Microscopy

Question 1

Microscopy findings supportive of a diagnosis of candida?

Answer 1

Presence of: - blastospores - pseudohyphae - neutrophils

Gram +ve yeast
Abundant branched pseudohyphae
Blastoconidia = grape like clusters along the length of the hyphae
can form a biofilm

Question 2

On microscopy what may the presence of neutrophils in vaginal secretions suggest

Answer 2

an inflammatory response and therefore presence of infection

Question 3

On microscopy what may the absence of neutrophils in the presence of candida in vaginal secretions suggest

Answer 3

Absence of neutrophils in the presence of Candida is likely to represent colonisation

Question 4

Appearance of GC on microscopy

Answer 4

monomophicgram negativediplococciwithin polymorphonuclear leukocytes

Question 5

On a microscope what is the name of the place where you put the specimen that you want to examine

Answer 5

Stage

Question 6

On a microscope what are the names of the metal parts that hold th slide in place

Answer 6

Stage clips

Question 7

On a microscope what is the name of the part that you look down

Answer 7

Eyepiece or occular

Question 8

On a microscope what is the name of the part that holds 2+objective lenses and can be rotated to easily change power (magnification)

Answer 8

The revolving nosepiece or turret

Question 9

On a microscopewhat is the name of the 3or 4 parts which can be rotated to alter thepower

Answer 9

Objective Lenses

Question 10

On a microscopewhat is the name of the part where the light comes from

Answer 10

The illuminator

Question 11

On a microscopewhat is the name of the part which connects the eyepiece to the objective lenses

Answer 11

The tube

Question 12

On a microscopewhat is the name of two focus knobs

Answer 12

Coarse focus

Fine focus

Question 13

After Gram staining what colour do Gram-positive bacteria appear

Answer 13

Gram-positive bacteria = violet / purplein colour

after gram staining - retain most of the crystal violet

Question 14

After gram staining what colour do gram negative bacteria appear

Answer 14

gram-negative bacteria = pink colour

after Gram staining

Question 15

For gram stain what 4 reagents are used in sequence

Answer 15

sequential application of:
crystal violet 
iodine 
alcohol 
safranin

Question 16

What is the role of crystal violet in gram staining

Answer 16

Crystal violet = the basic dye which acts as the “Primary stain”
It stains all the micro-organisms to appear violet in colour
Applied 1st in gram staining

Question 17

what is the role of Grams Iodine in gram staining

Answer 17

Gram’s iodine -acts as “Mordant dye”
increases the affinity between the cell and stain by forming a complex with crystal violet known as “CV-I complex”

Applied 2nd

Question 18

What is the role of 95% Alcohol in the gram staining process

Answer 18

95% Alcohol -acts as the “Decolourizing agent”
functions as lipid solvent
dissolves the lipid present in the cell wall
causes decolourization of the cell by the removal of the CV-I complex from the bacterial cell

Applied 3rd

Question 19

what is the role of Safranin in the process of gram staining

Answer 19

Safranin = a “Counter stain”
stains the decolourized bacterial cell by giving pink colour to the cell

applied 4th

Question 20

what are the steps involved with preparing a gram stained slide

Answer 20

1. Take a clean dry slide and place one drop of distilled water at the centre.
2. Prepare bacterial smear by taking small inoculum from the bacterial culture by the help of the inoculating loop.
3. mix the inoculum with the water drop to prepare thin bacterial smear by rotating the inoculating loop in a clockwise and anticlockwise direction.
4. heat fixes the bacterial smear under the spirit lamp.
5. flood the bacterial smear with crystal violet and allow to stand for 1minute.
6. flood the smear with Gram’s iodine.
7. add the decolourizing agent i.e. 95% Ethanol to the smear.
8. add counter stain i.e. Safranin to the smear and allow to stand for 1 minute.
9. wash the slide with water and air dry .
10. observe the glass slide under the microscope by adding oil immersion to the stained cells to differentiate gram-positive and negative cells.

Question 21

Interpretation of Gram Staining - if abacteria cell isstained Violet

Answer 21

Gram-Positive - will stain Violet.

Question 22

Interpretation of Gram Staining -If a bacteria stains Pink

Answer 22

Gram-negative: will stain Pink.

Question 23

Interpretation of Gram Staining - what colour does a Gram-Positive bacteria stain

Answer 23

violet

Question 24

Interpretation of Gram Staining - what colour does a Gram negative bacteria stain

Answer 24

pink

Question 25

what is a positive gram reaction

Answer 25

stains puple

= Gram positive bacteria

Question 26

what is a negative gram reaction

Answer 26

Stains pink

= gram negative

Question 27

What part of thebacterial cell does the gram staining processadd colour to?

Answer 27

the cell wall

Question 28

what is the the difference between Gram-positive and Gram-negative bacteria

Answer 28

Difference in the cell wall
Gram-positive bacteria - high peptidoglycan content (70-80%) + low lipid content (1-4%) - lacks lipopolysaccharide - lacks porins on the outer membrane -has less periplasmic space

Gram-negative bacteria- low peptidoglycan content (10-20%) and high lipid content (20-30%) - outer membrane is present + contains lipopolysaccharide, porins and has more periplasmic space