Dolomite Formation Mediated by Halophilic Archaeal Cells under Different Conditions and Carboxylated Microspheres
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摘要: 白云石是沉积岩中广泛存在的碳酸盐矿物，其成因机制一直备受关注.野外调研发现现生白云石多分布于高盐环境，模拟实验也表明嗜盐微生物能诱导形成白云石，但微生物诱导白云石沉淀的机理仍不明确.分别用嗜盐古菌Natrinema sp.J7-1对数后期的活细胞、失去代谢活性的J7-1完整细胞（经线粒体氧化磷酸化解偶联剂处理）、表面蛋白质变性的J7-1细胞（经多聚甲醛和戊二醛处理）和表面富含羧基的微球，在盐度为280‰的沉淀体系中诱导白云石沉淀.分别利用X射线衍射（XRD）分析矿物的物相，扫描电子显微镜（SEM）分析矿物、微生物以及羧基微球的形貌，傅立叶红外光谱（FT-IR）分析细胞变性前后表面的官能团.结果表明，失去代谢活性的J7-1细胞与正常的对数后期细胞均能够诱导原白云石形成；经过多聚甲醛/戊二醛固定后，细胞表面羧基含量降低，不能诱导白云石沉淀；羧基微球能够诱导形成原白云石.以上研究证实细胞表面的羧基可能是微生物促进白云石沉淀的一种关键因素，而细胞的生长代谢在本研究的条件下不是控制白云石沉淀的主要因素.Abstract: Dolomite is a widespread carbonate mineral in sedimentary rocks, and its formation mechanism have always attracted much attention. Most modern dolomite occurred in hypersaline environments and several microbial isolates from these environments have been reported to be able to induce dolomite formation as well. However the detailed mechanism of microbial induced dolomite remains largely unclear to date. In this study, a halophilic archaea, Natrinema sp. J7-1, was used to investigate what exactly played an important role in the microbial dolomite formation. Normal cells of Natrinema sp. J7-1 in post-log phase, inactive cells treated with carbonyl cyanide 3-cholorophenylhydrazine (CCCP, amitochondrial inhibitor), denatured cells treated by paraformaldehyde and glutaraldehyde, as well as carboxylated microspheres were used to induce dolomite formation. The mineral phases were identified by X-ray Diffraction(XRD), the morphologies of minerals, cells and microspheres were observed by Scanning Electron Microscope(SEM) and the functional groups on the surface of normal cells and denatured cells were analyzed by Fourier Transform Infrared Spectroscopy(FT-IR). Results showed that normal cells, inactive cells and carboxylated microspheres induced proto-dolomite formation successfully at the salinity of 280‰. In contrast, denatured cells were not capable of inducing the formation of dolomite, and the percentage of carboxyl groups on their surface decreased compared with normal cells. It can be concluded that carboxyl is one critical factor for microbially mediated dolomite formation, but respiration may count for little under our conditions.
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Key words:
- dolomite /
- halophilic archaea /
- carboxyl /
- microorganism /
- mineralogy
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图 1 不同处理下的Natrinema sp. J7-1细胞及其诱导形成的矿物
a.对数后期的Natrinema sp. J7-1细胞；b.CCCP处理后的Natrinema sp. J7-1细胞；c.经1.5%多聚甲醛和2.5%戊二醛固定后的Natrinema sp. J7-1细胞；d.图a中细胞诱导形成的矿物；e.图b中细胞诱导形成的矿物；f.图c中细胞在沉淀体系中形成的矿物
Fig. 1. The Natrinema sp. J7-1 cells with different treatments and the precipitates they induced
图 2 羧基微球及其诱导形成的矿物
a.0.82 μm羧基微球；b.浓度为108个/ mL的羧基微球诱导形成的矿物
Fig. 2. Morphologies of carboxylated microspheres and the minerals they induced
图 3 不同处理后的Natrinema sp. J7-1细胞及羧基微球诱导沉淀的矿物XRD图谱
a.对数生长后期的正常细胞；b.CCCP处理后的失活细胞；c.经1.5%多聚甲醛和2.5%戊二醛固定后的变性细胞；d.羧基微球；e.空白对照
Fig. 3. Mineral XRD patterns of the precipitates induced by Natrinema sp. J7-1 with different treatments and carboxylated microspheres
图 4 正常Natrinema sp. J7-1细胞和经固定处理后Natrinema sp. J7-1细胞的FT-IR图谱
Fig. 4. FT-IR spectra of normal cells and fixed cells of Natrinema sp. J7-1
表 1 不同处理的样品的ATP测量值
Table 1. ATP values of samples with different treatments
样品 ATP值(RLU's) A B C 平均值 无菌水 0 0 0 0 280‰洗液 0 0 0 0 细胞培养液 686 604 624 638 细胞重悬液 13 780 14 516 16 067 14 788 CCCP处理的细胞重悬液 0 0 0 0 
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表 2 细胞固定前后FT-IR特征峰红外波长及其对应的官能团
Table 2. The infrared wave lengths of the typical peaks on the FTIR spectra of normal cells and fixed cells and the corresponding functional groups
吸收带 波数(cm-1) 官能团 1 3 500~3 400 酰胺类化合物中的N-H伸缩振动(Benning et al., 2004) 2 ~1 638 酰胺Ⅰ峰的C＝O伸缩振动(Yee et al., 2004；Tourney et al., 2008) 3 ~1 554 酰胺Ⅱ峰的N-H变形振动(Yee et al., 2004；Tourney et al., 2008) 4 ~1 400 羧基官能团中的C＝O对称伸缩振动(Yee et al., 2004；Dittrich and Sibler, 2005；Heinrich et al., 2007；Leone et al., 2007) 5 ~1 242 磷酰基中的P＝O非对称伸缩振动(Dittrich and Sibler, 2005) 6 1 200~900 多糖和脂类中的C-O-C，C-O-P，P-O-P振动(Dittrich and Sibler, 2005；Hadjoudja et al., 2010；Liu et al., 2015) 7 ~1 100 磷酰基中的P＝O非对称伸缩振动和多糖中的C-OH伸缩振动(Jiang et al., 2004；Dittrich and Sibler, 2005)
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