Recently, self-healing applications of cement-based materials have received a lot of interest. One major area of interest with respect to self-healing applications in cement-based systems focuses on using biomineralization processes. Biomineralization is biochemical process in which microorganisms stimulate the formation of minerals. The existing research on biomineralization in cement-based systems has showed promising results and the studies suggest that biomineralization could be a useful approach for remediation of cracks on the surface of concrete. This dissertation presents the results of an intensive study undertaken to understand the influence of vegetative bacteria, specifically Sporosarcina pasteurii (S. pasteurii), when it is incorporated within cement paste. Vegetative S. pasteurii cells were suspended in a urea-yeast extract medium and this medium was mixed with cement. The influence of the vegetative S. pasteurii cells on Portland cement paste properties, such as compressive strength, hydration kinetics, and setting time was evaluated. It was determined that the hydration kinetics was highly influenced when the bacterial medium was used to prepare cement paste, and severe retardation was observed. It was also observed that an increase in calcium carbonate precipitation, particularly calcite, occurred within cement paste when the bacterial medium was used. Furthermore, use of the bacterial medium resulted in reducing the porosity and increasing the compressive strength of the hardened paste. Ex-situ culture experiments were conducted to determine the impact of pH and calcium concentration on the morphology of calcium carbonate precipitate; the results indicated that the morphology of the precipitate was more influenced by calcium concentration. A key focus of this dissertation was to examine the viability of the vegetative cells that were inoculated in cement paste. Viable S. pasteurii cells were found to be present in hardened cement paste samples that were as old as 330-days, and 50% of the viable cells detected were defined as vegetative cells. At last, the use of including internal nutrient reservoirs as a means to extend the viability of the bacterial cells within hardened cement paste was explored. The results showed that the percentage of vegetative cells remaining was affected when internal nutrient reservoirs was incorporated into the system. / text

<p>To make bacterial-based self-healing concrete, alkaline-resistant bacterial spores, nutrient sources, and a calcium source are incorporated into a concrete matrix. Two ureolytic spore-forming bacteria, <i>Sporosarcina pasteurii</i>, <i>Lysinibacillus sphaericus</i>, and two non-ureolytic spore-forming bacteria, <i>Bacillus cohnii</i>, and <i>Bacillus pseudofirmus</i>, which have been used in previous studies as bacterial concrete healing agents, were compared in this study. The four bacteria were compared for their (1) sporulation rates on different sporulation agar plates, (2) growth in five liquid media, (3) survival rates in light weight aggregates (LWA) and in mortar samples, and (4) calcium carbonate precipitation rates from either calcium lactate or calcium nitrate. Sporulation was successfully induced after three-day incubation at 30°C on an appropriate sporulation medium. High sporulation rates of <i>B. cohnii</i>, and <i>B. pseudofirmus</i>(93% and 99% respectively) were found on alkaline R2A medium (AR2A). A sporulation rate (89%) of <i>S. pasteruii</i>was observed on tryptic soy agar supplemented with 2% urea (TSAU)<i>.</i>The highest sporulation rate (60%) of <i>L. sphaericus</i>was found on R2A medium supplemented with 2% urea (R2AU). In the growth study, tryptic soy broth supplemented with 2% urea (TSBU) was a positive control which supported rapid growth of all four bacteria. <i>Sporosarcina pasteurii </i>and <i>L. pasteurii</i>showed rapid growth rates in alkaline yeast extract broth (AYE) and yeast extract with 2% urea broth (YEU) respectively. In contrast, <i>B. cohnii</i>, and <i>B. pseudofirmus</i>grew poorly in all media except in the positive control. Viable counts of the four bacterial spores reduced (1.8–3.3 logs) during the first 24 h in mortar samples and then remained stable for next 27 days testing period. Among the four, <i>S. pasteurii</i>showed the smallest reduction of viable counts (1.8–2.5 logs) in mortar after one day of incubation. Both <i>S. pasteurii</i>and <i>L. sphaericus</i>showed high CaCO₃productions (>80%) after 24 h incubation at 30°C in YEU containing either calcium nitrate or calcium lactate. However, <i>B. pseudofirmus</i>and <i>B. cohnii </i>showed<i></i>low calcite recovery rates (<11%) in AYE containing either<i></i>calcium nitrate or calcium lactate under the same incubation condition. Overall, <i>S. pasteurii</i>was the best bacterial concrete healing agent of the four. This bacterium had (1) rapid growth rate in AYE, (2) about 90% sporulation rate within 3 days, (3) highest survival rates after 24 h in mortar samples and, (4) high CaCO₃precipitation rates, 82 or 98%, in broth containing calcium nitrate or calcium lactate respectively.</p><p>In addition, two different lightweight aggregates (LWA), expanded shale (ES) and expanded clay (EC), which were used as bacterial carriers and protective materials, were compared in this study. Each type of LWA was separated into three sizes (<0.85 mm, 0.85– 2.0 mm, and >2.0 mm) and immobilized with spores of <i>B. cohnii</i>or <i>B. pseudofirmus.</i>Viable counts recovered from EC and ES reduced <1.0 log after the immobilization process and remained stable during the 150 days testing period. Neither the type nor the particle sizes of the two LWA significantly affected the survival rates of the bacterial spores. This result showed that both EC and ES could be used as carriers for bacterial healing agents. It was also found that when the spores were immobilized with nutrients in LWA, their survival rates in mortar samples can be improved slightly (<1.0 log).</p><p><br></p>

Microbial-Induced Calcium Carbonate (CaCO3) Precipitation (MICP) is a biological process in which microbial activities alter the surrounding aqueous environment and induce CaCO3 precipitation. Because the formed CaCO3 crystals can bond soil particles and improve the mechanical properties of soils such as strength, MICP has been explored for potential engineering applications such as soil stabilisation. However, it has been difficult to control and predict the properties of CaCO3 precipitates, thus making it very challenging to achieve homogeneous MICP-treated soils with the desired mechanical properties. This PhD study investigates MICP at both micro and macro scales to improve the micro-scale understandings of MICP which can be applied at the macro-scale for improving the homogeneity and mechanical properties of MICP-treated sand. A microfluidic chip which models a sandy soil matrix was designed and fabricated to investigate the micro-scale fundamentals of MICP. The first important finding was that, during MICP processes, phase transformation of CaCO3 can occur, which results in smaller and less stable CaCO3 crystals dissolving at the expense of growth of larger and more stable CaCO3 crystals. In addition, it was found that bacteria can aggregate after being mixed with cementation solution, and both bacterial density and the concentration of cementation solution affect the size of aggregates, which may consequently affect the transport and distribution of bacteria in a soil matrix. Furthermore, bacterial density was found to have a profound effect on both the growth kinetics and characteristics of CaCO3. A higher bacterial density resulted in a quicker formation of a larger amount of smaller crystals, whereas a lower bacterial density resulted in a slower formation of fewer but larger crystals. Based on the findings from micro-scale experiments, upscaling experiments were conducted on sandy soils to investigate the effect of injection interval on the strength of MICP treated soils and the effects of bacterial density and concentration of cementation solution on the uniformity of MICP treated soils. Increasing the interval between injections of cementation solution (from 4 h to 24 h) increased the average size of CaCO3 crystals and the resulting strength of MICP-treated sand. An optimised combination of bacterial density and cementation solution concentration resulted in a relative homogeneous distribution of CaCO3 content and suitable strength and stiffness of MICP-treated sand. This thesis study revealed that a microfluidic chip is a very useful tool to investigate the micro-scale fundamentals of MICP including the behaviour of bacteria and the process of CaCO3 precipitation. The optimised MICP protocols will be useful for improving the engineering performance of MICP-treated sandy soils such as uniformity and strength.