CENSUS-Dioxane Metabolism

An increasing number of microorganisms have been isolated that utilize dioxane as a growth supporting substrate under aerobic conditions suggesting biodegradation is a viable attenuation mechanism. In aerobic dioxane utilizing bacteria including Pseudonocardia dioxanivorans CB1190, the first step in dioxane metabolism is mediated by a dioxane monooxygenase (DXMO). A crucial difference in the metabolism vs cometabolism of dioxane is processing of the resulting intermediates. In bacteria like CB1190 that can utilize dioxane as a growth supporting substrate, a key step is mediated by an aldehyde dehydrogenase enzyme (ALDH).

Therefore, qPCR assays have been developed to quantify the DXMO and ALDH genes to evaluate the potential for dioxane metabolism in environmental samples (Gedalanga et al. 2014).


MI Code 

  Relevance / Data Interpretation

Dioxane Monooxygenase
DXMO Initiates aerobic metabolism of 1,4-dioxane by

P. dioxanivorans CB1190 and other dioxane utilizing bacteria

Aldehyde dehydrogenase ALDH Catalyzes a key step in metabolism of a dioxane breakdown product.


CENSUS – Dioxane Cometabolism

Under aerobic conditions, dioxane is also amenable to cometabolism by several groups of organisms expressing monooxygenase genes for the metabolism of a variety of primary substrates. To date, cometabolic transformation of dioxane has been observed for monooxygenase-expressing bacteria utilizing methane, propane and other n-alkanes, tetrahydrofuran, and toluene as growth supporting substrates (Lan et al. 2013; Mahendra and Alvarez-Cohen 2006; Masuda et al. 2012; Vainberg et al. 2006). More specifically, methane oxidizing bacteria producing soluble methane monooxygenase (sMMO) have been shown to be capable of cometabolism of dioxane while those expressing particulate methane monooxygenase do not appear to co-oxidize dioxane. Likewise, the ability to cometabolize dioxane utilizing toluene as a growth supporting substrate appears to be pathway dependent. Bacterial strains expressing toluene-2- and toluene-4-monooxygenases which attack at the ring structure (ring-hydroxylating monooxygenases [RMO and RDEG]) can co-oxidize dioxane. Conversely, organisms initiating toluene metabolism via toluene dioxygenase (TOD) or side chain oxidation (TOL) do not appear to be capable of dioxane cometabolism (Mahendra and Alvarez-Cohen 2006).

The following table describes individual CENSUS targets and their importance in evaluating aerobic cometabolism as a treatment mechanism, and provides guidelines for integrating CENSUS results into routine groundwater monitoring for common corrective actions.


MI Code 

  Relevance / Data Interpretation

Soluble Methane Monooxygenase sMMO When expressed, sMMO is capable of co-oxidation of dioxane. Particulate methane monooxygenase however does not appear to co-oxidize dioxane.
Propane Monooxygenase PPO With addition of propane as a growth supporting substrate, aerobic propane utilizing bacteria are capable of co-oxidation of dioxane.
Ring Hydroxylating Toluene Monooxygenase RMO When expressed, a group of related toluene and phenol monooxygenases (RMO, RDEG, PHE) that perform the first and/or second step in aerobic biodegradation of BTEX also co-oxidize dioxane. More specifically, RMO quantifies a subfamily of toluene-3- and toluene-4-monooxygenase genes.
Ring Hydroxylating Toluene Monooxygenase RDEG RDEG targets groups of toluene-2-monoxygenase and phenol hydroxylase genes.
Phenol Hydroxylase PHE Targets phenol hydroxylase genes and benzene monoxygenase genes which catalyze both oxidation steps in BTEX metabolism.

Stable Isotope Probing

Stable Isotope Probing (SIP) may also be a viable option for evaluating biodegradation of dioxane. SIP is an innovative method to track the environmental fate of a “13C-labeled” contaminant of concern such as dioxane to unambiguously demonstrate biodegradation in the field. The label serves as a tracer which can be detected in the end products of biodegradation (biomass and CO2 or dissolved inorganic carbon).


Gedalanga, P.B., P. Pornwongthong, R. Mora, S.-Y.D. Chiang, B. Baldwin, D. Ogles, and S. Mahendra. 2014. Identification of Biomarker Genes To Predict Biodegradation of 1,4-Dioxane. Applied and Environmental Microbiology 80 no. 10: 3209-3218.

Lan, R.S., C.A. Smith, and M.R. Hyman. 2013. Oxidation of Cyclic Ethers by Alkane-Grown Mycobacterium vaccae JOB5. Remediation Journal 23 no. 4: 23-42.

Mahendra, S., and L. Alvarez-Cohen. 2006. Kinetics of 1,4-Dioxane Biodegradation by Monooxygenase-Expressing Bacteria. Environmental Science & Technology 40 no. 17: 5435-5442.

Masuda, H., K. McClay, R.J. Steffan, and G.J. Zylstra. 2012. Biodegradation of Tetrahydrofuran and 1,4-Dioxane by Soluble Diiron Monooxygenase in <b><i>Pseudonocardia</i></b> sp. Strain ENV478. Journal of Molecular Microbiology and Biotechnology 22 no. 5: 312-316.

Vainberg, S., K. McClay, H. Masuda, D. Root, C. Condee, G.J. Zylstra, and R.J. Steffan. 2006. Biodegradation of Ether Pollutants by Pseudonocardia sp. Strain ENV478. Applied and Environmental Microbiology 72 no. 8: 5218-5224.