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Streptomyces scabies is the causative agent of the common scab (CS) disease on tuber and root crops (potatoes, radishes, beets,…). Studies on S. scabies 87-22 are focused on understanding the molecular mechanisms and the molecules that confer this strain its virulent properties. The accessibility of its genome sequence and its subsequent analysis helped further revealing new strain-specific virulence determinants and additional features for this phytopathogen. 

The first objective of my Master II thesis is in line with previous works that is, deepening the knowledge related to the mechanisms involved in the molecular interactions existing with its host plants. This was performed by accurate genome mining of S. scabies. The biosynthetic gene clusters (BGCs) coding for (secondary) specialized metabolites predicted via the antiSMASH software have been subjected to a comparative study in order to assess, in terms of quantity and diversity, the cryptic BGCs for which no homologous BGC is known so far and therefore involved in the production of unknown compounds. Our work revealed that S. scabies possess 45 BGCs; beyond the 17 known BGCs mainly including siderophores and phytotoxins next to the core metabolites, we revealed 28 uncharacterized cryptic BGCs with poor or no homology to the currently known BGCs. Some of these BGCs might be involved in the production of cryptic new virulence key determinants, which will be confirmed or infirmed by further investigations.
The second objective of my thesis was to analyze the expression of the genes composing these 45 BGCs under culture conditions triggering the production of the main virulence determinant thaxtomin A. This work was performed by a transcriptomic study on RNA extracted when S. scabies was cultivated on minimal media containing the best known virulence triggering factors, namely cellobiose and cellotriose. With this approach we noticed that most plant-associated compounds of S. scabies followed the same transcriptional response as thaxtomin phytotoxins. The expression of most siderophore BGCs was also significantly enhanced thereby highlighting the crucial role of iron acquisition in virulence. Interestingly, the expression of several cryptic BGCs, and therefore, new molecules, was also drastically modified under virulence conditions suggesting that other and yet unknown molecules would play a significant role in the plant colonization/interaction by S. scabies.
Finally, the third objective of this work is related to the classification of S. scabies as a plant pathogen. Indeed, although CS is responsible for significant economic losses, the symptoms mostly affect the visual appearance of the vegetable, and the disease has never been reported as lethal. Could S. scabies instead contribute to the protection of its host against much more harmful pathogens by secreting antimicrobial compounds? In this work we therefore aim to provide the first evidence for answering the challenging question: “Is Streptomyces scabies really a plant-pathogen or instead a plant-protecting bacterium?”.

In order to answer this question, we assess the potential of S. scabies to produce bioactive compounds with antifungal and anti-oomycete activity by i) genome mining (also performed for the transcriptomic study), and ii) bioactivity assays. Our main focus was on the production of bioactive compounds against Phytophthora infestans - the oomycete responsible for late blight that also affects potato crops. In addition, interaction of S. scabies in antagonizing tests with ascomycetes plant pathogens such as Alternaria solani, Gibberella zeae, and Fusarium culmorum was studied as well. We reveal that S. scabies is able to produce antimicrobials (both diffusible and volatile compounds) against all pathogens tested. The identification of these compounds is key to fully understand the role of S. scabies as host for root and tuber plants.

Sciences du vivant > Microbiologie

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The research topic investigated during this master thesis focused on the induction of the pathogenic behaviour in Streptomyces scabies. The Streptomyces genus, well-known for its extensive production of valuable bioactive compounds, includes – among many saprophytic species – a few pathogens with S. scabies 87-22 as model species. This Gram-positive bacterium is responsible for the common scab disease affecting potatoes and other tuber plants in the fields, causing substantial economic losses. The signalling pathway leading to the production of the main phytotoxin – namely thaxtomin A – produced by S. scabies upon cello-oligosaccharides (cellobiose and cellotriose) feeding was recently described. However, the model is likely more complex than previously thought, and some key features of this model still need to be confirmed. We decided to investigate the genetic adaptations – either at the gene level or more subtle genetic mutations – responsible for the development of a pathogenic lifestyle in the Streptomyces genus.
This work was divided in two parts:
• The first aspect that we studied was the role of the beta-glucosidase BglC in the induction pathway of thaxtomin A biosynthesis. The mutant of S. scabies defective in bglC (ΔbglC) displayed two unexpected phenotypes regarding thaxtomin A production, i.e. reduced and increased production levels in the presence and absence of cello-oligosaccharides, respectively. Our study of the cello-oligosaccharides consumption and beta-glucosidase activity in the mutant allowed us to propose the awakening of alternative beta-glucosidase(s) as explanation for the reduced thaxtomin production observed in presence of cello-oligosaccharides. Our virulence bioassays confirmed that the overproduction of thaxtomin A by the bglC null mutant results in hypervirulence in media devoid of cello-oligosaccharides.
• The second aspect of our research focused on the impact of point mutations on the DNA-binding ability of CebR – the master repression of thaxtomin A production – when different cis-acting elements are encountered. In this work, we confirmed that single-nucleotide mutations caused a substantial and meaningful decrease of the affinity of CebR. Our results strengthen the idea that cellotriose, which is not the best allosteric effector of CebR, is likely able to remove CebR from its cis-acting binding sites located in the thaxtomin biosynthesis gene cluster and induce the pathogenic lifestyle of S. scabies.
Finally, our work brought new insights into the understanding of the mechanisms governing the induction of pathogenicity in S. scabies and opened new horizons for this research project.

Streptomyces scabies --- Pathogen --- Scab disease --- Thaxtomin A --- CebR --- BglC --- Signalling pathway --- Virulence induction --- Sciences du vivant > Microbiologie --- Sciences du vivant > Biochimie, biophysique & biologie moléculaire