Activity of Linezolid Tested Against Uncommonly Isolated Gram-positive ACCEPTED
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1 AAC Accepts, published online ahead of print on 8 January 2007 Antimicrob. Agents Chemother. doi: /aac Copyright 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved Activity of Linezolid Tested Against Uncommonly Isolated Gram-positive Organisms (3,251 strains): Report from the SENTRY Antimicrobial Surveillance Program Short running title: Note Ronald N. Jones 1,2 *, Matthew G. Stilwell 1, Patricia A. Hogan 3, and Daniel J. Sheehan 3 1 JMI Laboratories, North Liberty, IA, USA; 2 Tufts University School of Medicine, Boston, MA, USA; and 3 Pfizer Inc, New York, NY, USA *Corresponding Author: R.N. Jones, M.D. JMI Laboratories 345 Beaver Kreek Centre, Suite A North Liberty Iowa Phone: (319) Fax: (319) ronald-jones@jmilabs.com 1
2 Abstract Linezolid was tested against 32 species of uncommonly isolated gram-positive organisms (3,251 strains) by reference MIC methods and found to be highly active (MIC 50 range, µg/ml; and MIC 90 range, µg/ml). Only one isolate (viridans group streptococcus) was resistant (0.03% of tested strains) to linezolid. Keywords: Linezolid, gram-positive organisms, oxazolidinone activity, resistance, SENTRY Program 2
3 Linezolid, the only oxazolidinone used in clinical practice, has demonstrated potent activity against gram-positive pathogens including resistant variants such as methicillin-resistant Staphylococcus aureus or coagulase-negative staphylococcal (CoNS) species, vancomycin-resistant enterococci (VRE) and β-haemolytic streptococci (8, 14, 15). The oxazolidinone action against these organisms has been described as bacteriostatic and pre-marketing surveillance trials show near complete coverage (>99.9% susceptibility) against the indicated pathogens (2-5, 12, 15). Subsequent post-marketing resistance monitoring studies (ZAAPS, LEADER, SENTRY Antimicrobial Surveillance Program) have discovered linezolid-resistant isolates among staphylococci, enterococci and some viridans group streptococci species (1, 9, 13, 16). However, the rates of occurrence have remained far less than 1% but occasionally more frequent among some species (CoNS, E. faecium), as well as associated with long-term therapy ( 6 weeks) or suspected compromise of infection control practices (17, 19). Regulatory agencies and the Clinical and Laboratory Standards Institute (CLSI) have established indications for linezolid use and breakpoint interpretive criteria of 2 µg/ml as susceptible for streptococci or enterococci and 4 µg/ml for staphylococci (7). Resistance has only been defined at 8 µg/ml for enterococci (7). These criteria are very helpful for clinical laboratories in guiding therapy for serious Gram-positive infections; however, numerous other non-indicated gram-positive pathogens could require linezolid as a treatment option due to potential intolerance of other agents or frank resistance. To address this therapeutic possibility, we expand the knowledge of linezolid activity and spectrum by studying the compound against 32 gram-positive species that are uncommonly isolated from contemporary clinical infections. The organisms were obtained from the SENTRY Antimicrobial Surveillance Program ( ) from infections cultured in North America, Latin America and Europe (10). All strains were processed in a central, reference laboratory study design using CLSI M7-A7 broth microdilution methods and interpretive criteria (6, 7, 11). The cation-adjusted Mueller-Hinton broth was supplemented with 2-5% lysed horse blood when testing fastidious species (6, 7). To interpret the susceptibility of the rarer species, established breakpoints for organisms of the same genus (e.g. streptococci or enterococci) were applied for comparison purposes only. All organisms with linezolid MIC results of 8 µg/ml were tested by PCR techniques for rrna target mutations commonly associated with oxazolidinone resistances (14, 16). Also, staphylococci and enterococci with linezolid MIC results of 4 µg/ml were repeated to establish the reproducible MIC values that occurred near published breakpoints (7). All tests had concurrent quality control (QC) performed with CLSI recommended strains (6, 7). All QC results were within established MIC ranges (7). 3
4 A total of 3,251 organisms were analyzed as follows: Aerococcus spp. (22), Bacillus spp. (202; two species groups), Corynebacterium spp. (342; five species), enterococci (378; six species), Listeria monocytogenes (137), Micrococcus luteus (29), Rothia mucilaginosus (18), β-haemolytic streptococci not group A or B (865; three serogroups) and viridans group streptococci (1,258; 12 species); see Tables 1 and 2. All identifications were initially made at the participant sites and confirmed at the central laboratory by conventional reference methods and commercial systems (Vitek and Vitek 2, biomerieux, Hazelwood, MO). Table 1 provides the linezolid MIC distributions, MIC 50 and MIC 90 results from testing 1,128 isolates of Gram-positive cocci. Among these species, the Corynebacterium spp. (unspeciated, amycolatum, jeikeium, pseudiphtheriticum, striatum) were most susceptible to linezolid with a MIC 50 of only µg/ml. M. luteus and R. mucilaginosus strains were also very susceptible to linezolid (MIC 50, 0.5 µg/ml),followed by Aerococcus spp. and the two groupings of Bacillus spp. (MIC 50, 1 µg/ml). Enterococci (not E. faecalis or E. faecium) were clearly less susceptible to linezolid (MIC 50 and MIC 90, 2 µg/ml), but all enterococcal MIC results were 4 µg/ml, i.e. not resistant by CLSI criteria (7). No linezolid- resistant isolates were detected among the species summarized in Table 1. The reference linezolid MIC results for all streptococcal species of uncommon occurrence in documented clinical infections have been listed in Table 2. A very uniform pattern of linezolid susceptibility was noted with MIC 50 results ranging from 0.5 to 1 µg/ml and MIC 90 values from 1 to 2 µg/ml. All species or serogroups demonstrated complete (100%) susceptibility (MIC, 2 µg/ml) except for a single strain of S. oralis previously reported from this program (10, 16). This organism had a documented G2576T rrna mutation by PCR sequencing analysis (14, 18). These in vitro linezolid results document the wide potential clinical application to uncommonly isolated gram-positive species, and the infrequency of linezolid-resistant strains among indicated species (1-5, 9, 12, 13, 15, 16) illustrates the continued clinical utility of the oxazolidinone class. However, when therapy with linezolid would be considered, susceptibility tests should be performed by validated in vitro methods applying published interpretive criteria, if available (6, 7, 11). Results of such tests should be similar to those listed here for those rare species, and where linezolid MIC values of 8 µg/ml occur the results should be confirmed by a reference laboratory using standardized methods (6, 7). Sahm et al. (Abstr. 105 th General Meeting American Society for Microbiology, abstr C-320, 2005) recently noted discords between MIC results derived from the E-test (AB BIODISK, Solna, Sweden) and the reference broth microdilution method. Generally, the linezolid E-test MIC values presented in this study were reliable for S. aureus testing but were consistently higher for enterococcal tests (D.F. Sahm, D.C. Draghi, 4
5 R.S. Blosser, P.A. Hogan and D.J. Sheehan, Abstr. 105 th General Meeting American Society for Microbiology, abstr C-320, 2005). Appropriate interpretations of linezolid MIC endpoints requires the exclusions of trailing effects with the reference MICs and the application of an 80% inhibition endpoint to the hazy borders often seem with agar diffusion methods (disk diffusion and E-test) (6, 7) (S. Poppe, R. Schaadt, D. Sheehan, D. Sahm, G. Zurenko and D. Shinabarger, Abstr. 106 th General Meeting American Society for Microbiology, abstr. A-088, 2006). These technical details must be considered along with acceptable concurrent QC results (7) to avoid the reporting of false linezolid resistance. 5
6 112 References Anderegg, T. R., H. S. Sader, T. R. Fritsche, J. E. Ross, and R. N. Jones Trends in linezolid susceptibility patterns: Report from the worldwide Zyvox Annual Appraisal of Potency and Spectrum (ZAAPS) Program. Int J Antimicrob Agents 26: Ballow, C. H., D. J. Biedenbach, F. Rossi, and R. N. Jones Multicenter assessment of the linezolid spectrum and activity using the disk diffusion and E-test methods: report of the Zyvox(R) antimicrobial potency study in Latin America (LA-ZAPS). Braz J Infect Dis 6: Ballow, C. H., R. N. Jones, and D. J. Biedenbach A multicenter evaluation of linezolid antimicrobial activity in North America. Diagn Microbiol Infect Dis 43: Bell, J. M., J. D. Turnidge, C. H. Ballow, and R. N. Jones Multicentre evaluation of the in vitro activity of linezolid in the Western Pacific. J Antimicrob Chemother 51: Bolmstrom, A., C. H. Ballow, A. Qwarnstrom, D. J. Biedenbach, and R. N. Jones Multicentre assessment of linezolid antimicrobial activity and spectrum in Europe: Report from the Zyvox antimicrobial potency study (ZAPS-Europe). Clin Microbiol Infect 8: Clinical and Laboratory Standards Institute M7-A7, Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard - seventh edition. Wayne, PA: CLSI. 7. Clinical and Laboratory Standards Institute M100-S16, Performance standards for antimicrobial susceptibility testing; sixteenth informational supplement. Wayne, PA: CLSI Diekema, D. J., and R. N. Jones Oxazolidinone antibiotics. Lancet 358: Draghi, D. C., D. J. Sheehan, P. Hogan, and D. F. Sahm In vitro activity of linezolid against key gram-positive organisms isolated in the United States: Results of the LEADER 2004 surveillance program. Antimicrob Agents Chemother 49:
7 Jones, R. N Global epidemiology of antimicrobial resistance among community-acquired and nosocomial pathogens: A five-year summary from the SENTRY Antimicrobial Surveillance Program ( ). Semin Respir Crit Care Med 24: Jones, R. N., T. R. Anderegg, and D. J. Biedenbach Validation of commercial dry-form broth microdilution panels for susceptibility testing of AZD2563, a new long-acting oxazolidinone. Clin Microbiol Infect 9: Jones, R. N., C. H. Ballow, and D. J. Biedenbach Multi-laboratory assessment of the linezolid spectrum of activity using the Kirby-Bauer disk diffusion method: Report of the Zyvox Antimicrobial Potency Study (ZAPS) in the United States. Diagn Microbiol Infect Dis 40: Jones, R. N., J. E. Ross, T. R. Fritsche, and H. S. Sader Oxazolidinone susceptibility patterns in 2004: Report from the Zyvox Annual Appraisal of Potency and Spectrum (ZAAPS) Program assessing isolates from 16 nations. J Antimicrob Chemother 57: Meka, V. G., and H. S. Gold Antimicrobial resistance to linezolid. Clin Infect Dis 39: Mutnick, A. H., D. J. Biedenbach, J. D. Turnidge, and R. N. Jones Spectrum and potency evaluation of a new oxazolidinone, linezolid: Report from the SENTRY Antimicrobial Surveillance Program, Diagn Microbiol Infect Dis 43: Mutnick, A. H., V. Enne, and R. N. Jones Linezolid resistance since 2001: SENTRY Antimicrobial Surveillance Program. Ann Pharmacother 37: Potoski, B. A., J. Adams, L. Clarke, K. Shutt, P. K. Linden, C. Baxter, A. W. Pasculle, B. Capitano, A. Y. Peleg, D. Szabo, and D. L. Paterson Epidemiological profile of linezolid- resistant coagulase-negative staphylococci. Clin Infect Dis 43: Prystowsky, J., F. Siddiqui, J. Chosay, D. L. Shinabarger, J. Millichap, L. R. Peterson, and G. A. Noskin Resistance to linezolid: Characterization of mutations in rrna and 7
8 comparison of their occurrences in vancomycin-resistant enterococci. Antimicrob Agents Chemother 45: Rahim, S., S. K. Pillai, H. S. Gold, L. Venkataraman, K. Inglima, and R. A. Press Linezolid-resistant, vancomycin-resistant Enterococcus faecium infection in patients without prior exposure to linezolid. Clin Infect Dis 36:E
9 Table 1. Linezolid potency and spectrum when tested against 1,128 isolates uncommonly isolated of Gram-positive organisms. Cumulative % inhibited at MIC (µg/ml): MIC (µg/ml): Organism (no. tested) % 90% Aerococcus spp. (22) Bacillus spp. (142) B. cereus (60) Corynebacterium spp. (236) C. amycolatum (11) C. jeikium (59) C. pseudodiphtheriticum (11) C. striatum (25) Enterococcus avium (116) E. casseliflavus (65) E. durans (49) E. gallinarum (119) E. hirae (16) E. raffinosus (13) Listeria monocytogenes (137) Micrococcus luteus (29) Rothia mucilaginosus (18)
10 Table 2. Potency and spectrum of linezolid when tested against uncommonly isolated species/serogroups of β-haemolytic (865 strains) and viridans group streptococci (1,258 strains). Cumulative % inhibited at MIC (µg/ml): MIC (µg/ml): Organism (no. tested) % 90% β-haemolytic streptococci Group C (199) Group F (53) Group G (613) Viridans group streptococci - S. anginosus (104) S. bovis (223) a S. constellatus (89) S. gordonii (12) S. intermedius (66) S. mitis (341) S. mutans (19) S. oralis (139) b 1 1 S. parasanguis (21) S. salivarius (91) S. sanguis (142) S. vesicularis (11) a. Also known as S. gallolyticus. b. One documented strain resistant by a G2576T mutation. 10
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