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THE IDENTITY OF STICTA LIVIDA KREMP
Lichenologist 32(2): 197–200 (2000) doi:10.1006/lich.1999.0244 Available online at http://www.idealibrary.com on
SHORT COMMUNICATIONS THE IDENTITY OF STICTA LIVIDA KREMP. In the recent revision of Sticta (Schreb) Ach. in New Zealand (Galloway 1997), the taxon S. livida Kremp., collected by Charles Knight and described by Krempelhuber (Krempelhuber 1876) was excluded from the New Zealand flora since examination of a wide range of New Zealand material of Sticta, both in the field and in the herbarium over many years had failed to turn up any additional specimens. It was suggested that the material described by Krempelhuber might have been collected by Knight in Australia, as it has some resemblance to the Australian taxon S. camarae Müll. Arg. (Galloway 1998b), and Knight was known to have travelled several times in northern Australia in the 1880s subsequent to his retirement (Galloway 1998a). However, in a parcel of northern New Zealand lichens recently sent from AK for identification, two unidentified specimens of Sticta collected from offshore islands north and east of Auckland proved to be S. livida. Thus the occurrence of this taxon in New Zealand is unambiguously confirmed. A description and notes are given below, together with a revised key to Sticta in New Zealand. Sticta livida Kremp Verhandl. zool.-bot. Ges Wien 26: 448 (1876); type: Nova Zelandia [New Zealand], sine loco, C. Knight (W!—lectotype, fide Galloway 1997: 162).
Thallus 2–8(–10) cm diam.,loosely attached, with occasional small attaching fascicles of rhizines, margins and apices free, straggling in irregular patches, amongst bryophytes or overgrowing crustose lichens. Lobes rather narrow, 2–3(–5) mm wide, 0·5–2·5 cm long, subdichotomously branching to irregularly laciniate, discrete at margins, overlapping-complex centrally. Margins entire, slightly thickened above and below, sinuses smoothly rounded, without isidia or soredia, occasionally secondarily minutely lobulate, here and there with a thin fringe of projecting pale tomentum from the lower surface visible. Upper surface olive-green when moist, pale buff or yellow-brown to grey-fawn when dry, matt, minutely papillate (10 lens), irregularly wrinkled in parts, elsewhere undulate or smooth, rather thin, brittle when dry, pliable when moist, without isidia, maculae, phyllidia, pseudocyphellae or soredia. Photobiont green. Medulla white (K). Lower surface uniformly pale buff or tan, not darkening centrally, smooth or shallowly undulate, not conspicuously wrinkled, evenly tomentose from margins to centre, or occasionally with narrow glabrous patches at margins, tomentum thin, even, velvety, pale buff or tan, occasionally projecting at margins as a pale fringe. Cyphellae widely spaced, infrequent, pin-prick-like at margins, elsewhere shallowly to deeply crateriform, 0·1–0·2 (rarely up to 0·4) mm diam., round to somewhat 0024–2828/00/020197+08 r35.00/0
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irregular, margins thin, raised, projecting only slightly from tomentum, pit membrane smooth, creamish to pale buff. Pycnidia not seen. Thallus 150–400 m thick. Upper cortex 30–35 m thick, with a pale brownish upper zone, 10–12·5 m thick, and a colourless inner zone 15–25 m thick, of thick-walled, isodiametric cells 7·5–10 m diam. Photobiont zone up to 25 m thick, photobiont cells 5–7 m diam. Medulla 90–200(–300) m thick, of loosely interwoven, colourless hyphae, 3–5 m diam. Lower cortex up to 25 m thick, pale brownish, similar in cell structure to upper cortex. Rhizines projecting from lower cortex, cells septate, 4–6 m diam., 50–75 m long, colourless to pale brownish, rather thick-walled, clustered in fascicles of 2–6 rhizines. Apothecia not seen. Distribution. Known only from the northern offshore islands (Great Mercury Island, Little Barrier Island) north of Lat. 36S. Apparently an endemic species (it is not known from Australia). It is still very poorly collected in New Zealand. Habitat ecology. A corticolous taxon known from the bark of kanuka (Kunzea ericoides) and rewarewa (Knightia excelsa) in northern coastal forest, from sea level to 100 m. Distinguishing features. Sticta livida is characterized by a white medulla (K), a green photobiont, rather narrow, subdichotomously branching lobes with entire margins, and prominent, smoothly rounded sinuses, and here and there with small, projecting secondary lobules. The lower surface is pale buff and evenly short-tomentose with occasional scattered pin-prick-like to crateriform cyphellae. It is superficially similar to the Australian species S. camarae, but has much smaller, narrower, and more densely branching lobes, has an even, thinly tomentose lower surface, and lacks an attaching stalk and holdfast, which are characteristic of S. camarae. Specimens examined. New Zealand: North Auckland: Little Barrier Island, Track 17, on bark of kanuka in ridge-top forest, 9 May 1990, A. E. Wright 10002 (AK 193458). South Auckland: Great Merury Island, Peachgrove, on Rewarewa (Knightia excelsa) bark, May 1975, B. W. Hayward H 40.92 (AK 230721).
Revised key to species of Sticta in New Zealand 1
Photobiont green; thallus bright green or olive-green when wet . 2 Photobiont cyanobacterial; thallus dark blue-grey, olivaceous brownish to grey-black when wet . . . . . . . . . . . . . . . . . . . 11
2(1)
Thallus attached to substratum by a holdfast; lobes arising from a pronounced stalk . . . . . . . . . . . . . . . . . . . . . . . . . 3 Thallus not attached to substratum by a holdfast, attached over whole of lower surface; lobes not stalked . . . . . . . . . . . . 5
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3(2)
Thallus over 4 cm long, erect, attaching stalk long, discrete; lower surface pubescent or tomentose . . . . . . . . . . . . . . . . . 4 Thallus 1–2 cm long, entangled-spreading, decumbent, fragile, attaching stalk very short, often proliferating; lower surface glabrous . . . . . . . . . . . . . . . . . . . . . . . . S. lacera
4(3)
Thallus usually over 10 cm long; margins entire . . . S. latifrons Thallus 4–8 cm long; margins lacerate-incised . . . . . . . S. filix
5(2)
Without isidia or phyllidia . . . . . . . . . . . . . . . . . . . . . 6 Isidiate or phyllidiate . . . . . . . . . . . . . . . . . . . . . . . . 9
6(5)
Apothecia usually present (sometimes absent); thalli corticolous or saxicolous . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 Apothecia absent; thalli terricolous in subantarctic/subalpine grassland . . . . . . . . . . . . . . . . . . . . . . . . . . . S. colinii
7(6)
Lobes broadly rounded, 5–20(–25) mm broad, not dichotomously branching . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 Lobes narrow, 2–4(–5) mm broad, dichotomously branching . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S. livida
8(7)
Apothecial disc orange; ascospores 1–3-septate . . S. subcaperata Apothecial disc red to red-brown; ascospores 5–7-septate . . . . . . . . . . . . . . . . . . . . . . . . . . S. cinereoglauca
9(5)
Phyllidiate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 Isidiate, isidia dense, marginal and laminal, squamiform-imbricate . . . . . . . . . . . . . . . . . . . . . . . . . . . . S. squamata
10(9)
Cyphellae orange, pinkish or yellow; lower surface dark . . . . . . . . . . . . . . . . . . . . . . . . . . . S. babingtonii Cyphellae white; lower surface pale . . . . . . . . . . S. martinii
11(1)
Thallus broadly attached, without a stalk . . . . . . . . . . . . 12 Thallus palmate, attached by a narrow,terete stalk; margins isidiate . . . . . . . . . . . . . . . . . . . . . . . . S. caliginosa
12(11) Sorediate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 Isidiate . . . . . . . . . . . . . . . . . . . . . . . . . S. fuliginosa 13(12) Lobes cochleate or monophyllous; soralia marginal and laminal in erose patches, soredia not derived from isidia . . . S. limbata Lobes irregularly spreading not monophyllous; soralia linear, eroding lower surface of margins, never laminal, soredia derived from minute, granular to coralloid isidia . . . . . . S. sublimbata I am indebted to Doug Rogan, Botanical Curator at AK for the loan of material for study. Funds for this research were provided by the Foundation for Research, Science and Technology (FORST, Wellington, New Zealand) under Contract C09618. R Galloway, D. J. (1997) Studies on the lichen genus Sticta (Schreber) Ach. IV. New Zealand species. Lichenologist 29: 105–168.
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Galloway, D. J. (1998a) Joseph Hooker, Charles Knight, and the commissioning of New Zealand’s first popular flora: Hooker’s Handbook of the New Zealand Flora (1864–1867). Tuhinga 10: 31–62. Galloway, D. J. (1998b) Studies on the lichen genus Sticta (Schreber) Ach.: V. Australian species. Tropical Bryology 15: 117–160. Krempelhuber, A. von (1876) Neue Beiträge zur Flechten-Flora Neu Seelands. Verhandlungen der Zoologisch-Botanischen Gesellschaften in Wien 26: 447–460.
D. J. Galloway* *Landcare Research New Zealand Ltd, Private Bag 1930, Dunedin, New Zealand.
MYCOBIONT-SPECIFIC PCR PRIMERS FOR THE AMPLIFICATION OF NUCLEAR ITS AND LSU rDNA FROM LICHENIZED ASCOMYCETES Lichen thalli are composed of several genetically distinct organisms: a fungus, and one or several green algal or cyanobacterial species. Any DNA amplification process from lichen thalli must minimize the risk of targeting the wrong DNA. A simple way to achieve this is to use polymerase chain reaction (PCR) primers with enhanced specificity for the lichen mycobiont. Mycobiontspecific PCR primers for the nuclear small subunit (SSU) of the ribosomal DNA (rDNA) were designed by Gargas & Taylor (1992), and have enabled several investigations into lichen evolution and higher-level phylogeny (e.g. Gargas et al. 1995; Wedin et al. 1998). The SSU rDNA is, however, not variable enough for studies on lower taxonomic levels (Wedin & Döring 1999). Within the nuclear ribosomal gene cluster, the internal transcribed spacers (ITS) region (ITS1 and/or ITS2) has been extensively used for lower level phylogenies, species delimitation, and population studies in nonlichenized fungi, with some recent examples from lichen studies (e.g. Lohtander et al. 1998; Arup & Grube 1998). Only a few fungal-specific primers, primarily designed for studies of mycorrhiza, are available for ITS amplifications from lichens (ITS1F, Gardes & Bruns 1993; NL6A, Egger 1995). In our experience, amplification of the ITS region using these primers or some of the non-green algal primers located in the SSU rDNA, is still often non-specific with poor or multiple products. The nuclear large subunit rDNA (LSU), although more variable than the SSU, and extensively used for higher- and lower-level phylogenies of basidiomycetes and yeasts, have not yet been widely utilized in studies of ascomycete lichens. This is most likely due to the lack of suitable, fungal-specific PCR primers, and lack of relevant LSU sequence data in public databases. In order to enable studies into ecology and lower-level phylogeny of lichen-forming ascomycetes, we have designed specific PCR primers that do not amplify the ITS and LSU rDNA of lichen algae, and are located in the 5 part of the LSU. The primers were constructed using a manually adjusted MegAlign alignment (LaserGene, DNASTAR Inc.) of LSU sequences of representative
SHORT COMMUNICATIONS THE IDENTITY OF STICTA LIVIDA KREMP. In the recent revision of Sticta (Schreb) Ach. in New Zealand (Galloway 1997), the taxon S. livida Kremp., collected by Charles Knight and described by Krempelhuber (Krempelhuber 1876) was excluded from the New Zealand flora since examination of a wide range of New Zealand material of Sticta, both in the field and in the herbarium over many years had failed to turn up any additional specimens. It was suggested that the material described by Krempelhuber might have been collected by Knight in Australia, as it has some resemblance to the Australian taxon S. camarae Müll. Arg. (Galloway 1998b), and Knight was known to have travelled several times in northern Australia in the 1880s subsequent to his retirement (Galloway 1998a). However, in a parcel of northern New Zealand lichens recently sent from AK for identification, two unidentified specimens of Sticta collected from offshore islands north and east of Auckland proved to be S. livida. Thus the occurrence of this taxon in New Zealand is unambiguously confirmed. A description and notes are given below, together with a revised key to Sticta in New Zealand. Sticta livida Kremp Verhandl. zool.-bot. Ges Wien 26: 448 (1876); type: Nova Zelandia [New Zealand], sine loco, C. Knight (W!—lectotype, fide Galloway 1997: 162).
Thallus 2–8(–10) cm diam.,loosely attached, with occasional small attaching fascicles of rhizines, margins and apices free, straggling in irregular patches, amongst bryophytes or overgrowing crustose lichens. Lobes rather narrow, 2–3(–5) mm wide, 0·5–2·5 cm long, subdichotomously branching to irregularly laciniate, discrete at margins, overlapping-complex centrally. Margins entire, slightly thickened above and below, sinuses smoothly rounded, without isidia or soredia, occasionally secondarily minutely lobulate, here and there with a thin fringe of projecting pale tomentum from the lower surface visible. Upper surface olive-green when moist, pale buff or yellow-brown to grey-fawn when dry, matt, minutely papillate (10 lens), irregularly wrinkled in parts, elsewhere undulate or smooth, rather thin, brittle when dry, pliable when moist, without isidia, maculae, phyllidia, pseudocyphellae or soredia. Photobiont green. Medulla white (K). Lower surface uniformly pale buff or tan, not darkening centrally, smooth or shallowly undulate, not conspicuously wrinkled, evenly tomentose from margins to centre, or occasionally with narrow glabrous patches at margins, tomentum thin, even, velvety, pale buff or tan, occasionally projecting at margins as a pale fringe. Cyphellae widely spaced, infrequent, pin-prick-like at margins, elsewhere shallowly to deeply crateriform, 0·1–0·2 (rarely up to 0·4) mm diam., round to somewhat 0024–2828/00/020197+08 r35.00/0
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irregular, margins thin, raised, projecting only slightly from tomentum, pit membrane smooth, creamish to pale buff. Pycnidia not seen. Thallus 150–400 m thick. Upper cortex 30–35 m thick, with a pale brownish upper zone, 10–12·5 m thick, and a colourless inner zone 15–25 m thick, of thick-walled, isodiametric cells 7·5–10 m diam. Photobiont zone up to 25 m thick, photobiont cells 5–7 m diam. Medulla 90–200(–300) m thick, of loosely interwoven, colourless hyphae, 3–5 m diam. Lower cortex up to 25 m thick, pale brownish, similar in cell structure to upper cortex. Rhizines projecting from lower cortex, cells septate, 4–6 m diam., 50–75 m long, colourless to pale brownish, rather thick-walled, clustered in fascicles of 2–6 rhizines. Apothecia not seen. Distribution. Known only from the northern offshore islands (Great Mercury Island, Little Barrier Island) north of Lat. 36S. Apparently an endemic species (it is not known from Australia). It is still very poorly collected in New Zealand. Habitat ecology. A corticolous taxon known from the bark of kanuka (Kunzea ericoides) and rewarewa (Knightia excelsa) in northern coastal forest, from sea level to 100 m. Distinguishing features. Sticta livida is characterized by a white medulla (K), a green photobiont, rather narrow, subdichotomously branching lobes with entire margins, and prominent, smoothly rounded sinuses, and here and there with small, projecting secondary lobules. The lower surface is pale buff and evenly short-tomentose with occasional scattered pin-prick-like to crateriform cyphellae. It is superficially similar to the Australian species S. camarae, but has much smaller, narrower, and more densely branching lobes, has an even, thinly tomentose lower surface, and lacks an attaching stalk and holdfast, which are characteristic of S. camarae. Specimens examined. New Zealand: North Auckland: Little Barrier Island, Track 17, on bark of kanuka in ridge-top forest, 9 May 1990, A. E. Wright 10002 (AK 193458). South Auckland: Great Merury Island, Peachgrove, on Rewarewa (Knightia excelsa) bark, May 1975, B. W. Hayward H 40.92 (AK 230721).
Revised key to species of Sticta in New Zealand 1
Photobiont green; thallus bright green or olive-green when wet . 2 Photobiont cyanobacterial; thallus dark blue-grey, olivaceous brownish to grey-black when wet . . . . . . . . . . . . . . . . . . . 11
2(1)
Thallus attached to substratum by a holdfast; lobes arising from a pronounced stalk . . . . . . . . . . . . . . . . . . . . . . . . . 3 Thallus not attached to substratum by a holdfast, attached over whole of lower surface; lobes not stalked . . . . . . . . . . . . 5
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3(2)
Thallus over 4 cm long, erect, attaching stalk long, discrete; lower surface pubescent or tomentose . . . . . . . . . . . . . . . . . 4 Thallus 1–2 cm long, entangled-spreading, decumbent, fragile, attaching stalk very short, often proliferating; lower surface glabrous . . . . . . . . . . . . . . . . . . . . . . . . S. lacera
4(3)
Thallus usually over 10 cm long; margins entire . . . S. latifrons Thallus 4–8 cm long; margins lacerate-incised . . . . . . . S. filix
5(2)
Without isidia or phyllidia . . . . . . . . . . . . . . . . . . . . . 6 Isidiate or phyllidiate . . . . . . . . . . . . . . . . . . . . . . . . 9
6(5)
Apothecia usually present (sometimes absent); thalli corticolous or saxicolous . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 Apothecia absent; thalli terricolous in subantarctic/subalpine grassland . . . . . . . . . . . . . . . . . . . . . . . . . . . S. colinii
7(6)
Lobes broadly rounded, 5–20(–25) mm broad, not dichotomously branching . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 Lobes narrow, 2–4(–5) mm broad, dichotomously branching . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S. livida
8(7)
Apothecial disc orange; ascospores 1–3-septate . . S. subcaperata Apothecial disc red to red-brown; ascospores 5–7-septate . . . . . . . . . . . . . . . . . . . . . . . . . . S. cinereoglauca
9(5)
Phyllidiate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 Isidiate, isidia dense, marginal and laminal, squamiform-imbricate . . . . . . . . . . . . . . . . . . . . . . . . . . . . S. squamata
10(9)
Cyphellae orange, pinkish or yellow; lower surface dark . . . . . . . . . . . . . . . . . . . . . . . . . . . S. babingtonii Cyphellae white; lower surface pale . . . . . . . . . . S. martinii
11(1)
Thallus broadly attached, without a stalk . . . . . . . . . . . . 12 Thallus palmate, attached by a narrow,terete stalk; margins isidiate . . . . . . . . . . . . . . . . . . . . . . . . S. caliginosa
12(11) Sorediate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 Isidiate . . . . . . . . . . . . . . . . . . . . . . . . . S. fuliginosa 13(12) Lobes cochleate or monophyllous; soralia marginal and laminal in erose patches, soredia not derived from isidia . . . S. limbata Lobes irregularly spreading not monophyllous; soralia linear, eroding lower surface of margins, never laminal, soredia derived from minute, granular to coralloid isidia . . . . . . S. sublimbata I am indebted to Doug Rogan, Botanical Curator at AK for the loan of material for study. Funds for this research were provided by the Foundation for Research, Science and Technology (FORST, Wellington, New Zealand) under Contract C09618. R Galloway, D. J. (1997) Studies on the lichen genus Sticta (Schreber) Ach. IV. New Zealand species. Lichenologist 29: 105–168.
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Galloway, D. J. (1998a) Joseph Hooker, Charles Knight, and the commissioning of New Zealand’s first popular flora: Hooker’s Handbook of the New Zealand Flora (1864–1867). Tuhinga 10: 31–62. Galloway, D. J. (1998b) Studies on the lichen genus Sticta (Schreber) Ach.: V. Australian species. Tropical Bryology 15: 117–160. Krempelhuber, A. von (1876) Neue Beiträge zur Flechten-Flora Neu Seelands. Verhandlungen der Zoologisch-Botanischen Gesellschaften in Wien 26: 447–460.
D. J. Galloway* *Landcare Research New Zealand Ltd, Private Bag 1930, Dunedin, New Zealand.
MYCOBIONT-SPECIFIC PCR PRIMERS FOR THE AMPLIFICATION OF NUCLEAR ITS AND LSU rDNA FROM LICHENIZED ASCOMYCETES Lichen thalli are composed of several genetically distinct organisms: a fungus, and one or several green algal or cyanobacterial species. Any DNA amplification process from lichen thalli must minimize the risk of targeting the wrong DNA. A simple way to achieve this is to use polymerase chain reaction (PCR) primers with enhanced specificity for the lichen mycobiont. Mycobiontspecific PCR primers for the nuclear small subunit (SSU) of the ribosomal DNA (rDNA) were designed by Gargas & Taylor (1992), and have enabled several investigations into lichen evolution and higher-level phylogeny (e.g. Gargas et al. 1995; Wedin et al. 1998). The SSU rDNA is, however, not variable enough for studies on lower taxonomic levels (Wedin & Döring 1999). Within the nuclear ribosomal gene cluster, the internal transcribed spacers (ITS) region (ITS1 and/or ITS2) has been extensively used for lower level phylogenies, species delimitation, and population studies in nonlichenized fungi, with some recent examples from lichen studies (e.g. Lohtander et al. 1998; Arup & Grube 1998). Only a few fungal-specific primers, primarily designed for studies of mycorrhiza, are available for ITS amplifications from lichens (ITS1F, Gardes & Bruns 1993; NL6A, Egger 1995). In our experience, amplification of the ITS region using these primers or some of the non-green algal primers located in the SSU rDNA, is still often non-specific with poor or multiple products. The nuclear large subunit rDNA (LSU), although more variable than the SSU, and extensively used for higher- and lower-level phylogenies of basidiomycetes and yeasts, have not yet been widely utilized in studies of ascomycete lichens. This is most likely due to the lack of suitable, fungal-specific PCR primers, and lack of relevant LSU sequence data in public databases. In order to enable studies into ecology and lower-level phylogeny of lichen-forming ascomycetes, we have designed specific PCR primers that do not amplify the ITS and LSU rDNA of lichen algae, and are located in the 5 part of the LSU. The primers were constructed using a manually adjusted MegAlign alignment (LaserGene, DNASTAR Inc.) of LSU sequences of representative