RESUMEN
Ten isolates of the recently delineated genospecies Borrelia spielmanii were tested against antimicrobial agents used to treat Lyme disease and compared to eight isolates of the other three human-pathogenic borrelial genospecies. Despite some small but significant differences in four out of eight antibiotic agents, the susceptibility pattern of B. spielmanii mainly parallels that of the other known human-pathogenic members of the B. burgdorferi sensu lato complex.
Asunto(s)
Antiinfecciosos/uso terapéutico , Grupo Borrelia Burgdorferi/clasificación , Grupo Borrelia Burgdorferi/genética , Borrelia burgdorferi/genética , Enfermedad de Lyme/tratamiento farmacológico , Grupo Borrelia Burgdorferi/aislamiento & purificación , Guías como Asunto , Humanos , Enfermedad de Lyme/genética , Pruebas de Sensibilidad Microbiana , Factores de TiempoRESUMEN
Borrelia burgdorferi exploits multiple strategies to evade host immune responses. One central immune escape mechanism is the inactivation of the host complement attack by acquisition host complement regulators FHL-1 and factor H via complement regulator-acquiring surface proteins (BbCRASPs). The BbCRASP-1 protein is the first bacterial factor H/FHL-1-binding protein for which the atomic structure has been solved. Previously, 3 regions including the C terminus were identified as putative contact sites for the two complement regulators by the pepspot analysis. Based on the crystallographic structure an in vitro mutagenesis approach was conducted to identify amino acid residues which are relevant for FHL-1 and factor H binding by exchanging single or multiple residues in region 1 and the C-terminally located region 3. Single changes at 4 positions in region 1 either reduced (Lys136, Lys141, Glu147) or completely eliminated (Leu146) binding of both complement regulators. Substitutions clustered within the C-terminal region decreased (Glu234, Lys238, Tyr239, Lys241, Asp244, Thr245) or abolished binding (Lys240, Asp242, Leu246) of both complement regulators. Mapping the mutations onto the atomic structure of BbCRASP-1 reveals that, in contrast to earlier assumption, the C-terminal mutations act indirectly on FHL-1 and factor H binding, whilst the region 1 mutations map the site of direct complement regulator interaction. The elucidation of BbCRASP-1 structure - function may allow development of novel therapeutic strategies against Lyme disease.
Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Borrelia burgdorferi/química , Factor H de Complemento/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Mapeo de Interacción de Proteínas , Sustitución de Aminoácidos/genética , Proteínas Bacterianas/genética , Borrelia burgdorferi/genética , Proteínas Inactivadoras del Complemento C3b , Análisis Mutacional de ADN , Proteínas de la Membrana/genética , Modelos Moleculares , Mutagénesis Sitio-Dirigida , Unión Proteica , Estructura Terciaria de ProteínaRESUMEN
Tigecycline is a new glycylcycline that has recently been revealed to be very effective in vitro against a variety of Gram-negative and Gram-positive bacteria including multi-drug resistant microorganisms. Using a standardized microdilution susceptibility testing method, we determined the minimal inhibitory concentrations (MICs) and the minimal bactericidal concentrations (MBCs) of tigecycline, in parallel with doxycycline, tetracycline, and other antibiotic agents relevant for Lyme borreliosis treatment such as ceftriaxone and cefotaxime. The activity of all agents against 16 different Borrelia isolates belonging to all borrelial genospecies known to be pathogenic for humans was investigated and analyzed under standardized conditions. The overall rank order of MIC(90)s was tigecycline (≤0.016 mg/L) > ceftriaxone (0.03 mg/L) > cefotaxime (≤0.125 mg/L) > doxycycline (0.25 mg/L) > tetracycline (0.25 mg/L). The rank order of MBC(90)s was tigecycline (0.5 mg/L) > ceftriaxone (2 mg/L) > tetracycline (16 mg/L) > doxycycline (16 mg/L) > cefotaxime (>16 mg/L). High in vitro activity of the new glycylcycline against Borrelia was further substantiated by time-kill experiments performed with B. afzelii isolate EB1. Parallel testing of tigecycline and ceftriaxone demonstrated a bacteriostatic effect for 0.016 mg/L of tigecycline and for 0.03 mg/L for ceftriaxone after 72 h of incubation. Moreover, tigecycline was bactericidal at a concentration of 0.25 mg/L showing a >3 log(10) unit reduction of the initial inoculum, whereas for ceftriaxone a concentration of 2 mg/L was needed.