  Simultaneous identification and metabolic analysis of microbes with
single-cell resolution and high throughput is necessary to answer the question
of "who eats what, when, and where" in complex microbial communities. Here, we
present a mid-infrared photothermal-fluorescence in situ hybridization
(MIP-FISH) platform that enables direct bridging of genotype and phenotype.
Through multiple improvements of MIP imaging, the sensitive detection of
isotopically-labelled compounds incorporated into proteins of individual
bacterial cells became possible, while simultaneous detection of FISH labelling
with rRNA-targeted probes enabled the identification of the analyzed cells. In
proof-of-concept experiments, we showed that the clear spectral red shift in
the protein amide I region due to incorporation of $^{13}$C atoms originating
from $^{13}$C-labelled-glucose can be exploited by MIP-FISH to discriminate and
identify $^{13}$C-labelled bacterial cells within a complex human gut
microbiome sample. The presented methods open new opportunities for single-cell
structure-function analyses for microbiology.
