Michael G. Milgroom

Genome Expansion and Gene Loss in Powdery Mildew Fungi Reveal Tradeoffs in Extreme Parasitism

From Blight to Powdery Mildew Pathogenic effects of microbes on plants have widespread consequences. Witness, for example, the cultural upheavals driven by potato blight in the 1800s. A variety of microbial pathogens continue to afflict crop plants today, driving both loss of yield and incurring the increased costs of control mechanisms. Now, four reports analyze microbial genomes in order to understand better how plant pathogens function (see the Perspective by Dodds). Raffaele et al. (p. 1540) describe how the genome of the potato blight pathogen accommodates transfer to different hosts. Spanu et al. (p. 1543) analyze what it takes to be an obligate biotroph in barley powdery mildew, and Baxter et al. (p. 1549) ask a similar question for a natural pathogen of Arabidopsis. Schirawski et al. (p. 1546) compared genomes of maize pathogens to identify virulence determinants. Better knowledge of what in a genome makes a pathogen efficient and deadly is likely to be useful for improving agricultural crop management and breeding. A group of papers analyzes pathogen genomes to find the roots of virulence, opportunism, and life-style determinants. Powdery mildews are phytopathogens whose growth and reproduction are entirely dependent on living plant cells. The molecular basis of this life-style, obligate biotrophy, remains unknown. We present the genome analysis of barley powdery mildew, Blumeria graminis f.sp. hordei (Blumeria), as well as a comparison with the analysis of two powdery mildews pathogenic on dicotyledonous plants. These genomes display massive retrotransposon proliferation, genome-size expansion, and gene losses. The missing genes encode enzymes of primary and secondary metabolism, carbohydrate-active enzymes, and transporters, probably reflecting their redundancy in an exclusively biotrophic life-style. Among the 248 candidate effectors of pathogenesis identified in the Blumeria genome, very few (less than 10) define a core set conserved in all three mildews, suggesting that most effectors represent species-specific adaptations.

Clonal dispersal and spatial mixing in populations of the plant pathogenic fungus, Sclerotinia sclerotiorum

Two thousand seven hundred and forty‐seven isolates of Sclerotinia sclerotiorum were sampled from four field populations of canola in western Canada. Each field was sampled in a grid of 128 50‐m 50‐m quadrats plus four intensive quadrats each sampled in a diagonal transect. Sampling was done at two phases of the disease cycle: (1) from ascospore inoculum on petals and (2) from disease lesions in stems. A total of 594 unique genotypes was identified by DNA fingerprinting. In each field, a small group of clones represented the majority of the sample, with a large group of clones or genotypes sampled once or twice. Clone frequencies were compared by χ2 tests. The difference in profiles of clone frequencies for the two fields sampled in 1991 was not significant, but in 1992 the difference in profiles was marginally significant, indicating some local population substructure. The difference in profiles of clone frequencies for petals and lesions was not significant in each of the two fields sampled in 1991. In each of the two fields sampled in 1992, however, the difference was highly significant, consistent either with selection for some clones or with waves of immigration during the disease cycle. Nine of the 30 most frequently sampled clones from this study were previously recovered in a macrogeographical sample from western Canada in 1990. For spatial analyses, randomization tests indicated no significant spatial aggregation of either clones on petals or clones from lesions. Also, isolates of a clone on petals were not closer to isolates of the same clone from lesions than could be predicted by chance. Both observations suggest spatial mixing of ascospore inoculum from resident or immigrant sources.

Contributions of Population Genetics to Plant Disease Epidemiology and Management

Publisher Summary This chapter discusses contributions of population genetics to plant disease epidemiology and management. Population genetics and genetic variation in plant pathogens are subjects that have generated much interest since the late 1980s. Almost every recent issue of major plant pathological and mycological journals has at least one article on genetic variation of a plant pathogen species. Whether population genetics becomes an integral discipline within plant pathology depends, in part, on whether it can be integrated with epidemiology and disease management. Evolutionary biology and population genetics have the potential to deliver much basic information about plant pathogens. Population genetics is a field concerned with determining the extent and pattern of genetic variation in populations with the goal of understanding the evolutionary processes affecting the origin and maintenance of genetic variation. The conceptual framework is based on evolutionary biology and on the processes affecting the genetic composition of populations: selection, mutation, gene flow, genetic drift, and mating systems. The advances in technology also brought about a marked change in emphasis in population genetics of plant pathogens. The biology of pathogens at the population level and processes other than selection has been emphasized. Accurate population definition is essential so that sampling is done in a manner which will enable inferences to be made about the population of interest. From an operational perspective, recognition of population structure is very important when estimating population genetic parameters.

Intercontinental population structure of the chestnut blight fungus, Cryphonectria parasitica

The population structure of the chestnut blight fungus, Cryphonectria parasitica was analyzed using restriction fragment length polymorphisms (RFLPs). A total of 791 isolates were sampled from four...

Amplified fragment length polymorphism (AFLP) fingerprinting of symbiotic fungi cultured by the fungus-growing ant Cyphomyrmex minutus.

A PCR‐based fingerprinting technique based on amplified fragment length polymorphisms (AFLP) is used to screen symbiotic fungi of the fungus‐growing ant Cyphomyrmex minutus for genetic differences. AFLP fingerprints reveal several fungal ‘types’ that (a) represent distinct clones propagated vegetatively by the ant, or (b) correspond to free‐living fungi that may be acquired by the ant. Fungal types identified by AFLP fingerprints correspond to vegetative‐compatibility groups established previously, suggesting that vegetative compatibility can be used as a crude indicator of genetic differences between fungi of C. minutus.

Programmed cell death correlates with virus transmission in a filamentous fungus.

Programmed cell death (PCD) is an essential part of the defence response in plants and animals against pathogens. Here, we report that PCD is also involved in defence against pathogens of fungi. Vegetative incompatibility is a self/non–self recognition system in fungi that results in PCD when cells of incompatible strains fuse. We quantified the frequency of cell death associated with six vegetative incompatibility (vic) genes in the filamentous ascomycete fungus Cryphonectria parasitica. Cell death frequencies were compared with the effects of vic genes on transmission of viruses between the same strains. We found a significant negative correlation between cell death and virus transmission. We also show that asymmetry in cell death correlates with asymmetry in virus transmission; greater transmission occurs into vic genotypes that exhibit delayed or infrequent PCD after fusion with an incompatible strain. Furthermore, we found that virus infection can have a significant, strain–specific, positive or negative effect on PCD. Specific interactions between vic gene function and viruses, along with correlations between cell death and transmission, strongly implicate PCD as a host–mediated pathogen defence strategy in fungi.

Distribution and diversity of vegetative compatibility types in subpopulations of Cryphonectria parasitica in Italy

Using a medium that discriminated with high resolution, 20 vegetative compatibility (vc) types were detected among a sample of 716 isolates of Cryphonectria parasitica from 11 widely separated subpopulations throughout Italy. Each isolate was assigned unambiguously to a single vc type; no isolates were compatible with more than one vc type. Most isolates (85%) were in four common vc types; the other 16 vc types were represented by only a few isolates each. Vegetative compatibility type frequencies were markedly different between northern and southern Italy. Three of the most common vc types (I-1, I-2 and I-5) were found mostly in the North, where diversity was greatest, while two others (I-10 and I-12) were primarily found in the South.

The chestnut blight fungus world tour: successive introduction events from diverse origins in an invasive plant fungal pathogen

Clonal expansion has been observed in several invasive fungal plant pathogens colonizing new areas, raising the question of the origin of clonal lineages. Using microsatellite markers, we retraced the evolutionary history of introduction of the chestnut blight fungus, Cryphonectria parasitica, in North America and western Europe. Combining discriminant analysis of principal components and approximate Bayesian computation analysis, we showed that several introduction events from genetically differentiated source populations have occurred in both invaded areas. In addition, a low signal of genetic recombination among different source populations was suggested in North America. Finally, two genetic lineages were present in both invaded areas as well as in the native areas, suggesting the existence of genetic lineages with a high capacity to establish in diverse environments and host species. This study confirmed the importance of multiple introductions, but questioned the role of genetic admixture in the success of introduction of a fungal plant pathogen.

Phylogeography and population structure of the grape powdery mildew fungus, Erysiphe necator, from diverse Vitis species

BackgroundThe grape powdery mildew fungus, Erysiphe necator, was introduced into Europe more than 160 years ago and is now distributed everywhere that grapes are grown. To understand the invasion history of this pathogen we investigated the evolutionary relationships between introduced populations of Europe, Australia and the western United States (US) and populations in the eastern US, where E. necator is thought to be native. Additionally, we tested the hypothesis that populations of E. necator in the eastern US are structured based on geography and Vitis host species.ResultsWe sequenced three nuclear gene regions covering 1803 nucleotides from 146 isolates of E. necator collected from the eastern US, Europe, Australia, and the western US. Phylogeographic analyses show that the two genetic groups in Europe represent two separate introductions and that the genetic groups may be derived from eastern US ancestors. Populations from the western US and Europe share haplotypes, suggesting that the western US population was introduced from Europe. Populations in Australia are derived from European populations. Haplotype richness and nucleotide diversity were significantly greater in the eastern US populations than in the introduced populations. Populations within the eastern US are geographically differentiated; however, no structure was detected with respect to host habitat (i.e., wild or cultivated). Populations from muscadine grapes, V. rotundifolia, are genetically distinct from populations from other Vitis host species, yet no differentiation was detected among populations from other Vitis species.ConclusionsMultilocus sequencing analysis of the grape powdery mildew fungus is consistent with the hypothesis that populations in Europe, Australia and the western US are derived from two separate introductions and their ancestors were likely from native populations in the eastern US. The invasion history of E. necator follows a pattern consistent with plant-mediated dispersal, however, more exhaustive sampling is required to make more precise conclusions as to origin. E. necator shows no genetic structure across Vitis host species, except with respect to V. rotundifolia.

Molecular Characterization of Vegetative Incompatibility Genes that Restrict Hypovirus Transmission in the Chestnut Blight Fungus Cryphonectria parasitica

Genetic nonself recognition systems such as vegetative incompatibility operate in many filamentous fungi to regulate hyphal fusion between genetically dissimilar individuals and to restrict the spread of virulence-attenuating mycoviruses that have potential for biological control of pathogenic fungi. We report here the use of a comparative genomics approach to identify seven candidate polymorphic genes associated with four vegetative incompatibility (vic) loci of the chestnut blight fungus Cryphonectria parasitica. Disruption of candidate alleles in one of two strains that were heteroallelic at vic2, vic6, or vic7 resulted in enhanced virus transmission, but did not prevent barrage formation associated with mycelial incompatibility. Detailed characterization of the vic6 locus revealed the involvement of nonallelic interactions between two tightly linked genes in barrage formation, heterokaryon formation, and asymmetric, gene-specific influences on virus transmission. The combined results establish molecular identities of genes associated with four C. parasitica vic loci and provide insights into how these recognition factors interact to trigger incompatibility and restrict virus transmission.