title: A new typing technique for the Rickettsiales Ehrlichia ruminantium: multiple-locus variable number tandem repeat analysis.
abstract: Ehrlichia ruminantium (ER) is a member of the order Rickettsiales transmitted by Amblyomma ticks. This obligatory intracellular bacterium is the causative agent of a fatal disease in ruminants, named heartwater. It represents a constraint on breeding development in sub-Saharan Africa and in the Caribbean. The genetic diversity of the strains of ER, which could be a limiting factor to obtain effective vaccines, needs to be better characterized. For this purpose, we developed a molecular typing technique based on the polymorphism of variable number tandem repeat (VNTR) sequences, MLVA (multiple locus VNTR analysis). Eight (out of 21) VNTR candidates were validated using 17 samples representing a panel of ER strains from different geographical origins from West, South Africa, and Caribbean areas and in ER infected ticks and goat tissues. This result demonstrated the ability of these VNTRs to type a wide range of strains. The stability of the selected VNTR markers was very good, at the time scale needed for epidemiological purposes: in particular, no difference in the VNTR profiles was observed between virulent and attenuated strains (for Gardel and Senegal strains) and between strains (Gardel and Blonde strains) isolated in the same area 19years apart. We validated the strong discriminatory power of MLVA for ER and found a high level of polymorphism between the available strains, with 10 different profiles out of 13 ER strains. The MLVA scheme described in this study is a rapid and efficient molecular typing tool for ER, which allows rapid and direct typing of this intracellular pathogen without preliminary culture and gives reliable results that can be used for further epidemiological studies.
authors.affiliation: UPE, Ecole nationale vétérinaire d'Alfort, UMR BIPAR, ENVA, ANSES, UPEC, USC INRA, 23 Rue du Gl de Gaulle-94703 Maisons-Alfort, France.
authors.forename: Héloïse
authors.initials: H
authors.lastname: Pilet
authors.forename: Nathalie
authors.initials: N
authors.lastname: Vachiéry
authors.forename: Moez
authors.initials: M
authors.lastname: Berrich
authors.forename: Rim
authors.initials: R
authors.lastname: Bouchouicha
authors.forename: Benoît
authors.initials: B
authors.lastname: Durand
authors.forename: Ludovic
authors.initials: L
authors.lastname: Pruneau
authors.forename: Valérie
authors.initials: V
authors.lastname: Pinarello
authors.forename: Angélique
authors.initials: A
authors.lastname: Saldana
authors.forename: Catherine
authors.initials: C
authors.lastname: Carasco-Lacombe
authors.forename: Thierry
authors.initials: T
authors.lastname: Lefrançois
authors.forename: Damien F
authors.initials: DF
authors.lastname: Meyer
authors.forename: Dominique
authors.initials: D
authors.lastname: Martinez
authors.forename: Henri-Jean
authors.initials: HJ
authors.lastname: Boulouis
authors.forename: Nadia
authors.initials: N
authors.lastname: Haddad
mesh_terms: D000818:Animals; D002417:Cattle; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics; D016995:Ehrlichia ruminantium / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D006041:Goats; D006357:Heartwater Disease / Q000382:microbiology; D026863:Ixodidae / Q000382:microbiology; D018598:Minisatellite Repeats; D058889:Molecular Typing / Q000379:methods*; D011110:Polymorphism, Genetic; D015203:Reproducibility of Results
chemical_list: D004269:DNA, Bacterial