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Bacterial Testing of Platelet Components: 2008 Update

Bacterial Testing of Platelet Components: 2008 Update. William B. Lockwood, PhD, MD Clinical Professor Department of Pathology & Laboratory Medicine University of Louisville Director, Transfusion Services & Tissue/Bone Bank University of Louisville Hospital

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Bacterial Testing of Platelet Components: 2008 Update

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  1. Bacterial Testing of Platelet Components: 2008 Update William B. Lockwood, PhD, MD Clinical Professor Department of Pathology & Laboratory Medicine University of Louisville Director, Transfusion Services & Tissue/Bone Bank University of Louisville Hospital Director, Transfusion Services, Tissue/Bone Bank & Coagulation Lab Norton Hospital & Kosair Children’s Hospital Louisville, Kentucky wblock01@gwise.louisville.edu 502-852-5857

  2. Objectives • Describe the current FDA regulations and accreditation agency standards for testing platelet collections for bacterial contamination • Compare & contrast 3 methods of bacterial detection in platelet collections • Describe a validation plan for use of non-FDA approved bacterial detection methods PACE Program Number: 362-001-09 SCACM is approved as a provider of continuing education programs in the clinical laboratory sciences by the ASCLS P.A.C.E. Program. SCACM Conference Jan. 20, 2009

  3. Whole Blood Collection Set ca. 1900 TODAY! Whole Blood Glass Bottle ca. 1940 SCACM Conference Jan. 20, 2009

  4. Bacterial Contamination of Blood Components • Known to occur since 1900 when using vented glass bottles for whole blood collection • Continued with advent of plastic blood containers in 1950s • Platelets stored refrigerated had less of a contamination problem, but were not efficacious on transfusion*! *Murphy S, Gardner FH. NEJM 1969;380:1094-98 SCACM Conference Jan. 20, 2009

  5. Bacterial Contamination of Blood Components • Additional studies on platelet storage medium, storage temperature, pH, type of agitation, volume of platelets in container and size of container, plastic used for platelet bag from 1960s-1980s • Changes included: • Gentle horizontal agitation • Room temperature (RT) storage- 3 days to 5 days (1981); 5 days to 7 days (1984) • Reduction from 7 day to 5 day RT storage (1986)* *Shiffer CA et al. Blood 1986;67:1591-94 SCACM Conference Jan. 20, 2009

  6. Bacterial Contamination of Blood Components • US Food & Drug Administration (FDA) via blood regulatory department Center for Biologics Evaluation & Research (CBER) held industry workshops in 1995 & 1999 to discuss platelet bacterial contamination • Literature continues to report increasing platelet bacterial contamination (BaCon study of American Red Cross Blood Services) SCACM Conference Jan. 20, 2009

  7. Bacterial Contamination of Blood Components • Bacterial contaminated blood components cause of >10% (77/694) of recipient fatalities reported to FDA from 1985-1999* • 1st multicenter study of bacterial contamination of blood components published by American Red Cross** *Workshop on bacterial contamination of platelets.Bethesda: FDA, Center for Biologics Evaluation and Research,September 24, 1999. Available from http://www.fda.gov/cber/minutes/workshop-min.htm. **Kuehnert MJ, et al. Transfusion-transmitted bacterial infection in the United States, 1998 through 2000. Transfusion 2001;41:1493-99 SCACM Conference Jan. 20, 2009

  8. Bacterial Contamination of Blood Components • BaCon study • 23,711,169 RBCs • 1,804,725 single donor platelets (SDP) • 1,033,671 Pooled platelets (WBDP) SCACM Conference Jan. 20, 2009

  9. Bacterial Contamination of Blood Components • Transfusion-transmitted bacteremia rates: • SDP= 1:100,000 • WBDP= 1:100,000 • RBC= 1:8,000,000 SCACM Conference Jan. 20, 2009

  10. Bacterial vs. Virus: Infectious risk in the blood supply Hepatitis B HBsAg 1971 Anti HBsAg 1987 HIV P 24 1996 HIV PCR 2000 HIV Ab 1985 Hepatitis C HCV Ab 1990 HCV PCR 2000 HTLV 1/2 HTLV 1/2 1989 West Nile Virus WNV NAT 2003 No Practical Screening Tests Currently Available Bacteria - Platelets 1:2,000 Bacteria - Red Cells 1:20,000 Source: Ilert WE, et al. Transfusion Medicine 1995, 5:57-61 Brecher et al. Clinical Microbiology Reviews, 2005, 195-204 1:100,000 1:1,000,000 Incidence rate SCACM Conference Jan. 20, 2009

  11. Bacterial Contamination of Blood Components Sources of Bacterial Contamination Skin Surface Contamination Phlebotomy Core Donor Bacteremia Containers and Disposables Environment SCACM Conference Jan. 20, 2009

  12. Bacterial Contamination of Blood Components ORGANISMS INVOLVED Exogenous Normal skin Staphylococcus epidermidis Staphylococcus aureus Diphteroids spp Micrococcus spp Pseudomonas spp Bacillus cereus Propionibacterium acnes Flavobacterium spp SCACM Conference Jan. 20, 2009

  13. Bacterial Contamination of Blood Components ORGANISMS INVOLVED Endogenous Staphylococcus spp TeethStreptococcus viridans Serratia liquefaciens Yersinia enterocolitica Intestines Salmonella spp Campylobacter spp Osteomyelitis Staphylococcus S. cholera suis SCACM Conference Jan. 20, 2009

  14. Bacterial Contamination of Blood Components Prevention and Detection Options • Donor screening – not feasible except for arm screening. Can’t detect asymptomatic bacteremic donors • Arm Preparation-Limited effectiveness of arm scrub • Better phlebotomy methods and initial blood diversion • Bacterial contamination testing offers best confirmatory option • Pathogen Inactivation--INTERCEPT SCACM Conference Jan. 20, 2009

  15. FDA Regulations • FDA is silent about Whole-blood Derived Platelets (WBDP) bacterial testing guidelines [FDA guidelines are recommendations but are not legally binding until published as a Code of Federal Regulations (CFR)] • HOWEVER- establishments requesting a Biological License Application (BLA) or Supplement (BLS) must follow the Title 21 CFR Parts 200 and 600 SCACM Conference Jan. 20, 2009

  16. FDA Regulations • Part 200 relates to Good Manufacturing Practice, current (cGMP) of biologics • Part 600 relates to blood components - safety, quality, purity, potency, identification • Part 640- platelets • Subsections itemize various manufacturing regulations, - source, donor suitability, collections, QC • QC elements- platelet count of donor, donor weight, pH at outdate (≥6.2),sterility testing, etc • Manufacturer’s requirements for automated plateletpheresis collection SCACM Conference Jan. 20, 2009

  17. FDA Regulations • Revised regulation effective in 2008 (1st was in 1988, draft in 2005!!) • “Revisions to the Requirements Applicable to Blood, Blood Components and Source Plasma”* • Pertains to those establishments that want to license their blood component • THEREFORE, FDA regulates platelet “manufacturing” through the BLA/BLS!! • Also, approval of manufacturer’s devices through Pre-Market Approval 510(k) application regulates equipment used in all phases of blood component production *Federal Register: August 16, 2007 (Volume 72, Number 158) SCACM Conference Jan. 20, 2009

  18. CAP Accreditation Checklist • 1st accreditation group to require bacterial testing of platelets! • CAP Checklist - 2002 • Phase I deficiency • Revised 12/29/2004 to Phase II deficiency SCACM Conference Jan. 20, 2009

  19. CAP Accreditation Checklist 2008 • TRM.44955 Phase II N/A YES NO • Does the laboratory have a validated system to detect the presence of bacteria in platelet components? • NOTE: For random donor platelets, any of the following testing methods satisfy this checklist question: detection of decreased pH or glucose by analytic instrument or dipstick; gram stain; acridine orange stain. Though of low sensitivity, these methods may detect units that are heavily contaminated by bacteria. Culture or FDA-approved commercial detection systems have greater sensitivity. The swirling technique is not recommended because of its very low sensitivity. SCACM Conference Jan. 20, 2009

  20. CAP Accreditation Checklist 2008 • Two commercial systems have been cleared by the FDA for in-process quality control culturing of platelet units; one detects the growth of bacteria by their generation of CO2, and the other detects growth by their consumption of O2. Another system has been cleared for bacterial detection by fluorescent staining. If this testing is performed by the supplier of platelet components, the transfusion service can satisfy this checklist requirement by having an agreement with the supplier to notify the transfusion service if any units suspected of containing bacteria have been transferred to the transfusion service. SCACM Conference Jan. 20, 2009

  21. AABB Accreditation • AABB Standards for Blood Banks & Transfusion Services, 23rd ed, Effective May 1, 2004 • Implement standard 5.1.5.1: The blood bank or transfusion service shall have methods to limit and detect bacterial contamination in all platelet components. Standard 5.6.2 applies • Standard 5.6.2- Protection Against Contamination: The venipuncture site shall be prepared so as to minimize risk of bacterial contamination. Green soap shall not be used. SCACM Conference Jan. 20, 2009

  22. AABB Accreditation con’t • Standard 5.1.5.1.1 (Standard 5.1.5.2, 25th ed, 2008) • When a true-positive result is obtained and an appropriate specimen is available, additional testing to identify the organism shall be performed. Additional testing and follow-up shall be defined. Standards 5.2.2 and 7.1 to 7.1.4 apply. • Standard 5.2.2: Donor Notification of Abnormal Findings and Test Results (Standard 5.2.3, 25th ed, 2008) • Standards 7.1-7.1.4: Nonconformances SCACM Conference Jan. 20, 2009

  23. AABB Accreditation con’t • Standard 5.6.6.1 (25th ed, 2008) Blood collection containers with draw line (inlet) diversion pouches shall be used for any collection of platelets, including whole blood from which platelets are made. SCACM Conference Jan. 20, 2009

  24. Bacterial Contamination of Blood Components • Bacterial detection tests • •3 devices are cleared for quality control monitoring of platelet collection process of leukoreduced platelets • BioMeriuex BacT/ALERT® • Pall eBDS • hemoSystems Scansystem™ • •Other non approved and non validated methods are also being used to meet the AABB standard for bacterial detection SCACM Conference Jan. 20, 2009

  25. Bacterial Contamination of Blood Components FDA concerns with bacterial detection as currently applied (Vostal JG, Jan. 28, 2005 CBER workshop) • Test performance characteristics unknown • Use of non-validated tests (glucose and pH by dipstick, swirling) • Non-standardized methodology even with culture-based devices • Potential for excessive false positives or negatives • Less reliable methods are used on whole blood derived platelets creating a two tiered safety system for apheresis and whole blood derived platelets SCACM Conference Jan. 20, 2009

  26. Bacterial Contamination of Blood Components • Manual tests- validation required for pH & glucose • pH • Glucose • Swirling SCACM Conference Jan. 20, 2009

  27. SCACM Conference Jan. 20, 2009

  28. Devices approved for QC detection of platelet bacterial contamination- Pall BDS SCACM Conference Jan. 20, 2009

  29. Pall eBDS • Sample set/Oxygen Analyzer • Sterile weld platelet component to set • Fill pouch with ~3 mL of product • Disconnect sample pouch from set and incubate at 35°C for 24-30 hrs • Measure the O2 content in the air above the plasma sample with insertion of analyzer probe into pouch • LED display will read PASS or FAIL SCACM Conference Jan. 20, 2009

  30. Pall eBDS SCACM Conference Jan. 20, 2009 Pall Medical eBDS Brochure, 2004

  31. BioMeriuex BacT/ALERT® • Colorimetric technology/Sensor Culture bottles • CO2 release causes sensor bottle to turn yellow • Instrument measures & detects color change, analyzes data to determine positivity, alerts when positive culture SCACM Conference Jan. 20, 2009

  32. hemoSystems Scansystem™ • Scansystem Platelet Kit/Scansystem Analyzer • After sample processing in platelet kit, bacteria are stained with a green fluorescence and retained on the surface of a dedicated membrane (platelets are aggregated and lysed) • Membrane is inserted in analyzer and scanned by an Argon laser • Each fluorescent spot on the membrane will be detected and analyzed • Analyze 3 platelet components (SDP, WBDP) SCACM Conference Jan. 20, 2009

  33. hemoSystems Scansystem™ Scansystem™ Brochure accessed at www.hemosystem.com SCACM Conference Jan. 20, 2009

  34. Preparing Platelets for Culture Sterile Connection Device SCACM Conference Jan. 20, 2009

  35. Connecting Platelets to Syringe Set SCACM Conference Jan. 20, 2009

  36. Tubing from syringe set Tubing from platelet bag SCACM Conference Jan. 20, 2009

  37. Aliquoting Platelet Samplefrom Transfusion Bag(Suspected Transfusion Reaction) Syringe set Transfusion set SDP SCACM Conference Jan. 20, 2009

  38. Aliquoting Platelet Samplefor Bacterial Detection Testing SCACM Conference Jan. 20, 2009

  39. Aliquoting Platelet Samplefor Bacterial Detection Testing SCACM Conference Jan. 20, 2009

  40. Inoculating Culture Bottle (pediatric size) SCACM Conference Jan. 20, 2009

  41. Sterile Connection Port SCACM Conference Jan. 20, 2009

  42. Bacterial Contamination of Blood Components-2008 • ARC Study of bacterial contamination before & after implementation of sample diversion pouch (Pall Acrodose with eBDS) • Prestorage pooling of WBDP with culture testing • Jan 2003-December 2006 Benjamin RJ, et al. Transfusion 2008;48:2348-55 SCACM Conference Jan. 20, 2009

  43. Bacterial Contamination of Blood Components-2008 Benjamin RJ, et al. Transfusion 2008;48:2348-55 SCACM Conference Jan. 20, 2009

  44. Bacterial Contamination of Blood Components-2008 Benjamin RJ, et al. Transfusion 2008;48:2348-55 SCACM Conference Jan. 20, 2009

  45. Bacterial Contamination of Blood Components-2008 PSP=prestorage pooled WBDP Benjamin RJ, et al. Transfusion 2008;48:2348-55 SCACM Conference Jan. 20, 2009

  46. SCACM Conference Jan. 20, 2009

  47. Blood Transfusion Safety-FDA Blood Transfusion Fatality Report SCACM Conference Jan. 20, 2009 http://www.fda.gov/Cber/

  48. Why test at point of issue ? Wide variation in rate of log Phase Growth Source: Goodnough LT et al. NEJM 1999, Klein et al.JAMA, vol 274, issue 1368 –1373, November 1,2005, Goodman et al. Bacterial Contamination of blood Components: Risk, Strategy and regulation, Hematology 2003 Wide variation in duration of Lag Phase SCACM Conference Jan. 20, 2009

  49. New FDA Approved Device for Transfusion Services • The Abbott Verax Platelet PGD® Test • A rapid, qualitative immunoassay for the detection of Aerobic and Anaerobic; Gram-positive and Gram-negative bacteria in leukocyte reduced apheresis platelets (SDP) • ~30 minute TAT • SDP must have undergone QC culture by blood supplier!! SCACM Conference Jan. 20, 2009

  50. New Device for Transfusion Services SCACM Conference Jan. 20, 2009

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