Methicillin-resistant Staphylococcus aureus isolated from an intensive care unit in Minas Gerais, Brazil, over a six-year period

w w w . e l s e v i e r . c o m / l o c a t e / b j i d

The

Brazilian

Journal

of

INFECTIOUS

DISEASES

Brief

communication

Methicillin-resistant

Staphylococcus

aureus

isolated

from

an

intensive

care

unit

in

Minas

Gerais,

Brazil,

over

a

six-year

period

Thiago

C.

Nascimento

_,

_Cláudio

_G.

_Diniz

_,

_Vânia

_L.

_Silva

_,

Alessandra

B.

Ferreira-Machado

_,

_Marina

_O.

_Fajardo

_,

_Tamara

_Lopes

_R.

_de

_Oliveira

_,

Dennis

de

C.

Ferreira

_,

_Fernanda

_S.

_Cavalcante

_,

_Kátia

_R.

_Netto

_dos

_Santos

a_{UniversidadeFederaldeJuizdeFora,DepartamentodeParasitologia,MicrobiologiaeImunologia,JuizdeFora,MG,Brazil} b_{UniversidadeFederaldoTriânguloMineiro,DepartamentodeMicrobiologia,ImunologiaeParasitologia,Uberaba,MG,Brazil} c_{UniversidadeFederaldoRiodeJaneiro,InstitutodeMicrobiologiaPaulodeGóes,RiodeJaneiro,RJ,Brazil}

d_{UniversidadeVeigadeAlmeida,RiodeJaneiro,RJ,Brazil} e_{UniversidadeEstáciodeSá,RiodeJaneiro,RJ,Brazil} f_{UniversidadeFederaldoRiodeJaneiro,Macaé,RJ,Brazil}

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Articlehistory:

Received30August2017 Accepted20October2017

Availableonline21November2017

Keywords: Clonalcomplex MRSA PFGElineages SCCmec

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Tocharacterizemethicillin-resistantStaphylococcusaureusisolatesfromanintensivecare unitofatertiary-careteachinghospital,between2005and2010.Atotalof45isolateswere recoveredfrompatientsadmittedtotheintensivecareunitinthestudyperiod.Resistance rateshigherthan80%werefoundforclindamycin(100%),erythromycin(100%),levofloxacin (100%),azithromycin(97.7%),rifampin(88.8%),andgentamycin(86.6%).TheSCCmectyping revealedthattheisolatesharboredthetypesIII(66.7%),II(17.8%),IV(4.4%),andI(2.2%).Four (8.9%)isolatescarriednon-typeablecassettes.Most(66.7%)oftheisolateswererelatedtothe BrazilianendemicclonefromCC8/SCCmecIII,whichwasprevalent(89.3%)between2005and 2007,whiletheUSA100/CC5/SCCmecIIlineageemergedin2007andwasmorefrequentinthe lastfewyears.Thestudyshowedhighratesofantimicrobialresistanceamong methicillin-resistantS.aureusisolatesandthereplacementofBrazilianclone,awell-establishedhospital lineage,bytheUSA100inthelate2000s,attheintensivecareunitunderstudy.

©2017SociedadeBrasileiradeInfectologia.PublishedbyElsevierEditoraLtda.Thisisan openaccessarticleundertheCCBY-NC-NDlicense(http://creativecommons.org/licenses/ by-nc-nd/4.0/).

Staphylococcusaureusisoneofthemaincausesof healthcare-associatedinfections.1_{Methicillin-resistantS.aureus(MRSA)}

isagrowingproblemworldwideandisassociatedwith signifi-cantmorbidity,mortalityandincreasedcostsoftreatments.2,3

∗ _{Correspondingauthor.}

E-mailaddress:[email protected](K.R.Santos).

The majorityof MRSAisolatesare foundin intensivecare units(ICUs).4_{InBrazil,theserateshaveevenreached70%.}5

Methicillinresistanceislocatedinastaphylococcalcassette chromosome(SCCmec)andthemostfrequenttypesareII,III andIV.2

RecentstudieshavedescribedtheemergencyofMRSA lin-eagesinhospitalsworldwide.6,7 _{InBrazil,althoughisolates}

relatedtotheBrazilianendemicclone(BEC)/SCCmecIII/CC8

https://doi.org/10.1016/j.bjid.2017.10.004

1413-8670/©2017SociedadeBrasileiradeInfectologia.PublishedbyElsevierEditoraLtda.ThisisanopenaccessarticleundertheCC BY-NC-NDlicense(http://creativecommons.org/licenses/by-nc-nd/4.0/).

havecausedthemajorityofhospital-acquiredstaphylococcal infectionsinthepast,8_{duringthelastdecadeanincreased}

occurrenceofnosocomialinfectionsduetoisolatescarrying theSCCmecIVandIIhavebeendescribed.9–11_The

character-izationofMRSA isolatesfrompatients inICUshasalready beendescribedbyvariousauthors.9,11 _{However,these}

stud-iesonlyanalyzedafewICUisolateswithinalargerhospital collection,withouthighlightingthecharacterizationofthese isolates.Thisstudyinvestigatedthephenotypicand molecu-larcharacteristicsofacollectionofMRSAclinicalisolatesfrom patientsadmittedtoaBrazilianICUoverasix-yearperiod.

Thestudywasconductedatatertiary-careteaching hospi-talaffiliatedtotheFederalUniversityofJuizdeFora,Minas Gerais, Brazil. This is a 146-bed, six of them being ICU. TheMRSAisolateswereobtainedfrompatientsadmittedto theICU consecutively, between2005and2010, from differ-entsources,suchastrachealsecretion(53.3%),blood(20%), cathetertip(13.4%),and others(13.3%).Bacterial identifica-tionandsusceptibilitytomethicillinweredeterminedatthe hospitallaboratorybytheclassicalidentificationtests12_and

the cefoxitin(Oxoid,Basingstoke, UK)disk diffusiontest,13

respectively. Minimum inhibitory concentration (MIC) was assessedfor11drugs asrecommendedbythe CLSI.13

Bac-terialDNAwasextractedaspreviouslydescribed14 _andthe

determinationoftheSCCmectypeswasperformedaccording toMilheiric¸oetal.15_{AllMRSAisolatesweretypedbyPFGE}8

andtheclonalitywasdeterminedaccordingtoVanBelkum etal.16criteriausingpreviouslycharacterizedcontrolstrains, suchas:USA100/SCCmecII,USA400andUSA800/SCCmecIV9

andBEC/SCCmecIII.8_{OneisolaterepresentativeofeachPFGE}

profilewaschosenforcharacterizationoftheclonalcomplex (CC).17_{Statisticalcomparisonswereperformedbyanalysisof}

contingencytablesusingFisher’s exacttest;levelof signifi-cancewasestablishedat5%(p<0.05).

Out of 76 S. aureus recovered from patients admitted to the ICU, between January/2005 and November/2010, 45 (59.2%)MRSAisolatesrecoveredfrom45patientswere eval-uated.Exceptforvancomycinandlinezolid,whoseMIC90was

2.0␮g/mL,highratesofresistancewerefoundforsevenofthe 11antimicrobialstested(Table1).Amongthe45MRSAisolates,

30(66.7%)harboredtheSCCmecIII,8(17.8%)thetypeII,2(4.4%) thetypeIVand1(2.2%)thetypeI.Four(8.9%)MRSAisolates werenontypeable(NT).S.aureusisolatescarryingSCCmecIII were relatedtotheBEC/clonalcomplex (CC)8(Table2).All theeightisolatesthatcarriedtheSCCmecIIwere relatedto theUSA100/CC5lineage.TwoisolatescarryingtheSCCmecIV belongedtothelineagesUSA400/CC1andUSA800/CC5.The SCCmecIisolatewasassociatedtoUSA500/CC5andtheother fourMRSAisolatesdidnotbelongtoanyclonalitypreviously described(Table2).

Brazilian studies have evaluated the epidemiology of MRSA and the results indicate that several lineages ini-tiallyrestrictedtoothercontinentsareemerginginBrazilian hospitals.9–11 _{This study aimed to analyze the phenotypic}

andmolecularcharacteristicsofacollectionofMRSAisolates obtainedexclusivelyfrom patientsadmittedtoanICU and verified the replacementand emergence oflineagesin the periodunderinvestigation.Initiallytherewasahigh dissemi-nationofBEC/CC8/SCCmecIIIisolatesfrom2005to2007,with aprevalenceofthiscloneof89.3%amongtheisolates.The USA100/CC5/SCCmecIIlineageemergedin2007andwasmore frequentin2009and2010,whilesporadiclineagesoccurredin 2008.

The BEC, a well-established lineage in Brazilian hospi-tals,representingabout90%ofthenosocomialMRSAisolates in the late 1990s8 _{has been replaced inrecent decades by}

SCCmec IV and II carrying MRSAisolates.9–11 _{A study also}

conductedinanICUfromMinasGeraisevaluated36MRSA isolatedin2009andfoundthat58.3%ofisolatescarriedthe SCCmec II.18 _{In Rio de Janeiro, a study performed by our}

groupintwohospitals,between2004and2007,showedthat about50%ofMRSAisolateswererelatedtotheBEC/SCCmec III lineage, whileabout 35% ofisolates carried the SCCmec II or IV.9 _{In another study conducted by our group in a}

teaching hospital, between 2005and 2006, the majority of isolates carried the cassette type IV (49%) and BEC iso-lates accounted for 49% of them among 83 nasal MRSA isolates analyzed,11 _{confirming the replacing of this}

lin-eage forothers in the years 2000,as foundin the present study.

Table1–Antimicrobialresistanceof45MRSAisolatesrecoveredfrompatientsofanICUataMinasGeraisteaching

hospital,between2005and2010.

Antimicrobialagent MinimalInhibitoryConcentration(␮g/mL) No(%)ofresistant

isolates

MIC50 MIC90 Range

Azithromycin >1024.0 >1024.0 0.5–>1024.0 44(97.7) Chloramphenicol 32.0 64.0 4.0–128.0 29(64.4) Clindamycin >1024.0 >1024.0 512.0–>1024.0 45(100) Erythromycin 512.0 512.0 256.0–512.0 45(100) Gentamicin 128.0 1024.0 0.125–>1024.0 39(86.6) Levofloxacin 4.0 16.0 2.0–32.0 45(100) Linezolid 2.0 2.0 1.0–2.0 0(0) Rifampin 2.0 256.0 0.0625–>1024.0 40(88.8) Tetracycline 32.0 64.0 0.0625–128.0 31(68.8) Trimethoprim/sulfamethoxazole 32.0/608.0 128.0/2432.0 0.0625/2.3–1024.0/19,456.0 32(71.1) Vancomycin 1.0 2.0 0.5–2.0 0(0)

MIC50,minimalinhibitoryconcentrationthatinhibits50%ofbacterialpopulation;MIC90,minimalinhibitoryconcentrationthatinhibits90%of bacterialpopulation.

Table2–Characteristicsof45MRSAisolatedfrompatientsadmittedtoanICUofaMinasGeraisteachinghospital, between2005and2010. Isolate number Isolation date (mm/dd/yy) Bed Age (years) Clinical source Comorbidity SCCmec type Clonalitya_{/CC Antimicrobial} resistance profile

1 01/11/05 04 77 Blood Bladdercancer III BEC/8 ACEGLRST

2 01/18/05 01 79 TS Stomachcancer III BEC/8 ACEGLRST

3 03/08/05 03 35 Urine Respiratoryfailure III BEC/8 AHCEGLRST

4 05/17/05 03 78 TS Pneumonia III BEC/8 AHCEGLRST

5 07/18/05 01 69 Blood Bladdercancer III BEC/8 AHCEGLRST

6 07/24/05 02 40 TS Respiratoryfailure III BEC/8 ACEGLRST

7 07/25/05 01 48 TS Incarceratedhernia III BEC/8 ACEGLRST

8 08/31/05 04 25 PL Ascitis NT ND/ND ACEGLRST

9 09/08/05 03 69 Blood Coronaryarterydisease III BEC/8 AHCEGLRST

10 09/06/06 05 75 CT Fahrdisease III BEC/8 ACEGLRST

11 09/23/06 02 66 TS COPD III BEC/8 ACEGLRST

12 10/09/06 02 60 TS COPD III BEC/8 AHCEGLRST

13 12/06/06 03 81 Blood COPD III BEC/8 ACEGLRST

14 12/09/06 04 23 TS Acuteintermittentporphyria III BEC/8 AHCEGLRST

15 12/11/06 05 69 TS Chronickidneydisease III BEC/8 AHCEGLRST

16 12/18/06 03 53 CT Rheumatoidarthritis III BEC/8 AHCEGLRST

17 01/30/07 01 53 TS Rheumatoidarthritis III BEC/8 AHCEGLRST

18 02/04/07 01 56 TS Neurogenicbladder II USA100/5 AHCEGLR

19 02/28/07 01 41 PL Acutepancreatitis III BEC/8 AHCEGLRST

20 03/25/07 02 76 TS Cerebrovascularaccident III BEC/8 AHCEGLRST

21 04/02/07 04 54 Blood Hodgkinlymphoma II USA100/5 ACEGLR

22 06/25/07 03 61 CT Hemothorax III BEC/8 AHCEGLRST

23 07/08/07 04 42 SS Pancreatitis III BEC/8 AHCEGLRST

24 09/10/07 06 35 TS Aids III BEC/8 AHCEGLRST

25 09/18/07 03 74 TS Extrapontinemyelinolysis III BEC/8 AHCEGLRST

26 12/04/07 04 73 Blood Bladdercancer III BEC/8 AHCEGLRST

27 12/24/07 01 51 PL Cirrhosis III BEC/8 AHCEGLRST

28 12/31/07 01 73 Blood Malnutrition III BEC/8 ACEGLRST

29 01/14/08 02 23 CT Diabetesmellitus IV USA800/5 CEGLT

30 01/30/08 03 65 TS Chronickidneydisease III BEC/8 AHCELRST

31 02/03/08 02 84 TS Respiratoryfailure IV USA400/1 AHCEGL

32 02/07/08 06 59 CT Cirrhosis III BEC/8 AHCEGLRST

33 03/16/08 03 81 CT Acutekidneyinjury NT ND ACELS

34 08/16/08 06 61 Blood Myocardialinfarction NT ND ACEGLR

35 10/13/08 01 65 SS Appendicitis I USA500/5 ACEGL

36 07/18/09 03 42 TS Renaltransplantation NT ND AHCELR

37 08/03/09 05 19 TS Pneumonia II USA100/5 AHCELR

38 08/09/09 01 78 TS Breastcancer II USA100/5 AHCEGLR

39 08/17/09 06 79 TS COPD II USA100/5 AHCEGLR

40 12/14/09 01 83 TS Cholangitis III BEC/8 AHCEGLRST

41 02/06/10 02 44 TS Acutekidneyinjury II USA100/5 AHCEGLR

42 02/21/10 04 98 TS Aplasticanemia III BEC/8 AHCEGLRS

43 04/18/10 06 35 TS Aids III BEC/8 AHCEGLRST

44 08/25/10 06 59 Blood Necrosisinamputation II USA100/5 ACEL

45 11/30/10 04 62 TS Acutekidneyinjury II USA100/5 ACELR

CT,catetertip;PL,peritonealliquid;SS,surgicalsite;TS,trachealsecretion;COPD,chronicobstructivepulmonar;CC,clonalcomplex;NT, nontypeable;ND,notdetermined;A,Azithromycin;C,Clindamycin;E,Erythromycin;G,Gentamicin;H,Chloramphenicol;L,Levofloxacin;R, Rifampin;S,Trimethoprim/sulfamethoxazole;T,Tetracycline.

a _{AccordingtoCockfieldetal.}17

MRSA isolates carrying SCCmec II represented 17.8% of all ICU isolatesin the present study, and belongedto the USA100/CC5,a lineagevery common in USAhospitals.7 _In

2007,thislineageemergedinourICUandwasprevalent(60%) in 2009 and 2010, replacing the BEC. Caiaffa-Filho et al.10

evaluated50consecutivebloodMRSAisolatesduringa three-monthperiodin2010atahospitalinSãoPauloanddetected 60%carrying the SCCmec II,and 83% ofthemwere related

to the USA100 lineage. Chamon et al.19 _{recently evaluated}

a collection of 45 MRSA isolates from bloodstream infec-tions(BSI)obtainedattwodifferentpublichospitalsinRiode Janeirocity,between2008and2009.Theauthorsshowedthe complete substitutionofthe BEC/SCCmecIIIandthe preva-lenceofUSA100/SCCmecIIisolatesinthelate2000s.Similar results were found inthe present study showing the pre-dominance of USA100/SCCmec II between 2009 and 2010,

changing the epidemiological profile of MRSA in the ICU evaluated.

Inthepresentstudy,MRSAisolatespresentedhighrates of resistance over half of the evaluated antimicrobials. In general, isolatesbelonging to USA100/SCCmec typesII and BEC/typeIIIlineagepresenthighresistanceratesfor antimi-crobials unlike the type IV isolates.9,11,19 _{Multiresistance}

among isolates of these lineages could explain the abil-ity of them to persistin the hospital environment. While theresistanceratesfortrimethoprim/sulfamethoxazoleand tetracyclinewere100%and96.7%,respectivelyfortypeIII,all typeIIisolatesweresusceptibletobothdrugs(p<0.0001)(data notshown),afactalsoobservedbyCavalcanteetal.,20_who

proposedtousetheseantimicrobialsasmarkerstodistinguish MRSAisolates.

AlimitationofthisstudywasthenumberofMRSAisolates evaluatedsincethe ICUunderstudy hasonlysixbedsand becauseseveralclinicalstrainsisolatedduringthestudywere Gramnegativebacteria.Moreover,themodeofacquisitionof theisolateswasnotdescribed,althoughthemajorityofthe isolateswereofnosocomialorigin.

Our results showed that MRSA isolates from patients admitted to an ICU of a teaching hospital showed high ratesofresistanceoverhalfoftheevaluatedantimicrobials. Moreover,there was prevalence ofthe BEC/CC8/SCCmecIII lineage between 2005 and 2007 and the emergency ofthe USA100/CC5/SCCmecIIlineagein2007,whichwasmost fre-quentinthelate2000s.

Conflicts

of

interest

Theauthorsdeclarenoconflictsofinterest.

Acknowledgments

TheauthorsaregratefultotheProgramadePós-Graduac¸ão emSaúde–UniversidadeFederaldeJuizdeFora(PPGS/UFJF), Fundac¸ãodeAmparoàPesquisadeMinasGerais(FAPEMIG), Fundac¸ãodeAmparo àPesquisadoRiodeJaneiro(FAPERJ) andConselhoNacionaldeDesenvolvimentoCientíficoe Tec-nológico(CNPq)forfinancialsupport.Theauthors are also gratefultostafffromtheLaboratoryProf.MaurilioBaldi,and SuzaneF.Silva,PedroP.Castro,DéboraM.Coelho,for techni-calhelpwiththemedicalrecordsandrecordbooksfromthe clinicalmicrobiologylaboratory.

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