Microbial Calcium Carbonate Precipitation Research Articles

Microbial Precipitation of Calcium Carbonate for Crack Healing and Stabilization of Sandy Soils

Microbially induced calcium carbonate (CaCO3) precipitation (MICP) can improve the shear strength of soil via biocementation while reducing its porosity and hydraulic conductivity. The purpose of this study was to evaluate the effect of the addition of bacterial metabolites and montmorillonite on the crack healing and biocementation of sandy soil during the MICP process. Cracks were generated by drying wet soil samples in Petri dishes, after which they were sprayed with one of four treatments: deionized water, a cementation solution, bacteria mixed with the cementation solution, and bacterial metabolites mixed with the cementation solution. After five cycles of this spray treatment, the surface crack ratio was observed to decrease by about 71% when living cells were used and by about 80% when microbial metabolites were added. However, the crack reduction ratio was relatively low when treated with water (28%) and the cementation solution alone (48%). To investigate the effect of adding a phyllosilicate to improve the strength of sandy soil, MICP was induced in sand mixed with 0–30% montmorillonite (MMT). As a result, the soil strength increased with higher levels of MMT, indicating that MMT contributed to soil stabilization as a colloid for CaCO3 precipitation and via adhesion between sand grains. Therefore, for the crack healing and stabilization of sandy soil, the addition of bacterial metabolites and montmorillonite may enhance the effectiveness of the MICP process.

Experimental study on alkali reduction of film-coated porous ecological concrete by microbial calcium carbonate precipitation technology

Some plants do not grow due to the high pH levels of ecological concrete pore solutions. In this paper, we design and build an integrated device featuring a combined microbial film and a transverse/U-shaped grouting film. We have applied to the China Intellectual Property Office for an invention patent on this device. The device overcomes the blockage of the grouting port caused by microbial and vertical grouting. The vertical grouting tube leaves holes inside a specimen, reducing the compressive strength, while the integrated device optimizes and decreases variation in the recycling of microbial bacteria. The reduction in the pore alkalinity of porous ecological concrete resulting from the microbial grouting film of this device is larger than that resulting from a microbial sprayed film. The pH values of porous ecological concrete with microbial grouting films and microbial sprayed films are obtained by the pure slurry soaking method and solid–liquid extraction method, respectively. The pH value is lower for the film obtained by the pure slurry soaking method than for that obtained by the solid–liquid extraction method. Conversely, the pH value of porous ecological concrete with a microbial grouting film is reduced to approximately 8 at an age of 56 days. The compressive strengths of the porous ecological concrete specimens with the two films are almost the same. The results of this study provide the necessary theoretical basis for developing alkali reduction technology for porous ecological concrete with environmental and economic benefits.

Applications of microbial calcium carbonate precipitation in concrete through denitrification: a review

Applications of microbial calcium carbonate precipitation in concrete through denitrification: a review

Study of Concrete Crack Repair using Bacillus megaterium

The low yield of microbial calcium carbonate precipitation due to the strong pH environment has limited the development of biotechnology-based concrete crack repair techniques. In this study, Bacillus megaterium was selected to determine the optimal nutrient formulation through orthogonal tests. The growth and reproduction characteristics and urease activity of this strain were analysed by controlling different pH values, and the calcium carbonate precipitation yield under different pH conditions was investigated, and the lower precipitation yield under strong alkaline conditions was improved by using alkaline environment domestication with the addition of urea to the nutrient solution. It has been shown that higher pH results in lower growth and multiplication of Bacillus megaterium and weaker urease activity, and a strong alkaline environment can significantly suppress its growth and multiplication and urease activity. At pH 7, the growth and multiplication of Bacillus megaterium were the fastest, the urease activity was the strongest, and the precipitation yield was greater. The addition of urea to the nutrient solution to domesticate Bacillus megaterium in an alkaline environment can significantly increase the growth rate and precipitation yield, which can effectively solve the problem of insufficient calcium carbonate precipitation under alkaline conditions. Comparing the effectiveness of microbially induced calcium carbonate precipitation (MICP) in repairing concrete after damage before and after microbial domestication shows that the domesticated Bacillus megaterium is more effective in reducing the permeability characteristics of concrete and improving its strength.

Density-dependent microbial calcium carbonate precipitation by drinking water bacteria via amino acid metabolism and biosorption

Drinking water plumbing systems appear to be a unique environment for microorganisms as they contain few nutrients but a high mineral concentration. Interactions between mineral content and bacteria, such as microbial calcium carbonate precipitation (MCP) however, has not yet attracted too much attention in drinking water sector. This study aims to carefully examine MCP behavior of two drinking water bacteria species, which may potentially link scaling and biofouling processes in drinking water distribution systems. Evidence from cell density evolution, chemical parameters, and microscopy suggest that drinking water isolates can mediate CaCO3 precipitation through previously overlooked MCP mechanisms like ammonification or biosorption. The results also illustrate the active control of bacteria on the MCP process, as the calcium starts to concentrate onto cell surfaces only after reaching a certain cell density, even though the cell surfaces are shown to be the ideal location for the CaCO3 nucleation.

Effect of Hydrocarbon Contamination on Biostabilization of Soil Contaminated with Motor Oil and Gasoline

Environmental problems caused by soil contamination can cause changes in the physical, chemical and geotechnical properties of the soil. Research on a suitable method for soil improvement has focused on the use of biological methods for soil improvement and remediation. Microbial calcium carbonate precipitation has been applied to solve many geotechnical problems. In this study, sandy soil was mixed with two conventional hydrocarbon pollutants and classic soil mechanics tests (compaction, direct shear, uniaxial and permeability tests) were implemented. The use of two bacterial solutions, two-phase injection and bacterial flocculation with injection of cementation solution, produced favorable results. Considering the lower inhibition of gasoline in low dilution, it was possible to improve with both methods. In the case of soil contaminated with motor oil, limitations on the use of microbial calcium carbonate precipitation were improved with the use of bacterial flocculation. The results of this approach were more suitable because the bacteria stabilized between the soil grains. XRD, SEM, EDS and wet chemical analysis were carried out to help confirm and interpret the results.

Microbial precipitation of calcium carbonate in cementitious materials – A critical review

The article reviews the application of soil living bacteria in concrete. Formation of cracks and subsequent degradation in concrete are the critical durability phenomenon. For protection against cracking, the repair and regular maintenance by traditional methods are found to be costly. Taking notes from biomimicry and biotechnology in concrete from various literature, the role of bacteria on self-healing of cracks and the indirect benefits to strength and durability property of concrete were consolidated and presented. Amid the different microbial species investigated by researchers, bacillus sphaericus have shown greater precipitation of calcium carbonate (CaCO3) with the cell concentration of 105 cells/ml. The hydrolysis of urea has shown a significant result in the precipitation of CaCO3. Thus, the conjunction of bacillus sphaericus with urea hydrolysis could be an optimum way for biomineralization of CaCO3. The shell life of microbes can be extended by encapsulation into the carrier materials (Eg: expanded clay, perlite etc). The large-scale production of bacterial concrete requires sophisticated laboratory for culturing of microbes and preparation of media. Due to these technical constraints, the application of novel bacterial concrete is at the laboratory level.

Insight into the Properties of Surface Percolated Biocemented Sand

Most traditional soil improvement methods are time-consuming, expensive, require heavy machinery and environmentally detrimental. As a more ecofriendly ground improvement method, the biocementation of soil offers an alternative to traditional soil improvement techniques. This method is based on microbial precipitation of calcium carbonate serving as cementing or bonding agents between soil grains. To achieve this bonding, the method of in situ soil stabilization by surface percolation under free drainage of solutions containing either bacteria or calcium ions was chosen. This paper investigates the effects of this method on the strength improvement and microstructure of a dry loose poorly-graded sand after bioimprovement. To evaluate the success of treatment, a series of laboratory experiments was conducted, including, shear wave velocity, unconfined compressive strength, calcium carbonate content, triaxial shear test, scanning electron microscopy, etc. The study revealed that this newly emerging biomediated technique causes the improvement of soil strength as a result of the cementation of sand particles via crystal precipitation and the surface percolation method has the potential of stabilizing the loose sand with desirable depth. Results also showed that strength increase depends not only on calcium carbonate content but also on its precipitation location and particle-to-particle binding strength and numbers. In addition, scanning electron microscopy images and X-ray diffraction showed precipitation of biological calcium carbonate is different in polymorph type throughout the sand column.

Field detection of urease and carbonic anhydrase activity using rapid and economical tests to assess microbially induced carbonate precipitation.

Microbial precipitation of calcium carbonate is a widespread environmental phenomenon that has diverse engineering applications, from building and soil restoration to carbon sequestration. Urease-mediated ureolysis and CO2 (de)hydration by carbonic anhydrase (CA) are known for their potential to precipitate carbonate minerals, yet many environmental microbial community studies rely on marker gene or metagenomic approaches that are unable to determine in situ activity. Here, we developed fast and cost-effective tests for the field detection of urease and CA activity using pH-sensitive strips inside microcentrifuge tubes that change colour in response to the reaction products of urease (NH3 ) and CA (CO2 ). The urease assay proved sensitive and useful in the field to detect in situ activity in biofilms from a saline lake, a series of calcareous fens, and ferrous springs, finding relatively high urease activity in lake samples. Incubations of lake microbes with urea resulted in significantly higher CaCO3 precipitation compared to incubations with a urease inhibitor, showing that the rapid assay indicated an on-site active metabolism potentially mediating carbonate precipitation. The CA assay, however, showed less sensitivity compared to the urease test. While its sensitivity limits its utility, the assay may still be useful as a preliminary indicator given the paucity of other means for detecting CA activity in the field. Field urease, and potentially CA, activity assays complement molecular approaches and facilitate the search for carbonate-precipitating microbes and their in situ activity, which could be applied toward agriculture, engineering and carbon sequestration technologies.

The Influence of Microbiology on Soil Aggregation Stability

In the present study, this work investigated the effects of bacteria (Bacillus sp.) decay and biological aggregating factors (organic matter, microbial calcium carbonate precipitation) at various steps on soil aggregate stability. Experiments carried out at different cultivation time 2, 5, and 8 weeks. The soil samples were compared the control (pure soil) versus soil treated with bacteria. The results showed that bacterial activity played an essential function in the macro-aggregate foundation. Nevertheless, the influence of organic matter (OM) on soil aggregate stability was important, which might be related to the microbial activity of bacteria to decay the organic material to an organic mineral that is connected with soil particles to form stable micro-aggregates. The CaCO3 is one of the most stable bonding agencies in nature as it is called natural cement, The soil aggregates stability increased as calcium carbonate increased. No change of soil pH levels observed due to microbial reactions with soil. Hence, the appropriate pH was 7.5. In addition, the higher pH decreases enzymatic activity and carbonate tends to dissolve at pH levels are very low. This study affords the helpful of soil stabilization in the presence of natural cementation of bacteria in contrast with the previous studies.

Induction of microbial carbonate precipitation as a sustainable strategy for post-improvement and remediation of cold mix asphalt

Cold Mix Asphalt (CMA) is highly susceptible to moisture damage, manifesting itself in stripping, raveling, rutting, and cracking faults. This study aimed to tackle this concern with bio-treatment. In doing so, this effort investigates the efficacy of microbial calcium carbonate precipitation in (i) the post-bio-strengthening (PBS) of mechanical properties and (ii) bio-remediation (BR) of moisture damage in cold-mix asphalt emulsion mixtures. To study the post-bio-strengthening potential, a set of cold-mix asphalt specimens was sprayed and injected with bacterial culture medium and nutrient solution to induce calcium carbonate precipitation for the purpose of post-strengthening. Then, the effect of curing times of 7, 14, 28, and 56 days on the fixation of precipitates in the mixtures was examined. To study the bio-remediation potential, another set of cold-mix asphalt specimens was subjected to a freeze-thaw cycle (modified Lottman test) intended to cause moisture damage and bacterial culture medium and nutrient solution were injected, afterward. For these specimens, the appropriate curing time of the fixation process for the bio-remediation was determined. All specimens were subjected to the destructive and non-destructive tests of mechanical properties, semi-circular bending (SCB), moisture susceptibility and also Cantabro test to assess indirect tensile strength (ITS), Tensile strength ratio, Indirect tensile fracture energy ratio, percent of toughness loss, crack propagation by SCB fracture energy, ultrasonic pulse velocity ratio, electrical resistivity, as well as the abrasion resistance. The Analysis of Variance (ANOVA) of the results was performed at 95% confidence level. A Field Emission Scanning Electron Microscope (FESEM) examination was also performed to study the microstructure and precipitates formed in the mixtures. The results showed that the post-bio-strengthened specimens had significantly better mechanical and resistance to moisture damage properties than those of control. The moisture-damaged specimens that underwent the bio-remediation also showed significant improvements in mechanical properties, and in some cases were similar to the undamaged control specimens. The results indicate that microbial calcium carbonate precipitation (MCP) can serve as an effective and sustainable method for both post-strengthening and remediation of cold mix asphalt mixtures.

Seepage control in sand using bioslurry

Abstract This paper presents a new method for seepage control in sand using bioslurry, a suspension of CaCO3 crystals formed using a microbial calcium carbonate precipitation process (MICP). The bioslurry can be permeated through sand or deposit on top of a sand layer. The bioslurry contains high urease active bacteria cells and allows further MICP treatment when introducing cementation solution. In this way, the permeability of the bioslurry layer or bioslurry permeated sand layer could be further reduced to the order of 10−9 m/s through another 2–3 numbers of MICP treatment using a cementation solution with an optimum concentration of 1.6 M. Such a low permeability is hardly achievable using the conventional MICP method which would require many more numbers of treatment to reduce the permeability of sand to be below 10−7 m/s. Therefore, the proposed bioslurry method is not only more effective, but also more efficient. The water barrier layer formed using the proposed method is much less affected by wet and dry or temperature change cycles than compacted clay liners. It also allow cracks in the water barrier layer to be repaired if required.

An environmentally friendly soil improvement technology for sand and dust storms control

Background: Dust storms occur when unchecked, strong, or turbulent winds combine with exposed loose and dried soil surfaces. Sand and dust storms have a significant impact on society, economy, and environment at local, regional, and global levels. The environmental and health hazards of such storms cannot be permanently reduced, however, by taking appropriate measures, its impact can be reduced. The present study aimed to investigate the effects of microbial precipitation of calcium carbonate (CaCO3) as a biocompatible agent on soil stabilization and control of dust storms using ureaseproducing bacteria (UPB) as a biological improvement technique, which were isolated, identified, sprayed on the soil surface. Methods: For this purpose, the erosion of bio-cemented soil samples was investigated experimentally in a wind tunnel under the condition of wind velocity of 0 to 98 km.h-1 in two soil types with sandy and silty texture in a completely randomized design with three replicates. Results: The investigation of the threshold wind velocity of soil particles showed that soil particles began to move at velocity of 8 and 10 km.h-1 in silty and sandy soils, respectively, but in all biological samples (MICP), particles did not move until the wind speed reached 97 km.h-1. It was also revealed that the weight loss of all MICP-treated samples at different wind velocities was significantly reduced compared to the control group. Differences in the amount of soil loss among bio-cemented samples and control treatments were even superior at higher velocities, so that at velocities more than 57 km.h-1, soil losses increased significantly in the control group, while in soils treated with bacteria, soil loss was very low (about 2.5 kg.m-2.h-1). Comparison of the bacteria used in this study also showed that Bacillus infantis and Paenibacillus sp3 had high efficiency in controlling dust storms. Conclusion: The formation of abrasion-resistant surface layers on soil samples treated by biocementation showed that cementation by biological methods could be an effective way to stabilize surface particles and control sand and dust storms.

Microbial calcium carbonate precipitation with high affinity to fill the concrete pore space: nanobiotechnological approach.

Despite the advantages of concrete, it has a pore structure and is susceptible to cracking. The initiated cracks as well as pores and their connectivity accelerate the structure degradation by permitting aggressive substances to flow into the concrete matrix. This phenomenon results in a considerable repair and maintenance costs and decreases the concrete lifespan. In recent years, biotechnological approach through immobilization of bacteria in/or protective vehicles has emerged as a viable solution to address this issue. However, the addition of macro- or micro scale size particles can decrease the integrity of matrix. In this study, the immobilization of bacteria with magnetic iron oxide nanoparticle (ION) was proposed to protect the bacterial cell and evaluate their effect on healing the concrete pore space. The results show that the addition of immobilized bacteria with IONs resulted in a lower water absorption and volume of permeable pore space. Crystal analysis using scanning electron microscope (SEM) and energy dispersive X-ray spectroscopy (EDS) revealed that CaCO3 was precipitated in bio-concrete specimen as a result of microbial biosynthesis.

Utilization of carbon dioxide as an alternative to urea in biocementation

Ureolytic microbial calcium carbonate precipitation technology has recently emerged as a sustainable, eco-friendly approach for protection and remediation of building materials. However, use of urea has some disadvantages such as production of ammonia, which has raised environmental concerns, as well as risks of damage to concrete materials. In the present investigation, urea is replaced with direct influx of carbon dioxide and studied the precipitation of carbonates by bacteria (Bacillus megaterium SS3). The bacteria is able to grow well and precipitate carbonates well when CO2 influx of 99.5% pure passed (20ml/min) into the flasks for 60s. The amount of CaCO3 precipitated through CO2 influx is comparable with that of 2% urea used for precipitation. The percentage improvement in strength with respect to the control specimens by accelerated carbonation curing was 117% while it was 47% in urea treated specimens. Sorptivity was significantly reduced in accelerated carbonation specimens treated with bacteria compared to other treatments. Though carbonation increased compressive strength in control samples, but bacterial treatment significantly reduced the water absorption in concrete specimens. Present study results suggested that utilization of CO2 serves as an alternative to urea during CaCO3 precipitation.

Decrease in Hydraulic Conductivity of a Paddy Field using Biocalcification in situ

ABSTRACTThe hydraulic conductivity of a paddy field (Anthraquic Dystrustept), a silty clay soil containing more than 29% (w/w) of gravel, in Nagoya University Farm was reduced by in situ treatment of subsurface soil using bentonite and biocalcification (microbial calcium carbonate precipitation) through the addition of CaCl2, urea, and corn steep liquor (CSL). The treatment decreased the hydraulic conductivity of the field from an average of 10−3 cm/s to a range of 10−5 to 10−7 cm/s during 69 days, with reducing the proportion of pores of subsurface soil larger than 75 µm in diameter. The biocalcification effect was observed at 10-cm thickness from the treated subsurface. Laboratory soil core experiments demonstrated that the decrease in the hydraulic conductivity was not attributed to the effect of bentonite but mainly to the effect of biocalcification. The addition of CSL enhanced the urease activity of soil required for biocalcification, even at 4°C, as indicated by a decrease in urease activation energy temperature sensitivity. These experimental results agreed with the gradual decrease in hydraulic conductivity observed in the field when the average daily temperature was 7°C (days 24–69). It was suggested that the biocalcification is a potential technique to reduce the hydraulic conductivity of paddy field.

Assessment of the Parameters Influencing Microbial Calcite Precipitation in Injection Experiments Using Taguchi Methodology

Soil improvement is one of the major concerns in civil engineering. Therefore, a variety of approaches have been employed for different soil types. The loose granular soils and sediments have always imposed challenges due to their low strength and bearing capacity as well as presenting difficulties in drilling and excavation. Biomediated soil improvement, i.e., utilizing some bacteria to precipitate calcite on soil particles, has recently been introduced as a novel link of biotechnology and civil engineering to improve the problematic soils. Biogrout as a branch of biomediated soil improvement is based upon microbial calcium carbonate precipitation (MICP). In the present study, the Taguchi method with the aim of optimizing the process was utilized to design the experiments (DOE). A standard L9 orthogonal array with four parameters comprising bacterial cell concentration, molar concentration ratio of nutrient solution, curing time, and flow rate, each assigned to three levels, was selected. In this regard, soil samples were stabilized in sandy soil columns. Two-phase injections were conducted by injecting the bacterium Sporosarcina pasteurii PTCC 1642 in the first phase and nutrient in the second phase. Specimens were subjected to an unconfined compressive strength (UCS) test. ANOVA pointed out how effectual each parameter was. The most effective parameter was curing time, which accounted for 45.97% of the overall variance of the experimental data followed by bacterial cell concentration (22.01%), nutrient strength (19.98%), and flow rate (12.04%). Predicted UCS values for the optimum condition were validated in a confirmation test. Indeed, the UCS of the soil increased from 85 kPa in the control sample to 930 kPa for the optimally treated specimen. It was concluded that rather than curing time, the other parameters are almost equally influential in the applied injection procedure.

Biomineralization processes of calcite induced by bacteria isolated from marine sediments.

Biomineralization is a known natural phenomenon associated with a wide range of bacterial species. Bacterial-induced calcium carbonate precipitation by marine isolates was investigated in this study. Three genera of ureolytic bacteria, Sporosarcina sp., Bacillus sp. and Brevundimonas sp. were observed to precipitate calcium carbonate minerals. Of these species, Sporosarcina sp. dominated the cultured isolates. B. lentus CP28 generated higher urease activity and facilitated more efficient precipitation of calcium carbonate at 3.24 ± 0.25 × 10−4 mg/cell. X-ray diffraction indicated that the dominant calcium carbonate phase was calcite. Scanning electron microscopy showed that morphologies of the minerals were dominated by cubic, rhombic and polygonal plate-like crystals. The dynamic process of microbial calcium carbonate precipitation revealed that B. lentus CP28 precipitated calcite crystals through the enzymatic hydrolysis of urea, and that when ammonium ion concentrations reached 746 mM and the pH reached 9.6, that favored calcite precipitation at a higher level of 96 mg/L. The results of this research provide evidence that a variety of marine bacteria can induce calcium carbonate precipitation, and may influence the marine carbonate cycle in natural environments.

Functional gene diversity of oolitic sands from Great Bahama Bank

Despite the importance of oolitic depositional systems as indicators of climate and reservoirs of inorganic C, little is known about the microbial functional diversity, structure, composition, and potential metabolic processes leading to precipitation of carbonates. To fill this gap, we assess the metabolic gene carriage and extracellular polymeric substance (EPS) development in microbial communities associated with oolitic carbonate sediments from the Bahamas Archipelago. Oolitic sediments ranging from high-energy 'active' to lower energy 'non-active' and 'microbially stabilized' environments were examined as they represent contrasting depositional settings, mostly influenced by tidal flows and wave-generated currents. Functional gene analysis, which employed a microarray-based gene technology, detected a total of 12,432 of 95,847 distinct gene probes, including a large number of metabolic processes previously linked to mineral precipitation. Among these, gene-encoding enzymes for denitrification, sulfate reduction, ammonification, and oxygenic/anoxygenic photosynthesis were abundant. In addition, a broad diversity of genes was related to organic carbon degradation, and N2 fixation implying these communities has metabolic plasticity that enables survival under oligotrophic conditions. Differences in functional genes were detected among the environments, with higher diversity associated with non-active and microbially stabilized environments in comparison with the active environment. EPS showed a gradient increase from active to microbially stabilized communities, and when combined with functional gene analysis, which revealed genes encoding EPS-degrading enzymes (chitinases, glucoamylase, amylases), supports a putative role of EPS-mediated microbial calcium carbonate precipitation. We propose that carbonate precipitation in marine oolitic biofilms is spatially and temporally controlled by a complex consortium of microbes with diverse physiologies, including photosynthesizers, heterotrophs, denitrifiers, sulfate reducers, and ammonifiers.

Microbially Induced Calcium Carbonate Precipitation on Surface or in the Bulk of Soil

Microbial precipitation of calcium carbonate takes place in nature by different mechanisms. One of them is microbially induced carbonate precipitation (MICP), which is performed due to bacterial hydrolysis of urea in soil in the presence of calcium ions. The MICP process can be adopted to reduce the permeability and/or increase the shear strength of soil. In this paper, a study on the use of Bacillus sp., which was isolated from tropical beach sand, to perform MICP either on the surface or in the bulk of sand is presented. If the level of calcium salt solution was below the sand surface, MICP took place in the bulk of sand. On the other hand, if the level of calcium salt solution was above the sand surface, MICP was performed on the sand surface and formed a thin layer of crust of calcium carbonate. After six sequential batch treatments with suspension of urease-producing bacteria and solutions of urea and calcium salt, the permeability of sand was reduced to 14 mm/day (or 1.6×10−7 m/s) in both cases of bulk and surface MICP. Quantities of precipitated calcium after six treatments were 0.15 and 0.60 g of Ca per cm2 of treated sand surface for the cases of bulk or surface MICP, respectively. The stiffness of the MICP treated sand also increased considerably. The modulus of rupture of the thin layer of crust was 35.9 MPa which is comparable with limestone.