Biochemical Tests

Streptococcus pneumoniae

______________: is a gram-positive, encapsulated, lancet-shaped diplococci, most commonly causing otitis media, pneumonia, sinusitis, and meningitis. 

Streptococcus viridans 

_______________: Consist of Strep. mutans and Strep mitis found in the normal flora of the oropharynx commonly that cause dental carries and subacute bacterial endocarditis (Strep. sanguinis).

Streptococcus pyogenes

_______________: is a gram-positive group A cocci which can cause pyogenic infections (pharyngitis, cellulitis, impetigo, erysipelas), toxigenic infections (scarlet fever, necrotizing fasciitis), and immunologic infections (glomerulonephritis and rheumatic fever).

Streptococcus agalactiae

_______________:  Is a gram-positive group B cocci that colonize the vagina and is found mainly in babies. Pregnant women need screening for Group-B Strep (GBS) at 35 to 37 weeks of gestation. 

Enterococci

________________: Is a gram-positive group D cocci found mainly in the colonic flora and can cause biliary tract infections and UTIs. Vancomycin-resistant enterococci (VRE) are an important cause of nosocomial infections.

Bacitracin Test

________________: The test is used for presumptive identification and differentiation of beta-hemolytic group A streptococci (Streptococcus pyogenes– susceptible) from other beta-hemolytic streptococci.

• It is also used to distinguish staphylococci species (resistant) from micrococci (susceptible).

Bacitracin Test (Principle)

The antibiotic bacitracin inhibits the synthesis of bacterial cell walls by interfering the peptidoglycan synthesis of bacteria.

Mostly, blood agar plate is used (Trypticase soy agar + 5% sheep blood).

Bacitracin Test (Medium)

Bacitracin Test (Expected results)

______________:

• Positive: Any zone of inhibition greater than 10 mm; susceptible.

• Negative: No zone of inhibition; resistant

Beta Lactamase Test

________________: To detect the enzyme beta-lactamase, which confers penicillin resistance to various bacterial organisms.

Beta Lactamase Test (Principle)

______________: The chromogenic cephalosporinase test detects β-lactamase by using a chromogenic cephalosporin substrate on a test disk. If the organism produces β-lactamase, it hydrolyzes the β-lactam ring, releasing a visible colored product (deep pink). No color change means β-lactamase is absent.

Beta Lactamase Test (Expected results)

_____________:

• Positive: Yellow to pink-red color change on the area where the culture is applied.

• Negative: No color change.

Beta Lactamase Test (Uses)

Useful applications include detection of:

• N. gonorrhoeae resistance to penicillin

• H. influenzae resistance to ampicillin

• Staphylococcal resistance to penicillin

Bile Esculin Test

______________: This test detects organisms that grow in bile and hydrolyze esculin. Hydrolysis produces esculetin, which reacts with iron to form a black color, indicating a positive result commonly seen in enterococci and some streptococci. 

Bile Esculin Test (Expected results)

_______________:

• Positive: Growth and blackening of the agar slant

• Negative: Growth and no blackening of medium ; No growth

Bile Solubility Test (Principle)

_______________: This test distinguishes S. pneumoniae from other alpha-hemolytic streptococci. Bile salts activate the bacterium’s autolytic enzyme, causing cell lysis and clearing, which indicates a positive result.

Bile Solubility Test (Expected results)

______________:

Positive Result: Suspension clears in tube labelled test and remains turbid in control tube.

Negative Result: Suspension remains turbid.

CAMP Test (Principle)

______________: Used to identify Streptococcus agalactiae (Group B). Its CAMP factor enhances S. aureus beta-hemolysin, producing an arrowhead-shaped zone of hemolysis on blood agar.

CAMP Test (Expected results) 

_______________: 

• Positive: Enhanced hemolysis is indicated by an arrow head-shaped zone of beta-hemolysis at the junction of the two organisms.

• Negative: No enhancement of hemolysis.

CAMP Test (Quality control)

_______________: 

• Positive: Streptococcus agalactiae—enhanced arrowhead hemolysis.

• Negative: Streptococcus pyogenes—beta-hemolysis without enhanced arrowhead formation.

Catalase Test (Principle)

______________: This test detects the enzyme catalase, which breaks down hydrogen peroxide into water and oxygen. A positive result is shown by rapid bubble formation, while no or weak bubbles indicate a negative result.

Catalase Test (Quality control)

_______________: 

• Positive: Staphylococcus aureus

• Negative: Enterococcus faecalis

Coagulase Test (Principle)

_______________: Differentiates S. aureus from other staphylococci. S. aureus produces bound coagulase (clumping factor) and free coagulase, both of which act on fibrinogen to cause clumping or clot formation.

Coagulase Test (Quality control)

________________:    

• Positive: Staphylococcus aureus 

• Negative: Staphylococcus epidermidis 

Hippurate Hydrolysis Test

_______________: Detects hippuricase enzyme that breaks down hippurate into glycine and benzoic acid. Glycine reacts with ninhydrin to form a purple color, used to identify Group B streptococci and some other bacteria.

Hippurate Hydrolysis Test (Expected results)

______________:

• Positive: A positive test is indicated by the appearance of a deep blue/violet color in 30 minutes.

• Negative: Colorless or slightly yellow pink color

Deoxyribonuclease (DNase) Test (Principle)

________________: The DNase test detects bacteria that produce deoxyribonuclease, which breaks down DNA and creates a clear zone on the agar. It’s mainly used to identify Staphylococcus aureus.

Deoxyribonuclease (DNase) Test (Expected results)

________________:

• Positive: Medium is colorless around the test organism.

• Negative: If no degradation of DNA occurs, the medium remains green.

Leucine Amino Peptidase (LAP) Test (Principle)

_________________: Detects leucine aminopeptidase in bacteria. A positive reaction turns the disk red, helping identify catalase-negative, gram-positive cocci like streptococci.

Leucine Amino Peptidase (LAP) Test (Expected results)

_______________: 

• Positive: Development of a red/pink color

• Negative: No change or a slight yellow color