The Influence of Diet and Exposure to Antibiotics on Gut Microbiota Development in Early Life

Research output: Book/ReportPh.D. thesisResearch

  • Shamrulazhar Bin Shamzir Kamal
The human gut microbiota (GM) has a strong mutual relationship with its host. It plays vital functions in postnatal development of the immune system, metabolism homeostasis and protection against pathogens. At birth, the human gastrointestinal tract (GIT) is nearly sterile, but right after birth, it is initially colonized by facultative anaerobes such as Enterobacteriaceae, Streptococcus, Enterococcus and Staphylococcus spp. Subsequently, the GM gradually becomes dominated by bifidobacteria and obligate anaerobic bacteria such as Bacteroides and clostridia. The progression of early life GM colonization and maturation are influenced by a range of factors, for example, mode of delivery, maternal microbiota, gestational age, genetics, mode of feeding, and exposure to antibiotics.
In the past few decades, we have witnessed an immense increase in our understanding of the factors that shape our GM, but despite this progress, it remains only partly understood how early life perturbations such as exposure to antibiotics and different feeding regimes influence the development of the GM. This notion is true for infants delivered at term, but even more profound for preterm infants, where huge knowledge caps on e.g. the interactions between feeding regimes, GM development and short and long-term health of the infants still exist.
Preterm birth is associated with immaturity of many organs, including the GIT. The colonization of the GIT with microorganisms may affect organ maturation and host immunity. Due to prematurity of the immune system and vital organs such as lung, brain, and GIT, infants born very preterm (26-32 weeks of gestation) have an increased risk of developing serious postnatal infections such as necrotizing enterocolitis (NEC). The causes of NEC are still not fully understood, but some studies have revealed that too rapid advancement of feeding (formula milk) is linked with higher NEC risk, but delaying the feeding (or advancing too slow) is also associated with adverse outcomes. The right timing of feeding and availability of specific bioactives are thus crucial issues for the preterm infant’s overall organ development at in early age.
This PhD forms a part of a larger gut‐brain‐immunity-gut microbiota research consortium, (“Early milk and microbiota to support immunity, gut and brain development, NEOMUNE”) focusing on preterm neonates and investigating the influence of external factors on early life GM colonization and immunity. Specifically, this PhD project aimed at evaluating the influences of gestational age, feeding regimes and the effect of antibiotic exposure in early life on the composition of GM, bacterial load, bacterial resistance towards antibiotics and the production of metabolites with particular focus on short chain fatty acids (SCFA).
The main findings of the present PhD-thesis have been published in 4 manuscripts. In Manuscript 1, we first investigated the effect of feeding regime (early enteral feeding (EEF) and total parenteral nutrition (TPN)) and gestational age (preterm (PT) and term (T)) on the establishment of GM of neonates short and more long-term using piglets as a model for infants. Here, we hypothesized that reduced gestational age at birth (versus birth at normal term) and the first enteral feeding (versus no enteral feeding) induce changes in gut colonization that last beyond the first weeks after birth. The study aimed to investigate the effects of early life feeding on gut microbiota composition at day 5 and day 26 after birth. GM composition was determined by 16S rRNA gene-amplicon sequencing and qPCR. The piglets were fed (day 0 till day 5) with two different diet regimes, EEF supplemented with bovine colostrum and TPN. The composition of GM, quantity of bacteria and bacterial metabolites (SCFA) were also evaluated. We demonstrated that the establishment of GM in early life (Day 5) was influenced by early feeding strategy as well as the gestational age of the neonates. However, only the gestational age had a profound effect on the GM composition and its SCFA production more long term (26 days).
In Manuscript 2 and 3 we studied the effects of early life antibiotic administration enterally or parenterally using preterm piglets as a model for preterm infants. Preterm infants are at increased risk of developing infections and necrotizing enterocolitis (NEC). Antibiotics are thus commonly used to prevent and treat NEC and other morbidities. Prematurity, abnormal gut bacterial colonization and aggressive enteral nutrition are important factors that increase the risk of NEC development. We hypothesized that only enteral antibiotics promote maturation of systemic immunity and delay gut bacterial colonization and these thereby may have protected preterm neonates against both NEC and bacteraemia. It was found that early life administration of antibiotics influenced gut bacterial colonization and reduced inflammation, and NEC lesions in new-born, formula-fed preterm pigs. The delayed bacterial colonization of the gut might support intestinal structure, function, and immunity in the immediate postnatal period of formula-fed preterm neonates. Delaying gut microbiota colonization by using enteral antibiotics within the first few days after preterm birth may protect and support the intestine to mature because there is less interference from colonizing microbes and bacterial fermentation products.
In Manuscript 4 we investigated the effect of the very early life (perinatal) antibiotic exposure (cefuroxime) on GM colonization of caesarean section (CS) delivered infants. In this experiment, we randomly assigned forty-two pregnant women scheduled for elective CS to receive a single prophylactic dose of cefuroxime (1500mg) either shortly before skin incision or immediately after the umbilical cord clamping. In the latter case, the infant will be exposed to antibiotics via the umbilical cord. We hypothesized that very early life exposure to antibiotics potentially changes the composition of the GM of infants and it may also increase the occurrence of antibiotic-resistant bacteria in the gut. Fecal samples were collected at day 10 and 10 months from the infants and the GM composition were then determined by 16S ribosomal RNA gene amplicon high-throughput sequencing. Further, Gram-positive cocci and Enterobacteriaceae were isolated and identified before antimicrobial susceptibility tests were performed by disk diffusion. Our study found that there are no clear differences in GM composition between infants whose mothers received cefuroxime either shortly before or after CS. Though surprisingly, at 10 months of age, but not at 10 days of age, the number of observed species was higher in the infants where the mothers received cefuroxime after cord clamping. Additionally, antimicrobial susceptibility testing similarly showed no susceptibility differences among Enterobacteriaceae, Enterococcus spp., and Staphylococcus spp. isolated at 10 days of age between the two groups of infants.
In conclusion, the present PhD thesis and the publications forming part of the thesis have contributed to our knowledge and understanding on gut microbiota colonization in early life and the factors that influence the colonization, more specifically early diet and gestational age, mode of antibiotic administration (enteral and parenteral) and antibiotic administration during CS. Our first conclusion suggests that early diet and gestational age have a profound effect on the GM very early in life (age of 5 days) but only gestational age showed more long-term (4 weeks) effects on GM. Secondly, we also investigated the mode of antibiotic delivery and its effect on infants GM early in life. Our study suggests enteral administered antibiotics contrary to parenteral antibiotics which had a strong effect on GM in term of abundance. Furthermore, limiting or delaying gut bacterial colonization in preterm neonates may benefit systemic immunity and protect against NEC. Finally, we show that the timing of antibiotic administration
to mothers undergoing CS (term infants) has no or only negligible effect on GM colonization short and more long term (10 months of life).
Original languageEnglish
PublisherDepartment of Food Science, Faculty of Science, University of Copenhagen
Number of pages105
Publication statusPublished - 2021

ID: 286303231