Functional Single-Cell Approach to Probing Nitrogen-Fixing Bacteria in Soil Communities by Resonance Raman Spectroscopy with 15N2 Labeling |
Cui,L(Cui,Li);Yang,K(Yang,Kai);Li,HZ(Li,Hong-Zhe);Zhang,H(Zhang,Han);Su,JQ(Su,Jian-Qiang);Paraskevaidi,M(Paraskevaidi,Maria);Martin,FL(Martin,FrancisL.);Ren,B(Ren,Bin);Zhu,YG*(Zhu,Yong-Guan) |
Nitrogen (N) fixation is the conversion of inert nitrogen gas (N2) to bioavailable N essential for all forms of life. N2-fixing microorganisms (diazotrophs), which play a key role in global N cycling, remain largely obscure because a large majority are uncultured. Direct probing of active diazotrophs in the environment is still a major challenge. Herein, a novel culture-independent single-cell approach combining resonance Raman (RR) spectroscopy with 15N2 stable isotope probing (SIP) was developed to discern N2-fixing bacteria in a complex soil community. Strong RR signals of cytochrome c (Cyt c, frequently present in diverse N2-fixing bacteria), along with a marked 15N2-induced Cyt c band shift, generated a highly distinguishable biomarker for N2 fixation. 15N2-induced shift was consistent well with 15N abundance in cell determined by isotope ratio mass spectroscopy. By applying this biomarker and Raman imaging, N2-fixing bacteria in both artificial and complex soil communities were discerned and imaged at the single–cell level. The linear band shift of Cyt c versus 15N2 percentage allowed quantification of N2 fixation extent of diverse soil bacteria. This single-cell approach will advance the exploration of hitherto uncultured diazotrophs in diverse ecosystems. |
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Volume:90 Page:5082-5089 Journal:ANALYTICALCHEMISTRY |
http://dx.doi.org/10.1021/acs.analchem.7b05080
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