In vitro activity of ceftaroline and comparator antimicrobials against European and Middle East isolates from complicated skin and skin-structure infections collected in 2008–2009

International Journal of Antimicrobial Agents 40 (2012) 227–234

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In vitro activity of ceftaroline and comparator antimicrobials against European and Middle East isolates from complicated skin and skin-structure infections collected in 2008–2009 Ian Morrissey ∗ , Anne Leakey, John Blackman Northwood Quotient Bioresearch Ltd., Fordham, UK

a r t i c l e

i n f o

Article history: Received 2 March 2012 Accepted 14 May 2012 Keywords: Ceftaroline fosamil Novel cephalosporin Europe Middle East

a b s t r a c t The activities of ceftaroline, the active metabolite of the pro-drug ceftaroline fosamil, a novel antimeticillin-resistant staphylococcal cephalosporin, and nine comparators were determined against surveillance isolates collected in 2008–2009. Over 3000 isolates associated with complicated skin and skin-structure infections (cSSSIs) were collected from 106 centres in 19 countries. MICs were determined using CLSI broth microdilution methodology. Clonal relatedness of meticillin-resistant Staphylococcus aureus (MRSA) with raised ceftaroline MICs (2 mg/L) was assessed by MLST, PFGE and mec typing. The presence of Panton–Valentine leukocidin in these isolates was also determined. Ceftaroline was active against 500 MRSA and 479 meticillin-susceptible S. aureus, with MIC50/90 values of 0.5/2 mg/L and 0.25/0.25 mg/L, respectively. For coagulase-negative staphylococci (CoNS), the ceftaroline MIC50/90 values for meticillin-resistant strains (n = 159) were the same as those seen for MRSA. Meticillin-susceptible CoNS (n = 113) had the same MIC90 as that seen with S. aureus, but the MIC50 was lower at 0.06 mg/L. Ceftaroline was also active against ␤-haemolytic streptococci (n = 526; MIC50/90 = 0.004/0.015 mg/L), other streptococci (n = 75; 0.015/0.06 mg/L), common Enterobacteriaceae (n = 897; 0.25/≥128 mg/L) and Enterococcus faecalis (n = 329; 1/16 mg/L). Those MRSA with ceftaroline MICs of 2 mg/L were found to be from four clonal groups associated with the country of origin. These data confirm the broad-spectrum in vitro activity of ceftaroline against cSSSI pathogens. Ceftaroline is unique among clinically available cephalosporins, having good in vitro activity against MRSA and meticillin-resistant CoNS and moderate activity against Gram-negative bacteria. © 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

1. Introduction Ceftaroline fosamil (formerly PPI-0903 and TAK-599), the prodrug of the active compound ceftaroline, is a broad-spectrum, parenteral cephalosporin that was recently approved in the USA for the treatment of acute bacterial skin and skin-structure infections (SSSIs) and community-acquired bacterial pneumonia. It is active in vitro against Gram-positive species [1–3], including meticillinresistant Staphylococcus aureus (MRSA), and variably active against Gram-negative species owing to lack of activity against class A and C ␤-lactamases [1–5]. The aim of this study was to determine the in vitro activity of ceftaroline compared with that of a number of other antimicrobials against isolates from Austria, Bulgaria, Czech Republic, Egypt, France, Germany, Greece, Ireland, Israel, Italy, Poland, Portugal,

∗ Corresponding author. Present address: IHMA Europe Sàrl, 9A Route de la Corniche, 1066 Epalinges, Switzerland. Tel.: +44 1279 724 929. E-mail address: [email protected] (I. Morrissey).

Russia, Slovakia, Spain, Switzerland, The Netherlands, Turkey and the UK collected from hospitalised patients with complicated SSSIs (cSSSIs) during 2008–2009. The collection would be expected to include a diverse population of clonal types, particularly MRSA, which will differ from those in North America.

2. Materials and methods 2.1. Isolates and identification In total, 3078 clinical isolates from cSSSIs were collected from 106 centres in 19 countries across Europe and the Middle East between May 2008 and June 2009 (Table 1). Only isolates considered to be clinically significant by a microbiologist or infectious disease specialist were included in the study, which included Staphylococcus spp., ␤-haemolytic and other Streptococcus spp., Escherichia coli, Klebsiella pneumoniae, Enterobacter spp. and Enterococcus faecalis (Table 2). The majority of the isolates were from wounds (75.3%) and skin (16.1%); the remaining isolates were

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I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

Table 1 Countries and number of sites participating in the study. Country Austria Bulgaria Czech Republic Egypt France Germany Greece Ireland Israel Italy Poland Portugal Russia Slovakia Spain Switzerland The Netherlands Turkey UK Total

No. of isolates

No CLSI or EUCAST breakpoints are currently available for ceftaroline. No. of centres

58 75 32 11 193 371 88 41 38 253 196 88 478 23 400 104 76 280 273

2 2 1 1 9 12 3 1 2 13 5 3 10 1 15 4 3 11 8

3078

106

2.3. Molecular characterisation of MRSA with raised ceftaroline minimum inhibitory concentrations MRSA isolates (n = 54) from sites where more than one isolate had raised ceftaroline MIC values of 2 mg/L were genotyped to assess the presence of Panton–Valentine leukocidin (PVL) [9] and the staphylococcal cassette chromosome mec (SCCmec) type (types I–VI) [10] as well as performing multilocus sequence typing (MLST) [11] and pulsed-field gel electrophoresis (PFGE) [12]. Clonal nomenclature was based on that described by Enright et al. [13]. Isolates were grouped by MLST, followed by PFGE patterns for isolates within the same MLST sequence type (where there were more than three isolates present in the clonal group). PFGE patterns were analysed using Phoretix 1D Advanced software v5.20 and Phoretix 1D Database v2.00 (Nonlinear Dynamics Ltd., Newcastle upon Tyne, UK). Isolates that showed ≥80% similarity were considered to be within the same PFGE group [14]. 3. Results

sourced from blood (3.0%), abscess, ulcer or cellulitis (1.3%), pus (1.1%), tissue (0.5%) and other (2.7%). The identities of the isolates were re-confirmed at a central laboratory using standard methods as follows: S. aureus, DNAase and Staphytect plus (Oxoid Ltd., Basingstoke, UK); other Staphylococcus spp., Id 32 Staph (bioMérieux UK Ltd., Basingstoke, UK); and ␤-haemolytic Streptococcus spp., Lancefield group (Oxoid Streptococcal Grouping Kit; Oxoid Ltd.). API 20 Strep (bioMérieux) was used for streptococci that did not belong to Groups A, B, C or G as well as enterococci. Enterobacteriaceae were identified using API 20 E (bioMérieux). The zone diameter obtained with a 30 ␮g cefoxitin disk was used to determine meticillin resistance in staphylococci. 2.2. Minimum inhibitory concentration (MIC) determination MICs were determined by the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method [6] using plates prepared in-house and stored at −80 ◦ C until required. Cation-adjusted Mueller–Hinton II broth (Becton, Dickinson U.K. Ltd., Oxford, UK) was used to prepare the plates. The comparator antimicrobials were cefuroxime, ceftazidime, ciprofloxacin, amoxicillin/clavulanic acid, daptomycin, levofloxacin, linezolid, moxifloxacin, penicillin, piperacillin/tazobactam, tigecycline, teicoplanin and vancomycin. Broth used for tigecycline was prepared on the day of plate manufacture and plates were frozen within 8 h of broth preparation. The calcium ion content of the broth used for daptomycin MIC determination was adjusted to a final concentration of 50 mg/L. Each batch of plates was subjected to full quality control using CLSI reference strains, including some or all of E. coli ATCC 25922 and ATCC 35218, S. aureus ATCC 29213, E. faecalis ATCC 29212 and Streptococcus pneumoniae ATCC 49619. Susceptibility for comparator antimicrobials was determined using breakpoints set by the CLSI [7], except where European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints [8] were used, as follows: • ␤-haemolytic streptococci: moxifloxacin, tigecycline and teicoplanin; • other streptococci: cefuroxime, tigecycline and teicoplanin; • Enterobacteriaceae: moxifloxacin and tigecycline; • staphylococci: tigecycline • enterococci: tigecycline.

The ceftaroline MIC distribution for each bacterial species is shown in Table 3. Summary MIC data for ceftaroline and comparators are shown in Tables 4 and 5. No significant differences were noted between ceftaroline MIC results from the different countries and between patients in different age groups (data not shown). 3.1. Activity against staphylococci The maximum ceftaroline MIC obtained for all the S. aureus isolates was 2 mg/L (Table 3). MIC50 and MIC90 values (MICs for 50% and 90% of the organisms, respectively) for meticillin-susceptible S. aureus (MSSA) were both 0.25 mg/L, whilst the MIC50 of MRSA was only one dilution higher at 0.5 mg/L (MIC90 = 2 mg/L). This is considerably more active than other ␤-lactams and fluoroquinolones (Table 4). There were two MRSA (one from Russia and the other from Spain) with a vancomycin MIC of 4 mg/L (intermediate susceptibility), both of which had a ceftaroline MIC of 1 mg/L and were susceptible to daptomycin and linezolid (data not shown). Meticillin-susceptible coagulase-negative staphylococci (MSCoNS) were generally very susceptible to ceftaroline, with a MIC50 of 0.06 mg/L and a MIC90 of 0.25 mg/L. Meticillin-resistant coagulase-negative staphylococci (MR-CoNS) had the same MIC50 and MIC90 values as MRSA. One isolate of meticillin-resistant Staphylococcus capitis (from Italy) was exceptional, with a ceftaroline MIC of ≥128 mg/L. Three MR-CoNS isolates had a ceftaroline MIC of 4 mg/L [1 S. capitis (Russia) and 2 Staphylococcus haemolyticus (Portugal)]. All staphylococci were susceptible to daptomycin and linezolid, whilst 1% (12 isolates) were nonsusceptible to tigecycline: 2 MRSA (Russia), 2 MSSA (Turkey and Poland), 1 meticillin-susceptible S. haemolyticus (Greece), 1 meticillin-susceptible Staphylococcus epidermidis (Poland), 2 meticillin-resistant S. haemolyticus (Poland) and 4 meticillinresistant S. epidermidis (2 Italy, 1 Turkey and 1 UK). 3.2. Activity against streptococci Ceftaroline was very active against the ␤-haemolytic streptococci (Table 3). The MIC90 was 0.015 mg/L with a maximum MIC of 0.03 mg/L for all the ␤-haemolytic groups. The least effective antimicrobial was levofloxacin at 98.5% susceptible (Table 4). The levofloxacin-non-susceptible isolates (n = 8) were 1 Group A streptococcus (Italy), 2 Group B streptococci (Italy and Spain), 2 Group

Table 2 Distribution of test isolates by geographical location. Group

Pathogen

Country of origin Austria Bulgaria Czech Republic

Other streptococci (N = 75)

Enterobacteriaceae (N = 897)

Staphylococcus aureus (N = 979) CoNS (N = 272)

Enterococci (N = 329)

Group A Group B Group C Group G anginosus group bovis group mitis group salivarius group All Enterobacter spp. E. aerogenes E. amnigenus E. asburiae E. cloacae E. gergoviae Escherichia coli Klebsiella pneumoniae MRSA MSSA All MR-CoNS All MS-CoNS S. epidermidis (MR, 104) S. haemolyticus (MR, 32) S. hominis (MR, 12) S. capitis (MR, 6) Other CoNS (MR, 5) E. faecalis

6 3 3 5

3 5

1 1 1 1

1 1 5

6

5

1 1 5

21 19 9 21

5 3

2

3

6

2

4

3 1

1

17

36

5

6

6

6

1

2 2

3

7 11 4 5

5 6 1 4

14 9 20 14

4

1

1

1

2

2

1

22

16

1 3

15 17

39 22 3 7

12 6 4 11

4 3 2 3

11 6

8

3

1

1

1 1

5 3

4

16

57

1

1

3

1 8

24 21 11 21

196 156 61 113

2 1 4 1

1 4 1

32 1 35 7

30

19

295

3

3

1

39

6

5

2

1 33

4

8

27

16

11

30

4

6

1

17

15

16

54

1

11 6

46 30

12 5

8 6

5 3

36 21

31 20

8 4

91 29

2

1

45 29

7 3

7 9

47 19

20 12

35 2 1 256 1 401 201

2 1

34 38

53 55

16 15

2

9 8

30 38

25 32

14 11

74 63

3 6

71 66

17 16

14 10

60 51

56 44

500 479

5 8 8

16 10 21

5 5 6

5 2 2

4

20 12 19

10 8 10

5 2 4

20 8 7

1 1

19 9 14

6 10 11

5 3 7

17 11 17

9 11 12

159 113 164

1

1

1

2

5

3

2

12

3

6

2

38

1

1

3 1 4

2 1 2

1

1 6 2

4

3 2

1 3 2

20 17 33

33

24

9

19

19

329

5

6 1

5 2

3 11

11 10

6 4

5 4 9

7 9 11

1 1

3 8

5

3 3

1

3 1 14 1

5

22 19 2 13

5

7

3

2 2

13

45

1 1

13

4

2 2 1 5

3

63

1

3

7

5

44

5

6

I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

␤-Haemolytic streptococci (N = 526)

Egypt France Germany Greece Ireland Israel Italy Poland Portugal Russia Slovakia Spain Switzerland The Turkey UK Total Netherlands

MRSA, meticillin-resistant S. aureus; MSSA, meticillin-susceptible S. aureus; CoNS, coagulase-negative staphylococci; MR, meticillin-resistant; MS, meticillin-susceptible.

229

0.004 0.015 0.015 0.008 0.03 8 ≥128 64 ≥128 ≥128 ≥128 2 0.25 1 2 0.25 0.5 0.5 0.12 2 16 0.004 0.015 0.008 0.004 0.015 0.015 0.5 2 0.5 0.12 0.12 0.5 0.25 0.25 0.5 0.06 0.25 0.25 0.06 2 1

MIC50 (mg/L)

43 2 41 45 39 0 0 1 1 0 0 0 0 0 11

≥128

0 0 0 0 0 1 12 0 12 29 5 158 2 16 16 0 4 4 0 8 153 0 0 0 0 0 2 8 3 4 119 54 0 1 63 3 60 44 2 42 0 0 140 3 23 58 0 6 Group A streptococci (196) Group B streptococci (156) Group C streptococci (61) Group G streptococci (113) Anginosus group streptococci (32) Mitis group streptococci (35) Enterobacter spp. (295) Enterobacter aerogenes (35) Enterobacter cloacae (256) Escherichia coli (401) Klebsiella pneumoniae (201) MRSA (500) MSSA (479) All CoNS (272) MR-CoNS (159) MS-CoNS (113) S. epidermidis (164) MR S. epidermidis (104) MS S. epidermidis (60) S. haemolyticus (38) E. faecalis (329)

6 0 1 0 1 2

32 2 2 2 0 0

16 38 24 49 4 4 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0

2 113 10 4 16 7 0 0 0 4 0 1 0 4 0 4 1 0 1 0 1

0 0 1 0 11 9 2 0 0 30 3 0 0 6 0 6 4 0 4 0 0

0 0 0 0 0 0 48 9 38 77 62 0 68 36 10 26 18 8 10 1 0

0 0 0 0 0 2 81 1 80 42 13 15 398 77 62 15 57 54 3 5 1

0 0 0 0 0 1 33 4 29 25 12 242 9 45 43 2 36 36 0 3 11

0 0 0 0 0 0 8 1 7 4 6 84 1 21 21 0 0 0 0 19 52

0 0 0 0 0 0 3 1 2 4 4 0 0 3 3 0 0 0 0 2 9

0 0 0 0 0 1 11 2 9 4 1 0 0 0 0 0 0 0 0 0 60

13 3 10 2 0 0 0 0 0 0 0 0 0 0 12

12 5 7 4 0 0 0 0 0 0 0 0 0 0 16

21 4 17 12 1 0 0 0 0 0 0 0 0 0 3

64 32 16 8 4 2 1 0.5 0.25 0.12 0.06 0.03 0.015 0.008 0.004 0.002 ≤0.001

No. of isolates at ceftaroline MIC (mg/L) Group/species (N)

Table 3 Minimum inhibitory concentration (MIC) distribution for ceftaroline against all bacterial species collected.

MIC50/90 , MIC for 50% and 90% of the isolates, respectively; MRSA, meticillin-resistant S. aureus; MSSA, meticillin-susceptible S. aureus; CoNS, coagulase-negative staphylococci; MR, meticillin-resistant; MS, meticillin-susceptible.

I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

MIC90 (mg/L)

230

C streptococci (Poland) and 3 Group G streptococci (2 Ireland and 1 UK). The other streptococci generally exhibited a greater range of MICs across all the antimicrobials tested, with 21 isolates (28%) being penicillin-non-susceptible (Table 4); 5 isolates (all mitis group) were penicillin-resistant (2 Turkey, 2 UK and 1 France) and the remaining 16 were penicillin-intermediate (13 mitis group and 3 salivarius group from various countries). Like the ␤-haemolytic streptococci, the other streptococci were all susceptible to daptomycin, linezolid, teicoplanin and vancomycin. The ceftaroline MIC90 was 0.06 mg/L, with only two isolates (both Streptococcus mitis from Turkey) having a ceftaroline MIC ≥ 1 mg/L. 3.3. Activity against Enterococcus faecalis The E. faecalis were more susceptible to ceftaroline than to cefuroxime (MIC50 = 1 mg/L and ≥128 mg/L, respectively) (Table 4). There were only 17 vancomycin-resistant isolates among all the E. faecalis. The majority of these (16 isolates) had a ceftaroline MIC of ≥8 mg/L compared with only 86 isolates (27.6%) within the vancomycin-susceptible enterococci population (data not shown). There were 13 amoxicillin/clavulanic acid-resistant E. faecalis, 12 of which had a ceftaroline MIC of ≥8 mg/L (5 from Egypt and 1 each from Germany, Greece, Poland, Russia, Spain, Turkey and the UK). Only daptomycin was 100% effective; 2 isolates were linezolid-intermediate (Spain and Turkey), 1 was tigecyclineresistant (Spain) and 3 were tigecycline-intermediate (Italy, Russia and UK). Approximately 40% of the E. faecalis were fluoroquinoloneresistant (Table 4). 3.4. Activity against Enterobacteriaceae The ceftaroline MIC distribution was lowest for E. coli followed by K. pneumoniae and all Enterobacter spp. (Table 3). A high ceftaroline MIC was found in some Enterobacteriaceae and almost all of these were ceftazidime-resistant (Table 5). The ceftazidimeresistant subsets were also highly resistant to other classes of antimicrobials (Table 5) and probably reflect extended-spectrum ␤-lactamase-producers or isolates with de-repressed AmpC. 3.5. Molecular characterisation of MRSA with raised ceftaroline minimum inhibitory concentrations At the time of molecular analysis, 1 of 54 isolates eligible for testing was not available owing to loss of viability during storage. Of the remaining 53 isolates, all were negative for the presence of PVL; 17 were SCCmec type I, 3 were type III and 33 were an undetermined variant of type III. These data suggest that the MRSA with raised ceftaroline MICs were hospital-associated strains. MLST analysis identified four clonal groups: • ST111-MRSA-I (7 from Bulgaria) • ST228-MRSA-I [1 from Germany, 4 from Italy (one with a novel single-locus variant of ST228) and 4 from Spain] • ST239-MRSA-III [3 from Greece, 33 from Turkey with an undetermined variant of SCCmec type III (including 6 containing one of three novel single-locus variants of ST239)] • ST461-MRSA-I (1 isolate from The Netherlands). Analysis of the PFGE patterns of clonal groups ST239-MRSA-III (n = 36) showed five PFGE clusters of ≥80% similarity (A–E; Fig. 1) and three unrelated isolates with <80% similarity to any of the other isolates tested. Two of the three isolates from Greece grouped into cluster E and one showed <80% similarity to any other isolate. The 33 isolates from Turkey were fairly evenly distributed into clusters

I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

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Table 4 Summary of minimum inhibitory concentration (MIC) results of all species tested for ceftaroline and comparator antimicrobials against Gram-positive bacteria. Species (N)

Antimicrobial

Susceptibility %S

MIC (mg/L) %I

%R

Min.

MIC50

MIC90

Max.

MRSA (500)

Ceftaroline Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanin Daptomycin

– 24.0 0.2 24.2 30.6 9.4 12.4 39.6 99.6 100 99.6 100 100

– 8.8 0 0 0 1.0 1.8 6.0 – – 0.4 0 –

– 67.2 99.8 75.8 69.4 89.6 85.8 54.4 – – 0 0 –

0.015 2 0.12 0.5 1 0.12 0.12 0.015 0.06 0.5 0.25 0.25 ≤0.015

0.5 ≥128 32 16 16 ≥8 ≥16 2 0.25 2 1 0.5 0.25

2 ≥128 64 ≥64 ≥256 ≥8 ≥16 8 0.5 2 1 2 0.5

2 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 1 4 4 8 1

MSSA (479)

Ceftaroline Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanin Daptomycin

– 98.3 22.1 99.0 99.4 91.0 96.2 97.9 99.6 100 100 100 100

– 0.2 0 0 0 4.2 0.4 0.4 0 – 0 0 –

– 1.5 77.9 1.0 0.6 4.8 3.3 1.7 0 – 0 0 –

0.06 0.25 ≤0.03 0.06 ≤0.06 0.06 0.06 0.008 0.06 1 0.25 0.12 0.06

0.25 1 2 0.5 1 0.5 0.25 0.06 0.25 2 1 0.5 0.25

MR-CoNS (159)

Ceftaroline Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanin Daptomycin

– 63.5 2.5 66.7 64.8 28.9 28.9 52.8 96.2 100 100 98.1 100

– 7.6 0 0 0 1.3 2.5 18.2 – – 0 1.9 –

– 28.9 97.5 33.3 35.2 69.8 68.6 28.9 – – 0 0 –

0.06 0.12 0.06 0.12 0.25 0.06 0.12 0.015 0.06 0.25 0.25 0.12 ≤0.015

0.5 4 8 2 4 ≥8 4 0.5 0.5 1 2 4 0.25

MS-CoNS (113)

Ceftaroline Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanin Daptomycin Ceftaroline

– 99.1 38.1 100 99.1 88.5 90.3 92.0 98.2 100 100 98.2 100 –

– 0 0 0 0 0 1.8 3.5 – – 0 1.8 – –

– 0.9 62.0 0 0.9 11.5 8.0 4.4 – – 0 0 – –

0.015 0.06 ≤0.03 ≤0.03 ≤0.06 0.008 0.03 0.03 0.03 0.25 0.25 0.12 0.03 ≤0.001

0.06 0.5 0.5 0.12 0.5 0.25 0.25 0.06 0.25 1 1 1 0.25 0.004

␤-Haemolytic streptococci (526)

Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacina Tigecyclinea Linezolid Vancomycin Teicoplanina Daptomycin Ceftaroline

– 100 – – – 98.5 99.1 99.8 100 100 100 100 –

– 0 – – – 0.6 0.6 0.2 – 0 0 – –

– 0 – – – 1.0 0.4 0 – 0 0 – –

≤0.015 ≤0.008 ≤0.008 0.03 0.06 ≤0.03 0.03 0.015 0.12 0.06 ≤0.03 ≤0.015 ≤0.001

≤0.015 ≤0.008 0.015 0.06 0.5 0.5 0.12 0.06 1 0.5 0.06 0.06 0.015

Other streptococci (75)

Cefuroximea Penicillin AMC TZP Ciprofloxacin

78.7 72.0 – – –

0 21.3 – – –

21.3 6.7 – – –

≤0.015 0.015 ≤0.008 0.03 0.12

0.25 0.06 0.06 0.12 1

0.25 2 16 1 2 1 0.5 0.06 0.25 4 1 1 0.25 2 ≥128 ≥128 32 ≥256 ≥8 ≥16 4 0.5 2 2 8 0.5 0.25 1 2 0.5 1 ≥8 1 0.25 0.5 2 2 4 0.5 0.015 0.06 0.06 0.06 0.25 1 1 0.25 0.12 1 0.5 0.12 0.12 0.06 2 1 1 2 4

2 ≥128 ≥128 ≥64 64 ≥8 ≥16 8 1 4 2 4 1 ≥128 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 2 4 4 16 1 0.5 32 16 2 16 ≥8 ≥16 2 1 4 4 16 1 0.03 0.25 0.12 0.25 0.5 32 32 4 0.5 2 1 0.5 0.5 ≥4 ≥32 ≥32 32 ≥128 32

232

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Table 4 (Continued) Species (N)

Antimicrobial

Susceptibility %S

Enterococcus faecalis (329)

MIC (mg/L) %I

%R

Min.

MIC50

MIC90

Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanina Daptomycin

97.3 – – 100 100 100 100

1.3 – – – 0 0 –

1.3 – – – 0 0 –

0.12 ≤0.015 0.015 0.25 0.25 ≤0.03 ≤0.015

1 0.12 0.06 1 0.5 0.06 0.25

2 0.25 0.12 1 1 0.12 0.5

Ceftaroline Cefuroxime Penicillin AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea Linezolid Vancomycin Teicoplanin Daptomycin

– – 93.3 96.1 – 54.4 57.5 – 98.8 99.4 94.5 95.1 100

– – 2.4 0 – 4.0 0.9 – 0.9 0.6 0.3 0.3 –

– – 4.3 4.0 – 41.6 41.6 – 0.3 0 5.2 4.6 –

0.015 1 0.12 0.06 0.12 0.25 0.5 0.06 0.03 0.5 0.25 0.12 0.12

1 ≥128 2 0.5 4 1 2 0.25 0.12 2 1 0.5 0.5

16 ≥128 8 1 16 ≥8 ≥16 ≥16 0.25 2 2 1 1

Max. 8 1 0.5 2 1 0.5 1 ≥128 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 1 4 ≥128 ≥64 2

S, Susceptible; I, intermediate; R, resistant; MIC50/90 , MIC for 50% and 90% of the isolates, respectively; MRSA, meticillin-resistant Staphylococcus aureus; MSSA, meticillin-susceptible S. aureus; CoNS, coagulase-negative staphylococci; MR, meticillin-resistant; MS, meticillin-susceptible; AMC, amoxicillin/clavulanic acid; TZP, piperacillin/tazobactam. a European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints used where indicated (see Section 2.2).

A–D (7, 7, 8 and 9 isolates, respectively), but two isolates showed <80% similarity to any other isolate tested. The ST228-MRSA-I distributed into three clusters (F, G and H) with two isolates in each cluster (Fig. 2), and 3 showed <80% similarity to any other isolate tested. Six of the seven ST111-MRSA-I grouped into a single cluster (Fig. 3), but DNA from the seventh isolate was not cleaved by SmaI and could not be evaluated.

4. Discussion Ceftaroline is the active component of the inactive pro-drug ceftaroline fosamil. It has potent activity against the penicillin-binding protein 2a (PBP2a), which is encoded by the mecA gene located on the staphylococcal cassette chromosome [2,15]. This activity is reflected by the low MIC results for MRSA in this study (≤2 mg/L) and confirms ceftaroline staphylococcal MIC results obtained in

Table 5 Summary of minimum inhibitory concentration (MIC) results of all species tested for ceftaroline and comparator antimicrobials against Gram-negative bacteria. Species (N)

Antimicrobial

Susceptibility %S

MIC (mg/L) %I

%R

Min.

Enterobacteriaceae (897)

Ceftaroline Ceftazidime Cefuroxime AMC TZP Ciprofloxacin Levofloxacin Moxifloxacina Tigecyclinea

– 75.3 60.8 45.7 78.2 77.2 78.4 74.7 88.6

– 2.3 9.8 13.3 8.5 0.8 2.9 3.9 7.0

– 22.4 29.4 41.0 13.4 22.1 18.7 21.4 4.4

≤0.008 ≤0.03 0.5 ≤0.5 0.25 ≤0.002 0.008 0.008 0.12

Ceftazidime-resistant Enterobacteriaceae (201)

Ceftaroline Ceftazidime Cefuroxime AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecycline

– 0 12.4 9.5 34.8 51.2 55.2 48.3 74.1

– 0 1.5 20.4 22.9 2.0 6.5 8.0 16.4

– 100 86.1 70.2 42.3 46.8 38.3 43.8 9.5

0.03 16 1 1 1 0.008 0.015 0.015 0.25

Ceftazidime-susceptible Enterobacteriaceae (675)

Ceftaroline Ceftazidime Cefuroxime AMC TZP Ciprofloxacin Levofloxacin Moxifloxacin Tigecyclinea

– 100 77.0 57.9 92.0 85.5 85.9 83.3 93.2

– 0 12.6 10.2 3.6 0.4 1.8 2.7 4.3

– 0 10.4 31.9 4.4 14.1 12.3 14.1 2.5

≤0.008 ≤0.03 0.5 ≤0.5 0.25 ≤0.002 0.008 0.008 0.12

MIC50 0.25 0.5 8 16 4 0.015 0.06 0.06 0.5 ≥128 64 ≥128 ≥64 64 1 1 1 1

S, Susceptible; I, intermediate; R, resistant; MIC50/90 , MIC for 50% and 90% of the isolates, respectively. a European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints used where indicated (see Section 2.2).

0.12 0.25 4 8 4 0.015 0.06 0.06 0.5

MIC90

Max.

≥128 ≥128 ≥128 ≥64 128 ≥8 ≥16 ≥16 2

≥128 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 ≥8

≥128 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 2

≥128 ≥128 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 ≥8

1 1 32 ≥64 16 ≥8 8 8 1

≥128 4 ≥128 ≥64 ≥256 ≥8 ≥16 ≥16 ≥8

I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

233

Fig. 1. Dendrogram showing the pulsed-field gel electrophoresis (PFGE) relatedness of clonal group ST239-MRSA-III.

several previous studies [1,4,5,8,15,16]. Other studies have shown that this is associated with bactericidal activity [1,17–19]. CoNS, whilst not necessarily being the aetiological agent of cSSSI, are frequently associated with these infections. In this study, the MIC90 both of MS-CoNS and MR-CoNS were the same as those of the MSSA and MRSA, respectively.

Those MRSA isolates with raised ceftaroline MIC values (2 mg/L) that were tested were all PVL-negative. These strains are associated historically with healthcare-associated infections and are less invasive than their PVL-positive counterparts. There were a number of different clones identified by MLST, including ST239-MRSA-III (also known as the Hungarian/Brazilian clone), ST228-MRSA-I (also

Fig. 2. Dendrogram showing the pulsed-field gel electrophoresis (PFGE) relatedness of clonal group ST228-MRSA-I.

234

I. Morrissey et al. / International Journal of Antimicrobial Agents 40 (2012) 227–234

Fig. 3. Dendrogram showing the pulsed-field gel electrophoresis (PFGE) relatedness of clonal group ST111-MRSA-I.

known as the southern Germany clone) and ST111-MRSA-I, which is a variant of the southern Germany clone. The clones present were generally specific to the country from which they were isolated. Interestingly, ST239 has been found to be the sole clone associated with a raised ceftaroline MIC of 4 mg/L in a previous global surveillance study (although in the current study ceftaroline MICs were not found to be as high as this) [3]. The ceftaroline results for ␤-haemolytic streptococci, E. faecalis and Enterobacteriaceae were similar to those obtained in other studies [1,4,5,15,16]. These data confirm those from other studies demonstrating the potent in vitro activity of ceftaroline against common aetiological agents of cSSSI, namely ␤-haemolytic streptococcal species and staphylococci, especially meticillin-resistant strains. Ceftaroline fosamil is unique among clinically available cephalosporins, having good in vitro activity against MRSA and MRCoNS as well as moderate activity against Gram-negative bacteria.

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Acknowledgments

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The authors would like to thank Regina Janes, John Dallow, Daniel Knight, Maria-Luisa Cassettari, Sarah Farrell, Angela Brook, Deanna Cooper, Edward Siegwart, Rosemary Maxfield, Mohammed Adil and Mikhail Caga-Anan from Quotient Bioresearch (Fordham, UK) for their technical support. The authors would also like to thank Dr Jane Ambler (AstraZeneca Pharmaceuticals) for assistance with the manuscript and helpful comments. Funding: This work was supported by educational/research grants, and the authors received compensation fees for services in relation to preparing the manuscript, which was funded by AstraZeneca Pharmaceuticals. Competing interests: None declared. Ethical approval: Not required.

[12]

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