Susan H. Ferguson

Using Real-Time PCR to Assess Changes in the Hydrocarbon-Degrading Microbial Community in Antarctic Soil During Bioremediation

A real-time polymerase chain reaction (PCR) method to quantify the proportion of microorganisms containing alkane monooxygenase was developed and used to follow changes in the microbial community in hydrocarbon-contaminated Antarctic soil during a bioremediation field trial. Assays for the alkB and rpoB genes were validated and found to be both sensitive and reproducible (less than 2% intrarun variation and 25–38% interrun variation). Results from the real-time PCR analysis were compared to analysis of the microbial population by a culture-based technique [most probable number (MPN) counts]. Both types of analysis indicated that fertilizer addition to hydrocarbon-contaminated soil stimulated the indigenous bacterial population within 1 year. The proportion of alkB containing microorganisms was positively correlated to the concentration of n-alkanes in the soil. After the concentration of n-alkanes in the soil decreased, the proportion of alkane-degrading microorganisms decreased, but the proportion of total hydrocarbon-degrading microorganisms increased, indicating another shift in the microbial community structure and ongoing biodegradation.

Treatability studies: Microcosms, mesocosms, and field trials

Introduction In order to demonstrate the effectiveness of a bioremediation project, one must have an accurate measure of the contaminants, both at the start of the project and throughout the treatment process. The measurement of the contaminants throughout the process is important to demonstrate that the treatment is successful and to identify advances or set-backs quickly and effectively. Proving the disappearance of hydrocarbons is important to the success of a bioremediation project. An accurate measurement of hydrocarbons and their biodegradation products is needed to confirm that petroleum was actually consumed by bacteria (discussed in Chapter 7, Section 7.3). One method of confirming biodegradation of petroleum is the coupled measurement of biodegradation rates by proxy methods and the disappearance of the contaminant. Biodegradation rates do not, in and of themselves, prove the decomposition of contaminants. Measurement of biodegradation rates, however, can be an easy way to demonstrate that the potential exists for contaminant removal. While measures of biodegradation rates are often used to estimate time to closure for a site, or proof of technology, biodegradation rates can be unreliable. Common measures of aerobic biodegradation are loss of contaminants, oxygen (O 2 ) consumption, and carbon dioxide (CO 2 ) evolution. Unfortunately, the CO 2 can result from non-biological sources (see Chapter 7, Section 7.2.2.2 for additional discussion). Particularly in low pH groundwater, pH adjustment made during bioremediation could result in CO 2 off-gassing from groundwater. Oxygen depletion in the subsurface is also not proof of biodegradation.

Correction to Determining the Extent of Biodegradation of Fuels Using the Diastereomers of Acyclic Isoprenoids

The retention time shown in Figure 1 is incorrect. In this case, the gas chromatograph was run using constant pressure conditions (25 psi) rather than with the constant flow conditions (1.2 mL/min) detailed in the methods section. As a result, the peaks eluted around 15 min later. Figure 2 shows the correct retention times for the constant flow method detailed in the methods section. Also, Figure S2 in the Supporting Information is correct and also shows the retention times when run with constant pressure conditions (25 psi), hence it has later eluting peaks as in Figure 1.