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Background Bloodstream infections remain one of the major challenges in intensive care units, leading to sepsis or even septic shock in many cases. Due to the lack of timely diagnostic approaches with sufficient sensitivity, mortality rates of sepsis are still unacceptably high. However a prompt diagnosis of the causative microorganism is critical to significantly improve outcome of bloodstream infections. Although various targeted molecular tests for blood samples are available, time-consuming blood culture-based approaches still represent the standard of care for the identification of bacteria. Methods Here we describe the establishment of a complete diagnostic workflow for the identification of infectious microorganisms from seven septic patients based on unbiased sequence analyses of free circulating DNA from plasma by next-generation sequencing. Results We found significant levels of DNA fragments derived from pathogenic bacteria in samples from septic patients. Quantitative evaluation of normalized read counts and introduction of a sepsis indicating quantifier (SIQ) score allowed for an unambiguous identification of Gram-positive as well as Gram-negative bacteria that exactly matched with blood cultures from corresponding patient samples. In addition, we also identified species from samples where blood cultures were negative. Reads of non-human origin also comprised fragments derived from antimicrobial resistance genes, showing that, in principle, prediction of specific types of resistance might be possible. Conclusions The complete workflow from sample preparation to species identification report could be accomplished in roughly 30 h, thus making this approach a promising diagnostic platform for critically ill patients suffering from bloodstream infections.

Sepsis is a frequently fatal condition characterized by an uncontrolled and harmful host reaction to microbial infection. Despite the prevalence and severity of sepsis, we lack a fundamental grasp of its pathophysiology. Here we report that the cytokine interleukin-3 (IL-3) potentiates inflammation in sepsis. Using a mouse model of abdominal sepsis, we showed that innate response activator B cells produce IL-3, which induces myelopoiesis of Ly-6Chigh monocytes and neutrophils and fuels a cytokine storm. IL-3 deficiency protects mice against sepsis. In humans with sepsis, high plasma IL-3 levels are associated with high mortality even after adjusting for prognostic indicators. This study deepens our understanding of immune activation, identifies IL-3 as an orchestrator of emergency myelopoiesis, and reveals a new therapeutic target for treating sepsis.

Cytotoxic T cells are responsible for the elimination of infected cells and are key players in the control of viruses. CD8 + T cells with an effector phenotype express cytotoxic molecules and are able to perform target cell killing. COVID-19 patients with a mild disease course were analyzed for the differentiation status and cytotoxic profile of CD8 + T cells. SARS-CoV-2 infection induced a vigorous cytotoxic CD8 + T cell response. However, this cytotoxic profile of T cells was not detected in COVID-19 patients over the age of 80 years. Thus, the absence of a cytotoxic response in elderly patients might be a possible reason for the more frequent severity of COVID-19 in this age group than in younger patients. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection induces a T cell response that most likely contributes to virus control in COVID-19 patients but may also induce immunopathology. Until now, the cytotoxic T cell response has not been very well characterized in COVID-19 patients. Here, we analyzed the differentiation and cytotoxic profile of T cells in 30 cases of mild COVID-19 during acute infection. SARS-CoV-2 infection induced a cytotoxic response of CD8 + T cells, but not CD4 + T cells, characterized by the simultaneous production of granzyme A and B as well as perforin within different effector CD8 + T cell subsets. PD-1-expressing CD8 + T cells also produced cytotoxic molecules during acute infection, indicating that they were not functionally exhausted. However, in COVID-19 patients over the age of 80 years, the cytotoxic T cell potential was diminished, especially in effector memory and terminally differentiated effector CD8 + cells, showing that elderly patients have impaired cellular immunity against SARS-CoV-2. Our data provide valuable information about T cell responses in COVID-19 patients that may also have important implications for vaccine development. IMPORTANCE Cytotoxic T cells are responsible for the elimination of infected cells and are key players in the control of viruses. CD8 + T cells with an effector phenotype express cytotoxic molecules and are able to perform target cell killing. COVID-19 patients with a mild disease course were analyzed for the differentiation status and cytotoxic profile of CD8 + T cells. SARS-CoV-2 infection induced a vigorous cytotoxic CD8 + T cell response. However, this cytotoxic profile of T cells was not detected in COVID-19 patients over the age of 80 years. Thus, the absence of a cytotoxic response in elderly patients might be a possible reason for the more frequent severity of COVID-19 in this age group than in younger patients.

The complex process of manual biomarker extraction from body composition analysis (BCA) has far restricted the analysis of SARS-CoV-2 outcomes to small patient cohorts and a limited number of tissue types. We investigate the association of two BCA-based biomarkers with the development of severe SARS-CoV-2 infections for 918 patients (354 female, 564 male) regarding disease severity and mortality (186 deceased). Multiple tissues, such as muscle, bone, or adipose tissue are used and acquired with a deep-learning-based, fully-automated BCA from computed tomography images of the chest. The BCA features and markers were univariately analyzed with a Shapiro–Wilk and two-sided Mann–Whitney-U test. In a multivariate approach, obtained markers were adjusted by a defined set of laboratory parameters promoted by other studies. Subsequently, the relationship between the markers and two endpoints, namely severity and mortality, was investigated with regard to statistical significance. The univariate approach showed that the muscle volume was significant for female ( p severity  ≤ 0.001, p mortality  ≤ 0.0001) and male patients ( p severity  = 0.018, p mortality  ≤ 0.0001) regarding the severity and mortality endpoints. For male patients, the intra- and intermuscular adipose tissue (IMAT) ( p  ≤ 0.0001), epicardial adipose tissue (EAT) ( p  ≤ 0.001) and pericardial adipose tissue (PAT) ( p  ≤ 0.0001) were significant regarding the severity outcome. With the mortality outcome, muscle ( p  ≤ 0.0001), IMAT ( p  ≤ 0.001), EAT ( p  = 0.011) and PAT ( p  = 0.003) remained significant. For female patients, bone ( p  ≤ 0.001), IMAT ( p  = 0.032) and PAT ( p  = 0.047) were significant in univariate analyses regarding the severity and bone ( p  = 0.005) regarding the mortality. Furthermore, the defined sarcopenia marker ( p  ≤ 0.0001, for female and male) was significant for both endpoints. The cardiac marker was significant for severity (p female  = 0.014, p male  ≤ 0.0001) and for mortality (p female  ≤ 0.0001, p male  ≤ 0.0001) endpoint for both genders. The multivariate logistic regression showed that the sarcopenia marker was significant ( p severity  = 0.006, p mortality  = 0.002) for both endpoints (OR severity  = 0.42, 95% CI severity : 0.23–0.78, OR mortality  = 0.34, 95% CI mortality : 0.17–0.67). The cardiac marker showed significance (p = 0.018) only for the severity endpoint (OR = 1.42, 95% CI 1.06–1.90). The association between BCA-based sarcopenia and cardiac biomarkers and disease severity and mortality suggests that these biomarkers can contribute to the risk stratification of SARS-CoV-2 patients. Patients with a higher cardiac marker and a lower sarcopenia marker are at risk for a severe course or death. Whether those biomarkers hold similar importance for other pneumonia-related diseases requires further investigation.

In order to identify biomarkers for earlier prediction of COVID-19 outcome, we collected blood samples from patients with fatal outcomes (non-survivors) and with positive clinical outcomes (survivors) at ICU admission and after seven days. COVID-19 survivors and non-survivors showed significantly different transcript levels for 93 genes in whole blood already at ICU admission as revealed by RNA-Seq. These differences became even more pronounced at day 7, resulting in 290 differentially expressed genes. Many identified genes play a role in the differentiation of hematopoietic cells. For validation, we designed an RT-qPCR assay for C-type lectin domain family 12 member A ( CLEC12A ) and acetylcholinesterase ( ACHE ), two transcripts that showed highest potential to discriminate between survivors and non-survivors at both time points. Using our combined RT-qPCR assay we examined 33 samples to accurately predict patient survival with an AUROC curve of 0.931 (95% CI = 0.814–1.000) already at ICU admission. CLEC12A and ACHE showed improved prediction of patient outcomes compared to standard clinical biomarkers including CRP and PCT in combination (AUROC = 0.403, 95% CI = 0.108–0.697) or SOFA score (AUROC = 0.701 95% CI = 0.451–0.951) at day 0. Therefore, analyzing CLEC12A and ACHE gene expression from blood may provide a promising approach for early risk stratification of severely ill COVID-19 patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a worldwide health threat. In a prospective multicentric study, we identify IL-3 as an independent prognostic marker for the outcome during SARS-CoV-2 infections. Specifically, low plasma IL-3 levels is associated with increased severity, viral load, and mortality during SARS-CoV-2 infections. Patients with severe COVID-19 exhibit also reduced circulating plasmacytoid dendritic cells (pDCs) and low plasma IFNα and IFNλ levels when compared to non-severe COVID-19 patients. In a mouse model of pulmonary HSV-1 infection, treatment with recombinant IL-3 reduces viral load and mortality. Mechanistically, IL-3 increases innate antiviral immunity by promoting the recruitment of circulating pDCs into the airways by stimulating CXCL12 secretion from pulmonary CD123 + epithelial cells, both, in mice and in COVID-19 negative patients exhibiting pulmonary diseases. This study identifies IL-3 as a predictive disease marker for SARS-CoV-2 infections and as a potential therapeutic target for pulmunory viral infections. Here, the authors identify interleukin-3 as a predictive marker for severity and outcome of SARS-CoV-2 infection in a multi-center, prospective study and find that patients with severe COVID-19 have reduced circulating plasmacytoid dendritic cell levels compared to non-severe COVID-19 patients.

Extracorporeal membrane oxygenation (ECMO) is widely used to manage acute respiratory distress syndrome (ARDS); however, biofilm formation on cannulas may contribute to infections. This study investigated the prevalence, timing, and microbial profiles of infectious complications following ECMO decannulation. This prospective, single-center cohort study was conducted in the mixed medical-surgical intensive care unit (ICU) at the University Hospital Essen (01/2022–01/2023). Adults who received ECMO for > 48 h were included. Microbiological sampling (blood cultures and wound swabs) was performed before and after decannulation. Cannulas were assessed for biofilms, and plasma samples were analyzed using next-generation sequencing (NGS) of microbial cell-free DNA (cfDNA). Sepsis was defined according to the most recent Sepsis-3 criteria. The study included 18 patients (56% men); 17 received VV-ECMO and one received VA-ECMO. Post-decannulation sepsis occurred in 10 of 18 patients (56%; 95% CI: 31–79%), and overall infectious complications were observed in 72% of patients. A strong negative correlation was evident between pre-ECMO ventilation duration and biofilm-forming bacteria in the blood during ECMO ( r = − 0.7, p  = 0.002). Blood cultures obtained within 10 min of decannulation were positive in 6 of 18 (33%) patients. NGS identified pathogens in 9 of 12 patients (75%), with 5 (42%) revealing additional organisms not detected by conventional methods. Viral pathogens were detected in 3 of 12 (25%) patients using NGS. Patients with sepsis demonstrated higher antibiotic consumption ( p  = 0.012) and more frequently met SOFA-based sepsis criteria ( p  = 0.007), whereas other parameters were comparable. Sepsis and infection frequently occur after ECMO decannulation and may be associated with biofilm-related pathogens. NGS has improved pathogen detection beyond standard diagnostics, although the findings require clinical correlation. These prospective findings support the need for further investigation of advanced integrated microbiological surveillance post-ECMO in larger multicenter studies. Trial registration : DRKS, DRKS00024842. Registered 12 Apr 2021, https://drks.de/search/de/trial/DRKS00024842 .

We describe screening results for detection of co-infections with Legionella pneumophila in patients infected with severe acute respiratory syndrome coronavirus 2. In total, 93 patients were tested; 1 was positive (1.1%) for L. pneumophila serogroup 1. Co-infections with L. pneumophila occur in coronavirus disease patients and should not be missed.

Invasive infections with Aspergillus fumigatus in ICU patients are linked to high morbidity and mortality. Diagnosing invasive pulmonary aspergillosis (IPA) in non-immunosuppressed patients is difficult, as Aspergillus antigen (galactomannan [GM]) may have other causes. This retrospective study analyzed 160 ICU surgical patients with positive GM in broncho-alveolar lavage fluid (BALF), classifying them based on AspICU criteria for suspected IPA (pIPA) or aspiration. Patients with pIPA had higher disease severity than those with aspiration, including higher dialysis rates, organ transplantation, corticosteroid use, and Sequential Organ Failure Assessment (SOFA) score. Aspergillus culture was positive in 47.0% of pIPA cases but only 2.6% of aspiration cases ( p  < 0.001). SOFA score at first positive GM in BALF independently predicted 28-day mortality. In surgical patients with a positive GM in BALF, aspiration is more likely if there’s no corticosteroid therapy, negative Aspergillus culture, and a history of aspiration events. Diagnosis of pIPA requires Aspergillus culture or prior corticosteroid therapy in this cohort of critically ill patients.

Background Extracorporeal membrane oxygenation (ECMO) cannulas are potential reservoirs for pathogens, yet their role in bacteremia and sepsis following decannulation remains poorly understood. This proof-of-concept study aims to characterize bacterial colonization of ECMO cannulas, identify potential sources of these bacteria, and assess their association with post-decannulation bloodstream infections and sepsis. Methods We conducted a single-center observational study including 10 patients receiving venovenous ECMO support between January 2022 and January 2023. Microbial colonization of cannulas, skin sites, and plasma was analyzed using culture-based methods and 16S rDNA amplicon sequencing. Alpha and beta diversity analyses were performed, and findings were correlated with clinical outcomes, including sepsis and bacteremia. Results A total of 117 samples yielded ~ 11 million sequencing reads. Bacteria colonizing ECMO cannulas matched pathogens causing prior bacteremia during ECMO support in all affected patients. Bacteria detected on cannulas and insertion sites were frequently recovered in plasma following decannulation. Notably, 16S rDNA analysis detected circulating pathogens that conventional cultures missed, often those from prior infections that were thought to be eradicated by antibiotics. Patients who developed sepsis post-decannulation exhibited higher bacterial diversity on cannulas and a higher overall abundance of Pseudomonas , while non-septic patients had greater Enterococcus abundance. Conclusions Our results confirm that ECMO cannulas serve as pathogen reservoirs, with decannulation enabling bacterial translocation into the bloodstream and contributing to post-decannulation sepsis. 16S rDNA sequencing exhibited greater sensitivity than cultures for detecting bloodstream pathogens. These findings support re-assessment of prophylactic measures during ECMO decannulation and lay the groundwork for developing early sepsis risk stratification tools.