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BACKGROUND The oxazolidinone PNU-100480 is superior to linezolid against experimental murine tuberculosis. Two metabolites contribute to but do not fully account for its superiority. This study examined the safety, tolerability, pharmacokinetics, and mycobactericidal activity of single ascending doses of PNU-100480. METHODS Nineteen healthy volunteers received 2 escalating single oral doses (35-1500 mg) of PNU-100480 or placebo. Eight subjects received 4 daily doses of 300 mg of linezolid. Drug concentrations and bactericidal activity against Mycobacterium tuberculosis in whole-blood bactericidal culture were measured. RESULTS All doses were safe and well tolerated. PNU-100480 doses to 1000 mg were well absorbed and showed approximately proportional increases in exposures of parent and metabolites. The geometric mean maximal concentrations of PNU-100480, PNU-101603, and PNU-101244 (sulfoxide and sulfone metabolites) at 1000 mg were 839, 3558, and 54 ng/mL, respectively. The maximal whole-blood bactericidal activity (-0.37 +/- .06 log/day) occurred at combined PNU levels > or =2 times the minimum inhibitory concentration. The observed geometric mean maximal concentration for linezolid was 6425 ng/mL. Its maximal whole-blood bactericidal activity also occurred at > or =2 times the minimum inhibitory concentration, but it was only -0.16 +/- .05 log/day (P< .001) Neither drug showed enhanced activity at higher concentrations. CONCLUSIONS Single doses of PNU-100480 to 1000 mg were well tolerated and exhibited antimycobacterial activity superior to 300 mg of linezolid at steady state. Additional studies are warranted to define its role in drug-resistant tuberculosis.

New treatments are urgently required for infections caused by meticillin-resistant Staphylococcus aureus (MRSA) as these strains are often resistant to multiple conventional antibiotics. Earlier studies showed that ranalexin, an antimicrobial peptide (AMP), in combination with lysostaphin, an antistaphylococcal endopeptidase, synergistically inhibits the growth of MRSA, meaning that it deserved consideration as a new anti-S. aureus therapy. Using haemolysis and Vero cell viability assays, ranalexin with lysostaphin is proven to be non-toxic at antibacterial concentrations. In human serum, ranalexin with lysostaphin is significantly more effective against MRSA than treatment with either component alone. In a rabbit model of wound infection, ranalexin with lysostaphin reduced MRSA in the wound by ca. 3.5log(10) colony-forming units (CFU) compared with the untreated control. The combination is significantly more effective than treatment with ranalexin or lysostaphin alone. In a mouse model of systemic infection, ranalexin with lysostaphin reduced MRSA kidney burden by ca. 1log(10)CFU/g compared with untreated controls or treatment with ranalexin or lysostaphin alone. Importantly, the combination is synergistically bactericidal against various S. aureus isolates in vitro, including those with reduced susceptibility to lysostaphin or vancomycin. Ranalexin and lysostaphin could be incorporated in wound dressings for the prevention and treatment of topical S. aureus infections. That AMPs can enhance the antibacterial effectiveness of lysostaphin in vivo highlights a new avenue of research in the fight against drug-resistant staphylococci.

The amino terminal region of group A streptococcal M proteins evokes type-specific immunity while the conserved C-repeat epitopes evoke cross-protective immunity against multiple serotypes. The present studies were undertaken to compare the protective efficacy of vaccines containing either type-specific (hexavalent vaccine) or conserved C-repeat (J14 vaccine) M protein epitopes and to determine if combination vaccines resulted in enhanced levels of protection. Our results indicated that the protective efficacy of the type-specific vaccine was significantly greater than that of J14 and that the addition of J14 to vaccine formulations did not enhance the level of protection achieved with type-specific formulations.

To evaluate the synergistic activity of erythromycin and human polymorphonuclear cells (PMNs) on the binomial erythromycin-resistant (ERY(R)) Streptococcus pyogenes/host, the phagocytic and bactericidal activities of PMNs against ERY(R) streptococcal strains (cMLS(B), M, and iMLS(B) A, B and C phenotypes) were assessed in the presence of the macrolide. The results showed that when erythromycin, PMNs and streptococci [both erythromycin-sensitive (ERY(S)) and ERY(R)] were simultaneously present in the culture medium, PMN phagocytic activity was similar to that of drug-free controls. In contrast, the results emphasised a significant high increase in intracellular killing by PMNs in the presence of erythromycin not only for ERY(S) streptococci but also for ERY(R)S. pyogenes with high (cMLS(B), iMLS(B) A and iMLS(B) B phenotypes) and moderate (M and iMLS(B) C phenotypes) erythromycin resistance compared with controls without drug. From literature data it emerged that, even if intracellularly concentrated, erythromycin is relatively inactive because of its instability. The results indicate that the enhanced intra-PMN streptococcal killing detected is mainly attributable to PMN bactericidal systems that synergise with intracellular erythromycin in eradicating ERY(R)S. pyogenes strains (both with high and moderate resistance). These data confirm that the antibiotic resistance detected in vitro does not always imply a failure of antimicrobial treatment.

To compare the pharmacokinetics of vancomycin administration by continuous infusion and intermittent infusion.A prospective, randomized, two-way crossover study of 12 patients with methicillin-resistant Staphylococcus aureus infections was conducted. All patients were randomized to receive vancomycin in both regimens consecutively: (i) infusion of 15 mg/kg of vancomycin as a loading dose for 1 h followed by 30 mg/kg of vancomycin as a continuous infusion over 24 h for 48 h; and (ii) intermittent infusion of 15 mg/kg of vancomycin for 1 h every 12 h for 48 h. Vancomycin pharmacokinetic studies were carried out during hours 24-48 after the start of both regimens.For the continuous infusion regimen, the mean highest steady-state concentration was 24.88 +/- 12.75 microg/ml and the mean lowest steady-state concentration was 19.89 +/- 10.15 microg/ml. For the intermittent infusion regimen, the mean peak and trough serum concentrations were 55.02 +/- 17.36 and 12.43 +/- 12.86 microg/ml, respectively. After 10 days of vancomycin treatment, the MRSA infections were eradicated in all patients. Moreover, during both methods of infusion, no adverse events related to the use of vancomycin were observed.Either continuous infusion or intermittent infusion can be used as an effective mode of vancomycin administration to achieve bactericidal activity.

Background Leptospirosis is a zoonosis of worldwide distribution caused by infection with pathogenic serovars of Leptospira spp. The most common species, L. interrogans, can survive in the environment for lengthy periods of time in between infection of mammalian hosts. Transmission of pathogenic Leptospira to humans mostly occurs through abraded skin or mucosal surfaces after direct or indirect contact with infected animals or contaminated soil or water. The spirochete then spreads hematogenously, resulting in multi-organ failure and death in severe cases. Previous DNA microarray studies have identified differentially expressed genes required for adaptation to temperature and osmolarity conditions inside the host compared to those of the environment. Results In order to identify genes involved in survival in the early spirochetemic phase of infection, we performed a transcriptional analysis of L. interrogans serovar Copenhageni upon exposure to serum in comparison with EMJH medium. One hundred and sixty-eight genes were found to be differentially expressed, of which 55 were up-regulated and 113 were down-regulated. Genes of known or predicted function accounted for 54.5 and 45.1% of up- and down-regulated genes, respectively. Most of the differentially expressed genes were predicted to be involved in transcriptional regulation, translational process, two-component signal transduction systems, cell or membrane biogenesis, and metabolic pathways. Conclusions Our study showed global transcriptional changes of pathogenic Leptospira upon exposure to serum, representing a specific host environmental cue present in the bloodstream. The presence of serum led to a distinct pattern of gene expression in comparison to those of previous single-stimulus microarray studies on the effect of temperature and osmolarity upshift. The results provide insights into the pathogenesis of leptospirosis during the early bacteremic phase of infection.

We recently demonstrated that the accumulation of ceramide in Cftr-deficient epithelial cells is important for the pathophysiology of CF. However, the role of ceramide in other lung cells, particularly lung macrophages, requires definition. In this study, we report that ceramide is accumulated in Cftr-deficient lung macrophages. Alveolar macrophages contain a vesicle population, which is stained with LysoSensor probes but not by tetramethylrhodamine dextran. These vesicles, presumably secretory lysosomes, exhibit a higher pH in Cftr-deficient macrophages than the corresponding vesicles in lung macrophages isolated from wild-type (WT) mice. Alkalinization of these vesicles in Cftr-deficient macrophages correlates with a failure of the macrophages to respond to infection with various Pseudomonas aeruginosa strains by acutely activating acid sphingomyelinase, releasing ceramide, forming ceramide-enriched membrane platforms that serve to cluster gp91phox, and, most importantly, releasing reactive oxygen species (ROS). In contrast, these events occur rapidly in WT lung macrophages postinfection. Inhibiting ROS in WT macrophages prevents the killing of P. aeruginosa. These findings provide evidence for a novel pH-controlled pathway from acid sphingomyelinase activation via ceramide and clustering of gp91phox to the release of ROS in lung macrophages.

The role of tolerance was investigated in the prophylaxis of Staphylococcus aureus endocarditis with cloxacillin in rats. The effect of a single dose of 500 mg/kg, two 80 mg/kg doses 3 h apart, and a single dose of 80 mg/kg, alone or in combination with a single dose of 4 mg of gentamicin/kg, were compared for a tolerant strain of S. aureus and its isogenic nontolerant variant. At all dosages, cloxacillin was significantly less effective in preventing endocarditis with the tolerant strain than with the nontolerant variant. With the high dose of cloxacillin or two successive lower doses, nearly complete protection could be obtained against the nontolerant strain. However, for the tolerant strain, only the combination of cloxacillin and gentamicin afforded almost complete protection. For the tolerant strain, no serum bactericidal activity was found at the time of bacterial challenge after an injection of any dose of cloxacillin. These results suggest that the in vitro phenomenon of tolerance may have relevance in vivo.

14-Hydroxyclarithromycin, an active metabolite of clarithromycin, was compared for antimicrobial activity alone and in combination with the parent compound. The 14-hydroxyclarithromycin potency was comparable to that of clarithromycin, but was more active against Haemophilus influenzae (MIC50, 1 microgram/ml). Combination MICs at pharmacokinetic ratios produced end points equal to the most active component of the combination. However, checkerboard MICs and kill-curve studies suggested enhanced interactive effects. Partial synergy and additive interactions were demonstrated in 96% of strains tested with synergy (partial) most often observed among the Enterococcus faecalis, H. influenzae, and staphylococci. To determine the best in vitro test methods for predicting the value of 14-hydroxyclarithromycin, combination disks or ratio MIC tests may not be practical. A modification of the proposed clarithromycin-susceptible breakpoint (less than or equal to 2 micrograms/ml) upward to less than or equal to 4 micrograms/ml (greater than or equal to 14 mm) was suggested to recognize the additional activity contributed by the 14-hydroxy metabolite. This modification should be applied, limited to susceptibility tests of H. influenzae and possibly the enterococci.

Meningococcal factor H-binding protein (fHbp) is a promising vaccine candidate that elicits serum bactericidal antibodies in humans. Based on sequence variability of the entire protein, fHbp has been divided into three variant groups or two sub-families. We recently reported that the fHbp architecture was modular, consisting of five variable segments, each encoded by genes from one of two lineages. Based on combinations of segments from different lineages, all 70 known fHbp sequence variants could be classified into one of six modular groups. In this study, we analyzed sequences of 172 new fHbp variants that were available from public databases. All but three variants could be classified into one of the six previously described modular groups. Among systematically collected invasive group B isolates from the U.S. and Europe, modular group I overall was most common (60%) but group IV (natural chimeras) accounted for 23% of UK isolates and

To optimize the serum bactericidal assay (SBA), detect and analyze the bactericidal antibody level against Neisseria meningitidis serogroup C strains after divalent polysaccharide (A plus C) vaccine immunization.Two Neisseria meningitidis serogroup C strains, vaccine candidate strain (C11) and epidemic strain (053442), were selected as targets. The national Neisseria meningitidis standardized serum was used as reference serum. Pel-Freez infant rabbit complements was available. The optimized SBA method was used to detect bactericidal antibody against strain C11 and 053442 for 122 pairs of sera before and after immunization with a divalent polysaccharide (A and C) vaccine.The strain C11 and 053442 both could be used as targeted strain for SBA. The optimized concentration of targeted strain was achieved when a whole-cell suspension of 0.35 A at 600 nm was diluted 4 x 10(4) times. Before immunization, SBA geometric mean titers (GMT) of 122 sera against strain C11 and 053442 were 1:1.75 and 1:2.63 respectively, and the protective rates were 9.8% and 17.2% respectively. After immunization, the GMTs and the protective rates of 122 sera both rose significantly (P0.01), the GMTs against strain C11 and 053442 were 1:483.73 and 1:412.57 respectively. The protective rates against strain C11 and 053442 were 100% and 95.9% respectively.Immunization with a divalent polysaccharide (A and C) vaccine could elevate remarkably the population SBA titer against Neisseria meningitidis serogroup C strains of different subtypes, but the surveillance of vaccine effect against different targeted strains remains necessary.

Factor H binding proteins (fHBP), are bacterial surface proteins currently undergoing human clinical trials as candidate serogroup B Neisseria meningitidis (MnB) vaccines. fHBP protein sequences segregate into two distinct subfamilies, designated A and B. Here, we report the specificity and vaccine potential of mono- or bivalent fHBP-containing vaccines. A bivalent fHBP vaccine composed of a member of each subfamily elicited substantially broader bactericidal activity against MnB strains expressing heterologous fHBP than did either of the monovalent vaccines. Bivalent rabbit immune sera tested in serum bactericidal antibody assays (SBAs) against a diverse panel of MnB clinical isolates killed 87 of the 100 isolates. Bivalent human immune sera killed 36 of 45 MnB isolates tested in SBAs. Factors such as fHBP protein variant, PorA subtype, or MLST were not predictive of whether the MnB strain could be killed by rabbit or human immune sera. Instead, the best predictor for killing in the SBA was the level of in vitro surface expression of fHBP. The bivalent fHBP vaccine candidate induced immune sera that killed MnB isolates representing the major MLST complexes, prevalent PorA subtypes, and fHBP variants that span the breadth of the fHBP phylogenetic tree. Importantly, epidemiologically prevalent fHBP variants from both subfamilies were killed.

Vancomycin-resistant enterococci pose an emerging health risk. The limitation in therapeutic options has resulted in the development of new drugs such as quinupristin/ dalfopristin and linezolid. AIM, SETTING AND DESIGN: This study investigated the species prevalence and antibacterial resistance among enterococci isolated in selected Tehran hospitals.Between March 2006 and August 2007, 200 enterococcal isolates from urine, blood, stool and wound were recovered in 2 teaching hospitals of Tehran province. Susceptibility of all isolates was tested against vancomycin, teicoplanin and linezolid antibiotics by disk diffusion and agar dilution method.Seventeen (8.5%), 6 (3%) and 4 (2%) of the isolates were resistant to vancomycin, teicoplanin and linezolid, respectively. Within the vancomycin-resistant isolates, 6 (35.2%), 4 (25%) and 1 (5.88%) showed vanA, vanB and vanC genotype patterns, respectively. Four (23.5%) of VRE isolates were resistant to linezolid with minimum inhibitory concentrations between 16 and 32 microg/mL. Two linezolid vancomycin resistant enterococci were E. faecium.

The objectives of this study were to determine pharmacokinetics of intravenous (i.v.) ceftiofur in foals, to compare ultra-high performance liquid chromatography tandem mass spectometry (UPLC-MS/MS) and microbiologic assay for the measurement of ceftiofur concentrations, and to determine the minimum inhibitory concentration (MIC) of ceftiofur against common equine bacterial pathogens. In a cross-over design, ceftiofur sodium was administered i.v. to six foals (1-2 days-of-age and 4-5 weeks-of-age) at dosages of 5 and 10 mg/kg. Subsequently, five doses of ceftiofur were administered i.v. to six additional foals between 1 and 5 days of age at a dose of 5 mg/kg q 12 h. Concentrations of desfuroylceftiofur acetamide (DCA), the acetamide derivative of ceftiofur and desfuroylceftiofur-related metabolites were measured in plasma, synovial fluid, urine, and CSF by use of UPLC-MS/MS. A microbiologic assay was used to measure ceftiofur activity for a subset of plasma samples. Following i.v. administration of ceftiofur at a dose of 5 mg/kg to 1-2 day-old foals, DCA had a t(1/2) of 7.8 +/- 0.1 h, a body clearance of 74.4 +/- 8.4 mL/h/kg, and an apparent volume of distribution of 0.83 +/- 0.09 L/kg. After multiple i.v. doses at 5 mg/kg, DCA concentrations in CSF were significantly lower than concurrent plasma concentrations. Ceftiofur activity using a microbiologic assay significantly underestimated plasma concentrations of DCA. The MIC of ceftiofur required to inhibit growth of 90% of isolates of Escherichia coli, Pasteurella spp, Klebsiella spp, and beta-hemolytic streptococci was

Triazolylmethyl oxazolidinones are novel potent antibacterial agents recently synthesized in our laboratory. PURPOSE. Development of a rapid and specific LC-MS method for evaluating the stability of three potentially active antibacterial triazolylmethyl oxazolidinone compounds, namely PH-27, PH-38 and PH-41, in human plasma. METHODS. Ion-trap mass spectrometry using +APCI-MS at m/z 348.1 (PH-27), 389.1 (PH-38) and 451.1 (PH-41) combined with liquid chromatographic analysis using C18 reversed-phase column and a mobile phase consisting of 20 mM ammonium acetate solution and acetonitrile (50:50 v/v) was used. Plasma samples were pre-treated with acetonitrile to precipitate proteins prior to LC-MS analysis. Linezolid was used as an internal standard whose predominant ion at m/z 338.1 was monitored and used to construct the calibration curves. An accelerated stability study under controlled experimental conditions was conducted at -20 oC for a 5-weeks period. RESULTS. Calibration curves of the examined oxazolidinones in human plasma were established (r: 0.99) up to the concentration of 20.0 µg ml-1 with LLQ of 0.1 (PH-27), 0.5 (PH-38) and 1.0 µg ml-1 (PH-41). Validation data showed appropriate intra- and inter-day precision and accuracy as indicated from RSD% 1.3 - 8.6 and % DEVs -12.7 to +11.3, respectively. Stability studies gave the kinetic degradation parameters of kdeg 0.0191- 0.0857 week-1, t1/2 8.1-36.3 weeks and t90 1.2 -5.5 weeks. CONCLUSIONS. The developed +APCI-LC-MS assay is simple, fast and specific method that could be applied for the routine analysis of the selected oxazolidinones during stability and pharmacokinetic studies. The examined compound PH-27 was proved to be relatively more stable and resistant to degradation in human plasma compared to PH-38 and PH-41, as indicated from the kinetics and recovery studies at -20 oC.

A high-throughput bioluminescence screening procedure for identification of virulence genes in bacteria was developed and applied to the fish pathogen Edwardsiella ictaluri . A random transposon mutant library expressing bioluminescence was constructed and robotically arrayed on 384-well plates. Mutants were cultivated and mixed with catfish serum and neutrophils in 96-well plates, and bioluminescence was used to detect mutants that are more susceptible to killing by these host factors. The virulence and vaccine efficacy of selected mutants were determined in channel catfish. Transposon insertion sites in 13 mutants attenuated in the natural host were mapped to the E. ictaluri genome. Ten unique genes were mutated, including genes encoding a negative regulator of sigmaE activity, a glycine cleavage system protein, tricarboxylic acid cycle enzymes, an O polysaccharide biosynthesis enzyme, proteins encoded on the native plasmid pEI1, and a fimbrial chaperon protein. Three of these mutants were found to have potential as live attenuated vaccines. This study demonstrates a novel application of bioluminescence to identify bacterial genes required for host resistance; as a result, efficacious and genetically defined live attenuated vaccine candidates were developed.

A dynamic in vitro model was used to determine the killing kinetics of daptomycin against 15 vancomycin-resistant clinical isolates of Enterococcus faecium. Concentration profiles simulating those observed in serum following administration of both low-dose (2 mg/kg) and high-dose (6 mg/kg) daptomycin were bactericidal within 5.5 and 2.8 h, respectively. In contrast, when albumin was added to the growth medium, the corresponding bacterial killing times were slowed to greater than 24 h and 7 h; these results suggest that in the clinical setting, daptomycin dosages of approximately 6 mg/kg are required to achieve bactericidal activity against vancomycin-resistant Enterococcus faecium.

To better define the pharmacokinetics and serum bactericidal activity (SBA) of ciprofloxacin and other antimicrobial agents in the elderly, six healthy (greater than 65 years) volunteers with normal renal function were given ciprofloxacin alone orally, ciprofloxacin plus rifampin orally, ciprofloxacin plus clindamycin orally, rifampin alone orally (three volunteers), and, for comparison of SBA against gram-positive cocci, vancomycin intravenously. Mean peak ciprofloxacin concentrations and other pharmacokinetic parameters were not altered significantly by coadministration of either rifampin or clindamycin. Ciprofloxacin had somewhat greater SBA against the oxacillin-susceptible and oxacillin-resistant Staphylococcus aureus strains tested than did vancomycin, but rifampin was by far the most active single agent tested. The SBA of rifampin against S. aureus was modestly antagonized during combination therapy with ciprofloxacin, but substantial SBA still was present. The ciprofloxacin SBA against S. aureus was completely antagonized by clindamycin if the strains were susceptible to the latter agent. Ciprofloxacin had modest SBA against group A streptococci and no SBA against the three pneumococcal strains tested. All of the regimens had poor to absent SBA against Enterococcus faecalis. By contrast, ciprofloxacin had excellent SBA against Escherchia coli and Klebsiella pneumoniae and moderate SBA against Pseudomonas aeruginosa. Combination therapy with rifampin or clindamycin in general enhanced the SBA against the nonenterococcal streptococci and had no effect on the SBA against the gram-negative bacilli.

Summary The influence of a new oral cephalosporin cefpodoxime, on several interactions of the host-parasite relationship was investigated. Pretreatment of polymorphonuclear granulocytes (PNG) with cefpodoxime at therapeutical dosage in vitro did not have a significant effect on chemotaxis and luminol-dependent chemiluminescence. On the other hand, human serum did not exert a considerable effect on the bactericidal activity of cefpodoxime on Klebsiella pneumoniae and Staph. aureus . Serum resistance of Klebsiella pneumoniae was not altered after pretreatment with cefpodoxime 1116 MIC (0.03 μg/ml) for 3 to 27 hours.

A colorimetric assay was developed for quantitating bovine neutrophil bactericidal activity against Staphylococcus aureus. The procedure used the tetrazolium compound, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The assay was conducted by incubating antibody-opsonized S. aureus with neutrophils in microtiter plates for 1 h at a ratio of 10 bacteria per neutrophil. Neutrophils were then lysed with saponin. The MTT was added and samples were incubated for 10 min. Live S. aureus reduced MTT to purple formazan. Dead bacteria and lysed neutrophils did not react with MTT. Bacterially-reduced formazan was solubilized by adding isopropanol and formazan production was quantitated by measuring absorption at 560 nm. Absorption of formazan was directly related to viable bacteria cell number and was used to determine the number of S. aureus not killed by neutrophils. The percentage of bacteria killed by neutrophils was determined by extrapolation from a standard formazan curve that was derived by incubating MTT with known numbers of S. aureus. The colorimetric MTT assay detected suppressed bactericidal activity after in vitro treatment of bovine neutrophils with colchicine, cytochalasin B, or phorbol 12-myristate 13-acetate. In vitro treatment of neutrophils with low levels of recombinant bovine interferon gamma (rBoIFN-gamma) enhanced bactericidal activity, whereas high levels decreased activity. These results suggest the colorimetric MTT bactericidal assay is efficacious in detecting modulation of bovine neutrophil bactericidal activity. Furthermore, the MTT assay has many advantages over traditional bactericidal assays in that it is sensitive, inexpensive, requires less than 3 h to complete, and can analyze many neutrophil samples in a single day.

Cephalosporins differ in their ability to prevent staphylococcal wound infection. Although the reasons for the observed differences are not fully understood, the susceptibility of cephalosporins to hydrolysis by staphylococcal beta-lactamase has been correlated with failures of prophylaxis. To investigate the effect of beta-lactamase stability and other in-vitro parameters of the bacterial-antimicrobial interaction on the efficacy of antimicrobial prophylaxis, two beta-lactamase-stable agents, cefuroxime and cefmetazole were compared to cefazolin and cefamandole in an in-vivo model of intradermal infection employing four strains of Staphylococcus aureus. Following intraperitoneal administration of a single dose of cephalosporin or placebo, guinea pigs were inoculated at multiple intradermal sites with 2 x 10(7) cfu of a strain of staphylococcus. For three strains, the area of induration at 24 h following inoculation was significantly smaller in guinea pigs receiving prophylaxis with cephalosporins versus placebo; no cephalosporin was effective against the fourth strain. Differences were also noted among the cephalosporins; prophylaxis with cefuroxime and cefmetazole resulted in smaller lesions than seen in animals given cefazolin or cefamandole. Poor correlation was noted between results of the in-vivo model and in-vitro determinants of the bacterial-antimicrobial interaction which were MIC values, time-kill curves, and the rates of beta-lactamase-mediated cephalosporin hydrolysis by the different strains. The model demonstrated unexplained failures of prophylaxis and unexpected differences in efficacy of various cephalosporins as has been described before. This study highlights the need for an improved animal model of surgical antimicrobial prophylaxis and the identification of in-vitro determinants that predict in-vivo prophylactic efficacy more accurately.

Sera from healthy infants (under 1 year old), toddlers (3–4 years) and adults (18–65 years) were assayed for their ability to bind to inner core (ic) lipopolysaccharide (LPS) epitopes of Neisseria meningitidis. Antibodies (Abs) reacting to inner core structures, including different substitutions of the first heptose (HepI) and second heptose (HepII) residues of the LPS backbone, truncated and fully extended LPS glycoforms, were detected and for each structure, these inner core antibodies showed an age-related pattern of acquisition. A novel column-based methodology was used to affinity purify IgG antibodies in which purified inner core LPS (derived from a mutant MC58) was covalently linked to Sepharose 4B. Comparison of reactivity before and after affinity purification of the pooled sera showed that the purified Abs bound to the surface of N. meningitidis organisms displaying truncated and extended LPS with a homologous inner core region, promoted the deposition of C3b, were opsonophagocytic in vitro and decreased bacteraemia when used to passively protect infants rats. In addition, the purified Abs were bactericidal in vitro against the mutant strain displaying truncated LPS with a homologous inner core region. These results demonstrate that naturally occurring serum human antibodies to N. meningitidis LPS can access inner core epitopes of encapsulated organisms with a fully extended LPS.

We addressed the hypothesis that meningococcal secreted proteins (MSPs) can elicit protective immunity against meningococcal disease. Endotoxin-depleted MSP preparations were used to immunise a group of 15 six-week-old BALB/c mice (25 μg MSPs/dose mixed with Freund's complete adjuvant) on days 0, 14 and 21. Mice were challenged 2 weeks later with 107 colony forming units of live Neisseria meningitidis strain MC58 (serogroup B, ET-5). Negative and positive control groups of 15 mice each were injected with adjuvant only, or a live attenuated strain of MC58, respectively. Seven out of 15 mice (47%) from the negative control group died after 72 h of challenge, whereas none of test or positive control group died. Protection afforded by the anti-MSP immune response can be at least partly attributed to complement-mediated bacterial lysis, detectable in vitro using the serum of immunised mice. Murine anti-MC58 MSP sera were bactericidal against homologous and five unrelated ET-5 serogroup B strains. However, failed to kill strains from other hypervirulent clonal lineages belonging to the same or different serogroups, despite the presence of cross-reactive antibodies detectable by immunoblotting. Similar sera raised against MSPs from an isolate belonging to the ET-37 electropherotype lineage were bactericidal against all tested isolates of this lineage and, in addition, against some but not all isolates belonging to the ET-5 lineage. FACS analysis of intact bacteria treated with anti-MSPs confirmed surface-binding of antibodies.

The effect of subinhibitory concentrations of amikacin and ciprofloxacin on the susceptibility of Escherichia coli strains to the bactericidal action of normal human serum, outer membrane protein (OMP) expression and cell morphology was investigated. Sub-minimal inhibitory concentrations (sub-MICs) of both antibiotics were found to alter the expression of some OMPs of serum-resistant E. coli rods, the morphology of their cells and made them sensitive to the bactericidal activity of normal human serum.

Rationale: The effectiveness and safety of aztreonam lysine for inhalation (AZLI) in patients with cystic fibrosis (CF) on maintenance treatment for Pseudomonasaeruginosa (PA)airway infection was evaluated in this randomized, double-blind, placebo-controlled study. Objectives: To evaluate the safety and efficacy of inhaled aztreonam lysine in controlling PA infection in patients with CF. Methods: After randomization and a 28-day course of tobramycin inhalationsolution (TIS),patients(n 5211; >6yr;>3TIScourseswithin previous year; FEV1 > 25% and

The immunological status was studied in 335 patients with urolithiasis and pyelonephritis. The techniques used for assessment of the immunological status provided a multifactorial control over immune system function in inflammation and urolithiasis: registered a significant rise of the level of proinflammatory and anti-inflammatory mediators of the immune response (IL-1beta, IL-4, IL-6, TNF), of neutrophil metabolic activity with parallel suppression of neutrophil phagocytic and absorbing functions, inhibition of bactericidal activity of the serum, a significant fall in the number of mature T-lymphocytes (CD3+) and T-helpers (CD4+). The immune status indices in urolithiasis patients with secondary pyelonephritis are closely linked with clinico-laboratory manifestations of the inflammatory process and renal function. Thus, the results of the study give grounds for developing indications for use of immunomodulating drugs in patients with secondary pyelonephritis associated with urolithiasis.

Context Immunization with a meningococcal tetravalent (serogroup ACWY) glycoconjugate vaccine is recommended for all US adolescents. However, the currently licensed vaccine is poorly immunogenic in infancy, when the highest rates of disease are observed. Objective To determine the immunogenicity of a novel tetravalent CRM197-conjugated meningococcal vaccine (MenACWY) in infants. Design, Setting, and ParticipantsRandomized, open-label, controlled study of 225 UK and 196 Canadian 2-month-olds from August 2004 to September 2006.Intervention UK infants received a primary course of MenACWY (at 2, 3, and 4 months or 2 and 4 months) or Neisseria meningitidis serogroup C monovalent meningococcal glycoconjugate vaccine (MenC) (at 2 and 4 months). All received MenACWY at 12 months. Canadian infants received MenACWY at 2, 4, and 6 months or 2 and 4 months; at 12 months they received MenACWY, a plain tetravalent polysaccharide vaccine, or no vaccine. Main Outcome MeasurePercentage of infants with a human complement serum bactericidal activity (hSBA) titer ≥1:4 after a primary course of MenACWY and after a 12-month booster. Safety and reactogenicity of MenACWY were also assessed.Results According to the prespecified per-protocol analysis, the percentages (95% CIs) of MenACWY 2-, 3-, and 4-month recipients with hSBA titers ≥1:4 after primary immunization were serogroup A, 93% (84%-98%); C, 96% (89%-99%); W-135, 97% (90%-100%); and Y, 94% (86%-98%). With a post hoc intention-to-treat analysis with imputed values for missing data, these values were unchanged for serogroups C and Y; for serogroup A, values were 92% (84%-97%), and for W-135, 97% (91%-99%). For the per-protocol analysis of MenACWY 2-, 4-, and 6-month recipients, the percentages (95% CIs) of responders were A, 81% (71%-89%); C, 98% (92%-100%); W-135, 99% (93%-100%); and Y, 98% (92%-100%). With the imputed value analysis, these values were A, 83% (74%-89%); C, 98% (93%-99%); W-135, 99% (94%-100%); and Y, 98% (92%-99%). At least 84% of MenACWY 2- and 4-month recipients achieved hSBA titers ≥1:4 for serogroups C, W-135, and Y after primary immunization, as did at least 60% for serogroup A (per-protocol and imputation analysis). At least 95% of primary and booster MenACWY recipients achieved hSBA titers ≥1:4 for serogroups C, W-135, and Y at 13 months, as did at least 84% for serogroup A (per-protocol and imputation analysis). During the primary immunization course, postimmunization pain on leg movement was observed in 2% of UK MenACWY 2- and 4-month recipients and 4% of MenC 2- and 4-month recipients; a temperature of 38°C or greater was observed in 4% and 2% in these groups, respectively. ConclusionMenACWY is well tolerated and immunogenic in infancy.Trial Registration clinicaltrials.gov Identifier: NCT00262002

The etiology of acute lower respiratory tract infection (ALRI) was identified in 235 (43.8%) of 537 hospitalized children less than 5 years of age. Clinical evidence of measles was found in 258 (48.0%) patients, of whom 59 had a second viral infection. A viral agent was identified in an additional 121 patients, so that a total of 379 (70.6%) had viral infections. After measles, respiratory syncytial virus was the most common respiratory virus. Bacteremia was noted in 72 children (13.4%), occurring as frequently in children with measles (14.8%) as in those without (12.1%); Haemophilus influenzae and Salmonella typhi were predominant in the former, and H. influenzae, Staphylococcus aureus, and Streptococcus pneumoniae were prominent in the latter. The presence of bacterial antigen in urine was not helpful in identifying bacterial infection. Extrapulmonary and intrapleural complications, concomitant measles, complicated ALRI, female gender, and malnutrition were associated with increased mortality among children with ALRI. The importance of measles immunization, vitamin A supplementation for alleviation of defects associated with malnutrition, and timely antimicrobial therapy is emphasized.

The effect of sublethal concentrations of antibiotics on the serum sensitivity of Pseudomonas aeruginosa was examined. Cefepime, ceftazidime, and imipenem but not amikacin nor ciprofloxacin increased the serum bactericidal activity of pooled normal human serum. Killing was both serum- and antibiotic dose-dependent. Increased sensitivity to the bactericidal action of serum in the presence of cefepime was observed for several different clinical isolates. With the use of C8-deficient sera, the late components of the complement pathway were shown to be essential for bacterial killing. A significant increase in the amount of [125I]C9 based on bacterial mass was observed with bacteria incubated with cefepime compared with non-antibiotic- or amikacin-treated controls. No major change in the amount of type of lipopolysaccharide was observed when cefepime-treated and control bacteria were compared. The data show that cefepime and other cephalosporins at sublethal concentrations increase the complement-mediated bactericidal activity of serum against P. aeruginosa.

To better define the pharmacokinetics and serum bactericidal activity (SBA) of the expanded-spectrum cephalosporins in the elderly, we administered single 2-g intravenous infusions of cefoperazone, cefotaxime, ceftriaxone, ceftazidime, and ceftizoxime to six healthy volunteers over the age of 65 years. Serum was collected over 24 h, and concentrations were determined by high-performance liquid chromatography; pharmacokinetic parameters were determined for each drug. SBA was measured against representative strains of Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Enterobacter aerogenes, and Pseudomonas aeruginosa. All agents tested had excellent SBAs against E. coli and K. pneumoniae, often for a longer duration than would be expected on the basis of conventional dosing regimens. Ceftazidime had the greatest SBA against E. aerogenes and was the only agent with a substantial SBA against P. aeruginosa. Although ceftizoxime had the greatest SBA against S. aureus, none of these cephalosporins had substantial antistaphylococcal SBAs. Pharmacokinetic analysis revealed that cefoperazone and ceftriaxone had markedly different concentration-time profiles in the elderly volunteers than would have been expected on the basis of existing data from younger volunteers. For older patients, dosing guidelines for these two agents may need to be altered.

We studied the prevention of experimental aortic endocarditis caused by a beta-lactamase-producing, aminoglycoside-resistant strain of Enterococcus faecalis (HH22) in 146 catheterized rabbits. Both vancomycin and ampicillin-sulbactam readily killed this resistant enterococcus strain in vitro. At a challenge inoculum of approximately 10(9) CFU, vancomycin (40 mg/kg intravenously [i.v.]), ampicillin (40 mg/kg i.v.), or a combination of ampicillin plus a beta-lactamase inhibitor, sulbactam (20 mg/kg, i.v.), did not prevent the development of endocarditis in any of the animals, although mean intravegetation bacterial densities were significantly lower in animals that received vancomycin than they were in animals that received other therapies (P less than 0.001). At a challenge inoculum of 10(6) CFU, vancomycin was 100% effective in preventing enterococcal endocarditis compared with ampicillin (29%; P less than 0.00001) and ampicillin-sulbactam (65%; P less than 0.01). Factors associated with the superior prophylactic efficacy of vancomycin in this model included prolonged serum inhibitory activity and time above MICs. Factors not associated with the antienterococcal prophylactic efficacy of vancomycin included the duration of the in vitro postantibiotic effect of the drug and the magnitude of the ability of this drug to enhance enterococcal in vitro opsonophagocytic killing by polymorphonuclear leukocytes. The superior prophylactic efficacy of vancomycin in this endocarditis model related to the superior pharmacokinetic profile of the drug when it was given intermittently at dose intervals of every 6 h.

The bactericidal activity of daptomycin and vancomycin alone in cation-supplemented Mueller-Hinton broth and in human serum against clinical isolates of Staphylococcus aureus, Staphylococcus epidermidis, and Enterococcus faecalis was evaluated by exposing replicating microorganisms to concentrations ranging from 2 to 128 micrograms/ml for 24 hr. In addition, the possibility of emergence of resistance, the stability of each agent in the respective medium, and the percent of protein binding by human serum for each agent was evaluated. We found that a concentration of less than or equal to 8 micrograms/ml of daptomycin was sufficient to achieve bactericidal activity (greater than or equal to 99.9% killing of the inoculum) in cation-supplemented Mueller-Hinton broth for all staphylococcal isolates tested; a concentration of less than or equal to 16 micrograms/ml of daptomycin was required for bactericidal activity in cation-supplemented Mueller-Hinton broth for enterococcal isolates. In human serum, comparable bactericidal activity with daptomycin was achieved only with concentrations 8-16 times higher. A similar but less pronounced effect in human serum was seen for vancomycin. Neither daptomycin nor vancomycin was appreciably degraded in human serum over a 24-hr period. It is likely that the clinical efficacy of daptomycin in humans would be enhanced by higher dosing than has been studied to date.

The influence of protein binding on anti-staphylococcal activity of cefotaxime, ceftazidime and ceftriaxone was evaluated using an ex vivo cross-over study in human volunteers. Serum inhibitory and bactericidal titres were measured using samples collected 1 h and 6 h after infusion of 2 g of cefotaxime and ceftazidime and 1 g of ceftriaxone. Time-kill curves were done using the same samples. Twelve strains of Staphylococcus aureus were studied including six capsulated strains of known capsule types. Two media were compared: Mueller-Hinton and 'capsule' medium. The MICs and MBCs of oxacillin-susceptible strains measured in capsule medium were higher than in Mueller-Hinton, hence the higher serum inhibitory and bactericidal titres in the latter medium. The effect of protein binding was evaluated, using the difference between expected serum titres, from serum concentration and MICs or MBCs measured in serum-free medium, and the area under the time-kill curve corrected for the serum concentration and the sensitivity of the strains. The anti-staphylococcal activity of ceftriaxone was markedly affected by its strong protein binding.

Bactericidal activity in compromised host's serum i.e. patients with cancer, the elderly, diabetes, was stronger than that in normal serum against Proteus mirabilis (P. mirabilis) but was weaker against Klebsiella pneumoniae (K. pneumoniae). Against Escherichia coli (E. coli), bactericidal activity on serum of patients with cancer was weaker in the following order, that in elderly serum, that in diabetic serum. Against Proteus vulgaris (P. vulgaris), bactericidal activity in elderly serum was similar to that in normal serum but was stronger than that in serum of patients with cancer. Against Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus), bactericidal activity in elderly serum and diabetic serum was similar to that in normal serum but was weaker than that in serum of patients with cancer. Piperacillin showed bactericidal activity in nutrient broth, normal serum and compromised host's serum at a concentration of 1/4 MIC against E. coli, P. mirabilis and P. aeruginosa. Aspoxicillin showed bactericidal activity in nutrient broth, and bactericidal or bacteriostatic activity in normal serum and serum of patients with cancer against E. coli and P. aeruginosa. While cefazolin and cefmetazole slightly inhibited the growth of bacteria in nutrient broth, they showed hardly any bactericidal activity in normal serum and compromised host's serum.

We compared the bactericidal activity of serum obtained from healthy volunteers after single intravenous infusions of the combination of ceftizoxime (1 g) plus metronidazole (1 g) and after infusions of ceftizoxime (2 g), clindamycin (900 mg), and imipenem (1 g) against six obligate anaerobes. All agents were bactericidal but only the combination regimen resulted in bactericidal titres greater than 1:2 at 12 h for all the Bacteroides fragilis group organisms. High titres against Fusobacterium necrophorum and anaerobic Gram-positive cocci were attained with ceftizoxime/metronidazole, ceftizoxime, and imipenem 12 h after the dose. Imipenem and the combination of ceftizoxime/metronidazole had the greatest area under the bactericidal curve (AUBC) against the Bacteroides species. Clindamycin had a significantly smaller AUBC than other regimens for all strains tested except B. thetaiotaomicron. Clinical trials are needed to assess the efficacy and cost-benefit ratio of a 12 h dosing regimen of ceftizoxime in combination with metronidazole for treating mixed aerobic/anaerobic infections.

The Wzz proteins are important for determining the length of the O-antigen side chain attached to lipopolysaccharide (LPS). Several bacteria, including Pseudomonas aeruginosa strain PAO1 (serogroup O5), produce two such proteins responsible for the preference of two different chain lengths on the surface. Our group has previously identified one wzz gene ( wzz1 ) within the O-antigen locus of P. aeruginosa strain PA103 (serogroup O11). In this study we have identified the second wzz gene ( wzz2 ), located in the same region of the genome and with 92% similarity to PAO1's wzz2 gene. Mutations were generated in both wzz genes by interruption with antibiotic resistance cassettes, and the effects of these mutations were characterized. Wild-type PA103 prefers two O-antigen chain lengths, referred to as long and very long. The expression of the long O-antigen chain length was reduced in the wzz1 mutant, indicating the Wzz1 protein is important for this chain length preference. The wzz2 mutant, on the other hand, was missing O-antigens of the very long chain length, indicating the Wzz2 protein is responsible for the production of very long O-antigen. The effects of the wzz mutations on virulence were also investigated. In both serum sensitivity assays and a mouse pneumonia model of infection, the wzz1 mutants exhibited greater defects in virulence compared to either wild-type PA103 or the wzz2 mutant, indicating the long chain length plays a greater role during these infectious processes.

The Yersinia adhesin A (YadA) is a trimeric autotransporter adhesin of enteric yersiniae. It consists of three major domains: a head mediating adherence to host cells, a stalk involved in serum resistance, and an anchor that forms a membrane pore and is responsible for the autotransport function. The anchor contains a glycine residue, nearly invariant throughout trimeric autotransporter adhesins, that faces the pore lumen. To address the role of this glycine, we replaced it with polar amino acids of increasing side chain size and expressed wild-type and mutant YadA in Escherichia coli . The mutations did not impair the YadA-mediated adhesion to collagen and to host cells or the host cell cytokine production, but they decreased the expression levels and stability of YadA trimers with increasing side chain size. Likewise, autoagglutination and resistance to serum were decreased in these mutants. We found that the periplasmic protease DegP is involved in the degradation of YadA and that in an E. coli degP deletion strain, mutant versions of YadA were expressed almost to wild-type levels. We conclude that the conserved glycine residue affects both the export and the stability of YadA and consequently some of its putative functions in pathogenesis.

The “gold standard” assay for measuring serologic protection against Neisseria meningitidis group B (MenB) is the serum bactericidal antibody (SBA) assay. Of vital importance to the outcome of the SBA assay is the choice of the target strain(s), which is often chosen on the basis of phenotype or genotype. We therefore investigated the effect on the results produced by the SBA assay of using phenotypically indistinguishable but geographically distinct MenB isolates. Nine PorA P1.19,15 and 11 PorA P1.7-2,4 MenB isolates were incorporated into the SBA assay using human complement and were assayed against sera obtained either before or after outer membrane vesicle vaccination. Large differences in the results produced by the isolates in the SBA assay were demonstrated. These included differences as great as 5.8-fold in SBA geometric mean titers and in the proportions of subjects with SBA titers of ≥4. Ranges of as many as 9 SBA titers were achieved by individual sera across the panels of isolates. To determine the reasons for the differences observed, investigations into the expression of capsular polysaccharide, PorA, PorB, Opc, and lipooligosaccharide (LOS) and into LOS sialylation were completed. However, minor differences were found between strains, indicating similar expression and no antigen masking. These results have implications for the choice of MenB target strains for inclusion in future studies of MenB vaccines and highlight the requirement for standardization of target strains between laboratories.

The genes coding for the five outer membrane proteins (OMPs) of the Omp25/Omp31 family expected to be located in the outer membrane (OM) of rough virulent Brucella ovis PA were inactivated to evaluate their role in virulence and OM properties. The OM properties of the mutant strains and of the mutants complemented with the corresponding wild-type genes were analyzed, in comparison with the parental strain and rough B. abortus RB51, in several tests: (i) binding of anti-Omp25 and anti-Omp31 monoclonal antibodies, (ii) autoagglutination of bacterial suspensions, and (iii) assessment of susceptibility to polymyxin B, sodium deoxycholate, hydrogen peroxide, and nonimmune ram serum. A tight balance of the members of the Omp25/Omp31 family was seen to be essential for the stability of the B. ovis OM, and important differences between the OMs of B. ovis PA and B. abortus RB51 rough strains were observed. Regarding virulence, the absence of Omp25d and Omp22 from the OM of B. ovis PA led to a drastic reduction in spleen colonization in mice. While the greater susceptibility of the Δ omp22 mutant to nonimmune serum and its difficulty in surviving in the stationary phase might be on the basis of its dramatic attenuation, no defects in the OM able to explain the attenuation of the Δ omp25d mutant were found, especially considering that the fully virulent Δ omp25c mutant displayed more important OM defects. Accordingly, Omp25d, and perhaps Omp22, could be directly involved in the penetration and/or survival of B. ovis inside host cells. This aspect, together with the role of Omp25d and Omp22 in the virulence both of B. ovis in rams and of other Brucella species, should be thoroughly evaluated in future studies.

Vibrio vulnificus is a bacterial species that is virulent for humans and fish. Human isolates are classified into biotypes 1 and 3 (BT1 and BT3) and fish isolates into biotype 2 (BT2). However, a few human infections caused by BT2 isolates have been reported worldwide (zoonosis). These BT2 human isolates belong to serovar E (SerE), which is also present in diseased fish. The aim of the present work was to characterize a new BT2 serovar [serovar A (SerA)], which emerged in the European fish-farming industry in 2000, by means of phenotypic, serological and genetic [plasmid profiling, ribotyping and random amplified polymorphic DNA (RAPD)] methodologies. The results confirmed that SerA constitutes a homogeneous O-serogroup within the species that shares plasmidic information with SerE. Like SerE, this new serogroup was resistant to fresh fish serum, as well as being highly virulent for fish. In contrast, it was sensitive to human serum and avirulent for mice, even after pretreatment with iron. The two serovars presented different biochemical profiles as well as specific patterns by ribotyping and RAPD analysis. In conclusion, SerA seems to constitute a different clonal group that has recently emerged within the species V. vulnificus, with pathogenic potential for fish but not for humans.

Following a large Neisseria meningitidis W135 (NmW135) epidemic in Burkina Faso (BF) during 2002, a newly licensed trivalent A/C/W135 meningococcal polysaccharide vaccine was introduced in 2003. We conducted a case-control study to assess the vaccine effectiveness (VE) against meningococcal disease. Thirty-two N. meningitidis A (NmA) and 3 NmW135 meningitis cases were enrolled and matched by age-neighborhood to 103 controls. After adjusting for confounding risk factors, VE against NmA or NmW135 was 83.6% (95% CI 31.8-97.0, p=0.01) for persons with verified vaccination. VE against probable/definite NmA alone was 94.0% (95% CI 58.7-99.0, p=0.0003). Low number of NmW135 cases did not allow estimation of VE against NmW135 alone. The vaccine was highly effective against the epidemic. Since 2003, the trivalent vaccine continues to be effectively used in Africa for the control of meningococcal disease epidemics.

Two poly(DL-lactide-co-glycolide) microsphere formulations (A, 10% wt/wt, and B, 23% wt/wt, 1-10 microns) were evaluated for intracellular delivery of rifabutin using the J774 murine and Mono Mac 6 (MM6) human monocytic cell lines. Within 7 days, formulation A released 100% in both cell lines and B released 53 and 67% in the J774 and MM6, respectively. Intracellular release of rifabutin with both formulations caused significant reduction of intracellularly replicating Mycobacterium avium (MAC). In MAC-infected beige mice, formulation B (50 mg, intraperitoneal days 0 and 7) completely eliminated infection by 21 days (p < 0.001), similar to a rifabutin daily oral regimen.

We studied the serum bactericidal activity (SBA) of moxifloxacin and levofloxacin against common pathogens associated with complicated intra-abdominal infections. Ten healthy volunteers received a single dose of moxifloxacin (400 mg) and levofloxacin (750 mg) and serum samples were collected at 2, 4, 8, 12, and 24h after the dose of each drug. Bactericidal titers in serum over time were determined for aerobic gram-negative bacilli (Escherichia coli, Klebseilla pneumoniae, and Enterobacter cloacae) and anaerobic bacteria (Bacteroides fragilis, Bacteroides thetaiotaomicron, Prevotella bivia, and Finegoldia magna). Both fluoroquinolones provided rapid (2h) attainment and prolonged (24h) SBA (titersor = 1:8) against each of the aerobic bacilli studied. SBA was observed for at least 12h against B. fragilis strains with MICsor = 2 microg/ml to moxifloxacin andor = 4 microg/ml to levofloxacin. Prolonged (12h) SBA (titersor = 1:2) was also observed against isolates of B. thetaiotaomicron, P. bivia, and F. magna with moxifloxacinor = MICs 2 microg/ml.

Urinary tract infections are frequently caused by Proteus mirabilis strains. In the previous studies there were defined the complete structures of O-polysaccharide parts of lipopolysaccharides from strains: P. mirabilis O3 (S1959), P. mirabilis O9 and P. mirabilis O18. In the present study it was investigated bactericidal effect of normal human serum (NHS) to P. mirabilis strains. We also focused on the diversity of outer membrane proteins (OMPs) being separated on a gel isolated from tested strains. Serial passage of P. mirabilis O18 in 90% normal bovine serum (NBS) contributed to over-expressing some classes of OMPs.

There is uniform consensus that flucytosine blood concentrations should be measured to avoid toxicity and ensure adequate efficacy.The purpose of this study was to evaluate all flucytosine levels performed in a regional centre in the UK from October 1991 to May 2006. Concentrations were measured by bioassay.We reviewed 1071 flucytosine levels in 233 patients, including 33 neonates. Overall, only 20.5% of levels were in the expected therapeutic range. Low levels were observed in 40.5%, of which 5.1% were undetectable levels (12.5 mg/L). High levels occurred in 38.9%, of which 9.9% were considered potentially toxic (100 mg/L). High flucytosine levels occurred more frequently amongst neonates, which could be related to an immature renal system resulting in drug accumulation.Our findings reveal that the vast majority of patients were out of range for flucytosine levels. These data emphasize the importance of monitoring flucytosine levels.

The purpose of this experiment was to evaluate the pharmacokinetics and serum bactericidal titers (SBTs) of daptomycin alone and in combination with gentamicin against strains of Staphylococcus aureus and enterococci to determine if there might be any benefit to the addition of the aminoglycoside. A multiple-dose, randomized crossover study was performed in 11 healthy volunteers to evaluate the steady-state pharmacokinetic profile of 6 mg/kg of body weight daptomycin once daily with or without 1 mg/kg gentamicin every 8 h. SBTs were determined against clinical isolates of nosocomial (MRSA 494) and community-acquired (CA-MRSA 44) methicillin-resistant S. aureus , vancomycin-susceptible Enterococcus faecalis (VSEF 49452), vancomycin-resistant Enterococcus faecium (VREF 80), and quality control strains of methicillin-susceptible S. aureus (ATCC 29213) and vancomycin-susceptible E. faecalis (ATCC 29212). Enhancement of bactericidal activity was evaluated by calculating and comparing the areas under the bactericidal curve (AUBC) for each dosing regimen against each isolate. The area under the concentration-time curve from 0 to 24 h and clearance for daptomycin alone were 645 ± 91 μg · h/ml and 9.47 ± 1.4 mg/h/kg, respectively, compared with 642 ± 69 μg · h/ml and 9.45 ± 1.0 mg/h/kg for daptomycin plus gentamicin. Daptomycin alone displayed sustained bactericidal activity against five of the six isolates over the entire 24-h dosing interval; bactericidal activity was maintained for 8 h against VREF 80. Mean AUBCs for daptomycin alone ranged from 935 to 1,263 and 36 to 238 against staphylococcal and enterococcal isolates, respectively, compared with 902 to 972 and 34 to 213 against staphylococci and enterococci when coadministered with gentamicin. The results of this study suggest that the addition of gentamicin does not alter the pharmacokinetic profile or enhance the bactericidal activity of daptomycin against staphylococcal or enterococcal isolates.

During June-August 2005, CDC and the Food and Drug Administration (FDA) were notified of seven clusters of inadvertent subcutaneous (SC) misadministration of the new meningococcal conjugate vaccine (MCV4, Menactra) (Sanofi Pasteur, Inc., Swiftwater, Pennsylvania), which is licensed for intramuscular (IM) administration only. A total of 101 persons in seven states were reported to have received MCV4 by the SC route. Of these, 100 were contacted by their health-care providers and advised of the administration error. CDC conducted an investigation to determine whether SC administration of MCV4 resulted in a protective immunologic response. This report describes the results of that investigation, which indicated that, despite the misadministration, persons vaccinated by the SC route were sufficiently protected and that revaccination was not necessary.

Since 1989, we have been involved in the development of a vaccine against Haemophilus influenzae type b. The new vaccine is based on the conjugation of synthetic oligosaccharides to tetanus toxoid. Our main goals have been (i) to verify the feasibility of using the synthetic antigen and (ii) to search for new production alternatives for this important infant vaccine. Overall, eight trials have already been conducted with adults, children (4 to 5 years old), and infants. We have described herein the details from the first two phase I clinical trials conducted with human adult volunteers under double blind, randomized conditions. The participants each received a single intramuscular injection to evaluate safety and initial immunogenicity. We have found an excellent safety profile and an antibody response similar to the one observed for the control vaccine.