FIG 3.

A focus on metabolomics as the methodology of choice for direct pathway reconstruction. Microbiome alterations in health and disease through sequencing of the 16S rRNA gene might not represent real changes in the active fraction (the 16S rRNA gene that is being transcribed) of microbes that change under a certain condition. Nevertheless, integration of 16S rRNA gene (rDNA) sequencing with shotgun metagenome sequencing and metaproteomics provides a more precise understanding of the function of the system. As an alternative solution, metabolomics alone could provide the pieces missing in pathway reconstruction and allow direct interference of function. Further integration with shotgun metagenomics, metaproteomics, or target gene amplicon sequencing would incorporate all layers of complexity in the biological system, allowing deep understanding of the mechanisms within.