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Objectives

Objectives. By the end of this lecture the student must be: A) Identify the genus Pseudomonadaceae & Vibrionaceae B) describe the chemical tests for this genus C ) Differentiate between different sps . D) List and match the symptoms, diagnosis and treatment for different sps.

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Objectives

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  1. Objectives • By the end of this lecture the student must be: • A) Identify the genus Pseudomonadaceae & Vibrionaceae • B) describe the chemical tests for this genus • C) Differentiate between different sps. • D) List and match the symptoms, diagnosis and treatment for different sps.

  2. Pseudomonas aeruginosa • Gram-negative bacilli belonging toPseudomonadaceae • Motile by single polar flagellum, non spore • Non fastidious- Minimal nutritional requirements • Produces grape-like odor and blue-green pus and colonies • Two types of soluble pigments: • Pyoverdin: Yellow-green pigment & fluorescent • Pyocyanin: Blue-green pigment and non-fluorescent • Strict aerobic, O+/F-, Oxidase and catalase positive • Optimum temperature is 37 0C & able to grow at 42 0C • Resistant to dyes, weak antiseptics and many antibiotics

  3. Epidemiology • The most important pathogenic is Ps. aeruginosa • Opportunistic pathogen • Important agent in causing nosocomial infections • Ubiquitous in moist environment, hospital, in soil and water • Colonized in the intestine in10% of human • Represents 10-20% of hospital-acquired infections • Transiently colonize respiratory & GIT of hospitalized patients: • Treated with broad-spectrum antibiotics • Exposed to respiratory therapy equipment • Hospitalized for long periods

  4. Pathogenesis 1. Adhesins • Fimbriae, Pilli, Alginate (mucoid exo-polysaccharide) • Alginate slim forms biofilm that protect from antibodies, complement, phagocytosis & antibiotic • Invasins • Proteases- Inactivate IFN and TNF • Elastase • Break down of elastin-containing tissues e.g. blood vessels, lung tissue, skin • Cleaves collagen, IgG, IgA and complement) • Produce hemorrhagic lesions associated with disseminated infection • Alkaline protease (lyses fibrin) • Hemolysins (phospholipase & lecithinase) & Leukocidin

  5. Pathogenesis • Toxins • Exotoxin A & S • Inhibit protein synthesis (Like diphtheria toxin) • Immunosuppressive • Causes dermatonecrosis in burn wounds, corneal damage in ocular infections, and tissue damage in chronic pulmonary infections • Pyocyanin – inhibits mitochondrial enzymes, impairs ciliary function, mediates tissue damage • LPS (endotoxin)

  6. Clinical Finding • UTIs (Catheterized patients) • Wound, burn & other skin & soft tissues infections • Echtyma Gangrenosum • Pneumonia (Cystic Fibrosis) • Eye infection- Contaminated contact lens clearing fluids • Chronic otitis media & otitis externa (swimming pool) • Meningitis- following lumbar puncture • Systemic infections- Septicemia in debilitated patients • Pseudomonal Endocarditis

  7. Echtyma Gangrenosum Causative agent:   Ps. aeruginosa Under risk: immunocompromised, burn patients, and other critically ill patients Round or oval (1-15cm) severe invasive cutaneous ulceritic single or multiple  lesion with halo of erthryma Pseudomonas exotoxin: Tissue destruction   Elastase degrades elastin in blood vessels wall Phospholipase C degrades phospholipids in cell membranes Pyocyanin generates reactive oxygen species  

  8. Typing methods • Bacteriocin typing • Three types of bacteriocins are produced-R,F,S • Pyocin produced by test strain is employed to assess the growth inhibition of 13 indicators strains • Depending upon the growth inhibition of 13 indicators strains,105 types are recognized • Most popular method used • Phage typing • Serotyping-based on O & H,17 serotypes are recognized

  9. Laboratory diagnosis • Specimen • Urine, pus, sputum, CSF, blood, skin swap • Microscopical Examination • Gram-negative rods and motile • Cultural Characteristics • On Nutrient agar: • Colonies are surrounded by bluish green coloration • On selective media "Cetermide" • Pigments are more obvious • On Blood agar: -hemolytic colonies • On MacConkey agar • Pale yellow colonies i.e. non lactose fermenters • Ps. aeruginosa able to grow at 42 0C for 3 days • Pyocin typing: Identification for epidemiological purpose

  10. Vibrionaceae • General characteristics • Gram negative, curved, comma shaped bacilli • Motile by single polar flagella • Non spore, Non capsulated • Facultative anaerobes, Fermentative • Oxidase and catalase positive • Most vibrios have relatively simple growth factor requirements and grow well in alkaline pH • Natural inhabitants : Aquatic environment

  11. Vibriocholerae • >200 serogroups based on O-antigen • V. cholerae O1 (classical and El Tor biotypes) and O139 are primarily responsible for cholera outbreaks • V. El Tor biotype causes cholera-like but milder • Recently, V. cholerae O75 and O141 strains has been associated with cholera-like diarrhea • Some O1 strains do not produce cholera enterotoxin (atypical or nontoxigenic O1 V. cholerae) • Cholera is endemic in southern Asia (India, Pakistan and Bangladesh), Latin America

  12. Pathogenesis • Mode of Transmission • Consumption of contaminated water or food • Incubation period:1-4 days • High infectious dose: >108 CFU • V. cholerae attach to mucosa of small intestine • V. cholerae multiply & secrete enterotoxin (cholargen) • Toxin binds to specific receptor on the intestinal mucosal cell • Toxin stimulates activity of cAMP, resulting in active secretion of chloride & secondary loss of Na and H2O • The patient loss 20 Litre fluids/day • V. cholerae cause same disease as ETEC but more severe

  13. Diagnosis of V. cholerae • Specimen: Rice watery stool or rectal swab • Culture on Alkaline Peptone Water (APW) then subculture on selective differential medium Thiosulfate Citrate Bile Salts Sucrose agar • Selective due to alkalinity pH9 & contains bile salts • Differential because contains sucrose • Sucrose fermenting V. choleraeyellow colonies • Sucrose non fermenting V. parahemolyticusappears as blue to green colonies • Sucrose fermentation is gold standard in identification

  14. Diagnosis of V. cholerae • Any sucrose fermenting colonies were subjected to Gram stain and oxidase test • Gram stain • Gram negative short rods, comma shaped, motile • Biochemical reactions: • Oxidase positive • O+/F+ • Cholera red reaction • TCBS agar (Figure) (Thiosulfate-citrate-bile salts-sucrose agar) • Serology: • Diagnosis confirmed & serotyping done by agglutinationwith specific antisera (O1, O139)

  15. V. parahaemolyticus • It is the cause of acute gastroenteritis following ingestion of contaminated sea-food such as raw fish • Marine organism which live in high salt conc. • So it requires 2% Na Cl in the isolation medium • It is invasive affecting the colon • V. cholerae is noninvasive, affecting the small intestine through secretion of an enterotoxin • Allied Vibriosare large group of organisms; some are saprophytic while others cause disease in animals

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