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Decrypting the role of bacterial signals in microbial interactions to enhance Lysobacter establishment in the rhizosphere

Periodic Reporting for period 1 - RhizoTalk (Decrypting the role of bacterial signals in microbial interactions to enhance Lysobacter establishment in the rhizosphere)

Reporting period: 2018-06-15 to 2020-08-14

Bacterial biocontrol agents represent a promising strategy for the sustainable management plant pathogens

In agriculture, beneficial microorganisms thriving in association and interacting with plant roots play an essential role in plant growth and disease suppression and represent a potential solution to replace chemical fertilizers and pesticides. Beneficial bacteria displaying biocontrol activities, for instance, are regarded as a promising strategy for the sustainable management of soil-borne plant pathogens. However, their widespread use is still limited, mainly due to their difficulty to adapt to new conditions in the soil and integrate within indigenous soil microbial communities.
The recruitment and maintenance of the soil microbiota is conditioned by plant root exudates, as well as by signalling molecules produced and secreted by both plants and microbes. Bacterial signalling occurs mainly via quorum-sensing (QS) allowing bacterial communities to form and synchronize their behaviour. It is therefore conceivable that, when arriving in the soil, newcomer biocontrol bacteria will perceive and interpret these signals coordinating their establishment and controlling their functioning in the soil microbial community.
The Marie Skłodowska-Curie Action RhizoTalk focused on solving this problem and dedicated to increase knowledge of molecular interactions between microorganisms in the soil by unveiling the response of the model bacterial BCA Lysobacter capsici AZ78 (AZ78) to main bacterial QS signals in the soil and developing consortia formulations that guarantee the establishment bacterial BCA in the field.
The implementation of RhizoTalk was broken down in five work packages (WPs), three of which were entirely result-oriented. WP1 was devoted to the identification of communication systems in AZ78 and the characterization of its response to main soil bacterial communication signals. WP2 was devoted to the identification of soil bacterial strains producing signals relevant for AZ78 and the characterization of bipartite interactions between those and AZ78. The third WP (WP3) was instead dedicated to the development and validation of biocontrol formulations of AZ78 in consortium with other bacterial species. WP4 and WP5 addressed dissemination and outreach activities, as well as issues related to career development and project management, and consequently lasted during the entire duration of the project.

A transcriptome approach to biopesticide development

The presence of genes involved in cell-cell signalling systems in the genome of AZ78 was determined by genome mining, and production of quorum-sensing signals by AZ78 was evaluated using reporter strains. Next, a functional analysis of bacterial signalling compounds was performed in vitro. Firstly, at a microbiological level, by evaluating the effect of main QS signals on AZ78 biocontrol activity, and secondly, at a molecular level by evaluating the transcriptome response of AZ78 to QS molecules by RNA-Seq. Our findings showed that AZ78 perceive and produce different kinds of QS signals. Moreover, it allowed for the identification of important genes modulated by QS signals, shedding a light on the molecular mechanisms involved in AZ78 establishment in the soil.

Bacterial formulations as biopesticides against soil borne pathogens

A collection of bacterial isolates deriving from copper contaminated soils was screened for pathogen inhibition in binary combinations with AZ78. Each strain was tested for potential enhancement of AZ78 biocontrol activity and tolerance to desiccation and UV light. Strains were taxonomically identified using 16S rDNA PCR and characterised for their plant growth promoting traits and their ability to produce QS signals. Strains enhancing AZ78 biocontrol activity and resistance to environmental stresses were designated as helper bacterial strains (HBS) and were further formulated together with AZ78. AZ78 and HBS were blended, supplemented with additives and incorporated to inorganic carriers to produce water dispersible granules. A total of 16 formulations consisting of AZ78 and HBS were developed and characterized. Shelf life was monitored under different storage conditions and best performing formulations were validated under greenhouse conditions.

Results derived from RhizoTalk have been disseminated in conferences and seminars. Moreover, RhizoTalk have given rise to a book chapter, and 3 peer-reviewed publications will be soon available. Additionally, RhizoTalk has taken part in events addressed to the public like the European Researchers' Night.
The Marie Skłodowska-Curie Action RhizoTalk has, in particular, contributed to determine the impact that QS signals have on AZ78 behaviour, uncover mechanisms involved in the interaction between Lysobacter BCAs and other soil-living bacteria, and ultimately generate knowledge that can be translated into agricultural applications. All in all, the outcome of RhizoTalk supposes a milestone in environmental microbiology. By means of novel conceptual frameworks invigorated by modern molecular and chemical techniques, Rhizotalk has contributed to highlight belowground interactions and shed light on their ecological roles. Finally, results achieved in RhizoTalk might fill the knowledge gap that severely hampers our ability to understand and predict the functioning of the soil biota. Nevertheless, further work is still needed to translate knowledge of microbial interactions into a predictable science that could be harnessed.
In view of future, we are currently exploring the possibilities of additional funding and material transfer agreements with agricultural companies for consolidating this research line.