SRA

Summary: Incidence of necrotizing enterocolitis (NEC), which remains a leading and intractable cause of mortality in preterm infants, is attenuated by feeding human breast milk (HBM). We report that this diminution correlates with alterations in the gut microbiota, particularly enrichment of Propionibacterium species. Transfaunation of microbiota from HBM-fed preterm infants, or a Propionibacterium strain, P. UF1, cultured therefrom, to mice conferred protection against pathogens and correlated with profoundly increased intestinal Th17 cells, sustained regulatory T cells (Tregs), and reduced pathogen-induced inflammation. Th17 cell differentiation was dependent on dihydrolipoamide acetyltransferase (DlaT) of the bacterial surface layer. Binding of P. UF1 with its cognate receptor, SIGNR1, resulted in the activation of intestinal phagocytes. Importantly, P. UF1 significantly mitigated induced NEC in neonatal mice. Together, these results mechanistically elucidate the protective effects of HBM and P. UF1-induced immunoregulation that safeguard against proinflammatory diseases, including NEC. Purpose: The goal of this study is to discover differentially epxressing genes between wild-type P. UF1 and DdlaT P. UF1 bacteria to reveal the critical role of DlaT in P. UF1 metabolism and potentially link it with P. UF1-induced protective immunity in the mice Methods: Total RNA was extracted from P. UF1 and ?dlaT P. UF1 strains cultured in MRS medium. The ScriptSeq Complete Gold Kit was used for rRNA depletion and the cDNA library generation. Subsequently, cDNA libraries were evaluated, pooled and sequenced on the Illumina MiSeq machine, which generates 7-8 million paired-end reads (~75nt) per sample. The sequence reads first underwent data preprocessing and quality evaluation including trimming off the low quality ends of reads. Rockhopper was used to map sequence reads to annotated P. UF1 genome, normalize the counts values by upper quartile normalization and perform differential expression analysis using NB statistical model Results and conclusion: Data demonstrated that deletion of dlaT in P. UF1 resulted in significant activation of genes involved in the glycolytic pathway and significant changes in metabolic pathways, potentially influencing DC and T cell regulatory responses. However, these induced metabolic alterations are not critical for antigen-dependent Th17 response accordingly. Overall design: mRNA profiles of wild-type and DdlaT P. UF1 bacteria were generated by deep sequencing, in triplicate, using Illumina Miseq.