16Enterococcus faecalis is an early coloniser of the human infant gut and contributes to the development 17 of intestinal immunity. To better understand the functional capacity of E. faecalis we constructed a 18 broad host range RP4 mobilisable vector, pEHR513112, that confers chloramphenicol resistance and 19 used a metaparental mating approach to isolate E. faecalis AHG0090 from a faecal sample collected 20 from a healthy human infant. We demonstrated that E. faecalis AHG0090 is genetically tractable and 21 could be manipulated using traditional molecular microbiology approaches. E. faecalis AHG0090 was 22 comparable to the gold-standard anti-inflammatory bacterium Faecalibacterium prausnitzii A2-165 in 23 its ability to suppress cytokine mediated NF-B activation in human gut derived LS174T goblet cell-24 like and Caco-2 enterocyte-like cell lines. E. faecalis AHG0090 and F. prausnitzii A2-165 produced 25 secreted low molecular weight NF-B suppressive peptidic bioactives. Both bioactives were sensitive 26 to heat and proteinase K treatments although the E. faecalis AHG0090 bioactive was more resilient to 27 both forms of treatment. As expected, E. faecalis AHG0090 suppressed IL-1 induced NF-B-p65 28 subunit nuclear translocation and expression of the NF-B regulated genes IL-6, IL-8 and CXCL-10. 29Finally, we determined that E. faecalis AHG0090 is distantly related to other commensal strains and 30 likely encodes niche factors that support effective colonisation of the infant gut. 31