An integrative study of Pannaria hookeri (Ascomycota lichenized) from Argentina and the update of the taxon circumscription based on specimens from the Northern and Southern Hemispheres

ABSTRACT

Pannaria hookeri (Borrer ex Sm.) Nyl. is reported as a bipolar cyanolichen that occurs in more than twenty countries in both Hemispheres. In the Northern Hemisphere, a minor variation is attributed to the species, whereas larger morphological and chemical variations have been reported in specimens from the Southern Hemisphere. Six P. hookeri DNA sequences are available in GenBank, and only one (from Australia) is associated with a published morphological description. In this study, we performed a preliminary phylogenetic analysis including seven nuITS and mtSSU sequences from eight specimens collected in south Argentina. In addition, we provide an update of the taxon circumscription based on 50 specimens from 13 countries. Here, the molecular analyses revealed a well-supported monophyletic group formed by the P. hookeri sequences from Argentina, Australia, Norway and USA (Alaska). Regarding the morphological and anatomical characteristics, the Southern Hemisphere specimens did not present the historical reported variations, resulting in the proposition of a concise species description that accomplishes most of P. hookeri known geographical distribution.

Keywords:
species circumscription; cyanolichen; lichenized fungi; nuITS; taxonomy; Tierra del Fuego

Introduction

Pannaria Delise ex Bory is one of the largest genera within Pannariaceae (lichenized Ascomycota), in which the lichen-forming fungi can be associated with different photobiont partners (Ekman et al. 2014; Magain & Sérusiaux 2014). Usually, the thalli are bipartite and the mycobiont is associated with Nostoc cyanobacteria. Still, tripartite thalli are also found within the genus where the mycobiont is associated with green algae, as the main photobiont, and the cyanobacteria are confined in structures called cephalodia, for example in Pannaria durietzii (P. James & Henssen) Elvebakk & D.J. Galloway (Elvebakk et al. 2008). The Pannaria species are characterized by squamulose or foliose thalli, apothecia with thalline margins, an amyloid hymenium (I+ blue), asci without internal amyloid apical structures, secondary metabolism with pannarin and related substances, and occurrence in tropical, subtropical, temperate and polar regions from both Hemispheres (Ekman et al. 2014).

Pannaria hookeri (Borrer ex Sm.) Nyl. is a species with bipolar distribution (Jørgensen 1978; Kantvilas & Gueidan 2018), which has already been reported from Austria, Canada, Faeroes Island, Finland, France, Greenland, Iceland, Ireland, Italy, Norway, Russia, Sweden, Switzerland, and U.S.A. in the Northern Hemisphere (Jørgensen 1978; 2000b; 2003; 2007; Thomson 1984); as well as from the Antarctic Peninsula, Argentina, Australia, Chile, New Zealand, and for the subantarctic islands and archipelagos Bouvetøya, Kerguelen, Marion, South Georgia, South Orkney, and South Shetlands in the Southern Hemisphere (Lindsay 1974; 1976; Redón 1985; Jørgensen 1978; 1986; 2000a; 2007; Olech 1989; 2001; Øvstedal & Lewis Smith 2001; Calvelo & Fryday 2006; Kantvilas & Gueidan 2018; Passo et al. 2020). In addition, P. hookeri was recorded at high altitudes in Ecuador (5050 m. a.s.l., Z. Palice 4598; Jørgensen & Palice 2010) and points from Kenya (4430 m a.s.l. and at 4640 m a.s.l.; Frisch & Hertel 1998). The species was originally described from Scotland (Smith & Sowerby 1811), and its first record in the Southern Hemisphere was made from a South African specimen, initially identified as P. leucolepis (Wahlenb.) Nyl. (Stizenberger 1890; Doidge 1950) and posteriorly included in the synonym list of P. hookeri (Jørgensen 2006). Nevertheless, there is no consensus about the identification of the South African material, and also from others localities from Southern Hemisphere, which can also be an extreme form of P. rubiginosa (Thunb.) Delise (Jørgensen 2003).

Regarding the morphological descriptions, a minor variation is reported to P. hookeri from Europe (Jørgensen 1978), while a larger variation has been described to P. hookeri from the Southern Hemisphere (Jørgensen 2000a). In the same way, the chemical profile differs according to the region studied. Traces of atranorin and ergosterol peroxide was found in specimens from Robert Island (Quilhot et al. 1989), the absence of secondary substances to Bouvetøya specimens (Jørgensen 1986), and pannarin or related substances were reported to specimens from Argentina (Passo et al. 2020), Australia (Kantvilas & Gueidan 2018), and Europe (Jørgensen 1978; 2007), inferring cryptic lineages and taxonomical problems in the morphological and chemical studies.

Despite the observed inconsistencies, the wide geographical distribution of P. hookeri was confirmed when the sequences of the nuITS and mtSSU regions were generated from an Australian specimen and clustered with P. hookeri sequences from Norway (Kantvilas & Gueidan 2018). Nowadays, only six specimens have available sequences in the GenBank: one from Australia, three from Norway, one from USA (Alaska) and one of unknown origin (Ekman & Jørgensen 2002; Spribille & Muggia 2013; Kantvilas & Gueidan 2018; Park et al. 2018; Marthinsen et al. 2019). Therefore, better knowledge is still needed for the circumscription characters that belong to P. hookeri, including genetic analysis with Southern Hemisphere specimens.

The aim of this study was to a) perform a phylogenetic analysis of P. hookeri from Argentina and sequences available on GenBank, generating the first molecular sequences from the South America; and b) to update the species circumscription based on specimens from the Northern (including topotypes) and Southern Hemispheres, establishing a unified morphological description.

Material and methods

Sampling

Fresh samples were collected in the National Park of Tierra del Fuego, Ushuaia, extreme south of the Argentina, during the summer of 2016-2017 (Proyecto Número 105-CPA-2016, supported by CONICET and PROANTAR), and specimens were found on rock and saxicolous mosses.

Phylogenetic analysis

Eight fresh-collected specimens were used in the molecular analysis. DNA extraction, amplification, and sequencing of the nuITS (nuclear internal transcribed spacer) and mtSSU (mitochondrial small subunit) regions were performed according to Kitaura et al. (2018). However, the specimens requested as loans have more than 10 years, and the DNA extraction was not performed. In addition, four sequences of P. hookeri of each nuITS and mtSSU regions were retrieved from GenBank. These sequences were generated from Australian, Norwegian, U.S.A., and unknown origin specimens (Tab. 1).

Table 1
Dataset used in the present study for the phylogenetic analysis. Species, geographical origin, voucher information, nuITS and mtSSU GenBank accession codes and references. Sequences generated in this study are marked in bold.

Species	Origin	Voucher	GenBank Accession numbers		Reference
			nuITS	mtSSU
Pannaria andina	Chile	Elvebakk 06-245	GQ927268	-	(Elvebakk et al. 2010)
Pannaria athroophylla	Argentina	Passo 181	EU885295	EU885317	(Passo et al. 2008)
Pannaria athroophylla	Argentina	Passo 251	EU885303	EU885325	(Passo et al. 2008)
Pannaria calophylla	Argentina	Passo 101	EU885296	EU885318	(Passo et al. 2008)
Pannaria conoplea	Norway	Ekman 3188	AF429281	-	(Ekman & Jørgensen 2002)
Pannaria contorta	Argentina	Passo 142	EU885297	EU885319	(Passo et al. 2008)
Pannaria farinosa	Argentina	Passo 119	EU885299	EU885321	(Passo et al. 2008)
Pannaria hookeri	Australia	GK102/16	MG786563	MG792317	(Kantvilas & Gueidan 2018)
Pannaria hookeri	Norway	Jørgensen s.n.	AF429282	KC608083	(Ekman & Jørgensen 2002)
Pannaria hookeri	Norway	NK-325	KY350562	-	(Park et al. 2018)
Pannaria hookeri	Norway, NO, Nordland	JT Klepsland	MK812291	-	(Marthinsen et al. 2019)
Pannaria hookeri	USA, Alaska	Spribille 29292 (KLGO)	-	JX464134	(Spribille & Muggia 2013)
Pannaria hookeri	Unknown	Jørgensen s.n. (BG)	-	KC608083	Unpublished
Pannaria hookeri	Argentina	MJK3314	MT755913	MN63424	This study
Pannaria hookeri	Argentina	MJK3319	MT755912	MN634242	This study
Pannaria hookeri	Argentina	MJK3323	MT755911	MN634243	This study
Pannaria hookeri	Argentina	MJK4022	-	MN634244	This study
Pannaria hookeri	Argentina	MJK4028	MT755910	MN634245	This study
Pannaria hookeri	Argentina	MJK4036	MT755909	MN634246	This study
Pannaria hookeri	Argentina	MJK4041	MT755908	MN634247	This study
Pannaria hookeri	Argentina	MJK4045	MT755907	-	This study
Pannaria immixta	Unknown	Elvebakk 02-352b	-	KC608084	(Kantvilas & Gueidan 2018)
Psoroma implexa	Argentina	Passo 84	-	EU885333	(Passo et al. 2008)
Pannaria insularis	Japan	Kashiwadani 43760	KC618716	KC608085	(Kantvilas & Gueidan 2018)
Pannaria leucosticta	Unknown	Hur 041227	EU266107	-	(Kantvilas & Gueidan 2018)
Pannaria lurida subsp. lurida	Unknown	Kashiwadani 43861	-	KC608086	(Kantvilas & Gueidan 2018)
Pannaria lurida subsp. russellii	Unknown	Tønsberg 22565	-	KC608087	(Kantvilas & Gueidan 2018)
Pannaria microphyllizans	Argentina	Passo 264	EU885300	EU885322	(Passo et al. 2008)
Pannaria multifida	Unknown	Schumm & Frahm s.n.	KC618717	KC608088	(Kantvilas & Gueidan 2018)
Pannaria pallida	Argentina	Passo 249	EU885301	EU885323	(Passo et al. 2008)
Pannaria rubiginella	Canada	Thor 10050	-	GQ259037	(Wedin et al. 2009)
Pannaria rubiginella	Unknown	Tønsberg 32508	KC618718	KC608089	(Kantvilas & Gueidan 2018)
Pannaria rubiginosa	Unknown	Anonby 870/Purvis s.n.	KC618717	AY340513	(Kantvilas & Gueidan 2018)
Pannaria sphinctrina	Argentina	Passo 221	EU885302	EU885324	(Passo et al. 2008)
Pannaria subfusca	Unknown	Tønsberg 33592	KC618719	-	(Kantvilas & Gueidan 2018)
Pannaria tavaresii	Argentina	Passo 122	EU885294	EU885316	(Passo et al. 2008)
Pannaria tavaresii	Unknown	Schumm s.n.	KC618720	-	(Kantvilas & Gueidan 2018)
Staurolemma oculatum	China	Aptroot 55941	KC618738	GQ259045	(Kantvilas & Gueidan 2018)
Staurolemma omphalarioides	Turkey	Tibell s.n.	KJ533487	KJ533439	(Bendiksby et al. 2014)

Sequences of Staurolemma oculatum P.M. Jørg. & Aptroot (GenBank accession numbers KC618738 and GQ259045) and S. omphalarioides (Anzi) P.M. Jørg. & Henssen (KJ533487 and KJ533439) were used as outgroups according to Kantvilas & Gueidan (2018) (Tab. 1).

The alignments were performed in Geneious v9.1.2 (Kearse et al. 2012) with the MAFFT v7.308 algorithm (Katoh et al. 2002) using default settings, and the Gblocks web server (http://molevol.cmima.csic.es/castresana/Gblocks_server.html) was used to exclude unreliable aligned sites using the less stringent options. The alignments were used to infer trees from nuITS, mtSSU, and concatenated regions. Phylogenetic trees were estimated using the Bayesian (BA) and Maximum Likelihood (ML) approaches, and analyses for each region separately and for a combined data matrix were performed. The nucleotide substitution and site heterogeneity models were inferred following the Bayesian Inference Criterion in jModelTest2 on XSEDE, CIPRES Science Gateway V.3.3 (Guindon & Gascuel 2003; Miller et al. 2010; Darriba et al. 2012). The BA was performed in MrBayes v.3.2.7a on XSEDE, available in the CIPRES Science Gateway V.3.3 (Huelsenbeck & Ronquist 2001; Ronquist & Huelsenbeck 2003; Miller et al. 2010) using TrNef+I+G as substitution model for nuITS, TPM2uf+I+G for mtSSU regions and TIM1+I+G for the concatenated dataset, and their respective base frequencies were settled. The settings were: two independent runs with four chains (length of 10,000,000 generations), with trees sampled every 1,000 steps. The first 25 % of the generated trees were discarded as burn-in, and a 50 %-majority-rule tree was built. The ML trees were built with the RaxML-HPC2 v.8.2.12 on XSEDE available in the CIPRES Science Gateway V.3.3 (Miller et al. 2010; Stamatakis 2014), using GTR+G+I as nucleotide substitution model and 1,000 bootstrap replicates. FigTree v1.4.2 (http://tree.bio.ed.ac.uk/software/figtree/) was used to edit the trees. Branches with posterior probability values above 0.95 (BA) and bootstrap above 75 (ML) were considered significantly supported.

Morphological and anatomical studies

The AAS, C, COLO, E, H, M, O, and S herbaria sent 35 specimens from the Northern Hemisphere and seven from the Southern Hemisphere. The specimens received as loan were morphologically examined, and the determinations were confirmed (Text S1), when compared with eight fresh P. hookeri specimens.

The P. hookeri description was built according to Jørgensen (1978). Then, our description was compared with other available in the literature (Lindsay 1974; Redón 1985; Jørgensen 1986, 2007; Øvstedal & Lewis Smith 2001; Kantvilas & Gueidan 2018; Passo et al. 2020), and the variable characters were highlighted in Table 2 and discussed here. The specimens were tested chemically by applying PD spot tests, and orange reactions were interpreted as indicating the presence of pannarin. Therefore, TLC and HPLC analyses are necessary to confirm the substances.

Results

Molecular analysis

The dataset of the present study was composed of sequences of the nuITS and mtSSU regions generated from eight specimens of Pannaria hookeri collected in southern Argentina. Furthermore, six P. hookeri sequences retrieved from GenBank composed the dataset together with other Pannaria species sequences and the selected outgroups (Tab. 1). The final dataset comprised 31 sequences and an alignment 1105 base long, with 423 of the nuITS and 682 of the mtSSU regions.

Trees of both BA and ML approaches, among genes and concatenated datasets, showed congruent topology, so only the concatenated tree is presented, with the values of the posterior probabilities and bootstrap values shown (Fig. 1). The sequences of the specimens identified as P. hookeri, newly produced and those obtained from GenBank, presented 0.89/95 of posterior probability and bootstrap values, respectively, grouping the specimens from Argentina, Australia, Norway, and the USA in a well-supported monophyletic group. The divergence in the nuITS region, considered the universal barcode DNA of fungi (Schoch et al. 2012), revealed a low variation, with the mean percentage of identity of 99.34 % (98.25 % to 100 %) among the sequences, without deeply divergent populations or cryptic lineages.

Figure 1
Bayesian Maximum Clade Credibility tree based on nuITS and mtSSU datasets showing the phylogenetic relationships within the Pannaria and P. hookeri sequences from Argentina, Australia, Norway and USA (Alaska). Posterior probabilities and Bootstrap values are above branches, respectively. Bold branches have posterior probabilities (PP) >0.95 % or bootstrap values >75. Sequences obtained in this study are marked in bold. Text after species names corresponds to voucher information (Tab. 1).

The morphological descriptions of Pannaria hookeri from the literature were gathered and used for the circumscription of the species and for the observation of inconsistencies in the species descriptions and identifications, with the differences found highlighted in the sections "Description of Species" and "Discussion" below (Tab. 2).

Table 2
Morphological descriptions of Pannaria hookeri reported in the literature and in the present study.

	Lindsay (1974 )	Redón (1985 )	Jørgensen (1986 )	Jørgensen (2000 a)	Øvstedal & Lewis Smith (2001 )	Jørgensen (2007 )	Kantvilas & Gueidan (2018 )	Passo et al. (2020 )	This study
Upper surface color	Greenish grey, streaked with white towards tips of squamules	Greenish grey	Brownish, white-grey striated and with distinctly white-grey margins which are sometimes blackened	Brownish grey, but dominated by the whitish margins and the laminal white striae	Grey to brownish, also to Pannaria caespitosa P.M. Jørg	Grey-brown though whitish striate	Pale brownish grey to smoky bluish grey	Bluish grey to brownish	Whitish beige to brownish-grey
Branches	Not mentioned	Up to 3 mm broad	Up to 4 mm broad	To 2 mm	3-4 mm, also to Pannaria caespitosa P.M. Jørg	Up to 3 mm	0.3-1.0 mm wide	0.5-1.0 mm diam.	0.5-1.0(-2.0) mm broad
Marginal lobes		Enlarged	Enlarged	Deeply divided, finger like peripheral lobes	Fan-shaped	Radiated	± effigurate and spathulate	Discrete	Enlarged in the apices
Central part		Areolate to granulose	Not mentioned	Disintegrating in the subcrustose	Not mentioned	Centrally often verrucosely fragmented and partially detached	Irregularly rhomboidal and delimited by deep cracks	Grouped	Verrucose and fragmented or often partially detached
Apothecia	Up to 5 mm diam.	0.5-2.0 (-5.0) mm diam.	Up to 3 mm broad	To at most 2 mm diam	Up to 3.5 mm, also to Pannaria caespitosa P.M. Jørg	Up to 2 mm	0.4-1.3 mm diam.	0.4-1.3 mm diam.	0.4-1.5(-2.0) mm diam.
Discs	Some shade of brown	Brown to black	Black or dark brown	Usually with black	Pale brown	Black to rarely dark brown	Dark brown to black	Black to dark brown	Black discs
Ascospores	Ellipsoid, 14-16 × 7-8 µm	Ellipsoid, (11.5-)14-16(-17) × 7-8(-11) µm	Ellipsoid to globular, 12-14 × 10-12 µm	Nearly globular, 12-15 × 8-11 µm	Subglobose, 12-15 × 10-13 µm	Subglobose, 12-15 × 8-11 µm	Ellipsoid to ovate, (10-)12-14.0-16(-18) × 6-7.6-9(-10) µm	Ellipsoid to ovate, 12-16 × 6-9 µm	Ellipsoid to spherical, (7.5-)12.5-15.0(-17.5) × 5.0-12.5 µm
Examined specimens	South Georgia	South Shetland Islands	Bouvetøya Island	Canada, Greenland, Argentina, Chile, Kerguelen, Marion I., New Zealand, and South Shetland Isl.	Antarctica and South Georgia Island	Nordic Region	Tasmania	Argentina	14 localities from the northern and southern Hemispheres

Description of species

Pannaria hookeri (Borr. ex Sm.) Nyl., Mém. Soc. Sci. Nat. Math. Cherbourg 5: 109 (1857). (Fig. 2 A-F)

Description. Thallus placodioid. Lobes at the circumference narrow, 0.5-1.0(-2.0) mm broad and 1.0-2.5 mm long, convex, enlarged in the apices, centrally verrucose and fragmented or often partially detached. Upper surface whitish beige to brownish-grey under fluorescent light, whitish beige to whitish brown under the stereomicroscope. Hypothallus inconspicuous at marginal branches. Apothecia frequent, 0.4-1.5(-2.0) mm diam., with black discs. Thalline margin conspicuous, usually crenulate, concolorous with the thallus. Thallus 200-400 µm thick. Upper cortex 10-50 µm (2-12 layers of cells) thick, proso- to colloparaplectenchymatous tissue; cells thick-walled cells, 2.5-5.0 µm diam. Photobiont layer dense and massive, probably Nostoc clusters, spherical cells ca. 5 µm diam.; numerous vertically orientated fascicles of hyphae derived from the medulla penetrating into the photobiont layer. Medulla is hyaline with some photobionts. Lower cortex 12.5-25.0 µm (ca. 3 cells) thick when on the moss, blackish. Apothecia with a thick thalline margin to 250 µm, containing a large number of algal cells; cortex of thalline exciple colloplectenchymatous or with amorphous cells, 5.0-10.0 µm (2 cells) thick at the apices, 7.5-35.0 µm (2-8 cells) thick at the middle, indistinct at the base. Proper exciple ca. 50 µm thick, colloplectenchymatous to elongated cells. Subhymenial layers yellowish to hyaline, 25-65 µm thick, usually prosoplectenchymatous. Hymenium 100-200 µm thick, partially I+ persistently deep blue, mainly in the vicinity of the asci. Simple paraphyses, up to 2.5 µm thick, slightly clavate at the apices. Asci 50-65 × ca. 20 µm. Spores 8 per ascus, (7.5-)12.5-15.0(-17.5) × 5.0-12.5 µm, colorless, ellipsoid to spherical, smooth-walled. Pycnidia were not observed.

Spot tests: K-, C-, KC-, P+ orange (probably pannarin).

Notes: Pannaria hookeri is characterized by presence of thallus placodioid with marginal lobes enlarged (less than 2 mm), central part often verrucose, fragmented and partially detached, and by presence of pannarin (P+ orange). The lecanorine apothecia have 0.4-1.5(-2.0) mm diam., with the blackish discs, subhymenium of prosoplectenchymatous tissue, and hypothecium and parahymenium of colloplectenchymatous tissue and elongated cells. The ascospores have always smooth epispores.

We examined specimens are from Austria (4 specimens), Canada (2), Denmark (Faroe Islands, 1), Greenland (1), Iceland (1), Italy (1), Norway (16), Russia (1), Scotland (4), and Sweden (4) from the Northern Hemisphere; and Argentina (11), Chile (1), Kerguelen Island (2), and New Zealand (1) from the Southern Hemisphere. The determination of the specimens received on loan from herbaria was confirmed only through morphological approach. Unfortunately, most of the received material was collected more than 10 years ago (Text S1), and we were not able to generate genetic sequences. When the P. hookeri characters were not observed in the material, the specimen was excluded of the study.

Figure 2
Pannaria hookeri from Argentina. (A): Specimen on the habitat. (B): Detail of laciniate branches on moss (MJK3314). (C): Saxicolous specimen (MJK4041). (D): Transversal section of the thallus. (E): Section of apothecium on the squamule. (F): Diametral section of apothecium. Symbols. apo = apothecium; black arrowhead = parahymenial tissue; black arrow = moss; hym = hymenium; hyp = hypothallum; lc = lower cortex; mh = medullar hyphae; ph = photobiont layer; squ = squamule; sub = subhymenium; uc = upper cortex; white arrow = P+ orange (pannarin).

Discussion

In this study, we re-confirmed the wide geographical distribution of Pannaria hookeri through phylogenetic and morphological evidence, now including the first P. hookeri sequences from the southern South America, Argentina. The phylogenetic analysis resulted in a well-supported clade with P. hookeri sequences from Argentina, Australia, Norway, and the USA, whereas the morphological analysis included specimens from Argentina, Austria, Canada, Chile, Denmark (Faroe Islands), Greenland, Iceland, Italy, Kerguelen Island, New Zealand, Norway, Russia, Scotland and Sweden, generating a concise morphological description. The materials from Scotland are topotypes (Text S1) and were compared with fresh specimens.

The bipolar distribution has been concepted as species that are disjunctly distributed in both hemispheres, mostly in polar and subpolar regions, and are largely absent in the tropics. However, many lichens reported as bipolar are also distributed in temperate and tropical latitudes - usually in high mountains, where their ecological requirements are met (Garrido-Benavent & Pérez-Ortega 2017), which is the case of P. hookeri, and should be carefully revised and possibly considered as (sub)cosmopolitan. Nowadays, the species is mentioned in high altitudes of Ecuador and Kenya (material not revised in the present study) (Frisch & Hertel 1998; Jørgensen & Palice 2010), but efforts are still needed to confirm or not the species distribution through the tropical latitudes. Studies including DNA analysis of species reported as bipolar has confirmed this remarkable distribution, for example in Austroplaca, Cetraria and Cladonia (Myllys et al. 2003; Søchting & Castello 2012; Fernández-Mendoza & Printzen 2013), while others has resulted in the discovery of cryptic lineages or restricted distribution, such as in Parmelia, Sphaerophorus and Rusavskia (Crespo et al. 2002; Högnabba & Wedin 2003; Scur et al. 2022), indicating that phylogeographical studies should be performed to revise the distribution stated to P. hookeri.

Historically, Pannaria hookeri specimens from the Southern Hemisphere were reported with a larger morphological plasticity than specimens from Northern Hemisphere (Jørgensen 2000a), and the taxonomic problems has invalited other results, e.g. the chemical study (Quilhot et al. 1989). The variations in the color of the thallus, branching, diameter of apothecia, and color of the apothecium discs, which were reported in the literature from Antarctic specimens (Lindsay 1974; Redón 1985; Jørgensen 1986; 2000a; Øvstedal & Lewis Smith 2001), were not observed in the material confirmed by us and can be characters of other cryptic species. For example, apothecia up to 5.0 mm diam. were reported to the specimens from South Georgia and South Shetland Islands (Lindsay 1974; Redón 1985), apothecia up to 3.0 mm diam. in specimens from Bouvetøya Island (Jørgensen 1986), but only apothecia up to 2 mm diam. were observed during the present study.

In the same way, Øvstedal & Lewis Smith (2001) reported the species to South Georgia, Bouvetøya, South Orkney, South Shetland Islands, and Antarctic Peninsula, but posteriorly transferred the most of examined specimens within P. caespitosa P.M. Jørg. (Jorgensen 2000a). Only two specimens, SJA 168 and RILS 8347, were kept within P. hookeri, but they have the presence of pale brown discs in the apothecia (Øvstedal & Lewis Smith 2001) which do not match with the dark brown to blackish discs of P. hookeri (Tab. 2). Therefore, a revision is still required to the both specimens.

The material received as loan that were not confirmed as P. hookeri are from Antarctic and subantarctic Islands (Antarctica Peninsula and Kerguelen, Marion, South Georgia, South Orkney and King George islands - Text S1), highlighting the taxonomic problem in the region. In addition, fresh specimens collected during four Brazilian Antarctic Expeditions in the Antarctic Peninsula (near to Esperanza and Primavera stations), James Ross Island, Marambio Island, and South Shetland Islands (Deception, Greenwich, King George, Livingston, Nelson, Robert, and Snow Islands) did not cluster with the P. hookeri sequences and they were also excluded from our analysis. An integrative revision of the Psoroma-complex is also required to Antarctica as well as a complete identification key of the group; P. hookeri in Antarctica can be more restricted than previously thought.

Previously, P. hookeri was considered as strictly saxicolous (Jørgensen 2000a). However, we observed in Argentina that the species can grow on the rock (Fig. 1A), and also on saxicolous moss (Fig. 1B), expanding the species' known ecology. Furthermore, the species were usually found on the mountains in Argentina, Australia and Chile (Jørgensen 2000a; Kantvilas & Gueidan 2018). Calvelo & Fryday (2006) reported P. hookeri on rock outcrops along a stream the sea-level in Argentina, but the material has not been reviewed by us.

Our description was based on 50 specimens from 13 countries, corroborating with the descriptions provided by Jørgensen (1978; 2000a; 2007), Kantvilas & Gueidan (2018) and Passo et al. (2020). Therefore, the large morphological variations reported for specimens from the Southern Hemisphere are potentially characters of different Psoroma species. Studies including more genetic sequences, chemical and morphological variation analysis should be performed to dept the evolutionary history and concept of Pannaria hookeri.

Acknowledgments

The authors thanks Dr. A.E., Dr. A.P. and others anonymous reviewers that substantially corroborated to better the manuscript. This work was financed in part by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), under the “Ação Transversal nº 64/2013 - Chamada MCTI/CNPq/FNDCT”; and by the Fundação Universidade Federal de Mato Grosso do Sul - UFMS/MEC - Brazil. Marcos J. Kitaura was supported in part by the Fundação de Apoio ao Desenvolvimento do Ensino, Ciência e Tecnologia do Estado de Mato Grosso do Sul (FUNDECT - Chamada 04/2019), by the CNPq, and by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) - Finance Code 001". Mayara C. Scur was supported by the CAPES - Finance Code 001".

References

• Bendiksby M, Mazzoni S, Jørgensen MH, Halvorsen R, Holien H. 2014. Combining genetic analyses of archived specimens with distribution modelling to explain the anomalous distribution of the rare lichen Staurolemma omphalarioides: long-distance dispersal or vicariance? Journal of Biogeography 41: 2020-2031.
• Calvelo S, Fryday AM. 2006. New reports of lichens from Argentine Tierra del Fuego and the Falkland Islands (Islas Malvinas). The Bryologist 109: 372-380.
• Crespo A, Molina MC, Blanco O, Schroeter B, Sancho LG, Hawksworth DL. 2002. rDNA ITS and β -tubulin gene sequences analyses reveal two monophyletic groups within the cosmopolitan lichen Parmelia saxatilis Mycological Research 106: 788-795.
• Darriba D, Taboada GL, Doallo R, Posada D. 2012. JModelTest 2: More models, new heuristics and parallel computing. Nature Methods 9: 772.
• Doidge EM. 1950. The South African Fungi and Lichens to the end of 1945. Bothalia 5: 1-1094.
• Ekman S, Jørgensen PM. 2002. Towards a molecular phylogeny for the lichen family Pannariaceae (Lecanorales, Ascomycota). Canadian Journal of Botany 80: 625-634.
• Ekman S, Wedin M, Lindblom L, Jørgensen PM. 2014. Extended phylogeny and a revised generic classification of the Pannariaceae (Peltigerales, Ascomycota). Lichenologist 46: 627-656.
• Elvebakk A, Papaefthimiou D, Robertsen EH, Liaimer A. 2008 Phylogenetic patterns among Nostoc cyanobionts within bi- and tripartite lichens of the genus Pannaria. Journal of Phycology 44: 1049-1059.
• Elvebakk A, Robertsen EH, Park CH, Hong SG. 2010. Psorophorus and Xanthopsoroma, two new genera for yellow-green, corticolous and squamulose lichen species, previously in Psoroma. The Lichenologist 42: 563-585.
• Fernández-Mendoza F, Printzen C. 2013. Pleistocene expansion of the bipolar lichen Cetraria aculeata into the Southern hemisphere. Molecular Ecology 22: 1961-1983.
• Frisch A, Hertel H. 1998. Flora of Macrolichens in the Alpine and Subalpine zones of Mt. Kenya. Sauteria: 363-370.
• Garrido-Benavent I, Pérez-Ortega S. 2017. Past, present, and future research in bipolar lichen-forming fungi and their photobionts. American Journal of Botany 104:1660-1674.
• Guindon S, Gascuel O. 2003. A Simple, Fast, and Accurate Algorithm to Estimate Large Phylogenies by Maximum Likelihood. Systematic Biology 52: 696-704.
• Högnabba F, Wedin M. 2003. Molecular phylogeny of the Sphaerophorus globosus species complex. Cladistics 19: 224-232.
• Huelsenbeck JP, Ronquist F. 2001. MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics 17: 754-755.
• Jørgensen PM. 1978. The lichen family Pannariaceae in Europe. Opera Botanica 45: 1-123.
• Jørgensen PM. 1986. Macrolichens of Bouveteya. Norsk Polarinstitutt Skrifter 185: 23-34.
• Jørgensen PM. 2000a. Studies in the lichen family Pannariaceae IX - A revision of Pannaria subg. Chryopannaria. Nova Hedwigia 71: 405-414.
• Jørgensen PM. 2000b. Survey of the Lichen family Pannariaceae on the American Continent, North of Mexico. The Bryologist 103: 670-704.
• Jørgensen PM 2003. Notes on African Pannariaceae (lichenized ascomycetes). Lichenologist 35: 11-20.
• Jørgensen PM. 2006. Conspectus familiae Pannariaceae (Ascomycetes lichenosae). Revised version. Botanisk Institutt. Universitetet I Bergen.
• Jørgensen PM. 2007. Pannaria In: Ahti T, Jørgensen PM, Kristinsson H, Moberg R, Søchting U, Thor G. (eds.) Nordic Lichen Flora, Cyanolichens. Vol.3.Museum of Evolution, Uppsala Iniversity on behalf of Nordic Lichen Society. pp. 105-107.
• Jørgensen PM, Palice Z. 2010. Additions to the lichen family Pannariaceae in Ecuador. Nordic Journal of Botany 28: 623-628.
• Kantvilas G, Gueidan C. 2018. Pannaria hookeri (lichenised Ascomycetes) - a remarkable new record for Australia. Muelleria 36: 74-80.
• Katoh K, Misawa K, Kuma K, Miyata T. 2002. MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform. Nucleic Acids Research 30: 3059-3066.
• Kearse M, Moir R, Wilson A, et al 2012. Geneious Basic: An integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 28: 1647-1649.
• Kitaura MJ, Scur MC, Spielmann AA, Lorenz-Lemke AP. 2018. A revision of Leptogium (Collemataceae, lichenized Ascomycota) from Antarctica with a key to species. Lichenologist 50: 467-485.
• Lindsay D. 1974. The macrolichens of South Georgia. British Antarctic Survey Reports: 1-91.
• Lindsay DC. 1976. Two new lichens from Marion Island, southern Indian Ocean. Nova Hedwigia: 877-880.
• Magain N, Sérusiaux E. 2014. Do photobiont switch and cephalodia emancipation act as evolutionary drivers in the lichen symbiosis? A case study in the Pannariaceae (Peltigerales). PLoS ONE 9. doi:10.1371/journal.pone.0089876
  » https://doi.org/10.1371/journal.pone.0089876
• Marthinsen G, Rui S, Timdal E. 2019. OLICH: A reference library of DNA barcodes for Nordic lichens. Biodiversity Data Journal 7: e36252. doi: 10.3897/BDJ.7.e36252
  » https://doi.org/10.3897/BDJ.7.e36252
• Miller MA, Pfeiffer W, Schwartz T. 2010. Creating the CIPRES Science Gateway for inference of large phylogenetic trees. Gateway Computing Environments Workshop: 1-8.
• Myllys L, Stenroos S, Thell A, Ahti T. 2003. Phylogeny of bipolar Cladonia arbuscula and Cladonia mitis ( Lecanorales , Euascomycetes). Molecular Phylogenetics and Evolution 27: 58-69.
• Olech M. 1989. Lichens from the Admiralty Bay region, King George Island (South Shetland Islands, Antarctica). Acta Societatis Botanicorum Poloniae 58: 493-512.
• Olech M. 2001. Annotated checklist of Antarctic lichens and lichenicolous fungi. Kraków, The Institute of Botany of the Jagiellonian University.
• Øvstedal DO, Lewis Smith RI. 2001. Lichens ofAntarctica and South Georgia: A Guide to their Identification and Ecology. Cambridge, Cambridge University Press.
• Park CH, Hong SG, Elvebakk A. 2018. Psoroma antarcticum, a new lichen species from Antarctica and neighbouring areas. Polar Biology 41: 1083-1090.
• Passo A, Stenroos S, Calvelo S. 2008. Joergensenia, a new genus to accommodate Psoroma cephalodinum (lichenized Ascomycota). Mycological Research 112: 1465-1474.
• Passo A, Díaz Dominguez RE, Rodríguez JM. 2020. El género Pannaria (Pannariaceae) en la Argentina: nuevos registros y actualización del conocimiento. Boletín de la Sociedad Argentina de Botánica 55: 339-357.
• Quilhot W, Piovano M, Arancibia H, Garbarino JA, Gambaro V. 1989. Studies on Chilean lichens, XII. Chemotaxonomy of the genus Psoroma Journal of Natural Products 52: 191-192.
• Redón J. 1985. Liquenes antarticos. Santiago, Instituto Antatico Chileno.
• Ronquist F, Huelsenbeck JP. 2003. MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19: 1572-1574.
• Schoch CL, Seifert KA, Huhndorf S, et al 2012. Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi. Proceedings of the National Academy of Sciences of the United States of America 109: 6241-6246.
• Scur MC, Kitaura MJ, de Paula JB, Spielmann AF, Lorenz AP. 2022. Contrasting variation patterns in Austroplaca hookeri and Rusavskia elegans (Teloschistaceae, lichenized Ascomycota) in maritime Antarctica. Polar Biology 45: 101-111.
• Smith JE, Sowerby J. 1811. English Botany; or, coloured figures of British Plants, with their essential characters, synonyms and places of growth: to which will be added, Occasional Remarks by James Edward Smith.
• Søchting U, Castello M. 2012. The polar lichens Caloplaca darbishirei and C. soropelta highlight the direction of bipolar migration. Polar Biology 35: 1143-1149.
• Spribille T, Muggia L. 2013. Expanded taxon sampling disentangles evolutionary relationships and reveals a new family in Peltigerales (Lecanoromycetidae, Ascomycota). Fungal Diversity 58: 171-184.
• Stamatakis A. 2014. RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30: 1312-1313.
• Stizenberger E. 1890. Lichenaea africana. Bericht über die Thatigkeit der St. Gallischen Naturwissenschaftlichen Gesellschaft 2: 133-268.
• Thomson JW. 1984. The lichens of Arctic America. Part II. Microlichens. New York, University Press.
• Wedin M, Wiklund E, Jørgensen PM, Ekman S. 2009. Slippery when wet: Phylogeny and character evolution in the gelatinous cyanobacterial lichens (Peltigerales, Ascomycetes). Molecular Phylogenetics and Evolution 53: 862-871.

Publication Dates

• Publication in this collection
  21 Nov 2022
• Date of issue
  2022

History

• Received
  05 Dec 2021
• Accepted
  29 Aug 2022