RESUMEN
Proteins belonging to the Gls24 superfamily are involved in survival of pathogenic Gram-positive cocci under oligotrophic conditions and other types of stress, by a still unknown molecular mechanism. In Firmicutes, this superfamily includes three different valine-rich orthologal families (Gls24A, B, C) with different potential interactive partners. Whereas the Streptococcus pneumoniae Δgls24A deletion mutant experienced a general long growth delay, the Δgls24B mutant grew as the parental strain in the semisynthetic AGCH medium but failed to grow in the complex Todd-Hewitt medium. Bovine seroalbumin (BSA) was the component responsible for this phenotype. The effect of BSA on growth was concentration-dependent and was maintained when the protein was proteolyzed but not when heat-denatured, suggesting that BSA dependence was related to oligopeptide supplementation. Global transcriptional analyses of the knockout mutant revealed catabolic derepression and induction of chaperone and oligopeptide transport genes. This mutant also showed increased sensibility to cadmium and high temperature. The Δgls24B mutant behaved as a poor colonizer in the nasopharynx of mice and showed 20-fold competence impairment. Experimental data suggest that Gls24B plays a central role as a sensor of amino acid availability and its connection to sugar catabolism. This metabolic rewiring can be compensated in vitro, at the expenses of external oligopeptide supplementation, but reduce important bacteria skills prior to efficiently address systemic virulence traits. This is an example of how metabolic factors conserved in enterococci, streptococci, and staphylococci can be essential for survival in poor oligopeptide environments prior to infection progression.
Asunto(s)
Aminoácidos Esenciales/metabolismo , Proteínas Bacterianas/metabolismo , Infecciones Neumocócicas/microbiología , Streptococcus pneumoniae/metabolismo , Animales , Proteínas Bacterianas/genética , Medios de Cultivo/metabolismo , Regulación Bacteriana de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos BALB C , Familia de Multigenes , Eliminación de Secuencia , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/crecimiento & desarrolloRESUMEN
OBJECTIVES: Many outer multidomain proteins play fundamental virulent roles in an allele-dependent manner. We aimed to investigate the influence of the outer SP1992 protein, here renamed DiiA (Dimorphic invasion-involved A), in pneumococcal disease. METHODS: The presence and type of diiA allele was screened by PCR in 560 clinical isolates. Isogenic mutants carrying progressive diiA deletions were constructed and checked in mouse models of infection. DiiA binding to human molecules was carried out by surface plasmon resonance. RESULTS: The diiA gene is exclusive of Streptococcus pneumoniae and included in the core genome. DiiA variants contain one or two imperfect repeats (R1 and R2), an unstructured region and a cell-wall anchor domain. Clonal complexes carrying both repeats were associated with invasive disease, while those carrying R2 preferentially caused non-invasive syndromes in patients with underlying risk factors. Mutants lacking both repeats were less efficient in nasopharyngeal colonization and dissemination from lungs. Moreover, the ΔdiiA defective strain suffered a severe impairment in bacterial proliferation in blood. Purified DiiA bound to collagen and lactoferrin with high affinity. CONCLUSIONS: DiiA is a distinctive pneumococcal virulence factor contributing to colonization and long-term invasion in this pathogen.
Asunto(s)
Proteínas Bacterianas/metabolismo , Pared Celular/metabolismo , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/patología , Streptococcus pneumoniae/patogenicidad , Factores de Virulencia/metabolismo , Adulto , Animales , Proteínas Bacterianas/genética , Colágeno/metabolismo , Modelos Animales de Enfermedad , Eliminación de Gen , Humanos , Lactoferrina/metabolismo , Masculino , Ratones Endogámicos C57BL , Reacción en Cadena de la Polimerasa , Unión Proteica , Resonancia por Plasmón de Superficie , Factores de Virulencia/genéticaRESUMEN
BACKGROUND: The major Gram-positive coccoid pathogens cause similar invasive diseases and show high rates of antimicrobial resistance. Uncharacterised proteins shared by these organisms may be involved in virulence or be targets for antimicrobial therapy. RESULTS: Forty four uncharacterised proteins from Streptococcus pneumoniae with homologues in Enterococcus faecalis and/or Staphylococcus aureus were selected for analysis. These proteins showed differences in terms of sequence conservation and number of interacting partners. Twenty eight of these proteins were monodomain proteins and 16 were modular, involving domain combinations and, in many cases, predicted unstructured regions. The genes coding for four of these 44 proteins were essential. Genomic and structural studies showed one of the four essential genes to code for a promising antibacterial target. The strongest impact of gene removal was on monodomain proteins showing high sequence conservation and/or interactions with many other proteins. Eleven out of 40 knockouts (one for each gene) showed growth delay and 10 knockouts presented a chaining phenotype. Five of these chaining mutants showed a lack of putative DNA-binding proteins. This suggest this phenotype results from a loss of overall transcription regulation. Five knockouts showed defective autolysis in response to penicillin and vancomycin, and attenuated virulence in an animal model of sepsis. CONCLUSIONS: Uncharacterised proteins make up a reservoir of polypeptides of different physiological importance and biomedical potential. A promising antibacterial target was identified. Five of the 44 examined proteins seemed to be virulence factors.