Biotransformation and detoxification of chloroacetanilide herbicides by Trichoderma spp. with plant growth-promoting activities.

Due to the increasing use of chlorinated organic compounds, environmental pollution is a key issue in agricultural and industrial areas. In this study, biodegradation of chloroacetanilide herbicides, such as alachlor and metolachlor, by eight fungal strains of Trichoderma spp. originating from different microorganism collections was investigated. The tested fungi converted 80-99% of alachlor and 40-79% of metolachlor after 7 days of incubation. Biotransformation of herbicides was performed mainly by dechlorination and hydroxylation reactions. Eight alachlor metabolites and four byproducts of metolachlor conversion were detected in Trichoderma cultures, including two metolachlor intermediates for the first time identified in fungi. Moreover, in the cultures of six Trichoderma strains supplemented with chloroacetanilides, a decrease in toxicity was observed toward tested Artemia franciscana crustaceans. Simultaneously, 7 days after the application of the spores of T. koningii IM 0956, T. citrinoviride IM 6325, T. harzianum KKP 534, T. viride KKP 792 and T. virens DSM 1963 the length of roots and shoots of rapeseed seedlings treated with alachlor or metolachlor significantly increased. All tested strains exhibited plant growth-promoting traits, such as siderophore production, 1-aminocyclopropane-1-carboxylate deaminase (ACCD) activity, and phosphate solubilization, even in the presence of chloroacetanilide herbicides.

Assessment of oxidative stress and phospholipids alterations in chloroacetanilides-degrading Trichoderma spp.

To investigate the induction of oxidative stress and antioxidant response in the chloroacetanilides-degrading Trichoderma spp. under alachlor and metolachlor exposure, a comparative analysis using popular biomarkers was employed. An increased intracellular level of reactive oxygen species (ROS; especially superoxide anion [O2-]) as well as products of lipid and protein oxidation after 24 h incubation with the herbicides confirmed chloroacetanilide-induced oxidative stress in tested Trichoderma strains. However, the considerable decline in the ROS levels and the carbonyl group content (biomarkers of protein peroxidation) in a time-dependent manner and changes in the antioxidant enzyme activities indicated an active response against chloroacetanilide-induced oxidative stress and the mechanism of tolerance in tested fungi. Moreover, the tested herbicides clearly modified the phospholipids (PLs) content in Trichoderma spp. in the stationary phase of growth, which was manifested through the difference in phosphatidic acid (PA), phosphatidylethanolamine (PE) and phosphatidylcholines (PC) levels. Despite enhanced lipid peroxidation and changes in PLs in most tested fungi, only a slight modification in membrane integrity of Trichoderma spp. under chloroacetanilides exposure was noted. The obtained results suggest that the alterations in the antioxidant system and the PLs profile of Trichoderma spp. might be useful biomarkers of chloroacetanilide-induced oxidative stress.

Trichoderma harzianum diminished oxidative stress caused by 2,4- dichlorophenoxyacetic acid (2,4-D) in wheat, with insights from lipidomics.

2,4-dichlorophenoxyacetic acid (2,4-D) is among the most commonly used herbicides applied for weed control during wheat cultivation. However, its application could affect wheat growth. The present study investigates the effect of the ascomycetous fungus Trichoderma harzianum on lipid peroxidation, phospholipids, signaling lipids and phospholipase D in the seedlings of wheat (Triticum aestivum L.) treated with 2,4-D (2.5 mg L-1). In the group of 4-day-old seedlings exposed to the herbicide, increased lipid peroxidation and inhibition of growth were observed in shoots and roots. Moreover, elevated levels of oxylipins were noted. Among them, the amount of 13-HOTrE oxygenated from linolenic acid (18:3) increased the most significantly. Concurrently, in the seedlings inoculated with T. harzianum, growth was stimulated when the level of phosphatidylcholine (PC) increased. Moreover, in wheat seedlings treated with 2,4-D and T. harzianum, the level of lipid peroxidation was similar to that in the control and there was no increase observed in oxylipins and phospholipase D activity. T. harzianum might have partly alleviated the toxic effect of 2,4-D on wheat seedlings.

Potential of Trichoderma koningii to eliminate alachlor in the presence of copper ions.

The filamentous fungus Trichoderma koningii is capable of fast and effective eliminate alachlor (90% after 72 h when added separately and 80-60% in the presence of 1-5 mM of copper). After 168 h over 99% elimination of alachlor resulted in detoxification and was connected with the mitigation of reactive oxygen species (ROS) production. Using MS/MS techniques, seven dechlorinated and hydroxylated metabolites were identified. Cytochrome P450 and laccase participate in biotransformation of the herbicide by this non-ligninolytic fungus. Laccase activity is stimulated both by copper and the mixture of copper and alachlor, which seems to be important for combined pollutants. T. koningii is characterized by high tolerance to copper (up to 7.5 mM). The metal content in mycelia reached 0.9-7.76 mg in 1 g of dry biomass. Our results suggest that T. koningii strain seems to be a promising tool for bioremediation of agricultural areas co-contaminated with copper-based fungicides and chloroacetanilide herbicides.

2,4-dichlorophenoxyacetic acid-induced oxidative stress: Metabolome and membrane modifications in Umbelopsis isabellina, a herbicide degrader.

The study reports the response to herbicide of the 2,4-dichlorophenoxyacetic acid (2,4-D)-degrading fungal strain Umbelopsis isabellina. A comparative analysis covered 41 free amino acids as well as 140 lipid species of fatty acids, phospholipids, acylglycerols, sphingolipids, and sterols. 2,4-D presence led to a decrease in fungal catalase activity, associated with a higher amount of thiobarbituric acid-reactive substances (TBARS). Damage to cells treated with the herbicide resulted in increased membrane permeability and decreased membrane fluidity. Detailed lipidomic profiling showed changes in the fatty acids composition such as an increase in the level of linoleic acid (C18:2). Moreover, an increase in the phosphatidylethanolamine/phosphatidylcholine ratio was observed. Analysis of fungal lipid profiles revealed that the presence of 2,4-D was accompanied by the accumulation of triacylglycerols, a decrease in ergosterol content, and a considerable rise in the level of sphingolipid ceramides. In the exponential phase of growth, increased levels of leucine, glycine, serine, asparagine, and hydroxyproline were found. The results obtained in our study confirmed that in the cultures of U. isabellina oxidative stress was caused by 2,4-D. The herbicide itself forced changes not only to membrane lipids but also to neutral lipids and amino acids, as the difference of tested compounds profiles between 2,4-D-containing and control samples was consequently lower as the pesticide degradation progressed. The presented findings may have a significant impact on the basic understanding of 2,4-D biodegradation and may be applied for process optimization on metabolomic and lipidomic levels.

Elimination and detoxification of 2,4-D by Umbelopsis isabellina with the involvement of cytochrome P450.

The chemical 2,4-dichlorophenoxyacetic acid (2,4-D) is used in agriculture as a herbicide. Its intensive use has an adverse effect on the environment. This study involved examining the degradation of 2,4-D compound by the filamentous fungus Umbelopsis isabellina. After 5 days of incubation, 98% of the herbicide (added at 25 mg L-1) was found to be removed. The elimination of 2,4-D by U. isabellina was connected with the formation of 2,4-dichlorophenol (2,4-DCP), which resulted in a 60% decrease in the sample toxicity toward Artemia franciscana larvae. The metabolism of 2,4-D was inhibited by the addition of metyrapone, a known cytochrome P450 inhibitor. It provides evidence that cytochrome P450 system is involved in 2,4-D metabolism in U. isabellina.