Human Milk Oligosaccharides, Gut Bifidobacterium, Vitamin D, and Infant Immunity (HMO-BIFIDI)

Human Milk Oligosaccharides in Breast Milk and Its Relation to Gut Bifidobacterium, Vitamin D and Immune Modulation in Infants

The goal of this observational study is to learn how certain natural sugar components in breast milk may be linked with vitamin D level, healthy gut bacteria, and immune markers in healthy breastfed infants. The study will include exclusively breastfeeding mothers and their healthy infants aged 1 to 2 months.

The main questions it aims to answer are:

• Is this natural sugar in breast milk linked with the baby's vitamin D level?
• Is this natural sugar in breast milk linked with healthy gut bacteria?
• Is this baby's vitamin D level linked with healthy gut bacteria?
• Is this natural sugar linked with immune factors in breast milk? Participants will answer health questions from questionnaire, undergo a brief physical examination, allow breast milk and a small baby blood sample to be collected, and provide an baby stool sample.

Researchers will test these samples in the laboratory and analyze the results to answer the research questions.

Study Overview

• Status: Active, not recruiting
• Conditions: Healhty

Detailed Description

Subject recruitment was conducted during immunization activities at community health centers (Puskesmas) and integrated health posts (Posyandu). The subject collection technique used the quota sampling method, a non-probability subject selection technique carried out by determining the desired sample size (quota) from a population with predetermined criteria, and researchers collected sample units until the target quota was reached. Researchers provided an explanation of the research protocol and obtained informed consent from participants regarding their willingness to participate in the study. Participants who met the eligibility criteria (inclusion and exclusion criteria) were declared as research subjects. Next, a research questionnaire was completed, including recording the subject's identity (mother and baby), the mother's health and nutritional history, childbirth and breastfeeding history, observation of the baby's stool, history and risk factors for allergies, and a general physical examination.

Mature breast milk samples will be analyzed to determine maternal secretion status, HMO status, Bifidobacterium in breast milk, and the status of immune markers such as sCD14, TGF-beta, and IgA in breast milk. Stool samples will be analyzed to determine the infant's intestinal Bifidobacterium. Blood samples will also be analyzed to determine vitamin D levels. Statistical correlations between these parameters will be performed to determine the research objectives.

The collected data is grouped based on the purpose and type of data, then the appropriate statistical method is selected, namely:

1. Univariate analysis (Used to describe basic data characteristics, in the form of frequency, mean value, standard deviation, and range)

2. Bivariate analysis

   1. Comparison Test (To see differences between groups)

      • Chi-Square Test → If the variable is categorical (e.g., the relationship between maternal secretory status (Secretory vs. Non-Secretory) and delivery method (Vaginal vs. Cesarean section). An alternative test, the Fisher's Exact test, is performed if the basic assumptions of the Chi-Square are not met.
      • T-Test (Independent T-Test) → If you want to compare the means of two groups (e.g., the difference in vitamin D levels between male and female infants). An alternative test, the Mann-Whitney test, is used if the data is not normally distributed.
      • ANOVA Test → If you want to compare the means of more than two groups (e.g., the difference in vitamin D levels based on the timing of early breastfeeding initiation: <1 hour, 1-6 hours, >6 hours). An alternative test, the Kruskal-Wallis test, is used if the data is not normally distributed.

   2. Correlation Analysis (To see the relationship between two numeric variables)

      • Pearson Correlation → If the data is normally distributed.
      • Spearman Correlation → If the data is not normally distributed or is ordinal

• Study Type: Observational
• Enrollment (Actual): 100

Contacts and Locations

Study Locations

• Indonesia
  • South Sulawesi
    • Makassar, South Sulawesi, Indonesia, 90245
      • Hasanuddin University

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Child; Adult; Older Adult
• Accepts Healthy Volunteers: Yes

• Sampling Method: Non-Probability Sample

Study Population

The population in this study was a group of breastfeeding mothers and their babies who came to visit immunization activities at integrated health posts/community health centers in Makassar City, Indonesia

Description

Inclusion Criteria:

1. Maternal Subject :

   • Women aged at least 18 years who are physically healthy;
   • Have a healthy baby aged 1-2 months with a full-term gestational age (37-42 weeks) and a normal birth weight (2500-4000 grams);
   • Exclusively breastfeeding;
   • Agree to participate in this study by signing an informed consent form.

2. Infant Subjects :

   • All infants of mothers who meet the inclusion criteria

Exclusion Criteria:

1. Maternal Subject :

   • Currently participating in another clinical trial;
   • Having a condition such as a breast abscess or other breast pathology;
   • Currently taking medications for conditions that may affect breast milk, such as antibiotics;
   • Subjects who cannot comply with the study protocol

2. Infant Subjects:

   • Currently participating in another clinical trial;
   • Having a history of antimicrobial treatment (oral or parenteral antibiotics);
   • Subjects are unable to comply with the research protocol.

Study Plan

How is the study designed?

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Analysis maternal Human Milk Oligosaccharides (HMO) and Breast Milk (BM) Bifidobacterium and their correlation to gut Bifidobacterium, serum vitamin D and immune regulatory status in infants	This outcome analyze the correlation between maternal human milk oligosaccharide (secretor status, 2FL' level), Breast Milk (BM) Bifidobacterium, gut Bifidobacterium, serum vitamin D and immune regulatory status in infants in infants aged 1 to 2 months.	At enrollment (single time point, cross-sectional assessment)

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Association between maternal HMO and infant gut Bifidobacterium	Association between maternal human milk oligosaccharide (secretor status and 2FL' level) and Bifidobacterium abundance in infant stool.	At enrollment (single time point, cross-sectional assessment)
Association between maternal HMO and infant vitamin D level	Association between maternal human milk oligosaccharide (secretor status and 2FL' level) and vitamin D level in infant serum.	At enrollment (single time point, cross-sectional assessment)
Association between maternal HMO and breast milk immune markers	This outcome assesses the association between human milk oligosaccharide (HMO) (secretor status and 2FL' level) in breast milk and immune markers in breast milk, including soluble CD14 (sCD14), transforming growth factor beta (TGF-beta), and immunoglobulin A (IgA).	At enrollment (single time point, cross-sectional assessment)
Association between breast milk Bifidobacterium and infant gut microbiota	This outcome assesses the association between Bifidobacterium composition in breast milk and gut microbiota (Bifidobacterium) composition in infant stool samples.	At enrollment (single time point, cross-sectional assessment)
Association between breast milk Bifidobacterium and infant vitamin D level	This outcome assesses the association between Bifidobacterium composition in breast milk and vitamin D level in infant serum.	At enrollment (single time point, cross-sectional assessment)
Association between breast milk Bifidobacterium and breast milk immune marker	This outcome assesses the association between Bifidobacterium composition in breast milk and immune markers in breast milk, including soluble CD14 (sCD14), transforming growth factor beta (TGF-beta), and immunoglobulin A (IgA).	At enrollment (single time point, cross-sectional assessment)

Collaborators and Investigators

• Sponsor: Hasanuddin University
• Collaborators: DSM-Firmenich AG
• Investigators: Principal Investigator: Bahrul Fikri, MD, Ph.D, Department of Pediatrics, Faculty of Medicine, Hasanuddin University; Study Director: Andi Raisyiah Akrimah Imran, MD, MHA, Department of Public Health and Family Medicine, Faculty of Medicine, Hasanuddin University

Study record dates

Study Major Dates

• Study Start (Actual): July 30, 2025
• Primary Completion (Actual): October 23, 2025
• Study Completion (Estimated): May 30, 2026

Study Registration Dates

• First Submitted: April 29, 2026
• First Submitted That Met QC Criteria: April 29, 2026
• First Posted (Actual): May 6, 2026

Study Record Updates

• Last Update Posted (Actual): May 6, 2026
• Last Update Submitted That Met QC Criteria: April 29, 2026
• Last Verified: April 1, 2026

More Information

Terms related to this study

• Keywords: Vitamin D; Gut microbiota; Bifidobacterium; Breast milk; HMO; Immune modulation
• Other Study ID Numbers: UH25030200

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: NO
• IPD Plan Description: No individual participant data will be made publicly available. The study is supported by commercial funding from DSM-Firmenich, and participant-level data are subject to confidentiality and data-sharing restrictions under the research collaboration agreement. Any data access or sharing would require prior written approval from DSM-Firmenich and must comply with applicable ethical and regulatory requirements.

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No