Effects of Storage on Vancomycin and Daptomycin MIC in susceptible. blood isolates of Methicillin-Resistant Staphylococcus aureus

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1 JCM Accepts, published online ahead of print on 1 August 2012 J. Clin. Microbiol. doi: /jcm Copyright 2012, American Society for Microbiology. All Rights Reserved. 1 2 Effects of Storage on Vancomycin and Daptomycin MIC in susceptible blood isolates of Methicillin-Resistant Staphylococcus aureus 3 Running title: Effects of Storage on Susceptibility of MRSA Isolates 4 Authors: Franziska Ludwig, 1 Becky Edwards, 1# Timothy Lawes, 2 Ian M Gould Institution: This work was performed at the Medical Microbiology Department, Aberdeen Royal Infirmary, Aberdeen, United Kingdom. 7 Author affiliations: 8 1 Medical Microbiology Department, Aberdeen Royal Infirmary, Aberdeen, United Kingdom. 9 2 Raigmore Hospital, Inverness, United Kingdom. 10 Correspondent footnote: Address correspondence to Becky Edwards, bex.edwards@gmail.com Present addresses: Becky Edwards, Microbiology Department, Southern General Hospital, 1345 Govan Road, Glasgow, G51 4TF Timothy Lawes, 1A Drum Court, Raigmore Hospital, Inverness, United Kingdom, IV2 3UJ By repeating Etests on 36 blood isolates of MRSA over 9 months we explored the effects of isolate storage on vancomycin and daptomycin MIC. We identified overall declines in vancomycin and daptomycin MIC taken from the same isolates at 3 monthly intervals (P <0.001). Declines 1 doubling dilution were observed in 75% and 67% of isolates for vancomycin and daptomycin MIC respectively. Effects of storage should be considered in evidence around MIC creep. 1

2 Decreasing effectiveness of vancomycin in the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections has been linked to increasing vancomycin minimum inhibitory concentration (MIC) within the susceptible range ( 2mg/L). (5) Sub-inhibitory vancomycin exposure may also be a risk-factor for daptomycin heterogenous susceptibility. (18, 28, 22) However, evidence of MIC creep is mixed (24) and the implications of a high but susceptible vancomycin MIC for treatment decisions uncertain. (5) We previously demonstrated that detection of vancomycin and daptomycin MIC creep may be method dependent. (7) In this study we further explored the hypothesis that isolate storage may explain divergent results in investigations of susceptibility trends A prospective repeated-measures design involved susceptibility testing of MRSA blood isolates at the time of isolation and at 3 monthly intervals. A 9-month follow up period was chosen on the basis of previous data suggesting that changes in susceptibility occurred within this time. Our sample included all non-replicate MRSA bloodstream isolates (BSI) identified in adult, non-obstetric, patients at Aberdeen Royal Infirmary (Scotland) between January and March Susceptibility testing at the time of isolation was by Etest performed as per manufacturer s guidelines on Mueller-Hinton agar (Oxoid), and read blind in duplicate. Each isolate was then stored in Cryobank storage containers (Mast diagnostics) which contain glycerol, peptones, sucrose and saline, at C, recovered from storage at 3,6 and 9 months, and sub-cultured twice prior to repeat Etests. Assessors were blinded to previous MIC results Inter-observer agreement was assessed by weighted Cohen s kappa. Friedman tests were used to test for overall difference in MIC across repeated readings with post-hoc analyses by Wilcoxon-signed rank tests with Bonferonni adjustment (α = ). Variation in trends were investigated by independent-samples t-test to assess differences in mean of slope in isolates grouped by high (>median) or low (< median) baseline MIC. All analyses were repeated with MIC from Etest converted to doubling dilutions by rounding up of intermediate increments To evaluate the potential for systematic bias arising from differences in storage of isolates before susceptibility testing in studies reporting MIC creep, we searched Medline and PubMed databases for 2

3 original articles published after 2000 (all languages) using the following search terms: ( MIC Creep or resistance trend* or susceptibility trend* or decreas* susceptibility) and ( vancomycin or glycopeptide ) and ( Staphylococcus aureus or MRSA or Gram-positive). Reference lists of original articles were also searched. We excluded case-studies, studies of treatment effects, cross-sectional designs, susceptibility testing by automated agar dilution methods, and reports of vancomycin resistance without reference to actual MICs Thirty-six MRSA BSI were identified between January and March All duplicate readings were within one E-test increment with excellent inter-observer agreement (Κ= 0.980). There were significant differences in vancomycin and daptomycin (P <0.001) MIC from the same isolates measured at 3 monthly intervals - table 1. Findings were unchanged by use of doubling-dilutions. Declines 1 and 2 doubling dilution were observed in 67%-75% and 11% of isolates respectively figure 1. Post-hoc comparisons (see supplemental data) found that declines in vancomycin and daptomycin MICs occurred within 3 months and after 6 months respectively figure 1. Average decline in vancomycin MIC was significantly steeper in isolates with higher MIC (> 1.0 mg/l) at baseline (-0.08 vs month -1 ; P =0.002) Trends in daptomycin susceptibility were not related to baseline MIC A literature search identified 376 papers. After exclusions 16 were relevant. Review of references in these papers identified 3 other studies. Both studies performed at the time of isolation suggested evidence of vancomycin MIC creep, however creep was less consistently found by Etest or broth microdilution (BMD) after storage, although heterogeneity in study population and size was noted see supplemental data This prospective longitudinal study found evidence that in daptomycin and vancomycin susceptible blood isolates, MIC results from Etest were inversely related to the duration of isolate storage prior to susceptibility testing. A review of studies describing vancomycin MIC creep suggested failure to account for storage may introduce systematic errors. 3

4 Our study had a number of limitations. A small sample size may have caused under- or overestimation of the effects of storage. Consistency in the direction and the size of changes in MIC suggested that trends were not explained by lack of precision in testing method. Finally, we did not define MRSA strains of isolates which may be important for generalizability. We have previously reported dominance of MLST clonal complexes 22 and 30 in MRSA blood isolates in the same population and differences in MIC after storage irrespective of strain. (7) Although a number of studies have found vancomycin and daptomycin MIC and MIC creep to be method dependent (19, 25, 27, 31) we are not aware of any detailing effects of storage of MRSA isolates on MIC in susceptible isolates. van Griethuysen et al. found a relationship between loss of the meca gene in MRSA isolates and duration of storage, suggesting that meca negative cells may have a survival advantage, or that loss of the meca occurs during storage: such deletions have been associated with Vancomycin Intermediate S.aureus (VISA). (15) A larger study found no evidence of loss of resistance but suggested variable genetic stability of MRSA strains and freeze-thaw strains of cryostocking may be important to reliability of susceptibility testing in frozen isolates. (33) The vancomycin resistant phenotype is known to be unstable in the absence of selective pressure (15) with vancomycin-susceptible revertant mutations observed after repeated passages on non-selective media. (3) Genetic instability may impact phenotypic characteristics correlated with low-level vancomycin resistance including cell-wall thickness. (4, 12, 15) Differences in results from prospective and retrospective susceptibility testing raise concerns around interpretation of studies conducted on isolates after extended periods of storage. A recent metaanalysis finding association between high vancomycin MIC within the susceptible range and outcomes in MRSA infections accounted for susceptibility testing method but not storage of isolates. (32) There is a need to clarify the relevance of high but susceptible vancomycin MICs to treatment decisions (5) which should also be informed by : baseline health; site (34) and severity of infection; source elimination; patient and population antibiotic exposures (9, 21); MRSA strain (15) and regional factors. (8,17) The performance of routinely used methods in detecting low-level glycopeptide resistance has been questioned, particularly given apparent instability of resistant 4

5 phenotypes in-vitro. (10, 16) and our study suggested effects of storage should be included in standardisation of susceptibility testing. (15) Interpretation of evidence around MIC creep and effects on treatment efficacy should account for susceptibility testing methods used and storage prior to testing. We suggest that clinical relevance of findings will be optimised where MIC results are from susceptibility testing around the time of isolation. This may be particularly relevant to surveillance programmes receiving stored isolates ACKNOWLEDGEMENTS No direct funding was attached to this study. We are grateful to the staff in the diagnostic laboratory and media departments of Aberdeen Royal Infirmary for their help IM Gould has received personal and grant financial support from companies manufacturing diagnostics and therapeutics for MRSA. B. Edwards has received grant financial support from Novartis. T. Lawes and F. Ludwig have no conflicts of interest to declare. 116 REFERENCES Adam HJ, Louie L, Watt C, Gravel D, Bryce E, Loeb M, Matlow A, McGeer A, Mulvey MR, Simor AE Canadian Nosocomial Infection Surveillance Program. Detection and characterization of heterogeneous vancomycin-intermediate Staphylococcus aureus isolates in Canada: results from the Canadian Nosocomial Infection Surveillance Program, Antimicrob Agents Chemother. 54(2): Alós JI, García-Cañas A, García-Hierro P, Rodríguez-Salvanés F Vancomycin MICs did not creep in Staphylococcus aureus isolates from 2002 to 2006 in a setting with low vancomycin usage. J Antimicrob Chemother. 62(4):

6 Boyle-Vavra S, Berke SK, Lee JC, Daum RS Reversion of the glycopeptide resistance phenotype in Staphylococcus aureus clinical isolates. Antimicrob Agents Chemother. 44(2): Cui L, Ma X, Sato K, Okuma K, Tenover FC, Mamizuka EM, Gemmell CG, Kim MN, Ploy MC, El-Solh N, Ferraz V, Hiramatsu K Cell wall thickening is a common feature of vancomycin resistance in Staphylococcus aureus. J Clin Microbiol. 41(1): Dhand A, Sakoulus G Reduced vancomycin susceptibility among clinical Staphylococcus aureus isolates ( the MIC Creep ): implications for therapy. F1000 Medicine Reports.4: Dhawan B, Gadepalli R, Rao C, Kapil A, Sreenivas V Decreased susceptibility to vancomycin in meticillin-resistant Staphylococcus aureus: a 5 year study in an Indian tertiary hospital. J Med Microbiol. 59(3): Edwards B, Milne K, Lawes T, Cook I, Robb A, Gould IM Is Vancomycin MIC "Creep" Method Dependent? Analysis of Methicillin-Resistant Staphylococcus aureus Susceptibility Trends in Blood Isolates from North East Scotland from 2006 to J Clin Microbiol. 50(2): Gasch O, Ayats J, Angeles Dominguez M, Tubau F, Liñares J, Peña C, Grau I, Pallarés R, Gudiol F, Ariza J, Pujol M Epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infection: secular trends over 19 years at a university hospital. Medicine (Baltimore). 90(5): Han JH, Bilker WB, Edelstein PH, Mascitti KB, Lautenbach E. 10 April Derivation and validation of clinical prediction rules for reduced vancomycin susceptibility in Staphylococcus aureus bacteraemia. Epidemiol Infect.1-9. doi: /S Hawkey PM Low-level glycopeptide resistance in methicillin-resistant Staphylococcus aureus and how to test it. Clin Microbiol Infect. 15 Suppl 7:2-9. 6

7 Hawser SP, Bouchillon SK, Hoban DJ, Dowzicky M, Babinchak T Rising incidence of Staphylococcus aureus with reduced susceptibility to vancomycin and susceptibility to antibiotics: a global analysis Int J Antimicrob Agents. 37(3): Hiramatsu K. Vancomycin-resistant Staphylococcus aureus: a new model of antibiotic resistance. Lancet Infect Dis. 1(3): Ho PL, Lo PY, Chow KH, Lau EH, Lai EL, Cheng VC, Kao RY. Vancomycin MIC creep in MRSA isolates from 1997 to 2008 in a healthcare region in Hong Kong. J Infect. 2010;60(2): Holmes RL, Jorgensen JH Inhibitory activities of 11 antimicrobial agents and bactericidal activities of vancomycin and daptomycin against invasive methicillin-resistant Staphylococcus aureus isolates obtained from 1999 through Antimicrob Agents Chemother. 52(2): Howden BP, Davies JK, Johnson PD, Stinear TP, Grayson ML Reduced vancomycin susceptibility in Staphylococcus aureus, including vancomycin-intermediate and heterogeneous vancomycin-intermediate strains: resistance mechanisms, laboratory detection, and clinical implications. Clin Microbiol Rev. 23(1): Jones RN Microbiological features of vancomycin in the 21st century: minimum inhibitory concentration creep, bactericidal/static activity, and applied breakpoints to predict clinical outcomes or detect resistant strains. Clin Infect Dis. 42 Suppl 1:S Kehrmann J, Kaase M, Szabados F, Gatermann SG, Buer J, Rath PM, Steinmann J Vancomycin MIC creep in MRSA blood culture isolates from Germany: a regional problem? Eur J Clin Microbiol Infect Dis. 30(5): Kelley PG, Gao W, Ward PB, Howden BP Daptomycin non-susceptibility in vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous-visa (hvisa): implications for therapy after vancomycin treatment failure. J Antimicrob Chemother. 66(5):

8 Kruzel MC, Lewis CT, Welsh KJ, Lewis EM, Dundas NE, Mohr JF, Armitige LY, Wanger A Determination of Vancomycin and Daptomycin MICs by Different Testing Methods for Methicillin-Resistant Staphylococcus aureus. J. Clin. Microbiol. 49(6): Lagacé-Wiens P, Adam H, Nichol K, Decorby M, Mulvey M, Karlowsky J, Zhanel1 GG, Hoban1 D Vancomycin and Daptomycin MIC Creep in Staphylococcus spp.in Canada Analysis of 4642 S. aureus and 497 S. epidermidis isolates from Poster presented at Interscience Conference on Antimicrobial Agents and Chemotherapy, Chicago, USA, September 17 20, Lubin AS, Snydman DR, Ruthazer R, Bide P, Golan Y Predicting high vancomycin minimum inhibitory concentration in methicillin-resistant Staphylococcus aureus bloodstream infections. Clin Infect Dis. 52(8): Moise PA, North D, Steenbergen JN, Sakoulas G Susceptibility relationship between vancomycin and daptomycin in Staphylococcus aureus: facts and assumptions. Lancet Infect Dis. 9(10): Musta AC, Riederer K, Shemes S, Chase P, Jose J, Johnson LB, Khatib R Vancomycin MIC plus heteroresistance and outcome of methicillin-resistant Staphylococcus aureus bacteremia: trends over 11 years. J Clin Microbiol. 47(6): Pitz AM, Yu F, Hermsen ED, Rupp ME, Fey PD, Olsen KM Vancomycin susceptibility trends and prevalence of heterogeneous vancomycin-intermediate Staphylococcus aureus in clinical methicillin-resistant S. aureus isolates. J Clin Microbiol. 49(1): Prakash V, Lewis II JS, and Jorgensen JH Vancomycin MICs for Methicillin-Resistant Staphylococcus aureus Isolates Differ Based upon the Susceptibility Test Method Used. Antimicrob. Agents Chemother. 52(12):

9 Robert J, Bismuth R, Jarlier V Decreased susceptibility to glycopeptides in methicillin- resistant Staphylococcus aureus: a 20 year study in a large French teaching hospital, J Antimicrob Chemother. 57(3): Sader HS, Rhomberg PR, Jones RN Nine-hospital study comparing broth microdilution and Etest method results for vancomycin and daptomycin against methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 53(7): Sakoulas G, Alder J, Thauvin-Eliopoulos C, Moellering RC Jr, Eliopoulos GM Induction of daptomycin heterogeneous susceptibility in Staphylococcus aureus by exposure to vancomycin. Antimicrob Agents Chemother. 50(4): Steinkraus G, White R, Friedrich L Vancomycin MIC creep in non-vancomycin- intermediate Staphylococcus aureus (VISA), vancomycin-susceptible clinical methicillin-resistant S. aureus (MRSA) blood isolates from J. Antimicrob. Chemother. 60: van Griethuysen A, van Loo I, van Belkum A, Vandenbroucke-Grauls C, Wannet W, van Keulen P, Kluytmans J Loss of the meca gene during storage of methicillin-resistant Staphylococcus aureus strains. J Clin Microbiol.43(3): van Hal SJ, Barbagiannakos T, Jones M, Wehrhahn MC, Mercer J, Chen D, Paterson DL, Gosbell IB Methicillin-resistant Staphylococcus aureus vancomycin susceptibility testing: methodology correlations, temporal trends and clonal patterns. J Antimicrob Chemother. 66(10): van Hal SJ, Lodise TP, Paterson DL The clinical significance of vancomycin minimum inhibitory concentration in Staphylococcus aureus infections: a systematic review and meta-analysis. Clin Infect Dis. 54(6): Veguilla W, Peak KK, Luna VA, Roberts JC, Davis CR, Cannons AC Two-year study evaluating the potential loss of methicillin resistance in a methicillin-resistant Staphylococcus aureus culture collection. J Clin Microbiol. 46(10):

10 Walraven CJ, North MS, Marr-Lyon L, Deming P, Sakoulas G, Mercier RC Site of infection rather than vancomycin MIC predicts vancomycin treatment failure in methicillin-resistant Staphylococcus aureus bacteraemia. J Antimicrob Chemother. 66(10): Wang G, Hindler JF, Ward KW, Bruckner DA Increased Vancomycin MICs for Staphylococcus aureus Clinical Isolates from a University Hospital during a 5-Year Period. J Clin Microbiol. 44(11): Zhao C, Sun H, Wang H, Liu Y, Hu B, Yu Y, Sun Z, Chu Y, Cao B, Liao K, Lei J, Hu Z, Zhang L, Zhang X, Xu Y, Wang Z, Chen M. 21 April Antimicrobial resistance trends among 5608 clinical Gram-positive isolates in China: results from the Gram-Positive Cocci Resistance Surveillance program ( ). Diagn Microbiol Infect Dis. doi: /j.diagmicrobio Table 1: MIC (mg/litre) for vancomycin and daptomycin from Etests repeated on same isolates at 3 month intervals No.(%) of isolates with No. of Mean MIC (95% Modal MIC > baseline Time of testing isolates CI) MIC MIC range median MIC50 MIC Vancomycin Baseline (0.63 to 1.79) to (47%) months (0.15 to 1.54) to (14%) months (0.38 to 1.16) to (0%) months (0.25 to 1.04) to (0%) Daptomycin Baseline (0.00 to 0.47) to (50%) months (0.01 to 0.43) to (42%) months (-0.03 to 0.44) to (39%) months (0.04 to 0.20) to (3%)

11 Figure 1: Change in (a) vancomycin and (b) daptomycin MIC by time period as measured in doubling dilutions and Etest increments and trend in mean MIC (Etest) by baseline MIC * Significant difference between these datapoints by related-samples Wilcoxon-signed rank-test

12 ) (a) Vancomycin No. of isolates months 6 months 9 months * MIC (mg/l) All (with 95% CI) MIC 1 (with 95% CI) MIC > 1 mg/l (with 95% CI) * No. of MIC increments different (doubling dilutions) No. of MIC increments different (E-test increments) m 3m 6m 9m Time from isolation (b) Daptomycin All (with 95% CI) MIC 0.19 (with 95% CI) No. of isolates MIC (mg/l) MIC > 0.19 mg/l (with 95% CI) * m 3m 6m 9m No. of MIC increments different (doubling dilutions) No. of MIC increments different (Etest increments) Time from isolation

13 3m 6m 9m 0m 3m 6m 9m 0m 3m 6m 9m 0m 3m 6m m 6m 9m 0m 3m 6m 9m m 3m 6m 9m 0m 3m 6m 22 3m 6m 9m 0m 3m 6m 9m m 3m 6m 9m 0m 3m 6m 28

14 3m 6m 9m 0m 3m 6m 9m 0m 3m 6m 9m 0m 3m m 6m 9m 0m 3m 6m 9m m 3m 6m 9m 0m 3m m 6m 9m 0m 3m 6m 9m 8 9 0m 3m 6m 9m 0m 3m 6 10