ISSN 00036838, Applied Biochemistry and Microbiology, 2013, Vol. 49, No. 6, pp. 570–580. © Pleiades Publishing, Inc., 2013. Original Russian Text © T.V. Fedorova, N.V. Shakhova, O.I. Klein, O.A. Glazunova, L.G. Maloshenok, N.A. Kulikova, N.V. Psurtseva, O.V. Koroleva, 2013, published in Prikladnaya Biokhimiya i Mikrobiologiya, 2013, Vol. 49, No. 6, pp. 570–579. Comparative Analysis of the Ligninolytic Potential of Basidiomycetes Belonging to Different Taxonomic and Ecological Groups T. V. Fedorovaa, N. V. Shakhovab, O. I. Kleina, O. A. Glazunovaa, L. G. Maloshenoka, N. A. Kulikovaa, c, N. V. Psurtsevab, and O. V. Korolevaa a Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow, 119071 Russia email: [email protected] b Komarov Botanical Institute, Russian Academy of Sciences, St. Petersburg, 197376 Russia c Department of Soil Science, Moscow State University, Moscow, 119991 Russia Received June 7, 2013 Abstract—Screening of the ligninolytic activity of basidiomycetes from the Komarov Botanical Institute Basidio mycetes Culture Collection (LEBIN), Russian Academy of Sciences, belonging to different taxonomic and eco logical groups was performed. The patterns of the position of taxa of active producers of ligninolytic enzymes in the modern system of fungi were identified. Cluster analysis showed that the group of fungi with the greatest ligni nolytic and degradation potential includes representatives of the families Pleurotaceae, Polyporaceae, and Phan erochaetaceae, which perform the first stages of wood decomposition. As a result, species of interest for the further study of their oxidative potential and use in biotechnology were selected. DOI: 10.1134/S0003683813060082 Currently, the interest towards the bioconversion of mechanism of detoxification of both lignin degrada lignocellulosic waste is steadily growing, which is tion products and various xenobiotics [6, 7]. For this determined by an increased volume of waste of the reason, researchers constantly work to search for, iso pulp and paper industry (PPI) and the woodworking late, and study new strains of basidiomycetes promis industry and, as a consequence, an increased anthro ing for use in bioconversion and bioremediation tech pogenic impact on the environment [1]. For example, nologies [8–10]. according to the Federal State Statistics Service, the This study is the first step in the development of a total amount of waste in the Russian pulp and paper biotechnology for the processing of solid waste from industry has increased from 2991.2 million tons in the pulp and paper industry using basidiomycetes. 2005 to 4303.3 million tons in 2011. At the same time, The goal of this study was to screen and assess the the main methods for disposing of waste in our coun ligninolytic and degradation potential of whiterot try remain warehousing in landfills and incineration fungi from different ecological and taxonomic groups after dewatering and compaction. and to select the most promising strains for use in the The use of biotechnological approaches for con utilization of lignincontaining waste of the pulp and verting the pulp and paper industry’s waste is limited paper industry. by its toxicity due to the presence of compounds, such as polynuclear aromatic hydrocarbons, high concen MATERIALS AND METHODS trations of xenobiotics (dioxins, chlorine derivatives, etc.), and lignin, which is one of the most difficult Screening of cultures. More than 520 cultures of biopolymers to degrade. That is why basidio 330 species of higher (basidiomycete) fungi belonging mycetes—the most active lignin destructors—attract to different taxonomic and ecological groups from the the attention of researchers as the key component of Komarov Botanical Institute Basidiomycetes Culture the lignocellulosic waste bioconversion technology. Of Collection, Russian Academy of Sciences (St. Peters all known representatives of Basidiomycota, whiterot burg), were used to assess the ligninolytic and degrada fungi are the most efficient lignin destructors [2]. The tion potential by rapid methods [11]. The most active process of degradation of lignocellulosic substrates by strains shown in Table 1 were selected for further work. these fungi consists of a large number of steps and All cultures were stored in tubes on wort agar slants depends on the presence of an enzymatic system (a (wort 4%, agar 1.5%) at 4°C. unique ligninolytic complex consisting of laccase (EC Species identification. The species of fungi were 1.10.3.2) and various peroxidases, including the man verified morphologically using conventional methods ganese (EC 1.11.1.13) and lignin (EC 1.11.1.14) per with subsequent genotyping. oxidases) in them [3–5]. It is known that some species Strains were grown on an MEA medium with 2% of whiterot fungi (wood destructors) have a unique malt extract (DiaM, Russia) and 1.6% agar (Difco, 570 COMPARATIVE ANALYSIS OF THE LIGNINOLYTIC POTENTIAL OF BASIDIOMYCETES 571 Table 1. Characteristics of fungal strains studied Taxonomic Strain Species affiliation Geographic origin Species ecology affiliation LEBIN Antrodiella fagi Steccheri Russia, the Far East. Secondary xylotroph (typical of heavily decayed 1998 nea Vampola & naceae, The substrate is dry wood or remaining on it for 15 years and longer), Pouzar Polyporales branches of deciduo which causes white rot. Colonizes dead twigs, large us trees branches, and trunks of deciduous trees, more rarely shrubs, of the genera Alnus, Populus, Salix, Padus, Corylus, and Syringa. LEBIN Coriolopsis Polyporaceae, Cuba Xylotroph, which causes white rot. Lives on decom 0677 caperata (Berk.) Polyporales posed wood of angiosperms in serried forest commu Murrill nities and tropical mangroves. LEBIN Lenzites Polyporaceae, Finland, Lahti district. Wound primary xylotroph performing the first stage 2047 betulina (L.) Fr. Polyporales The substrate is a birch of wood decomposition (first seven years), which cau Betula sp. trunk in a co ses white rot. Lives on wood of deciduous trees, mo niferous forest stly on species of the genera Betula, as well as Acer, Al nus, Carpinus, Corylus, Fagus, Fraxinus, Juglans, Pru nus, Quercus, Salix, Sorbus, Tilia, and Ulmus. LEBIN Byssomerulius Phanerocha Russia, the Far East. Xylotroph, which causes white rot. Lives on dead and 2009 avellaneus etaceae, Poly The substrate is snag deadanddown branches, as well as on litter of broad (Bres.) J. Erikss. porales branches in an oak leaved trees of the genera Acer, Alnus, Carpinus, and & Hjortstam grove Quercus. LEBIN Pleurotus ostrea Pleurotaceae, Russia, Sochi Primary wound xylotroph (performing the first stage of 0432 tus (Jacq.) P. Agaricales wood decomposition (first seven years), which causes Kumm. mixed yellow rot. It lives on weakened and dying trees and dead branches and stumps of many deciduous and more rarely coniferous (Picea, Araucaria) tree species. LEBIN Peniophora lycii Peniophora Russia, Rostov oblast. Xylotroph, which lives on dead branches of various de 2142 (Pers.) Höhn. & ceae, Russu The substrate is creta ciduous (Acer, Fagus, Rhamnus, and Fraxinus) and oc Litsch. lales ceous sediments in the casionally coniferous (Pinus) trees. steppe LEBIN Steccherinum Steccheri Russia, the Far East, Secondary xylotroph typical of heavily decayed wood 1963 murashkinskyi naceae, Poly Kamchatka region. or remaining on it for 15 years and longer. Causes (Burt) Maas porales Found on a charred white rot. Lives on dead trunks and branches of va Geest. trunk of the birch Be rious deciduous (Acer, Betula, Populus, Salix, and tula platyphylla Quercus) and occasionally coniferous (Abies) trees. LEBIN Trametes gibbosa Polyporaceae, Russia, Samara Xylotroph, which causes actively spreading white 1911 (Pers.) Fr. Polyporales oblast, Stavropol dis rot. Lives on dead (mostly largesized) wood (dead trict. Found on lime wood, stumps, snags, and treated wood) of decidu ous (usually, on Betula and, more rarely, on Alnus, Carpinus, Cerasus, Fagus, Populus, Salix, Tilia, etc.) and, occasionally, coniferous (Abies, Picea) tree spe cies. A highly competitive species able to slowly dis place many species of wooddecaying fungi on the majority of substrates. LEBIN Xerula radicata Marasmi Russia, Samara oblast, Xylotroph, which develops on the roots (and possibly on 1795 (Relhan) Dörfelt aceae, Agari Stavropol district. At the buried wood) of mostly deciduous trees (Quercus, cales the base of a heavily Fagus). Fruiting bodies are usually found at the base of damaged stump old heavily damaged tree stumps, which allows this spe cies to be regarded as a secondary xylotroph. United States) in Petri dishes (90 mm) at 25°С in the ods [12, 13]. The macromorphological characteristics dark. The growth of the strains was characterized by included descriptions of the aerial mycelium, color, the diameter of the colony on days 7 and 14 of growth. smell, and reversum of the colonies. The micromor The characterization of the macromorphological and phological description included the characteristic fea micromorphological features of the mycelium was tures of the hyphal system and the presence of swell performed for 4weekold colonies by standard meth ings, buckles, and anamorphic elements (asexual APPLIED BIOCHEMISTRY AND MICROBIOLOGY Vol. 49 No. 6 2013 572 FEDOROVA et al. reproductive structures) on the mycelium. The teleo inoculation, the inoculum was crushed with porcelain morph stage (sporocarp formation) in the culture was beads at 180 rpm for 20 min to form a homogeneous obtained on substrate blocks (birch sawdust with suspension,