LAB PRACTICAL

Aseptic technique

preventing contamination of cultures, media, and environment

Inoculation

transfer of microorganisms into media

Broth culture

liquid medium containing bacteria

Agar plate

solid medium used to grow bacteria

Slant agar

solid agar in a tube at an angle

Smear

thin layer of bacteria spread on a slide

Heat fix

process of fixing bacteria to slide using heat

Cocci

round-shaped bacteria

Bacilli

rod-shaped bacteria

Spirilla

spiral-shaped bacteria

Strepto

chain arrangement of cells

Staphylo

cluster (grape-like) arrangement

Sarcina

cubical arrangement of 8 cells

Gram stain

differential stain that separates bacteria based on cell wall structure

Crystal violet

primary stain in Gram stain (colors all cells purple)

Iodine

mordant that fixes crystal violet to cell wall

Decolorizer (ethanol)

removes stain from Gram-negative cells

Safranin

counterstain that colors Gram-negative cells pink

Gram-positive

bacteria with thick peptidoglycan wall, appear purple

Gram-negative

bacteria with thin wall, appear pink

Endospore stain

stain used to detect spores in bacteria

Malachite green

primary stain that colors endospores green

Safranin (endospore stain)

counterstain that colors vegetative cells pink

Endospore

resistant structure formed by some bacteria

Streak plate

method used to isolate single bacterial colonies

Colony

visible growth of bacteria on solid media

Psychrophile

bacteria that grow best in cold temperatures

Mesophile

bacteria that grow best at moderate temperatures (~37°C)

Thermophile

bacteria that grow best in high temperatures

Osmotic pressure

effect of solute concentration on cells

Halophile

organism that thrives in high salt conditions

FTM (Fluid Thioglycolate Medium)

medium used to determine oxygen requirements

Obligate aerobe

requires oxygen to grow

Obligate anaerobe

cannot survive in oxygen

Facultative anaerobe

can grow with or without oxygen

Thermal death time

time required to kill bacteria at a certain temperature

Disinfectant

chemical used to kill microbes on surfaces

Antiseptic

chemical used to kill microbes on living tissue

Mannitol Salt Agar (MSA)

selective and differential medium for Staphylococcus

Selective media

allows some organisms to grow while inhibiting others

Differential media

distinguishes organisms based on biochemical reactions

Phenol red

pH indicator that turns yellow in acid and pink in base

Mannitol fermentation

breakdown of mannitol producing acid (yellow color)

Eosin Methylene Blue (EMB)

selective and differential medium for Gram-negative bacteria

Lactose fermentation

breakdown of lactose producing acid

Green metallic sheen

strong lactose fermentation (E. coli)

Hektoen Enteric Agar (HE)

selective medium for Gram-negative enteric pathogens

Bile salts

inhibit Gram-positive bacteria

Hydrogen sulfide (H2S) production

results in black precipitate

Phenol Red Broth

test for carbohydrate fermentation

Durham tube

small tube that traps gas produced during fermentation

Citrate test

determines if bacteria can use citrate as carbon source

Citrate permease

enzyme needed to transport citrate into cell

Bromothymol blue

pH indicator that turns blue in alkaline conditions

Catalase test

detects presence of catalase enzyme

Catalase

enzyme that breaks down hydrogen peroxide into water and oxygen

Starch hydrolysis

breakdown of starch by amylase

Amylase

enzyme that digests starch

Iodine (starch test)

indicator that turns black in presence of starch

Gelatin hydrolysis

breakdown of gelatin by gelatinase

Gelatinase

enzyme that digests gelatin into amino acids

Urea hydrolysis

breakdown of urea by urease

Urease

enzyme that converts urea into ammonia

Oxidase test

detects cytochrome c oxidase enzyme

Cytochrome c oxidase

enzyme in electron transport chain

DNA hydrolysis

breakdown of DNA by DNase

DNase

enzyme that degrades DNA

SIM medium

test for sulfur reduction, indole production, and motility

Sulfur reduction

production of hydrogen sulfide (black color)

Indole production

breakdown of tryptophan into indole

Kovac’s reagent

indicator that turns red if indole is present

Motility

ability of bacteria to move

Blood agar

enriched and differential medium with red blood cells

Hemolysis

breakdown of red blood cells

Alpha hemolysis

partial hemolysis (green color)

Beta hemolysis

complete hemolysis (clear zone)

Gamma hemolysis

no hemolysis

Kirby-Bauer test

method to test antibiotic susceptibility

Mueller-Hinton agar

medium used for antibiotic testing

Zone of inhibition

clear area where bacteria did not grow around antibiotic

Sensitive (S)

bacteria are inhibited by antibiotic

Resistant (R)

bacteria are not affected by antibiotic

Intermediate (I)

bacteria show partial sensitivity

Gram stain order

crystal violet → iodine → alcohol → safranin

Endospore stain key step

steaming allows stain to enter spores

Streak plate goal

isolate single colonies

Aseptic technique key step

flame loop and tube openings

Gram positive result

purple

Gram negative result

pink

MSA positive

yellow (mannitol fermentation)

MSA negative

pink

EMB E. coli result

green metallic sheen

Hektoen Salmonella result

black precipitate

Catalase positive

bubbles

Oxidase positive

purple

Starch positive

clear halo after iodine

Gelatin positive

liquid

Urea positive

pink

Citrate positive

blue