Supplementary Material for: Physical Activity and Gastric Residuals as Biomarkers for Region-Specific NEC Lesions in Preterm Neonates

<b><i>Background:</i></b> Necrotizing enterocolitis (NEC) is a serious feeding-related inflammatory gut disease with high mortality. Early clinical markers of NEC are of great importance for optimizing preventive interventions. <b><i>Objective:</i></b> Using preterm pigs as models, we hypothesized that an early postnatal onset of NEC can be predicted by decreased physical activity during the first few days after birth.<b><i> Methods:</i></b> Cesarean-delivered preterm pigs were fed parenteral nutrition and increasing amounts of formula for 5 days after birth (n = 120). Their physical activity was quantified by a continuous camera surveillance system and they were evaluated twice daily for clinical signs of apathy, discoloration, respiratory distress, abdominal distension and diarrhea. The volume of gastric residuals and the presence of macroscopic NEC-like lesions in the stomach, intestine and colon were recorded at euthanasia on day 5.<b><i> Results:</i></b> Half of the pigs (48%) showed clear NEC-like lesions on day 5, and these individuals had more adverse clinical symptoms from day 3 but decreased physical activity already from day 2 relative to the unaffected pigs (both p < 0.05). Only animals with NEC lesions in the small intestine had lower physical activity on days 2 and 3, and the increased volume of gastric residuals was specifically related to colon lesions (both p < 0.05). <b><i>Conclusions:</i></b> Decreased physical activity precedes the clinical symptoms of NEC in the small intestine of preterm pigs, and increased gastric residuals predict NEC lesions in the colon. Physical activity and gastric residuals may function as clinical biomarkers for region-specific NEC lesions in preterm neonates.