Research Article |
Corresponding author: Emilia Ossowska ( emilia.ossowska@ug.edu.pl ) Academic editor: Thorsten Lumbsch
© 2019 Emilia Ossowska, Beata Guzow-Krzemińska, Marta Kolanowska, Katarzyna Szczepańska, Martin Kukwa.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Ossowska E, Guzow-Krzemińska B, Kolanowska M, Szczepańska K, Kukwa M (2019) Morphology and secondary chemistry in species recognition of Parmelia omphalodes group – evidence from molecular data with notes on the ecological niche modelling and genetic variability of photobionts. MycoKeys 61: 39-74. https://doi.org/10.3897/mycokeys.61.38175
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To evaluate the importance of morphological and chemical characters used in the recognition of species within the Parmelia omphalodes group, we performed phylogenetic, morphological and chemical analyses of 335 specimens, of which 34 were used for molecular analyses. Phylogenetic analyses, based on ITS rDNA sequences, show that P. pinnatifida is distinct from P. omphalodes and the most important difference between those species is the development of pseudocyphellae. In P. pinnatifida, they are mostly marginal and form white rims along lobes margins, but laminal pseudocyphellae can develop in older parts of thalli and are predominantly connected with marginal pseudocyphellae. In contrast, in P. omphalodes laminal pseudocyphellae are common and are predominantly not connected to marginal pseudocyphellae. Chemical composition of secondary lichen metabolites in both analysed species is identical and therefore this feature is not diagnostic in species recognition. Few samples of P. discordans, species morphologically similar to P. omphalodes and P. pinnatifida, were also included in the analyses and they are nested within the clade of P. omphalodes, despite the different chemistry (protocetraric acid present versus salazinic acid in P. omphalodes). All taxa of the P. omphalodes group occupy similar niches, but their potential distributions are wider than those currently known. The absence of specimens in some localities may be limited by the photobiont availability. Parmelia omphalodes and P. pinnatifida are moderately selective in photobiont choice as they form associations with at least two or three lineages of Trebouxia clade S. Parmelia pinnatifida, as well as P. discordans are associated with Trebouxia OTU S02 which seems to have a broad ecological amplitude. Other lineages of Trebouxia seem to be rarer, especially Trebouxia sp. OTU S04, which is sometimes present in P. pinnatifida. This study indicates the importance of extensive research including morphology, chemistry and analysis of molecular markers of both bionts in taxonomical studies of lichens.
Ascomycota, Parmeliaceae, parmelioid lichens, ITS rDNA, secondary metabolites, morphology, photobiont, ecological niche modelling
The genus Parmelia Ach. (Parmeliaceae, Ascomycota) currently comprises ca. 40 species (
The P. omphalodes group includes three taxa, often treated at the species level, i.e. P. discordans Nyl., P. omphalodes (L.) Ach. and P. pinnatifida Kurok. (
The second issue is related to the differences between the species.
The species of the Parmelia omphalodes group are rare in most parts of their distributional ranges. Parmelia discordans is reported from Europe only (
According to literature, all Parmelia species form associations with green algae of the genus Trebouxia de Puymaly (
During our study of P. omphalodes and P. pinnatifida specimens, important differences between published data and the results of our own studies were observed. For example, lobaric acid was identified in the specimens with marginal pseudocyphellae (thus morphologically similar to P. pinnatifida) or both lobaric acid and fatty acids were absent in specimens with marginal and laminal pseudocyphellae (thus morphologically similar to P. omphalodes). The differences between our results and literature data prompted more detailed morphological, chemical and phylogenetic studies on those two species, which are also relatively common and thus easy to be sampled for molecular analyses. We also included a few samples of P. discordans to better understand the differences amongst all three species of P. omphalodes group, especially in the case of photobiont associations. In the study, we used the nuclear ribosomal internal transcribed spacer region (ITS), which is considered as a universal barcode marker for fungi in many taxonomic groups (e.g.
The main goals of this paper are to study the phylogenetic relationships between P. discordans, P. omphalodes and P. pinnatifida, to determine, based on molecular evidence, the diagnostic characters separating P. omphalodes and P. pinnatifida and to study the photobionts genetic variation in all three species. As not much is known about their ecology, the evaluation of the ‘ecological niche similarity’ is also presented.
In total, 335 herbarium specimens deposited in B, H, HBG, LD, S, UGDA and UPS were used for morphological, chemical and ecological niche modelling (ENM) study: 61 of P. discordans, 113 of P. pinnatifida and 161 of P. omphalodes. A total of 34 specimens were selected for molecular study using the nuclear internal transcribed spacer region (ITS rDNA). Thirty four ITS rDNA sequences of the mycobionts and 17 ITS rDNA sequences of their photobionts were newly generated (Table
Specimens used in this study with the locality, voucher information, references and GenBank accession numbers. Sequences generated during this study are in bold.
Species/OTU | Voucher/ References | Fungal ITSrDNA | Algal ITSrDNA |
---|---|---|---|
Parmelia discordans | Sweden, S-F284965, Odelvik 15-293 | MN412798 | MN412816 |
Sweden, S-F252494, Odelvik 13-147 et al. | MN412800 | MN412815 | |
Sweden, UGDA L-23627, Kukwa 12278 | MN412799 | – | |
UK, MAF-Lich 10232, ( |
AY583212 | – | |
Parmelia ernstiae | Germany, HBG 4619 ( |
AF410833 | – |
Latvia, UGDA L-19917 ( |
KU845673 | – | |
Parmelia imbricaria | Canada, TG 08-108 ( |
KT625503 | – |
Parmelia mayi | USA, MAF 15765 ( |
JN609439 | – |
USA, MAF 15766 ( |
JN609438 | – | |
USA, MAF 15767 ( |
JN609437 | – | |
Parmelia omphalodes | Sweden, S-F236118, Odelvik 12163 | MN412792 | MN412806 |
Sweden, S-F300480, Odelvik 16-490 | MN412794 | MN412805 | |
Sweden, S-F252845, Odelvik 13-113 | MN412793 | MN412808 | |
UK, 2240 ( |
EF611295 | – | |
Finland ( |
AY251440 | – | |
Spain, MAF 7062 ( |
AY036998 | – | |
Spain, MAF 7044, ( |
AY036999 | – | |
Sweden, S-F238139, Odelvik 12238 | MN412796 | MN412803 | |
Sweden, UGDA L- 23632, Kukwa 12283 | MN412795 | MN412817 | |
Parmelia pinnatifida | Norway, S-F254099, Odelvik 13-439 | MN412790 | MN412804 |
Sweden, S-F299936, Odelvik 16-276 | MN412791 | – | |
Sweden, S-F252763, Odelvik 13-225 et al. | MN412797 | MN412807 | |
Sweden, S-F285120, Odelvik 15-294 et al. | MN412789 | MN412802 | |
Poland, UGDA L-24300, Ossowska 118 et al. | MN412774 | – | |
Poland, UGDA L-24301, Ossowska 119 et al. | MN412775 | MN412813 | |
Poland, UGDA L-24302, Ossowska 120 et al. | MN412776 | – | |
Poland, UGDA L-24304, Ossowska 123 et al. | MN412777 | – | |
Poland, UGDA L-24305, Ossowska 124 et al. | MN412778 | MN412814 | |
Poland, UGDA L-24306, Ossowska 127 et al. | MN412779 | – | |
Poland, UGDA L-24307, Ossowska 132 et al. | MN412780 | – | |
Poland, UGDA L-24308, Ossowska 133 et al. | MN412781 | – | |
Poland, UGDA L-24310, Ossowska 137 et al. | MN412783 | – | |
Poland, UGDA L-24311, Ossowska 138 et al. | MN412782 | – | |
Poland, UGDA L-24318, Ossowska 150 et al. | MN412785 | MN412812 | |
Poland, UGDA L-24319, Ossowska 152 et al. | MN412784 | MN412818 | |
Poland, UGDA L-24313, Ossowska 143 et al. | MN412786 | – | |
Poland, UGDA L-24312, Ossowska 139 et al. | MN412787 | MN412811 | |
Poland, UGDA L-24316, Ossowska 147 et al. | MN412788 | – | |
Poland, UGDA L-24294, Szczepańska s.n. | MN412772 | MN412810 | |
Poland, UGDA L-24293, Szczepańska 1040 | MN412770 | MN412809 | |
Poland, UGDA L-24296, Szczepańska 1049 | MN412767 | – | |
Poland, UGDA L-24297, Szczepańska 1052 | MN412768 | – | |
Poland, UGDA L-24298, Szczepańska 1080 | MN412769 | – | |
Poland, UGDA L-24295, Szczepańska 1126 | MN412773 | – | |
Poland, UGDA L-24299, Szczepańska 1135 | MN412771 | – | |
Austria ( |
EF611300 | – | |
Russia, MAF 7272 ( |
AY036988 | – | |
Russia, MAF 7274 ( |
AY036987 | – | |
Parmelia saxatilis | Czech Republic, UGDA L-21245 ( |
KU845667 | – |
Sweden, S-F300671, Odelvik 16-669 & Hedenäs | MN412801 | – | |
Sweden, MAF 6882 ( |
AF350028 | – | |
Parmelia serrana | Poland, UGDA L-21210 ( |
KU845669 | – |
Spain, MAF 9756 ( |
AY295109 | – | |
Parmelia skultii | Canada, LD 795 ( |
AY251456 | – |
Greenland, 311C ( |
FJ425881 | – | |
Parmelia submontana | Poland, UGDA L-21213 ( |
KU845664 | – |
Morocco, MAF 15440 ( |
JN609434 | – | |
Parmelia sulcata | Ireland, MAF 15421 ( |
JN118597 | – |
OTU I01 | USA, I01_RH_shus_usa_UT_saxi_544 ( |
– | KR913803 |
OTU I02 | USA, I02_ME_subau_usa_MI_cort_4176 ( |
– | KR913865 |
OTU I03 | Estonia, I03_MH_exata_estonia_unk_cort_4110 ( |
– | KR913991 |
OTU I04 | Russia, I04_RH_chryC_russia_Orenb_saxi_6890 ( |
– | KR914011 |
OTU I05 | USA, I05_PUN_rud_usa_OH_cort_3157 ( |
– | KR914027 |
OTU I06 | Canada, I06_MH_infum_canada_BC_saxi_4834 ( |
– | KR914029 |
OTU I07 | USA, I07_ME_elber_usa_MN_cort_5773 ( |
– | KR914035 |
OTU I08 | China, I08_MH_subexata_china_richuan_cort_3649 ( |
– | KR914042 |
OTU I09 | USA, I09_MH_halei_usa_NC_cort_4008 ( |
– | KR914044 |
OTU I10 | Argentina, I10_MH_ushua_argentina_unk_saxi_6045 ( |
– | KR914047 |
OTU I11 | Russia, I11_MH_oliva_russia_Prim_cort_6012 ( |
– | KR914050 |
OTU I12 | Russia, I12_MH_oliva_russia_Prim_cort_5998 ( |
– | KR914053 |
OTU I13 | USA, I13_PUN_cas_usa_OH_cort_3161 ( |
– | KR914054 |
OTU I14 | Russia, I14_MH_oliva_russia_Prim_cort_5973 ( |
– | KR914055 |
OTU I15 | Kenya, I15_PUN_rud_kenya_unk_cort_1195 ( |
– | KR914056 |
OTU S01 | Canada, S01_LE_lupina_canada_BC_cort_FJ170511 ( |
– | FJ170511 |
OTU S02 | UK, S02_CE_acul_ant_shetland_terr_GQ375315 ( |
– | GQ375315 |
OTU S03 | Canada, S03_LE_vulpina_canada_BC_cort_FJ170752 ( |
– | FJ170752 |
OTU S04 | Canada, S04_MH_exula_canada_BC_cort_5194 ( |
– | KR914114 |
OTU S05 | USA, S05_LE_vulpina_usa_CA_cort_FJ170727 ( |
– | FJ170727 |
OTU S06 | USA, S06_MH_eltula_usa_CO_cort_4212 ( |
– | KR914169 |
OTU S07 | USA, S07_MH_eltula_usa_WA_cort_4343 ( |
– | KR914185 |
OTU S08 | Spain, S08_CE_acul_spain_unk_terr_GQ375345 ( |
– | GQ375345 |
OTU S09 | Turkey, S09_CE_acul_turkey_unk_terr_GQ375351 ( |
– | GQ375351 |
OTU S10 | S10_TRE_simplex_SAG101_80_cult_FJ626735 ( |
– | FJ626735 |
OTU S11 | S11_TRE_australis_SAG2250_cult_FJ626726 ( |
– | FJ626726 |
OTU S12 | USA, S12_CE_acul_usa_AK_terr_GU124701 ( |
– | GU124701 |
OTU S13 | S13_TRE_brindabellae_SAG2206_FJ626727 ( |
– | FJ626727 |
OTU G01 | Canaries, G01_PMT_pse_CANAR_gome_cort_3730 ( |
– | KR913271 |
OTU G02 | Canaries, G02_PMT_per_CANAR_gome_cort_3751 ( |
– | KR913285 |
OTU G03 | G03_TRE_usneae_UTEX2235_cult_AJ249573 ( |
– | AJ249573 |
OTU G04 | Canaries, G04_PMT_per_CANAR_gome_cort_3746 ( |
– | KR913286 |
OTU G05 | G05_TRE_galapagensis_UTEX2230_AJ249567 ( |
– | AJ249567 |
OTU A01 | USA, A01_LEC_garov_usa_ID_saxi_078 ( |
– | KR912351 |
OTU A02 | USA, A02_LEC_garov_usa_ID_saxi_108 ( |
– | KR912568 |
OTU A03 | Sweden, A03_ME_fulig_swe_Skane_cort_3935 ( |
– | KR912760 |
OTU A04 | USA, A04_XA_chE2_usa_ID_terr_201 ( |
– | KR912832 |
OTU A05 | Mexico, A05_ORO_bicolor_mexico_OAX_cort_4043 ( |
– | KR912913 |
OTU A06 | USA, A06_XA_coE3_usa_CO_saxi_6618 ( |
– | KR912989 |
OTU A07 | USA, A07_XA_chE2_usa_UT_terr_008 ( |
– | KR913034 |
OTU A08 | USA, A08_RH_mela_usa_UT_saxi_614 ( |
– | KR913115 |
OTU A09 | USA, A09_XA_coE3_usa_UT_saxi_064 ( |
– | KR913162 |
OTU A10 | Canada, A10_XA_cuF1_canada_BC_saxi_1007 ( |
– | KR913184 |
OTU A11 | USA, A11_XA_idBX_usa_WY_terr_787 ( |
– | KR913199 |
OTU A12 | USA, A12_XA_chE3_usa_UT_terr_126 ( |
– | KR913203 |
OTU A13 | UK, A13_LEC_disp_uk_unk_saxi_6407 ( |
– | KR913212 |
OTU A14 | USA, A14_XA_maricopF2_usa_A2_saxi_6699 ( |
– | KR913215 |
OTU A15 | A15_TRE_gigantea_UTEX2231_cult_AF242468 ( |
– | AF242468 |
OTU A16 | Canada, A16_XA_caB1_canada_BC_terr_901 ( |
– | KR913224 |
OTU A17 | Peru, A17_ORO_unk_peru_unk_cort_1602 ( |
– | KR913235 |
OTU A18 | USA, A18_LEC_garov_usa_UT_saxi_140 ( |
– | KR913237 |
OTU A19 | Canaries, A19_PMT_per_CANAR_gome_cort_3742 ( |
– | KR913241 |
OTU A20 | USA, A20_XA_meF2_usa_A2_saxi_147 ( |
– | KR913248 |
OTU A21 | USA, A21_XA_caB3_usa_ID_terr_334 ( |
– | KR913250 |
OTU A22 | USA, A22_XA_chE2_usa_UT_terr_007 ( |
– | KR913255 |
OTU A23 | A23_TRE_showmanii_UTEX2234_cult_AF242470 ( |
– | AF242470 |
OTU A24 | USA, A24_ME_calif_usa_CA_cort_4088 ( |
– | KR913251 |
OTU A25 | USA, A25_XA_mariF2_usa_A2_saxi_6698 ( |
– | KR913259 |
OTU A26 | USA, A26_XA_coE3_usa_UT_saxi_073 ( |
– | KR913261 |
OTU A27 | USA, A27_XA_chE3_usa_WY_terr_110 ( |
– | KR913264 |
OTU A28 | Mexico, A28_XA_diA1_mex_PU_saxi_098 ( |
– | KR913265 |
OTU A29 | Japan, A29_MO_predis_japan_Shinano_saxi_8597 ( |
– | KR913266 |
OTU A30 | USA, A30_XA_cuE2_usa_UT_saxi_036 ( |
– | KR913267 |
OTU A31 | USA, A31_XA_coE1_usa_UT_saxi_030 ( |
– | KR913268 |
OTU A32 | USA, A32_XA_cuE1_usa_UT_saxi_075 ( |
– | KR913269 |
OTU A33 | A33_TRE_decolorans_UTEXB781_cult_FJ626728 ( |
– | FJ626728 |
OTU A34 | USA, A34_XA_mariF2_usa_AZ_saxi_6702 ( |
– | KR913270 |
The upper surfaces of all specimens were examined to determine the type of pseudocyphellae orientation such as: only marginal, marginal with few laminal in older parts of thalli and marginal and laminal in young and older parts of thalli. Pseudocyphellae were analysed on the whole thalli surfaces. Moreover, the length (distance between points of lobe branching) and width (distance between two adjacent lobe edges at the point of their branching) of lobes were also measured. Based on morphology and chemistry (see below), the studied specimens were divided into groups, which are characterised in Table
Diagnostic morphological and chemical features in species from Parmelia omphalodes group analysed in this study with their classification after molecular research (ATR – atranorin, SAL – salazinic acid with consalazinic acid, LOB – lobaric acid, PRC – protocetraric acid, LICH – lichesternic acid, PRL – protolichesterinic acid).
Chemistry | Orientation of pseudocyphellae | Lenght (L) and width (W) of lobes (mm) | Voucher of specimens used in molecular research | Classification after molecular research |
ATR, SAL, LOB | marginal | L 1.5–2; W 1 | S-F299936 | Parmelia pinnatifida |
S-F254099 | ||||
ATR, SAL, LOB | marginal, laminal in older lobes | L 2; W 2 | UGDA L-24310 | Parmelia pinnatifida |
S-F252763 | ||||
ATR, SAL, LOB, LICH, PRL | marginal | L 1–2; W 0.5–1.5 | UGDA L-24295 | Parmelia pinnatifida |
UGDA L-24311 | ||||
UGDA L-24319 | ||||
UGDA L-24294 | ||||
UGDA L-24296 | ||||
UGDA L-24298 | ||||
UGDA L-24305 | ||||
UGDA L-24306 | ||||
ATR, SAL, LOB, LICH, PRL | marginal, laminal in older lobes | L 1.5–2; W 1.5–2 | UGDA L-24313 | Parmelia pinnatifida |
UGDA L-24308 | ||||
UGDA L-24293 | ||||
UGDA L-24297 | ||||
ATR, SAL, LOB, PRL | marginal | L 0.5–2; W 0.5–1 | UGDA L-24299 | Parmelia pinnatifida |
UGDA L-24300 | ||||
UGDA L-24307 | ||||
UGDA L-24318 | ||||
ATR, SAL | marginal | L 1; W 1 | UGDA L-24304 | Parmelia pinnatifida |
MAF 7274 | ||||
ATR, SAL | marginal, laminal in older lobes | L 1.5 ,W 1 | UGDA L-24312 | Parmelia pinnatifida |
ATR, SAL, LICH, PRL | marginal | L 2; W 1 | UGDA L-24301 | Parmelia pinnatifida |
ATR, SAL, PRL | marginal | L 1.5–2; W 1.5–1 | UGDA L-24302 | Parmelia pinnatifida |
S-F285120 | ||||
ATR, SAL, PRL | marginal, laminal in older lobes | L 1.5; W 1 | UGDA L-24316 | Parmelia pinnatifida |
ATR, PRC, LOB | marginal | L 3; W 1–2 | S-F284965 | Parmelia discordans |
S-F252494 | ||||
MAF 10232 | ||||
ATR, PRC | marginal and laminal on young thalli | L 3; W 2 | UGDA L-23627 | Parmelia discordans |
ATR, SAL, LOB | marginal, laminal | L 3–4; W 2–3 | S-F300480 | Parmelia omphalodes |
S-F252845 | ||||
S-F238139 | ||||
S-F236118 | ||||
UGDA L-23632 | ||||
MAF 7044 | ||||
ATR, SAL | marginal, laminal | L 2; W 1.5 | MAF 7062 | Parmelia omphalodes |
Secondary lichen compounds were identified using thin-layer chromatography (TLC) in solvents A and C (
Total genomic DNA was extracted using the Sherlock AX Kit (A&A Biotechnology, Poland) in accordance with the manufacturer’s protocol, with slight modifications described by
Fungal ITS rDNA was amplified using the primers ITS1F and ITS4A (
The amplifications were performed in an Eppendorf thermocycler and carried out using the following programme: for fungal ITS rDNA marker: initial denaturation at 94 °C for 3 min and 33 cycles of: 94 °C for 30 sec; annealing at 52 °C for 45 sec; extension at 72 °C for 1 min and final extension at 72 °C for 10 min. For green-algal ITS: initial denaturation at 94 °C for 3 min and 35 cycles of: 94 °C for 45 sec; annealing at 55 °C for 45 sec; extension at 72 °C for 90 sec and final extension at 72 °C for 7 min.
The PCR products were purified using Wizard SV Gel and PCR Clean Up System (Promega, US), according to the manufacturer’s instruction. The cleaned DNA was sequenced using Macrogen sequencing service (http://www.macrogen.com).
The newly generated mycobiont sequences, together with selected representatives of Parmelia spp., were automatically aligned in Seaview (
The newly generated photobiont sequences, together with representative Trebouxia OTUs, downloaded from Dryad database (Dryad Digital Repository) (
The final alignment of photobionts consisted of 84 ITS rDNA sequences and 580 characters. The names of operational taxonomic units (OTU) for Trebouxia ITS rDNA sequences were given according to
The GTR+I+G best-fit evolutionary model was selected for the mycobiont dataset, based on Akaike Information Criterion (AIC) (
Bayesian analysis was carried out using the Metropolis-coupled Markov chain Monte Carlo (MCMCMC) method by using the Markov chain Monte Carlo (MCMC) method, in MrBayes v. 3.2.6 (
A Maximum Likelihood (ML) analysis was performed using RAxML-HPC2 v.8.2.10 (
Phylogenetic trees were visualised using FigTree v. 1.4.2 (
Phylogenetic relationships of Parmelia discordans, P. omphalodes and P. pinnatifida, based on Bayesian analysis of the ITS rDNA dataset. Posterior probabilities and maximum likelihood bootstrap values are shown near the internal branches. Newly generated sequences are described with herbarium numbers following the species names. GenBank Accession numbers of sequences downloaded from GenBank follow the species names. Clades with Parmelia discordans, P. omphalodes and P. pinnatifida are highlighted.
Phylogenetic placement of Trebouxia photobionts from selected Parmelia spp., based on Bayesian analysis of the ITS rDNA dataset. Posterior probabilities and maximum likelihood bootstrap values are shown near the internal branches. Newly generated sequences are in bold, with collecting numbers preceding the species names. Representative Trebouxia OTUs, as described in
Sequences of ITS rDNA from specimens belonging to P. discordans and P. omphalodes were aligned using Seaview software (
Haplotype network showing relationships between ITS rDNA sequences from Parmelia discordans and P. omphalodes. The names of species are followed with herbarium numbers of specimens or GenBank Accession Numbers. Mutational changes are presented as numbers in brackets near lines between haplotypes.
To evaluate the similarity of niches occupied by all studied taxa, ecological niche modelling (ENM) was applied.
The database of localities of P. discordans, P. omphalodes and P. pinnatifida was compiled, based on information provided on labels of herbarium specimens. The geographic coordinates provided on the herbarium sheet labels were verified. If there were no information about the latitude and longitude on the herbarium sheet label, we followed the description of the collection site and assigned coordinates as precisely as possible to this location. Google Earth (Google Inc.) was used to validate all gathered information. In total, 61 records of P. discordans, 161 of P. omphalodes and 113 of P. pinnatifida were used to perform ENM analysis (Figure
The maximum entropy method, as implemented in Maxent version 3.3.2 software, was used to create models of the suitable niche distribution (
Twelve bioclimatic variables in 2.5 minutes developed by
bio1 | annual mean temperature |
bio2 | mean diurnal range (mean of monthly (max temp - min temp)) |
bio3 | isothermality (mean diurnal range / temperature annual range * 100) |
bio4 | temperature seasonality (standard deviation *100) |
bio5 | max temperature of the warmest month |
bio8 | mean temperature of the wettest quarter |
bio12 | annual precipitation |
bio13 | precipitation of the wettest month |
bio14 | precipitation of the driest month |
bio15 | precipitation seasonality (coefficient of variation) |
bio18 | precipitation of the warmest quarter |
bio19 | precipitation of coldest quarter |
The differences amongst the niches occupied by the populations of three studied lichens were evaluated using the niche identity indices: Schoener’s D (D) and I statistic (I) as available in ENMTools v1.3 (
Principal components analysis (PCA) was performed to explain the general variation pattern amongst the studied species, based on 12 bioclimatic factors used in ENM analysis. Statistical computations were performed with the programme PAST v. 3.0 (
Trees of similar topologies were generated using the maximum likelihood method (RaxML; best tree likelihood LnL = −1512.540166) and the Bayesian approach (BA; harmonic mean was −1667.09). The Bayesian tree is presented in Figure
Within the larger clade of P. omphalodes, four sequences obtained from specimens containing protocetraric acid and determined as P. discordans are nested. Three of those specimens form a highly supported lineage (1 PP and 93 BS), while the fourth sample of P. discordans is placed outside this subclade (Figure
So far, the taxonomy of P. omphalodes group was unclear.
The distinguishing character between P. omphalodes and P. pinnatifida is the development of pseudocyphellae; however, the determination of the type and orientation of pseudocyphellae requires checking of the entire thallus surface, not only marginal or central parts of the thalli. We concluded that P. pinnatifida has mostly marginal pseudocyphellae forming white rims around lobes margins (Figure
A Parmelia discordans, with marginal and laminal pseudocyphellae, laminal pseudocyphellae mostly not connected with marginal ones (S F-252494) B P. omphalodes, with marginal and laminal pseudocyphellae, laminal pseudocyphellae mostly not connected with marginal ones (S F-252845) C P. pinnatifida, with marginal pseudocyphellae (UGDA L-24298) D P. pinnatifida, with marginal and laminal pseudocyphellae, laminal pseudocyphellae starting predominantly from pseudocyphellae formed at the edge of lobes (S F-239397). Scale bars: 200 μm (A, B, D), 150 μm (C).
The presence of lobaric and fatty acids cannot be treated as diagnostic for the separation of P. omphalodes and P. pinnatifida, as it does not correspond with molecular data. Until now, P. pinnatifida was characterised as a species lacking lobaric acid (
Morphological and chemical characteristics of all taxa of the group are summarised in Table
Historical and present overview of species delimitations within the Parmelia omphalodes group with their morphological and chemical characteristics (ATR – atranorin, SAL – salazinic acid with consalazinic acid, LOB – lobaric acid, PRC – protocetraric acid, PRL – protolichesterinic acid, FAT – fatty acids; + present in all specimens; ± sometimes present).
Taxa | Morphology | Chemistry | |
|
P. discordans | pseudocyphellae marginal and laminal; lobules absent; lobes 1–2.5 mm wide | ATR (+), PRC (+), LOB (+), FAT (±) |
P. omphalodes | pseudocyphellae marginal and laminal; lobules present | ATR (+), SAL (+), LOB (+) | |
P. pinnatifida | pseudocyphellae marginal; narrow lobules present; lobes repeatedly branched | ATR (+), SAL (+), FAT (+) | |
|
P. omphalodes subsp. discordans | pseudocyphellae sparse and marginal in young lobes; lobes diameter 0.13–2.8 mm | ATR (+), PRC (+), LOB (+), PRL (+) |
P. omphalodes subsp. omphalodes | pseudocyphellae marginal and laminal; lobes up to 3.5 mm diameter | ATR (+), SAL (+), LOB (+), PRL (±) | |
P. omphalodes subsp. pinnatifida | pseudocyphellae marginal, in old lobes laminal; lobes narrow, 0.13–2.9 mm diameter | ATR (+), SAL (+), PRL (±) | |
|
P. discordans | pseudocyphellae marginal, few also laminal; lobes 1–3 mm wide | ATR (+), PRC (+), LOB (+), unidentified FAT (±) |
P. omphalodes | pseudocyphellae mostly marginal; lobes wide 1–4 mm | ATR (+), SAL (+), LOB (±), PRL (+)* | |
|
P. discordans | pseudocyphellae linear; lobes overlapping, 1–3 mm wide | PRC (+), LOB (+) |
P. omphalodes | lobes 4 mm wide | ATR (+), SAL (+), LOB (+), PRC (±) | |
P. pinnatifida | pseudocyphellae restricted to the margins; lobes narrow, repeatedly branched and overlapping | ATR (+), SAL (+), PRL (+) | |
|
P. discordans | pseudocyphellae indistinct; lobes narrow | ATR (+), PRC (+), LOB (+) |
P. omphalodes | – | ATR (+), SAL (+), LOB (+), PRL (+), PRC (±) | |
P. pinnatifida | pseudocyphellae marginal; lobes narrow | ATR (+), SAL (+), PRL (+), PRC (±) | |
This study | P. discordans | pseudocyphellae marginal and laminal, laminal pseudocyphellae at least partly not starting from the lobe margins; lobes narrow and sublinear, about 1–3 mm wide and 1–3 mm length | ATR (+), PRC (+), LOB (±), FAT (±) |
P. omphalodes | pseudocyphellae marginal and laminal, laminal pseudocyphellae mostly not starting from the lobe margins; lobes broad and sublinear, about 2–3 mm wide and 3–4 mm length | ATR (+), SAL (+), LOB (±), FAT (±) | |
P. pinnatifida | pseudocyphellae marginal, in older parts of thalli with few laminal connected to the lobes margins; lobes narrow, sublinear, about 1–2 mm wide and 0.5–2 mm length | ATR (+), SAL (+), LOB (±), FAT (±) |
Trees of similar topologies were generated using maximum likelihood (RaxML; best tree likelihood LnL = -7013.073328) and Bayesian analysis (BA; harmonic mean was -6996.31). The Bayesian tree is presented in Figure
Association network between lichen mycobionts of P. omphalodes group (i.e. Parmelia discordans, P. omphalodes and P. pinnatifida) and photobiont OTUs. The line width is proportional to the number of specimens forming the association with the particular OTU. SUn1 and SUn2 represent unnamed lineages of Trebouxia belonging to clade S.
According to
Lichens that reproduce sexually via independent dispersal of fungal spores, undergo a process of re-lichenisation. This means that the germinating spore of the mycobiont can easily exchange its autotrophic partner, in contrast to asexually reproducing lichens distributing both partners together, which allows continuation of the symbiosis without the need to re-associate with another biont (
The ecological ’lichen guilds‘ hypothesis, i.e. communities of lichens growing on the same type of habitat and forming associations with the same photobiont species, have been proposed for cyanobacterial lichens (
In this study, we found that the most common photobiont in P. pinnatifida was Trebouxia OTU S02. All samples of P. pinnatifida were collected from rocks; however, some authors previously reported the same Treboxia OTU S02 from terricolous, saxicolous and corticolous Parmeliaceae (i.e. genera Cetraria Ach., Melanohalea O.Blanco et al., Montanelia Divakar et al., Protoparmelia M.Choisy and Rhizoplaca Zopf and species Xanthoparmelia coloradoensis Hale and Vulpicida juniperinus (L.) J.-E.Mattsson & M.J.Lai) (
Interestingly, although P. omphalodes was found to associate with two lineages of Trebouxia photobionts (i.e. OTU S05 and an unidentified lineage SUn1), it does not associate with Trebouxia OTU S02, which, on the other hand, was found to associate with P. discordans (two samples). However, P. discordans also associates with Trebouxia OTU S05. As those species differ in morphology and chemistry, we suggest that those differences might be related to the photobiont type. Although some researchers did not find any correlation between different chemotypes and the associated photobionts (e.g.
The created models, derived from MaxEnt, received high AUC scores, indicating high reliability of analyses (Table
Northern Hemisphere | Eurasia | America | |
P. discordans | 0.993 (SD = 0.001) | 0.992 (SD = 0.001) | – |
P. omphalodes | 0.980 (SD = 0.003 | 0.982 (SD = 0.002) | 0.767 (SD = 0.101) |
P. pinnatifida | 0.981 (SD = 0.003 | 0.986 (SD = 0.002) | 0.819 (SD = 0.064) |
The distribution of P. discordans is limited mainly by precipitation of the driest month (bio14), but two other factors that can influence the occurrence of this taxon, varied in analyses conducted for the Northern Hemisphere and Eurasia separately. While in the former analysis, annual mean temperature (bio1) and mean diurnal range (bio2) gave important contributions to the model, the latter analysis indicated maximum temperature of the warmest month (bio5) and temperature seasonality (bio4) as significant limiting factors. Additionally, in cases of P. omphalodes and P. pinnatifida, different variables gave various contributions to the models created for different study areas. Mean diurnal range (bio2) was the crucial limiting factor for Eurasian populations of P. omphalodes, while within the American range of this species, its occurrence depends on precipitation of the driest month (bio14). For the American distribution of P. pinnatifida, the annual mean temperature (bio1) significantly influenced the model and the distribution of Eurasian populations appears limited by the maximum temperature of the warmest month (bio5) (Table
Estimates of relative contributions of the environmental variables to the Maxent model.
Northern Hemisphere | Eurasia | America | |
P. discordans | bio14 (25.6) | bio14 (35.9) | – |
bio1 (18.8) | bio5 (15.2) | ||
bio2 (15.4) | bio4 (14.6) | ||
P. omphalodes | bio19 (21.1) | bio2 (27.8) | bio14 (48.2) |
bio4 (21) | bio19 (24.8) | bio15 (20.3) | |
bio2 (17.7) | bio4 (14.2) | bio2 (10.9) | |
P. pinnatifida | bio5 (17.7) | bio5 (24.6) | bio1 (42.2) |
bio14 (17.3) | bio14 (19.1) | bio14 (18) | |
bio4 (14.1) | bio4 (15.7) | bio8 (11.1) |
The PCA diagram (Figure
D\I | P. discordans | P. omphalodes | P. pinnatifida |
P. discordans | x | 0.791 | 0.703 |
P. omphalodes | 0.544 | x | 0.840 |
P. pinnatifida | 0.441 | 0.581 | x |
D\I | P. omphalodes | P. pinnatifida |
P. omphalodes | x | 0.968 |
P. pinnatifida | 0.821 | x |
D\I | P. discordans | P. omphalodes | P. pinnatifida |
P. discordans | x | 0.828 | 0.729 |
P. omphalodes | 0.587 | x | 0.820 |
P. pinnatifida | 0.468 | 0.564 | x |
According to published data (
PCA (Figure
Trebouxia OTUs associating with species from P. omphalodes group with the information about their distribution, substrata preferences and references.
OTUs | Distribution | Substrata | References |
S02 | Antarctica, Austria, Canada, Chile, Germany, Greenland, Iceland, Morocco, Norway, Poland, Portugal, Russia, Slovakia, Spain, Sweden, UK, USA | corticolous, saxicolous and terricolous |
|
S04 | Canada, Estonia, Germany, Netherlands, Poland, Sweden, Turkey, USA | corticolous and saxicolous |
|
S05 | Canada, Finland, Italy, Norway, Spain, Sweden, Turkey, USA | corticolous, saxicolous and terricolous |
|
SUn1 | Canada, Finland, Spain, Sweden | corticolous and terricolous |
|
SUn2 | Canada, Norway, Russia, Sweden | corticolous and terricolous |
|
1 | Pseudocyphellae marginal | 2 |
– | Pseudocyphellae marginal and laminal (at least in older parts of thalli) | 3 |
2 | Salazinic acid present | P. pinnatifida |
– | Protocetraric acid present | P. discordans (young thalli, rare) |
3 | Lobes 0.5–2 mm long and 1–2 mm wide, laminal pseudocyphellae predominantly connected with marginal pseudocyphellae, very few pseudocyphellae not starting from the lobe edges | P. pinnatifida |
– | Lobes 1–4 mm long and 1–3 mm wide, laminal pseudocyphellae predominantly not connected to the lobe margins | 4 |
4 | Protocetraric present | P. discordans |
– | Salazinic acid present | P. omphalodes |
We are grateful to the curators of all herbaria for the loan of specimens and reviewers for their helpful comments, Agnieszka Jabłońska and Magdalena Kosecka for help with molecular study, Paulina Dygner for help with TLC study and Andrzej Szczepański for help during field research. The research was supported by the Ministry of Science and Higher Education, project no. 2012/07/N/NZ8/00061 and BW/538-L150-B257-16 from the University of Gdansk, granted to EO.
Table S1. Database of localities used in the analyses with the bioclimatic values for each record
Data type: occurrence
Figure S2. PNO profiles created for P. discordans (A), P. omphalodes (B) and P. pinnatifida (C) in Northern Hemisphere
Data type: multimedia
Figure S3. PNO profiles created for P. discordans (A), P. omphalodes (B) and P. pinnatifida (C) in Eurasia
Data type: multimedia
Figure S4. PNO profiles created for P. omphalodes (A) and P. pinnatifida (B) in America
Data type: multimedia