Sains Malaysiana 48(8)(2019): 1593–1600

http://dx.doi.org/10.17576/jsm-2019-4808-04

Phylogenetic Analysis of Fusarium oxysporum f. sp. cubense Associated with Fusarium Wilt of Bananas from Peninsular Malaysia

(Analisis Filogenetik Fusarium oxysporum f. sp. cubense Berkait dengan Penyakit Layu Fusarium Pisang dari Semenanjung Malaysia)

CLEMENT KIING FOOK WONG¹, DZARIFAH ZULPERI¹*, GANESAN VADAMALAI¹, NOOR BAITY SAIDI² & CHUI YAO TEH²

¹Department of Plant Protection, Faculty of Agriculture, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia

²Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, UPM Serdang, Selangor Darul Ehsan, Malaysia

Received: 3 November 2018/Accepted: 12 May 2019

ABSTRACT

Fusarium wilt of bananas caused by Fusarium oxysporum f. sp. cubense (Foc) has been known to hamper banana production. In Malaysia, most of the edible banana cultivars are susceptible to the fungal strain Foc tropical race 4 (Foc TR4). To date, molecular characterisation of Foc TR4 in this country remains elusive. Thus, this study aimed to characterise the diversity of Foc TR4 in Peninsular Malaysia based on the phylogenetic analysis of transcription elongation factor (TEF-1α) sequences. From March 2016 to December 2016, 17 isolates of Foc were isolated from nine outbreak states in Peninsular Malaysia consisting of Penang, Perak, Selangor, Negeri Sembilan, Melaka, Johor, Pahang, Kelantan and Terengganu, with disease incidence exceeding 70% in several plantations. Morphological, molecular identification using TR4 specific primers and pathogenicity assay confirmed the identity of the fungal pathogen. Pathogenicity assays indicated that the aggressiveness level varies among the isolates. Phylogenetic analyses of TEF1-α sequences showed that there was no genetic variation among the Foc isolates by clustering them into four vegetative compatibility groups (VCGs) of 01213 (TR4), 01216 (TR4), 01213/16 (TR4) and 0121 (R4). PCR amplification using VCG specific primers have further categorised 16 Foc isolates as VCG01213 and a single isolate as VCG01213/16. In addition, no correlation between TEF-1α sequences and aggressiveness of isolates could be established.

Keywords: Aggressiveness; banana; fusarium wilt; phylogenetic analysis; TEF-1α

ABSTRAK

Penyakit layu Fusarium pisang disebabkan oleh kulat Fusarium oxysporum f. sp. cubense (Foc) telah menjejaskan pengeluaran pisang. Kebanyakan kultivar pisang yang boleh dimakan di Malaysia mudah dijangkiti oleh strain kulat Foc tropical race 4 (Foc TR4). Sehingga kini, pencirian Foc TR4 di peringkat molekul dalam negara ini masih tidak dikaji. Justeru, kajian ini bertujuan untuk mencirikan kepelbagaian Foc TR4 di Semenanjung Malaysia berdasarkan analisis filogenetik jujukan faktor transkripsi pemanjangan (TEF-1α). Dari Mac 2016 hingga Disember 2016, sebanyak 17 pencilan Foc telah dipencilkan dari sembilan kawasan berpenyakit di Semenanjung Malaysia terdiri daripada Pulau Pinang, Perak, Selangor, Negeri Sembilan, Melaka, Johor, Pahang, Kelantan dan Terengganu dengan kejadian penyakit melebihi 70% di beberapa ladang pisang. Pengenalpastian morfologi dan molekul dengan menggunakan primer khusus TR4 dan ujian kepatogenan telah mengesahkan identiti patogen kulat. Ujian kepatogenan menunjukkan bahawa tahap agresif antara pencilan Foc adalah berbeza. Analisis filogenetik jujukan TEF-1α menunjukkan bahawa tiada variasi genetik antara pencilan Foc dengan dibahagikan kepada empat vegetative compatibility groups (VCGs) iaitu 01213 (TR4), 01216 (TR4), 01213/16 (TR4) dan 0121 (R4). Amplifikasi PCR dengan menggunakan primer khusus VCG telah selanjutnya mengkategorikan 16 pencilan Foc sebagai VCG01213 dan satu pencilan sebagai VCG01213/16. Tambahan pula, tiada korelasi antara jujukan TEF-1α dan tahap agresif pencilan Foc dapat ditunjukkan.

Kata kunci: Analisis filogenetik; penyakit layu fusarium; pisang; tahap agresif; TEF-1α

REFERENCES

Buddenhagen, I.W. 2009. Understanding strain diversity in Fusarium oxysporum f. sp. cubense and history of introduction of ‘Tropical Race 4’ to better manage banana production. Acta Horticulturae 828: 193-204.

Costa, S.N., Brangaςa, C.A.D., Ribeiro, L.R., Amorim, E.P., Oliveira, S.A.S., Dita, M.A., Laranjeira, F.F. & Haddad, F. 2015. Genetic structure of Fusarium oxysporum f. sp. cubense in different region of Brazil. Plant Pathology 64: 137-146.

Cunha, C.M., Hinz, R.H., Pereira, A., Tcacenco, F.A. & Stadnik, M.J. 2015. Aggressiveness and genetic diversity of Fusarium oxysporum f. sp. cubense from Santa Catarina, Southern Brazil. Tropical Plant Pathology 40: 326-334.

Dita, M.A., Waalwijk, C., Buddenhagen, I.W., Souza Jr., M.T. & Kema, G.H.J. 2010. A molecular diagnostic for tropical race 4 of the banana fusarium wilt pathogen. Plant Pathology 60(2): 348-357.

Ellis, M.L., Lanubile, A., Garcia, C. & Munvold, G.P. 2016. Association of putative fungal effectors in Fusarium oxysporum with wilt symptoms in soybean. Phytopathology 106(7): 762-773.

Fourie, G., Steenkamp, E.T., Gordon, T.R. & Vijoen, A. 2009. Evolutionary relationships among the Fusarium oxysporum f. sp. cubense vegetative compatibility groups. Applied Environmental Microbiology 75: 4770-4781.

Groenewald, S., Van der Berg, N., Marasas, W.F.O. & Viljoen, A. 2006. The application of high-throughput, AFLP in assessing genetic diversity in Fusarium oxysporum f. sp. cubense. Mycology Research 110: 297-305.

Katan, T. 1999. Current status of vegetative compatibility groups in Fusarium oxysporum. Phytoparasitica 27: 72-77.

Komada, H. 1975. Development of a selective medium for quantitative isolation of Fusarium oxysporum from natural soil. Plant Protection Research 8: 114-125.

Kumar, B.H., Shankar, U.A.C., Kini, R.K., Prakash, H.S. & Shetty, S.H. 2006. Genetic variation in Fusarium oxysporum f. sp. cubense isolates based on random amplified polymorphic DNA and intergenic spacer. Archives of Phytopathology and Plant Protection 39: 151-160.

Leong, S.K., Latiffah, Z. & Baharuddin, S. 2010. Genetic diversity of Fusarium oxysporum f. sp. cubense isolates from Malaysia. African Journal of Microbiology Research 4: 1026-1037.

Leslie, J.F. & Summerell, B.A. 2006. The Fusarium Laboratory Manual. UK: Wiley Blackwell.

Li, C.Y., Mostert, G., Zuo, C.W., Beukes, I., Yang, Q.S., Sheng, O., Kuang R.B., Wei, Y.R., Hu, C.H., Rose, L., Karangwa, P., Yang, J., Deng, G.M., Liu, S.W., Gao, J., Viljoen, A. & Yi, G.J. 2013. Diversity and distribution of the banana wilt pathogen Fusarium oxysporum f. sp. cubense in China. Fungal Genomics and Biology doi: 10.4172/2165-8056.1000111.

Lin, Y.H., Chang, J.Y., Liu, E.T., Chao, C.P., Huang, J.H. & Chang, P.F.L. 2009. Development of a molecular marker for specific detection of Fusarium oxysporum f. sp. cubense race 4. European Journal of Plant Pathology 123: 353-365.

Mostert, D., Molina, A.B., Daniells, J., Fourie, G., Hermanto, C., Chao, C.P., Fabregar, E., Sinohin, V.G., Masdek, N., Thangavelu, R., Li, C., Yi, G., Mostert, L. & Viljoen, V. 2017. The distribution and host range of the banana Fusarium wilt fungus, Fusarium oxysporum f. sp. cubense, in Asia. PLoS ONE 12: e0181630.

O’Donnell, K., Kistler, H.C., Cigelnik, E. & Ploetz, R.C. 1998. Multiple evolution origins of the fungus causing Panama disease of banana: Concordant evidence from nuclear and mitochondrial gene genealogies. Applied Biological Sciences 95: 2044-2049.

Ploetz, R.C. 2015. Fusarium wilt of banana. Phytopathology 105: 1512-1521.

Ploetz, R.C. 2006. Fusarium wilt of banana is caused by several pathogens referred to as Fusarium oxysporum f. sp. cubense. Phytopathology 96: 653-656.

Ploetz, R.C. & Pegg, K.G. 1997. Fusarium wilt of banana and Wallace’s line: Was the disease originally restricted to his Indo-Malayan region? Australasian Plant Pathology 26: 239-249.

Ploetz, R.C., Herbert, J., Sebasigari, K., Hernandez, J.H., Pegg, K.G., Ventura, J.A. & Mayato, L.S. 1990. Importance of fusarium wilt in different banana-growing regions. In Fusarium Wilt of Banana: Proceedings of International Conference on Fusarial Wilt of Banana 1989, edited by Ploetz, R.C. USA: APS Press. pp. 9-26.

Puhalla, J.E. 1985. Classification of strains of Fusarium oxysporum on the basis of vegetative compatibility. Canadian Journal of Botany 63: 179-183.

Purwati, R.D., Hidayah, N., Sudjindro. & Sudarsono. 2008. Inoculation methods and conidial densities of Fusarium oxysporum f. sp. cubense in Abaca. HAYATI Journal of Biosciences 15: 1-7.

Stover, R.H. 1972. Banana, Plantain and Abaca Diseases. Surrey, England: Commonwealth Mycological Institute.

Sutherland, R., Viljoen, A., Myburg, A.A. & Van den Berg, N. 2013. Pathogenicity associated genes in Fusarium oxysporum f. sp. cubense race 4. South African Journal of Science 109: 1-10.

Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. & Kumar, S. 2011. MEGA 5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular Biology and Evolution 28: 2731-2739.

Visser, M., Gordon, T., Fourie, G. & Viljoen, A. 2010. Characterisation of South African isolates of Fusarium oxysporum f. sp. cubense from Cavendish Bananas. South African Journal of Science 106: 1-6.

Zulperi, D., Sijam, K., Ahmad, M., Zainal, A., Awang, Y. & Sulaiman Rahid, T. 2014. Occurrence of Ralstonia solanacearum race 2 biovar 1 associated with moko disease of banana (Musa paradisiaca cv. Nipah) in Malaysia. Journal of Phytopathology 162: 697-702.

*Corresponding author; email: dzarifah@upm.edu.my