Early life gut microbiota profiles linked to synbiotic formula effects: a randomized clinical trial in European infants.

The American journal of clinical nutrition. 2023;117(2):326-339
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Microbial colonisation of the intestine after birth is a central event that influences infant health with life-long consequences. Although improvement of hygienic conditions reduces infant mortality due to infections, environments with low microbial biomass counteract natural colonisation by commensal microbes. The aim of this study was to assess the effects of a synbiotic intervention formula (IF) on faecal microbiota. This study was a multicentre, randomised, controlled, double-blind intervention trial which enrolled 540 infants. Infants whose parents had chosen not to breastfeed or were not able to breastfeed prior to study inclusion were allocated randomly to 1 of 2 formula groups (n = 230 control formula, n = 230 IF). The infants in the breastfed reference group (n = 80) were mainly fed human milk. Results showed that synbiotic intervention influenced the gut microbiota and milieu parameters during early life to resemble some major characteristics found in breastfed infants (higher relative abundances of bifidobacteria, lower richness, lower faecal pH and butyrate concentrations), and effects depended on the ecosystem profile of the infants. Authors conclude that specific randomised, controlled studies that focus on infants born by Caesarean section and how early nutrition can support the beneficial development of their microbiota are needed.

Abstract

BACKGROUND Microbial colonization of the gastrointestinal tract after birth is an essential event that influences infant health with life-long consequences. Therefore, it is important to investigate strategies to positively modulate colonization in early life. OBJECTIVES This randomized, controlled intervention study included 540 infants to investigate the effects of a synbiotic intervention formula (IF) containing Limosilactobacillus fermentum CECT5716 and galacto-oligosaccharides on the fecal microbiome. METHODS The fecal microbiota from infants was analyzed by 16S rRNA amplicon sequencing at 4, 12, and 24 months of age. Metabolites (e.g., short-chain fatty acids) and other milieu parameters (e.g., pH, humidity, and IgA) were also measured in stool samples. RESULTS Microbiota profiles changed with age, with major differences in diversity and composition. Significant effects of the synbiotic IF compared with control formula (CF) were visible at month 4, including higher occurrence of Bifidobacterium spp. and Lactobacillaceae and lower occurrence of Blautia spp., as well as Ruminoccocus gnavus and relatives. This was accompanied by lower fecal pH and concentrations of butyrate. After de novo clustering at 4 months of age, overall phylogenetic profiles of the infants receiving IF were closer to reference profiles of those fed with human milk than infants fed CF. The changes owing to IF were associated with fecal microbiota states characterized by lower occurrence of Bacteroides compared with higher levels of Firmicutes (valid name Bacillota), Proteobacteria (valid name Pseudomonadota), and Bifidobacterium at 4 months of age. These microbiota states were linked to higher prevalence of infants born by Cesarean section. CONCLUSIONS The synbiotic intervention influenced fecal microbiota and milieu parameters at an early age depending on the overall microbiota profiles of the infants, sharing a few similarities with breastfed infants. This trial was registered at clinicaltrials.gov as NCT02221687.

Expert Review


Conflicts of interest: None

Take Home Message:
  • Infant gut colonisation differs in vaginal versus cesarean section deliveries and between breastfed and infant formula practices.
  • Both enriched strain-specific probiotic and standard infant formula were shown to have a marked effect on microbiota colonisation in infants at age 4 months.
  • By the age of 2 years, however, there is no significant difference between breastfed and formula fed infants.

Evidence Category:
  • A: Meta-analyses, position-stands, randomized-controlled trials (RCTs)
  • X B: Systematic reviews including RCTs of limited number
  • C: Non-randomized trials, observational studies, narrative reviews
  • D: Case-reports, evidence-based clinical findings
  • E: Opinion piece, other

Summary Review:
Introduction

This randomised controlled intervention study compared gut health parameters with the use of a synbiotic pre- and probiotic strain enriched infant formula with human milk and standard formula at three intervals over a period of 2 years.

Methods

This was a double-blinded controlled study of 540 infants from France and Belgium. Participants were randomly allocated to 2 formula groups (n = 230 Control Formula (CF), n = 230 Intervention Formula (IF)) and the breastfed reference group (n = 80) as well as delivery mode (Cesarean and vaginal delivery). The synbiotic IF was a standard infant formula enriched with prebiotic GOS (0.02 g/g) and the probiotic strain L. fermentum CECT5716 (at least 1.0 × 106 cfu/g).

Stool analysis was conducted at three time intervals, 4, 12, and 24 months (infant age). Biomarkers included short chain fatty acids, pH, secretory IgA, calprotectin, and various bacterial phyla via microbiota analysis.

Results

  • At 4 months, the IF group tested higher for Bifidobacterium spp., and Lactobacillaceae and lower occurrence of Blautia spp., as well as Ruminoccocus gnavus and relatives compared to CF. They also had lower fecal pH and butyrate levels
  • Both the formula cohorts had lower SigA and more basic pH values than the human milk cohort, as well as higher prevalence of anaerobes belonging to the bacterial genera Akkermansia, Collinsella, and Faecalibacterium.
  • By age 24 months, the IF cohort exhibited increased levels of Akkermansia, Escherichia-Shigella, and R.gnavus. However there were no significant differences between the formula fed and human milk cohort at this time interval.
  • The differences observed at 4 months disappeared over time, except for a significantly higher relative abundance of bifidobacteria and Faecalibacterium spp. in IF infants at 12 months compared with CF infants.

Conclusion:

Although prominent differences between the cohorts were observed at 4 months, it appears that by the age of 2 years, there is little observable difference. This is most likely due to gut ecosystem maturation. The paper draws attention to the fact that changes to microbiota following treatment were more pronounced in infants who tested lower in occurrences of Bacteroides spp at age 4 months. Of note is the prevalence of cesarean birth deliveries in this cohort thereby indicating potential improved alternative feeding options when breastfeeding is not possible for these infants.

Clinical practice applications:
  • Probiotic L.fermentum and prebiotic galacto-oligosaccharide enriched infant formula appears to the improve infant microbiome, when compared to that of breastfed infants.
  • The most receptive infants were those born via cesarean section.

Limitations to consider:

  • The sample groups were from France and Belgium, with no indication as to culture, socio-economic, or sex distribution.
  • The two infant formula groups were n=230 each with only 80 infants in the breastfed reference group.
  • There was no indication of maternal diet practices pre-, during, and post- pregnancy.
  • Stool samples were not collected from the infants at baseline visit prior to formula intervention.

Considerations for future research:
  • Future studies need to include more diverse cultural and socio-economic cohorts to ascertain the potential influence of parental diet in baseline infant microbiome.
  • It is imperative to establish what role solid food choices, generally introduced at 6 months, might have on gut ecosystem maturation.
  • It would be useful to have a larger cesarean section birth cohort to compare to vaginal deliveries for more definitive results.

Lifestyle medicine

Fundamental Clinical Imbalances : Digestive, absorptive and microbiological
Patient Centred Factors : Mediators/Gut microbiota
Environmental Inputs : Microorganisms
Personal Lifestyle Factors : Environment
Functional Laboratory Testing : Stool
Bioactive Substances : Probiotics ; Synbiotic

Methodological quality

Jadad score : 4
Allocation concealment : Yes

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