The development of early life gut microbiota in health and allergic disease

The development of the gut microbiome from birth through to childhood is thought to be important for establishing a healthy symbiosis, but much is yet to be learnt about this phase of microbiome development. Perturbations of gut microbiome during this development have been associated with the pathology of diseases, such as allergy. In Chapter 2, we describe this dynamic period of gut microbiome development, the various factors involved in shaping its composition and the importance for the concurrent maturation of the immune system. Environmental factors including birth mode, exposure to antibiotics and household exposures (such as siblings and furry pets) represent important factors impacting its development, and which also have been epidemiologically implicated in the risk to develop allergic disease. Several paediatric studies indeed associated altered gut microbiota with development of allergic disease. Breastfeeding represents the most significant factor in shaping early life microbiome and is associated with several short-term and long-term health benefits, including a lower risk of developing allergic disease. Specific prebiotic or synbiotic (when combined with probiotics) ingredients added to infant formula may exert similar effects on gut microbiota composition and activity, which may benefit infants for whom breastfeeding is not possible.

Bifidobacteria are the most abundant bacteria in breastfed infants, but often under-represented in 16S rRNA surveys of diversity, due to poor DNA extraction techniques, poor PCR primer choice or a combination of both. In Chapter 3, we optimized a commonly used “universal” PCR primer set and demonstrated the effective recovery of this genus without compromising the detection of other genera. In Chapter 4 we applied the optimized pyrosequencing method described in Chapter 3 to analyse the gut microbiota of infants at high-risk of developing allergy, who participated in a clinical trial that investigated the effects of a partially hydrolysed protein formula supplemented with prebiotics on the prevention of eczema. We showed that the taxonomic composition of infants receiving the prebiotic-supplemented formula was closer to that of breastfed infants when compared to infants receiving an infant formula based on intact protein without prebiotics, which was driven by increased abundance of Bifidobacterium spp. and decreased abundances of Clostridium spp. and Lachnospiraceae spp. Importantly this also led to specific changes in gut eco-physiology characterized by a more acidic pH and increased levels of lactate and acetate as also characteristic for breastfed infants. In a nested case-control, we found that infants who developed eczema by 18 months of age showed an altered development of bacterial taxa and metabolites around the time that complementary feeding was started. The patterns identified suggested that the establishment of specific bacteria that utilize lactate and acetate to produce butyrate may have a role in protecting from the development of eczema.

Infants who suffer from severe CMA often rely on cow's milk protein avoidance and, when breastfeeding is not possible, on specialised infant formulas such as amino-acid based formulas (AAF) to meet their nutritional needs and in order to resolve the allergic symptoms they suffer from. In Chapter 5, we investigated the modulatory effects of an AAF supplemented with synbiotics on the gut microbiota and showed that both composition and activity approximated that of an age-matched breastfed reference group as opposed to infants receiving the AAF without synbiotics. Similar as observed in Chapter 4 this was driven by an increase of the Bifidobacterium spp./ Lachnospiraceae spp. ratio (B/L-ratio).

In Chapter 6 we screened the intestinal microbiota of a small set CMA-infants and healthy controls to select donor samples for faecal microbiota transplantation into germ-free mice. We confirmed the decreased B/L-ratio in CMA versus healthy infants as observed in Chapter 5, which was maintained upon transplantation into the germ-free mice. Herein, we showed that CMA-associated infant microbiota resulted in an atopic orientation, with increased immunoglobulin E levels and an enhanced responsiveness to cow’s milk allergen upon sensitization, which suggested that the pathobiology of allergic disease is mediated at least in part by gut microbiome perturbation.