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1. A comparative approach for selecting orthologous candidate genes underlying signal in genome-wide association studies across multiple species

   https://doi.org/10.1101/2023.10.05.561051  

   Whitt, L; Mahood, E H; Ziegler, G; Luebbert, C; Gillman, J D; Norton, G J; Price, A H; Salt, DE; Dilkes, B; Baxter, I (October 2023, bioRxiv)

   Advances in quantitative genetics have enabled researchers to identify genomic regions associated with changes in phenotype. However, genomic regions can contain hundreds to thousands of genes, and progressing from genomic regions to candidate genes is still challenging. In genome-wide association studies (GWAS) measuring elemental accumulation (ionomic) traits, a mere 5% of loci are associated with a known ionomic gene - indicating that many causal genes are still unknown. To select candidates for the remaining 95% of loci, we developed a method to identify conserved genes underlying GWAS loci in multiple species. For 19 ionomic traits, we identified 14,336 candidates across Arabidopsis, soybean, rice, maize, and sorghum. We calculated the likelihood of candidates with random permutations of the data and determined that most of the top 10% of candidates were orthologous genes linked to GWAS loci across all five species. The candidate list also includes orthologous genes with previously established ionomic functions in Arabidopsis and rice. Our methods highlight the conserved nature of ionomic genetic regulators and enable the identification of previously unknown ionomic genes. 

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2. Morphological and Phylogenetic Characterization of Alternaria Section Undifilum Fungal Endophytes from Astragalus and Swainsona spp.

   https://doi.org/10.3390/jof11070541  

   Neyaz, Marwa; Adebisi, Olabisi; Cook, Daniel; Creamer, Rebecca (July 2025, Journal of Fungi)

   The locoweeds Astragalus and Oxytropis in the Americas and China, as well as Swainsona in Australia, harbor swainsonine-producing endophytes responsible for “locoism” or “pea struck” syndromes in grazing animals. Demonstration of Alternaria section Undifilum spp. requires demonstration of morphological characters such as a wavy germ tube and slow growth. While Astragalus wetherilli, A. pubentissimus, Swainsona canescens, and S. galegifolia plants have been shown to contain swainsonine, and fungi isolated from the plants have been partially characterized genetically, the fungi have not been characterized morphologically. This work sought to complete morphological characterization and determine species for those fungi and from fungi associated with Swainsona luteola and S. brachycarpa. The fungi were isolated from their hosts onto media and exhibited slow growth, resulting in a colony diameter of approximately 10 mm after 30 days. Morphological identification revealed production of conidia that produced a wavy germ tube for the endophytes from Astragalus pubentissimus species, Swainsona canescens, and S. galegifolia. Sequence analyses of the ITS region and the swnK-KS and swnK-TR genes of these fungi suggest that the fungi isolated from Astragalus are closely related and distinct from the fungi isolated from Swainsona. Presence of the swnK gene demonstrates that all the fungi have a necessary component to produce swainsonine. Fungi isolated from Astragalus spp. differed in color, growth, and conidium size, and/or their sequences. While the fungi isolated from Swainsona canescens and S. galegifolia endophytes differed in color, growth, and conidium size, those isolated from Swainsona luteola and S. brachycarpa did not produce conidia. Sequences from all Swainsona endophytes were almost identical and were concluded to be the same species. The new species described here are Alternaria wetherii, A. pubentissima, A. pubentissimoides, and A. swainsonii. 

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3. Analysis of the Mycotoxin Levels and Expression Pattern of SWN Genes at Different Time Points in the Fungus Slafractonia leguminicola

   https://doi.org/10.3390/microorganisms12040670  

   Das, Sumanjari; Gardner, Dale R; Cook, Daniel; Creamer, Rebecca (April 2024, Microorganisms)

   de_Paula, Renato G; Silva, Roberto N (Ed.)

   The fungal plant pathogen Slafractonia leguminicola produces two mycotoxins that affect animals: slaframine, which causes slobbers, and swainsonine, which causes locoism. Slafractonia leguminicola contains the swainsonine-associated orthologous gene clusters, “SWN”, which include a multifunctional swnK gene (NRPS-PKS hybrid), swnH1 and swnH2 (nonheme iron dioxygenase genes), swnN and swnR (reductase genes), and swnT (transmembrane transporter). In addition to these genes, two paralogs of swnK, swnK1 (paralog1) and swnk2 (paralog2), are found in S. leguminicola. cDNAs from total mRNA were isolated from the S. leguminicola mycelia grown in the culture plates as well as from leaves inoculated with the fungal mycelia at different time points, and expression pattern of the SWN genes were analyzed using RT-qPCR. The concentrations of swainsonine and slaframine production from this fungus at different time points were also examined using liquid chromatography–mass spectrometry. The timing of gene expression was similar in cultured fungus and inoculated leaves and agreed with our proposed biosynthetic pathway. Substantially more swainsonine was produced than slaframine during time course studies. 

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4. Evolutionary characteristics of intergenic transcribed regions indicate rare novel genes and widespread noisy transcription in the Poaceae

   https://doi.org/10.1038/s41598-019-47797-y  

   Lloyd, John_P; Bowman, Megan_J; Azodi, Christina_B; Sowers, Rosalie_P; Moghe, Gaurav_D; Childs, Kevin_L; Shiu, Shin-Han (August 2019, Scientific Reports)

   Abstract Extensive transcriptional activity occurring in intergenic regions of genomes has raised the question whether intergenic transcription represents the activity of novel genes or noisy expression. To address this, we evaluated cross-species and post-duplication sequence and expression conservation of intergenic transcribed regions (ITRs) in four Poaceae species. Among 43,301 ITRs across the four species, 34,460 (80%) are species-specific. ITRs found across species tend to be more divergent in expression and have more recent duplicates compared to annotated genes. To assess if ITRs are functional (under selection), machine learning models were established inOryza sativa(rice) that could accurately distinguish between phenotype genes and pseudogenes (area under curve-receiver operating characteristic = 0.94). Based on the models, 584 (8%) and 4391 (61%) rice ITRs are classified as likely functional and nonfunctional with high confidence, respectively. ITRs with conserved expression and ancient retained duplicates, features that were not part of the model, are frequently classified as likely-functional, suggesting these characteristics could serve as pragmatic rules of thumb for identifying candidate sequences likely to be under selection. This study also provides a framework to identify novel genes using comparative transcriptomic data to improve genome annotation that is fundamental for connecting genotype to phenotype in crop and model systems. 

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5. Transcriptional Divergence Underpinning Sexual Development in the Fungal Class Sordariomycetes

   https://doi.org/10.1128/mbio.01100-22  

   Kim, Wonyong; Wang, Zheng; Kim, Hyeonjae; Pham, Kasey; Tu, Yujia; Townsend, Jeffrey P.; Trail, Frances (June 2022, mBio)

   Goldman, Gustavo H. (Ed.)

   ABSTRACT Gene expression divergence through evolutionary processes is thought to be important for achieving programmed development in multicellular organisms. To test this premise in filamentous fungi, we investigated transcriptional profiles of 3,942 single-copy orthologous genes (SCOGs) in five related sordariomycete species that have morphologically diverged in the formation of their flask-shaped perithecia. We compared expression of the SCOGs to inferred gene expression levels of the most recent common ancestor of the five species, ranking genes from their largest increases to smallest increases in expression during perithecial development in each of the five species. We found that a large proportion of the genes that exhibited evolved increases in gene expression were important for normal perithecial development in Fusarium graminearum . Many of these genes were previously uncharacterized, encoding hypothetical proteins without any known functional protein domains. Interestingly, the developmental stages during which aberrant knockout phenotypes appeared largely coincided with the elevated expression of the deleted genes. In addition, we identified novel genes that affected normal perithecial development in Magnaporthe oryzae and Neurospora crassa , which were functionally and transcriptionally diverged from the orthologous counterparts in F. graminearum . Furthermore, comparative analysis of developmental transcriptomes and phylostratigraphic analysis suggested that genes encoding hypothetical proteins are generally young and transcriptionally divergent between related species. This study provides tangible evidence of shifts in gene expression that led to acquisition of novel function of orthologous genes in each lineage and demonstrates that several genes with hypothetical function are crucial for shaping multicellular fruiting bodies. IMPORTANCE The fungal class Sordariomycetes includes numerous important plant and animal pathogens. It also provides model systems for studying fungal fruiting body development, as its members develop fruiting bodies with a few well-characterized tissue types on common growth media and have rich genomic resources that enable comparative and functional analyses. To understand transcriptional divergence of key developmental genes between five related sordariomycete fungi, we performed targeted knockouts of genes inferred to have evolved significant upward shifts in expression. We found that many previously uncharacterized genes play indispensable roles at different stages of fruiting body development, which have undergone transcriptional activation in specific lineages. These novel genes are predicted to be phylogenetically young and tend to be involved in lineage- or species-specific function. Transcriptional activation of genes with unknown function seems to be more frequent than ever thought, which may be crucial for rapid adaption to changing environments for successful sexual reproduction. 

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