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Comparison of the amino acid sequences of the VGSith that of S. pneumoniae R6 showed a change in ParCf Asn91Asp and in GyrA of Ser114Gly in all VGS anal-sed, independently of the ciprofloxacin MIC. These changesave been detected previously in VGS and in ciprofloxacin-esistant S. pneumoniae strains with mosaic structure in theiropoisomerase genes, suggesting that these strains have orig-nated by recombination with VGS [4].

All isolates with a ciprofloxacin MIC ≥8 mg/L had aminocid substitutions involving ParC (Ser79 → Phe/Tyr) andyrA (Ser81 → Phe/Tyr). One strain with a MIC of 4 mg/L

or ciprofloxacin had a Ser79 substitution in ParC. No muta-ions were detected in the seven strains with ciprofloxacin

ICs of 2 mg/L. Efflux was present in all isolates withiprofloxacin MICs of 4 mg/L (≥4-fold change). Efflux phe-otype was not present in isolates with ciprofloxacin MIC8 mg/L. Analysis of the QRDR of ParC and GyrA showed

hat low-level ciprofloxacin-resistant strains had mutationsltering the QRDR of topoisomerase IV and that high-evel ciprofloxacin-resistant strains had changes affectingRDRs both of ParC and GyrA. We found that reserpine-

nhibited efflux contributes primarily to a lower level ofiprofloxacin resistance and may do so either alone or tolesser extent by complementing QRDR substitutions in

arC. These results are in agreement with previous reports7,8].

Our results highlight the importance of accurate identifica-ion of VGS at species level, as differences in fluoroquinoloneusceptibility are observed.

Since first-step mutations and reserpine-inhibited effluxre predominantly found among VGS with a MIC ≥4 �g/mL,ur findings suggest that this MIC value should be consideredhe resistance breakpoint for this group of microorganisms.

VGS could act as donors in the horizontal transfer of fluo-oquinolone resistance genes to S. pneumoniae, constituting aeservoir for fluoroquinolone resistance. It is therefore impor-ant to undertake continued surveillance studies to monitorheir resistance status and its possible dissemination.

cknowledgment

This work was supported by grant FIS PI050344 from theondo de Investigacion Sanitaria, Madrid, Spain.

eferences

1] Gershon AS, de Azavedo JCS, McGeer A. Activities of new fluoro-quinolones, ketolides and other antimicrobials against blood cultureisolates of viridans group streptococci from across Canada. Antimicrob

Agents Chemother 2002;46:1553–6.

2] Diekema DJ, Beach ML, Pfaller MA, Jones RN. SENTRY ParticipantsGroup. Antimicrobial resistance in viridans group streptococci amongpatients with and without the diagnosis of cancer in the USA, Canadaand Latin America. Clin Microbiol Infect 2001;7:152–7.

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ntimicrobial Agents 29 (2007) 471–483

3] de Azavedo JCS, Trpeski L, Pong-Porter S, et al. In vitro activitiesof fluoroquinolones against antibiotic-resistant blood culture isolatesof viridans group streptococci from across Canada. Antimicrob AgentsChemother 1999;43:2299–301.

4] Balsalobre L, Ferrandiz MJ, Linares J, Tubau F, de la Campa AG.Viridans group streptococci are donors in horizontal transfer of topoi-somerase IV genes to Streptococcus pneumoniae. Antimicrob AgentsChemother 2003;47:2072–81.

5] Coykendall AL. Classification and identification of the viridans strepto-cocci. Clin Microbiol Rev 1989;2:315–28.

6] Clinical and Laboratory Standards Institute. Performance standardsfor antimicrobial susceptibility testing. 15th Informational Supplement.M100-S15. Wayne, PA: CLSI; 2005.

7] Gonzalez I, Georgiu M, Alcaide F, Balas D, Linares J, de la Campa AG.Fluoroquinolone resistance mutations in the parC, parE, and gyrA genesof clinical isolates of viridans group streptococci. Antimicrob AgentsChemother 1998;42:2792–8.

8] Munoz R, de la Campa AG. ParC subunit of DNA topoisomerase IV ofStreptococcus pneumoniae is a primary target of fluoroquinolones andcooperates with DNA gyrase A subunit in forming resistance phenotype.Antimicrob Agents Chemother 1996;40:2252–7.

9] Marron A, Carratala J, Alcaide F, Fernandez-Sevilla A, Gudiol F. Highrates of resistance to cephalosporins among viridans-group strepto-cocci causing bacteraemia in neutropenic cancer patients. J AntimicrobChemother 2001;47:87–91.

Iciar Rodrıguez-Avial ∗Carmen Rodrıguez-Avial

Esther CulebrasJuan J. Picazo

Servicio de Microbiologıa, Hospital Clınico San Carlos,Plaza de Cristo Rey s/n, 28040, Madrid, Spain

∗ Corresponding author. Tel.: +34 913 303 486;fax: +34 913 303 478.

E-mail address: iciaravial@hotmail.com(I. Rodrıguez-Avial)

oi:10.1016/j.ijantimicag.2006.12.006

aematological safety of long-term therapy with linezolid

Sir,Linezolid exhibits a broad spectrum of activity against

ram-positive bacteria, including methicillin-resistanttaphylococcus aureus (MRSA) and vancomycin-resistantnterococci (VRE).

The major side effect associated with linezolid administra-ion is reversible myelosuppression, mostly thrombocytope-ia, the risk of which appears to increase for treatment longerhan 28 days [1–4], ranging from 1 to 13%. Since in most clin-cal studies linezolid was administered for 2–4 weeks, littles known about the long-term incidence of haematologicalbnormalities. We analysed haematological and clinical dataf patients with Gram-positive infections receiving long-terminezolid therapy.

Outpatients treated for >4 weeks were prospectivelynalysed. Side effects, including haematological abnormal-ties, were recorded. Thrombocytopenia was defined as alatelet count <150 000 cells/�L or a reduction >25% from

Letters

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Editor

/InternationalJournalofAntim

icrobialAgents

29(2007)

471–483481

Table 1Demographics and haematological parameters of the study population

Patient no. Sex Age(years)

Disease Treatmentduration(weeks)

Pathogens Side effects Pre-/post-treatmentplatelet count (×103/mL)

Pre-/post-treatmentHgb (g%)

Pre-treatmentwith glycopeptides

1 M 35 Acute post-traumatic osteomyelitis(humerus)

16 MRSA – 265/274 12.8/12.5 Vancomycin

2 F 38 Acute post-traumatic osteomyelitis(radius)

7 MRSA Skin rash 273/234 14.9/14.6 –

3 M 39 Acute post-traumatic osteomyelitis(femur)

6 MRSA Skin rash 210/186 13.1/13.0 Teicoplanin

4 M 35 Acute post-traumatic osteomyelitis(femur)

14 MRSA – 231/219 13.8/13.3 Vancomycin

5 F 80 Acute post-traumatic osteomyelitis(radius)

6 MRSA Oral paraesthesia 234/187 11.3/11.1 –

6 F 40 Acute post-traumatic osteomyelitis(femur)

35 MRSA, Enterobactercloacae, Pseudomonasaeruginosa

– 261/246 12.7/12.2 Vancomycin

7 M 76 Chronic osteomyelitis (tibia) 10 MRSA – 233/199 14.2/12.9 –8 M 24 Chronic osteomyelitis (tibia) 12 MRSA – 206/165 15.5/14.8 Teicoplanin9 M 30 Chronic osteomyelitis (tibia) 5 MRSA – 209/193 16.1/15.9 Teicoplanin

10 M 77 Chronic osteomyelitis (tibia) 6 MRSA, Proteus mirabilis,P. aeruginosa

Oral cheilitis 203/170 13.3/12.9 –

11 M 34 Chronic osteomyelitis (humerus) 33 VRE – 234/257 11.3/11.6 Teicoplanin12 F 45 Chronic osteomyelitis (calcaneus) 16 MRSA, CoNS – 328/276 13.2/11.3 Teicoplanin13 M 50 Chronic osteomyelitis (femur) 13 MRSA – 296/266 12.0/13.4 Teicoplanin14 F 65 Chronic osteomyelitis (sternum) 5 MRSA Thrombocytopenia 204/94 14.7/12.8 Teicoplanin15 M 33 Septic arthritis (knee) 27 MSSA – 196/187 13.3/13.0 Vancomycin16 F 33 Septic arthritis (knee) 25 CoNS – 184/192 13.1/12.8 Vancomycin17 F 73 Total knee prosthesis infection 12 MRSA Nausea, vomiting 252/222 12.7/11.6 Teicoplanin18 M 63 Total knee prosthesis infection 12 MRSA – 259/212 15.8/15.4 Teicoplanin19 F 67 Total knee prosthesis infection 7 CoNS Thrombocytopenia 237/95 12.6/11.2 Teicoplanin20 F 62 Total knee prosthesis infection 15 CoNS – 371/282 12.5/11.4 Teicoplanin21 F 77 Total knee prosthesis infection 7 MRSA Nausea, vomiting 236/198 12.2/13.0 –22 F 72 Total hip prosthesis infection 7 MSSA Anaemia 313/245 12.4/9.8 Teicoplanin23 F 55 Total hip prosthesis infection 17 MSSA, Serratia

marcescens– 275/211 12.5/11.9 Teicoplanin

24 M 65 Total hip prosthesis infection 11 CoNS – 237/195 13.1/13.9 –25 F 47 Soft tissue infection 16 MRSA – 233/184 13.8/12.9 –26 M 62 Soft tissue infection 12 MRSA – 180/166 13.6/14.5 Teicoplanin27 F 66 Soft tissue infection 6 MRSA, P. aeruginosa – 306/356 12.8/12.0 Teicoplanin28 M 22 Soft tissue infection 9 MRSA Nausea, vomiting 203/198 12.1/11.9 –29 M 38 Maxillofacial prosthesis infection 6 MRSA – 249/189 12.4/12.1 –30 M 38 Maxillofacial prosthesis infection 7 MRSA, CoNS – 246/268 12.3/11.8 –31 F 27 Cerebral and pulmonary abscesses 37 MRSA – 303/231 12.7/12.9 –

Hgb, haemoglobin; MRSA, methicillin-resistant Staphylococcus aureus; VRE, vancomycin-resistant enterococci; CoNS, coagulase-negative staphylococci; MSSA: methicillin-sensitive S. aureus.

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82 Letters to the Editor / International Jour

aseline, and anaemia was defined either as haemoglobinHgb) levels <11.0 g% or Hgb reduction >25% fromaseline.

Bacteria were demonstrated by culture of drainage mate-ial or tissue samples. Species identification was performedsing the API system (bioMerieux, Marcy l’Etoile, France),nd antibiotic susceptibility was determined according to theuidelines of the Clinical and Laboratory Standards Institute5–8].

Thirty-one patients (16 male and 15 female; median age7 years) were studied. Twenty-six patients had an osteoar-icular infection, four had a soft tissue infection and one hadulmonary and cerebral abscesses (Table 1). Ten patients hadignificant co-morbidities.

All patients received oral linezolid 600 mg twice a day,xcept for four patients treated intravenously. The medianuration of treatment was 12 weeks (range 5–37 weeks).

MRSA was isolated in 23 patients (74%), of whom twoere also infected with coagulase-negative staphylococci

CoNS). Three patients were infected by methicillin-sensitivetaphylococcus aureus, four by CoNS only and one by VRE.

polymicrobial infection (i.e. infection both with Gram-egative and Gram-positive organisms) was documented inour cases.

Rifampicin was co-administered in three patients,nd treatment for a Gram-negative infection either withuinolones, �-lactams or amikacin was added in 12 patients.

Linezolid was well tolerated and no serious drug-relateddverse events leading to discontinuation were recorded.

The mean reductions from baseline in platelet countnd Hgb level were 14% and 3%, respectively. Twoatients developed thrombocytopenia: one had a mildhrombocytopenia (94 × 106 platelets/mL; baseline level04 × 106 platelets/mL) during the fifth and last week ofreatment; and the second had a reduction in platelet countrom 234 × 106/mL at baseline to 95 × 106/mL after 4 weeks,ut completed 6 weeks of treatment.

No significant changes in Hgb levels were recorded,xcept for one case of anaemia (Hgb = 9.8 g%) after 5 weeksf therapy (baseline level 12.4 g%), not requiring treatmentnterruption.

On Kaplan–Meyer analysis, the risk of developingnaemia or thrombocytopenia was higher between day 28 anday 35 of treatment, ranging from 3 to 10%, but cumulativencidence did not increase with longer administration.

Twenty patients (65%) received a glycopeptide (15eicoplanin and 5 vancomycin) in the 2 weeks precedinginezolid therapy. Both patients who developed throm-ocytopenia had previously received teicoplanin: baselinearameters and mean duration of teicoplanin treatmentere not significantly different compared with otheratients.

One patient suffered from perioral paraesthesia, one fromral cheilitis and two patients developed a non-itching skineaction on the face: all events resolved after treatment com-letion. No peripheral neuropathy was observed.

ntimicrobial Agents 29 (2007) 471–483

Linezolid is unique among new antibacterial agents owingo its oral bioavailability being almost equivalent to thatchieved intravenously and the excellent penetration intosteoarticular, pulmonary and soft tissues.

In phase III clinical studies, linezolid treatment >4 weeksas associated with a higher risk of reversible myelosuppres-

ion. Our study, although considering a small population,s one of the few reports of the haematological safety ofong-term linezolid administration: the incidence of throm-ocytopenia, similar to that previously reported [1,2], did notncrease with prolonged duration of treatment, suggestinghat the pathogenetic mechanism of thrombocytopenia, stillebated, is not related to the total amount of drug adminis-ered.

In critically ill patients with nosocomial pneumonia, theisk of developing thrombocytopenia (6.6%) was similar forinezolid and vancomycin [9]. A higher risk of thrombocy-openia was described in patients with orthopaedic infectionsreated with vancomycin within the 2 weeks before startinginezolid [10]. In our study, both patients with thrombocy-openia were previously treated with teicoplanin, supportinghe hypothesis that previous administration of a glycopeptideould represent a risk factor for linezolid-associated throm-ocytopenia.

Our study has two limitations. First, our patients were notritically ill and did not have oncohaematological diseaser renal failure, three categories with an increased risk ofhrombocytopenia. Second, the low rate of linezolid-relatednaemia may be accounted for by the low mean age of patientsnd the good levels of pre-treatment haemoglobin [11]. How-ver, our data demonstrate a good haematological tolerance ofrolonged linezolid administration in outpatients with a sat-sfactory immune status. Oral treatment with linezolid mayecome the drug of choice in outpatients requiring long-asting antimicrobial therapies. However, further studies areeeded to clarify the pathogenetic mechanism of linezolid-elated thrombocytopenia and to identify the patients atigher risk of myelosuppression.

eferences

[1] Attassi K, Hershberger E, Alam R, Zervos MJ. Thrombocytopeniaassociated with linezolid therapy. Clin Infect Dis 2002;34:695–8.

[2] Birmingham MC, Rayner CR, Meagher AK, Flavin SM, Batts DH,Schentag JJ. Linezolid for the treatment of multidrug-resistant, Gram-positive infections: experience from a compassionate-use program. ClinInfect Dis 2003;36:159–68.

[3] Gerson S, Kaplan S, Bruss J, et al. Hematologic effects of line-zolid: summary of clinical experience. Antimicrob Agents Chemother2002;46:2723–6.

[4] Waldrep TW, Skiest DJ. Linezolid-induced anaemia and thrombocy-topenia. Pharmacotherapy 2002;22:109–12.

[5] Clinical and Laboratory Standards Institute. Performance standards for

antimicrobial disk susceptibility tests, 9th ed. Approved standard. M02-A9; Wayne, PA: CLSI, 2006.

[6] Clinical and Laboratory Standards Institute. Performance standardsfor antimicrobial susceptibility testing. Sixteenth informational sup-plement. M100-S16. Wayne, PA: CLSI, 2006.

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E-mail address: silvia.garazzino@inwind.it

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[7] Clinical and Laboratory Standards Institute. Methods for dilutionantimicrobial susceptibility tests for bacteria that grow aerobi-cally, 7th ed. Approved standard. M07-A7. Wayne, PA: CLSI,2006.

[8] Clinical and Laboratory Standards Institute. Methods for antimicrobialsusceptibility testing of anaerobic bacteria, 6th ed. Approved standard.M11-A6. Wayne, PA: CLSI, 2006.

[9] Nasraway SA, Shorr AF, Kuter DJ, O’Grady N, Le VH,Cammarata SK. Linezolid does not increase the risk of throm-bocytopenia in patients with nosocomial pneumonia: comparativeanalysis of linezolid and vancomycin use. Clin Infect Dis 2003;11:1609–16.

10] Rao N, Ziran BH, Wagener MM, Santa ER, Yu VL. Similar hematologiceffects of long-term linezolid and vancomycin therapy in a prospectiveobservational study of patients with orthopedic infections. Clin Infect

Dis 2004;38:1058–64.

11] Senneville E, Legout L, Valette M, et al. Risk factors for anaemiain patients on prolonged linezolid therapy for chronic osteomyeli-tis: a case-control study. J Antimicrob Chemother 2004;54:798–802.

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ntimicrobial Agents 29 (2007) 471–483 483

Silvia Garazzino ∗Francesco Giuseppe De Rosa

Olivia BargiacchiSabrina Audagnotto

Agostino MaielloGiovanni Di Perri

University of Turin, Department of Infectious Diseases,Ospedale Amedeo di Savoia, Corso Svizzera 164,

10149 Turin, Italy

∗ Corresponding author. Tel.: +39 011 439 3926;fax: +39 011 439 3977.

(S. Garazzino)

oi:10.1016/j.ijantimicag.2006.12.007