Biotech for Sustainable Agriculture
Angharad M. R. Gatehouse
Biotechnology for Sustainable Agriculture: The Future?
Angharad M. R. Gatehouse*, and Martin G. Edwards
School of Natural and Environmental Sciences, Newcastle University, Newcastle upon Tyne, UK
Food Security is a major political and societal challenge. However, achieving this goal is threatened both by climate change and an ever-increasing global population, already exceeding 8.23 billion. Thus, the need for sustainable crop production has never been greater, requiring the development of efficacious and cost-effective crop protection strategies that pose negligible risks to the environment at large. This goal requires a paradigm shift in how we produce and cultivate our crops to be resilient to both abiotic and biotic stressors to meet the challenges of feeding a predicted population of 9.8 billion people by 2050 (SDG 2). Technologies such as RNA interference (RNAi) provide an attractive alternative approach to crop protection, providing control by silencing essential genes of pest insects in a highly sequence-specific manner. However, this technology is not without its problems. These include stability of the dsRNA, both in planta and within the target insect itself, and effective delivery mechanisms when used as a biopesticide. Similarly, gene editing techniques, particularly CRISPR/Cas, provide transformative agricultural tools for enhancing crop protection against stress. This presentation will provide an overview of the potential of these emerging technologies for enhancing crop protection against both biotic (diseases, pests, pathogens) and abiotic (drought, heat, cold, salinity) stresses, with examples of their current commercial status and those in development.
Baltasar Escriche
- Session: Biotechnology in Alternative Proteins - Focus on Bt toxins
- Presentation: Speaker
New challenges for Bacillus thuringiensis in crop protection
Yolanda Bel & Baltasar Escriche
Biotechnological pest control laboratory, Institute of Biotechnology and Biomedicine (BIOTECMED), Universitat de València, 46100-Burjassot, Valencia, Spain
Author for correspondence: baltasar.escriche@uv.es
Crop protection against a wide range of pests has been a constant challenge since the introduction of agriculture. The various strategies developed over time to control them have enabled the production of high-yield, commercially valuable crops. However, the overuse of synthetic chemical insecticides has raised environmental concerns and promoted the development of sustainable, low-impact pest control alternatives. Among these, the microbial insecticide particularly based on the bacterium Bacillus thuringiensis (Bt) have achieved remarkable success.
During sporulation, Bt produces highly compact parasporal crystals composed of proteins (Cry prpteins) with strong insecticidal activity. Each Cry protein displays a high degree of specificity against certain target organisms.The activity of some Cry proteins against major lepidopteran pests is well known, and several have been expressed in transgenic crops (Bt crops) that are inherently protected. Large areas of maize and cotton are currently cultivated using this approach in certain countries.
In contrast, plant-parasitic nematodes have received comparatively little attention, although globally distributed and economically damaging. These pests weaken plants and reduce yields, often without causing visible above-ground symptoms. Several Cry proteins from Bt have been identified with nematicidal activity, including members of the Cry5, App6, Cry12, Cry13, Cry14, Cry21, and Xpp55 groups.
Our group screened 846 Bt strains to determine the diversity and distribution of nematicidal cry genes. PCR analyses followed by sequencing confirmed the presence of at least one nematicidal cry gene in 164 isolates (20%). Cry5 and App6 were the most abundant, and Cry5 frequently occurred together with Cry21. The frequency of specific genes varied markedly between collections, reflecting differences in sampling time, geographic origin, and source material.
We also evaluated the toxicity of purified Bt crystal proteins Cry5, Cry21, App6, and Xpp55 against two root-knot nematodes, Meloidogyne incognita and M. javanica. Once solubilized, all four proteins were highly toxic to both species in laboratory assays. To assess the potential field use of Bt strains producing these nematicidal proteins, we conducted in planta experiments with M. javanica using wild-type Bt strains that produced Cry5 alone or in combination with App6. Cucumber and tomato plants were infested with nematodes and irrigated with spore–crystal mixtures of the respective strains. Results revealed that nematicidal activity was plant-dependent: Bt significantly reduced the emergence of infective juveniles (J2) in tomato but not in cucumber. Moreover, the in planta toxicity was much lower than expected from in vitro results, underscoring the difficulty of achieving effective nematode control with proteins delivered as crystalline inclusions.
These findings highlight that the delivery method of Bt nematicidal proteins is critical for their success against root-knot nematodes. In this context, transgenic plants expressing these proteins may represent the most effective and reliable strategy for sustainable nematode management.
Federico Ariel
RNA Sprays: Harnessing Noncoding RNAs for Sustainable Crop Protection and Resilience
Federico Ariel
APOLO Biotech, Argentina. Instituto de Fisiología, Biología Molecular y Neurociencias (CONICET and Universidad de Buenos Aires, Argentina).
The success of RNA vaccines has brought RNA molecules to the forefront of biotechnology, and agriculture is now poised to benefit from this molecular revolution. RNA-based technologies, particularly RNA interference (RNAi), offer a sustainable alternative to conventional pesticides by providing species-specific, environmentally safe, and human health–friendly solutions. At APOLO Biotech, we have developed an integrated platform to advance RNA applications in agriculture. This includes: (i) bioinformatics pipelines for the rational design of RNA solutions; (ii) the first pilot plant for RNA production in Latin America, established in Argentina, enabling scalable and cost-effective manufacturing; and (iii) machine learning–driven computer vision systems to systematically evaluate plant responses and disease control efficacy. Through this pipeline, we currently target over 10 fungal diseases across more than 15 crops, demonstrating both effectiveness in field trials and scalability. Beyond crop protection, RNA technologies are opening new opportunities in plant resilience. Once considered genomic “dark matter,” noncoding RNAs (ncRNAs) are now recognized as key regulators of gene expression in plants. By leveraging long and small ncRNAs, we are designing RNA sprays to promote crop adaptation to environmental stresses, thereby mitigating yield losses while preserving genetic diversity. RNA-based agtech innovations can boost crop productivity, reduce reliance on synthetic chemicals, and contribute to global food security and sustainable agriculture.
Gabriela Alejandra Massa
Gene-Edited Potato Varieties: From Laboratory Innovation to Agricultural and Industrial Application
Potato (Solanum tuberosum) is the third most consumed food crop worldwide after rice and wheat, and represents a strategic crop for global food security. In Argentina, modern potato production is concentrated mainly in the provinces of Buenos Aires and Córdoba, where the leading cultivars are Spunta for fresh consumption and Atlantic for the potato chip industry. Despite its importance, potato breeding faces major challenges related to post-harvest quality, processing characteristics, environmental resilience, and disease resistance. Among the most critical industrial traits to improve are enzymatic browning and cold-induced sweetening, two physiological processes that strongly reduce both the quality of fresh tubers and processed products. On the agronomic side, breeding goals also include the development of cultivars with improved water-use efficiency and increased tolerance to major pathogens such as Phytophthora infestans, the causal agent of late blight. Traditional breeding of potato is particularly difficult due to its polyploid, highly heterozygous, and vegetatively propagated nature, which makes the process slow and labor-intensive, often requiring 10-15 years to release a new cultivar. In this context, advanced biotechnological tools such as CRISPR/Cas9-mediated genome editing provide a powerful alternative to accelerate the development of improved potato varieties with specific, well-defined traits. Our research group has been pioneering the application of genome editing to potato improvement, targeting genes associated with both quality and resilience. Our first achievement was the development of bruise-resistant potatoes derived from cv. Desiree. This was obtained through knockout (KO) of the StPPO2 gene, which encodes a polyphenol oxidase enzyme responsible for enzymatic browning. We also generated cv. Atlantic lines resistant to cold-induced sweetening via KO of the vacuolar invertase gene (StInvVac), a key enzyme regulating the accumulation of reducing sugars during cold storage. In addition, KO of the StCBP80 gene in cv. Spunta, a regulator within the abscisic acid (ABA) signaling pathway, resulted in plants with enhanced water-use efficiency. More recently, we are advancing the development of cv. Spunta potatoes edited simultaneously in StPPO2 and StInvVac, aiming to combine resistance to both enzymatic browning and cold-induced sweetening within the same genetic background. Importantly, we are also focusing on increasing resistance to late blight by editing host susceptibility genes. Ongoing work is targeting StDMR6 (Downy Mildew Resistance 6) and StSR1 (Signal Responsive 1), two genes implicated in plant-pathogen interactions and previously validated as promising candidates for broad-spectrum disease resistance. Edited Spunta lines carrying modifications in these genes are currently under evaluation for their response to P. infestans infection. CONABIA determined that lines resistant to enzymatic browning, cold-induced sweetening, and improved water-use efficiency are not considered GMOs. In addition, the enzymatic browning resistant line and the cold-induced sweetening resistant line are currently under evaluation by INASE for registration as new varieties. Overall, our results demonstrate that genome editing can deliver potato products with superior industrial quality, reduced post-harvest losses, improved resilience, and enhanced disease resistance. These developments will benefit farmers, the food industry, and consumers, while positioning Argentina at the forefront of innovative, sustainable, and socially accepted crop biotechnology.
Hector Ruiz
Title: Bioprocesses and biorefineries as a strategy for the conversion of lignocellulosic and algal biomass using high-pressure hydrothermal technology
Abstract: The development of society leads us to seek alternatives in the production and use of fuels and compounds of industrial interest to meet our needs. One concept that has been developed in recent years for this purpose is biorefineries, which are analogous to petroleum refineries, using different types of biomass as feedstock. This conference aims to showcase, analyze, and discuss technological advances in the development of second-generation (lignocellulosic biomass) and third-generation (algae) biorefineries for the production of biofuels and high-value-added compounds. This talk will focus on the operation of the biomass fractionation process, bioprocess conversion, specifically high-pressure hydrothermal processes, and on the design and operation of pilot-scale bioethanol production as a biofuel. In addition, microalgae cultivation systems, focusing on the design of photobioreactors and alternative cultivation media for the production of microalgal biomass with applications in bioenergy and food will be addressed.
Janice Barbosa
Unraveling Gall Ontogeny and Regeneration: Pathways to Combat Parasitic Nematodes
Janice de Almeida Engler
Plant-parasitic nematodes pose a significant threat to global agriculture, aggravated by global warming, making the development of effective and sustainable control strategies a pressing priority. A fundamental understanding of the cellular and molecular mechanisms underlying nematode infection is essential for developing crop varieties with enhanced resistance.
Our research explores gall ontogeny and regeneration in response to root-knot nematode (RKN) infection, with a particular focus on cytoskeletal dynamics, nuclear organization, and cell cycle regulation in nematode-induced giant cells. Disruption of microtubules during infection appears to be a deliberate strategy by the nematode to facilitate nutrient acquisition, identifying the host cytoskeleton as a potential target for nematode control. Notably, studies on nuclear cluster organization revealed the presence of nucleotubes structures connecting lobed nuclei within clusters which resemble features observed in cultured tumor-derived brain cancer cells. This suggests a conserved cellular mechanism supporting multinucleate cell function and survival.
Gall formation involves a sophisticated reprogramming of the host cell cycle machinery. RKNs induce tumor-like structures within the root vascular cylinder, where cells retain proliferative competence. Functional analyses revealed the differential activation of key cell cycle regulators that control mitotic and endocycle processes to maintain gall homeostasis.
Furthermore, we investigated whether nematode-induced cellular damage also triggers a regeneration-like response within galls. Our results show that the transcription factors ERF115 and PAT1 are activated during infection, initiating a regeneration program within gall tissues. Ongoing collaborative studies with the PSB/VIB/Ghent University (Belgium), LIMPP/Embrapa-Cenargen and the LBMP / IBqM / UFRJ aim to elucidate the downstream signaling components of this pathway.
Together, these findings deepen our understanding of gall ontogeny, highlight parallels between plant and animal cell strategies for maintaining giant cell integrity, and uncover novel aspects of host regeneration responses. Future efforts will focus on extending these findings to crop species and fully characterizing the ERF115-dependent signaling cascade. Ultimately, overall findings may support the development of innovative biotechnological approaches for sustainable nematode control.
Main References: Doi: 10.1111/nph.19399; Doi.org/10.1111/nph.70066; DOI: 10.1105/tpc.11.5.793; Doi: 10.1111/j.1365-313X.2004.02019.x.; Doi.org/10.1111/nph.70227; Doi.org/10.3389/fpls.2021.636663; Doi:10.1111/nph.16185; Doi: 10.1111/nph.12255.
Jeremie Bazin
Exogenous RNA Targeting of Non-Coding Regions Enables Modulation of Plant Stress Responses and Development
Fabio Ferrario, Soledad Traubenik, Martin Crespi, Jeremie Bazin.
Institute of Plant Sciences Paris-Saclay (IPS2), CNRS, INRAE, Université Paris Saclay –Evry, Université de Paris, Gif sur Yvette, France
Conventional breeding and genetic modification approaches for crop improvement are often time-consuming and may encounter regulatory or societal hurdles. Exogenous RNA applications, such as RNA sprays, represent a promising alternative to transiently modulate endogenous gene expression without altering the genome. Spray-Induced Gene Silencing (SIGS) has emerged as an effective RNA-based strategy for pest control, but its use to directly regulate endogenous plant traits remains largely unexplored.
In this study, we investigated whether double-stranded RNA (dsRNA) delivered via nanoparticles could be used to influence plant development and environmental stress responses. Two experimental systems were analyzed: heat stress tolerance in tomato and cluster root formation in white lupin. Initial attempts to silence genes by targeting exonic coding regions were ineffective in both species, suggesting that endogenous plant genes may be protected from RNA silencing. We therefore shifted focus to non-coding regions of the genome.
In tomato, we characterized alternative splicing of heat stress response genes and identified intron-containing isoforms specifically induced under stress conditions. Targeting these introns with dsRNA enabled isoform-specific silencing, which in turn modulated thermotolerance. In white lupin, dsRNA targeting of precursor miRNAs reduced mature miRNA levels, leading to detectable developmental defects in root architecture.
Together, these findings demonstrate that non-coding genomic regions provide effective targets for exogenous RNA-mediated regulation of plant traits. This work establishes a new strategy to manipulate endogenous gene expression without genetic modification, opening avenues for sustainable, non-GMO approaches to improve plant growth and stress resilience.
Jose Barrero
GM CROPS FOR FOOD SECURITY: THE CASE OF AN INSECT-RESISTANT COWPEA IN NIGERIA
Jose M. Barrero Sanchez - CSIRO Agriculture and Food, Canberra, ACT, Australia
Cowpeas are an essential crop in West Africa, providing vital protein for over 200 million people. However, their production faces severe yield losses due to insect pests, particularly the destructive pod-borer Maruca vitrata. Traditional breeding methods have been ineffective in developing resistant varieties due to the lack of natural resistance, so farmers have relied on chemical insecticides, posing health and environmental risks. To address this problem, our international team developed a Bt pod-borer resistant (PBR) cowpea variety, the world's first genetically modified cowpea, which provides full protection against Maruca. It was released in Nigeria in 2020, providing farmers with an affordable, safe, and accessible solution, which is projected to yield over USD$336 million in benefits over the next 25 years. This project has triggered a transformative effect on seed systems in the region, with the establishment of national seed companies aiming to reach 25% of farmers within five years. The impact of this breakthrough extends beyond Nigeria, as the PBR cowpea was also recently released in Ghana in 2024. The release of a GM food crop in Africa is a significant achievement, and it highlights the pivotal role of biotechnology in addressing global food security challenges and the UN Sustainable Development Goals. Our team is now focused on developing a second-generation PBR cowpea, incorporating two different Bt genes to prevent insect resistance.
Lorenzo Miguel
Abstract
Europe's biotechnology landscape operates as a connected research–industry–policy ecosystem where national societies catalyze talent, translation and public engagement. This talk surveys that ecosystem with a focus on the scientific and community-building roles of Spain's SEBiot and Portugal's Sociedade Portuguesa de Biotecnologia (SPBt), together with Portugal's bioindustry association P-BIO. We highlight how their flagship activities—SEBiot's BIOTEC congress, SPBt's Microbiotec meeting, and P-BIO's BIOMEET—create on-ramps for collaboration across health, agri-food, industrial biotech and the bioeconomy.
Building on this Iberian platform, we present FIAMBIOT, the Ibero-American Federation of Biotechnology founded by Spain, Portugal, Mexico and Brazil to connect scientific societies, industry clusters and early-career communities across regions. We argue that FIAMBIOT is strategically positioned to accelerate talent circulation, cross-testing of pilot/GMP infrastructures, and a shared policy voice on standards and sustainability. As a concrete pathway, we propose a "twin-track" program—Science (microbiomes, biomanufacturing, sustainable processes) and Business (startup dealrooms, investor roundtables, industry days)—co-curated by the Portuguese and Spanish societies with Brazilian and Mexican partners.
Expected outcomes include co-badged sessions, short-term exchanges, co-funded pilots and new consortia. We conclude with a call to action for Brazilian stakeholders to co-lead FIAMBIOT working groups and synchronize Brazil-hosted activities with European events, turning transatlantic goodwill into measurable impact within one annual cycle.
Keywords: Biotechnology, Europe, SEBiot, SPBt, P-BIO, FIAMBIOT, Ibero-America, bioeconomy, international collaboration, talent mobility.
Neena Mitter
RNA based biopesticides for sustainable crop protection
Professor Neena Mitter FTSE FNAAS GAICD. Charles Sturt University, Wagga Wagga, NSW, Australia.
Innovation driving sustainable crop protection resonating with health of planet and the future consumer is key to food and nutritional security and to meet the daily need of two trillion calories for 10 billion people by 2050. Management of pests and diseases in sustainable and safe manner is imperative to meet the targets reduction in pesticide emissions and still meet the growing demand for food and value-added products. The usefulness of chemical pesticides suffers from issues such as residual toxicity, run-off, pest specificity and resistance.
The discovery of RNAi as a natural regulatory mechanism has proved to be a powerful strategy to engineer disease resistance The introduction of RNA interference -based biopesticides based on topical application of double-stranded RNA (dsRNA) presents a transformational opportunity to manage pests and diseases as a biological alternative. RNA based biopesticides or ‘RNA sprays’ for plants without the need for genetic modification is gaining momentum globally, with pests and pathogens targeted with accuracy and specificity.
The efficacy of RNAi-based crop protection depends on multiple parameters across the design–delivery continuum. Particle based delivery of dsRNA and formulation chemistry has the potential to provide stability, rain fastness, enhanced uptake and slow release of dsRNA extending the window of protection as compared to naked dsRNA. Delivery systems, such as Layered Double Hydroxide (LDH) clay nanoparticles (BioClay™) and other delivery approaches are being investigated protection against viruses, insect pests, and fungal pathogens under laboratory and field conditions.
Real-world application of RNA-based biopesticides with sustainable credentials will be governed by factors such as cost-effective production of dsRNA, delivery platforms and formulations, the regulatory landscape and social licensing. This innovative approach represents a significant step towards sustainable, environmentally responsible agricultural pest management as one of the tools in integrated pest and disease management, addressing the urgent need for alternative crop protection strategies.
Nathan Pumplin
Abstract:
The Purple Tomato represents a new chapter in consumer-focused biotechnology. Developed through decades of research in basic plant biology with a focus on improving nutrition, this tomato contains elevated levels of anthocyanins—the same health-promoting bioactives found in blueberries and blackberries—while maintaining the flavor and productivity growers expect. This talk will trace the journey of the Purple Tomato from early genetic discoveries in the laboratory to its path through regulatory review, market testing, and commercial launch. Along the way, I will highlight lessons learned in building trust, navigating public perception, and creating value across the supply chain. The Purple Tomato illustrates how thoughtful innovation can bridge science, agriculture, and consumer choices.
Shuangxia Jin
Genome Editing Tools and Their Applications for Cotton Molecular Breeding
Shuangxia JIN 1*, Xianlong ZHANG 1, Zhongping XU1, Lu YU1, Guangying WANG1.Fengjiao Hu1, Muna ALARIQI1
Corresponding author: *jsx@mail.hzau.edu.cn
1 National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, China
Genome editing tools for cotton: CRISPR/Cas9, Cas12a, Cas12b, Cas13, Off-target analysis, Base editors, dCas9-TV (transcription activation). Cotton (G. hirsutum) is an allotetraploid with a complex genome. Most genes have multiple copies that belong to At and Dt subgenomes. Sequence similarity is also very high between gene homologues. To efficiently achieve site/gene-specific mutation is quite needed. In 2018, we successfully developed CRISPR/Cas 9 system with an average 65-85% efficiency. Then, we performed the whole genome sequencing to investigate the off-target in the CRISPR/Cas9 edited cotton plants. The results showed that of 2000+ potential off-targets sites, the WGS data revealed that only four are bona fide off-target mutations suggested that CRISPR/Cas9 system is highly specific for the editing of genes of polyploid plant species. Then, we further developed a high-throughput genome editing system in cotton. A sgRNAs library (containing 1100 sgRNAs targeted to 600 independent genes) was constructed and cloned into the CRISPR/Cas 9 vector. By this way, we can edit several hundred target genes in one transformation. Recently, CRISPR/Cas12a and Cas12b were developed and both are belonging to class 2 CRISPR/Cas system with features distinct from those of Cas9. Our data showed that both work very well with high efficiency and accuracy in cotton. At the same time, we developed several base editing tools (CBE, ABE8e, Dural base editor) for cotton by fusion of dCas9/nCas9 with activation-induced cytidine deaminase (AID) or engineered adenosine deaminase (TadA), creating specific point mutations. Most recently, several CRISPR/Cas13 systems (13a, b, c, d) were developed in cotton to target the RNA for down-regulating the gene transcription (knock-down). On the contrary, a transcription activation (dCas9-TV) system was establishing by fusion transcriptional activators VP64, TAL and EDLL with dCas9 to knock-up gene’s transcription.
Cotton Molecular Breeding using Genome editing: The high oleic acid content, nontransgenic allotetraploid cotton was generated by knockout of GhFAD2 genes with CRISPR/Cas9 system in our lab. Similarly, by targeting GhPGF with Cas12, we efficiently created non-transgenic and gossypol-free cotton plants. These gossypol-free, high-oleic, nontransgenic mutants provide useful parents in breeding programs to introgress agronomically valuable traits into commercial cotton varieties. Moreover, we have extended the application of base editors to cotton plants by editing the non-coding and coding regions of the GhTFL1 gene in a high-throughput approach to create ideotype cotton germplasm with moderate height, shortened fruiting branches, compact branches. In addition, an in vivo maternal haploid induction system was developed in cotton by knock out the GhDMP through CRISPR/Cas9 exhibits a haploid induction rate of 1.06% in F1 progeny as the haploid inducer line. Most recently, the optimized dCas9-TV system was recruited to activate the GhEPSPS to generate glyphosate-resistant cotton with the transcription of GhEPSPS increased by 16-fold, resulting in moderate resistance to glyphosate without growth deficit. Dr.Jin’s lab has published 20 peer review papers regarding cotton genome editing (https://www.researchgate.net/profile/Shuangxia-Jin) .
Stefanie Menezes
Resumo 1:
Title: Innovative dsRNA designs for RNAi-based pest control: paperclip RNA and structured viroid-like approaches
Authors: Stefanie Menezes de Moura; Thuanne Pires Ribeiro; Nazanin Amanat; Leonardo Lima Pepino Macedo; Steve Whyard; Maria Fátima Grossi-de-Sá; Juan Luis Jurat-Fuentes
Presenter: Stefanie Menezes de Moura
Institution: University of Tennessee, Knoxville (UTK), TN, USA
Section: 1.2 - Formulations and Nanobiotechnology in Agriculture
Abstract
The lepidopteran pests Spodoptera frugiperda (fall armyworm, FAW) and Helicoverpa armigera (cotton bollworm, CBW) are major threats to global agriculture. Insecticidal gene silencing by RNA interference (RNAi) has emerged as a promising and sustainable strategy for pest management; however, its efficacy in these species is often limited by physiological barriers
and inefficient RNA delivery. In this study, we evaluated the performance of two structured dsRNA designs: (i) paperclip RNA (pcRNA) and (ii) viroid-like structured dsRNA constructs. When tested in diet-based assays, pcRNA molecules exhibited increased stability in FAW larvae gut extracts compared to linear dsRNA and induced specific gene silencing. Bioassays showed that pcRNA formulations targeting individual genes reduced FAW adult emergence, while pcRNA cocktails combining multiple targets produced the highest mortality rates, indicating additive or synergistic effects. In contrast, a viroid-structured dsRNA pyramiding three genes for FAW did not induce significant mortality when delivered via diet-based application. Complementarily, when the viroid-like structured dsRNA was expressed in transgenic tobacco plants transformed
via nuclear or chloroplast integration, both transformation strategies resulted in stable transgene expression and efficient silencing of target genes in insects feeding on transgenic tissues. Feeding assays demonstrated larval mortalities exceeding 70% for both FAW and CBW when fed on transformed leaves, compared to insects feeding on wild-type plants. Altogether, the data obtained
so far in both lepidopteran pests suggest the superior performance of the paperclip RNA design for multiplex gene targeting in non-transformative delivery, and high efficiency of the viroid-like structured RNA in transformative, plant-based delivery. These findings underscore the importance of RNA structure and delivery context for optimizing RNAi efficacy and advancing nextgeneration RNA-based pest control technologies.
Keywords: Spodoptera frugiperda, Helicoverpa armigera, RNAi, paperclip RNA, viroid-like dsRNA, gene silencing, pest management.
Resumo 2:
Title: Symbionts to Solutions: Targeting Agricultural Pests with Nematode-Associated Bacteria
Authors: Stefanie Menezes de Moura, Heba Abdelgaffar, David Shapiro-Ilan, Shaohui Wu, Juan Luis Jurat-Fuentes
Presenter: Stefanie Menezes de Moura
Institution: University of Tennessee, Knoxville (UTK), TN, USA
Section: 2.3 – Biotechnology in Alternative Proteins – Focus on Pesticide Proteins
Abstract
The development of environmentally sustainable biopesticides is essential to address the growing challenges of insect resistance and the ecological impacts of chemical insecticides. Bacteria such as Photorhabdus luminescens, a symbiont of entomopathogenic nematodes (EPNs), and Bacillus thuringiensis (Bt) are promising sources of insecticidal proteins and metabolites for sustainable pest management. In this study, we evaluated the insecticidal activity of cell-free supernatant and cell pellet fractions from a strain of P. luminescens and B. thuringiensis serovar berliner ATCC 10792 against four major agricultural pests: Spodoptera frugiperda (fall armyworm, FAW), Helicoverpa zea (corn earworm, CEW), Ostrinia nubilalis (European corn borer, ECB), and Leptinotarsa decemlineata (Colorado potato beetle, CPB). After seven days of exposure, both P. luminescens fractions (supernatant and pellet) caused 100% mortality in CPB larvae, while Bt fractions also exhibited high toxicity, with supernatants inducing 100% mortality and pellets exceeding 50%. In contrast, none of the fractions from either bacterium caused statistically significant mortality in the lepidopteran species when compared to the negative control (TSY medium). To investigate whether the toxic activity observed against CPB was associated with proteinaceous components or small-molecule metabolites, additional bioassays were conducted with proteinase K–treated samples. Remarkably, protease treatment did not affect the toxicity of either P. luminescens or Bt fractions against CPB larvae, suggesting that the active insecticidal compounds are non-proteinaceous metabolites. These findings highlight the potent nature of the bioactive compounds produced by P. luminescens and reinforce its potential as a versatile source of insecticidal metabolites for pest control. Ongoing analyses aim to further characterize these
bioactive molecules and insecticidal proteins in these samples. Overall, these results reinforce the potential of both P. luminescens and B. thuringiensis as valuable microbial resources for the development of novel biopesticides targeting lepidopteran and coleopteran pests.
Keywords: Photorhabdus luminescens, Bacillus thuringiensis, entomopathogenic bacteria,
biopesticides, insecticidal proteins, sustainable agriculture
Yiping Qi
Resumo 1: opening plenary
Promoting Plant Regeneration and Genome Editing in Multiple Crops by CRISPR-Combo
Yiping Qi
Department of Plant Science and Landscape Architecture, University of Maryland, College Park.
Email: yiping@umd.edu
RNA-guided CRISPR activation (CRISPRa) systems such as CRISPR-Act3.0 have been developed for the efficient activation of endogenous plant genes. Based on CRISPR-Act3.0, we further developed a versatile CRISPR-Combo platform for simultaneous genome editing (targeted mutagenesis or base editing) and gene activation in plants. We reasoned that by activation of endogenous morphogenic genes, we may be able to promote plant tissue culture and regeneration of genome-edited plants. It is indeed a great success when demonstrating this idea in rice. Here, we expanded this approach to screen a list of morphogenic genes in multiple dicot species, including potato, strawberry, citrus, and populus. We have successfully identified promising morphogenic genes in all these plant species, where their activation by CRISPR-Combo confers enhanced plant tissue regeneration and efficient genome editing. Conventionally, a gene stacking approach is used to express morphogenic genes to aid plant tissue culture and regeneration of genome-edited plants. By contrast, our CRISPR-Combo approach relies on a single Cas9 protein with multiplexed gRNAs, presenting a simple and promising strategy to accelerate and revolutionize plant genome engineering in many crops.
Keywords: CRISPR activation, CRISPR-Combo, morphogenic genes, plant regeneration, genome editing
Resumo 2:
CRISPR-Cas12a genome editing tools for plant research and crop improvement
Yiping Qi
Department of Plant Science and Landscape Architecture, University of Maryland, College Park.
Email: yiping@umd.edu
CRISPR-Cas12a (Cpf1) has rapidly emerged as a versatile genome editing tool with unique features that make it particularly attractive for plant research and crop improvement. Compared with earlier sequence-specific nucleases, Cas12a provides distinct advantages, including self-processing of crRNA arrays, a T-rich PAM requirement, and the ability to generate staggered DNA breaks that facilitate precise editing outcomes. In this presentation, I highlight the development and application of Cas12a systems across diverse plant species. First, optimized expression systems were established that enable highly efficient and often biallelic editing in crops such as rice and maize. Second, Cas12a was demonstrated to efficiently disrupt microRNA genes, with downstream effects on plant traits such as grain quality. Third, Cas12a-based promoter editing, termed CRISPR-Cas12a promoter editing (CAPE), was used to fine-tune quantitative traits by modulating cis-regulatory elements, exemplified by engineering starch biosynthesis and plant architecture genes. Fourth, Cas12a has been harnessed for precise gene replacement through homology-directed repair (HDR) as well as for multiplexed cytosine and adenine base editing, broadening the scope of targeted nucleotide modifications. Finally, protein engineering and temperature optimization have produced next-generation Cas12a variants with superior activity, broader PAM recognition, and improved performance in both monocot and dicot species, including rice and poplar. Collectively, these advances establish CRISPR-Cas12a as a robust and flexible platform for plant functional genomics and trait development, with wide-ranging applications from basic research to crop biotechnology. Ongoing collaborations and technology improvements will continue to expand its impact on agriculture.
Keywords: CRISPR-Cas12a, T-rich PAM, large deletions, plant genome editing, crop improvement