Lab Two, Part 2: Blood and MSA, pg 429

Lab Two, Part 1: Coagulase Test, pg 429

Exp 29: Catalase Test

Physical Factors

Exp 15: Temperature

Exp 16: pH

Exp 17/18: O2 Requirements

Week 10WWeek 10W

Exp 62: ID of Human Staph. and Strep. Pathogens

Lab Two, pg 196add H2O2

Last week: New This Week: Gram (+) Bacteria ID Review Gram + and - tests

New This Week: Gram (+) Bacteria ID Review Gram + and - tests

Gram staining practice

Syllabus Corrections

Pg 3– Week 12: Assign. #7 due– Week 13: Assign. #6 due (not # 8)

Pg 8– Week 11: Prepare Assign. #7 (not #6)– Week 12: Quiz 9 covers definitions from

Assign. #7 (not #6)

: Prepare Assign. #6 (not #8)

Exp 62: ID of Human Staph. And Strep. Pathogens: Blood and MSA Review

An MSA plate with Micrococcus sp. (1), Staphylococcus epidermis (2) and S. aureus (3) colonies.

  Red blood cells on an agar plate are used to diagnose infection. On the left is a positive Staphylococcus infection, on the right a positive Streptococcus pyogenes culture.

Exp 62: ID of Human Staph. And Strep. Pathogens:

Coagulase Test Coagulase reacts with the fibrinogen in plasma,

and precipitates into fibrin cells clump together , look “lumpy”.

Test useful for differentiating Staphylococcus aureus from other Gram positive, catalase + cocci.

Any degree of clotting, from a loose clot suspended in the plasma to a solid, immovable clot is a positive result. Plasma which liquid remained is a negative result.

Exp 62: ID of Human Staph. And Strep. Pathogens:

Coagulase Test

fibrin

Exp. 29: Catalase Test

o Hydrogen peroxide (H2O2) accumulation is toxic to cells. o Presence of bubbling or foaming when adding H2O2 to a culture, indicates that the hydrogen peroxide is being broken down by catalase, with the release of oxygen gas.o Bubbling = pos. catalase o Absence of foaming/bubbling= neg. catalase (catalase is not present in cell)

Results To ID Gram Positive Bacteria

 

Notation

MICROORGANISM

Staph. epidermidis

(SE)

Staph. aureus

(SA)

B. subtilis (BS)

Strep. salivarius (SS)

M. luteus

(ML)

Morphology

shape/ arrangement cocci/ cluster

cocci/ cluster

rods cocci/ chaincocci/ cluster

TSA (cultural characteristics)

color, size, form, margin,

elevation, texture

         

+/- growth        ■

MSAMF or NMF/

color         

Blood Agar (cultural characteristics)

color, size, form, margin,

elevation, texture

         

Blood Hemolysis

У α β         

Catalase Activity

+/- catalase        ■

Coagulase Actvity

+/- coagulase          

Seafloor bacteria on ocean-bottom rocks are more abundant and

diverse than previously thought.

Grand Prismatic Spring, Yellowstone National Park (thermophiles)

Below Freezing Salt Water Pockets in Antarctica

Bacteria Around Nuclear Reactors

Biofilm build-up on tank walls—

guaranteed tocontaminate any fuel

Exp 15: Physical Factors: Temperature

Exp 16: Physical Factors: pH of the Extracellular Environment

Exp 17/18: Physical Factors: Atmospheric Oxygen RequirementsCategory Oxygen Requirement Production of

Catalase

Obligate Aerobe

Requires oxygen and oxygen is not toxic to it. Produces catalase.

Microaerophile Requires oxygen at low levels and atmospheric levels of oxygen are toxic to it. Prefers oxygen

at a concentration lower than 10%.

Produces catalase

sometimes.

Facultative Anaerobe

Uses oxygen preferentially when oxygen is present, but can grow without oxygen (slower growth than with oxygen). Oxygen is not toxic

to it.

Produces catalase.

Obligate anaerobe

Does not use oxygen and oxygen is toxic to it. No catalase production.

Aerotolerant Does not use oxygen, but oxygen is not toxic to it.

Produces catalase.

How is distribution of growth affected by atmospheric oxygen ……..?

TABLES 5 & 6Exp. 17/18: Atmospheric Oxygen RequirementsMedia: 3Thioglycollate broth 3 NUT brothCulture: EC, ML & CSProcedure: (refer to pg. 127)• Label ea. tube w/org & media.• Inoculate each org. in thioglycollate and NUT broth.• Place thioglycollate tubes in anaerobic GasPack chamber.•Incubate all at 37oC for 24 hrs.

Week 10WWeek 10W

GLU SUC LAC

TABLES 1 & 2

Exp 15: Temperature

Media: 12 NUT slant

Culture: SA , ML, BST

Procedure: (refer to pg. 111)

• Label each tube w/ org. and temp.

• Inoculate with loop 4 slants w/each org.

• Incubate 1 slant from each org. at either 4oC, 25oC, 37oC, and 55oC for 24 hrs.

TABLES 3 & 4

Exp 16: pH of the Extracellular Environ.

Media:

2 NUT broth @ pH 2

2 NUT broth @ pH 5

2 NUT broth @ pH 7

2 NUT broth @ pH 9

Culture: EC & SA

Procedure: (refer to pg. 116)•

Label each tube w/ org. and pH.

• Inoculate with ea. org in various pH tubes.

• Incubate at 37oC for 24 hrs.

BST- Bacillus stearothermophilusCS- Clostridium sporogenes

Gram staining practice