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The discovery of cyanobacteria fossils in microbialite prompts the investigation of carbonate biomineralization using cyanobacteria. However, the impact of coexisting magnesium and iron in microbialite on carbonate biomineralization has been overlooked. Here, Synechocystis sp. PCC 6803 was used to induce calcium carbonate in the presence of coexisting magnesium and ferric ions. The findings demonstrate that cell concentration, pH, carbonic anhydrase activity, and carbonate and bicarbonate concentrations decreased with increasing concentrations of magnesium and calcium ions. Ferric ions yielded a contrasting effect. The levels of deoxyribonucleic acid, protein, polysaccharides, and humic substances in extracellular polymeric substances increased in the presence of separated or coexisting calcium, magnesium, and ferric ions. Magnesium ions inhibited calcium ion precipitation, whereas ferric ions exhibited the opposite effect. Protein secondary structures became more abundant and O-C=O and N-C=O contents increased with increasing ion concentrations by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy analyses. Scanning electron microscopy revealed that ferric ions lead to rougher surfaces and incomplete rhombohedral structures of calcite, whereas magnesium ions promoted greater diversity in morphology. Magnesium ions enhanced the incorporation of ferric ions. This work aims to further understand the effect of magnesium and ferric ions on calcium carbonate biomineralization induced by cyanobacteria. [ABSTRACT FROM AUTHOR]

In this study, a facultative anaerobic strain isolated from marine sediments and identified as Citrobacter freundii, was used to induce the precipitation of carbonate and phosphate minerals in the laboratory under anaerobic conditions. This is the first time that the ability of C. freundii ZW123 to precipitate carbonate and phosphate minerals has been demonstrated. During the experiments, carbonic anhydrase, alkaline phosphatase and ammonium released by the bacteria not only promoted an increase in pH, but also drove the supersaturation and precipitation of carbonate and phosphate minerals. The predominant bio-mediated minerals precipitated at various Mg/Ca molar ratios were calcite, vaterite, Mg-rich calcite, monohydrocalcite and struvite. A preferred orientation towards struvite was observed. Scanning transmission electron microscopy (STEM) and elemental mapping showed the distribution of magnesium and calcium elements within Mg-rich calcite. Many organic functional groups, including C=O, C-O-C and C-O, were detected within the biominerals, and these functional groups were also identified in the associated extracellular polymeric substances (EPS). Fifteen kinds of amino acid were detected in the biotic minerals, almost identical to those of the EPS, indicating a close relationship between EPS and biominerals. Most amino acids are negatively charged and able to adsorb cations, providing an oversaturated microenvironment to facilitate mineral nucleation. The X-ray photoelectron spectroscopy (XPS) spectrum of struvite shows the presence of organic functional groups on the mineral surface, suggesting a role of the microorganism in struvite precipitation. The ZW123 bacteria provided carbon and nitrogen for the formation of the biotic minerals through their metabolism, which further emphasizes the close relationship between biominerals and the microorganisms. Thermal studies showed the enhanced thermal stability of biotic minerals, perhaps due to the participation of the bacteria ZW123. The presence of amino acids such as Asp and Glu may explain the high magnesium content of some calcites. Molecular dynamics simulations demonstrated that the morphological change and preferred orientation were likely caused by selective adsorption of EPS onto the various struvite crystal surfaces. Thus, this study shows the significant role played by C. freundii ZW123 in the bioprecipitation of carbonate and phosphate minerals and provides some insights into the processes involved. [ABSTRACT FROM AUTHOR]

Although biomineralization of minerals induced by microorganisms has been widely reported, the mechanisms of biomineralization and the characteristics of the biominerals precipitated needs to be studied further. In this study, Staphylococcus warneri YXY2, a moderate halophile, was used to induce the precipitation of carbonate minerals at various Mg/Ca molar ratios. To investigate the biomineralization mechanism, the growth curve, pH changes, ammonia test, the concentration of bicarbonate and carbonate ions, and the activity of carbonic anhydrase (CA) and alkaline phosphatase (ALP) were determined. X-ray powder diffraction (XRD), scanning electron microscopy - energy disperse spectroscopy (SEM-EDS), Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), scanning transmission electron microscopy (STEM), and stable carbon isotope analyses were used to characterize the minerals. The obtained biotic minerals were calcite, vaterite, Mg-rich calcite, and aragonite crystals. The crystallinity of aragonite decreased with increasing Mg/Ca ratios. The preferred orientation, diverse morphologies, organic substances, and more negative stable carbon isotope values proved the biogenesis of these carbonate minerals. The presence of Mg in the biotic aragonite crystals was likely related to the acidic amino acids which also facilitated the nucleation of minerals on/in the extracellular polymeric substances (EPS). Mg2+ and Ca2+ ions were able to enter into the YXY2 bacteria to induce intracellular biomineralization. Dynamics simulation using Material Studio software proved that different adsorption energies of Glutamic acid (Glu) adsorbed onto different crystal planes of aragonite led to the preferred orientation of aragonite. This study helps to deepen our understanding of biomineralization mechanisms and may be helpful to distinguish biotic minerals from abiotic minerals. [ABSTRACT FROM AUTHOR]

The halophilic bacterium Halomonas smyrnensis from a modern salt lake used in experiments to induce biomineralization has resulted in the precipitation of monohydrocalcite and other carbonate minerals. In this study, a Halomonas smyrnensis WMS-3 (GenBank:MH425323) strain was identified based on 16S rDNA homology comparison, and then cultured in mediums with 3% NaCl concentration to induce monohydrocalcite at different Mg/Ca molar ratios of 0, 2, 5, 7, and 9. The growth curve of WMS-3 bacteria, pH values, NH4+ concentration, HCO3− and CO32− concentration, carbonic anhydrase (CA) activity, and the changes in Ca2+ and Mg2+ ion concentration were determined to further explore the extracellular biomineralization mechanism. Moreover, the nucleation mechanism of monohydrocalcite on extracellular polymeric substances (EPS) was analyzed through studying ultrathin slices of the WMS-3 strain by High resolution transmission electron microscopy (HRTEM), Selected area election diffraction (SAED), Scanning transmission electron microscopy (STEM), and elemental mapping, besides this, amino acids in the EPS were also analyzed. The results show that pH increased to about 9.0 under the influence of ammonia and CA activity. The precipitation ratio (%, the ratio of the mass/volume concentration) of the Ca2+ ion was 64.32%, 62.20%, 60.22%, 59.57%, and 54.42% at Mg/Ca molar ratios of 0, 2, 5, 7, and 9, respectively, on the 21st day of the experiments, and 6.69%, 7.10%, 7.74%, 8.09% for the Mg2+ ion concentration at Mg/Ca molar ratios 2, 5, 7, and 9, respectively. The obtained minerals were calcite, Mg-rich calcite, aragonite, and hydromagnesite, in addition to the monohydrocalcite, as identified by X-ray diffraction (XRD) analyses. Monohydrocalcite had higher crystallinity when the Mg/Ca ratio increased from 7 to 9; thus, the stability of monohydrocalcite increased, also proven by the thermogravimetry (TG), derivative thermogravimetry (DTG) and differential scanning calorimetry (DSC) analyses. The C=O and C–O–C organic functional groups present in/on the minerals analyzed by Fourier transform infrared spectroscopy (FTIR), the various morphologies and the existence of P and S determined by scanning electron microscope-energy dispersive spectrometer (SEM-EDS), the relatively more negative stable carbon isotope values (−16.91‰ to −17.91‰) analyzed by a carbon isotope laser spectrometer, plus the typical surface chemistry by XPS, all support the biogenesis of these mineral precipitates. Moreover, Ca2+ ions were able to enter the bacterial cell to induce intracellular biomineralization. This study is useful to understand the mechanism of biomineralization further and may provide theoretical reference concerning the formation of monohydrocalcite in nature. [ABSTRACT FROM AUTHOR]

Removal of calcium and magnesium ions through biomineralization induced by bacteria has been proven to be an effective and environmentally friendly method to improve water quality, but the process and mechanism are far from fully understood. In this study, a newly isolated probiotic Bacillus licheniformis SRB2 (GenBank: KM884945.1) was used to induce the bio-precipitation of calcium and magnesium at various Mg/Ca molar ratios (0, 6, 8, 10, and 12) in medium with 30 g L−1 sodium chloride. Due to the increasing pH and HCO3− and CO32− concentrations caused by NH3 and carbonic anhydrase, about 98% Ca2+ and 50% Mg2+ were precipitated in 12 days. The pathways of bio-precipitation include extracellular and intracellular processes. Biominerals with more negative δ13C values (−16‰ to −18‰) were formed including calcite, vaterite, monohydrocalcite, and nesquehonite with preferred orientation. The nucleation on extracellular polymeric substances was controlled by the negatively charged amino acids and organic functional groups. The intracellular amorphous inclusions containing calcium and magnesium also contributed to the bio-precipitation. This study reveals the process and mechanism of microbial desalination for the removal of calcium and magnesium, and provides some references to explain the formation of the nesquehonite and other carbonate minerals in a natural and ancient earth surface environment. [ABSTRACT FROM AUTHOR]

Biomineralization induced by microorganisms has become a hot spot in the field of carbonate sedimentology; however, the mechanisms involved still need to be explored. In this study, the bacterium Bacillus subtilis J2 (GenBank MG575432) was used to induce the precipitation of calcium carbonate minerals at Mg/Ca molar ratios of 0, 3, 6, 9, and 12. Bacillus subtilis J2 bacteria released ammonia to increase pH, but the ammonia released only made the pH increase to 8.25. Carbonic anhydrase was also produced to catalyze the hydration of carbon dioxide, and this process released carbonate and bicarbonate ions that not only increased pH but also elevated carbonate supersaturation. The biominerals formed at a Mg/Ca molar ratio of 0 were spherulitic, elongated, dumbbell-shaped, and irregularly rhombohedral calcite; at a Mg/Ca molar ratio of 3, the biominerals were calcite and aragonite, the weight ratio of calcite decreased from 26.7% to 15.6%, and that of aragonite increased from 73.3% to 84.4% with increasing incubation time. At higher Mg/Ca molar ratios, the biominerals were aragonite, and the crystallinity and thermal stability of aragonite decreased with increasing Mg/Ca molar ratios. FTIR results showed that many organic functional groups were present on/within the biominerals, such as C–O–C, N–H, C=O, O–H, and C–H. HRTEM-SAED examination of the ultra-thin slices of B. subtilis J2 bacteria showed that nano-sized minerals with poor crystal structure had grown or been adsorbed on the EPS coating. The EPS of the B. subtilis J2 strain contained abundant glutamic acid and aspartic acid, which could be deprotonated in an alkaline condition to adsorb Ca2+ and Mg2+ ions; this made EPS act as the nucleation sites. This study may provide some references for further understanding of the mechanism of biomineralization induced by microorganisms. [ABSTRACT FROM AUTHOR]

Carbonate precipitation induced by microorganisms has become a hot topic in the field of carbonate sedimentology, although the effects of magnesium on biomineral formation have rarely been studied. In experiments described here, magnesium sulfate and magnesium chloride were used to investigate the significant role played by Mg2+ on carbonate precipitation. In this study, Staphylococcus epidermidis Y2 was isolated and identified by 16S ribosomal DNA (rDNA) homology comparison and ammonia, pH, carbonic anhydrase, carbonate, and bicarbonate ions were monitored during laboratory experiments. The mineral phase, morphology, and elemental composition of precipitates were analyzed by XRD and SEM-EDS. Ultrathin slices of bacteria were analyzed by HRTEM-SAED and STEM. The results show that this bacterium releases ammonia and carbonic anhydrase to increase pH, and raise supersaturation via the large number of carbonate and bicarbonate ions that are released through carbon dioxide hydration catalyzed by carbonic anhydrase. The crystal cell density of monohydrocalcite is lower in a magnesium chloride medium, compared to one of magnesium sulfate. Crystals grow in the mode of a spiral staircase in a magnesium sulfate medium, but in a concentric circular pattern in a magnesium chloride medium. There was no obvious intracellular biomineralization taking place. The results presented here contribute to our understanding of the mechanisms of biomineralization, and to the role of Mg2+ in crystal form. [ABSTRACT FROM AUTHOR]

Biomineralization has become a research hotspot and attracted widespread attention in the field of carbonate sedimentology. In this study, precipitation of carbonate minerals was induced by Bacillus licheniformis DB1-9 bacteria, (identity confirmed with its phylogenetic tree), to further explore the biomineralization mechanisms. During experiments, lasting up to 24 days with varying Mg/Ca molar ratios and regular monitoring of conditions, ammonia and carbonic anhydrase are released by the bacteria, resulting in a pH increase. Carbonic anhydrase could have promoted carbon dioxide hydration to produce bicarbonate and carbonate ions, and so promoted supersaturation to facilitate the precipitation of carbonate minerals. These include rhombohedral, dumbbell-shaped, and elongated calcite crystals; aragonite appears in the form of mineral aggregates. In addition, spheroidal and fusiform minerals are precipitated. FTIR results show there are organic functional groups, such as C–O–C and C=O, as well as the characteristic peaks of calcite and aragonite; these indicate that there is a close relationship between the bacteria and the minerals. Ultrathin slices of the bacteria analyzed by HRTEM, SAED, EDS, and STEM show that precipitate within the extracellular polymeric substances (EPS) has a poor crystal structure, and intracellular granular areas have no crystal structure. Fluorescence intensity and STEM results show that calcium ions can be transported from the outside to the inside of the cells. This study provides further insights to our understanding of biomineralization mechanisms induced by microorganisms. [ABSTRACT FROM AUTHOR]