The lichen genus Pertusaria in Poland I. P . multipuncta and P . ophthalmiza

The lichen . (2): 231–238, 2010. The paper presents results of study on two morphologically very similar lichens in Poland , P. multipuncta (Turner) Nyl. and P. ophthalmiza (Nyl.) Nyl. (Pertusariales, Ascomycota). Previously, specimens were determined using only thallus characters and spot test reaction, with no data on lichen substances. This led to several misidentifications. After the revision of all available material of P . multipuncta from Poland it appeared to be reported correctly only from one locality in Gorce Mts. Most of other specimens belong to P . ophthalmiza , which has not been recorded in Polish lichenological literature until 2008.

thin layer chromatography (TLC) in the country. Previously, specimens were determined using only thallus characters and spot test reaction, and very rarely the ascus type was studied. This could have led to many misidentifications.
Based on morphology and TLC-data, the present paper focuses on two taxa, namely P. multipuncta (Turner) Nyl. and P. ophthalmiza (Nyl.) Nyl., which are very similar in morphology, but have different chemistry. Both produce ascomata, which are covered with soredia-like granules, and thus resemble soralia. P. multipuncta was reported in Poland more often than P. ophthalmiza, as it could have been easily determined with the aid of the key by Nowak and Tobolewski (1975). Until recently P. ophthalmiza has been reported from Poland only by , who cited single, old collection. That record was not included in the recent Polish checklist of lichens and lichenicolous fungi (see Fałtynowicz 2003), and its presence in the country has remained unnoticed until the paper by Kukwa et al. (2008). These authors reported also new Polish localities of the species.
As preliminary study suggested that the name P. multipuncta might have been misapplied in Poland for the material of P. ophthalmiza, we decided to revise all available specimens of both taxa. The aim of this paper is to present the results of study on the chemistry, morphology, habitat requirements and distribution of P. multipuncta and P. ophthalmiza in Poland. This paper is the first of a series of articles devoted to a revision of Pertusaria in Poland.

MATErIAL AND METHODS
The present study is based on the specimens deposited in the Polish lichen herbaria: GPN, KTC, KrAM, LBL, LOD and UGDA. The morphology of the specimens was studied using a stereomicroscope. The thickness, morphology and colour of thallus, colour, shape and size of apothecia were noted. Secondary metabolites were identified by thin layer chromatography (TLC) according to the methods of Orange et al. (2001). The chromatograms were developed in solvent C. Localities of all Polish material examined are mapped according to the ATPOL grid square system (Zając 1978;modified by Cieśliński and Fałtynowicz 1993; see also Kukwa et al. 2002).

CHArACTErISTIC OF THE GENUS PerTusarIa
The genus Pertusaria (Pertusariales, Ascomycota; see Schmitt et al. 2006) includes corticolous, saxicolous, and terricolous crustose taxa, most of which do not grow on pure limestone or other strongly calcareous substrata (Fałtynowicz 2003;Chambers et al. 2009). Many species occur in dry, open habitats, but some prefer humid localities (Nowak, Tobolewski 1975;Fałtynowicz 2003;Chambers et al. 2009). The genus is cosmopolitan and its members are found from the Arctic and Antarctic to the tropics in both Hemispheres (e.g., Dibben 1980;Archer 1997;Chambers et al. 2009). Nevertheless, there are still substantial gaps in the distribution of many taxa in various regions of the world. In many cases, published data should be treated with caution, since the determination of species is often based mainly on morphological characters, but not secondary chemistry, which is an important systematic character set (e.g., Dibben 1980;Archer 1997;Chambers et al. 2009).
the taxonomy of Pertusaria and the delimitation from other phylogeneticaly related genera are still not satisfactorily settled. Traditionally, members of the genus Pertusaria are characterized by rather large (sometimes up to 4 mm in diameter) apothecia with hemiangiocarpous type of development, with open or almost closed, perithecia-like disk, hamathecium of branched and richly anastomosing, lax paraphysoids, thick-walled, amyloid asci with 1-8 spores, and thick spore wall. Thallus is crustose, moderately thick or ± immersed, continuous to rimose-cracked, fissuredareolate or warted. In many species soralia and/or isidia are produced. In some taxa apothecial disk can be occluded with granular soredia and appearing soralium-like. Secondary lichen metabolites are very diverse in Pertusaria, and these can be divided into four chemosyndromes, each consisting of either xanthones, fatty acids, depsides or depsidones (e.g., Dibben 1980;Archer 1997;Chambers et al. 2009).
After the recent molecular studies by Schmitt and Lumbsch (2004), Pertusaria appeared as polyphyletic in its current circumscription and can be divided into three well supported groups, Pertusaria s.str.-group, Varicellaria-group and Variolariagroup. The first one is characterized by amyloid, 2-8-spored asci with distinctive ocular chamber, non-amyloid hymenial gel, thick or thin walled spore wall, and the presence of chlorinated xanthones, gyrophoric and planaic acids. Apothecia can be open with plane disc or closed, and then resembling perithecia. Members of the Varicellaria-group has strongly amyloid, 1-2-spored asci with no recognizable apex structures, strongly amyloid hymenial gel, ± thick spore wall, disciform apothecia, production of lecanoric acid and absence of chlorinated xanthones. At present 3 taxa of Pertusaria are known to belong in this group, P. hemisphaerica (Flörke) Erichsen, P. lactea (L.) Arnold and P. velata (Turner) Nyl., and they are close to the monotypic genus Varicellaria Nyl. (with V. rhodocarpa (Körb.) Th. Fr.). The third, Variolariagroup, is characterized by strongly amyloid, 1-spored asci with no recognizable apex structures, weakly or not amyloid hymenial gel, thin spore wall, open apothecia, production of picrolichenic and thamnolic acids, and absence of chlorinated xanthones (Schmitt and Lumbsch 2004). However, since that time no taxonomical segregation has been proposed yet, therefore all taxa are still kept in Pertusaria.
Syn. Variolaria multipuncta Turner, Trans. Linn. Soc. London 9: 137 (1806) CharaCteristiC of the speCies. P. multipuncta is distinguished from superficially similar species by the white, well-delimited, scattered, sorediate warts containing apothecia, 1-spored asci and the chemistry (physodalic acid and often protocetraric acid) Chambers et al. 2009). Thallus of the species can be very variable; on sheltered, smooth bark it is often thin, continuous and even, but in some exposed habitats it is more robust and the surface is coarsely warted (Chambers et al. 2009). Thallus and soralia of P. multipuncta are K+ yellow, C-, KC+ yellow, PD+ orange-red, UV-or faintly glaucous (Chambers et al. 2009). In Polish specimen physodalic and protocetraric acids were detected.
affinities. in europe P. multipuncta can be mistaken for the morphologically similar P. ophthalmiza. Both species produce similar soralium-like apothecia, but they can be easily separated by spot test reaction PD and thin layer chromatography: P. multipuncta reacts PD+ orange-red and contains physodalic acid, often together with protocetraric acid, and P. ophthalmiza is PD-and has fatty acids Chambers et al. 2009). They differ also morphologically. P. multipuncta has fertile thallus warts measuring 0.5-1.5 mm diam., wide apothecial disc, and continuous and ± even exciple; in P. ophthalmiza the fertile warts are smaller (0.3-0.8 mm diam.), disc is up to twice narrower, and the exciple irregular and crenate (Chambers et al. 2009).
P. albescens and P. amara can also resemble P. multipuncta in morphology, but they are usually sterile, soredia are coarse (very often forming consoredia) and produced in true soralia; they posses also different lichen substances: P. albescens contains fatty acids, whereas P. amara picrolichenic acid, often accompanied by protocetraric acid (Tønsberg 1992;Chambers et al. 2009).
habitat. P. multipuncta is a typical epiphytic lichen species, but it can rarely grow also on siliceous rocks (Chambers et al. 2009). In Poland it was found on bark of Fagus sylvatica in beech forest.
Distribution. in Poland P. multipuncta was reported from northern and southern part of Poland, but rather scarcely (e.g., Nowak and Tobolewski 1975;Fałtynowicz 2003 and literature cited therein; Motiejūnaitė et al. 2004). After the revision of all available material from Poland it appeared that most records were based on misidentifications. Only one specimen from Gorce Mts appeared to belong to this species. Most of other specimens belong to P. ophthalmiza, which has not been recorded in Polish lichenological literature until 2008 (see Kukwa et al. 2008, also below). Some few additional specimens identified as P. multipuncta were rather small and difficult to identify, however they certainly do not belong either to P. multipuncta or P. ophthalmiza. P. multipuncta species was reported from many countries, however, as this name was often misapplied in the past (see , several records are doubtful. Confirmed records of P. multipuncta appears to be confined to western and southern Europe, where it has been so far reported from Belgium, British Isles, Denmark, France, Germany, Italy, Luxembourg, Slovenia, Spain, Sweden Suppan et al. 2000;Søchting and Alstrup 2002;Nimis and Martellos 2008;Diederich et al. 2009). According to Chambers et al. (2009) it occurs also in Macaronesia, Africa and Asia.    CharaCteristiC of the speCies. This species is anatomically and morphologically very similar to P. multipuncta, but it can be distinguished predominantly by the different chemical constituents; for the differences see under P. multipuncta. According to  and Chambers et al. (2009), P. ophthalmiza contains 4 fatty acids, but in most Polish specimens 2-3 fatty acids were found. Only in three specimens 4 substances were found, but also 1 was detected in two samples. This variation is most probably caused by rather low quantity of material taken for the analyses, as most specimens were quite small.
affinities. P. ophthalmiza can be mistaken for chemically and, when young, also morphology similar P. albescens. Both species produce fatty acids, but they predominantly differ in thallus characters: P. albescens produces true soralia with granular soredia, whereas the sorediate warts of P. ophthalmiza contain apothecia Tønsberg 1992;Chambers et al. 2009).
Superficially P. ophthalmiza can resemble also Ochrolechia turneri (Sm.) Hasselrot. This species often produces regular and separate soralia, which may resemble fertile warts of P. ophthalmiza. Both taxa can be very easily separated by the C reaction of soralia and content of lichen substances. P. ophthalmiza is C negative and produces fatty acids, whereas soralia of O. turneri react C+ yellow and the species contains variolaric acid Tønsberg 1992;Kukwa 2008).
habitat. research on Polish material indicated that P. ophthalmiza is a corticolous species. It has been rather frequently found on Tilia cordata (13 specimens). Much less commonly it was found on alnus spp. (4 specimens), Carpinus betulus (2 specimen) and Fraxinus (1 specimen), always in old, humid woodland. According to Chambers et al. (2009) this taxon can occur on acid bark (chiefly birches, oaks and coniferous trees) and overgrow epiphytic mosses, but in Poland it appears to be strictly corticolous on deciduous trees with more neutral bark.

cOnclusiOns
The study has shown, that the secondary lichen metabolites are invaluable taxo-• nomic characters, that helps to evaluate the occurrence and distribution of at least some lichens. Pertusaria multipuncta • and P. ophthalmiza are morphologically very similar and visually almost indistinguishable, but they differ in secondary chemistry. The analyses of lichen substances proved that P. multipuncta was mostly misidentified, and at present it is known from only one locality in Gorce Mts; most of remaining records of this species belong to P. ophthalmiza. Fig. 1. Distribution of Pertusaria multipuncta and P. ophthalmiza in Poland in the atPOl grid square system; empty circle -locality of P. multipuncta, black circles -localities of P. ophthalmiza based on revised specimens; square -historical record of P. ophthalmiza from . P. ophthalmiza • appears to occur in well preserved natural ecosystems in North Eastern Poland and Tatra Mts. Surprisingly, it was not found in Bieszczady Mts, the mountain range with suitable habitats for this species.