Integrative taxonomy and genus delimitation in the Rhizocarpaceae (lichenized Ascomycota)

Abstract

The Rhizocarpaceae, a family of lichenized fungi within the Ascomycota, comprises approximately 160 species within five genera: Catolechia, Epilichen, Haugania, Poeltinula, and Rhizocarpon. Rhizocarpon is the most species-rich, with about 150 species predominantly inhabiting siliceous rock in boreal and arctic-alpine environments. Molecular phylogenetic studies have revealed that current taxonomy, heavily reliant on morphology, chemistry, and life strategies, renders Rhizocarpon paraphyletic. This study aims to elucidate the phylogenetic relationships and clarify genus delimitation within the Rhizocarpaceae using an integrative taxonomic approach that combines three genetic markers and a diversity of taxa covering the morphological, chemical, and ecological spectrum of the family. Our comprehensive sampling includes 50 species across the Rhizocarpaceae collected from diverse geographical locations and ecological settings. Our phylogenetic hypothesis is based on a concatenated dataset of two nuclear (ITS and MCM7) and one mitochondrial (mtSSU) genetic marker. Ascospore characteristics and thallus pigmentation alongside secondary metabolite profiles were mapped onto this DNA-based evolutionary framework. Our results underscore significant refinements in the classification of the Rhizocarpaceae, highlighting the inadequacy of traditional taxonomic markers alone to infer robust phylogenetic affiliations. We advocate for new circumscriptions of Catolechia, Poeltinula, and Rhizocarpon based on the molecular phylogeny and propose synonymizing Epilichen with Catolechia, the transfer of the species in the R. hochstetteri complex to Poeltinula, and the resurrection of Rehmia. We hence propose 24 new combinations and three typifications. Collectively, this study sets the groundwork for future research and stability in the systematics of the Rhizocarpaceae, augmenting our understanding of their diversity and evolutionary dynamics.

Citation: Möller EJ, Timdal E, Haugan R, Bendiksby M (2025). Integrative taxonomy and genus delimitation in the Rhizocarpaceae (lichenized Ascomycota). Fungal Systematics and Evolution 16: 215–231. doi: 10.3114/fuse.2025.16.12

INTRODUCTION

The Rhizocarpaceae (Ascomycota: Lecanoromycetes: Lecanoromycetidae: Rhizocarpales) is a family of lichenized fungi that consists of ca 160 currently accepted species unevenly distributed across five genera: Catolechia (one species), Epilichen (three species), Haugania (two species), Poeltinula (three species), and Rhizocarpon (ca 150 species) (Fryday et al. 2024). In addition, Hafellner (1984) lists nine generic names as synonyms of these (Catillariopsis, Catocarpus, Dimaura, Diphaeis, Diphanis, Lepidoma, Phalodictyum, Rehmia, and Siegertia). The current generic circumscriptions are based mainly on thallus morphology and ecology, and some phylogenies render Rhizocarpon paraphyletic, suggesting that the current taxonomy needs revision (Ihlen & Ekman 2002, Miadlikowska 2014). In this study we include three genetic markers and a broad taxonomic sampling to investigate this suspected paraphyly.

The family is characterized by having a crustose thallus (lobate in one species) or lacking a thallus in some lichenicolous species; green, unicellular photobionts (very rarely also cyanobacteria in cephalodia); lecideine apothecia (i.e., having a proper exciple without a thalline margin); richly branched, anastomosing, and strongly conglutinated paraphyses; and asci with a moderately amyloid tholus with an internal, apical, and deeper pigmented amyloid cap (a small structure at the apex of the ascus; Honegger 1980, Hafellner 1984). The ascospores, usually eight per ascus (sometimes a reduced number), are 1-septate to eumuriform, hyaline to green or dark brown, and often surrounded by a gelatinous perispore.

Of the five genera in the Rhizocarpaceae, Rhizocarpon is by far the most speciose with ca 150 species. They grow mainly on rocks in the boreal and arctic-alpine regions, but the genus also occurs throughout the temperate, subtropical, and even tropical regions (GBIF 2025). The type species, R. geographicum was first described by Linnaeus (1753) as Lichen geographicus. The species is both abundant and globally distributed, sometimes completely dominating alpine rock surface ecosystems. The species is, however, very diverse and poorly understood and must be considered a species complex (Roca-Valiente et al. 2016).

Rhizocarpon was described by De Candolle in Lamarck & De Candolle (1805) and represents one of the earliest described lichen genera. The species in the genus grow mainly on siliceous rock. Although most species are autonomous lichens from the start, some start their lives growing on other lichens (lichenicolous) and carry on autonomously after consuming their host, and some species are persistent obligately lichenicolous on other crustose lichens (Holtan-Hartwig & Timdal 1987, Timdal & Holtan-Hartwig 1988, Poelt 1990). A typical lichenized Rhizocarpon thallus is areolate (or sometimes more continuous and cracked) with a conspicuous, usually black hypothallus from which the areolae and apothecia originate. A few species are mainly sterile and propagate vegetatively by isidia or soredia. The number of species in the genus is difficult to estimate accurately as many areas of the world lack lichen checklists and those that exist have a partly incompatible taxonomy for Rhizocarpon. We estimate roughly 150 species, although Lücking et al. (2017) indicates there may be as many as 225 species and MycoBank (2025) lists 258 accepted species names out of 361 legitimate names.

Catolechia is currently a monotypic genus. This narrow circumscription was established by Koerber (1855) after exclusion of some additional species placed in the genus by Flotow (1850; the protologue) and Massalongo (1852). Catolechia wahlenbergii was first described as Lecidea wahlenbergii and moved to Catolechia by Koerber in 1855. Catolechia was placed in the Rhizocarpaceae by Hafellner (1984) based primarily on ascus type. It can be found growing on soil and moss-carpets in shaded crevices of humid rock walls in boreal and arctic-alpine habitats. According to GBIF (2025), the species is found mainly in the Northern Hemisphere, with some outliers in the tropics that may need confirmation.

The three species of Epilichen are all small and lichenicolous on crustose lichens. The genus was described by Clements (1909) and placed in the Rhizocarpaceae by Hafellner (1984), also based mainly on ascus type. The type species, E. scabrosus, and E. glauconigellus grow on terricolous Baeomyces spp. (Ihlen 1997). The former is known from throughout the Northern Hemisphere, southernmost South America and New Zealand, while the latter is only registered from Europe and Greenland (Fryday 2019, GBIF 2025). The third species, E. stellatus, parasitizes Lecidea tessellata and is reported from the type locality in the Russian Far East and from North America (Triebel 1989, Triebel et al. 1991).

The Catolechia and Epilichen species lack a perispore around their ascospores. Otherwise, the two genera are anatomically similar to the rest of the family by having Rhizocarpon-type asci and brown, 1-septate ascospores. Both C. wahlenbergii and E. scabrosus contain two yellow pigments, apparently pulvinic acid derivatives, but not the common yellow pigment of Rhizocarpon, rhizocarpic acid. In addition to their different ecologies, Hafellner (1979) notes that the two species mainly differ in thallus thickness and apothecial ontogeny. There is also a significant size difference between the two species. While C. wahlenbergii is one of the larger species in the family, with thalli often reaching 5–10 cm or more in diameter, Epilichen species rarely grow larger than a centimetre in diameter.

Haugania was described as a segregate of Rhizocarpon in Fryday et al. (2024), based on its phylogenetic position (to be presented here), the rusty thallus colour, and metallophilic habitat preference. The genus was established for two species, H. oederi and H. pycnocarpoides, both of which grow on heavy metal rock-substrates, often in abandoned mining areas. Both species have halonate, hyaline ascospores; H. oederi having three-septate to submuriform ascospores and H. pycnocarpoides having sub- to eumuriform ascospores.

Poeltinula was introduced as a new name for Melanospora Mudd (nom. illeg.) and originally included only a single species, P. cerebrina (Hafellner 1984). Later, three more species have been included in the genus, P. cacuminum, P. cerebrinella, and P. interjecta. The latter species is currently included in Melaspilea (e.g., by BLS 2025), a view we follow here although its generic position is not supported by any public DNA sequence. None of the Poeltinula species make conspicuous thalli, but are instead endolithic and appear only as small, black apothecia with furrows of sterile tissue on calcareous rock. The species all have 1-septate, halonate Rhizocarpon-type ascospores; those of P. cerebrina and P. cacuminum soon becoming green. Poeltinula was placed in the Rhizocarpaceae by Hafellner (1984) based mainly on ascus type. According to GBIF (2025), the type species, P. cerebrina, occurs in the Alps and the British Isles, while P. cacuminum occurs only in the Alps. The third species, P. cerebrinella, is known only from the type locality on Kerguelen and from the Marion Island in the Antarctic Sea and must be considered poorly understood.

Even though the Rhizocarpaceae, especially Rhizocarpon, is both widespread and abundant, there is no comprehensive molecular phylogenetic study focusing on the genus or family level relationships. Most published Rhizocarpon sequences were provided for phylum and class level phylogenetics (e.g., Lutzoni et al. 2001, Miadlikowska et al. 2014, Kraichak et al. 2018) or, alternatively, for studies focusing on some other lichen family (e.g. Buschbom & Mueller 2004). At the other end of the scale, molecular phylogenies have been provided in support of descriptions of new species (McCune et al. 2016, Davydov & Yakovchenko 2017, Halici et al. 2022, Timdal et al. 2024).

The only study known to us that addresses inter-genus relations and character evolution in the Rhizocarpaceae is Ihlen & Ekman (2002), which presented a phylogeny based on ITS and mtSSU sequences of 15 species of Rhizocarpon, Catolechia and Poeltinula. In their study, R. hochstetteri is the phylogenetic sister to P. cerebrina. These two species do not resemble each other morphologically (Fig. 1), although they do share one key character: sharply delimited pigmented apical caps on the otherwise hyaline paraphyses (Fryday 2002, Möller 2021), a character state that is mostly lacking in the rest of the Rhizocarpaceae. Pigmented paraphysis caps, albeit not as swollen and less sharply delimited, are also reported in some yellow species of Rhizocarpon (Runemark 1956). In the study by Ihlen & Ekman (2002), the Poeltinula-R. hochstetteri-clade was either sister to the rest of the family or sister to the rest of Rhizocarpon, with Catolechia recovered as sister to the rest of the family. In the large phylogeny of the Lecanoromycetes by Miadlikowska et al. (2014), Poeltinula is not included, but the R. hochstetteri group is phylogenetic sister to Catolechia, and H. oederi (as R. oederi) is sister to the rest of the family [with the exception of an accession of R. geographicum (AFTOL-ID 2067) that appears to be based on two specimens, from which one of the three markers, nucSSU GenBank AF088246, seems to be a contamination].

Fig. 1.

Habitus photographs of selected species of the Rhizocarpaceae. A. Rhizocarpon geographicum, Norway, O-L-161808. B. R. norvegicum, Norway, O-L-163568. C. R. parvum, Norway, O-L-225584. D. R. eupetraeum, Greenland, O-L-139134. E. R. hochstetteri, Norway, O-L-149415. F. Poeltinula cacuminum, Austria, Hafellner 63791, GZU. G. R. caesium, Iceland, O-L-160733. H. R. amphibium, Norway, O-L-169822. I. R. umbilicatum, Norway, O-L-123119. J. R. cinereovirens, Norway, O-L-163425. K. R. lavatum, Norway, O-L-169833. L. Epilichen scabrosus, Norway, O-L-164936. M. Catolechia wahlenbergii, Norway, not collected. N. Haugania oederi, Norway, O-L-51867. O. H. pycnocarpoides, Norway, O-L-184267. Photo credits: E. Timdal.

The position of the Rhizocarpaceae in the Lecanoromycetidae is still not resolved. Some studies have placed the family as the phylogenetic sister to the Sporastatiaceae and the Rhizocarpaceae + Sporastatiaceae as sister to the rest of the Lecanoromycetidae (Miadlikowska et al. 2014, Kraichak et al. 2018). Others placed the Sporastatiaceae as sister to the Rhizocarpaceae + the rest of the Lecanoromycetidae (e.g., Bendiksby & Timdal 2013). Whichever of these two topologies is correct, the long genetic distance between the Rhizocarpaceae and its nearest neighbour makes investigations of the deep topology and selection of a suitable outgroup for the family challenging. This was also highlighted by Ihlen & Ekman (2002).

According to the molecular phylogenetic hypotheses by Ihlen & Ekman (2002) and Miadlikowska et al. (2014), Rhizocarpon is, as currently circumscribed, paraphyletic. In both studies, the R. hochstetteri complex seems to be more closely related to one of the smaller genera, either Poeltinula or Catolechia, than to Rhizocarpon.

In this study, we aim to discover, understand, delimit, and circumscribe the genera of the Rhizocarpaceae, reveal their phylogenetic interrelations, and identify informative diagnostic characters at genus level. To reach that goal, we include a set of species that covers most of the anatomical, morphological, chemical and ecological variation of the family. We study these features integratively within the framework of a phylogenetic hypothesis based on three genetic markers to conclude meaningful taxon limits.

MATERIAL AND METHODS

Taxon sampling

In this study, we have included 87 specimens representing 50 species of the Rhizocarpaceae, both freshly collected from field work in Norway during the summers of 2019 and 2020 and as dried specimens from various fungaria (BG, E, GZU, MSC, O, OSC, hb Wheeler; Table 1). The fungarium material includes specimens from Austria, Canada, Iceland, Japan, Norway, Russia, Sweden, UK and USA. Specimens were selected to represent most of the breadth of variation in ascospore morphology (i.e., septation, pigmentation, and number of spores per ascus), chemistry, and ecology in the Rhizocarpaceae. Lecanora cavicola and L. coronulans were used as outgroup taxa, as they are phylogenetically far from the Rhizocarpaceae and had all the included markers available in GenBank.

Table 1.

Voucher specimen table. Current name, previous name, voucher ID, country, GenBank accession numbers for the ITS, mtSSU, and MCM7 markers (sequences produced for this study are in bold), and references for existing sequences are provided. Taxonomic novelties are highlighted in bold text in the first column.

Current name	Previous name	Voucher	Country	ITS	mtSSU	MCM7	Reference
Catolechia glauconigella	Epilichen glauconigellus	O-L-141634	Norway	MK811705	—	—	Marthinsen et al. (2019)
		O-L-179907	Norway	MK812566	—	—	Marthinsen et al. (2019)
		O-L-239178	Norway	PV665039	—	—	This study
Catolechia scabrosa	Epilichen scabrosus	O-L-164936	Norway	MK812386	PV661952	PV738713	Marthinsen et al. (2019)
		O-L-179955	Norway	MK812558	—	—	Marthinsen et al. (2019)
		O-L-225908	Norway	—	PV661953	—	This study
		O-L-227683	Norway	PV665040	—	PV738714	This study
Catolechia wahlenbergii		O-L-226116	Norway	PV665035	PV661948	PV738709	This study
		O-L-226139	Norway	PV665036	PV661949	PV738710	This study
		O-L-227703	Norway	PV665037	PV661950	PV738711	This study
		O-L-227789	Norway	PV665038	PV661951	PV738712	This study
Haugania oederi		O-L-151401	Norway	PV665041	PV661954	—	This study
		O-L-28978	Norway	AF483612	AF483180	—	Ihlen & Ekman (2002)
		Spribille 36629 (MSC)	USA, Alaska	MN437622	MN508296	MN437622	Spribille et al. (2020)
Haugania pycnocarpoides		O-L-179560	Norway	KR780753	PV661955	—	Westberg et al.(2015)
Lecanora cavicola		Flakus29582	Bolivia	OL604041	OL604121	OK665503	Medeiros et al. (2021)
Lecanora coronulans		Flakus29260	Bolivia	OL604008	OL604090	OK665498	Medeiros et al. (2021)
Poeltinula cacuminum		Hafellner 63781 (GZU)	Austria	PV665042	—	—	This study
		Hafellner 63791 (GZU)	Austria	PV665043	—	—	This study
Poeltinula caesia	Rhizocarpon caesium	E-456329 (Holotype)	Great Britain	PV665049	—	—	This study
		O-L-160733	Iceland	PV665050	PV661959	—	This study
Poeltinula cerebrina		Mayrhofer et al. 12838 (GZU)	Austria	AF483173	AF483606	—	Ihlen & Ekman (2002)
Poeltinula hensenniae	Rhizocarpon hensenniae	O-L-209863	Japan	PV665058	PV661967	PV738719	This study
		O-L-209874	Japan	—	PV661968	PV738720	This study
Poeltinula hochstetteri	Rhizocarpon hochstetteri	O-L-222653	Norway	PV665059	PV661969	PV738721	This study
		O-L-225971	Norway	PV665060	PV661970	PV738722	This study
		O-L-227846	Norway	PV665061	—	—	This study
		O-L-227912	Norway	PV665062	PV661971	PV738723	This study
Poeltinula infernula	Rhizocarpon infernulum	McCune 32921 (OSU)	USA, Alaska	—	PV661972	—	This study
Poeltinula sylvatica	Rhizocarpon infernulum f. sylvaticum	O-L-119602	Norway	AF483607	—	—	Ihlen & Ekman (2002)
		O-L-228034	Norway	PV665063	—	PV738724	This study
Rehmia amphibia	Rhizocarpon amphibium	BG-L-68527	Sweden	AF483611	AF483179	—	Ihlen & Ekman (2002)
		O-L-163528	Norway	PV665045	PV661957	—	This study
Rehmia caeruleoalba	Rhizocarpon caeruleoalbum	9.8.1987 Poelt (GZU)	Austria	PV665046	—	—	This study
		hb Wheeler 5259	USA, Alaska	PV665047	—	—	This study
		UPS-L-67821	Norway	PV665048	—	—	This study
Rehmia chionea	Rhizocarpon chioneum	O-L-175723	Norway	PV665051	PV661960	—	This study
		O-L-195535	Norway	KY266951	—	—	Westberg et al. (2015)
Rehmia cinereovirens	Rhizocarpon cinereovirens	O-L-174113	Norway	—	PV661961	—	This study
Rehmia furfurosa	Rhizocarpon furfurosum	O-L-169766	Norway	PV665056	PV661963	—	This study
Rehmia haidensis	Rhizocarpon haidense	Fryday 10680 (MSC)	USA, Alaska	MN483143	MN508295	—	Spribille et al. (2020)
Rehmia lavata	Rhizocarpon lavatum	O-L-228204	Norway	PV665066	—	—	This study
		O-L-28923	Norway	AF483610	AF483178	—	Ihlen & Ekman (2002)
Rehmia petraea	Rhizocarpon petraeum	O-L-169123	Norway	PV665072	PV661979	—	This study
		O-L-225882	Norway	PV665073	PV661980	PV738725	This study
		O-L-29039	Norway	AF483609	AF483177	—	Ihlen & Ekman (2002)
Rehmia reducta	Rhizocarpon reductum	McCune 36587 (OSU)	USA, Oregon	PV665077	PV661985	PV738729	This study
Rehmia roridula	Rhizocarpon roridulum	O-L-201497	Norway	PV665079	PV661986	—	This study
Rehmia rubescens	Rhizocarpon rubescens	UPS-L-131191	Sweden	PV665080	—	—	This study
Rehmia tetramera	Rhizocarpon tetramerum	O-L-29034	Norway	PV665083	—	—	This study
Rehmia timdalii	Rhizocarpon timdalii	O-L-71409	Norway	PV665084	PV661989	—	This study
		UPS-L-160175	Sweden	PV665085	—	—	This study
Rehmia umbilicata	Rhizocarpon umbilicatum	O-L-163744	Norway	PV665086	—	—	This study
		O-L-163789	Norway	PV665087	—	—	This study
		O-L-184352	Norway	PV665088	—	—	This study
Rhizocarpon alpicola		O-L-170541	Norway	PV665044	PV661956	—	This study
		O-L-223755	Canada, British Columbia	ON332060	ON324015	—	Halici et al. (2022)
Rhizocarpon atroflavescens		O-L-228046	Norway	—	PV661958	—	This study
Rhizocarpon dinothetes		O-L-184631	Norway	PV665052	—	—	This study
Rhizocarpon distinctum		O-L-28901	Norway	AF483615	—	—	Ihlen & Ekman (2002)
Rhizocarpon effiguratum		O-L-223731	Canada, Alberta	PQ219499	ON324016	PV738715	ITS: Timdal & Rui (2024) | mtSSU: Halici et al. (2022)
Rhizocarpon eupetraeum		O-L-163557	Norway	PV665053	—	—	This study
		O-L-165382	Norway	PV665054	—	PV738716	This study
Rhizocarpon ferax		O-L-175699	Norway	PV665055	PV661962	—	This study
Rhizocarpon geographicum		O-L-175717	Norway	PV665057	PV661964	—	This study
		O-L-223664	Canada, Alberta	PQ219488	PV661965	PV738717	Timdal & Rui (2024)
		O-L-223711	Canada, Alberta	PQ219494	PV661966	PV738718	Timdal & Rui (2024)
Rhizocarpon intermediellum		O-L-149126	Norway	PV665064	PV661973	—	This study
Rhizocarpon intersitum		O-L-227971	Norway	PV665065	PV661974	—	This study
Rhizocarpon lecanorinum		McCune 36309 (OSU)	USA, Alaska	PV665067	PV661975	—	This study
		O-L-169130	Norway	PV665068	PV661976	—	This study
Rhizocarpon macrosporum		McCune 32022 (OSU)	USA, Montana	PV665069	PV661977	—	This study
Rhizocarpon norvegicum		O-L-163568	Norway	PV665070	PV661978	—	This study
Rhizocarpon parvum		O-L-225584	Norway	PV665071	—	—	This study
Rhizocarpon polycarpum		McCune 36183 (OSU)	USA, Alaska	PV665074	PV661981	PV738726	This study
		O-L-175739	Norway	PV665075	PV661982	—	This study
		O-L-227898	Norway	PV665076	PV661983	PV738727	This study
Rhizocarpon pusillum		O-L-223718	Canada	PQ219496	PV661984	PV738728	Timdal & Rui (2024)
Rhizocarpon richardii		O-L-145354	Norway	PV665078	—	—	This study
Rhizocarpon saurinum		hb Wheeler 6026 p.p	USA, Montana	PV665081	PV661987	—	This study
Rhizocarpon subgeminatum		O-L-165477	Norway	KU687452	—	—	McCune et al. (2016)
		O-L-166501	Norway	KU687457	—	—	McCune et al. (2016)
Rhizocarpon suomiense		O-L-169818	Norway	PV665082	PV661988	—	This study
		O-L-19770	Norway	AF483613	AF483181	—	Ihlen & Ekman (2002)
Rhizocarpon viridiatrum		McCune 34748 (OSU)	USA, Oregon	PV665089	PV661990	—	This study
		O-L-160701	Iceland	PV665090	—	—	This study
		O-L-183807	Norway	PV665091	—	—	This study
Rhizocarpon vorax		O-L-20249	Norway	PV665092	PV661991	—	This study

Ascospore characters, thallus colour, and secondary metabolites

The morphology in the group has been studied for decades, including by co-authors of this study, and the microscopical characters and secondary metabolites for the species concepts are documented by, e.g., Timdal (1986), Holtan-Hartwig & Timdal (1987), Timdal & Holtan-Hartwig (1988), Purvis et al. (1992), Fryday (2002), Feuerer & Timdal (2004), Ihlen (2004), McCune et al. (2016), Halici et al. (2022), Fryday et al. (2024), and Timdal et al. (2024). In this study, the material sequenced by us was identified by microscopical and chemical methods described by Timdal et al. (2024). The photographs of ascospores and microtome cross-sections of apothecia were made by a Nikon Z 6II camera attached to a Leitz Aristoplan compound microscope. Ascospore characters (i.e., pigmentation, septation, and number of septa), thallus colour, amyloidity in the medulla, and presence of secondary metabolites were mapped onto the phylogeny using ggtree (Yu et al. 2017).

DNA extraction and amplification

Total genomic DNA was extracted from 1–5 apothecia per specimen or a small thallus piece if apothecia were scarce or missing. We used two different extraction protocols: (1) the E.Z.N.A.® SP Plant DNA Kit (Omega Bio-Tek, Georgia, U.S.A) following a modified protocol (Bendiksby & Timdal 2013) and (2) the Chelex 100 protocol (Bio-Rad, Hercules, CA, USA) according to Ferencova et al. (2017); the latter selected for speed in the second period of the project. For each accession, we sequenced up to three genetic markers: the nuclear ribosomal internal transcribed spacer (ITS), the Minichromosome Maintenance Complex Component 7 (MCM7), and the mitochondrial ribosomal small subunit (mtSSU). For the amplification reactions, the Illustra™ PuReTaq Ready-To-Go™ PCR Beads (GE Healthcare, Buckinghamshire, UK) were used to amplify the markers, following the manufacturer’s protocol, except for using half reactions (i.e., 9.95 uL water, 1.25 µL MgCl2, 0.3 µL of each primer, 0.7 µL template DNA = 12.5 µL per reaction). The ITS and MCM7 loci were mainly amplified as single fragments using the primer pairs ITS1F/ITS4 (White et al. 1990, Gardes & Bruns 1993) and MCM7-709for/MCM7-1348rev (Schmitt et al. 2009), respectively. In some cases, the ITS locus was amplified as two shorter fragments using the primer pairs ITS1F/ITS2 and ITS3/ITS4 (White et al. 1990). The mtSSU locus was mostly amplified in two fragments using the primer pairs mtSSU1/mtSSU-RhiR (Zoller et al. 1999; this study: 5’-AAT AAC ATA CTT CAC TAC TGG T-3’, respectively) and mtSSU-RhiF/mtSSU3R (this study: 5’-ACC AGT AGT GA A GTA TGT TAT T-3’; Zoller et al. 1999, respectively).

DNA sequence analysis

The ITS and mtSSU markers were aligned using PASTA (Mirarab et al. 2014), as it performs well for variable markers and in cases where many gaps can be expected. The more conserved, protein coding MCM7 marker was aligned using MAFFT (algorithm E-INS-i) (Katoh et al. 2019). For both alignment methods, the same settings were used as in Svensson & Fryday (2022). For viewing the alignments, AliView was used (Larsson 2014). Parsimony informative sites were inspected in MEGA v. 11 (Tamura et al. 2021). Gene trees were inferred for all markers using RAxML-NG-MPI v. 1.0.2. (Stamatakis 2006, Kozlov et al. 2019) with 1000 bootstrap repeats (Felsenstein 1985). The three markers were also concatenated, and the concatenated alignment was used for both Bayesian inference (BI) using MrBayes v. 3.2.7a (Ronquist et al. 2012), and Maximum Likelihood (ML) using RAxML-NG-MPI v. 1.0.2. with 1000 bootstrap repeats. The MrBayes analysis ran until the average standard deviation of split frequencies (ASDSF) reached below 0.01, with four runs, four chains, and a relative burn-in of 25 %. Effective sample sizes (ESS) were examined in Tracer v. 1.7.1 (Rambaut et al. 2018). The resulting phylogenies were edited in ggtree and Inkscape (Inkscape 2023).

RESULTS

Microscopic characters and chemistry

Ascospore morphology, thallus colour and secondary metabolites are shown in the character matrix in Fig. 2. The character states shown represent what is known for the species concepts, not necessarily the individual accessions [e.g., R. hochstetteri does not always contain stictic acid, but it is the only secondary metabolite that has been detected in this species (Fryday et al. 2024)].

Fig. 2.

Best tree from the ML analysis of the concatenated alignment, branches with BS < 50 % are collapsed. Numbers above branches indicate PP/BS support from the BI/ML analyses. The character matrix shows character states known for the species concepts (not necessarily the specimen): 1: ascospore pigmentation; 2: ascospore septation; 3: ascospores per ascus; 4: lichenicolous lifestyle; 5: rusty thallus; 6: yellow pigment; 7: thallus amyloidity; 8–13: secondary metabolites known to occur in the species.

Molecular phylogeny

The ITS alignment of the 83 accessions was 717 base pairs (bp) long. Of the 357 variable sites in the ingroup 315 were parsimony informative. The MCM7 alignment was 609 bp long with 24 accessions and the ingroup contained 188 parsimony informative sites. The mtSSU alignment was 901 bp long with 56 accessions and the ingroup contained 126 parsimony informative sites. As there were no supported incongruences [a branch is here considered supported in the BI analysis if the posterior probability (PP) exceeds 0.95 and in the ML analysis if the bootstrap support (BS) exceeds 70] between the gene trees (supplementary material), ML and BI phylogenies were inferred for a concatenated alignment (2227 bp long with 88 accessions). The BI analysis converged after 2.2 million generations and all ESS values exceeded 200. All alignments and phylogenies are deposited at Figshare [doi (alignments and trees, respectively): 10.6084/m9.figshare.28596479.v2 and 10.6084/m9.figshare.28596530.v2; Möller et al. 2025a, b].

The two analyses showed no supported incongruences and display five distinct, highly supported major clades (Fig. 2): (1) Catolechia (three species), (2) Haugania (two species), (3) Poeltinula (seven species), (4) Rehmia (14 species; named so because the generic name is available for the clade), and (5) Rhizocarpon s. str. (20 species). Catolechia and Poeltinula are supported as sisters in the BI analysis (PP = 0.98). This relation is not significantly supported in the ML analysis (BS = 60). Four accessions, representing two species of Rhizocarpon, are not assigned to any of the above listed five major clades and are hereafter referred to as Rhizocarpon rest group.

Catolechia (PP = 1 / BS = 97):

Catolechia wahlenbergii and the two Epilichen species form a highly supported clade, with E. glauconigellus as sister to a genetically uniform clade consisting of C. wahlenbergii and E. scabrosus.

Haugania (PP = 1 / BS = 100):

The two species H. oederi and H. pycnocarpoides form a fully supported clade.

Poeltinula (PP = 1 / BS = 99):

The five species/taxa in the Rhizocarpon hochstetteri complex (R. caesium, R. hensseniae, R. hochstetteri, R. infernulum, and R. infernulum f. sylvaticum) and the two Poeltinula species (P. cacuminum and P. cerebrina) form a fully supported clade.

Rehmia (PP = 1 / BS = 89):

The 14 species Rhizocarpon amphibium, R. caeruleoalbum, R. chioneum, R. cinereovirens, R. furfurosum, R. haidense, R. lavatum, R. tetramerum, R. timdalii, R. petraeum, R. umbilicatum, R. reductum, R. roridulum, and R. rubescens, form a highly supported clade.

Rhizocarpon s. str. (PP = 1 / BS = 100):

The 20 species R. alpicola, R. atroflavescens, R. eupetraeum, R. dinothetes, R. distinctum, R. effiguratum, R. ferax, R. geographicum, R. intermediellum, R. lecanorinum, R. macrosporum, R. norvegicum, R. parvum, R. polycarpum, R. pusillum, R. richardii, R. saurinum, R. viridiatrum and R. vorax form a fully supported clade.

Rhizocarpon rest group (PP = 1 / BS = 94):

The two species R. subgeminatum and R. suomiense are not assigned to any of the major clades, but form a highly supported clade which is sister to Rhizocarpon s. str. with PP = 0.94 / BS = 64.

Taxonomy

The following names include all accepted genera (including synonyms) of the Rhizocarpaceae and all accepted species (excluding synonyms) with the exception of the numerous species of Rhizocarpon.

Catolechia

Flot., Bot. Zeitung (Berlin) 8: 367. 1850.

Typification: Lecidea wahlenbergii Ach., designated by Koerber, Systema Lichenum Germaniae (2): 181. 1854–1855.

Synonyms: Dimaura Norman, Conatus praemissus redactionis novae generum nonnullorum lichenum: 23. 1852.

Typification: Lecidea wahlenbergii Ach., designated by Hafellner, Beiheft zur Nova Hedwigia 79: 330. 1984.

Xanthopsis Acloque, Les Lichens: 346. 1893.

Typification: Lecidea wahlenbergii Ach.

Epilichen Clem., The Genera of Fungi: 69, 174. 1909.

Typification: Lecidea scabrosa Ach.

Catolechia glauconigella

(Nyl.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859240.

Basionym: Lecidea glauconigella Nyl., Lichenes Scandinaviae: 238. 1861.

Catolechia scabrosa

(Ach.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859241.

Basionym: Lecidea scabrosa Ach., Methodus: 48. 1803.

Catolechia stellata

(Triebel) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859242.

Basionym: Epilichen stellatus Triebel, Biblioth. Lichenol. 35: 136. 1989.

Catolechia wahlenbergii

(Ach.) Körb., Systema Lichenum Germaniae (2): 181. 1854–1855.

Basionym: Lecidea wahlenbergii Ach., Methodus: 81. 1803

Typification: «Lapponia» H9510095 (H-ACH 219D, lectotype designated here) MBT 10026483.

Haugania

E.J. Möller & Timdal in Fryday et al., Revisions of British and Irish Lichens 41: 27. 2024.

Typification: Lecidea oederi Ach.

Haugania oederi

(Ach.) E.J. Möller & Timdal in Fryday et al., Revisions of British and Irish Lichens 41: 27. 2024.

Basionym: Lecidea oederi Ach., Methodus: 49. 1803 (nom. cons.).

Haugania pycnocarpoides

(Eitner) E.J. Möller & Timdal in Fryday et al., Revisions of British and Irish Lichens 41: 27. 2024.

Basionym: Rhizocarpon pycnocarpoides [“pycnocappoides”] Eitner, Jahresber. Schles. Ges. Vaterl. Cult. 88: 46. 1911.

Poeltinula

Hafellner, Nova Hedwigia Beih. 79: 330. 1984.

Basionym: Melanospora Mudd, A Manual of British Lichens: 226. 1861 (nom. illegit., Art. 53.1), non Melanospora Corda, Icones Fungorum Tomus 1: 24. 1837.

Typification: Opegrapha cerebrina DC.

Poeltinula cacuminum

(Asta, Clauzade & Cl.Roux) Clauzade & Cl. Roux, Bull. Soc. Bot. Centre-Ouest. 7: 827. 1985

Basionym: Encephalographa cerebrina subsp. cacuminum Asta, Clauzade & Cl.Roux, Bull. Soc. Linn. Provence 30: 11. 1977.

Poeltinula caesia

(Fryday) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859243.

Basionym: Rhizocarpon caesium Fryday, Lichenologist 34: 458. 2002.

Poeltinula cerebrina

(DC.) Hafellner, Nova Hedwigia Beih. 79: 330. 1984.

Basionym: Opegrapha cerebrina DC. in Lamarck & de Candolle, Flore Française. ed. 3, 2: 312. 1805.

Poeltinula cerebrinella

(Nyl.) Øvstedal, S. African J. Bot. 67: 563. 2001.

Basionym: Lecidea cerebrinella Nyl., J. Bot. 14: 22. 1876.

Poeltinula hensseniae

(Brodo) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859244.

Basionym: Rhizocarpon hensseniae Brodo, Biblioth. Lichenol. 38: 32. 1990.

Poeltinula hochstetteri

(Körb.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859245.

Basionym: Catillaria hochstetteri Körb., Parerga Lichenologica: 195. 1861.

Poeltinula infernula

(Nyl.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859246.

Basionym: Lecidea infernula Nyl., Flora 68: 440. 1885.

Poeltinula sylvatica

(Fryday) E.J. Möller, Timdal, Haugan & Bendiksby, comb. et stat. nov. MB 859247.

Basionym: Rhizocarpon infernulum f. sylvaticum Fryday, Lichenologist 34: 468. 2002.

Rehmia

Kremp., Denkschr. Königl.-Baier. Bot. Ges. Regensburg 4, 2: 211. 1861.

Typification: Rehmia caeruleoalba Kremp.

Rehmia amphibia

(Fr.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859248.

Basionym: Lecidea amphibia Fr., Kongl. Vetensk. Acad. Handl. 1822: 262. 1822.

Rehmia anapera

(Vain.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859249.

Basionym: Lecidea anapera Vain., Meddeland. Soc. Fauna Fl. Fenn. 10: 141. 1883.

Rehmia caeruleoalba

Kremp., Denkschr. Königl.-Baier. Bot. Ges. Regensburg 4, 2: 211. 1861.

Rehmia chionea

(Norman) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859250.

Basionym: Catillaria chionea Norman, Kongel. Norske Videnskabers Selsk. Skr. 19de Aarhundr. 5: 355. 1868.

Rehmia cinereovirens

(Müll. Arg.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859251.

Basionym: Patellaria cinereovirens Müll. Arg., Flora 51: 49. 1868.

Rehmia furfurosa

(H. Magn. & Poelt) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859252.

Basionym: Rhizocarpon furfurosum H. Magn. & Poelt, Verh. Zool.-Bot. Ges. Wien 95: 110. 1955.

Rehmia haidensis

(Brodo & Fryday) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859253.

Basionym: Rhizocarpon haidense Brodo & Fryday, Lichenologist 52: 116. 2020.

Rehmia lavata

(Fr.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859254.

Basionym: Lecidea atroalba var. lavata Fr., Nova Schedulae Criticae de Lichenius Suecanis: 18. 1827.

Synonym: Lecidea lavata (Fr.) Nyl., Flora 56: 23. 1873.

Rehmia petraea

(Wulfen) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859255.

Basionym: Lichen petraeus Wulfen, Schriften Ges. Naturf. Freunde Berlin 8: 89. 1787.

Rehmia postuma

(Nyl.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859256.

Basionym: Lecidea postuma Nyl., Flora 51: 345. 1868.

Rehmia reducta

(Th. Fr.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859257.

Basionym: Rhizocarpon reductum Th. Fr., Lichenographia Scandinavica Pars prima (2): 633. 1874.

Rehmia roridula

(Th. Fr.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859258.

Basionym: Rhizocarpon obscuratum ssp. roridulum Th. Fr., Lichenographia Scandinavica Pars prima (2): 629. 1874.

Synonym: Lecidea roridula (Th. Fr.) Stizenb., Ber. Thätigk. St. Gallischen Naturwiss. Ges. 1874–1875: 235. 1876.

Rehmia rubescens

(Th. Fr.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859259.

Basionym: Rhizocarpon rubescens Th. Fr., Lichenographia Scandinavica Pars prima (2): 631. 1874.

Typification: Sweden, Uppland, Lyran prope Stockholm, 1870, S. Almquist (UPS L-107090, lectotype designated here) MBT 10026484.

Rehmia subpostuma

(Arnold) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859260.

Basionym: Rhizocarpon subpostumum Arnold, Verh. K. K. Zool.-Bot. Ges. Wien. 27: 554. 1877.

Rehmia tetramera

(Vain.) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859261.

Basionym: Lecidea postuma var. tetramera Vain., Meddeland. Soc. Fauna Fl. Fenn. 10: 141. 1883.

Synonym: Rhizocarpon tetramerum (Vain.) Vain., Acta Soc. Fauna Fl. Fenn. 51, 1: 316. 1922.

Rehmia timdalii

(Ihlen & Fryday) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859262.

Basionym: Rhizocarpon timdalii Ihlen & Fryday, Lichenologist 34: 96. 2002.

Rehmia umbilicata

(Ramond) E.J. Möller, Timdal, Haugan & Bendiksby, comb. nov. MB 859263.

Basionym: Lecidea umbilicata Ramond, Mém. Acad. Sci. Inst. France 6: 128. 1827.

Rhizocarpon

Ramond ex DC., Flore française, Ed. 3, 2: 365. 1805.

Typification: Lichen geographicus L., designated by Fink, Contr. U. S. Natl. Herb. 14: 97. 1910.

Lepidoma Link, Neues J. Bot. 3: 5. 1809.

Typification: Lichen geographicus L.

Catocarpus (Körb.) Arnold, Flora (Regensburg) 54: 147. 1871.

Basionym: Buellia sect. Catocarpus Koerb. Systema Lichenum Germaniae: 223. 1855.

Typification: Lecidea badioatra Flörke ex Schaer. (type designated here, MBT 10026487).

Notes: The genus is currently regarded as typified by Dodge (1973: 88). Dodge (1973), however, merely states (erroneously) that the genus was based on Lecidea badioatra Flörke. There are four species in the protologue (Buellia badioatra “Flörke”, B. ocellata “Flörke”, B. coracina “Hoffm.” and Catolechia lactea A. Massal.), and we designate the first as the type in accordance with how the name is usually applied. The typification of Lecidea badioatra Flörke ex Schaer. [i.e., the name that we think is the basionym of Buellia badioatra “Flörke”] was treated in Timdal et al. (2024).

Diphaeis Clem., The Genera of Fungi: 77, 174. 1909.

Typification: Lecidea badioatra Flörke ex Schaer.

Diphanis Clem., The Genera of Fungi: 77, 174. 1909.

Typification: Lecidea confervoides var. polycarpa Hepp.

Catillariopsis (Stein) M. Choisy, Bull. Mens. Soc. Linn. Lyon. 19: 158. 1950.

Basionym: Catocarpus sect. Catillariopsis Stein, Kryptogamen-Flora von Schlesien Zweiter Band, zweite Häfte, Flechten: 225. 1879.

Typification: Lecidea confervoides var. polycarpa Hepp, designated by Hafellner, Beiheft zur Nova Hedwigia 79: 329. 1984.

Rhizocarponomyces Cif. & Tomas., Atti Ist. Bot. Lab. Crittog. Univ. Pavia 10: 38, 65. 1953 (nom. illegit., Art. 52.1).

Typification: The type of Rhizocarpon DC. (Art. 7.5).

Excluded genera

The following genera are listed as synonyms of Rhizocarpon in MycoBank (2025), but are here excluded from the Rhizocarpaceae.

Cormothecium

A. Massal., Geneacaena Lichenum noviter proposita ac descripta: 8. 1854.

Typification: Lecidea duebenii Fr.

Note: Lecidea duebenii is currently regarded as a synonym of Monerolechia badia (Fr.) Kalb (Caliciaceae) [e.g., Nimis et al. 2018, as Buellia duebenii (Fr.) Hellb.]. Cormothecium actually predates Monerolechia Trev. (priority 1857).

Phalodictyum

Clem., The Genera of Fungi: 77, 174. 1909.

Typification: Lecidea petraea var. obscurata Ach.

Note: The type [syn. Rhizocarpon obscuratum (Ach.) A. Massal.] is a synonym of Fuscidea lygaea (W. Mann) V. Wirth & Vězda (Fusideaceae) (lectotypified by Ihlen 2004: 556). Phalodictyum actually predates Fuscidea V. Wirth & Vězda (priority 1972).

Siegertia

Körb., Parerga Lichenologica (2): 180. 1860.

Typification: Lichen calcareus L.

Notes: Although the protologue apparently describes the genus here named Rehmia, the only included species was named “S. calcarea (Weis) Kbr.” with reference to the species described in Koerber (1855: 220) as “Diplotomma calcarea Weis”. This can only be Lichen calcarius in Weis (1770) which refers (indirectly, through phrase name) to Lichen calcareus L. That name is the basionym of the species currently named Circinaria calcarea (L.) A. Nordin, Savić & Tibell (Megasporaceae). Siegertia antedates Circinaria Link (priority 1809).

DISCUSSION

Our phylogenetic results show that Rhizocarpon, as currently circumscribed, is paraphyletic with Catolechia, Epilichen, and Poeltinula nested within it, corroborating the findings of prior studies with a more limited taxon sampling (Ihlen & Ekman 2002, Miadlikowska et al. 2014). Our integrated assessment of molecular, anatomical and chemical evidence in a molecular phylogenetic framework suggests that the family consists of at least five major lineages. These five clades do not reflect current generic circumscriptions.

One solution is to synonymize Catolechia, Epilichen, Poeltinula, and Haugania with Rhizocarpon. However, this would have rendered Rhizocarpon an even larger genus with few unifying characters and the only genus in the Rhizocarpales. Under the current classification, Rhizocarpon species are characterized by being saxicolous and having a crustose thallus with apothecia originating from the hypothallus, in addition to having 1- to pluriseptate ascospores with perispores. Catolechia and Epilichen species are terricolous or lichenicolous on terricolous lichens and do not have perispores. Poeltinula species barely form visible thalli. Synonymizing all these genera with Rhizocarpon would result in a quite large genus (150–200 species) being the single genus in the family or, possibly, order. The only common character would be the ascus anatomy (Hafellner 1984).

Here, we propose to instead accept the five major genetic lineages as distinct genera (Catolechia, Haugania, Poeltinula, Rehmia, and Rhizocarpon), distinguishable by sets of traits (i.e., septation and pigmentation of ascospores, number of ascospores per ascus, hymenial characters, thallus colour, amyloidity, chemistry and ecology). In the following discussion we refer to the species under the old generic names, even though the new combinations were already made above.

Catolechia:

Morphologically and ecologically, C. wahlenbergii and the two Epilichen species are quite different: C. wahlenbergii is autonomous, up to 10 cm in diameter, and grows almost exclusively in shaded crevices in more or less humid, rock walls, while the Epilichen species are obligate parasites on other lichens, rarely more than 1 cm in diameter. However, species in this clade are similar in microscopic characters, unique in the family, by having ascospores without a gelatinous perispore. Catolechia wahlenbergii and E. scabrosus are both yellow due to two yellow pigments, apparently pulvinic acid derivatives that are not rhizocarpic acid. Also, the clade is unique in the family in being terricolous or lichenicolous on other terricolous lichens.

We include sequences from four specimens of C. wahlenbergii (all three markers for all four specimens) and four specimens of E. scabrosus (three ITS sequences, two MCM7 sequences, and two mtSSU sequences). Curiously, C. wahlenbergii and E. scabrosus are nested with each other on all three markers (see gene trees, supplementary material), and consequently in the species tree. Normally, the microscopical, chemical and genetic evidence provided here would merit synonymization. Due to the large morphological and ecological differences, the lack of intermediates, and the fact that we use only three markers, we do not synonymize the two species here. Further investigations are needed to confidently revise this species complex. Regardless of whether the two species are conspecific, there is no doubt that Epilichen should be synonymized with Catolechia.

Haugania:

Haugania oederi and H. pycnocarpoides form a fully supported clade. The species are similar in having eight, halonate, hyaline ascospores, rust red thalli, and by growing exclusively on heavy metal rich rocks. These characters are admittedly not unique within the family and hardly of high taxonomic value, but, as a combination, a key character for the genus. Haugania is here accepted based on the phylogenetic reconstruction.

Poeltinula:

The species in the R. hochstetteri complex are characterized by being crustose, non-yellow, having a distinct apothecial margin, a non-amyloid medulla, swollen and sharply delimited paraphysis caps, and 1-septate, halonate, hyaline ascospores (Fig. 3). Together with the two included Poeltinula species, which have an endolithic thallus, furrowed apothecia, green, 1-septate ascospores, and pigmented, albeit substantially smaller, paraphysis caps, the complex constitutes a fully supported clade. Species in the clade either have stictic acid or no secondary metabolites.

Fig. 3.

Poeltinula anatomy: Transverse sections through apothecia (A, D, G), tip of paraphyses (B, E, H), and ascospores (C, F, I). A–C. Rhizocarpon hochstetteri, Norway, O-L-227846. D–F. R. infernulum f. sylvatica, Norway, O-L-228034. G–I. P. cacuminum, Austria, Wilfling & Möslinger 569, GZU. Photo credits: E. Timdal. Scale bars: A, D, G = 100 µm; B, C, E, F, H, I = 10 µm.

Runemark (1956) reports observing pigmented paraphysis caps in some yellow species, but his description of the caps differs from the ones observed in Poeltinula and the R. hochstetteri complex by being more diffuse and not as sharply delimited. Furthermore, hyaline, 1-septate ascospores also occur in R. polycarpum and R. richardii, but those species have a strongly amyloid medulla. Rehmia caeruleoalba, R. chionea, R. cinereovirens and R. haidensis, also have similar ascospores, but lack the distinct apothecial margin and the swollen pigmented paraphysis caps.

Poeltinula is the only available genus name for this group, and we propose to include Rhizocarpon hochstetteri, R. caesium, R. infernulum and R. hensseniae in that genus. Additionally, we accept R. infernulum f. sylvaticum as a distinct species, since the branch lengths in all gene trees are equivalent to the accepted species in the group.

Rehmia:

The 14 species that constitute the Rehmia clade are similar in having a non-amyloid medulla and halonate, hyaline, mainly muriform ascospores (some have 1-septate ascospores and those of Rhizocarpon tetramerum are 3-septate ascospores) (Fig. 4). The four species that have 1-septate ascospores can be distinguished from Poeltinula by lacking an apothecial margin and swollen paraphysis caps. In addition, R. caeruleoalbum and R. chioneum have white thalli similar to R. umbilicatum. Hyaline, muriform ascospores also occur in species outside of Rehmia: Rhizocarpon subgeminatum and R. suomiense both have less than eight ascospores per ascus, whereas all Rehmia species have eight, and Rhizocarpon distinctum, which is retained in Rhizocarpon, has a strongly amyloid medulla, whereas all Rehmia species have a non-amyloid medulla.

Fig. 4.

Rehmia anatomy: Transverse sections through apothecia (A, C, E) and ascospores (B, D, F). A, B. Rhizocarpon amphibium, Norway, O-L-163528. C, D. R. caeruleoalbum, Norway, UPS L-67821. E, F. R. lavatum, Norway, O-L-28923. Photo credits: E. Timdal. Scale bars: A, C, E = 100 µm; B, D, F = 10 µm.

The clade here named Rehmia largely corresponds to the so-called Rhizocarpon obscuratum group (i.e., non-yellow species with hyaline, muriform ascospores) studied by Ihlen (2004). Of the 16 species placed in the group by Ihlen (2004), we include 12 in Rehmia (nine from molecular evidence, three from morphology) and exclude four (three from molecular evidence, i.e., Rhizocarpon distinctum, R. subgeminatum, and R. suomiense) and one from preliminary, incomplete molecular data indicating distant relationship (R. sublavatum; Möller 2021). Our molecular evidence clearly shows that ascospore morphology alone is not diagnostic for the circumscription of Rehmia, but that amyloidity and number of ascospores per ascus must also be taken into account. This is emphasized by the fact that Rhizocarpon caeruleoalbum, R. cinereovirens, R. chioneum and R. haidense, all with 1-septate ascospores and hence not included in the R. obscuratum group, belong in the Rehmia clade.

The clade is phylogenetically well separated from the rest of the family, and is anatomically distinguishable by the traits mentioned above. We propose accepting it as a distinct genus and resurrect the name Rehmia. The type species of Rehmia is Rhizocarpon caeruleoalbum, in our tree represented by three specimens collected in North America, Norway and Austria. In addition to the fourteen species in the Rehmia clade, we also transfer three non-sequenced species to Rehmia based on the morphological features given above.

Rhizocarpon s. str.:

The clade that contains the type species of the genus (Rhizocarpn geographicum) is a diverse group and has few traits that unite them. However, there are trends, albeit with many exceptions. All the species of the Rhizocarpaceae with an amyloid medulla belong here, as do all the species with secondary compounds other than barbatic, stictic, and norstictic acids (i.e. confriesiic, diffractaic, gyrophoric, psoromic, and rhizocarpic acids). The only Rhizocarpaceae species in our phylogeny known to contain barbatic acid outside Rhizocarpon s. str. is R. subgeminatum of the Rhizocarpon rest group. Most of the species have pigmented ascospores, but they vary in number of septa. All of the lichenicolous species except the two Epilichen species also belong here. It is likely that the group is even more diverse than shown here, since this study did not include all known Rhizocarpon species.

Rhizocarpon rest group:

Rhizocarpon subgeminatum and R. suomiense are morphologically and anatomically similar to Rehmia species, but are easily distinguishable by the number of ascospores per ascus (all Rehmia species have eight ascospores per ascus, while Rhizocarpon subgeminatum and R. suomiense have two). They were included in the study because they have hyaline ascospores, and could therefore be suspected to belong to either the Rehmia or the Poeltinula clade. Here, we show that they do not appear in either of those clades, but rather form a well-supported, distinct clade with uncertain placement in the presented phylogeny. We do not formally describe the clade as a genus because we do not consider evidence from so few markers for only four accessions (ITS and mtSSU for R. suomiense and only ITS for R. subgeminatum) to be sufficient for describing these two species as a distinct genus. We know from preliminary analyses that the group adjacent to what we here refer to as Rhizocarpon s. str. requires more data to be properly treated.. We have ITS sequences for ca 80 accepted species of Rhizocarpon, but here we excluded many species and groups that we know from preliminary analyses do not belong to any of the groups we treat taxonomically. These species include putatively monophyletic groups, like the R. geminatum complex (i.e. R. geminatum, R. disporum, and R. sulphurosum) and the R. badioatrum complex (Timdal et al. 2024), and several more species that so far form a polytomy close to Rhizocarpon s. str. (e.g. R. inarense, R. jemtlandicum, R. leptolepis, and R. superficiale) (Möller 2021). To our best knowledge, we included all accepted species that do not belong in or are closely related to Rhizocarpon s. str., with the exception of R. sublavatum, which has an uncertain placement based on preliminary studies (Möller 2021). All of these species would be interesting to analyse further with a more complete dataset.

Outgroup:

In the previous genus level study of the Rhizocarpaceae, the authors disclose that finding a suitable outgroup for the family is challenging due the uncertain placement of the Rhizocarpaceae (Ihlen & Ekman 2002). At the time, the family was hypothesized to belong to the Lecanorales, but has since been described as an order itself, possibly together with the Sportastatiaceae (e.g. Bendiksby & Timdal 2013, Miadlikowska et al. 2014, Kraichak et al. 2018). In this study, we had similar issues with outgroup selection as Ihlen & Ekman (2002). In the end, Lecanora cavicola and L. coronulans were selected as an outgroup based on having all the three markers publicly available for both accessions. It should be noted that we analysed the dataset multiple times with different outgroups, or without outgroups, and the same five major clades appear. For future studies that intend to investigate the deep topology of the family, we recommend acquiring additional conserved markers for e.g. the Sporastatiaceae. With the advent of phylogenomics, the answer to the placement of the Rhizocarpaceae within the Lecanoromycetes may be closer.

CONCLUSION

The molecular results clearly show that the taxonomy of the Rhizocarpaceae needs revision. The current circumscription is largely based on morphology and life-strategy, and renders Rhizocarpon paraphyletic. Our phylogeny indicates that microscopical traits (e.g. spore morphology and hymenial characters) are more informative.

We have presented evidence for making nomenclatural changes in the clades Catolechia, Poeltinula and Rehmia: specifically, we propose to synonymize Epilichen with Catolechia, to move the species in the Rhizocarpon hochstetteri complex to Poeltinula, and to resurrect the genus Rehmia for a set of species previously placed in Rhizocarpon.

Conflict of Interest:

The authors declare no conflicts of interest.

Supplementary Material: http://fuse-journal.org

Fig. S1

Best tree from the ML analysis of the ITS alignment, branches with BS < 50 % are collapsed. Numbers above branches indicate BS support.

Fig. S2

Best tree from the ML analysis of the mtSSU alignment, branches with BS < 50 % are collapsed. Numbers above branches indicate BS support.

Fig. S3

Best tree from the ML analysis of the MCM7 alignment, branches with BS < 50 % are collapsed. Numbers above branches indicate BS support.