Some endophytes of Juncus trifidus from Tatra Mts. in Poland

This is a first part of work devoted highland rush endophytes: Penicillium expansum, Cladosporium oxysporum, Arthrinium state of Apiospora montagnei and Aureobasidium pullulans . The basidiomycete strain, possibly Lagarobasidium detriticum was also isolated.

Host plant.Juncus trifidus l. (highland rush) belongs to the section Steirochloa which is a sister group of Luzula (Drábkova et al. 2003).J. trifidus subsp.trifidus is the amphi-atlantic perennial plants of boreal-arctic-alpine distribution.The species is divided into three subspecies.Of them J. trifidus subsp.carolinianus occurs in north America, J. trifidus subsp.monanthos is restricted to calcareous area of Alps and Appenines.The typical taxon J. trifidus subsp.trifidus occurs in silicate mountains of europe, Greenland and northern Asia.The geographical distribution of the plant indicates its european origin (böcher 1972).Plants from north America resembling those from Greenland and Scandinavia and differ in many characters from strains from Pyrenees and Tatra Mts.(böcher 1972).during the Pleistocene it spread in the Arctic, reaching central Siberia in the East (Kulczyński 1924).In Poland it occurs in some localities such as the karkonosze Mts.(the Sudetes), Mt.Babia Góra Range and Tatra Mts.(the Carpathians).The locality in Mt.Śnieżnik in the Central Sudetes, reported by Ciaciura (1988), is situated on the Czech side of this mountain.highland rush is perennial caespitose rush forming dense tufts.Aerial stems are stiff and erected, leaves alternate, simple with acuminate apices.Inflorescence cymose or head-like.Number of chromosomes of Polish populations 2n=30 (Skalińska et al.1957).It is pioneering alpine species with effective strategy of colonization of disturbed sites (Marchand, Roach 1980).

MeThodS
Material was collected on NE slope of Mt.Małołączniak in the Tatra Mts.(49˚ 55'N, 19˚ 14'E).19 specimens of Juncus trifidus were screened for the presence of fungal endophytes.Small part of tuft including culms with rots were removed from the soil into sterile unclosed zip-lock plastic bags and putted in the refrigerator.After 3 days they were moving from the bags and processed according to isolation procedures (bacon 1990).Three green culms of each sample were selected, central parts of each culm were cut into two segment ca 3 cm long.Six segments from each sample were used (totally 114 segments).All segments were dipped for 1 min in 96% ethanol, then surface sterilized for 3 min in 3% chlorox (naoCl), rinsed in ethanol for 1 min and transferred to 90 mm Petri plates with Ferency medium and then were incubated in room temperature in daylight.obtained isolates of different fungi were then removed on PDA and MAA media and incubated in room temperature.Taxa of fungi were identified on the basis of cultural characteristics and the morphology of hyphae and conidia.

reSulTS
Totally 114 segments were used.11 strains of different fungi were isolated.of them five taxa are presented in this article.Sacc., Nuovo Giorn. Bot. Ital. 7: 306, 1875.Colonies superficial, widely effused, produced mycelial mats, mycelium colourless to pale olivaceous brown, composed of network of septate hyphae anastomosing and branched, smooth ca 3 μm diam, conidiophores colourless, smooth 1.5-2 μm diam., conidia brown 5-9 × 4-5 μm with a hyaline band at the junction of the two sides.The fungus started growth after 5 months from two segments and overgrow every mycelia in the dishes.

Aureobasidium pullulans (De
Comments.This black yeast-like fungus (meristematic fungus) is ubiquitous saprophyte of phyllosphere and other aerial plant parts (domsch, Gams and Anderson 1980).It is frequently noted as endophyte of different plants, among them in bark and xylem of Eucalyptus (bettucci et al. 1999), leaf fern Pteridium aquilinum (Fisher 1996), twigs, leaves and buds of Acer pseudoplatanus and other trees (Pugh, buckley 1971).A. pullulans can be antagonistic against a number of phytopathogenic fungi (Andrews et al. 1983).Schena et al. (2002) noted high protection levels of A. pullulans against Penicillium digitatum, Botrytis cinerea and Monilia laxa. A. pullulans is also noted on painted wood, stone and glass (Yurlova et al. 1999).

Penicillium expansum
Comments.Common fungus on fruits, meat, paper, various rotting substrates and soil (Domsch et al. 1980), known as Sorbus endophyte (Samson, Frisvad 2004).Also batista et al. (2003) and Vega & Posada (2006) informed that fungi from the genera Penicillium and Aspergillus where isolated as endophytes.P. expansum strongly suppressed growth of black yeast strain growing on nearest segment.The white mycelium was growing on five segments of culm.Mycelium transferring on MAA medium has 54 mm diam.after five days and 80 mm after seven days.Mycelium white, velvety, pale yellow underside.At the beginning aerial mycelium cover all dish.Hyphae 3-5 μm diam., with clamps, spores smooth, ellipsoid 6-6.5 × 5-5.5 μm diam.with a small apiculus and big oil drop.Crystals not seen.Cystidia not present, however similar structures were seen.

Basidiomycete endophyte, possibly
Comments.Size of spores resemble L. detriticum.Eriksson and Ryvarden (1976) noted L. detriticum on leaves of Juncus, Carex, Eqisetum, ferns, on rotten woods of Juniperus, Alnus, Betula, Populus and Sorbus.basidiomycete endophytes are not often isolated from plants.Some species from the genus Peniophora were isolated from xylem and bark of Eucalyptus globulus and they probably initiated the decomposition process (bettucci et al. 1999).diSCuSSion Totally 11 strains of different fungi were isolated.of them a black yeast, basidiomycete fungus, DSE and non-sporulating colonies will be subject of the next article.In this article are presented first result of presence of endophytes in the highland rush stems.Four isolated species of fungi have worldwide general distribution and were earlier noted as endophytes of various plants.The precise determination of the most interesting basidiomycete strain need sequencing of iTS region (nilson 2003).Also comparison of frequency of various ecological groups of endophytes needs further investigation.So far any endophytes were isolated from stems of highland rush.
Acknowledgements.I especially thank to anonymous reviewer for important comments.This work was supported by the Ministry of Science and Information Society Technologies, Poland (Project No. 2 P04F 066 28 and N N304 328336).