Process of Carbonate Precipitation by Deleya halophila

Scanning electron microscopy and X-ray dispersive energy microanalysis were used to investigate the formation of carbonate crystals by Deleya halophila. The formation of calcium carbonate crystals (polymorphous aragonite) by D. halophila is a sequential process that commences with a nucleus formed by the aggregation of a few calcified bacterial cells and the subsequent accumulation of more calcified cells and carbonate, which acts to weld the bacteria together. The process leads to the formation of spherical bioliths measuring approximately 50 μm in diameter. The mechanism of carbonate precipitation by D. halophila under our working conditions represents a process of induced biomineralization.     

Biocalcification by halophilic bacteria for remediation of concrete structures in marine environment

Microbial carbonate precipitation has emerged as a promising technology for remediation and restoration of concrete structures. Deterioration of reinforced concrete structures in marine environments is a major concern due to chloride-induced corrosion. In the current study, halophilic bacteria Exiguobacterium mexicanum was isolated from sea water and tested for biomineralization potential under different salt stress conditions. The growth, urease and carbonic anhydrase production significantly increased under salt stress conditions. Maximum calcium carbonate precipitation was recorded at 5 % NaCl concentration. Application of E. mexicanum on concrete specimens significantly increased the compressive strength (23.5 %) and reduced water absorption about five times under 5 % salt stress conditions compared to control specimens. SEM and XRD analysis of bacterial-treated concrete specimens confirmed the precipitation of calcite. The present study results support the potential of this technology for improving the strength and durability properties of building structures in marine environments.     

Bacillus megaterium mediated mineralization of calcium carbonate as biogenic surface treatment of green building materials

Microbially induced calcium carbonate precipitation is a biomineralization process that has various applications in remediation and restoration of range of building materials. In the present study, calcifying bacteria, Bacillus megaterium SS3 isolated from calcareous soil was applied as biosealant to enhance the durability of low energy, green building materials (soil–cement blocks). This bacterial isolate produced high amounts of urease, carbonic anhydrase, extra polymeric substances and biofilm. The calcium carbonate polymorphs produced by B. megaterium SS3 were analyzed by scanning electron microscopy, confocal laser scanning microscopy, X-ray diffraction and Fourier transmission infra red spectroscopy. These results suggested that calcite is the most predominant carbonate formed by this bacteria followed by vaterite. Application of B. megaterium SS3 as biogenic surface treatment led to 40 % decrease in water absorption, 31 % decrease in porosity and 18 % increase in compressive strength of low energy building materials. From the present investigation, it is clear that surface treatment of building materials by B. megaterium SS3 is very effective and eco friendly way of biodeposition of coherent carbonates that enhances the durability of building materials.     

Conservation of Ornamental Stone by Myxococcus xanthus-Induced Carbonate Biomineralization

Increasing environmental pollution in urban areas has been endangering the survival of carbonate stones in monuments and statuary for many decades. Numerous conservation treatments have been applied for the protection and consolidation of these works of art. Most of them, however, either release dangerous gases during curing or show very little efficacy. Bacterially induced carbonate mineralization has been proposed as a novel and environmentally friendly strategy for the conservation of deteriorated ornamental stone. However, the method appeared to display insufficient consolidation and plugging of pores. Here we report that Myxococcus xanthus-induced calcium carbonate precipitation efficiently protects and consolidates porous ornamental limestone. The newly formed carbonate cements calcite grains by depositing on the walls of the pores without plugging them. Sonication tests demonstrate that these new carbonate crystals are strongly attached to the substratum, mostly due to epitaxial growth on preexisting calcite grains. The new crystals are more stress resistant than the calcite grains of the original stone because they are organic-inorganic composites. Variations in the phosphate concentrations of the culture medium lead to changes in local pH and bacterial productivity. These affect the structure of the new cement and the type of precipitated CaCO₃ polymorph (vaterite or calcite). The manipulation of culture medium composition creates new ways of controlling bacterial biomineralization that in the future could be applied to the conservation of ornamental stone.     

Preliminary study on the correlation between the trace Mn2+ and the calcite polymorph in gallstones containing calcium carbonate from the northeast China via electron spin resonance

Gallstones containing calcium carbonate (GCCC) from the northeast China were analyzed using X-ray diffraction (XRD), infrared spectroscopy (IR), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), and electron spin resonance (ESR). The sextet signal arising from the allowed transitions of the trace Mn²⁺ ions in GCCC was found to be ESR-detectable and strong. The XRD technique revealed the crystal habit of calcite in GCCC. Of the three polymorphs of calcium carbonate, no calcite was present as a solitary crystallization form, accompanied by aragonite or vaterite or both. The sextet ESR signal and the (104) main XRD peak at 2θ = ∼29.4° were employed as two probes to explore the relationship between trace Mn²⁺ and calcite. The Mn content can be considered as an indicator of the amount of calcite in GCCC because of the existence of a correlation between Mn²⁺ and calcite. The correlation between Mn²⁺ and calcite, the relation between the levels of Mn²⁺ and the type of gallstones, the structural preference of Mn²⁺ to the calcite polymorph, and the influence of dietary habits on calcite in calcium carbonate gallstones are discussed.     

厚壳贻贝whirlin类贝壳基质蛋白的重组表达与功能分析

贝壳历来是生物工程和材料学研究的重要对象。贝壳中的贝壳基质蛋白质在贝壳的形成与发育过程中具有重要的调控作用。Whirlin类蛋白质(Whirlin-like protein,WLP)是一种从厚壳贻贝(Mytilus coruscus)中鉴定的新型贝壳基质蛋白质。序列分析结果显示,该蛋白质含有PDZ(postsynaptic density/Discs large/Zonula occludens)结构域,而该结构域对贝壳生物矿化的影响目前尚无报道。为深入了解WLP在贝壳形成中对碳酸钙晶体的影响,在序列分析基础上,采用密码子优化结合原核重组表达,获得其重组表达产物后,开展了重组WLP对碳酸钙晶体形貌及晶型的影响研究,结晶速度抑制以及碳酸钙晶体结合分析。分析结果表明,重组WLP能诱导文石型碳酸钙晶体的形貌和方解石型碳酸钙晶体的晶型发生改变;同时重组WLP对碳酸钙晶体具有结合作用,且能抑制碳酸钙晶体的结晶速度。上述结果表明,WLP对贝壳的形成及发育具有重要影响,并可能在贝壳肌棱柱层的形成中发挥了重要作用。     

Deoxyribonucleic acid relatedness among strains of the moderately halophilic speciesDeleya halophila

The genomic relatedness among 16 strains assigned to the moderately halophilic speciesDeleya halophila and other 20 representative strains of halophilic and nonhalophilic species was estimated by determination of deoxyribonucleic acid (DNA) base composition and by DNA-DNA hybridization studies. The guanine-plus-cytosine (G + C) base contents, determined from the melting temperature of DNAs ofD. halophila strains, were 66.0–68.8 mol %. DNA-DNA homology studies, determined by membrane filter technique, indicate that the 16 strains ofD. halophila comprise a genetically homogeneous group. High homology (70–100%) was obtained between the type strainD. halophila CCM 3662 and the otherD. halophila strains studied; however, very low DNA relatedness was found between the representative strains ofD. halophila and otherDeleya species (13-0%), as well as other moderately halophilic, marine, or nonhalophilic bacteria investigated.     

Production of nanocalcite crystal by a urease producing halophilic strain of <Emphasis Type="Italic">Staphylococcus saprophyticus</Emphasis> and analysis of its properties by XRD and SEM

Cementation of salt-containing soils can be achieved by salt-tolerant or halophilic calcite precipitation bacteria. Therefore, the isolation of calcite-producing bacteria in the presence of salt is the first step in the microbial cementation of saline soils. Urease producing bacteria can cause calcite nano-crystals to precipitate by producing urease in the presence of urea and calcium. The purpose of this study was to isolate urease producing halophilic bacteria in order to make calcite precipitate in saline soil. The calcite and the properties of the strains were further analyzed by X-ray diffraction (XRD) and scanning electron microscope equipped with an energy dispersive X-ray detector. In this study, a total of 110 halophilic strains were isolated, from which 58 isolates proved to have the ability of urease production. Four strains were identified to produce nano-calcite using urease activity in the precipitation medium. The XRD studies showed that the size of these particles was in the range of 40–60 nm. Strain H3 revealed that calcite is mostly produced in the precipitation medium containing 5% salt in comparison with other strains. This strain also produced calcite precipitates in the precipitation medium containing 15% salt. Phylogenetic analysis indicated that these isolates are about 99–100% similar to Staphylococcus saprophyticus.     

Chemical Purity of Shewanella oneidensis-Induced Magnetites

Magnetite is a common iron oxide produced both inorganically and biogenically. Biologically-induced magnetite is often originated, under appropriate conditions, as a result of the Fe³⁺ reduction by dissimilatory iron reducing bacteria, which are usually found in anoxic environments or at the oxic-anoxic interface. Such a Fe³⁺ bioreduction occurs upon this cation acting as an electron acceptor of an anaerobic respiration, thus creating favorable conditions for magnetite precipitation. This biologically-induced magnetite is an important biomineral in the environments inhabited by iron reducing bacteria. The presence of a variety of cations may influence both the biomineralization process and the resulting biomineral, however this phenomenon has not been investigated extensively. In the present study, we study the effect on the magnetite biomineralization process of the presence of calcium, magnesium and manganese in the culture medium where Shewanella oneidensis lives. We also test the incorporation of these cations into the crystalline structure of inorganic and biogenic magnetite induced by S. oneidensis. According to our findings, manganese ions likely become incorporated into the crystal structure of biologically produced magnetites, while magnesium ions are incorporated in inorganic magnetites, and calcium ions are excluded from the crystal structure of both inorganic and biotic magnetites. We hypothesize that the incorporation of cations into magnetite depends not only on the relative cation radii, but also on the mechanisms of magnetite formation.     

Intracellular amorphous Ca-carbonate and magnetite biomineralization by a magnetotactic bacterium affiliated to the Alphaproteobacteria

Bacteria synthesize a wide range of intracellular submicrometer-sized inorganic precipitates of diverse chemical compositions and structures, called biominerals. Their occurrences, functions and ultrastructures are not yet fully described despite great advances in our knowledge of microbial diversity. Here, we report bacteria inhabiting the sediments and water column of the permanently stratified ferruginous Lake Pavin, that have the peculiarity to biomineralize both intracellular magnetic particles and calcium carbonate granules. Based on an ultrastructural characterization using transmission electron microscopy (TEM) and synchrotron-based scanning transmission X-ray microscopy (STXM), we showed that the calcium carbonate granules are amorphous and contained within membrane-delimited vesicles. Single-cell sorting, correlative fluorescent in situ hybridization (FISH), scanning electron microscopy (SEM) and molecular typing of populations inhabiting sediments affiliated these bacteria to a new genus of the Alphaproteobacteria. The partially assembled genome sequence of a representative isolate revealed an atypical structure of the magnetosome gene cluster while geochemical analyses indicate that calcium carbonate production is an active process that costs energy to the cell to maintain an environment suitable for their formation. This discovery further expands the diversity of organisms capable of intracellular Ca-carbonate biomineralization. If the role of such biomineralization is still unclear, cell behaviour suggests that it may participate to cell motility in aquatic habitats as magnetite biomineralization does.Subject terms: Phylogenetics, Biodiversity, Biogeochemistry, Water microbiology     

Microbially Mediated Calcium Carbonate Precipitation: Implications for Interpreting Calcite Precipitation and for Solid-Phase Capture of Inorganic Contaminants

Microbial degradation of urea was investigated as a potential geochemical catalyst for Ca carbonate precipitation and associated solid phase capture of common groundwater contaminants (Sr, UO₂, Cu) in laboratory batch experiments. Bacterial degradation of urea increased pH and promoted Ca carbonate precipitation in both bacterial control and contaminant treatments. Associated solid phase capture of Sr was highly effective, capturing 95% of the 1 mM Sr added within 24 h. The results for Sr are consistent with solid solution formation rather than discrete Sr carbonate phase precipitation. In contrast, UO₂ capture was not as effective, reaching only 30% of the initial 1 mM UO₂ added, and also reversible, dropping to 7% by 24 h. These results likely reflect differing sites of incorporation of these two elements-Ca lattice sites for Sr versus crystal defect sites for UO₂. Cu sequestration was poor, resulting from toxicity of the metal to the bacteria, which arrested urea degradation and concomitant Ca carbonate precipitation. Scanning electron microscopy (SEM) indicated a variety of morphologies reminiscent of those observed in the marine stromatolite literature. In bacterial control treatments, X-ray diffraction (XRD) analyses indicated only calcite; while in the presence of either Sr or UO₂, both calcite and vaterite, a metastable polymorph of Ca carbonate, were identified. Tapping mode atomic force microscopy (AFM) indicated differences in surface microtopography among abiotic, bacterial control, and bacterial contaminant systems. These results indicate that Ca carbonate precipitation induced by passive biomineralization processes is highly effective and may provide a useful bioremediation strategy for Ca carbonate-rich aquifers where Sr contamination issues exist.     

Precipitation of calcium carbonate byDeleya halophila in media containing NaCl as sole salt

The precipitation of calcium carbonate by 27 strains ofDeleya halophila using solid and liquid media containing different NaCl concentrations (2.5, 7.5, or 20%, wt/vol) as sole salt, and two incubation temperatures (22° and 32°C) have been studied. All the strains tested were able to precipitate calcium carbonate under the different environmental conditions assayed. Crystals formed were calcite and vaterite; the ratio of calcite to vaterite was dependent on total salts and on the type of medium.     

Biomineralization of carbonate and phosphate by moderately halophilic bacteria

We investigated the precipitation of carbonate and phosphate minerals by 19 species of moderately halophilic bacteria using media with variable Mg(2+)/Ca(2+) ratios. The precipitated minerals were calcite, magnesium (Mg) calcite, and struvite (MgNH(4)PO(4) x 6H(2)O) in variable proportions depending on the Mg(2+)/Ca(2+) ratio of the medium. The Mg content of the Mg-calcite decreased with increasing Ca(2+) concentration in the medium. According to the saturation indices, other minerals could also have precipitated. We observed important differences between the morphology of carbonate and phosphate, which may help us to recognize these minerals in natural systems. We studied the growth and pH curves of four bacteria in media specific for carbonate and struvite precipitation. We consider the biomineralization processes that produce carbonate and phosphate minerals, and propose a hypothesis for the lack of struvite in natural environments and ancient rocks.     

Carbonate Precipitation of Bacterial Strains Isolated from Sediments and Seawater: Formation Mechanisms

This article presents a research study on carbonate formation in solid and liquid media by Thalassospira sp., Halomonas sp., Bacillus pumilus, and Pseudomonas grimontii, four bacterial strains isolated from sediments and deep seawater. As part of this study, we analyzed carbonic anhydrase activity, pH, adsorption of calcium and magnesium ions, and total organic and inorganic carbon. The geochemical program PHREEQC was also used to calculate the mineral saturation indexes in all the cultures. The minerals formed were studied with X-ray diffraction, X-ray dispersive energy microanalysis, and scanning electron microscopy. In addition, all four bacterial strains were found to induce carbonate precipitation and to have carbonic anhydrase activity. Sterile control experiments did not precipitate carbonate. In solid M1 and B4 media, all of the strains precipitated magnesium calcite, whereas in the liquid media, they precipitated different percentages of magnesium calcite, aragonite, and monohydrocalcite. In both cases, small amounts of amorphous precipitates were also produced. This article discusses carbonate formation and the possible role played by metabolic activity, bacterial surfaces and carbonic anhydrase in this process. Finally, the results obtained lead to a hypothesis regarding the importance of carbonate precipitation for the survival of bacteria populations in certain habitats.     

Biological nitrogen fixation and phosphate solubilization by bacteria isolated from tropical soils

Introduction

In addition to fixing atmospheric nitrogen, some bacterial isolates can also solubilize insoluble phosphates, further contributing to plant growth.

Aims

The objectives of this study were the following: isolate, select, and identify nodulating bacteria in the cowpea that are efficient not only in biological nitrogen fixation (BNF) but also in the solubilization of insoluble inorganic phosphates; identify and quantify the organic acids produced; and establish the relationship between those acids and the solubilizing capacity.

Methods

The bacteria were captured from two soils containing high concentrations of insoluble phosphorus from the cities of Lavras and Patos de Minas, using the cowpea [Vigna unguiculata (L.) Walp.] as bait. We obtained 78 strains, which were characterized according to their cultural attributes in culture medium 79 with the strains UFLA 03-84, INPA 03-11B, and BR3267 (approved by the Ministry of Livestock and Supply Agriculture—MAPA, as inoculants for the cowpea) and Burkholderia cepacia (LMG1222^T), which was used as a positive control for phosphate solubilization. Strains that were selected for their efficiency in both processes were identified by 16S rDNA sequence analysis. We evaluated the symbiotic efficiency (BNF) in a greenhouse and the solubilization efficiency of CaHPO₄, Al(H₂PO₄)₃, and FePO₄.2H₂O in solid and liquid GELP media. Strains that excelled at the solubilization of these phosphate sources were also evaluated for the production of the following organic acids: oxalic, citric, gluconic, lactic, succinic, and propionic.

Results

The presence of Acinetobacter, Bacillus, Firmicutes, Microbacterium, Paenibacillus, and Rhizobium was detected by 16S rDNA sequencing and analysis. Bacterial strains obtained from cowpea nodules varied greatly in the efficiency of their BNF and phosphate solubilization processes, especially in the strains UFLA 03-09, UFLA 03-10, UFLA 03-12, and UFLA 03-13, which were more efficient in both processes. More strains were able to solubilize insoluble inorganic calcium and iron phosphates in liquid medium than in solid medium. The production of organic acids was related to the solubilization of CaHPO₄ and FePO₄.2H₂O for some strains, and the type and concentration of the acid influenced this process.

Conclusions

These are the first results obtained with bacterial isolates from tropical soils in which the production of organic acids was detected and quantified to examine the solubilization of insoluble inorganic phosphates.     

Subscribed Content Calcium Carbonate Precipitation by Ureolytic Subsurface Bacteria

Coprecipitation in carbonate minerals offers a means of slowing the transport of divalent radionuclides and contaminant metals (e.g.,⁹⁰Sr²⁺, UO²⁺, Co²⁺) in the subsurface. It may be possible to accelerate this process by stimulating the native microbial community to generate chemical conditions favoring carbonate precipitation. In a preliminary evaluation of this approach, we investigated the ability of ureolytic subsurface bacteria to produce alkaline conditions conducive to calcium carbonate precipitation. Groundwater samples from the Eastern Snake River Plain (ESRP) aquifer in Idaho were screened for urea-hydrolyzing microorganisms; three isolates were selected for further evaluation. Analysis of 16S rRNA gene sequences indicated that two of the ESRP isolates were of the genus Pseudomonas , and the other was a Variovorax sp. The specific urease activities of the ESRP isolates appeared to be similar to each other but less than that of Bacillus pasteurii , a known urease-positive organism. However, calcium carbonate was rapidly precipitated in all cultures that were supplied with urea and calcium, and X-ray diffraction analyses indicated that calcite was always the predominant carbonate polymorph produced. The correspondence between measured calcium concentrations and equilibrium predictions suggested that the rate of calcite precipitation was directly linked to the rate of urea hydrolysis. These results are promising with respect to the potential utility of this approach for in situ remediation and indicate that further evaluation of this approach under conditions more closely simulating environmental conditions is warranted.     

Fertilization enhancing carbon sequestration as carbonate in arid cropland: assessments of long-term experiments in northern China

Aims

Soil inorganic carbon (SIC), primarily calcium carbonate, is a major reservoir of carbon in arid lands. This study was designed to test the hypothesis that carbonate might be enhanced in arid cropland, in association with soil fertility improvement via organic amendments.

Methods

We obtained two sets (65 each) of archived soil samples collected in the early and late 2000’s from three long-term experiment sites under wheat-corn cropping with various fertilization treatments in northern China. Soil organic (SOC), SIC and their Stable ¹³C compositions were determined over the range 0–100 cm.

Results

All sites showed an overall increase of SIC content in soil profiles over time. Particularly, fertilizations led to large SIC accumulation with a range of 101–202 g C m^?2 y^?1 in the 0–100 cm. Accumulation of pedogenic carbonate under fertilization varied from 60 to 179 g C m^?2 y^?1 in the 0–100 cm. Organic amendments significantly enhanced carbonate accumulation, in particular in the subsoil.

Conclusions

More carbon was sequestrated in the form of carbonate than as SOC in the arid cropland in northern China. Increasing SOC stock through long-term straw incorporation and manure application in the arid and semi-arid regions also enhanced carbonate accumulation in soil profiles.     

The Roles of Mg over the Precipitation of Carbonate and Morphological Formation in the Presence of Arthrobacter sp. Strain MF-2

Microbial mineralization of carbonate is a research subject widely studied in the past decades. The magnesium ions (Mg²⁺), present in water systems, are a key determinant in biomineralization process of carbonate and they are widely found in calcium-based biominerals as an accessory component. However, the crystallization mechanism and morphological change of carbonate polymorphs in the presence of Mg²⁺ ions has not been clarified sufficiently. In this report, a series of culture experiments were performed for 50 days using the Arthrobacter sp. strain MF-2 in a M2 culture medium using Mg/Ca molar ratios (R) of 0, 1.5, 3, 6, 9, and 12 in solution. And the roles of Mg²⁺ ions on the crystal growth and morphological change of biogenic carbonate were investigated. Experimental results show: (1) MF-2 could induce aragonite, high-Mg calcite, and Ca-dolomite formation in M2 culture media with different R values. The increased stability of amorphous calcium carbonate suggests Mg²⁺ ions inhibit carbonate crystallization. (2) The mineral morphologies were varied (rod-shaped, dumbbell-shaped, cauliflower-like, spherical, etc.) in the medium with R = 1.5, but they became simple (spherical and lamellar) in high Mg²⁺ concentrations (Mg > 0.15 M, R > 3). (3) The increased ionic strength of Mg²⁺ ions in the environment has an influence on the polymorphs and morphologies of carbonate formed by controlling the metabolism of strain MF-2 and the activity of carbonic anhydrase.     

A Bivalve Biomineralization Toolbox

Mollusc shells are a result of the deposition of crystalline and amorphous calcite catalyzed by enzymes and shell matrix proteins (SMP). Developing a detailed understanding of bivalve mollusc biomineralization pathways is complicated not only by the multiplicity of shell forms and microstructures in this class, but also by the evolution of associated proteins by domain co-option and domain shuffling. In spite of this, a minimal biomineralization toolbox comprising proteins and protein domains critical for shell production across species has been identified. Using a matched pair design to reduce experimental noise from inter-individual variation, combined with damage-repair experiments and a database of biomineralization SMPs derived from published works, proteins were identified that are likely to be involved in shell calcification. Eighteen new, shared proteins likely to be involved in the processes related to the calcification of shells were identified by the analysis of genes expressed during repair in Crassostrea gigas, Mytilus edulis, and Pecten maximus. Genes involved in ion transport were also identified as potentially involved in calcification either via the maintenance of cell acid–base balance or transport of critical ions to the extrapallial space, the site of shell assembly. These data expand the number of candidate biomineralization proteins in bivalve molluscs for future functional studies and define a minimal functional protein domain set required to produce solid microstructures from soluble calcium carbonate. This is important for understanding molluscan shell evolution, the likely impacts of environmental change on biomineralization processes, materials science, and biomimicry research.