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      The meeting convened at 8:00 a.m. in the
 10  Plaza Ballroom of the Hilton Washington
     DC/Rockville Executive Meeting Center, 1750
 11  Rockville Pike, Rockville, Maryland, Frederick
 12  P. Siegal, M.D., Chairman, presiding.
     FREDERICK P. SIEGAL, M.D., Chairman
 14  JUDITH R. BAKER, M.H.S.A., Consumer
 15  MARK BALLOW, M.D., Member
     ARTHUR W. BRACEY, M.D., Temporary Voting
 17  HENRY M. CRYER, III, M.D., Ph.D., Member
 18  MAUREEN A. FINNEGAN, M.D., Member
     THOMAS R. FLEMING, Ph.D., Temporary Voting
 19   Member
     SIMONE A. GLYNN, M.D., M.Sc., M.P.H., Member
     LOUIS M. KATZ, M.D., Non-Voting Industry
 21   Representative
     MATTHEW J. KUEHNERT, M.D., Member
 22  ROSHNI KULKARNI, M.D., Member
 23  This transcript has not been edited or
     corrected, but appears as received from the
     commercial transcribing service. Accordingly,
 25  the Food and Drug Administration makes no
     representation as to its accuracy.
      Page 2
     FERNANDA LESSA, M.D., M.P.H., Temporary Non-
  2   Voting Member (Topic I)
     KATHERINE A. McCOMAS, Ph.D., Member
  3  FRANCISCO J. RENTAS, Ph.D., Member
  4  BARRY S. SKIKNE, M.D., Temporary Voting Member
      (Topic II)
  5  ANN B. ZIMRIN, M.D., Member
     DONALD W. JEHN, M.S., Designated Federal
  7   Official
  8  JAY EPSTEIN, M.D., Director, Office of Blood
      Research and Review, CBER
  9  JESSE GOODMAN, M.D., M.P.H., Director, CBER
     SALIM A. HADDAD, M.D., Division of Hematology,
 10   Office of Blood Research and Review,
 11  LESLIE HOLNESS, M.D., Chief, Blood and Plasma
 12   Branch, Division of
      Blood Applications, CBER
 13  JARO VOSTAL, M.D., Ph.D., Office of Blood
      Research and Review, CBER
 15  MARK E. BRECHER, M.D., University of North
      Carolina School of Medicine
     GARY M. BRITTENHAM, M.D., College of
 17   Physicians and Surgeons, Columbia
 18  BARBARA BRYANT, M.D., University of Texas,
      Medical Branch, Galveston
 19  RITCHARD G. CABLE, M.D., American Red Cross
      Blood Services
     SARAH E. CUSICK, Ph.D., U.S. Centers for
 21   Disease Control and Prevention
     JERRY A. HOLMBERG, Ph.D., Executive Secretary,
 22   Advisory Committee on Blood Safety and
 23  KARIN MAGNUSSEN, M.D., Copenhagen University
 25   Ehrlich Institute
     DAN A. WAXMAN, M.D., Indiana Blood Center
      Page 3
  1   T-A-B-L-E O-F C-O-N-T-E-N-T-S
  2  Statement of Conflicts of Interest,
     Acknowledgment of Members . . . . . . . . . . .4
  4  Opening Remarks . . . . . . . . . . . . . . . 14
  5  Committee Update. . . . . . . . . . . . . . . 14
  6  Topic I. Strategies to Enhance Bacterial
     Safety of Seven-Day Platelets for Transfusion
  8   Jaro Vostal, M.D. . . . . . . . . . . . 35
      Mark Brecher, M.D.. . . . . . . . . . . 47
  9   Thomas Montag-Lessing, M.D. . . . . . . 78
      Louis Katz, M.D.. . . . . . . . . . . .103
 10   Salim Haddad, M.D.. . . . . . . . . . .130
 11  Open Public Hearing . . . . . . . . . . . . .151
     Open Committee Discussion and Questions . . .152
     Topic II: Iron Status in Blood Donors
      Leslie Holness, M.D.. . . . . . . . . .287
 15   Gary Brittenham, M.D. . . . . . . . . .296
      Sarah Cusick, Ph.D. . . . . . . . . . .331
      Karin Magnussen, M.D. . . . . . . . . .347
 17   Barbara Bryant, M.D.. . . . . . . . . .362
      Dan Waxman, M.D.. . . . . . . . . . . .400
 18   Ritchard Cable, M.D.. . . . . . . . . .416
 19  Open Public Hearing . . . . . . . . . . . . .458
     Questions for the Committee
 21   and Discussion. . . . . . . . . . . . .467
 22  Adjourn
      Page 4
  1   P-R-O-C-E-E-D-I-N-G-S
  2   8:07 a.m.
  3   CHAIRMAN SIEGAL: We're already a
  4  little late, so let's try and get started. If
  5  everybody could settle down.  
  6   MR. JEHN: Okay. Mr. Chairperson,
  7  members of the Committee, invited guests,
  8  temporary members and participants, I'd like
  9  to welcome you to this 92nd meeting of the
 10  Blood Products Advisory Committee.
 11   I'm Donald Jehn, the Designated
 12  Federal Official for this meeting.
 13   This meeting will be completely
 14  open to the public.  
 15   At this time I'd like to introduce
 16  the individual seated at the head table for
 17  today. Would the members and temporary
 18  members please raise their hand as their name
 19  is called.
 20   To the left of me is our BPAC
 21  Chairperson Dr. Frederick Siegal, Medical
 22  Director of Comprehensive HIV Center, St.
      Page 5
  1  Vincent's Catholic Medical Centers, New York.
  2   To the right of me and going
  3  around the table counterclockwise, Dr.
  4  Fernanda Lessa, she's going to be a nonvoting
  5  temporary member for Topic I only. She's from
  6  the CDC in the Epidemic Intelligence Service
  7  Officer, Division of Health Care Control and
  8  Prevention.
  9   Next, Ms. Judith Baker, our
 10  Consumer Rep. She's Regional Administrative
 11  Director of Federal Hemophilia Treatment
 12  Centers, Regions 9.
 13   Dr. Mark Ballow, Chief, Division
 14  of Allergy & Immunology, Department of
 15  Pediatrics, SUNY, Buffalo.
 16   Dr. Henry Cryer, Chief of Trauma
 17  and Critical Care, Division of General
 18  Surgery, UCLA.
 19   Dr. Adrian Di Bisceglie, Professor
 20  of Internal Medicine, Chief of Hematology,
 21  Saint Louis University.
 22   Dr. Arthur Bracey, Associate Chief
      Page 6
  1  Department of Pathology, at St. Luke's
  2  Episcopal Hospital, Houston.
  3   Dr. Edwards will join us in the
  4  afternoon.
  5   Dr. Maureen Finnegan, Associate
  6  Professor Department of Orthopedic Surgery,
  7  University of Texas, Southwestern Medical
  8  Center.
  9   Dr. Thomas Fleming, Professor of
 10  Department of Biostatistics, University of
 11  Washington.
 12   Going around to the other side,
 13  Dr. Harvey Klein, Chief, Division of
 14  Transfusion Medicine, NIH.
 15   Dr. Matthew Kuehnert, Director,
 16  Office of Blood, Organ, and other Tissue
 17  Safety, Division of Health Care Quality
 18  Promotion, CDC.
 19   Dr. Roshni Kulkarni, Professor,
 20  Department of Pediatrics and Human
 21  Development, Michigan State University.
 22   Dr. Katherine McComas, Assistant
      Page 7
  1  Professor, Department of Communications,
  2  Cornell.
  3   Dr. Francisco Rentas, Chief, Blood
  4  Services and RMC Blood Manager.
  5   Dr. Ann Zimrin, Associate
  6  Professor Division of Hematology Oncology,
  7  University of Maryland School of Medicine.
  8   Dr. Lewis Katz, our industry rep.
  9  He's the Executive Vice President of Medical
 10  Affairs, Mississippi Valley Regional Blood
 11  Center.
 12   The Committee members not in
 13  attendance are Drs. Manno, Toxel and Trunkey
 14  for this meeting.
 15   I'd like to thank you all for
 16  attending this meeting.
 17   I now have the COI statement to
 18  read. Please bear with me.
 19   The Food and Drug Administration,
 20  FDA, is convening the September 10th to 11th,
 21  2008 meeting of the Blood Products Advisory
 22  Committee under the authority of the Federal
      Page 8
  1  Advisory Committee Act, FACA, of 1972. With
  2  the exception of the industry representative
  3  all participants of the Committee are special
  4  Government employees or regular Federal
  5  employees from other agencies and are subject
  6  to the Federal Conflict of Interest laws and
  7  regulations.
  8   The following information on the
  9  status of this Advisory Committee's compliance
 10  with federal ethics and conflict of interest
 11  of laws including but not limited to 18 U.S.
 12  Code 208 and 712 of the Federal Food, Drug and
 13  Cosmetic Act are being provided to
 14  participants of this meeting and to the
 15  public.
 16   FDA has determined that all
 17  members of this Advisory Committee are in
 18  compliance with the Federal ethics and
 19  conflict of interest laws under 18 U.S. Code
 20  208. Congress has authorized FDA to grant
 21  waivers to special Government employees and
 22  regular Government employees who have
      Page 9
  1  financial conflicts when it is determined that
  2  the agency's need for a particular
  3  individual's service outweighs his or her
  4  potential financial conflicts of interest.
  5   Under 712 of the Food and Drug and
  6  Cosmetic Act Congress has authorized FDA to
  7  grant waivers to special Government employees
  8  and regular Government employees with
  9  potential financial conflicts when necessary
 10  to afford the Committee their essential
 11  expertise.
 12   Related to the discussions of this
 13  meeting members and consultants of this
 14  Committee have been screened for potential
 15  financial conflicts of interest of their own
 16  as well as those imputed to them, including
 17  those of their spouses or minor children. And
 18  for the purposes of 18 U.S. Code 208 their
 19  employers. These interests may include
 20  investments, consulting, expert witness
 21  testimony, contract and grants, CRADAs,
 22  teaching, speaking, writing, patents and
      Page 10
  1  royalties and primary employment.
  2   The Committee will discuss
  3  strategies to enhance bacterial safety of
  4  seven-day platelets for transfusion. This is
  5  a particular matter involving specific parties
  6  Topic I.
  7   For Topic II the Committee will
  8  discuss iron status in blood donors. This is
  9  a particular matter of general applicability.
 10   For Topic III the Committee will
 11  discuss options for blood donor screening and
 12  reentry for malaria. This is a particular
 13  matter involving specific parties.
 14   In addition, the Committee will
 15  hear updates and informational presentations
 16  on several topics. These updates and
 17  presentations are not for discussion by the
 18  Committee, and therefore the Committee members
 19  were not screened for financial interests
 20  relating to these presentations and
 21  informational updates.
 22   Based on the agenda and all
      Page 11
  1  financial interests reported by members and
  2  consultants no conflict of interest waivers
  3  were issued under 18 U.S. Code 208(b)(3) or
  4  712 of the Food, Drug and Cosmetic Act.
  5   With regard to FDA's guest
  6  speakers the agency has determined that the
  7  information being provided is essential. The
  8  following information is being made public to
  9  allow the audience to objectively evaluate any
 10  presentation and/or comments.
 11   For Topic I, Dr. Mark Brecher is a
 12  science advisor and speaker for several firms
 13  and receive research support from several
 14  firms that could be effected by the Committee
 15  discussions.
 16   For Topic II, Dr. Barbara Bryant
 17  is a principal investigator on an NIH protocol
 18  that could be affected by the Committee
 19  discussions. She recently was employed by the
 20  Department of Transfusion Medicine, NIH,
 21  though she's now actually at the University of
 22  Texas, Galveston.
      Page 12
  1   Dr. Richard Cable is the PI on a
  2  grant that could be affected by the Committee
  3  discussions.
  4   Dr. Karen Magnussen is an advisor
  5  for the Danish Blood Donor Organization.
  6   Dr. Louis Katz is serving as the
  7  industry representative acting on behalf of
  8  all related industry and is employed by the
  9  Mississippi Valley Regional Blood Centers. In
 10  addition, Dr. Katz is employed part-time with
 11  the Scott County Health Department, Iowa and
 12  the Genesis Health System in Davenport.
 13   Dr. Katz is a member and chair of
 14  various committees with the America's Blood
 15  Center and the American Association of Blood
 16  Banks.
 17   Industry representatives are not
 18  special Government employees and do not vote.
 19   In addition, there may be
 20  regulated industry or other outside
 21  organization speakers making presentations.
 22  These speakers may have financial interests
      Page 13
  1  associated with their employer and with other
  2  regulated firms. The FDA asks in the interest
  3  of fairness that they address any current or
  4  previous financial involvement with any firm
  5  whose product they may wish to comment upon.
  6   These individuals were not
  7  screened by the FDA for conflicts of interest.
  8   This conflict of interest
  9  statement will be available for review at the
 10  registration table.
 11   We would like to remind members,
 12  consultants and participants that if the
 13  discussions involve any other products or
 14  firms not already on the agenda for which an
 15  FDA participant has a personal or an imputed
 16  financial interest, the participants need to
 17  exclude themselves from such involvement and
 18  their exclusion will be noted for the record.
 19   FDA encourages all other
 20  participants to advise the Committee of any
 21  financial relationships that you may have with
 22  the sponsor, its product and, if known, its
      Page 14
  1  direct competitors.
  2   Thank you.
  3   Now it over to the Chair, Dr.
  4  Siegal.
  5   CHAIRMAN SIEGAL: I'd like to
  6  welcome everyone back from a somewhat
  7  tempestuous and somewhat actually continuingly
  8  tempestuous summer. Hopefully this summer
  9  won't be as agitated as the summer was.
 10   As you've heard from Don, we'll be
 11  talking first about strategies to enhance the
 12  bacterial safety of seven-day platelets and
 13  this afternoon we'll be considering iron
 14  status and blood donors. And I think perhaps
 15  we should start right away with a Committee
 16  update. This will be a summary by Jerry
 17  Holmberg of the May 29th to 30th Meeting of
 18  the DHHS Advisory Committee on Blood Safety
 19  and Availability.
 20   Dr. Holmberg?
 21   DR. HOLMBERG: Thank you, Mr.
 22  Chair.
      Page 15
  1   Since our last time meeting at the
  2  Blood Products Advisory Committee we did have
  3  a meeting of the Advisory Committee for Blood
  4  Safety and Availability on May 29th and 30th.
  5  And, sir, with you permission, I am going to
  6  spend a little bit more time talking about
  7  some of the topics that were discussed and
  8  specifically dwell on the recommendations.
  9   Because I do believe that there's
 10  a lot of synergy that has taken place between
 11  the Advisory Committee which is the
 12  Secretary's Committee for Blood Safety and
 13  Availability and also the FDA's Blood Products
 14  Advisory Committee and some of the topics that
 15  you will be discussing today.
 16   First of all, I do want to go
 17  through and just explain a little bit about
 18  the recommendation that came out of the
 19  January meeting of the Advisory Committee on
 20  Blood Safety and Availability.
 21   And I apologize for the print
 22  being pretty small there for those people in
      Page 16
  1  the audience but, hopefully, the Committee has
  2  handouts and also can read this along. I will
  3  read this for you.
  4   But at the January meeting of the
  5  Advisory Committee there was a recommendation
  6  on pathogen reduction and the Department
  7  looked at various aspects of this and the
  8  Secretary, or the Assistant Secretary for
  9  Health, Dr. Garcia, responded to Dr. Bracey
 10  with a letter just recently, and I will read
 11  that to you.
 12   "The decisions and the
 13  recommendations made by the Committee at the
 14  January 2008 meeting are substantial to
 15  supplying an additional layer of security to
 16  the safety of blood products. This area of
 17  new technology is exciting and has generated
 18  much discussion within the Department.  
 19   The Department fully supports a
 20  cooperative effort within its public health
 21  agencies together with stakeholders to advance
 22  development and validation of pathogen
      Page 17
  1  reduction technologies for all transfusable
  2  product components. We are committed to
  3  providing regulatory, scientific, and
  4  surveillance advice to facilitate the
  5  development of such products.  
  6   This summer the National Heart,
  7  Lung, Blood Institute will review the current
  8  status of pathogen reduction, identify primary
  9  critical non-infectious related research
 10  needs, and identify action that could advance
 11  this important area.
 12   Funding to support any new
 13  initiative is always a challenge. However,
 14  with the support of the Department, its
 15  operating divisions and other stakeholders,
 16  this additional layer of security could become
 17  a reality.
 18   In the long-run cost
 19  neutralization and even cost reduction
 20  avoidance could be possible with the
 21  elimination of procedures, e.g., irradiation
 22  and leukoreduction and/or reduction of
      Page 18
  1  emerging transfusion transmitted infectious
  2  diseases."
  3   We were all very pleased with this
  4  response from the Assistant Secretary for
  5  Health and we are moving forward to see how
  6  the government can move forward in this area.
  7   The next topic that was really
  8  discussed was some of these areas that were
  9  discussed at your last Blood Products Advisory
 10  Committee; but for completeness, I'm going to
 11  reiterate some of the information that was
 12  presented. I'm going to be looking at the
 13  blood safety team that was presented by Dr.
 14  Holness. Dr. Solomon presented the Tissue
 15  Safety Committee and then the area that was
 16  not discussed here at the Blood Products
 17  Advisory Committee was Dr. Rios' discussion on
 18  organ transplant related serious adverse
 19  events.
 20   First of all, just to reiterate
 21  some of the things that Dr. Holness mentioned
 22  in his presentation. This was based on 2007
      Page 19
  1  data. You can see that even in 2007 TRALI
  2  still remains to be a number one reported
  3  fatality.  
  4   I would also like to draw your
  5  attention to the six microbial deaths and
  6  three of them are Babesia related. I would
  7  also draw your attention that on Friday there
  8  will be a workshop on Babesia that the FDA is
  9  sponsoring.
 10   If we look at this information
 11  based over 2005, 2006, and 2007 you can see
 12  the percentages and also the number of
 13  reported cases for TRALI and also for
 14  microbial infections. This is a composite
 15  number so just to draw your attention to that.
 16   Also, Dr. Holness brought the
 17  attention of the Advisory Committee the
 18  information that there is a pending rule out
 19  there that was posted on March 14, 2003,
 20  Safety Reporting Requirements for Human Drug
 21  & Biological Products.  
 22   I think all of this information
      Page 20
  1  and some of the information that I'm
  2  presenting on the safety teams and also the
  3  organs and tissue all relate to what we are
  4  trying to do in conjunction with the AABB and
  5  UNOS on biovigilance. Just to draw your
  6  attention to the comments from this, even
  7  though it was published in 2003, are still
  8  under review for safety reporting.
  9   Also moving on to the tissue
 10  aspect of the presentation, some of the things
 11  that tissues get reports in from tissue
 12  establishments and Dr. Solomon brought a lot
 13  of this to our attention. Again, some of
 14  these same reporting mechanisms are mechanisms
 15  that we eventually will integrate into the
 16  biosurveillance process.
 17   Again, the tissue, just highlight
 18  some of the areas between 2006 and 2007 with
 19  the adverse reactions. You can see with the
 20  bone and the eye and muscular tissue and also
 21  skin being really high in the reported adverse
 22  reactions.
      Page 21
  1   If we look at the data and compare
  2  it to infectious versus noninfectious, you can
  3  see that even in 2006 and 2007 56 percent of
  4  the cases and 74 percent were related to
  5  infectious agents.
  6   Now switching to organs. What
  7  I've done is I've taken Dr. Ortiz-Rios'
  8  presentation and I've summarized it into just
  9  several slides. Primarily looking at
 10  infections if we take the organs and the cause
 11  of death being infections in 2001 through
 12  2006, about 21.5 percent of the causes of
 13  death in kidneys were due to infections. You
 14  can see 19 and 18 and hearts.  
 15   If we look at the period of time
 16  in 2001 to 2004 in a period of greater than
 17  one year or less than three years you can see,
 18  of course, that the infectious cause of death
 19  has dropped off in these various organ
 20  transplants.
 21   This is a slide that combines both
 22  the unknown cause of death and the missing
      Page 22
  1  data in the organ transplants. You can see
  2  that in kidney and liver and heart that the
  3  percentage is 16.8 to 16 percent but over the
  4  period of greater than one year to three years
  5  the amount of missing data or unknown cause of
  6  death increases the number of cases that are
  7  reported. You see a total of 14 percent going
  8  to 28 percent.
  9   There was a similar slide showing
 10  that definitely with the expanded number of
 11  years there was also an increased number of
 12  cause of death for malignancy. Now, the organ
 13  procurement network database has some problems
 14  with it. This is operated by UNOS. There is
 15  no data collected for serious viral, bacteria,
 16  fungal, or parasitic infections.  
 17   Donor related malignancy data, I
 18  should say, are collected but not nearly
 19  inclusive. Also that the OPTN has a Disease
 20  Transmission Advisory Group but it's voluntary
 21  reporting, no enforcement and limited follow-
 22  up.
      Page 23
  1   Some of the things that HRSA, one
  2  of the operating divisions within HHS, is
  3  dealing with is to define the reporting
  4  expectations of the Organ Procurement
  5  Organizations and CDC and also the state
  6  health authorities. To strengthen their
  7  Advisory Committee interaction for
  8  determination of donor transmission.  
  9   To strengthen Donor Transmission
 10  Advisory Committee's communication on
 11  potential tissues and organ destinations.
 12  Also to clarify and expedite CDC involvement
 13  in the investigation as an event unfolds.
 14   HRSA has asked OPTN when NAT
 15  testing for HIV could become nationally
 16  instituted. Also they are continuing to work
 17  with CDC to study events for guidance on how
 18  to screen more effectively without loss of
 19  organs due to false positives, and to
 20  investigate the possibility of an explicit
 21  approach to -- to investigate the possibility
 22  of an explicit approach to relating the risk
      Page 24
  1  without versus the risk of transmission with
  2  the transplant for an individual case.
  3   I bring that up because there is a
  4  recommendation that comes out of some of those
  5  observations and I will present that later on.
  6   One of the things we did have,
  7  moving on to the main focus of the meeting,
  8  Dr. Wright, who is the Principal Deputy
  9  Assistant Secretary for Health, did charge the
 10  Committee on, first of all, bacterial
 11  contamination of platelet concentrates, both
 12  apheresis and whole blood derived, and also
 13  the red cell age as a variable in transfusion
 14  outcome.
 15   As far as the platelets, he asked
 16  the Committee to think about is the risk
 17  associated with bacterial contamination of
 18  platelet concentrates and subsequent detection
 19  for both apheresis and whole blood derived
 20  platelets acceptable? If the risk associated
 21  with the available detection systems is
 22  unacceptable, what does the Committee
      Page 25
  1  recommend for next steps?
  2   As far as red cell age, do current
  3  data support a change in medical practice from
  4  transfusion of red cells stored for as long as
  5  42 days to a shorter period? If so, what
  6  would be the impact? Is there a need for
  7  additional research? What impact would change
  8  have on blood availability? Should the blood
  9  banking industry strive to produce improved
 10  red blood cell products?
 11   As we moved into the discussions
 12  on the platelets, our first discussion was Dr.
 13  Murphy's discussion in which he did a study on
 14  bacterial contamination of platelet
 15  concentrates where he cultured at four days.
 16  The conclusion of that was that there is low
 17  sensitivity of cultures due to the low number
 18  of bacteria and delay or slow growth of the
 19  bacteria.
 20   Basically the comment was that
 21  sampling will never reach an acceptable level
 22  of detection no matter how large the sample or
      Page 26
  1  sensitivity of the test. This will lead to
  2  increased morbidity, more recalls, and loss of
  3  product.
  4   I can't remember whether it was
  5  Dr. Murphy or someone on the Committee and we
  6  do have various Committee members at the table
  7  that probably can clarify this if I say it
  8  wrong but I think the statement that really
  9  caught my attention was that we will never be
 10  able to culture to sensitivity -- to
 11  sterility, I should say. Culture to
 12  sterility.
 13   Dr. Dumont presented the passport
 14  and the risk assessment. Dr. Benjamin
 15  presented the ARC experience, especially with
 16  the diversion pouch. Also, Dr. Jacobs
 17  presented information on the detection and
 18  some of the challenges of bacterial
 19  contamination of platelets.
 20   I do want to highlight some of the
 21  comments that Dr. Jacobs presented, primarily
 22  because I think it will help you in some of
      Page 27
  1  your discussions today. First of all, his
  2  comment was that whole blood derived platelets
  3  should be cultured early to bring their level
  4  of testing up to that of apheresis platelets.
  5   The optimum volume and condition
  6  to provide the most cost effective method for
  7  detection of bacterial contamination by early
  8  culture needs to be further studied. Also,
  9  the value of the anaerobic culture needs to be
 10  clarified. The real incidence of bacterial
 11  contamination needs to be studied by
 12  quantitative cultures at time of issue to
 13  assess the value of preventative and detection
 14  methods.  
 15   Also, the quantitative cultures of
 16  platelet units need to be performed on units
 17  with positive early culture broth cultures.
 18  And the clinical efficacy and cost
 19  effectiveness of point of issue assays by
 20  hospital transfusion services either as a
 21  stand-alone test or in addition to early
 22  culturing needs to be determined. The safety,
      Page 28
  1  efficacy, and cost effectiveness of pathogen
  2  inactivation in the eradication of bacterial
  3  contamination needs to be determined.
  4   We then moved on to the discussion
  5  on the age of the red cell which I think that
  6  may have been what the Chair was referring to
  7  as some of the storms experienced over the
  8  summer. There has been quite a bit of
  9  literature recently on the age of the red cell
 10  and who gets fresh red cells and who does not.
 11   We started looking at the
 12  discussion of the red cell storage lesion.
 13  Dr. Bunn presented quite a comprehensive
 14  review of that. Also the new insights into
 15  the healthy red cells with the nitric oxide.
 16   Also Dr. Koch from Cleveland
 17  Clinic presented her data on the 6,000 reviews
 18  that she did in cardiac surgery on the age of
 19  red cell. Then also Dr. McMahon from Duke
 20  presented some of the changes again with the
 21  nitric oxide effect in aged red cells.
 22   Dr. Glynn presented some of the
      Page 29
  1  things that the NHLBI was working on. In
  2  fact, the Transfusion Medicine and Hemostasis
  3  Network has put together a protocol called
  4  RECESS.  
  5   By the way, I just wanted to let
  6  you know that the Transfusion Medicine and
  7  Hemostasis Network has been in existence for
  8  about six years and is really a very powerful
  9  arm to get some of these studies done. Dr.
 10  Steiner presented what they are proposing for
 11  that and also that the NHLBI has put forward
 12  a funding opportunity announcement for basic
 13  research on transfusable red cells.
 14   So let me just try to finish up
 15  with less than a minute to go. I think I'll
 16  probably go over my time. Anyway, the
 17  recommendation on bacterial contamination is
 18  that the Committee appreciates that
 19  intervention including culture and diversion
 20  have reduced the risk of bacterial
 21  transmission.
 22   However, more effective methods
      Page 30
  1  are needed to further limit or eliminate the
  2  risk of bacterial contamination of platelets.
  3  Additionally, the current status of disparate
  4  levels of safety for platelet products is
  5  highly problematic.  
  6   The Committee recommends that
  7  additional measures, e.g., prevention,
  8  detection, pathogen inactivation, be adopted
  9  to reduce the difference in safety profile
 10  between whole blood and apheresis platelets
 11  and reduce the overall risk of bacterial
 12  contamination of platelets.  
 13   The Department should monitor the
 14  current status of platelet availability and
 15  potential for meeting future needs. Support
 16  should be established for initiatives to
 17  extend platelet storage life as a strategy to
 18  improve platelet availability.
 19   I'm sure you'll have a lively
 20  discussion on the platelets today and I am
 21  very appreciative that several of our
 22  Committee members are also on your Committee
      Page 31
  1  with Dr. Bracey being the chair of our
  2  Advisory Committee for Blood Safety and
  3  Availability being a temporary voting member
  4  for this meeting.
  5   Going on for the red cell age in
  6  transfusion, the Committee recommended that
  7  based on the available scientific data of red
  8  cell storage the Committee is concerned about
  9  the potential toxicity associated with
 10  progressive storage of red blood cells,
 11  particularly in certain clinical settings,
 12  e.g., cardiac surgery, ICU, and trauma.  
 13   However, absent the availability
 14  of definitive safety data from adequately
 15  controlled clinical trials, and in the absence
 16  of any analysis of the impact of shortened red
 17  cell dating on blood availability, the
 18  Committee believes that a change in practice
 19  is premature.
 20   The Committee recommends efforts
 21  to optimize blood transfusion practices in
 22  these areas through research and promulgation
      Page 32
  1  of clinical practice guidelines based on
  2  scientific evidence of safety and efficacy.
  3  As needed, the Committee recommends the
  4  Department be supportive of operations
  5  research on management of blood inventory.
  6   The Committee finds that the
  7  available scientific data from observational
  8  and limited prospective clinical studies are
  9  insufficient to resolve concerns regarding
 10  safety of progressive stored red cells.
 11   Therefore, adequately controlled
 12  clinical research is needed to correlate basic
 13  science findings on the adverse effects of
 14  progressive red cell storage with clinical
 15  outcomes. In parallel, studies are needed to
 16  establish the efficacy of transfusion
 17  therapies in various clinical settings.
 18  The Committee recommends new and sustained
 19  investment in basic and clinical research in
 20  this area.
 21   Then the meat of the
 22  recommendation was that whereas the HHS
      Page 33
  1  Advisory Committee on Blood Safety and
  2  Availability is charged with advising the
  3  Assistant Secretary on public health issues
  4  related to the safety of tissue and organ
  5  transplantation and after review of the
  6  current status of safety and utilization
  7  reporting for organs and tissue the Committee
  8  recommends enhanced acquisition of data on
  9  tissue distribution and utilization to allow
 10  current surveillance activity to better
 11  determine the frequency of adverse events.
 12   Capture of appropriate data
 13  regarding etiological agents of infection
 14  reported following organ transplant to allow
 15  for better assessment of infectious risk
 16  related to transplantation.
 17   And to support the acceleration of
 18  the development of a rapid infectious disease
 19  assays for use in the organ transplant setting
 20  as a strategy to improve both safety and
 21  availability of organs.  
 22   And to also enhance utilization of
      Page 34
  1  the Center for Medicare and Medicaid Services
  2  and other available databases to improve
  3  monitoring of organ transplant practices and
  4  related outcomes through cooperative
  5  arrangements with other agencies. So the last
  6  recommendation was actually dealing with the
  7  first discussion points on the safety teams
  8  and also the adverse events related to tissue
  9  and organs.
 10   Just to follow up here, the
 11  nomination for ACBSA closed on June 30th and
 12  are currently being vetted. The ACBSA Charter
 13  is in the process of being renewed. We are
 14  under the Sunset Law and do have to be renewed
 15  every two years.
 16   We are continuing to work on the
 17  biovigilance coordination throughout the
 18  Department. We will be reporting soon,
 19  hopefully within the next month, the National
 20  Blood Collection and Utilization Survey. Our
 21  meeting is upcoming in December on the 16th
 22  and 17th of December right here in this hotel
      Page 35
  1  and this is our website. Thank you.
  2   DR. SIEGAL: Thank you, Dr.
  3  Holmberg. Let's move directly into the Topic
  4  I on seven-day platelets. We are going to
  5  hear first an introduction from Jaro Vostal
  6  from FDA.  
  7   Dr. Vostal.DR. VOSTAL: Good
  8  morning. It's my pleasure today to be able to
  9  introduce this topic that we are going to
 10  discuss today and the topic on the agenda is
 11  Strategies to Enhance Bacterial Safety of
 12  Seven-Day Platelets for Transfusion.
 13   As a way of introduction, as
 14  pretty much everyone is currently aware,
 15  bacterial contamination of platelet products
 16  remains a serious problem. It occurs
 17  frequently to the best of our knowledge.
 18  Approximately one per 2,000 to one per 5,000
 19  collected units are contaminated.
 20   Bacteria can grow in platelet
 21  products to very high levels during storage at
 22  room temperatures. This brings on the serious
      Page 36
  1  consequences that can lead to morbidity,
  2  mortality of patients who are transfused with
  3  these contaminated products. This is
  4  underscored by the reports to the FDA that 60
  5  deaths were caused by bacterially contaminated
  6  platelets between the years 1995 and 2004.
  7   So part of the problem that we
  8  have with bacterial contamination of platelets
  9  is that platelet storage supports bacterial
 10  growth. Platelets are collected -- collected
 11  platelets are stored at room temperature in
 12  gas permeable bags and so far we have not been
 13  able to move away from room temperatures.
 14  Storage of 4 degrees is associated with rapid
 15  clearance of platelets from circulation.
 16   Storage is also in plasma, a
 17  medium that can support bacterial
 18  proliferation. Based on these issues the
 19  current shelf life of platelets is five days.
 20  There were two historic attempts to extend
 21  shelf life of platelets to seven days and this
 22  was in 1985, the mid '80s, and more recently
      Page 37
  1  in 2005. Unfortunately, both times the shelf
  2  life was reduced back to five days over
  3  concerns of increased bacterial contamination.
  4   Now, there are methods currently
  5  available to decrease bacterial contamination
  6  of platelet products and these include skin
  7  disinfection, the diversion of initial small
  8  volume of blood that is collected away from
  9  the final collection bag because it's thought
 10  that some of the initial blood actually
 11  carries the bacteria to the final product.
 12   Then there are detection methods
 13  using cultured based bacterial detection
 14  devices and more recently cleared antibody
 15  based bacterial detection devices with a rapid
 16  turn around.
 17   The clearance of these bacterial
 18  detection devices by the FDA is based on the
 19  intended use for these devices. The FDA
 20  recognizes two intended uses right now. One
 21  is for quality control and the intent here is
 22  to monitor blood collection process for
      Page 38
  1  increased rates of contamination.
  2   This type of intended use can be
  3  cleared based solely on the in vitro data
  4  performed by the device. The other intended
  5  use is a release of products for transfusion.
  6  The intent here is to monitor each product to
  7  determine suitability for transfusion. This
  8  is a more rigorous process and FDA clearance
  9  is based on evidence of clinical performance
 10  in addition to in vitro data.
 11   These are the bacterial detection
 12  devices that are currently on the market. The
 13  culture based devices are the BacT/ALERT
 14  device manufactured by bioMerieux. It has a
 15  Q/C indication. There is an eBDS devices
 16  manufactured by Pall Corporation. It also has
 17  a Q/C indication.  
 18   And there is the antibody based
 19  rapid detection device by Verax that has
 20  received an indication or a clearance as an
 21  adjunct to a Q/C device indication. That is
 22  because on its own it didn't have sufficient
      Page 39
  1  sensitivity to be placed in the Q/C indication
  2  category.
  3   The application of these bacterial
  4  detection devices to extend shelf life was
  5  used around the mid 2000's, around 2005. It
  6  was focused on the BacT/ALERT device. The
  7  device, as I mentioned, was cleared for a Q/C
  8  indication but the manufacturer did not pursue
  9  a release indication.
 10   It fell onto the manufacturers of
 11  blood storage bags, Gambro and Fenwal, to take
 12  up this indication or take up this objective.
 13  They started out by validating their own bags
 14  to show that they can store platelets up to
 15  seven days.
 16   Then Gambro went on to collect
 17  available performance data on the Q/C
 18  application of the BacT/ALERT system and
 19  submitted this data to the FDA with the intent
 20  to clear the system, meaning clear the bags as
 21  well as the bacterial detection device for
 22  storage of platelets up to seven days.
      Page 40
  1   Part of this clearance was that
  2  the company, Gambro, committed to a post-
  3  market study of BacT/ALERT performance for
  4  detection of contaminated platelet products
  5  stored to seven days in the Gambro collection
  6  container. This became known as the PASSPORT
  7  study. Subsequently, Gambro was joined by
  8  Fenwal in the study thus permitting
  9  cooperating users of the Fenwal collection
 10  container to release seven-day platelets.
 11   Now, the PASSPORT study was
 12  designed to validate the performance of the
 13  BacT/ALERT bacterial detection device by
 14  retesting platelet products at outdate.
 15  During the study products were tested at day
 16  one and the ones that were negative were
 17  placed into clinical use and the outdated
 18  products were retested at outdate to make sure
 19  they were still negative based on the initial
 20  determination by the device.
 21   Unfortunately, approximately one-
 22  and-a-half years after initiation of the
      Page 41
  1  PASSPORT study it was terminated over concerns
  2  of increased bacterial contamination of seven-
  3  day platelets and platelet storage reverted
  4  back to five days.
  5   What the PASSPORT study pointed
  6  out along with several other studies is that
  7  the bacterial detection as currently applied
  8  is missing contaminated products. Several of
  9  these studies indicate that testing with the
 10  BacT/ALERT device during the first day post
 11  collection identifies only a fraction of the
 12  contaminated products.
 13   There was an American Red Cross
 14  study that looked at five-day-old platelets.
 15  They were tested with a BacT/ALERT device and
 16  documented an increased septic reaction rate.
 17  There was a PASSPORT study, as I mentioned,
 18  and this documented higher than expected
 19  contamination rate of the retested products at
 20  outdate.  
 21   Finally, there was an Irish Blood
 22  Bank study on seven-day platelets. They were
      Page 42
  1  also retested on day four and documented a
  2  high contamination rate at the outdate of day
  3  seven. These three studies suggested the way
  4  we are applying these bacterial detection
  5  devices is not sufficient to capture all
  6  contaminated platelet products.
  7   The objective for today's
  8  discussion is to discuss ways to apply
  9  existing bacterial detection technology to
 10  reduce the bacterial risk of six and seven-day
 11  platelets and bring seven-day platelets back
 12  to the market.
 13   As you are contemplating some of
 14  these issues that we are going to hear today,
 15  I wanted to make sure that you thought about
 16  some of these things that are needed to
 17  consider for successful detection of
 18  bacterially contaminated platelet units.
 19   First of all, the initial level of
 20  contamination at the time of collection is
 21  very low and it is not exactly clear how low
 22  it is but it could be one to ten bacteria per
      Page 43
  1  bag and one bacteria is called 1 CFU, or
  2  colony forming unit.
  3   The bag volume ranges from 250 to
  4  400 ml. The initial bacterial concentration
  5  in the bag at the time of contamination could
  6  be very low in the order of 0.01 CFU/ml. Even
  7  though it's such a low initial contamination,
  8  during storage the bacteria can grow to levels
  9  of greater than 10 to the 12 CFU/ml.  
 10   The sensitivity of the culture-
 11  based devices is on the order of one to five
 12  CFU/ml. The strategy for successful detection
 13  need to take into account the device
 14  sensitivity for both aerobes and anaerobes
 15  bacteria, the amount of volume sampled from
 16  the product, and also the time for bacteria to
 17  proliferate into detectable range. This could
 18  vary for the slow-growing bacteria versus the
 19  fast growing bacteria.
 20   Just to graphically demonstrate
 21  the issue that is facing these devices, the
 22  way to detect contaminated platelet products
      Page 44
  1  is to be able to avoid sample error at the
  2  beginning. This is a hypothetical situation
  3  where you have a bag that is contaminated with
  4  a very low level of bacteria on day one. You
  5  can see that the bacteria will proliferate
  6  into a very high bacterial load at day seven.
  7   Now, if you are trying to detect
  8  bacteria on day one you have to be fortunate
  9  enough to actually capture a bacteria in your
 10  sample. If you don't manage to do that, then
 11  the device has no chance of detecting a
 12  contaminated unit. Your chances of detecting
 13  a contaminated sample improve over the time of
 14  storage and by day two and day three your
 15  chances of detecting a contaminated unit
 16  improves substantially.
 17   This is also demonstrated in this
 18  graph which compares the strategies for fast
 19  and slow growing bacteria. Again, this is the
 20  hypothetical growth curve. Here is the
 21  storage days for platelets. Here is the
 22  bacterial load inside the bag. Bacteria start
      Page 45
  1  off with very low contamination here and they
  2  proliferate for fast growing bacteria within
  3  a day or two.
  4   So if this is the sensitivity of
  5  your bacterial detection device, if you are
  6  sampling at day one, you're likely to pick up
  7  one of these fast growing bacteria. But this
  8  other one being that it's slower growing based
  9  on bacterial species, or based on the
 10  conditions in the platelet bag such as some
 11  plasma products or some platelet products
 12  support bacteria better than others, based on
 13  that the growth may be just a little bit
 14  slower and would cause the device to miss this
 15  contaminated bag.
 16   The situation gets even worse with
 17  slow growing bacteria that don't start to
 18  proliferate until days three and four. If you
 19  are sampling, let's say, at day three or day
 20  four, you are stretching the ability of the
 21  device to detect a contaminated unit. You
 22  might have to wait until day five to seven to
      Page 46
  1  actually have a better chance of detecting
  2  that.
  3   A few other additional points to
  4  consider for today is what is the optimal way
  5  to monitor the performance for sensitivity of
  6  the day one culture and this would be a
  7  discussion on whether it is appropriate to
  8  have a confirmatory culture at outdate versus
  9  a septic transfusion reaction of the patient
 10  population that is transfused with these
 11  products.  
 12   Finally, what is the appropriate
 13  intervention that will assure the safety of
 14  day six and seven platelets if you decide to
 15  put these products back on the market.
 16   Thank you very much. These are
 17  some of the questions that you will be -- that
 18  we will be asking you later. One of them is
 19  does the Committee agree with the FDA that the
 20  reporting of sepsis should be active and not
 21  passive?
 22   Another question is in addition to
      Page 47
  1  the reporting of sepsis does the Committee
  2  agree with the FDA that: (a) additional
  3  aerobic and anaerobic cultures should be
  4  performed on day five both to increase the
  5  safety of platelets on days six and seven and
  6  as a baseline measure? Finally, (b)
  7  surveillance cultures should be performed at
  8  outdate after day seven to provide a
  9  bacteriological endpoint for this study.
 10   With that introduction we have a
 11  panel of distinguished speakers that will
 12  actually provide further data on these very
 13  issues. Thank you very much.
 14   DR. SIEGAL: Thank you, Dr.
 15  Vostal.
 16   We will now hear from Mark Brecher
 17  from the University of North Carolina School
 18  of Medicine, Issues and Detection of
 19  Bacterially Contaminated Platelet Products.
 20   Dr. Brecher.
 21   DR. BRECHER: Thank you. I was
 22  asked to provide some background and go over
      Page 48
  1  some of the current issues about bacteria
  2  contamination of platelets. A couple of
  3  potential conflicts. I actually don't own any
  4  shares in any company like my family only has
  5  two shares of stock. Each of my daughters has
  6  one share of Disney stock.
  7   We have done a great job in
  8  reducing risk from various transfusion
  9  transmitted diseases. HIV, which used to be
 10  1 percent of units in the early '80s, is now
 11  down to one in 2 million. But for the longest
 12  time we really didn't do much about bacterial
 13  contamination of platelets which through
 14  multiple studies generally using aerobic at
 15  outdate found rates of about one in 2,000
 16  units were bacterially contaminated.
 17   Through the years bacterial death
 18  from contaminated units has been one of the
 19  leading causes of death from transfusion.
 20  Sixty cases over 10 years from bacteria
 21  contaminated platelets so roughly six cases
 22  per year the majority being gram negative
      Page 49
  1  organisms. This is a repeated finding from
  2  multiple countries that the bugs that kill
  3  people roughly two-thirds of them are gram
  4  negative organisms.
  5   I was heartened by the data that
  6  Dr. Holmberg presented this morning that in
  7  2007 the FDA only had three cases of bacterial
  8  deaths if you exclude the Babesia. It looks
  9  like we certainly are heading in the right
 10  direction but it's thought that many cases go
 11  unrecognized and that this is simply the tip
 12  of the iceberg.
 13   Just to remind people about the
 14  different types of platelets, this is somewhat
 15  of an old slide but blood can be donated and
 16  it goes into a receptacle. In the old days it
 17  used to be a glass bottle. We generally don't
 18  do direct donor to recipient transfusions but
 19  comic books have historically used this as a
 20  plot device in many cases for the passive
 21  transfer of super powers.  
 22   If you are keenly paying
      Page 50
  1  attention, you are going to ask yourself how
  2  did they get the needle through Superman's
  3  skin? They do explain this. They have a
  4  doctor here saying, "Thank you for puncturing
  5  your skin with your fingernail," which raises
  6  all kinds of questions about super infections.
  7   The other way to -- this is how we
  8  would make random platelets from a whole blood
  9  donation. The other way is to hook up a donor
 10  to an apheresis machine, some of the older
 11  Fenwal CS-3000s, and just selectively take off
 12  a therapeutic dose of platelets from a donor.
 13  See this happy donor here. I like this donor
 14  so much I married her.
 15   Where were we just a few years
 16  ago? Roughly 4 million platelet bags were
 17  transfused per year in the U.S. Roughly a
 18  million apheresis and 3 million random
 19  platelets with a contamination rate of 1 in
 20  1,000 to 1 in 2,000. There were 2,000 to
 21  4,000 bacterially contaminated bags that we
 22  were handing out and transfusing every year.
      Page 51
  1   People often say, "Well, most of
  2  those wouldn't have caused sepsis." There is
  3  some estimates that as low as 1 in 10 may have
  4  caused clinical sepsis. If you look to the
  5  literature the number is probably higher. It
  6  depends on whether you're talking about gram
  7  positive or gram negative organisms.  
  8   There is some data from the
  9  University Hospital of Cleveland from Roslyn
 10  Yomtovian and Mike Jacobs that they for years
 11  cultured platelets and they screened them with
 12  a gram stain. When they pulled out the most
 13  heavily contaminated units with the gram
 14  stain, they still had a 40 percent incidence
 15  of clinical sequelae in recipients.  
 16   Probably the real answer is
 17  somewhere in here but maybe a little but
 18  higher but we're talking about 200 to 1,600
 19  cases of clinical sepsis. Not all of them
 20  will die again depending on the gram positive
 21  or gram negative.  
 22   Maybe one-fifth to one-third will
      Page 52
  1  result in a fatality. We are talking about
  2  perhaps 40 to 500 deaths per year from
  3  bacterial contaminated platelets or 1 in 7,500
  4  to 1 in 100,000 fatalities per unit. These
  5  are a lot of cascading assumptions. Is there
  6  any validity to this? You can again go to the
  7  literature.  
  8   Data from Johns Hopkins found that
  9  with a pool of six random platelets there was
 10  a rate of 1 in 17,000. For apheresis platelet
 11  it was roughly 1 in 68,000. Very similar to
 12  data from the University Hospital of Cleveland
 13  of 1 in 58,000 for apheresis and from France
 14  1 in 150,000.
 15   So are there a lot of assumptions
 16  going down this cascade. They do seem to be
 17  real so there was probably about 100 to 200
 18  deaths per year from bacterially contaminated
 19  platelets.
 20   Some of the data from the
 21  University Hospital of Cleveland they had 32
 22  recipients of which 13 had reactions of 41
      Page 53
  1  percent, nine had severe reactions, 28
  2  percent. In three cases the patients died so
  3  that was 10 percent. Again, they screened out
  4  the most heavily contaminated units so you
  5  would have expected that the fatality rate
  6  would have been higher.
  7   The FDA has convened several
  8  meetings through the years to discuss
  9  bacterial contamination. This was a summary
 10  from Ed Synder from Yale at the end of a
 11  workshop in 1999 where he felt that the sense
 12  of the meeting was the imperative was to act.
 13  You don't have to explain yourself on
 14  Nightline. I have often said we just have to
 15  wait until a celebrity dies and then it's
 16  going to hit the fan.
 17   Regulation is necessary to achieve
 18  the goals. Nothing says I care like a page of
 19  483s. Those are the deficiencies that the FDA
 20  hands out on their inspections. When all else
 21  fails do something. Give us a mandate and we
 22  will do the rest.  
      Page 54
  1   There was a feeling in the blood
  2  banking industry that we wanted to do
  3  something but our hospital administrators
  4  wouldn't let us do something unless there was
  5  a mandate to do it because it would cost
  6  money.
  7   In 2002 things began to change.
  8  There were two companies that brought to
  9  market quality control testing. One was the
 10  bioMerieux BacT/ALERT which is an automated
 11  liquid culture system that has color metric
 12  sensors at the bottom of the bottles that as
 13  C02 is generated in the bottles the color
 14  changes from, I guess, green for go to yellow
 15  for caution.  
 16   This method looks at both the
 17  absolute color change and the rate of change
 18  so there is a little bit of applied calculus.
 19  Much of the data was generated in my lab for
 20  this submission. That was in February of
 21  2002. I think it was in September or October
 22  of 2002 the Pall Bacterial Detection System,
      Page 55
  1  BDS, was brought to market.  
  2   This looks at the P02 in the head
  3  space of a bag so as the bacteria grow they
  4  consume the oxygen. I used to say it couldn't
  5  be a Pall product if it didn't have a filter
  6  so there was the filter. The problem with
  7  this first model was that the filter took out
  8  50 percent of the bacteria.  
  9   That wasn't a good thing. They
 10  modified it. They took the filter out and
 11  made a few other changes and they had the
 12  enhanced bacterial detection system on the
 13  market.
 14   Another meeting with the FDA
 15  addressed bacteria and a variety of other
 16  issues but mostly pathogen reduction occurred
 17  in the summer of 2002. After the meeting
 18  several of the speakers and moderators got
 19  together and they issued a public letter to
 20  the blood banking industry.  
 21   In this letter they said that
 22  pathogen reduction isn't going to be here
      Page 56
  1  anytime soon. This is Jim Aubuchon from
  2  Dartmouth; Ros Yomtovian, University Hospital
  3  of Cleveland; Mo Blajchman, McMaster; Paul
  4  Ness, Johns Hopkins, and myself.  
  5   We said that bacteria
  6  contamination is a problem. We can do
  7  something about that and the blood banking
  8  industry needs to go ahead and do that. A lot
  9  of times people write these sort of open
 10  letters and nothing much ever happens but this
 11  letter really seemed to be a tipping point in
 12  bacteria contamination and it garnered a lot
 13  of attention.
 14   Shortly thereafter the two
 15  voluntary accrediting agencies in the U.S.,
 16  the American Association of Blood Banks and
 17  the College of American Pathology issued
 18  standards. The ABB, the blood bank or
 19  transfusion service, shall have methods to
 20  detect bacterial contamination.  
 21   From CAP the laboratory shall have
 22  a system to detect the presence of bacteria in
      Page 57
  1  platelet components. That is really what got
  2  things moving. We finally had a mandate from
  3  someone that we had to do something.
  4   What some of you may not know is
  5  that right before the ABB standard went into
  6  effect in March 2004 the Acting Assistant
  7  Secretary of Health, Christina Beato, wrote a
  8  letter asking that it be delays because they
  9  were worried about its affects on the
 10  availability of platelets. This is really at
 11  the 11th hour. This is, I think, February
 12  24th was the date on this.
 13   The ABB responded that further
 14  delaying implementation will compromise
 15  patient safety and public health. The country
 16  did go on to implement bacterial detection.
 17  ABB task force looked at the impact shortly
 18  after the implementation of the testing and
 19  they found that for the majority of blood
 20  centers, hospital blood banks and transfusion
 21  services that their ability to provide
 22  platelets was not really effective after the
      Page 58
  1  implementation.
  2   Similarly, the majority said they
  3  were not experiencing increased platelet
  4  outdating so for the majority of centers it
  5  went down pretty well. One interesting fact
  6  that came out of the survey was that for
  7  apheresis platelets the vast majority of
  8  people were doing one of the culture methods.
  9   However, for random platelets most
 10  people were doing non-culture surrogate
 11  methods such as monitoring the pH of the
 12  random unit or the glucose often using urine
 13  dip sticks which is a very insensitive method.
 14  I say this and I'm one of the ones who put it
 15  in the literature but it only picks up about
 16  10 to the 7th colony forming units per ml.
 17   The other interesting thing, the
 18  true positive pickup rate was about 1 in
 19  4,000. You will see that using early culture
 20  of 1 in 4,000, 1 in 5,000 seems to be the
 21  recurring number that we are picking up. With
 22  the non-culture method it was only one in
      Page 59
  1  roughly 18,500 or a 4.6 fold difference in
  2  sensitivity.  
  3   We know from many culture studies
  4  that the contamination rate per bag should be
  5  the same so clearly this method of using pH
  6  and glucose was not really addressing a
  7  majority of the problem.
  8   I just wanted to mention that the
  9  College of American Pathology has a proposal
 10  on the table to change their standards in 2009
 11  where they will no longer allow for
 12  insensitive methods such as swirling pH or
 13  glucose.  
 14   The sensitivity of the method must
 15  be at least 10 CFU/mL 24 hours after the
 16  collection or at least 10 to the 5th CFU/mL 72
 17  hours after collection. Hopefully this will
 18  get to the dichotomy of safety that we
 19  currently have.
 20   Now, there are several ways that
 21  you can reduce the risk of bacterial
 22  contamination, one of which is going to all
      Page 60
  1  apheresis platelet supply. At Johns Hopkins
  2  in the mid 80's 52 percent of their platelets
  3  were apheresis platelets, 48 percent were
  4  pooled random platelets.  
  5   They made an active effort to go
  6  to 100 percent apheresis platelets. By 1998
  7  99 percent of their platelets were apheresis
  8  platelets. The reaction rate during this time
  9  period went from 1 in 5,000 to 1 in 15,000.
 10   When you teased out the
 11  differences between random platelets and
 12  apheresis platelets there is about a 5.6 fold
 13  difference in reaction rate. Don't forget
 14  they were transfusing six packs, pools of six.
 15  It, again, gets back to that the risk is per
 16  bag of platelet product.
 17   One of the other things that came
 18  out of the survey is we went back through the
 19  literature and we plotted out what percentage
 20  of apheresis platelets were being transfused
 21  in this country. Interestingly it made almost
 22  a perfect straight line so that by 2004 we
      Page 61
  1  were at roughly 79 percent of all doses handed
  2  out in this country were apheresis platelets.
  3  Jim Aubuchon extended my line saying that by
  4  2010 we'll be almost all apheresis platelets.
  5  I think that is a bit optimistic.  
  6   The most current data, preliminary
  7  data from a survey of transfusion practice inn
  8  2006 estimates that we are currently sitting
  9  around 88 or maybe 89 percent of all platelets
 10  handed out in this country are apheresis
 11  platelets. I'm not sure when we are going to
 12  cross the line of standard of care. I'm not
 13  sure what is the standard of care. Is it 95
 14  percent? I'm not sure.
 15   Other interventions that we've
 16  tried is diverting the first volume of the
 17  donation. We say we sterilize the skin. We
 18  don't really sterilize the skin with iodine
 19  solutions. What we do is we do a bacterial
 20  load reduction. You cannot get all the
 21  bacteria off the skin. There are bacteria in
 22  the hair follicles, spacious glands, sweat
      Page 62
  1  glands that the iodine never reaches. A
  2  needle is going through there and is making a
  3  core and pulling that up the needle.
  4   However, data from a variety of
  5  studies and I just picked two. This one is
  6  from France where the first 15 mL 76 were
  7  contaminated but if they diverted the second
  8  15 mL only 21 of the 76 were contaminated.
  9   Study from the Netherlands.
 10  Actually, the author is out in the audience.
 11  18,000 collections, .35 percent were
 12  contaminated. If they diverted the first 10
 13  percent it dropped to .21 percent.
 14   You can decrease the bacterial
 15  contamination in the bag. However, as Ros
 16  Yomtovian likes to say, we should not be
 17  diverted by diversion. Two-thirds of the
 18  fatalities are gram negative organisms. What
 19  we are dealing with here are mostly gram
 20  positive organisms.  
 21   Yes, we can decrease gram
 22  positives but we are not really attacking
      Page 63
  1  those bugs that cause death which are mainly
  2  gram negatives and are thought to principally
  3  come from donors who has a asymptomatic
  4  transient bacteremia.
  5   The other thing with putting a
  6  diversion pouch on a bag is that you would
  7  think it would be a very simple thing to do
  8  but it has had its own set of problems. Those
  9  first 15 mL or so of blood are often then used
 10  to do your viral tests. Even that had
 11  problems.  
 12   Recent data from the American Red
 13  Cross published last year where they looked at
 14  the true positive contamination rate using an
 15  early culture found that with two-arm
 16  apheresis procedures the rate of bacterial
 17  contamination was twice as high as a single-
 18  arm procedure. The COBE Trima here only does
 19  a single-arm procedure. It came to light that
 20  the diversion pouch of both of these
 21  manufacturers was put on the wrong line.
 22   It was put on the return line, not
      Page 64
  1  the draw line, so the bacteria coming up into
  2  the collection set was contaminating your
  3  collection set and then it was being diverted.
  4  Little things. Here is a picture Richard
  5  Benjamin provided of that diversion pouch that
  6  you see here which is actually on the return
  7  line. These sets have been modified.
  8   So the question is have we really
  9  done anything. Have we accomplished anything
 10  with the bacterial culturing of early units.
 11  The first large paper in this country was from
 12  the American Red Cross and they looked at the
 13  first 10 months of having bacterial culture
 14  versus the 10 months prior where they were not
 15  doing bacterial culture.  
 16   What they found was with the
 17  septic reactions -- that's not good. From
 18  March through December 2003 they had 15 septic
 19  reactions, 12 were high probability, two were
 20  fatal. In the same period there were only
 21  eight involving apheresis but only three were
 22  such a high probability so it was a 75 percent
      Page 65
  1  drop in high probability septic events.  
  2   Well, that was pretty good.
  3  Subsequent data from the Red Cross looking at
  4  a longer time period suggested maybe only 50
  5  percent. Clearly it seems like we impacted on
  6  the number of septic transfusion reactions.
  7   However, those reactions that do
  8  occur tend to occur on older units. What we
  9  don't know, however, is how the breakdown of
 10  the yes is in this country. For example, here
 11  we have 13 cases that occurred on day five but
 12  we don't know, for example, are 60 percent of
 13  the platelets transfused on day five and only
 14  10 percent on day two?  
 15   We don't really know the
 16  denominator for these cases. Again, it's
 17  worrisome that the older the unit the more
 18  risk of a septic transfusion reaction. What
 19  we also don't know is are the bacteria that
 20  are slow growing are they plateauing around
 21  day five. We don't know if day six or day
 22  seven platelets would actually have a higher
      Page 66
  1  septic rate per unit.
  2   This is data from Hema-Quebec
  3  putting in a variety of interventions. They
  4  were measuring the septic transfusion reaction
  5  rate in thousands. The line stopped here
  6  because in this presentation of ABB they
  7  didn't have anymore reactions. As far as they
  8  could tell 100 percent had been presented by
  9  all these interventions.
 10   This is data looking at 123,000
 11  apheresis platelets from blood systems, the
 12  second largest collector of blood in this
 13  country. What they found in the 24-month
 14  interval that there had been three known
 15  septic reactions in the time period before but
 16  none after. We are clearly preventing many of
 17  these cases.
 18   Data from the Red Cross. Their
 19  septic transfusion reaction rate was 1 in
 20  40,000. When they began culturing it dropped
 21  to 1 in 75,000. After they added diversion in
 22  their current estimates are now 1 in 175,000
      Page 67
  1  so we are impacting.
  2   Data using the Pall eBDS is a
  3  little harder to find but here is a report
  4  from 118,000 apheresis and random platelets
  5  from 23 U.S. blood centers. Their true
  6  positive contamination rate was 1 in 5,000,
  7  very similar to using the BacT/ALERT. There
  8  was one reported case of a missed staph epi so
  9  this also seems to be effective.
 10   This is combined data from the Red
 11  Cross and blood systems looking at 1.23
 12  million apheresis products, roughly a year's
 13  worth of apheresis platelets in this country,
 14  207 isolates many of which are gram negatives,
 15  chierchia, E. coli that invariably would have
 16  killed people. Are we saving lives? There is
 17  no question in my mind we have saved a lot of
 18  lives by doing these interventions.
 19   However, there is some data that
 20  continues to be worrisome. This has already
 21  been alluded to. This is a paper from Murphy
 22  from Ireland. What they found was that when
      Page 68
  1  they did their early culture they detected 35
  2  cases out of 42,000 or .08 percent.  
  3   Another 3,000 were recultured on
  4  day four and they picked up another .12
  5  percent. Of 8,000 that outdated they found 18
  6  that were positive or .22 percent. They said
  7  that the early culture only had a sensitivity
  8  of less than 40 percent which was not optimal
  9  at all.
 10   However, when you look at papers
 11  like this you need to understand the full
 12  picture. What they do in Ireland is not what
 13  we do here. They, for example, culture their
 14  apheresis platelets after 12 hours from
 15  collection. We wait at least 24 hours,
 16  sometimes 36 hours before we take our sample
 17  to minimize the sampling error.  
 18   I'm not sure you can carry this
 19  data forward. The other thing is they are not
 20  looking at the transfusion septic reaction
 21  rate whereas we are. They are just looking at
 22  the culture positive rate. Some of these bugs
      Page 69
  1  probably wouldn't hurt anybody if they had
  2  gotten them.
  3   Another vexing problem that
  4  remains is that of anaerobic bacteria. There
  5  have been a few cases of sepsis and death from
  6  anaerobic organisms from platelets and from
  7  red cells. This is from the literature. Two
  8  cases of clostridium. The FDA is aware of
  9  another couple cases of clostridium from red
 10  cells, clostridium from platelets and U
 11  bacterium from platelets. Use of anaerobic
 12  model could potentially interdict these few
 13  cases, these rare cases.
 14   The other interesting thing about
 15  anaerobic model it's a different media than
 16  was in the aerobic model. Some organisms like
 17  that media better. For example, if you look
 18  at various strep organisms from the data from
 19  my lab, strep pyogenes the aerobic model takes
 20  19 hours, the anaerobic model 13.8 on average.
 21   Strep viridans 43 hours in the
 22  aerobic model, 21 in the anaerobic model. I
      Page 70
  1  have argued for a long time that difference in
  2  time may make the difference between whether
  3  you are able to interdict a unit or not.
  4   There is also some data at low
  5  concentrations of some bacteria. This is a
  6  staph lugdunensis that actually killed a
  7  recipient. At low concentrations the
  8  anaerobic model seems to be much faster than
  9  the -- the anaerobic model seems to be much
 10  faster than the aerobic model.  
 11   This is looking at a preparation
 12  of 1.5 CFU/ml. We put 1 mL into a bottle, 2
 13  mL, 3 mL, 4, 5, 6, 8, and 10. Then we made up
 14  the volume difference with saline so there was
 15  no dilutional effect.
 16   There is a suggestion that at very
 17  low concentrations the anaerobic model may be
 18  even more sensitive. Here we had three out of
 19  three that came up positive in the anaerobic
 20  model but in the aerobic model only one of
 21  three came up positive.
 22   Okay. Other initiatives that move
      Page 71
  1  toward rapid detection. This was discussed at
  2  BPAC in March 2006. Several companies then
  3  were pursuing this. Immunetics has a
  4  peptoglycan based assay. GloBac looking at
  5  bacterial ATP there are two systems that use
  6  lateral flow devices.  
  7   Verax is one that is currently
  8  approved and you've already heard about that
  9  today. There was one other company that
 10  actually had an interesting method that used
 11  bacillus spores that would fluoresce if there
 12  was another bacteria nearby but they have sort
 13  of dropped out of the race.
 14   At this meeting Ros Yomtovian and
 15  Mike Jacobs presented their data on their long
 16  surveillance at University Hospital of
 17  Cleveland. They have subsequently gone on and
 18  published this experience in two different
 19  papers, same figure in both papers.
 20   What they presented was that at 10
 21  to the 5th CFU/mL it would have prevented all
 22  fatal reactions, 91 percent of serious
      Page 72
  1  reactions, 79 percent of all reactions. 10 to
  2  the 3rd would have prevented all serious
  3  reactions, 79 percent of all cases and 95
  4  percent of all reactions.  
  5   It's from data like this that the
  6  feeling is that we need to have a sensitivity
  7  of around 10 to the 4th CFU/mL for a rapid
  8  test. They were really only evaluating for
  9  acute reactions.
 10   Random platelets you could pool
 11  them. They used to only be good for four
 12  hours. Acrodose System came online in 2005
 13  with FDA approval. You can now pre-pool and
 14  keep the entire shelf life at the banks.
 15   Preliminary data from the Red
 16  Cross shows a 5.8 fold higher true positive
 17  rate of cultures compared to apheresis
 18  platelets so this is worrisome. We don't
 19  really have separate transfusion reaction
 20  data. I think this is a worrisome
 21  development.
 22   Cases continue to slide through.
      Page 73
  1  MMWR reporting some cases in 2004. The CDC
  2  sort of dinged the Red Cross for not following
  3  the manufacturer's recommendations regarding
  4  volume for testing in one case. This is a
  5  case from Kansas City. It's an E. coli death.
  6   In this case they have their
  7  BacT/ALERT cabinets in a back room. They
  8  loaded the bottles on Friday, closed the door,
  9  and walked away for the weekend. They had a
 10  computer interface but the computer interface
 11  went down.  
 12   There is a flashing red screen on
 13  the BacT/ALERT if there is a positive
 14  detection but, of course, no one could see it.
 15  There is an audible alarm. They turned off
 16  that audible alarm. They walked in Monday
 17  morning, saw that the machine was flashing,
 18  tried to call back that unit but it had been
 19  transfused within the previous hour at a
 20  hospital so mistakes can still happen. No
 21  system is going to be perfect.
 22   This is a case of streptococcal
      Page 74
  1  death from Florida that is about to be
  2  published where they were actually culturing
  3  the random platelets by taking a bit of the
  4  tubing and pulling it with several other units
  5  together but they were doing their sampling
  6  two hours after the production of platelets.
  7   They weren't waiting for the 24
  8  hours. There is no surprise that there is a
  9  sampling error that they missed. Again, they
 10  were not following the package insert
 11  recommendations for culturing which is to wait
 12  at least 24 hours.
 13   Another thing that worries me is
 14  that there are cases that we are just not
 15  aware of that are slipping through our
 16  fingers. This is a paper from the NIH
 17  Clinical Center from 1973 where one year they
 18  noticed they had all these salmonella cases
 19  being reported.  
 20   They brought the CDC in and did
 21  this being epidemiologic search. Long story
 22  short they found out that every patient --
      Page 75
  1  when they checked further everyone had a
  2  platelet transfusion from the same donor.
  3   Working up the donor extensively
  4  they found he had osteomyelitis. The
  5  interesting thing is how many days from the
  6  transfusion until they became sick. It was on
  7  average 8.6 days. By then people had
  8  forgotten they had even given a platelet. It
  9  worries me there may be other cases out there.
 10  There were several deaths, multiple
 11  recurrences of the salmonella.
 12   Similarly, just a few years ago
 13  there was a recall of a couple lots of Heparin
 14  that was in a catheter set that was
 15  contaminated with Pseudomonas. CDC followed
 16  up on many of the people who had received this
 17  contaminated Heparin and it turned out that
 18  there were 15 patients in Michigan and 13 in
 19  South Dakota who had a delayed onset of
 20  Pseudomonas fluorescence.  
 21   This ranged from 84 days to 421
 22  days. If the CDC hadn't been following up, do
      Page 76
  1  you think anyone would have realized that it
  2  came from the Heparin that they used a year
  3  ago? There is a concern that any bacteria
  4  could be bad bacteria.
  5   Pathogen reduction we already had
  6  mentioned this morning. There was a Canadian
  7  conference last year where the FDA finds it
  8  problematic which is why we don't have
  9  pathogen reduction yet in this country and the
 10  Canadian regulators had basically a similar
 11  opinion.
 12   I'm running out of time. Across
 13  the world most people are using the
 14  BacT/ALERT. Many countries have gone to seven
 15  days. A few notable exceptions. Japan is not
 16  doing any bacterial detection. They were only
 17  keeping their platelets for three days. I
 18  understand now it's four. France is gradually
 19  implementing pathogen reduction.
 20   Okay. Several years ago I was
 21  invited to speak at a workshop on bacterial
 22  contamination and it was entitled, "Bacterial
      Page 77
  1  Contamination. So Have we Missed the Boat?"
  2  I thought that was the wrong -- where is my
  3  sound? Let's try this again. No sound.
  4  Okay. Let's turn the volume up. Thank you.
  5  Okay. Sorry for this technological delay.
  6   Okay. I thought that was the
  7  wrong question. This is the first
  8  undocumented use of this well-known phrase,
  9  "Have we missed the boat." I thought that was
 10  the wrong question. The better question is do
 11  we know where we're going?
 12   (Whereupon, a video was played.)
 13   DR. BRECHER: So it's important
 14  that you have to know where you're going. By
 15  the way, if you're curious the subtitles were
 16  in Swedish. This is a company that makes GPS
 17  navigation gear.
 18   Do we know where we're going?
 19  We're going toward better patient safety.
 20  Have we done that? No question. We have
 21  saved many lives from what we've already done.
 22  Now I'm done. Sorry for running a little
      Page 78
  1  over.
  2   DR. SIEGAL: Thank you, Dr.
  3  Brecher.
  4   Do we want to take questions at
  5  this point or go on? Let's go on for now.
  6  Next is Thomas Montag-Lessing, M.D. from Paul-
  7  Ehrlich Institute on bacterial detection in
  8  blood components.
  9   DR. MONTAG-LESSING: Mr. Chairman,
 10  ladies and gentlemen, I am pleased to have
 11  this opportunity to speak here in front of the
 12  Blood Products Advisory Board. My topic is
 13  bacterial detection in blood components.
 14  Since Jay Epstein asked me to report something
 15  about our work in the past 10 years regarding
 16  blood standards, of course we do it with
 17  pleasure.
 18   Considering the introduction of
 19  Dr. Vostal and the excellent review of Mark
 20  Brecher, there is no need for me to discuss
 21  residual risk and whatever. I would like to
 22  start immediately with the question what is
      Page 79
  1  PEI. PEI means Paul-Ehrlich Institute.  
  2   Ten years ago we understood that
  3  the usual reference strains coming or used in
  4  microbiology, for instance, coming from the
  5  so-called from the ATCC, American Type Culture
  6  Collection, cannot automatically be used in
  7  blood components since a lot of them are not
  8  able to multiply to grow up or they remain
  9  without any reaction and so on so we started
 10  to collect strains which are able to grow up
 11  in platelet concentrates.
 12   The next question we asked as
 13  microbiologist is are you able to grow up in
 14  platelets from as much as possible different
 15  donors in order to exclude impairments or any
 16  interferences from the immune systems of the
 17  different donors. So our strain standards are
 18  characterized in platelets from at least a
 19  hundred different donors.
 20   The next step is following a
 21  specially developed procedure our standards
 22  are deep frozen and ready to use, stable, and
      Page 80
  1  shapeable and so they are defined and consist
  2  mainly of living cells. That is the
  3  prerequisite that we are able to perform real
  4  life spiking of blood components.  
  5   That means contamination of one
  6  platelet bank with 10 corresponding to .03 per
  7  mL. That point had been mentioned already by
  8  Jaro Vostal. Under real life conditions that
  9  means imagining the actual level is acceptable
 10  in the given blood bank. This is a real life
 11  contamination. That's what microbiologists
 12  don't think at all but our approach in low
 13  saturation mimicking or the simulation of real
 14  life contamination.
 15   In consequence, PEI standards are
 16  two to four and that is very important.
 17  Objective validation and assessment of methods
 18  for screening and especially for pathogen
 19  reduction methods because it is very important
 20  to have objective data after validation in
 21  order to assess these approaches.
 22   Two years ago we proposed a WHO
      Page 81
  1  whether they are interested in installation of
  2  these bacteria standards, WHO bacteria
  3  standards, since there is no reference
  4  material available at all following a proposal
  5  of my colleague who is in the audience. We
  6  just started a discussion with the American
  7  Type Culture Collection to involve them in the
  8  current process.
  9   Inside the working party for
 10  transfusion transmitted infectious diseases
 11  from the International Society of Blood
 12  Transfusion. The bacteria group is a subgroup
 13  of the working party. We agreed last year
 14  that we should perform in collaboration
 15  between WHO and ISBT an international
 16  validation study in order to characterize the
 17  strains as a crucial prerequisite for
 18  installation of the strains as WHO standards.
 19   Just for illustration I'm going to
 20  show you that we worked so-called PASSPORT of
 21  our blood standards in order to prevent any
 22  confusion with the PASSPORT study headed by
      Page 82
  1  Larry Dumont, of course, we will change the
  2  name to certificate but this is the former
  3  version as discussed in the last ISBT meeting
  4  in China in Macau.
  5   My post-doc included a photograph
  6  as a PASSPORT procedure. Okay. The
  7  certificates contain information regarding the
  8  gross conditions of the given strain in
  9  platelet concentrates under real life
 10  conditions. It's very important.  
 11   We learned only 10 years ago that
 12  there is no chance to reduce the volume of the
 13  platelet concentrate or to work with smaller
 14  batches. I like to call it two poly-
 15  dimensional systems which are interacting and
 16  there is no chance using my small brain I have
 17  to understand what is happening in such a
 18  complex situation.
 19   Okay. Furthermore, we included
 20  the information how the bacteria are growing
 21  on typical culture plates and what about the
 22  picture of the gram staining. Furthermore,
      Page 83
  1  our bacteria strains are genome sequenced.
  2  That means basing on the 16S small subunit
  3  ribosomal RNA bacteria can be identified to
  4  know exactly regarding their species.
  5   One further information. In order
  6  to prevent any mixing up or any change during
  7  different cultures we characterize our strains
  8  using the so-called randomly primed rapid PCR
  9  fingerprinting working with random primers
 10  which is giving a clear picture regarding
 11  stability of a given strain.
 12   That is the method actually used
 13  by the colleagues from forensic medicine who
 14  trained or whatever for identification of
 15  gangsters or whatever you would imagine.
 16   This is an illustration of a
 17  prevalidation study we did together with the
 18  National Guard Service in London with the help
 19  of Dr. Carl McDonald. I like to say these are
 20  the numbers on the left-hand side estimated
 21  regarding the count of bacteria in my lab and
 22  here are the results produced in the London
      Page 84
  1  lab and some of them are looking like
  2  certifications and we are very proud of the
  3  result.
  4   The plan which The National
  5  Validation Study should go on -- this is a
  6  list of partners involved up to now. We are
  7  happy that we could involve colleagues from
  8  Asia, from Hong Kong, up to colleagues from
  9  South Africa.  
 10   The study will start with the
 11  shipping and sending out of -- and that's in
 12  September 2008, this year. The next ISBT
 13  meeting is in March of next year in Cairo,
 14  Egypt. We will have a workshop which will
 15  discuss the results of this study.
 16   This PI blood standards hopefully
 17  in the future WHO blood standards can be used.
 18  I would like to start with one example
 19  regarding validation of pathogen reduction
 20  methods we did two years ago or two-and-a-half
 21  years ago with Cerus or, this time, the Baxter
 22  Company.
      Page 85
  1   Here we can see one experiment
  2  dealing with isolated spores from bacillus
  3  cereus. You know bacillus cereus belongs to
  4  the so-called spore forming bacteria and
  5  bacteria spores that are very, very stable
  6  because they are dry. They are heat stable
  7  and stable against irradiation and so on.
  8   The microbiologist wait when we
  9  spike platelet concentrates with altogether
 10  three times 10 to the 6th isolated spores for
 11  bacillus cereus corresponding to 10 to the
 12  four per mL. Then no spore will be killed
 13  after the treatment because, as I mentioned
 14  already, the spores are dry. Where there is
 15  no water there is no effusion and there is no
 16  biochemistry and that will be dead away.
 17   Surprising was the next experiment
 18  in which we tried to follow real-life
 19  conditions as mentioned already. That means
 20  we contaminated platelet bags with 10 spores
 21  per bag, stored the bags overnight as a usual
 22  procedure in a blood facility, and thereafter
      Page 86
  1  we performed treatment. As we can see here
  2  the spores survived. The products were not
  3  sterile.  
  4   The question, of course, was what
  5  about the lifestyle of the spore-forming
  6  bacteria in PC. There were no spores when we
  7  contaminated 10 spores per bag and store in
  8  the platelet bank overnight. Then the spores
  9  should germinate into vegetative forms.
 10   Vegetative forms of bacteria, of
 11  course, can be killed without any problem.
 12  Why did bacillus cereus survive? The
 13  explanation you can see here in this
 14  experiment we did in my department.  
 15   We contaminated platelet
 16  concentrates and parallel typical usual
 17  microbiological culture medium and followed --
 18  monitored all together 40 hours, following
 19  sample drawing every hour. This could be done
 20  only because it was summertime and the
 21  trainees were students so I could ask them for
 22  something so often all of the day up to 24
      Page 87
  1  hours.
  2   Okay. As you can see there was a
  3  very interesting result. All of the time we
  4  found in the PCs concentrate residual spores.
  5  The red points demonstrate what happened in
  6  the classical microbiological culture medium.,
  7  and me as a microbiologist I since studied one
  8  year microbiology and that is described in the
  9  textbooks.  
 10   Spores will germinate in
 11  microbiological culture medium and over a
 12  given time, in this case around 30 hours, I
 13  have vegetative cells only. In platelets
 14  there is heavier spores and the consequence is
 15  very clearly there is no chance to evacuate
 16  spore-forming bacteria by the intercept
 17  procedure or any other methods.
 18   We are just going to perform in
 19  October this year the same experiments with
 20  the riboflavin procedure by Navigant. The
 21  company has a new name but I'm always
 22  forgetting the new name.
      Page 88
  1   Okay. I would like to add the
  2  question are spore-forming bacteria relevant.
  3  A short look into the PubMed educates you that
  4  there are a lot of severe events up to fatal
  5  cases induced by spore-formers. Mark Brecher
  6  already mentioned the cause of klebsiella
  7  pneumonia, a lot of people are doing the same.
  8   Here I listed the frequency of
  9  spore-forming bacteria in several publications
 10  up to the number here mentioned in a paper,
 11  with one in 500. Then I remembered the cake
 12  shown by Mark Brecher here. There were around
 13  five percent spore-forming bacteria in the
 14  isolates.
 15   The reason is very easy to
 16  understand. The alcoholic disinfectants we
 17  are using because they are working very, very
 18  fast are not sporicidal at all so spore-
 19  forming bacteria, typical environmental
 20  bacteria can come into blood components.
 21   Coming to the validation of
 22  screening methods I would like to focus to
      Page 89
  1  rapid methods as already announced in the
  2  title. Allow me to repeat shortly what is the
  3  crux in bacteria screening of blood
  4  components. Usually contamination happens
  5  with very, very low counts of bacteria.  
  6   That means 10 to 100 CFU/mL
  7  corresponding to .03 to .3 CFU/mL. There are
  8  also mentioned .01 CFU/mL that can happen, of
  9  course, too. Thereafter the bacteria are able
 10  to grow up. I would like to repeat that we
 11  developed our bacteria standards in such a
 12  manner that we are able to imitate real life
 13  conditions in contamination.
 14   That is the extractum of what is
 15  happening after a bacteria entered a blood
 16  component during the blood donation. Starting
 17  with the count around .03 CFU/mL there is a
 18  given phase, an adaptation phase of the
 19  bacteria to the new environment as usually
 20  happens. Then there's the growth stage
 21  usually the geometric growth stage, and then
 22  the stationary phase is reached.
      Page 90
  1   I like to call this approach shown
  2  in detail by Mark Brecher Dutch approach
  3  because the Dutch colleagues were actually the
  4  first worldwide who installed platelet
  5  bacteria screening as mandatory nationwide in
  6  2001. We called it early sampling combined
  7  with culture methods and the idea was to say
  8  considering that the contaminating or the
  9  count of contaminated material is so low we
 10  should wait at least one day.  
 11   Let's give the contaminating
 12  bacterial strains a chance to start its
 13  multiplication or having the chance to have at
 14  least one bacterium in my bottle before
 15  analyzed. The two approaches available on the
 16  market are the BacT/ALERT machine and the Pall
 17  eBDS as demonstrated by Mark Brecher.  
 18   I would like to show you one of
 19  the cases we had to deal with the last time
 20  inside a National Red Cross study in Germany.
 21  One apheresis platelet concentrate consisting
 22  of two therapeutic units had been -- oh, thank
      Page 91
  1  you. Sorry. I didn't know. I hope you could
  2  hear me up to now.
  3   A sample had been formed from the
  4  apheresis units 20 hours after donation. Both
  5  samples were incubated into a BacT/ALERT
  6  machine as well as Pall eBDS the cartridges
  7  remained sterile and transfusion of the
  8  products happened. They produced two fatal
  9  cases in the patients.
 10   Because we have a very good
 11  contact list the colleagues from the
 12  University Hospital in Germany we could get
 13  the platelets back and we could analyze them
 14  in my department and we found in one 10 to the
 15  8th CFU/mL of klebsiella pneumonia and the
 16  second bag 10 to the 9th CFU/mL klebsiella
 17  pneumonia including tons of endotoxin of
 18  course you know, klebsiella negative bacteria.
 19   This event fitted very well with
 20  the clinical outcomes. I had to write the
 21  assessment for the prosecutor so I could study
 22  the clinical data, too. That means all these
      Page 92
  1  things were very, very understandable. What
  2  we did is we characterized the strain
  3  following our approach. That means spiking
  4  different platelet bags or platelet bags from
  5  different donors with .03 CFU/mL and saw a
  6  very interesting outcome.
  7   In several cases the klebsiella
  8  pneumonia strain grew as other strains we
  9  observed before already. That means very
 10  fast. But there were platelet bags in which
 11  a very much prolonged adaptation phase or --
 12  I just forgot the other term. The strain
 13  needed up to three days to enter the cutoff
 14  for the determination of this experiment.
 15  That means 10 CFU/mL.
 16   So the diagnostic that I described
 17  to you before, that means we contaminated
 18  platelet bags with .03 CFU/mL, draw a sample
 19  after 31 hours with a negative result. When
 20  we repeated a sample drawing two days after
 21  contamination we could produce a positive
 22  result.
      Page 93
  1   The consequence is that in biology
  2  there is a diagnostic window in platelet
  3  bacteria screening and this window can be
  4  dependent on the given strain and donor
  5  properties can be changed or can be prolonged.
  6  The consequence, the conclusion is that we
  7  should draw the sample as late as possible
  8  imagining that the contaminating bacterium,
  9  Vostal mentioned the one bacterium per bag,
 10  Jaro Vostal mentioned, give the bacteriums a
 11  chance to grow as long as possible, to
 12  multiply as long as possible.
 13   Let's hold the entity of platelet
 14  back together and let's draw the sample as
 15  late as possible in order to increase the
 16  chance to get the bacteria in our method or in
 17  order to overcome the sampling error. This
 18  approach had been proposed by the already
 19  mentioned ISBT working party on Transfusion
 20  Transmitted Infectious Diseases subgroup in
 21  Transfusion Today in March 2006.
 22   What rapid methods, feasible
      Page 94
  1  methods for rapid bacteria detection do we
  2  have? First, I would like to show you flow
  3  cytometry developed by Becton Dickinson
  4  Biosciences Europe. The Verax system had been
  5  mentioned already. Then universal bacteria
  6  PCT NAT where no -- where still no commercial
  7  variant is available.
  8   Then I would like to show you some
  9  data with a newly developed system which is
 10  able to measure without pH inside blood
 11  effects without opening the bag, the rapid
 12  detection of microbes in PCs using flow
 13  cytometry. As you can see here, it's a very
 14  simple and very easy procedure.  
 15   A sample of 50 microliters drawn
 16  from the platelet bag is mixed with 450
 17  microliters of a lysis buffer containing the
 18  fluorescent dye Thiazole orange. Then
 19  incubation for five minutes measuring for 30
 20  seconds. That means result within 10 minutes.
 21  These are some results obtained with different
 22  bacteria. You can read it yourself, staph
      Page 95
  1  aureus, staph epidermidis, bacillus cereus,
  2  klebsiella pneumoniae are the bacteria left
  3  inside the gray events or the platelet debris.
  4   Very important ingredient here
  5  marked by the red events absolute count beads
  6  implemented into the lysis buffer which
  7  guarantee that during measuring there is not
  8  any impairment of the machine when the sample
  9  is drawn and so on and so on and so on.
 10   These are data we performed in my
 11  department in order to characterize how flow
 12  cytometry can be used and how the signals are
 13  produced after artificial contamination with
 14  .03 CFU/mL. You see the time after
 15  contamination and the bacteria is clearly
 16  growing up.
 17   As you can see here in these two
 18  pictures there is a problem regarding the
 19  differentiation between the platelet debris
 20  and the bacteria. In consequence a two-step
 21  method has been developed which prolongs a
 22  little bit the time up to diagnosis from 10 to
      Page 96
  1  20 minutes but is producing much better
  2  discrimination between the platelet debris and
  3  the bacteria signal.
  4   Okay. Next information and then I
  5  would like to finish with the flow cytometry
  6  data. There is a way to increase sensitivity
  7  of rapid bacteria detection by flow cytometry
  8  simply by drawing a sample, take the sample
  9  into tube, incubate the tube at 37 degrees.
 10  This is shown here. That means sample drawn
 11  16 hours after contamination almost no signal.
 12   Then pre-incubation at 37 degrees
 13  for one hour, for two hours, and that is the
 14  sample drawn from the platelet after 18 hours.
 15  You can see you have to compare it with the
 16  pre-incubated sample. It's an impressive
 17  increase in sensitivity of the method. All
 18  together the increase per one hour incubation
 19  at 37 degrees is around one lock step.
 20   Summarizing rapid bacteria
 21  detection using flow cytometry detects
 22  bacteria and fungi because of resin-sized
      Page 97
  1  nuclear-phillic cowering or laboring genome
  2  and ribosomes of microbes. It's feasible
  3  sensitivity is 103 to 104 CFU/mL can be
  4  improved by pre-incubation. Result is
  5  available after 20 minutes.
  6   Rapid bacteria detection using Pan
  7  Genera detection system developed by Verax and
  8  now offered in collaboration with Abbott.
  9  Mark Brecher mentioned it already. We do not
 10  have up to now our own experiences on the
 11  basis of the available data. I would like to
 12  assess it as a feasible approach too. It
 13  detects a broad spectrum of bacteria. It's
 14  not able to detect fungi. Sensitivity between
 15  103 to 105 CFU/mL result after around one
 16  hour.
 17   Rapid detection of microbes using
 18  real time universal bacteria and, of course,
 19  fungi, PCR NAT. The basis is that on the
 20  ribosomal RNA of microbes bacteria as well as
 21  fungi they are highly conserved sequences so
 22  there is a chance to perform in one PCR
      Page 98
  1  identification and analyzing detection of any
  2  bacteria species or any fungal species
  3  imaginable.
  4   These are data from my post-doc
  5  Dr. Melanie Stomer in my department working
  6  with bacterial suspensions alone there is a
  7  sensitivity in RTPCR of 16 CFU/mL. Not bad
  8  but when we are going into platelet
  9  concentrates one of the scientific backgrounds
 10  for this sensitivity is that the mitochondrial
 11  DNA from human beings has cross-reacting
 12  regions with the ribosomal RNA from bacteria.
 13  That means sensitivity in PCs for several
 14  reasons is 102 to 103 CFU/mL.
 15   In my knowledge, there is no
 16  commercial variant available up to now from
 17  universal bacteria PCR. Me, personally, I
 18  know a lot of different groups at least in
 19  Europe which are dealing with and which
 20  approved the approach. From my perspective,
 21  sensitivity is acceptable. There is one
 22  problem considering real time PCR, especially
      Page 99
  1  reverse transcriptase, real time PCR. Of
  2  course, I need a few hours for production of
  3  the result and so PCR is not a real rapid
  4  method when compared with the other methods I
  5  showed you.
  6   A few data regarding an
  7  interesting approach which is available or
  8  have been made available by a company from
  9  Seattle, BCSI, Blood Cell Storage, Inc.
 10  Actually they performed or they developed a
 11  procedure for in-line pH measuring in platelet
 12  bags without any opening of the system by a
 13  scanning procedure of resin-sized - labeled
 14  here to a membrane inside this blue cylinder.
 15   What we did we contaminated the
 16  platelet bags following the approach .03
 17  CFU/mL starting dose. Here you can see the
 18  data regarding bacterial growth. This is the
 19  pH. The pH will drop down after the bacteria
 20  had reached a high number. In this case
 21  klebsiella pneumoniae strain 108 per mL. That
 22  could be abated.  
      Page 100
  1   That had been published by Steve
  2  Wagner in the middle of the '90s evaluating
  3  the pH measuring as a screening method.
  4  Surprisingly after this decrease there was a
  5  re-increase of the pH inside the platelets.
  6  I have not time enough to discuss it up to the
  7  last detail. In my eyes, it is at least a
  8  very interesting result. 
  9   The same holds true, for instance,
 10  for yeast in this case -- this one was from
 11  Canada. This is a summary of the results
 12  obtained with different strains, different
 13  blood bacteria standards in comparison to the
 14  negative control. At least this approach
 15  could be used or would produce better results
 16  than the approach mentioned by Mark Brecher.
 17   That means the pH measuring with
 18  dip sticks or strips or whatever. Of course,
 19  generally pH measuring in PCs is not accepted
 20  to be a solitary approach for platelet
 21  bacteria screening because of its low
 22  sensitivity. On the other hand there are time