Organisms Diversity & Evolution

, Volume 17, Issue 2, pp 351–363 | Cite as

Using multi-locus sequence data for addressing species boundaries in commonly accepted lichen-forming fungal species

  • Xin Zhao
  • Samantha Fernández-Brime
  • Mats Wedin
  • Marissa Locke
  • Steven D. Leavitt
  • H. Thorsten Lumbsch
Original Article

Abstract

Accurate species delimitations are of great importance for effectively characterizing biological diversity. Our criteria for delimiting species have changed dramatically over the last decades with the increasing availability of molecular data and improvement of analytical methods to evaluate these data. Whereas reciprocal monophyly is often seen as an indicator for the presence of distinct lineages, recently diverged species often fail to form monophyletic groups. At the same time, cryptic species have repeatedly been detected in numerous organismal groups. In this study, we addressed the species delimitation in the crustose lichen-forming fungal genus Diploschistes using multilocus sequence data from specimens representing 16 currently accepted species. Our results indicate the presence of previously undetected, cryptic species-level lineages in the subgenus Limborina. In the subgenus Limborina, samples from different continents currently classified under the same species were shown to be only distantly related. At the same time, in parts of subgen. Diploschistes characterized by short branches, none of the currently accepted species formed monophyletic groups. In spite of the lack of monophyly in phylogenetic reconstructions, a multispecies coalescent method provided support for eight of the nine accepted species in subgen. Diploschistes as distinct lineages. We propose to reduce D. neutrophilus to synonymy with D. diacapsis and point out that additional sampling will be necessary before accepting additional species in subgen. Limborina.

Keywords

BPP Diploschistes Graphidaceae Molecular phylogeny Species delimitation 

References

  1. Alors, D., Lumbsch, H. T., Divakar, P. K., Leavitt, S. D., & Crespo, A. (2016). An integrative approach for understanding diversity in the Punctelia rudecta species complex (Parmeliaceae, Ascomycota). PloS One, 11(2). doi:10.1371/journal.pone.0146537.
  2. Amo de Paz, G., Crespo, A., Cubas, P., Elix, J. A., & Lumbsch, H. T. (2012). Transoceanic dispersal and subsequent diversification on separate continents shaped diversity of the Xanthoparmelia pulla group (Ascomycota). PloS One, 7(6), e39683.CrossRefPubMedGoogle Scholar
  3. Arguello, A., Del Prado, R., Cubas, P., & Crespo, A. (2007). Parmelina quercina (Parmeliaceae, Lecanorales) includes four phylogenetically supported morphospecies. Biological Journal of the Linnean Society, 91(3), 455–467.CrossRefGoogle Scholar
  4. Camargo, A., Morando, M., Avila, L. J., & Sites, J. W., Jr. (2012). Species delimitation with ABC and other coalescent-based methods: a test of accuracy with simulations and an empirical example with lizards of the Liolaemus darwinii complex (Squamata: Liolaemidae). Evolution, 66, 2834–2849, doi:10.1111/j.1558-5646.2012.01640.x.Google Scholar
  5. Camargo, A., & Sites, J. (2013). Species delimitation: a decade after the renaissance, the species problem - ongoing issues. In I. Pavlinov (Ed.), InTech, ISBN: 978–953–51-0957-0. doi:10.5772/52664. Available from: http://www.intechopen.com/books/the-species-problem-ongoing-issues/species-delimitation-a-decade-after-the-renaissance.
  6. Carstens, B. C., Pelletier, T. A., Reid, N. M., & Satler, J. D. (2013). How to fail at species delimitation. Molecular Ecology, 22(17), 4369–4383. doi:10.1111/mec.12413.CrossRefPubMedGoogle Scholar
  7. Castresana, J. (2000). Selection of conserved blocks from multiple alignments for their use in phylogenetic analysis. Molecular Biology and Evolution, 17(4), 540–552.CrossRefPubMedGoogle Scholar
  8. Clauzade, G., & Roux, C. (1985). Likenoj de Okcidenta Europo. Ilustrita Determinlibro: Bulletin de la Societe Botanique du Centre-Ouest, Nouvelle Serie, Numero Special 7. Royan, France.Google Scholar
  9. Cornejo, C., & Scheidegger, C. (2015). Multi-gene phylogeny of the genus Lobaria: evidence of species-pair and allopatric speciation in East Asia. American Journal of Botany, 102(12), 2058–2073.CrossRefPubMedGoogle Scholar
  10. Coyne, J. A., & Orr, H. A. (2004). Speciation. Sunderland: Sinauer Associates.Google Scholar
  11. Crespo, A., & Lumbsch, H. T. (2010). Cryptic species in lichen-forming fungi. IMA Fungus, 1, 167–170.CrossRefPubMedPubMedCentralGoogle Scholar
  12. Darriba, D., Taboada, G. L., Doallo, R., & Posada, D. (2012). jModelTest 2: more models, new heuristics and parallel computing. Nature Methods, 9(8), 772.CrossRefPubMedPubMedCentralGoogle Scholar
  13. Degnan, J. H., & Rosenberg, N. A. (2009). Gene tree discordance, phylogenetic inference and the multispecies coalescent. Trends in Ecology & Evolution, 24(6), 332–340. doi:10.1016/j.tree.2009.01.009.CrossRefGoogle Scholar
  14. Divakar, P. K., Figueras, G., Hladun, N. L., & Crespo, A. (2010). Molecular phylogenetic studies reveal an undescribed species within the north American concept of Melanelixia glabra (Parmeliaceae). Fungal Diversity, 42, 47–55.CrossRefGoogle Scholar
  15. 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. Lichenologist, 32, 200–204.Google Scholar
  16. Fernandez-Brime, S., Llimona, X., Molnar, K., Stenroos, S., Hognabba, F., Bjoerk, C., et al. (2011). Expansion of the Stictidaceae by the addition of the saxicolous lichen-forming genus Ingvariella. Mycologia, 103(4), 755–763. doi:10.3852/10-287.CrossRefPubMedGoogle Scholar
  17. Fernández-Brime, S., Llimona, X., Lutzoni, F., & Gaya, E. (2013). Phylogenetic study of Diploschistes (lichen-forming Ascomycota: Ostropales: Graphidaceae), based on morphological, chemical, and molecular data. Taxon, 62(2), 267–280.CrossRefGoogle Scholar
  18. Fujisawa, T., & Barraclough, T. G. (2013). Delimiting species using single-locus data and the generalized mixed yule coalescent (GMYC) approach: a revised method and evaluation on simulated datasets. Systematic Biology, 62(5), 707–724. doi:10.1093/sysbio/syt033.CrossRefPubMedPubMedCentralGoogle Scholar
  19. Gardes, M., & Bruns, T. D. (1993). ITS primers with enhanced specificity for basidiomycetes - Application to the identification of mycorrhizae and rusts. Molecular Ecology, 2, 113–118.Google Scholar
  20. Guderley, R., Lumbsch, H. T., & Feige, G. B. (1997). Ingvariella, a new genus in the Thelotremataceae (lichenized Ascomycotina). Nova Hedwigia, 64(1–2), 147–154.Google Scholar
  21. Hodkinson, B. P., & Lendemer, J. C. (2011). Molecular analyses reveal semi-cryptic species in Xanthoparmelia tasmanica. Bibliotheca Lichenologica, 106, 108–119.Google Scholar
  22. Jaklitsch, W. M., Baral, H. O., Lücking, R., & Lumbsch, H. T. (2016). Ascomycota. In W. Frey (Ed.), Syllabus of plant families - Adolf Engler’s syllabus der Pflanzenfamilien (Vol. 1/2, 13th ed., p. 150). Stuttgart: Gebr. Borntraeger Verlagsbuchhandlung.Google Scholar
  23. Katoh, K., Asimenos, G., & Toh, H. (2009). Multiple alignment of DNA sequences with MAFFT. Methods in Molecular Biology, 537, 39–64.CrossRefPubMedGoogle Scholar
  24. Kingman, J. (1982). The coalescent. Stochastic Processes and their Applications, 13, 235–248.CrossRefGoogle Scholar
  25. Knowles, L. L., & Carstens, B. C. (2007). Delimiting species without monophyletic Gene trees. [article]. Systematic Biology, 56(6), 887–895. doi:10.1080/10635150701701091.CrossRefPubMedGoogle Scholar
  26. Kraichak, E., Parnmen, S., Lücking, R., & Lumbsch, H. T. (2014). Gintarasia and Xalocoa, two new genera to accommodate temperate to subtropical species in the predominantly tropical Graphidaceae (Ostropales, Ascomycota). Australian Systematic Botany, 26, 466–474.Google Scholar
  27. Kraichak, E., Divakar, P. K., Crespo, A., Leavitt, S. D., Nelsen, M. P., Lücking, R., et al. (2015). A tale of two hyper-diversities: diversification dynamics of the two largest families of lichenized fungi. Scientific Reports, 5, e10028.CrossRefGoogle Scholar
  28. Larena, I., Salazar, O., González, V., Julián, M. C., & Rubio, V. (1999). Design of a primer for ribosomal DNA internal transcribed spacer with enhanced specificity for ascomycetes. Journal of Biotechnology, 75, 187–194.Google Scholar
  29. Leavitt, S. D., Fankhauser, J. D., Leavitt, D. H., Porter, L. D., Johnson, L. A., & St Clair, L. L. (2011). Complex patterns of speciation in cosmopolitan "rock posy" lichens - Discovering and delimiting cryptic fungal species in the lichen-forming Rhizoplaca melanophthalma species-complex (Lecanoraceae, Ascomycota). Molecular Phylogenetics and Evolution, 59, 587–602, doi:10.1016/j.ympev.2011.03.020.Google Scholar
  30. Leavitt, S. D., Johnson, L., & St Clair, L. L. (2011a). Species delimitation and evolution in morphologically and chemically diverse communities of the lichen-forming genus Xanthoparmelia (Parmeliaceae, Ascomycota) in western North America. American Journal of Botany, 98(2), 175–188. doi:10.3732/ajb.1000230.CrossRefPubMedGoogle Scholar
  31. Leavitt, S. D., Johnson, L. A., Goward, T., & St. Clair, L. L. (2011b). Species delimitation in taxonomically difficult lichen-forming fungi: an example from morphologically and chemically diverse Xanthoparmelia (Parmeliaceae) in North America. Molecular Phylogenetics and Evolution, 60, 317–332.CrossRefPubMedGoogle Scholar
  32. Leavitt, S. D., Esslinger, T. L., Divakar, P. K., & Lumbsch, H. T. (2012). Miocene and Pliocene dominated diversification of the lichen-forming fungal genus Melanohalea (Parmeliaceae, Ascomycota) and Pleistocene population expansions. BMC Evolutionary Biology, 12, 176.CrossRefPubMedPubMedCentralGoogle Scholar
  33. Leavitt, S. D., Lumbsch, H. T., Stenroos, S., & St Clair, L. L. (2013). Pleistocene speciation in north American lichenized fungi and the impact of alternative species circumscriptions and rates of molecular evolution on divergence estimates. PloS One, 8(12). doi:10.1371/journal.pone.0085240.
  34. Leavitt, S. D., Divakar, P. K., Ohmura, Y., Wang, L. S., Esslinger, T. L., & Lumbsch, H. T. (2015a). Who’s getting around? Assessing species diversity and phylogeography in the widely distributed lichen-forming fungal genus Montanelia (Parmeliaceae, Ascomycota). Molecular Phylogenetics and Evolution, 90, 85–96. doi:10.1016/j.ympev.2015.04.029.CrossRefPubMedGoogle Scholar
  35. Leavitt, S. D., Moreau, C. S., & Lumbsch, H. T. (2015b). The dynamic discipline of species delimitation: progress toward effectively recognizing species boundaries in natural populations. In D. K. Upreti, P. K. Divakar, V. Shukla, & R. Bajpai (Eds.), Recent advances in lichenology (pp. 11–44). India: Springer.CrossRefGoogle Scholar
  36. Leavitt, S. D., Divakar, P. K., Crespo, A., & Lumbsch, H. T. (2016). A matter of time – understanding the limits of the power of molecular data for delimiting species boundaries. Herzogia, submitted. Herzogia, 29, submtted.Google Scholar
  37. Liu, Y. J., Whelen, S., & Hall, B. D. (1999). Phylogenetic relationships among ascomycetes: evidence from an RNA polymerase II subunit. Molecular Biology and Evolution, 16, 1799–1808.Google Scholar
  38. Llimona Pages, X. (1974). Las Communidades de Liquenes de los Yesos de Espana: Universidad de Barcelona. Barcelona: Secretariado de Publicationes.Google Scholar
  39. Lumbsch, H. T. (1988). The identity of Diploschistes gypsaceus. The Lichenologist, 20(1), 19–24.CrossRefGoogle Scholar
  40. Lumbsch, H. T. (1989). Die holarktischen Vertreter der Flechtengattung Diploschistes (Thelotremataceae). Journal of the Hattori Botanical Laboratory, 66, 133–196.Google Scholar
  41. Lumbsch, H. T., & Elix, J. A. (1989). Taxonomy of some Diploschistes spp (lichenized ascomycetes, Thelotremataceae) containing gyrophoric acid. Plant Systematics and Evolution, 167(3–4), 195–199.CrossRefGoogle Scholar
  42. Lumbsch, H. T., & Huhndorf, S. M. (2010). Myconet volume 14. Part one. Outline of Ascomycota - 2009. Fieldiana (Life and Earth Sciences), 1, 1–42.CrossRefGoogle Scholar
  43. Lumbsch, H. T., & Leavitt, S. D. (2011). Goodbye morphology? A paradigm shift in the delimitation of species in lichenized fungi. Fungal Diversity, 50, 59–72.CrossRefGoogle Scholar
  44. Lumbsch, H. T., & Tehler, A. (1998). A cladistic analysis of the genus Diploschistes (Ascomycotina, Thelotremataceae). Bryologist, 101(3), 398–403.CrossRefGoogle Scholar
  45. Mangold, A., Martin, M. P., Lücking, R., & Lumbsch, H. T. (2008). Molecular phylogeny suggests synonymy of Thelotremataceae within Graphidaceae (Ascomycota : Ostropales). Taxon, 57, 476–486.Google Scholar
  46. Martín, M. P., LaGreca, S., & Lumbsch, H. T. (2003). Molecular phylogeny of Diploschistes inferred from ITS sequence data. The Lichenologist, 35(1), 27–32.CrossRefGoogle Scholar
  47. Matheny, P. B., Liu, Y. J., Ammirati, J. F., & Hall, B. D. (2002). Using RPB1 and RPB2 nucleotide sequences (Inocybe; Agaricales). American Journal of Botany, 89, 688–698.Google Scholar
  48. Mayr, E. (1963). Animal species and evolution. Cambridge: Harvard University Press.CrossRefGoogle Scholar
  49. Nixon, K., & Wheeler, Q. (1990). An amplification of the phylogenetic species concept. Cladistics, 6, 211–223.CrossRefGoogle Scholar
  50. Nylander, J. A. A., Wilgenbusch, J. C., Warren, D. L., & Swofford, D. L. (2008). AWTY (are We there yet?): a system for graphical exploration of MCMC convergence in Bayesian phylogenetics. Bioinformatics, 24, 581–583.CrossRefPubMedGoogle Scholar
  51. Otálora, M. A. G., Martínez, I., Aragón, G., & Molina, M. C. (2010). Phylogeography and divergence date estimates of a lichen species complex with a disjunct distribution pattern. American Journal of Botany, 97, 216–223.CrossRefPubMedGoogle Scholar
  52. Pant, G., & Upreti, D. K. (1993). The lichen genus Diploschistes in India and Nepal. The Lichenologist, 25(1), 33–50.CrossRefGoogle Scholar
  53. Parnmen, S., Rangsiruji, A., Mongkolsuk, P., Boonpragob, K., Nutakki, A., & Lumbsch, H. T. (2012). Using phylogenetic and coalescent methods to understand the species diversity in the Cladia aggregata Complex (Ascomycota, Lecanorales). PloS One, 7(12), e52245. doi:10.1371/journal.pone.0052245.CrossRefPubMedPubMedCentralGoogle Scholar
  54. Parnmen, S., Cáceres, M. E. S., Lücking, R., & Lumbsch, H. T. (2013). Myriochapsa and Nitidochapsa, two new genera in Graphidaceae (Ascomycota: Ostropales) for chroodiscoid species in the Ocellularia clade. Bryologist, 116, 127–133.CrossRefGoogle Scholar
  55. Printzen, C. (2010). Progress in botany. In U. Lüttge, W. Beyschlag, B. Büdel, & D. Francis (Eds.), Lichen systematics: the role of morphological and molecular data to reconstruct phylogenetic relationships (Vol. 71, pp. 233–275). Botany: Progress in Botany.Google Scholar
  56. Rambaut, A. (2009). FigTree 1.2.2. http://tree.bio.ed.ac.uk/software/figtree/.
  57. Rannala, B. (2015). The art and science of species delimitation. Current Zoology, 61(5), 846–853.CrossRefGoogle Scholar
  58. Rannala, B., & Yang, Z. (2013). Improved reversible jump algorithms for Bayesian species delimitation. Genetics, 194, 245–253.CrossRefPubMedPubMedCentralGoogle Scholar
  59. Rivas Plata, E. (2011). Historical biogeography, ecology and systematics of the family Graphidaceae (lichenized Ascomycota: Ostropales). Chicago: University of Illinois at Chicago.Google Scholar
  60. Rivas Plata, E., Lücking, R., Sipman, H. J. M., Mangold, A., & Lumbsch, H. T. (2010). A world-wide key to the thelotremoid Graphidaceae, excluding the Ocellularia-Myriotrema-Stegobolus clade. The Lichenologist, 42, 139–185.CrossRefGoogle Scholar
  61. Rivas Plata, E., Parnmen, S., Staiger, B., Mangold, A., Frisch, A., Weerakoon, G., et al. (2013). A molecular phylogeny of Graphidaceae (Ascomycota: Lecanoromycetes: Ostropales) including 437 species. MycoKeys, 6, 55–94.CrossRefGoogle Scholar
  62. Ronquist, F., Teslenko, M., van der Mark, P., Ayres, D. L., Darling, A., Höhna, S., et al. (2012). MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology, 61, 539–542.CrossRefPubMedPubMedCentralGoogle Scholar
  63. Schmitt, I., Crespo, A., Divakar, P. K., Fankhauser, J., Herman-Sackett, E., Nelsen, M. P., et al. (2009). New primers for single-copy protein-coding genes for fungal systematics. Persoonia - Molecular Phylogeny and Evolution of Fungi, 23, 35–40.Google Scholar
  64. Sites, J. W., & Marshall, J. C. (2004). Operational criteria for delimiting species. Annual Review of Ecology, Evolution, and Systematics, 35(1), 199–227. doi:10.1146/annurev.ecolsys.35.112202.130128.CrossRefGoogle Scholar
  65. Stamatakis, A. (2006). RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics, 22, 2688–2690.CrossRefPubMedGoogle Scholar
  66. Stiller, J. W., & Hall, B. D. (1997). The origin of red algae: implications for plastid evolution. Proceedings of the National Academy of Sciences of the United States of America, 94, 4520-4525.Google Scholar
  67. Thell, A., Elix, J. A., & Søchting, U. (2009). Xanthoparmelia lineola s. L. in Australia and North America. Bibliotheca Lichenologica, 99, 393–404.Google Scholar
  68. 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.Google Scholar
  69. Yang, Z. (2015). The BPP program for species tree estimation and species delimitation. Current Zoology, 61(5), 854–865.CrossRefGoogle Scholar
  70. Yang, Z., & Rannala, B. (2010a). Bayesian species delimitation using multilocus sequence data. Proceedings of the National Academy of Sciences, 107(20), 9264–9269. doi:10.1073/pnas.0913022107.CrossRefGoogle Scholar
  71. Yang, Z., & Rannala, B. (2010b). Bayesian species delimitation using multilocus sequence data. Proceedings of the National Academy of Sciences of the United States of America, 107, 9264–9269.CrossRefPubMedPubMedCentralGoogle Scholar
  72. Yang, Z., & Rannala, B. (2014). Unguided species delimitation using DNA sequence data from multiple loci. Molecular Biology and Evolution, 31, 3125–3135.CrossRefPubMedPubMedCentralGoogle Scholar
  73. Zhang, C., Zhang, D. X., Zhu, T., & Yang, Z. (2011). Evaluation of a Bayesian coalescent method of species delimitation. Systematic Biology, 60, 747–761.Google Scholar
  74. Zhao, X., Zhang, L. L., Zhao, Z. T., Wang, W. C., Leavitt, S. D., & Lumbsch, H. T. (2015). A molecular phylogeny of the lichen genus Lecidella focusing on species from mainland China. PloS One, 10(9). doi:10.1371/journal.pone.0139405.
  75. Zhou, S., & Stanosz, G. R. (2001). Primers for amplification of mt SSU rDNA, and a phylogenetic study of Botryosphaeria and associated anamorphic fungi. Mycological Research, 105, 1033–1044.Google Scholar
  76. Zoller, S., Scheidegger, C., & Sperisen, C. (1999). PCR primers for the amplification of mitochondrial small subunit ribosomal DNA of lichen-forming ascomycetes. Lichenologist, 31, 511–516.Google Scholar

Copyright information

© Gesellschaft für Biologische Systematik 2017

Authors and Affiliations

  • Xin Zhao
    • 1
    • 2
  • Samantha Fernández-Brime
    • 3
  • Mats Wedin
    • 3
  • Marissa Locke
    • 4
  • Steven D. Leavitt
    • 5
  • H. Thorsten Lumbsch
    • 5
  1. 1.College of Life SciencesLiaocheng UniversityLiaochengPeople’s Republic of China
  2. 2.College of Life SciencesShandong Normal UniversityJinanPeople’s Republic of China
  3. 3.Department of BotanySwedish Museum of Natural HistoryStockholmSweden
  4. 4.Department of Pathology and ImmunologyWashington University School of MedicineSt. LouisUSA
  5. 5.Science and EducationThe Field MuseumChicagoUSA

Personalised recommendations