CLINICAL RESEARCH ARTICLE Bifidobacterium breve BBG-001 and intestinal barrier function in preterm babies: Exploratory Studies from the PiPS Trial Paul Fleming 1,2 , Mark Wilks 3,4 , Simon Eaton 5 , Nicola Panton 4 , Richard Hutchinson 2 , Abena Akyempon 1 , Pollyanna Hardy 6 , Michael R. Millar 3,4 and Kate Costeloe 1,2 BACKGROUND: Uncertainty remains about the role of probiotics to prevent necrotising enterocolitis (NEC) some of which arises from the variety of probiotic interventions used in different trials, many with no prior evidence of potential efficacy. Mechanistic studies of intestinal barrier function embedded in a large probiotic trial could provide evidence about which properties of probiotics might be important for NEC prevention thus facilitating identification of strains with therapeutic potential. METHODS: Intestinal permeability, stool microbiota, SCFAs and mucosal inflammation were assessed from the second postnatal week in babies enrolled to a randomised controlled trial of B. breve BBG-001 (the PiPS trial). Results were compared by allocation and by stool colonisation with the probiotic. RESULTS: Ninety-four preterm babies were recruited across six nested studies. B. breve BBG-001 content was higher by allocation and colonisation; Enterobacteriaceae and acetic acid levels were higher by colonisation. No measure of intestinal barrier function showed differences. The PiPS trial found no evidence of efficacy to reduce NEC. CONCLUSIONS: That the negative results of the PiPS trial were associated with failure of this probiotic to modify intestinal barrier function supports the possibility that the tests described here have the potential to identify strains to progress to large clinical trials. Pediatric Research (2021) 89:1818–1824; https://doi.org/10.1038/s41390-020-01135-5 IMPACT: ● Uncertainty about the therapeutic role of probiotics to prevent necrotising enterocolitis is in part due to the wide range of bacterial strains with no previous evidence of efficacy used in clinical trials. ● We hypothesised that mechanistic studies embedded in a probiotic trial would provide evidence about which properties of probiotics might be important for NEC prevention. ● The finding that the probiotic strain tested, Bifidobacterium breve BBG-001, showed neither effects on intestinal barrier function nor clinical efficacy supports the possibility that these tests have the potential to identify strains to progress to large clinical trials. INTRODUCTION Necrotising enterocolitis (NEC) is the major gastro-intestinal emergency following pre-term birth—it is a devastating disease carrying high mortality and long-term morbidity. 1 The pathogen- esis remains incompletely understood and is complex and heterogeneous involving functional and anatomical immaturity of both the intestinal barrier and gastrointestinal immune systems, abnormal patterns of intestinal microbial colonisation, abnormal- ities in gastro-intestinal blood flow and a possible genetic predisposition. 2,3 Figure 1 outlines some of the characteristics of the foetal and preterm neonatal intestine that may predispose preterm babies to the development of NEC. There is no clear case definition for NEC, and while the global incidence varies widely, it is estimated to affect between 2 and 9% of babies born <1500 g birth weight. 4 Clinical management involving ‘resting’ the bowel and giving antibiotics with recourse to surgical intervention if necessary 5 has not changed for decades, and there is urgent need for evidence-based interventions both to prevent and treat disease. Over the past 20 years, there has been interest in the potential of enteral probiotics to prevent NEC. 6–10 There have been a number of meta-analyses all of which suggest evidence of benefit, one of the most recent including 38 trials involving 10,520 participants. 11 Despite this, clinicians remain divided about the therapeutic role of probiotics. Various aspects of the trials have been criticised, one of the frequent comments being the heterogeneity of the interventions in terms of bacterial strain, dosage and the number of strains given. 12 In the majority of these trials, the choice of probiotic appears to be based on availability rather than any pre-clinical evidence of benefit. The largest of the trials included in these meta-analyses, the Probiotics in Preterm Babies (PiPS) trial, was a placebo controlled trial of the probiotic Received: 2 March 2020 Revised: 9 August 2020 Accepted: 11 August 2020 Published online: 18 September 2020 1 Homerton University Hospital, NHS Foundation Trust, London, UK; 2 Centre for Genomics and Child Health, Queen Mary University of London, London, UK; 3 Department of Infection, Barts Health NHS Trust, London, UK; 4 Blizard Institute, Queen Mary University of London, London, UK; 5 UCL Great Ormond Street Institute of Child Health, University College London, London, UK and 6 Birmingham Clinical Trials Unit, University of Birmingham, Birmingham, UK Correspondence: Paul Fleming (Paul.fleming@qmul.ac.uk) www.nature.com/pr © International Pediatric Research Foundation, Inc 2020 1234567890();,: