Nonadditive gene expression is correlated with nonadditive phenotypic expression in interspecific triploid hybrids of willow (Salix spp.)

Many studies have highlighted the complex and diverse basis for heterosis in inbred crops. Despite the lack of a consensus model, it is vital that we turn our attention to understanding heterosis in undomesticated, heterozygous, and polyploid species, such as willow (Salix spp.). Shrub willow is a dedicated energy crop bred to be fast-growing and high yielding on marginal land without competing with food crops. A trend in willow breeding is the consistent pattern of heterosis in triploids produced from crosses between diploid and tetraploid species. Here, we test whether differentially expressed genes are associated with heterosis in triploid families derived from diploid S. purpurea, diploid S. viminalis, and tetraploid S. miyabeana parents. Three biological replicates of shoot tips from all family progeny and parents were collected after 12 weeks in the greenhouse and RNA extracted for RNA-Seq analysis. This study provides evidence that nonadditive patterns of gene expression are correlated with nonadditive phenotypic expression in interspecific triploid hybrids of willow. Expression-level dominance was most correlated with heterosis for biomass yield traits and was highly enriched for processes involved in starch and sucrose metabolism. In addition, there was a global dosage effect of parent alleles in triploid hybrids, with expression proportional to copy number variation. Importantly, differentially expressed genes between family parents were most predictive of heterosis for both field and greenhouse collected traits. Altogether, these data will be used to progress models of heterosis to complement the growing genomic resources available for the improvement of heterozygous perennial bioenergy crops.

Invertebrate and Plant Community Diversity of an Illinois Corn–Soybean Field with Integrated Shrub Willow Bioenergy Buffers

Perennial bioenergy crop production within intensively managed agricultural landscapes has the potential to improve the sustainability, resiliency, and diversity of these landscapes. Perennial crops are ideal because of their high production potential on marginal lands relative to grain crops (e.g., corn and soybean) and their ability to provide additional ecosystem service benefits. When agricultural landscapes are designed to target specific services, determining the non-targeted services of perennial bioenergy crops can further promote their adoption. This 3-year study addresses this proposition by evaluating the canopy invertebrates and understory plant (non-target crop) communities using bee bowls and point measurement of ground coverage, respectively, within a grain field integrated with shrub willow buffer systems designed for nutrient loss reduction. Greater plant diversity and richness were observed under willow than under grain, resembling that of the surrounding riparian community with more perennial, native species. However, the same relationship did not hold true for invertebrates, with seasonality having a significant influence resulting in similar communities observed in willow and grain plots. The presence of unique plant and invertebrate species in both willow and grain crops as well as foraging pollinators on both crop and non-target crop species highlights the importance of habitat heterogeneity for supporting biodiversity and the potential benefits of buffer bioenergy landscape designs.

Nonadditive gene expression is correlated with nonadditive phenotypic expression in interspecific triploid hybrids of willow (Salix spp.)

Many studies have highlighted the complex and diverse basis for heterosis in inbred crops. Despite the lack of a consensus model, it is vital that we turn our attention to understanding heterosis in undomesticated, heterozygous, and polyploid species, such as willow (Salix spp.). Shrub willow is a dedicated energy crop bred to be fast-growing and high yielding on marginal land without competing with food crops. A trend in willow breeding is the consistent pattern of heterosis in triploids produced from crosses between diploid and tetraploid species. Here, we test whether differentially expressed genes are associated with heterosis in triploid families derived from diploid S. purpurea, diploid S. viminalis, and tetraploid S. miyabeana parents. Three biological replicates of shoot tips from all family progeny and parents were collected after 12 weeks in the greenhouse and RNA extracted for RNA-Seq analysis. This study provides evidence that nonadditive patterns of gene expression are correlated with nonadditive phenotypic expression in interspecific triploid hybrids of willow. Expression-level dominance was most correlated with heterosis for biomass yield traits and was highly enriched for processes involved in starch and sucrose metabolism. In addition, there was a global dosage effect of parent alleles in triploid hybrids, with expression proportional to copy number variation. Importantly, differentially expressed genes between family parents were most predictive of heterosis for both field and greenhouse collected traits. Altogether, these data will be used to progress models of heterosis to complement the growing genomic resources available for the improvement of heterozygous perennial bioenergy crops.

The Melampsora americana population on Salix purpurea in the Great Lakes region is highly diverse with a contributory influence of clonality

Shrub willows (Salix spp.) are emerging as a viable lignocellulosic, second-generation bioenergy crop with many growth characteristics favorable for marginal lands in New York State and surrounding areas. Willow rust, caused by members of the genus Melampsora, is the most limiting disease of shrub willow in this region and remains extremely understudied. In this study, genetic diversity, genetic structure, and pathogen clonality were examined in Melampsora americana over two growing seasons using genotyping-by-sequencing to identify single nucleotide polymorphism markers. In conjunction with this project, a reference genome of rust isolate R15-033-03 was generated to aid in variant discovery. Sampling between years allowed for regional and site-specific investigation into population dynamics, in the context of both wild and cultivated hosts within high density plantings. This work revealed that this pathogen is largely panmictic over the sampled areas with few sites showing moderate genetic differentiation. This data supports the hypothesis of sexual recombination between growing seasons as no genotype persisted across the two years of sampling. Additionally, clonality was determined as a driver of pathogen populations within cultivated fields and single shrubs, however there is also evidence of high genetic diversity of rust isolates in all settings. Together, this work provides a framework for M. americana population structure in the Great Lakes region, providing crucial information that can aid in future resistance breeding efforts.

Phytomanagement of soil and groundwater at the Niagara Falls Storage Site (NFSS) using hybridized trees

The Manhattan Engineer District previously used the 191-acre Niagara Falls Storage Site (NFSS) in Niagara County, New York, to store radioactive residues and wastes from uranium (U) ore processing. At present, management practices will determine whether enhanced evapotranspiration rates produced by hybridized shrub willow cuttings planted in 2016 will affect groundwater hydrology. Two shrub willow varieties were planted in an approximately one-half acre area to examine growth performance along a U impacted sanitary sewer line. Additionally, control plots will compare the effectiveness of shrub willows to unplanted areas. Observations of the planted area after 18 months showed success of shrub willow growth with increasing biomass. Chemical analysis from tree tissue samples of the field study showed no significant uptake of U or thorium (Th) to date. A greenhouse study conducted in parallel to the field study tested the willows under controlled greenhouse conditions and evaluated their ability to grow and accumulate contaminants under controlled conditions. Results from the greenhouse study demonstrated that U accumulation was minimal. Thus, this study demonstrates that the shrub willows are not accumulators of U or Th, an advantageous characteristic that implies stabilized contaminants in the soil and no translocation of U into the aboveground biomass.

Plant growth and nutrient composition in shrub and arbor willows grown in Cu contaminated soil as affected by flooding

Flooding can adversely worsen the metal contaminated soil and plant growth thus, it is crucial to explore the ecophysiological responses of plants upon co-exposure to heavy metals and flooding. Here, the plant growth, photosynthesis, and nutrient elements composition in arbor willow (Salix jiangsuensis ‘J172’) and shrub willow (Salix integra ‘Yizhibi’) were studied using a pot experiment with Cu contaminated soil (239.51 mg∙kg-1) under flooded versus non flooded condition. Salix integra showed larger BCFs than Salix jiangsuensis in both treatments, soil flooding significantly decreased the Cu contents and BCF while obviously increased TF values in both willow species (p < 0.05). Soil flooding markedly enhanced the leaf C:P and N:P ratios, while significantly decreased root C:P and N:P ratios, as compared to non flooded condition. The shrub willow exhibited better tolerance to soil flooding with little alteration in biomass and photosynthetic rate, and showed greater potential of Cu accumulation capacity, even though its total biomass was significantly lower than arbor willow. Our study also helps further understanding the nutrient balance and stoichiometry of willows in Cu contaminated soil and their response to soil flooding, helping the management of Cu-contaminated flooded soils.

Comparative Transcriptomics And eQTL Mapping Of Response To Melampsora Americana In Selected Salix Purpurea F2 Progeny

Background: Melampsora spp. rusts are the greatest pathogen threat to shrub willow (Salix spp.) bioenergy crops. Genetic resistance is key to limit the effects of these foliar diseases on host response and biomass yield, however, the genetic basis of host resistance has not been characterized. The addition of new genomic resources for Salix provides greater power to investigate the interaction between S. purpurea and M. americana, species commonly found in the Northeast US. Here, we utilize 3' RNA-seq to investigate host-pathogen interactions following controlled inoculations of M. americana on resistant and susceptible F2 S. purpurea genotypes identified in a recent QTL mapping study. Differential gene expression, network analysis, and eQTL mapping was used to contrast the response to inoculation and to identify associated candidate genes.Results: Controlled inoculation in a replicated greenhouse study identified 19 and 105 differentially expressed genes between resistant and susceptible genotypes at 42 and 66 HPI, respectively. Defense response gene networks were activated in both resistant and susceptible genotypes and enriched for many of the same defense response genes, yet the hub genes of these common response modules showed greater mean expression among the resistant plants. Further, eight and six eQTL hotspots were identified at 42 and 66 HPI, respectively. The combined results of the three analyses highlight 124 candidate genes in the host for further analysis while analysis of pathogen RNA showed differential expression of 22 genes, two of which are candidate pathogen effectors. Conclusions: We identified two differentially expressed M. americana transcripts and 124 S. purpurea genes that are good candidates for future studies to confirm their role in conferring resistance.