Seroprevalence of hepatitis E virus in chronic hepatitis C in Brazil

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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

Original

article

Seroprevalence

of

hepatitis

E

virus

in

chronic

hepatitis

C

in

Brazil

Guilherme

Bricks

a,∗

_,

_Jorge

_Figueiredo

_Senise

a

_,

_Henrique

_Pott

_Junior

a

_,

_Giuliano

_Grandi

a

_,

Amanda

Passarini

a

_,

_Débora

_Bellini

_Caldeira

a

_,

_Dimas

_Carnaúba

_Junior

b

_,

Hamilton

Antonio

Bonilha

de

Moraes

c

_,

_Celso

_Franscisco

_Hernandes

_Granato

a

_,

Adauto

Castelo

a

a_Universidade_Federal_de_São_Paulo,_Departamento_de_Medicina,_Divisão_de_Doenças_Infecciosas,_São_Paulo,_SP,_Brazil b_Hospital_de_Transplante_Euryclides_Jesus_Zerbini,_Ambulatório_de_Doenças_Infecciosas,_São_Paulo,_SP,_Brazil c_Instituto_de_Vacinação_e_Infectologia_de_Piracicaba,_Piracicaba,_SP,_Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory:

Received28September2017

Accepted2February2018

Availableonline27February2018

Keywords: HepatitisEvirus HepatitisCvirus Brazil Seroprevalence

a

b

s

t

r

a

c

t

Backgroundandaims:HepatitisEvirusinfectioninpatientswithunderlyingchronicliver

diseaseisassociatedwithliverdecompensationandincreasedlethality.Theseroprevalence

ofhepatitisEvirusinpatientswithchronichepatitisCinBrazilisunknown.Thisstudyaims

toestimatetheseroprevalenceofhepatitisEvirusinpatientswithchronichepatitisCand

todescribeassociatedriskfactors.

Methods:Atotalof618patientschronicallyinfectedwithhepatitisCvirusfromthree

refer-encecentersofSãoPaulo,Brazilwereincluded.Presenceofanti-HEVIgGwasassessedby

enzyme-linkedimmunosorbentassay(WANTAIHEV-IgGELISA).

Results:Outofthe618patientstested,10.2%turnedoutpositiveforanti-HEVIgG(95%CI

8.0–12.8%).Higherseroprevalencewasfoundindependentlyassociatedwithageover60

years(OR=2.04;p=0.02)andpreviouscontactwithpigs(OR=1.99;p=0.03).

Conclusions: PatientswithchronichepatitisCareunderriskofhepatitisEvirus

superinfec-tioninSãoPaulo.Contactwithpigsisariskfactorfortheinfection,suggestingapossible

zoonosiswithoraltransmission.

openaccessarticleundertheCCBY-NC-NDlicense(http://creativecommons.org/licenses/

by-nc-nd/4.0/).

Introduction

HepatitisEvirus(HEV)isthemostcommonhepatotropicvirus

intheworld.1–3_The_World_Health_Organization_estimates_the

∗ _{Corresponding}_author.

E-mailaddress:guibricks@gmail.com(G.Bricks).

occurrenceof20millioninfectionsperyearworldwide,with

around3.3millionsymptomaticinfections,and44,000deaths

in2015.4

HEV includes four main genotypes that infect humans

with different epidemiological characteristics. Genotypes 1

and2,describedonlyinhumans,istheetiologicagentoflarge

fecal-oral transmissionepidemics, typicalofoverpopulated

regions andwith poorsanitaryconditions. Genotype1has

https://doi.org/10.1016/j.bjid.2018.02.001

(2)

beendescribedinAsiaandAfrica,andgenotype2inMexico

andAfrica.Genotype3,describedinEurope,theAmericasand

Japan,andgenotype4,describedinAsia,infectalargevariety

ofmammalsandoccurintheformofsporadiccases.Ingestion

ofundercookedporkmeatisimplicatedinthetransmission

ofsuchzoonosis.HEVcanalsobetransmittedalternativelyby

parenteralroute,suchastransfusionofbloodproducts.2

BrazilexhibitsanintermediateHEVseroprevalence,with

ratesof2–9.8%reportedinblooddonors,5,6_12%_in_intravenous

drugusers,7_and_15%_in_renal_transplant_patients.8_Previous

study ofHEVseroprevalence amongpigs inBrazildetected

seropositivityin97.3%oftheanimalsolderthan25weeksof

age.9 _Genotype₃_was_identiﬁed_in_humans_and_pigs_in_this

country.8,10,11

Itisestimatedthat1.6–3.2millionpeoplearechronically

infectedwithhepatitisCvirus (HCV)inBrazil.12,13 _A

previ-ousstudyfoundhighmorbidityand mortalityratesofHEV

infectioninpatientswithunderlyingliverdisease.14

HEVseroprevalencein patientswithchronichepatitisC

virus infection isunknown in Brazil. Theobjective ofthis

studywastoestimatetheprevalenceofanti-HEVantibodies

inchronicHCVinfectedpatientsintheStateofSãoPaulo,as

wellastoevaluateassociatedriskfactors.

Material

and

methods

Studydesign

Across-sectionalstudyofseroprevalencewasconductedat

thehepatitisoutpatientservicesofUniversidadeFederalde

SãoPaulo,HospitaldeTransplanteEuryclidesJesusZerbini,

andInstitutodeVacinac¸ãoeInfectologiadePiracicaba,inthe

StateofSãoPaulo,Brazil.

BetweenOctober2015andDecember2016patientsofboth

sexes,olderthan18 years,and withchronicHCVinfection

conﬁrmedbyRT-PCRRNAwereincluded.

Samplesizecalculation

AssuminganexpectedprevalenceofHEVinfectionof8%,a

conﬁdencelevelof99%,andatotalwidthofconﬁdence

inter-val(CI)aroundtheexpectedproportionof6%,atotalof576

HCVinfectedpatientswouldbenecessary.

Anti-HEVantibodiesdetection

Thepresenceofanti-HEVIgGwasassessedbyenzyme-linked

immunosorbent assay (WANTAI HEV-IgG ELISA), strictly

accordingtothemanufacturer’srecommendations.Patients

positiveforHEVIgGwerefurthertestedforthepresenceof

HEVIgMantibodiesusingaspeciﬁckitfromthesame

manu-facturer.

Datacollection

ThefollowingvariablespossiblyassociatedwithHEV

infec-tion were assessed: sex, age, socioeconomic class, HIV

co-infection,previoushistoryofcontactwithpigs(haveever

livedinaplacewherepigswereraised),consumptionofpork

meat(never,seldom,often),previoususeofintravenousdrugs,

priorsurgery,transfusionofbloodproducts,presenceof

tat-toos,andhemodialysis.

Forthecharacterizationofthesocioeconomicclassthe

val-idatedquestionnaireofeconomicclassiﬁcationdevelopedby

theBrazilianAssociationofResearchCompanies(ABEP),

ver-sion201515_was_used.

Statisticalanalysis

Seroprevalence ofIgGantibodies ispresentedwiththe

rel-evant 95% conﬁdence interval, calculated according to the

Wilsonmethod.

Student’st-testwasusedtocomparemeans.Proportions

werecomparedbytheChi-squaretestwithYatescorrection,

orbytheFisher’sexacttest,whenappropriate.

Associationswithsigniﬁcancelevelwithp<0.20in

univari-ateanalysesweresubsequentlyincludedinmultiplelogistic

regressionmodels.Statisticallysigniﬁcantindependent

asso-ciationwasconsideredifp<0.05aftermultivariateanalysis.

AlltheanalysesandchartsweredrawnusingprogramR,

version3.3.2(TheRFoundationforStatisticalComputing).

Ethicalaspects

Thestudyprotocolconformstotheethicalguidelinesofthe

1975DeclarationofHelsinkiasreﬂectedinapriorapproval

bytheInstitutionalEthicsReviewBoardoftheFederal

Univer-sityofSãoPaulo(0350/2015).Awritteninformedconsentwas

obtainedfromeachpatientincluded.

Results

A total of 618 patients with chronic HCV infection was

included. Table 1summarizes the distribution ofthe

vari-ablesevaluated.Fifty-threepercentofthesampleweremales,

mean age of53.8 years,minimum22 and maximum of86

years.Historyofpreviouscontactwithpigswasreportedby

30.4%oftheassessedpatients.Twenty-sixcasesofHIV

infec-tion(4.2%),ﬁvehemophiliacs,threepregnantwomen,andtwo

transplantedpatientswereincluded.

Atotalof63caseswereanti-HEVIgGreactive,

determin-ing seroprevalenceof10.2%(95%CI8.0–12.8%).Threecases

(0.5%)hadanindeterminateresultandtheinformationwas

consideredasnegativeforhypothesistesting.Anti-HEVIgM

wasscreenedinallreactiveandindeterminatecasesforIgG

andallofthemturnedoutnon-reactive.

Table 2 summarizes the results of univariate analyses

for the dependent variable anti-HEV IgG and the assessed

variables. Onlythe variables ageand history ofhome

con-tact with pigs were signiﬁcantly associated with anti-HEV

IgG. Hemophiliaexhibitedalevel ofborderlinesigniﬁcance

(Fisher’sexacttestp=0.09).

Fig.1illustratestheconditionaldensitycurveofanti-HEV

IgGassociatedwithage,showingaseeminglylinearincrease

from30to70years,withprevalencerangingfromzeroupto

20%betweenthesetwopoints.Themeanageofpatientswith

positiveandnegativeanti-HEVIgGwas57.98and53.36years

(3)

Table1–DemographicandclinicalcharacteristicsoftheHCVinfectedpatients. Variable Distributionn(%) *Median[p25–p75] N(%) Sex 618(100) Male 326(52.8) Female 292(47.2) Age *54[46–62] 618(100) 22–29years 7(1.1) 30–39years 70(11.3) 40–49years 130(21.1) 50–59years 199(32.3) 60–69years 160(25.9) 70–86years 51(8.3) Socioeconomicclass 449(72.7) A 22(4.9) B 155(34.5) C 249(55.5) D–E 23(5.1) Parenteralexposure 485(78.5) Intravenousdrug 94(19.4) Tattoo 109(22.5)

Transfusionofbloodproducts 188(38.9)

Previoussurgery 352(72.6)

Hemodialysis 10(2.1)

Porkmeatconsumption 475(76.9)

Never 53(11.2)

Rarely 207(43.6)

Often 215(45.3)

Contactwithpigs 144(30.4) 474(76.7)

HAVIgG+ 352(91.7) 385(62.3)

ToxoplasmosisIgG+ 95(70.4) 135(21.8)

HIVinfection 26(4.2) 618(100)

respectively.Seroprevalenceabove60years(15.2%)was sig-niﬁcantlyhigherthaninthoseagedlessthan60yearsofage (8.4%)[OR:1.96;p=0.014].Theprevalenceofreactiveanti-HEV IgGincaseswho hadcontactwithpigswas14.58%and in thosewithnocontacthistoryitwas7.58%(OR:2.08;p=0.028). Themeanageofthecaseswithahistoryofcontactwithpigs (56.5years)wassigniﬁcantlyhigherthanthemeanageof indi-vidualswithout ahistory ofcontactwithpigs(52.81years) [Student’st-testwithp<0.001].

Variables withalevel ofsignificance lower than 0.20 in theunivariateanalysiswereincludedinthemultiplelogistic regressionmodel. Hemophiliawasexcluded from the mul-tivariate analysis due to presence in only five cases and, consequently,alargeconfidenceinterval.Agewasincluded inthemodelasadichotomousvariable,withcutoffpointat 60yearsasindicatedbyROCcurve analysis.Table3shows

theﬁnalmultivariatemodelwithadjustedORswith95%CI

andtheirrelevantlevelsofsigniﬁcance.Ageandprevious

con-tactwithpigswereremainedindependentlyassociatedwith

reactiveanti-HEVIgG.

Discussion

Thepresentstudydemonstrated10.2%(95%CI8.0–12.8%)HEV

seroprevalenceinpatientswithchronicHCVinfection.Older

ageandpriorcontactwithpigswereindependentlyassociated

withreactiveanti-HEVIgG.

ThisstudyistheﬁrstevaluationofHEVseroprevalencein

patientswithchronicHCVinfectioninBrazil.Itisimportant

tonotethatthesamplestudiedisnotnecessarily

representa-tiveoftheHCV-infectedpopulationinthecountry.Onlythree

centersintheStateofSãoPauloparticipatedinthestudyand

inclusionstartedinOctober2015,aperiodconcomitantwith

thereleaseofnewdrugsforthetreatmentofHCVinBrazil.

Thus,theparticipatingcentersbegantoexperiencea

predom-inantﬂowofpatientswithindicationofHCVtreatment,the

majoritywithadvancedliverﬁbrosis (equivalenttoMetavir

scoreF3orF4).Inthesampleassessed,45%ofthepatients

hadlivercirrhosis.

In618HEVsserologiesonlythreecaseswithundetermined

resultsweredetected,showingahighdiscriminatorypower

of the test used. No reactive IgM case was detected thus

indicatingnorecentHEVinfection.TheHEVseroprevalence

(10.2%)inthe samplestudiedwassimilar tothatobserved

ina recentstudy in500blood donorsfrom thecity ofSão

Paulo(9.8%)usingthesameserologickit.5_High_{seroprevalence}

wasalsodescribedincertainspeciﬁcgroups,suchas

profes-sionalsexworkers(17.7%,N=214),16_intravenous_drug_users

(11.8%,N=102),7_cases_of_acute_hepatitis_(17.7%,_N₌_79),16_renal

transplant(15.0%,N=192),8_and_HIV_infected_patients_(10.7%,

(4)

Table2–HEVseroprevalenceaccordingtotheindependentvariablesassessedamongHCVinfectedpatients. Variable IgG+ n(%) OR 95%CIOR p-Value Sex Male 37(11.4) 1.30 [0.77–2.21] 0.39 Female 26(9.0) Age(years) ≥60 30(14.4) 1.88 [1.11–3.19] 0.002 <60 33(8.1) Socialclass NS NS 0.57 A 2(9.1) B 12(7.7) C 22(8.9) D-E 2(8.7) Exposuretopigs 0.04 Contact 20(14.0) 1.99 [1.06–3.72] Nocontact 25(7.6)

Porkmeatconsumption NS NS 0.88

Never 5(9.4) Rarely 22(10.2) Often 28(8.8) Intravenousdrug 0.34 Yes 12(12.9) 1.50 [0.74–3.02] No 35(9.0) Tattoo 0.13 Yes 6(5.6) 0.48 [0.19–1.16] No 41(11.0)

Transfusionofbloodproducts 1.00

Yes 18(9.6) 0.99 [0.52–1.79] No 29(9.9) Previoussurgery 0.84 Yes 33(9.4) 0.88 [0.46–1.71] No 14(10.5) Hemodialysis 1.00 Yes 1(10.0) 1.02 [0.13–8.28] No 46(9.8) HIV 1.00 Yes 3(11.5) 1.15 [0.33–3.94] No 60(10.2) Hemophilia Yes 2(40.0) 5.99 [0.98–36.5] 0.09 No 61(10.0)

Variableswithsignificantassociation:ageandcontactwithpigs.Hemophiliaexhibitedlevelofborderlinesignificance.NS,notsignificant.

Temporal, geographical and methodological differences limit comparison between the different surveys. Previous studieshavedemonstratedsigniﬁcantdifferencein sensitiv-itybetweenthedifferentserologicalkits.18–21_Therefore,_it_is

possiblethatpreviousstudieshaveunderestimatedHEV

sero-prevalenceinthiscountry.Itisimportanttoemphasizethat,

todate,noserologicaltestforHEVhasbeenreleasedbythe

BrazilianFoodandDrugAdministration(ANVISA)forusein

Brazil.Therefore,sincethereisnotestavailableforroutine

use, theinfection isnotdiagnosedinour country.Existing

information was produced using imported kits authorized

onlyforuseinclinicalresearch,andpointedatmore

com-monHEV infection thanclinically suspected.To date,only

anecdotalcasesofactiveHEVinfectionhavebeenreported

inBrazil,allofthemcausedbygenotype3.8,10,11_Taking_into

accountthecurrentstudyandotherspreviouslyconductedin

thecountry,itmaybereckonedthatHEVisthesecondmost

commonhepatotropicvirusinBrazil,comingonlyafter

hep-atitisAvirus(HAV).Morecomprehensiveandrepresentative

seroprevalencestudiesarerequiredtoconﬁrmthis

hypothe-sis.

Few studies havedirectly evaluatedHEV seroprevalence

in patients with HCV. A case-control study conducted in

Italy, publishedin1994,comparedsamplestakenfrom 100

anti-HCV reactive individuals with 100 non-reactive

anti-HCV. Anti-HEV was reactive in 27% and 2%, respectively

(p=0.00012).22_However,_the_age_difference_between_the_groups

(5)

30

Age conditional density on HEV

40 50 Age HEV IgG+ IgG − 60 70 0.0 0.4 0.8 80

Fig.1–HEVseroprevalenceassociatedwithage.Lightgray=non-reactiveIgG;darkgray=reactiveIgG;axisy=proportionof cases.

Table3–Multiplelogisticregressionﬁnalmodelforthe dependentvariableanti-HEVIgG.

Variable Coefﬁcienta _OR _IC_95%_OR _p-Value

Age≥60a 0.71 2.04 1.09–3.79 0.0237

Contactwithpigs 0.68 1.99 1.06–3.71 0.0298

a _Coefﬁcient_value_in_logit_function._Excluding₁₄₄_cases_of

incom-pletedata.

alsosigniﬁcant(medianage60versus42years).Patientsolder

than 50 yearspresentedanti-HEV seroprevalence of24.5%,

comparedto2.2%ofcasesunder50years ofage (OR:14.3;

p=0.0002).Themethodologyusedinthisstudy,wherecases

andcontrolswerenotmatchedforage,castsdoubtaboutthe

validityofthehigherprevalenceofanti-HEVinpatientswith

reactiveanti-HCV,withagebeinganimportantconfounding

factor.

AstudyconductedinTurkeyevaluatedtheprevalenceof

anti-HEVinpatientswithchronichepatitisBandCand

con-trols.Thecontrolgroup,withoutHBVorHCVinfectionand

withoutknownliverdisease,presentedreactiveanti-HEVIgG

in15.7%ofthecases(28/178).TheHBVgroupdidnotpresenta

statisticallysigniﬁcantdifference,whereastheHCVgroup

pre-sentedarateof54%(94/174).SuchahighprevalenceofHEV

foundamongHCVreactiveindividualssuggestsacommon,

probablyparenteral,routeoftransmission.23

Our study shows higher seroprevalence in hemoplilic

patients (40% versus 10%; p=0.09). Because of the limited

number of cases with this condition and the consequent

large95%CI,hemophiliawasnotincludedinthemultivariate

model.Takingintoaccountthatthisisagroupofpatientswith

ahistoryofmultipletransfusionsofbloodcomponents,

char-acterizingahighriskofacquisitionofblood-borneinfections,

it canbehypothesized thatthe infection ofthesepatients

occurredbybloodtransfusion.Similarly,studiesconductedin

Japan24_and_in_Tunisia25_identiﬁed_{signiﬁcantly}_higher

sero-prevalenceofHEVinhemophiliacpatients(7.5%and16.3%)

whencomparedtoblooddonors(4.5%and3.7%),respectively.

Arecentstudyevaluated500bloodbanksamplesfromSão

Paulo. NoHEV RNA was detectedin any sample,but only

samplespositivefor anti-HEVIgG were submittedforRNA

detection.5_On_the_other_hand,_a_study_carried_out_in_England

with225,000blood banksamplesdetected79casesofHEV

genotype3viremia(1:2848).Follow-upof43patientswhohad

received viremicblood identiﬁed18 casesofHEVinfection

(infectionrate=42%).26_The_high_{seroprevalence}_(15%)_of_HEV

foundinrenaltransplantpatientsinSãoPaulocorroborates

thehypothesisofaparenteraltransmissionroute.8

Rivero-Juarez et al. when assessing 894 HIV-infected

patients, 461 HIV/HCV co-infected patientsand 399

mono-infectedHIVpatients,foundnodifferenceinHEV

seropreva-lence between HCV infected (9.7%) and HCV non-infected

(10%)patients.Thisstudydidnotevaluateriskvariablesof

parenteral exposure.27 _It_is_possible _that_both _reactive _and

non-reactiveHCVpatientshadasimilarparenteralrisk,since

inmorethan60% ofthe patientsenrolled,allofthemHIV

infected,therewasnoriskofsexualexposure.Therefore, it

canbehypothesizedthatsimilarparenteralexposurebetween

reactiveandnon-reactiveHCVpatientswouldmakeit

impos-sibletodetectthedifferenceinHEVseroprevalencebetween

thetwogroups.

Passos-Castilho et al. found an HEV seroprevalence of

9.8% (95% CI 7.5–12.7%) using the Wantai kit in 500blood

donors from thecity ofSão Paulo. Theprevalenceof

anti-HEV IgG was signiﬁcantly increased in older age groups.5

Similarly,ourstudyhasdemonstratedasigniﬁcantand

inde-pendent association between HEV seroprevalence and age

increase.Actually,noreactiveanti-HEVIgGcasewasdetected

inpatientsunder30yearsofage.However,areactiveresult

wasobservedin14.2%ofpatientsover60yearsofage.The

differencemaybeexplainedasfollows:antibodiesoftheIgG

class persistforanundetermined time,making

seropreva-lence a cumulative phenomenon as the population grows

older;someotherunassessedriskfactormaybeassociated

withbothageandHEVinfection;andthepossibilitythatin

thepast,theincidenceofHEVinfectionwashigher.Studies

carriedoutindevelopedcountries,withapredominanceof

genotype 3HEV,havealsodescribedaprogressiveincrease

inseroprevalence withaging, particularlyin theage group

above50years.2_Thus,_it_is_possible_that_most_of_the_patients

includedinthepresentstudy,withapresumedHCVinfection

formorethantwodecades,becameinfectedbyHEVatalater

time.

Animportantassociationofcontactwithpigsrearing

his-torywasalsodetected(adjustedOR=1.99;95%CI1.06–3.70).

(6)

bygenotype3HEVwithhighgeneticsimilaritywiththevirus

detectedinhumans.28,29_Previous_study_of_HEV_{seroprevalence}

amongpigsinBrazildetectedseropositivityin63.6%ofthe

ani-malstested(N=357).Inaddition,97.3%ofthepigsolderthan

25weeksofagewereHEVIgGreactive,demonstratingearly

infectionandthepossibilityofactiveinfectionduringtheage

priortoslaughterforconsumption.9_HEV_RNA_was_recovered

from118liversamples(1.7%) andbile(0.84%)fromhealthy

pigsofapigfarminthestateofParaná.30_Another_study

car-riedoutinthestateofRiodeJaneirodetectedHEVRNAin11

outof115(9.6%)swinebilesamplesandinthreeoutofsix

(50%)samplesobtainedinthedrainagepipesoftheevaluated

breedingsites.31_In_both_studies,_genotype₃_HEV_was_detected

asbeingtheunderlyingetiologicagent.

Althoughthisstudydetectedastrongassociationof

reac-tiveHEVIgGwithahistoryofcontactwithpigs,it didnot

detectanyassociation withporkmeatconsumption. Afew

reasonsmayexplainthislackofcorrelation:insufﬁcientpower

todetect the difference, given thelow proportion ofcases

whoreportednotconsumingpork;possibilityofan

alterna-tiverouteofHEVinfection,suchasinter-humanfecal-oral,

directcontactwiththeanimalsorwiththeirwaste;andthe

semi-quantitativecharacteristics ofthefood questionnaire,

withinaccuratequestionsandanswerswhichmayhavenot

reﬂectedtheactualrespondents’consumption.Previous

stud-ies identiﬁed HEV genotype 3 inuntreated sewage,32 _river

waters,33_and _bivalve_molluscs,34_{corroborating} _the

hypoth-esisthatHEVinfectioncanbetransmittedbycontaminated

waterandthattheconsumptionofinfectedmeatperhapsis

notthemainformoftransmissionofthiszoonosis.

Previousstudies have shown ahigh riskof severe HAV

andHBVsuperinfectioninpatientswithchronicHCVorliver

disease.35,36 _Thus, _the _indication _of _immunization _against

HAVand HBVinsuchsituations isa consensus,whenthe

serologicalproﬁleissusceptible.37_Similarly,_high_lethality_of

HEVgenotype1infectedpatientswasdemonstratedin

cir-rhoticpatients (70%at 12months).14 _However, _the _routine

serologicalinvestigationofHEVisnotrecommendedbythe

current management guidelines forchronic HCV infection.

Theseroprevalence ofHEV foundin this and other recent

studiesconductedinBrazilsupporttheinvestigationofthe

serologicalproﬁleofHEVinpatientswithchronichepatitisC

andprophylacticmeasuresagainstHEVinfection.

Thegeneralizabilityoftheresultsofthisstudyislimited

asithasincludedonlythreecentersintheStateofSãoPaulo.

Inaddition,themajorityofstudysubjectshadadvancedliver

ﬁbrosis(equivalenttoMetavirscoreF3orF4)andtherefore

atendencytobeolder.Therefore,anoverrepresentation of

older patientsmight haveoverestimated the prevalenceof

HEVinfectionamongHCVinfectedpatients.

Authors’

contribution

Guilherme Bricks:Main Author; study concept and design;

acquisitionofdata;analysisandinterpretationofdata;

draft-ingofthemanuscript;statisticalanalysis;fundraising.Jorge

FigueiredoSenise:Acquisitionofdata;critical reviewofthe

manuscriptforintellectualcontent.HenriquePottJunior,

Giu-lianoGrandi,DimasCarnaúbaJuniorandHamiltonAntonio

BonilhadeMoraes:DataAcquisition.AmandaPassariniand

DéboraBelliniCaldeira:Laboratorysupport.CelsoFrancisco

HernandesGranato:Laboratorysupport,criticalreviewofthe

manuscript forintellectual content. AdautoCastelo Junior:

Studysupervision,concept anddesign;acquisition ofdata;

analysisandinterpretationofdata;statisticalanalysis;fund

raising;criticalreviewofthemanuscriptforintellectual

con-tent.

Conﬂicts

of

interest

Theauthorsdeclarenoconﬂictsofinterest.

Acknowledgement

This study was supportedby the National Council for the

ImprovementofHigherEducation(CAPES).

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