Metabolic Profiling and Comparative Proteomic Insight in Respect of Amidases during Iprodione Biodegradation.

The fungicide iprodione (IPR) (3-(3,5-dichlorophenyl) N-isopropyl-2,4-dioxoimidazolidine-1-carboxamide) is a highly toxic compound. Although IPR has been restricted, it is still being applied in many places around the world, constituting an environmental risk. The biodegradation of IPR is an attractive option for reducing its residues. In this study, we isolated thirteen IPR-tolerant bacteria from a biopurification system designed to treat pesticides. A study of biodegradation using different strains was comparatively evaluated, and the best degradation rate of IPR was presented by Achromobacter sp. C1 with a half-life (T1/2) of 9 days. Based on a nano-LC-MS/MS analysis for the strains, proteins solely expressed in the IPR treatment were identified by highlighting the strain Achromobacter sp. C1, with 445 proteins primarily involved in the biosynthesis of secondary metabolites and microbial metabolism in diverse environments. Differentially expressed protein amidases were involved in six metabolic pathways. Interestingly, formamidase was inhibited while other cyclases, i.e., amidase and mandelamide hydrolase, were overexpressed, thereby minimizing the effect of IPR on the metabolism of strain C1. The dynamic changes in the protein profiles of bacteria that degrade IPR have been poorly studied; therefore, our results offer new insight into the metabolism of IPR-degrading microorganisms, with special attention paid to amidases.

Biosurfactant Production by Bacillus amyloliquefaciens C11 and Streptomyces lavendulae C27 Isolated from a Biopurification System for Environmental Applications.

Biosurfactant-producing bacteria can be found in contaminated environments such as biopurification systems (BPS) for pesticide treatments. A total of 18 isolates were screened to determine their ability to produce extracellular biosurfactants, using olive oil as the main carbon source. Out of the eighteen isolates, two strains (C11 and C27) were selected for biosurfactant production. The emulsification activities of the C11 and C27 strains using sunflower oil was 58.4 and 53.7%, respectively, and 46.6 and 48.0% using olive oil. Using molecular techniques and MALDI-TOF, the strains were identified as Bacillus amyloliquefaciens (C11) and Streptomyces lavendulae (C27). The submerged cultivation of the two selected strains was carried out in a 1 L stirred-tank bioreactor. The maximum biosurfactant production, indicated by the lowest surface tension measurement, was similar (46 and 45 mN/m) for both strains, independent of the fact that the biomass of the B. amyloliquefaciens C11 strain was 50% lower than the biomass of the S. lavendulae C27 strain. The partially purified biosurfactants produced by B. amyloliquefaciens C11 and S. lavendulae C27 were characterized as a lipopeptide and a glycolipid, respectively. These outcomes highlight the potential of the selected biosurfactant-producing microorganisms for improving pesticides' bioavailability and therefore the degradational efficacy of BPS.

Bioconversion of potato solid waste into antifungals and biopigments using Streptomyces spp.

Potato waste was processed and used as a sole substrate for simultaneously producing antifungals and biopigments using Streptomyces spp. Out of three different Streptomyces isolates, strain SO6 stood out due to its ability to produce antifungals against economically important fungal phytopathogens and intracellular biopigments using potato waste powders without additional nutrients. This strain also showed the potential to secrete a broad range of enzymes for fermentation of eight sugars that could be involved in potato waste bioconversion. The results of the fermentation assay indicated that Streptomyces sp. strain SO6 degrades potato wastes during submerged fermentation, diminishing total dry weight and increasing reducing sugars from 0.3 to 3.6 mg·mL-1 and total proteins from 70.6 to 187.7 µg·mL-1. The results showed that Streptomyces strain SO6 was able to convert the potato waste into 0.96 mg·g-1 of diffusible antifungals and 1.75 mg·g-1 of reddish-purple biopigments. On the contrary, an absence of pigment production was observed during the fermentation of the commercial medium used as reference. According to our results, replacement of commercial culture media with available low-cost agroindustrial wastes for producing bioactive chemicals is a real opportunity to enhance the Streptomyces pigment production and antibiotic sustainability with cost-competitiveness. To our knowledge, this is the first report on the simultaneous production of biopigments and diffusible antifungal antibiotics produced by Streptomyces spp. using potato solid waste as the sole nutrient source.

Pesticide-tolerant bacteria isolated from a biopurification system to remove commonly used pesticides to protect water resources.

In this study, we selected and characterized different pesticide-tolerant bacteria isolated from a biomixture of a biopurification system that had received continuous applications of a pesticides mixture. The amplicon analysis of biomixture reported that the phyla Proteobacteria, Firmicutes, Bacteroidetes and Actinobacteria were predominant. Six strains grew in the presence of chlorpyrifos and iprodione. Biochemical characterization showed that all isolates were positive for esterase, acid phosphatase, among others, and they were identified as Pseudomonas, Rhodococcus and Achromobacter based on molecular and proteomic analysis. Bacterial growth decreased as both pesticide concentrations increased from 10 to 100 mg L-1 in liquid culture. The Achromobacter sp. strain C1 showed the best chlorpyrifos removal rate of 0.072-0.147 d-1 a half-life of 4.7-9.7 d and a maximum metabolite concentration of 2.10 mg L-1 at 120 h. On the other hand, Pseudomonas sp. strain C9 showed the highest iprodione removal rate of 0.100-0.193 d-1 a half-life of 4-7 d and maximum metabolite concentration of 0.95 mg L-1 at 48 h. The Achromobacter and Pseudomonas strains showed a good potential as chlorpyrifos and iprodione-degrading bacteria.

Performance of a continuous stirred tank bioreactor employing an immobilized actinobacteria mixed culture for the removal of organophosphorus pesticides.

In this study, we evaluated polyurethane foam (PF), volcanic rock (VR), and a modified plastic cap (MPC) as supports for the immobilization of organophosphorus (OP) pesticide-degrading actinobacterial strains. The colonization and activity of four streptomycetes were favoured by PF, which was selected as the carrier to use in a continuous stirred tank bioreactor (CSTR) that can be operated at increasing inflows of a pesticide mixture that contains the insecticides chlorpyrifos (CP) and diazinon (DZ). Our results demonstrate that the CSTR can be operated at flow rates of 10 and 40 mL h-1 with greater than 85% removal of the pesticides in the short term. A significant decrease in the efficiency of CP removal was observed at the highest inflows into the reactor. The CP and DZ loading rates in the bioreactor ranged from 0.44 to 1.68 mg L-1 h-1 and from 0.50 to 2.17 mg L-1 h-1, respectively. Although the treated wastewater exhibited moderate toxicity for Raphanus sativus, a bioreactor inoculated with a mixed culture formed by Streptomyces spp. strains AC5, AC9, GA11 and ISP13 may provide an effective biotechnological strategy for the reduction of OP pesticide residues produced during agronomic and manufacturing practices and therefore prevent environmental pesticidal pollution.

Biodegradation inducers to enhance wheat straw pretreatment by Gloeophyllum trabeum to second-generation ethanol production.

The native state of lignocellulosic biomass is highly resistant to enzymatic hydrolysis and the fermentation process of biofuel production. Brown-rot fungi use an extracellular Fenton system to degrade lignocellulosic biomass in the initial stages of decay. In this work, the combined effects of Mn2+, Fe2+, and NO3- inducers were evaluated based on the activities of hydrolytic enzymes and Fe3+ reduction as well as the catechol-type compound production during wheat straw pretreatment by the brown-rot fungus Gloeophyllum trabeum. Weight loss and chemical changes were evaluated to establish the culture conditions for stimulating wheat straw degradation using a central composite design. The results showed that weight loss and the Fe3+-reducing activity were promoted at the highest concentrations of Fe2+. A positive effect on catechol compound production by the addition of Mn2+ and NO3- was observed. Cellulase activity was increased at the highest concentration of NO3-. The multiple optimizations of G. trabeum culture conditions in wheat straw resulted in 11.3% weight loss and 0.47 total crystallinity index at 0.24 M NO3-, 0.95 mM Fe2+, and 0.85 mM Mn2+ after 40 days. The wheat straw pretreatment by G. trabeum for 10 days increased glucose recovery. The results indicated that the wheat straw pretreatment using G. trabeum with biodegradation inducers could be a complementary step to physicochemical pretreatment of lignocellulosic biomass for production of second-generation ethanol.

Green synthesis of silver nanoparticles: effect of synthesis reaction parameters on antimicrobial activity.

In this work, the biosynthesis of silver nanoparticles by Galega officinalis extract using AgNO3 as a precursor was reported. The reaction parameters for the biosynthesis and efficiency in their antimicrobial control against Escherichia coli, Staphylococcus aureus and Pseudomonas syringae were determined. For biosynthesis, a central composite design combined with response surface methodology was used to optimize the process parameters (pH, AgNO3 and extract concentration), and the design was assessed through the size distribution, zeta potential and polydispersity index of the nanoparticles. The results demonstrated that at pH 11, 1.6 mM of AgNO3 and 15% vv-1 of G. officinalis extract were the optimal reaction parameters. Transmission electron microscope (TEM) images and X-ray diffraction (XRD) confirmed the formation of small spherical silver nanoparticles. Antimicrobial assays showed a high inhibitory effect against E. coli, S. aureus and P. syringae, and that effect was larger with silver nanoparticles of a smaller size (23 nm). This work demonstrates that G. officinalis extract is a feasible medium for the synthesis of silver nanoparticles and that the control of the reaction parameters can determine the nanoparticle characteristics and therefore their antimicrobial effectiveness.

Sequential white-rot and brown-rot fungal pretreatment of wheat straw as a promising alternative for complementary mild treatments.

White-rot and brown-rot fungi have complementary mechanisms to selectively degrade lignin and holocellullose, respectively. Thereby, a fungal co-culture of a white-rot and a brown-rot fungal could result in efficient strategy for a mild lignocellulosic biomass pretreatment. In this work, single, sequential and co-inoculation of the selective-lignin degrading white-rot fungus Ganoderma lobatum and the brown-rot fungus Gloeophyllum trabeum were evaluated as biological pretreatments of wheat straw to enhance enzymatic hydrolysis of cellulose. The single cultures of G. lobatum and G. trabeum exhibited preferential degradation of lignin and hemicellulose, respectively. The total crystallinity index decreased in samples pretreated with G. trabeum but not with G. lobatum. The pretreatment with single cultures of G. lobatum or G. trabeum increased glucose yields by 43.6% and 26.1% respectively compared to untreated straw. Although co-inoculation resulted in higher yields of glucose when compared with single cultures, only a slight synergistic effect between fungi was observed. Contrary, the sequential inoculation of G. lobatum incubated for 10 days followed by G. trabeum incubated for 10 days more showed a strong synergic effect on enzymatic hydrolysis. This sequential culture showed the highest glucose yield (191.5 mg g-1 wheat straw), 2.8-fold higher than untreated wheat straw, and 140-150% higher than the single-cultures of G. lobatum and G. trabeum, respectively.

Organophosphorus pesticide mixture removal from environmental matrices by a soil Streptomyces mixed culture.

The current study aimed to evaluate the removal of a pesticide mixture composed of the insecticides chlorpyrifos (CP) and diazinon (DZ) from liquid medium, soil and a biobed biomixture by a Streptomyces mixed culture. Liquid medium contaminated with 100 mg L-1 CP plus DZ was inoculated with the Streptomyces mixed culture. Results indicated that microorganisms increased their biomass and that the inoculum was viable. The inoculum was able to remove the pesticide mixture with a removal rate of 0.036 and 0.015 h-1 and a half-life of 19 and 46 h-1 for CP and DZ, respectively. The sterilized soil and biobed biomixture inoculated with the mixed culture showed that Streptomyces was able to colonize the substrates, exhibiting an increase in population determined by quantitative polymerase chain reaction (q-PCR), enzymatic activity dehydrogenase (DHA) and acid phosphatase (APP). In both the soil and biomixture, limited CP removal was observed (6-14%), while DZ exhibited a removal rate of 0.024 and 0.060 day-1 and a half-life of 29 and 11 days, respectively. Removal of the organophosphorus pesticide (OP) mixture composed of CP and DZ from different environmental matrices by Streptomyces spp. is reported here for the first time. The decontamination strategy using a Streptomyces mixed culture could represent a promising alternative to eliminate CP and DZ residues from liquids as well as to eliminate DZ from soil and biobed biomixtures.

Combined effect of enzyme inducers and nitrate on selective lignin degradation in wheat straw by Ganoderma lobatum.

Lignin is one of the main barriers to obtaining added-value products from cellulosic fraction of lignocellulosic biomass due to its random aromatic structure and strong association with cellulose and hemicellulose. Inorganic and organic compounds have been used as enzyme inducers to increase the ligninolytic potential of white-rot fungi, without considering their effect on the selectivity of degradation. In this study, the selective lignin degradation in wheat straw by Ganoderma lobatum was optimized using a central composite design to evaluate the combined effect of Fe2+ and Mn2+ as inducers of ligninolytic enzymes and NO3- as an additional nitrogen source. Selective lignin degradation was promoted to maximize lignin degradation and minimize weight losses. The optimal conditions were 0.18 M NO3-, 0.73 mM Fe2+, and 1 mM Mn2+, which resulted in 50.0% lignin degradation and 18.5% weight loss after 40 days of fungal treatment. A decrease in absorbance at 1505 and 900 cm-1 in fungal-treated samples was observed in the FTIR spectra, indicating lignin and cellulose degradation in fungal-treated wheat straw, respectively. The main ligninolytic enzymes detected during lignin degradation were manganese-dependent and manganese-independent peroxidases. Additionally, confocal laser scanning microscopy revealed that lignin degradation in wheat straw by G. lobatum resulted in higher cellulose accessibility. We concluded that the addition of enzyme inducers and NO3- promotes selective lignin degradation in wheat straw by G. lobatum.