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BACTERIAL CLASSIFICATION AND DIAGNOSIS OF BACTERIAL DISEASE

BACTERIAL CLASSIFICATION AND DIAGNOSIS OF BACTERIAL DISEASE. Disease causing agent Bacterial, fungal, viral or other? Treatment Antibiotic sensitivity Source of infection Food, air or contact? Epidemiology Cholera, Diphtheria Prevention E. coli O157:H7 outbreak, Shigella, Listeria.

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BACTERIAL CLASSIFICATION AND DIAGNOSIS OF BACTERIAL DISEASE

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  1. BACTERIAL CLASSIFICATION AND DIAGNOSIS OF BACTERIAL DISEASE • Disease causing agent • Bacterial, fungal, viral or other? • Treatment • Antibiotic sensitivity • Source of infection • Food, air or contact? • Epidemiology • Cholera, Diphtheria • Prevention • E. coli O157:H7 outbreak, Shigella, Listeria

  2. Antibiotic sensitivity

  3. Site of sampling • Sterile sites • Blood • Cerebrospinal fluid (CSF) • Body fluids (Peritoneal and pleural) • Non-sterile (normal flora) • Respiratory tract • Ear, eye and mouth • Skin (wound and abscess) • Urine (mid-stream) • Feces

  4. Bacterial classification • Wall structure • Gram + • Staphylococcus, Streptococcus, Clostridium, Bacillus • Gram - • Enteric, respiratory and others • Acid-fast • Mycobacterium • Wall-less • Mycoplasma • Unusual • Obligate intracellular • Rickettsia, Chlamydia Bacteria G+ G- AF WL IC

  5. Bacterial classification • Cell morphology • Shapes • Rod • Cocci • Spiral • Associations • Individual • Diplo- • Staphylo- • Strepto- Bacteria G+ G- AF WL IC Rod Cocci Rod Cocci Spiral

  6. Bacterial classification • Growth characteristics • Oxygen requirement • Aerobic • Anaerobic • Microaerophilic, aerotolerant • Facultative • Spore formation • Intracellular/extracellular • Fastidious/non-fastidious Bacteria G+ G- AF WL IC Rod Cocci Rod Cocci Spiral + +/- -O2 + spore -

  7. Classification & Diagnosis • Type of colonies • Appearance • Color, shape, size and smoothness • On differential media • Blood, MacConkey, EMB • On selective media • MacConkey, Thayer-Martin

  8. Classification & Diagnosis • Metabolism • Utilization of specific substrates • Lactose (Sal/Shi/Yer/)- • Citrate (E. coli-/Klebsiella+) • Production of certain end products • Fermentation end products • Acid (acetate, propionic acid, butyric acid etc.) • Acetoin • Alcohol • Amine • H2S

  9. Classification & Diagnosis • Specialized tests • Immunological • O-, H- & K-Ag (serotype) • Precipitation, agglutination • Specialized enzymes • Catalase--- Staph+. vs. Strep-. • Coagulase---S. aureus+ vs. S. epidermidis- • Oxidase---Neisseria gonorrhoea+ • Urease---Proteus+, Helicobacter+ • Antibiogram pattern • Phage typing • Fatty acid profile

  10. Immunological detection

  11. Conventional diagnosis methods

  12. Conventional method • Depend on ability to culture • Treponema pallidum • Slow, esp. for fastidious species • Mycobacteriumspp. • Not always definitive

  13. Molecular diagnosis • Ribotyping • Restriction fragment length polymorphism (RFLP) • DNA hybridization • PCR, RT-PCR and RAPD • Nucleic acid sequence analysis • Phage-GFP (TB)

  14. RFLP GGATCC CCTAGG

  15. DNA hybridization

  16. In situHybridization

  17. PCR RT-PCR Rate of increase 2n

  18. RAPD of P. aeruginosa

  19. Reduce reliance on culture Faster More sensitive More definitive More discriminating Techniques adaptable to all pathogens Technically demanding Relatively expensive Can be too sensitive Provides no information if results are negative Molecular diagnosis

  20. Bioterrorism • Pathogen detection • Fast and accurate • Mobile • Inexpensive • Source investigation

  21. Differentiating Staphylococci from Streptococci • Gram stain and morphology • Both Gram + • Staphylococci: bunched cocci • Streptococci: chained cocci (S. pneumoniae form diplococcus) • Enzyme tests • Staphylococci: catalase + • Streptococci: catalase - • Growth • Staph.: large colonies (non-fastidious), some hemolytic • Strep.: small colonies (fastidious), many hemolytic (a or b)

  22. Staphylococci • S. aureus: coagulase + • S. epidermidis: coagulase -

  23. Streptococci

  24. Differentiating the Gram- bacteria • Cocci • Neisseria • Rods • Type of disease they cause • Enteric Gram- rods • API test • Curved • Vibrio, Campylobacter, Helicobacter • Spiral Gram- organisms • Spirochetes

  25. Gram negative Curved rods Straight rods Lactose+ Lactose- TCBS agar Yellow Oxidase+ Vibrio Campy blood agar 42oC+ 25oC- Campylobacter Citrate+ Citrate- H2S+ H2S- Klebsiella E. coli Salmonella Shigella

  26. Bacteria Intra Cellular Wall Less Gram+ Gram- Acid Fast Cocci Rod Spiral Rod Cocci M.t. M.l. N.c. Rickettsia Coxiella Erlichia Chlamydia Mycoplasma Staph. Strep. Non-spore Spore Treponema Borrelia Leptospira Neisseria Moraxella Fil Rod +O2 -O2 Curve B.a. B.c. C.b. C.t. C.p. C.d. S. a. S. e. S. s. A B Pn Vir A.i. C.d. L. m. Straight Vibrio Campylobacter Helicobacter +O2 +/-O2 -O2 Other P.a. Enteric Bact. Resp. Zoo GU H. ducreyi Gardnerella Calymmatobacterium Yersinia Pasteurella Brucella Francisella Streptobacillus Bordetella. H. influenzae Legionella

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