CENSUS – Chlorinated Benzenes

Detect and quantify bacteria responsible for biodegradation of Chlorinated Benzenes

Chlorinated benzenes are an important class of industrial solvents and chemical intermediates in the productions of drugs, dyes, herbicides and insecticides. The physical-chemical properties of chlorinated benzenes as well as susceptibility to biodegradation are functions of their degree of chlorination and the positions of chlorine substituents.

In general, chlorobenzenes with four or less chlorine groups are susceptible to aerobic biodegradation even serving as growth supporting substrates.

Under anaerobic conditions, reductive dechlorination of higher chlorinated benzenes including hexachlorobenzene (HCB), pentachlorobenzene (PeCB), tetrachlorobenzene (TeCB) isomers, and trichlorobenzene (TCB) isomers by halorespiring bacteria has been well documented (Field and Sierra-Alvarez 2008).

  • The dichlorobenzene (DCB) isomers and chlorobenzene (CB) were considered relatively recalcitrant under anaerobic conditions.
  • However, new evidence has demonstrated reductive dechlorination of DCBs to CB and CB to benzene (Fung et al. 2009) with corresponding increases in concentrations of Dehalobacter spp. (Nelson et al. 2011).

CENSUS Targets for Reductive Dechlorination

The following table describes the individual CENSUS targets, their importance in evaluating reductive dechlorination as a treatment mechanism.


MI Code 

  Relevance / Data Interpretation

Dehalococcoides DHC Although biodegradation of individual compounds and specific isomers does vary somewhat between isolates, Dehalococcoides such as strain CBDB1 have identified which reductively dechlorinate, hexachlorobenzene (HCB), pentachlorobenzene (PeCB), all three tetrachlorobenzene (TeCB) isomers, 1,2,3-TCB and 1,2,4-TCB (Adrian et al. 2000; Jayachandran et al. 2003).
Dehalobacter DHB While considered relatively recalcitrant under anaerobic conditions, recent work has implied Dehalobacter spp. in the reductive dechlorination of DCBs and CB (Nelson et al. 2011).
Dehalobium DECO Dehalobium chlorocoercia  DF-1 has been shown to be capable of reductive dechlorination of HCB, PeCB and 1,2,3,5-TeCB (Wu et al. 2002).
Methanogens MGN Methanogens utilize hydrogen and carbon dioxide to produce methane.  While common in the anaerobic environments conducive to reductive dechlorination, methanogens can compete with dechlorinating bacteria for available hydrogen.
Sulfate Reducing Bacteria APS This assay targets a gene found in sulfate reduction.  As with methanogens, SRBs can compete with dechlorinating bacteria for available hydrogen.
Total Bacteria EBAC This assay quantifies total bacterial biomass.


CENSUS Targets for Aerobic Biodegradation 


MI Code 

  Relevance / Data Interpretation

Phenol Hydroxylase PHE Phenol hydroxylase catalyzes the continued oxidation and in some cases, the initial oxidation of a variety of monoaromatic compounds.  In an independent study, significant increases in numbers of bacteria containing PHE genes corresponded to increases in biodegradation of DCB isomers (Dominguez et al. 2008).
Toluene Dioxygenase TOD Toluene dioxygenase has relatively relaxed substrate specificity and mediates the incorporation of both atoms of oxygen into the aromatic ring of benzene and substituted benzenes (toluene and chlorobenzene).  Comparison of TOD levels in background and source zone samples from a CB impacted site suggested that CBs promoted growth of TOD containing bacteria (Dominguez et al. 2008).
Ring hydroxylatingtoluene monooxygenase RMO Similar to PHE, ring hydroxylating monooxygenases (RMO) catalyze the initial and in some cases second oxidation of a variety of monoaromatic compounds including BTEX and CB.
Trichlorobenzene Dioxygenase TCBO The TCBO assay targets the genes encoding aromatic dioxygenases responsible for initiating aerobic biodegradation of a number of chlorinated benzenes including chlorobenzene, 1,2-dichlorobenzene, 1,2,4-trichlorobenzene, and 1,2,4,5-tetrachlorobenzene.



Abraham W-R, Wenderoth DF, GlaBer W (2005). “Diversity of biphenyl degraders in chlorobenzene polluted aquifer”. Chemosphere 58(4): 529-533.

Adrian L, Szewzyk U, Wecke J, Gorisch H (2000). “Bacterial dehalorespiration with chlorinated benzenes”. Nature 408 (6812):580-583.

Beil S, Happe B, Timmis KN, Pieper DH (1997). “Genetic and Biochemical Characterization of the Broad Spectrum Chlorobenzene Dioxygenase from Burkholderia sp. PS12- Dechlorination of 1,2,4,5-Tetrachlorobenzene”. European Journal of Biochemistry 247 (1): 190-199.

Dominguez R, da Silva M, McGuire T, Adamson D, Newell C, Alvarez P (2008). “Aerobic bioremediation of chlorobenzene source-zone soil in flow-through columns: performance assessment using quantitative PCR”. Biodegradation 19(4):545-553.
Field J, Sierra-Alvarez R(2008). “Microbial degradation of chlorinated benzenes”. Biodegradation 19(4):463-480.

Fung JM, Weisenstein BP, Mack EE, Vidumsky JE, Ei TA, Zinder SH. (2009). “Reductive Dehalogenation of Dichlorobenzenes and Monochlorobenzene to Benzene in Microcosms”. Environmental Science and Technology 43 (7):2302-2307.

Javachandran G, Gorisch H, Adrian L. (2003). “Dehalorespiration with hexachlorobenzene and pentachlorobenzene by Dehalococcoides strain CBDB1”. Archives of Microbiology 180(6):411-416.
Nelson JL, Fung JM, Cadillo-Quiroz H, Cheng X, Zinder SH. (2011). “A role for Dehalobacter spp. in the Reductive Dehalogenation of Dichlorobenzenes and Monochlorobenzene”. Environmental Science and Technology 45(16):6806-6813.

van der Meer JR, van Neerven AR, de Vries EJ, de Vos WM, Zehnder AJ. (1991). “Cloning and characterization of plasmid-encoded genes for the degradation of 1,2 dichloro-, 1,4-dichloro-, and 1,2,4-trichlorobenzene of Pseudomonas sp. strain P51″. Journal of Bacteriology 173(1):6-15.

Wu Q, Miliken CE, Meier GP, Watts JEM, Sowers KR, May HD. (2002). ” Dechlorination of Chlorobenzenes by a Culture Containing Bacterium DF-1, a PCB Dechlorinating Microorganism”. Environmental Science and Technology 36(15):3290-3294.