Daily Sepsis Research Analysis
Three papers advanced sepsis research today: a validated low-volume dual RNA-seq workflow enabling simultaneous host–pathogen transcriptomics from clinical blood, a large real-world cohort showing mortality and length-of-stay benefits of COVID-19 antivirals in viral sepsis, and a multi-omic analysis linking ARDS/sepsis inflammatory phenotypes to mortality via distinct metabolic-immune pathways.
Summary
Three papers advanced sepsis research today: a validated low-volume dual RNA-seq workflow enabling simultaneous host–pathogen transcriptomics from clinical blood, a large real-world cohort showing mortality and length-of-stay benefits of COVID-19 antivirals in viral sepsis, and a multi-omic analysis linking ARDS/sepsis inflammatory phenotypes to mortality via distinct metabolic-immune pathways.
Research Themes
- Host–pathogen transcriptomics in clinical sepsis
- Antiviral therapy effectiveness in viral sepsis
- Precision phenotyping and metabolic-immune mechanisms in ARDS/sepsis
Selected Articles
1. Dual RNA isolation from blood: an optimized protocol for host and bacterial RNA purification for dual RNA-sequencing analysis in whole blood sepsis samples.
The authors present DRIB, a clinically compatible workflow that simultaneously stabilizes and purifies host and bacterial RNA from just 0.5 mL of whole blood, enabling dual RNA-seq in sepsis. In a pilot cohort, dual rRNA depletion and sequencing produced 16.6–24.8 million reads/sample with ~63% unique mapping, capturing both host (51–68% of reads) and bacterial (0.5–6.7%) transcripts and revealing immune metabolic and metal-binding pathways.
Impact: This methodological advance removes a key barrier to studying in vivo host–pathogen interactions in human sepsis from low-volume blood, enabling discovery of actionable biomarkers and mechanisms.
Clinical Implications: While not immediately practice-changing, this workflow enables translational studies toward dual transcriptomic diagnostics and mechanism-guided therapies in sepsis, including pediatrics and low-volume settings.
Key Findings
- Developed DRIB to simultaneously isolate host leukocyte and bacterial RNA from 0.5 mL whole blood compatible with clinical workflows.
- Dual rRNA depletion and RNA-seq yielded 16.6–24.8 million reads/sample with ~63% uniquely mapped reads.
- Host reads comprised 51–68%, while 0.5–6.7% mapped to bacterial genomes, enabling dual-species expression profiling.
- Pathway analyses highlighted immune metabolism and metal-ion binding signatures in host and bacterial responses.
Methodological Strengths
- Low-volume (0.5 mL) protocol compatible with routine clinical sampling and high-throughput sequencing.
- Dual-species rRNA depletion with robust mapping performance enabling simultaneous host–pathogen transcriptomics.
Limitations
- Pilot validation with limited sample numbers; larger, time-resolved cohorts are needed.
- Bacterial reads are a small fraction (0.5–6.7%), increasing susceptibility to contamination and stochasticity.
Future Directions: Scale DRIB to multicenter cohorts, include pediatric neonates, integrate with single-cell and cell-free RNA, and evaluate diagnostic/prognostic performance in sepsis endotypes.
Dual RNA-sequencing (dual RNA-seq) holds significant promise for deciphering bacterial virulence mechanisms during systemic infections. However, its application in sepsis research is hindered by technical challenges, including a low bacterial burden in blood and limited sample volumes and RNA yield from vulnerable populations, such as neonates. We developed an optimized protocol [dual RNA isolation from blood (DRIB)] for simultaneous stabilization, isolation and purification of high-quality host leukocyte and bacterial RNA from low-volume whole blood samples (0.5 ml). This protocol is compatible with clinical sample collection workflows and high-throughput RNA sequencing. The feasibility of DRIB for dual RNA-seq was validated using a pilot cohort of clinical adult sepsis samples, enabling the investigation of host-bacterial gene expression during sepsis. The DRIB protocol yielded 2.10-6.91 µg of total RNA per clinical sample in our pilot cohort. Dual-species ribosomal RNA (rRNA) depletion and RNA-seq generated 16.6-24.8 million filtered reads per sample, with 63±7% of reads uniquely mapped to host or bacterial sequences. Host genes accounted for 51-68% (8.4-10.9 million) reads, while 0.5-6.7% (79,496-789,808 reads) mapped to bacterial genomes. Bioinformatic analysis revealed that both shared and individual transcriptional patterns were identified in host and bacterial responses, including pathways related to immune metabolism and metal-ion binding. Our optimized DRIB protocol and RNA-seq pipeline effectively captured both host and bacterial RNA transcription in clinical sepsis samples. Expanding this approach to larger cohorts and varying disease timepoints will provide crucial new insights into host-bacterial gene co-expression dynamics in sepsis progression and outcomes.
2. Effectiveness of Antivirals Nirmatrelvir-Ritonavir and Molnupiravir in Viral Sepsis: Retrospective Cohort Study.
In 15,599 hospitalized COVID-19 patients without secondary infections, molnupiravir and nirmatrelvir-ritonavir were associated with reduced in-hospital mortality, particularly in those without organ dysfunction, and shortened length of stay in several organ dysfunction subgroups. Benefits persisted in propensity score–matched analyses across respiratory failure, acute kidney injury, and coagulopathy strata.
Impact: This large real-world analysis reframes antivirals as disease-modifying therapy in viral sepsis, demonstrating mortality and resource-use benefits across clinically relevant organ dysfunction strata.
Clinical Implications: Findings support early consideration of nirmatrelvir-ritonavir or molnupiravir in hospitalized viral sepsis (COVID-19 without secondary infection), with prioritization in patients before multiorgan dysfunction and attention to organ-specific benefits.
Key Findings
- Molnupiravir reduced mortality both with any organ dysfunction (HR 0.75, 95% CI 0.58–0.96) and without organ dysfunction (HR 0.29, 95% CI 0.15–0.56).
- Nirmatrelvir-ritonavir reduced mortality in respiratory failure (absolute risk difference 9.5%) and in patients without organ dysfunction (HR 0.17, 95% CI 0.05–0.56).
- Both antivirals shortened length of stay across subgroups (e.g., −3.37 days in respiratory failure for nirmatrelvir-ritonavir; −6.7 days in AKI for molnupiravir).
Methodological Strengths
- Large sample size with propensity score matching and organ dysfunction–stratified analyses.
- Reported both hazard ratios and absolute/mean differences, enhancing interpretability for outcomes.
Limitations
- Retrospective design with potential residual confounding and indication bias.
- Generalizability beyond COVID-19 and culture-negative cohorts is uncertain; timing of antiviral initiation not randomized.
Future Directions: Prospective randomized trials in viral sepsis (including non-COVID pathogens) to define optimal timing, patient selection, and organ-specific benefits; evaluation of combination strategies.
BACKGROUND: Viral infections, including those leading to sepsis, are common but often overlooked in clinical practice, yet the treatment strategies for viral sepsis remain inadequately defined. OBJECTIVE: This study aims to investigate the effectiveness of antivirals nirmatrelvir-ritonavir and molnupiravir in the treatment of culture-negative sepsis. METHODS: This retrospective cohort study was conducted across public hospitals in Hong Kong. We included patients diagnosed with COVID-19 between February 22, 2022, and June 30, 2023, who had no secondary bacterial or fungal infections. Propensity score matching was used to assess the efficacy of the antivirals nirmatrelvir-ritonavir and molnupiravir in patient subgroups with or without organ dysfunction at hospital admission, including circulatory shock, respiratory failure, acute kidney injury, coagulopathy, acute liver impairment, a composite of all organ dysfunctions, or no organ dysfunction. Key outcomes were in-hospital mortality and length of stay, reported as hazard ratios (HR) and mean differences, respectively. RESULTS: The study included 15,599 COVID-19 patients with a mean age of 75.1 (SD 15.9) years. Molnupiravir treatment was associated with a significantly lower risk of mortality in patients in both the presence of any organ dysfunction (HR 0.75, 95% CI 0.58 to 0.96) and without organ dysfunction (HR 0.29, 95% CI 0.15-0.56). Nirmatrelvir-ritonavir was associated with decreased mortality with respiratory failure (absolute risk difference: 9.5%, 95% CI 6.26-12.72) and without organ dysfunction (HR 0.17, 95% CI 0.05-0.56). Antivirals also reduced the length of hospital stay; nirmatrelvir-ritonavir reduced length of stay in respiratory failure by an average of 3.37 (95% CI 2.32-4.42) days, acute kidney injury by 7.25 (95% CI 2.97-11.52) days, and coagulopathy by 7.04 (95% CI 2.99-4.05) days. Molnupiravir reduced the length of stay in acute kidney injury by an average of 6.7 (95% CI 2.39-11.08) days and coagulopathy by 5.68 (95% CI 1.20-10.16) days. CONCLUSIONS: Antivirals reduced mortality among hospitalized COVID patients, with the greatest reduction observed in patients without organ dysfunction. Antivirals were also effective in reducing the length of hospital stay.
3. Longitudinal multi-omic signatures of ARDS and sepsis inflammatory phenotypes identify key pathways associated with mortality.
Integrating metabolomics and transcriptomics in ARDS patients, the study identifies phenotype-specific and phenotype-independent molecular signatures tied to mortality, including innate immune–glycolysis coupling, hepatic/immune dysfunction with impaired beta-oxidation, and interferon suppression with altered mitochondrial respiration. These signatures were validated in an independent sepsis cohort, suggesting targets for precision therapeutics.
Impact: Provides mechanistic clarity linking ARDS/sepsis inflammatory phenotypes to mortality via distinct metabolic and immune programs, validated across cohorts, enabling phenotype-informed trial design.
Clinical Implications: Supports phenotype-based risk stratification and the development of targeted interventions (e.g., metabolic modulation, interferon pathway support) in ARDS/sepsis; encourages adaptive trial designs aligned to inflammatory endotypes.
Key Findings
- Three Hyperinflammatory-associated mortality signatures: innate immune activation with increased glycolysis; hepatic/immune dysfunction with impaired fatty acid beta-oxidation; interferon suppression with altered mitochondrial respiration.
- A fourth mortality signature (redox impairment and cell proliferation) was independent of inflammatory phenotype.
- All mortality-associated signatures were validated in an independent sepsis cohort (EARLI), supporting generalizability.
Methodological Strengths
- Integrative multi-omics (transcriptomics + metabolomics) with phenotype-stratified analyses.
- Independent external validation in a separate sepsis cohort enhances robustness.
Limitations
- Preprint status without peer review; methodological details may evolve.
- Observational associations cannot prove causality; sampling timing may influence signatures and is not interventional.
Future Directions: Test targeted metabolic or interferon-modulating therapies within phenotype-enriched adaptive trials; extend validation across diverse ICUs and integrate with proteomics/single-cell data.
Critically ill patients with acute respiratory distress syndrome (ARDS) and sepsis exhibit distinct inflammatory phenotypes with divergent clinical outcomes and apparent heterogeneity of treatment effects, but the underlying molecular mechanisms remain poorly understood. These phenotypes, derived from clinical data and protein biomarkers, were associated with metabolic differences in a prior pilot study. This study investigated the metabolomic and transcriptomic differences between Hyperinflammatory and Hypoinflammatory phenotypes through integrative multi-omics analysis of blood samples from ARDS patients in the ROSE trial. Multi-omics integration revealed three molecular signatures strongly associated with the Hyperinflammatory phenotype and with mortality: enhanced innate immune activation coupled with increased glycolysis, hepatic dysfunction and immune dysfunction paired with impaired fatty acid beta-oxidation, and interferon program suppression coupled with altered mitochondrial respiration. A fourth molecular signature, not associated with inflammatory phenotype, identified redox impairment and cell proliferation pathways associated with mortality. Integrated multi-omics analysis within each inflammatory phenotype revealed distinct pathways associated with mortality. All mortality-associated molecular signatures including those within phenotypes were validated in an independent cohort of critically ill patients with sepsis (EARLI). These findings reveal distinct molecular mechanisms underlying ARDS/sepsis phenotypes and suggest potential therapeutic targets for precise treatment strategies in critical illness.