A metaproteomic pipeline to identify newborn mouse gut phylotypes

In order to characterize newborn mouse gut microbiota phylotypes in very early-life stages, an original metaproteomic pipeline, based on LC-MS(2)-spectra and Mascot driven NCBI non-redundant repository database interrogation was developed. An original computational analysis assisted in the generation of a taxonomic gut architecture from protein hits to operational taxonomic units (OTUs) and related functional categories. Regardless of the mouse's genetic background, a prevalence of Firmicutes (Lactobacillaceae) and Proteobacteria (Enterobacteriaceae) was observed among the entire Eubacteria taxonomic node. However, a higher abundance of Firmicutes was retrieved for Balb/c gut microbiota compared to Rag2(ko) mice, the latter was mainly characterized by a Proteobacteria enriched microbiota. The metaproteomic-obtained OTUs were supported, for the identification (ID) of the cultivable bacteria fraction, corroborated by axenic culture-based MALDI-TOF MS IDs. Particularly, functional analysis of Rag2(ko) mice gut microbiota proteins revealed the presence of abundant glutathione, riboflavin metabolism and pentose phosphate pathway components, possibly related to genetic background. The metaproteomic pipeline herein presented may represent a useful tool to investigate the highly debated onset of the human gut microbiota in the first days of life, when the bacterial composition, despite its very low diversity (complexity), is still very far from an exhaustive description and other complex microbial consortia.