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Publié par | biomed |
Publié le | 01 janvier 2012 |
Nombre de lectures | 12 |
Langue | English |
Extrait
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RESEARCH
OpenAccess
Clinical,epidemiologicalandmolecularfeatures
oftheHIV-1subtypeCandrecombinantforms
thatarecirculatinginthecityofSãoPaulo,Brazil
RosanaAlcalde
1*
†
,MonickLGuimarães
3
†
,AlbertoJSDuarte
1
andJorgeCasseb
1,2*
Abstract
Background:
ThecityofSaoPaulohasthehighestAIDScaserate,withnearly60%inBrazil.Despite,several
studiesinvolvingmolecularepidemiology,lackofdataregardingalargecohortstudyhasnotbeenpublishedfrom
thiscity.
Objectives:
ThisstudyaimedtodescribetheHIV-1subtypes,recombinantformsanddrugresistancemutations,
accordingtosubtype,withemphasisonsubtypeCandBCrecombinantsinthecityofSãoPaulo,Brazil.
Studydesign:
RNAwasextractedfromtheplasmasamplesof302HIV-1-seropositivesubjects,ofwhich211were
drug-naiveand82wereexposedtoART.HIV-1partial
pol
regionsequenceswereusedinphylogeneticanalysesfor
subtypingandidentificationofdrugresistancemutations.TheenvelopegeneofsubtypeCandBCsampleswas
alsosequenced.
Results:
Frompartial
pol
geneanalyses,239samples(79.1%)wereassignedassubtypeB,23(7.6%)wereF1,
16(5.3%)weresubtypeCand24(8%)weremosaics(3CRF28/CRF29-like).ThesubtypeCandBCrecombinants
weremainlyidentifiedindrug-naïvepatients(72.7%)andtheheterosexualriskexposurecategory(86.3%),whereas
forsubtypeB,thesevalueswere69.9%and57.3%,respectively(p=0.97andp=0.015,respectively).Anincreasing
trendofsubtypeCandBCrecombinantswasobserved(p<0.01).
Conclusion:
TheHIV-1subtypeCandCRFsseemtohaveemergedoverthelastfewyearsinthecityofSãoPaulo,
principallyamongtheheterosexualpopulation.Thesefindingsmayhaveanimpactonpreventivemeasuresand
vaccinedevelopmentinBrazil.
Keywords:
HIV-1,Subtypes,Recombinants,Resistance,SãoPaulo,Brazil
Background
genotypingsystems[1-3].Thus,HIV-1subtypesalso
ThehugegeneticvariabilityofHIV-1resultsinacom-contributetothecapacityofHIV-1toevadethehost
plexanddynamicmolecularclassificationoftypes(HIV-immuneresponse,[4]whichcanaffecttheresponseto
1and2),groups(M,N,O,P),andthepandemicgroupMantiretroviraltreatmentand,consequently,totheemer-
couldbedividedintosubtypes(A-D,F-H,J,K)andrecom-genceofdrugresistance[5].Somestudieshavesug-
binantforms,suchascirculatingrecombinantformsgestedthatcoreceptorswitchingfromCCR5toCXCR4
(CRF)anduniquerecombinantforms(URF).SuchHIV-islesscommoninHIV-1subtypeC,[6]showingalower
1variationhasanimportantimpactondiagnosis,viralrateofaccumulationofmutationsthatconferresistance
loadmeasurementandtheperformanceofHIV-1thansubtypeB[5].Concerningthereplicationfitnessof
subtypeC,thestudieswerecontroversial,[7-9]andin
*Correspondence:rosana117@yahoo.com;jcasseb@usp.br
relationtotransmissioninutero,thissubtypepresented
†
Equalcontributors
1
LaboratoryofDermatologyandImmunodeficiencies,Departmentof
highefficiencycomparedtosubtypesAorD[8,10,11].
Dermatology,MedicalSchoolofSãoPauloUniversity,LIM56/FMUSP.Av.Dr.
Thesepropertiessuggestanoveralltransmissionadvan-
EneasdeCarvalhoAguiar,470,IMTII,3°andar,05403-000SãoPaulo,SP,
tage,inpart,possiblyrelatedtoefficientreplicationin
Brazil
Fulllistofauthorinformationisavailableattheendofthearticle
©2012Alcaldeetal.;licenseeBioMedCentralLtd.ThisisanOpenAccessarticledistributedunderthetermsoftheCreative
CommonsAttributionLicense(http://creativecommons.org/licenses/by/2.0),whichpermitsunrestricteduse,distribution,and
reproductioninanymedium,providedtheoriginalworkisproperlycited.
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dendriticcells,thetargetsoftheonsetofHIV-1infec-
tion[11].
AlthoughHIV-1subtypeBisthemoststudied,HIV-1
Cpredominatesgloballyandisresponsibleforapproxi-
mately50%ofinfections[12].Thissubtypeisnotwide-
spread,butitisthemostprevalentinsub-Saharan
Africaandinthepopulouscountries,suchasIndiaand
China,whereHIVinfectionratesarehighestamonghet-
erosexuals[12,13].Uptonow,fivecirculatingrecombin-
antforms(CRFs)involvingthissubtypehavebeen
detected,threeofthempresentingrecombinationsbe-
tweensubtypesBandC,CRF07,CRF08,describedin
China,[14,15]andCRF31_BC,describedinBrazil[16].
Theothertwopresentrecombinationsbetweensubtypes
CandD,CRF10_CDandCRF41_CD[17,18].Overall,
morethan18%ofnewinfectionshavebeenattributedto
HIV-1recombinants[19].
TheBrazilianAIDSepidemicismainlydrivenby
subtypesB,F1,andBF1recombinants;however,inthe
Southernregion,subtypeCandBCrecombinantscan
representupto50%ofcases,dependingofthegeo-
graphicalstudiedregion[20-27].Despitethedocumen-
tedearlycirculationofHIV-1subtypeCoutsideofthe
southernregionin1992,onecasefromthecityofSão
Paulo[28]andanotherin1986inSantos,[29]only
recentlyhasacontinuousincreaseincasesbeen
observed[30-32].
ThecurrentstudywasconductedtodetermineHIV-1
geneticdiversityandPI/RTIresistanceassociatedmuta-
tionsamongHIV-1infecteddrugnaïveandHAART
patientsinacohortinthecityofSãoPaulo.Moreover,
patientsclassifiedasHIV-1subtypeCandBCrecombi-
nantswereanalyzedinmoredetails.
Objectives
ThepresentstudyaimedtodescribeHIV-1subtypes,
CRFsanddrugresistancemutationsindrug-naïveand
failingHAARTindividualslivinginthecityofSão
Paulo,inparticularattentiontothoseclassifiedassub-
typeCandBC.
Studydesign
Studypopulation
ThestudypopulationwascomposedbyHIV-1-infected
patientswhohavebeenfollowed-upattheAmbulatory
ServiceoftheDepartmentofSecondaryImmunodeficiency
ClinicoftheClinicalHospital,UniversityofSãoPaulo
MedicalSchool(HC/FMUSP),SãoPaulo,SP,Brazil,oneof
thelargestteachingandresearchhospitalsinBrazil.
AccordingtotheBrazilianMinistryofHealth,HIV
genotypingtestshouldberoutinelyperformedin
patientsunderantiretroviraltherapywhopresenta
plasmaviralloadover2000copies/mL[33].Following
thisdirective,atotalof82patientsweregenotypedat
Page2of10
HC/FMUSPfromMarch2002toJune2010.Duringthe
sameperiod,211drug-naiveindividualsandnineindivi-
dualswhohadunknownstatusinrelationtoantiretro-
viraltherapy(ART)werealsogenotyped.Demographic
andclinicaldatawereobtainedfromclinicalchartsor
directinterviewandthepatientsindividuallyallowus
forpublication,asshownatTable1.Thisstudyprotocol
wasapprovedbytheResearchEthicsCommitteeofHC/
FMUSP,underprotocolnumber774/99,andwritten
informedconsentwasobtainedfromallpatients.During
+thefollow-up,CD4TcellsandHIV-1viralloadwere
determinedusingtheBrazilianMinistryofHealth
guidelines.
RNAisolation,RT-PCRandDNAsequencing
HIV-1viralRNAwasisolatedfromplasmasamples
usingtheQIAamp
W
ViralRNAMiniKit(Qiagen,Hilden,
Germany)extractionmethod,inaccordancewiththe
manufacturer
’
sprotocol.ComplementaryDNAwasper-
formedbyRT-PCRprotocolwithrandomprimersand
usingthePR/RTPCRstrategypreviouslydescribedby
Gonzalesetal.[34,35].ThePCRproductswerepuri-
fiedusingQIAquick
W
(Qiagen,Hilden,Germany),in
accordancewiththemanufacturer
’
sprotocol.ThePCR
purifiedproductsweresequencedutilizingABIPrismBig
DyeTerminatorReadyReactionKitversion3.0(Applied
Biosystems
W
,FosterCity,CA),inaccordancewiththeirre-
spectiveprotocols.Thereactionswereanalyzedusingthe
ABIPrism3100GeneticAnalyzer(AppliedBiosystems
W
,
FosterCity,CA).
Sequenceanalysis
ThechromatogramsofallsequencedDNAwerevisu-
allyusingtheSequencherprogramversion4.0.5(Gene
Codes)andmanuallyedited.PR/RTsequencingwas
conductedtodetectdrugresistancemutationsand
HIV-1subtypes.Majorantiretroviraldrugresistance
mutationswereclassifiedaccordingtoStanfordUni-
versityonlineHIVDrugResistanceDatabase,[36].
TheInternationalAIDSSociety-USAPanelandUp-
dateoftheDrugResistanceMutationsinHIV-1,
Spring2008[37-39].
NucleotidesequenceswerealignedusingtheClustalX
program[40]andlaterhandeditedforminoradjust-
mentsandgap-stripped.Analignmentof860bpthat
partiallycoveredthePR/RTregion(nucleotides2343-
3203)wasusedforphylogeneticinferencesunder
Neighbor-Joining(NJ)algorithminMega4.0.2program
[41].Inordertoconfirmtheclassificationofthenon-B
sequences,Bayesiananalysiswasperformed.ThejMo-
deltest0.1.1program[42]wasusedtoselectthebest-fit
modelofnucleotidesubstitutionundertheAkaikeinfor-
mationcriteria,resultinginthechoiceoftheGTR+I+
Gmodel,[43]asimplementedinMrBayesv3.1.2[44].
Table1Epidemiological,virological,immunologicalcharacteristicsandresistanceassociatedmutationsofPR/RTregionfrompatientsclassifiedasHIV-1
I.D.LIM56DateAgeGenderVLCD4TransmissionMutationsMutationsotherMutationsMutationsMutationsother(RT)
collection(years)modeProtease(Protease)(NRTI)(NNRTI)
orinmA-subtypeC
0122/09/200348
0225/09/200363
0318/04/200531
0423/05/200522
5006708009
0111
21
31
14
11/08/200537
13/09/200537
10/04/200639
11/05/200644
03/08/200623
09/04/200725
4/5/200930
4/5/200929
8/4/201031
1