Molecular Detection of Plasmodiophora brassicae, Causal Agent of Clubroot of Crucifers, in Plant and Soil.

Clubroot of crucifers, caused by Plasmodiophora brassicae, recently has been identified in canola (Brassica napus) fields in Alberta, Canada. An effective strategy for managing the disease is to avoid planting cruciferous crops in P. brassicae-infested soil, because the pathogen produces resting spores that can remain infectious for many years. A simple, one-step polymerase chain reaction (PCR) protocol was developed to detect the pathogen in plant and soil samples. The primers TC1F and TC1R, based on a P. brassicae partial 18S ribosomal RNA (rRNA) gene sequence from GenBank, yielded a 548-bp product in the optimized PCR. A second pair of primers, TC2F and TC2R, which amplified a fragment of the 18S and internal transcribed spacer (ITS) 1 regions of the rDNA repeat, also was tested and produced a 519-bp product. Neither set of primers amplified any DNA fragment from noninfected plant hosts, noninfested soil, or common soil fungi and bacteria tested in this study. Quantities of 100 fg or less of total P. brassicae DNA, or 1 × 103 resting spores per gram of soil, could be detected consistently using these primers and PCR protocol, corresponding to an index of disease of 11% or lower when the soil was bioassayed. The protocol also enabled detection of P. brassicae in symptomless root tissue 3 days after inoculation with the pathogen. Therefore, the PCR assay described in this study could provide a reliable diagnosis for routine detection of P. brassicae in plant and soil materials in a specific and rapid manner.

Genetic diversity and recombination within populations of Fusarium pseudograminearum from western Canada.

Genetic diversity within populations of Fusarium pseudograminearum isolated from wheat grains from the Canadian provinces of Alberta and Saskatchewan was investigated. Three restriction enzymes (EcoRI, HaeIII, and PstI) were used to carry out restriction analysis of the nuclear ribosomal DNA (nrDNA) intergenic spacer region (IGS region) and eight primers were used to generate inter-simple sequence-repeat (ISSR) molecular markers. Our study indicated substantially high genetic diversity within these two populations, but low genetic differentiation and frequent gene flow among populations. The IGS data showed no genetic distinction between the two Alberta populations and only minor genetic differentiation between the Saskatchewan and Alberta populations. Analysis of molecular variance indicated that most genetic variability resulted from differences among isolates within populations. Multilocus linkage disequilibrium analysis suggested a panmictic population genetic structure and the occurrence of significant recombination in F. pseudograminearum. Regular gene flow and random mating between isolates from different populations could result in novel genotypes with both improved pathological and biological traits.

Genetic diversity and recombination within populations of Fusarium pseudograminearum from western Canada / Diversidad genética y recombinación en poblaciones de Fusarium pseudograminearum del oeste de Canadá

Genetic diversity within populations of Fusarium pseudograminearum isolated from wheat grains from the Canadian provinces of Alberta and Saskatchewan was investigated. Three restriction enzymes (EcoRI, HaeIII, and PstI) were used to carry out restriction analysis of the nuclear ribosomal DNA (nrDNA) intergenic spacer region (IGS region) and eight primers were used to generate inter-simple sequence-repeat (ISSR) molecular markers. Our study indicated substantially high genetic diversity within these two populations, but low genetic differentiation and frequent gene flow among populations. The IGS data showed no genetic distinction between the two Alberta populations and only minor genetic differentiation between the Saskatchewan and Alberta populations. Analysis of molecular variance indicated that most genetic variability resulted from differences among isolates within populations. Multilocus linkage disequilibrium analysis suggested a panmictic population genetic structure and the occurrence of significant recombination in F. pseudograminearum. Regular gene flow and random mating between isolates from different populations could result in novel genotypes with both improved pathological and biological traits (AU)

Se investigó la diversidad genética en poblaciones de Fusarium pseudograminearum aisladas de semillas de trigo de las provincias canadienses de Alberta y Saskatchewan. Se usaron tres enzimas de restricción (EcoRI, HaeIII, and PstI) para analizar los marcadores moleculares de las regiones espaciadoras intergénicas (IGS) del DNA ribosómico del núcleo (nrDNA) y de los fragmentos generados entre repeticiones de secuencias sencillas (fragmentos ISSR). Nuestro estudió reveló una gran diversidad genética en ambas poblaciones, pero poca diferenciación genética y un flujo genético frecuente entre las poblaciones. Los datos relativos a las IGS no mostraron diferencias genéticas entre las dos poblaciones de Alberta estudiadas y sólo una ligera diferenciación entre las poblaciones de Alberta y de Saskatchewan. El análisis de la varianza molecular indicó que la variabilidad genética respondía en su mayor parte a diferencias entre aislamientos dentro de las poblaciones. El análisis del desequilibrio en el ligamiento genético sugería una estructura genética de la población de tipo panmíctico y la existencia de una recombinación significativa en F. pseudograminearum. Un flujo genético regular y el apareamiento al azar de aislamientos de poblaciones distintas podría producir nuevos genotipos con características biológicas mejores o patológicas (AU)

Dynamics of Rhynchosporium secalis pathotypes in relation to barley cultivar resistance.

Rhynchosporium secalis isolates E97-2 and H97-2, represented the major pathotypes in populations on barley in Alberta, Canada, but differed widely in their virulence. Following greenhouse co-inoculation with the two pathotypes, E97-2, originally isolated from resistant cv. 'CDC Earl', predominated over H97-2, isolated from the susceptible cv. 'Harrington', from the first to the last of four infection cycles on both 'CDC Earl' and 'Harrington'. These results indicated that the host can rapidly influence pathotype composition and that pathotype E97-2 may have a competitive advantage over H97-2 on these cultivars. DNA polymorphisms were found between isolates from single or mixed inoculations on cvs 'CDC Earl' and 'Harrington' for four successive cycles. Co-inoculation with the two isolates resulted in a shift to a molecular phenotype more similar to E97-2 than H97-2. The competitive advantage of E97-2 over H97-2, combined with the selective pressure exerted by the host, would explain the increased susceptibility of cv. 'CDC Earl' and other cultivars with similar sources of scald resistance, in fields across Alberta. However, H97-2 will likely remain one of the major pathotypes in Alberta due to the relatively large acreage of cv. 'Harrington' in this province.