Loss of penicillin tolerance by inactivating the carbon catabolite repression determinant CcpA in Streptococcus gordonii

Objectives Antibiotic tolerance is a phenomenon allowing bacteria to withstand drug-induced killing. Here, we studied a penicillin-tolerant mutant of Streptococcus gordonii (Tol1), which was shown to be deregulated in the expression of the arginine deiminase operon (arc). arc was not directly responsible for tolerance, but is controlled by the global regulator CcpA. Therefore, we sought whether CcpA might be implicated in tolerance. Methods The ccpA gene was characterized and subsequently inactivated by PCR ligation mutagenesis in both the susceptible wild-type (WT) and Tol1. The minimal inhibitory concentration and time-kill curves for the strains were determined and the outcome of penicillin treatment in experimental endocarditis assessed. Results ccpA sequence and expression were similar between the WT and Tol1 strains. In killing assays, the WT lost 3.5 ± 0.6 and 5.3 ± 0.6 log10 cfu/mL and Tol1 lost 0.4 ± 0.2 and 1.4 ± 0.9 log10 cfu/mL after 24 and 48 h of penicillin exposure, respectively. Deletion of ccpA almost totally restored Tol1 kill susceptibility (loss of 2.5 ± 0.7 and 4.9 ± 0.7 log10 cfu/mL at the same endpoints). In experimental endocarditis, penicillin treatment induced a significant reduction in vegetation bacterial densities between Tol1 (4.1 log10 cfu/g) and Tol1ΔccpA (2.4 log10 cfu/g). Restitution of ccpA re-established the tolerant phenotype both in vitro and in vivo. Conclusions CcpA, a global regulator of the carbon catabolite repression system, is implicated in penicillin tolerance both in vitro and in vivo. This links antibiotic survival to bacterial sugar metabolism. However, since ccpA sequence and expression were similar between the WT and Tol1 strains, other factors are probably involved in toleranc

2014, dos años de datos para grafica

El articulo presenta los datos y logros de la edición de grafica durante el año 2014. Se aportan datos objetivos del número de registros, del número de lectores y también se relaciona los títulos de los artículos publicados. Además, se informa de la frecuencia de las palabras clave utilizadas en los artículos, la metodologia utilitzada por los autortes y la orientación en investigación en base a si se formulan preguntes de investigación o bien hipòtesis. Todos estos datos son comparados con los del año anterior lo que permite compararlos y visualitzar major los cambios. Por otra parte, se hace especial mención a la indexación de la revista en latindex y en el directorio DOAJ de revistes editades en Open Accés. Al complir con 30 de los 36 criterios de Latindex se hap odido quedar listada en diferentes bases de datos y catálogos como son ISOC, Dialnet.The article presents the data of grafica during the year 2014. The text explains the Objective data, readers and it also relates the titles of articles published. Additionally, it informs about the keywords and the methodology used by the authors and the typology of approaches: questions research or hypotheses. All these details are compared with previous year, which permits to obtain better point of view about grafica. The text relate to the indexing of grafica in Latindex, in the Directory of Open Access Journals (DOAJ), in ISOC and also in Dialnet.L'article presenta les dades i èxits de l'edició de gràfica durant l'any 2014. S'aporten dades objectives del nombre de registres, del nombre de lectors i també es relaciona els títols dels articles publicats. A més, s'informa de la freqüència de les paraules clau utilitzades en els articles, la metodologia utilitzada pels autors i l'orientació en investigació basant-se si es formulen preguntes de recerca o bé hipòtesis. Totes aquestes dades són comparats amb els de l'any anterior el que permet visualitzar millor els canvis. D'altra banda, es fa especial esment a la indexació de la revista en Latindex i en el directori DOAJ de revistes editades en Open Accés. En acomplir amb 30 dels 36 criteris de Latindex ha quedat llistada en diferents bases de dades i catàlegs com ISOC, Dialnet

In vitro activities of tigecycline combined with other antimicrobials against multiresistant Gram-positive and Gram-negative pathogens

Objectives To test the activity of tigecycline combined with 16 antimicrobials in vitro against 22 Gram-positive and 55 Gram-negative clinical isolates. Methods Antibiotic interactions were determined by chequerboard and time-kill methods. Results By chequerboard, of 891 organism-drug interactions tested, 97 (11%) were synergistic, 793 (89%) were indifferent and 1 (0.1%) was antagonistic. Among Gram-positive pathogens, most synergisms occurred against Enterococcus spp. (7/11 isolates) with the tigecycline/rifampicin combination. No antagonism was detected. Among Gram-negative organisms, synergism was observed mainly with trimethoprim/sulfamethoxazole against Serratia marcescens (5/5 isolates), Proteus spp. (2/5) and Stenotrophomonas maltophilia (2/5), with aztreonam against S. maltophilia (3/5), with cefepime and imipenem against Enterobacter cloacae (3/5), with ceftazidime against Morganella morganii (3/5), and with ceftriaxone against Klebsiella pneumoniae (3/5). The only case of antagonism occurred against one S. marcescens with the tigecycline/imipenem combination. Selected time-kill assays confirmed the bacteriostatic interactions observed by the chequerboard method. Moreover, they revealed a bactericidal synergism of tigecycline with piperacillin/tazobactam against one penicillin-resistant Streptococcus pneumoniae and with amikacin against Proteus vulgaris. Conclusions Combinations of tigecycline with other antimicrobials produce primarily an indifferent response. Specific synergisms, especially against enterococci and problematic Gram-negative isolates, might be worth investigating in in vitro models and/or in animal models simulating the human environmen

New concepts in the pathophysiology of infective endocarditis

Endocarditis pathogens colonize valves with pre-existing sterile vegetations or valves with minimal endothelial lesions. Inflamed endothelia produce cytokines, integrins, and tissue factor, which in turn attract fibronectin, monocytes, and platelets. Bacteria attaching to such structures further activate the cascade, becoming embedded and protected from host defenses. Staphylococcus aureus also actively invade the endothelium, causing apoptosis and endothelial damage. Knowledge of this interplay identifies host factors as potential therapeutic targets. Blocking infection by modulating host factors might be opportune because host factors are conserved. In contrast, interfering with bacterial virulence factors might be more complicated because they vary among different bacteri

Efficacy of daptomycin in the treatment of experimental endocarditis due to susceptible and multidrug-resistant enterococci

Objectives: Daptomycin was tested in vitro and in rats with experimental endocarditis against the ampicillin-susceptible and vancomycin-susceptible Enterococcus faecalis JH2-2, the vancomycin-resistant (VanA type) mutant of strain JH2-2 (strain JH2-2/pIP819), and the ampicillin-resistant and vancomycin-resistant (VanB type) Enterococcus faecium D366. Methods: Rats with catheter-induced aortic vegetations were treated with doses simulating intravenously kinetics in humans of daptomycin (6 mg/kg every 24 h), amoxicillin (2 g every 6 h), vancomycin (1 g every 12 h) or teicoplanin (12 mg/kg every 12 h). Treatment was started 16 h post-inoculation and continued for 2 days. Results: MICs of daptomycin were 1, 1 and 2 mg/L, respectively, for strains JH2-2, JH2-2/pIP819 and D366. In time-kill studies, daptomycin showed rapid (within 2 h) bactericidal activity against all strains. Daptomycin was highly bound to rat serum proteins (89%). In the presence of 50% rat serum, simulating free concentrations, daptomycin killing was maintained but delayed (6-24 h). In vivo, daptomycin treatment resulted in 10 of 12 (83%), 9 of 11 (82%) and 11 of 12 (91%) culture-negative vegetations in rats infected with strains JH2-2, JH2-2/pIP819 and D366, respectively (P < 0.001 compared to controls). Daptomycin efficacy was comparable to that of amoxicillin and vancomycin for susceptible isolates. Daptomycin, however, was significantly (P < 0.05) more effective than teicoplanin against the glycopeptide-susceptible strain JH2-2 and superior to all comparators against resistant isolates. Conclusions: These results support the use of the newly proposed daptomycin dose of 6 mg/kg every 24 h for treatment of enterococcal infections in human

The Impact of Penicillinase on Cefamandole Treatment and Prophylaxis of Experimental Endocarditis Due to Methicillin-Resistant Staphylococcus aureus

β-lactams active against methicillin-resistant Staphylococcus aureus (MRSA) must resist penicillinase hydrolysis and bind penicillin-binding protein 2A (PBP 2A). Cefamandole might share these properties. When tested against 2 isogenic pairs of MRSA that produced or did not produce penicillinase, MICs of cefamandole (8-32 mg/L) were not affected by penicillinase, and cefamandole had a ⩾40 times greater PBP 2A affinity than did methicillin. In rats, constant serum levels of 100 mg/L cefamandole successfully treated experimental endocarditis due to penicillinase-negative isolates but failed against penicillinase-producing organisms. This suggested that penicillinase produced in infected vegetations might hydrolyze the drug. Indeed, cefamandole was slowly degraded by penicillinase in vitro. Moreover, its efficacy was restored by combination with sulbactam in vivo. Cefamandole also uniformly prevented MRSA endocarditis in prophylaxis experiments, a setting in which bacteria were not yet clustered in the vegetations. Thus, while cefamandole treatment was limited by penicillinase, the drug was still successful for prophylaxis of experimental MRSA endocarditi

Comparative Genomics Analysis of Streptococcus tigurinus Strains Identifies Genetic Elements Specifically and Uniquely Present in Highly Virulent Strains.

Streptococcus tigurinus is responsible for severe invasive infections such as infective endocarditis, spondylodiscitis and meningitis. As described, S. tigurinus isolates AZ_3aT and AZ_14 were highly virulent (HV phenotype) in an experimental model of infective endocarditis and showed enhanced adherence and invasion of human endothelial cells when compared to low virulent S. tigurinus isolate AZ_8 (LV phenotype). Here, we sought whether genetic determinants could explain the higher virulence of AZ_3aT and AZ_14 isolates. Several genetic determinants specific to the HV strains were identified through extensive comparative genomics amongst which some were thought to be highly relevant for the observed HV phenotype. These included i) an iron uptake and metabolism operon, ii) an ascorbate assimilation operon, iii) a newly acquired PI-2-like pilus islets described for the first time in S. tigurinus, iv) a hyaluronate metabolism operon, v) an Entner-Doudoroff pathway of carbohydrates metabolism, and vi) an alternate pathways for indole biosynthesis. We believe that the identified genomic features could largely explain the phenotype of high infectivity of the two HV S. tigurinus strains. Indeed, these features include determinants that could be involved at different stages of the disease such as survival of S. tigurinus in blood (iron uptake and ascorbate metabolism operons), initial attachment of bacterial pathogen to the damaged cardiac tissue and/or vegetation that formed on site (PI-2-like pilus islets), tissue invasion (hyaluronate operon and Entner-Doudoroff pathway) and regulation of pathogenicity (indole biosynthesis pathway)

Primer simposi grafica

Primer simposi grafica, editorial, 201

Importance of Genotypic and Phenotypic Tolerance in the Treatment of Experimental Endocarditis Due to Streptococcus gordonii

Genotypic and phenotypic tolerance was studied in penicillin treatment of experimental endocarditis due to nontolerant and tolerant Streptococcus gordonii and to their backcross transformants. The organisms were matched for in vitro and in vivo growth rates. Rats with aortic endocarditis were treated for 3 or 5 days, starting 12, 24, or 48 h after inoculation. When started at 12 h, during fast intravegetation growth, 3 days of treatment cured 80% of the nontolerant parent compared with <30% of the tolerant derivative (P < .005). When started at 24 or 48 h and if intravegetation growth had reached a plateau, 3 days of treatment failed against both bacteria. However, a significant difference between the 2 organisms was restored when treatment was extended to 5 days. Thus, genotypic tolerance conferred a survival advantage in both fast- and slow-growing bacteria, demonstrating that the in vitro-defined tolerant phenotype also carried the risk of treatment failure in viv