bluuebunny/crossref_metadata_2025 huggingface dataset:
OXA-18, a class D clavulanic acid-inhibited extended-spectrum beta-lactamase from Pseudomonas aeruginosa
Authors: Philippon L N , Naas T , Bouthors A T , Barakett V , Nordmann P
Published: October 2018

Clinical isolate Pseudomonas aeruginosa Mus showed resistance both to extended-spectrum cephalosporins and to aztreonam. We detected a typical double-disk synergy image when ceftazidime or aztreonam was placed next to a clavulanic acid disk on an agar plate. This resistance phenotype suggested the presence of an extended-spectrum beta-lactamase. Isoelectric focusing revealed that this strain produced three beta-lactamases, of pI 5.5, 7.4, and 8.2. A 2.6-kb Sau3A fragment encoding the extended-spectrum beta-lactamase of pI 5.5 was cloned from P. aeruginosa Mus genomic DNA. This enzyme, named OXA-18, had a relative molecular mass of 30.6 kDa. OXA-18 has a broad substrate profile, hydrolyzing amoxicillin, ticarcillin, cephalothin, ceftazidime, cefotaxime, and aztreonam, but not imipenem or cephamycins. Its activity was totally inhibited by clavulanic acid at 2 microg/ml. Hydrolysis constants of OXA-18 (Vmax, Km) confirmed the MIC results. Cloxacillin and oxacillin hydrolysis was noticeable with the partially purified OXA-18. The blaOXA-18 gene encodes a 275-amino-acid protein which has weak identity with all class D beta-lactamases except OXA-9 and OXA-12 (45 and 42% amino acid identity, respectively). OXA-18 is likely to be chromosomally encoded since no plasmid was found in the strain and because attempts to transfer the resistance marker failed. OXA-18 is peculiar since it is a class D beta-lactamase which confers high resistance to extended-spectrum cephalosporins and seems to have unique hydrolytic properties among non-class A enzymes.
Widespread detection of PER-1-type extended-spectrum beta-lactamases among nosocomial Acinetobacter and Pseudomonas aeruginosa isolates in Turkey: a nationwide multicenter study
Authors: Vahaboglu H , Oztürk R , Aygün G , Coşkunkan F , Yaman A , Kaygusuz A , Leblebicioglu H , Balik I , Aydin K , Otkun M
Published: October 2018

We studied the prevalence and molecular epidemiology of PER-1-type beta-lactamases among Acinetobacter, Klebsiella, and Pseudomonas aeruginosa strains isolated over a 3-month period in eight university hospitals from distinct regions of Turkey. A total of 72, 92, and 367 Acinetobacter, Klebsiella, and P. aeruginosa isolates were studied, respectively. The presence of blaPER was determined by the colony hybridization method and later confirmed by isoelectric focusing. We detected PER-1-type beta-lactamases in 46% (33/72) of Acinetobacter strains and in 11% (40/367) of P. aeruginosa strains but not in Klebsiella strains. PER-1-type enzyme producers were highly resistant to ceftazidime and gentamicin, intermediately resistant to amikacin, and susceptible or moderately susceptible to imipenem and meropenem. Among PER-1-type-beta-lactamase-positive isolates, five Acinetobacter isolates and six P. aeruginosa isolates from different hospitals were selected for ribosomal DNA fingerprinting with EcoRI and SalI. The EcoRI-digested DNAs were later hybridized with a digoxigenin-labelled PER-1 probe. The ribotypes and the lengths of blaPER-carrying fragments were identical in four Acinetobacter strains. A single isolate (Ac3) harbored a PER gene on a different fragment (approximately 4.2 kbp) than the others (approximately 3.4 kbp) and showed a clearly distinguishable ribotype. Ribotypes of P. aeruginosa strains obtained with EcoRI showed three patterns. Similarly, in Pseudomonas strains two different EcoRI fragments harbored blaPER (approximately 4.2 kbp in five isolates and 3.4 kbp in one isolate). PER-1-type beta-lactamases appear to be restricted to Turkey. However, their clonal diversity and high prevalence indicate a high spreading potential.
Absorption of ofloxacin isomers in the rat small intestine
Authors: Rabbaa L , Dautrey S , Colas-Linhart N , Carbon C , Farinotti R
Published: October 2018

Ofloxacin, a chiral fluoroquinolone, possesses two optical isomers. The antibacterial activity of S-(-)-ofloxacin is 8 to 128 times higher than that of R-(+)-ofloxacin. In the rat, a saturable absorption process has been described for racemic ofloxacin. In the present study we investigated the mechanism underlying the in vivo intestinal absorption of ofloxacin enantiomers in the rat. Blood samples were collected from the portal vein. Our results show that the intestinal absorption of ofloxacin isomers is pH dependent, both enantiomers being best absorbed at neutral pH. S-(-)-Ofloxacin seems to have a greater affinity for the intestinal transporter (initial concentrations at 5 min [C(init)] are 0.17 +/- 0.04 and 0.12 +/- 0.03 microg/ml for S-(-)- and R-(+)-ofloxacin, respectively). Dipeptides fail to modify ofloxacin absorption, but amino acids reduce both isomers' absorption (C(init) is reduced by 53 and 33% with glycine for S-(-)- and R-(+)-ofloxacin, respectively, and by 59 and 42% with L-leucine). Gamma amino butyric acid interferes with the absorption of ofloxacin isomers, but less seriously than do amino acids. Furthermore, ofloxacin competes with other fluoroquinolones or P-glycoprotein substrates for a common secretory pathway, resulting in an increased rate of absorption for both ofloxacin isomers; this is probably an indirect result of their reduced efflux from the apical side of intestinal cells.
Low-level release of Shiga-like toxin (verocytotoxin) and endotoxin from enterohemorrhagic Escherichia coli treated with imipenem
Authors: Takahashi K , Narita K , Kato Y , Sugiyama T , Koide N , Yoshida T , Yokochi T
Published: October 2018

Shiga-like toxin (SLT) and endotoxin may participate in the pathogenesis of enterohemorrhagic Escherichia coli (EHEC) infection. Levels of release of SLT and endotoxin from EHEC treated in vitro with antibiotics were estimated. There were differential levels of release of SLT and endotoxin from EHEC treated with different antibiotics. Treatment of EHEC strains, namely, E. coli O157, O111, and O26, with imipenem induced much lower levels of release of SLT and endotoxin than treatment with ceftazidime.
Activity of SCH 56592 compared with those of fluconazole and itraconazole against Candida spp
Authors: Law D , Moore C B , Denning D W
Published: October 2018

The in vitro activity of Schering 56592, a new azole drug, was compared with those of fluconazole and itraconazole against 103 isolates of Candida comprising 10 different species. Schering 56592 was more active than itraconazole and fluconazole, and it was active against many fluconazole-resistant isolates.
Carbapenem resistance in a clinical isolate of Citrobacter freundii
Authors: Mainardi J L , Mugnier P , Coutrot A , Buu-Hoï A , Collatz E , Gutmann L
Published: October 2018

Carbapenem resistance was studied in two sets of Citrobacter freundii strains: (i) strain CFr950, resistant to imipenem (MIC, 16 microg/ml) and isolated in vivo during imipenem therapy, and strain CFr950-Rev, the spontaneous, imipenem-susceptible revertant of CFr950 selected in vitro, and (ii) strains CFr801 and CFr802, two imipenem-resistant mutants selected in vitro from the susceptible clinical isolate CFr800. In all strains, whether they were imipenem-susceptible or -resistant strains, production of the cephalosporinase was derepressed and their Km values for cephaloridine were in the range of 128 to 199 microM. No carbapenemase activity was detected in vitro. The role of cephalosporinase overproduction in the resistance was demonstrated after introduction of the ampD gene which decreased the level of production of cephalosporinase at least 250-fold and resulted in an 8- to 64-fold decrease in the MICs of the carbapenems. The role of reduced permeability in the resistance was suggested by the absence, in CFr950 and CFr802, of two outer membrane proteins (the 42- and 40-kDa putative porins whose levels were considerably decreased in CFr801) and the reappearance of the 42-kDa protein in imipenem-susceptible strain CFr950-Rev. This role was confirmed after introduction of the ompF gene of Escherichia coli into the CFr strains, which resulted in 8- to 16-fold decreases in the MICs of carbapenems for CFr802 and CFr950. We infer from these results that the association of reduced, porin-mediated permeability with high-level cephalosporinase production, observed previously in other gram-negative bacteria, may also confer carbapenem resistance on C. freundii.
Regulation of heme polymerizing activity and the antimalarial action of chloroquine
Authors: Fitch C D , Chou A C
Published: October 2018

Mice infected with Plasmodium berghei served as donors of erythrocytes with a high level of parasitemia for the study of ferriprotoporphyrin IX (FP) polymerization. Six hours after treatment of these mice with 3 micromol of chloroquine per 25 g of body weight, there were significant losses of heme polymerase I (HPA I). For chloroquine-susceptible (CS) P. berghei, the rate of FP polymerization decreased from 541 +/- 42 (mean +/- standard deviation; n = 12) to 51 +/- 19 (n = 8) nmol of FP polymerized per h per ml of packed erythrocytes (normalized to represent 1,000 parasites per 1,000 erythrocytes). For chloroquine-resistant (CR) P. berghei, the rate decreased from 284 +/- 19 (n = 16) to 124 +/- 11 (n = 6) nmol per h per ml. The chloroquine-induced loss of HPA I was accompanied by the accumulation of unpolymerized FP in CS P. berghei but not in CR P. berghei, which is consistent with the hypothesis that FP mediates the antimalarial action of chloroquine. Quinine treatment partially reversed the effects of chloroquine in CS P. berghei but not in CR P. berghei. Cycloheximide treatment antagonized the effects of chloroquine in both lines of parasites. To explain these findings, we propose that chloroquine, quinine, and cycloheximide perturb a regulatory process for HPA I. Furthermore, we propose that when chloroquine engages its target in the regulatory process, it initiates a chain of events which culminates in increased production, accessibility, or reactivity of a regulator (inactivator) of HPA I.
Pharmacokinetic interaction of megestrol acetate with zidovudine in human immunodeficiency virus-infected patients
Authors: Van Harken D R , Pei J C , Wagner J , Pike I M
Published: October 2018

This nonrandomized, two-period crossover study was performed to assess whether concomitant administration of megestrol acetate influences the steady-state pharmacokinetics of zidovudine and its inactive 5'-O-glucuronide metabolite. Twelve HIV-positive, asymptomatic male volunteers received a 100-mg oral capsule dose of zidovudine at least 30 min before meals five times a day at 0700, 1100, 1500, 1900, and 2300 h on study days 1 to 3 and a single 100-mg dose at 0700 h on day 4. On days 5 to 17, 800 mg of megestrol acetate, as a 40-mg/ml aqueous suspension, was administered orally immediately before the 0700 h dose of zidovudine. On days 5 to 16, zidovudine was also administered at 1100, 1500, 1900, and 2300 h. Serial blood samples were collected for 12 h after the single 100-mg dose of zidovudine on days 4 and 17; trough samples were also obtained just before the 0700 h dose on days 2 to 4 and 15 to 17. Levels of zidovudine and its glucuronide in plasma were assayed by a validated radioimmunoassay. Statistical analysis of trough plasma level data indicated that steady-state levels of zidovudine and its glucuronide in plasma had been attained when pharmacokinetic assessments were made on days 4 and 17. When megestrol acetate and zidovudine were coadministered for 13 days, differences of -14, -6.5, and -4.6% in mean zidovudine peak concentration and areas under the curve at 0 to 4 and 0 to 12 h, respectively, +22.5% in mean trough concentration, +2.6% in mean plasma half-life, and no change in median time to peak were observed compared to conditions when zidovudine was administered alone; for zidovudine 5'-O-glucuronide the respective differences were -9, -7.3, -4.4, +2.3, and +10% and no change. None of the differences were statistically significant (P > 0.05). Concomitant therapy with megestrol acetate, at the dose employed to treat anorexia, cachexia, or an unexplained, significant weight loss in AIDS patients, did not alter the steady-state pharmacokinetics of zidovudine or its 5'-O-glucuronide metabolite.
In vitro and in vivo antibacterial activities of CS-940, a new fluoroquinolone, against isolates from patients with respiratory infections
Authors: Miyazaki S , Domon H , Tateda K , Ohno A , Ishii Y , Matsumoto T , Furuya N , Yamaguchi K
Published: October 2018

We compared the in vivo and in vitro activities of CS-940, a new fluoroquinolone, with those of a group of other drugs. The activities of CS-940 against gram-positive cocci and gram-negative rods, including methicillin-susceptible Staphylococcus aureus and penicillin-resistant Streptococcus pneumoniae, were comparable to those of tosufloxacin, with MICs at which 90% of the strains were inhibited (MIC90s) of 0.5 microg/ml or less. Against methicillin-resistant S. aureus, CS-940 was as active as tosufloxacin, with a MIC90 of 16 microg/ml. The efficacy of CS-940 against murine respiratory infections due to S. pneumoniae or Haemophilus influenzae was better than those of tosufloxacin and sparfloxacin. The efficacy of oral doses of CS-940 reflected not only potent in vitro activity but also a high transmigration ratio from the bloodstream to lung tissues.
Antimicrobial susceptibility testing of 230 Helicobacter pylori strains: importance of medium, inoculum, and incubation time
Authors: Hartzen S H , Andersen L P , Bremmelgaard A , Colding H , Arpi M , Kristiansen J , Justesen T , Espersen F , Frimodt-Møller N , Bonnevie O
Published: October 2018

No standardized method of susceptibility testing for Helicobacter pylori is currently available, so before a large agar dilution study comprising 230 H. pylori strains belonging to more than 80 genetically different groups was initiated, we performed a relatively small preliminary study to determine the influences of medium, inoculum density, and incubation time. Seven media were investigated and were primarily evaluated on the basis of their abilities to support growth both semiquantitatively and qualitatively; Iso-Sensitest agar supplemented with 10% horse blood was found to be well suited for the purpose; this was closely followed by Mueller-Hinton agar with 10% horse blood, Mueller-Hinton with 10% sheep blood, and finally, 7% lysed horse blood agar. Investigations of two inoculum densities and two incubation times resulted in recommendations for the use of 10(9) CFU/ml (10[6] CFU/spot) as the inoculum and 72 h as the incubation time. A modest inoculum effect was noted for amoxicillin and metronidazole. By the methodology derived from our preliminary study, the susceptibilities of 230 H. pylori strains to six antibiotics were subsequently determined. The results were generally in accord with those of others, and apart from metronidazole, the MIC of which for approximately 25% of the strains tested was >8 microg/ml, resistance was low in Denmark. The situation might, however, quickly change when and if the number of indications for antibiotic therapy for H. pylori infections increase. Consequently, susceptibility testing of all H. pylori strains is recommended in order to survey the development of resistance, and in our hands the described methodology was relatively easy to perform and the results were easy to read.
Pharmacokinetics of imipenem-cilastatin in critically ill patients undergoing continuous venovenous hemofiltration
Authors: Tegeder I , Bremer F , Oelkers R , Schobel H , Schüttler J , Brune K , Geisslinger G
Published: October 2018

The pharmacokinetics of imipenem-cilastatin were investigated in 12 critically ill patients with acute renal failure (ARF) managed by continuous veno-venous hemofiltration (CVVH) while receiving a fixed combination of 500 mg of imipenem-cilastatin intravenously three or four times daily. No adverse drug reactions were observed. Plasma and hemofiltrate samples were taken at specified times during one dosing interval, and the concentrations of imipenem and cilastatin were determined by high-performance liquid chromatography. Pharmacokinetic variables were calculated by a first-order, two-compartment pharmacokinetic model for both substances. Total clearances of imipenem and cilastatin (mean +/- standard deviations) were 122.2 +/- 28.6 and 29.2 +/- 13.7 ml/min, respectively, with hemofiltration clearances of 22.9 +/- 2.5 and 16.1 +/- 3.1 ml/min, respectively, and nonrenal, nonhemofiltration clearances of 90.8 +/- 26.3 and 13.2 +/- 13.9 ml/min, respectively. Mean imipenem dosage requirements were approximately 2,000 mg/24 h (2,111.8 +/- 493.4 mg/24 h). They were calculated in order to achieve an average steady-state concentration of 12 mg/liter to ensure that concentrations in plasma exceeded the MICs at which 90% of intermediately resistent bacteria are inhibited (8 mg/liter) during the majority of the dosing interval. By contrast, the recommended dosage for patients with end-stage renal failure (ESRF) and infections caused by intermediately resistant bacteria is 1,000 mg/24 h. This remarkable difference may be due (i) to differences in the nonrenal clearance of imipenem between patients with ARF and ESRF and (ii) to the additional clearance by the hemofilter. Since the total clearance of cilastatin was low, marked accumulation occurred, and this was particularly pronounced in patients with additional liver dysfunction. Thus, in patients with ARF managed by CVVH, rather high imipenem doses are required, and these inevitably result in a marked accumulation of cilastatin. The doses of imipenem recommended for patients with ESRF, however, will lead to underdosing and inadequate antibiotic therapy.
Loss of intrinsic aminoglycoside resistance in Acinetobacter haemolyticus as a result of three distinct types of alterations in the aac(6')-Ig gene, including insertion of IS17
Authors: Rudant E , Courvalin P , Lambert T
Published: October 2018

The distribution of the aac(6')-Ig gene, encoding aminoglycoside 6'-N-acetyltransferase-Ig [AAC(6')-Ig], was studied in 96 Acinetobacter haemolyticus strains and 12 proteolytic Acinetobacter strains, including Acinetobacter genomospecies 6, 13, and 14 and 3 unnamed species assigned to this genomic group by DNA-DNA hybridization. This gene was detected by DNA-DNA hybridization in all 96 A. haemolyticus strains and by PCR in 95 strains but was not detected in strains of other species, indicating that it may be used to identify A. haemolyticus. Three A. haemolyticus strains were susceptible to tobramycin and did not produce an aminoglycoside 6'-N-acetylating activity, although they contained aac(6')-Ig-related sequences. An analysis of three susceptible A. haemolyticus strains indicated that aminoglycoside resistance was abolished by the following three distinct mechanisms: (i) a point mutation in aac(6')-Ig that led to a Met56-->Arg substitution, which was shown by analysis of a revertant to be responsible for the loss of resistance; (ii) a polythymine insertion that altered the reading frame; and (iii) insertion of IS17, a new member of the IS903 family. These observations indicated that AAC(6')-Ig is not essential for the viability of A. haemolyticus, although the aac(6')-Ig gene was detected in all members of this species.
Concentrations in plasma and safety of 7 days of intravenous itraconazole followed by 2 weeks of oral itraconazole solution in patients in intensive care units
Authors: Vandewoude K , Vogelaers D , Decruyenaere J , Jaqmin P , De Beule K , Van Peer A , Woestenborghs R , Groen K , Colardyn F
Published: October 2018

Pharmacokinetics and safety of a hydroxy-beta-propyl solution of itraconazole were assessed in 16 patients in an intensive care unit. On the first 2 days, four 1-h infusions of 200 mg were given at 0, 8, 24, and 32 h. From day 3 to 7, inclusive, a single 1-h infusion of 200 mg of itraconazole was given daily. The intravenous (i.v.) treatment was directly followed by repeated administrations of an oral solution of itraconazole at a dosage of either 200 mg once daily or 200 mg twice daily (b.i.d.). During i.v. treatment, steady-state concentrations of itraconazole and hydroxy-itraconazole in plasma were reached within 48 and 96 h, respectively. At the end of i.v. treatment, mean (+/- standard deviation) itraconazole and hydroxy-itraconazole trough concentrations in plasma were 0.344 +/- 0.140 and 0.605 +/- 0.205 microg/ml, respectively. After the 2-week oral follow-up of 200 mg once daily the mean trough concentration had decreased to 0.245 microg/ml, whereas after 200 mg b.i.d. it increased to 0.369 microg/ml. Diarrhea during oral treatment appeared to be dose related and may be due to the solvent hydroxypropyl-beta-cyclodextrin. More severe laboratory abnormalities were noted during the i.v. than the oral treatment phase, probably related to more severe illness in that period of intensive care, but none proved clinically important. These results suggest that plasma itraconazole levels above 0.250 microg/ml may be achieved and maintained with the 1-week i.v. schedule followed by b.i.d. oral administration, whereas the once-daily oral follow-up seems to be a suboptimal treatment.
Immunomodulating effect of fosfomycin on gut-derived sepsis caused by Pseudomonas aeruginosa in mice
Authors: Matsumoto T , Tateda K , Miyazaki S , Furuya N , Ohno A , Ishii Y , Hirakata Y , Yamaguchi K
Published: October 2018

We evaluated the protective effect of fosfomycin (FOM) and an enantiomer of fosfomycin [FOM (+); an isomer of FOM with no bactericidal activity] on murine gut-derived sepsis caused by Pseudomonas aeruginosa. Endogenous bacteremia was induced by administering cyclophosphamide (CY) and ampicillin to specific-pathogen-free mice fed P. aeruginosa. Treatment of mice with FOM at 250 mg/kg of body weight per day twice a day after the second CY administration significantly increased the survival rate compared to that for control mice treated with saline. Treatment with FOM (+) at 20 and 100 mg/kg also significantly increased the survival rate (from 30% for control mice to 80% for treated mice). The bacterial counts in the liver and blood were both significantly lower in FOM(+)-treated mice in comparison with those in liver and blood of saline-treated control mice. FOM(+) administration affected neither the bacterial colonization in the intestinal tract nor the leukocyte counts in the peripheral blood of the mice. After intravascular inoculation of P. aeruginosa, treatment of mice with FOM (+) did not enhance bacterial clearance from the blood of mice pretreated or not enhance bacterial clearance from the blood of mice pretreated or not pretreated with CY, FOM(+) significantly suppressed tumor necrosis factor alpha, interleukin-1 beta, and interleukin-6 levels in the serum of mice after gut-derived sepsis. These results indicate that both FOM and FOM(+) have protective effects against P. aeruginosa bacteremia, despite a lack of specific activity of FOM(+), and suggest that FOM may possess immunomodulating activity and that it induces a protective effect. The protective mechanism is speculated to be that FOM modulates the vivo production of inflammatory cytokines.
Characterization of the 6'-N-aminoglycoside acetyltransferase gene aac(6')-Im [corrected] associated with a sulI-type integron
Authors: Hannecart-Pokorni E , Depuydt F , de wit L , van Bossuyt E , Content J , Vanhoof R
Published: October 2018

The amikacin resistance gene aac(6')-Im [corrected] from Citrobacter freundii Cf155 encoding an aminoglycoside 6'-N-acetyltransferase was characterized. The gene was identified as a coding sequence of 521 bp located down-stream from the 5' conserved segment of an integron. The sequence of this aac(6')-Im [corrected] gene corresponded to a protein of 173 amino acids which possessed 64.2% identity in a 165-amino-acid overlap with the aac(6')-Ia gene product (F.C. Tenover, D. Filpula, K.L. Phillips, and J. J. Plorde, J. Bacteriol. 170:471-473, 1988). By using PCR, the aac(6')-Im [corrected] gene could be detected in 8 of 86 gram-negative clinical isolates from two Belgian hospitals, including isolates of Citrobacter, Klebsiella spp., and Escherichia coli. PCR mapping of the aac(6')-Im [corrected] gene environment in these isolates indicated that the gene was located within a sulI-type integron; the insert region is 1,700 bases long and includes two genes cassettes, the second being ant (3")-Ib.
Decoy approach using RNA-DNA chimera oligonucleotides to inhibit the regulatory function of human immunodeficiency virus type 1 Rev protein
Authors: Nakaya T , Iwai S , Fujinaga K , Sato Y , Otsuka E , Ikuta K
Published: October 2018

Human immunodeficiency virus type 1 (HIV-1) encodes two regulatory proteins, Tat and Rev, that bind to target RNA sequences. These are the trans-activation responsive (TAR) RNA and the Rev-responsive element (RRE), respectively. The Rev protein shifts RNA synthesis to viral transcripts by binding to the RRE within the env gene. In the present study we prepared a RNA-DNA chimera consisting of 29 or 31 nucleotides to inhibit the Rev regulatory function by means of the decoy approach. The chimera oligonucleotides (anti-Rev oligonucleotides [AROs]) contained an RNA "bubble" structure (13 oligonucleotides; the Rev-binding element in RRE) that bound Rev with a high affinity in an in vitro assay. The controls were RNA-DNA chimera oligonucleotides (negative control oligonucleotides [NCOs]) similar to ARO, but without the bubble structure, that bound with considerably less affinity to Rev. When the inhibitory effects of these decoys on HIV-1 replication were examined, we found that AROs, but no NCOs, reduced more than 90% of the HIV-1 production generated by productively infected human T-cell lines. The production of primary HIV-1 isolates in healthy donor-derived peripheral blood mononuclear cells was also similarly inhibited by AROs. In addition, the induction of viral mRNAs and antigens in latently HIV-1-infected ACH-2 cells by tumor necrosis factor alpha was specifically inhibited by AROs, but not by NCOs. No apparent cytotoxicity was caused by either decoy. Thus, the use of a Rev-binding element-based decoy, the RNA-DNA chimera oligonucleotide, may represent a safer approach to gene therapy for reducing the virus load in HIV-1-infected individuals.
Differential distributions in tissues and efficacies of aztreonam and ceftazidime and in vivo bacterial morphological changes following treatment
Authors: Turcotte A , Simard M , Morin N J , Beauchamp D , Bergeron M G
Published: October 2018

The differential tissue distributions of aztreonam and ceftazidime within fibrin clots infected with Pseudomonas aeruginosa, Enterobacter cloacae, and Serratia marcescens, their efficacies, and the in vivo bacterial morphological changes induced by these drugs were evaluated. Rabbits were given intravenously a single dose of 100 mg of either agents/kg of body weight. In the cores of the clots, the peak levels of both drugs were much lower than those observed in the peripheries and in serum. Aztreonam's half-lives within the peripheries and in the cores of the fibrin clots were up to six times higher than observed in serum, while ceftazidime's half-lives in clots were twice that observed in serum. This resulted in a much greater penetration ratio for aztreonam than for ceftazidime. Both drugs controlled the growth of P. aeruginosa in vivo, but E. cloacae and S. marcescens responded better to ceftazidime. Morphological changes were more abundant in the peripheries than in the cores of the clots. In the control group, P. aeruginosa's morphology in the cores was different than that in the peripheries of the clots. Against P. aeruginosa, aztreonam did induce morphological changes in the cores while ceftazidime did not. Electron microscopic studies revealed that morphological changes associated with aztreonam seemed different than those of ceftazidime. Along with elongation of bacteria, more bow tie and herniated bacteria were observed with aztreonam. Though both agents selectively affect PBP 3, as manifested by elongated bacteria, they induce in the peripheries of the clots thickening, breaks, and detachment in bacterial cell walls, alterations which are generally associated with antibiotics affecting PBP 1a and 1b.
Inhibition of multiple phases of human immunodeficiency virus type 1 replication by a dithiane compound that attacks the conserved zinc fingers of retroviral nucleocapsid proteins
Authors: Rice W G , Baker D C , Schaeffer C A , Graham L , Bu M , Terpening S , Clanton D , Schultz R , Bader J P , Buckheit R W , Field L , Singh P K , Turpin J A
Published: October 2018

The human immunodeficiency virus type 1 (HIV-1) nucleocapsid p7 protein contains two retrovirus-type zinc finger domains that are required for multiple phases of viral replication. Chelating residues (three Cys residues and one His residue) of the domains are absolutely conserved among all strains of HIV-1 and other retroviruses, and mutations in these residues in noninfectious virions. These properties establish the zinc finger domains as logical targets for antiviral chemotherapy. Selected dithiobis benzamide (R-SS-R) compounds were previously found to inhibit HIV-1 replication by mediating an electrophilic attack on the zinc fingers. Unfortunately, reaction of these disulfide-based benzamides with reducing agents yields two monomeric structures (two R-SH structures) that can dissociated and no longer react with the zinc fingers, suggesting that in vivo reduction would inactivate the compounds. Through an extensive drug discovery program of the National Cancer Institute, a nondissociable tethered dithiane compound (1,2-dithiane-4,5-diol, 1,1-dioxide, cis; NSC 624151) has been identified. This compound specifically attacks the retroviral zinc fingers, but not other antiviral targets. The lead compound demonstrated broad antiretroviral activity, ranging from field isolates and drug-resistant strains of HIV-1 to HIV-2 and simian immunodeficiency virus. The compound directly inactivated HIV-1 virions and blocked production of infectious virus from cells harboring integrated proviral DNA. NSC 624151 provides a scaffold from which medicinal chemists can develop novel compounds for the therapeutic treatment of HIV infection.
Nonlinear intestinal absorption kinetics of cefuroxime axetil in rats
Authors: Ruiz-Balaguer N , Nacher A , Casabo V G , Merino M
Published: October 2018

Cefuroxime is commercially available for parenteral administration as a sodium salt and for oral administration as cefuroxime axetil, the 1-(acetoxy)ethyl ester of the drug. Cefuroxime axetil is a prodrug of cefuroxime and has little, if any, antibacterial activity until hydrolyzed in vivo to cefuroxime. In this study, the absorption of cefuroxime axetil in the small intestines of anesthetized rats was investigated in situ, by perfusion at four concentrations (11.8, 5, 118 and 200 microM). Oral absorption of cefuroxime axetil can apparently be described as a specialized transport mechanism which obeys Michaelis-Menten kinetics. Parameters characterizing absorption of prodrug in free solution were obtained: maximum rate of absorption (Vmax) = 289.08 +/- 46.26 microM h-1, and Km = 162.77 +/- 31.17 microM. Cefuroxime axetil transport was significantly reduced in the presence of the enzymatic inhibitor sodium azide. On the other hand, the prodrug was metabolized in the gut wall through contact with membrane-bound enzymes in the brush border membrane before absorption occurred. This process reduces the prodrug fraction directly available for absorption. From a bioavailability point of view, therefore, the effects mentioned above can explain the variable and poor bioavailability following oral administration of cefuroxime axetil. Thus, future strategies in oral cefuroxime axetil absorption should focus on increasing the stability of the prodrug in the intestine by modifying the prodrug structure and/or targeting the compound to the absorption site.
Increased activation of the combination of 3'-azido-3'-deoxythymidine and 2'-deoxy-3'-thiacytidine in the presence of hydroxyurea
Authors: Palmer S , Cox S
Published: October 2018

The intracellular phosphorylation of 3'-azido-3'-deoxythymidine and 2'-deoxy-3'-thiacytidine was increased two- to threefold by the addition of hydroxyurea (HU) to the single drugs or to the two drugs in combination. The ratios of drug triphosphate to competing cellular deoxynucleoside triphosphate were increased two- to threefold for both 3'-azido-3'-deoxythymidine and 2'-deoxy-3'-thiacytidine in the presence of HU. These HU-induced increases in 3'-azido-3'-deoxythymidine and 2'-deoxy-3'-thiacytidine metabolism may further enhance the anti-human immunodeficiency virus activity of these two drugs.
Characterization of pncA mutations in pyrazinamide-resistant Mycobacterium tuberculosis
Authors: Scorpio A , Lindholm-Levy P , Heifets L , Gilman R , Siddiqi S , Cynamon M , Zhang Y
Published: October 2018

Pyrazinamide (PZA) is a first-line drug for short-course tuberculosis therapy. Resistance to PZA is usually accompanied by loss of pyrazinamidase (PZase) activity in Mycobacterium tuberculosis. PZase converts PZA to bactericidal pyrazinoic acid, and the loss of PZase activity is associated with PZA resistance. The gene (pncA) encoding the M. tuberculosis PZase has recently been sequenced, and mutations in pncA were previously found in a small number of PZA-resistant M. tuberculosis strains. To further understand the genetic basis of PZA resistance and determine the frequency of PZA-resistant strains having pncA mutations, we analyzed a panel of PZA-resistant clinical isolates and mutants made in vitro. Thirty-three of 38 PZA-resistant clinical isolates had pncA mutations. Among the five strains that did not contain pncA mutations, four were found to be falsely resistant and one was found to be borderline resistant to PZA. The 33 PZA-resistant clinical isolates and 8 mutants made in vitro contained various mutations, including nucleotide substitutions, insertions, or deletions in the pncA gene. The identified mutations were dispersed along the pncA gene, but some degree of clustering of mutations was found at the following regions: Gly132-Thr142, Pro69-Leu85, and Ile5-Asp12. PCR-single-strand conformation polymorphism (SSCP) analysis was shown to be useful for the rapid detection of pncA mutations in the PZA-resistant strains. We conclude that a mutation in the pncA gene is a major mechanism of PZA resistance and that direct sequencing by PCR or SSCP analysis should help to rapidly identify PZA-resistant M. tuberculosis strains.
Direct evidence for uptake of intact liposomes encapsulating silver sulfadiazine by cultured human keratinocytes based on combined transmission electron microscopy and X-ray microanalysis
Authors: Schaller M , Wurm R , Korting H C
Published: October 2018

Cultured human keratinocytes were exposed to liposomally encapsulated silver sulfadiazine, free silver sulfadiazine, silver sulfadiazine cream, and a corresponding vehicle for 5 min to 24 h. Phagocytosis of intact liposomes by keratinocytes was demonstrated in vitro by combined X-ray microanalysis and electron microscopy. Silver as an active part of the antimicrobial served as an electron-dense marker.
High incidence of erythromycin-resistant streptococci in Taiwan
Authors: Wu J J , Lin K Y , Hsueh P R , Liu J W , Pan H I , Sheu S M
Published: October 2018

The activities of nine antimicrobial agents against 247 isolates of group B, C, F, and G streptococci and viridans group streptococci were studied by the broth microdilution method. Erythromycin resistance was found in 29.7, 41.7, 81.8, 23.5, and 53.3% of the strains of group B, C, F, and G streptococci and viridans group streptococci tested, respectively. Macrolides are not considered an optimal alternative to penicillin in the treatment of streptococcal infections, at least empirically, in Taiwan.
In vitro activity of fluvastatin, a cholesterol-lowering agent, and synergy with flucanazole and itraconazole against Candida species and Cryptococcus neoformans
Authors: Chin N X , Weitzman I , Della-Latta P
Published: October 2018

Fluvastatin, a cholesterol-lowering drug, exhibited minimal activity (MICs of 64 to >128 microg/ml) against Candida species and Cryptococcus neoformans. When fluvastatin was combined with fluconazole or itraconazole, both synergistic and additive effects were noted (fractional inhibitory concentration indices of < or = 0.156 to 0.625; fractional lethal concentration indices of < or = 0.156 to 0.75). This combined fungicidal activity was confirmed by time-versus-killing studies.
Sensitivity of Aeromonas hydrophila carbapenemase to delta3-cephems: comparative study with other metallo-beta-lactamases
Authors: Felici A , Perilli M , Franceschini N , Rossolini G M , Galleni M , Frere J M , Oratore A , Amicosante G
Published: October 2018

Ceftriaxone and ceftriaxone S-oxide behaved as inactivators against the metallo-beta-lactamase of Aeromonas hydrophila AE036 and as substrates for the zinc beta-lactamase produced by Bacillus cereus (569/H/9) and Stenotrophomonas maltophilia ULA 511. Moreover, RO 09-1428, a catechol-cephalosporin, was not recognized by the A. hydrophila enzyme. Panipenem, cephalosporin C, cephalosporin C-gamma-lactone, and loracarbef were substrates for the three studied beta-lactamases.
Efficacy of trimethoprim in murine experimental infection with a thymidine kinase-deficient mutant of Escherichia coli
Authors: Tokunaga T , Oka K , Takemoto A , Ohtsubo Y , Gotoh N , Nishino T
Published: October 2018

The antimicrobial activity of trimethoprim is antagonized by thymidine in in vitro susceptibility tests. The purpose of this investigation was to determine whether this antagonism also occurred during experimental infection in mice, which have high serum thymidine concentrations. We derived a mutant strain of Escherichia coli, TT-48, incapable of utilizing exogenous thymidine from parent strain E. coli KC-14 and then investigated the in vitro and in vivo antimicrobial activities of trimethoprim, sulfamethoxazole, cefdinir, and ofloxacin against these strains. E. coli TT-48 lacked the activity of thymidine kinase, which catalyzes the conversion of thymidine to thymidylate, but its growth curve remained close to that of the parent strain. The MICs of all of the antimicrobial agents tested, except cefdinir, for the mutant strain were slightly inferior to those for the parent strain. The bactericidal effect of trimethoprim against the parent strain was antagonized by thymidine at concentrations of more than 1 microg/ml, while that against the mutant strain was not affected by thymidine even at the highest concentration (10 microg/ml). The therapeutic efficacy of trimethoprim in experimental murine infections was significantly higher when the mutant rather than the parent strain was used, whereas the therapeutic efficacy of cefdinir or ofloxacin, whose antimicrobial action is independent of folic acid synthesis, was the same with both strains. Unexpectedly, sulfamethoxazole also had similar efficacy against both strains. Thus, high thymidine concentrations antagonized the antimicrobial activity of trimethoprim in vitro and in vivo.
Pharmacokinetics of zidovudine after rectal administration in human immunodeficiency virus-infected patients
Authors: Wintergerst U , Rolinski B , Bogner J R , Notheis G , Goebel F D , Roscher A A , Belohradsky B H
Published: October 2018

We evaluated the pharmacokinetics of rectally administered zidovudine (ZDV) in 10 human immunodeficiency virus-infected adults. After rectal administration of an aqueous ZDV solution (250 mg of ZDV), mean peak ZDV levels were 1.3 +/- 0.7 micromol/liter (mean +/- standard deviation) versus 5.0 +/- 2.2 micromol/liter (P < 0.0001) after oral intake of a 250-mg ZDV capsule. The half-life at beta phase was 87.8 +/- 39.6 min for rectally administered ZDV versus 55.8 +/- 20.1 min (P = 0.035) for orally administered ZDV. The mean area under the concentration-time curve from 0 min to infinity was 232 +/- 181 micromol/liter x min after rectal administration versus 362 +/- 110 micromol/liter x min after oral intake. Although the two routes were not bioequivalent, ZDV was absorbed considerably after rectal administration, with a pharmacokinetic profile resembling that of a sustained-release device.
Axenically cultured amastigote forms as an in vitro model for investigation of antileishmanial agents
Authors: Sereno D , Lemesre J L
Published: October 2018

Using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide microassay, previously described as a means of quantifying Leishmania amazonensis in vitro at the amastigote stage (D. Sereno and J. L. Lemesre, Parisitol. Res., in press), we have compared the activities of seven drugs, including those currently used to treat leishmaniasis, against axenically grown amastigote and promastigote forms of three Leishmania species (L. amazonensis, L. mexicana, and L. infantum, responsible for diffuse cutaneous, cutaneous, and visceral leishmaniasis, respectively). The ability of axenically cultured amastigote organisms to be used in an investigation of antileishmanial agents was first evaluated. We have confirmed the toxicities of sodium stibogluconate (Pentostam), pentamidine, and amphotericin B to active and dividing populations of axenically cultured amastigotes. The toxicity of potassium antimonyl tartrate trihydrate, which is generally higher than that of Pentostam, seemed to indicate that pentavalent antimony can be metabolized in vivo to compounds, possibly trivalent in nature, which are more active against the amastigote organisms. When the drug susceptibilities of parasites at both stages were compared, great variations were found for all the drugs studied. These major differences, which show the specific chemosusceptibility of the parasite at the mammalian stage, demonstrate the potential of using cultured amastigotes instead of promastigotes in a drug-screening procedure for early detection. This in vitro model may help in the isolation of active compounds, particularly those with low-grade activities, against the mammalian stage of the parasite.
Galactose oxidase-glucan binding domain fusion proteins as targeting inhibitors of dental plaque bacteria
Authors: Lis M , Kuramitsu H K
Published: October 2018

In order to inhibit the growth of bacteria present in the human oral cavity, a novel system which targets antimicrobial agents to dental plaque has been developed. This system involves a hybrid protein consisting of a peptide expressing the bactericidal properties of galactose oxidase (GAO) fused to the glucan binding domain (GBD) of the Streptococcus mutans glucosyltransferase-S enzyme. A gene encoding GAO from the fungus Fusarium sp. has been inserted into an Escherichia coli expression vector and fused to sequences encoding the GBD, which binds to the glucans synthesized by oral streptococci. Bacterial extracts expressing the hybrid protein were tested for their ability to target the GAO activity to an in vitro plaque model consisting of streptococcal cells bound to microtiter plate wells. The binding of the hybrid protein to the streptococcal cells through its GBD and the dependence of binding on the production of glucans by bacteria were demonstrated. Furthermore, killing of three different species of oral streptococci by bound hybrid protein in conjunction with the galactose-lactoperoxidase-iodide cytotoxic system has been demonstrated. These results suggest a novel strategy for controlling dental plaque formation as well as dental caries in humans.
Therapeutic efficacies of isoniazid and rifampin encapsulated in lung-specific stealth liposomes against Mycobacterium tuberculosis infection induced in mice
Authors: Deol P , Khuller G K , Joshi K
Published: October 2018

One recent promising development in the modification of drug formulations to improve chemotherapy is the use of a liposome-mediated drug delivery system. The efficacies of isoniazid and rifampin encapsulated in lung-specific stealth liposomes were evaluated by injecting liposomal drugs and free drugs into tuberculous mice twice a week for 6 weeks. Liposome-encapsulated drugs at and below therapeutic concentrations were more effective than free drugs against tuberculosis, as evaluated on the basis of CFUs detected, organomegaly, and histopathology. Furthermore, liposomal drugs had marginal hepatotoxicities as determined from the levels of total bilirubin and hepatic enzymes in serum. The elimination of mycobacteria from the liver and spleen was also higher with liposomal drugs than with free drugs. The encapsulation of antitubercular drugs in lung-specific stealth liposomes seems to be a promising therapeutic approach for the chemotherapy of tuberculosis.
Antimicrobial activity of DU-6681a, a parent compound of novel oral carbapenem DZ-2640
Authors: Tanaka M , Hohmura M , Nishi T , Sato K , Hayakawa I
Published: October 2018

The in vitro antibacterial activity of DU-6681a, a parent compound of DZ-2640, against gram-positive and -negative bacteria was compared with those of penems and cephalosporins currently available. MICs at which 90% of the isolates are inhibited (MIC90s) of the compound for clinical isolates of methicillin-susceptible and -resistant Staphylococcus aureus and Staphylococcus epidermidis, including methicillin-susceptible and -resistant strains, were 0.10, 25, and 12.5 microg/ml, respectively. DU-6681a inhibited the growth of all strains of Streptococcus pyogenes and of penicillin-susceptible and -insusceptible Streptococcus pneumoniae at 0.006, 0.025, and 0.20 microg/ml, respectively, and MIC90s of the compound were 6.25 and >100 microg/ml for Enterococcus faecalis and Enterococcus faecium, respectively. MIC90s of DU-6681a were 0.20, 0.10, and 0.025 microg/ml for Haemophilus influenzae, Moraxella catarrhalis, and Neisseria gonorrhoeae, respectively. For Pseudomonas aeruginosa, the MIC50 and MIC90 of DU-6681a were 25 and 50 microg/ml, respectively. DU-6681a activity was not affected by different media, varied inoculum size (10(4) to 10(7) CFU), or the addition of human serum but was decreased under acidic conditions against gram-negative bacteria, under alkaline conditions against gram-positive bacteria, and in human urine, as was the activity of the other antibiotics tested. The frequency of spontaneous resistance to DU-6681a was less than or equal to those of the reference compounds. Time-kill curve studies demonstrated the bactericidal action of DU-6681a against S. aureus, S. pneumoniae, Escherichia coli, and H. influenzae.