Genus Streptococcus Gram positive, Catalase negative - PowerPoint Presentation

Slide1

Genus Streptococcus

Gram positive, Catalase negative cocci

Practical

No. 9

Slide2

GENERAL CHARACTERISTIC:*

G+ve cocci , arrange in chains or pairs.* Some strains are capsulated* Majority are facultative anaerobic, few are obligatory anaerobic.* Catalase –ve

* Non motile.* Non spore forming* Fastidious microorganism

Slide3

Members of the genus

Streptococcus are responsible for disease as well as being part of the

normal flora of humans. Among the diseases caused are:bacterial pneumonia

meningitis

Tonsillitis

Endocarditis

scarlet fever

Erysipelas

urinary tract infections.

Streptococcus

species are also

found

normally in the

mouth

and on the

skin surface

.

The streptococci are classified by

two major methods

:

1. hemolytic

activity

2. serologic

classification of Lancefield.

Slide4

Slide5

When grown on sheep blood agar, streptococci display one

of three types of hemolysis of the red blood cells in the agar.

Alpha hemolysis--The red blood cells in

the media are partially digested producing a greening of the

agar.

Beta

hemolysis-

-The red blood cells in the media are completely digested producing a clearing of the

agar.

Gamma

hemolysis-

-No change is noted in the agar. The red blood cells are not lysed.

Streptococcus

pyogenes

always beta

hemolylic

Streptococcus

agalactiae

usually beta hemolytic

Streptococcus

pneumoniae

and

Viridans

streptococci

are always alpha

hemolylic

Slide6

Alpha hemolysis

on

blood

agar

(partial hemolysis).

Beta hemolysis

on blood agar

(complete hemolysis).

No hemolysis on blood agar

Hemolytic Streptococci

Alpha hemolysis (green discoloration)

(Beta hemolysis)

clear zone around colonies

Slide7

DISEASES S. pyogenes

Slide8

Lab dx.Specimens: sputum, throat swab, nasopharyngeal swab, blood, CSF…etc.

Gram stain: G+ve cocci, arrange in chains.Culture: on blood agar

pinpointed, Grayish white, translucent, matte or glossy colonies with large zone of β- hemolysis.

S.pyogenes on blood agar (beta hemolysis)

Slide9

Bacitracin Susceptibility tests

Commercially available paper disks saturated with a solution containing Bacitracin will inhibit about 97% of all strains of Group A streptococci; other groups of beta-hemolytic streptococci will not be affected.

Streptococcus pyogenes

group

A is inhibited

by

small amount of

bacitracin in the disk other B hemolytic streptococci usually are not.Streptococcus pyogenes Strept. pyogenes Strept. agalactiae

Slide10

  A blood test that checks for a strep infection

. When you come into contact with harmful bacteria, your body produces antibodies to defend itself against these bacteria. Your body produces antibodies specific to the bacteria they fight.The

antistreptolysin O (ASO) titer testThe

ASO titer test

measures antibodies produced by your body in response to a toxin known as

streptolysin

O.

Streptolysin

O is a toxin produced by group A 

Streptococcus (GAS) bacteria. Your body makes the antistreptolysin O antibodies when you have a strep infection caused by GAS bacteria.

Slide11

A positive test usually is >200 units/mL, but normal ranges vary from laboratory to laboratory and by age. Normal

levels is

less than 160 unit per

milliliter.

Slide12

S. agalactia

(Group B β- hemolytic):

Slide13

Identification of group B streptococci. (S. agalactiae

)The CAMP Test

The laboratory identification of Group B hemolytic streptococci .Streptococcus agalactiae

produce

CAMP factor (a diffusible extracellular protein) that synergistically act with the beta –

lysin

of

Staphylococcus

aureus and enhances the lysis of RBC.The CAMP test is performed by making a single streak of the streptococcus perpendicular to a strain of Staphylococcus aureus that is known to produce ß - l

ysin. The two streak lines must not touch one another. The inoculated plate must be incubated with room temperature. Any Bacitracin-negative, CAMP-positive, bile-

esculin

-negative streptococcus can be reported as: Group B streptococcus

CAMP (Christie, Atkins and Munch – Peterson) test

Slide14

CAMP test

Slide15

The CAMP Test

Slide16

The Hippurate test:

Is a qualitative procedure for determining the ability of bacteria to enzymatically hydrolyze sodium

hippurate into glycine and benzoic acid by action of hippuricase enzyme present in

bacteria

.

An

oxidizing agent

ninhydrin

is used as an indicator

.Ninhydrin react with glycine to form a deep blue or purple color.

Slide17

Slide18

Slide19

Non-beta hemolytic (alpha- and non hemolytic) streptococci

The non-beta hemolytic streptococci include the S.

pneumoniae, Viridans

streptococci

, Group D streptococci and enterococci.

Identification of group D Streptococci:

Lancefield Group D streptococci are divided into two groups

:

(1)

Enterococci (2) Non enterococci. Enterococcus

faecalis and Enterococcus faecium are the species of Enterococcus

.

Strep

.

bovis

and

Strep.

equinus

are species of "Group-D streptococci, not Enterococci"

Slide20

Bile-Esculin Hydrolysis Test:

The purpose of this test is to determine the ability of an organism to hydrolyze the glycoside esculin to

esculatin and glucose in the presence of bile (10 - 40%). This test aids in the differentiation of group D streptococci from other "not group D streptococci". Procedure:

1. Inoculate the organism to be tested into the bile

esculin

medium. Incubate at 37

o

C for 24

hours.

Positive

Test: Presence of a black to dark brown color on the slant -(Enterococcus faecalis) Negative Test: No blackening of the medium - (Streptococcus agalactiae or Streptococcus pyogenes)

Slide21

Bile

Esculin

hydrolysis

Ability to grow in 40% bile and hydrolyze

Esculin

are features of streptococci that possess Group D antigen.

The

organism can hydrolyze

esculin

and produce black color in the media.

Bile-Esculin Hydrolysis Test:

Slide22

Viridans streptococcus

Slide23

S. pneumoniae

(Pneumococci,

Diplococcus pneumoniae)

Capsular polysaccharide

Gram +VE diplococcus

Slide24

S.

pneumoniae

Slide25

Identification of

S. pneumonia and viridans

streptococci The alpha-hemolytic streptococci, or viridans

(

green

)

group, are

normal inhabitants of the

mouth,

nasopharynx

, and respiratory tract. Alpha-hemolytic streptococci may be isolated by blood culture in cases of bacterial endocarditis. Specimen sputum, blood, throat swab, ear swab, CSF

Slide26

1-Gram stain of Streptococcus

pneumoniae and

viridans Streptococci: Gram stain of

S.

pneumoniae

and a

viridans

streptococcus obtained from the

blood

agar

plate.

Capsular polysaccharide

Gram +VE

diplococcus

Strept

.

pneumoniae

gram-positive

diplococci

. Notice the characteristic elongated shape of the

diplococci

Slide27

2-The

Quellung reaction to differentiate S. pneumoniae

from viridans streptococci.

The

Quellung

reaction

(capsular swelling reaction)

is a

biochemical

reaction in which antibodies bind to the capsule of Streptococcus pneumoniae, Klebsiella

pneumoniae, Neisseria meningitidis and

Haemophilus

influenzae

and thus allow them to be visualized under a microscope

.

If the reaction is positive, the capsule becomes

opaque

and appears to enlarge

.

Slide28

Optochin disc for S. pneumococci

3. Susceptibility test

Optochin

“P”

disk

Differentiates

S.

pneumoniae

from other

alpha-hemolytic streptococci (

Viridans

group

)

S. pneumococci

Slide29

To determine

the ability of bacterial cells to lyse in the presence of bile salts.

4.  Bile solubility test 

The 

test

 is primarily used to

differentiate 

bile

soluble

 

Streptococcus pneumoniae from bile insoluble alpha- hemolytic streptococci.

Slide30

5-Inulin fermentation test

Pneumococci

there will be a change in the color of the media due to acid formation.

Procedure

:

Inoculate a test tube containing nutrient broth + inulin + the organism, (red

color).

Then incubate over night, if the

color

remains red the organism is

Viridans

Streptococcus, If the organism has fermented the inulin resulting in acid production. The acid lowers the pH of the media and the indicator (phenol red) turns yellow.

Slide31

Observe for the presence of a

yellow

color (center tube), indicative of acid formation from the fermentation of inulin.

Red

indicates no inulin fermentation (left tube; right tube is

uninoculated

control).

Inulin

fermentation test

Slide32

CHARACTER

PNEUMOCOCCI

VIRIDANS STREPTOCOCCIMorphology

Ovoid or

lanceolate

diplococci

Rounded

cocci

in short or long chains.

Capsule

PresentAbsentOptochin sensitivity +ve-veBile solubility

+ve-veCapsular swelling test (Quelling reaction

)

+ve

-ve

Virulence in mice

+

ve

-

ve

Differentiation of alpha and non-hemolytic streptococci

Slide33

Hemolysis Aids in Classification

Slide34

Thank you