title: Characterisation of a Leishmania mexicana knockout lacking guanosine diphosphate-mannose pyrophosphorylase.
abstract: In eukaryotes, the enzyme GDP-mannose pyrophosphorylase (GDP-MP) is essential for the formation of GDP-mannose, the donor of activated mannose for all glycosylation reactions. Unlike other eukaryotes, where deletion of GDP-mannose pyrophosphorylase is lethal, deletion of this gene in Leishmania mexicana has no effect on viability, but leads to the generation of avirulent parasites. In this study, we show that the null mutants have a perturbed morphology and cytokinesis, retarded growth and increased adherence to the substratum where they form large colonies. The null mutants attach avidly to mouse macrophages, but unlike the wild type organisms, they do not bind to the complement receptor 3 and are slow to induce phagocytosis. Once internalised, they localise to the phagolysosome, but in contrast to wild type organisms which transform into the intracellular amastigote and establish in the macrophage, they are cleared by 24 h in culture and by 5 h in vivo. The null mutants are hypersensitive to human but not mouse complement and to temperature and acidic pH. Surprisingly, in view of the lack of several known host-protective antigens, injection of the mutant parasites into BALB/c mice confers significant and long lasting protection against infection, suggesting that these temperature sensitive mutants are an attractive candidate for a live attenuated vaccine.
authors.affiliation: The University of Melbourne, Parkville, Vic. 3052, Australia.
authors.forename: James
authors.initials: J
authors.lastname: Stewart
authors.forename: Joan
authors.initials: J
authors.lastname: Curtis
authors.forename: Timothy P
authors.initials: TP
authors.lastname: Spurck
authors.forename: Thomas
authors.initials: T
authors.lastname: Ilg
authors.forename: Attila
authors.initials: A
authors.lastname: Garami
authors.forename: Tracey
authors.initials: T
authors.lastname: Baldwin
authors.forename: Nathalie
authors.initials: N
authors.lastname: Courret
authors.forename: Geoffrey I
authors.initials: GI
authors.lastname: McFadden
authors.forename: Antony
authors.initials: A
authors.lastname: Davis
authors.forename: Emanuela
authors.initials: E
authors.lastname: Handman
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line; D048749:Cytokinesis / Q000502:physiology; D005260:Female; D006155:Guanosine Diphosphate Mannose / Q000235:genetics; D006790:Host-Parasite Interactions; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007894:Leishmania mexicana / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D016177:Macrophage-1 Antigen / Q000276:immunology; D008264:Macrophages / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D010641:Phenotype; D013696:Temperature; D014611:Vaccination / Q000379:methods; D014774:Virulence
chemical_list: D000906:Antibodies; D016177:Macrophage-1 Antigen; D006155:Guanosine Diphosphate Mannose