Promise and Pitfalls of Endemic Resistance for Cultural Resources Threatened by Phytophthora ramorum.

Invasive forest pathogens can harm cultural, economic, and ecological resources. Here, we demonstrate the potential of endemic tree pathogen resistance in forest disease management using Phytophthora ramorum, cause of sudden oak death, in the context of management of tanoak (Notholithocarpus densiflorus), an ecologically unique and highly valued tree within Native American communities of northern California and southern Oregon in the United States. We surveyed resistance to P. ramorum on the Hoopa Valley Indian Reservation and Yurok Indian Reservation in a set of study sites with variable management intensities. Variation in resistance was found at all sites with similar mean and variation across stands, and resistance tended to have a random spatial distribution within stands but was not associated with previous stand management (thinning or prescribed fire) or structural characteristics such as tree density, basal area, or pairwise relatedness among study trees. These results did not suggest host, genetic, management, or environment interactions that could be easily leveraged into treatments to increase the prevalence of resistant trees. We applied epidemiological models to assess the potential application of endemic resistance in this system and to examine our assumption that in planta differences in lesion size-our measure of resistance-reflect linkages between mortality and transmission (resistance) versus reduced mortality with no change in transmission (tolerance). This assumption strongly influenced infection dynamics but changes in host populations-our conservation focus-was dependent on community-level variation in transmission. For P. ramorum, slowing mortality rates (whether by resistance or tolerance) conserves host resources when a second source of inoculum is present; these results are likely generalizable to pathogens with a broader host range. However, when the focal host is the sole source of inoculum, increasing tolerant individuals led to the greatest stand-level pathogen accumulation in our model. When seeking to use variation in mortality rates to affect conservation strategies, it is important to understand how these traits are linked with transmission because tolerance will be more useful for management in mixed-host stands that are already invaded, compared with single-host stands with low or no pathogen presence, where resistance will have the greatest conservation benefits.

The escalatory Red Queen: Population extinction and replacement following arms race dynamics in poplar rust.

Host-parasite systems provide convincing examples of Red Queen co-evolutionary dynamics. Yet, a key process underscored in Van Valen's theory - that arms race dynamics can result in extinction - has never been documented. One reason for this may be that most sampling designs lack the breadth needed to illuminate the rapid pace of adaptation by pathogen populations. In this study, we used a 25-year temporal sampling to decipher the demographic history of a plant pathogen: the poplar rust fungus, Melampsora larici-populina. A major adaptive event occurred in 1994 with the breakdown of R7 resistance carried by several poplar cultivars widely planted in Western Europe since 1982. The corresponding virulence rapidly spread in M. larici-populina populations and nearly reached fixation in northern France, even on susceptible hosts. Using both temporal records of virulence profiles and temporal population genetic data, our analyses revealed that (i) R7 resistance breakdown resulted in the emergence of a unique and homogeneous genetic group, the so-called cultivated population, which predominated in northern France for about 20 years, (ii) selection for Vir7 individuals brought with it multiple other virulence types via hitchhiking, resulting in an overall increase in the population-wide number of virulence types and (iii) - above all - the emergence of the cultivated population superseded the initial population which predominated at the same place before R7 resistance breakdown. Our temporal analysis illustrates how antagonistic co-evolution can lead to population extinction and replacement, hence providing direct evidence for the escalation process which is at the core of Red Queen dynamics.

Sudden oak death: interactions of the exotic oomycete Phytophthora ramorum with naïve North American hosts.

Ten years after a threatening and previously unknown disease of oaks and tanoaks appeared in coastal California, a significant amount of progress has been made toward the understanding of its causal agent Phytophthora ramorum and of the novel pathosystems associated with this exotic organism. However, a complete understanding of the ecology and epidemiology of this species still eludes us. In part, our inability to fully understand this organism is due to its phylogenetic, phylogeographic, phenotypic, and epidemiological complexities, all reviewed in this paper. Most lines of evidence suggest that the high degree of disease severity reported in California is not simply due to a generalized lack of resistance or tolerance in naïve hosts but also to an innate ability of the pathogen to survive in unfavorable climatic conditions and to reproduce rapidly when conditions become once again favorable.

Innate Resistance and Phosphite Treatment Affect Both the Pathogen's and Host's Transcriptomes in the Tanoak-Phytophthora ramorum Pathosystem.

Phosphites have been used to control Sudden Oak Death; however, their precise mode of action is not fully understood. To study the mechanism of action of phosphites, we conducted an inoculation experiment on two open-pollinated tanoak families, previously found to be partially resistant. Stems of treatment group individuals were sprayed with phosphite, and seven days later, distal leaves were inoculated with the Sudden Oak Death pathogen Phytophthora ramorum. Leaves from treated and untreated control plants were harvested before and seven days after inoculation, and transcriptomes of both host and pathogen were analyzed. We found that tanoak families differed in the presence of innate resistance (resistance displayed by untreated tanoak) and in the response to phosphite treatment. A set of expressed genes associated with innate resistance was found to overlap with an expressed gene set for phosphite-induced resistance. This observation may indicate that phosphite treatment increases the resistance of susceptible host plants. In addition, genes of the pathogen involved in detoxification were upregulated in phosphite-treated plants compared to phosphite-untreated plants. In summary, our RNA-Seq analysis supports a two-fold mode of action of phosphites, including a direct toxic effect on P. ramorum and an indirect enhancement of resistance in the tanoak host.

Evolution of morphological but not aggressiveness-related traits following a major resistance breakdown in the poplar rust fungus, Melampsora larici-populina.

Crop varieties carrying qualitative resistance to targeted pathogens lead to strong selection pressure on parasites, often resulting in resistance breakdown. It is well known that qualitative resistance breakdowns modify pathogen population structure but few studies have analyzed the consequences on their quantitative aggressiveness-related traits. The aim of this study was to characterize the evolution of these traits following a resistance breakdown in the poplar rust fungus, Melampsora larici-populina. We based our experiment on three temporal populations sampled just before the breakdown event, immediately after and four years later. First, we quantified phenotypic differences among populations for a set of aggressiveness traits on a universally susceptible cultivar (infection efficiency, latent period, lesion size, mycelium quantity, and sporulation rate) and one morphological trait (mean spore volume). Then, we estimated heritability to establish which traits could be subjected to adaptive evolution and tested for evidence of selection. Our results revealed significant changes in the morphological trait but no variation in aggressiveness traits. By contrast, recent works have demonstrated that quantitative resistance (initially assumed more durable) could be eroded and lead to increased aggressiveness. Hence, this study is one example suggesting that the use of qualitative resistance may be revealed to be less detrimental to long-term sustainable crop production.

Genetic signatures of a range expansion in natura: when clones play leapfrog.

The genetic consequences of range expansions have generally been investigated at wide geographical and temporal scales, long after the colonization event. A unique ecological system enabled us to both monitor the colonization dynamics and decipher the genetic footprints of expansion over a very short time period. Each year an epidemic of the poplar rust (Melampsora larici-populina) expands clonally and linearly along the Durance River, in the Alps. The colonization dynamics observed in 2004 showed two phases with different genetic outcomes. Upstream, fast colonization maintained high genetic diversity. Downstream, the colonization wave progressively faltered, diversity eroded, and differentiation increased, as expected under recurrent founder events. In line with the high dispersal abilities of rust pathogens, we provide evidence for leapfrog dispersal of clones. Our results thus emphasize the importance of colonization dynamics in shaping spatial genetic structure in the face of high gene flow.

Detection and Quantification of Phytophthora ramorum from California Forests Using a Real-Time Polymerase Chain Reaction Assay.

ABSTRACT The timely and accurate detection of pathogens is a critical aid in the study of the epidemiology and biology of plant diseases. In the case of regulated organisms, the availability of a sensitive and reliable assay is essential when trying to achieve early detection of the pathogen. We developed and tested a real-time, nested polymerase chain reaction (PCR) assay for the detection of Phytophthora ramorum, causal agent of sudden oak death. This technique then was implemented as part of a widespread environmental screen throughout California. The method here described is sensitive, detecting less than 12 fg of pathogen DNA, and is specific for P. ramorum when tested across 21 Phytophthora spp. Hundreds of symptomatic samples from 33 sites in 14 California counties were assayed, resulting in the discovery of 10 new host species and 23 infested areas, including 4 new counties. With the exception of a single host, PCR-based discovery of new hosts and infested areas always was confirmed by traditional pathogen isolations and inoculation studies. Nonetheless, molecular diagnostics were key in early pathogen detection, and steered the direction of further research on this newly discovered and generalist Phytophthora species.

Scaling up from greenhouse resistance to fitness in the field for a host of an emerging forest disease.

Forest systems are increasingly threatened by emergent, exotic diseases, yet management strategies for forest trees may be hindered by long generation times and scant background knowledge. We tested whether nursery disease resistance and growth traits have predictive value for the conservation of Notholithocarpus densiflorus, the host most susceptible to sudden oak death. We established three experimental populations to assess nursery growth and resistance to Phytophthora ramorum, and correlations between nursery-derived breeding values with seedling survival in a field disease trial. Estimates of nursery traits' heritability were low to moderate, with lowest estimates for resistance traits. Within the field trial, survival likelihood was increased in larger seedlings and decreased with the development of disease symptoms. The seed-parent family wide likelihood of survival was likewise correlated with family predictors for size and resistance to disease in 2nd year laboratory assays, though not resistance in 1st year leaf assays. We identified traits and seedling families with increased survivorship in planted tanoaks, and a framework to further identify seed parents favored for restoration. The additive genetic variation and seedling disease dynamics we describe hold promise to refine current disease models and expand the understanding of evolutionary dynamics of emergent infectious diseases in highly susceptible hosts.

AFLP and phylogenetic analyses of North American and European populations of Phytophthora ramorum.

The genetic structure within and between USA and European populations of the emerging phytopathogen Phytophthora ramorum was examined. Four primer combinations were used for amplified fragment length polymorphism (AFLP) fingerprinting of 67 USA isolates from California and Oregon, and 18 European isolates from Belgium, Germany, The Netherlands, Spain and the UK. In addition, three DNA regions (ITS, cox II, and nad 5) of additional Phytophthora species were amplified by polymerase chain reaction, sequenced, and analysed to provide better phylogenetic understanding of P. ramorum within the genus Phytophthora. AFLP banding patterns indicate that the 85 isolates form two distinct lineages within a monophyletic group, distinct from the closely related outgroup species P. lateralis. With the exception of two isolates from an Oregon nursery, European and USA isolates clustered separately within individual clades. The AFLP profiles also indicate that a single clonal lineage dominates the North American population, while the European population consists of an array of mainly unique, closely related AFLP types. Sequences from the three DNA regions were identical among all P. ramorum isolates, and phylogenetic analysis indicates that P. ramorum is closely related to P. lateralis and P. hibernalis.