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.     

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

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 monitored in response to the application of linuron and to environmental perturbations in agricultural soil microcosms and microcosms 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 linuron mineralization capacity. In the soil microcosms and one biopurification system setup, libA numbers responded to the application of linuron and environmental changes in congruency with the modulation of linuron mineralization capacity and the occurrence of a particular Variovorax phylotype (phylotype A). However, in another biopurification system setup, no such correlations were found. Our data suggest that in the simulated environmental settings, the occurrence of libA can be linked to the linuron mineralization 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 linuron mineralization in the environment.     

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.     

Inoculation with a Mixed Degrading Culture Improves the Pesticide Removal of an On-Farm Biopurification System

To investigate whether the pesticide removal in on-farm biopurification systems (BPS) filled with two different types of substrata (biomix and plastic carriers) is affected by inoculation with a pesticide-degrading strain or mixed culture, lab-scale BPS used to treat chloropropham point source contaminations were bioaugmented with either a specialized chloropropham-degrading strain or a chloropropham-degrading enrichment culture. Application of both inoculum types leads to an accelerated degradation activity in the columns filled with plastic carriers. For both substratum types, inoculation with the mixed culture resulted in a lower breakthrough of the toxic intermediate 3-chloroaniline at high hydraulic loads, compared to inoculation with the pure isolate and no inoculation. This study suggests that the use of plastic carrier materials could be a proficient alternative to the use of a conventional biomix as a substratum in on-farm BPS and that inoculation with a mixed degrading culture can reduce the leaching of more mobile toxic intermediates.     

Constitutive mineralization of low concentrations of the herbicide linuron by a Variovorax sp. strain

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.     

Effect of propanil,linuron, and dicamba on the degradation kinetics of 2,4‐dichlorophenoxyacetic acid by Burkholderia sp. A study by differential analysis of 2,4‐dichlorophenoxyacetic acid degradation data

The successive application of distinct pesticides, or mixtures of them, is a frequent practice that could adversely affect the microbial species inhabiting soil and aquatic ecosystems. The ability of soil or aquatic microbiota to degrade a pesticide could be affected by the presence of another. If the degradation rate of the first compound is inhibited, its dissipation half‐life in the environment could be hazardously enlarged. Few studies have been made to quantify the impact on the biodegradation rate of pesticides in soils or water by the presence of other pesticides. In this work, a method for assessing the effect of a pesticide on the biodegradation rate of another, measuring its effect on the biodegradation kinetics of a single bacterial strain is presented. The mathematical analysis is a powerful tool to study the stoichiometry and kinetics of microbial processes, which was used to evaluate independently, in detail, the effect of three pesticides (propanil, linuron, and dicamba) on the biodegradation kinetics of 2,4‐dichlorophenoxyacetic acid by a strain of Burkholderia sp. It was evidenced that linuron and dicamba caused a decay of more than 40% in the top instantaneous degradation rate of 2,4‐dichlorophenoxyacetic acid, while propanil showed a minimal effect.     

Effects of temperature on microbial utilization of lignocellulosic detritus in a thermally impacted stream

The effects of temperature on rates of mineralization of [¹⁴C]lignocellulose were investigated in water and sediment from a thermally impacted stream and from a nearby unimpacted swamp at the Savannah River Plant, South Carolina. The temperature optimum for lignocellulose mineralization remained near 35°C at the unimpacted site throughout the sampling period from November 1986 to May 1987. The temperature optimum for lignocellulose mineralization in the thermally impacted stream was near 45°C when thermal effluents from a nuclear reactor were released to the stream, and was near 35°C when the reactor was not operating. Microbial populations capable of rapidly degrading lignocellulose at higher temperatures (45–55°C) developed between 9 and 27 days under conditions of thermal stress, indicating that under favorable conditions thermophilic microorganisms became dominant components of the microbiota. Removal of thermal stress for periods of 75 days or less resulted in a collapse of the thermophilic degrading population.     

Interannual Invariability of Forest Evapotranspiration and Its Consequence to Water Flow Downstream

Although drought in temperate deciduous forests decreases transpiration rates of many species, stand-level transpiration and total evapotranspiration is often reported to exhibit only minor interannual variability with precipitation. This apparent contradiction was investigated using four years of transpiration estimates from sap flux, interception–evaporation estimates from precipitation and throughfall gauges, modeled soil evaporation and drainage estimates, and eddy covariance data in a mature oak-hickory forest in North Carolina, USA. The study period included one severe drought year and one year of well above-average precipitation. Normalized for atmospheric conditions, transpiration rates of some species were lower in drought than in wet periods whereas others did not respond to drought. However, atmospheric conditions during drought periods are unlike conditions during typical growing season periods. The rainy days that are required to maintain drought-free periods are characterized by low atmospheric vapor pressure deficit, leading to very low transpiration. In contrast, days with low air vapor pressure deficit were practically absent during drought and moderate levels of transpiration were maintained throughout despite the drying soil. Thus, integrated over the growing season, canopy transpiration was not reduced by drought. In addition, high vapor pressure deficit during drought periods sustained appreciable soil evaporation rates. As a result, despite the large interannual variation in precipitation (ranging from 934 to 1346 mm), annual evapotranspiration varied little (610–668 mm), increasing only slightly with precipitation, due to increased canopy rainfall interception. Because forest evapotranspiration shows only modest changes with annual precipitation, lower precipitation translates to decreased replenishment of groundwater and outflow, and thus the supply of water to downstream ecosystems and water bodies.     

土壤有效硅对大豆生长发育和生理功能的影响

人工调节土壤有效硅含量及盆栽试验,研究土壤有效硅对大豆生长发育和生理功能的影响.结果表明,土壤有效硅含量在55.1～202.8mg·kg^-1范围内,随着土壤有效硅含量的提高,大豆种子萌发过程中蛋白酶和脂肪酶活性升高,淀粉酶活性无显著变化,呼吸速率加快,种子活力升高,萌发速度加快,种子萌发率无显著变化;幼苗生长过程中叶片叶绿素含量无显著变化,光合速率加快,根系活力、硝酸还原酶活力升高,蒸腾强度减弱,水分利用效率和叶含水量升高,抗旱保水能力提高.大豆幼苗含硅量与土壤有效硅含量呈线性正相关趋势(r=0.994).土壤有效硅含量大于202.8mg·kg^-1,生理功能不再显著变化,说明土壤中的硅被大豆吸收后,改善了大豆萌发种子和幼苗的生理功能,使种子萌发和幼苗生长加快.     

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

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.     

Quantifying the Contribution of Entire Free-Living Nematode Communities to Carbon Mineralization under Contrasting C and N Availability

To understand the roles of nematodes in organic matter (OM) decomposition, experimental setups should include the entire nematode community, the native soil microflora, and their food sources. Yet, published studies are often based on either simplified experimental setups, using only a few selected species of nematode and their respective prey, despite the multitude of species present in natural soil, or on indirect estimation of the mineralization process using O₂ consumption and the fresh weight of nematodes. We set up a six-month incubation experiment to quantify the contribution of the entire free living nematode community to carbon (C) mineralization under realistic conditions. The following treatments were compared with and without grass-clover amendment: defaunated soil reinoculated with the entire free living nematode communities (+Nem) and defaunated soil that was not reinoculated (-Nem). We also included untreated fresh soil as a control (CTR). Nematode abundances and diversity in +Nem was comparable to the CTR showing the success of the reinoculation. No significant differences in C mineralization were found between +Nem and -Nem treatments of the amended and unamended samples at the end of incubation. Other related parameters such as microbial biomass C and enzymatic activities did not show significant differences between +Nem and -Nem treatments in both amended and unamended samples. These findings show that the collective contribution of the entire nematode community to C mineralization is small. Previous reports in literature based on simplified experimental setups and indirect estimations are contrasting with the findings of the current study and further investigations are needed to elucidate the extent and the mechanisms of nematode involvement in C mineralization.     

雪被去除对川西亚高山云杉林土壤活性氮的影响

气候变化引发的季节性雪被改变可能对高寒森林土壤氮循环产生深刻影响.以遮雪棚去除雪被,研究了雪被去除样方和对照样方在不同关键时期(雪被初期、深雪被期和雪被融化期)土壤氮库和矿化速率的变化.结果表明: 季节性雪被对土壤具有良好的保温作用,雪被去除使得5 cm深度土壤平均温度和最低温度分别降低0.33和1.17 ℃,并明显增加了土壤冻结深度和冻融循环.土壤活性氮在不同雪被时期存在显著差异.雪被去除使得冬季土壤铵态氮、硝态氮和可溶性有机氮增加38.6%、23.5%和57.3%.此外,雪被去除也促进了融化期土壤硝化和矿化速率的增加.因此,未来气候变暖引起的雪被减少可能加快川西亚高山森林冬季土壤氮循环.     

污染土壤中有机磷农药降解菌的分离及其多样性

采用添加有机磷农药的选择性培养基,在长期受有机磷农药污染的土壤中分离到7株有机磷农药降解菌,经生理生化鉴定和系统发育分析,菌株mp-1鉴定为Pseudaminobactersp.,菌株mp-2为Alcaligenessp.,菌株mp-7为Brucellasp.,其他菌株为Ochrobactrumsp.。16SrDNA序列同源性比较、系统发育分析和染色体ERIC-PCR指纹图谱扩增表明有机磷农药长期污染的土壤中有机磷农药降解菌具有丰富的多样性。     

The quantity and quality of dissolved organic matter as supplementary carbon source impacts the pesticide-degrading activity of a triple-species bacterial biofilm

Effects of environmental dissolved organic matter (eDOM) that consists of various low concentration carbonic compounds on pollutant biodegradation by bacteria are poorly understood, especially when it concerns synergistic xenobiotic-degrading consortia where degradation depends on interspecies metabolic interactions. This study examines the impact of the quality and quantity of eDOM, supplied as secondary C-source, on the structure, composition and pesticide-degrading activity of a triple-species bacterial consortium in which the members synergistically degrade the phenylurea herbicide linuron, when grown as biofilms. Biofilms developing on 10 mg L^?1 linuron showed a steady-state linuron degradation efficiency of approximately 85 %. The three bacterial strains co-localized in the biofilms indicating syntrophic interactions. Subsequent feeding with eDOM or citrate in addition to linuron resulted into changes in linuron-degrading activity. A decrease in linuron-degrading activity was especially recorded in case of co-feeding with citrate and eDOM of high quality and was always associated with accumulation of the primary metabolite 3,4-dichloroaniline. Improvement of linuron degradation was especially observed with more recalcitrant eDOM. Addition of eDOM/citrate formulations altered biofilm architecture and species composition but without loss of any of the strains and of co-localization. Compositional shifts correlated with linuron degradation efficiencies. When the feed was restored to only linuron, the linuron-degrading activity rapidly changed to the level before the mixed-substrate feed. Meanwhile only minor changes in biofilm composition and structure were recorded, indicating that observed eDOM/citrate effects had been primarily due to repression/stimulation of linuron catabolic activity rather than to biofilm characteristics.     

HylA,an Alternative Hydrolase for Initiation of Catabolism of the Phenylurea Herbicide Linuron in Variovorax sp. Strains

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 K_m, 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.     

草甸草原土壤碳/氮矿化潜力及土壤微生物水分敏感性对极端干旱的响应

土壤碳、氮矿化是生态系统养分循环的关键过程,受到水分供给的强烈影响。本研究对极端干旱处理(连续3年生长季减少66%降水)的内蒙古草甸草原野外取土,采用超低温冻干后再调节土壤水分至3%、8%、13%、18%、25%和35% 6个水平进行室内培养,研究极端干旱处理后土壤碳/氮矿化潜力以及土壤微生物对水分变化的敏感性。结果表明: 与对照(自然降雨)相比,极端干旱处理后,6个培养水平的平均土壤氮矿化潜力显著提高14.2%,但未显著影响土壤碳矿化潜力。极端干旱显著提高土壤微生物生物量氮和土壤可溶性有机碳26.8%和26.9%。无论是对照还是极端干旱处理,土壤氮矿化潜力、碳矿化潜力和微生物生物量碳和氮均随着土壤含水量增加而增加,而可溶性有机碳从较低水分的培养水平(3%和8%)到较高水分的培养水平(>13%)显著降低,表明底物的扩散起到重要作用。极端干旱处理显著提高了碳矿化初始脉冲强度,表明极端干旱提高了土壤微生物对水分的敏感性。极端干旱显著降低了土壤碳矿化潜力/氮矿化潜力的比值,表明长期干旱可能会降低土壤碳、氮循环过程的耦合作用。极端干旱对土壤碳矿化和氮矿化过程的影响存在差异,激发了土壤微生物对水分的敏感性,弱化了碳、氮循环过程的耦合关系,并进一步影响中国北方草甸草原生态系统的生物地球化学循环过程及草地生产力。     

Reappraisal of drying and wetting effects on C and N mineralization and fluxes in soils

In the next decades, many soils will be subjected to increased drying/wetting cycles or modified water availability considering predicted global changes in precipitation and evapotranspiration. These changes may affect the turnover of C and N in soils, but the direction of changes is still unclear. The aim of the review is the evaluation of involved mechanisms, the intensity, duration and frequency of drying and wetting for the mineralization and fluxes of C and N in terrestrial soils. Controversial study results require a reappraisal of the present understanding that wetting of dry soils induces significant losses of soil C and N. The generally observed pulse in net C and N mineralization following wetting of dry soil (hereafter wetting pulse) is short‐lived and often exceeds the mineralization rate of a respective moist control. Accumulated microbial and plant necromass, lysis of live microbial cells, release of compatible solutes and exposure of previously protected organic matter may explain the additional mineralization during wetting of soils. Frequent drying and wetting diminishes the wetting pulse due to limitation of the accessible organic matter pool. Despite wetting pulses, cumulative C and N mineralization (defined here as total net mineralization during drying and wetting) are mostly smaller compared with soil with optimum moisture, indicating that wetting pulses cannot compensate for small mineralization rates during drought periods. Cumulative mineralization is linked to the intensity and duration of drying, the amount and distribution of precipitation, temperature, hydrophobicity and the accessible pool of organic substrates. Wetting pulses may have a significant impact on C and N mineralization or flux rates in arid and semiarid regions but have less impact in humid and subhumid regions on annual time scales. Organic matter stocks are progressively preserved with increasing duration and intensity of drought periods; however, fires enhance the risk of organic matter losses under dry conditions. Hydrophobicity of organic surfaces is an important mechanism that reduces C and N mineralization in topsoils after precipitation. Hence, mineralization in forest soils with hydrophobic organic horizons is presumably stronger limited than in grassland or farmland soils. Even in humid regions, suboptimal water potentials often restrict microbial activity in topsoils during growing seasons. Increasing summer droughts will likely reduce the mineralization and fluxes of C and N whereas increasing summer precipitation could enhance the losses of C and N from soils.     

模拟干旱和磷添加对热带低地雨林氮矿化过程的影响

土壤氮矿化作为氮转化的主要过程决定土壤供氮能力。热带森林生态系统往往受磷限制, 氮矿化过程对干旱的响应是否受磷限制的调控值得探讨。该研究以海南三亚甘什岭自然保护区热带低地雨林为研究对象, 利用2019年建立的林内穿透雨减少(50%)及磷添加双因素交互实验平台, 通过野外树脂芯原位培养法研究模拟干旱及磷添加对土壤无机氮(包括铵态氮和硝态氮)含量和氮矿化过程的影响。结果表明: 1)减雨处理显著降低了5和15 cm深度土壤的水分含量, 而对土壤温度没有显著影响。2)减雨处理和减雨与磷添加共同处理无论在旱季还是湿季对0-10 cm土壤无机氮含量均没有产生显著影响, 但磷添加处理在旱季显著降低了土壤硝态氮含量, 表明磷添加处理对氮有效性的影响主要体现在旱季, 而非湿季。3)干旱处理在旱季和湿季均显著降低了土壤净氨化速率和净氮矿化速率, 而磷添加处理和减雨与磷添加共同处理无论在旱季还是湿季对净氨化速率、净硝化速率和净氮矿化速率均没有产生显著影响, 结果表明了干旱能够显著降低土壤净氮矿化速率。4)土壤水分含量与土壤净氨化速率和净氮矿化速率呈显著正相关关系, 同时减雨处理显著影响了土壤净氨化速率与铵态氮含量的关系, 并且在铵态氮含量相等的情况, 随着干旱的影响净氨化速率下降得更快。这表明土壤水分含量变化是影响该研究样地土壤氮矿化的主要因素。上述研究结果说明, 降水变化对热带低地雨林中土壤氮矿化有重要影响, 短期磷添加没有显著影响, 减雨与磷添加对土壤氮矿化过程并没有交互效应。