MICROBIOLOGY

Gram-Positive Cocci (Staphylococci)

Dr. Billones
CHARACTERISTICS OF STAPHYLOCOCCI 4 MOST FREQUENLY ENCOUNTERED SPECIES ASSOCIATED WITH HUMAN DISEASE
A. Morphology: 1. S. aureus – coagulase positive, major pathogen to humans. Causes
 Spherical; approximately 1 µm in diameter diseases like minor skin infection, food poisoning, and toxic shock
 Arranged in grapelike clusters (single cocci, pairs, tetrads, syndrome
and chains may be seen in liquid cultures) 2. S. epidermidis- coagulase negative. Capable of establishing an
 Non-motile; non-spore formers opportunistic infection. It is part of the normal human flora and can
also cause infection to those who are immunocompromised
B. Culture: 3. S. saprophyticus- can also opportunistic infection. Can cause UTI
 Grow readily on most bacteriologic culture media among sexually active women
 Aerobic or Microaerophilic environment 4. S. lugdunensis
 Rapid growth at 37oC; pigment production best at 20oC-
Note: S. aureus is the only staphylococci that is coagulase positive, the rest are
25oC
coagulase negative
 Colonies on solid media- round, smooth, raised,
glistening ANTIGENIC STRUCTURES OF STAPHYLOCOCCI
o S. aureus- gray to deep golden yellow 1. Cell Wall Components
colonies; (+) varying degrees of hemolysis on a. Teichoic Acids
BAP (Blood Agar Plate) o Important for the attachment of the
o S. epidermidis- gray to white on primary staphylococci on the mucosal surface
isolation → (+) pigments on prolonged o Polymers of polyribitol phosphate; cross-
incubation linked to peptidoglycan
o Effects: antigenic → anti-teichoic acid
C. Growth Characteristics: antibodies found in patients with active S.
 (+) Catalase Production → differentiate from Streptococci aureus endocarditis
All staphylococcus are catalase positive b. Peptidoglycan
S. aureus is coagulase positive o Polysaccharide polymer; destroyed by strong
acid or exposure to lysozyme
 Slowly ferment carbohydrates → (+) lactic acid o Effects/Function:
production, (-) gas 1. Provides rigid exoskeleton of the
 Relative resistance to: cell wall
1. Drying 2. Elicits production of IL-1
2. Heat (withstand 50oC for 30 minutes) (endogenous pyrogen) and
3. 9% NaCl opsonizing antibodies
 Inhibited by certain chemicals (e.g. 3% hexachlorophene) 3. Chemoattractant for PMNs
4. With endotoxin-like activity
D. Mechanisms of Antimicrobial Resistance: 5. Activate complement
1. Production of ß-lactamase – common; plasmid- c. Protein A
mediated; resistance to penicillins (pen G, amipicillin, o Characterized among a group of adhesins
ticarcillin, pepracillin, and similar drugs) called microbial surface components
2. Encoding of gene sequences that provides resistance to recognizing adhesive matrix molecules
specific antimicrobials (MSCRAMMS) – mediate bacterial attachment
o Nafcillin resistance – region of chromosome to host cell
called staphylococcal cassette chromosome mec o Other actions: bind to Fc portion of IgG
(SCCmec) → mecA gene → encodes a low-affinity (EXCEPT IgG3) → can combine with a specific
penicillin binding protein (PBP2a) antigen. Prevents antibodies from binding to
o Methicillin resistance- SCCmec type IV the bacteria, hindering opsonization and acid
o Hospital-acquired infections- SCCmec type I, II, III complement activation
 Used as reagent in immunology and
3. Vancomycin resistance diagnostic laboratory →
o Intermediate resistance (vancomycin- agglutinates bacteria that have an
intermediate S. aureus or VISA)- due to
antigen (coagglutination)
increased cell wall synthesis and alteration in 2. Polysaccharide Capsule
the cell wall; patients with complex infections o At least 11 serotypes → types 5 & 8 responsible for
under prolonged vancomycin therapy majority of infections
4. Plasmid-mediated resistance to tetracyclines, o Action: inhibit phagocytosis by PMNs leukocytes
erythromycins, aminoglycosides, and other drugs
5. Development of tolerance
o Organism is inhibited but not killed by the drug
o Patients with endocarditis
o Due to lack of activation of autolytic enzymes
in the cell wall

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 MICROBIOLOGY
Gram-Positive Cocci (Staphylococci)
Dr. Billones
VIRULENCE FACTORS: ENZYMES & TOXINS
1. ENZYMES
a) Catalase- all Staphylococci are catalase (+) [Strept. are catalase (-)] c) Toxic Shock Syndrome Toxin 1 (enterotoxin F)
 Used for conversion of H2O2 (hydrogen peroxide) to water  Chromosomal-mediated toxin; gene found in 20% of S.
(H2O) and oxygen (O) aureus isolates
 Catalase test is used to differentiate Staphylococci from  Superantigen
Streptococci  Bind to MHC class II molecules → (+) T cell activation
 Catalase test is positive if there is bubble formation,
indicating the presence of oxygen d) Enterotoxins
 A-D, G-J, K-R, U and V; superantigens; 50% of isolates
b) Coagulase- mainly produced by S. aureus  Heat-stable & resistant to GIT enzymes
 Clots oxalated or citrated plasma  Produced when S. aureus grows in carbohydrate and
 Binds prothrombin → (+) fibrin polymerization → fibrin protein foods → important cause of food poisoning
deposited on the surface of the organism → impair  Enterotoxin → causes vomiting and diarrhea
engulfment by phagocytic cells o Enterotoxin A – most common
 Associated with invasive potential o Enterotoxin B
 Also called “Clumping Factor”
 An MSCRAMM → adherence of organism to fibrinogen Stimulate neural receptors in the GIT → activate
and fibrin vomiting center in the CNS → (+) emesis (vomiting)
 Form clumps when mixed with plasma
e) Panton-Valentine Leukocidin (PVL)
 Induces a strong immunogenic response in the host
c) Other enzymes:  Two components (S and F) → same mechanism as gamma
hemolysin
o Hyaluronidase- spreading factor
o Staphylokinase- cause fibrinolysis  Important virulence factor for CA-MRSA (Community
o Proteinases- degrade proteins Associated Methicillin-Resistant S. Aureus)
o Lipases- degrade lipids
o ß-Lactamases- most of Staphylococci are resistant to CLINICAL SYNDROMES
penicillin 1. Staphylococcal Scalded Skin Syndrome (SSSS)
 Bullous exfoliative dermatitis
 Blister- clear fluid without organisms
2. TOXINS  7-10 days
a) Cytolysins (Hemolysins)  Due to exfoliative toxin- a protease that can cleave the
 Target: Eukaryotic cell membranes desmoglein (desmoglein is the one holding the
 It can lead to tissue destruction and abscess formation granulosum and spinosum layer of the skin)
 Four types:  In SSSS, there is separation of stratum granulosum and
1. Alpha-hemolysin – lyse erythrocytes; damage spinosum due to exfoliative toxin
platelets and macrophages; severe tissue
damage 2. Toxic Shock Syndrome (TSS)
2. Beta-hemolysin (Sphingomyelinase C) – acts  Abrupt onset of high fever, diarrhea with vomiting,
on sphingomyelin in the plasma membrane of
myalgias, scarlatiniform rash
erythrocytes
 Severe cases: hypotension with cardiac and renal failure
3. Delta-hemolysin- disrupts biologic
membranes; possible role in S. aureus  Onset within 5 days after onset of menses in young
diarrheal diseases women using high-absorbency tampons
4. Gamma-hemolysin- a leucocidin  May occur in children and men with staphylococcal
 Interacts with Pantov-Valentine wound infection
leucocidin (PVL) → form 6 potential 2-  Organism found in the vagina, tampons, in wounds, other
component toxins → increased cation localized infections, or in the throat
permeability → release of IL-8,
leukotriene, and histamine → 3. Food Poisoning
necrosis + severe inflammation
 Intoxication
 Incubation: 1-8 hours
b) Exfoliative toxin A (epidermolytic toxin)
 Due to enterotoxin
 Encoded by eta gene; superantigen
 Nausea, vomiting, diarrhea without fever (or with low-
 Plasmid-mediated toxin
grade fever [38oC])
 Cause dissolution of mucopolysaccharide matrix of
epidermis → cause Staphylococcal Scalded Skin
4. Staphylococci of low invasiveness
Syndrome (SSSS) or Ritter disease
 Skin infections (e.g. acne, pyoderma, impetigo)
 Also implicated in bullous impetigo
 Epidermolytic toxin A – a chromosomal gene; heat stable
 Epidermolytic toxin B – produced by plasmid; heat labile

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 MICROBIOLOGY
Gram-Positive Cocci (Staphylococci)
Dr. Billones
Clinical Syndromes continued….. S. aureus S. epidermidis S. saprophyticus
5. Wound infection or infection after trauma Colony Color Yellow, white White White variable
 Due to direct contamination Hemolysis (Old
+ +/- -
 Examples: meningitis after skull fracture, open fracture Agar)
(chronic osteomyelitis) Coagulase test + - -
Catalase test + + +
6. Bacteremia Glucose
+ + -
 Endocarditis, acute hematogenous osteomyelitis Fermentation test
(metaphysic of long bones), meningitis, or pulmonary Mannitol
+ - -
Fermentation test
infection (empyema), sepsis
Novobiocin Susceptible Susceptible Resistant
EPIDEMIOLOGY
 Ubiquitous TREATMENT
 Chief sources of infection:  Antimicrobials; choice depends on:
1. Shedding human lesions 1. Type of infection
2. Fomites contaminated from human lesions- clothing, bed 2. Location of infection
linens, etc. 3. Resistance mechanism of organism
3. Respiratory tract  Guidelines for choice of antimicrobials:
4. Skin 1. Organism rapidly develops resistance – perform
5. Hospitals- high risk areas include newborn nurseries, ICUs, antimicrobial susceptibility testing
operating rooms, and cancer chemotherapy wards 2. Drugs are incapable of acting on the central necrotic part
 Normal flora, including respiratory & GI tract- S. epidermis of the abscess
 Transient skin flora- S. aureus 3. It is difficult to eradicate the carrier state of S. aureus
4. Bacteremia and conditions resulting from spread of the
 Colonization with S. aureus:
organism will require prolonged IV therapy with a ß-
 Neonates- umbilical stump, skin and perineum
lactamase resistant penicillin
 Older children and adults- anterior nasopharynx (20-50%
5. Infection with S. epidermidis occur in prosthetic devices
of humans)
→ enable the organism to be sequestered in a biofilm →
PATHOGENESIS more resistant to antimicrobials than S. aureus
 Pathogenicity dependent on interaction of extracellular factors,
toxins, and invasiveness PREVENTION AND CONTROL
 Pathogenic invasive S. aureus  Local antisepsis to prevent recurrent skin infections
o (+) yellow pigment in culture with production of  Exclusion of hospital personnel with active lesions or are carriers
hemolysis; coagulase (+) from high risk areas in the hospital
 Non-pathogenic, non-invasive staphylococci  For carriers:
o Coagulase (-); non-hemolytic o Application of topical antiseptics such as mupirocin to
o Non-suppurative infections nasal or perineal carriage sites to decrease shedding of
o May infect patients with orthopedic or cardiovascular the organism
prostheses as well as immunocompromised patients o Rifampin + oral anti-staphylococcal drug- possible long-
 Produce biofilms → refractory to treatment term suppression and cure of nasal carriage
 S. lugdunensis  Monitoring high risk patients for anterior nares colonization
o Virulent; similar to S. aureus in disease spectrum,
hemolysis & clumping factor
 S. saprophyticus REFERENCES
o Non-pigmented; Novobiocin resistant; nonhemolytic  Microbiology Manual (2018)
o UTI in young women  Dr. Billones Recordings

PATHOLOGY:
 Infection characterized by focal suppuration (abscess)
 Dissemination via lymphatics and bloodstream → suppuration
within veins → (+) thrombosis

LABORATORY DIAGNOSIS
1. Smears
2. Culture
3. Catalase test- identifying Staphylococcus from Streptococcus
4. Coagulase test- identifying S. aureus
5. Susceptibility testing
6. Serologic and typing test

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