Taxonomic recognition of some species-level lineages circumscribed in nominal Rhizoplaca subdiscrepans s. lat. (Lecanoraceae, Ascomycota)

Taxon sampling

Material used in this study originated from the following herbaria: ASU, CANL (CMN), KRAM, M, MIN, PRA, WRSL and the private herbarium of K. Szczepańska (hb. Szczepańska). Our sampling focused on genera of lecanoroid lichens with placodioid thalli as follows: Lecanora (L. pseudomellea), Protoparmeliopsis (P. garovaglii, P. muralis) and Rhizoplaca (R. chrysoleuca, R. opiniconensis, R. phaedrophthalma, R. subdiscrepans); holotypes of Lecanora pseudomellea, Rhizoplaca opiniconensis and R. phaedrophthalma were also analysed. Unfortunately, we were unable to study the original collection of R. subdiscrepans housed at the PC herbarium since type material at PC is unavailable for loan and we had no opportunity to visit the herbarium. All of the specimens listed in the text were included in the morphological, anatomical and chemical studies, but due to the age and poor condition of some samples, it was not possible to obtain sequences and include these in our phylogenetic analyses. New sequences generated for this study have been deposited in GenBank.

Morphological and chemical study

All specimens were examined using light microscopy for the assessment of lobes, areoles, squamules and apothecia, especially the morphological characters mentioned in Leavitt et al. (2011) such as point of attachment (umbilicate/squamulose), thallus form (polyphyllous/monophyllous), lobe morphology (distinct/indistinct), appearance of upper surface (dull/glossy), upper surface colour (yellow-green/olive), apothecia morphology (sessile/constricted), apothecia pruinosity (pruinose/epruinose), ascospore shape (ellipsoid/subglobose) and ascospore size. Apothecial margin (persistent/excluded), height of hymenium, and size and shape of conidia (straight/arched) were also analysed.

For light microscopy, vertical, free-hand sections of apothecia were cut by a razor blade and mounted in water. Hymenium measurements were made in water and ascospore measurements in 10% KOH (K); the structure and coherency of paraphyses and the solubility of granules in the epihymenium were also tested with K. At least 10 measurements of morphological variables and measurements of 20 spores were made for each sample and their minimum and maximum values calculated.

Chemical examinations included colour reactions and thin-layer chromatography (TLC). Spot test reactions of thalli, apothecial margins and discs were made with KOH, sodium hypochlorite [commercial laundry bleach] (C) and paraphenylenediamine [solution in 95% ethyl alcohol] (PD). TLC analyses were performed in solvent systems A, B and C using the standardized method of Culberson (1972) and following Orange, James & White (2001).

Descriptions of the species are based on our own observations, measurements and TLC analyses of specimens cited in this paper. The terminology used in the descriptions of the species follows Ryan et al. (2004).

Molecular methods: DNA extraction, PCR amplification and DNA sequencing

To infer relationships between species of lichenized fungi studied, the ITS rDNA region, that contains ITS1, 5.8S, ITS2 sequences, was used. Total DNA was extracted from specimens using a CTAB method (Doyle & Doyle, 1987). Dried samples were mechanically disintegrated using Mixer Mill MM400 (Retsch; Haan, Germany). The quality of the DNA was checked by electrophoresis on agarose gel (1%) with Simply Safe staining chemical (Eurx, Gdańsk, Poland). The complete ITS rDNA region was amplified using primers ITS1F (Gardes & Bruns, 1993) and ITS4 (White et al., 1990). The PCR reaction mix included 1U Taq recombinant polymerase (Thermo-Fisher Scientific, Waltham, USA), 10x Taq Buffer, 1 mM MgCl₂, 0.5 M of each primer, 0.4 mM dNTP and 1 µl DNA template. Amplification cycles were performed with a Veriti Thermal Cycler (Life Technologies, Carlsbad, USA) and involved 8 min at 95 °C, followed by 32 cycles for 45 s at 95 °C and 45 s at 52 °C (annealing), and 1 min at 72 °C, with the a final extension step of 10 min at 72 °C. Amplified PCR products were purified using GeneJet PCR Purification Kit (Thermo-Fisher Scientific, Waltham, USA). This was accomplished at the Laboratory of Molecular Biology (Environmental and Life Sciences) at Wroclaw University. Sequencing of PCR products, post-reaction purification, forward and reverse directions reading were done by sequencing service Genomed (Genomed, Warsaw, Poland), using an ABI 377XL Automated DNA Sequencer (Applied Biosystems, Carlsbad, USA).

Alignment assembly and molecular phylogenetic analyses

The obtained ITS rDNA sequences were assembled and manually edited using Geneious Pro, version 8.0. (Biomatters Ltd). Sequences of Rhizoplaca subdiscrepans (putative name for our collection) together with related species, namely Lecanora pseudomellea, Protoparmeliopsis garovaglii, P. muralis, Rhizoplaca chrysoleuca, R. melanophthalma, R. novomexicana, R. occulta, R. opiniconensis (as Lecanora in GeneBank), R. parilis and R. phaedrophthalma, were included in the analysis. Our final ITS data-set included 21 sequences newly generated for this study and 95 sequences downloaded from GenBank (Table 1). The final alignment was performed on Geneious Pro using the MAFFT algorithm (Katoh et al., 2005), then re-checked and improved. Ambiguously aligned regions were removed in GBlocks (Castresana, 2000). The nucleotide substitution models were separately searched for in each partition of the ITS region (ITS1, 5.8S, ITS2) to find the best-fitting model using the corrected Akaike information criterion (AICc) as an optimality model criterion in a greedy algorithm search as implemented in PartitionFinder version 1.0.1 (Lanfear et al., 2012). The GTR+G model for ITS1 and ITS2 and K80 for the 5.8S partitions were selected.

The phylogenetic reconstruction was generated using the CIPRES Scientific Gateway (http://www.phylo.org/portal2/) (Miller, Pfeiffer & Schwartz, 2010). Maximum likelihood (ML) bootstrap tree with simultaneous heuristic search was undertaken as implemented in RAxML–HPC2 on XSEDE (Stamatakis, 2006) under the GTRGAMMA substitution model and 1,000 bootstrap resamples. Bayesian inference was carried out using Markov Chain Monte Carlo (MCMC) implemented in MrBayes 3.2.6 on XSEDE (Ronquist et al., 2012). MrBayes was set to two independent parallel runs each initiated with four incrementally heated chains; the run length was settled to 20M generations, and to infer convergence the average standard deviation of split frequencies was printed every 1000th generation discarding the first 50% of the trees sampled as a burn-in fraction. The analyses were stopped after 1M generations when the standard deviation had dropped below 0.01. The resulting phylogenetic trees were visualized in Figtree software (Rambaut, 2014). The alignment of ITS sequences described here is deposited in TreeBASE with number TB2: S26365.

Taxonomy

Rhizoplaca opiniconensis (Brodo) Leavitt, Zhao Xin & Lumbsch, in Zhao et al., Fungal Diversity 78: 302. 2016. ≡ Lecanora opiniconensis Brodo, Mycotaxon 26: 309. 1986.

Figs. 2A, 2B.

Type: [CANADA], ONTARIO: Leeds Co., Snake Island, in Lake Opinicon, Queen’s University Biological Station, Chaffeys Locks, on S-facing rockface, 1.0–1.5 m above water, 2 Feb. 1985, I.M. Brodo 25117 (holotype, CANL!).

Thallus lichenized, placodioid, polyphyllous, usually distinctly rounded, irregular when older, thick, loosely attached to the substrate, 2–5 cm diam. or more. Marginal lobes distinctly developed, short, 0.5–1 mm long and 0.2–0.6 mm wide, plane to slightly convex, broadened and crenate-incised at the tips. Thallus centre squamulose-areolate to squamulose-bullate, irregularly cracked, thick, loosely attached to the substrate. Squamules crowded, flat to strongly convex, irregular, 0.2–1.5 mm diam. Upper surface smooth, dull, yellowish-green to yellowish-orange (more evident in herbarium material), marginal lobes usually darker than the thallus centre. Apothecia numerous, sessile to constricted, 0.2–2 mm diam. circular, older apothecia irregular and distorted. Margin thin, concolourous with thallus or paler, flexuose, crenate, usually persistent, sometimes excluded, disc pale yellowish-brown to yellowish-orange, epruinose, plane, rarely convex, dull. Hymenium colourless, 60–70 μm high, hypothecium colourless. Epihymenium orange-brown to olive-brown, with small granules dissolving in K. Ascospores 8 per ascus, hyaline, simple, ellipsoid, 10–12 × 6–7 μm. Pycnidia immersed, simple with bluish-black ostioles, conidia hyaline, filiform, mostly arched 15–25 × 0.5 μm.

Chemistry. Thallus K–, C–, KC+ yellow, PD–, medulla K+ yellow, C–, KC+ yellow, PD–. Secondary metabolites detected by TLC: isousnic (+∕ −), usnic (+) and placodiolic (+) acids, as well as some unidentified terpenoids (+∕ −).

Ecology and distribution. On quartzite and volcanic rocks, at higher elevations (c. 1,200–3,000 m), in shaded places near streams, in North America and Eastern Asia.

Comments. Characteristic features of R. opiniconensis are the mostly arched conidia and thallus colour, which changes to a more orange colour in herbarium specimens. Some specimens are morphologically similar to R. subdiscrepans s. str., in which case proper identification of both taxa may be difficult. However, in contrast to R. subdiscrepans s. str., R. opiniconensis does not produce fatty acids and was genetically confirmed in North American and Asian but not from European specimens. Additionally, the species has also specific habitat preferences.

Specimens examined. CANADA. ONTARIO: West edge of Marathon, shore of Lake Superior and back on rock ridges with black spruce and scattered quaking aspen, alt. 629 ft, 5 Aug. 2002, C.M. Wetmore 88134 (MIN). MEXICO. MEXICO: Baja California Sur, below top of Sierra Agua Verde (part of the Sierra San Francisco), on rhyolite, N-facing cliff, alt. c. 1300 m, 1 Jan. 1998, T.H. Nash III 40100 (ASU); Chihuahua, along secondary dirt road to Casa Grandes from Bavispe, Sonora, oak woodland area with large outcrops of rhyolite, on rhyolite, alt. c. 2050 m, 18 July 1994, T.H Nash III 36458 (ASU). USA. ARIZONA: Apache Co., Apache National Forest, Mt Baldy Wilderness area, trail from Phelp’s cabin along East Fork of Little Colorado River, on rock, alt. 3001 m, 1 July 1990, T.H Nash III 27051 (ASU); Apache Co., Petrified Forest National Park, south of I-40, east of Petrified Forest Rd, near Crystal Forest Trail, on sandstone, alt. 1669 m, 14 May 1990, W.C. Davis 735 (ASU); Cochise Co., Chiricahua Mountains, Colorado National Forest, cliff area above Mormon Springs in Mormon Canyon, oak woodland, on exposed cliff-face, alt. c. 2070 m, 24 Nov. 1995, T.H. Nash III 37156 (ASU); Cochise Co., Chiricahua Mountains, Chiricahua National Monument, along the Loop Trail above Rhyolite Canyon and below “Heart of the Rocks”, oak-pine forest, on rhyolite, alt. c. 2000 m, 5 June 1998, T.H. Nash III 41767 (ASU); Cochise Co., Chiricahua Mountains, Chiricahua National Monument, just before junction of trail to Heart of Rocks and trail to Massai Point, steep NE-facing part of rhyolite boulders under tall rock formation, alt. c. 2020 m, 5 June 1993, B.D. Ryan 30861 (ASU); Graham Co., E side of the Pinaleno Mountains, along trail above Shannon Campground, mixed conifer-oak forest, on acidic rock, alt. c. 2000 m, 15 July 1994, T.H Nash III 36056 (ASU); Pima Co., east side of Baboquivari Peak, on exposed granite, alt. c. 1920 m, 13 Oct. 1990, T.H. Nash III 27444 (ASU); Pima Co., Santa Catalina Mountains, Coronado National Forest, upper part of Sabino Canyon, near Marshall Gulch, granite boulders in mixed conifer forest, on granite, alt. c. 2310 m, 3 June 1998, T.H. Nash III 41709 (ASU). MINNESOTA: Cook Co., Grand Portage State Forest, Lake Superior shore NE of Horseshoe Bay (16 miles SW of Grand Portage), shrubby wet cobble shore, alt. 603 ft, 28 Sept. 2001, C. Reschke 1567 (MIN); Cook Co., Grand Portage National Monument, north side of Mt Rose, rock cliffs and large rock blocks below with some white birch and white spruce, alt. 867 ft, 13 July 2012, C.M. Wetmore 100545 (MIN); Pope Co., Glacial Lakes State Park, 5 miles S of Starbuck, Prairie area with border woods on esker 0.1 mile NE of Mountain Lake, 7 Aug. 1993, J.P. Schuster 3098 (ASU); Lake of the Woods Co., Clementson (8 miles east of Baudette), at mouth of Rapid River to Rainy River, on hillside with bur oak, ash, white birch and balsam poplar, also on rocks above rapids, 18 Aug. 1994, C.M. Wetmore 74873 (MIN); Otter Tail Co., Inspiration Peak State Scenic Wayside Park, 12 miles west of Parkers Prairie, on hill with grass and rock on top and on hillside with red oak and bur oak, 14 Aug. 1994, C.M. Wetmore 74475 (MIN); Washington Co., Lost Valley Prairie Scientific and Natural Area, prairie area with limestone outcroppings and cedar trees, surrounded by aspen and mixed hardwoods, 3 miles NNW of junction of St Croix and Mississippi Rivers, 3 miles NNE of Hastings, 5 Aug. 1990, J.P. Schuster 2460 (KRAM). NEW MEXICO: Cibola Co, Cibola National Forest, Zuni Canyon 0.2 miles SW of Log Chute (8.5 miles SW of Grants), small N-S canyon with rocks, douglas fir, oak and few ponderosa pine, alt. 7434 ft, 17 June 2010, C.M. Wetmore 99445 (MIN); Cibola Co., El Malpais National Conservation Area above Ranger Station (12.8 miles S of Grants), ridge S of ranger station with juniper, sage and rocks, alt. 6837 ft, 15 June 2010, C.M. Wetmore 99205 (KRAM); Cibola Co, El Malpais National Conservation Area, 9 miles S of Grants, lava beds with scattered juniper, alt. 6553 ft, 15 June 2010, C.M. Wetmore 99234 (MIN); McKinley Co., Cibola National Forest, 0.5 mile E of Quaking Aspen campground on USFS 162 (15 miles SE of Gallup), valley with rock ledges, ponderosa pine, juniper and oak, alt. 7735 ft, 18 June 2010, C.M. Wetmore 99527 (KRAM); San Miguel Co., near town of Villanueva (42 miles SW of Santa Fe), hillside and ridge near dam with juniper and pinyon pine, alt. 5900 ft, 11 June 2010, C.M. Wetmore 98925 (KRAM).

Rhizoplaca phaedrophthalma (Poelt) Leavitt, Zhao Xin & Lumbsch, in Zhao et al., Fungal Diversity 78: 302. 2016 ≡ Lecanora phaedrophthalma Poelt, Mitt. Bot. Staatss. München 2: 483. 1958.

Figs. 2C, 2D.

Type: [NEPAL], NÖRDL. ZENTRALNEPAL: Manangbhot (oberes Marsyandi –Tal) Felsen nördlich Banphag, alt. 4100 m, 13 June 1955, Fr. Lobbichler (holotype, M 122–86/40!).

Thallus lichenized, placodioid, polyphyllous, rounded to irregular when older, 2–5 cm diam. Marginal lobes usually weakly developed, 0.5–1.2 mm long and 0.2–0.7 mm wide, short, plane to slightly convex, broadened at the tips and flabellate. Thallus centre squamulose-areolate to squamulose-bullate, irregularly cracked, thick, loosely attached to the substrate. Squamules crowded, flat to slightly convex, irregular, 0.5–2 mm diam., usually with bluish-black tinges on the margins and underside towards edges. Upper surface smooth, dull, pale and light yellow, pruinose in parts, marginal lobes concolourous with thallus centre. Apothecia numerous, sessile, dispersed to grouped mainly in the centre, 0.5–1.2 mm diam., circular, older irregular. Margin thin, smooth, concolourous with thallus, slightly raised when young, then excluded, disc orange-brown to reddish-brown, usually contrasting with the thallus, epruinose, strongly convex, dull. Hymenium colourless 50–60 μm high, hypothecium colourless. Epihymenium orange-brown, with small granules dissolving in K. Ascospores 8 per ascus, hyaline, simple, subglobose, 8–10 × 5–7 μm. Pycnidia immersed, simple with bluish-black ostioles, conidia hyaline, filiform, mostly straight, 15–30 × 0.5 μm.

Comments. The main characteristic feature of R. phaedrophthalma is the morphology of apothecia, which are strongly convex, with reddish-brown, epruinose discs and excluded margin. Additionally, mature specimens of the species, in contrast to R. opiniconensis and R. subdiscrepans s. str., are usually poorly placodioid, with weakly define marginal lobes. R. phaedrophthalma seems to be also characterized by the size of its conidia, which are slightly larger than in other species treated, as well as by subglobose rather than ellipsoid spores. Concerning secondary metabolites produced by the species, neither isousnic acid nor fatty acids were seen in TLC, unlike from the thalli of other taxa.

Specimens examined. CANADA. BRITISH COLUMBIA: Buse Hill, on exposed rock near top of hill, alt. c. 950 m, 27 Aug. 1994, B.D. Ryan 31877 (ASU); near Quilchena Hotel, alt. c. 625 m, 28 Aug. 1994, B.D. Ryan 31889 (ASU). RUSSIA. ORENBURG REGION: Surroundings of water reservoir “Iriklinskoe vodokhranilishche”, Iriklinskiy, vill. Vishevoe, rocks 2 km E of village, in valley of stream, alt. 250–270 m, on acidic schist, 10 June 2011, J. Vondrák 9406 & Frolov (PRA, two samples). USA. COLORADO: Park Co., Pennsylvania Mt, Pike National Forest, on rock, c. 3688 m, 1 June 1990, M.A. Thomas 32870 (ASU). IDAHO: Twin Falls Co., Hagerman Fossil Beds National Monument, 0.5–1 km E of S end of 500 East Rd, on basalt, alt. c. 900 m, 10 Sept 1998, B.D. Ryan 32870 (ASU). MONTANA: Gallatin Co., Yellowstone National Park, below Black Butte along highway 191 near NW corner of park, on talus slope and along highway, alt. 6700 ft, 15 July 1998, C.M. Wetmore 80571 (MIN); Park Co., Yellowstone National Park, Grazing enclosure 1 mile W of Garsiner at northern edge of park. Open grassland on knoll with sagebrush and rock outcrop, alt. 5300 ft, 21 July 1998, C.M. Wetmore 80979 (MIN). OREGON: Deschutes Co., 6 miles W of Redmond, Juniper and sage area with lots of loose soil, alt. 3020 ft, 18 July 2006, C.M. Wetmore 95059 (MIN); Malheur Co., along US20 above Malheur River 13 miles SW of Harper, Sage brush prairie on north slope, alt. 2790 ft, 19 July 2006, C.M. Wetmore 5101 (MIN). WYOMING: Park Co., West and of Lamar Canyon, 6 miles E of Tower Junction, on south facing hillside with large granitic glacial erratics and scattered douglas fir, alt. 6500 ft, 19 July 2006, C.M. Wetmore 81235 (MIN); Park Co., Yellowstone National Park. Pebble Creek Trail at rocks above cliffs, below switchback, 0.25 miles up from camp-ground, alt. 6900 ft, 27 July 1998, C.M. Wetmore 81446 (MIN); Park Co., Yellowstone National Park, Sheepeater Cliffs, 5.5 miles S of Mammoth, on south-facing columnar cliffs and talus above stream, alt. 7200 ft, 20 July 1998, C.M. Wetmore 80866 (MIN).

Rhizoplaca subdiscrepans (Nyl.) R. Sant., The Lichens of Sweden and Norway: 278. 1984. ≡ Squamaria chrysoleuca var. subdiscrepans Nyl., Flora 44: 718. 1861. ≡ Lecanora subdiscrepans (Nyl.) Stizenb., Berichtüber die Tätigkeit der St. Gallischen Naturwissenschaftlichen Gesellschaft 1880-1881: 341. 1882.

Figs. 2G, 2H.

Type: [EUROPE], Helvetiae et Tyroliae (PC).

Thallus lichenized, placodioid, polyphyllous, usually rounded, thick, convex, loosely attached to the substrate, 1–5 cm or more in diam. Marginal lobes distinctly developed, short, 1–2.5 mm long and 0.5–1.5 mm wide, discrete or contiguous to slightly overlapping, convex, broadened and crenate-incised at the tips. Thallus centre squamulose-bullate, composed of crowded strongly convex subunits. Squamules rounded, sinusoid and plicate, 0.25–1 mm diam. Upper surface smooth, glossy, greenish-yellow, in shade becoming greyish-green, marginal lobes concolourous with thallus centre, upper cortex interspersed with brownish granules soluble in K. Apothecia usually numerous and grouped, 0.5–2.5 mm diam., rounded, sessile, adnate. Margin prominent, rather thick, concolourous with thallus, shiny, entire to crenate when older, disc plane, bright orange, yellowish-orange to brownish-orange, matte, epruinose. Hymenium colourless, 50–80 μm high, hypothecium colourless, paraphyses simple or weakly branched with swollen apices, subhymenium well-differentiated, pale grey. Epihymenium orange-brown, interspersed with small granules dissolving in K. Ascospores 8 per ascus, hyaline, simple, ellipsoid, 11–12  ×  6–7 μm. Pycnidia immersed, simple with bluish-black ostioles, conidia hyaline, filiform, mostly straight 10–2 × 0.5 μm.

Chemistry. Thallus K+ yellow, C+ pale yellow, KC+ yellow, PD–, medulla K+ yellow, C+ yellow, KC+ yellow, PD–. Secondary metabolites detected by TLC: isousnic (+∕ −), usnic (+) and placodiolic (+) acids, as well as unidentified fatty acid (+∕ −) and unidentified terpenoids (+).

Ecology and distribution. On volcanic and granite rocks in warm and sunny places, at lower elevations (200–400 m), and very rare at higher elevations (c. 800 m), in Europe (Poland and Ukraine) and Western Asia.

Comments. Thallus morphology of specimens representing R. subdiscrepans s. str. is rather homogeneous and similar to R. opiniconensis. Both species have a placodioid thallus, squamulose-bullate in the centre, however R. opiniconensis usually has an orange tint to the thallus, especially visible in older collections. Nevertheless, R. subdiscrepans s. str. produces fatty acids that have not been found in other taxa of R. subdiscrepans s. lat.

Specimens examined. HUNGARY: Montes Matra, region montis Kékes, in rupibus Saskö dictis, on andesitic rock, alt. 880 m, 6 June 1974, G. Kiszely & A. Vezda (KRAM, two specimens). POLAND. SUDETY MTS: Przedgórze Sudeckie foreland, Wzgórza Strzegomskie Hills, Góra Krzyzowa Mt, alt. 354 m, on basalt rocks in open place, 4 Oct. 2013, K. Szczepanska 923, 967 (KRAM, hb. Szczepanska). RUSSIA. CHELYABINSK REGION: Kizilskoe, Obruchevka (c. 20 km E of Kizilskoe), Mt Razbornaya, c. 8 km W of village, alt. c. 450 m., on granite outcrops and stones, 25 June 2011, J. Vondrák 9408 & Vondráková (PRA); Kizilskoe, Obruchevka (c. 20 km E of Kizilskoe), Mt Razbornaya, c. 8 km W of village, alt. c. 450 m., on sun-exposed granite outcrops and stones, 25 June 2011, J. Vondrák 9418 & Vondráková (PRA). Orenburg region: surroundings of water reservoir “Iriklinskoe vodokhranilishche”, Iriklinskiy, vill. Chapaevka, volcanic and limestone rocks on opposite slope of lake W of village, alt. 270–290 m, on sunny volcanic rock, 11 June 2011, J. Vondrák 9384, 9385, 9415 & Frolov (PRA), 9422 & Frolov (PRA, two samples); Kuvandik, vill. Maloe Churaevo (c. 25 km N of Kuvandic) camp c. 2 km W of illage, steppes and Quercus robur-Tilia cordata-Ulmus laevis woodland areas around camp, alt. 250–500 m, on sun-exposed siliceous outcrops, 27 June 2011, J. Vondrák 9412 & Vondráková (PRA); surroundings of water reservoir “Iriklinskoe vodokhranilishche”, vill. Chapaevka, on opposite slope of lake, volcanic rock in valley of stream “Verkhnaya Orlovka”, alt. 270–300 m, on sun-exposed siliceous outcrops, 11 June 2011, J. Vondrák 9416 & Frolov (PRA), 9420 & Frolov (PRA, four samples). REPUBLIC OF BASHKORTOSTAN: surroundings of water reservoir “Iriklinskoe vodokhranilishche”, vill. Tashtugay, rocks in valley of river Tanalik, c. 4 km S of village, alt. 260–300 m, on acid schist, 10 June 2011, J. Vondrák 9411 & Frolov (PRA). SWEDEN. Ngermanland: Vibyggeråpar., Värna, Valaberget (Mt c. 32 km SSE of Örnskoldsvik), steep southern slope facing the sea, on steep rocks to S, alt. 10 m, 11 Aug. 1986, R. Moberg 6937 (KRAM).

New combination

Rhizoplaca pseudomellea (B.D. Ryan) Szczepanśka, Rodriguez-Flakus & Śliwa, comb. nov. ≡ Lecanora pseudomellea B.D. Ryan, Bryologist 96: 295. 1993.

MB 835038

Figs. 2E, 2F.

Type: [USA]. CALIFORNIA: Alpine Co., 1 mi E of Monitor Pass, along Calif. Hwy 89, 16 mi E of Markleeville, alt. 2440 m, field of small rocks among Sagebrush, 24 June 1985, B.D. Ryan 13609 (holotype, ASU!).

Thallus lichenized, placodioid, usually distinctly rosette, moderately attached to the substrate, 1–4 cm diam. Marginal lobes distinctly developed, 1–3.5 mm long and 0.7–2 mm wide, slightly convex, smooth, contiguous, sinusoid, minutely broadened and grey, greenish-black to black at the tips. Thallus centre areolate, areoles convex, bullate, rounded to irregular, smooth or slightly rough, 0.5–2 mm diam. Upper surface smooth, glossy, yellowish-brown, orange-brown to reddish- or rusty brown, marginal lobes paler, yellowish, except darker tips. Apothecia grouped in the thallus centre, 0.5–2.5 mm diam., rounded to irregular when older, sessile. Margin thin, concolourous with thallus, shiny, flexuose, slightly raised when young, becoming excluded, disc concolourous with thallus or darker, reddish-brown, glossy, plane to slightly convex, epruinose. Hymenium colourless, 60–70 μm high, with colourless hypothecium. Epihymenium orange-brown, interspersed with small granules dissolving in K. Ascospores 8 per ascus, hyaline, simple, ellipsoid, 10–12 × 6–7 μm. Pycnidia punctiform, globose, immersed, simple with bluish-black ostioles, c. 260 μm diam., conidia hyaline, filiform, 10–25 ×  0.5 μm.

Chemistry. Thallus K–, C–, KC–, PD–, medulla K–, C–, KC–, PD–. Secondary metabolites detected by TLC: isousnic ( +∕ −), usnic (+∕ −), psoromic (+∕ −) and fatty acids (+∕ −).

Ecology and distribution. On granite and volcanic rocks, in dry and sunny stations, at higher elevations (1,500–2,500 m) in North America.

Comments. R. pseudomellea is characterized mainly by its distinct rosette thallus with long, sinusoid, darker at the tips marginal lobes and orange-brown to reddish-brown, glossy upper surface. Compared to other species treated here, R. pseudomellea has a darker colour, longer and much less convex marginal lobes, as well as a distinctly areolate not squamulose thallus centre. However, its apothecia are usually very similar in colour and morphology to R. phaedrophthalma. R. pseudomellea has very variable secondary chemistry. The most often secondary metabolite occurring in the thallus is isousnic acid, but it may also contains psoromic acid.

Specimens examined. USA. CALIFORNIA: Kern Co., Eastern slope of the southern Sierra Nevada, BLM land on the divide between Oil Canyon and Pine Tree Canyon, along 4WD road which also serves as part of the Pacific Crest Trail, in a pinyon pine and scrub oak woodland, on white shale-like rocks, alt. c. 6150 ft, 19 Apr. 1997, J.R. Shevock 15124 (ASU). OREGON: Harney Co., Steen Mountain (68 ml SSE of Burns), NW slope near snow patch, alt. 8700 ft, 18 July 2006, C.M. Wetmore 95084, 95079 (MIN); Lake Co., Fremont National Forest, Along Dairy Creek, SE part of Gearhart Wilderness, alt. c. 1915 m, 27 Aug 1991, B.D. Ryan 28456 (ASU); Lake Co., Fremont National Forest, Palisade Rocks, SE part of Gearhart Wilderness, c. 1950 m, 11 Sept. 1991, B.D. Ryan 28652 (ASU); Tulare Co., Sequoia National Park, Milk Ranch Peak at border of park E of headquarters, around peak with white fir, incense cedar, pines and rocks, alt. 6100 ft, 21 May 1984, C.M. Wetmore 51151 (MIN); Tuolumne Co., Winter Sports Area, N of Hwy 108, 5 km SW of Fraser Flat Campground, alt. 1700 m, on volcanic rock, pine, oak, 13 Aug. 1989, B.D. Ryan 24402 (MIN).