Gabriela Briceño

Atrazine dissipation and its impact on the microbial communities and community level physiological profiles in a microcosm simulating the biomixture of on-farm biopurification system

The effects of repeated atrazine application (40 mg a.i.kg(-1)) on its degradation, microbial communities and enzyme activities were studied in a peat based biomixture composed by straw, soil and peat in the volumetric proportions of 2:1:1 that can be used in on-farm biopurification system. Atrazine removal efficiency was high (96%, 78% and 96%) after each atrazine application and did not show a lag phase. Microbial enzyme activities were reduced significantly with atrazine application but rapidly recovered. Microbial diversity obtained by BiologEcoplate was similar after the first and second atrazine application. However, an inhibitory effect was observed after the third application. After each atrazine application, culturable fungi were reduced, but rapidly recovered without significant changes in culturable bacteria and actinomycetes compared to the control. Denaturing gradient gel electrophoresis (DGGE) patterns demonstrated that microbial community structure remained relatively stable in time when compared to the controls. In conclusion, our results demonstrated that after successive ATZ applications, the peat based biomixture had a good degradation capacity. Moreover, microbiological assays demonstrated the robustness of the peat based biomixture from a microbiological point of view to support pesticide degradation.

Carbendazim dissipation in the biomixture of on-farm biopurification systems and its effect on microbial communities.

The impact of repeated carbendazim (CARB) applications on the extent of CARB dissipation, the microbial diversity, the community level physiological profile (CLPP), and the enzymatic activity within the biomixture of an on-farm biopurification system was evaluated. After three successive CARB applications, the CARB dissipation efficiency was high; the efficiency of dissipation was 87%, 94% and 96% after each application, respectively. Although microbial enzymatic activity was affected significantly by CARB application, it could recover after each CARB pulse. Likewise, the numbers of cultivable bacteria, fungi and actinomycetes (as measured in CFUs) were slightly affected by the addition of CARB, but the inhibitory effect of the pesticide application was temporary. Denaturing gradient gel electrophoresis (DGGE) and Biolog Ecoplate assays demonstrated that the microbial populations remained relatively stable over time when compared to the control. The results obtained herein therefore demonstrate the high dissipation capacity of this biomixture and highlight the microbiological robustness of this biological system.

Enhanced removal of a pesticides mixture by single cultures and consortia of free and immobilized Streptomyces strains.

Pesticides are normally used to control specific pests and to increase the productivity in crops; as a result, soils are contaminated with mixtures of pesticides. In this work, the ability of Streptomyces strains (either as pure or mixed cultures) to remove pentachlorophenol and chlorpyrifos was studied. The antagonism among the strains and their tolerance to the toxic mixture was evaluated. Results revealed that the strains did not have any antagonistic effects and showed tolerance against the pesticides mixture. In fact, the growth of mixed cultures was significantly higher than in pure cultures. Moreover, a pure culture (Streptomyces sp. A5) and a quadruple culture had the highest pentachlorophenol removal percentages (10.6% and 10.1%, resp.), while Streptomyces sp. M7 presented the best chlorpyrifos removal (99.2%). Mixed culture of all Streptomyces spp. when assayed either as free or immobilized cells showed chlorpyrifos removal percentages of 40.17% and 71.05%, respectively, and for pentachlorophenol 5.24% and 14.72%, respectively, suggesting better removal of both pesticides by using immobilized cells. These results reveal that environments contaminated with mixtures of xenobiotics could be successfully cleaned up by using either free or immobilized cultures of Streptomyces, through in situ or ex situ remediation techniques.

Effect of liquid cow manure on andisol properties and atrazine adsorption.

Application of animal manure amendments to agricultural soils is a common practice to improve soil fertility through the addition of essential plant nutrients. This practice may increase the potential for atrazine (2-chloro-4-ethylamino-6-isopropylamino-1, 3, 5-triazine) leaching due to competition for adsorption sites between the pesticide and dissolved organic carbon (DOC) added through manure. We evaluated the influence of liquid cow manure (LCM) application on soil properties, atrazine adsorption, and the physicochemical controlling mechanisms in an Andisol. The LCM was applied at rates equivalent to 0, 100,000, 200,000, and 300,000 L ha(-1), resulting in treatments S-0, S-100, S-200, and S-300, respectively. The LCM application increased DOC and pH of the soils immediately on addition, but pH returned to S-0 values 30 d after application. The LCM application did not modify atrazine adsorption with the two lowest application rates (S-100 and S-200), but atrazine adsorption was decreased in S-300 (K(f) = 0.96) compared with the control (S-0) (K(f) = 1.19), possibly due to the competitive adsorption of DOC with the pesticide. The Fourier-transformed infrared analysis showed that LCM increased aliphaticity and presence of N-containing groups and polysaccharide-like groups in amended soils; however, these properties did not modify the atrazine interaction in the studied amended soils. Interestingly the addition of DOC to soil at the high application rate (S-300) reduced atrazine adsorption in this rich OM Andisol despite the LCM not raising the concentration of stable organic matter. The application of high rates of liquid manure containing DOC incurs an increased risk of pesticide leaching.

Effect of dairy manure rate and the stabilization time of amended soils on atrazine degradation

The application rate of liquid cow manure (LCM) in the field and the stabilization time of amended soils before application of pre-plant herbicides are factors that determine their efficiency. This study includes evaluation of residual atrazine (2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine) in soil and amended soils with equivalent rate of 100,000; 200,000; and 300,000 L ha(-1) of LCM and the effect of pre-incubation time of amended soils on atrazine degradation. The study was carried out under controlled conditions using an Andisol with previous historical application of atrazine. The respiratory activity and fluorescein diacetate (FDA) studies indicated that the time necessary for stabilization of amended soils is over 20-30 d. During the measurement of respiratory and FDA activity, no significant differences were observed when atrazine was applied. The half-life of atrazine ranged from 5 to 8d and the relative distribution of degradation products seem to be affected by the application of LCM. The pre-incubation time of amended soil and LCM dose would not affect atrazine degradation rate, when the soil has a history of herbicide application. However, repeated applications of LCM in a long period of time could change the soil pH and increase the content of dissolved organic carbon (DOC) which could further contribute to a faster degradation of atrazine. Both effects would reduce the effectiveness of atrazine in weed control.

Effect of cow slurry amendment on atrazine dissipation and bacterial community structure in an agricultural Andisol.

Atrazine is a commonly used herbicide for maize production in Chile, but it has recently been shown to be ineffective in soils that receive applications of cow slurries generated from the dairy industry. This effect may be caused either by the sorption of the pesticide to organic matter or more rapid degradation in slurry-amended soils. The objectives of this study were to evaluate the effects of cow slurry on atrazine dissipation, the formation of atrazine metabolites and the modification of bacterial community in Andisol. The cow slurry was applied at doses of 100,000-300,000 Lha(-1). After 4 weeks, atrazine was applied to the slurry-amended soils at concentrations of 1-3 mg kg(-1). The amounts of atrazine and its metabolites were determined by high performance liquid chromatography (HPLC). The soil microbial community was monitored by measurement of CO(2) evolution and changes in bacterial community using PCR-DGGE of 16S rRNA genes. The results show that cow slurry applications had no effect on atrazine dissipation, which had a half-life of 15-19 days. The atrazine metabolites were detected after 20 days and were significantly higher in soils amended with the slurry at both 20 and 40 days after application of the herbicide. Respiration rates were elevated after 10 days in all soils with atrazine addition. Both the atrazine and slurry amendments altered the bacterial community structures, indicated by the appearance of specific bands in the DGGE gels after 10 days. Cloning and sequencing of the 16S rRNA genes from the DGGE gels showed that the bands represented various genera of beta-proteobacteria that appeared in response to atrazine. According to our results, further field studies are required to explain the lower effectiveness of atrazine in weed control. These studies may include the effect of dissolved organic carbon on the atrazine mobility.

Removal of the insecticide diazinon from liquid media by free and immobilized Streptomyces sp. isolated from agricultural soil.

From an agricultural soil that had received continuous applications of organophosphorus pesticides, 30 actinobacteria strains were isolated. Two strains, identified as Streptomyces sp. AC1‐6 and Streptomyces sp. ISP4, were selected because of their tolerance to diazinon and based on the relationship between diazinon removal and microbial growth. In liquid medium with diazinon at concentrations of 25 and 50 mg L−1, both strains were able to remove approximately 40–50% and 70–90% of the initial diazinon after 24 and 96 h of incubation, respectively. This diazinon removal was accompanied by microbial growth of the strains, an initial pH decrease, and glucose consumption in the liquid medium. Evaluation of the diazinon removal achieved by the free actinobacteria and Streptomyces sp. AC1‐6 immobilized on alginate beads revealed that the immobilized cells exhibited a 60% higher diazinon removal compared with the free cells. The reusability of the encapsulated biomass was confirmed, and a diazinon removal rate of more than 50% was obtained after the second batch. This work constitutes one of the few reports that describe Streptomyces strains as diazinon degraders. Given the high diazinon removal found, the streptomycetes exhibit suitable potential as diazinon‐degrading actinobacteria for elimination of diazinon from liquid residues.

USE OF SLUDGE FROM KRAFT MILL WASTEWATER TREATMENT AS IMPROVER OF VOLCANIC SOILS: EFFECT ON SOIL BIOLOGICAL PARAMETERS

Sludge from kraft mill wastewater treatment was applied on two soils derived from volcanic ashes and, the effect on biological parameters of the soils was evaluated. The soils used in this study were an Andisol belonging of Gorbea Series and an Ultisol belonging of Collipulli Series. The sludge was added at rates of 0, 10, 20, 30 and 50 t ha -1 . Previously, germination of red clover (Trifolium pratense), white clover (Trifolium repens), alfalfa (Medicago sativa) and ryegrass (Lolium perenne) was tested to assess phytotoxicity of the sludge. The different soil-sludge mixtures were incubated at controlled temperature for 60 days and microbial respiration, microbial biomass carbon, fluorescein diacetate hydrolysis and acid phosphatase activity were evaluated throughout the incubation time. All biological parameters evaluated were sensitive enough to shown the effect of sewage sludge application on soil microorganisms. The sludge application at different rates increased significantly (P<0.05) the microbial activity and enzymatic activity of the sludge amended soils. The maximum levels of activity were observed between 15 and 30 days after sludge application in both Gorbea soil and Collipulli soil, obtaining the highest values when were applied between 30 and 50 t ha -1 . The results of this study suggest that sludge from kraft mill wastewater treatment may have potential as a beneficial soil amendment for improving biological properties of the soils.

Rhizosphere effect on pesticide degradation in biobeds under different hydraulic loads

Interactions between microorganisms and root exudates in a biobed system with vegetal (grass) cover could enhance pesticide degradation. Otherwise, a high water load may generate high concentrations of pesticides in lixiviates. We studied the effect of the vegetal cover on the degradation of a mixture of atrazine (ATZ), chlorpyrifos (CHL) and iprodione (IPR) (35 mg L -1 each) in a biobed system operated with two different hydraulic loads (0.6 and 1.2 L of tap water per day). The concentration of the pesticides and their main metabolites were measured in the lixiviates during 60 days, as well as in the biomixtures at the end of the study. Dehydrogenase activity in the biomixtures and organic acid exudation from the vegetal cover were also analysed. The vegetal cover diminished the lixiviation of pesticides and their metabolites mainly at the lower hydraulic load used. The degradation of the pesticides was high (>95%) and increased in biobeds with vegetal cover and low hydraulic load. Degradation metabolites of CHL and IPR were formed during pesticide degradation; however they were degraded in the biobed and were not detected in lixiviates at the end of the study. In general, an increase in organic acid exudation by vegetal cover was observed caused by chemical stress after pesticide application. The increase was similar at both hydraulic loads. Efficient colonisation of wheat straw by fungi was observed by confocal microscopy.

Volatiles from white-rot fungi for controlling plant pathogenic fungi

Fungal volatile secondary metabolites play important roles in mediating antagonistic and beneficial interactions among organisms. The effects of eight strains of white-rot fungi cultured on potato waste against the mycelial growth of Botrytis cinerea, Fusarium oxysporum and Mucor miehei were investigated using a bi-compartmented Petri dish assay. The chemical composition of volatile organic compounds released from the strain with the highest inhibitory effect was also investigated by headspace solid-phase microextraction and gas chromatography/mass spectrometry analysis. Of the eight white-rot fungi evaluated, Anthracophyllum discolor. Sp4 showed a high inhibitory activity against M. miehei (approximately 76%) and B. cinerea (approximately 20%). F. oxysporum was inhibited to a lesser extent (approximately 10%) by A. discolor and T. versiscolor. The gas chromatography/mass spectrometry analysis showed nine main volatile compounds released from A. discolor Sp4, among them are the sesquiterpenesα-bisabolene and bulnesene, and the chlorinated aromatic compounds 1,5-dichloro-2,3-dimethoxybenzene, 3,5-dichloro-4-methoxybenzaldehyde and 3-chloro-4-methoxybenzaldehyde. Some of which have been reported previously with antimicrobial activity. The antifungal activity and volatile profile of A. discolor have not been previously reported.