Wide Variety Of Gram-positive Bacteria Research Articles

Determining optimal dosing regimen of oral administration of dicloxacillin using Monte Carlo simulation.

BackgroundDicloxacillin, a semisynthetic isoxazolyl penicillin, exhibits antimicrobial activity against a wide variety of Gram-positive bacteria, as well as stability against penicillinases and low level of toxicity. The objective of this study was to obtain optimal dosing regimen of oral administration of dicloxacillin by analyzing the pharmacokinetic (PK) index in healthy volunteers and in vitro antibacterial activity by using Monte Carlo simulation.Materials and methodsA total of 867 clinical isolates from community-onset infections were collected from 31 secondary hospitals in People’s Republic of China. The minimum inhibitory concentration (MIC) values of dicloxacillin were determined by the agar dilution method. Based on the MICs and the PK parameters of different dosage regimens, Monte Carlo simulation was performed to simulate the PK/pharmacodynamic indices of 250 mg once-daily (qd), 500 mg qd, 1,000 mg qd, 2,000 mg qd, 250 mg every 6 hours (q6h), and 500 mg q6h, respectively. The probability of target attainment was estimated at each MIC value, and the cumulative fraction of response (CFR) was calculated to evaluate the efficacy of these regimens.ResultsDicloxacillin showed poor antibacterial activity against Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae. Resistance to dicloxacillin was observed in 7.5% of coagulase-negative Staphylococcus (CNS) isolates and 9.2% of other Streptococcus isolates, whereas 1.5% of methicillin-sensitive Staphylococcus aureus (MSSA) was resistant to dicloxacillin. Multiple-dose regimens could obtain higher CFR than single-dose regimens against H. influenza and S. pneumoniae. However, all dosing regimens against MSSA achieved CFR ≥$90%. Meanwhile, dosing regimen of 2,000 mg qd, 250 mg q6h, and 500 mg q6h could achieve >90% of CFR for CNS. For other Streptococcus isolates, multiple-dose regimens achieved CFR ≥90%.ConclusionDicloxacillin has a significant antibacterial activity against MSSA, CNS, and other Streptococcus isolates. The simulation results suggest that dicloxacillin 250 mg q6h and 500 mg q6h dosing regimens may be recommended for clinical applications, especially for community-onset infections.

Safety and pharmacokinetics of dicloxacillin in healthy Chinese volunteers following single and multiple oral doses.

BackgroundDicloxacillin, a semisynthetic isoxazolyl penicillin antibiotic, has antimicrobial activity against a wide variety of gram-positive bacteria including Staphylococcus aureus, Streptococcus pyogenes, Streptococcus pneumonia, Streptococcus epidermidis, Streptococcus viridans, Streptococcus agalactiae, and Neisseria meningitidis. The objective of this study was to evaluate the safety and pharmacokinetic profile of dicloxacillin after single and multiple oral dose in healthy Chinese volunteers.MethodsA single-center, open-label, randomized, two-phase study was conducted in 16 subjects. In the single-dose phase, subjects were randomly assigned to receive single doses of 0.25, 0.5, 1.0, and 2.0 g of dicloxacillin sodium capsule in a 4-way crossover design with a 5-day washout period between administrations. In the multiple-dose phase, subjects were assigned to receive 0.25 or 0.5 g every 6 hours for 3 days in a 2-way crossover design. Plasma and urine pharmacokinetic samples were assayed by a validated high-performance liquid chromatography-tandem mass spectrometry method. Pharmacokinetic parameters were calculated and analyzed statistically. Safety assessments were conducted throughout the study.ResultsFollowing a single oral dose of 0.25–2.0 g dicloxacillin sodium, the maximum plasma drug concentration (Cmax) and the corresponding values for the area under the concentration– time curve from 0 to 10 hours (AUC0–10 h) increased in a dose-proportional manner. The mean elimination half-life (t1/2) was in the range of 1.38–1.71 hours. Dicloxacillin was excreted in its unchanged form via the kidney, with no tendency of accumulation, and varied from 38.65% to 50.10%. No appreciable accumulation of drug occurred with multiple oral doses of dicloxacillin. No serious adverse events were reported. Adverse events were generally mild.ConclusionDicloxacillin was safe and well tolerated in the volunteers and displayed linear increases in the Cmax and AUC0–10 h values.

Transcriptional attenuation controls macrolide inducible efflux and resistance in Streptococcus pneumoniae and in other Gram-positive bacteria containing mef/mel(msr(D)) elements.

Macrolide resistance, emerging in Streptococcus pneumoniae and other Gram-positive bacteria, is increasingly due to efflux pumps encoded by mef/mel(msr) operons found on discrete mobile genetic elements. The regulation of mef/mel(msr) in these elements is not well understood. We identified the mef(E)/mel transcriptional start, localized the mef(E)/mel promoter, and demonstrated attenuation of transcription as a mechanism of regulation of macrolide-inducible mef-mediated macrolide resistance in S. pneumoniae. The mef(E)/mel transcriptional start site was a guanine 327 bp upstream of mef(E). Consensus pneumococcal promoter -10 (5′-TATACT-3′) and -35 (5′-TTGAAC-3′) boxes separated by 17 bp were identified 7 bp upstream of the start site. Analysis of the predicted secondary structure of the 327 5’ region identified four pairs of inverted repeats R1-R8 predicted to fold into stem-loops, a small leader peptide [MTASMRLR, (Mef(E)L)] required for macrolide induction and a Rho-independent transcription terminator. RNA-seq analyses provided confirmation of transcriptional attenuation. In addition, expression of mef(E)L was also influenced by mef(E)L-dependent mRNA stability. The regulatory region 5’ of mef(E) was highly conserved in other mef/mel(msr)-containing elements including Tn1207.1 and the 5612IQ complex in pneumococci and Tn1207.3 in Group A streptococci, indicating a regulatory mechanism common to a wide variety of Gram-positive bacteria containing mef/mel(msr) elements.

Antibacterial activity of a cell wall hydrolase from Lactobacillus paracasei NRRL B-50314 produced by recombinant Bacillus megaterium.

The cell-free supernatant (CFS) from Lactobacillus paracasei NRRL B-50314 culture has been previously reported as containing antibacterial activity against a wide variety of Gram-positive bacteria. The CFS protein gel slice corresponding to antibacterial activities was subjected to trypsin digestion and ion trap MASS (Gel/LC-MS/MS) analysis. BlastP search of the resulted IQAVISIAEQQIGKP sequence led to a hypothetical cell-wall associated hydrolase (designated as CWH here) from Lactobacillus paracasei ATCC 25302. Further analyses of CWH revealed that the IQAVISIAEQQIGKP belongs to a highly conserved region of the NlpC/P60 superfamily. The L. paracasei NRRL B-50314 CWH gene, cloned in pStrepHIS1525CWH477, was introduced into Bacillus megaterium MS 941. The production of CWH477 protein was induced by xylose. The CWH477 protein was purified by using NiNTA column, and elution fraction E2 showed highest antibacterial activity. This study and bioinformatics analyses suggested that the antibacterial activity of CWH could originate from its cell wall degrading enzymatic function.

A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria

The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria.

Novel Antibacterial Polypeptide Laparaxin Produced By Lactobacillus Paracasei Strain NRRL B-50314 Via Fermentation

This study reports the production and characterization of a novel antibacterial polypeptide, designated laparaxin, which is secreted by Lactobacillus paracasei NRRL B-50314. Crude laparaxin has antibacterial activity against a wide variety of Gram-positive bacteria, including: lactic acid bacteria (Lactococcus lactis and Lactobacillus buchneri), food-borne pathogens (Listeria monocytogenes), gastrointestinal pathogens (Enterococcus faecalis), and opportunistic pathogens (Staphylococcus aureus methicillin-sensitive (MSSA) and methicillin resistant (MRSA) strains, a hetero-vancomycin-intermediate methicillin resistant strain (Hetero VISA also MRSA) MM66, and homogeneous vancomycin intermediate (Homo VISA). Using L. lactis as an indicator strain, the inhibitory activity of crude laparaxin was detected originally in early log phase, and the activity maximizes at the early stationary phase and remains stable after prolonged incubation. Laparaxin activity is stable after 30 min of incubation at 94°C. Higher concentrations of inhibitory activity are produced when glucose, fructose and sucrose are used as carbon-sources in growth media. Crude laparaxin has potential applications in food and feed industries, as well as in clinical and veterinary medicine.

Oritavancin Population Pharmacokinetics in Healthy Subjects and Patients with Complicated Skin and Skin Structure Infections or Bacteremia

Oritavancin is a novel glycopeptide antimicrobial agent with potent in vitro activity against a wide variety of gram-positive bacteria, including multidrug-resistant strains of staphylococci and enterococci. A population pharmacokinetic model was developed to describe the disposition of oritavancin with data from a pooled population of phase 1 healthy subjects and phase 2 and 3 patients with complicated skin and skin structure infections or Staphylococcus aureus bacteremia. In addition, the potential influence of factors such as the subject's age, gender, and clinical laboratory measures on oritavancin disposition was evaluated. Oritavancin was administered as both single- and multiple-dose intravenous (i.v.) infusions in fixed doses ranging from 100 to 800 mg or weight-based doses ranging from 0.02 to 10 mg/kg of body weight, with infusion durations ranging from 0.13 to 6.5 h across all studies. The most robust fit to the data (n = 6,290 oritavancin plasma concentrations from 560 subjects) was obtained using a three-compartment model with zero-order i.v. infusion and first-order elimination. The model was parameterized using total clearance (CL), volume of central compartment (Vc), distributional clearances from the central to both the first and second peripheral compartments, and volumes of distribution for both the first and second peripheral compartments. Weight and study phase (phase 1 versus phase 2/3) were identified as significant predictors of the interindividual variability in CL, while body surface area and age were significant for Vc. These results suggest that dose modification may be warranted in patients weighing >110 kg. However, the mild nature of the observed relationships for Vc suggest that dosing adjustments are not necessary for elderly patients.

A type IV-secretion-like system is required for conjugative DNA transport of broad-host-range plasmid pIP501 in gram-positive bacteria.

Plasmid pIP501 has a very broad host range for conjugative transfer among a wide variety of gram-positive bacteria and gram-negative Escherichia coli. Functionality of the pIP501 transfer (tra) genes in E. coli was proven by pIP501 retrotransfer to Enterococcus faecalis (B. Kurenbach, C. Bohn, J. Prabhu, M. Abudukerim, U. Szewzyk, and E. Grohmann, Plasmid 50:86-93, 2003). The 15 pIP501 tra genes are organized in a single operon (B. Kurenbach, J. Kopeć, M. Mägdefrau, K. Andreas, W. Keller, C. Bohn, M. Y. Abajy, and E. Grohmann, Microbiology 152:637-645, 2006). The pIP501 tra operon is negatively autoregulated at the transcriptional level by the conjugative DNA relaxase TraA. Three of the 15 pIP501-encoded Tra proteins show significant sequence similarity to the Agrobacterium type IV secretion system proteins VirB1, VirB4, and VirD4. Here we report a comprehensive protein-protein interaction map of all of the pIP501-encoded Tra proteins determined by the yeast two-hybrid assay. Most of the interactions were verified in vitro by isolation of the protein complexes with pull-down assays. In conjunction with known or postulated functions of the pIP501-encoded Tra proteins and computer-assisted prediction of their cellular location, we propose a model for the first type IV-secretion-like system encoded by a conjugative plasmid from gram-positive bacteria.

Daptomycin

Daptomycin (Cubicin) is the first of a new class of antibacterials, the cyclic lipopeptides, and is approved for use in the treatment of complicated skin and soft-tissue or skin-structure infections (hereafter referred to as cSSTI) caused by Gram-positive bacteria and Staphylococcus aureus bacteraemia including right-sided infective endocarditis.Daptomycin has activity in vitro against a wide variety of Gram-positive bacteria, including meticillin-resistant S. aureus (MRSA) and vancomycin-resistant enterococci. When administered as a once-daily intravenous infusion, daptomycin was not inferior to standard parenteral therapy (vancomycin or semi-synthetic penicillins) in terms of clinical and microbiological efficacy in patients with cSSTI or S. aureus bacteraemia with or without infective endocarditis (including MRSA infection) and was well tolerated. With the advantage of once-daily administration and a low potential for drug interactions, daptomycin is a useful addition to the range of parenteral antibacterial agents available for the treatment of patients with cSSTI or S. aureus bacteraemia with or without right-sided infective endocarditis. Efficacy against both meticillin-susceptible S. aureus (MSSA) and MRSA infections makes daptomycin suitable for empirical therapy in patients with serious Gram-positive infections.

SecA2 is distinct from SecA in immunogenic specificity, subcellular distribution and requirement for membrane anchoring inStreptococcus parasanguis

A secA2 gene is present in the genomes of a wide variety of Gram-positive bacteria. In Streptococcus parasanguis, a primary colonizer of the tooth surface, secA2 is involved in the secretion of a small group of proteins including the fimbrial adhesin, Fap1. Although the substrate specificity is different, SecA2 is predicted to be similar to SecA in structure and function based on the homology between these two proteins. In this study, polyclonal antibodies against SecA2 and SecA did not cross-react with each other, indicating that these two proteins possessed distinct immunogenic epitopes. Fractionation analysis demonstrated that SecA2 was not evenly distributed between the cytoplasmic membrane and the cytoplasm as was noted for SecA. SecA2 was associated with the membrane in the wild type and in secA2 mutants with different regions deleted. The subcellular distribution of SecA2 was not dependent on secY2, suggesting that the membrane association is not through SecY2. These data suggested that SecA2 is distinct from SecA in many respects such as substrate specificity, immunogenic specificity, subcellular distribution and requirement for membrane anchoring.

Lytic enzyme, labiase for a broad range of Gram-positive bacteria and its application to analyze functional DNA/RNA

The lytic activity of labiase and achromopeptidase for bacterial DNA/RNA extraction were compared. Rapid lysis of many bacterial strains was observed with labiase followed by SDS treatment. Both labiase and achromopeptidase showed high lytic activity against bacterial strains with the A1α chemotype (e.g., Aerococcus viridans) and the A3α chemotype (e.g., Staphylococcus epidermidis) for cell wall peptidoglycan structures. The lytic activity of labiase was higher than that of achromopeptidase against strains with the A1γ chemotype (e.g., Bacillus subtilis). The activity of labiase was not detrimentally affected with increasing NaCl concentration. Labiase lysates were successfully used for rapid extraction of DNA and RNA, whereas achromopeptidase lysates degraded RNA. The DNA and RNA obtained were successfully used for 16 S rRNA amplification and real-time RT-PCR detection. It is concluded that labiase is useful for rapid lysis of a wide variety of Gram-positive bacteria and can be used for DNA/RNA isolation protocols.

Mechanism of action of the mannopeptimycins, a novel class of glycopeptide antibiotics active against vancomycin-resistant gram-positive bacteria.

The naturally occurring mannopeptimycins (formerly AC98-1 through AC98-5) are a novel class of glycopeptide antibiotics that are active against a wide variety of gram-positive bacteria. The structures of the mannopeptimycins suggested that they might act by targeting cell wall biosynthesis, similar to other known glycopeptide antibiotics; but the fact that the mannopeptimycins retain activity against vancomycin-resistant organisms suggested that they might have a unique mode of action. By using a radioactive mannopeptimycin derivative bearing a photoactivation ligand, it was shown that mannopeptimycins interact with the membrane-bound cell wall precursor lipid II [C(55)-MurNAc-(peptide)-GlcNAc] and that this interaction is different from the binding of other lipid II-binding antibiotics such as vancomycin and mersacidin. The antimicrobial activities of several mannopeptimycin derivatives correlated with their affinities toward lipid II, suggesting that the inhibition of cell wall biosynthesis was primarily through lipid II binding. In addition, it was shown that mannopeptimycins bind to lipoteichoic acid in a rather nonspecific interaction, which might facilitate the accumulation of antibiotic on the bacterial cell surface.

A Pharmacokinetic Evaluation of Concomitant Administration of Linezolid and Aztreonam

Linezolid, a new oxazolidinone antimicrobial agent, has a spectrum of activity encompassing a wide variety of Grampositive bacteria. The purpose of this study was to evaluate the pharmacokinetics of linezolid and aztreonam, an antimicrobial agent with selective activity against Gram-negative bacteria, when given alone and in combination. Healthy subjects were randomized to receive single, 30-minute intravenous infusions of (1) linezolid 375 mg, (2) aztreonam 1000 mg, and (3) linezolid 375 mg plus aztreonam 1000 mg in an open-label, crossover manner. The only statistically significant differences observed with combination treatment relative to each drug alone were an increase in the maximum plasma concentration of linezolid (approximately 18%) and an approximate 7% decrease in the apparent elimination rate of aztreonam, neither of which are expected to be clinically significant. In healthy subjects, the combination of linezolid and aztreonam was safe and well tolerated compared with each agent used alone. Pharmacokinetic data demonstrate that coadministration of linezolid and aztreonam does not alter the disposition of either agent under single-dose conditions. Therefore, it is not expected that a dose alteration of either agent will be necessary in a clinical setting.

Integrative recombination of Lactobacillus delbrueckii bacteriophage mv4: functional analysis of the reaction and structure of the attP site.

The integrase of the temperate bacteriophage mv4 catalyzes site-specific recombination between the phage attP site and the attB site of the host during lysogenization of Lactobaccillus delbrueckii subsp. bulgaricus. The mv4 integrase also functions in a wide variety of gram-positive bacteria and in Escherichia coli. In this report, in vitro and in vivo recombination assays were developed and the integrase was purified in order to study in greater detail the mv4 attP x attB recombination event. In a cell-free extract of E. coli at 42 degrees C, the mv4 integrase promotes efficient in vitro recombination between a supercoiled attP-containing plasmid and a linear attB fragment. The integrase, which was purified to apparent homogeneity, showed no absolute requirement for accessory factors, unlike the majority of the lambda Int family of recombinases. Deletion derivatives of the attP site were constructed and tested for recombination with the attB site in vitro. A 234-bp DNA fragment containing five scattered putative mv4 Int-binding sites was sufficient for function of the attP site. In contrast to the right arm of attP, most of the left arm could be deleted without drastically reducing the recombination efficiency. In vivo in E. coli, mv4 Int catalyzed recombination in trains between attP and attB sites present on two separate plasmids. This property was used to confirm in vivo the results of the deletion analysis of the attP site performed in vitro.

Sporeamicin A, a new macrolide antibiotic. III. Biological properties.

Sporeamicin A is a new erythromycin-type antibiotic isolated from a species of Saccharopolyspora. It was active in vitro against a wide variety of Gram-positive bacteria. In vitro studies indicated that the sporeamicin A was stable in the presence of human serum, although it was bound to serum proteins. Sporeamicin A was effective in the mouse protection test against Staphylococcus aureus, Streptococcus pyogenes and Streptococcus pneumoniae. Sporeamicin A attained higher plasma and tissue levels in the rat than did erythromycin stearate.

Identity of antibiotic P-42-1 elaborated by Actinomyces tumemacerans with kanchanomycin and albofungin.

Antibiotic P-42-1 was isolated from the culture filtrate of Actinomyces tumemacerans strain INMI.P-42, and its detailed physicochemical and biological properties were investigated. The antibiotic was obtained as yellow crystals and its molecular formula was estimated to be G27H24N2O9. It was found to be highly active on a wide variety of gram-positive bacteria and fungi, whereas much less active on gram-negative bacteria. The antibiotic was cytotoxic to HeLa cell cultures and exhibited antitumor activity on EHRLICH ascites tumor in mice. Antibiotic P-42-1 was found to be closely related to antibiotic BA180265 A (kanchanomycin) and albofungin. According to structural studies on albofungin recently conducted by Russian workers, antibiotic P-42-1 proved to be identical with albofungin and kanchanomycin.

SURVEY OF THE BACTERIOCINES OF ENTEROCOCCI.

Brock, Thomas D. (Indiana University, Bloomington), Barbara Peacher, and Deborah Pierson. A survey of the bacteriocines of enterococci. J. Bacteriol. 86:702-707. 1963.-A survey has been made of bacteriocine production by a wide variety of well-characterized strains of group D streptococci. On the basis of spectrums and sensitivity to chloroform, heat, and proteolytic enzymes, five distinct bacteriocines can be defined. Type 1 is produced by all Streptococcus zymogenes (S. faecalis var. zymogenes) strains, is active against a wide variety of gram-positive bacteria, and is also a hemolysin. Type 2 is produced by some S. liquefaciens (S. faecalis var. liquefaciens) strains, and acts on many enterococci as well as on certain other lactic acid bacteria. Type 3 is produced by certain strains of both S. faecalis and S. faecium, and inhibits a wide variety of group D streptococci, but is inactive against all other lactic acid bacteria tested except Leuconostoc citrovorum. Type 4 is produced by certain S. faecium strains and resembles in certain ways the type 3 activity, but differs from it in other ways. Type 5 has been found to be produced by only one proteolytic strain of S. zymogenes, and this bacteriocine has a very narrow spectrum. The strain that produces this bacteriocine also produces type 1 activity. No strain is sensitive to a bacteriocine of the type it produces.