Micrococcus spp. Flashcards by Nicole Angeline

Micrococcus spp. can be mistaken for?

Staphylococcus spp.

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Micrococcus spp. colonies

✦ Tetrads or cuboidal packets
✦ Non-hemolytic, lemon yellow colonies
on BAP

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Micrococcus spp. characteristics

✦ Utilizes glucose oxidatively
✦ With growth on 6.5% NaCl
✦ Nitrite (-)
✦ Not lysed with lysostaphin
✦ resistant ot furazolidone
✦ Susceptible to bacitracin (0.04 units)

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Culture media for Micrococcus spp.

Sheep blood agar ( BAP )
Mannitol salt agar (MSA)
Columbia colistin nalidixic acid (CNA)
Phenylethyl alcohol (PEA) agar

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BAP

to see hemolytic pattern

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MSA

selective
and differential culture medium; it has
increased level of salt (7.5% NaCl

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CNA & PEA

selective culture media for

gram-positive cocci

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Mannitol Salt Fermentation Test

Culture medium: MSA
• Contains a high concentration of salt
(7.5%) — most bacteria can’t grow
in this level of salt
• Sugar: mannitol
• pH indicator: phenol red — acid
production turns phenol red to
yellow
• Selective differential culture media; it
is selective for organisms that can
tolerate high levels of salt and it is
differential for mannitol and nonmannitol fermenters.

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Coagulase test

Best single criterion of pathogenicity

of Staphylococcus aureus

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2 methods of coagulase test

slide and tube method

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slide method

Slide method
‣ Detects cell-bound coagulase /
clumping factor
‣ Reagent: human, rabbit, pig
plasma —usually human/rabbit
plasma
‣ Positive result: clumping or
agglutination
‣ Positive: Staphylococcus aureus
‣ Other cell-bound coagulase
positive organisms:
-Staphylococcus lugdunensis
-Staphylococcus schleiferi

If the slide test is negative, you can
proceed to tube method.

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tube method

‣ Detects unbound / extracellular
coagulase
‣ Reagent: human, rabbit, pig
plasma
‣ Positive result: clot formation
after 4 hours of incubation at
37C; extend incubation if
negative result is observed.
‣ Positive: Staphylococcus aureus
‣ Other extracellular coagulase
positive organisms:
-Staphylococcus hyicus
-Staphylococcus intermedius
-Staphylococcus delphini
-Staphylococcus schleiferi subsp.
coagulans

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PYR detection

(+) red color

• The substrate pyroglutamyl-Bnaphtylamide is hydrolyzed to Lpyrrolidone and B-naphtylamine
which combines to pDAB to form a
red compound
• Positive:
• S. lugdenensis
• S. intermedius
• S. scleiferi
• Negative: S. aureus

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Voges-Proskauer (VP) test

Formation of acetoin from glucose
or pyruvate
• Reagent: a-naphthol and 40% KOH
• Positive: S. aureus
• Negative: S. Intermedius

also for identification of enterobacerioceae

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Aerobic utilization of glucose

(+) staphylococci

(-) micrococci

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Susceptibility to: Lysostaphin

S - staphylococci

R- micrococci

Susceptibility to: Bacitracin

R - staphylococci

S -micrococci

Susceptibility to: Furazolidolne

S - staphylococci

R - micrococci

Catalase test

+ both

Modified oxidase test

(-) staphylococci

(+) micrococci

Antimicrobial susceptibility testing

✦ Most S. aureus are resitant to penicillin due to B-lactamase (Penicillinase)

✦ Penicillin-resistant strains require
treatment with penicillinase-resistant
penicillina

penicillinase-resistant

penicillins

methicillin, oxacilin, nafcilin

methicillin-resistant

spahylococci- MRSA

✦ Isolates that are resistant to nafcillin

or oxacillin

Methicillin-Resistant Staphylococcus

aureus (MRSA)

Associated risk: recent
hospitalization, long-term care,
dialysis and indwelling devices.
✦ Primary mechanis: production of
altered penicillin-binding protei
(PBP2a/PBP2’) which renders all
currently available B-lactans
essentially ineffective
✦ Gene that encodes for PBP 2a- mecA
gene

Oxacillin

was generally used for detection of methicillin resistance from staphylococci species

Cefoxitin

better inducer of mec-Amediated rsistance

Lab diagnosis for methicillin resistant s. aureus

oxacillin- salt agar plate and cefoxitin disk diffusion

oxacillin- salt agar plate

- (+ 2-4% NaCl) - for screening of MRSA in clinical samples or as a way to differentiate MRSA isolates from those that are hyperproducers of B-lactamase (organisms that are resistant with Blactams not because they have the gene but because they are hyperproducers), known as borderline oxacillin-resistant S. aureus (BORSA)

Cefoxitin Disk Diffusion

- (30ug) - detect methicillin resistance in S. aureus and S. lugdunensis. • Resistant: inhibition zone < 21mm • Susceptible: inhibition zone > 22mm • CoNS- oxacillin testing of > 25 mm is susceptible and 24mm is resistant ** Susceptibility testing with cefoxitin is the recommended method for the detection of the susceptibility or resistance of penicillinase-resistant penicillins

Chromogenic selective, differential | media (CHROMagar MRSA)

``` • branded type of culture medium that is specific or selective for MRSA • Specimen of choice: nasal samples • Cefoxitin is incorporated in the media • 24-48 hours incubation • MRSA isolates will produce a colored colony ```

``` Nucleic acid probe or Polymerase Chain Reaction (PCR) amplification ```

* “gold standard” for MRSA detection | * Specimen: anterior nare swabs

Vancomycin

drug of choice for | serious staphylococcal infections

Vancomycin agar plate

enhance the detection of Vancomycin-intermediate S. aureus (VISA) and Vancomycinresistant S. aureus

Macrolide resistance

``` - for treatment of resistance, drugs involved: Erythromycin and Clindamycin - a constitutive mechanism or inducible mechanism that is activated by the presence of erythromycin ```

Macrolide resistance mechanism

• msrA gene- coded for an efflux mechanism, which results in resistance to erythromycin but susceptibility to clindamycin. • erm gene- confers cross resistance to the Macrolides-Lincosamidestreptogramin B (MLSB); may be expressed as a constitutive or inducible mechanism.

Double-Disk Diffusin (D-test)

``` ✦ To determine the organism’s susceptibility to clindamycin ✦ Two antibiotic disks are used, clindamycin (2ug) is placed 15mm from an erythromycin disk (15ug) on a Mueller Hinton Agar plate. ```