bio318L exam 2: lab#8

What is Staphylococcus?

What is Staphylococcus?

• gram positive cocci (in clumps like grapes)

• all members are highly resistant to drying, extremes in pH, and high temperatures

• increase in community acquired cases, with infections associated with schools, prisons, and shared athletic equipment

S. epidermis

common cause of hospital-acquired infections in immunocompromised patients

S. saprophyticus

an occasional cause of UTIs

S. aureus

common cause of hospital-acquired skin infections

• normal flora of humans: nasal membranes & skin

• opportunistic infections

Latex Agglutination Test

rapid identification for S. aureus

• Protein A is a virulence factor found on the cell wall of only S. aureus

• antibodies in the test solution bind antigens on Protein A, resulting in a positive result of clumping

Latex Agglutination Test Results

positive = S. aureus

• clumping occurs because the antibodies in the test solution bind with the antigens found on Protein A which is located on the cell wall of only S. aureus

negative = not S. aureus

• no clumping occurs due to the absence of Protein A

Catalase Test

key test in differentiating Gram (+) cocci

• substrate is hydrogen peroxide

• distinguishes Staph from Strep

Catalase Test Results

positive = Staph

• bubbling will occur due to the presence of catalase

• the catalase reacts with the hydrogen peroxide, resulting in oxygen to be released as seen by the bubbling

negative = Strep

• no bubbling occurs due to the absence of catalase and thus no release of oxygen

Mannitol Salt Agar

Selective & differential media for Staphylococcus

• contains 7.5% salt to inhibit the growth of most bacteria

• contains phenol-red dye to detect mannitol fermentation

• highly selective for S. aureus and some strains of S. saprophyticus

Mannitol Salt Agar Test Results

positive = S. aureus or S. saprophyticus

• a color change from red to yellow indicates the fermentation of mannitol, which S. aureus and some strains of S. saprophyticus are able to do

negative = S. epidermis

• no color change indicates the inability to ferment mannitol

Blood Agar Plates

Tests for hemolysis ability

• contains Tripticase Soy Agar (TSA) and 5% sheeps blood

• beta, alpha, and gamma

Blood Agar Plates Results

beta = S. aureus

• indicates complete breakdown of red blood cells

• if the plate is held up to a light, you will be able to see through it

partial = none

• indicates partial breakdown of red blood cells

• seen by a green tint around the growth

gamma = S. saprophyticus & S. epidermis

• indicates no breakdown of red blood cells

• if the plate is held up to a light, you will not be able to see through it

Novobiocin Susceptibility

Distinguishes coagulase-negative Staph species

• does not apply to S. aureus

Novobiocin Susceptibility Test Results

positive for coag-negative = S. epidermis

• seen by a zone of inhibition

negative for coag-negative = S. saprophyticus

• seen by no zone of inhibition

What is Streptococcus?

• gram positive cocci (in a chain-like pattern)

• highly environmentally sensitive

• normal human commensal

  • mouth, skin, intestine, and upper respiratory tract

Strep. mutans

most common cause of dental plaque

Strep. pneumoniae

most common cause of bacterial pneumonia

Strep. pyogenes

common cause of throat infections; “Strep throat”

• responsible for pharyngitis and systemic infection like scarlet fever

• able to completely lyse RBC

4 Streptococcus strains identified in lab

• S. pyogenes

• S. agalactiae

• S. equi

• E. faecalis

CAMP Test

Identification for Group B or S. agalactiae

Bacitracin Susceptibility Test

Identification for Group A or S. pyogenes

Bacitracin Susceptibility Test Results

positive = S. pyogenes or Group A

• seen by a zone of inhibition around the disk

negative = not S. pyogenes or Group A

• seen by no zone of inhibition around the disk

SXT Sensitivity Test

Identification for Group C or S. equi

• enzyme inhibitor (blocks folic acid synthesis)

SXT Sensitivity Test Result

positive = S. equi or Group C

• seen by a zone of inhibition around the disk

negative = not S. equi or Group C

• seen by no zone of inhibition around the disk

Optochin Sensitivity Test

Identification for S. pneumoniae

• was not performed in lab

• a zone of inhibition indicates a positive result for S. pneumoniae

Bile Esculin Media

Identification for Group D or E. faecalis

• bile salts inhibit non-Group D species

• esculin hydrolysis forms black precipitate

Bile Esculin Media Results

positive = E. faecalis or Group D

• seen by the formation of black precipitate

• bacteria is able to hydrolyze esculin

negative = not E. faecalis or Group D

• seen by no change in the test tube

• bacteria is unable to hydrolyze esculin