Medical MicrobiologyMedical Microbiology

Lecture 3Lecture 3

Dr. Saleh M Y OTHDr. Saleh M Y OTH

PhDPhDMedical Molecular Biotechnology and Infectious DiseasesMedical Molecular Biotechnology and Infectious Diseases

02/10/201002/10/2010IMS - MSUIMS - MSU

Systemic Bactreiology

STREPTOCOCCI Systemic Bactreiology

Words must to be known1.1. Group A streptococcus Group A streptococcus (S. pyogenes(S. pyogenes))2.2. Lancefield groupsLancefield groups3.3. Hemolysis (alpha, beta, gamma)Hemolysis (alpha, beta, gamma)4.4. Bacitracin susceptibility test Bacitracin susceptibility test 5.5. M, T, R proteinsM, T, R proteins6.6. Streptolysins O and SStreptolysins O and S7.7. F protein/lipoteichoic acidF protein/lipoteichoic acid8.8. Rheumatic fever/carditis/arthritisRheumatic fever/carditis/arthritis9.9. GlomerulonephritisGlomerulonephritis10.10. Scarlet feverScarlet fever11.11. Toxic shock-like Toxic shock-like

syndrome/bacteremiasyndrome/bacteremia12.12. ““Flesh-eating bacteria”Flesh-eating bacteria”13.13. Pyrogenic toxinPyrogenic toxin14.14. Erythrogenic toxinErythrogenic toxin

1. Group B streptococcus (S.agalactiae)

2. Neonatal septicemia/meningitis 3. CAMP test4. Hippurate hydrolysis test5. Group D streptococcus6. Urinary tract infection/ endocarditis 7. Bile-esculin test8. Enterococci 9. Non-enterococci10. Large colony11. Minute colony

12. Viridans streptococci13. Dental caries/endocarditis

Streptococcus pyogenesStreptococcus pyogenes

During the infection, this bacteria produce pus During the infection, this bacteria produce pus

- One of the most important pathogens- One of the most important pathogens- Gram positive cocci in chains- Gram positive cocci in chains- Lancefield Serological Group A- Lancefield Serological Group A- Beta Hemolytic on blood agar- Beta Hemolytic on blood agar

Gram Stain of S. pyogenes

Streptcoccus pyogenes S. agalactiaS. peumoniae

Hemolysis on Blood Agar PlatesHemolysis on Blood Agar Plates

Beta hemolysisBeta hemolysis-organisms excretes potent -organisms excretes potent hemoysins which completely lyse RBCs (complete hemoysins which completely lyse RBCs (complete hemolysis) thus a clear zone appears around hemolysis) thus a clear zone appears around colony. colony. S. pyogenesS. pyogenes

Alpha hemolysisAlpha hemolysis-organism excretes hemolysins -organism excretes hemolysins which partially break down rbc (incomplete which partially break down rbc (incomplete hemolysis) thus a greenish zone appears around hemolysis) thus a greenish zone appears around colony. colony. S. pneumoniaeS. pneumoniae

S.pyogenes S. pneumonia

Beta hemolysisAlpha hemolysis

StreptococciStreptococci – facultative anaerobefacultative anaerobe– Gram-positiveGram-positive– chains or pairs chains or pairs – catalase negativecatalase negative

(staphylococci are catalase positive)

Streptococcus in chains (Gram stain)

Streptococci • Lancefield groups

*one or more species per group*carbohydrate antigens

Groupable streptococci 1- A, B and D

frequent2- C, G, F

less frequent

Non-groupable

1- S. pneumoniaepneumonia

2- viridans streptococcie.g. S. mutans

* dental caries

hemolysis reaction - sheep blood agar

a- (beta-β) complete clearing A and B

b- (alpha-α)partial hemolysis

green color

c- (gamma-δ) no lysis

White colonies

Hemolysis1- Groups A an B

β-hemolysis

2- Group D α or δ-hemolysis

3- S. pneumoniae and S. viridans α-hemolysis

hemolysis reaction + one characteristic presumptive identification

Group A streptococcus (S. pyogenes)

Group A streptococcal infections affect all agespeak incidence at 5-15 years of age

S. pyogenes –suppurative

1- non-invasive - pharyngitis - skin infection, impetigo

2- invasive bacteremia - toxic shock-like syndrome - "flesh eating" bacteria- pyrogenic toxin

Pyrogenic toxinStreptococcus pyogenes and Staphylococcus aureus express pyrogenic toxin superantigens (PTSAgs) that are associated with toxic shock syndrome (TSS) and staphylococcal food poisoning (SFP).

Most PTSAgs cause TSS in deep-tissue infections, whereas only TSS toxin 1 (TSST-1) is associated with menstrual, vaginal TSS.

In contrast, SFP has been linked only with staphylococcal enterotoxins (SEs).

Pyrogenic toxin …Because of the differential abilities of PTSAgs to cause systemic or localized symptoms in a site-dependent manner, these toxins have abilities to cross mucosal barriers.

The activity of three PTSAgs when delivered orally, vaginally, or intravenously to rabbits and orally to monkeys in some experimental studies.

Pyrogenic toxin1- superantigen2- T cell mitogen 3- activates immune system

Scarlet fever

- rash - erythrogenic toxin

non-suppurativerheumatic fever

- inflammatory disease- life threatening- chronic sequalae

• fever • heart • joints

• rheumatic NOT rheumatoid arthritisRheumatic fever is an inflammatory disease that occurs following a Group A streptococcal infection, (such as strep throat or scarlet fever). Believed to be caused by antibody cross-reactivity that can involve the heart, joints, skin, and brain, the illness typically develops two to three weeks after a streptococcal infection. Acute rheumatic fever commonly appears in children between the ages of 5 and 15, with only 20% of first-time attacks occurring in adults. The illness is so named because of its similarity in presentation to rheumatism.

Rheumatic fever -etiology• M protein

– cross-reacts heart myosin – autoimmunity

• Cell wall antigens – poorly digested in vivo– persist indefinitely

Rheumatic fever• Penicillin

- terminates pharyngitis- decreases carditis

Acute glomerulonephritisimmune complex disease of kidney

Major pathogenesis factors lipoteichoic acid /F protein

fimbriaebinds to epithelial cells

M protein anti-phagocytic

S. pyogenes

fibronectin

lipoteichoic acid/F-protein

epithelial cells

M protein

M proteinM protein

fibrinogenfibrinogen

rrr

peptidoglycanpeptidoglycan

rrr

IgGIgG

Complement Complement IMMUNEIMMUNE

NON-IMMUNENON-IMMUNE

M proteinmajor target

natural immunitystrain variation

antigenicityre-infection

occurs with different strain

CapsulesAnti-phagocytic

mucoid strains

Isolation and identificationIsolation and identification β-hemolytic colonies

bacitracin inhibits growth

β-hemolytic colonies group A antigen

Bacitracin is a mixture of related cyclic polypeptides produced by organisms of the licheniformis group of Bacillus subtilis var Tracy, isolation of which was first reported in 1945.As a toxic and difficult-to-use antibiotic, bacitracin does not work well orally. However, it is very effective topically, and is a common ingredient of eye and skin antibiotic preparations. Its action is on gram-positive cell walls. It can cause contact dermititis due to allergic sensitivity to it.

ΒΒ--hemolysishemolysishemolysin O hemolysin O

sensitive oxygensensitive oxygenhemolysin Shemolysin S

insensitive oxygeninsensitive oxygen

Modern Rapid “Strep” TestModern Rapid “Strep” TestThroat swab extract Throat swab extract (+/- streptococcal antigen)(+/- streptococcal antigen)

AntibodyAntibody

Liposome

++

-

Streptococcal antigenStreptococcal antigen

Post-infectious diagnosis Post-infectious diagnosis (serology)(serology)

antibodies to streptolysin O antibodies to streptolysin O important if delayed clinical important if delayed clinical

sequelae occursequelae occur

Traditional serotyping of Traditional serotyping of proteins:proteins:- M - M - T - T - R- R

TypingTyping

Modern:Modern:- Sequencing of M protein gene- Sequencing of M protein gene

Group B streptococcusGroup B streptococcusneonatal meningitisneonatal meningitissepticemiasepticemia transmission transmission

vaginal floravaginal flora

Group B streptococcus Group B streptococcus - identification- identification

hemolysishemolysishippurate hydrolysishippurate hydrolysisCAMP reactionCAMP reaction

increases increases hemolysis of hemolysis of S. aureusS. aureus

Group D streptococcusGroup D streptococcus

Growth on bile esculin agarGrowth on bile esculin agarblack precipitate black precipitate

6.5% saline6.5% saline growgrow

enterococcienterococci no growth no growth

non-enterococcinon-enterococci

EnterococciEnterococci

distantly related to other streptococcidistantly related to other streptococci genus genus EnterococcusEnterococcus gut floragut flora

urinary tract infection urinary tract infection fecal contaminationfecal contamination

opportunistic infectionsopportunistic infectionsparticularly endocarditisparticularly endocarditis

most common most common E. (S.) faecalisE. (S.) faecalis

EnterococciEnterococci resistant to many antibiotics resistant to many antibiotics

including vancomycinincluding vancomycin terminal D-ala replaced by D-lactate terminal D-ala replaced by D-lactate

Minute colony streptococciMinute colony streptococciVarious groups/hemolysis (e.g. group A)Various groups/hemolysis (e.g. group A)

– genetically distinct genetically distinct * from large colony (e.g. S. pyogenes)from large colony (e.g. S. pyogenes)

– no rheumatic feverno rheumatic fever

Large colonyLarge colony Minute colonyMinute colony

Viridans streptococciViridans streptococci diverse species diverse species oral oral dental cariesdental caries hemolytic and negative for other tests hemolytic and negative for other tests non-groupable.non-groupable. includes includes S. mutansS. mutans

endocarditis endocarditis tooth extractiontooth extraction