Stains and Cultures Simplified: High-Yield Microbiology

Scientists and Stains

Negative Stains (b/w)

• Staining the background to highlight the organisms
• Cryptococcus causes cryptococcal meningitis Sample : CSF
  • Capsulated fungi, that that doesn't take up any colour
  • E.g., India ink stain, Nigrosin stain
  • Nigrosin

Impregnation Stains

• Deposit stain on the surface of the object to make it look thicker.
• Silver stains- black colour
• For thin structures
  • Flagella
  • Spirochetes (Eg: Treponema caused syphilis)
    • Syphilis patient with Genital ulcers
• Fontana stain- fluid samples
• Levaditi's stain-tissue samples

Differential Stains

• Game positive cell wall contains a lot of peptidoglycans, which retain the dye longer
• Gram-negative cell wall contains lipopolysaccharides (LPS) making it permeable to the
• secondary dye after decolorization (as alchohol/acetone dissolves & washes away all the lipids in bacterial cell wall).
• Poorly gram staining:(gram stain doesn't work well)
  • Mycoplasma
  • Rickettsia
  • Chlamydia
  • Spirochetes

Albert Stain

• Albert solution 1
  • Malachite Green- Organism
  • Toludine blue - stains volutin granules (Metachromatic stain- 2 colours)
    • Actual colour- blue
    • On sample- purple
  • Glacial Acetic acid
• Albert solution 2: Iodine
• Used for Volutin / Babes Ernt granules
  • present in C-diphtheria
  • located at 2 poles, it is also called bipolar Granules
  • Metachromatic Granules-stained by Toluidine blue.
• Stains for Volutin granules:
  • Ponder's Stain
  • Loeffler methylene Blue (best)
  • Albert Stain
  • Neisser stain
• Volution granules also seen in Spirillum, Gardnerella, Yersinia pestis, yeast, MTB

Spore Stain

• Spore- resting/ dormant form of bacteria.
• Schaeffer and Fulton stain/ Modified Ashby stain
  • Malachite green- spore
  • Heat- Mordant
  • Water- decolorizer
  • Safranin Red- bacteria/organism

MTB Stained with Auramine & Rhodamine dye

A dichroic mirror is used in a fluorescence microscope alongside a strong light source, an excitation filter, and an emission filter.

Blood Agar

• Autoclaved nutrient agar cooled to 50°C.
• 5-10% sterile sheep RBCs
• Made rich by adding blood.
• This is a differential media.

Chocolate Agar

• Autoclaved nutrient agar cooled to 70°C.
• 5-10% sterile sheep RBCs
• More nutritious
• Releases factor: V and X factors

Loeffler's Serum Slope

• Corynebacterium diphtheria
  • Aka Klebs Loffler's media
  • Enriched media
  • time taken to grow - 8 hrs

Lowenstein Jensen Medium: Used for Tuberculosis

1. Coagulated hen's egg
2. Mineral salt solution
3. Asparagine
4. Malachite green (inhibit other bacteria)

MacConkey Agar

• Composition: Made of (mnemonic PLANT)
  • Peptone
  • Lactose (differential medium: lactose fermentation +/-)
  • Agar
  • Neutral Red (indicator medium): Acts as an indicator
    • Fermentation of lactose is pink.
    • If no fermentation, looks pale
  • Taurocholate (Bile salt) (selective)

CLED

• Cysteine Lactose Electrolyte Deficient.
• On lactose fermentation yellow colonies are formed.
• Indicator: Bromothymol blue
• Better than MacConkey- as it is less inhibitory (non-selective)

Mannitol Salt agar

• Selective Media (has salt- only allows S.aureus grow).
• Mannitol fermentation (differential media)
• Yellow color
• Indicator Media: Phenol Red

Thiosulfate Citrate Bile Salt Sucrose (TCBS)

• Vibrio
• Bile salt makes it selective.
• Indicator: Bromothymol blue.
• Differential media (sucrose fermentation +/-)

Potassium Tellurite Agar

• Black color
• C. diphtheriae
  • Enriched media(loeffler serum slope): 8 hours
  • Selective media(PTA): 48 hours

Culture Technologies

Streak Culture

• Take a nichrome loop, sterilize and dip the sample
• Start streaking 90 degree.
• To Obtain Pure colonies.

Stroke Culture

• Done in a test tube: Sterilize, pickup sample and streak again
• Stroke culture made in tubes containing agar slope/slant
• Uses: Provide a pure growth for the bacterium of slide agglutination and other diagnostic tests
• Example: Test for urease.

Lawn Culture

• Lawn cultures are obtained by flooding the plate's surface with the bacterium's suspension.
• AKA carpet culture.
• Mainly used for
  • Antibiotic sensitivity testing.
  • Preparation of bacterial antigens and vaccines.
  • Bacteriophage typing.
• Take the sample directly and it put on a culture plate.

Stab Culture

• It is performed by a straight wire, charged with culture material, by puncturing deep inside the agar.
• Straight wire and stab once: put the wire, in and out only once.
• It is used to:
  • Demonstrate gelatin liquefaction.
  • Oxygen requirement of the bacterium.
  • To maintain stock culture for the preservation of bacteria.

Download the PrepLadder app now and unlock a 24-hour FREE trial of premium high-yield content. Access Smarter Video Lectures also in हिंglish, Game Changing Qbank, Audio QBank, Structured Notes, Treasures, Mock test for FREE to ace your NEET PG preparation. Elevate your study experience and gear up for success. Start your journey with PrepLadder today!