From: Dr. Rick Willer [rwiller@azda.gov] Sent: Friday, December 17, 2004 10:39 AM To: 'fdadockets@oc.fda.gov' Subject: Docket Nos. 1996P-0418, 1997P-0197, 1998P-0203, and 2000N-0504 an d RIN number 0910-AC14 United States Animal Health Association 8100 Three Chopt Road, Suite 203 P. O. BOX K227 RICHMOND, VIRGINIA 23288 804- 285-3210 FAX 804-285-3367 E-Mail: usaha@usaha.org Web Site: www.usaha.org December 16, 2004 Food and Drug Administration Division of Dockets Management 5630 Fishers Lane, Rm. 1061 Rockville, MD 20852 fdadockets@oc.fda.gov Re: Docket Nos. 1996P-0418, 1997P-0197, 1998P-0203, and 2000N-0504 and RIN number 0910-AC14 To Whom It May Concern: These comments are submitted on behalf of the United States Animal Health Association (USAHA) in reference to Docket Nos. 1996P-0418, 1997P-0197, 1998P-0203, and 2000N-0504 and RIN number 0910-AC14, Egg Safety; Proposed Rule for Prevention of Salmonella Enteritidis in Shell Eggs During Production; Public Meetings. USAHA is a 108 year-old science based, dues supported, voluntary national organization of state and federal animal health agencies and other governmental departments, animal agriculture industries, university animal scientists, and veterinary laboratory diagnosticians that addresses issues of food safety, animal health and disease control, homeland security, animal welfare and public health. USAHA serves as a clearinghouse for new information and methods that may be incorporated into laws, regulations, policy and programs. It acts to develop solutions to animal health and food safety issues based on science, new information and methods, public policy, risk/benefit analysis and the ability to develop consensus for changing law, regulations, policies and programs. Devising a national program to reduce Salmonella enteritidis (SE) human infections using on-farm methods of reducing infection of egg layers is a very difficult task. Variations exist among the different regions and states regarding SE positivity rate, egg industry infrastructure, etc. Many egg quality assurance programs have developed out of necessity to maintain egg markets and have been successful. As many regional differences exist in management, housing types, training levels, etc., supervision of the program should be state or regionally based. To be effective in this endeavor, the program must not place adverse economic hardship on the producer yet obtain a significant reduction in SE human infections. It is therefore important to structure the program as a risk reduction program and not an eradication program. Partnerships between state organizations such as departments of agriculture or poultry associations and FDA should be established to carry out the program similar to how the National Poultry Improvement Plan (NPIP) program operates between the United States Department of Agriculture (USDA) and the state organizations. These organizations are knowledgeable about the industry and are usually best able to coordinate laboratory activities, training, and personnel to carry out various aspects of the program. NPIP has a successful track record in the eradication of egg transmitted SE in the egg layer breeder industry. The basis of an SE risk reduction program should be results (or process) based and not regulation based meaning that if an egg producer shows that his flocks are low risk flocks, no regulatory action is taken. Components of successful SE risk reduction programs include both steps to reduce exposure of the layers to SE and building immunity to SE in case of exposure. To reduce exposure to SE, cleaning and disinfection of SE contaminated houses, preventing introduction of SE from outside sources, and effectively controlling rodent populations have been emphasized. Vaccination has been a very valuable tool for producers with contaminated facilities to reduce infection with SE as producers have not been able to eliminate SE exposure once they are contaminated. Use of vaccine, probiotics, cleaning and disinfection between flocks, and increased efforts in controlling rodents over several successive flocks has allowed a gradual reduction of SE contamination of facilities to near non-detectable levels in many industry and state egg quality assurance programs (EQAPs); i.e. Ohio and California now find only 2 % positive environments and Pennsylvania data show only 5.1% SE manure positive flocks in 2004 compared to 38% when the programs were first started. The exemption of flocks with 3,000 layers or less from the program is seen as a weakness in the proposed program. The smaller flocks of layers are felt to more likely be exposed to SE as these flock are housed in open housing, have increased exposure to rodents and wildlife, plus spread among the flock mates is much more likely. Specific comments submitted by USAHA follow. The topic areas of these specific comments are separated into ten sections. The ten sections are: I. Pest Control II. Cleaning and Disinfection Between Flocks III. Use of Vaccines IV. Funding Issues Related to Manure Testing, Egg Testing, and the Negative Impact of Egg Diversion and Cleaning and Disinfection of Houses. V. Training of Personnel VI. Laboratory Issues VII. Model Food Code VIII. Biosecurity IX. Refrigeration Requirements X. Diversion of Eggs to Pasteurization Issues I. PEST CONTROL Rodent control is a major issue in SE control. Important issues include: 1) Use of trained rodent control personnel familiar with specific poultry housing, management and other environmental issues or personnel working under the supervisor of such knowledgeable personnel. 2) Verification of effectiveness. A part of the effectiveness evaluation could be the use the rodent index using Tin-Cats as in the Pennsylvania EQAP. 3) Continuous program evaluation. 4) Records to verify actions/procedures. 5) A certain level of flexibility in the control program should be allowed depending on such factors for instance as house construction that may markedly reduce rodent access. Fly control is NOT a major issue: 1) Although there undoubtedly can be some mechanical transfer of SE by flies, flies have not been considered a major factor in SE control. 2) Flies may be more important as a carrier of SE in areas with a high density of egg layers. 3) The possibility of a hen eating a contaminated fly is believed to be extremely less likely than a hen consuming a mouse dropping in the feed trough. 4) Fly control is complex and can vary markedly from one season to another and from one flock to another. 5) Efforts to reduce flies should probably best be handled as a recommendation rather than defining a specific mandatory program. II. CLEANING AND DISINFECTION BETWEEN FLOCKS: Between-flock cleaning and disinfection (C&D) of houses that contained manure positive flocks has been a part of most state and industry SE reduction programs in the United States. This between-flock period of time while the house is vacated of birds gives the opportunity to reduce the number of SE bacteria in the house and also to reduce rodent populations. Egg layer production facilities are not easily decontaminated hence complete sterilization of the facility is not the goal, only reduction. Historically, removing organic material using water has been used successfully to control viral diseases such as Marek’s Disease. It has been determined that wet washing is not necessary for desired results of SE negative manure tested flocks the next cycle. In fact, there is evidence that the addition of water to the layer house environment may actually increase the numbers of bacteria. In addition, wet washing is costly, leads to greater equipment damage due to corrosion, waste water causes disposal issues in many cases, houses in cold climates are very difficult to wet wash as freezing may occur. The Pennsylvania EQAP has allowed between-flock C&D without wet washing since early in 2003 (Pennsylvania EQAP Operations Annex section IV). Houses must qualify for this program however with a relatively low percent of positive manure samples (25% or less) and a low rodent index the last 3 months of lay (2 or less). The producer agrees to 1) vaccinate the incoming pullets using an approved program with bacterin, live vaccines, or both, 2) completely remove all organic material using physical means, 3) fumigate with formalin after dry cleaning, and 4) allow a minimum of a 10-day down period between flocks. In determining if dry cleaning without wet washing was a feasible option, 7 houses in which wet washing C&D was performed in houses with manure positive layers, 100% of the next flocks were tested manure positive. These same SE contaminated houses were then allowed to use the Dry Clean Program (dry cleaned, fumigated with formalin, and placement of SE vaccinated pullets). Only 2 of these 7 flocks tested manure positive at either 30 or 45 weeks of age following the use of the Dry Clean Program. The use of vaccine is partly, if not largely, responsible for the success of this program but is one method of increasing vaccine use by allowing a trade-off for not having to use wet washing. A major effort in reducing rodents during the between flock period is stressed heavily. At this time with a lack of feed and organic material, the rodents are much more easily baited plus harborage and entry sites can more easily be filled with rodent-proof materials. The local/state SE Program authority, with the on-farm SE program coordinator, should coordinate the entire process of between flock C&D planning, execution, and evaluation of results. III. USE OF VACCINES: Killed vaccines for bacterial diseases have been used for many years. As an example, much success has come from the use of commercial vaccines for Pasteurella multocida in turkeys. In addition, Erysipelothrix bacterins for hogs and turkeys and Salmonella cholerasuis vaccines for hogs have been widely used. Salmonella vaccines have been used routinely in turkey breeder programs to prevent specific Salmonella infections in poults. With the advent of SE in layers it is only natural that killed vaccines should be attempted. The development of the live vaccines by recombinant gene research has been a tremendous benefit. One of the problems with killed vaccines is the loss in body weight due to the bacterial toxins. Two injections of a killed bacterial vaccine will result in a .3 to .5 lb loss in body weight before 16 weeks of age. The advantage of the live recombinants is the oral or spray application and no injection. Since the vaccine can be used in young birds the immune system is prepared for the killed vaccine or another stimulation by the live vaccine booster. Another live vaccine developed for the European and South American conditions has been the 9R Salmonella gallinarum vaccine used in breeders conditions where challenge with Salmonella gallinarum has been very severe. This vaccine which is also a group D vaccine has been very successful as a live vaccine without the spread of the original organism. USAHA has in the past opposed the use of 9R vaccine due to concerns about safety as the U.S. has eradicated Salmonella gallinarum. Technical proof of efficacy There are many researchers who have studied the use of vaccines in layers. The Salmonella vaccine section of Diseases of Poultry, by Dr. Richard Gast summarizes published research results on this topic. The numbered references (in brackets) for the information in this section of his book are found at the end of this section. “The degree of protection afforded by Salmonella vaccines (including cross-protection by vaccines of serotypes heterologous to the challenge strain) has generally been good, although rarely complete. Such protection rarely approaches 100% efficacy and can usually be overcome by high challenge doses. However, even partial protection can be very useful toward achieving the long-term goal of reducing SE incidence in successive laying flocks in a house or farm. Data from the European programs that use vaccines certainly demonstrate that vaccination can contribute to SE control in a very positive manner. In my opinion, one of the real weaknesses of the "test and divert" emphasis of the FDA proposal is that it only address the public health risk of currently infected flocks, and has minimal value for preventing recurrence of infection in subsequent flocks on the same site. Vaccination is a powerful tool that should be utilized (in addition to cleaning and disinfection) for this purpose. Identifying infected flocks might protect consumers against an immediate disease threat, but true long-term protection of public health will require additional efforts to break the cycle of continuous re-infection of flocks from environmental reservoirs.” “Vaccination with either killed or live preparations has been found to reduce the susceptibility of poultry to paratyphoid Salmonella infection. Live Salmonella vaccines have often been associated with a stronger or longer-lasting protective response in poultry, perhaps either because of adverse effects on relevant protective antigens during the preparation of killed vaccines or because live vaccines present relevant antigens to the host immune system more persistently (23). Killed vaccines may also fail to fully elicit the cell-mediated portion of the protective response (343). Nevertheless, both killed and live vaccines have been associated with significant protection against Salmonellae, although neither type of vaccine has consistently provided an impenetrable barrier against infection. Moreover, feed or water deprivation and environmental stresses such as heat may compromise the effectiveness of vaccines (346). Like competitive exclusion, vaccination is most effectively used as a component in a comprehensive program of risk reduction practices.” “Interest in the use of killed vaccines (bacterins) in poultry has been renewed in recent years by escalating concerns about S. enteritidis. Subcutaneous administration of adjuvanted oil-emulsion bacterins to laying hens has been reported to reduce significantly the incidence of fecal shedding and the numbers of S. enteritidis shed in the feces, the frequency of isolation of S. enteritidis from various internal tissues, and the incidence of production of eggs with contaminated contents following subsequent oral challenge (173, 174). Chickens vaccinated with bacterins have been reported to exhibit reductions in mortality, lesions, clinical signs, and organ invasion for up to 12 wk post vaccination when challenged with S. enteritidis by intravenous or intramuscular routes (441, 442). Field studies have associated bacterin administration with a reduced incidence of S. enteritidis infection in Dutch broiler breeder flocks (145), but could not document any significant effect of vaccination on the isolation of S. enteritidis from the environments of Pennsylvania laying flocks (122). Subunit vaccines, composed of Salmonella outer-membrane proteins administered with adjuvants or incorporated into lipid-conjugated immunostimulating complexes, have been efficacious against S. enteritidis in chickens (332) and against S. enteritidis or S. heidelberg in turkeys (62, 63). Another potential benefit from vaccination was evident in a study that reported reduced multiplication of S. enteritidis in eggs from bacterin-treated hens compared to eggs from control hens (238).” “Live attenuated vaccines need to persist in tissues long enough to induce a protective immune response, but should be avirulent and eventually cleared from vaccinated birds. PT Salmonella vaccine strains attenuated by several different approaches have been tested for their protective efficacy in poultry. Oral or intramuscular administration of various aroA mutants of S. enteritidis (auxotrophs that do not grow well in vivo because of their inability to synthesize particular aromatic compounds) reduced organ invasion after intravenous or aerosol challenge and reduced fecal shedding, horizontal transmission, and egg contamination after oral challenge (27, 76, 77, 78). This protection has been found to persist for up to 23 wk after administration of the vaccine strain (79). An orally administered cya crp S. typhimurium strain (a double mutant with deletions of both adenylate cyclase and the cyclic AMP receptor protein) provided very strong protection against intestinal colonization and organ invasion by S. typhimurium (204). A temperature-sensitive mutant (59) and a strain attenuated by repeated passage in chicken heterophils (282) have also been shown to protect chickens against S. enteritidis infection. Hassan and Curtiss (205) reported that vaccination of hens with an avirulent S. typhimurium strain reduced intestinal colonization of their progeny when challenged with virulent wild-type strains. Evidence for cross-protection by live vaccine strains against other epidemiologically important Salmonella serotypes has been inconsistent. An avirulent S. typhimurium vaccine reduced colonization, organ invasion, and egg contamination by S. enteritidis (206), but aroA- S. enteritidis strains did not cross-protect very effectively against S. typhimurium challenge (78). Several safety concerns about live Salmonella vaccines have been raised recently, based on evidence that some vaccine strains may be genetically unstable (19) and can be detected for longer than anticipated in vaccinated hens if sufficiently sensitive culturing methods are used (428).” References for the above section – Diseases of Poultry by Dr. Richard Gast 19. Barbezange, C., G. Ermel, C. Ragimbeau, F. Humbert, and G. Salvat. 2000. Some safety aspects of Salmonela vaccines for poultry: in vivo study of the genetic stability of three Salmonella typhimurium live vaccines. FEMS Microbiol Lett 192:101-106. 23. Barrow, P.A., J. Hassan, and A. Berchieri, Jr. 1990. Reduction in faecal excretion of Salmonella typhimurium strain F98 in chickens vaccinated with live and killed S. typhimurium organisms. Epidemiol Infect 104:413-426. 24. Barrow, P.A., M.B. Huggins, and M.A. Lovell. 1994. Host specificity of Salmonella infection in chickens and mice is expressed in vivo primarily at the level of the reticuloendothelial system. Infect Immun 62:4602-4610. 27. Barrow, P.A., M.A. Lovell, and A. Berchieri. 1991. The use of two live attenuated vaccines to immunize egg-laying hens against Salmonella enteritidis phage type 4. Avian Pathol 20:681-692. 59. Cerquetti, M.C. and M.M. Gherardi. 2000. Orally administered attenuated Salmonella enteritidis reduces chicken cecal carriage of virulent Salmonella organisms. Vet Microbiol 76:185-192. 62. Charles, S.D., K.V. Nagaraja, and V. Sivanandan. 1993. A lipid-conjugated immunostimulating complex subunit vaccine against Salmonella infection in turkeys. Avian Dis 37:477-484. 63. Charles, S.D., I. Hussain, C.-U. Choi, K.V. Nagaraja, and V. Sivanandan. 1994. Adjuvanted subunit vaccines for the control of Salmonella enteritidis infection in turkeys. Am J Vet Res 55:636-642. 76. Cooper, G.L., L.M. Venables, and M.S. Lever. 1996. Airborne challenge of chickens vaccinated orally with the genetically-defined Salmonella enteritidis aroA strain CVL30. Vet Rec 139:447-448. 77. Cooper, G.L., L.M. Venables, R.A.J. Nicholas, G.A. Cullen, and C.E. Hormaeche. 1992. Vaccination of chickens with chicken-derived Salmonella enteritidis phage type 4 aro A live oral Salmonella vaccines. Vaccine 10:247-254. 78. Cooper, G.L., L.M. Venables, R.A. J. Nicholas, G.A. Cullen, and C.E. Hormaeche. 1993. Further studies of the application of live Salmonella enteritidis aroA vaccines in chickens. Vet Rec 133:31-36. 79. Cooper, G.L., L.M. Venables, M.J. Woodward, and C.E. Hormaeche. 1994. Vaccination of chickens with strain CVL30, a genetically defined Salmonella enteritidis aroA live oral vaccine candidate. Infect Immun 62:4747-4754. 122. Davison, S., C.E. Benson, D.J. Henzler, and R.J. Eckroade. 1999. Field observations with Salmonella enteritidis bacterins. Avian Dis 43:664-669. 145. Feberwee, A., T.S. de Vries, A.R.W. Elbers, and W.A. de Jong. 2000. Results of a Salmonella enteritidis vaccination field trial in broiler-breeder flocks in the Netherlands. Avian Dis 44:249-255. 173. Gast, R.K., H.D. Stone, P.S. Holt, and C.W. Beard. 1992. Evaluation of the efficacy of an oil-emulsion bacterin for protecting chickens against Salmonella enteritidis. Avian Dis 36:992-999. 174. Gast, R.K., H.D. Stone, and P.S. Holt. 1993. Evaluation of the efficacy of oil-emulsion bacterins for reducing fecal shedding of Salmonella enteritidis by laying hens. Avian Dis 37:1085-1091. 204. Hassan, J.O., and R. Curtiss III. 1994. Development and evaluation of an experimental vaccination program using a live avirulent Salmonella typhimurium strain to protect immunized chickens against challenge with homologous and heterologous Salmonella serotypes. Infect Immun 62:5519-5527. 205. Hassan, J.O. and R. Curtiss III. 1996. Effect of vaccination of hens with an avirulent strain of Salmonella typhimurium on immunity of progeny challenged with wild-type Salmonella strains. Infect Immun 64:938-944. 206. Hassan, J.O. and R. Curtiss III. 1997. Efficacy of a live avirulent Salmonella typhimurium vaccine in preventing colonization and invasion of laying hens by Salmonella typhimurium and Salmonella enteritidis. Avian Dis 41:783-791. 238. Holt, P.S., H.D. Stone, R.K. Gast, and R.E. Porter, Jr. 1996. Growth of Salmonella enteritidis (SE) in egg contents from hens vaccinated with an SE bacterin. Food Microbiol 13:417-426. 282. Kramer, T. T. 1998. Effects of heterophil adaptation on Salmonella enteritidis fecal shedding and egg contamination. Avian Dis 42:6-13. 332. Meenakshi, M., C.S. Bakshi, G. Butchaiah, M.P. Bansal, M.Z. Siddiqui, and V.P. Singh. 1999. Adjuvanted outer membrane protein vaccine protects poultry against infection with Salmonella enteritidis. Vet Res Communic. 23:81-90. 343. Muotiala, A., M. Hovi, and P.H. Makela. 1989. Protective immunity in mouse salmonellosis: Comparison of smooth and rough live and killed vaccines. Microb Pathog 6:51-60. 346. Nakamura, M., N. Nagamine, T. Takahashi, S. Suzuki, and S. Sato. 1994. Evaluation of the efficacy of a bacterin against Salmonella enteritidis infection and the effect of stress after vaccination. Avian Dis 38:717-724. 428. Tan, S., C.L. Gyles, and B.N. Wilkie. 1997. Evaluation of an aroA mutant Salmonella typhimurium vaccine in chickens using modified semisolid Rappaport Vassiliadis medium to monitor faecal shedding. Vet Microbiol 54:247-254. 441. Timms, L.M., R.N. Marshall, and M.F. Breslin. 1990. Laboratory assessment of protection given by an experimental Salmonella enteritidis PT4 inactivated, adjuvant vaccine. Vet Rec 127:611-614. 442. Timms, L.M., R.N. Marshall, and M.F. Breslin. 1994. Laboratory and field trial assessment of protection given by a Salmonella enteritidis PT4 inactivated, adjuvant vaccine. Br Vet J 150:93-102.” USAHA is a strong proponent of vaccines for Salmonella. At the present time, we have the emergence of antibiotic resistant strains of Salmonella ex. DT104, of Salmonella typhimurium, SE PT4, and penta-resistant strains of other Salmonella. The poultry industry is constantly under the threat of losing the availability of antibiotics. Available tools are decreasing. We have to look for alternate ways, which are successful in reducing the risk of spreading Salmonella. Vaccines are one of the effective tools we have today both in human and animal industry. If Salmonella is cycling in a breeder flock the best way to stop cycling of Salmonella in that flock is through vaccination. This has been demonstrated in many experimental trials both in turkeys and chickens. Both killed and live vaccines have their own inherent advantages and disadvantages. Nevertheless, the use of vaccines will play a significant role in reducing Salmonella in poultry. Birds vaccinated will be less susceptible for infection with Salmonella and shed less into the environment. In addition to the above comments USDA approved vaccine efficacy studies by several commercial vaccines companies are available through USDA. These studies represent many years of research, which have resulted in the granting of many licenses to manufacture and distribute under a USDA approval based on efficacy and safety in the United States. Experiences in use of vaccine in the field Inactivated vaccines have been a very effective means of reducing environmental contamination when used in combination with rodent control, sanitation and housing of SE clean replacements. The use of vaccines is one of the most prudent (effective, practical) SE risk measures to be taken for multiple in-line layer complexes. There is overwhelming information available from abroad and the U.S. on its effectiveness and limitations. Vaccines have been consistently very successful in reducing detectable environmental contamination in our experience. The verdict on live vaccines is still out. I agree with the FDA proposal that vaccinations should not be required, but left to local veterinary/company decision makers. It has been shown that the control of SE cannot be done without vaccine use in conjunction with rodent control, dry cleaning, disinfection, and biosecurity measures. Killed and live Salmonella vaccines have been used to: 1.. Eliminate an SE break in a large complex by testing, removing a flock, dry cleaning a house, and vaccinating the new flock. The 30 plus house complex is now negative and has been for several years. 2.. Vaccinate all flocks going into a complex resulting in a conversion of many houses from positive to negative. 3.. Revaccinate positive molted flocks with low incidence SE, which converted to negative and maintained their negative status until the end of lay. 4.. Spot vaccinate resulting in a lowering of incidence to negative. 5.. Vaccinate pullets during grow, move into a positive layer house in a complex and never have a positive SE isolation after 2 more flocks in the same house. 6.. Vaccinate a positive pullet flock in a pullet complex and with cleaning, isolation and disinfection not have another occurrence in other houses or in the same house the next grow period or on the complex again. The Pennsylvania EQAP has already reported the results of several years of vaccination. The data associated with that program was reported in the Maryland SE open meetings held by FDA. International programs and benefits The international community has had many experiences with Salmonella enteritidis PT4. Only in the past few years has England been able to reduce their problems by requiring mandatory vaccination of birds where eggs are being sent to the market for consumer use. In Germany where just a few years ago the incidence of Salmonella was approaching epidemic proportions until a vaccine was required. Currently, as reported by the Euro- Surveillance Monthly data from 15 countries in Europe report the incidence of PT4 has declined from 61.8% in 1998 to 32.1% in 2003. The European Food Safety Authority (EFSA) has issued an opinion on the use of vaccines for the control of Salmonella in poultry in October 2004 that contains supportive information regarding vaccination to prevent SE in eggs. This document can be accessed at the EFSA website at: http://www.efsa.eu.int/science/biohaz/biohaz_opinions/721_en.html. Summary of Section III comments – Use of Vaccines The evidence for the efficacy of the Salmonella vaccines although not perfect is very good under the most challenging of research and field circumstances. When either live or killed vaccines are used with appropriate programming, and associated with bio-security measures, testing, and rodent control the result will be a reduction of SE. An SE program is always a work in progress and not a course to elimination. IV. FUNDING ISSUES RELATED TO MANURE TESTING, EGG TESTING, AND NEGATIVE IMPACT OF EGG DIVERSION, CLEANING /DISINFECTION: It is doubtful that anyone can come even close to accurately predicting what the economic impact of such a national regulatory measure will be. It will be fairly easy to come up with an estimate of manure testing costs once the laboratories can furnish costs estimates of their needed expansions to meet the increased testing need. However, the costs related to egg testing will depend upon the manure testing results. A lot of positive environmental tests will, of necessity, increase the number of required egg tests. The industry costs related to the diversion of positive eggs will depend upon the number of positive egg test results and the availability of breakers that will purchase those eggs at a fair price. Most doubt that there will be either the breaking capacity or the market capacity to accept the increased amount of product from diverted eggs. Without the sale of diverted eggs at a fair price, hens will have to be depopulated. It is not unusual for hens to be the collateral securing bank loans for their purchase so flock destruction could have very serious consequences to some producers. It seems clear that under the proposed regulation, whether or not an egg producing company can financially survive will depend upon the SE status of their flocks and whether or not they have their own breaking/pasteurization capability. Without some sort of compensation program to offset inevitable diversion losses for those that lack egg breaking/pasteurization capability, there will likely be an increased number of bankrupt egg producers. The primary uncertainties that limit the usefulness of a discussion on the need for indemnification to cover testing, egg diversion and cleaning /disinfection costs depends upon the extent of environmentally positive houses and then the extent of SE positive eggs requiring diversion (which will not likely be a viable marketing option for many producers). The proposed program is presented as being for the "public good". There is the common belief that such programs should be funded by those who the program is designed to benefit: the public. Compensation can be through increased egg prices or through the government from public tax funds. One thing seems clear: without a testing compensation plan and an indemnification program for diverted egg losses or for losses due to layer depopulation because of a lack of breaker capacity or adequate liquid egg markets, many egg producers could be put out of business. We recommend that FDA contract with each State Department of Agriculture that has a credible Egg Quality Assurance Program for oversight of this program. States are in the best position to administer the program. There must be some flexibility to address unique state and regional production/management systems. Adequate funding should be provided for state cooperative agreements. V. TRAINING OF PERSONNEL: D. official quality control supervisor The following recommendations are based on experience with existing proven Egg Quality Assurance Programs (EQAP’s): 1. We recommend to FDA that the person responsible for overseeing SE prevention measures be trained in the manner adopted by EQAP’s using process control principles. The following are administrative requirements of the California EQAP: Administrative a.. Develop a farm/premises flock egg quality assurance plan. b.. Designate an employee or employees as the official quality control supervisor(s) for in-house operations and for follow-up training. 2. The official quality control supervisor responsible for overseeing SE prevention measures should receive comprehensive training in the following areas: Preparing a Quality Assurance Plan, Egg Handling, Flock Health Management, Cleaning, Disinfection and Biosecurity, Vector Control and Biosecurity, and Environmental Monitoring and Sampling. The training material for these topic areas from the California EQAP is available at: http://animalscience.ucdavis.edu/avian/train.html. 3. Each Company should designate a person responsible for overseeing the core process control elements of a comprehensive SE reduction program. This person received training from qualified poultry professionals from academia, industry and state agencies. 4. Each State should develop regionally adapted training programs based on industry and marketing structure, with particular consideration for site-specific specific management and housing systems used. 5. Special training provisions must be made for States without a pre-existing EQAP and for small producers that cannot easily get away to attend training. It is recommended that FDA provide qualified poultry professionals and adequate resources from States with a proven existing plan to assist in developing new training pertinent to that State. In some remote areas, it may be necessary to provide a home study curriculum for core components and continuing education credits in order to make training available to all producers and to provide cost effective training across the United States. 6. Training should include pre-test and post-test (with an appropriate passing score) assessment of the trainees’ performance in order to assess competence and quality of training materials. 7. There should be required annual continuing education for all official quality control supervisors by qualified poultry professionals. 8. The official quality control supervisor will develop and review the basic premises quality assurance plan at least once annually or when changing circumstances dictate. 9. That FDA acknowledges the importance of official quality control supervisor by granting an official certificate or qualification upon completion of training and that annual re-certification be contingent upon completion of mandatory continuing education. A passing grade of 90% must be achieved in each module (see Appendix A). 10. That FDA audit training records from each State to assure equivalence in the degree of training required but that it allow for flexibility in the designing the specific program elements. D. state agency representatives 1. That all State agency representatives overseeing SE prevention measures be required to receive the same training required of the official quality control supervisor. 2.. That all State agency representatives receive training necessary to conduct uniformly administered audits of egg production facilities. 3.. That all State and Federal agency representatives receive standardized training on conducting trace back investigations through a formal cooperative agreement among agencies. D. laboratory personnel 1. That all laboratory personnel conducting Salmonella testing receive standardized training through a formal cooperative agreement among agencies. D. sample collectors 1.. Who will be approved to collect samples is yet to be resolved by FDA. Persons collecting samples must be trained in proper handling and sampling techniques using scientifically valid principles including randomization. 2.. In order for the regulatory program to be credible, chain of custody issues must also be addressed and implemented. VI. LABORATORY ISSUES: A. Environmental testing The FDA proposal states, “you must conduct environmental testing for SE as an indicator of whether your SE prevention measures are working effectively”. We agree that the best screening method to determine if a flock is positive for SE is environmental testing. However, finding SE in the environment of a poultry house does not necessarily indicate that the birds are colonized or infected with SE nor does it indicate that eggs may be contaminated with SE. 1. Testing times The FDA proposal specifies testing a house when “each group of laying hens in the flock in that house are 40-45 weeks of age and, if molted, approximately 20 weeks after the end of any molting process”. Most of EQAP’s specify testing the environments of layer houses at the end of the laying period prior to depopulation. The purpose of the test is to determine if SE has contaminated the environment of the house. If the house environment is found to be positive, then the subsequent flock to be housed is at risk of being colonized with SE. Therefore the company should reevaluate their control program and may choose to intensify their routine cleaning and disinfections of the house, increase their rodent control program, or vaccinate the incoming flock. The Ohio EQAP tests layer house environments from 2 to 10 weeks prior to depopulation. Using this testing scheme, the Ohio EQAP has shown a reduction in the percentage of SE positive flocks from 20% in 1997 to 2.1% in 2003. The California EQAP tests in a similar time frame and has had success in reducing both environmental positives and human disease. Since 2000, there have been no outbreaks of SE associated with consumption of California eggs. In 1997, 12 SE egg-related food borne disease outbreaks occurred in California and since the year 2000, no egg related SE outbreaks have been linked to California eggs. Human SE has declined in California from 3.60 cases per 100,000 population in 1993 to 1.76 cases per 100,000 population in 2003, representing a 51% decrease in the per capita human incidence. The percentage of California outbreak cases due to SE was at a maximum of 21.84% in 1996 and was recently at 3.4% of outbreak cases in 2003, representing an 84.4% decline in cases. It is evident that with existing programs and epidemiological trends, the need for further action through regulation is debatable from a cost: benefit perspective. Thus the objectives of Healthy People 2010 have been exceeded in California and perhaps in other States at the present time. Better allocation of funds can be achieved through risk-based regional prevention and control efforts. The prevalence of SE positive environmental samples from California layer ranches declined generally over time from 9.78% (95% CI: 4.85, 18.21) in 1991 to 2.0% (95% CI: 1.17, 3.38) in 2004, indicating a statistically significant change over time. It is likely that the CEQAP and other egg quality assurance programs have played a contributing role in declining human illness due to SE in eggs, although it is important to note that many other factors are involved. We are not aware of any data that indicates that 40-45 wks of age is the best time to monitor flocks. The reference cited in the proposed rule is a memo from Richard Wood of Food Animal Concerns Trust to FDA. Programs that require testing at the end of lay have proven efficacious in reducing both environmental positives and human disease. We recommend that the time for the environmental testing of a layer house be sometime at the end of lay. The testing should be done early enough to allow the company to respond accordingly if SE is isolated. This would also eliminate the costly and problematic requirements for egg testing and diversion. 2. Sampling methods The FDA proposal recognizes the data concerning manure sampling versus egg machinery sampling and concludes “tentatively … that environmental testing of the manure or (bold and italics added) egg machinery in a poultry house is an appropriate method for screening the environment for SE and should be used as one indicator of the effectiveness of your SE prevention measures”. FDA further states that for verification of an on-farm egg program that the manure is the preferred sample type. A drag swab should be dragged the entire length of one side of the row/bank. Another drag swab should be dragged along the entire length of the other side of the row/bank. These should be placed into separate sample bags. a. Types of samples Developing an equitable program for environmentally sampling layer houses is a challenge because of the vast number of styles or types of layer houses. Considering only different types of manure collection/disposal systems, these may include high-rise deep pit, shallow pit, manure belt, shallow pit flush, and cage-free floor systems. These may vary greatly in a given geographic area and across geographic regions. Because of the difference in the manure collection/disposal systems, it is difficult to test these various houses on an equivalent basis. Many of the EQAP’s specify the manure-type sample as the sample of choice. However, there are others that prefer the egg machinery-type sample (e.g. UEP 5-Star) or a combination of the two. The Pennsylvania SE Pilot Project (SEPP, SEPP Progress Report, 1995) reported that 15% of the manure drag swabs, 14% of the manure scraper swabs, 17% of the egg machinery samples, 12% of fan swabs, and 18% of walkway drag swabs were positive for SE. Schlosser et al. (1999), reporting on the data from the SEPP, indicated that egg machinery and manure drag swabs together are better predictors of the potential of a flock to produce positive eggs than is either sample type alone. However, the Pennsylvania EQAP recommends the collection of manure samples only, unless these samples cannot be obtained safely or because of house design. The USDA National Animal Health Monitoring System Layer ‘99 study found that 48% of egg belts, 45% of elevators, 18% of walkways, and 17% of manure were positive for SE. However egg roll-outs may not be as sensitive as egg belts. In a study presented at the 1998 USAHA Salmonella Workshop, various sample types and isolation protocols for detecting Salmonella from layer houses were compared. The study did not detect any SE, however, there was a high prevalence of other Salmonella serotypes present. It was found that 60% -77% of manure samples were positive (depending on type of manure sample), 67% of walkway drag swabs were positive, and about 50% of egg machinery samples were positive for Salmonella. This study involved several different housing types, with the only common environmental sample being walkway drag swabs. In a study of layer houses in California, it was concluded that the manure swabs were better than egg belts/roll-outs samples. The Ohio EQAP prefers the manure sample. They believe that the manure sample is directly related to the chicken and has proven to identify SE far better than egg machinery testing. The Maine EQAP specifies collecting egg machinery, manure, and walkway samples. This is based on the finding that 5.3% of manure samples and 6.1% of egg conveyors were SE positive and often only one or the other sample type is positive. Egg conveyors may be more of an indicator for contamination from rodents and dust in the house and manure may be an indicator of mice, dust, and birds shedding SE. Because of the variations in housing types and management systems it is difficult to specify a single sampling procedure. The California EQAP recommends that the types of samples that are collected should depend on the type of house that is to be sampled. For example, a hand gathered house in California that does not have egg belts or conveyors, only egg roll-outs, would probably only be sampled by manure drag swabs because egg roll outs have not shown to be an effective source for SE. However, a deep pit high-rise house in Georgia that has easy access to both the manure pits and egg machinery may include both manure and egg machinery samples. b. Collection of samples Individuals representing the layer company, state or laboratory personnel, or accredited veterinarians qualify for collecting samples provided they have been trained in the proper methods for collecting and transporting the samples. FDA or state authorities should be responsible for training individuals. c. Number of samples The number of samples collected needs to be practical and take into consideration the cost of testing, while adequately assessing the presence of SE in the house. The various programs recommend different numbers of samples depending on the size and type of house. The Pennsylvania EQAP recommends collecting 2 manure drag swabs per row/bank. The Ohio EQAP requires 2 drag swabs per row and then pools the 2 swabs from a row. The California EQAP recommends collecting a standard number of 16 manure swabs from each house regardless of the house type and to pool these samples down to 4 samples. The 16 swabs were calculated using a binomial distribution model assuming 10% of the drag swab area was contaminated with SE. Based on this assumption, the use of 16 swabs gives an 81% certainty of detecting SE with a confidence level of 95%. California has compared the isolation of SE from layer houses using 16 individual manure swabs versus 4 pooled manure swabs and found no significant differences. The use of pooling samples has the advantage of reducing the number of samples to be tested. Because of the large variation in the size and types of layer houses, we recommend using a standard number of samples per house as opposed to a number dependent on rows or some other parameter. The California EQAP has developed a system based on a standard number of 16 swabs per house that has been shown to be effective. Therefore we recommend, as a minimum requirement, that a standard number of 16 swabs be collected from each housing environment. The types of samples to be collected will depend on the particular type of layer house. For example, all 16 samples may come from manure or there may be 8 samples from manure and 8 samples from the egg machinery. These 16 samples may be pooled into 4 samples provided that each pool is made up of the same type of sample (i.e. all manure or egg machinery and not a mixture of manure and egg machinery swabs). In large houses, the areas to be swabbed should be randomized to be representative of the entire house. d. SE outbreak trace back testing The FDA proposal makes a distinction between the sampling plan for verifying an on-farm egg program and the sampling plan for an outbreak trace back. In addition to the manure drag swabs recommended for the on-farm program, sampling of the egg machinery, fans, and walkways would be a part of a trace back test. There should also be a distinction made between the level of testing necessary for an on-farm monitoring program and for a trace back evaluation. 3.. Culture methods The FDA proposes that “you must test for SE in environmental samples according to the method “Detection of Salmonella in Environmental Samples from Poultry Houses”. FDA states that these methods are required unless you test for SE in environmental samples using other methods that are at least equivalent in accuracy, precision, and sensitivity in detecting SE. The FDA proposal suggests using a pre-enrichment (BPW) followed by a dual selective enrichment (TT and RV) method for culturing environmental samples. The samples are pre-enriched in BPW at 35 C for 24 hours. One ml of the pre-enriched sample is inoculated into 10 ml of TT enrichment broth and 0.1 ml inoculated into 10 ml of RV enrichment broth. Both enrichment broths are incubated at 42 C for 24 hours. Each enrichment broth is plated onto BS, BGN, and XLT4 media. After 24 hours incubation (BS is 24 and 48 hours), 5 suspect colonies are picked to TSI and LIA for screening. The referenced FDA procedure does not offer any supporting data or reference any scientific reports in deriving the recommended procedures. The NPIP has adopted two separate protocols for the isolation of Salmonella from environmental samples. The first protocol involves direct enrichment of the sample into Tetrathionate (TT) enrichment broth followed by the delayed secondary enrichment (DSE) procedure. The second, more recently adopted, procedure involves pre-enriching the sample in buffered peptone water (BPW) followed by selectively enriching into both TT and Rappaport-Vassiliadis (RV) enrichment broths. After enrichment, the broths are inoculated onto highly selective media; generally brilliant green agar supplemented with novobiocin (BGN) and xylose lysine tergitol 4 (XLT4) agar. After incubation, suspect colonies are screened using triple sugar iron (TSI) agar and lysine iron (LI) agar. The FDA proposed method is similar to the second NPIP method. The NPIP methodology was developed primarily for the detection of Salmonella from the environments of breeder houses. However, this methodology has been shown to be effective with other types of environments, including layer houses. There are some important aspects of layer houses that make the isolation of Salmonella, especially SE, more difficult. The environmental samples of layer houses, especially manure samples, have a much higher level of background bacteria, they have a greater percentage of Salmonella positive samples, and they typically have multiple Salmonella serotypes present. In a study reported at the 1998 USAHA Salmonella Workshop, several isolation protocols for detecting Salmonella from environmental samples from layer houses were compared. The methods included, direct enrichment, pre-enrichment followed by selective enrichment, and delayed secondary enrichment. Direct enrichment varied from 54% to 65% Salmonella positive, depending on the enrichment used. Pre-enrichment followed by selective enrichment varied from 48% to 66%, depending on the enrichments used. Delayed secondary enrichment procedures ranged from 75% to 79% based on the incubation temperature. If the results of the pre-enrichment followed by TT (63%) and pre-enrichment followed by RV (66%) were combined (essentially the FDA proposed method), 76.5% of the samples were positive compared to 83% Salmonella positive combining direct and DSE. The California Animal Health and Food Safety Laboratory System has found that the DSE procedure increases the isolation rate compared to direct enrichment and is particular important when pooling samples. In other studies, of the various culture methods tested, the best result involves direct enrichment with TT broth followed by DSE. We recommend that FDA adopt the methodology used by the NPIP for isolating Salmonella from poultry environmental samples. This would allow the use of either isolation procedures, however, we emphasize that the use of DSE would provide increased sensitivity. There is a great deal of comparative work on the use of selective plating media with environmental samples. The plating media specified by AOAC/BAM, i.e. BS, HE, and XLD, have been shown to perform very poorly with environmental samples. The primary difficulty is the level of false positives on these media, which results in a great deal of screening and a reduction in the sensitivity due to the number of false positive colonies. Of the three media, BS has the most problems. The newer, highly selective media such as the novobiocin supplemented media (BGN, HEN, XLDN), the tergitol 4 containing media (XLT4, MM), and some of the chromogenic agars have been shown to be very effective. Bismuth sulfite agar is primarily used for the isolation of the human serotype-specific Salmonellae, S. typhi. Also, it is recommended for the isolation of lactose-fermenting Salmonellae that are occasionally isolated from milk- or dairy-type samples and some reptilian samples. The presence of lactose-fermenting Salmonella has not been shown to be a common finding with poultry environmental samples. Moreover, if the Salmonella isolate produces hydrogen sulfide, it would still be isolated on the hydrogen-sulfide indicator media, even if the isolate fermented lactose. We strongly oppose the use of BS agar as unnecessary and a waste of time and money. Because of the high percentage of Salmonella that are typically present in layer house environmental samples and the presence of multiple serotypes, it is important to screen at least 5 suspect colonies from each plate. Typically, this screening begins with inoculation into TSI and LI agar slants. After incubation, if either media shows a reaction typical of Salmonella the isolate should be screened with antisera to Salmonella serogroup D1. If any of the isolates agglutinate with the D1 antisera, they should be sent to NVSL for serotyping. All confirmed S. enteritidis isolates should be phage typed. 4. Reporting All Salmonella Group D isolates must be sent to the USDA, National Veterinary Services Laboratory (NVSL) or a USDA accredited regional laboratory for identification to rule out false positive isolates. Once the NVSL confirms the Salmonella isolate is SE, they report the results to the submitting laboratory. The laboratory then notifies the State or FDA representative and the layer company official of this finding. D. Egg testing The isolation of SE from environmental samples does not necessarily indicate that the birds are producing SE contaminated eggs. In order to directly evaluate if the eggs are contaminated with SE, FDA requires egg testing. Testing times FDA proposes “that you must review implementation of your SE prevention measures and begin egg testing within 24 hours of receiving notification of the positive environmental test”. Once the layer company receives notification of having a SE positive house, the State/FDA responsible entity, layer company and laboratory should collectively determine the earliest time for collecting and testing eggs from that flock. The current FDA proposal requires eggs to be tested within 24 hours of notification. This is not practicable. For example, what if the company was notified on Friday? There is no way a laboratory is going to set up and test over the weekend. Few laboratories are routinely set up to test eggs. Laboratories will require some time to set up for this testing. Also the laboratory that routinely tests the environmental samples may not be able to do the egg testing. There needs to be some leeway in scheduling the egg testing. Sampling methods The FDA proposal requires a random collection of 1000 eggs to be tested every 2 weeks for 4 collection periods. A trained company representative, accredited veterinarian, or state or laboratory technician should be responsible for randomly collecting the 1000 eggs. Realistically, more than 1000 eggs should be collected to allow for discarding any dirty or cracked eggs. The person collecting the eggs must coordinate the collection of the eggs with the laboratory that will be doing the testing. Culture methods The FDA proposed method references an article by Valentin-Bon and involves the pooling of 20 eggs. The pooled eggs are incubated at room temperature for 4 days. From each egg pool a 25 ml portion is added to 225 ml of TSB supplemented with ferrous sulfate and mixed. The pH was adjusted to 6.8. The sample is incubated for 24 hours at 35 C. One ml of the sample is inoculated into TT enrichment broth and 0.1 ml is inoculated into RV enrichment broth. The TT broth is incubated at 35 C and the RV broth is incubated at 42 C, both for 24 hours. Both enrichment cultures are plated onto BS, BG, BGN, XLD, and XLT4 media. Presumptive colonies are picked to TSI and LIA media. They state that these methods are required unless you test for SE in eggs using other methods that are at least equivalent in accuracy, precision, and sensitivity in detecting SE. The currently accepted egg testing methodology recommends pooling either 10 or 20 eggs. The eggs are aseptically broken and the contents thoroughly mixed. They are incubated at room temperature for 3 to 5 days and then inoculated onto selective agar plates. A modification of this procedure can be used by transfering 1 ml of the egg mixture after incubation at room temperature for 5 days into TT enrichment. This procedure can detect as few a 3 CFU per 20-egg pool. The proposed method also recommends the pooling of eggs and incubating them for 4 days at room temperature, but then recommends transferring 25 ml of the egg mixture into non-selective, iron supplemented media followed by transfer into 2 selective enrichments. We do not see the usefulness of the iron-supplemented non-selective enrichment (TSB). Whole egg is one of the most nutritious media available. An article by Seo et al. (2003) that appeared in the same journal as the procedural paper by Valentin-Bon documents levels of SE in egg pools of greater than 8 logs after initial inoculation of 10 SE per 10 egg pool and incubation at 37 C for 48 hours. The egg mixture was plated without additional enrichment. Levels of Salmonella in whole egg mixtures of 6 logs or better are readily detected by direct plating. The use of selective enrichments (TT and RV) is also not necessary because the levels of background flora are very low in whole egg. The procedure recommended by FDA also inoculates both selective enrichments onto 5 selective agar plates. This would result in the use and inoculation of 500 agar plates (50 egg pools x 2 enrichments x 5 plates), which is a tremendous waste of time and money. There is no valid reason to use all of these plates. The BG and BGN are the same basic media, except BGN has been supplemented with novobiocin and proven to be more effective because of it. XLT4 and XLD have similar basic formulations, however, XLT4 has been shown to be much more effective in poultry. We have already discussed the difficulties of BS agar. Therefore we recommend the use of highly selective media such as BGN and XLT4 agar. We recommend the currently accepted egg testing procedure be included as the accepted procedure. D. Laboratory involvement Laboratory requirements The FDA proposal does not specify which laboratories will be able to conduct the testing or if there are any requirements that must be met in order to conduct such testing. We propose that laboratories must agree to follow the proposed minimal standard guidelines for isolating SE or use other procedures that have been shown to be as effective. Such laboratories would include laboratories that are authorized by the NPIP or those that submit to the NVSL check test. Cost of testing The FDA proposal states that the cost of environmentally testing the average layer house (a 6-row house and 12 samples) would be $541 (~$45/sample). This includes the cost of sample collection, shipping and lab costs. California has calculated the costs (labor and materials) for culturing samples by direct selective enrichment and by direct selective enrichment followed by DSE. He calculated the cost of testing a layer house assuming a prevalence of 10% SE and rule out of 90% non-group D samples, and found the following: Direct selective enrichment $/sample $/32 samples No Salmonella, no suspects $8.95 $ 286.40 No Salmonella, suspects to rule out $20.85 $ 667.20 Salmonella detected, not group D1 $41.55 $1196.60 Salmonella detected, Group D1 $58.60 $1384.00 Direct selective enrichment followed by DSE $/sample $/8 pools No Salmonella, no suspects $32.10 $ 256.80 No Salmonella, suspects to rule out $44.10 $ 352.80 Salmonella detected, not group D1 $64.70 $ 465.84 Salmonella detected, Group D1 $81.75 $ 531.24 Cost was found to depend on the culture result. The use of the DSE procedure allowed the use of pooled samples and thereby reduced the costs by over half. The Ohio Department of Agriculture states that just the laboratory costs for a negative environmental sample is $33, a positive environmental sample is $49.50, and negative egg test is $32.50, and a positive egg test is $41.25. FDA determined the cost of egg testing to be $1,859/1,000 eggs. This included collecting the eggs, the value of the eggs collected, shipping, and lab costs. Therefore the cost of 4 egg tests would be $7,436. In addition to the under estimation of the actual costs of doing the testing, the proposed rule does not address the labor involved in doing some of the testing, especially the egg testing. In a time, when laboratory budgets are being cut and personnel are being reduced, many laboratories may not be able to handle this increased volume of samples. For example the Ohio Department of Agriculture submitted a breakdown in the manpower requirements for culturing 1,000 eggs as follows: prep time – 0.5 man-hours (1 person, 30 min), breaking eggs – 10 man-hours (10 people, 1 hour each), culturing - 2 man-hours (2 people, 1 hour each), and reading a negative culture – 0.5 man-hours (1 person, 30 min). Another aspect of this additional testing is the amount of space and equipment that will be necessary to meet the requirements. These costs are very difficult to pass on to the customers or, in the case of space, not achievable. Therefore it seems essential to minimize the testing program as much as possible and still obtain the required results. This proposal comes on the heels of another new poultry monitoring program. The monitoring program for Avian Influenza (AI) will dramatically increase the number of samples that will be tested for AI in this country and many of the laboratories that would be testing for Salmonella will be also responsible for that program. Further stretching laboratory space, personnel and funds. Asking the layer companies and state laboratories to fund this mandatory testing program is unrealistic. If FDA is requiring a mandatory testing program then there should be federal funds allocated to cover all or part of the costs. Notification of results Laboratories testing these samples should screen for the presence of Salmonella and then test for agglutination with serogroup D1 antisera. Any serogroup D1 isolates should be sent to the NVSL or an accredited regional laboratory for serotyping. Any isolates confirmed as SE should also be phage typed. The USDA-NVSL or USDA accredited regional laboratory will notify the submitting laboratory of the identification of the isolates. Once the laboratory receives the confirmation of SE from NVSL, the laboratory should notify the State or FDA responsible entity and the respective company having the SE positive house. D. Other comments (Section VI – Laboratory Issues) Based on the definitions of farm, house, and flock in the FDA proposed rule, we would like to clarify a possible testing scenario. Given a 10-house layer complex, essentially every month one or more of the houses will be environmentally tested based on the age of the birds in a house. If, for example, house 4 becomes environmentally positive this would not result in any additional environmental sampling of other houses. Furthermore, only the eggs from house 4 will be tested for SE. Also any required C & D that results from the positive environmental sample will only apply to house 4. VII. MODEL FOOD CODE: Salmonella control requires an integrated Farm to Fork approach. Only an inclusive program will achieve the desired results of human disease reduction. The proposed program only addresses the on-farm aspect of control. A program that includes processing and most importantly food service portions of egg distribution will have the greatest impact on human health. USAHA supports a much more comprehensive approach to egg safety. We recommend mandatory adoption, and adequate funding, to enforce the egg safety requirements in FDA's Model Food Code, to assure proper handling and preparation of eggs. As an example, only pasteurized eggs should be used in an institutional or retail setting where food preparation requires pooling of eggs. Many current outbreaks continue to be the direct result of improper food handling, an important risk factor that must be more effectively addressed. Most importantly, bulk pooling of eggs should only be done with pasteurized product, because it is a high-risk activity. In addition to pasteurization requirements, we specifically recommend enforcement of the following measures of the Model Food Code in order to meet the stated human health goals. 1. Proper storage of eggs including refrigeration and hygiene standards 2. Prohibit pooling of > 2 eggs at a time 3. Monitoring of adequate time and temperature parameters when cooking eggs 4. Health standards of food workers VIII. BIOSECURITY: There are five biosecurity points in the proposed rule but they are quite simply not appropriate for SE control. Further, the proposed biosecurity rules are said to be important to “ensure” freedom from SE. Particularly with a disease organism with multiple hosts, it is not possible to ensure freedom from infection with any biosecurity program. Any biosecurity program for SE must be harmonized with biosecurity standards set for Avian Influenza and Exotic Newcastle Disease. Maintaining complete isolation of houses on multi-age layer complexes is not practical and maybe impossible. Efforts should concentrate on eliminating transmission of disease between farms and not between houses on farms. The biosecurity portions of the proposal do not take into account the most effective control methods for house-to-house exposure. These intervention points overlap with pest control and vaccination. Rodents are far more important in house-to-house transmission than clothing worn between houses. Efforts can be made to prevent transmission between houses. Interventions such as vaccination of adjacent houses will help stop the spread of the bacteria across the farm/ranch more so than changing clothes. Every location will have its own unique challenges and solutions. Only general rules of biosecurity should be in the final rule leaving the details to the flock owners. Well-educated flock owners will then be able to design program directed at preventing introduction of SE onto their farms and containing the infection to a single house in the event of an outbreak. The following are known to be important in reducing the risk of SE transmission between farms: decontamination and control of a.) Vehicles and people used to move pullets between the growing house and layer house; b.) Vehicles and people used to move out spent fowl; c.) people entering the farm; d.) Equipment entering the farm; e.) Egg flats, racks, pallets, etc. and f.) On-farm traffic between egg rooms and chicken houses. IX. REFRIGERATION REQUIREMENTS: Experimental evidence demonstrates that actual deposition of SE inside the contents of the yolk is exceedingly rare (to the point of being almost inconsequential from an epidemiological standpoint). Supporting this line of thinking, he has seen that 90% of freshly laid eggs from hens given huge oral doses of SE harbored 7 or fewer SE cells per ml of liquid contents, and none harbored more than 26 cells per ml. Moreover, Dr. T. Humphrey in the UK (Humphrey, T.J. 1990. Growth of Salmonella in Intact Shell Eggs: Influence of Storage Temperature. Vet. Rec. 1236:292) has reported that small numbers of SE deposited in albumen will rarely grow to reach high levels before the yolk membrane begins to deteriorate after 2-3 weeks of storage at room temperature. Therefore, the intrinsic physical and biochemical defense mechanisms of eggs help limit bacterial growth except in the rare event of initial deposition inside yolks. University of Minnesota research (Kim, C., et al. 1989. Effect of Time and Temperature Growth of Salmonella Enteritidis in Experimentally Inoculated Eggs. Avian Diseases. 33:735–742) showed that with low number of SE organisms injected into the albumen grew minimally for the first 10 days of storage at temperatures in excess of 60F. Why omit producers with less than 3000 layers from cooling provisions? The environmental conditions on many small farms are likely to present a greater risk of supporting the introduction and perpetuation of pathogens than those present on larger operations. These producers should do environmental testing. The only adjustment that should be made for small producers is that the number of eggs sampled on environmentally positive small farms should be proportionately reduced to an appropriate level. Why cool to 45 F those eggs destined for treatment? In the absence of evidence that heat-treated eggs cause a public health problem there is no reason to insist upon a stringent 36 hour 45 F rule. Eggs held at 45 F on the farm will have condensation problems, thermal checks during washing and reduced yield when broken. Some eggs that go to a breaking plant are from breeder flocks and such eggs cannot be held at 45F. A lower standard of cooling (in terms of either promptness or temperature) might be justifiable for eggs destined for pasteurization than for table eggs. Why 36 hours? It is possible that eggs could be held at 55 to 60 degrees at the farm for up to 7 days without any significant impact on public health. Bacteria deposited in albumen are unlikely to grow in such a short period of time (and in fact are unlikely to grow much for at least 2 to 3 weeks). Although there are very minimal solid data available regarding the true location at which SE is deposited in naturally contaminated eggs, most indirect evidence (such as the fact that highly contaminated eggs are quite rare, even when experimentally infected hens are given large oral doses of SE) suggests that albumen contamination is a much more common event than yolk contamination. The idea that naturally contaminated eggs will use their natural biochemical defenses to resist bacterial growth for several weeks is a generalization that is usually but not always true. Consider a hypothetical scenario: If 1 in 20,000 eggs is contaminated with SE (USDA Risk Assessment calculation), and if 1% of these involve yolk contamination, than about 35,000 of the ~70 billion eggs produced in the USA every year could have SE in the yolk. These eggs would have already spent 24 hours in the oviduct at about 106 F (and thus experienced rapid bacterial growth) before refrigeration begins. Maximal bacterial growth in such eggs may occur before refrigeration temperatures could be achieved inside the egg no matter when or at what temperature refrigeration began. It is conceivable that many human outbreaks are actually caused by such eggs after SE multiplication produces dangerously high infectious doses. The risk to public health of failure to refrigerate yolk-contaminated eggs as soon as possible is unknown. Impact of moving cold eggs in warm humid weather? Moving cold eggs in warm humid weather will induce a significant amount of condensation of water on the shell. High moisture levels on eggshells are likely to increase opportunities for penetration by pathogens (numerous Salmonella serotypes are commonly found on egg shells). Impact of washing cold egg in warm water? Washing cold eggs in warm water is certain to induce an undesirable degree of thermal cracking, resulting in either loss of marketable table eggs or in an increased risk of pathogen contamination of contents when cracks are not detected. Summary – Section IX – Refrigeration Requirements In summary, we feel that a change from the presently used cooler room temperature of 55F to holding eggs at 45F prior to processing would not contribute to the reduction of SE infections in humans. The presently used holding temperature of 55F for eggs prior to processing should be continued. X. DIVERSION OF EGGS TO PASTEURIZATION ISSUES: The following comments seem to be made from the thought rather than the data mostly because little data is available. What are the alternatives for handling egg positive flocks? A few options for handling a positive flock are sending eggs to pasteurization, hard-cooking, destruction of the flock, vaccination (plus or minus probiotics) in lay and testing until a negative test, wait it out and hope they test clean, inedible processing, dump eggs in pit, or send eggs to rendering. All of the alternatives result in a possible break-even to a gross loss either as eggs or birds. In none of these instances are the eggs used for human food unless the egg-testing program allows the return the egg to a shell egg market. The last alternative is to break the eggs and use them as a further processed product from pasteurization or hard-boiling. What would be the impact on the total egg market? The total egg market now is currently about 30% breaking and 70% shell egg. First, what theoretically would be the number of eggs to be diverted? If 9.7% of the flocks are positive and of those flocks 26% have one egg positive, 2.5% of the shell egg flocks’ eggs will have to be diverted. Can this number be diverted into breaking? The market is fluid. As eggs are presented for breaking and shell eggs are low, available eggs will flow into the shell egg market. In theory, the breaking market has enough flexibility to take the 2.5% with no problem in most cases. Will these eggs be purchased at the available market levels? No, they will most likely be discounted. Trucking to available plants will be expensive. Plant location may be result in impractical trucking distances. Small producers who niche market products and have higher per dozen costs will experience a greater loss because the difference between the shell egg price and breaking price will be greater than the large producer and transportation costs on a per dozen basis will be greater. On a national level, breaking capacity may be adequate. However, breaking plant distribution is not even across the country. Several egg production centers, such as the west and the northeast, developed around a model of shell egg production centered on urban areas. These regions have seen a decrease in breaking capacity in recent years. Hawaii does not have a single breaking plant in the entire state. A positive flock there might require depopulation and indemnification. Other regions have extensive breaking capacity to the point on farms dedicated 100% to liquid egg product. These in-line plants are not always designed to absorb outside eggs into their production system and if they do create bio-security challenges. Additionally, some farms have no means to reroute eggs from their breaking plants to the shell egg market. What effect will this number have on the shell egg market economically? After some discussion with marketers the answer as to the effects will be the result of supply and demand. If the year happens to have an excess of eggs, the market will not be affected since excess eggs will flow into the shell egg supply. If the available excess eggs are in short supply the price may be 20 cents higher than normal like the last year. Late 2003/early 2004 price spike coincided with a 3% decrease in national flock size due to Exotic Newcastle Disease depopulations There seems to be no way of determining the market reaction to egg diversion strategies. However, if the market cannot absorb the projected 2.5% positive portion of the national flock, and depopulation or diversion to a non-human food product becomes the alternative. The resulting changes in production could have a destabilizing influence on shell egg market. There are two possible points with no specific answers. 1.. Egg processors currently have set supplies of eggs to meet quality requirements for customers. If customers balk at using eggs from SE positive flocks, breakers will be forced to not use these eggs, which will result in the flocks having to be killed rather than the eggs processed. 2.. What is the evidence that diversion is the action of choice for positive flocks considering that transovarian transfer is variable and mostly when birds are shedding at higher rates of infectivity? Treatments have shown helpful but inconsistent abilities to reduce shed while vaccines have shown a more consistent ability to prevent transovarian transfer. Our conclusion is that on the national level the breaking capacity exists for the projected infection rate. However, variance in regional infection rates and regional access to breaking plants will impact this model making the economic effects impossible to predict. Positive eggs will be discounted if sold to the breaking market and/or at no better than current farm prices. The big unknown will be whether consumers (or special interest groups) will consider this SE positive egg an adulterant and ask breaker plants not to use them. Sincerely, Richard Willer, President United States Animal Health Association 8100 Three Chopt Road, Suite 203 P. O. BOX K227 RICHMOND, VIRGINIA 23288 804- 285-3210 FAX 804-285-3367 E-Mail: usaha@usaha.org Web Site: www.usaha.org