Heavy water-labeled Raman spectroscopy reveals carboxymethylcellulose-degrading bacteria and degradation activity at the single-cell level
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Biodegradation of cellulose-rich substrates is an indispensable process for soil carbon replenishment in various ecological niches. Biodegradation of cellulose has been studied extensively via an enzyme assay to quantify the amount of cellulase with a view to identify effective cellulose degraders. However, a bulk enzyme assay undermines the role of physiological heterogeneity between cells; it is therefore imperative to opt out for a more effective method such as single-cell Raman spectroscopy combined with heavy water (D2O) to reveal active cellulose degraders. Cellular incorporation of D2O-derived D produces a new C-D Raman band which can act as a quantitative indicator of microbial activity. In this study, metabolic responses of seven cellulose-degrading bacteria to carboxymethylcellulose (CMC) and glucose were evaluated via the C-D Raman band. On the basis of % C-D, CMC was demonstrated to be most efficiently metabolized by Bacillus velezensis 2a-9 and Providencia vermicola 5a-9(b). Metabolic activity between individual cells of B. velezensis and P. vermicola towards CMC ranged from approximately 8 to 27% and 6 to 16%, respectively, clearly indicating heterogeneous degradation activities among isogenic populations. Linear correlation between % C-D and specific endoglucanase activity validated Raman results on CMC-degrading activity. Also, % C-D obtained from bacteria cultivated with only glucose was around 60% higher than that obtained from CMC, indicating the preference of bacteria for simple sugar glucose than CMC. In conclusion, Raman spectroscopy combined with heavy water is a sensitive analytical technique to reveal cellulose degraders and their degrading activities.
KeywordsSingle-cell microbiology Raman spectroscopy Heavy water isotope labeling Cellulose biodegradation Heterogeneous degradation activity
This study was funded by the National Key Research and Development Program of China (2017YFD0200201, 2017YFE0107300), Natural Science Foundation of China (21777154), the Strategic Priority Research Program of Chinese Academy of Sciences (XDB15020302, XDB15020402), CAS President’s International Fellowship Initiative (PIFI) for 2017, and K.C.Wong Education Foundation.
Compliance with ethical standards
This research does not involve the use of human or animals.
Conflict of interest
The authors declare that they have no conflicts of interest.
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