Resident microbiota influence physiologic responses in many organs and are linked to many inflammatory, metabolic, neoplastic and behavioral disorders. Altered microbiota (dysbiosis) is associated with these conditions by vastly improved deep sequencing, metabolomic and analytic techniques. However, functional consequences, the primary vs. secondary nature of these compositional changes and role of individual bacterial species and combinations remain unknown. These functional properties are mechanistically addressed in gnotobiotic mice by precisely manipulating the microbiota in selectively colonized germ-free (GF) wild-type and genetically-engineered inbred mice with single or multiple resident or pathogenic microbial strains. Fecal microbial transplants (FMT) transfer healthy or diseased human or experimental animal’s fecal microbiota to GF mice to explore functional properties of dysbiotic complex bacterial communities. However, microbial and clinical heterogeneity between individuals with human complex disease are enormous and greatly affect FMT results. Most investigators use single or small numbers of fecal donors for FMT, leading to marked variability in results from different human donors. An important unmet research need is an easily accessible source of broadly representative and fully characterized human fecal samples for colonizing GF mice t achieve consistent reproducible results. Human FMT into gnotobiotic mice is a popular approach to test in vivo functional properties of dysbiotic bacterial communities from diseased patients and enhance clinical relevance of murine preclinical therapeutic models. We show low efficiency transfer of human fecal bacteria to GF mice, but enhanced transfer efficiency with this material from the murine recipient to another GF mouse. Transplant of this mouse-adapted (MA) human microbiota to other mice yields more reproducible colitis than did FMT of the original pooled human fecal samples, but with retention of the transferred human phenotype. Other uncertainties that limit reproducibility of human FMT to GF mice are the variable methods of fecal microbial transfer; frequency of administration, duration of equilibration after shipping mice prior to FMT, since most of our users do not have local gnotobiotic facilities to maintain sterility after receiving our mice, and type of housing after transfer. Carefully dissecting these variables will develop optimal protocols for handling and housing mice before and after FMT. We hypothesize that human fecal bacterial transfer to GF mice can be improved by optimizing transfer techniques and transferring fully characterized mouse-adapted pooled human fecal transplants into gnotobiotic inbred mice will yield highly reproducible results and stable phenotypes in recipient mice. Aim: Improve fecal transfer techniques, optimize housing conditions for recipient mice and characterize pooled complex mouse-adapted human fecal microbial communities from healthy donors and th...