Microbial decomposition of wood in streams: distribution of microflora and factors affecting [C]lignocellulose mineralization

The distribution and lignocellulolytic activity of the microbial community was determined on a large log of Douglas fir (Pseudotsuga menziesii) in a Pacific Northwest stream. Scanning electron microscopy, plate counts, and degradation of [C]lignocelluloses prepared from Douglas fir and incubated with samples of wood taken from the surface and within the log revealed that most of the microbial colonization and lignocellulose-degrading activity occurred on the surface. Labeled lignocellulose and surface wood samples were incubated in vitro with nutrient supplements to determine potential limiting factors of [C]lignocellulose degradation. Incubations carried out in a nitrogenless mineral salts and trace elements solution were no more favorable to degradation than those carried out in distilled water alone. Incubations supplemented with either (NH(4))(2)SO(4) or organic nitrogen sources showed large increases in the rates of mineralization over incubations with mineral salts and trace elements alone, with the greatest effect being observed from an addition of (NH(4))(2)SO(4). Subsequent incubations with (NH(4))(2)SO(4), KNO(3), and NH(4)NO(3) revealed that KNO(3) was the most favorable for lignin degradation, whereas all three supplements were equally favorable for cellulose degradation. Supplementation with glucose repressed both lignin and cellulose mineralization. The results reported in this study indicate that nitrogen limitation of wood decomposition may exist in streams of the Pacific Northwest. The radiotracer technique was shown to be a sensitive and useful tool for assessing relative patterns of lignocellulose decay and microbial activity in wood, along with the importance of thoroughly characterizing the experimental system before its general acceptance.     

Nitrogen dynamics in stream wood samples incubated with [C]lignocellulose and potassium [N]nitrate

Surface wood samples obtained from a Douglas fir log (Pseudotsuga menziesii) in a Pacific Northwest stream were incubated in vitro with [C]lignocellulose in a defined mineral salts medium supplemented with 10 mg of N liter of N-labeled NO(3) (50 atom% N). Evolution of CO(2), distribution and isotopic dilution of N, filtrate N concentrations, and the rates of denitrification, N(2) fixation, and respiration were measured at 6, 12, and 18 days of incubation. The organic N content of the lignocellulose-wood sample mixture had increased from 132 mug of N to a maximum of 231 mug of N per treatment after 6 days of incubation. Rates of [C]lignocellulose decomposition were greatest during the first 6 days and then began to decline over the remaining 12 days. Total CO(2) evolution was also highest at day 6 and declined steadily over the remaining duration of the incubation. Filtrate NH(4)-N increased from background levels to a final value of 57 mug of N per treatment. Filtrate NO(3) N completely disappeared by day 6, and organic N showed a slight decline between days 12 and 18. The majority of the N that could be recovered appeared in the particulate organic fraction by day 6 (41 mug of N), and the filtrate NH(4) N fraction contained 11 mug of N by day 18. The N enrichment values of the filtrate NH(4) and the inorganic N associated with the particulate fraction had increased to approximately 20 atom% N by 18 days of incubation, whereas the particulate organic fraction reached its highest enrichment by day 6. Measurements of N(2) fixation and denitrification indicated an insignificant gain or loss of N from the experimental system by these processes. The data show that woody debris in stream ecosystems might function as a rapid and efficient sink for exogenous N, resulting in stimulation of wood decomposition and subsequent activation of other N cycling processes.     

Thermophilic anaerobic biodegradation of [C]lignin, [C]cellulose, and [C]lignocellulose preparations

Thermophilic (55 degrees C) anaerobic enrichment cultures were incubated with [C-lignin]lignocellulose, [C-polysaccharide]lignocellulose, and kraft [C]lignin prepared from slash pine, Pinus elliottii, and C-labeled preparations of synthetic lignin and purified cellulose. Significant but low percentages (2 to 4%) of synthetic and natural pine lignin were recovered as labeled methane and carbon dioxide during 60-day incubations, whereas much greater percentages (13 to 23%) of kraft lignin were recovered as gaseous end products. Percentages of label recovered from lignin-labeled substrates as dissolved degradation products were approximately equal to percentages recovered as gaseous end products. High-pressure liquid chromatographic analyses of CuO oxidation products of sound and degraded pine lignin indicated that no substantial chemical modifications of the remaining lignin polymer, such as demethoxylation and dearomatization, occurred during biodegradation. The polysaccharide components of pine lignocellulose and purified cellulose were relatively rapidly mineralized to methane and carbon dioxide; 31 to 37% of the pine polysaccharides and 56 to 63% of the purified cellulose were recovered as labeled gaseous end products. An additional 10 to 20% of the polysaccharide substrates was recovered as dissolved degradation products. Overall, these results indicate that elevated temperatures can greatly enhance rates of anaerobic degradation of lignin and lignified substrates to methane and low-molecular-weight aromatic compounds.     

Simulated nitrogen deposition affects wood decomposition by cord-forming fungi

Anthropogenic nitrogen (N) deposition affects many natural processes, including forest litter decomposition. Saprotrophic fungi are the only organisms capable of completely decomposing lignocellulosic (woody) litter in temperate ecosystems, and therefore the responses of fungi to N deposition are critical in understanding the effects of global change on the forest carbon cycle. Plant litter decomposition under elevated N has been intensively studied, with varying results. The complexity of forest floor biota and variability in litter quality have obscured N-elevation effects on decomposers. Field experiments often utilize standardized substrates and N-levels, but few studies have controlled the decay organisms. Decomposition of beech (Fagus sylvatica) blocks inoculated with two cord-forming basidiomycete fungi, Hypholoma fasciculare and Phanerochaete velutina, was compared experimentally under realistic levels of simulated N deposition at Wytham Wood, Oxfordshire, UK. Mass loss was greater with P. velutina than with H. fasciculare, and with N treatment than in the control. Decomposition was accompanied by growth of the fungal mycelium and increasing N concentration in the remaining wood. We attribute the N effect on wood decay to the response of cord-forming wood decay fungi to N availability. Previous studies demonstrated the capacity of these fungi to scavenge and import N to decaying wood via a translocating network of mycelium. This study shows that small increases in N availability can increase wood decomposition by these organisms. Dead wood is an important carbon store and habitat. The responses of wood decomposers to anthropogenic N deposition should be considered in models of forest carbon dynamics.     

The influence of substrate quality and stream size on wood decomposition dynamics

Woody materials decayed more rapidly in a first order stream than in larger streams in eastern Quebec, Canada. The rate of annual mass loss (k) was highest (k=1.20) for alder wood chips in a first order stream and lowest (k=0.04) for black spruce wood chips in a ninth order stream. Decay rates for woody materials in a first order stream were inversely related to their initial lignin to nitrogen ratios. In larger streams, decay rates of woody materials were inversely related to their initial lignin concentrations. A number of quantifiable relationships were found to exist between the initial lignin and nitrogen contents of woody materials and the nitrogen dynamics of decaying wood.     

不同碳源和氮源对金针菇降解木质纤维素酶活性的影响

以3株栽培的金针菇Flammulina velutipes为材料,研究它们在玉米芯和棉子壳以及不同碳源、氮源培养条件下纤维素、半纤维素和木质素降解酶活性的规律。结果表明,不同金针菇菌株的羧甲基纤维素酶、木聚糖酶和漆酶活力显著不同（P<0.001）,同时,培养条件对羧甲基纤维素酶、木聚糖酶和漆酶的活力都有显著影响（P<0.001）。在简单碳源存在的条件下,金针菇的羧甲基纤维素酶和木聚糖酶活力远远低于复杂碳源培养基（P<0.05）。全营养培养基上生长的金针菇的羧甲基纤维素酶和木聚糖酶活力低于缺乏碳源和氮源的培养基（P<0.05）。漆酶活力在无简单氮源培养基上低于全培养基（P<0.05）和无葡萄糖培养基（P<0.05）,即复杂碳源和氮源培养基上的漆酶活力低于简单碳源和氮源培养基（P<0.05）。     

氮、磷养分有效性对森林凋落物分解的影响研究进展

通过对相关研究文献的综述结果表明,氮(N)和磷(P)是构成蛋白质和遗传物质的两种重要组成元素,限制森林生产力和其他生态系统过程,对凋落物分解产生深刻影响。大量的凋落物分解试验发现在土壤N有效性较低的温带和北方森林,凋落物分解速率常与底物初始N浓度、木质素/N比等有很好的相关关系,也受外源N输入的影响;而在土壤高度风化的热带亚热带森林生态系统中,P可能是比N更为重要的分解限制因子。然而控制试验表明,N、P添加对凋落物分解速率的影响并不一致,既有促进效应也有抑制效应。为了深入揭示N、P养分有效性对凋落物分解的调控机制,"底物的C、N化学计量学"假说、"微生物的N开采"假说以及养分平衡的理论都常被用于解释凋落物分解速率的变化。由于微生物分解者具有较为稳定的C、N、P等养分需求比例,在不同的养分供应的周围环境中会体现出不同的活性,某种最缺乏的养分可能就是分解的最重要限制因子。未来的凋落物分解研究,应延长实验时间、加强室内和野外不同条件下的N、P等养分添加控制试验,探讨驱动分解进程的微生物群落结构和酶活性的变化。     

Acetate production from hydrogen and [13C]carbon dioxide by the microflora of human feces

Fecal suspensions from humans were incubated with 13CO2 and H2. The suspensions were from subjects who harbored 10(8) and 10(10) methanogens per g (dry weight) of feces, respectively, and from a subject who did not harbor methanogens. Quantitative nuclear magnetic resonance spectroscopy showed that acetate labeled in both the methyl and carboxyl groups was formed by suspensions from the subject without methanogens and the subject with the lower concentrations of methanogens. The amounts of labeled acetate formed were in agreement with the amounts expected based on measurements of H2 utilization. No labeled acetate was formed by suspensions from the subject with the higher concentrations of methanogens, and essentially all of the H2 used was accounted for by CH4 production. Suspensions from the subject with lower concentrations of methanogens produced both methane and acetate from H2 and CO2. The results indicate that reduction of CO2 to acetate may be a major pathway for microbial production of acetate in the human colon except when very high concentrations of methanogens (ca. 10(10) per g [dry weight] of feces) are present. Double-labeled acetate was also formed from H2 and 13CO2 by fecal suspensions from nonmethanogenic and moderately methanogenic rats.     

Effects of nitrogen and phosphorus availability on the decomposition of aquatic plants

The responses of decomposition to nitrogen (N) and phosphorus (P) supply were investigated in three leaf species: Eichhornia crassipes, Vallisneria natans, and Potamogeton maackianus. Decomposition was fastest in E. crassipes (0.047–0.099 day⁻¹), intermediate in V. natans (about 0.030 day⁻¹), and slowest in P. maackianus (about 0.010 day⁻¹). Increase in P-availability increased the decomposition rate of E. crassipes by 68–87%, whereas the impact of N-availability alone was insignificant. Both N- and P-availability in waters had no significant impact on the decomposition rates of V. natans and P. maackianus (P > 0.05). The effects of P-availability on the N and P content levels of the three species were significant (P < 0.01), except for the impact on N content of V. natans (P = 0.526). In contrast, environment N-availability was insignificant. These results indicate that the responses of decomposition to nutrient availability depend on plant species and nutrient type. P-availability has stronger effects on litter nutrient dynamics than N-availability.     

Acetate production from hydrogen and [13C]carbon dioxide by the microflora of human feces.

Fecal suspensions from humans were incubated with 13CO2 and H2. The suspensions were from subjects who harbored 10(8) and 10(10) methanogens per g (dry weight) of feces, respectively, and from a subject who did not harbor methanogens. Quantitative nuclear magnetic resonance spectroscopy showed that acetate labeled in both the methyl and carboxyl groups was formed by suspensions from the subject without methanogens and the subject with the lower concentrations of methanogens. The amounts of labeled acetate formed were in agreement with the amounts expected based on measurements of H2 utilization. No labeled acetate was formed by suspensions from the subject with the higher concentrations of methanogens, and essentially all of the H2 used was accounted for by CH4 production. Suspensions from the subject with lower concentrations of methanogens produced both methane and acetate from H2 and CO2. The results indicate that reduction of CO2 to acetate may be a major pathway for microbial production of acetate in the human colon except when very high concentrations of methanogens (ca. 10(10) per g [dry weight] of feces) are present. Double-labeled acetate was also formed from H2 and 13CO2 by fecal suspensions from nonmethanogenic and moderately methanogenic rats.     

Effects of dietary nonspecific nitrogen on [14C]glutamate and [14C]proline incorporation into bone proteins in chicks

The incorporation of [14C]glutamic acid into EDTA-soluble and -insoluble calvaria protein in vitro and [14C]proline into EDTA-insoluble femur protein in vivo was determined in chicks fed inadequate and adequate levels of nonspecific nitrogen (glutamic acid). In each instance, the amount of amino acid incorporated into bone protein was reduced by the low level of nonspecific nitrogen. It was concluded that the high incidence of leg abnormalities observed in chicks fed purified diets containing adequate levels of indispensable amino acids but lacking in total nitrogen might be associated with an inability to form bone matrix protein.     

Impact of Nitrogen and Phosphorus on [14C]Lignocellulose Decomposition by Stream Wood Microflora

Nutritional and physical factors affecting the decomposition of [¹⁴C]lignocellulose prepared from Douglas fir (Pseudotsuga menziesii) were examined by incubating the labeled substrate with homogenized surface wood scrapings obtained from a Douglas fir log in a Pacific Northwest stream. Incubations were conducted in distilled water, in stream water collected from four different sources, or in a defined mineral salts solution with or without supplemental N (KNO₃). Decomposition rates of [¹⁴C]lignocellulose, as measured by ¹⁴CO₂ evolution, were greater in each of the four filter-sterilized sources of stream water than in distilled water alone. Decomposition experiments conducted in stream water media with the addition of defined mineral salts demonstrated that [¹⁴C]cellulose decomposition was stimulated 50% by the addition of either KNO₃ or KH₂PO₄/K₂HPO₄ and further enhanced (167%) by a combination of both. In contrast, [¹⁴C]lignin decomposition was stimulated (65%) only by the addition of both N and P. Decomposition of [¹⁴C]lignocellulose was greatest when supplemental KNO₃ was supplied in concentrations of at least 10.0 mg of N liter⁻¹ but not increased further by higher concentrations. The decomposition of [¹⁴C]lignocellulose increased as the incubation temperature was raised and NO₃⁻¹-N supplementation further increased these rates between three-and sevenfold over the range of temperatures examined (5 to 22°C). Accumulation of NH₄⁺ (2 to 4 mg of N liter⁻¹) was always observed in culture filtrates of incubations which had been supplemented with KNO₃, the quantity being independent of NO₃⁻ concentrations ≥ 10 mg of N liter⁻¹. The role of supplemental NO₃⁻ in the decomposition of [¹⁴C]lignocellulose is discussed in relation to wood decomposition and the low concentrations of N found in stream ecosystems of the Pacific Northwest.     

Anaerobic degradation of soluble fractions of [C-lignin]lignocellulose

[C-lignin]lignocellulose was solubilized by alkaline heat treatment and separated into different molecular size fractions for use as the sole source of carbon in anaerobic enrichment cultures. This study is aimed at determining the fate of low-molecular-weight, polyaromatic lignin derivatives during anaerobic degradation. Gel permeation chromatography was used to preparatively separate the original C-lignin substrate into three component molecular size fractions, each of which was then fed to separate enrichment cultures. Biodegradability was assessed by monitoring total carbon dioxide and methane production, evolution of labeled gases, loss of C-activity from solution, and changes in gel permeation chromatographic elution patterns. Results indicated that the smaller the size of the molecular weight fraction, the more extensive the degradation to gaseous end products. In addition, up to 30% of the entire soluble lignin-derived carbon was anaerobically mineralized to carbon dioxide and methane.     

Effects of stream order and season on mineralization of [14C]-phenol in streams

Mineralization of trace levels of [¹⁴C]-phenol by heterotrophic microorganisms was quantified at 4 sites along a river continuum in southwestern Virginia. Significant phenol mineralization rates were detected in surface sediment and seston samples at all sites from August 1985 through May 1986. Phenol degradation was strongly affected by season (ANOVA; P < 0.0001). From a baseline rate in August (range: 1.19 × 10^-5 to 897 × 10^-4 mg phenol mineralized mg AFDW^-1 h^-1) phenol mineralization rose to a yearly maximum in October (range: 1.21 × 10^-4 to 1.16 × 10^-3 mg phenol mineralized mg AFDW^-1 h^-1) despite decreasing stream temperatures. This autumnal peak in phenol degradation was attributed to the pulsed input of allochthonous detritus, especially leaf litter, which contains substantial quantities of phenols and related compounds. Although phenol mineralization was significant in these streams, phenols were metabolized at much slower rates than more labile compounds present in the dissolved organic matter (DOM) pool. Estimates of turnover rates for three major components of DOM revealed that glucose and glutamate turnover rates (0.064–0.140 h^-1 mg sediment AFDW^-1 and 0.140–0.610 h^-1 mg sediment AFDW^-1, respectively) were, respectively, 2.2–4.7 × and 9.6–16.9 × greater than phenol turnover rates (0.015–0.064 h^-1 mg sediment AFDW^-1). Although the relatively low rates of utilization of refractory phenolic materials suggest that these compounds may accumulate and become more prevalent components of the DOM pool, phenol concentrations at the 4 study sites remained below detectable levels (i.e., < 1 g 1^-1) throughout the study. Consequently, it seems that although phenolic materials are metabolized more slowly than labile DOM, phenols are degraded at rates which preclude accumulation in the water column.     

Solubilisation and mineralisation of [14C]lignocellulose from wheat straw by Streptomyces cyaneus CECT 3335 during growth in solid-state fermentation

Nine Streptomyces strains were screened for their ability to solubilise and mineralise ¹⁴C-labelled lignin during growth in solid-state fermentation. Streptomyces viridosporus was confirmed as an active lignin-degrading organism along with a new isolate, Streptomyces sp. UAH 15, further classified as Streptomyces cyaneus CECT 3335. This organism was able to solubilise and mineralise the [¹⁴C]lignin fraction of lignocellulose (44.96 ± 1.77% and 3.41 ± 0.48% respectively) after 21 days of incubation. Cell-free filtrates from Streptomyces sp. grown in solid-state fermentation were capable of solubilising up to 20% of the [¹⁴C]lignin after 2 days incubation, with most of the product detected in the acid-soluble rather than in the water-soluble fraction. Identification of the extracellular enzymes produced during growth of S. cyaneus CECT 3335 revealed that extracellular peroxidase and phenol oxidase activities were present, with the activity of phenol oxidase being 100 times greater than peroxidase activity. The activity of these two enzymes was found to correlate with both solubilisation and mineralisation rates. This is the first report of phenol oxidase activity produced by a Streptomyces strain during growth in solid-state fermentation. A role for the enzyme in the solubilisation and mineralisation of lignocellulose by S. cyaneus is suggested. Received: 12 May 1997 / Accepted: 19 May 1997     

Bioconversion of alpha-[14C]zearalenol and beta-[14C]zearalenol into [14C]zearalenone by Fusarium roseum 'Gibbosum'

Cultures of Fusarium roseium 'Gibbosum' on rice were treated with [14C]zearalenone, alpha[14C]zearalenol, or beta-[14C]zearalenol to determine whether a precursor-product relationship exists among these closely related fungal metabolites. Culture extracts were purified by silica gel column chromatography and fractionated by high-pressure liquid chromatography, and the level of radioactivity was determined. Within 7 days, the beta-[14C]zearalenol was converted to zearalenone, and no residual beta-[14C]zearalenol was detectable. Most of the alpha-[14C]zearalenol added was also converted into zearalenone with 14 days. In cultures treated with [14C]zearalenone, no radioactivity was noted in any other components.     

Bioconversion of alpha-[14C]zearalenol and beta-[14C]zearalenol into [14C]zearalenone by Fusarium roseum 'Gibbosum'.

Cultures of Fusarium roseium 'Gibbosum' on rice were treated with [14C]zearalenone, alpha[14C]zearalenol, or beta-[14C]zearalenol to determine whether a precursor-product relationship exists among these closely related fungal metabolites. Culture extracts were purified by silica gel column chromatography and fractionated by high-pressure liquid chromatography, and the level of radioactivity was determined. Within 7 days, the beta-[14C]zearalenol was converted to zearalenone, and no residual beta-[14C]zearalenol was detectable. Most of the alpha-[14C]zearalenol added was also converted into zearalenone with 14 days. In cultures treated with [14C]zearalenone, no radioactivity was noted in any other components.     

Anaerobic biodegradation of the lignin and polysaccharide components of lignocellulose and synthetic lignin by sediment microflora

[This corrects the article on p. 998 in vol. 47.].     

Formation of [13C]- and [14C]zearalenone by Fusarium graminearum DSM 4529

Summary To raise the yields for the production of ¹⁴C-labelled zearalenone in Fusarium cultures the influence of growth conditions and known effectors or precursors of toxin biosynthesis was studied. Benzoic acid and 2,4-dihydroxybenzoic acid used as precursors decreased toxin formation; in the presence of different pesticides such as 2,4-dichlorophenoxyacetic acid, however, toxin production increased up to 140%. The known pathway of zearalenone biosynthesis could be confirmed from the relative extents of ¹³C-incorporation into the zearalenone molecule by incubating Fusarium graminearum DSM 4529 with d-(+)-[1-¹³C]glucose as carbon source. When grown in the presence of d-[U-¹⁴C]glucose or [2-¹⁴C]malonic acid the strain produced [¹⁴C]zearalenone with specific activities of 0.07 and 0.09 Ci/mg, the ¹⁴C-incorporation rates being 0.34% and 0.48%, respectively.