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In vitro evaluation of grapevine endophytes, epiphytes and sap micro-organisms for potential use to control grapevine trunk disease pathogens.

Grapevine trunk diseases (GTDs) threaten the economic sustainability of viticulture worldwide causing a significant reduction of both yields and quality of grapes. Biological control presents a promising sustainable alternative to cultural and chemical methods to mitigate the effects of pathogens causing GTDs, including Botryosphaeria dieback, Eutypa dieback and Esca. This study aimed to identify naturally occurring potential biological control agents from a variety of grapevine tissues, including sap, cane and pith and evaluate their antagonistic activity against selected fungal pathogens responsible for GTDs in vitro. Bacterial and fungal isolates were preliminary screened in vitro to determine their antifungal activity via a dual culture assay against Neofusicoccum parvum and Eutypa lata. Among the fungal isolates, Trichoderma spp. inhibited E. lata mycelial growth up to 64% and N. parvum mycelial growth up to 73% with overgrowth and stopped growth being the likely antagonistic mechanisms. Among the bacterial isolates, Bacillus spp. inhibited E. lata mycelial growth up to 20% and N. parvum mycelial growth up to 40%. Select antagonistic isolates of Trichoderma, Bacillus and Aureobasidium spp. were subject to further dual culture antifungal analysis against Diplodia seriata and Diaporthe ampelina, with Trichoderma isolates consistently causing the greatest inhibition. Volatile organic compound antifungal analysis revealed that these Trichoderma isolates resulted significantly inhibited mycelial growth of N. parvum, E. lata and D. ampelina causing up to 20.11%, 60.55% and 70.9% inhibition respectively (P[≤]0.05). Multilocus sequence analysis revealed that the Trichoderma isolates are most closely related to Trichoderma asperellum and Trichoderma hamatum. This study identifies grapevine sap as a novel source of potential biological control agents for control of GTDs to support existing efforts to control GTDs. Further testing will be necessary to fully characterize these microbes mode of antagonism and assess their efficacy for pruning wound protection in planta.

plant biology↗

Tabula Glycine: The whole-soybean single-cell resolution transcriptome atlas

Soybean (Glycine max) is an essential source of protein and oil with high nutritional value for human and animal consumption. To enhance our understanding of soybean biology, it is essential to have accurate information regarding the expression of each of its 55,897 protein-coding genes. Here, we present "Tabula Glycine", the soybean single-cell resolution transcriptome atlas. This atlas is composed of single-nucleus RNA-sequencing data of nearly 120,000 nuclei isolated from 10 different Glycine max organs and morphological structures comprising the entire soybean plant. These nuclei are grouped into 157 different clusters based on their transcriptomic profiles. Among genes, the pattern of activity of transcription factor genes is sufficient to define most cell types and their organ/morphological structure of origin, suggesting that transcription factors are key determinants of cell identity and function. This unprecedented level of resolution makes the Tabula Glycine a unique resource for the plant and soybean communities.

plant biology↗

Semi-automated workflow for high-throughput Agrobacterium-mediated plant transformation

High-throughput experiments in plants are hindered by long generation times and high costs. To address these challenges, we present an optimized pipeline for Agrobacterium tumefaciens transformation and simplified a protocol to obtain stable transgenic lines of the model liverwort Marchantia polymorpha, paving the way for efficient high-throughput experiments for plant synthetic biology and other applications. Our protocol involves freeze-thaw Agrobacterium transformation method in 6-well plates that can be adapted to robotic automation. Using the Opentrons open-source platform, we implemented a semi-automated protocol showing similar efficiency compared to manual manipulation. Additionally, we have streamlined and simplified the process of stable transformation and selection of M. polymorpha, reducing cost, time, and manual labour without compromising transformation efficiency. The addition of sucrose in the selection media significantly enhances the production of gemmae, accelerating the generation of isogenic plants. We believe these protocols have the potential to facilitate high-throughput screenings in diverse plant species and represent a significant step towards the full automation of plant transformation pipelines. This approach allows testing [~]100 constructs per month, using conventional plant tissue culture facilities. We recently demonstrated the successful implementation of this protocol for screening hundreds of fluorescent reporters in Marchantia gemmae.

synthetic biology↗

New molecular players: siRNA expression and gene regulation in Hordeum vulgare ageing seeds after germination

Small interfering RNAs (siRNAs), a subclass of small non-coding RNAs, play crucial roles in regulating seed germination and viability through epigenetic mechanisms like RNA-directed DNA methylation (RdDM). This study presents the first comprehensive investigation of siRNA profiles linked to seed viability and germination in barley (Hordeum vulgare), utilizing a unique set of seeds from a single batch subjected to controlled long-term storage. Some seeds lost viability due to moisture exposure from unsealing, creating a natural experimental model to explore vigor effects. sRNA sequencing revealed 85,728 differentially expressed siRNAs, with distinct patterns between regenerated, high-viability, and low-viability seeds. Notably, trans-acting siRNAs (ta-siRNAs) showed peak abundance at different imbibition times depending on seed quality, suggesting dynamic regulation. Around 46% of siRNAs were 21 nucleotides, and 54% were 22 nucleotides long. Gene Ontology and degradome analyses confirmed siRNA target genes involved in vital biological processes such as cytochrome complex function, root development, cell maturation, and carbohydrate metabolism. Despite RNA degradation in low-viability seeds, siRNAs remained relatively stable, indicating their potential role in maintaining seed metabolic activity during dormancy release and germination initiation. This pioneering research uncovers novel insights into siRNA-mediated control of seed longevity and germination, highlighting the innovative use of stable, well-characterized plant material to disentangle molecular mechanisms underpinning seed vigor and germination success.

plant biology↗

New algorithm for pearl millet modelling in APSIM allowing a mechanistic simulation of tillers

We present a new algorithm for pearl millet simulation in APSIM. Compared to the actual released model, this new model increases the ability to simulate dynamic tillers by integrating recent progresses about biological understanding of the tillering mechanism. The new algorithm also offers the possibility to have an increased genetic control over key functions like canopy development and tillering through additional genotype related parameters. Next to model description, we also present the parametrization of 9444 and HHB 67-2, two genotypes broadly used in India. Overall, we could show that the new algorithm is able to reconstruct the main plant function like biomass accumulation and tillering. Some margin of improvement remains concerning the simulation of tiller cessation.

plant biology↗

Variability of soybean response to rhizobia inoculant, Vermicompost, and a legume-specific fertilizer blend in Siaya County of Kenya

Rhizobia inoculation can increase soybean yield, but its performance is influenced by soybean genotype, rhizobia strains, environment, and crop management among others. The objective of the study was to assess soybean response to rhizobia inoculation when grown in soils amended with urea or Vermicompost to improve nitrogen levels. Two greenhouse experiments and one field trial at two sites were carried out. The first greenhouse experiment included soils from sixty locations, sampled from smallholder farms in Western Kenya. The second greenhouse experiment consisted of one soil selected from soils used in the first experiment where inoculation response was poor. The soil was amended with Vermicompost or urea. In the two greenhouse experiments, Legumefix(R) (inoculant) + Sympal (legume fertilizer blend) were used as a standard package. Results from the second greenhouse experiment were then validated in the field. In the first greenhouse trial, soybean response to inoculation was significantly affected by soil fertility based on nodule fresh weight and shoot biomass. Soils with low nitrogen had low to no response to inoculation. After amendment, nodule fresh weight, nodule effectiveness, nodule occupancy, and shoot dry biomass were greater in the treatment amended with Vermicompost than those amended with urea (Legumefix(R) + Sympal + Vermicompost and Legumefix(R) + Sympal + urea). Under field conditions, trends were similar to the second experiment for nodulation, nodule occupancy, and nitrogen uptake resulting in significantly greater grain yields (475, 709, 856, 880, 966 kg ha-1) after application of Vermicompost at 0, 37, 74, 111, and 148 kg N ha-1, respectively. It was concluded that soybean nodulation and biological nitrogen fixation in low fertility soils would not be suppressed by organic amendments like Vermicompost up to 148 kg N ha-1.

plant biology↗

Cell specialization and coordination in Arabidopsis leaves upon pathogenic attack revealed by scRNA-seq.

Plant defense responses involve several biological processes that allow plants to fight against pathogenic attacks. How these different processes are orchestrated within organs and depend on specific cell types is poorly known. Here, using scRNA-seq technology on three independent biological replicates, we identified 10 distinct cell populations in wild-type Arabidopsis leaves inoculated with the bacterial pathogen Pseudomonas syringae DC3000. Among those, we retrieved major cell types of the leaves (mesophyll, guard, epidermal, companion and vascular S cells) to which we could associate characteristic transcriptional reprogramming and regulators, thereby specifying different cell-type responses to the pathogen. Further analyses of transcriptional dynamics, based on inference of cell trajectories, indicated that the different cell types, in addition to their characteristic defense responses, can also share similar modules of gene reprogramming, allowing for instance vascular S cells, epidermal cells and mesophyll cells to converge towards an identical cell fate, mostly characterized by lignification and detoxification functions. Moreover, it appeared that the defense responses of these three cell types can evolve along a second separate path. As this divergence does not correspond to the differentiation between immune and susceptible cells, we speculate that this might reflect the discrimination between cell-autonomous and non-cell-autonomous responses. Altogether our data provide an upgraded framework to describe, explore and explain the specialization and the coordination of plant cell responses upon pathogenic challenge.

plant biology↗

Ubiquitin-related genes are differentially expressed in isogenic lines contrasting for pericarp cell size and grain weight in hexaploid wheat

BackgroundThere is an urgent need to increase global crop production. Identifying and combining genes controlling indidviual yield components, such as grain weight, holds the potential to enhance crop yields. Transcriptomics is a powerful tool to gain insights into the complex gene regulatory networks that underlie such traits, but relies on the availability of a high-quality reference sequence and accurate gene models. Previously, we identified a grain weight QTL on wheat chromosome 5A (5A QTL) which acts during early grain development to increase grain length through cell expansion in the pericarp. In this study, we performed RNA-sequencing on near isogenic lines (NILs) segregating for the 5A QTL and used the latest gene models to identify differentially expressed (DE) genes and pathways that potentially influence pericarp cell size and grain weight in wheat.\n\nResultsWe sampled grains at four and eight days post anthesis and found genes associated with metabolism, biosynthesis, proteoloysis and defence response to be upregulated during this stage of grain development in both NILs. We identified a specific set of 112 transcripts DE between 5A NILs at either time point, including seven potential candidates for the causal gene underlying the 5A QTL. The 112 DE transcripts had functional annotations including non-coding RNA, transpon-associated, cell-cycle control, and ubiquitin-related processes. Many of the wheat genes identified belong to families that have been previously associated with seed/grain development in other species. However, few of these wheat genes are the direct orthologs and none have been previously characterised in wheat. Notably, we identified DE transcripts at almost all steps of the pathway associated with ubiquitin-mediated protein degradation. In the promoters of a subset of DE transcripts we identified enrichment of binding sites associated with C2H2, MYB/SANT, YABBY, AT HOOK and Trihelix transcription factor families.\n\nConclusionsIn this study, we identified DE transcripts with a diverse range of predicted biological functions, reflecting the complex nature of the pathways that control early grain development. Further functional characterisation of these candidates and how they interact could provide new insights into the control of grain size in cereals, ultimately improving crop yield.

plant biology↗

Afrothismiaceae (Dioscoreales), a new fully mycoheterotrophic family endemic to tropical Africa

Afrothismia is a genus of non-photosynthetic mycoheterotrophs from the forests of continental tropical Africa. Multiple phylogenetic inferences using molecular data recover the genus as sister to a clade comprising mycoheterotrophic Thismiaceae and the photosynthetic family Taccaceae, contrary to earlier placements of Afrothismia and Thismiaceae within Burmanniaceae. Morphological support for separating Afrothismia from the rest of Thismiaceae has depended on the zygomorphic flowers of Afrothismia (although some South American species of Thismia are also zygomorphic) and their clusters of root tubers, each with a terminal rootlet. The number of described species of Afrothismia has recently increased substantially, from four to 16, which has provided additional morphological characters that support its distinction from Thismiaceae. Most notably, the ovary in Afrothismia has a single stalked placenta, and circumscissile fruits from which seeds are exserted by placental elevation (in Thismiaceae, in contrast, there are three placentas, a deliquescing fruit lid, and the seeds are not exserted). Afrothismia stamens are inserted in the lower perianth tube where they are attached to the stigma, and individual flowers are subtended by a single large dorsal bract (in Thismiaceae, stamens are inserted at the mouth of the tube, free of and distant from the stigma, and each flower is subtended by a loose whorl of (2-)3(-4) bracts). Here we formally characterise Afrothismiaceae and review what is known of its development, seed germination, interactions with mycorrhizal Glomeromycota, biogeography, phylogeny and pollination biology. All but one (Afrothismia insignis; Vulnerable) of the 13 species assessed on the IUCN Red List of Threatened Species are either Endangered or Critically Endangered; one species (A. pachyantha Schltr.) is considered to be extinct.

plant biology↗

Transcriptome analysis of mycorrhizal and non-mycorrhizal soybean plantlets upon infection with Fusarium virguliforme, one causal agent of sudden death syndrome

Soilborne pathogens represent a threat to agriculture causing important yield losses. The \"Sudden Death Syndrome\" (SDS), a severe disease in soybean is caused by a complex of Fusarium species. This pathosystem has been widely investigated and several strategies were proposed to manage SDS. Although a decrease in symptoms and in the level of root tissue infection particularly by F. virguliforme was observed in presence of arbuscular mycorrhizal fungi (AMF), biological control based on AMF has received less attention. Here we report the results, under strict in vitro culture experimental conditions, a transcriptional analysis in mycorrhizal versus non-mycorrhizal soybean plantlets upon infection by F. virguliforme. An important transcriptional reprogramming was detected following infection by the pathogen. Results revealed 1768 and 967 differentially expressed genes in the AMF-colonized (+AMF+Fv) and non-colonized (-AMF+Fv) plants, respectively. Major transcriptional changes, corresponded to defence response related genes belonging to secondary metabolism, stress and signalling categories. The +AMF+Fv treatment showed the largest number of upregulated genes related to defence, as those encoding for disease resistance proteins, WRKY transcription factors, auxins, receptors kinases, and proteases. Only few genes had primed expression in +AMF+Fv treatment, as those coding for a thaumatin-like protein (TLP) and a pleiotropic drug resistance (PDR) protein. Moreover, +AMF+Fv showed a significant number of downregulated genes related to cell wall modification and peroxidases than - AMF+Fv treatment. This detailed insight increases our knowledge on the transcriptional changes and the potential metabolic pathways involved in the enhanced resistance/tolerance of mycorrhizal plants upon infection with F. virguliforme.

plant biology↗

SWEET11 and SWEET12 transporters function in tandem to modulate sugar flux in Arabidopsis: An account of the underlying unique structure-function relationship

Sugar will eventually be exported transporters (SWEETs) have been identified as a unique class of sugar efflux transporters in all biological kingdoms. AtSWEET11 and AtSWEET12 in Arabidopsis act synergistically to perform distinct physiological roles, particularly in apoplasmic phloem loading, seed filling, and sugar level alteration at the site of pathogen infection. Plasma membrane-localized AtSWEET11 and AtSWEET12 transporters exclusively facilitate sucrose transport along the concentration gradient. This article examines the sucrose binding pocket of AtSWEET11 and AtSWEET12 using docking studies, and how they act synergistically in various functions throughout plant development and during abiotic and biotic stresses. Further, we highlight the phylogenetic and the in-silico analyses of AtSWEET11 and AtSWEET12 orthologs from 39 economically important plant species that could provide new platforms for future studies on sugar allocation mechanisms across the different plant families. In-depth understanding of these transporters and their molecular regulatory mechanisms could be harnessed for crop improvement and crop protection.

plant biology↗

Protein Interaction Patterns in Arabidopsis thaliana Leaf Mitochondria Change in Response to Illumination

Mitochondrial biology is underpinned by the presence and activity of large protein complexes participating in the organelle-located parts of cellular respiration, the TCA cycle and oxidative phosphorylation. While the enzymatic roles of these complexes are undisputed, little is known about the interactions of the subunits beyond their presence in the monomeric protein complexes and their functions in regulating mitochondria metabolism. By applying one of the most important regulatory cues for plant metabolism, the presence or absence of light, we here assess the changes in the composition and molecular mass of known mitochondrial protein complexes by employing a differential complexome profiling strategy. Covering a mass range up to 25 MDa, we demonstrate dynamic associations of TCA-cycle enzymes and of OXPHOS components. The data presented here form the basis for future studies aiming to advance our understanding of the role of protein:protein interactions in the regulation of plant mitochondrial functions.

plant biology↗

Genome-wide association mapping of bruchid resistance loci in soybean

Soybean is a globally important industrial, food, and cash crop. Despite its importance in present and future economies, its production is severely hampered by bruchids (Callosobruchus chinensis), a destructive storage insect pest, causing considerable yield losses. Therefore, the identification of genomic regions and candidate genes associated with bruchid resistance in soybean is crucial as it helps breeders develop new soybean varieties with improved resistance and quality. In this study, 6 multi-locus methods of the mrMLM model for genome-wide association study were used to dissect the genetic architecture of bruchid resistance using 5 traits: percentage adult bruchid emergence (PBE), percentage weight loss (PWL), growth index (GI), median development period (MDP), and Dobie susceptibility index (DSI) on 100 diverse soybean genotypes, genotyped with 14,469 single-nucleotide polymorphism (SNP) markers. Using the best linear unbiased predictors (BLUPs), 13 quantitative trait nucleotides (QTNs) were identified by the mrMLM model, 3 of which, rs16_14976250, rs1_22916615, and rs16_14975721, were associated with more than 1 bruchid resistance trait. As a result, the identified QTNs linked with resistance traits can be employed in marker-assisted breeding for the accurate and rapid screening of soybean genotypes for resistance to bruchids. Moreover, a gene search on the Phytozome soybean reference genome identified 27 potential candidate genes located within a window of 478.45 kb upstream and downstream of the most reliable QTNs. These candidate genes exhibit molecular and biological functionalities associated with various soybean resistance mechanisms and, therefore, could be incorporated into the farmers preferred soybean varieties that are susceptible to bruchids.

plant biology↗

Immunovisualization of spatial changes in leaves and root tissue associated with drought stress in wheat (Triticum aestivum L.)

Background and AimsPlants have evolved complex cell-type-specific processes to adapt to a dynamic environment, exhibiting distinct signals in response to emerging drought stress. We propose an advanced qualitative and quantitative analysis approach, demonstrating tissue specificity in drought adaptation, which in turn may provide novel biological insights. MethodsWe performed immunofluorescence labeling of specific cellular components in situ, and the acquired data were analyzed in terms of changes in quantitative and spatial fluorescence intensity. ResultsThe qualitative analysis revealed differences in terms of individual components and individual days of the experiment. The quantitative analysis of leaf anatomy showed that the most pronounced changes were observed in the level of proteoglycans (JIM13, JIM15) and polysaccharides (LM5, LM16, LM20). The leaves of plants growing in drought were characterized by destroyed fragments, in which increased secretion of extensins, AGPs, galactans, hemicelluloses, and RG-I was noted. In turn, the qualitative analyses of the microscopy images of roots, along with fluorescence intensity analyses, revealed a significantly higher content of AGP and arabinoxylan in the exodermis in plants grown under drought stress. ConclusionOur research has revealed that the changes at the tissue level are targeted and highly specific. The obtained results also emphasize the importance of in planta analyses, which indicate that findings from only single ex planta studies may distort the entire image of changes occurring in the plant as a result of stress. HIGHLIGHT STATEMENTOne of the strategies employed by plants to mitigate the effects of water loss is the mechanical protection of organs through targeted and highly specific modifications in their cellular architecture.

plant biology↗

YamOmics: A comprehensive data resource on yam multi-omics

Yams (Dioscorea spp.) are a highly important class of horticultural crops, serving as a staple food for millions of people in Africa and contributing significantly to food security. They are also widely cultivated in East Asia as medicinal herbs, bringing substantial economic incomes. Diverse omics data play a pivotal role in advancing yam research and breeding. However, these data are often scattered, lacking in systematic organization and analysis, which underscores the need for centralized and comprehensive data management. In view of this, we gathered extensive omics data and developed the Yam Omics Database (YamOmics; https://biotec.njau.edu.cn/yamdb). The database currently offers a vast and diverse range of omics data, covering genomic, transcriptomic and plastomic data from 41 distinct yam species, along with detailed records of genomic variants from [~]1000 germplasms, and gene expression profiles from [~]200 samples. Additionally, the database features thorough annotations, encompassing aspects like genome synteny, ortholog groups, signaling pathways, gene families and protein interactions. To support yam basic biology and breeding research, it is also equipped with a suite of user-friendly online tools, including PCR primer design, CRISPR design, expression analysis, enrichment analysis, and kinship analysis tools.

plant biology↗

Hijacking host microPEP: pathogens modulate the microRNA-microPEP loop to promote infection

The partners of an ecological association tend to copy the biological system of their hosts. We hypothesized that microorganisms, particularly pathogens, have acquired the ability to express short peptides (pathoPEPs) homologous to host microPEPs (miPEPs) thus modulating the expression of the corresponding host microRNA (miRNA) and the function of miRNA-targeted genes. The pathosystem involving interactions between Brassica napus and its pathogen Plasmodiophora brassicae was studied. Using in silico analysis and ribosomal profiling, we identified three putative pathoPEPs produced by P. brassicae and their targeted plant miRNA genes. A link between the level of infection of B. napus by P. brassicae and the expression of pathoPEPs and their targeted miRNA genes was found, with the expression of the latter two being inversely related. Finally, we identified differential expression and translation of genes predicted to be targets of pathoPEP-regulated miRNAs. These genes, involved in auxin pathway, immune defense, root architecture or carbohydrate metabolism, are thought to enable P. brassicae, through its pathoPEPs, to hijack plants metabolic pathways (hormonal pathways, sugar synthesis, root morphology), thereby facilitating its invasion. Using computational in silico approaches, the involvement of miPEPs from plant pathogens as a host post-transcriptional regulatory pathway is described herein for the first time.

plant biology↗

Inhibition of auxin biosynthesis in early rice grains leads to extensive post-fertilisation grain abortion

In spite of its abundant presence in rice grains, auxin functions during grain development are not understood well. Absence of grain-specific auxin biosynthesis mutants in rice further limits our understanding in this respect. Here, we report a chemical biology approach to inhibit auxin biosynthesis specifically in early rice grains as well as its subsequent effects on final grain yield. Exogenous application of two auxin biosynthesis inhibitors, namely L-kynurenine (L-Kyn) and 4-phenoxyphenylboronic acid (PPBo), to spikelets daily from three to ten days after pollination (DAP) significantly reduced grain levels of indole-3-acetic acid (IAA), the predominant in planta auxin. The inhibitor-treated panicles showed extensive post-fertilisation seed abortion, leading to drastic reduction in total panicle weight at maturity. Locally synthesised auxin during early grain development may therefore play a crucial role in grain retention. This chemical biology approach can be an easy and cost-effective way to study auxin biosynthesis and signalling during grain development in rice and possibly other cereal crops. HighlightAuxin synthesised in early rice grains may play a crucial role in grain retention.

plant biology↗

The Activity of the Stress Modulated Arabidopsis Ubiquitin Ligases PUB46 and PUB48 is Partially Redundant

The Arabidopsis ubiquitin ligases PUB46, PUB47 and PUB48 are encoded by paralogus genes; pub46 and pub48 mutants display increased drought sensitivity compared to wild type (WT). Although the phenotype displayed in the single gene mutants, suggest that each has specific biological activity, PUB46 and PUB48 activity may be also redundant. To test functional redundancy between two gene products requires a double mutant. Unfortunately, the close proximity of the PUB46 and PUB48 gene loci precludes obtaining a double mutant by crossing the available single mutants. We thus applied microRNA technology to reduce the activity of all three gene products of the PUB46-48 subfamily by constructing an artificial microRNA (aMIR) targeted to this subfamily. Expressing aMIR46-48 in WT plants resulted in increased drought-sensitivity, a phenotype resembling that of the single pub46 and pub48 mutants, and enhanced sensitivity to methyl viologen, similar to that observed for the pub46 mutant. Furthermore, the WT plants expressing aMIR46-48 also revealed reduced inhibition by ABA at seed germination, a phenotype not evident in the single mutants. Expressing aMIR46-48 in pub46 and pub48 mutants further enhanced the drought sensitivity of each parental single mutant and of WT expressing aMIR46-48. Thus, whereas the gene-specific activity of the PUB46 and PUB48 E3s is partially redundant in that absence of either E3 leads to drought sensitivity, our ability to eliminate the activity of both PUB46 and PUB48 in the same plant reveals additional gene specific facets of their activity in the reaction to abiotic stress.

plant biology↗