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Micromorphological and Biocalcification Effects of Sporosarcina Pasteurii and Sporosarcina Ureae in Sandy Soil Columns

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Micromorphological and Biocalcification Effects of Sporosarcina Pasteurii and Sporosarcina Ureae in Sandy Soil Columns J. Agr. Sci. Tech. (2014) Vol. 16: 681-693 Micromorphological and Biocalcification Effects of Sporosarcina pasteurii and Sporosarcina ureae in Sandy Soil Columns M. Sarmast 1, M. H. Farpoor 1*, M. Sarcheshmehpoor 1, and M. K. Eghbal 2 ABSTRACT Microbial calcium carbonate, by bridging sand particles, can play an important role in sand dune stability. A study was carried out on the cementation of sand grains and infilling of pore spaces by CaCO 3. Two bacterial species ( Sporosarcina pasteurii and Sporosarcina ureae ), three reactant concentrations (0.5, 1 and 1.5M), and six reaction times (12, 24, 48, 96, 192 and 288 hours) were tested in factorial experiment. Bacterial inocula and reactant solutions were added daily to sandy soil columns (6.5 cm height and inner diameter of 7.7 cm), while precipitation of CaCO 3 being investigated within 0-1.5, 1.5-3, 3-4.5 and 4.5-6 cm intervals. Chemical and micromorphological analyses revealed that CaCO 3 formation, inorganic C sequestration, and depth of cementation were more profound for S. pasteurii as compared with S. ureae . Both microbial CaCO 3 precipitation and inorganic C sequestration increased with increase in reaction time from 12 to 288 hours. Increase in reactant concentration also caused an increase in CaCO 3 precipitation (by 12%). Micromorphological observations showed a high degree of calcite crystals’ bridging, coating on sand particles and as well infilling of pore spaces. S. pasteurii is thus recommended for being used in stabilization of sand dunes; due to its significant effects on CaCO 3 deposition and as well on sand grain cementation. Keywords: Biomineralization, Inorganic C sequestration, Sand dune fixation, Ureolytic bacteria. INTRODUCTION One mole of urea is hydrolyzed intracellularly into 2 moles of ammonia and Biocalcification is the process of calcite 1 mole of CO 2 [Equations (1) and (2)] through urease enzyme activity (Hammes et Downloaded from jast.modares.ac.ir at 21:24 IRST on Friday September 24th 2021 formation resulting from metabolic activities of autotrophic and heterotrophic organisms. al ., 2003). Soil pH increases as a result of Using metabolic pathway of ureolytic this enzymatic reaction, and CaCO 3 [Equations (3) and (4)] will be precipitated hydrolysis through nitrogen cycle, some of 2- 2+ the heterotrophic bacteria are of the potential from CO 3 and Ca ions (Siddique and Chahal, 2011). Meanwhile, produced CO 2 is of providing high concentrations of CaCO 3 in short periods of time (Castanier et al ., trapped in CaCO 3 preventing CO 2 emission into atmosphere. 1999; De Muynck et al ., 2010). Bacillus , → Sporosarcina , Sporolactobacillus , CO(NH 2)2 + H 2O NH 2COOH + NH 3 (1) Clostridium and Desulfotomaculum genera → are among the most urease producing NH 2COOH + H 2O NH 3 + CO 2 + H 2O bacteria in soils (Ivanov and Chu, 2008). (2) _____________________________________________________________________________ 1 Department of Soil Science, Faculty of agriculture, Shahid Bahonar University of Kerman, Kerman, Islamic Republic of Iran. 2 Department of Soil Science, Faculty of Agriculture, Tarbiat Modares University, Tehran, Islamic Republic of Iran. * Corresponding author; e-mail: [email protected] 681 ______________________________________________________________________ Sarmast et al. - + CO 2 + H 2O ↔ H 2CO 3 ↔ HCO 3 + H ↔ 100 grams sterile sand treated by Bacillus 2- + CO 3 + 2H (3) pasteurii were used by Stocks-Fischer et al . 2- 2+ CO 3 + Ca → CaCO 3 (4) (1999) to study plugging of porous media. Landi et al . (2003) demonstrated that the XRD and SEM analysis showed that calcite soils of the prairies and of the forests have was precipitated in between sand particles. sequestered biogenic C from plant respiration Using urease enzyme in glass columns (H: 30 and residues in the form of pedogenic and ID: 2.5 cm) packed with a mixture of sand carbonates and Ca is the rate-limiting factor to and glass beads, Nemati and Voordouw (2003) be precipitated with CO 2. Microbial activity created calcite cementation, which in turn helps inorganic sequestration of C in CaCO 3 caused a 98% decrease in permeability. form. Geological sequestration of CO 2 and its Porosity and permeability decrease induced by conversion into carbonate solid phases were Sporosarcina pasteurii in sand columns has studied by Dupraz et al . (2009) using Bacillus also been reported by Cunningham et al . pasteurii . In a study of factors controlling (2011) and Whiffin et al . (2007). urease enzyme and microbial deposition of Role of microbial deposition of calcium CaCO 3 using Sporosarcina pasteurii carbonate on soil physical properties (porosity, previously known as Bacillus pasteurii (Yoon permeability, and as well hydraulic et al ., 2001), Okwadha and Li (2010) found conductivity) has been investigated by several that CO 2 sequestration rate caused by ureolytic researchers (Whiffin et al ., 2007; Cunningham hydrolysis was directly proportional to et al ., 2011; Sarmast et al ., 2011a, b). Vast microbial CaCO 3 formation. areas in arid and semi-arid ecosystems are Calcium carbonate precipitation induced by covered by sand dunes, the stabilization of bacterial activity has extensively been which is of great importance. Besides, employed to increase stability of structures in micromorphological observations, civil engineering. Ghosh et al . (2006) used demonstrating microbial calcite morphology thermophilic and anaerobic bacteria within and pattern that help bridge and cement cement-sand mortar/concrete to develop particles are lacking in literature. Despite the bioconcrete material. Results of the study fact that genesis of calcium carbonate in indicated that compressive strength of concrete calcareous soils of Iran is reported in the and cement-sand mortar increased literature (Khormali and Nabiollahi, 2009; significantly, due to the development of Farpoor et al ., 2011; Moazallahi and Farpoor, crystals within the pores of cement sand 2012; Owliaie, 2012), however, limited matrix. Effects of pure Bacillus sphaericus and information is presently available on biocalcite Downloaded from jast.modares.ac.ir at 21:24 IRST on Friday September 24th 2021 a mixed culture of ureolytic bacteria on formation in soils and sediments in Iran. The microbial CaCO 3 deposition and durability of present research was conducted to: mortar and concrete were studied by De 1). Study the levels, reaction time, and depth Muynck et al . (2008a, b). They reported that of secondary carbonates. deposited calcite on the surface of samples by 2). Compare Sporosarcina pasteurii vs. bacteria decreased capillary water uptake as Sporosarcina ureae microbial activity on well as permeability towards gas. Carbonate calcium carbonate formation. precipitation induced by microbial activity 3). Determine the micromorphology of using Bacillus pasteurii , an endospore forming bridging and cementing particles. soil microorganism, takes place with the highest urease activity, Sarda et al . (2009) on MATERIALS AND METHODS brick samples to increase the strength and durability. Microbial deposition of calcium carbonate The research was conducted in the has also been employes to improve physical framework of a factorial, completely properties of soils and other porous media. randomized design experiment of 3 Plastic syringe columns (60 mL) containing replicates. Effects of two bacterial species 682 Microbial Calcium Carbonate Formation in Soils _________________________________ (Sporosarcina pasteurii and Sporosarcina Soil Column Parameters ureae ), 3 concentrations of urea and CaCl 2 mixture (0.5, 1 and 1.5M), along with 6 time Plastic columns with heights of 6.5 and intervals (12, 24, 48, 96, 192 and 288 hours) inner diameters of 7.7 cm were packed with were studied on microbial calcium carbonate 500 grams of sterile sandy soil to study the formation. depth of bacterial activity. The daily flows including inoculum, fixation flow of 50 mM Soil Sample CaCl 2 and cementation flow (urea + CaCl 2) were added uniformly and continuously to Soil sample was taken from sand dunes of the columns. The sandy soil columns were Jupar area in Kerman Province. Air dried incubated at 28±2˚C. Flow rates were soil sample was sieved through a 2 mm adjusted according to the hydraulic conductivity of the sandy soil (10 -2-10 -3 cm sieve and routine soil physicochemical -1 properties determined. Pipette method was sec ). employed for particle size distribution analysis of the soil (Gee and Bauder, 1986). Microorganisms and Growth Electrical conductivity of the saturated Conditions extract (Rhoades, 1982), and pH of the saturated paste (Mclean, 1982) were Sporosarcina pasteurii (PTCCi 1645: determined applying Jenway EC and pH previously known as Bacillus pasteurii ; meters. Calcium carbonate equivalent (CCE) Yoon et al ., 2001) and Sporosarcina ureae was investigated through back titration (PTCCi 1642) were obtained from Iran (Nelson, 1982). Gypsum content was Research Organization of Science and measured through acetone method (Nelson Technology. Both species are gram positive, and Sommers, 1982). No gypsum was endospore forming, of the potential of being detected during chemical analyses and grown in alkaline environments, and bear through morphological observations. Table 1 urease activity. Bacterial inoculums, using shows physicochemical properties of the soil appropriate medium cultures (Table 2), were sample used in the experiment. prepared at 30˚C, being shaken for 24 hours and at 140 rpm. The pH of the media was Table 1 . Selected physicochemical
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