Impact of syndromic molecular diagnostics on antimicrobial adequacy and time to therapy in critically ill patients with pneumonia: a systematic review and meta-analysis of randomized trials

Abstract

Background

Pneumonia is a leading cause of ICU admission and mortality, requiring prompt and adequate antimicrobial therapy to improve outcomes. Conventional cultures are slow and often insensitive, delaying targeted treatment. Syndromic PCR panels offer rapid identification of pathogens and resistance genes directly from respiratory samples, potentially improving early antibiotic optimization. However, the true clinical benefit of these diagnostics in critically ill patients remains uncertain.

Methods

We conducted a systematic review and meta-analysis of randomized controlled trials (RCTs) comparing PCR-based molecular diagnostics with standard culture techniques in predominantly adult ICU patients with severe community-acquired, hospital-acquired, or ventilator-associated pneumonia. Literature searches were performed in PubMed, Embase, and Cochrane CENTRAL from inception to July 16, 2025. The primary outcome was in-hospital mortality. Secondary outcomes included adequacy of initial antimicrobial therapy and time to effective antibiotic administration. Data were synthesized using random-effects models.

Results

We included five randomized controlled trials comprising 2,466 patients. Syndromic PCR testing did not significantly reduce in-hospital mortality, the primary outcome (RR 1.04; 95% CI: 0.90–1.21; p = 0.57; I² = 0%). However, PCR testing was associated with a higher rate of adequate initial antimicrobial therapy (RR 1.82; 95% CI: 1.10–3.00; p = 0.02; I² = 97%) and a reduction in time to effective antibiotic administration (mean difference − 27.98 h; 95% CI: − 46.07 to − 9.89; p = 0.002; I² = 94%).

Conclusions

Syndromic PCR diagnostics did not reduce in-hospital mortality in critically ill patients with pneumonia but were associated with improved adequacy of initial antimicrobial therapy and faster initiation of effective treatment. These findings support their role as a complementary tool in ICU-based antimicrobial stewardship.

Trial registration

PROSPERO CRD420251006301.

Supplementary Information

The online version contains supplementary material available at 10.1186/s13054-025-05623-0.

Background

Pneumonia remains a major cause of morbidity and mortality among critically ill patients and is a common reason for ICU admission and antimicrobial initiation worldwide [1, 2]. Accurate and timely identification of the etiologic pathogen is essential to guide early and appropriate antimicrobial therapy, minimize unnecessary broad-spectrum antibiotic exposure, and improve clinical outcomes [3]. However, conventional microbiological cultures often have low sensitivity, are affected by prior antibiotic administration, and can require 48–72 h to yield actionable results [4, 5].

In recent years, molecular diagnostic techniques, particularly multiplex polymerase chain reaction (PCR) assays, have been introduced into clinical practice to address these limitations. These platforms allow for the rapid and simultaneous detection of a wide range of bacterial and viral pathogens directly from respiratory specimens, with results typically available within hours [6, 7]. Several syndromic panels also incorporate resistance gene detection, potentially enabling earlier antimicrobial tailoring and stewardship interventions [5, 8].

Although the theoretical advantages of syndromic PCR are considerable, the clinical impact of these diagnostic tests in ICU patients with pneumonia remains uncertain. While some studies report improved pathogen detection, more frequent and earlier antibiotic de-escalation, and reduced antibiotic duration [9–11], others fail to show consistent benefits in terms of mortality or stewardship endpoints [12, 13]. Notably, the time to effective antibiotic administration is an actionable metric in critical care, as delays in initiating appropriate therapy are strongly associated with worse outcomes, including prolonged organ dysfunction and increased mortality. Syndromic PCR may help shorten this interval by providing rapid, targeted results, but the extent to which this translates into clinical benefit remains uncertain. Moreover, concerns persist regarding the potential for overdiagnosis, misinterpretation of colonization, and cost-effectiveness [4, 14].

Given the ongoing uncertainties surrounding the clinical utility of syndromic molecular diagnostics in critically ill patients with pneumonia, we conducted a systematic review and meta-analysis of randomized controlled trials comparing these platforms with standard microbiological culture. The primary outcome was in-hospital mortality, selected as the most objective and patient-centered endpoint. Secondary outcomes included the adequacy of initial antimicrobial therapy and time to effective antibiotic administration, both representing key process-of-care measures linked to prognosis in severe infections. This outcome framework was designed to capture not only the potential downstream impact on survival but also the diagnostic and therapeutic relevance of syndromic PCR in the acute management of pneumonia in the intensive care setting.

Methods

This systematic review and meta-analysis was prospectively registered in the International Prospective Register of Systematic Reviews (PROSPERO; registration number CRD420251006301) and conducted in accordance with the Cochrane Handbook for Systematic Reviews of Interventions [15] and the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 guidelines [16].

Eligibility criteria

We included studies that met the following criteria: [1] peer-reviewed randomized controlled trials (RCTs); [2] ) enrolling predominantly (80% of trial’s population) critically ill adult patients (≥ 18 years old) with severe community-acquired pneumonia (CAP), hospital-acquired pneumonia (HAP), ventilator-associated pneumonia (VAP), or other nosocomial pneumonia, as defined by the original studies; [3] comparing molecular diagnostic methods using polymerase chain reaction (PCR) to conventional microbiological cultures for etiological identification; and [4] reporting at least one of the following outcomes: in-hospital mortality, adequacy of initial antibiotic therapy, or time to effective antibiotic administration. We excluded trials focusing exclusively on single-pathogen pneumonia (e.g., Pneumocystis jirovecii, Staphylococcus aureus).

Search strategy

A comprehensive literature search was performed in PubMed, Embase, and the Cochrane Central Register of Controlled Trials (CENTRAL) from inception to July 16, 2025. The search strategy combined controlled vocabulary (e.g., MeSH, Emtree) and free-text terms related to pneumonia, intensive care, and PCR-based diagnostics. The full strategy is available in Supplementary Table S1. Only English-language articles were included. We conducted manual searches of reference lists of included studies and other relevant reviews.

Study selection

Two investigators independently extracted data from each included trial using a standardized data collection form. We extracted all available data on study design, year, setting, sample size, patient characteristics (age, sex, pneumonia type, immunosuppression), diagnostic platform used and timing of use, in-hospital mortality, adequacy of antibiotic therapy, and time to effective treatment. Disagreements were resolved by consensus. The study selection process is illustrated in the PRISMA 2020 flow diagram (Fig. 1).

Fig. 1.

Prisma flow diagram of study screening and selection

Data extraction

Data were extracted independently by two reviewers using a standardized form and included:

• Study design, year, setting, sample size, patient characteristics (age, sex, pneumonia type, immunosuppression).

• Diagnostic platform used and timing .

• Reported outcomes: mortality, adequacy of antibiotic therapy and time to effective treatment.

Any disagreements were resolved by consensus.

Risk of bias assessment

The risk of bias for each included study was assessed using the Cochrane Risk of Bias 2.0 (RoB 2) tool [17], evaluating five domains: randomization, deviations from intended interventions, missing outcome data, measurement of outcomes, and selection of reported results. Two authors performed the assessments independently, with consensus resolution of disagreements. Studies were classified as having low risk, some concerns, or high risk of bias.

Certainty of evidence

To evaluate the overall certainty of the evidence for each outcome, we applied the GRADE (Grading of Recommendations Assessment, Development and Evaluation) framework. Factors considered included study limitations (risk of bias), inconsistency, indirectness, imprecision, and publication bias. Each outcome was rated as having high, moderate, low, or very low certainty. GRADE assessments were based on the consensus of two authors and summarized in Supplementary Table S4.

Outcomes and sensitivity analyses

The primary outcome of this meta-analysis was in-hospital mortality. Secondary outcomes included adequacy of initial antimicrobial therapy and time to effective antibiotic administration.

In-hospital mortality was defined as death occurring during the index hospitalization, regardless of the duration of hospital stay. For each trial, the longest available follow-up during hospitalization (ranging from 28 to 90 days) was considered for outcome assessment.

Adequacy of initial antimicrobial therapy was defined as the use of antibiotics that were both appropriate, meaning active in vitro or presumed active based on the combination of identified pathogens and resistance gene profiles, and proportionate, that is, not unnecessarily broad in spectrum relative to the likely or confirmed pathogens. In studies employing molecular diagnostics, adequacy was determined based on organism and resistance gene detection, with the recognition that this approach does not equate to conventional phenotypic susceptibility testing. Differences in the range and sensitivity of resistance markers across platforms were acknowledged as a source of potential variability.

Time to effective antibiotic administration was defined as the interval from diagnostic sampling (typically bronchoalveolar lavage or tracheal aspirate collection) to administration of an antibiotic regimen with in vitro activity against the identified pathogen(s), as determined either by conventional susceptibility testing or by inference from PCR-detected resistance genes. Sensitivity analyses were conducted to explore the impact of individual studies on outcomes with substantial heterogeneity, rather than being based solely on risk of bias.

Data synthesis and statistical analysis

Meta-analyses were conducted using Review Manager (RevMan) version 5.4. Risk ratios (RR) and odds ratios (OR) with 95% confidence intervals (CIs) were calculated for dichotomous outcomes, and mean differences (MD) for continuous outcomes. For continuous outcomes reported as medians and interquartile ranges, we estimated means and standard deviations using the validated method proposed by Wan et al. and Luo et al. [18, 19] A random-effects model (DerSimonian and Laird) was applied to account for anticipated clinical and methodological heterogeneity.

Statistical heterogeneity was assessed via the I² statistic and Cochran’s Q test. I² >50% or p < 0.10 indicated substantial heterogeneity. Funnel plots were used to assess publication bias for the secondary outcomes. A two-sided p-value < 0.05 was considered statistically significant.

Results

Study selection and characteristics

The PRISMA flow diagram of the study selection process is shown in Fig. 1. The initial database search yielded 1,030 records. After removing duplicates and applying eligibility criteria, nine studies remained for full-text review. Four of these were excluded for not meeting inclusion criteria, resulting in the inclusion of five randomized controlled trials (RCTs), comprising a total of 2,387 patients [20–24]. Detailed characteristics of the included studies are presented in Table 1 and Supplementary Table S3.

Table 1.

Baseline characteristics of included studies

Study	Design	Country	Patients MP/SC	Male, % MP/SC	Age†, y MP/SC	CAP , % MP/SC	Immunosuppression, % MP/SC	Sample Type	PCR test	Primary Outcome	Follow-up†, MP/SC days
INHALE WP3 2025 18	RCT	England	223/219	66/70	58/59	70/68	5/5	Endotracheal aspirate, sputum, BAL	FilmArray®	- Superiority in antibiotic stewardship at 24 h - Clinical cure at 14 days.	28
FLAGSHIP II 2022 19	RCT	Switzerland	100/108	59/63	60/60	74/77	57/56	BAL	MAGPIX, RespiFinder-22, Seegene	- Time on inappropriate antibiotics (hours)	30
Virk 2024 22	RCT	USA	563/589	62/65	62/62	19/22	40/43	Sputum, tracheal secretions, BAL	FilmArray®	- Median time to first antibiotic modification at 96 h	30
Poole 2022 21	RCT	United Kingdom	100/100	68/88	58/56	42/43	7/8	Endotracheal aspirate, sputum, BAL	FilmArray®	- Proportion of patients receiving therapy guided by PCR results	60
MULTICAP 2025 20	RCT	France	188/197	69/71	65/67	100/100	0/0	Endotracheal aspirate, sputum, BAL	FilmArray®	- Number of antibiotic-free days	90

^†mean or median; BAL: bronchoalveolar lavage CAP: Community-acquired pneumonia; MP: Molecular Panel; NA: not available; PCR: Polymerase Chain Reaction; RCT: randomized controlled trial; SC: Standard Culture; USA: United States of American

Pooled analysis of all studies

In the pooled analysis of five randomized trials, in-hospital mortality did not differ significantly between patients managed with syndromic PCR and those receiving conventional microbiological testing (risk ratio 1.04; 95% CI: 0.90–1.21; p = 0.57; I² = 0%) (Fig. 2). By contrast, adequacy of initial antimicrobial therapy was significantly higher in the syndromic PCR group (risk ratio 1.82; 95% CI: 1.10–3.00; p = 0.02; I² = 97%) (Fig. 3). Additionally, time to effective antibiotic administration was significantly reduced with syndromic testing, with a pooled mean difference of − 27.98 h (95% CI: − 46.07 to − 9.89; p = 0.002; I² = 94%) (Fig. 4).

Fig. 2.

Forest plot of In-hospital mortality. There was no statistically significant difference in in-hospital mortality between patients evaluated with syndromic PCR panels and those assessed by conventional microbiology (risk ratio 1.04; 95% confidence interval: 0.90–1.21; p = 0.57; I² = 0%)

Fig. 3.

Forest plot of adequacy of initial antimicrobial therapy. Use of syndromic molecular panels (PCR panel) was associated with significantly higher rates of adequate initial antimicrobial therapy compared to conventional microbiology (risk ratio 1.78; 95% confidence interval: 1.23–2.57; p = 0.002; I² = 95%)

Fig. 4.

Forest Plot of Time to Effective Antimicrobial Therapy. Syndromic molecular panel (PCR Panel) testing significantly reduced the time to effective antibiotic administration compared to conventional microbiology (mean difference − 28.03 h; 95% confidence interval: −47.49 to − 8.58; p = 0.005; I² = 95%)

Sensitivity analysis

Given the high degree of heterogeneity observed in the outcomes of both time to targeted therapy (I² = 94%) and adequacy of initial antimicrobial therapy (I² = 95%), we conducted sensitivity analyses to assess the robustness of all primary outcomes.

For the adequacy of initial antimicrobial therapy, sequential leave-one-out analyses were conducted, excluding each trial individually. The direction and magnitude of the pooled effect size remained consistent with the primary analysis across all models. Notably, statistical significance was preserved in all sensitivity analyses except when the INHALE WP3 trial was excluded (risk ratio 2.04; 95% CI: 0.94–4.44; p = 0.07; I² = 96%), suggesting overall robustness of the findings (Supplementary Figures S3–S6).

For the outcome of time to effective antibiotic therapy, sensitivity analyses excluding each trial in turn demonstrated greater variability in statistical significance. While the exclusion of the Poole (p = 0.09) or FLAGSHIP II (p = 0.07) trials yielded nonsignificant results, the exclusion of the Virk trial restored statistical significance (mean difference: − 28.03 h; 95% CI: − 47.49 to − 8.58; p = 0.005; I² = 95%), indicating that the overall effect was not driven by a single outlier but remained sensitive to the inclusion of individual studies (Supplementary Figures S7–S9).

Meta-regression analysis

To explore potential sources of heterogeneity in antimicrobial adequacy, we conducted an exploratory meta-regression using the proportion of patients with community-acquired pneumonia (CAP) as a moderator (Supplementary Table S7). The analysis included all four studies and revealed that this variable explained only a small proportion of the between-study heterogeneity (R² = 2.5%), with substantial residual heterogeneity remaining (τ² = 0.2374; I² = 94.8%). The association between CAP proportion and antimicrobial adequacy did not reach statistical significance (coefficient = 0.86; 95% CI: -0.70 to 2.41; p = 0.28), and the test of moderators was also non-significant (QM = 1.17; p = 0.280) (Supplementary Table S6 and Figure S10) .These findings suggest that pneumonia subtype alone does not account for the variability in adequacy outcomes across studies, and other unmeasured clinical or methodological factors may be contributing to the observed heterogeneity.

Quality assessment

Three trials were judged to have an overall low risk of bias across all domains, while two studies (FLAGSHIP II and MULTI-CAP) [21, 22] was rated as having “some concerns” due to deviations from intended interventions and selective reporting (Supplementary Table S2). No study was classified as having a high overall risk of bias.

Potential publication bias was assessed using funnel plots for the two secondary outcomes: adequacy of initial antimicrobial therapy and time to targeted therapy. For the adequacy outcome, visual inspection suggested a degree of asymmetry, with a relative absence of small studies showing neutral or negative effects. No substantial asymmetry was observed in the funnel plot for time to therapy. Nevertheless, interpretation remains limited due to the small number of included studies (n = 4), and Egger’s regression test was not performed due to insufficient power. The corresponding funnel plots are shown in Supplementary Figures S1 and S2.

Discussion

In this systematic review and meta-analysis of randomized controlled trials, we found that the use of syndromic PCR-based molecular diagnostics in critically ill patients with pneumonia did not significantly reduce in-hospital mortality. Nonetheless, the intervention was associated with improved adequacy of initial antimicrobial therapy and a shorter time to effective, targeted treatment. These process-of-care benefits underscore the potential of rapid molecular diagnostics to enhance early therapeutic decision-making in the ICU, even if such improvements have not yet translated into consistent reductions in mortality.

The improved adequacy of empirical therapy likely stems from the ability of syndromic PCR panels to rapidly identify pathogens and resistance markers directly from respiratory samples. In the ICU setting, where delays or inappropriate therapy are strongly linked to adverse outcomes including septic shock and death, these diagnostic advantages are particularly valuable [1–3]. Our findings are consistent with previous observational studies and meta-analyses demonstrating that molecular testing can support antimicrobial stewardship by enabling earlier antibiotic optimization [25–28].

Importantly, this is the first meta-analysis to systematically evaluate clinical outcomes, including in-hospital mortality, associated with syndromic PCR in critically ill patients with pneumonia. By synthesizing randomized trial data, our study provides novel insights into the real-world clinical utility of these platforms in ICU practice.

The lack of a mortality benefit should be interpreted in context. Most patients in both intervention and control arms received early broad-spectrum antibiotics, potentially attenuating the incremental value of faster pathogen identification. Additionally, although all included trials assessed antimicrobial adequacy, definitions and methodologies varied considerably (Supplementary Table S5). Some studies used culture-based susceptibility data, while others relied on the detection of resistance genes by PCR, and the timing and context of sample collection also differed. These inconsistencies may have introduced indirectness and contributed to the observed heterogeneity. Standardized and clinically grounded definitions of adequacy are needed to facilitate meaningful comparisons in future studies.

Mortality is a multifactorial outcome in critical illness, influenced not only by appropriateness of antibiotics but also by comorbidities, organ failure severity, source control delays, and complications unrelated to infection. Although molecular diagnostics can improve process-of-care measures such as timeliness and appropriateness of therapy, these improvements may be insufficient to reverse the trajectory of critically ill patients with advanced organ dysfunction. Moreover, most trials were underpowered to detect modest differences in mortality due to low event rates and heterogeneous populations. This underscores the limitations of using mortality as a primary endpoint to assess diagnostic interventions in the ICU.

Our findings also underscore the challenges in applying microbiological adequacy definitions across varied clinical contexts. In HAP and VAP, for example, empiric broad-spectrum coverage is often justified. As a result, treatments deemed “inadequate” by microbiological criteria may have been clinically appropriate. Future studies should adopt pragmatic definitions of adequacy that reflect both pathogen sensitivity and clinical context.

Recent expert consensus has emphasized the need for structured diagnostic stewardship programs to ensure that PCR results are interpreted and acted upon effectively. Indeed, some included trials showed that PCR testing influenced prescribing behavior and facilitated earlier de-escalation, reinforcing its value as a stewardship tool. The substantial heterogeneity in time to effective therapy (I² = 95%) likely reflects variability in trial design, stewardship implementation, and institutional response to PCR results.

We also emphasize technical limitations of widely used panels, such as the BioFire Pneumonia FilmArray, which does not detect Stenotrophomonas maltophilia, a relevant pathogen in critically ill patients. Moreover, its resistance gene coverage is limited for non-fermenting Gram-negative bacilli, whose resistance mechanisms are complex and not reliably monogenic. These shortcomings may impair the ability of PCR platforms to fully inform empirical therapy decisions in some ICU contexts. Additionally, the clinical significance of the semi-quantitative reporting bins remains uncertain, and these values are often misinterpreted as surrogates for pathogen burden.

To further explore potential sources of heterogeneity, we conducted an exploratory meta-regression using pneumonia subtype (community-acquired vs. nosocomial) as a moderator of the effect size for antimicrobial adequacy (Table S. Although the interaction was not statistically significant (p = 0.280), the model substantially reduced residual heterogeneity (I² = 0%), suggesting that pneumonia subtype may partly explain inter-study diferences (Supplementary Table S6 and Figure S10).Given the small number of included trials, however, this finding should be considered hypothesis-generating rather than conclusive.

Generalizability is another important consideration. All included studies were conducted in high-resource settings with access to advanced diagnostic infrastructure and infectious disease support. The performance and impact of syndromic PCR in lower-resource ICUs, or in the absence of robust antimicrobial stewardship programs, remains uncertain.

This review has limitations. The number of included trials was small, and despite a large pooled sample size, the analysis may have been underpowered to detect differences in mortality. The limited number of studies also restricts the reliability of funnel plot assessments. Additionally, definitions of antimicrobial adequacy were heterogeneous, and other clinically meaningful outcomes such as treatment duration, resistance development, and cost-effectiveness were inconsistently reported. While we performed sensitivity analyses and assessed study quality rigorously, unmeasured confounding and clinical heterogeneity remain possible.

Finally, one included study enrolled both adults and children. Although over 80% of participants were adults, the inclusion of pediatric patients may have introduced minor clinical heterogeneity. Nonetheless, we considered the aggregate data appropriate for inclusion given the predominance of adult subjects.

Conclusions

In conclusion, our findings suggest that syndromic PCR-based diagnostics represent a promising technological advance for the rapid identification of pathogens in critically ill patients with pneumonia. While these platforms appear to enhance microbiological yield and may support antimicrobial stewardship through earlier targeted therapy, the clinical impact remains uncertain. Future large-scale, multicenter randomized trials are needed to determine whether these diagnostic improvements translate into meaningful patient-centered outcomes.

Supplementary Information

Below is the link to the electronic supplementary material.

Abbreviations

ICU

Intensive care unit

PCR

Polymerase chain reaction

RCT

Randomized controlled trial

CAP

Community-acquired pneumonia

HAP

Hospital-acquired pneumonia

VAP

Ventilator-associated pneumonia

PROSPERO

International prospective register of systematic reviews

OR

Odds ratio

RR

Risk ratio

CI

Confidence interval

MD

Mean difference

RoB 2

Risk of bias 2.0 Tool

GRADE

Grading of recommendations, assessment, development and evaluation

CENTRAL

Cochrane central register of controlled trials

SC

Standard culture

MP

Molecular panel

Author contributions

YAPS and FJSR conceived the study idea and defined the review objectives and methodology. YAPS, FJSR, BMT, and MHCS conducted the systematic literature search and independently extracted data from eligible studies. YAPS and ELQC performed the statistical analysis and meta-analytical synthesis. ELVC, LPJ, MHCS, and FJSR contributed to the interpretation of the results and the assessment of study quality. YAPS drafted the initial manuscript. All authors critically revised the manuscript for important intellectual content and approved the submitted version.

Funding

This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors. No institutional or departmental funds were used.

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

Not applicable. This study is a systematic review and meta-analysis of previously published randomized controlled trials and did not involve access to individual patient data.

Consent for publication

Not applicable. This manuscript does not contain any individual person’s data in any form.

Competing interests

FJSR reports speaker fees from bioMérieux. The other authors declare that they have no competing interests.