Plant Research Research Articles

Evaluating the efficiency of Chelex 100 for DNA extraction in orange (Citrus sinensis)

Plant research has recently concentrated on the molecular level, which requires reliable, inexpensive, and fast protocols for extracting DNA. Many researchers have identified Chelex-based DNA extraction methods as having the potential to reduce costs. However, Chelex can also prevent DNA degradation by chelating metal ions. Citrus is one of the top three crops grown globally, with oranges being a well-known species. Therefore, study on the molecular of C. sinensis is essential. This study was conducted in 2022 at the genetic lab of the University of Limpopo- to compare two different extraction methods for DNA from C. sinensis. The DNA from the C. sinensis leaves was extracted using Chelex "method 1" (overnight incubation at 56°C) and "method 2" (Ten-minute incubation at 95°C). A spectrophotometer was employed to measure the extracted DNA's qualitative and quantitative parameters accurately. Furthermore, all samples were subjected to PCR reactions with primers for 28S. The results indicated that "method 2" (1.59-1.84) proved to be the most effective method to obtain high-quality extracted DNA. Additionally, the quantitative and qualitative tests for PCR reaction demonstrated that "method 2" produced better results. In conclusion, although both methods yielded favourable outcomes, "method 2" was ultimately the more successful approach.

Revisiting the Role of Sensors for Shaping Plant Research: Applications and Future Perspectives.

Plant health monitoring is essential for understanding the impact of environmental stressors (biotic and abiotic) on crop production, and for tailoring plant developmental and adaptive responses accordingly. Plants are constantly exposed to different stressors like pathogens and soil pollutants (heavy metals and pesticides) which pose a serious threat to their survival and to human health. Plants have the ability to respond to environmental stressors by undergoing rapid transcriptional, translational, and metabolic reprogramming at different cellular compartments in order to balance growth and adaptive responses. However, plants' exceptional responsiveness to environmental cues is highly complex, which is driven by diverse signaling molecules such as calcium Ca2+, reactive oxygen species (ROS), hormones, small peptides and metabolites. Additionally, other factors like pH also influence these responses. The regulation and occurrence of these plant signaling molecules are often undetectable, necessitating nondestructive, live research approaches to understand their molecular complexity and functional traits during growth and stress conditions. With the advent of sensors, in vivo and in vitro understanding of some of these processes associated with plant physiology, signaling, metabolism, and development has provided a novel platform not only for decoding the biochemical complexity of signaling pathways but also for targeted engineering to improve diverse plant traits. The application of sensors in detecting pathogens and soil pollutants like heavy metal and pesticides plays a key role in protecting plant and human health. In this review, we provide an update on sensors used in plant biology for the detection of diverse signaling molecules and their functional attributes. We also discuss different types of sensors (biosensors and nanosensors) used in agriculture for detecting pesticides, pathogens and pollutants.

Advances in Endangered Plant Research: Ammopiptanthus’s Responses to Biotic and Abiotic Stressors

Ammopiptanthus, a unique evergreen broadleaf shrub endemic to the desert regions of Northwest China, exhibits a remarkable capacity for aeolian erosion control and dune stabilization. Attributed to its robust tolerance in terms of xerotolerance, halophytic adaptations, extreme thermotolerance, resistance to biotic and abiotic degradation and its defensive strategies against herbivory, Ammopiptanthus has emerged as an exemplary model organism for the study of plant resilience to diverse environmental stressors. Current research on Ammopiptanthus is scattered, lacking a systematic review, which poses a disadvantage for subsequent in-depth studies and the effective conservation of this endangered resource. In recent years, natural Ammopiptanthus communities have been severely disrupted, and the species’ natural range is rapidly shrinking. Here, this review summarizes the signaling pathways in the Ammopiptanthus response to biotic stress (especially the early signaling events), as well as the research advances in the resistance interactions between biotic and abiotic stresses. Then, the synergistic effects of multiple environmental pressures on Ammopiptanthus could be established, which may provide guidance for further studies on the resistance mechanism of Ammopiptanthus and be beneficial to its natural community protection and reconstruction.

Organ Segmentation and Phenotypic Trait Extraction of Cotton Seedling Point Clouds Based on a 3D Lightweight Network

Cotton is an important economic crop; therefore, enhancing cotton yield and cultivating superior varieties are key research priorities. The seedling stage, a critical phase in cotton production, significantly influences the subsequent growth and yield of the crop. Therefore, breeding experts often choose to measure phenotypic parameters during this period to make breeding decisions. Traditional methods of phenotypic parameter measurement require manual processes, which are not only tedious and inefficient but can also damage the plants. To effectively, rapidly, and accurately extract three-dimensional phenotypic parameters of cotton seedlings, precise segmentation of phenotypic organs must first be achieved. This paper proposes a neural network-based segmentation algorithm for cotton seedling organs, which, compared to the average precision of 75.4% in traditional unsupervised learning, achieves an average precision of 96.67%, demonstrating excellent segmentation performance. The segmented leaf and stem point clouds are used for the calculation of phenotypic parameters such as stem length, leaf length, leaf width, and leaf area. Comparisons with actual measurements yield coefficients of determination R2 of 91.97%, 90.97%, 92.72%, and 95.44%, respectively. The results indicate that the algorithm proposed in this paper can achieve precise segmentation of stem and leaf organs, and can efficiently and accurately extract three-dimensional phenotypic structural information of cotton seedlings. In summary, this study not only made significant progress in the precise segmentation of cotton seedling organs and the extraction of three-dimensional phenotypic structural information, but the algorithm also demonstrates strong applicability to different varieties of cotton seedlings. This provides new perspectives and methods for plant researchers and breeding experts, contributing to the advancement of the plant phenotypic computation field and bringing new breakthroughs and opportunities to the field of plant science research.

The Function of RNA Interference (RNAi) in Crop Enhancement- Mechanism and Applications: A Review

RNA interference (RNAi) is a conserved molecular mechanism that plays a critical role in post-transcriptional gene silencing across diverse organisms. This review delves into the role of RNAi in plant functional genomics and its applications in crop improvement, highlighting its mechanistic insights and practical implications. The overview starts with the fundamental finding of the mechanism of RNAi, following its journey from petunias to its ubiquitous presence in a wide range of species. The discovery of several kinds of regulatory non-coding small RNAs, including as phasiRNAs, miRNAs, and siRNAs, has broadened our knowledge of RNAi-mediated gene regulation beyond what is previously thought. These RNA types have an impact on gene expression through complex post-transcriptional and epigenetic mechanisms. RNA interference (RNAi) has become a potent tool for comprehending gene activities in the context of crop improvement. It has shown to be useful in determining the functions of genes for metabolic pathways, stress tolerance, and more. Furthermore, RNAi-based strategies have potential for sustainable agriculture and integrated pest management, supporting international efforts to ensure food security. The many uses of RNA interference (RNAi) are covered in this article, including improving crop nutrition, changing plant architecture, and increasing shelf life. The challenges and future prospects of RNAi technology are also explored. The worldwide geography of RNA interference research is emphasized, with notable contributions from North America, Europe, and China. In conclusion, RNA interference (RNAi) continues to be a flexible and essential tool in contemporary plant research, providing new pathways for understanding gene activities and enhancing crop characteristics. Its combination with other biotechnological techniques, such as gene editing, might influence how agriculture and sustainable food production develop in the future.

A survey of listeners' perceptions of an extension-produced invasive plant podcast

Science communication and university Extension initiatives are evolving in response to society’s needs and an ever-changing digital landscape. The UF/IFAS Center for Aquatic and Invasive Plants (CAIP) has recently launched a podcast called Working in the Weeds (WITW). We surveyed podcast listeners to find out their motivations for listening and asked for their feedback on current episodes and future content. The survey revealed that a majority of respondents were individuals who work in the world of invasive plants. Many respondents were introduced to the podcast through work and listen to learn more about relevant research and information, to feel connected to the Center, and to be entertained. Overall, the respondents were satisfied with the podcast production elements and the variety of episode styles. Based on open-ended responses, listeners described that the podcast shares science in a meaningful way and the Center should continue producing episodes that highlight invasive plant research. Some respondents suggested future episodes should cover specific invasive plants, environmental impacts, herbicide use and safety, management techniques, and social issues surrounding invasive plant management.

Crop HTP Technologies: Applications and Prospects

In order to rapidly breed high-quality varieties, an increasing number of plant researchers have identified the functions of a large number of genes, but there is a serious lack of research on plants’ phenotypic traits. This severely hampers the breeding process and exacerbates the dual challenges of scarce resources and resource development and utilization. Currently, research on crop phenotyping has gradually transitioned from traditional methods to HTP technologies, highlighting the high regard scientists have for these technologies. It is well known that different crops’ phenotypic traits exhibit certain differences. Therefore, in rapidly acquiring phenotypic data and efficiently extracting key information from massive datasets is precisely where HTP technologies play a crucial role in agricultural development. The core content of this article, starting from the perspective of crop phenomics, summarizes the current research status of HTP technology, both domestically and internationally; the application of HTP technology in above-ground and underground parts of crops; and its integration with precision agriculture implementation and multi-omics research. Finally, the bottleneck and countermeasures of HTP technology in the current agricultural context are proposed in order to provide a new method for phenotype research. HTP technologies dynamically monitor plant growth conditions with multi-scale, comprehensive, and automated assessments. This enables a more effective exploration of the intrinsic “genotype-phenotype-environment” relationships, unveiling the mechanisms behind specific biological traits. In doing so, these technologies support the improvement and evolution of superior varieties.

Effects of nitrogen deficiency and drought stresses on miRNA expressions in Arabidopsis thaliana

MicroRNAs are now known to have an important role in regulating gene expression of eukaryotic organisms. miRNA research in plants gained importance after the discovery that several stress factors alter certain miRNA expressions, which subsequently regulate their target gene expressions and affect development and growth of plants. In this study, two of the widely studied abiotic stress conditions for plants, nitrogen deficiency and drought were used individually and as a combined stress treatment on Arabidopsis thaliana callus tissues to observe the change of expressions in certain miRNAs, when multiple stressors are encountered. Combined stress strongly inhibited callus growth compared to other conditions, while strongly altering certain miRNA expressions. Compared to control, in 7-day stress treated callus, miR165a-3p,b, miR319a,b, miR396b-5p, miR399d and miR827 showed significant downregulation for all stress treatments, while 7-day N deficiency caused miR167 upregulation. Stress treatments for 7 days mostly downregulated miR167c-5p, miR319c, miR399a, miR399e expressions except for the N deficient samples. After 14 days of stress, miR165a-3p,b, miR396a-5p, miR399b, miR399d were downregulated. During 14-day drought and combined stress, miR399a and miR396b-5p expressions were upregulated, respectively. The differences observed in this study between stress responses of 7 and 14-day-long treatments are believed to be valuable to further elucidate the associated molecular mechanisms of miRNAs, determination and validation of miRNA targets, and how plants respond to stress conditions via various genetic regulations.

Development and application of an RNA nanostructure to induce transient RNAi in difficult transgenic plants.

The development of RNA interference (RNAi) is crucial for studying plant gene function. Its use, is limited to a few plants with well-established transgenic techniques. Spray-induced gene silencing (SIGS) introduces exogenous double-stranded RNA (dsRNA) into plants by spraying, injection, or irrigation, triggering the RNAi pathway to instantly silence target genes. As is a transient RNAi technology that does not rely on transgenic methods, SIGS has significant potential for studying gene function in plants lacking advanced transgenic technology. In this study, to enhance their stability and delivery efficiency, siRNAs were used as structural motifs to construct RNA nanoparticles (NPs) of four shapes: triangle, square, pentagon, and hexagon. These NPs, when synthesized by Escherichia coli, showed that triangular and square shapes accumulated more efficiently than pentagon and hexagon shapes. Bioassays revealed that RNA squares had the highest RNAi efficiency, followed by RNA triangles, with GFP-dsRNA showing the lowest efficiency at 4 and 7 days post-spray. We further explored the use of RNA squares in inducing transient RNAi in plants that are difficult to transform genetically. The results indicated that Panax notoginseng-derived MYB2 (PnMYB2) and Camellia oleifera-derived GUT (CoGUT) were significantly suppressed in P. notoginseng and C. oleifera, respectively, following the application of PnMYB2- and CoGUT-specific RNA squares. These findings suggest that RNA squares are highly effective in SIGS and can be utilized for gene function research in plants.

Spatial transcriptomics techniques and its applications in plant research

The heterogeneity of gene expression in plant cells plays a crucial role in determining the functional differences among tissues. Recent advancements in spatial transcriptome (ST) technology have significantly contributed to the study of specific biological questions in plants. This technology has been successfully applied to examine cell development, identification, and stress resistance. This review aims to explore the application of ST technology in plants by reviewing three aspects: the development of ST technology, its current application in plants, and future research directions. The review provides a systematic description of the development process of ST technology, with a focus on analyzing its progress in studying plant cell growth and differentiation, plant cell identification, and stress resistance. In addition, the challenges faced by ST technology in plant applications are summarized, along with proposed future directions for plant research, including the advantages of combining other omics technologies with ST technology to tackle scientific challenges in the field of plants.

Biological culture module for plant research from seed-to-seed on the Chinese Space Station

The long-term cultivation of higher plants in space plays a substantial role in investigating the effects of microgravity on plant growth and development, acquiring valuable insights for developing a self-sustaining space life supporting system. The completion of the Chinese Space Station (CSS) provides us with a new permanent space experimental platform for long-term plant research in space. Biological Culture Module (GBCM), which was installed in the Wentian experimental Module of the CSS, was constructed with the objective of growing Arabidopsis thaliana and rice plants a full life cycle in space. The techniques of LED light control, gas regulation and water recovery have been developed for GBCM in which dry seeds of Arabidopsis and rice were set in root module of four culture chambers (CCs) and launched with Wentian module on July 24, 2022. These seeds were watered and germinated from July 28 and grew new seeds until November 26 within a duration of 120 days. To this end, both Arabidopsis and rice plants completed a full life cycle in microgravity on the CSS. As we know, this is the first space experiment achieving rice complete life cycle from seed-to-seed in space. This result demonstrates the possibility to cultivate the important food crop rice throughout its entire life cycle under the spaceflight environment and the technologies of GBCM have effectively supported the success of long-term plant culture experiments in space. These results can serve as invaluable references for constructing more expansive and intricate space plant cultivation systems in the future.

Sexy ways: approaches to studying plant sex chromosomes.

Sex chromosomes have evolved in many plant species with separate sexes. Current plant research is shifting from examining the structure of sex chromosomes to exploring their functional aspects. New studies are progressively unveiling the specific genetic and epigenetic mechanisms responsible for shaping distinct sexes in plants. While the fundamental methods of molecular biology and genomics are generally employed for the analysis of sex chromosomes, it is often necessary to modify classical procedures not only to simplify and expedite analyses but sometimes to make them possible at all. In this review, we demonstrate how, at the level of structural and functional genetics, cytogenetics, and bioinformatics, it is essential to adapt established procedures for sex chromosome analysis.

Improving transformation and regeneration efficiency in medicinal plants: Insights from other recalcitrant species.

Medicinal plants are integral to traditional medicine systems world-wide, being pivotal for human health. Harvesting plant material from natural environments, however, has led to species scarcity, prompting action to develop cultivation solutions that also aid conservation efforts. Biotechnological tools, specifically plant tissue culture and genetic transformation, offer solutions for sustainable, large-scale production and enhanced yield of valuable biomolecules. While these techniques are instrumental to the development of the medicinal plant industry, the challenge of inherent regeneration recalcitrance in some species to in vitro cultivation hampers these efforts. This review examines the strategies for overcoming recalcitrance in medicinal plants using a holistic approach, emphasising the meticulous choice of explants, e.g. embryonic/meristematic tissues; plant growth regulators, e.g. synthetic cytokinins; and use of novel regeneration-enabling methods to deliver morphogenic genes e.g. GRF/GIF chimeras and nanoparticles, which have been shown to contribute to overcoming recalcitrance barriers in agriculture crops. Furthermore, it highlights the benefit of cost-effective genomic technologies that enable precise genome editing and the value of integrating data-driven models to address genotype-specific challenges in medicinal plant research. These advances mark a progressive step towards a future where medicinal plant cultivation is not only more efficient and predictable but also inherently sustainable, ensuring the continued availability and exploitation of these important plants for current and future generations.

Why a strategic shift in action is needed to recognise and empower Indigenous plant pathology knowledge and research

Plant pathology researchers play a pivotal role in thought leadership and its translation to action regarding the recognition and demonstration of the value of Indigenous knowledge and science. For many scientists, navigating the space of Indigenous rights and perspectives is challenging. In pursuit of a cultural shift in research and development within the field of plant pathology, the 2019–2021 Management Committee of the Australasian Plant Pathology Society (APPS) undertook a review and modernization of the Society’s Constitution. The aim was to ensure its alignment with principles that foster inclusivity of Indigenous peoples in the development and implementation of relevant research projects impacting their communities. Additionally, a dynamic repository of guidelines and resources was compiled. These resources are designed to assist plant pathologists, while respecting and not superseding the guidance provided by local Indigenous researchers, practitioners, and advisors. The collective efforts of plant pathologists hold immense potential in championing Indigenous Peoples and their rights, steering the field toward a more inclusive and equitable future. This paper builds upon the thesis presented in the APPS Presidential Address at the Biennial APPS Conference in 2021, held virtually in lutruwita (Tasmania) on the unceded lands of the Palawa people. It underscores the potential impact when plant pathologists unite in advocating for Indigenous Peoples and their rightful place within the field.

High-quality chromosome-level genome assembly of Nicotiana benthamiana

Nicotiana benthamiana is a fundamental model organism in plant research. Recent advancements in genomic sequencing have revealed significant intraspecific genetic variations. This study addresses the pressing need for a precise genome sequence specific to its geographic origin by presenting a comprehensive genome assembly of the N. benthamiana LAB strain from the Republic of Korea (NbKLAB). We compare this assembly with the widely used NbLAB360 strain, shedding light on essential genomic differences between them. The outcome is a high-quality, chromosome-level genome assembly comprising 19 chromosomes, spanning 2,762 Mb, with an N50 of 142.6 Mb. Comparative analyses revealed notable variations, including 46,215 protein-coding genes, with an impressive 99.5% BUSCO completeness score. Furthermore, the NbKLAB assembly substantially improved the QV from 33% for NbLAB360 to 49%. This refined chromosomal genome assembly for N. benthamiana, in conjunction with comparative insights, provides a valuable resource for genomics research and molecular biology. This accomplishment forms a strong foundation for in-depth exploration into the intricacies of plant genetics and genomics, improved precision, and a comparative framework.

Rationalising Optimal Dosing of Phytotherapeutics For Use In Children: Current Status - Potential Solutions - Actions Needed.

"Children are not small adults with respect to the treatment with medicinal products." This statement of the WHO was the basis for the initiative of the European Commission for the establishment of a paediatric regulation in 2007 to improve the health of children by facilitating the development of medicines for children and adolescents. Seventeen years later, in the field of herbal medicinal products, results are still sobering. Therefore, the Foundation Plants for Health, Society for Medicinal Plants and Natural Products Research, and German Society for Phytotherapy organised a symposium to assess the status quo for the paediatric use of herbal medicinal products (HMPs), to analyse the causes of the current situation, and to discuss strategies for establishing the proof of safe and efficacious HMPs for children.The current situation for HMPs and their use in children is not fulfilling the requirements of legislation. HMPs in paediatrics are effective and safe, but considering the needs of children is necessary. In European countries, the use, registration, and marketing of HMPs are different, depending on the respective national regulations and specific traditions. EU herbal monographs are the best common denominator for such procedures. Emerging safety discussions must be considered. New approaches with real-world data might be a solution. The regulatory framework is to be adapted. Defining rationalised dosing for HMPs can be achieved by the extrapolation of data from adults, by using existing clinical data for children, and by using RWD. Therefore, a strong need for revising restrictions for the use of HMPs in children and rationalising defined dosage regimes is obvious.

Harnessing Nature's Pharmacy: Biotechnology Advances in Medicinal Plant Research

The integration of biotechnology into medicinal plant research has revolutionized the pharmaceutical industry, offering unprecedented opportunities for the development of novel therapeutics and healthcare solutions. By tapping into the vast biochemical diversity of plants, scientists have unlocked a treasure trove of potential drug candidates, many of which have been used in traditional medicine for centuries. Biotechnological tools such as genetic engineering, tissue culture, metabolomics, and bioinformatics have enabled the precise manipulation and optimization of plant-derived compounds, leading to the production of high-value pharmaceuticals with enhanced efficacy and safety profiles. Moreover, biotechnology facilitates sustainable practices by enabling the cultivation of medicinal plants under controlled conditions, reducing reliance on wild harvesting and alleviating pressure on endangered species. As our understanding of plant biology continues to deepen, the synergy between biotechnology and medicinal plant research holds tremendous promise for addressing global health challenges and improving patient outcomes.

Vom Chaos zur Klarheit. Die Neuordnung der Apfel TCP Gen-Benennung

Apple Proliferation (AP) is a widespread disease affecting apple orchards throughout Europe, including South Tyrol.The disease is associated with a phytoplasma, 'Candidatus Phytoplasma mali' ('Ca. P. mali'), which manipulates the plant's physiological processes through the secretion of small peptides called effectors. One well-studied phytoplasma effector is SAP11, which targets Teosinte Branched 1/Cycloidea/Proliferating Cell Factor 1 (TCP) genes in multiple plant species, including apple. TCP genes encode plant-specific transcription factors involved in various biological processes, such as growth, development, and responses to stimuli. The identification and naming of TCP genes have been inconsistent, leading to confusion and redundancy in sequence databases. In apple, 52 TCP genes (MdTCP) were identified in 2014 on the 2010 assembly of the apple genome, which is now considered outdated. To address this issue, a comprehensive investigation was conducted to identify and name TCP genes in apple using the high-quality genome assembly, GDDH13v1.1. Existing TCP sequences were aligned with the genome excluding redundant, fragmented, and non-TCP sequences. The revised set comprised 40 unique MdTCPs, including three novel genes. To establish a standardised nomenclature, each MdTCP was BLASTedagainst the Arabidopsis thaliana gene database, in order to identify the best hits for each MdTCP. The MdTCP genes were then renamed, incorporating the letter "a" or "b" to differentiate between MdTCPs showing homology to the same AtTCP, and also between the existing and proposed nomenclatures. The present study highlights the need for clarity and organisation in sequence databases, especially in respect of TCP genes. The presence of redundant and fragmented sequences in databases complicates accurate identification and annotation of genes. By providing a comprehensive and standardised nomenclature system, we aim to enhance the coherence and interoperability of future TCP gene research in plant and crop science. In our view, this work will serve as a valuable resource for researchers studying TCP genes in apple and will provide insights into the evolutionary dynamics and functional roles of TCP genes in plants.

Construction and initial performance evaluation of a newly developed plant PET system

In plant research, positron emission tomography (PET) is occasionally employed for physiological studies, offering valuable insights. However, the generally high cost of PET systems and their suboptimal design for plant research pose challenges to their application in this field. To address these issues, we have developed a new PET system optimized specifically for plant research. The PET detector ring was positioned vertically to enable measurements of plants in their normal upright position. The developed plant PET system features a transaxial field of view (FOV) of approximately 12 cm and an axial FOV slightly larger than 9 cm, allowing for the imaging of relatively small-sized plants. To facilitate imaging of taller plants, the PET system can expand the axial FOV by changing the subject height using a lab jack, enabling the imaging of taller plant species. The measured spatial resolution at the central FOV was 3.3 mm FWHM, and the sensitivity was 3.7%. The timing resolution was 6.78 ns FWHM with a lower energy threshold set to 350 keV. Phantom images simulating plants were successfully measured using the developed plant PET system. We conclude that the developed plant PET system holds promise for effective plant imaging.

First report of Ralstonia pseudosolanacearum causing bacterial wilt on Rosa sp. in Greece.

In March 2021, a sample of nine-month-old, non-grafted, diseased rose (Rosa sp.) plants was sent by a grower to the Benaki Phytopathological Institute for examination. The plants exhibited symptoms of dieback with black necrosis of pruned shoots, brown discoloration of shoot and root vascular tissues, and whitish slime exudation on cutting wounds of the shoots. The symptoms resembled those caused by Ralstonia pseudosolanacearum (Tjou-Tam-Sin et al. 2016). According to the sample's information sheet, the sample had been collected in a commercial greenhouse rose crop for cut flowers with a 10% disease incidence in the area of Troizinia-Methana (Regional Unit of Islands, Greece). Microscopic examination of symptomatic shoot and root vascular tissues revealed masses of bacterial cells streaming out of them. Sections of symptomatic tissues were suspended in water and in the resulting suspension, bacteria of the R. solanacearum species complex (RSSC) were detected by an indirect immunofluorescence (IF) assay using polyclonal antibodies (Plant Research International, the Netherlands) and a qPCR assay (RS-I-F/RS-II-R primers, RSP-55T probe) (Vreeburg et al. 2016). Furthermore, colonies with typical characteristics of RSSC were isolated from vascular tissues of shoots and roots on non-selective (NA) and semi-selective (mSMSA) media (EPPO 2022), and their identification as RSSC was confirmed by the above-mentioned IF and qPCR assays. Also, the isolates were assigned to: i) biovar 3, based on their ability to metabolize three disaccharides (maltose, lactose, D(+) cellobiose) and three hexose alcohols (mannitol, sorbitol, dulcitol) producing acid (EU 2006) and ii) phylotype I, by multiplex conventional PCR (Opina et al. 1997; Fegan and Prior 2005). A representative isolate was selected for sequencing part of the genes: 16S rDNA (1464bp), mutS (729bp) and egl (795bp) with GenBank Accession Nos. OR102443, OR683617 and OR702781, respectively. Blast analysis of these sequences showed 100% identity with those of various RSSC strains (e.g. GenBank Ac. Nos. CP025741.1, CP021762.1, MF141029.1, respectively). The obtained egl sequence conforms with the characteristics of phylotype I based on the DNA barcoding tool (EPPO 2021) and is 100% identical to that of the Dutch strain PD7216 (MF141029.1) reported to be sequevar I-33 (Bergsma-Vlami et al. 2018). The pathogenicity of two isolates was tested by inoculating: i) tomato seedlings (cv. 'Belladona') at their stem between the cotyledons and the first true leaf (EU 2006) and b) rose plants (cv. 'Aqua' and 'Papa Meilland') at their shoot base (Tjou-Tam-Sin et al. 2016), with bacterial suspensions in water (108 cfu/ml). The inoculated plants were maintained at a day/night temperature about 28/20°C with tomato plants exhibiting leaf wilting (7-17 dpi) and rose plants exhibiting chlorosis and necrosis of leaves (17 dpi). The pathogen was re-isolated on mSMSA from both artificially infected plant species and identified by the IF assay described above, thus fulfilling Koch's postulates. This is the first diagnosis in Greece of: i) rose plants infected by a Ralstonia species and ii) a crop infected by R. solanacearum phylotype I that corresponds to the R. pseudosolanacearum species (EPPO 2022). Official phytosanitary measures imposed in the affected area include an annual survey of rose crops for the presence of this pathogen, aiming at an early detection and prevention of its spread in such a highly valued ornamental crop.