Microbialpathogensandstrategiesforcombatingthem:science,technologyandeducation(A.

MndezVilas,Ed.)
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Modulationofmucosalantiviralimmuneresponseby
immunobioticlacticacidbacteriaPartII:the
respiratorymucosa
1,2,4,*

1,3

1,2

1,2

J.Villena
,M.G.VizosoPinto ,V.Rodriguez ,H.Kitazawa ,S.Salva and
1,2,5,**
S.Alvarez
1

ImmunobioticResearchGroup2ReferenceCentreforLactobacilli(CERELA
CONICET).Tucuman,Argentina.3INSIBIOCONICET,BiomedicalDepartment,Faculty
ofMedicine,NationalUniversityofTucumn,Argentina.4FoodandFeedImmunology
Group,GraduateSchoolofAgriculturalScience,TohokuUniversity.Sendai,Japan.
5
InstituteofAppliedBiochemistry,FacultyofBiochemistryandPharmacy,National
UniversityofTucumn.Tucuman,
Argentina.*Correspondingauthor.MailingaddressforJulioVillena:Laboratoryof
Immunobiotechnology,ReferenceCentrefor
Lactobacilli(CERELACONICET).Tucuman,Argentina.Phone:+543814310465.
Fax:+543814005600.Email:
jcvillena@cerela.org.ar
**Correspondingauthor.MailingaddressforSusanaAlvarez:Laboratoryof
Immunobiotechnology,ReferenceCentreforLactobacilli(CERELACONICET).
Tucuman,Argentina.Phone:+543814310465.Fax:+543814005600.Email:
salvarez@cerela.org.ar
Virusesarethemostimportantcauseofseveremucosalinfectionsworldwideespecially
inhighriskpopulationssuchasininfants,youngchildren,elderlyand
immunocompromisedhosts.Lacticacidbacteria(LAB)aretechnologicallyand
commerciallyimportantandhavevariousbeneficialeffectsonhumanhealth.Several
studieshavedemonstratedthatcertainLABstrainscanexerttheirbeneficialeffectonthe
hostthroughtheirimmunomudulatoryactivity.Thesestrains,termedimmunobiotics,
havebeenusedforthedevelopmentofprobioticfoodswiththeabilitytostimulate
mucosalantiviralimmunity.Inthisreviewweexaminethecurrentscientificliterature
concerningtheadvancesinourunderstandingofhowprobioticmicroorganismsareable
tomodulaterespiratoryviralimmunityandaffecttheoutcomeofviraldiseases.
Moreover,thisreviewexplorestherecentadvancesofourlaboratoriesregardingthe

cellularandmolecularinteractionsbetweenimmunobioticsandhostscellsandhowthis
interactionmodulatetheresistanceagainstrespiratoryviralinfections.Researchfromthe
lastdecadedemonstratesthatimmunobioticLABrepresentapromisingresourceforthe
developmentofpreventionstrategiesagainstviralinfectionsthatcouldbeeffectivetools
formedicalapplication.
Keywordslacticacidbacteria,immunobiotics,respiratoryviralinfection1.

Modulationofrespiratoryantiviralimmuneresponseby
immunobioticLAB
RespiratorySyncytialVirus(RSV)isamajorrespiratorypathogenofinfantsand
childrenandanemergingpathogenoftheelderly.Otherimportantviralpathogen
isInfluenzaVirusthatisthemostcommoncauseofhumanrespiratoryinfection,
andisamongthemostsignificantbecauseitcauseshighmorbidityandmortality.
Severallinesofevidenceshowedthatmucosaladministrationofimmunobioticsis
abletoincreaseresistanceagainstrespiratoryviralinfections[1].
Thefirststudydemonstratingthebeneficialeffectofimmunobioticsoninfluenza
infectionwasperformedwithBifidobacteriumbreveYIT4064.Thisstrainisable
toinducetheproductionoflargequantitiesofIgAinamurinePeyersPatches
(PPs)cellculturemethod[2].Moreover,invitrostudiesdemonstratedthatthe
YIT4064strainenhancestheproductionofantiinfluenzavirus,antirotavirus,and
antipoliovirusantibodiesbyPPscellsinresponsetothechallengeswiththe
respectiveviruses[3].ItwasshownthattheoraladministrationofB.breve
YIT4064protectedmiceagainstinfluenzaviruschallenge[4].Theauthors
demonstratedthatoraladministrationofB.breveYIT4064significantlydecreased
theaccumulatedsymptomrateofinfluenzavirusinfectionandimprovedthe
survivalrate.Theseprotectiveeffectswererelatedtoaugmentedlevelsofanti
influenzavirusIgGinserum[4].Therefore,Yasuiet.aldemonstratedthatB.
breveYIT4064protectagainstrespiratoryviralinfectionthroughanenhancement
ofhumoralimmuneresponse.Recently,itwasshownthatoraladministrationof
heatkilledlactobacilliareabletoimproveIgAandIgGantiinfluenzaantibodies
intheairwaymucosaandlung,andtoaugmentprotectionagainstinfluenzavirus
infectioninmice[5,6].Then,immunobioticsarecapabletomodulatethe
productionofsystemicandmucosalantibodiesagainstrespiratoryviruses(Table
1).
Horietal.[7]studiedtheeffectofthenasaladministrationofanonviableL.casei
Shirotaonrespiratoryimmunityandobservedthatnasaltreatmentofadult
BALB/cmicewiththisstrainstimulatedcellularimmunityintherespiratorytract
andsignificantlyincreasedtheresistanceofmicetoinfluenzavirusinfection.The

authorsinvestigatedtheproductionofvariouscytokinesbymediastinallymphoid
nodescellsinmicereceivingL.caseiShirotaintranasally.Itwasfoundthatthe
ShirotastrainstronglyinducedproductionofIL12inthesecells,whichisan
importantcytokineforcytotoxicTcellsandNKcellsstimulationand
enhancementofTh1cytokinesproductionandTh1cellsproliferation.Moreover,
bothIFNandTNFlevelswereimprovedinmediastinallymphoidnodescell
culturesfrommiceadministeredL.caseiShirotaintranasally,afterinfluenzavirus
challenge[7].Thiswasthefirststudydemonstratingthat
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nasallyadministeredLABwereabletoactivatecellularimmunityinthe
respiratoryimmunesystemandprotectedagainstarespiratoryviralinfection.
Later,thegroupinvestigatedwhetheroraladministrationofL.caseiShirota
activatesthesystemicandrespiratoryimmunesystemsandwhetheritameliorates
influenzavirusinfectionintherespiratorytractofaged[8]andinfantmice[9].
TheseworksshowedthatorallyadministeredL.caseiShirotaaugmentedNKcell
activityinsplenocytesandlungsandenhancedIFNandTNFproductionof
nasallymphocytesofagedandinfantBALB/cmice.Thesestudiesalsofoundthat
viraltitersinL.caseiShirotagroupsweresignificantlylowerthanthatinthe
controlgroups[8,9].TheauthorspostulatedthatL.caseiShirotawastakenupby
McellsinthePPsandstimulatedTh1cellsandNKcells,whichmigratedtothe
mesentericlymphoidnodesandthen,viathethoracicductandbloodstream
migratetothespleen,lungs,andnasalassociatelymphoidtissue.
OtherstudieshavealsoemphasizedtheimportanceofIFNproductionandNK
cellsactivationfortheprotectiveeffectofimmunobioticsagainstinfluenza
infection(Table1)[1012].Recently,Kawaseetal.[13]showedthatoral
administrationofheatkilledL.gasseriTMC0356resultedinsignificantlyhigher
expressionofpulmonaryIFNinmicewhencomparedtocontrols.Moreover,
authorsevaluatedtheeffectoforallyadministeredlyophilizedL.rhamnosusGG
andL.gasseriTMC0356intheoutcomeofinfluenzavirusinfectioninmiceand
foundsignificantdifferencesinpulmonaryvirustitresbetweencontroland
lactobacillitreatedmice.Inaddition,histologicaltissueexaminationshowed
bronchialmucosaepitheliumhypertrophyandinfiltrationofleukocytesinthe
bronchialsubmucosaandpulmonarycoreofinfectedcontrolmicewhilesimilar
pathologicalchangeswerenotobservedinlactobacillitreatedmice[13].Takeda

etal.[14]showedthattheoraladministrationofL.plantarum06CC2isableto
increaseIFNexpressioninPPsandlungs.ImprovedrespiratoryIFNinduced
bythe06CC2strainwasassociatedwithaugmentationofNKcellactivityand
correlatedwiththealleviationofinfluenzainfectioninmice.Moreover,authors
demonstratedthattheimmunobiotictreatmentdifferentiallymodulatedthe
productionofcytokinesduringinfluenzainfection.ThelevelsofIFN,IL12,
andIFNininfectedmiceadministeredthe06CC2strainweresignificantly
higherthanthoseinthecontrolswhilethelevelofTNFwassignificantlylower
thanthatinthecontrolmice[14].
Maedaetal.[15]reportedthatoraladministrationofheatkilledLactobacillus
plantarumL137enhancesprotectionagainstinfluenzavirusinfectionby
stimulationoftypeIinterferonproduction,demonstratingthatorallyadministered
immunobioticsareabletostimulateinnateantiviralresponsesintherespiratory
tract.Thestudyshowedthatallthemiceinfectedwithamouseadaptedvirulent
strainofinfluenzavirusH1N1diedwithin14daysafterintranasalchallenge.
However,themeansurvivaltimewassignificantlyprolongedinL.plantarumL
137treatedmice[15].IFNwashardlydetectedintheserumofcontrolmiceon
day1,2,3or6afterinfection,whilesignificantlevelsofIFNweredetectedin
theserumofL137treatedmiceattheearlystagedaysafterchallenge.Thisstudy
demonstratedthattheincreasedproductionofIFNinducedbytheimmunobiotic
straincontributestotheincreasedresistanceagainstinfluenzavirusinfection;
however,detailedstudiestoinvestigatetheimmunemechanismsinvolvedinthis
effectwerenotperformed.
Morerecently,Leeetal.,[16]investigatedwhetherthesublingualrouteisuseful
forthedeliveryofprobioticsagainstinfluenzavirusinfection.Theauthors
demonstratedthatsublingualadministrationofL.rhamnosusenhancedprotection
againstinfluenzavirusinfectionbyenhancingofmucosalsecretoryIgA
productionandTcellandNKcellactivity.Moreover,IL12levelsinthelungsof
L.rhamnosustreatedmiceincreasedsignificantlywhencomparedtocontrols
whileIL6andTNFlevelsdecreasedsignificantly.ConsideringthatIL6and
TNFlevelspositivelycorrelatewithlunginflammationandvascular
dysfunction,theseresultssuggestthatthedecreasedlevelsofproinflammatory
citokinesmightalsocontributetotheprotectionagainstinfluenzavirusinfection
[16].
Collectively,thesestudiesindicatethat:a)orallyadministeredimmunobioticscan
increaserespiratoryimmunityagainstinfluenzavirus,b)theeffectismediatedby
theimprovementofrespiratorytractandsystemiclevelsofIFNandNKcells
+
activity,c)theactivationofaTh1responsewithIFN Tcellsmobilizationfrom

guttorespiratorytractwouldbethemechanisminvolved,d)nasalprimingwith
immunobioticLABisalsoaninterestingalternativetoimproverespiratory
antiviraldefencesand,e)immunobioticLABhavethepotentialtobesuccessfully
usedasadjuvantsforantiviralvaccines.
Thestudiesdescribedaboveshowedthatmucosaladministrationof
immunobioticshadthepotentialtoimprovetheoutcomeofinfluenzavirus
infection.However,thecapacityofimmunobioticstoimproveprotectionagainst
pneumovirusesofthefamilyParamyxoviridaesuchasRSVwasnotinvestigated
indetail.Inanefforttoevaluatethecapacityoflactobacillitoreducethe
pathogenesisofseverepneumovirusinfectioninvivo,Gabryszewskietal.,[17]
developedamodelpneumoniavirusofmice(PVM)infection.Theauthors
demonstratedthatlactobacilli,whentargetedtotherespiratoryepithelium,are
highlyeffectiveatsuppressingPVMinducedinflammationandprotectingagainst
lethaldisease.Wildtypemiceprimedviaintranasalinoculationwithliveorheat
inactivatedLactobacillusplantarumorLactobacillusreuteriwerecompletely
protectedagainstlethalinfectionwithPVM.Primingwithlivelactobacilliresulted
indiminishedgranulocyterecruitment,diminishedexpressionofmultiplepro
inflammatorycytokines(CXCL10,CXCL1,CCL2,andTNF),andreducedvirus
recovery.Moreover,Lactobacillusprimingalsoresultedinprolongedsurvivaland
/
protectionagainstthelethalsequelaeofPVMinfectioninMyD88 mice,
suggestingthattheprotectivemechanismsmaybeTLRindependent(Table1).
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Ourlaboratoryperformedarandomizedcontrolledtrialinordertoevaluatethe
effectofaprobioticyogurtcontainingtheimmunobioticstrainL.rhamnosus
CRL1505onbothgutandnongutrelatedillnessesamongchildren[18].We
demonstratedthatadministrationofL.rhamnosusCRL1505improvedmucosal
immunityandreducedtheincidenceandseverityofintestinalandrespiratory
infectioninchildren.Ourrandomizedclinicalstudydemonstratedasignificant
reductioninoccurrenceofinfectiouseventsassociatedwithconsumptionofL.
rhamnosusCRL1505[18].Wealsoevaluatedthepresenceorabsenceoffever
duringinfectiouseventsaswellastheneedofantibiotictreatmentinchildrenwho
hadinfections,asindicatorsofseverity.Therewasasignificantdecreaseinthe
presenceoffeverinchildrenwhoconsumedprobioticyogurtaswellasaslight
decreaseintheneedforantibiotictreatment,indicatinglessseriousinfectionsin

relationtotheplacebogroup[16].Althoughwedidnotevaluatetheetiologyof
respiratoryinfectionsintheclinicalstudy,previousevaluationshaveshownthat
viralpathogens,suchasRSV,humanmetapneumovirus,influenzaAvirus,
parainfluenzaviruses,andrhinovirusesareconsideredthemajorvirusesthatcan
causerespiratorytractdiseasesinchildren[19].Therefore,thefindingsofour
studysuggestthatadministrationofL.rhamnosusCRL1505mayprovideoneof
thepotentialinterventionstoreducetheburdenofcommonchildhoodmorbidities,
especiallythoseassociatedtoviralinfections[18].
Later,weevaluatedtheeffectoftheoraladministrationoftwoLactobacillus
strains,L.rhamnosusCRL1505andL.rhamnosusCRL1506,onmucosalantiviral
immunity.Invivoexperimentsdemonstratedthattheadministrationoflactobacilli
strainssignificantlyaugmentedtheexpressionofIFNinPPscomparedwiththe
control[20,21].Moreover,wefoundthatL.rhamnosusCRL1505wasmore
efficientthanCRL1506strainforincreasingthelevelsofIFN,IL10andIL6in
theintestine.Onthecontrary,L.rhamnosusCRL1506showedahighercapacityto
improvelevelsofIFN,IFNandTNFinthegutwhencomparedwith
CRL1505.WhenweevaluatedthelevelsofserumcytokineswefoundthatL.
rhamnosusCRL1506wasmoreefficientthanL.rhamnosusCRL1505toincrease
IFN,IFNandTNF,whileserumIFN,IL10andIL6levelsweremore
efficientlyimprovedbyL.rhamnosusCRL1505.Thesechangesintheprofileof
serumcytokinesweresimilartothosefoundintheintestinalfluid,indicatingthat
levelsofserumcytokinesareareflectionofintestinalchanges[20].Onthe
contrary,theanalysisofrespiratorycytokinesshowedthatonlyL.rhamnosus
CRL1505wasabletoincreasethelevelsofIFN,IL10andIL6[21].While
thesearethesamecytokinesthatwereincreasedbythisstraininserum,wecannot
attributeadirectcorrelationbetweenthetwoincreases,aswedidnotfound
increasedlevelsofIFN,IFNorTNFintherespiratorytractofL.
rhamnosusCRL1506treatedmice.Therefore,andtakingintoaccountthecapacity
+
+
+
ofL.rhamnosusCRL1505ofincreasingthenumberofCD3 CD4 IFN Tcells
inPPsandthestudiesmentionedabove,wehypothesizedthatL.rhamnosus
CRL1505wouldbeabletoinduceamobilizationofthesecellsintotherespiratory
mucosa.Wedemonstratedthatthishypothesiswastruesinceincreasednumbers
+
+
+
ofCD3 CD4 IFN TwerefoundinlungsofL.rhamnosusCRL1505treated
+
+
mice[21].Furthermore,wecanspeculatethatthemobilizationofCD3 CD4 IFN
+
TcellsfromtheintestinetotheairwaysandtheimprovedproductionofIFN
couldbeinvolvedintheprotectiveeffectagainstviralinfectionsinducedbyL.
rhamnosusCRL1505thatwasobservedinclinicalstudies[18].
TomimictheproinflammatoryandphysiopathologicalconsecuencesofRNA
viralinfectionsinthelung,weusedanexperimentalmodeloflunginflammation

basedontheadministrationoftheartificialTLR3/RIGIligandanddsRNAanalog
poly(I:C).Nasaladministrationofpoly(I:C)toBALB/cmiceinducedamarked
impairmentoflungfunctionthatwasaccompaniedbytheproductionofpro
inflammatorymediatorsandinflammatorycellrecruitmentintotheairways[21]in
accordancewithresultspublishedbyStowelletal.[61].Exposuretopoly(I:C)
inducedrespiratoryepithelialcelldeathandimpairedepithelialbarrierfunctionas
demonstratedbytheincreasedlevelslactatedeshidrogenase(LDH)activityand
albuminconcentrationinbronchoalveolarlavages(BAL).Moreover,intranasal
administrationofthreeoncedailydosesofpoly(I:C)resultedinneutrophilsand
mononuclearcellsinfluxintothelung[60].
Invitrostudieshavedemonstratedthatstimulationoflungepithelialcellswith
poly(I:C)elicitedthesecretionofmultiplecytokines,chemokines,theinductionof
transcriptionfactorsandincreasedexpressionofTLRs[22].Inourinvivomodel
increasedlevelsofTNF,IL6,IL8andMCP1wereobservedintherespiratory
tract,thereforealikelysourceofcytokinesfollowingpoly(I:C)administration
maybetheairwayepithelium.Inaddition,theexperimentalmodelusedinthis
workresemblesRSVinfectionsincethisrespiratoryvirusisabletoinducea
profileofproinflammatorycytokinessimilartothatobservedfollowinginvivo
poly(I:C)challengeinmice[22,23].Infact,naturalhumanRSVinfectionin
childrenandexperimentalRSVinoculationinmiceresultinprominentlocal
secretionofproinflammatorycytokines,suchasTNF,IL6,andCXC/CC
chemokines,includingIL8,MIP1,RANTES,andMCP1.Thecoordinated
actionsofseveralofthesecytokinesstronglypromotetherecruitmentand
activationofneutrophilsandmonocytes/macrophages[24],alsoobservedinour
experimentalmodel[21].
Duringacutevirallunginfection,itisimperativethatthehostsinflammatory
responseistightlyregulated,enablingpathogeneliminationbutlimitingthe
detrimentaleffectsofinflammationonthegasexchange.Anappropriatebalance
ofantiinflammatoryandproinflammatorymediatorsisessentialforasafeand
effectiveantiviralimmuneresponse.Thus,anexcessiveTNF/IL8/MCP1
responsecanleadtoincreasedimmunopathology,whileexuberantIL10
productioncanresultindelayedpathogenclearance[25].Inthissense,ithasbeen
shownthatTNFcontributestoclearanceofthevirusduringtheearlystagesof
RSVinfection,whichismostlikelyaresultoftheNKcellresponse.But
continuedproductionofTNFexacerbatesillnessandtissueinjuriesduringthe
latestagesofRSVinfection[26].
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Interestingly,recentstudiesdemonstratearoleforIL10incontrolling
immunopathologyduringrespiratoryviralinfections.Sunetal.[27]showedthat
IL10preventsimmunopathologyandlethaldiseaseduringacuteinfluenzavirus
infection.Ontheotherhand,IL10alsoseemstoplayacrucialroleincontrolling
diseaseseverityinRSVinfection[27,28].ItwasfoundthatIL10deficiency
duringRSVchallengedidnotaffectviralload,butledtomarkedlyincreased
diseaseseveritywithenhancedweightloss,delayedrecoveryandagreaterinflux
ofinflammatorycellsintothelungandairwaysandenhancedreleaseof
inflammatorymediators[29].ThepreventiveadministrationofL.rhamnosus
CRL1505reducedtheproductionofTNF,IL6,IL8andMCP1inthe
respiratorytractafterthechallengewithpoly(I:C).Therefore,thereductionof
theseproinflammatorymediatorscouldexplainatleastpartiallythereducedlung
injuriesintheL.rhamnosusCRL1505treatedgroup[21].Moreover,L.
rhamnosusCRL1505treatmentpriortopoly(I:C)challengeinducedasignificant
increaseinIL10inlungandserum.Consequently,IL10wouldbevaluablefor
attenuatinginflammatorydamageandpathophysiologicalalterationsinlungs
challengedwiththeviralpathogenassociatedmolecularpatternpoly(:IC).
Accordingtotheseresults,L.rhamnosusCRL1505treatmentwouldbeneficially
regulatethebalancebetweenproinflammatorymediatorsandIL10,allowingan
effectiveinflammatoryresponseagainstinfectionandavoidingtissuedamage.
OraltreatmentwithCRL1505strainalsoincreasedlevelsofIFN inBALafter
poly(I:C)challenge[21].ThehigherlevelsofrespiratoryIFNafterpoly(I:C)
challengeinL.rhamnosusCRL1505treatedmicecouldbeexplainedbythe
+
+
+
highernumberofCD3 CD4 IFN Tcellsandbyanimprovedactivationofthese
cellsbylungDCs.WhenweanalyzedlungDCsinL.rhamnosusCRL1505treated
miceafterthenasalchallengewithpoly(I:C)wefoundincreasedlevelsofboth
+
high
CD103 andCD11b DCs.Moreover,bothDCspopulationsshowedhigher
expressionofMHCIIwhencomparedwithcontrols.However,IL12andIFN
+
wereincreasedonlyinCD103 DCs[21].Consistentwithourresults,ithasbeen
+
high
demonstratedthatCD4 CD62L DO11.10Tcells,whichhavebeenprimedwith
+
+
lungCD103 DCsinducedhigherfrequenciesofCD4 TcellsproducingIFN
thanIL4[30].
TheseresultsclearlyindicatedthatL.rhamnosusCRL1505wasapotentinducer
ofantiviralcytokinesandmaybeusefulasaprophylacticagenttocontrol
respiratoryvirusinfectionsasobservedintheclinicalstudy.However,further

studieswereneededinordertoconclusivelydemonstratetheprotectiveeffectof
L.rhamnosusCRL1505.Therefore,wenextaimedtoinvestigatewhetheroral
administrationofL.rhamnosusCRL1505wasabletoimproveresistanceagainst
RSVininfantmiceandevaluatedtheimmunologicalmechanismsinvolvedinthe
probioticeffect.WehaverecentlydemonstratedthatoraladministrationofL.
rhamnosusCRL1505to3weekoldBALB/cmicesignificantlyreducelungviral
loadsandtissueinjuriesafterthechallengewithRSV[31].Moreover,ourstudy
showedthattheprotectiveeffectachievedbytheCRL1505strainwasrelatedto
itscapacitytodifferentiallymodulaterespiratoryantiviralimmuneresponse.
NaturalhumanRSVinfectioninchildrenandexperimentalRSVinoculationin
miceresultinprominentlocalsecretionofproinflammatorycytokines,suchas
TNF,IL6,IL8,MIP1,RANTES,andMCP1[24].TheexcessiveTNF ,IL
8andMCP1responsecanleadtoincreasedimmunopathology.Thesepro
inflammatorycytokinescontributetoclearanceofthevirusduringtheearlystages
ofRSVinfection,howevercontinuedproductionofthesefactorsexacerbate
illnessandtissueinjuriesduringthelatestagesofinfection[26].Asmentioned
before,anadequatebalanceofproinflammatoryandantiinflammatoryfactorsis
essentialforasafeandeffectiveantiviralimmuneresponseenablingvirus
eliminationbutlimitingthedetrimentaleffectsofinflammationonthelungtissue.
WedemonstratedthattheCRL1505strainbeneficiallymodulatethebalance
betweenproandantiinflammatorycytokinesinresponsetoRSVinfection[31].
WeobservedthatCRL1505treatedmicewereabletoearlyincreasethelevelsof
TNFandIL6intherespiratorytractwhencomparedtocontrols.Theearly
increaseofthesecytokinestogetherwiththeimprovedlevelsofIFN should
explainthehighercapacityofCRL1505treatedmicetoreduceviralloads.In
addition,orallyadministeredL.rhamnosusCRL1505significantlyincreasedIL10
levelsthatwouldcontributetoprotectionagainstinflammatorydamage[31].
ImprovedproductionofIL10inCRL1505treatedmicecouldhaveother
beneficialeffectssuchasthemodulationofNKcellsactivity.Interestingly,itwas
demonstratedinIL10knockoutmicethattheabsenceofIL10alteredtheNKcell
responseinthelungafterRSVinfection.AuthorsfoundadecreaseinNKcells
numbersandreducedgranzymeBexpressionbyNKcellsinthelungsandairways
afterRSVinfection[29].Inaddition,itwasreportedthatIL10Rblockadeduring
acutemurinecytomegalovirusinfectionresultedinimpairedNKcell
responsivenessandproposedthatIL10actstopromoteNKcellactivationand
survivalinthelung[32].Then,consideringthereportsaboutthebeneficialeffect
ofprobioticsagainstinfluenzainfectionthroughmodulationofNKcellactivities
[33],itwouldbeimportanttoevaluatetheeffectoforallyadministeredL.
rhamnosusCRL1505onlungNKcellsandtheirrelationshipwithIL10

productionwhicharebothinterestingtopicsforfutureresearch.
DCshaveacentralroleintheshapeofinnateandacquiredimmuneresponses
then,wealsoexaminedtheeffectofimmunobioticsonlungDCsactivation.We
+
observedthattheCRL1505strainimprovedthenumbersofbothCD103 and
high
CD11b DCspopulationsaswellastheexpressionofMHCIIinresponseto
RSVchallenge[31].Thiseffectwouldhaveasignificantimpactintheimmune
responseagainstRSVsinceitwasreportedthatbothDCspopulationsare
+
+
importantinthegenerationofRSVspecificCD4 andCD8 Tcells.Inthisregard,
Lukensetal.[34],usingeffectorsTcellsasareadoutsystemtomeasureantigen
+
displaybyMHCIandMHCIImolecules,foundthatbothmigratingCD103 and
high
+
+
CD11b lungDCspresentedRSVderivedantigenstoCD4 andCD8 Tcells.
Then,
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consideringthatRSVinteractionwithlungDCsresultsinactivationand
maturationeventsthatplayimportantrolesinestablishingvirusspecificimmunity
and,thattheseearlyeventsduringtheinitialimmuneresponsemaydeterminethe
qualityanddurabilityofhostimmunityandinfluencesusceptibilitytoreinfection,
wecanspeculatethatL.rhamnosusCRL1505preventivetreatmentwould
significantlyimproveresistanceagainsttheinfectionbybeneficiallymodulating
DCsactivityforthegenerationofaprotectiveadaptiveimmuneresponse.
ItwasreportedthataTh2immuneresponseisfavoredduringRSVinfection,
especiallyinyoungerhosts.RSVusesmultiplemechanismstoinduceaTh2cell
responseinthehost,includingRSVGproteinmediatedeffects[35],increasing
IL4productionfrombasophils[36]andinductionofalternativelyactivated
macrophages[37].Inaddition,itwassuggestedthatRSVinfectioninducesTh2
likeinflammationinthelung,whichpromotesaTh2likeeffectorphenotypein
Tregcellsandalossofsuppressivefunction.Then,itisconsideredthatstrategies
abletoimproveTh1responsesagainstRSVwouldbeneficiallymodulatethe
outcomeoftheinfectionespeciallyinyoungindividuals.Itwasdemonstratedthat
IFNisabletoupregulatetheexpressionofMHCIIandMCHImoleculesin
APCsandtherebyenhancethecellularimmuneresponsetoviralinfectionand
suppresstheproliferationofTh2typeTcells[38].Consistentwiththisnotion,we
showedthattreatmentofinfantmicewithL.rhamnosusCRL1505significantly

improvedtheproductionofIFNinresponsetoRSVinfectionandincreasedthe
capacityofmicetoclearthevirus.Then,modulationofrespiratoryandsystemic
immunitypotentiatedbytheCRL1505strainmightcontributetoanimprovedTh1
toTh2shiftandtherebyfavorprotectiveimmunityagainstviralinfectionssuchas
RSV.WehavedemonstratedthatorallyadministeredL.rhamnosusCRL1505
+
+
+
induceamobilizationofCD3 CD4 IFN Tcellsfromthegutintotherespiratory
mucosaandimprovelocalproductionofIFN[21].Probably,IFNsecretedin
responsetoL.rhamnosusCRL1505stimulationwouldmodulatethepulmonary
innateimmunemicroenvironmentconductingtotheactivationofDCsandthe
generationofaTh1responsewiththeconsequentattenuationofthestrongand
damagingTh2reactionsassociatedwiththesubsequentintranasalRSVchallenge
[31].
RSVisthechiefcauseofbronchiolitisandviralpneumoniainyoungerchildren.
ThebronchiolitisandpneumoniainducedbyRSVinfectionhavebeenbelievedto
beimmunopathologicalinnature,becausealargenumberofinflammatorycells
areaccumulatedandactivatedinthelungsafterinfection.Thestudiesdescribed
beforeshowedthatimmunobioticadministrationisabletomodulateinflammatory
responsesinducedbypneumovirusesoftheParamyxoviridaefamilyinmice,
demonstratingthatimmunobioticsareaninterestingalternativetoachievean
immunoprotectiveeffectduringparamixovirusinfections.
Table1Effectofimmunobioticsonviralrespiratoryinfections.

Strain
B.breve
YIT4064
L.caseiShirota
L.caseiShirota
L.caseiShirota
L.plantarumL137
Viability
Heatkilled

Heatkilled
Heatkilled
Viable
Heatkilled
Mice
Adult
Adult
Aged
Infant
Adult
Route
Oral
Nasal
Oral
Oral
Oral
Challenge
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
Protectiveeffect
ReductionofaccumulatedsymptomrateImprovementofsurvivalrate

ReductionofvirustiterinnasalwashImprovementofsurvivalrate
Reductionofvirustiterinnasalwash
ReductionofvirustiterinnasalwashReductionofaccumulatedsymptomrate
ReductionofvirustiterinlungImprovementofsurvivalrate
ImmunoregulatoryRef.effect
Improvementofserum[4]IgG
ImprovementofIL12,[7]TNFandIFNinMLN
ImprovementofNKcell[8]activityinspleenandlung
andincreaseofTNFandIFNinnasallymphocytes
ImprovementofNKcell[9]activityinlungandlevels
ofIL12inMLN
Improvementofserum[15]IFN
1818
FORMATEX2013
Microbialpathogensandstrategiesforcombatingthem:science,technologyandeducation(A.
MndezVilas,Ed.)
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______________

L.gasseriLyophilizedTMC0356
L.pentosusSHeatkilledPT84
L.rhamnosusLyophilizedGG
B.longumLyophilizedBB536
L.plantarumHeatkilled06CC2
L.pentosusHeatkilledb240
L.plantarumViableATCCBAA793Heatkilled
L.plantarumViableATCC23272Heatkilled

L.rhamnosusViableCRL1505
LactobacilliFormalinmixturetreated
Adult
Adult
Adult
Adult
Adult
Adult
Adult
MyD88/
Adult
MyD88/
Adult
Adult
Oral
Nasal
Nasal
Oral
Oral
Oral
Nasal
Nasal
Oral
Oral

InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
InfluenzavirusH1N1
Pneumoniavirusofmice
Pneumoniavirusofmice
Poly(I:C)
InfluenzavirusH1N1
ReductionofvirustiterinlungReductionofclinicalscoresReductionoflunginjury
ReductionofvirustiterinBALF
ImprovementofsurvivalrateReductionofaccumulatedsymptomrateReductionoflung
injury
ReductionofsymptomscoreReductionoflunginjuryReductionbodyweighloss
ReductionofvirustiterinlungReductionbodyweighloss
ImprovementofsurvivalrateReductionofvirustiterinlung
ImprovementofsurvivalrateReductionofvirustiterinlung
ImprovementofsurvivalrateReductionofvirustiterinlung
Reductionoflunginjury
ImprovementofsurvivalrateReductionoflunginjury
Notstudied[13]
ImprovementofNKcell[10]activityinlungandlevels
ofIL12andIFNinBALF
ImprovementofNKcell[11]activityandlevelsofIL

1,TNF,MCP1andIFNinlung
ImprovementofIL1,[12]IL6andIFNinlung
ImprovementofNKcell[14]activityinspleenandIFN,IFN,IFN,
TNF,IL12andIL6inBALF
Reductionofinfiltratedneutrophils
ImprovementofBALF[5]IgAandIgG
Suppressionofvirus[17]inducedCXCL10,CCL2,CXCL1,CCL9,TNFandCCL24in
aMyD88TLR
signalingindependentmanner
Suppressionofvirus[17]inducedCXCL10,CCL2,CXCL1,CCL9,TNFandCCL24in
aMyD88TLR
signalingindependentmanner
ImprovementofDCsand[21]CD4+IFN+Tcellsin
lungandlevelsofIFN,IL10andIL6inBALF
ImprovementoflungIgA[6]Improvementoflung
TNFandIL12
FORMATEX2013
1819

Lactobacillimixture
L.rhamnosus
L.rhamnosus
CRL1505
Formalintreated
Lyophilized
Viable

Adult
Infant
Infant
Nasal
Sublingual
Oral
InfluenzavirusH1N1
InfluenzavirusH1N1
Respiratorysyncytialvirus
ImprovementofsurvivalrateReductionoflunginjuryReductionofvirustiterinlung
Reductionoflunginjury
ReductionofvirustiterinlungReductionoflunginjury
ImprovementoflungIgA[6]Improvementoflung
TNFandIL12
ImprovementoflungIgA,[16]IL12andNKcellactivity
andreductionofIL6andTNF
ImprovementofDCsand[1]CD4+IFN+Tcellsin
lungandlevelsofIFN,IL10andIL6inBALF
Microbialpathogensandstrategiesforcombatingthem:science,technologyandeducation(A.
MndezVilas,Ed.)
______________________________________________________________________________
______________

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