Corynebacterium.ppt 3se2q

Definition • Diphtheria is an acute, toxinmediated disease caused by toxigenic Corynebacterium diphtheria. • It’s a very contagious and potentially life-threatening bacterial disease.

Definition • It’s a localized infectious disease, which usually attacks the throat and nose mucous membrane

Etiology • C. diphtheriae is an aerobic grampositive bacillus. – Pleomorphic, club-end – Non-spore-forming – Non-acid-fast – Non-motile

Etiology • The major virulence determinant is an exotoxin, diphtheria toxin. After binding to the host cells, the active subunit will interrupt the protein synthesis of the target host cell and results in cell death. • Toxoid made from diphtheria toxin can be used as vaccine.

Etiology • There are three biotypes — gravis, Intermedius, and mitis. The most severe clinical type of this disease is associated with the gravis biotype, but any strain may produce toxin.

Pathogenesis • Entry ------ the bacilli multiply locally in the throat and elaborate a powerful exotoxin ----produce local and systemic symptoms. Local lesions : • Exotoxin causes necrosis of the epithelial cells and liberates serous and fibrinous material which forms a grayish white pseudo membrane • The membrane bleeds on being dislodged • Surrounding tissue is inflamed

Typical Presentation of Bull Neck

Local manifestation Depend on the site of lesion: Nasal diphtheria : • Unilateral or bilateral serosanguineous ( blood and serous fluid ) discharge from the nose • Excoriation of upper lip • Toxemia is minimal

Faucial diphtheria : • Exudates form white pseudo membrane • Regional lymph nodes are inflamed • Sore throat and • dysphagia

Corynebacterium diphtheria • • • • •

Slender rods Clubbing at both ends Pleomorphic Non capsulate / Gram + Granules are composed of polymetapohosphate • Staining with Loffler's methylene blue show bluish purple metachromatic granules. with polar bodies,

Staining methods • Grams method • • • •

Albert's stain Neissers stain Ponders stain On staining seen as Pairs, Appear as v and L letters, resembling Chinese letter pattern or also called cuneiform arrangement.

Cultural characters • • • • • • •

Need enrichment Media Contain Blood, Serum or Egg 37 c ph 7.4 Aerobic/Facultative anaerobic. Commonly used medium Loeffler serum, Tellurite Blood agar,

Colonies

Culturing

• Selective & differential medium • Corynebacterium are resistant to tellurite – Reduced to tellurium

• Forms deposit in colonies – Colonies appear dark

• Biotypes – gravis, Intermedius, mitis

Biochemical Reactions • Acid

Glucose,Galactose Maltose, Dextrin Do not produce acid with Lactose, Mannitol, sucrose. All fermentation reactions tested in Hiss serum sugars Urease test negative. Proteolytic

Toxin • Pathogenicity associated with Toxin • Gravis/Intermedius 95-99% are toxigenic • Mitis 80 – 85% • Some abundant others poorly • Toxin M W 62,000 0.0001 can kill guinea pig

Toxin ( Contd ) • Toxin contain two components A 24,000 B 38,000 A produce toxigenicity by proteolytic effect B Produce binding Toxin + Formalin = Toxoid What is Toxoid – Antigenic, not toxigenic Tox + Corynephage Toxin production

Resistance • Can be killed at 580 c in 10 mt 1000 c in 1 mt Survive in Blankets, Floor dust, toys

Clinical features • • • • • • •

Malaise, Sore throat, Fever Adherent grey pseudo membrane Nasal ulcers, Obstruction of larynx and lower airways, Difficulty in swallowing Lead to Myocarditis, Peripheral neuritis, Paralysis of limbs,

Diphtheria Clinical Features Incubation period 2-5 days May involve any mucous membrane Classified based on site of infection anterior nasal pharyngeal and tonsillar laryngeal cutaneous ocular genital

Thick Membrane

Pseudo membrane

Skin Lesions

Complications • Asphyxia - causing mechanical obstruction. • May need tracheotomy • Circulatory failure. • Post Diphtheria paralysis

Non toxigenic clinical manifestations • Bacteria can produce 1. Endocarditis, 2.Meingitis, 3 Cerebral abscess. 4 Osteoarthritis.

Laboratory Diagnosis • Specific treatment is more important than Laboratory Diagnosis. 1 Isolation of Diphtheria bacilli. 2.Testing for toxigenicity,

Collection of Specimens • Throat swabs • Smear examinations Gram s staining, Albert's, Ponders Immunoflorescent methods Cultures on Loeffers serum slope Tellurite Blood agar, Blood agar.

Isolation of C.diptheria • • • •

Serum slope – Growth in 6 – 8 hours, Stain with Neissers stain Albert's stain Bacilli have metachromatic granules, Tellurite Blood agar takes two days for manifestation of colonies,

Virulence tests, • In Vivo and In Vitro • In Vivo in Animals • Subcutaneous tests Inject broth from culture into two Guinea pigs, 0.8 ml One animal given 500 units of antitoxin Other no Vaccine. Animal not given antitoxin will die Loss of Animals. Restricts its testing.

Intracutaneous Method • One animal given 500 units before toxin • Other 50 units after Toxin • So the Animals can be saved

In Vitro Testing • Elek s Gel precipitation testing • Filter paper impregnated with Diphtheria antitoxin 1000 Units / ml • Tested on the horse serum agar • Positive / Negative /Test strains tested for Immunodiffusion • Line of precipitation – test positive • Other methods testing in Tissue cultures.

Toxigenicity Tests In Vitro Elek test In Vivo Animal inoculation rabbit skin testnecrosis guinea pig challenge testlethal low [Fe 2+] induces toxin

Schick Test • Injection of toxin I D • Produces redness/erythemati c in 2-4 days • No reaction – Protective immunity present.

Epidemiology • • • • • •

Eradicated in developed nations, Children between 2 – 5 years. A symptomatic carriers Person to person . Carriers spread. Prolonged .

Active Immunization. • • • • • • • • •

Toxoid – Toxin treated with Formaldehyde Absorbed Toxoid Given by Intramuscular route Given in DTP –Triple Vaccine Primary Immunization Three Doses of DPT at least 4 weeks apart. Non vaccinated Three doses of Toxoid four weeks apart One dose after One Year.

Prevention Vaccination: Immunisation with diphtheria toxoid,

combined with tetanus and pertussis toxoid (DTP vaccine), should be given to all children at two, three and four months of age. Booster doses are given between the ages of 3 and 5 . The child is given a further booster vaccine before leaving school and is then considered to be protected for a further 10 years (16 – 18 years).

ive Immunization • • • • •

Given in Acute infections Give Subcutaneously 500 – 1000 Units of Antitoxin Given as Horse Serum Combined in Acute Infections ( Both Active Immunization with Toxoid and Antitoxin.

Diphtheria Epidemiology Reservoir Human carriers Usually asymptomatic Transmission Respiratory Skin and fomites rarely Temporal pattern

Winter and spring

Communicability Up to several weeks without antibiotics

Treating s • All s are advised to receive 500 mg Erythromycin 4 times a day.