Abstract
The complex of Diaporthe (asexual morph) species occurring on soybean constitutes an important pathogenic group associated with diseases such as pod and stem blight, seed decay and stem canker. Stem canker, caused by Diaporthe aspalathi, has been reported as the most aggressive form of canker and its occurrence has limited soybean crop productivity in the southern United States. The main form of pathogen control is the use of stem canker resistant soybean varieties. In this study, strains of Diaporthe and Phomopsis were isolated from stem and seeds of soybean in different locations in South America during the years 1989–2014. Genomic DNA from 26 isolates were analyzed by PCR-restriction fragment length polymorphism (RFLP) and Amplified Fragment Length Polymorphism (AFLP) techniques, and sequencing of internal transcribed spacer (ITS) regions of ribosomal DNA. The molecular analysis of ITS sequences by alignment with those of ex-type strains deposited in GenBank and morphological characteristics allowed the identification of Phomopsis longicolla, D. phaseolorum var. sojae, D. caulivora and D. aspalathi. An analysis of the pathogenicity of 13 isolates of D. aspalathi inoculated in soybean genotypes carrying different resistance genes to stem canker (Rdm1, Rdm2, Rdm3, Rdm4, Rdm5 and Rdm?) enabled us to identify the occurrence of at least three races of D. aspalathi occurring in Brazil. Among the isolates identified as D. aspalathi, both molecular and phenotypic analyses showed clustering depending on the date of collection and pathogenicity, which revealed the existence of variability of the pathogen.
Similar content being viewed by others
References
Alexopoulos, C. J., Mims, C. W., & Blackwell, M. (1996). Introductory mycology. New York: John Wiley & Sons.
Almeida, A. M. R., & Seixas, C. D. S. (2010). Soja: Doenças radiculares e de hastes e inter-relações com o manejo do solo e da cultura. Londrina: Embrapa – Soja.
Athow, K. L., & Caldwel, R. M. (1954). A comparative study of Diaporthe stem canker and stem blight of soybeans. Phytopatology, 44, 319–325.
Backes, R. L., Reis, M. S., Sediyama, T., Cruz, C. D., & Backes, F. A. A. L. (2005). Reaction of soybean lines to stem canker. Crop Breeding and Applied Biotechnology, 5, 223–228.
Backman, P. A., Weaver, D. B., & Morgan-Jones, G. (1985). Soybean stem canker: An emerging disease problem. Plant Disease, 69, 641–647.
Barbieri, R. L., & Carvalho, F. (2001). Coevolução de plantas e fungos patogênicos. Revista Brasileira de Agrociência, 7, 79–83.
Bowers, G. R., Ngeleka, K., & Smith, O. D. (1993). Inheritance of stem canker resistance in soybean cultivars Crockett and Dowling. Crop Science, 33, 67–70.
Cheng, Z., Tang, W., Xu, S., Sun, S., Huang, B. Y., Yan, X., Chen, Q., & Lin, Y. (2008). First report of an endophyte (Diaporthe phaseolorum var. sojae) from Kandelia candel. Journal of Forestry Research, 19, 277–282.
Chiesa, M. A., Pioli, R. N., & Morandi, E. N. (2009). Specific resistance to soybean stem canker conferred by the Rdm4 locus. Plant Pathology, 58, 1032–1038.
Coelho, A. S. G. (2001) Software: Dboot—Avaliação de dendrogramas baseados em estimativas de distâncias/similaridades genéticas através do procedimento de bootstrap, versão 3.0. Universidade Federal de Goiás, Goiânia.
Costamilan, L. M., Yorinori, J. T., Almeida, A. M. R., Seixas, C. D. S., Binneck, E., Araújo, M. R., & Carbonari, J. A. (2008). First report of Diaporthe phaseolorum var. caulivora infecting soybean plants in Brazil. Tropical Plant Pathology, 33, 381–385.
Diogo, E. L. F., Santos, J. M., & Phillips, A. J. L. (2010). Phylogeny, morphology and pathogenicity of Diaporthe and Phomopsis species on almond in Portugal. Fungal Diversity, 44, 107–115.
Doyle, J. J., & Doyle, J. L. (1987). A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin, 19, 11–15.
Felsenstein, J. (1985). Phylogenies and comparative method. The American Naturalist, 125, 1–15.
Fernández, F. A., & Hanlin, R. T. (1996). Morphological and RAPD analyses of Diaporthe phaseolorum from soybean. Mycologia, 88, 425–440.
Fernández, F. A., Phillips, D. V., Russin, J. S. & Rupe, J. C. (1999). Stem canker. In G. L. Hartman, J. B. Sinclair, & J. C. Rupe (Eds.), Compendium of soybean diseases, (4th ed, pp. 33–35). St. Paul: American Phytopathological Society.
Figueiredo, M. B., & Pimentel, P. V. C. (1975). Métodos utilizados para conservação de fungos na Micoteca da Seção de Micologia Fitopatológica do Instituto Biológico. Summa Phytopatologica, 1, 299–302.
Hillen, T., Juliatti, F. C, Polizel, A. C., Hamawaki, O. T., Brito, C. H. (2006). Reação de genótipos de soja quanto a resistência ao cancro da haste. Bioscience Journal, 22, 27–35.
Hobbs, T. W., Schmitthenner, A. F., & Kuter, G. A. A. (1985). New Phomopsis species from soybean. Mycologia, 77, 535–544.
Keeling, B. L. (1982). A seedling test for resistance to soybean stem canker caused by Diaporthe phaseolorum var. caulivora. Phytopathology, 72, 807–809.
Keeling, B. L. (1988). Influence of temperature on growth and pathogenicity of geographical isolates Diaporthe phaseolorum var. caulivora. Plant Diseases, 72, 220–222.
Kilen, T. C., Keeling, B. L., & Hartwig, E. E. (1985). Inheritance of reaction to stem canker in soybean. Crop Science, 25, 50–51.
Kulik, M. M. (1984). Failure of Phomopsis phaseoli to produce mature pycnidia in senescent soybean stems at the end of the growing season. Mycologia, 76, 863–867.
Lehman, S. G. (1923). Pod and stem blight of soybean. Annals of the Missouri Botanical Garden, 10, 111–178.
Lu, X., Robertson, A. E., Byamukama, E. Z., & Nutter, F. W. (2010). Evaluating the importance of stem canker of soybean in Iowa. Plant Disease, 94, 167–173.
Luttrell, E. S. (1947). Diaporthe phaseolorum var. sojae on crop plants. Phytopathology, 37, 445–465.
Mahmodi, F., Kadir, J. B., Puteh, A., Wong, M. Y., & Nasehi, A. (2013). First report of pod and stem blight of lima bean caused by Diaporthe phaseolorum var. sojae in Malaysia. Plant Disease, 97, 287.
Morgan-Jones, G (1989) The Diaporthe/Phomopsis complex: Taxonomic considerations. Proceedings. Buenos Aires, Associatión Argentina de la Soya.
Naruzawa, E. S., Dalla Vale, R. K., Silva, C. M., & Camargo, L. E. A. (2011). Genetic diversity of Podosphaera xanthii assessed by AFLP markers and ITS sequences. Summa Phytopathologica, 37, 94.
Nozaki, M. H., Holanda, M., Andrade, A. P., & Barreto, M. (2006). Caracterização molecular através da técnica fAFLP de isolados de Diaporthe citri. Summa Phytopathologica, 32, 147–150.
Pioli, R. N., Morandi, E., Martinez, M., Tozzini, A., Bisaro, V., & Hopp, E. (2003). Morphologic, molecular and pathogenic caracterizacion of Diaporthe phaseolorum variability in the core soybean producing area of Argentina. Phytopatology, 99, 136–146.
Querales, P. J. (2010) Caracterização morfológica e genética de Fusarium spp. isolados de sementes e associados à podridão do colmo de milho (Zea mays L.). Dissertation, Escola Superior de Agricultura Luiz de Queiroz, Piracicaba.
Reck, M., Benício, L. M., Ruas, E. A., Rodrigues, L. A., Ruas, P. M., Ortiz, M. A., Talavera, S., Urtubey, E., Stuessy, T., & Weiss-Schneeweiss, H. (2011). Karyotype and AFLP data reveal the phylogenetic position of the Brazilian endemic Hypochaeris catharinensis (Asteraceae). Plant Systematics and Evolution, 296, 231–243.
Reis, G. M. (2009). Variabilidade Genética de cepas de Aspergillus flavus isoladas de amendoim. Dissertation, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo.
Rensburg, J. C. J. V., Lamprecht, S. C. L., Groenewald, J. Z., Lisa, A., Castlebury, L. A., & Crous, P. W. (2006). Characterisation of Phomopsis spp. associated with die-back of rooibos (Aspalathus linearis) in South Africa. Studies in micology, 55, 65–74.
Rohlf, F. J. (1993) NTSYS – pc: Em unumerical taxonomy and multivariate system. Version 2.9. New York. Applied Biostatistics.
Roy, K. W. (1997). Colonization of weeds by Phomopsis longicolla. Plant Pathology, 19, 193–196.
Sanogo, S., & Etarock, B. F. (2009). First report of Phomopsis longicolla causing stem blight of Valencia peanut in New Mexico. Plant Disease, 93, 965.
Santos, J. M., Vrandecic, K., Cosic, J., Duvnjak, T., & Phillips, A. J. L. (2011). Resolving the Diaporthe species occurring on soybean in Croatia. Persoonia, 27, 9–19.
Says-Lesage, V., Roeckel-Drevet, P., Viguié, A., Tourvielle, J., Nicolas, P., & Tourvieille, D. (2002). Molecular variability within Diaporthe/Phomopsis helianthi from France. Phytopathology, 92, 308–313.
Shearin, Z. P. (2011) Facilitanting breeding for resistance to sourthern stem canker and elevated seed oleic acid content in soybean. Dissertation, University da Georgia, Athenas.
Shimomoto, Y., Sato, T., Hojo, H., Morita, Y., Takeuchi, S., Mizumoto, H., Kiba, A., & Hikichi, Y. (2011). Pathogenic and genetic variation among isolates of Corynespora cassiicola in Japan. Plant Pathology, 60, 253–260.
Shu, C., Chen, J., Huang, H., He, Y., & Zhou, E. (2014). First report of Phomopsis longicolla causing stem canker of eggplant in Guangdong, China. Plant Disease, 98, 426.
Soldera, M. C. (2010) A. Diversidade genética de isolados do complexo Diaporthe/ Phomopsis em soja no Brasil. Dissertation, Universidade Estadual de Maringá.
Sun, S., Van, K., Kim, M. Y., Min, K. H., Lee, Y. M., & Lee, S. H. (2012). Diaporthe phaseolorum var. caulivora, a causal agent for both stem canker and seed decay on soybean. Plant Pathology, 28, 55–59.
Tyler, J. M. (1996). Characterization of stem canker resistance in “Hutcheson” soybean. Crop Science, 36, 591–593.
Udayanga, D., Liu, X., Crous, P. W., Mckenzie, E. H. C., Chukeatirote, E., & Hyde, K. D. (2012). A multi-locus phylogenetic evaluation of Diaporthe (Phomopsis). Fungal Diversity, 56, 157–171.
Udayanga, D., Castlebury, L. A., Rossman, A. Y., Chukeatirote, E., & Hyde, K. D. (2014). Insights into the genus Diaporthe: Phylogenetic species delimitation in the D. eres species complex. Fungal Diversity, 67, 203–229.
Vos, R., Hogers, R., Bleeker, M., Reijans, M., Van de Lee, T., Homes, M., Frijters, A., Pot, J., Peleman, J., & Kuiper, M. (1995). AFLP: A new concept for DNA fingerprinting. Nucleic Acids Research, 23, 4407–4414.
Vrandecic, K., Cosic, J., Riccioni, L., Duvnjak, T., & Jurkovic, D. (2005). Isolation of Diaporthe phaseolorum var. caulivora from Abutilon theophrasti in Croatia. Plant Pathology, 54, 576.
Welch, A. W., & Gilman, J. C. (1948). Hetero- and homo-thallic types of Diaporthe on soybeans. Phytopathology, 38, 628–637.
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). New York: Academic Press.
Wysmierski, P. T., Vello, N. A. (2013). The genetic base of Brazilian soybean cultivars: evolution over time and breeding implications. Genetics and Molecular Biology, 4, 547–555.
Yorinori, J. T. (1991). Metodologia para avaliação da resistência ao cancro da haste da soja. Fitopatologia Brasileira, 16, 58.
Yorinori, J. T. (1996) Cancro da haste da soja: Epidemiologia e controle. Embrapa Soja. Circular Técnica, 14.
Zhang, A. W., Riccioni, L., Pedersen, W. L., Kollipara, K. P., & Hartman, G. L. (1998). Molecular identification and phylogenetic grouping of Diaporthe phaseolorum and Phomopsis longicolla isolates from soybean. Phytopathology, 88, 1306–1314.
Acknowledgements
We thank the support Guilherme Goulart Filho, Luis Cajiolla (Laboratório de Fitopatologia da Embrapa Soja) in the phenotypic evaluations, Robson Emidio de Sousa (Banco Ativo de Germoplasma) in planting, Sergio Fernando Haas Herculano (Laboratório de Microbiologia da Embrapa Soja) in microscopy analysis, Adair Vicente Carneiro in the pictures, Marcia Kamogae Kuwahara, Silvana Regina Rockenbach Marin, Ana Maria Avelino, Adriana Santos for molecular techniques (Laboratório de Biotecnologia Vegetal e Seleção Assistida por Marcadores Moleculares da Embrapa Soja), and all colleagues from the Plant Biotechnology Laboratory of EMBRAPA Soybean. This worked was financed by CAPES, CNPq and EMBRAPA (Govt. of Brazil).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Rights and permissions
About this article
Cite this article
Brumer, B.B., Lopes-Caitar, V.S., Chicowski, A.S. et al. Morphological and molecular characterization of Diaporthe (anamorph Phomopsis) complex and pathogenicity of Diaporthe aspalathi isolates causing stem canker in soybean. Eur J Plant Pathol 151, 1009–1025 (2018). https://doi.org/10.1007/s10658-018-1436-5
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10658-018-1436-5