Micro Lab Exam 2

The types of Media are

Enriched, Selective, and Differential

Enriched

contains one or more additives to promote

Selective

contains one or more additives to suppress

Differential

contains an indicator to distinguish between different types of bacteria

What is the selective for Phenylethanol Agar Plate (PEA)

for Gram-positive organisms (phenylethanol)

What is the selective for Mannitol Salt Agar Plate (MSA)

for salt tolerance (Micrococcus and Staphylococcus) (salt tolerance (7.5% NaCl))

What is the Differential for Mannitol Salt Agar Plate (MSA)

for mannitol fermentation – (+) = yellow halo in agar (phenol red indicator)

What is the selective for MacConkey Agar Plate (MAC)

for Gram-negative organisms (crystal violet, bile)

What is the Differential for MacConkey Agar Plate (MAC)

for lactose fermentation – (+) = pink bacteria
(neutral red indicator)

What is the selective for Eosin Methylene Blue Agar Plate (EMB)

for Gram-negative organisms (methylene blue, EosinY)

What is the Differential for Eosin Methylene Blue Agar Plate (EMB)

for lactose fermentation – (+) = shiny green bacteria (methylene blue, EosinY)

What is the Enriched for Blood Agar Plate (BAP)

with 5% sheep RBC

What is the Differential for Blood Agar Plate (BAP)

based on hemolysis (destruction of RBCs)

Types of Hemolysis

β: ?
α: ?
γ: ?

β: complete
α: partial
γ: none

What degrees and how long should you incubate all plates

all plates at 37˚C for 24-48hrs

Phenylethanol Agar (PEA) Serratia marcescens has

no growth (gram negative)

Phenylethanol Agar (PEA) Staphylococcus
epidermidis has

Growth (gram positive)

Mannitol Salt Agar (MSA) Escherichia
coli has

no growth (gram negative)

Mannitol Salt Agar (MSA) Staphylococcus
aureus has

Growth (gram positive)

Yellow halo = mannitol fermenter

Mannitol Salt Agar (MSA) Staphylococcus
epidermidis has

Growth (gram positive)

No yellow halo = not a mannitol fermenter

MacConkey Agar (MAC) Escherichia
coli has

Growth = gram positive
Pink with bile precipitate haze = lactose
fermentation

MacConkey Agar (MAC) Serratia marcescens has

Growth = gram positive
No pink or bile precipitate haze = not a lactose
fermenter

MacConkey Agar (MAC) Staphylococcus epidermidis has

No growth = gram negative

Eosin Methylene Blue Agar (EMB) has Escherichia coli

Growth = gram positive
Green metallic sheen = lactose fermentation

Eosin Methylene Blue Agar (EMB) has Serratia
marcescens

Growth = gram positive
No green metallic sheen = not a lactose fermenter

Eosin Methylene Blue Agar (EMB) has Staphylococcus epidermidis

No growth = gram negative

Blood Agar (BAP) Staphylococcus
aureus has

growth = gram positive

clear halo = β (complete) hemolysis

Blood Agar (BAP) Staphylococcus epidermidis has

growth = gram positive

Red agar/no change = γ (no) hemolysis

Blood Agar (BAP) Streptococcus
viridans has

growth = gram positive

Brownish green = α (partial)hemolysis

What do Biochemical tests do

helps us establish the bacteria’s identity

Phenol Red Broth (PR) tests detect?

The fermentation of sugars

what is the level of phenol Red Broth (PR) pH indicator that is yellow

below pH 6.8 (positive)

Durham tube is used to detect

the formation of gas products

Catalase is produced by what to break down what

Catalase is produced by aerobic and facultative bacteria to break down the hydrogen peroxide

Oxidase Test detects

Detects cytochrome c oxidase

During the Oxidase Procedure positive results will turn the indicator pad what color

dark purple

Citrate Test determine

if sodium citrate is used as a sole carbon source

the pH indicator Bromthymol Blue

Blue – pH 7.6 (positive)

Organisms that can remove the amine group from the amino acid phenylalanine do this by producing an enzyme called

phenylalanase

After incubation, what reagent is added to the agar slant

Ferric chloride

During the phenylalanine Deaminase Test what color forms and is it positive or negative?

Formation of a green color = positive

Starch Hydrolysis Test determines

if the microbe of interest produces the enzyme
amylase

if starch is broken down in a Starch Hydrolysis Test what happens

the iodine cannot combine with it, and no
purple/black color will be formed = positive for amylase production

Urea can be hydrolyzed using what enzyme

urease

Urea Hydrolysis Test pH level

pink – above pH7.4 (positive)

What does SIM stand for?

Hydrogen Sulfide, Indole, and Motility Test

(H2S)

Hydrogen Sulfide

SIM Medium

Positive = blackening of the media

Indole

by-product that is produced when the bacteria
hydrolyze tryptophan utilizing the enzyme tryptophanase

Presence of indole can be determined
by adding what reagent

Kovac’s after incubation

Positive = pink

Motility

Diffuse growth through the media (hazy) = positive for motility

Phenol red:

yellow – below pH6.8 (positive)

Phenol Red Broth Durham tube:

bubble = positive for gas

Catalase Test Bubbles

positive

Catalase Test Oxidase Test Dark Purple

positive

Citrate Test blue

pH7.6 (positive)

What do you have to add one dropper full of Ferric Chloride
to each tube

Phenylalanine Deaminase

Phenylalanine Deaminase Formation of a green color

positive

What type of plate do you have to ass enough Iodine Reagent to cover the plate

Starch Hydrolysis

Starch Hydrolysis Clear zone in agar around growth

positive

Urease Test

pink – above pH7.4 (positive)

What test do you have to add 5 drops of Kovac’s Reagent to

Indole SIM test

SIM Test

Indole: pink Kovac’s reagent =

positive

SIM test

Sulfur reduction: black =

positive

SIM test

Motility: diffuse (hazy) growth away from stab line =

positive

The Staphylococci

Gram positive

Grow in clusters

Staphylococcus aureus

30% of population is a S. aureus carrier – normal flora

Staphylococcus epidermidis

Normal flora of everyone’s skin

The Enterobacteriaceae

Gram-negative

ADD PICURE SLIDE 11 chapt 11 the Enterobacteriaceae grow as

single bacilli

Pathogenesis

major source of food poisoning and water contamination

Shigella

Cause of food poisoning and water contamination

Pathogenesis cont. Escherichia

Most common cause of water contamination

To ensure water is safe for consumption…

it is screened for E. coli, which is referred to in this instance as a coliform.

Unknown Project includes

two different organisms

MSA isolates

Staphylococcus organism

MAC isolates

Enterobacteriaceae organism

Members of the Enterobacteriaceae family,
such as coliforms (E. coli), are

gram-negative

Bottled spring water =

generally potable because it’s regulated

Water straight from a natural spring =

not guaranteed safe unless tested

Mannitol fermentation

MSA - Staphylococci

Lactose fermentation

MAC - Enterobacteriaceae

MSA – Staphylococci Tests
BAP/Novobiocin Susceptibility Test

what do you use in BAP/Novobiocin Susceptibility Test

1 Novobiocin antibiotic disk

MSA – Staphylococci Tests

Coagulase

MAC – Enterobacteriaceae Tests

Citrate Utilization

With Citrate Utilization what do you have to do

Cap tube loosely to ensure bacteria have enough oxygen

MAC – Enterobacteriaceae Tests

SIM Test

The Streptococci

Gram positive, chains of catalase negative cocci

Streptococci species are divided into

groups

Epidemiology

the study of the causes, occurrence, transmission, distribution, and prevention of disease in the population

Common modes of transmission:

Inhalation, Direct skin contact, Ingestion, etc

Index Case:

the first person that is infected with a disease

Prevalence Rate:

the total number of cases of a disease occurring at a specific time point in a defined population

Calculation of Prevalence Rate formula

(Total Number of Cases)/(Number of People in the population) = (make sure to  convert to percent)

This is the initiation of the epidemic –
the index case will be infected” if they have the one candy that is covered in a powder that glows under ____ (fomite)

blacklight

How does disease spreads

through shaking hands (direct skin contact)

Expected Results Mannitol Fermentation yellow =

positive

Expected Results BAP Clearing of blood agar =

complete (β) hemolysis

Expected Results BAP Brownish green blood agar =

partial (α) hemolysis