Assessing Role of Probiotics in Children Aged 6-36 Months Treated for Pneumonia

This study is a randomized, double-blind, placebo-controlled clinical trial conducted at Phu Tho Provincial Obstetrics and Pediatrics Hospital to evaluate Bacillus spore probiotics delivered by nasal-oropharyngeal spray (LiveSpo® Navax) with or without an oral probiotic suspension (LiveSpo® Preg-Mom) as an adjunct to standard medical treatment in children aged 6-36 months hospitalized with pneumonia.

A total of 330 eligible children will be randomly assigned in a 1:1:1 ratio into three groups (n = 110 per group):

the Control group receiving 0.9% sodium chloride nasal-oropharyngeal spray and oral RO water, the Navax group receiving Bacillus spore probiotic nasal-oropharyngeal spray and oral RO water, and the Navax-PregMom group receiving Bacillus spore probiotics both as nasal spray and oral suspension. All participants receive standard-of-care treatment according to hospital guidelines, with differences between groups limited to the assigned study interventions.

The nasal-oropharyngeal spray is administered as three sprays per nostril and throat per dose, three times daily, and the oral intervention is administered at a daily dose of 5 mL, twice daily. Interventions are initiated during hospitalization and continued for up to two months from admission.

Rationale and need: Pediatric pneumonia often proceeds upper-airway infection and is characterized by pathogen colonization of-and inflammation within the naso-/oropharyngeal mucosa. Accordingly, a nasal-throat Bacillus spore spray could serve as a local adjunct to maintain airway hygiene and mucosal restitution while potentially mitigating the load of respiratory viruses and bacterial co-pathogens at the site of infection. Concomitant interventions for hospitalized children with pneumonia, including supportive care and administration of medications (such as antibiotics), could perturb the gut microbiota and lead to gastrointestinal symptoms before or after treatment. An oral Bacillus spore suspension is expected to complement the local intervention by acting through the gut-immune interface, thereby supporting systemic immune modulation and recovery beyond the upper airway.

Crucially, most of the available clinical trials examine probiotics as monotherapy (e.g., oral or nasal alone), and did not sufficiently address outcomes related to combining both routes in the same treatment. The purpose of this trial is to directly examine the hypothesis that there will be an additive effect of providing concurrent Bacillus nasally (B. subtilis ANA4 + B. clausii ANA39) and orally (B. subtilis ANA46 + B. clausii ANA39 + B. coagulans ANA40) administered probiotics in hospitalized children with pneumonia. The expected mechanism is that the combined regimen may better support treatment and prevention of recurrence by reducing viral load, bacterial co-infection, and pro-inflammatory cytokines, while improving nasal and gut microbiota profiles.

After written informed consent is obtained, baseline demographic and clinical data are collected using standardized questionnaires and medical records.

During inpatient treatment, physicians and nurses monitor and record daily clinical symptoms, including fever, cough, tachypnea, chest indrawing, wheezing, crackles, and gastrointestinal manifestations, as well as length of hospital stay and use of medications such as antibiotics, antiviral agents, and supportive therapies. Nasopharyngeal, blood and stool samples are collected at predefined time points according to the protocol (mandatory at day 0 and day 3, and optional at day 30 and day 60). All of these samples are processed to quantify inflammatory cytokines (IL-6, IL-8, IL-10, IL-17, IL-23, TNF-α) and IgA using ELISA, and to detect respiratory viruses, bacterial co-pathogens, and B. subtilis, B. clausii, and B. coagulans by real-time PCR. Stratified subsamples of stool and nasopharyngeal samples (n = 15-20 per group x 2 time points) undergo 16S rRNA gene sequencing to assess changes in respiratory (at day 30 vs. day 0) and gut microbiota (at day 30 vs. day 0) diversity and taxonomic composition. After discharge, children are followed up at day 30 and/or day 60 through outpatient visits or structured interviews with parents or caregivers to document recurrence of respiratory or gastrointestinal symptoms, rehospitalization, and post-discharge medication use. Parents or caregivers are instructed not to administer other probiotic products (except yoghurt) or Bacillus spore nasal preparations during the study period.

All medical records are collected, scanned, and stored in electronic format. Patient data are systematically organized into databases for analysis. Reduction in pathogen load is assessed by comparing real-time PCR cycle threshold (Ct) values at discharge or follow-up with baseline using the 2^-ΔCt method. Changes in cytokine and IgA concentrations are evaluated by comparing median values before and after intervention. The presence of B. subtilis, B. clausii, and B. coagulans in samples is expressed using Ct values from TaqMan probe assays. Statistical analyses and graphical presentations are performed using GraphPad Prism (version 10.2.3), IBM SPSS Statistics (version 27), and R software (version 4.4.2), with a two-sided significance level set at p < 0.05. Randomization and double blinding are applied to minimize random and systematic bias, and treatment allocation codes remain concealed until completion of data analysis, except in cases of serious adverse events requiring emergency unblinding.

Inclusion criteria

• Children (male/female) aged 6-36 months diagnosed and hospitalized for treatment for pneumonia.
• Parents or guradians of the children agree to participate in the study, explain, and sign the research consent form.

Exclusion criteria

• Children who need ventilator support.
• Children who have had rhinoplasty surgery, nasal ulcers, nasal polyps or any nasal deformity that affects the use of the study product.
• Children who get transferred or leave the treatment unit (not for professional reasons).
• Children who have experienced congenital immunodeficiency or infectious disease (eg, HIV).
• Children who are concurrently participating in another clinical trial or participated in a clinical trial within 30 days before selection.
• Children who are under long-term medication (more than 7 consecutive days) within 30 days before selection.
• Children with allergy or hypersensitivity to any ingredient of the study product.