18 -chapter_20_-_curved_gnb

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18 -chapter_20_-_curved_gnb

  1. 1. CURVED GRAM-NEGATIVE Chapter 20 BACILLI
  2. 2. OBJECTIVES Describe the gram stain morphology of Vibrio species. Discuss the clinical significance of Vibrio cholerae. List the selective media for the isolation of Vibrio species and describe the different colony morphologies observed on the media. Discuss the mode of transmission of Vibrio cholerae. Identify Vibrio cholerae based on biochemical reactions. Explain the role of the vibrio toxin the pathogenesis of cholera infections. Differentiate the species of Vibrio species discussed in this unit based on colony morphology on Thiosulfate Citrate Bile Salts Sucrose (TCBS) and clinical symptoms of patient.
  3. 3. VIBRIO SPECIES
  4. 4. GENERAL CHARACTERISTICS More than 30 recognized species Approximately 12 species have been associated with human disease Persists in the environment:  Grows in saltwater, freshwater  Causes human disease because it can adapt to colonize intestinal tract if ingested
  5. 5. PROGRESSION OF INFECTION
  6. 6. MICROSCOPIC CHARACTERISTICS Gram-negative, curved rod Motile  single polar flagellum  “Darting” motility  “Shooting star” motility
  7. 7. GENERAL BIOCHEMICAL CHARACTERISTICS Most are:  Indole positive  Urea negative  Nitrate positive  Most are oxidase positive Posses the vibriostatic compound O/129
  8. 8. MEDIA AND GROWTH Media  Grow quite readily on various routine media  Use of Thiosulfate-Citrate-Bile Salts-sucrose agar (TCBS) has enhanced recovery  Alkaline peptone water - enrichment broth Conditions for growth  Facultative anaerobes  Cultures should be incubated at 35°C in air or CO 2
  9. 9. COLONY MORPHOLOGY On TCBS agar:  Sucrose fermenters are yellow  V. cholera and V. alginolyticus  Non-sucrose fermenters are green  V. parahaemolyticus On BAP and chocolate agar:  Most are iridescent with a greenish hue
  10. 10. VIBRIO CHOLERAE
  11. 11. EPIDEMIOLOGY Mostly found in aquatic environments Transmitted:  Fecal-oral route  Contact with contaminated water  Ingestion of contaminated shellfish or other seafood
  12. 12. VIRULENCE FACTORSProduction of Cholera toxin or choleragen  Provokes an accumulation of cyclic AMP in the cell membrane which causes mucosal cells to hypersecrete electrolytes and water into the lumen of the GI tract which results in watery diarrhea and fluid loss = rice -water stools  Such a rapid fluid and electrolyte loss leads to severe dehydration, hypovolemic shock, metabolic acidosis and death in just a matter of hours
  13. 13. CLINICAL SIGNIFICANCE Etiologic agent of cholera Cholera is mostly seen in Asia and South America Causes both intestinal and extraintestinal infections Infections are acquired by ingesting contaminated food or water Characteristic “rice-water” stools
  14. 14. EPIDEMIOLOGYContaminated water enters stomachBacterial cells adhere to gastric and intestinal mucosal epithelial cellsEnterotoxin is produced  Increase in cyclic AMP - prevents the re-absorption of Na+ ions and the excretion of sodium bicarbonate and potassium  Causes water to leave the epithelial cells into intestinal lumen causing "rice water" stoolsIt may lead to death in matter of hours
  15. 15. SPECIMEN MANAGEMENT Stool specimens are preferred - must be transported in Cary - Blair medium Rectal swabs acceptable
  16. 16. MEDIA FOR ISOLATION Will grow on BAP, Choc, and MAC (non -lactose fermenter on MAC) Alkaline peptone water broth for enrichment Thiosulfate Citrate Bile Salts Sucrose (TCBS) - selective and differential media (yellow or green colonies)  Yellow - ferment sucrose  Green - do not ferment sucrose
  17. 17. COLONY MORPHOLOGYVibrio cholerae - BAP
  18. 18. COLONY MORPHOLOGYVibrio cholerae - MAC
  19. 19. COLONY MORPHOLOGYVibrio cholerae - TCBS
  20. 20. BIOCHEMICAL CHARACTERISTICS Oxidase positive Sucrose positive (yellow on TCBS) Lactose negative (clear on MAC) Lysine positive Ornithine positive Arginine negative String test positive – 0.5% deoxycholate
  21. 21. BIOCHEMICAL CHARACTERISTICS String test  Principle:  Addition of 0.5% sodium deoxycholate causes most Vibrio spp. to lyse and release DNA, which can be pulled up into a string with a inoculating loop
  22. 22. TREATMENT Therapy - fluid and electrolyte replacement Antimicrobial therapy will shorten the course of the disease and may decrease bacterial excretion Tetracycline is the drug of choice
  23. 23. VIBRIO ALGINOLYTICUS
  24. 24. CLINICAL SIGNIFICANCE Least pathogenic for humans and least isolated Wound, ears, or eyes after trauma and sea water exposure
  25. 25. MEDIA AND GROWTH Strict halophile = requires added NaCl for optimal growth Yellow colonies on TCBS
  26. 26. COLONY MORPHOLOGYVibrio alginolyticus - BAP
  27. 27. COLONY MORPHOLOGYVibrio alginolyticus - TCBS
  28. 28. BIOCHEMICAL CHARACTERISTICS Sucrose positive
  29. 29. VIBRIOPARAHAEMOLYTICUS
  30. 30. CLINICAL SIGNIFICANCE Acute gastroenteritis associated with the ingestion of raw contaminated seafood, particularly oysters Usually mild but can be fatal Endemic in Japan; #1 cause of “summer diarrhea” Usually self-limiting disease Rare extraintestinal pathogen
  31. 31. COLONY MORPHOLOGYVibrio parahemolyticus - BAP
  32. 32. COLONY MORPHOLOGYVibrio parahemolyticus - TCBS
  33. 33. BIOCHEMICAL CHARACTERISTICS Sucrose negative Beta-hemolytic on blood agar
  34. 34. VIBRIO VULNIFICUS
  35. 35. CLINICAL SIGNIFICANCE Second most pathogenic Causes wound infections:  Wound infections – after exposure to marine animals or marine environment Wound infections may lead to septicemia  Primary septicemia – 24 hours after ingestion of raw oysters
  36. 36. COLONY MORPHOLOGY
  37. 37. COLONY MORPHOLOGYVibrio vulnificus - green or yellow on TCBS Agar
  38. 38. BIOCHEMICAL CHARACTERISTICS Sucrose variable
  39. 39. CAMPYLOBACTER SPECIES & HELICOBACTER PYLORI
  40. 40. OBJECTIVES Discuss the clinical significance of Campylobacter and Helicobacter pylori. Summarize specimen collection and transport procedures for Campylobacter and Helicobacter. Select appropriate culture media and incubation conditions for Campylobacter cultures. Compare the methods for obtaining microaerobic conditions. Describe the microscopic morphology of Campylobacter and Helicobacter. Differentiate species of Campylobacter based on biochemical identification. Explain the principle and procedure of the CLO test and breath tests for the detection of Helicobacter.
  41. 41. CAMPYLOBACTER SPECIES
  42. 42. CLINICAL SIGNIFICANCE Gastroenteritis - Campylobacter spp. is one of the leading causes of bacterial diarrhea worldwide Septicemia
  43. 43. EPIDEMIOLOGY Transmitted via contaminated food, milk, or water
  44. 44. SPECIMEN MANAGEMENT Stool and rectal swabs - if specimen cannot be cultured within 4 hours of collection, place in Cary -Blair transport medium Blood
  45. 45. GROWTH CONDITIONS Optimal temperature for growth is 42 ̊C Microaerophilic (5-10% oxygen) Capnophilic (8-10% CO 2 ) Needs at least 48-72 hours for growth
  46. 46. MICROSCOPIC CHARACTERISTICSCurved gram negative rodsComma shapes, "S" shapes and gull wing forms are seenOrganisms may occur in short or occasionally long chainsOxidase positiveNon-sporeformingCells have a single polar unsheathed flagellum at one or both ends = “darting motility”
  47. 47. MEDIA FOR ISOLATION Media  Campy BAP (most commonly used)  Butzler Medium  Skirrow medium  Campy-Thio for enrichment Examine at 24, 48 and 72 hours
  48. 48. COLONY MORPHOLOGY Colonies smooth Convex Translucent May be flat and watery with irregular edges They may be pinpoint to spreading over large areas of the plate
  49. 49. COLONY MORPHOLOGY
  50. 50. BIOCHEMICAL CHARACTERISTICS Catalase positive Oxidase positive Darting motility
  51. 51. BIOCHEMICAL CHARACTERISTICS C. coli  Hippurate negative  Resistant to cephalothin  Susceptible to nalidixic acid C. jejuni  Hippurate positive  Resistant to cephalothin  Susceptible to nalidxic acid
  52. 52. TREATMENT Antimicrobic therapy is not recommended in most cases of diarrhea Supportive care and fluid replacement When needed:  Erythromycin is the drug of choice Systemic infections are treated with Gentamycin
  53. 53. HELICOBACTER PYLORI
  54. 54. CLINICAL SIGNIFICANCE Found only on mucus-secreting epithelial cells of the stomach Causative agent of active chronic gastritis (type B gastritis) = Peptic ulcer disease
  55. 55. MICROSCOPIC MORPHOLOGY Gram negative Small curved (gull wing and U shapes) Slightly plump bacilli
  56. 56. SPECIMENS Stool cultures are not often done for detection of Helicobacter pylori Specimens/test of choice include:  Invasive (tissue biopsy)  CLO rapid urease test  Non-invasive  Urea breath test  Serological detection of IgG
  57. 57. BIOCHEMICAL CHARACTERISTICS Oxidase positive Catalase positive Urease positive
  58. 58. WHAT’S NEXT? Review Mahon, chapter 20 Complete AUMoodle Review

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