Next-Generation Sequencing Diagnostics of Bacteremia in Sepsis

Study Overview
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Brief Summary
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Sepsis remains a major challenge, even in modern intensive care medicine. The identification of the causative pathogen is crucial for an early optimization of the antimicrobial treatment regime in patients with sepsis. In this context, culture-based diagnostic procedures (e.g. blood cultures) represent the standard of care, although they are associated with relevant limitations. Therefore, culture independent methods (e.g. Next-Generation Sequencing (NGS)) seem to be an attractive alternative. By the identification of circulating cell-free DNA in the blood and the use of the quantitative sepsis indicating quantifier (SIQ) score, causing pathogens can be identified and potential contaminations can be excluded. The goal of the presented study is therefore, to assess the diagnostic performance of a NGS-based approach for the detection of relevant infecting organisms in a big cohort of septic patients (n=500). Moreover, the plausibility of this NGS-based approach will be estimated by a panel of independent clinical specialists, retrospectively identifying potential changes in patients´ management based on NGS results.

Detailed Description
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Sepsis remains a major challenge, even in modern intensive care medicine. The identification of the causative pathogen is crucial for an early optimization of the antimicrobial treatment regime in patients with sepsis. In this context, culture-based diagnostic procedures (e.g., blood cultures) represent the standard of care, although they are associated with relevant limitations. Accordingly, culture-independent molecular diagnostic procedures might be of help for the identification of the causative pathogen in infected patients. Especially the concept of an unbiased sequence analysis of circulating cell-free DNA (cfDNA) from plasma samples of septic patients by next-generation sequencing (NGS) has recently been identified to be a promising diagnostic platform for critically ill patients suffering from bloodstream infections. Although this new approach might be more sensitive and specific than culture-based state-of-the-art technologies, additional clinical trials are needed to exactly define the performance as well as clinical value of a NGS-based approach. Next GeneSiS is a prospective, observational, non-interventional, multicenter study to assess the diagnostic performance of a NGS-based approach for the detection of relevant infecting organisms in patients with suspected or proven sepsis (according to recent sepsis definitions [sepsis-3]) by the use of the quantitative sepsis indicating quantifier (SIQ) score in comparison to standard (culture-based) microbiological testings. Moreover, the clinical value of this NGS-based approach will be estimated by a panel of independent clinical specialists, retrospectively identifying potential changes in patients´ management based on NGS results. Further subgroup analyses will focus on the clinical value especially for patients suffering from a failure of empiric treatment within the first three days after onset (as assessed by (1.) death of the patient or lack of improvement of the patient´s clinical condition (in terms of an inadequate decrease of SOFA-score) or (2.) persistent high procalcitonin levels). This prospective, observational, non-interventional, multicenter study trial for the first time investigates the performance as well as the clinical value of a NGS based approach for the detection of bacteremia in patients with sepsis and may therefore be a pivotal step toward the clinical use of NGS in this indication. Two sets of blood cultures (2x aerobic / 2x anaerobic) will be collected at study inclusion (=Onset) as well as 72 hours afterwards (=72h). In parallel, plasma samples for NGS-based measurements need to be obtained as described previously. Further blood samples for NGS-based measurements can be collected whenever physicians order blood cultures (2x aerobic / 2x anaerobic) because of the clinical suspicion of a bloodstream infection (BSI) within the first 3 days after study inclusion. Results of microbiological routine diagnostics in specimens different from blood (e.g. body fluid, tissue, bronchoalveolar lavage, endotracheal aspirate) will be used for further analyses when they are obtained within a timeframe of ≤72 hours prior or after the timepoints for NGS-based measurements. Clinical data collection and (if possible) PCT measurements will be performed at Onset as well as at 72h after study inclusion. The final outcome evaluation of patients will be performed at 28 days.

Official Title
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Next-Generation Sequencing Diagnostics of Bacteremia in Sepsis (Next GeneSiS-Trial) - Study Protocol for a Prospective, Observational, Non-interventional, Multicenter, Clinical Trial

Conditions
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Sepsis, Septic Shock

Intervention / Treatment
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* Diagnostic Test: Next-Generation Sequencing


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Age ≥18yr Informed consent Sepsis (with an onset ≤24h): Patients with a life-threatening organ dysfunction caused by a dysregulated host response to a suspected or proven infection. Organ dysfunction can be identified as an acute change in total SOFA score ≥2 points consequent to the infection. Patients can also be promptly identified at the bedside with qSOFA, ie, alteration in mental status, systolic blood pressure ≤100mmHg, or respiratory rate ≥22/min. or Septic shock (with an onset ≤24h): Patients with septic shock can be identified with a clinical construct of sepsis with persisting hypotension requiring vasopressors to maintain mean arterial pressure (MAP) ≥65mmHg and having a serum lactate level >2mmol/L (18mg/dL) despite adequate volume resuscitation. Exclusion Criteria: Age ≤18yr Refusal to give consent Patient will probably be discharged from the ICU within the first 72h following inclusion Palliative treatment intent Clinician is not committed to aggressive treatment Death is deemed imminent and inevitable Patients who had previously been included, but are readmitted to the ICU during the same hospitalization, will not be included a second time.

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 99 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Sepsis Group<br>Patients (n=500) with suspected or proven sepsis or septic shock (according to the Sepsis-3 definitions). | Diagnostic Test: Next-Generation Sequencing<br>* In 500 patients with suspected or proven sepsis or septic shock (according to the Sepsis-3 definitions), patients´ characteristics and routine blood parameters will be determined at sepsis onset as well as 72 hours afterwards. At the same time points, 2 sets of blood cultures and one blood tube for Next-Generation Sequencing (NGS)-diagnostics will be collected. An evaluation of outcome will be performed at 28 days after sepsis onset.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Sensitivity [%] (of the NGS-based SIQ-score in comparison to blood culture diagnostics, as the goldstandard for the identification of the causative pathogen in sepsis) | Proportion of positives that are correctly identified as such. | 2 years | 
| Specificity [%] (of the NGS-based SIQ-score in comparison to blood culture diagnostics, as the goldstandard for the identification of the causative pathogen in sepsis) | Proportion of negatives that are correctly identified as such. | 2 years | 
| Positive predictive value [%] (of the NGS-based SIQ-score in comparison to blood culture diagnostics, as the goldstandard for the identification of the causative pathogen in sepsis) | Proportion of positives that are true positive. | 2 years | 
| Negative predictive value [%] (of the NGS-based SIQ-score in comparison to blood culture diagnostics, as the goldstandard for the identification of the causative pathogen in sepsis) | Proportion of negatives that are true negative. | 2 years | 
| Cohen's kappa coefficient [no measuring unit; 0</=k</=1] (of the NGS-based SIQ-score in comparison to blood culture diagnostics, as the goldstandard for the identification of the causative pathogen in sepsis) | Measurement of the interobserver agreement between both items. | 2 years | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Plausibility [%] (of the NGS-based SIQ-score) | Using a majority rule, SIQ-score results will be evaluated for plausibility by a panel of three independent clinical specialists not associated with the study site. Therefore, the panel will be provided with clinical case summaries, NGS results and standard-of-care results from all samples tested. Results of microbiological routine diagnostics in specimens different from blood (e.g. body fluid, tissue, bronchoalveolar lavage, endotracheal aspirate) will be included when they have been obtained within a timeframe of ≤72 hours prior or after the timepoints for NGS-based measurements. | 2,5 years | 
| Changes in therapy [%] (that may have occurred if the NGS-based SIQ-score had been available for clinical use) | Using a majority rule, the clinical value of the NGS-based approach will be estimated by a panel of three independent clinical specialists not associated with the study site, retrospectively identifying potential changes in patients´ management based on NGS results. Therefore, the panel will be provided with clinical case summaries, NGS results and standard-of-care results from all samples tested. Results of microbiological routine diagnostics in specimens different from blood (e.g. body fluid, tissue, bronchoalveolar lavage, endotracheal aspirate) will be included when they have been obtained within a timeframe of ≤72 hours prior or after the timepoints for NGS-based measurements. To identify potential changes in antimicrobial management that may have occurred if the results from the NGS technology had been available for clinical use, the panel will be provided with a special questionnaire. | 2,5 years | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Sepsis, Septic shock, Next-Generation Sequencing, Bacteremia