Evidences on the Ability of Mycorrhizal Genus Piloderma to Use Organic Nitrogen and Deliver It to Scots Pine

RESEARCH ARTICLE Evidences on the Ability of Mycorrhizal Genus Piloderma to Use Organic Nitrogen and Deliver It to Scots Pine Jussi Heinonsalo1*, Hui Sun1,2, Minna Santalahti1, Kirsi Bäcklund1, Pertti Hari2, Jukka Pumpanen2 1 Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland, 2 Department of Forest Sciences, University of Helsinki, Helsinki, Finland a11111 * Abstract OPEN ACCESS Citation: Heinonsalo J, Sun H, Santalahti M, Bäcklund K, Hari P, Pumpanen J (2015) Evidences on the Ability of Mycorrhizal Genus Piloderma to Use Organic Nitrogen and Deliver It to Scots Pine. PLoS ONE 10(7): e0131561. doi:10.1371/journal. pone.0131561 Editor: Erika Kothe, Friedrich Schiller University, GERMANY Received: April 2, 2015 Accepted: May 2, 2015 Published: July 1, 2015 Copyright: © 2015 Heinonsalo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper and its Supporting Information files, and Sequence data is deposited in EMBL database under accession numbers AM910819, LK052852, LK052848, FN556986, LK052844, AM910820 and LK939126-LK939128. Funding: This research was supported by grants from the Academy of Finland to JH (263858), JP (130984, 218094 and 255576) and KB (ICOS 271878, ICOS-Finland 281255 and ICOS-ERIC 281250). HS and MS were supported by the Research Funds of the University of Helsinki Ectomycorrhizal (ECM) symbiosis has been proposed to link plant photosynthesis and soil organic matter (SOM) decomposition through the production of fungal enzymes which promote SOM degradation and nitrogen (N) uptake. However, laboratory and field evidence for the existence of these processes are rare. Piloderma sp., a common ECM genus in boreal forest soil, was chosen as model mycorrhiza for this study. The abundance of Piloderma sp. was studied in root tips and soil over one growing season and in winter. Protease production was measured from ectomycorrhiza and soil solution in the field and pure fungal cultures. We also tested the effect of Piloderma olivaceum on host plant organic N nutrition in the laboratory. The results showed that Piloderma sp. was highly abundant in the field and produced extracellular proteases, which correlated positively with the gross primary production, temperature and soil respiration. In the laboratory, Piloderma olivaceum could improve the ability of Pinus sylvestris L. to utilize N from extragenous proteins. We suggest that ECM fungi, although potentially retaining N in their hyphae, are important in forest C and N cycling due to their ability to access proteinaeous N. As Piloderma sp. abundance appeared to be seasonally highly variable, recycling of fungal-bound N after hyphal death may therefore be of primary importance for the N cycling in boreal ecosystems. Introduction The balance between soil organic matter (SOM) decomposition and litter accumulation is of primary importance to the boreal forest carbon (C) budget. Boreal forest soil is one of the largest terrestrial C pools [1] and any changes in the C balance in this ecosystem could have global consequences [2]. Boreal forests are typically dominated by trees that form symbiosis with ectomycorrhizal fungi (ECM) and by understory vegetation that forms ericoid mycorrhizal (ERM) associations [3]. In the ECM symbiosis, the plant delivers C compounds fixed during photosynthesis to the ECM which in turn assists the host plant in nutrient and water uptake [3]. ECM also play a role in protecting the plant against attack by harmful microbial pathogens PLOS ONE | DOI:10.1371/journal.pone.0131561 July 1, 2015 1 / 17 Piloderma sp. Help Host Plants Use Soil Organic N (490127). In addition, this study was part of the Academy of Finland Finnish Centre of Excellence program (project 1118615). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. [3]. In ECM ecosystems, SOM accumulates and nutrients are present in an organic form resulting in an ‘organic nutrient economy’ [4]. In order to acquire N and P, trees translocate large amounts of carbohydrates belowground to microbes and particularly ECM, which links soil processes to forest productivity [3]. Surprisingly ECM and ERM ecosystems have been shown to store almost twice as much C per unit of soil N than arbuscular mycorrhizal ecosystems, indicating that the type of mycorrhizal symbiosis is an important determinant for soil C storage [5]. The incorporation of interactions between the C and N cycles into climate system models has therefore been recommended [6]. Recently it has been shown that the decomposition of recalcitrant, slow-cycling SOM pool is accelerated by easily available sources of energy, such as cellulose or glucose [7,8], in a process called priming [9]. The easily utilizable carbohydrates facilitate the production of extracellular enzymes needed for SOM decomposition by microorganisms. The priming effect is not only dependent on a C source but also on other factors, including the nutritional status of the soil, and in some cases, neither priming nor even ‘negative priming’ has been observed [9]. Recent studies on boreal forest soils have shown that the priming effect is connected to N uptake, and as boreal forests are often N-limited, N scavenging from complex organic molecules has been proposed to be the main trigger of the priming effect [10, 11]. There is increasing body of evidence, which suggests that ECM fungi play a key role in priming, soil organic N transformations and SOM decomposition [12–14]. In contrast to wood-decomposing fungi, ECM fungi are not known to produce significant quantities of peroxidases (e.g. manganese peroxidase, MnP) that degrade complex, lignin-containing structures. However, there is evidence that MnP-like peroxidases may be produced by the common ECM genus Cortinarius, and class II peroxidase-encoding genes have been identified in a wide range of ECM fungi [15]. High peroxidase activity and DNA quantity from Cortinarius species were co-localized in boreal forest soil [13]. Recently, Rineau et al. [16] demonstrated that ECM fungus Paxillus involutus may degrade SOM using a Fenton-reaction based mechanism in a similar way as brown-rot fungi. Laccase, a non-specific oxidative enzyme, is known to be produced by many fungi, including ECM [17–19]. Multiple laccase genes are encoded in the genome of Piloderma sp. [20] and their expression was related to N availability [17]. Heinonsalo et al. [19] screened 23 fungal strains and showed that laccase activities in Piloderma olivaceum pure culture were triggered by the addition of humus and Scots pine (Pinus sylvestris) sa (...truncated)