Plant transformation remains a major bottleneck to the improvement of plant science, both on fundamental and practical levels. The recalcitrant nature of most commercial and minor crops to genetic transformation slows scientific progress for a large range of crops that are essential for food security on a global scale. Over the years, novel stable transformation strategies loosely grouped under the term “in planta” have been proposed and validated in a large number of model (e.g. Arabidopsis and rice), major (e.g. wheat and soybean) and minor (e.g. chickpea and lablab bean) species. The in planta approach is revolutionary as it is considered genotype-independent, technically simple (i.e. devoid of or with minimal tissue culture steps), affordable, and easy to implement in a broad range of experimental settings. In this article, we reviewed and categorized over 300 research articles, patents, theses, and videos demonstrating the applicability of different in planta transformation strategies in 105 different genera across 139 plant species. To support this review process, we propose a classification system for the in planta techniques based on five categories and a new nomenclature for more than 30 different in planta techniques. In complement to this, we clarified some grey areas regarding the in planta conceptual framework and provided insights regarding the past, current, and future scientific impacts of these techniques. To support the diffusion of this concept across the community, this review article will serve as an introductory point for an online compendium about in planta transformation strategies that will be available to all scientists. By expanding our knowledge about in planta transformation, we can find innovative approaches to unlock the full potential of plants, support the growth of scientific knowledge, and stimulate an equitable development of plant research in all countries and institutions.

EDITORIAL article Front. Plant Sci., 20 June 2023Sec. Plant Cell Biology Volume 14 - 2023 | https://doi.org/10.3389/fpls.2023.1213511

Fusarium graminearum is the causal agent of Fusarium Head Blight, a serious disease affecting grain crops worldwide. Biological control involves the use of microorganisms to combat plant pathogens such as F. graminearum. Strains of Bacillus velezensis are common biological control candidates for use against F. graminearum and other plant pathogens, as they can secrete antifungal secondary metabolites. Here we study the interaction between B. velezensis E68 and F. graminearum DAOMC 180378 by employing a dual RNA-seq approach to assess the transcriptional changes in both organisms. In dual culture, B. velezensis up-regulated genes related to sporulation and phosphate stress and down-regulated genes related to secondary metabolism, biofilm formation and the tricarboxylic acid cycle. F. graminearum up-regulated genes encoding for killer protein 4-like proteins and genes relating to heavy metal tolerance, and down-regulated genes relating to trichothecene biosynthesis and phenol metabolism. This study provides insight into the molecular mechanisms involved in the interaction between a biocontrol bacterium and a phytopathogenic fungus.

Plant growth-promoting rhizobacteria (PGPR) influence plant health. However, the genotypic variations in host organisms affect their response to PGPR. To understand the genotypic effect, we screened four diverse B. distachyon genotypes at varying growth stages for their ability to be colonized by B. velezensis strain B26 . We reasoned that B26 may have an impact on the phenological growth stages of B. distachyon genotypes. Phenotypic data suggested the role of B26 in increasing the number of awns and root weight in wild type genotypes and overexpressing transgenic lines. Thus, we characterized the expression patterns of flowering pathway genes in inoculated plants and found that strain B26 modulates the transcript abundance of flowering genes. An increased root volume of inoculated plants was estimated by CT-scanning which suggests the role of B26 in altering the root architecture. B26 also modulated plant hormone homeostasis. A differential response was observed in the transcript abundance of auxin and gibberellins biosynthesis genes in inoculated roots. Our results reveal that B. distachyon plant genotype is an essential determinant of whether a PGPR provides benefit or harm to the host and shed new insight into the involvement of B. velezensis in the expression of flowering genes.

Global demand for pesticide-free food products is increasing rapidly. Crops of all types are, however, under constant threat from various plant pathogens. To achieve adequate control with minimal pesticide use, close monitoring is imperative. Many plant pathogens spread through the air, so the atmosphere is composed of a wide variety of plant pathogenic and non-plant pathogenic organisms, in particular in agricultural environments. Aerobiology is the science that studies airborne microorganisms and their distribution, especially as agents of infection. Although this discipline has existed for decades, the development of new molecular technologies is contributing to an increase in the use of aerobiological data for several purposes, from day-to-day monitoring to improving our understanding of pathosystems. Although the importance of knowing the size and composition of plant pathogen populations present in the air is recognized, technical constraints hinder the development of agricultural aerobiology. Here we review the application of spore sampling systems in agriculture and discuss the main considerations underlying the implementation of airborne inoculum monitoring. The results of this literature review confirm that the use of aerobiological data to study the escape of inoculum from a source and its role in the development of diseases is well mastered, but point at a lack of knowledge to proceed with the deployment of these systems at the landscape scale. Thus, we conclude that airborne inoculum surveillance networks are still in their early stages and although more and more initiatives are emerging, research must be conducted primarily to integrate evolving technologies and improve the access, analysis, interpretation and sharing of data. These tools are needed to estimate short- and medium-term risks, identify the most appropriate control measures with the lowest environmental risk, develop indicators to document the effects of climate change, and monitor the evolution of new genotypes at multiple scales.

Several Peronospora species are carried by wind over short and long distances, from warmer climates where they survive on living plants to cooler climates. In eastern Canada, this annual flow of sporangia was thought to be the main source of Peronospora destructor responsible for onion downy mildew. However, the results of a recent study showed that the increasing frequency of onion downy mildew epidemics in eastern Canada is associated with warmer autumns, milder winters, and previous year disease severity, suggesting overwintering of the inoculum in an area where the pathogen is not known to be endogenous. In this study, genotyping by sequencing was used to investigate the population structure of P. destructor at the landscape scale. The study focused on a particular region of southwestern Québec-Les Jardins de Napierville-to determine if the populations were clonal and regionally differentiated. The data were characterized by a high level of linkage disequilibrium, characteristic of clonal organisms. Consequently, the null hypothesis of random mating was rejected when tested on predefined or nonpredefined populations, indicating that linkage disequilibrium was not a function of population structure and suggesting a mixed reproduction mode. Discriminant analysis of principal components performed with predefined population assignment allowed grouping P. destructor isolates by geographical regions, while analysis of molecular variance confirmed that this genetic differentiation was significant at the regional level. Without using a priori population assignment, isolates were clustered into four genetic clusters. These results represent a baseline estimate of the genetic diversity and population structure of P. destructor.

Plant growth-promoting rhizobacteria (PGPR) are associated with plant roots and use organic compounds that are secreted from root exudates as food and energy source. Root exudates can chemoattract and help bacteria to colonize the surface of plant roots by inducing chemotactic responses of rhizospheric bacteria. In this study, we show that root colonization of Brachypodium distachyon by Bacillus velezensis strain B26 depends on several factors. These include root exudates, organic acids, and their biosynthetic genes, chemotaxis, biofilm formation and the induction of biofilm encoding genes. Analysis of root exudates by GC-MS identified five intermediates of the TCA cycle; malic, fumaric, citric, succinic, oxaloacetic acids, and were subsequently evaluated. The strongest chemotactic responses were induced by malic, succinic, citric, and fumaric acids. In comparison, the biofilm formation was induced by all organic acids with maximal induction by citric acid. Relative to the control, the individual organic acids, succinic and citric acids activated the epsD gene related to EPS biofilm, and also the genes encoding membrane protein (yqXM) and hydrophobin component (bslA) of the biofilm of strain B26. Whereas epsA and epsB genes were highly induced genes by succinic acid. Similarly, concentrated exudates released from inoculated roots after 48 h post-inoculation also induced all biofilm-associated genes. The addition of strain B26 to wild type and to icdh mutant line led to a slight induction but not biologically significant relative to their respective controls. Thus, B26 has no effect on the expression of the ICDH gene, both in the wild type and the mutant backgrounds. Our results indicate that root exudates and individual organic acids play an important role in selective recruitment and colonization of PGPR and inducing biofilm. The current study increases the understanding of molecular mechanisms behind biofilm induction by organic acids.

Summary Plants that successfully acclimate to stress can resume growth under stressful conditions. The grass Brachypodium distachyon can grow a cold‐adaptive morphology during cold acclimation. Studies on transcriptional memory (TM) have revealed that plants can be primed for stress by adjusting their transcriptional responses, but the function of TM in stress acclimation is not well understood. We investigated the function of TM during cold acclimation in B. distachyon. Quantitative polymerase chain reaction (qPCR), RNA‐seq and chromatin immunoprecipitation qPCR analyses were performed on plants exposed to repeated episodes of cold to characterize the presence and stability of TM during the stress and growth responses of cold acclimation. Transcriptional memory mainly dampened stress responses as growth resumed and as B. distachyon became habituated to cold stress. Although permanent on vernalization gene VRN1, TMs were short‐term and reversible on cold‐stress genes. Growing under cold conditions also coincided with the acquisition of new and targeted cold‐induced transcriptional responses. Overall, TM provided plasticity to cold stress responses during cold acclimation in B. distachyon, leading to stress habituation, acquired stress responses, and resumed growth. Our study shows that chromatin‐associated TMs are involved in tuning plant responses to environmental change and, as such, regulate both stress and developmental components that characterize cold‐climate adaptation in B. distachyon.

Onion downy mildew (ODM) caused by Peronospora destructor has been increasing annually in south-western Québec since the early 2000s, reaching 33% of affected onion fields in 2014. Using observational data collected over a period of 31 consecutive years, this study aimed to investigate the variations in ODM incidence and epidemic onset and identify the meteorological variables that influence its polyetic development. A logistic model was fitted to each ODM epidemic to estimate and compare the onset of epidemics on a regional basis. Results of this analysis showed that the first observation date, 10% epidemic onset (b10) and mid-time (b) were, on average, 30.4, 15.1 and 11.3 days earlier in 2007–2017 than in 1987–1996. Results of a principal component analysis suggested that regional disease incidence was mostly influenced by the precipitation regime, the final regional disease incidence the previous year, and warmer temperature during the harvest period the previous fall. Subsequently, the data were divided in three periods of 10, 10 and 11 years, and a discriminant analysis was performed to classify each year in the correct period. Using a sufficient subset of five discriminating variables (temperature and rainfall at harvest the previous fall, winter coldness, solar radiation, and disease incidence the previous year), it was possible to classify 93.5% of the ODM epidemics in the period where they belong. These results suggest that P. destructor may overwinter under northern latitudes and help to highlight the need for more research on overwintering and for the development of molecular-based tools enabling the monitoring of initial and secondary inoculum.

Onion downy mildew (ODM), caused by Peronospora destructor, is a serious threat for onion growers worldwide. In southwestern Québec, Canada, a steady increase in occurrence of ODM has been observed since the mid-2000s. On onion, P. destructor can develop local and systemic infections producing numerous sporangia which act as initial inoculum locally and also for neighboring areas. It also produces oospores capable of surviving in soils and tissues for a prolonged period of time. A recent study showed that ODM epidemics are strongly associated with weather conditions related to production and survival of overwintering inoculum, stressing the need to understand the role of primary (initial) and secondary inoculum. However, P. destructor is an obligate biotrophic pathogen, which complicates the study of inoculum sources. This study aimed at developing a molecular assay specific to P. destructor, allowing its quantification in environmental samples. In this study, a reliable and sensitive hydrolysis probe-based assay multiplexed with an internal control was developed on the internal transcribed spacer (ITS) region to quantify soil- and airborne inoculum of P. destructor. The assay specificity was tested against 17 isolates of P. destructor obtained from different locations worldwide, other members of the order Peronosporales, and various onion pathogens. Validation with artificially inoculated soil and air samples suggested a sensitivity of less than 10 sporangia g-1 of dry soil and 1 sporangium m-3 of air. Validation with environmental air samples shows a linear relationship between microscopic and real-time quantitative PCR counts. In naturally infested soils, inoculum ranged from 0 to 162 sporangia equivalent g-1 of dry soil, which supported the hypothesis of overwintering under northern climates. This assay will be useful for primary and secondary inoculum monitoring to help characterize ODM epidemiology and could be used for daily tactical and short-term strategic decision-making.

Anthropogenic climate change precipitates the need to understand plant adaptation. Crucial in temperate climates, adaptation to winter is characterized by cold acclimation and vernalization, which respectively lead to freezing tolerance and flowering competence. However, the progression of these responses during fall and their interaction with plant development are not completely understood. By identifying key seasonal cues found in the native range of the cereal model Brachypodium distachyon, we designed a diurnal-freezing treatment (DF) that emulates summer-to-winter change. DF induced unique cold acclimation and vernalization responses characterized by low VERNALIZATION1 (VRN1) expression. Flowering under DF is characterized by an up-regulation of FLOWERING LOCUS T (FT) postvernalization independent of VRN1 expression. DF, while conferring flowering competence, favors a high tolerance to freezing and the development of a winter-hardy plant structure. The findings of this study highlight the contribution of phenotypic plasticity to freezing tolerance and demonstrate the integration of key morphological, physiological, and molecular responses in cold adaptation. The results suggest a fundamental role for VRN1 in regulating cold acclimation, vernalization, and morphological development in B. distachyon This study also establishes the usefulness of reproducing natural cues in laboratory settings.

Histone methylation plays a fundamental role in the epigenetic regulation of gene expression driven by developmental and environmental cues in plants, including Arabidopsis. Histone methyltransferases and demethylases act as 'writers' and 'erasers' of methylation at lysine and/or arginine residues of core histones, respectively. A third group of proteins, the 'readers', recognize and interpret the methylation marks. Emerging evidence confirms the crucial roles of histone methylation in multiple biological processes throughout the plant life cycle. In this review, we summarize the regulatory mechanisms of lysine methylation, especially at histone H3 tails, and focus on the recent advances regarding the roles of lysine methylation in Arabidopsis development, from seed performance to reproductive development, and in callus formation.

Crop plant resistance against pathogens is governed by dynamic molecular and biochemical responses driven by complex transcriptional networks. However, the underlying mechanisms are largely unclear. Here we report an interesting role of HvWRKY23 transcription factor (TF) in modulating defense response against Fusarium head blight (FHB) in barley. The combined approach of gene silencing, metabolomics, real time expression analysis and ab initio bioinformatics tools led to the identification of the HvWRKY23 role in FHB resistance. The knock-down of HvWRKY23 gene in the FHB resistant barley genotype CI9831, followed by inoculation with Fusarium graminearum, led to the down regulation of key flavonoid and hydroxycinnamic acid amide biosynthetic genes resulting in reduced accumulation of resistant related (RR) secondary metabolites such as pelargonidin 3-rutinoside, peonidin 3-rhamnoside-5-glucoside, kaempferol 3-O-arabinoside and other flavonoid glycosides. Reduced abundances of RR metabolites in TF silenced plants were also associated with an increased proportion of spikelets diseased and amount of fungal biomass in spikelets, depicting the role of HvWRKY23 in disease resistance. The luciferase reporter assay demonstrated binding of HvWRKY23 protein to promoters of key flavonoid and hydroxycinnamic acid amides (HCAA) biosynthetic genes, such as HvPAL2, HvCHS1, HvHCT, HvLAC15 and HvUDPGT. The accumulation of high abundances of HCAAs and flavonoid glycosides reinforce cell walls to contain the pathogen to initial infection area. This gene in commercial cultivars can be edited, if non-functional, to enhance resistance against FHB.

Plant endophytes are a group of microorganisms that reside asymptomatically within the healthy living tissue. The diversity and molecular and biochemical characterization of industrial hemp-associated endophytes have not been previously studied. This study explored the abundance and diversity of culturable endophytes residing in petioles, leaves, and seeds of three industrial hemp cultivars, and examined their biochemical attributes and antifungal potential. A total of 134 bacterial and 53 fungal strains were isolated from cultivars Anka, CRS-1, and Yvonne. The number of bacterial isolates was similarly distributed among the cultivars, with the majority recovered from petiole tissue. Most fungal strains originated from leaf tissue of cultivar Anka. Molecular and phylogenetic analyses grouped the endophytes into 18 bacterial and 13 fungal taxa, respectively. The most abundant bacterial genera were Pseudomonas, Pantoea, and Bacillus, and the fungal genera were Aureobasidium, Alternaria, and Cochliobolus. The presence of siderophores, cellulase production, and phosphorus solubilization were the main biochemical traits. In proof-of-concept experiments, re-inoculation of tomato roots with some endophytes confirmed their migration to aerial tissues of the plant. Taken together, this study demonstrates that industrial hemp harbours a diversity of microbial endophytes, some of which could be used in growth promotion and (or) in biological control designed experiments.

BACKGROUND:Histone deacetylases (HDACs) function as key epigenetic factors in repressing the expression of genes in multiple aspects of plant growth, development and plant response to abiotic or biotic stresses. To date, the molecular function of HDACs is well described in Arabidopsis thaliana, but no systematic analysis of this gene family in soybean (Glycine max) has been reported.RESULTS:In this study, 28 HDAC genes from soybean genome were identified, which were asymmetrically distributed on 12 chromosomes. Phylogenetic analysis demonstrated that GmHDACs fall into three major groups previously named RPD3/HDA1, SIR2, and HD2. Subcellular localization analysis revealed that YFP-tagged GmSRT4, GmHDT2 and GmHDT4 were predominantly localized in the nucleus, whereas GmHDA6, GmHDA13, GmHDA14 and GmHDA16 were found in both the cytoplasm and nucleus. Real-time quantitative PCR showed that GmHDA6, GmHDA13, GmHDA14, GmHDA16 and GmHDT4 were broadly expressed across plant tissues, while GmHDA8, GmSRT2, GmSRT4 and GmHDT2 showed differential expression across various tissues. Interestingly, we measured differential changes in GmHDACs transcripts accumulation in response to several abiotic cues, indicating that these epigenetic modifiers could potentially be part of a dynamic transcriptional response to stress in soybean. Finally, we show that the levels of histone marks previously reported to be associated with plant HDACs are modulated by cold and heat in this legume.CONCLUSION:We have identified and classified 28 HDAC genes in soybean. Our data provides insights into the evolution of the HDAC gene family and further support the hypothesis that these genes are important for the plant responses to environmental stress.

Tomato bacterial canker disease, caused by Clavibacter michiganensis subsp. michiganensis (Cmm) is a destructive disease and has been a serious concern for tomato industries worldwide. Previously, a rhizosphere isolated strain of Pseudomonas sp. 23S showed antagonistic activity toward Cmm in vitro. This Pseudomonas sp. 23S was characterized to explore the potential of this bacterium for its use in agriculture. Pseudomonas sp. 23S possesses ability to solubilize inorganic phosphorus, and to produce siderophores, indole acetic acid, and hydrogen cyanide. The strain also showed antagonistic activity against Pseudomonas syringae pv. tomato DC 3000. A plant assay indicated that Pseudomonas sp. 23S could promote growth of tomato seedlings. The potential of treating tomato plants with Pseudomonas sp. 23S to reduce the severity of tomato bacterial canker by inducing systemic resistance (ISR) was investigated using well characterized marker genes such as PR1a [salicylic acid (SA)], PI2 [jasmonic acid (JA)], and ACO [ethylene (ET)]. Two-week-old tomato plants were treated with Pseudomonas sp. 23S by soil drench, and Cmm was inoculated into the stem by needle injection on 3, 5, or 7 days post drench. The results indicated that plants treated with Pseudomonas sp. 23S, 5 days prior to Cmm inoculation significantly delayed the progression of the disease. These plants, after 3 weeks from the date of Cmm inoculation, had significantly higher dry shoot and root weight, higher levels of carbon, nitrogen, phosphorus, and potassium in the leaf tissue, and the number of Cmm population in the stem was significantly lower for the plants treated with Pseudomonas sp. 23S. From the real-time quantitative PCR (qRT-PCR) analysis, the treatment with Pseudomonas sp. 23S alone was found to trigger a significant increase in the level of PR1a transcripts in tomato plants. When the plants were treated with Pseudomonas sp. 23S and inoculated with Cmm, the level of PR1a and ACO transcripts were increased, and this response was faster and greater as compared to plants inoculated with Cmm but not treated with Pseudomonas sp. 23S. Overall, the results suggested the involvement of SA signaling pathways for ISR induced by Pseudomonas sp. 23S.

The objective of this study was to establish the nitrogen, phosphorus, and potassium fertilization requirements of industrial hemp grown in eastern Canada. Earn 0.5 CEUs in Nutrient Management by reading this article and taking the quiz at www.certifiedcropadviser.org/certifications/self-study/780 .

Industrial hemp ( Cannabis sativa L.) is a multipurpose crop for which there is growing interest in Canada. There is, however, currently limited information on the adaptation and production potential of government approved cultivars in eastern Canada. Eleven cultivars were evaluated in terms of biomass and seed yields and composition, when grown in seven contrasted environments in the province of Québec. Dry matter yields varied considerably across cultivars and environments, ranging between 172 and 8837 kg ha −1 (average of 3226 kg ha −1 ) for biomass, and between 47 and 3781 kg ha −1 (average of 1315 kg ha −1 ) for seeds. Despite the presence of cultivar × environment interactions for both biomass and seed yields, the cultivars Anka, Ferimon, and Jutta consistently produced high yields across environments. Averaged across environments, these cultivars respectively had biomass and seed yields of 4591 and 1397, 5827, and 1878, and 4096 and 1408 kg DM ha −1 . Limited variation was observed among cultivars in terms of composition. Cellulose was the largest biomass constituent (average of 562 g kg −1 DM across cultivars and environments), followed by hemicellulose and lignin (averages of 123 and 93 g kg −1 DM, respectively), while seed crude protein (CP) and oil (ether extract) concentrations averaged 237 and 288 g kg −1 DM, respectively. Considerable variation in yield potential and adaptation were observed among the cultivars of industrial hemp evaluated in the province of Québec; while biomass yields were moderate, the seed yield potential appears to be high locally compared to western Canada. Core Ideas Biomass and seed dry matter yields of the 11 industrial hemp cultivars evaluated varied considerably across cultivars and environments. Despite the presence of cultivar × environment interactions for biomass and seed yields, some cultivars consistently produced high yields of both biomass and seeds. Limited variation was observed among cultivars in terms of biomass and seed chemical composition.

Industrial hemp (Cannabis sativa L.) production has increased in Canada in recent years, and interest for this multipurpose crop remains high. The lack of agronomic guidelines for Eastern Canada represents, however, a limiting factor for local hemp production. This study assessed biomass and seed yields and composition of two hemp cultivars (CRS-1 and Anka), following various N, P, and K fertilization treatments (0, 50, 100, 150, and 200 kg N or K ha(-1); 0, 25, 50, 75, and 100 kg P ha(-1)). The experiment was conducted in multiple environments in the province of Quebec. Positive linear and quadratic responses of biomass yield, seed yield, and seed crude protein concentration to N fertilization were observed in all environments; the magnitude of the response depended, however, on the environment and cultivar. Across environments and cultivars, biomass and seed yields increased from 1674 to 4209 kg ha(-1) and from 519 to 1340 kg ha(-1), respectively, with the application of 200 kg N ha(-1) when compared with the unfertilized control. Nitrogen fertilization affected biomass cellulose and hemicellulose concentrations, but the overall response remained minimal. Phosphorus and potassium fertilizations had very limited effect on biomass and seed yields and composition in all environments. In conclusion, while industrial hemp responded to N fertilization up to 200 kg N ha(-1), response to P and K fertilization remained limited.