BACKGROUND AND OBJECTIVES: Soil frost heaving causes significant destruction to road pavements, railways, pipelines, and other lifeline infrastructures. The conventional methods for dealing with the soil frost heave are primarily based on using the materials whose production and use are harmful to the environment. Due to the recent ecological concerns, developing novel alternative methods has received much attention. This study aims to investigate the possibility of using the microbially INDUCED CALCITE PRECIPITATION method to control soil frost heave for less pollution introduction to the soil. METHODS: In this study, the Sporosarcina Pasteurii bacterium was used for CALCITE PRECIPITATION. The influence of three factors in four levels, including bacteria concentration, cementing solution concentration, and curing time, was investigated based on a plan set by Taguchi design of experiment method. The results were obtained by analysis of means and analysis of variance statistical methods and compared with the conventional frost heave reduction methods. FINDINGS: The results were presented in terms of heave ratio. Based on the testing results, the heave ratios (frost heave ratios of the treated to untreated samples) were obtained to be in the range of 0. 21 to 0. 42. The results showed that bacteria concentration was the most influential factor in the total frost heave of the treated soil. The influence of curing time was in second place, and the effect of cementing solution concentration was relatively less. The minimum frost heave was achieved in 10 colony-forming units per milliliter bacteria concentration, 0. 6 mole per litre cementing solution concentration, and 21 days of curing. CONCLUSION: The findings indicated that the used method could be efficiently used to reach the desired objective. The heave ratios obtained by this method were promising to a great extent compared to the conventional methods. The reduction of frost heave due 8 to the application of this method was attributed to the precipitated CALCITE within the soil voids and was justified by the scanning electron microscopy images of the treated soil samples. This study proved that the proposed method might be utilized as a potential ecological-friendly approach in the future researches.

This study assessed the role of picocyanobacterial photosynthesis in the induction of CALCITE PRECIPITATION. It aimed at establishing whether photosynthetic uptake of bicarbonate by Synechoccoccus cells leads to CALCITE nucleation. The PRECIPITATION of CALCITE was initiated by addition of previously washed cyanobacterial cells of Synechococcus strain PCC 7942 to solutions of calcium carbonate at different saturation levels with respect to CALCITE. PRECIPITATION experiments were performed under controlled laboratory conditions in two set-ups: one in which photosynthesis was inhibited using a herbicide called Diuron and the other one in which photosynthesis was taking place. During the experiments, a pH meter monitored the pH and ion selective electrodes monitored concentrations of carbonate and calcium ions. The morphology of the precipitated crystals was analysed using Scanning Electron Microscopy. When the kinetics of calcium carbonate nucleation by the Synechococcus cells were compared for the two sets of experiments, there were very little differences. In fact, the induction times for PRECIPITATION reactions with photosynthesis were shorter due to the uptake of carbon dioxide. It is therefore ‘ concluded that photosynthesis does not directly influence the nucleationof CALCITE at the surface of Synechococcus cells with sufficient supply of carbon dioxide, i.e. cells took up carbon dioxide and not bicarbonate.The microscopic observations, however, provided some evidence that picocyanobacterial cell walls act as a template for CALCITE nucleation.

Biological cementation is a new process in which urea hydrolysis bacteria or free urease enzyme decompose urea and increases the pH of the environment and chemical interactions in the presence of calcium ions to form CALCITE. Nowadays, nano-CALCITE is widely used in engineering, such as increasing the strength of soil and concrete, as well as in medicine, such as drug delivery and cancer treatment. This study aimed to investigate the laboratory conditions for producing nano-CALCITE particles with appropriate quality, size and purity by Sporosarcina pasteurii enzyme extract for use in medical and engineering studies. This investigation aimed to make CALCITE by S. pasteurii enzyme extract and optimize influential factors in CALCITE production. For this purpose, the bacterium S. pasteurii was cultured in nutrient broth containing urea and nickel, and upon reaching the appropriate time, the cells were separated and washed. Then, their enzyme extract was prepared by sonication, and CALCITE PRECIPITATION was studied in different amounts of urea, calcium chloride, enzyme and temperature. The quality of produced CALCITE crystals and their ratio compared to other crystals were investigated by XRD and SEM analyses. According to the results of XRD analysis, it was found that in 0.5 M urea and 0.25 M calcium chloride, the highest amount of CALCITE is produced with 96%, and the least side products are produced. Examining the particle size histogram in the sample containing 0.5 M urea and 0.25 M calcium chloride revealed that the range of particles were between 50 and 100 nm. The nature and type of crystals were studied by electron microscopy, and EDX analysis showed the presence of calcium, oxygen, and carbon. According to the results, it was found that by the increase of the concentrations of urea and calcium, the range of particle size became larger. Also, the percentage of CALCITE produced in low urea and calcium chloride concentrations is higher than those in high concentrations.

Introduction: The present study was focused on the statistical optimization of growth parameters for enhancing the Microbially INDUCED CALCITE PRECIPITATION (MICP) using ureolytic yeast strain. Materials and Methods: Thirteen yeast strains were tested for the synthesis of urease enzyme by phenol-hypochlorite assay and were further evaluated for CALCITE PRECIPITATION test. The growth parameters were optimized using the best ureolytic strain by Box-Behnken Design (BBD) and the extracted MICP was characterized through instrumental analysis. Results: Among thirteen yeast strains, Candida tropicalis NN4, Spathospora sp. NN04, Wickerhamomyces anomalus VIT-NN01 and Candida dubliniensis NN03 showed positive results for the synthesis of urease enzyme. Spathospora sp. was found to be the most potent strain for MICP. A significant enhancement in MICP by Spathospora sp. was observed under optimized conditions viz. A-urea concentration (80. 0 g/L), B-calcium chloride (45. 0 g/L), C-pH (9. 0) and D-inoculum dosage (8%, v/v). The actual value (34. 4± 0. 12 g/L) was in agreement with predicted value (34. 7± 0. 01g/L) with the R 2 value (0. 9900), confirming the validity of the model. The FTIR of MICP confirmed the fundamental bands of CO3 stretching and bending vibrations, observed at 1394. 23 and 874. 85 cm-1. The Scanning Electron Microscope (SEM) images of biomotar revealed aggregated polymorphs of MICP interconnected with yeast mycelium and spores. The Energy Dispersive X-Ray Spectrometer (EDX) analysis indicated the presence of CALCITE in the biomotar. A remarkable improvement in the compressive strength (28 to 44 MPa) and morphological changes were observed in biocement mortar as compared to cement mortar. Conclusions: This result is the first report on the implementation of ureolytic Spathospora towards the application of biocementation through MICP using BBD.

Microbial-INDUCED CALCITE PRECIPITATION (MICP) is a relatively green and sustainable soil improvement technique. It refers to a chemical reaction network that is managed and controlled within soil through biological activity and whose byproducts alter the engineering properties of soil. To treat soil, first, the microbial population in-situ is augmented by the injection of additional urease positive bacteria and then reagents are added. This paper provides an overview of the factors affecting the MICP in soil. Several factors including nutrients, bacteria type, geometric compatibility of bacteria, bacteria cell concentration, fixation and distribution of bacteria in soil, temperature, reagents concentration, pH, and injection method are introduced. These factors were found to be essential for promoting successful MICP soil treatment. Furthermore, a preliminary laboratory test was carried out to investigate the potential application of the technique in improving the strength and impermeability of a sand specimen and utilized techniques, materials, methods and empirical process during the test are explained. The results showed that as a result of the CALCITE PRECIPITATION, shear wave velocity increased up to 1000m/s and UCS strength increased to about 300Kpa and permeability of soil decreased significantly upon MICP treatment.

One of the most important causes of soil desertification is wind erosion. Deserts are known for their poor physical, chemical, and biological properties. In order to solve the problem of loose sand, many solutions have been thought and used so far. Today, scientists have greatly benefited from the capabilities of MICP in the field of sand dune stabilization, which seems to be an environmentally friendly solution. In Khuzestan, a lot of "vinasse" enter the environment every year, which is considered a pollutant. Combining it with beneficial native microorganisms can turn it into a valuable ingredient. The objectives of this study were to investigate the microbial activity of bacteria isolated from the sand dunes of Khuzestan in Iran using vinasse as a carbon source for growth. In order to eliminate the effect of competition and investigate the activity of the studied bacterial isolates (Bacillus licheniformis MZ057843, MZ057842, OP329211, and Sporosarcina pasteurii), the samples of sand dunes collected from Khuzestan were first sterilized. Then, inoculated vinasses were added to them, and the prepared samples were incubated for five weeks. The experiments (counting bacteria by plate count technique, Soil catalase by the method of titration with KMnO4, and Soil microbial respiration by measuring the amount of CO2) were measured after the first, third and fifth weeks of incubation time. Then, the data were analyzed by split plot in time design. The current study showed significant effects of vinasse, bacteria, time and their interaction in all three tests (p < 0.0001). Isolate B1 was able to cause the highest amount of microbial population and microbial respiration, and isolate B3 produced the highest amount of catalase enzyme in the soil. Among the three types of vinasse, V3 had the highest amount of catalase and microbial population, and V2 had the highest soil microbial respiration rate. The best incubation time for the microbial respiration and population tests was the first week, and third week of incubation for the catalase test. Generally, native Bacillus bacteria grew in vinasse as a carbon source and perhaps the potential of vinasse as a mulch can be explored. However, more research is needed to use it in soil stabilization operations.