R. D. Shelby1, L. Mashburn-Warren2, J. B. Navarro2, N. Tengberg1, J. Allen2, M. T. Bailey2, S. D. Goodman2, G. E. Besner1 1Nationwide Children’s Hospital, The Research Institute, Center for Perinatal Research,Pediatric Surgery,Columbus, OH, USA 2Nationwide Children’s Hospital, The Research Institute,Center For Microbial Pathogenesis,Columbus, OH, USA
Introduction: Despite decades of research, necrotizing enterocolitis (NEC) remains the most common gastrointestinal (GI) surgical emergency in the premature infant. While the pathogenesis of NEC remains unclear, alterations in the intestinal microbiome likely play a mechanistic role. Our laboratory has demonstrated the efficacy of Lactobacillus reuteri(Lr) in protecting the intestines from experimental NEC when the probiotic is administered in its biofilm state. When grown as a biofilm on maltose-loaded dextranomer microspheres (DM), the Lr-DM-maltose product significantly restores the intestinal microbiome, and decreases the incidence and severity of NEC after a single dose. Lr exhibits both anti-inflammatory and anti-microbial properties – the latter attributed to the production of reuterin, an anti-microbial compound that inhibits harmful gut bacterial growth. In the current study, we developed a reuterin-deficient mutant formulation of Lr (Lr-gldC) to examine the importance of the antimicrobial effects of Lr in protecting the intestines from NEC.
Methods: Using a validated rodent model of NEC, premature rat pups were delivered via C-section and subjected to repeated episodes of hypercaloric formula feeding, hypoxia, and hypothermia for 96 hours. Rat pups were randomized to one of several treatment groups prior to initiation of formula feeding and stress: 1) NEC + sterile water (vehicle control, N=51), 2) NEC + planktonic (free-living) Lr (N=45), 3) NEC + Lr on maltose loaded microspheres (Lr + DM-maltose; N=56), 4) NEC + Lr-gldC (mutant form of Lr) (N=12); or 5) NEC + Lr-gldC + DM-maltose (N=42). Control pups were unstressed and breast fed (N=13).Rat pups were sacrificed when they exhibited signs and symptoms of NEC, or after 96 hours. Intestinal tissue was collected at sacrifice for blinded histopathological analysis.
Results: As expected, compared to no treatment, administration of a single dose of Lr in its biofilm state (Lr+ DM-maltose) significantly decreased the incidence of NEC (67% vs. 18%, p< 0.0001), whereas administration of Lr in its planktonic state had no significant effect. In contrast, compared to administration of Lr+ DM-maltose, administration of Lr-gldC + DM-maltose resulted in significant (albeit not total) loss of beneficial effects (18% vs.41%, p=0.008).
Conclusion: In conclusion, the ability of Lr to produce reuterin contributes to, but is not solely responsible for, the ability of the probiotic to reduce both the severity and incidence of NEC. Other factors such as the anti-inflammatory properties of Lr may also play a role, and will be investigated in future studies. These results may contribute to our understanding of the etiology of NEC, and will be important as we translate this technology to the bedside.
