Bruce A. McDonald

Emergence of a new disease as a result of interspecific virulence gene transfer

New diseases of humans, animals and plants emerge regularly. Enhanced virulence on a new host can be facilitated by the acquisition of novel virulence factors. Interspecific gene transfer is known to be a source of such virulence factors in bacterial pathogens (often manifested as pathogenicity islands in the recipient organism) and it has been speculated that interspecific transfer of virulence factors may occur in fungal pathogens. Until now, no direct support has been available for this hypothesis. Here we present evidence that a gene encoding a critical virulence factor was transferred from one species of fungal pathogen to another. This gene transfer probably occurred just before 1941, creating a pathogen population with significantly enhanced virulence and leading to the emergence of a new damaging disease of wheat.

The population genetics of plant pathogens and breeding strategies for durable resistance

The durability of disease resistance is affected by the evolutionary potential of the pathogen population. Pathogens with a high evolutionary potential are more likely to overcome genetic resistance than pathogens with a low evolutionary potential. We will propose a set of guidelines to predict the evolutionary potential of pathogen populations based on analysis of their genetic structure. Under our model of pathogen evolution, the two most important parameters to consider are reproduction/mating system and gene/genotype flow. Pathogens that pose the greatest risk of breaking down resistance genes are those that possess a mixed reproduction system, with at least one sexual cycle per growing season and asexual reproduction during the epidemic phase, and a high potential for gene flow. The lowest risk pathogens are those with strict asexual reproduction and low potential for gene flow. We will present examples of high- and low-risk pathogens. Knowledge of the population genetic structure of the pathogen may offer insight into the best breeding strategy for durable resistance. We will present broad guidelines suggesting a rational method for breeding durable resistance according to the population genetics of the pathogen.

The Origins of Plant Pathogens in Agro-Ecosystems

Plant pathogens can emerge in agricultural ecosystems through several mechanisms, including host-tracking, host jumps, hybridization and horizontal gene transfer. High-throughput DNA sequencing coupled with new analytical approaches make it possible to differentiate among these mechanisms and to infer the time and place where pathogens first emerged. We present several examples to illustrate the different mechanisms and timescales associated with the origins of important plant pathogens. In some cases pathogens were domesticated along with their hosts during the invention of agriculture approximately 10,000 years ago. In other cases pathogens appear to have emerged very recently and almost instantaneously following horizontal gene transfer or hybridization. The predominant unifying feature in these examples is the environmental and genetic uniformity of the agricultural ecosystem in which the pathogens emerged. We conclude that agro-ecosystems will continue to select for new pathogens unless they are re-engineered to make them less conducive to pathogen emergence.

The Population Genetics of Fungi: Tools and Techniques

Over the last 10 years plant pathologists have begun to realize that more knowledge about the genetic structure of populations of plant pathogens is needed to implement effective control strategies (48). Research on the genetic structure of fungal populations has mushroomed, and review papers that summarize these studies are numerous (7,27,33,34,38). Although the number of fungal studies has increased greatly, the most comprehensive work has focused on a small number of plant-pathogenic fungi. The majority of these fungi can be recognized easily by their fruiting bodies or disease symptoms on aboveground plant parts. It has proven more difficult to assess the genetic structure of fungal populations that exist mainly belowground, because the distribution of individuals cannot be visualized directly and appropriate sampling procedures are less obvious and more cumbersome. Nevertheless, substantial progress has been made in interpreting the population genetic structure of some soilborne fungi (1,17). The purpose of this paper is to provide an overview of the tools and techniques of fungal population genetics. I will try to emphasize approaches that may be applied to studies of soilborne fungi. Instead of providing detailed methods, I will cite recent references where appropriate. There are many opinions regarding which techniques and tools are best suited to studies of fungal populations. I will give a personal and biased viewpoint, which I believe will be most useful to those who are just entering the field.

The global genetic structure of the wheat pathogen Mycosphaerella graminicola is characterized by high nuclear diversity, low mitochondrial diversity, regular recombination, and gene flow

A total of 1673 Mycosphaerella graminicola strains were assayed for DNA fingerprints and restriction fragment length polymorphism (RFLP) markers in the nuclear and mitochondrial genomes. The isolates were collected from 17 wheat fields located in 11 countries on five continents over a six year period (1989-1995). Our results indicate that genetic diversity in the nuclear genome of this fungus was high for all but three of the field populations surveyed and that populations sampled from different continents had similar frequencies for the most common RFLP alleles. Hierarchical analysis revealed that more than 90% of global gene diversity was distributed within a wheat field, while approximately 5% of gene diversity was distributed among fields within regions and approximately 3% was distributed among regions on different continents. These findings suggest that gene flow has occurred on a global scale. On average, each leaf was colonized by a different nuclear genotype. In contrast, only seven mtDNA haplotypes were detected among the 1673 isolates and the two most common mtDNA haplotypes represented approximately 93% of the world population, consistent with a selective sweep. Analysis of multilocus associations indicated that all field populations were in gametic equilibrium, suggesting that sexual recombination is a regular occurrence globally.

Dothideomycete–Plant Interactions Illuminated by Genome Sequencing and EST Analysis of the Wheat Pathogen Stagonospora nodorum

Stagonospora nodorum is a major necrotrophic fungal pathogen of wheat (Triticum aestivum) and a member of the Dothideomycetes, a large fungal taxon that includes many important plant pathogens affecting all major crop plant families. Here, we report the acquisition and initial analysis of a draft genome sequence for this fungus. The assembly comprises 37,164,227 bp of nuclear DNA contained in 107 scaffolds. The circular mitochondrial genome comprises 49,761 bp encoding 46 genes, including four that are intron encoded. The nuclear genome assembly contains 26 classes of repetitive DNA, comprising 4.5% of the genome. Some of the repeats show evidence of repeat-induced point mutations consistent with a frequent sexual cycle. ESTs and gene prediction models support a minimum of 10,762 nuclear genes. Extensive orthology was found between the polyketide synthase family in S. nodorum and Cochliobolus heterostrophus, suggesting an ancient origin and conserved functions for these genes. A striking feature of the gene catalog was the large number of genes predicted to encode secreted proteins; the majority has no meaningful similarity to any other known genes. It is likely that genes for host-specific toxins, in addition to ToxA, will be found among this group. ESTs obtained from axenic mycelium grown on oleate (chosen to mimic early infection) and late-stage lesions sporulating on wheat leaves were obtained. Statistical analysis shows that transcripts encoding proteins involved in protein synthesis and in the production of extracellular proteases, cellulases, and xylanases predominate in the infection library. This suggests that the fungus is dependant on the degradation of wheat macromolecular constituents to provide the carbon skeletons and energy for the synthesis of proteins and other components destined for the developing pycnidiospores.

Population Structure of Mycosphaerella graminicola: From Lesions to Continents

ABSTRACT The genetic structure of field populations of Mycosphaerella graminicola was determined across a hierarchy of spatial scales using restriction fragment length polymorphism markers. The hierarchical gene diversity analysis included 1,098 isolates from seven field populations. Spatial scales ranged from millimeters to thousands of kilometers, including comparisons within and among lesions, within and among fields, and within and among regions and continents. At the smallest spatial scale, microtransect sampling was used to determine the spatial distribution of 15 genotypes found among 158 isolates sampled from five individual lesions. Each lesion had two to six different genotypes including both mating types in four of the five lesions, but in most cases a lesion was composed of one or two genotypes that occupied the majority of the lesion, with other rare genotypes interspersed among the common genotypes. The majority (77%) of gene diversity was distributed within plots ranging from approximately 1 to 9 m(2) in size. Genotype diversity (G / N) within fields for the Swiss, Texas, and Israeli fields was high, ranging from 79 to 100% of maximum possible values. Low population differentiation was indicated by the low G(ST) values among populations, suggesting a corresponding high degree of gene flow among these populations. At the largest spatial scale, populations from Switzerland, Israel, Oregon, and Texas were compared. Population differentiation among these populations was low (G(ST) = 0.05), and genetic identity between populations was high. A low but significant correlation between genetic and geographic distance among populations was found (r = -0.47, P = 0.012), suggesting that these populations probably have not reached an equilibrium between gene flow and genetic drift. Gene flow on a regional level can be reduced by implementing strategies, such as improved stubble management that minimize the production of ascospores. The possibility of high levels of gene flow on a regional level indicates a significant potential risk for the regional spread of mutant alleles that enable fungicide resistance or the breakdown of resistance genes.

An analysis of the durability of resistance to plant viruses

ABSTRACT Genetic resistance often fails because a resistance-breaking (RB) pathogen genotype increases in frequency. On the basis of an analysis of cellular plant pathogens, it was recently proposed that the evolutionary potential of a pathogen is a major determinant of the durability of resistance. We test this hypothesis for plant viruses, which differ substantially from cellular pathogens in the nature, size, and expression of their genomes. Our analysis was based on 29 plant virus species that provide a good representation of the genetic and biological diversity of plant viruses. These 29 viruses were involved in 35 pathosystems, and 50 resistance factors deployed against them were analyzed. Resistance was found to be durable more often than not, in contrast with resistance to cellular plant pathogens. In a third of the analyzed pathosystems RB strains have not been reported, and in another third RB strains have been reported but have not become prevalent in the virus population. The evolutionary potential of the viruses in the 35 pathosystems was evaluated with a compound risk index based on three evolutionary factors: the population of the pathogen, the degree of recombination, and the amount of gene and genotype flow. Our analysis indicates that evolutionary potential may be an important determinant of the durability of resistance against plant viruses.

QoI resistance emerged independently at least 4 times in European populations of Mycosphaerella graminicola

BACKGROUND QoI fungicides or quinone outside inhibitors (also called strobilurins) have been widely used to control agriculturally important fungal pathogens since their introduction in 1996. Strobilurins block the respiration pathway by inhibiting the cytochrome bc1 complex in mitochondria. Several plant pathogenic fungi have developed field resistance. The first QoI resistance in Mycosphaerella graminicola (Fuckel) Schroter was detected retrospectively in UK in 2001 at a low frequency in QoI-treated plots. During the following seasons, resistance reached high frequencies across northern Europe. The aim of this study was to identify the main evolutionary forces driving the rapid emergence and spread of QoI resistance in M. graminicola populations. RESULTS The G143A mutation causing QoI resistance was first detected during 2002 in all tested populations and in eight distinct mtDNA sequence haplotypes. By 2004, 24 different mtDNA haplotypes contained the G143A mutation. Phylogenetic analysis showed that strobilurin resistance was acquired independently through at least four recurrent mutations at the same site of cytochrome b. Estimates of directional migration rates showed that the majority of gene flow in Europe had occurred in a west-to-east direction. CONCLUSION This study demonstrated that recurring mutations independently introduced the QoI resistance allele into different genetic and geographic backgrounds. The resistant haplotypes then increased in frequency owing to the strong fungicide selection and spread eastward through wind dispersal of ascospores.

Genetic Structure of Rhynchosporium secalis in Australia

ABSTRACT Restriction fragment length polymorphism (RFLP) markers were used to determine the genetic structure of Australian field populations of the barley scald pathogen Rhynchosporium secalis. Fungal isolates were collected by hierarchical sampling from five naturally infected barley fields in different geographic locations during a single growing season. Genetic variation was high in Australian R. secalis populations. Among the 265 fungal isolates analyzed, 214 distinct genotypes were identified. Average genotype diversity within a field population was 65% of its theoretical maximum. Neis average gene diversity across seven RFLP loci was 0.54. The majority (76%) of gene diversity was distributed within sampling site areas measuring 1 m(2); 19% of gene diversity was distributed among sampling sites within fields; and 5% of gene diversity was distributed among fields. Fungal populations from different locations differed significantly both in allele frequencies and genotype diversities. The degree of genetic differentiation was significantly correlated with geographic distance between populations. Our results suggest that the R. secalis population in Western Australia has a different genetic structure than populations in Victoria and South Australia.