Practical Lab Final

Question 1

Define ocular lens (what is it called, what does it do, and what is its magnification?)

Answer 1

Binocular lens. Allows both eyes to focus on the specimen. Has a magnification of 10x.

Question 2

Define ocular adjustment

Answer 2

Allows you to adjust each separate ocular eyepiece for people who have differences between their eyes

Question 3

Define revolving nosepiece/ turret

Answer 3

Holds objective lenses. Rotation of this allows the objective lenses to ‘click’ into place for viewing

Question 4

1) Define objective lenses.
2) What magnifications are they?
3) What total magnifications do they provide?

Answer 4

1) The lenses on the revolving turret. Provide the major magnification of the microscope.
2) 4x, 10x, 40x, 100x.
3) The ocular lens (10x) times the magnification (ex: 40x * 10x = 400x total)

Question 5

1) Where is a slide placed on a microscope?
2) What holds the slides in place for observation?
3) What knob moves the stage to view different parts of the slide?

Answer 5

1) The stage
2) The stage clips/ slide clamp
3) The stage adjustment

Question 6

1) What is the series of lenses that focuses the light traveling from the light source onto the specimen and allow for bright field microscopy?
2) What expands or reduces the amount of light hitting the specimen?
3) What is a power control that adjusts the power from the light source? What does higher power equal?

Answer 6

1) Condenser
2) Iris diaphragm
3) Rheostat; higher power = brighter light. Ranges 0-10.

Question 7

1) What is the light bulb on a microscope called?
2) What type of focus should only be used on lower power objectives (because it moves the stage in large increments)?
3) What type of focus can be used on higher power objectives?

Answer 7

1) Light source
2) Course focus
3) Fine focus

Question 8

What turns the microscope on and off?

Answer 8

Power switch

Question 9

Gram stain:
1) What color are gram + species? Give examples.
2) What color are gram - species? Give examples.

Answer 9

1) Purple. Ex: Staphylococcus aureus & S. epidermidis, B. subtilis and B. cereus.
2) Red. Ex: E. coli, Salmonella enteritidis, S. sonnei, K. pneumoniae, P. vulgaris.

Question 10

1) What color are bacteria on an endospore stain? What color are endospores?
2) Acid fast stain: What color are acid-fast bacteria? Why? What color are non-acid-fast bacteria? Why?

Answer 10

1) Red; endospores are green
2) Acid-fast: red (carbol fuchsin); non-acid-fast: blue (methylene blue)

Question 11

1) What is a statistically valid plate count?
2) How do you count the number of cells on a plate?
3) What is the equation for colony count?

Answer 11

1) Between 30 and 300 colonies
2) #colonies / TDF = # of cells
3) #cells per mL X TDF = colony count

Question 12

1&2) What are the two methods used to generate a bacterial growth curve in lab?
3) What are the 4 phases of a bacterial growth curve?

Answer 12

1) A viable plate count and 2) turbidity measurement using a Nephlo flask and a spectrophotometer
3) Lag, log, stationary, and death

Question 13

Which two enzyme tests required a stab inoculation?

Answer 13

Gelatinase and SIM

Question 14

Which 3 enzyme tests were streaked for isolated colonies?

Answer 14

1) SM110
2) MS
3) TSA

Question 15

Why does the lysine test require mineral oil?

Answer 15

To make the broth an anaerobic environment to force the culture to ferment glucose

Question 16

Which 3 enzyme tests required a loopful into broth?

Answer 16

1) Nitrate
2) Coagulase (TSB)
3) Urea

Question 17

Which 2 enzyme tests required a line inoculation?

Answer 17

1) DNase
2) Starch (amylase)

Question 18

Name 5 tests that differentiate between S. aureus and S. epidermidis

Answer 18

1) MSA
2) BA
3) SM-110
4) Coagulase
5) DNase

Question 19

What 2 tests differentiate between BS and BC?

Answer 19

1) MSA
2) LM

Question 20

Name 2 tests useful in the identification of the genus Proteus

Answer 20

1) PHE
2) Urea

Question 21

Which tests should be used for G+ rods? (1)

Answer 21

LM (litmus milk)

Question 22

Which tests should be used for G+ cocci? (5)

Answer 22

1) BA
2) MSA
3) Coagulase (SA+; SE-)
4) SM-110
5) MS

Question 23

Which tests should be used for G- rods (6)?

Answer 23

1) SS
2) MAC
3) TSI
4) Citrate (EC-; EA+)
5) VP (EC-; EA+)
6) Methyl Red (EC+; EA+)

Question 24

Which tests should be used for G- cocci? (1)

Answer 24

1) SIM

Question 25

Which tests should be used for G- bacteria in general? (1)

Answer 25

EMB

Question 26

Which tests are differential only? (6)

Answer 26

1) LM 2) TSI 3) Citrate 4) VP 5) MR 6) SIM

Question 27

Which tests are selective only? (2)

Answer 27

Coagulase and BA

Question 28

Which tests are both selective and differential? (6)

Answer 28

1) SS 2) SM-110 3) MAC 4) MSA 5) MS 6) EMB

Question 29

List the enzymes the following tests look for the presence of, and state what each enzyme degrades: 1) DNase 2) Urea 3) Nutrient starch 4) LYS 5) Nutrient gelatin 6) Catalase 7) PHE 8) ONPG 9) Coagulase 10) MS

Answer 29

1) DNase: DNase, DNA 2) Urea: Urease, urea 3) Nutrient starch: Amylase, starch 4) LYS: Lysine decarboxylase, amino acid 5) Nutrient gelatin: Gelatinase, gelatin 6) Catalase: Catalase, H2O2 7) PHE: Phenylalanine deaminase, phenylalanine 8) ONPG: B-galactosidase; lactose fermentation 9) Coagulase: Coagulase, protein clotting 10) MS: Saccharase; sucrose

Question 30

What question does a NO3 (nitrate) test answer?

Answer 30

Does bacteria use NO3- as a terminal electron acceptor in anaerobic respiration (and denitrify it to N2)?

Question 31

1) What does a SIM test test for? (3 things) 2) What does an EMB test test for? (2 things)

Answer 31

1) SIM: Motility and H2S to FeS production (some G- cocci); indole production (from tryptophan breakdown) (EC+; EA-) 2) EMB: Ability to grow on EMB's eosin Y & methylene blue (G-) and coliform differentiation (by color and lactose fermentation)

Question 32

1) What does an SM-110 test test for? (3 things) What bacteria are positive? 2) What does an MS test test for? What bacteria are positive?

Answer 32

1) Ability to grow on NaCl (G+); gelatinase presence; glycolysis of fructose-phosphate from mannitol (G+ cocci: SA+; SE-) 2) Ability to grow on MS's sucrose due to saccharase (G+ cocci: SM and EF+)

Question 33

What do the following tests look for? 1) Methyl red 2) VP 3) Citrate

Answer 33

1) Ability to ferment (make) mixed acids, decreasing pH (EC+, EA-) 2) Ability to ferment alcohol, creating intermediate acetylmethylcarbinol (EC-; EA+) 3) Ability to grow on citrate which releases ammonia, increasing pH (EC-; EA+)

Question 34

What does a TSI test look for? (3 things) Include the color of each thing

Answer 34

1) Ability to ferment glucose (red over yellow, K/A) 2) Ability to ferment lactose and/or sucrose (all yellow, A/A) 3) H2S production (black)

Question 35

What does an MAC test look for? (2 things) Include what species are positive for each thing.

Answer 35

1) Ability to grow on MAC's CV and bile salts (G-) 2) Ability to ferment lactose (EC+, red)

Question 36

What does an SS test look for? (3 things). Include what species are positive for each thing.

Answer 36

1) Ability to grow on SS's bile salts (G- non-coliform rods) 2) Ability to ferment lactose (EC+, red). Neutral red indicator 3) H2S production (Salmonella+)

Question 37

What does an MSA test look for? (2 things) Include what species are positive for each thing.

Answer 37

1) Ability to grow on MSA's high NaCl (G+ cocci: Bacillus and staphylococcus) 2) Ability to ferment mannitol (SA+)

Question 38

1) What genuses are a BA test differential for? 2) What question does a BA test answer? Include which species are positive.

Answer 38

1) Staph and strep 2) Does bacteria do gamma, alpha, or beta hemolysis? (G+ cocci: SA does beta; SE does gamma)

Question 39

What 2 things does a LM test look for? Include what species is positive.

Answer 39

1) Ability to ferment lactose and produce acid, decreasing pH (Bacillus) 2) Ability to break down casein protein in milk

Question 40

1) What 2 tests use neutral red indicator? 2) What color is neutral red indicator when introduced to an acid?

Answer 40

1) SS and MAC 2) Red: acidic

Question 41

1) What 3 tests use phenol red indicator? 2) What color is phenol red when introduced to an acid?

Answer 41

1) MSA, TSI, Urea 2) Yellow: acidic

Question 42

1) What test uses litmus milk as an indicator? What color is it when positive? 2) What test uses methyl green as an indicator? What color is it when positive? 3) What test uses bromcresol purple as an indicator? 4) What test uses bromothymol blue as an indicator?

Answer 42

1) LM: litmus milk (white: acidic = +) 2) DNase: methyl green (clearing = +) 3) LYS: bromocresol purple (purple/black: basic = +) 4) Citrate: bromothymol blue (vivid blue: basic = +)

Question 43

Which 4 tests can help differentiate between Staphylococcus aureus (SA) and Staphylococcus epidermidis (SE)? Report when each is positive and negative

Answer 43

1) Coagulase (TSB): SA+; SE- 2) BA: SA beta; SE gamma 3) MSA: SA+; SE- 4) SM-110: SA+; SE-

Question 44

Salmonella and shigella grow well on which two media?

Answer 44

MAC and SS

Question 45

1) What two bacterial species ferment mannitol (MSA test)? What color does this result in? 2) What two bacterial species do not ferment on MSA? What color does this result in?

Answer 45

1) S. aureus (SA) and B. subtilis (BS): turn yellow (acidic) 2) S. epidermidis (SE) and B. cereus (BS): stay red (basic)

Question 46

What 7 media can help differentiate between E. coli (EC) and E. aerogenes?

Answer 46

1) VP: EC-; EA+ 2) Citrate: EC-; EA+ 3) Indole: EC+; EA- 4) Methyl red: EC+; EA- 5) MAC: EC+; EA- 6) SS: EC+; EA- 7) SIM: EC+; EA-

Question 47

List the 3 media that use phenol red and what color they are when positive

Answer 47

1) MSA: Turns yellow when positive 2) TSI: Red is 'k', yellow is 'a' 3) Urea: Turns red when positive

Question 48

Name the 4 antiseptics/ disinfectants used in the bacterial death exercise

Answer 48

1) Clorox (QAC) 2) Pinesol (DET) 3) Greenworks (CIT) 4) Iodine (IO)

Question 49

What type of disinfectant/ antiseptic is clorox?

Answer 49

A quaternary ammonium compound

Question 50

What type of disinfectant/ antiseptic is pinesol?

Answer 50

A detergent

Question 51

What is greenworks?

Answer 51

Citric acid

Question 52

What two characteristics would you look for in a test regarding the efficacy of chemical agents on bacteria?

Answer 52

1) Quantitative 2) Reproducible

Question 53

What 3 categories of techniques were used to kill bacteria in the bacterial death exercise?

Answer 53

1) Disinfectants/ antiseptics 2) Antibiotics 3) UV radiation

Question 54

What 4 categories antibiotics were used to inhibit bacterial growth in the bacterial death lab? What antibiotic was used to represent each category?

Answer 54

1) Nucleic acid inhibitor: RIF 2) Cell wall inhibitor: AMP 3) Protein synthesis inhibitor: TET 4) Structural analog: SXT

Question 55

1) What type of test was used to measure the efficacy of disinfectants/ antiseptics and antibiotics on bacteria? 2) How is this test interpreted?

Answer 55

1) Disk diffusion test 2) If there is an area of no growth around the disc, the agent was effective

Question 56

How many seconds of UV radiation did each condition get in the bacterial death lab?

Answer 56

0, 10s, 30s, 1m, 2m, 5m, 10m

Question 57

1) What was the first test used to measure the efficacy of chemical agents on bacteria? Why was it not preferable? 2) What test replaced that test? 3) What test replaced the second test?

Answer 57

1) Broth dilution testing; time consuming and resource intensive 2) Disk diffusion testing 3) E-test strips

Question 58

What are the steps of a disk diffusion test?

Answer 58

1) Streak a plate to create a lawn of bacteria 2) Apply antibiotic or disinfectant disk 3) Incubate 4) Zone of inhibition is created around the disk if agent is effective (i.e. no bacteria around disk)

Question 59

Why is there a gradient observed around the disk in the disk diffusion test?

Answer 59

Because the agent diffuses from the disk into the agar medium

Question 60

Why did the BS 5-day old culture grow better than the newer culture under UV conditions?

Answer 60

Because the bacteria had sporulated

Question 61

Is UV radiation more effective against G+ or G- bacteria? Why?

Answer 61

More effective against G- bacteria; G+ bacteria that can form spores are more protected

Question 62

Name 5 environmental conditions that can influence bacterial growth

Answer 62

1) Oxygen 2) Osmotic pressure 3) pH 4) Temperature ranges 5) Liquid water presence

Question 63

1) What one species (that we work with) clarifies litmus milk? 2) What species is a denitrifier? 3) What 2 things makes P. vulgaris unique?

Answer 63

1) B. subtilis 2) Ps. aeruginosa 3) It makes an alkaline broth in urea and breaks down phenylalanine to make PPA

Question 64

1) How does a Brewer jar work? 2) How does a candle jar work?

Answer 64

1) They become anaerobic by using a GasPak; the Pak generates H2 and CO2; the H2 reacts with the oxygen in the jar to form water vapor by the use of a catalyst. 2) A lit candle is added to a jar and tightly sealed; the flame makes CO2 and extinguishes itself when the O2 concentration is too low (6-8%).

Question 65

1) Name 3 ways to enumerate (count) bacteria 2) What is one way to estimate bacteria count?

Answer 65

1) Viable plate count, MPN, and turbidity measurements using a flow cytometer 2) Spectrophotometer to measure turbidity.

Question 66

1) How can you test to see if someone has S. aureus on their skin or throat? 2) How can you test to see if someone has yeast on their skin?

Answer 66

1) MSA plate (turns yellow) 2) SDA (sabouraud dextrose agar)

Question 67

What two places of the body were microbes isolated from?

Answer 67

1) The skin (behind the ear) 2) The oral cavity (back of the throat)

Question 68

What plates were inoculated with the oral cavity swab?

Answer 68

1) TSA 2) BA 3) MSA

Question 69

What plates were inoculated for the skin (behind ear) swab?

Answer 69

1) TSA 2) MSA 3) SDA

Question 70

Why was the SDA plate used for the skin sample?

Answer 70

To isolate any yeast growing on skin

Question 71

Name 2 types of bacteria that typically colonize the skin (exterior)

Answer 71

1) Micrococcus 2) Staphylococcus (aureus and epidermidis)

Question 72

Name the 3 typical mucosal (mouth) colonizers

Answer 72

1) Staph aureus 2) Streptococci 3) Enterococcus 4) Corynebacterium

Question 73

Where are bacteria typically abundant in the oral cavity?

Answer 73

Underneath the gums

Question 74

1) What are the yellow bacteria typically isolated from the skin? 2) What are the white bacteria isolated from the skin?

Answer 74

1) Micrococcus: Yellow 2) S. epidermidis and S. aureus: White

Question 75

Name the 3 bacteria typically seen in the mouth

Answer 75

1) Streptococci (tiny colonies) 2) Haemophilus 3) Cyanobacterium

Question 76

What does clearing on a plate of isolated mouth bacteria indicate?

Answer 76

Beta hemolysis, which means it's more likely a pathogen

Question 77

1) What commensal microbes live on the skin? 2) What commensal microbes live in the mouth?

Answer 77

1) Staphylococcus, Micrococcus, Corynebacterium, Pityrosporum (yeast) 2) Staphylococcus, Micrococcus, Streptococcus

Question 78

Name 5 potential pathogens

Answer 78

1) S. aureus 2) St. pyogenes 3) Neisseria 4) Haemophilus 5) S. epidermidis

Question 79

1) What is agar? 2) What should bacterial media contain?

Answer 79

1) Carrageenan (seaweed) 2) Major elements (CHONPS), minor elements (Fe,Ca,K, Mg), trace needs (vitamins), an energy source, buffering capacity

Question 80

Differentiate between complex and defined media

Answer 80

1) Complex: major constituents are organic plant or animal components (ex: TSB); exact nutritional amounts are unknown. Grow many types of bacteria. 2) Defined: synthetic media that contain precisely measured components. Generally grow a narrow range of bacteria.

Question 81

How does autoclaving work?

Answer 81

High pressure saturated steam at 121c works because moist heat kills: 1) Increases motion of particles 2) Cleavage of H bonds between proteins 3) H2O molecules in steam are highly energized. 4) More penetration into liquids/ surfaces

Question 82

Name 3 reasons why aseptic technique is important

Answer 82

1) Safety (of you) 2) Sterility (of substances) 3) Purity (of cultures)

Question 83

Name and describe 3 ways in which we isolated cultures

Answer 83

1) Streak plate isolation: streak for isolated colonies 2) Pour plate isolation: dilute bacterial samples multiple times prior to pouring into molten agar. 3) Spread plate isolation: dilute bacterial samples multiple times prior to streaking a plate.

Question 84

1) What test was created due to a problem with water contamination? When is it positive? 2) How are the results calculated?

Answer 84

1) Phenol red lactose fermentation. Yellow tube = positive. 2) MPN (most probable number) (number of positive tubes + chart).

Question 85

How do you calculate the resulting dilution found on a plate?

Answer 85

TDF of last tube x amount transferred to plate

Question 86

How do you calculate the number of bacteria per ml in each tube of a dilution? (if you're given the original culture's cell count)

Answer 86

Last tube's number of cells/ml x SDF

Question 87

How do you find the dilution used at each step of a dilution scheme? (the ratio)

Answer 87

Amount transferred: Amount already in tube + amount transferred

Question 88

1) How else can 1:100 be written? 2) How else can 1:10 be written? 3) How else can 1:2 be written?

Answer 88

1) 10^-2 2) 10^-1 3) 5 x 10^-1

Question 89

1) How do you calculate number of cells? 2) How do you calculate number of colonies?

Answer 89

1) #colonies / TDF 2) #cells per mL x TDF

Question 90

1) What 2 types of bacteria are acid-fast stains best for? 2) Why are acid-fast stains used on these bacteria?

Answer 90

1) Genus Mycobacterium or Norcardia 2) These cells have a waxy component in their cell walls composed of mycolic acid, which makes them resistant to penetration by aqueous- based stains

Question 91

1) What are the 2 reasons for heat-fixing and air-drying bacteria? 2) What two things does a gram stain tell you?

Answer 91

1) Kills the bacteria and makes sure they adhere to the slide 2) Shape and Gram + or -

Question 92

Name and describe the 4 stages of differential stains

Answer 92

1) Primary stain: initial staining of all cells 2) Mordant: chemical reaction that anchors primary stain to one cell type 3) Decolorizer: chemical reaction that selectively removes primary stain 4) Counter stain: stains the cells that lost the primary stain