RESUMO
Rhodococcus equi preferentially infects macrophages causing pyogranulomatous pneumonia in young foals. Both the vapA and rhbC genes are up-regulated in an iron (Fe)-deprived environment, such as that found within macrophages. Chloroquine (CQ) is a drug widely used against malaria that suppresses the intracellular availability of Fe in eukaryotic cells. The main objective of this study was to evaluate the ability of CQ to inhibit replication of virulent R. equi within murine (J774A.1) and foal alveolar macrophages (AMs) and to verify whether the mechanism of inhibition could be Fe-deprivation-dependent. CQ effect on R. equi extracellular survival and toxicity to J774A.1 were evaluated. R. equi survival within J774A.1 and foal AMs was evaluated under CQ (10 and 20µM), bovine saturated transferrin (bHTF), and bovine unsaturated transferrin (bATF) exposure. To explore the action mechanism of CQ, the superoxide anion production, the lysozyme activity, as well as the relative mRNA expression of vapA and rhbC were examined. CQ at≤20µM had no effect on R. equi extracellular multiplication and J774A.1 viability. Exposure to CQ significantly and markedly reduced survival of R. equi within J774A.1 and foal AMs. Treatment with bHTF did not reverse CQ effect on R. equi. Exposure to CQ did not affected superoxide anion production or lysozyme activity, however vapA and rhbC expression was significantly increased. Our results reinforce the hypothesis that intracellular availability of Fe is required for R. equi survival, and our initial hypothesis that CQ can limit replication of R. equi in J774A.1 and foal AMs, most likely by Fe starvation.
Assuntos
Cloroquina/farmacologia , Macrófagos Alveolares/microbiologia , Rhodococcus equi/efeitos dos fármacos , Animais , Proteínas de Bactérias/genética , Linhagem Celular , Regulação da Expressão Gênica/efeitos dos fármacos , Cavalos , Ferro/metabolismo , Macrófagos Alveolares/citologia , Camundongos , Viabilidade Microbiana/efeitos dos fármacos , Muramidase/metabolismo , Rhodococcus equi/citologiaRESUMO
OBJECTIVE: To compare isolates of Rhodococcus equi on the basis of geographic source and virulence status by use of pulsed-field gel electrophoresis (PFGE). SAMPLE POPULATION: 290 isolates of R equi (218 virulent isolates from foals and 72 avirulent isolates from feces, soil, and respiratory tract samples) obtained between 1985 and 2000 from horses and horse farms from 4 countries. PROCEDURE: DNA from isolates was digested with the restriction enzyme Asel and tested by use of PFGE. Products were analyzed for similarities in banding patterns by use of dendrograms. A similarity matrix was constructed for isolates, and the matrix was tested for nonrandom distributions of similarity values with respect to groupings of interest. RESULTS: There was little grouping of isolates on the basis of country, virulence status, or region within Texas. Isolates of R equi were generally < 80% similar, as determined by use of PFGE. Isolates from the same farm generally were rarely of the same strain. CONCLUSIONS AND CLINICAL RELEVANCE: Considerable chromosomal variability exists among isolates of R equiobtained from the same farm, sites withinTexas, or among countries from various continents. Only rarely will it be possible to link infections to a given site or region on the basis of analysis of isolates by use of PFGE of chromosomal DNA.