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Ectomycorrhizal Communities in a Tuber Aestivum Vittad. Orchard in Poland

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Ectomycorrhizal Communities in a Tuber Aestivum Vittad. Orchard in Poland Open Life Sci. 2016; 11: 348–357 Research Article Open Access Dorota Hilszczańska*, Hanna Szmidla, Jakub Horak, Aleksandra Rosa-Gruszecka Ectomycorrhizal communities in a Tuber aestivum Vittad. orchard in Poland DOI 10.1515/biol-2016-0046 Received July 6, 2016; accepted October 27, 2016 1 Introduction Abstract: Cultivation of the Burgundy truffle, Tuber Burgundy truffle (Tuber aestivum Vittad.) is an aestivum Vittad., has become a new agricultural alternative ectomycorrhizal fungus that forms edible hypogeous in Poland. For rural economies, the concept of landscaping ascocarps of considerable economic value. It is well- is often considerably more beneficial than conventional documented in literature that T. aestivum grows in an agriculture and promotes reforestation, as well as land-use ectomycorrhizal symbiosis with many different trees and stability. Considering examples from France, Italy, Hungary shrubs belonging to genera such as Carpinus, Fagus, Tilia, and Spain, truffle cultivation stimulates economic and Populus, Quercus and Corylus [1-3]. social development of small, rural communities. Because Cultivation of the fungus is starting to become a there is no long tradition of truffle orchards in Poland, promising agroforestry alternative for rural areas in knowledge regarding the environmental factors regulating Poland. For a long time, truffles, especially the species the formation of fruiting bodies of T. aestivum is limited. praised by chefs and gourmets for their scent and taste, Thus, knowledge concerning ectomycorrhizal communities were considered rare in Poland, and the Burgundy truffle of T. aestivum host species is crucial to ensuring successful was recorded only once after the Second World War [4]. In Burgundy truffle production. We investigated the the last decade, new data on the distribution of T. aestivum persistence of T. aestivum ectomycorrhizae on roots of and other species of truffles have been reported from hazel (Corylus avellana L.) and oak (Quercus robur L.) and Poland [5,6], as well as from other countries of Central, checked the host-species influence on community structure Eastern and Northern Europe [7-11]. of ectomycorrhizal fungi. The study was conducted in an The persistence of truffle ectomycorrhizae on experimental plantation located in eastern Poland and inoculated seedlings outplanted in the field is one of the established in 2008. We demonstrated that the number most important factors of truffle cultivation success [12-14]. of fungal taxa was not significantly different between However, in many cases, the targeted truffle ectomycorrhizae oak and hazel. However, the species composition differed is replaced by other competitive ectomycorrhizal species between these two host trees. During the three-year study, [15,16]. Understanding the conditions that promote truffle we observed that species richness did not increase with the fructification is essential for detecting species that have age of the plantation. a high possibility of outcompeting targeted truffles [16]. Knowledge about the diversity of species is crucial for Keywords: Burgundy truffle, ectomycorrhizae, hazel, successful conservation efforts and future plantation oak, truffle orchard management. Comparison of the ectomycorrhizal fungi that are associated with different host species within environments with the same conditions would help clarify *Corresponding authors: Dorota Hilszczańska, Department of the importance of host preference in structuring the Forest Ecology, Forest Research Institute, Braci Leśnej 3 Str., ectomycorrhizal community [17]. Sękocin Stary, 05-090 Raszyn, Poland, The aims of our study were i) to examine whether E-mail: [email protected] and how differently T. aestivum mycorrhizae persist in Hanna Szmidla, Aleksandra Rosa-Gruszecka, Department of Forest Protection, Forest Research Institute, Braci Leśnej 3 Str., Sękocin roots of hazel (Corylus avellana L.) and oak (Quercus Stary 05-090 Raszyn, Poland robur L.), ii) to characterize other (non-Tuber) species Jakub Horak, Department of Forest Protection and Entomology, present in the orchard, and iii) to estimate, based on the Faculty of Forestry and Wood Sciences, Czech University of Life ectomycorrhizal fungi community (ECM), the future T. Sciences, Kamýcká 1176, CZ-165 21, Prague 6 – Suchdol, Czech aestivum fructification in the orchard. Republic © 2016 Dorota Hilszczańska et al., published by De Gruyter Open. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivs 3.0 License. Ectomycorrhizal communities in a Tuber aestivum Vittad. orchard in Poland 349 2 Materials and Methods 2.2 Sampling and morphological analyses To analyse the ectomycorrhizal morphotypes present in root 2.1 Study area tips, five hazel trees and five oaks were randomly chosen (every year, analyses of mycorrhizae were performed on The study was conducted in a plantation located in the same chosen trees). Three soil cores (6 cm diameter, 20 eastern Poland (latitude 51° 8’ 51’’; longitude 23° 28’ 29’’) cm deep) were obtained 25 cm from the base of each tree. at 200 m a.s.l. where the soil parent material is Triassic These three cores per tree were combined to yield a single Muschelkalk. The climate is continental with a mean composite sample. The samples were collected every year annual rainfall of 550 mm and mean annual temperature of the study (2012-2014), in September. Each composite of 8.0°C. The truffle orchard was established in 2008 by sample was individually soaked overnight in tap water and planting 134 seedlings of Quercus robur and 150 seedlings sieved to separate the root fragments and ectomycorrhizae of Corylus avellana, all of which were inoculated with (ECM) from the soil. The clean roots were placed in Petri spores of native Tuber aestivum [18]. The soil physical and dishes filled with water. Only vital roots (identified as chemical properties are shown in Table 1 and were similar swollen, without root hair or covered by fungal mantles) in all areas of the orchard. were considered ECM-colonized roots. The different types of ectomycorrhizae found in each sample were morphotyped Table 1. Physical and chemical properties of the soil in the using a stereomicroscope and a light microscope, with plantation. the anatomo-morphological characteristics described by Measured parameter Agerer [19,20] and two online databases, EctoMycorrhizal Sand (0.2-0.063 mm) % 32.62 Community DataBase (www.emyco.uniss.it) and DEEMY Clay (0.002 mm) % 35.29 (http://www.deemy.de), as references [21,22]. Silt (0.063-0.002 mm) % 32.09 Morphotypes were identified by colour and shape P 0 g kg-1 2.52 2 5 of the mycorrhiza, characteristics of the mantel surface, Calcium cmol+kg-1 27.96 Magnesium cmol+kg-1 0.32 ramification system, and presence and structure of Potassium cmol+kg-1 0.60 rhizomorphs, emanating hyphae and other elements. The water pH 7.6 morphology and colour of the ECMs were evaluated with a CaCO % 19.56 3 dissecting microscope on freshly isolated ectomycorrhizal Organic matter % 2.3 tips. The ectomycorrhizae formed by T. aestivum were C/N 24.3 identified by its ochreous and chestnut brown colour and by the presence of swollen fine roots with tips embedded The experimental plantation of 0.44 ha was established in radially twisted hyphae. The number of living on land that had been abandoned for 19 years prior to mycorrhizae of each morphotype was recorded separately outplanting of seedlings inoculated with of T. aestivum. for each sample. Roots were cut into 2-cm pieces and a Before outplanting of seedlings, the soil was ploughed total of 100 vital tips were counted. to a depth of 50 cm to promote deep soil aeration, and superficial tilling with several passes was later applied to remove herbaceous vegetation. The site was chosen 2.3 Data analysis because truffle species, such as Tuber rufum and T. excavatum, had been found in the forest surrounding the The diversity of ectomycorrhizae on hazel and oak was area. This forest was mainly composed of Q. robur, Carpinus expressed as the number of identified ectomycorrhizal betulus and Populus tremula, with C. avellana in the species (species richness). The relative abundance of each understorey. Seedlings were arranged with 3 m between morphotype (number of root tips of each morphotype / the rows and 4.5 between individuals within the rows. total number of mycorrhizae) was calculated separately Green woven polypropylene fabrics (99 g m-2) were used for each sample. Ecological indicators, such as species in rows for reducing herbaceous cover in this plantation. richness, Shannon diversity index and Simpson In order to control the rapid growth of weeds, soil tilling dominance index, were calculated using EcoComPaC with cultivator tines set at 15 cm depth and manual weed Version 1 [23] (http://prf.osu.cz/kbe/dokumenty/sw/ control with a hoe were performed every year – namely, ComEcoPaC/ComEcoPaC.xls.) in Excel. in May and in August or September, depending on weed Statistical analyses were performed in EstimateS 8.2. growth. and CANOCO 4.5. We focused on predictors that were 350 D. Hilszczańska, et al. testable only within a limited spatial scale. Unreplicated Morphological observations revealed a total of 14 fungal treatments were the only option for our study [24]. We taxa: 10 in oak and 10 in hazel (Table 2 and Fig. 1 a-n). controlled this problem by using randomized techniques Of these, one was assigned at the order level (Pezizales), for taxa richness data [25], as recommended and used seven at the genus level (Inocybe sp., Lactarius sp. 1, elsewhere [26-28]. Lactarius sp. 2, Tomentella sp., Geopora sp., Peziza sp. For the analysis of species richness and for and Alnicola sp.) and five at the species level (Tuber comparisons between tree species and over time, we used aestivum, Amphinema byssoide, Cenococcum geophilum, sample-based species rarefactions (Mao Tau function) Scleroderma areolatum, Laccaria tortilis and Thelephora with 95% confidence intervals [29]. The number of terrestris). In our study, T. aestivum ectomycorrhizae were randomizations was set at 1,000 in EstimateS.
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