Cephalotrichum stemonitis as a biofilm inhabitant in the gold mine in Poland

Cephalotrichum stemonitis and its synanamorph Echinobotryum atrum isolated from bacterial biofilm is presented.

basidiomycete detritus, manore pile, bronchial washing, right feel, leaves of needleleafed tree (Abbott 2000). C. stemonitis was noted in coyote and rat dung, indoor air of honeybee (Apis mellifera) overwintering facility, cone of white spruce, sandy soil, decayed wood of white spruce, soil of elm woods, agricultural soil. It is also important colonizer of pPVC buried in soil (Sabev et al. 2006). these authors mentioned it as Doratomyces spp. in the article, but in GenBank it is more precisely determined as D. stemonitis. Malloch and Hubart (1987) described unnamed species of Microascus from Ramoil Cave. It means that fungi from this group can inhabit also underground environments. Fassatiova (1970) noted C. stemonitis on wood in uranium mine in Czech Republic.

StuDY AREA AND MEtHoDS
Gray biofilms of rock-inhabiting bacteria in Gertruda Adit in a closed gold mine located in Złoty Stok in Lower Silesia were chosen for microbial analysis. This mine posseses constant conditions such as low temperature ca 10º C, darkness, high humidity, high As concentration and other toxic substances (Chlebicki et al. 2005). So far 12 species of fungi were noted in this mine (Chlebicki et al. 2005;Chlebicki, Lorenc 2006). Bacteria with fungi were collected in sterile plastic tubes and refrigeratorated at 10º C. Fungal growth was performed on DRBC, RBC, YMA and PDA media. Inoculated media on Petri dishes were putted in incubator at 10º C. the morphological characters of the living fungi were examined in water and cotton blue in lactophenol using light microscopy (Nikon SMZ 1500, Nikon Labophot 2 and Nikon Eclipse 800). Microphotographs were taken with these microscopes equipped with a digital camera. For scanning electron microscope (SEM) studies mycelium was coated with gold, and photographed using a LEo 1430 VP Zeiss microscope with a working distance of ca 10 mm. Fungus-species nomenclature follows Abbott (2000).  (1837) Description: mycelium creamy-white to dark brown, after 7 days on PDA 17,5-21 mm diam., on RBC 17,0 mm diam., and on DRBC 13-17 mm, all in room temperature (Fig. 1A, B, D). First conidiophores of synanamorph -Echinonobotryum atrum -appeared throughout the mycelium after 12 days on PDA. Mycelium growing on RBC after two weeks produced rings of densely distributed synnemata of C. stemonitis in central part of the mycelium. Mycelium on PDA formed at the beginning E.
Material examined: on rock bacterial biofilm in Gertruda Adit in gold mine in Złoty Stok, Lower Silesia, Poland, 16 October 2006, coll. A. Chlebicki. First culture obtained on DRBC medium, then transferred to other media.

DISCuSSIoN
So far two species of the genus Cephalotrichum Link were noted in Poland. Dominik and Majchrowicz (1965) and Dominik (1970)  on the basis of specimen isolated from the soil. Conidia of this specimen are something smaller than typical variety, 4-6 x 2.5-4 μm and 'very slightly rough' (Dominik 1970). It resembles Cephalotrichum microsporum (Sacc.) P. M. Kirk. Moreover these authors did not mention the presence of Echinobotryum synanamorph which is diagnostic character of Cephalotrichum stemonitis. the next species Cephalotrichum putredinis (Corda) S. P. Abbott was reported by Dominik (1970) as Doratomyces albus (Szilvinyi) Dominik. unfortunately, these collections are not available for investigation.
Presence of Cephalotrichum stemonitis in bacterial biofilm is accidental. However, as indicate informations of Fassatiova (1970) and Malloch and Hubart (1987), such fungi were noted in subterranean environments. Mille-Lindblom (2005), Hogan and Kolter (2002) and Kirkwood (2002) noted mostly antagonistic relation between fungi and bacteria. Fungi were always negatively affected by presence of bacteria. Penetration of fungal hyphae of Fusarium oxysporum was not observed where microcolonies of Pseudomonas were present, moreover Pseudomonas bacteria attached and colonized fungus hyphae (Bolwerk et al. 2003). Also growth of Cephalotrichum stemonitis was suppressed by bacteria from the genus Pseudomonas isolated from biofilm.