title: Development of a live, attenuated, potential vaccine strain of R. equi expressing vapA and the virR operon, and virulence assessment in the mouse.
abstract: Pneumonia caused by Rhodococcus equi remains a significant problem in foals. The objective of this study was to develop a safe and efficacious attenuated strain of R. equi for eventual use in oral immunization of foals. The approach involved expression of vapA in a live, virulence plasmid-negative, strain of R. equi (strain 103-). PCR-amplified fragments of the vapA gene, with and without the upstream genes virR, orf5, vapH, orf7 and orf8 (orf4-8), were cloned into a shuttle vector pNBV1. These plasmids, named pAW48A and pAWVapA respectively, were electroporated into strain 103-. The presence of the recombinant vectors in the attenuated strain (103-) and the integrity of the inserted genes were confirmed, and both constructs expressed VapA. The virulence of the two strains was compared to that of wild type R. equi 103+ and negative controls by their intravenous inoculation into mice, followed by examination of liver clearance 4 days later. Mice inoculated with R. equi 103-, 103-/pAWVapA and 103-/pNBV1 completely cleared infection, whereas strain 103-/pAW48A persisted in 47% of mice.
authors.affiliation: Department of Clinical Studies, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
authors.forename: Ashley E
authors.initials: AE
authors.lastname: Whitehead
authors.forename: Valeria R
authors.initials: VR
authors.lastname: Parreira
authors.forename: Joanne
authors.initials: J
authors.lastname: Hewson
authors.forename: Johanna L
authors.initials: JL
authors.lastname: Watson
authors.forename: John F
authors.initials: JF
authors.lastname: Prescott
mesh_terms: D000193:Actinomycetales Infections / Q000276:immunology / Q000517:prevention & control / Q000662:veterinary*; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology; D001428:Bacterial Vaccines / Q000235:genetics* / Q000276:immunology; D015153:Blotting, Western / Q000662:veterinary; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D005260:Female; D006734:Horse Diseases / Q000276:immunology / Q000382:microbiology / Q000517:prevention & control; D006736:Horses / Q000276:immunology; D051379:Mice; D009876:Operon / Q000235:genetics / Q000276:immunology; D016133:Polymerase Chain Reaction / Q000662:veterinary; D016929:Rhodococcus equi / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity; D014157:Transcription Factors / Q000235:genetics* / Q000276:immunology; D014613:Vaccines, Attenuated / Q000235:genetics / Q000276:immunology; D014774:Virulence / Q000235:genetics / Q000276:immunology
chemical_list: D001426:Bacterial Proteins; D001428:Bacterial Vaccines; D014157:Transcription Factors; D014613:Vaccines, Attenuated; C541879:VapA protein, Rhodococcus equi; C523274:virR protein, Rhodococcus equi