D2O同位素标记方法，通过单细胞拉曼光谱法在复杂土壤社区中探测可溶解磷酸盐的细菌。,Analytical Chemistry

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D2O同位素标记方法，通过单细胞拉曼光谱法在复杂土壤社区中探测可溶解磷酸盐的细菌。
Analytical Chemistry ( IF 6.7 ) Pub Date : 2019-01-04 00:00:00 , DOI: 10.1021/acs.analchem.8b04820
Hong-Zhe Li _{1,

2} , Qing-Fang Bi _{1,

3} , Kai Yang _{1,

2} , Bang-Xiao Zheng _{1,

2} , Qiang Pu _{1,

2} , Li Cui ₁

Affiliation

通过磷酸盐增溶细菌（PSB）提高土壤中固定磷（P）的生物利用度是有效的可持续农业利用P和缓解P危机的有效策略。在这里，D ₂ O同位素标记与单细胞拉曼光谱法（Raman–D ₂ O）结合被开发为一种有效的基于活性的方法，可以以与培养物无关的方式表征PSB的存在和活性。根据发现，在存在不溶性P的情况下，PSB的活性明显高于非PSB，这是由于D _2的主动同化引起的C–D拉曼谱带O衍生的D被确立为可溶解无机磷酸盐的细菌和可溶解有机磷酸盐的细菌的生物标记。由于C–D比率和C–D比率之间的一致性，进一步确定了C–D比率（C–D频带的强度相对于C–D和C–H频带的强度的百分比）作为P释放活动的半定量指标。通过常规的批量测定法测量的溶解的磷酸或酸性磷酸酶活性的浓度。通过应用拉曼成像，单细胞拉曼– D ₂在混合土壤细菌培养物中，甚至在复杂的土壤群落中，O都能清楚地识别出PSB。在单细胞水平上发现了微生物活性的显着异质性，范围从2％到30％（分别接近无磷培养基和具有足够可溶性磷的培养基）。溶解磷。这项工作不仅能够探测PSB及其P的释放活性，而且在禁止获得相关同位素标记的底物时也为探索更多种微生物资源打开了一个窗口。

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D2O-Isotope-Labeling Approach to Probing Phosphate-Solubilizing Bacteria in Complex Soil Communities by Single-Cell Raman Spectroscopy.

Increasing the bioavailability of immobilized phosphorus (P) in soil by phosphate-solubilizing bacteria (PSB) is an effective strategy for sustainable agronomic use of P and for mitigating the P crisis. Here, D₂O isotope labeling combined with single-cell Raman spectroscopy (Raman–D₂O) was developed as an efficient activity-based approach to characterizing the presence and activity of PSB in a culture-independent way. On the basis of the finding that PSB were significantly more active than non-PSB in the presence of insoluble P, a C–D Raman band from active assimilation of D₂O-derived D was established as a biomarker for both inorganic-phosphate-solubilizing bacteria and organic-phosphate-solubilizing bacteria. C–D ratios (intensities of C–D bands as percentages of the intensities of both the C–D and C–H bands) were further established as semiquantitative indicators of P-releasing activities because of the consistency between the C–D ratio and the concentration of solubilized phosphate or acid phosphatase activity as measured by conventional bulk assays. By applying Raman imaging, single-cell Raman–D₂O clearly discerned PSB in a mixed-soil bacterial culture and even in complex soil communities. Remarkable heterogeneity of microbial activity, ranging from 2 to 30% (close to that in medium without P and that in medium with sufficient soluble P, respectively), was revealed at the single-cell level and clearly illustrated the subpopulation of soil bacteria active in solubilizing P. This work not only enables probing PSB and their P-releasing activities but also opens a window to explore more diverse microbial resources when obtaining related isotope-labeled substrates is prohibitive.

更新日期：2019-01-04

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