General Microbiology LaboratoryGeneral Microbiology Laboratory

Isolation and Identification of Gram Isolation and Identification of Gram Positive CocciPositive Cocci

Mohammed laqqan

Identified method for Identified method for StaphylococciStaphylococci

Gram-stain Isolation and culture Pure culture

Direct identification

Mohammed laqqan

Staphylococci are Gram-positive cocci typically arranged in clumps or Grape-like clusters

Mohammed laqqan

Mohammed laqqan

DirectDirect identifidentificationication

The mannitol fermentation test The Coagulase Test The DNase Test Novobiocin (NB) disc

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

Identified method for Identified method for StaphylococciStaphylococci

Gram-stain Isolation and culture Pure culture

Direct identification

Mohammed laqqan

Staphylococci are Gram-positive cocci typically arranged in clumps or Grape-like clusters

Mohammed laqqan

Mohammed laqqan

DirectDirect identifidentificationication

The mannitol fermentation test The Coagulase Test The DNase Test Novobiocin (NB) disc

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

Staphylococci are Gram-positive cocci typically arranged in clumps or Grape-like clusters

Mohammed laqqan

Mohammed laqqan

DirectDirect identifidentificationication

The mannitol fermentation test The Coagulase Test The DNase Test Novobiocin (NB) disc

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

Mohammed laqqan

DirectDirect identifidentificationication

The mannitol fermentation test The Coagulase Test The DNase Test Novobiocin (NB) disc

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

DirectDirect identifidentificationication

The mannitol fermentation test The Coagulase Test The DNase Test Novobiocin (NB) disc

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

The mannitol fermentation testThe mannitol fermentation test

Inoculate the bacteria into a mannitol micro-tubeincubate at 370C for 18hSaureus will ferment mannitol to produce acidwhich causes the medium to turn yellow

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

TestStaphylococcus aureusStaphylococcus epidermidis

Staphylococcus saprophyticus

Mannitol fermentationPositiveNegativeUsually positive

Mohammed laqqan

The Coagulase TestThe Coagulase Test

Coagulase is an enzyme converting fibrinogen into fibrin promoting blood clotting

It might be a virulence factor with the coagulated blood around the bacteria protecting them from the immune system

Coagulase-negative strains are often as pathogenic as coagulase-positive strains

Mohammed laqqan

The DNase TestThe DNase Test

Inoculate DNase agar plates with a loop so that the growth is in plaques about 1 cm in diameterI

ncubate at 370C overnightFlood the plate with 1 N hydrochloric acidClearing

around the colonies indicates DNase activityThe hydrochloric acid reacts with unchanged

deoxyribonucleic acid to give a cloudy precipitateA few other bacteriaeg Serratiamay give a positive

reaction

Mohammed laqqan

Staphylococcus aureus Growing on DNase AgarStaphylococcus aureus Growing on DNase Agar

Mohammed laqqan

Blood agar with a novobiocin (NB) discBlood agar with a novobiocin (NB) disc

Staphylococcus aureus Growing on Blood Agar Staphylococcus saprophyticus

Growing on Blood Agar

Mohammed laqqan

Identification of Gram-positive cocciIdentification of Gram-positive cocci

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

The DNase TestThe DNase Test

Inoculate DNase agar plates with a loop so that the growth is in plaques about 1 cm in diameterI

ncubate at 370C overnightFlood the plate with 1 N hydrochloric acidClearing

around the colonies indicates DNase activityThe hydrochloric acid reacts with unchanged

deoxyribonucleic acid to give a cloudy precipitateA few other bacteriaeg Serratiamay give a positive

reaction

Mohammed laqqan

Staphylococcus aureus Growing on DNase AgarStaphylococcus aureus Growing on DNase Agar

Mohammed laqqan

Blood agar with a novobiocin (NB) discBlood agar with a novobiocin (NB) disc

Staphylococcus aureus Growing on Blood Agar Staphylococcus saprophyticus

Growing on Blood Agar

Mohammed laqqan

Identification of Gram-positive cocciIdentification of Gram-positive cocci

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Staphylococcus aureus Growing on DNase AgarStaphylococcus aureus Growing on DNase Agar

Mohammed laqqan

Blood agar with a novobiocin (NB) discBlood agar with a novobiocin (NB) disc

Staphylococcus aureus Growing on Blood Agar Staphylococcus saprophyticus

Growing on Blood Agar

Mohammed laqqan

Identification of Gram-positive cocciIdentification of Gram-positive cocci

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Blood agar with a novobiocin (NB) discBlood agar with a novobiocin (NB) disc

Staphylococcus aureus Growing on Blood Agar Staphylococcus saprophyticus

Growing on Blood Agar

Mohammed laqqan

Identification of Gram-positive cocciIdentification of Gram-positive cocci

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Identification of Gram-positive cocciIdentification of Gram-positive cocci

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

types Of Streptococcus types Of Streptococcus

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

β haemolytic steps Lancefield groupsβ haemolytic steps Lancefield groups

Historically A B C F and G group specific antigens cell wall polysaccharide group D and Enterococci lipoteichoic acid moiety Not always β haemolyticNow 18 different groups of strep from A-H and from K-T

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

(β) Haemolytic Streptococcus groups(β) Haemolytic Streptococcus groups

S pyogenesbull Group A S agalactiaebull Group B (occasionally α or none) Streptococcus equismilisbull Group C (occasionally α or none) Streptococcus bovisbull Group D (α or none occasionally β) Streptococcus anginosusbull Group F amp G

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Gp A strep S pyogenes Laboratory IDGp A strep S pyogenes Laboratory ID

Gram +ve Catalase Haemolysis small round grey colonies β haemolysis zone is large Gp A antigen

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)Isolation and Identification of group A beta hemolytic (Streptococcus pyogenes)

1 on Blood agar Streptococcus pyogenes produces a zone of beta hemolysis around 2-3mm in diameter surrounding each colony

2 Sensitivity to the antibiotic bacitracin

3 Group A Strep is senstive other b haemolytic streps are resistant

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Bacitracin sensitivityBacitracin sensitivity

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Group B S agalactiaeGroup B S agalactiae

S agalactiae

Identification Catalase negative Appearance on BA Larger colonies than Gp A Small zone of hemolysis Latex agglutination Gp B Bacitracin resistant CAMP positive Growth on Mac (weak)

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Presumptive ID Tests - CAMPPresumptive ID Tests - CAMP

The ldquoCAMPrdquo test (acronym for developers) is used to differentiate S agalactiae (GBS) which is positive from other beta hemolytic strep all of which are negative

ldquoCAMP factorrdquo is a soluble hemolysin produced by GBS that combines in a synergistic way with a similar hemolysin of S aureus to form an arrowhead zone of clearing See image

The staph is streaked perpendicularly to the unknown strep After appropriate incubation time an ldquoarrowhead-shapedrdquo clear zone of hemolysis will appear

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

S agalactiaeS agalactiae

Synergistic haemolysisobserved between betahemolyticStaphylococcusaureus and group Bstreptococci1048698 Positive reaction arrowhead haemolysis pattern

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

α ndash Haemolytic streps Viridans strepsα ndash Haemolytic streps Viridans streps

Not groupable by Lancefield groups No group specific CHO Streptococcus pneumoniae Primary human pathogenSsanguis Smitis Smutans Ssalivarius

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

The Pneumococcus (Streptococcus pneumoniae)The Pneumococcus (Streptococcus pneumoniae)

1 Isolation on Blood agar bull Pneumococci frequently require enriched media and increased

CO2 tension for initial isolation bull They are usually isolated on Blood agar and incubated in a

candle jar (a closed container in which a lit candle is placed to remove O2 and increase CO2 ) at 37C On Blood agar colonies appear small shiny and translucent

bull They are surrounded by a zone of alpha hemolysis 2 Optochin sensitivity bull Pneumococci are the only streptococci that are sensitive to the

drug optochin(ethylhydrocupreine hydrochloride) 3 Gram stain gram-positive diplococci

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

The Genus EnterococcusThe Genus Enterococcus

Enterococci are gram-positive streptococci typically occurring in pairs and short chains that are normal flora of the intestinal tract Enterococci responsible for a variety of opportunistic infections in humans and serologically belong to Lancefield group D streptococci

On Bile Esculin agar Unlike most bacteria the enterococci will grow in the

presence of the bile salts in the medium They hydrolyze the esculin producing esculetin

which reacts with the iron salts in the medium turning the agar black

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

EnterococcusEnterococcus

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

Laboratory ID EnterococcusLaboratory ID Enterococcus

white colonies alpha and no haemolysis (rarely beta) Also grow on MacConkey Resembles S pneumoniae in Gram stains Bile esculin positive Grows on Mac

Mohammed laqqan

End of lecture

Mohammed laqqan

End of lecture