M

E
D WEEK 7
I
C
A
L

L
A
B
O
R
A
T
O
R
Y
CANELLIE P. CANLAS, RMT, MSMT
S
C References:
I
E
N Textbook of Diagnostic Microbiology 5th edition by Mahon
C Diagnostic Microbiology 12th edition by Bailey & Scott
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E STAPHYLOCOCCI
D
I
C - Catalase positive
A - Gram positive cocci
L
- spherical cells 0.5 - 1.5 um appear in singly, in pairs, in clusters
L - Staphylococcus, is derived from the Greek term staphle, meaning “bunches of grapes.”
A
B
- non-motile
O - non spore forming
R - aerobic & facultatively anaerobic except : S. saccharolyticus (oblogate anaerobe)
A
T
O Rare strains of Staphylocci are Fastidious, they require: These so called small
R
Y 1. colony variants , forming
2. colonies about 1/10 the
S
C
3. size after 48 hours
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E MICROCOCCUS
D
I
C
A
- Catalase positive OTHER GRAM-POSITIVE COCCI
L - Coagulase negative OCCASSIONALLY RECOVERED
L - Gram positive cocci found in the WITH STAPHYLOCOCCI:
A
B
O
environment, member of skin 1. Rothia mucilaginos
R
A
microbiota 2. Aerococcus
T
O
- Often recovered with 3. Alloiococcus otitis
R
Y
Staphylococci recovered from middle ear fluid

S
- Some of them produce Yellow
C
I
Pigment
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E MICROCOCCUS
D
I
C
A
- They will alsogrow well in broth-blood culture systems and
L
common nutrient broths, such as thioglycollate and brain-
L
A
heart infusion.
B
O
- Gram stains should be performed on young cultures,
R
A
because very old cells may lose their ability to retain crystal
T
O
violet and may appear gram-variable or gram-negative.
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E STAPHYLOCOCCI
D
I
C -
A
L

L
A (Remel, Lenexa, KS)
B
O
R
A
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E STAPHYLOCOCCI
D
I
C - Staphylococcal species are differentiated by COAGULASE TEST
A
L (+) clot formation STAPHYLOCOAGULASE

L
A
B
STAPHYLOCOAGULASE-PRODUCING OCCASIONALLY MISTAKEN AS
O (COAGULASE-POSITIVE) COAGULASE-POSITIVE
R
A STAPHYLOCOCCI: STAPHYLOCOCCI:
T
O
1. S. lugdunensis
R 1. S. hyicus 2. S. schleiferi CLUMPING FACTOR
Y
2. S. intermedius
S
C 3. S. lugdunensis
I
E
4. S. aureus
N 5. S. delphini
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E STAPHYLOCOCCI
D
I
C
A
COAGULASE NEGATIVE
L
STAPHYLOCOCCI (CoNS)
L
A
B
O
R
1. S. saprophyticus
A
T 2. S. haemolyticus
O
R 3. S. epidermidis
Y

S
4. S. lugdunensis
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C - most clinically significant species of staphylococci
A ENTEROTOXINS together with
L - important cause of nosocomial infections TSST-1 are superantigens that
have ability to react with T cells
L
and activate aggresive immune
A VIRULENCE FACTORS
B response
O 1. ENTEROTOXIN stable at 100oC for 30 mins
R
A
- heat-stable exotoxins causes diarrhea and vomiting
T - 9 serologically distinct identified to fall into enterotoxin groups: A,E,G,J
O
R - ENTEROTOXINS A , B , D causes: Staphylococcal food poisoning
Y
- ENTEROTOXINS B & C sometimes G & I associated by:
S
C
Toxic Shock Syndrome
I
E
N - ENTEROTOXIN B: staphylococcal pseudomembranous
C enterocolitis
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
2. Toxic Shock Syndrome Toxin-1 Previously Enterotoxin F
A
L
- causes nearly all cases of
L
A
menstruating-associated TSS
B
O
- is a superantigen stimulating
R
A
T-cell proliferation and the
T
O
subsequent production of a
R
Y
large amount of cytokines that
S
are responsible for the
C
I
symptoms.
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C 3. EXFOLIATIVE TOXIN Epidermolytic Toxin
A
L - It causes the epidermal layer of the skin to
L
slough off
A - causes: Staphylococcal SSS or Ritter's Disease
B
O - implicated also in Bullous impetigo
R
A
T
O
4. CYTOLYTIC TOXINS
R - other extracellular proteins that affect red blood cells and leukocytes
Y
- HEMOLYSINS and LEUKOCIDINS
S
C
- 4 hemolysins produced by S. aureus : alpha, beta, gamma, delta
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
L a. Alpha - can damage platelets and macrophage
L b. Beta - acts on sphingomyelin in the plasma membrane
A
B of erythrocytes and is also called the “hotcold” lysin.
O
R c. Delta - found in a higher percentage of S. aureus stains
A
T and some CoNS, is considered less toxic to cells
O
R d. Gamma - associated with Panton-Valentine leukocidin
Y
(PVL) Staphylococcal Leukocidin
S
C exotoxin lethal to PMN that suppress
I
E Phagocytosis
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
5. ENZYMES
L a. Protease
L b. Coagulase
A
B
c. Hyaluronidase - hydrolyzes hyaluronic acid present in the
O intracellular ground substance that makes up connective tissues,
R
A permitting the spread of bacteria during infection.
T
O d. Lipase - act on lipids present on the surface of the skin, particularly
R
Y
fats and oil secreted by the sebaceous glands.
S
C 6. PROTEIN A
I
E
- S. aureus is its ability to bind the Fc portion of immunoglobulin
N G (IgG).
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C INFECTIONS CAUSED BY S. aureus :
A
L

L
1. SKIN AND WOUND INFECTIONS
A - suppurative
B
O a. folliculitis - mild inflammation of a hair follicle or oil gland; the infected
R
A
area is raised and red
T b. furuncles - which can be an extension of folliculitis, are large, raised,
O fever and chills, systemic spread of
R superficial abscesses bacteria
Y
c. carbuncles - occur when larger, more invasive lesions develop from multiple
S
C
furuncles, which can progress into deeper tissues
I d. bullous impetigo - highly contagious infection that is easily
E
N spread by direct contact, fomites, or autoinoculation
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
2. SCALDED SKIN SYNDROME
L - bullous exfoliative dermatitis that occurs
L primarily in newborns and previously healthy
A
B young children.
O
R
- Cases of SSS in adults occur most commonly in
A patients with chronic renal failure
T
O and in patients with compromised immune systems
R
Y
3. TOXIC SHOCK SYNDROME Todd, 1978
S
C - rare but potentially fatal, multisystem disease
I
E - fever, chills, vomiting, diarrhea, muscle aches, and
N
C
rash, which can quickly progress to hypotension
E and shock
Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
4. TOXIC EPIDERMAL NECROLYSIS
L
- commonly drug induced
L
A
- some cases have been linked to
B
O
infections and vaccines
R
A
- very similar initial presentation to
T
O
that of SSS, treatments differ
R
Y
- can be resolved by the
S
administration of steroids early in
C
I
the initial stages of presentation,
E
N
whereas steroids aggravate SSS
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
5. FOOD POISONING
L
- most commonly A (78%), D (38%), and B (10%)
L
A
- associated with gastrointestinal disturbances
B
O
- The source of contamination is usually an infected food
R
A
handler
T
O
- Foods kept at room temperature are especially susceptible
R
Y
to higher levels of toxin production
S
- The enterotoxins do not cause any detectable odor or
C
I
change in the appearance or taste of the food.
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
6. Staphylococcal bacteremia leading to secondary
L pneumonia and endocarditis
L
A
- observed among intravenous drug users.
B
O
- The organisms gain entrance to the bloodstream via
R
A
contaminated needles or from a focal lesion present on the
T
O
skin or in the respiratory or genitourinary tract
R
Y
- Bacteria may lodge in the diaphysis of the long bones and
S
establish an infection. manifestation: OSTEOMYELITIS
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus aureus
D
I
C
A
-
L

L
A
B
O
R
A
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus epidermidis
D
I
C
A
- hospital acquired
L
- predisposing factors:
L
A
cathetherization
B
O
medical implantation
R
A
immunosuppresive therapy
T
O
- causes: Prosthetic valve
R
Y
endocarditis
S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus saprophyticus
D
I
C
A
- adheres more effectively to the epithelial cells lining
L
the urogenital tract
L
A
- associated with UTIs in young women.
B
O
- When present in urine cultures, S. saprophyticus may be
R
A
found in low numbers (<10,000 colony-forming units/mL)
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Staphylococcus lugdunensis
D
I
C
A
- although it can give a positive clumping factor test result,
L
but it has a negative tube coagulase reaction.
L
A
- This organism can be more virulent and can clinically mimic
B
O
S. aureus infections
R
A
- has been known to contain the gene mecA that encodes
T
O
oxacillin resistance.
R
Y
- It is an important pathogen in infective endocarditis,
S
septicemia, meningitis, skin and soft tissue infections, UTIs,
C
I
and septic shock.
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E OTHER COAGULASE NEGATIVE STAPHYLOCOCCI
D
I
C
A 1. S. warneri
L
2. S. capitis
L
A
B
3. S. simulans
O
R
4. S. hominis - some strains are Vancomycin resistant
A
T - wounds, bacteremia, endocarditis, and
O
R
Y
UTIs
S
6. S. schleiferi
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C
A
-
L

L
A
B
O
R
A
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C
A
1. Sheep's Blood Agar
L
2. Mannitol Salt Agar
S. aureus
L
A
3. CHROMagar (Becton Dickinson, Franklin Lakes, NJ)
B
O
4. Colistin-Nalidixic Acid Agar for heavily contaminated
R
5. Phenylethyl Alcohol Agar specimens
A
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C
A
- Staphylococci ferment glucose, whereas micrococci fail to
L
produce acid under anaerobic conditions.
L
A
- Staphylococci that fail to grow or produce acid
B
O
anaerobically, such as:
R
A
S. saprophyticus
T
O
S. hominis
R
Y
S. auricularis
S
S. cohnii
C
I
S. xylosus
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C COAGULASE TEST B. TUBE COAGULASE (Confirmatory) -
A
L enzyme coagulase produced by detects FREE coagulase or
L S.aureus binds plasma fibrinogen and STAPHYLOCOAGULASE
A
B
activate cascade of reaction that **Isolates suspected of being S.
O causes plasma to clot. aureus, but failing to produce bound
R
A coagulase
T
O
A. SLIDE COAGULASE (Presumptive) -
R detects BOUND coagulase
Y
**Approximately 10% to 15% of
S
C strains may give a negative slide
I
E
coagulase test.
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C
A
PYRROLIDONYL ARYLAMIDASE
L

L
A
- differentiate S. aureus (negative)
B
O
from S. lugdunensis, S. intermedius,
R
A
and S. schleiferi (positive)
T
O
R PRINCIPLE: (L-pyrrolidonyl-β-
Y
naphthylamide; PYR), is
S
C hydrolyzed to L-pyrrolidone and β-
I
E
naphthylamine, which combines with p-
N dimethylaminocinnamaldehyde to form a
C
E red compound
Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C animal pathogen, and most human
A VOGUE-PROSKAUER TEST infections
L
are associated with animal bites.
L
A differentiating S. aureus (positive) from S. intermedius
B
O (negative)
R
A Also positive:
T
O S. lugdunensis
R
Y S. haemolyticus
S S. schleiferi
C
I
E
N (+) formation of acetoin from glucose or pyruvate
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E Isolation and Identification
D
I
C
A
- Used to differentiate Coagulase-
L
Negative Staphylococci
L
A
5ug
B
O
R
A
Positive: ZOI (CoNS)
T
O
Negative: no ZOI (S. saprophyticus)
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)
M
E
D
I
C
A
L

L
A
B
O
R
A
T
O
R
Y

S
C
I
E
N
C
E

Our Lady of Fatima University - Valenzuela

BACTERIOLOGY (LECTURE)