JAC Advance Access originally published online on July 2, 2007
Journal of Antimicrobial Chemotherapy 2007 60(3):568-574; doi:10.1093/jac/dkm236
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Antistaphylococcal activities of CG400549, a new bacterial enoyl-acyl carrier protein reductase (FabI) inhibitor
1 School of Life and Food Sciences, Handong Global University, Pohang 791-708, South Korea 2 CrystalGenomics, Inc., 388-1 Pungnap-dong, Songpa-gu, Seoul 138-736, South Korea
* Corresponding author. Tel: +82-54-260-1353; Fax: +82-54-260-1925; E-mail: jhkwak{at}handong.edu
Received 6 March 2007; returned 16 April 2007; revised 15 May 2007; accepted 5 June 2007
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Abstract |
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Objectives: This study was performed to analyse in vitro and in vivo activities of CG400549, a new FabI inhibitor, against clinical isolates of staphylococci. The mode of action of CG400549 and resistance mechanism of Staphylococcus aureus against CG400549 were also investigated by genetic approaches.
Methods: In vitro activity of CG400549 was evaluated by the 2-fold agar sdilution method as described by the CLSI, and compared with those of oxacillin, erythromycin, ciprofloxacin, sparfloxacin, moxifloxacin, gemifloxacin, vancomycin, linezolid and quinupristin-dalfopristin. In vivo activity of CG400549 was determined against systemic infections in mice. Time–kill curves of CG400549 were analysed at concentrations of 1 x , 2 x and 4 x MIC against S. aureus strains.
Results: CG400549 had the lowest MICs among the test compounds against 238 clinical isolates of S. aureus (MIC90, 0.25 mg/L) and 51 clinical isolates of coagulase-negative staphylococci (MIC90, 1 mg/L). The activity of CG400549 was irrespective of whether the strains were methicillin-susceptible or -resistant. Furthermore, CG400549 was effective by oral or subcutaneous administration against systemic infections in mice. In a time–kill study, CG400549 at concentrations of 1 x MIC, 2 x MIC and 4 x MIC had a bacteriostatic activity during 24 h. A FabI-overexpressing S. aureus strain gave rise to an increase in the MIC of CG400549 compared with the parental strain, while the susceptibilities of the FabI-overexpressing S. aureus strain to the other antibacterial agents such as oxacillin, erythromycin and ciprofloxacin were not affected. This result showed that the mode of action of CG400549 was via inhibition of FabI, which is involved in biosynthesis of fatty acids in bacteria. Study of the resistance mechanism of S. aureus showed that CG400549-resistant mutants of S. aureus had an alteration in FabI at Phe-204 to Leu.
Conclusions: CG400549 had potent in vitro and in vivo activity against staphylococci, including methicillin-, ciprofloxacin- and multidrug-resistant staphylococci strains. This compound could be a good candidate for clinical development as a novel anti-MRSA drug.
Keywords: antibacterial activity , mode of action , resistance mechanism
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Introduction |
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The emergence of multidrug-resistant Gram-positive pathogens, such as methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant coagulase-negative staphylococci (MRCoNS), penicillin-resistant Streptococcus pneumoniae and vancomycin-resistant enterococci, has generated worldwide concern in the medical community.1,2
S. aureus is an important pathogenic organism causing a variety of local and systemic infections. It has developed resistance to most classes of antimicrobial agents soon after their introduction into clinical use. MRSA has been a serious problem in hospitals only, but recently, an increasing incidence of community-acquired MRSA is becoming an important public health problem.3 Although vancomycin provides effective therapy against most strains of MRSA, the first clinical isolate of S. aureus with decreased susceptibility to vancomycin was isolated in Japan in 1997, and other staphylococcal strains with reduced susceptibility to vancomycin have been identified in several countries since then.4–7
There is an urgent need to develop new classes of antibiotics to tackle the increase of resistance in Gram-positive bacteria.8 The recent trend to develop new antibiotics is to identify and exploit new molecular targets of pathogenic strains.9,10 Many novel validated targets have been identified from bacterial genome information. Among the new antibacterial targets, several enzymes involved in bacterial fatty acid biosynthesis have been receiving increased attention as attractive targets for the development of novel antibacterial agents.
Bacterial fatty acid biosynthesis is an essential process that supplies precursors for the assembly of important cellular components, including phospholipids, lipoproteins, lipopolysaccharides, mycolic acids and the cell envelope. In the type I system of mammals including humans, fatty acid synthase is a single, large polypeptide composed of several distinct domains. In the type II system of bacteria, the fatty acid synthase components including the acyl carrier protein (ACP) exist as discrete proteins. This difference in organization makes the bacterial fatty acid biosynthetic enzymes potentially selective antibacterial targets.11 The last step in the fatty acid biosynthetic pathway is performed by enoyl-ACP reductase (FabI), which is responsible for reduction of the double bond in the enoyl-ACP derivative. In addition, FabI is an important enzyme because reduction of enoyl-ACP derivatives is thought to regulate the ratio of saturated to unsaturated fatty acids and to coordinate fatty acid and phospholipid syntheses, and it plays a determinant role in completing cycles of fatty acid elongation.12,13 In S. aureus, this enzyme has been shown to be the antibacterial target of triclosan, thereby demonstrating the essentiality of FabI in this organism.14,15
CG400549 (Figure 1), a novel FabI inhibitor, was synthesized by the CrystalGenomics, Inc. (Seoul, Republic of Korea). In this study, we compared in vitro activities of CG400549 with those of nine other antibiotics against a variety of clinical isolates of staphylococci. We examined in vivo efficacy of CG400549 against systemic infection in mice and performed a time–kill study of CG400549 against methicillin-susceptible S. aureus (MSSA) and MRSA. We also characterized the mode of action of CG400549 by using a FabI-overexpressing S. aureus strain, and the resistance mechanism of S. aureus against CG400549 by genetic analysis of resistant mutants.
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Materials and methods |
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Antimicrobial agents
The test compounds were obtained as follows. CG400549 was synthesized at CrystalGenomics. Linezolid and quinupristin/dalfopristin were extracted from commercial tablets and purified by HPLC at CrystalGenomics. Ciprofloxacin, moxifloxacin, sparfloxacin and gemifloxacin were obtained from the R&D Center, Dong Wha Pharmaceutical Company, Anyang, Republic of Korea. Oxacillin, kanamycin, ampicillin, vancomycin and erythromycin were purchased from Sigma Chemical Co., St Louis, MO, USA.
Test organisms used in this study were originally isolated from human clinical specimens. These were obtained from several hospitals in Seoul, Republic of Korea, between 2001 and 2005. One hundred and sixty-nine MRSA strains, 69 MSSA strains, 36 MRCoNS strains and 15 methicillin-susceptible coagulase-negative staphylococci (MSCoNS) strains were used for this study. The challenge organisms used in systemic infections in mice were as follows: S. aureus giorgio was kindly provided by LG life sciences Ltd, Daejeon, Republic of Korea,16 and S. aureus P197 (MRSA), S. aureus B1588 (MRSA) and quinolone-resistant S. aureus P128 (QRSA) were selected through the screening of clinical isolates.17
The MICs were determined by the 2-fold agar dilution method as described by the CLSI (formerly NCCLS).18 Test strains were grown for 18 h at 37°C in tryptic soy broth and diluted with the same fresh medium to a density of ~107 cfu/mL. These were applied to Mueller–Hinton agar (MHA) plates containing serial dilutions of antimicrobial agents using a multi-pin inoculator to yield 105 cfu/spot. Plates were incubated in air at 37°C for 18 h and were examined for growth. MIC was considered to be the lowest concentration that completely inhibited growth on agar plates, disregarding a single colony or a faint haze caused by the inoculum.
In vivo activity of compounds was determined against systemic infections in mice. The experimental protocol was approved by the Ethics Review Committee for Animal Experimentation of Handong Global University School of Life and Food Sciences. The design of this study was in accordance with the institutional guidelines for animal experiments and the care and use of laboratory animals. Four-week-old male IcrTacSam:ICR mice weighing 18–22 g were purchased from Samtaco Co., Ltd, Osan, Republic of Korea. They were maintained in animal rooms at 23 x 2°C with 55 x 20% relative humidity. Test organisms for infection were cultured on tryptic soy agar (TSA) at 37°C for 18 h. For use as inoculum, all bacterial strains were suspended in 0.9% saline solution containing 5% gastric mucin (Difco). Mice were used in groups of six each and were challenged intraperitoneally with a single 0.5 mL portion of the bacterial suspension corresponding to an inoculum range of 10–100 times the MLD of bacteria. CG400549 at four dose levels was orally or subcutaneously administered to mice four times at 1, 4, 10 and 24 h post-infection. Mortality was recorded for 7 days and the median effective dose needed to protect 50% of the mice (ED50) was calculated by the Probit method.19 The challenge inoculum was sufficient to kill 100% of the untreated control mice, which died within 48 h post-infection.
The time–kill analyses were performed by the method of the CLSI M26-A.20 Test organisms incubated on TSA for 18 h at 37°C were diluted with fresh Mueller–Hinton broth to ~105 cfu/mL, and the diluted cultures were pre-incubated for 2 h. Each drug was added to the cultures at concentrations of 0.25 x , 0.5 x , 1 x , 2 x and 4 x MIC. Aliquots (0.1 mL) of the cultures were removed at 0, 2, 4, 6 and 24 h of incubation and serial 10-fold dilutions were prepared in saline as needed. The numbers of viable cells were determined on drug-free MHA plates after 24 h of incubation. The compound was considered bactericidal at the concentration that reduced the original inoculum by 3 log10 cfu/mL (99.9%) at each of the time periods, or considered bacteriostatic if the inoculum was reduced by 0–3 log10 cfu/mL.20
The wild-type fabI gene from S. aureus RN4220 was amplified by PCR and cloned into vector pE194 yielding recombinant plasmid pE194-fabI.21 pE194 and recombinant pE194-fabI were introduced into electrocompetent S. aureus RN4220 via electroporation.22 Expression levels of FabI in the wild-type and FabI-overexpressing strains were examined by western immunoblotting. By using a methodology described previously, a FabI-overexpressing S. aureus strain was used to confirm that the mode of antibacterial action of CG400549 was via inhibition of FabI.15
Frequency of spontaneous single-step mutation
Approximately 109–1010 cfu of S. aureus RN4220 and S. aureus giorgio were plated on brain heart infusion (BHI) agar containing CG400549 (2 x MIC or 4 x MIC), respectively. The plates were incubated aerobically at 37°C for 48 h. The resistant colonies were subcultured on BHI agar plates containing the same concentration of CG400549. To ensure the reproducibility and reliability of single-step mutation, MIC test was performed for all resistant mutants. The resistance frequency at each MIC was calculated as the number of resistant colonies per inoculum.
Resistance mechanism of S. aureus to CG400549
To characterize any changes in the fabI gene of resistant mutants, a PCR method was used to amplify the fabI gene using the following two primers: fabI(seq-f), 5'-GTCATCATGGGAATCGCTAAT-3'; and fabI(seq-r), 5'-CGTGGAATCCGCTATCTACAT-3'. Each PCR fragment was purified using the AccuPrep PCR product purification kit (Bioneer Co., Ltd, Daejeon, Republic of Korea) before being used for DNA sequence determinations by Solgent Co., Ltd (Daejeon, Republic of Korea). Sequencing traces were analysed using the Lasergene software program.
To study whether resistance in these mutants was due to an efflux mechanism, the MICs for each resistant mutant were determined by agar dilution in the presence of reserpine (10 mg/L), a known efflux pump inhibitor.
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In vitro antibacterial activities
The MICs of test compounds for clinical isolates of S. aureus are shown in Table 1. On the basis of their susceptibilities to ciprofloxacin, 98% of MSSA isolates were quinolone-susceptible and almost all ( > 99%) MRSA isolates were quinolone-resistant (Table 1). The MIC90s of CG400549 for MSSA and MRSA (MICs at which 90% of the isolates tested were inhibited) were 0.25 mg/L. Among the test compounds, CG400549 had the lowest MICs for staphylococci, followed by quinupristin/dalfopristin, vancomycin, linezolid, gemifloxacin, moxifloxacin, sparfloxacin, ciprofloxacin, oxacillin and erythromycin. CG400549 was four to eight times more active than vancomycin and linezolid. The MICs of test compounds for clinical isolates of CoNS are shown in Table 2. Against MRCoNS and MSCoNS, CG400549 was as active as quinupristin/dalfopristin. However, it was 2-fold more active than vancomycin and linezolid. In general, the MICs of CG400549 were slightly higher (but still in the susceptible range) for coagulase-negative strains than for S. aureus strains.
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In vivo activity against systemic infections
The protective efficacy of CG400549 against systemic infections in mice is presented in Table 3. Against infection caused by S. aureus giorgio (MSSA), the ED50 of CG400549 was 4.38 mg/kg of body weight when it was administered by subcutaneous route. CG400549 was also active when it was administered orally (ED50, 18.85 mg/kg). CG400549 had similar in vivo efficacy against systemic infections caused by antibiotic-resistant strains such as S. aureus P197 (MRSA), S. aureus B1588 (methicillin-resistant but quinolone-susceptible) and S. aureus P128 (methicillin-resistant and quinolone-resistant). The ranges of ED50s were 5.12–10.36 (oral route) and 25.83–34.45 mg/kg (subcutaneous route). CG400549 was more active when it was administered by subcutaneous route.
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Time–kill studies
The killing kinetics of CG400549 against S. aureus strains are presented in Figure 2. The MICs of CG400549 against S. aureus giorgio, S. aureus P197, S. aureus B1588 and S. aureus P128 were 0.25 mg/L. The results of the time–kill assays showed that CG400549, at concentrations of 1 x MIC, 2 x MIC and 4 x MIC, had a bacteriostatic activity against all strains during 24 h.
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Mode of action
An increase in the MIC for the strain overexpressing FabI relative to that for the wild-type is indicative of FabI being the mode of antibacterial action. To test this hypothesis, the MICs of CG400549, triclosan (FabI inhibitor which is known to inhibit FabI and exhibit antibacterial activity against S. aureus) and ten other antimicrobial compounds with different mechanisms of action (such as DNA synthesis, protein synthesis and cell wall synthesis) were determined against S. aureus RN4220 (wild-type), S. aureus RN4220 (pE194) and S. aureus RN4220 (pE194-fabI). As shown in Table 4, the overexpression of FabI in S. aureus RN4220 (pE194-fabI) gave rise to an increase in the MIC of CG400549 from 0.25 to 16 mg/L. Triclosan showed the same pattern as CG400549 in susceptibility. In contrast, the MICs of nine other compounds (except erythromycin) not targeting FabI were not changed in the FabI-overexpressing strain. S. aureus RN4220 (pE194) was highly resistant to erythromycin because pE194 plasmid had an erythromycin resistance gene [erm(C)] as a selection marker. On the other hand, S. aureus RN4220 (pE194) did not show increased resistance to CG400549.
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Resistance mechanism of S. aureus to CG400549
The frequencies of single-step mutants at concentration of 2 x or 4 x MIC of CG400549 are summarized in Table 5. The frequency at which spontaneous CG400549-resistant mutants arose in S. aureus was 1.18 x 10–9–8.75 x 10–10. Thirteen S. aureus mutants resistant to CG400549 were selected to characterize any changes in the fabI gene of the resistant mutants (Table 6). Ten strains had a mutation in FabI; phenylalanine at position 204 was replaced by leucine. These mutant strains showed a 16-fold increase in the MIC of CG400549 compared with their corresponding parent strain. Three resistant mutants (ER5M-2, ER5M-7 and ER5M-12) had no mutations in the fabI gene, and these mutant strains showed an 8-fold increase in their MICs. Treatment with reserpine, the efflux pump inhibitor, did not affect the MICs of CG400549 (Table 6).
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Discussion |
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Staphylococci are important nosocomial pathogens. They are able to adapt to the hospital environment by becoming resistant to many antibiotics, including methicillin, quinolones and vancomycin. Recently, linezolid, quinupristin/dalfopristin, daptomycin and tigecycline have been approved as new anti-MRSA agents.23 However, not long after linezolid and quinupristin/dalfopristin were used clinically, bacteria with resistance to both agents was reported.24
FabI is an attractive target for antibiotic therapy, because it is an essential enzyme that is ubiquitous in pathogenic bacteria and as it has no mammalian counterpart. Recently, a number of inhibitors of FabI have been described to have antibacterial activity.25–28 However, analysis of key bacterial genomes demonstrated that FabI is absent in some organisms, and an alternative enoyl-ACP reductase, FabK, is present in several important clinical pathogens, including S. pneumoniae, Enterococcus faecalis and Pseudomonas aeruginosa.29 Consequently, FabI represents a selective antibacterial target for those pathogens such as S. aureus wherein FabI is the sole enoyl-ACP reductase.
CG400549 was identified as a highly selective FabI inhibitor. This compound had a potent in vitro antibacterial activity against staphylococci and it was four to eight times more active than vancomycin and linezolid. CG400549 showed a good protective efficacy against systemic infections caused by S. aureus in mice when it was administered by oral or subcutaneous route. Especially, CG400549 was equally active against MSSA and MRSA, suggesting the possibility of using this compound for treating infections caused by staphylococci. But CG400549 had no activity against streptococci, enterococci and Gram-negative bacteria (data not shown).
Previous studies showed that triclosan resistance in S. aureus can occur as a result of overexpression of FabI or amino acid changes in FabI.30 And clinical studies with S. aureus have shown that mutations in FabI (G23S, Y147H and F204C) and their overexpression cause decreases in susceptibility to triclosan.31,32 Interestingly, the predicted FabI amino acid change (F204L), which occurred in CG400549-resistant mutants, is different from the previously reported FabI mutations of G23S, Y147H and F204C. In addition, nucleotide changes in the fabI gene were not found in three CG400549-resistant mutants (ER5M-2, ER5M-7 and ER5M-12). The lack of fabI mutations in these mutants suggested that genetic loci other than fabI may also be involved in the reduction in susceptibility to CG400549. CG400549-resistant mutants showed cross-resistance to triclosan in our study (data not shown). Therefore, there is a possibility of occurrence of CG400549-resistant mutants due to the increasing use of triclosan, although CG400549 is very active against clinical isolates of S. aureus including MRSA.
In conclusion, CG400549 was identified as a highly selective FabI inhibitor. This compound was highly active in vitro and in vivo against staphylococci. CG400549, at concentrations of 1 x , 2 x and 4 x MIC, showed bacteriostatic activity against S. aureus during 24 h. CG400549 could be a good candidate for clinical development as an anti-MRSA drug.
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This work was supported by a Korea Research Foundation grant (KRF-2004-042-E00163).
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None to declare.
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References |
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1 Bell JM, Turnidge JD. High prevalence of oxacillin-resistant Staphylococcus aureus isolates from hospitalized patients in Asia-Pacific and South Africa: results from SENTRY Antimicrobial Surveillance Program, 1998–1999. Antimicrob Agents Chemother (2002) 46:879–81.
2 Levy SB, Marshall B. Antibacterial resistance worldwide: causes, challenges and responses. Nat Med (2004) 10:s122–129.[CrossRef][Web of Science][Medline]
3 Zetola N, Francis JS, Nuermberger EL, et al. Community-acquired methicillin-resistant Staphylococcus aureus: an emerging threat. Lancet Infect Dis (2005) 5:275–86.[CrossRef][Web of Science][Medline]
4
Hiramatsu K, Hanaki H, Ino T, et al. Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother (1997) 40:135–6.
5 Cosgrove SE, Carroll KC, Perl TM. Staphylococcus aureus with reduced susceptibility to vancomycin. Clin Infect Dis (2004) 39:539–45.[CrossRef][Web of Science][Medline]
6
Linda MW, Clewell DB, Gill SR, et al. Genetic analysis of a high-level vancomycin-resistant isolate of Staphylococcus aureus. In: Science (2003) 302:1569–71.
7
Song JH, Hiramatsu K, Suh JY, et al. Emergence in Asian countries of Staphylococcus aureus with reduced susceptibility to vancomycin. Antimicrob Agents Chemother (2004) 48:4926–8.
8 Bax R, Mullan N, Verhoef J. The millennium bugs: the need for and development of new antibacterials. Int J Antimicrob Agents (2000) 16:51–9.[CrossRef][Web of Science][Medline]
9 Lynn M, Jonathan G, Todd AB. Genetic strategies for antibacterial drug discovery. Nature (2003) 4:442–56.
10 Molly BS. Seeing is believing: the impact of structural genomics on antimicrobial drug discovery. Nat Rev Microbiol (2004) 2:739–46.[CrossRef][Web of Science][Medline]
11 Heath RJ, White SW, Rock CO. Lipid biosynthesis as a target for antibacterial agents. Prog Lipid Res (2001) 40:467–97.[CrossRef][Web of Science][Medline]
12
Heath RJ, Rock CO. Enoyl-acyl carrier protein reductase (fabI) plays a determinant role in completing cycles of fatty acid elongation in Escherichia coli. J Biol Chem (1995) 270:26538–42.
13
Heath RJ, Rock CO. Regulation of fatty acid elongation and initiation by acyl-acyl carrier protein in Escherichia coli. J Biol Chem (1996) 271:1833–6.
14 McMurry LM, Oethinger M, Levy SB. Triclosan targets lipid synthesis. Nature (1998) 394:531–2.[CrossRef][Medline]
15
Slater-Radosti C, Van-Aller G, Greenwood R, et al. Biochemical and genetic characterization of the action of triclosan on Staphylococcus aureus. J Antimicrob Chemother (2001) 48:1–6.
16 Oh JI, Paek KS, Ahn MJ, et al. In vitro and in vivo evaluations of LB20304 a new fluoronahthyridone. Antimicrob Agents Chemother (1996) 40:1564–8.[Abstract]
17
Park HS, Kim HJ, Seol MJ, et al. In vitro and In vivo antibacterial activities of DW-224a, a new fluoronaphthyridone. Antimicrob Agents Chemother (2006) 50:2261–4.
18 Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Fifteenth Informational Supplement M100-S15 (2005) Wayne, PA, USA: CLSI.
19 Bliss CI. Statistics in Bioassay (1985) New York: Academic Press, Inc.
20 National Committee for Clinical Laboratory Standards. Methods for Determining Bactericidal Activity of Antimicrobial Agents: Approved Guideline M26-A (1999) Wayne, PA, USA: NCCLS.
21
Kwak JH, Weisblum B. Regulation of plasmid pE194 replication: control of cop-repF operon transcription by Cop and of repF translation by countertranscript RNA. J Bacteriol (1994) 176:5044–51.
22 Kraemer GR, Iandolo JJ. High frequency transformation of Staphylococcus aureus by electroporation. Curr Microb (1990) 21:373–6.[CrossRef]
23 Spellberg B, Powers JH, Brass EP, et al. Trends in antimicrobial drug development: implications for the future. Clin Infect Dis (2004) 38:1279–86.[CrossRef][Web of Science][Medline]
24
Luh KT, Hsueh PR, Teng LJ, et al. Quinupristin-dalfopristin resistance among Gram-positive bacteria in Taiwan. Antimicrob Agents Chemother (2000) 44:3374–80.
25
Losee LL, Xian J, Ali S, et al. Identification and characterization of inhibitors of bacterial enoyl-acyl carrier protein reductase. Antimicrob Agents Chemother (2004) 48:1541–7.
26 Moir DT. Identification of inhibitors of bacterial enoyl-acyl carrier protein reductase. Curr Drug Targets Infect Disord (2005) 5:297–305.[CrossRef][Medline]
27
Payne DJ, Miller WH, Berry W, et al. Discovery of a novel and potent class of FabI-directed antibacterial agents. Antimicrob Agents Chemother (2002) 46:3118–24.
28 Seefeld MA, Miller WH, Newlander KA, et al. Indole naphthyridinones as inhibitors of bacterial enoyl-ACP reductases FabI and FabK. J Med Chem (2003) 46:1627–35.[CrossRef][Web of Science][Medline]
29 Marrakchi H, Dewolf WE, Quinn C, et al. Characterization of Streptococcus pneumoniae enoyl-(acyl-carrier protein) reductase (FabK). Biochem J (2003) 370:1055–62.[CrossRef][Web of Science][Medline]
30
Heath RJ, Yu UT, Shapiro MA, et al. Inhibition of the Staphylococcus aureus NADPH-dependent enoyl-acyl carrier protein reductase by triclosan and hexachlorophene. J Biol Chem (2000) 275:4654–9.
31
Frank F, Yan K, Wallis NG, et al. Defining and combating the mechanisms of triclosan resistance in clinical isolates of Staphylococcus aureus. Antimicrob Agents Chemother (2002) 46:3343–7.
32 Brenwald NP, Fraise AP. Triclosan resistance in methicillin-resistant Staphylococcus aureus. J Hosp Infect (2003) 55:141–4.[CrossRef][Web of Science][Medline]
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