Abstract
The transformation of explosives, including hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), by xenobiotic reductases XenA and XenB (and the bacterial strains harboring these enzymes) under both aerobic and anaerobic conditions was assessed. Under anaerobic conditions, Pseudomonas fluorescens I-C (XenB) degraded RDX faster than Pseudomonas putida II-B (XenA), and transformation occurred when the cells were supplied with sources of both carbon (succinate) and nitrogen (NH4 +), but not when only carbon was supplied. Transformation was always faster under anaerobic conditions compared to aerobic conditions, with both enzymes exhibiting a O2 concentration-dependent inhibition of RDX transformation. The primary degradation pathway for RDX was conversion to methylenedinitramine and then to formaldehyde, but a minor pathway that produced 4-nitro-2,4-diazabutanal (NDAB) also appeared to be active during transformation by whole cells of P. putida II-B and purified XenA. Both XenA and XenB also degraded the related nitramine explosives octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine and 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaazaisowurtzitane. Purified XenB was found to have a broader substrate range than XenA, degrading more of the explosive compounds examined in this study. The results show that these two xenobiotic reductases (and their respective bacterial strains) have the capacity to transform RDX as well as a wide variety of explosive compounds, especially under low oxygen concentrations.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Adrian NR, Arnett CM (2004) Anaerobic biodegradation of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX) by Acetobacterium malicum strain HAAP-1 isolated from a methanogenic mixed culture. Curr Microbiol 48:332–340. doi:https://doi.org/10.1007/s00284-003-4156-8
Adrian NR, Arnett CM (2006) Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) serves as a carbon and energy source for a mixed culture under anaerobic conditions. Curr Microbiol 53:129–134. doi:https://doi.org/10.1007/s00284-005-0348-8
Adrian N and Sutherland K (1999) RDX biodegradation by a methanogenic enrichment culture obtained from an explosives manufacturing wastewater treatment plant. U.S. Army Corps of Engineers, Construction Engineering Research Laboratories. 99/15
Adrian NR, Arnett CM, Hickey RF (2003) Stimulating the anaerobic biodegradation of explosives by the addition of hydrogen or electron donors that produce hydrogen. Water Res 37:3499–3507. doi:https://doi.org/10.1016/S0043-1354(03)00240-9
Balakrishnan VK, Monteil-Rivera F, Gautier MA, Hawari J (2004) Sorption and stability of the polycyclic nitramine explosive CL-20 in soil. J Environ Qual 33:1362–1368
Bhatt M, Zhao J-S, Monteil-Rivera F, Hawari J (2005) Biodegradation of cyclic nitramines by tropical marine sediment bacteria. J Indust Microbiol Biotechnol 32:261–267. doi:https://doi.org/10.1007/s10295-005-0239-9
Bhushan B, Halasz A, Spain JC, Hawari J (2002) Diaphorase catalyzed biotransformation of RDX via N-denitration mechanism. Biochem Biophys Res Commun 296:779–784
Bhushan B, Paquet L, Spain JC, Hawari J (2003) Biotransformation of 2,4,6,8,10,12-hexanitro-2,3,6,8,10,12-hexaazaisowurtzitane (CL-20) by denitrifying Pseudomonas sp. strain FA1. Appl Environ Microbiol 69:5216–5221. doi:https://doi.org/10.1128/AEM.69.9.5216-5221.2003
Bhushan B, Halasz A, Hawari J (2004a) Nitroreductase catalyzed biotransformation of CL-20. Biochem Biophys Res Commun 322:271–276. doi:https://doi.org/10.1016/j.bbrc.2004.07.115
Bhushan B, Halasz A, Spain JC, Hawari J (2004b) Initial reaction(s) in biotransformation of CL-20 is catalyzed by salicylate 1-monooxygenase from Pseudomonas sp. strain ATCC 29352. Appl Environ Microbiol 70:4040–4047. doi:https://doi.org/10.1128/AEM.70.7.4040-4047.2004
Binks PR, Nicklin S, Bruce NC (1995) Degradation of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX) by Stenotrophomonas maltophilia PB1. Appl Environ Microbiol 61:1318–1322
Blehert DS, Knoke KL, Fox BG, Chambliss GH (1997) Regioselectivity of nitroglycerin denitration by flavoprotein nitroester reductases purified from two Pseudomonas species. J Bacteriol 179:6912–6920
Blehert DS, Fox BG, Chambliss GH (1999) Cloning and sequence analysis of two Pseudomonas flavoprotein xenobiotic reductases. J Bacteriol 181:6254–6263
Boopathy R, Gurgas M, Ullian J, Manning JF (1998) Metabolism of explosive compounds by sulfate-reducing bacteria. Curr Microbiol 37:127–131. doi:https://doi.org/10.1007/s002849900350
Chang H-W, Kahng H-Y, Kim S-I, Chun J-W, Oh K-H (2004) Characterization of Pseudomonas sp. HK-6 cells responding to explosive RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine). Appl Microbiol Biotechnol 65:323–329. doi:https://doi.org/10.1007/s00253-004-1556-z
Clausen J, Robb J, Curry D, Korte N (2004) A case study of contamination on military ranges: Camp Edwards, Massachusetts, USA. Environ Pollut 129:13–21. doi:https://doi.org/10.1016/j.envpol.2003.10.002
Coleman NV, Nelson DR, Duxbury T (1998) Aerobic biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) as a nitrogen source by a Rhodococcus sp., strain DN22. Soil Biol Biochem 30:1159–1167. doi:https://doi.org/10.1016/S0038-0717(97)00172-7
Crocker FH, Thompson KT, Szecsody JE, Fredrickson HL (2005) Biotic and abiotic degradation of CL-20 and RDX in soils. J Environ Qual 34:2208–2216
Hareland WA, Crawford RL, Chapman PJ, Dagley S (1975) Metabolic function and properties of 4-hydroxyphenylacetic acid 1-hydroxylase from Pseudomonas acidovorans. J Bacteriol 121:272–285
Hawari J (2004) Microbial Degradation of RDX and HMX (Project ER-1213). Strategic Environmental Research and Development Program (SERDP)—Final Report (http://www.serdp.org/Research/upload/CU-1213-FR-01.pdf)
Hawari J, Beaudet S, Halasz A, Thiboutot S, Ampleman G (2000a) Microbial degradation of explosives: biotransformation versus mineralization. Appl Microbiol Biotechnol 54:605–618. doi:https://doi.org/10.1007/s002530000445
Hawari J, Halasz A, Sheremata T, Beaudet S, Groom C, Paquet L, Rhofir C, Ampleman G, Thiboutot S (2000b) Characterization of metabolites during biodegradation of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX) with municipal anaerobic sludge. Appl Environ Microbiol 66:2652–2657
Hawari J, Halasz A, Beaudet S, Paquet L, Ampleman G, Thiboutot S (2001) Biotransformation routes of octahydro-1, 3, 5, 7-tetranitro-1, 3, 5, 7-tetrazocine by municipal anaerobic sludge. Environ Sci Technol 35:70–75. doi:https://doi.org/10.1021/es0013531
Holmes RM, Aminot A, Kérouel R, Hooker BA, Peterson BJ (1999) A simple and precise method for measuring ammonium in marine and freshwater ecosystems. Can J Fish Aquat Sci 56:1801–1808
Jackson RG, Rylott EL, Fournier D, Hawari J, Bruce NC (2007) Exploring the biochemical properties and remediation applications of the unusual explosive-degrading P450 system XplA/B. Proc Nat Acad Sci 104:16822–16827
Jenkins TF, Pennington JC, Ranney TA, Berry TE, Jr., Miyares PH, Walsh ME, Hewitt AD, Perron NM, Parker LV, Hayes CA, Wahlgren EG (2001) Characterization of explosives contamination at military firing ranges. U.S. Army Corps of Engineers, Engineer Research and Development Center. ERDC TR-01-5
Kitts CL, Cunningham DP, Unkefer PJ (1994) Isolation of three hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine-degrading species of the family Enterobacteriaceae from nitramine explosive contaminated soil. Appl Environ Microbiol 60:4608–4611
Kitts CL, Green CE, Otley RA, Alvarez MA, Unkefer PJ (2000) Type I nitroreductase in soil enterobacteria reduce TNT (2, 4, 6-trinitrotoluene) and RDX (hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine). Can J Microbiol 46:278–282
Maloney SW, Adrian NR, Hickey RF, Heine RL (2002) Anaerobic treatment of pinkwater in a fluidized bed reactor containing GAC. J Hazard Mater 92:77–88. doi:https://doi.org/10.1016/S0304-3894(01)00375-2
McCormick NG, Cornell JH, Kaplan AM (1981) Biodegradation of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine. Appl Environ Microbiol 42:817–823
Monteil-Rivera F, Paquet L, Deschamps S, Balakrishnan VK, Beaulieu C, Hawari J (2004) Physico-chemical measurements of CL-20 for environmental applications: Comparison with RDX and HMX. J Chromatogr A 1025:125–132. doi:https://doi.org/10.1016/j.chroma.2003.08.060
Nejidat A, Kafka L, Tekoah Y, Ronen Z (2008) Effect of organic and inorganic nitrogenous compounds on RDX degradation and cytochrome P-450 expression in Rhodococcus strain YH1. Biodegradation 19:313–320. doi:https://doi.org/10.1007/s10532-007-9137-3
Nishino SF, Spain JC, Lenke H, Knackmuss H-J (1999) Mineralization of 2, 4- and 2, 6-dinitrotoluene in soil slurries. Environ Sci Technol 33:1060–1064. doi:https://doi.org/10.1021/es9808301
Nishino SF, Paoli GC, Spain JC (2000) Aerobic degradation of dinitrotoluenes and pathway for bacterial degradation of 2, 6-dinitrotoluene. Appl Environ Microbiol 66:21239–22147
Pak JW, Knoke KL, Noguera DR, Fox BG, Chambliss GH (2000) Transformation of 2, 4, 6-trinitrotoluene by purified xenobiotic reductase B from Pseudomonas fluorescens I-C. Appl Environ Microbiol 66:4742–4750
Pennington JC, Jenkins TF, Ampleman G, Thiboutot S, Brannon JM, Hewitt AD, Lewis J, Brochu S, Diaz E, Walsh MR, Walsh ME, Taylor S, Lynch JC, Clausen J, Ranney TA, Ramsey CA, Hayes CA, Grant CL, Collins CM, Bigl SR, Yost SL, Dontsova KM (2006) Distribution and fate of energetics on DoD test and training ranges: Final report. U.S. Army Engineer Research and Development Center, Environmental Laboratory. ERDC TR-06-13
Pudge IB, Daugulis AJ, Dubois C (2003) The use of Enterobacter cloacae ATCC 43560 in the development of a two-phase partitioning bioreactor for the destruction of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX). J Biotechnol 100:65–75. doi:https://doi.org/10.1016/S0168-1656(02)00229-8
Seth-Smith HMB, Edwards J, Rosser SJ, Rathbone DA, Bruce NC (2008) The explosive-degrading cytochrome P450 system is highly conserved among strains of Rhodococcus spp. Appl Environ Microbiol 74:4550–4552
Thompson KT, Crocker FH, Fredrickson HL (2005) Mineralization of the cyclic nitramine explosive hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine by Gordonia and Williamsia spp. Appl Environ Microbiol 71:8265–8272. doi:https://doi.org/10.1128/AEM.71.12.8265-8272.2005
Tipton DK, Rolston DE, Scow KM (2003) Transport and biodegradation of perchlorate in soils. J Environ Qual 32:40–46
Trott S, Nishino SF, Hawari J, Spain JC (2003) Biodegradation of the nitramine explosive CL-20. Appl Environ Microbiol 69:1871–1874. doi:https://doi.org/10.1128/AEM.69.3.1871-1874.2003
Yamamoto H, Morley MC, Speitel GE Jr, Clausen J (2004) Fate and transport of high explosives in a sandy soil: adsorption and desorption. Soil Sediment Contam 13:459–477. doi:https://doi.org/10.1080/10588330490500419
Zhang C, Hughes JB (2003) Biodegradation pathways of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX) by Clostridium acetobutylicum cell-free extract. Chemosphere 50:665–671. doi:https://doi.org/10.1016/S0045-6535(02)00639-2
Zhao J-S, Halasz A, Paquet L, Beaulieu C, Hawari J (2002) Biodegradation of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine and its mononitroso derivative hexahydro-1-nitroso-3, 5-dinitro-1, 3, 5-triazine by Klebsiella pneumoniae strain SCZ-1 isolated from an anaerobic sludge. Appl Environ Microbiol 68:5336–5341. doi:https://doi.org/10.1128/AEM.68.11.5336-5341.2002
Zhao J-S, Spain J, Hawari J (2003) Phylogenetic and metabolic diversity of hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX)-transforming bacteria in strictly anaerobic mixed cultures enriched on RDX as a nitrogen source. FEMS Microbiol Ecol 46:189–196. doi:https://doi.org/10.1016/S0168-6496(03)00216-2
Acknowledgments
The investigators acknowledge and thank the Strategic Environmental Research and Development Program (Mark Fuller, P.I., Project ER-1378) and NSF MCB-0316232 (Brian Fox., P.I.) for support of this research. Views, opinions, and/or findings contained herein are those of the authors and should not be construed as an official department of the army position or decision unless so designated by other official documentation. Thomas E. Malone was a trainee of the NIH Institutional Biotechnology Pre-Doctoral Training Grant T32 GM08349.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Fuller, M.E., McClay, K., Hawari, J. et al. Transformation of RDX and other energetic compounds by xenobiotic reductases XenA and XenB. Appl Microbiol Biotechnol 84, 535–544 (2009). https://doi.org/10.1007/s00253-009-2024-6
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-009-2024-6