II: Gram Negative Bacilli Flashcards
Normal Intestinal Flora
UTI associated
Often resistant to antimicrobial therapy
Providencia
Providencia spp. urease positive
P. rettgeri
Providencia spp. urease negative/variable
P. stuartii
Proteus spp that is Indole (-)
P. mirabilis
Proteus spp. Indole (+)
P. vulgaris
- Gram-negative Bacilli
- Non-Lactose Fermenters
- Isolated from Cold blooded and warm blooded animals
Edwardsiella
Edwardsiella spp. occurs in most human species
E. tarda
E. tarda has same IMVIC result to
E.coli (++–)
To differentiate E. tarda from E.coli
E. tarda has
H2S:
NLF / RLF:
H2S: +
NLF / RLF: NLF
To differentiate E. tarda from E.coli
E.coli has
H2S:
NLF / RLF:
H2S: -
NLF / RLF: RLF
Late-Lactose Fermenter
Gram-Negative Bacilli
Causes Black Death/Bubonic Plaque
Yersinia pestis
Vector of Yersinia pestis
Xenopsylla cheopsis (Rat flea)
Yersinia pestis in broth agar exhibits clump of cells adhere to one side of tube known as
Stalactite pattern
Y. pestis preferred temp for growth
25-30 deg Cel
Y. pestis exhibit ____ appearance after 48 hrs in BAP
Cauliflower
Y. pestis inclusion bodies forms
safety pin appearance
Y. pestis inclusion bodies are stained using
Wayson stain
Gram-negative Bacilli
Late-Lactose Fermenters
Causes ENTEROCOLITIS (fever, abdominal pain, diarrhea); Appendicitis in children
Y. enterocolitica
Associated with Arthitis, Erythrema Nodosum
Gram-negative Bacilli
Late Lactose Fermenters
Y. enterocolitica
Y.enterocolitica is motile @ _______ deg and non motile @ ___________
Motile - Room Temp
Non-Motile - 37 deg Cel
Y. enteroclotica is culture in CIN agar; CIN means
Cefsulodin Irgasan Novobiocin Agar
Y. enterocolitica in CIN agar appears
Bull’s Eye colonies
H2S (+)
SPACEd
Salmonella
Proteus
Arizonae
Citrobacter freundii
Edwardsiella
Microorganism with Bipolar Inclusion Bodies
Bipolar si BAY
B. pseydomallei
A. actinomycetemcomitans
Yersinia
LISTAHAN NG MGA BULL-BULLL
Bull-eyes colonies on CIN
Y. Enterocolitica
Aeromonas
LISTAHAN NG MGA BULL-BULLL
Fish eye colonies on EMB
Enterobacter
LISTAHAN NG MGA BULL-BULLL
Bull’s eye amoeba
E. histolytica
LISTAHAN NG MGA BULL-BULLL
Bull’s Eye Rash
Borellia burdorgferi
LISTAHAN NG MGA BULL-BULLL
Bull’s Neck Infection
C. diptheriae
LISTAHAN NG MGA BULL-BULLL
Buffalo Hump
Cushing Syndrome
Agar that uses Ferric Ammonium Citrate for indication of H2S
SSA, XLD, HEA, SIM , LIA
Considered as INITIAL STEP for the identification of Enterobacteriaceae
Triple Sugar Iron Agar (TSIagar)
TSI Agar Component:
CHO (3)
1% Lactose (Slant)
1% Sucrose (Slant)
0.1% Glucose (Butt)
TSI Agar component:
pH indicator
Phenol Red
TSI Agar component:
Sulfur Source:
Sodium thiosulfate
TSI Agar Component:
H2S indicator:
Ferrous sulfate
H2S + Ferric Ions =
Ferrous sulfide (black precipitate)
TSI Reactions spp.
A/AG H2S (-)
EKE
Escherichia
Klebsiella
Enterobacter
TSI Reactions spp.
K/AG H2S (+)
SPACEd
Salmonella
Proteus
Arizonae
Citrobacter freundii
Edwardsiella tarda
TSI Reactions spp.
K/A H2S (-)
Shigella
Providencia
Morganella
TSI Reactions spp.
K/K H2S (-)
Pseudomonas
Burkholderia
Test primarily for ENTERIC pathogen
Lysine Iron Agar (LIA)
test used to detect if organism can deaminate Lysine (red) or Decarboxylase Lysine (purple)
Lysine Iron Agar
Lysine Iron Agar Composition:
Amount of protein must be in:
a. Small
b. Large
Small amount of protein
Lysine Iron Agar Composition:
CHO
Glucose
Lysine Iron Agar Composition:
pH indicator
Bromcresol purple
Lysine Iron Agar Composition:
H2S indicator
Ferric Ammonium Citrate
LIA Reaction spp.
K/K H2S (+)
Salmonella
LIA Reaction spp.
K/A H2S (-)
Shigella
LIA Reaction spp.
R/A H2S (-)
PPM
Proteus
Providencia
Morganella
*Proteus
- H2S (+) but does deaminate lysine = alkaline = inc. ammonia = Alk cannot produce H2S since; H2S can only be produced in ACIDIC ENVIRONMENT
Selective Differential Media for Gram-Negative Enteric Bacilli (5)
- Eosin Methylene Blue
- Macconkey Agar
- Hektoen Enteric Agar (HEA)
- Salmonella Shigella Agar
- XLD Agar
Eosin-Methylene Blue Agar Composition:
CHO:
Inhibitor & Indicator:
CHO: Lactose and Sucrose
Inhibitor & Indicator: Eosin Methylene Blue
Eosin-Methylene Blue Result:
Lactose Fermenters:
Non Lactose Fermenters:
LF: Pink to Purple Colonies
NLF: Colorless
Eosin-Methylene Blue Result:
Pink to Purple Colonies with Green Metallic Sheen
E.coli
Eosin-Methylene Blue Result:
Pink mucoid colonies
Klebsiella
Eosin-Methylene Blue Result:
Pink colonies with Black Center “Fish Eye Colonies”
Enterobacter
Macconkey Agar Composition:
Inhibitor
Crystal Violet
Bile Salt
Macconkey Agar Composition:
pH indicator
Neutral Red
Macconkey Agar Composition:
CHO
Lactose
Macconkey Agar Result:
NLF:
LF:
NLF: Colorless colonie
LF: Pink
Neutral Red
Acidic < 6.8
Alkaline 6.8 -8.0
Acidic < 6.8 = Pink-red
Alkaline 6.8 -8.0 = Brown
Hektoen Enteric Agar Composition:
CHO
Lactose, Sucrose, Salicilin
Hektoen Enteric Agar Composition:
Inhibitor
Bile Salts
Hektoen Enteric Agar Composition:
pH indicator
Bromthymol Blue
Hektoen Enteric Agar Composition:
H2S indicator
Ferric Ammonium Citrate
Hektoen Enteric Agar Result:
LF (H2S +)
NLF (H2S -)
LF (H2S +) - Yellow-Orange with Black Center
NLF (H2S -) - Green-Blue w/o Black Center
Salmonella-Shigella Agar Composition:
CHO
pH indicator
H2S indicator
CHO: Lactose
pH indicator: Neutral Red
H2S indicator: Ferric Ammonium Citrate
Salmonella-Shigella Agar Result:
Colorless Black Center
H2S +
NLF
Salmonella
Salmonella-Shigella Agar Result:
Colorless w/o Black Center
H2S -
NLF
Shigella
Salmonella will have Mauve-colored colonies; Enterobacter will have blue-white colonies
CHROMagar Salmonella
NLF spp. produces lysine decarboxylase showing Red Colonies w/ Black Center in this agar
XLD Agar
XLD agar composition:
CHO
Sulfur Source
pH indicator
H2S indicator
Inhibitor
CHO - Lactose, Sucrose, Xylose
Sulfur Source - Sodium thiosulfate
pH indicator - Phenol Red
H2S indicator - Ferric Ammonium Citratre
Inhibitor - Sodium desoxycholate
XLD agar Result:
Red w/ Black Center or Colorless w/ Black Center
Salmonella
IMVIC
Indole
Methyl Red
Voges Proskauer
Citrate
Indole detects for
Tryptophanase
Indole reagent
Kovacs/Ehrlich Reagent (PDAB)
Kovac’s Reagent
Positive Control:
Negative Control:
Positive Control: E. coli
Negative Control: K. pneumoniae
Ehrlich Reagent
Positive Control:
Negative Control:
Positive Control: H. influenzae
Negative Control: H. parainfluenzae
Indole Test
Medium uses
Sulfur Indole Motility (SIM) / Tryptone Broth
Indole Test Result:
Positive:
Negative:
Positive: Red Ring
Negative: w/o color change
Indole Test Positive spp
May PEKPEC Po?
Morganella
Proteus vulgaris
E. tarda
K. oxytoca
Providencia
E. coli
Citrobacter koseri
Plesiomonas
Test for Mixed Acid Fermentation Pathway
Methyl Red
Methyl Red - Mixed Acid Fermentation = LAFS
Lactic
Acetic
Formic
Succinyl acid
Methyl Red QC:
Positive
Negative
Positive: E.coli
Negative: Enterobacter aerogenes
Methyl Red Result
Positive:
Negative:
Positive: Red
Negative: Yellow
Test used to detect Acetoin (ButyleneGlycol Pathway)
Voges-Proskauer Test
Voges Proskauer Reagent
a-naphtol (4drops)
40% KOH (2 drops)
Voges Proskauer QC:
Positive
Negative
Positive: Enterobacter aerogenes
Negative: E.coli
Voges Proskauer Result:
Positive
Negative
Positive: Red
Negative: Yellow
Voges-Proskauer Positive Spp.
KEESH
Klebsiella
Ewingella
Enterobacter
Serratia
Hafnia
Citrate Utilization Test Composition:
Medium:
pH indicator:
Simmon’s Citrate
Bromthymol blue
Citrate Utilization Test Result:
Positive:
Negative:
Positive: Growth/Blue
Negative: No growth/ green
Citrate Positive Test:
Pero Sa CHEEKS
Pero Sa CHEEKS
Providencia
Serratia
Citrobacter
Erwingella
Enterobacter
Klebsiella
Salmonella enteritidis
Malonate utilization has the same result w/ (3)
Acetamide
Acetate
Citrate
Malonate Utilization Result
Positive
Negative
Positive: blue
Negative: green or yellow
Urease Test Composition
Medium:
pH indicator:
Positive will produce:
Medium: Stuart’s Urea broth/ Christensen’s Urea Agar
pH indicator: Phenol Red
Positive will produce: Ammonia
Urease Test
Rapid Urease will produce:
TPUNCH
Rapid Urease will produce: Pink to Red colonies w/in 2-4 hrs
TPUNCH
T. mentagrophytes
PPM
Ureaplasma
Nocardia
C. neoformans
H. pylori
Urease Test
Slow Urease will produce:
CKEYS
Slow Urease will produce: Orange after 4hrs
CKEYS
Citrobacter
Klebsiella
Enterobacter
Yersinia enterocolitica
Serratia
ONPG test is a rapid test to detect
B-galactosidase
ONPG substrate
O-nitrophenyl-beta-D-galactopyranoside
ONPG is hydrolyze to form O-nitrophenol forming
Yellow colonies (LF)
ONPG Result:
Positive:
Negative:
Positive: Yellow
Negative: Colorless
Test to detect B-gluconidase
MUG test
All E.coli are MUG TEST POSITIVE EXCEPT
E. coli 0157: H7
Test that detects the ability of an organims to remove carboxyl group from amino acid
Decarboxylase Test
Decarboxylase Test Composition:
Medium:
Indicator:
CHO:
Amino acid:
Medium: Decarboxylase Basal Medium (Moller’s Medium)
Indicator: Bromcresol Purple
CHO: Glucose
Amino acid: Lysine, Ornithin, Arginine
Decarboxylase Test
Positive Result
Purple-yellow-purple
Test to detect organism’s ability to remove amino group from an amino acid
Phenylalanine Deaminase
Phenylalanine Deaminase composition:
Phenylalanine Agar
Overnight Culture
10% FECI
Phenylalanine Agar
Overnight Culture
10% FECI3
Phenylalanine Deaminase Result
Positive
Green Slant and fluid
Routinely Performed Test for Enterobacteriaceae (6)
- IMVIC, MOtility
- Growth in KIA/TSI - LF, H2S
- Urease, ONPG
- LDC, ODC, ADC
- PPA/PDA
- Mannitol Fermentation