Studies On The Adsorption Of Bacteria By Clay Minerals And Soil Particles

Soil particles and clay minerals such as kaolinite, montmorillonite, and goethite were selected and Bacillus thuringiensis and Pseudomonas putida were used in this study. The correlation of bacterial number with total bacterial protein was explored; bacterial adsorption amount was represented as total bacterial protein; based on the former two and Nycodenz separation reagent with equal density gradient, the measurement method was established about bacterial adsorption on clay minerals. Separation efficiency was compared between 1.31 g ml-1 of sucrose solution and Nycodenz solution, and experimental conditions of bacterial adsorption were investigated on soil sand and silt. Adsorption isotherm and relatively influencing factors were probed and then the isotherm was fitted to Langmuir equation. On the basis of surfacee physicochemical properties of soil minerals and bacteria such as specific surface area, point of zero charge, zeta potential and contact angle, adsorption mechanisms were preliminarily analyzed using DLVO theory and other theories from the view of physical chemistry. The main results were listed as following:1. This study firstly tried to separate free cells from adsorbed cells in suspension of clay minerals and bacteria and managed to establish a measurement method of bacterial adsorption on clay minerals. It was optimum to control bacterial number of 12-15×108 cells and mineral concentration of 1.75-2.50 mg ml-1. After approximate adsorption equilibrium of 60 min, sucrose solution of 60% (w/v) and Nycodenz solution of 3-5 ml were used to separate free cells from adsorbed cells, respectively. Adsorption amount of bacteria was characterized as total bacterial protein determined by Bradford method.2. Separation efficiency was clear through comparing 1.31 g ml-1 of sucrose solution with Nycodenz solution, and experimental conditions of bacterial adsorption on soil sand and silt were surveyed. The results showed that 1.31 g ml-1 of sucrose solution could replace Nycodenz solution. In the experiment of bacterial adsorption on soil particles, the concentrations of soil sand and silt suspension were 60-120 mg ml-1,4.5-7.5 mg ml-1, respectively and proper adsorption equilibrium time was 45-60 min.3. The measurement method of bacterial adsorption on clay minerals was firstly established. Adsorption isotherm was determined between kaolinite, montmorillonite and goethite and B. thuringiensis and P. putida. Effect of environmental factors, several low molecular weight organic acid and phosphate ligands, and bacterial growth phase and activity on bacterial adsorption by minerals was measured. The results revealed that (1) adsorption isotherms accorded with Langmuir equation about two kinds of bacteria with kaolinite, montomorillonite and goethite; maximum amount of bacteria adsorbed on goethite and kaolinite was larger than that of montmorillonite; the affinity of B. thuringiensis onto kaolinite and montmorillonite was bigger than the affintity of P. putida whereas goethite presented bigger affinity with P. putida relative to B. thuringiensis; (2) maximum amount of bacteria adsorbed was occurring at the temperature of 15-25℃; amount of bacteria adsorbed increased with the decreasing pH, and the decreasing order was goethite> kaolinite> montmorillonite; ionic strength could greatly promote adsorption of bacteria onto clay minerals and in the view of minerals, the promoting order was montmorillonite> kaolinite> goethite, and as to bacteria, P. putida> B. thuringiensis; Mg2+expressed stronger promotion effect relative to Na+and the effect increased with increasing pH; (3) most ligands inhibited adsorption of bacteria on minerals except that acetate accelerated adsorption of bacteria by kaolinite and goethite at pH 5.5; strong inhibition happened at low concentration of ligands (0-10 mmolL-1 or 20 mmolL-1), whereas inhibition decreased or levelled off at high concentration (20 mmolL-1 or 50-80 mmolL-1 or 200 mmolL-1); inhibition of several ligands on adsorption of bacteria by minerals correlated with mineral types and solution pH values; effect of inhibition of ligands on adsorption of P. putida was stronger than on B. thuringiensis; (4) adsorption isotherm of bacteria whether viable or dead by kaolinite, montmorillonite and goethite corresponded to Langmuir equation. Compared with bacteria from the stationary phase, adsorption of viable B. thuringiensis from exponential phase by kaolinite and montmorillonite took on decline, however, maximum amount of viable P. putida adsorbed from exponential phase by the above minerals showed slight increase at the pH of 5.5 and decline at the pH of 7.0; moreover, maximum amounts of two viable bacteria adsorbed from exponential phase by goethite displayed increase; there existed decline in the maximum amounts of the both dead bacteria adsorbed by kaolinite and montmorillonite but increase by goethite; compared with both bacteria, the maximum amount of dead bacteria P. putida adsorbed by kaolinite and montmorillonite decreased greatly relative to that of B. thuringiensis and showed greater decline for dead bacteria from exponential phase than from stationary phase; nevertheless, the maximum amount of dead bacteria of P. putida adsorbed expressed greater increase than that of B. thuringiensis.4. This study pioneered on measuring adsorption isotherm of P. putida and B. thuringiensis on red soil particles, and influence of environmental factors, several low weight organic acids and phosphate, bacterial growth phase and activity on bacterial adsorption was also determined. The results indicated that (1) adsorption isotherm for both bacteria accorded with Langmuir equation; the amount of adsorbed bacteria by soil particles with different size followed the sequence of clay> slit> fine sand> coarse sand, which by clay was 3.7-4.9,44.3-89.2 and 262-799 times larger than by silt, fine sand and coarse sand; the difference was 389 and 857 times between maximum and minimum amount of B. thuringiensis and P. putida adsorbed; the affinity of B. thuringiensis onto soil particles of the same size was higher than that of P. putida and B. thuringiensis displayed higher affinity onto soil particles of the same size with organic matter removed (soil inorganic particles) than with organic matter left (soil organic particles); (2) maximum amount of bacteria adsorbed achieved at temperature of 15-25℃; bacterial amount of soil particles with different size decreased with increasing pH and the decrease followed the sequence of coarse sand> fine sand> silt> clay, soil organic particles> soil inorganic particles and amount of P. putida adsorbed by soil particles appeared great decline relative to that of B. thuringiensis; ionic strength presented great promotion effect on bacterial adsorption, in which Mg2+was greater than Na+and the amount of bacteria adsorbed followed the order:coarse sand> fine sand> silt> clay and soil organic particles> soil inorganic particles; on the effect of Mg2+, the largest increasing amount of bacteria adsorbed was 10.7 times in the case of organic coarse sand and P. putida whereas the smallest decreasing amount showed 1.6 times in the case of inorganic clay and B. thuringiensis; (3) ligands inhibited adsorption of bacteria on red soil particles with different size, besides acetate with accelerating effect of adsorption on clay separated from red soil, in which the decreasing followed the sequence of clay> silt> fine sand> coarse sand, and in the same size, inorganic particles > organic particles; stronge inhibition happened at low concentration of ligands (0-10 mmolL-1 or 20 mmolL-1), whereas inhibition decreased or levelled off at high concentration (>20 mmolL-1 or 50 mmolL-1); inhibition effect of ligands on adsorption of bacteria was relative to particle size and particles of the same size with organic matter removed or left; (4) adsorption isotherm of viable bacteria adsorbed in exponential phase and dead bacteria adsorbed in exponential and stationary phases by soil particles corresponded to Langmuir equation; compared with stationary phase, viable bacteria of B. thuringiensis in exponential phase presented decreasing adsorption and amount of P. putida adsorbed by sand increased slightly whereas decreased mildly by silt and clay; amount of dead bacteria adsorbed decreased by all different particles and amount followed the order of coarse sand> fine sand> silt>clay, at the same size, organic particles> inorganic particles; compared with both bacteria, amount of dead P. putida adsorbed in exponential phase decreased by 31.1% larger than 25.6% in stationary phase, and however, amount of dead B. thuringiensis adsorbed in stationary phase decreased by 22.0% larger than 18.7% in exponential phase.Based on comprehensive analysis, DLVO theory could be used to elucidate bacterial adsorption onto clay minerals and soil particles. Adsorption behavior was controlled by surface physicochemical properties such as surface charge and hydrophobicity of soil minerals and particles and bacteria, in addition, the key role of electrostatic attraction. Surface properties of clay and soil particles would be influenced by different types of clay minerals, soil particles of diverse size, composition and properties, furthermore, organic matter existing or not, solution chemistry properties such as various temperature, pH, ionic strength and low molecular weight organic and inorganic ligands, and also solution chemistry would impact bacterial surface properties. As an organism, bacteria would have dissimilar surface structure and character during different growth phases and activity. Therefore, solution chemistry properties were of fundamental significance in bacterial adsorption onto soil particles and minerals.