Kinetic analysis of data obtained from studies on microbial degradation of cement waste forms, using shrinking core models.

Model equations based on analytical solutions of two shrinking core models (acid dissolution or shrinking unreacted core (SUC) model, and bulk diffusion model), were used to analyze the kinetics of microbial degradation of cement waste forms. Two current approaches of waste form microbial stability evaluation (Nuclear Regulatory Commission (NRC) method and refined biofilm formation) were used to generate the data. Good linear correlations with R(2)>0.95 were obtained for the leaching data from both the NRC and biofilm approaches, using the model equation based on the bulk diffusion concept. Analyses using the model equation based on the acid dissolution model generally gave poor correlations except when data obtained from biofilm formation method was normalized.

Stability evaluation of a cement based waste form to microbially induced degradation.

In this study the current Nuclear Regulatory Commission (NRC) protocol is used to evaluate the stability of Tuskegee cement/cobalt chloride waste form in the presence of Thiobacillus thiooxidans (T. thiooxidans). A critical examination of this protocol and identified limitations are reported also. Tuskegee cement/cobalt chloride waste forms were shown to exhibit considerable instability to microbial degradation as indicated by significant physical deterioration, and increased leaching of calcium and cobalt on exposure to T. thiooxidans. The instability was aggravated with higher levels of cobalt chloride content of the waste forms. The degradative capability of T. thiooxidans closely followed its ability to significantly decrease the pH of its environment. Inherent limitations in the NRC protocol were observed which could lead to serious result interpretation errors. The use of a T. thiooxidans culture that is significantly lower in pH in comparison to the control medium could lead to an overestimation of the degradative effect of T. thiooxidans, while the use of a culture that is substrate limited could result in an underestimation of T. thiooxidans capability.

Production of asporogenous mutants of Bacillus sphaericus 2362 in continuous culture.

AIMS: To report the production of asporogenous mutants (Spo-) of Bacillus sphaericus 2362 in continuous culture. METHODS AND RESULTS: Microbial culture samples were taken at 0.05 h-1 dilution rate and plated out on nutrient agar plates. Translucent colonies were obtained with vegetative morphology under phase contrast microscope. Heat resistance evaluations at different temperature settings showed that the Spo- mutants had lower heat resistance than the Spo+ wild type. Western blots analyses carried out on both wild type and the mutants indicated the presence of binary protein toxins of 42 and 51 kDa in both. Bioassays carried out on the wild type and the Spo- mutants against mosquitoes showed the mutants to be 100-fold less toxic in comparison to the wild type. CONCLUSION: Existence and production of asporogenous mutants of Bacillus sphaericus 2362 in continuous culture at low dilution rates is demonstrated by this study. The organism's ability to produce toxins appears to be significantly reduced by the mutational process. SIGNIFICANCE AND IMPACT OF THE STUDY: The production of asporogenous mutants had not been reported previously among strains of Bacillus sphaericus. The present report on the toxigenic capability of asporogenous mutants also raises the possibility of using continuous culture to significantly improve the productivity of toxin production in future.

A refinement of the biofilm formation method for waste forms stability evaluation.

A refinement of the biofilm formation method for waste form stability evaluation was carried out in this study. Refinement of the biofilm formation method became necessary because of the reduced contrast in degradation between control and experimental samples. The reduction in contrast was occasioned by the long duration of exposure (12 days) of the control samples to sterile medium of low pH in the first stage. Results of evaluation carried out reveal that the duration of the first stage of the biofilm formation method can be reduced to 24 h, with substantial increase in the contrast between degradations experienced by control and experimental samples. Reduction of the first stage can be done without compromising the efficiency of the inoculation process, which the longer duration of the first stage was originally intended to ensure. A doubt as to actual formation of biofilms on experimental samples, resulting from the use of non-sterile tubings and glass wares in the second stage, was also addressed in this study. Results reveal that substantial attachment of microbes occur on the surfaces of experimental samples in the first stage, thus any supply of microbes via the tubings and glass wares in the second stage is only additional and inconsequential.

Development of a biofilm formation method for waste forms stability evaluation.

The development of an accurate assessment protocol is critical for the prediction of long-term performance of waste disposal systems under field conditions. In this study, the development of a biofilm formation method for the evaluation of waste forms stability to microbially induced degradation (MID) is reported. The development process involved significant modifications to the existing Nuclear Regulatory Commission (NRC) approach. In the biofilm formation method, the control media and fermenter broths are designed to be of similar pH to avoid overestimation of the microbe's capability to degrade the waste forms. In the NRC approach, the pH values are different. The existing one-stage process of the NRC approach is also replaced with a two-stage process in the biofilm formation method. This is to ensure full evaluation of the microbe's involvement in waste forms degradation. The first stage of the two-stage process is for biofilm formation and the second is for biofilm evaluation. The use of a two-stage process eliminates the possibility of substrate limitation, resulting in values of degradation indices that are about two times higher than those obtained using the single-stage NRC approach. Two waste forms (100% Tuskegee cement and 21% cobalt chloride/79% cement) were used in the development of the biofilm formation method. Both waste forms showed evidence of biofilm formation. The formation of biofilm on the cobalt-containing waste form indicates a lack of anti-microbial capability of cobalt.