Abstract:P. infestans isolates collected from potato and tomato crops from various parts of Nepal during the 1999 and 2000 crop seasons were characterized for nuclear and mitochondrial DNA polymorphisms using restriction fragment length polymorphism markers. The nuclear DNA probe RG57 detected 11 multilocus genotypes among 280 isolates. Three genotypes were detected 21 times or more, constituting 94% of the total population, whereas frequencies of other genotypes ranged from 0.004 to 0.014. The overall genotypic diversity as estimated by the Gleason index was 1.78. Most of the overall diversity was present at the highest level (i.e., interregional, 46%), indicating limited gene flow among regions. Cluster analysis of multilocus genotypes derived from RG57 and mating type data for Nepalese isolates and representative isolates worldwide showed Nepalese isolates grouping into four clusters. Characterization of 67 isolates for mitochondrial DNA polymorphisms revealed the presence of two mt-haplotypes, Ia and Ib with the proportions of 0.88 and 0.12, respectively. Polymorphisms in nuclear and mitochondrial DNA revealed a moderate level of diversity in this population. Genotype NP3 had an identical RG57 fingerprint to US1 and had mt-haplotype Ib, confirming the presence of an old population in Nepal. Most of the genotypes had a different RG57 fingerprint than that of US1 and mt-haplotype Ia, the common characteristics of new populations. The presence of a new population at high proportions in Nepal was consistent with the global trend of mt-haplotype distribution, and suggests the displacement of old populations. This study indicates at least three possible introductions of P. infestans to Nepal.
Abstract: A new species of Phytophthora was isolated from stem and root rot of chrysanthemum in the Gifu and Toyama prefectures of Japan. The species differs from other Phytophthora species morphologically, and is characterized by nonpapillate, noncaducous sporangia with internal proliferation, formation of both hyphal swellings and chlamydospores, homothallic nature, distinctive intercalary antheridia, and funnel-shaped oogonia. The new species can grow even at 35°C, with an optimum growth temperature of 30°C in V8 juice agar medium. In phylogenetic analyses based on five nuclear regions (LSU rDNA; genes for translation elongation factor 1α, β-tubulin, 60 S ribosomal protein L10, and heat shock protein 90), the isolates formed a monophyletic clade. Although the rDNA ITS region shows a high resolution and has proven particularly useful for the separation of Phytophthora species, it was difficult to align the sequences for phylogenetic analysis. Therefore, ITS region analysis using related species as defined by the multigene phylogeny was performed, and the topology of the resulting tree also revealed a monophyletic clade formed by the isolates of the species. The morphological characteristics and phylogenetic relationships indicate that the isolates represent a new species, Phytophthora chrysanthemi sp. nov. In pathogenicity tests, chrysanthemum plants inoculated with the isolates developed lesions on stems and roots within 3 days, and the symptoms resembled the ones originally observed. Finally, the pathogen’s identity was confirmed by re-isolation from lesions of infected plants.
Abstract: Eight isolates of Phytophthora infestans were recovered from late blight infected samples collected from the districts of Mbale and Mbarara in the Eastern and Western highlands of Uganda in 2001 and analysed using mitochondrial deoxyribonucleic acid (DNA)haplotype and Amplified Fragment Length Polymorphism (AFLP) markers. Polymerase chain reaction amplification with the P2 primer followed by digestion with MspI yielded a three-fragment pattern characteristic of isolates belonging to the US-1 clonal lineage; the polymorphism was confirmed by DNA sequencing. AFLPanalysis yielded 60 markers, analysis of which clustered the Ugandan isolates with reference to US-1 isolates (US930258 and US940501). These results suggest that the examined Ugandan isolates belong to the US-1 clonage lineage.
Abstract: A total of 2691 single-lesion isolates of Phytophthora infestans was established from samples of late-blight disease from 354 commercial and garden/allotment sites in Scotland, England and Wales over four growing seasons, 1995–98. The A2 mating type was rare (3·0% of isolates) and was detected at only 34 sites. In vitro tests of sensitivity to the phenylamide fungicide metalaxyl showed that 316 sites yielded isolates with some insensitivity (resistant and/or intermediate); these were more often commercial sites than garden/allotment sites. Over the four seasons, the frequency of isolates with intermediate fungicide sensitivity increased, while the frequency of resistant phenotypes decreased. Resistant isolates were always of A1 mating type. A subset of 1459 isolates from 326 sites was analysed for molecular diversity. Mitochondrial DNA (mtDNA) haplotype Ia predominated (91·0% of isolates); haplotype IIa was present at 54 sites and both haplotypes at 33 sites. The multilocus RFLP probe RG57 detected 30 fingerprints. Four fingerprints were particularly common (RF002, RF006, RF039 and RF040) and 10 were unique to a single site in a single year. The three commonest fingerprints (RF039 >RF002 > RF006) were of A1 mating type and the fourth (RF040) was A2. RF002 isolates were resistant to the phenylamide metalaxyl and were more common in Scotland than in England and Wales. Small sample sizes limited the usefulness of estimates of diversity. Although approximately half of all sites appeared to be colonized by RF039 genotypes, some sites (both commercial and garden/ allotment) showed a higher diversity, having both common and unique genotypes. The genotypic diversity within isolates collected from commercial sites and those from garden/allotment sites were similar. The contributions of sexual reproduction and alternatives to sex to the generation of variation are discussed.
Abstract: Single-strand-conformation polymorphism (SSCP) of ribosomal DNA of 29 species (282 isolates) of Phytophthora was characterized in this study. Phytophthora boehmeriae, Phytophthora botryosa, Phytophthora cactorum, Phytophthora cambivora, Phytophthora capsici, Phytophthora cinnamomi, Phytophthora colocasiae, Phytophthora fragariae, Phytophthora heveae, Phytophthora hibernalis, Phytophthora ilicis, Phytophthora infestans, Phytophthora katsurae, Phytophthora lateralis, Phytophthora meadii, Phytophthora medicaginis, Phytophthora megakarya, Phytophthora nicotianae, Phytophthora palmivora, Phytophthora phaseoli, Phytophthora pseudotsugae, Phytophthora sojae, Phytophthora syringae, and Phytophthora tropicalis each showed a unique SSCP pattern. Phytophthora citricola, Phytophthora citrophthora, Phytophthora cryptogea, Phytophthora drechsleri, and Phytophthora megasperma each had more than one distinct pattern. A single-stranded DNA ladder also was developed, which facilitates comparison of SSCP patterns within and between gels. With a single DNA fingerprint, 277 isolates of Phytophthora recovered from irrigation water and plant tissues in Virginia were all correctly identified into eight species at substantially reduced time, labor, and cost. The SSCP analysis presented in this work will aid in studies on taxonomy, genetics, and ecology of the genus Phytophthora.
Abstract: Phytophthora species are devastating plant pathogens in both agricultural and natural environments. Due to their significant economic and environmental impact, there has been increasing interest in Phytophthora genetics and genomics, culminating in the recent release of three complete genome sequences (P. ramorum, P. sojae, and P. infestans). In this study, genome and other large sequence databases were used to identify over 225 potential genetic markers for phylogenetic analyses. Here, we present a genus-wide phylogeny for 82 Phytophthora species using seven of the most informative loci (approximately 8700 nucleotide sites). Our results support the division of the genus into 10 well-supported clades. The relationships among these clades were rigorously evaluated using a number of phylogenetic methods. This is the most comprehensive study of Phytophthora relationships to date, and many newly discovered species have
been included. A more resolved phylogeny of Phytophthora species will allow for better interpretations of the overall evolutionary history of the genus.
Abstract: A molecular phylogenetic analysis of the genus Phytophthora was performed, 113 isolates from 48 Phytophthora species were included in this analysis. Phylogenetic analyses were performed on regions of mitochondrial (cytochrome c oxidase subunit 1; NADH dehydrogenase subunit 1) and nuclear gene sequences (translation elongation factor 1x; b-tubulin) and comparisons made to test for incongruence between the mitochondrial and nuclear data sets. The genus Phytophthora was confirmed to be monophyletic. In addition, results confirm that the classical taxonomic grouping as described by [Waterhouse (1963)] does not reflect true phylogenetic relations. Phytophthora species were redistributed into 8 clades, providing a more accurate representation of phylogenetic relationships within the genus Phytophthora. The evolution and transition of morphological, pathogenic, and reproductive traits was inferred from the cladogram generated in this study. Mating system was inferred to be a homoplasious trait, with at least eight independent transitions from homothallism to heterothallism observed.
Abstract: Phenotypes of species hybrids created from in vitro fusion of zoospores from Phytophthora nicotianae and P. capsici were characterized
and compared. The species hybrids were created as part of a study of sources of genetic variation in populations of the parent species that are pathogenic over a similar range of plants. Four isolates of species hybrids proved to be similar to both P. capsici and P. nicotianae in relation to vegetative and reproductive morphologies. As in a previous study, DNA of P. capsici was detected more readily than that of P. nicotianae in all hybrid isolates. In the present study, DNA of P. nicotianae was detected in three of four hybrids by hybridization of RAPD-PCR products with species-specific DNA from P. nicotianae. By thermal denaturation analyses, DNA melting temperatures and GC contents of parent species and species hybrids were similar. The mean GC content of 47–2% was similar to GC contents reported for other Phytophthora spp. Additionally, the distributions of GC-rich regions of hybrids were more similar to the distribution in P. capsici than in P. nicotianae. By these molecular analyses, the hybrids were shown to be more similar to P. capsici than to P. nicotianae. Even though interspecific somatic fusion is likely to occur rarely under natural conditions, it could contribute to the genetic diversity of heterothallic species of Phytophthora.
Abstract: Seventy-three isolates of Phytophthora cinnamomi were collected from diseased Eucalyptus marginata (jarrah) and Corymbia calophylla (marri) trees in two forest communities in the southwest of Western Australia. Both populations of P. cinnamomi were examined for phenotypic and genotypic variation. Microsatellite DNA analysis showed that all isolates were of the same clonal lineage. We show, for the first time for P. cinnamomi, that morphological and pathogenic variation between populations of the clonal lineage are very broad and continuous. The phenotypes examined included growth rates and colony morphology on potato dextrose agar at different temperatures, sporangial and gametangial morphology, ability to form lesions in detached jarrah and marri stems, and ability to cause deaths of clonal jarrah plants in a glasshouse trial. Phenotype variation was derived asexually. All phenotypes investigated varied independently from one another. Cluster analysis of 24 morphological and pathogenicity phenotypes identified two main clusters of isolates corresponding to each population. The ability to cause deaths in both populations ranged from killing all plants within 59 d to plants being symptomless 182 d after inoculation.
Abstract: Fifty-five Phytophthora sojae isolates were collected from soil samples and diseased soybean plants from Illinois, Indiana, Iowa, and Minnesota in 1994 and 1995. Races for the isolates were determined. DNA of P. sojae isolates was amplified with 16 Operon decanucleotide primers. Twenty-three of 75 amplified fragments were polymorphic. Based on the 23 RAPD markers, a dendrogram depicting the relatedness of the isolates was constructed using UPGMA. The P. sojae isolates clustered into four distinct groups. The isolates of races 3, 4 and 25 clustered into group I. The isolates of races 1, 8 and 13 clustered into group II. The isolates of race 5 clustered into group III, and the isolates of race 7 clustered into group IV. Genetic diversity was detected among isolates of races 1, 3, 4, 5, 7 and 25 but not among isolates of races 8 and 13.
Abstract