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HLSC 124 - Microbiology for Health Professionals > Chapter 3 - Observing Microorganisms Through a Microscope > Flashcards

Chapter 3 - Observing Microorganisms Through a Microscope Flashcards

1
Q

Ocular Lens/Eyepiece

A

-remagnifies the image formed by the objective lens
-the part you look through

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2
Q

Body Tube

A

-transmits the image from the objective lens the the ocular lens

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3
Q

Objective Lenses

A

-primary lenses that magnify the specimen
-the rotating piece with different magnifications

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4
Q

Stage

A

-holds the microscope slide in position

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5
Q

Condenser

A

-located underneath the stage
-focuses light through the specimen

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6
Q

Diaphragm

A

-controls amount of light entering the condenser

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7
Q

Illuminator

A

-light source

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8
Q

Total Magnification

A

-calculated by multiplying the objective lens magnification by the ocular lens magnification

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9
Q

Resolution (aka resolving power)

A

-ability of lenses to distinguish fine detail and structure

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10
Q

Refractive Index

A

-a measure of the light-bending ability of a medium

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11
Q

Electron Microscope

A

-beam of elections is used instead of light
-resolving power is greater

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12
Q

Staining

A

-colouring the microorganisms with a dye that emphasizes certain structures
-the structures must be fixed (attached) to the slide

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13
Q

Stain Composition

A

-salts composed of a positive and negative ion

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14
Q

Smear

A

-thin film containing the microorganism

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15
Q

Chromophore

A

-either the positive or negative ion that is coloured

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16
Q

Basic Dyes

A

-most common stain used in microbiology
-dyes in the cation (positive ion)

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17
Q

Acidic Dyes

A

-rarely used in microbiology
-dyes on the anion (negative ion)
-ie. Glycocalyx Satin

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18
Q

Why is basic dye more popular?

A

-bacteria are slightly negatively charged at pH 7
-the coloured cation in a basic dye is attracted to the negatively charged bacterial cell

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19
Q

Why is acidic dye less common?

A

-not attracted to most types of bacteria because the dyes negative ions are repelled by the negatively charged bacteria surface
-stain colours the background instead

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20
Q

Negative Staining

A

-preparing colourless bacteria against a coloured background
-valuable for observing cell shapes, sizes, and capsuls

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21
Q

Simple Stain

A

-aqueous or alcohol solution composed of one basic dye
-used to highlight the entire microorganism
-recognizes shape and structure
-too simple for healthcare

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22
Q

Mordant

A

-chemical added to the solution to intensify the stain
-makes it darker, hold the stain, or enlarge the specimen

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23
Q

2 Functions of a Mordant

A

1) Increased the affinity of a stain
2) Coat a structure (ie. flagellum)

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24
Q

Differential Stains

A

-react differently with different kinds of bacteria
-can be used distinguish them
-2 types: Gram stain and acid-fast stain

25
Gram Stain
-developed by Hans Christian Gram -classifies bacteria into two large groups: Gram-positive and gram-negative
26
Gram Stain Procedure Step 1
-Primary stain -Crystal violet -smear is covered with this purple and colours all cells -Results: both bacteria turn purple
27
Gram Stain Procedure Step 2
-Mordant -Iodine -the purple dye is washed off and the smear is covered with iodine -Results: both bacteria turn purple
28
Gram Stain Procedure Step 3
-Decolourizing Agent -Acetone alcohol -the slide is washed with alcohol solution -removed purple from the gram negative cells -gram positive cells remain purple
29
Gram Stain Procedure Step 4
-Counterstain -Safranin (red) -alcohol is rinsed off and the slide is stained with red dye -smear is washed and examined -Results: gram positive remains purple, gram-negative turns red/pink
30
Gram-positive Bacteria
-retain the purple colour after alcohol has attempted to decolourize
31
Gram-negative Bacteria
-bacteria that lose the purple colour after decolorization -made visible after losing colour by Safranin
32
What differs between gram-positive and gram-negative?
-because of structural differences in the cell walls -affect the retention or escape of the crystal violet-iodine (CV-1) complex
33
Gram-positive Bacteria explanation
-have a thicker peptidoglycan layer -CV-1 complex is larger -compound gets stuck and can't be decolourized
34
Gram-negative Bacteria explanation
-have a thinner peptidoglycan layer -have a lipopolysaccharide layer in their cell wall -alcohol wash disrupts outer lipopolysaccharide layer and CV-1 complex is washed out
35
CV-1
-crystal violet or iodine alone are water soluble and drain out of the cell -CV-1 is insoluble in water so it doesn't easily leave the cell wall
36
Antibiotics
-gram + easily killed by some antibiotics while gram - are more resistant since antibiotics can't penetrate lipopolysaccharide layer
37
Acid-Fast Stain
-binds strongly only to bacteria that have a waxy material (Mycolic acid) in their cell walls -used to detected Mycobacterium tuberculosis and Mycobacterium leprae (leprosy)
38
Acid-Fast Stain Procedure Step 1
-primary stain -carbolfuchsin (red dye) applied to slide and heated for several minutes -results: both cells appear red
39
Heating Smears
-increases penetration and retention of the dye
40
Acid-Fast Stain Procedure Step 2
-Decolorization -acid-alcohol -slide is cooled and washed with water -acid-alcohol removes red stain from bacteria that are not acid-fast
41
Why do acid fast keep their colour?
-carbolfuchsin is more soluble in the cell wall lips than in the acid-alcohol
42
Why do non-acid fast lose their colour?
-cell wall lacks lipid components -able to decolourize leaving cells colourless
43
Acid-Fast Stain Procedure Step 3
-counterstain -methylene blue -smear is stained with blue dye -non-acid-fast cells appear blue -acid-fast remain red
44
So which cells would be positive for TB/Leprosy?
-the red acid-fast cells would be positive for the pathogen -the blue cells are just tissue cells
45
Special Stains
-used to colour parts of microorganisms such as endospores, flagella, or capsules
46
Capsule Stain (Negative Staining)
-determines the organisms virulence -capsular materials are water soluble and may be dislodged during washing (makes this stain difficult) -glycocalyx (acid stain) -a quick stain
47
Capsule
a gelatinous covering of a microorganism
48
Virulence
-the degree to which a pathogen can cause disease
49
Capsule Stain Procedure Step 1
-acidic stain -colours the background (negative stain) -uses India ink or nigrosin to colour background dark blue -allows cells to shine through
50
Capsule Stain Procedure Step 2
-Water wash -negative results can occur from capsular materials washing away
51
Capsule Stain Procedure Step 3
-simple stain -Safranin (red dye) -capsules don't accept most dyes and thus appear as halos surrounding the red cells
52
Endospore (spore) Staining
-7ish min stain -require special stain cause simple dyes don't penetrate endospores wall
53
Endospore
-resistant, dormant structure formed within a cell that protects it from adverse environments
54
Endospore Stain Procedure Step 1
-primary stain -malachite green -applied and heated for 5mins
55
Endospore Stain Procedure Step 2
-water wash -removes the green from all the cells parts except endospores
56
Endospore Stain Procedure Step 3
-counterstain -safranin -stains portions of the cell other than endospores -endospores appear green within the red cell
57
Flagella Stain
-tedious and delicate procedure -2 min stain -flagella are delicate and can break off easily, resulting in a misdiagnosis
58
Flagella
-structures of movement too small to be seen under light microscope without staining
59
Flagella Stain Procedure
-carbolfuchsin (red) and a dark red mordant (potassium alum) are used to build up the flagella diameters until they become visible

HLSC 124 - Microbiology for Health Professionals (18 decks)

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