The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Dirk Springael - One of the best experts on this subject based on the ideXlab platform.
-
dna sip and repeated isolation corroborate variovorax as a key organism in maintaining the genetic memory for Linuron biodegradation in an agricultural soil
FEMS Microbiology Ecology, 2021Co-Authors: Harry Lerner, R. De Mot, Basak Ozturk, Anja B Dohrmann, Joice Thomas, Kathleen Marchal, Wim Dehaen, Christoph C Tebbe, Dirk SpringaelAbstract:The frequent exposure of agricultural soils to pesticides can lead to microbial adaptation, including the development of dedicated microbial populations that utilize the pesticide compound as a carbon and energy source. Soil from an agricultural field in Halen (Belgium) with a history of Linuron exposure has been studied for its Linuron-degrading bacterial populations at two time points over the past decade and Variovorax was appointed as a key Linuron degrader. Like most studies on pesticide degradation, these studies relied on isolates that were retrieved through bias-prone enrichment procedures and therefore might not represent the in situ active pesticide-degrading populations. In this study, we revisited the Halen field and applied, in addition to enrichment-based isolation, DNA stable isotope probing (DNA-SIP), to identify in situ Linuron-degrading bacteria in Linuron-exposed soil microcosms. Linuron dissipation was unambiguously linked to Variovorax and its Linuron catabolic genes and might involve the synergistic cooperation between two species. Additionally, two novel Linuron-mineralizing Variovorax isolates were obtained with high 16S rRNA gene sequence similarity to strains isolated from the same field a decade earlier. The results confirm Variovorax as a prime in situ degrader of Linuron in the studied agricultural field soil and corroborate the genus as key for maintaining the genetic memory of Linuron degradation functionality in that field.
-
dna sip and repeated isolation corroborate variovorax as a key organism in maintaining the genetic memory for Linuron biodegradation in an agricultural soil
bioRxiv, 2020Co-Authors: Harry Lerner, R. De Mot, Basak Ozturk, Anja B Dohrmann, Joice Thomas, Kathleen Marchal, Wim Dehaen, Christoph C Tebbe, Dirk SpringaelAbstract:The frequent exposure of agricultural soils to pesticides often leads to microbial adaptation, including the development of dedicated microbial populations that utilize the pesticide compound as a carbon and energy source. Soil from an agricultural field in Halen (Belgium) with a history of Linuron exposure has been studied for its Linuron-degrading bacterial populations at two time points over the past decade and Variovorax was appointed as a key Linuron degrader. Like most studies on pesticide degradation, these studies relied on isolates that were retrieved through bias-prone enrichment procedures and therefore might not represent the in situ active pesticide-degrading populations. In this study, we revisited the Halen field and applied, in addition to enrichment-based isolation, DNA stable isotope probing (DNA-SIP), to identify the in situ Linuron degrading bacteria. DNA-SIP unambiguously linked Variovorax and its Linuron catabolic genes to Linuron dissipation, likely through synergistic cooperation between two species. Additionally, two Linuron mineralizing Variovorax isolates were obtained with high 16S rRNA gene sequence similarity to strains isolated from the same field a decade earlier. The results confirm Variovorax as the in situ degrader of Linuron in the studied agricultural field and corroborate the genus as key in the maintenance of a robust genetic memory regarding Linuron degradation functionality in the examined field.
-
comparative genomics suggests mechanisms of genetic adaptation toward the catabolism of the phenylurea herbicide Linuron in variovorax
Genome Biology and Evolution, 2020Co-Authors: Basak Ozturk, Johannes Werner, Jan P Meierkolthoff, Boyke Bunk, Cathrin Sproer, Dirk SpringaelAbstract:Biodegradation of the phenylurea herbicide Linuron appears a specialization within a specific clade of the Variovorax genus. The Linuron catabolic ability is likely acquired by horizontal gene transfer but the mechanisms involved are not known. The full-genome sequences of six Linuron-degrading Variovorax strains isolated from geographically distant locations were analyzed to acquire insight into the mechanisms of genetic adaptation toward Linuron metabolism. Whole-genome sequence analysis confirmed the phylogenetic position of the Linuron degraders in a separate clade within Variovorax and indicated that they unlikely originate from a common ancestral Linuron degrader. The Linuron degraders differentiated from Variovorax strains that do not degrade Linuron by the presence of multiple plasmids of 20-839 kb, including plasmids of unknown plasmid groups. The Linuron catabolic gene clusters showed 1) high conservation and synteny and 2) strain-dependent distribution among the different plasmids. Most of them were bordered by IS1071 elements forming composite transposon structures, often in a multimeric array configuration, appointing IS1071 as a key element in the recruitment of Linuron catabolic genes in Variovorax. Most of the strains carried at least one (catabolic) broad host range plasmid that might have been a second instrument for catabolic gene acquisition. We conclude that clade 1 Variovorax strains, despite their different geographical origin, made use of a limited genetic repertoire regarding both catabolic functions and vehicles to acquire Linuron biodegradation.
-
proma plasmids are instrumental in the dissemination of Linuron catabolic genes between different genera
bioRxiv, 2019Co-Authors: Johannes Werner, Dirk Springael, Boyke Bunk, Cathrin Sproer, Eman Nour, Kornelia Smalla, Basak OzturkAbstract:PromA plasmids are broad host range plasmids, which are often cryptic and hence have an uncertain ecological role. We present three novel PromA {gamma} plasmids which carry genes associated with degradation of the phenylurea herbicide Linuron, two (pPBL-H3-2 and pBPS33-2) of which originate from unrelated Hydrogenophaga hosts isolated from different environments, and one (pEN1) which was exogenously captured from an on-farm biopurification system. Both Hydrogenophaga plasmids carry all three necessary gene clusters determining the three main steps for conversion of Linuron to Krebs cycle intermediates, while pEN1 only determines the initial Linuron hydrolysis step. Linuron catabolic gene clusters that determine the same step were identical on all plasmids, encompassed in differently arranged constellations and characterized by the presence of multiple IS1071 elements. In all plasmids except pEN1, the insertion spot of the catabolic genes in the PromA {gamma} plasmids was the same. Highly similar PromA plasmids carrying the Linuron degrading gene cargo at the same insertion spot were were previously identified in Linuron degrading Variovorax sp. Interestingly, in both Hydrogenophaga populations not every PromA plasmid copy carries catabolic genes. The results indicate that PromA plasmids are important vehicles of Linuron catabolic gene dissemination, rather than being cryptic and only important for the mobilization of other plasmids.
-
comparative genomics unravels mechanisms of genetic adaptation for the catabolism of the phenylurea herbicide Linuron in variovorax
bioRxiv, 2019Co-Authors: Basak Ozturk, Johannes Werner, Jan P Meierkolthoff, Boyke Bunk, Cathrin Sproer, Dirk SpringaelAbstract:Abstract Biodegradation of the phenylurea herbicide Linuron appears a specialization within a specific clade of the Variovorax genus. The Linuron catabolic ability is likely acquired by horizontal gene transfer but the mechanisms involved are not known. The full genome sequences of six Linuron degrading Variovorax strains isolated from geographically distant locations were analyzed to acquire insight in the mechanisms of genetic adaptation towards Linuron metabolism in Variovorax. Whole genome sequence analysis confirmed the phylogenetic position of the Linuron degraders in a separate clade within Variovorax and indicated their unlikely origin from a common ancestral Linuron degrader. The Linuron degraders differentiated from non-degraders by the presence of multiple plasmids of 20 to 839 kb, including plasmids of unknown plasmid groups. The Linuron catabolic gene clusters showed (i) high conservation and synteny and (ii) strain-dependent distribution among the different plasmids. All were bordered by IS1071 elements forming composite transposon structures appointing IS1071 as key for catabolic gene recruitment. Most of the strain carried at least one broad host range plasmid that might have been a second instrument for catabolic gene acquisition. We conclude that clade 1Variovorax strains, despite their different geographical origin, made use of a limited genetic repertoire to acquire Linuron biodegradation. Importance The genus Variovorax and especially a clade of strains that phylogenetically separates from the majority of Variovorax species, appears to be a specialist in the biodegradation of the phenyl urea herbicide Linuron. Horizontal gene transfer (HGT) likely played an essential role in the genetic adaptation of those strain to acquire the Linuron catabolic genotype. However, we do not know the genetic repertoire involved in this adaptation both regarding catabolic gene functions as well as gene functions that promote HGT neither do we know how this varies between the different strains. These questions are addressed in this paper by analyzing the full genome sequences of six Linuron degrading Variovorax strains. This knowledge is important for understanding the mechanisms that steer world-wide genetic adaptation in a particular species and this for a particular phenotypic trait as Linuron biodegradation.
Karolien Bers - One of the best experts on this subject based on the ideXlab platform.
-
functional redundancy of Linuron degradation in microbial communities in agricultural soil and biopurification systems
Applied and Environmental Microbiology, 2016Co-Authors: Benjamin Horemans, Karolien Bers, R. De Mot, Erick Ruiz Romero, Eva Pose Juan, Vincent Dunon, Dirk SpringaelAbstract:The abundance of libA , encoding a hydrolase that initiates Linuron degradation in the Linuron metabolizing Variovorax sp. SRS16, was previously found to correlate well with Linuron mineralization in various environments, but not in all. Recently, an alternative Linuron hydrolase HylA was identified in Variovorax sp. WDL1, a strain that initiates Linuron degradation in a Linuron mineralizing commensal bacterial consortium. The discovery of alternative Linuron hydrolases poses questions about the respective contribution and competitive character of hylA and libA carrying bacteria as well as the role of Linuron mineralizing consortia versus single strains in Linuron exposed settings. Therefore, dynamics of hylA as well as dcaQ as a marker for downstream catabolic functions involved in Linuron mineralization, in response to Linuron treatment in agricultural soil and on-farm biopurification systems (BPS), were compared with previously reported libA dynamics. The results suggest that (i) organisms containing libA and hylA both contribute simultaneously to Linuron biodegradation in the same environment albeit to a varying extent, (ii) that environmental Linuron mineralization depends on multispecies bacterial food webs and (iii) initiation of Linuron mineralization can be governed by currently unidentified enzymes. Importance Several years ago, Linuron degrading bacteria and a cooperative consortium were enriched and isolated from environmental samples that were exposed to Linuron for a period of time. However, it9s an unwarranted assumption that these bacterial isolates would be key protagonists in the in situ removal of the pesticide in the samples where they were isolated from. In this study, the complementary involvement of two initiators of Linuron degradation executing the first step in the degradation of Linuron and essential for its final mineralization, as well as the involvement of metabolite mineralizing bacterial populations, was illustrated. This study is a first in illustrating the importance of the synergistic metabolism of Linuron in agricultural soils and biopurification systems.
-
Dynamics of the Linuron Hydrolase libA Gene Pool Size in Response to Linuron Application and Environmental Perturbations in Agricultural Soil and On-Farm Biopurification Systems
2015Co-Authors: Karolien Bers, Kristel Sniegowski, René A De Mot, Dirk SpringaelaAbstract:libA, a gene encoding a novel type of Linuron hydrolase, was recently identified in the Linuron-mineralizing Variovorax sp. strain SRS16. In order to assess the contribution of libA to Linuron degradation in environmental settings, libA abundance was moni-tored in response to the application of Linuron and to environmental perturbations in agricultural soil microcosms andmicro-cosms simulating the matrix of on-farm biopurification systems. libA numbers were measured by real-time PCR and linked to reported data of Variovorax community composition and Linuronmineralization capacity. In the soil microcosms and one bio-purification system setup, libA numbers responded to the application of Linuron and environmental changes in congruency with the modulation of Linuronmineralization capacity and the occurrence of a particular Variovorax phylotype (phylotype A). How-ever, in another biopurification system setup, no such correlations were found. Our data suggest that in the simulated environ-mental settings, the occurrence of libA can be linked to the Linuronmineralization capacity and that libA is primarily hosted by Variovorax phylotype A strains. However, the results also suggest that, apart from libA, other, as-yet-unknown isofunctional genes play an important role in Linuronmineralization in the environment. Linuron is a phenylurea herbicide that is widely used to controla diversity of annual and perennial broadleaf and grassy weeds in various crops and in orchards (7). As a consequence, Linuron is detected in soil, groundwater, and surface water (17). The long history of Linuron exposure of agricultural soils apparently re
-
in situ response of the Linuron degradation potential to Linuron application in an agricultural field
FEMS Microbiology Ecology, 2013Co-Authors: Karolien Bers, R. De Mot, Dirk SpringaelAbstract:To assess the involvement of the genus Variovorax and the Linuron hydrolase gene libA in in situ Linuron degradation in agricultural fields, changes in Variovorax community size and composition, in libA abundance and in Linuron mineralization capacity were monitored in field soil plots either treated or not with a Linuron-containing herbicide mixture. Changes in Variovorax community composition, due to the proliferation of a hereto unknown Variovorax phylotype D, and increases in libA numbers occurred concomitant to increases in Linuron mineralization capacity in the plot treated with the herbicide mixture. The observations suggest that Variovorax and libA proliferated as a response to Linuron and hence their contribution to in situ Linuron degradation. The involvement of Variovorax phylotype D and libA in Linuron degradation in the examined soil was supported by laboratory soil microcosm experiments. Attempts to enrich in suspended cultures and isolate the organism corresponding to phylotype D from the soil were unsuccessful as the enrichment resulted in replacement of Variovorax phylotype D by other Variovorax phylotypes. This illustrates that Linuron-degrading strains isolated by liquid enrichment cultures are not always representatives of those responsive to Linuron in the field, although the genus specificity of Linuron degradation was retained.
-
HylA, an alternative hydrolase for initiation of catabolism of the phenylurea herbicide Linuron in Variovorax sp. strains.
Applied and environmental microbiology, 2013Co-Authors: Karolien Bers, Isabelle Batisson, Paul Proost, Ruddy Wattiez, R. De MotAbstract:Variovorax sp. strain WDL1, which mineralizes the phenylurea herbicide Linuron, expresses a novel Linuron-hydrolyzing enzyme, HylA, that converts Linuron to 3,4-dichloroaniline (DCA). The enzyme is distinct from the Linuron hydrolase LibA enzyme recently identified in other Linuron-mineralizing Variovorax strains and from phenylurea-hydrolyzing enzymes (PuhA, PuhB) found in Gram-positive bacteria. The dimeric enzyme belongs to a separate family of hydrolases and differs in Km, temperature optimum, and phenylurea herbicide substrate range. Within the metal-dependent amidohydrolase superfamily, HylA and PuhA/PuhB belong to two distinct protein families, while LibA is a member of the unrelated amidase signature family. The hylA gene was identified in a draft genome sequence of strain WDL1. The involvement of hylA in Linuron degradation by strain WDL1 is inferred from its absence in spontaneous WDL1 mutants defective in Linuron hydrolysis and its presence in Linuron-degrading Variovorax strains that lack libA. In strain WDL1, the hylA gene is combined with catabolic gene modules encoding the downstream pathways for DCA degradation, which are very similar to those present in Variovorax sp. SRS16, which contains libA. Our results show that the expansion of a DCA catabolic pathway toward Linuron degradation in Variovorax can involve different but isofunctional Linuron hydrolysis genes encoding proteins that belong to evolutionary unrelated hydrolase families. This may be explained by divergent evolution and the independent acquisition of the corresponding genetic modules.
-
minimal pesticide primed soil inoculum density to secure maximum pesticide degradation efficiency in on farm biopurification systems
Chemosphere, 2012Co-Authors: Kristel Sniegowski, Karolien Bers, Peter Jaeken, Jaak Ryckeboer, Pieter Spanoghe, Dirk SpringaelAbstract:Abstract Addition of pesticide-primed soil containing adapted pesticide degrading bacteria to the biofilter matrix of on farm biopurification systems (BPS) which treat pesticide contaminated wastewater, has been recommended, in order to ensure rapid establishment of a pesticide degrading microbial community in BPS. However, uncertainties exist about the minimal soil inoculum density needed for successful bioaugmentation of BPS. Therefore, in this study, BPS microcosm experiments were initiated with different Linuron primed soil inoculum densities ranging from 0.5 to 50 vol.% and the evolution of the Linuron mineralization capacity in the microcosms was monitored during feeding with Linuron. Successful establishment of a Linuron mineralization community in the BPS microcosms was achieved with all inoculum densities including the 0.5 vol.% density with only minor differences in the time needed to acquire maximum degradation capacity. Moreover, once established, the robustness of the Linuron degrading microbial community towards expected stress situations proved to be independent of the initial inoculum density. This study shows that pesticide-primed soil inoculum densities as low as 0.5 vol.% can be used for bioaugmentation of a BPS matrix and further supports the use of BPS for treatment of pesticide-contaminated wastewater at farmyards.
Jens Aamand - One of the best experts on this subject based on the ideXlab platform.
-
constitutive mineralization of low concentrations of the herbicide Linuron by a variovorax sp strain
Fems Microbiology Letters, 2009Co-Authors: Sebastian R Sorensen, Allan Simonsen, Jens AamandAbstract:The mineralization of the herbicide Linuron at concentrations of μg and mg L−1 was studied in liquid batch experiments with Variovorax sp. strain SRS16. The strain was highly efficient at mineralizing a range of Linuron concentrations (0.002–10 mg L−1) with 20–60% of the added 14C-ring-labeled Linuron metabolized to 14CO2 within hours to days depending on the initial Linuron concentration and incubation period. At mg L−1 Linuron concentrations the mineralization activity by SRS16 was inducible and a shift to constitutive mineralization activity was apparent with a reduction in the Linuron concentration to μg L−1 levels. This study revealed that strain SRS16 is a promising candidate for bioaugmentation of water or soil resources contaminated with low Linuron concentrations.
-
spatial variability in the mineralisation of the phenylurea herbicide Linuron within a danish agricultural field multivariate correlation to simple soil parameters
Pest Management Science, 2005Co-Authors: Jim Rasmussen, Jens Aamand, Per Rosenberg, Ole Stig Jacobsen, Sebastian R SorensenAbstract:The spatial variability in the mineralisation rate of Linuron [N-(3,4-dichlorophenyl)-N'-methoxy-N'-methylurea] was studied within a previously treated Danish agricultural field by sampling soils from eleven different plots randomly distributed across an area of 20 x 20m. The soils were characterised with respect to different abiotic and biotic properties including moisture content, organic matter content, pH, nutrient content, bacterial biomass, potential for mineralisation ofMCPA [(4-chloro-2-methylphenoxy)acetic acid] and Linuron. Five soils had a potential for mineralisation of Linuron, with 5-15% of the added [ring-U- 14 C]Linuron metabolised to 14 CO 2 within 60 days at 10°C, while no extensive mineralisation of Linuron was observed in the six remaining soils within this period. A TLC analysis of the methanol-extractable residues showed no development of 14 C-labelled metabolites from Linuron in any of the samples. Multivariate analysis was conducted to elucidate relationships between the intrinsic properties of single soil samples and initial rate of Linuron mineralisation. The analysis indicated that important soil parameters in determining the spatial heterogeneity included the C total /N total ratio, pH and the water-extractable potassium contents, with the first of these highly negatively correlated and the last two highly positively correlated to the initial Linuron mineralisation rate. This study shows that enhanced biodegradation of Linuron may develop with successive field treatments, but that considerable in-field spatial heterogeneity in the degradation rate still exists. Combined with a parallel enrichment study focused on the underlying microbial processes, the present results suggest that intrinsic soil properties affect the Linuron-metabolising bacterial population and thereby determine the spatial variability in the Linuron mineralisation activity.
-
elucidating the key member of a Linuron mineralizing bacterial community by pcr and reverse transcription pcr denaturing gradient gel electrophoresis 16s rrna gene fingerprinting and cultivation
Applied and Environmental Microbiology, 2005Co-Authors: Sebastian R Sorensen, Jim Rasmussen, Ole Stig Jacobsen, Carsten S Jacobsen, Rene K Juhler, Jens AamandAbstract:The phenylurea herbicide Linuron [N-(3,4-dichlorophenyl)-N′-methoxy-N′-methylurea] is used worldwide in the conventional production of corn, cereals, vegetables, and fruit. The rates of dissipation in agricultural soils determined by laboratory and field experiments are highly variable, with values ranging from days to several years (2, 8, 9, 11, 16, 23). Linuron is frequently detected in surface and ground waters near or below areas with intensive use, and in one extreme case, Linuron was detected in a drinking-water well in concentrations up to 2,800 μg liter−1 (2). Unfortunately, Linuron and some of its metabolites are suspected of being endocrine disruptors (12) and of having toxic effects on various aquatic and soil organisms (2, 22), which has stimulated research aimed at studying Linuron-mineralizing microorganisms from agricultural soils. Mixed bacterial cultures able to mineralize Linuron have been derived from extensively treated British and Belgian agricultural soils (4, 7, 15). Similar enrichments based on related phenylurea herbicides (1, 3, 6, 19) suggest that this group can serve as carbon and nitrogen sources for bacterial metabolism in agricultural soils. Several attempts to cultivate phenylurea-metabolizing soil bacteria from degradative enrichment cultures, however, were unsuccessful (e.g., studies described in references 7, 15, and 18), and the active bacteria seem reluctant to grow on agar media. Recently, however, the first Linuron-mineralizing bacterium, Variovorax sp. strain WDL1 (4), was isolated from previously treated Belgian agricultural soil. Strain WDL1 appeared to be an ineffective Linuron degrader in pure culture and dependent on four other consortium members (4). The phenomenon of synergistic bacterial interactions has also been described for the herbicide isoproturon (19, 20). An extensively Linuron-treated Danish agricultural field harboring a potential for rapid Linuron mineralization was located among three investigated fields. The objective of this study was to obtain Linuron-mineralizing enrichment cultures and pinpoint degradative microorganisms by molecular and cultivation-based techniques.
-
elucidating the key member of a Linuron mineralizing bacterial community by pcr and reverse transcription pcr denaturing gradient gel electrophoresis 16s rrna gene fingerprinting and cultivation
Applied and Environmental Microbiology, 2005Co-Authors: Sebastian R Sorensen, Jim Rasmussen, Ole Stig Jacobsen, Carsten S Jacobsen, Rene K Juhler, Jens AamandAbstract:A bacterial community from Danish agricultural soil was enriched with Linuron [N-(3,4-dichlorophenyl)-N′-methoxy-N′-methylurea] as the sole carbon and nitrogen source. The community mineralized [ring-U-14C]Linuron completely to 14CO2 and 14C-biomass. Denaturing gradient gel electrophoresis analysis and cultivation revealed that a Variovorax sp. was responsible for the mineralization activity.
Philip Breugelmans - One of the best experts on this subject based on the ideXlab platform.
-
environmental dissolved organic matter governs biofilm formation and subsequent Linuron degradation activity of a Linuron degrading bacterial consortium
Applied and Environmental Microbiology, 2013Co-Authors: Benjamin Horemans, Erik Smolders, Philip Breugelmans, Johan Hofkens, Dirk SpringaelAbstract:ABSTRACT It was examined whether biofilm growth on dissolved organic matter (DOM) of a three-species consortium whose members synergistically degrade the phenylurea herbicide Linuron affected the consortium9s integrity and subsequent Linuron-degrading functionality. Citrate as a model DOM and three environmental DOM (eDOM) formulations of different quality were used. Biofilms developed with all DOM formulations, and the three species were retained in the biofilm. However, biofilm biomass, species composition, architecture, and colocalization of member strains depended on DOM and its biodegradability. To assess the Linuron-degrading functionality, biofilms were subsequently irrigated with Linuron at 10 mg liter −1 or 100 μg liter −1 . Instant Linuron degradation, the time needed to attain maximal Linuron degradation, and hence the total amount of Linuron removed depended on both the DOM used for growth and the Linuron concentration. At 10 mg liter −1 , the final Linuron degradation efficiency was as high as previously observed without DOM except for biofilms fed with humic acids which did not degrade Linuron. At 100 μg liter −1 Linuron, DOM-grown biofilms degraded Linuron less efficiently than biofilms receiving 10 mg liter −1 Linuron. The amount of Linuron removed was more correlated with biofilm species composition than with biomass or structure. Based on visual observations, colocalization of consortium members in biofilms after the DOM feed appears essential for instant Linuron-degrading activity and might explain the differences in overall Linuron degradation. The data show that DOM quality determines biofilm structure and composition of the pesticide-degrading consortium in periods with DOM as the main carbon source and can affect subsequent pesticide-degrading activity, especially at micropollutant concentrations.
-
a molecular toolbox to estimate the number and diversity of variovorax in the environment application in soils treated with the phenylurea herbicide Linuron
FEMS Microbiology Ecology, 2011Co-Authors: Karolien Bers, Kristel Sniegowski, Philip Breugelmans, Pieter Albers, Larissa Hendrickx, R. De MotAbstract:Real-time PCR and PCR-denaturing gradient gel electrophoresis (DGGE) approaches that specifically target the Variovorax 16S rRNA gene were developed to estimate the number and diversity of Variovorax in environmental ecosystems. PCR primers suitable for both methods were selected as such that the enclosed sequence showed maximum polymorphism. PCR specificity was maximized by combining PCR with a targeted endonuclease treatment of template DNA to eliminate 16S rRNA genes of the closely related Acidovorax. DGGE allowed the grouping of PCR amplicons according to the phylogenetic grouping within the genus Variovorax. The toolbox was used to assess the Variovorax community dynamics in agricultural soil microcosms (SMs) exposed to the phenylurea herbicide Linuron. Exposure to Linuron resulted in an increased abundance within the Variovorax community of a subgroup previously linked to Linuron degradation through cultivation-dependent isolation. SMs that were treated only once with Linuron reverted to the initial community composition 70 days after Linuron exposure. In contrast, SMs irrigated with Linuron on a long-term base showed a significant increase in Variovorax number after 70 days. Our data support the hypothesis that the genus Variovorax is involved in Linuron degradation in Linuron-treated agricultural soils.
-
response to mixed substrate feeds of the structure and activity of a Linuron degrading triple species biofilm
Research in Microbiology, 2010Co-Authors: Philip Breugelmans, Benjamin Horemans, Johan Hofkens, Dirk SpringaelAbstract:We sought to determine whether the pesticide-degrading performance of a multi-species bacterial biofilm is affected by co-occurrence of multiple nutrient sources. Thus, the 3-(3,4-dichlorophenyl)-1-methoxy-1-methyl urea (Linuron)-degrading activity of a triple-species Linuron-degrading consortium, cultivated in continuous flow biofilm systems, was monitored when exposed to mixed substrate feeds which contained, in addition to Linuron, readily assimilated carbon (i.e. citrate and trypticase soy broth) and/or nitrogen (i.e. ammonium) sources. The addition of alternative carbon sources at different concentrations resulted in diminished Linuron degradation efficiency. In addition, the efficiency of removal of the Linuron metabolite 3,4-dichloroaniline was affected. These effects might be attributed to catabolic repression of the Linuron metabolic pathway in the presence of alternative carbon substrates. Moreover, each nutrient condition resulted in a particular biofilm composition and a particular spatial and structural organization, which might also be related to the performance of the biofilm community. Results show that the activity of pesticide-degrading biofilms strongly depends on prevailing nutrient conditions and that the ideal biofilm configuration and activity, as observed under selective conditions, does not exist in real-life environmental conditions where mixtures of substrates are often present.
-
proteomic study of Linuron and 3 4 dichloroaniline degradation by variovorax sp wdl1 evidence for the involvement of an aniline dioxygenase related multicomponent protein
Research in Microbiology, 2010Co-Authors: Philip Breugelmans, Karolien Bers, Ruddy Wattiez, R. De Mot, Winnie Dejonghe, Baptiste Leroy, Dirk SpringaelAbstract:A proteomic approach was used to explore the metabolism of the phenylurea herbicide Linuron and 3,4-dichloroaniline (3,4-DCA) in Variovorax sp. WDL1. This bacterium grows on Linuron as sole source of carbon, nitrogen and energy, while it transiently accumulates 3,4-DCA as a metabolite. Differential protein expression analysis of Variovorax sp. WDL1 grown in a heterotrophic medium in the presence and absence of Linuron or 3,4-DCA was conducted using 2-D PAGE. Selected up- and downregulated proteins were identified with nanoLC-ESI-MS/MS. In the 3,4-DCA-supplemented culture, upregulation of several proteins showing high amino acid sequence similarity to different components of the multicomponent aniline dioxygenase in aniline-degrading Proteobacteria was observed. For one of the components, multiple variant proteins were detected, suggesting that strain WDL1 harbors several copies of the aniline dioxygenase (AD) gene cluster which are simultaneously expressed in the presence of 3,4-DCA. A number of unidentifiable proteins, which were upregulated in the Linuron- and/or 3,4-DCA-supplemented cultures, might represent up to now uncharacterized proteins with a role in Linuron and/or 3,4-DCA degradation in strain WDL1. In addition, several stress-related proteins were differentially expressed.
-
architecture and spatial organization in a triple species bacterial biofilm synergistically degrading the phenylurea herbicide Linuron
FEMS Microbiology Ecology, 2008Co-Authors: Philip Breugelmans, Kim Bundvig Barken, Tim Tolkernielsen, Johan Hofkens, Winnie DejongheAbstract:Members of a triple-species 3-(3,4-dichlorophenyl)-1-methoxy-1-methyl urea (Linuron)-mineralizing consortium, i.e. the Linuron- and 3,4-dichloroaniline-degrading Variovorax sp. WDL1, the 3,4-dichloroaniline-degrading Comamonas testosteroni WDL7 and the N,O-dimethylhydroxylamine-degrading Hyphomicrobium sulfonivorans WDL6, were cultivated as mono- or multi-species biofilms in flow cells irrigated with selective or nonselective media, and examined with confocal laser scanning microscopy. In contrast to mono-species biofilms of Variovorax sp. WDL1, the triple-species consortium biofilm degraded Linuron completely through apparent synergistic interactions. The triple-species Linuron-fed consortium biofilm displayed a heterogeneous structure with an irregular surface topography that most resembled the topography of Linuron-fed mono-species WDL1 biofilms, indicating that WDL1 had a dominating influence on the triple-species biofilm architecture. This architecture was dependent on the carbon source supplied, as the biofilm architecture of WDL1 growing on alternative carbon sources was different from that observed under Linuron-fed conditions. Linuron-fed triple-species consortium biofilms consisted of mounds composed of closely associated WDL1, WDL7 and WDL6 cells, while this association was lost when the consortium was grown on a nonselective carbon source. In addition, under Linuron-fed conditions, microcolonies displaying associated growth developed rapidly after inoculation. These observations indicate that the spatial organization in the Linuron-fed consortium biofilm reflected the metabolic interactions within the consortium.
R. De Mot - One of the best experts on this subject based on the ideXlab platform.
-
dna sip and repeated isolation corroborate variovorax as a key organism in maintaining the genetic memory for Linuron biodegradation in an agricultural soil
FEMS Microbiology Ecology, 2021Co-Authors: Harry Lerner, R. De Mot, Basak Ozturk, Anja B Dohrmann, Joice Thomas, Kathleen Marchal, Wim Dehaen, Christoph C Tebbe, Dirk SpringaelAbstract:The frequent exposure of agricultural soils to pesticides can lead to microbial adaptation, including the development of dedicated microbial populations that utilize the pesticide compound as a carbon and energy source. Soil from an agricultural field in Halen (Belgium) with a history of Linuron exposure has been studied for its Linuron-degrading bacterial populations at two time points over the past decade and Variovorax was appointed as a key Linuron degrader. Like most studies on pesticide degradation, these studies relied on isolates that were retrieved through bias-prone enrichment procedures and therefore might not represent the in situ active pesticide-degrading populations. In this study, we revisited the Halen field and applied, in addition to enrichment-based isolation, DNA stable isotope probing (DNA-SIP), to identify in situ Linuron-degrading bacteria in Linuron-exposed soil microcosms. Linuron dissipation was unambiguously linked to Variovorax and its Linuron catabolic genes and might involve the synergistic cooperation between two species. Additionally, two novel Linuron-mineralizing Variovorax isolates were obtained with high 16S rRNA gene sequence similarity to strains isolated from the same field a decade earlier. The results confirm Variovorax as a prime in situ degrader of Linuron in the studied agricultural field soil and corroborate the genus as key for maintaining the genetic memory of Linuron degradation functionality in that field.
-
dna sip and repeated isolation corroborate variovorax as a key organism in maintaining the genetic memory for Linuron biodegradation in an agricultural soil
bioRxiv, 2020Co-Authors: Harry Lerner, R. De Mot, Basak Ozturk, Anja B Dohrmann, Joice Thomas, Kathleen Marchal, Wim Dehaen, Christoph C Tebbe, Dirk SpringaelAbstract:The frequent exposure of agricultural soils to pesticides often leads to microbial adaptation, including the development of dedicated microbial populations that utilize the pesticide compound as a carbon and energy source. Soil from an agricultural field in Halen (Belgium) with a history of Linuron exposure has been studied for its Linuron-degrading bacterial populations at two time points over the past decade and Variovorax was appointed as a key Linuron degrader. Like most studies on pesticide degradation, these studies relied on isolates that were retrieved through bias-prone enrichment procedures and therefore might not represent the in situ active pesticide-degrading populations. In this study, we revisited the Halen field and applied, in addition to enrichment-based isolation, DNA stable isotope probing (DNA-SIP), to identify the in situ Linuron degrading bacteria. DNA-SIP unambiguously linked Variovorax and its Linuron catabolic genes to Linuron dissipation, likely through synergistic cooperation between two species. Additionally, two Linuron mineralizing Variovorax isolates were obtained with high 16S rRNA gene sequence similarity to strains isolated from the same field a decade earlier. The results confirm Variovorax as the in situ degrader of Linuron in the studied agricultural field and corroborate the genus as key in the maintenance of a robust genetic memory regarding Linuron degradation functionality in the examined field.
-
functional redundancy of Linuron degradation in microbial communities in agricultural soil and biopurification systems
Applied and Environmental Microbiology, 2016Co-Authors: Benjamin Horemans, Karolien Bers, R. De Mot, Erick Ruiz Romero, Eva Pose Juan, Vincent Dunon, Dirk SpringaelAbstract:The abundance of libA , encoding a hydrolase that initiates Linuron degradation in the Linuron metabolizing Variovorax sp. SRS16, was previously found to correlate well with Linuron mineralization in various environments, but not in all. Recently, an alternative Linuron hydrolase HylA was identified in Variovorax sp. WDL1, a strain that initiates Linuron degradation in a Linuron mineralizing commensal bacterial consortium. The discovery of alternative Linuron hydrolases poses questions about the respective contribution and competitive character of hylA and libA carrying bacteria as well as the role of Linuron mineralizing consortia versus single strains in Linuron exposed settings. Therefore, dynamics of hylA as well as dcaQ as a marker for downstream catabolic functions involved in Linuron mineralization, in response to Linuron treatment in agricultural soil and on-farm biopurification systems (BPS), were compared with previously reported libA dynamics. The results suggest that (i) organisms containing libA and hylA both contribute simultaneously to Linuron biodegradation in the same environment albeit to a varying extent, (ii) that environmental Linuron mineralization depends on multispecies bacterial food webs and (iii) initiation of Linuron mineralization can be governed by currently unidentified enzymes. Importance Several years ago, Linuron degrading bacteria and a cooperative consortium were enriched and isolated from environmental samples that were exposed to Linuron for a period of time. However, it9s an unwarranted assumption that these bacterial isolates would be key protagonists in the in situ removal of the pesticide in the samples where they were isolated from. In this study, the complementary involvement of two initiators of Linuron degradation executing the first step in the degradation of Linuron and essential for its final mineralization, as well as the involvement of metabolite mineralizing bacterial populations, was illustrated. This study is a first in illustrating the importance of the synergistic metabolism of Linuron in agricultural soils and biopurification systems.
-
in situ response of the Linuron degradation potential to Linuron application in an agricultural field
FEMS Microbiology Ecology, 2013Co-Authors: Karolien Bers, R. De Mot, Dirk SpringaelAbstract:To assess the involvement of the genus Variovorax and the Linuron hydrolase gene libA in in situ Linuron degradation in agricultural fields, changes in Variovorax community size and composition, in libA abundance and in Linuron mineralization capacity were monitored in field soil plots either treated or not with a Linuron-containing herbicide mixture. Changes in Variovorax community composition, due to the proliferation of a hereto unknown Variovorax phylotype D, and increases in libA numbers occurred concomitant to increases in Linuron mineralization capacity in the plot treated with the herbicide mixture. The observations suggest that Variovorax and libA proliferated as a response to Linuron and hence their contribution to in situ Linuron degradation. The involvement of Variovorax phylotype D and libA in Linuron degradation in the examined soil was supported by laboratory soil microcosm experiments. Attempts to enrich in suspended cultures and isolate the organism corresponding to phylotype D from the soil were unsuccessful as the enrichment resulted in replacement of Variovorax phylotype D by other Variovorax phylotypes. This illustrates that Linuron-degrading strains isolated by liquid enrichment cultures are not always representatives of those responsive to Linuron in the field, although the genus specificity of Linuron degradation was retained.
-
HylA, an alternative hydrolase for initiation of catabolism of the phenylurea herbicide Linuron in Variovorax sp. strains.
Applied and environmental microbiology, 2013Co-Authors: Karolien Bers, Isabelle Batisson, Paul Proost, Ruddy Wattiez, R. De MotAbstract:Variovorax sp. strain WDL1, which mineralizes the phenylurea herbicide Linuron, expresses a novel Linuron-hydrolyzing enzyme, HylA, that converts Linuron to 3,4-dichloroaniline (DCA). The enzyme is distinct from the Linuron hydrolase LibA enzyme recently identified in other Linuron-mineralizing Variovorax strains and from phenylurea-hydrolyzing enzymes (PuhA, PuhB) found in Gram-positive bacteria. The dimeric enzyme belongs to a separate family of hydrolases and differs in Km, temperature optimum, and phenylurea herbicide substrate range. Within the metal-dependent amidohydrolase superfamily, HylA and PuhA/PuhB belong to two distinct protein families, while LibA is a member of the unrelated amidase signature family. The hylA gene was identified in a draft genome sequence of strain WDL1. The involvement of hylA in Linuron degradation by strain WDL1 is inferred from its absence in spontaneous WDL1 mutants defective in Linuron hydrolysis and its presence in Linuron-degrading Variovorax strains that lack libA. In strain WDL1, the hylA gene is combined with catabolic gene modules encoding the downstream pathways for DCA degradation, which are very similar to those present in Variovorax sp. SRS16, which contains libA. Our results show that the expansion of a DCA catabolic pathway toward Linuron degradation in Variovorax can involve different but isofunctional Linuron hydrolysis genes encoding proteins that belong to evolutionary unrelated hydrolase families. This may be explained by divergent evolution and the independent acquisition of the corresponding genetic modules.