The goal of this clinical trial is to investigate whether administering a probiotic (Infloran®) to infants who received antibiotics in the first 28 days of life can restore or enhance their immune response to routine vaccines.

Antibiotic use in the first weeks of life can lower the levels of beneficial gut bacteria, such as bifidobacteria, which play a key role in immune function. As a result, infants treated with antibiotics may produce fewer antibodies after routine vaccinations, leaving them less protected against infections.

The main questions this study aims to answer are:

• Does treatment with the probiotic Infloran® improve the geometric mean concentrations (GMCs) of anticapsular antibodies against at least 11 serotypes included in the pneumococcal conjugate vaccine (PCV20) in serum samples collected at 6 months of age compared with placebo in infants treated with antibiotics in the neonatal period?
• Does treatment with the probiotic Infloran® improve the GMCs for the pneumococcal conjugate vaccine (PCV20) at 12 months of age compared with placebo in infants treated with antibiotics in the neonatal period?
• Does treatment with the probiotic Infloran® improve the GMCs of other routine childhood vaccines at 6 and 12 months of age compared with placebo in infants treated with antibiotics in the neonatal period?
• Does treatment with the probiotic Infloran® increase the proportion of infants achieving seroprotective antibody levels for pneumococcal antigens compared to placebo in infants treated with antibiotics in the neonatal period?
• What are the differences in antigen specific T cell responses, flow cytometry, blood transcriptomics, and gut microbiota composition in the probiotic (Infloran®) vs placebo groups in infants treated with antibiotics in the neonatal period?

Researchers will compare infants who receive Infloran® (a probiotic containing Bifidobacterium bifidum and Lactobacillus acidphilus) with those who receive a placebo (which contains the same excipients as Infloran® but does not contain any bacterial strains).

Participants will:

• Be randomly assigned to receive either a 14-day course of probiotic Infloran® or a placebo.
• Provide blood samples (3-5 mL) at 6 weeks, 6.5 weeks (optional blood-draw for exploratory endpoint), 6 months and 12 months of age.
• Provide stool samples at four timepoints: prior to starting the intervention (probiotic/placebo), on day 7, on day 14 after completion of the study supplement, and prior to their first vaccination at 6 weeks of age.
• Receive routine vaccinations at 6 weeks, 4 months and 6 months in line with the National Immunisation Program
• Complete surveys to collect information regarding probiotic/placebo administration and vaccination related side effects

This study aims to recruit 360 infants to assess whether this probiotic treatment following antibiotic exposure improves the immunogenicity of vaccinations. The information from this study will improve our understanding of how probiotic intervention can support optimal immune responses to vaccination in early life. The findings could potentially influence public health strategies, offering a new way to support optimal vaccine responses in antibiotic-treated infants.

Inclusion Criteria:

1. Infants administered antibiotics in the first 28 days of life. Infants must have documented direct antibiotic exposure - defined as having received at least 36 hours of antibiotic treatment in the neonatal period (the first 28 days after birth).
2. Gestational age ≥ 35 weeks.
3. Birth weight ≥ 2500g.
4. Mother aged at least 18 years and able and willing to provide written informed consent for themselves and their infant.
5. Parent/guardian agrees to not give any other probiotics to their infant prior to vaccination at 6 weeks, including any formula that contains probiotics.
6. Infant planning to receive all nationally approved vaccines during next 6 months.

Exclusion Criteria:

1. Significant medical condition in either the mother or infant that, in the opinion of a medical investigator, may interfere with the study.
2. Infant had confirmed sepsis or other serious infection in the neonatal period.
3. Infants with known congenital diseases or who are immunocompromised or considered medically at risk (MAR).
4. Infant participating in another interventional trial during the trial period.

Non-identifiable individual participant data, from participants who give specific consent, may be made available long-term for use by future researchers from a recognised research institution whose proposed use of the data has been ethically reviewed and approved by an independent committee and who accept SAHMRI's conditions, under a collaborator agreement, for accessing:

• Individual participant data that underlie the results reported in our articles after deidentification (text, tables, figures and appendices)
• Study protocol, Statistical Analysis Plan, PICF

Molecular data generated from this study (i.e. transcriptomic, epigenomic and metagenomic) will be labelled with a unique identification code and stored on an appropriate data repository (e.g. Gene Expression Omnibus, Sequence Read Archive) with demographic information (age and sex). No identifying participant information will be shared.

At the time of publication. Data will be available indefinitely.

Researchers from a recognised research institution can approach SAHMRI for access of data. The researcher will need to provide evidence that the proposed use of the data has been ethically reviewed and approved by an Institutional Review Board (IRB)/ Human Research Ethics Committee (HREC), and accept SAHMRI's conditions, under a collaborator agreement.