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Identification flow chartsBergey’s Manual of Determinative BacteriologyAll of the unknowns will fall into the following groups in Bergey's Manual of DeterminativeBacteriology (The pink book on the shelf in the laboratory).GROUP 4Description: Gram Negative, Aerobic/Microaerophilic rods and cocciKey differences are: pigments/fluorescent, motility, growth requirements, denitrification,morphology, and oxidase, read Genera descriptionsExamples: Acinetobacter, Pseudomonas, Beijerinckia, AcetobacterGROUP 5Description: Facultatively Anaerobic Gram negative rodsKey differences are: growth factors, morph., gram rxn., oxidase rxn., read Genera descriptionsExamples: Family Enterobacteriaceae and VibrionaceaeGROUP 17Description: Gram-Positive CocciKey differences are: oxygen requirements, morph., growth requirements (45 C and supplements),read Genera descriptionsExamples: Micrococcus, Staphylococcus, Streptococcus, Enterococcus, Lactococcus, AerococcusGROUP 18Description: Endospore-Forming Gram positive rods and cocciKey differences are: oxygen requirements, motility, morph, catalaseExamples: Bacillus, ClostridiumGROUP 19Description: Regular, Nonsporlating Gram positive rodsKey differences are: morph., oxygen require, catalaseExamples: Lactobacillus, ListeriaGROUP 20Description: Irregular, Nonsporlating Gram-positive rodsKey differences are: catalase, motility, morph., read Genera descriptionsExamples: Actinomyces, Corynebacterium, Arthrobacter, PropionibacteriumGROUP 21Description: Weakly Gram-Positive Nonsporlating Acid Fast Slender RodsKey differences are: acid fast, growthExamples: Mycobacterium1

Identification flow chartsDifferentiation via Gram stains and cell morphology.Gram Stain & MorphologicalFlowchartSome ExamplesGram PositiveCocciBacilliFilamentousRound in clusters& tetrads:StaphylococcusMicrococcusPeptococcusOval shape usClub-shaped and/or ycobacterium (Acid Fast)PropionobacteriumSpore-bearing,large, nomycesArachniaNocardia (partiallyacid-fast)Branching rudimentry or tomycesGram coccobacillary& acterium2Exaggeratedpointed ends:FusobacteriumnucleatumCoiled AlcaligenesChromobacterium

Identification flow chartsGram Positive Rods ID FlowchartGram Positive RodsBacillus spp.Clostrididium spp.Corynebacterium spp.Lactobacillus spp.Mycobacterium spp.Mycobacterium smegmatisSpore Forming Corynebacterium spp.Lactobacillus spp.Mycobacterium spp.Bacillus spp.Clostridium sppAcid FastStrict AnaerobesCorynebacterium sppLactobacillus spp.- Clostridium spp.You sure you have this?Go to Bergey's.Bacillus spp.See separate flowchart.Catalase -Corynebacterium spp.Lactobacillus spp.Starch HydrolysisGlucose Ferm. Activity Corynebacterium kutsceriAcidAcid & GasCorynebacterium xerosisLactobacillus fermentiLactobacillus caseiLactobacillus delbrueckiiMannitol Lactobacillus casei3Lactobacillus delbrueckii

Identification flow chartsBacillus spp. ID FlowchartBacillus spp.B. subtilisB. cereusB. megateriumB. stearothermophilusB. polymyxaB. mycoidesB. maceransB. thuringiensisB. macquariensisB. licheniformisB. lentusB. alveiB. anthracisB. alcalophilusB. coagulansB. brevisB. circulansB. firmusB. pantothenticusStarch Hydrolysis(amylase) VP B. subtilisB. cereusB. polymyxaB. mycoidesB. thuringiensisB. licheniformisB. alveiB. anthracisB. coagulans(makes insecticidalprotein)B. mycoides(round spore, restare oval)B. popilliaeB. pasteuriiB. lentimorbusB. azotoformansB. badiusB. insolitusB. larvaeB. laterosporus(add creatine as catalystw/ reagents)B. megateriumB. stearothermophilusB. maceransB. pantothenticusB. macquariensisB. lentusB. alcalophilusB. badiusB. brevisB. circulansB. popilliae-Catalase(spore is notround, rest are)-(colony has arhizoidal appearance)B. azotoformansSwollen CellSwollen Cell-B. stearothermophilusB. megaterium(growth at 55 C-rest neg.) (citrate pos.)B. maceransB. pantothenticusB. macquariensisB. lentusB. badius(citrate neg.)-B. sphaericusB. pasteuriiB. laterosporus(oval spore,rest are round)B. alcalophilusB. brevisB. circulans(Motile - It is thisone )B. larvae(gelatinase pos.)B. popilliae(gelatinase neg.)(containing spore)(catalase neg.-rest pos.)B. cereusB. insolitus(acid from glucose)B. marinus(acid from glucose)Nitrate Reduction -B. pasteuriiB. sphaericusCitrate6.5% NaCl Growth- B. coagulansB. licheniformisB. subtilis-B. insolitusB. laterosporusB. pasteuriiB. marinusB. sphaericus B. subtilisB. polymyxaB. lichenifomisB. alveiB. coagulansCitrate B. badius(containing spore) B. thuringiensisB. azotoformansB. larvaeB. pasteurii(grows at 55 C, rest no)(the width)(non-motile)(VP is pos., rest are neg.)B. marinusB. sphaericusB. schlegelii-Cell Diamenter ! 1"mB. anthracisB. pumilusB. polymyxa(manitol pos.)B. alvei(manitol neg.)- B. maceransB. macquariensisB. alcalophilusB. brevisB. circulansB. pantothenticus(acid via arabinose neg.)B. circulans(acid via arabinose pos.)6.5% NaCl Growth B. subtilis(no growth at 55 C)-Acid from Arabinose-B. coagulansB. licheniformisB. macquariensis(growth at 55 C)(methyl red pos.)B. brevis(methyl red neg.)4 B. macerans(gas from glucose pos.)B. alcalophilus(gas from glucose neg.,no growth pH 7.0)B. circulans(gas from glucose neg.,growth pH 7.0)

Identification flow chartsGram Positive Cocci ID FlowchartGram Positive CocciCatalase- Micrococcus spp.Staphylococcus spp.Streptococcus spp.(see Bergey’s)-Micrococcus spp.Staphylococcus spp.Streptococcus spp.Enterococcus spp.Bile Esculin(growth with esculin hydrolysis)Streptococcus pneumoniaeStreptococcus mitisStreptococcus pyogenes Group AStreptococcus spp. Group BEnterococcus spp.Streptococcus spp.Enterococcus faecalisEnterococcus faeciumEnterococcus hirae Enterococcus mundtiiEnterococcus spp.Streptococcus spp.Growth w/ TelluriteGamma (!)Mannitol FermentationHemolysis- Staphylococcus aureus -Enterococcus faecalisEnterococcus faecium(mannitol pos., white colony)Staphylococcus spp.Micrococcus spp.Enterococcus hirae(mannitol neg.)Enterococcus mundtii(mannitol pos., yellow colony)Beta (")Yellow Pigment (Colony)Alpha (#)Streptococcus pyogenes Group AStreptococcus spp. Group B- Micrococcus spp.Staphylococcus spp.Pyrrolidonearylamidase(PYR Test)Streptococcus pneumoniaeStreptococcus mitisNovobiocin SensitivityGlucose Fermentation Micrococcus varians YesStreptococcus pyogenes Group AStaphylococcus epidermidis- Micrococcus luteusStreptococcus spp. Group B(Streptococcus agalactiae)NoStaphylococcus saprophyticusOptochin SensitivityYesStreptococcus pneumoniae5NoStreptococcus mitis

Identification flow chartsGram Negative Rods ID FlowchartGram Negative RodsOxidase Test Enterobacteriaceae(See Separate Flowchartt)Aeromonas, Pseudomonas, Vibrio AcidVibrio spp., Aeromonas spp.Na Required for Growth-Glucose Fermentation- AcidPseudomonas spp.Fluorescent Diffusible Yellow Pigment(Fluorescein)—use Psuedo F AgarAeromonas spp.- VPVibrio spp.- A. hydrophilaA. salmonicidaLuminescentOther Aeromonas spp.(See Bergey's)(subsp. masoucida)P. aeruginosaP. chlororaphisP. cichoriiP. fluorescensP. putidaP. syringaeOther Pseudomonas spp.(See Bergey's)(Oxidase Neg.)A. sobria(no gas from glucose)A. veronii- V. fischeriV. logeiV. orientalisV. splendidiusNon-Fluorescent Diffusible Blue PigmentOther Vibrio spp.(See Bergey's)(Pyocyanin)—use Psuedo P AgarMotility-Pigment (Yellow Colony)A. hydrophilaA. veronii -V. fischeriV. logeiV. orientalisV. splendidiusA. salmonicida(subsp. masoucida)P. chlororaphisP. cichoriiP. fluorescensP. putidaP. syringae P. aeruginosaH2S ProductionNitrate Reduction A. hydrophila30 C Growth 37 C Growth V. fischeriA. veroniiP. chlororaphis V. logeiV. orientalis(Colony Pigmented)P. fluorescens-(To differentiate Biovarsdo a lecithinase test)V. splendidius6P. putida(Lecithinase Neg.)P. syringae(Oxidase Neg.)P. cichorii(Lecithinase Pos.)

Identification flow chartsFamily Enterobacteriaceae Lactose Positive ID FlowchartFamily EnterobacteriaceaeLactose Fermentation Citrobacter diversusCitrobacter freundiiEnterobacter aerogenesEnterobacter cloacaeEnterobacter amnigenusEnterobacter intermediusErwinia carotovoraErwinia chrysanthemiEscherichia coliKlebsiella oxytocaKlebsiella pneumoniaeSee Lactose Negative Flowchartfor Enterobacteriaceae(subsp. ozaenae and pneumoniae)Serratia fonticolaSerratia rubidaeaIndoleThe Rest Citrobacter diversusEscherichia coliErwinia chrysanthemiKlebsiella oxytocaMR-VP / -/ Klebsiella pneumoniae(subsp. pneumoniae)Enterobacter intermedius /Citrobacter freundiiSerratia fonticolaKlebsiella pneumoniaeCitrateEnterobacter aerogenesEnterobacter cloacaeEnterobacter amnigenusErwinia carotovoraSerratia rubidaea(subsp. ozaenae)Lysine Decarboxylase-Citrobacter diversusEscherichia coliKlebsiella pneumoniae subsp. pneumoniaeEnterobacter cloacaeSerratia rubidaeaEnterobacter amnigenusNon-motile, No Pigment(VP neg.)Erwinia chrysanthemiAcid from sorbitolMotile, Red pigment(VP and H2S pos.)(VP pos and H2S neg.)No acid from sorbitolEnterobacter aerogenesH2S ProductionKlebsiella oxytoca- Motile, No pigment?Erwinia carotovoraLysine unknown but Gelatinasepositive; none above are positive.- Citrobacter freundiiSerratia fonticolaMotility pos.Klebsiella pneumoniae subsp. ozaenaeMotility neg.7

Identification flow chartsFamily Enterobacteriaceae Lactose Negative ID FlowchartFamily Enterobacteriaceae ContinuedLactose Negative Flow ChartEdwardsiella tardaErwinia cacticidaMorganella morganiiProteus mirabilisProteus penneriProteus vulgarisProvidencia stuartiiSalmonella bongoriSalmonella entericaEdwardsiella tardaMorganella morganiiProteus vulgarisProvidencia stuartiiSerratia marcescensSerratia liquefaciensShigella spp. (boydii, dysenteriae, flexneri)Shigella sonneiYersinia enterocoliticaYersinia pestisYersinia pseudotuberculosisErwinia cacticidaProteus mirabilisProteus penneriSalmonella bongoriSalmonella entericaSerratia marcescensSerratia liquefaciensShigella sonneiYersinia pestisYersinia pseudotuberculosis- Indole TestUrease ?Please See Bergey’s to ID:Shigella spp.Proteus mirabilisYersinia enterocoliticaProteus penneriErwinia cacticidaSalmonella bongoriSalmonella entericaSerratia marcescensSerratia liquefaciensShigella sonneiYersinia pestisOrnithine decarb. pos.(boydii, dysenteriae, flexneri)Ornithine decarb. neg., motileH2SYersinia pseudotuberculosisOrnithine decarb. neg., non-motile- Edwardsiella tardaProteus vulgarisMorganella morganiiProvidencia stuartiiMotility -Erwinia cacticidaShigella sonneiYersinia pestis(lysine decarb. neg., the rest pos.)UreaseSalmonella bongoriSalmonella choleraesuisSerratia marcescensSerratia liquefaciensOrnithine Decarboxylase Proteus vulgarisProvidencia stuartii-Edwardsiella tardaOrnithine DecarboxylaseMorganella morganiiH2S - Salmonella bongoriSerratia marcescensSalmonella entericaSerratia liquefaciensKCN growth pos.KCN growth neg.Red pigmentNo Red pigment(It is this one)8Shigella sonneiYersinia pestis

Bergey’s Manual of Determinative Bacteriology All of the unknowns will fall into the following groups in Bergey's Manual of Determinative Bacteriology (The pink book on the shelf in the laboratory). GROUP 4 Descripti