A role of Bradyrhizobium elkanii and closely related strains in the degradation of methoxychlor in soil and surface water environments.

We have reported that a leguminous bacterial strain, Bradyrhizobium sp. strain 17-4, isolated from river sediment, phylogenetically very close to Bradyrhizobium elkanii, degraded methoxychlor through O-demethylation and oxidative dechlorination. In the present investigation, we found that B. elkanii (USDA94), a standard species deposited in the Culture Collection, degraded methoxychlor. Furthermore, Bradyrhizobium sp. strain 4-1, also very close to B. elkanii, isolated from Japanese paddy field soil, degraded methoxychlor. These B. elkanii and closely related strains degraded methoxychlor through almost identical metabolic pathways, and cleaved the phenyl ring and mineralized. In contrast, another representative Bradyrhizobium species, B. japonicum (USDA110), did not degrade methoxychlor at all. Based on these findings, B. elkanii and closely related strains are likely to play an important role not only in providing the readily biodegradable substrates but also in completely degrading (mineralizing) methoxychlor by themselves in the soil and surface water environment.

O-Demethylation and successive oxidative dechlorination of methoxychlor by Bradyrhizobium sp. strain 17-4, isolated from river sediment.

O-Demethylation of insecticide methoxychlor is well known as a phase I metabolic reaction in various eukaryotic organisms. Regarding prokaryotic organisms, however, no individual species involved in such reaction have been specified and characterized so far. Here we successfully isolated a bacterium that mediates oxidative transformation of methoxychlor, including O-demethylation and dechlorination, from river sediment. The isolate was found to be closely related to Bradyrhizobium elkanii at the 16S rRNA gene sequence level (100% identical). However, based on some differences in the physiological properties of this bacterium, we determined that it was actually a different species, Bradyrhizobium sp. strain 17-4. The isolate mediated O-demethylation of methoxychlor to yield a monophenolic derivative [Mono-OH; 1,1,1-trichloro-2-(4-hydroxyphenyl)-2-(4-methoxyphenyl)ethane] as the primary degradation product. The chiral high-performance liquid chromatography (HPLC) analysis revealed that the isolate possesses high enantioselectivity favoring the formation of (S)-Mono-OH (nearly 100%). Accompanied by the sequential O-demethylation to form the bis-phenolic derivative Bis-OH [1,1,1-trichloro-2,2-bis(4-hydroxyphenyl)ethane], oxidative dechlorination of the side chain proceeded, and monophenolic carboxylic acid accumulated, followed by the formation of multiple unidentified polar degradation products. The breakdown proceeded more rapidly when reductively dechlorinated (dichloro-form) methoxychlor was applied as the initial substrate. The resultant carboxylic acids and polar degradation products are likely further biodegraded by ubiquitous bacteria. The isolate possibly plays an important role for complete degradation (mineralization) of methoxychlor by providing the readily biodegradable substrates.

Reductive dechlorination of methoxychlor by bacterial species of environmental origin: evidence for primary biodegradation of methoxychlor in submerged environments.

Methoxychlor [1,1,1-trichloro-2,2-bis(4-methoxyphenyl)ethane] is an organochlorine insecticide that undergoes dechlorination in natural submerged environments. We investigated the ability to dechlorinate this compound in seven environmental bacterial species ( Aeromonas hydrophila , Enterobacter amnigenus , Klebsiella terrigena , Bacillus subtilis , Achromobacter xylosoxidans , Acinetobacter calcoaceticus , and Mycobacterium obuense ) and the enteric bacterium Escherichia coli as a positive control. In R2A broth at 25 °C under aerobic, static culture, all species except Ach. xylosoxidans were observed to convert methoxychlor to dechlorinated methoxychlor [1,1-dichloro-2,2-bis(4-methoxyphenyl)ethane]. The medium was aerobic at first, but bacterial growth resulted in the consumption of oxygen and generated microaerobic and weakly reductive conditions. Replacement of the headspace of the culture tubes with nitrogen gas was found to decrease the dechlorination rate. Our findings suggest that extensive bacterial species ubiquitously inhabiting the subsurface water environment play an important role in the primary dechlorination of methoxychlor.

An environmental fate study of methoxychlor using water-sediment model system.

Agricultural waste water containing pesticides can reach the sea via rivers and estuaries, including brackish lakes. We studied the metabolic fate of methoxychlor [MXC; 1,1,1-trichloro-2,2-bis(4-methoxyphenyl)ethane] in a model system consisting of sediment and associated water collected from two sampling sites: a brackish lake and a freshwater river. MXC degraded rapidly and was finally mineralized in both sediment systems. The first step of degradation was dechlorination to yield 1,1-dichloro-2,2-bis(4-methoxyphenyl)ethane [de-Cl-MXC] or CN-replacement to yield 2,2-bis(4-methoxyphenyl)acetonitrile [MXC-CN], followed by O-demethylation. Although the metabolites were common to the two sediments, the dynamics of the metabolites over time were clearly distinct. In the brackish lake sediment, de-Cl-MXC accumulated transiently, whereas in the river sediment, it was rapidly converted to its demethylated metabolite. We also found that dechlorination and CN-replacement proceeded in autoclave-sterilized river sediment. In the river sediment, the abiotic reaction mediated by abundant humic acid and low oxygen level also appeared to contribute to the overall MXC metabolism.

Bioconcentration and biotransformation of [¹4C]methoxychlor in the brackish water bivalve Corbicula japonica.

To obtain basic information on the metabolic fate of xenobiotics in the brackish water, bivalve Corbicula japonica, bioconcentration and biotransformation experiments were performed using methoxychlor (MXC) as a model compound. Bivalves were exposed to [ring-U-¹4C]MXC (10 µg L⁻¹) for 28 days under semi-static conditions followed by a 14-day depuration phase. The ¹4C concentration in the bivalves rapidly increased and reached a steady state after exposure for 7 days (BCFss = 2010); however, it rapidly decreased with a half-life of 2.2 days in the depuration phase. Mono- and bis-demethylated MXC, and their corresponding sulphate conjugates, were identified as minor metabolites. No glycoside conjugates (including glucuronide and glucoside) were detected. Despite this biotransformation system, bivalves were found to excrete retained MXC mostly unchanged although its relatively hydrophobic nature.

Bioaccumulation and metabolism of [14C]bisphenol A in the brackish water bivalve Corbicula japonica.

The brackish water bivalve mollusk Corbicula japonica was exposed to brackish water containing approximately 9 microg/l [(14)C]bisphenol A (BPA) for 168 h (the uptake phase), and subsequently transferred to clean brackish water for 144 h (the depuration phase) under semi-static conditions. Mono and disulfate conjugates of BPA were detected in the bivalves as major metabolites. At the end of the uptake phase, the visceral mass contained the highest (14)C-concentration, and the monosulfate conjugate of BPA was a major metabolite in the visceral mass. These data suggest that the visceral mass is the major tissue/organ to take up and metabolize BPA in these bivalves. The BPA concentration in the bivalves readily reached steady state during the uptake phase and immediately decreased in the depuration phase. The accumulation and elimination rates of the mono and disulfate conjugates of BPA were slower than those of BPA.