Microbial pretreatment of cotton stalks by submerged cultivation of Phanerochaete chrysosporium

This study used the fungus, Phanerochaete chrysosporium, to pretreat cotton stalks with two methods, shallow stationary and agitated cultivation, at three supplemental salt concentrations. Pretreatment efficiencies were compared by evaluating lignin degradation, solid recovery and carbohydrate availability over a 14-day period. Shallow stationary cultivation with no salts gave 20.7% lignin degradation along with 76.3% solid recovery and 29.0% carbohydrate availability. The highest lignin degradation of 33.9% at a corresponding solid recovery and carbohydrate availability of 67.8% and 18.4%, respectively, was obtained through agitated cultivation with Modified NREL salts. Cultivation beyond 10 days did not significantly increase lignin degradation during 14 days of pretreatment. Manganese addition during shallow stationary and agitated cultivation resulted in higher solid recoveries of over 80% but lower lignin degradation. Although agitated cultivation resulted in better delignification, results indicate that pretreatment under submerged shallow stationary conditions provides a better balance between lignin degradation and carbohydrate availability.     

巴西蘑菇对木质纤维素的降解与转化*

巴西蘑菇能够降解棉籽壳和麦草两种培养基中木质纤维素复合体中的全部组分,属于白腐真菌;巴西蘑菇降解的有机物质的绝大部分被菌体的呼吸过程消耗掉,其绝对生物学效率较低,仅为4.41%~5.25%;在栽培前期木质素的降解速率大于纤维素和半纤维素,这对纤维素和半纤维素的降解十分有利;非木质纤维素组分主要在菌丝生长阶段被利用,而木质纤维素是子实体生长发育阶段的主要碳源;就整个栽培过程而言,巴西蘑菇生长发育所需要的82.39%~84.50%的碳源来自木质纤维素。     

一株地膜降解真菌的筛选及其降解性能分析

【目的】分离并鉴定具有聚乙烯材料降解能力的微生物菌株,探究其降解农用地膜的效能,为地膜的微生物降解途径提供支撑。【方法】以线性低密度聚乙烯粉末为唯一碳源的培养物中分离出1株具有降解聚乙烯材料能力的真菌,利用分子生物学方法结合菌株的培养性状对该菌株进行鉴定,通过观察聚乙烯粉末降解情况和测定地膜失重率,结合红外扫描、高分辨场发射扫描电子显微镜分析该菌株对农用地膜的降解效果。【结果】筛选获得1株具有农用地膜降解效果的真菌菌株PT1,经鉴定为桔青霉(Penicillium citrinum),桔青霉PT1菌株能以重均分子量(Mw)2000和400000的聚乙烯粉末作为唯一碳源生长,经红外扫描、电镜观察发现桔青霉PT1可侵蚀传统聚乙烯地膜。桔青霉PT1能快速利用聚酯类生物降解地膜生长,35 d地膜失重率达50%左右。【结论】本文筛选到具有地膜降解特性的桔青霉PT1真菌,丰富了降解聚乙烯材料的微生物类群,同时也为废弃农用地膜的处理提供了环保的处理途径。     

贝叶多孔菌营养生理生化特性的研究

本文对贝叶多孔菌(Polyporus frondosus)在柞树木屑——麦麸基物上生长期间基物的降解特性和培养物中胞外纤维素酶、半纤维素酶和淀粉酶在培养过程中的活性变化规律及其它一些生物化学性质进行了研究。认为贝叶多孔菌是白腐型木腐真菌。在培养初期主要利用基物中可溶性糖类为碳源,在子实体发育期,碳源基本由降解基物中木质纤维素所提供。     

Evariquinone,isoemericellin, and stromemycin from a sponge derived strain of the fungus Emericella variecolor

From a strain of the fungus Emericella variecolor derived from the marine sponge Haliclona valliculata, two new natural products, evariquinone and isoemericellin, were isolated after HPLC-UV, -MS, and -NMR studies of the extract and their structures were elucidated by mass spectrometry and NMR experiments. Evariquinone showed antiproliferative activity towards KB and NCI-H460 cells at a concentration of 3.16 microg/ml. Furthermore, the fungus was found to produce the known metabolites stromemycin, shamixanthone, and 7-hydroxyemodin. Chemical degradation, NMR decoupling experiments, and spin-system simulation provided evidence for the double bonds in stromemycin to be all E-configured. ROESY experiments established the monosaccharide moiety to be glucose.     

Influence of PAHs on ligninolytic enzymes of the fungus <Emphasis Type="Italic">Pleurotus ostreatus</Emphasis> D1

Polycyclic aromatic hydrocarbons (PAHs), their derivatives, and their degradation products were assayed for the ability to enhance activities of ligninolytic enzymes (laccase and versatile peroxidase) of the fungus Pleurotus ostreatus D1. The activities of both laccase and versatile peroxidase were induced by the PAHs, their derivatives, and their degradation products. Laccase was produced mostly in the first 7–10 days, whereas the production of versatile peroxidase began after 5–7 days of cultivation. Non-denaturing PAGE showed the presence of additional forms of laccase and versatile peroxidase in the presence of the xenobiotics in the cultivation medium. The difference in the production time for these enzymes may reflect that laccases are involved in the first stages of PAHs degradation and that versatile peroxidase can be necessary for oxidation of some degradation products. This is the first report on versatile peroxidase induction by PAHs and their derivatives.     

Interaction of the rumen fungus Orpinomyces joyonii with Megasphaera elsdenii and Eubacterium limosum

The degradation and fermentation of microcrystalline cellulose were studied in monoculture of the polycentric anaerobic fungus Orpinomyces joyonii and in co-cultures with the rumen bacteria Megasphaera elsdenii and Eubacterium limosum. More than 25% of cellulose hydrolysis products (glucose and cellodextrins) were released by the fungus into the medium after 8 d of cultivation. These products were metabolized by bacteria in mixed cultures. In co-culture with the fungus M. elsdenii and E. limosum . increased the extent of microcrystalline cellulose degradation by 10·12% and 7·96%, respectively. Biomass yield in co-cultures was increased by 89·9% and 59·4% for M. elsdenii and E. limosum . Y_cellulose for fungus alone was 52·29 g dry matter mol^-1 glucose. These values were 64·93 and 55·92 g mol^-1 glucose unit in co-culture with M. elsdenii and E. limosum , respectively.     

EEG source localization: comparative study of classical and neural network methods.

We address the problem of estimating biopotential sources within the brain, based on EEG signals observed on the scalp. This problem, known as the inverse problem of electrophysiology, has no closed-form solution, and requires iterative techniques such as the Levenberg-Marquardt (LM) algorithm. Considering the nonlinear nature of the inverse problem, and the low signal to noise ratio inherent in EEG signals, a backpropagation neural network (BPN) has been recently proposed as a solution. The technique has not been properly compared with classical techniques such as the LM method, or with more recent neural network techniques such as the Radial Basis Function (RBF) network. In this paper, we provide improved strategies based on BPN and consider RBF networks in solving the inverse problem. We compare the performances of BPN, RBF and a hybrid technique with that of the classical LM method.     

Influence of cultivation conditions on pyrene degradation by the fungus Pleurotus Ostreatus D1

For the first time the dependence of completeness of pyrene degradation by the white-rot fungus Pleurotus ostreatus D1 on cultivation conditions was found. In Kirk’s medium about 65.6 ± 0.9% of the initial pyrene was metabolized after 3 weeks, with pyrene-4,5-dihydrodiol accumulating. This process was accompanied by laccase production only. In basidiomycetes rich medium, P. ostreatus D1 metabolized up to 89.8 ± 2.3% of pyrene within 3 weeks without pyrene-4,5-dihydrodiol accumulation throughout the time of cultivation. Phenanthrene and phthalic acid were identified as the metabolites produced from pyrene degradation under these conditions. Accumulation of phenanthrene with its subsequent disappearance was observed. One more metabolite probably was the product of phenanthrene degradation. Pyrene metabolism in basidiomycetes rich medium was accompanied first by laccase and tyrosinase production and later by versatile peroxidase production. The cell-associated activities of laccase, tyrosinase, and versatile peroxidase were found. The data obtained indicate that both enzymes (laccase and versatile peroxidase) are necessary for complete degradation of pyrene. Furthermore, both cell-associated and extracellular laccases can catalyse the first stages of pyrene degradation, and versatile peroxidase can be necessary for oxidation of the resulting metabolites.     

Studies on xylanase fromBasidiomycetes

Formation of extracellular xylanase was studied in 10 strains of wood-destroying fungi belonging to Basidiomycetes during their submerged cultivation with willow sawdust. The highest enzyme activity was found in the fungus Trametes hirsuta (Wulf.) Pilát. The effect of sources of carbon and nitrogen, cultivation time and initial pH of the cultivation solution on the formation of xylanase by the fungus Trametes hirsuta was investigated. The highest production of the enzyme was reached during cultivation in the presence of willow sawdust, asparagine and at the initial pH of 5.0. The presence of xylanase, cellulase, mannanase and amylase as well as of beta-xylosidase, beta-glucosidase, beta-mannosidase and beta-galactosidase was demonstrated in the enzyme preparation obtained after a 10-day submerged cultivation of Trametes hirsuta under optimal conditions.     

Effect of carbon source on the production of oxalic acid and hydrogen peroxide by brown-rot fungus Poria placenta

Oxalic acid and hydrogen peroxide have been suggested to be essential in the degradation of wood carbohydrates by brown-rot fungi. The production of oxalic acid, hydrogen peroxide and endo-β-1,4-glucanase activity by the brown-rot fungus Poria placenta was studied on crystalline cellulose, amorphous cellulose and glucose media. Oxalic acid and hydrogen peroxide by P. placenta were clearly produced on culture media containing either crystalline or amorphous cellulose. Oxalic acid and hydrogen peroxide were formed simultaneously and highest amounts of oxalic acid (1.0 g l⁻¹) and hydrogen peroxide (39.5 μM) were obtained on amorphous cellulose after 3 weeks cultivation. On glucose medium the amounts were low. The endoglucanase activity was observed to increase during the cultivation and was most pronounced on glucose medium and thus indicated the constitutive characteristics of the brown-rot cellulases.     

Specificity in the symbiotic association between fungus-growing ants and protective Pseudonocardia bacteria

Fungus-growing ants (tribe Attini) engage in a mutualism with a fungus that serves as the ants' primary food source, but successful fungus cultivation is threatened by microfungal parasites (genus Escovopsis). Actinobacteria (genus Pseudonocardia) associate with most of the phylogenetic diversity of fungus-growing ants; are typically maintained on the cuticle of workers; and infection experiments, bioassay challenges and chemical analyses support a role of Pseudonocardia in defence against Escovopsis through antibiotic production. Here we generate a two-gene phylogeny for Pseudonocardia associated with 124 fungus-growing ant colonies, evaluate patterns of ant-Pseudonocardia specificity and test Pseudonocardia antibiotic activity towards Escovopsis. We show that Pseudonocardia associated with fungus-growing ants are not monophyletic: the ants have acquired free-living strains over the evolutionary history of the association. Nevertheless, our analysis reveals a significant pattern of specificity between clades of Pseudonocardia and groups of related fungus-growing ants. Furthermore, antibiotic assays suggest that despite Escovopsis being generally susceptible to inhibition by diverse Actinobacteria, the ant-derived Pseudonocardia inhibit Escovopsis more strongly than they inhibit other fungi, and are better at inhibiting this pathogen than most environmental Pseudonocardia strains tested. Our findings support a model that many fungus-growing ants maintain specialized Pseudonocardia symbionts that help with garden defence.     

Metagenomic and metaproteomic insights into bacterial communities in leaf-cutter ant fungus gardens

Herbivores gain access to nutrients stored in plant biomass largely by harnessing the metabolic activities of microbes. Leaf-cutter ants of the genus Atta are a hallmark example; these dominant neotropical herbivores cultivate symbiotic fungus gardens on large quantities of fresh plant forage. As the external digestive system of the ants, fungus gardens facilitate the production and sustenance of millions of workers. Using metagenomic and metaproteomic techniques, we characterize the bacterial diversity and physiological potential of fungus gardens from two species of Atta. Our analysis of over 1.2 Gbp of community metagenomic sequence and three 16S pyrotag libraries reveals that in addition to harboring the dominant fungal crop, these ecosystems contain abundant populations of Enterobacteriaceae, including the genera Enterobacter, Pantoea, Klebsiella, Citrobacter and Escherichia. We show that these bacterial communities possess genes associated with lignocellulose degradation and diverse biosynthetic pathways, suggesting that they play a role in nutrient cycling by converting the nitrogen-poor forage of the ants into B-vitamins, amino acids and other cellular components. Our metaproteomic analysis confirms that bacterial glycosyl hydrolases and proteins with putative biosynthetic functions are produced in both field-collected and laboratory-reared colonies. These results are consistent with the hypothesis that fungus gardens are specialized fungus–bacteria communities that convert plant material into energy for their ant hosts. Together with recent investigations into the microbial symbionts of vertebrates, our work underscores the importance of microbial communities in the ecology and evolution of herbivorous metazoans.     

Effect of Hirsutella rhossiliensis on Infection of Potato by Pratylenchus penetrans

We evaluated the ability of the nematode-pathogenic fungus Hirsutella rhossiliensis (Deuteromycotina: Hyphomycetes) to reduce root penetration and population increase of Pratylenchus penetrans on potato. Experiments were conducted at 24 C in a growth chamber. When nematodes were placed on the soil surface 8 cm from a 14-day-old potato cutting, the fungus decreased the number entering roots by 25%. To determine the effect of the fungus on population increase after the nematodes entered roots, we transplanted potato cuttings infected with P. penetrans into Hirsutella-infested and uninfested soil. After 60 days, the total number of nematodes (roots and soil) was 20 ± 4% lower in Hirsutella-infested than in uninfested soil.     

Non-linear global optimization via parameterization and inverse function approximation: an artificial neural networks approach

In this article, a novel technique for non-linear global optimization is presented. The main goal is to find the optimal global solution of non-linear problems avoiding sub-optimal local solutions or inflection points. The proposed technique is based on a two steps concept: properly keep decreasing the value of the objective function, and calculating the corresponding independent variables by approximating its inverse function. The decreasing process can continue even after reaching local minima and, in general, the algorithm stops when converging to solutions near the global minimum. The implementation of the proposed technique by conventional numerical methods may require a considerable computational effort on the approximation of the inverse function. Thus, here a novel Artificial Neural Network (ANN) approach is implemented to reduce the computational requirements of the proposed optimization technique. This approach is successfully tested on some highly non-linear functions possessing several local minima. The results obtained demonstrate that the proposed approach compares favorably over some current conventional numerical (Matlab functions) methods, and other non-conventional (Evolutionary Algorithms, Simulated Annealing) optimization methods.     

Chrysene bioconversion by the white rot fungus <Emphasis Type="Italic">Pleurotus ostreatus</Emphasis> D1

The effect of cultivation conditions on chrysene bioconversion by the fungus Pleurotus ostreatus D1 was shown. Under the laccase production conditions, transformation of this polycyclic aromatic hydrocarbon occurs with accumulation of the quinone metabolite. Under both the laccase and versatile peroxidase production conditions, chrysene degradation occurs, with the stages leading to phthalic acid formation and its further utilization. The formation of phthalic acid as a metabolite of chrysene degradation by white rot fungi was revealed for the first time. The data obtained suggest that the laccase revealed on the mycelial surface and the extracellular laccase are probably involved at the initial stages of chrysene metabolism, whereas versatile peroxidase seems to be required for oxidizing the metabolites formed.     

Screening of oxylipins for control of oilseed rape (Brassica napus) fungal pathogens

Oxylipins are products of oxygenase-catalyzed reactions of fatty acids. Oxylipins have been found or implied to participate in a variety of different functions in or between organisms. In this report we investigated the potential of various naturally occurring oxylipins found in plants for their effects as fungicides on a number of fungal pathogens interfering with Brassica cultivation. The fungi investigated were Alternaria brassicae, Leptosphaeria maculans, Sclerotinia sclerotiorum and Verticillium longisporum. An in vitro growth inhibition assay was used, where the relative growth rate of the fungi were determined in the presence of various concentrations of oxylipins. While no universal fungicidic effect was found for the 10 compounds investigated there were examples of oxylipins having inhibitory effects. In certain cases the inhibitory effects was overcome by time, however. Since several of the oxylipins tested were found to be stable in the absence of the fungus this effect could be explained by induction of the degrading capacity of the fungus or increased tolerance. Several of the oxylipins also inhibited germination of L. maculans spores but the relative potency differed compared to the effects on hyphae. The study suggests that selected oxylipins may be used for disease control on Brassica plants.     

Complement-mediated tail degradation of Neodiplostomum seoulense cercariae

The furcocercus cercariae of Neodiplostomum seoulense (Digenea: Neodiplostomidae) penetrate the skins of tadpoles and shed their tails. The speculated mechanism of this tail loss was physical efforts required to produce a vigorous zigzag motion during skin penetration; no other mechanism has been proposed. We examined the relationship between the host serum and cercarial tail loss. Cercariae of N. seoulense were collected from experimentally infected Segmentina hemisphaerula, and lots of 300 cercariae were cultured in medium 199 contained several types of sera. Cercarial tail degradation was induced in all media, but all the cercariae cultured except those cultured in media containing fetal bovine serum (FBS) died within 48 hr. After 72 hr cultivation in media containing FBS, cercarial tail degradation was induced in 67.0%; in continuous cultivation 13.3% of larvae survived for 7 days. Tail degradation did not occur in the absence of serum and when serum was heat inactivated at 56 degrees Celsius for 30 min. The addition of 20 mM ethylenediaminetetraacetic acid (EDTA) blocked cercarial tail degradation completely. Moreover, the addition of 20 mM MgCl2 restored tail degradation blocked by EDTA. These results suggest that the alternative complement pathway is related with the N. seoulense cercarial tail degradation induced by serum.     

Loss of pectin is an early event during infection of cocoyam roots by Pythium myriotylum

Cocoyam (Xanthosoma sagittifolium) is an important tuber crop in most tropical zones of Africa and America. In Cameroon, its cultivation is hampered by a soil-borne fungus Pythium myriotylum which is responsible for root rot disease. The mechanism of root colonisation by the fungus has yet to be elucidated. In this study, using microscopical and immunocytochemical methods, we provide a new evidence regarding the mode of action of the fungus and we describe the reaction of the plant to the early stages of fungal invasion. We show that the fungal attack begins with the colonisation of the peripheral and epidermal cells of the root apex. These cells are rapidly lost upon infection, while cortical and stele cells are not. Labelling with the cationic gold, which binds to negatively charged wall polymers such as pectins, is absent in cortical cells and in the interfacial zone of the infected roots while it is abundant in the cell walls of stele cells. A similar pattern of labelling is also found when using the anti-pectin monoclonal antibody JIM5, but not with anti-xyloglucan antibodies. This suggests that early during infection, the fungus causes a significant loss of pectin probably via degradation by hydrolytic enzymes that diffuse and act away from the site of attack. Additional support for pectin loss is the demonstration, via sugar analysis, that a significant decrease in galacturonic acid content occurred in infected root cell walls. In addition, we demonstrate that one of the early reactions of X. sagittifolium to the fungal invasion is the formation of wall appositions that are rich in callose and cellulose.     

Patterns of functional enzyme activity in fungus farming ambrosia beetles

ABSTRACT: INTRODUCTION: In wood-dwelling fungus-farming weevils, the so-called ambrosia beetles, wood in the excavated tunnels is used as a medium for cultivating fungi by the combined action of digging larvae (which create more space for the fungi to grow) and of adults sowing and pruning the fungus. The beetles are obligately dependent on the fungus that provides essential vitamins, amino acids and sterols. However, to what extent microbial enzymes support fungus farming in ambrosia beetles is unknown. Here we measure (i) 13 plant cell-wall degrading enzymes in the fungus garden microbial consortium of the ambrosia beetle Xyleborinus saxesenii, including its primary fungal symbionts, in three compartments of laboratory maintained nests, at different time points after gallery foundation and (ii) four specific enzymes that may be either insect or microbially derived in X. saxesenii adult and larval individuals. RESULTS: We discovered that the activity of cellulases in ambrosia fungus gardens is relatively small compared to the activities of other cellulolytic enzymes. Enzyme activity in all compartments of the garden was mainly directed towards hemicellulose carbohydrates such as xylan, glucomannan and callose. Hemicellulolytic enzyme activity within the brood chamber increased with gallery age, whereas irrespective of the age of the gallery, the highest overall enzyme activity were detected in the gallery dump material expelled by the beetles. Interestingly endo-beta-1,3(4)-glucanase activity capable of callose degradation was identified in whole-body extracts of both larvae and adult X. saxesenii, whereas endo-beta-1,4-xylanase activity was exclusively detected in larvae. CONCLUSION: Similar to closely related fungi associated with bark beetles in phloem, the microbial symbionts of ambrosia beetles do not degrade cellulose. Instead, their enzyme activity is directed mainly towards comparatively more easily accessible hemicellulose components of the ray-parenchyma cells in the wood xylem. Furthermore, the detection of xylanolytic enzymes exclusively in larvae and not in adults indicates that larvae (pre-) digest plant cell wall structures exclusively in larvae (which feed on fungus colonized wood) and not in adults (which feed only on fungi). This implies that in X. saxesenii and likely also in many other ambrosia beetles, adults and larvae do not compete for the same food within their nests - in contrast, larvae increase colony fitness by facilitating enzymatic wood degradation and fungus cultivation.