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

Abstract

In previous work, we found that an anaerobic sludge efficiently degraded hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), but the role of isolates in the degradation process was unknown. Recently, we isolated a facultatively anaerobic bacterium, identified as Klebsiella pneumoniae strain SCZ-1, using MIDI and the 16S rRNA method from this sludge and employed it to degrade RDX. Strain SCZ-1 degraded RDX to formaldehyde (HCHO), methanol (CH 3 OH) (12% of total C), carbon dioxide (CO 2 ) (72% of total C), and nitrous oxide (N 2 O) (60% of total N) through intermediary formation of methylenedinitramine (O 2 NNHCH 2 NHNO 2 ). Likewise, hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine (MNX) was degraded to HCHO, CH 3 ). These findings suggested the possible involvement of a common initial reaction, possibly denitration, followed by ring cleavage and decomposition in water. The trace amounts of MNX detected during RDX degradation and the trace amounts of hexahydro-1,3-dinitroso-5-nitro-1,3,5-triazine detected during MNX degradation suggested that another minor degradation pathway was also present that reduced ONO 2 groups to the corresponding ONO groups.Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) is a widely used explosive that severely contaminates soil and groundwater (13,22). RDX has been found to be toxic in various terrestrial and aquatic environments (26,29). The toxicity of cyclic nitramines means that contaminated soil and groundwater must be remediated, preferably by bioremediation. Biodegradation of RDX with anaerobic sludge was extensively studied by McCormick et al. (21) and Kaplan (19), who proposed a pathway based on the sequential reduction of RDX to hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine (MNX), hexahydro-1,3-dinitroso-5-nitro-1,3,5-triazine (DNX), and hexahydro-1,3,5-trinitroso-1,3,5-triazine (TNX). It was proposed that the nitroso compounds undergo further transformation to hydroxylamino derivatives (HOHN-RDX), which are subsequently cleaved to produce formaldehyde (HCHO), methanol (CH 3 OH), hydrazine (NH 2 NH 2 ), and dimethyl hydrazine [(H 3 C) 2 NNH 2 ].Other research groups have employed mixed anaerobic microbial cultures that have included methanogens (1, 2), acetogens (4), nitrate reducers (12), and individual isolates such as isolates of Desulfovibrio sp. and Serratia marcescens (31) to biotransform RDX, but in most cases no clear details concerning ring cleavage products or mineralization were provided in the descriptions. Subsequent work in our laboratory (14,15,16) showed that RDX could be degraded in anaerobic sludge via intermediary formation of methylenedinitramine (O 2 NNHCH 2 NHNO 2 ), which decomposes in water to nitrous oxide (N 2 O) and HCHO. Recently, Oh et al. (23) confirmed the formation of O 2 NNHCH 2 NHNO 2 during RDX degradation with anaerobic sludge. Despite these previous efforts, there is still insufficient information regarding the roles of individual anaerobic isolates and their products during RDX degradation. Without these details, particularly details concerning...

Involvement of cytochrome c CymA in the anaerobic metabolism of RDX by Shewanella oneidensis MR-1

“…Chromatograms were taken at a wavelength of 225 nm. HCHO was analyzed as its derivative with 2,4-pentanedione (Zhao et al 2002). N 2 O was analyzed in the headspace of the samples using an Agilent 6890 gas chromatograph equipped with an electron capture detector (ECD).…”

“…Protein concentration was determined using the bicinchoninic acid method by following the instruction of the BCA Protein Assay kit (Pierce, Rockford, Illinois). Some microcosms were spiked with L-[U-14 C]RDX (0.038 µCi (1 Ci = 37 Gbq)) to measure mineralization (liberated 14 CO 2 ) with a Tri-Carb 4530 liquid scintillation counter (model 2100 TR; Packard Instrument Company) as described previously (Zhao et al 2002).…”

The ways to increase efficiency of soil bioremediation

“…The strains of Dechloromonas (β-proteobacteria) completely oxidize benzene under anaerobic conditions, using nitrate as an electron acceptor [31]. Moreover, in the studies by Zhao et al [32], Klebsiella pneumoniae strain SCZ-1, a facultative anaerobic microorganism isolated from domestic anaerobic sludge, was used to successfully degrade hexahydro-1,3,5-trinitrio-1,3,5 triazine (RDX) and its mono-nitroso derivative (MNX) under aerobic conditions. The strain SCZ-1 degraded RDX to HCHO, CH 3 OH (12%), CO 2 (72%) and N 2 O (60%) through the intermediary formation of methylenedinitramine.…”

Molasses enhanced phyto and bioremediation treatability study of explosives contaminated Hawaiian soils

“…Various microorganisms isolated from RDX contaminated soil were able to degrade RDX [12,13]. The degradation of RDX can occur via the two-electron reductive pathway (Nitroso route) and via denitration [14,15]. Enterobacteria, and Escherichia coli are some of the bacteria that can degrade RDX using the reductive degradation pathway.…”

“…The denitration pathway can be aerobic or anaerobic. Bacteria like Klebsiella pneumoniae can denitrate RDX anaerobically [15] and Williamsia, and Gordonia can denitrate RDX aerobically [16].…”