bluuebunny/crossref_metadata_2025 huggingface dataset:
In Vivo Efficacies of Amoxicillin and Cefuroxime against Penicillin-Resistant
Streptococcus pneumoniae
in a Gerbil Model of Acute Otitis Media
Authors: Cenjor Carlos , Ponte Carmen , Parra Araceli , Nieto Eva , García-Calvo Gloria , Giménez María José , Aguilar Lorenzo , Soriano Francisco
Published: October 2018

The comparative efficacies of amoxicillin and cefuroxime against acute otitis media caused by a penicillin-resistant (MIC, 2 μg/ml)
Streptococcus pneumoniae
strain were assessed in a gerbil model by challenging each ear with 10
7
bacteria through transbullar instillation. Each antibiotic was tested at two doses (5 and 20 mg/kg of body weight) administered at 2, 10, and 18 h postinoculation. Samples were obtained from the middle ear (ME) on days 3 and 7 postinoculation for determination of bacterial counts. Only amoxicillin, at both doses, was able to significantly halt the weight loss in animals, reducing both the number of culture-positive animals and the bacterial concentration in ME samples versus the values for untreated animals. Comparison of the efficacies between the antibiotics, determined by their ability to achieve culture-negative ME specimens, showed that amoxicillin at 5 mg/kg was significantly more active than cefuroxime at the same dose. The use of higher doses of either amoxicillin or cefuroxime did not produce significantly better results than those obtained with the lower dose but caused a greater inflammatory response. The more favorable results obtained with amoxicillin compared with those obtained with cefuroxime could be related to the antimicrobial susceptibility of the pneumococcal strain (MICs and minimum bactericidal concentrations of 1 and 1 μg/ml and 4 and 4 μg/ml for amoxicillin and cefuroxime, respectively) as well as to the better pharmacokinetic parameters obtained with amoxicillin.
Assessment of Antifungal Activities of Fluconazole and Amphotericin B Administered Alone and in Combination against
Candida albicans
by Using a Dynamic In Vitro Mycotic Infection Model
Authors: Lewis Russell E. , Lund Brian C. , Klepser Michael E. , Ernst Erika J. , Pfaller Michael A.
Published: October 2018

We evaluated the pharmacodynamic activities of fluconazole and amphotericin B given alone and in combination against
Candida albicans
by using an in vitro model of bloodstream infection that simulates human serum pharmacokinetic parameters for these antifungals. Fluconazole was administered as a bolus into the model to simulate regimens of 200 mg every 24 h (q24h) and 400 mg q24h. Amphotericin B was administered at doses producing the peak concentration (2.4 μg/ml) observed with a regimen of 1 mg/kg of body weight q24h. A combination regimen of fluconazole (400 mg q24h) and amphotericin B (1 mg/kg q24h) administered simultaneously and as a staggered regimen (amphotericin B bolus given 8 h after fluconazole bolus) was also simulated in the model to characterize possible antagonism between these agents. Fluconazole alone and amphotericin B alone demonstrated fungistatic (<99.9% reduction in numbers of CFU per milliliter from the starting inoculum) and fungicidal (>99.9% reduction) activity, respectively. When fluconazole and amphotericin B were administered simultaneously, fungicidal activity similar to that observed with amphotericin B alone was observed. Staggered administration of fluconazole and amphotericin B, however, resulted in a substantial reduction of the fungicidal activity of amphotericin B, producing fungistatic activity similar to that observed with noncombination fluconazole regimens. These results demonstrate the usefulness of this model for comparing the in vitro pharmacodynamic characteristics of different antifungal regimens and support the theory of azole-polyene antagonism. The effects of this antagonism on the in vivo activity and clinical usefulness of combination antifungal therapy, however, remain to be determined.
Novel Antibiotic Susceptibility Tests by the ATP-Bioluminescence Method Using Filamentous Cell Treatment
Authors: Hattori Noriaki , Nakajima Moto-O , O’Hara Koji , Sawai Tetsuo
Published: October 2018

Antimicrobial susceptibility testing by the ATP-bioluminescence method has been noted for its speed; it provides susceptibility results within 2 to 5 h. However, several disagreements between the ATP method and standard methodology have been reported. The present paper describes a novel ATP method in a 3.5-h test which overcomes these deficiencies through the elimination of false-resistance discrepancies in tests on gram-negative bacteria with β-lactam agents. In our test model using
Pseudomonas aeruginosa
and piperacillin, it was shown that ATP in filamentous cells accounted for the false resistance. We found that 0.5% 2-amino-2-methyl-1,3-propanediol (AMPD) extracted ATP from the filamentous cells without affecting normal cells and that 0.3 U of adenosine phosphate deaminase (APDase)/ml simultaneously digested the extracted ATP. We used the mixture of these reagents for the pretreatment of cells in a procedure we named filamentous cell treatment, prior to ATP measurements. This novel ATP method with the filamentous cell treatment eliminated false-resistance discrepancies in tests on
P. aeruginosa
with β-lactam agents, including piperacillin, cefoperazone, aztreonam, imipenem-cilastatin, ceftazidime, and cefsulodin. Furthermore, this novel methodology produced results which agreed with those of the standard microdilution method in other tests on gram-negative and gram-positive bacteria, including
P. aeruginosa
,
Escherichia coli
,
Staphylococcus aureus
, and
Enterococcus faecalis
, for non-β-lactam agents, such as fosfomycin, ofloxacin, minocycline, and aminoglycosides. MICs obtained by the novel ATP method were also in agreement with those obtained by the agar dilution method of susceptibility testing. From these results, it was shown that the novel ATP method could be used successfully to test the activities of antimicrobial agents with the elimination of the previously reported discrepancies.
Analysis of the Pharmacokinetic Interaction between Cephalexin and Quinapril by a Nonlinear Mixed-Effect Model
Authors: Padoin C. , Tod M. , Perret G. , Petitjean O.
Published: October 2018

Oligopeptidic drugs such as β-lactams and angiotensin-converting enzyme inhibitors share the same carriers in humans and animals, which results in possible pharmacokinetic interactions. To model such interactions, the effects of quinapril on cephalexin pharmacokinetics were investigated in rats. Blood cephalexin concentrations were measured by liquid chromatography, and the data were analyzed by a noncompartmental method and by fitting a bicompartmental model by a nonlinear mixed-effect modeling approach. Five groups of eight rats were examined. In the first three groups, cephalexin elimination kinetics after intra-arterial administration alone or in combination with quinapril given by the parenteral or the oral route were studied, and the occurrence of a pharmacokinetic interaction was not revealed. The absence of an effect of quinapril on cephalexin elimination after parenteral administration might be explained either by the higher affinity of cephalexin for the renal anionic transport system than that of quinapril or by the much higher concentrations of cephalexin than those of quinapril. In the last two groups, cephalexin was administered by the oral route alone or in combination with quinapril. The mean area under the concentration-time curve (AUC) for cephalexin was increased by ca. 30% by coadministration of quinapril (40.1 versus 31.4 mg · h/liter;
P
= 0.04). The mean elimination clearance of cephalexin was significantly decreased by quinapril, from 0.81 to 0.64 liter/h/kg of body weight (
P
< 0.05), probably by competitive inhibition of cephalexin secretion at the tubular level. The mean absorption rate constant of cephalexin was significantly lowered by quinapril (from 0.249 to 0.177 h
−1
;
P
< 0.01), without modification of the extent of absorption (89%). This pharmacokinetic interaction could be explained by competitive inhibition of cephalexin active transport by quinapril at the intestinal level.
Delivery of the Non-Membrane-Permeative Antibiotic Gentamicin into Mammalian Cells by Using
Shigella flexneri
Membrane Vesicles
Authors: Kadurugamuwa Jagath L. , Beveridge Terry J.
Published: October 2018

We developed a model to test whether non-membrane-permeative therapeutic agents such as gentamicin could be delivered into mammalian cells by means of bacterial membrane vesicles. Many gram-negative bacteria bleb off membrane vesicles (MVs) during normal growth, and the quantity of these vesicles can be increased by brief exposure to gentamicin (J. L. Kadurugamuwa and T. J. Beveridge, J. Bacteriol. 177:3998–4008, 1995), which can be entrapped within the MVs. Gentamicin-induced MVs (g-MVs) were isolated from
Shigella flexneri
and contained 85 ± 2 ng of gentamicin per μg of MV protein. Immunogold electron microscopic labeling of thin sections with antibodies specific to
S. flexneri
lipopolysaccharide (LPS) demonstrated the adherence and subsequent engulfment of MVs by the human Henle 407 intestinal epithelial cell line. Further incubation of g-MVs with
S. flexneri
-infected Henle cells revealed that the g-MVs penetrated throughout the infected cells and reduced the intracellular pathogen by ∼1.5 log
10
CFU in the first hour of incubation. Antibiotic was detected in the cytoplasms of host cells, indicating the intracellular placement of the drug following the penetration of g-MVs. Soluble antibiotic, added as a fluid to the tissue culture growth medium, had no effect on intracellular bacterial growth, confirming the impermeability of the cell membranes of the tissue to gentamicin. Western blot analysis of MVs with
S. flexneri
Ipa-specific antibodies demonstrated that the invasion protein antigens IpaB, IpaC, and IpaD were present in MVs. Being bilayered, with outer faces composed of LPS and Ipa proteins, these MVs were readily engulfed by the otherwise impermeable membranes and eventually liberated their contents into the cytoplasmic substance of the host tissue.
Efficacies of Zinc-Finger-Active Drugs againstGiardia lamblia
Authors: Nash Theodore , Rice William G.
Published: October 2018

Twenty-nine of 34 (85%) Zn-finger-active compounds at 300 μM or less inhibited the growth ofGiardia lamblia. The most active compound, disulfiram (Antabuse), was cidal at 1.23 ± 0.32 μM. In the adult mouse model, significant in vivo activity was demonstrated by increased cure rates and decreased parasite burdens.
Oral Cimetidine Prolongs Clarithromycin Absorption
Authors: Amsden Guy W. , Cheng Kit L. , Peloquin Charles A. , Nafziger Anne N.
Published: October 2018

The present study was conducted to identify any potential interaction between oral cimetidine and clarithromycin. Twelve healthy subjects were administered single doses of clarithromycin alone and with oral cimetidine dosed to steady state. Cimetidine prolonged the absorption of clarithromycin, as evidenced by decreased peak concentrations of both clarithromycin and 14-OH-clarithromycin (14OHC) in serum (46 and 43%, respectively), a delay in the formation of 14OHC (increase of 68%), and increases in both of their half-lives (75 and 82%, respectively), despite no changes in total oral clearance or area under the concentration-time curve for either compound. No mechanism for this interaction has been identified.
Urinary Excretion and Bactericidal Activities of a Single Oral Dose of 400 Milligrams of Fleroxacin versus a Single Oral Dose of 800 Milligrams of Pefloxacin in Healthy Volunteers
Authors: Naber Kurt G. , Theuretzbacher Ursula , Kinzig Martina , Savov Orlin , Sörgel Fritz
Published: October 2018

Twelve healthy volunteers participated in this randomized crossover study to compare the concentrations and recovery levels of fleroxacin and pefloxacin in urine and to assess their bactericidal activities against 12 strains of urinary pathogens with different susceptibilities over a wide range of MICs. The volunteers received a single oral dose of 400 mg of fleroxacin or 800 mg of pefloxacin. The mean cumulative renal excretion of unchanged fleroxacin,
N
-demethyl-fleroxacin, and
N
-oxide-fleroxacin accounted for 67, 7, and 6% of the total dose, respectively. The total urinary recovery of pefloxacin and the active metabolite norfloxacin was 34%. In the time-kill and the urinary bactericidal titer (UBT) studies, only the subjects’ urine not supplemented with broth was used. With most tested organisms and both quinolones it took more than 8 h to achieve a reduction in CFU of 99.9% (3 log units). Overall, there was a good correlation between UBTs and MICs for the strains. Against
Escherichia coli
ATCC 25922 the median UBTs were similar for both antibiotics and at least 1:8 for 96 h; against the
E. coli
strain for which the MIC was 0.5 μg/ml the UBT was at least 1:4 for 48 h. The UBTs of both drugs against
Klebsiella pneumoniae
were at least 1:16 for 72 h. The UBTs for
Staphylococcus aureus
(the MIC for which was 16 μg/ml) of both antibiotics were low, and in some of the samples, no bactericidal titers were observed. UBTs for
Proteus mirabilis
of pefloxacin are significantly higher than those of fleroxacin. For
Pseudomonas aeruginosa
the median UBTs were present for the 24-to-48-h interval. The same is true for
Enterococcus faecalis
. Against
Staphylococcus saprophyticus
, UBTs were present for at least 48 h with both quinolones. Overall, a single oral dose of 400 mg of fleroxacin exhibits UBTs comparable to those of 800 mg of pefloxacin. Therefore, it may be expected that half of the dose of fleroxacin gives comparable results in the treatment of urinary tract infections; this should be substantiated in comparative clinical trials.
16S rRNA Mutation Associated with Tetracycline Resistance in a Gram-Positive Bacterium
Authors: Ross Jeremy I. , Eady E. Anne , Cove Jonathan H. , Cunliffe William J.
Published: October 2018

A genetic basis for tetracycline resistance in cutaneous propionibacteria was suggested by comparing the nucleotide sequences of the 16S rRNA genes from 16 susceptible and 21 resistant clinical isolates and 6 laboratory-selected tetracycline-resistant mutants of a susceptible strain. Fifteen clinical isolates resistant to tetracycline were found to have cytosine instead of guanine at a position cognate with
Escherichia coli
16S rRNA base 1058 in a region important for peptide chain termination and translational accuracy known as helix 34. Cytosine at base 1058 was not detected in the laboratory mutants or the tetracycline-susceptible strains. The apparent mutation was recreated by site-directed mutagenesis in the cloned
E. coli
ribosomal operon,
rrnB
, encoded by pKK3535.
E. coli
strains carrying the mutant plasmid were more resistant to tetracycline than those carrying the wild-type plasmid both in MIC determinations and when grown in tetracycline-containing liquid medium. These data are consistent with a role for the single 16S rRNA base mutation in clinical tetracycline resistance in cutaneous propionibacteria.
Multidose Pharmacokinetics of Ritonavir and Zidovudine in Human Immunodeficiency Virus-Infected Patients
Authors: Cato Allen , Qian Jiang , Hsu Ann , Levy Benjamin , Leonard John , Granneman Richard
Published: October 2018

The effect of coadministration of ritonavir and zidovudine (ZDV) on the pharmacokinetics of these drugs was investigated in a three-period, multidose, crossover study. Eighteen asymptomatic, human immunodeficiency virus-positive men were assigned randomly to six different sequences of the following three regimens: ZDV (200 mg every 8 h [q8h]) alone for 4 days, ritonavir (300 mg q6h) alone for 4 days, and ZDV with ritonavir for 4 days. Ritonavir pharmacokinetics were unaffected by coadministration with ZDV. However, ZDV exposure was reduced by about 26% (
P
< 0.05) in the presence of ritonavir. The maximum concentration in (
C
max
) of ZDV plasma decreased from 748 ± 375 (mean ± standard deviation) to 546 ± 296, and area under the concentration-time curve from 0 to 24 h (AUC
0–24
) decreased from 3,052 ± 1,007 to 2,261 ± 715 when coadministered with ritonavir. In contrast, the ZDV elimination rate constant was unaffected by ritonavir, suggesting that there was no change in ZDV systemic metabolism. Correspondingly, differences in ZDV-glucuronide
C
max
and AUC were not statistically significantly different between regimens (
P
> 0.31). Also, there were no apparent differences in the formation of 3′-amino-3′-deoxythymidine or in the adverse event profiles between the regimens. The lack of change in ritonavir pharmacokinetics suggests that dosage adjustment of ritonavir is unnecessary when it is administered concurrently with ZDV. The clinical relevance of a 26% reduction in ZDV exposure when ZDV is administered with ritonavir is unknown. In addition to other multidrug regimens, the long-term safety and efficacy of coadministration of ritonavir and ZDV is being investigated.
Effect of Growth Rate on Resistance of
Candida albicans
Biofilms to Antifungal Agents
Authors: Baillie George S. , Douglas L. Julia
Published: October 2018

A perfused biofilm fermentor, which allows growth-rate control of adherent microbial populations, was used to assess whether the susceptibility of
Candida albicans
biofilms to antifungal agents is dependent on growth rate. Biofilms were generated under conditions of glucose limitation and were perfused with drugs at a high concentration (20 times the MIC). Amphotericin B produced a greater reduction in the number of daughter cells in biofilm eluates than ketoconazole, fluconazole, or flucytosine. Similar decreases in daughter cell counts were observed when biofilms growing at three different rates were perfused with amphotericin B. In a separate series of experiments, intact biofilms, resuspended biofilm cells, and newly formed daughter cells were removed from the fermentor and were exposed to a lower concentration of amphotericin B for 1 h. The susceptibility profiles over a range of growth rates were then compared with those obtained for planktonic cells grown at the same rates under glucose limitation in a chemostat. Intact biofilms were resistant to amphotericin B at all growth rates tested, whereas planktonic cells were resistant only at low growth rates (≤0.13 h
−1
). Cells resuspended from biofilms were less resistant than intact biofilm populations but more resistant than daughter cells; the susceptibilities of both these cell types were largely independent of growth rate. Our findings indicate that the amphotericin B resistance of
C. albicans
biofilms is not simply due to a low growth rate but depends on some other feature of the biofilm mode of growth.
Antibacterial Activity of Gatifloxacin (AM-1155, CG5501, BMS-206584), a Newly Developed Fluoroquinolone, against Sequentially Acquired Quinolone-Resistant Mutants and the
norA
Transformant of
Staphylococcus aureus
Authors: Fukuda Hideyuki , Hori Satoshi , Hiramatsu Keiichi
Published: October 2018

Alternate mutations in the
grlA
and
gyrA
genes were observed through the first- to fourth-step mutants which were obtained from four
Staphylococcus aureus
strains by sequential selection with several fluoroquinolones. The increases in the MICs of gatifloxacin accompanying those mutational steps suggest that primary targets of gatifloxacin in the wild type and the first-, second-, and third-step mutants are wild-type topoisomerase IV (topo IV), wild-type DNA gyrase, singly mutated topo IV, and singly mutated DNA gyrase, respectively. Gatifloxacin had activity equal to that of tosufloxacin and activity more potent than those of norfloxacin, ofloxacin, ciprofloxacin, and sparfloxacin against the second-step mutants (
grlA gyrA
; gatifloxacin MIC range, 1.56 to 3.13 μg/ml) and had the most potent activity against the third-step mutants (
grlA gyrA grlA
; gatifloxacin MIC range, 1.56 to 6.25 μg/ml), suggesting that gatifloxacin possesses the most potent inhibitory activity against singly mutated topo IV and singly mutated DNA gyrase among the quinolones tested. Moreover, gatifloxacin selected resistant mutants from wild-type and the second-step mutants at a low frequency. Gatifloxacin possessed potent activity (MIC, 0.39 μg/ml) against the NorA-overproducing strain
S. aureus
NY12, the
norA
transformant, which was slightly lower than that against the parent strain SA113. The increases in the MICs of the quinolones tested against NY12 were negatively correlated with the hydrophobicity of the quinolones (correlation coefficient, −0.93;
P
< 0.01). Therefore, this slight decrease in the activity of gatifloxacin is attributable to its high hydrophobicity. Those properties of gatifloxacin likely explain its good activity against quinolone-resistant clinical isolates of
S. aureus
harboring the
grlA
,
gyrA
, and/or
norA
mutations.
Efficacy of Nitazoxanide against
Cryptosporidium parvum
in Cell Culture and in Animal Models
Authors: Theodos Cynthia M. , Griffiths Jeffrey K. , D’Onfro Jennifer , Fairfield Alexandra , Tzipori Saul
Published: October 2018

Nitazoxanide (NTZ), a drug currently being tested in human clinical trials for efficacy against chronic cryptosporidiosis, was assessed in cell culture and in two animal models. The inhibitory activity of NTZ was compared with that of paromomycin (PRM), a drug that is partially effective against
Cryptosporidium parvum
. A concentration of 10 μg of NTZ/ml (32 μM) consistently reduced parasite growth in cell culture by more than 90% with little evidence of drug-associated cytotoxicity, in contrast to an 80% reduction produced by PRM at 2,000 μg/ml (3.2 mM). In contrast to its efficacy in vitro, NTZ at either 100 or 200 mg/kg of body weight/day for 10 days was ineffective at reducing the parasite burden in
C. parvum
-infected, anti-gamma-interferon-conditioned SCID mice. Combined treatment with NTZ and PRM was no more effective than treatment with PRM alone. Finally, NTZ was partially effective at reducing the parasite burden in a gnotobiotic piglet diarrhea model when given orally for 11 days at 250 mg/kg/day but not at 125 mg/kg/day. However, the higher dose of NTZ induced a drug-related diarrhea in piglets that might have influenced its therapeutic efficacy. As we have previously reported, PRM was effective at markedly reducing the parasite burden in piglets at a dosage of 500 mg/kg/day. Our results indicate that of all of the models tested, the piglet diarrhea model most closely mimics the partial response to NTZ treatment reported to occur in patients with chronic cryptosporidiosis.
In Vivo Activity and Pharmacokinetic Evaluation of a Novel Long-Acting Carbapenem Antibiotic, MK-826 (L-749,345)
Authors: Gill Charles J. , Jackson Jesse J. , Gerckens Lynn S. , Pelak Barbara A. , Thompson Randall K. , Sundelof Jon G. , Kropp Helmut , Rosen H.
Published: October 2018

MK-826 (formerly L-749,345), is a potent 1-β-methyl carbapenem with a long half-life and broad spectrum of activity. This compound is presently in phase-II clinical trials. Its activity against a number of gram-positive and gram-negative organisms was compared to those of imipenem (IPM) and eight other β-lactam agents in two in vivo murine infection models. The distribution in tissue and pharmacokinetic properties of MK-826 and ceftriaxone (CTRX) were also evaluated in CD-1 mice following a single intraperitoneal dose (10 mg/kg of body weight). In addition, concentrations in plasma as well as biliary and urinary recovery of MK-826 were compared to that of CTRX in a cannulated rat model. In a localized murine thigh infection model, MK-826 and IPM were superior to a variety of β-lactam antibiotics in reduction of
Staphylococcus aureus
CFU compared with results from nontreated controls (eliminating ≥4 log
10
CFU). Similar activities of IPM and MK-826 were observed in a gram-positive bacterial murine systemic infection model. While IPM demonstrated greater efficacy than MK-826 against
Enterobacter cloacae
(50% effective doses [ED
50
s] of 0.062 and 0.227 mg/kg, respectively) and
Pseudomonas aeruginosa
(ED
50
s of 0.142 and 3.0 mg/kg, respectively) systemic infections, MK-826 was 8- to 350-fold more efficacious than IPM against all other gram-negative organisms in this infection model. In mice, MK-826 demonstrated a higher peak concentration in serum (62.8 versus 42.6 μg/ml) and a larger area under the curve (AUC) (150.8 versus 90.0 μg · hr/ml) than CTRX. The concentrations of MK-826 and CTRX in serum declined slowly, with levels of 3.6 and 2.0 μg/ml remaining, respectively, at 6 h posttreatment. The rat pharmacokinetic model showed the average AUC of MK-826 to be greater than that of CTRX (284 versus 142 μg · hr/ml) following a single 10-mg/kg dose. Also, a half-life of MK-826 longer than that of CTRX (3.2 versus 2.3 h) was observed in this species. The total amount of drug excreted in the bile in 8 h was greater for CTRX (55 to 64% of the dose) than for MK-826 (6 to 12.5% of the dose). Urinary recovery was similar for both antibiotics, with 16 to 18% of the dose recovered over an 8-h period. This excellent broad-spectrum in vivo efficacy of MK-826, together with advantageous pharmacokinetics, supports the argument for its further clinical development.
Activities of Three Quinolones, Alone and in Combination with Extended-Spectrum Cephalosporins or Gentamicin, against
Stenotrophomonas maltophilia
Authors: Visalli Melissa A. , Jacobs Michael R. , Appelbaum Peter C.
Published: October 2018

The present study examined the activities of trovafloxacin, levofloxacin, and ciprofloxacin, alone and in combination with cefoperazone, ceftazidime, cefpirome, and gentamicin, against 100 strains of
Stenotrophomonas maltophilia
by the MIC determination method and by synergy testing of the combinations by the time-kill and checkerboard titration methods for 20 strains. The respective MICs at which 50% and 90% of isolates were inhibited for the drugs used alone were as follows: trovafloxacin, 0.5 and 2.0 μg/ml; levofloxacin, 2.0 and 4.0 μg/ml; ciprofloxacin, 4.0 and 16.0 μg/ml; cefoperazone, >128.0 and >128.0 μg/ml; ceftazidime, 32.0 and >128.0 μg/ml; cefpirome, >128.0 and >128.0 μg/ml; and gentamicin, 128.0 and >128.0 μg/ml. Synergistic fractional inhibitory concentration indices (≤0.5) were found for ≥50% of strains for trovafloxacin-cefoperazone, trovafloxacin-ceftazidime, levofloxacin-cefoperazone, levofloxacin-ceftazidime, ciprofloxacin-cefoperazone, and ciprofloxacin-ceftazidime, with other combinations affecting fewer strains. For 20 strains tested by the checkerboard titration and time-kill methods, synergy (≥100-fold drop in count compared to the count achieved with the more active compound) was more pronounced after 12 h due to regrowth after 24 h. At 12 h, trovafloxacin at 0.004 to 0.5 μg/ml showed synergy with cefoperazone for 90% of strains, with ceftazidime for 95% of strains with cefpirome for 95% of strains, and with gentamicin for 65% of strains. Levofloxacin at 0.03 to 0.5 μg/ml and ciprofloxacin at 0.5 to 2.0 μg/ml showed synergy with cefoperazone for 80% of strains, with ceftazidime for 90 and 85% of strains, respectively, with cefpirome for 85 and 75% of strains, respectively, and with gentamicin for 65 and 75% of strains, respectively. Time-kill assays were more discriminatory than checkerboard titration assays in demonstrating synergy for all combinations.
Rapid Detection and Evaluation of Clinical Characteristics of Emerging Multiple-Drug-Resistant Gram-Negative Rods Carrying the Metallo-β-Lactamase Gene
bla
IMP
Authors: Hirakata Yoichi , Izumikawa Koichi , Yamaguchi Toshiyuki , Takemura Hiromu , Tanaka Hironori , Yoshida Ryoji , Matsuda Junichi , Nakano Michiko , Tomono Kazunori , Maesaki Shigefumi , Kaku Mitsuo , Yamada Yasuaki , Kamihira Shimeru , Kohno Shigeru
Published: October 2018

Gram-negative rods (GNR) carrying the transferable carbapenem resistance gene
bla
IMP
, including
Pseudomonas aeruginosa
and
Serratia marcescens
, have been isolated from more than 20 hospitals in Japan. Although the emergence of such multiple-drug-resistant bacteria is of utmost clinical concern, little information in regard to the distribution of
bla
IMP
-positive GNR in hospitals and the clinical characteristics of infected patients is available. To address this, a system for the rapid detection of the
bla
IMP
gene with a simple DNA preparation and by enzymatic detection of PCR products was developed. A total of 933 ceftazidime-resistant strains of GNR isolated between 1991 and 1996 at Nagasaki University Hospital, Nagasaki, Japan, were screened for the
bla
IMP
gene; 80 isolates were positive, including 53
P. aeruginosa
isolates, 13 other glucose-nonfermenting bacteria, 13
S. marcescens
isolates, and 1
Citrobacter freundii
isolate. Most of the patients from whom
bla
IMP
-positive organisms were isolated had malignant diseases (53.8%). The organisms caused urinary tract infections, pneumonia, or other infections in 46.3% of the patients, while they were just colonizing the other patients evaluated. It was possible that
bla
IMP
-positive
P. aeruginosa
strains contributed to the death of four patients, while the other infections caused by GNR carrying
bla
IMP
were not lethal. DNA fingerprinting analysis by pulsed-field gel electrophoresis suggested the cross transmission of strains within the hospital. The isolates were ceftazidime resistant and were frequently resistant to other antibiotics. Although no particular means of pathogenesis of
bla
IMP
-positive GNR is evident at present, the rapid detection of such strains is necessary to help with infection control practices for the prevention of their dissemination and the transmission of the resistance gene to other pathogenic bacteria.
Population Pharmacokinetics of Rifabutin in Human Immunodeficiency Virus-Infected Patients
Authors: Gatti Giorgio , Papa Pietro , Torre Donato , Andreoni Massimo , Poggio Antonio , Bassetti Matteo , Marone Piero
Published: October 2018

Rifabutin pharmacokinetics were studied by the population approach (NONMEM) with 40 human immunodeficiency virus-infected patients receiving rifabutin at different doses for prophylaxis or therapy of mycobacterial infections. A two-compartment open model with first-order absorption was used as the structural pharmacokinetic model. Parameter estimates were the absorption rate constant (0.201/h), clearance/bioavailability (CL/
F
; 60.9 liters/h), volume of the central compartment/bioavailability (231 liters), intercompartmental clearance (60.3 liters/h), and volume of the peripheral compartment/bioavailability (

V
p
/F

; 1,050 liters). The distribution and elimination half-lives were 1.24 and 25.4 h, respectively. The covariates tested for influence on CL/
F
and

V
p
/F

were sex, age, weight, height, body surface area, tobacco smoking, drug addiction, alanine aminotransferase levels, creatinine clearance, total protein, bilirubin, numbers of CD4
+
cells, presence of diarrhea, cachexia index, rifabutin use (prophylaxis versus therapy), rifabutin dose, study site, and the concomitant administration of clarithromycin, fluconazole, phenobarbital, ciprofloxacin, azithromycin, or benzodiazepines. The only statistically significant effects on rifabutin pharmacokinetic parameters were a 27% decrease in

V
p
/F

due to the concomitant administration of azithromycin and a 39% increase in

V
p
/F

due to tobacco smoking. Such effects may be considered clinically unimportant. Our results confirm the lack of a correlation of rifabutin pharmacokinetic parameters with parameters of disease progression and gastrointestinal function. Also, the lack of a correlation with covariates which were previously found to be significant, such as concomitant fluconazole and clarithromycin use, may suggest that the effect of such covariates may be less important in the real clinical setting, in which several concomitant factors may influence pharmacokinetic parameters, with an overall effect of no apparent correlation.
In Vitro Susceptibility of
Mycobacterium ulcerans
to Clarithromycin
Authors: Portaels F. , Traore H. , De Ridder K. , Meyers W. M.
Published: October 2018

Buruli ulcer (BU), caused by
Mycobacterium ulcerans
, was recently recognized by the World Health Organization as an important emerging disease. While antimycobacterial therapy is often effective for the earliest nodular or ulcerative lesions, medical management of BU lesions in patients presenting for treatment is usually disappointing, leaving wide surgical excision the only alternative. Advanced ulcerated lesions of BU rarely respond to antimycobacterial agents; however, perioperative administration of such drugs may prevent relapses or disseminated infections. Clarithromycin possesses strong activity in vitro and in vivo against most nontuberculous mycobacteria. In this study we determined the antimycobacterial activity of this drug in vitro against 46 strains of
M. ulcerans
isolated from 11 countries. The MIC of clarithromycin was determined at pH 6.6 (on 7H11 agar) and at pH 7.4 (on Mueller-Hinton agar). The MICs ranged from 0.125 to 2 μg/ml at pH 6.6 and from <0.125 to 0.5 μg/ml at pH 7.4. For the majority of the strains, geographic origin did not play a significant role. Thirty-eight strains (83%) were inhibited by 0.5 μg/ml at pH 7.4. These MICs are below peak therapeutic concentrations of clarithromycin obtainable in blood. These results suggest that clarithromycin is a promising drug both for the treatment of early lesions of
M. ulcerans
and for the prevention of hematogenous dissemination of the etiologic agent during and after surgery. Studies should be initiated to evaluate the effects of clarithromycin in combination with ethambutol and rifampin on
M. ulcerans
both in vitro and in experimentally infected mice. Multidrug regimens containing clarithromycin may also help control the secondary bacterial infections sometimes seen in BU patients, most importantly osteomyelitis.
Confirmatory Evidence from 18S rRNA Gene Analysis for In Vivo Development of Propamidine Resistance in a Temporal Series of
Acanthamoeba
Ocular Isolates from a Patient
Authors: Ledee Dolena R. , Seal David V. , Byers Thomas J.
Published: October 2018

DNA sequences of three 18S rRNA gene alleles present in trophozoites obtained before and after therapy for
Acanthamoeba
keratitis substantiate a previous report that the infection was due to a single
Acanthamoeba
strain. Thus, the possibility that propamidine resistance which developed during therapy was due to a mixed infection was ruled out.
Influence of the TonB Energy-Coupling Protein on Efflux-Mediated Multidrug Resistance inPseudomonas aeruginosa
Authors: Zhao Qixun , Li Xian-Zhi , Mistry Anita , Srikumar Ramakrishnan , Zhang Li , Lomovskaya Olga , Poole Keith
Published: October 2018

TonB couples the energized state of the cytoplasmic membrane to the operation of outer membrane receptors responsible for Fe(III) siderophore uptake across the outer membrane of gram-negative bacteria. AtonBmutant ofPseudomonas aeruginosadeficient in iron siderophore uptake was shown in the present study to be hypersusceptible to a wide variety of antibiotics, reminiscent of the phenotype of mutants defective in themexAB-oprMantibiotic efflux operon. This was not related to influences of atonBmutation on the iron status of the cell, and indeed, intrinsic antibiotic susceptibility andmexAB-oprMexpression were unaffected by iron levels in the growth medium. The presence oftonBon a multicopy plasmid increased the level of resistance of a MexAB-OprM+strain but not that of a MexAB-OprM−strain to a variety of antimicrobial agents.mexAB-oprMexpression was not, however, altered in atonBdeletion mutant, indicating that any influence of TonB on MexAB-OprM-mediated multidrug resistance was at the level of pump activity. Consistent with this, drug accumulation assays revealed that thetonBdeletion mutant exhibited decreased levels of drug efflux. Still, the multidrug resistance of analBstrain was not wholly abrogated by atonBmutation, indicating that it is likely not an essential component of the efflux apparatus. Similarly, elimination oftonBfrom annfxBstrain only partially compromised MexCD-OprJ-mediated multidrug resistance. Intriguingly, the drug susceptibility of amexAB-oprMdeletion strain was increased following deletion oftonB, suggesting that TonB may also influence antibiotic resistance mediated by determinants other than MexAB-OprM (and MexCD-OprJ). Thus, TonB plays an important role in both intrinsic and acquired antibiotic resistance inP. aeruginosa.
Incidence of Foscarnet Resistance and Cidofovir Resistance in Patients Treated for Cytomegalovirus Retinitis
Authors: Jabs Douglas A. , Enger Cheryl , Forman Michael , Dunn J. P. , Retinitis for The Cytomegalovirus , Group Viral Resistance Study
Published: October 2018

Cytomegalovirus (CMV) retinitis is a common opportunistic infection in patients with AIDS. With long-term therapy for CMV retinitis, resistant CMV may develop. In a prospective study of 122 patients with CMV retinitis, 2.4 and 0.8% of patients had foscarnet-resistant blood culture isolates (50% inhibitory concentration [IC
50
], >400 μM) and urine culture isolates, respectively, at diagnosis of CMV retinitis prior to treatment, whereas 4.1 and 6.6% had cidofovir-resistant (IC
50
, >2 μM) blood and urine culture isolates, respectively. Patients were treated according to best medical judgement. Of 44 foscarnet-treated patients, 26% had a resistant blood or urine culture isolate by 6 months of treatment and 37% had a resistant isolate by 9 months; of 13 cidofovir-treated patients, 29% had a resistant blood or urine culture isolate by 3 months of therapy. The probabilities of developing foscarnet resistance while on foscarnet and developing cidofovir resistance while on cidofovir were not significantly different from that for developing ganciclovir resistance while on ganciclovir (odds ratios, 1.87 [
P
= 0.19] and 2.28 [
P
= 0.15], respectively).
Antimalarial Synergy of Cysteine and Aspartic Protease Inhibitors
Authors: Semenov Andrey , Olson Jed E. , Rosenthal Philip J.
Published: October 2018

It has been proposed that the
Plasmodium falciparum
cysteine protease falcipain and aspartic proteases plasmepsin I and plasmepsin II act cooperatively to hydrolyze hemoglobin as a source of amino acids for erythrocytic parasites. Inhibitors of each of these proteases have potent antimalarial effects. We have now evaluated the antimalarial effects of combinations of cysteine and aspartic protease inhibitors. When incubated with cultured
P. falciparum
parasites, cysteine and aspartic protease inhibitors exhibited synergistic effects in blocking parasite metabolism and development. The inhibitors also demonstrated apparent synergistic inhibition of plasmodial hemoglobin degradation both in culture and in a murine malaria model. When evaluated for the treatment of murine malaria, a combination of cysteine and aspartic protease inhibitors was much more effective than higher concentrations of either compound used alone. These results support a model whereby plasmodial cysteine and aspartic proteases participate in the degradation of hemoglobin, and they suggest that combination antimalarial therapy with inhibitors of the two classes of proteases is worthy of further study.
Influence of Assay Methodology on the Measurement of Free Serum Ceftriaxone Concentrations
Authors: Kohlhepp Sue J. , Gilbert David N. , Leggett James E.
Published: October 2018

The influence of assay methodology on the measurement of the active free fraction of ceftriaxone in plasma was determined. The free fraction was measured by three methods: agar diffusion bioassay, precipitation of plasma protein with methanol followed by high-performance liquid chromatography (HPLC) of the supernatant, and ultrafiltration of plasma followed by HPLC of the filtrate. In human serum, the free ceftriaxone levels were significantly lower (
P
= 0.03) when measured on ultrafiltrates compared to the other two methods. This difference disappeared when dolphin serum was studied. After ultrafiltration, human serum was shown, by Scatchard plot analysis, to have two ceftriaxone binding sites. Species differences were also demonstrated. Hence, in humans, determination of free plasma ceftriaxone varies with the assay method employed.
In Vitro Inactivation of
Chlamydia trachomatis
by Fatty Acids and Monoglycerides
Authors: Bergsson Gudmundur , Arnfinnsson Jóhann , Karlsson Sigfús M. , Steingrímsson Ólafur , Thormar Halldor
Published: October 2018

The antichlamydial effects of several fatty acids and monoglycerides were studied by incubating
Chlamydia trachomatis
bacteria with equal volumes of lipid solutions for 10 min and measuring the reduction in infectivity titer compared with that in a control solution without lipid. Caprylic acid (8:0), monocaprylin (8:0), monolaurin (12:0), myristic acid (14:0), palmitoleic acid (16:1), monopalmitolein (16:1), oleic acid (18:1), and monoolein (18:1) at concentrations of 20 mM (final concentration, 10 mM) had negligible effects on
C. trachomatis
. In contrast, lauric acid (12:0), capric acid (10:0), and monocaprin (10:0) caused a greater than 10,000-fold (>4-log
10
) reduction in the infectivity titer. When the fatty acids and monoglycerides were further compared at lower concentrations and with shorter exposure times, lauric acid was more active than capric acid and monocaprin was the most active, causing a greater than 100,000-fold (>5-log
10
) inactivation of
C. trachomatis
at a concentration of 5 mM for 5 min. The high levels of activity of capric and lauric acids and particularly that of monocaprin are notable and suggest that these lipids have specific antichlamydial effects. The mode of action of monocaprin was further studied by removal of the lipid by centrifugation before inoculation of
Chlamydia
onto host cells and by electron microscopy. The results indicate that the bacteria are killed by the lipid, possibly by disrupting the membrane(s) of the elementary bodies. A 50% effective concentration of 30 μg/ml was found by incubation of
Chlamydia
with monocaprin for 2 h. The rapid inactivation of large numbers of
C. trachomatis
organisms by monocaprin suggests that it may be useful as a microbicidal agent for the prevention of the sexual transmission of
C. trachomatis
.
Effect of Food on the Bioavailability of Stavudine in Subjects with Human Immunodeficiency Virus Infection
Authors: Kaul Sanjeev , Christofalo Barbara , Raymond Ralph H. , Stewart Michael B. , Macleod Catherine M.
Published: October 2018

A randomized, three-way crossover study was carried out to determine the effects of food ingestion on the pharmacokinetics of stavudine (d4T). Fifteen subjects with human immunodeficiency virus (HIV) infection and CD4
+
cell counts of ≥200/μl received 70 mg of d4T in a fasting state or 1 h before or 5 min after a standardized high-fat breakfast. A 7- to 15-day washout period was included between treatments. Blood and urine were collected before and for 10 h after dosing, and plasma and urine d4T concentrations were determined with a validated radioimmunoassay. Plasma drug concentration-time data were analyzed with a noncompartmental model. The mean maximum plasma drug concentration (
C
max
) and the time to
C
max
(
T
max
) for administration of d4T after a meal were significantly lower and longer (
P
= 0.0001 for both measures) than those observed in the fasting state, although the area under the concentration-time curve from time zero to infinity (AUC
0–∞
) was not significantly different. Neither of these parameters was significantly altered when d4T was taken 1 h before a meal. The bioavailability of d4T taken after a meal was 95% of that observed in the fasting state, and it was 97% when d4T was administered before a meal (
P
> 0.05 for both comparisons with the fasting state). The results of this study indicate that (i) ingestion of food does not affect the bioavailability of d4T and that patients with HIV infection can take it without regard to meals, and (ii) absorption is essentially complete within 1 h when d4T is administered in the fasted state.
Activity of Voriconazole Combined with Neutrophils or Monocytes against
Aspergillus fumigatus
: Effects of Granulocyte Colony-Stimulating Factor and Granulocyte-Macrophage Colony-Stimulating Factor
Authors: Vora Shefali , Chauhan Sharda , Brummer Elmer , Stevens David A.
Published: October 2018

Voriconazole (VCZ) was tested for antifungal activity against
Aspergillus fumigatus
hyphae alone or in combination with neutrophils or monocytes. Antifungal activity was measured as percent inhibition of hyphal growth in assays using the dye MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] or XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2
H
-tetrazolium-5-carboxanilide]. With both assays, VCZ inhibited hyphal growth at concentrations of <1 μg/ml and was almost as active as amphotericin B. VCZ (0.6 μg/ml) was sporicidal, as was amphotericin B (0.4 μg/ml). With both the MTT and XTT assays, neutrophils alone inhibited hyphae; when combined with VCZ, there was additive activity. Both granulocyte colony-stimulating factor- and granulocyte-macrophage colony-stimulating factor (GM-CSF)-treated polymorphonuclear neutrophils (PMN) had enhanced inhibition of hyphal growth. Moreover, such treatment of PMN also enhanced the collaboration of PMN with VCZ. Monocytes inhibited hyphal growth. When VCZ was combined with monocytes or monocytes were treated with GM-CSF, inhibition was significantly increased, to similar levels. However, the combination of VCZ with GM-CSF treatment of monocytes did not significantly increase the high-level inhibition by monocytes with either agent alone.
Pharmacokinetics of Meropenem in Critically Ill Patients with Acute Renal Failure Treated by Continuous Hemodiafiltration
Authors: Krueger W. A. , Schroeder T. H. , Hutchison M. , Hoffmann E. , Dieterich H.-J. , Heininger A. , Erley C. , Wehrle A. , Unertl K.
Published: October 2018

The pharmacokinetics of meropenem were studied in nine anuric critically ill patients treated by continuous venovenous hemodiafiltration. Peak levels after infusion of 1,000 mg over 30 min amounted to 103.2 ± 45.9 μg/ml, and trough levels at 12 h were 9.6 ± 3.8 μg/ml. A dosage of 1,000 mg of meropenem twice a day provides plasma drug levels covering intermediately susceptible microorganisms. Further reductions of the dosage might be appropriate for highly susceptible bacteria or when renal replacement therapies with lower clearances are applied.
Prevalence and Characterization of the Mechanisms of Macrolide, Lincosamide, and Streptogramin Resistance in Isolates of
Streptococcus pneumoniae
Authors: Johnston Nicole J. , de Azavedo Joyce C. , Kellner James D. , Low Donald E.
Published: October 2018

Of a total of 147 erythromycin-resistant
Streptococcus pneumoniae
isolates, 64 (43.5%) were resistant to erythromycin, clindamycin, and streptogramin B (MLS
B
phenotype), 57 of which possessed the
ermB
gene. Eighty-two (55.8%) were resistant to erythromycin alone (M phenotype), 81 of which possessed the
mefE
gene. One was erythromycin and streptogramin B resistant but susceptible to clindamycin (MS phenotype) and possessed neither the
erm
gene nor the
mefE
gene.