DAVAO DOCTORS COLLEGE

MEDICAL LABORATORY SCIENCE DEPARTMENT

STUDENT NOTES: MICRO 1
STAPHYLOCOCCI

GENERAL CHARACTERISTICS: - facultative anaerobe and exhibits GOLDEN YELLOW

-non-motile, catalase-producing, gram (+) cocci PIGMENTATION due to the action of the pigment
-Greek word “staphle” = bunch of grapes STAPHYLOXANTHIN
-smear: exhibit spherical cells
-colonies: medium-sized, β hemolytic colonies that are creamy or white VIRULENCE FACTORS:
& rarely gold 1. ENTEROTOXINS
-members of the family Staphylococcaceae - are acid-stable & heat stable
-resemble some members of the family Micrococcaceae, such as the - 9 serologically distinct enterotoxins have been
genus Micrococcus: catalase- producing, coagulase (-), gram (+) cocci identified
- A: food poisoning
CHO OXIDATION FERMENTATION TEST - B: staphylococcal pseudomembranous colitis
- indicator: bromthymol blue - C & D: contaminated milk
- medium: Hugh & Leifson tubes 2. TOXIC SHOCK SYNDROME TOXIN-1 (TSST-1)
- formerly referred to as Enterotoxin F
MODIFIED OXIDASE TEST - only toxin that is SYSTEMIC
Principle: - common in: MENSTRUATING WOMEN using
Micrococcus organisms possess CYTOCHROME C in the cytochrome TAMPONS
oxidase system, while Staphylococci do not.
Reagent: 3. EXFOLIATIVE TOXIN (Epidermolytic Toxin)
6% tetramethylphenylenediamine hydrochloride in dimethyl sulfoxide - causes the epidermal layer of the skin to slough off and
or Microdase Disk (with impregnated Modified oxidase is known to cause STAPHYLOCOCCAL SSS
reagent) 4. CYTOLYTIC TOXINS
Expected Results: - α HEMOLYSIN: disrupts smooth muscle in blood vessels
(+): Dark-blue - β HEMOLYSIN (SPHINGOMYELINASE C): “hot-cold
(-): No color change lysin”; acts on sphingomyelin in the plasma membrane of
RBC
LYSOSTAPHIN SUSCEPTIBILITY TEST - δ HEMOLYSIN: destroy surfactants in cell membrane
Principle: (like detergent) affecting RBC and is considered less toxic
Lysostaphin is an endopeptidase that cleaves the glycine-rich peptide to cells than α & β hemolysin
linkages, rendering the cells susceptible to osmotic lysis - γ HEMOLYSIN: toxic to WBC which causes lysis to the
(lysostaphin susceptible). cell and often found only in association with Panton-
The interpeptide bridge of Micrococcus does not contains glycine. Valentine Leukocidin (PVL)
5. PROTEIN A
TEST Staphylococci Micrococci - ability to bind to the Fc portion of IgG
6. ENZYMES
- COAGULASE: conversion of fibrinogen to fibrin
CHO Utilization FERMENTERS OXIDIZERS - STAPHYLOKINASE (fibrinolysin):
dissolves fibrin clot and may enable release of bacteria →
Oxygen Requirement FACULTATIVE STRICTLY spread of infection begins
ANAEROBE AEROBE -LIPASE: hydrolyze lipids in plasma & skin
-HYALURONIDASE: “spreading factor”; hydrolyzes
Modified Oxidase (-) (+) hyaluronic acid present in connective tissue causing the
spread of infection
Bacitracin (0.04 units) R S - DNAse: hydrolyzes DNA
- β- LACTAMASE (Penicillinase): hydrolysis &
Lysostaphin Test S R inactivation of penicillin

Staphylococcus aureus Staphylococcus epidermidis

- most clinically significant species of staphylococci -former name is Staphylococcus albus
-infections caused are predominantly hospital acquired such as
PROSTATIC HEART-VALVE ENDOCARDITIS
-BIOFILM PRODUCTION
DDC Medical Laboratory Science Program | 2020-2021 Prepared by: Rachel Ann O. Eromon, RMT
 4. BETA-LACTAMASE (CEFINASE) TEST
Staphylococcus saprophyticus ✓ detect β lactamase production
- associated with UTIs IN YOUNG WOMEN ✓ CEFINASE DISK is impregnated with the CHROMOGENIC
- in urine cultures, S. saprophyticus may be found in low CEPHALOSPORIN NITROCEFIN
numbers (<10,000 colony-forming units/ mL) and still be ✓ (+): yellow → RED
considered SIGNIFICANT ✓ (-): NO COLOR CHANGE
5. DNA HYDROLYSIS/ DNAse TEST
BIOCHEMICAL CHARACTERISTICS: - detects the ability of an organism to hydrolyze DNA with DNAse
1. CATALASE TEST 6. Growth on MANNITOL SALT AGAR
• Staphylococci VS. Streptococci ✓ inhibitor: 7.5% NaCl
• reagent: 3% H䔖䔖 (+) result is EFFERVESCENCE/ ✓ CHO: MANNITOL
✓ pH indicator: PHENOL RED
Bubble Formation ✓ Mannitol fermenter: “yellow halo” (Staphylococcus aureus)
• (+) Staphylococci ✓ Non-mannitol fermenter: CoNS (Staphylococcus
• (-) Streptococci saprophyticus & Staphylococcus epidermidis)
2. COAGULASE TEST
Staphylococcus aureus VS. CoNS
2 forms: cell bound coagulase (“clumping factor”)
free coagulase (“staphylocoagulase”)
(+): Staphylococcus aureus
(-): CoNS (Staphylococcus saprophyticus & Staphylococcus
epidermidis)

A. SLIDE COAGULASE TEST

✓ for detecting CELL BOUND COAGULASE
✓ Test: Rabbit Plasma + Organism
✓ Control: NSS + Organism
✓ (+) MACROSCOPIC CLUMIPING in 10 seconds
✓ (-) NONE, proceed to tube test to confirm
B. TUBE COAGULASE TEST
✓ for detecting FREE COAGULASE
✓ sterile test tube with 0.5 mL RABBIT PLASMA
- then inoculate colonies to give MILKY SUSPENSION
✓ incubate 35°C for 24 hrs.
✓ (+) COAGULUM/ CLOT FORMATION
3. NOVOBIOCIN SUSCEPTIBILITY TEST
PURPOSE: to differentiate S. saprophyticus from S. epidermidis
PRINCIPLE: after incubation with 5µg NOVOBIOCIN, S.
saprophyticus is not inhibited by the antibiotic, while S. REFERENCES:
epidermidis is susceptible Delost, Maria Dannessa. Introduction to Diagnostic Microbiology for
RESULTS: the Laboratory Sciences. 1st Edition, 2014
Susceptible: zone is >16mm Mahon, Connie & Lehman, Donald. Textbook of Diagnostic
Resistant: zone is <16mm Microbiology, 6th Edition, 2018
Tille Patricia M. Bailey and Scott’s Diagnostic Microbiology, 14th
Edition, 2018

DDC Medical Laboratory Science Program | 2020-2021 Prepared by: Rachel Ann O. Eromon, RMT