Fig. 1

A mixed Flow Cytometry-Next Generation Sequencing strategy to identify human host-microbial associations. Body samples (saliva, faeces, urine, mucosa, milk, etc.) are disaggregated by vortexing and mild sonication. Microbial cells are fixed in 4 % paraformaldehyde previous to staining with fluorescent markers to detect cells (e.g. by DNA markers [6]), active cells (e.g. by RNA markers [17]) and specific antibodies (e.g. by anti-human IgA) through a flow cytometer. Microbial load can also be accurately estimated by cell counting. Cells are sorted depending on whether they are opsonized with either IgA or IgG antibodies. Each bacterial population can then be PCR-amplified and pyrosequenced, characterizing the microorganisms which evade the immune system and those which are recognized by each immunoglobulin. The application of the technique to healthy and diseased individuals may unravel the contribution of the immune response to microbial infections and polymicrobial diseases