Differential Media:
Overview of Some Common Enteric Plating Media

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Page 1

  • On this page and continuing on "Page 2": MacConkey ("regular" and the Bact. 102 modification), EMB, Hektoen Enteric, Brilliant Green and XLD Agars.
  • Supplementary Information:
    • Our old explanatory material on MacConkey, Hektoen Enteric, Brilliant Green and XLD Agars is archived on "Page 3."
    • Our newly-revised and simplified Bacteriology 102 demonstration on enteric plating media which utilizes only Modified MacConkey Agar and XLD Agar is on "Page 4."
  • Additional Notes:
    • On this site: "Screening media" that are inoculated from enteric plating media are outlined and illustrated here.
    • Some of these media (along with Bismuth Sulfite Agar) are included in a lecture outline which begins here and includes a list of common pH indicators and selective agents.
    • Media formulated for isolation and characterization of Edwardsiella tarda are analyzed and discussed here.

xAs an introduction or review: Many of our pH-based differential media have one or more of the features shown on the following table which should be helpful in analyzing the reactions of certain selective-differential media used in the isolation of enterics. The introductory page of the Differential Media Site addresses the concept of "programming" differential media to detect certain physiological types, and a good practical overview of the general concept can be found here.

Aerobic or Anaerobic ReactionSubstrateMicrobial ActivityApparent Reaction (noted with appropriate pH or H2S indicator in medium)Some Examples
aerobicvarious amino acids in peptones, yeast extract, etc.deaminationalkalineGlucose O/F and Fermentation Media, KIA, TSI, and many other differential media
anaerobicone or more specific sugarsin relatively large amountfermentationrelatively large amount of acidGlucose O/F and Fermentation Media, MacConkey Agar, Brilliant Green Agar, XLD Agar, KIA, TSI
in relatively small amountfermentationrelatively small amount of acidXLD Agar, KIA, TSI
specific amino acid in relatively large amountdecarboxylationalkalineXLD Agar, MIO, Lysine Broth
thiosulfatereduction with formation of H2S black color
(not a pH-related reaction)
Modified MacConkey Agar (a Bact. 102 exclusive), XLD Agar, KIA, TSI

The following two tables summarize six selective-differential plating media used in the isolation of enterics. When formulating such media, one takes advantage of the enterics' gram-negativity, and one or more agents that inhibit gram-positive bacteria are added. In the isolation of the enteric pathogens Salmonella, Shigella and Edwardsiella from food or clinical samples, an important consideration is the fact that they usually do not ferment certain sugars, especially lactose and sucrose. Therefore, one tends to pick colonies showing a net alkaline reaction on media containing the appropriate sugar(s), ignoring the acidic colonies which coliforms and various other enterics tend to produce.

In Bacteriology 102 for the enteric isolation experiment, we make use of MacConkey Agar which is modified by the addition of the H2S indicator system possessed by XLD Agar – i.e., sodium thiosulfate and ferric ammonium citrate. As far as we know, no such medium exists out there in the "real world," but we use it to illustrate some features of multipurpose selective-differential media one might encounter in a clinical laboratory.

Dr. Scott Cayley's explanation of the selective aspect of MacConkey Agar is shown here.

A study of the special differential features of XLD Agar is especially rewarding. This medium incorporates lysine which may be decarboxylated, resulting in an alkaline reaction that may or may not overneutralize any acid formed from fermentation of the various sugars. As shown below and on the following pages, incorporation of lysine in XLD Agar "tips the balance" to distinguish strains of Salmonella (which characteristically decarboxylate lysine and thus produce a net alkaline reaction on XLD Agar) from the "Salmonella-like strains" of Citrobacter. (Many other strains of Citrobacter rapidly ferment lactose with the production of acid and gas and would be termed "coliform strains" of Citrobacter, remembering the definition of coliform.) Without the lysine present, both organisms would produce a net acid reaction (from the fermentation of xylose), and the medium would be like other isolation media which render these two organisms indistinguishable from each other – making the isolation process for Salmonella a bit more tedious.

usual formulation
special Bact. 102 modification
Levine's formulation
selective agent(s)bile salts,
crystal violet,
neutral red
bile salts,
crystal violet,
neutral red
eosin Y,
methylene blue
source of amino acids which may be deaminated (alkaline rx.)peptone,
proteose peptone
proteose peptone
amino acid added for detection of decarboxylation (alkaline rx.)nonenonenone
fermentable sugar(s) (acid rx.)lactose (1%)lactose (1%)lactose (1%)
pH indicatorneutral red:
net acid = red,
net alkaline = whitish/light
neutral red:
net acid = red,
net alkaline = whitish/light
eosin Y and methylene blue:
net acid = dark,
net alkaline = light
source from which H2S may be producednoneNa thiosulfatenone
indicator of H2S productionnoneferric ammonium citratenone

selective agent(s)bile saltsbrilliant greenNa desoxycholate
source of amino acids which may be deaminated (alkaline rx.)proteose peptone,
yeast extract
proteose peptone,
yeast extract
yeast extract
amino acid added for detection of decarboxylation (alkaline rx.)nonenonelysine
fermentable sugar(s) (acid rx.)lactose (1.2%),
sucrose (1.2%),
salicin (0.2%)
lactose (1%),
sucrose (1%)
lactose (0.75%),
sucrose (0.75%),
xylose (0.375%)
pH indicatorbrom thymol blue + acid fuchsin:
net acid = yellow-orange,
net alkaline = blue-green
phenol red:
net acid = yellow,
net alkaline = red
phenol red:
net acid = yellow,
net alkaline = red
source from which H2S may be producedNa thiosulfatenoneNa thiosulfate
indicator of H2S productionferric ammonium citratenoneferric ammonium citrate

Characteristic reactions observed on these media are shown and explained on the next page.

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the Differential Media site.

Page last modified on 5/26/18 at 7:00 PM, CDT.
John Lindquist:  new homepage, complete site outline.
Department of Bacteriology, U.W.-Madison