bluuebunny/crossref_metadata_2025 huggingface dataset:
Efficacy of the Carbocyclic 2′-Deoxyguanosine Nucleoside BMS-200475 in the Woodchuck Model of Hepatitis B Virus Infection
Authors: Genovesi E. V. , Lamb L. , Medina I. , Taylor D. , Seifer M. , Innaimo S. , Colonno R. J. , Standring D. N. , Clark J. M.
Published: October 2018

Daily oral treatment with the cyclopentyl 2′-deoxyguanosine nucleoside BMS-200475 at doses ranging from 0.02 to 0.5 mg/kg of body weight for 1 to 3 months effectively reduced the level of woodchuck hepatitis virus (WHV) viremia in chronically infected woodchucks as measured by reductions in serum WHV DNA levels and endogenous hepadnaviral polymerase activity. Within 4 weeks of daily therapy with 0.5 or 0.1 mg of BMS-200475 per kg, endogenous viral polymerase levels in serum were reduced about 1,000-fold compared to pretreatment levels. Serum WHV DNA levels determined by a dot blot hybridization technique were comparably decreased in these treated animals. In the 3-month study, the sera of animals that had undetectable levels of WHV DNA by the dot blot technique were further analyzed by a highly sensitive semiquantitative PCR assay. The results indicate that BMS-200475 therapy reduced mean WHV titers by 10
7
- to 10
8
-fold, down to levels as low as 10
2
to 10
3
virions/ml of serum. Southern blot hybridization analysis of liver biopsy samples taken from animals during and after BMS-200475 treatment showed remarkable reductions in the levels of WHV DNA replicative intermediates and in the levels of covalently closed circular viral DNA. WHV viremia in BMS-200475-treated WHV carriers eventually returned to pretreatment levels after therapy was stopped. These results indicate that BMS-200475 should be evaluated in clinical trials for the therapy of chronic human hepatitis B virus infections.
ABT-378, a Highly Potent Inhibitor of the Human Immunodeficiency Virus Protease
Authors: Sham Hing L. , Kempf Dale J. , Molla Akhteruzammen , Marsh Kennan C. , Kumar Gondi N. , Chen Chih-Ming , Kati Warren , Stewart Kent , Lal Ritu , Hsu Ann , Betebenner David , Korneyeva Marina , Vasavanonda Sudthida , McDonald Edith , Saldivar Ayda , Wideburg Norm , Chen Xiaoqi , Niu Ping , Park Chang , Jayanti Venkata , Grabowski Brian , Granneman G. Richard , Sun Eugene , Japour Anthony J. , Leonard John M. , Plattner Jacob J. , Norbeck Daniel W.
Published: October 2018

The valine at position 82 (Val 82) in the active site of the human immunodeficiency virus (HIV) protease mutates in response to therapy with the protease inhibitor ritonavir. By using the X-ray crystal structure of the complex of HIV protease and ritonavir, the potent protease inhibitor ABT-378, which has a diminished interaction with Val 82, was designed. ABT-378 potently inhibited wild-type and mutant HIV protease (

K
i

= 1.3 to 3.6 pM), blocked the replication of laboratory and clinical strains of HIV type 1 (50% effective concentration [EC
50
], 0.006 to 0.017 μM), and maintained high potency against mutant HIV selected by ritonavir in vivo (EC
50
, ≤0.06 μM). The metabolism of ABT-378 was strongly inhibited by ritonavir in vitro. Consequently, following concomitant oral administration of ABT-378 and ritonavir, the concentrations of ABT-378 in rat, dog, and monkey plasma exceeded the in vitro antiviral EC
50
in the presence of human serum by >50-fold after 8 h. In healthy human volunteers, coadministration of a single 400-mg dose of ABT-378 with 50 mg of ritonavir enhanced the area under the concentration curve of ABT-378 in plasma by 77-fold over that observed after dosing with ABT-378 alone, and mean concentrations of ABT-378 exceeded the EC
50
for >24 h. These results demonstrate the potential utility of ABT-378 as a therapeutic intervention against AIDS.
An
Escherichia coli
Expression Assay and Screen for Human Immunodeficiency Virus Protease Variants with Decreased Susceptibility to Indinavir
Authors: Melnick Laurence , Yang Shiow-Shong , Rossi Rick , Zepp Charlie , Heefner Donald
Published: October 2018

We have developed a recombinant
Escherichia coli
screening system for the rapid detection and identification of amino acid substitutions in the human immunodeficiency virus (HIV) protease associated with decreased susceptibility to the protease inhibitor indinavir (MK-639; Merck & Co.). The assay depends upon the correct processing of a segment of the HIV-1 HXB2
gag-pol
polyprotein followed by detection of HIV reverse transcriptase activity by a highly sensitive, colorimetric enzyme-linked immunosorbent assay. The highly sensitive system detects the contributions of single substitutions such as I84V, L90M, and L63P. The combination of single substitutions further decreases the sensitivity to indinavir. We constructed a library of HIV protease variant genes containing dispersed mutations and, using the
E. coli
recombinant system, screened for mutants with decreased indinavir sensitivity. The discovered HIV protease variants contain amino acid substitutions commonly associated with indinavir resistance in clinical isolates, including the substitutions L90M, L63P, I64V, V82A, L24I, and I54T. One substitution, W6R, is also frequently found by the screen and has not been reported elsewhere. Of a total of 12,000 isolates that were screened, 12 protease variants with decreased sensitivity to indinavir were found. The L63P substitution, which is also associated with indinavir resistance, increases the stability of the isolated protease relative to that of the native HXB2 protease. The rapidity, sensitivity, and accuracy of this screen also make it useful for screening for novel inhibitors. We have found the approach described here to be useful for the detection of amino acid substitutions in HIV protease that have been associated with drug resistance as well as for the screening of novel compounds for inhibitory activity.
In Vitro Activities of Ketolide HMR 3647, Macrolides, and Clindamycin against Coryneform Bacteria
Authors: Martínez-Martínez Luis , Pascual Alvaro , Suárez Ana Isabel , Perea Evelio J.
Published: October 2018

The in vitro activity of ketolide HMR 3647 against coryneform bacteria isolated from clinical samples was evaluated. Except against
Corynebacterium jeikeium
and
C. urealyticum
, HMR 3647 showed high activity against
Corynebacterium
spp., being more active than 14- and 16-membered macrolides, azithromycin, or clindamycin. HMR 3647 also had high in vitro activity against
Brevibacterium
spp. and
Listeria monocytogenes
.
In Vitro Susceptibilities of the Microsporidia
Encephalitozoon cuniculi
,
Encephalitozoon hellem
, and
Encephalitozoon intestinalis
to Albendazole and Its Sulfoxide and Sulfone Metabolites
Authors: Ridoux Olivier , Drancourt Michel
Published: October 2018

In vitro comparisons demonstrated that the efficacy of albendazole, albendazole-sulfoxide, and albendazole-sulfone against pathogenic
Encephalitozoon
species was proportional to the degree of oxidation at a concentration of >10
−3
μg/ml. Furthermore, at a concentration of <10
−2
μg/ml, benzimidazoles were more effective against
Encephalitozoon cuniculi
and
Encephalitozoon hellem
than against
Encephalitozoon intestinalis
.
Efficacies of Cefepime, Ceftazidime, and Imipenem Alone or in Combination with Amikacin in Rats with Experimental Pneumonia Due to Ceftazidime-Susceptible or -Resistant
Enterobacter cloacae
Strains
Authors: Mimoz Olivier , Jacolot Anne , Leotard Sophie , Hidri Nadia , Samii Kamran , Nordmann Patrice , Petitjean Olivier
Published: October 2018

The antibacterial activities of human regimens of cefepime, ceftazidime, and imipenem alone or in combination with amikacin against an isogenic pair of
Enterobacter cloacae
strains (wild type and its corresponding derepressed cephalosporinase mutant) were compared by using our nonlethal model of pneumonia with 180 immunocompetent rats. Compared with untreated animals, all β-lactam-treated rats, except those inoculated with the mutant isolate and receiving ceftazidime, had significantly lower bacterial counts in their lungs 60 h after the onset of therapy. Although the combination of a β-lactam and amikacin was more bactericidal than each corresponding antimicrobial agent alone, true synergy was noted only with cefepime and imipenem against the constitutive derepressed strain.
Amphotericin B Colloidal Dispersion Combined with Flucytosine with or without Fluconazole for Treatment of Murine Cryptococcal Meningitis
Authors: Diamond DeAnn M. , Bauer Madeline , Daniel Barbra E. , Leal Mary Ann E. , Johnson Debra , Williams Byron K. , Thomas Ann M. , Ding James C. , Najvar Laura , Graybill J. Richard , Larsen Robert A.
Published: October 2018

Studies with animals and in vitro studies have demonstrated that flucytosine plus amphotericin B or fluconazole has significantly improved mycologic activity against meningitis caused by
Cryptococcus neoformans
compared to the activity of amphotericin B or fluconazole used alone. However, few doses have been tested in combination. This study evaluated the antifungal efficacy of amphotericin B colloidal dispersion (ABCD) combined with flucytosine with and without fluconazole in a murine model of cryptococcal meningitis. The following dosages were tested: ABCD at 0 to 12.5 mg/kg of body weight given intravenously 3 days/week, flucytosine at 0 to 110 mg/kg/day, and fluconazole at 0 to 50 mg/kg/day. Meningitis was established in male BALB/c mice by intracerebral injection of
C. neoformans
. Treatment with flucytosine with or without fluconazole dissolved in the sole source of drinking water was started on day 2; animals were sacrificed at 16 days, and the numbers of fungal colonies in the brain were quantified. A survival rate of 100% was achieved with ABCD plus flucytosine without fluconazole; however, the addition of fluconazole was required to prevent weight loss (
P
< 0.00001) and to achieve the maximum antifungal effect (
P
< 0.00001). The only region of dose combinations for which the 99% confidence intervals were less than 100 CFU/g of brain was defined by ABCD at 5.0 to 7.5 mg/kg combined with flucytosine at 20 to 60 mg/kg/day and fluconazole at 30 to 40 mg/kg/day. The triple combination of ABCD plus flucytosine and fluconazole was necessary to achieve the greatest antifungal activity.
The Gene Encoding the Low-Affinity Penicillin-Binding Protein 3r in
Enterococcus hirae
S185R Is Borne on a Plasmid Carrying Other Antibiotic Resistance Determinants
Authors: Raze Dominique , Dardenne Olivier , Hallut Séverine , Martinez-Bueno Manuel , Coyette Jacques , Ghuysen Jean-Marie
Published: October 2018

Two plasmid-derived
Nco
I DNA fragments of 14 and 4.5 kb, respectively, have been isolated from the multidrug-resistant strain
Enterococcus hirae
S185R and analyzed. The 14-kb fragment contains two inverted (
L
and
R
) IS
1216
insertion modules of the ISS
1
family. These modules define a Tn
5466
transposon-like structure that contains one copy of the methylase-encoding
ermAM
conferring erythromycin resistance and one copy of the adenylyl-transferase-encoding
aadE
conferring streptomycin resistance. Immediately on the left side of IS
1216L
there occurs a copy of
pbp3r
encoding the low-affinity penicillin-binding protein (PBP) PBP3r, itself preceded by a
psr
-like gene (
psr3r
) that controls the synthesis of PBP3r.
ermAM
,
aadE
, and the transposase gene (
tnp
) of IS
1216R
have the same polarities, and these are opposite those of
psr3r
,
pbp3r
, and the
tnp
gene of IS
1216L
. The 4.5-kb fragment is a copy of the 4.5-kb sequence at the 5′ end of the 14-kb fragment, although it is not a restriction product of the 14-kb fragment. It contains three genes with the same polarity:
psr3r
,
pbp3r
, and
tnp
in an IS
1216
element. Because of the very high degree of identity (99%) with the chromosomal
psrfm
and
pbp5fm
genes of
Enterococcus faecium
D63R, it is proposed that both the
psr3r
and
pbp3r
genes were transferred from an
E. faecium
strain and inserted in a plasmid of
E. hirae. E. hirae
is the first known bacterial species in which a low-affinity PBP-encoding gene has been found to be plasmid borne.
Rifampin Increases Cytokine-Induced Expression of the CD1b Molecule in Human Peripheral Blood Monocytes
Authors: Tentori L. , Graziani G. , Porcelli S. A. , Sugita M. , Brenner M. B. , Madaio R. , Bonmassar E. , Giuliani A. , Aquino A.
Published: October 2018

In recent years, it has been shown that a nonclassical, major histocompatibility complex-independent system (i.e., CD1-restricted T-cell responses) is involved in T-cell immunity against nonpeptide antigens. The CD1 system appears to function by presenting microbial lipid antigens to specific T cells, and the antigens so far identified include several known constituents of mycobacterial cell walls. Among the four known human CD1 isoforms, the CD1b protein is the best characterized with regard to its antigen-presenting function. Expression of CD1b is upregulated on human blood monocytes upon exposure to granulocyte/macrophage-colony stimulating factor, alone or in combination with interleukin-4 (IL-4) (S. A. Porcelli, Adv. Immunol. 59:1–98, 1995). Rifampin (RFP) and its derivatives are widely used for chemoprophylaxis or chemotherapy against
Mycobacterium tuberculosis
. However, this agent was found to reduce the mitogen responsiveness of human B and T lymphocytes, chemotaxis, and delayed-type hypersensitivity. The present study extends the immunopharmacological profile of RFP by examining its effects on CD1b expression by human peripheral blood monocytes exposed to GM-CSF plus IL-4. The results showed that clinically attainable concentrations (i.e., 2 or 10 μg/ml for 24 h) of the agent produced a marked increase in CD1b expression on the plasma membrane, as evaluated by fluorescence-activated cell sorter analysis, whereas it had no effect on cytosolic fractions, as indicated by Western blot analysis. This was found to be the result of increased CD1b gene expression, as shown by Northern blot analysis of CD1b mRNA. These results suggest that RFP could be of potential value in augmenting the CD1b-restricted antigen recognition system, thereby enhancing protective cellular immunity to
M. tuberculosis.
Reduced Clinical Efficacy of Pazufloxacin against Gonorrhea Due to High Prevalence of Quinolone-Resistant Isolates with the GyrA Mutation
Authors: Tanaka Masatoshi , Matsumoto Tetsuro , Sakumoto Misao , Takahashi Koichi , Saika Takeshi , Kabayashi Intetsu , Kumazawa Joichi , Group The Pazufloxacin Std
Published: October 2018

Forty-two men with gonococcal urethritis were treated with an oral dosage of 200 mg of pazufloxacin, a new fluoroquinolone, three times daily for 3 days. Only 28 of the 42 men (66.7%) had negative culture results for
Neisseria gonorrhoeae
during follow-up. Of the 42 isolates, 41 could be recultured for antibiotic susceptibility testing and DNA sequencing. In 26 of the 41 isolates (63.4%), GyrA mutations with or without ParC mutations were identified. Among the 26 isolates, 23 contained a single GyrA mutation, 1 contained two GyrA mutations, and 2 contained three mutations including double GyrA and single ParC mutations. A single Ser-91-to-Phe mutation, which was detected in 14 of the 26 isolates, was the most common GyrA mutation, followed by an Ala-75 to Ser mutation and an Asp-95 to Asn or Gly mutation in GyrA. All three isolates with two or three mutations contained the Ser-91-to-Phe GyrA mutation. Eleven of the 14 isolates with the single Ser-91-to-Phe mutation within GyrA and all 3 isolates with two or three mutations persisted after pazufloxacin treatment. On the other hand, all 15 wild-type and 9 mutant isolates with a substitution at codon Ala-75 or Asp-95 were eradicated. The mean MIC of pazufloxacin for mutants with the single Ser-91-to-Phe mutation in GyrA was 66-fold higher than that for the wild type. The results obtained in this study suggest that a high prevalence of fluoroquinolone-resistant gonococcal isolates with the Ser-91-to-Phe mutation in GyrA reduced the efficacy of pazufloxacin as treatment for gonococcal urethritis.
Antibiotic Treatment of Experimental Pneumonic Plague in Mice
Authors: Byrne William R. , Welkos Susan L. , Pitt M. Louise , Davis Kelly J. , Brueckner Ralf P. , Ezzell John W. , Nelson Gene O. , Vaccaro Joseph R. , Battersby Luann C. , Friedlander Arthur M.
Published: October 2018

A mouse model was developed to evaluate the efficacy of antibiotic treatment of pneumonic plague; streptomycin was compared to antibiotics with which there is little or no clinical experience. Infection was induced by inhalation of aerosolized
Yersinia pestis
organisms. Antibiotics were administered by intraperitoneal injection every 6 hours for 5 days, at doses that produced levels of drug in serum comparable to those observed in humans treated for other serious infections. These studies compared in vitro to in vivo activity and evaluated the efficacy of antibiotics started at different times after exposure. Early treatment (started 24 h after challenge, when 0 of 10 mice tested had positive blood cultures) with netilmicin, ciprofloxacin, ofloxacin, ceftriaxone, ceftazidime, aztreonam, ampicillin, and rifampin (but not cefazolin, cefotetan, or ceftizoxime) demonstrated efficacy comparable to streptomycin. Late treatment (started 42 h after exposure, when five of five mice tested had positive blood cultures) with netilmicin, ciprofloxacin, ofloxacin, and a high dose (20 mg/kg of body weight every 6 h) of gentamicin produced survival rates comparable to that with streptomycin, while all of the beta-lactam antibiotics (cefazolin, cefotetan, ceftriaxone, ceftazidime, aztreonam, and ampicillin) and rifampin were significantly inferior to streptomycin. In fact, all groups of mice treated late with beta-lactam antibiotics experienced accelerated mortality rates compared to normal-saline-treated control mice. These studies indicate that netilmicin, gentamicin, ciprofloxacin, and ofloxacin may be alternatives for the treatment of pneumonic plague in humans. However, the beta-lactam antibiotics are not recommended, based upon poor efficacy in this mouse model of pneumonic plague, particularly when pneumonic plague may be associated with bacteremia.
Effect of Terbinafine on Theophylline Pharmacokinetics in Healthy Volunteers
Authors: Trépanier Eric F. , Nafziger Anne N. , Amsden Guy W.
Published: October 2018

Twelve healthy volunteers were enrolled in an open-label, randomized, crossover study. Subjects received single doses of theophylline (5 mg/kg) with and without multiple-dose terbinafine, and 11 blood samples were collected over 24 h. The study phases were separated by a 4-week washout period. Theophylline serum data were modeled via noncompartmental analysis. When the control phase (i.e., no terbinafine) was compared to the treatment phase (terbinafine), theophylline exposure (the area under the serum concentration-time curve from time zero to infinity) increased by 16% (
P
= 0.03), oral clearance decreased by 14% (
P
= 0.04), and half-life increased by 24% (
P
= 0.002). No significant changes in other theophylline pharmacokinetic parameters were evident.
Antimicrobial Resistance in Enterococci Isolated from Turkey Flocks Fed Virginiamycin
Authors: Welton L. A. , Thal L. A. , Perri M. B. , Donabedian S. , McMahon J. , Chow J. W. , Zervos M. J.
Published: October 2018

From 125 separate cloacal cultures from three turkey flocks fed virginiamycin, 104
Enterococcus faecium
and 186
Enterococcus faecalis
isolates were obtained. As the turkeys aged, there was a higher percentage of quinupristin-dalfopristin-resistant
E. faecium
isolates, with isolates from the oldest flock being 100% resistant. There were no vancomycin-resistant enterococci. Results of pulsed-field gel electrophoresis (PFGE) indicated there were 11 PFGE types of
E. faecalis
and 7 PFGE types of
E. faecium
that were in more than one group of flock cultures.
Relationship between Didanosine Exposure and Surrogate Marker Response in Human Immunodeficiency Virus-Infected Outpatients
Authors: Adams John M. , Shelton Mark J. , Hewitt Ross G. , Grasela Thaddeus H. , DeRemer Mary , Morse Gene D.
Published: October 2018

We used information available from routine clinic visits to characterize the pharmacokinetics of didanosine in 82 human immunodeficiency virus-infected patients. A total of 271 blood samples were collected for the measurement of didanosine concentrations in plasma (mean ± standard deviation [SD], 3.30 ± 2.21 samples/patient). Bayesian estimates of didanosine oral clearance (CL
oral
) were obtained for these patients by the POSTHOC option within the NONMEM software package. Population priors from a previous NONMEM analysis of didanosine pharmacokinetics were used. The mean ± SD CL
oral
was 132 ± 27.7 liters/h, which agrees reasonably well with estimates obtained from previous pharmacokinetic studies of didanosine. Estimates of individual didanosine exposure were then used to consider potential relationships between drug exposure and surrogate marker response over a 6-month period. No correlations were found between the didanosine area under the concentration-time curve from 0 to 6 months and the absolute CD4 cell count (
r
= 0.305; 0.1 <
P
< 0.2), weight response (
r
= 0.0857;
P
> 0.4), or percentage of CD4 lymphocytes (
r
= 0.0559;
P
> 0.4). Future efforts to characterize didanosine exposure in outpatients by random sampling methods should involve more directed efforts to limit residual variability in the data.
Cefotaxime-Resistant
Enterobacteriaceae
Isolates from a Hospital in Warsaw, Poland: Identification of a New CTX-M-3 Cefotaxime-Hydrolyzing β-Lactamase That Is Closely Related to the CTX-M-1/MEN-1 Enzyme
Authors: Gniadkowski Marek , Schneider Ines , Pałucha Andrzej , Jungwirth Renate , Mikiewicz Barbara , Bauernfeind Adolf
Published: October 2018

A group of cefotaxime-resistant
Citrobacter freundii
and
Escherichia coli
isolates were collected by a clinical laboratory in a hospital in Warsaw, Poland, in July 1996. Detailed analysis has shown that all of these produced a β-lactamase (pI, 8.4) belonging to the CTX-M family, one of the minor extended-spectrum β-lactamase families with a strong cefotaxime-hydrolyzing activity. Sequencing has revealed that
C. freundii
isolates produced a new CTX-M-3 enzyme which is very closely related to the CTX-M-1/MEN-1 β-lactamase, sporadically identified in Europe over a period of 6 years. Amino acid sequences of these two β-lactamases differ at four positions: Val77Ala, Asp114Asn, Ser140Ala, and Asn288Asp (the first amino acid of each pair refers to CTX-M-1/MEN-1 and second refers to CTX-M-3). The partial sequence of the
E. coli
CTX-M gene was identical to the corresponding region of
bla
CTX-M-3
, but a transconjugant of the
E. coli
isolate expressed higher levels of resistance to β-lactams than did
C. freundii
transconjugants. These resistance differences correlated with differences in plasmid DNA restriction patterns. Our results suggest that CTX-M genes have been spread among different species of the family
Enterobacteriaceae
in the hospital and that the CTX-M-3-expressing
C. freundii
strain causing routine urinary tract infections has been maintained for a relatively long time in the hospital environment.
Unique Metabolism of a Novel Antiviral
l
-Nucleoside Analog, 2′-Fluoro-5-Methyl-β-
l
-Arabinofuranosyluracil: a Substrate for Both Thymidine Kinase and Deoxycytidine Kinase
Authors: Liu Shwu-Huey , Grove Kristie L. , Cheng Yung-Chi
Published: October 2018

2′-Fluoro-5-methyl-β-
l
-arabinofuranosyluracil (
l
-FMAU) is the first
l
-nucleoside analog with low cytotoxicity discovered to have potent antiviral activities against both hepatitis B virus and Epstein-Barr virus but not human immunodeficiency virus. This spectrum of activity is different from those of the other
l
-nucleoside analogs examined.
l
-FMAU enters cells through equilibrative-sensitive and -insensitive nucleoside transport as well as through nonfacilitated passive diffusion.
l
-FMAU is phosphorylated stepwise in cells to its mono-, di-, and triphosphate forms. In the present study the enzymes responsible for the first step of
l
-FMAU phosphorylation were identified. This is the first thymidine analog shown to be a substrate not only for cytosolic thymidine kinase and mitochondrial deoxypyrimidine kinase but also for deoxycytidine kinase. This finding suggests that the antiviral activity of
l
-FMAU will not be limited by the loss or alteration of any of these deoxynucleoside kinases.
Cyclosporin Analogs Inhibit In Vitro Growth ofCryptosporidium parvum
Authors: Perkins Margaret E. , Wu Teresa W. , Le Blancq Sylvie M.
Published: October 2018

Cyclosporine and nonimmunosuppressive cyclosporin (CS) analogs were demonstrated to be potent inhibitors of the growth of the intracellular parasiteCryptosporidium parvumin short-term (48-h) in vitro cultures. Fifty-percent inhibitory concentrations (IC50s) were 0.4 μM for SDZ 033-243, 1.0 μM for SDZ PSC-833, and 1.5 μM for cyclosporine. Two other analogs were less effective than cyclosporine: the IC50of SDZ 205-549 was 5 μM, and that of SDZ 209-313 was 7 μM. These were much lower than the IC50of 85 μM of paromomycin, a standard positive control for in vitro drug assays for this parasite. In addition, intracellular growth of excysted sporozoites that had been incubated for 1 h in cyclosporine was significantly reduced, suggesting that the drug can inhibit sporozoite invasion. The cellular activities of the CS analogs used have been characterized for mammalian cells and protozoa. The two analogs that were most active in inhibitingC. parvum, SDZ PSC-833 and SDZ 033-243, bind weakly to cyclophilin, a peptidyl proline isomerase which is the primary target of cyclosporine and CS analogs. However, they are potent modifiers of the activity of the P glycoproteins/multidrug resistance (MDR) transporters, members of the ATP-binding cassette (ABC) superfamily. Hence, both cyclophilin and some ABC transporters may be targets for this class of drugs, although drugs that preferentially interact with the latter are more potent. Cyclosporine (0.5 μM) had no significant chemosensitizing activity. That is, it did not significantly increase sensitivity to paromomycin, suggesting that an ABC transporter is not critical in the efflux of this drug. Cyclosporine at concentrations up to 50 μM was not toxic to host Caco-2 cells in the CellTiter 96 assay. The results of this study complement those of studies of the inhibitory effect of cyclosporine and CS analogs on other apicomplexan parasites,Plasmodium falciparum,Plasmodium vivax, andToxoplasma gondii.
Impact of Bacterial Biofilm Formation on In Vitro and In Vivo Activities of Antibiotics
Authors: Schwank Silvia , Rajacic Zarko , Zimmerli Werner , Blaser Jürg
Published: October 2018

The impact of bacterial adherence on antibiotic activity was analyzed with two isogenic strains of
Staphylococcus epidermidis
that differ in the features of their in vitro biofilm formation. The eradication of bacteria adhering to glass beads by amikacin, levofloxacin, rifampin, or teicoplanin was studied in an animal model and in a pharmacokinetically matched in vitro model. The features of
S. epidermidis
RP62A that allowed it to grow on surfaces in multiple layers promoted phenotypic resistance to antibiotic treatment, whereas strain M7 failed to accumulate, despite initial adherence on surfaces and growth in suspension similar to those for RP62A. Biofilms of
S. epidermidis
M7 were better eradicated than those of strain RP62A in vitro (46 versus 31%;
P
< 0.05) as well as in the animal model (39 versus 9%;
P
< 0.01).
Fluconazole Pharmacokinetics in Burn Patients
Authors: Boucher Bradley A. , King Stephen R. , Wandschneider Heidi L. , Hickerson William L. , Hanes Scott D. , Herring Vanessa L. , Canada Todd W. , Hess Mary M.
Published: October 2018

The pharmacokinetics of fluconazole in nine adult patients with severe (30 to 95% total body surface area) burns were studied. There was no significant difference in half-life (t1/2), clearance (CL), or volume of distribution (V) over time in five patients on days 3 and 8 of the study (P> 0.05). Combined parameter estimates (means ± standard deviations) for all nine patients for the two study periods were as follows:t1/2, 24.4 ± 5.8 h; CL, 0.36 ± 0.09 ml/min/kg; andV, 0.72 ± 0.12 liters/kg. These estimates oft1/2and CL in burn patients were approximately 13% shorter and 30% more rapid, respectively, than the most extreme estimates reported for other populations.
Ribosomes from an Oxazolidinone-Resistant Mutant Confer Resistance to Eperezolid in a
Staphylococcus aureus
Cell-Free Transcription-Translation Assay
Authors: Murray Robert W. , Schaadt Ronda D. , Zurenko Gary E. , Marotti Keith R.
Published: October 2018

Oxazolidinone-resistant mutants of
Staphylococcus aureus
, isolated with a spiral plating technique, had a 16-fold higher MIC (2 versus 32 μg/ml) of eperezolid when compared to the parental sensitive strain. Eperezolid inhibited in vitro protein translation with 50% inhibitory concentrations of 30 μM for the oxazolidinone-sensitive S30 extract and 75 μM for the resistant extract. Experiments mixing various combinations of S100 and crude ribosome preparations from oxazolidinone-sensitive and -resistant
S. aureus
strains in a transcription-translation assay demonstrated that the resistant determinant resided within the ribosomal fraction. Ribosomes from the oxazolidinone-resistant strain bound less drug than ribosomes from the sensitive strain, indicating that the ribosome is the site of action for the oxazolidinones. These experiments demonstrate that an alteration of the ribosome is responsible for some or all of the oxazolidinone resistance observed in the
S. aureus
mutant.
Transferable, Plasmid-Mediated VanB-Type Glycopeptide Resistance in
Enterococcus faecium
Authors: Rice Louis B. , Carias Lenore L. , Donskey Curtis L. , Rudin Susan D.
Published: October 2018

An approximately 60-kb transferable,
vanB
-carrying plasmid has been identified in a clinical
Enterococcus faecium
strain. A similar plasmid has been observed in an unrelated
E. faecium
strain, suggesting that plasmid transfer of
vanB
operons occurs in nature and plays a role in the dissemination of VanB-type resistance among strains of
E. faecium.
Alginate Lyase Promotes Diffusion of Aminoglycosides through the Extracellular Polysaccharide of MucoidPseudomonas aeruginosa
Authors: Hatch Richard A. , Schiller Neal L.
Published: October 2018

We demonstrated that a 2% suspension ofPseudomonas aeruginosaalginate completely blocked the diffusion of gentamicin and tobramycin, but not that of carbenicillin, illustrating how alginate production can help protectP. aeruginosagrowing within alginate microcolonies in patients with cystic fibrosis (CF) from the effects of aminoglycosides. This aminoglycoside diffusion barrier was degraded with a semipurified preparation ofP. aeruginosaalginate lyase, suggesting that this enzyme deserves consideration as an adjunctive agent for CF patients colonized by mucoid strains ofP. aeruginosa.
Susceptibilities of Clinical and Laboratory Isolates of
Blastomyces dermatitidis
to Ketoconazole, Itraconazole, and Fluconazole
Authors: Chapman Stanley W. , Rogers P. David , Rinaldi Michael G. , Sullivan Donna C.
Published: October 2018

Eighteen isolates of
Blastomyces dermatitidis
were evaluated for their in vitro susceptibilities to ketoconazole, itraconazole, and fluconazole. The MIC ranges were 0.1 to 0.4 μg/ml for ketoconazole, ≤0.018 to 0.07 μg/ml for itraconazole, and 2.5 to 4.0 μg/ml for fluconazole. The ranges for the minimal lethal concentrations were 0.2 to 0.8 μg/ml for ketoconazole, ≤0.018 to 0.07 μg/ml for itraconazole, and 10 to 40 μg/ml for fluconazole. Itraconazole was the most active agent against
B. dermatitidis
in vitro, while fluconazole was the least active. These results correlate with the clinical efficacies noted to date with doses of these agents used to treat blastomycosis.
Fosfomycin Reduces CD15s-Related Antigen Expression ofStreptococcus pyogenes
Authors: Hirota Katsuhiko , Murakami Kinya , Nemoto Ken , Ono Tsuneko , Matsuo Takashi , Kumon Hiromi , Miyake Yoichiro
Published: October 2018

We have previously shown the immunological mimicry of human sialyl-Lewisx(CD15s) by a surface antigen ofStreptococcus pyogenes. This mimicking surface antigen may act as a ligand to the selectin family and may induce antibody production against CD15s on host cells, suggesting a possible role in the pathogenesis ofS. pyogenes. In this study, the effects of antibiotics on the CD15s-related antigen expression ofS. pyogeneswere examined at a concentration below the MIC (sub-MIC). The amounts of CD15s on the surfaces ofS. pyogenescells and on the surfaces ofS. pyogenesbiofilms were determined by a whole-cell enzyme-linked immunosorbent assay and by laser scanning fluorescence microscopy, respectively, by using an anti-CD15s monoclonal antibody. At the sub-MICs, fosfomycin (1R,2S-1,2-epoxypropyl phosphonic acid), its enantiomer (1S,2R-1,2-epoxypropyl phosphonic acid), and benzylpenicillin significantly inhibited the CD15s expression of all strains studied. The effects of fosfomycin and its enantiomer on biofilms were also observed by scanning electron microscopy. Incubation ofS. pyogeneswith the sub-MIC of fosfomycin or its enantiomer, which has no antibacterial activity, reduced the amount of CD15s on the biofilm surface and made it smooth. These results suggest that fosfomycin or its enantiomer might be useful for preventingS. pyogenesadherence to human CD15s receptors and the resulting immunological pathogenicity.
Characterization of Vancomycin-Resistant
Enterococcus faecium
Isolates from the United States and Their Susceptibility In Vitro to Dalfopristin-Quinupristin
Authors: Eliopoulos G. M. , Wennersten C. B. , Gold H. S. , Schülin T. , Souli M. , Farris M. G. , Cerwinka S. , Nadler H. L. , Dowzicky M. , Talbot G. H. , Moellering R. C.
Published: October 2018

In the course of clinical studies with the investigational streptogramin antimicrobial dalfopristin-quinupristin, isolates of vancomycin-resistant
Enterococcus faecium
were referred to our laboratory from across the United States. Seventy-two percent of the strains were of the VanA type, phenotypically and genotypically, while 28% were of the VanB type. High-level resistance to streptomycin or gentamicin was observed in 86 and 81%, respectively, of the VanA strains but in only 69 and 66%, respectively, of the VanB strains. These enterococci were resistant to ampicillin (MIC for 50% of the isolates tested [MIC
50
] and MIC
90
, 128 and 256 μg/ml, respectively) and to the other approved agents tested, with the exception of chloramphenicol (MIC
90
, 8 μg/ml) and novobiocin (MIC
90
, 1 μg/ml). Considering all of the isolates submitted, dalfopristin-quinupristin inhibited 86.4% of them at concentrations of ≤1 μg/ml and 95.1% of them at ≤2 μg/ml. However, for the data set comprised of only the first isolate submitted for each patient, 94.3% of the strains were inhibited at concentrations of ≤1 μg/ml and 98.9% were inhibited at concentrations of ≤2 μg/ml. Multiple drug resistance was very common among these isolates of vancomycin-resistant
E. faecium
, while dalfopristin-quinupristin inhibited the majority at concentrations that are likely to be clinically relevant.
Bactericidal Activity of a Single-Dose Combination of Ofloxacin plus Minocycline, with or without Rifampin, against
Mycobacterium leprae
in Mice and in Lepromatous Patients
Authors: Ji Baohong , Sow Samba , Perani Evelyne , Lienhardt Christian , Diderot Vimala , Grosset Jacques
Published: October 2018

To develop a fully supervisable, monthly administered regimen for treatment of leprosy, the bactericidal effect of a single-dose combination of ofloxacin (OFLO) and minocycline (MINO), with or without rifampin (RMP), against
Mycobacterium leprae
was studied in the mouse footpad system and in previously untreated lepromatous leprosy patients. Bactericidal activity was measured by the proportional bactericidal method. In mouse experiments, the activity of a single dose of the combination OFLO-MINO was dosage related; the higher dosage of the combination displayed bactericidal activity which was significantly inferior to that of a single dose of RMP, whereas the lower dosage did not exhibit a bactericidal effect. In the clinical trial, 20 patients with previously untreated lepromatous leprosy were treated with a single dose consisting of either 600 mg of RMP plus 400 mg of OFLO and 100 mg of MINO or 400 mg of OFLO plus 100 mg of MINO. The OFLO-MINO combination exhibited definite bactericidal activity in 7 of 10 patients but was less bactericidal than the RMP-OFLO-MINO combination. Both combinations were well tolerated. Because of these promising results, a test of the efficacy of multiple doses of ROM in a larger clinical trial appears justified.
Sorivudine versus Acyclovir for Treatment of Dermatomal Herpes Zoster in Human Immunodeficiency Virus-Infected Patients: Results from a Randomized, Controlled Clinical Trial
Authors: Gnann John W. , Crumpacker Clyde S. , Lalezari Jacob P. , Smith Jean A. , Tyring Stephen K. , Baum Kenneth F. , Borucki Michael J. , Joseph W. Patrick , Mertz Gregory J. , Steigbigel Roy T. , Cloud Gretchen A. , Soong Seng-jaw , Sherrill Lanette C. , DeHertogh Deborah A. , Whitley Richard J. ,   the Collaborative Antiviral Study Group (Casg)/aids Clinical Trials Group (Actg) Herpes Zoster Study Group
Published: October 2018

The present randomized, double-blind, placebo-controlled, multicenter clinical trial was designed to compare the efficacy and tolerability of sorivudine [1-β-
d
-arabinofuranosyl-E-(2-bromovinyl)uracil] and acyclovir for the treatment of dermatomal herpes zoster in human immunodeficiency virus (HIV)-seropositive patients. A total of 170 HIV-seropositive adults presenting with herpes zoster (confirmed by direct fluorescent-antigen testing and/or viral culture) were enrolled and randomized to receive a 10-day course of orally administered sorivudine (40 mg once daily plus acyclovir placebos) or acyclovir (800 mg five times daily plus sorivudine placebo). Patients were monitored daily to document the events of cutaneous healing, pain, zoster-related complications, and drug-related adverse events. Patients were reassessed on days 21 and 28 and then once monthly for 1 year. The primary efficacy endpoint was time to the cessation of new vesicle formation. Secondary efficacy endpoints included times to other events of cutaneous healing, resolution of pain, and frequency of dissemination and zoster recurrence. In a multivariate analysis, sorivudine was superior to acyclovir for reducing the times to the cessation of new vesicle formation (relative risk [RR] = 1.54, 95% confidence interval [CI] = 1.00 to 2.36;
P
= 0.049) and total lesion crusting (RR = 1.48, 95% CI = 1.07 to 2.04;
P
= 0.017). In a univariate analysis, there was a trend favoring sorivudine for the cessation of new vesicle formation (median of 3 versus 4 days;
P
= 0.07) and a significant advantage for time to total lesion crusting (median of 7 versus 8 days;
P
= 0.02). The time to the resolution of zoster-associated pain, the frequency of dissemination, and the frequency of zoster recurrence were not different between the two treatment groups. Both drugs were well tolerated. Sorivudine is an effective drug for the treatment of herpes zoster in HIV-infected patients and results in accelerated cutaneous healing when compared with acyclovir therapy.
Photoaffinity Analog of the Semisynthetic Echinocandin LY303366: Identification of Echinocandin Targets in
Candida albicans
Authors: Radding Jeffrey A. , Heidler Steven A. , Turner William W.
Published: October 2018

The echinocandins are a family of cyclic lipopeptides with potent antifungal activity. These compounds inhibit the synthesis of β-1,3-glucan in fungi. The new semisynthetic echinocandin LY303366 was derivatized to produce a photoactivatable cross-linking echinocandin analog with antifungal activity. This analog was radioiodinated and used as a probe in microsomal membrane preparations of
Candida albicans
which contain glucan synthase activity. The photoaffinity probe identified two major proteins of 40 and 18 kDa in both membrane preparations. Labeling of these proteins was specific in that it required irradiation with UV light and was effectively competed against with unlabeled echinocandin analogs. In addition, the abilities of echinocandin analogs to compete with the photoaffinity probe correlated to their relative antifungal potencies and glucan synthase inhibition. The 40-kDa protein was isolated, and partial sequences were obtained from internal peptide fragments of the protein. Analysis of the sequences of these internal peptides of the 40-kDa protein revealed that it was a new protein not previously described as being involved in glucan synthesis or the mode of action of echinocandins.
Risk of Development of In Vitro Resistance to Amoxicillin, Clarithromycin, and Metronidazole inHelicobacter pylori
Authors: Sörberg Mikael , Hanberger Håkan , Nilsson Maud , Björkman Anders , Nilsson Lennart E.
Published: October 2018

We have studied initial killing, morphological alterations, the frequency of occurrence, and the selective growth of resistant subpopulations ofHelicobacter pyloriduring exposure to amoxicillin, clarithromycin, or metronidazole by bioluminescence assay of intracellular ATP levels, microscopy, and a viable count assay. We found an induction of spheroplasts and a decrease in intracellular ATP levels after 21 h of exposure to high concentrations of amoxicillin. During clarithromycin exposure the onset of a decrease in intracellular ATP levels started after prolonged incubation, and with the highest concentration of clarithromycin an induction of coccoid forms was seen after 68 h. Metronidazole exposure resulted in the strongest initial decrease in intracellular ATP levels, and coccoid forms were seen after 21 h of exposure to high concentrations of metronidazole. Amoxicillin caused a low-level increase in resistant subpopulations, which indicates a need for surveillance of the amoxicillin susceptibility ofH. pyloriin order to detect decreasing susceptibility. No increase in the numbers of resistant subpopulations was demonstrated during clarithromycin exposure. Metronidazole selected resistant subpopulations, which caused high-level resistance inH. pylori.
Meropenem versus Cefuroxime plus Gentamicin for Treatment of Serious Infections in Elderly Patients
Authors: Jaspers C. A. J. J. , Kieft H. , Speelberg B. , Buiting A. , van Marwijk Kooij M. , Ruys G. J. H. M. , Vincent H. H. , Vermeulen M. C. A. , Olink A. G. , Hoepelman I. M.
Published: October 2018

In this multicenter study, the efficacy of and tolerability for meropenem were compared with those for the combination of cefuroxime-gentamicin (±metronidazole) for the treatment of serious bacterial infections in patients ≥65 years of age. A total of 79 patients were randomized; thirty-nine received meropenem (1 g/8 h), and 40 received cefuroxime (1.5 g/8 h) plus gentamicin (4 mg/kg of body weight daily) for 5 to 10 days. Metronidazole (500 mg/6 h) could be added to the cefuroxime-gentamicin regimen for the treatment of intra-abdominal infections (
n
= 10). Seventy patients were evaluable for clinical efficacy; the primary diagnoses were as follows: pneumonia in 41 patients (20 treated with meropenem, 21 treated with cefuroxime-gentamicin), intra-abdominal infection in 10 patients (7 meropenem, 3 cefuroxime-gentamicin-metronidazole), urinary tract infection (UTI) in 11 patients (6 meropenem, 5 cefuroxime-gentamicin), sepsis syndrome in 7 patients (4 meropenem, 3 cefuroxime-gentamicin), and “other” in 1 patient (cefuroxime-gentamicin). The pathogens isolated from 18 patients with bacteremia were as follows:
Staphylococcus
spp. (
n
= 2),
Streptococcus
spp. (
n
= 2), members of the family
Enterobacteriaceae
(
n
= 11), and
Bacteroides
spp. (
n
= 3). A satisfactory clinical response at the end of therapy was achieved in 26 of 37 (70%) and 24 of 33 (73%) evaluable patients treated with meropenem and combination therapy, respectively. Clinical success was achieved in 23 of 31 (74%) and 21 of 28 (75%) evaluable patients with infections other than UTIs, respectively. A satisfactory microbiological response occurred in 15 of 22 (68%) patients in the meropenem group compared with 12 of 19 (63%) treated with combination therapy. Renal failure occurred during therapy in 2 of 39 (5%) meropenem recipients compared with 5 of 40 (13%) of those treated with combination therapy. The findings in this small study indicate that meropenem is as efficacious for and as well tolerated by elderly patients as the combination of cefuroxime-gentamicin (±metronidazole).
Sequence of the Gene Encoding a Plasmid-Mediated Cefotaxime-Hydrolyzing Class A β-Lactamase (CTX-M-4): Involvement of Serine 237 in Cephalosporin Hydrolysis
Authors: Gazouli Maria , Tzelepi Eva , Sidorenko Sergei V. , Tzouvelekis Leonidas S.
Published: October 2018

The sequence of the gene encoding a novel cefotaxime-hydrolyzing β-lactamase (CTX-M-4) was determined. It was located in a plasmid harbored by a
Salmonella typhimurium
strain. CTX-M-4 was similar to the plasmidic cefotaxime-hydrolyzing β-lactamases CTX-M-2 and Toho-1 and related to the chromosomal β-lactamase of
Klebsiella oxytoca
. A Ser-237→Ala substitution, introduced by site-directed mutagenesis, caused minor alterations in the interaction of CTX-M-4 with β-lactams, reducing slightly the relative hydrolytic activity against cefotaxime and the susceptibility to inhibition by clavulanate.
A Triazine Dye, Cibacron Blue F3GA, Decreases Oxacillin Resistance Levels in Methicillin-Resistant
Staphylococcus aureus
Authors: Shirai Chiharu , Sugai Motoyuki , Komatsuzawa Hitoshi , Ohta Kouji , Yamakido Michio , Suginaka Hidekazu
Published: October 2018

Cibacron blue F3GA (CB) was found to reduce the MIC of oxacillin for methicillin-resistant
Staphylococcus aureus
(MRSA). This effect was not observed with methicillin-susceptible
S. aureus
. CB alters the resistance level of MRSA through a factor(s) other than
mecA
-related products, major autolysins, or
femAB
products. The exact target(s) of CB in causing the effect is unknown.
Properties of Mutant SHV-5 β-Lactamases Constructed by Substitution of Isoleucine or Valine for Methionine at Position 69
Authors: Giakkoupi Panagiota , Miriagou Vivi , Gazouli Maria , Tzelepi Eva , Legakis Nicholas J. , Tzouvelekis Leonidas S.
Published: October 2018

The effect of replacement of Met-69 by Ile or Val on the properties of the extended-spectrum β-lactamase SHV-5 was studied. Mutant enzymes were constructed by site-specific mutagenesis and expressed under isogenic conditions in
Escherichia coli
DH5α cells. Compared with SHV-5, the mutant β-lactamases conferred lower levels of β-lactam resistance and were less efficient in hydrolyzing ampicillin, cephalothin, and cefotaxime. The substitutions rendered SHV-5 less susceptible to inhibition by clavulanate, sulbactam, and tazobactam; however, the MICs of penicillin-inhibitor combinations remained similar, suggesting an attenuation of penicillinase activity.