美国FDA细菌学分析手册第八版沙门氏菌.docx
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美国FDA细菌学分析手册第八版沙门氏菌.docx
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美国FDA细菌学分析手册第八版沙门氏菌
BAM:
Salmonella
NOTICE:
IfyouarelookingforBAMChapter5:
Salmonella(December2007Edition)thatisincorporatedbyreferencein21CFRParts16and118:
FederalRegisterFinalRule1(July9,2009,74FR33030):
PreventionofSalmonellaEnteritidisinShellEggsDuringProduction,Storage,andTransportation,pleaseusethese versionsoftheBAM SalmonellaChapter2(PDF,189Kb) and Appendix1:
RapidMethodsforDetectingFoodbornePathogens3(PDF,195Kb).
Themostrecent EditionofBAMChapter5:
Salmonellaisavailablebelowthisnotice.
November2011Version
BacteriologicalAnalyticalManual
Chapter5
Salmonella
Authors:
WallaceH.AndrewsandThomasHammack
RevisionHistory:
∙November2011-AdditiontoSectionC:
PreparationoffoodsforisolationofSalmonella:
Leafygreenvegetablesandherbs.
∙February2011– RemovedlinktoAppendix1:
RapidMethodsforDetectingFoodbornePathogens(nowarchived).
∙December2007–Mameypulpmethodadded,andSectionDrevised.
∙June2006 –Eggsmethodrevisedforshelleggsandliquidwholeeggs.
∙April2003–Froglegsmethod,Lacticcasein,Rennetcasein,SodiumcaseinateandRabbitcarcassmethodsrevised,topearsandotherdogchewtoysadded.RemovedsectionA.25,Mechanicalshaker.
∙October25,2001–ExtensionoftheapplicabilityoftheorangejuicemethodinsectionC.19toapplejuiceandapplecider.
∙1999-DEC,2000-MAR,and2000-AUGFinalrevisionon2000-NOV-14(seetheIntroductionforasummaryofchanges).
Toobtainacopyofapriorversionnotcurrentlyposted,pleasecontactThomasHammack
ChapterContents
∙Introduction
∙EquipmentandMaterials
∙MediaandReagents
∙PreparationoffoodsforisolationofSalmonella
∙IsolationofSalmonella
∙IdentificationofSalmonella
∙References
Introduction
SeveralchangesarebeingintroducedinthiseditionofBAM(8thEdition).ThefirstchangeinvolvestheexpandeduseofRappaport-Vassiliadis(RV)medium4forfoodswithbothhighandlowlevelsofcompetitivemicroflora.Inthepreviousedition,RVmediumwasrecommendedonlyfortheanalysisofshrimp.BasedonthecompletionofAOACprecollaborative(5,6)andcollaborative(7,8)studies,RVmediumisnowbeingrecommendedfortheanalysisofhighmicrobialandlowmicrobialloadfoods.RVmediumreplacesselenitecystine(SC)brothfortheanalysisofallfoods,exceptguargum.Inaddition,RVmediumreplaceslauryltryptosebrothforusewithdryactiveyeast.Tetrathionate(TT)5brothcontinuestobeusedasthesecondselectiveenrichmentbroth.However,TTbrothistobeincubatedat43°Cfortheanalysisofhighmicrobialloadfoodsandat35°Cfortheanalysisoflowmicrobialloadfoods,includingguargum.
Thesecondchangeinvolvestheoptionofrefrigeratingincubatedpreenrichmentsandselectiveenrichmentsoflow-moisturefoodsforupto72h.Withthisoption,sampleanalysescanbeinitiatedaslateasWednesdayorThursdaywithoutweekendworkbeinginvolved.
Thethirdchangeinvolvesreducingtheperiodofincubationofthelysineironagar(LIA)6slants.Intheformeredition(BAM-7),triplesugarironagar(TSI)7andLIAslantswereincubatedat35°Cfor24±2hand48±2h,respectively.Unpublisheddatahavedemonstratedthatthe48hreadingofLIAslantsiswithoutdiagnosticvalue.Of193LIAslantsexamined,allgavedefinitiveresultswithin24±2hofincubation.Nosignificantchangesalteredthefinaltestresultwhentheslantswereincubatedanadditional24h.Thus,boththeTSIandLIAslantsarenowincubatedfor24±2h.
Thefourthchangeinvolvestheprocedureforsurfacedisinfectionofshelleggs.Inthepreviousedition(BAM-7),eggshellsweresurface-disinfectedbysoakingin0.1%mercuricchloridesolutionfor1hfollowedbysoakingin70%ethanolfor30min.Mercuricchlorideisclassifiedasahazardouswaste,andisexpensivetodisposeofaccordingtoEnvironmentalProtectionAgencyguidelines.Inthisedition(BAM-8)eggshellsarenowsurface-disinfectedbysoakingforatleast10secina3:
1solutionconsistingof3partsof70%alcohol(ethylorisopropyl)to1partofiodine/potassiumiodidesolution.
Thefifthchangeinvolvesthesamplepreparationofeggs.Eggcontents(yolkandalbumen)arethoroughlymixedbeforeanalysis.Aftermixingtheeggcontents,25g(ml)areaddedto225mltrypticase(tryptic)soybrothsupplementedwithferroussulfate.
Amethodfortheanalysisofguargumhasbeenincluded.Whenguargumispreenrichedata1:
9sample/brothratio,ahighlyviscous,nonpipettablemixtureresults.Additionoftheenzymecellulasetothepreenrichmentmedium,however,resultsinareadilypipettablemixture.
Amethodfororangejuice(pasteurizedandunpasteurized)hasbeenincludedduetorecentorangejuice-relatedoutbreaks.
Thedirectionsforpickingcoloniesfromtheselectiveplatingagarshavebeenmademoreexplicittoreflecttheintentofthemethod.Intheabsenceoftypicalorsuspectcoloniesontheselectiveplatingagars,itisrecommendedthatatypicalcoloniesbepickedtoTSIandLIAslants.Thisrecommendationisbasedonthefactthatupto4%ofallSalmonellaculturesisolatedbyFDAanalystsfromcertainfoods,especiallyseafoods,duringthepastseveralyearshavebeenatypical.
Finally,sincethepublicationofBAM-7,a6-waycomparisonwasconductedoftherelativeeffectivenessofthethreeselectiveplatingagarsrecommendedintheBAM(bismuthsulfite8,Hektoenenteric9,andxyloselysinedesoxycholateagars10)andthreerelativelynewagars(EF-18,xyloselysineTergitol4,andRambachagars).Ourresults(9)indicatednoadvantageinreplacinganyoftheBAM-recommendedagarswithoneormoreoftheneweragars.Thus,thecombinationofselectiveplatingagarsrecommendedinBAM-7remainsunchanged.
A.EquipmentandMaterials
1.Blenderandsterileblenderjars(seeChapter111)
2.Sterile,16oz(500ml)wide-mouth,screw-capjars,sterile500mlErlenmeyerflasks,sterile250mlbeakers,sterileglassorpaperfunnelsofappropriatesize,and,optionally,containersofappropriatecapacitytoaccommodatecompositedsamples
3.Sterile,bentglassorplasticspreaderrods
4.Balance,withweights;2000gcapacity,sensitivityof0.1g
5.Balance,withweights;120gcapacity,sensitivityof5mg
6.Incubator,35±2°C
7.Refrigeratedincubatororlaboratoryrefrigerator,4±2°C
8.Waterbath,49±1°C
9.Waterbath,circulating,thermostatically-controlled,43±0.2°C
10.Waterbath,circulating,thermostatically-controlled,42±0.2°C
11.Sterilespoonsorotherappropriateinstrumentsfortransferringfoodsamples
12.Sterileculturedishes,15x100mm,glassorplastic
13.Sterilepipets,1ml,with0.01mlgraduations;5and10ml,with0.1mlgraduations
14.Inoculatingneedleandinoculatingloop(about3mmidor105l),nichrome,platinum-iridium,chromelwire,orsterileplastic
15.Steriletestorculturetubes,16x150mmand20x150mm;serologicaltubes,10x75mmor13x100mm
16.Testorculturetuberacks
17.Vortexmixer
18.Sterileshears,largescissors,scalpel,andforceps
19.Lamp(forobservingserologicalreactions)
20.FisherorBunsenburner
21.pHtestpaper(pHrange6-8)withmaximumgraduationsof0.4pHunitspercolorchange
22.pHmeter
23. Plasticbags,28x37cm,sterile,withresealabletape.(Items23-24areneededintheanalysisoffroglegsandrabbitcarcasses.)
24.Plasticbeakers,4liter,autoclavable,forholdingplasticbagduringshakingandincubation.
25.Sponges,non-bactericidal(Nascocat#B01299WA),orequivalent.
26.Swabs,non-bactericidal,cotton-tipped.
B. Media12andReagents13
-967.28inOfficialMethodsofAnalysis
(1).
1.Lactosebroth(M7414)
2.Nonfatdrymilk(reconstituted)(M11115)
3.Selenitecystine(SC)broth(M13416)
4.Tetrathionate(TT)broth(M14517)
5.Rappaport-Vassiliadis(RV)medium(M13218).NOTE:
RVmediummustbemadefromitsindividualingredients.Commercialformulationsarenotacceptable.
6.Xyloselysinedesoxycholate(XLD)agar(M17919)
7.Hektoenenteric(HE)agar(M6120)
8.Bismuthsulfite(BS)agar(M1921)
9.Triplesugarironagar(TSI)(M14922)
10.Tryptone(tryptophane)broth(M16423)
11.Trypticase(tryptic)soybroth(M15424)
12.Trypticasesoybrothwithferroussulfate(M18625)
13.Trypticasesoy-tryptosebroth(M16026)
14.MR-VPbroth(M10427)
15.Simmonscitrateagar(M13828)
16.Ureabroth(M17129)
17.Ureabroth(rapid)(M17230)
18.Malonatebroth(M9231)
19.Lysineironagar(LIA)(EdwardsandFife)(M8932)
20.Lysinedecarboxylasebroth(M87)33
21.Motilitytestmedium(semisolid)(M10334)
22.Potassiumcyanide(KCN)broth(M12635)
23.Phenolredcarbohydratebroth(M12136)
24.Purplecarbohydratebroth(M13037)
25.MacConkeyagar(M9138)
26.Nutrientbroth(M11439)
27.Brainheartinfusion(BHI)broth(M2440)
28.Papainsolution,5%(M56a41)
29.Cellulasesolution,1%(M18742)
30.Tryptosebloodagarbase(M16643)
31.Universalpreenrichmentbroth(M18844)
32.Universalpreenrichmentbroth(withoutferricammoniumcitrate)(M188a45)
33.Bufferedpeptonewater(M19246)
34.Dey-Engleybroth(M19347)
35.Potassiumsulfitepowder,anhydrous
36.Chlorinesolution,200ppm,containing0.1%sodiumdodecylsulfate(R12a48)
37.Ethanol,70%(R2349)
38.Kovacs'reagent(R3850)
39.Voges-Proskauer(VP)testreagents(R8951)
40.Creatinephosphatecrystals
41.Potassiumhydroxidesolution,40%(R6552)
42.1NSodiumhydroxidesolution(R7353)
43.1NHydrochloricacid(R3654)
44.Brilliantgreendyesolution,1%(R855)
45.Bromcresolpurpledyesolution,0.2%(R956)
46.Methylredindicator(R4457)
47.Steriledistilledwater
48.TergitolAnionic7(R7858)
49.TritonX-100(R8659)
50.Phys
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