Gram Positive Cocci

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50 Terms

1
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Family: Micrococcaeae includes genera ___, but Staphylococcus is now __.

Micrococcus, Rothia, Staphylococcus;

it’s own family = Staphylococcaceae

2
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Staphylococceae general characteristics

  • GPC

  • single, pairs, tetrads, or clusters

  • catalase + except Rothia

  • col entire, smooth, opaque / white & butyrous growth

3
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What test diff Staph from Strep? How does it work

catalase test

  • reagent = 3% H2O2

  • + → production of O2 and H2O (bubbles)

  • - → no bubbles

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Which genera are catalase + and -?

catalase + = Staphylococcus & Micrococcus

catalase - = Streptococcus & Enterococcus

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What can cause a false +?

if pick up blood/rbc from agar → contains catalase → false +/bubbles

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Which bacti is catalase + besides Staphylococcus? How to diff b/t other Staphylococceae?

Rothia mucilaginosa (sticky staph)

Rothia can’t grow in 5% NaCl

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once catalase +, what test is done next to determine if S. aureus?

slide coagulase

  • media = rabbit plasma w/EDTA

  • rapid

  • tests for coagulase enzyme (which alters fibrinogen → clot), specifically clumping factor/cell-bound coagulase

  • if (-) → confirm w/ tube coag test

    • if (+) → S. aureus

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why does slide coagulase test need negative control? what to do to if this occurs?

neg control checks for bacti that auto-agglutinate in saline → if auto-agglut → tube coag test

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when is tube coagulase test done? what is it and what to look out for?

when (-) from slide coag test → will test for free coagulase

  • inoculate loop of bacti into 0.5 mL of rabbit plasma → incubate 35C for 4 hr → observe at 4 hr mark in case of staphylokinase which lyses clot → if (-), incubate overnight

pos = clot formation or aggulatination

neg = none

10
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Staph Latex / hemagglutination test

  • detect what?

    • advantage/disadvantages?

detect: clumping factor, protein A (all S. aureus have virulence factors), and antigenic proteins

  • rapid & more definitive, if + → call it S. aureus

  • false - b/c some strains don’t produce enough of ^^^

  • false + from certain Strep A, C, and G

    • get + from other Staph sp like S. lugdenesis, schleiferi, etc…

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S. aureus

  • infects where

  • usually founds in __

  • produces enterotoxins ___

in skin (furuncles, carbuncles, cellulitis, impetigo), pneumonia, meningitis, endocarditis

  • thermostable enterotoxin (food poisoning): projectile vomiting & fast incubation time

  • TSST-1 → toxic shock syndrome

  • exfoliatin → scalded skin syndrome in kids

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ID of S. aureus

  • grows in 18-24h at 37C

  • turns golden yellow w/ age

  • typically beta-hem rxn

  • coagulase +

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S. aureus epidemiology

Most are resistant to __?

a significant nosocomial pathogen (hospital acquired)

95% resistant to penicilin due to Beta-lactamase or cell wall binding protein

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MRSA stands for

what happens if pt is + for MRSA?

Methicillin/oxacillin resistant S. aureus due to mecA or mecC gene → codes for PBP2A (penicillin binding protein)

15
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Why is cefoxitin disk better for detecting MRSA than oxacillin disk or agar?

What will it look like?

cefoxitin (= a cephalosporin, also has beta-lactam ring) → detects heteroresistant strains of MRSA (=don’t express Abx-R genes)

MRSA → cefoxitin-R → report 6mm ZOI

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What does MRSA look like on chromogenic media?

Spectra MRSA → denim blue col

CHROMagar MRSA → mauve col

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Methods to rapidly detect MRSA?

detection of mecA gene via PCR/probe, fluorescence tests, or slide latex agglutination

18
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MRSA are resistant to ___ →

methicillin, oxacillin, nafcillin (semisynthetic penicillins) → other beta-lactam drugs are reported as resistant even if in vitro tests appear sensitive

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What may be misID’d as S. aureus?

S. lugdunensis is misID’d b/c can look + on slide coag test (but is coag -) bc may have clumping factor and yellow pigment col

  • confirm w/tube coag test that it’s coag -

  • part of normal skin flora (toe or inguinal fold) → but can cause soft tissue infns

  • AST interpretation reported like a S. aureus, NOT CNS

  • ornithine decarboxylase +

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Coagulase-negative Staph (CNS)

significant species and when to w/u?

Staph epidermidis - bad bc produce slime → adhere to catheters & prosthetics

S. saprophyticus - ID from novobiocin-R, UTI in young females

  • only do w/u if multiple + BC or +BC AND +catheter tip

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S. saprophyticus is resistant to ___.

novobiocin

< 16 mm ZOI

22
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How to diff Micrococcus luteus from Staph spp.?

bright yellow col, coag neg

diff from Staph

  • glucose oxidizer (Staph ferm glucose)

  • bacitracin-S (Staph are R)

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Streptococcus & related general traits

GPC in pairs and chains

facultative anaerobes

translucent, shiny, grey col

catalase -

24
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What is the Lancefield Grouping system?

used for beta-hemolytic Streptococci

based on carbohydrate cell wall Ag → detected by kits via heat, chemical, enzyme

groups include:

  • A and B - can use biochem tests to ID

  • C, F, G - need serology tests to ID

    • D = Enterococcus?

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Group A Strep aka __. Colony morph? Virulence mechanism?

Strep. pyogenes

tiny, beta-hemolytic, with larger zone of hemolysis

can be elongated cocci in chains

Streptolysin O (oxygen labile ie breaks down in O2) → if stab into agar → enhanced zone of beta-hemolysis

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GAS clinical manifestations

spread via droplets or direct contact

most common cause of pharyngitis

impetigo, erysipelas

necrotizing fasciitis (got thigh swab from unknowns!)

strep toxic shock syndrome

after infection (sequelae) →

rheumatic fever: Ab from infn attack heart tissue

glomerulonephronitis - Ab/Ag complex deposits in kidneys

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Media used for GAS?

Bacitracin → ?

PYR → ?

SXT or SSA

bAcitracin-S (A on disk is for group A strep)

PYR + (pink throat)

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GAS rapid tests

use throat specimen

positives are conclusive, but neg must have culture

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Group B Strep (Strep. agalactiae)

colony morph

found where?

clinical manifestations

larger col w/ narrow zone of beta-hemolysis (or diffuse zone)

normal flora of GI & genitourinary tracts

GBS is screened for in pregnant women in 3rd trimester → neonatal pneumonia, septicemia, meningitis

in adults: UTI, bacteremia, endocarditis, pneumonia, osteomyelitis

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media for GBS?

Hippurate hydrolysis → ?

CAMP test → ?

use CNA & LIM broth (contain colistin & nalidixic acid)

hippurate hydrolysis + (hippy girls/bros like camping and gardening)

CAMP test + (like to camp in the vag/rectal area)

**hip gals (galactiae) camp positive

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hippurate hydrolysis test

principle

steps

result

sodium hippurate → benzoic acid + glycine

glycine + ninhydrin → purple complex

inoculate bact in 0.5 mL distilled water in tube → add sodium hippurate disk → 35C for 2hr → 0.2 mL ninhydrin reagent

pos = blue/purple w/in 5 min

neg = no color change

<p>sodium hippurate → benzoic acid + glycine </p><p>glycine + ninhydrin → purple complex</p><p></p><p>inoculate bact in 0.5 mL distilled water in tube → add sodium hippurate disk → 35C for 2hr  → 0.2 mL ninhydrin reagent </p><p>pos = blue/purple w/in 5 min</p><p>neg = no color change</p>
32
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Hippurate hydrolysis test is positive for

GBS, Listeria monocytogenes, Gardnerella vaginalis, Campylobacter jejuni

“Hippy Girls Like Camping & Gardening”

33
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CAMP test

principle

steps

results

ID for GBS

free CAMP factor reacts w/Staphylococcal beta lysin to lyse rbcs

single straight streaks of test bact & CAMP S. aureus are made perpendicular to each other on SBA

pos = enhanced zone of beta hemolysis in shape of arrowhead pointing toward S.aureus

neg = no enhanced zone of hemolysis

<p>ID for GBS</p><p>free CAMP factor reacts w/Staphylococcal beta lysin to lyse rbcs</p><p>single straight streaks of test bact &amp; CAMP S. aureus are made perpendicular to each other on SBA</p><p>pos = enhanced zone of beta hemolysis in shape of arrowhead pointing toward S.aureus</p><p>neg = no enhanced zone of hemolysis </p>
34
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Group C, F, and G Streptococcus

sources

clin manif?

how to ID?

still beta-hemolytic

normal flora of skin, nasopharynx, GI and genital tracts

infns in immunocompromised pt → similar infn as GAS and GBS, but no sequelae like GAS

ID w/kits

35
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Group D Strep includes __

morphology

clinical significance

intrinsically resistant to __?

Enterococcus & Group, non-Enterococcus

grey-white, translucent, alpha or non-hemo, some rare beta-hemo

found in soil, food, water, normal flora of birds, animals, human (GI and female GU tract) → can cause UTI, bacteremia, endocarditis, mix wound infn

cephalosporins and aminoglycosides → need to treat w/multiple Abx for synergistic effect

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How to distinguish Enterococcus spp vs Group D non-Enterococcus?

bile esculin

PYR

salt broth 6.5% NaCl

both have group D Ag

bile esculin → all group D + (black → dlack)

PYR → only Enterococcus + (pink)

salt broth → only Enterococcus + (purple or yellow growth)

37
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bile esculin slant

reaction

results

40% bile → inhibits most G+ → used to diff Group D from non-group D strep

esculin → esculetin + dextrose → esculetin + ferric citrate → black pigment

pos = >1/2 slant turned black

neg = <1/2 slant black

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PYR test

principle

steps

result

used to diff Enterococcus spp from Group D, non Enterococcus

PYR enzyme made by GAS & Enterococcus.

PYR -L pyrglutamyl aminopeptidase → free beta-naphthylamine —cinnamaldehyde → red complex

moisten substrate disk w/sterile water → touch colony to disk → incubate 2 min → add cinnamaldehyde

pos = red color on disk → GAS or Enterococcus spp.

neg = no color change

39
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PYR + organisms

GAS → r/o Group B, C, F, G Strep, S. viridans

Enterococcus spp. → r/o S. gallolyticus

(some Staphylococcus strains, Micrococcus, Gemella, Lactococcus)

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6.5% NaCl test

used to ID Enterococcus spp

inoculate and incubate overnight 35C

pos = growth (w or w/o change to yellow)

neg = no growth

<p>used to ID Enterococcus spp</p><p>inoculate and incubate overnight 35C</p><p>pos = growth (w or w/o change to yellow)</p><p>neg = no growth</p>
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Enterococcus faecalis

causes most human enterococcal infn

non-hemo

** Cali = huge population, most infns, non-hemo bc californians are chill → not resistant ie VRE

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Enterococcus faecium

alpha-hemo

causes 10-15% enterococcal infn

more commonly vancomycin-R (VRE)

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Enterococcus gallinarum/cassliflavus

motile

intrinisically vancomycin intermediate

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Group D non Enterococcus aka

Streptococcus bovis (old name) → S. gallolyticus

assoc’d w/colon cancer

45
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Strep pneumoniae

morphology

clin manif

GPC in pairs, lancet shaped, may have capsule

always alpha-hemo, older colonies may be donut-shaped (autolysing)

leading cause of bact meningitis and pneumonia; bacteremia, sinusitis, otitis media

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ID’ing Strep pneumoniae

bile solubility

Optochin test

Quellung reaction

bile solubility → +, colony dissolves → can see alpha-hemo

Optochin → S

Quellung rxn (only for epidemiology) → capsule is enhanced for certain S. pneumo strains

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Viridans Group Streptococcus

morphology

clinical signifance

typically alpha-hemo, but can be non or beta (viridans vAries)

butterscotch odor (sweet → viridans (veneers) → endocarditis)

opportunistic pathogens, normal flora of oral cavity, GI tract, female genital tract → poor dental care → bacteremia, endocarditis, meningitis

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Viridans Group Strep ID

PYR and salt

bile esculin

optochin

bile solubility

all negative!

w/u only if multiple +BC

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Abiotrophia spp aka ___

requires ___ to grow → grows on __ agar not SBA.

Will grow on SBA if?

PYR → ?

Granulicatella spp.

pyridoxal or thiol cmpds or cysteine; choc

linked to endocarditis (normal oral cavity flora)

beta-hemolytic Staph on SBA → B-hemo → sattelite streptococci in lysed region

PYR → +

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other catalase - GPC