Jeffrey L. Bennetzen

Key message: Although herbicide resistance in Setaria is rampant and cosmopolitian across four herbicide families, we encountered little evidence of target-site resistance, indicating diverse non-target mechanisms of metabolizing, sequestering, and overwhelming herbicides. Setaria viridis is a cosmopolitan weed and model genetic system with increasing reports of resistance to multiple classes of herbicides. Our goal was to assess the herbicide resistance and allelic diversity in herbicide target genes in a collection of Setaria genotypes from North America and Eurasia, and identify the occurrence of novel and known target-site mutations that led to resistance. A total of 214 Setaria genotypes were exposed to commonly used herbicides that inhibit specific genes: herbicide action class (HRAC) group 1 herbicides targeting acetyl-CoA carboxylase (ACCase), HRAC 2 targeting acetolactate synthase (ALS), HRAC 9 targeting 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase, and HRAC 10 targeting glutamine synthetase. ACCase and ALS genes in 53 accessions were PCR-amplified and sequenced. Whole-genome sequencing reads covering the target genes were analyzed for an additional 98 genotypes. Herbicide trials showed that 30% of our accessions set seed following application of at least one herbicide, and 13 accessions were resistant to multiple classes of herbicides. Although there were numerous SNPs, including some known to lead to resistance, in our target genes, SNPs found predominantly in herbicide-resistant genotypes were largely intronic or synonymous. A small number of amino acid substitutions in ALS and ACCase indicated potential and incomplete resistance to HRAC 1 and 2 herbicides, but no SNPs putatively associated with herbicide resistance were identified in the other 6 target-site genes. The broader pattern of herbicide resistance in S. viridis is likely driven by non-target mutations that detoxify or compartmentalize applied herbicides.

Societal Impact Statement: Sorghum bicolor is a globally important cereal crop with annual yields exceeding 50 million tons across more than 100 countries and can be grown on marginal lands where conventional agriculture is limited. We examined how eight genetically diverse sorghum genotypes shaped arbuscular mycorrhizal fungal (AMF) community assembly across two contrasting field environments. We found that sorghum genotype influences AMF community composition and that this showed a pattern over time. These findings suggest that crop genotype selection can be a useful practical tool to modulate root‐microbial interactions in low‐input and marginal agricultural systems, with implications for sustainable intensification strategies and microbiome‐informed crop breeding programs. Summary: Arbuscular mycorrhizal fungal (AMF) symbiosis can influence crop production but can be variable across environmental conditions, host‐partner complementarity, and temporal dynamics. Understanding how these factors interact to shape AMF community assembly allows for the selection of crop genotypes that may maximally utilize AMF associations in agricultural systems. We assessed the development of AMF communities colonizing the roots of eight genetically diverse genotypes of Sorghum bicolor across a growing season. We used two field sites with contrasting environments and management histories. Sorghum cultivated in Georgia (GA) sorghum harbored 3.7‐fold more species than Arizona (AZ). We observed evidence of host‐filtering of AMF communities, with genotypes displaying more distinct associations in GA than AZ. AZ showed rapid shifts from early Funneliformis mosseae dominance to increasing dominance by either Entrophospora etunicata or Diversispora aurantia . In GA, we observed temporal variation associated with turnover of low‐abundance taxa contributing to larger family‐level patterns. Our findings demonstrate that there is potential for leveraging intra‐species genetic variation in AMF community assembly as an extended plant phenotype.

Switchgrass (Panicum virgatum L.) is an important cellulosic biofuel grass native to North America. Rust, caused by Puccinia spp. is the most predominant disease of switchgrass and has the potential to impact biomass conversion. In this study, virulence patterns were determined on a set of 38 switchgrass genotypes for 14 single-spore rust isolates from 14 field samples collected in seven states. Single nucleotide polymorphism (SNP) variation was also assessed in 720 sequenced cloned amplicons representing 654 base pairs of the elongation factor 1-α gene from the field samples. Five major haplotypes were identified differing by 11 out of the 39 SNP positions identified. STRUCTURE, Principal Coordinate Analysis, and phylogenetic analyses divided the rust population into two genetic clusters. Virginia and Georgia had the highest and lowest rust genetic diversity, respectively. Only nine accessions showed a differential disease response between the 14 isolates, allowing the identification of eight races, differing by 1–3 virulence factors. Overall, the results suggested clonal reproduction of the pathogen and a North–South differentiation via local adaptation. However, similar haplotypes and races were also recovered from several states, suggesting migration events, and highlighting the need to further investigate the switchgrass rust population structure and evolution in the USA.

Introduction: Although a few bacteria have been studied in great depth, relatively little is known about the characteristics of microbe-microbe interactions that occur within ecosystems on a daily basis. A simple, robust technique was developed to set up the foundation for investigating pairwise bacterial-bacterial interactions, using cell-cell binding as a self-selective mechanism to identify interesting bacterial species pairs.Methods: Using a Serratia marcescens strain (SMC43) isolated from Georgia soil as a “bait”, specific bacteria were purified by their specificity in binding SMC43 bacteria that were themselves attached to a wooden applicator stick.Results: The isolated Microbial Partners (MiPners) were greatly enriched for members of the genera Sphingobium and Caulobacter. Two streaked MiPners were unable to grow on the plates employed after separation from SMC43to be separated from, and grow on the plate type tested without, SMC43.Discussion: This suggests that the MiPner technology will be one strategy for purifying bacteria that were previously recalcitrant to culturing.

Drought stress significantly impairs growth, and microbial interactions in sorghum. This study explores the transcriptional and microbial shifts in sorghum under drought, revealing key adaptations to water deficit. LC-MS (Liquid Chromatography-Mass Spectrometry) analyses revealed that drought stress induced abscisic acid while significantly reducing jasmonic acid levels in sorghum roots, likely due to resource conservation strategies during drought. Transcriptional reprogramming highlighted the upregulation of genes in the roots involved in mineral homeostasis ( Ferritin 1 , Iron dehydrogenase , Nitrate transporter 1 ), hormone signaling ( Ethylene-insensitive protein 3 , Gibberellin 2-oxidase ), and osmotic regulation ( Aquaporin , Dehydrin ), underlining key adaptive responses to maintain nutrient uptake, redox status, and cellular turgor. In Fe-supplemented plants, increased Fe in roots correlated with increased Ferritin 1 expression, improved plant health, and reduced Fenton reaction rate and H□O□ levels. This suggests that ferritin helps minimize oxidative stress under drought in sorghum. Drought reduced root-associated bacterial diversity and richness while enriching drought tolerance-associated genera, such as Burkholderia, Caballeronia and Paraburkholderia , known for promoting plant growth through auxin production, phosphate solubilization, and siderophore-mediated iron acquisition. In contrast, fungal diversity and richness remained unchanged, dominated by Talaromyces , which showed a statistically non-significant increase under drought. Random forest models could not identify functional predictors for fungi but revealed a shift in bacterial functional groups under drought, with enrichment in phototrophy, methylotrophy, and nitrate reduction, traits emphasizing microbial roles in nutrient cycling and drought adaptation of sorghum. This study provides insights into the role of ferritin and potential bacterial bioinoculants that could enhance sorghum resilience to drought. Future research should validate these findings in the field to develop sustainable strategies for drought-tolerant sorghum and climate-resilient agriculture.

We conducted a comprehensive analysis of the total microbiome and transcriptionally active microbiome communities in the roots and root nodules of Prosopis cineraria , an important leguminous tree in arid regions of many Asian countries. Mature P. cineraria trees growing in the desert did not exhibit any detected root nodules. However, we observed root nodules on the roots of P. cineraria growing on a desert farm and on young plants growing in a growth chamber, when inoculated with rhizosphere soil, including with rhizosphere soil from near desert tree roots that had no nodules. Compared to nearby soil, non-nodulated roots were enriched with Actinobacteria (e.g., Actinophytocola sp.), whereas root nodules sampled from the desert farm and growth chamber had abundant Alphaproteobacteria (e.g., Ensifer sp.). These nodules yielded many microbes in addition to such nitrogen-fixing bacteria as Ensifer and Sinorhizobium species. Significant differences exist in the composition and abundance of microbial isolates between the nodule surface and the nodule endosphere. Shotgun metagenome analysis of nodule endospheres revealed that the root nodules comprised over 90% bacterial DNA, whereas metatranscriptome analysis showed that the plant produces vastly more transcripts than the microbes in these nodules. Control inoculations demonstrated that four out of six Rhizobium, Agrobacterium, or Ensifer isolates purified from P. cineraria nodules produced nodules in the roots of P. cineraria seedlings under greenhouse conditions. The best nodulation was achieved when seedlings were inoculated with a mixture of those bacterial strains. Though root nodulation could be achieved under water stress conditions, nodule number and nodule biomass increased with copious water availability. . IMPORTANCE: Microbial communities were investigated in roots and root nodules of Prosopis cineraria , a leguminous tree species in arid Asian regions that is responsible for exceptionally important contributions to soil fertility in these dramatically dry locations. Soil removed from regions near nodule-free roots on these mature plants contained an abundance of bacteria with the genetic ability to generate nodules and fix nitrogen but did not normally nodulate in their native rhizosphere environment, suggesting a very different co-evolved relationship than that observed for herbaceous legumes. The relative over-expression of the low-gene-density plant DNA compared to the bacterial DNA in the nodules was also unexpected, indicating a very powerful induction of host genetic contributions within the nodule. Finally, the water dependence of nodulation in inoculated seedlings suggested a possible link between early seedling growth (before a deep root system can be developed) and the early development of nitrogen-fixing capability.

Summary: Grasses (Poaceae) comprise around 11,800 species and are central for human livelihoods and terrestrial ecosystems. Knowing their relationships and evolutionary history is key to comparative research and crop breeding. Advances in genome-scale sequencing allow for increased breadth and depth of phylogenomic analyses, making it possible to infer a new reference species tree of the family. We inferred a comprehensive species tree of grasses by combining new and published sequences for 331 nuclear genes from genome, transcriptome, target enrichment and shotgun data. Our 1,153-tip tree covers 79% of grass genera (including 21 genera sequenced for the first time) and all but two small tribes. We compared it to a 910-tip plastome tree. The nuclear phylogeny matches that of the plastome at most deep branches, with only a few instances of incongruence. Gene tree–species tree reconciliation suggests that reticulation events occurred repeatedly in the history of grasses. We provide a robust framework for the grass tree of life to support research on grass evolution, including modes of reticulation, and genetic diversity for sustainable agriculture.

Polyploidy (genome duplication) is a pivotal force in evolution. However, the interactions between parental genomes in a polyploid nucleus, frequently involving subgenome dominance, are poorly understood. Here we showcase analyses of a bamboo system (Poaceae: Bambusoideae) comprising a series of lineages from diploid (herbaceous) to tetraploid and hexaploid (woody), with 11 chromosome-level de novo genome assemblies and 476 transcriptome samples. We find that woody bamboo subgenomes exhibit stunning karyotype stability, with parallel subgenome dominance in the two tetraploid clades and a gradual shift of dominance in the hexaploid clade. Allopolyploidization and subgenome dominance have shaped the evolution of tree-like lignified culms, rapid growth and synchronous flowering characteristic of woody bamboos as large grasses. Our work provides insights into genome dominance in a remarkable polyploid system, including its dependence on genomic context and its ability to switch which subgenomes are dominant over evolutionary time.