2. Clinical Microbiology MLCM- 201) Prof. Dr. Ebtisam. F. El Ghazzawi. Medical Research Institute (MRI) Alexandria University

3. Intended Learning Outcomes By the end of this lecture the student should be able to understand all about Gram Positive Rods Types pathogens and there Laboratory Diagnostic tests.

5. There are four mainly important genera of gram positive rods: bacillus, clostridium, corynebacterium and listeria. Bacillus and clostridium form spores, whereas corynebacterium and listeria do not. Members of the genus bacillus are aerobic, whereas those of the genus clostridium are anearobic.

6. The gram-positive rods can also be distinguished based on their appearance on gram stain. Bacillus and clostridium species are longer and more deeply staining than those of corynebacterium and listeria species. Corynebacterium species are club-shaped, i.e. they are thinner on one end than the other. Corynebacterium and listeria species characteristically appear as V or L-shaped rods.

7. Many important infections due to bacteria are common. The infections are often polymicrobial, that is, the anaerobic bacteria are found in mixed infections with other anaerobes, facultative anaerobes and aerobes. Anaerobic bacteria are found throughout the human body, on the skin, on mucosal surfaces and in high concentrations in the mouth and gastrointestinal tract, as part of the normal flora.

8. Infection results when anaerobes and other bacteria of the normal flora contaminate normally sterile body sites. Several important diseases are caused by anaerobic Clostridium species from the environment or from normal flora: botulism, tetanus, gas gangrene, food poisoning and pseudomembranous colitis.

9. There are four medically important species: Cl. tetani, Cl. botulinum, Cl. perfringens (which causes either gas gangrene or food poisoning), and Cl. difficili. All Clostridia are anaerobic, spore forming, gram-positive rods.

10. Cl. tetani causes tetanus (lockjaw). Spores are widespread in soil. The portal of entry is usually wound site. e.g. where a nail penetrates the foot. Germination of the spores is favored by necrotic tissue and poor blood supply in the wound.

11. Neonatal tetanus in which the organism enters through a contaminated umbilicus or circumcision wound is a major problem in some developing countries. Once the organism grow it elaborate a toxin that spreads along nerves to the central nervous system (CNS) and leads to muscle spasm. Death results from inability to breath.

12. There is no microbiologic or serologic diagnosis. Organisms are rarely isolated from the wound site. Cl. tetani produces a terminal spore i.e. spore at the end of the rod. This gives the organism the characteristic appearance of a “tennis racket”.

13. Cl. botulinum causes botulism. Spores widespread in soil, contaminated vegetables and meats. When these foods are canned or vacuum-packed without adequate sterilization, spores survive and germinate in the anaerobic environment.

14. Toxin is produced within the canned food and ingested performed. The highest risk foods are: 1.Alkaline vegetables such as green beans, peppers and mushrooms. 2.Smoked fish. The toxin is relatively heat labile, it is inactivated by boiling for several minutes. Thus, disease can be prevented by sufficient cooking.

15. The organism is usually not cultured. Botulism toxin is demonstrated by injecting a mice with a sample of clinical specimen, where it will dies unless protected by antitoxin.

16. Cl. perfringens causes two distinct diseases, gas gangrene and food poisoning. Spores are located in the soil, vegetative cells are members of the normal flora of the colon and vagina. Gas gangrene is associated with war wounds, automobile and motorcycle accidents and septic abortions.

17. Pain, oedema and cellulitis occur in the wound area. Crepitation indicates the presence of gas in tissues. Hemolysis and jaundice are common as are blood-tinged exudates. Shock and death can occur. Mortality rates are high.

18. Smears of tissue and exudate samples show large gram-positive rods. The organisms are cultured anaerobically. On blood agar Cl. perfringens colonies exhibit a double zone of hemolysis.

19. There are two important pathogens in this group: Corynebacterium diphtheriae and Listeria monocytogenes. C. diphtheriae causes diphtheria. Other Corynebacterium species (diphtheroids) are implicated in opportunistic infections.

20. Corynebacteria are gram-positive rods that appear club-shaped (wider at one end) and are arranged in palisades or V or L-shaped formation or Chinese like structure. Humans are the only natural host of C. diphtheriae. The organisms reside in the upper respiratory tract and are transmitted by airborne droplets.

21. The host response to C. diphtheriae consist of: 1.Local inflammation in the throat with a fibrinous exudate that forms the tough, adherent, gray pseudomembrane over the tonsils and throat characteristic of the disease. 2.Antibody that can neutralize exotoxin activity.

22. Involves both isolating the organism and demonstrating toxin production. The decision to treat with antitoxin is a clinical one and cannot wait for the laboratory results. A throat swab should be cultured on Löffler’s medium and blood agar.

23. Immunization of children by DPT (Diphtheria pertusis and tetanus toxoids) in the age of 3, 4 and 6 months with a boosters at 1 and 6 years of age.

24. Assignment Escherichia Coli.

25. Questions Complete the following : There are four mainly important genera of gram positive rods………..,…………,……….and……. Cl. tetani causes………(…………). Cl. botulinum causes …………….

26. Recommended Textbooks Manual of Clinical Microbiology, Vols. 1 and 2: Eighth Edition Patrick R. Murray