MICrobiota, KEY between Mother and Offspring USsing SEquencing techniques

Necrotising enterocolitis (NEC) is the most prevalent acute
gastro-enterological disease in the Neonatal Intensive Care Unit (NICU). Its
incidence varies from 1-5 per 1000 live born children, and is associated with
prematurity and low birth-weight. NEC occurs in ~6% of premature children born
with a gestational age of 28-32 weeks, but increases to ~15% in children born <
28 weeks.1 Diagnosis is often difficult, just as predicting which child will
develop the disease. Signs and symptoms are often non-specific, such as
abdominal distension, gastric retention or feeding intolerance. Laboratory and
radiological tests also have limited diagnostic accuracy. The disease often
progresses rapidly, with potentially life-threatening complications such as
bowel perforation necessitating urgent laparotomy in around 50% of cases.
Morbidity and mortality are therefore high, in some series up to 40%.
NEC is a multifactorial disease. Several predisposing factors have been
identified from which aberrant bacterial colonization of the gut, formula
feeding, mode of delivery, an immature immune system, aberrant bile salts and
changes in intestinal perfusion and oxygenation are major predisposing factors.
Despite all research, this complex interplay of both maternal and neonatal
factors has not been unraveled yet. Furthermore, no study has ever combined
analyses of all these predisposing factors in the development of NEC in one
cohort. Doing so will give us the chance to investigate the relation and
interaction between the different aspects of the development of NEC and to
predict or diagnose NEC at an earlier stage.

In the present study we aim to address several relevant clinical and
pathophysiological questions. Data from the present study will be used to
identify a bundle of interventions we can subsequently test to decrease the
incidence of NEC. More specifically we aim to:

1. Identify the origin of the gut microbiota in neonates at a high risk for NEC
by investigating the maternal microbiota in amniotic fluid, placenta, vagina,
rectum and in mother*s milk as well as in formula feeding and correlate this
with the developing gut microbiota of the infant.
2. Investigate the neonatal innate immune response in umbilical cord blood and
correlate this immune response with microbiotal colonization and markers for
gut wall intergrity and gut wall inflammation and oxygenation.
3. Identify quantitative differences in DNA methylation of marker genes in
enterocyte DNA isolated from stool samples.
4. Investigate the role of specific fecal bile salts in NEC development and
investigate whether bile salts are related to certain microbiota, gut wall
integrity, breastfeeding and/or antibiotics.
5. Generate the fecal SCFA profile in neonates with imminent NEC.
6. Generate the metabolic profiles in neonates with imminent NEC using
metabolomics technology.
7. Relate intestinal perfusion and oxygenation levels and variability to
neonatal gut microbiome, and metabolic profiles.
8. Relate tissue perfusion and oxygenation levels and variablity to levels of
ischemia modified albumin (IMA).
9. Analyze DNA damage in buccal cells and relate this to the neonatal innate
immune response in umbilical cord blood and to microbiotal colonization.
10. Investigate whether any of the parameters mentioned above can be used to
predict NEC early, properly diagnose NEC, either alone or in combination.

Prospective cohort study

This study will be performed on neonates and their mothers. As there are no
extra invasive measurements, risks and burden can be considered negligible,
with the exception of the minimal burden for mothers regarding the swabs
antenatally. Cerebral and abdominal NIRS is already performed on a routine
clinical basis, and all neonates are continuously clinically monitored in the
neonatal intensive care setting. As its sets out to identify prognostic factors
for the development of necrotising enterocolitis, the study can only be
performed in neonates as NEC only occurs in neonates. Results from this study
can be important in the future care of babies at a high risk for NEC or its
complications.