UNITED STATES OF AMERICA

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FOOD AND DRUG ADMINISTRATION

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TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES

ADVISORY COMMITTEE (TSEAC)

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BETHESDA, MARYLAND

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WEDNESDAY,

JULY 17, 2003

This transcript has not been edited

or corrected, but appears as received

from the commercial transcribing

service. Accordingly, the Food and

Drug Administration makes no

representation as to its accuracy.

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The Advisory Committee met in the Versailles Room at the Holiday Inn
Select, 8120 Wisconsin Avenue, Bethesda, Maryland 20814, at 8:00 a.m.,
with Suzette A. Priola, Ph.D., Chair, presiding.

[PRESENT]{.underline}:

SUZETTE A. PRIOLA, Ph.D., Chair

JOHN C. BAILAR, III, M.D., Ph.D., Member

ARTHUR W. BRACEY, M.D., Member

LISA A. FERGUSON, D.V.M., Member

PIERLUIGI GAMBETTI, M.D., Member

R. NICK HOGAN, M.D., Ph.D., Member

RICHARD T. JOHNSON, M.D., Member

RIMA F. KHABBAZ, M.D., Member

SIDNEY M. WOLFE, M.D., Member

CHARLES E. EDMISTON, JR., Ph.D., Temporary Voting Member, Topics , 3 & 4

KENRAD E. NELSON, M.D., Temporary Voting Member, Topics 2, 3 & 4

TERRY V. RICE, Temporary Voting Member,

Topics 2, 3 & 4

DAVID F. STRONCEK, M.D., Temporary Voting Member, Topics 2, 3 & 4

SHIRLEY J. WALKER, Consumer Representative

STEPHEN R. PETTEWAY, JR., Ph.D., Non-Voting Industry Representative

SHEILA D. LANGFORD, Staff

[ALSO PRESENT]{.underline}:

DR. DAVID M. ASHER, OBRR, CBER, FDA

DR. STANLEY BROWN, CDRH

DR. YUAN-YUAN CHIU, CDER, FDA

DR. MICHAEL DUNN, V. Pres., Chairman of the Regulatory Committee, GMIA

DR. JAY EPSTEIN, Director, OBRR, CBER, FDA

DR. ROBERT HILLS, Health Canada, Ottawa

DR. GEORGE MASSON, President GMIA

DR. TERRY MORRIS, APHIS

DR. PEDRO PICCARDO, CJD

DR. MORRIE POTTER, CFSAN, FDA

CAPTAIN EDWARD RAU, Environmental Health Officer, NIH

DR. RON ROGERS, Health Canada, Ottawa

DR. ROBERT ROHWER, Director Molecular Neuro-Virology Unit, VA Medical
Center, Baltimore

DR. WILLIAM RUTALA, UNC

REINHARD SCHRIEBER, Chief Manufacturing Officer, Deutsche Gelatine

DR. ROBERT SOMERVILLE, IAH Edinburgh, UK

FABRIKEN STOESS, AG, Gelita Group

DR. DAVID TAYLOR, SEDECON 2000, UK

NELSON BROOKLANG, Ortech International

DANIEL R. DWYER, ESQ., Kleinfeld, Kaplan & Becker, Counsel to GME

CHARLES FILLBURN, Nutranax Laboratories

PAUL HAFFENDEN, TerraCell

MERLYN SAYERS, M.B., B.Ch., Ph.D., Carter BloodCare

WAYNE E. VAZ, Serologicals Corporation

[AGENDA ITEM]{.underline} [PAGE]{.underline}

[OPENING/INTRODUCTION]{.underline}:

William Freas 6

[CONFLICT OF INTEREST STUDY]{.underline}:

Katherine McComas 11

[TOPIC 1 - SAFETY OF BOVINE BONE GELATIN]{.underline}:

[Background & Introduction]{.underline}:

Morrie Potter 13

[Questions to the Committee]{.underline}:

Yuan-Yuan Chiu 18

[Market Trend in the U.S.]{.underline}:

George Masson 21

[Questions/Comments]{.underline}: 27

[Manufacturing Process for Bone Gelatin in U.S.]{.underline}:

Michael Dunn 39

[Questions/Comments]{.underline}: 54

[Manufacturing Process Bone Gelatin in Europe]{.underline}:

Reinhard Schrieber 60

[Questions/Comments]{.underline}: 74

[GME Validation Studies of Bone Gelatin]{.underline}:

Robert Somerville 75

[Questions/Comments]{.underline}: 94

[Gelatin Manufacturing Process]{.underline}:

Robert Rohwer 101

[Questions/Comments]{.underline}: 116

[Risk Analysis of Infectivity]{.underline}:

Ron Rogers 120

[Questions/Comments]{.underline}: 135

[USDA Gelatin Policy]{.underline}:

Terry Morris 136

[AGENDA ITEM]{.underline} [PAGE]{.underline}

[Questions/Comments]{.underline}: 142

[Open Public Hearing]{.underline}:

William Freas 144

Daniel Dwyer 145

[Committee Discussion & Voting]{.underline}: 150

[TOPIC 2 - BSE IN CANADA]{.underline}:

[Potential Exposure of Blood Donors in]{.underline}

[North America to the BSE Agent]{.underline}:

Jay Epstein 192

[Questions/Comments]{.underline}: 193

[Review of BSE in Canada]{.underline}:

Robert Hills 195

[Questions/Comments]{.underline}: 210

[Open Public Hearing]{.underline}:

Wayne Vaz 222

Merlyn Sayers 229

[Questions/Comments]{.underline}: 236

[Committee Discussion]{.underline}: 238

[TOPICS 3 & 4 - TSEs AND DECONTAMINATION OF MEDICAL EQUIPMENT AND
FACILITIES]{.underline}:

[TSEs, Decontamination & FDA Regulated Products]{.underline}:

David Asher 241

[Principles of TSE Inactivation]{.underline}:

Robert Rohwer 251

[Questions/Comments]{.underline}: 269

[Basis for WHO Recommendations]{.underline}:

David Taylor 270

[Questions/Comments]{.underline}: 289

[AGENDA ITEM]{.underline} [PAGE]{.underline}

[Reducing Risk of CJD Transmission Through]{.underline} [Surgical
Procedures: Experience in UK]{.underline}:

Philippa Edwards (Pedro Piccardo) 299

[Questions/Comments]{.underline}: 311

[TSE Agents/Infection Control in USA Hospitals]{.underline}:

William Rutala 315

[Questions/Comments]{.underline}: 330

[Infectivity of Air Emissions & Residues from]{.underline}

[Simulated Incineration of Scrapie Tissues]{.underline}:

Edward Rau 338

[Questions/Comments]{.underline}: 350

[TSE Infectivity: Experience With Models for]{.underline}

[Validating Decontamination of Surfaces]{.underline}:

Stanley Brown 351

David Asher 361

[Questions/Comments]{.underline}: 375

[ADJOURN]{.underline}:

Suzette Priola 379

P-R-O-C-E-E-D-I-N-G-S

8:03 a.m.

SECRETARY FREAS: Dr. Priola, members of the public, invited guests and
public participants, I would like to welcome all of you to this our
14^th^ meeting of the Transmissible Spongiform Encephalopathies Advisory
Committee. I am Bill Freas. I am the executive secretary for this
Committee. At this time, I would like to go around and introduce to you
the members at the head table, starting on the right hand side of the
room.

The first chair will soon be occupied very shortly by Dr. Pierluigi
Gambetti. He is a professor and director Division of Neuropathology
Case, Western Reserve University. Okay. Then the second chair will soon
be occupied by Dr. Richard Johnson, professor of neurology at Johns
Hopkins University. And then going around the table, the people who are
here, Dr. Arthur Bracey, associate chief Department of Pathology, Saint
Lukes Episcopal Hospital. Next is Dr. Lisa Ferguson, a senior staff
veterinarian, U.S. Department of Agriculture.

Next is Dr. Nick Hogan, associate professor of ophthalmology, University
of Texas, Southwestern Medical School. Next is Dr. Rima Khabbaz,
associate director for Epidemiologic Science, National Center for
Infectious Diseases, Atlanta, Georgia. Around the corner of the table is
a gentleman, whom I\'m going to ask to join us at lunch time, if that is
okay, Dr. Nelson. Could I ask you to join us at lunch time instead of in
the morning?

DR. NELSON: Certainly.

SECRETARY FREAS: This is my mistake. I apologize. Dr. Nelson will be a
temporary voting member, and he will join us right at lunch time, and if
you could just sit over in the FDA section up until Topic 1 is over. And
when I read the Conflict of Interest statement, hopefully, that will be
explained. My apologies for not checking before I started. Okay.

Next is our Chair, Dr. Suzette Priola. She is an investigator of
Laboratory of Persistent and Viral Diseases of the Rocky Mountain
Laboratories. Next is our consumer representative, Ms. Shirley Walker,
vice president of the Health and Human Services, Urban League of Greater
Dallas in north central Texas. Next is Dr. Sidney Wolfe, director of
Public Citizen Health Research Group, Washington, D.C.

Next is Dr. John Bailar, professor in America\'s University of Chicago.
Next is our non-voting industry representative, Dr. Stephen Petteway,
director of Pathogen Safety and Research, Bayer Corporation. Three
Committee members in addition to the two that are joining us shortly
could not be with us at all for this meeting. They are Mr. Val Bias,
consumer representative, Lynn Creekmore, staff veterinarian and Dr.
Stephen DeArmond from the University of California.

I would now like to read into the public record the Conflict of Interest
statement for this meeting. \"The following announcement is made part of
the public record to preclude even the appearance of a Conflict of
Interest at this meeting. Pursuant to the authority granted under the
Committee Charter, the Director Center for Biologics Evaluation and
Research has appointed Mr. Terry Rice and Drs. Kenrad Nelson, who I just
asked to leave the table, and David Stroncek as temporary voting members
for Topics 2, 3 and 4 of this meeting.

In addition, the associate commissioner of External Relations of FDA has
appointed Dr. Charles Edmiston as a temporary voting member for Topics
2, 3 and 4 of this meeting. Based on the agenda, it has been determined
that the Committee will not be providing advice on specific firms or
products at this meeting. The topics deemed discussed by the Committee
in open session are considered general matters issues.

To determine if Conflicts of Interest exist, the Agency reviewed the
agenda and all relevant reported financial interests from meeting
participants. The Food and Drug Administration prepared general matters
waivers for special Government employees, who required a waiver under 18
U.S. Code 208. Because general matters topics impact on so many
entities, it is not prudent to recite all potential Conflicts of
Interest as they apply to each member.

FDA acknowledges that there may be potential Conflicts of Interest, but
because of the general nature of the discussion before the Committee,
these potential conflicts are mitigated. We would like to note for the
record that Dr. Stephen Petteway is serving as a non-voting industry
representative member for this Committee. He is employed by Bayer and
thus has interests in his employer and other similar firms.

Listed on the agenda are speakers making industry presentations and/or
updates. These speakers have financial interests associated with their
employer and with other regulated firms. These speakers were not
screened for these Conflicts of Interests. With regard to FDA\'s invited
guest speakers, that\'s all other speakers, except those from industry,
the Agency has determined that the services of these speakers are
essential.

The following interests are being made public to allow the meeting
participants to objectively evaluate their presentations and comments
that they may make. Dr. Robert Rohwer has disclosed he has financial
interest with various firms that could be affected by the Committee
discussions. Dr. William Rutala receives consultant fees from several
firms that could be affected by the Committee discussions. Dr. Robert
Somerville has research supported by the Gelatin Manufacturers of
Europe. His expenses to this meeting were also paid by the Gelatin
Manufacturers of Europe. Dr. Charles Weissmann holds patents related to
Prion Disease work.

Members and consultants are aware of the need to exclude themselves from
the discussions involving specific products or firms which they have not
been screened for the Conflict of Interest. Their exclusion will be
noted in the public record. With respect to all other meeting
participants, we ask, in the interest of fairness, that they address any
current or previous financial involvement with any firm whose product
they may wish to comment upon. Waivers may be available by written
request to the Freedom of Information Office.\"

That\'s the end of the Conflict of Interest statement. I do ask that
throughout this meeting before it starts if you would check your cell
phone or your pager and, please, put it in the silent mode, so it won\'t
disrupt those people sitting next to you.

Next, the FDA is continually trying to improve its Advisory Committee
Program and to reduce any perceived Conflicts of Interest. It has asked
Dr. Katherine McComas from the University of Maryland to conduct a
survey of this program, and I would like to give her the opportunity to
tell us how we can help her with this survey and how the survey is being
conducted. Dr. McComas, either place. Keep talking and they\'ll turn the
mike volume up.

DR. MCCOMAS: Okay. Good morning and thank you. I\'m Katherine McComas
and I\'m a faculty member at the University of Maryland, and I\'m here
today to conduct a study of what people know and understand about the
Conflict of Interest procedures that the FDA uses to monitor real or
potential Conflicts of Interest of its Advisory Committee members. This
is a study that is being conducted across multiple meetings. This is the
11^th^ meeting I have attended across the centers at FDA, including
CBER.

For those of you in the audience, I\'ve distributed a questionnaire on
your chair and I have also distributed a different questionnaire to the
Advisory Committee members. If you have an opportunity today to complete
this questionnaire or tomorrow, there is a box on the registration desk
where you can drop it. Otherwise, there is a business reply envelope
that you can just drop it in the mail as soon as you can. Your
participation is voluntary, but it is important. The more responses we
get, the better we are to provide feedback to the FDA about what people
know and understand about the Conflict of Interest procedures, and what
may be done to improve satisfaction, if necessary, with the Advisory
Committee process.

Again, I appreciate your participation and if you have any questions, my
contact information is included in the letter, in the questionnaire and
I would be happy to provide any answers. Also, when the study is done,
the responses will be available in summary form to everyone who is
interested. So thank you very much for your time and have a great day.

SECRETARY FREAS: So if you got here early and did not receive a
questionnaire, the questionnaires are on the table outside and, please,
everybody on the FDA staff will be more than glad to help you if you
have any questions with this questionnaire.

Dr. Priola, I turn the microphone over to you to start the meeting.
Thank you.

DR. PRIOLA: So since we have a very full agenda today, we will just get
started with the first speaker, who is Dr. Potter, who will give us
background on Topic 1.

DR. POTTER: Good morning. FDA has been considering the safety of gelatin
with regard to BSE for a number of years, and has come to this Committee
on a number of occasions to get its recommendations on FDA\'s guidance
to gelatin manufacturers and users. The safety of gelatin is determined
as you\'ve told us before by the safety, the source materials in the
degree to which the gelatin manufacturing process destroys prions that
enter the system.

Questions to the Committee have dealt with these two factors and how
well knowledge about TSE\'s was reflected in FDA guidance for assuring
the safety of gelatin for food and cosmetic use. Before 1996, FDA did
not include gelatin within its recommended restrictions concerning
bovine ingredients in FDA regulated products. In 1996, FDA began to
review its position on gelatin in light of new information that
associated BSE exposure with Variant CJD in humans and new data from a
study on the effect on infectivity of gelatin processing that suggested
only partial effectiveness.

In 1997, this Committee met to consider the safety of gelatin and to
provide an assessment on the overall risk to humans associated with
imported gelatin. This Committee made the following observations: First,
that the scientific information available no longer justified excepting
gelatin from restrictions recommended by FDA for other bovine derived
materials from BSE countries. Second, that bovine gelatin injected or
implanted forms posed a higher risk of transmitting BSE to humans than
gelatin that is ingested. Third, that brains and spinal cords from
cattle from BSE countries should be excluded from raw materials used to
produce gelatin for human consumption. Fourth, alkaline or acid
processing in gelatin manufacturer may only reduce rather than eliminate
BSE infectivity, and the Committee called for better validation studies,
particularly to investigate the other steps of gelatin manufacture. And
finally, that porcine gelatins appear to pose no known risk of
transmitting TSEs to humans.

After the 1997 TSEAC meeting, FDA issued its gelatin guidance document
which remains the current FDA position in policy on the production of
gelatin. In this guidance, FDA proposed the following recommendations
concerning the acceptability of gelatin in FDA regulated products
intended for human use. First, that importers, manufacturers and
suppliers should determine the tissue, species and country source of all
materials used in processing gelatin for human use.

Second, that bone and hides from cattle from any source country that
show signs of neurologic disease should not be used as raw materials.
Third, gelatin production from bones and hides obtained from cattle that
reside in BSE countries or countries that do not meet the latest BSE
related OIE standards should not be used in injectable, ophthalmic or
implanted FDA regulated products or in their manufacture, but may be
used in FDA regulated products for oral consumption and cosmetic use by
humans if the cattle come from BSE-free herds and if the slaughter house
removes heads, spines and spinal cords directly after slaughter.

Fourth, gelatin produced from bovine hides from any source country may
be used in FDA regulated products for oral consumption and cosmetic use
by humans if processors insure that the hides have not been contaminated
with brain, spinal cord or ocular tissues of cattle residing in or
originating from BSE countries. Fifth, gelatin produced from bovine
hides and bones may be used in FDA regulated products for human use if
the gelatin is produced from raw materials from countries like the
United States that observe OIE standards and have not diagnosed BSE in
their national cattle herd, that is RBSE-free. And finally, gelatin
produced from porcine skins from any source country may be used in FDA
regulated products for human use.

In 1998, this Committee met again to discuss gelatin among other issues.
FDA\'s guidance, based on the 1997 TSEAC recommendations, was presented
to the Committee to consider several new pieces of relevant information.
For example, the infectivity of dorsal root ganglia and low level
infectivity in bone marrow and the growing number of BSE cases being
discovered in Europe. The Committee considered this new information and
decided gelatin could be safely sourced from bones and hides of cattle
in BSE countries as long as the recommendations in the guidance were
met. That is that the cattle came from BSE-free herds and the high-risk
materials were removed after slaughter.

And this is at present the status of the safe source factor for gelatin.
Continuing on with the other key factor, that of validated effectiveness
in the manufacturing process, in June 2001, the Committee was given an
update from the Gelatin Manufacturers of Europe on the interim
validation study results on the inactivation of BSE through the gelatin
manufacturing process. This was an information sharing meeting only and
no questions were posed to the Committee.

The Committee reviewed the study design and the preliminary data and
requested a presentation of the final results as soon as they were
available. The Committee is now about to get its wish as GME will
present their completed studies, and we will hear other marketing and
manufacturing information on gelatin in North America and Europe. After
you have heard this new information, we would like you to comment on the
studies and to consider the current gelatin guidance in light of these
completed studies and other relevant information.

And I think, according to my schedule, Yuan-Yuan will now charge up the
Committee. Thank you.

CHAIR PRIOLA: Thank you, Dr. Potter. Dr. Chiu will now present the
questions for Committee.

DR. CHIU: Good morning. First, I would like to thank Dr. Priola and the
Committee members to take the time to come out and also we have sent you
a huge package, gelatin studies protocols and procedures and the
results. We appreciate how much time you need to really review those
studies. In the early days, in the 1998 year, when the Agency and the
Committee together made a decision for the Agency\'s recommendation on
gelatin was based on previous study which the Committee thought was
somewhat flawed.

So generally, this reason follows the advice of the Committee to then
redesign the studies and then today, you know, we have the new study
results. You did not review the interim results, but today we have the
final results off of five studies. We\'re hoping, you know, with the
presentation today and the background information you have you will be
able to help the Agency to answer two questions.

Next slide, the first question is \"Do those results of these new
studies demonstrate a reduction in infectivity that is sufficient to
protect human health?\" And we are only limited to hear the question to
bovine bone gelatin is consumed by humans through oral or topical
administration. The question is not for gelatin of other
administrations, such as the injection, you know, implantable. We would
like the Committee to focus on oral and topical administration.

Next slide, now, the first question, you know, the answer could be yes
or no or in between regardless, you know, the answer we also would like
you to answer the second question. There are two parts. The first part
is \"Do the scientific data and the information available support the
current FDA recommendations on bovine bone gelatin for oral and topical
administration?\"

The current recommendations, next slide, is on this slide. The general
policy of FDA is for FDA regulated products, the bovine derived material
should come from cattle not bone residing as slaughtered in BSE
countries, but the Agency also provides some exemptions. The exemption
could be a total exemption unconditional, such as milk, dairy products
and the milk derived product. But some of the substances, you know, the
Agency provide conditional exemptions, and the gelatin for oral and
topical use are giving conditional exemptions.

So if the cattle actually is coming from BSE countries, then that
condition is the cattle must be from a BSE-free herd and also at the
slaughter house the head, the spine and spinal cord should be removed.
And this is from BSE countries. Now, some countries may not have BSE
cases, but there is consider of high-risk of BSE. Then the
recommendation is the heads, the spine and the spinal cord should be
removed as the first step in the slaughter house. So the first question
is whether this current recommendation still is valid, based on the
scientific information we have today.

Next slide, if the answer is yes, then that\'s the end of it. If the
answer is no, then we would like to know what changes the Committee
would like to recommend to our current policy. The changes can be in all
different directions. You may consider we can actually grant a total
exemption to the gelatin for oral and topical use or you may consider to
modify the current recommendation the FDA has, either by strengthening
or by relaxing the conditions. So we are anxious and grateful you will
give Agency your deliberation. Thank you.

CHAIR PRIOLA: Okay. Thank you, Dr. Chiu. Our next speaker is Mr. Masson,
who will discuss market trends in the U.S.

DR. MASSON: Yes, good morning everybody. Madam Chairman, I would just
like to thank the Committee and the FDA, in particular, about the
opportunity to address the Committee. As we have heard from Drs. Potter
and Chiu, it has been a long and winding road, the saga of gelatin, and
we hope today that we can reach a satisfactory conclusion and see
gelatin taken off the file, so to speak, having reassured you of its
safety.

My first slide, please. Can I have the first slide, please? Okay. Thank
you. Well, just an introduction of I\'m currently the president of our
industry association, the GMIA, and also president and CEO of one of its
members at Russelot. The next slide? A bit of history as to GMIA and
credentials, so to speak. Our association was formed in 1956. We have
six members, all NAFTA based, four from the U.S., one in Mexico and one
in Canada. And we\'ve listed here the typical working committees by
which we run the institute. There is no particular order of precedence,
but the technical and regulation committees as you can imagine, indeed,
are the primary focus of most of our work, I guess.

Next, in terms of what we represent, we, as you see, represent roughly
22 percent of the global gelatin production, and almost 100 percent of
all the gelatin made in North America. And, indeed, three of our members
are also affiliates of the Gelatin Manufacturers of Europe. And I should
have added actually that one of the other members is an affiliate of the
Japanese Gelatin Manufacturing Group.

Next, please. This lists our objectives. As you see, we try to monitor
and inform our members of any and all regulations which can impact
gelatin. We are the liaison with FDA, USDA and other regulatory
authorities, and we gather and distribute technical information to our
members, endeavor to promote a broader knowledge of gelatin and
encourage its wider consumption. And we provided the forum as you\'ve
seen from our committee information on all of the major aspects
concerning technical, environmental and safety issues.

And as time has gone by and as other industry associations have been
formed around the world in Japan, South America and so on, a major
function which has emerged has been to liaise with them to ensure that
technical information and regulation information, etcetera, is shared
with the other associations around the world. And I just participated,
for instance, in the Japanese meeting or the Asia Pacific meeting, which
was held in Japan, just last month, as an example of the increasing
international corporation among the industry associations.

Next, please. This slide lists the primary uses of the concentrated,
obviously for today\'s purposes, bovine bone gelatin and, as you see
here, this is a list of the major uses for bovine bone gelatin in the
United States. It is listed in the standing order of use with
photographic still being the largest consumer going on down to food.
There\'s less and less bovine bone gelatin used in food products, by
food we mean confectionery and marshmallows or whatever else.

It is being, I guess, more replaced there by pigskin porcine gelatins.
But anyway, those are the primary uses and, as you see, no matter what
the end use, all the gelatins are produced through the same
manufacturing processes and my colleagues will be describing those in
some detail in a few minutes.

Next, please. To give you an idea of the scale of the gelatin business
globally, and in particular the bovine part of that production, we have
listed here the various theaters, so to speak. Europe is still the
biggest gelatin producer. Significantly so with 117,000 tons out of a
total of some 270,000 tons around the world. Above that, I would have to
say, over 25 percent or so is actually bone gelatin. The U.S. in total
we make something like 60,000 tons. And again, these are all gelatins,
whether bovine or porcine or bovine hide or bovine bone, porcine skin.

And as you see, of the 60,000, about 17,000 tons is actually bone
gelatin. Other covers is Asia Pacific, the Asia Pacific regions and
South America, and you see about a third of their gelatin is of bovine
bone origin. So totally, bovine bone represents almost 80,000 out of the
total of 270,000 tons. And to give you an idea again of this
international value industry total globally is not that big. It\'s 1.5
billion dollars equivalent.

Next, please. What we tried to do here is to put to ourselves a few
questions, the elimination of which, I think, will be helpful to the
committee in looking at bovine bone gelatin, in particular, in the U.S.
The first question, as you can see is, \"Can the U.S. gelatin industry
supply total U.S. capsule industry\'s needs?\" The answer is no. And
this illustrates how that is the case.

As you saw earlier, the U.S. bovine bone gelatin production totals some
17,000 tons, but of that 11,500 are needed for photographic and other
non-capsule uses. So you see that that remains, there remains only about
5,500 tons which can be used by the capsule industry, but the total
needs, in fact, are 10,000 tons, and this means that the shortfall
roughly 4,500 tons of bovine bone gelatin has to be imported and they
come primarily from Europe, also from Japan and India, and immediately
derived nevertheless from U.S. bones or from bones from other countries.

Next, just continuing that theme, of those 4,500 tons of which we need
to import, \"Can they be derived solely from U.S. bones, even if it\'s
not actually manufactured in the U.S.?\" Again, the answer is no. As you
see here, the total amount of U.S. bones which are made available to the
gelatin business is roughly 130,000 tons and because of its use in
photographic production, whether in Europe or in the U.S., and also for
manufacturing bovine bone gelatin by other companies outside of the
U.S., the amount remaining available for pharmaceutical gelatin
production here is only 28,000 tons.

The next line which is in bold print illustrates that we need roughly 6
tons of bones to make a ton of gelatin. So that the 10,000 tons of
gelatin, which the capsule industry needs, is actually equivalent to
60,000 tons of bovine bones and, consequently, you see the shortfall
here is roughly 32,000 tons, so to speak, to be able to make all of the
capsule industry requirements strictly from U.S. bones. So in other
words, the deficit has to be sourced from bone suppliers outside of the
United States.

Next, so then I apologize, this is a little bit of a busy slide, but the
bottom line is that there are, indeed, other sources outside of the
United States, but even though the quantity is maybe available for
various reasons in terms of surveillance, the inspection procedures and
so on, it\'s not so obvious that the quantities, the tonnages, which are
listed in the second line or the second section are, indeed, available
and because of the various restrictions and so on, you see, in fact,
that the bone and that those numbers diminished to rather smaller
numbers.

And this really drives to the heart of the matter. This is the crux
really of what we want to get at today and my colleagues will be
addressing this individually and then in the public comment session
later, the question of how we can determine the BSE status adherents and
also the question as Dr. Chiu referred to of just when they have to be
removed in the gelatin bone process. Again, we\'ll be traveling to that
in some much more detail in later presentations.

Next, please. I guess that concludes my presentation, unless there are
any questions.

CHAIR PRIOLA: Yes, are there any questions for Mr. Masson?

DR. MASSON: Thank you.

CHAIR PRIOLA: Oh, Dr. Hogan?

BOARD MEMBER HOGAN: I have one question.

CHAIR PRIOLA: Just a second, Mr. Masson, there\'s a question.

BOARD MEMBER HOGAN: Sorry. I had one question, perhaps it is contained
in this information you provided us, which is quite huge. In terms of
the amount of gelatin that is derived from Europe, could you tell us
something about the country breakdown, that is it\'s most from the UK,
France, Switzerland, etcetera?

DR. MASSON: Yes, I think you\'ll find in the information packet there is
a detailed breakdown of the various imports. The consumption in the
U.S., actually the total marketing, is closer to 80,000 tons. And as you
saw, we make 60,000. There is a net import/export situation. The U.S.
does export gelatin, but basically to get to the 80,000 that we need, we
need effectively a net import of 20,000 tons.

Those 20,000 tons come from quite a variety of countries and, indeed,
they are listed in the information packet. We didn\'t go into the detail
of it here, because it\'s somewhat difficult to differentiate, certainly
differentiate country by country. It\'s a little bit more difficult to
differentiate within certain countries whether it is bovine gelatin or
porcine gelatin, which is actually being imported. But basically the
primary countries who do export into the States would be France,
Germany, not so much any more from UK, for obvious reasons, Brazil,
Argentina, Japan, India. Those would represent the large majority of the
total import picture.

And again, the variety of gelatins some of that is bovine bone, for
sure, but also a lot of bovine hide gelatin comes, for instance, from
South America, and bovine bone also from India. It\'s quite a variety of
types from those principle countries.

CHAIR PRIOLA: Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yes, I didn\'t hear you well and I apologize. When
you said in foods increasingly, there\'s less bovine gelatin and an
increased used of porcine gelatin. Was that porcine skin?

DR. MASSON: Yes, one can make and, indeed, one does use porcine bones,
but the large majority of porcine gelatin made around the world is from
porcine skins. And again just to elaborate on that point, the food
industry, the present manufacturing process of bone gelatin, which
we\'ll hear much more about in a few minutes, is a very long process.
It\'s a very costly process. Whereas porcine gelatin and hide gelatin,
certain portions, is a much sorter process. And economically, therefore,
it\'s much more viable to utilize porcine gelatin, in particular, in the
food industry compared to bone, you know.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: The numbers went by pretty rapidly, but it looked
to me like the proportionate shortfall from U.S. production is about the
same as the proportionate shortfall when you add production from U.S.
bones processed elsewhere. Is that correct? I\'m looking at the second
and third from the last slides.

DR. MASSON: And again, could you just repeat that?

BOARD MEMBER BAILAR: Well, in the answer here to question 1, the third
from last slide.

DR. MASSON: Yes.

BOARD MEMBER BAILAR: There was a shortfall of 4,500 tons and a need of
10,000. And in the next one, it was a shortfall of 32,000 tons and a
total need of, was it, 60,000, maybe I have misread this. Yes, 60,000.
It\'s about the same proportions, but I understand why these includes
other production and the other does not.

DR. MASSON: The shortfall with the U.S., as you see, makes 17,000 tons,
that is equivalent to over 100,000 tons of bones, and as we said,
basically the cattle industry needs 10,000 tons of gelatin and only half
of that effectively is made here in the States. The other half, because
of lack of availability of bones and lack of capacity in the States for
bovine bone production, has to come from outside of the States, and
that\'s, as you mention, roughly the same proportion. It\'s almost
50/50. Does that help?

BOARD MEMBER BAILAR: If I understand correctly then, adding the U.S.
bone processed elsewhere doesn\'t help much at present?

DR. MASSON: Excuse me, adding?

BOARD MEMBER BAILAR: Adding gelatin from U.S. bones processed elsewhere
does not, at present, help very much.

DR. MASSON: No, because again the total demand for U.S. bones, because
of the other applications, particularly for photographic and other
European and other countries utilization of U.S. bones, they don\'t
always end up as pharmaceutical gelatin. The end up more often as
photographic gelatin, so there\'s just not the amount of U.S. bones
going overseas which can come back to the U.S. as pharmaceutical gelatin
for capsule production.

BOARD MEMBER BAILAR: At what point is the distinction made regarding the
ultimate use of the gelatin?

DR. MASSON: Regarding what, sir?

BOARD MEMBER BAILAR: Regarding the ultimate use of the gelatin. Is it
all processed? I thought it was all processed in the same way.

DR. MASSON: Well, my colleagues will describe that in a great deal of
detail, and it is more or less, yes.

CHAIR PRIOLA: And Dr. Wolfe?

BOARD MEMBER WOLFE: This is sort of a follow-up on John\'s question. You
mentioned two factors that are rate limited, so to speak, in terms of
the use of U.S. bones. One was the capacity, presumably, to convert U.S.
bones into gelatin, and secondly, was the unavailability or the shortage
of U.S. bones. I can\'t believe that the second one is really a problem.
It is likely that only a small fraction of U.S. bones are currently
being exported to other countries for reprocessing. I mean, is that
correct or not? I mean, it must be a limitation on production, not a
limitation on U.S. bones, and that gets to the issue of why there
couldn\'t be an increase. If the capacity is the problem, why there
couldn\'t be an increased export of U.S. bones to European countries to
use them, preferentially, in favor of bones from BSE countries.

DR. MASSON: Yeah, your point is well- taken. The problem, however, is
that the largest consumer of bovine bone, as you see, is the
photographic industry. Out of the 130,000 tons, which is produced in the
states, over 100,000 or approximately 100,000 goes to the photographic
industry. And by definition, therefore, the remainder simply isn\'t
satisfactory, and we can\'t drive \-- the gelatin industry is at the
bottom of the totem pole, so to speak, in terms of creating greater
availability of bones. The different industries sell so much bone
that\'s made available to us basically, and there is only so much.

BOARD MEMBER WOLFE: Are you saying that the bone either goes to
photographic industry or elsewhere and that there isn\'t, at the present
time, bone from U.S. beef that is not being converted into gelatin? I
mean, what percentage of the, theoretically, available bone from U.S.
beef is, in fact, being converted to some kind of gelatin? Because my
question is sort of getting to the issue of whether or not it is
possible to divert or not to divert, but just to increase the use of
bone from U.S. beef, even though you want to \-- you said there\'s a tug
between photographic gelatin and other gelatins if the total amount of
bone was available, you could satisfy both of them. So just,
specifically, how much of U.S. bone is, in fact, getting converted into
some kind of gelatin? Half of it, two thirds of it, all of it? What?

DR. MASSON: Well, the bone that is available is being converted. Again,
there are only a few bone producers of the major beef players, but only
a few of them actually make gelatin bone at some of their facilities.
And again, it\'s supply and demand. We can\'t. Basically, there\'s not
enough demand from our side that would force them or encourage them,
let\'s say, to produce still more bone. It\'s that simple.

BOARD MEMBER WOLFE: Okay.

CHAIR PRIOLA: Dr. Johnson?

BOARD MEMBER JOHNSON: Yeah, I may understand the way this is processed.
But it seems to me that the ready solution would be that U.S. bone would
be used for all consumables, whether they be dietary supplements and
then you could use the foreign bone for photographic materials. It\'s
about even.

DR. MASSON: On paper that\'s true, but that\'s \--

BOARD MEMBER JOHNSON: That\'s what I\'m looking at.

DR. MASSON: Yes. But it\'s rather simplistic, because again we can\'t
make that determination. It\'s those industries who make that
determination. The photographic industry has determined that they will
use bovine bone, and that\'s their prerogative that we can\'t influence
it.

BOARD MEMBER JOHNSON: So a solution would be if we deregulated
photographic bovine bone, and that would be a possibility. Does FDA
regulate photographic gelatin?

DR. MASSON: No.

BOARD MEMBER BAILAR: No, they can use whatever they want.

DR. MASSON: Yes.

BOARD MEMBER JOHNSON: So you could split it up.

DR. MASSON: I\'m sorry?

BOARD MEMBER JOHNSON: If there\'s no regulation on photographic gelatin,
you\'re subtracting it out to produce all this shortfall, why not make
the photographic gelatin from British bones?

DR. MASSON: I\'m sure some is, but again, we as an industry can\'t make
that determination. It\'s the photographic people who make that
determination.

CHAIR PRIOLA: Dr. Bracey?

BOARD MEMBER BRACEY: Yes, in the information that you present, the
majority of the gelatin is used for photographic purposes. It seems to
me that there has been a major move away from film based photography
towards digital. Have you seen a reduction in the demand and, in
essence, your picture is a static picture, but what does it look like
really as far as the demand for photographic gelatin in the future?

DR. MASSON: That\'s a very good question. As you rightly observed,
digital photography is here in a big way and will continue to grow. But
there is some complimentality between silver halide, the traditional
silver halide process, which does utilize photographic gelatin and the
digital business. So that the two things, digital is growing certainly
at a much more rapid rate, but photographic traditional silver halide
photography is still very much en vogue and, indeed, you know, the last
photographic companies, Kodak, Fuji and so on still continue to invest
quite significantly in the traditional side of the business as well. So
the two things, I\'m not sure of \--

BOARD MEMBER BRACEY: Well, I guess, what I\'m wondering is over the
years the data in terms of total demand has been static or has it been
actually declining?

DR. MASSON: I would say it is fairly static. There has been a diminution
for sure in some sectors of the traditional silver halide, photographic
side of the graphic arts, for instance, probably uses any photographic
gels any more. That has gone totally, more or less totally, to the
digital side. But the traditional film that you or I shoot, the amateur
film, medical x-ray and other types of cinema, film photography for
movies, those are still the traditional situation, and that demand is
still very much there.

CHAIR PRIOLA: Yes, is there a question from this side or answer?

MR. SCHRIEBER: Thank you, Madam Chairman. I would like to make \--
Reinhard Schrieber.

CHAIR PRIOLA: Could you identify yourself?

MR. SCHRIEBER: From GME, and I would like to make a remark about
potential replacement for the photographic industry of domestic bones
and imported bones. The following situation is the biggest manufacturer
of photographic gelatin is Eastman-Kodak sitting here in the United
States. They are forced to use domestic bones, because as a ban on
import of bones from out of the U.S. into U.S., because the risk of
bringing in bones from maybe BSE risk countries is tremendously high to
bring in just in case by the bones BSE into the United States.

So gelatin is safe to be imported, but importing bones from other
countries, I think, is of high-risk for this community here, so
therefore it would really replace and most probably negligible risk with
gelatin by a big risk by importing bones, degreased bones from other
countries, who therefore is a replacement in this way, I think, is not a
good idea for the U.S. On the other hand, I think it is really
impossible to force Eastman-Kodak just out of using bones from the U.S.
I don\'t know how their reply would be in this case.

And maybe one more question, answer to your question about from which
European countries is sourced in Europe has no bone at all coming for
the last 20 years from UK, so the European gelatin industry did not
source bone from UK. We do not source bone from Ireland. We do not
source bone from Switzerland, Portugal, the so-called higher risk
countries in Europe. All the bones, proven bones used by the European
industry are coming from either Germany, France, Belgium, Netherlands or
Austria. These are the source countries. Thank you.

SECRETARY FREAS: Because our meetings are being transcribed, we\'re
asking everybody who uses a microphone other than at the table to
identify themselves. That was Mr. Schrieber, the chief manufacturing
officer of the Gelatin Group.

CHAIR PRIOLA: Okay. I think we\'ll move on to our next speaker. Thank
you, Mr. Masson.

DR. MASSON: Thank you.

CHAIR PRIOLA: Our next speaker is Dr. Dunn, who is going to explain some
of the manufacturing processes for gelatin in the U.S. and that might
address some of the questions that have arisen.

DR. DUNN: I also would like to thank the FDA and the Committee for the
opportunity to come in and speak with you today about the practices of
the U.S. gelatin manufacturers. My name is again Michael Dunn. I\'m
currently vice president of Gelita North America, and I also serve as
the chairman of the Regulatory Committee for GMIA. As you can tell on
this slide, there are two current manufacturers of bone gelatin here in
the United States, Eastman Gelatin, who provides to Kodak, they are
primarily producing photographic gelatin, and GELITA USA, who is
primarily a pharmaceutical producer.

When we put those together, though, the majority of this gelatin goes to
the photographic applications, although there is a substantial quantity
that does go to the pharmaceutical sector as well. The limed share of
the gelatin that we produce is limed bone gelatin. We do, however,
produce a small amount of what we call Type A or acid bone gelatin, but
this is a very small quantity.

I also wanted to note that the practices that I\'m going to be talking
about today, as well as the processes, apply to both GELITA USA as well
as Eastman Gelatin. Could I have the next slide? So just to set the
overall objectives, they basically are two-fold today. I want to
adequately describe for you today what our current sourcing practices
are, as well as the processing conditions that we use to manufacture
bone gelatin in the United States.

I also want to clearly confirm that the bone gelatin processing
conditions that we employ here are virtually the same that are currently
used in Europe. And more importantly, they meet or exceed the minimum
processing requirements that were spelled out in the GME TSE
Inactivation Study Protocol. This I want to make clear, because we want
to make sure that any of the results, we want to make sure that they are
applicable to what we are producing here in the United States, as well
as what is being produced in Europe.

Could I have the next slide? So when we get to sourcing, in the U.S.
degreased gelatin bone is sourced exclusively from USDA inspected beef
processing facilities in the United States, and this raw material is
derived solely from healthy cattle that have been deemed fit for human
consumption based upon both anti and postmortem inspections.

Could I have the next slide, please? When it comes to SRMs, the U.S.
gelatin bone suppliers have been removing SRMs with the exception of
vertebrae since as early as 1998. And right now, limited quantities of
vertebrae-free gelatin bone have been available from as early as fall of
2002. Currently, there are no FDA or USDA requirements for the removal
of SRMs in the United States. We primarily do the two above bullet
points primarily because of EU regulations and we supply a large number
of customers that have business in Europe that must comply with those
kinds of regulations.

Could I have the next slide, please? So let\'s go on to the process.
What I have outlined here is an overview of what happens in a daily
gelatin production. The major input, of course, to this is the degreased
gel bone. We\'re on the order of about 100,000 pounds of gel bone per a
production day. And we have an equivalent amount of hydrochloric acid,
so another 100,000 pounds of hydrochloric acid would go into this next.
We use at least a half a million gallons of water in the production and,
of course, there is a lot of labor and energy that goes into this as
well.

What I\'ll be talking about primarily today is what goes on in this blue
box here, in terms of the DTL processing conditions. The output we\'re
looking for, of course, is gelatin. On a base of 100,000, you get out
about 25,000 pounds of gelatin, and then about 50,000 pounds of
dicalcium phosphate, which is the primary byproduct of this process.

Could I have the next slide? So overall, what we\'re trying to achieve
here, we\'re starting with the protein we call collagen, which is an
extremely fibrous insoluble protein and we\'re going to transform that
into a protein that is fragmented and soluble, but has a variety of very
interesting functionalities, which makes gelatin such an interesting
business. So there are three major things we are trying to achieve here.

Initially, we need to hydrolyze the collagen. We do this by breaking,
there is intra and inter molecular cross links between the adjacent
chains. We start to break up peptide bonds, so that we\'re able to water
extract this material from the ossein that we\'re producing. Subsequent
to that, we spend a lot of time purifying and concentrating the gelatin.
When we do that initial extraction, it\'s a very dilute solution about 5
percent, so we have to take a lot of water back out of that and then we
purify the material from both a chemical, physical and micrological
point of view.

If I could take the next slide, please? So the incoming gel bone comes
to us. It\'s delivered by a truck or rail car and these are just simply
typical characteristics of that material, and we would use the same
material to make either the Type B or the Type A gelatin. So the fat
content ranges from 1 to 2.5 percent. The size of these chips is an
1/8^th^ inch to 5/8^th^ inch. The mineral protein ratio is about 2 to 1.
And the moisture content is about 6 to 9 percent. And in contrast, it\'s
worth mentioning in the EU all of the producers there have their own
degreasing facilities, which is different than the way things are done
here in the United States. The big meat producers have their own gel
bone processing facilities, and they supply us with this finished bone
chip.

Next slide, please. The first pretreatment step is what we call
acidulation. But what is happening here is the demineralization of the
bone. This is where all that hydrochloric acid comes into place. What
we\'re trying to achieve here is the production of what we call ossein,
which is this demineralized bone material. There\'s a number of
washings, hydrochloric acid washings during this process. We also remove
a lot of non-collagen impurities that come in with the raw bone.

The concentrations that we\'re looking at here, maximum, 4 to 6 percent.
The way this works is it\'s a counter-current distribution process. We
start out with a dilute hydrochloric acid concentration, that\'s what
the initial bone is exposed to, and it\'s gradually raised up over this
4 to 5 days. It\'s a very exothermic reaction, and this is why it takes
to long to carry this out to dissolve out all of this material. The
typical ambient range as far as temperature after this process is done,
the residual acid, is washed out for about a 24 hour period before we go
on to the next step, which is on the alkaloid side of things.

Could I have the next slide? So if we choose to lime, at this point,
this is the breaking point we choose to make either lime bone or acid
bone, at this point. In the case of liming, this is a lime pit that
you\'re seeing up there in the picture. Again, we being, this is where
we continue to hydrolyze the collagen molecules and there\'s a lot of
washing that goes on here with the refreshing of the lime solution, so
we\'re moving impurities.

There is also something important that happens here chemically that is
different than porcine gelatin. You hydrolyze away the asparagine and
glutamine. You deanimate those and form their respective acids which
drops the iso-electric point of that molecule from about 9 down to about
5. So electrically, the porcine and the bovine gelatins are quite
different. We use a saturated lime slurry to do this. The pH is
approximately 12.5. The liming time is 25 to 70 days that we\'re tying
up this material in production for a long period of time before we can
make gelatin out of it. Again, and the temperatures, these lime pits are
agitated on a daily basis. We\'re there to make sure we\'re getting
proper exposure to the alkaline material to the bone chips that are in
the pit. And these lime slurries are completely refreshed on a weekly
basis.

Next slide, please. After that, there is a washing and acidification
step. We want to neutralize the excess lime, again remove, wash out
additional non-collagen impurities, and we want to adjust the pH of the
ossein slurry, so we can prepare it for extraction. So this wash out
period under alkaline conditions is 24 to 48 hours under vigorous
agitation, temperatures from 45 to 70 degrees. The neutralizing or
souring of acids in this case are either hydrochloric or sulfuric acid,
and our target pH for this part of the process range between 5 to 7.

Could I have the next slide? In lieu of liming or alkaline which is what
we do most of the time, we\'re only talking a few percent of the time we
do this process. We can do an acid treatment and produce Type A or acid
bone gelatin. So the purpose of this process here is to condition and
ready the ossein material for an extraction at a very low pH. In the
traditional process, we use a sulfuric acid and we expose the ossein to
a pH of about in the range of 1 to 2 for about 6 hours, and then we
rinse that back to a pH ranging from 2.8 to 3.2. And this is where we
will extract the gelatin. This is, I mean, usually pH to extract
gelatin. Most gelatin is extracted at much higher, more neutral pH.

We also have an alkaline pretreatment option that we\'re looking at,
that some of our customers are looking at, because of all the discussion
around sodium hydroxide pretreatment. In this situation, you would do
this alkaline pretreatment prior to the ossein treatment. And in this
case, you are able to maintain the pH at 13 or greater with sodium
hydroxide for a period of three hours.

Next slide, please. Okay. Now, we\'ve finished with the pretreatment,
whether it be for acid bone gelatin or lime bone gelatin and the rest of
this will be common to both of these types of gelatins. Now, we extract
the gelatin. This is where we\'ve wetted the gelatin, we\'ve hydrolyzed
it, now we\'re going to actually pull this, extract this out of that
ossein particle to produce the gelatin.

We use demineralized water. What you\'re seeing up there is a typical
gelatin extractor. We do a series of extractions. I said 4 to 6
depending on the plant and the company, the way they do that. But the
initial extracts are done at a lower temperature, and what you will get
out is a material that typically has a higher molecular weight, a higher
viscosity, a higher bloom strength.

As you go to subsequent extracts, that material will become more
degraded. It will have a longer profile of treatment with time and
temperature. And those ending extracts will conversely have higher
collagens, lower molecular rates, lower viscosities and so on and so
forth. So the temperature range is from about 120 to 200 as you go
through that series of separate gelatin extracts that you are pulling
out. The conditioning time for each extraction ranges from 1 to 6 hours
and it\'s 4 to 6 extracts.

Next slide, please. When that extract comes off, it\'s a typical, very
dilute solution somewhere in the range of about 4 to 6 percent. So
you\'re saying to get to a dry product, we got to pull a lot of water
out of here as well. So we have initial filtration, this is a U.S. type
filter, vertical leaf type filter. It\'s precoated with diatomaceous
earth and cellulose. And that basically is to give us initial and
improvement in the clarity. The solution will also go on to ion
exchange. We want to protect these ion exchange collagens.

Could I have the next slide, please? So you\'re looking here at an ion
exchange battery. You see three columns in the forefront and three in
the background. Those are batteries of cation in that exchange columns.
Of course, the objective here is to deionize this material, depending on
whether it is pharmaceutical or photographic. It gets more exposure to
those columns depending on what is needed.

Primarily, the cations we\'re removing are calcium magnesium and iron.
On the anion side, it would depend on the acid that we were souring the
material with before we extracted it. And sometimes we use hydrochloric
and sometimes sulfuric. So those would be the primary anions that would
be removed under those conditions. And the finished product from an ash
standpoint would be somewhere between .1 and 1 percent, depending on the
product that we\'re making.

Could I have the next slide, please? Now, we begin to remove water, and
we do this by using evaporative means initially. So we have this 5
percent solution that we\'re going to drive up to a 15 to 25 percent
concentration. The evaporator you see there in the picture is a triple
effect plate and frame type evaporator. The output temperature is not
too high. It usually runs about 125 to 130 degrees on the average.
Basically, a temperature that will just make sure the gelatin doesn\'t
gel up in the production plant.

Could I have the next slide? Then we have another filtration. We heat it
again. We\'ve concentrated that material, so there is more particulate
becoming apparent, in certain cases, and then there is a chance that you
may get some coagulated protein, so we have another clarification step
here. The medium we use are exactly the same in the prior filtration
cellulose and diatomaceous earth, but we use a plate and frame pressure
filter. The viscosity of this solution is increasing now as we move
along in the process, and this is what requires a completely different
configuration for filtration.

Could I have the next slide, please? Then we take the opportunity to
adjust the pH, at this point. The final pH targets of the finished
product are usually in the range of 5 to 7. At this point, it\'s usually
just a fine adjustment and most typically it\'s done with sodium
hydroxide.

Could I have the next slide? Then we do our final concentration with
evaporative means. Again, this solution is becoming quite viscus, so
we\'re concentrating our thick, what we call at this point, our thick
gelatin liquor. This is an example of a double effect plate and frame
type evaporator as well. And the concentration here will be a fairly
broad range here from 25 to 50 percent, and this is because, I talked
earlier about your initial extracts are much higher viscosity, so you
only will be able to drive those up to about a 25 percent. However, the
latter extracts, which have a much lower viscosity, you\'re able to
drive those up to a much higher concentration level, and that\'s what is
done.

Next slide, please. Then we go through a sterilization step at the end
of the liquid phase. After this, we\'re going to be going into a more
solid mode with the gelatin production, so this is our last opportunity
to do something with the liquid phase. So we use direct steam injection.
We use a temperature that ranges anywhere from 138 to 149 C for 8 to 16
seconds, and this is primarily to ensure the product, hygiene of the
product.

Next slide, please. Then we\'re taking another tack here in terms of
drying the gelatin. We\'re beyond evaporative means, so what we do is to
increase the surface area, then able to dry this material, we cool it
down from about 120 down to about 70 degrees where the gelatin actually
sets, starts to set, and this is down with a glycol cooled heat
exchanger. Then it is extruded out through these perforated heads to
form these noodles, which will range in size from under 2 feet long and
about an 1/8^th^ inch thick, and they are deposited on the front end of
a dryer, which is in the next slide.

These dryers are typically about 12 feet wide and about 150 feet long.
The air quality we use is heated, dehumidified and filtered air. The
object is to produce a stable product. It has very low water activity.
Typically, it has 10 to 12 discrete zones with different temperatures.
There\'s a gradient that ranges from about 80 to 160 degrees fahrenheit
that goes across that entire dryer. It takes about like 2 to 3 hours to
get through this system, and the final moisture content of the gelatin
product is about 10 to 12 percent.

It\'s a very touchy process. It\'s very easy to melt the gelatin. If you
try to dry it too fast, you know, with too much water, the melting point
is lower and it is going to melt down or you can get case hardening.
It\'s a very delicate process drying this gelatin effectively.

Next slide, please. Then we do a milling after the drying and our size
is typically 8 mesh. That\'s our kind of working mesh size. We can do a
variety of mesh sizes in the finished product, but most of our
intermediate products we\'re producing these intermediate extracts that
we use to do our final finished blending, and it\'s typically about 8
mesh.

Next slide, please. So as these individual extracts, whether it be 4 or
6, come off there, they are separated on the dryer as discrete extracts.
Those individual extracts from daily production are individually blended
to make sure that there is no lack of homogeneity as that material is
processed across that dryer. So we blend those with homogeneity. We
sample those materials, as intermediate product, and those that go in
the dryers are weighed and go into storage as intermediate product for
future blending and mixing.

Next slide, please. So there is our inventory that we\'re building up
with our daily production, and then based on the specifications of our
customers, we build mixes and we formulate mixes with these individual
extracts that we have been producing. These are much larger blends. Some
of these are 10, 20, 40,000 pound mixes, so now we have a high capacity
blender that allows us to put those together.

Many times we\'ll make a much smaller small scale mix to make sure that
we can blend it properly, particularly if it\'s a new product. We can
hit the specification before we go to the large scale blend. So
sometimes there is a series of analysis that we\'ve done it two or three
times before we finish off the finished product.

Next slide, please. And then we provide that product once we are ensured
that it meets the specifications of customers. We\'ll package that up
using drums, FIBCs or small bags and then it is off to the customer.

I hope that has given you a quick \-- I had to go through that rapidly.
There is a lot of information to cover there, but you\'ve got that in
your handouts there. So I hope that was useful and I would be glad to
entertain any questions you have. And I also would like to invite you to
come out to see our facility in Sioux City, Iowa if you would like to
see first hand how we make gelatin.

CHAIR PRIOLA: Yes, Dr. Bracey?

BOARD MEMBER BRACEY: Yes, I have one question. You said in the cation
exchange process that you treat the product in a different manner
depending upon the end use, i.e., photographic versus other. So, in
essence, that suggests that there is the potential for control.

DR. DUNN: That\'s right. That\'s right. I mean, there are certain types
of food products where you may not go through the columns at all. I
mean, it depends on the ash content. Typically, the ash if it was
unprocessed, it could be as high as 2 percent, okay. In some cases,
there would be no need. And it would get very sophisticated with the
photographic realm whether you are interested in anions and cations, you
go through a cation and bypass an anion or you may go through a
secondary column.

You know, we have a battery with three columns of each type. Usually,
one is a lead column, lag column and then there is a regenerate one
under regeneration. So there is a variety of ways to go through that ion
exchange system, depending on what the specifications of the customer
are. You might have a food customer who says well, ash is less than 2 or
you might have a photo customer and it has got to be between .1 and 2.5
or .1 and .25 or something like this. There is all kinds of variations
on the thing in terms of exposure to ion exchange.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: I understand from Dr. Chiu that it is the
processors who are responsible for the safety of supplies. How is that
monitored or enforced here and abroad?

DR. DUNN: You\'re talking about the supply of our gel bone?

BOARD MEMBER BAILAR: Right.

DR. DUNN: Okay. We audit our suppliers. One of the things that makes it
a little bit easier here in the States is we only have a few. We
basically have \-- it depends on the company. Between the two companies,
I think, we have five or at most six different suppliers. So it\'s not
an unmanageable deal to go in and audit these customers on a regular
basis. We also know that USDA is in these plants. They help us with
this. As a partnership, they are in there auditing all the time.

For example, when we worked with the USDA because of these European
regulations to start taking our SRMs, back in 1998, they worked with us
to do that, to go in and validate those procedures and so on. So we have
an ongoing program in that respect and we work with the USDA sometimes
to do various things as well.

BOARD MEMBER BAILAR: What about foreign supplies?

DR. DUNN: All of our suppliers here in the United States, everything we
source is here in the United States right now.

CHAIR PRIOLA: Can you remind me, you said there were Type B and Type A
\--

DR. DUNN: That\'s right.

CHAIR PRIOLA: \-- process and the Type A is acid?

DR. DUNN: Type A is the acid. Type B means base.

CHAIR PRIOLA: Right. And why do you choose one of those others?

DR. DUNN: Like I said, we do very little Type A. I mean, very little.
We\'re talking probably less than a couple percent, something like 2 to
3 percent, and that\'s all directed to the pharmaceutical capsule
industry, and there are reasons for that. Because of the way we process
this material, the ratio of viscosity bloom and the ratio of viscosity
to concentration is very different. We can acquire a very low viscosity
concentration ratio with this process for acid bone.

And sometimes those customers who make the capsules require that they
have a higher concentration. And the limit usually is viscosity. So if
they can get a gelatin that has a lower viscosity to concentration
ratio, that allows them to bring more gelatin into that capsule, and
sometimes in the soft gel, it depends on the drug fill and what is going
on there, that can be very important. So it\'s very important for a
number of applications in the soft gel area.

CHAIR PRIOLA: All right. So even though it\'s a small percent of the
time you do this process, most of it goes to the pharmaceutical
industry?

DR. DUNN: That\'s right.

CHAIR PRIOLA: Then the sodium hydroxide option, the base treatment, you
said that\'s under review. Is that to see how that might effect \--

DR. DUNN: That\'s right. That\'s under review for acid bone. The most
important thing it\'s under review by our customers, and they are
currently evaluating that to see if there is not any other shortcomings
of the fact that the sodium is there as opposed to the calcium from the
lime.

CHAIR PRIOLA: Does it seem to change the end product at all?

DR. DUNN: From our prospective, it doesn\'t, but that\'s why we\'re
relying on the capsule manufacturers to do their full evaluation and
that\'s what we\'re looking for. Okay. So we can do it. It\'s easy for
us to do. It\'s not a problem for us to do that.

CHAIR PRIOLA: Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yeah, I have a question regarding the bovine bone
sourcing practices. You said since 1998, you have been removing the
specified risk materials, except for vertebrae.

DR. DUNN: Yes.

BOARD MEMBER KHABBAZ: Why that exception and is it still practiced?

DR. DUNN: That\'s a very difficult thing to do, and there is really up
until recent times there has been no requirement. There are EU
regulations now developing and that\'s why there is concern there that
that may be a requirement coming into place as early as the end of this
year. We\'re not sure how this is going to roll out, so we\'re looking
at this strategically. Right now, there is not a requirement, but there
is a big hurdle there in terms of industry\'s ability to do this.

This will cost us more money. It will reduce the amount of bone
available. Right now, if you take the vertebrae and take it somewhere
else, you reduce the quantity right there by 25 to 50 percent. And then
there will be certain facilities that will just not be able to do this
with the equipment they have. They won\'t be able to make this change
without investing new capital. But anyway, the prices we are seeing now,
you can get this material, small masses of this material now, but it is
going to cost you 50 to 100 percent more than the traditional. So, I
mean, nobody wants to go there unless we have to. It\'s going to be very
costly for us, our suppliers and our customers.

CHAIR PRIOLA: Okay. Thank you very much, Dr. Dunn.

DR. DUNN: Thank you.

CHAIR PRIOLA: I think we\'ll move on to the next speaker. It will be Mr.
Schrieber, who will describe the European manufacturing processes for
gelatin.

MR. SCHRIEBER: First slide, please. I would like to thank you, Dr.
Priola, this Committee and the FDA for the opportunity of presenting on
behalf of the Gelatin Manufacturers Association, GME. Again, details
about raw materials sourcing and the bone gelatin manufacturing
practices in Europe. My name is Reinhard Schrieber. I\'m the chief
manufacturing officer operating GELITA Gelatin Group. I\'m 36 years in
the gelatin business, and I have served at European Gelatin Association
for many years as president, chairman of the regulatory committee and
the chairman of our BSE Steering Committee.

After my American colleague, Mr. Dunn, has already substantially
presented the details of the bone gelatin manufacturing process, I would
like to go only relatively shortly into this issue. The manufacturing
processes in general and although the bone gelatin manufacturing
processes in particular are very similar to each other, not only in the
U.S. and Europe but all over the world. The main differences which can
be noticed between the continents are related to the safety status of
the raw material and the sourcing systems in place.

This is why I like to focus more on these topics, whereas I would like
to try as well to connect the connections and the conditions of our
study to those existing in reality.

Next slide, please. GME members have taken several voluntary steps to
ensure the safety of the raw materials. Long before the emergence of
BSE, the European gelatin industry has decided to use, and this applies
for all types of raw materials, only raw material coming from healthy
slaughtered animals and released for human consumption, regardless of
whether this was mandatory or not in different member states. So we
don\'t use any materials from fallen or sick animals.

So traditionally, no material from fallen animals have been used by
European manufacturers. The three bones gelatin manufactures in Europe
have never used UK bones, but when BSE in the UK became evident, they
confirmed immediately in writing not to use UK bones. After the
condition of BSE to humans was detected, the GME members committed
themselves to stop the use of skull bones, the target which was reached
in 1997. This was further followed by the complete removal of spinal
cord by European meat packers only on request of the European gelatin
industry.

In parallel, our industry started to replace European bones to a certain
extent by imported bone chips, mainly from the United States, but also
from other countries outside Europe. In 1999, the European gelatin
industry was able to convince its suppliers to remove vertebrae from
bovine bones of all ages, which again was more than European law
required.

Next slide, please. As I stated before, on top of our European sourcing
of our demand for bovine bones can only be covered with additional
imports from different countries. So we always force our suppliers in
GBR II countries to voluntarily take measures in order to increase the
safety of our raw materials. GBR II country means that there are so far
no BSE case detected and the European has assessed that it is unlikely
that there will be a case, but it cannot be excluded.

The U.S. is and Canada has been until recently GBR II countries.
Together with our American colleagues, we implemented the removal of
spinal cord, also in the U.S., and one year before we succeeded in doing
so in Europe, we had forced our suppliers in India, Pakistan, Nigeria to
remove the vertebrae as a precautionary measure.

Next slide, please. Most of the measures which we had already
implemented became mandatory by regulation in Europe some years later.
On top came the postmortem rapid testing of all cattle older than 30
months. Furthermore, the removal of vertebrae as requested now by law
only for animals older than 12 months, but again in the bones we use in
Europe, there are no vertebrae in at all. So in practice, the vertebrae
is removed from all cattle in the European Union if the bones are
intended to be supplied to the gelatin industry.

I assume that you are aware of all those regulations presented to you, I
think, by Dr. David Asher in February of this year. With gelatin
regulations, the EU fixed raw material sourcing conditions and certain
safety relevant procedures to all kinds of food grade gelatin. This has
been presented to this Committee two years ago by my colleague, Dr.
Scheigas. Those requirements are in line with the new study conditions,
and our regular intervals controlled by public veterinarians responsible
for the supervision of our plants, although the FDA has made audits to
the gelatin bone manufacturers in Europe two years ago, they went to all
plants.

Next slide, please. Because of the steps taken by the industry, there
was always only a very little chance that BSE infectivity could be
present in the raw materials used to produce bovine-origin. To date, due
to additional more recently implemented controls like the postmortem BSE
testing and the careful removal of all SRM, it is almost impossible for
highly infected material to enter our supply chain.

Next slide, please. As with any process and systems, there is a certain
possibility of error. What could happen, for example, animals with very
low infectivity might not be detected by the rapid BSE test. But they
are considered today as to posting no risk to human health. The
surveillance systems in place might not be adequate in all countries.
The removal of SRM may not be done perfectly. The infectivity of bone
marrow has not been finally clarified. Based on our experience, we
believe that those risks are low, but they are not negligible. They will
be quantified by the Scientific Steering Committee of the European Union
and then used in the coagulation of the quantitative risk assessment,
which is currently under development.

Next slide, please. Last year, more than 9 million normal slaughtered
animals were tested on BSE within the whole European Union, including
the UK. And 287 positive cases were found, which gives a ratio of 1 to
50,000. But our tests which had been done and our study has assumed that
all animals used were clinically infective. Supposing that the removal
of SRM is not effected perfectly and that those impurities may not be
detected by the gelatin industry when inspecting the incoming fresh
bones, some might enter the process. Again, our tests and our study have
assumed that the bones from all animals contained the food quantity of
infective spinal cord and dorsal root ganglia. Well, this gives a huge
safety margin between the study conditions and reality.

Next slide, please. Here again, the major production steps applied
during the commercial and the study manufacturing processes, most of
them have already been described by Dr. Dunn. All plants in Europe are
ISO 9000 certified for the quality management and they apply the HACCP
principles. The combination of those is about equivalent to GMP. FDA
audits have been successfully conducted in all European bone gelatin
operations two years ago. And a further round of audits is scheduled for
the end of August and early September this year.

It has to be noted that SGS and independent institute specialized in
quality certification carried out a validation audit. And each of the
bone gelatin plants of GME in Europe and there are no known GME bone
gelatin plants in Europe, and by these inspections all processed
parameters of our study design have been validated against minimum
production conditions in place in those plants.

Just to clarify what this means, minimum conditions. In certain plants,
for example, a higher concentration of the hydrochloric acid or a longer
liming time might be applied by one or the other manufacturer compared
to the conditions of this study. But we used in our study the minimum
conditions applied at least by all manufacturers.

Next slide, please. One of the differences in Europe compared with the
rest of the world is the fact that in Europe bone gelatin manufacturers
have their own bone degreasing plants. In other countries, like the
U.S., degreasing is part of the meat packers work. In the Far East, for
example, it is effected by independent specialized companies. As
mentioned before, only bones from healthy slaughtered animals released
for human consumption following audit and postmortem inspection are
collected from the meat processors, who do then later the deboning of
the carcasses.

In the U.S., slaughtering and deboning is done normally at the same
premises. In Europe, we have very often different locations. So this
means that the carcasses of the animals are transported to a sausage
manufacturer, to a meat packer at a different place and during this
transport, the bones are still with the carcass. Only the SRM, the
spinal cord, the heads are gone, spinal cord is out, but the bones are
still with the meat.

The incoming uncrushed bones are then inspected by the gelatin industry
on sorting belts for extraneous materials, including potential SRM
contamination. Then the bones are crushed to small chips of about
5/8^ths^ of an inch, this fingernail size. Then the bones \-- this means
after crushing that we have a big surface. And for example, with the
hollow long bones the inside would as well become an outside.

These small bone particles are then degreased by hot water in a
continuous flow process at approximately 185 degree fahrenheit of an
average period of about 20 minutes in equipment with high education.
This mix of water, temperature and movement separates fat and soft
tissue from the solid bone particles. The little ones are then separated
by sieves and cyclones, dried with hot air, but the surface temperature
of the bone particles will stay below 150 degrees fahrenheit to avoid
degradation. Then they are sieved to remove fine particles and stored in
silos.

Next slide, please. Demineralization to remove the phosphates from the
bones is carried out at the same conditions like in the U.S. in a
conduct current system. The total treatment is about 4 days with
hydrochloric acid of 4 percent. The remaining protein matrix of the
bones is called ossein.

Next slide, please. To cut the cross veins of the collagen acid or
alkaline can be applied. This was addressed just before. For a small
portion of the total bovine bone gelatin production, it is about 2 to 3
percent for special pharmaceutical soft gel capsules. The ossein is
treated again for 24 hours with sulfuric acid at the low pH and after
some washes, the gelatin can be extracted at a pH between 2 and 3.

So standard bovine bone gelatin is normally extracted at a pH between 6
and 7. And the ossein is treated before the saturated or over saturated
lime solution for at least 20 days. As you have heard, the pH of this
lime solution, which is replaced several times during the process, is
around pH 12.5.

Next slide, please. To make sure that acid bone will be as safe as lime
bone, our industry looked into an alternative process which would
include an alkaline pretreatment, but without working the special
physical and chemical properties of this pharmaceutical as in bone
gelatin. Based on the knowledge that after the bones are crushed
potential infectivity would sit on the surface of the bones and not
inside the bone matrix, we assumed that a short time treatment of about
2 hours with .3 molar sodium hydroxide solution should be enough to
inactivate infectivity if this pH is kept at 13 for this time.

Our study results have shown that this treatment is very effective. But
our study has also shown that gelatin made by the traditional acid bone
process did not show any detectable remaining infectivity, which means
there is a demand for this type of gelatin is still very rare. You\'ve
heard that we are depending, of course, on our suppliers to do it or not
to do it.

Next slide, please. During extraction of the pretreated raw material,
several single extracts are collected, each with different physical
properties due to an ongoing hydrolysis during the extraction. It has to
be stated that due to the different requirements of the gelatin using
industry, quite often photographic, pharmaceutical and food grade
gelatins manufactured from the same raw material batch in sequence.
Also, Eastman-Kodak is manufacturing some pharmaceutical and some food
grade gelatin.

This means that all gelatin of one production day, including the
photographic gelatin, have to comply with the regulatory requirements
for food and pharma. When talking about food and pharma, one has to keep
in mind, as well, that the same capsules might be filled today with
nutritional products, being food, and tomorrow with Rx drugs.

Next slide, please. For further clarification, the diluted gelatin
solution is filtered by different types of equipment and filter media in
the ossein and ion-exchange columns and concentrated in the apparatus.

Next slide, please. So final concentrated gelatin solution is sterilized
by direct steam injection. The temperature is at 4 bar. The pressure in
the liquid phase, which is very important, is a minimum of 280 degree
fahrenheit and the temperature stays for at least 4 seconds.

Next slide, please. Finally, the sterilized gelatin solution is chilled
to set and then dried with purified and conditioned air on belt dryers.
Each production batch, which is a single extract, is then tested on
physical, chemical and bacteriological properties. According to customer
specification, different production batches are then dry blended. The
final blends are again tested under compliance with regulatory and
customer requirements and then released for shipment. These are the
common processes applied by the European industry.

Next slide, please. There is one special process which is done by only
one company in Europe to manufacture gelatin with low gelling strength
for limited applications. The degreasing is done of the bones in the
common way, but then the bone chips are autoclaved for at least 20
minutes under 3 bar pressure and 270 degree fahrenheit. After the
autoclaving, bone chips are rinsed with salt water. A certain quantity
of gelatin goes into solution.

After this gelatin solution is taken out, autoclaving at lower
temperature and shorter time is repeated several times. Then these
different extracts are collected, flocculated, ion-exchanged and
evaporated, drying, testing, blending, retesting and shipping is
effected, like with all other gelatins. Low gelling strengths, the
gelatin is used only for certain applications, and the Committee members
might remember that two years ago at this meeting, we already explained
that the main application is a confectionery licorice, although this
process has been successfully simulated during our study.

Next slide, please. What are the conclusions which we have drawn from
this review presented here? So commercial mineral manufacturing
conditions are reflected by the GME study conditions. The GME plants and
process parameters have been validated for conformity against the study
design. The inactivation results of the study, which will be presented
next, are therefore fully applicable to the practical gelatin
manufacturing processes. The study demonstrates the ability of the
gelatin manufacturing process to remove and inactivate infectivity even
under conditions in which raw material contain unrealistically high
infectivity levels.

Last slide, please. So safety of European bone bovine gelatin is
established on two principles. The safety of the raw material as
required by GME practices and EU law and the safety of our manufacturing
processes as demonstrated by the GME study. The Scientific Steering
Committee of the European Union has concluded based on all these
principles, in it\'s opinion, on the safety of gelatin that the risk is
close to zero.

Madam Chairman, Committee, that concludes my presentation. I would like
to thank you and the Committee for your attention. Thank you.

CHAIR PRIOLA: Okay. Thank you, Mr. Schrieber. Are there any questions?
Okay. Thank you very much. Our schedule says there is a break, I
believe, after Mr. Schrieber, so we can adjourn and return at 10:00, so
that\'s about 15 minutes from now. All right. Thank you.

(Whereupon, at 9:42 a.m. a recess until 10:03 a.m.)

SECRETARY FREAS: We\'re going to go ahead and resume the meeting.

CHAIR PRIOLA: Okay. I would like to go ahead and get started. Dr. Hogan
had a question for our last speaker, Dr. Schrieber, that he would like
the Committee to hear the answer to. So, Dr. Hogan, do you want to?

BOARD MEMBER HOGAN: Mr. Schrieber, I asked just after our last talk
about how the meat processors were audited, in terms of providing safety
of the raw materials to the gelatin manufacturers. Could you address
that, Mr. Schrieber, please?

MR. SCHRIEBER: Surely. The standard procedure in Europe is that in every
slaughter house, every meat packing operation there is a public vet
present all time, every day as long as this operation works to supervise
that regulation is followed, removal of SRM is done and so on. And
besides this, the gelatin manufacturer are auditing their suppliers on a
regular basis, normally once a month or every two months, again
inspecting as well the commercial documents about where the animals have
been sourced, because commercial document which is required as well by
law. So there\'s a double-fold. But the main thing is that the public
vet is present all day, all the time. Thank you.

CHAIR PRIOLA: Okay. Thank you, Mr. Schrieber. We\'ll go on to our first
speaker for this later morning session. That\'s Dr. Robert Somerville,
who is going to discuss the GME validation studies on bone gelatin.

DR. SOMERVILLE: Okay. Thank you, Madam Chairman. It\'s a pleasure to be
back in the USA where I\'ve spent quite a few happy years working a
couple of decades ago. My task is to describe to you the actual
validation studies that were performed in three labs actually over
several years. There were several people involved and I want to mention
them. First, Ad Grobben is perhaps the most important one of them all,
because he, as an employee of gelatin, which was a member of GME,
actually performed or was participating in all three studies in
Edinburgh, in Holland and here in the USA in Baltimore.

Phil Steele, actually I should say that, I know a consultant to GME and
is present in the audience and I hope will assist in any different
questions you might ask me later on. Phil Steele is a technician in my
group and he assisted at not only in the work he did in Edinburgh, but
also in Holland to assist in the experiments there.

David Taylor was my predecessor in running the inactivation group, and
he initiated the studies that we\'re about to describe and collaborated
in setting up the whole thing. He again is here in the audience. I
inherited the work from David and responsibility for the work when David
retired in 2000, so it\'s my duty to report the results, but all the
hard work was done before that.

The work I\'m specifically going to concentrate is on the
Neuropathogenesis Unit, which is part of the Institute for Animal Health
in Edinburgh. It was funded by GME with further support from the
European Union. I should also say that at the end of my presentation, I
suggested that Bob Rohwer, who performed the Baltimore studies, spend a
few minutes describing the work that was done in Baltimore.

The next slide, please. I thought it would be helpful to describe the
basic mechanisms of TSE inactivation first, and there are three ways in
which inactivation or removal can take place. The first is through some
form of destruction through combustion, incineration, oxidation with
hypochlorite, hydrolysis of extreme pHs or with very high concentrations
of highly effected proteases and radiation can have an affect at very
high doses.

Next section, please. What possibly concerns us mainly today is
denaturation type of processes where materials hydrated, in particular,
will have a degree of inactivation effect and exposure to chemicals,
such as strong detergents or chaotropes, can also have an effect.

Next. And we have to look at treatment variables. There are several
biological parameters that we must consider. The strain of the TSE agent
is particularly important and I will illustrate that in a couple of
slides time. The PrP genotype may well be important, and we have to
consider that. The tissue and the state in which the tissue is presented
in the experiment are also important. We have to consider
physicochemical parameters such as heat temperature, pH and the kind of
chemicals that one uses. And finally, the dynamics and kinetics of the
reaction have to be considered, the time, concentration of any chemicals
involved and the temperature are particularly critical.

Next. Okay. This shows a slide of some data that was originally
published in 1983 by Kimberlin, et al, where TSE infectivity from two
TSE strains was heated for various lengths of time shown on the axis. On
the Y axis is the titre that was recovered after these treatments. Two
strains were used, as I say the 22A strains and the 139A strain, and you
can see that there is a lot of rapid reduction in the amount of
infectivity present, first. Then a plateau. So the reaction is biphasic
with respect to time, and there is little effect of time after initial
exposure.

The second point to notice is that was a strain difference, so the 22A
strain on this particular example is much more resistent to an
activation than the 139A strain.

Next. Now, in this slide, we\'re looking at what happens when we heat at
a constant time, 30 minutes, with a range of temperatures, and what we
can see here is that there is little reduction in infectivity to start
off with until we reach an inflection point, and then the amount of
infectivity drops rather rapidly, and that happens for both the TSE
strains that we\'re looking at here. But you can see that the inflection
point for these two strains differs, so that for 22C, it\'s rather
thermolabile, which might be a surprise to some. In fact, this
temperature which starts to inactivate is only about 70 to 75 degrees
centigrade.

With 22A it\'s higher, about 97 degrees. But we haven\'t specifically
done experiments comparable to this BSE or BSE derived strains, although
I\'m hoping to do them in the near future. But from the data that we
have available, we think that BSE derived strains are even more
resistent to inactivation than the 22A strain here in red, which is the
more thermostable of the ones we have seen. So we can say the
inactivation process is biphasic with respect to temperature and
dependent on temperature. TSE strain, and I mentioned the hydration
state, and I\'ll come on to that in one moment.

Next slide. Okay. This slide shows the effect of hitting again at 126
degrees centigrade for 30 minutes autoclave, three strains of TSE, and
you can see with the 22C strain that all infectivity shown in red has
been destroyed. The blue shows the starting titres. The two different
blue bands are indicative of two different PrP genotypes that the TSE
strains were passaged in, and there is no effective PrP genotype in this
experiment available.

With ME7, we cover both the types, a little infectivity, but with 301V,
we cover a lot more. Now, 301V is important to the rest of this talk.
301V is the most thermostable TSE strain that has been derived from the
passage of BSE through a particular strain of mice, the VM strain of
mice. And it has certain advantages to these studies. Notably, it is
very high thermostability, and that makes it a greater challenge to the
studies that we are performing.

On the right hand side of the panel, you can see a different experiment
where material was heated in a dry oven to 200 degrees centigrade for
either 20 minutes or 60 minutes. And I think the contrast between what
happened in the autoclave and the dry oven is really quite remarkable.
We get much less reduction in infectivity and we\'ve lost our strain
differentiation. So there\'s no strain difference in the results. And
also material survives the dry oven much better than it does in the
autoclave. So that emphasized the point about hydration status. I think
if we dry out infectivity, we make it much more resistent to
inactivation.

Can I have the next one? Okay. This is an experiment where we have
combined temperature treatments with a range of pHs. I don\'t suppose
you can read this, but each line represents a different pH from pH 7 up
to pH 12 with three strains of TSE again, 301V, ME7 and 22C. And the
point is to say that with 301V, in particular, we got very little
reduction in the amount of infectivity up to 100 degrees. Certainly up
to pH of 11.

We didn\'t measure what happened at pHs greater than 11, whether we were
getting any reduction infectivity at pH 12 up to 100 degrees, but at 60
and below there was very little reduction in the amount of infectivity
recovered there. You do start to see, in effect, that pH 12 with the
more thermolabile strains ME7 and 22C. So the suggestion is that high pH
acts synergistically with temperature when TSEs are inactivated.

Can I have the next slide, please? So on the left you can see a list of
the things that I have been showing on all previous, three or four
slides, and results and conclusions. Thermostability is an intrinsic
property of TSE agents developed to kinetic mother which I\'m not going
into today, and so forget about the rest. Thank you.

Next slide. Okay. I want to move on now to reduction of risk of TSE
infectivity in gelatin. The challenges that we face, there is very high
resistent to inactivation, and the resistance increases on drying up
infectivity. There are several available approaches. We can remove by
filtration, for example. We can denature with heat and high pH or we can
use at very high pH, we can get hydrolysis of infectivity.

Looking specifically at the risk reduction steps that are available in
gelatin manufacturing, the sourcing of bones, which Mr. Schrieber has
just described, is important as practices in precleaning the raw
materials. I\'m not going to discuss this. The standard gelatin
extraction methods are thought, were thought to be effective, and that
is what the valid study of it we\'ve been involved in is designed to
test. And then the sterilization steps are steps which may specifically
move TSE infectivity and, of course, other contaminants.

Next slide. Now, this slide shows the results from the very first
studies that were performed in the gelatin manufacturing process. What
was done by Inveresk was to take any 7 grain homogenate and look at two
components of the process, either treating with hydrochloric acid, the
liming step or the combination of the two. The reduction in titre after
exposure to the hydrochloric acid was about 1 log, 1.2 was measured. So
that\'s 10-fold roughly.

Exposure to lime for 20, 45 or 60 days resulted in a reduction in titre
of about 2 to 2.3 logs. And you can see that even after 60 days, these
values are very similar. So there was a small reduction of about
100-fold, but the time of exposure had new extra effects. And the
combined treatment results in the reduction of nearly 3 logs, but you
can see that adding these two values together does not come to 2.8. So
there isn\'t complimentary effect, but the treatments are not completely
out of it.

Can you give me that slide, please? So there is a reduction in
infectivity titre measured by the acid and alkali in combined treatment
of any 7 homogenate. The combined treatment is more effective than
either single treatment, but they are not titre, and time of exposure to
costs in hydroxide does not effect infectivity titre. And these
processes were not representative of the actual process involved in the
plant.

There is another study by Manske, et al, which showed that there was
removal of proteins under industrial degreasing conditions. These
initial studies led to the desire for more systematic studies to be
performed.

Next slide, please. As I\'ve already indicated, there was several
experiments performed. In Edinburgh we performed four experiments, two
alkaline treatments using two TSE strains, the 301V strain, which I have
described, also the 263K strain, which is a hamster-facade strain, which
we believe is reasonably thermostable, but may not be quite as
thermostable as 301V.

We also looked at an acid process and we tested the addition of an NaOH
treatment in the acid process. Mr. Schrieber has described the Dutch
heat and pressure method and an experiment was performed in the
Netherlands to look at that process. And as I say, Bob Rohwer will
describe the sterilization filtration experiments later.

Next slide. Okay. The rational of the experimental design. The TSE
source is a high titre BSE derived model. It\'s thermostable. It\'s
readily assayed in experimental mice. We feel that the total titre is a
likely BSE contamination event during industrial processing the gelatin,
as Mr. Schrieber has already suggested. Short incubation periods, but we
have to be aware that we occasionally see very extended incubation
periods after heat treatments. And so we kept the mice under observation
for up to 600 days.

The limits of detection depend on concentration of the sample and the
toxicity of the sample. We cannot inject material that is toxic,
obviously, to the mice. So sometimes dilution factors had to be included
so that we could inject the mice and not limit the clearance levels that
we can measure. However, we feel that near optimum demonstrate clearance
levels are demonstrated from this model.

The scaled down to simulate typical gelatin manufacturing conditions was
performed by Ad Grobben from earlier and that was reviewed by an
international panel prior to initiation, and as already indicated, Ad
Grobben is here to answer specific questions on that matter. And the
quality of gelatin was checked as the experiments proceeded and again I
can address those questions.

Next slide. Okay. So this is what was done. The raw materials were 1.5
kilograms of fresh crushed bones and 500 grams of intact calf backbone,
spiked with approximately 10 grams of TSE infected grain homogenate.
Half this back was injected into the spinal column and the remainder
smeared onto bones and dried onto the surface, and the backbone was then
sawed into pieces. There was a degreasing process where the bone chips
were washing at 85 degrees centigrade to remove soft tissue and fat
after the spike had been added, and then dried in the hot air at 120
degrees centigrade.

Then the demineralization step was performed. The bones were soaked in
hydrochloric acids of increasing concentrations. The ossein, of course,
remains as already described. Then the liming process, the ossein was
exposed to saturated calcium hydroxide of pH 12.5 for the minimum of
three weeks, and then neutralized. On the acid treatment left over
night, also a pH 3 and then washed in water. The NaOH treatment, which
is included in the acid experiment, one acid experiment, the ossein was
exposed to .3 molars of sodium hydroxide pH 13 for two hours.

Then the extraction process ossein was stirred gently with water at
temperatures from 60 to 90 degrees to a final gelatin concentration of
2.8 percent. And then purification steps were performed, depth
filtration, ion-exchange, heat sterilization and drying, and all steps
were designed to accurately represent the conditions of the industrial
process.

And it should be pointed out that in the larger process we used indirect
heating, but in the industrial skill process, of course, direct steam
injection is used.

Could I have the next slide? Okay. The spike, as I\'ve already
indicated, we use the 301V strain in three of the four experiments, and
we use it because it is the most thermostable TSE strain tested so far,
and it also is BSE derived. We actually titrated the spike on three
separate occasions. We actually had two spikes, Pool 1 and Pool 2, and
you can see the values are very similar in all three titrations that we
performed with a value of about 7.7 in each case.

And as I\'ve already mentioned, all clinically negative animals were
observed for at least 600 days, and then we examined the brains for any
evidence of pathological lesions of TSE infection afterwards. And all
positive clinical cases were confirmed by pathological examination.

Next slide. Okay. Some results. So this is the first experiment where
the bone was spiked with 301V. The steps were performed degreasing,
demineralization and DCP, the dicalcium phosphate, which is a byproduct
of the gelatin manufacturing process, was also tested for residual
infectivity and we find little. The extract sample after the liming
initialization extraction had a little bit of infectivity here, and you
can see the individual numbers on the left, and that calculates,
according to the Carver Method, to titre of less than or equal to
10^1.8^ ID~50~ per mil.

I say less than or equal to, because if you use the Carver Method, you
have to make it \-- and you\'ve got incomplete groups at either end of
your dilution series, you have to make assumptions about what happened
in that group. So we don\'t have a 10^+1^ group, but we assume to get
the number 1.8 that that value, all the mice would have gone down the
10^+1^ dilution. So the number over here is the total recovery
calculated against the 10 gram spike that was used. So we got from total
infective load to 10^8.7^ to total recovery of 10.5.

Then the sample was taken through the filtration ion-exchange in
concentration steps and the sterilization steps, and a sample was then
also measured for infectivity, and no infectivity at all was recovered.
And in this case, what these data say is that we couldn\'t detect
anything. We don\'t know what would have happened if we had been able to
inject a more concentrated sample again. So again, we can only say this
is the limit of the clearance that we have achieved. So the total
recovery is less than 10^3.8^ starting with the 10^8.7^.

Okay, next slide, please. Okay. So this is the second experiment in the
alkaline process where we used the other strain, 263K, and I will go
through this a bit quicker. The total infective load is 10^9^. We
recovered a little infectivity again and the DCP, the dicalcium
phosphate, and we also recovered a little bit of infectivity in the
extract sample totaling out to a total recovery of less than or equal to
10^4.3^.

Next slide. The acid process here we again had a spike which had a total
infectivity of 10^8.8^ and following the steps of degreasing,
demineralization, then the acid treatment and extraction we had a
recovery of infectivity of 10^6.2^. In this case, we got a clear end
point to the experiment, because the neat fraction, all the animals came
down to 10^-2^, none did, so a nice, neat Carver calculation of 10^6.2^.

After the filtration, ionization, concentration, sterilization steps no
infectivity was recovered and we can say that is less than or equal to
10^4^ logs of infectivity were recovered.

Next slide. Okay. Now, this is the variation on the acid treatment where
the sodium hydroxide step was included after the acid treatment had been
performed. And when this was done, we find that no infectivity at all
was recovered in the titration and we can calculate total recovery of
less than or equal to the 10^3.3^.

Next slide. Okay. Looking at the data across the way, that\'s the first
alkaline process and you can see it went from 8.7 to less than 5 down to
3.8 with no positives.

Next. Next? Oh, there we go. Thank you. And this is the alkaline process
with 263K, and now you can compare the numbers directly with each other.
So we start off with slightly higher spiked titre and slightly lower
recovery of infectivity at this point on the crude gelatin extract.

Next. And next again. And with the two acid process experiments, we
start off with the spike of 8.8, 8.7. We recover a little bit more
infectivity than in the alkaline process at the crude gelatin extract,
but again when we look at the purified material, no infectivity is
recovered and we can, as I already said, indicate the clearance values
from that part of the process. And you can see now that the acid process
with the included NaOH treatment in here resulted in no infectivity
being recovered.

Next. So this summarizes the data and now I have included on the right
hand side the clearance factors that have been obtained from the
experiment. So we can say that the alkaline process, the crude gelatin
extracts have a clearance of greater than 3.7, logs of infectivity and
for the 263K experiment it was greater than or equal to 4.7. The
finishing, the purification and sterilization steps have additional
clearance factors that we have demonstrated of greater than 1.2 and that
totals over the two parts of the process to greater than or equal to
4.9.

In the acid process, we got a clearance from the \-- in the crude
gelatin extract of 2.6. The sterilized gelatin after finishing has got
an additional demonstrated clearance of greater than 2.2 and adding that
together, we\'ve demonstrated a clearance for greater than or equal to
4.8. And then in the acid process with the additional NaOH treatment,
the overall clearance demonstrated is a value of greater than or equal
to 5.4 logs of infectivity.

Next. Okay. So that summarizes what we have. From the acid bone process,
we have got substantial infectivity measured before purification in the
third experiment, but complete appearance after \-- complete clearance
after purification, including sterilization. Complete clearance, no
infectivity detected before purification if an additional sodium
hydroxide step is included.

With the alkaline process, there is greater removal of infectivity than
after equivalent acid hydrolysis procedure and there was complete
clearance. No infectivity detected after purification, including
sterilization.

Next. So our conclusions are that the gelatin manufacturing procedure
was successfully scaled down and normal bone gelatin was produced. Both
degreasing and the standard acid and alkaline treatments alone remove
most, but not all, of the implied infectivity before final purification
of gelatin. The liming or alkaline procedure was more effective and the
additional sodium hydroxide step in the acid procedure inactivates a
residual detectable infectivity before purification. After purification,
all samples do not show any detectable infectivity. And again, this was
pointing out the removal of infectivity is cumulative, but not
necessarily additive.

Okay. I want to move on to report the data obtained by the Dutch
experiment where they applied pressure treatment to produce their
gelatin. They started off with titre of their spike, total titre of 9.2.
They went through the standard procedures of degreasing and preheating,
then did the pressure treatment at 3 bar, 20 minutes, 130 degrees
centigrade, then extracted the gelatin. In the crude gelatin extract,
they showed that no infectivity could be recovered, and the volume that
comes to is less than or equal to 0.2.

So they record a clearance factor of greater than or equal to 6.8 with
this process. They did nothing \-- well, they didn\'t follow this, the
purification steps. They didn\'t test that, but there was no infectivity
in the gelatin solution that would have come through this procedure
anyway, so it would have been a waste of time.

Next. So risk reduction and gelatin. We\'ve had several descriptions of
this already in the earlier session about sourcing using only animals
passed fit for human consumption, omission of head bones and vertebrae
from source material in BSE infected countries, and we have shown the
removal or inactivation steps or removal of TSE infectivity during the
gelatin extraction and purification process.

It\'s also worth noting that the species barrier would reduce the effect
of titre or BSE being \-- if humans were exposed to BSE from the source.
It is worth also noting that the acids were performed by injection
intracerebrally, and this is by far the most efficient route of
infection, other routes of infection are less efficient.

Next. I\'ll skip that. That\'s it. Okay. Thank you very much.

CHAIR PRIOLA: Are there any questions for Dr. Somerville? Dr. Bailar?

BOARD MEMBER BAILAR: I have a couple of related questions. First, I find
the time deactivation curves somewhat troubling. They suggest that some
of the infected agent is being protected somehow. What is your take on
that?

MR. SOMERVILLE: Exactly that. That there is \-- I didn\'t want to get
too much into the fundamental thoughts that I\'m having at the moment
about that, but I think we\'re getting a dissociation reaction and a
protective reaction occurring when inactivation, heat inactivation is
attempted. And the protected species that is formed or the stabilized
species is much more difficult to inactivate. It may be similar to the
dried material that I was showing in some of the earlier slides, too,
and that we know is much more difficult to inactivate.

BOARD MEMBER BAILAR: Well, there are at least a couple of other possible
explanations.

MR. SOMERVILLE: Sure.

BOARD MEMBER BAILAR: One is that some of the agents being protected
inside little particles. There many be subtle differences in the
chemical structure of the ones that survive versus those that don\'t.
Which leads me to my second question. In the intact animal, the
infection occurs while the animal is alive. It gets circulated and I
would presume gets distributed throughout all the tissues and whatever
titre is appropriate for that. In the experiment, the infective agent
was added to the bone chips, that is at a considerably later stage of
things, where it might be more on the surface of any particles that
remain or it might stay on particles and so forth. So I\'m asking if you
have looked into this, and if there is any reason for concern about this
difference in the sequence of when the infection is added to the
materials that you are processing.

MR. SOMERVILLE: Well, let me answer this, your question this way. I
don\'t know if it actually addresses what you are saying. But the reason
for doing the experiment the way we did it was to try and maximize the
exposure in the experiment. So the thinking was that the greatest risk
of BSE contaminating bones was that during the slaughter process, and
that spinal cord, for example, would get spread down the vertebrae
column included with it and dry onto it. So that was what was attempted
to be mimicked in the experiment.

I\'m not \-- I suppose the other side of the question is how much
infectivity in living animals associated with bone and bone related
tissue? Our primary concerns in that respect again is to do with spinal
cord in BSE infected cattle with spinal cord and ganglia and related
nerves and, of course, the brain in the skull. These should be removed,
and again we\'re asking the question what happens if they don\'t, and
we\'ve tried to include that kind of thought in the design of the
spiking of the experiment.

BOARD MEMBER BAILAR: Have you tried to grow out the infective agent that
survives the steps for 20, 40, 60 minutes to see if it remains highly
resistent?

DR. SOMERVILLE: No, not formally. It\'s an experiment I want to do,
obviously, but I haven\'t formed it. I don\'t think \-- David Taylor
whether he has actually done that experiment, either. My prediction is
that it would not be in the protected form after passage through an
animal, but we have to do the experiment. Thank you.

CHAIR PRIOLA: Dr. Petteway?

DR. PETTEWAY: Thanks. I just have a couple of questions about the
process of doing the studies and setting them up. Just to make sure I
understand, these were scaled down, coupled steps, so that the spike was
at the initial step and then removal was monitored throughout the
process without respiking it each additional step, correct?

DR. SOMERVILLE: That\'s right. Yes, that\'s correct.

DR. PETTEWAY: Okay.

DR. SOMERVILLE: I think the experiments that Dr. Rohwer will describe
are looking at process of the final steps in the process with spiking at
the beginning of those individual steps.

DR. PETTEWAY: Exactly. So that your final removal shows the cumulative
effect of the process to remove the input spike.

DR. SOMERVILLE: Yes, yes.

DR. PETTEWAY: I have one other question and that\'s with the magnitude
of the clearance numbers.

DR. SOMERVILLE: Yes.

DR. PETTEWAY: And the less than or equal to or greater than or equal to.
The magnitude reflects the limit of detection of the assay.

DR. SOMERVILLE: Precisely.

DR. PETTEWAY: As opposed to what may actually be the magnitude of
removal. The magnitude of removal is likely to be much greater than the
numbers reflect, because of the limit of detection of the assay, right?

DR. SOMERVILLE: Basically, yes. We can only report what we observe.

DR. PETTEWAY: Right.

DR. SOMERVILLE: But we can also make some predictions about what we know
from other parts of the process.

DR. PETTEWAY: Right.

DR. SOMERVILLE: And that is why, as you\'ve said it before, it\'s
important not only to look at the overall process, but to look at
individual steps and evaluate what they may be contributing to the
inactivation process or removal process. However, as the study
illustrated, for example, we also have to be aware that individual steps
will not be additive and that one part of the process may remove the
same thing as a later part, later stage might also remove, so you have
to be very careful when you\'re doing that.

DR. PETTEWAY: But we can be confident in the linking of these studies
that based on the input spike that there was no detectable infectivity
based on the limited detection of the assay at the end of the process?

DR. SOMERVILLE: Yes, yes.

DR. PETTEWAY: And then the last question I have is the additional step
with the sodium hydroxide. That was evaluated independently?

DR. SOMERVILLE: What? It was a separate experiment, if that\'s what you
mean.

DR. PETTEWAY: Yes, that was a separate experiment?

DR. SOMERVILLE: Yes.

DR. PETTEWAY: Evaluated independently. Okay.

DR. SOMERVILLE: Right.

DR. PETTEWAY: Thanks.

CHAIR PRIOLA: Dr. Hogan?

BOARD MEMBER HOGAN: Very nice studies, Bob. I had a question on when you
are calculating the clearance factor here, you\'ve listed an equation
that says clearance factor is equal to gram spike times 10 the log titre
spike divided by milliliters of gelatin times correct factor times 10 to
the log titre reduction or gelatin. For somebody that can\'t balance
their checkbook, what do you mean by correction factor in the
denominator and why was that entered?

DR. SOMERVILLE: Okay. The correction factors are to account for the
inherent losses in the process by taking a sample out for intermediate
titration or other evaluations. So there is natural loss in the amounts
going through the process. Does that deal with that?

BOARD MEMBER HOGAN: Yes, that\'s great and it makes good sense. The
second question is did you look at any place in the process where titre
might have accumulated or concentrated, such as inner vessels or on any
of the columns or anything like that?

DR. SOMERVILLE: I think the short answer is no. Unless Dr. Grobben would
like to comment on that. But as far as I\'m aware, there was no specific
attempt to evaluate that.

MR. GROBBEN: I do want to. I would like to comment to that, I think. No
attempt was done to try to measure the infectivity which remains in the
equipment, especially for filtration and ion-exchange, because of the
problem to extract that infectivity from that equipment, so that was not
done. We just measured what was left in the gelatin.

CHAIR PRIOLA: Go ahead.

BOARD MEMBER HOGAN: That\'s what I presumed. It\'s just very difficult
to get that stuff off to measure it regardless. Now, am I to understand
that in the gelatin processing process that these filters would be
reused batch after batch or are new filters introduced in the
manufacturing process either in Europe or the United States?

MR. SCHRIEBER: May I answer this. There is no reuse. It\'s a one time
use. It may be the answer as well as with the ion-exchange columns. They
are regenerated with either alkaline or assay to purify for the next
round of ion-exchange. So there is a constant chemical treatment after
the gelatin has passed those columns.

CHAIR PRIOLA: Okay. If there are no further questions, we\'ll move on.
Thank you very much, Dr. Somerville, and Dr. Rohwer is going to present
some data.

DR. ROHWER: Can we go to the next slide, the first slide here or do I
control it? Are you controlling it or am I? Where is it here? Oh. Yes,
please, go to the first slide. Thanks. The gelatin manufacturing process
is a diverse one. It has many generic features like the contractionation
for plasma, at least I see it that way having worked in both areas. So
they needed a protocol representing as much of their collective
production as possible. And the steps that we were asked to validate in
our laboratory were on bone gelatin.

Next. And we used the process parameters that were selected by GME,
their scale down and this took a lot of time setting this up. Ad Grobben
deserves a lot of credit for this, as was mentioned. And from our end,
our major concern was about hazard control, and we spent quite a bit of
time on this as well. We did this study at a scale that was much larger
than we typically use in the laboratory. We were using meters and liters
instead of 100 mls at a time and some of these steps did not fit easily
into the valid safety cabinets and that type of thing, so we had to
figure out other ways to do them. But in the end, we were successful and
it worked without a hitch when we finally got down to doing it.

Next. The filtrations, the way this was done is we tested several
different types of filtrations that are used across the industry and
then pulled the filtrates, and that\'s what was actually titred. I\'ll
show you that in a moment. And getting them all done though it took
quite a lot of time, because of the scale and the precautions we had to
take to do it safely.

Also, all of the work that is done with gelatin has this complication
that it is only liquid above 50 degrees centigrade, so you have to keep
things warm. You have to keep them warm on a large scale, and so we
developed a lot of technologies for doing that, which the tempering
beaker turned out to be one of our best tools, but circulating baths and
hot pads were also useful.

Next. Next, please. About hazard control, we used safety cabinets, bags
to cover everything up during the actual processes. All joints between
chromatography column unions and filtration things were \-- transfers
were covered with plastic sleeves in case they leaked, put things in
large pots when we could. We poured nothing. Everything was done by
pumping from one vessel to another in a safety cabinet.

Next. We are also concerned about cross-contamination simply because of
the scale that we were doing this on and also because of the sensitivity
of the results. And as a consequence, all new dedicated equipment was
used for these steps. Everything was disposed \-- most things were
disposable. The only things that weren\'t were the stainless steel
filtration vessels and a couple of other things which could be
autoclaved under sodium hydroxide for reuse.

Next. Next, please. We had a question about \-- some discussion about
the spike earlier, and I think this was a very gratifying experiment for
me. We\'ve been trying to figure out whether our spikes are relevant in
our plasma studies and that type of thing. But in the case of bone
gelatin, the most likely source of infectivity is CNS tissue. And as a
consequence, in this particular case, at least we can say that the brain
derived spike is probably the most appropriate spike for testing removal
from this type of study.

And personally, I think this is the relevant tissue and we can use it
with confidence. There are issues about whether 263K or the less adapted
BSE strain is more relevant. My feeling is that there are advantages to
both. Actually, clinically, hamster 263K looks a lot more like BSE than
the less adaptive 301V strain. On the other hand, this is a strain that
was devised from BSE and so we use that as well. The important thing is
when you do two different strains, is what you\'re looking for is the
point of convergence between the two to give you some confidence that
the result you are getting are generalized more.

Next. The continuous process was done at the Institute for Animal
Health, and we are only working on the end stage process right here.
Robert has discussed the rest of this. The continuity was maintained by
Ad Grobben, who took copious notes, and we also have a lot of further
documentation, which I\'ll share some of that with you in a moment.

Next. So here is the process we have been looking at. This is the part
that Robert has been describing right here. Well, actually, they carried
it through this stage as well, but the only part of this process that
we\'re going to be dealing with is this part right here at the bottom.
The so-called purification steps, the filtration, ion-exchange and UHT
sterilization, and we\'re going to look at step wise removals.

Actually, we\'re going to gang these two together in one experiment.
We\'re going to look at them independently as well, and we\'re going to
compare the cumulative versus the serial with the individual testing of
these two steps. This was done just individually.

Next. Here is the basic layout of these experiments. We have the
infectivity spike. It goes into the crude gelatin, which is taken
directly from production at the same stage of production. It\'s passed
through the filtration device. And in the filtration experiments, on one
arm, we took the filtrates and took it straight through the ion-exchange
columns and then titrated it. On another arm, we took it over here and
respiked it, figuring that we may have \-- hopefully, we had removed
something in the filtration.

This spike, at the most, would only double the titre that we started out
with here by respiking. If we got any kind of removal here, we\'re just
starting over at this point. And then testing the ion-exchange by
itself. We wanted to use this so that we had filtrated material to test
the ion-exchange process with.

Next. The filtration steps, in fact, involved five different protocols
with various compositions of cellulose, sources of cellulose and
formulations depending on different manufacturing setups across the
industry. The filtrates from those were all pooled and then they were
titrated by themselves before being passed to the ion-exchange column
over here or respiked on this arm and passed through the ion-exchange
columns here. The ion-exchange consisted of two columns, the cation
exchange followed by an anion exchange, and what we assayed was the
eluate from both, the final eluate from both.

Next. And then in terms of the UHT sterilization, the ultra high
temperature sterilization, we again started with gelatin from
production, infected that, spiked that with infectivity and then did the
UHT test and titred that. So it\'s a much simpler pathway.

Next. Next, please. Here is a picture of Ad Grobben setting up the
filtration experiments. This is the filter apparatus over here. We\'re
transferring, I believe at this step, we\'re preheating the filter with
hot water that has been heated over here, and I\'m not sure that\'s what
we\'re doing there, but that\'s what this is doing. This is the hot
water. It had to be preheated so that it was warm enough to keep the
gelatin melted once we put it in there. We\'ve got another bath heating
up the gelatin to dry through the filter.

Next. Here\'s the filter being assembled. It\'s quite a large apparatus,
compared to what we\'re used to, but we were able to do all of this
within the hood, though the transfers had to be through this pump on the
outside. There\'s the filter A being added.

Next. Here is the filter A being stirred in the filter and then it\'s
drained to form the filter cake in the bottom.

Next. Here is the filtration apparatus setup being done. Here is the
vent in case, because you have to vent some air out of it in the early
stages, and through HEPA filter here. And here is the assembly after the
filtration is over.

Next. This was a keeper in that experiment, and this was a failure. We
always inspected the filter cakes after the filtrations to make sure
they were intact. There were no possibilities of leaks and that kind of
thing before we would keep the filtration as a successful one. And so
the only thing that goes into this study were successful filtrations.

Next. Here is a picture of the column apparatus, the ion-exchange
apparatus. These were gigantic by our standards. We set them up on a
mobile cart on a chromatography rack that we can roll, so that once we
got everything setup and ready to go, we could roll the cart over a very
large plastic bag and then cover the whole thing in this plastic bag and
seal it up during the actual experiment in case there were leaks. Thank
God there were none.

Next. And here is the apparatus that we used for the UHT Inactivation
Experiment, and I can make that a little clearer in the next slide,
which is diagrammatic.

Next. The principles that we are trying to employ in the UHT study that
we were trying to mimic from the actual production environment is, from
my prospective in studies that I\'ll talk about later this afternoon,
infectivity is not intrinsically resistent. The problem is delivery of
the inactivant and the inactivant finding sanctuaries to hide from the
steam, and drying is \-- drying into a film is one of the biggest
problems.

And one of the nice features of the UHT process is the gelatin is being
pumped through a pipe in which live steam is being injected. There is no
head space. There are no sanctuaries. There is no place for this stuff
to dry. There is no place for it to escape from the hydrolytic
environment. We wanted to duplicate that as best as we could.

Next. Next, please. So we did that with this apparatus where we filled
this stainless steel capillary and we used this capillary so that we
could affect a very rapid heating and cooling, because the whole
process, the UHT process, is a 4 second exposure to 140 degrees
centigrade. So how do you do that in 4 seconds? Well, you have to get
the heat to it in a hurry. We didn\'t try to do it dynamically. We did
it statically. But we did it in this way.

So we have this chromatography capillary here. We have a thermocouple,
which is embedded in the tube. The probe is right about here. We have
another thermocouple on the outside to track what is happening in the
bath. And then to relieve any over pressure in the device, we have a
water column going here to a back pressure gauge, which ultimately if it
were to leak, it would go into this tube right here. And this relieves
the hydrostatic pressure that is developed by the fact that we\'re
heating this gelatin up in here. But we\'ve got within the gelatin
column itself, we have no head space.

Next. We\'ll take that and the way we get our rapid heating is through
trial and error. We set up a protocol where we can dip this thing,
hooked to its various thermocouples, up to this recording device into
our 160 degree oil bath and then as we see the temperature hit our
target transfer temperature, which was about 80 degrees, we quickly dump
it into the 140 degree bath and it comes to equilibrium in the
successive period. We then take it from there after 4 seconds has
elapsed and dump it into the other temperature. We\'re tracking this
whole thing on the computer. We\'re watching it in real time as we\'re
doing it.

Next. So we get curves like this. This is seconds down here. This is
degrees over here. This is the outside of the capillary tube. So we\'re
seeing the capillary dipped into the 160 degree bath, and here we\'re
seeing the transfer into the 140 degree bath, and we\'re seeing it come
to \-- and this is the internal thermocouple and it is coming to
temperature very quickly thereafter, and then at 4 seconds we plunge it
into the water bath and that\'s the way there.

Now, what I showed you first was the hamster experiment. We have now
advanced to the mouse experiment. And there is one important difference.
As we got more and more experienced with this, we were able to get this
ramp time down to shorter and shorter periods. We had about 4 seconds on
the first one and about 1 second here. We did four or five trials,
actually, three in the end we focused on once we got the method working.
And then we picked the best of those trials. And what I have shown you
is the temperature records for the two best trials for mouse and hamster
and that\'s what got titrated.

Next. Next, please. This is the results of all these experiments. The
pooled filtrates gave a very disappointing clearance. I was expecting
much higher than that. The respiked column gave this, only about a half
log removal, and remember for these types of input titrations, we got
about a .3 log error associated with these numbers. The successive
filtration and ion-exchange gave about 1.8, which very interestingly,
but probably somewhat randomly, is exactly the same as the added values
between these two.

But I think it gives us some confidence that putting these things
together, even though the removal at each step is low, we are getting
some significant removal here of about 1.5 to 2 logs. The UHT
sterilization by comparison gave a much better result. Even that 4
second exposure is giving us about 4 logs of removal. Attached to this,
we have about 6 logs cumulative. And I think it is legitimate to attach
these, because these are quite different methods of removal versus
inactivation.

Next. And this is just a comment on that, these things were showing
independent removal to the extent that we could detect it with the lower
levels that we saw there. But in terms of looking at the total, at what
was actually going on there, the serial experiment is actually the best
one to use, and that\'s the one I think we should focus on. But both of
these steps were much less effective than I had expected. And I don\'t
know whether it is because of the matrix, the apparatus, the gelatin
itself, but in the next slide I\'ll show you some data.

Next, please. In our experience, this is about half these experiments
were done by us. The other half were done by other folks, but they were
compiled for a former presentation of the FDA and a TSE Advisory
Committee meeting in October 7, 1997. And typically, and we\'ve done
more of these since then, especially these depth filtrations, and they
are typically removing 3 logs or better.

So there\'s something different about gelatin. And it\'s either the way
we did the experiment or it could be that the gelatin is so overwhelming
in terms of a competitive binder for the matrix that we\'re not getting
removal because of that. But anyway, it doesn\'t fall into expectation.
There is a warning in this though, which says that you have to check
these things. You can\'t extrapolate from this cumulative experience and
presume that it\'s going to work in all cases.

Next. I\'ve already dealt with this. Let\'s go on. Next, please. I just
want to point out that this UHT result is the worst case result. We did
it under static conditions. It is heated from the outside instead of the
inside. 4 seconds is a minimum exposure that is seen in the industry.
And we\'re using crude brain homogenate instead of material that has
been already refined by the process. And my guess is that the stuff that
has been through the process may be even more susceptible, but that\'s a
guess.

Next. I want to make one final point and that is that the total exposure
that these samples got really begins, at least for sure, with the 263K
case, with the 80 degree exposure. I mean, somewhere between 80 and 100
degrees. We have a series of experiments which are actually on the next
slide that I did in the \'80s showing that we start to see affect around
100 degrees, and we get total killing in a few seconds at 121.

So this ramp temperature is also contributing to the inactivation here.
And if we take these ramp temperatures and add it from 100, the ramp
exposure plus the exposure temperature for the 263K case and the 301V
case and plot them on the same curve, which I\'m going to do next.

Next, please. This is just showing you that there is an effect at 100
degrees and above for the 263K case, at least.

Next. These are the ramp times for those former experiments.

Next. Let\'s go on. Next. I just want to show you this last slide. If
you plot the data from the 301V case and the 263K case on the same time
axis down here, including these earlier studies out here which were done
at 121 versus 140, and plot it back to the origin, you get a straight
line through these things. Well, I first draw the line through them. And
what that is telling me is two things.

One, there isn\'t really any significant difference between the
sensitivity of these two agents to this process. And two, it gives me
some confidence in saying that if you are to extend this process to 10
or 12 minutes, you get another 4 logs or so removal. This is something
that should be checked with actual kinetic experiment and kinetic
measurements, but it seems to me that this being a minimum is a very \--
this 4 seconds being a minimum exposure is a very encouraging feature of
this experiment.

Next. In conclusion, the purification steps are removing 4 to 6 logs and
the UHT step, in particular, provides a potentially very secure
inactivation step. Thank you.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: The next to the last slide you showed the
susceptibility to heat over time, and what you had you mentioned the
straight line fit, but I didn\'t see any intermediate points there that
could really detect curvature in the line.

DR. ROHWER: No, there isn\'t. What I\'m saying is we\'re working with
the data that I have. And I think I also said at the same time that it
would be very nice to do a complete kinetic study on this.

BOARD MEMBER BAILAR: Yes, but I would not conclude from that that it\'s
a straight line.

DR. ROHWER: Oh, I see what you\'re saying. It may not be. You\'re right.
From here to here, well, from here to here, extrapolation, I don\'t
know. I mean, it\'s hard. I guess, what would you say? You could have
something like that, I guess.

BOARD MEMBER BAILAR: I would say you do not have the evidence on which
to detect whether there is any curvature.

DR. ROHWER: Okay. Well, I\'ll grant you that. And all I\'m saying is
that this is \-- let\'s put it this way, this data is consistent with a
first order process here, with these two samples behaving very, very
similarly.

BOARD MEMBER BAILAR: Okay. That\'s all.

CHAIR PRIOLA: Dr. Petteway?

DR. PETTEWAY: That\'s a very impressive set of experiments, Bob,
especially dealing with the scale down, handling it all. That\'s an
extremely difficult thing to do. But the 4.2 logs, was that the
magnitude with some residual infectivity found?

DR. ROHWER: Oh, yes.

DR. PETTEWAY: Okay.

DR. ROHWER: Yes, I mean, we started with 7.5 logs.

DR. PETTEWAY: Yes.

DR. ROHWER: So there\'s still 3 or 4 logs left.

DR. PETTEWAY: And that was at 4 seconds which is worst case?

DR. ROHWER: Yes.

DR. PETTEWAY: And what you\'re saying, I mean, even given other points
that would show a change in that curve, the likelihood is 8, 10, 12
seconds, there would be nothing left is the point?

DR. ROHWER: I was very interested this morning when Michael Dunn pointed
out in his presentation that in North America anyway the typical time is
8 to 16 seconds, as opposed to 4 seconds, and apparently gelatin can
tolerate that quite well. If you would like to say something about that?
Well, that\'s up to Sue. Sorry.

CHAIR PRIOLA: Dr. Dunn, do you want to comment on that?

DR. DUNN: Could you say it again?

DR. ROHWER: Yes, if I could repeat that, what I just heard here is that
there is apparently no problem extending that time for 8 to 16 seconds.

CHAIR PRIOLA: And Dr. Hogan?

BOARD MEMBER HOGAN: Well, the question is why does the European process
use 4 seconds and is there a ramp up time to that or is it just the
batch is brought in, zap 4 seconds and then it is taken out?

DR. ROHWER: I would like to defer to Mr. Schrieber, if I could.

CHAIR PRIOLA: Yes, Mr. Schrieber?

MR. SCHRIEBER: What I explained in my presentation already is that we
used the softest condition we have found in one of the gelatin plants in
Europe. So it\'s not uncommon to have like in the States a longer
temperature or even a somewhat higher, longer time or even somewhat
higher temperature, but we had to choose the minimum conditions founded
in the three or four studies, and that\'s what it is. You are right if
the time would be expanded to 6 seconds or the temperature would go up
to 140 instead of 138, this would not really harm the quality of the
gelatin.

CHAIR PRIOLA: All right. If there are no other questions, thank you very
much, Bob.

(Applause)

CHAIR PRIOLA: I would just like to say having gone through the bulk of
this, these infectivity studies in our rather thick handout, that it is
very impressive the work that Drs. Taylor, Somerville, Rohwer, Ad
Grobben and Schrieber have done studying inactivation of TSE infectivity
through the gelatin processes. It\'s a lot of real nice work.

I would like now to ask Dr. Morris to come up and explain to us the
USDA\'s gelatin policy.

DR. MORRIS: Okay. Good morning and thank you for the opportunity to
speak with your Committee regarding APHIS\'s policies regarding the
importation of gelatin.

COURT REPORTER: Dr. Morris, hit the volume button.

DR. MORRIS: Thank you.

CHAIR PRIOLA: I\'m sorry. My apologies. It was supposed to be Dr.
Rogers. I\'m very sorry. That\'s my error. Can we just go with that?

DR. ROGERS: Yes.

CHAIR PRIOLA: Okay. My apologies. I\'m sorry. You should have told me.
I\'m misaligned in the agenda. Okay. So, in fact, we\'re not going to
hear from Dr. Morris yet. It\'s Dr. Rogers who is going to give us a
risk analysis of infectivity.

DR. ROGERS: Well, I guess the slide has disappeared for a minute there,
so don\'t start the thing until \-- the timer until it shows up. Is the
mike on? Okay. Thanks for inviting us down here today from Canada. I\'m
from Health Canada.

CHAIR PRIOLA: We can\'t hear.

DR. ROGERS: So is the mike on? Oh, it\'s on now? Oh, closer. Taller and
closer. How\'s that? Okay. At Health Canada we have been doing a number
of quantitative risk assessments and part of our topic today that we
will be covering is what\'s on your agenda. But I did want to say that
what we\'re really looking at is the varying-CJD risk to consumers
eating foods containing small amounts of processed ruminant products.
And I want to talk about some of our modeling functions.

Next slide, please. We have just completed a quantitative risk
assessment for basically products that contain beef extracts and the
beef extract industry certainly has a lot of similarities to the gelatin
industry, so some of the information I\'m going to provide today will
certainly with some understanding of the overall picture. I do want to
present today like the quantitative model parameters for the evaluation
on pairing CJD risks. I want to focus on the front end parameters for
risk analysis, and I want to provide some information on evaluating
uncertainty in the parameters and provide information on variability
that we\'re using in our models.

Next slide, please. The purpose of our risk assessments are really to
provide information on two risk outcomes of the probability of
individuals acquiring varying-CJD through the consumption of a product
and the annual number of varying-CJD infections that could be predicted.

Next slide, please. The approach that we\'re using, basically, the first
thing that we look at is the length from BSE agent to varying-CJD. To
date, there is no direct evidence linking the acquiring of varying-CJD
to particular products. And I want to emphasize that certainly the only
information we had previously was the work of Simon Cozens of the UK for
food products which had some implicated meat pies, sausages, these types
of things in his work, but he has, in fact, recalculated his and
reevaluated some of that publications and in Edinburgh last year he has,
in fact, shown there has been no statistical significance to particular
food products and varying-CJD. So that\'s an important picture.

The presence of the BSE agent in the product of concern are not
measurable by our current techniques. The only thing that we can
actually still seem to have some type of laboratory analysis for is the
presence of CNS materials through IHCGFP and some neuro analyst
techniques. The hazard identification basically has established that
there is a route from BSE to consumption exists. And so that\'s the
reason for the presentation of the modeling.

Next, please. In Canada, we are using the model that has basically been
setup by the Kodak element. We have an issued statement. We do hazard
identification, hazard characterization, exposure assessment and risk
characterization, but nothing goes forward until you have hazard
identification.

Next slide, please. Our structure in our risk characterization is
depicted here. Basically, we are looking for these probability
statements in the middle, which are outcomes, but we are looking at the
infectivity consumed, which really comes through our exposure
assessments and the consumption frequencies from the exposure
assessment, and then the dose response models that we have been
developing, which are in the hazard characterization.

Next slide, please. Our structure in hazard characterization, basically,
the main things, variables that we would be looking at are the
susceptibility in human population. We can say that certainly the we in
our risk assessment are looking at worst case assumptions. In fact, with
the human population, we are not looking at divergence, for instance,
because of met type of codons, we say that all humans are susceptible.
We\'re not looking at immuno-compromised or younger children. There\'s
no infant instances of that, so like our population characteristics say
that all populations are susceptible.

Infectivity accumulation is one of the things we are looking at
particularly with dose response type of modeling. Our species barrier
from bovines to humans, I guess, what I would say there again is that we
are looking at the worst case. We\'re saying that there is no species
barriers. It\'s a 1 to 1 ratio, but certainly when we\'re looking at the
advice that we get from the Scientific Steering Committee over in Europe
that they say that we should use the range of 1 to 10,000.

And, in fact, the latest publications do say for oral transmission to
food products, you probably should be looking at 10,000. And we are
looking at risk assessments to be tried and say, for instance, they do
want things to be practical and realistic, but then we are going with
due caution. And so some of these products that we\'re looking at to
start with, we are emphasizing generally worst case assumptions and
seeing what those numbers generated look like, and so that\'s what we
have been doing.

Next slide, please. In particular, it\'s the dose response area for
varying-CJD. And in our models we are using a threshold dose response as
well as an accumulation dose response. I have a lot of slides here, so
I\'ll have to hurry along with this. We\'ll talk about that a little
further along.

Next slide, please. The structure and our exposure assessment just
basically analyze the model that we use.

Next slide, please. And particularly, which is of interest to this
particular audience is the fact that we are looking for the presence of
BSE in cattle populations. Our models are setup in such a way that we do
know that the disease status of a country changes and I think we have
that from our own experience, but we have them working along the lines,
for instance, that food products and gelatin products are produced over
periods of time, so the BSE status and the amount of BSE possibly
infected cattle in the country change, and so we really want to be able
to adapt that to the different lots and processes.

The tissue infectivity information that we use, basically, a lot of it
from the oral pathogenesis studies to start with from Dr. Wells group
and that continues on in the UK. Our source of infectivity in the
slaughtering areas, our sources of infectivity certainly depend on the
tissues that are used in the products, and I think you\'ve been
discussing a lot of those today with specified risk materials, for
instance.

And this area here I put the word gelatin for the commercial product of
what we\'re looking at. In fact, this model was developed particularly
for beef extracts, but it is the front end that I wanted to talk about
today. Because what we\'ve been doing as well as we do look at
consumption, the actual final products, the amount of material that are
in final products, and then the amount that go to consumers and consumer
individual servings.

Next slide, please. So our quantitative model prevalence of BSE into \--
the BSE infected bovines in populations in the screening procedures. The
inference from the countries of BSE surveillance, first of all, I want
to say one thing and that is that the products we were looking at were
generally ones that were coming from the European Union or could have
been coming from other places. But when we were looking at prevalence
data itself, because of the wide enhanced surveillance targets that have
been going on in the EU that we particular have some good observational
data there to work backwards from.

And I must say that certainly the EU has also been doing a lot of
missions out to their member states to go for audit and compliance, and
they have been doing a number of good reports on that. That\'s why we
get some excellent data to sort of give some parameters around to put in
models. However, I\'ll tell you that particularly our concerns are
detected diagnosed cases are removed from \-- are diverted from food
chains.

But the incubating cattle are a question in the amount of infectivity in
incubating cattle, certainly one of the major things that we have been
struggling with. But in our particular assessments, we\'re using 4
incubating cattle per adult cattle diagnosed and we\'ve done that,
basically, from talking to experts in Europe. And the other thing that
we\'re doing, though, is that we\'re talking about the infectivity of
tissues.

We started off with giving them exactly the same infectivity as the
clinical animals to run through the worst case numbers, and then we
scaled that backwards. But I\'ll explain a little bit later. But what
we\'ve done is basically we try to group countries into low, medium and
high prevalence rates and so then I do have numbers on that, but again
this presentation is going to be a little small for that, but we\'ll get
to that.

As far as the abattoir screening itself it\'s concerned that certainly
now there are rapid tests involved, and we\'ve seen that they were
talking about 100 percent sensitivity, 100 percent specificity. There\'s
a number of rapid tests out there. We have done an evaluation of them
and we\'ve used the worst case sensitivity for one particular rapid
test, because we cannot tell because there are varied tests that have
been used in similar countries and so you don\'t necessarily have all
that information.

But for modeling purposes, again, we\'re using the worst case. And for
the ante-mortem, postmortem inspections only for diverting BSE infected
cattle from the food chain, we\'re using a 2.5 percent removal from
ante-mortem. And I think most people know how very difficult it is to
diagnose TSE diseases and that they are very complicated.

However, they have gone back and it is a requirement in the European
Union to, in fact, state where you are during your diagnoses, are they
ante-mortem or are they rapid test. And so that the range of variability
in ante-mortem depends on the country and in the awareness, the
education and in a lot of infrastructure elements. However, I can tell
you that for what we\'ve done, I\'ve only seen six reports so far from
mission compliance audit states and the lowest amount of divergence is 3
percent out of Belgium from ante-mortem and up to in the high 30
percents in Germany. So there\'s a lot of variability in the amount of
BSE infected animals that are removed postmortem. And this goes into the
models as well.

Next, please. Next, please. Oh, sorry, I couldn\'t see it right. Okay.
So now, I\'m looking at tissue infectivity. I just want to give a brief
run down here that we are using .1 gram of raw unprocessed brain tissue
from a clinically infected bovine as the minimal or as the threshold
dose in our models, at this time. I think that most of you are aware
that that is the amount of unprocessed tissue now that has been orally
given to a cow that has come down with BSE in the UK in the latest
pathogenesis studies. That animal was 52 months. Again, like a very low
dose, but it is our starting point.

However, we do put uncertainty around these things, again, up to, per
program, 10^1^ to 10^3^ infectious doses. And then the infectivity that
we are assumingly using the same infectivity scaling standardization to
the trigeminal root ganglia, the dorsal root ganglia, the spinal cord
and emboli could possibly go into this slaughtering and stunning
procedures. And I have mentioned already we are looking at the
incubating bovine, and particularly the sensitivity issue around that,
and so we have a scale at different levels in our final results.

Next, please. Our sources of infectivity, particularly, when we are
looking at raw materials, it could be going into things like gelatin.
Our CNS emboli in the blood, possibly spinal column
cross-contaminations, blood itself, edible fat contaminations, bone
marrow, spinal column, and trigeminal ganglia.

Next, please. And, in fact, this is the way that we have started in our
beef extract risk assessment that we sort of look at in terms of tissue
restrictions and no tissue restrictions, and particularly, although I
guess this was the top line here that would be very much parallel to
what could, in fact, be going on in the gelatin industry, because in
beef extract, we do have some productions that only use muscles only.

Next, please. One of the things that we see are really the number of
bovines that are, in fact, going into batches and lot production and
silo storage in beef extract and this is very similar to some of the
things you are seeing in the gelatin production. So that we do have, in
fact, calculated the probability of a batch contamination, lot
contaminations by the prevalence rates and by \-- well, it\'s a little
complicated here for me just to go over that quickly, but it\'s those
calculations that we were looking for to say that there\'s a probability
that the consumer product is made from a contaminated product at that
end.

And so we are looking at the number of infected bovine tissues that go
into the batch or lots based on those country prevalence ratios as well.
We also are looking at the infectivity reductions, because in beef
extract production, as well, you get a lot of heating, wet heat,
filtering, decanting and denaturation and, in fact, we have tried to mix
estimations on the log reductions there.

Next, please. In terms of defining our concerns and characteristics for
these products, and I suppose this is one of the difficulties that we do
have with food products that contain small amounts of a ruminant product
ingredient, not always on the labels and not always necessarily going to
the ingredients. In the beef extract production business, for instance,
there is no GME type organization.

We, in fact, had to go to every country, major country, around the world
that does have beef extract production and do our individual
investigations by companies to find out the capacity of their equipment,
the number of animals, for instance, that \-- first of all, one animal
contributes so much tissue per lot, and so there is a range of animals
that go into lots or batches. And so the probabilities are all derived
from that type of information.

And so like that\'s something I\'m \-- actually, I skipped over that a
little bit, but it\'s very important for this type of estimation of, for
instance, that even if you did have a BSE infected cow going to gelatin
production, for instance, you have to know the capacities of the
equipment and the type of equipment and the different processes and
certainly that the gelatin industry have indicated that there basically
are very similar processes, a little different in the other areas.

I see I\'m at stop time already, but I just need to go to the next
slide, please. Basically, these were the variabilities and the
components that we\'ve been looking at, and I\'m probably way over time
here. But can I just kind of continue on just real quick? Okay? Because
certainly like our particular interests is really in the production
methods themselves, the production practices, the sources of
infectivity, all of these that we\'ve quite clearly documented in our
written reports, which we would be glad to share with this Committee at
a later time.

And the consumer product characteristics themselves, because there is
differences amongst the groups of products and within the groups of
products. And so we have gone through actual analysis of the amounts of
materials that go in there, and then the consumption characteristics
themselves, because each product has a different consumption
characteristic and so we\'ve tried to work that through with a point
estimate of the maximum values.

Next, please. And the uncertainty issues that we really weren\'t looking
at in our reports and reporting them as sensitivity, you really have to
do a tissue infectivity incubating bovines or species bearing the dose
response.

Next, please. And if you want to \-- next, please. Because this is
basically the charts that we prepare, and we are providing like product
groups within our report.

Next, please. The BSE prevalence is basically put into our charts.

Next. Our abattoir screening techniques.

Next. For divergence of BSE animals.

Next. Other production methods that we have been going through with all
information we collect and we can provide that as a range.

Next. And then we\'ve done production parameters which are a range of
ranges depending on the type of processing, etcetera, and the types of
tissues that are added.

Next, please. And I think I\'ll skip this one right now. Next, please.
Next. And this basically is just giving us some information on if
you\'re using rapid test and ante-mortem tests.

Next, please. Because it was this type of scatter diagram that we\'re
trying to provide to sort of show or give the information on the
probability of the consumer batches themselves. If you\'ve got large
batches, small batches, high prevalence, low prevalence, and so we tried
to do some diagrammatic information in our reports to give some idea of
the dilution of tissues with no infectivity or with infectivity.

Next, please. And the difficulties I have talked to, and next, please.
Bottom line is that these are the individual outcomes that we have been
trying to quantify in our risk assessments, and particularly, though
that we \-- you will find that you can have a lot of problems with
surveys, nutrition surveys, etcetera, for the types of details that you
would be looking for for trying to get estimates on consumption values.

I\'m sorry to have gone over.

CHAIR PRIOLA: Okay. Well, thank you. Are there any questions for Dr.
Rogers? Yes?

BOARD MEMBER WOLFE: You mentioned earlier on that your assumption of the
ratio of incubating cows to infectiveness is 4 to 1. What is the basis
for that?

DR. ROGERS: Excuse me, well, that\'s basically expert opinion from
Europe, because like we had talked to people that had the experience
with BSE for a number of years, and so that that is just strictly an
expert opinion. There\'s no rationale for that except there is a range
of incubating cattle that we do use, but I can tell you the reason why
we\'re using 4, at this time, is because we have implied such harsh
assumptions to the fact that there is, it has the same amount of
infectivity as the clinical animal.

BOARD MEMBER WOLFE: Can you tell me what the range is that you said that
you use it for? What is the range or ratio?

DR. ROGERS: 4 to 10.

BOARD MEMBER WOLFE: 4 to 10. Thank you.

CHAIR PRIOLA: Okay. Thank you again, Dr. Rogers. After carefully
checking my agenda, now, we\'re going to hear from Dr. Morris from the
USDA. I apologize again for putting you on the spot earlier.

DR. MORRIS: Good morning. Thank you again for the opportunity to share
our Agency\'s policy on gelatin. Next slide, please.

I am Dr. Terry Morris with the National Center for Import/Export. I am
representing the United States Department of Agriculture, Animal Implant
Health Inspection Service, Veterinary Services.

Next slide, please. We are headquartered out of Riverdale, Maryland.

Next slide. And we are under the direction of Dr. Karen James-Preston.

Next slide, please. Title 9, Code of Federal Regulations, Part 94, 95,
121 and 122 gives APHIS the authority to regulate animal products. Part
94 gives us the authority to regulate gelatin.

Next slide, please. We regulate gelatin based on the presence or absence
of BSE and the association with the BSE affected region, BSE being
Bovine Spongiform Encephalopathy. We have lumped gelatin into one of
three categories. One category would be gelatin that is derived from
non-ruminant species. A second category would include ruminant gelatin
that is derived from cattle that have no association with a BSE affected
region. And the third category is ruminant gelatin that has been derived
and has an association with a BSE affected region. For the gelatin that
has an association with a BSE affected region, those regulations are
found in Part 94, Section 18(c).

Next slide, please. And pretty much to summarize, 94.18(c), the gelatin
that is derived from ruminants and the ruminants are from a BSE affected
region, that gelatin is prohibited entry, unless the gelatin is imported
for human food purposes, pharmaceutical products, photography or any
other similar uses that would not result in the gelatin coming in
contact with ruminants in the United States.

Next slide, please. 94.19 addresses gelatin derived from non-ruminant
species. This would include your pig, horse, poultry and fish gelatin.
On May 27, 2003, an interim rule was signed that modified the current
verbiage in 94.19.

Currently, next slide, please, the gelatin that is imported that is
derived from pigs, horses, birds and fish species must be accompanied by
an original official certificate endorsed by the full-time salaried
veterinarian responsible for animal health of the exporting country, and
it must state the animal species of origin.

Next slide, please. 94.19 also deals with gelatin derived from
ruminants, provided those ruminants have not been in a BSE affected
region.

Next slide, please. This part of the regulation requires that each
shipment should be accompanied by an official original certificate
endorsed by the full-time salaried veterinarian responsible for animal
health of the exporting government, and that certificate must state four
things. The first thing it must state the animal species from which the
material is derived. The second statement must include the region in
which the facility where the material was processed is located. The
third statement would include a statement that the material was derived
only from ruminants that have never been in a BSE affected region. And
the fourth statement must address dedicated facility conditions, meaning
the facility cannot receive, store or process any ruminant material from
any BSE affected region.

Next slide, please. The last category deals with ruminant gelatin that
has been associated with a BSE affected region.

Next slide, please. Ruminant gelatin that has been associated with a BSE
affected region must be accompanied by a veterinarian import permit. A
permit is a legal document that authorizes the importation of controlled
materials or organisms or vectors into the United States. For ruminant
gelatin associated with a BSE affected region, the permit would address
the country of origin. It would address the animal tissue species,
meaning hide or bone. It would address the exporting and the processing
country of origin. Again, we\'re looking at BSE-free versus BSE
affected.

Next slide, please. The next few slides depict scenarios that address
how APHIS would regulate the importation of ruminant material under
certain circumstances. In this scenario, the ruminant material whether
it be hide or bone is derived of ruminants from a BSE-free country, but
it is processed and exported in a BSE affected country. In this case, we
would issue a permit for this material.

The permit would require that the government certify the country of
origin of the raw animal materials and the government would also have to
certify specific conditions that exist within that facility and the BSE
affected region. Again, that facility would have to be a dedicated
facility, meaning it cannot store, receive or process any ruminant
material from any BSE affected region, with the exception of milk and
hides.

Next slide, please. In the second scenario, we address high derived
gelatin only, sourced from ruminants. In this case, whether the hide is
derived from ruminants from a BSE-free or a BSE affected region, the
fact that it is processed in a BSE affected region requires the need for
the permit. The permit, when issued, would require that the government
certify that the gelatin is hide derived only, and again because the
facility is in a BSE affected region, the facility would have to be
dedicated.

Next slide, please. The last scenario addresses bone derived gelatin.
For bone derived gelatin, and in this case, the ruminants are from a BSE
affected region. This material is allowed entry, provided the individual
obtains a permit. And when we issue the permit, the permit would require
that the individual maintain affidavits that they obtained from
individuals who they distribute this gelatin to. The affidavits would
require that the individual certifies that the material will not be used
as livestock feed ingredient.

The material cannot be incorporated into veterinary pharmaceutical uses
or the material cannot be incorporated into veterinary biologic
products. And this goes back to 94.18(c), which says that the material
can be imported, provided it is imported for human food, pharmaceutical
products and other uses, photography, and other uses that does not
result in the material being introduced to U.S. ruminants.

Next slide, please. To complete the process for obtaining an import
permit, you have to submit an application, which is VS form 16-3. It
takes about 2 to 3 weeks between the time that we can process the
application and turn around a permit to you. The application can be
submitted electronically through our website. It can be submitted by fax
or my mailing it into our office. The permit is good for one year, and
the permit will only allow for the specific commodity requested from the
specific exporters, and it would have to go to the importer that
requested the permit.

Next slide, please. This is my contact information, in the event that
you need to contact our office.

Next slide, please. And again, I wanted to thank the Committee for the
opportunity to share APHIS policies regarding the importation of
gelatin. I\'m happy to answer any questions you may have.

CHAIR PRIOLA: Dr. Wolfe?

BOARD MEMBER WOLFE: This is not meant to put you on the spot, but as you
know, the Department of Agriculture is seriously considering, we have
heard from others, on the verge of, lifting the ban on importation of
cattle, beef, from Canada to this country. You\'ve outlined a thoughtful
and, I think, careful permit process that affects gelatin, for instance,
which would come from a BSE affected region, such as Canada.

Do you not think that there is somewhat of a contradiction between being
so tight properly and restrictive about allowing gelatin to come in from
there, but seriously considering lifting the ban on meat from what would
be the first time the United States would ever have lifted a ban that
previously existed from a BSE affected country?

DR. MORRIS: Yes, sir, the APHIS TSE working group has devised a list of
low-risk commodities and a list of mitigation factors under which those
low-risk commodities can be imported, the specific criteria under which
we would consider accepting these low-risk commodities. That list has
been presented through channels to the White House and it is our
understanding that the White House has disseminated that list to the
trading partners, so it is in negotiation to make sure that all of our
trading partners are aware of what the potential actions would be.

BOARD MEMBER WOLFE: So you\'re saying that beef is presumably on a list
as a low-risk commodity? Is that what I interpreter you\'re saying?

DR. MORRIS: And I would have to look at the list, but it\'s specific
categories and it\'s specific ages. And, Lisa, if you want to help me
out here? Thank you.

BOARD MEMBER FERGUSON: Yes, I\'ll try and help you out. Speaking for the
Department, first of all, I would like to reiterate the point that you
probably shouldn\'t necessarily believe all the rumors that are in the
press and everything that you hear. There are lots of things under
consideration, not only at USDA, but through the entire Administration,
so all of the departments are contributing to these discussions.

And the discussions are centered around, you know, is there a
science-based way to look at the situation? Are there things that we can
do, that are based on known science to address the situation with
Canada? And as Terry has described, at least, we, at APHIS, have
provided some recommendations for certain products that perhaps could be
considered low-risk and could initially be allowed for import under
certain conditions.

I\'m not at liberty necessarily to say what is on that list, what might
not be on that list, but we have tried to address it. Okay. First of
all, products that are accepted internationally not to present a risk of
transmission, obviously, are not affected already. But we are looking at
a range of things to say these could be considered lower risk than other
things. And really it\'s a wide ranging list that goes over a lot of
issues.

BOARD MEMBER WOLFE: Thank you.

DR. MORRIS: Thank you.

CHAIR PRIOLA: Okay. Thank you very much, Dr. Morris. We\'ll now move on
to the open public hearing portion of the morning. So, Dr. Freas?

SECRETARY FREAS: As part of the Advisory Committee program, we hold open
public hearings, so that members of the public may wish to make comments
to the Advisory Committee will have the opportunity to do so. At this
time, I have received two requests. One is a written request. This
written request was run off for the Committee members, posted in the
viewing notebook out on the table and some copies were provided for the
public if you were here early.

The second request is from Mr. David Bieging and he is at the microphone
right now. Welcome.

MR. DWYER: Actually, I\'m Dan Dwyer. Dave Bieging made the request and
I\'m going to speak. I\'m Dan Dwyer. I represent the Gelatin
Manufacturers of Europe, and I\'m also speaking today on behalf of the
Gelatin Manufacturers Institute of America. You\'ve already heard from
representative of these two associations this morning. These
associations represent virtually all of the gelatin produced in Europe
and in the United States.

These two associations have been working for many years, as you know, to
ensure that gelatin is safe and we\'ve been pleased to be able to do so
in cooperation with the FDA. As we\'ve discussed with FDA previously, we
would like, at this time, to comment on the questions that FDA has asked
this Committee to address today.

Specifically, FDA\'s Question 1 currently reads \"Do the results of
these new studies demonstrate a reduction in infectivity that is
sufficient to protect human health?\" This question must be interpreted
in light of the normal circumstances surrounding gelatin production. In
particular, the question focuses only on the manufacturing processes
that were studied, but in practice, as you\'ve heard today, the safety
of gelatin is based on two principles.

The first principle is the use of raw materials. As you know, in Europe
this involves controls on raw materials imposed by the European Union
and by GME members. The second principle is the use of manufacturing
processes that can eliminate any potential infectivity that might
theoretically be present in the raw materials. In Europe, this involves
the use of the processes that you\'ve already heard discussed today and
that have been studied by GME.

These two principles of gelatin safety apply as well to all bone gelatin
regardless of geographic origin. Therefore, we request that when the
Committee considers FDA\'s Question 1 it take these two principles into
consideration, that is we would recommend that the question be revised
to read \"Based on the use of raw material sources and gelatin
manufacturing processes, as described in the information presented to
the Committee today, do the results of these new studies demonstrate a
reduction in infectivity that is sufficient to protect human health?\"

FDA\'s Question 2 addresses the Agency\'s guidance on gelatin. As you
have heard already today from Dr. Potter, in 1997, FDA issued a guidance
document that established certain parameters for the sourcing and
processing of gelatin in order to avoid BSE risk. At that time, the
effect of the gelatin manufacturing process on in infectivity had not
been proven. The data discussed with the Committee today, however, in
our view, provides a basis for concluding that FDA\'s guidance is no
longer necessary.

Indeed, as Dr. Chiu mentioned to you earlier, you may decide that
gelatin should be exempt that gelatin should be exempt from any FDA
restrictions. At a minimum, we believe that the guidance should be
modified so as to improve the opportunity for European raw materials to
be brought into compliance with the guidance while at the same time
maintaining appropriate controls on the use of European raw materials
and, as Mr. Masson expressed before, ensuring a continued adequate
supply of gelatin for pharmaceutical use.

If the Committee takes the approach of modifying the guidance in this
way, we request that the Committee consider two potential modifications
to the guidance. The current text of the guidance has been provided
already to you by FDA and, indeed, our recommended changes to the text
have also already been provided to you for your consideration.

First, FDA\'s guidance currently requires that \"cattle come from
BSE-free herds.\" As a practical matter, the term BSE-free herd refers
to a herd in which there has not been a single animal identified with
BSE. In Europe, it is mandatory, as you\'ve heard, that animals over 30
months of age be tested for BSE, whereas animals under that age are
normally not tested, because they have not been defined, at this time,
to pose a risk to human health.

Thus, in practice, a BSE-free herd is a herd in which BSE has not been
detected in tested animals. FDA\'s guidance in this regard would be
clearer if it were to include a brief explanation of the term BSE-free
herd by stating \"BSE-free herd as determined by generally accepted
testing procedures.\"

The second modification to the guidance that we would ask the Committee
to consider is one that this Committee has considered before. FDA\'s
guidance currently requires that heads, spines and spinal cords be
removed from gelatin raw materials \"directly after slaughter.\" In
1998, this Committee recommended that the removal of spines may be done
at any time during the deboning process. Indeed, the removal of heads
and spinal cords is not an issue as you heard, because they are already
removed before or at the time of slaughter.

Therefore, it continues to be appropriate for FDA\'s guidance to be
modified to permit the removal of spines at any time during the deboning
process. As the Committee considers FDA\'s Question 2B then, we request
that these proposed modifications to the guidance be taken into
consideration. A copy of our recommended changes to the guidance has
been distributed to you already for your consideration, and it also has
been made available to the public.

Thank you very much. We appreciate the opportunity to appear before you
today.

SECRETARY FREAS: Thank you for your comments. Is there anyone else in
the audience who would like to address the Committee, at this time? If
not, Dr. Priola, I would like to state that we all have three more open
public hearings throughout this meeting, and we do encourage the public
participation. Thank you.

CHAIR PRIOLA: Okay. So the questions put to us by the FDA are now open
for discussion and voting. Do we have the questions to put up? So the
first question, while they\'re getting it up there, is simply, well,
hopefully simply, do the results of these new studies demonstrate a
reduction in infectivity that is sufficient to protect human health? Are
there any comments or any discussion from the Committee? Yes, Dr. Hogan?

BOARD MEMBER HOGAN: Since nobody else is biting, let me take this
opportunity to say that when I reviewed your article and when I started
reading this voluminous amount of material, I sort of looked with the
same sort of skeptical eye that I do when I accept papers for
publication, and I initially had, when I started reading, several
questions about processing and scale-down issues and residual
infectivity, etcetera. But as I got deeper and deeper in this, we concur
that those had been addressed.

So the initial questions that I have asked today, I am extremely
personally pleased with the results of these studies. And while no study
can be absolutely perfect, and I think that all the questions that the
original Committee in 1997 had regarding the data, in my mind, have been
answered.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: I agree that these are very important experiments.
They were very well done. I read the reports also as somebody who has
done a lot of reviewing. I do have one remaining question or set of
questions. I\'m not sure that we know enough about the time course of
deactivation and why some of the infective agents seem to be so
resistent.

CHAIR PRIOLA: Well, maybe Dr. Rohwer would like to address that more
specifically, but, well, would you, Bob, would you like to, since this
is your day. I don\'t want to speak for you.

DR. ROHWER: You\'re asking a fundamental question of TSE science,
actually. And it\'s something that is going to get a lot of discussion
this afternoon. And so I don\'t know, I mean, I have another talk that
I\'ll be giving and it goes directly to that question, and Robert
Somerville has given his perspective on it, and we\'re going to hear
from David Taylor as well. And I think, is there anybody else? I
honestly can\'t remember. Well, and David Asher has some new data on
this area as well.

And you, yourself, put your finger, I think, on the central issues in
your first question to the panel, I mean, to the speakers earlier this
morning about the biphasic nature of these inactivations and what is
behind them. And we don\'t know for sure. My own bias is quite different
from Robert\'s. I mean, I don\'t think there is any intrinsic difference
between these agents. What we\'re talking about is sanctuaries and an
inability to actually reach all of the agent. But there are other
interpretations.

You pointed at one which is a genetic one and there are different ways
that you can look at these kinds of protections. We don\'t have the
answers to that. And I think it is a residual question that haunts every
single validation study, inactivation study that is done, is to know
just how far you can extrapolate this data to zero.

I would like to point out that this is not a new issue. It\'s something
that has bedeviled the vaccine industry, water purification, virtually
any area in which you want to assure that something is sterile, but you
have no way of measuring the entire production lot to find out whether
it is or not. And we\'re kind of in the same boat here.

BOARD MEMBER BAILAR: Yes, but I\'m not concerned about extrapolating
down past the last data point. You have data showing that the curve
flattens out, at least, to a considerable extent.

DR. ROHWER: Yes, and the point that I\'ll be making this afternoon is
that the place where that flattening out occurs is very context
dependent. And you can force it down or up depending on what kind of
mixture you are inactivating, what the conditions are and that type of
thing. And so the one thing I can say about these studies is that the
knowledge that that occurs was part of the design of the study.

And your other question about intrinsic versus extrinsic infectivity,
the idea that you have to introduce the spike into this spine
preparation at the beginning, and you can\'t know for sure whether you
have really mimicked the invito situation in which you would find the
infectivity in a BSE infected cow is a very appropriate one. However, in
this particular case, I feel more comfortable with it than practically
any other study like this that I have done, because it is the spinal
cord and the ganglia that we feel are the threat. They are extrinsic to
the bone.

And what was done here is the stuff was actually injected into the
spinal cord and smeared on the bone, actually given an opportunity to
dry on the bone, which is something that probably actually happens, and
is something that is very, in my opinion, probably very dangerous to do
with TSE infectivity. And so the original for the total process
experiment, which by the way I wasn\'t part of the experiment, but
nevertheless, my perspective on that is that that was probably just
about as good a spike as you could devise. And I mean, I can\'t think of
anything better.

You could always argue with the downstream position of spiking
homogenate into these things, but even there I think we\'re talking
about a worst case spike in the sense that the homogenate is completely
unrefined. And having taken this through the process, you\'re liable to
have stripped away some of the fats and things like that that may be
protective to these agents in pure brain type associations. But that\'s
speculation on my part. I can\'t satisfy your basic underlying concern
there, because we don\'t have data on that point.

BOARD MEMBER BAILAR: Well, I remain a little bit concerned, because I
recall reading basically in the [Daily Press]{.underline} that in the
usual method of slaughter, bits of CNS material do get into the
peripheral tissues. Is that correct?

DR. ROHWER: With penetrating concussive slaughter, I think, it is
without a question that that happens. And that\'s \-- I don\'t want to
comment on that. There are people here from the USDA who can probably
tell us just whether that practice still occurs there or not. I\'m not
sure.

BOARD MEMBER FERGUSON: I\'ll answer that. Actually, the issue is with
air injected stunning, where you\'ve got a captive bolt and then you\'ve
got holes drilled at the end of it, and you inject a bolt, a blast of
air, and that type of nomadic air injected stunning is not used in the
U.S. industry any more. Our colleagues at the Food Safety Inspection
Service are actually in the process of promulgating regulations that
officially prohibit that, but based on our understanding of slaughter
practices, it is not used in the U.S. Is it used elsewhere?

BOARD MEMBER WOLFE: Well, like in the countries where we\'re talking
about, the European countries?

BOARD MEMBER FERGUSON: In Europe it also prohibited by regulation within
Europe.

BOARD MEMBER WOLFE: Within all of Europe?

BOARD MEMBER FERGUSON: Yes, yes. Actually, well, within the EU. EU
regulations prohibit it. So within the community, I think, you can
probably also then assume that any of those countries that are exceeding
to the community, the same thing applies.

CHAIR PRIOLA: Dr. Somerville, I think, also wanted to address part of
your question, Dr. Bailar. Thanks, Bob.

DR. SOMERVILLE: Can I just \-- is this on? Okay. Just to add to what Bob
was saying and to emphasize what I said at the beginning of my talk, was
that in processes that were considering its denaturation reaction which
is, I suggest, leading to the stabilization of the aging, past the
drying processes that Barbara has just mentioned. There are other
processes involved in the gelatin extraction procedure which may assist
in its destruction or removal.

I suggest that possibly there is a degree of hydrolysis of infectivity
which would not necessarily depend on the stability of the agent in
terms of its denaturation properties, and also, of course, the
filtration properties described are of importance in removing, in the
totality of the removal of infectivity from the process.

CHAIR PRIOLA: Yes, I think it is also worth remembering that having sat
through many of these Committee meetings and always asking for data, I
now have before me 2 inches of data, all of which point to the same
thing. That in the worst case scenario you can still inactivate these
huge doses of infectivity. And then in the real world we\'re talking
about starting material that doesn\'t even have, at least from the
European point of view, as we\'ve heard, since they are now removing the
vertebrae, it doesn\'t even have that material there to start.

So whatever contamination may be present is going to be significantly
lower than anything that has been discussed here today. So at every step
of the process, precautions are being taken that should also be taken
into consideration when you\'re thinking about these things about total
inactivation and sequestering evasion.

Are we ready to vote, dare I ask? Does anyone else have anything they
would like to say now? Shall we call for a vote then?

SECRETARY FREAS: There are currently nine voting members at the table. I
will go around the table starting with Dr. Johnson over there. Dr.
Johnson, how would you like to vote?

BOARD MEMBER JOHNSON: I vote yes.

SECRETARY FREAS: Dr. Bracey?

BOARD MEMBER BRACEY: I vote yes.

SECRETARY FREAS: Dr. Ferguson?

BOARD MEMBER FERGUSON: Yes.

SECRETARY FREAS: Dr. Hogan?

BOARD MEMBER HOGAN: Yes.

SECRETARY FREAS: Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yes.

SECRETARY FREAS: Dr. Priola?

CHAIR PRIOLA: Yes.

SECRETARY FREAS: Ms. Walker?

MS. WALKER: Yes.

SECRETARY FREAS: Dr. Wolfe?

BOARD MEMBER WOLFE: Abstain.

SECRETARY FREAS: Dr. Bailar?

BOARD MEMBER BAILAR: No.

SECRETARY FREAS: The tally is 7 yes votes, 1 abstain vote, and 1 no
vote.

CHAIR PRIOLA: Okay. So we can move on to Part A of the second question,
which is due to scientific data and information available support the
following current FDA recommendation on bone gelatin. And we can keep in
mind that we can modify as the FDA has said we can modify this question
if we think it is necessary for this recommendation. So that\'s open for
discussion. Dr. Bailar?

BOARD MEMBER BAILAR: Before we vote on this, could we have somebody from
FDA say whether the modifications suggested are acceptable?

CHAIR PRIOLA: I\'m sorry, the modifications suggested by the gelatin
manufacturers?

BOARD MEMBER BAILAR: Yes.

CHAIR PRIOLA: Yes. Would someone from FDA, yes, Dr. Chiu.

DR. CHIU: I would put the question back to the Committee. If the
Committee think, you know, the modification suggested by industry is
acceptable, then we will take that recommendation back to the Agency and
then have internal discussion.

CHAIR PRIOLA: Comments from the Committee? I would like to read through
the gelatin manufacturers recommendations. Is there any overwriting
reason that anyone can see here to alter what the FDA already has down,
which seems to cover what it should in terms of removing risk materials?
Dr. Hogan?

BOARD MEMBER HOGAN: No, I don\'t think it should ever go under
non-exempt. I think that this is good. The question is from the
industry, why is it important to \-- when you say BSE-free herds, that
covers it. I guess what you\'re not allowed to use then are herds which
contain animals that are younger than 30 months, and you would like to
be able to do that. Is that the sense of why you want the modification?
Since animals that are less than 30 months are already assumed to be
BSE-free.

CHAIR PRIOLA: Dr. Schrieber, Mr. Schrieber?

MR. SCHRIEBER: This request for modification is based of an opinion
expressed by the USDA. USDA has stated to FDA we do not consider any
herd in Europe being BSE-free. So this means the current text, the way
this is written, would exclude altogether all European bones to be used
for gelatin manufacturing and then exported into the U.S. So therefore
we need the clarifications that under certain circumstances, and that\'s
what we have described, that the animals are tested according to current
procedures in Europe, that this would be, let\'s call it, equivalent to
the BSE-free herds. So that\'s one point.

And the other request for the modification is what I said before. Due to
the transport of the carcasses from a slaughter house to a meat packer
to sausage companies, with bone in, if the request will stay, removal of
spine, I\'m not talking about spinal cord, this is directly removed
after slaughter. But removal of spine has to be taking place directly
after slaughter, this would as well totally exclude the use of European
bones, because this is not the standard procedure.

So we need some time frame during the further processing, because
deboning is done somewhere else and transport of carcasses without the
bones is not possible. This is the ratio behind our request.

BOARD MEMBER HOGAN: Well, then I would ask Lisa, is that true the USDA
considers no herds in Europe BSE-free, despite testing?

BOARD MEMBER FERGUSON: Well, I think what we\'re dealing here with is
the way our regulations are written. And our regs prohibit the entry of
ruminant from any country that is on the BSE restricted list. So, you
know, since our regs are clearly prohibiting all these animals, we
can\'t necessarily make an exemption and say yes, okay, something is
free, something is not free.

CHAIR PRIOLA: I\'m sorry. Dick, go ahead.

BOARD MEMBER JOHNSON: Yes. If this were modified by this Committee, that
would not affect the FDA regulations, and then you would have two
conflicting rules, right? Is that right?

CHAIR PRIOLA: Well, you\'re not necessarily going to have two
conflicting rules. You know, the way our regs are written, we prohibit
gelatin from entering, as Dr. Morris has described, unless it can be
demonstrated that it is not going to go for animal use. Okay.

BOARD MEMBER FERGUSON: So we don\'t make this type of an exemption, you
know, for stuff going for animal use, if that\'s clear.

BOARD MEMBER JOHNSON: I thought it was all products derived from cattle
that were from BSE positive countries that you don\'t permit. But as
long as we eat it, it\'s all right? As long as humans eat it.

BOARD MEMBER FERGUSON: APHIS\' authority is related to animal health
issues. APHIS\' authority is not related to public health issues, so our
regs are written based on that authority.

BOARD MEMBER JOHNSON: But doesn\'t your animal health issue say that
products derived from, cattle products derived from BSE positive
countries cannot be brought into the country?

BOARD MEMBER FERGUSON: Correct. Our regs in general prohibit not just
bovine products, but most ruminant products.

BOARD MEMBER JOHNSON: Yes.

BOARD MEMBER FERGUSON: From countries on our BSE restricted list.
However, I think, as Dr. Morris explained in her presentation, there are
certain things in the regs that can be allowed entry and one of those
things is gelatin under certain conditions that is not going for animal
use.

BOARD MEMBER JOHNSON: That\'s in your exemptions at FDA?

BOARD MEMBER FERGUSON: Correct.

CHAIR PRIOLA: This is USDA, Dick, so, yes. They are USDA.

BOARD MEMBER JOHNSON: USDA, that\'s okay.

CHAIR PRIOLA: Yes, the FDA is strictly concerned with oral or topical
applications in humans of gelatin, so the USDA regs aren\'t our concern.
It\'s the FDA. It\'s this specific recommendation by the FDA.

BOARD MEMBER JOHNSON: Except it isn\'t \-- wouldn\'t it be a regulation,
for instance, that you couldn\'t bring in cattle hides from Europe under
the safety of animals?

BOARD MEMBER FERGUSON: No, hides and skins are exempted from our regs.

BOARD MEMBER JOHNSON: They are exempted? Okay.

BOARD MEMBER FERGUSON: They are considered, yes, not to present a risk
of transmission.

BOARD MEMBER JOHNSON: Okay.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: How is herd defined? Is that all the animals on a
single farm or ranch?

BOARD MEMBER FERGUSON: I don\'t know why you guys are looking at me,
because these aren\'t our regs. Actually, I have to admit, I mean, these
are the types of things that you always run into when you put that type
of a thing in a reg. It\'s very difficult to define that. When we look
at it from an animal health point of view, it\'s a group of animals that
is housed together. And if a premise has, you know, two separate groups
of animals that never come into contact with each other and are managed
completely differently, those could, technically, be considered two
different herds. But essentially, it\'s a group of animals that are
managed together and handled together.

BOARD MEMBER BAILAR: Okay.

CHAIR PRIOLA: Mr. Dwyer has been standing there for a couple of minutes.
Would you like to make a comment?

MR. DWYER: Yes, thank you. As you\'ve explained, there is a complete
distinction between the FDA\'s guidance and the USDA\'s regulations. The
USDA\'s regulations are intended only to protect animals and not to deal
with anything that the FDA has going on here. If you go back and look at
the early meetings and transcripts of this Advisory Committee when FDA
was discussing with the Committee the formulation of this guidance,
you\'ll see that the requirement for BSE-free herd restriction was put
in as one of a series of restrictions in FDA\'s guidance that were
intended to protect the safety of gelatin.

There wasn\'t much discussion, at that time, of what a BSE-free herd
meant or how that would be defined. Because it is obviously possible to
define it in many different ways, what the industry was looking for is a
way of defining it in a logical, rational way that is consistent with
current practice in Europe. And that\'s basically it.

BOARD MEMBER BRACEY: Although \--

CHAIR PRIOLA: Go ahead.

BOARD MEMBER BRACEY: Although I think the safety of gelatin has been
certainly demonstrated to be rather robust today, what bothers me is, in
essence, a disconnect between two levels of animals. One is the human
where we are considering saying that well, it\'s okay, based on the data
that we see, to allow humans to ingest these materials. Whereas, on the
other hand, another arm of the Government says that another animal,
which some of us think would be on a lower level perhaps than the human,
that it is not acceptable.

And, you know, I really feel that we need to have some sort of
harmonization, because the message, I think, that \-- if I were the
public, I would be somewhat concerned about the message that we would be
issuing.

BOARD MEMBER FERGUSON: Yeah, that\'s a valid point. I would ask
everybody, however, to keep in mind that, you know, it\'s one thing to
talk about an agent that is coming from cattle and going directly back
into cattle versus an agent that is coming from cattle and is going into
a different species. Granted, it has been demonstrated that there is
that transmission, but you do still have somewhat of a species barrier
there.

CHAIR PRIOLA: Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yeah, and actually, listening to the USDA
presentation, I have that same reaction saying there is an apparent
contradiction here between conditions for animals and humans. But in
thinking about it, I mean, you have a potential amplification. I mean,
these are some different issues that go into place with animals and
that\'s why I didn\'t comment. But there is an apparent contradiction. I
agree.

CHAIR PRIOLA: Yes, Lisa?

BOARD MEMBER FERGUSON: Can I go back to the term BSE-free herd? That\'s
very difficult to define and I don\'t want to necessarily sound too
harsh, but in some ways it is sort of meaningless. I know we have
struggled with those types of definitions as we tried to setup our
scrapie or CWD eradication programs. And, you know, specifically, as
we\'re doing our CWD program, we don\'t necessarily define under the
auspices of that program as herd as free until they have gone through a
5 year period with extensive surveillance and a lot of that. So it is a
bit difficult to define.

I guess I\'m not quite sure exactly what level of risk mitigation it\'s
necessarily adding in this guidance. Probably more of the risk
mitigation is coming from removing those tissues that are at highest
risk and also just through the inactivation of the process itself. So
perhaps what we should consider is that specific point even necessary in
there or does it just cause more confusion than it is really worth?

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: I feel like I just don\'t know enough about all
this. And I am concerned about the definition of a herd. Does this
include animals that come from the same source, prior to the time they
are parceled out into different farms? Does it include any element of
time? That is, you know, if all those animals there today are gone and
you bring in new ones, is that part of the same herd? What about overlap
in time, which I understand is common in the industry, that there is a
continuing flow of young ones in and older ones out. I just don\'t know
enough about it.

CHAIR PRIOLA: Dr. Chiu, do you want to comment on what the FDA means by
BSE-free herd or is there something more specific you can tell us?

DR. CHIU: Well, I will try. If I didn\'t get the picture across right,
then I will ask Dr. David Asher to add it. I think in our original
discussion we were thinking a herd is a group of animals managed, you
know, by the same people and also physically they are together, so they
are sort of separated from another group of animals. And also we think
when we say BSE-free means, you know, that group of animals in the past
there was never, you know, a BSE case among that group. In addition, we
were also thinking, you know, that group of animals were never fed with
meat and bone marrow, so therefore they don\'t have that kind of risk to
contact BSE.

CHAIR PRIOLA: Dr. Wolfe, did you want to say something?

BOARD MEMBER WOLFE: I just wanted to ask Lisa, just from your
perspective, what do you think the difference is between this guidance
or recommendation as it now exists and the way that the industry would
like to redefine it? I mean, the reason I\'m asking you is (a) you\'re
from the USDA, but (b) you have just gotten done saying you don\'t think
the phrase herd has any meaning at all. So if it doesn\'t have any
meaning, then what is the difference between our current version and
what they propose?

BOARD MEMBER FERGUSON: That\'s a good point, and actually I don\'t
really see a whole lot of difference in true meaning between what the
industry has proposed and what currently exists. My sense of what
industry has proposed is trying to make it more realistic and to make it
more meaningful in what fits with industry practices, which is a very
valid point, especially this one about the removal of tissues and where.

BOARD MEMBER WOLFE: I\'m specifically just talking about the herd
definition.

BOARD MEMBER FERGUSON: Actually, I mean, after what Dr. Chiu has just
said, you know, if those are the specific issues that FDA is intending
to address with that point, then I guess my suggestion would be to put
that in there as the guidance to say that these animals have not been
fed meat and bone meal, those types of things. That is a more accurate
definition of the risk mitigation measure and is more easily
understandable and leads to less confusion.

CHAIR PRIOLA: Looking though this, I don\'t have any trouble. I think
that\'s an excellent suggestion actually for the FDA to modify it
according to what they mean by BSE-free herd. The other suggested
modification by industry down there at the bottom, I\'m somewhat
uncomfortable with, but you had mentioned that you weren\'t as
uncomfortable. Why exactly is that?

BOARD MEMBER FERGUSON: Well, I think that\'s probably just because of my
understanding of slaughterhouse practices. And if this is saying, you
know, as it currently says, let me find it, \"and if the slaughterhouse
removes the heads, spines and spinal cords directly after slaughter,\"
that lends itself to a lot of interpretation.

First of all, talking about spine directly after slaughter, does that
mean right after the animal is stunned and, you know, hung up on the
rail and bled out? if so, that\'s not necessarily common practice. You
need that vertebral column there to give some structure to the carcass
that\'s moving through the plant. You know, and I think the point is
that those tissues are removed at some point in time during the process.
Although they are not going into the start of the gelatin manufacturing
process. It\'s not as much a point as specifically when are they
removed, it\'s that they are removed.

CHAIR PRIOLA: Which the current guidance says anyway. I mean, I don\'t
see where the industry modification makes that much of a difference if,
in fact, they take that out at the level of the slaughterhouse they take
out that requirement. The way I read it.

BOARD MEMBER FERGUSON: Well, I guess from an interesting point of view
and actually let me rephrase that. From a Government point of view, as a
federally employed Government veterinarian that might be put in a
position to certify to this, I probably couldn\'t. And it is just
because of the way that that is worded, where this stuff is removed
directly after slaughter.

CHAIR PRIOLA: Where does it say directly after slaughter?

BOARD MEMBER FERGUSON: Right in the text, yes. If the slaughterhouse
removes the head, spines and spinal cords directly after slaughter.

CHAIR PRIOLA: I just have after slaughter. Do I have the wrong one?

BOARD MEMBER FERGUSON: Can we put it up?

CHAIR PRIOLA: Oh, I see. You\'re looking in the \-- I see. It says
directly after slaughter if it\'s from a BSE herd. Later in the
recommendation it says if the slaughterhouse removes after slaughter. So
there is two different ones.

BOARD MEMBER FERGUSON: Yes, but even, I mean, the later one remove head,
spines and spinal cords as a first procedure following slaughter, that
just leaves open a lot of ambiguity and, you know, there are some of our
folks who are very literal, you know, when they would read that and say
oh, no, they didn\'t, you know, stun this animal, bleed her out and then
immediately remove things, therefore, I can\'t attest to that type of
certification.

CHAIR PRIOLA: I guess again, could we ask FDA, is there \-- since
that\'s a USDA interpretation of this recommendation, does the FDA have
the same sort of reservations or are they concerned about those same
sort of reservations as to when exactly the tissue is taken after
slaughter or is the discussion enough?

DR. ASHER: No, I think the discussion is very useful. My recollection of
the intent of the FDA with both those issues was that the reason why
BSE-free herds was specified but not defined was just to put the
industry on notice that under no circumstances did we consider material
from a herd recognized to have BSE as being an acceptable source for any
kind of gelatin entering the United States. No effort at the time was
made to define a BSE-free herd.

If one were to try to define an acceptable BSE-free source, I would
certainly agree with Dr. Chiu that it would not simply be all tests of
30 month old animals going to slaughter are negative. The herd would
have to have a certified history of never using food supplements
containing prohibited proteins. There would have to be an adequate
surveyance program, not just 30 months slaughter animals.

And my personal opinion would have to include a sufficient number of
older sentinel animals and, of course, careful veterinary surveyance to
make sure that all sick animals were recognized. My personal opinion
also is that this Committee not entertain an assertion that an animal
that tests negative at 30 months poses no threat to the public health. I
say both those things without attempting to influence the discussions of
the Committee. Thank you.

CHAIR PRIOLA: Would it be sufficient to say something like a BSE-free
herd is defined by the FDA, if that is in fact defined somewhere,
clearly?

DR. CHIU: No, we have not put in writing. And regarding the slaughter,
you know, the first procedure are directly after slaughter, I remember
our discussion in the past, was because if you remove spinal cord, it is
not possible, you know, to make sure entire cord, everything is removed.
You might have residual, you know, tissues. And if you carry that to
somewhere else and then remove the spine, then create contamination of
other tissues, in the bones of other tissues. So we thought, you know,
it would be better to remove, you know, the spine, the vertebrae at the
slaughterhouse. That was the thought at that time.

CHAIR PRIOLA: I guess the other thing to consider is, again, given all
the data we have seen showing inactivation of infectivity following the
gelatin extraction process, the issue of contamination,
cross-contamination by a spinal cord being removed at a different part
of the slaughter process may not be as major an issue given the fact
that now there are these five individual studies, all of which saying
that the gelatin process itself, as you get to the end, can remove
extremely high levels of infectivity under worst case conditions. So
it\'s possible that this discussion as to when things are removed and
may not, given that data, be as critical as it might have been before we
had access to this data. Dr. Bailar?

BOARD MEMBER BAILAR: Dr. Priola, we have had questions about some of the
wording in this recommendation, this draft recommendation. I have a
question about the last sentence that the processors are responsible for
the safety of what comes into them. Without offering any guidance about
that, would it be appropriate? I don\'t want to vote against this. On
the other hand, I\'m not very comfortable about voting for it.

Would it be appropriate to defer action until the next meeting with a
request that FDA consider revising the wording? I think the intent is
fine. I have no particular quarrel with the intent of the changes
proposed by the industry, but I think it needs some tightening up.

CHAIR PRIOLA: Well, I think in a way that\'s what the FDA is actually
asking us to discuss. Given what we have heard today and the current
discussion, how can we modify this or should we modify it in a way that
addresses the concerns of the Committee? So this, I would think would be
an opportunity to make that known, how you would want to do that.

BOARD MEMBER BAILAR: I\'m not sure we can modify it on the fly this way.
That\'s why I would like to allow a little bit of time for people who
know a lot more about the process, the problems, than some of us on the
Committee, and time for some reflection about the implications of any
changes.

BOARD MEMBER WOLFE: I would agree with John, because I think based on
what Lisa has said, which, I think, amplifies the understanding of the
process somewhat and what other people are saying, that the FDA has
gotten some input from us, which is what Question 2B is, and it would
make most sense for us to get at the next meeting the new version of the
recommendation to vote on.

CHAIR PRIOLA: Do you have suggestions for changes that we can make? I
mean, we still have to actually vote on Question 2A, but would you have
suggested for recommendations?

BOARD MEMBER WOLFE: I mean, defining, as Lisa suggested, what a BSE-free
herd is, sorting out the differences between directly after, immediately
after, first process or just after. I mean, there are three different
ways of describing in the current recommendation the timing between
slaughter and removal of spine, spinal cord and so forth, so I think, I
mean, those are, I think, two areas that need to be neatened or
tightened up.

CHAIR PRIOLA: Yes. Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yes, it\'s a question to the FDA. Can we vote on
this recommendation and then leave to the FDA to wordsmith the BSE-free
herd and the timing of removal based on the discussion that they heard?

DR. CHIU: I think we definitely can go back to before BSE-free herd, you
know. We have some idea, you know, over the years, you know, we have in
mind. We would like to get advised whether to remove the spinal, the
spine, the vertebras in the slaughterhouse is needed or not because of
the results you have seen, you know, from the validation studies.

CHAIR PRIOLA: Dr. Hogan?

BOARD MEMBER HOGAN: Well, it seemed, the validation studies suggested if
you can start with really high titre material, that you get rid of
almost, I mean, virtually totally. So I think what you start with in the
real world is sort of irrelevant, because it\'s never going to be as
high as what they are starting with in these validation studies.

Now, I am a little concerned that if you leave the spinal cord in and
then you drive, you know, 200 miles to have the spinal cord removed, it
is going to dry during that time period. Is that going to sequester some
agent that might be more difficult to remove later? But as I just said,
I think the titres will be much less than the validation studies. So I
personally wouldn\'t have a problem with that.

CHAIR PRIOLA: Mr. Dwyer, do you have a brief comment?

MR. DWYER: Thank you. Just as a reminder, this Committee voted in 1998
to agree with or recommend to FDA the removal of the spines in the
manner that we have suggested in our draft modifications to the
guidance. We have attempted to craft our draft modifications to the
guidance with respect to this issue, that is spine removal in a way that
is consistent with what this Committee recommended in 1998 as reflected
in the transcript of the meeting from April 1998. Thank you.

CHAIR PRIOLA: Dr. Johnson and then Dr. Ferguson.

BOARD MEMBER JOHNSON: Well, I think it\'s very impressive how much this
does decrease the infectivity. On the other hand, we should go back to
remember that there is that inactivated tail or whatever you want to
call it, so there is inactivated particles, and think back to the
Committee hearings after the Cutter episode with Jonas Salk where they
forgot, they neglected looking at the unneutralized tail, which caused
the whole Cutter episode.

Is it there and I think we should consider that. I don\'t think that\'s
enough to change the rules, but I don\'t think it\'s enough to say well,
let\'s not worry about splashing a little spinal cord around. I think we
still ought to keep that as tight as absolutely possible to keep the
contamination membranes of the spinal cord down.

So I would agree that I don\'t think we need to change. I voted yes on
1, but on this I would not want to see it made more permissive for the
possibility of contamination despite the good inactivation studies.

BOARD MEMBER FERGUSON: I guess I would like to briefly run through sort
of the standard slaughter practice at least in the U.S., and ask
everybody to sort of think about the possibilities for contamination.
You know, an animal comes in. It is stunned, rendered unconscious, then,
essentially, hooks are applied to the rear legs and it is bled out. The
animal is skinned. The head is removed and then, generally, the carcass
is split, at that point in time.

The standard practice is to go ahead and remove the spinal cord, at that
point in time. That is the easiest time to do it. But the issue is not
necessarily the removal of the cord. The issue is the removal of the
spine and that, you know, vertebral structure that allows the carcass to
sort of hold together and it\'s going through the rest of the processing
process.

So if concerns are about cross-contamination from removal of the
vertebral column later in the process, I\'m not quite sure where that
cross-contamination is going to come from, especially on bones and bone
chips that are going into a gelatin derived process, because even if you
assume okay, you can get some contamination when you split that carcass
in half or if you have a missplit, you\'re getting aerosolized cord that
is going on the surface of that carcass, and the bones aren\'t on the
surface of that carcass. The meat on those bones is removed elsewhere in
the meat cuts, and the contamination isn\'t necessarily going to be on
those bones, per se, which is what is going into the gelatin process.

CHAIR PRIOLA: I think we could \-- at least, the sense I\'m getting is
the recommendation as it stands needs some tightening up in terms of
clarifying definitions of BSE herd and when after slaughter things need
to be removed. The primary question prior to this is do the scientific
data and information available support the following FDA recommendation?
If the answer is no, what changes? Are there changes other than
tightening up these definitions that anyone would like to recommend?

For myself, the removal of the vertebral column, I think, is a big issue
for European countries because of the European BSE, but given that here
in the United States there is as yet no BSE and they haven\'t yet moved,
if I remember correctly from this morning, to removal of the entire
vertebral column, is that right, that has happened. There is a
significantly different level of risk, if I understand correctly. So
these rules seem to apply to really primarily European BSE countries.

Should we call for a vote on Part 2A and, if necessary, move on to Part
B with specifics? Are there any objections to that? If the FDA has
gotten what they need from the discussion, which I think they have, we
can move on to a vote for 2A.

SECRETARY FREAS: I will go around and poll the table exactly as last
time. Dr. Johnson?

BOARD MEMBER JOHNSON: With the likely changes made by FDA, do we vote
yes or no?

CHAIR PRIOLA: I think you vote no.

BOARD MEMBER JOHNSON: You vote?

CHAIR PRIOLA: And then we ask what changes for 2B. Isn\'t that right?
Well, actually, I think yes. Well, because I think that the

scientific \--

BOARD MEMBER JOHNSON: I would vote.

CHAIR PRIOLA: Yes.

BOARD MEMBER JOHNSON: If I looked at that just as it says, which states
there on the board, my answer would be yes.

CHAIR PRIOLA: Yes. I think you can vote yes or no and we can still make
modifications in 2B, because this Committee has never hesitated to make
modifications.

BOARD MEMBER JOHNSON: Then my vote stands, Sue.

CHAIR PRIOLA: So I\'m sorry, so what is it again, Dick, officially?

BOARD MEMBER JOHNSON: It\'s a yes.

SECRETARY FREAS: Dr. Bracey?

BOARD MEMBER BRACEY: I would second that yes.

SECRETARY FREAS: Dr. Ferguson?

BOARD MEMBER FERGUSON: Yes.

SECRETARY FREAS: Dr. Hogan?

BOARD MEMBER HOGAN: Yes, but we need modification.

SECRETARY FREAS: Dr. Khabbaz?

BOARD MEMBER KHABBAZ: Yes.

SECRETARY FREAS: Dr. Priola?

CHAIR PRIOLA: Yes.

SECRETARY FREAS: Ms. Walker?

MS. WALKER: Abstain.

SECRETARY FREAS: Dr. Wolfe?

BOARD MEMBER WOLFE: No.

SECRETARY FREAS: Dr. Bailar?

BOARD MEMBER BAILAR: No.

SECRETARY FREAS: The industry, would you, please, express your comments
on this? Okay. Out of the nine voting members at the table, we have 2
nos, 1 abstention and 6 yeses.

CHAIR PRIOLA: Okay. Under the part of 2B, even though we answered yes,
if I tallied right, there are three specific things that we would ask
the FDA to clarify. And that would be the definition of a BSE-free herd,
to make the recommendations at slaughter, directly after slaughter, you
know, just after slaughter, if they could be more specific as to when
the vertebral column should be removed, and also, Dr. Chiu had asked
specifically about whether removal of the vertebral column is necessary.
They wanted some clarification on that, too, I believe. Are there any
comments, Dr. Bailar?

BOARD MEMBER BAILAR: I would add a point also about some clarification
about the insurance by the processors that their supplies are adequately
protected.

CHAIR PRIOLA: Well, I think that \-- isn\'t that in the last? That is in
the last sentence, right, gelatin processes should ensure?

BOARD MEMBER BAILAR: It says the processors should ensure, and I would
like to know more about that.

BOARD MEMBER HOGAN: Well, unfortunately, Dr. Gambetti isn\'t here, but
from my experience in removing spinal cords, there can be left dorsal
root ganglia and other nervous tissues depending on how you do it. So I
think the issue of vertebral column if you\'re asking just for some
comments, may be important if you want to reduce that last little bit.

CHAIR PRIOLA: Does anybody want to recommend any specific language if we
can, I don\'t know if we can, to give the FDA some further guidance? You
know, for example, industry recommendation for BSE-free herd. Is that an
appropriate way to qualify it, to introduce the concept of testing
according to standard procedures? Lisa?

BOARD MEMBER FERGUSON: I guess I\'m uncomfortable with having testing in
there as the only thing that\'s qualifying the herd. You know, I don\'t
think that testing is necessarily the critical thing to hang your hat
on. I think the point is lack of exposure, and that should probably be
where that definition heads.

CHAIR PRIOLA: Are there any other comments as to specificity as to
modifications of the recommendation? Okay. Sidney, do you have anything
in terms of the slaughterhouse issue directly after? I mean, how
specific should specific be, given again all the data we have heard this
morning?

BOARD MEMBER WOLFE: No, I understand that, but, I mean, we heard that
there is a vastly different slaughtering process in Europe versus here,
so what I thought I heard this morning from the Europeans was that since
it\'s done in a different place, it\'s not even within the
slaughterhouse. I mean, is that correct? I mean, in Europe, you just
repeat what you said is the main difference between European
slaughtering techniques in terms of bone, in terms of getting the bone
for gel, as opposed to this country?

MR. SCHRIEBER: The difference is only the place of the removal of the
bones.

BOARD MEMBER WOLFE: The place, right.

MR. SCHRIEBER: Just the place.

BOARD MEMBER WOLFE: So it\'s not \--

MR. SCHRIEBER: Slaughtering practice is exactly the same, I think.

BOARD MEMBER WOLFE: But in one case, in this country, the removal of the
bones is in the slaughterhouse and there, somewhere else? That\'s the
difference. So it\'s the issue of transport and whatever. So it\'s
beyond just where in the slaughterhouse. It\'s in the slaughterhouse or
not. The issue is whether we think that it\'s okay for \-- which is the
issue the industry raised, whether we think it\'s okay for the bone
removal to be somewhere else with at least risks to workers and others
that are different than they would be if it were all done within the
slaughterhouse.

CHAIR PRIOLA: Go ahead.

MR. SCHRIEBER: In addition to this, the places of the removal, which
other meat processes are, are exactly under the same supervision of the
authorities of the public like the slaughterhouse itself. They are meat
processors, so they have to follow the same rules. It\'s just a question
of distance. It\'s not a question of how procedures are done, whether
they are inspected, whether they are controlled. That\'s all the same
whether it\'s here or there.

CHAIR PRIOLA: Are there any other comments? I guess I should ask the
FDA. Do you have sufficient information in terms of what the Committee
is asking for, for modification to the recommendation based on the
discussion and what was just said?

DR. CHIU: Well, in my mind, I\'m still not quite clear, you know. We
have read and heard, you know, the study results and as many of you
expressed, it\'s quite impressive. So I am not quite clear, you know,
whether we get any advice. Is it necessary to remove spine and if it\'s
necessary, then when it should be done? So if we can, you know, have a
little more discussion whether the spine, the vertebrae, actually needs
to be removed.

CHAIR PRIOLA: Lisa?

BOARD MEMBER FERGUSON: Yes, I guess I\'ll throw my two cents worth in
here and everybody else can have at it. I guess, I think it\'s important
to essentially limit the use of vertebral column in the production of
gelatin or say, you know, you are not using vertebral column in the
production of gelatin. I don\'t think it makes any difference where or
when that is removed, but to say yes, it\'s not included, it\'s not
going into the gel balm is important.

CHAIR PRIOLA: Yes, I would actually agree with that, that it\'s
important that it is being removed given the data where heard. Where
exactly it\'s removed may not be that big of an issue since you can
activate, apparently, quite effectively quite a bit of infectivity that
might be residual on the bone surface after removal of the spinal cord.

I am actually comfortable if the FDA does tighten up the definition of
BSE-free herd. I am comfortable for myself with the recommendation, how
it sits, with just some tightening up of those definitions, BSE-free
herd, as well as being careful when you describe when the vertebral
column should be removed after slaughter. I think in Europe, all the
vertebral columns are removed anyway, so that is moot. It\'s just where
the removal is, and that is not a primary concern for myself.

Would anybody else like to contribute? Is it just too near to lunch? Are
we running out of steam? Well, if anyone has any \-- I mean, so I guess
we have addressed the questions and if anyone else would like to say
anything after lunch, feel free to do that. When we restart the session
at 1:30, 1:40?

SECRETARY FREAS: Let\'s try 1:30.

CHAIR PRIOLA: Okay. Let\'s go for 1:30.

(Whereupon, the hearing was recessed at 12:45 p.m. to reconvene at 1:37
p.m. this same day.)

A-F-T-E-R-N-O-O-N S-E-S-S-I-O-N

1:37 p.m.

SECRETARY FREAS: Okay. Thank you very much for rushing through lunch. In
the afternoon, we are very fortunate. We will be joined by four new
temporary voting members. I would like to go around and introduce them.
I won\'t introduce the whole table, just the four new temporary voting
members. Well, I will introduce at least three of the new temporary
voting members.

On the far side of the table is Mr. Terry Rice, Board of Directors,
Committee of 10,000 from Windham, Maine. Would you raise your hand, Mr.
Rice? At the corner of the table right in front of the screen is a new
voting temporary member for today, Dr. Charles Edmiston. He is associate
professor of surgery, Medical College of Wisconsin, and he is also chair
of the CDRH General Hospital in Personal Use Device Panel, and he will
be taking these issues from today to his center\'s panels.

And we will be very shortly joined by Dr. Kenrad Nelson, who will be
sitting next to Dr. Priola, our Chair, and Dr. Nelson is a professor,
Department of Epidemiology, Johns Hopkins University School of Hygiene
and Public Health, and he is chair of the Center of Biologics Blood
Products Advisory Committee. And there is one more person, and that is
also from the Blood Products Advisory Committee. That is Dr. David
Stroncek, Chief Lab Service Section, Department of Transfusion Medicine,
NIH. And to all four of you, I would like to welcome you to the table
and thank you.

CHAIR PRIOLA: Okay. We\'ll go on to starting with Topic 2, which is BSE
in Canada, and I want to let the Committee know that this is an
informational topic only. It\'s for our benefit. We\'re here to listen,
and there are no questions being posed to us. This is an informational
topic only. The first speaker is Dr. Jay Epstein from FDA.

DR. EPSTEIN: Thank you, Dr. Priola. Before Mr. Hills makes a
presentation on what is known about the reported case of BSE in Canada,
I would just like to take a moment to make a brief statement about
FDA\'s current thinking in regard to potential implications of that case
report for blood safety policy.

FDA is undertaking an assessment of the BSE exposure risk to blood
donors in the U.S. and Canada in light of the single BSE case that has
recently been reported in Canada. Although, it is premature for the FDA
to present any results of this assessment now, we believe that the
likelihood of exposure to the BSE agent for both Canada and the U.S. is
and has been very small.

The exact magnitude of BSE risks for Canada and the U.S. will be
difficult to quantify because of methodological limitations. However,
preliminary considerations suggest first that the risk of exposure of
blood donors in North America to the BSE agent has been extremely low
and is even lower now than it was several years ago. And then secondly,
in particular, implementation of the feed ban of 1997 both in the U.S.
and in Canada significantly reduced the likelihood of human exposure to
the BSE agent for both countries.

FDA does not believe that there are sufficient data, at this time, to
warrant changing our blood donor deferral guidance. However, we will
continue to study this issue and will take further action as
appropriate. Thank you very much.

BOARD MEMBER WOLFE: I have a question.

CHAIR PRIOLA: Dr. Wolfe?

BOARD MEMBER WOLFE: Given that the spectrum of countries for which there
are limitations on blood donation go from UK, lots of cases, to EU with
some countries with very small numbers of cases in just cattle, and that
Canada is still a \"moderate risk country\" that has had a case, the
benefit risk equation is always important, and have you at least tried
to get some data, so that this question can be answered better later as
to what fraction of the blood supply in this country would be affected
if there was some sort of constriction on the ability of people who have
spent whatever amount of time in Canada?

Is there at least some effort to collect that, because otherwise, as we
learn more about the possible risk, small though it may be, and the
benefit, which is having a blood supply that is not impaired some comes
up to it? So just a simple question. Is someone trying to get a hold of
those kinds of data?

DR. EPSTEIN: Yes, Dr. Wolfe. Thank you for that question. We are mindful
of the need to try to assess the impact on the blood supply of any
potential change to our donor exclusion policy and, indeed, we have
already had dialogue with major blood organizations on both the
feasibility and scope of surveys that could establish the impact of any
candidate deferral policy related to Canadian exposure on the U.S. blood
supply.

And more broadly speaking also, we are thinking about similar questions
as they might pertain to say exposure in Japan or other countries that
have had case reports of BSE in cattle. So that enterprise is recognized
and is ongoing.

BOARD MEMBER WOLFE: So you are going to be collecting data on what this
impact would be?

DR. EPSTEIN: Well, blood organizations have been asked \--

BOARD MEMBER WOLFE: Right.

DR. EPSTEIN: \-- if they would collect such data, and we have had
preliminary statements of agreement.

BOARD MEMBER WOLFE: Okay. Thank you.

CHAIR PRIOLA: Any other questions for Dr. Epstein? Okay. Thank you very
much.

DR. EPSTEIN: Thank you.

CHAIR PRIOLA: Our next speaker will be Dr. Robert Hills from Health
Canada Ottawa who will discuss the review of Bovine Spongiform
Encephalopathies in Canada.

DR. HILLS: All right. First, thanks very much for inviting me to give
you a little update of what the situation is in Canada right now with
respect to BSE. I will just wait for the slides to come up. There is a
fair bit of information on the slides, so what I will try to do is go
through it relatively quickly. All right. Thanks a lot.

First of all, I thought what I would do is give you a little bit of
background to what Canada has been doing with respect to BSE before we
found a case in May of this year. First of all, there was a prohibition
of the importation of products assessed to have a high-risk of
introducing BSE in Canada. There was importation of meat and meat
products only from countries that Canada recognized as being BSE-free.

In 1990, there was a designation of BSE as being a reportable disease in
Canada, and any suspect case of BSE would be reported to a federal
veterinarian. In 1992, there was the creation of the National BSE
Surveillance Program. In 1997, the same year as the U.S. did, as well,
there was the implementation of the feed ban of feeding rendered protein
products from ruminant animals to other ruminants. In the year 2001,
there was the creation of the Canadian Cattle Identification Program for
Cattle and Bison making it possible to trace individual animal movements
from the herd of origin to the slaughter.

Next slide, please. This is just a quick pictorial of sort of how Canada
has approached it. Canada has adhered to the OIE guidelines on TSE risk
management. Up until our finding of the case, we were considered to be
provisionally free, and we have also done a risk assessment that was
completed in December of 2002 with respect to Bovine BSE cattle in
Canada, and in that risk assessment, we actually determined that the
likelihood of finding BSE in Canada would be remote. That has changed,
but still remote.

Next slide, please. Before we go on, what I would like to do though is
just bring you back a little bit in time, because we did have a case
previously. In 1993, we did diagnose a case of BSE in a beef cow that
was imported from the UK in 1987. The exposure of this animal to BSE
occurred prior to its arrival to Canada. The index herd and all the UK
animal imports were destroyed, at the time, and it was subsequently
determined that the UK herd, which was a source cow for this particular
animal, did have other infected animals, as well.

Next slide, please. This graph here is, again, a pictorial showing the
importation of animals into Canada, particularly, and we\'re it the
North American Disposition of Imported UK Cohort Members prior to the
index case discovery in 1993. So there was importation of animals prior
to our taking action in 1993 to eliminate those animals.

Next slide, please. So getting on to our first indigenous case. January
31, 2003, a 6 to 8 year-old downer beef cow from northern Alberta went
to slaughter to a provincially licensed meat facility. Alberta
Agricultural Food and Rural Development meat inspector condemns the
carcass as being unfit for human consumption.

At that time, the head was collected and submitted as part of our
National Surveillance Program, the Federal Provincial Surveillance
Program for BSE. And the carcass, at that time, because it was
condemned, was sent to inedible rendering.

Next slide, please. On May 16^th^, the testing was completed with a
tentative diagnosis of BSE by the Alberta Ministry of Agriculture. The
sample was then sent immediately to Canadian Food Inspection Agency\'s
National Center for Foreign Disease in Winnipeg, Manitoba where they
also confirmed BSE, and then the sample was subsequently sent to the
Veterinary Laboratory Agency in Weybridge, England, which is the OIE
Reference Center for BSE. And on May 20^th^, they actually confirmed our
diagnosis of it being an actual case of BSE. Immediately upon
notification, we notified the OIE, in fact, that we did have a case of
BSE.

Next slide, please. So what did we do from there? So we had a start in
epidemiological investigation. We basically broke it down into three
phases. The first phase, we\'re calling that the case itself, which
we\'ll call the animal trace back, its immediate management, which
we\'ll call the animal trace forward, and the most probably origins,
which is where did the animal get the exposure from?

Next slide, please. This is a well used graph. It is done in colors for
a particular reason. The red is the case herd where the index animal
was. The blue line is considered the primary line of inquiry where we
think the source animal came from, which is in Saskatchewan. The yellow
line is considered the secondary line of inquiry. That was an Alberta
line, and the green one in the middle is, in fact, that we did discover
that there was some commingling with another herd. So those are the
areas, which we were tracing out as part of this investigation.

Next slide, please. As I said earlier, the index case was a 6 to 8
year-old Angus. It was a member of a herd that was recently established
within a two year period between 2001 and 2002, and the animals that
made up that herd were from two farms. What we believe initially from
the age of the animal was that the expression of the clinical BSE at
this age offers the first epidemiological insight, which would probably
mean it was a low level BSE exposure given the age of the animal.

As I mentioned earlier in a previous slide, the Saskatchewan blue line
of inquiry was the most probable avenue for which the positive animal
moved to the Alberta farm. That particular line of inquiry, the animals
were culled and depopulated and tested, and all tests were negative by
Prionics Western Blot and immunohistochemistry.

Next slide, please. At the same time as we were culling and depopulating
and testing, we were also trying to confirm through DNA testing the
origin of the particular index animal. Unfortunately, the DNA testing
did not come back with a clear definitive result, and as a result, we
needed to then move down the second line of inquiry.

So we proceeded with the depopulation and testing of animals in the
Alberta line of inquiry, which was that yellow line in the previous
pictorial that I showed you. We culled those animals and tested those
animals and all tests came back negative again from Western Blot and
immunohistochemistry. Even though we didn\'t have a confirmed definitive
answer for the DNA, it is still a probably line of inquiry, and most
probable one, is the Saskatchewan blue line for the introduction of the
animal to this farm.

Next slide, please. The next phase, which was the Animal Trace Forward
Investigation, was to determine what would happen with the animals that
left the farm. So there was movement. We looked for the movement of the
cattle from the index herd. We looked at, as I mentioned earlier there
was a green box, where there was some commingling. We traced out those
animals. We culled them. We depopulated them. We tested them. We found
that they were all negative by Western Blot and immunohistochemistry.

Next slide, please. So to summarize, the Trace Forward and Trace Back
Investigations, we had 15 premises that were quarantined, an additional
25 herds were scrutinized and the tracing-out of single animals or
cohorts from the Saskatchewan line of inquiry.

The trace out also included the identification and notification of the
export of five animals to the United States, which you all should be
aware of, we did let you know as soon as we found that out. And in all,
we had a culling of more than 2,700 animals, 2,000 of which were 24
months of age and older, and all, as I mentioned earlier, have been
found negative by Prionics Western Blot and immunohistochemistry.

Next slide, please. This is just a pictorial or a graphic of the
disposition of the carcass of the particular BSE index case. It shows
the yellow line, which shows the line of investigation with respect to
the use of the products for laboratory testing. We have the lighter blue
line, which shows what happened with the processing of the hide. And
then, again, we have the mauve or the purple, which is moving the
carcass to inedible rendering and what happened to it from there. And as
you can see, it did get rendered into some meat and bone meal.

Next slide, please. So we then looked at the Feed Investigation. Since
the index cow was condemned unfit for human consumption, its carcass was
sent to an inedible rendering. And I would want to reemphasize because
it was sent to inedible rendering, there was no part of the animal that
actually went into the human food chain.

Next slide, please. The carcass of the index as I showed you in the
previous pictorial, the carcass of the index case was traced via
Canadian Food Inspection Agency from the abattoir, the renderer, the
feed mill, the producer continuum to its direct allocation into pet
food. And in the pet food case, we did find that actually there was some
pet food that actually was exported to the U.S. to which we notified the
U.S. when we found that out, and there was pet food in Canada, as well.
And there was the production of meat and bone meal.

What is important to take out of this though is the visit to the
renderer and the feed mills confirmed adherence to our feed ban or meat
and bone meal feed ban legislation on the product receipt, segregation,
labeling and distribution. So there was no breach in compliance, at that
point, at the renderer. So it was not labeled to be fed back to
ruminants.

Next slide, please. Further investigation was the trace out of the feed
to individual farms, and we did find that three additional farms were
quarantined when the investigation could not preclude exposure of 63
head of cattle to the feed destined for poultry feed. In that case, it
was evidenced that the farm itself had allowed poultry feed to be fed to
ruminants. The animals were culled and tested by Prionics Western Blot
and immunohistochemistry and, again, all those animals came back
negative.

Next slide, please. This is just a pictorial, again, of what I was
describing before from the inedible material from the index case going
to the renderer. It went up to pet food. It went into poultry and some
pet food, and there was the feed mills that we traced out afterwards. So
this is sort of managing the risk of the disposition of the material
from the index animal.

Next slide, please. There were other considerations that we wanted to
take into account as we proceeded in with the exposure investigation. We
did look at maternal transmission. We looked at contaminated meat and
bone meal used in feed products, particularly early risk factors, any UK
imports slaughtered prior to 1993 or other European imports. It was
figured into our investigation.

We looked at TSEs resident in other animals, CWD and scrapie as
examples, and we did look at the possibility of it being a spontaneous
case. Our investigation right now is at the point now, we are looking at
feed products that are considered the most probable root of exposure.

Next slide, please. Again, this is just a graphic again to illustrate
the hypothetical foreign domestic exposures of the index case, and it\'s
just groups of, you know, what the possibilities might be for the
exposure of this particular case.

Next slide, please. What we did find out in the investigation is there
was two potential meat and bone meal epidemiological exposure roots that
were identified. The first was a feed concentrate and the second was a
high energy feed block. Both have incorporated meat and bone meal, at
some point in time. The investigation did find though that the feed mill
records and compounding formulae confirmed that the meat and bone meal
incorporated in both of these products was curtailed in 1997 upon
implementation of the meat and bone meal feed ban.

Next slide, please. So what can we conclude? What we can conclude so far
is that discovery of BSE in Canada proves that the active surveillance
and the diagnostic programs were working, because we did find the case.
Epidemiological evidence supports the probability that BSE in this case
animal was associated to exposure to infected material through the
feeding system, at some point, early in the animal\'s life.

Next slide, please. What we felt that we needed to do was to ensure that
what we were doing was accepted and would be recognized, so it was
decided that what we would do is convene an expert panel to actually go
over our procedures and how we did it and what we were doing and what
actions we were going to be taking, and get their recommendations back
to us.

That particular panel comprised of Ullie Kihm from Switzerland, Will
Hueston from the USA, Dagmar Heim from Switzerland, and we did have
contact with Stuart MacDiarmid from New Zealand, as well. The first
three met on June 7^th^ to 9^th^ and met with the members of the
Canadian Food Inspection Agency and Health Canada to which the team was
provided with an overview of the epidemiological investigation. All
actions taken to date and the scope of the options and the measures
being considered to adjust domestic policies.

Subsequently to that meeting, the team went back and did actually do a
report, and what the panel did find was that the \-- they found that the
risk management measures put in place in Canada achieved the desired
outcome. The surveillance did detect the case with BSE. The animal did
not enter the food chain and the measures in place have reduced the
spread and amplification of BSE in Canada.

Next slide, please. They did come back with some specific
recommendations, however, for us to strengthen our current situation.
They did say that there should be a prohibition on Specified Risk
Materials in human food and animal feed, including advanced recovery
meat products, tighter controls on non-ruminant feed, enhanced audit and
compliance, strengthening the existing cattle identification tracking
and tracing systems that are existing in Canada, enhanced disease
testing and surveillance by increasing the coverage of fallen and dead
stock, downer and diseased animals, and to work at efforts to improve
the awareness among producers, veterinarians and the general public with
respect to BSE.

Next slide, please. So what will Canada do? Well, the government of
Canada will be responding to the recommendations of the International
Team. We\'ll respond through our consultation process with our provinces
or territories, the Canadian industry, our U.S. counterparts and other
trading partners, and there will be a new policy measure for Specified
Risk Materials as being the first step.

Next slide, please. I felt before we sort of should go a little bit
further, I would just give you a little bit of background about \--
because I\'m going to be talking about Specific Risk Materials Ban right
now, which is the first step, that in Canada, 95 percent of the
slaughters is in federally registered establishments and the majority of
those animals that are slaughtered are between the age of 18 and 24
months.

5 percent of the slaughter is in provincial abattoirs and the majority
of those being over 30 months of age. Only animals slaughtered in
registered establishments can be exported. The provincial slaughtered
animals can only be traded within provinces and sold within provinces.
If they are to leave provinces or to leave the country, they have to be
at a registered establishment. Removing SRMs at the point of slaughter
and disposing of them, we estimate removes about 99 percent of the human
exposure to potentially infected material.

Next slide, please. The immediate objective of the SRM policy is to
establish a requirement that the SRMs be removed at the time of
slaughter, and that they be removed from human use, human food and human
use. The new policy will define Specified Risk Materials and require
removal, as I mentioned earlier, at slaughter.

The list that I have right here are some of the things that we have been
considering as being the most probable. They will likely include the
brain, spinal cord, dorsal root ganglia, eyes, tonsils, skull and distal
ileum.

Next slide, please. Following our first step, as was recommended in the
expert panel report, there will be other measures that will be taken.
There will be areas that will be looked at with restrictions on animal
feed and process and protect human and animal health, that should be,
expanded surveillance, as was mentioned earlier, expanded food safety
plans, comprehensive tracking and tracing systems and national standards
and approaches will be implemented in Canada.

Next slide, please. And that concludes a very quick overview of what we
did. Hopefully, it did give you an idea of the scope in which we reacted
and what we looked at. I have listed here a number of different sites
that you can look at for updates. We\'re trying to be as open and as
transparent about our investigation or actions as we possibly can, and I
would encourage you to go to these sites and go to this to get the most
up to date information. That\'s it.

CHAIR PRIOLA: All right. Thank you, Dr. Hills. Dr. Wolfe?

BOARD MEMBER WOLFE: This morning, we sent a letter to the Secretary of
Agriculture, Veneman, strongly urging them not to lift the ban on meat
coming from Canada to this country, and one of the questions we have
was, and I will just read you three sentences, because it\'s really the
form of the questions.

Public information regarding the enforcement of the Canadian Feed Ban,
and we know it went into effect in \'97, but we also know the data from
the United States show very spotty and uneven enforcement, particularly,
the first few years of the ban, which is very similar to the U.S. ban
and was enacted about the same time. It\'s available, it\'s data
unenforcement. It\'s not on the website of the Canadian Food Inspection
Agency and a telephone call to the agency requesting these data has not
produced any information. Most tellingly, the report from the team of
international experts, which is, I assume, the one you just referred to.

DR. HILLS: Yes.

BOARD MEMBER WOLFE: That investigated the Canadian government\'s
response to the outbreak makes no mention of compliance with the feed
ban. It mentions the feed ban, of course, but data incompliance.

DR. HILLS: Yes.

BOARD MEMBER WOLFE: It is simply impossible to assess the wisdom of
lifting the ban you wisely put in place, you, Secretary Veneman, in this
case, on an emergency basis without these data.

So my question to you is what is your knowledge? I thought your
presentation was excellent. What is your knowledge of looking backward
how effective the enforcement of the feed ban has been from 1997 when it
was imposed to now?

DR. HILLS: Unfortunately, I don\'t have the history to go back in time
from \'97 backwards or forwards. I know it has come up in our
discussions numerous times, being able to put some sort of quantifiable
number to it and to try to do that. I have not yet seen that myself. We
are trying to determine that now, because we have had investigations. We
have looked at it. We have found that our plants themselves have been in
compliance.

As I noted in our Feed Investigation, we did find though that there was
a possibility that if a farm was coproducing, that there was a
possibility of cross-contamination, if you want to call it that, and
those are things that we\'re trying to address now to improve.

BOARD MEMBER WOLFE: Yes, I mean, needless to say, it\'s the essential
issue, because you have admitted that that\'s the most likely place that
the cow that got infected got infected from, and given \--

DR. HILLS: Well, no, that\'s not quite what I said.

BOARD MEMBER WOLFE: Well, I think it\'s all on your slides, for the most
part.

DR. HILLS: No, I think what I said was we found that there were three
slides as part of the investigation from the index case that actually
found that there were three farms in B.C. that actually were not in
compliance, because they inadvertently had fed or that we couldn\'t
definitively tell whether or not the feed that was destined for poultry
did not end up having inadvertently been fed to ruminants. What I said
was that our most likely possibility would be \--

BOARD MEMBER WOLFE: The feed.

DR. HILLS: \-- exposure before the feed ban.

BOARD MEMBER WOLFE: For that particular cow.

DR. HILLS: Yes.

BOARD MEMBER WOLFE: But, you know, again, are there going to be some
data on enforcement? I mean, I assume that once it went into effect,
there was some kind of government effort.

DR. HILLS: Yes.

BOARD MEMBER WOLFE: To check on enforcement.

DR. HILLS: Yes.

BOARD MEMBER WOLFE: When can we expect to see those data?

DR. HILLS: I can\'t give you a date on that. I can certainly find out
for you, but I cannot give you that, but I am not aware of what date
they are going to be able to make that information available.

BOARD MEMBER WOLFE: Okay. Thank you again, very good presentation.

CHAIR PRIOLA: Dr. Johnson?

BOARD MEMBER JOHNSON: Yes. Dr. Hills, a question that was brought up
early on was the possible U.S. origin of that cow, that it might be a
North Dakota or Montana cow, and it may have been our feed ban was the
problem. Is there any further data on that original origin of it, and
has the U.S. origin been ruled out?

DR. HILLS: The data we have right now suggests that the line of inquiry
was the source was the Saskatchewan farm right now. We have no
definitive evidence that would say that it was an animal that was
imported from the U.S.

CHAIR PRIOLA: Dr. Gambetti?

BOARD MEMBER GAMBETTI: Can you describe the procedure that Health Canada
uses to diagnose this particular animal and suspected animal in general?
You listed Prionics Western Blot and the immunohistochemistry. Is that
done in more than one area and in that particular animal that turned out
to be positive, were both positive? Can you, in other words, amplify a
little bit on how the animal or another animal are studied in Canada?

DR. HILLS: If I understood your question correctly, Canada has a
national TSE laboratory network. The gold standard, the
immunohistochemistry test, is a test that is used, was used in all these
laboratories, and so there was, as I mentioned earlier, the National
Surveillance Program.

The Prionics test for Western Blot was brought in and we were evaluating
it, at the time, but it was brought in for use mainly because we had so
many animals to do, at the time, that we needed to find some mechanism
by which we could up the volumes, and at the same time, we feel that it
was a mechanism by which we could do validation testing on the Prionics
test itself.

So in doing that, the work was done in the Alberta and Winnipeg labs for
the Prionics test, but we\'re in the process now of looking at and
getting that test now distributed across the TSE laboratory network.

BOARD MEMBER GAMBETTI: How many tissue samples or brain area were
examined, only one, the lower medulla or more than one and were all of
them, if there were more than one, were they all positive?

DR. HILLS: I believe there was more than one and yes, they were all
positive, but I can\'t give you the exact number.

CHAIR PRIOLA: Dr. Bracey?

BOARD MEMBER BRACEY: Perhaps along that line, I guess a question that
comes up is the issue of with any test, there is always the chance of
having a false positive the more times you do an assay. How are you all,
in essence, getting at that and is there a plan to actually look at
infectivity in some of these animals or do you feel, in essence,
confident enough with the assay system in terms of eliminating that rare
false positive?

DR. HILLS: Well, that was one of our concerns, was to go to tests other
than the immunohistochemistry analysis, but we felt that the information
that we have generated now based on the culling exercise that we have
gone through with the animals that we have right now, because each one
of those animals was testing in parallel with the gold standard test,
that we felt that we\'re starting to now get the numbers that would
indicate whether or not the test is, in fact, what the manufacturer
suggests, which is 100 percent no false negatives. So we are generating
the information now, and that is the only way that we can do it.

BOARD MEMBER BRACEY: What about the issue of false positives though?

DR. HILLS: Well, in doing the animals that we\'re doing now, we have
found no false positives, but that, as I said, was only 2,700 animals.
That is what we have done. That is the claim of the Prionics, I believe,
is what they are suggesting is it is being 100 percent accurate.

I did mention that the animals that we were talking about were \-- we
were targeting 24 months and older animals, because we still believe
that there is some possibility that the test will not work sufficiently
well for animals below that age, and so the testing system itself is
probably a little questionable for younger animals.

CHAIR PRIOLA: Dr. Nelson?

DR. NELSON: You mentioned that there were 2,700, I think, animals that
were tested and found not to be \-- not found any positives, but how
many were of the similar age to this animal? In other words, the
infection in this animal could have occurred six, seven years ago and
not shown up if the animals examined where younger.

DR. HILLS: Yes, that\'s a very good question. That\'s why when we were
looking at the culling and the depopulating, we were looking at the
specific herds, so we were getting an age distribution. I can\'t give
you the exact number of animals that were there, but because we were
targeting the animals older than 30 months, we would then take into
account some of those animals. And some of those herds on the
Saskatchewan side were breeding animals, so many of them were older. I
just can\'t give you the exact number.

CHAIR PRIOLA: Dr. Taylor, did you have a comment?

DR. TAYLOR: David Taylor from Edinburgh. You mentioned how you concluded
that because clinical disease emerged somewhere between when the animal
was somewhere between 6 and 8 years-old, that this may be reasonably
construed as evidence of low level challenge. Now, certainly, it is true
in the UK that incubation period as we have taken to broadly equate with
age, because meat and bone meal was usually and often only fed in calf
food.

But if the animal, in fact, received meat and bone meal A for the first
time or subsequently on several occasions after it was born, you can\'t
really pin down the incubation period. So it could still be a high level
dose if it got its meat and bone meal at a later stage.

DR. HILLS: Yes, you are correct. I have no trouble with what you\'re
saying, but I will go back to our feed ban that we have in place right
now. The fact that the feeding of animals back and forth, the feeding of
ruminant material back to ruminants is prohibited in Canada, so the
likelihood of that would be, in our estimation, remote, not nil, but it
would be remote. So yes, so that\'s what we\'re saying is we do think
it\'s preceding the feed ban, so that would make it then \'95, \'96,
somewhere around in there, and it is a possibility, yes, at that time.

CHAIR PRIOLA: Dr. Bailar? I\'m sorry, Dr. Bailar, excuse me.

BOARD MEMBER BAILAR: You mentioned the Canadian system for
identification, tracking and tracing of animals. How much help was that
to you in your investigation?

DR. HILLS: It was significantly helpful for us. Unfortunately, because
we instituted it in the year 2001, it really only was successful for
animals that were within that age group. Even though all animals are
tagged for movement now, what we can\'t do is really definitively say,
for example, the age of the animal, where the entire history of the
animal was, but for the younger animals, yes, we can do that. So after
2001, we can certainly trace it.

BOARD MEMBER BAILAR: Can anyone tell us about the present status of
moves to have such a system in the U.S.? I\'m sorry Dr. Ferguson isn\'t
here now. I\'m concerned, of course, about the possibility that there
might be a single case here sometime.

CHAIR PRIOLA: Yes, and I\'m not sure anyone here can answer that. That
is sort of a USDA issue, not an FDA one, I think. We\'re about to
proceed to the open public hearing portion. Do you gentlemen have a
couple of quick questions? You have been standing very patiently. Do you
have a couple of quick questions for the speaker?

MR. HAFFENDEN: Yes, Paul Haffenden from TerraCell. I would just like you
to comment. Several years ago, the European Scientific Steering
Committee assessed Canada and the U.S. as category 2 countries,
equivalent risk, given the \-- maybe you could comment on the movement,
high incidence of movement of animals between the two countries in both
directions, high incidence of movement of animal feed products between
the two countries in both directions, and then any comment on how you
think that might affect the adjustment and category risk now with this
case in Canada?

DR. HILLS: I don\'t think I can discuss anything about how that is going
to affect the categorization. I think that is something that somebody
else will determine, not me. But what I can say is that given the trade
between our two countries, there is movement of both animals and feed
across the border.

CHAIR PRIOLA: If it\'s a very quick question.

MR. BROOKLANG: Yes, Nelson Brooklang, Ortech International, New York.
You made a distinction between federally registered and provincially
registered abattoirs and the age of cattle that are processed in those,
and the fact that provincial cattle don\'t get exported. I wanted to ask
whether blood byproducts used in the biotech industry, what I am from,
like Bovine Serum Albumin transferred, purified from bovine blood could
be collected from provincial abattoirs in Canada and sold in the U.S.?

DR. HILLS: Good question. My recollection is that the material itself is
not. From the provincial licensed establishments, the provincial
government looks after that, and I do not believe it then goes to the
federally registered renderers, but I would have to reverify that. I\'m
not 100 percent sure.

CHAIR PRIOLA: Okay. Thank you very much for your presentation. I will
move on to the open public hearing portion.

SECRETARY FREAS: In response to our [Federal Register]{.underline}
announcement, I have received two requests to speak at today\'s open
public hearing for this afternoon, and the first one is Mr. Wayne Vaz.
Would you, please, come to the microphone and make your presentation?

MR. VAZ: Good afternoon. My name is Wayne Vaz. I am representing
Serologicals Corporation. We are a leading supplier of animal-based
products of the global health care industry. We are based in Atlanta,
Georgia with more than 800 employees worldwide. We greatly appreciate
the opportunity to be here today to talk about the TSE safety of our
bovine products and their critical importance in global health care.

Next, please. Our goal is to raise the level of awareness regarding the
pervasive use of bovine products in the production of life-saving drugs
and other essential health care products. We want to present the facts
according to high safety and qualify of Serologicals\' bovine-based
products, and we would like to work with the TSE Advisory Committee and
regulators to further develop industry guidelines to assure the
continued availability of bovine products.

Next, please. Serologicals is a global provider of biological products
and enabling technologies, which are essential for the research,
development and manufacturing of biologically-based life science
products. Some examples of our products include antibodies, cell culture
supplements, such as bovine albumin and other products for diagnostic
and research.

Next, please. This is a listing of our bovine-based products. We\'re
focusing on this, because these are the products that are believed to
offer a theoretical TSE risk. At present, we have two manufacturing
facilities, one in Toronto, Canada, the other in Kankakee, Illinois. We
have a third facility under construction in Lawrence, Kansas.

Next, please. Our bovine-based products are used in the development and
production of life-saving FDA-approved drugs, FDA licensed diagnostics,
medical devices and animal vaccines. Our bovine-based products are used
in the development and production of FDA approved biologics for
treatment of cancer, arthritis, Crohn\'s Disease, psoriasis, blood
clotting disorders, spondylitis, RSV and at least one genetically
predisposed orphan disease.

In the diagnostic area, our bovine products are used in the screening of
U.S. blood for key viruses, such as HIV and HCV, for screening of cancer
markers and in serological testing as a potentiator of blood typing
prior to transfusions. In medical devices, our bovine products are used
in the production of a medical device that is used during surgery, and
in animal vaccine, our products are used for the cultivation of
Leptospira, which is used to produce animal vaccines for the treatment
of Leptospirosis, which is a worldwide problem in livestock. Also, these
bovine products are used pervasively in life science research as
reagents in the lab for protein assays and other lab assays like
chromatography and electrophoresis.

Next, please. So in April of 2000, we received an update, which was
issued to manufacturers of biological products from Kathryn Zoon, the
former director of CBER, that essentially says avoid using ruminant
origin products derived from BSE-affected countries in the production of
FDA-regulated products that are intended for humans.

Make sure you identify all the ruminant materials used in production of
the regulated products, and document the country of origin, and make
sure you maintain traceability records for each lot. Of course, the
purpose of this guidance is to minimize the TSE threat to the public.

Next, please. So this creates some regulatory uncertainty regarding the
products under development today that are made using bovine ingredients.
Also, there is a risk of current production of approved drugs, which use
bovine ingredients. This may lead to a possible interruption to the
supply of these biotech drugs if BSE occurs in the U.S. Bovine-based
products provide unparalleled performance. There has been a few attempts
to replace these products in cell culture, but they typically result in
lower productivity and higher costs.

Next, please. I would like to switch gears for a minute and talk about
the prion infectivity clearance studies that Serologicals has conducted.
We scaled down our manufacturing processes and we used a hamster-adapted
strain of scrapie agent as a model to emulate the BSE. Like many
presentations before, we used a 263K agent. We spiked known titres of
infectivity prior to key process steps, and using serum tenfold
dilutions we titrated the infectivity downstream to measure the ability
of the intervening steps to reduce infectivity, looking at that in-vivo
using hamsters and looking at the clinical signs, abnormal gait,
tremors, ataxia or incoordination, also looking at a histopathological
examination of the brain tissue to confirm the clinical diagnosis, and
the characteristic protease resistance of the transformed prions.

The conclusion of these studies, if you look at our Bovine Serum Albumin
product line, our HS product line, looking at four process steps
achieving a total clearance of 16 log~10~, Bovine Aprotinin, a total of
17 logs and EX-CYTE completing one manufacturing step to date, achieving
3.7 log~10~.

Next, please. So we feel that these prion clearance studies offer some
objective evidence that these products are very safe from a TSE risk. In
summary, the high safety and quality of our bovine-based products are
summarized by the following.

One, many are manufactured from bovine blood, which is recognized as
being low-risk of TSE infectivity, according to the World Health
Organization and the EC. Moreover, Serologicals use either plasma or
serum for added safety. One of the theories is that it is believed that
prions may reside in the cellular fraction of blood, for example,
leucocytes. We only use bovines that are less than 30 months of age, and
they are typically less than 20 months of age.

According to the DEFRA statistics in Great Britain, no BSE reported in
cattle that is less than 20 months. It is uncommon for it to happen in
less than 30 months. We use only USDA-approved raw materials collected
in USDA-licensed establishments. All these products are manufactured
within an ISO 9002 registered GMP environment.

We have completed and published prion and viral clearance studies, and
this compounded with the similar clearance studies that our customers
have completed that are producing these biologics, we also maintain EDQM
certificates of suitability, which again is an assessment of low TSE
risk. We have a proven track record of safety in global health care.

Next, please. So this is an example of one of the viral clearance
studies that we have completed on our bovine product line. Due to time
constraints, I won\'t get into this other than to say that we have
demonstrated more that 6 log~10~ of bovine viruses.

Next, please. So in summary, one, bovine-based products are critical to
the production of life-saving health care products. Secondly,
manufacturers of FDA-regulated products cannot replace bovine
ingredients quickly, easily or economically. The high safety and quality
of Serologicals\' products is supported by the low TSE risk raw
materials that we use, the controlled production and the research
studies that we have conducted that demonstrates robust virus and prion
clearance ability of the manufacturing process, and our track record of
safety and success. We\'re pleased to work with the TSE Advisory
Committee to further develop the TSE risk guidelines covering these
important products to permit their continued use.

Next, please. Some considerations. In addition to using low TSE risk raw
materials, why not recognize the value of prion clearance studies and
let\'s establish minimum acceptance criteria. Let\'s have suppliers
perform prion clearance studies to provide objective evidence supporting
the product safety. Also, why not consider prohibiting the sourcing from
countries with a high incidence of BSE, rather than just one or two
cases?

And finally, when it comes to setting policy, we would request that the
FDA and the USDA carefully weigh the impact to the end consumers, i.e.,
the patients, producers of biomedical products, which are our customers
and supply chain producers like Serologicals.

Next, please. Finally, we would like to leave you with two contacts at
Serologicals Corporation. If any of you wish to discuss this further, we
would be happy to do that. Our email addresses are listed. Thank you
very much.

SECRETARY FREAS: Thank you for your comments. Our next speaker for this
open public presentation is Dr. Merlyn Sayers.

DR. SAYERS: Excuse me. Thanks for this opportunity to talk to you. See
if you can rustle up my first illustration. It\'s a brave blood bank of
the talks in the immediate shadow of the regulators, so I am indebted to
Dr. Hills and to Mr. Vaz for giving me some narrative separation from
Dr. Epstein.

No, let\'s have the earlier slide, please. I\'m speaking to you as the
CEO of Carter BloodCare, and that\'s the community independent blood
program that meets the transfusion needs of something like 150 hospitals
in the Dallas, Fort Worth area. I am also speaking to you as a former
chairman of the Blood Products Advisory Committee and as a consultant to
this group, and I only make those two comments to emphasize how keenly I
appreciate the challenges that the regulators confront and also advisory
groups like yours have to confront, as well.

By way of a preface, let me have the first illustration. Something like
40,000 Americans donate every day and their health history, their
screening for markers of diseases that are potentially transmissible by
transfusion, their subsequent counseling if that counseling is
indicated, these elements constitute what is perhaps the largest public
health exercise in the country and might even be the largest public
health exercise in the world.

Now, bear in mind that those 40,000 individuals that donate originally
were some 50,000 individuals, close to 50,000. Of course, many get
deferred during the history and examination even before any serological
testing is done on those folk. So what happens at a local level?

At Carter BloodCare, at our blood program, last year we registered
something like 270,000 individuals, 40,000 were deferred and they were
deferred either permanently or temporarily, as I say, even before
testing. The majority of these deferrals certainly are temporary
deferrals, and they are attributable to medications that those
individuals might have been on. They might have a low hematocrit. They
might have traveled to a malaria area.

But those temporarily deferred individuals are of particular interest to
us as blood bankers, because potentially those individuals are
individuals that we could get back to continue their donation to the
community\'s needs. We have been considering what has happened to
temporarily deferred blood donors for a long period of time.

May I have the next illustration, please? What is their subsequent
conduct? We looked to 500 temporarily deferred donors and followed them
for two years, and you can see from this illustration that 58.5 percent
returned to donate successfully. 8 percent returned only to be deferred
again, but 33.5 percent of that original starting temporarily deferred
group did not return.

So for one third of temporarily deferred donors, that temporary deferral
is so discouraging an experience that those individuals resist all our
entreaties for them to come back and donate again. Now, the likelihood
of deferral is obviously proportional to the amount of scrutiny that
these individuals are subjected to.

So let\'s have the next illustration. And certainly, the amount of
scrutiny that donors are being subjected to has increased dramatically.
You can see the number of questions that donors were asked in 1988 and
the number of questions donors are asked in 2003. For those of you that
are donors, the donor history questionnaire does not list 160 separate
questions. It\'s probably closer to 40, but each question has become so
complex. There are multiple questions. There are questions within
questions. There are nested questions. So what we now want are 160
pieces of information from donors.

So where do these increasing scrutinies relate to our consideration for
deferring donors who have some geographic risk and our need to
potentially exclude them from the donor base? Let\'s show in the next
illustration. The number of donors that we have deferred since the year
2000 now that we have introduced additional scrutiny with regard to
deferral for attempts to decrease the likelihood of transfusion
transmitted CJD.

You can see that at our blood program in the Dallas, Fort Worth area, we
have now deferred something like 3,500 donors, and this sad tally is a
significant underestimate of the actual number of donors that have been
deferred, and it is an underestimate, because many of these donors are
individuals who have paid attention, taken heed of our broadcasts, our
announcements, our publications urging them about the new restrictions.
We have no idea of what that number is. This number reflects only those
individuals who fail to appreciate the new restrictions that we are
publishing and who came to donate anyway.

So what sort of contributions might these individuals have made? Let\'s
have the next illustration. The next illustration. Thanks. How many
donations have these 3,500 individuals made? Well, they have made
something like 13,000 previous donations, and these are individuals.
These citizens are now people who are indefinitely deferred. They have
obviously made important contributions to the community blood program,
and it is interesting that in that conflict of interest questionnaire
that was handed out this morning, you were asked if you regarded it as
important that citizens affected by decisions are directly involved in
the Advisory Committee process. And certainly, I believe that citizens,
in this case donors, are important and should be involved, but they
frequently do not get that opportunity.

If you were to ask them how they respond to their temporary or permanent
deferral, let me show you some of the questions that have been posed to
us during counseling sessions when we have spoken to individuals
deferred as a result of some geographic exclusion.

Let\'s have the next illustration. This is what we get posed. What
should I tell my wife, my husband, children, my dentist? What should I
tell my family practitioner? Where can I get tested? Where can I get
treated? Will this affect my medical insurance, my disability insurance,
my life insurance? Will this count as a preexisting condition? Should I
reconsider having a family?

Let\'s have the next illustration. Why do the criteria change? Other
donors have asked questions along these lines. So if I had been in the
UK for one day short of three months, I would be safe? If you\'re
telling me that I can\'t donate anymore, what are you telling patients
who got my blood? Why didn\'t I hear about this from the military? How
many patients have got this disease from my blood? How many patients
have got this disease from a blood transfusion anywhere?

Now, these are tough questions and deferral criteria can be debated in
the relatively academic climate of these meetings, but justifications
for deferral that are acceptable here do not sit well when they are
explained to the donor community at a lay level. The donor deferral
process is essentially a contributor and an important contributor to all
those layers of transfusion safety that we recognize as being valuable,
but the process is also responsible for increasing numbers of former
numbers whose experience is marked by a sense of frustration and
alienation.

And this next illustration shows what has happened to the rate of
permanent deferrals amongst blood donors in our community. You can see
that between 1999 and 2003, there has been a threefold increase in the
risk of permanent deferral of individuals in the community. So just as
we are concerned about individuals who are temporarily lost and our
difficulty with getting them back, we are concerned, too, about the fact
that the rate of permanent deferrals significantly contributed to
geographic exclusion is increasing so dramatically.

So in closing then, a few comments. For many deferred donors, there is a
credibility gap that our explanations just do not bridge. And to avoid
the risk of transfusion safety being achieved at the expense of
availability of blood for transfusion, CBER\'s Blood Action Plan that
was promulgated in \'97, which addresses increasing the blood supply and
removing restrictions to safe donation really needs our enthusiastic
support and endorsement.

And then lastly, and this sincerely is not meant as a plea for less
safety or a plea for less regulation, let me just say that it is easy to
add eligibility restrictions, and there are many recent examples,
smallpox, SARS, West Nile Virus, but the more difficult task of lifting
restrictions that no longer serve a purpose is a task that also needs to
be addressed. Thanks.

SECRETARY FREAS: Thank you, Dr. Sayers. Is there anyone else in the
audience, at this time, who would like to address the Committee on this
topic before the Committee?

MR. FILLBURN: Charles Fillburn, Nutramax Laboratories. I would like to
ask Dr. Hills, does he exclude the possibility that the lone BSE animal
that was observed is not due to a mutation? Have you sequenced the gene?

SECRETARY FREAS: Could you come to a microphone, so our transcriber can
record the comments, please?

DR. HILLS: Bob Hills. Yes, we did look at the spontaneous possibility,
in other words the mutation of PrP gene. What I can say is that we did
look at it. We have excluded it right now and for other reasons, I
really can\'t comment. There are some publications coming out shortly
with respect to that.

MR. FILLBURN: Do you think it\'s possible that it could arise again
either here or in the United States simply as a consequence of mutation?

DR. HILLS: I \--

MR. FILLBURN: The reason I ask is we seem to assume that the only way
this can occur is through feed. If that\'s not the case, then we need to
be more aware that some of these restrictions that we\'re putting on may
be overkill.

DR. HILLS: Well, I think there are ways that we can look at the PrP gene
to determine whether or not it is spontaneous mutation based on that.
Now, whether or not you can determine whether or not you have sufficient
testing power to ensure that that one case you found is spontaneous or
not, I don\'t know.

MR. FILLBURN: Okay. I would just like to echo the comments of Mr. Vaz
that in how we react to the situation in Canada and our importation of
any products really has a dramatic \-- can have a dramatic effect on
health care in the United States and it may be an extreme overkill. I
would like to see the USDA and the FDA be on the same page in how they
try to treat this, that they be more realistic about it, and demand more
clearance work by processors who may be dealing with these types of
products.

SECRETARY FREAS: Thank you for your comments. Do you have a quick
comment?

MR. HAFFENDEN: I\'ll try to keep it really quick. I would like to echo
the same comments, the same that was expressed by Mr. Vaz. We do
collect, have up until May 24^th^ collected Canadian origin
animal-derived blood products that are sold into the veterinary and
pharmaceutical industry. We have collections in Australia and in the
United States. These are critical supply raw materials, and I believe
that we do need harmony between USDA and FDA on guidance. We also do
have an isolated herd, isolated BSE-free herd in Canada and would like
to volunteer to participate in a committee than can help to set those
guidelines and give some examples of what is really there.

I understand why the USDA in particular and the FDA have to react
quickly and close doors, but I believe we need to put the resources
shortly to analyzing products on a product by product basis, not a
global product entity, and reopen those doors for products that are
needed and critical.

SECRETARY FREAS: Thank you for your comments. There will be two more
open public hearings tomorrow, and at that time, we will be more than
glad to welcome your comments.

CHAIR PRIOLA: This topic is open for discussion by the Committee if
anyone would like to make a comment or have any additional questions. I
know I have one question that I actually forgot to ask Dr. Hills. You
said you ruled out the possibility of this case having originating in
Saskatchewan or no, sorry, excuse me.

As a consequence of exposure to CWD or scrapie, as well as as a
consequence of a spontaneous event, did you do that based on purely
pathological assessment or how did you come to the conclusion that this
was not a case of a cow coming into contact with a CWD infected deer or
elk, a scrapie infected sheep, especially since Saskatchewan is where
the CWD is, right?

DR. HILLS: Yes. That actually was a concern, which is why what we did
when we sent the sample over to Weybridge, we actually were asking them
to look specifically at the strain that they had in hand and to compare
that with the strains that they had and they saw no differences.

DR. ROGERS: Ron Rogers, a little bit more.

CHAIR PRIOLA: Okay.

DR. ROGERS: I just wanted to say that some samples have been sent over
to a stacks group in the UK, and like they are doing a differentiation
on the glyco-civilization patterns, and so the CWD \-- we had already
previously been involved with them in some research activities to look
at CWD profiling, I guess you have to say at this stage. So this did
have some positive material over there already, and so this material
also was brought over to sort of see if, in fact, we can get those kinds
of patterns.

CHAIR PRIOLA: Okay.

DR. ROGERS: So it\'s purely at the research level that this was ruled
out.

CHAIR PRIOLA: So basically, right now, pathological assessment, and
you\'re doing the molecular assessment of the PrP. Okay. Any other
questions or comments? Yes, Shirley?

MS. WALKER: I find that it is interesting that Canada is now moving to
provide the restriction to add SRMs after reviewing their new case, and
we have just been asked to look at that language in our present policy,
so we might be cognitive of looking at and changing our policy too
quickly.

CHAIR PRIOLA: Okay. If there are no other questions or comments from the
Committee, then I think we can move on to Topics 3 and 4, so the
remainder of the afternoon is going to be a general introduction to TSEs
and decontamination of medical equipment and facilities. The first
speaker will be Dr. David Asher from the FDA.

DR. ASHER: Thank you. Well, it\'s a great pleasure to open this session
on decontamination of TSE agents, which has been developed jointly by
the FDA Centers for Biologics and Devices. This topic is presented, next
slide, please, in response to a request from this Committee last year
for more specific information before members felt comfortable advising
the Agency concerning appropriate decontamination of tissue
establishments where the TSE agents might be encountered.

Next slide, please. FDA, of course, as part of its mission, is
responsible for helping industry to keep regulated products safe and
that includes keeping products free of pathogens, today\'s pathogen of
interest, of course, the TSE agent. Properties of the TSE agents
complicate those efforts. Thank you. You have already heard some
discussion about the context-dependency of inactivation of TSE agents. I
expect you will hear more.

Scrapie, of course, scrapie agent has not been completely inactivated
after exposure or after drying and then exposure to steam heat.
Fortunately, TSE agents are substantially inactivated in solutions of
sodium hydroxide, sodium hypochlorite and probably other chemicals.
Hence, the World Health Organization consultants have recommended
decontamination in health care environments using combined sodium
hydroxide or sodium hypochlorite and moist heat. Some other authorities
have doubted the need for such harsh chemical treatments, and we would
like the Committee today to consider these different points of view.

The situations for which we are soliciting advice today are for products
regulated by the Center for Devices, that is instruments and by the
Center for Biologics Instruments and Surfaces used in the production of
tissue products and plasma derivatives.

There are, of course, somewhat similar though not identical situations
that would involve other centers, our Center for Drugs is particularly
interested in today\'s discussion, because, of course, some drugs have
components of human or animal origin. We are aware that some of the
problems involved in the production of food or feeds are similar, but
the contexts are really quite different, for example, of course, sodium
hydroxide poses certain problems in proximity to food products.

We would certainly logically expect that some issues of the U.S.
Department of Agriculture, which regulates animal slaughter and meat
production in interstate commerce and the Environmental Protection
Agency, which regulates water affluence, including affluence from
autopsy rooms, that they would have relative issues, but this Committee
is not advisory to those agencies, and we do not solicit advice for
those problems.

Next slide, please. There is no question that contamination of classes
of products regulated by the Food and Drug Administration have
transmitted Creutzfeldt-Jakob Disease to human beings, fortunately few,
relatively few such cases have been recognized in the United States.

Next. One such class of contaminated products is reusable surgical
instruments of which a contaminated cortical electrode shown here in
pieces is the best known example.

Next slide, please. I am aware of only six cases summarized here in
which transmission of CJD has been plausibly attributable to
contaminated surgical instruments, and I would note that in none of
those was modern cleaning or steam water used to decontaminate the
instruments involved.

In addition, at least two epidemiological studies of which I am aware
have claimed to demonstrate some association with previous surgery,
though most studies have not found that and the association is not
particularly impressive.

Next slide, please. You have heard that inactivation by heating of
scrapie agent is very much context-dependent. This is a slide of data
from Bob Rohwer some 20 years ago demonstrating that scrapie infectivity
in aqueous suspension was reduced to the level of detection in less than
five minutes at 121 celsius. That was carefully suspended in aqueous
solution. Although, at boiling temperatures, although, there was also
prompt reduction in infectivity, a reduction, a resistant fraction,
that\'s the term of art for Dr. Johnson, a resistant fraction remained.

Next slide, please. But when dried onto surfaces, infectivity was
readily detected even after autoclaving for an hour at 134 degrees
celsius.

Next slide, please. Dr. Rohwer, who will be our second speaker in this
session, has confirmed that apparent\-- or rather, Dr. Taylor who will
follow Dr. Rohwer has confirmed that apparent paradox, and found that
different strains of TSE agent in similar preparations, at least,
appeared to have differences in thermal stability. The conclusion of
those studies were that in worst case scenarios, autoclaving has not
been validated to decontaminate all TSE agents completely.

Next slide. A number of factors listed here must be considered in
deciding whether there is a significant risk that a contaminated
instrument might transmit Creutzfeldt-Jakob Disease, including
infectivity of the contaminating material, the reduction in activity
achieved by cleaning and decontamination, and the root by which a
susceptible individual is exposed and Martha O\'Lone will talk more
about those things tomorrow. Our speakers later this afternoon and
tomorrow will address those and other issues.

Next slide, please. Central nervous system tissues for humans as for
animals have been consistently demonstrated to be infectious when
assayed in susceptible animals.

Next slide. But there is also \-- next slide, please. There is also a
significant though smaller likelihood that tissues of patients with
Creutzfeldt-Jakob Disease outside the central nervous system, and
that\'s not just Variant Creutzfeldt-Jakob Disease, that is typical
sporadic Creutzfeldt-Jakob Disease, will have some infectivity found and
here are positive tissues listed.

Next slide, please. Of course, fortunately, most human tissues, fluids,
excreta have never been found to be infectious.

Next slide. Although, confidence in the negative results is somewhat
tempered because of the very small number of samples studied, and the
fact that the assays used were animal transmissions, and there does
appear to be some species barrier even between human beings and
primates, which might raise the level of the limit of detection.

Next slide. Just a couple of examples of really how small the number of
tissues successfully studied has been.

Next slide, and the next slide. In human CNS tissues, the mean content
of infectivity measured in the NIH series was estimated to be about
10^5^ primate intracerebral lethal doses per gram, but note that one
brain was found to be infectious at a dilution of 10^-8^, and
considering both that and the species barrier, it might be prudent to
assume such high levels of infectivity for high-risk human tissues, at
least in an occasional patient.

Next slide. Because of the substantial uncertainties attendant to the
biology of the TSEs and the properties of the agent effects of cleaning
and decontamination, advice offered to public health authorities in the
United Kingdom and the USA concerning surgical instruments has not been
consistent, and I won\'t read these. But the UK CJD Incidence Panel has
advised incinerating instruments exposed to brains of patients with
known CJD where our respected authority, Bill Rutala, whose is going to
speak later, has felt that cleanable critical or even semi critical
devices in contact with high-risk tissues of CJD patients can be cleaned
and sterilized by autoclaving either at 134 celsius or 121 celsius,
etcetera. Our proponents of both points of view are present here today,
and we really would encourage a discussion of these conflicting points
or, at least, apparently conflicting points of view.

Next slide, please. The two FDA centers who developed today\'s program
have generally relied on advice concerning decontamination that came
from a consultation convened at the World Health Organization in 1999
published the following year. That consultation was chaired by Paul
Brown, who was then the chair of this Committee, and our next two
speakers, who are among the most influential of the participants, are
two of this Committee\'s former CDC members, both of whom are in
attendance today, were also in attendance at that meeting. The
consultation identified recipients of potentially contaminated products
as being the group of persons at the greatest risk of iatrogenic CJD.

Next slide, please. And they offered the following general advice. They
acknowledged that decontamination is context-dependent and that one
method may not be completely effective in all circumstances. Cleaning
facilitates decontamination using the best validated methods available,
essentially meaning based on actual pilot studies. And then they advised
using what we call an orthogonal strategy, that is using two different
methods, methods based on different physical chemical principles
whenever that is possible.

FDA staff might add that in choosing those orthogonal methods, that a
method that inactivates the agent is generally considered more reliable
than one that simply removes it, because when an agent is removed, there
is always the danger that it can be reintroduced back into the product
of interest.

Next slide, please. They recommended single-use to instruments,
destroying reusable instruments wherever possible, but they acknowledged
that there are obvious situations in which instruments simply cannot be
discarded, and that less effective methods than destruction may have to
be used.

Next slide, please. The consultation recommended a series of
decontamination methods in order of decreasing probable effectiveness.

Next slide. I expect that they recommended six such methods. I won\'t go
through them all here. I expect that they will be discussed by several
of the speakers later today and tomorrow, but note that the first four
all include the use of either sodium hydroxide or sodium hypochlorite
either with or followed by steam autoclaving.

Next slide. The last of the six was to autoclave 134 celsius 18 minutes
with the caveat that in worst case scenarios, that is where brain tissue
has been baked, dried under surfaces, the infectivity will be largely,
but not completely removed.

Next slide, please. And for surfaces or heat sensitive instruments, they
recommended sodium hydroxide or sodium hypochlorite at room temperature.

Next slide. A number of other treatments listed here were dismissed as
being inadequate.

Next slide. We tend to think of the decisions regarding effective
decontamination in these four general categories, and I suggest that it
might be useful if the Committee addressed them in this way, as well.
The surface of these situations are surfaces or instruments potentially
contaminated with either high-risk tissue or lower-risk tissue from a
subject with definite or probable TSE, and then the same categories for
someone where TSE is not suspected.

Next slide. We\'re fortunate to have with us today Bob Rohwer, who has
already spoken, David Taylor to review the general principles of TSE
decontamination and the basis for the WHO recommendations.
Unfortunately, Dr. Philippa Edwards of the UK, CJD Incidence Panel, is
ill, but she kindly emailed a talk for us, and that will be delivered by
Pedro Piccardo, who is an alumnus of this Committee and a most welcome
recent addition to our CBER staff, dealing with TSE issues.

Bill Rutala, who is consultant to the CDC on hospital infection control,
will share his extensive experience here in the USA. Ed Rau will then
report on interesting studies of incineration that he and Paul Brown
have been conducting. And finally, today we\'ll end with Stan Brown of
our Center for Devices, and I, who will report some early results of
models that we have been studying based in part on the work of Charles
Weissmann, who will speak to us tomorrow.

Next slide. Then tomorrow the issue for discussion will be presented for
CDRH by Lillian Gill, Martha O\'Lone and Charles Durfor, and for CBER by
Ruth Solomon and Dorothy Scott. Ellen Heck will review the needs of Eye
Bank. Christoph Kempf and Andrew Bailey will represent the Plasma
Proteins Therapeutics Association in discussing the needs of plasma
processes, including a study that they will propose.

Please, note that, again, although the discussions will doubtlessly be
of great interest to other agencies of the U.S. Government, and
especially to our Center for Drugs, we do not solicit advice for these
other agencies, only for FDA-regulated devices, tissue products and
blood products.

Last slide, please. With that, I hope that you find the program that we
have assembled informative, and we anticipate having useful comments
during our open public hearing, and discussions and votes by the
Committee. Thank you very much.

CHAIR PRIOLA: Okay. Thank you, Dr. Asher. Our next speaker will be Dr.
Bob Rohwer.

DR. ROHWER: Thank you, and let me begin with an apology to those people
who have heard this talk before. It is one that I have given to this
Committee actually in an earlier form of it in the past, and have given
it fairly frequently over the last few years, but I was asked to do it
again just because David felt that the place needed revisiting. And so
we\'ll begin with the first slide. The main points I\'m going to make in
this talk are that the susceptibility to inactivation of TSEs
infectivity is within the normal range for viruses and spores, but that
the TSE infectivity is resistant to disinfection or sterilization. That
may seem like incompatible statements, but I will try to show you what I
mean in just a minute.

The susceptibility to inactivation is an intrinsic property of the
agent, and this susceptibility is normal. That\'s what I\'m saying, but
the resistance is context-dependent and a property of the environment of
the infectivity.

Next slide. The best sources for this at the current time are these old
papers of mine in [Science and Nature]{.underline}, which have the
complete experiments behind the kinetics that I\'m going to show you
here and this review, which puts it into a larger context.

Next. And then the WHO report, which Dr. Asher just reviewed is also a
very excellent source.

Next. I am going to begin by just talking about the inactivation process
itself. And this is actual data taken from an activation process with
the scrapie agent. This is was a hypochlorite experiment, I believe. I
have forgotten now. I put this together so long ago, but the main points
are the following.

If I can have the next. We\'re going to build this slide as we go
through it. The inactivation process, one way to think about this is if
we think of this is surviving fraction on this axis over here where we
start with 100 percent survival, no killing. At 10^-1^ survival, we have
destroyed 90 percent of the population.

Next slide. So for example here, I mean, here we\'re starting. If we
convert this to a 100 individuals, what we have got by the time we\'re
here on this inactivation curve is only 10 percent of the population
left. We have killed 90 percent of the population in these very first
moments of exposure, and by the time we get to the second log of
inactivation, we\'re down to one out of 100 survivors. This is just by
way of review of things that you probably had in your elementary
chemistry class, but we sometimes forget this with time.

Next. And then if we look down here, we notice that 90 percent of the
kill occurs during this first tiny interval. Only 10 percent of the kill
occurs during this next interval. Only 1 percent during this, a 10^th^
of a percent, a 100^th^ of a percent during this interval right here.
We\'re getting less and less effect as we go along. The vast majority of
what is happening to this population is occurring right here in the very
first moments of exposure.

Next. And this is reflected by this component of the inactivation, which
is reflected by this line right here, and the inactivation rate constant
for this line is the inactivation rate constant, which is defining the
behavior of the vast majority of the individuals in this population.

Next. Next. This line describes a second component, and it is
describing, next, a much smaller proportion of the population. About one
in 10,000 of the individuals behaves like this.

Next. So what is going on here? We have susceptibility to inactivation
as defined by this initial rate of inactivation, is intrinsic to the
agent. It is actually less complex and there are fewer controlling
parameters. Whereas, over here, this population that is being
inactivated at a different rate could be gaining those properties in a
number of context-dependent ways, and these are different for each
environment and they are much more complex.

It could be due to the container. It could be a factor. I mean, it could
be any combination of these factors, as well, but among the things that
we have to consider are the container, rough or smooth surfaces,
reactive surfaces, porous surfaces, cofactors like fats, proteins,
oxidants, reductants, water, air, in the case of autoclaving, buffers,
pH, temperature can all affect the inactivation suspension, whether what
the source of the tissue is and its composition, the procedure for
making it, how it was homogenized, the dose rate, various transfers.

And then we have these kind of procedural problems with making these
measurements themselves, which are the accuracy limitations of the assay
and its reproduce-ability. This is a real issue in the case of end point
dilution titration of TSE infectivity, which is only good to about .3
logs. And cross-contamination is an issue, and it becomes an especially
important issue when we\'re talking about very low levels of survival at
the very end of these inactivation curves.

How do we know that this survival isn\'t due to something that got
transferred from here, which has almost 100,000 times more infectivity
in it? The way we know is we\'re very careful when we do these
experiments, but it\'s something that you have to be very careful about.

Next. Next, please, and next. Let\'s go to the next slide. Click through
to the next slide. Thanks. In comparing agent properties, the properties
that are intrinsic to the agent are reflected in the initial rate of
inactivation. That is when the vast majority is being inactivated, and
the interpretation is less complex. The size of the residual fraction is
a complex function of environmental parameters, and cannot be used to
compare the intrinsic sensitivities of agent strains.

This is where I differ with Robert\'s perspective that he gave this
morning. In other words, I consider these plateaus to be a very
important public health and agricultural problem in terms of infection
control, but they are not telling us that much about the intrinsic
properties of the agent. They are telling us about the context of the
agent. They are telling us something about the context of the agent, and
it has been very difficult for us to figure out what that is.

Next. So let\'s consider an example from chemical inactivation.

Next. This is a hypochlorite inactivation in which we can see that. In
the scrapie curve right here, we have \-- this is surviving fraction,
the same kind of curve I just showed you minutes ago down here, that on
contact with hypochlorite, .5 percent, this is a normal concentration,
which bleaches use, and we get an initial very rapid killing down to
about 3 logs, but then we hit a plateau and there is at the level of 1
part per 1,000 or a 10^th^ of a percent, we have got something in this
population that is protected from hypochlorite. It\'s not seeing it.

By the way, we checked. The available chlorine did not change
significantly over the course of this infection. We did this same
experiment with some controlled viruses. These are bacteria phages. They
are non-involute viruses. They are very easy to assay and they are
reasonably robust in some ways and not others. But here is PhiX 174
showing the exact same phenomenon. It plateaus at a lower level. FD and
M13 like phages doing the same thing over here.

Here are these two. These two viruses were spiked into the same kind of
normal brain homogenate that the scrapie brain was in, and they
exhibited this behavior in a purified form in PDS. They were inactivated
to the limit of detection almost instantly on contact with bleach.
Another example of context.

Next. Next, please. Well, two of the things that we are going to discuss
here are the things that work best for TSE agents, and bleach is one of
them, and I hope that David will be sharing. He has a lot more data on
bleach than I do, and I hope he will be sharing that with us. But sodium
hydroxide is something that I have been pursuing for a long time, and
this was an experiment a long time ago with Paul Brown, one of our
initial experiments, comparing CJD and 263 scrapie.

Again, in a very highly dispersed 10 percent brain homogenate of these
two infectious agents, and adding sodium hydroxide at these
concentrations, and I would just ask you to concentrate on these first
three lines here. At 60 minutes with one normal, we had limited
detection killing here and here for both CJD and scrapie. By 15 minutes,
we had almost as much inactivation. A 10^th^ normal did almost as well
as one normal. It\'s a very effective method.

On the other hand, next slide, please, this is a table of \-- it\'s now
out of date. There are more experiments would could be put on here, but
at the time I made this, these were all the sodium hydroxide experiments
that were in the literature, and we got very, very good inactivation by
sodium hydroxide, but there are examples here. These are the experiments
that we had done, at that time, but there are examples here where there
is some activity left after considerable amount of exposure, and that
always was very puzzling to me, but my guess is that it has to do with
how the stuff is presented and homogenized.

Next. I went back and revisited. We have revisited this subject with a
kinetic experiment on sodium hydroxide, which is presented here. And in
this case, the black circles indicate infectivity, and this is time of
exposure, and we\'re seeing something that is very similar to the sodium
hypochlorite effect, except much more dramatic even. We\'re getting a
huge reduction on contact, essentially, with sodium hydroxide. This is
the point that was taken in the shortest amount of time, interval, that
we could effect between adding the sodium hydroxide and then adding the
acid to neutralize it, and then taking the points. So it\'s right around
30 seconds. This is two minutes, etcetera.

But on this same curve, I have got two other plots. One is a plot, which
I am labeling denaturation in one hydrolysis. And what do I mean by
that? Well, we went back later and used a Western Blot on these samples
to see whether we could recover Western Blot signal or not from these
various fractions. And the Western Blot, especially at the time that we
did this, was not as sensitive as the infectivity assay, so we couldn\'t
detect it over as long a range, but it was very clear that upon contact
with sodium hydroxide, we destroyed the pk resistance of PrP. It was
gone. It was showing the same inactivation kinetics, essentially, as the
infectivity.

Whereas, if we didn\'t pk digest and we just put the stuff on the gel to
see whether there was anything left, it also disappeared, and this is a
disappearance by hydrolysis. The protein is being hydrolyzed. It no
longer shows up on the gel, and it is showing quite different kinetics.
So one of the points that we can take home from this is that to the
extent that infectivity and PrP are related and the prion protein are
related, and I am not entirely convinced of that, but nevertheless, to
the extent that they are, it\'s denaturation that is the correlate with
inactivation of infectivity not hydrolysis. This is basically good news,
because it\'s much easier to denature something than hydrolyze it.

Next. Heat inactivation will be the next topic.

Next. This is a \-- I have just taken the 121 degree autoclave
experiment out of that family of curves that Dave just showed you and
that I showed you earlier in the day, because it makes the points best
in my opinion. Here is a case where, at the time we did this experiment,
the story was that you couldn\'t kill this stuff with autoclaving, you
know, that 121 degrees was not sufficient to destroy the infectivity
from 263K hamster scrapie.

This is a kinetic experiment showing that, and this was done not in an
autoclave. It was done in an oil bath. The samples were sealed in
ampoules. They were plunged into the oil bath, so that we could control
their \-- and I was using thermistors, at that time. The temperature was
being recorded, so I knew when they got to 121 degrees. I knew what the
ramp time was. I had that on my recorder, and we could control the
actual time of exposure within very narrow limits.

So this first point on this curve was taken after the 58 second ramp
time to 121 degrees, so it had just got there. By the time it got to 121
degrees, we had already destroyed 99.9999 percent of the infectivity in
that sample. On the other hand, it took another 10 or 15 minutes to get
to the limit of detection of the infectivity. There was a residual
population that took longer.

And this is a concern, and this was a very highly dispersed sample again
of infectivity, and you get quite a different story, next, please, if
you do this type of experiment. Now, I think David is going to be
showing a lot more of this in a few minutes, but later on, David Taylor
started doing these experiments using brain macerates. Now, this is not
a homogenate. This is a mush of brain. It is not dispersed in fluid.
It\'s a paste, basically, and it is being exposed at these various
temperatures in the autoclave, and this is the untreated sample, and
these samples are getting incomplete killing even after these very
extreme treatments. I mean, this is quite extreme for steam
inactivation.

Well, you definitely have to say that this poses \-- you wouldn\'t want
this on your scissors when they go back into the next patient, for
example. This is an extremely important public health result. On the
other hand, what does it tell us about the agent in what we\'re dealing
with? Well, you can get a titre out of this, because you\'re at limiting
dilution here, and we can do something we call a parson, we get a parson
titre out of this type of sample. This is how we make our measurements
in low titre blood samples. And I have done this on the next slide and
just put these figures on next. This is where these samples would fall
on this curve that I just showed you. There has still been an extremely
high level of inactivation associated with these, but you have got
survival going all the way out to 134 degrees here, at 134 all the way
out to 60 minutes for some of these samples that were done in these
macerates.

Personally, I think we\'re talking about the same story here. It is just
a matter of what we\'re talking about, and the context has been ramped
up in the case of macerates versus homogenates, and the survival lingers
for longer periods of time.

Next. Next, please. So what are we dealing with here? These could be
intrinsic differences, and that was a question that came earlier in the
day from Dr. Bailar and it\'s a legitimate one. I think it needs more
study. A lot of us have this on our books. Robert said he has been
planning to do this. I have been doing it. I have got these samples in
the freezer. I just have not gone back and redone this experiment,
redone the kinetics on these, but it is on the books. Someday, it will
get done.

But when you talk about these heritable differences, the point that I
want to emphasize is that I feel that they have to be discussed on the
basis of inactivation rate, not residual infectivity. And my own
prejudice is that the rates will be exactly the same, because what
we\'re dealing with here is context, not intrinsic differences.

Aggregation is another issue. This is something that I was very
interested in early on in my career, but I think we have this under
control at the moment with the way we are homogenizing and dispersing
things, and aggregation would give you a recognizable difference in the
inactivation kinetics. It would not look like first order. It would be
first order with a delay. There would be a delay in something like that
contributing to that.

The most likely reason for this, in my opinion, is compartmentalization.
The inactivant is not actually reaching the infectivity, and our
challenge before us is to find ways to open and destroy this compartment
to get at the infectivity.

Next. I just have a couple more here. So if we compare these two moduses
of investigation, what we are using is 10 percent homogenate sonicated
highly dispersed versus whole brain macerate. This is sealed in a serum
bottle. I can\'t remember, David. Do I have this wrong? I think David
will correct me if I have got this not exactly correct here on how he
has got these set up. We are using an oil bath versus an autoclave. Our
samples were being constantly stirred while we were inactivating them
versus static. And, in fact, this is kind of an idealized type of
inactivation to get at the properties of the phenomenon. Whereas, this
is a worst case scenario, which gets at the worst case problems that
might be confronted in the public health or agricultural context.

Next. Okay. Now, we had some dry heat data earlier in the day from
Robert Somerville, so I\'m not going to go over this, except to say that
if you dry this material onto a surface, the inactivation properties
become completely different. It becomes much, much more resistant to
inactivation. However, this isn\'t a completely unfamiliar phenomenon.
It happens with spores and it happens with other microbes, as well.

Next. And so, in fact, my own interpretation of this in a nutshell is
that what is happening in these experiments and where the source of
residual infectivity may be coming from in our ampoule type of
experiments is that as we stick our ampoule into the oil bath, it boils
and flashes off immediately, and we throw things up on the walls and
they dry. We get little specks drying on the walls. I was very religious
about trying to recover everything when I went back to reanalyze this
material, so I scraped the walls and got everything back into the test
tube.

And what if what is happening is we have the infectivity in a form in
which it is basically anhydrous. We have little drips and drops here
that end up in little droplets of fat. Fat when it is oxidized becomes a
varnish, which is, essentially, a plastic. And so, basically, what
we\'re subjecting this to is a dry heat sterilization at the rate of
parts per million in our case. It\'s not something that\'s happening
very often, but we create a dry heat environment for a very small part
of this infectivity, and that is what is escaping. If the reagent can\'t
kill it, if you can\'t reach it, you can\'t kill it.

The other example I like to give is that if you put brain homogenate in
a Zip Lock bag and throw it into one normal sodium hydroxide, nothing
will happen to that either. And so it has to be available.

Next. Next, please. Next. Not that one. That\'s not supposed to be
there. So the point I want to make here is that 132 degrees uses a
significantly higher temperature than 121 for steam sterilization where
the inactivation takes place in minutes or even seconds, but 132 degrees
is only incrementally more effective than a 121 degree centigrade
environment for dry heat sterilization where the inactivation takes
hours to days at those temperatures depending on what you\'re talking
about.

So this does form, I think, a rationalization for what we\'re seeing in
this situation, and it also tells us \-- and this was the
rationalization for trying to remove all headspace from those devices in
which we did the gelatin inactivations that I showed you this morning.
We didn\'t want any opportunity, any place for drying to occur.

Next. I think there is just two more. Steam sterilization, the agent is
not intrinsically resistant to steam sterilization. There are problems
with delivery.

Next. And for effective delivery, we recommend surfactants,
homogenization, high levels of dispersion, eliminate sanctuaries,
agitation is helpful. My guess is that a refinant will also reduce the
potential for protective associations and will improve the ability to
inactivate.

Next. Prevent drying, immerse in water prior to enduring steam
sterilization and combine two or more methods. And the processing
details can be critical. Adhere closely to validated approaches, and
this is referring to this stainless steel result we\'ll hear more about
tomorrow.

And where we should go with this. We need to know more about the
underlying principles of resistance, and we definitely need more robust
methods for sterilization, which will actually get at these last little
bits of infectivity.

Why don\'t you end right there, and let me just end by saying that the
way we inactivate in the laboratory, our own instruments, is for
stainless steel and things that can take it and things that are recycled
and go back into animals, immediately after use they go into one normal
sodium hydroxide. They are immersed in one normal sodium hydroxide for
at least an hour, and then if they can take it, they are put through the
autoclave under one normal sodium hydroxide. They are cleaned after
decontamination under those conditions, and then they are reprocessed in
sterile packs back into the facility for further use.

CHAIR PRIOLA: Okay. Thank you very much, Dr. Rohwer. Are there any
questions before we move on to Dr. Taylor? Okay. If there are none,
we\'ll go on. Oh, I\'m sorry. David, go ahead.

DR. ASHER: Can you comment on aluminum vessels, please?

DR. ROHWER: I didn\'t hear that.

DR. ASHER: Aluminum vessels.

DR. ROHWER: I still didn\'t hear it.

DR. ASHER: Can you comment on the use of aluminum vessels?

DR. ROHWER: Oh, yes, right. We use a lot of sodium hydroxide in our
environment, and we learned early on that you don\'t mix sodium
hydroxide with aluminum. And, in fact, aluminum and sodium hydroxide in
an autoclave can explode and can be quite dangerous, so you have to be
very careful about that. So we, essentially, have no aluminum in our
BL3.

CHAIR PRIOLA: I think I\'ll check our BL3. I\'m not sure if we have
aluminum. Dr. Taylor, if you would? Our next speaker is Dr. Taylor, and
he is going to talk about decontamination of TSE agents and the WHO
recommendations.

DR. TAYLOR: Thank you very much. Well, thank you for the invitation to
speak this afternoon. As you can see, coming from the UK, I\'m using
thumb roll technologies, slides and overheads. I was warned there could
be problems with the electronic system, so I didn\'t bother with the
front-line. I just brought the backup.

As has already been discussed and as this group will appreciate, there
has been accumulating evidence over decades that TSE type agents are
remarkably resistant to a wide variety of decontamination methods, which
are quite effective with conventional microorganisms. This does not mean
to say that these methods have no effect, but rather that they are
impractical for usage in medical settings, etcetera. These include
things like strong oxidizing agents, phenolic disinfectants and even
ionizing radiation.

Because of this general resistance, there have been some known examples
of iatrogenic transmission where instruments or devices that were in
contact with the brains of CJD infected individuals went on to cause
accidental transmission in subsequent patients despite having been
processed in some fashion or another.

Now, I use the phrase in some fashion or another advisably, because the
methods that were used would not, in fact, be used nowadays, but David
Asher showed you the x-ray of implantation electrodes, which would be
put into a marmoset to look for infectivity after this was suspected of
causing this disease in humans through accidental transmission. In this
case, the electrodes were washed in benzine and in a well meaning
exercise to try and sterilize them, they were then exposed to alcohol
and formaldehyde, which we now know is not terribly good as far as TSE
agents are concerned.

The second example, which David Asher also listed, was instruments used
on a suspect case of CJD, neurosurgical instruments, I should say, were
exposed to hot air, 180 degrees centigrade, for two hours before reuse,
and there was transmission from patient to patient.

Now, as I said, and as David Asher referred to, there is actually no
convincing evidence that we have seen accidental transmissions through
neurosurgical instruments, but some data suggests epidemiologically that
there is perhaps some evidence of this, but there is no hard and fast
evidence.

Nevertheless, with such dreadful diseases that are incurable,
untreatable, there has been a constant nagging doubt about transmission
of CJD. To some extent, this was aggravated when Bob Will reported in
the UK the emergence of Variant CJD. As you know, the number of cases
has risen into the hundreds now and is still mainly confined to the UK.
The worrying aspect of that, of course, was that the work of Moira Bruce
clearly demonstrated that the agent causing Variant CJD was identical to
the BSE agent in cattle and quite dissimilar to any other TSE agent that
had ever been discovered.

Concerns regarding accidental transmission of Variant CJD between
patients was elevated by the finding that New Variant CJD
lymphoreticular system tissues in the patient examined, infectivity or
at least positive PrP was detected with 100 percent of these samples
compared with nil percent of the iatrogenic sporadic cases that were
examined or in other controls. That was a study here.

We also know that in a limited number of studies, if you have archival
tissue from patients who end up with Variant CJD, in this case,
appendix. You can find PrP in the appendix at the time when the patient
had no clinical signs of disease. So the potential for accidental
transmission through surgery is, at least, in theory enhanced by the
fact that surgeons compared with neurosurgical would much more commonly
be invading lymphoreticular tissues either deliberately or incidentally.

Now, I would like to just show a few overheads if I may. Both David
Asher and Bob Rohwer referred to the WHO meeting in 1999, which resulted
in guidelines being issued. It was related to not only clinical aspects
of CJD-like diseases, but also to concerns for the practical issues,
such as protection of laboratory staff, pathologists, surgeons,
etcetera.

Now, within the guidelines, there is this table here, which you may not
all be able to see, which is almost a short form of what I started with,
talking about ineffective methods. And the only thing I would say here
is that I will go in to talk a little about this procedure here, which
is regarded as variably or partially effectively boiling in 3 percent
sodium dodecyl sulfate, SDS, because this has been commonly banded
around as a probably relative effective procedure.

In terms of the actual processes recommended, and David Asher did show
you a summary of this, incineration, I will say nothing about, because
there will be something said about that coming up shortly. These
procedures, they were based on what was known from the literature on TSE
inactivation at the time of the meeting. To my knowledge, not much has
happened since then to alter the views and recommendations in these
guidelines, and they are listed in their perceived order of
effectiveness.

So we start with emersion in sodium hydroxide and heating in an
autoclave, as opposed to going on here to immersing in hydroxide then
transferring into water and going on to autoclaving. Also, here, the
alternative is to immerse in sodium hypochlorite, and then going on to
autoclave.

Here, we have emersion in hydroxide or hypochlorite, and then going into
an open pan and then autoclaving. This is because one of the options
here is the 134 degree centigrade porous load cycle in which you cannot
put fluids. So if you\'re putting instruments through these after the
fluid treatment, you must remove them from the fluid.

We then go on to suggestions for boiling. These are listed in order of
decreasing perceived effectiveness, bearing in mind that WHO
recommendations are, essentially, for the health community worldwide,
and that facilities and equipment availability will vary tremendously,
especially in some more deprived areas of the world.

Finally, we go on to talk about autoclaving at 134 for 18 minutes. And
then when it comes to things like surfaces, we revisit procedures like
sodium hydroxide and sodium hypochlorite. Then you can just do thorough
cleaning if you can\'t do anything else. And then there are some
questions about dry goods and autoclaving.

So I have been asked to address or discuss with you the data, in a
sense, that we use to back up these recommendations, which I will do and
finish with one or two bits of additional, perhaps anecdotal
information.

Will you go back to the slides now, please? I did mention that I would
talk briefly about SDS, because simply boiling in sodium dodecyl sulfate
or concentrations as low as 3 percent has been widely, well, fairly
widely recommended as a very effective procedure. However, in our own
experiments where we used 5 percent of this compound and even went on to
autoclave at 121 degrees centigrade, we certainly did not completely
inactivate.

Now, we did get down to almost a limiting dilution. In other words, we
have reduced infectivity probably in the region of 10,000 fold or
something like that, but within a medical care context to have surviving
infectivity at this level would be a concern. And so I present this
simply to discuss an idea that hot SDS is a universal panacea.

Now, Bob Rohwer discussed with you his hydroxide data that he
co-published with Paul Brown in 1986, I think, and he also showed a list
of, if you like, some contradictory data. These are publications, which
are all saying much of the same thing, and that is that sodium hydroxide
looks to be pretty effective, but not completely so. A suggestion is
that you are knocking down infectivity, because this is at room
temperature, by the way.

And the one comment I would make to Bob about certainly our experiments
compared to his, I can\'t talk for many of the others, but clearly, you
found complete inactivation, but we didn\'t know it. It\'s acknowledged
in your paper that the sensitivity of your assays were slightly reduced
because of the toxicity of the hydroxide to the examples. In other
words, you diluted these to make them so that they could be tolerated by
the hamsters.

In our own experiments, what we found is that if we fiddled around
considerably, we could actually neutralize, get the pH down to neutral
in the end products, and provided they were injected very quickly into
the brains of mice, we didn\'t need to dilute. So there is a slight
difference in sensitivity between the tests. I\'m not saying that is
necessarily the explanation, but it is possibly so, because we do have a
solid bank of data saying cooled hydroxide is not completely effective.

Right. In our own studies, what we found was that after exposure and,
again, room temperature, this is a hamster agent, one molar hydroxide,
two molars for two hours. We brought the infectivity level something
down, certainly, but we were left with about 4 logs of infectivity.

Now, if we combine the hydroxide treatment with heat as has been
recommended, then, in fact, we find complete inactivation either when
you add hydroxide to the samples and immediately autoclave or when you
hold in hydroxide for an hour and then go and autoclave. And in other
studies, such as those from the Rocky Mountain Lab, they found that if
you held in hydroxide, then neutralized the pH and went on to autoclave,
you still got inactivation.

And these are the various publications, which all come to the same
viewpoint, somewhat unusual in TSE studies to have so many publications
saying the same thing, that hot hydroxide is effective, whether this is
a sequential process or whether the hydroxide treatment is at the same
time as your autoclaving.

Now, in terms of sodium hypochlorite, which is one of the recommended
procedures, we did some studies quite some time ago with two strains of
mouse agent exposed to sodium hypochlorite containing various
concentrations of available chlorine, and we found that once you got up
to about 8,250 parts per million of available chlorine, you had a
complete effect.

Now, the data here, and these were studies that were done on behalf of
the Department of Health in the UK some time ago, and being extremely
conservative, the Department of Health accepted the data, but said well,
to play it safe, we\'ll make the recommendation that you should use
sodium hypochlorite containing 20,000 parts per million of available
chlorine, which considerably exceeds the lowest levels of efficiency
here, but that\'s where the recommendation came from to use 20,000 parts
per million.

Somewhat later, using two sources of BSE infected cow brain, we tested
sodium hypochlorite once again. Alongside it, we also tested sodium
dichloroisocyanurate, which is another chlorine releasing compound,
which is generally considered to have a comparable efficiency compared
with hypochlorite when compared at the same levels of available
chlorine.

In these studies of these various concentrations of available chlorine,
there was no infectivity detected in any of the BSE cow brain samples
treated with hypochlorite. But when you looked at the samples treated
with the dichloroisocyanurate at comparable levels of available
chlorine, there were, in fact, a significant number of positives.

This came as rather a surprise, but we found then by doing assays on the
chlorine content left after the exposure periods, that the sodium
hypochlorite compared with the dichloroisocyanurate, if you look at the
starting and finishing concentrations of chlorine, the hypochlorite much
more readily gave up its available chlorine during these decontamination
procedures compared with this compound. It may be that longer exposures
might be effective, but we are already up to two hours, which it\'s
getting a bit impractical to extend things beyond that.

Mention was made of boiling. Well, we certainly do have data, which have
only actually ever appeared in an abstract sort of meeting. They have
never been formally published, but we did find with 301V, that if you
boiled for one minute, that we have no detectable infectivity left
compared with material exposed to hydroxide at room temperature or
microwaved for one minute.

Bob mentioned the data produced based on 134 to 138 degrees centigrade
porous load autoclaving. This was in either BSE infected cow brain,
scrapie infected sheep brain or scrapie infected hamster brain, and we
had survival rates as shown here, which, as Bob suggested from this
graph, fall pretty far down on his survival curve. And, indeed, when we
titrated, the starting titre here again was 9 and a half logs. It came
down to about 2 logs or less. So substantial inactivation, but, in fact,
in terms of health care, still a worrying amount of infectivity left.

In terms of more recent studies using 301V, we had really surprising
data for this experiment where we autoclaved either at 134 or 138
degrees centigrade for these periods of time with these weights of
tissue. Now, the norm is, of course, as you increase autoclaving time
and/or temperature, you expect the efficiency of decontamination to
increase.

In these studies, the reverse was true. In fact, we had more cases of
TSE in the case injected with the samples from the 138 compared with the
134 process, which was statistically significant. If done on a one off
basis, I would have had severe doubts about the technical quality of our
experiments here, but, in fact, we had other experiments running at the
same time, which showed the same trends, perhaps not so impressively as
here, but definitely showed the same trends.

Also, there were studies being carried out on behalf of the Department
of Health who insisted quite correctly that all of the equipment and the
processes should be independently monitored. And so we had a third party
monitoring the progress of these experiments, thermocoupling of blanks
for every single stage of the process. And there are some of you that
know there is still, I think, a T-chest full of trace-outs for all these
experiments. So I have no doubt that we\'re seeing a genuine trend here.

I mentioned that we\'re using 301V and we do know, as Robert Somerville
mentioned this morning, that 301V within the spectrum of the agents that
we have tested is certainly far more thermostable than others. These
strains here are all most precise scrapie agents. 301V is our most
precise BSE agent. There is a survival after autoclaving and the blue
bars are the untreated samples. And you can compare the titre losses
with the different agents after the autoclaving process.

My take on what was happening in these experiments was that in the past
where we found much more efficient inactivation or in some cases,
complete inactivation, we often used intact pieces of brain tissue. In
the more recent experiments, as Bob Rohwer said, we were using brain
macerate. This is undiluted brain, which is just mixed up, so it\'s a
homogenous sample to give you a blancmange like material for
autoclaving.

Now, in putting these samples into what Ron described as long neck
tubes, not terribly long neck, but there is almost inevitably some
smearing and drying of the infectivity onto the tubes before you get to
the autoclaving stage.

My concept, my take of what is happening and what explains the results
is that during the porous load autoclaving process, which I must tell
you, unlike the gravity displacement system where there is usually a
slow buildup of steam, the porous load system involves a huge and rapid
admission of steam into the chambered autoclave, which, in my simple
hypothetical structure here, is able to fix any proteinaceous material
in these fringes, and that paradoxically, if that protein is PrP
protein, the actual fixation process, which occurs early and rapidly at
the beginning of the steam process actually protects that infectivity
from the subsequent sterilization of the steam effect.

If that was so, that would explain why the 138 degree samples were more
positive than the 134 since you would expect the rapidity and efficiency
of that heat fixation to be greater at 138 compared to 134. I hope to
show you in the next few slides that this is not all quite cuckoo land.

We do know that if you fix infectivity or fix infected tissues with
formaldehyde, you make that infectivity colossally more resistant to
inactivation by autoclaving. Here, we have 50 milligram fragments or
whole mouse brains that are infected with the strain called 22A, fixes
in formalin and then autoclaved. And, in fact, 100 percent of the
recipient animals have gone down in disease. Whereas, in these
experiments, we were completely able to inactivate infectivity in these
samples if they were simply emerged in saline.

Similarly, if you immerse infected mouse brains in ethanol, another
protein fixative, and then autoclave, you get remarkable survival of
infectivity even though ethanol fixed in autoclave, 100 percent
recipient animals going down. So there is clear evidence that if you fix
the PrP protein by whatever means, you, in fact, stabilize it to the
extent that it is not normally taken out by the standard autoclaving
procedures that we\'re looking at.

And to test the hypothesis a bit further, we picked up on the
experiments of David Asher\'s going back, I think, to the 1980s. I think
he was among the first to observe that with scrapie-like agents, if the
materials are dried onto surfaces, they become extremely difficult to
inactivate.

Here, we have an infected brain homogenate simply autoclaved and then
injected into mice, and in this case, one of the eight animals went
down. If on the other hand, we took the homogenate, dried it onto a
slide, autoclaved it and then reconstituted it, scrape the top of the
infectivity again and try to challenge animals, 100 percent of these
recipient animals went down.

That, again, would be compatible with the idea that this thin sheet of
material on a microscope slide in autoclave would be subject to very
rapid and efficient heat fixation. And there is one more experiment that
we carried out with this in mind where we knew that dry heat at 160 for
an hour would not inactivate the agent, but that autoclaving without any
other processing was effective. We dry heated, which would heat fix, and
then autoclaved and, again, we had substantially more survival of
infectivity.

So my interpretation is that the effects that we\'re seeing of smearing
and drying of tissue in tubes in autoclaving experiments may well be
down to heat fixation. I make no apology for the fact that many of the
experiments that I have done have used brain macerate and not brain
homogenate, that these have all been more scarce conditions, because
many have been driven by public health concerns funded by the Department
of Health who do actually want to know what happens under worst case
circumstances that could reflect conditions relating to tissues dried on
instruments, etcetera.

In terms of concern over instruments, the Department of Health has
funded quite a number of studies relating to decontamination,
disinfection. They are quite interested in the combined hydroxide and
heating effect, and one of the concerns is what effect does this have on
stainless steel instruments and devices?

So one of the studies being carried out in Edinburgh is to look at test
pieces made of stainless steel before and after various hydroxide
treatments. It is mainly facilitated by collaboration with the
engineering department who have a scanning white light interferometer
where you can compare the roughness indexes of surfaces before and after
various treatments. It will print out different graphs giving you the
roughness indexes.

And here, just to the naked eye are test pieces, which on the left hand
side are all untreated. These are different grades of stainless steel.
On the right hand side are pieces that have been subjected to
autoclaving at 121 centigrade for 24 hours. And as you can see, in some
cases, there is hardly any difference, but in some cases, there is a
darkening of the testing piece.

As I understand it, this is due to precipitation of chromium salts and
what I\'m unaware of is whether you can clean these chromium salts off
and start again with a pristine surface. This is what was started before
I left the unit, so I\'m not quite sure of the current state of play.

I will finish off with just three slides containing anecdotal
information, which may be of some interest. One is that a low formalin
fixed tissue is incredibly difficult to inactivate, and one would
recourse usually to incineration for its disposal. We did find that the
hot hydroxide process, when applied to infected brain tissue fixed in
formalin was, in fact, effective at removing that.

And I will finish up with two more overheads, if I may. One goes back to
the GME study, and the figures may have changed slightly, but the
principles are nevertheless the same. It was discussed how the
superimposing of our sodium hydroxide step, especially to the ossein
material that remains after the acidic extraction process where there
was a significant amount of infectivity surviving, at that point. If you
then applied \-- I\'m sorry, this should be hydrochloric acid up here.
If you then apply .3 molar sodium hydroxide for two hours at ambient
temperature, there was no infectivity detectable in the resulting
gelatin.

This clearly suggests that earlier studies using infected brain
suggested that one more sodium hydroxide is quite effective and not
completely so, these studies suggest that when you get the circumstances
in an environment such as ossein where you are largely devoid of any
extraneous lipids or proteins, that the hydroxide process is much more
effective.

And I will leave you with some recent data from a commercial study,
which I have some sketch information for you from. This involves a
process where raw materials exposed to saturated lime calcium hydroxide,
and then it goes on to hot lime at 80 degrees centigrade, here we are,
sorry, and thereafter onto even hotter lime at greater than 140 degrees
centigrade under pressure, much of the same conditions if not higher
conditions than those described that are completely effective.

Now, the thing to be on your mind here is that the pH of the lime, the
maximum pH of lime is significantly lower than that of one molar sodium
hydroxide. What we seem to be finding here is that after the exposure to
saturated lime for three hours at 80, we do have some titre loss. The
expectation then might be that when you go on to this very high
pressure, high temperature process, that you might lose all the
infectivity, but, in fact, you do not.

To me, this demonstrates potentially two things. One is that molarities
of hydroxide lower than one molar may not be truly effective under the
high pressure conditions and/or separately than any surviving
infectivity from this stage, which is carried out at 80 degrees
centigrade, the heat fixation, which goes on in here, all the surviving
infectivity may, in fact, render it more resistant to inactivation at
this level. So these are speculative comments, but they all contribute
to the general arguments about heat and hydroxide. And I will leave it
there. Thank you.

CHAIR PRIOLA: Are there any questions? Oh, please, Dr. Edmiston.

DR. EDMISTON: I have a comment, which I want to direct to the speakers,
the previous speaker and Dr. Taylor, and also a general comment to the
members of the panel in terms of how this applies to surgical
instruments in the operating room.

I am not surprised that you haven\'t achieved complete inactivation,
because as a rule, it\'s a general trend we sort of adhere to, is as
long as there is biological material present or, I should say, as long
as the organic component is still there, it\'s unlikely you\'re going to
see complete inactivation.

From a surgical perspective, one needs to recognize, and I\'m not quite
clear on what my colleagues are doing in Europe, but at least from the
U.S. perspective, we just don\'t take surgical instruments and put them
into an autoclave. There is a pretreatment facility, which reduces
organic comment, and I know the next speakers will address that probably
in some detail.

Actually, I am heartened by some of the data you have shown in terms of
inactivation, especially in the presence of high organic content. The
fact that in these high carbon environments, you are able to reduce the
number of viable particles, so I think we need to think about this two
step process as we procedure through the next day and a half in that
we\'re just not talking about instruments being directly sterilized.
We\'re talking about a process in which instruments are being rendered
sterile by virtue of not only a sterilization process itself, but also
the removal of organic material prior to sterilization.

DR. TAYLOR: Yes. Could I make one comment now? There is, at least in the
UK, what I consider to be a worrying trend, and that is that
traditionally in a very common sense fashion, it was common in wards and
even theaters for certain instruments to be washed in the sink before
they went on for washing in the Central Sterilization Department.

That process is increasingly being discouraged for health and safety
reasons. It\'s resulting in an increasing number of instruments reaching
the Central Sterilization Department with absolutely dried on blood,
tissue and whatever, and this is the problem that, I think, you are
referring to. The washing processes as they exist at the moment, at
least in the UK, are largely incapable of dealing with the situation
where you have material that is absolutely dried or baked on.

DR. EDMISTON: And I think the recommendations and this Committee needs
to anticipate the fact that that is a problem. Therefore, the
recommendations not only from this Committee, but from other
professional organizations such as APEC and others would suggest that
pretreatment of these instruments is mandatory.

CHAIR PRIOLA: Yes, Bob?

DR. ROHWER: Yes. I am aware that that\'s how it\'s done in the hospital
setting. I work in a hospital, but the problem that we have with that is
the potential for cross-contamination at the level of the cleaning, and
especially in a laboratory environment at least, that would be a
disaster for us to spread this stuff around in our sinks and cleaning
stations before it ever got to the autoclave.

So we want to make absolutely sure that we know that our instruments are
contaminated. You don\'t necessarily know that yours are, and so we want
to make absolutely sure that everything is gone before we even handle
them, and we do that by going to these extreme measures.

On the other hand, the only thing that I find encouraging about what you
do is what standard practice is in the hospital, is that slide that Dr.
Asher showed in his introduction. There really isn\'t any evidence that
instruments cleaned and sterilized in the way that is specified by
current practice are causing CJD infections, and I think we have to give
a lot of weight to that.

On the other hand, I think it\'s also very important to think about the
cleaning step and what kind of potential that poses for having a major
accident if you don\'t contain that particular environment, as well,
because I consider that a high-risk environment.

DR. EDMISTON: Well, you need to know that most of us have had a high
threshold interest in this for several years, and more and more of
neurosurgical instruments are being triaged and actually being treated
separately in separate kits. And for the most part, and I will say for
the most part, because there are exceptions, are not getting into the
main surgical instrument stream.

And we\'re spending a lot of time and effort with our neurosurgical
colleagues to first of all identify potential patients or suspected
patients, but overall, I can tell you most surgical departments, most
hospitals, will have unique surgical, neurosurgical kits, and this is
becoming more and more common for the reason that you just mentioned.

DR. ROHWER: I guess the thing is I would like to know more about how
that segregation takes place, because it\'s not particularly comforting
to me to know that the neurosurgical instruments are being segregated.
Neurosurgery is potentially the biggest hazard in terms of passaging the
disease, so you run through a set of CJD exposed instruments, and then
that is followed by a set of cleaned instruments, you know, coming from
a normal patient or something like that. How do you assure yourself that
you\'re not getting cross-contamination at the level of neurosurgical
instruments in that type of environment?

DR. EDMISTON: I won\'t go into a lot of detail on this, because I know
my colleague over here will discuss it, but when patients are
identified, those instruments are quarantined and sequestered, so that
they are treated entirely separate from the rest of the general
instruments. So that is the policy that most of us have developed over
the years in dealing with these suspected patients.

Now, the other issue is well, how about all of the other neurosurgical
patients, which you find out about anecdotally? Now, that is an
important process to discuss, but in terms of those that we identify or
we suspect, those instruments are quarantined and they are triaged and
segregated out of the system.

DR. ROHWER: Okay. Can I say anything more? Are you tired of hearing me?
I guess my rebuttal to that would be that my guess is that the greatest
part of the risk comes from people that you will never, ever identify as
even carrying the disease, and that\'s the greatest part of your
exposure. You will never know about it, and what I see a need for is
some way to actually effectively sterilize the cleaning environment
between uses.

CHAIR PRIOLA: Are there other questions for the speakers? I have one
quick one for Dr. Taylor about the experiment you showed where you
exposed material to dry heat at 160 degrees, and you had complete
transmission. And then you took the material, exposed it to dry heat and
then, if I remember, you autoclaved following the dry heat, and that
dropped to almost 50 percent survival or you get 50 percent survivors.

What implications do you think that has or does it have any implications
for multiple rounds of autoclaving, say wet autoclaving or multiple
rounds of sterilization and getting rid of that residual activity?

DR. TAYLOR: It\'s difficult to answer your question within the context
of infectivity dried onto surfaces. All I can say is that I have done
one experiment where I didn\'t make any attempt to smear and dry, but
just using standard samples with the hamster agent where after one round
of sterilization, and I am quoting figures very crudely here.

In the first round of a standard autoclaving procedure, I lost something
like 4 logs, somewhere about there. And when that material was taken and
just reautoclaved, the loss on the second round was about 1.7 logs. So
the second autoclaving, even under these conditions, was certainly not
very efficient, and I suspect would have been even poorer if this had
been agent that partially survived after smearing and drying.

CHAIR PRIOLA: Dr. Gambetti?

BOARD MEMBER GAMBETTI: Listening to all these presentations, of course,
are very informative. One, however, wish that experiments were available
in which decontamination of surgical instruments is monitored under more
realistic conditions. For example, one wished that there would be some
data on decontamination of surgical instruments used experimentally in a
more surgical operation on a CJD brain, and then see how this level of
contamination that is a classic level of contamination that you may
expect from a CJD brain in surgery, how the decontamination is effective
on those particular conditions.

Vice versa, one would like to know how much decontamination is achieved
on contaminated surgical instruments after the routine sterilization
that the surgical instruments undergo, as I said, under routine
conditions. Those are the data that I would like to know whether they
are available at all. I have never seen, so I think those would be very
useful data to have for this discussion.

CHAIR PRIOLA: Dr. Taylor, do you have a response to that?

DR. TAYLOR: Yes, I have an experiment that I started before I retired,
and I\'m going to throw the buck right over to Robert Somerville here as
my successor. In this experiment, the very question that you\'re asking
was asked. In other words, how realistic or how appropriate are the
inactivation we\'re achieving to real life situations?

Now, we weren\'t doing neurosurgery on human patients, but we were daily
doing surgical interventions within the brains of infected animals. So
we took deliberately infected instruments that had been deliberately
traumatized into animal brain, subjected them to routine washing
procedures, and then proceeded to reuse these instruments again
neurosurgically or in subsequent animals.

My take on things before I left, and I haven\'t looked at the data
since, was that even the washing processes in the lab, which were not
anything up to the Central Sterilization Department were having a
useful, if not complete effect. But by the time we got to reuse of these
instruments on animals, they weren\'t, as measured at that time,
producing any significant levels of infections in the animals. I don\'t
know if Robert can add anything to these data or are they still lying
buried?

DR. SOMERVILLE: I think they are still lying buried, David. I don\'t
have access to the data at present. What I would say to Professor
Gambetti though is that attempts, which I think Professor Weissmann is
addressing the Committee about tomorrow, I think model the kind of
situation that you are trying to \-- the kind of question that you\'re
asking, and that is the implantation of contaminated surgical
instruments, stainless steel. Professor Weissmann has already done some
studies with contaminated wares and our lab is also hoping to initiate
this kind of system with different grades of stainless steel.

One of the problems that you have to appreciate is that surgical
instruments made out of various kinds of stainless steel, and that is
one of the challenges of actually set up these kinds of experiments, is
how you model the different kinds of surfaces that will be involved in
real life. But to summarize, I think the best way of testing your
question is through this kind of model.

CHAIR PRIOLA: Okay. I think we\'ll move on to the next speaker who, as
Dr. Asher mentioned, was supposed to be Dr. Philippa Edwards, but she
has taken ill and is unable to attend, so Dr. Pedro Piccardo from the
FDA has graciously agreed at the very last minute to give her
presentation. Dr. Piccardo?

DR. PICCARDO: Thank you. Well, obviously, Dr. Edwards could not attend,
and yesterday I was given somehow the daunting task of presenting the
information that she provided. I will try to do this as objectively as I
can.

Next one. Okay. It has been settled already iatrogenic transmission of
Transmissible Spongiform Encephalopathies from person to person has
occurred in non-Variant CJD, and this has instated already, and here are
the numbers that were provided by a publication called [Brown and
Neurology in the Year 2000]{.underline}. And as you see, the bulk goes
to growth hormone treatment and dura mater grafting.

However, there are here five cases implicating which neurosurgery,
meaning contaminated instruments, have been implicated. On top of that,
we have a few cases following treatment with gonadotrophin, chromium
transplants and, of course, electrodes here.

The next, please. However, one of the big problems came when in 1986,
Bovine Spongiform Encephalopathy was described in the UK, and the
problem became humongous when in 1996, vCJD was described in humans. As
you see here, I mean, obviously, there are a numbers of barriers that
have been established to try to prevent the transmission of vCJD, the
agent, from animals to humans. However, the big question here is humans
are being exposed. Humans died with vCJD, and the question is we don\'t
know how many people has been exposed, how many people could be
infected.

The next one, please. And, of course, we don\'t know how many people may
be asymptomatic, at this time and carriers.

The next one, please, the next one, the next one. Okay. The next one,
please. The next one, please, the next one, the next one. So due to
great uncertainties, risk assessment has been considered. What happened?
Oh, okay. Here we go. Due to great uncertainties, risk assessment has
been considered. The risk assessment has considered a wide range of
scenarios.

And why the risk assessment was done? Basically, for two reasons. One
was to determine the risk of transmission of vCJD through surgical
instruments, and the second one to indicate what measures could be the
most effective to reduce the risk.

The next one, please, the next one, next one, next one, next one. The
guidance follows the assumption that an average of 10 milligrams of
material could remain in instruments, and this information I gather from
the document that was provided by the CJD Incidence Panel.

Next one, please. Go ahead again, again. The risk could be calculated
for different scenarios, and the effect of different actions could be
estimated.

Next, please. Next, please. Okay. Improving the standards of
decontamination is one of the main objectives of the UK policy, and
single-use instruments have been considered, for example for extraction
of CSF, and the idea was to use as much as possible single-use
instruments without compromising the clinical standards, of course. And
a pilot program was established to use single-use instruments for
tonsillectomies.

Next one, please. Go ahead. Why tonsillectomies? Why was this chosen?
Because infectivity is present in vCJD in tonsils. I mean, PrP has been
found in tonsils from patients with vCJD. The other thing was the
relatively large number of operations and the other thing is the young
patients usually with long life expectancy go through this type of
surgery, and these are instruments that can be identified.

Next one, please. Okay. But there were some adverse reactions. I mean,
why there were problems? One was you cannot probably think that the
problems raised from the quality of the sets, the surgeon preferences
and the other problems were unrelated to the use of single-use
instruments. So at this time, there is an audit on this situation.

Next one, please. So what Dr. Edwards tried to convey, the message that
she tried to convey with this cartoon is that while trying to solve one
problem, you create another.

The next one, please, again. The best decontamination available cannot
be guaranteed to remove all sorts of infectivity, and single-use
instruments definitely is a situation that is not possible for all kinds
of surgery, so we must bear that in mind.

Next one, please. Okay. Here, we have on this panel, a tissue forceps,
the tip of a forceps that has been routinely decontaminated. Here, we
have electromicroscopy, and this what we can see here in green is
material that remains on the tip of that forceps. This is the kind of
material that remains. This is florescent staining for protein, and this
is the superimposition of these two images gave this image. So,
obviously, there is a significant amount of material that remains, a lot
of which is protein following routine decontamination.

The next one, please. So to reduce the risk of transmission of TSE from
person to person, the Department of Health seek guidance from the
Advisory Committee.

Next, please. Next, please. And a first version was done in 1998 and
now, there is a revised version, June 2003.

Next, please. This presentation concentrates mainly on the risk arising
from the care of patients.

Next, please. Yes, okay. So well, one of the issues is dealing with
symptomatic patients. I mean, when dealing with patients with CJD, there
are three types of definition. One is a definite case. By definite we
mean something that has been clinical and pathologically confirmed.
Probable case, which is has clinical, but on top of that usually, there
is electron encephalographic analysis and there is MRI imaging analysis,
and possible CJD when usually is by clinical presentation.

Next, please. Now, when we are dealing with asymptomatic patients, when
we talk about risk in the case of \-- when we talk about asymptomatic
patients and we talk about risk, we have to consider two situations. One
is in the case of inherited diseases, and by inherited diseases, we
consider that there are two or more blood relatives are affected by a
prion disease or one or more blood relative showed genetic testing, show
a mutation in the prion protein gene. Usually what is done is PCR
sequence, the open reading frame of the protein, and then from there you
can detect mutations.

Next, please. Now, the other is the iatrogenic risk, and this case
already was mentioned treatment with hormones, dura mater grafts, and
that is why, I mean, obviously, the Department of Health seeks advice.

Next, please. This table is based on what we currently understand about
the distribution of infectivity in sporadic CJD or in non-Variant CJD,
and obviously, when we talk about tissue infectivity as it has been said
already many times, the highest amount of infectivity is here in the CNS
or retina and low medium type of infectivity in the eye and olfactory
epithelium.

Next, please. Now, when we talk about risk of different tissues in
Variant CJD, the situation varies, because we have to introduce into the
medium risk tissue lymphoid tissues. The rest remains the same.

Next, please. So we don\'t have a problem here and we don\'t have a
problem here, because this is by genetic testing or what was said
already, is we can know who these people are, and we understand who
these people are. But the problem is when we deal with sporadic or when
we deal with variant, people that are asymptomatic, at this time, but
might have infectivity.

The next, please. Yes, okay. So the problem comes or starts when,
obviously, a CJD patient is diagnosed, and immediately the question
should be has that patient had surgery or donated blood, etcetera, and
then try to assess what is the risk to other patients that have been
exposed to instruments that have been used on this patient.

Next, please. And the risk, basically, will depend on the type of tissue
that we are talking about, because we said already that there are
tissues with high levels of infectivity and in this case of vCJD, the
lymphoid tissue corresponds with tissues with medium levels or medium
risk.

Next, please. So this graph is an estimate that comes from animal
studies, so this is an estimate that comes from animal studies. And,
obviously, the paren of tissue infectivity in vCJD probably could follow
this, and this is the onset of clinical symptoms. So if we go, let\'s
say that the surgery was done way before the development of clinical
symptoms, probably the amount of infectivity will be very low, and
because we are dealing with vCJD in this graph, we have two parameters
or two tissues to consider. One is the CNS and the other is lymphoid
tissues.

Next, please. So, as I said already, the risk depends basically on the
type of \-- I mean, depends once again on the type of tissue where the
surgery is performed, and if there is variable time between surgery and
onset of disease. Well, this basically refers to the previous graph.

Next, please. Okay. One of the issues here is that the risk depends \--
let me see. In the document provided by the CJD Incidence Panel, it is
stated that the first washing and autoclaving would achieve at least a
10^5.4^ reduction of infectivity, and this was already mentioned by Dr.
Taylor before, and that subsequent cycles of decontamination reduce the
infectivity, but it\'s much less effective.

The next one, please. So at this time, the Department of Health is in
discussion with manufacturers of surgical instruments. I mean, the
discussion is based on what is the probability of using single-use
instruments or to replace parts or to provide instruments that could be
easily decontaminated.

Next, please. So what are the aims of the CJD Incidence Panel? Well, it
seems to be quite obvious, which is to protect the patients. Let me see.

Next, please. The aims. Go ahead again, again. Next, please. No, yes,
yes, here we go. Obviously, the aims are to protect the patients and to
inform potentially exposed patients, and to inform the public and to
increase the knowledge.

Next, please. So in management of risk, I mean, what is being done is
quarantine the instruments during the risk assessment, and instruments
that have undergone less than 10 cycles of decontamination should be
incinerated.

Next, please. Again, next, please. Okay. So the patients will be
contacted to alert them of their possible exposure and to take health
protective actions. So these are the patients that should be contacted
under these circumstances. If the index patient goes through, I mean,
the material went through high-risk procedures, the first six patients
that follow that first surgery should be contacted. This is for tissues
with less amounts of infectivity, and these are the amount of patients
that should be contacted.

Next, please. So this is sort of the kind of data that has been gathered
during the last two years of experience, and this is the incidence
reported to August of 2002, and definitely we have 39 cases implicated
in Variant CJD, 39 cases implicated in sporadic CJD, and that there are
few that correspond to familial or non CJD or unclear, cases that could
not be determined.

The next one, please. The type of surgery is 131. So obviously, before
were 87, I believe. Yes, 87, and now, we are talking about 131
surgeries, and the reason is that some patients went through more than
one surgical procedure. And what we see here is that the GI surgery
takes the bulk followed by obstetric and gynecology and here we have
neurology, neurosurgery.

Next one, please. So in 76 incidents, tracing was sought. Some or all
were traceable, that means in 34. In 18, it was not possible to trace
them. And in 24, there is incomplete information, at this time.

The next one, please. So instruments that have been quarantined are 48
and that have been not quarantined are 39.

The next one, please. The fate of the quarantined instruments, we have
21 that have been returned to use, because it was assessed that the risk
was not higher than the usual risk for the UK population. Here, we have
four that have been completely destroyed, the whole panel was destroyed,
because it was not possible to identify exactly which of the instruments
was involved. And in 23 cases, the hospital directly decided to take
care of the instruments and destroy them.

The next one, please. So, obviously, this is a very difficult task and
there are a number of dilemmas and difficulties, and there are a number
of scientific uncertainties, and it is very difficult to trace back
instruments and sometimes patients and, of course, there are ethical
issues that are involved.

The next one, please. Go ahead, yes. And these are the websites that Dr.
Edwards suggested consulting for further information. I think this is
it. Thank you.

CHAIR PRIOLA: Dr. Khabbaz?

DR. PICCARDO: Oh, before \-- excuse me, sorry. Yesterday, after I
learned that I had to give this presentation, I called the UK
immediately, right away, and the first question I asked before you ask
me the question was what do I do with \-- how do I handle the questions?

So the thing is we will make clear note of your questions, and then we
will forward the questions to the UK, and Dr. Edwards has been kind
enough to review them and, hopefully, if she feels well enough, to
provide the answer tomorrow. So we are in business. Anyhow, if you want
to ask the question, go ahead.

CHAIR PRIOLA: So that gets you off the hook, doesn\'t it, Pedro? Yes.

BOARD MEMBER KHABBAZ: You may know the answer. I had actually a couple
of questions. One has to do with the adverse events related to
single-use of instruments for tonsillectomy. Do you have any idea what
types of adverse events would occur?

DR. PICCARDO: Yes, the answer is bleeding, bleeding. That is the answer
to that.

BOARD MEMBER KHABBAZ: Thanks. The second question is I don\'t think \--
I may not have understood you correctly. I think when you talked about
the various types of CJD, you mentioned for inherited and iatrogenic not
concerned for infectivity?

DR. PICCARDO: Sorry, come again. I mentioned \--

BOARD MEMBER KHABBAZ: No infectivity for inherited and iatrogenic versus
sporadic and Variant CJD. Is that in terms of how they got it or for
peripheral tissues?

DR. PICCARDO: Let me see. Can you pose the question again? I have a
problem hearing, too.

BOARD MEMBER KHABBAZ: Okay.

DR. PICCARDO: Listening to the question. Yes, go ahead.

CHAIR PRIOLA: Yes. I think you\'re referring to the slide you showed
where you had sporadic patients, iatrogenic and inheritable. And are you
asking if there was infectivity associated with those patients?

BOARD MEMBER KHABBAZ: I didn\'t understand the statement that there is
no infectivity related to inherited and iatrogenic.

CHAIR PRIOLA: Oh, I don\'t think that\'s what you said.

DR. PICCARDO: No, well, I would be happy to review Dr. Edwards\' slide.
However, my answer to that is that there is no difference between
inherited and sporadic. We probably put them in the same box. We will do
a difference when we deal with Variant CJD, because that is when we have
tonsils and we have the lymphoreticular system involved that we tend not
to have in sporadic or other forms of CJD.

CHAIR PRIOLA: Right, and I think that was one to identify, patients at
risk, right, prior to use of instruments. If someone has inheritable
mutation, then that is a patient that you identify as being at risk of
possibly transmitting to somebody else. I think that\'s what that \--

DR. PICCARDO: Right, in terms of risk. I mean, if you have a patient
that, obviously, comes from a family and has the mutation, etcetera,
etcetera, you know that patient is at risk already, so it\'s very easy
to recognize that patient. It\'s also very easy to recognize a patient
that, let me see, that went through surgery that has a dura mater graft.
It could be a patient at risk. However, if you say well, let\'s take
sporadic CJD, maybe I am incubating sporadic CJD and I don\'t know and
no one will know.

CHAIR PRIOLA: Dr. Nelson?

DR. NELSON: How would you classify cerebrospinal fluid that, let\'s say,
has lymphocytes or is an inflammatory cerebrospinal fluid? Would that be
the same as blood being low-risk or would it be closer to CNS tissue?

DR. PICCARDO: Well, I would like someone else, probably Dr. Asher, to
attend that. Before we go ahead with that, Dr. Edwards made clear that
they provide disposable instruments for CSF extractions, so now, it\'s
single-use. Go ahead.

DR. ASHER: Yes. In the NIH series, about 15 percent of spinal fluids
from subjects with mostly sporadic CJD did transmit disease to primates.
So the risk of spinal fluid is comparable to the risk of some non CNS
solid tissues, lymphoid tissue, liver, kidney, spleen, lung.

DR. NELSON: But it\'s definitely higher than blood?

DR. ASHER: Higher, definitely higher than blood.

CHAIR PRIOLA: Yes, Dr. Gambetti?

BOARD MEMBER GAMBETTI: I believe that the experiment that, David, you
are quoting included not only sporadic, but also Kuru patients, and
three of 37 or so, spinal fluid tested transmitted the disease. Do you
know whether some of the CSF that transmitted the disease were actually
from Kuru patients, rather than sporadic case? Do you know that?

DR. ASHER: I think it\'s in the \'93 article, but I don\'t remember.

CHAIR PRIOLA: Okay. I think we\'ll move on. Thank you, Dr. Piccardo.
We\'ll move on to our last speaker before the break if there are no
other questions, and that is Dr. Bill Rutala who is going to discuss TSE
agents and infection control in U.S. hospitals.

DR. RUTALA: Thank you very much and good afternoon. What I would like to
do very quickly, and certainly by looking at the next slide, review the
recommendations on and the practices in U.S hospitals as it pertains to
the prevention of cross transmission from medical devices contaminated
with prions and, hopefully, have a few minutes to discuss how important
methodology is, and how important methodology is from the standpoint
that we can fail to inactivate even easy to kill microorganisms like
bacteria with FDA cleared sterilization processes dependent upon the
methodology that is employed to include the absence of cleaning.

Next slide. Let\'s begin with the rationale for the U.S.
recommendations, and these recommendations have existed for decades, the
recommendations in infection control literature, surgical literature,
certainly, essential processing literature and so forth.

But let\'s look at the next slide. As we know as far as the epidemiology
of prion transmission, we know that it\'s not spread by contact. It is
not spread by airborne. It is not spread by environment, but we are
concerned about the iatrogenic spread.

Next slide. We can see here that contaminated medical instruments have
been implicated in disease transmission, and we\'ll discuss that in just
a minute.

Next slide. Well, let\'s look at this issue of prion transmission via
surgical instruments.

Looking at the next slide, we see, essentially, the two confirmed cases
that have already been mentioned. Those two confirmed cases, of course,
were reprocessed by a method that we never use in U.S. hospitals, a
combination of benzine, alcohol and formaldehyde vapor, and then we also
have four suspected cases.

Those four suspected cases are involved with CJD that has occurred in
persons following brain surgery. However, only one of the four had an
index CJD case identified. These cases occurred before 1980 and there
has been no known failure of steam sterilization to date.

Next slide. How about the infectivity of human tissue as we discuss this
rationale? As we already know by looking at the next several slides, we
used epidemiology data and, of course, we used experimental data and
infectivity data. We know that there is evidence of transmission via eye
and brain from an epidemiology standpoint, and we know that
experimentally we can inoculate animals, susceptible animals, and
demonstrate that certain body fluids and tissues transmit CJD.

And we have already discussed the contents of the next slide, which is
that there are certain tissues that are considered high-risk, certain
tissues are considered low-risk and, of course, some tissues that are
considered no risk.

Next, we see the issue of removing microbes by cleaning, something that,
certainly, we need to discuss a little bit more as it pertains to the
methodology and how methodology is so important in this issue of prion
inactivation.

In the next slide, we will see, essentially, something that has already
been mentioned by one of the panelists. The issue that effectiveness
should not consider only the effectiveness of the disinfection of
sterilization procedure, but also has to consider the effectiveness of
removal by cleaning. And, of course, the probability of a device
remaining capable of transmitting disease is related to not only the
initial concentration of that prion on the surgical instrument, but also
it is related to the effectiveness not only of disinfection and
sterilization, but also cleaning.

And there are literally dozens of studies in the literature, which show
how effective cleaning is. Cleaning will reduce anywhere from 4 to 6
logs of microorganisms by a manual or a mechanical cleaning procedure.
We don\'t have as much data regarding protein reduction, but there are a
few papers in the literature that demonstrates there is, approximately,
a 2 log reduction of protein by the various cleaning procedures.

In the next slide, we see the prion inactivation studies. We don\'t need
to go over this very much. We\'re just going to very quickly go through
a few slides. We could possibly put a question mark up here with prions.
The question mark I would put up there is related to the fact that maybe
the studies that have been done are artifactual in nature and,
essentially, a reflection of the methodologies that are employed, and I
think I can show you data that would be supportive of that.

And then also, we would see here that other microorganisms fall below
possibly prions and spores and it pertains to the susceptibility to
disinfection and sterilization procedures. And in just a minute, I\'m
going to show you some data where bacteria will survive FDA cleared
sterilization processes, because precleaning did not precede the
sterilization process.

Next slide. We know, of course, there are many procedures that are
ineffective or partially effective.

Next slide. We know also that there are some gaseous sterilization
procedures and, of course, physical procedures that are also ineffective
or partially effective processes.

Next slide. We can see, of course, that there are some effective
disinfectants and, of course, by effective here, we\'re saying a 4 log
reduction decrease in the ID~50~ within one hour and, certainly, among
them include sodium hydroxide and sodium hypochlorite.

The next slide will just tell us what the effective processes are as it
pertains to sterilization, and this is what we use in U.S. hospitals.
We, of course, use sterilization primarily by steam sterilization with a
prevacuum sterilizer at 134 for 18 minutes. Sometimes, the combination
of sodium hydroxide and steam sterilization is employed, but it\'s not
widely employed because of some of the deleterious issues associated
with the combination of sodium hydroxide and steam, deleterious, of
course, to the instruments, deleterious to the sterilizer and, of
course, the vaporization of sodium hydroxide to staff. But we certainly
recognize the effectiveness, and that is an option for hospitals to
choose.

Next slide. As it pertains to risk associated with instruments, let\'s
see what we have with that.

Next slide. I just wanted to mention, essentially, that there are
certain categories of instruments in every health care institution, not
only in the United States, but in the world. There are certain
instruments that we consider must be sterile. They are instruments that
have contact with sterile tissue or the vascular system. We consider, of
course, then to be very critical.

There are other instruments like in endoscopes that have contact with
mucous membranes or skin that is not intact, and we have a very high
level of disinfection associated with those instruments. And the other
instruments are noncritical, only have contact with intact skin and,
essentially, are not involved in disease transmission.

The reason for mentioning that is seen in the next slide and that is,
essentially, in a minute we\'re going to develop, essentially, the
scheme for how we disinfect and sterilize instruments in health care
setting in the United States.

As it pertains to surgical instruments, a question was just asked. What
is the microbial load associated with surgical instruments? Actually, we
do know the microbial load associated with surgical instruments. A few
studies have actually evaluated the microbial load. Of course, it\'s not
for prions. It\'s for other microorganisms, and microbial load 80
percent of the time is less than 100 organisms. Rarely does it exceed
1,000 organisms. Many surgeries, many different investigators have made
that observation.

Next slide. Well, this is how we decide how to, essentially, employ
special prion precautions in U.S. hospitals. We, essentially, assess the
patient, assess the tissue and assess the device. Of course, we consider
whether it\'s a high-risk patient, a high-risk tissue and a high-risk
medical device, again, those critical and semicritical devices.

Next slide. As far as that is concerned, most U.S. hospitals then would
do special prion reprocessing, and that would be those higher
temperatures or a combination of sodium hydroxide with steam, special
prion reprocessing when it\'s a high-risk tissue, a high-risk patient
and a high-risk medical device, and for all other situations with one
possible exception, it would just be conventional disinfection and
sterilization.

The one possible exception would be a high-risk patient, a critical and
semicritical device and low-risk tissue. Some hospitals do treat
low-risk tissues from a high-risk patient, critical and semicritical
device as instruments requiring special prion reprocessing, so possibly
this would go into that category.

Next slide. So the conclusions of this, of course, is that from an
epidemiology standpoint, we have two cases of disease transmission that
are definitive, possibly four other cases. The guidelines that we have
discussed and are used in the U.S. are based upon epidemiological
evidence, tissue infectivity, the risk of disease associated with
certain medical devices and, of course, inactivation data, and the risk
assessment is based again on patient, tissue and device. And only when
there is critical and semicritical devices, contacting high-risk tissue
and possible low-risk tissue from high-risk patients do we require,
essentially, special prion reprocessing.

Now, the next slide, we see, essentially, what those special
reprocessing procedures are. We have already mentioned that this is the
preferred procedure that many hospitals use, the 134 at 18 and a
prevacuum sterilizer. There is no low temperature sterilization
technology that is recommended and we know, essentially, that there are
some disinfectants that have activity against CJD.

Next slide. So this is what we\'re talking about here. We\'re talking
about used instruments, and this is true for all used surgical
instruments. They are kept wet. They are not allowed to dry. They are,
essentially, cleaned before they are sent to central processing. We
don\'t let tissue and fluid dry on them. When they get to central
processing area, an area where all instruments are received for a
quality control standpoint, the instruments generally go into a
mechanical washer disinfector. In the case of special prion
reprocessing, there would be a special steam sterilization cycle, and
that instrument would be returned to health care.

Now, we have already mentioned, essentially, the rationale. The last
thing I want to do is very quickly look at methodology and how
methodology affects results. And I am going to show you some slides, and
let\'s begin with the next slide where, essentially, we can fail to kill
easy to kill microorganisms by methodological manipulations, and I will
call them manipulations, because all we do is we don\'t add cleaning to,
essentially, the process. And, of course, we have already seen how
important cleaning is.

Next slide. This is the issue. I don\'t want to get too involved in
this, but, essentially, the point needs to be made that there are a
number of studies that have been done, and most of those studies are
done in a worst case scenario, of course, and, of course, we try to
achieve sterilization by using appropriate reprocessing procedures.

And there are no studies, including the studies that have been published
involving cleaning, that reflect the reprocessing procedures in a
clinical setting. We use enzymatic cleaners. We use mechanical
sterilizers in a closed system and we use mechanical washer disinfectors
in a closed system.

Next slide. Now, I want to talk briefly how factors affected
sterilization, and many factors affect sterilization, but I am only
going to choose a couple. First, let\'s look at protein and salt.

Next slide. If we just put, essentially, some microorganisms on a
penicylinder like this right here, next slide, and then we let it dry
for 30 minutes, next slide, and then we put those penicylinders that are
inoculated with easy to kill microorganisms, such as E. coli and
pseudomonas and enterococcus faecalis in a FDA cleared sterilization
process like ethylene oxide or hydrogen peroxide gas plasma, in the
absence of serum or salt, you get 100 percent kill.

In the presence of serum or salt, you get 40 percent failure, as well as
in this case, 63 percent failure, a significant amount of failure,
because cleaning did not precede the sterilization procedure. Now, the
amount of salt and serum is not really that high. The amount of salt is
.65 percent. The amount of serum is 10 percent, but failure in the
absence of any cleaning.

Next slide. If we use, essentially, a lumen device and do the same
experiment, next slide, we see, essentially, 60 percent failure. Again,
we\'re failing to sterilize instruments that are contaminated with easy
to kill microorganisms, because we failed to clean them. And it\'s not
really the lumen device that is causing the problem, because we see here
in the absence of serum or salt, those organisms are killed. So the
failure to clean allows the survival of easy to kill microorganisms in a
sterilization process, such as low temperature sterilization.

Next slide. So all technologies have limitations. Salt and serum provide
protections for spores and bacteria, and salt and serum with a lumen
carrier even provides extraordinary protection.

Next slide. Now, let\'s look at the issue of cleaning and let\'s look at
spores.

Next slide. Let\'s just put some spores on a stainless steel scalpel and
we\'ll see on the next slide. We\'re going to put about 10^6^
geobacillus stearothermophilus spores on this stainless steel scalpel,
and then we\'re going to put that scalpel in a low temperature
sterilization technology, such as hydrogen peroxide, gas plasma in the
absence of cleaning, and we can see complete failure here. 60 out of 60
positive stainless steel scalpels.

Now, let\'s look at the next slide where the only thing we did is we
again put the spores on the stainless steel scalpel.

And then the next slide. All we did was place the stainless steel
scalpel into either distilled or tap water for 60 seconds, just placing
it there for 60 seconds, taking it out and then putting it into low
temperature sterilization technology.

Next slide. You can see that there is complete success, a complete
ability to kill microorganisms to include spores just because of a
static soak. Of course, you can see here a very light rinse also was
successful.

Next slide. Here, we are going to, essentially, try to identify why this
is happening, and what we\'re really looking at is what is going on as
far as chloride, protein and spore concentration by just doing that
static soak.

In the next slide, we see, essentially, in a matter of seconds we see
the salt, protein and spores released from the fetal bovine serum dried
on stainless steel blades and placed into deionized water at room
temperature. In a matter of seconds, essentially, you get significant
reductions in salt, proteins and spores.

Next slide. Now, so what we found is, essentially, inorganic, organic
and microbial contaminants on the device are dramatically reduced during
washing and, of course, there is a significant reduction of spores.

Next slide. Well, let\'s see if that is effective for steam
sterilization. Right now, we\'re really talking about these low
temperature sterilization technologies. Does the same thing happen with
steam sterilization? This is a study from Doyle and Ernst in 1967 where,
essentially, all they did was monitor the effect of spore occlusion and
calcium carbonate crystals in inactivation in steam dry heat and
ethylene oxide sterilization processes.

They were just inoculating 10^3^ or 8 times 10^3^, bacillus subtilis
spores, and let\'s see what happened because of the spore occlusion and
calcium carbonate. Here, we see steam at 121 degree centigrade in the
unoccluded spores, the biological challenge. It only takes 10 seconds.
You can kill 10^4^ in 10 seconds, no time. But in the presence of the
calcium carbonate, to kill that 10^4^ took 150 minutes. For dry heat,
it\'s three and a half hours. In the presence of the calcium carbonate,
it\'s 50 hours.

Next slide. So, essentially, a number of things have been found, that is
contact with water or cleaning for just a short period of time rapidly
leads to the dissolution of crystals, of course, removed microorganisms
and, of course, also has an effect on protein elimination. And, of
course, minimal cleaning eliminates the effects of these salts, which
effect the effectiveness of sterilization processes. And simulated use
tests that do not include washing would not represent conditions that
exist in clinical use situations.

Next slide. And this is, essentially, what you see in electron
micrograph. If you look at, essentially, .75 sodium chloride in the
presence of spores, you see the salt crystal, essentially, occluding the
microorganisms from exposure and, essentially, cleaning dramatically
effected those results.

Next slide. The point that we need to make for all these studies that
have involved prion inactivation is that you can clean without
sterilization, but you never can sterilize without cleaning. That is a
point, a principle, that is known to every professional in health care,
certainly, every professional that is involved in reprocessing
instruments.

Next slide. So the conclusions would be all sterilization processes are
effective in killing spores. Salts favor crystal formation and impairs
sterilization not only for low temperature sterilization, but also high
temperature sterilization. Cleaning removes salts and proteins and must
precede sterilization. Failure to clean or ensure exposure of
microorganisms to the sterilant could affect the effectiveness of the
sterilization process. We say repeatedly if the organism does not have
exposure to the germicide or the sterilant, inactivation will not occur.
And, of course, these salts and protein materials and possibly other
environmental conditions to include surfaces affect that exposure. And
lastly, CJD inactivation studies should be consistent with actual
clinical practice.

I think we have done what we said we were going to do. We have looked at
the recommendations from the U.S. We have talked a little bit about
methodology and how methodology affects results not only for hard to
kill organisms such as spores, but easy to kill microorganisms such as
bacteria.

Next slide. I thank you very much for your attention.

CHAIR PRIOLA: Okay. Thank you, Dr. Rutala. Are there are questions from
the Committee or from Dr. Taylor? Would you like to make a comment?

DR. TAYLOR: Just a couple of quick comments. A very nice talk and very
much to the point, I think. Two comments, one is mentioned that
generally, the washing procedure is usually pretty effective in taking
off bacteria and spores, etcetera. The one comment here is that one
might anticipate that TSE infectivity might, nevertheless, be somewhat
more adherent to instruments because of the hydrophobicity of the PrP
protein.

The other comment is that there are concerns about damaged autoclaves by
hydroxide, but that is certainly not inevitable, because it depends on
the grade. The commercial company that we asked where do you dispose of
animal carcasses, in reactor vessels with hot hydroxide, have had
vessels running for many years now and they have certainly subjected
these to x-ray analysis, etcetera, and they are absolutely fine.

DR. RUTALA: To your two points, I certainly agree with the first point.
Certainly, the data that I presented, of course, are non-prion proteins,
as well as microorganisms and, certainly, the same type of analysis
needs to be done with prion proteins, and I support that work.

In regard to the second point, the effect of sodium hydroxide on
sterilizers, certainly some sterilizer manufacturers have threatened the
owner of the sterilizer that in the hospital, if they use sodium
hydroxide in the sterilizer, they will nullify the warranty, which, of
course, affects the utilization of sodium hydroxide.

But there are ways, as you probably know better than I do, to limit that
vaporization and contain the vaporization with, for example, containers
that have lids. And certainly, Dr. Asher knows more about that, and
possibly that can be discussed.

CHAIR PRIOLA: I have one very quick question before Dr. Rohwer makes a
comment, and that is you stress very strongly that instruments are
always kept wet, so that you don\'t have this problem of material drying
on the instrument and then perhaps adversely affecting its ability to be
sterilized.

During a surgery \-- I mean, I know that when I do my little surgeries
on mice that as you\'re doing it, stuff does dry on the instrument just
as you\'re poking around, so how is that dealt with?

DR. RUTALA: Well, many times \-- it is dealt with in different ways.
Many hospitals, essentially, don\'t even take the surgical instrument
and put it on a dry tray. They very commonly place it in a basin, which
has, for example, saline or water or possibly even a germicidal agent,
so it doesn\'t go into a setting, which is going to allow dry fluids and
tissues to be achieved.

And then, of course, the other issue is that there is sometimes
precleaning before it is sent to central processing. Central processing
does not want instruments that are contaminated with tissue and blood.
They won\'t accept instruments that are contaminated with tissue and
blood. Sometimes, there is also a washer sterilizer that, essentially,
is a precleaning procedure before it goes to central sterilization.

So different hospitals do different things, but the one thing in common
is there is an effort to keep it wet, and there is an effort to keep it
clean, because most central processing areas won\'t accept surgical
instruments that have dried tissue on them or bloody instruments.

CHAIR PRIOLA: You know, I understand that. I guess my point was more
during the procedure as you use the instrument. Just as you\'re using
it, it\'s going to air dry, because it is exposed to the environment,
and so you can\'t keep it wet the entire time. I mean, you\'re just
going to have some dried material that will probably be taken care of
possibly by the cleaning and whatnot.

DR. RUTALA: Yes.

CHAIR PRIOLA: But some drying will occur no matter what you do.

DR. RUTALA: And by immersion in the bath, but some drying, depending
upon the level, of course, will occur.

CHAIR PRIOLA: Okay. Dr. Rohwer, what was your comment?

DR. ROHWER: Yes, I have a couple of comments and also would like to get
Dr. Rutala\'s opinion on something, and that is first, I would highly
advise not autoclaving with the lid on the vessel, and I am also
mystified by this concern about sodium hydroxide vapors. As far as I
know, sodium hydroxide has no measurable vapor pressure, and a properly
operating autoclave should not be aerosolizing it either, because it
shouldn\'t boil on the way down. That is my first point.

But what I would like to have you address is this issue of
cross-contamination at the level of washing, because that is our major
objection with that approach. We\'re talking about an agent that is very
difficult to get rid of. If it gets spread around the laboratory and the
environment then, we just couldn\'t tolerate that. You know, we don\'t
want it in our sinks. We don\'t want it on our surfaces, etcetera. And
so how is it that you deal with the eluates and the washers that come
off of a set of instruments, which you know have been exposed to a
Creutzfeldt-Jakob Disease patient, for example? How are the washers
sterilized? How do you dispose of that?

It seems to me you create a cascading level of problems that have to be
dealt with, and I am absolutely willing to concede all the points that
you are making, except that in the case of this particular agent, it is
very difficult for me to accommodate this idea of spreading this stuff
around, exposing myself to it willingly before it has actually been
decontaminated.

DR. RUTALA: Well, to your point, certainly, this practice has been
employed for a number of years, and I think it has been practiced for
the following reasons. One, of course, in a decontamination area in
central processing, the persons in that area wear personal protective
equipment. They wear gowns, gloves, protective masks. Second is that
nearly every hospital where decontamination takes place in central
processing, it\'s a closed unit. It\'s a washer disinfector completely
closed. That is there is no aerosolization of droplets, of fluid that
are related to the washing procedure. It\'s a closed procedure.

The closed procedure, of course, has many steps to it. Some of them are
the use of enzymatic detergents, high temperatures, rinses and so forth.
And I guess the third point related to that, protective apparel, a
closed system, is the issue that while there is a recognition that some
prion proteins may go down the drain connected to a sanitary sewer, we
don\'t believe that that\'s the only source of prions reaching,
essentially, the sanitary sewer system.

And to that point also, prions, of course, are unlike many other
infectious diseases, are not transmitted by direct contact, indirect
contact, droplets spread, airborne or the environment. So I am not sure
what the level of concern is as far as transmission. It certainly
wouldn\'t be, as I look at the issue, transmission to health care
workers, it could possibly be contamination of the environment, but I
believe that there is other forms of contamination in the environment
outside that setting.

CHAIR PRIOLA: Dr. Bailar?

BOARD MEMBER BAILAR: Very nice talk. I do have a question about this
very simple straightforward slide you had on the decreasing order of
resistance of microorganisms disinfected in sterilants. You didn\'t go
into the details of that, but I suspect it\'s ranked on the basis of
things we know are effective at the susceptible end, that is the
bacteria and the enveloped viruses.

Is anybody looking at other kinds of agents, you might call them
unconventional agents, that may not be very effective at that end, but
might be pretty good with prions?

DR. RUTALA: As far as that slide is concerned, you know, it is a general
slide. There is an exception to that slide as it pertains to germicidal
agents and particular groups of microorganisms, but it is a general
slide. As your question intimates, most of the data, and there is
literally hundreds of papers that support that slide, and most of the
data is with the types of germicides that you have seen today and in my
presentation and in other presentations. That is the conventional, not
the nonconventional germicides and sterilization processes, products
such as the alcohols, the phenols, the ethylene oxide, the steam
sterilization and dry heat and so forth. Does that answer your question?

BOARD MEMBER BAILAR: Yes, it leaves me wondering whether it might be
worthwhile for somebody to try some of these things, but I am no expert
in this field.

DR. RUTALA: Well, to your point, I think that there are people
interested in that.

CHAIR PRIOLA: If there are no more questions, we have two more speakers,
but I think we should take maybe a 10 minute break. We\'re about 15
minutes behind, but let\'s take a 10 minute break and reconvene at 5:10.

(Whereupon, at 5:06 p.m. a recess until 5:18 p.m.)

CHAIR PRIOLA: If I could have the Committee members take their seats, so
we can conclude this session. Okay. Our next speaker is Captain Edward
Rau, Environmental Health Officer for NIH, and he is going to discuss
infectivity of air emissions and the incineration of scrapie tissue.

CAPTAIN RAU: Thank you very much. Unfortunately, our only TSE expert in
our group, Paul Brown, couldn\'t be here today. He is on some kind of a
hardship assignment in southern France at the beach right now, so he has
left that up to me to take care of.

The other disclaimer is that the results that I am going to present here
are really very preliminary. Our experiments are still in progress. The
paper is not written. None of the data is published. So with that,
we\'ll go ahead here.

Could I have the next slide, please? I don\'t need to elaborate on all
of the difficulties there are in inactivating the prion agents, and that
the resistance to thermal inactivation is, of course, highest under
conditions of dry heating. And that has led some concerns about even
incineration being an effective technology to dispose of certain TSE
waste.

Sitting here this afternoon, I think I have a new definition for
incineration. It\'s that process, which incorporates all of the things
that make inactivation difficult to do. We start out with a material
that has been smeared and mashed around by all the handling of the
medical waste process. It has not been precleaned. In fact, it is, in
some cases, pure dirt. Then we\'re going to take that into a process,
which begins by a drying and probably fixation step, and maybe melt a
few varnish like materials over it before we really get into the
combustion process.

Next slide, please. As you are aware, incineration is still the
technology of choice for disposing of most medical waste that contain
TSEs, and it is also being used to dispose of large volumes of animal
products, contaminated carcasses and so forth, some of which are still
in large quantities in storage. The potential for TSEs being in
emissions from combustion processes is of public concern, and has
received very little investigation so far.

Next slide, please. We have published some previous experiments
documenting the unprecedented level of resistance to thermal
inactivation, both crude brain tissue and purified PrP from the 263
scrapie infected hamsters. That included a partial inactivation after
heating for 300 degrees for 15 minutes, and several transmissions after
actually ashing brain material at 600 degrees C. And at those higher
temperatures, there were similar patterns of resistance in both formalin
fixed and non- fixed tissues.

Next. The objectives of our experiments in progress are first to confirm
the results from our previous study. Some people were a little bit
skeptical about the transmission after 600 degrees C. Others might have
even labeled it science fiction. And secondly, we wanted to investigate
the potential for transmission via the air emission that might come from
a medical waste incinerator.

Our previous experiments were rather primitive in that we merely headed
brain tissue macerate in vented crucibles. The new experiments, we are
actually going to simulate the conditions of humidity and the gas
mixtures and so forth that occur in two types of incinerators.

The first are the reducing environment or starved air incinerators.
These are the most commonly used type of incinerator in the United
States. A synonym for that is the controlled air incinerator. The other
situation we wanted to look at was an oxidizing environment or referred
to as a normal or excess air incinerator. And in this study, we repeated
the temperatures that were used in the previous study, 600 degrees and
1,000 degrees C.

Next. Materials, our tissues samples were cooled, hamster brain tissue
macerates from terminal animals with the 263 scrapie strain, about 10
logs of infectivity per gram, and for controls we had tissue from normal
animals. The incineration situation was referred to as a Lindberg
Furnace in a quartz reactor tube, the removal of the specimen crucible
and holder. The gas supply coming into the incineration unit was normal
air or nitrogen with flow and humidity controls. And then coming out of
the unit was an impinger train and terminal filter for collection of the
air emissions.

Next slide. This is a photograph of the main part of the simulator. It
all fit in a large chemical fume hood. The incoming gases come into the
furnace here. The quartz tube is contained inside of the Lindberg
Furnace. This is a pyrometer. And then the outflow coming out of the
combustion process goes into this impinger system, a series of
collectors, the first one being ice water bath. The second one, dry ice,
and then out through a terminal filter.

Next. This schematic gives an idea of the inside of the reactor tube,
inside of the furnace. The tube is, approximately, one inch diameter,
all quartz construction. At this end, we can remove the plug and insert
into a thermocouple to directly measure the temperature or we can insert
the sample on a glass rod, which is housed right here.

The gas flow comes from this direction in the top. It passes the sample.
Exists through a ball joint and then on to the impinger train. In
designing this, we tried to ensure that all the components in the system
were inert. We used quartz and teflon joints as the materials.

Next. This schematic shows a little bit more information about the
impinger system. It\'s a rather challenging design, because we really
didn\'t know what we were trying to trap coming out of this process.
Again, we have the ice water bath and the crushed dry ice bath following
into a cartridge filter and exhausting into the hood.

Next. The methods began by introducing a one gram sample of the brain
tissue into the reactor tube. We incinerated that for 15 minutes at
either 600 or 1,000 degrees in either normal air or starved air
conditions. Following the process, we collected the air and air emission
samples separately from each run, and then replaced the impinger train
between each run. We didn\'t have enough reactor tubes to use one for
each experiment, so we disinfected those with bleach after each test.

Next. So each experimental run gave us three different samples. We had
the ash residue that was collected in the crucible. The small amount of
residue that formed in the reactor tubing as it exited the quartz
reactor and came out cooled down at the border of the furnace, and then
the emissions collected in the impinger traps.

Next. This gives you the array of samples that we collected. We ran both
normal and infected tissues in the two different gases, two different
temperatures, and for the infected material, that gave us three
different samples. We combined samples for two of the normals and we did
not run some of the exit tubes and traps on those. The reason for that
is simply economics. We\'re dealing with about 500 animals to be
maintained here, a great cost and time.

Next slide, please. The bioassay method, we concentrated the samples of
the ash and emissions from each test into, approximately, a one
milliliter volume in saline, and that was intracerebrally injected into,
approximately, 30 Wingling hamsters. That is about 3 hundredths of a
milliliter per animal. So the entire emission from each test was
injected. We\'re not taking a sub sample out of those emissions. The
entire emission from each burn was injected into animals.

Animals, of course, were segregated by test group. We observed them over
12 months for symptoms and then examined all of the brains for the
presence of prion proteins by Western Blot testing. That testing is
still in progress on the negative appearing animals.

Next. Results, we had no transmissions from the controls. There was some
possibility, my commentor thought, we might be able to get some kind of
symptoms as artifacts of this trauma and injecting into the animals,
this residue. We just wanted to rule that out. We had no transmissions
from any of the materials collected at the 1,000 degree C burn, and we
had no positives from any of the residues collected in the impingers or
the end of the reactor tube.

However, we did get two transmissions, and these were after very long
incubation time from the ash from the crucible from the 600 degree group
in normal air. The asymptomatic animals, again, we\'re still testing
those for silent infections. We\'re not finished with that, so we have
to call our results preliminary, at this point.

Next. Conclusions from the experiment. First, that the results were very
similar from the previous study showing that there is, apparently, a
threshold transmission from tissues at about 600 degrees C. The low
transmission rate and very long onset time for the symptoms suggest,
again, that that is the extinction temperature or very near it. We found
no evidence of infectivity in the air emission samples.

Next. Speculate a little bit about what the environmental implications
of this are. First, we did not see any evidence of transmission in the
air emissions, so it\'s probably unlikely that will have a possible
emission to the air from a properly operated medical waste incinerator.
It is possible that some survival of the agent could occur in ash if
there is not enough penetration of the temperature and time of exposure
in the ash bed.

I wanted to kind of put this in perspective a little bit though, because
I think it\'s a very low potential for transmission. First off, as Dr.
Taylor said, we don\'t see environmental sources in transmission going
on. Secondly, what we are seeing these two positives on is a simulated
burn load that is pure material with 10 logs of infectivity per gram,
and we\'re collecting virtually of the residue from that and injecting
it into, approximately, 100 animals. So there is a very low level of
infectivity in that material that is coming out.

Probability of survival in ash not only depends on a lot of factors, the
load density, the turbulence, the type of equipment, other operational
factors. And as we heard earlier today about the importance of context,
I really can\'t imagine a more complicated context than a medical waste
incinerator and all the combustion and mixing and reactions and things
that are going on inside of that process.

Next. Our colleagues at EPA felt that these factors probably would be
the most important in increasing the potential for prion survival in ash
from medical waste incinerators. Under normal conditions, there are
certain design factors, particularly in the grate oriented designs that
might allow some of the material to not be treated for 15 minutes. It
falls through the grates or it somehow gets passed on through the system
faster than the nominal residence time for the solids.

Particularly, as things are just inserted into the incinerator, you tend
to get a boil off of some of the material, a flash burn. That can be
carried over very quickly into the second chamber. The other factor is
that the ash bed temperatures often may run 100 degrees C lower than the
actual air temperature.

Reported temperatures for incinerators refer to the air. That is what is
being monitored, and not the actual temperature in the ash. Under
abnormal conditions, a lot of things can really go wrong, cold start up
conditions, overloading, inadequate control of the under fire air flow.

Next. We tried to compare what our experimental conditions were with the
conditions in actual medical waste incinerators and some other types of
processors used for bone meal products, and the most common incinerator
in the U.S., the controlled air or starved air type of incinerator, in
that primary chamber, you have temperatures of about 760 to 980. If you
subtract about 100 degrees from that, allowing for some cooler
temperatures in the ash, we\'re right on that threshold of survival that
we saw in our experiments.

The secondary chamber, which mostly sees the pyrolysis products and not
the ash is usually well up into that 1,000 degree temperature. I don\'t
have much information on the temperatures in the excess air
incinerators. They are probably quite variable because of the way that
process is run. The secondary chamber is, again, quite hot, sufficient
probably to inactivate prions.

There is some information on the other types of burn units that are
being used in Europe for disposal of the meat by-product material. One
of the articles had indication that there is actually a measurement of
temperature at the ash grate in this 800 to 1,000 degree range, which
looks pretty good for inactivation.

Next. There are other possible incineration options. If we get into a
situation where we have a large amount of material to dispose of, the
mass burn municipal waste incinerator in the United States operates at
about 1,000 degrees, so that would likely inactivate prions.

Western Europe is looking at a variety of other types of industrial
incinerators, fuel burners and so forth. Again, they have some high
temperature and residence times. In one system, a holding time of 30 to
40 minutes, which is very encouraging.

I think that\'s the last slide. Next. I believe that is the last slide.
Again, our results are very preliminary. So far, all of the testing on
the negative appearing animals is confirming that\'s the case, but
we\'re not quite finished with that yet. Any questions?

CHAIR PRIOLA: Any questions from the Committee? All right. Thank you
very much, Captain Rau.

CAPTAIN RAU: Thank you.

CHAIR PRIOLA: Our final talk of the \-- oh, I\'m sorry. You had a
question? Oh, sorry. Yes, go ahead.

DR. ROHWER: One issue I had with the original study was that the
thermocouples were not actually in the sample, and the sample was loaded
wet, and it wasn\'t clear here whether you\'re starting with wet tissue
or dry tissue, and where the temperature measurement is actually being
made, vis-a-vis, the sample. And the reason I bring this up is that a
wet sample will not spend as much time at 600 degrees as a dry one,
because you got to boil off the water first, and that could actually
take some time.

CAPTAIN RAU: We did start with wet samples, wet tissue samples. The burn
time is 15 minutes, however, in here, so I think we\'re probably
spending most of that time at temperature. With regard to the
thermocouple, before each sample was inserted, it was measured, then the
thermocouple was withdrawn. We still have a pyrometer on the outside of
the tube that we\'re confirming temperatures with, and that is really
the best we could do. There is just not a way to have the thermocouple
in there and be able to insert and withdraw a sample out of there. I
agree with your boil off concern, but that is also real world, what\'s
happening in the incinerator.

DR. ROHWER: In the original experiment, there was a thermocouple between
the crucibles, so I took that to mean that there are thermocouples that
can survive those kinds of temperatures. Is that incorrect?

CAPTAIN RAU: Yes, the problem is getting the output out of the burn
chamber. It was a design issue. But in the first experiment, the
thermocouple was right adjacent to the crucible and we were able to
measure that in the muffle furnace.

CHAIR PRIOLA: Okay. Thank you. I think we\'ll move on to our last
speaker who is Dr. David Asher from the FDA, as well as Dr. Brown, Dr.
Stanley Brown. Oh, actually, you\'re going to start.

DR. BROWN: Actually, the last team is \-- I am the rigger coming in from
CDRH. Could I have my first slide, because it tells the whole story?
Let\'s see, it worked on my computer. It was created on Terry\'s
computer.

Well, anyway, I\'ll start. I\'m Stan Brown. I am an engineer from the
Center for Devices. I will present the first half of the team effort
between myself and David Asher\'s group, which was funded by the FDA
Office of Science, and these data, some have been published. Some are
preliminary. Some are in manuscripts in preparation, and this is not
good news on the screen.

Basically, what we were doing in my side of town was to look at four
questions. The first question is can you safely autoclave in sodium
hydroxide without wrecking your autoclave? The second question is what
are the effects of the WHO protocols on surgical instruments? The third
part of that was to develop an experimental instrument that could be
used in a simulated instruments contamination study that would be
compatible with the hamster model that David Asher has. And the fourth
was to do some \-- there we go. Okay. Let\'s click through here.

First of all, instruments from CDRH, we were thinking about primarily
reuse, as you have heard, or reusables, but there is also growing
concern about these things called SUDs or single-use devices, and with
the law we are now reevaluating how we assess the reprocessing and
validation of some of the reusables, particularly those that are
neurological or other type of tissue contact. From the CBER point of
view, as you know, we are talking about contact of instruments that you
use for tissue preparation.

Next. Disclaimer. We developed these methods, because they fit within
the financial constraints and the laboratory constraints. In our
laboratories, these presentations do not constitute a regulatory
endorsement for these methods. They are simply methods we thought would
get answers that we\'re after.

Next. You know all about this story and we\'re primarily interested in
the sodium hydroxide autoclaving phase and the soaking in sodium
hydroxide or bleach.

Next. So if you go to the CDC website, you will see that there are a
couple of warnings. One is that if you autoclave in sodium hydroxide,
you wreck your autoclave and two, if you soak in bleach, you will wreck
your instruments. And these are based on the studies that we started a
few years ago, and I think CDC may actually have now inserted the
reference from our work on that.

Next. Again, there are the four questions that we are trying to answer.

Next. The first one has just been published in the [American Journal of
Infection Control]{.underline} with Kathy and myself.

Next. She did the work and I wrote the paper, so I got to be first
author. And again, the question is the autoclave manufacturer said if
you autoclave in sodium hydroxide, you don\'t have a warranty on your
autoclave. Knowing, of course, this must be done in a gravity
displacement, so it doesn\'t fit in the standard central storage big
prevacuum type autoclave. It has got to be controlled with a liquid
cycle.

Next. Two approaches. One was we would put a liter of sodium hydroxide
in a pan and cover it. Two, we would put some sodium hydroxide in a
beaker and put the beaker in a pan and cover it. And then we put it in
an autoclave. We did repeat one hour sterilizations, and we did these at
134C just to be a little more extreme, and we did them for an hour, up
to five cycles, and we put pH paper inside, outside, everywhere. We put
pH meters inside and outside, and it is a closed system for the little
tabletop with six liters of water reservoir. We put it through five
cycles to see what happens to the water in the reservoir to see, again,
what happens to the autoclave.

Next. And we got thinking about pans and lids. This one probably, the
condensate will get on the top and drip down and wreck your autoclave.
This one, it might wreck your autoclave and it might drip inside. Some
of them have little nipples or construction bars. To me, that is a Black
Iron Dutch Oven where these drips, so that they roast while you\'re
doing it. And then you have got some that actually have gutters that
contain the lid within the pan. So part of this is what kind of pan and
lid design you have.

Next. The two that we used successfully was a Nalgene Instrument tray
shown here. This was filled with a liter of sodium hydroxide and closed.

Next slide. And if you look here, you can see this is a drain on that
gutter, so the condensate goes back into the pan. It doesn\'t go out.

Next. And there you see the lid that is fully contained within the
gutter and, of course, this has been used for years. It\'s for control
of human waste and biohazards.

Next. The other type of pan was a Lid (D), which has a lip on the lid.
It also has crossbars that act as condensate drip spots.

Next. And the results of this were no pH changes outside the
containment. Inside the lids were very caustic. The bottom of the pans
were very caustic. There was lots of vaporization, condensation going
inside, but it was all contained within the containment vessel. So we
conclude that if you use this kind of \-- if you use the right kind of
pans and lid, you can do it without wrecking your autoclave. And, of
course, those of you who have been doing this in the lab for years know
that. Obviously, hot caustic is hot and you have to be careful. It
probably cannot be done in a standard central storage autoclave and it
may require larger approved type pans.

Next question. What do these things do to your instruments? Next. What
we did was we bought surgical instruments from Roboz, which is a medical
device supplier in Rockville, and we bought lab stuff from VWR. Some of
them are labeled Germany with CE marks and some of them were labeled
Pakistan, and there are some members in the audience who will appreciate
this. We put them through repeat cycles of the WHO including autoclave
and water.

Next. And here you see some carbide tipped needle holders. This one has
been through five times autoclave and sodium hydroxide, five hours in
sodium hydroxide. There is a little bit of blackening in the box joint.
This one was soaked for one hour in Clorox, and you can see there is a
tremendous amount of corrosion going on at the box joint and around the
carbides.

Next. These had beautiful gold handles, high quality instruments, and
that is one hour in bleach. So if you got gold handles, don\'t bleach
them. This is five times gold handles autoclaved in sodium hydroxide. It
looks fine.

Next. This is a German pair of scissors versus a Pakistani pair of
scissors, five times in sodium hydroxide. You can\'t photograph shiny,
but this is shiny. It just looked great, and this looked really dark and
dingy.

Next. This is Germany versus Pakistan, and you notice the Roboz label on
this thing as stainless steel, and this tubing clamp around this weld
really took it with the Clorox. This tubing clamp after five hours in
bleach, this is the 6 percent, which is what, 2,800 parts per million.
It looked fine. So some go, some are fine.

Next. So the conclusion of this, and I didn\'t show you any pictures.
Titanium really does not like sodium hydroxide, and this is well-known
in the material science literature, as well. Soaking in sodium
hydroxide, they couldn\'t care less, none of them. Soaking in bleach,
some did fine, some didn\'t. The problems were the gold handles and the
welds. But the important message here is if it\'s going to corrode,
it\'s going to do it first time. So you don\'t have to do a long
experiment. If you put it in Clorox and it comes out rusty, you know
it\'s going to rust and you go on and find a better instrument.

Next. So Part 3. We wanted pins and this is part to lead into an animal
model of the simulated instrument for David\'s studies. He was using a
25 gauge needle on a half cc syringe in his animal work for injection,
and we wanted to make pins instruments that were like that, but we also
wanted to be able to suspend them over 96 weld plates, so we could do
serial dilutions of bacteria, viruses in brain homogenate, and the
system needed to be autoclavable.

Next. So there is the syringe needle that he was using. That is a good
old copper penny, and what we did was we took Eppendorf tips. My wife is
a microbiologist, and so is Kathy. We took half millimeter stainless
steel pins. We took some epoxy. We used the Eppendorf to draw the epoxy
up into the tip, stuck the metal pin into it, hung it in a rack, put it
in an oven to cure and now, you have things that can be autoclaved, and
it\'s the same size as the needle that is used. So from an ergonomics
point of view, it\'s something that he would feel comfortable with, I
think.

Next. And here, you see the setup. This is your standard Eppendorf rack
with modified stainless screws to adjust the height, and the pins were
sitting in the wells of a 96 weld plate, and Kitty, that after a little
bit of practice, she could actually get all the needles into all the
holes, right?

Next. Finally, we wanted to do some preliminary of adhesion of both
blood and tissue and looking at WHO protocols, and one question was what
about damage and adherence? So we were using stainless steel pins and we
also made pins out of piano wire, which really did not have a good time
in Clorox.

Next. Pins are placed in a rack and stuck into a slab of liver for an
hour, and then we left to dry as a worst case. The pins were stuck in a
96 weld plate in sheep\'s blood for an hour, and then they were left to
dry. We went through ultrasonic cleaning, which the standard protocol is
60 degrees C with an enzyme cleaner. We autoclaved in sodium hydroxide.
We soaked in bleach and we got unclean controls.

The results were the unclean ones, that the protein was more adherent
from liver than it was from blood, and the amount was using Bradfords
reagent. It\'s about the equivalent of \-- our minimum was one
microliter detection limit. Damaged pins did not seem to be more
adherent, so that the blackening in the box from autoclaving and sodium
hydroxide is probably not a problem. Repeat exposure did not show
accumulative effect.

Next. So then Kathy wanted to do some bacteriology, and she soaked them
in a solution of staph epi. for 24 hours, let them dry and did the same
kind of cleaning things, and then stuck them into an agar in a test tube
and incubated for 24 hours.

Next. Lo and behold, autoclaving in bleach killed everything. So we had
to try some modified WHO. So we dropped the sodium hydroxide autoclave
and the ultrasonic cleaning was done at room temperature.

Next. So what we found was that only the pins and bleach showed no
growth, but, of course, we don\'t know if we cleaned it or we just
killed them. The other produce showed fewer protocol than the untreated
control, but bacteria was still present. And then the question is are we
cleaning or are we just killing?

Next. And we tried some SEM work, and you can see a little small column
of staph epi. here on the uncleaned tip. It was very unconvincing in
terms of whether we really were cleaning or we were just killing.

Next. So our conclusions were, first of all, yes, you can safely
autoclave in sodium hydroxide with the right pan and lid. Some WHO
protocols can damage some instruments. Discoloration does not seem to
impair function or cleaning. The bacteria leave a lot of unanswered
questions, and the questions for prions, of course, we don\'t touch them
in CDRH, that is David\'s role.

So next, and I will turn the podium over to David to talk about his
hamster studies.

DR. ASHER: Thanks. You can go right to the next slide, please. Thank
you. I can\'t believe that it\'s almost 6:00 and we\'re still giving
talks. Quite a few people in CBER participated in TSE related
activities. These studies are really involved only people in CBER in my
own little group, and especially Kitty Pomeroy who I think is still here
in the back. Without her holding the whole enterprise together, there is
no way that we could have done this.

Next slide, please. And among the staff at CDRH, of course, Kathy
Merritt and Stan Brown, who has just spoken, and I don\'t believe that
we would have gotten the funding to do what we have been able to do
without Stan\'s efforts.

Next slide, please. We have developed two simple methods to evaluate
methods for decontamination of TSE agents dried on surfaces. In this
talk, I am going to concentrate on two regimens that more or less
replicate recommendations of the World Health Organization consultation.
We used two general models for both of which 263K scrapie was the agent.

The first model was a modification of a method for evaluating virucides
that was described years ago by John Chen of the Environmental
Protection Agency. He dried viral agents onto glass cover slips, treated
them and then assayed residual virus.

The second method was stimulated by two reports from Charles
Weissmann\'s group, and Professor Weissmann will speak here tomorrow
morning. They dried scrapie agent onto steel wires implanted into mouse
brains. We didn\'t do that, but as you saw from the devices that Stan
showed you, our model was very similar.

Years ago, we had done a couple of experiments with model squirting
scrapie through actual hypodermic needles, but it was very cumbersome,
and we never followed up on it. We have used the first method.

Next slide, please. The first method for many years, simply dropping
suspensions of scrapie infected hamster brains onto glass cover slips.

Next slide, please. Then they are dried in a petri dish in a hood.

Next slide. And then they can be exposed to any number of disinfectant
or decontamination regimens, here potassium permanganate solution that
can be autoclaved, that can be soaked. You can do all kinds of things
with it.

Next slide, please. Then they can be ground up and supernatant fluid
assayed. Now, we used plastic pipettes and tubes for the whole
procedure, because they are disposable. We don\'t have to worry about
potential carryover of infectivity. Although, the original method
described by Chen used 10 brack tissue grinders of the kind seen here.
So the slips are ground up in an ml of diluent.

Next slide, please. The glass is allowed to settle out, and then the
supernatant fluids are assayed for infectivity by intracerebral
injection of hamsters.

Next slide. If the hamsters get sick, their brains are removed and then
they are checked for protease resistant prion protein as evidence that
scrapie agent was present and was not eliminated by the decontamination
regimen.

Next slide, please. For reasons that may become clear at the end of this
talk, I have about six minutes left, we think that it might be useful to
do immunohistochemistry on some of these brains, as well. Although, we
haven\'t done that yet.

Next slide, please. One advantage of the method is that you can rid of
residual toxic disinfectants, Robert Somerville talked about that
problem this morning, by simply rinsing the cover slips in distilled
water to get rid of things like Clorox, which is really terrible for
assay animals.

As you see here, this is sort of an upside down dose response curve.
It\'s hamster survival times plotted against the dilution of supernatant
fluid. And here are three curves, one for unsoaked or dipped slides, one
that has been soaked in water, and the other that was soaked in water
and then dipped in water a second time. And you can see that they are
virtually superimposable. None of the infectivity appeared to come off
on this exposure to distilled water.

Next slide, please. And it was using this method that we first
demonstrated the resistant fraction of infectivity that survived drying
on glass and then prolonged exposures to steam autoclaving at elevated
temperatures.

Next slide. To investigate some of the performance characteristics of
the model, you can imagine doing many, many assays in hamsters is
extremely cumbersome. We tried drying specimen samples of polio virus
and two other viruses suspended in brain onto glass, and then titrating
multiple samples. The results summarized here suggest that the agreement
from test to test and day to day was reasonably good, but that there was
enough variability, so that a controlled titration really should be done
in each test. And it might even be reasonable to consider putting in a
test decontamination, a reference decontamination treatment. Although,
we have never had enough resources to support that.

Next slide, please. When we began to work with the scrapie brain
suspensions dried onto the steel needles that Stan Brown has just showed
you, we decided first to do some similar preliminary studies with
conventional virus suspended in saline containing 10 percent brain
extracts to get some idea of how the scrapie agent might be expected to
behave, so we wouldn\'t waste months and months on a model that
wouldn\'t get enough infectivity on.

But we were surprised when neither polio virus nor porcine parvo virus
suspended in brain appeared to stick to the steel needles at all or at
least we couldn\'t detect any of them in cell culture assays. We had no
trouble getting them to stick to glass, but we couldn\'t find detectable
porcine parvo virus or polio virus dried onto steel needles.

So now wanting to take a chance that we would set up our test with the
limited funding available and the find that we hadn\'t had enough
challenge agent stuck to the steel needle, so we decided to suspend the
263K scrapie hamster brain in a normal brain paste, and we used a calf
brain as the source of the normal brain material.

Next slide, please. I must say we decided to check out our concern. We
had enough hamsters to do a rough titre of 263K scrapie diluted in
phosphate-buffered saline without any additional brain material, and it
appears that the scrapie infectivity in the saline suspension did stick
to the steel wires. You will notice we get positives out to a dilution
of 10^-5^, so that the behavior of the polio virus and the porcine parvo
virus does not appear to have predicted the behavior of the scrapie
agent.

But the studies that I am going to summarize in the next few slides use
scrapie infected brains as a paste to charge the needles. The glass was
charged in the way that I described previously with saline suspensions.

Next slide, please. So let me summarize for you the general design of
the efforts. We looked at two variations of two kinds of decontamination
regimens that generally resemble those recommended by the WHO
consultation, and then after that, I will add some other results that we
thought you might find of some interest.

First, as we have mentioned, infected brain is dried on the objects,
either a saline suspension on glass slides or tissue paste onto steel
needles. Then come the decontamination steps, which are either a
chemical soak in one normal sodium hydroxide in the autoclave for 30
minutes or a soak in sodium hypochlorite, full strength chlorine bleach
from the grocery store at room temperature for 60 minutes followed by an
autoclaving at 121 celsius for 30 minutes or at 134 celsius for 90
minutes. The autoclaving with sodium hydroxide is in the sodium
hydroxide. The autoclaving with bleach is after it is in water.

Following that, all these materials were cleaned in an ultrasonic
cleanser using a laboratory proprietary detergent with a pH of 9.45. All
of them got this, because, as pointed out by Dr. Rutala, it is important
to try and replicate the conditions under which these things would be
done in a hospital. The sonic cleaner was cranked up the highest
temperature that it would take, which was over 60 degrees, although,
somewhat variable, and for the longest time the timer kept, which was
for 90 minutes.

This was done by putting each object into a separate tube filled with
the cleaner, and then the tubes were immersed in more cleaner in the
chamber. Following that, there was a water rinse and then, finally, a
terminal sterilization in the autoclave at 121 celsius for 20 minutes in
order to model what we took to be standard hospital practice.

Next slide, please. As I mentioned, the sonicator was set at maximum
temperature and time. We only did the one set of conditions. We made no
effort to select a better cleaning solution. I am sure there are many
others, that many others are available.

Okay. Next slide, please. First, the assay technique for residual
infectivity on the glass slips. Each experiment, a positive control
consisting of 10 slips each holding .1 ml of dried on scrapie infected,
10 percent hamster brain dried down, not exposed to any decontamination
regimen, ground to a powder and a ml of PBS glass allowed to settle.
Fluids were then pooled, tenfold dilutions performed in
phosphate-buffered saline, each dilution assayed in four hamsters, the
same volumes that Dr. Rau showed you, .03 ml each intracerebrally into
the left frontal lobe. That is simply so that we would know in each test
how much infectivity had been used.

Next. Hang on a second. Let me finish. Each of 10 slips then was exposed
to some decontamination regimen, and then also ground to a powder in
phosphate-buffered saline, the fluid assayed as for the controls, so
that each experiment on glass involved 10 slips and 40 hamsters.

As David Taylor had mentioned to you earlier, we deducted incidental
deaths. We took 45 days as the cutoff between considering it a death
incidental. Perhaps we shouldn\'t have done that.

Next slide, please. For the steel needles, the positive controls were
tenfold, dilutions of infected hamster brain as a 10 percent paste in
normal calf brain, and then serial dilutions were done with normal calf
brain paste. Four needles were charged for each dilution, dried and then
a separate hamster was assayed for each needle. For the actual test, 40
needles charged with 10 percent hamster brain and normal calf brain,
dried, tested and then assayed as for the control above. Less incidental
deaths occurring before four days.

Next slide, please. This is just to show you what a titration on glass
looked like, the interim score here at eight and a half months. Notice
that the last positive animal so far as those inoculated with a dilution
of 10^-8^ calculated from the original brain tissue.

Next slide, please. And a similar titration for scrapie dried onto steel
needles, also positive to a reasonably high dilution. Actually, somewhat
higher than we got with the saline suspension, so we weren\'t sorry that
we had used the brain paste.

And you might notice that there is one negative at the lowest dilution.
That was an animal that died at 55 days. Brain was negative, and that is
why we\'re wondering whether 45 days might have been the best date to
estimate incidental deaths.

Next slide, please. Before I move on to the actual results, we were
interested to see what ultrasonic cleaning in hot alkaline detergent
alone without any other treatment would do, so we did a titration from
that and found a substantial reduction, both of the infectivity on glass
and on steel needles from the hot ultrasonic cleaning alone. The log
reduction factor is slightly over 5 logs. Although, for both models
there was some residual infectivity left on the surface.

Again, we made no effort to optimize, to modify or optimize the
procedure. We presume that most of the infectivity probably went into
the liquid, but we haven\'t made any attempt to find out whether that is
true.

Next slide, please. So here are the WHO studies. After exposure of glass
slips, there are the glass slips, to sodium hydroxide or to sodium
hypochlorite with autoclaving at either 121 celsius or at 134 celsius,
there was obviously a dramatic removal of infectivity, but darn, one of
the animals assaying material exposed to one normal sodium hydroxide at
134 autoclave, 134 celsius for 90 minutes has come down positive.

Next slide, please. And similar experiments with steel yielded
relatively similar results, at least two, maybe three of the assay
animals have had positive Western Blots. We are going to have to check
those out. Obviously, these stray positives have been seen before, and
we have to convince ourselves as to whether they are really positives or
whether it\'s inadequately digested PrP in the Western Blot or whether
it\'s real.

So the methods are, obviously, highly effective. They saved almost all
the hamsters and removed so much infectivity that most of the objects
assayed didn\'t show evidence of contamination. Remember that each of
these objects was charged with at least a million lethal doses of
scrapie agent, but we can\'t say that they are perfect.

Next slide, please. We have seen similar results in the past using
single chemical soaks. These are all done with the Chen glass test. And,
again, we have frequently seen, these are sodium hydroxide soaks at
various temperatures, an occasional stray positive.

Next slide. Some tests have found no positives at all, but remember with
the Chen glass test, we sample only about 12 percent of the supernatant
fluid from each slip, so that these results are not necessarily
different from the ones that show single positives. There is a
substantial sampling problem when you\'re dealing with very small
amounts of infectivity.

Next slide, please. Here is another result with sodium hypochlorite
where we had no positive animals. I marked these all as interim, because
we haven\'t finished all the Western Blots even though some of these are
not new experiments.

Next slide, please. And finally, I would like to say that some other
chemical agents are probably also very effective. Here are some results
using concentrated formic acid, which is used to treat tissues for
histology and immunohistochemistry. Note that there is only a single
positive animal out of 37 tested. Reports of a commercial phenolic
disinfection, at least temporarily unavailable here in the United States
is reported to be very effective, and we have heard that there are other
decontamination regimens in development not yet ready to share with the
FDA or the public that are showing promise.

Let me conclude now by, next slide, please, just summarizing that
methods developed to evaluate the effects of virucides are adaptable to
evaluate decontamination of TSE agents. Studies with two models both
suggested that exposure to 263K scrapie agent dried on surfaces to
solutions of sodium hydroxide, sodium hypochlorite with simultaneous or
sequential autoclaving and ultrasonic cleaning in hot alkaline detergent
markedly reduced amounts of infectivity, and the risk that any object
would retain detectable amounts of agent.

Other chemical treatments may also be effective, but uncertainties
remain. One, the reliability of the decontamination procedures, not only
the fact that we see stray positives, but also there is a theoretical
concern that the predictive value of these results, the results from
such models, may not adequately predict the behavior of decontamination
regimens in the actual health care or manufacturing setting, concern
that there may be sanctuaries of the kind that Bob Rohwer and David
Taylor have discussed that might occur in manufacturing processes or
health care setting that would impair the ability of otherwise effective
decontamination regimens to act.

It is quite late, but I am happy to answer questions for anybody who has
got the energy still to ask them. Thank you.

CHAIR PRIOLA: Are there any questions for Dr. Asher or Dr. Brown? All
right. If not, I \-- okay, Dr. Somerville has one for you.

DR. SOMERVILLE: I\'m just going to make a brief comment about the first
part of the talk, and that is that in our experience, in our survey, we
find that the various different grades of stainless steel are used from
the manufacturer of surgical instruments and with various different
finishings, and they have different responses to the kinds of treatment
that Stan Brown was trying on the instruments.

The one brief question I have is have you tried anything other than
visual inspection to see what the degree of damage is being done to the
instruments?

CHAIR PRIOLA: Can Dr. \-- oh, he is coming up there. Dr. Brown can
answer that.

DR. BROWN: The answer, at this point, is we have done nothing other than
visual, and part of the next generation of study is to be looking at
some of the different alloys, some of the different corrosion test
methods. Some of these effects are so blatant that why both to \-- I
mean, I cut up the gold handles and put them in the SEM just to make
sure it really was gold.

And, in fact, there was gold on those handles, but no, we haven\'t gone
any further. But one of the questions is are the different grades, you
know \-- in talking with the instrument manufacturers and the people who
do chemical analysis of instruments, there are a whole wide range of
grades of stainless steels, but the manufacturers will tell you what
probably is the most important is actually the mechanical treatment in
terms of how they make them, coworking, etcetera. And it may not be a
matter of chemistry, but it\'s a matter of mechanical parts.

The finger rings very typically are attacked by Clorox, and that is an
area where there has been a lot of mechanical cowork to form the rings.
Whereas, elsewhere on the same instrument, the surface looks fine. So
it\'s not just the chemistry, but it\'s actually the mechanical
processes used in the forming or fabricating. And again, if it\'s going
to go, it\'s going to go the first time you throw it in bleach.

BOARD MEMBER HOGAN: Dr. Brown, before you leave, I have one more
question. Dr. Brown, could you get rid of the black deposit that formed
on the sodium hydroxide instruments?

DR. BROWN: First of all, we didn\'t do any other cleaning. We just over
and over and over, autoclave and bleach. We didn\'t use what do they
call it, milk, the cleaning milk that is used in standard central
storage.

BOARD MEMBER HOGAN: So you didn\'t try? Is that it?

DR. BROWN: So what I did actually on some of them is I did a bit of
gentle scrubbing in the box joints to see if it would come off. It
wouldn\'t come off much by general scrubbing. Actually, if you reuse
them, you can begin to wear off the blackening. But it\'s really a very
superficial kind of blackening, and then the thing with the protein
adherence with the piano wire, they really did corrode and at least the
serum protein stuff we did didn\'t show any difference.

CHAIR PRIOLA: Dr. Edmiston?

DR. EDMISTON: I know it\'s late and I don\'t want to hold anybody up,
but I really want to commend Dr. Asher and Dr. Brown. You are heading in
the right direction. The question that I have is do you contemplate
looking at this in devices that have larger bores in terms of if you\'re
looking at a hollow device, are you looking at other devices that may
have a larger bore where the cleaning process may be expedited, normal
cleaning process may be expedited on the basis of having a larger
internal diameter?

DR. BROWN: These are solid pins.

DR. EDMISTON: These are solid pins?

DR. BROWN: These are solid pins. They were not needles.

DR. EDMISTON: Okay.

DR. BROWN: So the idea was that David had been using a needle. He was
used to the feel of that size needle, and I made solid pins to match. So
these were not hollow.

DR. EDMISTON: So you don\'t know what would happen with a hollow bore
device?

DR. BROWN: No. One can sort of guess, but I think \--

DR. EDMISTON: Right.

DR. BROWN: You know, this term of, you know, the nooks and crannies and
the hiding places, I think that\'s the next generation of the studies.
Polymer coated, we have got some that, apparently, are even teflon
coated that are part of the next step in the study.

DR. ASHER: Yes. As I mentioned, years ago I did some standard hypodermic
needles, just squirting suspensions of scrapie through and letting the
needles dry and autoclave. You know, you\'re not surprised to hear that
they were not sterilized.

DR. EDMISTON: I think our experiences have been that, especially in the
case of neurosurgery, that those patients who fall into that risk
category, a lot of us are moving towards the use of disposable biopsy
needles.

CHAIR PRIOLA: Dick, did you have a question? I\'m sorry, can you what?

BOARD MEMBER JOHNSON: Can we leave our papers behind?

CHAIR PRIOLA: I think you \--

SECRETARY FREAS: If you want it tomorrow morning, I would really
recommend you take it to your room. I do have a couple of quick
announcements. This morning, we passed out about 200 Conflict of
Interest questionnaires and we got about five of them back. I would like
to encourage you to look at the questionnaires and if you could drop
them off on your way out, we\'ll pass out another 100 tomorrow and,
hopefully, we\'ll got some back.

Also, somebody left behind a Palm Pilot. It looks like it\'s a very
expensive Palm Pilot, and if you can identify it, it\'s yours. Tomorrow
morning, we\'ll be seeing you at 8:00 sharp.

CHAIR PRIOLA: Okay. I would like to thank all of the speakers for
presenting published and unpublished data to the Committee, and we\'re
adjourned until 8:00 a.m. Thank you.

(Whereupon, at 6:18 p.m. the meeting was adjourned.)