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Background: E. coli is a leading cause of neonatal sepsis. Newborns ingest E. coli, which transcytoses the gut producing bacteremia. Lactobacillus spp. decrease E. coli gut transcytosis but the mechanisms involved in this protective effect are not well understood.

Objective: To determine the effects of Lactobacillus pretreatment on the intestinal microbiota and inflammation in neonatal rats orally infected with E. coli.

Design/Methods: Newborn rats were orally pretreated on day of life (DOL) 1 and 2 with four doses of 107 colony forming units (CFU) of Lactobacillus rhamnosus GG (LGG) or PBS. On DOL 2, pups received orally 106 CFU of neonatal E. coli bacteremia strain SCB34 or PBS. On DOL 7, distal colon with stool were collected for 16S sequencing. AbundantOTU+ was used to generate de novo operational taxonomic units (OTUs). Observed, ACE, Shannon, and Simpson indices were used for alpha diversity. Bray-Curtis and Jaccard indices were used for beta diversity. Linear discriminant analysis effect size (LEfSe) was used to determine differences at the genus level. Expression of ICAM-1, GRO1, Toll-like receptor 4 (TLR4), and Single-Immunoglobulin Interleukin-1 Related Receptor (SIGIRR) was measured in ileal homogenates by real-time PCR.

Results: Phyla distribution clearly distinguished the SCB34-infected experimental groups (Fig. 1). We also observed greater alpha diversity in the SCB34-infected groups (Fig. 2; A-D). Bray-Curtis analyses showed differences in microbial abundance between the groups receiving LGG pretreatment vs. PBS prior to SCB34 infection. However, the microbial composition of these two groups was similar per Jaccard index (Fig. 2; E-F). LEfSe results showed greater abundance of Lactobacillus in both SCB34-infected groups, and surprisingly, lower abundance of Escherichia-Shigella genera in these groups compared to controls (Fig. 3; A, B). LGG pretreatment produced a significant decrease in anaerobes including Clostridium, Romboutsia and Veillonella (Fig. 3; C-E). LGG prophylaxis significantly suppressed expression of ICAM-1, GRO-1 and TLR4, while inducing the TLR4-inhibitor, SIGIRR (Fig. 4).

Conclusion(s): LGG pretreatment significantly modified clinically relevant microbiota features of neonatal pups orally infected with E. coli, and attenuated E. coli-induced intestinal inflammation. Harnessing these relevant mechanisms afforded by probiotics such as LGG will provide novel preventive and therapeutic interventions against gut-derived neonatal sepsis.

Presented at the 2021 PAS Virtual Conference

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Infectious Disease | Pediatrics

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Presented at the 2021 PAS Virtual Conference

Neonatal Gut Microbiota Alterations and Local Inflammation Induced by Escherichia coli Infection are Modified by Lactobacillus rhamnosus Prophylaxis