Rapid detection of Colletotrichum gloeosporioides in infected strawberry plants using loop-mediated isothermal amplification

Katoh, Hiroshi; Fukuda, Takashi; Nishigawa, Hisashi; Natsuaki, Tomohide

doi:10.1007/s10327-016-0665-8

Cited by 15 publications

(17 citation statements)

References 41 publications

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“…LAMP amplification uses repetition of two types of elongation reactions via multiple primer sets and is rapid, simple, and highly specific [Notomi et al, Identification of the Colletotrichum species associated with mango diseases and a universal LAMP detection method for C. gloeosporioides species complex 2015]. Recent examples of LAMP-based detection of Anthracnose fungal disease include the identification of Anthracnose on strawberry using ITS1-5.8S-ITS2 sequences [Katoh et al, 2016] and the identification of Anthracnose on soybean, grape, pear, rubber, navel orange, and fishtail palm using glutamine synthetase sequences [Wang et al, 2017]. The LAMP primer sets used in these studies were designed to detect several specific crop diseases.…”

Section: Introductionmentioning

confidence: 99%

Identification of the Colletotrichum species associated with mango diseases and a universal LAMP detection method for C. gloeosporioides species complex

Hattori,

Nakashima,

Kitabata

et al. 2021

PLANT &Amp; FUNGAL RESEARCH

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The Colletotrichum gloeosporioides species complex contains plant pathogens linked to Anthracnose diseases afflicting various crops. In this study, we designed a loop-mediated isothermal amplification assay (LAMP) primer set based on calmodulin gene coding region sequences from taxonomically authorized isolates of species from this complex to rapidly detect the presence of fungi associated with Anthracnose diseases. This test can be employed at any point between cultivation and sale. Moreover, we examined the specificity and detectable range of this primer set using isolates selected from species of the genus Colletotrichum. This test was able to specifically detect members of the C. gloeosporioides species complex, including C. gloeosporioides, C. aotearoa, C. fructicola, C. horii, C. kahawae, C. musae, C. siamense, C. theobromicola, and C. tropicale. Key Words: Anthracnose, diagnosis, phylogeny, plant disease

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Section: Introductionmentioning

confidence: 99%

Identification of the Colletotrichum species associated with mango diseases and a universal LAMP detection method for C. gloeosporioides species complex

Hattori,

Nakashima,

Kitabata

et al. 2021

PLANT &Amp; FUNGAL RESEARCH

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“…Farklı kültür bitkilerinde son yıllarda Colletotrichum'a bağlı türlerin ekonomik ve hızlı tanısında LAMP tekniği, sürdürülebilir ve hassas bir yöntem olarak kabul edilmeye başlanmıştır. Çilek bitkilerini enfekte eden C. gloeosporioides türü (Katoh et al 2016), şekerkamışı bitkisinde kırmızı çürüklük patojeni C. falcatum türü (Chandra et al 2015), biberde antraknoza neden olan C. capsici fungal patojeni (Kandan et al 2016) ve soya fasulyesi bitkisinde antraknoza yol açan C. acutatum türünün (Tian et al 2017) tespiti ve tanısında LAMP tekniğinin hızlı, güvenilir, ekonomik ve yararlı sonuçlar ortaya koyduğu laboratuvar ve saha çalışmaları ile açık ve net olarak görülmektedir. Bu bulgular, çalışmamızda elde ettiğimiz sonuçlarla uyumlu ve bunları destekler niteliktedir.…”

Section: Tartişma Ve Kaniunclassified

“…). Enfekteli çilek bitkilerinde C. gloeosporioides'in hızlı tespiti için yapılan çalışmada(Katoh et al 2016), LAMP tekniği ile C. gloeosporioides'in hızlı ve hassas bir şekilde tespit edildiği ve C. acutatum, C. aenigma ve C. siamense gibi farklı Colletotrichum türlerinin de ayırt edilebildiği bildirilmiştir. Ayrıca, LAMP tekniği kullanarak C. acutatum'un neden olduğu soya antraknozunun hızlı tespiti şeklinde çalışma yapılmıştır.…”

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“…Öte yandan Hindistan'ın farklı bölgelerinden toplanan enfekteli biber tohumlarından C. capsici'nin hızlı ve hassas tespiti için yapılan çalışmada, klasik PCR'a göre, LAMP tekniğinin güvenilir bir şeklide kullanılabileceği kanıtlanmıştır (Kandan et al 2016). Enfekteli çilek bitkilerinde C. gloeosporioides'in hızlı tespiti için yapılan çalışmada (Katoh et al 2016) Şekil 2. LAMP tekniği ve PCR C. gloeosporioides ve F. solani'nin elektroforez jel performansları (A1, size marker: LAMP tekniği, A2-A5: C. gloeosporioides, A6: F. solani, A11-A12:GAPDH primeri ile C. gloeosporioides'in PCR analizi)…”

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Rapid Diagnosis of Citrus Anthracnose Caused by Colletotrichum gloeosporioides Using a LAMP (Loop-Mediated Isothermal Amplification) Assay

Uysal

Kurt

2020

Bitki Koruma Bülteni

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Anthracnose disease in citrus occurs as dieback in shoots, spot on leaves, early leaf and fruit drop and anthracnose symptoms in fruits. Colletotrichum gloeosporioides, a causal agent of anthracnose, causes severe infections in many citrus species and varieties, including lemon. Fungal pathogen C. gloeosporioides can be diagnosed by morphological, molecular methods and protein-based spectrum analyzes. This study was conducted to perform a rapid identification of C. gloeosporioides, using the LAMP technique. Specific primers for the LAMP method were designed using the computer program (Primary Primer Explorer V3) (http://primerexplorer.jp/e), with primer sets specific to each gene region (F3 / B3 and FIP / BIP) of the gene sequences of the C. gloeosporioides strain. The specificity of the primers intended for use in the LAMP reaction was evaluated by using different three isolates of C. gloeosporioides and Fs4 isolates of Fusarium solani using a total of 3 DNA samples. Genomic DNA was used in the amount of 1 ng µl-1 in the PCR study and 10 fg µl-1 in the LAMP study. As a result, C. gloeosporioides isolates glowed in tubes when examined with fluorescent dye. However, no glare was observed in the tube with F. solani. LAMP analysis showed that it successfully detected genomic DNA from C. gloeosporioides, but F. solani was unable to detect the genomic DNA obtained. After 2 hours of analysis with LAMP technique, the pathogen was detected in a short time with the successful results.

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“…Loop-Mediated Isothermal Amplification has already been deployed in the detection of important strawberry pathogens with success [21][22][23][24][25][26][27][28]. Additionally, the LAMP assay for the detection of Phytophthora spp.…”

Section: Introductionmentioning

confidence: 99%

“Shining a LAMP” (Loop-Mediated Isothermal Amplification) on the Molecular Detection of Phytopathogens Phytophthora spp. and Phytophthora cactorum in Strawberry Fields

2021

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Phytopathogenic microorganisms belonging to the genus Phytophthora have been recognized many times as causal agents of diseases that lower the yield of many plants important for agriculture. Meanwhile, Phytophthora cactorum causes crown rot and leather rot of berry fruits, mainly strawberries. However, widely-applied culture-based methods used for the detection of pathogens are time-consuming and often inaccurate. What is more, molecular techniques require costly equipment. Here we show a rapid and effective detection method for the aforementioned targets, deploying a simple molecular biology technique, Loop-Mediated Isothermal Amplification (LAMP). We optimized assays to amplify the translation elongation factor 1-α (EF1a) gene for two targets: Phytophthora spp. And Phytophthora cactorum. We optimized the LAMP on pure strains of the pathogens, isolated from organic plantations of strawberry, and successfully validated the assay on biological material from the environment including soil samples, rhizosphere, shoots and roots of strawberry, and with SYBR Green. Our results demonstrate that a simple and reliable molecular detection method, that requires only a thermoblock and simple DNA isolation kit, can be successfully applied to detect pathogens that are difficult to separate from the field. We anticipate our findings to be a starting point for developing easier and faster modifications of the isothermal detection methods and which can be applied directly in the plantation, in particular with the use of freeze-dried reagents and chemistry, allowing observation of the results with the naked eye.

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Rapid detection of Colletotrichum gloeosporioides in infected strawberry plants using loop-mediated isothermal amplification

Cited by 15 publications

References 41 publications

Identification of the Colletotrichum species associated with mango diseases and a universal LAMP detection method for C. gloeosporioides species complex

Identification of the Colletotrichum species associated with mango diseases and a universal LAMP detection method for C. gloeosporioides species complex

Rapid Diagnosis of Citrus Anthracnose Caused by Colletotrichum gloeosporioides Using a LAMP (Loop-Mediated Isothermal Amplification) Assay

“Shining a LAMP” (Loop-Mediated Isothermal Amplification) on the Molecular Detection of Phytopathogens Phytophthora spp. and Phytophthora cactorum in Strawberry Fields

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