Detection and molecular characterization of Macrophomina phaseolina isolates in sesame using species-specific markers

Abstract

Sesame (Sesamum indicum L.) is a vital oilseed crop cultivated globally, yet its production is significantly threatened by charcoal rot disease caused by Macrophomina phaseolina. This soilborne pathogen leads to severe yield losses, affecting sesame-growing regions globally. In this study, we conducted a molecular characterization of M. phaseolina isolates collected from infected sesame plants in Diyarbakır and Şanlıurfa, Türkiye. During the field survey, the characteristic symptoms such as wilting, stunted growth, and blackened stems were observed and samples were collected to isolate the targeted pathogen. Seven isolates were successfully isolated and analyzed for their morphological and microscopic features, confirming the presence of M. phaseolina. The DNA extraction and PCR amplification using species-specific primers (MpKRI/MpKFI) successfully detected the pathogen in all isolates, producing a distinct 350 bp amplicon. The results validate the reliability of molecular markers in identifying M. phaseolina with high specificity. This study highlights the importance of molecular tools for precise pathogen detection, aiding in disease management strategies. The findings contribute to a better understanding of M. phaseolina’s genetic diversity and offer valuable insights for breeding programs aimed at developing resistant sesame cultivars. Future research should focus on integrating molecular diagnostics with breeding efforts to enhance sesame's resilience against charcoal rot disease.

Keywords

• Sesamum indicum
• Macrophomina phaseolina
• molecular characterization
• fungal pathogens
• sesame breeding

Supporting Institution

This study is part of the Ph.D. thesis of Amjad Ali, the first author, accepted by the Graduate School of Sivas Science and Technology University.

Ethical Statement

There is no need to obtain permission from the ethics committee for this study.

Türlere özgü markörler kullanılarak susamda Macrophomina phaseolina izolatlarının tespiti ve moleküler karakterizasyonu

Abstract

Susam (Sesamum indicum L.), dünya genelinde yetiştirilen önemli bir yağ bitkisi olup, üretimi Macrophomina phaseolina’nın neden olduğu kömür çürüklüğü hastalığı tarafından ciddi şekilde tehdit edilmektedir. Toprak kökenli bu patojen, susam yetiştirilen bölgelerde önemli verim kayıplarına yol açmaktadır. Bu çalışmada, Diyarbakır ve Şanlıurfa, Türkiye susam üretim alanlarındaki enfekte susam bitkilerinden izole edilen M. phaseolina izolatlarının moleküler karakterizasyonu gerçekleştirilmiştir. Arazi çalışmaları sırasında solgunluk, bodur büyüme ve siyahlaşmış gövdeler gibi karakteristik semptomlar tespit edilmiştir. Yedi izolat başarıyla elde edilerek morfolojik ve mikroskobik özellikleri incelenmiş ve M. phaseolina'nın varlığı doğrulanmıştır. Türlere özgü primerler (MpKRI/MpKFI) kullanılarak gerçekleştirilen DNA ekstraksiyonu ve PCR amplifikasyonu, tüm izolatlarda patojeni başarıyla tespit etmiş ve 350 bp’lik özgün bir amplikon ürünü üretmiştir. Sonuçlar, moleküler markörlerin M. phaseolina’nın yüksek özgüllükle tanımlanmasındaki güvenilirliğini doğrulamaktadır. Bu çalışma, hastalık yönetim stratejilerine katkı sağlamak için moleküler araçların hassas patojen tespitindeki önemini vurgulamaktadır. Bulgular, M. phaseolina’nın genetik çeşitliliğinin daha iyi anlaşılmasına katkıda bulunarak, kömür çürüklüğüne dayanıklı susam çeşitlerinin geliştirilmesine yönelik ıslah programları için değerli bilgiler sunmaktadır. Gelecek araştırmalar, susamın kömür çürüklüğüne karşı direncini artırmak amacıyla moleküler tanı yöntemleri ile ıslah çalışmalarının entegrasyonuna odaklanmalıdır.

Keywords

• Sesamum indicum
• Macrophomina phaseolina
• moleküler karakterizasyon
• fungal patojenler

References

1. Adhikary, N.K., Chowdhury, M.R., Begum, T. & Mallick, R. (2019). Integrated management of stem and root rot of sesame (Sesamum indicum L.) caused by Macrophomina phaseolina (Tassi) Goid. International Journal of Current Microbiology & Applied Sciences, 8(4), 22-24.
2. Akhtar, K.P., Sarwar, G. & Arshad, H.M.I. (2011). Temperature response, pathogenicity, seed infection and mutant evaluation against Macrophomina phaseolina causing charcoal rot disease of sesame. Archives of Phytopathology and Plant Protection, 44(4), 320-330.
3. Babu, B.K., Saxena, A.K., Srivastava, A.K. & Arora, D.K. (2007). Identification and detection of Macrophomina phaseolina by using species-specific oligonucleotide primers and probe. Mycologia, 99(6), 797-803.
4. Balabaskar, P., Renganathan, P. & Sanjeevkumar, K. (2015). Occurrence of sesame root rot disease in Cuddalore district of Tamil Nadu and analysis of the variability in cultural characteristics and pathogenicity among isolates of Macrophomina phaseolina (Tassi) Goid. European Journal of Biotechnology and Bioscience, 3(8), 55-59.
5. Beas-Fernández, R., De Santiago-de, S.A., Hernández, D.S. & Mayek-Pérez, N. (2006). Characterization of Mexican and non-Mexican isolates of Macrophomina phaseolina based on morphological characteristics, pathogenicity on bean seeds and endoglucanase genes. Journal of Plant Pathology, 88, 53-60.
6. Chowdhury, S., Basu, A., Ray Chaudhuri, T. & Kundu, S. (2014). In-vitro characterization of the behaviour of Macrophomina phaseolina (Tassi) Goid at the rhizosphere and during early infection of roots of resistant and susceptible varieties of sesame. European Journal of Plant Pathology, 138, 361-375.
7. Gupta, R.N., Saharan, H.S., Rathi, A.S., Avtar, R. & Mandhania, S. (2020). Role of non-conventional chemicals in biochemical defence against charcoal rot of sesame incited by Macrophomina phaseolina (Tassi) Goid. International Journal of Current Science, 8(2), 656-660.
8. Iqbal, U. & Mukhtar, T. (2014). Morphological and pathogenic variability among Macrophomina phaseolina isolates associated with mungbean Wilczek from Pakistan. The Scientific World Journal, 1-9.

9. Khan, A.N., Shair, F., Malik, K., Hayat, Z., Khan, M.A., Hafeez, F.Y. & Hassan, M.N. (2017). Molecular identification and genetic characterization of Macrophomina phaseolina strains causing pathogenicity on sunflower and chickpea. Frontiers in Microbiology, 8, 1309.
10. Mahdy, E.E., El-Shimy, A.A., Sayed, M.A., Mahmoud, A.F. & Salah, A. (2023). Pedigree selection in improving seed yield and tolerance of sesame to Macrophomina phaseolina (Tassi) Goid. Egyptian Journal of Agronomy, 45(1), 31-44.
11. Mahmoud, S.A., El-Sharkawy, E.S. & Emam, M.A. (2024). Breeding sesame for resistance to charcoal rot caused by Macrophomina phaseolina. SVU-International Journal of Agricultural Sciences, 6(2), 18-35.
12. Nayar, N.M. & Mehra, K.L. (1970). Sesame: Its uses, botany, cytogenetics, and origin. Economic Botany, 24, 20-31.
13. Romanelli, A.M., Fu, J., Herrera, M.L. & Wickes, B.L. (2014). A universal DNA extraction and PCR amplification method for fungal rDNA sequence-based identification. Mycoses, 57, 612-622.
14. Sarkar, T.S., Biswas, P., Ghosh, S.K. & Ghosh, S. (2014). Nitric oxide production by necrotrophic pathogen Macrophomina phaseolina and the host plant in charcoal rot disease of jute: complexity of the interplay between necrotroph–host plant interactions. PLoS One, 9(9), e107348.
15. Yilmaz, M. (2022). Determination of yield and yield components of some sesame (Sesamum indicum L.) genotypes in the eastern Mediterranean transitional conditions. Ege Üniversitesi Ziraat Fakültesi Dergisi, 59, 709-715.
16. Yousef, H. (2021). Pathogenic variation and molecular characterization of Macrophomina phaseolina, the cause of sesame charcoal rot. Egyptian Journal of Phytopathology, 49(1), 151-165.