Intro to Microbiology Lab Exam 2

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Final Exam

Primary Isolation Media Purpose

-can help differentiate between types of bacteria
-different types of media can be used to encourage growth and to suppress growth

Types of Media

enriched, selective, differential

Enriched Media

contains one or more additives to promote bacterial growth

Selective Media

contains one or more additives to suppress bacterial growth

Differential Media

contains an indicator to distinguish between different types of bacteria

Blood Agar Plate

-(BAP)
-Enriched: with 5% sheep RBC
-Differential: based on hemolysis

Types of Hemolysis

Beta: complete
Alpha: partial
Gamma: none

Mannitol Salt Agar Plate

-(MSA)
-Selective: for Micrococcus and Staphylococcus (salt tolerance 7.5% NaCL)
-Differential: for mannitol fermentation (phenol red indicator)
*phenol red indicator turns yellow under acidic conditions

MacConkey Agar Plate

-(MAC)
-Selective: for Gram negative organisms (crystal violet, bile)
-Differential: for lactose fermentation (neutral red indicator)
*neutral red indicator turns pink under acidic conditions

Eosin Methylene Blue Agar Plate

-(EMB)
-Selective: for Gram negative organisms (methylene blue, EosinY)
-Differential: for lactose fermentation
*indicator causes bacteria to have a greenish-gold metallic sheen under acidic conditions

Phenylethanol Agar Plate

-(PEA)
-Selective: for Gram positive organisms (phenylethanol)

Lysozyme

An enzyme that degrades the peptidoglycan layer in the cell wall. This degradation caused the cell wall to weaken, and the cell lyses (bursts).
-Found in tears, saliva, mucous

Purpose of Lysozyme

-1st line of defense for the body
-Non-specific immune mechanism

Purpose of Biochemical Testing

-Used to identify bacteria
-Enzymes are responsible for biochemical reactions
-Biochemical tests determine what enzymes are being produced by the bacteria, which helps to establish the bacteria's identity

Simple Carbohydrate Fermentation

-sugars (carbs) are fermented by bacteria for energy
-the end products of this fermentation are acids, gasses, and/or alcohols
-media: nutrient broth, carbohydrate, pH indicator, Durham tube
-pH indicator reacts with the acid products of fermentation and will indicate if fermentation has occured
-common pH indicators: Phenol Red, bromcresol purple (both turn yellow under acidic conditions
-Durham tube is used to detect the formation of gas products from fermentation

Hydrolysis of Starch

-hydrolysis=to split molecule apart
-starch is a complex carb (polysaccharide)
-some microorganisms can hydrolyze (split apart) complex carbs into simple carbs utilizing the enzyme amylase
-when starch combines with iodine a purple/black color is formed
-if starch is broken down, the iodine cannot combine with it, and no purple/black color is formed

Production of Hydrogen Sulfide

-gas that is produced when an amino acid that contains sulfur is metabolized by a microorganism
-produced by the reduction thiosulfates, sulfates, or sulfites by microorganism
-presence can be determined by a blackening of the media

Indole

-a by-product that is produced when the bacteria hydrolyze (split apart) tryptophan utilizing the enzyme tryptophanase
-pyruvic acid and ammonia are used by the bacteria, indole is left over
-presence of indole can be determined by adding a reagant called Kovac's after incubation

Motility

-stab innoculation of this media will aid in determining motility
-distinct line of growth at the stab line-non-motile
-diffused growth through the media-notile

Activity of Urease

-urea can be hydrolyzed by some microorganisms to CO2 (carbon monoxide) and NH3 (ammonia)
-NH3 is detected by a pH indicator=phenol red
-NH3=alkaline (basic) turns the phenol red hot pink

Activity of gelatinase

-some microorganisms can hydrolize gelatin utilizing the enzyme gelatinase
-gelatin is semisolid at room temp and liquid at 37C
-refrigeration after incubation will determine if gelatin was hydrolyzed
-liquid media=hydrolyzation of gelatin
-solid media=no hydrolyzation of gelatin

Activity of Phenylalanine

-Phenylalanase removes an amino group from phenylalanine.
-Ammonia is used by the bacteria, and phenylpyruvic acid is left over
-presence of phenylpyruvic acid can be determined by the addition of a reagent called ferric chloride after incubation

Activity of Catalase

-hydrogen peroxide is toxic to cells. catalase is produced by aerobic and facultative bacteria to break down the hydrogen peroxide
-the exceptions to this are the bacteria in the genera of Streptococcus and Lactobacillus

Staphylococci

-gram positive
-facultative anaerobes
-grow in clusters
-normal flora on skin, mucous membranes and throats
-differentiation between staph species is done through appearance on BAP, coagulase activity, mannitol fermentation, and S or R novobiocin
-75% of all staphylococcal infections are from S. aureus, S. epidermidis, and S. saprophyticus

BAP

-S. aureus: colonies appear yellow, beta hemolytic
-S. epidermidis: colonies appear white, gamma hemolytic
-S. saprophyticus: colonies appear white or yellow, gamma hemolytic

Mannitol Fermentation

-S. aureus: +
-S. epidermidis: -
-S. saprophyticus: +/-

Novobiocin

-S. aureus: S
-S. epidermidis: S
-S. saprophyticus: R

Coagulase

-Enzyme that reacts with plasma to form a clot around the bacteria that is producing the coagulase.
-Protects the bacteria from the host's defenses
-S. aureus: +
-S. epidermidis: -
-S. saprophyticus: -

Pathogenesis: Staphylococcus aureus

-primary staph infection
-can cause: skin infections (boils, pimples, impetigo), toxic shock syndrome, food poisoning
-30% of population is a S. aureus carrier-normal flora

Pathogensis: Staphylococcus epidermidis

-normal flora of skin, mucous membranes
-usually not pathogenic
-can become pathogenic if it enters the body through abnormal sites

Pathogenesis: Staphylococcus saprophyticus

-not a part of normal flora
-can cause: UTI's in females

Enteric bacilli

-gram negative
-facultative anaerobe
-inhabit the intestinal tract

Enteric pathogens

-enter the body when contaminated food and water is ingested and causes illness
-of 97 species, 25 are pathogenic
-main enteric pathogens: salmonella, shigella, yersinia, escherichia (enteropathogenic)

Opportunistic pathogens

usually cause infection outside the intestines when the host's condition is compromised
-infections caused by members of the Enterobacteriaceae family that are not one of the main enteric pathogens are opportunistic

E. coli

-normally in the large intestine
-usually an opportunistic pathogen, but general pathogenic strains do exist
-can cause: traveler's diarrhea, hemorrhagic colitis, kidney failure

Salmonella and Shigella

-isolation of either of these implies disease or that the host is a carrier
-can cause: bacterial diarrhea, food poisoning

Blood agar plates (BAP) are enriches with 5% sheep red blood cells and are __________ for hemolysis. The three types of hemolysis are ___ which is complete, ______ which is partial, and ________ which is no hemolysis

differential, beta, alpha, gamma

Eosin Methylene Blue agar plates (EMB) are selective for ________ and are differential for _________. The addition of methylene blue and eosin Y is what makes this media selective.

gram negative organisms, lactose fermentation

Mannitol Salt agar plates (MSA) are selective for ______ and are differential for mannitol fermentation. The addition of ______ to the agar is what makes this media selective.

Micrococcus and staphyloccus, 7.5% NaCl

Phenylethanol agar plates (PEA) are ________ for gram ________ organisms

selective, positive

Lysozyme works against bacteria by degrading the _________ layer in the cell wall. This enzyme can be found in body fluids like ____________

peptidoglycan, saliva, tears, mucous membranes

True or False: A mannitol fermenter appears yellow on Mannitol Salt agar (MSA).

True

Carbohydrate fermentation ______________
a. May produce gas
b. produces acids that lower the pH of the media
c. produces acids that raise the pH of the media
d. both a and b

D. both a and b

________ is an enzyme that breaks down hydrogen peroxide.
a. Hydrolase peroxide
b. Amylase
c. peroxidase
d. catalase

d. catalase

the enzyme urease convers urea into ammonia (NH3) and __________

carbon dioxide (CO2)

What color will a lactose fermenter appear on MacConkey agar (MAC)?
a. pink
b. yellow
c. green metallic
d. red

a. pink

A durham tube is used to detect ________ produced by carbohydrate fermentation.

gas

SIM media detects the production of hydrogen sulfide, indole production, and motility. A positive indole test indicates the enzyme ________ was produced by the bacterial strain.
a. phenylalanase
b. tryptophanase
c. indolase
d. urease

b. tryptophanase

How does beta hymolysis appear on blood agar?
a. A clear zone surrounding the colony
b. no zone surrounding the colony
c. a green/brown zone surrounding the colony
d. a black zone surrounding the colony

a. a clear zone surrounding the colony

The enzyme phenylalanase converts phylalaine into ________.
a. glucose
b. amino acids
c. ammonia and co2
d. ammonia and phenylpyruvic acid

d. ammonia and phenylpyruvic acid

To test for indole production in a SIM tube, you would need to ____________
a. add ferric chloride and look for a yellow color
b. look for a black color in the tube
c. add kovac's reagent and check for a red/pink color
d. add iodine and look for a blue/black color

c. add Kovac's reagent and check for a red/pink color

Name one of the three species of Staphylococci that are responsible for 75% of all Staphylococcal infections.

Staphylococcus aureus

A positive test for gelatinase would result in ___________
a. a yellow color
b. a liquid test tube after refrigeration
c. a solid test tube after refrigeration
d. a pink color

b. a liquid test tube after refrigeration

which of the following organisms produces the enzyme coagulase?
a. staphylococcus aureus
b. staphylococcus epidermidis
c. staphylococcus saprophyticus
d. escherichia coli

a. staphylococcus aureus

True or False: to test for starch production, Kovac's reagent was added to the starch agar after incubation.

False

Which of the following organisms are resistant to antibiotic Novobiocin?
a. Staphylococcus aureus
b. Staphylococcus epidermidis
c. Staphylococcus saprophyticus
d. Escherichia coli

c. Staphylococcus saprophyticus

The reagent used to test for phenylalanine deamination is _________; the reagent turns _____________ as a postive result.
a. phenol red, yellow
b. phenol red, red/pink
c. ferric chloride, yellow
d. ferric chloride, green

d. ferric chloride, green

Which of the following organisms is part of the normal flora of the intestinal tract?
a. Staphylococcus aureus
b. Staphylococcus epidermidis
c. Staphylococcus saprophyticus
d. Escherichia coli

d. escherichia coli

True or False: Salmonella species can cause food poisoning.

True

Name one disease that staphylococcus aureus can cause.

Kidney failure, boils, pimples

Which of the following would be a positive result for the production of the enzyme coagulase?
a. bubbling
b. no bubbling
c. the formation of a clot in the test tube
d. no formation of a clot in the test tube

c. the formation of a clot in the test tube

True or False: the only member in group A Streptococcus in Streptococcus agalactiae.

False

Which of the following is NOT a part of the definition for a coliform?
a. facultative anaerobes
b. ferments lactose to acid and gas
c. gram positive
d. non-sporulating

c. gram positive

the most pathogenic streptococci are in group _________ and display ___________ hemolysis.
a. A, alpha
b. A, beta
c. B, alpha
d. C beta

B. A, beta

True or False: the directogen test (rapid Group A strep test) is based on antigen/antibody reactions.

True

Which of the following organisms is a member of group C Streptococcus?
a. Streptococcus pyogenes
b. Streptococcus equi
c. Enterococcus faecalis
d. Streptococcus agalactiae

B. Streptococcus equi

What is the term that is used if water is coliform free and drinkable?

potable

Which of the following organisms is used as an indicator of fecal contamination of water?
a. S. pyogenese
b. salmonella
c. shigella
d. E. coli

D. E. coli

The citrate test is a biochemical test that determines which of the following traits about the organism?
a. if it is gram positive or gram negative
b. if it can ferment citrate
c. if it can use sodium citrate as a sole carbon source
d. if it can perform aerobic respiration

c. if it can use sodium citrate as a sole carbon source

Name one disease that Streptococcus pyogenese can cause.

Scarlet Fever

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