An iron-acquisition-deficient mutant of Corynebacterium pseudotuberculosis efficiently protects mice against challenge

Abstract

Caseous lymphadenitis (CLA) is a chronic disease that affects sheep and goats worldwide, and its etiological agent is Corynebacterium pseudotuberculosis. Despite the economic losses caused by CLA, there is little information about the molecular mechanisms of bacterial pathogenesis, and current immune prophylaxis against infection has been unable to reduce the incidence of CLA in goats. Recently, 21 different mutant strains of C. pseudotuberculosis were identified by random mutagenesis. In this study, these previously generated mutants were used in mice vaccination trials to develop new immunogens against CLA. Based on this analysis, CZ171053, an iron-acquisition-deficient mutant strain, was selected. After challenge with a virulent strain, 80% of the animals that were immunized with the CZ171053 strain survived. Furthermore, this vaccination elicited both humoral and cellular responses. Intracellular survival of the bacterium was determined using murine J774 cells; in this assay, the CZ171053 had reduced intracellular viability. Because iron acquisition in intracellular bacteria is considered one of their most important virulence factors during infection, these results demonstrate the immunogenic potential of this mutant against CLA.

Corynebacterium pseudotuberculosiscp09 mutant and cp40 recombinant protein partially protect mice against caseous lymphadenitis

“…The protection in animals immunised only with CP09 mutant strain resulted in 50% survival after virulent challenge, which is in agreement with previously published results [ 7 ]. Ribeiro et al [ 12 ] described a high protection rate (80%) in a murine model when using an iron acquisition-deficient mutant obtained with the same methodology employed to develop the CP09 mutant, showing that C. pseudotuberculosis live attenuated mutants can be promising tools for protection against caseous lymphadenitis.…”

“…Kanamycin (kanamycin sulphate 25 μg/mL; Sigma-Aldrich, USA) was added to the mutant growth media. T1 and MIC-6 strains were isolated from caseous lesions of goats and have previously been employed in vaccination trials using goats and mice [ 11 , 12 ].…”

Evaluation of the aroA mutant of Corynebacterium pseudotuberculosis in cellular and murine models

“…Thus, the mutant failed to confer at least 80 % protection, which is desirable for an immunogen, in the vaccinated mice 19 . Previously, an immunogen against caseous lymphadenitis was developed by deleting the pld gene of C. pseudotuberculosis, which is related to the production of phospholipase D, an exotoxin that is considered the major bacterial virulence factor 25 .…”

“…To lyse the erythrocytes, the pellet was re-suspended in 1 ml of 0.17 M ammonium chloride per spleen for 5 min at 4 °C. Three more washes with DMEM supplemented with 10 % FBS, 2 mM L-glutamine, and 0.1 mM non-essential amino acids (Gibco) were performed 19 . Subsequently, 5 x 10 6 lymphocyte cells were extracted from the mutant-vaccinated and control mice spleens and placed in 24-well polystyrene plates.…”

Identification of new Corynebacterium pseudotuberculosis antigens by immunoscreening of gene expression library

“…A recent study used an attenuated strain of C. pseudotuberculosis in which the ciuA gene (gene involved in the transport iron citrate to the bacterium metabolism) was knocked out. This vaccine induced both cellular and humoral responses after challenge with the wild type strain [ 15 ].…”