bluuebunny/crossref_metadata_2025 huggingface dataset:
EFFECTS OF DIFFERENT FACTORS ON SENSORY ATTRIBUTES, OVERALL ACCEPTANCE AND PREFERENCE OF ROOIBOS (ASPALATHUS LINEARIS) TEA
Authors: DOS AYSEGUL , AYHAN ZEHRA , SUMNU GULUM
Published: July 2005

Effects of serving size, gender, age, tea type, tea‐making technique and water quality on the sensory attributes (appearance, aroma and taste), overall acceptability and preference of Rooibos tea were investigated. In general, there were significant effects of age and main factor interactions on the attributes and product acceptability (P ≤ 0.05). The interaction of serving size, gender and age affected the acceptability of Rooibos (P ≤ 0.05). Although there was significant difference between natural and vanilla‐flavored tea in terms of color, clarity, odor and taste (P ≤ 0.05), no difference was observed in the product acceptability (P > 0.05). The tea‐making technique only affected the odor, with traditionally brewed tea having lower intensity scores than the infused tea. The tea prepared using natural spring water was found to be clearer and had higher acceptability scores than the tea prepared using tap water (P ≤ 0.05). There was no existing significant preference for the natural Rooibos over the vanilla‐flavored one, and the traditionally brewed tea over the infused one; however, the tea prepared using the spring water was preferred over the one prepared using tap water.
Electron Localization Determines Defect Formation on Ceria Substrates
Authors: Esch Friedrich , Fabris Stefano , Zhou Ling , Montini Tiziano , Africh Cristina , Fornasiero Paolo , Comelli Giovanni , Rosei Renzo
Published: July 2005

The high performance of ceria (CeO
2
) as an oxygen buffer and active support for noble metals in catalysis relies on an efficient supply of lattice oxygen at reaction sites governed by oxygen vacancy formation. We used high-resolution scanning tunneling microscopy and density functional calculations to unravel the local structure of surface and subsurface oxygen vacancies on the (111) surface. Electrons left behind by released oxygen localize on cerium ions. Clusters of more than two vacancies exclusively expose these reduced cerium ions, primarily by including subsurface vacancies, which therefore play a crucial role in the process of vacancy cluster formation. These results have implications for our understanding of oxidation processes on reducible rare-earth oxides.
Inactivation of the
FCY2
Gene Encoding Purine-Cytosine Permease Promotes Cross-Resistance to Flucytosine and Fluconazole in
Candida lusitaniae
Authors: Chapeland-Leclerc Florence , Bouchoux Julien , Goumar Abdelhak , Chastin Christiane , Villard Jean , Noël Thierry
Published: July 2005

In a previous work, we described the possible relationship between a defect of purine-cytosine permease and the acquisition of a cross-resistance to the antifungal combination flucytosine (5FC) and fluconazole (FLC) in
Candida lusitaniae
(T. Noël, F. François, P. Paumard, C. Chastin, D. Brethes, and J. Villard, Antimicrob. Agents Chemother. 47:1275-1284, 2003). Using degenerate PCR and chromosome walking, we cloned two
FCY2
-like genes in
C. lusitaniae
. Northern blot analysis revealed that only one gene was expressed; it was named
FCY2
. The other one behaved as a pseudogene and was named
FCY21
. In order to better characterize the possible role of
FCY2
in cross-resistance to 5FC-FLC, disruption experiments with auxotrophic strain 6936
ura3
(
D95V
)
FCY2
with an integrative vector carrying the
URA3
gene and a partial sequence of the
C. lusitaniae FCY2
gene were undertaken. Southern blot analysis revealed that homologous recombination events occurred in all transformants analyzed at rates of 50% at resident locus
FCY2
and 50% at resident locus
URA3
, resulting in the genotypes
ura3 fcy2
::
URA3
and
ura3
::
URA3 FCY2
, respectively. It was then demonstrated that only transformants harboring a disrupted
fcy2
gene were resistant to 5FC, susceptible to FLC, and resistant to the 5FC-FLC combination. Finally, complementation experiments with a functional
FCY2
gene restored 5FC and FLC susceptibilities to the wild-type levels. The results of this study provide molecular evidence that inactivation of the sole
FCY2
gene promotes cross-resistance to the antifungal association 5FC-FLC in
C. lusitaniae
.
Efficacy of Telavancin in a Rabbit Model of Aortic Valve Endocarditis Due to Methicillin-Resistant
Staphylococcus aureus
or Vancomycin-Intermediate
Staphylococcus aureus
Authors: Madrigal Andres G. , Basuino Li , Chambers Henry F.
Published: July 2005

The activities of telavancin and vancomycin were compared in vitro and in the rabbit model of aortic valve endocarditis against a methicillin-resistant
Staphylococcus aureus
strain, COL, and a vancomycin-intermediate
S. aureus
(VISA) strain, HIP 5836. Telavancin was bactericidal in time-kill studies at a concentration of 5 μg/ml against both COL and HIP5836. Vancomycin was bacteriostatic at 5 μg/ml and bactericidal at 10 μg/ml against COL and was bacteriostatic at 10 μg/ml against VISA strain HIP 5836. Compared to untreated controls, a twice-daily regimen of 30 mg/kg of telavancin reduced mean aortic valve vegetation titers of the COL strain by 4.7 log
10
CFU/g after 4 days of therapy and sterilized 6/11 vegetations compared to 3.4 log
10
CFU/g with 3/10 vegetations sterilized for a regimen of twice-daily vancomycin, 30 mg/kg; these differences were not statistically significant. Telavancin was significantly more effective than vancomycin in the VISA model, producing a 5.5 log
10
CFU/g reduction versus no reduction in CFU with vancomycin. In experiments comparing 2-day regimens of telavancin at 30 mg/kg and 50 mg/kg twice daily, organisms were rapidly eliminated from vegetations, but the effect was not different between the two doses. These results suggest that telavancin may be an effective treatment for endocarditis and other serious staphylococcal infections accompanied by bacteremia, including infections caused by staphylococci not susceptible to vancomycin.
Fluconazole MIC and the Fluconazole Dose/MIC Ratio Correlate with Therapeutic Response among Patients with Candidemia
Authors: Clancy Cornelius J. , Yu Victor L. , Morris Arthur J. , Snydman David R. , Nguyen M. Hong
Published: July 2005

We tested 32
Candida
isolates recovered in the early 1990s from the bloodstreams of patients with candidemia for in vitro susceptibility to fluconazole and determined if MIC and/or the daily dose of fluconazole/MIC ratio correlated with the response to therapy. This is a unique data set since 87.5% (28/32) of patients were treated with fluconazole doses now considered to be inadequate (≤200 mg), which contributed to high therapeutic failure rates (53% [17/32]). The geometric mean MIC and dose/MIC ratio for isolates associated with therapeutic failure (11.55 μg/ml and 14.3, respectively) differed significantly from values associated with therapeutic success (0.95 μg/ml and 219.36 [
P
= 0.0009 and 0.0004, respectively]). The therapeutic success rates among patients infected with susceptible (MIC ≤ 8 μg/ml), susceptible-dose dependent (S-DD) (MIC = 16 or 32 μg/ml), and resistant (MIC ≥ 64 μg/ml) isolates were 67% (14/21), 20% (1/5), and 0% (0/6), respectively. A dose/MIC ratio >50 was associated with a success rate of 74% (14/19), compared to 8% (1/13) for a dose/MIC ratio ≤50 (
P
= 0.0003). Our data suggest that both fluconazole MIC and dose/MIC ratio correlate with the therapeutic response to fluconazole among patients with candidemia. In clinical practice, dose/MIC ratio might prove easier to interpret than breakpoint MICs, since it quantitates the effects of increasing fluconazole doses that are alluded to in the S-DD designation.
Pharmacodynamic Evidence that Ciprofloxacin Failure against Tuberculosis Is Not Due to Poor Microbial Kill but to Rapid Emergence of Resistance
Authors: Gumbo Tawanda , Louie Arnold , Deziel Mark R. , Drusano George L.
Published: July 2005

Studies of early bactericidal activity provide a fast and economic way to evaluate the clinical efficacy of potential agents for the treatment of tuberculosis. Based on good early bactericidal activity data, ciprofloxacin entered further studies and is now recommended as part of treatment for multidrug-resistant tuberculosis. We examined the relationship between ciprofloxacin bactericidal activity and the emergence of resistance in an in vitro pharmacodynamic infection model in which we exposed
Mycobacterium tuberculosis
to simulated free-drug ciprofloxacin serum concentration-time profiles that mimic those encountered in humans treated with ciprofloxacin orally for 2 weeks.
Mycobacterium tuberculosis
cultures were sampled during the experiment in order to determine the effect of therapy on the total microbial population as well as the drug-resistant population. The ciprofloxacin regimen, which achieved a ratio of the area under the concentration time curve from 0 to 24 h to MIC of 80.4, resulted in a rapid microbial kill similar to that encountered in humans during studies of early bactericidal activity. However, despite this impressive bactericidal activity, resistance emerged quickly. By the end of the first week, most of the microbial population had been replaced by a ciprofloxacin-resistant population. Given the MICs encountered in clinical isolates of
M. tuberculosis
, we estimate that most clinically tolerable doses of ciprofloxacin will lead to emergence of resistance, especially when used as the only effective component of regimens given for treatment of multidrug-resistant tuberculosis. One of the explanations for why early bactericidal activity fails to predict sterilization may be the emergence of a resistant subpopulation, which only becomes ≥1% at the end of the early bactericidal activity studies.
In Vitro and In Vivo Evaluations of the Activities of Lauric Acid Monoester Formulations against
Staphylococcus aureus
Authors: Rouse Mark S. , Rotger Margalida , Piper Kerryl E. , Steckelberg James M. , Scholz Matthew , Andrews Jeffrey , Patel Robin
Published: July 2005

Due to increasing mupirocin resistance, alternatives for
Staphylococcus aureus
nasal decolonization are needed. Lauric acid monoesters combined with lactic, mandelic, malic, or benzoic acid are being evaluated as possible alternatives. We determined the in vitro activity of 13 lauric acid monoester (LAM) formulations and mupirocin against 30 methicillin-susceptible
S. aureus
(MSSA) isolates and 30 methicillin-resistant
S. aureus
(MRSA) isolates. We then used a murine model of MRSA nasopharyngeal colonization to compare the in vivo activity of mupirocin with three LAM formulations. MSSA and MRSA MIC
90
values were 0.25 μg/ml for mupirocin and ≤4 μl/ml for all LAM formulations tested. Hsd:ICR mice were challenged with 10
8
CFU/naris MRSA. Five days later,
S. aureus
colonization was documented by culture. Treatment with bland, mupirocin, or one of three LAM ointments was then administered unblinded thrice daily for 2 days. Three days after treatment, both anterior nares were cultured for
S. aureus
. Administration of 128774-49E or 128774-53A was associated with greater eradication of MRSA carriage (24/34 [71%] or 33/40 [83%]) of animals, respectively) than bland ointment (12/38 [32%]) (
P
< 0.005). 128774-53A administration resulted in greater MRSA carriage eradication than mupirocin (19/38 [50%]) (
P
< 0.005) in this model. LAM formulations warrant evaluation for
S. aureus
nasal decolonization in humans.
Molecular Analysis of Cross-Resistance to Capreomycin, Kanamycin, Amikacin, and Viomycin in
Mycobacterium tuberculosis
Authors: Maus Courtney E. , Plikaytis Bonnie B. , Shinnick Thomas M.
Published: July 2005

Capreomycin, kanamycin, amikacin, and viomycin are drugs that are used to treat multidrug-resistant tuberculosis. Each inhibits translation, and cross-resistance to them is a concern during therapy. A recent study revealed that mutation of the
tlyA
gene, encoding a putative rRNA methyltransferase, confers capreomycin and viomycin resistance in
Mycobacterium tuberculosis
bacteria. Mutations in the 16S rRNA gene (
rrs
) have been associated with resistance to each of the drugs; however, reports of cross-resistance to the drugs have been variable. We investigated the role of
rrs
mutations in capreomycin resistance and examined the molecular basis of cross-resistance to the four drugs in
M. tuberculosis
laboratory-generated mutants and clinical isolates. Spontaneous mutants were generated to the drugs singularly and in combination by plating on medium containing one or two drugs. The frequencies of recovery of the mutants on single- and dual-drug plates were consistent with single-step mutations. The
rrs
genes of all mutants were sequenced, and the
tlyA
genes were sequenced for mutants selected on capreomycin, viomycin, or both; MICs of all four drugs were determined. Three
rrs
mutations (A1401G, C1402T, and G1484T) were found, and each was associated with a particular cross-resistance pattern. Similar mutations and cross-resistance patterns were found in drug-resistant clinical isolates. Overall, the data implicate
rrs
mutations as a molecular basis for resistance to each of the four drugs. Furthermore, the genotypic and phenotypic differences seen in the development of cross-resistance when
M. tuberculosis
bacteria were exposed to one or two drugs have implications for selection of treatment regimens.
Peptide Nucleic Acid Antisense Oligomer as a Therapeutic Strategy against Bacterial Infection: Proof of Principle Using Mouse Intraperitoneal Infection
Authors: Tan Xin-Xing , Actor Jeffrey K. , Chen Yin
Published: July 2005

Antisense oligodeoxynucleotides (ODNs) and their analogs have been successfully utilized to inhibit gene expression and bacterial growth in vitro or in cell culture. In this study,acpP-targeting antisense peptide nucleic acid (PNA) and its peptide conjugate were tested as potential antibacterial agents in two groups of experiments using a mouse model. In the first group,Escherichia colimutant strain SM101 with a defective outer membrane was used to induce bacteremia and peritonitis in BALB/c mice by intraperitoneal (i.p.) injection. The resulting bacteremia was fatal within 48 h. A single i.p injection of 5 nmol (or more) of PNA administered 30 min before bacterial challenge significantly reduced the bacterial load in mouse blood. Reductions in serum concentrations of the proinflammatory cytokines tumor necrosis factor alpha, interleukin-1β (IL-1β), IL-6, and IL-12 were also observed. PNA treatment was effective in rescuing 100% of infected animals. In the second group, bacteremia in BALB/c mice was induced by i.p. injection ofE. coliwild-type strain K-12. The infected mice were treated by a single intravenous injection of peptide-PNA conjugate 30 min after bacterial challenge. Treatment with the peptide-PNA conjugate significantly reduced the K-12 load, with modest reduction in cytokine concentrations. The conjugate treatment was also able to rescue up to 60% of infected animals. This report is the first demonstration of ODNs' antibacterial efficacy in an animal disease model. The ability of PNA and its peptide conjugate to inhibit bacterial growth and to prevent fatal infection demonstrates the potential for this new class of antibacterial agents.
Activity of the De Novo Engineered Antimicrobial Peptide WLBU2 against
Pseudomonas aeruginosa
in Human Serum and Whole Blood: Implications for Systemic Applications
Authors: Deslouches Berthony , Islam Kazi , Craigo Jodi K. , Paranjape Shruti M. , Montelaro Ronald C. , Mietzner Timothy A.
Published: July 2005

Cationic amphipathic peptides have been extensively investigated as a potential source of new antimicrobials that can complement current antibiotic regimens in the face of emerging drug-resistant bacteria. However, the suppression of antimicrobial activity under certain biologically relevant conditions (e.g., serum and physiological salt concentrations) has hampered efforts to develop safe and effective antimicrobial peptides for clinical use. We have analyzed the activity and selectivity of the human peptide LL37 and the de novo engineered antimicrobial peptide WLBU2 in several biologically relevant conditions. The host-derived synthetic peptide LL37 displayed high activity against
Pseudomonas aeruginosa
but demonstrated staphylococcus-specific sensitivity to NaCl concentrations varying from 50 to 300 mM. Moreover, LL37 potency was variably suppressed in the presence of 1 to 6 mM Mg
2+
and Ca
2+
ions. In contrast, WLBU2 maintained its activity in NaCl and physiologic serum concentrations of Mg
2+
and Ca
2+
. WLBU2 is able to kill
P. aeruginosa
(10
6
CFU/ml) in human serum, with a minimum bactericidal concentration of <9 μM. Conversely, LL37 is inactive in the presence of human serum. Bacterial killing kinetic assays in serum revealed that WLBU2 achieved complete bacterial killing in 20 min. Consistent with these results was the ability of WLBU2 (15 to 20 μM) to eradicate bacteria from ex vivo samples of whole blood. The selectivity of WLBU2 was further demonstrated by its ability to specifically eliminate
P. aeruginosa
in coculture with human monocytes or skin fibroblasts without detectable adverse effects to the host cells. Finally, WLBU2 displayed potent efficacy against
P. aeruginosa
in an intraperitoneal infection model using female Swiss Webster mice. These results establish a potential application of WLBU2 in the treatment of bacterial sepsis.
Ciprofloxacin Induction of a Susceptibility Determinant in
Pseudomonas aeruginosa
Authors: Brazas Michelle D. , Hancock Robert E. W.
Published: July 2005

With few novel antimicrobials in development, resistance to the current selection of antibiotics increasingly encroaches on our ability to control microbial infections. One limitation in our understanding of the basis of the constraints on current therapies is our poor understanding of antibiotic interactions with bacteria on a global scale. Custom DNA microarrays were used to characterize the response of
Pseudomonas aeruginosa
to ciprofloxacin, a fluoroquinolone commonly used in therapy against chronic infections by this intrinsically resistant bacterium. Of the approximately 5,300 open reading frames (ORFs) on the array, 941 genes showed statistically significant (
P
≤ 0.05) differential expression in response to 0.3× MIC of ciprofloxacin; 554 were promoted and 387 were repressed. Most striking among the responsive genes was the region between PA0613 and PA0648, which codes for the bacteriophage-like R2/F2 pyocins. In this region, virtually every ORF was increased by 0.3× MIC of ciprofloxacin and even more dramatically up-regulated (7- to 19-fold) following treatment with 1× MIC of ciprofloxacin. Pyocin gene expression was confirmed with
lux
reporter mutants and real-time PCR studies; pyocin-like particles were also present in transmission electron micrographs of supernatants from cells treated with 1× MIC of ciprofloxacin. Interestingly, mutants in this region exhibited ≥8-fold-increased resistance to ciprofloxacin and other fluoroquinolones, demonstrating that this region is a susceptibility determinant. Since this region is known to be variably present in the genomes of clinical isolates of
P. aeruginosa
(R. K. Ernst et al., Environ. Microbiol.
5:
1341-1349, 2003, and M. C. Wolfgang et al., Proc. Natl. Acad. Sci. USA
100:
8484-8489, 2003), these findings demonstrate that the R2/F2 pyocin region is a “loaded gun” that can mediate fluoroquinolone susceptibility in
P. aeruginosa
.
Effect of Ciprofloxacin-Induced Prostaglandin E2on Interleukin-18-Treated Monocytes
Authors: Takahashi Hideo Kohka , Iwagaki Hiromi , Xue Dong , Katsuno Goutarou , Sugita Sachi , Mizuno Kenji , Mori Shuji , Saito Shinya , Yoshino Tadashi , Tanaka Noriaki , Nishibori Masahiro
Published: July 2005

Ciprofloxacin, a fluorinated 4-quinolone, is useful for the clinical treatment of infections due to its antibacterial properties and also modulates the immune response of monocytes isolated from human peripheral blood mononuclear cells. In the present study, we found that ciprofloxacin induced the production of prostaglandin E2in monocytes in a concentration-dependent manner regardless of the presence of interleukin-18 by enhancing the expression of cyclooxygenase-2 protein and that this in turn led to the elevation of intercellular cyclic AMP in monocytes via the stimulation of prostaglandin receptors. The prostaglandin E2and cyclic AMP production increased by ciprofloxacin was inhibited by indomethacin, a nonselective cyclooxygenase-2 inhibitor, and NS398, a selective cyclooxygenase-2 inhibitor. In addition, ciprofloxacin suppressed the interleukin-18-induced production of tumor necrosis factor alpha, gamma interferon, and interleukin-12 in peripheral blood mononuclear cells by inhibiting the expression of intercellular adhesion molecule 1, B7.1, B7.2, and CD40 on monocytes, and this effect could be reversed by the addition of indomethacin or NS398. These results indicate that ciprofloxacin exerts immunomodulatory activity via the production of prostaglandin E2and imply therapeutic potential of ciprofloxacin for the treatment of systemic inflammatory responses initiated by interleukin-18.
In Vitro and In Vivo Antibacterial Activities of CS-023 (RO4908463), a Novel Parenteral Carbapenem
Authors: Koga Tetsufumi , Abe Tomomi , Inoue Harumi , Takenouchi Takashi , Kitayama Akiko , Yoshida Tatsuhiko , Masuda Nobuhisa , Sugihara Chika , Kakuta Masayo , Nakagawa Miyuki , Shibayama Takahiro , Matsushita Yoko , Hirota Takashi , Ohya Satoshi , Utsui Yukio , Fukuoka Takashi , Kuwahara Syogo
Published: July 2005

CS-023 (RO4908463, formerly R-115685) is a novel 1β-methylcarbapenem with 5-substituted pyrrolidin-3-ylthio groups, including an amidine moiety at the C-2 position. Its antibacterial activity was tested against 1,214 clinical isolates of 32 species and was compared with those of imipenem, meropenem, ceftazidime, ceftriaxone, ampicillin, amikacin, and levofloxacin. CS-023 exhibited a broad spectrum of activity against gram-positive and -negative aerobes and anaerobes, including methicillin-resistant
Staphylococcus aureus
(MRSA), methicillin-resistant
Staphylococcus epidermidis
, penicillin-resistant
Streptococcus pneumoniae
(PRSP), β-lactamase-negative ampicillin-resistant
Haemophilus influenzae
, and
Pseudomonas aeruginosa
. CS-023 showed the most potent activity among the compounds tested against
P. aeruginosa
and MRSA, with MICs at which 90% of isolates tested were inhibited of 4 μg/ml and 8 μg/ml, respectively. CS-023 was stable against hydrolysis by the β-lactamases from
Enterobacter cloacae
and
Proteus vulgaris
. CS-023 also showed potent activity against extended-spectrum β-lactamase-producing
Escherichia coli
. The in vivo efficacy of CS-023 was evaluated with a murine systemic infection model induced by 13 strains of gram-positive and -negative pathogens and a lung infection model induced by 2 strains of PRSP (serotypes 6 and 19). Against the systemic infections with PRSP, MRSA, and
P. aeruginosa
and the lung infections, the efficacy of CS-023 was comparable to those of imipenem/cilastatin and vancomycin (tested against lung infections only) and superior to those of meropenem, ceftriaxone, and ceftazidime (tested against
P. aeruginosa
infections only). These results suggest that CS-023 has potential for the treatment of nosocomial bacterial infections by gram-positive and -negative pathogens, including MRSA and
P. aeruginosa
.
Comparison of Four Methods for Determining Lysostaphin Susceptibility of Various Strains of
Staphylococcus aureus
Authors: Kusuma Caroline M. , Kokai-Kun John F.
Published: July 2005

Lysostaphin is an endopeptidase that cleaves the pentaglycine cross-bridges of the staphylococcal cell wall rapidly lysing the bacteria. Recently, lysostaphin has been examined for its potential to treat infections and to clear
Staphylococcus aureus
nasal colonization, requiring a reliable method for determining the lysostaphin susceptibility of strains of
S. aureus
. We compared four methods for determining the lysostaphin susceptibility of 57 strains of methicillin-sensitive
S. aureus
, methicillin-resistant
S. aureus
, vancomycin intermediately susceptible
S. aureus
(VISA), mupirocin-resistant
S. aureus
, and various defined genetic mutants of
S. aureus
. Three reference lysostaphin-resistant
S. aureus
variants were also included in the assays as negative controls. The assays examined included turbidity, MIC, minimum bactericidal concentration (MBC), and disk diffusion assays. All of the strains of
S. aureus
tested, including a VISA strain which had previously been reported to be lysostaphin resistant, were susceptible to lysostaphin by all four methods. The three reference lysostaphin-resistant variants were resistant by all four methods. The disk diffusion assay was the simplest method to differentiate lysostaphin-susceptible
S. aureus
strains from lysostaphin-resistant variants, while the MBC assay could be used as a follow-up assay if required. In the disk diffusion assay, all strains of
S. aureus
tested revealed zones of inhibition of ≥11 mm using a 50-μg lysostaphin disk, while the three reference lysostaphin-resistant
S. aureus
variants had no zones of inhibition. In MBC assays, concentrations of lysostaphin ranging from 0.16 μg/ml to 2.5 μg/ml were found to cause a 3 log or greater drop from the initial CFU of
S. aureus
within 30 min for all strains tested.
Secretion of Proteases by
Pseudomonas aeruginosa
Biofilms Exposed to Ciprofloxacin
Authors: Ołdak Ewa , Trafny Elżbieta A.
Published: July 2005

Pseudomonas aeruginosa
proteases are considered important virulence factors which damage host tissues and interfere with host antibacterial defense mechanisms.
P. aeruginosa
biofilm cells are not completely killed by antibacterials, and therefore this study addresses the question whether ciprofloxacin attenuates the virulence of biofilm communities by abolishing their secretion of proteases. The surviving cells of the colony biofilms studied, despite their cyclical exposure to four doses of ciprofloxacin at bactericidal concentrations (one dose a day), still secreted active proteases to the environment surrounding the biofilms. The biofilm cells secreted elastase B (LasB) over the duration of the experiments as confirmed by Western immunoblot analysis. The colony biofilms did not secrete LasA—a protease with staphylolytic activity. The same profiles on zymogram gels with gelatin were observed for the proteases secreted by both ciprofloxacin-exposed and unexposed (control) biofilms. Total proteolytic activities of the colony biofilms studied were significantly reduced after exposure to ciprofloxacin at bactericidal concentrations—after 96 h of exposure they dropped to 38% for the strain intermediate resistant to ciprofloxacin and to 65% for the strain highly resistant to the antibiotic, relative to the control biofilms. The surviving cells of the colony biofilms after their release into a fresh medium displayed transient increased resistance to ciprofloxacin compared to their planktonic counterparts.
Systemic Antibacterial Activity of Novel Synthetic Cyclic Peptides
Authors: Dartois Véronique , Sanchez-Quesada Jorge , Cabezas Edelmira , Chi Ellen , Dubbelde Chad , Dunn Carrie , Granja Juan , Gritzen Colleen , Weinberger Dana , Ghadiri M. Reza , Parr Thomas R.
Published: July 2005

Cyclic peptides with an even number of alternatingd,l-α-amino acid residues are known to self-assemble into organic nanotubes. Such peptides previously have been shown to be stable upon protease treatment, membrane active, and bactericidal and to exert antimicrobial activity againstStaphylococcus aureusand other gram-positive bacteria. The present report describes the in vitro and in vivo pharmacology of selected members of this cyclic peptide family. The intravenous (i.v.) efficacy of six compounds with MICs of less than 12 μg/ml was tested in peritonitis and neutropenic-mouse thigh infection models. Four of the six peptides were efficacious in vivo, with 50% effective doses in the peritonitis model ranging between 4.0 and 6.7 mg/kg against methicillin-sensitiveS. aureus(MSSA). In the thigh infection model, the four peptides reduced the bacterial load 2.1 to 3.0 log units following administration of an 8-mg/kg i.v. dose. Activity against methicillin-resistantS. aureuswas similar to MSSA. The murine pharmacokinetic profile of each compound was determined following i.v. bolus injection. Interestingly, those compounds with poor efficacy in vivo displayed a significantly lower maximum concentration of the drug in serum and a higher volume of distribution at steady state than compounds with good therapeutic properties.S. aureuswas unable to easily develop spontaneous resistance upon prolonged exposure to the peptides at sublethal concentrations, in agreement with the proposed interaction with multiple components of the bacterial membrane canopy. Although additional structure-activity relationship studies are required to improve the therapeutic window of this class of antimicrobial peptides, our results suggest that these amphipathic cyclicd,l-α-peptides have potential for systemic administration and treatment of otherwise antibiotic-resistant infections.
Safety, Tolerability, and Pharmacokinetics of Micafungin (FK463) in Febrile Neutropenic Pediatric Patients
Authors: Seibel Nita L. , Schwartz Cindy , Arrieta Antonio , Flynn Patricia , Shad Aziza , Albano Edith , Keirns James , Lau Wendi M. , Facklam David P. , Buell Donald N. , Walsh Thomas J.
Published: July 2005

Micafungin (FK463) is a new parenteral echinocandin. A multicenter, phase I, open-label, sequential-group dose escalation study was conducted to assess the safety, tolerability, and pharmacokinetics of micafungin in neutropenic pediatric patients. A total of 77 patients stratified by age (2 to 12 and 13 to 17 years) received micafungin. Therapy was initiated at 0.5 mg/kg per day and escalated to higher dose levels of 1.0, 1.5, 2.0, 3.0, and 4.0 mg/kg per day. Micafungin was administered within 24 h of initiating broad-spectrum antibacterial antibiotics for the new onset of fever and neutropenia. The most common overall adverse events in the study population were diarrhea (19.5%), epistaxis (18.2%), abdominal pain (16.9%), and headache (16.9%). Nine patients (12%) experienced adverse events considered by the investigator to be possibly related to the study drug. The most common related events were diarrhea, vomiting, and headache, all occurring in two patients each. There was no evidence of a dose-limiting toxicity as defined within the prespecified criteria of this clinical protocol. There was one death during the study due to septic shock. The pharmacokinetic profiles for micafungin over the 0.5- to 4.0-mg/kg dose range demonstrated dose linearity. Clearance, volume of distribution, and half-life remained relatively constant over the dose range and did not change with repeated administration. The overall plasma pharmacokinetic profile was similar to that observed in adults. However, there was an inverse relation between age and clearance. For patients 2 to 8 years old, clearance was approximately 1.35 times that of patients ≥9 years of age. In summary, micafungin over a dosage range between 0.5 and 4.0 mg/kg/day in 77 febrile neutropenic pediatric patients displayed linear pharmacokinetics and increased clearance as a function of decreasing age.
Anti-Human Immunodeficiency Virus Type 1 Activity and Resistance Profile of 2′,3′-Didehydro-3′-Deoxy-4′-Ethynylthymidine In Vitro
Authors: Nitanda Takao , Wang Xin , Kumamoto Hiroki , Haraguchi Kazuhiro , Tanaka Hiromichi , Cheng Yung-Chi , Baba Masanori
Published: July 2005

2′,3′-Didehydro-3′-deoxy-4′-ethynylthymidine (4′-Ed4T) has been identified as a novel nucleoside analog with potent and selective anti-human immunodeficiency virus type 1 (HIV-1) activity and weak cytotoxicity in cell cultures. 4′-Ed4T proved to be 5- to 10-fold more active than its structurally related compound, stavudine (d4T). However, the drug resistance profile of 4′-Ed4T was different from those of d4T and other existing HIV-1 nucleoside reverse transcriptase inhibitors (NRTIs). Approximately 6- to 11-fold decreases in susceptibility to 4′-Ed4T were observed for HIV-1 carrying NRTI-associated mutations (D67N, K70R, T215F, and K219Q) or the lamivudine (3TC)-resistant mutation M184V. In contrast, the susceptibility of the virus carrying the K65R mutation or the multidrug-resistant mutation with the Q151M complex (A62V, V75I, F77L, F116Y, and Q151M) was not altered. Furthermore, the activity of 4′-Ed4T appeared to be enhanced in the presence of K103N, a major nonnucleoside reverse transcriptase inhibitor-resistant mutation. Although 4′-Ed4T was 4.5- to 17.5-fold less active against multidrug-resistant clinical isolates than against a reference strain isolated from a treatment-naïve patient, it was still inhibitory to these isolates at low concentrations. Analysis of 4′-Ed4T-resistant HIV-1 obtained through in vitro selection revealed that the virus was also resistant to 3TC and had two amino acid mutations (P119S and T165A) in addition to the M184V mutation. Since 4′-Ed4T has increased anti-HIV-1 activity, decreased cytotoxicity, and a different resistance profile, it should be considered for further development as a new member of NRTIs.
Hypermutation Is a Key Factor in Development of Multiple-Antimicrobial Resistance inPseudomonas aeruginosaStrains Causing Chronic Lung Infections
Authors: Maciá María D. , Blanquer David , Togores Bernat , Sauleda Jaume , Pérez José L. , Oliver Antonio
Published: July 2005

Pseudomonas aeruginosais the most relevant pathogen producing chronic lung infections in patients with chronic underlying diseases such as cystic fibrosis (CF), bronchiectasis, and chronic obstructive pulmonary disease (COPD). Hypermutable (or mutator)P. aeruginosastrains, characterized by increased (up to 1,000-fold) spontaneous mutation rates due to alterations of the DNA mismatch repair (MMR) system have been found at high frequencies in the lungs of CF patients, but their role in other chronic processes is still unknown. Sixty-twoP. aeruginosaisolates from 30 patients with underlying non-CF chronic respiratory diseases (22 with bronchiectasis and 8 with COPD) and documented chronic infection were studied. Antibiotic susceptibility profiles and mutation frequencies were determined, and complementation assays using the cloned wild-typemutSgene and molecular epidemiology studies (pulsed-field electrophoresis, [PFGE]) were performed with these strains. Thirty-three (53%) of the isolates were hypermutable, and 17 (57%) of the 30 patients were colonized by hypermutable strains. Strains from 11 of the 17 patients were found to be defective in the MMRmutSgene by complementation assays. Interpatient transmission of strains was ruled out by PFGE. Multiple-antimicrobial resistance was documented in 42% of the hypermutable strains in contrast to 0% resistance in the nonhypermutable strains (P< 0.0001). HypermutableP. aeruginosastrains are extremely prevalent in chronic infections in contrast to what has been described in acute processes, suggesting a role of hypermutation in bacterial adaptation for long-term persistence. Furthermore, hypermutation is found to be a key factor for the development of multiple-antimicrobial resistance, and therefore these findings are expected to have important consequences for the treatment of chronic infections.
Azithromycin Inhibits the Formation of Flagellar Filaments without Suppressing Flagellin Synthesis in
Salmonella enterica
Serovar Typhimurium
Authors: Matsui Hidenori , Eguchi Masahiro , Ohsumi Katsufumi , Nakamura Akio , Isshiki Yasunori , Sekiya Kachiko , Kikuchi Yuji , Nagamitsu Tohru , Masuma Rokuro , Sunazuka Toshiaki , Omura Satoshi
Published: July 2005

The present study shows that a sub-MIC of the macrolide antibiotic azithromycin (AZM) diminishes the virulence function of
Salmonella enterica
serovar Typhimurium. We first constructed an AZM-resistant strain (MS248) by introducing
ermBC
, an erythromycin ribosome methylase gene, into serovar Typhimurium. The MIC of AZM for MS248 exceeded 100 μg/ml. Second, we managed to determine the efficacy with which a sub-MIC of AZM reduced the virulence of MS248 in vitro. On the one hand, AZM (10 μg/ml) in the culture medium was unable to inhibit the total protein synthesis, growth rate, or survival within macrophages of MS248. On the other hand, AZM (10 μg/ml) reduced MS248's swarming and swimming motilities in addition to its invasive activity in Henle-407 cells. Electron micrographs revealed no flagellar filaments on the surface of MS248 after overnight growth in L broth supplemented with AZM (10 μg/ml). However, immunoblotting analysis showed that flagellin (FliC) was fully synthesized within the bacterial cells in the presence of AZM (10 μg/ml). In contrast, the same concentration of AZM reduced the export of FliC to the culture medium. These results indicate that a sub-MIC of AZM was able to affect the formation of flagellar filaments, specifically by reducing the amount of flagellin exported from bacterial cells, but it was not involved in suppressing the synthesis of flagellin. Unfortunately, AZM treatment was ineffective against murine salmonellosis caused by MS248.
Selection of a Macaque Fab with Framework Regions Like Those in Humans, High Affinity, and Ability To Neutralize the Protective Antigen (PA) of
Bacillus anthracis
by Binding to the Segment of PA between Residues 686 and 694
Authors: Laffly Emmanuelle , Danjou Ludivine , Condemine Florence , Vidal Dominique , Drouet Emmanuel , Lefranc Marie-Paule , Bottex Chantal , Thullier Philippe
Published: July 2005

Human anthrax infection cannot always be treated successfully by antibiotics, as highlighted by recent bioterrorist attacks; thus, adjunct therapies are clearly needed for the future. There is a particular need to further develop adjunct therapies that can neutralize secreted toxins, such as antibodies directed towards the 83-kDa protective antigen (PA
83
). In the absence of human donors, we immunized a macaque (
Macaca fascicularis
) with PA
83
to obtain such antibodies suitable as an adjunct therapy for human anthrax infection. By using bone marrow as a template, we PCR amplified specific Fab-encoding genes and cloned them as an immune library (10
7
clones). We isolated a high-affinity (equilibrium dissociation constant [

K
D

], 3.4 nM), highly neutralizing (50% inhibitory concentration, 5.6 ± 0.13 nM) Fab (designated 35PA
83
) from this library by panning. Its epitope was localized by Pepscan analysis between residues 686 and 694 of PA
83
and is part of the region which directly interacts with the cell receptor. 35PA
83
may thus neutralize the anthrax toxin by competing directly for its receptor. The genes encoding 35PA
83
were similar to those of a human immunoglobulin germ line and were assigned to subgroups of human V, (D), or J genes by IMGT/V-QUEST analysis. The 35PA
83
framework regions were 92% identical to a representative allele of each subgroup. When compared to framework regions coded by related human germ line genes, only 2 of 74 (VH) or 75 (VK) analyzed amino acids of 35PA
83
have different chemical characteristics. A very high degree of identity with human framework regions makes 35PA
83
well suited for expression as a whole primatized immunoglobulin G and demonstrates the practicality of using macaque Fabs when immunized human plasma cell donors are not available.
Amino Acid Residues That Contribute to Substrate Specificity of Class A β-Lactamase SME-1
Authors: Majiduddin Fahd K. , Palzkill Timothy
Published: July 2005

Carbapenem antibiotics are used as antibiotics of last resort because they possess a broad spectrum of antimicrobial activity and are not easily hydrolyzed by β-lactamases. Recently, class A enzymes, such as the SME-1, NMC-A, and IMI-1 β-lactamases, have been identified with the capacity to hydrolyze carbapenem antibiotics. Traditional class A β-lactamases, such as TEM-1 and SHV-1, are unable to hydrolyze carbapenem antibiotics and exhibit some differences in sequence from those that are able to hydrolyze carbapenem antibiotics. The positions that differ may contribute to the unique substrate specificity of the class A carbapenemase SME-1. Codons in the SME-1 gene representing residues 104, 105, 132, 167, 237, and 241 were randomized by site-directed mutagenesis, and functional mutants were selected for the ability to hydrolyze imipenem, ampicillin, or cefotaxime. Although several positions are important for hydrolysis of β-lactam antibiotics, no single position was found to uniquely contribute to carbapenem hydrolysis. The results of this study support a model whereby the carbapenemase activity of SME-1 is due to a highly distributed set of interactions that subtly alter the structure of the active-site pocket.
Establishment of a Superficial Skin Infection Model in Mice by Using
Staphylococcus aureus
and
Streptococcus pyogenes
Authors: Kugelberg Elisabeth , Norström Tobias , Petersen Thomas K. , Duvold Tore , Andersson Dan I. , Hughes Diarmaid
Published: July 2005

A new animal model for the purpose of studying superficial infections is presented. In this model an infection is established by disruption of the skin barrier by partial removal of the epidermal layer by tape stripping and subsequent application of the pathogens
Staphylococcus aureus
and
Streptococcus pyogenes
. The infection and the infection route are purely topical, in contrast to those used in previously described animal models in mice, such as the skin suture-wound model, where the infection is introduced into the deeper layers of the skin. Thus, the present model is considered more biologically relevant for the study of superficial skin infections in mice and humans. Established topical antibiotic treatments are shown to be effective. The procedures involved in the model are simple, a feature that increases throughput and reproducibility. This new model should be applicable to the evaluation of novel antimicrobial treatments of superficial infections caused by
S. aureus
and
S. pyogenes
.
Triazine Inhibits
Toxoplasma gondii
Tachyzoites In Vitro and In Vivo
Authors: Mui Ernest J. , Jacobus David , Milhous Wilbur K. , Schiehser Guy , Hsu Honghue , Roberts Craig W. , Kirisits Michael J. , McLeod Rima
Published: July 2005

The triazine WR99210 [4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine] inhibits
Toxoplasma gondii
in vitro at nanomolar levels (
P
< 0.05). The 50% inhibitory concentration (IC
50
) was approximately 50 nM. It is a potent inhibitor in vitro and is also effective in vivo. Administration of WR99210 parenterally (i.e., intraperitoneally) reduced the mean number of RH strain tachyzoites present in peritoneal fluid substantially 4 days after intraperitoneal infection of mice. There was a mean of approximately 35 million parasites in control mice as contrasted with approximately 2 million parasites in mice treated with 1.25 mg WR99210/kg of body weight in a representative experiment (
P
< 0.05). In addition the prodrug PS-15 N′-[3-(2,4, 5-trichlorophenoxy)propyloxy]-N9-(1-methylethyl) imidocarbonimidicdiamide is converted to 4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine in vivo when the prodrug is administered orally. PS-15 administered by gavage also reduced intraperitoneal RH strain
T. gondii
tachyzoite numbers. WR99210 has high efficacy and relatively low toxicity because of its substantial effect on
T. gondii
dihydrofolate reductase (DHFR) but not the mammalian host DHFR. Amino acid sequences of
T. gondii
,
Plasmodium falciparum
, and
Homo sapiens
DHFRs were compared. It is of interest that of the DHFR amino acids considered to be interacting with WR99210 in
P. falciparum
within interatomic distances within 3 to 5 Å, four of eight were shared with
T. gondii
DHFR.
H. sapiens
also shared four amino acids thought to be interacting with WR99210. Efficacy of intraperitoneal administration of WR99210 and peroral administration of PS-15 demonstrate the potential usefulness of this class of compounds in treatment of toxoplasmosis administered either parenterally or perorally. The recent development program for this class of antimicrobials as antimalarials makes our proof of principle of improved efficacy of triazines (compared with the gold standard treatment, pyrimethamine) against
T. gondii
especially promising.
TAK-220, a Novel Small-Molecule CCR5 Antagonist, Has Favorable Anti-Human Immunodeficiency Virus Interactions with Other Antiretrovirals In Vitro
Authors: Tremblay Cécile L. , Giguel Françoise , Guan Yongbiao , Chou Ting-Chao , Takashima Katsunori , Hirsch Martin S.
Published: July 2005

TAK-220 is a CCR5 antagonist, part of the new class of anti-human immunodeficiency virus type 1 (anti-HIV-1) entry inhibitors. We evaluated the anti-HIV-1 interactions between TAK-220 and various antiretrovirals in vitro. Synergy was observed with all drugs at the 90 and 95% inhibitory concentrations. The favorable drug interactions observed suggest that further clinical evaluation is warranted.
First Detection of a Carbapenem-Hydrolyzing Metalloenzyme in Two
Enterobacteriaceae
Isolates in Spain
Authors: Tórtola M. Teresa , Lavilla Susana , Miró Elisenda , González Juan José , Larrosa Nieves , Sabaté Montserrat , Navarro Ferran , Prats Guillermo
Published: July 2005

Two strains of
Enterobacteriaceae
,
Escherichia coli
and
Klebsiella pneumoniae
, producing VIM-1 were isolated for the first time in Spain. In both strains,
bla
VIM-1
was found to be carried on a gene cassette inserted into a class 1 integron. The
bla
VIM-1
-containing integron was located on a transferable plasmid.
In Vitro Synergistic Interaction between Amphotericin B and Micafungin against
Scedosporium
spp
Authors: Yustes Clara , Guarro Josep
Published: July 2005

The in vitro interaction between amphotericin B and micafungin against 36 isolates of
Scedosporium
spp. has been evaluated using checkerboard assays and the minimal effective concentration endpoint. Synergy was found for 82.4% of
Scedosporium prolificans
isolates and for 31.6% of
Scedosporium apiospermum
isolates. Antagonism was not observed.
Antimicrobial Activity and Spectrum of PPI-0903M (T-91825), a Novel Cephalosporin, Tested against a Worldwide Collection of Clinical Strains
Authors: Sader Helio S. , Fritsche Thomas R. , Kaniga Koné , Ge Yigong , Jones Ronald N.
Published: July 2005

PPI-0903M is a novel
N
-phosphono-type cephalosporin active against oxacillin-resistant staphylococci and many other gram-positive organisms. This study evaluated the in vitro activity and spectrum of PPI-0903M against 1,478 recent clinical isolates collected from 80 medical centers (22 countries). PPI-0903M demonstrated broader in vitro activity against gram-positive bacteria, particularly against multidrug-resistant staphylococci and streptococci of current clinical concern, than currently available extended-spectrum cephalosporins while maintaining similar activity against gram-negative pathogens.
Effects of Daptomycin, Linezolid, and Vancomycin on Establishment of Intestinal Colonization with Vancomycin-Resistant Enterococci and Extended-Spectrum-β-Lactamase-Producing
Klebsiella pneumoniae
in Mice
Authors: Pultz Nicole J. , Stiefel Usha , Donskey Curtis J.
Published: July 2005

In mice, vancomycin and linezolid treatment disrupted the anaerobic intestinal microflora, based on denaturing gradient gel electrophoresis analysis, and promoted colonization by
Klebsiella pneumoniae
and vancomycin-resistant enterococci. However, the effects varied depending on dose and duration of treatment. Daptomycin treatment did not disrupt the anaerobic microflora or promote either pathogen.
Molecular Characterization of the First Telithromycin-Resistant
Streptococcus pneumoniae
Isolate in Germany
Authors: Reinert Ralf René , van der Linden Mark , Al-Lahham Adnan
Published: July 2005

A total of 486
Streptococcus pneumoniae
isolates were collected in 2003 and 2004 in Germany and revealed the following resistance rates: penicillin G (7.2%) and erythromycin A (18.9%). Telithromycin exhibited good in vitro activity (MIC at which 90% of the isolates tested were inhibited, 0.125 μg/ml). However, one
erm
(B)-positive isolate was found to be telithromycin resistant (MIC, 8 μg/ml).