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Microbio Lab 4

Microbio Lab 4

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Published by: api-3743217 on Oct 15, 2008
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03/18/2014

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MICROBIOLOGY LABORATORY 4

USTMED \u201907 Sec C \u2013 AsM; Photos provided by JV.N & MeaM.
Culture of Microorganisms
Different Streaking Methods

Culture Media Used: Eosin Methylene Blue
Organisms Used: Escherichia Coli
Method of Streaking: Simple Streaking

Illustration:

Culture Media Used: Eosin Methylene Blue
Organisms Used: Escherichia Coli
Method of Streaking: 4 Quadrant Method of Streaking

Note: This method of streaking is used for better isolation of
the organism
Illustration:

Culture Media Used: Eosin Methylene Blue
Organisms Used: Escherichia Coli
Method of Streaking: Overlapping method of streaking

Note: This method of streaking is used for sensitivity testing
Illustration:
Demo slides:

Gram stain of pus from an epyema cavity showing long and short chains of Gram-positive streptococci and large numbers of pus cells (stained red) x3,900.

Staphylococcus aureus. Gram stain of culture showing characteristic irregular clusters of gram-positive cocci. There are no spores or capsules.

Gram stain of pus from cervix \u2013 Gonorrhea
Neisseria gonorrhea \u2013 Gram stain
Neisseria gonorrhea \u2013
Stanford\u2019s stain

Clinical specimen: Urethral Discharge
Staining used: Gram Staining
Results: Gram Negative Cocci in Pairs Intracellularly located

Staphylococcus aureus
Gram Positive Cocci in clusters
Catalase Test \u2013 Positive
(best single test for the pathogenecity of Staph. Aureus)
Colonies on plated media:
1. Blood agar plate (BAP) \u2013 Beta hemolytic (complete
hemolysis)
2. Mannitol salt agar (MSA) \u2013 Yellow colonies (due to
mannitol fermentation)

Staphylococcus aureus on mannitol Salt Agar (MSA), differentiating S. aureus from other catalase positive gram posititve cocci. The medium contains 7.5% NaCl which inhibits the growth of many organisms. S. aureus ferments mannitol resulting to acid end product causing the phenol red indicator to change from pink to yellow.

Staphylococcus aureus on Mannitol Salt Agar (MSA)
Showing yellow colonies due to Mannitol fermentation

Mannitol Salt Agar (MSA) differentiates S. aureus from other catalase-positive gram-positive cocci. The medium contains 7.5%NaCl which inhibits the growth of many orgs.

(+) results \u2013 fermentation of mannitol resulting to acid end product causes the phenol red indicator to change from pink to yellow (right half of the plate)

(-) result \u2013 no fermentation, no chage of color (left half)
Colonies of Staph. Aureus on MSA (close-up). Medium
surrounding colonies has turned yellow

Coagulase test confirms identiy as Staphylococcus aureus (coagulase positive) and Staphylococcus epidermidis (coagulase negative):

1. Slide coagulase test \u2013 when a saline suspension of staphylococci is mixed with human or rabbit plasma on a slide, there is immediate clumping with coagulase positive staphylococci.

2. Tube coagulase test \u2013 when diluted plasma is inoculated with S. aureus, jellification occurs w/in 2- 4 hours. This result is due to coagulase converting fibrinogen in the plasma to fibrin and a positive test confirms a positive slide test.

Staphylococcus aureus on Blood Agar Plate (BAP)
Showing beta hemolytic hemolysis; complete hemolysis

Staphylococcus aureus on BAP colonies showing beta or complete type of hemolysis.

Growth of staphylococcus on
BAP
with

characteristic pigmentation (golden-yellow) and \u2018oil paint\u2019 appearance.

Streptococcus pneumoniae
-
gram positive cocci in pairs
-
catalase negative
-
alpha hemolytic/incomplete hemolysis on BAP
-
Sensitive to Optochin Disk Test as seen in Chocolate
Agar Plate (CAP)
Streptococcus viridans
-
gram positive cocci in chain
-
catalase negative
-
alpha hemolytic/incomplete hemolysis on BAP
-
Resistant to OPtochin Disk Test as seen in CAP
Streptococcus pyogenes
-
gram positive cocci in chains
-
catalase negative
-
beta hemolytic/complete hemolysis on BAP
-
resistant to optochin disk
Optochin
sensitivity
of
pneumococci in contrast with
Streptococcus viridanswhich
also
produces

alpha- haemolytic colonies on blood agar.

Streptococcus viridans on BAP
-
produces alpha hemolytic/incomplete hemolysis on
BAP

Growth of a beta-haemolitic stretptococcus on blood agar medium showing large zones of clear haemolysis around small transparent colonies.

Growth of pneumococci on blood agar showing characteristic
\u2018draughtsman\u2019 colonies with concentric surface rings.

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