﻿Revision of Immersaria and a new lecanorine genus in Lecideaceae (lichenised Ascomycota, Lecanoromycetes)

﻿Abstract The species Immersariacupreoatra has been included in Bellemerea. This caused us to reconsider the relationships between Bellemerea and the lecanorine species of Immersaria and to question the monophyly of Immersaria. Amongst 25 genera of the family Lecideaceae, most have lecideine apothecia, the exceptions being Bellemerea and Koerberiella, which have lecanorine apothecia. According to previous classifications, Immersaria included species with both lecanorine and lecideine apothecia. A five-loci phylogenetic tree (nrITS, nrLSU, RPB1, RPB2, and mtSSU) for Lecideaceae showed that Immersaria was split into two clades: firstly, all the lecideine apotheciate species and secondly, all the lecanorine apotheciate species. The latter clade was closely related to the remaining lecanorine apotheciate genera: Bellemerea and Koerberiella. Therefore, the genus concept of Immersaria is revised accordingly and a new lecanorine genus Lecaimmeria is proposed. Furthermore, four new species for Immersaria and seven new species and three new combinations for the new genus Lecaimmeria are proposed. Keys to Immersaria and the new genus Lecaimmeria are provided.

The species Immersaria cupreoatra (Nyl.) Calat. & Rambold (= Lecanora cupreoatra Nyl.) was previously included in Bellemerea Hafellner & Cl. Roux (Clauzade and Roux 1984), then into Immersaria by Calatayud and Rambold (1998). This caused us to reconsider the relationships between Bellemerea and the lecanorine species of Immersaria and to question the monophyly of Immersaria. The family Lecideaceae Chevall originally included all the crustose lecideoid genera, but now only 25 genera have been retained. Most of these are monospecific genera or small genera with under five species (Fryday and Hertel 2014;McCune et al. 2017). Most genera in Lecideaceae have lecideine apothecia. Three exceptions are Bellemerea, Immersaria and Koerberiella Stein, which have lecanorine apothecia. Only Immersaria has both lecanorine and lecideine apothecia, according to the previous circumscription (Calatayud and Rambold 1998;Valadbeigi et al. 2011). Calatayud and Rambold (1998) indicated that the presence of "two types of ascomata" represent different stages of ontogeny. However, there was no molecular evidence that could clarify the species-level phylogenetic relationships within Immersaria. In the two-loci phylogenetic tree of Buschbom and Mueller (2004), the lecideine species Immersaria usbekica (Hertel) M. Barbero, Nav.-Ros. & Cl. Roux was related to Lecidea tessellata Flörke. However, because only two loci of one lecideine species were included, this tree was insufficient to clarify the relationship of the lecanorine apotheciate species in Immersaria.
In this study, a phylogenetic tree of Lecideaceae, based on five loci, is established in order to verify the monophyly of Immersaria. The results show that Immersaria is split into two clades. One clade includes all the lecideine apotheciate species, which is sister to Lecidea tessellata, L. auriculata Th. Fr., Cyclohymenia epilithica McCune & M.J. Curtis and the Porpidia albocaerulescens group and the Porpidia speirea group. The second clade contains all the lecanorine apotheciate species and is closely related to the rest of the lecanorine apotheciate genera within this family: Bellemerea and Koerberiella. Therefore, the genus concept of Immersaria is revised, retaining only the species with lecideine apothecia. The lecanorine species of Immersaria are excluded and proposed as a new genus, Lecaimmeria C.M. Xie, Lu L. Zhang & Li S. Wang. Furthermore, four new species for Immersaria and seven new species and three new combinations for the new genus Lecaimmeria are proposed, based on the four-loci phylogenetic trees. Keys to Immersaria and the new genus are provided below.

Morphological analysis
All the materials for this study were collected in mainland China, mostly from the Qinghai-Tibetan Plateau, during the authors' participation in The Second Tibetan Plateau Scientific Expedition and Research Program. These specimens were stored in the Herbarium of the Kunming Institute of Botany, Chinese Academy of Sciences (KUN) and the Lichen Section of the Botanical Herbarium, Shandong Normal University (SDNU). Type specimens were loaned from the University of Helsinki (H) and Universität Wien (WU). Highresolution photographs of type specimens were provided by the curators of H or obtained from the website Global Plants (https://plants.jstor.org/). Morphological descriptions were made from under a dissecting microscope COIX. Anatomical descriptions were based on observations made from hand-cut sections, mounted in water and using a NIKON microscope. Usually, twenty ascospores were measured and the values of measurement means smallest measured-largest measured, with outlying values in brackets. Photographs were captured with a NIKON Eclipse 50i microscope, equipped with a NIKON digital camera (DSFi2 high-definition colour camera head, NIKON, Japan). The specimens were tested with a 10% aqueous solution of potassium hydroxide (K), a solution of aqueous sodium hypochlorite (C) and 3% Lugol's iodine (I) in the medulla and the surface of the thallus. Secondary metabolites of all the specimens were examined by thin-layer chromatography (TLC) methods, using Solvents A, B and C, as described by Orange et al. (2001).

Phylogenetic analysis
Molecular analysis was carried out on the selected specimens. Genomic DNA was extracted from dry or fresh specimens using a DNAsecure Plant Kit (Tiangen), following the manufacturer's instructions. Five gene loci were amplified by using the following primers: ITS1F (Larena et al. 1999), ITS4 (White et al. 1990), LR0R (Rehner and Samuels 1994), LR5 (Vilgalys and Hester 1990), gRPB1a (Stiller and Hall 1997), fRPB1c (Matheny et al. 2002), RPB2-6f, RPB2-7cr (Liu et al. 1999), mrSSU1 and mrSSU3R (Zoller et al. 1999). The 25 μl PCR mixture consisted of 2 μl DNA, 1 μl of each primer, 12.5 μl 2 × Taq PCR MasterMix (Aidlab) (Taq DNA Polymerase [0.1 unit/ ml]; 4 mM MgCl 2 ; and 0.4 mM dNTPs) and 8.5 μl ddH 2 O. Conditions for PCR of nrITS, nrLSU and mtSSU were set for an initial denaturation at 94 °C for 10 min, followed by 34 cycles of denaturation at 95 °C for 45 s, annealing at 50 °C for 45 s, extension at 72 °C for 90 s and a final extension at 72 °C for 10 min. For RPB1 and RPB2, the parameters were set to an initial denaturation at 94 °C for 10 min, followed by 34 cycles of denaturation at 95 °C for 45 s, annealing at 52 °C for 50 s, extension at 72 °C for 60 s and a final extension at 72 °C for 5 min. The PCR products were sequenced using Sanger technology by the company of Tsingke Biological Technology (Beijing).
The raw sequences were assembled and edited using SeqMan v.7.0 (DNAstar packages). Sequences extracted from new materials with each gene locus were aligned with additional sequences that were available from GenBank (Suppl. material 1: Table S1), by using MEGA v.10.0 and an online version of MAFFT v.7.0 to generate nrITS-nrLSU-RPB1-RPB2-mtSSU or nrITS-nrLSU-RPB1-RPB2 matrices. The five or four gene matrices were combined by SequenceMatrix v.1.7.8. and the concatenated alignments were estimated by PartitionFinder 2 (Lanfear et al. 2017), based on the Bayesian Information Criterion (BIC), to find the most appropriate nucleotide substitution model for each of the five loci.
Phylogenetic relationships were inferred using Bayesian Inference (BI) and Maximum Likelihood (ML). ML analyses were performed with RAxMLHPC using the general time reversible model of nucleotide substitution with the gamma model of rate heterogeneity (GTRGAMMA or GTRCAT). The analyses were run with a rapid bootstrap analysis using 1000 replicates with data partitioned. The Bayesian method was performed with MrBayes v.3.1.2 (Huelsenbeck and Ronquist 2001). Four Markov chains were run with 2 million generations for each dataset and trees were sampled every 100 generations. It was ensured that the average standard deviation of split frequencies was lower than 0.01. Posterior probabilities above 0.9 and bootstrap support above 70% were considered significant supporting values. All the trees were visualised with FigTree v. 1.4.0 (Rambaut 2012).

Results
A total of 172 sequences of the nrITS, nrLSU, RPB1, RPB2, and mtSSU were generated from 61 specimens representing 57 species. Although the five-loci tree only poorly resolved the hierarchy of genera within the family Lecideaceae and the split between the lecanorine and lecideine genera of Lecideaceae was without robust support, nonetheless the results revealed that the genus Immersaria was not a monophyletic lineage. Rather, it was divided into two distant and well-supported lineages: clade 1 which contained the lecideine apotheciate species and clade 2 which contained the lecanorine apotheciate species (Fig. 1).
Clade 1, together with Amygdalaria Norman, Cyclohymenia McCune & M.J. Curtis, Lecidea s str. and Porpidia Körb. (Fig. 1) formed a well-supported clade (93% MLBS and 0.99 PP), all of which have lecideine apothecia. Amygdalaria, Porpidia and Lecidea s str. were nested together, which was consistent with the results of previous research (Buschbom and Mueller 2004;). However, the relationships between these genera still need further research. There was a high level of support for a monophyletic lineage of lecideine apotheciate species of Immersaria, with these being sister to the Lecidea tessellata, L. auriculata, Cyclohymenia, the Porpidia albocaerulescens group and the Porpidia speirea group. The type species of Immersaria (I. athroocarpa) was included in the lineage. Thus, only those Immersaria species with lecideine apothecia belong to Immersaria s. str. This revised concept of the genus Immersaria is as follows: a glossy surface of thallus with an epinecral layer, immersed lecideine apothecia with a reduced margin and Porpidia-type asci with halonate ascospores. There was also a high level of support for clade 2 as a monophyletic lineage (100% MLBS and 1.00 PP), which was clustered with other genera of Lecideales with lecanorine apothecia: Bellemerea and Koerberiella (Fig. 1). Bellemerea could be distinguished from Clade 2 by its amyloid ascospores and Koerberiella by its adnate apothecia. Although the topology of clade 2 for Bellemerea and Koerberiella is not robust, there are conspicuous differences in their morphology and significant differences between the bases in their nucleotide sequences. Since clade 2 is monophyletic with strong support, a new genus, Lecaimmeria, is proposed to accommodate clade 2. The new genus has immersed lecanorine apothecia with a white margin and a distinct plectenchyma developed on top of the orange epihymenium.
Two additional phylogenetic trees were constructed, based on four loci (nrITS, nrLSU, RPB1, and RPB2), in order to assess the phylogenetic position of species within Immersaria and Lecaimmeria, respectively. The phylogenetic tree of Immersaria was comprised of one highly supported clade with five separate lineages, based on 105 sequences from 37 specimens (Fig. 2). All the species with brown, orange, irregular or aggregate thalli formed respective monophyletic lineages. Immersaria shangrilaensis C.M. Xie & Lu L. Zhang formed a well-supported clade and the aggregate areolae clearly distinguished I. shangrilaensis from other species. Immersaria ferruginea C.M. Xie & Li S. Wang also formed a well-supported clade and differed from other species by its greyish-brown thallus. It seems that Immersaria shangrilaensis is sister to I. ferruginea, but the nodes were without support. In addition, the morphology is distinct between Immersaria shangrilaensis and I. ferruginea. The robust lineage Immersaria aurantia C.M. Xie & Li S. Wang was distinguished by its irregular, conspicuously orange thallus and green epihymenium. Immersaria athroocarpa was sister to I. venusta C.M. Xie & Xin Y. Wang, but differed in its convex, polygon areolae and densely crowded apothecia.
The phylogenetic tree of Lecaimmeria was comprised of one well-supported clade with nine separate lineages, based on 140 sequences from 61 specimens (Fig. 3). "Immersaria" sp. and Lecaimmeria iranica (Valadb., Sipman & Rambold) C.M. Xie comprised the basal group. "Immersaria" sp. has only been recorded from Macedonia and Lecaimmeria iranica has been recorded from Inner Mongolia in China and from Iran. Wang, but differed in its crowded apothecia. Lecaimmeria orbicularis formed a highly supported sister group to L. lygaea and L. tibetica, but differed in its round apothecia and the white margin of the apothecia. Lecaimmeria lygaea was seemingly sister to L. tibetica and differed in its areolae having a black margin and with a well-developed prothallus between areolae.

Revised boundaries of Immersaria
Formerly, the boundaries for Immersaria species were: lecanorine or lecideine type of immersed apothecia, production of confluentic acid and gyrophoric acid and ostiole or stellate shapes of conidiomata. However, these characters were not good characters by which to distinguish this genus. The lecanorine species Immersaria cupreoatra was previously included in Bellemerea. Based on many specimens from China, it was also discovered that the ostiole or stellate shapes of conidiomata appeared in different stages of ontogeny. The main substances produced in the genus are confluentic acid and gyrophoric acid; confluentic acid only occurs in lecideine species, whereas gyrophoric acid appears in lecanorine species, with the exception of one lecideine species Immersaria usbekica. Furthermore, these characters, the types of apothecia and the shapes of conidiomata could not be applied as proper delimitations to classify species within Immersaria, neither were they supported by the phylogeny. Phylogenetic tree constructed from Maximum Likelihood analyses in Immersaria, based on the concatenated nrITS-nrLSU-RPB1-RPB2 dataset. Maximum Likelihood bootstrap probabilities above 70% (left) and Bayesian Inference posterior probabilities above 0.9 (right) are given at the nodes. Solid brown rectangle: thallus brown; solid orange rectangle: thallus yellow brown to orange brown; solid red rectangle: thallus rusty; hollow brown rectangle: thallus pale yellow brown. Solid green circle: green epihymenium; solid grey circle: without apothecia; hollow green circular: brown epihymenium.
The five-loci based analysis ( Fig. 1) was incompatible with previous circumscriptions of the genus Immersaria, the members of which in this study are defined by their lecideine immersed apothecia, brown areolae with an epinecral layer and brown/green epihymenium without a plectenchyma. Consequently, a new genus, Lecaimmeria, Figure 3. Phylogenetic tree constructed from Maximum Likelihood analyses in Lecaimmeria, based on the concatenated nrITS-nrLSU-RPB1-RPB2 dataset. Maximum Likelihood bootstrap probabilities above 70% (left) and Bayesian Inference posterior probabilities above 0.9 (right) are given at the nodes. Solid purple rectangle: areolae margin white; solid grey rectangle: areolae margin black; hollow purple rectangle: areolae margin jade green pigmented. Solid blue circle gyrophoric acid; hollow blue circle: none. is established to accommodate the excluded lecanorine species. The new taxonomic system, proposed here, revised the classification boundaries between Immersaria and Lecaimmeria, but it may still be difficult to distinguish between them in cases when apothecia are absent. In this case, they could be distinguished by the substances produced or by molecular methods.

Diagnostic traits within species of Immersaria and Lecaimmeria
Species of Immersaria could be identified by their different thallus colours (indicated in Fig. 2). Immersaria ferruginea has a conspicuously greyish-brown thallus, whereas I. athroocarpa, I. aurantia, I. shangrilaensis and I. venusta have a reddish-brown thallus. Immersaria athroocarpa (indicated in Fig. 2) is the species that mostly has a green epihymenium, whereas the other species mostly have a brown epihymenium. Almost all these species contain confluentic acid, which is often accompanied by 2'-O-methylmicrophyllinic acid. Planaic acid, which is newly reported from this genus, is only presented in specimens of Immersaria aurantia, I. shangrilaensis and I. venusta. All the characters discussed above were supported by the phylogeny, thus could be used as key characters to differentiate species in Immersaria.
Species of the new genus Lecaimmeria could be delimited by the colours of their areolae and margins, the existence of an apothecial margin and usually by the lack of substances. The margin of areolae (indicated in Fig. 3) was usually white, but rarely black or jade green. Lecaimmeria lygaea could be easily distinguished by the black margin of the areolae. The jade green margin occurs in Lecaimmeria tuberculosa, which grows on Qilian jade. The areolae margin of Lecaimmeria qinghaiensis is white, but is occasionally pigmented with very slightly green colour. The margin of the apothecia is absent in Lecaimmeria tuberculosa and L. iranica, whereas the apothecia of the other species have white margins. Most species of Lecaimmeria lack secondary metabolites, while gyrophoric acid was detected only in Lecaimmeria botryoides, L. iranica and L. mongolica (indicated in Fig. 3). In addition, an orange thallus appeared only in Lecaimmeria mongolica and L. tibetica, whereas the remaining species were brownish.
Chemistry. Thallus K-, C-. Medulla I+ violet. Confluentic acid, often accompanied with 2'-O-methylmicophyllinic acid, planaic acid or no substances detected by TLC. The compound planaic acid is newly reported in this genus.
Ecology and distribution. In China, growing on bare rock, sandstone or granite, from elevations of 3800 to 4500 m in the alpine zone of west China and elevations of 1200 to 1900 m in the steppe of north China. Worldwide distribution.
Notes. Species with lecanorine apothecia were previously included in Immersaria (Calatayud and Rambold 1998;Valadbeigi et al. 2011), but the five-loci phylogenetic analysis excluded these species from Immersaria. This exclusion entails a restricted concept of the genus. Immersaria is now defined by its orange-brown, yellow-brown, sometimes rusty coloured thallus, the amyloid medulla, the glossy surface of areolae with a pruinose margin, the black immersed lecideine apothecia with a reduced proper margin, the brown epihymenium and the Porpidia-type asci with eight halonate and non-amyloid ascospores. The members of this genus occur in alpine habitats.
Species of Sporastatia A. Massal. might be misidentified as members of Immersaria because of field observations of their glossy areolae and the immersed lecideine apothecia. However, they are characterised by multi-spored asci and their yellowbrown thallus. Additionally, Miriquidica Hertel & Rambold resembles Immersaria by its glossy areolae and the lecideine apothecia, but differs in its black brown thallus, its Lecanora-type asci with non-halonate ascospores and often containing miriquidic acid. The immersed apothecia of Immersaria may resemble Aspicilia A. Massal. and Acarospora A. Massal., but Aspicilia has a white or grey thallus, the Aspicilia-type asci with non-halonate ascospores; Acarospora has multi-spored asci.
Although four known species, Immersaria carbonoidea (J.W. Thomson) Esnault & Cl. Roux, I. fuliginosa Fryday, I. olivacea Calat. & Rambold and I. usbekica, currently lack molecular data, they are temporarily left in Immersaria due to their morphology which corresponds to that of Immersaria. Our morphological comparisons were based on high-resolution photographs of type materials and the original descriptions.
Ecology and distribution. In China, growing on granite in arid and semi-arid steppe habitats at elevations of 1200-1950 m. Worldwide distribution. This species is known from Inner Mongolia and Mt. Changbai (Hertel and Zhao 1982) in China.
Notes. The lectotype grows on siliceous rock and contains several intact apothecia. The materials from Inner Mongolia are identical with the lectotype, based on comparisons with high-resolution photographs and the description given by Hertel (1977). It is, therefore, treated as Immersaria athroocarpa at present. Some Inner Mongolian materials contain an unknown substance, but form a well-supported clade with other materials. Immersaria athroocarpa is characterised by the convex, yellow-brown areolae and the large sizes of ascospores. In this genus, only this species has ascospores up to 20.0 μm long.
Ecology and distribution. In China, growing on rock at elevations of 3900-4300 m in the alpine zone. This species is known from Qinghai, Sichuan Province and Tibet of China.
Notes. Immersaria aurantia is characterised by its distinct orange, irregular areolae and the mostly green epihymenium. Immersaria athroocarpa and I. venusta are similar to I. aurantia, but I. athroocarpa differs in the convex, regularly polygonal areolae and the more crowded apothecia; I. venusta differs in having yellow-brown, often rusty, cracked areolae and flat apothecia. Additionally, confluentic acid and planiaic acid do not appear simultaneously in Immersaria aurantia, whereas I. venusta always contains both compounds. Ecology and distribution. In China, growing on quartz sandstone or granite at elevations of 3800-4300 m in the alpine zone. This species is known from Sichuan Province and Tibet of China.
Ecology and distribution. In China, growing on granite at elevations of 4300-4500 m in the alpine zone. This species is known from Yunnan Province of China.
Ecology and distribution. In China, growing on quartz sandstone or granite at elevations of 3900-4300 m in the alpine zone. This species is known from Qinghai Province of China.
Notes. Immersaria venusta is characterised by its yellow-brown, cracked areolae, its flat apothecia and brown epihymenium. It resembles Immersaria shangrilaensis by its cracked areolae, but its areolae have the tendency to split into several patches, but not aggregate like those of I. shangrilaensis. Immersaria athroocarpa is similar to I. venusta in the brown appearance of its thallus and in forming a sister group in the phylogenetic tree, but it differs in its yellow brown thallus, convex areolae, densely crowded apothecia and larger ascospores (17.5-20.0 × 10.0 μm). Immersaria venusta is also similar to I. aurantia (see notes for I. aurantia). The brown thallus of Immersaria venusta possibly resembles that of I. olivacea, but the latter differs in its simple or one-septate ascospores, pyriform conidia and dark bluish-green epihymenium. Description. Thallus crustose, red-brown, orange-brown or dark brown, continuous or not; areolae irregular or tending to rectangular, with a glossy surface (atrobrunnea-type) caused by a layer of dead, colourless cells above the upper cortex; margin white or black; prothallus distinct at the margin of thallus or absent, sometimes developed between areolae. Upper cortex orange; epinecral layer colourless; algal layer continuous; medulla filled with grey granules. Apothecia lecanorine, immersed, round or irregular; disc red-brown, dark red-brown or dark orange-brown, flat or concave; margin present or absent, black or white, rarely green, pruinose or not. Exciple reduced, tissue at the lateral sides of the hymenium corresponding to the upper cortex and the algal layer of the vegetative areolae and to hypothecial hyphal cells when apothecia reach the margin of the areole (indicated in Figs. 7c, g, 8c, 9c, h, 10c, h). Hymenium colourless; paraphyses simple, rarely branched, anastomosing or not; epihymeinum orange, orange-brown, rarely brown, with a plectenchyma. Asci Porpidia-type (indicated in Fig 9d), cylindrical, eight-spored; ascospores ellipsoid, halonate, nonamyloid. Conidiomata present or absent, immersed, rarely convex, linear or stellate, rarely tuberculiform; conidia bacilliform.

Specimens examined (KUN
Chemistry. Thallus K-, C+/-. Medulla I+ violet. Gyrophoric acid, 4-O-demethylplanaic acid or no substances detected by TLC. Ecology and distribution. In China, growing on rock, sandstone, granite or Qilian jade (rare), from elevations of 3100 to 4800 m in the alpine zone of west China and from 1200 to 1900 m in the steppe of north China. This genus is known from China, Europe, Iran, Mongolia, Romania, Russia, and USA.
Notes. The five-loci phylogenetic analysis showed that the species with lecanorine apothecia formed a novel lineage and should be excluded from Immersaria; thus, they are here treated as a new genus Lecaimmeria. Lecaimmeria is distinguished from related genera by its glossy surface, orange or red-brown areolae with margins, the amyloid medulla, the red-brown immersed lecanorine apothecia, the orange epihymenium with a plectenchyma and the Porpidia-type asci with eight halonate and non-amyloid ascospores. In China, the genus is distributed in alpine areas, high altitude desertsteppe areas or high latitude steppe. Almost all the species of Lecaimmeria grow on granite or sandstone, with the exception of one species, L. tuberculosa, which grows on jade. Interestingly, the margin of conidiomata and areolae of Lecaimmeria tuberculosa appear with heavily jade-green pruinose.
The immersed apothecia and brown thallus of Lecaimmeria often resemble those of Immersaria, but Lecaimmeria differs in its red-brown lecanorine apothecia, often with a white margin, their orange epihymenium with a plectenchyma and the thallus containing gyrophoric acid. This genus might be confused with Bellemerea by its lecanorine apothecia and the Porpidia-type asci with halonate ascospores, but the latter genus differs in its amyloid ascospores.
Three species, previously included in Immersaria, I. cupreoatra, I. iranica and I. mehadiana, have lecanorine apothecia, but two of these, I. cupreoatra and I. mehadiana, currently lack molecular sequences. We suggest that these three species should be transferred to Lecaimmeria, based on the following factors. Their morphology is consistent with Lecaimmeria according to molecular results and comparisons with type specimens, high-resolution photographs of the type materials and the original descriptions. One unknown "Immersaria" species from Macedonia is sister to Lecaimmeria iranica in the phylogenetic tree (Fig. 3), but comparison with high-resolution photograph and previous records (Malíček and Mayrhofer 2017) show that it differs in its black margin of areolae and absence of gyrophoric acid. This unknown species with lecanorine apothecia is possibly a member of Lecaimmeria, but descriptions are lacking and the specimens were not seen. Thus, this species is temporarily retained in Immersaria. Notes. Lecaimmeria botryoides is characterised by its discontinuous thallus, densely crowded apothecia while immature and the orange epihymenium. Lecaimmeria orbicularis is similar to L. botryoides in its red-brown thallus, but differs in its round, flat apothecia and continuous thallus. The red-brown thallus of Lecaimmeria botryoides resembles L. cupreoatra, but the latter differs in the black margin of its apothecia and its distinct black prothallus between areolae.
Chemistry. Thallus K-, C-. Medulla I+ violet. Unknown fatty acid by TLC. Ecology and distribution. In China, growing on sandstone at elevations of 4000-4200 m in the alpine zone. This species is known from the Tibet Region of China.

Lecaimmeria mongolica
Chemistry. Thallus K-, C+. Medulla I+ violet. Gyrophoric acid. Ecology and distribution. In China, growing on granite at elevations of 1400-2000 m in steppe or mountains. This species is known from Inner Mongolia of China.
Chemistry. Thallus K-, C-. Medulla I+ violet. None. Ecology and distribution. In China, growing on granite or sandstone at elevations of 3700-4200 m in the alpine zone. This species is known from Qinghai and Sichuan Provinces of China.
Notes. Lecaimmeria orbicularis is characterised by its orange brown thallus, neatly arranged areolae and round, flat apothecia. Lecaimmeria botryoides is similar to L. orbicularis (see notes for L. botryoides). Lecaimmeria mongolica might be confused with L. orbicularis due to its large apothecia with a white margin, but differs in its red-brown thallus and distribution in steppes. The red-brown thallus of Lecaimmeria cupreoatra resembles that of L. orbicularis, but differs in the black margin of its apothecia and its distinct black prothallus between areolae.
Chemistry. Thallus K-, C-. Medulla I+ violet. None. Ecology and distribution. In China, growing on rock at elevations of 4600-4900 m in the alpine zone. This species is known from Qinghai Province of China.
Chemistry. Thallus K-, C-. Medulla I+ violet. None. Ecology and distribution. In China, growing on quartz sandstone at elevations of 4300-4600 m in the alpine zone. This species is known from Tibet, China.
Chemistry. Thallus K-, C-. Medulla I+ violet. No substances by TLC. Ecology and distribution. In China, growing on the Qilian jade or sandstone at elevations of 3900-4200 m in the alpine zone. This species is known from Qinghai Province and Gansu Province of China.
Notes. Lecaimmeria tuberculosa is characterised by its red-brown thallus, the jadegreen pruinose colour at the margin of its areolae, its red-brown, concave apothecia without a proper margin and tuberculiform conidiomata. Lecaimmeria qinghaiensis is similar to L. tuberculosa (see notes for L. qinghaiensis). Lecaimmeria tuberculosa usually grows on jade and, interestingly, the margin of the conidiomata and areolae of the species are heavily jade-green pigmented. Lecaimmeria iranica resembles L. tuberculosa by the absence of an apothecial margin, but differs in its immersed conidiomata and the white margin of its areolae.