lab 2, 3, 4

Question 1

agar

Answer 1

solidifies medium
relatively stable polysaccharide
wide variety of heterotrophic microbes will grow on the surface of a t-soy plate by absorbing the nutrients out of the solid plate beneath them
agar cant be degraded by most terrestrial microbes, so the plate remains solid even when colonies have consumed most of the other nutrients in the plate

Question 2

to count bacteria perform a standard plate count

Answer 2

a bacterial culture or a sample of water or food is spread on an agar plate and the number of colonies is counted through a simple calculation, you can determine the titre of bacteria in the original sample
the titre or concentration of bacteria determined by a standard plate count is expressed as the number of colony forming units per ml or (cfu/ml)

Question 3

to carry out standard plate count there are two techniques

Answer 3

pour plate

spread plate

Question 4

serial dilutions

Answer 4

a bacterial culture may have billions of bacterial cells in it. to makes this number more manageable, we must dilute the original sample so that we can obtain a plate with a countable number of colonies

Question 5

ten fold serial dilution

Answer 5

0.1

Question 6

only use plate counts from…

Answer 6

30-300

Question 7

TNTC

Answer 7

too numerous to count

Question 8

streak plate technique

Answer 8

dilute the culture as you spread it around so that in the end, you have nicely isolated colonies

Question 9

an isolated colongy is a

Answer 9

colony that is not touching any other colonies on the plate

Question 10

pure culture

Answer 10

if all of your colonies appear exactly the same

Question 11

inoculating loop

Answer 11

used to pic colonies off a plate or to obtain a drop of liquid culture
the wire heats up and cools down very quickly

Question 12

inoculating wire

Answer 12

pick up very small amounts of bacteria
pick a colony from a plate that is located very close to another colony
and to inoculate stab tubes

Question 13

media (lab manual page 36)

Answer 13

singular medium
nutrient source for growing bacteria
can be solid liquid and can be prepared in a variety of containers such as tubes , flasks or plates

Question 14

pour plate technique is commonly used to enumerate bacteria in food samples

Answer 14

because it does an excellent job of isolating colonies

Question 15

it is difficult to observe bacteria using a brightfield microscope without

Answer 15

staining first as most bacteria are colourless

Question 16

stain is composed of salts of coloured compounds suspended in a liquid usually water

Answer 16

charged portion of the salt molecule, called the chromophore, can have either gram positive or gram negative charge
bacteria have a net negative charge on their outer surface

Question 17

basic stain AKA cationic stain

Answer 17

positively charged chromophore
these stains are attracted to the negatively charged surface of the cell and can be used for staining cell surfaces and structures 
-crystal violet
-safranin
-methylene blue

Question 18

acidic stain or anionic

Answer 18

negatively charged chromophore
these stains are repelled by the negatively charged cell surface and can be used to stain the background of the slide in a process called negative staining
-nigrosin
-india ink

Question 19

bacteria most common shapes are

Answer 19

cocci, bacilli

spirillia

Question 20

not all bacteria have a characteristic arrangement

Answer 20

singular
or random
most rod shaped bacteria occur as singles but can look like pairs when they divide

Question 21

Micrococcus luetus simple stain

Answer 21

very small cocci arranged in tetrads

gram positive

Question 22

gram positives

Answer 22

have a negative charge on their surface because of teichoic acids embedded in the thick peptidoglycan layer

Question 23

gram negatives

Answer 23

have a negative charge on their surface bc of the charged sugars and phosphates in the lipopolysaccharide component of the outer membrane

Question 24

both gram negative and gram positives have

Answer 24

negatively charged surfaces

Question 25

E. coli

Answer 25

gram negative
bacilli
singular

Question 26

B. su

Answer 26

gram positive
bacilli
either singular or streptobacilli

Question 27

s. ep

Answer 27

gram positive
coccus
staphylococci

Question 28

why is alcohol destaining the differential step

Answer 28

bc gram positives and gram negatives will react differently to this treatment
this is the step that lets you see a difference between the two cell types
-ethanol shrinks the pores of the peptidoglycan layer in the gram positive bacteria, trapping the crystal violet iodine complex
-in gram negative the ethanol disrupts the outer membrane which allows the loosely trapped crystal violet molecules to be washed out of the thing peptidoglycan layer

Question 29

safranin is a basic counter stain

Answer 29

a counterstain is a stain that sticks to structures not bound by a primary stain

• when counterstains are used, they are different colour from the primary stain
• applied after the primary stain so that cells that did not retain the primary stain take on the colour of safranin
• colouring the bacteria that was destained
• bacteria that have retained the crystal violet even after destained will be stained with safranin, but since crystal violet is much darker the cells remain purple

Question 30

crystal violet basic stain

Answer 30

cv particles get trapped in the peptidoglycan layer of the cell wall

• gram positive bacteria have a thick layer of peptidoglycan, therefore many dye particles are trapped
• in gram negative bacteria a semi permeable outer membrane only allows a few dye particles to pass through and get tapped in the thin peptidoglycan layer but the negative charge on the outer surface of the cell also attracts the crystal violet

Question 31

iodine a mordant

Answer 31

mordant can be anything that acts as a helper to increase the intensity of the primary stain
heat is used as a mordant for some stains
for gram stain iodine is the mordant
iodine acts by forming complexes with the crystal violet particles, causing them to increase in size and be trapped more effectively by the peptidoglycan

Question 32

heat fixing

Answer 32

attaches the cells to the slide so that they dont come off after repeated washes

Question 33

KOH string test

Answer 33

-rapid means of confirming the gram reaction
-by adding 3% KOH to bacteria , two reactions may happen
-gram positives will exhibit no reaction at all
-gram negative will form a slimy viscous solution that strings as the loop is pulled away from the slide
the gram negative cell walls are disrupted by the alkaline conditions created by the KOH and the strings form from the release of DNA from the inside the cell
gram positive cell walls are not susceptible to these conditions and dont break down as readily

Question 34

m. lu

Answer 34

umbonate elevation 
yellow
circular
entire margin
opaque 
glossy