Severe pneumonia continues to represent a substantial global burden of mortality.​ (Ward and Goldie, 2024). ​Current epidemiological data estimate that pneumonia necessitates ICU admission for approximately 100,000 patients worldwide each year (Mandell et al., 2007), where critical illness is associated with mortality rates reaching 20–50%​ (Lee et al., 2020). Emerging evidence reveals that the lungs, akin to the nasopharynx and gut, harbor a unique microbial ecosystem composed predominantly of bacteria, fungi, and viruses (Yu et al., 2016; Man et al., 2017). At a steady state, the lung microbiota critically maintains immune-metabolic balance, promoting pathogen clearance and suppressing maladaptive inflammation.​ (Dethlefsen et al., 2007; Mendez et al., 2019; Tomé, 2021). Notably, the diminished diversity of the respiratory microbial community has been proposed as an ecological hallmark of infection (Abreu et al., 2012). Clinically relevant perturbations characterize the lung microbiome in pulmonary disorders (Kim et al., 2017; He et al., 2022; Montassier et al., 2023; Li et al., 2024), and dysbiosis—imbalances in microbial structure and abundance—may influence disease onset, progression, and prognosis (Whiteside et al., 2021). Consequently, the respiratory microbiota ​represents a novel diagnostic biomarker enabling real-time disease surveillance​.

Streptococcus is the dominant colonizing flora in the lower respiratory tract (Woods et al., 2021). Streptococcus is taxonomically dominant within the conserved pulmonary microbiome, serving as a critical indicator of respiratory ecosystem health (Dickson et al., 2017). Among the ​primary commensals​ colonizing the human nasopharynx within hours of parturition (Bloch et al., 2024), Streptococcus participates in shaping complex microbial communities under both healthy and diseased conditions. ​Clinically, it sustains mucosal immune homeostasis and governs ecologically balanced colonization in the oropharyngeal microbiome (Abranches et al., 2018), serving as an initial dominant population in the human oral microbiome. This genus contributes to amino acid and nucleotide biosynthesis and is a key participant in bacterial quorum sensing (Su et al., 2023). Under specific conditions, Streptococcus acts as a commensal, promoting normal immune system development and inhibiting pathogen colonization (Lang et al., 2010); its metabolites (e.g., short-chain fatty acids) facilitate epithelial barrier repair and suppress excessive inflammation (Su et al., 2023). However, previous research has predominantly focused on the pathogenic potential of Streptococcus (Yumoto et al., 2019). Multiple studies (Erb-Downward et al., 2011; Bousbia et al., 2012) have reported no significant differences in Streptococcus detection rates between patient cohorts and healthy controls, reinforcing its role as a core respiratory tract colonizer. In asthma patients, lower inflammatory levels have been associated with pulmonary Streptococcus colonization during homeostasis (Bernasconi et al., 2016; Sharma et al., 2020). Similarly, reduced inflammation linked to Streptococcus has been observed in lung transplant recipients (Bernasconi et al., 2016), whereas in idiopathic pulmonary fibrosis (IPF) patients, Streptococcus colonization correlates with faster disease progression (O’Dwyer et al., 2019; Invernizzi et al., 2020); Streptococcus biomass accumulation remodels the lung tumor immune microenvironment via PI3K/ERK-dependent epithelial reprogramming, potentiating myeloid cell infiltration (Gustafson et al., 2010). These findings underscore the significant yet context-dependent role of Streptococcus colonization across respiratory pathologies. A microbiome investigation of severe pneumonia stratified patients into deceased and survivor cohorts according to 28-day survival outcomes. The survivor group exhibited Streptococcus as a core genus within the respiratory microbiota. Nevertheless, this study did not elucidate the mechanistic association between Streptococcus colonization and severe pneumonia prognosis (Huang et al., 2024).

Therefore, this study aimed to investigate the association between Streptococcus colonization and clinical outcomes in patients with severe pneumonia using metagenomic next-generation sequencing (mNGS).​ By analyzing the microbial composition data obtained from mNGS and the clinical data, we attempt to describe the clinical characteristics and prognostic analysis of Streptococcus colonization.

​To our knowledge, this study is the first to uniquely focus on elucidating the association between Streptococcus colonization and prognosis in severe pneumonia patients. We collected respiratory tract specimens from patients and employed both metagenomic next-generation sequencing (mNGS) and culture-based microbial testing (CMT) to determine Streptococcus colonization status in severe pneumonia cases, with final clinical adjudication performed by physicians. The results suggest that Streptococcus colonization may exert potential beneficial effects on improving prognosis in critically ill patients.

Our study observed a 28-day mortality rate of 41.32% post-admission. ​Systematic reviews report 20-50% mortality among ICU-admitted severe pneumonia patients, consistent with multinational registry data (Lee et al., 2020), which aligns with our findings. However, selection bias may lead to underestimation of true mortality rates, as the cohort excluded critically ill patients who did not meet the criteria for metagenomic next-generation sequencing (mNGS) sampling.​ Additionally, no statistically significant difference in 28-day mortality was observed between the two patient groups (p > 0.05). Based solely on this pivotal clinical endpoint, Streptococcus colonization status did not exert a significant impact on short-term survival outcomes. Nevertheless, upon further analysis of secondary clinical outcomes, we identified significantly shorter total hospital stays and ICU lengths of stay (LOS) in the Streptococcus-colonized group compared to the non-colonized group (p < 0.05). These findings preliminarily suggest that Streptococcus colonization may exert a beneficial effect on reducing total hospital stay and ICU length of stay (LOS). Notably, the results remained highly consistent before and after propensity score matching (p < 0.05), further reinforcing the reliability and robustness of this observation. Importantly, a previous study investigating microbial characteristics in survivors versus non-survivors of severe pneumonia identified Streptococcus as a core genus in the survivor group, with positive correlations observed across its subspecies. (Huang et al., 2024) This aligns with our current findings in severe pneumonia patients, implying that Streptococcus may play a multifaceted role in disease progression and influence short-term prognosis. Subsequent analysis of bacterial detection profiles before/after matching and interventions (Figure 4), revealed comparable prevalence of Acinetobacter and Klebsiella between Streptococcus-colonized and non-colonized groups. However, Stenotrophomonas detection was significantly lower (8.1% vs. 21.6%), and Haemophilus prevalence was higher (16.2% vs. 6.0%) in the colonized group. However, Stenotrophomonas detection was significantly lower (8.1% vs. 21.6%) and Haemophilus prevalence was higher (16.2% vs. 6%) in the colonized group. This microbial shift may arise from interspecies interactions: prior studies have documented Haemophilus-Streptococcus co-colonization (Lewnard et al., 2019), while research on pediatric Mycoplasma pneumoniae pneumonia demonstrated that increased Streptococcus and Prevotella abundance displaces Stenotrophomonas, with their metabolic production of lactate promoting Veillonella growth and anti-inflammatory short-chain fatty acids (SCFAs) (Wei et al., 2024). Analogous mechanisms might operate in severe pneumonia. However, the lack of longitudinal microbial dynamics precluded further mechanistic exploration. A key finding of the present study was that the detection rates of herpes simplex virus type 1 (HSV-1, 31.76% vs 14.19%) and cytomegalovirus (CMV, 16.89% vs 9.46%) were significantly higher in the Streptococcus-negative group (SP-) than in the Streptococcus-positive group (SP+). This observation suggested that Streptococcus colonization may be associated with the risk of respiratory herpesvirus infection. The above finding is consistent with the well-established consensus that interactions exist between the respiratory microbiota and viral pathogens, and also possesses distinct clinical implications. Our previous work has verified a prominent synergistic effect between HSV-1 and CMV, with their co-detection being closely correlated with an elevated 28-day mortality rate among critically ill patients (Liu et al., 2025). Moreover, accumulating evidence has demonstrated that respiratory commensal bacteria are capable of modulating host susceptibility to viral infections (Kim et al., 2019). Nevertheless, as a core commensal bacterium of the respiratory tract, the direct interaction between Streptococcus, HSV-1, and CMV has not been clearly elucidated in existing literature, which thus merits further in-depth investigations.

Previous studies have established colony-forming units (CFUs) as a critical metric for determining microbial pathogenicity (Bousbia et al., 2012). For defining lower respiratory tract colonization, a bronchoalveolar lavage (BAL) culture threshold of ≥10³ CFU·mL⁻¹ is widely adopted as evidence of bacterial colonization (Cabello et al., 1997), though some studies employ higher thresholds to refine pathogenicity assessments. Clinically, the principle “higher bacterial loads correlate with stronger infection relevance” guides pathogen identification (Chang et al., 2018), and positive cultures from lower respiratory specimens are typically classified as pathogens (Kitsios et al., 2018). However, metagenomic next-generation sequencing (mNGS)-based diagnosis represents an emerging field lacking standardized thresholds analogous to CFU-based criteria for distinguishing colonization from infection (Metlay et al., 2019). In practice, mNGS demonstrates a limited capacity for pathogen quantification, focusing primarily on presence/absence detection. Consequently, mNGS is rationally considered a qualitative assay (Diao et al., 2023), as it cannot discriminate between infection and colonization (Diao et al., 2023). Crucially, neither culture results nor microbial DNA sequencing alone suffices to delineate colonization versus infection; this requires comprehensive integration of clinical symptoms, radiological features, and systemic/local host responses. Therefore, this study utilized clinical microbiological testing (CMT) as the gold standard, wherein clinicians integrated multi-dimensional clinical parameters to adjudicate Streptococcus colonization status. Although this approach may introduce inter-rater variability, it aligns with real-world clinical decision-making workflows, thereby enhancing the clinical translatability of our findings.

The causal relationship between Streptococcus colonization and prognosis of severe pneumonia, as well as its colonization dynamics, remains under investigated, despite accumulating evidence highlighting its pathogenic significance in respiratory disorders (Preston et al., 2011; Segal et al., 2016; Invernizzi et al., 2020; Narciso et al., 2025). Notably, Streptococcus exerts heterogeneous functional roles in the lower respiratory tract across disease states—for instance, promoting pulmonary carcinogenesis in cancer patients and accelerating progression in idiopathic pulmonary fibrosis—precluding simplistic pathogenic classification (Tsay et al., 2018). Although the specific mechanisms underlying Streptococcus colonization-related effects remain unclear, prior studies have identified potential regulatory pathways: Streptococcus may ameliorate microbial dysbiosis via quorum-sensing-mediated oral biofilm stabilization (Su et al., 2023), reduce secondary infection risk through bacteriocin production (Wescombe et al., 2009; Khan et al., 2019), and modulate host immune responses (Preston et al., 2011), with preclinical evidence supporting its efficacy in alleviating recurrent pharyngitis and pathogen colonization (Deasy et al., 2015; Andaloro et al., 2019)Nevertheless, research on Streptococcus-supplemented probiotic interventions for respiratory diseases is still exploratory. Whether probiotics confer benefits in respiratory diseases by directly targeting lung parenchyma or restoring upper airway/intestinal microbiota homeostasis, along with their specific regulatory pathways, awaits further verification.

Streptococcus persists as a core genus in survivors of severe pneumonia. However, critical questions remain unresolved: Whether Streptococcus colonization synergizes with established pathogens to exacerbate pulmonary infection severity, whether it suppresses other pathogens through competitive inhibition, and whether it merely exists as a commensal without substantial clinical impact. Elucidating the ecological role of Streptococcus in this complex microenvironment holds significant implications for deciphering pulmonary infection pathogenesis, developing targeted therapeutic strategies, and improving patient prognosis.

Our study has several important limitations requiring careful consideration: Antimicrobial therapy, the cornerstone of severe pneumonia management (Zhao et al., 2023), was not systematically tracked.

Empirical broad-spectrum antibiotics are typically initiated within hours of ICU admission, profoundly altering lung microbiome composition within days – previous evidence shows sustained diversity reduction after just 72 hours of treatment (Torres et al., 2016). This missing pharmacological metadata may introduce potential confounding factors, as antibiotic-induced dysbiosis could independently affect outcomes. Single-timepoint mNGS profiling​ limited our capacity to characterize dynamic evolutionary profiles of Streptococcus and other genera at taxonomic resolution. The temporal dynamics of microbiome restructuring were not systematically analyzed. However, the observed depletion of Streptococcus (a core microbiome component) aligns with previous reports linking microbial community destabilization to adverse outcomes (Huang et al., 2024), reinforcing the pathological significance of pulmonary microbiome imbalance. Notably, although inter-group mortality showed no statistical difference, the significant disparities in total hospital stays and ICU length of stay (LOS) suggest pulmonary microbiome modulation could emerge as a therapeutic target for improving prognosis. Promising preclinical evidence exists: murine models demonstrate airway-targeted microbial modulation suppresses allergic immunity in asthma (Preston et al., 2011), while bacterial oral/nasal sprays reduce pharyngitis recurrence and pathogen colonization in clinical trials (Deasy et al., 2015; Andaloro et al., 2019). Nevertheless, key questions remain unresolved: Whether probiotic supplementation or microbial consortia transplantation can restore respiratory homeostasis; Whether direct lung parenchymal delivery outperforms upper airway/gut microbiome modulation. These knowledge gaps underscore the need for mechanistic studies exploring microbial therapeutics in critical respiratory illnesses.