Aerobic Degradation of Dinitrotoluenes and Pathway for Bacterial Degradation of 2,6-Dinitrotoluene

Abstract

An oxidative pathway for the mineralization of 2,4-dinitrotoluene (2,4-DNT) by Burkholderia sp. strain DNT has been reported previously. We report here the isolation of additional strains with the ability to mineralize 2,4-DNT by the same pathway and the isolation and characterization of bacterial strains that mineralize 2,6-dinitrotoluene (2,6-DNT) by a different pathway. Burkholderia cepacia strain JS850 and Hydrogenophaga palleronii strain JS863 grew on 2,6-DNT as the sole source of carbon and nitrogen. The initial steps in the pathway for degradation of 2,6-DNT were determined by simultaneous induction, enzyme assays, and identification of metabolites through mass spectroscopy and nuclear magnetic resonance. 2,6-DNT was converted to 3-methyl-4-nitrocatechol by a dioxygenation reaction accompanied by the release of nitrite. 3-Methyl-4-nitrocatechol was the substrate for extradiol ring cleavage yielding 2-hydroxy-5-nitro-6-oxohepta-2,4-dienoic acid, which was converted to 2-hydroxy-5-nitropenta-2,4-dienoic acid. 2,4-DNT-degrading strains also converted 2,6-DNT to 3-methyl-4-nitrocatechol but did not metabolize the 3-methyl-4-nitrocatechol. Although 2,6-DNT prevented the degradation of 2,4-DNT by 2,4-DNT-degrading strains, the effect was not the result of inhibition of 2,4-DNT dioxygenase by 2,6-DNT or of 4-methyl-5-nitrocatechol monooxygenase by 3-methyl-4-nitrocatechol.2,6-Dinitrotoluene (2,6-DNT) and 2,4-dinitrotoluene (2,4-DNT) occur as soil and groundwater contaminants at former 2,4,6-trinitrotoluene (TNT) production sites and in the wastewater from the commercial production of feedstocks for polyurethane foam (23). Twenty years after the cessation of TNT production in the United States, the manufacturing sites are still heavily contaminated with both 2,4-and 2,6-DNT even though 2,4-DNT-mineralizing bacteria can be readily isolated from the contaminated material (26). Commercial manufacture of DNT results in the release of DNT to industrial and municipal waste treatment systems (information found at the Environmental Health Center website [http://safety.webfirst .com/ehc/ew/chemical.htm] and in the TOXNET Toxics Release Inventory [http://six.nlm.nih.gov/sis1]). The unpredictable presence of DNT in the waste streams sent to the treatment plants can cause upsets in the ability of activated sludges to effectively remove the organic components in the waste streams (11). 2,4-and 2,6-DNT are priority pollutants (13), and industrial waste streams from DNT-manufacturing facilities are specifically regulated by the U.S. Environmental Protection Agency (40 CFR 261.32).Contaminated munitions manufacturing sites are ready sources of bacteria able to mineralize 2,4-DNT, but bacteria able to grow on 2,6-DNT have been more elusive. The bacterial pathway for degradation of 2,4-DNT (8, 28) is initiated by dioxygenation of 2,4-DNT, which results in the formation of 4-methyl-5-nitrocatechol (4M5NC) and the release of nitrite; monooxygenation of 4M5NC then yields 2-hydroxy-5-methylquinone, which is subsequently reduced to 2,4...

Environmental Transport and Fate Process Descriptors for Propellant Compounds

“…Aerobic transformation of 2,4-DNT to carbon dioxide and nitrite has been documented in soil slurries and munitions plant wastewater inoculated with known DNT-degrading microbes Spain 1995;Nishino et al 1999;Nishino et al 2000;Christopher et al 2000;Zhang et al 2000;Snellinx et al 2003), and by native microbes in DNTcontaminated soil and aquifer material (Bradley et al 1994;Bradley et al 1997). However, the persistence of 2,4-DNT in soils colonized by DNTdegrading microbes suggests slow rates of mineralization, possibly due to inhibition by 2,6-DNT or nitrite (Smets and Mueller 2001;Fortner et al 2003).…”

Aerobic biotransformation of 2,4-dinitroanisole in soil and soil Bacillus sp.

“…Bacterial species of Pseudomonas, Burkholderia, Comamonas, Arthrobacter, Acidovorax and Rhodococcus have been reported to grow on several nitroaromatic compounds including nitrobenzenes, nitrotoluenes and nitrophenols as the sole nitrogen, carbon and energy sources (Jain et al 1994;Nishino and Spain 1995;He and Spain 1999;Nishino et al 2000;Ju and Parales 2010). Rhodococcus strains AS2 and AS3 were shown to use 4-nitroanisole as the sole source of carbon and energy (Schäfer et al 1996).…”

TNT biotransformation potential of the clinical isolate of Salmonella typhimurium - potential ecological implications

“…[22,23] The level of denitration was substantially higher in static cultures than in orbital shaken cultures (data not shown). In fact, orbital agitation of TNT enriched culture was detrimental to TNT degradation, in contrary to (NH 4 ) 2 SO 4 -supplemented cultures.…”