Streptococcus

July, 2013

Outline
Introduction General features Classification Epidemiology Virulence and Pathogenesis Clinical manifestations/Diseases Laboratory Diagnostic tests Treatment Prevention and Control

Introduction
Gram positive in reaction Forms pairs / chains during growth Found everywhere Contain normal flora & pathogenic Spp. Produce extra cellular substances & enzymes Gram + ve cocci in pair or chain, non motile, non spore forming Catalase negative

General Features
Produce variety of infections, ranging from pharyngitis, cellulites to sepsis Includes more than 50 species Clinically important genera include Streptococcus and Enterococcus Most are facultative anaerobes & some are obligate anaerobes

Classification
Classified based on Hemolytic capacity on BA Alpha ()- incomplete, green hemolysis Beta ()- clear, complete lysis of red cells Gamma()- no hemolysis The antigenicity of a carbohydrate occurring in their cell walls (A-V) (Lancefield antigen) classified by Rebecca Lancefield Capsular antigens: S. pneumonia into 84 types Biochemical Tests: Sugar fermentation rxns Tests for the presence of enzymes Tests for susceptibility/resistance to certain ABCs

Lysis of red blood cells caused by specific bacterial enzymes

Partial lysis of RBCs green Alpha discoloration of agar around colonies Complete lysis of RBCs clearing of Beta agar around colonies (Gamma) No lysis of RBCs No change in agar around colonies

Classification contd
Species S.pyogenes Hemolysis Group Antigen A Common Terms Group A streptococci Group B streptococci Disease Association(s) Pharyngitis; scarlet fever pyoderma; rheumatic fever; AGN Neonatal sepsis; puerperal fever; pyogenic infections; pneumonia; meningitis Pharyngitis; impetigo; pyogenic infections Urinary tract infections Wound infections Bacteremia; Endocarditis Urinary tract; pyogenic infections; Endocarditis infections Bacteremia; pneumonia; meningitis; Endocarditis S.agalactiae B

S. equisimilis E. faecalis E. faecium E. durans S. bovis S. equinus

Alpha or no hemolysis ( rarely ) Alpha ()or none (rarely ) Alpha () hemolysis Alpha () hemolysis or no hemolysis

C D

Group C streptococci Enterococci

Nonenterococci

S. pneumoniae

Pneumococcus

Viridans and Nonhemolytic S. sanguis S. salivarius S. mitis or nonhemolytic S. milleri S. mutans Other species

Viridans strep

Dental caries

Streptococcus : Cell Wall Structure

Thick peptidoglycan layer Teichoic acid C=carbohydrate layer present except in viridans group Capsule in S. pneumoniae and in young cultures of most species

Sterptococcus Pyogenes
Pyogenes means pus producing One of the most important pathogens Gram positive cocci in chains Lancefield Serological Group A Beta Hemolytic on blood agar Some strains produce capsule and pathogenic strain contain M protein (attachment factor, antigenic and anti-phagocytic The most pathogenic member of the genus Produces a large number of powerful enzymes and toxins. Present as a commensal in the nasopharynx of healthy adults, and more commonly in children (10% carriage)

Virulence and pathogenesis

Virulence Factors Group specific polysaccharide antigen (C carbohydrate) Type specific protein antigen M- protein- major virulence factor T & R protein- not related to virulene Exotoxin Pyrogenic exotoxins A - C (Erythrogenic toxin) Hemolysin Streptolysin O oxygen labile Causes Beta hemolysis Antigenic- for ASO testing Streptolysin s Also responsible for Beta hemolysis Not Antigenic Capsular hyaluronic acid

Enzymes
Streptokinase (Fibrinolysin) Hyaluronidase Diphosphopyridine Nucleotidase (DNAase) Nicotineamid Adenine Dinucleotidase (NADase) Anti-C5a peptidase

Pathogenesis
Causes disease by three main mechanisms: 1. Inflammation Tonsillitis, pharyngitis, cellulites, otitis media, etc Impetigo, Erysipelas, Cellulitis The enzymes contribute for the invasiveness includes: Hyaluronidase- spreading factor Streptokinase- dissolves fibrin in clots. DNase - depolymerizes DNA in exudates or necrotic tissue

Pathog cont
2. Exotoxin and hemolysin production Erythrogenic toxin Streptolysin O Streptolysin S Pyogenic exotoxin A toxin 3. Immunologic Is due to the inflammation caused by immunologic response to streptococcal M proteins that cross react with human tissue Rheumatic fever: is due to cross-reaction between antibody & human heart & joint tissue; occurs after 2 weeks of pharyngitis Acute glomerulonephritis (AGN): caused by immune complexes bound to glomeruli; occurs 2 3 weeks skin or respiratory infection

Clinical Manifestations
Diseases caused 1. Strep throat (Streptococcal pharyngitis) Most common of all Spread by saliva or nasal secretions Incubation period 2-4 days Sore throat, slight fever (101) Important to treat immediately to avoid post strep diseases If the strain of S. pyogenes is lysogenic for a particular phage which expresses an erythrogenic toxin the result is Scarlet fever Rash appears and characteristic is the strawberry colored

Clinical Manifestations
2. Streptococcal skin infections Impetigo Folliculitis Erysipelas Cellulitis- could be life treatening 3. Invasive Strep A infections Necrotizing fasciitis- infection along the fascia Scarlet Fever Myositis:- resembles clostridial gangrene Toxic shock-like syndrome (STSS):-Multi-organ system failure

Clinical Manifestations
4. Delayed Antibody mediated Disease Rheumatic fever- involving heart valves, joints, nervous system Follows a strep throat By antibody cross reactivity between the cell wall of S. pyogenes and heart muscle Glomerulonephritis(Brights Disease) inflamatory disease of renal glomeruli and structures involved in blood filter of kidney Due to deposition of Ag/Ab complexes Symptoms include fever, malaise, edema, hypertension and blood or protein in urine

Erysipelas

Diseases caused by S.pyogenes

Laboratory Diagnosis
Specimen- throat swab, pus, blood Grams rxn - gram positive cocci in chains Culture- grow aerobic or anaerobically at temp 35- 37% Do not grow on MacConkey agar Shows clear zone of hemolysis on blood Agar Biochemical Test and Sensitivity Test Catalase-negative Bacitracin-susceptible PYR-positive Bile-esculinnegative 6.5% NaCl-negative

S.Pyogens Gramstain and culture chxs

S. Pyogens Gram rxn and morphology

S. Pyogens hemolysis on BA

Group A streptococcus is susceptible to Bacitracin disk (left)

S. Pyogens growing in the presence of SXT

Group B beta-Hemolytic Streptococci (S. agalactiae)

S. agalactiae is found in the vaginocervical tract of female carriers, and the urethral mucous membranes of male carriers, as well as in the GI tract A major cause of bovine mastitis They are also an occasional cause of infections in postpartum women (endometritis) and individuals with impaired immune systems, in whom the organism may cause septicemia or pneumonia Transmission occurs from an infected mother to her infant at birth, and venereally (propagated by sexual contact) among adults

S. agalactiae cont
Transmission occurs from an infected mother to her infant at birth, and venereally among adults Samples of blood, cervical swabs, sputum or spinal fluid can be obtained for culture on blood agar Group B streptococci are -hemolytic with larger colonies and less hemolysis than group A. Most isolates remain sensitive to penicillin G and ampicillin, which are still the antibiotics of choice

Laboratory Diagnosis
Specimens:
Cerebrospinal fluid Ear swab Blood for culture from neonates. High vaginal swab is required from women with suspected sepsis.

Group B streptococci are Gram positive cocci, occurring characteristically in short chains but also in pairs and singly The organisms are non-motile. Most strains are capsulated. Blood agar: Most strains of S. agalactiae produce grey mucoid colonies about 2 mm in diameter, surrounded by a small zone of beta haemolysis. About 5% of strains are nonhaemolytic Placing discs of penicillin and gentamicin on the plate can help to identify these strains (penicillin sensitive, gentamicin resistant).

Lab Diagn cont

MacConkey agar: Most strains grow on this medium Neomycin blood agar: A useful selective medium for isolating S. agalactiae from urogenital specimens CAMP (Christie, Atkins, Munch, Peterson) test to identify presumptively S. Agalactiae
This test requires the use of a beta-lysin producing strain of S. aureus (NCTC 1803 or ATCC 25923) to detect the CAMP factor, i.e. extracellular diffusible protein produced by S. agalactiae. This protein interacts with the staphylococcal beta-lysin on sheep (or ox) blood agar producing enhanced haemolysis

CAMP test

Group B streptococci showing the classical arrowshaped hemolysis near the staphylococcus streak

Lab Diagn cont

Direct detection of Group B streptococcal antigen in CSF When Group B streptococcal meningitis is suspected, a rapid diagnosis can be made by detecting Group B streptococcal antigen directly in CSF serum or urine using a latex or coagglation slide test. Direct tests are expensive. They are particularly useful if antibiotic treatment has been started and it is not possible to isolate S. agalactiae culturally Treatment S. agalactiae has the same susceptibility profile as S. pyogenes

Group D streptococci
Streptococcus bovis is the most clinically important of the nonenterococcus group D streptococci Part of normal fecal flora, they are either or nonhemolytic S. bovis occasionally causes
Urinary tract infections and Subacute bacterial endocarditis (especially in association with colon cancer).

Is bile- and esculin-positive, but is PYR-negative, and does not grow in 6.5 percent salt Tends to be sensitive to penicillin and other antibiotics

Identification of Hemolytic Colonies

Groups C & G
Occur in nasopharynx Cause sinusitis, bacteremia, or endocarditis hemolytic on blood agar Identified by rxns with specific antisera for Grps C & G

viridans streptococci
Although often alpha-haemolytic on blood agar, the viridans group of streptococci can also be nonhaemolytic and occasionally beta-haemolytic A few species are pathogenic (e.g. S. mutans, S. sanguis, S. mitior) causing endocarditis, bacteraemia, and dental caries Normal human GI tract flora Nasophrynx Cause Dental Infection Endocarditis Abscesses

viridans streptococci cont

The following are the main features which differentiate S. pneumoniae from viridans streptococci:

Enterococcus species
E. faecalis (formerly classified Streptococcus. faecalis) is the main pathogen in the genus Enterococcus, causing about 95% of enterococcal infections Infections include- urinary tract, biliary tract, ulcers (e.g. bed sores), wounds (particularly abdominal) and occasionally endocarditis or meningitis Normal commensal of the vagina and intestinal tract. A minority of infections are caused by E. faecium.

Lab Diagnosis
Enterococcus species are Gram positive cocci, occurring in pairs or short chains Are non-capsulate and the majority are non-motile Enterococci are aerobic organisms capable of growing over a wide temperature range, 1045 C Are mainly nonhaemolytic but some strains show alpha or beta-haemolysis E. faecalis ferments lactose, producing small dark-red magenta colonies on MacConkey agar and small yellow colonies on CLED (cysteine lactose electrolyte-deficient) agar

Lab Diagn cont

Catalase negative Are also able to grow in the presence of 6.5% sodium chloride and 40% bile. Ferment lactose (also mannitol and other sugars) Reduce litmus milk Treatment Most enterococci are susceptible to ampicillin and resistant to cephalosporins Resistance is shown against penicillin.

Streptococcus Pneumoniae (Pneumococcus)

S. pneumoniae are gram-positive, nonmotile, encapsulated cocci Often part of normal flora of respiratory track , becomes infective once hosts resistance is lowered. Classified as an endogenous infection In tissue, pus or sputum are typically arranged in pairs (diplococci) each coccus some what elongated and pointed at one end but rounded at the other (lanceolate) and the two members of a pair point away from each other surrounded by a polysaccharide capsule Serotypes: Over 80 capsular serotypes Less than 15 serotypes responsible for infections

Lancet shape

Pathogenesis
Primary virulence factor is the capsular polysaccharide which protects the organism against phagocytosis Pathogenesis is due to rapid growth of bacteria in alveolar spaces Symptoms Onset abrupt Chills Chest pains Labored breathing

Predisposing factors: upper respiratory viral infection, diabetes, alcoholism

Pathogenesis cont
S. pneumoniae causes lobar pneumonia, bronchitis (often with H. influenzae), meningitis bacteraemia otitis media, sinusitis and conjunctivitis Severe infections can occur in the elderly and those already in poor health, HIV or immunosuppressed. Risk of infection is increased following splenectomy Also a common cause of childhood pneumonia and serious infections in patients with sickle cell disease Pneumococci form part of the normal microbial flora of the upper respiratory tract

Laboratory Diagnosis
Diagnosis Chest Xray Culture and staining Biochemical tests Specimens -nasopharyngeal swab, blood, pus, sputum, or spinal fluid. S.pneumonea resembles the streptococci already described on its nutritional and environmental requirements except its growth is facilitated in 5-10%CO2 atmosphere. -Hemolytic on blood agar overnight under aerobic conditions at 37C Lancet-shaped, gram-positive diplococci . Capsular swelling is observed when the pneumococci are treated with type-specific antisera (the Quellung reaction).

Optochin sensitivity
Pneumococci are sensitive to optochin (ethylhydrocupreine hydrochloride). Placing a disc (5g) on a primary sputum culture and culturing the plate aerobically can help to provide a rapid presumptive identification of S. Pneumoniae The zone of inhibition should be at least 10 mm. Most viridans streptococci and other alpha haemolytic streptococci are resistant to optochin. If the zone of inhibition is less than 10 mm the colonies should be tested for bile solubility Others Rapid latex and coagglutination tests are available to detect capsular pneumococcal antigen in CSF, pleural fluid, serum and urine

Hemolytic, Biochemical, and Cultural Reactions of Common Streptococci and Enterococcia

Treatment of S.pneumonia
Early treatment usually results in rapid recovery Penicillin G is the drug of choice penicillin, erythromycin, co-trimoxazole Some penicillin-resistant strains are resistant to cefotaxime. Resistance to tetracycline and erythromycin occurs also Pneumococci remain susceptible to vancomycin. It is possible to immunize individuals with type-specific polysaccharides. Such vaccines can probably provide 90% protection against bacteremic pneumonia. Seven-valent vaccine is recommended for all children aged 2 23 months, to help prevent ear infections, and for selected children aged 2459 months.

Summary