Transcriptome Analysis of Poplar during Leaf Spot Infection with Sphaerulina spp.
ABSTRACT: Diseases of poplar caused by the native fungal pathogen Sphaerulina musiva and related species are of growing concern, particularly with the increasing interest in intensive poplar plantations to meet growing energy demands. Sphaerulina musiva is able to cause infection on leaves, resulting in defoliation and canker formation on stems. To gain a greater understanding of the different responses of poplar species to infection caused by the naturally co-evolved Sphaerulina species, RNA-seq was conducted on leaves of Populus deltoides, P. balsamifera and P. tremuloides infected with S. musiva, S. populicola and a new undescribed species, Ston1, respectively. The experiment was designed to contain the pathogen in a laboratory environment, while replicating disease development in commercial plantations. Following inoculation, trees were monitored for disease symptoms, pathogen growth and host responses. Genes involved in phenylpropanoid, terpenoid and flavonoid biosynthesis were generally upregulated in P. balsamifera and P. tremuloides, while cell wall modification appears to play an important role in the defense of P. deltoides. Poplar defensive genes were expressed early in P. balsamifera and P. tremuloides, but their expression was delayed in P. deltoides, which correlated with the rate of disease symptoms development. Also, severe infection in P. balsamifera led to leaf abscission. This data gives an insight into the large differences in timing and expression of genes between poplar species being attacked by their associated Sphaerulina pathogen.
Project description:Diseases of poplar caused by the fungal pathogen Sphaerulina musiva and related species are of growing concern, particular with the increasing interest in intensive popluliculture to meet increasing energy demands. S. musiva is able to cause infection on leaves, resulting in defoliation and canker formation on stems. To gain a greater understanding of the different responses of poplar species to infection with their natural Sphaerulina species, RNA-seq was conducted on leaves of Populus deltoides, P. balsamifera and P. tremuloides infected with S. musiva, S. populicola and a new undescribed species Ston1, respectively. Progression of disease symptoms, pathogen growth and host response were detected. Through the time course of infection, different and species-specific metabolic pathways were activated. In all three species, genes associated with growth and development were down-regulated, while genes involved the phenylpropanoid, terpenoid and flavonoid biosynthesis were up-regulated. Poplar defensive genes were expressed early in P. balsamifera and P. tremuloides, but delayed in P. deltoides, which correlated with the rate of disease symptoms development. This data gives an insight into the large differences in timing and expression of genes between poplar species being attacked with their native associated Sphaerulina pathogen. RNA-seq was conducted on leaves of Populus deltoides, P. balsamifera and P. tremuloides infected with S. musiva, S. populicola and a new undescribed species Ston1, respectively.
Project description:Hybrid genotypes that arise between plant species frequently have increased susceptibility to arthropod pests and fungal pathogens. This pattern has been attributed to the breakdown of plant defenses ('Hybrid susceptibility' hypothesis) and (or) to extended periods of susceptibility attributed to plant phenologies in zones of species overlap and (or) hybridization ('phenological sink' hypothesis). We examined these hypotheses by assessing the susceptibility of parental and hybrid Populus host genotypes to a leaf spot disease caused by the fungal pathogen Septoria musiva. For this purpose, 214 genotypes were obtained from morphologically pure zones of P. balsamifera and P. deltoides, and from an intervening zone of overlap and hybridization on the drainage of the Red Deer River, Alberta, Canada. Genotypes were identified as P. balsamifera, P. deltoides, or hybrid using a suite of 27 species-specific SNP markers. Initially the genetic structure of the hybrid zone was characterized with 27.7% of trees classified as admixed individuals. To test the hybrid susceptibility hypothesis, a subset of 52 genotypes was inoculated with four isolates of S. musiva. Levels of susceptibility were P. balsamifera > F1 hybrid > P. deltoides. A further 53 genotypes were grown in a common garden to assess the effect of genotype on variation in leaf phenology. Leaf phenology was more variable within the category of hybrid genotypes than within categories of either parental species. Leaf phenology was also more variable for the category of trees originating in the hybrid (P. balsamifera - P. deltoides [hybrid and parental genotypes combined]) zone than in adjacent pure zones of the parental species. The results from the inoculation experiment support the hybrid intermediacy hypothesis. The results from the common garden experiment support the 'phenological sink' hypothesis. These findings have greatly increased our understanding of the epidemiology and ecology of fungal pathogens in plant hybrid zones.
Project description:Trees bearing novel or exotic gene components are poised to contribute to the bioeconomy for a variety of purposes such as bioenergy production, phytoremediation, and carbon sequestration within the forestry sector, but sustainable release of trees with novel traits in large-scale plantations requires the quantification of risks posed to native tree populations. Over the last century, exotic hybrid poplars produced through artificial crosses were planted throughout eastern Canada as ornamentals or windbreaks and these exotics provide a proxy by which to examine the fitness of exotic poplar traits within the natural environment to assess risk of exotic gene escape, establishment, and spread into native gene pools. We assessed postzygotic fitness traits of native and exotic poplars within a naturally regenerated stand in eastern Canada (Quebec City, QC). Pure natives (P. balsamifera and P. deltoides spp. deltoides), native hybrids (P. deltoides × P. balsamifera), and exotic hybrids (trees bearing Populus nigra and P. maximowiczii genetic components) were screened for reproductive biomass, yield, seed germination, and fungal disease susceptibility. Exotic hybrids expressed fitness traits intermediate to pure species and were not significantly different from native hybrids. They formed fully viable seed and backcrossed predominantly with P. balsamifera. These data show that exotic hybrids were not unfit and were capable of establishing and competing within the native stand. Future research will seek to examine the impact of exotic gene regions on associated biotic communities to fully quantify the risk exotic poplars pose to native poplar forests.
Project description:In this article, we describe the presence of genes encoding close homologues of an endogenous plant peptide, rapid alkalinization factor (RALF), within the genomes of 26 species of phytopathogenic fungi. Members of the RALF family are key growth factors in plants, and the sequence of the RALF active region is well conserved between plant and fungal proteins. RALF1-like sequences were observed in most cases; however, RALF27-like sequences were present in the Sphaerulina musiva and Septoria populicola genomes. These two species are pathogens of poplar and, interestingly, the closest relative to their respective RALF genes is a poplar RALF27-like sequence. RALF peptides control cellular expansion during plant development, but were originally defined on the basis of their ability to induce rapid alkalinization in tobacco cell cultures. To test whether the fungal RALF peptides were biologically active in plants, we synthesized RALF peptides corresponding to those encoded by two sequenced genomes of the tomato pathogen Fusarium oxysporum f. sp. lycopersici. One of these peptides inhibited the growth of tomato seedlings and elicited responses in tomato and Nicotiana benthamiana typical of endogenous plant RALF peptides (reactive oxygen species burst, induced alkalinization and mitogen-activated protein kinase activation). Gene expression analysis confirmed that a RALF-encoding gene in F. oxysporum f. sp. lycopersici was expressed during infection on tomato. However, a subsequent reverse genetics approach revealed that the RALF peptide was not required by F. oxysporum f. sp. lycopersici for infection on tomato roots. This study has demonstrated the presence of functionally active RALF peptides encoded within phytopathogens that harbour an as yet undetermined role in plant-pathogen interactions.
Project description:Hybridization and introgression are pervasive evolutionary phenomena that provide insight into the selective forces that maintain species boundaries, permit gene flow, and control the direction of evolutionary change. Poplar trees (Populus L.) are well known for their ability to form viable hybrids and maintain their distinct species boundaries despite this interspecific gene flow. We sought to quantify the hybridization dynamics and postzygotic fitness within a hybrid stand of balsam poplar (Populus balsamifera L.), eastern cottonwood (P. deltoides Marsh.), and their natural hybrids to gain insight into the barriers maintaining this stable hybrid zone. We observed asymmetrical hybrid formation with P. deltoides acting as the seed parent, but with subsequent introgression biased toward P. balsamifera. Native hybrids expressed fitness traits intermediate to the parental species and were not universally unfit. That said, native hybrid seedlings were absent from the seedling population, which may indicate additional selective pressures controlling their recruitment. It is imperative that we understand the selective forces maintaining this native hybrid zone in order to quantify the impact of exotic poplar hybrids on this native system.
Project description:Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulose-degrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems.
Project description:Four highly inducible genes of poplar trees, PtdKTI5, PtdWIN4, PtdPOP3 from hybrid poplar (Populus trichocarpa × P. deltoides) and PtKTI2 from trembling aspen (Populus tremuloides Michx.) have been individually transformed into Arabidopsis thaliana for overexpression. High transcriptional level of each transgene in transgenic Arabidopsis lines was confirmed by RT-PCR analysis. The development, body weight and survivorship of cotton bollworm (Helicoverpa armigera) fed on four types of transgenic Arabidopis plants were evaluated in the laboratory. Our data indicated that these four Populus defense-related genes exhibited various degree of insectital activity on larval and postlarval development of cotton bollworm and may be utilized for herbivore resistance improvement in plant genetic engineering.
Project description:Invasive microbes causing diseases such as sudden oak death negatively affect ecosystems and economies around the world. The deployment of resistant genotypes for combating introduced diseases typically relies on breeding programs that can take decades to complete. To demonstrate how this process can be accelerated, we employed a genome-wide association mapping of ca 1,000 resequenced Populus trichocarpa trees individually challenged with Sphaerulina musiva, an invasive fungal pathogen. Among significant associations, three loci associated with resistance were identified and predicted to encode one putative membrane-bound L-type receptor-like kinase and two receptor-like proteins. A susceptibility-associated locus was predicted to encode a putative G-type D-mannose-binding receptor-like kinase. Multiple lines of evidence, including allele analysis, transcriptomics, binding assays, and overexpression, support the hypothesized function of these candidate genes in the P. trichocarpa response to S. musiva.
Project description:Schizoempodium mesophyllincola is an eriophyid mite that feeds in leaves of Populus trichocarpa in the central part of this cottonwood tree's range (i.e., coastal British Columbia, Washington and Oregon) in the Pacific Northwest (PNW) of North America, and on some interspecific hybrids planted in short-rotation, intensive forestry in the region. The mite, a leaf vagrant, sucks the contents of spongy mesophyll cells, causing leaf discoloration, or "bronzing." Here, we investigate the inheritance pattern of resistance to leaf bronzing using a three-generation Populus trichocarpa x P. deltoides hybrid pedigree. We found that resistance to the mite is an exaptation in that its source in two related F2 families of the TxD hybrid pedigree was the non-native host, P. deltoides. Two grandparental genotypes of the latter, 'ILL-5' and 'ILL-129', were completely free of the bronzing symptom and that phenotype was inherited in a Mendelian manner in the F1 and F2. Resistance to S. mesophyllincola is similar to resistance to many other regional pathogens of P. trichocarpa (e.g., Melampsora occidentalis, Venturia inopina, Sphaerulina populicola, and Taphrina sp.) in that it is inherited from the non-native grandparent (e.g., P. deltoides, P. nigra, or P. maximowiczii) in three-generation, hybrid pedigrees. In addition to finding evidence for Mendelian inheritance, we found two QTLs with LOD scores 5.03 and 3.12 mapped on linkage groups (LG) III and I, and they explained 6.7 and 4.2% of the phenotypic variance, respectively. The LG I QTL is close to, or synonymous with, one for resistance to sap-feeding arthropods and leaf developmental traits as expressed in a British study utilizing the same pedigree.
Project description:The deposition of antimicrobial plant resins in honey bee, Apis mellifera, nests has important physiological benefits. Resin foraging is difficult to approach experimentally because resin composition is highly variable among and between plant families, the environmental and plant-genotypic effects on resins are unknown, and resin foragers are relatively rare and often forage in unobservable tree canopies. Subsequently, little is known about the botanical origins of resins in many regions or the benefits of specific resins to bees. We used metabolomic methods as a type of environmental forensics to track individual resin forager behavior through comparisons of global resin metabolite patterns. The resin from the corbiculae of a single bee was sufficient to identify that resin's botanical source without prior knowledge of resin composition. Bees from our apiary discriminately foraged for resin from eastern cottonwood (Populus deltoides), and balsam poplar (P. balsamifera) among many available, even closely related, resinous plants. Cottonwood and balsam poplar resin composition did not show significant seasonal or regional changes in composition. Metabolomic analysis of resin from 6 North American Populus spp. and 5 hybrids revealed peaks characteristic to taxonomic nodes within Populus, while antimicrobial analysis revealed that resin from different species varied in inhibition of the bee bacterial pathogen, Paenibacillus larvae. We conclude that honey bees make discrete choices among many resinous plant species, even among closely related species. Bees also maintained fidelity to a single source during a foraging trip. Furthermore, the differential inhibition of P. larvae by Populus spp., thought to be preferential for resin collection in temperate regions, suggests that resins from closely related plant species many have different benefits to bees.