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Research Article
The genus Massalongia (lichenised ascomycetae) in the Southern Hemisphere
expand article infoPer M. Jørgensen, Heidi L. Andersen, Arve Elvebakk§
‡ University Museum of Bergen, Bergen, Norway
§ University of Tromsø – the Arctic University of Norway, Tromsø, Norway
Open Access

Abstract

The species of Massalongia recorded and described from the Southern Hemisphere are revised and it is shown that only one is present; M. patagonica which is widespread, with populations in Australia and New Zealand that differ from the South American populations, but at present best regarded as part of the variation of that species. Records from this hemisphere of all other species placed in the genus are incorrect. The type species, M. carnosa, is restricted to the Northern Hemisphere. Two species, M. antarctica and M. novozelandica cannot be identified precisely due to lack of sufficient type material and with the types as the only collections known of these, but none belongs in Massalongia according to available data. Massalongia griseolobata (from Gough Isl.) is shown here to belong in the Pannariaceae and is part of the parmelielloid clade. M. intricata (from South Georgia) and M. olechiana (from South Shetland) have both recently been correctly transferred to the genus Steinera in the Arctomiaceae.

Keywords

Peltigerales, Massalongiaceae, phylogeny, taxonomy, South Hemisphere

Introduction

The genus Massalongia was described by Körber (1855), based on the species Lichen carnosus described by J. Dickson in 1790 on material collected in Scotland, but later often called Pannaria muscorum (Ach.) Duby, an illegitimate, superfluous name. This reflects the difficulties which the early lichenologists had in classifying the species. Molecular studies (Wedin et al. 2007; Muggia et al. 2011) have shown that Massalongia does not belong in the Pannariaceae, but is best placed in a family of its own, Massalongiaceae, in the Peltigerales. There is, however, only one major study of the species and their variation, made by Henssen (1963) covering North America. She accepts two species; the widespread, variable M. carnosa and the nearly crustose, microphylline, local Californian endemic M. microphyllizans (Hasse ex Nyl.) Henssen. Jørgensen (2000), whilst revising the Pannariaceae, had studied the type of Massalongia fauriei (Hue) Zahlbr. and found the poor type (the only material existing) to belong in Fuscopannaria leucophaea s.lat., now transferred to the genus Vahliella (Jørgensen 2008; Wedin et al. 2010). An additional Asian species has been recorded from the Philippines based on Rehm (1916). This is, however, based on a misunderstanding of Massalongiella imperatae Rehm., which is a non-lichenised ascomycete as originally described. In Europe, Harmand recognised a variety which Gyelnik (1940), in his notorious treatment of the Pannariaceae in Rabenhorst’s Kryptogamenflora, raised to species rank as Massalongia meizospora, only representing a form with somewhat longer, 3-septate spores. In addition, he established a new Massalongia rabenhorstiana, the type of which has disappeared. It is most certainly only a synonym of M. carnosa, still the only species on the Northern Hemisphere in addition to M. microphyllizans, a species in need of a phylogenetic study.

The situation in the Southern Hemisphere is different, though it took a long time before any species in the genus was recognised. Zahlbruckner (1917) was the first when he recorded Massalongia carnosa from the Falkland Islands, followed by Lamb (1958) who recorded it from Patagonia. Later, it was mentioned from several regions in the Southern Hemisphere (Smith and Corner 1973; Lindsay 1974; Galloway 1985; Redon 1985; Jørgensen 1986; Jørgensen and Elix 1988; Øvstedal and Smith 2001). In addition, several new species were described from the Southern Hemisphere; M. antarctica Dodge (from the Antarctic Peninsula, Dodge 1971), M. novozelandica Dodge (from subantarctic New Zealand, Dodge 1971), M. griseolobata Øvstedal (from Gough Isl., Øvstedal and Gremmen 2010), M. intricata Øvstedal (from South Georgia, Øvstedal and Smith 2001) and M. olechiana Alstrup & Søchting (from South Shetland, Alstrup and Søchting 2011).

During fieldwork in Chile in 2015, one of the authors (A.E.) discovered a strange Pannaria-like lichen which, on closer inspection, proved to be a Massalongia with some differences from M. carnosa, as known by us from Norway. However, since this is a variable species, we felt that a more detailed study, including molecular screening, would be useful. This being done and the distinction of this material proven, we found it necessary to check on the surprisingly high number of species of Massalongia described from the Southern Hemisphere. This proved to be time-consuming and complicated, since it was difficult to get hold of suitable material and, when molecularly checked, often not giving clear results and involving quite unrelated lichen families. Fortunately, Ertz et al. (2017) solved some of our problems and, eventually, this new species was named by Kitaura and Lorenz in Liu et al. (2018). However, our project contains more data than their work includes and we present these here in an attempt to give full clarification of the taxonomic situation for the genus in the Southern Hemisphere. Some additional phylogenetic data were also added on the genus in the Northern Hemisphere.

Material and methods

The specimens

Specimens of Massalongia were obtained from various herbaria for phylogenetic analyses, see Tables 1 and 2. In addition, we microscopically studied material from the following herbaria: BAA, BG, BM, C, CANB, CANL, CHR, F, FH, H, MSC, NY, TROM and UPS. A total of 130 ascospores from collections from both hemispheres were drawn in detail and measured for comparison.

Table 1.

List of specimens used for phylogenetic analyses of the broad analysis of the Massalongiaceae and Pannariaceae, with vouchers and accession numbers from GenBank. Bold accession numbers are new in this study.

Species and ID Voucher mtSSU RPB1
Austrella arachnoidea Jørgensen 8200 (BG) KC608054 KC608108
Collema furfuraceum Wedin 6187 (BM) AY340488 GQ259048
Collema nigrescens Wedin 7046 (UPS) GQ259020 GQ259049
Collema parvum Nordin 5500 (UPS) GQ259021 GQ259050
Degelia duplomarginata Wedin 8023 (S) KC608058
Degelia durietzii Elvebakk 02-296 (TROM) KC608059
Degelia gayana Wedin 6112 (UPS) AY652619
Degeliella rosulata Galloway 840b (BG) KC608063
Degeliella versicolor Galloway 840a (BG) KC608064
Erioderma pedicellatum mrSSU: MacPitcher s.n. 2007 (BG-L85909), RPB1: MacPitcher s.n. 2007 (BG-L85911) KC608065 KC608110
Erioderma verruculosum AFTOL-ID 337 DQ972990 DQ973062
Fuscoderma applanatum Tibell 19076 (BG) KC608112 KC608066
Fuscopannaria pacifica Tønsberg 29359 (BG) KC608074 KC608118
Fuscopannaria praetermissa mrSSU: Tønsberg 36838 (BG)
RPB1: Wedin 7671 (UPS)
KC608075 GQ259056
Joergensenia cephalodina Passo 269 (BCRU 4895) EU885330
Lecidea fuscoatra Wedin 6860 (UPS) AY756401 AY756408
Leciophysma furfurascens Nordin 5695 (UPS) GQ259028 GQ259058
Leioderma erythrocarpum Schumm and Frahm s.n. 2009 (BG, dupl of hb. Schumm 15583) KC608078
Leioderma pycnophorum Wedin 8013 (S) GQ259031 GQ259059
Leptochidium albociliatum Tønsberg 29087 (BG) DQ900632 GQ259060
Muggia TSB38886 JF938191
Spribille 20997 (COLO) JF938193
Lobaria pulmonaria mrSSU: Wedin 6167 (UPS)
RPB1: Wedin 5092(UPS)
AY340503 GQ259068
Lobaria scrobiculata AFTOL-ID 128 AY584621 DQ883736
Massalongia carnosa Tønsberg 44267 (BG) MN708314 MN714653
Tønsberg 45410 (BG) MN708315 MN714654
Johnsen L-86694 (BG) MN708316
Ezhkin 1289 (SAK) MN708317 MN714655
Spribille 22021 (COLO) JF938205
Haikonen 20961 (H) EU558817
Hermansson 8916 (UPS) AY340509 GQ259071
Spribille 21565 (COLO) JF938204
Massalongia patagonica Elvebakk 99:775 (TROM) MN708318 MN714656
Elvebakk 15:033 (SGO) MN708319 MN714657
Buck 60287 (NY) MN708320 MN714658
Gremmen K-789 (BG) MN708321 MN714659
Galloway 5616 (CHR) MN708322 MN714660
Galloway s.n. (CHR) MN708323 MN714661
Kitaura 4181 (CGMS) MG243608
Kitaura 4188 (CGMS) MG243607
Kitaura 4168 (CGMS) MG243609
Massalongia griseolobata Gremmen 2006-91 (BG) MN708324
Nephroma parile Wedin 6169 (UPS) AY340512 GQ259072
Pannaria hookeri Jørgensen s.n. (BG) KC608083 KC608124
Pannaria immixta Elvebakk 02-352b (BG) KC608084 KC608125
Pannaria rubiginella Thor 10050 (S) GQ259037 GQ259074
Parmeliella appalachensis Lendemer 578 and Smith (BG) KC608090
Parmeliella miradorensis Tønsberg 23053 (BG) KC608094 KC608136
Parmeliella nigrocincta Elvebakk 02-356 (BG) KC608095 KC608137
Parmeliella pannosa Ståhl s.n. 1999 (BG) KC608096
Parmeliella triptophylla Wedin 7037 (UPS) AY652623 GQ259075
Pectenia atlantica Lindblom and Blom L251 (BG) KC608057 KC608109
Pectenia cyanoloma Purvis, James and Smith 1995 (BM) AY340491 GQ259052
Pectenia plumbea AFTOL-ID 1046 DQ912300 DQ912374
Placynthium nigrum Wedin 6778 (UPS) AY340518 GQ259079
Polychidium muscicola Obermayer 8547 (UPS) DQ900634 GQ959080
Spribille 26411 (KLGO) JF938220
Pseudocyphellaria aurata Purvis, James and Smith 7/5/1995 (BM) AY340520 GQ259082
Psoroma hypnorum Ihlen 453 (BG) KC608100 KC608142
Santessoniella arctophila Kristinsson s.n. (BG) KC608104 KC608145
Sticta fuliginosa Wedin 6078 (BM) AY340529 GQ259089
Vahliella carnifornica Tønsberg 26316 and Goward (BG) HQ268594 HQ268593
Vahliella leucophaea Wedin 6849 (UPS) AY652621 GQ259090
Table 2.

List of specimens used for phylogenetic analyses of the species delimitation in Massalongia, M. referring to Massalongia, P. to Polychidium and L. to Leptochidium, with vouchers and accession numbers from GenBank. Bold accession numbers are new in this study.

Species and ID Voucher Area ITS LSU mtSSU RPB1
M. carnosa Tønsberg 44267 (BG) USA: Alaska MN708327 MN708327 MN708314 MN714653
Tønsberg 45410 (BG) USA: Alaska MN708328 MN708328 MN708315 MN714654
Johnsen L-86694 (BG) Norway MN708329 MN708329 MN708316
Ezhkin 1289 (SAK) East Russia MN708330 MN708330 MN708317 MN714655
Spribille 22021 (COLO) USA: Montana JF938205
Hermansson 8916 (UPS) Sweden AY340554 AY340509 GQ259071
Rui andTimdal 13267 (O) Norway MG243601 MG243611
Hansen 1138 (COLO, H) Greenland MG243599 MG243615 MG243610
Hansen 1057 (H) Greenland MG243603 MG243616 MG243612
Türk 17280 (H) Austria MG243602 MG243614 MG243613
M. patagonica Elvebakk 99:775 (TROM) Chile MN708331 MN708331 MN708318 MN714656
Elvebakk 15:033 (SGO) Chile MN708332 MN708332 MN708319 MN714657
Buck 60287 (NY) Chile MN708333 MN708333 MN708320 MN714658
Gremmen K-789 (BG) Australia MN708321 MN714659
Galloway 5616 (CHR) New Zealand MN708334 MN708334 MN708322 MN714660
Galloway s.n. (CHR) New Zealand MN708335 MN708335 MN708323 MN714661
Kitaura 4181 (CGMS) Argentina MG243604 MG243617 MG243608
Kitaura 4188 (CGMS) Argentina MG243606 MG243619 MG243607
Kitaura 4168 (CGMS) Argentina MG243605 MG243618 MG243609
P. muscicola Obermayer 8547 (UPS) Austria DQ900647 DQ900634 GQ259080
L. albociliatum Tønsberg 29087 (BG) USA DQ900644 DQ900632 GQ259060

DNA extraction, Amplification and Sequencing

Total genomic DNA was extracted using DNeasy Plant Mini Kit (Qiagen). Four DNA markers where amplified; the mitochondrial small subunit rDNA (mtSSU rDNA: primers mrSSU1 and mrSSU3R (Zoller et al. 1999)), the internal transcribed spacer (ITS) and the large subunit (LSU) regions of the nuclear ribosomal RNA gene (primers ITS1f (Gardes and Bruns 1993), ITS4 (White et al. 1990), LSU155 and LSU362 (Döring et al. 2000), LSU635/LR3 and LSU1125/LR6 (Vilgalys and Hester 1990)) and the gene coding for the largest subunit of RNA polymerase II (RPB1: primers PRB1-BCR (Wedin et al. 2009), gRPB1-A (Stiller and Hall 1997) and fRPB1-C (Matheny et al. 2002)).

PCR reactions consisted of 1× GeneAmp PCR Buffer II (Applied Biosystems), 2.5 µM MgCl2 (Applied Biosystems), 20 µM dNTPs (Promega), 0.4 µM of each primer, 0.03 U AmpliTaq DNA Polymerase (Applied Biosystems), 2–5.0 µl of genomic DNA extract and distilled water to a total volume of 25 µl. PCR reactions were performed on a C1000 Touch thermal cycler (Bio-Rad Laboratories), with the following temperatures; initial denaturation at 94 °C for 4 min, followed by a 62–56 °C touchdown annealing for the first 6 cycles, ending with 30 cycles at 56 °C for 30 sec, polymerisation at 72 °C for 1 min 45 sec and a final elongation at 72 °C for 10 min.

Direct sequencing of PCR products was run with the PCR primers using a BigDye Terminator Cycle Sequencing kit (Applied Biosystems) on an ABI Prism 3700XL DNA analyser (Applied Biosystems) at the DNA Sequencing Facility (UiB), Norway. Sequences were assembled and edited using Geneious v.11.0.2 (Kearse et al. 2012).

Newly generated sequences with GenBank accession numbers are listed in Tables 1, 2, together with sequences downloaded from GenBank.

Phylogenetic analyses

To align the sequences, MAFFT v7.309 (Katoh et al. 2002; Katoh and Standley 2013) implemented in Geneious v.11.0.2 (Kearse et al. 2012) was used with default settings, followed by manual adjustments. Suitable substitution models for the separate datasets were identified using MrAIC v.1.4.6 (Nylander 2004).

Two different datasets were analysed; one broad analysis of Massalongiaceae and Pannariaceae to test whether the included species is part of Massalongia (Table 1) and a second analysis for species delimitation within Massalongia (Table 2). For the broader dataset, mtSSU and RPB1 were concatenated, using Lecidea fuscoatra as outgroup and for the species delimitation in Massalongia, mtSSU, LSU, ITS and RPB1 was concatenated using Polychidium muscicola as outgroup.

Separate analyses of all genes and concatenated datasets were run as Bayesian MCMC searches using MrBayes v.3.2.1 (Ronquist and Huelsenbeck 2003) with default options; substitution model GTR+G+I, 10 million generations starting with a random tree, four simultaneous chains and using the default temperature of 0.2. Every 1000th tree was saved. Phylogenetic trees were visualised using Geneious v. 11.0.2 (Kearse et al. 2012).

Results

Phylogeny

The two resulting concatenated datasets consisted for the broad analysis of Massalongiaceae and Pannariaceae of 63 taxa with 1435 characters, whereas for the species delimitation in Massalongia, of 21 taxa with 2983 characters (details in Table 3).

Table 3.

List of numbers of characters, taxa and constant variables, from the two concatenated datasets.

Dataset Numbers of characters Numbers of taxa Number of constant characters Number of variable characters
Broad analysis of the Massalongiaceae and Pannariaceae 1435 63 724 711
Species delimitation in Massalongia 2983 32 2745 238

The resulting phylogenetic consensus tree from the broad analysis of Massalongiaceae and Pannariaceae are given in Fig. 1. Both Massalongia carnosa and M. patagonica are with high support a part of the Massalongiaceae, together with Polychidium muscicola and Leptochidium albociliatum. M. griseolobata is a part of the Pannariaceae, in the “Parmelielloid” clade 1 from Ekman et al. (2014) with high support. Within this clade, M. griseolobata is a part of a supported group with no internal resolution, including Degeliella rosulata, Degeliella versicolor, Leioderma, Erioderma and Joergensenia.

Figure 1. 

The phylogenetic tree of a concatenated broad dataset of the Massalongiaceae and Pannariaceae, resulting from Bayesian MCMC analyses.

The resulting phylogenetic consensus tree from the species delimitation analysis of Massalongia is given in Fig. 2. M. patagonica from the Southern Hemisphere and M. carnosa from the Northern Hemisphere are nicely separated in two sister groups with high support.

Figure 2. 

The phylogenetic tree of a concatenated dataset for species delimitation of Massalongia, resulting from Bayesian MCMC searches.

The samples of M. patagonica from New Zealand are grouped in a separate subclade from the rest of the samples from Australia, Chile and Argentina. The phylogenetic tree indicates a high genetic variance within M. patagonica throughout the Southern Hemisphere, but further studies are necessary to evaluate these differences.

The samples from the Northern Hemisphere make a monophyletic clade with little variation between the samples and a sample from Sweden is practically identical to those analysed from Alaska and Greenland.

Taxonomy

The only species from the Southern Hemisphere which, according to our data belongs in Massalongia, is M. patagonica, the details of which are as follows:

Massalongia patagonica Kitaura & Lorenz in Liu et al.

MycoBank No: 824006

Massalongia patagonica Kitaura & Lorenz in Liu et al., Sydowia 70: 249–252 (2018) – Holotypus: Argentina, Ushuaia, National Park of Tierra del Fuego, Lapataia Bay, muscicolous on the rock, 54°50'41.42"S, 68°33'52.31"W, 6 m alt., 25 December 2016, leg.M.J.Kitaura, J.Bordin, A.A.Spielmann & D.Peralta 4188 (CGMS).

Description

M. patagonica (Fig. 3) is morphologically similar to M. carnosa. Generally, spore characters are the best distinguishing characters (Fig. 4). The spores of M. carnosa are longer and 92% of 72 measured spores were in the range 23–35 µm. By contrast, 70% of 58 measured spores of M. patagonica were in the range 15–22 µm. This means that there is an overlap in sizes between these two species. However, the spores of M. patagonica are often two-septate, sometimes three-septate, which is very rare in M. carnosa. Here follows a more detailed treatment of Massalongia patagonica Kitaura & Lorenz:

Figure 3. 

Massalongia patagonica, AE 15-033. Scale bar: 1 cm.

Figure 4. 

Massalongia ascospores, patagonica above, carnosa below. Scale bar: 20 µm.

Thallus foliose, forming rosettes up to 3 cm, mostly muscicolous; lobes 0.5–1.5 mm broad, up to 1 cm long, irregularly and repeatedly divided with isidioid marginal outgrowths, simple to sparingly branched, sometimes developing into branched lobule systems, lobules, 0.1 mm wide. Upper surface brown, glabrous and glossy; upper cortex 20–30 µm thick, paraplectenchymatic, of thick-walled (ca. 1.5–2 µm wide) cells with 7–12 µm large lumina; photobiont layer 40–60 µm thick, often also developed in the subhymenium; cyanobiont Nostoc, cells bluish-green, irregularly subglobose to ellipsoid, 5–9 × 6–11 µm in size, arranged within 20–40 µm large glomeruli without visible chain structures, chain structures visible in some liberated cells; medulla loose, 60–80 µm thick; lower cortex absent, with scattered rhizohyphae.

Apothecia common to scattered, substipitate, laminal, 1–2 mm wide; thalline excipulum lacking, true excipulum weakly prominent; epithecium 5–10 µm thick, of protruding brown and strongly swollen, pyriform paraphyse end cells, 4–6 µm wide, 7–10 µm long, paraphyses undivided to sparingly divided, 2–4 µm thick; hymenium ca. 60 µm thick, IKI + blue; asci clavate 50–70 × 10–15 µm, 8-spored, with distinct internal apical IKI + blue sheath-like structures, sometimes also with weak tube structures; ascospores narrowly ellipsoid, occasionally asymmetric, 1- to 2 (3)-septate, (13) 20–25 (28) × 5–7.5 µm. Hypothecium ca. 60 µm thick, weakly brownish, IKI negative. Conidiomata not seen.

Chemistry

All reactions negative, no lichen substances detected by TLC.

Habitat and distribution

This is a species of wet to dry rock surfaces or boulders, usually growing in between mosses or on plant remains. It has a widely scattered distribution in South America, ranging from the temperate forests of south-central Chile, including the Juan Fernandez Islands and Patagonia, with two widely separated collections from southernmost Chile and Argentinean Tierra del Fuego. In addition, it is known from the Falkland Islands, Antarctica, mountains of SE Australia, where it is rare and from several localities in New Zealand.

Specimens examined

Antarctica: South Shetland Islands, King George Isl., Admiralty Bay, creeping slopes above Paradise Cove, 26 Jan 1980, R. Ochyra 1224/80 (BG, H); Urbanek, Crag between Polar Committee Glacier and Ladies Icefall, in Ezcurra Inlet, 20 Feb 1980, R. Ochyra 2319/80 (BG, H).

Argentina: Patagonia: Chubut, Lago Verde, near Futalaufquen, 1 Feb 1950, I. M. Lamb 5877 (over mosses on a rock in open scrub, about 30 m above the lake), 5880 (over mosses on a rock in open forest about 15 metres above the lake) (CANL, UPS).

Australia: New South Wales, near the summit of Mt. Guthrie, Kosciusko National Park, on moss over granite rocks, 9 Feb 1978, J.A. Elix 4360 (CANB); Kosciusko National Park, near Digger’s Creek, 21 Jan 1976, J. A. Elix 1722 (CANB); Île Australia near Kerguelen Isl., on moss cushions, 492823S, 695329E, 45 m alt., 31 Dec 2003, NJM Gremmen, K-789 (BG).

Chile: IX Región de la Araucanía: Reserva Nacional Malalcahuello, W bank of Río Colorado, 500 m W of the Entrance/CONAF building and 200 m S of the junction between the paths Sendero Coloradito and Sendero Sierra de Colorado; 38°25'45"S, 71°32'44"W, 1380 m alt., over Grimmia mosses on a S-facing rock outcrop in a Nothofagus dombeyi-Araucaria araucana forest, probably affected by river water during high flooding events, 9 Jan 2015, A. Elvebakk 15:033 (SGO, BG, UPS, BM, TROM); Archipiélago de Juan Fernandez: Isla Alejandro Selkirk (Mas Afuera), Los Innocentes, 4 Dec 1965, H. Imshaug (MSC); Valdivia, Corral, R. Thaxter (MSC); XII Región de Magallanes y de la Antárctica Chilena, Provincia Magallanes, Morro Chico, 52°03'S, 71°28'W, 200 m alt., on acrocarpous mosses on a NW-facing rocky slope, 28 Nov 1999, A. Elvebakk 99:775 (TROM); Provincia Antártica Chilena, Comuna Cabo de Hornos, Isla Grande de la Tierra del Fuego, Bahía Yendegaia, NNE shore opposite Caleta Ferrari, 54°50'28"S, 68°47'52"W, 13 Jan 2013, W.R. Buck 60287 (NY 01886528).

Falkland Islands: W. Falkland, Chartres, Luxton NNR, 30 Jan 2015, A. Fryday 10999 (MSC).

New Zealand: Canterbury, Cass, between Sugar Loaf and Cass Hill, 761 m alt., 18 Feb 1991, A. J. Fife 9761 (CHR); Banks Peninsula, Mt. Sinclair, summit, 5 Feb 1970, D. J. Galloway (CHR); Mt. Cook National Park, D. J. Galloway (CHR); Otago, Deep Stream, above DCC water intake, 13 Feb 1998, D. J. Galloway 0170 (CHR); Otago, Old Man Range, N of Obelisk, 5 Feb 2009, D. J. Galloway 404009 (CHR); St. Mary’s Range, Anakin’s Skifield, 22 Feb 2006, D. J. Galloway (CHR); Lake Onslow near huts, amongst moss in drainage cracks of schist rock in grassland, 30 Jul 1998, D. J. Galloway 404012 (CHR); Otago, Pomahaka River- Hukarere, rock slabs above river, 13 Apr 1998, D.J. Galloway 404011 (CHR); North Rough Ridge, near “Great Tor”, 12 Apr 1998, D. J. Galloway (CHR).

The other taxa originally described from the Southern Hemisphere as Massalongia species, are listed alphabetically, according to the epithet at the end of the discussion.

Discussion

The result of the phylogenetic analyses of Massalongia (Fig. 2) show that Massalongia carnosa and M. patagonica are located in different supported clades, as separate species as also described by Kitaura and Lorenz in Liu et al. (2018), M. patagonica being restricted to the Southern Hemisphere, whereas M. carnosa occurs only in the Northern Hemisphere. The clade with M. carnosa includes one circumarctic and circumboreal species, with low genetic diversity, whereas M. patagonica is more variable and shows a geographic pattern within this species. The material from New Zealand groups in a distinct branch within the M. patagonica clade and is superficially much more similar to the material of M. carnosa, but has extra short ascospores measured in two samples from New Zealand, all spores were shorter than 23 µm. This could be a result of the preference for moist, mossy habitats (Galloway 2007) as opposed to the drier, often exposed habitats in Chile. The material from Australia and New Zealand is, therefore, best classified as part of the M. patagonica complex.

That species is also found as far west as the Juan Fernandez Islands and is also possibly present on the Antarctic Peninsula and the Bouvet Island, but the material examined was sparse, sterile and too old for molecular studies.

Still, M. patagonica is not morphologically easily distinguished from M. carnosa; the two species have different spores, although there is an overlap zone in both length and degree of septation. Both species have a gross morphology showing high variation, probably due to habitat modifications, depending on light exposure, competition, moisture and water availability.

Chilean material of M. patagonica tends to have thicker, narrower and clearly radiating lobes than most material of M. carnosa. However, in cases where habitat information is available, they appear to be dry, but exposed to nutrient supplies by spring flooding (the Río Colorado collection), wind-transported saline lake dust (the Morro Chico collection) or seashore spray (the Tierra del Fuego collection). The New Zealand material, on the other hand, treated as M. carnosa, is cited as widespread and from moist habitats by Galloway (2007).

This detailed phylogenetic signal within M. patagonica is the result of a long history of evolution and isolation in austral areas, although shorter than the split-up between M. carnosa and M. patagonica. There is a record of M. carnosa from Mt. Kinabalu on Borneo (Sipman 1993) which could have indicated a migration route between a northern and a southern distribution area of the genus; however, a check of the material deposited at herbarium B revealed that it instead represents a sterile, richly squamulose specimen of a Parmeliella species. Future studies should investigate phylogeographic relationships between the three accepted species and the molecular distances between M. patagonica in New Zealand, Australia and South America/West Antarctica.

The examination of all relevant material from the Southern Hemisphere, shows the following, treated alphabetically according to the epithet:

Massalongia antarctica Dodge is a species only known from the type specimen from Lambda Island at the tip of the Antarctic Peninsula (Siple 380c-2, FH!). The type specimen is minute and sterile and consists of two different species, none of which belongs in Massalongia. The one fitting best with the description has a crustose, hemi-gelatinous thallus in accordance with species of the Arctomiaceae. There are no apothecia present in the collection and the description of the apothecia, given by Dodge (1968), is at variance with characters of Massalongia, indicating a species of the Arctomiaceae, most probably in Arctomia. There is, however, no known species with such a distinctly crustose thallus. More material is needed to identify this taxon more exactly. The sample also contains squamules with a trebouxioid photobiont and this is possibly Pertusaria corallifera Vain. as pointed out by Castello and Nimis (1995).

Massalongia griseolobata Øvstedal is a species only known from the type specimen (from Gough Isl., coll. Gremmen 2006-91, BG!). Even if only incipient apothecia were found, we do not hesitate to place this species in the Pannariaceae, based on morphology and the original description of the asci. They are recorded to have apically blue in tholus in iodine with a weak ring-structure. (Massalongia has sheet-like structures, Jørgensen 2007). The molecular study confirms this (Fig. 1). The species groups in the parmelielloid clade (Clade 1) in the tree by Ekman et al. (2014), with Degeliella, Leiderma, Erioderma and Joergensiana. This is an unresolved group of subantarctic taxa (Jørgensen and Andersen 2015) in need of further studies.

Massalongia intricata Øvstedal was correctly transferred to the genus Steinera by Ertz et al. (2017). S. intricata has a semi-gelatinous thallus producing apothecia on special lobules, just as species in the Arctomiaceae.

Massalongia novozelandica Dodge was recorded by Galloway (2007), but the holotype (the only material) has not been possible to obtain. However, the original description of the spores being brownish at maturity with disappearing septae (clearly pseudoseptae) is at variance with characters found in Massalongia. We agree with Galloway that this is probably a parasite growing on the thallus of a species in the Pannariceae.

Massalongia olechiana Alstrup and Søcht. was correctly transferred to the genus Steinera in the Arctomiaceae by Ertz et al. (2017).

Massalongia patagonica Kitaura and Lorenz, the recently described species (Fig. 3), belongs in the genus and, according to our phylogenetic tree (Figs 1, 2), prove to be distinct from M. carnosa, the latter being restricted to the Northern Hemisphere. The two species are morphologically variable due to the ecological conditions, but have different spores (Fig. 4), usually shorter than 25 µm in M. patagonica, but variable both in length and number of septae in both species. Chilean material of M. patagonica tends to have thicker, narrower and clearly radiating lobes than most material of M. carnosa.

Conclusion

From these facts, we conclude that there is only one, widespread species of Massalongia in the Southern Hemisphere, M. patagonica, though the populations in Australia and New Zealand differ somewhat molecularly, but more data is necessary to decide their taxonomic status. M. patagonica has a wider distribution than indicated in the original paper, also southwards and westwards. Previous records of several species in the Southern Hemisphere proved incorrect, most of them belonging in other genera.

The type species M. carnosa is restricted to the Northern Hemisphere, where it is widespread and variable, but without distinct molecular groupings requiring taxonomic recognition. There is also a local endemic, M. microphyllizans in California.

Acknowledgements

We are indebted to the directors and curators of the cited herbaria, to Louise Lindblom, Univ. Bergen for technical assistance and Mari Karlstad, Tromsø University Museum for photographs. Our sincerest thanks to all of you. Corporación Nacional Forestal (CONAF) in Chile kindly granted permission to collect there and the Olaf Grolle Olsen Fund generously gave financial support.

References

  • Alstrup V, Søchting U (2011) Massalongia olechiana (Massalongiaceae, Peltigerales), a new lichen species from the Antarctic. Polish Polar Research 32: 117–121. https://doi.org/10.2478/v10183-011-0011-y
  • Castello M, Nimis PL (1995) A critical revision of Antarctic lichens described by C. W. Dodge. Nova Hedwigia 57: 71–92.
  • Dickson J (1790) Fasciculus Secundus Plantarum Cryptogamicarum. Britanniae, London, 29 pp.
  • Dodge CW (1968) Lichenological note on the flora of the Antarctic continent and the subantarctic islands VIII. Nova Hedwigia 15: 435–502.
  • Dodge CW (1971) Lichenological notes on the flora of the Antarctic continent and the subantarctic islands IX-XI. Nova Hedwigia 19: 439–502.
  • Döring H, Clerc P, Grube M, Wedin M (2000) Mycobiont-specific PCR primers for the amplification of nuclear ITS and LSU rDNA from lichenized Ascomycetes. The Lichenologist 32: 200–204. https://doi.org/10.1006/lich.1999.0250
  • Ekman S, Wedin M, Lindblom L, Jørgensen PM (2014) Extended phylogeny and a revised generic classification of the Pannariaceae (Peltigerales, Ascomycota). The Lichenologist 46: 627–656. https://doi.org/10.1017/S002428291400019X
  • Ertz D, Poulsen RS, Charrier M, Søchting U (2017) Taxonomy and phylogeny of the genus Steinera (Arctomiales, Arctomiaceae) in the subantarctic islands of Crozet and Kerguelen. Phytotaxa 324: 201–238. https://doi.org/10.11646/phytotaxa.324.3.1
  • Galloway DJ (1985) Flora of New Zealand lichens (1st edn.). Wellington.
  • Galloway DJ (2007) Flora of New Zealand lichens. Revised Second Edition Including Lichen- Forming and Lichenicolous Fungi. Manaaki Whenua Press, Lincoln.
  • Gyelnik V (1940) Pannariaceae. In: Rabenhorst L (Ed.) Kryptogamenflora Vol. 9, Div 2(2) Cyanophili, 135–272.
  • Henssen A (1963) The North American species of Massalongia and generic relationships. Canadian Journal of Botany 41: 1331–1346. https://doi.org/10.1139/b63-114
  • Jørgensen PM (1986) Macrolichens of Bouvetøya. Norsk Polarinstitutt Skrifter 185: 23–34.
  • Jørgensen PM (2000) Notes on some South-Asian species of the lichen genus Fuscopannaria. Journal of the Hattori Botanical Laboratory 89: 247–259.
  • Jørgensen PM (2007) Massalongia in Nordic Lichen Flora 3: 88–89.
  • Jørgensen PM, Elix JA (1988) Additional lichen records from Australia. I Massalongia, a lichen genus new to Australia. Australasian Lichenological Newsletter 22: 23–34.
  • Jørgensen PM, Andersen HL (2015) The lichen genus Psoromidium (Pannariaceae) re- evaluated, with nomenclatural notes on Degeliella and Psoromaria. The Lichenologist 47(5): 343–348. https://doi.org/10.1017/S0024282915000171
  • Katoh K, Misawa K, Kuma K-I, Miyata T (2002) MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform. Nucleic Acids Research 30: 3059–3066. https://doi.org/10.1093/nar/gkf436
  • Katoh K, Standley DM (2013) MAFFT Multiple Sequence Alignment Software version 7: Improvements in performance and usability. Molecular Biology and Evolution 30: 772–780. https://doi.org/10.1093/molbev/mst010
  • Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrocks S, Buxton S, Cooper A, Markowitz S, Duran C, Thierer T, Ashton B, Mentjies P, Drummond A (2012) Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 28: 1647–1649. https://doi.org/10.1093/bioinformatics/bts199
  • Körber GW (1855) Systema Lichenum Germaniae. Breslau.
  • Lamb IM (1958) La vegetacion liquenica de los parques nacionales patagonicos. Anales de parques nacionales VII.
  • Lindsay DC (1974) The Macrolichens of South Georgia. Cambridge, British Antarctic Survey, 91 pp. [British Antarctic Survey Scientific Reports, 89]
  • Liu LN, Razaq A, Atri NS, Bau T, Belbahri L, Chenari Bouket A, Chen L-P, Deng C, Ilyas S, Khalid AN, Kitaura MJ, Kobayashi T, Li Y, Lorenz AP, Ma Y-H, Malysheva E, Malysheva V, Nuytinck J, Qiao M, Saini MK, Scur MC, Sharma S, Shu L-L, Spirin V, Tanaka Y, Tojo M, Uzuhashi S, Valério-Júnior C, Verbeken A, Verma B, Wu R-H, Xu J-P, Yu Z-F, Zeng H, Zhang B, Banerjee A, Beddiar A, Bordallo JJ, Dafri A, Dima B, Krisai-Greilhuber I, Lorenzini M, Mandal R, Morte A, Nath P.S, Papp V, Pavlík J, Rodríguez A, Ševcˇíková H, Urban A, Voglmayr H, Zapparoli G (2018) Fungal Systematics and Evolution: FUSE 4. Sydowia 70: 211–286.
  • Matheny PB, Liu YJ, Ammirati JF, Hall BD (2002) Using RPB1 sequences to improve phylogenetic inference among mushrooms (Inocybe, Agaricales) American Journal of Botany 89: 688–698. https://doi.org/10.3732/ajb.89.4.688
  • Muggia L, Nelson P, Wheeler T, Yakovchenko LS, Tønsberg T, Spribille T (2011) Convergent evolution of a symbiotic duet: the case of the lichen genus Polychidium (Peltigerales, Ascomycota). American Journal of Botany 98: 1647–1656. https://doi.org/10.3732/ajb.1100046
  • Nylander JAA (2004) MrAIC.pl. Program distributed by the author. Evolutionary Biology Centre, Uppsala University.
  • Redon J (1985) Liquenes Antarcticos. Santiago de Chile.
  • Rehm H (1916) Ascomycetes philippinenses – VIII. Leaflets of Philippine Botany 8: 2935–2961.
  • Smith RIL, Corner RWH (1973) Vegetation of the Arthur Harbour-Argentine Islands Region of the Antarctic Peninsula. British Antarctic Survey Bulletins 33–34: 89–122.
  • Stiller JW, Hall BD (1997) The origin of red alga: Implications for plastid evolution. Proceedings of the National Academy of Sciences of the United States of America 94: 4520–4525. https://doi.org/10.1073/pnas.94.9.4520
  • Øvstedal DO, Gremmen NJM (2010) New Lichen species from Tristan da Cunha and Gough Island. Folia Cryptogamica Estonica 47: 43–49.
  • Øvstedal DO, Smith RIL (2001) Lichens of the Antarctic Peninsula and South Georgia. Cambridge.
  • Vilgalys R, Hester M (1990) Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. Journal of Bacteriology 172: 4238–4246. https://doi.org/10.1128/jb.172.8.4238-4246.1990
  • Wedin M, Jørgensen PM, Wiklund E (2007) Massalongiaceae fam. nov., an overlooked monophyletic group among the cyanobacterial lichens (Peltigerales, Lecanoromycetes, Ascomycetes). The Lichenologist 39: 61–67. https://doi.org/10.1017/S002428290700655X
  • Wedin M, Wiklund E, Jørgensen PM, Ekman S (2009) Slippery when wet: Phylogeny and character evolution in the gelatinous cyanobacterial lichens (Peltigerales, Ascomycota). Molecular Phylogenetics and Evolution 53: 862–871. https://doi.org/10.1016/j.ympev.2009.08.013
  • White TJ, Bruns T, Lee S, Taylor JW (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (Eds) PCR Protocols: A Guide to Methods and Applications. Academic Press, New York, 315–322. https://doi.org/10.1016/B978-0-12-372180-8.50042-1
  • Zahlbruckner A (1917) Botanische Ergebnisse der schwedischen Expedition nach Patagonien und dem Feuerlande 1907–1909. VI. Die Flechten. Kungliga Svenska Vetenskapsakademiens Handlingar 57: 1–62.
  • Zoller S, Scheidegger C, Sperisen C (1999) PCR primers for the amplification of mitochondrial small subunit ribosomal DNA of lichen-forming Ascomycetes. The Lichenologist 31: 511–516. https://doi.org/10.1006/lich.1999.0220