Formatted Title
Characterizing the Kinetics of Microbial Growth and Substrate Utilization in Reductive Dehalogenation and Chain Elongation
Background/Objectives
Globally, chlorinated solvents are some of the most ubiquitous groundwater contaminants. Dehalococcoides mccartyi is a key microbe in remediating chlorinated solvents, as various strains are capable of dechlorinating perchloroethene (PCE), trichloroethene (TCE), dichloroethene (DCE), and vinyl chloride (VC) in their energy metabolism. In reductive dechlorination, D. mccartyi uses H2 as its sole electron donor, and H2 limitation may contribute to the disconnect observed between the efficient dechlorination of TCE-containing microcosms in the laboratory, which can take a matter of days, and the time required to remediate and close sites, which can take months, years, or decades. During in situ bioremediation, the H2 requirement of D. mccartyi is commonly fulfilled by adding organic substrates such as lactate, glucose, or emulsified vegetable oil, which undergo fermentation. Over ten times the theoretical amount needed must be added to account for H2 use by competing microbial processes. Preliminary data support the viability of developing reductive dechlorination approaches based on the microbial chain elongation (MCE) metabolism of Clostridium kluyveri, which consumes ethanol and acetate to produce short and medium chain fatty acids (MCFAs) butyrate, caproate, and caprylate, as well as H2 gas. The primary goal of this project is to gain insights into the kinetic and ecological interactions that govern the two processes, which will be essential for engineering more effective bioremediation approaches.
Approach/Activities
Growth, substrate utilization, and kinetic parameters of interest include Ks, the concentration of the substrate at which the consumption rate is half of the maximum (Ms/L3), qmax, the maximum specific rate of substrate utilization (Ms/MxT), Y, the cell yield coefficient (Mx/Ms), and b, the endogenous-decay coefficient (1/T) (M = mass, L = length, T = time, S = substrate, and X = microbial biomass). Batch experiments in 160-mL glass serum bottles containing reduced anaerobic medium were performed with both pure cultures of D. mccartyi strain GT (reduces TCE to ethene) and mixed microbial communities.
Results/Lessons Learned
A mixed culture named MAT has been established in our lab that is capable of reductively dechlorinating TCE to non-toxic ethene and elongating acetate and ethanol to C4-C8 carboxylates. The D. mccartyi culture was fed with 0.2-0.8 mmol L-1 TCE and stoichiometric amounts of H2 gas. The mixed culture MAT was fed with acetate + ethanol and butyrate + ethanol at various molar ratios. Throughout the batch kinetic experiments, TCE and its daughter products cis-DCE, VC, and ethene was measured regularly via gas chromatography with flame ionization detection. H2 gas was monitored using a reduced gas analyzer and a gas chromatograph equipped with a thermal conductivity detector. Chain elongation substrates and products were quantified via high performance liquid chromatography. These experiments are on-going. At the time of the conference, the growth, substrate utilization, and kinetic parameters will presented. This systematic kinetic study will provide insights into the rate dependence of reductive dehalogenation on the rate and end products of chain elongation, thereby filling a critical knowledge gap and supporting the development of bioremediation strategies based on microbial chain elongation.