In:
Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 112, No. 2 ( 2015-01-13)
Kurzfassung:
Microbial communities are essential to the function of virtually all ecosystems and eukaryotes, including humans. However, it is still a major challenge to identify microbial cells active under natural conditions in complex systems. In this study, we developed a new method to identify and sort active microbes on the single-cell level in complex samples using stable isotope probing with heavy water (D 2 O) combined with Raman microspectroscopy. Incorporation of D 2 O-derived D into the biomass of autotrophic and heterotrophic bacteria and archaea could be unambiguously detected via C-D signature peaks in single-cell Raman spectra, and the obtained labeling pattern was confirmed by nanoscale-resolution secondary ion MS. In fast-growing Escherichia coli cells, label detection was already possible after 20 min. For functional analyses of microbial communities, the detection of D incorporation from D 2 O in individual microbial cells via Raman microspectroscopy can be directly combined with FISH for the identification of active microbes. Applying this approach to mouse cecal microbiota revealed that the host-compound foragers Akkermansia muciniphila and Bacteroides acidifaciens exhibited distinctive response patterns to amendments of mucin and sugars. By Raman-based cell sorting of active (deuterated) cells with optical tweezers and subsequent multiple displacement amplification and DNA sequencing, novel cecal microbes stimulated by mucin and/or glucosamine were identified, demonstrating the potential of the nondestructive D 2 O-Raman approach for targeted sorting of microbial cells with defined functional properties for single-cell genomics.
Materialart:
Online-Ressource
ISSN:
0027-8424
,
1091-6490
DOI:
10.1073/pnas.1420406112
Sprache:
Englisch
Verlag:
Proceedings of the National Academy of Sciences
Publikationsdatum:
2015
ZDB Id:
209104-5
ZDB Id:
1461794-8
SSG:
11
SSG:
12
