Xylopsora canopeorum Timdal, Reese Næsborg & Bendiksby is described as a new species occupying the crowns of large Sequoia sempervirens trees in California, USA. The new species is supported by morphology, anatomy, secondary chemistry and DNA sequence data. While similar in external appearance to X. friesii, it is distinguished by forming smaller, partly coralloid squamules, by the occurrence of soralia and, in some specimens, by the presence of thamnolic acid in addition to friesiic acid in the thallus. Molecular phylogenetic results are based on nuclear (ITS and LSU) as well as mitochondrial (SSU) ribosomal DNA sequence alignments. Phylogenetic hypotheses obtained using Bayesian Inference, Maximum Likelihood and Maximum Parsimony all support X. canopeorum as a distinct evolutionary lineage belonging to the X. caradocensis–X. friesii clade.

California, epiphytic, Hypocenomyce , integrative taxonomy, morphology, multiple DNA sequence alignment, phylogeny, redwood forest, TLC

The squamulose lichen genus Xylopsora Bendiksby & Timdal consists of two species, X. caradocensis (Nyl.) Bendiksby & Timdal and X. friesii (Ach.) Bendiksby & Timdal. The two species were formerly placed in Hypocenomyce M. Choisy and referred to as the H. friesii group (Timdal 1984, 2001) until Bendiksby and Timdal (2013) showed that Hypocenomyce was highly polyphyletic. Xylopsora is the phylogenetic sister of the clade consisting of the two foliose genera Lasallia Mérat and Umbilicaria Hoffm. Those three genera make up the sister clade of the genus Fulgidea Bendiksby & Timdal, another Hypocenomyce segregate. The four genera together constitute the Umbilicariaceae (Bendiksby and Timdal 2013). Fulgidea consists of two species, F. oligospora (Timdal) Bendiksby & Timdal and F. sierrae (Timdal) Bendiksby & Timdal. Fulgidea and Xylopsora are morphologically, anatomically and ecologically very similar and differ mainly in secondary chemistry; alectorialic acid and thamnolic acid occur in the former, friesiic acid (= “friesii unknown”) in the latter (Timdal 2001, 2002). All species of Fulgidea and Xylopsora grow on bark and wood and, with the exception of X. caradocensis, show preference for burnt stumps and trunks of conifers.

Coast redwood (Sequoia sempervirens) forests are an important component of California’s ecosystems. Spanning more than six degrees of latitude along the Pacific coast (Van Pelt 2001) and containing individual trees that can live for more than 2000 years (Rocky Mountain Tree Ring Research 2017), these forests provide important habitats for many terrestrial species (Sawyer et al. 1999). However, biodiversity occupying the redwood forest canopies remains relatively under-explored because access into the tree crowns, which often grow to over 100 m in height, is challenging. The epiphytic lichen flora in old-growth redwood forests appears to be particularly species rich; an epiphyte survey of just nine large redwood trees yielded 137 lichen species including a new species of Calicium (Williams and Sillett 2007, Williams and Tibell 2008).

Recent epiphyte surveys in the crowns of additional large coast redwood trees in the southern part of the geographic range (Reese Næsborg 2017) revealed a previously undescribed species of Xylopsora. Here the authors have provided detailed morphologic, anatomic, chemical and molecular description of this new species, as well as characterising the habitat and substrates it occupies.

Establishing a multiple DNA sequence alignment (MSA) of non-coding loci, which often have unequal lengths due to indels, can be both time-consuming and highly subjective with regard to structural correctness. There has been great activity in recent years in the development of multiple sequence alignment tools (reviewed by Kamena and Notredame 2009). Moreover, there is no single recommendation as to what phylogenetic algorithm to use to transform the MSA into a reliable phylogenetic hypothesis. The current, moderately sized dataset has been used to test whether a less time-consuming and more objective approach, SATé-II (Liu et al. 2012), provides similar and meaningful results. Both manual and automatic approaches have been used to establish a concatenated MSA of three loci, of which one is non-coding and highly variable (nrITS). Different methods representing three classes of phylogenetic inference (Bayesian, likelihood and parsimony) have also been used.

Four nrITS and three mtSSU sequences were generated (GenBank accession numbers MG309307–MG309313; Table 1). Preliminary parsimony jack-knife analyses of the individual three loci, regardless of the alignment approach, produced congruent gene-trees that were resolved to various extents (not shown). In subsequent analyses, the three loci were analysed in concert. The MSAmanual alignment (i.e. three loci, manually aligned) was 10 characters shorter than MSAsate (i.e. three loci, automatically aligned) and had 12 fewer parsimony informative characters (PIC; Table 2). Both alignments are provided as Suppl. material 1, 2 (MSAmanual.nex, MSAsate.nex). The MSAmanual dataset produced 1220 most parsimonious trees (MPTs) of length 1479, whereas MSAsate produced 10 MPTs of length 1485. Homoplasy measures (Farris 1989) differ negligibly between the two (RC: 46.6 vs 46).The likelihood scores from the RAxML analyses in SATé-II were very similar (Table 2). The ASDSF fell below 0.01 faster in the BI analysis of MSAsate (at generation 820) than in the BI analysis of MSAmanual (around generation 1300). All significantly supported clades were congruent amongst the BI, ML and MP analyses, regardless of the dataset analysed (MSAmanual vs MSAsate). Only results from analyses of the MSAsate dataset are shown (Fig. 1). The authors regarded clade support of at least 60% jack-knife (JK) and at least 0.9 posterior probability (PP) as significant.

The four accessions of the tentatively new species group with significant support and showed themselves as sister to a clade consisting of three accessions of Xylopsora friesii (2, 3 and 4; Fig. 1). Four characters varied amongst the four accessions of the tentatively new species (three in the nrITS and one in mtSSU), none of which were parsimoniously informative within the group. Xylopsora cf friesii 1 differed from X. friesii 2, 3 and 4 in eight characters in the nrLSU, at least four in the mtSSU and at least 16 in the nrITS. The MP analyses supported Elixia as monophyletic (JK MSAmanual = 94 %; JK MSAsate = 87 %) with Meridianelia as sister (JK MSAmanual = 95 %; JK MSAsate = 93 %). In the ML and BI analyses, Elixia monophyly was not supported, as Meridianelia grouped with accessions of E. flexella and Elixia sp. and excluded Elixia cretica. This topology was not significantly supported by any analyses. The sister-relation between the Elixiaceae and the Ophioparmaceae was significantly supported only by Bayesian PP (Fig. 1).

Figure 1. 

Hypothesis of the phylogenetic relationships and placement of the potentially undescribed species of Xylopsora based on DNA sequence data. The depicted topology is based on an automated alignment (MSAsate) of two nuclear (ITS and LSU) and one mitochondrial (SSU) ribosomal loci and is the “best tree” from a RAxML analysis using SATé-II. Clade support over certain values from Bayesian inference (posterior probability; PP) and parsimony jackknifing (JK) analyses are superimposed: PP >0.9 and JK>60% (PP/JK). Clades receiving maximum PP support (1.0) and at least 90% JK support are indicated with a black dot. Multiple accessions of the same taxon are numbered according to Table 1 (also corresponding to the numbering in Bendiksby & Timdal 2013). Family and genus circumscriptions are indicated. Abbreviation: Fusc. = Fuscideaceae. One accession in the tree (Meridianelia maccarthyana) appeared on a very long branch that is manually shortened (arrow tipped) to reduce the size of a broad figure.

Forest canopies in general are relatively understudied because accessing the tree crowns requires technical expertise and equipment (Lowman et al. 2012). Therefore, the potential for encountering new species is relatively high compared to more easily accessible forest floor environments. The new species presented here, Xylopsora canopeorum, has so far only been registered from the crowns of large coast redwood trees, but other similar habitats, like the fibrous bark of other large members of Cupressaceae, should be explored for the species. The collected Xylopsora canopeorum specimens occurred on stable bark surfaces of old, large redwood trees together with several species previously classified in the genus Hypocenomyce (e.g. Carbonicola anthracophila, Fulgidea oligospora, F. sierrae, and H. scalaris), which was recently shown to be highly polyphyletic (Bendiksby and Timdal 2013). Together, these species covered a substantial portion of the trunk surface.

We would like to thank Cameron Williams, Chuck Vanderpool, Wendy Baxter, Allison Kidder and Samuel Hargrove for field assistance. Cameron Williams provided constructive comments on the manuscript. Emily Bertram, Supervising State Park Peace Officer in Big Basin Redwoods State Park and Brendan O’Neil, Senior Environmental Scientist in Armstrong Redwoods State Natural Reserve facilitated the field work. Funding was provided by Save the Redwoods League. Sonja D. Kistenich assisted at the DNA lab at O. We also find it relevant to acknowledge the support of the Norwegian Biodiversity Information Centre (NBIC; ArtsDatabanken) that largely funded the establishment of the DNA sequence framework (project Nos. 70184216, 70184227).