Intestinal ecosystem of the very and extremly preterm infant: microbiota analysis, implications in short and long term outcomes – EPIFLORE

Included infants belong to the French national cohort EPIPAGE 2. The follow up will be over 12 years. The birth gestional age was less than 32 weeks. Fecal samples were collected at one week, one month and at hospital discharge. A supplementary sample was collected in case of NEC. Microbiota analyses are performed by pyrosequencing of the ARN 16S gene, and also by culture in case of NEC.

The first results concern NEC cases. Fourteen cases matched with 79 healthy controls were analyzed. By culture, a significant association between NEC and colonization by Clostridium and by Staphylococcus aureus has been found. Pyrosequencing analyses confirm these results. Concomitant analyses of the 110 NEC cases of the whole Epipage 2 cohort confirm the major role of microbiota in NEC onset.

The current management of NEC is empiric due to the lack of knowledge of its pathogenesis. Evidence between a specific bacterial pattern and the onset of NEC can lead to have an early biological marker of NEC. This could allow to propose a specific treatment of NEC and a reliable preventive approach as well.

A manuscript about the clinical and biological factors associated to NEC cases is in progress, based on clinical data from EPIPAGE 2 and microbiological data from EPIFLORE.

Prematurity, and more particularly severe prematurity, is currently increasing and represent a major cause of mortality and morbidity. During their first weeks of life, very preterm infants, i.e. born at a gestational less (GA) than 33 weeks, are at an increased risk of infections such as late-onset sepsis and gastrointestinal diseases, in particular neonatal necrotizing enterocolitis (NEC). Besides, relationships between prematurity and/or small birthweight and further chronic diseases have been reported. Currently, an ever-increasing body of evidence implicates the gut microbiota in defining states of health and disease. Indeed, dysbiosis is thought to lead to a multitude of ailments, from the obvious case of inflammatory diseases to allergy or obesity and development of the gut microbiota over the first weeks of life appears therefore a crucial step. This has led to the interest in the development of strategies that aim at manipulating the gut bacterial colonization using administration of probiotics and/or prebiotics.
Recent studies have described the abnormal gut colonization in preterm infants, especially very preterm ones. This abnormal pattern is thought to be a risk factor for the onset of early or late onset diseases. Hence, probiotics supplementation in preterm infants has been recommended by some expert committees. However, if several meta-analyses have shown the interest of such supplementation, controversies have emerged, in particular for very preterm neonates. An increase of knowledge of the gut microbiota in very preterm infants is therefore needed.
The project EPIFLORE is lying within this context, and aims at conducting a large scale analysis of the gut microbiota establishment in very preterm infants. For this purpose, we take the opportunity of two French cohorts, set up in 2011: (1) EPIPAGE 2 that included all infants born preterm in France and (2) ELFE that included fullterm and late preterm infants. All included infants will be followed until 12 years of age. The project EPIFLORE will have three aims: (1) to study on a very large scale the gut microbiota establishment in very preterm infants through a multicentric study taking into account both the variability between infants and between neonatal units, (2) to investigate the short and long term clinical evolution of very preterm infants as compared with fullterm and late preterm infants, and (3) to investigate the relationships between gut microbiota patterns and occurrence of diseases such as early- and late-onset sepsis and NEC in very preterm infants. Six hundred and sixty very preterm infants from 19 neonatal intensive care units (level III) and belonging to the cohort EPIPAGE 2 have been included. Fecal samples (about 1900), prospectively collected between day 3 and 7, day 23 and 30, and at hospital discharge (i.e. between 34 and 36 weeks corrected age) will be analyzed using a cutting-edge high-throughput technique, i.e. 454-GS FLX-Titanium pyrosequencing of the amplified bacterial 16S ribosomal RNA, which will give a qualitative and quantitative molecular species composition. Fecal samples collected at maternity discharge (between day 3 and 5) from 200 late preterm and fullterm infants belonging to the cohort ELFE will be included as a control group. Perinatal determinants which could influence the bacterial establishment will be analyzed. Comparison of short and long term clinical evolution between infants belonging from both cohorts allow determining the risk for developing short and long term outcomes for very preterm infants.