The Use of Nested PCR for the Detection of Phytophthora Fragariae in Strawberry

Bonants, P.J.M.; Weerdt, M. Hagenaar-de; Gent-Pelzer, M.P.E. van; Lacourt, Isabelle; Cooke, D.E.L.; Duncan, James M.

doi:10.1007/978-94-009-0043-1_9

Cited by 2 publications

(2 citation statements)

References 11 publications

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“…The DNA of the different isolates was amplified by polymerase chain reaction (PCR) using ITS 4 and DC 6 set of primers. Forward primer DC 6 (5´-GAGGGACTTTTGGGTAATCA-3´) and the reverse primer ITS4 (5´-TCCTCCGCTTATTGATATGC-3´) were designed based on sequences of 18S rDNA from a number of fungi (White et al 1990;Bonants et al 1997). The final 50 µl reaction mixture contained 1× PCR buffer (NEB, England), 1 nmol of dNTPs, 1 pmol of 2 mM MgSO 4 , 0.25 pmol of forward and reverse primers, 1 unit Taq DNA polymerase (NEB, England) and 10 µl template DNA (Barakat et al 2017).…”

Section: Molecular Identificationmentioning

confidence: 99%

Journal of Plant Protection Research

Elshahawy¹,

Abouelnasr²,

Lashin³

et al. 2018

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In August 2016, tomato plants grown during a hot, wet summer with heavy soil flooding, displaying symptoms of wilting, dead plant, root rot with crown and stem rot, at Beni Suef and Fayoum governorates were examined. A number of 16 fungal isolates were isolated from tomato plants displaying the above symptoms. These isolates were classified as belonging to six species, namely: Alternaria solani, Chaetomium globosum, Fusarium solani, Fusarium oxysporum, Pythium spp. and Rhizoctonia solani. Isolates of Pythium spp. were prevalent and were found to be more pathogenic than the other fungal isolates. This species causes damping-off, root rot, sudden death, stem rot and fruit rot. The pathogen was identified as Pythium aphanidermatum based on morphological, cultural, and molecular characteristics. Biogenic silver nanoparticles (AgNPs) were produced using the F. oxyspo rum strain and characterized by transmission electron microscopy (TEM). The size of these spherical particles ranged from 10 to 30 nm. In vitro, biogenic AgNPs showed antifungal activity against P. aphanidermatum. In greenhouse and field experiments, AgNPs treatment significantly reduced the incidence of dead tomato plants due to root rot caused by P. aphanidermatum compared to the control. All of the investigated treatments were effective and the treatment of root dipping plus soil drenching was the most effective. To the best of our knowledge, this study describes P. aphanidermatum on tomato in Egypt for the first time. Also, biogenic AgNPs could be used for controlling root rot disease caused by this pathogen.

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Section: Molecular Identificationmentioning

confidence: 99%

Journal of Plant Protection Research

Elshahawy¹,

Abouelnasr²,

Lashin³

et al. 2018

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“…shown that the fungus can specifically and sensitively be detected in roots with nested PCR (Bonants et al, 1996(Bonants et al, , 1997, as well as in water samples. This nested PCR procedure was used to screen many root and water samples from bait tests.…”

Section: P Fragariae Have Been Developed Previous Research Hasmentioning

confidence: 99%

Characterization and detection of Phytophthora fragariae in plant, water and soil by molecular methods*

Bonants¹,

Gent-Pelzer²,

Weerdt³

2000

EPPO Bulletin

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Phytophthora fragariae var. frugariae is pathogenic to strawberry only and is an EPPO A2 quarantine pest. Seventy-five isolates of P. fruguriae from all over the world have been characterized by amplified fragment length polymorphism (AFLP) analysis. However, a sensitive, specific and fast method of detection is now needed, and polymerase chain reaction (PCR)-based methods have now been set up. Based upon sequences of the ITS (internal transcribed spacer) regions of rDNA of several Phytophthora species, PCR primers specific for P.fragariae have been developed together with SCRI (Dundee, GB). These primers were tested on DNA extracted from water, soil and plant material. A nested PCR procedure has been included in the currently used bait test to improve sensitivity and reliability. Also several methods have been studied to detect the amplicon of the PCR reaction: gel electrophoresis, DIAPOPS (detection of immobilized amplified products in a one-phase system), PCR-ELISA, TaqMan and molecular beacon. With the last two methods, using the ABI 7700 detection system, quantification of target DNA is possible.

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The Use of Nested PCR for the Detection of Phytophthora Fragariae in Strawberry

Cited by 2 publications

References 11 publications

Journal of Plant Protection Research

Journal of Plant Protection Research

Characterization and detection of Phytophthora fragariae in plant, water and soil by molecular methods*

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