Temporal changes in diversity and expression patterns of fungal laccase genes within the organic horizon of a brown forest soil Harald Kellner a, b, * , Patricia Luis c , Bettina Schlitt a, b , François Buscot a, b a UFZ - Helmholtz Centre for Environmental Research (Leipzig-Halle) Ltd., Department of Soil Ecology, Theodor-Lieser-Str. 4, D-06120 Halle/Saale, Germany b University of Leipzig, Institute of Botany -Terrestrial Ecology, Johannisallee 21, D-04103 Leipzig, Germany c Universite´ de Lyon, Universite´ Lyon 1, Ecologie Microbienne, UMR CNRS 5557, USC INRA 1193, Ba ˆtiment Andre ´ Lwoff, 43 Boulevard du 11 Novembre 1918, F-69622 Villeurbanne Cedex, France article info Article history: Received 11 December 2008 Received in revised form 10 March 2009 Accepted 23 March 2009 Available online 21 April 2009 Keywords: Ascomycetes Basidiomycetes Laccase gene expression Lignin degradation abstract Temporal fluctuations of present and expressed fungal genes encoding the ligninolytic enzyme laccase were examined bimonthly between March 2004 and April 2005 in the organic horizon of a beech forest Cambisol. Using two sets of degenerate primer pairs, we detected 73 different basidiomycete laccase genes from soil DNA extracts and 42 different transcripts of asco- and basidiomycetes from soil RNA extracts (cDNA). Phylogenetic analysis related the sequences to fungal taxa. The highest fungal laccase gene richness in soil DNA and RNA samples were found in August, October and January, and followed the input of fresh litter into soil. The highest change of the fungal laccase gene population was observed from October to January, but no distinct temporal change in the total soil laccase activity was measured. Present and expressed laccase gene populations were highly different, implying different subsets amplified with our primer sets and likely impacting future research strategies. Despite considerable variations in gene presence and expression, we found a steady expression and high soil enzyme activity of fungal laccases at each sampling date, thus presumably laccases have major impacts to soil organic matter turnover and stabilization processes. Ó 2009 Elsevier Ltd. All rights reserved. 1. Introduction The accumulation and turnover of soil organic matter (SOM) result from a balance between mineralization and stabilization due to the recalcitrance of some compounds and the formation of low bioavailable organo-mineral complexes (von Lu ¨ tzow et al., 2006). Soil microorganisms are largely involved in both mineralization and stabilization processes, and in forest ecosystems fungi are key players (Berg and McClaugherty, 2003; Osono, 2007). In deciduous forests, plant litter is one major annual resource input for soil fungal communities, and it is colonized in relatively short time scales (Berg and Gerstberger, 2004). The first colonizers, i.e. yeasts, zygomycetes and several ascomycetes, use easy available soluble compounds before being replaced by asco- and basidiomycetes able to attack insoluble and more recalcitrant substances such as cellulose and lignin (Dighton, 1997; Koide et al., 2005). Recently, molecular techniques allow a better understanding and monitoring of these fungal dynamics directly in soil. In a recent study on the degradation of leaf litter in a native and foreign habitat, it was demonstrated that the general degradation capability was site specific and probably driven by fungal adaptation to the local leaf litter type (Aneja et al., 2006). Thus, degradation of sugars, starch, cellulose and lignin by the native communities was higher. In course of the degradation, the typical increase in the proportion of lignin was observed (Aneja et al., 2006). Lignin, the second most abundant biopolymer in nature, is largely resistant against microbial attack (Ko ¨ gel-Knabner, 2002) but is degraded by fungi producing ligninolytic oxidative exoenzymes (Thorn, 1997). Monitoring changes in fungal communities that produce such exoenzymes provides information on their potential role in temporal SOM degradation processes. Instead of monitoring fungal species based on their ribosomal RNA genes or spacer regions, studies following this line should analyze genes encoding exoenzymes such as phenol oxidases (laccases), manganese- and lignin-peroxidases, b-glucosidases or cellobiohydrolases that degrade plant litter (Zak et al., 2006; Edwards et al., 2008). Within fungal oxidative exoenzymes, laccases are one of the most universal. They catalyze the oxidation of various aromatic * Corresponding author. FUSAGx, Unite ´ de Biologie animale et microbienne, Avenue Mare ´chal Juin 6, B-5030 Gembloux, Belgium. Tel.: þ32 81 622355; fax: þ32 81 611555. E-mail address: mail@haraldkellner.com (H. Kellner). URL: http://www.haraldkellner.com/html/laccase_project.html Contents lists available at ScienceDirect Soil Biology & Biochemistry journal homepage: www.elsevier.com/locate/soilbio 0038-0717/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2009.03.012 Soil Biology & Biochemistry 41 (2009) 1380–1389