Skip to main content Skip to main navigation menu Skip to site footer
Type: Article
Published: 2024-06-13
Page range: 20-36
Abstract views: 8
PDF downloaded: 0

Morphological and multi-gene phylogenetic analyses of two new species of Phyllosticta (Phyllostictaceae, Botryosphaeriales) from Southwestern China

International Fungal Research and Development Centre, Institute of Highland Forest Science, Chinese Academy of Forestry, Kunming 650224, Yunnan, P. R. of China. Nanjing Forestry University, Nanjing 210037, P. R. of China.
International Fungal Research and Development Centre, Institute of Highland Forest Science, Chinese Academy of Forestry, Kunming 650224, Yunnan, P. R. of China.
International Fungal Research and Development Centre, Institute of Highland Forest Science, Chinese Academy of Forestry, Kunming 650224, Yunnan, P. R. of China.
International Fungal Research and Development Centre, Institute of Highland Forest Science, Chinese Academy of Forestry, Kunming 650224, Yunnan, P. R. of China.
2 new species microfungi saprobe sexual stage taxonomy Fungi


Phyllosticta is an important genus of pathogenic fungi with worldwide distribution. Phyllosticta species also occur as endophytes and saprobes on various plant hosts. Multi-gene phylogenetic analysis based on sequence data of ITS, LSU, TEF1 and morphological evidence supported the introduction of two new species, which were isolated from dead leaves in China. Phylogenetically, the new species, P. savannaensis and P. ovalina, formed two well-separated lineages in the Phyllosticta capitalensis and Phyllosticta cruenta species complexes, respectively. P. savannaensis is closely related to P. capitalensis, but formed a separate branch (BS= 100%/BYPP= 1.0), while P. ovalina and P. hubeiensis clustered together with high support (BS= 100%/BYPP= 1.0). Both new species exhibited the typical characteristics of the genus Phyllosticta but could be distinguished from closely related species. P. savannaensis can be differentiated from the sexual stage of other Phyllosticta species by the length to width ratio of the ascospores and shape of appendages. The ascospores of P. ovalina are oval, which distinguish it from other species of Phyllosticta.


  1. van der Aa, H.A. (1973) Studies in Phyllosticta. Vol. 5. Centraalbureau voor Schimmelcultures, 110 pp.
  2. van der Aa, H.A. & Vanev, S. (2002) A revision of the species described in Phyllosticta. Netherlands (Utrecht): Centraalbureau voor Schimmelcultures (CBS). 510 pp.
  3. Baayen, R.P., Bonants, P.J.M., Verkley, G., Carroll, G.C., Van Der Aa, H.A., De Weerdt, M., Van Brouwershaven, I.R., Schutte, G.C., Maccheroni, W., De Blanco, C.G. & Azevedo, J.L. (2002) Nonpathogenic isolates of the citrus black spot fungus, Guignardia citricarpa, identified as a cosmopolitan endophyte of woody plants, G. mangiferae (Phyllosticta capitalensis). Phytopathology 92 (5): 464–477.
  4. Bhunjun, C.S., Niskanen, T., Suwannarach, N., Wannathes, N., Chen, Y.-J., McKenzie, E.H.C., Maharachchikumbura, S.S.N., Buyck, B., Zhao, C.-L., Fan, Y.-G., Zhang, J.-Y., Dissanayake, A.J., Marasinghe, D.S., Jayawardena, R.S., Kumla, J., Padamsee, M., Chen, Y.-Y., Liimatainen, K., Ammirati, J.F., Phukhamsakda, C., Liu, J.-K, Phonrob, W., Randrianjohany, É., Hongsanan, S., Cheewangkoon, R., Bundhun, D., Khuna, S., Yu, W.-J, Deng, L.-S, Lu, Y.-Z, Hyde, K.D. & Lumyong, S. (2022) The numbers of fungi: are the most speciose genera truly diverse? Fungal Diversity 114 (1): 387–462.
  5. Carbone, I. & Kohn, L.M. (1999) A method for designing primer sets for speciation studies in filamentous ascomycetes. Mycologia 91 (3): 553–556.
  6. Crous, R.W., Seifert, K.A. & Ruiz, R.C. (1996) Microfungi associated with Podocarpus leaf litter in South Africa. South African Journal of Botany 62 (2): 89–98.
  7. Crous, P.W., Slippers, B., Wingfield, M.J., Rheeder, J., Marasas, W.F.O., Philips, A.J.L., Alves, A., Burgess, T., Barber, P. & Groenewald, J.Z. (2006) Phylogenetic lineages in the Botryosphaeriaceae. Studies in Mycology 55 (1): 235–253.
  8. Fries, E.M. (1849) Summa Vegetabilium Scandinaveae. Sectio posterior. Typographia Academica, Leipzig, pp. 259–572.
  9. Glienke, C., Pereira, O.L., Stringari, D., Fabris, J., Kava-Cordeiro, V., Galli-Terasawa, L., Cunnington, J., Shivas, R.G., Groenewald, J.Z. & Crous, P.W. (2011) Endophytic and pathogenic Phyllosticta species, with reference to those associated with citrus black spot. Persoonia 26 (1): 47–56.
  10. Guarnaccia, V., Groenewald, J.Z., Li, H., Glienke, C., Carstens, E., Hattingh, V., Fourie, P. & Crous, P.W. (2017) First report of Phyllosticta citricarpa and description of two new species, P. paracapitalensis and P. paracitricarpa from citrus in Europe. Studies in Mycology 87 (1): 161–185.
  11. Guarnaccia, V., Gehrmann, T., Silva-Junior, J., Fourie, P.H., Haridas, S., Vu, D., Vu, D., Spatafora, J., Martin, F.M., Robert, V., Grigoriev, I.V., Groenewald, J.Z. & Crous, P.W. (2019) Phyllosticta citricarpa and sister species of global importance to Citrus. Molecular Plant Pathology 20 (12): 1619–1635.
  12. Hall, T.A. (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. In Nucleic Acids Symposium Series 41: 95–98.
  13. Hyde, K.D. (1995) Fungi from palms. XX. The genus Guignardia. Sydowia 47: 180–198.
  14. Jiang, Y., Zhang, Z., Zhang, J., Wang, S. & Zhang, X. (2023) Morphological and phylogenetic analyses reveal three new species of Phyllosticta (Botryosphaeriales, Phyllostictaceae) in China. Journal of Fungi 10 (1): 7.
  15. Li, X.-H, Wu, H.-X., Li, J.-C, Chen, H. & Wang, W. (2020) The insights into the evolutionary history of Translucidithyrium: based on a newly-discovered species. MycoKeys 76: 1.
  16. Li, X.-H., Wu, H.-X., Li, J.-C., Song, J.-Y., Wang, Q., Promputtha, I. & Wijayawardene, N.N. (2022a) Two new species of Allodiatrype from monsoon evergreen broad-leaved forest in Pu’er, Yunnan, China. Chiang Mai Journal of Science 49 (3): 565–580.
  17. Li, J.-C., Wu, H.-X., Li, Y., Li, X.-H., Song, J.-Y., Suwannarach, N. & Wijayawardene, N.N. (2022b). Taxonomy, phylogenetic and ancestral area reconstruction in Phyllachora, with four novel species from Northwestern China. Journal of Fungi 8 (5): 520.
  18. Li, J.-C., Wu, H.-X. & Song, J.-Y. (2023) Two new Phyllachora species in Southwest China. Phytotaxa 578 (3): 275–285.
  19. Lin, S., Sun, X., He, W. & Zhang, Y. (2017) Two new endophytic species of Phyllosticta (Phyllostictaceae, Botryosphaeriales) from southern China. Mycosphere 8 (2): 1273–1288.
  20. Liu, J.-K., Phookamsak, R., Doilom, M., Wikee, S., Li, Y.-M., Ariyawansha, H., Boonmee, S., Chomnunti, P., Dai, D.-Q., Bhat, J.D., Romero, A.I., Zhuang, W.-Y., Monkai, J., Jones, E.B.G., Chukeatirote, E., Ko Ko, T.W.K., Zhao, Y.-C., Wang, Y. & Hyde, K.D. (2012) Towards a natural classification of Botryosphaeriales. Fungal Diversity 57: 149–210.
  21. Miller, M.A., Pfeiffer, W., Schwartz, T. (2010) Creating the CIPRES Science Gateway for inference of large phylogenetic trees. [http://www.phyloorg/sub_sections/portal]
  22. Norphanphoun, C., Hongsanan, S., Gentekaki, E., Chen, Y.J., Kuo, C.H. & Hyde, K.D. (2020) Differentiation of species complexes in Phyllosticta enables better species resolution. Mycosphere 11: 2542–2628.
  23. Okane, I., Nakagiri, A., Ito, T. & Lumyong, S. (2003) Extensive host range of an endophytic fungus, Guignardia endophyllicola (anamorph: Phyllosticta capitalensis). Mycoscience 44 (5): 353–363.
  24. Paradis, E. & Schliep, K. (2019) APE 5.0: an environment for modern phylogenetics and evolutionary analyses in R. Bioinformatics 35 (3): 526–528. bioinformatics/bty633
  25. Persoon, C.H. (1818) Traité sur les champignons comestibles, contenant l’indication des espèces nuisibles; a l’histoire des champignons. Paris (France): Belin-Leprieur.
  26. Rashmi, M., Kushveer, J.S. & Sarma, V.V. (2019) A worldwide list of endophytic fungi with notes on ecology and diversity. Mycosphere 10 (1): 798–1079.
  27. Ronquist, F., Teslenko, M., Van Der Mark, P., Ayres, D.L., Darling, A., Höhna, S., Larget, B., Liu, L., Suchard, M.A. & Huelsenbeck, J.P. (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology 61 (3): 539–542.
  28. Schoch, C.L., Shoemaker, R.A., Seifert, K.A., Hambleton, S., Spatafora, J.W. & Crous, P.W. (2006) A multigene phylogeny of the Dothideomycetes using four nuclear loci. Mycologia 98 (6): 1041–1052.
  29. Seaver, F.J. (1922) Phyllostictaceae. North American Flora 6: 3–84.
  30. Song, J.-Y., Wu, H.-X., Li, J.-C., Ding, W.-F., Gong, C.-L., Zeng, X.-Y., Wijayawardene, N. & Yang, D.-X. (2023) Taxonomy and evolution history of two new litter-decomposing Ciliochorella (Amphisphaeriales, Sporocadaceae). MycoKeys 100: 95–121.
  31. Stamatakis, A. (2014) RAxML version 8: A tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30 (9): 1312–1313.
  32. Sui, X.-N., Guo, M.-J., Zhou, H. & Hou, C.-L. (2023) Four new species of Phyllosticta from China based on morphological and phylogenetic characterization. Mycology 14 (3): 190–203.
  33. Viala, P. & Ravaz, L. (1892) Sur la dénomination botanique (Guignardia bidwellii) du black-rot. Typogr. Charles Boehm.
  34. Vilgalys, R. & Hester, M. (1990) Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. Journal of Bacteriology 172 (8): 4238–4246.
  35. Wang, X., Chen, G., Huang, F., Zhang, J., Hyde, K.D. & Li, H. (2012) Phyllosticta species associated with citrus diseases in China. Fungal diversity 52: 209–224.
  36. Wang, C.-B., Yang, J., Li, Y., Xue, H., Piao, C.-G. & Jiang, N. (2023a) Multi-gene phylogeny and morphology of two new Phyllosticta (Phyllostictaceae, Botryosphaeriales) species from China. MycoKeys 95: 189.
  37. Wang, W., Xiong, T., Zeng, Y., Li, W., Jiao, C., Xu, J. & Li, H. (2023b) Clonal expansion in multiple Phyllosticta species causing citrus black spot or similar symptoms in China. Journal of Fungi 9 (4): 449.
  38. White, T.J., Bruns, T., Lee, S. & Taylor, J.W. (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR protocols: a guide to methods and applications 18 (1): 315–322.
  39. Wijayawardene, N.N., Hyde, K.D., Dai, D.-Q., Sánchez-García, M., Goto, B.T., Saxena, R.K., Erdoğdu, M., Selçuk, F., Rajeshkumar, K.C., Aptroot, A., Błaszkowski, J., Boonyuen, N., da Silva, G.A., de Souza, F.A., Dong, W., Ertz, D., Haelewaters, D., Jones, E.B.G., Karunarathna, S.C., Kirk, P.M., Kukwa, M., Kumla, J., Leontyev, D.V., Lumbsch, H.T., Maharachchikumbura, S.S.N., Marguno, F., Martínez-Rodríguez, P., Mešić, A., Monteiro, J.S., Oehl, F., Pawłowska, J., Pem, D., Pfliegler, W.P., Phillips, A.J.L., Pošta, A, He, M.-Q., Li, J.X., Raza, M., Sruthi, O.P., Suetrong, S., Suwannarach, N., Tedersoo, L., Thiyagaraja, V., Tibpromma, S., Tkalčec, Z., Tokarev, Y.S., Wanasinghe, D.N., Wijesundara, D.S.A, Wimalaseana, S.D.M.K, Madrid, H., Zhang, G., Gao, Y., Sánchez-Castro, I., Tang, L.-Z., Stadler, M., Yurkov, A.M. & Thines, M. (2022) Outline of Fungi and fungus-like taxa—2021. Mycosphere 13 (1): 53–453. https://doi. org/10.5943/mycosphere/13/1/2
  40. Wikee, S., Udayanga, D., Crous, P.W., Chukeatirote, E., McKenzie, E.H., Bahkali, A.H., Dai, D.Q. & Hyde, K.D. (2011) Phyllosticta—an overview of current status of species recognition. Fungal Diversity 51 (1): 43–61.
  41. Wikee, S., Lombard, L., Crous, P.W., Nakashima, C., Motohashi, K., Chukeatirote, E., Alias, S.A., McKenzie, E.H.C. & Hyde, K.D. (2013a) Phyllosticta capitalensis, a widespread endophyte of plants. Fungal Diversity 60 (1): 91–105.
  42. Wikee, S., Lombard, L., Nakashima, C., Motohashi, K., Chukeatirote, E., Cheewangkoon, R., McKenzie, E.H.C., Hyde, K.D. & Crous, P.W. (2013b) A phylogenetic re-evaluation of Phyllosticta (Botryosphaeriales). Studies in Mycology 76 (1): 1–29.
  43. Wong, M.-H., Crous, P.W., Henderson, J., Groenewald, J.Z. & Drenth, A. (2012) Phyllosticta species associated with freckle disease of banana. Fungal Diversity 56 (1): 173–187.
  44. Wu, H.-X., Li, Y.-M., Ariyawansa, H.A., Li, W.-J., Yang, H. & Hyde, K.D. (2014) A new species of Microthyrium from Yunnan, China. Phytotaxa 176: 213–218.
  45. Wu, H.-X., Ren, W.-B., Li, X.-H. & Yang, Z.-X. (2021) A new species of foliicolous lichenized fungi from southwest China: Calopadia ruiliensis sp. nov. Turkish Journal of Botany 45 (4): 371–377.
  46. Wulandari, N.F., To-Anun, C., Hyde, K.D., Duong, L.M., de Gruyter, J., Meffert, J.P, Groenewald, J.Z. & Crous, P.W. (2009) Phyllosticta citriasiana sp. nov., the cause of Citrus tan spot of Citrus maxima in Asia. Fungal Diversity 34 (1): 23–39.
  47. Wulandari, N.F., To-Anun, C., McKenzie, E.H.C. & Hyde, K.D. (2011) Guignardia bispora and G. ellipsoidea spp. nov. and other Guignardia species from palms (Arecaceae). Mycosphere 2 (2): 115–128.
  48. Wulandari, N.F., Bhat, D.J. & To-anun, C. (2013) A modern account of the genus Phyllosticta. Plant Pathology & Quarantine 3 (2): 145–159.
  49. Zhang, Z., Liu, X., Zhang, X. & Meng, Z. (2022) Morphological and phylogenetic analyses reveal two new species and a new record of Phyllosticta (Botryosphaeriales, Phyllostictaceae) from Hainan, China. MycoKeys 91: 1–23.