Isolation and characterization of indigenous bacterial assemblage for biodegradation of persistent herbicides in the soil.

Extensive pesticides (herbicides) use is negatively disturbing the environment and humans. Pesticide bioremediation with eco-friendly techniques bears prime importance. This study aimed to isolate and characterize three different herbicides (metribuzin, clodinafop- propargyl, MCPA (2-methyl, 4 chlorophenoxyacetic acids) and Bromoxynil) degrading bacterial strains from agricultural fields of Punjab University, Pakistan. Among the 12 bacterial isolates, 5 were metribuzin degrading, 3 were clodinafop propargyl degrading and, 4 were MCPA and Bromoxynil degrading bacteria. Morphological, microscopic, and molecular characterization revealed that the majority of these bacterial strains were gram-negative and belonged to Bacillus and Pseudomonas genera. The isolates A6, B3, and C1 were subjected to respective herbicide degradation and the data was confirmed through GC-MS analysis. The effect of herbicide concentrations, pH, and temperature on bacterial growth was determined at OD600. The strain A6 degraded 14.8% metribuzin out of the provided concentration of 50 ppm by following the deamination pathway. While the isolates B3 and C1 degraded 23.2% and 33.9% clodinafop, MCPA and bromo-xynil, respectively, at a spiking concentration of 50ppm. The clodinafop, MCPA and Bromoxynil were metabolized into less toxic products i.e., dicarboxylic acids and 2-methyl phenol respectively, and metabolized via decarboxylation and dehalogenation mechanism. The present study evaluates the herbicides degrading bacterial strains that could potentially be used for bioremediation of agricultural contaminated sites.

Survey of organochlorine-tolerant culturable mycota from contaminated soils, and 2,4-D removal ability of Penicillium species in synthetic wastewater.

Agrochemical wastewater, which is produced by the extensive use of herbicides, has become a serious environmental pollutant. In this study, culturable mycota were isolated from soils contaminated with herbicides 2,4-dichlorophenoxyacetic acid (2,4-D) and 4-chloro-2-methylphenoxyacetic acid (MCPA), and their ability to tolerate and remove 2,4-D was assessed. The mycota were isolated on solid medium supplemented with 10 mmol L-1 of MCPA or 2,4-D. Tolerance and removal assays were performed in synthetic wastewater, and removal was quantified by HPLC-UV and MS/MS. Fusarium spp., Aspergillus spp., and Penicillium spp. were the most frequently isolated genera. Six Penicillium strains were able to tolerate up to 25 mmol L-1 of 2,4-D. Within this group, two P. crustosum strains (RCP4 and RCP13) degraded more than 50% of the 2,4-D in the medium during the first 7 days of incubation. Removal percentages reached 54% for RCP4 and 75% for RCP13 after 14 days. These two strains, therefore, could potentially be considered for the design of bioaugmentation strategies aimed at reducing contamination by 2,4-D in wastewater.

Spatial Control of Microbial Pesticide Degradation in Soil: A Model-Based Scenario Analysis.

Microbial pesticide degraders are heterogeneously distributed in soil. Their spatial aggregation at the millimeter scale reduces the frequency of degrader-pesticide encounter and can introduce transport limitations to pesticide degradation. We simulated reactive pesticide transport in soil to investigate the fate of the widely used herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) in response to differently aggregated distributions of degrading microbes. Four scenarios were defined covering millimeter scale heterogeneity from homogeneous (pseudo-1D) to extremely heterogeneous degrader distributions and two precipitation scenarios with either continuous light rain or heavy rain events. Leaching from subsoils did not occur in any scenario. Within the topsoil, increasing spatial heterogeneity of microbial degraders reduced macroscopic degradation rates, increased MCPA leaching, and prolonged the persistence of residual MCPA. In heterogeneous scenarios, pesticide degradation was limited by the spatial separation of degrader and pesticide, which was quantified by the spatial covariance between MCPA and degraders. Heavy rain events temporarily lifted these transport constraints in heterogeneous scenarios and increased degradation rates. Our results indicate that the mild millimeter scale spatial heterogeneity of degraders typical for arable topsoil will have negligible consequences for the fate of MCPA, but strong clustering of degraders can delay pesticide degradation.

The efficient persistence and migration of Cupriavidus gilardii T1 contribute to the removal of MCPA in laboratory and field soils.

The application of exogenous biodegradation strains in pesticide-polluted soils encounters the challenges of migration and persistence of inoculants. In this study, the degradation characteristics, vertical migration capacity, and microbial ecological risk assessment of an enhanced green fluorescent protein (EGFP)-tagged 2-Methyl-4-chlorophenoxyacetic acid (MCPA)-degrading strain Cupriavidus gilardii T1 (EGFP) were investigated in the laboratory and field soils. The optimum remediation conditions for T1 (EGFP) was characterized in soils. Meanwhile, leaching experiments showed that T1 (EGFP) migrated vertically downwards in soil and contribute to the degradation of MCPA at different depths. After inoculation with T1 (EGFP), a high expression levels of EGFP gene was observed at 28 d in the laboratory soil and at 45 d in the field soil. The degradation rates of MCPA were ≥ 60% in the laboratory soil and ≥ 48% in the field soil, indicating that T1 (EGFP) can efficiently and continuously remove MCPA in both laboratory and field conditions. In addition, the inoculation of T1 (EGFP) not only showed no significant impact on the soil microbial community structure but also can alleviate the negative effects induced by MCPA to some extent. Overall, our findings suggested that T1 (EGFP) strain is an ecologically safe resource for the in situ bioremediation of MCPA-contaminated soils.

Characterization of Opposing Responses to Phenol by Bacillus subtilis Chemoreceptors.

Bacillus subtilis employs 10 chemoreceptors to move in response to chemicals in its environment. While the sensing mechanisms have been determined for many attractants, little is known about the sensing mechanisms for repellents. In this work, we investigated phenol chemotaxis in B. subtilis. Phenol is an attractant at low, micromolar concentrations and a repellent at high, millimolar concentrations. McpA was found to be the principal chemoreceptor governing the repellent response to phenol and other related aromatic compounds. In addition, the chemoreceptors McpC and HemAT were found to govern the attractant response to phenol and related compounds. Using chemoreceptor chimeras, McpA was found to sense phenol using its signaling domain rather than its sensing domain. These observations were substantiated in vitro, where direct binding of phenol to the signaling domain of McpA was observed using saturation transfer difference nuclear magnetic resonance. These results further advance our understanding of B. subtilis chemotaxis and further demonstrate that the signaling domain of B. subtilis chemoreceptors can directly sense chemoeffectors. IMPORTANCE Bacterial chemotaxis is commonly thought to employ a sensing mechanism involving the extracellular sensing domain of chemoreceptors. Some ligands, however, appear to be sensed by the signaling domain. Phenolic compounds, commonly found in soil and root exudates, provide environmental cues for soil microbes like Bacillus subtilis. We show that phenol is sensed as both an attractant and a repellent. While the mechanism for sensing phenol as an attractant is still unknown, we found that phenol is sensed as a repellent by the signaling domain of the chemoreceptor McpA. This study furthers our understanding of the unconventional sensing mechanisms employed by the B. subtilis chemotaxis pathway.

Efficient Degradation of Phenoxyalkanoic Acid Herbicides by the Alkali-Tolerant Cupriavidus oxalaticus Strain X32.

Phenoxyalkanoic acid (PAA) herbicides are mainly metabolized by microorganisms in soils, but the degraders that perform well under alkaline environments are rarely considered. Herein, we report Cupriavidus oxalaticus strain X32, which showed encouraging PAA-degradation abilities, PAA tolerance, and alkali tolerance. In liquid media, without the addition of exogenous carbon sources, X32 could completely remove 500 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) or 4-chloro-2-methylphenoxyacetic acid within 3 days, faster than that with the model degrader Cupriavidus necator JMP134. Particularly, X32 still functioned at pH 10.5. Of note, with X32 inoculation, we observed 2,4-D degradation in soils and diminished phytotoxicity to maize (Zea mays). Furthermore, potential mechanisms underlying PAA biodegradation and alkali tolerance were then analyzed by whole-genome sequencing. Three modules of tfd gene clusters involved in 2,4-D catabolism and genes encoding monovalent cation/proton antiporters involved in alkali tolerance were putatively identified. Thus, X32 could be a promising candidate for the bioremediation of PAA-contaminated sites, especially in alkaline surroundings.

Occurrence and transformation of phenoxy acids in aquatic environment and photochemical methods of their removal: a review.

The article presents the behavior of phenoxy acids in water, the levels in aquatic ecosystems, and their transformations in the water environment. Phenoxy acids are highly soluble in water and weakly absorbed in soil. These highly mobile compounds are readily transported to surface and groundwater. Monitoring studies conducted in Europe and in other parts of the world indicate that the predominant phenoxy acids in the aquatic environment are mecoprop, 4-chloro-2-methylphenoxyacetic acid (MCPA), dichlorprop, 2,4-dichlorophenoxyacetic acid (2,4-D), and their metabolites which are chlorophenol derivatives. In water, the concentrations of phenoxy acids are effectively lowered by hydrolysis, biodegradation, and photodegradation, and a key role is played by microbial decomposition. This process is determined by the qualitative and quantitative composition of microorganisms, oxygen levels in water, and the properties and concentrations of phenoxy acids. In shallow and highly insolated waters, phenoxy acids can be decomposed mainly by photodegradation whose efficiency is determined by the form of the degraded compound. Numerous studies are underway on the use of advanced oxidation processes (AOPs) to remove phenoxy acids. The efficiency of phenoxy acid degradation using AOPs varies depending on the choice of oxidizing system and the conditions optimizing the oxidation process. Most often, methods combining UV radiation with other reagents are used to oxidize phenoxy acids. It has been found that this solution is more effective compared with the oxidation process carried out using only UV.

Hybrid electrochemical and biological treatment of herbicidal ionic liquids comprising the MCPA anion.

The present study was focused on the application of an electrochemical oxidation process combined with biodegradation for the removal of novel Herbicidal Ionic Liquids (HILs) -promising protection plant products which incorporate herbicidal anions and ammonium cations. The influence of carbon chain length (n = 8, 10, 12, 14, 16, 18) in the dialkyldimethylammonium cations on electrochemical oxidation kinetics, degradation efficiency and biodegradation by activated sludge was investigated. It was established that the applied cation influenced the heterogeneous rate constant and diffusion coefficient of electrochemical oxidation. The oxidation efficiency ranged from 17% in case of HILs with C8 alkyl chain to approx. 60% in case of HILs comprising C14 and C16 alkyl chains after 3 h of electrochemical treatment. Subsequent biodegradation studies revealed that electrochemical oxidation improved the mineralization efficiency of the studied HILs. The mineralization efficiency of electrochemically-treated HILs ranged from 28% in case of HILs comprising the C8 alkyl chain to 57% in case of HILs with C14 and C16 alkyl chains after 28 days. In case of untreated HILs, the corresponding mineralization efficiency ranged from 0 to 8%, respectively. This confirms the feasibility of a hybrid electrochemical-biological approach for treatment of herbicidal ionic liquids based on MCPA.

Transformation, CO2 formation and uptake of four organic micropollutants by carrier-attached microorganisms.

A tiered process was developed to assess the transformation, CO2 formation and uptake of four organic micropollutants by carrier-attached microorganisms from two municipal wastewater treatment plants. At the first tier, primary transformation of ibuprofen, naproxen, diclofenac, and mecoprop by carrier-attached microorganisms was shown by the dissipation of the target compounds and the formation of five transformation products using LC-tandem MS. At the second tier, the microbial cleavage of the four organic micropollutants was confirmed with 14C-labeled micropollutants through liquid scintillation counting of the 14CO2 formed. At the third tier, microautoradiography coupled with fluorescence in situ hybridization (MAR-FISH) was used to screen carrier-attached microorganisms for uptake of the four radiolabeled micropollutants. Results from the MAR-FISH screening indicated that only a small fraction of the microbial community (≤1‰) was involved in the uptake of the radiolabeled micropollutants and that the responsible microorganisms differed between the compounds. At the fourth tier, the microbial community structure of the carrier-attached biofilms was analyzed by 16S rRNA gene amplicon sequencing. The sequencing results showed that the MAR-FISH screening targeted ∼80% of the microbial community and that several taxonomic families within the FISH-probed populations with MAR-positive signals (i.e. Firmicutes, Gammaproteobacteria, and Deltaproteobacteria) were present in both biofilms. From the broader perspective of organic micropollutant removal in biological wastewater treatment, the MAR-FISH results of this study indicate a high degree of microbial substrate specialization that could explain differences in transformation rates and patterns between micropollutants and microbial communities.

Cupriavidus pinatubonensis AEO106 deals with copper-induced oxidative stress before engaging in biodegradation of the herbicide 4-chloro-2-methylphenoxyacetic acid.

BACKGROUND: Microbial degradation of phenoxy acid (PA) herbicides in agricultural soils is important to minimize herbicide leaching to groundwater reservoirs. Degradation may, however, be hampered by exposure of the degrader bacteria to toxic metals as copper (Cu) in the soil environment. Exposure to Cu leads to accumulation of intracellular reactive oxygen species (ROS) in some bacteria, but it is not known how Cu-derived ROS and an ensuing oxidative stress affect the degradation of PA herbicides. Based on the previously proposed paradigm that bacteria deal with environmental stress before they engage in biodegradation, we studied how the degradation of the PA herbicide 2-methyl-4-chlorophenoxyacetic acid (MCPA) by the model PA degrader Cupriavidus pinatubonensis AEO106 was affected by Cu exposure. RESULTS: Exposure of C. pinatubonensis in batch culture to sublethal concentrations of Cu increased accumulation of ROS measured by the oxidant sensing probe 2,7-dichlorodihydrofluorescein diacetate and flow cytometry, and resulted in upregulation of a gene encoding a protein belong to the Ohr/OsmC protein family. The ohr/osmC gene was also highly induced by H2O2 exposure suggesting that it is involved in the oxidative stress response in C. pinatubonensis. The increased ROS accumulation and increased expression of the oxidative stress defense coincided with a delay in the catabolic performance, since both expression of the catabolic tfdA gene and MCPA mineralization were delayed compared to unexposed control cells. CONCLUSIONS: The current study suggests that Cu-induced ROS accumulation in C. pinatubonensis activates a stress response involving the product of the ohr/osmC gene. Further, the stress response is launched before induction of the catabolic tfdA gene and mineralization occurs.

Evolution of Sphingomonad Gene Clusters Related to Pesticide Catabolism Revealed by Genome Sequence and Mobilomics of Sphingobium herbicidovorans MH.

Bacterial degraders of chlorophenoxy herbicides have been isolated from various ecosystems, including pristine environments. Among these degraders, the sphingomonads constitute a prominent group that displays versatile xenobiotic-degradation capabilities. Four separate sequencing strategies were required to provide the complete sequence of the complex and plastic genome of the canonical chlorophenoxy herbicide-degrading Sphingobium herbicidovorans MH. The genome has an intricate organization of the chlorophenoxy-herbicide catabolic genes sdpA, rdpA, and cadABCD that encode the (R)- and (S)-enantiomer-specific 2,4-dichlorophenoxypropionate dioxygenases and four subunits of a Rieske non-heme iron oxygenase involved in 2-methyl-chlorophenoxyacetic acid degradation, respectively. Several major genomic rearrangements are proposed to help understand the evolution and mobility of these important genes and their genetic context. Single-strain mobilomic sequence analysis uncovered plasmids and insertion sequence-associated circular intermediates in this environmentally important bacterium and enabled the description of evolutionary models for pesticide degradation in strain MH and related organisms. The mobilome presented a complex mosaic of mobile genetic elements including four plasmids and several circular intermediate DNA molecules of insertion-sequence elements and transposons that are central to the evolution of xenobiotics degradation. Furthermore, two individual chromosomally integrated prophages were shown to excise and form free circular DNA molecules. This approach holds great potential for improving the understanding of genome plasticity, evolution, and microbial ecology.

Distribution and incorporation mode of the herbicide MCPA in soil derived organo-clay complexes.

The incorporation of xenobiotics into soil, especially via covalent bonds or sequestration has a major influence on the environmental behavior including toxicity, mobility, and bioavailability. The incorporation mode of 4-chloro-2-methylphenoxyacetic acid (MCPA) into organo-clay complexes has been investigated under a low (8.5 mg MCPA/kg soil) and high (1000 mg MCPA/kg soil) applied concentration, during an incubation period of up to 120 days. Emphasis was laid on the elucidation of distinct covalent linkages between non-extractable MCPA residues and humic sub-fractions (humic acids, fulvic acids, and humin). The cleavage of compounds by a sequential chemical degradation procedure (OH-, BBr3, RuO4, TMAH thermochemolysis) revealed for both concentration levels ester/amide bonds as the predominate incorporation modes followed by ether linkages. A possible influence of the soil microbial activity on the mode of incorporation could be observed in case of the high level samples. Structure elucidation identified MCPA as the only nonextractable substance, whereas the metabolite 4-chloro-2-methylphenol was additionally found as bioavailable and bioaccessible compound.

Conservative tracer bromide inhibits pesticide mineralisation in soil.

Bromide is a conservative tracer that is often applied with non-conservative solutes such as pesticides to estimate their retardation in the soil. It has been applied in concentrations of up to 250 g Br L-1, levels at which the growth of single-celled organisms can be inhibited. Bromide applications may therefore affect the biodegradation of non-conservative solutes in soil. The present study investigated the effect of potassium bromide (KBr) on the mineralisation of three pesticides - glyphosate, MCPA and metribuzin - in four agricultural A-horizon soils. KBr was added to soil microcosms at concentrations of 0, 0.5, 2.5 and 5 g Br- L-1 in the soil solution. The study concluded that KBr had a negative effect on pesticide mineralisation. The inhibitory effect varied depending on the KBr concentration, the type of pesticide and the type of soil. Furthermore, 16 S amplicon sequencing revealed that the KBr treatment generally reduced the abundance of bacteroidetes and proteobacteria on both an RNA and DNA level. Therefore, in order to reduce the effect of KBr on the soil bacterial community and consequently also on xenobiotic degradation, it is recommended that KBr be applied in a concentration that does not exceed 0.5 g Br- L-1 in the soil water.

Adhesion to sand and ability to mineralise low pesticide concentrations are required for efficient bioaugmentation of flow-through sand filters.

Pesticide-polluted drinking water may be remediated by inoculating waterworks sand filters with specific degrading bacteria. However, degradation efficiency is often hampered by the poor adhesion behaviour of the introduced bacteria. The phenoxy acid herbicide 4-chloro-2-methyl-phenoxy-acetic acid (MCPA) is a widespread groundwater contaminant. The aim of this study was to investigate whether specific surface characteristics of MCPA-degrading bacteria could be linked to their degrading capabilities in sand filters. Four MCPA degraders with different taxonomic affiliations and original habitats (Sphingomonas sp. PM2, Sphingomonas sp. ERG5, Burkholderia sp. TFD34, Cupriavidus sp. TFD38) were characterised with regard to their motility, cell surface hydrophobicity, biofilm formation, adhesion behaviour and ability to mineralise MCPA. Strains PM2 and ERG5 were non-motile and hydrophobic, whilst strains TFD34 and TFD38 were motile and less hydrophobic. All the strains except ERG5 showed low biofilm formation on polystyrene, although it was significantly higher on glass. PM2 was the most efficient MCPA degrader as it displayed no lag phase and reached >50 % mineralisation at all concentrations (0.0016-25 mg L-1). PM2 adhered significantly better to sand than the other strains. No link was found between motility, biofilm formation and the ability to adhere to sand. PM2 completely removed MCPA for 14 days when inoculated in sand columns with a constant inlet of 1 mg L-1 MCPA. These results demonstrate that besides the ability to degrade the contaminant, surface hydrophobicity and adherence abilities are significant parameters controlling sustained degradation in flow-through sand columns and must be considered when selecting bacteria for bioaugmentation.

Biodegradation of the Herbicide 2,4-Dichlorophenoxyacetic Acid by a New Isolated Strain of Achromobacter sp. LZ35.

In this study, a bacterial strain of Achromobacter sp. LZ35, which was capable of utilizing 2,4-dichlorophenoxyacetic acid (2,4-D) and 2-methyl-4-chlorophenoxy acetic acid (MCPA) as the sole sources of carbon and energy for growth, was isolated from the soil in a disused pesticide factory in Suzhou, China. The optimal 2,4-D degradation by strain LZ35 occurred at 30 °C and pH 8.0 when the initial 2,4-D concentration was 200 mg L-1. Strain LZ35 harbored the conserved 2,4-D/alpha-ketoglutarate dioxygenase (96%) and 2,4-dichlorophenol hydroxylase (99%), and catabolized 2,4-D via the intermediate 2,4-dichlorophenol. The inoculation of 7.8 × 106 CFU g-1 soil of strain LZ35 cells to 2,4-D-contaminated soil could efficiently remove over 75 and 90% of 100 and 50 mg L-1 2,4-D in 12 days and significantly released the phytotoxicity of maize caused by the 2,4-D residue. This is the first report of an Achromobacter sp. strain that was capable of mineralizing both 2,4-D and MCPA. This study provides us a promising candidate for its application in the bioremediation of 2,4-D- or MCPA-contaminated sites.

Degradation and enantiomeric fractionation of mecoprop in soil previously exposed to phenoxy acid herbicides - New insights for bioremediation.

Phenoxy acid-contaminated subsoils are common as a result of irregular disposal of residues and production wastes in the past. For enhancing in situ biodegradation at reducing conditions, biostimulation may be an effective option. Some phenoxy acids were marketed in racemic mixtures, and biodegradation rates may differ between enantiomers. Therefore, enantio-preferred degradation of mecoprop (MCPP) in soil was measured to get in-depth information on whether amendment with glucose (BOD equivalents as substrate for microbial growth) and nitrate (redox equivalents for oxidation) can stimulate bioremediation. The degradation processes were studied in soil sampled at different depths (3, 4.5 and 6m) at a Danish urban site with a history of phenoxy acid contamination. We observed preferential degradation of the R-enantiomer only under aerobic conditions in the soil samples from 3- and 6-m depth at environmentally relevant (nM) MCPP concentrations: enantiomer fraction (EF)<0.5. On the other hand, we observed preferential degradation of the S-enantiomer in all samples and treatments at elevated (µM) MCPP concentrations: EF>0.5. Three different microbial communities were discriminated by enantioselective degradation of MCPP: 1) aerobic microorganisms with little enantioselectivity, 2) aerobic microorganisms with R-selectivity and 3) anaerobic denitrifying organisms with S-selectivity. Glucose-amendment did not enhance MCPP degradation, while nitrate amendment enhanced the degradation of high concentrations of the herbicide.

Evidence for the importance of litter as a co-substrate for MCPA dissipation in an agricultural soil.

Environmental controls of 2-methyl-4-chlorophenoxyacetic acid (MCPA) degradation are poorly understood. We investigated whether microbial MCPA degraders are stimulated by (maize) litter and whether this process depends on concentrations of MCPA and litter. In a microcosm experiment, different amounts of litter (0, 10 and 20 g kg(-1)) were added to soils exposed to three levels of the herbicide (0, 5 and 30 mg kg(-1)). The treated soils were incubated at 20 °C for 6 weeks, and samples were taken after 1, 3 and 6 weeks of incubation. In soils with 5 mg kg(-1) MCPA, about 50 % of the MCPA was dissipated within 1 week of the incubation. Almost complete dissipation of the herbicide had occurred by the end of the incubation with no differences between the three litter amendments. At the higher concentration (30 mg kg(-1)), MCPA endured longer in the soil, with only 31 % of the initial amount being removed at the end of the experiment in the absence of litter. Litter addition greatly increased the dissipation rate with 70 and 80 % of the herbicide being dissipated in the 10 and 20 g kg(-1) litter treatments, respectively. Signs of toxic effects of MCPA on soil bacteria were observed from related phospholipid fatty acid (PLFA) analyses, while fungi showed higher tolerance to the increased MCPA levels. The abundance of bacterial tfdA genes in soil increased with the co-occurrence of litter and high MCPA concentration, indicating the importance of substrate availability in fostering MCPA-degrading bacteria and thereby improving the potential for removal of MCPA in the environment.

Impact of dry-wet and freeze-thaw events on pesticide mineralizing populations and their activity in wetland ecosystems: A microcosm study.

Riparian wetlands are proposed to mitigate diffuse pollution of surface water by pesticides in agricultural landscapes. Wetland ecosystems though are highly dynamic environments and seasonal disturbances such as freezing and drying can affect microbial population sizes in the sediment and their functionality including pesticide biodegradation, which has hardly been studied. This study examined the effect of artificially induced dry-wet or freeze-thaw events on the mineralization of the pesticides isoproturon (IPU) and 2-methoxy-4-chlorophenoxy acetic acid (MCPA) in wetland microcosms, either without or with prior enrichment of IPU/MCPA degrading populations. Without prior enrichment, mineralization of IPU and MCPA was significantly reduced after exposure to especially freeze-thaw events, as evidenced by lower mineralization rates and longer lag times compared to non-exposed microcosms. However, herbicide mineralization kinetics correlated poorly with cell numbers of herbicide mineralizers as estimated by a most probable number (MPN) approach and the number of IPU and MCPA mineralizers was unexpectedly higher in freeze-thaw and dry-wet cycle exposed setups compared to the control setups. This suggested that the observed effects of season-bound disturbances were due to other mechanisms than decay of pesticide mineralizers. In addition, in systems in which the growth of pesticide mineralizing bacteria was stimulated by amendment of IPU and MCPA, exposure to a freeze-thaw or dry-wet event only marginally affected the herbicide mineralization kinetics. Our results show that season bound environmental disturbances can affect pesticide mineralization kinetics in wetlands but that this effect can depend on the history of pesticide applications.

The potential for bioaugmentation of sand filter materials from waterworks using bacterial cultures degrading 4-chloro-2-methylphenoxyacetic acid.

BACKGROUND: The herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) is found frequently in Danish groundwater in concentrations exceeding the EU threshold limit of 0.1 µg L(-1) . Groundwater is used for drinking water, and one potential remediation strategy is bioaugmentation using inoculation of sand filters at affected waterworks with degrader bacteria. Numerous bacteria degrading phenoxyacetic acid herbicide have previously been isolated, and they may be candidates for bioaugmentation processes. Designing the optimum inoculum, however, requires knowledge of the capacity for degrading realistically low herbicide concentrations and the robustness of the bacteria when inoculated into sand filter materials. RESULTS: Testing a range of different MCPA-mineralising bacterial combinations, using a high-throughput microplate radiorespirometric mineralisation assay, highlighted three efficient cocultures for mineralising low MCPA concentrations. Cocultures demonstrating a shorter time delay before initiation of (14) C-ring-labelled MCPA mineralisation to (14) CO2 , and a more extensive mineralisation of MCPA, compared with those of single strains, were found. When inoculated into different sand filter materials, the coculture effect was diminished, but several single strains enhanced MCPA mineralisation significantly at low MCPA concentrations. CONCLUSION: This study shows that an increase in the potential for mineralisation of low herbicide concentrations in sand filter materials can be achieved by inoculating with bacterial degrader cultures. © 2014 Society of Chemical Industry.

The effects of woodchip- and straw-derived biochars on the persistence of the herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) in soils.

Sorption and degradation are the primary processes controlling the efficacy and runoff contamination risk of agrochemicals. This study assessed the influence of two biochars, made from woodchips and straw at a pyrolysis temperature of 725°C and applied to a loamy sand and a sandy soil in the concentration of 5.3 g 100 g(-1) sandy soil and 4.1 g 100 g(-1) loamy sand soil, or 53 t ha(-1) for both soil types, on degradation of the herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA). Soils were spiked with 50 mg MCPA kg(-1) soil. In the sandy soil, significantly more MCPA remained after 100 days if amended with straw-derived biochar in comparison to wood-derived biochar. Both biochars types significantly increased urease activity (p<0.05) after 37 days in the loamy sand soil, but these differences disappeared after 100 days. A root and shoot elongation test demonstrated that the soils containing straw-derived biochar and spiked with MCPA, showed the highest phytotoxicity. Both biochars were found to retard MCPA degradation in loamy sand and sandy soils. This effect could not be explained only by sorption processes due to comparatively low developed micro/mesoporous structure of both biochars shown by BET surface analysis. However, an enhanced MCPA persistence and soil toxicity in sandy soil amended with straw biochar was observed and further studies are needed to reveal the responsible mechanisms.