ex. 14: skin cultures and importance of selective and differential media for isolating gram-positive...
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Ex. 14: Skin Cultures and Importance of Selective and
Differential Media for Isolating Gram-Positive Cocci
Objectives??
Table 14-2: Bacteria Commonly Found on Human Skin
Bacterium Frequency of presence
Propionibacterium spp. ++
Staphylococcus epidermidis ++
Staphylococcus aureus +
Micrococcus spp. +
Corynebacterium spp. ++
Streptococcus pyogenes +/++ = nearly always present; + = commonly present; +/ = rare
MSA: Selective and Differential Medium for Gram + Cocci
Composition: meat extract, casein, peptones; 7.5% NaCl, mannitol as only fermentable carbohydrate, phenol red indicator.
Table 14-1: Mannitol Salt Agar (MSA) Results and Interpretation
Result Interpretation Presumptive ID
Poor growth or no growth (P)
Organism is inhibited by NaCl Not Staphylococcus
Good growth (G) Organism is not inhibited by NaCl
Possible Staphylococcus
Medium turns yellow (Y)
Organism produces acid (A) from mannitol fermentation
Possible pathogenic Staphylococcus aureus
Medium remains red (R)
Organism does not ferment mannitol. No reaction (NR)
Nonpathogenic Staphylococcus
Staphylococcus saprophyticus on MSA plate
Materials needed per team of two students: Three MSA plates Two sterile cotton swab Two tubes with sterile saline
Materials needed per table: Slant cultures of the following bacterial species: 1.Micrococcus luteus2.Staphylococcus aureus3.Staphylococcus
epidermidisWork as a team of two for the control organisms, then perform the skin swab
individually!
Day 1
Control Organisms Skin Inoculum
Materials needed per table: Gram staining reagents Hydrogen peroxide (3%) , glass slides Rabbit plasma vials for coagulase test
Day 2
Catalase Test
H2O2 is by-product of aerobic respiration. Lethal to the cells.
Most aerobic organisms produce catalase for protection
Catalase test differentiates among morphologically similar Gram+ cocci.
Table 14-2: Catalase Test Results and Interpretation
Result Interpretation Symbol
Bubbles Catalase is present +
No bubbles Catalase is absent
Coagulase Test
Coagulase activates fibrin and leads to clot formation Protective barrier around bacterial cells Virulence factor
Test determines presence of coagulase enzyme
Used for differentiating S. aureus from other Staphylococci.
Tube Coagulase Test
check no later than 24 hours after inoculation!
Tests for bound of free coagulase
Slide Coagulase Test
Tests for bound coagulase
Positive if agglutination within 1 to 2 minutes
Table 14-5: Results of MSA Plates
ColonyColony
description Pigment Mannitol
fermentation Gram stainCatalase reaction
S. aureus
S. epidermidis
M. luteus
Skin isolate #1
Skin isolate #2
Skin isolate #3