Objectives and Study

High protein intake during infancy results in accelerated early weight gain and potentially later obesity. Infant formula with reduced protein concentration, closer to that of breastmilk, may reduce this risk. In the present study, we evaluate effects on growth and metabolism at 12 months after intervention with modified protein-reduced formulas from 2 to 6 months. We recently reported appropriate growth in the low-protein formula groups up to 6 months, and similar weight gain 4-6 months in infants receiving low-protein formula with alpha-lactalbumin enriched whey (α-lac-EW) and breastfed (BF) infants. Protein metabolism was closer to BF infants in the modified protein-reduced formula groups.

Methods

In a randomized, double-blinded, controlled prospective trial 245 healthy term infants received low-protein formulas with either α-lac-EW (1.75 g protein/100 kcal) or casein glycomacropeptide-reduced whey (CGMP-RW, 1.76 g protein/100 kcal) or standard infant formula (SF, 2.2 g protein/100 kcal) from 2 to 6 months of age. 83 BF infants served as reference group. At follow-up at 12 months, anthropometrics and dietary intake were assessed, and blood analysed for insulin, C-peptide and insulin-like growth factor 1 (IGF-1).

Results

At 12 months, 85% of the children remained in the study. Weight gain (g/d) between 6 and 12 months and body mass index at 12 months were higher in SF than in the BF group (p=0.019, p < 0.001, respectively), but comparable in the low-protein formula groups and BF group. SF group had higher serum insulin (p <0.001) and C-peptide (p=0.003) than the BF group, whereas both low-protein formula groups had serum concentrations comparable to the BF group. Serum IGF-1 was similar in all study groups as was dietary intake of macronutrients and energy.

Conclusions

Reduced protein formula enriched with α-lac-EW or CGMP-RW during early infancy results in growth and metabolic profile more comparable to breast-fed infants at 12 months.

Objectives and Study

Addition of bovine milk fat globule membrane (bMFGM) in infant formula may better approximate complex milk lipid composition in human milk and support development of the maturing gut microbiota.

Methods

This multicenter, double-blind, controlled, parallel-group, pilot study compared microbiota outcomes in healthy term infants (7-18 days of age) randomized to receive cow’s milk-based infant formula (Control, n=18) or similar formula with added whey protein-lipid concentrate (5 g/L, source of bMFGM; INV-MFGM, n=19) over a 60-day feeding period. A human milk reference group (HM, n=17) received mother’s-own milk over the same period. Stool and oral swabs were collected at Baseline (BL; 4-18 days of age) and Study Day 60 (+7 days). DNA was extracted, 16S rRNA genes sequenced, and amplicon sequence variants (ASV) assigned with DADA2. Alpha-diversity was calculated in R. ASV abundance was compared using Kruskal-Wallis and DESeq2.

Results

Stool data was available for 33 participants. Baseline alpha-diversity measures were similar by group and remained stable in the HM group (Figure, A). Alpha-diversity increased from BL to Day 60 for INV-MFGM and Control groups and both groups were higher than HM by Day 60, primarily due to increases in Clostridia species. Streptococcus, Klebsiella, and Clostridium species were higher and Lachnoclostridium, Ruminococcus, and two Bifidobacterium species were significantly lower for HM vs formula-fed groups at Day 60 (Figure, B). Akkermansia, Bacteroides, and Hungatella species were higher in INV-MFGM and Streptococcus species were lower compared to Control. No group or longitudinal differences in oral microbiota were detected.

Conclusions

Distinct patterns of neonatal microbiome establishment were demonstrated for infants receiving mother’s-own milk compared to routine cow’s milk-based infant formulas with or without added bovine MFGM, including higher prevalence of some Bifidobacterium species in infants receiving formula.

Objectives and Study

Toddlerhood is a critical period for bone mass acquisition. This study aimed at evaluating the impact of the consumption of a young child formula (YCF) containing Limosilactobacillus (L., previously Lactobacillus) reuteri + galacto-oligosaccharides (GOS) on bone quality in healthy young children.

Methods

In this randomized, double-blind controlled clinical trial performed in the Philippines, children aged 2-3 years received a YCF containing a synbiotic composed of L. reuteri DSM 17938 (at concentration that guarantees 10exp8 cfu/day) + GOS (4g/L) (experimental, EXPL, n=91) or a minimally fortified powdered milk mimicking local traditional milks (control, CTRL, n=91) for 6 months. Children were advised to consume 2 servings/day of the investigational products. The tibia quality was assessed at baseline and after 3- and 6-months measuring speed of sound (m/s) by ultrasound as an integrative readout of bone density, geometry, and microstructure. A linear mixed model with repeated measures was applied with fixed effects for treatment group, gender, visit, visit x treatment interaction, and baseline values for speed of sound, blood vitamin D and body mass index).

Results

Mean age at baseline was 29.6 ± 3.6 months and 53% were female. The tibia speed of sound measured at 6 months as primary outcome was significantly higher in EXPL compared to CTRL. The estimated average difference was 58.91 (95% CI = (21.34, 96.48) m/s), p=0.002, in the full analysis set (FAS). A positive effect was already observed after 3 months of intervention 53.00 (95% CI = (7.05, 98.95) m/s), p=0.024 (Figure). The effect estimated on the per protocol population at 6 months was supportive of the primary endpoint (p=0.005).

Conclusions

Consumption of a young child formula containing the synbiotic L. reuteri + GOS promotes bone strength, improving bone quality in healthy young children.

Objectives and Study

We previously reported that partly fermented infant milk formula (IMF) lowers the incidence of infantile colic in healthy term-born infants (Vandenplas et al. 2017). This IMF contains a specific fermented milk formula (Lactofidus^TM fermentation process using bacterial strains Streptococcus thermophilus 065 and Bifidobacterium breve C50) with prebiotic oligosaccharides scGOS/lcFOS (0.8g/100ml, 9:1). So far, mechanistic insight in how this IMF, currently designated IMF with postbiotics, impacts gut homeostasis is limited. Therefore, our current objective was to assess if IMF with postbiotics affects visceral sensitivity, and to define possible underlying mechanisms.

Methods

Wistar rats received either IMF with postbiotics and prebiotics, IMF with or without prebiotics (1ml/day) or NaCl as vehicle control (0.9%, 1 mL/day) for 14 days. Abdominal contractions upon colorectal distension (CRD) were measured under basal conditions and after partial restrained stress to determine visceral sensitivity. Metabolomics was performed on IMFs. Effects of selected metabolites were further investigated using cell culture.

Results

Under basal conditions, none of the IMFs modified visceral sensitivity in response to CRD compared to control. Only IMF with postbiotics limited both stress-induced visceral hypersensitivity and gut hyperpermeability.

Metabolomic analysis identified imidazole-lactate (IML) and indole-lactate (ILA) as being increased in IMF with postbiotics with levels closer to what is found in human breast milk.

In vitro we found that addition of IML, but not ILA, to Caco-2 cells reduced the acetylcholine-induced release of serotonin, a modulator of pain perception. Further, AhR signaling, which can also regulates (neuropathic) pain, was activated by the combination of IML and ILA.

Conclusions

IMF with postbiotics reduces stress-induced visceral hypersensitivity and hyperpermeability. Cell culture studies suggest that certain metabolites that are higher in IMF with postbiotics, such as IML and ILA, influence AhR and serotonin signaling pathways and might thereby influence visceral sensitivity. Further mechanistic studies are ongoing.

Objectives and Study

Infections caused by drug resistant Gram-negative bacteria (DR-GNB) are a major health concern for hospitalized preterm neonates, globally. The aim of this study was to investigate the effect of a multi-strain probiotic on the incidence of peri-rectal colonization with DR-GNB in preterm neonates.

Methods

A double-blind, placebo-controlled, randomized clinical trial was conducted including 200 neonates, randomly allocated to a probiotic (n = 100) or placebo (n = 100) group. Neonates received daily supplementation with either a multi-strain probiotic or placebo for up to 28 days. Sigma transwabs were used to perform peri-rectal swabs at enrollment, day 7 and day 14 of life. Swab tips were inoculated onto selective media (ChromID ESBL and ChromID CarbaSmart) for isolation of Extended-spectrum beta-lactamases- and carbapenem resistant GNB (including carbapenemase-producing Enterobacterales). The organisms were identified to species level using API10E strips.

Results

Fifteen percent of the neonates showed peri-rectal colonization with DR-GNB on the day of enrolment indicating probable maternal-to-neonate (vertical) bacterial transmission or environmental acquisition at time of delivery, with no difference between groups. Acquisition of further DR-GNB colonization was rapid, with an increase from 15% on the day enrolment to 77% by day 7 and 83% by day 14 of life. By day 7 (corresponding to early gut colonization), neonates in the probiotic group were 57% less likely to have peri-rectal DR-GNB colonization [OR: 0.43 (0.20–0.95); p = 0.04] and by day 14 (corresponding to late gut colonization), neonates in the probiotic group were 93% less likely to have peri-rectal DR-GNB colonization [OR: 0.07 (0.02–0.23); p < 0.001].

Conclusions

Hospitalized neonates showed substantial peri-rectal colonization with DR-GNB at enrolment and further rapid acquisition of DR-GNB in the first 2 weeks of life. The use of a multi-strain probiotic was effective in reducing early (day 7) and late (day 14) neonatal gut colonization with DR-GNB.

Objectives and Study

In the first period of life, foundations of lifelong immune homeostasis and microbial colonization in the gut are established. Optimized diets during this vulnerable period may be most effective for improving health. Here, we address this window of opportunity through advanced integration of ex vivo and artificial intelligence (AI) technologies that represent the infant/piglet gut environment and use this novel platform to evaluate gut health benefits of early life nutrition ingredients.

Methods

Early life-specific gut health was modelled by integrating two technologies, 1) ex vivo human infant-like gut tissue model (early life piglet InTESTine™), and 2) AI modeling of human infant gut-immune function into a visualization tool, a ‘gut health space model’. Docosahexaenoic acid (DHA) was selected as benchmark food ingredient to set-up and validate the novel research pipeline.

Results

Gut tissue of pre-weaned piglet was used to configure the early life InTESTine™. Six-hour exposure of colon tissue to 50 µM DHA resulted in proper tissue functionality (transcellular/paracellular transport >2), intact tissue integrity (FITC-dextran 4000 leakage maximally 1.4%/h) and preserved tissue viability (cumulative lactate dehydrogenase release of 5.9%). Furthermore, 2174 differentially expressed genes (DEGs) were discovered by RNA sequencing. Gene set enrichment and pathway analysis indicated that DHA affects multiple mechanisms such as decreased allergy-related processes and inflammation, increased wound healing and gut barrier function. These predictions of gut health outcomes were in line with clinical data from literature.

Conclusions

Here, we demonstrated a novel integrated early life-specific platform to evaluate early life nutrition ingredients in order to support and improve gut health in early life. Next, we will incorporate an in vitro human infant microbiome screen (early life i-screen) as third technology into the workflow. This, as well as testing other early life nutrition (benchmark) ingredients will further shape the gut health space model.