Lab practical 1 Flashcards
What is the purpose of aseptic technique?
To prevent contamination during microbial transfer and culture
What are the three types of culture media used in pure culture techniques?
Broth, slant, agar
What is the purpose of the streak technique?
To isolate pure colonies from a mixed culture
What does ‘parfocal’ mean in microscopy?
The ability to keep an object in focus when switching between different objectives
What is total magnification?
ocular lens magnification X objective lens magnification
Differentiate between bright-field, phase contrast, and dark field microscopy.
Bright-field: standard light, stained, both living & dead
Phase contrast: enhances contrast, living cells, unstained
Dark field: illuminates specimen against a dark background, living cells, unstained
What is a common problem in making a smear?
Overheating the slide or using too much sample
What is the difference between a simple stain and a differential stain?
-Simple stain uses one dye (highlight the size, shape & arrangement of bacteria ); differential stain uses multiple dyes to differentiate cell types/structures ,(Gram stain, acid fast stain, endospore stain)
What are the two types of dyes used in staining?
Acidic and basic dyes
What is the significance of cell morphology?
It helps identify the shape and arrangement of cells
What reagents are used in the Gram stain?
Crystal violet, iodine, alcohol, safranin
What could go wrong during the Gram staining procedure?
Improper timing or washing steps leading to incorrect results
How do you interpret Gram reaction?
Gram-positive: purple; Gram-negative: pink
What reagents are used in the spore stain?
Malachite green, safranin
What is the purpose of the acid-fast stain?
To identify acid-fast bacteria such as Mycobacterium
What is the purpose of a capsule stain?
To visualize the capsule surrounding certain bacteria
What is the indicator used in TTC motility media?
TTC (2,3,5-triphenyltetrazolium chloride)
How can you distinguish between motile and non-motile bacteria?
By observing growth patterns in motility media
What is the purpose of a serial dilution?
To reduce the concentration of bacteria for accurate counting
What is the function of a Quebec colony counter?
To count bacterial colonies on a plate
What is the purpose of the pour plate technique?
To isolate and quantify bacteria in a sample
How does UV light affect bacteria?
It causes DNA damage leading to cell death
What is the difference between resident and transient flora?
Resident flora are permanent; transient flora are temporary
What is measured in the zones of inhibition?
The effectiveness of antibiotics on bacterial growth
What are the major structural differences between yeasts and molds?
Yeasts are unicellular; molds are multicellular with hyphae
How are protozoa classified?
Based on their movement and reproductive methods
Fill in the blank: The major classes of fungi include _______.
[Ascomycetes, Basidiomycetes, Zygomycetes, Chytridiomycetes]
what is a broth?
-a liquid medium that supports bacterial growth
-grows large # of bacteria, does NOT isolate cultures
-no agar agar for fast luxuriant growth
what is a slant?
Agar solidified in a tube with a slanted surface store bacterial long term, grow in oxygen-rich areas
what is an agar?
-solid medium used in petri dishes, isolating and growing bacteria
-solidifying agent (1.5%-2%) for isolation & purification of microbes
How to prepare a specimen?
1.) A small amount of water
2.) spread specimen very thinly
3.) Air dry
4.) Pass over flame once to heat fix bacteria ( kill specimen protein coagulation & adherence to slide)
what is a basic dye?
positive charge
-attracted to slightly neagative surface of the cell
ex: crystal violet, safranin, methylene blue, malachite green, carbol fuschin
what is an acidic dye?
negative charge
-repelled by surface of the cell
used to stain area around the cell
ex: nigrosine, congo red
what is a gram stain?
1.)crystal violet (primary dye-binds with peptidoglycan)
1 min
2.)Grams iodine (mordant)
1 min
rinse with water
3.)Aceton-alcohol (decolorizer)=causes it to shrink
10 sec
rinse with water
4.)Safranin (counterstain)
1min
rinse with water
What is an endospore stain?
1.) add paper towel flood with malachite green steam 5 min (primary stain)
2.) rinse with water
3.) Counterstain-safranin 1 min (binds to vegatative cell
what is Acid-fast staining?
1.) add paper towel with carbol fuchsin steam. 5 min
2.)remove paper & rinse with water
3.) decolorizer-acid alcohol, 10 sec, rinse with water
3.) counterstain with methylene blue 1min & rinse with water
what is a capsule stain
thick polysaccharide layer surrounding the cell
-helps attach to surfaces
-protects against phagocytes
stain background and cell
protects being ingested by white blood cells
capsule is nonionic
dyes dont bind
Streptococcus pneumonia
known for having a prominent capsule that aids in virulence
klebsiella & Neisseria
most likely to produce a capsule
Dilution factor formula
amount of sample/amount of sample + amount in tube
total dilution formula
Tube 1 dilution X Tube 2 dilution
how does radiation kill
induces DNA mutations
ionizing radiation
(x-ray & gamma-ray)
shorter wavelength=stronger effect
pulls electrons from DNA
UV
-causes thymine dymers
-affects the cells ability to replicate DNA
-UV-c is the most germicidal but does not penetrate plastic/glass/clothes
acidophile
below pH 5
acid tolerant
prefer neutral, but can grow in acid
neutrophile
6.5-7.5 pH
alkalinophile
pH 8+
mesophile
20-40 deg C
Body 37 deg C
pathogen
(normal typical microbes)
psychrophiles
0-15 deg C
die above 20 deg
Live in the snow, ice
Not pathogenic
reason for spoilage in fridge
(like the cold)
Thermophiles
40 deg + C
compost hot springs
(very warm)
Hyperthermophiles
80-100 deg C
Hydrothermal vents
(boiling/extremeophiles)
what does a larger zone indicate
senstivity/ susceptibility
what does it mean when the bacteria is resistent to the antibiotic
bacterial culture shows NO zone of inhibition around an antibiotic disk
what is an antibiotic
antimicrobial chemical
(made by microbes to kill microbes)
