Ophiocordyceps stylophora (Ophiocordycipitaceae) in Poland: new localities and host species

Ophiocordyceps stylophora, a rare fungus species in Europe, was first recorded in one locality in Poland in 2011. This paper concerns 33 specimens found between 2011 and 2015 in 23 new localities in five areas. Amongst the described specimens, one was found on the Tenebrionidae beetle, and others grew on Elateridae larvae. For the first time, Tenebrionidae larvae have been observed as hosts of O. stylophora. Five new positions have been located in the southern part of Poland in the area of Oszast and Reberce nature reserves, Babia Góra and Roztocze national parks, and one has been found in the northern part of Poland, in the Elbląg Upland Landscape Park.


Introduction
Ophiocordyceps stylophora (Berk. & Broome) G. H. Sung et al. is a member of Hypocreales (Ascomycota). It was described in 1857, and classified within genus Cordyceps s. l. In 2007, it was transferred to the genus Ophiocordyceps [1,2]. This group of fungi exhibits parasitic tendencies [1]. In most cases, they parasitize Coleoptera larvae, but some species have been recorded on insects in the Lepidoptera, Heteroptera, Homoptera, Hymenoptera, and Araneae orders, as well as on fruit bodies of the hypogeous fungi from the genus Elaphomyces [1]. In Poland, 16 species of Cordyceps s. l. have already been recorded [3][4][5].
Ophiocordyceps stylophora is an entomopathogen that parasitizes beetle larvae from Cerambycidae, Elateridae, Scarabaeidae, and Chrysomelidae families [1]. The hosts are slightly recessed in wood (up to a few centimeters) when fungal structures are developed and visible on the wood surface. It is a rare species found in few countries in the world [3,6]. Recently, it has been observed in several places in Europe including Latvia [7], Norway [8], and Slovakia [9][10][11] (Fig. 1). Until now, recorded observations of this species in Poland have been limited to the Pieniny National Park [3]. Further fieldwork conducted between 2011 and 2015 revealed several other locations throughout Poland (Fig. 1).
The aim of the research is to present new localities of O. stylophora in Poland with reports on their hosts and description of its habitats. The ecology of the species is also briefly discussed.  [12] and Mains [13]. Preparations of dry specimens mounted in water, Melzer's reagent, and Congo red were microscopically observed using the PZO 14 light microscope with Nomarski interference contrast [14]. Macro-and microfeatures of the specimens did not differ significantly from the specimens previously collected in the Pieniny National Park [3]. The Latin names of liverworts are derived from Szweykowski [15], those of mosses from Ochyra et al. [16], those of vascular plants from Mirek et al. [17], and those of plant communities from Matuszkiewicz [18,19]. Names and numbers of geographical regions were adopted from Kondracki [20]. Site localizations were marked using GPS Garmin III+. The localities were assigned to 10 × 10-km ATPOL squares calculated according to Snowarski's calculator [21]. Herbarium materials were deposited in P. Chachuła's private collection.

Discussion
The localities presented in this paper are upland, foothill, and mountain regions with elevations ranging from 100 to 1,120 m a.s.l. It should be emphasized that out of the 15 studied regions, O. stylophora was found in five. Of these, localities ( Fig. 1) are located in areas under strict protection, excluded from forest management activities. They are in the Reberce and Oszast nature reserves (Fig. 1) and the Babia Góra National Park (Fig. 1). One site in the Roztocze National Park is located in an area of active protection. The site located in Elbląg Upland Landscape Park (Fig. 1) is managed by the State Forests National Forest Holding. In this area, suitable conditions for the development of O. stylophora were created by leaving dead wood in inaccessible places, such as the deep valleys of the streams. Among six areas in 23 localities, 33 specimens of O. stylophora were observed, including 30 in the teleomorphic stage and three in the anamorphic stage. This fungus is most commonly found on click beetle larvae from the Cerambycidae, Elateridae, Scarabaeidae, and Chrysomelidae families [1][2][3]7,8,[10][11][12][13]. A similar pattern was observed in this study of the 33 specimens, the hosts for the fungus in 29 cases were larvae from the Elateridae family (in the case of three specimens from Oszast the hosts were not identified). The exception was the fungi specimen observed in the Kumiel (Dzikuska) Stream in Elbląg (Fig. 1), where the stroma of O. stylophora emerged from a larva of the Tenebrionidae family. The insects from this family were never mentioned in literature as hosts for O. stylophora and our finding extends the list of potential hosts for this species [1].
In reports on O. stylophora, the authors rarely described the habitat of this species precisely [3,7]. Generally, it can be said that vegetation in which this species is found is diverse. In Elbląg Upland and Roztocze National Park, specimens were found in the riparian forest in valleys of streams, similar to regions in Latvia and Norway [7,8]. In higher-elevation areas of the Reberce nature reserve and Babia Góra National Park, this fungus grew in old beech and fir, such as those in the Pieniny National Park [3]. In Slovakia, they were found in beech forests [10,11] while in Mexico, they were found in oak and pine forests [6].Although most observations from Europe [7][8][9][10][11]22] have been done in October and one in April [3], the observations described in this paper indicate that the anamorphic and teleomorphic stages of Ophiocordyceps stylophora can be found year-round. Moreover, unlike Mains' conclusions [13,23], the results demonstrate that the mature stage of O. stylophora can also be observed at nearly anytime in the year (February-June, September-November) (Tab. 1).
The substrates in which beetle larvae were observed were coniferous and deciduous wood. The following tree species are mentioned in previous studies: Alnus glutinosa [7,8], Fagus sylvatica [10,11], and Abies alba [3]. In this respect, the following correlation was observed in Poland: in the natural ranges of Abies alba, O. stylophora was found in fir logs, while beyond the area of the fir range, larvae infected by this fungus were found in deciduous wood.
The observation of O. stylophora, a notably rare taxon, and the recent discovery of new Cordyceps s. l. species from Poland [4] prove that the fungal species in Poland have not been comprehensively investigated.