bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Single Oral Doses of Gemifloxacin (320 Milligrams) versus Trovafloxacin (200 Milligrams) on Fecal Flora in Healthy Volunteers
Authors: Garcia-Calvo Gloria , Molleja Ana , Giménez Maria J. , Parra Araceli , Nieto Eva , Ponte Carmen , Aguilar Lorenzo , Soriano Francisco
Published: July 2002

Gemifloxacin and trovafloxacin were administered to 12 volunteers in a randomized crossover trial with a 2-week washout period. Stool samples were collected predose and 1, 2, and 3 days postdose. Both quinolones reduced the number of organisms of the family
Enterobacteriaceae
and aerobic gram-positive organisms.
Escherichia coli
reduction was greater with gemifloxacin than with trovafloxacin, with postdose isolation of quinolone-resistant strains for which MICs of trovafloxacin were higher than those of gemifloxacin.
Pharmacokinetics of a New Parenteral Oligosaccharide Antibiotic, SCH27899 (Ziracin), in Healthy Subjects
Authors: Kozawa Osamu , Uematsu Toshihiko , Matsuno Hiroyuki , Niwa Masayuki , Kohno Ken-Ichi , Kawato Asakazu , Takahashi Kazuyoshi , Nagashima Satoru , Kanamaru Mitsutaka
Published: July 2002

The pharmacokinetic properties of an everninomicin antibiotic (SCH27899; Ziracin) were studied with healthy Japanese male volunteers by single (1, 3, 6, and 9 mg/kg of body weight) and multiple 60-min intravenous infusions (3, 6, and 9 mg/kg once daily for 10 consecutive days following a 2-day interval after the initial dose). At single doses the peak serum concentration and the area under the serum concentration-time curve linearly increased with the dose. While total body clearance (CL; 31.2 to 45.6 ml/kg/h) and percent cumulative urinary recovery as unchanged drug (4.9 to 7.1%) were rather constant irrespective of doses, the terminal half-life of γ phase (
t
1/2γ
; 14.2 to 19.6 h) were slightly prolonged at the higher two doses compared with the lower two doses. With repeated doses of SCH27899, a statistically significant decrease and increase were found in CL and
t
1/2γ
of about 36 and 21%, respectively, although these changes may be clinically irrelevant. The most commonly reported adverse events were local reactions such as erythema, pain, and palpable venous cord of mild to moderate degree around the injection site, which could be managed by changing the injection sites.
Efficacies of Gel Formulations Containing Foscarnet, Alone or Combined with Sodium Lauryl Sulfate, against Establishment and Reactivation of Latent Herpes Simplex Virus Type 1
Authors: Piret Jocelyne , Lamontagne Julie , Désormeaux André , Bergeron Michel G.
Published: July 2002

The influence of sodium lauryl sulfate (SLS) on the efficacies of gel formulations of foscarnet against herpes simplex virus type 1 (HSV-1) cutaneous lesions and on the establishment and reactivation of latent virus has been evaluated in a murine model of orofacial infection. Topical treatments were given twice daily for 3 days and were initiated at 6, 24, and 48 h after virus inoculation. The gel formulation that contained both 3% foscarnet and 5% SLS and that was administered within 48 h postinfection reduced the rate of development of herpetic skin lesions. This formulation also significantly decreased the viral content in skin tissues and in ipsilateral trigeminal ganglia when it was given within 24 and 6 h postinfection, respectively. A lower level of efficacy was observed for the gel formulation containing 3% foscarnet alone. Of prime interest, the gel formulation containing 5% SLS reduced significantly the mortality rate among mice in a zosteriform model of infection. Both formulations of foscarnet had no effect on the mean titers of reactivated virus in explant cultures of ipsilateral and contralateral trigeminal ganglia from latently infected mice. The use of a gel formulation containing combinations of foscarnet and SLS could represent an attractive approach for the treatment of herpetic mucocutaneous infections.
Specific Inhibition of Coxsackievirus B3 Translation and Replication by Phosphorothioate Antisense Oligodeoxynucleotides
Authors: Wang Aikun , Cheung Paul K. M. , Zhang Huifang , Carthy Christopher M. , Bohunek Lubos , Wilson Janet E. , McManus Bruce M. , Yang Decheng
Published: July 2002

The 5′ and 3′ untranslated regions (UTRs) of coxsackievirus B3 (CVB3) RNA form highly ordered secondary structures that have been confirmed to play important regulatory roles in viral cap-independent internal translation initiation and RNA replication. We previously demonstrated that deletions in different regions of the 5′ UTR significantly reduced viral RNA translation and infectivity. Such observations suggested strongly that viral RNA translation and replication could be blocked if highly specific antisense oligodeoxynucleotides (AS-ODNs) were applied to target crucial sites within the 5′ and 3′ UTRs. In this study, seven phosphorothioate AS-ODNs were synthesized, and the antiviral activity was evaluated by Lipofectin transfection of HeLa cells with AS-ODNs followed by infection of CVB3. Analysis by Western blotting, reverse transcription-PCR, and viral plaque assay demonstrated that viral protein synthesis, genome replication, and infectivity of CVB3 were strongly inhibited by the AS-ODNs complementary to different regions of the 5′ and 3′ UTRs. The most effective sites are located at the proximate terminus of the 5′ UTR (AS-1), the proximate terminus of the 3′ UTR (AS-7), the core sequence of the internal ribosome entry site (AS-2), and the translation initiation codon region (AS-4). These AS-ODNs showed highly sequence-specific and dose-dependent inhibitory effects on both viral protein synthesis and RNA replication. It is noteworthy that the highest inhibitory activities were obtained with AS-1 and AS-7 targeting the termini of the 5′ and 3′ UTRs. The percent inhibition values of AS-1 and AS-7 for CVB3 protein VP1 synthesis and RNA replication were 70.6 and 79.6 for AS-1 and 73.7 and 79.7 for AS-7, respectively. These data suggest that CVB3 infectivity can be inhibited effectively by AS-ODNs.
In Vitro Activities of Six Quinolones and Mechanisms of Resistance in
Staphylococcus aureus
and Coagulase-Negative Staphylococci
Authors: Linde Hans-Jörg , Schmidt Mario , Fuchs Emmi , Reischl Udo , Niller Hans-Helmut , Lehn Norbert
Published: July 2002

Of 94 clinical isolates of
Staphylococcus aureus
(
n
= 51) and coagulase-negative staphylococci (CNS) (
n
= 43), mutations in the quinolone resistance-determining region of topoisomerases GrlA, GrlB, GyrA, and GyrB together with MICs of six quinolones were analyzed. Amino acid substitutions at identical residues (GrlA residues 80 and 84; GyrA residues 84 and 88) were found in
S. aureus
and CNS. Active efflux, as suggested by blocking by reserpine, contributed substantially to the resistance phenotype in some strains. Among ciprofloxacin, clinafloxacin, levofloxacin, nalidixic acid, trovafloxacin, and sparfloxacin, a 0.5-μg/ml concentration of sparfloxacin discriminated best between strains with two or three mutations and those with no mutations.
Synergistic Interactions between Mammalian Antimicrobial Defense Peptides
Authors: Yan Hong , Hancock Robert E. W.
Published: July 2002

A single animal can express several cationic antimicrobial peptides with different sequences and structures. We demonstrate that mammalian peptides from different structural classes frequently show synergy with each other and selectively show synergy with human lysozyme.
Macrolide Resistance in
Streptococcus pneumoniae
in Hong Kong
Authors: Ip Margaret , Lyon Donald J. , Yung Raymond W. H. , Chan Colin , Cheng Augustine F. B.
Published: July 2002

Erythromycin resistance rates among penicillin-susceptible
Streptococcus pneumoniae
were 38 and 92% among penicillin-intermediate and -resistant
S. pneumoniae
isolates from Hong Kong, respectively, and 27% (43 of 158) of the isolates showed the MLS
B
phenotype, and the majority carried the
ermB
gene; 73% (115 of 158) displayed the M phenotype, and all possessed the
mef
gene. The MLS
B
phenotype was predominant in penicillin-susceptible, macrolide-resistant isolates and in penicillin-nonsusceptible isolates of serotype 6B, whilst the M phenotype was predominant in penicillin-intermediate or -resistant isolates belonging to serotype 23F or 19F. Extensive spread of clones of drug-resistant pneumococci has led to the widespread presence of macrolide resistance in
S. pneumoniae
in Hong Kong.
Mechanism of Synergy between Epigallocatechin Gallate and β-Lactams against Methicillin-Resistant
Staphylococcus aureus
Authors: Zhao Wei-Hua , Hu Zhi-Qing , Okubo Sachie , Hara Yukihiko , Shimamura Tadakatsu
Published: July 2002

Compared to MICs (more than 800 μg/ml) of (−)-epigallocatechin gallate (EGCg) against
Escherchia coli
, MICs of EGCg against methicillin-susceptible and methicillin-resistant
Staphylococcus aureus
(MSSA and MRSA) were 100 μg/ml or less. Furthermore, less than 25 μg EGCg per ml obviously reversed the high level resistance of MRSA to all types of tested β-lactams, including benzylpenicillin, oxacillin, methicillin, ampicillin, and cephalexin. EGCg also induced a supersusceptibility to β-lactams in MSSA which does not express
mecA
, encoding penicillin-binding protein 2′ (PBP2′). The fractional inhibitory concentration (FIC) indices of the tested β-lactams against 25 isolates of MRSA were from 0.126 to 0.625 in combination with 6.25, 12.5 or 25 μg of EGCg per ml. However, no synergism was observed between EGCg and ampicillin against
E. coli
. EGCg largely reduced the tolerance of MRSA and MSSA to high ionic strength and low osmotic pressure in their external atmosphere, indicating damage of the cell wall. Unlike dextran and lipopolysaccharide, peptidoglycan from
S. aureus
blocked both the antibacterial activity of EGCg and the synergism between EGCg and oxacillin, suggesting a direct binding of EGCg with peptidoglycan on the cell wall. EGCg showed a synergistic effect with
dl
-cycloserine (an inhibitor of cell wall synthesis unrelated to PBP2′) but additive or indifferent effect with inhibitors of protein and nuclear acid synthesis. EGCg did not suppress either PBP2′ mRNA expression or PBP2′ production, as confirmed by reverse transcription-PCR and a semiquantitative PBP2′ latex agglutination assay, indicating an irrelevance between the synergy and PBP2′ production. In summary, both EGCg and β-lactams directly or indirectly attack the same site, peptidoglycan on the cell wall. EGCg synergizes the activity of β-lactams against MRSA owing to interference with the integrity of the cell wall through direct binding to peptidoglycan.
Durability of Anti-Infective Effect of Long-Term Silicone Sheath Catheters Impregnated with Antimicrobial Agents
Authors: Tcholakian Robert K. , Raad Issam I.
Published: July 2002

This study was performed to test the long-term antimicrobial efficacy of impregnated silicone catheters comprising an antimicrobial layer sandwiched between an external surface sheath and a luminal surface silicone sheath. The design of the catheter permits the introduction of various antimicrobials in addition to anticoagulants or antifibrins in the antimicrobial layer and allows their gradual release over a period of months after insertion. The in vitro data presented show that the catheter can provide antimicrobial activity for 90 days, after being replated for 15 7-day cycles of replating. When the catheters were immersed in human serum and incubated at 37°C, they demonstrated significant antimicrobial activity after more than 325 days of incubation. The significant long-term in vitro antimicrobial activity observed may imply effective in vivo activity for almost 1 year after insertion and could serve as a cost-effective alternative to surgically implantable silicone catheters.
Alterations in MurM, a Cell Wall Muropeptide Branching Enzyme, Increase High-Level Penicillin and Cephalosporin Resistance in
Streptococcus pneumoniae
Authors: Smith Anthony M. , Klugman Keith P.
Published: July 2002

We report that alteration in MurM, an enzyme involved in the biosynthesis of branched-stem cell wall muropeptides, is required for maximal expression of penicillin and cefotaxime resistance in the pneumococcus. Hungarian isolate 3191 (penicillin MIC, 16 μg/ml; cefotaxime MIC, 4 μg/ml) was a source of donor DNA in transformation experiments. Penicillin-binding protein DNA was insufficient to transform recipient strain R6 to full resistance. Further transformation with altered
murM
DNA was required for full expression of donor penicillin and cefotaxime resistance.
Aminoglycoside Resistance with Homogeneous and Heterogeneous Populations of Antibiotic-Resistant Ribosomes
Authors: Recht Michael I. , Puglisi Joseph D.
Published: July 2002

Aminoglycosides bind to rRNA in the small subunit of the bacterial ribosome. Mutations in the decoding region of 16S rRNA confer resistance to specific subsets of aminoglycoside antibiotics. The two major classes of 2-deoxystreptamine aminoglycosides are the 4,5- and the 4,6-disubstituted antibiotics. Antibiotics of the 4,5-disubstituted class include neomycin, paromomycin, and ribostamycin. Gentamicins and kanamycins belong to the 4,6-disubstituted class of aminoglycosides. Structural studies indicated the potential importance of position 1406 (
Escherichia coli
numbering) in the binding of ring III of the 4,6-disubstituted class of aminoglycosides to 16S rRNA. We have introduced a U1406-to-A mutation in a plasmid-encoded copy of
E. coli
16S rRNA which has been expressed either in a mixture with wild-type ribosomes or in a strain in which all rRNA is transcribed from the plasmid-encoded
rrn
operon. High-level resistance to many of the 4,6-disubstituted aminoglycosides is observed only when all the rRNA contains the U1406-to-A mutation. In contrast to the partial dominance of resistance observed with other mutations in the decoding region, there is a dominance of sensitivity with the 1406A mutation. Chemical footprinting experiments indicate that resistance arises from a reduced affinity of the antibiotic for the rRNA target. These results demonstrate that although position 1406 is an important determinant in the binding and action of the 4,6-disubstituted aminoglycosides, other rRNA mutations that perturb the binding of ring I of both classes of 2-deoxystreptamine aminoglycosides confer higher levels of resistance as well as a partial dominance of resistance.
Antipneumococcal Activity of BMS 284756 Compared to Those of Six Other Agents
Authors: Pankuch Glenn A. , Nagai Kensuke , Davies Todd A. , Jacobs Michael R. , Appelbaum Peter C.
Published: July 2002

Antipneumococcal activity of BMS 284756 was compared to those of six agents by MIC and time-kill methodologies. BMS 284756 had the lowest MICs compared to those of ciprofloxacin, levofloxacin, and moxifloxacin against quinolone-susceptible (≤0.016 to 0.06 μg/ml) and quinolone-resistant (0.03 to 1 μg/ml) pneumococci. BMS 284756 was bactericidal against 11 of 12 strains at two times the MIC after 24 h.
Efficacy of Ceftriaxone and Gentamicin Given Once a Day by Using Human-Like Pharmacokinetics in Treatment of Experimental Staphylococcal Endocarditis
Authors: Gavaldà Joan , López Pedro , Martín Teresa , Gomis Xavier , Ramírez José Luis , Azuaje Carlos , Almirante Benito , Pahissa Albert
Published: July 2002

We compared the efficacy of ceftriaxone combined with gentamicin, both given once a day, with that of cloxacillin given every 4 h plus gentamicin given once a day or in three daily doses (t.i.d.) for the treatment of experimental methicillin-susceptible staphylococcal endocarditis. The antibiotics were administered by using human-like (H-L) pharmacokinetics that simulated the profiles of these drugs in human serum. Animals with catheter-induced endocarditis were infected intravenously with 10
5
CFU of
Staphylococcus aureus
S5 (MICs and minimal bactericidal concentrations of cloxacillin, ceftriaxone, and gentamicin, 0.5 and 2 μg/ml, 4 and 8 μg/ml, and 0.5 and 1 μg/ml, respectively). The animals were then treated for 24 h with cloxacillin at a dose of 2 g that simulated H-L pharmacokinetics (H-L 2 g) every 4 h alone or combined with gentamicin (administered at doses of H-L 1 mg/kg of body weight every 8 h or H-L 4.5 mg/kg every 24 h) or with ceftriaxone at H-L 2 g every 24 h alone or combined with gentamicin (administered at doses of H-L 1 mg/kg every 8 h or H-L 4.5 mg/kg every 24 h). The results of therapy for experimental endocarditis due to the S5 strain showed that (i) cloxacillin alone is more effective than ceftriaxone alone in reducing the bacterial load (
P
< 0.01), (ii) the combination of cloxacillin or ceftriaxone with gentamicin is more effective than each of these drugs alone (
P
< 0.01), and (iii) Ceftriaxone H-L plus gentamicin H-L 4.5 mg/kg, both administered every 24 h, showed efficacy similar to that of the “gold standard,” cloxacillin H-L plus gentamicin H-L 1 mg/kg t.i.d. (
P
> 0.05). An increase in the interval of administration of gentamicin to once daily resulted in a reduction in the numbers of bacteria in the vegetations equivalent to that achieved with the recommended regimen of cloxacillin plus gentamicin t.i.d. in the treatment of experimental endocarditis due to methicillin-susceptible
S. aureus
. Ceftriaxone plus gentamicin, both administered once a day, may be useful for home-based therapy for selected cases of staphylococcal endocarditis.
Expression of
lux
Genes in a Clinical Isolate of
Streptococcus pneumoniae
: Using Bioluminescence To Monitor Gemifloxacin Activity
Authors: Beard S. J. , Salisbury V. , Lewis R. J. , Sharpe J. A. , MacGowan A. P.
Published: July 2002

A clinical isolate of
Streptococcus pneumoniae
was transformed with a plasmid containing the
lux
operon of
Photorhabdus luminescens
that had been modified to function in gram-positive bacteria. Cells containing this plasmid produced light stably and constitutively, without compromising the growth rate. Light output was correlated with measurements of optical density and viable counts during exponential growth and provided a sensitive, real-time measure of the pharmacodynamics of the fluoroquinolone gemifloxacin.
Susceptibility to Telithromycin in 1,011
Streptococcus pyogenes
Isolates from 10 Central and Eastern European Countries
Authors: Nagai Kensuke , Appelbaum Peter C. , Davies Todd A. , Kelly Linda M. , Hoellman Dianne B. , Andrasevic Arjana Tambic , Drukalska Liga , Hryniewicz Waleria , Jacobs Michael R. , Kolman Jana , Miciuleviciene Jolanta , Pana Marina , Setchanova Lena , Thege Marianne Konkoly , Hupkova Helena , Trupl Jan , Urbaskova Pavla
Published: July 2002

Among 1,011 recently isolated
Streptococcus pyogenes
isolates from 10 Central and Eastern European centers, the MICs at which 50% of isolates are inhibited (MIC
50
s) and the MIC
90
s were as follows: for telithromycin, 0.03 and 0.06 μg/ml, respectively; for erythromycin, azithromycin, and clarithromycin, 0.06 to 0.125 and 1 to 8 μg/ml, respectively; and for clindamycin, 0.125 and 0.125 μg/ml, respectively. Erythromycin resistance occurred in 12.3% of strains.
Erm
(A) [subclass
erm
(TR)] was most commonly encountered (60.5%), followed by
mef
(A) (23.4%) and
erm
(B) (14.5%). At <0.5 μg/ml, telithromycin was active against 98.5% of the strains tested.
Ribosomal Mutations in
Streptococcus pneumoniae
Clinical Isolates
Authors: Pihlajamäki Marja , Kataja Janne , Seppälä Helena , Elliot John , Leinonen Maija , Huovinen Pentti , Jalava Jari
Published: July 2002

Eleven clinical isolates of
Streptococcus pneumoniae
, isolated in Finland during 1996 to 2000, had an unusual macrolide resistance phenotype. They were resistant to macrolides and streptogramin B but susceptible, intermediate, or low-level resistant to lincosamides. No acquired macrolide resistance genes were detected from the strains. The isolates were found to have mutations in domain V of the 23S rRNA or ribosomal protein L4. Seven isolates had an A2059C mutation in two to four out of the four alleles encoding the 23S rRNA, two isolates had an A2059G mutation in two alleles, one isolate had a C2611G mutation in all four alleles, and one isolate had a
69
GTG
71
-to-
69
TPS
71
substitution in ribosomal protein L4.
Characterization of a New Extended-Spectrum β-Lactamase (TEM-87) Isolated in
Proteus mirabilis
during an Italian Survey
Authors: Perilli Mariagrazia , Segatore Bernardetta , Massis Maria Rosaria De , Franceschini Nicola , Bianchi Ciro , Rossolini Gian Maria , Amicosante Gianfranco
Published: July 2002

A new natural TEM derivative, named TEM-87, was identified in a
Proteus mirabilis
isolate from an Italian hospital. Compared to TEM-1, TEM-87 contains the following mutations: E104K, R164C, and M182T. Kinetic analysis of TEM-87 revealed extended-spectrum activity against oxyimino cephalosporins (preferentially ceftazidime) and aztreonam. Expression of
bla
TEM-87
in
Escherichia coli
decreased the host susceptibility to these drugs.
In Vitro Antifungal Susceptibilities of
Trichosporon
Species
Authors: Paphitou Niki I. , Ostrosky-Zeichner Luis , Paetznick Victor L. , Rodriguez Jose R. , Chen Enuo , Rex John H.
Published: July 2002

The in vitro activities of amphotericin B, itraconazole, fluconazole, voriconazole, posaconazole, and ravuconazole against 39 isolates of
Trichosporon
spp. were determined by the NCCLS M27-A microdilution method. The azoles tested appeared to be more potent than amphotericin B. Low minimal fungicidal concentration/MIC ratios were observed for voriconazole, posaconazole, and ravuconazole, suggesting fungicidal activity.
In Vitro and In Vivo Activities of Anti-Influenza Virus Compound T-705
Authors: Furuta Y. , Takahashi K. , Fukuda Y. , Kuno M. , Kamiyama T. , Kozaki K. , Nomura N. , Egawa H. , Minami S. , Watanabe Y. , Narita H. , Shiraki K.
Published: July 2002

T-705 (6-fluoro-3-hydroxy-2-pyrazinecarboxamide) has been found to have potent and selective inhibitory activity against influenza virus. In an in vitro plaque reduction assay, T-705 showed potent inhibitory activity against influenza A, B, and C viruses, with 50% inhibitory concentrations (IC
50
s) of 0.013 to 0.48 μg/ml, while it showed no cytotoxicity at concentrations up to 1,000 μg/ml in Madin-Darby canine kidney cells. The selectivity index for influenza virus was more than 2,000. It was also active against a neuraminidase inhibitor-resistant virus and some amantadine-resistant viruses. T-705 showed weak activity against non-influenza virus RNA viruses, with the IC
50
s being higher for non-influenza virus RNA viruses than for influenza virus, and it had no activity against DNA viruses. Orally administered T-705 at 100 mg/kg of body weight/day (four times a day) for 5 days significantly reduced the mean pulmonary virus yields and the rate of mortality in mice infected with influenza virus A/PR/8/34 (3 × 10
2
PFU). These results suggest that T-705 may be a compound that is useful and highly selective against influenza virus infections and that has a mode of action different from those of commercially available drugs, such as amantadine, rimantadine, and neuraminidase inhibitors.
Anti-Human Immunodeficiency Virus Interactions of SCH-C (SCH 351125), a CCR5 Antagonist, with Other Antiretroviral Agents In Vitro
Authors: Tremblay Cécile L. , Giguel Françoise , Kollmann Christopher , Guan Yongbiao , Chou Ting-Chao , Baroudy Bahige M. , Hirsch Martin S.
Published: July 2002

SCH-C (SCH 351125) is a small-molecule antagonist of the human immunodeficiency virus type 1(HIV-1) coreceptor CCR5. It has in vitro activity against R5 viruses with 50% inhibitory concentrations ranging from 1.0 to 30.9 nM. We have studied anti-HIV-1 interactions of SCH-C with other antiretroviral agents in vitro. Synergistic interactions were seen with nucleoside reverse transcriptase inhibitors (zidovudine and lamivudine), nonnucleoside reverse transcriptase inhibitors (efavirenz), and protease inhibitors (indinavir) at all inhibitory concentrations evaluated. We have also studied antiviral interactions between the HIV-1 fusion inhibitor T-20 and SCH-C against a panel of R5 HIV-1 isolates. We found synergistic interactions against all the viruses tested, some of which harbored resistance mutations to reverse transcriptase and protease inhibitors. Anti-HIV-1 synergy was also observed between SCH-C and another R5 virus inhibitor, aminooxypentane-RANTES. These findings suggest that SCH-C may be a useful anti-HIV drug in combination regimens and that a combination of chemokine coreceptor/fusion inhibitors may be useful in the treatment of multidrug-resistant viruses.
Activity of the Ketolide Telithromycin Is Refractory to Erm Monomethylation of Bacterial rRNA
Authors: Liu Mingfu , Douthwaite Stephen
Published: July 2002

Methylation of specific nucleotides in rRNA is one of the means by which bacteria achieve resistance to macrolides-lincosamides-streptogramin B (MLS
B
) and ketolide antibiotics. The degree of resistance is determined by how effectively the rRNA is methylated. We have implemented a bacterial system in which the rRNA methylations are defined, and in this study we investigate what effect Erm mono- and dimethylation of the rRNA has on the activity of representative MLS
B
and ketolide antibiotics. In the test system, >80% of the rRNA molecules are monomethylated by ErmN (TlrD) or dimethylated by ErmE. ErmE dimethylation confers high resistance to all the MLS
B
and ketolide drugs. ErmN monomethylation predictably confers high resistance to the lincosamides clindamycin and lincomycin, intermediate resistance to the macrolides clarithromycin and erythromycin, and low resistance to the streptogramin B pristinamycin IA. In contrast to the macrolides, monomethylation only mildly affects the antimicrobial activities of the ketolides HMR 3647 (telithromycin) and HMR 3004, and these drugs remain 16 to 250 times as potent as clarithromycin and erythromycin. These differences in the macrolide and ketolide activities could explain the recent reports of variation in the MICs of telithromycin for streptococcal strains that have constitutive
erm
MLS
B
resistance and are highly resistant to erythromycin.
Pharmacology of β-
l
-Thymidine and β-
l
-2′-Deoxycytidine in HepG2 Cells and Primary Human Hepatocytes: Relevance to Chemotherapeutic Efficacy against Hepatitis B Virus
Authors: Hernandez-Santiago B. , Placidi L. , Cretton-Scott E. , Faraj A. , Bridges E. G. , Bryant M. L. , Rodriguez-Orengo J. , Imbach J. L. , Gosselin G. , Pierra C. , Dukhan D. , Sommadossi J. P.
Published: July 2002

β-
l
-Thymidine (
l
-dT) and β-
l
-2′-deoxycytidine (
l
-dC) are potent and highly specific inhibitors of hepatitis B virus (HBV) replication both in vivo and in vitro (50% effective concentrations, 0.19 to 0.24 μM in 2.2.15 cells). The intracellular metabolisms of
l
-dT and
l
-dC were investigated in HepG2 cells and primary cultured human hepatocytes.
l
-dT and
l
-dC were extensively phosphorylated in both cell types, with the 5′-triphosphate derivative being the predominant metabolite. In HepG2 cells, the 5′-triphosphate levels were 27.7 ± 12.1 and 72.4 ± 1.8 pmol/10
6
cells for
l
-dT and
l
-dC, respectively. In primary human hepatocytes, the 5′-triphosphate levels were 16.5 ± 9.8 and 90.1 ± 36.4 pmol/10
6
cells for
l
-dT and
l
-dC, respectively. Furthermore, a choline derivative of
l
-dCDP was detected at concentrations of 15.8 ± 1.8 and 25.6 ± 0.1 pmol/10
6
cells in human hepatocytes and HepG2 cells, respectively. In HepG2 cells exposed to
l
-dC, the 5′-monophosphate and 5′-triphosphate derivatives of β-
l
-2′-deoxyuridine (
l
-dUMP and
l
-dUTP, respectively) were also observed, reaching intracellular concentrations of 6.7 ± 0.4 and 18.2 ± 1.0 pmol/10
6
cells, respectively. In human hepatocytes,
l
-dUMP and
l
-dUTP were detected at concentrations of 5.7 ± 2.4 and 43.5 ± 26.8 pmol/10
6
cells, respectively. It is likely that deamination of
l
-dCMP by deoxycytidylate deaminase leads to the formation of
l
-dUMP, as the parent compound,
l
-dC, was not a substrate for deoxycytidine deaminase. The intracellular half-lives of
l
-dTTP,
l
-dCTP, and
l
-dUTP were at least 15 h, with intracellular concentrations of each metabolite remaining above their respective 50% inhibitory concentrations for the woodchuck hepatitis virus DNA polymerase for as long as 24 h after removal of the drug from cell cultures. Exposure of HepG2 cells to
l
-dT in combination with
l
-dC led to concentrations of the activated metabolites similar to those achieved with either agent alone. These results suggest that the potent anti-HBV activities of
l
-dT and
l
-dC are associated with their extensive phosphorylation.
Antifungal Susceptibility ofCandidaBiofilms: Unique Efficacy of Amphotericin B Lipid Formulations and Echinocandins
Authors: Kuhn D. M. , George T. , Chandra J. , Mukherjee P. K. , Ghannoum M. A.
Published: July 2002

Biofilms, likely the predominant mode of device-related microbial infection, exhibit resistance to antimicrobial agents. Evidence suggests thatCandidabiofilms have dramatically reduced susceptibility to antifungal drugs. We examined antifungal susceptibilities ofCandida albicansandCandida parapsilosisbiofilms grown on a bioprosthetic model. In addition to conventional agents, we determined if new antifungal agents (triazoles, amphotericin B lipid formulations, and echinocandins) have activities againstCandidabiofilms. We also explored effects of preincubation ofC. albicanscells with subinhibitory concentrations (sub-MICs) of drugs to see if they could modify subsequent biofilm formation. Finally, we used confocal scanning laser microscopy (CSLM) to image planktonic- and biofilm-exposed blastospores to examine drug effects on cell structure.Candidabiofilms were formed on silicone elastomer and quantified by tetrazolium and dry weight (DW) assays. Susceptibility testing of fluconazole, nystatin, chlorhexidine, terbenafine, amphotericin B (AMB), and the triazoles voriconazole (VRC) and ravuconazole revealed resistance in allCandidaisolates examined when grown as biofilms, compared to planktonic forms. In contrast, lipid formulations of AMB (liposomal AMB and AMB lipid complex [ABLC]) and echinocandins (caspofungin [Casp] and micafungin) showed activity againstCandidabiofilms. Preincubation ofC. albicanscells with sub-MIC levels of antifungals decreased the ability of cells to subsequently form biofilm (measured by DW;P< 0.0005). CSLM analysis of planktonic and biofilm-associated blastospores showed treatment with VRC, Casp, and ABLC resulted in morphological alterations, which differed with each agent. In conclusion, our data show thatCandidabiofilms show unique susceptibilities to echinocandins and AMB lipid formulations.
Longitudinal Use of a Line Probe Assay for Human Immunodeficiency Virus Type 1 Protease Predicts Phenotypic Resistance and Clinical Progression in Patients Failing Highly Active Antiretroviral Therapy
Authors: Servais Jean , Lambert Christine , Plesséria Jean-Marc , Fontaine Elodie , Robert Isabelle , Arendt Vic , Staub Thérèse , Hemmer Robert , Schneider François , Schmit Jean-Claude
Published: July 2002

An observational study assessed the longitudinal use of a new line probe assay for the detection of protease mutations. Probe assays for detection of reverse transcriptase (Inno-LiPA HIV-1 RT; Innogenetics) and protease (prototype kit Inno-LiPA HIV Protease; Innogenetics) mutations gave results for 177 of 199 sequential samples collected over 2 years from 26 patients failing two nucleoside reverse transcriptase inhibitors and one protease inhibitor (first line: indinavir,
n
= 6; ritonavir,
n
= 10; and saquinavir,
n
= 10). Results were compared to recombinant virus protease inhibitor susceptibility data (
n
= 87) and to clinical and virological data. Combinations of protease mutations (M46I, G48V, I54V, V82A or -F, I84V, and L90M) predicted phenotypic resistance to the protease inhibitor and to nelfinavir. The sum of protease mutations was associated with virological and clinical outcomes from 6 and 3 months on, respectively. Moreover, a poorer clinical outcome was linked to the sum of reverse transcriptase mutations. In conclusion, despite the limited number of patients studied and the restricted number of codons investigated, probe assay-based genotyping correlates with phenotypic drug resistance and predicts new Centers for Disease Control and Prevention stage B and C clinical events and virological outcome. Line probe assays provide additional prognostic information and should be prospectively investigated for their potential for treatment monitoring.
Erythromycin Modulates Eosinophil Chemotactic Cytokine Production by Human Lung Fibroblasts in Vitro
Authors: Sato Etsuro , Nelson Dan K. , Koyama Sekiya , Hoyt Jeffrey C. , Robbins Richard A.
Published: July 2002

Recent studies suggest that erythromycin can suppress the production of some cytokines and may be an effective treatment for asthma. Eosinophil chemotactic cytokines have been suggested to contribute to the pathogenesis of asthma by the recruitment of eosinophils. We hypothesized that erythromycin modulates eosinophil chemotactic cytokine production. To test the hypothesis, we evaluated the potential of erythromycin to modulate the release of eosinophil chemoattractants from the human lung fibroblast cell line HFL-1. HFL-1 released eotaxin, granulocyte-macrophage colony-stimulating factor, and regulated and normal T-cell expressed and presumably secreted (RANTES) in response to interleukin-1β or tumor necrosis factor alpha. Erythromycin attenuated the release of these cytokines and eosinophil chemotactic activity by the HFL-1. The suppressive effect on eotaxin was the most marked of these cytokines. Erythromycin therapy also suppressed eotaxin mRNA significantly. These results suggest a mechanism that may account for the apparent beneficial action of macrolide antibiotics in the treatment of allergic airway disorders.
Rapid and Simple Determination of the
Escherichia coli
Phylogenetic Group
Authors: Clermont Olivier , Bonacorsi Stéphane , Bingen Edouard
Published: July 2002

Phylogenetic analysis has shown that
Escherichia coli
is composed of four main phylogenetic groups (A, B1, B2, and D) and that virulent extra-intestinal strains mainly belong to groups B2 and D. Actually, phylogenetic groups can be determined by multilocus enzyme electrophoresis or ribotyping, both of which are complex, time-consuming techniques. We describe a simple and rapid phylogenetic grouping technique based on triplex PCR. The method, which uses a combination of two genes (
chuA
and
yjaA
) and an anonymous DNA fragment, was tested with 230 strains and showed excellent correlation with reference methods.
Fate of Selenate and Selenite Metabolized by
Rhodobacter sphaeroides
Authors: Van Fleet-Stalder Verena , Chasteen Thomas G. , Pickering Ingrid J. , George Graham N. , Prince Roger C.
Published: July 2002

Cultures of a purple nonsulfur bacterium,
Rhodobacter sphaeroides
, amended with ∼1 or ∼100 ppm selenate or selenite, were grown phototrophically to stationary phase. Analyses of culture headspace, separated cells, and filtered culture supernatant were carried out using gas chromatography, X-ray absorption spectroscopy, and inductively coupled plasma spectroscopy-mass spectrometry, respectively. While selenium-amended cultures showed much higher amounts of SeO
3
2−
bioconversion than did analogous selenate experiments (94% uptake for SeO
3
2−
as compared to 9.6% for SeO
4
2−
-amended cultures from 100-ppm solutions), the chemical forms of selenium in the microbial cells were not very different except at exposure to high concentrations of selenite. Volatilization accounted for only a very small portion of the accumulated selenium; most was present in organic forms and the red elemental form.
Complete Nucleotide Sequence of Ubiquitous Plasmid pEA29 from
Erwinia amylovora
Strain Ea88: Gene Organization and Intraspecies Variation
Authors: McGhee Gayle C. , Jones Alan L.
Published: July 2002

The complete sequence of plasmid pEA29 from
Erwinia amylovora
strain Ea88 consists of 28,185 bp with a 50.2% G+C content. As deletions and insertions were detected in other derivatives of pEA29, its size actually varied from 27.6 to 34.9 kb. Thirteen open reading frames that encoded predicted proteins with similarities to known proteins from other bacteria were identified along with two open reading frames related to hypothetical proteins found in GenBank and six open reading frames with no similarities to existing GenBank entries. Predicted products of open reading frames with similarity to the thiamine biosynthetic genes
thiO
,
thiG
, and
thiF
; a
betT
gene coding for choline transport; an
msrA
gene for the enzyme methionine sulfoxide reductase; a putative methyl-accepting chemotaxis gene; an aldehyde dehydrogenase gene; an
hns
DNA binding gene; a LysR-type transcriptional regulator; and
parA
and
parB
partitioning genes were identified. A putative iteron-containing theta-type origin of replication with an AT-rich region and a gene for a RepA protein was identified.
Pst
I and
Kpn
I restriction patterns for pEA29 isolated from tree fruit strains of
E. amylovora
were homogenous and different from those for pEA29 isolated from
Rubus
(raspberry) strains. All
Rubus
derivatives of pEA29 contained a point mutation that eliminated a
Pst
I site and a 1,264-bp region that replaced 1,890 bp of sequence found in pEA29 from strain Ea88. This change eliminated a second
Pst
I site and increased the length of a
Kpn
I fragment. An insertion sequence, IS
Eam1
, was detected in one
Rubus
strain, and transposon Tn
5393
was detected in three apple strains in two separate locations on the plasmid. Plasmid-cured strains exhibited reduced virulence and modified colony morphology on minimal medium without thiamine, indicating that some of the genes in pEA29 play a role in the physiology or metabolism of
E. amylovora.
Bradyrhizobium
sp. Strains That Nodulate the Leguminous Tree
Acacia albida
Produce Fucosylated and Partially Sulfated Nod Factors
Authors: Ferro Myriam , Lorquin Jean , Ba Salif , Sanon Kadidia , Promé Jean-Claude , Boivin Catherine
Published: July 2002

We determined the structures of Nod factors produced by six different
Bradyrhizobium
sp. strains nodulating the legume tree
Acacia albida
(syn.
Faidherbia albida
). Compounds from all strains were found to be similar, i.e., O-carbamoylated and substituted by an often sulfated methyl fucose and different from compounds produced by
Rhizobium-Mesorhizobium-Sinorhizobium
strains nodulating other species of the Acaciae tribe.
Determination of
Escherichia coli
Contamination with Chromocult Coliform Agar Showed a High Level of Discrimination Efficiency for Differing Fecal Pollution Levels in Tropical Waters of Kampala, Uganda
Authors: Byamukama Dennis , Kansiime Frank , Mach Robert L. , Farnleitner Andreas H.
Published: July 2002

Escherichia coli
, total coliforms, fecal coliforms, and sulfite-reducing anaerobic spore formers from different polluted sites in a tropical environment were determined in order to test for their indication ability for fecal contamination. Quantification of
E. coli
contamination with Chromocult coliform agar proved to be efficient and feasible for determining fecal pollutions in the investigated area within 24 h. The other microbial parameters showed a lower ability to differentiate sites and cannot be recommended for monitoring fecal pollution in the studied tropical surface waters.
Identification of Motile
Aeromonas
Strains with the MicroScan WalkAway System in Conjunction with the Combo Negative Type 1S Panels
Authors: Vivas J. , Sáa A. I. , Tinajas A. , Barbeyto L. , Rodríguez L. A.
Published: July 2002

This study was performed to compare the MicroScan WalkAway automated identification system in conjunction with the new MicroScan Combo Negative type 1S panels with conventional biochemical methods for identifying 85 environmental, clinical, and reference strains of eight
Aeromonas
species.
Isolation of Adherent Polycyclic Aromatic Hydrocarbon (PAH)-Degrading Bacteria Using PAH-Sorbing Carriers
Authors: Bastiaens Leen , Springael Dirk , Wattiau Pierre , Harms Hauke , deWachter Rupert , Verachtert Hubert , Diels Ludo
Published: July 2002

Two different procedures were compared to isolate polycyclic aromatic hydrocarbon (PAH)-utilizing bacteria from PAH-contaminated soil and sludge samples, i.e., (i) shaken enrichment cultures in liquid mineral medium in which PAHs were supplied as crystals and (ii) a new method in which PAH degraders were enriched on and recovered from hydrophobic membranes containing sorbed PAHs. Both techniques were successful, but selected from the same source different bacterial strains able to grow on PAHs as the sole source of carbon and energy. The liquid enrichment mainly selected for
Sphingomonas
spp., whereas the membrane method exclusively led to the selection of
Mycobacterium
spp. Furthermore, in separate membrane enrichment set-ups with different membrane types, three repetitive extragenic palindromic PCR-related
Mycobacterium
strains were recovered. The new
Mycobacterium
isolates were strongly hydrophobic and displayed the capacity to adhere strongly to different surfaces. One strain,
Mycobacterium
sp. LB501T, displayed an unusual combination of high adhesion efficiency and an extremely high negative charge. This strain may represent a new bacterial species as suggested by 16S rRNA gene sequence analysis. These results indicate that the provision of hydrophobic sorbents containing sorbed PAHs in the enrichment procedure discriminated in favor of certain bacterial characteristics. The new isolation method is appropriate to select for adherent PAH-degrading bacteria, which might be useful to biodegrade sorbed PAHs in soils and sludge.
Thermostabilization of Proteins by Diglycerol Phosphate, a New Compatible Solute from the Hyperthermophile
Archaeoglobus fulgidus
Authors: Lamosa Pedro , Burke Anthony , Peist Ralf , Huber Robert , Liu Ming-Y. , Silva Gabriela , Rodrigues-Pousada Claudina , LeGall Jean , Maycock Christopher , Santos Helena
Published: July 2002

Diglycerol phosphate accumulates under salt stress in the archaeon
Archaeoglobus fulgidus
(L. O. Martins, R. Huber, H. Huber, K. O. Stetter, M. S. da Costa, and H. Santos, Appl. Environ. Microbiol. 63:896–902, 1997). This solute was purified after extraction from the cell biomass. In addition, the optically active and the optically inactive (racemic) forms of the compound were synthesized, and the ability of the solute to act as a protecting agent against heating was tested on several proteins derived from mesophilic or hyperthermophilic sources. Diglycerol phosphate exerted a considerable stabilizing effect against heat inactivation of rabbit muscle lactate dehydrogenase, baker's yeast alcohol dehydrogenase, and
Thermococcus litoralis
glutamate dehydrogenase. Highly homologous and structurally well-characterized rubredoxins from
Desulfovibrio gigas
,
Desulfovibrio desulfuricans
(ATCC 27774), and
Clostridium pasteurianum
were also examined for their thermal stabilities in the presence or absence of diglycerol phosphate, glycerol, and inorganic phosphate. These proteins showed different intrinsic thermostabilities, with half-lives in the range of 30 to 100 min. Diglycerol phosphate exerted a strong protecting effect, with approximately a fourfold increase in the half-lives for the loss of the visible spectra of
D. gigas
and
C. pasteurianum
rubredoxins. In contrast, the stability of
D. desulfuricans
rubredoxin was not affected. These different behaviors are discussed in the light of the known structural features of rubredoxins. The data show that diglycerol phosphate is a potentially useful protein stabilizer in biotechnological applications.
Mitotic Recombination and Genetic Changes inSaccharomyces cerevisiaeduring Wine Fermentation
Authors: Puig Sergi , Querol Amparo , Barrio Eladio , Pérez-Ortín JoséE.
Published: July 2002

Natural strains ofSaccharomyces cerevisiaeare prototrophic homothallic yeasts that sporulate poorly, are often heterozygous, and may be aneuploid. This genomic constitution may confer selective advantages in some environments. Different mechanisms of recombination, such as meiosis or mitotic rearrangement of chromosomes, have been proposed for wine strains. We studied the stability of theURA3locus of aURA3/ura3wine yeast in consecutive grape must fermentations.ura3/ura3homozygotes were detected at a rate of 1 × 10−5to 3 × 10−5per generation, and mitotic rearrangements for chromosomes VIII and XII appeared after 30 mitotic divisions. We used the karyotype as a meiotic marker and determined that sporulation was not involved in this process. Thus, we propose a hypothesis for the genome changes in wine yeasts during vinification. This putative mechanism involves mitotic recombination between homologous sequences and does not necessarily imply meiosis.
Multiplex PCR for the Detection of
Lactobacillus pontis
and Two Related Species in a Sourdough Fermentation
Authors: Müller Martin R. A. , Ehrmann Matthias A. , Vogel Rudi F.
Published: July 2002

A specific multiplex PCR assay based on the amplification of parts of the 16S rRNA molecule was designed. Primers derived from variable regions of the 16S rRNA provided a means of easily differentiating the species
Lactobacillus pontis
and
Lactobacillus panis
. They could be clearly discriminated from the phylogenetically related species
Lactobacillus vaginalis
,
Lactobacillus oris
, and
Lactobacillus reuteri
and from other lactobacilli commonly known to be present in sourdough. Other strains isolated together with
L. pontis
from an industrial sourdough fermentation could be clearly separated from these species by comparative sequence analysis and construction of a specific PCR primer. For a fast identification a DNA isolation protocol based on the ultrasonic lysis of cells from single colonies was developed. To demonstrate the potential of such techniques for tracking these organisms in a laboratory-scale fermentation, we combined the specific PCR assay with direct DNA extraction from the organisms in the sourdough without previous cultivation.
Characterization of a Salt-Tolerant Family 42 β-Galactosidase from a Psychrophilic Antarctic
Planococcus
Isolate
Authors: Sheridan Peter P. , Brenchley Jean E.
Published: July 2002

We isolated a gram-positive, halotolerant psychrophile from a hypersaline pond located on the McMurdo Ice Shelf in Antarctica. A phylogenetic analysis of the 16S rRNA gene sequence of this organism showed that it is a member of the genus
Planococcus
. This assignment is consistent with the morphology and physiological characteristics of the organism. A gene encoding a β-galactosidase in this isolate was cloned in an
Escherichia coli
host. Sequence analysis of this gene placed it in glycosidase family 42 most closely related to an enzyme from
Bacillus circulans
. Even though an increasing number of family 42 glycosidase sequences are appearing in databases, little information about the biochemical features of these enzymes is available. Therefore, we purified and characterized this enzyme. The purified enzyme did not appear to have any metal requirement, had an optimum pH of 6.5 and an optimum temperature of activity at 42°C, and was irreversibly inactivated within 10 min when it was incubated at 55°C. The enzyme had an apparent

K
m

of 4.9 μmol of
o
-nitrophenyl-β-
d
-galactopyranoside, and the
V
max
was 467 μmol of
o
-nitrophenol produced/min/mg of protein at 39°C. Of special interest was the finding that the enzyme remained active at high salt concentrations, which makes it a possible reporter enzyme for halotolerant and halophilic organisms.
Thiosulfate Disproportionation by
Desulfotomaculum thermobenzoicum
Authors: Jackson Bradley E. , McInerney Michael J.
Published: July 2002

Desulfotomaculum thermobenzoicum
, but not
Desulfotomaculum nigrificans
,
Desulfotomaculum ruminis
, or
Desulfosporosinus orientis
, grew by disproportionation of thiosulfate, forming stoichiometric amounts of sulfate and sulfide; sulfite was not disproportionated. The addition of acetate enhanced growth and thiosulfate disproportionation by
D. thermobenzoicum
compared to those observed with thiosulfate alone.