Detection and identification of Phytophthora fragariae Hickman by the polymerase chain reaction

Phytophthora fragariae Hickman, which causes strawberry red stele and raspberry root rot, is a quarantine organism for which specific and sensitive detection methods are required to test the health of planting material. Sequences of the internal transcribed spacer regions of the ribosomal gene repeat (rDNA) were used to develop primers for P. fragariae in a nested Polymerase Chain Reaction (PCR). The fungus was readily detected in infected but symptomless roots by nested, but not single-round, PCR. It was also detected in infested water samples obtained from the Dutch General Inspection Service by nested PCR. Detection of PCR products was at least 10-fold more sensitive by PCR-ELISA than by conventional visualisation on agarose gels.

[1]  J. Duncan,et al.  Effect of temperature and host genotype on the production of inoculum by Phytophthora fragariae var. fragariae from the roots of infected strawberry plants , 1995 .

[2]  David L. Hawksworth,et al.  Ainsworth & Bisby's Dictionary of the Fungi , 1972 .

[3]  R. Casper,et al.  Nachweis von Phytophthora fragariae Hickman in Wurzeln der Erdbeerkultursorte ‘Tenira’ mit Hilfe des enzyme‐linked immunosorbent assay (ELISA) , 1987 .

[4]  J. Duncan,et al.  Analysis of RFLPs in nuclear and mitochondrial DNA and the taxonomy of Phytophthora fragariae , 1993 .

[5]  R. Nazar,et al.  Potential use of PCR-amplified ribosomal intergenic sequences in the detection and differentiation of verticillium wilt pathogens , 1991 .

[6]  J. W. Taylor,et al.  Phylogeny of five fungus-like protoctistan Phytophthora species, inferred from the internal transcribed spacers of ribosomal DNA. , 1992, Molecular biology and evolution.

[7]  T. White.  Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics , 1990 .

[8]  T. White,et al.  Detection of Phytophthora species by oligonucleotide hybridization to amplified ribosomal DNA spacers , 1993 .

[9]  S. Mohan.  Cross‐reactivity of antiserum raised against Phytophthora fragariae with other Phytophthora species and its evaluation as a genus‐detecting antiserum , 1989 .

[10]  J. Duncan,et al.  The use of nested PCR in the polymerase chain reaction for the detection of Phytophthora fragariae and P. cactorum in strawberry , 1997 .

[11]  D. Cahill,et al.  Exploitation of zoospore taxis in the development of a novel dipstick immunoassay for the specific detection of Phytophthora cinnamomi , 1994 .

[12]  J. Dunez,et al.  Immunoenzymatic detection of Phytophthora fragariae in infected strawberry plants , 1988 .

[13]  J. Duncan,et al.  Phylogenetic analysis of Phytophthora species based on ITS1 and ITS2 sequences of the ribosomal RNA gene repeat , 1997 .

[14]  J. Russell,et al.  Relatedness of Group I species of Phytophthora as assessed by randomly amplified polymorphic DNA (RAPDs) and sequences of ribosomal DNA , 1996 .

[15]  F. Rollo,et al.  Highly sensitive and fast detection of Phoma tracheiphila by polymerase chain reaction , 1990, Applied Microbiology and Biotechnology.

[16]  W. Wilcox.  Identity, virulence, and isolation frequency of seven Phytophthora spp. causing root rot of raspberry in New York , 1989 .

[17]  J. Duncan.  Phytophthora species attacking strawberry and raspberry. , 1990 .

[18]  D. Maclean,et al.  Evolutionary relationships among Phytophthora species deduced from rDNA sequence analysis , 1996 .

[19]  C. Hickman.  The Red Core Root Disease of The Strawberry Caused By Phytophthora Fragariae n.sp. , 1941 .

[20]  C. Olsson.  Diagnosis of Phytophthora infections in raspberry and strawberry plants by ELISA tests , 1995 .

[21]  J. Duncan.  A technique for detecting red stele (Phytophthora fragariae) infection of strawberry stocks before planting. , 1980 .

[22]  J. Henson,et al.  Use of polymerase chain reaction to detect Gaeumannomyces graminis DNA in plants grown in artificially and naturally infested soil. , 1993 .

[23]  K. Mullis,et al.  Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. , 1987, Methods in enzymology.