Molecular processes underlying synergistic linuron mineralization in a triple-species bacterial consortium biofilm revealed by differential transcriptomics.

The proteobacteria Variovorax sp. WDL1, Comamonas testosteroni WDL7, and Hyphomicrobium sulfonivorans WDL6 compose a triple-species consortium that synergistically degrades and grows on the phenylurea herbicide linuron. To acquire a better insight into the interactions between the consortium members and the underlying molecular mechanisms, we compared the transcriptomes of the key biodegrading strains WDL7 and WDL1 grown as biofilms in either isolation or consortium conditions by differential RNAseq analysis. Differentially expressed pathways and cellular systems were inferred using the network-based algorithm PheNetic. Coculturing affected mainly metabolism in WDL1. Significantly enhanced expression of hylA encoding linuron hydrolase was observed. Moreover, differential expression of several pathways involved in carbohydrate, amino acid, nitrogen, and sulfur metabolism was observed indicating that WDL1 gains carbon and energy from linuron indirectly by consuming excretion products from WDL7 and/or WDL6. Moreover, in consortium conditions, WDL1 showed a pronounced stress response and overexpression of cell to cell interaction systems such as quorum sensing, contact-dependent inhibition, and Type VI secretion. Since the latter two systems can mediate interference competition, it prompts the question if synergistic linuron degradation is the result of true adaptive cooperation or rather a facultative interaction between bacteria that coincidentally occupy complementary metabolic niches.

Environmental dissolved organic matter governs biofilm formation and subsequent linuron degradation activity of a linuron-degrading bacterial consortium.

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 consortium's 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.

Improvement of pesticide mineralization in on-farm biopurification systems by bioaugmentation with pesticide-primed soil.

Microcosms were used to examine whether pesticide-primed soils could be preferentially used over nonprimed soils for bioaugmentation of on-farm biopurification systems (BPS) to improve pesticide mineralization. Microcosms containing a mixture of peat, straw and either linuron-primed soil or nonprimed soil were irrigated with clean or linuron-contaminated water. The lag time of linuron mineralization, recorded for microcosm samples, was indicative of the dynamics of the linuron-mineralizing biomass in the system. Bioaugmentation with linuron-primed soil immediately resulted in the establishment of a linuron-mineralizing capacity, which increased in size when fed with the pesticide. Also, microcosms containing nonprimed soil developed a linuron-mineralizing population, but after extended linuron feeding. Additional experiments showed that linuron-mineralization only developed with some nonprimed soils. Concomitant with the increase in linuron degradation capacity, targeted PCR-denaturing gradient gel electrophoresis showed the proliferation of a Variovorax phylotype related to the linuron-degrading Variovorax sp. SRS16 in microcosms containing linuron-primed soil, suggesting the involvement of Variovorax in linuron degradation. The correlation between the appearance of specific Variovorax phylotypes and linuron mineralization capacity was less clear in microcosms containing nonprimed soil. The data indicate that supplementation of pesticide-primed soil results in the establishment of pesticide-mineralizing populations in a BPS matrix with more certainty and more rapidly than the addition of nonprimed soil.

Rapid mineralization of the phenylurea herbicide diuron by Variovorax sp. strain SRS16 in pure culture and within a two-member consortium.

The phenylurea herbicide diuron [N-(3,4-dichlorophenyl)-N,N-dimethylurea] is widely used in a broad range of herbicide formulations, and consequently, it is frequently detected as a major water contaminant in areas where there is extensive use. We constructed a linuron [N-(3,4-dichlorophenyl)-N-methoxy-N-methylurea]- and diuron-mineralizing two-member consortium by combining the cooperative degradation capacities of the diuron-degrading organism Arthrobacter globiformis strain D47 and the linuron-mineralizing organism Variovorax sp. strain SRS16. Neither of the strains mineralized diuron alone in a mineral medium, but combined, the two strains mineralized 31 to 62% of the added [ring-U-(14)C]diuron to (14)CO(2), depending on the initial diuron concentration and the cultivation conditions. The constructed consortium was used to initiate the degradation and mineralization of diuron in soil without natural attenuation potential. This approach led to the unexpected finding that Variovorax sp. strain SRS16 was able to mineralize diuron in a pure culture when it was supplemented with appropriate growth substrates, making this strain the first known bacterium capable of mineralizing diuron and representatives of both the N,N-dimethyl- and N-methoxy-N-methyl-substituted phenylurea herbicides. The ability of the coculture to mineralize microgram-per-liter levels of diuron was compared to the ability of strain SRS16 alone, which revealed the greater extent of mineralization by the two-member consortium (31 to 33% of the added [ring-U-(14)C]diuron was mineralized to (14)CO(2) when 15.5 to 38.9 mug liter(-1) diuron was used). These results suggest that the consortium consisting of strains SRS16 and D47 could be a promising candidate for remediation of soil and water contaminated with diuron and linuron and their shared metabolite 3,4-dichloroaniline.

Riparian deforestation, stream narrowing, and loss of stream ecosystem services.

A study of 16 streams in eastern North America shows that riparian deforestation causes channel narrowing, which reduces the total amount of stream habitat and ecosystem per unit channel length and compromises in-stream processing of pollutants. Wide forest reaches had more macroinvertebrates, total ecosystem processing of organic matter, and nitrogen uptake per unit channel length than contiguous narrow deforested reaches. Stream narrowing nullified any potential advantages of deforestation regarding abundance of fish, quality of dissolved organic matter, and pesticide degradation. These findings show that forested stream channels have a wider and more natural configuration, which significantly affects the total in-stream amount and activity of the ecosystem, including the processing of pollutants. The results reinforce both current policy of the United States that endorses riparian forest buffers as best management practice and federal and state programs that subsidize riparian reforestation for stream restoration and water quality. Not only do forest buffers prevent nonpoint source pollutants from entering small streams, they also enhance the in-stream processing of both nonpoint and point source pollutants, thereby reducing their impact on downstream rivers and estuaries.