Abstract
Pine needle diseases, such as red band and brown spot needle blight, are serious pine diseases that threatens forests in many countries. Several outbreaks have been reported resulting in loss of productivity and mortality in both exotic and native plantations of Pinus spp. Symptomatology of these two diseases is quite similar and characterized by the appearance of yellowish areas/bands on hosts’ leaves that subsequently lead to the appearance of more extensive lesions and/or necrotic areas. In an attempt to understand the main causes of needle blight-like disease symptoms a study was carried in two pine stands that were apparently affected by red band and brown spot needle blights. Needles showing spots and/or bands with fruiting bodies were sampled. From 25 pine trees samples, 82 fungal isolates were successfully retrieved. The most common fungal genera were Pestalotiopsis (42.68%, n = 35), Rhizosphaera (28.04%, n = 23) and Cladosporium (9.75%, n = 8). Seven isolates could not be assigned to a species through molecular identification by ITS sequence analysis, potentially representing novel taxa. Based on multilocus phylogenetic analyses, using ITS, tub2 and tef1-α sequences, and morphological data, we propose three novel fungal species: Didymocyrtis pini sp. nov., Pestalotiopsis iberica sp. nov. and Rhizosphaera pinicola sp. nov. These species are potential active players in the symptomatology initially associated to red band and brown spot needle blight diseases. Although the pathogenicity of these fungi needs to be confirmed, this study suggests a high complexity underlying fungal species associated with these diseases which may impact disease epidemiology and management.
Similar content being viewed by others
References
Adams, J. C., & Orehart, A. L. (1982). Decline and death of Pinus spp. in Delaware Caused by Bursaphelenchus xylophilus. Journal of Nematology, 14(3), 382–385.
Alves, A., Correia, A., Luque, J., & Phillips, A. (2004). Botryosphaeria corticola sp. nov. on Quercus species, with notes and description of Botryosphaeria stevensii and its anamorph, Diplodia mutila. Mycologia, 96(3), 598–613. https://doi.org/10.1080/15572536.2005.11832956
Alves, A., Phillips, A. J. L., Henriques, I., & Correia, A. (2007). Rapid differentiation of species of Botryosphaeriaceae by PCR fingerprinting. Research in Microbiology, 158, 112–121. https://doi.org/10.1016/j.resmic.2006.10.003
Bednářová, M., Dvořák, M., Janoušek, J., & Jankovský, L. (2013). Other foliar diseases of coniferous trees. In P. Gonthier & G. Nicolotti (Eds.), Infectious forest diseases (pp. 458–487).
Bensch, K., Groenewald, J. Z., Dijksterhuis, J., Andersen, B., Summerell, B. A., Shin, H.-D., Dugan, F. M., Schroers, H.-J., Braun, U., & Crous, P. W. (2010). Species and ecological diversity within the Cladosporium cladosporioides complex (Davidiellaceae, Capnodiales). Studies in Mycology, 67, 1–94. https://doi.org/10.3114/sim.2010.67.01
Bezos, D., Martínez-Álvarez, P., Sanz-ros, A. V., Martín-García, J., Fernandez, M. M., & Diez, J. J. (2018). Fungal communities associated with Bark Beetles in Pinus radiata Plantations in Northern Spain affected by Pine Pitch Canker, with special focus on Fusarium Species. Forests, 9, 1–20. https://doi.org/10.3390/f9110698
Blank, L., Martín-García, J., Bezos, D., Vettraino, A. M., Krasnov, H., Lomba, J. M., Fernández, M., & Diez, J. J. (2019). Factors affecting the distribution of pine pitch canker in Northern Spain. Forests, 10, 1–16. https://doi.org/10.3390/f10040305
Botella, L., & Diez, J. J. (2011). Phylogenetic diversity of fungal endophytes in Spanish stands of Pinus halepensis. Fungal Diversity, 47, 9–18. https://doi.org/10.1007/s13225-010-0061-1
Bußkamp, J., Langer, G. J., & Langer, E. J. (2020). Sphaeropsis sapinea and fungal endophyte diversity in twigs of Scots pine (Pinus sylvestris) in Germany. Mycological Progress, 19, 985–999. https://doi.org/10.1007/s11557-020-01617-0
Crous, P. W., Wingfield, M. J., Schumacher, R. K., Summerell, B. A., Giraldo, A., Gené, J., Guarro, J., Wanasinghe, D. N., Hyde, K. D., Camporesi, E., Jones, E. B. G., Thambugala, K. M., Malysheva, E. F., Malysheva, V. F., Acharya, K., Álvarez, J., Alvarado, P., Assefa, A., Barnes, C. W., et al. (2014). Fungal Planet description sheets: 281–319. Persoonia, 33, 212–289. https://doi.org/10.3767/003158514X685680
Crous, P. W., Wingfield, M. J., Burgess, T. I., Hardy, G. E. S. J., Gené, J., Guarro, J., Baseia, I. G., García, D., Gusmão, L. F. P., Thangavel, R., Adamčík, S., Barili, A., Barnes, C. W., Bezerra, J. D. P., Bordallo, J. J., Santiago, A. L. C. M. D. A., De Oliveira, L. F., De Souza, C. A. F., & Déniel, F. (2018). Fungal Planet description sheets: 716–784. Persoonia, 40, 240–393. https://doi.org/10.3767/persoonia.2018.40.10
Diederich, P., & Kocourkova, J. (2007). The lichenicolous Phoma species (coelomycetes) on Cladonia. The Lichenologist, 39, 153–163. https://doi.org/10.1017/S0024282907006044
Diederich, P., Lawrey, J. D., & Ertz, D. (2018). The 2018 classification and checklist of lichenicolous fungi, with 2000 non-lichenized, obligately lichenicolous taxa. The Bryologist, 121(3), 340–425. https://doi.org/10.1639/0007-2745-121.3.340
Ertz, D., Diederich, P., Lawrey, J. D., Berger, F., Freebury, C. E., Coppins, B., Gardiennet, A., & Hafellner, J. (2015). Phylogenetic insights resolve Dacampiaceae (Pleosporales) as polyphyletic: Didymocyrtis (Pleosporales, Phaeosphaeriaceae) with Phoma - like anamorphs resurrected and segregated from Polycoccum (Trypetheliales, Polycoccaceae fam. nov.). Fungal Diversity, 74, 53–89. https://doi.org/10.1007/s13225-015-0345-6
European and Mediterranean Plant Protection Organization (EPPO). (2020). EPPO A1 and A2 Lists of pests recommended for regulation as quarantine pests. In EPPO Standards (Vol. 2). https://gd.eppo.int/download/standard/2/pm1-002-29-en.pdf
European Forest Institute (EFI). (2020). A Mediterranean Forest Research Agenda – MFRA 2010–2020.
Giordano, L., Gonthier, P., Varese, G. C., Miserere, L., & Nicolotti, G. (2009). Mycobiota inhabiting sapwood of healthy and declining Scots pine (Pinus sylvestris L.) trees in the Alps. Fungal Diversity, 38, 69–83.
Goldberg, N. P. (2017). Rhizosphaera needle cast disease of blue spruce.
Gonçalves, M. F. M., Esteves, A. C., & Alves, A. (2020). Revealing the hidden diversity of marine fungi in Portugal with the description of two novel species, Neoascochyta fuci sp. nov. and Paraconiothyrium salinum sp. nov. International Journal of Systematic and Evolutionary Microbiology, 70, 5337–5354. https://doi.org/10.1099/ijsem.0.004410
Hall, T. A. (1999). BioEdit a user-friendly biological sequence alignment editor and analysis program for windows 95/98/NT. Nucleic Acids Symposium, 41, 94–98.
Hunter, G. C., Wingfield, B. D., Crous, P. W., & Wingfield, M. J. (2006). A multi-gene phylogeny for species of Mycosphaerella occurring on Eucalyptus leaves. Studies in Mycology, 55, 147–161. https://doi.org/10.3114/sim.55.1.147
Ivanová, H. (2016). Comparison of the fungi Pestalotiopsis funerea (Desm.) Steyaert and Truncatella hartigii (Tubeuf) Steyaert isolated from some species of the genus Pinus L. in morphological characteristics of conidia and appendages. Journal of Forest Science, 62(6), 279–284.
Jansons, A., Zeltinš, P., Donis, J., & Neimane, U. (2020). Long-term effect of lophodermium needle cast on the growth of scots pine and implications for financial outcomes. Forests, 11, 1–12. https://doi.org/10.3390/f11070718
Jie, C., Xin, H., Xuefeng, L., & Ling, M. (2020). First report of Pestalotiopsis neglecta causing black spot needle blight of Pinus sylvestris var. mongolica in China. Plant Disease, 104(5), 1545–1545.
Juzwik, J., & Service, U. F., Central, N., Experiment, F., Avenue, F., & Paul, S. (1993). Morphology, cultural characteristics, and pathogenicity of Rhizosphaera kalkhoffii on Picea spp. in Northern Minnesota and Wisconsin. Plant Disease, 77(6), 630–634.
Korhonen, K., & Stahl, G. (2020). Maintenance and appropriate enhancement of forest resources and their contribution to global carbon cycles. In State of Europe’s Forests 2020.
Kowalski, T. (1993). Fungi in living symptomless needles of Pinus sylvestris with respect to some observed disease processes. Journal of Phytopathology, 145, 129–146. https://doi.org/10.1111/j.1439-0434.1993.tb01409.x
Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura, K. (2018). MEGA X: Molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 35(6), 1547–1549. https://doi.org/10.1093/molbev/msy096
Kumi, J., & Lang, K. J. (1979). The susceptibility of various spruce species to Rhizosphaera kalkhoffii and some cultural characteristics of the fungus in vitro. Journal of Forest Pathology, 9, 35–46.
Lamichhane, J. R., & Venturi, V. (2015). Synergisms between microbial pathogens in plant disease complexes: A growing trend. Frontiers in Plant Science, 6, 1–12. https://doi.org/10.3389/fpls.2015.00385
Lawrence, D. P., Gannibal, P. B., Dugan, F. M., & Pryor, B. M. (2014). Characterization of Alternaria isolates from the infectoria species-group and a new taxon from Arrhenatherum, Pseudoalternaria arrhenatheria sp. nov. Mycological Progress, 13, 257–276. https://doi.org/10.1007/s11557-013-0910-x
Lawrey, J., & Diederich, P. (2003). Lichenicolous Fungi: Interactions, Evolution, and Biodiversity. The Bryologist, 106(1), 80–120.
Lawrey, J. D., Diederich, P., Nelsen, M. P., Freebury, C., Van den Broek, D., Sikaroodi, M., & Ertz, D. (2012). Phylogenetic placement of lichenicolous Phoma species in the Phaeosphaeriaceae (Pleosporales, Dothideomycetes). Fungal Diversity, 55, 195–213. https://doi.org/10.1007/s13225-012-0166-9
Liu, F., Bonthond, G., Groenewald, J. Z., Cai, L., & Crous, P. W. (2019). Sporocadaceae, a family of coelomycetous fungi with appendage-bearing. Studies in Mycology, 92, 287–415. https://doi.org/10.1016/j.simyco.2018.11.001
Lopes, A., Phillips, A. J. L., & Alves, A. (2017). Mating type genes in the genus Neofusicoccum: Mating strategies and usefulness in species delimitation. Fungal Biology, 121, 394–404. https://doi.org/10.1016/j.funbio.2016.08.011
Madar, Z., Solel, Z., & Kimchi, M. (1991). Pestalotiopsis Canker of Cypress in Israel. Phytoparasitica, 19(1), 79–81.
Magnani, R. F., Rodrigues-Fo, E., & Daolio, C. (2003). Three highly oxygenated caryophyllene sesquiterpenes from Pestalotiopsis sp., a fungus isolated from Bark of Pinus taeda. Zeitschrift Für Naturforschung, 58, 319–324.
Maharachchikumbura, S. S. N., Guo, L.-D., Cai, L., Chukeatirote, E., Wu, W. P., Sun, X., Crous, P. W., Bhat, D. J., McKenzie, E. H. C., Bahkali, A. H., & Hyde, K. D. (2012). A multi-locus backbone tree for Pestalotiopsis, with a polyphasic characterization of 14 new species. Fungal Diversity, 56, 95–129. https://doi.org/10.1007/s13225-012-0198-1
Maharachchikumbura, S. S. N., Hyde, K. D., Groenewald, J. Z., Xu, J., & Crous, P. W. (2014). Pestalotiopsis revisited. Studies in Mycology, 79, 121–186. https://doi.org/10.1016/j.simyco.2014.09.005
Martínez, A. T., & Ramírez, C. (1983). Rhizosphaera oudemansii (Sphaeropsidales) associated with a needle cast of Spanish Abies pinsapo. Mycopathologia, 83, 175–182.
Martínez-Álvarez, P., Martín-García, J., Rodríguez-Ceinós, S., & Diez, J. J. (2012). Monitoring endophyte populations in pine plantations and native oak forests in Northern Spain. Forest Systems, 21, 373. https://doi.org/10.5424/fs/2012213-02254
Martínez-Álvarez, P., Fernández-González, R. A., Sanz-Ros, A. V., Pando, V., & Diez, J. J. (2016). Two fungal endophytes reduce the severity of pitch canker disease in Pinus radiata seedlings. Biological Control, 94, 1–10. https://doi.org/10.1016/j.biocontrol.2015.11.011
Mercader, G. M., Flores, S. Z., Vargas, G. G., & Von Stowasser, E. S. (2006). Screening to antagonistic fungi for Botrytis cinerea biocontrol in Chilean forest nurseries. Bosque, 27(2), 126–134. https://doi.org/10.4067/s0717-92002006000200007
Mesanza, N., Raposo, R., Elvira-Recuenco, M., Hernández-Escribano, L., Barnes, I., van der Nest, A., Pascual, M. T., Barrena, I., Martín, U. S., Cantero, A., & Iturritxa, E. (2021). New Hosts for Lecanosticta acicola and Dothistroma septosporum in Spain. Forest Pathology, 51, 1–6. https://doi.org/10.20944/PREPRINTS201912.0031.V1
Mittal, R. K., Singh, P., & Wang, B. S. P. (1987). Botrytis: A hazard to reforestation. Forest Pathology, 17, 369–384. https://doi.org/10.1111/j.1439-0329.1987.tb01330.x
Möller, E. M., Bahnweg, G., Sandermann, H., & Geiger, H. H. (1992). A simple and efficient protocol for isolation of high molecular weight DNA from filamentous fungi, fruit bodies, and infected plant tissues. Nucleic Acids Research, 20(22), 6115–6116. https://doi.org/10.1093/nar/20.22.6115
Morales-Rodríguez, C., Matteo, D. V., Aleandri, M., & Vannini, A. (2019). Pestalotiopsis biciliata, a new leaf pathogen of Eucalyptus spp. recorded in Italy. Forest Pathology, 1–7. https://doi.org/10.1111/efp.12492
Mullett, M., & Barnes, I. (2012). Dothistroma Isolation and molecular identification methods. In COST ACTION FP1102 determining invasiveness and risk of dothistroma (Issue May). https://doi.org/10.1007/978-1-4020-4585-1_266
Mullett, M. S., Adamson, K., Bragança, H., Bulgakov, T. S., Georgieva, M., Henriques, J., Jürisoo, L., Laas, M., & Drenkhan, R. (2018). New country and regional records of the pine needle blight pathogens Lecanosticta acicola, Dothistroma septosporum and Dothistroma pini. Forest Pathology, 48(5), 1–10. https://doi.org/10.1111/efp.12440
Muñoz-Adalia, E. J., Sanz-Ros, A. V., Flores-Pacheco, J. A., Hantula, J., Diez, J. J., Vainio, E. J., & Fernández, M. (2017). Sydowia polyspora dominates fungal communities carried by two Tomicus species in pine plantations threatened by Fusarium circinatum. Forests, 8, 1–16. https://doi.org/10.3390/f8040127
Ortíz de Urbina, E., Mesanza, N., Aragonés, A., Raposo, R., Elvira-Recuenco, M., Boqué, R., Patten, C., Aitken, J., & Iturritxa, E. (2016). Emerging Needle Blight Diseases in Atlantic Pinus Ecosystems of Spain. Forests, 8(1), 18. https://doi.org/10.3390/f8010018
Phookamsak, R., Liu, J.-K., Mckenzie, E. H. C., Manamgoda, D. S., Ariyawansa, H., Thambugala, K. M., Dai, D.-Q., Camporesi, E., Chukeatirote, E., Wijayawardene, N. N., Bahkali, A. H., Mortimer, P. E., Xu, J.-C., & Hyde, K. D. (2014). Revision of Phaeosphaeriaceae. Fungal Diversity, 68, 159–238. https://doi.org/10.1007/s13225-014-0308-3
Qi, M., Xie, C.-X., Chen, Q.-W., & Yu, Z.-D. (2021). Pestalotiopsis trachicarpicola, a novel pathogen causes twig blight of Pinus bungeana (Pinaceae : Pinoideae) in China. Antonie Van Leeuwenhoek, 114, 1–9. https://doi.org/10.1007/s10482-020-01500-8
Ronquist, F., & Huelsenbeck, J. P. (2003). MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics, 19(12), 1572–1574. https://doi.org/10.1093/bioinformatics/btg180
Schneider, S., Jung, E., Queloz, V., Meyer, J. B., & Rigling, D. (2019). Detection of pine needle diseases caused by Dothistroma septosporum, Dothistroma pini and Lecanosticta acicola using different methodologies. Forest Pathology, 49(2), 1–9. https://doi.org/10.1111/efp.12495
Schubert, K., Greslebin, A., Groenewald, J. Z., & Crous, P. W. (2009). New foliicolous species of Cladosporium from South America. Persoonia, 22, 111–122. https://doi.org/10.3767/003158509X449381
Silva, A. C., Diogo, E., Henriques, J., Ramos, A. P., Sandoval-Denis, M., Crous, P. W., & Bragança, H. (2020). Pestalotiopsis pini sp. nov., an Emerging Pathogen on Stone Pine (Pinus pinea L.). Forests, 11, 1–17.
Skilling, D. D., & Walla, J. A. (1986). Rhizosphaera needle cast of spruce. In J. W. Riffle (Ed.), Diseases of trees in the great plains (pp. 124–127).
Summerell, B. A. (2019). Resolving Fusarium: Current status of the genus. Annual Review of Phytopathology, 57, 323–339. https://doi.org/10.1146/annurev-phyto-082718-100204
Swofford, D. L. (1993). PAUP: Phylogenetic Analysis Using Parsimony. Mac Version 3. 1. 1. (Computer Program and Manual).
Syme, P. (2018). Werner’s nomenclature of colours: Adapted to zoology, botany, chemistry, mineralogy, anatomy, and the arts. Smithsonian Institution.
Thompson, J. D., Gibson, T. J., Plewniak, F., Jeanmougin, F., & Higgins, D. G. (1997). The CLUSTAL_X windows interface: Flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research, 25(24), 4876–4882.
Trakunyingcharoen, T., Lombard, L., Groenewald, J. Z., Cheewangkoon, R., Toanun, C., Alfenas, A. C., & Crous, P. W. (2014). Mycoparasitic species of Sphaerellopsis, and allied lichenicolous and other genera. International Mycological Association, 5(2), 391–414. https://doi.org/10.5598/imafungus.2014.05.02.05
Trivedi, P., Leach, J. E., Tringe, S. G., Sa, T., & Singh, B. K. (2020). Plant–microbiome interactions: From community assembly to plant health. Nature Reviews Microbiology, 18, 607–621. https://doi.org/10.1038/s41579-020-0412-1
Valenzuela-Lopez, N., Sutton, D. A., Cano-Lira, J. F., Paredes, K., Wiederhold, N., Guarro, J., & Stchigel, A. M. (2017). Coelomycetous fungi in the clinical setting: morphological convergence and cryptic diversity. Journal of Clinical Microbioly, 55, 552–567.
Xu, J., Ebada, S. S., & Proksch, P. (2010). Pestalotiopsis a highly creative genus: Chemistry and bioactivity of secondary metabolites. Fungal Diversity, 44, 15–31. https://doi.org/10.1007/s13225-010-0055-z
Zamora, P., Martínez-Ruiz, C., & Diez, J. J. (2008). Fungi in needles and twigs of pine plantations from northern Spain. Fungal Diversity, 30, 171–184.
Acknowledgements
Thanks are due to the Portuguese Foundation for Science and Technology (FCT/MCTES) for the financial support to CESAM (UIDP/50017/2020 + UIDB/50017/2020) and the PhD grants of Pedro Monteiro (SFRH/BD/143879/2019) and Micael Gonçalves (SFRH/BD/129020/2017). This study was also made possible through project PID2019-110459RB-I00 funded by MICINN (Spain) and FEDER (EU) budget as well as project VA208P20 funded by JCYL (Spain).
All principles of ethical and professional conduct have been followed during this research and elaboration of this manuscript.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Research involving human participants and/or animals
Not applicable.
Informed consent
All authors have reviewed the manuscript and approved its submission to the European Journal of Plant Pathology.
Conflict of interest
The authors declare that they have no conflict of interest.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Monteiro, P., Gonçalves, M.F.M., Pinto, G. et al. Three novel species of fungi associated with pine species showing needle blight-like disease symptoms. Eur J Plant Pathol 162, 183–202 (2022). https://doi.org/10.1007/s10658-021-02395-5
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10658-021-02395-5