micro lab test pt.1

enteric bacteria

“GUT bacteria → they EAT sugar”

E. coli, Enterobacter, Klebsiella, Salmonella

• Ferment glucose ✅

• Sometimes ferments lactose ✅

• Oxidase negative ❌

• convert nitrate to nitrite

non enteric bacteria

• Alcaligenes, Pseudomonas

• Do NOT ferment lactose ❌

• Often don't ferment glucose (or do weird metabolism) ❌

• Oxidase positive ✅

• nitrate to nitrite to ammonia or gas

🧠 EASY WAY TO STUDY- ENTERIC

• Enterics → oxidase NEGATIVE ❌

• Enterics = gut = ferment sugar = oxidase NEGATIVE

🧠 EASY WAY TO STUDY- NON ENTERIC

• Non-enterics = not gut = don't ferment lactose = oxidase POSITIVE

• Non-enterics → oxidase POSITIVE ✅

• ✔ Convert nitrate → nitrite → ammonia or gas

MacConkey agar (MAC) is used for what?

most important media for all clinical samples except stool samples

MacConkey agar (MAC) selective for?

Gram negative bacteria
selective agents
bile salts and crystal violet
inhibit Gram positive bacteria

✅ Question 1: Who is allowed to grow? MAC

• Bile salts + crystal violet = block Gram-positive bacteria ❌

• So only Gram-negative bacteria grow ✅

👉 So if something grows → it's Gram-negative

🍬 Question 2: What color are the colonies? PINK

MAC

This tells you about lactose fermentation

🩷 PINK / RED colonies

• Bacteria ferments lactose → makes acid

• Acid turns neutral red → pink

👉 Think: “Ferments sugar = turns pink”

🍬 Question 2: What color are the colonies? 🤍 WHITE / COLORLESS colonies

MAC

• Bacteria does NOT ferment lactose

• No acid → stays colorless

👉 Think: “No sugar fermentation = no color”

MacConkey agar (MAC)- differential?

lactose

MAC GRAPH

Result	Meaning
Growth	Gram-negative bacteria
Pink colonies	Lactose fermenter
White colonies	Non-lactose fermenter

🟢 Lactose fermenters (PINK)

• bacteria

• E. coli

• Enterobacter

• Klebsiella

👉 “Gut bacteria that eat sugar → pink”

🔵 Non-lactose fermenters (WHITE)

• MAC

salmonella

👉 “Don't eat lactose → stay clear”

Enterococcus- MAC does it grow?

Gram positive
does not grow on MAC

only gram negative bacteria grow

MacConkey agar-Selective agents

bile salts and crystal violet
inhibit Gram positives

Differential agent- MacConkey Agar

lactose

distinguishes Gram negatives by lactose fermentation

MAC ph indicator?

neutral red

Eosin Methylene Blue emb- used for?

water samples

Eosin Methylene Blue emb is what?

selective and differential agar

✅ Step 1: Who grows?

• EMB

• Eosin + methylene blue dyes + bile salts-

• ❌ Kill Gram-positive bacteria

• ✅ Allow Gram-negative bacteria

👉 If it grows → it's Gram-negative

Eosin Methylene Blue emb- selective agents?

eosin dye, methylene blue dye, bile salts

🟢 GREEN SHINY (metallic green) EMB

👉 ONLY E. coli

• Strong lactose fermentation

• Produces LOTS of acid

• Reacts with dyes → green metallic shine

🔥Think:
“E. coli = EXTREME fermenter = GREEN shine”

🩷 PINK / RED

EMB

👉 Other lactose fermenters/ other gram negatives

EXAMPLE: ENTEROBACTER OR KLEBSIELLA

• Ferment lactose

• Make acid (but not as much as E. coli )

👉Think:
“Normal fermenters = pink”

🤍 COLORLESS / CLEAR

EMB

👉 Non-lactose ferments

EXAMPLE: SALMONELLA

• No acid

• No color change

👉 Think:
“No sugar = no color”

Eosin Methylene Blue emb- differential agent?

lactose
distinguishes Gram negatives by lactose fermentation

EMB- no growth bacteria?

gram positive

staphylococcus aureus

Eosin Methylene Blue emb- does it have a ph indicator?

NO

Eosin Methylene Blue emb- E. coli why does it turn green?

due to high acidity, motility, and metachromatic properties of dyes

too much acid

Eosin Methylene Blue emb- Salmonella

whitish
does not ferment lactose to acid

Hektoen enteric agar (HE) is used for what?

used to evaluate stool samples

Hektoen enteric agar (HE) is what type of agar?

selective and differential agar

Hektoen enteric agar (HE) selective for who?

Gram negative enterics particularly Shigella and Salmonella

👉 Mainly looking for:

• Salmonella

• Shigella

Hektoen enteric agar (HE) distuinguishes who?

distinguishes Shigella from Salmonella

✅ 1. Who grows?

• HE

• Bile salts → kill Gram-positive ❌

• Only Gram-negative enterics grow ✅

Hektoen enteric agar (HE)- Selective agent

bile salts- inhibits Gram positives

Hektoen enteric agar (HE) - differential agent

lactose,

sucrose and salicin (sugar) fermentation

distinguishes Gram negatives by lactose fermentation

🍬 2. Do they ferment sugars?

HE

Sugars on HE:

• lactose

• sucrose

• salicin

🟡 ORANGE / YELLOW

👉 Ferments sugars → makes acid

Think:
“Normal gut bacteria = eat sugar = turn yellow”

🔵 GREEN / BLUE

👉 Does NOT ferment sugars

Think:
“Pathogens (Salmonella/Shigella) = don't ferment = stay green”

Hektoen enteric agar (HE) - pH indicator

bromothymol blue

💨 3. Do they produce H₂S (hydrogen sulfide)?

• HE

• Peptone iron detects H₂S

• H₂S + iron → BLACK color

---

⚫ BLACK (in colony or agar)

👉 H₂S positive

• Classic for Salmonella

Hektoen enteric agar (HE) -orange/ yellow colonies and agar

ferments lactose to acid

Hektoen enteric agar (HE)-greenish/ bluish colonies and agar

color of agar
does not ferment lactose

Hektoen enteric agar (HE) results- E. coli

orange on HE
ferments lactose to acid

HE GRAPH

Color	Meaning
Yellow/orange	Ferments sugars (normal bacteria)
Green/blue	No fermentation (possible pathogen)
Black	H₂S production (→ Salmonella)

Hektoen enteric agar (HE) results- Salmonella

greenish blue/ black on HE
does not ferment lactose to acid
-

• black- produces H2S

Hektoen enteric agar (HE) results- Proteus vulgaris, Citrobacter

yellow/ black on HE
ferments lactose to acid
black- produces H2S

what’s this?

Hektoen Enteric Agar- HE

what’s this?

Hektoen Enteric Agar- HE

oxidase test is what?

tests for cytochrome oxidase enzyme

checks if a bacterium has the enzyme “cytochrome oxidase”

👉 That enzyme is part of how some bacteria use oxygen to make energy.

oxidase test shows?

Indicates if bacterium is aerobic non enteric

cytochrome

oxidase test has what transport?

has electron transport chain similar to mitochondria ETC

oxidase reagent donates … to cytochrome oxidase

electrons

oxidase test- Enteric bacteria

cytochrome oxidase negative

oxidase test- Reagent

oxidase reagent

🔥 WHY THIS TEST MATTERS

OXIDASE test

Group	Oxidase
Enterics (gut bacteria)	❌ Negative
Non-enterics	✅ Positive

• Oxidase test = enzyme test

• Purple = positive

• No color = negative

• Enterics = negative

• Non-enterics = positive

oxidase test- Positive color?

blue/ black color on swab after 10 seconds
Alcaligenes - non enteric bacterium

oxidase test- Negative

not blue/ black on swab after 10 seconds
E. coli- enteric bacterium

🟣 OXIDASE POSITIVE

👉 Has cytochrome oxidase

👉 “P = Positive = Pseudomonas”

Examples:

• Pseudomonas

• Alcaligenes

👉 Think:
“Non-enterics = oxidase POSITIVE”

⚪ OXIDASE NEGATIVE

👉 Does NOT have cytochrome oxidase

👉 Enterics = NEGATIVE

Example:

• E. coli

👉 Think:
“Enterics = oxidase NEGATIVE”