MOD7 Enterobacterales Media

enterobacterales

order of facultative anaerobic GNB

aka coliform bacteria

two groups:

• spp that colonize human GI tracts/associated w human infcs → E.coli

• genera generally recognized as environmental inhabitants/colonize other animals — capable of colonizing humans, but rare → Shigella spp

Enterobacterales common characteristics

GNB

oxidase negative

reduces nitrate to nitrite

Enterobacterales appearance on BAP

large colonies → fast growers

convex

entire or irregular edges

grey-white

may be mucoid

may be beta or gamma → not important determinant of ID

Enterobacterales microscopic appearance

ends of bacilli stain darker → biopolar staining

ends appear roughed vs rounded

no spores (GNB cant) and not coccobacilli

nutrient agar

basic media ingredient to sustain bacterial growth

enriched agar

nutrient agar + special ingredients → blood, serum

selective agar

ingredients added to inhibit growth of some bacteria and allow growth of target organisms

differential agar

for enteric organisms → carbs + pH indicator

colour changes to colonies based on carb fermentation

selective and differential agar

media showing both abilities → ex MacConkey

enrichment/selective enrichment media

liquid media that inhibit normal flora and increase growth of pathogens

goal of stool/enteric culture media

facilitate detection → differentiate btwn pathogens and nonpathogens

reduce growth of other nonpathogenic organisms

reporting enteric pathogens

dont consider amount of growth → presence of pathogen indicates disease

additives in enteric culture media

bile salt → inhibit GPO

carbohydrate and pH indicator → fermentation

ferric ammonium citrate or sodium thiosulfate → presence of hydrogen sulfide H2S

purpose of MAC

isolate GNB + suppress GPO (if CV added)

differentiate isolates as lactose fermenters LF or nonlactose fermenters NLF

additives in MAC

carb: lactose

indicator: neutral red → A: red/pink, B: yellow/colourless

why differentiate LF and NLF

help with identification

most stool pathogens will never present as LF

what type of media is MAC

selective for GNB

differential for LF vs NLF

media types for Salmonella and Shigella

xylose lysine deoxycholate XLD

hektoen HEK

salmonella shigella SS

salmonella chromogenic medium

additives in xylose lysine deoxycholate XLD

sodium deoxycholate → GPO inhibition (also in bile solubility)

carbs: xylose, lactose, sucrose

indicator: phenol red → A: yellow // B: red (flipped)

sodium thiosulfate/ferric ammonium citrate → H2S detection

lysine → decarboxylation by isolates = alkaline

Salmonella spp on XLD

NLF; ferments xylose → small amount of acid produced

lysine in medium = decarboxylated by organism → alkaline

= acid production from xylose fermentation → neutralized

spp producing H2S → black centers

spp not producing H2S → no black centers

Shigella spp on XLD

no carb fermentation or H2S production → red colonies

nonpathogens on XLD

ferment carbs → yellow colonies

with or without black centers

(if see yellow, can ignore; non path)

additives in hektoen agar HEK

bile salts

carbs: lactose, sucrose, salicin

indicators: bromothymol blue → A: yellow, B: blue

acid fushin: adds red colour to carb fermentation

combo of BTB and acid fuchsin → orange colour

ferric ammonium citrate/sodium thiosulfate → H2S

pathogens on HEK

green/blue-green colonies

with or without black centers

non pathogens on HEK

yellow/salmon colonies

with or without black centers

additives in Salmonella Shigella agar SS

bile salts/brilliant green/sodium citrate → selectivity

carb: lactose

indicator: neutral red → A: red, B: yellow

ferric ammonium citrate/sodium thiosulfate → H2S

potential pathogens on SS

colourless colonies (NLF)

with or without black

normal flora on SS

pink colonies (LF)

with or without black

spp appearance on Salmonella chrom agar

Salmonella spp → mauve

nonpathogens → blue

types of enrichment broths

selenite F broth

tetrathionate broth

Gram negative broth GN

Thiosulfate Citrate Bile salts Sucrose (TCBS) agar use

Vibrio spp

used in conjunction w BAP for hemolysis

Vibrio spp cellular morph

comma shaped GNB

appearance on TCBS

sucrose fermenters → yellow

non sucrose fermenters → clear

cefsulodin-irgasan-novobiocin CIN use

Yersinia spp

abx combo suppresses GNO/GPO → proliferation of enteric Yersinia spp

Yersinia spp appearance

bullseye colonies

sorbitol macconkey SMAC use

E.coli O157

utilizes slow/inability to ferment sorbitol

nonpath strains of E.coli on SMAC

ferment sorbitol

red colonies

EC O157 on SMAC

non-sorbitol fermenting NSF / clear colonies

further investigation required

EC O157 on chromogenic agar

turquoise colonies

additives in Campylobacter spp selective media

abx to inhibit other organisms present

Campylobacter spp

most commonly isolate stool pathogen

GS: gullwing GNB

Campylobacter spp selective media growth conditions

incubated in microaerophilic 5% O2 environment

42deg

for further suppression of organisms