Métodos moleculares para la detección específica de Colletotrichum sansevieriae
DOI:
https://doi.org/10.15517/am.2025.62815Palabras clave:
RFLP, antractnosis, enfermedad vegetal, gen β-tubulina, pátogeno vegetal, diagnóstico fúngicoResumen
Introduction. Sansevieria anthracnose, caused by Colletotrichum sansevieriae, poses a significant risk to the cultivation and export of this ornamental plant. Effective and swift identification methods for this pathogen are crucial for implementing control measures to prevent its spread to uninfected areas. Objective. To implement and optimize molecular methods for the rapid and reliable identification of C. sansevieriae. Materials and methods. During 2016, a β-tubulin-2 (β-tub2) gene fragment of C. sansevieria isolated from a local farm in Alajuela, Costa Rica was analyzed. PCR-RFLP of the partial β-tubulin-2 (β-tub2) gene fragment was implemented with the enzyme MseI (Tru1I). In addition, species-specific primers for C. sansevieriae detection and PCR-RFLP analysis of the amplified fragment were applied. Results. The digestion consistently produced a two-band restriction pattern specific to C. sansevieriae. The designed primers successfully amplified a 383 bp fragment of the β-tub2 from all C. sansevieriae strains tested. No amplification was observed from other Colletotrichum species within the C. gloesporioides and C. acutatum complexes, as well as from C. truncatum and Fusarium oxysporum isolates. Moreover, this restriction site, located within the amplicon generated by the species-specific primers for C. sansevieriae, enabled successful validation of the species through digestion. Conclusions. Both PCR based methods proved sensitive enough to detect C. sansevieriae in naturally and artificially infected Sansevieria leaves without requiring isolation of the pathogen in pure cultures, making the diagnostic process more efficient and accessible.
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Afanador-Kafuri, L., González, A., Gañán, L., Mejía, J. F., Cardona, N., & Álvarez, E. (2014). Characterization of the Colletotrichum species causing anthracnose in Andean blackberry in Colombia. Plant Disease, 98(11), 1503-1513. https://doi.org/10.1094/PDIS-07-13-0752-RE
Aldaoud, R., DeAlwis, S., Salib, S., Cunnington, J. H., & Doughty, S. (2011). First record of Colletotrichum sansevieriae on Sansevieria sp. (mother-in-law’s tongue) in Australia. Australasian Plant Disease Notes, 6(1), 60-61. https://doi.org/10.1007/s13314-011-0020-z
Álvarez, E., Gañán, L., Rojas-Triviño, A., Mejía, J. F., Llano, G. A., & González, A. (2014). Diversity and pathogenicity of Colletotrichum species isolated from soursop in Colombia. European Journal of Plant Pathology, 139, 325-338. https://doi.org/10.1007/s10658-014-0388-7
Bhunjun, C. S., Phukhamsakda, C., Jayawardena, R. S., Jeewon, R., Promputtha, I., & Hyde, K. D. (2021). Investigating species boundaries in Colletotrichum. Fungal Diversity, 107(1), 107-127. https://doi.org/10.1007/s13225-021-00471-z
Brand, T., & Wichura, A. (2023). First report on Colletotrichum sansevieriae causing anthracnose of Sansevieria trifasciata in Germany. Gesunde Pflanzen, 75(1), 61-66. https://doi.org/10.1007/s10343-022-00777-1
Campoverde, E. V., & Palmateer, A. J. (2012). Colletotrichum sansevieriae causing anthracnose of Sansevieria trifasciata ‘Laurentii’ and ‘Moonshine’ in South Florida. Proceedings of the Florida State Horticultural Society, 125, 359-360. https://journals.flvc.org/fshs/issue/view/4046
Cao, L., Shi, K., Liu, Y., Xie, X., Sun, X., Dong, W., & Ma, L. (2024). Identification of specific genes as molecular markers for rapid and accurate detection of oil-tea Camellia anthracnose pathogen Colletotrichum fructicola in China. Frontiers in Microbiology, 15, Article 1442922. https://doi.org/10.3389/fmicb.2024.1442922
Chung, W. H., Chung, W. C., Peng, M. T., Yang, H. R., & Huang, J. W. (2010). Specific detection of benzimidazole resistance in Colletotrichum gloeosporioides from fruit crops by PCR-RFLP. Nature Biotechnology, 27(1), 17-24. https://doi.org/10.1016/j.nbt.2009.10.004
Damm, U., Cannon, P. F., Woudenberg, J. H. C., & Crous, P. W. (2012). The Colletotrichum acutatum species complex. Studies in Mycology, 73, 37-114. https://doi.org/10.3114/sim0010
Dela Cruz, M., Christensen, J. H., Thomsen, J. D., & Müller, R. (2014). Can ornamental potted plants remove volatile organic compounds from indoor air? — A review. Environmental Science and Pollution Research, 21(24), 13909-13928. https://doi.org/10.1007/s11356-014-3240-x
Doyle, J. J., & Doyle, J. L. (1987). A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin 19, 11–15.
Eaton, M., Edwards, S., Inocencio, H., Machado, F., Nuckles, E., Farman, M., Gauthier, N., & Vaillancourt, L. (2021). Diversity and cross-infection potential of Colletotrichum causing fruit rots in mixed-fruit orchards in Kentucky. Plant Disease, 105(4), 115-1128. https://doi.org/10.1094/PDIS-06-20-1273-RE
Ferrucho, R. L., Marín-Ramírez, G. A., Ochoa-Corona, F., & Ángel C., C. A. (2024). PCR-based detection for the quarantine fungus Colletotrichum kahawae, a biosecurity threat to the coffee (Coffea arabica) industry worldwide. Plant Disease, 108(9), 2615-2624. https://doi.org/10.1094/PDIS-09-23-1788-SR
Gang, G.-H., Cho, H. J., Kim, H. S., Kwack, Y.-B., & Kwak, Y.-S. (2015). Analysis of fungicide sensitivity and genetic diversity among Colletotrichum species in sweet persimmon. Plant Pathology Journal, 31(2), 115-122. https://doi.org/10.5423/PPJ.OA.03.2015.0033
Gautam, A. K., Avasthi, S., & Bhadauria, R. (2012). Colletotrichum sansevieriae on Sansevieria trifasciata — A report from Madhya Pradesh, India. Plant Pathology & Quarantine, 2(2), 190-192. https://plantpathologyquarantine.org/volume-2/issue-2.html
Glass, N. L., & Donaldson, G. C. (1995). Development of primer sets designed for use with the PCR to amplify conserved genes from filamentous ascomycetes. Applied and Environmental Microbiology, 61, 1323-1330. https://doi.org/10.1128/aem.61.4.1323-1330.1995
Grskovich, M., Creswell, T., & Bonkowski, J. (2024). First Report of Anthracnose Caused by Colletotrichum sansevieriae on Dracaena trifasciata (Snake Plant) in Tennessee. Plant Disease, 108(12), 3650. https://doi.org/10.1094/PDIS-05-24-1049-PDN
Hall, T. A. (1999). BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41, 95-98
.
Huang, R., Sun, W., Wang, L., Li, Q., Huang, S., Tang, L., ... & Hsiang, T. (2021). Identification and characterization of Colletotrichum species associated with anthracnose disease of banana. Plant Pathology, 70(8), 1827-1837. https://doi.org/10.1111/ppa.13426
Kee, Y. J., Zakaria, L., & Mohd, M. H. (2020a). Identification, pathogenicity, and histopathology of Colletotrichum sansevieriae causing anthracnose of Sansevieria trifasciata in Malaysia. Journal of Applied Microbiology, 129, 626-636. https://doi.org/10.1111/jam.14640
Kee, Y. J., Zakaria, L., & Mohd, M. H. (2020b). Morphology, phylogeny, and pathogenicity of Fusarium species from Sansevieria trifasciata in Malaysia. Plant Pathology, 69(3), 442-454. https://doi.org/10.1111/ppa.13138
Khalumba, M. L., Mbugua, P. K., & Kung’u, J. B. (2005). Uses and conservation of some highland species of the genus Sansevieria Thunb in Kenya. African Crop Science Conference Proceedings, 7, 527-532.
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
Lee, D. H., Kim, D.-H., Jeon, Y.-A., Uhm, J. Y., & Hong, S.-B. (2007). Molecular and cultural characterization of Colletotrichum spp. causing bitter rot of apples in Korea. Plant Pathology Journal, 23(2), 37-44. https://doi.org/10.5423/PPJ.2007.23.2.037
Li, Y. L., Zhou, Z., Lu, W., & Ye, J. R. (2013). First report of a Chaetomella sp. causing a leaf spot on Sansevieria trifasciata in China. Plant Disease, 97(7), Article 992. https://doi.org/10.1094/PDIS-10-12-0993-PDN
Li, J., Deng, H., Diao, L. G., Zhang, R. Q., Li, J. Y., Liu, H. X., & Jin, J. (2023). First Report of Anthracnose Caused by Colletotrichum sansevieriae on Sansevieria trifasciata var. laurentii in China. Plant Disease, 107(2), Article 583. https://doi.org/10.1094/PDIS-03-22-0673-PDN
Ma, Z., & Michailides, T. J. (2007). Approaches for eliminating PCR inhibitors and designing PCR primers for the detection of phytopathogenic fungi. Crop Protection, 26(2), 145-161. https://doi.org/10.1016/j.cropro.2006.04.014
McHenry, D. J., & Aćimović, S. G. (2024). New species-specific real-time PCR assays for colletotrichum species causing bitter rot of apple. Microorganisms, 12(5), Article 878. https://doi.org/10.3390/microorganisms12050878
Maharaj, A., & Rampersad, S. N. (2012). Genetic differentiation of Colletotrichum gloeosporioides and C. truncatum associated with anthracnose disease of papaya (Carica papaya L.) and bell pepper (Capsicum annuum L.) based on ITS PCR-RFLP fingerprinting. Molecular Biotechnology, 50, 237-249. https://doi.org/10.1007/s12033-011-9434-2
Martinez-Culebras, P. V., Barrio, E., Garcia, M. D., & Querol, A. (2000). Identification of Colletotrichum species responsible for anthracnose of strawberry based on the internal transcribed spacers of the ribosomal region. FEMS Microbiology Letters, 189, 97-101. https://doi.org/10.1111/j.1574-6968.2000.tb09213.x
Martinez-Culebras, P. V., Querol, A., Suarez-Fernandez, M. B., Garcia-Lopez, M. D., & Barrio, E. (2003). Phylogenetic relationships among Colletotrichum pathogens of strawberry and design of PCR primers for their identification. Journal of Phytopathology, 151, 135-143. https://doi.org/10.1046/j.1439-0434.2003.00694.x
Nakamura, M., Ohzono, M., Iwai, H., & Arai, K. (2006). Anthracnose of Sansevieria trifasciata caused by Colletotrichum sansevieriae sp. nov. Journal of General Plant Pathology, 72(4), 253-256. https://doi.org/10.1007/s10327-006-0280-1
O’Donnell, K., & Cigelnik, E. (1997). Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus Fusarium are nonorthologous. Molecular Phylogenetics and Evolution, 7(1), 103-116. https://doi.org/10.1006/mpev.1996.0376
O’Donnell, K., Kistler, H. C., Cigelnik, E., & Ploetz, R. C. (1998). Multiple evolutionary origins of the fungus causing Panama disease of banana: Concordant evidence from nuclear and mitochondrial gene genealogies. Proceedings of the National Academy of Sciences of the United States of America, 95(5), 2044-2049. https://doi.org/10.1073/pnas.95.5.2044
Palmateer, A. J., Tarnowski, T. L. B., & Lopez, P. (2012). First report of Colletotrichum sansevieriae causing anthracnose of Sansevieria trifasciata in Florida. Plant Disease, 96(2), Article 293. https://doi.org/10.1094/PDIS-06-11-0539
Park, J. H., Han, K. S., Kim, J. Y., & Shin, H.-D. (2013). First report of anthracnose caused by Colletotrichum sansevieriae on Sansevieria in Korea. Plant Disease, 97(11), Article 1510. https://doi.org/10.1094/PDIS-04-13-0402-PDN
Pérez-León, G., Castillo-Matamoros, R., Chavarría-Pérez, L., Brenes-Angulo, A., & Gómez-Alpízar, L. (2015). Combate químico de la antracnosis de Sansevieria Trifasciata Var. Hahnii en un sistema de hojas separadas. Agronomía Mesoamericana, 26(2), 305-313. http://dx.doi.org/10.15517/am.v26i2.19323
Pérez-León, G., Quesada, L., & Gómez-Alpízar, L. (2013). Identificación del agente causal de antracnosis enSansevieria spp. en Costa Rica. Agronomía Costarricense, 37(1), 39-50. https://doi.org/10.15517/rac.v37i1.10711
Pinzón Gutiérrez, Y. A., Bustamante, S. L., & Buitrago Hurtado, G. (2013). Diagnóstico molecular diferencial Colletotrichum gloeosporioides y Fusarium oxysporum en ñame (Dioscorea sp.). Revista Colombiana de Biotecnología, 15(1), 52-60.
Prakoso, A. B., Suryanti, S., & Widiastuti, A. (2019, April). Molecular detection of Colletotrichum spp. on postharvest commodities of horticulture in Central Java and Yogyakarta, Indonesia. AIP Conference Proceedings, 2099(1), 1-6. https://doi.org/10.1063/1.5098422
Ramdeen, S., & Rampersad, S. N. (2013). Intraspecific differentiation of Colletotrichum gloeosporioides sensu lato based on in silico multilocus PCR-RFLP fingerprinting. Molecular Biotechnology, 53(2), 170-181. https://doi.org/10.1007/s12033-012-9509-8
Ramdial, H., & Rampersad, S. N. (2015). Characterization of Colletotrichum spp. causing anthracnose of bell pepper (Capsicum annuum L.) in Trinidad. Phytoparasitica, 43, 37-49. https://doi.org/10.1007/s12600-014-0428-z
Rampersad, S. N. (2011). Molecular and phenotypic characterization of Colletotrichum species associated with anthracnose disease of papaya in Trinidad. Plant Disease, 95, 1244-1254. https://doi.org/10.1094/PDIS-02-11-0080
Rampersad, K., Ramdial, H., & Rampersad, S. N. (2016). Utility of internally transcribed spacer region of rDNA (ITS) and β-tubulin gene sequences to infer genetic diversity and migration patterns of Colletotrichum truncatum infecting Capsicum spp. Ecology and Evolution, 6(2), 593-606. https://doi.org/10.1002/ece3.1918
Rêgo, M., Lopes, A., Barros, R., Lamas, A. M., Costa, M. F., & Ferreira-Gomes, R. L. (2020). Morphological characterization and genetic diversity in ornamental specimens of the genus Sansevieria. Revista Caatinga, 33, 985-992. https://doi.org/10.1590/1983-21252020v33n413rc
Ruiz-Campos, C., Umaña-Rojas, G., & Gómez-Alpízar, L. (2017). Molecular and morphological identification of Colletotrichum species in papaya fruits (Carica papaya) ‘Pococí’ hybrid. Phytopathology, 107(7), S.4.16. https://apsjournals.apsnet.org/doi/epdf/10.1094/PHYTO-107-7-S4.7
Ruiz-Campos, C., Umaña-Rojas, G., & Gómez-Alpízar, L. (2022). Identificación multilocus de especies de Colletotrichum asociadas a la antracnosis de papaya. Agronomía Mesoamericana, 33(1), Artículo 45495. http://dx.doi.org/10.15517/am.v33i1.45495
Saghai-Maroof, M. A., Soliman, K. M., Jorgensen, R. A., & Allard, R. W. (1984). Ribosomal DNA spacer-length polymorphism in barley: Mendelian inheritance, chromosomal location and population dynamics. Proceedings of the National Academy of Sciences of the United States of America, 81(24), 8014-8019. https://doi.org/10.1073/pnas.81.24.8014
Schena, L., Mosca, S., Cacciola, S. O., Faedda, R., Sanzani, S. M., Agosteo, G. E., Sergeeva, V., & Magnano di San Lio, G. (2014). Species of the Colletotrichum gloeosporioides and C. boninense complexes associated with olive anthracnose. Plant Pathology, 63(2), 437-446. https://doi.org/10.1111/ppa.12110
Syafitri, L. M., Wibowo, A., Widiastuti, A., Subandiyah, S., & Harper, S. (2023). Molecular identification of Colletotrichum gloeosporioides causing anthracnose on shallot in Bantul, Yogyakarta, Indonesia. Biodiversitas Journal of Biological Diversity, 24(8), 4530-4534. https://doi.org/10.13057/biodiv/d240855
Silva, D. N., Talhinhas, P., Várzea, V., Cai, L., Paulo, O. S., & Batista, D. (2012). Application of the Apn1/MAT locus to improve the systematics of the Colletotrichum gloeosporioides complex: An example from coffee (Coffea spp.) hosts. Mycologia, 104, 396-409. https://doi.org/10.3852/11-145
Silva, A. O., Savi, D. C., Gomes, F. B., Gos, F. M. W. R., Silva, G. J., & Glienke, C. (2017). Identification of Colletotrichum species associated with postbloom fruit drop in Brazil through GAPDH sequencing analysis and multiplex PCR. European Journal of Plant Pathology, 147, 731-748.https://doi.org/10.1007/s10658-016-1038-z
Takawira, R., & Nordal, I. (2001). The genus Sansevieria (Family Dracaenaceae) in Zimbabwe. Acta Horticulturae, 552, 189-198. https://doi.org/10.17660/ActaHortic.2002.572.22
Takawira, R., & Stedje, B. (2011). Ethnobotanical studies in the genus Sansevieria Thunb. (Asparagaceae) in Zimbabwe. Ethnobotany Research and Applications, 9, 421-443. https://ethnobotanyjournal.org/index.php/era/article/view/657
Talhinhas, P., Sreenivasaprasad, S., Neves-Martins, J., & Oliveira, H. (2005). Molecular and phenotypic analyses reveal association of diverse Colletotrichum acutatum groups and a low level of C. gloeosporioides with olive anthracnose. Applied and Environmental Microbiology, 71(6), 2987-2998. https://doi.org/10.1128/AEM.71.6.2987-2998.2005
Tapia-Tussell, R., Quijano-Ramayo, A., Cortes-Velazquez, A., Lappe, P., Larque-Saavedra, A., & Perez-Brito, D. (2008). PCR-based detection and characterization of the fungal pathogens Colletotrichum gloeosporioides and Colletotrichum capsici causing anthracnose in papaya (Carica papaya L.) in the Yucatan Peninsula. Molecular Biotechnology, 40(3), 293-298. https://doi.org/10.1007/s12033-008-9093-0
Torres-Calzada, C., Tapia-Tussell, R., Quijano-Ramayo, A., Martin-Mex, R., Rojas-Herrera, R., Higuera-Ciapara, I., & Perez-Brito, D. (2011). A species-specific polymerase chain reaction assay for rapid and sensitive detection of Colletotrichum capsici. Molecular Biotechnology, 49, 48-55. https://doi.org/10.1007/s12033-011-9377-7
Untergasser, A., Nijveen, H., Rao, X., Bisseling, T., Geurts, R., & Leunissen, J. A. (2007). Primer3Plus, an enhanced web interface to Primer3. Nucleic Acids Research, 35(Suppl. 2), W71-W74. https://doi.org/10.1093/nar/gkm306
van Kleinwee, I., Larridon, I., Shah, T., Bauters, K., Asselman, P., Goetghebeur, P., Leliaert, F., & Veltjen, E. (2022). Plastid phylogenomics of the Sanseviera clade of Sanseviera (Asparagaceae) resolves a recent radiation. Molecular Phylogenetics and Evolution, 169, Article 107404. https://doi.org/10.1016/j.ympev.2022.107404
Watanabe, K., Ikeda, H., Sakashita, T., & Sato, T. (2016). Anthracnose of genus Mandevilla caused by Colletotrichum truncatum and C. siamense in Japan. Journal of General Plant Pathology, 82, 33-37. https://doi.org/10.1007/s10327-015-0635-6
Weir, B. S., Johnston, P. R., & Damm, U. (2012). The Colletotrichum gloeosporioides species complex. Studies in Mycology, 73, 115-180. https://doi.org/10.3114/sim0011
White, T. J., Bruns, T., Lee, S., & Taylor, J. W. (1990). Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In M. A. Innis, D. H. Gelfand, J. J. Sninsky, & T. J. White (Eds.), PCR Protocols: A Guide to Methods and Applications (pp. 315-322). Academic Press.
Yadav, A., & Yadav, K. (2025). Portable solutions for plant pathogen diagnostics: development, usage, and future potential. Frontiers in Microbiology, 16, Article 1516723. https://doi.org/10.3389/fmicb.2025.1516723
Yamagishi, N., Sato, T., Chuma, I., Ishiyama, Y., & Tosa, Y. (2016). Anthracnose of black locust caused by Colletotrichum nymphaeae (Passerini) Aa. Journal of General Plant Pathology, 82, 174-176. https://doi.org/10.1007/s10327-016-0649-8
Ye, J., Coulouris, G., Zaretskaya, I., Cutcutache, I., Rozen, S., & Madden, T. L. (2012). Primer-BLAST: A tool to design target-specific primers for polymerase chain reaction. BMC Bioinformatics, 13, Article 134. http://www.biomedcentral.com/1471-2105/13/134
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