I.

OVERVIEW OF BACTERIAL IDENTIFICATION METHODS AND STRATEGIES

TEST PURPOSE EXPECTED RESULTS POSITIVE CONTROL NEGATIVE CONTROL OTHERS
Acetamide utilization Differentiate based on the ability to use acetamide (+) Blue Pseudomonas aeruginosa Escherichia coli enzyme acylamidase
as the sole source of carbon. (-) Green ammonia=alkalinity
Acetate utilization Generally used to differentiate Shigella sp. from (+) Blue Escherichia coli Shigella sonnei
Escherichia coli. (-) Green
Bacitracin test Streptococcus pyogenes vs other beta-hemolytic (+) Any zone of inhibition Streptococcus pyogenes Streptococcus agalactiae Taxo A
streptococci. >10 mm 0.04 Units
(-) No zone
Bile Esculin agar enterococci and group D streptococci vs. (+) Blackening of agar Enterococcus faecalis Escherichia coli 40% Bile
non–group D viridans streptococci. (-) No blackening Indicator: Ferric ammonium citrate
Bile Solubility test Streptococcus pneumoniae Vs alpha hemolytic (+) Colony disintegrates Streptococcus pneumoniae Enterococcus faecalis Reagent= 10% Sodium desoxycholate
streptococci (-) Intact colonies Amidase= intracellular autolytic enzyme
Butyrate disk identification of Moraxella (Branhamella) (+) Blue color Moraxella catarrhalis Neisseria gonorrhoeae Reagent= Bromo-chloro-indolyl butyrate
catarrhalis (-) No color change enzyme butyrate esterase
CAMP test Streptococcus agalactiae vs. other streptococcal (+) Arrowhead zone of Streptococcus agalactiae Streptococcus pyogenes Other CAMP (+)
Christie species beta-hemolysis L. monocytogenes
Atkins (-) No enhancement of Reverse CAMP (+)
Munch hemolysis A. haemolyticum C. ulcerans
Peterson C. perfringens C. pseudotuberculosis
Catalase test Micrococcus and staph vs strep (+) Bubble formation Staphylococcus aureus Streptococcus pyogenes Reagent= 30% H2O2
(-) No or few bubbles Don’t use colonies from BAP= false (+)
Cetrimide isolate and purify Pseudomonas aeruginosa from (+) Growth Pseudomonas aeruginosa Escherichia coli
contaminated specimens. (-) No growth
Citrate utilization identify organisms capable of using sodium citrate (+) Growth w/ or w/o Enterobacter aerogenes Escherichia coli Medium: Simmon’s citrate agar
as the sole carbon source and inorganic ammonium change in color from green Indicator: Bromthymol blue
to blue Enzyme: citrate permease
(-) No growth
Coagulase test S. aureus vs. CONS a. Slide test Staphylococcus aureus Staphylococcus epidermidis Slide test: clumping factor/ bound coagulase
(+) Clumping Tube test: free coagulase
(-) No clumping Reagent= Rabbit EDTA plasma
b. Tube test
(+) Clot formation
(-) No clot
Decarboxylase test differentiate decarboxylase producing (+) Alkaline (purple color) Lysine Lysine Lysine- Cadaverine
(Moeller’s method) Enterobacteriaceae from other gram negative rods (-) Acid (yellow color) Klebsiella pneumoniae Enterobacter cloacae Ornithine Putrescine
Ornithine Ornithine Arginine Citrulline
Enterobacter cloacae Klebsiella pneumoniae
Arginine Arginine
Enterobacter cloacae Klebsiella pneumoniae
DNA Hydrolysis distinguish Serratia sp. from Enterobacter sp., S. (+) Colorless Staphylococcus aureus Staphylococcus epidermidis Medium: DNase agar
(DNase) aureus (positive) from other species, and M. (-) Green Serratia marcescens
catarrhalis (positive) from Neisseria sp.

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

Esculin Hydrolysis presumptive identification (+) Blackened medium Enterococcus faecalis Escherichia coli Enzyme: deoxyribonuclease.
and differentiation of Enterobacteriaceae. and loss of fluorescence
under Wood’s lamp
(-) No blackening and no
loss of fluorescence under
Wood’s lamp

TEST PURPOSE EXPECTED RESULTS POSITIVE CONTROL NEGATIVE CONTROL OTHERS

Fermentation Media differentiate a. Peptone medium a. Peptone medium a. Peptone medium = Peptone medium:
a. Peptone medium organisms based on their ability (+) Pink w/ or w/o gas formation w/ gas: Escherichia coli Pseudomonas aeruginosa Indicator: Andrade’s indicator
b. Heart Infusion to ferment carbohydrates (-) Growth but no change in color (Straw) w/o gas: Shigella flexneri =Heart infusion broth:
incorporated into the basal b. Heart infusion broth b. Brain-Heart infusion broth b. Brain-Heart infusion broth Indicator: Bromcresol purple
medium. (+) Yellow Escherichia coli Moraxella osloensis
(-) Growth but no change in color (purple)
Flagella Stain (+) Flagella Peritrichous: Klebsiella pneumoniae
(Wet-Mount technique) a. Peritrichous Escherichia coli
b. Lophotrichous Polar:
c. Polar Pseudomonas aeruginosa
Gelatin Hydrolysis Staphylococcus sp., (+) Partial or total liquefaction at 4’C within 14 Bacillus subtilis Escherichia coli enzyme: gelatinase
Enterobacteriaceae, days
and some gram-positive bacilli (-) Complete solidification at 4’C
Growth at 42’C differentiate a pyocyanogenic (+) Good growth at both 35’C and 24’C Pseudomonas aeruginosa Pseudomonas fluorescens
pseudomonads from other (-) No growth at 42’C but good growth at 35’C
Pseudomonas sp
Hippurate Hydrolysis enzyme hippuricaseused for (+) Deep purple color Streptococcus agalactiae Streptococcus pyogenes Other HH (+)= L. monocytogenes, C. jejuni
presumptive ID of diff org (-) Colorless or slightly yellow pink color G. vaginalis
hippuric acid to benzoic acid and glycine
(detected by Ninhydrin reagent)
Indole production identify organisms that (+) Pink- to wine-colored ring a. Kovac’s method a. Kovac’s method
a. Kovac’s method produce the enzyme (-) No color change Escherichia coli Klebsiella pneumoniae
b. Ehrlich’s method tryptophanase b. Ehrlich’s method b. Ehrlich’s method
Haemophilus influenza Haemophilus parainfluenzae
c. Ehrlich’s method (anaerobic) c. Ehrlich’s method (anaerobic)
P. asaccharolytica Bacteroides fragilis
LAP test presumptive identification (+) Red color Enterococcus faecalis Aerococcus viridans Reagent: cinnamaldehyde
of catalase-negative gram- (-) No color change or development of slight
positive cocci. yellow color
Litmus milk differentiates microorganisms Color of Indicator: Fermentation:
based on various metabolic Acid: Pink, mauve Clostridium perfringens
reactions in litmus milk, Alkaline: Blue
including fermentation, No change: Purple Acid:
reduction, clot formation, White: Reduction of indicator Lactobacillus acidophilus
digestion, and the formation of Consistency of Milk
gas. Clot/Coagulation: Acid or Alkaline pH Peptonization:
Digestion: Dissolution of clot w/ shrunken, Pseudomonas aeruginosa
insoluble pink clot (acid)

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 Peptonization: Dissolution of clot w/ shrunken,
insoluble blue clot (alk)

Lysine Iron Agar differentiate gram-negative K/K: Alkaline slant and butt: H2S (+): Decarboxylation: Butt (Purple)
bacilli based on decarboxylation (-) Glucose fermentation Citrobacter freundii Deamination: Slant (Red)
or deamination (+) Decarboxylation LIA indicator: Bromocresol purple,
of lysine and the formation of K/A: Alkaline slant and butt: H2S indicator: Ferric ammonium citrate
hydrogen sulfide (+) Glucose fermentation Escherichia coli
(H2S) (-) Decarboxylation
K/A or K/K w/ H2S: Alkaline slant and butt: H2S (+)
(+) Black precipitate (FeS) Salmonella typhimurium
R/A:
(+)Deamination Red slant, acid butt:
(+) Glucose fermentation Proteus mirabilis

TEST PURPOSE EXPECTED RESULTS POSITIVE CONTROL NEGATIVE CONTROL OTHERS

Methyl Red test differentiates members (+) Bright red color MR positive/VP negative: MR negative:/VP positive: MR detects mixed acid fermentation
of the Enterobacteriaceae (W+) Red-orange color Escherichia coli Enterobacter aerogenes Indicator: Methyl red
family (-) Yellow
Voges-Proskauer (+) Red color VP detects acetoin/acetylmethylcarbinol formation
(-) Yellow color Reagents
a. Barritt’s Method: α-naphthol + KOH
b. Coblentz Method: α-naphthol + 40% KOH w/ creatine
Microdase test differentiate gram-positive, (+) Blue to purple-blue color Micrococcus luteus Staphylococcus aureus Modified oxidase rgt:
catalase-positive cocci (-) No color change Tetramethyl-p-phenylenediamine dihydrochloride in
(micrococci from dimethylsulfoxide
staphylococci)
Motility testing determine whether an a. Semisolid agar deep Escherichia coli Klebsiella pneumoniae
enteric organism is motile. An (+) Motile: spread out into the medium
organism must from the site of inoculation
have flagella to be motile. (-) Nonmotile: remain at the site of
inoculation
MRS Broth determine whether an (+) Gas production Lactobacillus lactis Escherichia coli Durham tube detects gas formation
organism forms gas during (-) No gas production (+) bubbles
glucose fermentation. Some Lactobacillus spp. and Leuconostoc
sp. produce gas
MUG test presumptively identify (+) Electric blue fluorescence Escherichia coli Klebsiella pneumoniae Uses 366nm UV light
4-methylumbelliferyl-beta- various genera of (-) Lack of fluorescence
D-glucuronide Enterobacteriaceae and
verotoxin-producing E.coli.

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

Nitrate reduction All members of the After addition of solutions A and B: NO3+, no gas: Acinetobacter baumannii Reagents:
Enterobacteriaceae family (+) Red color Escherichia coli A: Sulfanilic acid
reduce nitrate, but some (-) Colorless NO3+, w/ gas: B: Alpha-naphthylamine
members further If colorless, add Zinc dust: Pseudomonas aeruginosa
metabolize nitrite to other (+) Colorless (no change in color) Zinc dust/powder= use to detect unreduced nitrate
compounds. (-) Red color (do not report)
Nitrite reduction Nitrtenitrogen gas (+) Colorless w/ gas Proteus mirabilis Acinetobacter baumannii
(-) Red w/o gas production
ONPG distinguishes LF from NLF of (+) Yellow = orthonitrophenol. Shigella sonnei Salmonella typhimurium determine the ability of an organism to produce β-
O-Nitrophenyl-beta-D- Enterobacteriaceae (-) Colorless galactosidase
galactopyranoside
Optochin test Optochin lyses pneumococci, (+) Zone of inhibition >14mm Streptococcus pneumoniae Streptococcus pyogenes Medium: 5% sheep BAP
Optochin/Taxo P= but alpha-streptococci are (-) No zone of inhibition
ethyl hydrocupreine resistant Equivocal: <14 mm zone of inhibition
hydrochloride (perform bile solubility test)
Oxidase test determines the presence of (+) Dark purple color within 10 secs Pseudomonas aeruginosa Escherichia coli Oxidase reagent
(Kovac’s method) cytochrome oxidase activity in (-) Absence of color Tetramethyl-p-phenylenediamine dihydrochloride
microorganisms for Other oxidase +  Colony
the identification of oxidase-  Filter paper
negativeEnterobacteriaceae,
PPAM ViC HeN  Filter paper disk
Pseudomonas, Plesiomonas,
Aeromonas, Vibrio,
Campylobacter, Helicobacter,
Neisseria

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 TEST PURPOSE EXPECTED RESULTS POSITIVE CONTROL NEGATIVE CONTROL OTHERS
Oxidation/Fermentation (OF) differentiate microorganisms (+) Yellow (Acid deep) Fermenter: Nonutilizer:
medium based on the ability to (W+) Deeper red (Alk) Escherichia coli Alcaligenes faecalis
(CDC method) oxidize or ferment (-) Red (alkaline deep) Oxidizer:
specific carbohydrates (NC or N): No change or Neutral, there is Pseudomonas aeruginosa
growth in the media
(NG): No growth
Phenylalanine Deaminase determine the ability of (+) Green color Proteus mirabilis Escherichia coli Rgt: 10% Ferric chloride
an organism to oxidatively (-) No color change (+) PMP
deaminate phenylalanine
to phenylpyruvic acid
PYR test for the presumptive (+) Bright red color Enterococcus faecalis Streptococcus agalactiae PYR: L-pyrrolidonyl-beta-naphthylamide
(PYRase) identification (-) No color or an orange color enzyme L-pyrrolidonyl arylamidase
of group A streptococci Reagent: N,N-Methylaminocinnamaldehyde
and enterococci
Pyruvate broth aids in the (+) Yellow Enterococcus faecalis Streptococcus bovis
differentiation between (-) No color change or yellow-Green
Enterococcus faecalis
and Enterococcus faecium
Salt Tolerance test differentiate enterococci (+) Visible turbidity w/ or w/o color Enterococcus faecalis Streptococcus bovis Indicator: Bromcresol purple
(positive) from change from purple to yellow Concentartion: 6.5% NaCl
nonenterococci (negative). (-) No turbidity and no color change
Spot Indole test rapid method (+) Blue color Escherichia coli Klebsiella pneumoniae Reagent= 1% paradimethylamino-cinnamaldehyde
that can be used in lieu of the (-) No color or slightly pink color
tube test

Triple Sugar Iron agar determine whether a K/K: A/A w/ gas: K/K: Peptone
(TSI) gramnegative Glucose, lactose and sucrose nonutilizer Escherichia coli Pseudomonas aeruginosa Indicators:
rod ferments glucose and K/A: K/A, +/− gas production, H2S+: pH: phenol red
1 part glucose lactose or Glucose fermentation only Salmonella typhimurium H2S: ferrous sulfate
10 parts lactose sucrose and forms hydrogen A/A: K/A, H2S+: Proteus mirabilis
10 parts sucrose sulfide (H2S) Glucose, sucrose, and/or lactose fermenter K/A: Shigella flexneri
Black butt ppt indicates: H2S production
Bubbles/cracks: CO2 or H2/gas prod

Urea Hydrolysis determine an organism’s (+)Change in color of slant from light Proteus vulgaris Escherichia coli Indicator: phenol red
(Christensen’s method) ability to produce the orange to magenta Weak positive: Klebsiella
enzyme urease, which (-) No color change pneumoniae
hydrolyzes urea.
X and V factor test used for differentiation of (+) Growth: Around XV disk: Haemophilus aphrophilus X factor: hemin/hematin
Haemophilus species Growth around XV disk= Haemophilus influenzae V factor: NAD
requirement for both factors Around XV and V disks: Medium: TSB
Growth around V disk, no growth around Haemophilus parainfluenzae
X disk, and light growth around XV disk: Haemophilus ducreyi :
=V factor requirement halo ofgrowth around the XV and
(-) Growth over entire surface of the agar X disks

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

Adapted from: Marjie S. Interno, RMT Lemar Review Hub
 BIOCHEMICAL PROPERTIES OF ENTEROBACTERIACEAE

TSI LIA I M Vi C U D L M
LACTOSE FERMENTER

Escherichia coli A/A + gas K/K + + - - - + + +

Klebsiella pneumoniae A/A + gas K/K - - + + + + + +

Enterobacter aerogenes A/A + gas K/K - - + + - + + +

Enterobacter cloacae A/A + gas K/A - - + + - + + +

LATE-LACTOSE FERMENTER

Arizona sp. A/A + gas + H2S K/K + H2S - + - + - + + +

Citrobacter freundii A/A + gas + H2S K/A - + - + - + + +

Citrobacter diversus A/A + gas K/A + + - + - + + +

NON-LACTOSE FERMENTER

Proteus vulgaris K/A + gas + H2S R/A + + - + + + - -

Proteus mirabilis K/A + gas + H2S R/A - + - + + + - -

Prodencia rettgeri K/A + gas R/A + + - + + + - +

Morganella morganii K/A + gas R/A + + - - + + - -

Salmonella typhi K/A + small HS2 K/K - + - - - + - -

Salmonella enteritidis K/A + gas + large H2S K/K - + - + - + - -

Shigella dyssenteriae K/A K/A - + - - - + - -

Shigella sonnei K/A K/A - + - - - + + +

Shigella flexnerii K/A K/A + + - - - + - +

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 Shigella boydii K/A K/A + + - - - + - +

Serratia marcescens K/A K/K + - + - + + + +

Biochemical Differentiation of Representative Enterobacteriacae

Yersinia enterocolitica
Cronobacter sakazakii
Pantoea agglomerans
Citrobacter Klebsiella Serratia Proteus Providencia

Morganella morganii
Enterobacter

Ewingella americana

Edwardsiella tarda

(was Enterobacter)
Escherichia coli

Shigella sonnei

Other Shigella

(formerly diversus)
Plesiomonas

S. enteritidis
shigelloides

K. pneumoniae
*oxidase+

S. marcescens
E. aerogenes
S. typhi
H. alvei

S. odorifera

P. mirabilis
K. oxytoxa
C. freundii

P. vulgaris
E. cloacae

P. stuartii
P.rettgeri
C. braakii

biotype 2
C. koseri
Indole + - - + - V - - + - -(v) + - + - - - -(v) - V + - + + + V
-
Methyl red + + -(v) V + + + + + + + + V - - - V V +(v) + + + + + +
(v)
Voges Proskauer - + +(v) - - - - - - - - - + + + + + +(v) + + - V - - - -
-
Simmons' citrate - + + - - - + - - + +(v) + + + + + + V + + +(v) - + + -
(v)
Hydrogen Sulfide (TSI) - - - - - - +(v) +w + + +(v) - - - - - - - - - + + - - - -
-
Urea - - - - - - - - - -(v) -(v) +(v) + + +(v) - - -(v) - + + + + -(v) +
(v)
Motility V +(v) + + - - + + + + + + - - + + + + + + + + V + +(v) -
Lysine decarboxylase +(v) - + + - - + + + - - - + + - + - - + + - - - - - -
Arginine dihydrolase -(v) - - + - V +(v) - - +(v) + + - - + - + - - - - - - - - -
Ornithine
+(v) - + + + - + - + - + + - - + + + - + - - + + - - +
decarboxylase
Phenylalinine
- - - - - - - - - - - - - - - - +(v) -(v) - - + + + + + -
deaminase
Gas from D-glucose + - + - - - + - + + + + + + + + + -(v) - - + + + - - -
Lactose + +(v) - V - - - - - +(v) + V + + + + + -(v) - + - - - - - -
Sucrose V - - - - - - - - +(v) - -(v) + + + + + +(v) + - + - - - V +

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

D-mannitol + + + - + + + + - + + + + + + + + + + + - - - + -(v) +
D-Sorbitol +(v) - - - - V + + - + + + + + + + - -(v) + + - - - - - +
KCN, growth in - - + - - - - - - + + - + + + + + -(v) + - + + + + + -
-
Gelatin (22oC) - - - - - - - - - - - - - - - - - - + + + - - - -
(v)
DNase - - - - - - - - - - - - - - - - - - + - - - - - - -

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 ACTINOBACILLLUS AGGREGATIBACTER CAPNOCYTOPHAGA SPP CARDIOBACTERIUM DYSGOMONAS KINGELLA SPP.
SPP. SPP. SPP. SPP.
COLONY Gram Stain: Exhibit Very short bacilli but Gram stain: fusiform shaped Gram Stain:
APPEARANCE: bipolar staining giving occasionally seen as bacilli with one rounded end & Plump coccobacilli with
an overall appearance filamentous forms. one tapered end and occasional squared-off ends that may
of the dots and dashes filamentous form. form chains.
of Morse code. 5%SBA: shiny, nonhemolytic
pale beige or yellowish color.
(HAZE) Gliding motility
similar to the swarming of
Proteus.
MEDIA Grow best in elevated moisture (candle jar with a -Requires CO2 and enriched Grow best in elevated Selective media
sterile gauze pad moistened with sterile water.) media. moisture (candle jar containing
-Does not grow in ambient air. with a sterile gauze pad Cefoperazone,
-selective media with moistened with sterile Vancomycin and
bacitracin, polymixin B, water). Amphotericin B
vancomycin, and trimethoprim,
or Thayer Matin and Martin
Lewes Agars.
-inhibited by SPS
OTHER INFO Actinobacillus equuli Facultative anaerobe
-Human respiratory -Nonmotile Kingella kingae
tract -β-hemolytic
Actinobacillus suis -Bone infection of childrent
-Human respiratory (3 yrs old and below)
tract *Most pathogenic specie of
Kingella
Source: Marjie Interno, Lemar Review Hub

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 A. segnis
A. Aggregatibacter (Haemophilus
actinomycetemcomitans aphrophilus segnis) Cardiobacterium hominis D. capnocytophagoides Kingella denitrificans
Gram Stain: One rounded end &
one tapered end giving the cell a
5% SBA: ▪Pinpoint colonies Gram Stain: rods or
Gram Stain: Very short teardrop appearance.
after 24 hrs; Gram stain: Pleomorphic coccobacilli
bacilli occasionally seen as -form clusters or rosettes when GS
▪sticky, adherent colonies, rods 5% SBA: (24 hrs) Pinpoint Small, nonhemolytic; frequently
filamentous. are prepared from 5% SBA.
slight greenish tinge after 48 5% SBA: Convex, colonies; (48 hrs) small, wet, pits agar; can grow on Neisseria
5% SBA: Round 5% SBA: (48-72 hrs) alpha
hrs. grayish white gray-white; some exhibit Beta gonorrhea selective agar (Thayer-
Chocolate Agar: convex, hemolytic, smooth, round, glistening
▪4-6 pointed star-liked Chocolate agar: Smooth hemolysis; Fruity Martin agar)
granular & yellow with an & opaque. Pitting may be
configuration resembling or granular at 48 hours strawberry-like odor or
opaque zone near center produced.
crossed cigars (BHI agar) bitter.
COLONY Yeast Extract Agar: Long
APPEARANCE: filaments, stick-like structure
Included in the
Aggregatibacter genus based Recovered from stool on CVA
(Haemophilus aphrophilus
on 16sRRNA sequencing. (Cefoperazone Vancomycin
and Haemophilus
-Ability to ferment mannitol Amphotericin Agar incubated
paraphrophilus reclassified
and sorbitol. at 35⁰C instead of 42⁰C
as a single specie)
-Have 6 serotypes (a-f) a, b NONMOTILE
●includes hemin-
&c are the most common. D. capnocytophagoides
dependent and hemin- Commonly mistaken as Neisseria
Requires V factor but Not (produces succinic &
independent isolates. gonorrhea. (separated by being
X factor. propionic acid) & D.
-H member of the HACEK catalase (+) and nitrate negative)
capnocytophaga (only
group
succinic acid) can be
-Does not require either X
separated by Gas-liquid
or V Factor.
Chromatography and
FOAM LOVING
distinguished by Cellular
fatty acid analysis.
OTHER INFO
Oxidase + + - - -
CATALASE: + - - +
Ferment
LACTOSE - + Not stated -
Ferment
SUCROSE - + Not stated Not stated
(+) extracted from xylene addtn of
Indole - - Erlich's reagent -

Nitrate reduction + + - -

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

Urea - - - -
Esculin
Hydrolysis - - - -

Adapted from: Marjie S. Interno, RMT Lemar Review Hub

 APPENDIX IV

BIOCHEM TEST FOR MYCOBACTERIA

TEST PURPOSE?PRINCIPLE POSITIVE NEGATIVE
Niacin(Nicotinic acid) Test Most common biochemical test for MTB -Formation of yellow liquid with addition of Liquid remains milky white/clear
identification cyanogen bromide
M. avium-intracellulare
M. tuberculosis, M.bovis
M. simiae
M. chelonae
Nitrate reduction The ability of acid-fast bacilli to reduce nitrate -Development of pink to red color No color development

M. tuberculosis M. bovis
M. kansasii
M. szulgai,
M. fortuitum
Semiquantitative Catalase Determination of catalase by the height of a >45 mm of bubbles <45 mm of bubbles
column of bubbles of oxygen NTM M. tuberculosis,
M. kansasii,
M. szulgai,
M. fortuitum
Heat stable (68ºC) Catalase Test Test for the ability of catalase enzyme to Stable at 68°C for 20 mins. Inactivated at 68°C for 20 mins
remain active after heating NTM M. tuberculosis, M. bovis, M. gastri,
M. haemophilum, M. marinum
Tween 80 hydrolysis Test for the ability of Mycobacteria to produce Positive in 5 days (RED) Negative (AMBER)
lipase Pink to red No color change
M. kansasii, M. gordonae
Positive in 10 days M. bovis
Pink to red M. tuberculosis M. avium

Tellurite Reduction Test for the ability of Mycobacteria to reduce Smooth, fine, black precipitate (smokelike
tellurite in 3 to 4 days to black metallic action) Gray clumps (no smokelike action)
tellurium Rapid growers
M. avium complex M. tuberculosis
M. kansasii
Arylsulfatase Test for the ability of Mycobacteria to produce Positive in 3 days Negative
arylsulfatase Pink to red M. fortuitum-chelonae Colorless, No color change
phenolphthalein disulfate  phenolphthalein Positive in 14 days M. tuberculosis)
(red with NaCO3) Pink to red M. marinum and M. szulgai

Adapted from: Marjie S. Interno, RMT Lemar Review Hub