Advances in detection methods for Shiga toxin ?producing Escherichia coli (STEC).

PADOLA, NORA LÍA.

Frontiers in cellular and Infection Microbiology

Frontiers Editorial

Lugar: Lausanne; Año: 2014

A commentaryon DetectionofShigatoxin-producing Escherichiacoli in groundbeefusing Genedisc real-timePCRsystem by Fratamico,P.M.,andBagi,L.K.(2012). Front.CellInfect.Microbiol.2:152.doi: 10.3389/fcimb.2012.00152 Shiga toxin-producing Escherichiacoli (STEC) comprise E. coli strainscapableof producingtoxinsnamedShigatoxintype 1 (Stx1),type2(Stx2),orboth,encoded by stx1 and stx2 genes,respectively.Stx2 is themostheterogeneousgrouppossess- ing subtypesthatdifferintheirassocia- tion withhumandisease(Friedrichetal., 2002). Othertypicalvirulencefactorsare intimin (encodedby eae gene), aplasmid- encodedenterohemolysinand,instrains lacking eae, anautoagglutinatingadhesin (Saa) (PatonandPaton,2002;Blancoetal., 2004). STEC arefoodbornepathogensthat havebeenassociatedworldwidewithout- breaksandsporadiccasesofhemolytic uremicsyndrome(HUS)inchildren, STEC O157:H7beingthemostnotori- ous agentofthegroup.However,there aremorethan400non-O157serotypes that havebeeninvolvedinhumandisease andisolatedfromreservoirsofinfection (Bettelheim,2007;Rivasetal.,2010;Mora et al.,2011). Itisimportanttonotethat not allnon-O157STEChavethecapac- itytocauseHUS,whichpresentsapublic health problemtoidentifyhigh-riskSTEC. STEC O157andnon-O157havebeen isolatedfrommeat,milkanddairyprod- ucts,water,unpasteurizedappledrinks, and vegetables.Additionally,directcon- tact withcattleandanimalenviron- ment arecurrentlyconsideredasasource of transmission(EtcheverríaandPadola, 2013). Since1982whenSTECO157:H7was first reported,theselectivediagnostic methods haveusedparticularfeatures of thisserotypetoincreaseitsisolation. Because oftheuseoftheseselectivemeth- ods, theprevalenceofnon-O157STEChas beenunderestimatedduringseveralyears. Both STECO157andnon-O157STEC havealsobeendetectedalsousingmolec- ular methodsthatdonotexertselection pressuretowardanyparticularserotype, such asdetecting stx in samplesbyPCR followedbyisolationofcolonies,detection of stx1, stx2, eae, saa, ehxA bymultiplex PCR andserotyping(Scheutzetal.,2001; Fernándezetal.,2010;Piazzaetal.,2010) but thisprocedureislaboriousandtime consuming.PCRprotocolshavealsobeen developedtodetectparticularserogroups and Hantigens. During2012,FoodSafetyand InspectionService(FSIS)determinedthe zero-tolerancepolicyforSTECO157:H7 includingthetop6non-O157STEC serogroupsinraw,nonintactbeefprod- ucts (Wangetal.,2013). Becauseof this, commercialtestkitsforscreen- ing fromenrichmentculturesbasedon immunologicalandmolecularmethods havebeendevelopedtodetect stx, eae, and serogroupsfrequentlyassociatedwith severehumandisease(O157,O26,O103, O111, andO145).Theseenrichmentsam- ples couldhaveindividualbacteriaor mixtureofdifferentbacteriaharboring stx, eae, and serogroupsthatshouldbe confirmedwithisolation. A largeoutbreakthatocurredin2011in GermanywascausedbyastrainO104:H4 harboring stx2. However,thisstrainwas not atypicalSTECbecausealsocarried a genomeofenteroaggregative E. coli (EAEC) (Mellmannetal.,2011). stx genes arecarriedbylysogenicphagesandcould transfertootherbacteriabyhorizontal transfer.Forthisreason,STECnon-O157 areconsideredemergingpathogensthat rapidlyevolveandnewserogroupsofsig- nificant publichealthriskshouldbecon- sideredbydiagnosticmethods(Coombes et al.,2011). Indeed,thepathogenicityof STEC shouldnotberestrictedtoapanel of serogroupsorasinglegenesuchas eae besides stx (EFSA PanelonBiological Hazards(BIOHAZ),2013). InordertoevaluateanewPCR-real time technologybasedonsimultaneous detectionofmultipletargets, Fratamico and Bagi(2012) haveusedaGeneDisc system. TheGeneDiscconsistsofadispos- able plasticdiscofthesizeacompactdisc with36microchamberspreloadedwith desiccatedPCRprimersandfluorescence labeled geneprobesthatuseaGeneDisc cycler(Beutinetal.,2009). Fratamicoand Bagi(2012) evaluatedthistechnologyto detect stx1, stx2, eae, ehxA, and O157fol- lowedbyasecondGeneDiscassaytarget- ing eightSTECserogroup:O26,O45,O91, O103, O111,O113,O121,O145inground beef. Notably,theseauthorshaveincor- poratedtwoimportantSTECserogroups: O91 andO113thatlackthe eae gene but carry ehxA. Bothserogroupshavebeen relatedtoHUScases. Inthisstudytwoenrichmentmedia werecompared,bufferedpeptonewater (BPW)recommendedbyGeneDiscand mTSBforSTECdetectionintheground beef artificiallyinoculated.Withboth media, PCRresultsweresimilarandtarget genes werecorrectlyidentifiedwithsimilar Ct values.Immunomagneticseparation (IMS) forserogroupsO26,O45,O103, O111, O121,O145,O157,andplating ontoRainbowagarO157wereusedand isolateswereconfirmedasthecorrect STEC serogroup.SerogroupsO91and O113 werenotsubjectedtoIMSandwere moredifficulttoisolateonRainbowAgar O157. Allisolateswereconfirmedbycon- ventionalPCRtargeting stx1, stx2, and eae. These authorshavedescribedtheuseof RainbowAgarO157forisolationofnon- O157 becausenon-O157strainsproduce coloniesofdifferentcolorsaccordingto serogroup. FratamicoandBagi(2012) demon-stratedthatthistechnologyisrapid,simple touseandsuitableforscreeningground beef andotherfoods,andcouldbeadapted todetectotherserogroupsofhighriskin public health.