Novel Loop-mediated Isothermal Amplification (LAMP) Assay with a Universal QProbe Can Detect SNPs Determining Races in Plant Pathogenic Fungi

Overview

Journal Sci Rep

Specialty Science

Date 2017 Jun 28

PMID 28652587

Citations 20

Authors

Yu Ayukawa

Saeri Hanyuda

Naoko Fujita

Ken Komatsu

Tsutomu Arie

Affiliations

Soon will be listed here.

Abstract

Tomato wilt pathogen Fusarium oxysporum f. sp. lycopersici (Fol) is grouped into three races based on their pathogenicity to different host cultivars. Rapid detection and discrimination of Fol races in field soils is important to prevent tomato wilt disease. Although five types of point mutations in secreted in xylem 3 (SIX3) gene, which are characteristic of race 3, have been reported as a molecular marker for the race, detection of these point mutations is laborious. The aim of this study is to develop a rapid and accurate method for the detection of point mutations in SIX3 of Fol. Loop-mediated isothermal amplification (LAMP) of SIX3 gene with the universal QProbe as well as two joint DNAs followed by annealing curve analysis allowed us to specifically detect Fol and discriminate race 3 among other races in about one hour. Our developed method is applicable for detection of races of other plant pathogenic fungi as well as their pesticide-resistant mutants that arise through point mutations in a particular gene.

Citing Articles

LAMP Reaction in Plant Disease Surveillance: Applications, Challenges, and Future Perspectives.

Aglietti C, Benigno A, Cacciola S, Moricca S Life (Basel). 2025; 14(12.

PMID: 39768257 PMC: 11678381. DOI: 10.3390/life14121549.

Rapid and accurate detection of f. sp. Lycopersici using one-pot, one-step LAMP-CRISPR/Cas12b method.

Dai S, Wu Y, Zhu N, Zhao Y, Mao M, Li Z Front Plant Sci. 2025; 15:1485884.

PMID: 39759228 PMC: 11695371. DOI: 10.3389/fpls.2024.1485884.

Rapid real-time quantitative colorimetric LAMP methodology for field detection of Verticillium dahliae in crude olive-plant samples.

Megariti M, Panagou A, Patsis G, Papadakis G, Pantazis A, Paplomatas E Plant Methods. 2024; 20(1):139.

PMID: 39252004 PMC: 11386372. DOI: 10.1186/s13007-024-01251-x.

Evolution of the Probe-Based Loop-Mediated Isothermal Amplification (LAMP) Assays in Pathogen Detection.

Zhang X, Zhao Y, Zeng Y, Zhang C Diagnostics (Basel). 2023; 13(9).

PMID: 37174922 PMC: 10177487. DOI: 10.3390/diagnostics13091530.

A highly multiplexed assay to monitor pathogenicity, fungicide resistance and gene flow in the fungal wheat pathogen Zymoseptoria tritici.

Bellah H, Gazeau G, Gelisse S, Amezrou R, Marcel T, Croll D PLoS One. 2023; 18(2):e0281181.

PMID: 36745583 PMC: 9901794. DOI: 10.1371/journal.pone.0281181.

References

Duan Y, Yang Y, Wang Y, Pan X, Wu J, Cai Y . Loop-Mediated Isothermal Amplification for the Rapid Detection of the F200Y Mutant Genotype of Carbendazim-Resistant Isolates of Sclerotinia sclerotiorum. Plant Dis. 2019; 100(5):976-983. DOI: 10.1094/PDIS-10-15-1185-RE. View

Badolo A, Bando H, Traore A, Ko-Ketsu M, Guelbeogo W, Kanuka H . Detection of G119S ace-1 (R) mutation in field-collected Anopheles gambiae mosquitoes using allele-specific loop-mediated isothermal amplification (AS-LAMP) method. Malar J. 2015; 14:477. PMC: 4665897. DOI: 10.1186/s12936-015-0968-9. View

Tani H, Miyata R, Ichikawa K, Morishita S, Kurata S, Nakamura K . Universal quenching probe system: flexible, specific, and cost-effective real-time polymerase chain reaction method. Anal Chem. 2009; 81(14):5678-85. DOI: 10.1021/ac900414u. View

Takken F, Rep M . The arms race between tomato and Fusarium oxysporum. Mol Plant Pathol. 2010; 11(2):309-14. PMC: 6640361. DOI: 10.1111/j.1364-3703.2009.00605.x. View

Biju V, Fokkens L, Houterman P, Rep M, Cornelissen B . Multiple Evolutionary Trajectories Have Led to the Emergence of Races in Fusarium oxysporum f. sp. lycopersici. Appl Environ Microbiol. 2016; 83(4). PMC: 5288826. DOI: 10.1128/AEM.02548-16. View

Joosten M, Cozijnsen T, De Wit P . Host resistance to a fungal tomato pathogen lost by a single base-pair change in an avirulence gene. Nature. 1994; 367(6461):384-6. DOI: 10.1038/367384a0. View

Badolo A, Okado K, Guelbeogo W, Aonuma H, Bando H, Fukumoto S . Development of an allele-specific, loop-mediated, isothermal amplification method (AS-LAMP) to detect the L1014F kdr-w mutation in Anopheles gambiae s. l. Malar J. 2012; 11:227. PMC: 3407793. DOI: 10.1186/1475-2875-11-227. View

Lievens B, Houterman P, Rep M . Effector gene screening allows unambiguous identification of Fusarium oxysporum f. sp. lycopersici races and discrimination from other formae speciales. FEMS Microbiol Lett. 2009; 300(2):201-15. DOI: 10.1111/j.1574-6968.2009.01783.x. View

Duan Y, Zhang X, Ge C, Wang Y, Cao J, Jia X . Development and application of loop-mediated isothermal amplification for detection of the F167Y mutation of carbendazim-resistant isolates in Fusarium graminearum. Sci Rep. 2014; 4:7094. PMC: 4235284. DOI: 10.1038/srep07094. View

10.

Zhang C, Yao Y, Zhu J, Zhang S, Zhang S, Wei H . Establishment and application of a real-time loop-mediated isothermal amplification system for the detection of CYP2C19 polymorphisms. Sci Rep. 2016; 6:26533. PMC: 4887897. DOI: 10.1038/srep26533. View

11.

Inami K, Yoshioka-Akiyama C, Morita Y, Yamasaki M, Teraoka T, Arie T . A genetic mechanism for emergence of races in Fusarium oxysporum f. sp. lycopersici: inactivation of avirulence gene AVR1 by transposon insertion. PLoS One. 2012; 7(8):e44101. PMC: 3428301. DOI: 10.1371/journal.pone.0044101. View

12.

Kurata S, Kanagawa T, Yamada K, Torimura M, Yokomaku T, Kamagata Y . Fluorescent quenching-based quantitative detection of specific DNA/RNA using a BODIPY((R)) FL-labeled probe or primer. Nucleic Acids Res. 2001; 29(6):E34. PMC: 29763. DOI: 10.1093/nar/29.6.e34. View

13.

Ikeda S, Takabe K, Inagaki M, Funakoshi N, Suzuki K . Detection of gene point mutation in paraffin sections using in situ loop-mediated isothermal amplification. Pathol Int. 2007; 57(9):594-9. DOI: 10.1111/j.1440-1827.2007.02144.x. View

14.

Heckman K, Pease L . Gene splicing and mutagenesis by PCR-driven overlap extension. Nat Protoc. 2007; 2(4):924-32. DOI: 10.1038/nprot.2007.132. View

15.

Platt A, Woodhall R, George Jr A . Improved DNA sequencing quality and efficiency using an optimized fast cycle sequencing protocol. Biotechniques. 2007; 43(1):58, 60, 62. DOI: 10.2144/000112499. View

16.

Houterman P, Cornelissen B, Rep M . Suppression of plant resistance gene-based immunity by a fungal effector. PLoS Pathog. 2008; 4(5):e1000061. PMC: 2330162. DOI: 10.1371/journal.ppat.1000061. View

17.

Ayukawa Y, Komatsu K, Kashiwa T, Akai K, Yamada M, Teraoka T . Detection and differentiation of Fusarium oxysporum f. sp. lycopersici race 1 using loop-mediated isothermal amplification with three primer sets. Lett Appl Microbiol. 2016; 63(3):202-9. DOI: 10.1111/lam.12597. View

18.

Thatcher L, Gardiner D, Kazan K, Manners J . A highly conserved effector in Fusarium oxysporum is required for full virulence on Arabidopsis. Mol Plant Microbe Interact. 2011; 25(2):180-90. DOI: 10.1094/MPMI-08-11-0212. View

19.

Sasaki K, Nakahara K, Tanaka S, Shigyo M, Ito S . Genetic and Pathogenic Variability of Fusarium oxysporum f. sp. cepae Isolated from Onion and Welsh Onion in Japan. Phytopathology. 2014; 105(4):525-32. DOI: 10.1094/PHYTO-06-14-0164-R. View

20.

Kashiwa T, Suzuki T, Sato A, Akai K, Teraoka T, Komatsu K . A new biotype of Fusarium oxysporum f. sp. lycopersici race 2 emerged by a transposon-driven mutation of avirulence gene AVR1. FEMS Microbiol Lett. 2016; 363(14). DOI: 10.1093/femsle/fnw132. View