Biodiversity and ecology of lichens of Kenai Fjords National Park, Alaska

We inventoried lichens in Kenai Fjords National Park in Alaska, USA We assembled the known information on occurrence and ecology of lichens in this park by combining field, herbarium, and literature studies. Our results provide baseline data on lichen occurrence that may be used in resource condition assessments, vulnerability assessments, long-term ecological monitoring, and resource management. We report a total of 616 taxa of lichenized fungi from the Park, plus an additional five subspecies and three varieties, all of which are new additions to the National Park Service database for this park unit. An additional five species of nonlichenized lichenicolous fungi are reported here. Eight non-lichenized fungi that are traditionally treated with lichens are also included, most of these associated with bark of particular host species. Four taxa new to North America are reported here (Arctomia delicatula var. acutior, Aspicilia dudinensis, Myriospora myochroa, and Ochrolechia bahusiensis), along with 44 species new to Alaska. Numerous species have been confirmed using ITS barcoding sequences. Also several records assigned to the genus level are reported, many of those are likely new species.


Introduction
Lichens are a major component of the biodiversity and function of high latitude ecosystems. Lichens are highly sensitive to environmental conditions, including airborne contaminants, substrate chemistry, and climate (Root et al. 2014). Such attributes make them useful indicators of species richness (Bergamini et al. 2005) and air quality, including the estimation of critical loads (Geiser & Neitlich 2007;Geiser et al. 2010). Although an ecologically important and conspicuous component of the vegetation in Alaska (e.g., Joly et al. 2003;Nelson et al. 2013Nelson et al. , 2015, lichens are a poorly known component of Kenai Fjords National Park (KEFJ or 'Kenai Fjords'). Located in the northern Gulf of Alaska, in western Prince William Sound, this park is managed by the National Park Service (NPS). Ecosystems in this region of south-central Alaska face an uncertain future of climate change effects, including effects on forest and riparian health (Werner et al. 2006;Ruess et al. 2009;Sherriff et al. 2011) and potential new resource development (Montgomery et al. 2003;Szumigala et al. 2010;Hite & Stone 2013). In Kenai Fjords, gold was discovered in 1918 in areas that would later become park land (Lanik et al. 2018). Mining activity, centered primarily on the Nuka Bay historic mining district, peaked in the 1930s (Richter 1970). The effects of mining are still felt today, and include both the mitigation of abandoned mines on park land, and the potential for future mineral development on non-federally owned lands (inholdings) within the park (Lanik et al. 2018). Documenting biodiversity is a first step in conserving biodiversity. This was the goal of this first comprehensive inventory of the lichens of Kenai Fjords. Krog (1968) sampled intensively in southeast Alaska and extreme western Alaska. She reported many interesting finds from the Aleutian Islands and the Bering Strait region, but made relatively few collections near Kenai Fjords and apparently none within the Park. The nearby sites that she visited were Marathon Mountain, west of Seward (elevations 62-900 m) and Seward, on the east and west sides of Resurrection Bay and in the forested area behind Seward Sanatorium, near sea level (Krog 1968, p. 27). Some of Krog's collections from Alaska have not been accessioned or fully identified, but are stored in Oslo (O;E. Timdal pers. comm. 2016), so at present we cannot provide a full accounting of her specimens from near Seward. A partial database of her collections provided by Einar Timdal, however, shows 17 specimens from Marathon Mountain and 22 specimens from Seward, all macrolichens. Walton et al. (2014) sampled epiphytic lichens in mature forest stands in Kenai Fjords National Park in 2012 and 2013 at 11, 0.38 ha plots (Appendix 2) using the standard Forest Inventory and Analysis protocol (USDA 2010). In addition, they collected specimens opportunistically in a range of other habitats and on other substrates. Their results have been summarized separately (92 lichen taxa; Walton et al. 2014), and the epiphyte data were included in studies of epiphytic lichen communities in relation to climate (Smith et al. 2017(Smith et al. , 2020. We incorporated these plot records into our database and their findings are integrated with ours in this paper. The most comprehensive studies of lichen diversity near Kenai Fjords National Park are McCune et al. (2018) from Katmai and Lake Clark National Parks, Spribille et al. (2010) from Klondike Gold Rush National Historic Park, and forthcoming studies from Glacier Bay National Park (Spribille et al. 2020). North and east of the study area Stehn et al. (2015) compiled a list of lichens of the Denali National Park region. In addition, Thomson (1984Thomson ( , 1997 included numerous records from the Kenai Peninsula, with some dots on his maps appearing near Seward. Farther south along the coast, Brodo and coauthors have critically examined numerous genera from Haida Gwaii in British Columbia (e.g., Brodo 1995Brodo , 2010Brodo & Ahti 1996;Brodo & Santesson 1997) We assembled a group of lichenologists to inventory lichens at Kenai Fjords National Park with the goals of (i) expanding the list of species known to occur in the park; (ii) compiling a reference collection of voucher specimens and associated habitat data; and (iii) compiling the associated geospatial data into a geodatabase.
Some of the results of this study have already been reported elsewhere (Fryday & Tønsberg 2015;Tønsberg 2016;Knudsen & Kocourková 2017;McCune 2018;McCune et al. 2019) and more are forthcoming. The purpose of the current publication is to present a comprehensive treatment of the lichens of Kenai Fjords National Park, bridging a gap in our knowledge of the lichen flora in south-central Alaska.

Study area
Kenai Fjords National Park ( Fig. 1) was established in 1980 through the Alaska National Interest Land Conservation Act (ANILCA) for the purpose of maintaining 'unimpaired the scenic and environmental integrity of the Harding Icefield, its outflowing glaciers, and coastal fjords and islands in their natural state (ANILCA 1980). Situated on Alaska's Kenai Peninsula, the park spans an area reaching from Resurrection Bay in the northeast to the Grewingk-Yalik Glacier Complex in the southwest. The Kenai Mountains form the western boundary of the park, with elevations ranging from sea level to 1996 m.
The climate is subpolar oceanic (Köppen-Geiger climate zone Cfc; Peel et al. 2007), implying a temperate climate without a dry season and with a cold summer. Only one long-term weather station exists near the Park, in the town of Seward, near sea level. Mean annual temperature at Seward, Alaska is 2.2°C, and total annual precipitation is 1712 mm (Seward 8 NW, AK, elevation 125 m;1981-2010; https://www.ncdc.noaa.gov/cdo-web/ datatools/normals).
The bedrock of Kenai Fjords consists primarily of the Valdez Group, consisting of Upper Cretaceous sandstone, siltstone, shale and minor conglomerate (Lanik et al. 2018). Areas of pillow basalt interbed with clastic sedimentary rocks. The southern and western portion of the park includes the McHugh Complex, consisting late Jurassic to Early Cretaceous rock, including greywacke, conglomerate, basalt, chert, gabbro, ultraplutonic rocks (e.g., granite), and limestone (Lanik et al. 2018). Surficial rocks in the park are dominated by metamorphosed sedimentary and granitic rocks. Calcareous rocks are rare and inaccessible.
The park has experienced three major intervals of glacial expansion in the late Holocene. Glacial advances occurred 3,600 years BP, in 600 A.D., and during the Little Ice Age (1300-1850 A.D.; Calkin et al. 2001). As of 2005, glacier cover in the park was 2,074 km 2 , roughly 1,800 km 2 of which was taken up by the Harding Icefield (Loso et al. 2014).
Recent deglaciation has exposed large areas that have been colonized by tall shrubs, primarily Sitka alder (Alnus viridis ssp. sinuata) and willow (Salix sitchensis) (Boggs et al. 2008). Alpine tundra grows on the higher ice-free ridges, transitioning to ericaceous heaths and Tsuga mertensiana krummholz in the subalpine. Towering rock walls along the ocean are cut by steep chutes and fringed with mature Sitka spruce (Picea sitchensis) and/or mountain hemlock (Tsuga mertensiana), and tall shrubs. Steep-sided glacial valleys penetrate from the mountains to the sea. The cold, wet climate results in Sphagnum peatlands at the lower elevations, even on steep slopes underlain by compact till or bedrock. These peatlands cover roughly 1% of the park, but support multiple sedge species (Carex spp.), cottongrass (Eriophorum spp.), ericaceous shrubs, and woodlands of dwarf Tsuga mertensiana or Picea sitchensis (Boggs et al. 2008). Low-elevation floodplains and benches are likewise dominated by P. sitchensis, T. mertensiana, and shrubs, with pockets of balsam poplar (Populus balsamifera) in some riparian areas. The 1964, the Great Alaska Earthquake (magnitude 9.2) resulted in 1.0-2.5 m of subsidence along the Kenai Fjords coastline, along with numerous landslides, widespread flooding and salt water incursion into previously forested areas (Lanik et al. 2018). Dead standing trees ('ghost forests') now occupy low-lying areas in the North and West Arm of Nuka Bay (Beauty Bay) and in McCarty Fjord (James Lagoon). Low-elevation mature and old-growth coniferous forest covers roughly 16% of the park, while tall shrubs, primarily alder, occupy roughly one quarter of the landscape (Boggs et al. 2008).

Materials and methods
We evaluated existing collections by NPS personnel for Kenai Fjords and surroundings; identified habitat and taxon gaps in the data; and conducted field inventories to fill gaps. In addition, we reviewed the existing literature, and to the extent possible, the specimens supporting that literature.
Although Kenai Fjords National Park is close to the city of Anchorage, most of it is inaccessible except by seaworthy boats and helicopter. The area of the Park close to the town of Seward probably has incidental collections by a number of lichenologists, though we have found only two of these by searching online collection databases. Near the town of Seward, only the Exit Glacier area is within the National Park. Indeed, that is the only area of the park that has frequent visitors.
We used subjective judgment to focus new surveys in areas where high diversity was expected. This method attempts to maximize rapid discovery of species diversity and is relatively cost-effective, but sacrifices park-level quantitative inference. Field observers and laboratory examiners included diverse taxonomic specialists in an effort to maximize the reliability and representativeness of important taxonomic groups. Data sources, including literature and new observations, are summarized in Table 1.
Our sampling design met the following criteria: (i) sampling occurred across an elevation gradient, from sea level to alpine, (ii) sampling occurred in a range of habitats, and (iii) voucher specimens were collected at each site, except for common macrolichens, unless precluded by low population size. Geologic maps, landcover maps and satellite imagery were used to identify accessible areas meeting those criteria. We chose to try to maximize species discovery rates rather than using a fixed area, fixed time, or otherwise equal effort among sites. The tradeoff for that choice is that the rigorous statistical comparisons of diversity are then impossible, because diversity estimates and community statistics are strongly affected by sample area and effort (e.g., McCune & Grace 2002, p. 27). While corrections to equal effort can be attempted, they all require tenuous assumptions with unknown effects on the conclusions.
In 2015, sites at Kenai Fjords (Appendix 1) were visited by boat by McCune, Rosentreter, Schultz, Tønsberg, and Walton. In addition, the team explored the Exit Glacier area near Seward by foot. Each collector focused on particular groups of lichens. Other authors contributed by examining specimens within their specialty. In addition to the sites visited by this group, we include many collections made by Walton and Hutten as part of their preliminary survey of lichens and bryophytes in the Park (Walton et al. 2014).
Locality information -Detailed locality information is given for the group sites (Appendix 1) and for specimens of particular interest. Other locality data are available from the first author or the NPS Southwest Alaska Network office in Anchorage (Park abbreviations: Table 2).
Abundance ratings -Abundance ratings are based on our experience and limited sampling. Abundance ratings are necessarily subjective and not given when there is little basis for doing so. For example, a single occurrence of an inconspicuous species may represent an overlooked common species or a truly rare species. The more conspicuous a species, the easier it is to state an abundance.
Noteworthy collections are listed individually. When one specimen is cited, all specimens of that species are cited, unless otherwise noted. When summarized verbally without citing individual specimens, we applied the following frequency classes: -very common (> 40 collections) -common (10-40 collections) -occasional (3-9 collections) -uncommon to rare (1-2 collections) Supplemental data such as anatomical details are reported for collections where the additional information may be helpful in either confirming unusual records or where the observations conflict or augment existing descriptions.
Chemistry is reported when thin-layer chromatography (TLC) results were available and the information was considered significant by us, either in separating the species from its relatives, in validating the species report, or in supplementing the known information about a species. In general, TLC protocols followed methods of Culberson & Kristinsson (1970), Culberson (1972), and the later modification by Culberson & Johnson (1982). All three solvent systems were used (A, B' and C) in most cases by Tønsberg. McCune used B' and C for Lecideaceae s.l. and Cladonia, A and B' for Umbilicaria, and A and C for most other genera.
DNA sequences were obtained for selected critical specimens and for many collections in particular groups, especially Lecideaceae, Teloschistaceae, Stereocaulon, and Umbilicaria. Various protocols were followed, differing by laboratory, as described in previous publications by various authors of this paper (e.g., Arup et al. 2015;Miadlikowska et al. 2018;McCune et al. 2019).
In each case where we report DNA sequences, the following procedure was generally used (with some variations) to place our specimens into the context of existing sequences. After an initial comparison with existing sequences using BLASTn, we created an alignment containing our new sequences and relevant existing sequences and several outgroup sequences. Sequences were aligned with MAFFT in Geneious (Kearse et al. 2012) using default settings (Auto algorithm selection), then adjusted manually when necessary. We then constructed phylogenetic trees, initially with neighbor joining methods to help refine the selection of sequences, then realignment and maximum likelihood analysis with 500 or 1000 bootstrap runs with the PhyML plugin (Guindon et al. 2010) to Geneious. We used PhyML with default The primary set of voucher specimens is housed at the NPS herbarium in Anchorage and in Fairbanks, Alaska (ALA), with portions of the collection housed at institutional herbaria of the authors while still under study (usually by the collector) or through loan agreements between institutions and the NPS. Individual collections were compiled in a Microsoft Access database that was then imported and archived into the NPS database. These data are available from the NPS on request.
For the most part, generic placement follows the most recent North American checklist (Esslinger 2019). Exceptions include cetrarioid lichens (Divakar et al. 2017) and cases where generic splits are not well supported by the data, insufficiently studied, or where authors have treated only some of the species from our region, making our species difficult to assign to genera in a consistent way. In these incomplete cases, we have retained a broader generic concept. Examples include Aspicilia and its segregates Circinaria and Sagedia, Lecanora and its segregates including Glaucomaria, Lecanoropsis, Protoparmeliopsis, and parts of Teloschistacaeae. Verrucaria determinations are by Breuss unless otherwise noted.
In most cases, material identified only to genus or tentatively to genus is omitted from the following list. We do, however, include some relatively distinctive specimens that could not be assigned to species, hoping to improve the chance that they might be included in future treatments of the genus.

Results and discussion
We recorded a total of 625 taxa of lichenized fungi from within or immediately adjacent to Kenai Fjords National Park (Table 3). This total includes 617 lichenized species, plus an additional five subspecies and three varieties. We did not attempt to represent nonlichenized, lichenicolous fungi, but recorded five of those. An additional eight species are apparently nonlichenized and nonlichenicolous, but traditionally treated with lichens. Four records of lichens are questionable including historical reports where a formerly broad species concept has narrowed substantially, creating uncertainty for existing records from Kenai Fjords.
In addition to the reports from Kenai Fjords, we include in a separate section at the end of the list of taxa, supplemental information for 14 taxa from nearby Katmai and Lake Clark National Parks, beyond that provided by McCune et al. (2018).
One species, Acarospora toensbergii, was described from our collections in the study area as new (Knudsen & Kocourková 2017) and two, Biatora troendelagica and Jamesiella scotica, were reported as new to North America, in previous papers (Tønsberg 2016;Tønsberg & Printzen 2018).

New to Alaska
Forty six species are new to Alaska, based on comparison of our results with an unpublished list for Alaska (Spribille et al. in prep.) and other recent publications not included in that compilation.

Rare species in Alaska
Of the 64 lichen species currently listed as 'rare' by the Alaska Center for Conservation Science (AKNHP 2015), we found 20% (13) in Kenai Fjords National Park. The list clearly needs to be updated, based on much lichenological work in Alaska since the list was created. No federally listed lichens are present in Alaska, but we presume this reflects the difficulties of the federal listing process for lichens and our lack of information rather than the occurrence of rare lichens (Allen et al. 2019).

Potential factors influencing lichen biodiversity
Compared to the interior side of the Kenai Peninsula and to Katmai and Lake Clark National Parks, Kenai Fjords has a distinctly oceanic climate and corresponding floristic composition. We recorded a lichen biota with a mix of arctic-alpine, boreal, and coastal elements. The Beringian element, which is evident on the Seward Peninsula and Aleutian Islands (e.g., Krog 1968;McCune 2008), was not found at Kenai Fjords. Slope bogs with rock outcrops provide a distinctive habitat rich in lichen species. These support Tsuga mertensiana, ericaceous shrubs, and Sphagnum with boggy slopes and ledges. Moist organic mats are colonized by lichens such as Dibaeis baeomyces and many species of Cladonia.
Similar to Katmai and Lake Clark National Parks (McCune et al. 2018), alpine sites are species rich, but lack many genera or species associated with interior alpine sites, such as Dactylina, Hypogymnia, and Rinodina.
Nitrophilous species (e.g., Caloplaca, Polycauliona, and Xanthoria) were sparse and very local, presumably associated with manuring by birds and other animals. We also encountered very few calciphiles, owing to the predominantly acidic rocks.
Much of the park has been heavily glaciated; in fact, much of the mountain mass is covered by a continuous ice sheet. The ice is penetrated by a number of nunataks (isolated peaks or ridge surrounded by extensive persistent icefield). Although access to these is very difficult, we did manage to sample two nunataks, as well as a few other subalpine and alpine sites. Whether or not these modern nunataks were ice free during the last glacial period is unknown. Vegetation sampling on modern nunataks (Miller et al. 2006) revealed a number of rare and disjunct vascular plant species. While that work listed a number of lichen species, it did not attempt a detailed inventory of lichens. In any event, we made an effort to include nunataks because of the possibility that they might be long-term refugia for alpine species. Given the uncertainties in the glacial history of the alpine sites and the small number that we visited, we cannot make firm conclusions regarding the biogeographic importance of nunataks, but we can say that numerous species were found at the alpine sites that were not recorded elsewhere, as described in the list of taxa.

Annotated list of taxa
The following list of taxa is annotated by substrate, frequency of occurrence, taxonomic notes, and type of record (  Arup (unpubl.) showed that this specimen belongs to the symmicta clade, perhaps even in a narrow sense. However, this group is not well understood and more taxa may be present within it. Closest to the sequence of this specimen are two sequences from GenBank named L. confusa, one from North America (GU480093) and one from Scotland (GU480120), but these are not L. confusa in a strict sense. Lecanora zosterae (Ach.) Nyl. -Near mouth of creek at north end of James Lagoon, on wood, exposed snag, Schultz 16819.   Lendemer's (2013b) data showed that a 1:1 correspondence between chemotypes and species is untenable in the L. neglecta group, it is clear that the chemotypes are phylogenetically structured: many supported clades are pure or nearly pure in chemotype (Lendemer 2013b, Fig. 4). Given the apparent diversity in both morphology and chemistry, as well as strong phylogenetic structure in this group, we prefer to continue to track both chemotypes and morphotypes in North America, pending systematic study with more molecular markers. Lepraria aff. borealis Loht. & Tønsberg -Part of Lepraria neglecta s.l., see Table 4. creek at north end of James Lagoon, on Salix, Tønsberg 45342 (ITS sequence, GenBank MN906302). The name for this will change soon because type of L. cookii, as given in Stone et al. (2016), was mistaken and resulted in L. cookii being a synonym of L. saturninum. The species concept as described for L. cookii will receive a new name (D. Stone, pers. comm. 2020 Stone et al. (2016) and confirmed by us, L. saturninum is polyphyletic, even after four new species were segregated, leaving two main clades of L. saturninum. The specimen we sequenced (Schultz 16702b) falls in the clade with the Scottish epitype (Stone et al. 2016, p. 415)   Hole Bay off Aialik Bay, on trunk of Tsuga mertensiana, Tønsberg 45528a (TLC: gyrophoric (major) and lecanoric acids; one apothecium, but not very well developed); on base of Picea sitchensis, Tønsberg 45534 (TLC: gyrophoric (major) and lecanoric acids); peninsula into Three Hole Bay off Aialik Bay, on trunk of Tsuga mertensiana, Tønsberg 45512 (this specimen also cited by Spribille et al. (2020); TLC: gyrophoric (major) and lecanoric acids; isidiate, sterile), Tønsberg 45513. The recently described species is distinctive in having coralloid isidia (Spribille et al. 2020 Thomson and Ahti (1994 bay on SW side of Portage Lake, on ± hard wood of Picea glauca on slope facing lake, Tønsberg 43918 (TLC: stictic and confriesiic acids, the latter missing or in low concentrations in one split of this specimen). Two ITS sequences (GenBank MN906296, MN906297) from different part of the same specimens, but similar morphologies -both sequences essentially identical.