bluuebunny/crossref_metadata_2025 huggingface dataset:
Low Doses of Caffeine: Enhancement of Physical Performance in Elite Adolescent Male Soccer Players
Authors: Ellis Matthew , Noon Mark , Myers Tony , Clarke Neil
Published: October 2018

Context: High doses of ∼6 mg·kg−1 body mass have improved performance during intermittent running, jumping, and agility protocols. However, there are sparse data on low doses of caffeine, especially in elite adolescent soccer players. Methods: A total of 15 elite youth soccer players (177.3 [4.8] cm, 66.9 [7.9] kg, and 16 [1] y) participated in the study, consuming 1, 2, or 3 mg·kg−1 caffeine in a gelatin capsule or a 2-mg·kg−1 placebo in a single-blind, randomized, crossover study design. Testing consisted of a 20-m sprint, arrowhead agility (change of direction [CoD] right or left), countermovement jump (CMJ), and Yo-Yo Intermittent Recovery Test Level 1 (Yo-Yo IR1). Postexercise CMJ performance was assessed as participants exited the Yo-Yo IR1. Data were analyzed using a Bayesian multilevel regression model to provide explained variance and probabilities of improvement (P = %). Results: Compared with placebo, 3 mg·kg−1 caffeine presented the highest probabilities of change across a range of tests (mean [SD], P = %). Times for 20-m sprint were 3.15 (0.10) s vs 3.18 (0.09) s (P = 73%), CoD-right times were 8.43 (0.24) s vs 8.55 (0.25) s (P = 99%), CoD-left times were 8.44 (0.22) s vs 8.52 (0.18) s (P = 85%), Yo-Yo IR1 distance was 2440 (531) m vs 2308 (540) m (P = 15%), and preexercise CMJ height was 41.6 (7.2) cm vs 38 (8.5) cm (P = 96%). Postexercise CMJ was higher with 3 mg·kg−1 than with placebo (42.3 [8] cm vs 36.6 [8] cm; P = 100%). Doses of 1 or 2 mg·kg−1 caffeine also demonstrated the ability to enhance performance but were task dependent. Conclusion: Low doses of caffeine improve performance but are dose and task dependent. A dose of 3 mg·kg−1 caffeine improved performance across the majority of tests with potential to further improve postexercise CMJ height.
Sequential Pulse Compression’s Effect on Blood Flow in the Lower-Extremity
Authors: Brock Kelly A. , Eberman Lindsey E. , Laird Richard H. , Elmer David J. , Games Kenneth E.
Published: October 2018

Context: Several interventions are available to reduce the intensity and duration of the unwanted effects (eg, muscle soreness) associated with physical activity, such as massage, compression garments, and sequential pulse compression (SPC). Such interventions aim to increase blood flow to alleviate symptoms. However, there is a lack of evidence to support the use of SPC to alter total hemoglobin concentration (THb) in active individuals. Objective: To examine the acute effects of a single session of SPC on hemoglobin concentration compared with a control condition. Design: Single cohort, crossover design. Participants: Thirty-four physically active and healthy participants (females = 12 and males = 22) completed the study. Interventions: The authors randomly assigned participants to first receive the experimental (SPC) or control condition. Measures were recorded precondition and postcondition. Participants returned to the laboratory to complete the second condition ≥24 hours after the first condition. Main Outcome Measures: Relative changes in THb, deoxygenated hemoglobin, and oxygenated hemoglobin measures were recorded using near-infrared spectroscopy placed on the muscle belly of the medial gastrocnemius of the dominant limb. Results: SPC significantly increased THb (P < .001, d = 0.505) and oxygenated hemoglobin (P < .001, d = 0.745) change scores compared with the control condition. No statistical difference in deoxygenated hemoglobin change scores was found between the SPC and control conditions, but a medium effect size suggests potential biological significance (P = .06, d = 0.339). Conclusions: Overall, SPC increases THb to the lower-extremity and may be a viable option in the management of muscle soreness related to physical activity.
Clinical and Biomechanical Efficacies of Mechanical Perturbation Training After Anterior Cruciate Ligament Rupture
Authors: Nawasreh Zakariya , Logerstedt David , Marmon Adam , Snyder-Mackler Lynn
Published: October 2018

Context: Manual perturbation training improves knee functional performance and mitigates abnormal gait in patients with anterior cruciate ligament (ACL) rupture. However, manual perturbation training is time- and labor-intensive for therapists. Objective: To investigate whether perturbation training administered using a mechanical device can provide effects similar to manual training on clinical measures and knee biomechanics after ACL rupture. Design: Prospective cohort (therapeutic) study. A 2 × 2 analysis of variance was used for statistical analysis. Setting: A clinical and biomechanical laboratory. Patients: Eighteen level I/II patients with acute ACL ruptures participated in this preliminary study. Intervention: Nine patients received mechanical perturbation training on an automated mechanical device (mechanical group), and 9 patients received manual perturbation training (manual group). Outcome Measures: Patients completed performance-based testing (quadriceps strength and single-legged hop tests), patient-reported questionnaires (Knee Outcome Survey-Activities of Daily Living Scale, Global Rating Score, and International Knee Documentation Committee 2000), and 3-dimensional gait analysis before (pretesting) and after (posttesting) training. Results: There was no significant group-by-time interaction found for all measures (P ≥ .18). Main effects of time were found for International Knee Documentation Committee 2000 (pretesting: 69.10 [10.95], posttesting: 75.14 [7.19]), knee excursion during weight-acceptance (pretesting: 16.01° [3.99°]; posttesting: 17.28° [3.99°]) and midstance (pretesting: 14.78° [4.13°]; posttesting: 16.92° [4.53°]) and external knee-flexion moment (pretesting: 0.43 [0.11] N m/kg/m; posttesting: 0.48 [0.11] N m/kg/m) (P ≤ .04). After accounting for pretesting groups’ differences, the mechanical group scored significantly higher on triple hops (mechanical: 96.73% [6.65%]; manual: 84.97% [6.83%]) and 6-m timed hops (mechanical: 102.07% [9.50%]; manual: 91.21 [9.42%]) (P ≤ .047) compared with manual group. Conclusion: The clinical significance of this study is the mechanical perturbation training produced effects similar to manual training, with both training methods were equally  effective at improving patients’ perception of knee function and increasing knee excursion and external flexion moment during walking after acute ACL rupture. Mechanical perturbation training is a potential treatment to improve patients’ functional and biomechanical outcomes after ACL rupture.
IL-17 integrates multiple self-reinforcing, feed-forward mechanisms through the RNA binding protein Arid5a
Authors: Amatya Nilesh , Childs Erin E. , Cruz J. Agustin , Aggor Felix E. Y. , Garg Abhishek V. , Berman Andrea J. , Gudjonsson Johann E. , Atasoy Ulus , Gaffen Sarah L.
Published: October 2018

The RNA binding protein Arid5a augments both mRNA stability and translation to promote cellular responses to IL-17.
Effects of Food and Omeprazole on a Novel Formulation of Super Bioavailability Itraconazole in Healthy Subjects
Authors: Lindsay Julian , Mudge Stuart , Thompson George R.
Published: October 2018

To address the limited bioavailability and intolerance of the conventional itraconazole (ITZ) formulations, a new formulation labeledsuperbioavailability (SUBA) itraconazole has been developed; however, the specific effects of food and gastric pH are unknown. This study evaluated the pharmacokinetic profile of SUBA itraconazole under fasting and fed conditions, as well as with the concomitant administration of a proton pump inhibitor.
Serum bactericidal activities and comparative pharmacokinetics of meropenem and imipenem-cilastatin
Authors: Dreetz M , Hamacher J , Eller J , Borner K , Koeppe P , Schaberg T , Lode H
Published: October 2018

The pharmacokinetics and serum bactericidal activities (SBAs) of imipenem and meropenem were investigated in a randomized crossover study. Twelve healthy male volunteers received a constant 30-min infusion of either 1 g of imipenem plus 1 g of cilastatin or 1 g of meropenem. The concentrations of the drugs in serum and urine were determined by bioassay and high-pressure liquid chromatography. Pharmacokinetic parameters were based on an open two-compartment model and a noncompartmental technique. At the end of infusion, the mean concentrations of imipenem and meropenem measured in serum were 61.2 +/- 9.8 and 51.6 +/- 6.5 mg/liter, respectively; urinary recoveries were 48.6% +/- 8.2% and 60.0% +/- 6.5% of the dose in 12 h, respectively; and the areas under the concentration-time curve from time zero to infinity were 96.1 +/- 14.4 and 70.5 +/- 10.3 mg.h/liter, respectively (P < or = 0.02). Imipenem had a mean half-life of 66.7 +/- 10.4 min; that of meropenem was 64.4 +/- 6.9 min. The volumes of distribution at steady state of imipenem and meropenem were 15.3 +/- 3.3 and 18.6 +/- 3.0 liters/70 kg, respectively, and the mean renal clearances per 1.73 m2 were 85.6 +/- 17.6 and 144.6 +/- 26.0 ml/min, respectively. Both antibiotics were well tolerated in this single-dose administration study. The SBAs were measured by the microdilution method of Reller and Stratton (L. B. Reller and C. W. Stratton, J. Infect. Dis. 136:196-204, 1977) against 40 clinically isolated strains. Mean reciprocal bactericidal titers were measured 1 and 6 h after administration. After 1 and 6 h the median SBAs for imipenem and meropenem, were 409 and 34.9 and 97.9 and 5.8, respectively, against Staphylococcus aureus, 19.9 and 4.4 and 19.4 and 4.8, respectively, against Pseudomonas aeruginosa, 34.3 and 2.2 and 232 and 15.5, respectively, against Enterobacter cloacae, and 13.4 and 2.25 and 90.7 and 7.9, respectively, against Proteus mirabilis. Both drugs had rather short biological elimination half-lives and a predominantly renal route of elimination. Both carbapenems revealed high SBAs against clinically important pathogens at 1 h; meropenem had a higher SBA against E. cloacae and P. mirabilis, and the SBA of imipenem against S. aureus was greater than the SBA of meropenem.
Clinical pharmacokinetics of piperacillin-tazobactam combination in patients with major burns and signs of infection
Authors: Bourget P , Lesne-Hulin A , Le Reveillé R , Le Bever H , Carsin H
Published: October 2018

The pathophysiology associated with major burns is complex and subject to a state of flux. The combination of beta-lactamase inhibitors with powerful penicillins is an interesting and an attractive potential solution to the emergence of bacterial resistance. The kinetics in serum and urine and the clinical safety of a fixed combination of 4 g of piperacillin (PPR) and 0.5 g of tazobactam (TZB) were studied in 10 patients (22 to 50 years old and weighing 45 to 105 kg) with major burns who were infected with Pseudomonas aeruginosa and various entero-bacteria. All of them received additional antimicrobial drugs. Treatment involved one dose every 6 h. The mean body surface area affected by third-degree burns was 30.0% +/- 4.0%. The study took place in accordance with current ethical guidelines. Two series of blood samples were drawn after the first (day 1) and ninth (day 3 at steady state) doses; urine was collected during the same periods. Levels of PPR and TZB in serum and urine were measured by high-pressure liquid chromatography. A noncompartmental method was used for kinetic and graphic analysis of concentration-time pairs. The safety of the treatment was excellent. There was no systemic accumulation of the beta-lactam combination. Residual concentrations measured on days 1 and 3 [mean (standard error of the mean)] were above the MIC for the organism responsible for infection; i.e., C(min)day1 = 26.3 (8.5) and C(min)day3 = 21.0 (9.1) for PPR and C(min)day1 = 1.9 (0.6) and C(min)day3 = 1.4 (0.3) for TZB. There was no statistically significant difference between pharmacokinetic parameters determined for day 1 and day 3. Evidence was found in burn patients, in contrast to healthy subjects, of a marked increase in apparent volumes of distribution, in such a way that the apparent elimination half-lives of the combination were notably prolonged, i.e., 1.8 (0.3) versus 1.5 (0.3) h for PPR in patients and healthy subjects, respectively, and 1.7 (0.3) versus 1.4 (0.3) h for TZB. These findings indicate the possibility of nonrenal translesional diffusion of PPR-TZB in burn patients. The polarity of the association would further support this hypothesis. It has been shown here that the recommended dosage regimen for administration of PPR-TZB must be high in major-burn patients, i.e., 4 g/0.5 g every 6 h. The data obtained provide valuable information, which is suitable for immediate application in everyday clinical practice.
Activity of pyrazinamide in a murine model against Mycobacterium tuberculosis isolates with various levels of in vitro susceptibility
Authors: Klemens S P , Sharpe C A , Cynamon M H
Published: October 2018

The activity of pyrazinamide (PZA) against eight isolates of Mycobacterium tuberculosis in a murine infection model was evaluated. M. tuberculosis isolates with various degrees of in vitro susceptibility to PZA (MIC range, 32 to > 2,048 micrograms/ml) were used. Four-week-old female mice were infected intravenously with approximately 10(7) viable M. tuberculosis organisms. PZA at 150 mg/kg of body weight was started 1 day postinfection and given 5 days/week for 4 weeks. Infected but untreated mice were compared with PZA-treated mice. Mice were sacrificed at the completion of the treatment period, and viable cell counts were determined from homogenates of spleens and right lungs. PZA had activity in the murine test system against M. tuberculosis isolates for which the MICs were < or = 256 micrograms/ml. However, there was an inconsistent correlation between the absolute MICs and the reductions in organ viable cell counts. Studies with drug-resistant M. tuberculosis isolates with an isogenic background would improve evaluation of drug efficacy in the murine test system. Further evaluation of antimycobacterial agents against monodrug-resistant isolates will provide data that will be useful for development of algorithms for treatment of infection with drug-resistant organisms.
Effects of cytokines on antiviral pharmacokinetics: an alternative approach to assessment of drug interactions using bioequivalence guidelines
Authors: Piscitelli S C , Amatea M A , Vogel S , Bechtel C , Metcalf J A , Kovacs J A
Published: October 2018

The effects of cytokines on the pharmacokinetics of nucleoside analogs were evaluated in two separate studies using zidovudine in combination with interleukin-2 and didanosine in combination with alpha interferon. In each study, drug interactions were evaluated by using both a standard method (Student's t test) and bioequivalence testing. Serial blood samples were collected from human immunodeficiency virus-infected patients prior to and during cytokine therapy for determination of nucleoside analog concentrations. Concentrations were fit separately to a two-compartment model by using the iterative two-stage approach to population analysis. No alterations in area under the curve or oral clearance were observed for either drug during combination therapy. In general, there was good agreement between statistical methods for determining if antiviral pharmacokinetic parameters were altered by concomitant cytokine therapy. However, large individual changes in the maximum concentration of zidovudine in serum were detected by bioequivalence testing but no difference was found by Student's t test. For didanosine, significant but clinically irrelevant decreases determined by standard hypothesis testing were seen for both the volume of the central compartment (1.91 to 1.86 liters) and the absorption rate constant (0.79 to 0.73 h-1) in the presence of alpha interferon. No interaction was noted for these parameters by using bioequivalence guidelines. Bioequivalence testing may provide an alternative approach to assessment of drug interactions. Interleukin-2 and alpha interferon do not alter the pharmacokinetics of zidovudine and didanosine, respectively.
Overproduction of a 37-kilodalton cytoplasmic protein homologous to NAD+-linked D-lactate dehydrogenase associated with vancomycin resistance in Staphylococcus aureus
Authors: Milewski W M , Boyle-Vavra S , Moreira B , Ebert C C , Daum R S
Published: October 2018

We previously reported the isolation of a laboratory-derived Staphylococcus aureus mutant, 523k, that has constitutive low-level resistance to vancomycin (MIC = 5 micrograms/ml) and teicoplanin (MIC = 5 micrograms/ml) and elaborates a ca. 39-kDa cytoplasmic protein that was not detected in the parent strain 523 (MIC = 1 micrograms/ml). We have now detected the protein in strain 523 by immunoblotting with antiserum raised against the protein. Consistent with our initial observations, densitometric analysis of the immunoblots revealed an increased production of the protein in 523k compared with that of the susceptible parent 523. The 5' region of the gene encoding the protein of interest was identified by nucleotide sequencing a PCR product amplified from the genome of 523k with degenerate primers designed to encode the amino acid sequence of proteolytic peptides obtained from the protein. The remainder of the gene was identified by library screening, PCR, and nucleotide sequencing. The gene encodes a 36.7-kDa protein with homology to a family of bacterial NAD+-dependent, D-specific 2-hydroxyacid dehydrogenases which includes both D-lactate dehydrogenase and the enterococcal vancomycin resistance protein VanH and is therefore designated ddh. Increased production of the product of ddh, Ddh, was associated with increased D-lactate dehydrogenase activity in 523k, a finding which suggested that Ddh is likely to be the D-lactate dehydrogenase previously identified in S. aureus. The increased D-lactate dehydrogenase activity in strain 523k and the structural similarities among Ddh, D-lactate dehydrogenase, and VanH suggest that overproduction of Ddh might play a role in vancomycin resistance in this strain.
Protection and rescue from 2',3'-dideoxypyrimidine nucleoside analog toxicity by hemin in human bone marrow progenitor cells
Authors: Fowler D A , Xie M Y , Sommadossi J P
Published: October 2018

Long-term therapy of AIDS patients with 3'-azido-3'-deoxythymidine (AZT) remains of concern because of resulting hematopoietic toxicity. While the mechanism(s) of this toxicity remains elusive, alternative strategies are being developed to reduce these toxic effects, including combination therapy with nonmyelotoxic antihuman immunodeficiency virus drugs and/or administration of protective or rescue agents, including cytokines and growth factors. By using a particularly relevant human CD34+ liquid culture system, the unique profiles of dideoxynucleoside (ddN) toxicities to both proliferation and differentiation were demonstrated, with decreased potencies in the order of 3'-fluoro-3'-deoxythymidine (FLT) = 3'-amino-3'-deoxythymidine (AMT) = 2',3'-dideoxycytidine > AZT for inhibition of proliferation and in the order of FLT = AMT > AZT > 2',3'-dideoxycytidine for inhibition of hemoglobin synthesis. Hemin selectively protected erythroid-lineage human burst-forming unit-erythroid cells from AZT- and AMT-induced inhibition but had no effect on FLT toxicity under similar conditions. Myeloid-lineage human CFU-granulocyte-macrophages were also not protected by hemin against all three ddN analogs. The simultaneous exposure of cells to hemin and AZT resulted in a complete protection of both cell proliferation and hemoglobin synthesis. In contrast, in reversal studies only the inhibition of the percentage of hemoglobin-synthesizing cells returned to control levels, but the inhibition of proliferation of cells previously exposed to AZT was not reversed by hemin. These studies further define the unique and multifactorial mechanism(s) of ddN-induced toxic effects during hematopoietic development of pluripotent stem cells and suggest that the use of hemin could be beneficial in alleviating the toxicity of certain ddN analogs.
In vivo maternal-fetal pharmacokinetics of stavudine (2',3'-didehydro-3'-deoxythymidine) in pigtailed macaques (Macaca nemestrina)
Authors: Odinecs A , Nosbisch C , Keller R D , Baughman W L , Unadkat J D
Published: October 2018

To determine whether stavudine (2',3'-didehydro-3'-deoxythymidine) is actively transported in vivo across the placenta and to determine the extent of its transfer, stavudine was administered as an intravenous bolus to four near-term macaques (Macaca nemestrina) (5 mg/kg of body weight via the femoral vein) or to their fetuses (10 mg/kg via the carorid artery) at gestational age 134 +/- 5 days, with the administrations about 1 week apart. Antipyrine (a passive diffusion marker) was always coadministered (20 mg/kg) with stavudine. Samples of maternal and fetal plasma and amniotic fluid were collected at frequent intervals up to 240 min after the dose. In a separate experiment, three animals received stavudine for 30 h at a low rate of infusion (22 micrograms/min/kg via the femoral vein) to the dam or at a 10-fold-higher rate of infusion (220 micrograms/min/kg), separated by at least one week, in order to determine if the transplacental transfer of stavudine is saturable. Antipyrine (41.7 micrograms/min/kg) was coinfused with stavudine. Samples of maternal and fetal plasma and amniotic fluid were collected at regular intervals for up to 30 h. The concentrations of stavudine and antipyrine were determined by high-performance liquid chromatography. The transplacental maternal-fetal drug clearances were compared by the paired Student t test. The clearance associated with maternal-fetal transfer of the drug (CLdf) (0.54 +/- 0.08 ml/min/kg) was not significantly different (P > 0.05) from the clearance associated with fetal-maternal transfer of the drug, CLfd (0.66 +/- 0.11 ml/min/kg). Also, CLdf was not significantly different (P > 0.05) from CLfd when normalized with respect to the corresponding transplacental clearance of antipyrine (0.23 +/- 0.04 versus 0.36 +/- 0.25). The ratios of the steady-state plasma stavudine concentration in the fetus to that in the dam were 0.77 +/- 0.06 at the low stavudine infusion rate and 0.81 +/- 0.09 at the high stavudine infusion rate. The obtained data indicate that transfer of stavudine across the placenta is passive and constant over the dose range studied.
Tandem duplication in ermC translational attenuator of the macrolide-lincosamide-streptogramin B resistance plasmid pSES6 from Staphylococcus equorum
Authors: Lodder G , Schwarz S , Gregory P , Dyke K
Published: October 2018

A tandem duplication of 23 bp in the ermC gene translational attenuator of plasmid pSES6 from Staphylococcus equorum which mediated constitutive resistance to macrolide-lincosamide-streptogramin B antibiotics was identified. This duplication included the ribosome binding site for the ermC gene as well as the first 5 bp of the ermC coding sequence. It was postulated that this sequence duplication affects the possible RNA conformations so that the ribosome binding site for ErmC synthesis is readily accessible to the ribosomes and thus constitutive expression of the ermC gene occurs.
Antiretroviral activity and pharmacokinetics in mice of oral bis(pivaloyloxymethyl)-9-(2-phosphonylmethoxyethyl)adenine, the bis(pivaloyloxymethyl) ester prodrug of 9-(2-phosphonylmethoxyethyl)adenine
Authors: Naesens L , Balzarini J , Bischofberger N , De Clercq E
Published: October 2018

Lipophilic ester prodrugs of 9-(2-phosphonylmethoxyethyl)adenine (PMEA), i.e., bis(pivaloyloxymethyl)-PMEA [bis(POM)-PMEA] and diphenyl-PMEA, have been synthesized in an attempt to increase the oral bioavailability of this broad-spectrum antiviral agent. The antiretroviral efficacy was determined in severe combined immune deficiency (SCID) mice infected with Moloney murine sarcoma virus (MSV). They were treated twice daily for 5 days after infection. Oral treatment with bis(POM)-PMEA at a dose equivalent to 100 or 50 mg of PMEA per kg of body weight per day proved markedly effective in delaying MSV-induced tumor formation and death of the mice. Oral bis(POM)-PMEA afforded anti-MSV efficacy equal to that of subcutaneous PMEA given at equimolar doses. Oral treatment with PMEA or diphenyl-PMEA proved less efficient. Similarly, in mice infected with Friend leukemia virus (FLV), oral treatment with bis(POM)-PMEA at a dose equivalent to 100 or 50 mg of PMEA per kg per day effected a marked inhibition of FLV-induced splenomegaly (87 and 48% inhibition, respectively), the efficacy being equal to that of PMEA given subcutaneously at equivalent doses. Pharmacokinetic experiments with mice showed that the oral bioavailabilities of PMEA following oral gavage of bis(POM)-PMEA, diphenyl-PMEA, or PMEA (at a dose equivalent to 50 mg of PMEA per kg) were 53,3, and 16%, respectively. These data were calculated from the levels of free PMEA in plasma. Also, the recoveries of free PMEA in the urine upon oral administration of bis(POM)-PMEA, diphenyl-PMEA, or PMEA (at a dose equivalent to 25 mg of PMEA per kg) were 48, 4, and 7%, respectively. Oral bis(POM)-PMEA was not recovered from plasma, suggesting that it was readily cleaved to free PMEA. In contrast, diphenyl-PMEA was not efficiently cleaved to free PMEA, resulting in a rather low oral bioavailability of PMEA from this prodrug. Bis(POM)-PMEA appears to be an efficient oral prodrug of PMEA that deserves further clinical evaluation in human immunodeficiency virus-infected individuals.
Penicillin tolerance and modification of lipoteichoic acid associated with expression of vancomycin resistance in VanB-type Enterococcus faecium D366
Authors: Gutmann L , Al-Obeid S , Billot-Klein D , Ebnet E , Fischer W
Published: October 2018

Induction of vancomycin resistance in Enterococcus faecium D366, which exhibits a VanB-type resistance, as well as its constitutive expression in MT9, a derivative of D366, was associated with penicillin tolerance as shown by decreased lysis and killing of the cells. This phenomenon was linked neither to decreased expression of the different autolysins nor to their decreased lytic activity on the different cell walls. The only change observed was that almost twice the normal amount of D-alanine was attached to the lipoteichoic acid.
Simple in vitro translation assay to analyze inhibitors of rhinovirus proteases
Authors: Heinz B A , Tang J , Labus J M , Chadwell F W , Kaldor S W , Hammond M
Published: October 2018

We have developed a simple in vitro translation method to analyze compounds that inhibit the rhinovirus 3C protease in peptide substrate assays but demonstrate no antiviral activity. This complementary assay, which provides both qualitative and quantitative results, detects the inhibition of the 3CD protease in the native polyprotein form.
Influence of pH on adaptive resistance of Pseudomonas aeruginosa to aminoglycosides and their postantibiotic effects
Authors: Xiong Y Q , Caillon J , Drugeon H , Potel G , Baron D
Published: October 2018

Adaptive resistance to aminoglycosides in Pseudomonas aeruginosa and other gram-negative bacilli is usually induced by the initial exposure to the drug. We investigated the influence of pH on the adaptive resistance of a clinical P. aeruginosa strain to aminoglycosides in vitro and on their postantibiotic effects. For adaptive resistance, the first-exposure concentrations of both amikacin and netilmicin were one, two, four, and eight times the MIC of each drug and the second-exposure concentrations were two times the MIC of each drug. Adaptive resistance was greater and more prolonged with higher initial aminoglycoside concentrations, and the bactericidal effects of the aminoglycosides were concentration dependent at pH 7.4. At pH 6.5, the killing rates of amikacin and netilmicin were far lower than those observed at pH 7.4. At pH 5.5, amikacin and netilmicin exerted practically no bactericidal effect on the P. aeruginosa strain used. However, with media at pH 5.5 and 6.5, adaptive resistance of P. aeruginosa preexposed to amikacin and netilmicin was also clearly exhibited, with the degree of adaptive resistance depending on the bactericidal effects of both drugs on nonpreexposed controls. Maximal adaptive resistance occurred between 0 and 4 h after preexposure. The postantibiotic effects of amikacin and netilmicin against the P. aeruginosa strain were shown to be concentration dependent and were reduced at acidic pHs. No changes in outer and inner membrane proteins occurred during the adaptive-resistance interval.
ParC subunit of DNA topoisomerase IV of Streptococcus pneumoniae is a primary target of fluoroquinolones and cooperates with DNA gyrase A subunit in forming resistance phenotype
Authors: Muñoz R , De La Campa A G
Published: October 2018

The genes encoding the ParC and ParE subunits of topoisomerase IV of Streptococcus pneumoniae, together with the region encoding amino acids 46 to 172 (residue numbers are as in Escherichia coli) of the pneumococcal GyrA subunit, were partially characterized. The gyrA gene maps to a physical location distant from the gyrB and parC loci on the chromosome, whereas parC is closely linked to parE. Ciprofloxacin-resistant (Cpr) clinical isolates of S. pneumoniae had mutations affecting amino acid residues of the quinolone resistance-determining region of ParC (low-level Cpr) or in both quinolone resistance-determining regions of ParC and GyrA (high-level Cpr). Mutations were found in residue positions equivalent to the serine at position 83 and the aspartic acid at position 87 of the E. coli GyrA subunit. Transformation experiments suggest that ParC is the primary target of ciprofloxacin. Mutation in parC appears to be a prerequisite before mutations in gyrA can influence resistance levels.
Pharmacokinetics of a water-soluble fullerene in rats
Authors: Rajagopalan P , Wudl F , Schinazi R F , Boudinot F D
Published: October 2018

Fullerenes are the recently discovered third allotropic form of carbon. The biological activities of these compounds are being studied for various purposes. The bis(monosuccinimide) derivative of p p'-bis(2-amino-ethyl)-diphenyl-C60 (MSAD-C60) is a water-soluble fullerene derivative. MSAD-C60 has been shown to have antiviral activity against human immunodeficiency virus types 1 and 2 in vitro and to have virucidal and anti-human immunodeficiency virus protease activities. Moreover, MSAD-C60 has been shown to be well tolerated in mice after intraperitoneal administration. The purpose of the present study was to develop a high-performance liquid chromatographic analytical methodology for MSAD-C60 and to characterize the preclinical pharmacokinetics of the compound in rats. Following intravenous administration of the fullerene derivative at a dose of 15 mg/kg of body weight, the concentrations of MSAD-C60 in plasma declined either bi- or triexponentially. The mean terminal-phase half-life of MSAD-C60 was 6.8 +/- 1.1 h (mean +/- standard deviation). Binding studies indicated that the compound is greater than 99% bound to plasma proteins. The average total clearance of the compound was 0.19 +/- 0.06 liter/h/kg. Urine samples obtained 24 h after intravenous administration did not contain detectable levels of the compound, indicating the absence of a significant renal clearance mechanism. The steady-state volume of distribution of MSAD-C60 averaged 2.1 +/- 0.8 liters/kg, indicating that the compound distributes into tissues. At a dose of 15 mg/kg, MSAD-C60 appeared to be well tolerated. However, a dose of 25 mg/kg resulted in shortness of breath and violent movement of the rats, followed by death within 5 min of dosing. Further controlled toxicity studies are needed to fully evaluate the toxicity of the compound.
Direct evidence for antipseudomonal activity of macrolides: exposure-dependent bactericidal activity and inhibition of protein synthesis by erythromycin, clarithromycin, and azithromycin
Authors: Tateda K , Ishii Y , Matsumoto T , Furuya N , Nagashima M , Matsunaga T , Ohno A , Miyazaki S , Yamaguchi K
Published: October 2018

Several previous investigators have reported that long-term administration of certain macrolides is efficacious in patients with persistent pulmonary Pseudomonas aeruginosa infections, even though the clinically achievable concentrations of these medications are far below their MICs. In the present study, we examined how sub-MICs of macrolide antibiotics affect the viability of and protein synthesis in several strains of P. aeruginosa. We report that 48 h, but not 12 or 24 h, of growth on agar containing a clinically achievable concentration of azithromycin (0.5 microgram/ml, 1/128 the MIC) significantly reduces the viability of strain PAO-1. Similar effects were seen with erythromycin and clarithromycin at 2 micrograms/ml (1/128 and 1/64 the respective MICs), whereas josamycin, oleandomycin, ceftazidime, tobramycin, minocycline, and ofloxacin had no effect on viability, even following 48 h of incubation with concentrations representing relatively high fractions of their MICs. The bactericidal activity of azithromycin seen following 48 h of incubation was not limited to strain PAO-1 but was also seen against 13 of 14 clinical isolates, including both mucoid and nonmucoid strains. Although viability was not decreased prior to 48 h, we found that 4 micrograms of azithromycin per ml inhibits protein synthesis after as little as 12 h and that protein synthesis continues to decrease in a time-dependent manner. We likewise found that P. aeruginosa accumulates azithromycin intracellulary over the period from 12 to 36 h. These results suggested that sub-MICs of certain macrolides are bactericidal to P. aeruginosa when the bacteria are exposed to these antibiotics for longer periods. Exposure-dependent intracellular accumulation of the antibiotic and inhibition of protein synthesis may partially account for the antipseudomonal activity of macrolides over relatively prolonged incubation periods.
Multidrug efflux in intrinsic resistance to trimethoprim and sulfamethoxazole in Pseudomonas aeruginosa
Authors: Köhler T , Kok M , Michea-Hamzehpour M , Plesiat P , Gotoh N , Nishino T , Curty L K , Pechere J C
Published: October 2018

Pseudomonas aeruginosa possesses at least two multiple drug efflux systems which are defined by the outer membrane proteins OprM and OprJ. We have found that mutants overexpressing OprM were two- and eightfold more resistant than their wild-type parent to sulfamethoxazole (SMX) and trimethoprim (TMP), respectively. For OprJ-overproducing strains, MICs of TMP increased fourfold but those of SMX were unchanged. Strains overexpressing OprM, but not those overexpressing OprJ, became hypersusceptible to TMP and SMX when oprM was inactivated. The wild-type antibiotic profile could be restored in an oprM mutant by transcomplementation with the cloned oprM gene. These results demonstrate that the mexABoprM multidrug efflux system is mainly responsible for the intrinsic resistance of P. aeruginosa to TMP and SMX.
In vitro inhibition of hepatitis C virus gene expression by chemically modified antisense oligodeoxynucleotides
Authors: Vidalin O , Major M E , Rayner B , Imbach J L , Trépo C , Inchauspé G
Published: October 2018

We have explored different domains within the hepatitis C virus (HCV) 5' noncoding region as potential targets for inhibition of HCV translation by antisense oligodeoxynucleotides (ODNs). Inhibition assays were performed with two different cell-free systems, rabbit reticulocyte lysate and wheat germ extract, and three types of chemical structures for the ODNs were evaluated: natural phosphodiesters (beta-PO), alpha-anomer phosphodiesters (alpha-PO), and phosphorothioates (PS). A total of six original ODNs, displaying sequence-specific inhibition ranging from 62 to 96%, that mapped in the pyrimidine-rich tract (nucleotides [nt] 104 to 127) and in the initiator AUG codon (nt 338 to 357) were identified. Two ODNs, which were targeted at the initiatory AUG (nt 341 to 367 and 351 to 377) and which had been previously described as active against genotype 1b and 2a sequences, were shown to exhibit inhibition of expression (> 95%) of a type 1a sequence. Control experiments with the irrelevant chloramphenicol acetyltransferase sequence as a marker and randomized ODNs demonstrated that levels of inhibition associated with the use of PS compounds (of as much as 94%) were mainly due to nonspecific effects. Both alpha- and beta-PO ODNs were found equally active, and no difference could be seen in the activity of beta-PO when it was tested in either rabbit reticulocyte lysate or wheat germ extract, suggesting that RNase H-independent mechanisms may be involved in the inhibitions observed. However, specific RNA cleavage products generated from beta-PO inhibition experiments could be identified, indicating that, with these compounds, control of translation also involves RNase H-dependent mechanisms. This study further delimits the existence of favorable target sequences for the action of ODNs within the HCV 5' noncoding region and indicates the possibility of using nuclease-resistant alpha-PO compounds in cellular studies.
Manipulation of the N-alkyl substituent in amodiaquine to overcome the verapamil-sensitive chloroquine resistance component
Authors: Hawley S R , Bray P G , O'Neill P M , Naisbitt D J , Park B K , Ward S A
Published: October 2018

Aminoquinoline resistance correlates with lipid solubility at pH 7.2. Consequently, the in vivo dealkylation of amodiaquine, to the less lipid-soluble desethylamodiaquine, is a likely contributor to therapeutic failure in vivo. Therefore, 4-aminoquinoline drugs with lipid solubilities similar to that of amodiaquine, but which are not subject to side chain modification in vivo, should be superior antimalarial agents. In this study, we have identified amopyroquine and N-tertbutylamodiaquine as two such compounds. The values for the logarithms of the partition coefficients for amopyroquine and N-tertbutylamodiaquine are between those for amodiaquine and its dealkylated metabolite, desethylamodiaquine. Both amopyroquine and N-tertbutylamodiaquine possess levels of antimalarial activity greater than that of desethylamodiaquine and significantly reduced cross-resistance patterns; i.e., the former two compounds are not subject to the verapamil-sensitive resistance mechanism. Simple in vitro markers of direct toxicity and potential reactive metabolite formation suggest that these two compounds are no more toxic than amodiaquine and desethylamodiaquine.
Characterization of the chromosomal aminoglycoside 2'-N-acetyltransferase gene from Mycobacterium fortuitum
Authors: Aínsa J A , Martin C , Gicquel B , Gomez-Lus R
Published: October 2018

A novel gene encoding an aminoglycoside 2'-N-acetyltransferase (AAC) was cloned from Mycobacterium fortuitum. DNA sequencing results identified an open reading frame that we have called aac(2')-Ib encoding a putative protein with a predicted molecular mass of 24,800 Da. The deduced AAC(2')-Ib protein showed homology to the AAC(2')-Ia from Providencia stuartii. This is the second member of a subfamily of AAC(2')-I enzymes to be identified. No homology was found with other acetyltransferases, including all of the AAC(3) and AAC(6') proteins. The aac(2')-Ib gene cloned in a mycobacterial plasmid and introduced in Mycobacterium smegmatis conferred resistance to gentamicin, tobramycin, dibekacin, netilmicin, and 6'-N-ethylnetilmicin. DNA hybridization with an intragenic probe of aac(2')-Ib showed that this gene was present in all 34 strains of M. fortuitum tested. The universal presence of the aac(2')-Ib gene in M. fortuitum was not correlated with any aminoglycoside resistance phenotype, suggesting that this gene may play a role in the secondary metabolism of the bacterium.
Structure-activity relationships of quinolone agents against mycobacteria: effect of structural modifications at the 8 position
Authors: Renau T E , Gage J W , Dever J A , Roland G E , Joannides E T , Shapiro M A , Sanchez J P , Gracheck S J , Domagala J M , Jacobs M R , Reynolds R C
Published: October 2018

A series of quinolones with substitutions at the 8 position has been prepared as part of a study to examine the relationship between structural modifications at this position and activity against mycobacteria. The compounds were prepared by procedures described in the literature and were evaluated for their activities against Mycobacterium fortuitum and Mycobacterium smegmatis. The activities of the compounds against these two organisms were used as a measure of Mycobacterium tuberculosis activity. The results demonstrate that the contribution of the 8 position to antimycobacterial activity was dependent on the substituent at N-1 and was in the order (i) COMe approximately CBr > CCI > CH approximately CF approximately COEt > N > CCF3 when N-1 was cyclopropyl; (ii) N approximately CH > CF > COMe when N-1 was 2,4-difluorophenyl; (iii) N > or = CH when N-1 was tert-butyl; and (iv) N > CH when N-1 was ethyl. In general, derivatives with piperazine substitutions at C-7 were slightly less active against mycobacteria than the analogs with pyrrolidine substitutions, regardless of the pattern of substitution at the 8 position. Several of the best compounds were evaluated for their potential side effects as well as their activities against Mycobacterium aurum, Mycobacterium avium-M. intracellulare, and M. tuberculosis. These agents exhibited biological profiles similar to or better than those of the positive controls ciprofloxacin and sparfloxacin.
Potent activity of 2'-beta-fluoro-2',3'-dideoxyadenosine against human immunodeficiency virus type 1 infection in hu-PBL-SCID mice
Authors: Ruxrungtham K , Boone E , Ford H , Driscoll J S , Davey R T , Lane H C
Published: October 2018

A new antiretroviral agent, 2'-beta-fluoro-2',3'-dideoxyadenosine (FddA), is an acid-stable compound whose triphosphate form is a potent reverse transcriptase inhibitor with in vitro anti-human immunodeficiency virus (HIV) activity and a favorable pharmacokinetic profile. Severe combined immunodeficiency (SCID) mice reconstituted with human peripheral blood leukocytes (hu-PBL-SCID mice) provide a useful small-animal model for HIV research. In the present study we utilized this experimental system for the in vivo evaluation of the anti-HIV activity of this new compound when administered prior to infection. Initial studies revealed that, following a challenge with 50 100% tissue culture infective doses of HIV type 1 lymphadenopathy-associated virus, 39 of 42 (93%) control mice developed HIV infection, as evidenced by positive coculture or positive PCR. Administration of zidovudine decreased the infection rate to 5 of 16 (31%), while administration of FddA decreased the infection rate to 0 of 44 (0%). In follow-up controlled studies, the anti-HIV activity of FddA was confirmed, with 18 of 20 control mice showing evidence of HIV infection, compared with 4 of 20 FddA-treated mice. In addition to having direct anti-HIV effects, FddA was found to have a protective effect on human CD4+ T cells in the face of HIV infection. Mice treated with FddA were found to have a significantly higher percentage of CD4+ T cells than controls (10.3% +/- 3.4% versus 0.27% +/- 0.21%; P = 0.01). Thus, FddA, with its potent anti-HIV activity in vivo, high oral bioavailability, long intracellular half-life, and ability to preserve CD4+ cells in the presence of HIV, appears to be a promising agent for clinical investigation.
Contributions of individual mechanisms to fluoroquinolone resistance in 36 Escherichia coli strains isolated from humans and animals
Authors: Everett M J , Jin Y F , Ricci V , Piddock L J
Published: October 2018

Twenty-eight human isolates of Escherichia coli from Argentina and Spain and eight veterinary isolates received from the Ministry of Agriculture Fisheries and Foods in the United Kingdom required 2 to > 128 micrograms of ciprofloxacin per ml for inhibition. Fragments of gyrA and parC encompassing the quinolone resistance-determining region were amplified by PCR, and the DNA sequences of the fragments were determined. All isolates contained a mutation in gyrA of a serine at position 83 (Ser83) to an Leu, and 26 isolates also contained a mutation of Asp87 to one of four amino acids: Asn (n = 14), Tyr (n = 6), Gly (n = 5), or His (n = 1). Twenty-four isolates contained a single mutation in parC, either a Ser80 to Ile (n = 17) or Arg (n = 2) or a Glu84 to Lys (n = 3). The role of a mutation in gyrB was investigated by introducing wild-type gyrB (pBP548) into all isolates; for three transformants MICs of ciprofloxacin were reduced; however, sequencing of PCR-derived fragments containing the gyrB quinolone resistance-determining region revealed no changes. The analogous region of parE was analyzed in 34 of 36 isolates by single-strand conformational polymorphism analysis and sequencing; however, no amino acid substitutions were discovered. The outer membrane protein and lipopolysaccharide profiles of all isolates were compared with those of reference strains, and the concentration of ciprofloxacin accumulated (with or without 100 microM carbony cyanide m-chlorophenylhydrazone [CCCP] was determined. Twenty-two isolates accumulated significantly lower concentrations of ciprofloxacin than the wild-type E. coli isolate; nine isolates accumulated less then half the concentration. The addition of CCCP increased the concentration of ciprofloxacin accumulated, and in all but one isolate the percent increase was greater than that in the control strains. The data indicate that high-level fluoroquinolone resistance in E. coli involves the acquisition of mutations at multiple loci.
Problem of antimicrobial resistance of fecal aerobic gram-negative bacilli in the elderly
Authors: Leistevuo T , Toivonen P , Osterblad M , Kuistila M , Kahra A , Lehtonen A , Huovinen P
Published: October 2018

In this study, we assessed the magnitude of risk (odds ratio [OR]) of patients being colonized with fecal aerobic gram-negative bacilli in two geriatric hospitals compared with the community, and we associated the use of antimicrobial agents with bacterial resistance. One fecal sample was collected from each of 341 patients, aged 60 years or older, during the hospital stay or when visiting the outpatient service. Samples were collected in 1988 and 1993 to 1994. The aerobic gram-negative bacilli from all samples were examined for resistance to seven antimicrobials by a replica plating method. The long-term-hospitalized patients had a significantly higher risk of being colonized with bacilli resistant to ampicillin (OR, 14.3; 95% confidence interval [95% CI], 6.0 to 34.1), cefuroxime (OR, 7.5; 95% CI, 2.7 to 20.8), trimethoprim (ORs, 22.3; 95% CI, 8.6 to 57.8), and tetracycline (OR, 5.2; 95% CI, 2.4 to 10.9) than the outpatients. The respective ORs among the short-term-hospitalized patients compared with the outpatients were 4.0 (95% CI, 1.9 to 8.4), 7.5 (95% CI, 2.7 to 20.8), 5.5 (95% CI, 2 to 14), and 2.0 (95% CI, 1 to 4). In 1993 to 1994 compared with 1988, in both hospitals there was a significantly increased risk of colonization by bacilli resistant to ampicillin (OR, 3.1; 95% CI, 1.9 to 5.1), cefuroxime (OR, 3.8; 95% CI, 2.1 to 6.7), and tetracycline (OR, 1.6; 95% CI, 1.0 to 2.5). However, the total use of antimicrobial agents increased only among the patients of the short-term-care hospital.
In vitro activities of two novel oxazolidinones (U100592 and U100766), a new fluoroquinolone (trovafloxacin), and dalfopristin-quinupristin against Staphylococcus aureus and Staphylococcus epidermidis
Authors: Mulazimoglu L , Drenning S D , Yu V L
Published: October 2018

Two oxazolidinones (U100592 and U100766), trovafloxacin, and a streptogramin combination (dalfopristin-quinupristin) were highly active in vitro against Staphylococcus aureus and Staphylococcus epidermidis, including methicillin-resistant strains. Trovafloxacin was more active than ciprofloxacin. Time-kill synergy studies demonstrated indifference for the oxazolidinones combined with vancomycin and rifampin against methicillin-resistant staphylococci. Spontaneous resistance was observed with all agents.