348 - Investigating fungal pathogenesis in necrotizing enterocolitis using a neonatal mouse model

Friday, April 28, 2023

5:15 PM – 7:15 PM ET

Poster Number: 348
Publication Number: 348.134

Dhirendra Kumar. Singh, North Carolina Children's Hospital, Chapel Hill, NC, United States; Claire M. Miller, University of North Carolina at Chapel Hill School of Medicine, Carrboro, NC, United States; Corey M. Jania, UNC, Chapel Hill, NC, United States; Stephen Mackay, UNC Chapel Hill, Chapel Hill, NC, United States; Natalia S.. Akopyants, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, NC, United States; Yukihiro Yamaguchi, University of North Carolina at Chapel Hill, Chapel Hill, NC, United States; Misty Good, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, NC, United States

Presenting Author(s)

Dhirendra Kumar Singh, Dr. (he/him/his)

Post Doctoral Researcher
North Carolina Children's Hospital
Chapel Hill, North Carolina, United States

Background: Necrotizing enterocolitis (NEC) is a destructive disease affecting the intestine of premature infants. Recent studies established the significance of the gut microbiome and the virome in NEC pathogenesis. Though several studies have shown a fungal association with inflammatory bowel disease, there have been no studies evaluating the host-fungal and bacteria-fungal interactions in NEC. Therefore, we sought to determine whether fungal pathogens play a role in the pathogenesis of NEC.

Objective: Investigation of host-fungal and fungal-bacterial interactions in a neonatal mouse NEC model.

Design/Methods:

Four-day-old neonatal mice were either dam-fed or were randomly assigned to the following groups: NEC (formula supplemented with lipopolysaccharide (LPS) and human microbiota from an infant with NEC), NEC with Candida albicans (C.a.): NEC+C.a.; or formula plus C.a. without LPS or microbiota (C.a.). To induce experimental NEC, mice were kept in an infant incubator at 37⁰C, and orally gavaged a mixture of formula, LPS, and enteric bacteria six times daily at regular intervals followed by 10 minutes of hypoxic stress twice a day for three days. For fungal infection, pups were gavaged daily with 1x10⁷ CFU of C. albicans (SC5314) per mouse. Body weights were monitored every day. At the end of each experiment, animals were sacrificed, and ileum was collected for qPCR analysis and histology. Spleen, liver, kidney, stomach, ileum, and colon were collected to monitor the fungal colonization, and stool was collected for microbiota analysis.

Results: Our data show a significantly higher death rate in NEC+C.a. group compared to both the dam-fed (p=0.0016) or C.a. groups (p=0.0053). We also found a higher fungal load in the ileum (p=0.057), colon (p=0.0030), and spleen (p=0.0465) of NEC+C.a. mice compared to C.a. alone mice, demonstrating higher fungal colonization and dissemination in this group. Remarkably, the NEC+C.a. mice had significantly higher ileal gene expression of the antimicrobial peptide Reg3γ compared to mice that were dam-fed (p< 0.0001) or subjected to NEC (p< 0.0001).

Conclusion(s): We established that NEC+C.a. mice had a compromised epithelial barrier with higher dissemination of C. albicans to the spleen. In addition, we found significantly higher expression of Reg3γ in NEC+C.a. compared to the dam-fed and NEC mice. This indicates a potential role that Reg3γ plays in the host-immune response related to microbial and mycobiome composition in NEC.
Future studies will be dedicated to dissecting the precise role of the mycobiome and microbiome in host immunity crosstalk in NEC pathogenesis