LAB - Staphylococci & MRSA

Question 1

Staph family

Answer 1

Micrococcaceae

Question 2

Major staph species associated with human infection

Answer 2

S. aureus
S. epidermidis
S. saprophyticus

Question 3

S. aureus

Answer 3

• skin infections
• wound infections
• enterotoxin-associated food poisoning
• endocarditis
• TSS
• scalded skin syndrome

Question 4

S. epidermidis

Answer 4

• normal flora

- hospital-acquired in immunosuppressed patients as a result of catheterization or valve replacement

Question 5

S. saprophyticus

Answer 5

• normal flora

- UTI in young females

Question 6

S. aureus is carried in the nose of ____% of the general population

Answer 6

20-40%

carriage is higher in hospital personnel

Question 7

Micrococcus colonies

Answer 7

• slower growing than staphylococci
• most produce yellow-orange pigment
• non-hemolytic on BAP
• larger cocci, often in tetrads

Question 8

Catalase test

Answer 8

• enzyme that converts hydrogen peroxide into water and oxygen = visible bubbles

Question 9

a hemoprotein similar in structure to Hb

Answer 9

catalase

- present in most cytochrome-containing aerobic and facultatively anaerobic bacteria

Question 10

pseudoperoxidase

Answer 10

RBCs produce a weakly + catalase so sample from tops of colonies!!!

Question 11

some bacteria in this family produce pseudocatalases which are capable of decomposing H2O2 and producing weak bubbles with some delay

Answer 11

Enterococci

- avoid by reading test within ten seconds

Question 12

T or F. Coagulase should be performed using selective media

Answer 12

F! use nonselective (BAP)

bc colonies isolated on selective media tend to agglutinate in saline, resulting in false positives

Question 13

Slide coagulase test

Answer 13

• bound coagulase or clumping factor (surface of cell wall of most strains of S. aureus); acts directly upon fibrinogen
• should be read within ten seconds
• screening procedure
• isolates positive for this are reported as presumptive S. aureus
• MRSA could be neg for this but pos for tube coag

Question 14

Tube coag test

Answer 14

• detects coagulase secreted extracellularly
• reacts with a substance in plasma called coag-reacting factor (CRF) = thrombin-like complex that converts fibrinogen to fibrin causing clot formation in plasma
• test read by gently tilting tube
• false pos or nef can occur with non-sterile plasma
• read within 4 hours but if neg, held overnight to inspect for delayed clot formation

Question 15

Latex agglutination

Answer 15

• replaces slide and tube coag
• detects bound coag and a S. aureus cell wall antigen = protein A
• late particles coated with fibrinogen and IgG

Question 16

DNAse test

Answer 16

• heat stable DNAse, thermonuclease test
• S. aureus produces heat-tolerant DNAse; different from other Micrococcus sp.
• medium containing DNA with methyl green or toluidine blue O as an indicator
• zone of clearing if DNAse is present for methyl green
• toluidine = positive will show pink around colonies; neg = remain blue

Question 17

Mannitol Salt Agar Test

Answer 17

• selective/differential
• high salt (7.5%) inhibit most bacteria except Staph species
• S. aureus ferments mannitol
• acid produced lowers pH; indicator phenol red changes to yellow
• other Staph don’t ferment mannitol so red colonies with red/purple zone

Question 18

Resistance to novobiocin is used as a presumptive test for recognition of this organism

Answer 18

S. saprophyticus

- zone of inhibition less than or equal to 16 mm are resistant

Question 19

Oxidase test with 6% TMPD for Micrococci vs. Staphylococci

Answer 19

cytochrome C in Micro but not in Staph

both are oxidase neg by traditional test

Question 20

Bacitracin for Micrococci vs. Staphylococci

Answer 20

micro is inhibited but staph shows no zone of inhibition

Question 21

Furazolidine

Answer 21

growth of Staph inhibited but Micro shows no zone of inhibition

Question 22

OF test

Answer 22

majority of Staph produce acid from glucose anaerobically (ferment), Micro are oxidative glucose utilizers

Question 23

This Staph enterotoxin is associated with majority of food outbreaks

Answer 23

Staphylococcus Enterotoxin A (SEA)

• intoxication
• detected in filtrates prepared from vomitus or food filtrates (not stool bc absorbed in gut)

Question 24

iatrogenic infection

Answer 24

physician transmitted

Question 25

These two methods are used to separate strains into specific groups or types (Staph strains)

Answer 25

• phage typing: based on susceptibility of Staph to certain bacterial phages or viruses
• DNA analysis

Question 26

Direct immunofluorescence antibody (DFA; 1 step)

Answer 26

• used to ID unknown bacterial antigen
• antiserum conjugated with fluorescein isothiocyanate is prepared + join with patient sample
• smear viewed under UV microscope; if fluorescence = bacteria is present
• faster = one incubation period
• more specific method for detecting antigens

Question 27

IFA (2 steps)

Answer 27

• detection of either Ab or Ag
• if detecting Abs in patient serum = prepare known Ags + patient sample = Ag-specific Abs will bind; wash away excess; THEN labelled antihuman globulin (AHG) applied = microscopy
• more sensitive method for detecting antigens

Question 28

MRSA identification strategies

Answer 28

• Oxacillin Agar Screen
• Oxacillin Disc Screen
• MRSA Chromagar
• PCR = gold standard (identifying methicillin resistance mecA gene)

Question 29

Oxacillin Agar Screen

Answer 29

• Mueller-Hinton agar medium = 6 micrograms of oxacillin/ml and 4% NaCl
• after incubation, any growth = resistance