Periodic Reporting for period 1 - SMART-AGRI-SPORE (Fighting the X. fastidiosa threat to EU agriculture - Novel smart biopesticides based on bacterial spores)
Reporting period: 2021-09-01 to 2024-08-31
The Mediterranean region represents one of the world’s most important agricultural production zones and is therefore inherently susceptible to emerging plant diseases and dependent on the availability of appropriate pesticides. In particular, olive agriculture presents an integral economic pillar, with 95% of the world’s olive tree population concentrated in this area. However, orchards are exposed to an unparalleled threat, caused by the bacterium Xylella fastidiosa, which is considered one of the most dangerous plant pests worldwide. Notwithstanding, currently no treatment of diseased trees under field conditions is available. At the same time, the European Union (EU) has committed to a sustainable food strategy through ambitious legislative proposals, such as the European Commissions’ Green Deal and the EU Farm to Fork strategy, which have a clear focus on reducing the dependency on pesticides and fostering the use of non-chemical alternatives. Nonetheless, plant diseases are omnipresent and plant protection products are indispensable to safeguard crops. Hence, these legislative requirements will have a considerable impact on both EU and international farming. Given that at this point, the availability and efficiency of non-chemical plant protection products are still limited, viable and effective alternatives are urgently needed.
Thus, the aim of this project was to develop a novel biological plant protection system (SMART-AGRI-SPORE) targeting X. fastidiosa infections in olive groves. It combined for the first time the use of bacterial spores with the controlled release of antimicrobial peptides in an agricultural application to provide a sustainable alternative to chemical pesticides.
The work could show that several indigenous spore-formers of Spanish olive groves inherently possess antagonistic activity against X. fastidiosa. These bacteria were derived from the soil, as well as from the leaf interior (endophytic).
Thus, the aim of this project was to develop a novel biological plant protection system (SMART-AGRI-SPORE) targeting X. fastidiosa infections in olive groves. It combined for the first time the use of bacterial spores with the controlled release of antimicrobial peptides in an agricultural application to provide a sustainable alternative to chemical pesticides.
The work could show that several indigenous spore-formers of Spanish olive groves inherently possess antagonistic activity against X. fastidiosa. These bacteria were derived from the soil, as well as from the leaf interior (endophytic).
During the project, 412 indigenous Bacillus strains were isolated from soil and leave samples derived from 5 olive orchards in the south of Spain (Jaén and Málaga region) termed the culturable olive sporobiota of Spanish olive groves. Selected candidates were identified by rep-PCR and sequencing and then characterised regarding environmental tolerance and resistance properties.
Based on the obtained results, the culturable olive sporobiota was screened for antimicrobial activity/ production of antimicrobial agents against 2 strains of Xylella fastidiosa as well as common human and plant pathogens. 5 candidates were of particular interest, as they showed antagonistic activity against Xylella fastidiosa. This activity was detected in bacterial cultures as well as cell free culture extracts. Another interesting finding was that these spore-formers were derived both from the soil close to the olive trees and the leaf interior. The 5 candidates were characterised in detail regarding their biotechnological and biochemical properties, as well as through genetic approaches.
The project then focused on the genetic engineering of Bacillus spp. to generate functionalised bioparticles with anti-Xylella fastidiosa activity. Following a combinatorial approach a cohort of Bacillus strains was engineered with different combinations of active agents and delivery systems. Culturable olive sporobiota strains with anti-Xf activity as well as genetically engineered Bacillus strains were then assessed and characterised.
In summary, the most important findings of our study were the identification and characterisation of the culturable sporobiota of Spanish olive groves. We furthermore identified 5 bacterial strains belonging to the Bacillus and Peribacillus clade, derived from the soil and plant interior (endophytic) with strong antagonistic activity against Xylella fastidiosa and other detrimental plant pests (e.g. Verticillium dahliae).
These results have been disseminated through open access articles in international peer-reviewed journals such as Microbiology Spectrum, Pest Management Science or Microbiology. The research has further been presented to scientific peers, stakeholders and policymakers through presentations in several international conferences, such as the 11th European Spore Conference, Encuentro Internacional Phytoma, or a stakeholder meeting with the International Olive Council. Finally, results of the project have been made available to the general public through several publications in media journals, radio interviews and News Op Eds.
Based on the obtained results, the culturable olive sporobiota was screened for antimicrobial activity/ production of antimicrobial agents against 2 strains of Xylella fastidiosa as well as common human and plant pathogens. 5 candidates were of particular interest, as they showed antagonistic activity against Xylella fastidiosa. This activity was detected in bacterial cultures as well as cell free culture extracts. Another interesting finding was that these spore-formers were derived both from the soil close to the olive trees and the leaf interior. The 5 candidates were characterised in detail regarding their biotechnological and biochemical properties, as well as through genetic approaches.
The project then focused on the genetic engineering of Bacillus spp. to generate functionalised bioparticles with anti-Xylella fastidiosa activity. Following a combinatorial approach a cohort of Bacillus strains was engineered with different combinations of active agents and delivery systems. Culturable olive sporobiota strains with anti-Xf activity as well as genetically engineered Bacillus strains were then assessed and characterised.
In summary, the most important findings of our study were the identification and characterisation of the culturable sporobiota of Spanish olive groves. We furthermore identified 5 bacterial strains belonging to the Bacillus and Peribacillus clade, derived from the soil and plant interior (endophytic) with strong antagonistic activity against Xylella fastidiosa and other detrimental plant pests (e.g. Verticillium dahliae).
These results have been disseminated through open access articles in international peer-reviewed journals such as Microbiology Spectrum, Pest Management Science or Microbiology. The research has further been presented to scientific peers, stakeholders and policymakers through presentations in several international conferences, such as the 11th European Spore Conference, Encuentro Internacional Phytoma, or a stakeholder meeting with the International Olive Council. Finally, results of the project have been made available to the general public through several publications in media journals, radio interviews and News Op Eds.
Most importantly, the project has identified new candidates for the development of sustainable plant protection products based on biological agents, notably with an activity against one of most detrimental plant pests for olive agriculture, i.e. Xylella fastidiosa but also other phytopathogens (e.g. Verticillium dahliae). The SMART-AGRI-SPORE project has furthermore provided important insights into the composition and characteristics of the spore-forming community of Spanish olive groves and described its genetic make-up and biotechnological properties. Finally, the project has developed a first prototype of a biological particle (SMART-AGRI-SPORE) with the potential to fight X. fastidiosa infections under field conditions.
Results derived from the project therefore can contribute to: (1) provide novel solutions for the treatment of X. fastidiosa; (2) greatly further our understanding of the use of biological compounds as non-chemical pesticides; as well as (3) expand our knowledge of targeted and sustainable compound delivery.
Results derived from the project therefore can contribute to: (1) provide novel solutions for the treatment of X. fastidiosa; (2) greatly further our understanding of the use of biological compounds as non-chemical pesticides; as well as (3) expand our knowledge of targeted and sustainable compound delivery.