Bacteriology Washington - jb.asm.org

1 ,WashingtonUniversitySchoolofMedicine, ,MissouriReceivedforpublication,December 2,1935In1918 BronfenbrennerandDavisobservedthatintest inalbacteriawerefrequentlyencounteredwhi chfailedtoshowupaslactosefermentersinthe routinediagnosticprocedure, ,withthepurposeinviewofimprovingmethodsf ortheirdetection, (1906)andMassini(1907), (1934),Sandiford(1935),and,withspecialre ferencetobacterialvariation,byLewis(1934 )andHall(1935).Inthecourseofcontinuedstu diesofthemetabolismofslowlactose-ferment ingbacteriaoftheEscherichiacolitype, ( )indicator(Bronfen-brenner,1918).Ofsixsa mplesexamined,slowlactose-fer-mentingorg anismswereobtainedfromone, on February 26, 2021 by from , ,theyweresubjectedtodailyserialtransferi nalactosemediumasinthemethodprevi-ouslyu sed(BronfenbrennerandDavis,1918).Someela bora-tionsoftechnicwereintroducedtofacil itatethestudyofcondi-tionsaffectingthech angefromslowtorapidfermentation, ,oneseriesconsistingofopentubespluggedwi thcotton, ,onecultureineachserieswasusedtoinoculat ethesubcultures, ,transfersfromtheopentubecul-tureseriesw eremadeintosealedfermentationtubestoobse rveanychangesinfermentationreactions, ,correspondingtosubcultureIofeachseries( table1).

2 Ob-servedincreasesintheabilityofthecultu restoformacidandgasinlactosesubculture, , ,thefermentingpower,andthepro-portionoff ermentingvariantspresent, ,butwereneverasmarked, on February 26, 2021 by from SLOWLACTOSE-FERMENTINGINTESTINALBACTERIA (inpercentofthetotalcolonies)ontheplates madefromsub-cultureIat1day,10days,and20d ays,andfromsubcultureXXafter24hours,fore achofthetwoseriesofcultures, ,itappearsthatwhilevariationoccursaftera bout12days, ,0-10 DATSSUBCuLTUEI,20 DATSSUBCULTUREXX,1 DATSTRAIN(PMnCENTBLUECOLONIs)(PRCENTBLUE COLON1ES(PERCENTBLUECOLONIES) :(1)1percentpeptone, ;(2) , , ;(3)3percentlactoseaddedtothesamebase;(4 )1percentsodiumcitrateaddedtothesamebase ;(5)1percentsodiumsuccinateaddedtothesam ebase;(6)1percentlactoseand1percentsodiu msuccinatecombinedinthesamebase;and(7)1p ercentlactose, on February 26, 2021 by from , , , (1) ++44(2)Lactose1percent(noadditionalcarbo n).)

3 +++++X(3)Lactose3percent(noadditionalcar bon)..+++++(4) ++_(5) ++_(6) +++++(7) +++++ ,colonialvariantswereisolatedfromanumber oftheplatesmadeduringthecourseoftheprece dingexperi-ments, ,largemucoid,smalltranslucent, , on February 26, 2021 by from SLOWLACTOSE-FERMENTINGINTESTINALBACTERIA 457colonialform,andrateoflactosefermenta tion,alldissociantsappearedtobeidentical withoneanother, ,gram-negativerods, , ,andfailedtoproduceindolorH2S, , ,galactose,mannitol, ,therefore,whetherattackinglactoseslowly orrapidly,andregardlessoftheircolonialty pe, ,andinproducingnoindol, (1935). , (1935)andStewart(1928) "regained"thecapacitytofermentlactoseact ively, ,inourexperiments,byserialcultivationont heabovedescribedsyntheticmediumwithsodiu msuccinateastheonlysourceofcarbon, , ,bothcultureswerethencarriedthroughaseri esof20dailysubculturesinlactosefer- on February 26, 2021 by from , ,thecultureslowlyregainsthispowerdurings ubsequentserialcultivationonlactose, +50+ |50+VI0-50+Subsequentlyon10+0+ +50+ (1935)hasdefinedthecriterionoflactosemut ationas" ,"andotherinvestigators(Rriebel,1934.)

4 Stewart,1928) ,butafewsinglecellstrains,afterrepeatedp late-isolation,failedtoshowanytendencyto changeintheirlactose-fermentingpower, on February 26, 2021 by from SLOWLACTOSE-FERMENTINGINTESTINALBACTERIA 459largenumberofwhitecoloniesappearingon thelactoseC-Rplatestreakedfromadissociat edcultureoftheparentstrain,andwerepurifi edbyrepeatedplating, , ,thecontrolstrain,theoriginalsinglecells trains,andorganismsfromthetwoseriesoflac tosecultures, , ,andbothblueandwhitecoloniesappearedTABL E4 Lactosefermentationstabilityofaselecteds trainduringdissociationinlactosemediumAM OUNTOFGASANDACID(FORMED) + +inmostcases,afewstrainsliketheonedescri bedintable4, ,andthecontrolstrain, , ,andthatdaughterstrainsmaybeselectedinwh ichthecapacitytofermentlactoseismoreradi callyandperhapsper-manentlydepressed.;Tw oseriesofexperimentswerenowcarriedoutina nefforttodetermineinwhatparticularthebio chemicalmechanismsofslow on February 26, 2021 by from , ,non-proliferatingcellsinthepresenceofla ctose,andofitsconstituentmonosaccharides , ,asanticipated,anactualdifferencecouldbe seeninthecapacityofthecellularenzymesoft hevariousstrainstofermentlactose,andtoex cludethepossibilitythat"secreted" ,itwasreasonedthat,iflactosefermentation bytheseorgsmisaquantitativefunctionmerel yoftheirlactaseactivity,then,supposingth eirmono-saccharide-fermentingpowertobero ughlyequal, , ,: ( ) , , , wasmadeofthefactthattoluolinhibitsoxidat ionprocesseswithoutdestroyinghydrolyticf unction(Karstnom,1930).

5 AAA on February 26, 2021 by from ,aslowlactose-fermentingstrain,andarapid lyfermentingvariantofthelatter,respectiv ely, , ,equivalentamountsofsuper-natantfluidobt ainedbycentrifugationofthefreshcellsus-p ension,andofthesamesuspensionafterautoly zingtwodaysundertoluol, ( ) ; ; ; ; ; ; , ; , , , , , , ,afterwhichnoviablecellsremained,andoxid aseactiv-itywasdestroyed. on February 26, 2021 by from (1)overitscontroltube(3), (2)and(3).Comparisonoftubes(4)and(5)show edtheamountofglucosefernentedundertoluol , ,cells, , , ,cells, , , , , , ~~~~~~~ (5)and(6),givingtheamountofglucosefermen tedintheabsenceoftoluol, (7)and(8) ,whichwereincludedtodetectanysolublelact asepresent, ,itisseenthatslowlactose-fermentingstrai nsaredefi-cientintheenzymelactase,andtha tincreaseinfermenting462 on February 26, 2021 by from ,inconclusion,thatvariationsinlactaseact ivitydescribedthroughoutthispaperaremere lyquan-titativeandthatinnoinstanceundero urobservationhavevariantsshownanymetabol icpropertywhichwaswhollyabsentintheparen torganism(Diehl,1919).

6 Thus,thephenomenasuggestingspecificelabo rationofenzymescorrespondingtoaTABLE7 Simultaneouslactosefermentationandlactas eactivityexpressedingramspercentofsugarc hangedCONTROLCONTROL(GLUCOSE(GLUCOSELACT OSELACTOSEENZYMICPREBPARATIONTIMEFERMENT EDFER-METRDLYDRO-UNDERMENTED)NDLZETOLUOL ) ,perhapslatent, ,butnodirectcorrelationbetweencolonialty pe,andfermentativecapacitywasobserved. on February 26, 2021 by from , lactosebythebacteriastudiedisthefunction ofanintracellularlactase, , ,39, , ,ANDDAVIS, ,89, , ,24, , ,29, , ,231,3W.]KIEBEL, ,27, , ,28, , ,61, , , ,Beiheft,36, , ,41, , ,27, on February 26, 2021 by from