Genomic mechanisms of climate adaptation in polyploid bioenergy switchgrass.

Long-term climate change and periodic environmental extremes threaten food and fuel security1 and global crop productivity2-4. Although molecular and adaptive breeding strategies can buffer the effects of climatic stress and improve crop resilience5, these approaches require sufficient knowledge of the genes that underlie productivity and adaptation6-knowledge that has been limited to a small number of well-studied model systems. Here we present the assembly and annotation of the large and complex genome of the polyploid bioenergy crop switchgrass (Panicum virgatum). Analysis of biomass and survival among 732 resequenced genotypes, which were grown across 10 common gardens that span 1,800 km of latitude, jointly revealed extensive genomic evidence of climate adaptation. Climate-gene-biomass associations were abundant but varied considerably among deeply diverged gene pools. Furthermore, we found that gene flow accelerated climate adaptation during the postglacial colonization of northern habitats through introgression of alleles from a pre-adapted northern gene pool. The polyploid nature of switchgrass also enhanced adaptive potential through the fractionation of gene function, as there was an increased level of heritable genetic diversity on the nondominant subgenome. In addition to investigating patterns of climate adaptation, the genome resources and gene-trait associations developed here provide breeders with the necessary tools to increase switchgrass yield for the sustainable production of bioenergy.

Nutri-cereal tissue-specific transcriptome atlas during development: Functional integration of gene expression to identify mineral uptake pathways in little millet (Panicum sumatrense).

Little millet (Panicum sumatrense Roth ex Roem. & Schult.) is an essential minor millet of southeast Asia and Africa's temperate and subtropical regions. The plant is stress-tolerant, has a short life cycle, and has a mineral-rich nutritional profile associated with unique health benefits. We report the developmental gene expression atlas of little millet (genotype JK-8) from ten tissues representing different stages of its life cycle, starting from seed germination and vegetative growth to panicle maturation. The developmental transcriptome atlas led to the identification of 342 827 transcripts. The BUSCO analysis and comparison with the transcriptomes of related species confirm that this study presents high-quality, in-depth coverage of the little millet transcriptome. In addition, the eFP browser generated here has a user-friendly interface, allowing interactive visualizations of tissue-specific gene expression. Using these data, we identified transcripts, the orthologs of which in Arabidopsis and rice are involved in nutrient acquisition, transport, and response pathways. The comparative analysis of the expression levels of these transcripts holds great potential for enhancing the mineral content in crops, particularly zinc and iron, to address the issue of "hidden hunger" and to attain nutritional security, making it a valuable asset for translational research.

miR156-PvSPL2 controls culm development by transcriptional repression of switchgrass CYTOKININ OXIDASE/DEHYDROGENASE4.

Culm development in grasses can be controlled by both miR156 and cytokinin. However, the crosstalk between the miR156-SPL module and the cytokinin metabolic pathway remains largely unknown. Here, we found CYTOKININ OXIDASE/DEHYDROGENASE4 (PvCKX4) plays a negative regulatory role in culm development of the bioenergy grass Panicum virgatum (switchgrass). Overexpression of PvCKX4 in switchgrass reduced the internode diameter and length without affecting tiller number. Interestingly, we also found that PvCKX4 was always upregulated in miR156 overexpressing (miR156OE) transgenic switchgrass lines. Additionally, upregulation of either miR156 or PvCKX4 in switchgrass reduced the content of isopentenyl adenine (iP) without affecting trans-zeatin (tZ) accumulation. It is consistent with the evidence that the recombinant PvCKX4 protein exhibited much higher catalytic activity against iP than tZ in vitro. Furthermore, our results showed that miR156-targeted SPL2 bound directly to the promoter of PvCKX4 to repress its expression. Thus, alleviating the SPL2-mediated transcriptional repression of PvCKX4 through miR156 overexpression resulted in a significant increase in cytokinin degradation and impaired culm development in switchgrass. On the contrary, suppressing PvCKX4 in miR156OE transgenic plants restored iP content, internode diameter, and length to wild-type levels. Most strikingly, the double transgenic lines retained the same increased tiller numbers as the miR156OE transgenic line, which yielded more biomass than the wild type. These findings indicate that the miR156-SPL module can control culm development through transcriptional repression of PvCKX4 in switchgrass, which provides a promising target for precise design of shoot architecture to yield more biomass from grasses.

The changing influence of host genetics on the leaf fungal microbiome throughout plant development.

Host genetics and the environment influence which fungal microbes colonize a plant. A new study in PLOS Biology finds that the relative influence of these factors changes throughout the development of the biofuel crop switchgrass growing in field settings.

Host genotype controls ecological change in the leaf fungal microbiome.

Leaf fungal microbiomes can be fundamental drivers of host plant success, as they contain pathogens that devastate crop plants and taxa that enhance nutrient uptake, discourage herbivory, and antagonize pathogens. We measured leaf fungal diversity with amplicon sequencing across an entire growing season in a diversity panel of switchgrass (Panicum virgatum). We also sampled a replicated subset of genotypes across 3 additional sites to compare the importance of time, space, ecology, and genetics. We found a strong successional pattern in the microbiome shaped both by host genetics and environmental factors. Further, we used genome-wide association (GWA) mapping and RNA sequencing to show that 3 cysteine-rich receptor-like kinases (crRLKs) were linked to a genetic locus associated with microbiome structure. We confirmed GWAS results in an independent set of genotypes for both the internal transcribed spacer (ITS) and large subunit (LSU) ribosomal DNA markers. Fungal pathogens were central to microbial covariance networks, and genotypes susceptible to pathogens differed in their expression of the 3 crRLKs, suggesting that host immune genes are a principal means of controlling the entire leaf microbiome.

A generalist-specialist trade-off between switchgrass cytotypes impacts climate adaptation and geographic range.

Polyploidy results from whole-genome duplication and is a unique form of heritable variation with pronounced evolutionary implications. Different ploidy levels, or cytotypes, can exist within a single species, and such systems provide an opportunity to assess how ploidy variation alters phenotypic novelty, adaptability, and fitness, which can, in turn, drive the development of unique ecological niches that promote the coexistence of multiple cytotypes. Switchgrass, Panicum virgatum, is a widespread, perennial C4 grass in North America with multiple naturally occurring cytotypes, primarily tetraploids (4×) and octoploids (8×). Using a combination of genomic, quantitative genetic, landscape, and niche modeling approaches, we detect divergent levels of genetic admixture, evidence of niche differentiation, and differential environmental sensitivity between switchgrass cytotypes. Taken together, these findings support a generalist (8×)­specialist (4×) trade-off. Our results indicate that the 8× represent a unique combination of genetic variation that has allowed the expansion of switchgrass' ecological niche and thus putatively represents a valuable breeding resource.

Biased mutations and gene losses underlying diploidization of the tetraploid broomcorn millet genome.

Broomcorn millet (Panicum miliaceum L.) is one of the earliest domesticated crops, and is a valuable resource to secure food diversity and combat drought stresses under the global warming scenario. However, due to the absence of extant diploid progenitors, the polyploidy genome of broomcorn millet remains poorly understood. Here, we report the chromosome-scale genome assembly of broomcorn millet. We divided the broomcorn millet genome into two subgenomes using the genome sequence of Panicum hallii, a diploid relative of broomcorn millet. Our analyses revealed that the two subgenomes diverged at ~4.8 million years ago (Mya), while the allotetraploidization of broomcorn millet may have occurred about ~0.48 Mya, suggesting that broomcorn millet is a relatively recent allotetraploid. Comparative analyses showed that subgenome B was larger than subgenome A in size, which was caused by the biased accumulation of long terminal repeat retrotransposons in the progenitor of subgenome B before polyploidization. Notably, the accumulation of biased mutations in the transposable element-rich subgenome B led to more gene losses. Although no significant dominance of either subgenome was observed in the expression profiles of broomcorn millet, we found the minimally expressed genes in P. hallii tended to be lost during diploidization of broomcorn millet. These results suggest that broomcorn millet is at the early stage of diploidization and that mutations likely occurred more on genes that were marked with lower expression levels.

Genome-wide profiling of histone (H3) lysine 4 (K4) tri-methylation (me3) under drought, heat, and combined stresses in switchgrass.

BACKGROUND: Switchgrass (Panicum virgatum L.) is a warm-season perennial (C4) grass identified as an important biofuel crop in the United States. It is well adapted to the marginal environment where heat and moisture stresses predominantly affect crop growth. However, the underlying molecular mechanisms associated with heat and drought stress tolerance still need to be fully understood in switchgrass. The methylation of H3K4 is often associated with transcriptional activation of genes, including stress-responsive. Therefore, this study aimed to analyze genome-wide histone H3K4-tri-methylation in switchgrass under heat, drought, and combined stress. RESULTS: In total, ~ 1.3 million H3K4me3 peaks were identified in this study using SICER. Among them, 7,342; 6,510; and 8,536 peaks responded under drought (DT), drought and heat (DTHT), and heat (HT) stresses, respectively. Most DT and DTHT peaks spanned 0 to + 2000 bases from the transcription start site [TSS]. By comparing differentially marked peaks with RNA-Seq data, we identified peaks associated with genes: 155 DT-responsive peaks with 118 DT-responsive genes, 121 DTHT-responsive peaks with 110 DTHT-responsive genes, and 175 HT-responsive peaks with 136 HT-responsive genes. We have identified various transcription factors involved in DT, DTHT, and HT stresses. Gene Ontology analysis using the AgriGO revealed that most genes belonged to biological processes. Most annotated peaks belonged to metabolite interconversion, RNA metabolism, transporter, protein modifying, defense/immunity, membrane traffic protein, transmembrane signal receptor, and transcriptional regulator protein families. Further, we identified significant peaks associated with TFs, hormones, signaling, fatty acid and carbohydrate metabolism, and secondary metabolites. qRT-PCR analysis revealed the relative expressions of six abiotic stress-responsive genes (transketolase, chromatin remodeling factor-CDH3, fatty-acid desaturase A, transmembrane protein 14C, beta-amylase 1, and integrase-type DNA binding protein genes) that were significantly (P < 0.05) marked during drought, heat, and combined stresses by comparing stress-induced against un-stressed and input controls. CONCLUSION: Our study provides a comprehensive and reproducible epigenomic analysis of drought, heat, and combined stress responses in switchgrass. Significant enrichment of H3K4me3 peaks downstream of the TSS of protein-coding genes was observed. In addition, the cost-effective experimental design, modified ChIP-Seq approach, and analyses presented here can serve as a prototype for other non-model plant species for conducting stress studies.

Recursive Test of Hardy-Weinberg Equilibrium in Tetraploids.

Testing for deviations from Hardy-Weinberg equilibrium (HWE) can provide fundamental information about genetic variation and evolutionary processes in natural populations. In contrast to diploids, where genotype frequencies remain constant after a single episode of random mating, polyploids, characterized by polysomic inheritance, approach HWE gradually. Here, we mathematically show the asymptotic trajectory of tetraploid equilibrium from any initial genotype frequencies. We formulate a statistical framework to test and estimate the degree of deviation from HWE at individual loci in allotetraploids and autotetraploids. Knowledge about HWE test fills an important gap in population genetic studies of tetraploids related to their evolution and ecology.

Genome-wide analysis of the switchgrass YABBY family and functional characterization of PvYABBY14 in response to ABA and GA stress in Arabidopsis.

BACKGROUND: The small YABBY plant-specific transcription factor has a prominent role in regulating plant growth progress and responding to abiotic stress. RESULTS: Here, a total of 16 PvYABBYs from switchgrass (Panicum virgatum L.) were identified and classified into four distinct subgroups. Proteins within the same subgroup exhibited similar conserved motifs and gene structures. Synteny analyses indicated that segmental duplication contributed to the expansion of the YABBY gene family in switchgrass and that complex duplication events occurred in rice, maize, soybean, and sorghum. Promoter regions of PvYABBY genes contained numerous cis-elements related to stress responsiveness and plant hormones. Expression profile analysis indicated higher expression levels of many PvYABBY genes during inflorescence development and seed maturation, with lower expression levels during root growth. Real-time quantitative PCR analysis demonstrated the sensitivity of multiple YABBY genes to PEG, NaCl, ABA, and GA treatments. The overexpression of PvYABBY14 in Arabidopsis resulted in increased root length after treatment with GA and ABA compared to wild-type plants. CONCLUSIONS: Taken together, our study provides the first genome-wide overview of the YABBY transcription factor family, laying the groundwork for understanding the molecular basis and regulatory mechanisms of PvYABBY14 in response to ABA and GA responses in switchgrass.

Genome-wide association study reveals marker-trait associations for major agronomic traits in proso millet (Panicum miliaceum L.).

MAIN CONCLUSION: The pilot-scale genome-wide association study in the US proso millet identified twenty marker-trait associations for five morpho-agronomic traits identifying genomic regions for future studies (e.g. molecular breeding and map-based cloning). Proso millet (Panicum miliaceum L.) is an ancient grain recognized for its excellent water-use efficiency and short growing season. It is an indispensable part of the winter wheat-based dryland cropping system in the High Plains of the USA. Its grains are endowed with high nutritional and health-promoting properties, making it increasingly popular in the global market for healthy grains. There is a dearth of genomic resources in proso millet for developing molecular tools to complement conventional breeding for developing high-yielding varieties. Genome-wide association study (GWAS) is a widely used method to dissect the genetics of complex traits. In this pilot study of the first-ever GWAS in the US proso millet, 71 globally diverse genotypes of 109 the US proso millet core collection were evaluated for five major morpho-agronomic traits at two locations in western Nebraska, and GWAS was conducted to identify single nucleotide polymorphisms (SNPs) associated with these traits. Analysis of variance showed that there was a significant difference among the genotypes, and all five traits were also found to be highly correlated with each other. Sequence reads from genotyping-by-sequencing (GBS) were used to identify 11,147 high-quality bi-allelic SNPs. Population structure analysis with those SNPs showed stratification within the core collection. The GWAS identified twenty marker-trait associations (MTAs) for the five traits. Twenty-nine putative candidate genes associated with the five traits were also identified. These genomic regions can be used to develop genetic markers for marker-assisted selection in proso millet breeding.

Nutritional and nutraceutical richness of neglected little millet genotypes from Eastern Ghats of India: implications for breeding and food value.

MAIN CONCLUSION: The study provides nutritional profiling of unexploited little millets from Eastern Ghats, which has ample opportunities for future breeding programs for enhancing the food quality and holds great potential in food industry. Little millet is an important small millet native to the Indian subcontinent and their nutritional value has been underutilized compared to other cereals. It's nutritional and nutraceutical profiling is essential to integrate the plants in developmental interventions. The present study evaluated comprehensive nutritional, nutraceutical and physico-functional properties of 14 selected little millet genotypes originated from Eastern Ghats of India and compared them with an improved variety (OLM 208) of the locality. The proximate compositions (per 100 g) showed significant variations, with moisture content ranging from 4.13 to 8.48 g, ash from 1.90 to 5.15 g, fat from 2.35 to 5.74 g, protein from 10.46 to 13.83 g, carbohydrate from 70.92 to 77.89 g, fiber from 2.03 to 7.82 g and energy from 372.8 to 391.1 kcal. These little millet flours are rich in phenol 5.37-12.73 mg/g, flavonoid 1.06-8.25 µg/g, vitamin C 12.72-22.86 µg/g, antioxidants 7.22-23.17%, iron 20.38 to 61.60 mg/ kg and zinc 17.47 to 37.59 mg/ kg. The first two components of principal component analysis captures 73.0% of the total variation, which reflected huge variability among the investigated genotypes. Maximum heritability and genetic advance were recorded in flavonoid, fiber, iron, zinc, phenol and vitamin C across the populations. Taken together, some indigenous little millet genotypes such as Mami, Kalia and Bada, were exceptionally rich in fiber, protein, energy, flavonoid, vitamin C and antioxidants and are nutritionally superior compared to other varieties from the locality. These nutrition rich little millet genotypes have ample opportunities for future breeding programs to enhance the cereal quality and holds great potential in food industry for making high value functional foods.

Blocking miR528 function promotes tillering and regrowth in switchgrass.

MiRNAs have been reported to be the key regulators involving a wide range of biological processes in diverse plant species, but their functions in switchgrass, an important biofuel and forage crop, are largely unknown. Here, we reported the novel function of miR528, which has expanded to four copies in switchgrass, in controlling biomass trait of tillering number and regrowth rate after mowing. Blocking miR528 activity by expressing short tandem target mimic (STTM) increased tiller number and regrowth rate after mowing. The quadruple pvmir528 mutant lines derived from genome editing also showed such improved traits. Degradome and RNA-seq analysis, combined with in situ hybridization assay revealed that up-regulation of two miR528 targets coding for Cu/Zn-SOD enzymes, might be responsible for the improved traits of tillering and regrowth in pvmir528 mutant. Additionally, natural variations in the miR528-SOD interaction exist in C3 and C4 monocot species, implying the distinct regulatory strength of the miR528-SOD module during monocot evolution. Overall, our data illuminated a novel role of miR528 in controlling biomass traits and provided a new target for genetic manipulation-mediated crop improvement.

Transcriptome and DNA methylome divergence of inflorescence development between 2 ecotypes in Panicum hallii.

The morphological diversity of the inflorescence determines flower and seed production, which is critical for plant adaptation. Hall's panicgrass (Panicum hallii, P. hallii) is a wild perennial grass that has been developed as a model to study perennial grass biology and adaptive evolution. Highly divergent inflorescences have evolved between the 2 major ecotypes in P. hallii, the upland ecotype (P. hallii var hallii, HAL2 genotype) with compact inflorescence and large seed and the lowland ecotype (P. hallii var filipes, FIL2 genotype) with an open inflorescence and small seed. Here we conducted a comparative analysis of the transcriptome and DNA methylome, an epigenetic mark that influences gene expression regulation, across different stages of inflorescence development using genomic references for each ecotype. Global transcriptome analysis of differentially expressed genes (DEGs) and co-expression modules underlying the inflorescence divergence revealed the potential role of cytokinin signaling in heterochronic changes. Comparing DNA methylome profiles revealed a remarkable level of differential DNA methylation associated with the evolution of P. hallii inflorescence. We found that a large proportion of differentially methylated regions (DMRs) were located in the flanking regulatory regions of genes. Intriguingly, we observed a substantial bias of CHH hypermethylation in the promoters of FIL2 genes. The integration of DEGs, DMRs, and Ka/Ks ratio results characterized the evolutionary features of DMR-associated DEGs that contribute to the divergence of the P. hallii inflorescence. This study provides insights into the transcriptome and epigenetic landscape of inflorescence divergence in P. hallii and a genomic resource for perennial grass biology.

Genetic linkage map construction and QTL analysis for plant height in proso millet (Panicum miliaceum L.).

KEY MESSAGE: A genetic linkage map representing proso millet genome was constructed with SSR markers, and a major QTL corresponding to plant height was mapped on chromosome 14 of this map. Proso millet (Panicum miliaceum L.) has the lowest water requirements of all cultivated cereal crops. However, the lack of a genetic map and the paucity of genomic resources for this species have limited the utility of proso millet for detailed genetic studies and hampered genetic improvement programs. In this study, 97,317 simple sequence repeat (SSR) markers were developed based on the genome sequence of the proso millet landrace Longmi 4. Using some of these markers in conjunction with previously identified SSRs, an SSR-based linkage map for proso millet was successfully constructed using a large mapping population (316 F2 offspring). In total, 186 SSR markers were assigned to 18 linkage groups corresponding to the haploid chromosomes. The constructed map had a total length of 3033.42 centimorgan (cM) covering 78.17% of the assembled reference genome. The length of the 18 linkage groups ranged from 88.89 cM (Chr. 15) to 274.82 cM (Chr. 16), with an average size of 168.17 cM. To our knowledge, this is the first genetic linkage map for proso millet based on SSR markers. Plant height is one of the most important traits in crop improvement. A major QTL was repeatedly detected in different environments, explaining 8.70-24.50% of the plant height variations. A candidate gene affecting auxin biosynthesis and transport, and ROS homeostasis regulation was predicted. Thus, the linkage map and QTL analysis provided herein will promote the development of gene mining and molecular breeding in proso millet.

Genetically correlated leaf tensile and morphological traits are driven by growing season length in a widespread perennial grass.

PREMISE: Leaf tensile resistance, a leaf's ability to withstand pulling forces, is an important determinant of plant ecological strategies. One potential driver of leaf tensile resistance is growing season length. When growing seasons are long, strong leaves, which often require more time and resources to construct than weak leaves, may be more advantageous than when growing seasons are short. Growing season length and other ecological conditions may also impact the morphological traits that underlie leaf tensile resistance. METHODS: To understand variation in leaf tensile resistance, we measured size-dependent leaf strength and size-independent leaf toughness in diverse genotypes of the widespread perennial grass Panicum virgatum (switchgrass) in a common garden. We then used quantitative genetic approaches to estimate the heritability of leaf tensile resistance and whether there were genetic correlations between leaf tensile resistance and other morphological traits. RESULTS: Leaf tensile resistance was positively associated with aboveground biomass (a proxy for fitness). Moreover, both measures of leaf tensile resistance exhibited high heritability and were positively genetically correlated with leaf lamina thickness and leaf mass per area (LMA). Leaf tensile resistance also increased with the growing season length in the habitat of origin, and this effect was mediated by both LMA and leaf thickness. CONCLUSIONS: Differences in growing season length may promote selection for different leaf lifespans and may explain existing variation in leaf tensile resistance in P. virgatum. In addition, the high heritability of leaf tensile resistance suggests that P. virgatum will be able to respond to climate change as growing seasons lengthen.

Rhizophagus irregularis and biochar can synergistically improve the physiological characteristics of saline-alkali resistance of switchgrass.

Inoculation of arbuscular mycorrhizal fungi (AMF) or biochar (BC) application can improve photosynthesis and promote plant growth under saline-alkali stress. However, little is known about the effects of the two combined on growth and physiological characteristics of switchgrass under saline-alkali stress. This study examined the effects of four treatments: (1) no AMF inoculation and no biochar addition (control), (2) biochar (BC) alone, (3) AMF (Rhizophagus irregularis, Ri) alone, and (4) the combination of both (BC+Ri) on the plant biomass, antioxidant enzymes, chlorophyll, and photosynthetic parameters of switchgrass under saline-alkali stress. The results showed that the above-ground, belowground and total biomass of switchgrass in the BC+Ri treatment group was significantly higher (+136.7%, 120.2% and 132.4%, respectively) than in other treatments compared with Control. BC+Ri treatment significantly increased plant leaves' relative chlorophyll content, antioxidant enzyme activity, and photosynthesis parameters. It is worth noting that the transpiration rate, stomatal conductance, net photosynthetic rate, PSII efficiency and other photosynthetic-related indexes of the BC+Ri treatment group were the highest (38% to 54% higher than other treatments). The fitting results of light response and CO2 response curves showed that the light saturation point, light compensation point, maximum carboxylation rate and maximum electron transfer rate of switchgrass in the Ri+BC treatment group were the highest. In conclusion, biochar combined with Ri has potential beneficial effects on promoting switchgrass growth under saline-alkali stress and improving the activity of antioxidant enzymes and photosynthetic characteristics of plants.

Comparative Multi-Omics Analysis of Broomcorn Millet in Response to Anthracocystis destruens Infection.

Anthracocystis destruens is the causal agent of broomcorn millet (Panicum miliaceum) smut disease, which results in serious yield losses in broomcorn millet production. However, the molecular basis underlying broomcorn millet defense against A. destruens is less understood. In this study, we investigated how broomcorn millet responds to infection by A. destruens by employing a comprehensive multi-omics approach. We examined the responses of broomcorn millet across transcriptome, metabolome, and microbiome levels. Infected leaves exhibited an upregulation of genes related to photosynthesis, accompanied by a higher accumulation of photosynthesis-related compounds and alterations in hormonal levels. However, broomcorn millet genes involved in immune response were downregulated post A. destruens infection, suggesting that A. destruens may suppress broomcorn millet immunity. In addition, we show that the immune suppression and altered host metabolism induced by A. destruens have no significant effect on the microbial community structure of broomcorn millet leaf, thus providing a new perspective for understanding the tripartite interaction between plant, pathogen, and microbiota.

The genetic basis of the root economics spectrum in a perennial grass.

Construction economics of plant roots exhibit predictable relationships with root growth, death, and nutrient uptake strategies. Plant taxa with inexpensively constructed roots tend to more precisely explore nutrient hotspots than do those with costly constructed roots but at the price of more frequent tissue turnover. This trade-off underlies an acquisitive to conservative continuum in resource investment, described as the "root economics spectrum (RES)." Yet the adaptive role and genetic basis of RES remain largely unclear. Different ecotypes of switchgrass (Panicum virgatum) display root features exemplifying the RES, with costly constructed roots in southern lowland and inexpensively constructed roots in northern upland ecotypes. We used an outbred genetic mapping population derived from lowland and upland switchgrass ecotypes to examine the genetic architecture of the RES. We found that absorptive roots (distal first and second orders) were often "deciduous" in winter. The percentage of overwintering absorptive roots was decreased by northern upland alleles compared with southern lowland alleles, suggesting a locally-adapted conservative strategy in warmer and acquisitive strategy in colder regions. Relative turnover of absorptive roots was genetically negatively correlated with their biomass investment per unit root length, suggesting that the key trade-off in framing RES is genetically facilitated. We also detected strong genetic correlations among root morphology, root productivity, and shoot size. Overall, our results reveal the genetic architecture of multiple traits that likely impacts the evolution of RES and plant aboveground-belowground organization. In practice, we provide genetic evidence that increasing switchgrass yield for bioenergy does not directly conflict with enhancing its root-derived carbon sequestration.

The effect of AMF combined with biochar on plant growth and soil quality under saline-alkali stress: Insights from microbial community analysis.

Arbuscular mycorrhizal fungi (AMF) and biochar application individually can enhance plant tolerance to saline-alkali stress and promote plant growth efficiency. However, little is known about the potential synergistic effects of their combination on improving plant growth and soil quality under saline-alkali stress. This experiment adopted the potted method to explore the effects of four treatments on switchgrass growth and soil quality: biochar (BC), Rhizophagus irregularis (Ri), biochar + Ri (BR) and a control without biochar or Ri (CK). Compared to the CK treatment, the switchgrass biomass increased by 92.4 %, 148.6 %, and 177.3 % in the BC, Ri, and BR treatment groups, respectively. Similarly, the rhizosphere soil quality index increased by 29.33 %, 22.7 %, and 49.1 % in the respective treatment groups. The BR treatment significantly altered the rhizosphere soil microbial composition and diversity. Notably, compared to the other treatments, the archaeal α-diversity in the BR group showed a significant decrease. BR treatment significantly increased the relative abundance of bacteria, fungi and archaea at the genus level (e.g., Bacillus, Trichome and candidatus_methanopenens). Network analysis showed that the complexity and closeness of interactions between different microbial taxa were stronger in the BC, Ri and BR treatments than in the CK treatment, with BR being the more prominent. In summary, biochar combined with Ri has a better effect on promoting the growth of switchgrass under saline-alkali stress, improving the quality of saline-alkali soil, and increasing soil microbial diversity. This study provides a new approach for the efficient development and utilization of saline-alkali land.