Biodegradation of 2,4,6-trinitrotoluene byPhanerochaete chrysosporium: Identification of initial degradation products and the discovery of a TNT metabolite that inhibits lignin peroxidases

Biodegradation of 2,4,6-trinitrotoluene (TNT) by the wood-rotting BasidiomycetePhanerochaete chrysosporium was studied in a fixed-film silicone membrane bioreactor and in agitated pellected cultures. The initial intermediate products of TNT biodegradation were shown to be 2-amino-4,6-dinitrotoluene (2amDNT) and 4-amino-2,6-dinitrotoluene (4amDNT). These intermediates were also degraded byP. chrysosporium. However, their rates of degradation were slow and appeared to represent rate-limiting steps in TNT degradation. The fact that 2amDNT and 4amDNT were further degraded is of importance. In most other microbial systems these compounds are typically not further degraded or are dimerized to even more persistent azo and azoxydimers. Similar to previous studies performed in stationary cultures, it was shown that substantial amounts of [¹⁴C]-TNT were degrade to [¹⁴C]-carbon dioxide in agitated pelleted cultures. Lignin peroxidase activity (assayed by veratryl alcohol oxidation) virtually disappeared upon addition of TNT to ligninolytic cultures ofP. chrysosporium. However, TNT, 2amDNT, and 4amDNT did not inhibit lignin peroxidase activity, nor were they substrates for this enzyme. Subsequent studies revealed that 4-hydroxylamino-2,6-dinitrotoluene, an intermediate in TNT reduction, was a potent lignin peroxidase inhibitor. Further studies revealed that this compound was also a substrate for lignin peroxidase H8.     

In vivo degradation of oxidized, regenerated cellulose

Oxidized, regenerated cellulose (ORC) was surgically implanted on the uterine horns of rabbits, and its biodegradation was studied in vivo. Samples of peritoneal lavages, serum, and urine were collected during the degradation process and analyzed for carbohydrate components utilizing high-performance liquid chromatography with pulsed amperometric detection (h.p.l.c.-p.a.d.). Degradation was rapid, and oligomeric products were evident primarily in the peritoneal fluid from the implantation site, with no apparent accumulation in either the serum or the urine. The size distribution and the amount of the oligomeric products decreased after day one, and by day four peritoneal lavages were essentially free of oligomers. The structure of the products formed was consistent with the lability of the polymer in solution, and the kinetics of degradation paralleled the results of the previously reported in vitro studies. Rabbit peritoneal macrophages, when incubated with ORC in vitro were observed to readily ingest and hydrolyze the polymeric material. A mechanism of degradation consisting of chemical depolymerization, followed by enzymatic hydrolysis mediated by glycosidases endogenous to peritoneal macrophages, is proposed.     

Disruption of a Global Regulatory Gene to Enhance Central Carbon Flux into Phenylalanine Biosynthesis in Escherichia coli

Genetic engineering of microbes for commercial metabolite production traditionally has sought to alter the levels and/or intrinsic activities of key enzymes in relevant biosynthetic pathway(s). Microorganisms exploit similar strategies for flux control, but also coordinate flux through sets of related pathways by using global regulatory circuits. We have engineered a global regulatory system of Escherichia coli, Csr (carbon storage regulator), to increase precursor for aromatic amino acid biosynthesis. Disruption of csrA increases gluconeogenesis, decreases glycolysis, and thus elevates phosphoenolpyruvate, a limiting precursor of aromatics. A strain in which the aromatic (shikimate) pathway had been optimized produced twofold more phenylalanine when csrA was disrupted. Overexpression of tktA (transketolase) to increase the other precursor, erythrose-4-phosphate, yielded ∼1.4-fold enhancement, while both changes were additive. These effects of csrA were not mediated by increasing the regulatory enzymes of phenylalanine biosynthesis. This study introduces the concept of “global metabolic engineering” for second-generation strain improvement. Received: 25 October 2000 / Accepted: 8 December 2000     

Biodegradation of phenanthrene by Phanerochaete chrysosporium: on the role of lignin peroxidase

Lignin peroxidase H8 from the wood rotting basidiomycete Phanerochaete chrysosporium is able to catalyse oxidation of 9-phenanthrol, forming phenanthrene-9, 10-quinone. This is of interest because 9-phenanthrol is an intermediate in the major pathway for phenanthrene degradation that occurs in this fungus under non-ligninolytic conditions whereas the product, phenanthrene-9, 10-quinone, is an intermediate in the pathway that occurs under ligninolytic conditions. It thus appears reasonable to suggest that, at the onset of idiophase (when cultures become ligninolytic), lignin peroxidases may function to link these two pathways.     

Crop residue harvest impacts wind erodibility and simulated soil loss in the Central Great Plains

Crop residue removal can affect the susceptibility to soil wind erosion in climates such as those of the Central Great Plains, United States. Six on‐farm trials were conducted in Kansas from 2011 to 2013 to determine the effects of winter wheat (Triticum aestivum L.), corn (Zea mays L.), and grain sorghum (Sorghum bicolor (L.) Moench), residue removal at 0, 25, 50, 75, and 100% of initial height on soil wind erosion parameters. Those parameters include soil surface random roughness (RR), and wind erodible fraction (EF; aggregates <0.84 mm), geometric mean diameter (GMD) and geometric standard deviation (GSD), stability of dry aggregates (DAS). Complete (100%) residue removal decreased the surface RR, increased EF, and decreased GMD. Overwinter EF values increased for five of six sites from fall 2011 to spring of 2012, particularly for the uppermost removal height (≥75%). Measured EF, GMD, GSD, DAS, and RR were also input into the Single‐event Wind Erosion Evaluation Program (SWEEP) to determine the effect of these parameters on simulated soil loss. The SWEEP simulated the wind velocity needed to initiate wind erosion as well as soil loss under each residue removal height at a wind velocity of 13 m s^?1 for three hours. Threshold wind velocity required to initiate wind erosion generally decreased with increasing crop residue removal height, particularly for >75% removal. Total estimated soil loss over the three‐hour event ranged from ≈2 to 25 Mg ha^?1, depending on EF, GMD, GSD, RR, and percent crop residue cover. Removing 75% residue increased simulated wind erosion at three of six sites while removing 50% appears sustainable at all six study sites. Findings reinforce the need for site‐by‐site consideration of the potential amount of crop residue that may be harvested while mitigating wind erosion. Study results indicate the value of maintaining residue at >75% of original height.     

Mast cells in the focus of an acute infectious inflammation]

On the model of E. coli-induced acute infectious peritonitis in rats it is established that the mast cell reaction and histamine level increase in exudate and inflamed mesentery tissue are biphase and are observed predominantly following the inflammatory agent action, in the period corresponding to the immediate phase of peritoneal cavity vessel permeability increases. The preliminary elimination of mast cells significantly inhibits a rise in the vascular permeability in the immediate phase and slightly affects the delayed phase, thus prolonging exudation. At the same time the dynamics of free histamine indicates its direct involvement in mediation and/or modulation as well as in subsequent inflammatory events. The common rules of mast cell involvement and vascular permeability increase in infectious and aseptic inflammation have been shown.     

Biodegradation of oxidized regenerated cellulose

The in vitro solubilization and degradation of regenerated cellulose was studied under conditions which approximate those found in vivo, when the material is used as an adhesion barrier to assist normal wound repair. Factors affecting solubilization which were examined included the effects of serum or plasma, and the presence of hydrolytic enzymes. Products of the solubilization and degradation processes were examined by high performance liquid chromatography coupled with pulsed amperometric detection. The oxidized polymer readily undergoes chain shortening to give oligomers which, in the presence of plasma or serum, are further hydrolyzed to smaller fragments, including glucuronic acid and glucose. Proposed mechanisms of degradation are discussed.