Enterobacterales

salmonella can cause:

septicemia and diarrhea

shigella can cause:

dysentery, diarrhea

E. coli can cause:

shiga toxin producing

yersinia pestis can cause:

plague

general stain characteristics

gram negative bacilli (rods)

temp for yersinia

at cold enrichment and room temp

colony morphology

All mostly large grey on blood agar, need selective/differential media, and biochemical tests

gram stain and colony of klebsiella

Mucoid, lactose fermenter, capsules around on gram stain.

gram staining on yersinia

Bipolar staining, (dark on both ends) Wayson stain

E coli hemoplysis:

large grey beta hemolytic

Enterobacter and klebsiella colony morphology

large mucoid on blood and MAC

Serratia marcescens colony morphology

pink/red on blood sometimes on MAC

Enterobacter (Pantoea) agglomerans colony morphology

large yellow on blood

proteus colony morphology

swarming on blood and CAP, concentric rings on agar

MacConkey agar MAC

Differential for lactose, selective with crystal violet and bile salts.
---pos is dark pink, neg is transparent

Eosin, Methylene Blue agar EMB

Differential for sucrose and lactose, selective with Eosin Y and Methylene Blue
—pos black/purple color, E. coli distinctive green metallic sheen, neg is transparent

Hektoen-Enteric Agar HE

Differential for lactose, sucrose, salicin, selective with bile salts, H2S indicator is sodium thiosulfate

—pos salmon colored, negative green to blue,
—H2S pos is black color

Salmonella and shigella agar SS

Differential for lactose, selective with brilliant green and bile salts, H2S indicator sodium thiosulfate

-Pos red, neg transparent
-H2S pos is black center
-used in all stool samples to differentiate the two bacteria

Xylose-Lysine Deoxycholate agar XLD

Differential for Lactose, sucrose, and xylose, selective with bile salts, H2S indicator sodium thiosulfate,

-Pos is yellow color, neg is transparent on red medium
-H2S pos is black
-red colony its non lactose, H2S neg

Strong lactose fermenters

Escherichia
Enterobacter
Klebsiella

Delayed lactose fermenters

Serratia (ONPG pos)
Citrobacter

Klebsiella

Lactose fermenter, H2S neg, A/A

Shigella

Non lactose, H2S neg, K/A

Salmonella

Non lactose, H2S pos, K/A with H2S+

H2S producers

Edwarsiella
Proteus
Citrobacter
Salmonella

Enrichment broths

Selenite and GN broth
-primarily for salmonella and shigella
-inhibit gram positive

Key characteristics

all oxidase neg except for some like pleisomonas
reduce nitrate to nitrite
ferment glucose
all motile except klebsiella, shigella and yersinia at 37 C ( it is at 22 C)

Nitrate reduction test

test if organism reduces nitrate NO# to nitrate NO2 or even all the way to nitrogen gas N2
—Inoculate a nitrate broth with organism, incubate in fresh incubator for 24 hours,
—then add sulfanic acid (nitrate reagent A/1) then alpha naphthylamine (nitrate reagent B/2)
—Negative is clear/yellow pos is red color

To tell if reduced all the way to nitrogen gas (remains clear) add zinc powder which reduces nitrate to nitrite
—pos is clear then yes it was all the way to gas, neg is red

TSI triple sugar iron slant

Fermentation of carbohydrates (sugars) into acid end products will result in a yellow color on the slant and/or butt
—Reported “A” for acid or “K” for alkaline
—Stab agar and streak it

Will turn yellow for glucose fermentation, the peptone utilization will yield alkaline products, proteins=alkaline, reversal to pink color
—Didn’t utilize sucrose or lactose
—Any yellow or black on the butt or bottom part of the tube means glucose fermenter
—The top part yellow means used sucrose and or lactose
—if top is pink= it used peptones or proteins
—Black is H2S pos
—if the whole thing is red/pink it doesn’t belong to that family

Motility test

Semi solid media
—stab media with needle, some have an indicator color, incubate for 24 hours
—looking if organism has moved away from stab site
—cloudiness or movement is pos

Enterobacteriaceae best identified by?

biochemical tests
—API system

Methyl red

mixed acid pathway
—strong acid
—pH to 4.5 or lower (MR+)

Voges Proskauer

Butylene glycol
—weak acids especially Acetoin reduced further into acetyl methyl carbinol
—wont reduce to less and 4.5 ph to turn it red
—VP+ after reagents 1 and 2
—tribe 5 including Klebsiella, Enterobacter, Serratia, Hafnia, and Pantoea are VP+

LIA Lysine iron agar slant

If it removes carboxyl (anaerobic process, bottom of slant) or amine group (aerobic process, top of slant)
—Bromocresol purple pH indicator color, sodium thiosulfate, and ferric ammonium citrate (H2S)
—Dark purple color at pH 6.8 or higher and yellow at pH 5.2 or lower

-Positive for decarboxylation = purple at the butt, Negative deamination = purple/port wine on slant
-Negative for decarboxylation = yellow at the butt, Positive for deamination = port wine color on slant

Byproducts

Proteus, Providencia, and Morganella positive for deamination, port wine color on slant

ONPG

To use lactose need Beta-galactoside permease (lactose permease) and Beta-galactosidase
—slow lactose fermenters don’t have Beta-galactoside permease
—detect delayed lactose fermenters, = yellow color, clear is non lactose fermenter

Spot Indole test

Indole is a degradation product, looks like the oxidase test on filter paper or circle.
—Saturate filter paper with reagent then with wooden stick add colonies to paper.
—Positive blue green color (ex": Proteus vulgaris, Klebsiella oxytoca, E. coli)
—Negative is no color change (ex: Proteus mirabilis, Klebsiella pneumonia)

Indole with Kovac’s reagent

Indole test in tube
—Positive is production of pink/red color after adding reagent
—Negative is production of yellow color after adding reagent

SImmons citrate medium slant

Ability of organism to use citrate as sole carbon source
--pH indicator is bromothymol blue
--neutral pH of this is green color in tube

Streak the slant part only and leave cap loose, incubate for 24 hours and read
—positive is blue (alkaline) color
—negative is it remains green color

Urea agar slant

Production of ammonium by splitting of urea by urease, increasing pH to alkaline
—color change from yellow to pink
—Pos is pink color neg remains yellow

Some urea positive organisms:
—Proteus, morganella, and Providencia rettgeri

Serratia is also:

Gelatinase Positive and DNase positive