Abstract
Fusarium wilt of lentil caused by Fusarium oxysporum f. sp. lentis (Fol) is a destructive pathogen limiting lentil production in India. In the present study, Secreted in Xylem (SIX) effectors genes were explored in Indian races of Fol and also a diagnostic tool for reliable detection of the disease was developed. Four SIX effectors genes, SIX11, SIX13, SIX6 and SIX2 were identified in 12 isolates of Fol belonging to seven races. SIX11 was present in all the races while SIX 13 was absent in race 6 and SIX6 was present only in race 4. The phylogenetic analysis revealed the conserved nature of the SIX genes within the forma specialis and showed sequence homology with F. oxysporum f. sp. pisi. The presence of three effectors, SIX11, SIX13 and SIX6 in race 4 correlates with high disease incidence in lentil germplasms. The in-silico characterization revealed the presence of signal peptide and localization of the effectors. Further SIX11 effector gene present in all the isolates was used to develop Fol-specific molecular marker for accurate detection. The marker developed could differentiate F. oxysporum f. sp. lycopersici, F. solani, F. oxysporum, Rhizoctonia solani and Sclerotium rolfsii and had a detection limit of 0.01ng μL− 1. The effector-based marker detection helps in the unambiguous detection of the pathogen under field conditions.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
No datasets were generated or analysed during the current study.
References
Adhikari TB, Gao A, Ingram T, Louws FJ (2020) Pathogenomics characterization of an emerging fungal pathogen, Fusarium oxysporum f. sp. lycopersici in greenhouse tomato production systems. Front Microbiol 11:1995
Ahmad S, Fook CWK, Vadamalai G, Wahab MA, Saidi NB, Zulperi D (2020) Molecular characterization of Fusarium oxysporum f. sp. cubense tropical race 4 (Foc-TR4) isolates from Malaysian banana using secreted in Xylem (SIX) effector genes. Archives Phytopathol Plant Prot 53:524–539
An B, Hou X, Guo Y, Zhao S, Luo H, He C, Wang Q (2019) The effector SIX8 is required for virulence of Fusarium oxysporum f. sp. cubense tropical race 4 to Cavendish banana. Fungal Biology 123:423–430. https://doi.org/10.1016/j.funbio.2019.03.001
Bayaa B, Erskine W (1990) A screening technique for resistance to vascular wilt in lentil. Arab J Plant Prot 8:30–33
Belabid L, Baum M, Fortas Z, Bouznad Z, Eujayl I (2004) Pathogenic and genetic characterization of Algerian isolates of Fusarium oxysporum f. sp. lentis by RAPD and AFLP analysis. Afr J Biotechnol 3:25–31. https://doi.org/10.5897/AJB2004.000-2005
Chakrabarti A, Rep M, Wang B, Ashton A, Dodds P, Ellis J (2011) Variation in potential effector genes distinguishing Australian and non-australian isolates of the cotton wilt pathogen Fusarium oxysporum f.sp. Vasinfectum. Plant Pathol 60:232–243. https://doi.org/10.1111/j.1365-3059.2010.02363.x
Chaudhary RG (2009) Pathogenic variability in isolates of Fusarium oxysporum f. sp. lentis. Trends Bioscience 2:50–52
Covey PA, Kuwitzky B, Hanson M, Webb KM (2014) Multilocus analysis using putative fungal effectors to describe a population of Fusarium oxysporum from sugar beet. Phytopathology 104:886–896. https://doi.org/10.1094/PHYTO-09-13-0248-R
Czislowski E, Fraser-Smith S, Zander M, O’Neill W, Meldrum RA, Tran‐Nguyen LT, Aitken EA (2018) Investigation of the diversity of effector genes in the banana pathogen, Fusarium oxysporum f. sp. cubense, reveals evidence of horizontal gene transfer. Mol Plant Pathol 19:1155–1171
De Oliveira CM, Do Carmo MGF, Ferreira LM, Höfte M, Do Amaral Sobrinho NMB (2021) Race identification of Fusarium oxysporum f. sp. lycopersici isolates obtained from tomato plants in Nova Friburgo, Brazil. Eur J Plant Pathol 161:273–287
Dean R, Van Kan JA, Pretorius ZA, Hammond-Kosack KE, Di Pietro A, Spanu PD, Rudd JJ, Dickman M, Kahmann R, Ellis J, Foster GD (2012) The top 10 fungal pathogens in molecular plant pathology. Mol Plant Pathol 13:414–430. https://doi.org/10.1111/j.1364-3703.2011.00783.x
Gawehns F, Houterman PM, Ichou FA, Michielse CB, Hijdra M, Cornelissen BJC, Takken FLW (2014) The Fusarium oxysporum effector Six6 contributes to virulence and suppresses I-2-mediated cell death. MPMI 27:336–348. https://doi.org/10.1094/MPMI-11-13-0330-R
Hiremani NS, Dubey SC (2018) Race profiling of Fusarium oxysporum f. sp. lentis causing wilt in lentil. Crop Prot 108:23–30. https://doi.org/10.1016/j.cropro.2018.02.010
Hiremani NS, Dubey SC (2019) Phylogenetic relationship among Indian population of Fusarium oxysporum f. sp. lentis infecting lentil and development of specific SCAR markers for detection. 3 Biotech 9(5):196. https://doi.org/10.1007/s13205-019-1734-4
Houterman PM, Speijer D, Dekker HL, De Koster CG, Cornelissen BJ, Rep M (2007) The mixed xylem sap proteome of Fusarium oxysporum-infected tomato plants. Mol Plant Pathol 8:215–221. https://doi.org/10.1111/j.1364-3703.2007.00384.x
Husien Al-Husien N, Hamwieh A, Ahmed S, Bayaa B (2017) Genetic diversity of Fusarium oxysporum f. sp. lentis population affecting lentil in Syria. J Phytopathol 165:306–312. https://doi.org/10.1111/jph.12563
Jiskani AM, Samo Y, Soomro MA, Leghari ZH, Gishkori ZGN, Bhutto SH, Majeedano AQA (2021) Destructive disease of lentil: Fusarium wilt of lentil. Plant Arch 21:2117–2127. https://doi.org/10.51470/PLANTARCHIVES.2021.v21.S1.350
Kannaiyan J, Nene YL (1978) Strains of Fusarium oxysporum f. sp. lentis and their pathogenicity on some lentil lines. LENS Newsl 5:8–10
Kelley LA, Mezulis S, Yates CM, Wass MN, Sternberg MJ (2015) The Phyre2 web portal for protein modeling, prediction and analysis. Nat Protoc 10:845–858. https://doi.org/10.1038/nprot.2015.053
Kumar A, Chordia N (2015) In silico PCR primer designing and validation. In: Basu C (ed) PCR primer design. Methods in Molecular Biology, vol 1275. Humana, New York, NY, pp 143–151. https://doi.org/10.1007/978-1-4939-2365-6_10
Laurence MH, Summerell BA, Liew ECY (2015) Fusarium oxysporum f. sp. canariensis: evidence for horizontal gene transfer of putative pathogenicity genes. Plant Pathol 64:1068–1075. https://doi.org/10.1111/ppa.12350
Leslie JF, Summerell BA (2006) The Fusarium Laboratory Manual. Blackwell Publishing, Oxford
Lievens B, Houterman PM, Rep M (2009) Effector gene screening allows unambiguous identification of Fusarium oxysporum f. sp. lycopersici races and discrimination from other formae speciales. FEMS Microbiol Lett 300:201–215
Ma LJ, Van Der Does HC, Borkovich KA, Coleman JJ, Daboussi MJ, Di Pietro A, Rep M (2010) Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium. Nature 464:367–373. https://doi.org/10.1038/nature08850
Meldrum RA, Fraser-Smith S, Tran-Nguyen LTT, Daly AM, Aitken EAB (2012) Presence of putative pathogenicity genes in isolates of Fusarium oxysporum f. sp. cubense from Australia. Australas. Plant Pathol 41:551–557. https://doi.org/10.1007/s13313-012-0122-x
Michielse CB, Rep M (2009) Pathogen profile update: Fusarium oxysporum. Mol Plant Pathol 10:311–324. https://doi.org/10.1111/j.1364-3703.2009.00538.x
Murray MG, Thompson W (1980) Rapid isolation of high molecular weight plant DNA. Nucleic Acids Res 8:4321–4326. https://doi.org/10.1093/nar/8.19.4321
Nishmitha K, Singh R, Kamil D, Dubey SC, Akthar J, Thripati K (2023) Resistance screening and in-silico characterization of cloned novel RGA from multi-race resistant Lentil germplasm against Fusarium wilt (Fusarium oxysporum f. sp. lentis). Front Plant Sci 14:1147220. https://doi.org/10.3389/fpls.2023.1147220
Pouralibaba HR, Rubiales D, Fondevilla S (2016) Identification of pathotypes in Fusarium oxysporum f. sp. lentis. Eur J Plant Pathol 144:539–549. https://doi.org/10.1007/s10658-015-0793-6
Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425. https://doi.org/10.1093/oxfordjournals.molbev.a040454
Sasaki K, Nakahara K, Tanaka S, Shigyo M, Ito SI (2015) Genetic and pathogenic variability of Fusarium oxysporum f. sp. cepae isolated from onion and Welsh onion in Japan. Phytopathology 105:525–532. https://doi.org/10.1094/PHYTO-06-14-0164-R
Sperschneider J, Dodds PN (2022) EffectorP 3.0: prediction of apoplastic and cytoplasmic effectors in fungi and oomycetes. Mol Plant Microbe Interact 35:146–156. https://doi.org/10.1094/MPMI-08-21-0201-R
Taylor A, Vágány V, Jackson AC, Harrison RJ, Rainoni A, Clarkson JP (2016) Identification of pathogenicity-related genes in Fusarium oxysporum f. sp. cepae. Mol Plant Pathol 17:1032–1047. https://doi.org/10.1111/mpp.12346
Thangavelu R, Edwinraj E, Gopi M, Pushpakanth P, Sharmila K, Prabaharan M, Uma S (2022) Development of PCR-based race-specific markers for differentiation of Indian Fusarium oxysporum f. sp. cubense, the causal agent of fusarium wilt in banana. J Fungi 8:53. https://doi.org/10.3390/jof8010053
Thatcher LF, Gardiner DM, Kazan K, Manners JM (2012) A highly conserved effector in Fusarium oxysporum is required for full virulence on Arabidopsis. Mol. Plant–Microbe Interact 25:180–190. https://doi.org/10.1094/MPMI-08-11-0212
Tintor N, Paauw M, Rep M, Takken FL (2020) The root-invading pathogen Fusarium oxysporum targets pattern‐triggered immunity using both cytoplasmic and apoplastic effectors. New Phytol 227:1479–1492. https://doi.org/10.1111/nph.16618
Untergasser A, Nijveen H, Rao X, Bisseling T, Geurts R, Leunissen JA (2007) Primer3Plus, an enhanced web interface to Primer3. Nucleic acids res 35(web server issue):W71-4. https://doi.org/10.1093/nar/gkm306
Van Dam P, Fokkens L, Ayukawa Y, Van der Gragt M, Ter Horst A, Brankovics B, Rep M (2017) A mobile pathogenicity chromosome in Fusarium oxysporum for infection of multiple cucurbit species. Sci Rep 7:9042. https://doi.org/10.1038/s41598-017-07995-y
Acknowledgements
The first author, K. Nishmitha acknowledges the senior research fellowship obtained from the Council of Scientific & Industrial Research (CSIR). Authors acknowledge ICAR-NBPGR for providing lentil seeds.
Funding
The authors declare that no funds, grants, or other support were received during the preparation of this manuscript.
Author information
Authors and Affiliations
Contributions
Conceptualization, K. Nishmitha, and Deeba Kamil; Material, S. C. Dubey; Methodology, Nishmitha K, Bishnu Maya Bashyal and S. C. Dubey; Software, Nishmitha K; Writing – original draft, Nishmitha K; Writing – review & editing, Bishnu Maya Bashyal and Deeba Kamil.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interests.
Additional information
Communicated by Nischitha R.
Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Nishmitha, K., Bashyal, B.M., Dubey, S.C. et al. Molecular characterization of Indian races of Fusarium oxysporum f. sp. lentis (Fol) based on secreted in Xylem (SIX) effector genes and development of a SIX11 gene-based molecular marker for specific detection of Fol. Arch Microbiol 206, 200 (2024). https://doi.org/10.1007/s00203-024-03945-1
Received:
Revised:
Accepted:
Published:
DOI: https://doi.org/10.1007/s00203-024-03945-1