ct values (low; Ct≤20, mid; 20&gt;Ct≤30, high; Ct&gt;30), serial PCR Ct values to CoLab-scores over time, and the association between within-patient delta Ct values and CoLab-scores.ResultsIn 324 patients, the median Ct was 33, and the median CoLab-score was −1.78. Mid (n=110) and low (n=41) Ct-categories had higher CoLab-scores over time (+0.60 points, 95 % CI; 0.04–1.17, and +0.28 points, 95 % CI −0.49 to 1.04) compared to the high Ct (n=87) category. Over time, higher serial Ct values were associated with lower serial CoLab-scores, decreasing by −0.07 points (95 % CI; −0.11 to −0.02) per day. Increasing delta Ct values were associated with a decreasing delta CoLab-score of −0.12 (95 % CI; −0.23; −0.01).ConclusionsThe study found an association between lower viral load on admission and reduced CoLab-score. Additionally, a decrease in viral load over time was associated with a decrease in CoLab-score. Therefore, the CoLab-score may make patient de-isolation an option based on the CoLab-score

External Validation of Mortality Prediction Models for Critical Illness Reveals Preserved Discrimination but Poor Calibration

OBJECTIVES: In a recent scoping review, we identified 43 mortality prediction models for critically ill patients. We aimed to assess the performances of these models through external validation. DESIGN: Multicenter study. SETTING: External validation of models was performed in the Simple Intensive Care Studies-I (SICS-I) and the Finnish Acute Kidney Injury (FINNAKI) study. PATIENTS: The SICS-I study consisted of 1,075 patients, and the FINNAKI study consisted of 2,901 critically ill patients. MEASUREMENTS AND MAIN RESULTS: For each model, we assessed: 1) the original publications for the data needed for model reconstruction, 2) availability of the variables, 3) model performance in two independent cohorts, and 4) the effects of recalibration on model performance. The models were recalibrated using data of the SICS-I and subsequently validated using data of the FINNAKI study. We evaluated overall model performance using various indexes, including the (scaled) Brier score, discrimination (area under the curve of the receiver operating characteristics), calibration (intercepts and slopes), and decision curves. Eleven models (26%) could be externally validated. The Acute Physiology And Chronic Health Evaluation (APACHE) II, APACHE IV, Simplified Acute Physiology Score (SAPS)-Reduced (SAPS-R)‚ and Simplified Mortality Score for the ICU models showed the best scaled Brier scores of 0.11‚ 0.10‚ 0.10‚ and 0.06‚ respectively. SAPS II, APACHE II, and APACHE IV discriminated best; overall discrimination of models ranged from area under the curve of the receiver operating characteristics of 0.63 (0.61-0.66) to 0.83 (0.81-0.85). We observed poor calibration in most models, which improved to at least moderate after recalibration of intercepts and slopes. The decision curve showed a positive net benefit in the 0-60% threshold probability range for APACHE IV and SAPS-R. CONCLUSIONS: In only 11 out of 43 available mortality prediction models, the performance could be studied using two cohorts of critically ill patients. External validation showed that the discriminative ability of APACHE II, APACHE IV, and SAPS II was acceptable to excellent, whereas calibration was poor

Applications of rotational thromboelastometry in heparin monitoring in critical COVID-19 disease: Observations in the Maastricht Intensive Care COVID cohort

Background: Critically ill COVID-19 patients are at risk for venous thromboembolism (VTE). Therefore, they receive thromboprophylaxis and, when appropriate, therapeutic unfractionated heparin (UFH) or low-molecular-weight heparin (LMWH). To monitor heparins in COVID-19 disease, whole-blood rotational thromboelastometry (ROTEM) may be a promising alternative to the aPTT and anti-Xa assays. Objective: To evaluate the ROTEM INTEM/HEPTEM ratios in mechanically ventilated COVID-19 patients treated with UFH and therapeutic LMWH. Material and methods: A subcohort of mechanically ventilated COVID-19 patients of the prospective Maastricht Intensive Care Covid (MaastrICCht) cohort was studied. Anti-Xa, aPTT, and ROTEM measurements following treatment with UFH or therapeutic dose of LMWH (nadroparin) were evaluated using uni- and multivariable linear regression analysis and receiver operating characteristics. Results: A total of 98 patients were included, of which 82 were treated with UFH and 16 with therapeutic LMWH. ROTEM-measured INTEM/HEPTEM CT ratio was higher in patients using UFH (1.4 [1.3–1.4]) compared to patients treated with LMWH (1.0 [1.0–1.1], p < 0.001). Both the aPTT and anti-Xa were associated with the CT ratio. However, the β-regression coefficient (95%CI) was significantly higher in patients on UFH (0.31 (0.001–0.62)) compared to therapeutic LMWH (0.09 (0.05–0.13)) for comparison with the anti-Xa assay. Furthermore, ROC analysis demonstrated an area under the curve for detecting UFH of 0.936(0.849–1.00), 0.851(0.702–1.000), and 0.645(0.465–0.826) for the CT ratio, aPTT, and anti-Xa, respectively. Conclusion: The ROTEM INTEM/HEPTEM CT ratio appears a promising tool to guide anticoagulant therapy in ICU patients with COVID-19 disease, but associations with clinical endpoints are currently lacking

Presence and impact of anemia in patients supported with left ventricular assist devices

Background: Data on anemia and its effects on patients supported with continuous-flow left ventricular assist devices (LVADs) are lacking. Objectives: This study sought to describe the presence of anemia over time and investigate its association with mortality, quality of life, exercise capacity, and adverse events in LVAD patients. Methods: Adults receiving durable LVADs between 2008 and 2017 were identified from the INTERMACS database. The full cohort was stratified according to anemia severity (no anemia, mild, and moderate-severe). Results: The analysis of 19,509 patients (females: 21.2%, age: 56.9 ± 12.9 years) showed that moderate-severe anemia affected 45.2% of patients at baseline, 33.5% of them at 6 months, and 32.3% in the fourth year after implantation. The presence of normal hemoglobin was 24.4% before surgery, 32.5% at 6 months, and 36.6% at 4 years after implantation. Multivariable linear mixed-effect regression revealed that the average hemoglobin over time was significantly lower (ß, -0.233, 95% confidence interval (CI): -0.282 to -0.185), and the reduction of hemoglobin over time was bigger (ß, -0.032 95% CI: -0.035 to -0.028) for LVAD nonsurvivors compared with LVAD survivors. Adjusted Cox regression showed that the severity of preimplant anemia was associated with higher mortality (HR, mild: 1.19; 95% CI: 1.05-1.35 and moderate-severe: 1.44; 95% CI: 1.28-1.62), with similar results in competing risk regression. Anemia progression during follow-up was associated with decreased Kansas City Cardiomyopathy Questionnaire scores and shorter 6-minute walk distances. Conclusions: In patients supported with LVADs, anemia is a frequent comorbidity, and deterioration over time is associated with poor prognosis