ABSTRACT
Downy mildew of sunflower (Helianthus annuus L.), caused by the pathogen Plasmopara halstedii, is a potentially devastating disease. Seventy-seven isolates of P. halstedii collected in twelve countries from four continents were investigated for RAPD polymorphism with 21 primers. The study led to a binary matrix, which was subjected to various complementary analyses. This is the first report on the international genetic diversity of the pathogen. Similarity indices ranged from 89% to 100%. Neither a consensus unweighted pair group method with arithmetic means (UPGMA) tree constructed after bootstrap resampling of markers nor a principal component analysis based on distance matrix revealed very consistent clusterings of the isolates, and groups did not fit race or geographical origins. Phylogenies were probably obscured by limited diversity. Analysis of molecular variance (AMOVA) and Nei's genetic diversity statistics gave similar conclusions. Most of the genetic diversity was attributable to individual differences. The most differentiated races also had the lowest within-diversity indices, which suggest that they appeared recently with strong bottleneck effects. Our analyses suggest that this pathogen is probably homothallic or has an asexual mode of reproduction and that gene flow among countries can occur through commercial exchanges. Knowledge of the downy mildew populations' structure at the international level will help to devise strategies for controlling this potentially devastating disease.
Subject(s)
Genetic Variation , Helianthus/microbiology , Oomycetes/genetics , Oomycetes/isolation & purification , DNA/isolation & purification , Oomycetes/classification , Oomycetes/physiology , Phylogeny , Plant Diseases/etiology , Polymorphism, Genetic , Random Amplified Polymorphic DNA Technique/methods , Reproduction, AsexualABSTRACT
The obligate parasitic fungus-like organism Plasmopara halstedii (Farl.) Berl. et De Toni, is the causal agent of downy mildew disease in sunflower (Helianthus annuus). New races of this economically important parasite are regularly detected throughout the world. In addition, fungicide-resistant isolates have been reported in Europe and North America. These observations of parasite evolution, as well as the risk of propagation of the disease by infected seeds, means that it is necessary to guarantee the absence of Plasmopara halstedii in seed shipments. We report here the development of a rapid assay that can be used to detect infection by Plasmopara halstedii in plant tissues. Based on the nucleotide sequence information obtained from one cloned random amplified polymorphic DNA fragment, specific oligonucleotides were designed and used as primers for in vitro DNA amplification by polymerase chain reaction. An amplification product was detected on agarose gel stained with ethidium bromide when DNA from various Plasmopara halstedii races was tested, whereas no amplified DNA was detected when DNA from other origins was tested, including DNA from the host plant. The sensitivity of the technique was evaluated. The assay successfully reveals the presence of Plasmopara halstedii in infected sunflower plants prior to sporulation.
Subject(s)
Helianthus/microbiology , Oomycetes/isolation & purification , Polymerase Chain Reaction/methods , DNA, Fungal/analysis , Oomycetes/genetics , Sensitivity and Specificity , Species SpecificityABSTRACT
Identification of some economically important Tuber species using classical morphological characteristics is sometimes difficult. We report here the molecular characterization of a species coming from China, Tuber indicum, mistaken with Tuber melanosporum species. Using restriction analysis of the amplified internal transcribed spacer (ITS) of rDNA, ITS sequence analysis, and sequence characterized amplified region markers, with DNA from fruit bodies or mycorrhizae, genetic variation was found between these two species, allowing to differentiate and characterize them.
Subject(s)
Ascomycota/classification , DNA, Ribosomal/analysis , Ascomycota/genetics , Base Sequence , China , Cloning, Molecular , DNA, Fungal/analysis , DNA, Fungal/genetics , Genes, Fungal , Genome, Fungal , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Random Amplified Polymorphic DNA Technique , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Nucleic AcidABSTRACT
The Pl1 locus in sunflower, Helianthus annuus L., conferring resistance to downy mildew, Plasmopara halstedii, race 1 has been located in linkage group 1 of the consensus RFLP map of the cultivated sunflower. Bulked segregant analyses were used on 135 plants of an F2 progeny from a cross between a downy mildew susceptible line, GH, and RHA266, a line carrying Pl1. Two RFLP markers and one RAPD marker linked to the Pl1 locus have been identified. The RFLP markers are located at 5.6 cM and 7.1 cM on either side of Pl1. The RAPD marker is situated at 43.7 cM from Pl1. The significance and applications of these markers in sunflower breeding are discussed.
