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THE NATIONAL ANTIMICROBIAL
RESISTANCE MONITORING SYSTEM MEETING

June 23, 2005

I N D E X

Introduction and Purpose of Meeting
by Linda Youngman, Ph.D.

NARMS Background
by David G. White, Ph.D.   

Budget Issues
by Robert Walker, Ph.D. 

Retail Meat Arm
by David G. White, Ph.D. 

Animal Arm
by Paula Fedorka-Cray, Ph.D.

Human Arm
by Tom Chiller, M.D., Ph.D. 
by Tim Barrett, Ph.D. (Continued) 

Animal Isolate Sampling
by Neena Anandaraman, DVM 

Retail Meat Sampling
by Terry Proescholdt   

Retail Meat Reporting
by Elvira Hall-Robinson, D.
V.M., Ph.D. 

Molecular Characterization
By Shaohua Zhao, D.V.M., Ph.D.

KEYNOTE:  "---" Indicates inaudible in transcript.
  " * " Indicates phonetic spelling.

Introduction and Purpose of Meeting
by Dr. Linda Youngman

(9:10 a.m.)
  
DR. YOUNGMAN:  Good morning.  We are just about ready to start.  So, if I could ask that you take your seats, please.
  (Pause.)
  
DR. YOUNGMAN:  Good morning.  I apologize for the delay in getting started this morning, but everything is here now, except for the booklets that we will be providing. The booklets will have the slides that people are presenting today.  Those will be arriving shortly.  I apologize that they are not here yet.
  
Anyway, we are very grateful that you all are here, and I want to do an introduction.  My name is Linda Youngman, and on behalf of the Food and Drug Administration’s Center for Veterinary Medicine, I would like to welcome you to this  NARMS Meeting.
  
I know that some of you have traveled quite a distance to be here, and we really appreciate you taking the time out of your busy schedules to be with us today.  We look forward to your comments and advice about our NARMS program.
  
Rear Admiral Tollefson is not here today.  I am filling in for her.  She sends her regrets.  She was called away by the Secretary of Health and Human Services for a bioterrorism exercise.  We were looking forward to having her here, and we are a bit lost without her, but we will struggle through nevertheless.
  
We have asked each of you here today and tomorrow because you possess individual expertise that we believe will be critical and invaluable in helping us to prepare a review of our NARMS program.  Today representatives from FDA, USDA and CDC will provide you with factual background and information on the NARMS program, and then we will ask for your individual comments once you have heard some of the background about NARMS.
  
For many of you this will be a repeat.  You already know a lot of this.  For some of you it will be completely new.  But for everyone we are hoping to provide an update.
  
The representatives who will be speaking today can provide clarification to the questions that we are asking you to consider, but they cannot actually engage in any discussion about any particular aspect of the NARMS program.  This meeting is not an advisory committee meeting, and we are asking you for your individual thoughts, rather than a consensus opinion, and there is not going to be any time allotted for you to prepare a consensus opinion.  So, please.  If we could ask for your individual thoughts, we would be grateful.
  
The meeting will be transcribed so that we can use it in drafting an internal assessment of the NARMS program, and we will draft this report to give to the Science Board, which is an external advisory board that has responsibilities to the FDA.  They will advise us on the direction of our science, and the Science Board will also be doing a review of our NARMS program.
  
We will draft an internal assessment of the NARMS program using the transcription of today’s meeting as one  piece of information that we will be providing to the FDA’s Science Board.  We will wait for the Science Board review to be completed and then we are planning a follow up with either a public meeting or a publication of the Science Board’s review of the program, and possibly, also including pieces of this internal assessment.  A Federal Register notice of availability with requests for comments will be put in the public docket.
  
Now for some housekeeping details.  Most of you know Aleta Sindelar.  Is she here?  I guess she is trying to get the booklets for you.  If you need something, Aleta can help you.  I know there were some questions about getting some copies made while we are here today.  She will help you with those kinds of issues.
  
The bathrooms are down the hall.  So, if you take a left and then a right where the TV screens are, the mens and ladies rooms and telephones are back there.

You should each have an agenda now.  Again, we apologize for the delay in starting, but we are going to try to stick closely to the agenda and try to make up for lost time this morning.  So, we are very grateful that you are here.  Thank you for coming.  Especially those of you who came from a long distance.  We appreciate it; flying through thunderstorms.
  
So, without further ado, I would like to turn this over to the first speaker, Dr. David White, who is going to be giving you an introduction to our NARMS program.
  
DR. VOGEL:  Linda, can I ask a question?
  
DR. YOUNGMAN:  Sure.
  
DR. VOGEL:  You mentioned that the speakers can provide clarification, but cannot enter into discussion.  Can you expand on that?  What do you really mean?
  
DR. YOUNGMAN:  We are looking for individual opinions, not a consensus; from the representatives who are here to give us their individual advice today.  So we don’t want to push people toward a certain conclusion.  We want to hear from each of you individually.
  
DR. VOGEL:  But we are free to question the presenters?
  
DR. YOUNGMAN:  Yes.  Yes.  Oh, I’m sorry.  I also forgot one other thing.  That is, parking.  If you parked in the garage today, the normal cost is $8.50 a day.  If you punch in this code, it will be free, and I think it is the same code tomorrow, but I will let you know tomorrow for sure.  If you punch in #1470, you will be able to park for free.  It is covered by your attending this conference.
  
DR. McEWEN:  I have one question as well.  Are you expecting our opinions as we go?  Or are we to consider things and prepare written notes to submit after we convene?
  
DR. YOUNGMAN:  No.  We are asking for your comments today.  But if you look at the agenda, we are providing questions and asking for answers after several of the presenters have presented.  We scheduled in time for the meeting for questions and answers after the talks.
  
I am sure some questions are going to come up during the talks, however, but again, the reason for this is that we don’t want a consensus and we don’t want you to ask you to draft a consensus opinion.  We are asking for your individual advice.
  
All right.  I will turn it over to Dr. Dave White.  Thank you.

NARMS Background
by Dr. David G. White  

DR. WHITE:   Thanks, Linda.  I just want to take a  quick second to look at my slides, if I can.
  (Pause.)
  
DR. WHITE:  Okay.  Thank you very much.  I think probably too though, Scott, in terms of your question, there is a notebook that is being created right now that will have a list of certain aspects we would like to address throughout the talk.  Unfortunately, you don’t have it in front of you right now, but hopefully, most of these talks will kind of steer you in the direction of questions that will probably be forthcoming from all of you folks.  And hopefully, we can answer it as best as we can.
  
DR. McEWEN:  Is it an oral exchange of information?  We won’t be preparing written answers to the questions?
  
DR. WHITE:  I believe it is oral.
  
Thank you very much for coming.  I appreciate it as well, and I would like to take about maybe 20, 25 minutes to go over the history of this program and try to go back in time to gleam the information from numerous documents about how this program came to be, where it has been going, and hopefully, where we are headed.
  (Slide)
  
We keep seeing more and more articles on antimicrobial resistance.  Of course, we only have to go back to 1944, when Penicillin was hailed as a miracle drug and a secret weapon of the allied effort.
  
Of course, my former mentor’s second edition of his Book, the Antibiotic Paradox.  A good reason for those of you who are interested. 
  (Slide)
  
In terms of resistance, we all know, I think, that it is a complex phenomenon.  The increasing incidence of resistance has serious implications for the future treatment and prevention of infectious diseases in both animals and humans.  Although much scientific information is available on this subject, many ecological aspects of the development and dissemination of antimicrobial resistance remain uncertain.
  (Slide)
  
What is known, however, is that the emergence and dissemination of bacterial antimicrobial resistance is the result of numerous complex interactions among antimicrobials, microorganisms and the surrounding environments.  A clearer understanding of these interactions is necessary if a science-based assessment of the relative risks concerning the use of antimicrobials in the animal husbandry environment is to be made.
  (Slide)
  
In terms of our strategy for CVM, it is aimed at assessing relationships between antimicrobial use in  agriculture and potential human health consequences.  We have adopted a multi-pronged approach that includes a revised safety assessment process.  For those of you not familiar with Guidance 152, it is a new process by which antimicrobial drugs are looked at.

We have increased our education outreach activities.  In cooperation with AVMA we published a series of prudent use guidelines on antimicrobial use in different food production environments.
  
Participation in international activities, like Codex and OIE.  Expanded research activities and enhanced surveillance activities.
  (Slide)
  
And, of course, the one that is relevant to us today is NARMS, in terms of surveillance activities.  This is what I tried to gleam from the history of NARMS.  It actually goes back to 1994, 11 years ago, when FDA held a joint Veterinary Medicine and an Anti-Infective Drugs Advisory Committee to address the specific issue of approval of fluoroquinolones for use in poultry.
  
The committee was united in advising the agency that if the products were to be approved, several restrictions should be placed on the use of these drugs in order to attempt to minimize the public health risks related to the development of resistant bacteria in animals.
  
The most relevant recommendation to us today was the establishment of a nationally representative surveillance system to monitor resistance trends among both human and animal enteric bacteria.  So, that was one of the recommendations from 11 years ago.
  (Slide)
  
So, from that the National Antimicrobial Resistance Monitoring System, Enteric Bacteria or, as we affectionately call it, NARMS, became operational in 1996.  Salmonella of animal and human origin was the initial organism accepted for surveillance, and NARMS combines the activities of FDA, CDC and USDA to create a nationwide monitoring system.
  
It was a tripartite design, development and implementation, and the three founding fathers, so to speak, were Dr. Linda Tollefson, from FDA, Dr. Fred Angulo, from CDC and Dr. Paula Fedorka-Cray, from USDA.
  
The sources and types of isolates have expanded over the time of the program.  Campylobacter was added in 1998 and Enterococcus and E. coli in 2000, and we added the retail arm in 2002.
  
Each year samples are collected from numerous origins and tested to determine if there have been changes in the susceptibility/resistance of certain enteric bacteria to selected antimicrobial drugs, and the antimicrobial drugs tested annually are selected based on their importance in both human and veterinary medicine.
  (Slide)
  
What NARMS allows the FDA to do is to monitor changes in susceptibility/resistance to drugs used in humans and food animals.  Very simply, the human samples are collected from ill people.  The animal samples are gathered from healthy farm animals, animal clinical specimens, carcasses of food animals at slaughter and processing plants and ground products at processing plants.  The retail meat samples are collected from grocery stores and shops that sell meat and poultry to the public.
  (Slide)
  
In terms of the goals of the NARMS program, it is to provide descriptive data and trends on antimicrobial susceptibility/resistance patterns in zoonotic foodborne pathogens and select commensal organisms.  It started out initially with the pathogens, but we have since added Enterococcus and E. coli as commensal organisms.
  
Also, the goal is to respond to unusual or high levels of bacterial drug resistance in humans, animals and retail meats in order to contain or mitigate resistance dissemination.  Also, to design follow-up epidemiology and research studies to better understand the emergence and transfer of antimicrobial resistance and assist the FDA in decision making on approving safe and effective drugs for humans and animals, as well as promote prudent and judicious use of antimicrobial drugs.
  (Slide)
  
There are three testing sites.  The human isolates are done at CDC, at the National Center for infectious Diseases, Foodborne and Diarrheal Diseases Laboratory in Atlanta, Georgia.  The animal isolates are tested at USDA, ARS in Athens, Georgia in the Bacterial Epidemiology and Antimicrobial Resistance Research Unit.  The retail meat isolates are tested at the Center for Veterinary Medicine Office of Research in Laurel, Maryland.
  (Slide)
  
From 1996 to the present, NARMS has been funded through FDA via IAGs, Interagency Agreements with both USDA and CDC, and we also have interagency agreements with APHIS and FSIS, I believe, through ARS.
  (Slide)
  
The animal, human, retail lab testing are comparable in terms of the methodologies used.  We have had methods meetings in 2002 and 2003 to standardize the methods between the three arms.  In the past we have had an annual NARMS meeting to present the data, but we did not have one this year due to budget limitations.  We hope to have one next year.  Hopefully, teamed up, possibly, with ICEID.
  
In terms of the susceptibility testing methods, they are all the same at all three arms.  We use the Sensititre System from Trek Diagnostics.  It is a broth microdilution panel for Salmonella, E. coli, Enterococcus, and hopefully, as of next week, we will be using the Campylobacter brother microdilution panel, which is going for approval this weekend at CLSI.
  
So, as of this year we will have all four bugs under surveillance being performed with broth microdilution.  In the past Campylobacter has been done by E-test and agar dilution.
  
We use the same antimicrobial plate formats.  We meet generally a couple of times a year to go over what antimicrobials are on the panels.  We try to come to some consensus about what should be on there.
  
In terms of the manufactured lots, Trek manufactures their susceptibility plates in certain lots.  We distributed them between the three sites so that each site has the same lots, and we try to use the same isolate handling procedures.
  
Quality assurance.  We have both internal and external programs to monitor quality assurance of susceptibility testing.  We follow CLSI/NCCLS standards where appropriate.  We do have several antimicrobials which do not have CLSI quality control or interpretive criteria, and we do note that in our reports.
  
In 2003 we adopted the WHO External Quality Assurance System for susceptibility testing and for Salmonella serology testing I believe.
  (Slide)
  
In terms of the bacteria tested, we do have some similarities among the sites, but each site does have its own unique isolates under surveillance.  CDC has Non-Typhi Salmonella, E. coli 0157:H7, Enterococcus, Campylobacter, Salmonella Typhi, Shigella, Listeria and Vibrio.
  
USDA has Non-Typhi Salmonella, E. coli 0157:H7 when available, Enterococcus and Campylobacter, and the retail meats do Non-Typhi Salmonella, E. coli, Enterococcus and Campylobacter.
  (Slide)
  
As I mentioned before with regards to the methods.  The major focus of this program is susceptibility testing, and we have designed this program so that comparable testing methods are used between all three arms.  We use the broth microdilution methods using the Sensititre
Semi-Automated system, and this is a picture here.
  
We have all started using the new ARIS systems, which have been a little problematic at the beginning.  We have started to use them more, and they have a new software that will allow us to, hopefully, dump the data right into an access program for analysis.
  
As I mentioned, the participating NARMS laboratories adhere to the appropriate published CLS/NCCLS performance standards.  Both the veterinary and the human.  The M100-S15 and the M31-A2 I believe.  Or, the M31-S1, which is coming out.
  (Slide)
  
So, in terms of the methods for Salmonella and E. coli, it is tested by broth microdilution, Sensititre.  The plates have changed over time for the past several years, but there are some core antimicrobials on the plates that have not changed over the years of the surveillance program. 
  
So, the core antimicrobials that have been tested over the length of the program include:  amikacin, amoxicillin-clavulanic acid, ampicillin, ceftiofur,  ceftriaxone, chloramphenicol, ciprofloxacin, gentamicin,  kanamycin, naldixic acid, streptomycin, tetracycline and trimethoprim-sulfamethoxazole.
  
Now, we did have sulfamethoxazole from 1987 to 2003.  We replaced it with sulfisoxazole in 2004, as the QC is only for sulfisoxazole with CLSI/NCCLS, not sulfamethoxazole. 
  
Ticarcillin was on one of the first plates but is geared as an anti pseudomonal.  Since we didn’t test Pseudomonas, we moved it off the plate.  We had apramycin on there for several years until the manufacture was discontinued by Elanco.  Apramycin is no longer being manufactured.  So, we removed it from our plate.
  
Florfenicol, we put on there as a snapshot to see if there was any changes of susceptibility of florfenicol, and we felt that chloramphenicol was suitable as a sentinel for both drugs, because most of the isolates that are Florfenicol resistant -- actually, I think almost every isolate that is florfenicol resistant is chloramphenicol resistant as well.
 
Imipenim we put on there again as a snapshot, and I don’t think we saw any resistance at all.  So, we took it off the plate.
  
Cephalothin we had on there for six years, but we didn’t feel it was giving us that much information.  So we removed it as well.  And we added cefoxitin in its place in 2000 to pick up primarily the blaCMY cephalosporinase that we are starting to see in our Salmonella and E. coli isolates.
  
So, over time the dilutions have improved to include full ranges in the past.  Some of our ranges do not include the appropriate quality control or the break points.  All of the dilutions on our plates today have both complete quality control ranges in there, as well as the interpretive criteria for SIR.
  
And we use the appropriate QC organisms for testing.  In the case of our Salmonella and E. coli, we used E. coli, S. Aureus, E. Faecalis and Pseudomonas Aeruginosa, depending on the drug and the range.
  
Enterococcus.  Again, it is much like Salmonella and E. coli.  It is tested by broth microdilution.  We do have several core antimicrobials that have remained the same over the plate, and these include bacitracin, chloramphenicol, ciprofloxacin, erythromycin, flavomycin, gentamicin, kanamycin, lincomycin, linezolid, nitrofurantoin, penicillin, salinomycin, streptomycin, Q/D, tetracycline, tylosin, vancomycin. 
  
A lot of these drugs you may not be familiar with because they are considered animal production drugs.  Salinomycin is a coccidiostat*, and we had that on the plate between 2000 and 2003.  We removed it from the plate in 2004, and we added Daptomycin once it was approved by FDA.  We felt that it was important to put it on our surveillance system.
  
Again, the dilutions have improved over time, much like the E. coli and Salmonella plates.  We have full ranges now for all of these antimicrobials, including the appropriate QC ranges, as well as the interpretative criteria.
  (Slide)
  
So, in terms of the ‘05 Salmonella/E. coli panel, this is probably our best panel we have put together, and like I said, we have worked hard to make sure that everything is quality controlled and we have the appropriate S/I/R in there.  We have several beta lactams under surveillance:  ampicillin, amoxicillin/clavulanic acid, cefoxitin, ceftiofur and ceftriaxone. 
  
We have nalidixic acid and cipofloxacin.  In terms of folic acid inhibitors we have sulfisoxazole and trimethoprim/sulfamethoxazole.  In aminoglycosides; we have streptomycin, kanamycin, gentamicin and amikacin.  There is no QC for streptomycin by CLSI/NCCLS standards.  And then we have chloramphenicol and tetracycline on the plate.  And the plate designation by Trek is CMV1AGNF.
  (Slide)
  
This is all going to be in your packet when you get it.  So, you don’t need to remember these numbers.  This is just to give you the transparency of th
e ranges that we employ in the testing and the break points used.
  As I mentioned in streptomycin we don’t have QC.  We don’t have interpretative criteria.  We do use a break point of 64 µg/mL.
  (Slide)
  
Enterococcus.  Just to give you a breakdown, we do have several production drugs, as I mentioned, that are used in veterinary medicine:  Bacitracin, flavomycin, lincomycin and tylosin.  We do have three aminoglycosides, and these are to measure high level aminoglycoside resistance.  These are all at MICs 256 or higher to strep, kan and gentamicin.
  
We have vancomycin on there, as well as the two newer drugs that have been approved for gram positive  infections in human medicine, daptomycin and linezolid.  We also have penicillin, Q/D, ciprofloxacin, nitrofurantoin, erythromycin, chloramphenicol and tetracycline for the Enterococcus plate.
  (Slide)
  
And again, this will be in your packet when you get it to give you an idea of the ranges used and the break points employed.  Again, the red asterisks show that we do not have QC for those drugs, as they are production drugs.  CLSI doesn’t do quality control for production drugs, but we hope to change that over time.
  (Slide)
  
Campylobacter surveillance was added in 1998 at USDA and CDC, and from 1998 to 2004 the E-test was employed for antimicrobials, including azithromycin, clindamycin, erythromycin, nalidixic acid, ciprofloxacin, gentamicin,  chloramphenicol and tetracycline.
  
When the NARMS retail arm began in 2002, it tested these isolates with E-test, as well as the agar dilution CLSI approved methodologies to ciprofloxacin, doxycycline, erythromycin, gentamicin and meropenem.
  (Slide)
  
Different methods though between the arms and the lack of reproducibility for some drugs led to the development of a new CLSI/NCCLS approved broth microdilution method for Campy in 2004 using C. jejuni ATCC 33560 as the quality control.
  
The QC MIC limits of the AST of Campylobacter have been accepted by the CLSI in January of ‘05, and will be published in the upcoming CLSI documents.  As I mentioned, the erythromycin was a little bit out of range and is being presented, I believe, this weekend at the VAST and AST meetings.
  
This broth method will be much easier to use than both agar dilution and E-tests and is amenable to semi-automation and has QC ranges for 14 different antimicrobial agents for Campylobacter.  So, we are very excited about this and really can’t thank enough the group at OR that was spearheaded by Dr. Pat McDermott, Dr. Bob Walker and Sonya Bodeis for helping to champion this system through.
  
So, as soon as we are ready to go with Campylobacter this year; we will use the broth microdilution method.  So, we are ready to jump in there and see what we get.
  (Slide)
  
These are the antimicrobials that are going to be on the new panel, and we have several macrolides and lincosamides:  azithormycin, erythromycin and clindamycin.  We have nalidixic acid and ciprofloxacin.  We added a new Ketolide, telithromycin, as well, and we have others on there:  florfenicol, gentamicin and tetracycline.  So, we will have nine antimicrobials on the Campy plate.
  (Slide)
  
These are the ranges that are going to be employed, and in terms of the breakpoints, this is all kind of based on populations since there is no interpretative criteria for Campylobacter for CLSI or NCCLS.  We have been using these types of breakpoints, and you will notice we don’t have
breakpoints for telithromycin or florfenicol at this point.
  (Slide)
  
Other methods used by NARMS. We do have our Salmonella and Campylobacter isolates characterized by Pulsed Field Gel Electrophoresis, and to give you some background, Dr. Zhao will talk about this later on.
  
Genomic DNA is cut with an enzyme and resulting fragments of different sizes can indicate mutations or insertions in the genomes.  Whereas PulseNet is an international program -- well, it is national and international now.  It assists investigations and improves outbreak detection through rapid linking of cases by comparing bacterial DNA fingerprinting patterns.  But a lot of our isolates are indeed now put into PulseNet.
  (Slide)
  
Other capabilities, of course, were warranted.  We all have molecular capabilities.  All three arms have molecular capabilities to investigate emerging phenotypes and go after the appropriate genotypes.  Here are just some examples of some PCRs that we have done looking at resistance genes for high level aminoglycoside resistance and then also we have several programs involved in looking at integrons among bacteria.  So, all three arms are able to do this.
  (Slide)
  
So again, just kind of summarizing everything up, this is the way NARMS is designed right now.  There are three arms, the USDA, CDC and CVM sides.
  (Slide)
  
We all have annual reports.  They are all linked on the CVM home page, but USDA and CDC have their own home pages as well for NARMS, and you can access all three of them on our web link up there.  These are just covers of the annual reports for the different sites.
  (Slide)
  
So, in conclusion, we need to remember that NARMS is a public health surveillance system.  It will enable  informed decision making by FDA, initiated response activities if data warrants.  We hope it promotes prudent and judicious use of antimicrobials and will help guide prescription practices.
  
We hope it encourages standardization of laboratory techniques, identify areas for more detailed investigation, promote collaboration between the three different government agencies involved, as well as academia.  And ultimately, prolong the efficacy and useful life of antimicrobials.
  
With that, I thank you very much.  Are we going to take questions now? 
 
DR. YOUNGMAN:  We can take questions.
  
DR. VOGEL:  David, you mentioned that Campylobacter has gone to the broth microdilution.  Two questions regarding that.  Number one, how is the results of that comparable to the E-test?  Is that going to affect your trend analysis or not?
  And secondly, how does that affect the ability for your workload, your sample throughput?  Will that increase your ability to do additional sampling for Campylobacter?
  
DR. WHITE:  Thanks, Lyle.  Those are good questions.  I will answer the second one first.  The broth dilution method is much easier than the E-test and agar dilution methods.  So, if anything, it will shorten the time it takes to do susceptibility testing on those organisms.
  
Allowing us to do more is all dependent on the budget and everything else.  You know, what we can collect for isolates.  The numbers we get now through retail meats -- and I am just speaking for myself; I can’t talk for Tom or Paula.  We get about now 600 to 800, and it takes a good six to eight months to do that by agar dilution.
  
This should probably drop us down to maybe two to three months once we are up and running.  So, we are really excited about that. 
  
The first question, in terms of the comparison to E-test, E-test is not a CLSI endorsed method.  So, if anything, you need to take the E-test data and compare it to the broth, not the other way around.
  
We have done some studies.  We have done both in the past comparing agar dilution and E-test at the same time, and that is probably because of our research laboratory and Dr. Walker’s expertise.  So, we have that data to show comparability for some drugs, but that is why we went to broth, because the E-test was not giving reproducible results for several drugs, whereas the broth microdilution method is a reproducible standardized method.
  
D
oes that help?  Okay.  Dan?
  
DR. SAHM:  Thanks for the overview, David.  The question that I would like to ask has to do with the determination -- the two key resistance mechanisms to extended cephalosporins that are being seen in enterics are extended spectrum, beta lactams and AMP C mutations or AMP C acquisition.
  
The selection of cephalosporins you have on your gram negative panel wouldn’t be the most sensitive set of cephalosporins to help detect ESBLs emerging, and I was wondering how that data is looked at, given the cephalosporins that you use as a sentinel for potential ESBL emergence in your study organisms.
  
DR. WHITE:  Thanks, Dan.  That is a great question.  Actually, we have -- that is a very good point.  A lot of ours are just kind of screening.  They may be adequate for 10 beta lactams and so forth, but the ESBLs -- we have actually created secondary plates.  I didn’t have a chance to talk about it.

We actually have a secondary ESBL plate that has on there cefotaxime*, cefotaxime/clavulanic acid, ceftazidime, ceftazidime/clavulanic acid, cefepime*  We have also added Cefquinome*, which is a fourth generation veterinary cephalosporin not approved in the United States. So, we further test anything that has a red flag for Cefoxitin.  We kind of take off and put on this ESBL plate and we look for the definite, the three-fold reduction, and then further on we try to characterize from there.
  
We actually have also a supplementary fluoroquinolone panel as well.  So, anything that is nalidixic acid resistant, we then put onto this plate that has on there marbofloxacin*, danofloxacin, enrofloxacin, levofloxacin, ciprofloxacin and so forth.
  
So, we try to -- you know, these are the second tier plates.  So once we get a red flag on a certain phenotype we go to that.  I apologize for not showing that.  I will try to get that for everybody for tomorrow.
  
DR. SAHM:  Just as a follow onto that, for instance, in the human isolates one of the more sensitive cephalosporins, and the flag for that is any MIC greater than or equal to one or two I believe.  I can’t remember.
  
DR. WHITE:  Right.
  
DR. SAHM:  Because of a large number of TEMS that won’t show up with this as well.  So, you might want to think about that flagging system.
  
DR. WHITE:  Okay.  Thank you.  Good point.
  
DR. ALTEKRUSE:  Is that being done in all three arms of NARSM?
  
DR. WHITE:  Those plates are available to all three arms.  I think right now CVM has probably done the most of that with that plate, but we had help with CDC.  So, all three arms use the plates.
  
We don’t see much though for ESBLs.  At least with the animal isolates and the retail isolates.  I think we have seen one ESBL E. coli that fits the classic definition.
  
DR. KOTARSKI:  I have two points to make.  The first point is not necessarily a question, and maybe we will come back to this later.  The objectives for the program that you have in your first slides don’t quite match up with what we received in our packages, and they are not necessarily the same as what is on the individual websites.
  
And so, as we go forward in the program, I would like to get some confirmation as to specifically what the objectives are.  That might be helpful for the panel in addressing the questions that we have outlined for the afternoon.  So, just a comment.
  
The second is a question.  Quickly, what do you use to establish -- what criteria do you use to establish breakpoints for Campylobacter as you work through?
  
DR. WHITE:  That is a great question.  I will let Dr. Walker answer that question.
  
DR. WALKER:  CLSI has a working group called, for lack of a better term, Orphan Organism, and they have been meeting -- we have been meeting for the last two years.  And one of the things that we are doing in that group is to take infrequently isolated organisms and try to establish testing methods and QC and interpretative criteria with Campylobacter, because we have already gone through CLSI to establish the testing methods and the QC ranges.
  
What we are going to do, like Dave said, at this meeting come up this weekend and this next week, is establish interpretative criteria for Campylobacter for some drugs.  So, CLSI is addressing this.  They are doing it off of population distributions for the most part, and I would suspect that come Tuesday afternoon there should be some recommendations as to interpretative criteria for some drugs for Campylobacter.
  
DR. KOTARSKI:  So then basically, it is a screening mechanism for increased MICs relative to the general population, what you would expect?  It is not based on therapeutic failures or anything?
  
DR. WALKER:  Yes.  And for those who are familiar with CLSI, you know that to -- generally what they do to generate interpretive criteria is to look at population distributions, look at pharmacokinetics, pharmacodynamics and look at the results of clinical trials.
  
And then those interpretive criteria go into books that are referred to as standards books.  For this particular bunch of organisms they are going to look at population distributions, PK/PD where they have it, but obviously the clinical trial is not going to be available.
  
So, this document will be called the M45, and it will be a guidance document.  It will not be a standards document.
  
DR. WHITE:  Great.  Thank you very much.
  
DR. MILLER:  One last question.  You mentioned C. jejuni for the microdilution new techniques.  Are you also moving in that direction for C. coli?
  
DR. WHITE:  It is for both.  All Campylobacter.
  
DR. ANGULO:  Dave, just one clarification.  CDC started Campylobacter in 1997, and thanks for highlighting the important innovation for having the broth microdilution for Campylobacter because that is really essential for us.
  
You will hear more, but we have changed the number of samples we are receiving.  And that is because we are able to test more samples because we are more efficient at testing what manner.  So, thanks for highlighting that.
  
DR. ALTEKRUSE:  You mentioned Listeria being done I think at CDC.  Is there any evidence from that that perhaps other arms should also be looking at Listeria?
  
DR. WHITE:  No.  I think every once in a while we take a snapshot of something and then we evaluate within our groups and we see.  It is kind of like imipenem.  We put imipenem on there for a year.  We didn’t see any elevated MICs.  I don’t think any of us did.  And so we said, okay, let’s yank it off.
  
Remember, we have 96 wells.  We are limited to 96 wells.  Three of those are control wells.  So, we are stuck with 93.  So any time we tweak something, we have to take something off. 
  
That is why we got rid of Cephalothin*, primarily because we didn’t think it was giving us information, and with those several wells that we had we were able to improve the QC ranges and cover all the ranges for all of the other drugs on the plate.
  
DR. ALTEKRUSE:  Okay.  Thanks.
  
DR. YOUNGMAN:  Before I introduce the next speaker, I just wanted to point out that while our panelists have microphones at their desks, for the rest of you there is a microphone which is live all the time that is here in the center of the room.  Please use it.  That helps our transcriber in making sure we accurately reflect your questions and comments.  Thank you.
  
If I could, I will introduce Dr. Robert Walker, who is going to be talking about the NARMS budget.

Budget Issues
by Dr. Robert Walker
  

DR. WALKER:  Good morning.  As Linda indicated,

Dr. Tollefson has been called away for other activities, and she was originally scheduled to discuss the budget.  So, what you are going to hear today is the budget from my perspective, which is kind of like looking at the universe through a pinhole.
  (Slide)
  
With that said, what I would like to do is to talk to you not so much about the specifics of the budget, but more of the mechanisms of how the NARMS budget is distributed.  When the money comes into CVM, CVM generates IAGs, Interagency Agreements, with the two participating laboratories, CDC and USDA.
  
The monies that are allocated in these IAGs are for personnel, supplies, equipment and overhead.  When we initiate get the IAGs, they may be broken down to a specific amount for personnel, a specific amount for overhead, et cetera.  But once the IAGs are in place, the agencies, the participating laboratories, can redistribute that money however they need to, as long as the monies are used to generate NARMS related work.
  
In other words, if we gave $800,000 for personnel and they found that they had an increased need for personnel, they could take monies from equipment or some other source and add it to the personnel part of the budget.  The only requirement is that however the money is spent, it is spent on NARMS-related activities.
  
Additional dollars that may go into the NARMS monies the labs receive is through supplies used for susceptibility testing, and Dave talked about this earlier.  What we have found is, in order to increase the uniformity in the susceptibility testing work that we do, if we order a large batch of plates, we get a cost break because of the large volume that we order.
  
We then take that volume of plates and divide it amongst the three labs so that everybody is using the same lot numbers.  We may have to do that several times throughout the year, but each time the lots are distributed throughout the three labs based on their needs.
  (Slide)
  
So, just to give you an idea of how the mechanics of this works, if we look at the USDA budget, in the IAG they had about $800,000 for personnel.  This person was a CVM person who was working at USDA to provide whatever assistance she could.  This represents the central supplies, the cost of plates and other supplies relating to testing those plates.  That comes out of CVM’s budget.
  
And then again, the different breakdown into the different types of testing that they are doing and the money that was allocated for that, for a total of a little over $2 million.  And again, the specifics on Campy testing or Salmonella PFGE, that is all controlled internally by Paula and her group.
  (Slide)
  
If we look at the CDC, it is kind of the same type of system where we have personnel monies, the CVM staff at CDC, central supplies.  CDC is a little bit different than USDA in that as part of the retail meat program, in order for us to gain access to the isolates from the retail meats, CDC has contracted with 10 FoodNet sites to go out into the community -- Terry  Proescholdt will talk about this later on -- and purchase meats, bring them back to their individual laboratories where they will isolate and identify the specific organisms.  These isolates are then sent to CVM where we confirm the identification their identification.  Once the identity is confirmed the isolates are subjected to susceptibility testing and whatever genetic analysis needs to be done.
  
CVM is not in a position to contract with these FoodNet sites or these state public health labs to go out and make these purchases.  So, CDC has contracted with them.  And the money -- approximately $500,000 goes from CVM into CDC for CDC to distribute it to the state public health labs that are participating in the retail meat study.
  (Slide)
  
For CVM, again it is broken down basically the same way.  And for those of you who are doing the math  you will realize that this number down here is not correct  This really should be $1,190,885.  You might want to make a note on that for your handouts when you get them.
  (Slide)
  
So , when you look at the expendable monies, and what we are talking about is the monies that are expendable from NARMS for the research or the surveillance program, this is how it breaks down for the animal arm, the human arm and the retail meat arm of NARMS.  If you look at this, plus this, this would be what CDC gets.  But keep in mind that not all of this money actually goes to CDC.  It passes through CDC to the state labs.
  
This is the amount, $1,190,000 is what goes into the retail meat arm.. An additional $100,000 of NARMS monies  goes to the WHO Global Salmonella Surveillance System.  This value then comes out to be 5,446,000 for those of you who like to keep track of numbers.
  
Keep in mind that this is the mechanisms by which the NARMS money comes into CVM and then is distributed to the other arms.  The specifics, in terms of the exact numbers, I think at this point in time this is not the issue.  The issue is the mechanism by which the money is distributed and the amount of expendable money that is available.  Are there any questions?
  
DR. ANGULO:  In the opening slide you mentioned discretion of the agencies to move money, but we don’t -- we have to get permission.  Well, CVM gives us permission to move money, but our finance office won’t allow us to move money, because we follow the interagency agreement as is given to us.  So, we have to keep the money in the categories where it is coded to us when it comes to us.  Unless we get an email or some written documentation from CVM for permission to move the money between. 
  
We can’t move supply money to personnel money, et cetera.  So, we have flexibility, but we have to get it in writing from CVM to make the changes and keep records of those changes.
  
DR. WALKER:  Yes.  That is an internal CDC thing, but I think you have found that if you communicate with us and ask for permission, it is given without any problem.
  
DR. ANGULO:  That is right.  We have always had permission to move, but we document the movement.
  
DR. WALKER:  Okay.  Yes, Paula.
  
DR. FEDORKA-CRAY:  Bob, I think it is important to point out that those figures --
  
MS. SINDELAR:  Can I make a comment here?  For the purpose of transcription, for her ease, that was Dr. Fred Angulo, from CDC.  And, Paula Cray.  So, please identify yourself prior to your comments.  Thank you.
  
DR. FEDORKA-CRAY:  This is Dr. Paula Cray, from the U.S. Department of Agriculture.  For the purpose of the interagency agreements, the interagency agreements do not reflect the numbers that have been shown on the slides, nor have these numbers been presented to at least the animal arm of NARMS in this particular fashion at any time during the past 11 years.
  
The inclusion of personnel and -- from CVM, Marcia Headrick was the NARMS coordinator.  Our understanding is that she was not a directly -- directly placed to work exclusively with USDA.  She coordinated, until very recently, all of the activities between CDC, FDA and USDA, and I think it would be inappropriate to count her salary toward a USDA figure.
  
In addition, the monies that have been expended on supplies, it was discussed early on in the program what the needs were with regard to the number of isolates that would be available, and it was always recognized that because of the -- particularly the slaughter and the diagnostic sources of isolates, that there would be an increased need for additional plates.
  
However, what the agency does not see is that in any given year we, between the three agencies, routinely trade or pass plates.  We provided CDC with over 1,000 plates at one time for Salmonella.  They provided us with 600 for Enterococci.
  
These numbers have gone back and forth over the year, in addition to supplies, and the supplies that FDA provides to us are strictly limited to Trek Diagnostic supplies, not other expendables that must go into the system, like gloves and tubes and gases and other media that are going to be required for the program.  So, I think that there needs to be some additional detail that is reflected in order to accurately assess the amount of monies that each arm receives.
  
DR. WALKER:  And I take what you say well.  What I wanted to do is talk more about the mechanics.  I am not involved in the budget.  I am not in a position to go into the exact specifics on the budget.
  
You would notice that there was $67,500 listed for USDA for other supplies, and I recognize that the central supplies were Trek related supplies monies.
  
DR. YOUNGMAN:  If I can just clarify a couple of things that you said though, Paula.  The monies that were presented here, the budget figures you saw here, were for 2005 only.  That is why you haven’t seen those exact numbers before.  We are just talking about this year’s money.
  
We can never be certain of our exact budget because our FDA budget changes from year to year, and we have to work with the budget constraints we are given. 
  
And also, to clarify about the CVM staff, the dollar figure that was put up there for a CVM staffer at USDA, we also supply a CVM staffer at CDC, and those monies do not come out of your IAG.  Those monies are paid for separately out of FDA monies toward NARMS.
  
DR. FEDORKA-CRAY:  This is Paula Cray again.  I think that then that there needs to be a clarification made on those tables if the experts are looking at those figures and they don’t have an accurate assessment.
  
The interagency agreement that USDA received was $1.5 and 6/7 million for 2005, not $2 million that is reflected in the figures that were presented to you.
  
DR. WALKER:  Right.  And most of those figures that are on that page came directly from the IAG.  The ones that would be the exception would be the $140,000 for Marcia and then the central supply monies would be different.
  
DR. MILLER:  Marissa Miller.  Bob, is there a special budgetary line item or appropriation for NARMS for FDA?  And if not, how does CVM protect that money to make sure?  I mean, a $5 million total is quite large part of the overall CVM budget I would think.
  
DR. WALKER:  Well, I wish Linda was here to give you more information on that.  I am not in the budget channel.  My area is in the science.  I’m not sure if this is a direct line.
  
DR. YOUNGMAN:  I apologize that Linda Tollefson is not here.  She might be able to answer your question.  In actual fact, she is really the one who could answer questions about the NARMS budget.  So, Bob and I are trying to cover as best we can for her today.
  
DR. WALKER:  Based on the things that I hear it may be a line item or at least it is specifically discussed by Congress.  But I don’t know the politics of how it channels down through the budgeting process.
  
DR. MILLER:  Do you happen to know?  Have you ever had a GAO audit and what the results were from that?
  
DR. WALKER:  Not as far as I know.  We have not had a GAO audit.
  
DR. McEWEN:  Scott McEwen.  Are there other sort of in-kind contributions that help to sustain and support NARMS that don’t appear in the budget?
  
DR. WALKER:  Are there other sources of funding that can be fed into it?
  
DR. McEWEN:  Well, funding or in-kind contributions from the different agencies.  Salaries of people I guess that don’t show up in the budget that participate in NARMS or other stakeholder groups that helps furnish samples.  That kind of thing.  I am just trying to get a feel for other activities that support NARMS that may not appear in the financial statement.
  
DR. WALKER:  I don’t know what CDC and USDA does.  I am sure that they have peripheral activities that all contribute to it.  I know at CVM we have a number of people that work  directly, and indirectly, with the NARMS program doing things like Dave talked about, the Pulsed-Field Gel Electrophoresis that are not figured into the NARMS budget.
  
In other words, we don’t have anybody at OR that is directly involved with NARMS who is doing Pulsed-Field Gel Electrophoresis, but we do have an OR team that is involved with Pulsed-Field Gel Electrophoresis that deal with a lot of NARMS isolates.
  
DR. ALTEKRUSE:  On that point, one example of -- and Dr. Kotarski brought this up.  One example of an in-kind contribution would be that both APHIS and FSIS isolate Salmonella and provide them to the USDA/ARS laboratory for the characterization.  So, that is a fairly substantial
in-kind contribution.
  
DR. WALKER:  Yes, Fred.
  
DR. ANGULO:  We all have in-kind additional support.  At CDC it is on the scale of -- about matching is the word that we get from the interagency agreement.  So, of all our NARMS activities, which include everything from education to core surveillance to applied epidemiology studies, et cetera, that are the total resources that are contributed to animal resistance testing and susceptibility testing, the interagency agreement provides about half of all of our resources.
  
DR. WALKER:  Okay.  Yes, Lyle.
  
DR. VOGEL:  Mostly just a comment.  I wish Linda Tollefson or Steve Sundlof were here to address the budget issues and questions, because one of our discussion questions was how do we suggest that we enhance or sustain funding for NARMS. 
  
I think it would be important for us to know that if, for example, we went to Congress and was able to either create a line item for NARMS, if that would be acceptable to the agency.  If not a line item, if we got increased funding specified for NARMS, would FDA use it for NARMS or would they divert to something else?
  
DR. WALKER:  I think those are good points.
  
DR. FEDORKA-CRAY:  This is Paula Cray again.  I just want to go on record, like Dr. Angulo, that the Agriculture Research Service and USDA, including, like Sean Altekruse said, APHIS and FSIS, would spend well in excess of what receive for the IAG if we looked at the isolation and receipt of all of the isolates and what it would cost.
  
In addition, we also have directed appropriations from within our unit and a number of permanent staff who are directly associated with NARMS.
  
DR. WALKER:  Okay.

DR. MILLER:  Bob, just looking at the possibility of any redundancies, at one time CVM was talking to the sponsors about doing post-marketing surveillance, either helping to pay for NARMS or doing their own post-marketing surveillance.  Particularly of the new fluoroquinolone approvals. 
  
Was that ever set up and are there data available from any post-marketing activities?
  
Also, at one point CDER was looking to set up a resistance monitoring, sort of a clinical failures sort of database, and has that been done?
 
DR. WALKER:  Dealing with the CDER question first, as far as I know, CDER has not done, and it is probably primarily due to budgets.  There also are two excellent surveillance systems in human medicine that may be able to address those issues.  One is Focus Technologies and the other is Sentry, which are very extensive surveillance monitoring systems that deal with hospitals throughout the country or throughout the world.
  
In terms of post-marketing for CVM, there is not a specific post-marketing program that has been set up.  The surveillance system that they have established is NARMS and NARMS deals with the pre-marketing and the post-marketing, to the pre-approval and the post-approval.  In other words,we would like to think that the data we generate is used by the pharmaceutical industry to help them in their drug approval process.
  
In fact, we had a VMAC Meeting this last year where one of the drug companies actually used some NARMS data to fill in some information for the Guidance 152 Document.
  
What we are seeing in terms of susceptibility isolates or susceptibility profiles on these organisms coming from retail meats, coming from zoonotic foodborne pathogens would represent a post-marketing approval for drugs that are being used in veterinary medicine, and we are seeing changes in susceptibility in the zoonotic foodborne pathogens.
  
DR. ANGULO:  I think one of the important background pieces that I think we should provide is that NARMS really went through a remarkable growth spurt.  I mean, we grew to this budget, our current budget approaching $2 million in this five-year window and had rapid increases, and that was because resources were provided to FDA.  Actually, the lead agency is CFSAN through the President’s -- at that time it was the President’s Food Safety Initiative.
  
President Clinton had a five-year Food Safety Initiative which brought important new resources to the government.  The first year award, for example, was $36 million and that money then got allocated to the different agencies that work in food safety, which was largely the Center for Food Safety and Applied Nutrition at FDA.  But a piece of that went to CVM for NARMS, and almost all of the CVM food safety allocated money has been used to support NARMS.
  
A piece actually went to CDC to support surveillance activities and food safety, which was FoodNet and PulseNet, which were largely funded through the Food Safety Initiative, and each year the Food Safety Initiative during this five-year period grew and had more resources.  And therefore, the CVM slice of the Food safety Initiative grew and NARMS resources grew.
  
That initiative is no longer active.  In fact, this year we see a decline in food safety resources
government-wide and it is quite marked actually at CFSAN because CFSAN had the biggest piece of the Food Safety Initiative.  It was also quite marked at CDC in our other activities related to food safety.
  
And also, as expected, we have a decline this year in CVM.  In fact, every year that there is not an increase there is a decline because we -- I hope people noticed on the breakdown that by far our biggest part of the budget, of course, is personnel and personnel have a seven percent increase every year.  So, if we don’t have an increase in the budget, in fact, it results in a decline.
  
So this year, the fact that we had a decline in the order of five or eight percent each, actually represents a 15 percent decline in our budget because of the increases in personnel.  All that is to say it is just a different climate in atmosphere in the government in terms of resources and all programs are being reduced, and we can clearly anticipate, in fiscal year ‘06, a decline in resources in food safety and a  decline, therefore, in NARMS.
  
I would expect that we would anticipate a
10-percent decline in our real resources, and we are wrestling with how to deal with that and I am sure other agencies are wrestling how to deal with that too.  But there is not anything on the horizon that we are aware of that would replace these funds. 
  
There is not new government initiatives in food safety, there is not a government-wide initiative on antibiotic resistance, which we have been trying to -- various groups have been trying to do.  But there is not a new initiative that would bring new resources for antibiotic resistance.  I think it is just the new reality that we can expect continued declines in the next five-year cycle.
  
DR. WALKER:  I appreciate what you are saying, Fred.  I think, with that in mind and we look at the 2006 and 2007 budget, while we don’t know what those budgets are going to be at this point in time, I think that we can safely assume that they will either be the same or a decrease.
  
And if they are the same, then just the cost of personnel is going to be a decrease to all of us.  So I think what it means is those of us who are involved in this work really need to look at the work that we are doing.
  
And going back to Lyle’s question of how can we sustain or enhance the funding, I think it is looking at the quality of the data that we are generating and making sure that all of that data that we generate maximizes the potential for human health care, and there are a number of ways of doing it.
  
One of the things that you will hear in this next couple of days is the reporting system that is going to be coming forth from the three different arms.  While it is important to generate data, it is also important to be able to report the data and do so in such a way so as to increase the visibility and the utility of the NARMS program to the country.
  
DR. ANGULO:  I agree complete.  And just to emphasize -- it gets exactly at Sue Kotarski’s point, which is to reassess the goals of NARMS and what are the real core functions, because the reality is we can expect these really steep cuts.
  
I have seen the pass through budget on the Food Safety Initiative for next year, and we are all anticipating important cuts in the food safety.  You are optimistic to expect equal funding.  Every scenario that we have worked on is anticipating deep cuts in food safety next year.
  
DR. WALKER:  Yes, Dan.
  
DR. SAHM:  Just a follow up on this topic and a point that Marissa brought up and then a question.  On the human side, as part of a new drug application, CDER does want to see the sponsor’s commitment to post-marketing surveillance, and it is on their dime to see how their drug continues to perform once it is being used in the clinical environment.  So, the precedent is set there on the human side.
  
So my question is how difficult or how much of an uphill climb would it be for CVM to have that same expectation for clients who are applying -- for sponsors applying for new drugs used in the animal industry?
  
DR. WALKER:  Well, that is a good question and that is a very complex question.  There are a couple of major differences between human medicine and veterinary medicine, in terms of the microbiology.
  
The first is that all of the human laboratories, or at least theoretically all of the human laboratories generating this information, do so under fairly regulated quality control testing methods, and they are tested I think twice yearly to assure that they are adhering to appropriate standard, quality control and testing methods.
  
One of the things that facilitates that is as a drug in human medicine moves through the approval process, one of the very first things that they do is they go to a contract laboratory and ask that laboratory to generate testing methods and quality control ranges for their drugs.  So then, when they go into the clinical phase of their durug testing all of the isolates can be tested under standardized testing methods.  From this the sponsor can establish a baseline data representing the pathogens susceptibility to their drug at the time of approval.
  
In veterinary medicine a pharmaceutical company does not have to go through quality control testing prior to the drug approval.  In other words, a lot of times sponsors will present drugs packages to CVM where they have generated clinical data, but it was generated without the benefit of QC organisms and quality control testing.  Without the benefit of appropriate testing methods, including the use of QC organisms and QC ranges the sponsors have no point of reference once their drug is approved.
  
The other thing is that a lot of times when drugs
gets approved in veterinary medicine, they do not have to go to CLSI for QC testing or interpretative criteria.  The pharmaceutical company can provide values to the laboratories that they have decided for QC range should and interpretive criteria without the benefit of a appropriate testing procedures but rather based on marketing strategies.
  
The other thing that comes into play here is that the veterinary diagnostic laboratories, or the laboratories doing the isolation, identification and susceptibility testing for veterinary isolates, while many of them may be laboratories associated with human medicine, the pathogens they encounter from animal sources are going to be different than what they normally see.  On the other hand, the majority of the laboratories processing specimens from animals are not human clinical laboratories and as such they do not use  standardized testing methods including an appropriate quality control program that  monitors the quality of data generated by those laboratories.
  
So again, we are dealing with, in human medicine, microbiological procedures that are fairly tightly controlled and regulated.  In veterinary medicine we haven’t reached that stage of perfection yet.  So I think it would be difficult for the pharmaceutical companies to generate the type of information that you are talking about.
  
I think it could be done if there was a  concentrated effort to get this ball rolling.  Probably within five years.  But I don’t see the concentrated effort being made to do that.
  
DR. SAHM:  Thanks.  That helps understand, Bob.  Just as a follow up to that, the guidance for industry from the FDA on the human side didn’t always require this either.  In fact, one of the major sections for the NDA in microbiology now is how well does your drug perform according to standard methodology?
  
It all rolls together, as you have pointed out.  So you can’t do one without first expecting that the clients or the sponsors or whatever, when they submit a drug -- do you have a good way of testing in the clinical laboratory?  Maybe the analogy would be do you have a good way of diagnostic veterinary laboratories testing your drug in the real world?
  
It does go hand-in-hand.  I understand what you are saying.
  
DR. WALKER:  I have looked at some packages that come in where the pharmaceutical company says that they have done the testing in accordance with CLSI.  And so, my question would be what QC organisms did you use, how often did you use them and what was your dilution range?
  
And when I get the information back, I find that they used the four standard QC organisms, but two of them had no QC range for the drug that was being tested.  One of them had a QC range below the dilution range that was tested.  The other one had a QC range above the dilution range that was tested.
  
So, in actual fact, they were using the CLSI QC organisms, and as such said they were following CLSI guidelines, but in actual fact, there was no QC there.  This is  a problem.
  
When I was in the vet diagnostic arena, that was one of the things that we tried to do; was to establish a quality control testing method among the participating diagnostic laboratories.  We tried to get all of them to buy into the CLSI testing procedures.
  
There are two other problems that we have in relation to this.  One is the veterinary pharmaceutical industry is somewhat reluctant, and perhaps rightfully so, to go public with their drug too early in the process because that gives their competitors an idea as to what is coming out.
  
The marketing in veterinary medicine  is very important.-- I have heard that in human medicine they won’t even consider an anti-infective unless it is going to make $1 million a day.  That is $365 million a year.  Veterinary medicine, a good product may make -- and Sue can tell me if I am wrong.  It may make $100 million a year.  Considerably less than anything that is a poor drug in human medicine.
  
The other thing is that if a sample goes into a human diagnostic lab or a human clinical micro lab for culture and susceptibility, I would imagine it could cost upward of $75 or more.  I know of some vet diagnostic labs that charge nothing because the money is just not there.
  
I know when I was in that arena I charged $25, and when I raised the price to $28, you could almost hear the "ah," but it was just to do the quality of testing that I thought we needed to do.

Okay.  Thank you.
  
DR. YOUNGMAN:  Thank you very much, Dr. Walker.  The next thing on our agenda is a 15-minute break.  We are going to try to keep on time as best we can, but I imagine we can’t really do a 10-minute.  So, if we could come back in 15 minutes, at about 10:35 or thereabouts, please, and we will start again.  Thank you.
  (Whereupon, at 10:21 a.m., a recess was taken.)
  
DR. YOUNGMAN:  In the interest of trying to keep on time, if we could start again, please.  I just want to remind everyone that the main purpose of our meeting today and tomorrow is to try and address the questions that are in the booklets that you have been provided that are headed with discussion points.
  
So, as the talks are occurring today, if you could consider those questions, those are the things that we would really like to hear your individual responses to.  And if we could keep the focus on that, that would be really, really helpful.
  
Our next speaker is Dr. David White, who is going to be talking about the Retail Meat Arm of NARMS.
  
DR. McEWEN:  In a quick leaf through here, I am not sure the questions are in here.
  (Pause.)
  
DR. YOUNGMAN:  I’m sorry for the confusion.  There were two different booklets that were passed out.  The panelists’ booklets do not have the discussion points in the front.  The booklet that I got does have the discussion points in the front.  We will make sure that the panelists get copies of the discussion points.
  (Pause.)
  
DR. YOUNGMAN:  I apologize.  We will make sure that the panelists get copies of the discussion points.  They were in the larger booklets that I had seen that the participants got.  I apologize for that, but we will make sure that you get them.
  
DR. McEWEN:  One other procedural point.  Do you suggest going through these question by question at some point?  You know, the panel member offering their perspective?  Or do you see us interjecting as we go along?
  
DR. YOUNGMAN:  Well, if you check the agenda, those are points that are going to come up later in the day.  Each of the questions that are part of the discussion points will be raised later in the day.
  
Okay.  And if I can, I will turn it over to Dr. White.
  
DR. CHILLER:  Dave, I just wanted to say one thing.  Tom Chiller.  I passed out to each of you guys CDC’s external review that was conducted last year.  It has the reviewer’s summary comments to the different questions we addressed.

Also, highlighted is sort of CDC’s response.
  
It is just an internal document that we put together, but I thought it might be helpful at least this evening, when you are sort of digesting all of this information, to see what we had done last year.  Obviously I will be talking about that in just a couple of talks from now too.

Retail Meat Arm
by Dr. David White

DR. WHITE:  I am going to take, hopefully, the next 20 to 25 minutes to talk about my real job.  I am a team leader for the NARMS retail part, and I would like to share with you how it started, the history of it and some pre-work we did before we went into the NARMS to become the third arm and end with a little of data.
  (Slide)
  
Of course, as background, the food supply, including meat and poultry, is an important source of enteric bacteria, including Salmonella, Campylobacter, E. coli and possibly enterococci.  I think the debate is still on enterococci coming from foods as pathogens.
  
Antimicrobial resistance among these foodborne bacteria is not uncommon and is often associated with the use of antimicrobial agents in food animals.  Retail food represents a point of exposure close to the consumer, and when combined with data from slaughter plants and on-farm studies, may provide an indication of the prevalence of resistance in foodborne pathogens.
  (Slide)
  
So, I would like to start with the history of microbiological surveys, focusing on animal derived meats from CVM’s perspective, and our primary objective has always been to characterize antimicrobial resistance among isolated from retail meats of animal origin in meat and poultry.
  
I will talk about three studies in particular.  The first one was our collaborations with the University of Maryland.  In particular, Dr. Jianghong Meng, looking at the greater Washington, D.C. area as our sampling areas.
  
We will talk a little bit about the Iowa project, which was our pilot project for NARMS retail, and then how we expanding into retail meats as part of the NARMS program.
  (Slide)
  
So, we started off with back in 1998/1999, partnering with Dr. Jianghong Meng, and we initiated a study looking at retail raw meats in the greater Washington, D.C. area.  We tested 825 samples of retail raw meats for E. coli and Salmonella, 719 for Campy and these were random samples taken from 59 stores, published in the Applied Environmental Microbiology in 2001.
  
Of course, one of the things that catches your eye, in terms of poultry, we had 71 percent positive for Campylobacter.  So, it was one of the first indications that Campylobacter was highly prevalent in chicken breast.  Now, this is pre-HACCP.  Okay?  These were samples pre-HACCP.
  (Slide)
  
We followed up with another study with Dr. Jianghong Meng that we were fortunate enough to publish in the New England Journal of Medicine looking at ground meat samples from the same area, and we looked at chicken, beef, poultry and pork, and these were ground samples.  We were able to go back and find out that there were four poultry processes, one pork processor and beef was ground in the store.
  
Again, we found Salmonella.  In this case from 20 percent of the ground samples.  We found a variety of serotypes, most frequently isolated from poultry meats, and we found eight isolates of Typhimurium.  The majority of those were DT104.
  (Slide)
  
And just quickly.  I don’t know if we can see this.  This is what caught our interest.  When we looked at those Salmonella isolates we did recover from the ground meats, we found the multi drug resistant to several antimicrobials.  In particular, S. agona.  We saw a lot of beta lactam resistance, strep-sul-tet and trimethoprim.  We did find some DT208s which were resistant to about nine to 12 drugs.  Very interesting.
These appear to be all plasmid mediated as well.
  (Slide)
  
So, these two studies led us to think that we needed to investigate further the role of retail meats as reservoirs of resistance, and we initiated a study called the Iowa Retail Meat Pilot Study in 2001 and 2002, and then we segued it into NARMS, the FoodNet -- I will go over the FoodNet Retail Meat Surveillance.  The first year was 2002.
  (Slide)
  
In terms of the Iowa Study, this was started in 2001 and goes into 2002.  We went to 24 samples a week, and

Dr. Terry Proescholdt will talk about sampling later on and how we developed the study.
  
But we picked four meats, much like we had done in the other studies in D.C.  Ground turkey, pork chops, ground beef and chicken breasts; 24 samples a week.  They went to six grocery stores and they picked four meats of each for 24 samples per week.
  
And ideally, what we were trying to determine was the prevalence and susceptibility patterns of Enterococcus, Salmonella, Campylobacter and suspect extended spectrum beta lactamase producing gram negative bacteria.  This goes back to Dan’s questions.
  
As part of our study we are trying to determine if we could find ESBLs on foods, and we did a whole screen with ceftazidime and cefotaxime with one microgram per ml.  The Salmonella and enterococcal plates were -- we used the NARMS plates to determine susceptibility, and for Campylobacter we used agar dilution and E-test methods.
  (Slide)
  
So, I will just give you some brief data.  The studies I mentioned went on for 15 months.  We had 24 samples per week.  We ended up sampling about 263 grocery stores; 209 for Campylobacter.  It took us several months to optimize the Campylobacter methods.  Overall though we had almost 1,000 meats analyzed, and as I mentioned, we looked for Salmonella, Campy, Enterococcus and ESBL-producing gram negative bacteria.
  
Susceptibility testing was done using the NARMS panel or agar dilution.  All Campylobacter and Salmonella were subjected to Pulsed Field Gel Electrophoresis as well using two enzymes, and we just finished and it has taken us two years to do that.  For those of you that do pulsed field, doing one enzyme is hard enough.  When you have to do two enzymes, you double your amount of work.
  
We were also able to get the human isolates from that same time from the State of Iowa.  So we were able to have comparisons from what we recovered on food to what was submitted to the clinical diagnostic lab at the University of Iowa as well.
  (Slide)
  
So, some quick data on that.  Of the 981 samples, 126 were positive for Salmonella.  So, about 13 percent overall.  Ground turkey and chicken breast accounted for the majority of those.  Forty-one percent of ground turkey samples were positive for Salmonella and 12 percent on chicken breasts. 
  
Overall, 21 percent of samples were contaminated with Campylobacter, the majority being chicken with 75 percent.  So, if you remember back to our original study in Washington, D.C., it was about 71.7 or pretty similar.  We are talking almost three quarters.
  (Slide)
  
And in terms of susceptibility, one thing I didn’t point out is that we did not recover Salmonella from any pork chop sampled.  They were completely free of Salmonella.  And all of our Salmonella from beef, there were five of them, were completely susceptible to all antimicrobials tested.
  
We saw the majority of resistance to isolates from either poultry, ground turkey or chicken breasts, and all isolates were susceptible to ciprofloxacin, amikacin and imipenem.
  (Slide)
  
In terms of pulsed fields, we just want to show you a couple of things where we did do Pulsed Field Gel Electrophoresis.  This was on Salmonella Heidelbergs that we found.  We did get several hits in terms of indistinguishable pulsed field patterns between those recovered from ground meats in Iowa and also those recovered from human illness.
  (Slide)
  
In terms of Campylobacter, this is the breakdown of what we found.  We found primarily the Campylobacter in the chicken breast, and we also had about 181 isolates that we had from humans that we were able to compare as well.
  (Slide)
  
This is just a -- I will give you quickly some ciprofloxacin resistance.  This is based on agar dilution in an interpretative resistance breakpoint of four µg/mL.  As you can see, that is supposed to be a micro.
  
But just to show you overall, 19 percent of the C. jejuni from chicken breast were cipro resistant at four microgram per mil or greater, 22 percent of C. coli.  None of the other C. jejuni were resistant to cipro from any other meats.  And overall, Campylobacter recovered from humans were about 12 percent resistant.  So, we are talking in the teens for ciprofloxacin resistance.
  (Slide)
  
Again, we did pulsed field on all Campylobacter  jejuni and on several occasions we got identical pattern matches between those Campylobacter jejuni from retail meats and from human illness in Iowa.
  (Slide)
  
Now, during the time of this study the WHO recommended a tripartite approach to include isolates from human clinical cases, food animals and retail meats when conducting antimicrobial resistance surveillance and monitoring for foodborne pathogens.  So they recommend this three-prong system of surveillance, if you are looking at foodborne pathogens of on-farm, slaughter, retail, slaughter/retail and human isolates.
  (Slide)
  
So therefore, in 2002 NARMS teamed up with FoodNet out of the Centers for Disease Control and Prevention in developing a program aimed at determining susceptibility of bacteria isolated from retail foods of animal origins, and this has become the third part of NARMS and the most recent.  It is located at the Office of Research in Laurel, Maryland.
  (Slide)
  
So, the overview.  We started in 2002 with six of the 10 FoodNet sites.  We had initially Connecticut, Georgia, Maryland, Georgia, Maryland, Minnesota and Tennessee started in January 2002.  Oregon joined us much later in the year, in September.  In January of 2003 we added New York and California, and in January of 2004 we added Colorado and New Mexico.
  
And currently we have 10 of the 11 sites.  I think the 11th site is Texas.  Correct?  And they have yet to join our surveillance system yet, but they are still in their first year of FoodNet as well.  So, we have the 10 major players.
  
Each of the 10 sites has a similar sampling scheme where the sites visit at least one grocery store per month.  In the past, between 2002 and 2004, they did this.  In 2005 we introduced a random sampling scheme.  Again, Terry Proescholdt will talk about that later on.
  
Each site purchases 40 meats per month.  Ten packages each of chicken breast, pork chops, ground turkey and ground beef.  All 10 sites culture the rinsates for Salmonella and Campylobacter, and due to the large numbers we recovered of Enterococcus and E. coli, we only have four sites culture the rinsates for E. coli and Enterococcus.
  
And I may have a slide to show you that.  The reason why is our Enterococcus were close to 95%  contamination.  So, if we had all 10 sites doing that, we are talking 5,000 Enterococcus isolates per year.  It would quickly overwhelm us.
  (Slide)
  
This just gives you a picture of the laboratories; where they are disbursed throughout the United States.  You can see we have Oregon and California on the West Coast, Colorado and New Mexico, Minnesota, Tennessee, Georgia, Maryland, Connecticut and New York.  So, these are the sites that are participating with us currently in the sampling.
  (Slide)
  
As I mentioned, we used similar standardized methods at each of the sites to isolate the respective bacteria.  They are adapted from the FDA BAM Manual.  All of the meats are initially rinsed in peptone broth and then the peptone broth is then put into the selective broth, depending on what isolate we are trying to recover.
  
And for Salmonella there are several broths.  Lactose broth, RVR10 and the M Broth, and we use a Tekra EIA Kit.  For Campylobacter we use Bolton broth.  For E. coli,  MacConkey broth.  Enterococcus is enterococcosel broth.  All of these broths have been incubated at appropriate incubation temperatures for an appropriate amount of time and then plated onto selective agars to isolate the respective bacteria.  XLD for Salmonella, CCA Agar for Campy, EMB for E. coli and Enterococcosel agar for Enterococcus.
  
The first two years we provided training where, we had the sites come out to our facility, and we had a week-long training to show them how we were doing it, because we wanted to make sure that all of the sites followed the same methods.  It is critical.
  (Slide)
  
So, the sites are the ones that isolate the bacteria, and once they isolate it, they presumptively identify it.  Then they send it to us at the Office of Research for confirmation of identification and antimicrobial susceptibility testing.
  
We confirm all bacterial species by biochemical identifications, primarily using the Vitek, and we also have API if we have some disparate results coming from the Vitek.  We have PCR capabilities for Campylobacter speciation using several genes that are specific for lari*, jejuni and coli.
  
We have our own serotyping laboratory that serotypes all of the Salmonella isolates that come again.  And again, we do susceptibility testing, which is our bread and butter.  We use broth microdilution/agar dilution, when appropriate, and as I mentioned before, we follow CLSI/NCCLS standards where available.
  (Slide)
  
And just to give you an idea what happens in this whole scheme of collection and reporting, the sites go out and buy the meat.  They isolate the bacterium from the different meats.  Again, on the selected medias.  There is a log sheet.  Dr. Elvira Hall-Robinson will talk about this later on.  It goes into our database.  It is an access database.  Again, that will be talked about later on by
Dr. Hall-Robinson.
  (Slide)
  
As part of the retail arm, all Salmonella and Campylobacter isolates are subjected to PFGE, which is a big burden on us, but we do them all.  I think it is very important information.
  
We follow CDC guidelines for PFGE analysis and
CVM-OR is a PulseNet certified laboratory.  Our PulseNet unit is headed up by Dr. Shaohua Zhao, who will talk about that later on.
  
All of our data goes into the PulseNet program, and our isolates can be used for future research projects for anyone who is interested in these isolates as well.  We have been involved in biosource tracking experiments, virulence studies and, of course, resistance studies when we try to find particular genotypes associated with particular phenotypes.
  
Enterococcus and E. coli.  Right now a lot of these should have question marks next to it because we have these isolates.  We are trying to determine what to do with them in terms of follow up studies.
  
Some questions that have come up are what virulence factors are present in Enterococcus isolates and are they present on the ones we see in foods?  Streptogramin resistance, of course, is a big interest to CVM, and we have initiated some studies to determine what streptogramin resistance genes are there.
  
We find that in a lot of our Enterococcus isolates that show high levels of streptogramin resistance do not have the typical genes that people have found in the literature.  So, there are genes yet to be identified, I believe, in these isolates.
  
E. coli.  We are going after generic E. coli.  We are not going after 0157 or shiga-toxin producing E. coli.  So, it is important to serotype these isolates?  Jim Johnson’s work out of Minnesota is showing that some of these E. coli from foods are potentially urinary tract pathogens.  We don’t know that.  And we do see fluoroquinolone resistance and ESBL phenotypes in our E. coli, and we are working those up via PCRs for mutation analysis and gyrase* and so forth and what type of beta lactamases are present.
  (Slide)
  
Just to give you some data on the three years, what I wanted to point out here is the number of meats that have increased from 2002 to 2004.  Our first year we had approximately 2,500 meats under analysis.  As of 2004 we were almost up to 5,000 meats.  So we are talking 4,600 and 4,700 meats in terms of our preliminary data, and the numbers are pretty stable across the board for Salmonella.
  
So, chicken breast was about 10 percent to 13 percent over the three years.  For ground turkey it is about 11 percent to 13 percent again down to 12 percent.  Our ground beef is almost identical over the years in terms of percent positive, and our pork chop is pretty similar as well.
  But it is interesting.  You can see it is mostly coming from poultry products.
  (Slide)
  
This is to compare two years of data for Salmonella, 2002 versus 2003, and I tried to group the beta lactams together on the left, then your aminoglycosides and then our other drugs.  You can see where we see a little more resistance in 2003 to several drugs, with the exception of TET where we did see a decrease.  So, we don’t have 2004 data yet, but I would be curious to see what we get.
  (Slide)
  
Again, I just wanted to show you that we are doing pulsed field, and Dr. Zhao will talk a little bit about this.  But we do see some hits between clinical isolates and what is getting through into the ground products.  Where it says turkey, they are actually clinical isolates from the diagnostic lab. 
  
Again, we see another hit here.  We see three indistinguishable clusters here, and this big cluster is mostly ground turkey.  You can see from multiple states.  Tennessee, Connecticut, Minnesota and different sampling times as well.  So, we are seeing the same clonal isolates out there in the food supply.
  (Slide)
  
In terms of Campy for the three years, it does appear like we have an increased rate of isolation of Campylobacter.  I think a lot of this is due to the laboratories improving their methods of isolation.  Our first year we had a lot of isolates sent to us from the laboratories that were indeed not Campylobacter.
  
Remember, we confirm everything at OR.  So we had to throw a lot out, because our initial rates were pretty similar to what we see here in 2003 and 2004.  But again, you can see we are not seeing much Campy coming from any other of the three meats sampled.  Primarily, when we recover it, it is coming from chicken breasts.
  (Slide)
  
And here is two years of data on resistance.  This is agar dilution data.  In 2002 we had approximately 300 Campy isolates.  In 2003, 464.  Our ciprofloxacin is pretty stable. About 14 percent to 15 percent.  Erythromycin went down from six percent to about three percent.  Doxycycline is fairly high, about 27 percent to 28 percent.  Gentamicin we -- and remember, these are interpretive criteria or breakpoints that we have determined.  They are not CLSI/NCCLS interpretative criteria.
  
We did see our first Gentamicin resistant Campylobacter this year, and it is at an MIC of 32 microgram per ml.  And interestingly, in erythromycin here these are all C. coli’s.  We don’t have a single C. jejuni that is erythromycin resistant.
  (Slide)
 
 On E. coli, I will quickly go over this.  This is, again, three years worth of data.  The numbers, remember, are lower because we only have four sites doing E. coli, and enterococci versus the 10 sites for Salmonella and Campylobacter.  But your numbers are pretty steady for the three years.
  We do seem to see this reproducible phenomenon with pork chops.  They seem to be less contaminated with E. coli than the other three meat types.
  (Slide)
  
And again, this is comparing two years of data.  You can see almost the E. coli from 2002 and 2003; they are almost like mirror images across the board in terms of percent resistance to the drugs under surveillance of NARMS.
  (Slide)
  
One thing we are trying to do now is to pull out to see if we see certain phenotypes associated with certain meat products, and this is an example of what we can do.  If you look at gentamicin and nalidixic Acid, among those isolates that were GEN resistant and NAL resistant, the majority of them are coming from poultry products. 
  
We see very little GEN or NAL resistance in other ground beef or pork chops, and they are primarily coming from chicken breast and ground turkey.  So, we can do this for all of the drugs under surveillance at NARMS.  At least for the retail arm.
  (Slide)
  
In summary, since its inception in 2002 NARMS retail has increased the number of sampling sites and retail samples analyzed and improved its sampling scheme.  For example, six sites in 2002 to 10 sites now in 2005.  We have almost doubled the number of meat and poultry samples under surveillance, about 2,500 to almost 5,000, and we introduced random sampling strategy in January 2005 that will be discussed later on.
  
Its primary purpose is to determine the prevalence of antimicrobial resistance among foodborne pathogens and commensal organisms, and the commensals that we are looking at, of course, are Enterococcus and E. coli as potential reservoirs of resistance from retail foods.
  
We hope the results will generate baseline data and establish a reference point for analyzing trends of resistance among these foodborne bacteria at a point closets to the consumer.  As I mentioned earlier, when combined with data from the rest of the NARMS program, it can give us an idea of what is happening and where we can target intervention and mitigation strategies.
  (Slide)
  
With that, I would like to thank -- there are a lot of people involved in this project and there are certainly many more players, but this is a good collaboration between CVM and primarily CDC, because without their help we can’t work with the FoodNet sites as well.  So, thanks.  I will take any questions.
  
DR. VOGEL:  I have got several questions.  I have always been curious why you chose ground turkey as one of your products to sample.  It doesn’t seem to me that per capita consumption is very high of ground turkey.
  
DR. WHITE:  That is a great question, Lyle.  During the Iowa Pilot Project we tried to determine what would be the most appropriate and the best handled in the laboratories, and we found out that buying the big carcasses was not the best way to go, because they were Fed Ex’ing them to us.  So we tried to go with a product that was easily obtainable.
  
I think Terry will talk about it later.  Ground turkey is actually the fastest growing commodity in the turkey industry being sold in grocery stores.  So, that is why we went with ground turkey.  But we could have easily gone with turkey legs and turkey thighs.  Absolutely.
  
The thing to remember too is that USDA has performance standards for ground turkey.  So that is one other thing we were looking at as well; to try to use that data to maybe help some other agencies when we get some data.
  
DR. SAHM:  Thanks, Dan Sahm.  Just in terms of trying to keep questions on target for the discussion points, with regard to sampling, now that what you have learned this data is sort of what we talked about at break.  In looking at this data, if I only had a dime to spend I would put it on poultry. 
  
How much information are you getting out of the other meat sections and would you not concentrate on poultry as being the highest risk of exposure to antibiotic resistant organisms for this program?
  
DR. WHITE:  I think for the overt pathogens.  Yes.  They are primarily coming from poultry products.  We are not getting that many isolates from either pork chops or ground beef.  Absolutely.  So in terms of bang for the buck, yes.  We are not getting that much for those meats.
  
DR. SAHM:  What would the risk be of going forward and not pursuing those other meats at this point if you wanted to concentrate more thoroughly on -- if people would agree that is where the risk is, to put your efforts in that area.
  
DR. WHITE:  That is a great question.  I think the major risk would be we would want to have information from those particular food commodities, and as we approve drugs in numerous animal classes we would only have information then from poultry.  We would not have that information from say swine and from the resulting meats that are obtained from the swine, as well as cattle as well.  So, we wouldn’t have that information, and we are trying to look at drug use in certain animals with what might be happening.
  
We wouldn’t be able to do that connect between say approval -- not to point out Sue’s drug, but ceftiofur is approved in numerous animals species.  Without looking at the different foods, we would not be able to see if there is anything going on in terms of decreased susceptibility or what have you.
  
DR. SAHM:  But in our surveillance efforts -- I just want to pursue this a little more to think through it.  You learn from your baseline data and you redesign.  It is analogous to your decision not to include imipenem anymore.
  
I just put that out as a point to consider based on the progress of surveillance of learning what you know and where you put your efforts going forward.
  
DR. WHITE:  I think it is a good point.  Thank you.  I agree.
  
DR. ALTEKRUSE:  I would just like to say from the FSIS standpoint we believe that also the greatest risk is for poultry and Salmonella.  Our programs are being sort of reconfigured to address that.  So we are going to put greater emphasis on the testing of poultry products.  That will have some implications for sampling issues later related to the FSIS isolates.
  
But I am also very intrigued in the finding of matches with the PFG patterns between human and food isolates, and I would like to hear your opinion on the potential of that to actually detect outbreaks or identify sources of infection.
  
DR. WHITE:  I think that is probably why we do it.  In terms of outbreak assistance I doubt that will be of much help at this point because we are not real time.  Our isolates are going in probably two years from when they were collected.  So really, what we are trying to do is expand the data at PulseNet so that if there is an outbreak and they submit a pattern and it does have a hit with one of ours, maybe they can start focusing their efforts on a particular food.
  
So, if they say I have a Salmonella Heidelberg and it is a match with a Heidelberg from our retail foods from ground beef, maybe that will help the health personnel to start focusing on where it is going to go.
  
However, we do have numerous pulse field patterns that are identical among the different meats, like Newport or Typhimurium.  So, it depends on the particular organism as well.
  
DR. ALTEKRUSE:  Just to take it one step further, the reason I am interested in that is because one of the triggers that FSIS that FSIS is likely to use for going into an establish and doing a really intensive assessment of their food safety system is the finding of some epidemiologic evidence, like a pulsed field match between human isolates and isolates coming from that plant.
  
DR. WHITE:  A lot of it too -- when people do those comparisons without, I think, a good epidemiological study to back that up, it is -- it just tells you there is a hit, but we need to do a lot more in between to see what is going on.  So that is why we do the pulsed field. 
  
I think it is very important to expand that database.  They don’t have much in their database from actual foods.  Now we are expanding with Paula’s isolates and CDC’s isolates as well.  Hopefully, we will get all of those in the system, and when we do have outbreaks they may be able to better target their intervention strategies.
  
DR. ALTEKRUSE:  I would like to ask you how -- what were the criteria to select the 10 sites that you include?
  
DR. WHITE:  Okay.  That is a great question.  The 10 sites were already existing as part of the FoodNet system.  So, they were already in existence.
  
DR. VOGEL:  In one of your slides you mentioned  that it was a samples from a diagnostic laboratory.  Are you obtaining samples from diagnostic laboratories yourself?  Or is that a sample that was obtained from USDA and then shared?  If you are doing it yourself, it seems like it is duplicative use of sources of resources here.
  
DR. WHITE:  Yes, I agree.  That was a diagnostic isolate from another study.   It is not from a NARMS study.

Remember, a lot of us are also Office of Research principle investigators.  We have our own studies underway as well besides NARMS.  That was a study that Dr. Zhao and Dr. Walker are PIs and were working with diagnostic labs.  But we do  the same thing.  We do susceptibility testing and pulsed field.
  
So then, when we run it through the system and we get hits, it would just happen to come out as one of those.
  
DR. VOGEL:  Can I ask another question?
  
DR. WHITE:  Sure.
  
DR. VOGEL:  On Campylobacter you were testing for Doxycycline why did you choose that instead of Tetracycline like the other arms?
  
DR. WHITE:  Yes.  Doxycycline I believe would be the actual drug that would be used to treat human illness.  I think more so than tetracycline.  But we have gone back to tetracycline for the microdilution.
  
DR. KOTARSKI:  Yes.  I come back to a comment that I made earlier in understanding the purposes of the sampling strategies.  Currently it looks like you are consisting sampling the same number from different -- getting the same number of meats from each store.
  
So, on the surface it looks at your sampling strategies are designed to look at prevalence of Salmonella or Campylobacter of meats in stores.  It is not set up exactly to examine exposure to humans.
  
DR. WHITE:  I agree.
  
DR. KOTARSKI:  Okay.  So that is understood.  Now you have generated data to show that the prevalence of Salmonella actually is very low in certain meats and that point has been made earlier.  I haven’t looked in detail at years 2003 and 2004, but I do recall, for example, your ground beef data for 2002.
  
You had very few isolates.  On the order of nine Salmonella isolates, as compared to hundreds maybe of isolates in poultry.  That being the case, to look at resistance trends of nine isolates in ground beef from one year to the next, probably your sampling error associated with only obtaining nine isolates of Salmonella, for example, in ground beef does not permit you to look at temporal trends that you would like to do without a lot of variability, just because of the number of isolates that you get.
  
So, I come back to the original point.  What is your objective in your sampling strategy for the retail meats?  Is it to understand the prevalence in meats, per se?  Understand resistance prevalence in retail stores?  Or is it to understand the prevalence in food consumption?
  
I think that those objectives should be clearly stated and then select your sampling strategy accordingly.
  
DR. WHITE:  Okay.
  
DR. ALTEKRUSE:  Many of the serotypes that you are isolating, for example, from poultry aren’t serotypes that are associated with human illness, and I wonder if those get the same priority or perhaps they are tested later.  Have you thought about it?
  
DR. WHITE:  In our system?
  
DR. ALTEKRUSE:  Right.
  
DR. WHITE:  Well, the first thing is serotyping takes the longest amount of time, besides pulsed field.  So actually, as soon as we get something in it is susceptibility tested.  So, they are done right away before we do serotyping.  You know what I mean?
  
We have different groups working on the bugs at the same time.  That is the best way to get things done the fastest, not wait until they are done.  Otherwise it would take years for us to get each year done.  But the majority of our Salmonella, as we see it, are the top five that CDC sees in human illness. 
  
We do see some other ones that we don’t traditionally see in human illness, but across the board they are typical household names that most people know.  Heidelberg, Newport, Typhimurium and so forth.
  
DR. VOGEL:  You mentioned that you are sampling E. coli and Enterococci from only four of the FoodNet sites, and I understand why, because of the number of isolates.  Have you looked to see if there is any geographic variation in E. coli or Enterococci?  If so, might you be better doing a systematic sampling from all 10 sites or all 11 sites instead of restricting yourself to four sites?
  
DR. WHITE:  Yes.  In our first annual report we tried to break it down by state, and I think at least with Enterococcus we do see a species distribution with -- Maryland sees more faecalis than faecium compared to the other three sites. 
  
We haven’t look at the 2003 data yet to see if that holds up in terms of a trend.  So, it is definitely something to throw out and look at the data.
  
E. coli seems to be different, we don’t see resistance associated with one state more so than another.  They are almost mirror images for the two years.  We try to pull out those things to see if there are differences between the states as well.
  
We did that with our Campy.  If you look at Campylobacter isolation, you see large variations between the states in terms of what they isolate over the year.  Okay.  Thank you.
  
DR. YOUNGMAN:  Thanks, Dave.
 
In the interest of time, if we can press ahead, our next presenter is Dr. Paula Cray, who is going to be talking about the animal arm of NARMS.

Animal Arm
by Dr. Paula Fedorka-Cray  

DR. CRAY:  I actually did these slides so they were done yesterday morning, as per Dave.  I called him.  I said I was sending them, and my father has a dial-up connection.  I dropped my son off at my parents’ house, and I tried to download a 10 megabyte file over a dial-up connection.  It was a rather painful experience for all concerned. 
  
I thank Bob Walker, Linda English, Dave and Pat for staying later in the evening to do that.  I could have driven here faster it turned out and xeroxed them and did what I did.
  (Slide)
  
Nonetheless, this is the animal arm of NARMS.  Dave went over some of this, but I will reiterate and give you a little bit of a different perspective on some aspects.  It began as a research project, the animal arm actually did, with Dr. Leanne Thomas and myself.  Dr. Leanne Thomas was ta the National Veterinary Services Laboratories and I was a scientist at the USDA National Animal Disease Center in Iowa.
  
We got together and we decided that at the time, in 1994 and 1995, there was an increased interest in antimicrobial resistance.  So we actually bought panels that just had breakpoints on them, and we were doing the testing when we received a call from Dr. Tollefson, who had heard what we were doing, and asking us if we would be interested in joining NARMS.
  
And we were certainly interested in doing that and NARMS officially launched in 1996, and we used up our first 1,500 plates of just breakpoint and then we went to the MIC format.  Although we didn’t have comparable plates until 1997/1998.
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The evolution is that NARMS is really a passive system and the original goals and objectives were to provide descriptive data on the extent and temporal trends of antimicrobial susceptibility in Salmonella and zoonotic enteric organisms from human and animal populations.  To identify resistance that arise. To provide timely information. To prolong the life span of approved drugs.  To identify more areas for more detailed investigation and guide responsibility.
  
It is actually a passive system and it relies primarily on the submission of isolates.  And really, all three arms either relies on the submission of isolates or the submission of some type of samples.  This isn’t an active system that we have gone out and actually designed ourselves.
  
We do believe it does provide a comprehensive snapshot, and that really is what it is.  It is a one time point snapshot of food animal production through the U.S.  We have all animal species that are representative in some way, shape or form.
  
We look at small, medium and large slaughter plants now, and Dr. Anandaraman will talk about it this afternoon.  This did not start until a later date in time.  We had the plants as they were coming onboard.  So the system was not as robust as it is now until 2000.
  
We also have isolates from ill animals.  There has been a lot of discussion about the necessity of having diagnostic isolates, but we believe that these can provide information on isolates that may be emerging or that we may see in the food chain over time, because these animals are  presumed to have undergone some type of treatment over time, and while they should not be part of the actual food that is being consumed, at least in the raw way, shape or form, they may, in fact, be representative of what is to come.
  
We can see in that some respect with Salmonella Uganda, which is first emerged as a diagnostic isolate and now we are trying to see more prevalence in some of our other samples.
  
Our isolates from on-farm samples are limited.  They are limited to the NAHMS studies, the National Animal Health Monitoring System studies that occur, and they are also limited to epidemiologic studies that we might conduct or our collaborators might conduct over time.
  
However, at some point in time, and for the most part, during the year we can receive isolates representative of all geographic regions in the United States.  So we have these three sources of isolates, on-farm, diagnostic and slaughter, and we looking for Salmonella and Salmonella is our sentinel organism, E. coli, Campylobacter and Enterococci.
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This just illustrates that and shows the differences between the human and the retail.  We have tested Listeria.  The question was asked earlier this morning.  We did a snapshot a couple of years ago on about 600 hot dog isolates.  We occasionally have some coming in on various other -- from other sources now.  We actually have one student who we acquired through a reorganization who is looking at Listeria for part of his project, which is not part of NARMS.
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Again, our isolates come from diagnostic and
non-diagnostic sources.  These diagnostic isolates are presumed to be associated with clinical illness.  The animals are not likely to enter the slaughter facility.  Right now we have 12 veterinary diagnostic labs.  California, Colorado, Iowa, Indiana, Kentucky, Michigan, Nebraska, New York, Oklahoma, Texas, Washington and Wisconsin who have submitted isolates over time.
  
We have approached Minnesota about submitting isolates, and Pennsylvania will actually join us, the University of Penn, later on this year.
  
Then we also go up twice a year and we collect isolates from the National Veterinary Services Laboratories, and what we are doing is we are collecting the diagnostic isolates from there that were associated with either a primary or a secondary etiologic outcome.  So, Salmonella has to be identified as the main cause of disease.
  
However, we exclude selection from the sentinel sites so we are not duplicating isolates.  So this way we then can saturate the rest of the United States, as far as the isolates coming from other states and the various regions.
  
For our non-diagnostic isolates we presume that all of these are coming from healthy animals.  Our on-farm isolates are part of the NAHMS studies, as I said; however, there is a five-year rotation between commodity.  So, for instance, there was a swine study in 1995.  There was a swine study in 2000.  There is scheduled to be a swine study coming up within this next year.
  
But when we look at these, again, they are  snapshots of time, although they do provide a statistical representation of greater than 95 percent of animal production on-farm in the United States for that particular time period.  So, they tend to be very invaluable as far as resources because they also collect management information at the same time, which gives us an idea of what antimicrobials may have been used in practice that may be affecting outcome of the resistance patterns that we may see.
  
I regret that our APHIS partners were not invited to the meeting, because I think that they could have provided some important additional information.
  
The slaughter isolates then are rinsates, carcass swabs and ground product.  Now we are receiving ready-to-eat products and isolates from eggs.  This is a comprehensive snapshot based on the FSIS compliance.  It is related to retail.  It is not statistically significant, but

Dr. Anandaraman will address that this afternoon.
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I think it is important for you to realize for you  that for the diagnostic isolates what we are just getting are NVSL isolates in the sentinel sites, and we receive anywhere from about 75 to 500 isolates from these various sentinel sites.  So, the larger the sentinel sites, like Michigan and New York, we tend to get about 500 of their diagnostic isolates because that is about what they get per year.  So we get all of them.
  
There are some smaller sites, and we actually get all of those too, like Washington State.  For actual diagnostic submissions we tend to get about 75 to 125 different isolates.
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All of this information is collected and reports are generated and sent back to the sentinel sites, and we give them free range to publish their own information, either including us as co-authors or citing the NARMS system as providing the information to begin with.
  
The compilation of data that you see on the reports is usually all of the sentinel sites together.
  
E. coli right now only comes from our sentinel sites, as far as diagnostic lab goes.  We get very few Enterococci, too little to really count, and we do get a few Campylobacter and they tend to be sporadic.  Those are available too in ancillary reports.
  
For our non-diagnostic isolates then all of the E. coli, Salmonella, Campy and Enterococci come from the raw product, federally inspected slaughter plants and from focus studies, but there is a caveat.  Only the Salmonella comes from the actual FSIS compliance testing.
  
For E. coli, Campylobacter and enterococci then we actually take the spent carcass rinsates that are sent to the eastern laboratory for FSIS.  Once they take their aliquot for Salmonella testing, anything that is left over that is greater than 10 mils is sent to our laboratory, and from that then we can isolate E. coli, Campylobacter and Enterococci.
  
Now, the FSIS laboratories, the eastern laboratory, doesn’t just receive samples from the eastern part of the United States.  The distribution is randomized so they are receiving them from all different -- from through the U.S. on a rotational basis.  But it is important to realize that these carcass rinsates are also only coming from chickens.
  
So, the information we get for Campy, E. coli and Enterococci through non-diagnostic slaughter isolates are limited to chicken carcass rinsates from spent Salmonella samples.
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This is what I see as one of the weaknesses of NARMS when we look at critiquing our own program and how we can improve the robustness of the program.  But farm sampling and diagnostic isolates are independent of the slaughter program.  So, we don’t have a flow through.
  
We have farm isolates that are coming because of NAHMS related studies, we have diagnostic isolates coming from the diagnostic labs and then we have the compliance testing out here.  And we really don’t have an association of what is happening between the farm then and the plant.
  
We believe that while you can say that, yes, there is some association -- and, of course, what we see on the farm is what we see, in part, of what is coming in the plant.  But we can’t say a lot of times what the use is associated at a plant, particularly with antimicrobials, and what is occurring then in the slaughter plant.
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If we move to looking at the individual organisms, Salmonella is our sentinel organism.  We have all sources.  We receive the isolates primarily.  We are not doing a lot of the isolation.
  
These serotypes will vary over time and vary by source, and we believe it is absolutely critical to take the information and break it down by serotype and by source because we see distinctly different patterns associated with these various sources and serotypes.
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When we look at our sampling scheme, what we were doing up until just recently took us several months to complete when we would receive an isolate, either slaughter isolates, which are the actual isolates themselves received weekly, or diagnostic isolate, which is the actual isolate.
  
On farm feces we have always been the testing laboratory for NAHMS studies.  So we actually receive the feces or farm sample, and we have to go through a seven to 10-day confirmation and isolation procedure.  For this we have used our published methods, which have been compared and contrasted and now are considered to be one of the gold standards in the industry.
  
Once we had the isolate then it went for serotyping, if we were doing the isolation, or it came with the serotype if they were slaughter and diagnostic isolates. They were susceptibility tested.  Then we freeze and store all of our isolates.  So we have every isolate that we have tested since 1995 in triplicate frozen in the building.
  
And right now, we are lyophilizing everything, and one isolate will be sent to the National Germ Plasma Repository in Ft. Collins for, hopefully, lifetime storage if anything happens to the Russell Building. 
  
If we don’t have a phage type, then we have to send it back out to NVSL for phagetyping, and because of our involvement now with USDA VetNet, we send all of our isolates to our pulsed field lab where they undergo Pulsed Field Gel Electrophoresis.  And then it goes on to preliminary data and then year-end analysis.
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At the request of FSIS, who has a need now to send information back to the plant on a regular basis, we have looked at how we can compress our time frame in getting this information back to FSIS that they might be able to use.  So,  particularly for our slaughter isolates now, we are looking at if everything works right in life and we have perfect gels and perfectly susceptibility tests, about a two-week turnaround time from receipt of isolates to a susceptibility pattern and a PFGE gel picture that we could send back to FSIS which they, in turn, we understand, will send back to the plants.

Now, in an testament to the compliance testing success, we see that the number of isolates from 2000 to 2004 has gone down from about 3,500 to 2,500 or so that we receive for a year from FSIS.  The number of isolates in toto that we receive for diagnostic are on-farm is also about another 2,000 to 3,000 isolates for any particular year.
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For generic E. coli this did not start for us until 2001.  We do have some diagnostic sources.  We have on-farm sources, which we have not included as part of NARMS.  But we have those isolates available that are for our own independent tests, and again, these are from the spent FSIS rinsates and only represent poultry.  Particularly, chicken.
  
There are thought to be reservoirs of resistance genes, which is why we are still testing them as part of the program.
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And the isolation procedure is almost identical to Salmonella, except that we use com-agar* for our isolation.  I will put a caveat in here; that we have done extensive and exhaustive testing of the different medias that you use for isolation and we can say that you will have different results on occasion, dependent upon the isolation methods that you are using.
  
So, for instance, if you use Campy line agar for a Campylobacter as opposed to cephex, you will get different populations than you will get of Campylobacter jejuni or coli.  The ratios will be different, and within those different populations from those different plates you will have different antimicrobial susceptibility patterns, and I think that is important to remember.
  
From 1998, which is our Campy started, through 2000 we had Campy obtained as an actual isola