Lionel Lebreton

French isolates of Phytophthora infestans from potato and tomato differ in phenotype and genotype

Prior to 1996, the A2 mating type of Phytophthora infestans was not detected on potato in France, but was found at one site on tomato in 1995. This finding lead to the question of the extent of differences and relationships existing between the populations of P. infestans present on each host. A collection of 76 isolates collected in France, mainly in 1996, from potato and tomato was characterised for mating type, allozyme genotype at the Gpi and Pep loci, and mitochondrial DNA haplotype; 74 of these isolates were also characterised for multilocus RFLP fingerprint, and 62 for virulence. All isolates except four showed allozyme genotypes (Gpi 90/100 or 100/100, Pep 83/100 or 100/100) and mtDNA haplotypes (Ia or IIa) characteristic of the populations introduced into Europe in the late 1970s. The four exceptions were isolates collected from tomato in Southern France in 1988-1991, which showed some characteristics of the former European populations (Gpi 86/100, Pep 92/100, mtDNA Ib). Both mating types were present among the collections from both hosts, but isolates with the A2 mating type were found on potato only in one garden crop, adjacent to tomato. Nine different RG57 fingerprints were observed, with a greater diversity among tomato isolates. Furthermore, tomato and potato collections differed markedly in the frequencies of genotypes present. Finally, tomato isolates generally had a lower virulence complexity than potato isolates. These data suggest that P. infestans populations on tomato and potato are largely separated, despite the occurrence of limited gene flow.

Aggressiveness and Competitive Fitness of Phytophthora infestans Isolates Collected from Potato and Tomato in France

ABSTRACT To test the hypothesis that host-related differences in the genotypic composition of populations of the late blight pathogen Phytophthora infestans can be explained by differential pathogenicity, the aggressiveness of isolates of the pathogen collected in France from potato and tomato was measured on detached leaflets of potato (cv. Bintje) and tomato (cv. Marmande). A preliminary trial with four isolates (two each from potato and tomato) showed that lesion appearance and development were similar for each isolate in detached leaflets and in whole plant tests in growth cabinets. Isolates collected from tomato were more pathogenic to tomato than isolates collected from potato. This was particularly the case for isolates belonging to the A2 mating type. Isolates originating from potato had a higher infection efficiency and a higher sporulation capacity on this host, but they induced lesions that generally spread more slowly than those caused by isolates from tomato. Extensive variation for components of aggressiveness on potato, and to a lesser extent on tomato, was observed in collections of isolates from each of the two hosts. Competition experiments between one potato isolate and one tomato isolate in field plots of the susceptible potato cv. Bintje clearly demonstrated the higher competitive fitness of the potato isolate on its host of origin. Therefore, differential pathogenicity to potato and tomato certainly contributes to the differentiation between P. infestans populations present on potato and tomato in France; however, additional factors, possibly related to survival ability or random genetic drift, are probably also involved and may explain the persistence of weakly pathogenic isolates in these populations.

Dominant colonisation of wheat roots by Pseudomonas fluorescens Pf29A and selection of the indigenous microflora in the presence of the take-all fungus

Increases in populations of fluorescent pseudomonads on wheat roots are usually associated with take-all decline, natural control of take-all, a disease caused by the fungus Gaeumannomyces graminis var. tritici (Ggt). Colonisation by Pseudomonas fluorescens strain Pf29A was assessed on the roots of healthy plants and of plants with take-all, and the effect of this bacterium on indigenous populations of fluorescent pseudomonads was studied. The efficacy of Pf29A as an agent for the biocontrol of take-all on five-week-old wheat seedlings was tested in non-sterile conducive soil in a growth chamber. RAPD (random amplification of polymorphic DNA) fingerprinting with a decamer primer was used to monitor strain Pf29A and culturable indigenous rhizoplane populations of fluorescent pseudomonad. Pf29A decreased disease severity and accounted for 44.6% of the culturable fluorescent pseudomonads on healthy plant rhizoplane and 75.8% on diseased plant rhizoplane. Fewer RAPD patterns were obtained when Pf29A was introduced into the soil with Ggt. In the presence of Ggt and necrotic roots, Pf29A became the dominant root coloniser and dramatically changed the diversity and the structure of indigenous fluorescent pseudomonad populations. The results show that Ggt and reduced lesion size on roots can trigger a specific increase in antagonist populations and that the introduction of a biocontrol agent in soil influences the structure of indigenous bacterial populations.

Strain-specific variation in a soilborne phytopathogenic fungus for the expression of genes involved in pH signal transduction pathway, pathogenesis and saprophytic survival in response to environmental pH changes

The soilborne fungus Gaeumannomyces graminis var. tritici (Ggt) causes take-all, a wheat root disease. In an original strain-specific way, a previous study indicates that inside the Ggt species, some strains grow preferentially at acidic pH and other strains at neutral/alkaline pH. The most important mechanism for a fungal response to the environmental pH is the Pal pathway which integrates the products of the six pal genes and the transcription factor PacC. To evaluate whether the Ggt strain-specific growth in function of the ambient pH is mediated via the Pal pathway, a transcriptional study of the genes encoding this pathway was carried out. This study provided the first evidence that the pH signalling pathway similar to those described in other fungi operated in Ggt. The pacC gene was induced at neutral pH whatever the strain. In an original way, the expression of Ggt genes coding for the different Pal proteins depended on the strain and on the ambient pH. In the strain growing better at acidic pH, few pal genes were pH-regulated, and some were overexpressed at neutral pH when regulated. In the strain growing better at neutral pH, underexpression of most of the pal genes at neutral pH occurred. The strains displayed higher gene expression in the ambient pH that unfavoured their growth as if it was a compensation system. All pH taken together, a globally weaker Pal transcript level occurred in the strains that were less sensitive to acidic pH, and on the contrary, the strain growing better on neutral pH showed higher Pal mRNA levels. The expression of genes involved in pathogenesis and saprophytic growth was also regulated by the ambient pH and the strain: each gene displayed a specific pH-regulation that was similar between strains. But all pH taken together, the global transcript levels of four out of six genes were higher in the strain growing better on neutral pH. Altogether, for the first time, the results show that inside a species, conditions affecting environmental pH modulate the expression of genes in an original strain-specific way.

Temporal dynamics of bacterial and fungal communities during the infection of Brassica rapa roots by the protist Plasmodiophora brassicae

The temporal dynamics of rhizosphere and root microbiota composition was compared between healthy and infected Chinese cabbage plants by the pathogen Plasmodiophora brassicae. When inoculated with P. brassicae, disease was measured at five sampling dates from early root hair infection to late gall development. The first symptoms of clubroot disease appeared 14 days after inoculation (DAI) and increased drastically between 14 and 35 DAI. The structure of microbial communities associated to rhizosphere soil and root from healthy and inoculated plants was characterized through high-throughput DNA sequencing of bacterial (16S) and fungal (18S) molecular markers and compared at each sampling date. In healthy plants, Proteobacteria and Bacteroidetes bacterial phyla dominated the rhizosphere and root microbiota of Chinese cabbage. Rhizosphere bacterial communities contained higher abundances of Actinobacteria and Firmicutes compared to the roots. Moreover, a drastic shift of fungal communities of healthy plants occurred between the two last sampling dates, especially in plant roots, where most of Ascomycota fungi dominated until they were replaced by a fungus assigned to the Chytridiomycota phylum. Parasitic invasion by P. brassicae disrupted the rhizosphere and root-associated community assembly at a late step during the root secondary cortical infection stage of clubroot disease. At this stage, Flavisolibacter and Streptomyces in the rhizosphere, and Bacillus in the roots, were drastically less abundant upon parasite invasion. Rhizosphere of plants colonized by P. brassicae was significantly more invaded by the Chytridiomycota fungus, which could reflect a mutualistic relationship in this compartment between these two microorganisms.

Computational analysis of the Plasmodiophora brassicae genome: mitochondrial sequence description and metabolic pathway database design

Plasmodiophora brassicae is an obligate biotrophic pathogenic protist responsible for clubroot, a root gall disease of Brassicaceae species. In addition to the reference genome of the P. brassicae European e3 isolate and the draft genomes of Canadian or Chinese isolates, we present the genome of eH, a second European isolate. Refinement of the annotation of the eH genome led to the identification of the mitochondrial genome sequence, which was found to be bigger than that of Spongospora subterranea, another plant parasitic Plasmodiophorid phylogenetically related to P. brassicae. New pathways were also predicted, such as those for the synthesis of spermidine, a polyamine up-regulated in clubbed regions of roots. A P. brassicae pathway genome database was created to facilitate the functional study of metabolic pathways in transcriptomics approaches. These available tools can help in our understanding of the regulation of P. brassicae metabolism during infection and in response to diverse constraints.

Influence of Belowground Herbivory on the Dynamics of Root and Rhizosphere Microbial Communities

Recent studies are unravelling the impact of microorganisms from the roots and rhizosphere on interactions between plants and herbivorous insects and are gradually changing our perception of the microorganisms’ capacity to affect plant defenses, but the reverse effect has seldom been investigated. Our study aimed at determining how plant herbivory influences the dynamics of root and rhizosphere microbial community assemblages and whether potential changes in root metabolites and chemical elements produced during herbivory can be related to microbial community diversity. We conducted our study on oilseed rape (Brassica napus) and its major belowground herbivore, the cabbage root fly (Delia radicum). We further assessed the influence of initial soil microbial diversity on these interactions. Different microbial diversities based on a common soil matrix were obtained through a removal-recolonization method. Root and rhizosphere sampling targeted different stages of the herbivore development corresponding to different perturbation intensities. Root bacterial communities were more affected by herbivory than some rhizosphere bacterial phyla and fungal communities, which seemed more resistant to this perturbation. Root herbivory enhanced the phylum of γ-Proteobacteria in the roots and rhizosphere, as well as the phylum of Firmicutes in the rhizosphere. Herbivory tended to decrease most root amino acids and sugars, and it increased trehalose, indolyl glucosinolates and sulfur. Higher abundances of four bacterial genera (Bacillus, Paenibacillus, Pseudomonas and Stenotrophomonas) were associated following herbivory to the increase of trehalose and some sulfur-containing compounds. Further research would help to identify the biological functions of the microbial genera impacted by plant infestation and their potential implications in plant defense.