This study is a 14-day, randomized, double-blind, placebo-controlled trial involving children aged 3 months to 6 years who meet clinical diagnostic criteria for mild to moderate upper respiratory tract infection (URTI). Participants are randomized to receive either a mixed probiotic preparation (Lactobacillus rhamnosus CRL1505 and Bifidobacterium breve M-16V) or a placebo. The primary clinical outcome assessed is the duration and severity of respiratory symptoms. Secondary outcomes include changes in gut microbiota composition, intestinal immune markers, and quality of life. To investigate potential mechanisms, fecal samples are collected before and after intervention for metagenomic sequencing to analyze microbial diversity and composition, alongside immunological assessments such as sIgA and calprotectin.

Upper respiratory tract infection (URTI) is one of the most common pediatric illnesses, particularly affecting young children whose immune systems are still developing. Children under six years of age are especially susceptible to frequent infections, which can negatively impact growth, daily functioning, and overall quality of life. Although most URTIs are mild and self-limiting, recurrent or prolonged symptoms can lead to increased healthcare utilization, caregiver burden, and inappropriate antibiotic use, contributing to antimicrobial resistance. Therefore, safe and effective interventions that can reduce symptom severity and duration are of significant clinical importance.

The pathogenesis of URTI is multifactorial, involving viral exposure, host immune responses, and microbial ecosystem balance. Increasing evidence highlights the role of the gut-lung axis, a bidirectional communication pathway between the intestinal microbiota and the respiratory system. The gut microbiota plays a critical role in immune system maturation and regulation. Disruptions in microbial balance (dysbiosis) may impair immune defenses, making children more vulnerable to respiratory infections.

Probiotics, defined as live microorganisms that confer health benefits when administered in adequate amounts, have shown promise in modulating the gut microbiota and enhancing immune responses. Specific strains such as Lactobacillus rhamnosus and Bifidobacterium breve have been associated with improved mucosal immunity, including increased production of secretory immunoglobulin A (sIgA), regulation of inflammatory responses, and enhanced resistance to pathogens. These effects may translate into reduced severity and shorter duration of respiratory symptoms.

Several biological mechanisms have been proposed to explain how probiotics exert beneficial effects on respiratory health. The gastrointestinal and respiratory tracts are interconnected components of the mucosal immune system. Activation of gut-associated lymphoid tissue by probiotics can stimulate systemic immune responses, including the production of cytokines and immunoglobulins that enhance respiratory defense. Additionally, probiotics may influence metabolic pathways through the production of short-chain fatty acids, which have anti-inflammatory and immunomodulatory properties. Maintaining microbial balance in both the gut and respiratory tract is essential for optimal immune function.

Children with respiratory infections often exhibit reduced levels of beneficial gut bacteria, such as Lactobacillus and Bifidobacterium, along with altered immune markers. Probiotic supplementation may help restore this balance, improve intestinal barrier function, and enhance systemic immunity. Clinical benefits reported in previous studies include reduced symptom severity, faster recovery, and improved quality of life. Importantly, probiotics are generally well tolerated and have a strong safety profile in pediatric populations.

Based on this scientific rationale, the present study focuses on children aged 3 months to 6 years diagnosed with mild to moderate URTI. Participants will receive a fixed-dose combination of Lactobacillus rhamnosus CRL1505 and Bifidobacterium breve M-16V (1 × 10¹⁰ CFU/day) or a placebo for 14 consecutive days. Clinical outcomes, including symptom duration and severity, will be evaluated alongside safety measures.

To further elucidate underlying mechanisms, fecal samples will be collected to assess gut microbiota composition using metagenomic sequencing, examining parameters such as alpha diversity, beta diversity, and relative abundance of key taxa. In addition, intestinal immune markers, including calprotectin (CALP) and secretory IgA (sIgA), will be measured. Quality of life will also be assessed using a pediatric respiratory questionnaire. Through the integration of clinical, microbiological, and immunological data, this study aims to clarify the therapeutic potential and mechanistic basis of probiotic intervention in pediatric URTI.

Inclusion Criteria:

• Children aged 3 months to 6 years, regardless of sex.
• Meet the clinical diagnostic criteria for upper respiratory tract infection, including but not limited to cough, rhinorrhea, nasal congestion, sore throat, and fever.
• Assessed by a clinician as having mild or moderate URTI, not requiring antibiotics or other systemic prescription medications, and suitable for observation and/or symptomatic management.
• Willing to discontinue other probiotics or supplements during the study period, except for the study product provided by the investigators.
• Agree to complete questionnaires and provide fecal biological samples during the study period.
• The child's parent(s) or legal guardian(s) are fully informed about the trial and voluntarily sign the written informed consent form before any study procedures begin.

Exclusion Criteria:

• Severe URTI as judged by the clinician, or symptoms severe enough to require antibiotics, corticosteroids, nebulized steroids/bronchodilators, or other systemic prescription medications, including suspected or confirmed bacterial infections such as bacterial tonsillitis, bacterial sinusitis, or acute suppurative otitis media.
• Acute lower respiratory tract infections, such as acute bronchitis, acute laryngitis/laryngospasm, or pneumonia.
• Respiratory tract infections secondary to underlying diseases, including primary immunodeficiency, acquired immunodeficiency syndrome, congenital airway malformation, congenital heart disease, gastroesophageal reflux, or abnormal lung development.
• Severe malnutrition.
• Severe chronic diseases, including severe asthma or other chronic respiratory diseases, chronic hepatitis or other liver diseases affecting liver function, renal insufficiency or other severe kidney diseases, neurological diseases such as epilepsy or neurodevelopmental disorders, and major genetic diseases such as chromosomal abnormalities or monogenic disorders.
• History of epilepsy or febrile convulsions.
• Planned vaccination during the trial period.
• Severe infection, severe trauma, or history of medium-to-major surgery within the past month, as judged by the investigator.
• Participation in other clinical intervention studies involving drugs, dietary supplements, probiotics, or prebiotics within the past 3 months.
• Known allergy to any component of the probiotic product.
• Any other condition that, in the opinion of the investigator, makes the participant unsuitable for this clinical study.