Corynebacterium and Other Non Spore-forming Gram-Positive Rods Chapter 16

1. Corynebacterium and OtherNon–Spore-forming Gram-Positive Rods – Chapter 16 Corynebacterium sp. Listeria monocytogenes Erysipelothrix rhusiopathiae

2. Corynebacteria Significant Corynebacterium species C. xerosis C. pseudodiphtheriticum C. pseudotuberculosis C. jekeium C. ulcerans Rhodococcus equi Arcanobacterium haemolyticum

3. Corynebacterium Species General characteristics Found as free-living saprophytes in fresh and salt water, in soil and in the air Members of the usual flora of humans and animals(often dismissed as contaminants) Often called “diphtheroids” Corynebacterium diphtheriae is the most significant pathogen Other species may cause infections in the immunocompromised hosts

4. Corynebacterium Species:General Characteristics Morphology Gram-positive, non–spore-forming rods Arrange in palisades:“L-V” shape; “Chinese characters” Pleomorphic: “club-ends” or coryneform Beaded, irregular staining

5. C. diphtheriae: Agent of Diphtheria Toxigenic Corynebacterium diphtheriae Worldwide distribution but rare in places where vaccination programs exist Exotoxin, Diphtheria toxin, as the virulence factor Not all C. diphtheriae strains produce toxin Toxin is produced by certain strains Lysogenized by bacteriophage with toxin gene (tox+) Toxin is antigenic

6. Toxigenic Corynebacterium diphtheriae Toxin consists of two fragments A: Active fragment Inhibits protein synthesis Leads to cell/tissue death B: Binding Binds to specific cell membrane receptors Mediates entry of fragment A into cytoplasm of host cell

7. Clinical Forms of Diphtheria Respiratory Acquired by droplet spray or hand to mouth contact Non-immunized individuals are susceptible Non-respiratory Systemic Skin and cutaneous forms

8. C. diphtheriae: Causative Agent of Diphtheria Respiratory disease–diphtheria Incubation period–2 to 5 days Symptoms: sore throat, fever, malaise Toxin is produced locally, usually in the pharynx or tonsils Toxin causes tissue necrosis, can be absorbed to produce systemic effects Forms a tough grey to white pseudomembrane which may cause suffocation

9. C. diphtheriae: Causative Agent of Diphtheria C. diphtheriae pseudomembrane WBC + organism

10. Clinical Infections: Non-Respiratory Disease Systemic infections Toxin is absorbed in the blood stream and carried systemically Affects the kidneys, heart, and nervous system Death occurs due to cardiac failure Cutaneous form More prevalent in the tropics Infections occur at the site of minor abrasions Maybe superinfected with Streptococcus pyogenes and/or Staphylococcus aureus

11. Treatment Infected patients treated with anti-toxin and antibiotics Anti-toxin produced in horses Antibiotics have no effect on circulating toxin, but prevent spread of the toxin Penicillin drug of choice

12. Laboratory Diagnosis Microscopic morphology Gram-positive, non–spore-forming rods, club-shaped, can be beaded Appear in palisades and give "Chinese letter" arrangement Produce metachromatic granules or “Babes’ Ernst” bodies (food reserves) which stain more darkly than remainder of organism

13. Laboratory Diagnosis: Cultural Characteristics Loeffler's slant or Pai's slant—Used to demonstrate pleomorphism and metachromatic granules ("Babes’ Ernst bodies“) Growth on Serum Tellurite or modified Tinsdale exhibits brown or grayish? to black halos around the colonies

14. Laboratory Diagnosis: Cultural Characteristics BAP@ 24-48 hours small, grey translucent colonies Small zone of ?- hemolysis also seen

15. Laboratory Diagnosis Identification Confirm identification by fermentation reactions(glucose +) Catalase positive Urease negative Non-motile

16. Laboratory Diagnosis Toxigenicity testing Elek test— p. 413 Immunodiffusion test Organisms are streaked on media with lox Fe content to maximize toxin production. Identification of C. diphtheriae does NOT mean the patient has dipheria. Must show the isolate produces the toxin.

17. C. diphtheriae Treatment: antitoxin Prevention: DPT immunization

18. C. jekeium Clinical Infections Septicemia Meningitis Pulmonary disease Populations Affected Immunosuppressed IV drug users Recent invasive procedure

19. C. jekeiumColony Morphology Isolation & identification BAP: 48-72 hours @ 35oC in ambient air or 5% CO2 small, gray-white colony, nonhemolytic Gram stain: pleomorphic, club-shaped GPR arranged in V forms or palisades

20. C. jekeiumLab Diagnosis Identification Nitrate reduction= negative Urea= negative Sucrose= negative Glucose= positive

21. C. jekeium Susceptibility testing Exhibits resistance to multiple antibiotics Susceptible to vancomycin

22. Differentiating Characteristics of Corynebacterium Species

23. Listeria monocytogenes:General Characteristics Gram-positive, non–spore-forming rods Only human pathogen in genus Widespread in nature Known to infect a wide variety of animals Human exposure is limited; direct or indirect Transient colonization occurs without disease

24. Listeria monocytogenes:Clinical Infections Adults Septicemia/meningitis in the compromised/elderly Mild flu-like syndrome in pregnant women could be fatal to fetus Ingestion of contaminated food (cottage cheese, coleslaw, chicken, hot dogs, lunch meat) Neonatal Early onset from intrauterine transmission results in sepsis; high mortality rate Late onset manifests as meningitis; lower mortality rate

25. Listeria monocytogenes: Virulence Factors Hemolysin ( Listeriolysin O)- damages macrophage Catalase Superoxide dismutase Phospholipid C P60 surface protein- induces phagocytosis thru adhesion and penetration

26. Laboratory Diagnosis: L. monocytogenes Identification Microscopic morphology Gram Positive non–spore-forming coccobacillary, pairs or short chains Colony Morphology Grows well on blood agar; colonies produce a narrow zone of hemolysis similar to Group B Streptococcus Small, round and translucent

27. Laboratory Diagnosis: L. monocytogenes Grows well at 0.5° C to 45° C Because of this temperature range, especially the cooler end of the range, this organism grows well in refrigerated products, such as cream, cheese, deli meats, etc. Can sometimes be isolated after “cold enrichment” (hold broth at 4° C for several weeks and subculture)

28. Laboratory Diagnosis: L. monocytogenes Identification Catalase positive Motility: Motile at 25o C; "umbrella" type ? Tumbling motility in hanging drop preparations (this can be seen on Gram Stain Tutor at www.medtraining.org)

29. Laboratory Diagnosis: L. monocytogenes Identification CAMP test Produces a “block” type of hemolysis in contrast to “arrow”-shape produced by Group B Streptococcus

30. Differentiating Characteristics between L. monocytogenes and Other Gram Positive Bacteria

31. Erysipelothrix rhusiopathiae:General characteristics Gram positive, non–spore-forming, pleomorphic rods (can produce long filaments) Distributed in nature Can cause disease in animals (swine, turkey, sheep); swine is the main reservoir Humans acquire the infection through occupational exposure, such as cuts & scratches (fish handlers, animal products)

32. Erysipelothrix rhusiopathiae: Clinical Infections Erysipeloid Self-limiting localized infection at the site of inoculation Produces painful swelling, usually on the hands or fingers Heals within 3 to 4 weeks Endocarditis May occur in those who have had valve replacements Disseminated infections may occur, but rarely

33. Laboratory Diagnosis: Erysipelothrix rhusiopathiae Microscopic Morphology Pleomorphic, gram-positive thin rods that may form long filaments, may be arranged singly, in short chains, or in a V shape

34. Laboratory Diagnosis: Erysipelothrix rhusiopathiae Identification Catalase negative CO2 is required Distinguishing characteristic: Production of H2S on TSI Microaerophilic Nonmotile Test tube brush growth in semisolid motility media

35. Laboratory Diagnosis: Erysipelothrix rhusiopathiae Colony Morphology Grows on blood or chocolate agar—colonies may appear gray or translucent, pinpoint with alpha hemolysis or nonhemolytic

36. Treatment: Erysipelothrix rhusiopathiae Penicillin, cephalosporin, erythromycin

37. Characteristics of Corynebacterium, Listeria, and Erysipelothrix

38. Lactobacillus Widely distributed in nature Normal flora of mouth, GI tract and female genital tract Treat with pencillin plus an aminoglycoside resistant to vancomycin (helps in diagnosis) Clinical Infections Bacterial vaginosis Bacteremia, endocarditis, meningitis (rare)

39. Lactobacillus Microscopic Morphology Long, slender gram positive pleomorphic bacilli Non-spore forming Colony Morphology Microaerophilic SBA: pinpoint, a- hemolytic colonies Lab Diagnosis Catalase negative