Distribution of glyphosate and methylphosphonate catabolism systems in soil bacteria <Emphasis Type="Italic">Ochrobactrum anthropi</Emphasis> and <Emphasis Type="Italic">Achromobacter</Emphasis> sp

Bacterial strains capable of utilizing methylphosphonic acid (MP) or glyphosate (GP) as the sole sources of phosphorus were isolated from soils contaminated with these organophosphonates. The strains isolated from MP-contaminated soils grew on MP and failed to grow on GP. One group of the isolates from GP-contaminated soils grew only on MP, while the other one grew on MP and GP. Strains Achromobacter sp. MPS 12 (VKM B-2694), MP degraders group, and Ochrobactrum anthropi GPK 3 (VKM B-2554D), GP degraders group, demonstrated the best degradative capabilities towards MP and GP, respectively, and were studied for the distribution of their organophosphonate catabolism systems. In Achromobacter sp. MPS 12, degradation of MP was catalyzed by C–P lyase incapable of degrading GP (C–P lyase I). Adaptation to growth on GP yielded the strain Achromobacter sp. MPS 12A, which retained its ability to degrade MP via C–P lyase I and was capable of degrading GP with formation of sarcosine, thus suggesting the involvement of a GP-specific C–P lyase II. O. anthropi GPK 3 also degraded MP via C–P lyase I, but degradation of GP in it was initiated by glyphosate oxidoreductase, which was followed by product transformation via the phosphonatase pathway.     

Microbial degradation of organophosphonates by soil bacteria

Bacteria that can utilize glyphosate (GP) or methylphosphonic acid (MPA) as a sole phosphorus source have been isolated from soil samples polluted with organophosphonates (OP). No matter which of these compounds was predominant in the native habitat of the strains, all of them utilized methylphosphonate. Some of the strains isolated from GP-polluted soil could utilize both phosphorus sources. Strains growing on glyphosate only were not isolated. The isolates retained high destructive activity after long-term storage of cells in lyophilized state, freezing to ?20°C, and maintenance on various media under mineral oil. When phosphorusstarved cells (with 2% phosphorus) were used as inoculum, the efficiency of OP biodegradation significantly increased (1.5-fold).     

Biodegradation of glyphosate by soil bacteria: Optimization of cultivation and the method for active biomass storage

Conditions for obtaining the active biomass of Ochrobactrum anthropi GPK 3 and Achromobacter sp. Kg 16, bacteria which are able to degrade the herbicide glyphosate (N-phosphonomethylglycine), were investigated. In the batch culture, degradation was most effective in the medium with pH 6.0–7.0 and aeration at 10–60% of air saturation supplemented with glutamate and ammonium chloride as sources of carbon and nitrogen, respectively. Due to the adaptation of the cells and induction of the relevant enzymatic systems, the inoculum grown in the presence of glyphosate exhibited 1.5–2-fold higher efficiency of xenobiotic degradation than that grown with other sources of phosphorus (orthophosphate and methylphosphonic acid). The efficiency of the toxicant decomposition increased with an increase in a specific load of glyphosate, which the cells were subjected to during the initial stage of growth. The specific load was regulated both by the initial cell concentration and the concentration of the phosphorus source, and the effect was probably determined by its availability to microorganisms. Storage of the liquid biopreparation as a paste with stabilizers (ascorbate, thiourea, and glutamate) at room temperature for 50 days resulted in high level of bacteria viability and a degrading activity approximately equal to that obtained when the bacteria were maintained on the agar medium containing glyphosate at 4°C with monthly transfers to the fresh culture medium.     

Sorption and microbial degradation of glyphosate in soil suspensions

Sorption and microbial destruction of glyphosate, the active agent of the herbicide Ground Bio, in suspensions of sod-podzol and gray forest soils has been studied. According to the adsorptive values (3560 and 8200 mg/kg, respectively) and the Freundlich constants (K_f, 15.6 and 18.7, respectively), these soils had a relatively high sorption capacity as related to the herbicide. Sorbed glyphosate is represented by extractable and bound (non-extractable) fractions. After long-term incubation of sterile suspensions, the ratio of these fractions reached 2: 1 for sod-podzol soil and 1: 1 for gray forest soil. Inoculation of a native suspension of sod-podzol soil with cells of a selected strain-degrader Ochrobactum anthropi GPK 3 resulted in a 25.4% decrease in the total glyphosate content (dissolved and extractable), whereas in a noninoculated suspension, the loss did not exceed 5.5%. The potential for the use of a selected bacterial strain in the glyphosate destruction processes in soil systems is demonstrated for the first time.     

Glyphosate bioavailability in soil

Biodegradation of glyphosate in sod-podzol soil by both the indigenous micro flora and the introduced strain Ochrobactrum anthropi GPK 3 was studied with respect to its sorption and mobility. The experiments were carried out in columns simulating the vertical soil profile. Soil samples studied were taken from soil horizons 0–10, 10–20, and 20–30 cm deep. It was found out that the most of the herbicide (up to 84%) was adsorbed by soil during the first 24 h; the rest (16%) remained in the soluble fraction. The adsorbed glyphosate was completely extractable by alkali. No irreversible binding of glyphosate was observed. By the end of the experiment (21st day), glyphosate was only found in extractable fractions. The comparison of the effect of the introduced O. anthropi GPK 3 and indigenous microbial community on the total toxicant content (both soluble and absorbed) in the upper 10 cm soil layer showed its reduction by 42% (21 mg/kg soil) and 10–12% (5 mg/kg soil), respectively. Simultaneously, 14–18% glyphosate moved to a lower 10–20 cm layer. Watering (that simulated rainfall) resulted in a 20% increase of its content at this depth; 6–8% of herbicide was further washed down to the 20–30 cm layer. The glyphosate mobility down the soil profile reduced its density in the upper layer, where it was available for biodegradation, and resulted in its concentration in lower horizons characterized by the absence (or low level) of biodegradative processes. It was shown for the first time how the herbicide biodegradation in soil can be increased manifold by introduction of the selected strain O. anthropi GPK 3.     

Bioremediation of glyphosate-contaminated soils

Based on the results of laboratory and field experiments, we performed a comprehensive assessment of the bioremediation efficiency of glyphosate-contaminated soddy-podzol soil. The selected bacterial strains Achromobacter sp. Kg 16 (VKM B-2534D) and Ochrobactrum anthropi GPK 3 (VKM B-2554D) were used for the aerobic degradation of glyphosate. They demonstrated high viability in soil with the tenfold higher content of glyphosate than the recommended dose for the single in situ treatment of weeds. The strains provided a two- to threefold higher rate of glyphosate degradation as compared to indigenous soil microbial community. Within 1–2 weeks after the strain introduction, the glyphosate content of the treated soil decreased and integral toxicity and phytotoxicity diminished to values of non-contaminated soil. The decrease in the glyphosate content restored soil biological activity, as is evident from a more than twofold increase in the dehydrogenase activity of indigenous soil microorganisms and their biomass (1.2-fold and 1.6-fold for saprotrophic bacteria and fungi, respectively). The glyphosate-degrading strains used in this study are not pathogenic for mammals and do not exhibit integral toxicity and phytotoxicity. Therefore, these strains are suitable for the efficient, ecologically safe, and rapid bioremediation of glyphosate-contaminated soils.     

Biodegradation of Organophosphorus Pollutants by Soil Bacteria: Biochemical Aspects and Unsolved Problems

Applied Biochemistry and Microbiology - The degradation of stable organophosphorus pollutants has been studied in six soil bacterial isolates and three strains of bacteria adapted to utilize...