ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Oct. 2009, p. 4217–4224 Vol. 53, No. 10 0066-4804/09/$08.00ϩ0 doi:10.1128/AAC.00742-09 Copyright © 2009, American Society for Microbiology. All Rights Reserved. Activity of Telavancin against Staphylococci and Enterococci Determined by MIC and Resistance Selection Studiesᰔ Klaudia Kosowska-Shick, Catherine Clark, Glenn A. Pankuch, Pamela McGhee, Bonifacio Dewasse, Linda Beachel, and Peter C. Appelbaum* Department of Pathology, Hershey Medical Center, Hershey, Pennsylvania 17033 Received 3 June 2009/Returned for modification 30 June 2009/Accepted 3 July 2009 This study used CLSI broth microdilution to test the activity of telavancin and comparator antimicrobial agents against 67 methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) isolates. Twenty-six vanco- mycin-intermediate S. aureus (VISA) strains were among the isolates tested; all strains were susceptible to telavancin at <1 ␮g/ml, whereas 12/26 (46%) of these isolates were nonsusceptible to daptomycin at the same concentration. All strains were susceptible to quinupristin-dalfopristin, while resistance was found to all other drugs tested. Telavancin demonstrated potent activity against all vancomycin-susceptible isolates as well as Downloaded from against heterogeneously VISA and VISA resistance phenotypes. In multistep resistance selection studies, telavancin yielded one stable mutant after 43 days in one MRSA strain out of the 10 MRSA strains tested with the MIC rising eightfold from 0.25 ␮g/ml (parent) to 2 ␮g/ml. MICs for this clone did not increase further when passages were continued for the maximum 50 days. In contrast, daptomycin selected stable resistant clones (MIC increase of >4؋) after 14 to 35 days in 4 of 10 MRSA strains with MICs increasing from 1 to 2 ␮g/ml (parents) to 4 to 8 ␮g/ml (resistant clones). Sequencing analysis of daptomycin resistance determinants aac.asm.org revealed point mutations in the mprF genes of all four stable daptomycin-resistant clones. Teicoplanin gave rise to resistant clones after 14 to 21 days in 2 of 10 MRSA strains with MICs rising from 1 to 2 ␮g/ml (parents) to4to16␮g/ml (stable resistant clones). Linezolid selected stable resistant clones after 22 to 48 days in 2 of 10 MRSA strains with MICs rising from 2 to 4 ␮g/ml (parents) to 32 ␮g/ml (resistant clones). Vancomycin yielded no resistant clones in 10 MRSA strains tested; however, MICs increased two- to fourfold from 1 to 8 at IOWA STATE UNIV on December 15, 2009 ␮g/ml to 2 to 16 ␮g/ml after 50 days. No cross-resistance was found with any clone/antimicrobial combination. The two enterococci developed resistance to daptomycin, and one developed resistance to linezolid. Single-step mutation frequencies for telavancin (<4.0 ؋ 10؊11 to <2.9 ؋ 10؊10 at 2؋ MIC) were lower than the spontaneous mutation frequencies obtained with the comparators. Staphylococcus aureus is becoming increasingly resistant to described the utility of the Etest macromethod using a double- antibiotics. Methicillin (meticillin)-resistant S. aureus (MRSA) sided vancomycin-teicoplanin Etest strip which accurately dif- strains are increasingly encountered all over the world and ferentiates between hVISA and VISA strains. Utilization of cannot be treated with existing ß-lactams. Additionally, the the latter method will surely increase reports of the incidence majority of hospital-acquired methicillin-resistant (and also of hVISA and VISA strains. As of this time, the pathogenicity some methicillin-susceptible) strains are resistant to all cur- of VRSA strains awaits confirmation. rently available quinolones. The situation has become more The situation has become further complicated by the ap- complicated by the appearance of heterogeneously vancomycin- pearance and rapid spread, especially in the United States, of intermediate S. aureus (hVISA) strains, vancomycin-interme- community-acquired MRSA strains that are especially viru- diate S. aureus (VISA) strains, and recently nine reported lent, possibly by virtue of production of Panton-Valentine leu- vancomycin-resistant S. aureus (VRSA) strains (1). Two recent kocidin (4, 10, 16, 28, 34, 37). Although these strains are papers emphasize the recent spread of VISA strains in Turkey currently more susceptible to antimicrobial agents than hospi- (47) and France (15), and a recent alert from the New York tal-acquired strains are, this situation will surely change. Ad- City Department of Health (11) has documented six cases of ditionally, treatment of the community-acquired MRSA strains VISA infections in New York City, NY, between February and with glycopeptides will increase the selective pressure leading October 2007, which led to four fatalities. It seems clear that to nonsusceptibility to vancomycin and teicoplanin. Recently, VISA phenotypes occur everywhere but that they are not being we and others have documented clinical development of dap- routinely detected due to lack of standardized methodology (1, tomycin resistance after daptomycin therapy (25), and not all 20). Recently, Rybak et al. (43) have indicated, with Etest VISA strains are daptomycin susceptible (1, 19, 25, 26). There macromethod and population analysis testing, that the inci- is an urgent need for new agents to treat MRSA infections. dence of hVISA strains has increased over the past 22 years to Telavancin is an investigational lipoglycopeptide active an overall incidence of 2.2%. Yusof et al. (55) have recently (MICs of Յ1 ␮g/ml) against gram-positive organisms including MRSA (17, 18, 22, 29, 31, 38, 48). Barcia-Macay and coworkers (3) have reported telavancin MICs of 0.5 ␮g/ml against two * Corresponding author. Mailing address: Department of Pathology, strains of VISA and of 2 to 4 ␮g/ml against two strains of Hershey Medical Center, P.O. Box 850, Hershey, PA 17033. Phone: (717) 531-5113. Fax: (717) 531-7953. E-mail: [email protected]. VRSA. Leuthner and coworkers in a study of 50 glycopeptide- ᰔ Published ahead of print on 20 July 2009. nonsusceptible staphylococci and 3 VRSA strains showed tela- 4217 4218 KOSOWSKA-SHICK ET AL. ANTIMICROB.AGENTS CHEMOTHER. TABLE 1. Microdilution MICs (␮g/ml) of 67 MRSA strains MICs for VSSA strains MICs for VISA strains ϭ MIC range for hVISA ϭ MIC range for VRSA Drug (n 33) (n 26) strains (n ϭ 2) strains (n ϭ 6) Range MIC50/MIC90 Range MIC50/MIC90 Telavancin 0.25–1 0.25/0.25 0.25–0.5 0.25–1 0.5/1 2–4 Vancomycin 0.5–1 1/1 2 2–8 4/8 Ͼ32 Teicoplanin 0.25–2 0.5/1 2–8 1–32 4/16 8–Ͼ32 Daptomycin 0.25–1 0.5/0.5 0.5–1 0.5–Ͼ11/Ͼ1 0.25–1 Linezolid 2–Ͼ4 4/4 2–4 Յ0.5–4 2/4 2–4 Telithromycin 0.12–Ͼ8 0.12/Ͼ8 Ͼ8 0.06–Ͼ8 Ͼ8/Ͼ8 Ͼ8 Oxacillin Ͼ4 Ͼ4/Ͼ4 Ͼ4 0.5–Ͼ4 Ͼ4/Ͼ4 Ͼ4 Quinipristin-dalfopristin 0.25–1 0.5/1 0.5 Յ0.12–2 0.5/1 0.5–1 Clindamycin Յ0.5–Ͼ4 Յ0.5/Ͼ4 Ͼ4 Յ0.5–Ͼ4 Ͼ4/Ͼ4 Ͼ4 Ciprofloxacin 0.25–Ͼ8 0.5/Ͼ8 Ͼ81–Ͼ8 Ͼ8/Ͼ8 Ͼ8 Erythromycin 2–Ͼ16 Ͼ16/Ͼ16 Ͼ16 0.25–Ͼ16 Ͼ16/Ͼ16 Ͼ16 Gentamicin 0.25–Ͼ16 0.5/1 0.25–0.5 0.25–Ͼ16 1/Ͼ16 0.5–Ͼ16 Trimethoprim-sulfamethoxazolea Յ0.5–Ͼ4 Յ0.5/Յ0.5 Յ0.5 Յ0.5–Ͼ4 Յ0.5/Ͼ4 Յ0.5–Ͼ4 a MICs reported based on trimethoprim concentration. Downloaded from vancin to be potent against all 37 hVISA and VISA strains (13 sputum samples from patients at Hershey Medical Center (25). The VISA strain coagulase-negative strains; resistance phenotypes not differen- isolated from a patient who developed the VISA and daptomycin nonsuscepti- tiated from one another) with MICs of Ͻ1 ␮g/ml, with higher bility phenotype from an MRSA strain while on therapy with the two respective agents has been reported in detail (25). ␮ aac.asm.org MICs of 2 to 4 g/ml against the 3 VRSA strains (33). The For single-step and multistep resistance selection studies, 10 MRSA strains as antibacterial mode of action of telavancin results from inhibi- well as two enterococci (all recent Hershey isolates) were tested. The MRSA tion of bacterial cell wall synthesis and interference with the strains comprised two hVISA strains, four VISA strains (including one strain that barrier function of the bacterial cell membrane (22). The mode developed the VISA and daptomycin resistance phenotype after sequential ther- apy with both compounds) and four vancomycin-susceptible strains. Addition- of action of daptomycin also involves targeting the bacterial at IOWA STATE UNIV on December 15, 2009 ally, one vancomycin-susceptible Enterococcus faecalis strain and one vancomy- cell membrane to initiate antimicrobial activity (2, 24, 26). This cin-susceptible Enterococcus faecium strain were tested. fact is supported by the involvement of staphylococcal genes MIC determinations. Both broth microdilution and broth macrodilution meth- regulating cell membrane surface charge (e.g., mprF) (39) and ods were used for MIC testing. Broth microdilution was conducted by the fatty acid synthesis (e.g., yycG) (36) in the development of Clinical and Laboratory Standards Institute (CLSI) method using commercially prepared trays (TREK, Inc., Cleveland, OH) (13), with added calcium for dap- daptomycin nonsusceptibility. tomycin and 24-h incubation for vancomycin. Broth macrodilution was also In an effort to expand the comparative activity of telavancin conducted according to CLSI methodology (13). Telavancin was obtained from against MRSA strains of various resistance phenotypes, we Theravance, Inc. (South San Francisco, CA), and the other antimicrobial agents have investigated the activity of telavancin against MRSA were obtained from their respective manufacturers. strains by determining the activities of telavancin and compar- Multistep resistance selection. Multistep resistance selection was performed as previously described using the broth macrodilution method (1, 6, 7, 13).