1. Welcome to MICROnesia 4 “Bug” Case Studies “Life of a Blood Culture” Slide Show Questions welcomed!

2. Case #1 UTI BUG Ambulatory 26 year old female with 101° temperature and painful urination Physician orders a urine culture with gram stain

3. Gram Stain Results Gram stain morphology shows many gram-positive cocci in pairs and chains

4. Urine Culture Setup Urine plated to agar plates 1/1000 ml inoculating loop used One big drop of urine is enough for a culture!

5. Urine Culture Results Culture grows >100,000 colonies of bacteria on a blood agar plate Patient’s UTI caused by a strep-like organism called Enterococcus

6. Identifying Enterococcus Produces an enzyme called PYRase Detectable in a two minute test

7. Normal sites for Enterococcus Upper respiratory tract Gastrointestinal tract Genitourinary tract

8. Enterococcus Infections UTI’s Nosocomial UTI’s Wound infections

9. Emerging Resistance Emerging strains showing resistance to Vancomycin Resistant strains called Vancomycin Resistant Enterococcus or VRE Bone marrow transplant and other immunocompromised patients at risk

10. Identifying VRE Identify VRE as an Enterococcus faecalis or faecium using biochemical tests interpreted by an automated instrument

11. Phoenix Automated Instrument Performs both biochemical tests and susceptibilities 100 organisms can be tested at a time

12. VRE on the rise Enterococcus showing resistance to Vancomycin E-strip VRE strains account for 6% of all Enterococcus Patients placed in isolation Reported to RN and Infection Control

13. Case #2 Wound Bug 65 year old male with 101° temperature after hip replacement surgery Develops redness, tenderness and drainage at incision site Physician orders a culture and gram stain on incision site

14. Incision site Gram Stain Gram stain shows few gram-positive cocci in clusters with few wbc’s

15. Bacterial culture results Staph aureus isolated on culture White colonies on blood agar

16. Identifying Staph aureus Latex agglutination test can identify an organism as Staph aureus in 10 seconds

17. Staph aureus infections Skin infections Scalded Skin Syndrome Toxic Shock Syndrome Osteomyelitis Food poisoning

18. Staph aureus reservoirs Carried in nose of 20-40% of adults Higher % in hospital personnel Transferred from nose to skin Passed to others by direct contact or droplets Primary way nosocomial infections occur

19. Staph aureus treatment Penicillin discovered in 1920 – worked great on Staph! More difficult to treat the last 50 years Some SA now showing resistant to methicillin, a commonly used drug

20. Identifying MRSA Strains resistant to methicillin are called MRSA Extraction test can identify SA as an MRSA strain in 15 minutes

21. Lots of MRSA Up to 50% of SA isolated are MRSA strains Carriage rate for MRSA higher in hospitals MRSA often found on health club gym equipment Pets can get MRSA from their owners

22. Wash Your Hands Good handwashing essential! Careful wound dressing technique Patients with MRSA placed in isolation Reported to RN and Infection Control

23. Case #3 GI BUG 38 year-old HIV positive male Several previous hospital admissions Taking AZT & Bactrim antibiotic therapy 3 day history of severe diarrhea with 10 pound weight loss and profound dehydration

24. Lab Results Stat Leukotest = negative (test for fecal wbc’s) Occult blood exam = negative Both tests usually positive with diarrhea caused by Salmonella or Shigella Negative Leukotest and Occult blood = noninflammatory diarrhea

25. Lab Results not Stat Ova & Parasite exam negative Stool culture negative for enteric pathogens Campylobacter EIA assay negative Shiga Toxin EIA assay negative

26. Other Findings No recent travel history Patient has not recently eaten shellfish

27. Clues from Patient History Severe diarrhea consistent with enterotoxigenic E.coli or Vibrio cholerae Endemic in limited regions Raw or undercooked shellfish may contain Vibrio cholerae Patient had not consumed shellfish

28. Suppressive Antibiotic Therapy Normal gut flora protects the bowel from invasive pathogens Antibiotics destroy large part normal flora Allows overgrowth of organisms usually suppressed

29. Responsible Bug Clostridium difficile frequently causes antibiotic-associated diarrhea Disrupted normal flora allows C. difficile to multiply Produces two different exotoxins

30. Patient’s Diagnosis Patient suffering from Clostridium difficile colitis “Pseudomembranous colitis”

31. More about Clostridium difficile C. difficile is an anaerobe Gram-positive rods on Gram Stain

32. Diagnosing C. difficile colitis Detect exotoxins in stool using EIA assay Performed twice daily in Microbiology Takes about 3 hours Pea-size amount of stool needed for testing Positive results called to patient’s RN

33. Important to Establish Cause of Diarrhea Many causes of diarrhea in AIDS patients untreatable C. difficile treatable with oral antibiotics Patient placed in isolation to avoid hospital outbreaks

34. Life of a BLOOD CULTURE Slide Show Drawn in yellow-top SPS tubes Full size & pedi-tube

35. Life of a BLOOD CULTURE 4 Kinds of Blood Culture Bottles Aerobic Anaerobic Pediatric ARD (Antimicrobial Removal Device)

36. Life of a BLOOD CULTURE Chlorhexidine preps or swabs disinfect venipuncture site Scrub arm for 30 seconds, not to exceed a 2 inch square surface Let arm air dry

37. Life of a BLOOD CULTURE Use of Chlorhexidine preps has decreased blood culture contamination rate by 50% Blood culture considered “contaminated” if common skin flora grows from one or both bottles in a set

38. Life of a BLOOD CULTURE Clean SPS tubes with alcohol and let air dry Draw 2 SPS tubes for each set of cultures 10 ml in each tube One tube –> aerobic One tube –> anaerobic Record collection site on label (peripheral, art line, etc.)

39. Life of a BLOOD CULTURE Recommended draw times: Two sets drawn at least 30 minutes apart in a 24 hour period Bacterial recovery rate increases by 57% when 2 sets are drawn

40. Life of a BLOOD CULTURE Bottles placed in an automated Bactec instrument Incubate for 5 days Monitored every 15 minutes for bacterial growth

41. Life of a BLOOD CULTURE Loud alarm sounds when growth is detected! Positive blood culture considered a STAT Subcultured to agar plates Plates incubate for 18 hours

42. Life of a BLOOD CULTURE Gram stain slide made from “positive” bottle

43. Life of a BLOOD CULTURE Gram Stain takes about two minutes Look for bacteria on slide under the microscope Gram stain results called to patient’s RN

44. Case #4 BLOOD BUG 37 year old man with sickle cell disease and numerous hospitalizations Porta-cath placed in right subclavian vein Patient admitted to ED two weeks after porta- cath placement

45. Emergency Department findings Patient has right arm discomfort and swelling Physician orders two sets of blood cultures One drawn through porta-cath One drawn through peripheral vein

46. Blood culture results Both sets of blood cultures show gram positive cocci in clusters on smear Both cultures grow the same organism

47. Responsible Bug Two positive blood cultures + porta-cath = probable line-related sepsis Most common bug causing line-related infection is Coagulase Negative Staph or CNS CNS  important cause of nosocomial bacteremia Foreign body devices act as source

48. Identifying CNS Grow as white colonies on blood agar plate Nonreactive in rapid latex tests

49. Sources of CNS Normal inhabitants of skin, mucous membranes and nares About 20 species of CNS Most common is Staph epidermidis

50. Slime Producers CNS secrete a virulence factor called slime Makes them “sticky” Stick to plastic surfaces like catheter tips Slime-producing strains more difficult to treat with antibiotics Indwelling catheters place patient at risk for infection

51. Diagnosing Line-Related Sepsis Draw 2 sets of blood cultures from a patient with fever or signs of infection at the IV site One set from catheter line One set from peripheral site

52. Two sites important CNS on skin can be a blood culture “contaminant” if blood not collected properly Single positive blood culture with CNS may be skin contamination and not true infection

53. Two sites important Negative peripheral culture and positive line culture with CNS may just show local infection of the catheter site Two Blood cultures with CNS from two different sites more likely represents true infection

54. Confirming line-related sepsis Confirm by performing a catheter tip culture Catheter is removed and sent to Micro Lab

55. Culturing the Catheter tip Catheter tip cut to 50mm Roll on surface of blood agar plate

56. Interpreting Catheter tip cultures Culture “positive” if 15 colonies grow from a 50 mm tip CNS growing on plate

57. Diagnosis confirmed If Catheter tip culture has CNS and blood cultures from both the line and peripheral draws have CNS  Patient has a confirmed line-related sepsis

58. Microbiology Art Thank you! Please call Microbiology with any questions: 2-2422 2-2435