Epidemiological surveillance of Ralstonia solanacearum, causal agent of bacterial wilt of solanaceous crops, in the South-West Indian Ocean islands and Eastern Africa: diversity and genetic structure of populations

Date de début de projet :

01/01/2015

Date de fin du projet :

01/01/2017

Objectifs

Epidemiological surveillance of plant pathogens allowing identification of emergingclones escaping control strategies and tracing of bacterial strains are of greatimportance for integrated plant protection and require extremely precise strainidentification. Ralstonia solanacearum is the causal agent of bacterial wilt attackingmore than 200 host species. Bacterial wilt is considered among the most damagingplant bacterial diseases worldwide (Mansfield et al. 2012). R. solanacearum has awide genetic diversity and virulence /aggressiveness and different lineages canshare the same ecological niche making this pathogen an excellent model forunderstanding epidemiological and evolutionary aspects of pathogen-hostinteractions. Though this plant pathogen is present in the Indian Ocean andEastern Africa, no data is available about its diversity and structure in this region.Our research project hence represents a pioneering study aiming to create anepidemiosurveillance network within the framework of partnerships betweeninternational research and education institutions (labex Agro Units, CGIARs,Universities), a national reference laboratory (ANSES-LSV), professional actors inthe agricultural world, and a start-up.The objectives of this project are to (i) thoroughly analyze the genetic diversity ofR. solanacearum in South-West Indian Ocean (SWIO) islands and Eastern Africa,(ii) characterize the evolutionary forces that shape populations of R. solanacearumin these geographic areas, (iii) evaluate genetic resources for resistance to3 of 21bacterial wilt (mainly for potato), and to (iv) validate molecular diagnostic toolsavailable or under development. This project will set up the epidemiologicalsurveillance of this pathogen in the Indian Ocean and Eastern Africa and ultimatelywill give guidance to decision makers in order to establish control strategies at alocal level, such as the choice of resistant varieties.To achieve these objectives, several actions are planned: (i) field surveys in eachcountry (collection of populations) ; (ii) molecular characterization of strains; (iii)genetic structure analysis of bacterial populations at different spatial and temporalscales; (iv) evaluation of genetic resources for resistance to bacterial wilt; (v)development and validation of diagnostic tools; and (vi) organization of kick-offand workshop meetings and training sessions for knowledge sharing and capacitybuilding both on R. solanacearum and the technologies.

Localisation

Réunion, Comoros, Mauritius and Seychelles

Description

A major initial effort will be required for sampling and collecting epidemiological and historical strains of R. solanacearum strains present on vegetable range of host crops (potato, tomato, eggplant...) across the SWIO islands (Réunion, Comoros, Mauritius and Seychelles) and Eastern Africa (Tanzania, Kenya). This step constitutes the cornerstone of the project and will be conducted through a statistical approach using a georeferenced sampling scheme (GPS). Sampled strains will complement the existing R. solanacearum collection from Mayotte (155 strains collected in 2012) and Madagascar (over 2500 strains collected in 2013) that are being analyzed in the context of S. Ravelomanantsoa’s PhD thesis (2013-2016) within a collaboration between UMR PVBMT and FIFAMANOR.

These strains will then be subjected to a thorough molecular characterization. First, determination of phylotype by multiplex PCR and of sequevar by endoglucanase (egl) gene sequencing will be conducted (Fegan and Prior, 2005). Second, genetic diversity within the R. solanacearum strains will be analyzed by MLSA/MLST. The approach will establish phylogenetic relationships among strains of R. solanacearum originated from SWIO islands and Eastern Africa and their position relative to worldwide reference strains already typed (Wicker et al., 2012). Furthermore, a MLVA scheme targeting tandem repeats with a rapid evolution rate will be used in order to decipher the evolutionary forces acting at the level of populations and therefore their structure. Several scales of analysis will be considered: country, island, region, and field. Spatial analyses will be performed at those different scales. Temporal analyses will be carried out on collections from Reunion to evaluate the level of clonality of R. solanacearum. Population genetics and molecular epidemiology analyses tools will be used to describe the intra- and inter- genetic diversity of these populations and the relationships between the individuals. Pathways of introduction and spread will be inferred using Bayesian scenario choice analyses using the genetic proximity of collected strains against an international reference collection, owned by our team in Reunion (INRA-CIRAD collection “RUN”), comprising over 3200 entries. RUN collection covers all phylotypes and sequevars of R. solanacearum described worldwide. These strains are characterized and maintained by cryoconservation and lyophilization. This collection will constitute a great asset to strengthen and optimize responsiveness in epidemiological surveillance, as molecular tools can be assessed and validated against a great diversity. This work will seek for new diagnostic markers and protocols in order to propose accurate, affordable, and convenient technologies suitable for these regions. Data originating from the population diversity study and the development of microsatellite markers will be strategically used for new application-oriented diagnostics. Furthermore, these tools and protocols will be shared within the SWIO and Eastern African laboratories. 
The genotyping data will be shared among partners and eventually been made public once scientific papers are released. For this, we will use the MLVA bank 8 of 21 database (http://www.biopred.net/MLVA/) which has been developed by two of the partners (UMR PVBMT and RPB/IPME) of the present project during a previous ANR-funded project (ANR-2010-BLANC-1723). The website allows viewing databases with sorting and clustering options, submitting queries, and sharing databases which are maintained and managed by different owners once a common agreement is achieved among partners. This database can handle different types of allelic data (MLVA, MLST, CRISPR…).

Finally, strains representative of the genetic diversity of R. solanacearum will be inoculated to solanaceous plant varieties in the P3 facilities held in CIRAD Reunion laboratories, in Kenya and in Mauritius to investigate the phenotypic diversity (virulence and aggressiveness) of the strains and then to identify genetic resources for resistance to bacterial wilt.

Partenaires

  • ANSES-Laboratoire de Santé des Végétaux - Unité Ravageurs et Agents Pathogènes Tropicaux - Réunion
  •  FDGDON (Fédération Départementale de Groupement de Défense contre les Organismes Nuisibles) - Réunion
  •  ARMEFHLOR (Association Réunionnaise pour la Modernisation de l’Economie Fruitière Légumière et Horticole) - Réunion
  •  VitroRun - Réunion
  • CRP RTB (CGIAR research program on Roots-Tubers-Bananas)
  •  IITA (Tanzania, Kenya)
  •  CIP (Kenya)
  •  University of Comoros
  •  INRAPE (Institut National de Recherche pour l’Agriculture, la Pêche et l’Environnement) - Comoros
  •  SAA (Seychelles Agricultural Agency) - Seychelles 
  •  University of Mauritius 
  •  FAREI (Food and Agricultural Research and Extension Unit) - Mauritius

Equipe

Poussier Stéphane,Professor, UMR PVBMT (University of Reunion)
Prior Philippe, Research Director UMR PVBMT(INRA-CIRAD)
Guérin Fabien, Associate professor UMR PVBMT (University of Reunion)
Pruvost Olivier,Researcher UMR PVBMT (CIRAD)
Vernière Christian,Researcher UMR PVBMT (CIRAD)
Boyer Karine, Lab assistant UMR PVBMT (CIRAD)
Chéron Jean-Jacques,Lab assistant UMR PVBMT (CIRAD)
Lebon Sylvain, field technician UMR PVBMT (CIRAD)
Cellier Gilles, Project manager, PhD ANSES- LSV Reunion
Hostachy Bruno,Laboratory unit head manager ANSES - LSVReunion
Yahiaoui Noura, PhD student CIRAD-ANSES Reunion
Ravelomanantsoa, Santatra PhD student CIRAD Reunion
Neema Claire, Professor UMR BGPI (Montpellier Supagro)
Ravigné Virginie,Researcher UMR BGPI(CIRAD)
Verdier Valérie,Researcher UMR RPB/IPME (IRD)
Koebnik Ralf , Researcher UMR RPB/IPME(IRD)
Minatchy Janice,Laboratory manager FDGDON
Willy Suzanne, Business manager VITRORUN
Bory Séverine,Researcher VITRORUN

Hamza Abdou Azali,associate professor University of Comoros
Issimaila Mohamed, head INRAPE, Comoros
Naiken Marc, Chief executive officer Seychelles Agricultural Agency
Petrousse Bobb,Laboratory technologist Seychelles Agricultura lAgency
Jaufeerally-FakimYasmina, Professor University of Mauritius
Benimadhu Shyam, Headplant pathology division FAREI - Mauritius
Coyne Danny, Researcher IITA -Kenya/Tanzania
Parker Monica,Researcher CIP - Kenya

Financement

Agropolis Fondation

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