Bacte Lec

Gram positive cocci- 

streptococcus and enterococcus 
 
Streptococci  ● Alpha prime
- Microscopy: ​gram positive - Haemolysis is an inner zone
spherical cells, chains or pairs of alpha hemo, and the outer
- Culture: grayish, pinpoint, zone of beta hemo
translucent to slightly opaque ● Alpha prime hemolysis
colonies; mucoid colonies, fastidious - colony with green
- Biochemical test: catalase (-), discoloration followed by a
oxidase (-), non-motile; ferments clear zone
carbohydrate 2. Lancefield classification (antigen
- All strep except viridans group have serogrouping)
a layer of C-carbohydrate a. Based on the antigenic
(polysaccharide) for serological nature of cell wall
classification. carbohydrates
- Streptococcus pyogenes and ( C-polysaccharides)
pneumoniae- most notorious b. Most significant in classifying
pathogens and identifying B-hemolytic
streptococcus (we use
antibodies so it is specific)
c. By Dr. Rebecca Lancefield

Group  A  streptococci  -  Streptococcus 

pyogenes  
- Not considered part of the normal
flora - pathogenic to man
- Transmission: contaminated droplets
Pinhead- mukhang pin, mataba (for staph)
by cough or sneeze (resides in the
upper part of respiratory tract)
- Resistant: drying (recovered from
swabs after several hours of
collection)
- You can still isolate the
pyogenes even the swab is
dried out
- The commensals will die but
not pyogenes
- Culture: small, translucent and smooth,
well-defined B- hemolysis

Virulence factor:
Pinpoint- size of tail of pin (for strep) 1. M-protein
a. Immunogenic (substances
Scheme of classification for  that can produce immune
Streptococcus   response ; ex. Antibody
formation)
1. Smith and Brown’s classification b. More than 200 (M1, M2,
a. Based on hemolytic pattern M3,...)
b. We use the BAP c. Problem: antibodies produce
are specific because there
are 200+ M-proteins
d. Coded by gene emm
 e. Most significant- 3. DNAse
anti-phagocytic a. Enzymes that destroys DNA,
2. Fibronectin-​ binding protein (protein lower viscosity of exudate
F) b. A, B, C , D - different
3. Lipoteichoic acid serotypes
4. Hyaluronic acid capsule i. Most common and
a. Capsule - inhibits specific - DNAse B
phagocytosis and used for 4. Streptokinase
attachment of bacteria to the a. Dissolves clot
host 5. Hyaluronidase
5. Hemolysins, toxin, and enzymes a. Spreading factor

Toxins produced by Streptococcus  Infections and diseases  

pyogenes   1. Pharyngitis or tonsillitis ​- “strep
1. Hemolysins throat”
● Streptolysin O- “oxygen labile” , a. Diagnosis relies on throat
highly antigenic culture or direct antigen
a. Responsible for subsurface detection
hemolysis on bap incubated 2. Scarlet fever
anaerobically - stab the agar a. Scarlatina
to create anaerobiosis ​(why b. Punctate exanthem overlying
do we need to stab the diffuse erythema and
agar?) appears initially on the neck
b. If the body is in infection with and upper chest , 1-2 days
pyogenes, it will produce following strep throat
Anti-streptolysin O c. Called as second disease-
c. Responsible for Anaerobic because 2nd most common
hemolysis cause of rashes amongst
d. Causes lysis of WBCs, children
platelets, tissue cells; d. Causes by the release of
induces Antibody response pyrogenic exotoxins
e. Anti-streptolysin O test- e. Communicable and spread
detects recent infection with by inhalations of infection
pyogenes (4-fold titer) respiratory droplet
f. Cardinal sign: strawberry
● Streptolysin s - “oxygen stable” , -colored tongue
nonantigenic g. Susceptibility test: Dick’s test
a. Responsible for aerobic (erythrogenic toxin)
hemolysis and surface i. (+) - erythema or
hemolysis on BAP redness of the test
b. Cause lysis of WBCs site
h. Diagnostic test: Schultz
2. Streptococcal pyrogenic exotoxins Charlton (anti-erythrogenic
a. Sertypes SpeA, SpeB, SpeC, toxin)
SpeF i. (+) - “​blanching
b. Formerly known as phenomenon​” - rash
erythrogenic toxins fade
c. Super antigens - causes non 3. Skin infections
specific activation of t cells (T a. Cellulitis, erysipelas,
cells should be activated by a impetigo
specific antigen) and can b. Caused by group a
cause autoimmune diseases streptococcus strains may
d. Exotoxin b- (cysteine also lead to necrotizing
protease) , degrades fasciitis “galloping gangrene
proteins, mediates rash in or flesh-eating bacteria
Scarlet fever syndrome)
 4. Complication of strep throat Group B streptococci- Streptococcus 
a. Rheumatic fever (heart is agalactiae  
affected) and AGN (acute
- Normal flora of the female genital
glomerulonephritis- because
tract and lower GIT
of cross reactivity)
- Nosocomially transmitted by
5. Streptococcal toxic shock
unwashed hands of mother or health
syndrome (strep TSS)
care personnel to the newborn or
a. Whole organ shuts down
infant
b. It can result to death
- Cause infections of fetus and
6. Necrotizing fasciitis
premature rupture of mothers
a. Invasive infection charac by
membranes
rapidly progressing
- Culture: grayish white, mucoid
inflammation and necrosis of
colonies w small zone of
the skin, subcutaneous fat,
b-hemolysis
and fascia
- PYRase - used to differentiate
between group A and B
Laboratory  diagnostics  tests  for  group  A  - Virulence factor- capsule (sialic
streptococcus   acid)
- Specimen: pharynx and tonsillar
swabs (throat swab) ** CDC recommendations: all pregnant
woman be screened for group B strep at
1. Bacitracin disc test / taxo A (0.04 35-37 weeks of gestation
units)  
a. Screening test for group A
Laboratory  diagnostic  test  for  group  B 
strep in throat cultures
b. (+) result: any ZOI streptococcus  
(susceptible) 1. CAMP test (​Christie
c. Group C and G are also -Atkins–Munch-Peterson)
susceptible to Bacitracin a. reagent - B-lysin producing
2. Sulfamethoxazole and strain of s. Aureus or B-lysin
trimethoprim (SXT) test disk
a. (+) - resistant (group A and b. (+) result- arrow-head
B) B-hemolysis near s. Aureus
b. Most interfering respiratory growth or (+) bowtie
microbiota will be inhibited by appearance
SXT
c. Group C strep is susceptible 2. Hippurate hydrolysis test
(negative result) to SXT a. Used to diff from agalactiae
3. L-pyrrolidonyl- a-naphthylamide to pyogenes
(PYR) test b. Streptococcus agalactiae
a. Specific than Bacitracin test possesses the enzyme
b. It detects the presence of hippuricase or hippurate
PYRase hydrolase
c. (+): bright/cherry red color c. Reagent: sodium hippurate +
(B-naphthylamide) ninhydrin
d. (-); no color change d. Results: (+) purple color after
e. Pyogenes is the only adding Ninhydrin
B-hemolytic streptococcus (-) no color change
that is PYR test (+)
f. Other PYR test (+) Na hippurate ---(Streptococcus
organisms: enterococcus, agalactiae)---> Na benzoate + glycine
aerococcus, and gemella Na benzoate + ninhydrin reagent -----> (+)
(not B-hemolytic) purple color
 1. Leucine aminopeptidase (LAP)
test
SXT Bacitracin PYR
Substrate: Leucine-B- naphthylamides
S. R S + Reagent: p-dimethylaminocinnamaldehyde
pyogenes (DMACA)
End product: B- naphthylamine (red color)
S. R R - LAP (+)- streptococcus, viridans
agalactiae strep, enterococcus and pediococcus
LAP (-); aerococcus and
leuconostoc

Group  C  and  G  streptococci  (Animal 

Pathogens)  
- They possess M protein just like the
group A streptococci
- Group C streptococci are the main
source of streptokinase
- Species: ​S. dysagalactiace subsp.
Equismilus, S. equi subsp. Leucine B-naphthylamide --(viridans)
zooepdemicus (LAP)--> B-naphthylamine
- Antimicrobial susceptibility tests: B-naphthylamine + DMACA ---> (+) red
- Group C streptococci is color
susceptible to bacitracin and
SXT 2. Voges Proskauer (VP) test
- Group G streptococci may be - It is used to distinguish the
bacitracin resistant or small-colony-forming B-hemolytic
susceptible anginosus group containing groups
A or C antigens from
Viridans streptococci   large-colony-forming pyogenic
- Also known as alpha-prime strains with the same antigens.
streptococci that lack the Lance - Have suspensions of bacteria in 2ml
group antigens and do not fall on the VP broth; incubate 35C for 6 hours
criteria for S. pneumoniae then ass the reagents.
- Are URT, Female Genital Tract, and - It detects acetoin production (pink or
GIT commensals red color) from glucose after 30
- Are the most common cause of SBE minutes of RT incubation.
-- individuals with rheumatic fever VP (+) - anginosus group
are susceptible to SBE.
** Viridans- green Laboratory  test  for  S.  bovis Group (Group 
D streptococci)  
Virulence factor
- Capsule, cytolysin, extracellular
dextran and adhesins Test Result

Growth in bile esculin +

Infections   medium
- SBE (S. mitis group), gingivitis,
dental caries, meningitis Growth in 6.5% NaCL -
osteomyelitis, empyema
PYR -
Laboratory tests:  
Bile insoluble (viridans in insoluble), Penicillin S
optochin resistant, no growth in 6.5% NaCl,
LAP (+) and PYR (-).
Notes to remember: (Cephalexin-aztreonam-arabi
● S. mutans group is the most nose agar)
commonly isolated viridans 3. Growth in Bile Esculin (BE)
streptococci medium
● S. bovis group and the enterococci ● reagent : esculin + 1% to 4%
possess the group D antigen. bile salt
● Bovis group is often isolated in blood ● (+) result: black color
culture of individuals with GI complex in the agar (both the
carcinoma. enterococci and S. bovis)
within 48 hours
Take note!
● DNA studies found that S. bovis and
S. equinus were the same species,
and the earlier name, S. equinus,
was adopted.

Enterococci  
● Natural inhabitants of the intestinal
tracts of humans and animals
● Are formerly known as Group D
Streptococci - all the species
produce the cell wall associated -
Group D antigen in the Lancefield
classification system
● They sometimes exhibit
pseudocatalase reaction - weak
bubbling in catalase test
● They can grow in extreme conditions
- alkaline pH, grown at 45C and salt
solutions.

Hemolytic patterns​: may be A, B, or

nonhemolytic
Virulence factors​: extracellular serine
protease, gelatinase and cytolysin (similar 4. 6.5% NaCl (nutrient broth base)
to bacteriocin) test
Species:​ E. faecalis (most common isolate) ● Result: (+)- turbidity -
E. faecium, E. avium, E. gallinarum enterococci
E. durans, E, raffinosus ● Group D and enterococci-
Related infections: UTI, endocarditis, use to grow in the 6.5 NaCl
bacteremia, and wound infection since enterococci can grow
in high salt concentration
Laboratory tests for Enterococci:  
Specimens: blood, urine, wound
secretions

1. Gram stain
2. Culture
● Culture media: TSH or BHI
with 5% Sheep’s blood
● Selective media: Bile Esculin
azide (BE Azide), Columbia
CNA agar, PEA agar
(​Phenylethyl alcohol agar),
CAA Agar
Notes to remember:
● Enterococci: (+) bile esculin, PYR,
and LAP tests
● (+) growth in 6.5% NaCl; penicillin
and vancomycin resistant
● All enterococci isolated from human
hydrolyze PYR

PYR Bile Esculin

S. pyogenes + -

Enterococci + +

Bile LAP 6.5% PYR

esculin NaCl

Enter + + + +
ococc
us

Grou + + - -
p D-
S.
bovis
group
● (+) growth in BE and 6.5% NaCl:
Leuconostoc, Pediococcus,
Globicatella, S. urinalis, and
Lactococcus