Operational stability to changes in composition of herbicide mixtures fed to a laboratory-scale biobarrier.

The main objective of this work was to evaluate the operational stability of a laboratory-scale aerobic biobarrier designed for the treatment of water contaminated by mixtures of three herbicides frequently found in agricultural runoffs, atrazine, simazine and 2,4-dichlorophenoxyacetic acid (2,4-D). The microbial consortium used to degrade the herbicides was composed by six cultivable bacterial strains, identified as members of the genera Variovorax, Sphingopyxis, Hydrocarboniphaga, Methylobacterium, Pseudomonas and Acinetobacter. The effect caused by a seventh member of the microbial consortium, a ciliated protozoa of the genus Colpoda, on the herbicides biodegradation kinetics, was also evaluated. The biodegradation of five combinations of the herbicides 2,4-D, atrazine and simazine was studied in the biobarrier, operated in steady state continuous culture at different volumetric loading rates. In all cases, removal efficiencies determined by chemical oxygen demand (COD) and HPLC were nearly 100 %. These results, joined to the null accumulation of aromatic byproducts of atrazine and simazine catabolism, show that after 495 days of operation, in the presence of the protozoa, the adaptability of the microbial consortium to changing environmental conditions allowed the complete removal of the mixture of herbicides.

Degradation kinetics of 4-amino naphthalene-1-sulfonic acid by a biofilm-forming bacterial consortium under carbon and nitrogen limitations.

By decolorization of azo dyes, caused by reductive cleavage of the azo linkage, toxic or recalcitrant amines are generated. The present study deals with the effect of the inflowing medium composition (C:N ratio) on the kinetic behavior of a bacterial biofilm-forming consortium, able to use as carbon, nitrogen and sulfur source, the molecule of 4-aminonaphthalene-1-sulfonic acid (4ANS), which is one of the most recalcitrant byproducts generated by decolorization of azo dyes. All the experiments were carried out at room temperature in a lab-scale packed-bed biofilm reactor. Because environmental conditions affect the bioreactor performance, two mineral salts media containing 4ANS, with distinct C:N ratios; 0.68 (carbon as the limiting nutrient) and 8.57 (nitrogen as the limiting nutrient) were used to evaluate their effect on 4ANS biodegradation. By HPLC and COD measurements, the 4ANS removal rates and removal efficiencies were determined. The cultivable bacterial strains that compose the consortium were identified by their 16S rDNA gene sequence. With the enrichment technique used, a microbial consortium able to use efficiently 4ANS as the sole carbon source and energy, nitrogen and sulfur, was selected. The bacterial strains that constitute the consortium were isolated and identified. They belong to the following genera: Bacillus, Arthrobacter, Microbacterium, Nocardioides, and Oleomonas. The results obtained with this consortium showed, under nitrogen limitation, a remarkable increase in the 4ANS removal efficiency η(ANS), and in the 4ANS volumetric removal rates R (V,4ANS), as compared to those obtained under carbon limitation. Differences observed in bioreactor performance after changing the nutrient limitation could be caused by changes in biofilm properties and structure.

Cyanuric acid biodegradation by a mixed bacterial culture of Agrobacterium tumefaciens and Acinetobacter sp. in a packed bed biofilm reactor.

Cyanuric acid (1,3,5-triazine-2,4,6-triol [OOOT]) is a common biodegradation byproduct of triazinic herbicides, frequently accumulated in soils or water when supplementary carbon sources are absent. A binary bacterial culture able to degrade OOOT was selected through a continuous selection process accomplished in a chemostat fed with a mineral salt (MS) medium containing cyanuric acid as the sole carbon and nitrogen source. By sequence comparison of their 16S rDNA amplicons, bacterial strains were identified as Agrobacterium tumefaciens, and Acinetobacter sp. When the binary culture immobilized in a packed bed reactor (PBR) was fed with MS medium containing OOOT (50 mg L(-1)), its removal efficiencies were about 95%; when it was fed with OOOT plus glucose (120 mg L(-1)) as a supplementary carbon source, its removal efficiencies were closer to 100%. From sessile cells, attached to PBR porous support, or free cells present in the outflowing medium, DNA was extracted and used for Random Amplification of Polymorphic DNA analysis. Electrophoretic patterns obtained were compared to those of pure bacterial strains, a clear predominance of A. tumefaciens in PBR was observed. Although in continuous suspended cell culture, a stable binary community could be maintained, the attachment capability of A. tumefaciens represented a selective advantage over Acinetobacter sp. in the biofilm reactor, favoring its predominance in the porous stone support.

Biodegradation of 2,4,6-trichlorophenol in a packed-bed biofilm reactor equipped with an internal net draft tube riser for aeration and liquid circulation.

For the aerobic biodegradation of the fungicide and defoliant 2,4,6-trichlorophenol (2,4,6-TCP), a bench-scale packed-bed bioreactor equipped with a net draft tube riser for liquid circulation and oxygenation (PB-ALR) was constructed. To obtain a high packed-bed volume relative to the whole bioreactor volume, a high A(D)/A(R) ratio was used. Reactor's downcomer was packed with a porous support of volcanic stone fragments. PB-ALR hydrodynamics and oxygen mass transfer behavior was evaluated and compared to the observed behavior of the unpacked reactor operating as an internal airlift reactor (ALR). Overall gas holdup values epsilon(G), and zonal oxygen mass transfer coefficients determined at various airflow rates in the PB-ALR, were higher than those obtained with the ALR. When comparing mixing time values obtained in both cases, a slight increment in mixing time was observed when reactor was operated as a PB-ALR. By using a mixed microbial community, the biofilm reactor was used to evaluate the aerobic biodegradation of 2,4,6-TCP. Three bacterial strains identified as Burkholderia sp., Burkholderia kururiensis and Stenotrophomonas sp. constituted the microbial consortium able to cometabolically degrade the 2,4,6-TCP, using phenol as primary substrate. This consortium removed 100% of phenol and near 99% of 2,4,6-TCP. Mineralization and dehalogenation of 2,4,6-TCP was evidenced by high COD removal efficiencies ( approximately 95%), and by the stoichiometric release of chloride ions from the halogenated compound ( approximately 80%). Finally, it was observed that the microbial consortium was also capable to metabolize 2,4,6-TCP without phenol as primary substrate, with high removal efficiencies (near 100% for 2,4,6-TCP, 92% for COD and 88% for chloride ions).

Chemostat selection of a bacterial community able to degrade s-triazinic compounds: continuous simazine biodegradation in a multi-stage packed bed biofilm reactor.

Using a successive transfer method on mineral salt medium containing simazine, a microbial community enriched with microorganisms able to grow on simazine was obtained. Afterwards, using a continuous enrichment culture procedure, a bacterial community able to degrade simazine from an herbicide formulation was isolated from a chemostat. The continuous selector, fed with a mineral salt medium containing simazine and adjuvants present in the commercial herbicide formulation, was maintained in operation for 42 days. Following the lapse of this time, the cell count increased from 5 x 10(5) to 3 x 10(8) CFU mL(-1), and the simazine removal efficiency reached 96%. The chemostat's bacterial diversity was periodically evaluated by extracting the culture's bacterial DNA, amplifying their 16S rDNA fragments and analyzing them by thermal gradient gel electrophoresis. Finally, a stable bacterial consortium able to degrade simazine was selected. By PCR amplification, sequencing of bacterial 16S rDNA amplicons, and comparison with known sequences of 16S rDNA from the NCBI GenBank, eight bacterial strains were identified. The genera, Ochrobactrum, Mycobacterium, Cellulomonas, Arthrobacter, Microbacterium, Rhizobium and Pseudomonas have been reported as common degraders of triazinic herbicides. On the contrary, we were unable to find reports about the ability of the genus Pseudonocardia to degrade triazinic compounds. The selected bacterial community was attached to a porous support in a concurrently aerated four-stage packed-bed reactor fed with the herbicide. Highest overall simazine removal efficiencies eta (SZ) were obtained at overall dilution rates D below 0.284 h(-1). However, the multistage packed bed reactor could be operated at dilution rates as high as D = 3.58 h(-1) with overall simazine removal volumetric rates R (v,SZ) = 19.6 mg L(-1) h(-1), and overall simazine removal specific rates R (X,SZ) = 13.48 mg (mg cell protein)(-1) h(-1). Finally, the consortium's ability to degrade 2-chloro-4,6-diamino-1,3,5-triazine (CAAT), cyanuric acid and the herbicide atrazine, pure or mixed with simazine, was evaluated in fed batch processes.