Background: Determining whether associations between gut microbiota characteristics and host physiology represent causal relationships is a fundamental challenge for microbiome research. One way these associations can be investigated is to instil donor faecal material into gnotobiotic mice and to assess the extent to which donor phenotype is recapitulated. However, the manner in which this process is performed varies considerably between studies, and assessment of microbiota re-establishment in recipient animals is not always carried out. We report a detailed investigation of microbiome assembly in germ-free mice and compare the effects of single and multiple rounds of faecal gavage, using both native and antibiotic-disrupted donor material.
Results: Levels of bacteria within the faeces of recipient animals increased rapidly following the instillation of donor material. However, considerable instability in microbiota composition continued during the first two weeks post-gavage, with substantial changes in taxon relative abundance occurring in parallel to declining faecal pH. Relative compositional stability was not achieved until day 28 and persistent differences between recipient and donor microbiota remained. These included an increased relative abundance of Bacteroidetes, and a reduced relative abundance of Firmicutes. Of taxa detected in donor material, 52% were represented in stable recipient microbiota following transplantation with native faecal material (single gavage), increasing to 66% following three rounds of gavage. These taxa accounted for 95% and 91% of total donor bacterial abundance, respectively. Performing multiple rounds of gavage significantly increased microbiota similarity between donor and recipient, and significantly reduced within-group dispersion (P<0.05). Instillation of antibiotic-associated microbiota resulted in substantially lower temporal and inter-animal variance, with multiple rounds of gavage providing no substantial benefit.
Conclusions: Microbiome assembly in recipient animals is not immediate and several weeks are required for microbiota stability to be achieved. Multiple rounds of faecal gavage result in greater similarity to donor microbiota and reduced inter-animal variance. The process of donor microbiota re-establishment, and therefore the interval required prior to investigations using recipient animals, is influenced by donor microbiota characteristics.