PBN Symposium 2023

The Plant Biologicals Network invites everyone interested to the annual PBN Symposium: An unique opportunity to learn more about plant biologicals and network with PBN members to share knowledge and discuss experiences.

Program

8:30-9:00

Registration

9:00-9:25

Welcome and opening address in the context of Biosolutions initiatives by Svend Christensen, PBN Chairperson and professor, University of Copenhagen

9:25-10:20

Session 1: Regulation

Domenico Deserio, Policy Officer, European Commission – DG SANTE (online)
BioControlAgents(BCA) in EU: State of play since 2011, trends and perspectives

Patrice Marchand, PhD, DSc., Head of Inputs Dept., Institut de L’Agriculture et de L’Alimentation Biologiques (ITAB) (online)
BioControlAgents(BCA) in EU: State of play since 2011, trends and perspectives

10:20-10:50

Group photo and coffee break

10:50-12.00

Session 2: Robustness/lab to field transition

Danny Geelen, Professor, University of Gent
Stimulating plant growth with biostimulants: mode of action

Jens Kreim Larsen, Customer Technology Specialist, Corteva
How to document the effect of BlueN in agricultural crops

3 x flash talks

12:00-12:45

Lunch

12:45-14.05

Session 3: End-user/hands-on experiences

Bent Jensen, Production and Propagation Manager, Gunnar Christensens Nursery
Application of biological control in field production of pot culture

Benny Jensen, CEO, BJAgro
Perspectives from the field on using biologicals – pros and cons from +25y of consulting in use of biologicals

3 x flash talks

14:05-14:45

Poster session and coffee

14:45-15:55

Session 4: Hot topics in biologicals

Tess van de Voorde, Researcher, Wageningen University & Research,
Wageningen Plant Research, Biointeractions and Plant Health
Stimulating soil micro-organisms to support sustainable agriculture

Robert M. Kennedy, Chief Scientific Officer, Vestaron
Development and Commercialization of SPEAR® – A peptide based bioinsecticide with a major new mode of action

3 x flash talks

15.55-16.20

Coffee break

16.20-17.20

Panel discussion

Topic
What role will plant biologicals play in the green transition and in securing food production? What are the scientific, political and practical barriers for increasing the use of biologicals? A panel of relevant stakeholders will discuss these and other questions about the potential of plant biologicals.

Panellists
Sofie Carsten Nielsen, EU Biosolutions Project Director, Danish Industry
Anna Bak Jäpelt, Head of sustainability and Project Manager, The Think Tank Frej
Burghard Liebmann, Director Global R&D Plant Health, FMC Corporation

Moderator
Svend Christensen, PBN Chairperson and professor, University of Copenhagen

17:20-17:30

Closing of the symposium and final words

17:30-18:00

Transport to dinner venue

18:00-20:30

Symposium dinner

Location

The Copenhagen Plant Science Centre auditorium
Department of Plant and Environmental Sciences
University of Copenhagen
Bülowsvej 21A
1871 Frederiksberg

Domenico Deserio

European Commission initiatives on microbials and other biological control agents

The European Commission’s (EC) Farm to Fork Strategy aims at reducing dependency on and use of chemical pesticides, and placing on the market of biological alternatives, especially micro-organisms (MO) that can be used in organic farming, is facilitated. As of today, more than seventy MO are approved in EU, and there is an increasing trend towards biological solutions. The EU Regulations setting out data requirements and principles for assessment of MO were recently updated and are applicable since November 2022. They reflect the latest scientific developments and focus the risk assessment on the biology and ecology of MO. The new rules intend to facilitate the approval of MO, so that EU farmers have faster access to biological alternatives and can protect crops in a more sustainable manner. To support the implementation of these new rules, the EC has also adopted specific guidance documents (e.g., SANCO/2020/12258, SANTE/2020/12260), and Commission Communications (C/2023/3552, C/2023/3548). On other biological control agents (e.g. semiochemicals, botanicals, macroorganisms) guidance documents are already available (e.g. SANCO/11470/2012), and more initiatives are under development.

Patrice Marchand

BioControlAgents(BCA) in EU : State of play since 2011, trends and perspectives

Biocontrol agents (BCA) plant protection active substances (AS)composed from microorganisms, semiochemicals and substances from natural origin are increasing in Europe since the entry into force of Regulation (EC) 1107/2009, in number and as a percentage of total AS. As they are included in the scope of plant protection products (PPPs), this raises the question as to whether they are only substitute active substances, certainly more socially acceptable, sustainable and environmentally preferable, or really another way of managing bioaggressors, pests and diseases. As we have conducted a survey of all active substances listed in all Parts of Regulation EU 540/2011, and compared chemical to BCA active substances, we describe here their evolution and characteristics since 2011 (categories, functions, usages) and predict the global perspective in the future years in favour of BCA AS. This work allows us to conclude on the evolution of crop protection and the means that must be implemented for BCA, especially natural substances, to face current and new threats.

Danny Geelen

Stimulating plant growth with biostimulants: mode of action

Plant biostimulants are substances or microorganisms that are applied to plants or the soil to enhance crop production under normal or stress conditions. Unlike fertilizers, which provide essential nutrients directly to plants, biostimulants work through various mechanisms to improve plant performance indirectly. Biostimulants may be generated from quite diverse biomass sources (plant extract, seaweed extract, hydrolysates, chitin etc) and various microbial species have been described that exert biostimulatory activity. Hence, the mode of action of plant biostimulants is likely very diverse. Moreover, many commercial biostimulants are mixtures, which may act in a concerted manner through different biochemical and physiological processes. However, some common modes of action include: (i) hormone regulation: biostimulants can influence the production, transport, and balance of plant hormones such as auxins and cytokinins with profound impact on root and shoot development, flowering, and fertility; (ii) enhanced nutrient uptake: biostimulants can improve the plant’s ability to absorb and utilize nutrients from the soil especially in conditions where nutrient availability is limited; (iii) stress tolerance: biostimulants can help plants better cope with various environmental stresses, such as drought, salinity, and extreme temperatures; (iv) microbial activity: biostimulants contain beneficial microorganisms (e.g., mycorrhizal fungi, rhizobacteria) that establish symbiotic relationships with plants improving nutrient uptake and resilience to stress and disease; (v) soil health: biostimulants can improve overall health of the soil and soil structure; (vi) photosynthesis and metabolism: biostimulants can stimulate photosynthesis and metabolic processes in plants, leading to increased energy production and growth. The effectiveness of biostimulants may depend on various factors, including plant species, environmental conditions, and application methods. More detailed insight into the mode of action will help reducing the variable success of biostimulant treatments and to tailor the application to specific agricultural contexts. In this talk I will show some of the complexity of underlying plant growth processes and examples how to manipulate these to improve crop growth and yield.

Jens Kreim Larsen

How to document the effect of BlueN in agricultural crops

Jens’ talk will be centered around the following topics/highlights: 1) Fundamentals: what cannot be explained cannot be defended – there must be a need in the crop. 2) The challenge between climate zones and biologicals: In the Nordic countries it is often cold and sometimes also wet, while many products are primarily developed in southern Europe where the climate is hot and dry (products are rarely customized to the climate zone for which they are intended). 3) New challenges: spring droughts are seen more often in the Nordics than in the past. 4) BlueN is a live bacterium that must be kept alive and protected during delivery. 5) Identification of crops and treatment times, GPS technology, redistribution in the field? What parameters, soil type drought, historical yield maps? 6) BlueN supplies nitrogen to hungry crops without negative effects such as leaching and ammonia evaporation.

Bent Jensen

Application of biological control in field production of pot culture

Hands-on experiences from crop protection with use of biologicals

Benny Jensen

Perspectives from the field on using biologicals – seen from the perspective of a consultant

Benny Jensen, Agronomist and owner of the consulting firm BJ-Agro, will share learnings and challenges in understanding and exploiting the possibilities of using biologicals in practical agriculture, based on more than 25 years of experience working with bio stimulants.

Tess van de Voorde

Stimulating soil micro-organisms to support sustainable agriculture

Soils suppressive to soil-borne diseases have attracted the attention of farmers and researchers for decades and many suppressive soils have been described. Enhancing soil suppressiveness against soil-borne pathogens or pests is a promising alternative strategy to chemical pesticides. Microorganisms and soil microbial communities involved in suppressiveness have been studied intensively, but the underlying mechanisms are still not completely understood. However, organic amendments have been shown to reduce crop diseases and pests. We studied which characteristics of organic products are correlated with disease suppression in several experiments where different types of organic amendments with different physicochemical properties were tested. Chitin and keratin products prooved to the be most efficient against fungal pathogens. We will discuss how suppressiveness can be influenced by agricultural practices, and how the use of organic amendments to steer the microbial community can be applied in agriculture.

Robert M. Kennedy

Development and Commercialization of SPEAR® – A peptide based bioinsecticide with a major new mode of action

In 2020 SPEAR® won the Green Chemistry Challenge award in the “The Design of Greener Chemicals” category. This family of insect control products is based on Vestaron’s first peptide -based active ingredient. The SPEAR® peptide active ingredient has a novel mode of action at the Nicotinic Acetylcholine receptor as an allosteric modulator site II (IRAC group #32). Importantly, it has the same pest control efficacy as conventional chemicals but is safer for the environment and is readily biodegradable with a 0 day pre-harvest interval and 4 hour re-entry interval. SPEAR® continues to be recognized, winning the Best New Biological Product (Biopesticide) of the year at the Crop Science Forum & Awards in 2021. Based on the same peptide technology platform, Vestaron has submitted to the EPA a regulatory filing for its next product family, BASIN®. This product is expected to earn another novel IRAC group number, adding to growers’ green chemistry tools for crop protection.

Abstracts for flash talks and poster session

Novel genus of bacteriophages targets Danish soft rot isolates and represent promising biocontrol agents

Julie Stenberg Pedersen^1,3, Alexander Byth Carstens¹, Magnus Mulbjerg Rothgardt¹, Anouk Viry¹, Witold Kot¹, Charles Franz², Frank Hille², Robert Edwards³, Lars Hestbjerg Hansen¹

¹ Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg 1871, Denmark, ² Department of Microbiology and Biotechnology, Max Rubner-Institute, Hermann-Weigmann-Str. 1, 24103 Kiel, Germany, ³ Flinders Accelerator for Microbiome Exploration, College of Science and Engineering, Flinders University, Adelaide, Australia

Abstract
The bacterial diseases black leg and soft rot in potatoes cause heavy losses of potatoes worldwide. Bacteria within the genus Pectobacteriacea are the causative agents of black leg and soft rot in Potatoes. The use of antibiotics in agriculture is heavily regulated and no other effective treatment currently exists, but bacteriophages (phages) have shown promise as a potential biocontrol agent. In this study we aimed to isolate phages targeting Danish soft rot bacteria isolated from tubers and plants symptomatic with soft rot or black leg disease. Using organic waste, we isolated 19 phages targeting different species within Pectobacterium. Here we present seven of these phages representing a new genus primarily targeting P. brasiliense; Ymer genus. TEM image of phage Ymer showed siphovirus morphology, and phages belong to the class Caudoviricetes, with double-stranded DNA genomes varying from 40kb to 42kb. A host-range experiment with 59 bacterial isolates from Danish tubers showed phages to primarily target P. brasiliense. Interestingly the seven phages displayed difference in host-range, with two of the phages being able to infect two or more species of Pectobacterium. In silico analysis of the P. brasiliense genomes showed difference in genes involved in the outer membrane, even in closely related isolates, which correlated with host range results. None of the phages encode any integrase or other genes typically associated with lysogeny. Based on the genome analysis together with the host range results these phages could have potential as biocontrol agents against soft rot and black leg in potatoes and should be tested further.

One Crop Health: A new approach for next-generation crop protection

Paul Neve¹, Jonathan Storkey², Sune Darkner³, Per Kudsk⁴

¹ Plant & Environmental Sciences, University of Copenhagen, Denmark. ² Protection Crops & the Environment, Rothamsted Research, United Kingdom, ³ Department of Computer Science, University of Copenhagen, Denmark, ⁴ Department of Agroecology, Aarhus University, Denmark.

Abstract
Current approaches for limiting crop yield loss to insect pests, plant pathogens and weeds are heavily reliant on pesticides, though environmental, regulatory, and agronomic pressures mean that pesticide-dominated crop protection is no longer sustainable. This newly funded project, which starts in January 2024, proposes a One Crop Health approach for crop protection that integrates the latest advances in agricultural technology, ecology, data science and modelling. Motivated by the One Health concept in human health, our vision asserts that plant health, soil health and agroecosystem health are intimately linked, and that only by recognising and optimising those links can we reduce the need for pest control interventions and associated negative environmental outcomes. To realise this potential, our systems-based research will emphasise agroecological approaches for pest, pathogen and weed prevention, digital agriculture-based solutions for enhanced detection and prediction, and robotics and biology-based solutions for more effective and environmentally benign control of biotic threats. The multi-scale approach that we propose spans farm networks, citizen science, experimental farm platforms, digital twins, and research to develop innovative solutions for pest, pathogen and/or weed prevention, detection, or control. We will develop a systems-level perspective that vertically integrates prevention, detection and control of individual biotic threats and horizontally integrates the management of pests, weeds, diseases, and functional biodiversity. Successful outcomes will include reduced use of pesticides, reduced pest pressure, increased cropping system resilience to biotic threats and enhanced biodiversity and soil health.

Integrating wheat genetics, natural product chemistry and microbial ecology approaches to improve sustainable crop production through biological nitrification inhibition (BNI)

Jasmeet Kaur Bhambra¹, Jonathan Sølve¹, Kenneth Madriz Ordenana³, Hans Thordal-Christensen³, Purna Kumar Khatri², Inge S. Fomsgaard², Cecile Gubry-Rangin³, Xiaoping Fan³, Kristian Koefoed Brandt³

¹Section for Microbial Ecology and Biotechnology, Department of Plant and Environmental Sciences, University of Copenhagen, Denmark, ²Section for Plant and Soil Sciences Department of Plant and Environmental Sciences, University of Copenhagen, Denmark, ³Department of Agroecology – Crop health, Aarhus University, Denmark,  ⁴School of Biological Sciences, Aberdeen University, Aberdeen, United Kingdom (UK)

Abstract
Climate change is a key challenge facing mankind. Anthropogenic activities contribute to nitrous oxide (N₂O) emissions, a long-lived and highly potent greenhouse gas accumulating in the atmosphere at an increasing rate. Agricultural fertilised soils are the primary source of N₂O emissions, where about 50% of fertiliser input in farmland is lost due to the microbial process of nitrification performed by nitrifiers that utilise fertiliser N for their growth. Reducing nitrification in agricultural soils efficiently reduces greenhouse gas emissions, improves crop yield by increasing fertiliser N use efficiency and reduces eutrophication from excessive N input. The synthetic nitrification inhibitors currently in use are inefficient and very costly. The release of nitrification inhibitors from plants through a process called biological nitrification inhibition (BNI), represents an attractive and sustainable approach to reducing nitrification and increasing the farmer economy. This project aims to further BNI knowledge in wheat agriculture by integrating wheat genetics with plant metabolite natural chemistry and its impact on nitrifier microbial diversity and soil activity. We will establish a high-throughput platform for identifying BNI capacity in old landraces and currently used wheat cultivars with the aim of breeding wheat lines with improved BNI efficiency. Here, we will present our preliminary data from old Iranian wheat landraces demonstrating that different wheat varieties have variable BNI efficiency based on nitrifier response and its links to the root exudate chemistry.

Herd immunity in food crops: efficient and durable use of biocontrol and chemical fungicides

Esther Kuper*, Mudassir Iqbal, Åsa Lankinen, Erik Andreasson, Johan A. Stenberg¹

¹ Department of Plant Protection Biology, Swedish University of Agricultural Sciences, Alnarp, Sweden

Abstract
Herd immunity is an epidemiological concept that describes the indirect protection of susceptible individuals when a sufficiently large proportion of the population is immune against a disease. Although widely discussed in human epidemiology (e.g. during the COVID-19 pandemic) this concept has never fully been explored in relation to crops. In this project we investigate whether herd immunity can be obtained in garden strawberries by 1) targeted application of the biological control fungus Aureobasidium pullulans on selected individual plants or 2) mixed cropping of susceptible and resistant cultivars. The grey mould-causing pathogen Botrytis cinerea is used as model organism. In a long-term field trial we evaluate the effect of A. pullulans compared to a common chemical fungicide (Switch^®) and explore disease prevalence and severity in populations with or without herd immunity treatment. The same experimental field matrix is used to monitor and compare the efficiency of biocontrol and chemical fungicides over time. As reports of B. cinerea strains resistant to chemical fungicides are increasing, we will test for resistances against biological control in B. cinerea field isolates that have repeatedly been exposed to A. pullulans.  We further hypothesize that the targeted application of A. pullulans in the herd immunity treatment creates evolutionary micro-refuges on untreated plants, preventing the pathogen from evolving counter-resistance. If we can observe herd immunity in strawberry populations it would result in a reduced use of plant protection products while nonetheless achieving sufficient disease control. If herd immunity additionally prevents evolution of counter-resistance in the pathogen, it would enable prolonged durability of plant protection products and resistant cultivars.

APEX™: A revolutionary pipeline of microbial solutions, fueled by nature to preserve nature

Charlotte Klank¹, Steven Vandenabeele¹

¹ Aphea.Bio NV, Gent, Belgium

Abstract
Aphea.Bio has developed the R&D platform APEX™, which combines unique know-how and disruptive technologies targeted towards selecting the ‘winner’ microorganisms out of millions present in and around the plant and developing them into an effective product ready to be used by farmers. APEX™ enables us to identify microorganisms that are effective against a certain target pest, disease or weed, or boost the plants in a highly time- & cost-efficient manner. During our discovery and development process, we identify candidate microorganisms that are highly efficient, while at the same time having a low risk profile, thus posing no harm to non-target organisms and plants as well as having no risk for human health. The versatile platform can be adapted towards different crops and traits, enabling the development of new biological products whose performance and consistency can complement as well as compete with or replace conventional products. Covering row crops such as wheat and maize as well as fruit & vegetables, our biological product portfolio will play a significant part in reducing the need of synthetic fertilizers and chemical pesticides by offering solutions that are not harmful to the environment and human health, thus enabling an agricultural system where ecosystem services can be protected, biodiversity in agricultural landscapes can be increased and food security can be ensured.

Accelerated biofungicide discovery using automated screening of fungal biodiversity

Parvathy Krishnan¹, Johan V. Christiansen¹, Vilhelm K. Møller¹, Søren D. Petersen¹, David L. Corcoles¹, Alexander R. Brems, Steen S. Brewer, Rune J. Brunshøj ¹, Belén C. Ramal², Emmanouil Chantzis², Joana Cachapa², Rikke A. Behrendt², Karolina Subko², Christopher B. C. Phippen², Burghard Liebmann², Lars Jelsbak¹, Rasmus Frandsen¹, Jakob B. Hoof ¹, Jens C. Frisvad, Niels B. Jensen^1*

¹Department of Bioengineering, Technical University of Denmark, Kongens Lyngby, Denmark
²FMC Agricultural Solutions A/S, European Innovation Center, Hørsholm, Denmark

Abstract
Crop losses due to fungal pathogens pose a significant challenge to the global food supply, with an estimated 20-40% of the world crop production being lost due to pests and diseases. While chemical pesticides offer a solution, they often introduce new environmental and health-related concerns. In pursuit of a sustainable biosolution, DTU has teamed up with FMC to rigorously screen fungi for their potential as future biological solutions against two defined wheat pathogens, Fusarium and Zymoseptoria. Using the extensive DTU/IBT strain collection, we have converted approximately 7,000 single-vial strains into a screenable 96-well plat format, with an end-project goal of 20,000 strains in this screenable format. These fungi are cultured on empirically-selected growth media optimized for secondary metabolite production. Our state-of-the-art robotics workflow allows for simultaneous co-culture assays, extract assays, and untargeted LC-MS data collection for about 800 strains every two to three weeks. Automated computational processing of the LC-MS data and a high-throughput genotyping workflow aimed at taxonomic identification, we can focus on commercial relevant lead candidates early in the process. By leveraging machine learning and image analysis, we aim to refine our selection criteria and reduce analysis time even further, pushing the frontier of fungal-derived biosolutions. In collaboration with our commercial partner, FMC, several strains are already under field trials in Europe.

Fungal compounds targeting the plant H+-ATPase

Anja Thoe Fuglsang¹

¹Section for Transport Biology, Department of Plant and Environmental Sciences, University of Copenhagen, Denmark

Abstract
The plant plasma membrane (PM) H⁺-ATPase is an essential enzyme controlling plant growth and development. It is an important factor in response to abiotic and biotic stresses and is subject to tight regulation. We are in demand for new sustainable natural growth regulators and as a key enzyme for regulation of transport into the plant cell the PM H⁺-ATPase is a potential target for these. We have screened collections of fungi for compounds with bioactivity towards the plant H⁺-ATPase and found both compounds with activating effect as well as an inhibitory effect. We have investigated a few compounds in details focusing on their mechanism of action and their potential as biologicals/growth regulators in plant production of future sustainable agriculture.

Viscosin synthesis promotes Pseudomonas fluorescens SBW25 root colonization and alters microbial assembly in wheat rhizoplane in a cultivar dependent manner

Ying Guan, Frederik Bak, Rosanna C. Hennessy, Christine Lorenzen Elberg, Dorte Bodin Dresbøll, Anne Winding, Rumakanta Sapkota, Mette H. Nicolaisen

Abstract
The importance of microbial functions for plant health and performance is unquestioned. Beneficial rhizosphere bacteria such as Pseudomonas sp. promote plant growth and provide protection against pathogens. In the rhizosphere, plant-associated pseudomonads are producers of a large number of bioactive compounds, including cyclic lipopeptides, which are particularly well-known for their antimicrobial activities. However, the role of cyclic lipopeptides in the interaction between beneficial bacteria and plants remains underexplored, especially in natural systems. In this study, the model strain Pseudomonas fluorescens SBW25 producing the cyclic lipopeptide viscosin was used to unravel the impact of viscosin on bacterial colonization potential and microbiome assembly at the wheat root. Two varieties of winter wheat were inoculated with either the SBW25 wild-type strain or a viscA mutant deficient in viscosin production. The ability of both strains to colonize the roots of each variety was quantified using qPCR analysis, and plant parameters including plant biomass and height were assessed. Our results indicate that SBW25 wild-type strain has stronger colonization ability than the mutant strain at the rhizoplane. In order to study the impact of viscosin on microbial assembly, rhizobiome16S rRNA gene and 18S rRNA gene amplicon sequencing were performed. Our results indicate that viscosin intervention enhanced bacterial community diversity and decreased protist community diversity in Sheriff and significantly altered the microbial community structure, whereas the viscosin producing strain did not alter the microbial community structure of Heerup. This study provide new insights into the natural importance of viscosin and specifically the role of viscosin in the colonization of plant roots and in shaping the microbial communities associated with different wheat varieties.

Streptomyces alleviate abiotic stress in plant by producing pteridic acids

Zhijie Yang¹, Emil Strøbech1, Yijun Qiao¹, Naga Charan Konakall², Pernille Harris³, Gundela Peschel⁴, Miriam Agler-Rosenbaum⁴, Tilmann Weber⁵, Erik Andreasson², Ling Ding¹,*

¹Department of Biotechnology and Biomedicine, Technical University of Denmark, Kgs. Lyngby, Denmark, ²Department of Plant Protection Biology, Swedish University of Agricultural Sciences, Alnarp, Sweden, ³Department of Chemistry, Technical University of Denmark, Kgs. Lyngby, Denmark, ⁴Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute (HKI), Jena, Germany, ⁵Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kgs. Lyngby, Denmark

Abstract
Soil microbiota can confer fitness advantages to plants and increase crop resilience to drought and other abiotic stressors. However, there is little evidence on the mechanisms correlating a microbial trait with plant abiotic stress tolerance. Here, we report that Streptomyces effectively alleviates the drought and salinity stress by producing spiroketal polyketide pteridic acid H (1) and its isomer F (2), both of which promote root growth in Arabidopsis at a concentration of 1.3 nM under abiotic stress. Pteridic acids induce stress response genes expression in salinitystressed Arabidopsis seedlings. The bifunctional biosynthetic gene cluster of pteridic acids and antimicrobial elaiophylin is confirmed in vivo and mainly disseminated by vertical transmission which is geographically distributed in various environments. This discovery reveals a perspective for understanding plant-Streptomyces interactions and provides a promising approach for utilising beneficial Streptomyces and their secondary metabolites in agriculture to mitigate the detrimental effects of climate change.

Nanoparticles and Plant Proteins: A Path to Sustainable Agriculture

Hoda Eskandari^1,2, Hossein Mohammad beigi², Duncan Sutherland^1*

¹ Interdisciplinary Nanoscience Center, Aarhus University, Denmark, ² Department of Biotechnology and Biomedicine, Technical University of Denmark, Denmark

Abstract
Nano-enabled agriculture offers a promising approach to tackle food security and sustainability challenges. This study investigates the interaction between plant macromolecules, especially proteins and nanoparticles to understand how they enhance nanoparticle biotransformation, uptake, and promote plant growth under abiotic stress. Silica nanoparticles (SNPs), known for their ability to enhance plant growth under heat stress, were selected for this study. Wheat and Lotus were chosen as model plants. The study involved exposing wheat and Lotus proteins to 50 nm SNPs at both 22°C and 43°C for 2 and 24 hours. We analyzed the formation of the protein corona using SDS-Page, dynamic light scattering (DLS), Nano-DSF, and transmission electron microscopy (TEM). SDS-Page results revealed distinct protein bands influenced by incubation time and temperature. DLS data showed a significant increase in SNP size and heterogeneity, particularly after the 2-hour and 24-hour exposures in the presence of proteins. Nano-DSF results highlighted changes in the stability of Lotus and wheat proteins in the presence of SNPs under heat stress. Additionally, TEM images visually confirmed the presence of protein coatings on the surface of SNPs, providing insights into the intricate interactions between nanoparticles and plant macromolecules. These findings have important implications for advancing agricultural practices and promoting sustainability.

Plant growth promoting rhizobacteria-mediated root architectural changes are correlating with systemic impact on antioxidant and carbohydrate metabolism

Chandana Pandey, Wenjing Tian, Victoria Naoumi, Daniel Buchvaldt Amby, Seyed Fazel Mirahmadi, Thomas Roitsch

University of Copenhagen, Department of Plant and Environmental Sciences, Denmark

Abstract
The structural features and traits of a plant’s root system play a vital role in securely anchoring the plant in the soil, while also facilitating the absorption of essential water and nutrients. These factors significantly influence both the health and overall productivity of the crop. A sustainable and environmentally friendly solutions such as plant growth promoting rhizobacteria (PGPR) contributes to an improved and stabilized crop yield. This case study aims to assess the barley (Hordeum vulgare cv Guld) responses to root growth and architecture under colonization of PGPR, Enterobacter and Pseudomonas spp. Preliminary experimental results have shown that the impact of PGPR is similar impact on root growth as cytokinins. Deep physiological phenotyping via enzyme activity profiling revealed an impact of PGPR on the activity signatures of key enzymes of carbohydrate and antioxidant metabolism. This specific effect on roots was not as prominent under controlled soil conditions. RNASeq was performed to study the early and late-stage interactions of Enterobacter on barley roots. Gene expression profiling identified differentially expressed genes and highlighted the enrichment oxidative stress and catabolic, metabolic, and biosynthetic processes. Based on these findings, it is evident that PGPR have the ability to influence plant metabolism and stress responses, which are critical for root development. In summary, this study offers significant evidence on the influence of Enterobacter and Pseudomonas spp. on barley plants, improves our comprehension of their mechanisms and providing potential value to researchers studying PGPR-plant interactions.

Production of viscosin in Pseudomonas fluorescens SBW25 is regulated by orphan LuxR regulators that bind to plant phenolics and quorum-sensing molecules

Kitzia Y. Molina-Zamudio¹, Nina Molin Høyland-Kroghsbo¹, Kasper Røjkjær Andersen², Rosanna C. Hennessy, Mette H. Nicolaisen¹*

¹Section for Microbial Ecology and Biotechnology, Department of Plant and Environmental Sciences, University of Copenhagen, Copenhagen, Denmark, ²Section of Plant Molecular Biology, Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark

Abstract
Viscosin is a cyclic lipopeptide (CLP) produced by Pseudomonas fluorescens SBW25 that facilitates bacterial plant-root colonization due to its biosurfactant properties. The biosynthesis of viscosin is regulated by two LuxR-type regulators, ViscAR and ViscBCR, but the exact regulatory mechanisms remain unclear. The production of CLPs in several Pseudomonads is regulated through quorum-sensing, where a LuxR transcriptional regulator binds to an acyl homoserine lactone (AHL) signaling molecule, forming a complex that induces the transcription of the target genes. Although ViscAR and ViscBCR belong to the LuxR family, they lack the conserved amino acids typically required for binding to AHLs. However, recent research suggests that in addition to binding to their cognate AHLs, several LuxR receptors can also interact with other compounds, including plant phenolics like arbutin, salicin, phenyl as well as flavonoids. We hypothesize that plant phenolics and/or AHLs in complex with ViscAR/BCR regulators, or other uncovered orphan LuxR regulators in the SBW25 genome may trigger up-regulation of viscosin biosynthetic genes. To investigate this, we tested a range of plant phenolics and quorum-sensing molecules as potential triggers for viscosin biosynthesis using a bioreporter assays. We determined that ViscAR up-regulates the expression of viscA in the presence of salicin, viscB in the presence of C6-HSL, and self-regulates its expression in the presence of 3OC6-HSL. Additionally, ViscBCR regulates viscA and viscAR expression via an unknown mechanism. Moreover, we discovered an orphan LasR-homolog that directly regulates viscA in the presence of 2-benzoxazolinone (BOA). An in silico structural analysis of ViscAR showed a non-archetypical ligand-binding domain potentially able to bind to AHLs and plant phenolics. The observed induction translated into significantly increased swarming motility of SBW25 in the presence of salicin, C6-HSL and 3OC6-HSL. This study shows that SBW25 can sense its environment and respond to both interspecies and interkingdom signals of importance for its dispersal.

Biological control of Zymoseptoria tritici in wheat

Hans J. L. Jørgensen, Birgit Jensen, Meike A. C. Latz, David B. Collinge

Department of Plant and Environmental Sciences, University of Copenhagen

Abstract
The disease Septoria tritici blotch (STB), caused by Zymoseptoria tritici, is one of the most important and yield-reducing constraints in wheat production worldwide. Control relies heavily on fungicides, but there is an increasing desire to reduce fungicide use and the pathogen readily develops resistance against commonly used products. Biological control, using living microorganisms, is an upcoming trend within disease control, also for STB. The mode of action of biocontrol agents is more complex than for traditional fungicides and therefore, the risk that they lose effect is considered much less than for chemical products. Potential biocontrol agents are often selected based on direct inhibitory effects in vitro and therefore, the mode of action is often not studied in detail. However, there is emerging evidence that one of the most important modes of action by biocontrol agents is induced resistance, which can only be discovered in assays using plants. We use different fungi (e.g. endophytes and Clonostachys rosea) and bacteria to control STB and have found significant reductions of disease severity using spray applications of fungal biocontrol agents under controlled and field conditions. Whereas in vitro studies showed limited inhibition of the pathogen, microscopy and transcriptomics implicated induced resistance as important mechanisms.