bluuebunny/crossref_metadata_2025 huggingface dataset:
Mechanisms of Fluoroquinolone Transport by Human Neutrophils
Authors: Walters John D. , Zhang Fanjie , Nakkula Robin J.
Published: October 2018

Neutrophils accumulate ciprofloxacin and other fluoroquinolones, a process that enhances the killing of intracellular pathogens and could facilitate the delivery of these agents to infection sites by migrating neutrophils. The mechanisms by which transport occurs have not been characterized. In the present study, quiescent neutrophils transported ciprofloxacin with an observed

K
m

of 167 μg/ml (501 μM) and a maximum velocity of 25.2 ng/min/10
6
cells. When neutrophils were stimulated with phorbol myristate acetate (PMA), a second component of ciprofloxacin transport was induced. This pathway had an apparent

K
m

of 9.76 μg/ml (29.3 μM) and a maximum velocity of 59.3 ng/min/10
6
cells. Transport by both pathways was Na
+
independent. Ciprofloxacin transport by quiescent cells was relatively insensitive to pH and
N
-ethylmaleimide but was competitively inhibited by adenine (

K
i

= 1.55 mM). Papaverine, a benzylisoquinoline known to inhibit nucleobase transport, also inhibited ciprofloxacin transport by quiescent cells. In contrast, transport by PMA-stimulated cells was enhanced at pH 8.2, inhibited at pH 6.2, and blocked by
N
-ethylmaleimide. Cationic and neutral amino acids and cystine competitively inhibited ciprofloxacin transport by PMA-stimulated neutrophils (

K
i

= 158 μM for ornithine) but had little effect on quiescent cells. PMA-activated transport was not inhibited when the Na
+
in the medium was replaced with K
+
or Li
+
, and the pattern of inhibition by cationic and neutral amino acids was similar. In summary, neutrophils continuously transport ciprofloxacin via a transport pathway shared by adenine. Activation by PMA induces a separate, higher-affinity transport pathway shared by a broad scope of amino acids. Neutrophils utilize one or both of these mechanisms to transport other fluoroquinolones.
Antimicrobial Susceptibility Testing of
Helicobacter pylori
in a Large Multicenter Trial: the MACH 2 Study
Authors: Mégraud Francis , Lehn Norbert , Lind Tore , Bayerdörffer Ekkehard , O’Morain Colm , Spiller Robin , Unge Peter , van Zanten Sander Veldhuyzen , Wrangstadh Michael , Burman Carl Fredrik
Published: October 2018

Culture and susceptibility testing of
Helicobacter pylori
strains was performed in a large multinational, multicenter randomized clinical trial. Culture was carried out on gastric biopsy samples obtained from 516 patients at entry and had a sensitivity of 99% when the [
13
C]urea breath test was used as a reference. Susceptibility testing was performed for clarithromycin and metronidazole on 485 strains by an agar dilution method and the epsilometer test (Etest) and for amoxicillin by an agar dilution method only. Resistance to clarithromycin (>1 μg/ml) was found in 3% of the
H. pylori
strains, with a perfect correlation between Etest and agar dilution methods. Resistance to metronidazole (>8 μl/ml) was found in 27% of the strains by agar dilution, but there were important discrepancies between it and the Etest method. No resistance to amoxicillin was found. The logarithms of the MICs of the three antibiotics against susceptible strains had a distribution close to normal. The impact of resistance was tested in the four arms of the trial. There were not enough clarithromycin-resistant strains to evaluate the impact of resistance on the cure rate of clarithromycin-based regimens. For metronidazole-resistant strains, the impact noted in the clarithromycin-metronidazole arm was partially overcome when omeprazole was added (76% eradication for resistant strains versus 95% for susceptible strains). Secondary resistance to clarithromycin occurred in strains from 12 of 105 patients (11.4%) after the failure of a clarithromycin-based regimen to effect eradication. The detection of point mutations in clarithromycin-resistant strains was performed by a combination of PCR and restriction fragment length polymorphism. Mutations (A2142G and 2143G) were found in all strains tested except one. This study stresses the importance of performing susceptibility tests in clinical trials in order to explain the results of different treatments.
Differential Modulatory Effects of Clarithromycin on the Production of Cytokines by a Tumor
Authors: Sassa Kazuhiko , Mizushima Yutaka , Kobayashi Masashi
Published: October 2018

In vitro treatment with clarithromycin inhibited the expression of the matrix metalloproteinase-9, transforming growth factor β, and tumor necrosis factor alpha genes in 13762NF rat mammary adenocarcinoma cells. Transient enhancement, rather than inhibition, was observed for the interleukin-6 gene, and no significant change was observed for the tissue inhibitor of metalloproteinase-2 gene. Such an effect was not observed for cefotiam or gentamicin.
Efficacies of High-Dose Fluconazole plus Amphotericin B and High-Dose Fluconazole plus 5-Fluorocytosine versus Amphotericin B, Fluconazole, and 5-Fluorocytosine Monotherapies in Treatment of Experimental Endocarditis, Endophthalmitis, and Pyelonephritis Due to
Candida albicans
Authors: Louie Arnold , Liu Weiguo , Miller Dorothy A. , Sucke Alana C. , Liu Qing-Feng , Drusano George L. , Mayers Martin , Miller Michael H.
Published: October 2018

We compared the efficacies of fluconazole (Flu), amphotericin B (AmB), and 5-fluorocytosine (5FC) monotherapies with the combination of Flu plus 5FC and Flu plus AmB in a rabbit model of
Candida albicans
endocarditis, endophthalmitis, and pyelonephritis. The dose of Flu used was that which resulted in an area under the concentration-time curve in rabbits equivalent to that seen in humans who receive Flu at 1,600 mg/day, the highest dose not associated with central nervous system toxicity in humans. Quantitative cultures of heart valve vegetations, the choroid-retina, vitreous humor, and kidney were conducted after 1, 5, 14, and 21 days of therapy. All untreated controls died within 6 days of infection; animals treated with 5FC monotherapy all died within 18 days. In contrast, 93% of animals in the other treatment groups appeared well and survived until they were sacrificed. At day 5, the relative decreases in CFU per gram in the vitreous humor were greater in groups that received Flu alone and in combination with 5FC or AmB than in groups receiving AmB or 5FC monotherapies (
P
< 0.005) but were similar thereafter. In the choroid-retina, 5FC was the least-active drug. However, there were no differences in choroidal fungal densities between the other treatment groups. On days 5 and 14 of therapy, fungal densities in kidneys of AmB recipients were lower than those resulting from the other therapies (
P
< 0.001 and
P
≤ 0.038, respectively) and AmB-plus-Flu therapy was antagonistic; however, all therapies for fungal pyelonephritis were similar by treatment day 21. While fungal counts in cardiac valves of Flu recipients were similar to those of controls on day 5 of therapy and did not change from days 1 to 21, AmB therapy significantly decreased valvular CFUs versus Flu at days 5, 14, and 21 (
P
< 0.005 at each time point). 5FC plus Flu demonstrated enhanced killing in cardiac vegetations compared with Flu or 5FC as monotherapies (
P
< 0.03). Similarly, the combination of AmB and Flu was more active than Flu in reducing the fungal density in cardiac vegetations (
P
< 0.03). However, as in the kidney, AmB plus Flu demonstrated antagonism versus AmB monotherapy in the treatment of
C. albicans
endocarditis (
P
< 0.05,
P
= 0.036, and
P
< 0.008 on days 5, 14, and 21, respectively).
Pharmacokinetics of Cefepime in Patients with Thermal Burn Injury
Authors: Bonapace Charles R. , White Roger L. , Friedrich Lawrence V. , Norcross E. Douglas , Bosso John A.
Published: October 2018

The pharmacokinetics of cefepime following administration of a single 2-g dose were evaluated for 12 adult patients with thermal burn injury and suspected or documented infection. Serial blood and urine samples for cefepime concentration determination were obtained for 24 h following drug administration. Serum and urine cefepime concentrations were determined by high-performance liquid chromatography and serum concentrations were fit to a two-compartment pharmacokinetic model. Mean (standard deviation [SD]) age, actual body weight (ABW), percent total body surface area burned, and days postburn at the time of study were 41 (13) years, 84 (22) kg, 36 (17)%, and 9 (3) days, respectively. Mean (SD) measured creatinine clearance (CL
CR
), total clearance (CL
T
), renal clearance (CL
R
), alpha phase half-life, beta phase half-life, and volume of distribution at steady state (
V
SS
) were 135 (31) ml/min, 8.8 (2.4) liters/h, 8.1 (2.0) liters/h, 0.33 (0.14) h, 2.8 (0.6) h, and 0.43 (0.10) liters/kg ABW, respectively. Cefepime CL
T
and CL
R
in burn patients were similar to previously reported values for healthy volunteers when normalized by CL
CR
. Stepwise multiple regression was used to associate CL
T
with CL
CR
and days postburn (
r
2
= 0.861), CL
R
with CL
CR
and days postburn (
r
2
= 0.773), nonrenal clearance with percent third-degree (% 3°) burn and albumin concentration (
r
2
= 0.550), and
V
SS
only with % 3° burn (
r
2
= 0.624). Simulated steady-state serum concentrations obtained by using the patients’ pharmacokinetic parameters exceeded the susceptibility interpretive standard (breakpoint) of cefepime for at least 60% of the dosing interval with dosing regimens of 1 g every 8 h (q8h), 2 g q8h, and 2 g q12h. Despite differences in pharmacokinetic parameters between our patients and healthy volunteers, it appears that these dosing regimens may be adequate in similar burn patients.
Comparison of In Vitro Activities of Camptothecin and Nitidine Derivatives against Fungal and Cancer Cells
Authors: Del Poeta Maurizio , Chen Shih-Fong , Von Hoff Daniel , Dykstra Christine C. , Wani Mansukh C. , Manikumar Govindarajan , Heitman Joseph , Wall Monroe E. , Perfect John R.
Published: October 2018

The activities of a series of camptothecin and nitidine derivatives that might interact with topoisomerase I were compared against yeast and cancer cell lines. Our findings reveal that structural modifications to camptothecin derivatives have profound effects on the topoisomerase I-drug poison complex in cells. Although the water-soluble anticancer agents topotecan and irinotecan are less active than the original structure, camptothecin, other derivatives or analogs with substitutions that increase compound solubility have also increased antifungal activities. In fact, a water-soluble prodrug appears to penetrate into the cell and release its active form; the resulting effect in complex with
Cryptococcus neoformans
topoisomerase I is a fungicidal response and also potent antitumor activity. Some of the compounds that are not toxic to wild-type yeast cells are extremely toxic to the yeast cells when the
C. neoformans
topoisomerase I target is overexpressed. With the known antifungal mechanism of a camptothecin-topoisomerase I complex as a cellular poison, these findings indicate that drug entry may be extremely important for antifungal activity. Nitidine chloride exhibits antifungal activity against yeast cells through a mechanism(s) other than topoisomerase I and appears to be less active than camptothecin analogs against tumor cells. Finally, some camptothecin analogs exhibit synergistic antifungal activity against yeast cells in combination with amphotericin B in vitro. Our results suggest that camptothecin and/or nitidine derivatives can exhibit potent antifungal activity and that the activities of camptothecin derivatives with existing antifungal drugs may be synergistic against pathogenic fungi. These new compounds, which exhibit potent antitumor activities, will likely require further structural changes to find more selective activity against fungal versus mammalian cells to hold promise as a new class of antifungal agents.
Hydroxyurea Potentiates the Antiherpesvirus Activities of Purine and Pyrimidine Nucleoside and Nucleoside Phosphonate Analogs
Authors: Neyts J. , De Clercq E.
Published: October 2018

Hydroxyurea has been shown to potentiate the anti-human immunodeficiency virus activities of 2′,3′-dideoxynucleoside analogs such as didanosine. We have now evaluated in vitro the effect of hydroxyurea on the antiherpesvirus activities of several nucleoside analogs (acyclovir [ACV], ganciclovir [GCV], penciclovir [PCV], lobucavir [LBV], (
R
)-9-[4-hydroxy-2-(hydroxymethyl)butyl]guanine [H2G], and brivudin and nucleoside phosphonate analogs (cidofovir [CDV] and adefovir [ADV]). When evaluated in cytopathic effect (CPE) reduction assays, hydroxyurea by itself had little effect on CPE progression and potentiated in a subsynergistic (herpes simplex virus type 1 [HSV-1]) to synergistic (HSV-2) fashion the antiviral activities of ACV, GCV, PCV, LBV, H2G, ADV, and CDV. Hydroxyurea also caused marked increases in the activities of ACV, GCV, PCV, LBV, and H2G (compounds that depend for their activation on a virus-encoded thymidine kinase [TK]) against TK-deficient (TK
−
) HSV-1. In fact, in combination with hydroxyurea the 50% effective concentrations of these compounds for inhibition of TK
−
HSV-1-induced CPE decreased from values of 20 to ≥100 μg/ml (in the absence of hydroxyurea) to values of 1 to 5 μg/ml (in the presence of hydroxyurea at 25 to 100 μg/ml). When evaluated in a single-cycle virus yield reduction assay, hydroxyurea at a concentration of 100 μg/ml inhibited progeny virus production by 60 to 90% but had little effect on virus yield at a concentration of 25 μg/ml. Under these assay conditions hydroxyurea still elicited a marked potentiating effect on the antiherpesvirus activities of GCV and CDV, but this effect was less pronounced than that in the CPE reduction assay. It is conceivable that the potentiating effect of hydroxyurea stems from a depletion of the intracellular deoxynucleoside triphosphate pools, thus favoring the triphosphates of the nucleoside analogues (or the diphosphates of the nucleoside phosphonate analogues) in their competition with the natural nucleotides at the viral DNA polymerase level. The possible clinical implications of these findings are discussed.
Expanded-Spectrum Nonnucleoside Reverse Transcriptase Inhibitors Inhibit Clinically Relevant Mutant Variants of Human Immunodeficiency Virus Type 1
Authors: Corbett Jeffrey W. , Ko Soo S. , Rodgers James D. , Jeffrey Susan , Bacheler Lee T. , Klabe Ronald M. , Diamond Sharon , Lai Chii-Ming , Rabel Shelley R. , Saye Jo Anne , Adams Stephen P. , Trainor George L. , Anderson Paul S. , Erickson-Viitanen Susan K.
Published: October 2018

A research program targeted toward the identification of expanded-spectrum nonnucleoside reverse transcriptase inhibitors which possess increased potency toward K103N-containing mutant human immunodeficiency virus (HIV) and which maintain pharmacokinetics consistent with once-a-day dosing has resulted in the identification of the 4-cyclopropylalkynyl-4-trifluoromethyl-3,4-dihydro-2(1
H
)quinazolinones DPC 961 and DPC 963 and the 4-cyclopropylalkenyl-4-trifluoromethyl-3,4-dihydro-2(1
H
)quinazolinones DPC 082 and DPC 083 for clinical development. DPC 961, DPC 963, DPC 082, and DPC 083 all exhibit low-nanomolar potency toward wild-type virus, K103N and L100I single-mutation variants, and many multiply amino acid-substituted HIV type 1 mutants. This high degree of potency is combined with a high degree of oral bioavailability, as demonstrated in rhesus monkeys and chimpanzees, and with plasma serum protein binding that can result in significant free levels of drug.
High-Dose Isoniazid Therapy for Isoniazid-Resistant Murine
Mycobacterium tuberculosis
Infection
Authors: Cynamon M. H. , Zhang Y. , Harpster T. , Cheng S. , DeStefano M. S.
Published: October 2018

The use of isoniazid (INH) for the treatment of INH-resistant
Mycobacterium tuberculosis
infection has been controversial. The purpose of the present studies was to determine if there is a dose response with INH for INH-susceptible
M. tuberculosis
Erdman (ATCC 35801), and whether high-dose INH (100 mg/kg of body weight) was more effective than standard-dose INH (25 mg/kg) for therapy of tuberculosis infections caused by INH-resistant mutants of
M. tuberculosis
Erdman. Six-week-old CD-1 mice were infected with approximately 10
7
viable mycobacteria. Early control groups of infected but untreated mice were euthanized by CO
2
inhalation 1 week later when treatment was initiated. INH (25, 50, 75, and 100 mg/kg) was given by gavage 5 days/week for 4 weeks. Late control groups of untreated mice and treated mice were sacrificed 2 days after the last dose of drug. Spleens and right lungs were removed aseptically and homogenized, and viable cell counts were determined by titration on 7H10 agar plates. In the next study, INH at 100 mg/kg was compared to INH at 25 mg/kg against an isogenic mutant of
M. tuberculosis
Erdman (INH MIC, 2 μg/ml) and the parent strain. This mutant was found to have a mutation in the KatG protein (Phe to Leu at position 183). In the first study, there was no dose response with increasing doses of INH. In the second study, there was no significant difference between the reduction of viable cell counts for mice treated with INH at 100 mg/kg and that for mice treated with INH at 25 mg/kg (parent or INH-resistant mutant). These preliminary results suggest that INH may be useful in combination therapy of
M. tuberculosis
infections caused by low-level INH-resistant organisms (INH MICs, 0.2 to 5 μg/ml) and that higher doses of INH are unlikely to be more efficacious than the standard 300-mg/day dose.
Multilocus Genotypes and DNA Fingerprints Do Not Predict Variation in Azole Resistance among Clinical Isolates of
Candida albicans
Authors: Cowen Leah E. , Sirjusingh Caroline , Summerbell Richard C. , Walmsley Sharon , Richardson Susan , Kohn Linda M. , Anderson James B.
Published: October 2018

If variation in azole resistance is due to inherent differences in strains of
Candida albicans
, as a predominantly clonal organism, then correlation between multilocus genotypes and drug resistance would be expected. A sample of 81 clinical isolates from patients infected with human immunodeficiency virus in Toronto, Canada, plus 3 reference isolates were genotyped at 16 loci, distributed on all linkage groups, by means of oligonucleotide hybridizations specific for each of the alleles at each locus. These multilocus genotypes were significantly correlated with DNA fingerprints obtained with the species-specific probe 27A, indicating widespread linkage disequilibrium in the genome. There were 64 multilocus diploid genotypes and 77 DNA fingerprint types delineated in this sample. Neither the multilocus genotyping nor DNA fingerprinting alone identified all of the 81 types identified by the combination of these two methods. Multilocus genotypes were not predictive of fluconazole resistance, suggesting that resistance is gained or lost too quickly to be predicted by linkage with neutral markers.
The
pfmdr1
Gene Is Associated with a Multidrug-Resistant Phenotype in
Plasmodium falciparum
from the Western Border of Thailand
Authors: Price R. N. , Cassar C. , Brockman A. , Duraisingh M. , van Vugt M. , White N. J. , Nosten F. , Krishna S.
Published: October 2018

On the western border of Thailand,
Plasmodium falciparum
has become resistant to almost all antimalarial agents. The molecular basis of resistance in these parasite populations has not been well characterized. This study assessed genetic polymorphisms in the
pfmdr1
gene in 54 parasites collected from the western border of Thailand to determine the relationship of
pfmdr1
copy number and codon mutations with parasite sensitivities to mefloquine, chloroquine, halofantrine, quinine, and artesunate assessed in vitro. A point mutation at codon 86 (resulting in a change of Asn to Tyr) was associated with a significantly lower 50% inhibitory concentration (IC
50
) of mefloquine (median, 9 ng/ml versus 52.4 ng/ml;
P
= 0.003). Overall 35% of the isolates (19 of 54) had an increase in
pfmdr1
copy number, and all 19 carried the wild-type allele at codon 86. Increased
pfmdr1
copy number was associated with higher IC
50
s of mefloquine (
P
= 0.04) and artesunate (
P
= 0.005), independent of polymorphism at codon 86. The relationship between
pfmdr1
and resistance to structurally distinct antimalarial agents confirms the presence of a true multidrug-resistant phenotype.
Determination of Zidovudine Triphosphate Intracellular Concentrations in Peripheral Blood Mononuclear Cells from Human Immunodeficiency Virus-Infected Individuals by Tandem Mass Spectrometry
Authors: Font Eva , Rosario Osvaldo , Santana Jorge , García Hermes , Sommadossi Jean-Pierre , Rodriguez Jose F.
Published: October 2018

Nucleoside reverse transcriptase inhibitors (NRTIs) used against the human immunodeficiency virus (HIV) need to be activated intracellularly to their triphosphate moiety to inhibit HIV replication. Intracellular concentrations of these NRTI triphosphates, especially zidovudine triphosphate (ZDV-TP), are relatively low (low numbers of femtomoles per 10
6
cells) in HIV-infected patient peripheral blood mononuclear cells. Recently, several methods have used either high-performance liquid chromatography (HPLC) or solid-phase extraction (SPE) coupled with radioimmunoassay to obtain in vivo measurements of ZDV-TP. The limit of detection (LOD) by these methods ranged from 20 to 200 fmol/10
6
cells. In this report, we describe the development of a method to determine intracellular ZDV-TP concentrations in HIV-infected patients using SPE and HPLC with tandem mass spectrometry for analysis. The LOD by this method is 4.0 fmol/10
6
cells with a linear concentration range of at least 4 orders of magnitude from 4.0 to 10,000 fmol/10
6
cells. In hispanic HIV-infected patients, ZDV-TP was detectable even when the sampling time after drug administration was 15 h. Intracellular ZDV-TP concentrations in these patients ranged from 41 to 193 fmol/10
6
cells. The low LOD obtained with this method will provide the opportunity for further in vivo pharmacokinetic studies of intracellular ZDV-TP in different HIV-infected populations. Furthermore, this methodology could be used to perform simultaneous detection of two or more NRTIs, such as ZDV-TP and lamivudine triphosphate.
Activity of Gemifloxacin against Penicillin- and Ciprofloxacin-Resistant
Streptococcus pneumoniae
Displaying Topoisomerase- and Efflux-Mediated Resistance Mechanisms
Authors: Heaton Victoria J. , Goldsmith Colin E. , Ambler Jane E. , Fisher L. Mark
Published: October 2018

Nine penicillin-resistant
Streptococcus pneumoniae
clinical isolates from Northern Ireland, resistant to ciprofloxacin (MICs, 2 to 64 μg/ml) through topoisomerase- and/or reserpine-sensitive efflux mechanisms, were highly susceptible to gemifloxacin (MICs, 0.03 to 0.12 μg/ml). Two strains (requiring a ciprofloxacin MIC of 64 μg/ml) carried known quinolone resistance mutations in
parC
,
parE
, and
gyrB
, resulting in S79F, D435V, and E474K changes, respectively. Thus, gemifloxacin is active against clinical strains exhibiting altered topoisomerase and efflux phenotypes.
Comparative In Vitro Antimicrobial Activities of the Newly Synthesized Quinolone HSR-903, Sitafloxacin (DU-6859a), Gatifloxacin (AM-1155), and Levofloxacin against
Mycobacterium tuberculosis
and
Mycobacterium avium
Complex
Authors: Tomioka Haruaki , Sato Katsumasa , Akaki Tatsuya , Kajitani Hiroko , Kawahara Shin , Sakatani Mitsunori
Published: October 2018

We compared the in vitro antimycobacterial activity of a new fluoroquinolone, HSR-903, with strong activity against gram-positive cocci with those of levofloxacin (LVFX), sitafloxacin (STFX), and gatifloxacin (GFLX). The MICs of the quinolones for
Mycobacterium tuberculosis
and
Mycobacterium avium
complex were in the order STFX ≈ GFLX < LVFX ≦ HSR-903 and STFX ≦ GFLX ≦ HSR-903 ≦ LVFX, respectively. HSR-903 effectively eliminated intramacrophagial
M. tuberculosis
, as did LVFX, and exhibited bacteriostatic effects against
M. tuberculosis
replicating in type II alveolar cells.
High-Level Aminoglycoside Resistance in the Beta-Hemolytic Group G
Streptococcus
Isolate BM2721
Authors: Galimand Marc , Lambert Thierry , Gerbaud Guy , Courvalin Patrice
Published: October 2018

The beta-hemolytic group G
Streptococcus
clinical isolate BM2721 was resistant to high levels of aminoglycosides by synthesis of AAC(6′)-APH(2"), APH(3′)-III, and ANT(6) modifying enzymes. The corresponding genes were found to be adjacent as the result of a recombination event between Tn
4001
and Tn
5405
, two transposons originating in staphylococci.
Distribution of β-Lactamases in Actinomycetes
Authors: Ogawara Hiroshi , Kawamura Nao , Kudo Takuji , Suzuki Ken-Ichiro , Nakase Takashi
Published: October 2018

The distribution of β-lactamase activities in a collection of actinomycete strains was surveyed. Six of 127 strains were found to produce β-lactamase. This low frequency was in contrast to the case with
Streptomyces
species. The producing strains were not related phylogenetically. MICs of benzylpenicillin did not correlate with β-lactamase production.
Genetic Characterization of Antimicrobial Resistance in Canadian Isolates of
Salmonella
Serovar Typhimurium DT104
Authors: Ng Lai-King , Mulvey Michael R. , Martin Irene , Peters Geoffrey A. , Johnson Wendy
Published: October 2018

PCR was used to identify antibiotic resistance determinants in 31 Canadian
Salmonella
serovar Typhimurium DT104 isolates. Genes encoding resistance to ampicillin (
pse1
or
blaP1
), chloramphenicol (
pasppflo
-like), streptomycin-spectinomycin (
aadA2
), sulfonamide (
sulI
), and tetracycline [
tet
(G)] were mapped to a 13-kb region of DNA of one isolate. Two copies of
sulI
were identified and mapped to the 3′ end of either
pse1
or
aadA2
integrons. The two integrons were separated by the
pasppflo
-like gene and the
tet
(G) gene. The kanamycin resistance determinant (
aphA-1
) was present on a 2.0-MDa plasmid (five isolates) or on the chromosome (three isolates).
Comparative In Vitro Activities of Meropenem, Imipenem, Temocillin, Piperacillin, and Ceftazidime in Combination with Tobramycin, Rifampin, or Ciprofloxacin against
Burkholderia cepacia
Isolates from Patients with Cystic Fibrosis
Authors: Bonacorsi Stephane , Fitoussi Frederic , Lhopital Sylvie , Bingen Edouard
Published: October 2018

We evaluated the activities of meropenem, imipenem, temocillin, piperacillin, and ceftazidime by determination of the MICs for 66 genotypically characterized
Burkholderia cepacia
isolates obtained from the sputum of cystic fibrosis patients. In vitro synergy assays, as performed by the time-kill methodology, of two- and three-drug combinations of the β-lactams with tobramycin, rifampin, and/or ciprofloxacin were also performed with 10 strains susceptible, intermediate, or resistant to fluoroquinolones. On the basis of the MICs, meropenem and temocillin were the most active β-lactam agents, with MICs at which 90% of isolates are inhibited of 8 and 32 μg/ml, respectively. The addition of ciprofloxacin significantly enhanced the killing activities of piperacillin, imipenem, and meropenem against the 10 strains tested (
P
< 0.05). The best killing activity was obtained with the combination of meropenem and ciprofloxacin, with bactericidal activity of 3.31 ± 0.36 log
10
CFU/ml (
P
< 0.05). Compared to the activity of the two-drug β-lactam–ciprofloxacin combination, the addition of rifampin or tobramycin did not significantly increase the killing activity (
P
> 0.05). The three-drug combinations (with or without ciprofloxacin) significantly enhanced the killing activities of piperacillin, imipenem, and meropenem relative to the activities of the β-lactams used alone (
P
< 0.05). The combination β-lactam–ciprofloxacin–tobramycin was the combination with the most consistently synergistic effect.
Amphotericin B and Fluconazole Affect Cellular Charge, Macrophage Phagocytosis, and Cellular Morphology ofCryptococcus neoformansat Subinhibitory Concentrations
Authors: Nosanchuk Joshua D. , Cleare Wendy , Franzot Sarah P. , Casadevall Arturo
Published: October 2018

Amphotericin B (AmB) and fluconazole (FLU) are the major antifungal drugs used in the treatment of cryptococcosis. Both drugs are believed to exert their antifungal effects through actions on cell membrane sterols. In this study we investigated whether AmB and FLU had other, more subtle effects onC. neoformansthat could contribute to their therapeutic efficacy.C. neoformanscells were grown in media with subinhibitory concentrations of either AmB or FLU and analyzed for cellular charge, phagocytosis by macrophages with antibody and complement opsonins, appearance by scanning electron and light microscopies, and release of the capsular polysaccharide glucuronoxylomannan into the culture medium. Growth in the presence of either AmB or FLU resulted in major reductions in cellular charge, as measured by determination of the zeta potential. Phagocytosis studies demonstrated that exposure ofC. neoformansto subinhibitory concentrations of AmB or FLU enhanced phagocytosis by macrophages. Scanning electron microscopy revealed that a large proportion of cells had an altered capsular appearance. Cells grown in medium with either AmB or FLU were smaller and released more glucuronoxylomannan into the culture medium than cells grown without antibiotics. The results suggest additional mechanisms of action for AmB and FLU that may be operative in body compartments where drug levels do not achieve the MICs. Furthermore, the results suggest mechanisms by which AmB and FLU can cooperate with humoral and cellular immune defense systems in controllingC. neoformansinfections.
glmM
Operon and Methicillin-Resistant
glmM
Suppressor Mutants in
Staphylococcus aureus
Authors: Glanzmann Philipp , Gustafson John , Komatsuzawa Hithoshi , Ohta Kouji , Berger-Bächi Brigitte
Published: October 2018

The
Staphylococcus aureus
phosphoglucosamine mutase gene
glmM
was shown to be the last gene of a three-cistron operon,
orf1-orf2-glmM
. One transcriptional start was identified upstream of
orf1
, and a second start producing a monocistronic transcript was identified upstream of
glmM
. Disruption of
glmM
abolished GlmM production, decreased methicillin resistance, and resulted in teicoplanin hypersusceptibility without affecting the production of the endogenous penicillin-binding proteins and PBP 2′. Complementation of the
glmM
mutation by the complete
glmM
operon restored both methicillin resistance and normal teicoplanin susceptibility. In contrast, a highly methicillin-resistant suppressor mutant obtained by selection for growth in the presence of methicillin remained GlmM deficient and teicoplanin hypersusceptible. The suppressor mutation was not linked to the
glmM
operon but was correlated with decreased autolysis and increased production of a 49-kDa protein, suggesting that there is an alternative pathway for glucosamine-1-phosphate synthesis in
S. aureus.
Comparison of a New Triazole Antifungal Agent, Schering 56592, with Itraconazole and Amphotericin B for Treatment of Histoplasmosis in Immunocompetent Mice
Authors: Connolly Patricia , Wheat Joe , Schnizlein-Bick Carol , Durkin Michelle , Kohler Steve , Smedema Melinda , Goldberg Janet , Brizendine Edward , Loebenberg David
Published: October 2018

A murine model of intratracheally induced histoplasmosis was used to evaluate a new triazole antifungal agent, Schering (SCH) 56592, for treatment of histoplasmosis. MICs were determined for SCH 56592, amphotericin B, and itraconazole by testing yeast-phase isolates from 20 patients by a macrobroth dilution method. The MICs at which 90% of the isolates are inhibited were for 0.019 μg/ml for SCH 56592, 0.5 μg/ml for amphotericin B, and ≤0.019 μg/ml for itraconazole. Survival studies were done on groups of 10 B6C3F1mice with a lethal inoculum of 105. All mice receiving 5, 1, or 0.25 mg of SCH 56592 per kg of body weight per day, 2.5 mg of amphotericin B per kg every other day (qod), or 75 mg of itraconazole per kg per day survived to day 29. Only 44% of mice receiving 5 mg of itraconazole/kg/day survived to day 29. Fungal burden studies done in similar groups of mice with a sublethal inoculum of 104showed a reduction in CFUs andHistoplasmaantigen levels in lung and spleen tissue in animals treated with 2 mg of amphotericin B/kg qod, 1 mg of SCH 56592/kg/day, and 75 mg of itraconazole/kg/day, but not in those treated with lower doses of the study drugs (0.2 mg of amphotericin B/kg qod, 0.1 mg of SCH 56592/kg/day, or 10 mg of itraconazole/kg/day). Serum drug concentrations were measured 3 and 24 h after the last dose in mice (groups of five to seven mice), each treated for 7 days with SCH 56592 (10 and 1 mg/kg/day) and itraconazole (75 and 10 mg/kg/day). Mean levels measured by bioassay were as follows: SCH 56592, 10 mg/kg/day (2.15 μg/ml at 3 h and 0.35 μg/ml at 24 h); SCH 56592, 1 mg/kg/day (0.54 μg/ml at 3 h and none detected at 24 h); itraconazole, 75 mg/kg/day (22.53 μg/ml at 3 h and none detected at 24 h); itraconazole, 10 mg/kg/day (1.33 μg/ml at 3 h and none detected at 24 h). Confirmatory results were obtained by high-pressure liquid chromatography assay. These studies show SCH 56592 to be a promising candidate for studies of treatment of histoplasmosis in humans.
Enterococci with Acquired Vancomycin Resistance in Pigs and Chickens of Different Age Groups
Authors: Butaye Patrick , Devriese Luc A. , Goossens Herman , Ieven Margaretha , Haesebrouck Freddy
Published: October 2018

Results for isolation of glycopeptide-resistant enterococci from fecal samples of pigs and chickens were found to differ strongly depending upon the type and age of animals and isolation technique (direct selective plate or broth enrichment). Isolations were frequent in broiler chickens and in sows but rare in layer chickens.
Fungal Lethality, Binding, and Cytotoxicity of Syringomycin-E
Authors: De Lucca A. J. , Jacks T. J. , Takemoto J. , Vinyard B. , Peter J. , Navarro E. , Walsh T. J.
Published: October 2018

Syringomycin-E (SE) was significantly lethal to
Aspergillus
and
Fusarium
species at between 1.9 and 7.8 μg/ml. SE complexed with the following fungal wall constituents (in order of binding): β-1,3-glucan > chitin > mannan > ergosterol = cholesterol. Cytotoxicity in HeLa cells was proportional to the SE concentration, while the amount required for cytotoxicity was 3 to 20 times that needed to kill 95% of the fungi tested.
Resistance of
Pseudomonas aeruginosa
to Imipenem Induced by Eluates from Siliconized Latex Urinary Catheters Is Related to Outer Membrane Protein Alterations
Authors: Martínez-Martínez Luis , Pascual Alvaro , del Carmen Conejo María , Picabea Leandro , Perea Evelio J.
Published: October 2018

The activity of imipenem against
Pseudomonas aeruginosa
HUS-3 decreased by 16 times in the presence of substances eluted from siliconized latex urinary catheters (SLUCs). SLUCs did not inactivate imipenem or increase β-lactamase activity. The outer membrane of
P. aeruginosa
HUS-3 grown in the presence of eluate lacked an OprD-like protein and expressed a new 50-kDa protein. The decreased activity of imipenem against
P. aeruginosa
in the presence of SLUCs is related to the loss of an OprD-like protein and the expression of a new outer membrane protein.
Mycobacteriocidal Action of Exogenous Nitric Oxide
Authors: Long Richard , Light Bruce , Talbot James A.
Published: October 2018

We tested the hypothesis that exposure of extracellularMycobacterium tuberculosisto low concentrations (<100 ppm) of nitric oxide (NO) for short periods (24 h or less) will result in microbial killing. We observed that NO had both dose- and time-dependent cidal effects that were very significant by two-way analysis of variance (Fratios of 13.4 [P< 0.001] and 98.1 [P< 0.0001], respectively). Conceivably, extracellular bacilli in patients with pulmonary tuberculosis might be vulnerable to exogenous NO.
Detection of
gyrA
Mutations among 335
Pseudomonas aeruginosa
Strains Isolated in Japan and Their Susceptibilities to Fluoroquinolones
Authors: Takenouchi Takashi , Sakagawa Eiko , Sugawara Mie
Published: October 2018

gyrA
point mutations in 335 clinical
Pseudomonas aeruginosa
isolates were examined mainly by nonisotopic single-strand conformation polymorphism analysis and direct sequencing. Seven types of missense
gyrA
mutations were observed in 70 of 335 strains (20.9%), and ciprofloxacin MICs were ≥3.13 μg/ml for 63 of 70 strains (90.0%). These included two double point mutations and three novel mutations (Ala-67→Ser plus Asp-87→Gly, Ala-84→Pro, and Gln-106→Leu). Thr-83→Ile mutants were predominantly observed (63 of 70 mutants) and showed high-level fluoroquinolone resistance (ciprofloxacin MIC at which 50% of isolates are inhibited, 25 μg/ml).
An Alternative Animal Model for Comparison of Treatments for Cryptococcal Meningitis
Authors: Najvar Laura K. , Bocanegra Rosie , Graybill John R.
Published: October 2018

Weanling outbred rats were infected with
Cryptococcus neoformans
by direct percranial puncture and inoculation into the cranium. A lethal infection ensued. Treatment with LY295337, a depsipeptide with antifungal activity, was effective in prolonging survival and reducing fungal counts in brain tissue. Weanling rats are an acceptable model for the study of central nervous system infection with
C. neoformans
.
In Vitro Activities of Aminomethyl-Substituted Analogs of Novel Tetrahydrofuranyl Carbapenems
Authors: Weiss William J. , Petersen Peter J. , Jacobus Nilda V. , Lin Yang-I , Bitha Panayota , Testa Raymond T.
Published: October 2018

CL 188,624, CL 190,294, and CL 191,121 are novel aminomethyl tetrahydrofuranyl (THF)-1β-methylcarbapenems. The in vitro antibacterial activities of these THF carbapenems were evaluated and compared with those of biapenem, imipenem, and meropenem against 554 recent clinical isolates obtained from geographically distinct medical centers across North America. The antibacterial activities of the THF carbapenems were equivalent to that of biapenem, and the THF carbapenems were slightly more active than imipenem and less active than meropenem against most of the members of the family
Enterobacteriaceae
but lacked significant activity against
Pseudomonas
isolates. In general, CL 191,121 was two- to fourfold more active than CL 188,624 and CL 190,294 against the staphylococcal and enterococcal isolates tested. CL 191,121 was twofold less active than imipenem against methicillin-susceptible staphylococci and was as activity as imipenem against
Enterococcus faecalis
isolates. Biapenem and meropenem were two- and fourfold less active than CL 191,121, respectively, against the methicillin-susceptible staphylococci and
E. faecalis
. All the carbapenems displayed equivalent good activities against the streptococci. Biapenem was slightly more active than the other carbapenems against
Bacteroides fragilis
isolates. Time-kill curve studies demonstrated that the THF carbapenems were bactericidal in 6 h against
Escherichia coli
and
Staphylococcus aureus
isolates. The postantibiotic effect exerted by CL 191,121 was comparable to or slightly longer than that of imipenem against isolates of
S. aureus
,
E. coli
, and
Klebsiella pneumoniae.
Influences of Urinary pH on Ciprofloxacin Pharmacokinetics in Humans and Antimicrobial Activity In Vitro versus Those of Sparfloxacin
Authors: Kamberi Marika , Tsutsumi Kimiko , Kotegawa Tsutomu , Kawano Koichi , Nakamura Koichi , Niki Yoshihito , Nakano Shigeyuki
Published: October 2018

The impact of acidification and alkalinization of urine on the pharmacokinetics of ciprofloxacin was investigated after single 200-mg oral doses were administered to nine healthy male volunteers. In addition, the effect of human urine on the MICs of ciprofloxacin and sparfloxacin against some common urinary tract pathogens such as
Escherichia coli
and
Pseudomonas aeruginosa
was investigated. Acidic and alkaline conditions were achieved by repeated oral doses of ammonium chloride or sodium bicarbonate, respectively. Plasma ciprofloxacin levels in all subjects were adequately described in terms of two-compartment model kinetics with first-order absorption. Acidification and alkalinization treatments had no effect on ciprofloxacin absorption, distribution, or elimination. The total amount of unchanged ciprofloxacin excreted over 24 h under acidic conditions was 88.4 ± 14.5 mg (mean ± standard deviation) (44.2% of the oral dose) and 82.4 ± 16.5 mg (41.2% of the oral dose) under alkaline conditions, while the total amount of unchanged drug excreted over 24 h in volunteers receiving neither sodium bicarbonate nor ammonium chloride was 90.53 ± 9.8 mg (45.2% of the oral dose). The mean renal clearance of ciprofloxacin was 16.78 ± 2.67, 16.08 ± 3.2, and 16.31 ± 2.67 liters/h with acidification, alkalinization, and control, respectively. Renal clearance and concentrations of ciprofloxacin in urine were not correlated with urinary pH. The antibacterial activity of ciprofloxacin and sparfloxacin against
E. coli
NIHJ JC-2 and
P. aeruginosa
ATCC 27853 was affected by human urine and in particular by its pH. The activities of both quinolones against
E. coli
NIHJ JC-2 were lower at lower urinary pH and rather uniform, while in the case of
P. aeruginosa
ATCC 27853 ciprofloxacin was more active than sparfloxacin.
Optimizing Aminoglycoside Therapy for Nosocomial Pneumonia Caused by Gram-Negative Bacteria
Authors: Kashuba Angela D. M. , Nafziger Anne N. , Drusano George L. , Bertino Joseph S.
Published: October 2018

Nosocomial pneumonia is a notable cause of morbidity and mortality and leads to increases in lengths of hospital stays and institutional expenditures. Aminoglycosides are used to treat patients with these infections, but few data on the doses and schedules required to achieve optimal therapeutic outcomes exist. We analyzed aminoglycoside treatment data for 78 patients with nosocomial pneumonia to determine if optimization of aminoglycoside pharmacodynamic parameters results in a more rapid therapeutic response (defined by outcome and days to leukocyte count resolution and temperature resolution). Cox proportional hazards, Classification and Regression Tree (CART), and logistic regression analyses were applied to the data. By all analyses, the first measured maximum concentration of drug in serum (
C
max
)/MIC predicted days to temperature resolution and the second measured
C
max
/MIC predicted days to leukocyte count resolution. For days to temperature resolution and leukocyte count resolution, CART analyses produced breakpoints, with an 89% success rate at 7 days of therapy for a
C
max
/MIC of >4.7 and an 86% success rate at 7 days of therapy for a
C
max
/MIC of >4.5, respectively. Logistic regression analyses predicted a 90% probability of temperature resolution and leukocyte count resolution by day 7 if a
C
max
/MIC of ≥10 is achieved within the first 48 h of aminoglycoside therapy. Aggressive aminoglycoside dosing immediately followed by individualized pharmacokinetic monitoring would ensure that
C
max
/MIC targets are achieved early in therapy. This would increase the probability of a rapid therapeutic response for pneumonia caused by gram-negative bacteria and potentially decreasing durations of parenteral antibiotic therapy, lengths of hospitalization, and institutional expenditures, a situation in which both the patient and the institution benefit.
Esterases in Serum-Containing Growth Media Counteract Chloramphenicol Acetyltransferase Activity In Vitro
Authors: Sohaskey Charles D. , Barbour Alan G.
Published: October 2018

The spirochete
Borrelia burgdorferi
was unexpectedly found to be as susceptible to diacetyl chloramphenicol, the product of the enzyme chloramphenicol acetyltransferase, as it was to chloramphenicol itself. The susceptibilities of
Escherichia coli
and
Bacillus subtilis
, as well as that of
B. burgdorferi
, to diacetyl chloramphenicol were then assayed in different media. All three species were susceptible to diacetyl chloramphenicol when growth media were supplemented with rabbit serum or, to a lesser extent, human serum. Susceptibility of
E. coli
and
B. subtilis
to diacetyl chloramphenicol was not observed in the absence of serum, when horse serum was used, or when the rabbit or human serum was heated first. In the presence of 10% rabbit serum, a strain of
E. coli
bearing the chloramphenicol acetyltransferase (
cat
) gene had a fourfold-lower resistance to chloramphenicol than in the absence of serum. A plate bioassay for chloramphenicol activity showed the conversion by rabbit, mouse, and human sera but not bacterial cell extracts or heated serum of diacetyl chloramphenicol to an inhibitory compound. Deacetylation of acetyl chloramphenicol by serum components was demonstrated by using fluorescent substrates and thin-layer chromatography. These studies indicate that esterases of serum can convert diacetyl chloramphenicol back to an active antibiotic, and thus, in vitro findings may not accurately reflect the level of chloramphenicol resistance by
cat
-bearing bacteria in vivo.
Influence of Renal Failure on Intestinal Clearance of Ciprofloxacin in Rats
Authors: Dautrey Sophie , Rabbaa Lydia , Laouari Denise , Lacour Bernard , Carbon Claude , Farinotti Robert
Published: October 2018

Following intravenous doses, ciprofloxacin pharmacokinetics in control and nephrectomized rats were studied. There were no differences between control and nephrectomized rats for area under the concentration-time curve in plasma or biliary clearance. The intestinal clearance of ciprofloxacin was increased in nephrectomized rats. Intestinal elimination seems to compensate partially for the decrease in urinary excretion of ciprofloxacin in nephrectomized rats.
Multiplex Sequence Analysis Demonstrates the Competitive Growth Advantage of the A-to-G Mutants of Clarithromycin-Resistant
Helicobacter pylori
Authors: Wang Ge , Rahman M. Sayeedur , Humayun M. Zafri , Taylor Diane E.
Published: October 2018

Clarithromycin resistance in
Helicobacter pylori
is due to point mutation within the 23S rRNA. We examined the growth rates of different types of site-directed mutants and demonstrated quantitatively the competitive growth advantage of A-to-G mutants over other types of mutants by a multiplex sequencing assay. The results provide a rational explanation of why A-to-G mutants are predominantly observed among clarithromycin-resistant clinical isolates.
Many Class I Integrons Comprise Distinct Stable Structures Occurring in Different Species of
Enterobacteriaceae
Isolated from Widespread Geographic Regions in Europe
Authors: Martinez-Freijo Pedro , Fluit Ad C. , Schmitz Franz-Joseph , Verhoef Jan , Jones Mark E.
Published: October 2018

Three sizes of inserted regions of DNA (800, 1,000, and 1,500 bp) were shown to be common among class I integrons in unrelated clinical isolates of
Enterobacteriaceae
from different European hospitals. Sequencing showed that 800-bp inserted regions comprised identical sequences including
aacA4
, that 1,000-bp inserted regions included
aadA
, and that 1,500-bp inserted regions included
dfrI
and
aadA1
, irrespective of host species and geographic origin. In addition promoter sequences were mostly identical for each size class. These data suggest that inserted gene cassettes and promoter regions of integrons are conserved and stable, with resistance genes transferred more often as part of the entire integron structure than as individual gene cassettes.
Declining Concentrations of Dihydroartemisinin in Plasma during 5-Day Oral Treatment with Artesunate for Falciparum Malaria
Authors: Khanh Nguyen Xuan , de Vries Peter J. , Ha Le Dang , van Boxtel Chris J. , Koopmans Richard , Kager Piet A.
Published: October 2018

Six patients with uncomplicated falciparum malaria received artesunate for 5 days. Plasma concentrations of artesunate and dihydroartemisinin were determined by high-performance liquid chromatography with electrochemical detection. The concentrations of dihydroartemisinin in plasma 2 h after a dose showed a time-dependent decline. Concentrations of artesunate in plasma especially after the last dose, were very low. Despite this, all patients responded with a fast recovery.