Biodegradation of Subsurface Pollutants by
Chlorate-Respiring Microorganisms

National Science Foundation (NSF). May1998 - April 2001.
Principal Investigator: Bruce Logan
Kappe Professor of Environmental Engineering
Penn State University
Phone: 814-863-7908, Fax: 814-863-7304, Email: blogan@psu.edu

Project Abstract

Subsurface bioremediation is limited in part by the availability of suitable electron acceptors for microorganisms capable of degrading target chemicals. Compounds investigated by others (such as nitrate, sulfate and hydrogen peroxide) have disadvantages that can include: low solubility, toxicity, low energy yields (resulting in slow growth rates of targeted microorganisms), and/or long acclimation times. It is proposed to investigate the potential for using chlorate (ClO3-) as an alternate electron acceptor for subsurface remediation processes coupled with injection of chlorate reducing microorganisms (CRMs). CRMs are good candidates for bioaugmentation for several reasons. Because chlorate is not a naturally occurring compound in nature, it is unlikely that there is large native chlorate-respiring population in-situ that would compete with injected microbes for chlorate. CRMs have they have high growth rates and high yields (characteristics which are more comparable to aerobic than anaerobic microbes) which should help them survive in a competitive soil community. The only byproduct of chlorate is chloride ion, resulting in no long term adverse effect of addition of chlorate to subsurface environments. It is hypothesized that simultaneous injection of chlorate and CRMs (acclimate to specific pollutants) into contaminated soils should result in high specificity of target pollutant degradation since only these targeted microorganisms will be capable of chlorate-supported growth.

CRMs are likely a subset of denitrifying microorganisms. From comparison with pollutant degradation abilities of denitrifiers, it is hypothesized chemicals degraded by chlorate reducers could include those degraded under denitrifying conditions. Chemicals selected for study that meet this criterion include: toluene, p-xylene, ethylbenzene, naphthalene, chlorophenol, chlorobenzoate, carbon tetrachloride, and phenol; two additional chemicals selected for study that are thought to be persistent under denitrifying conditions are benzene and pentane. To test our hypothesis that CRMs can be found that degrade these pollutants we will attempt to acclimate mixed cultures of CRMs to these chemicals and, if successful, demonstrate their degradative abilities in batch, chemostat, and column studies.

The physiology of microbial chlorate reduction is not well understood, and we also propose to study chlorate respiration to contrast the pathways for electron transport to chlorate with the pathways for molecular oxygen by using a series of respiratory inhibitors. In both cases, reducing equivalents enter the respiratory chain via dehydrogenases, such as NADH dehydrogenase, and are passed sequentially down a chain or carriers that can include iron-sulfur proteins, quinones, and a series of cytochromes. By inhibiting the action of these different carriers at selected points, it is possible to dissect the electron transport chain and compare components that participate in electron transport to oxygen, nitrate and chlorate.

It is envisioned that this research will show that co-injection of chlorate and chlorate+ pollutant-acclimated microbes is a viable method of selectively stimulating the degradation of target pollutants for subsurface bioremediation. Furthermore, the proposed research will also provide fundamental information on the properties of CRMs and the physiology of microbial chlorate respiration.

Meet the Graduate Students working on this project

Mr. Peter Mulvaney (ppm105@psu.edu) worked on isolating pure cultures of chlorate and perchlorate reducing bacteria (PDX and KJ1). He found that not all isolates that could respire using chlorate could degrade perchlorate.
Ms. Jun Wu (jxw310@psu.edu) evaluated the abundance of chlorate and perchlorate reducing micoorganisms in the natural environment. So found that while chlorate reducing microbes (CRMs) were relatively abundant in several systems, there are proportionately fewer perchlorate reducing microbes (PRMs) than CRMs. She also conducted soil column experiments and found that toluene degradation was enhanced in the presence of chlorate.
Mr. Hussen Zhang (hxz116@psu.edu) examined the kinetics of chlorate and perchlorate reduction by two isolates obtained in our laboraotory (KJ and PDX) in batch and chemostat cultures. He is shown here analyzing samples using an ion chromatograph.