Progress 04/25/01 to 02/11/05
Outputs
1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Foodborne disease is one of the most important issues in public health and animal agriculture. Changes in the production, processing, distribution and consumption of food have contributed to the emergence of new pathogens and to the increasing importance of old ones. This has markedly influenced both the domestic and international markets for meat and related products in the U.S. Prevention and control strategies for foodborne pathogens must be designed and implemented at all levels of the production cycle, including the preharvest period, to reduce the risk to consumers and economic impact on producers. The gastrointestinal tracts of livestock serve as the source of some of the more important bacteria which cause foodborne diseases, such as Escherichia coli O157:H7 and Salmonella typhimurium DT104.
Thus, understanding the epidemiology and ecology of livestock infection with such bacteria, as well as the interaction of bacteria with livestock and other host species, is required for rational design of control strategies in the preharvest portion of the production cycle. Developing improved diagnostic methods for important foodborne bacterial species allows for very accurate studies of natural infections in commercial and experimental herds of cattle. Sources of infection, dynamics of infection and effects of external influences (such as management, environment, etc.) are identified as potential control points to reduce prevalence of these bacteria in livestock, thereby, improving the safety of the U.S. food supply. This project supports National Program 108, Food Safety (100%). Determining how to reduce microbial pathogens in food products, throughout food operations from farm to fork, is the most urgent food safety problem today. A major goal of this program is to develop tests
that are precise and rapid enough to detect contamination in all foods prior to their entering into commerce. Equally important is the development of effective, reliable and cost-effective methods to control or eliminate pathogens in/on food producing animals throughout production and processing. Using epidemiological surveillance, a better definition of the economic impact of livestock diseases can be gained to better understand the ecology of emerging diseases and natural transmission cycles. This is needed to develop control strategies to prevent disease. The emergence of a new disease or introduction of an exotic disease into the U.S. could rapidly escalate into an epidemic due to the lack of resistance in host animals, absence of vaccines or effective drugs and limited resources to effectively manage the spread of these pathogens. 2. List the milestones (indicators of progress) from your Project Plan. Early in FY2005 this project was combined with CRIS Project #5438-32000-
020-00D. Milestones and accomplishments are highlighted in that project report. 4a What was the single most significant accomplishment this past year? Early in FY2005 this project was combined with CRIS Project #5438-32000- 020-00D. Milestones and accomplishments are highlighted in that project report. 4d Progress report. Early in FY2005 this project was combined with CRIS Project #5438-32000- 020-00D. Milestones and accomplishments are highlighted in that project report 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. This research addresses several goals of the National Program 108, Food Safety, Animal and Plant Products including: 1.1.1 - Sampling, isolation, identification, and quantification of pathogens in animal fluids and tissues, manure, and the environment, including feed, water, and wild animals. 1.2.1 - Ecology and assessment of risk factors of pathogens in food- producing animals, including those carrying
antibiotic resistance, outside of the host animal. 1.3.1 - Identify sites and mechanisms of colonization in animals, and identify and characterize virulence attributes which play a role in the host-pathogen relationship. Research activities are further related to ARS Strategic Plan Goal 3, Enhance Protection and Safety of the Nation's Agriculture and Food Supply. There have been four major accomplishments over the life of this project. First, we have shown that there is a clear and significant relationship between E. coli O157:H7 infection of live cattle and contamination of beef carcasses. This relationship underpins the entire concept of preharvest food safety. Second, we have developed an unsurpassed panel of monoclonal antibodies for the detection and characterization of EHEC and Salmonella, including type, group and virotype specific antibodies. Several of these antibodies have been formatted for diagnostic tests, including one test which is widely used in the U.S. meat industry.
Third, we have described the epidemiology of infection of cattle with EHEC and Salmonella typhimurium DT104 throughout the production cycle. These studies have shown the differences in progression of infection between these types of bacteria in cattle and have quantified the transmission between infected and naive cattle, important parameters for designing preharvest control methods. Epidemiologic studies were made possible by the development of sensitive and specific diagnostic methods. Using these methods it was shown that E. coli O157:H7 prevalence in beef cattle was underestimated by nearly 10-fold. Fourth, we have evaluated several potential intervention strategies to reduce fecal shedding of E. coli O157:H7. Results of these experiments will reduce utilization of plausible, but ineffective, control strategies for E. coli O157:H7 and have indicated strategies worthy of further development. 6. What science and/or technologies have been transferred and to whom? When is the science
and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? There has been great interest in the diagnostic techniques, reagents and protocols among industry, research, academic, diagnostic and regulatory groups. Scientists from major meat packing companies, several universities, regulatory and other Federal researchers have been provided with culture protocols and/or been trained in the use of this technique in our laboratories. In addition, we have responded to many requests for information about cross-species microarray hybridization. Constraints on the development of rational preharvest control strategies still remain, particularly incomplete knowledge of the ecology and epidemiology of the relevant bacteria in a production setting. 7. List your most important publications in the popular press and presentations to organizations and articles
written about your work. (NOTE: List your peer reviewed publications below). Early in FY2005 this project was combined with CRIS Project #5438-32000- 020-00D. Milestones and accomplishments are highlighted in that project report.
Impacts
(N/A)
Publications
- Bono, J.L., Keen, J.E., Clawson, M.L., Heaton, M.P., Chitko Mckown, C.G., Miller, L.C., Laegreid, W.W. 2005. Discrimintion of Escherichia coli O157:H7 isolates by genotyping single nucleotide polymorphisms. [abstract] American Society of Microbiologists. p. 318.
- Chitko Mckown, C.G., Green, B.T., Miller, L.C., Mckown, R.D., Laegreid, W. W. 2004. Development and characterization of three porcine monocyte- derived cell lines [abstract]. Research Workers in Animal Diseases Conference. p. 96.
- Laegreid, W.W., Bauer, N. Probiotics for pathogen control in poultry and livestock. 2004. American Meat Science Association Conference Reciprocal Proceedings.
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Progress 10/01/03 to 09/30/04
Outputs
1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Foodborne disease is one of the most important issues in public health and animal agriculture. Changes in the production, processing, distribution and consumption of food have contributed to the emergence of new pathogens and to the increasing importance of old ones. This has markedly influenced both the domestic and international markets for meat and related products in the U.S. Prevention and control strategies for foodborne pathogens must be designed and implemented at all levels of the production cycle, including the preharvest period, to reduce the risk to consumers and economic impact on producers. The gastrointestinal tracts of livestock serve as the source of some of the more important bacteria which cause foodborne diseases, such as Escherichia coli O157:H7 and Salmonella typhimurium DT104.
Thus, understanding the epidemiology and ecology of livestock infection with such bacteria, as well as the interaction of bacteria with livestock and other host species, is required for rational design of control strategies in the preharvest portion of the production cycle. Developing improved diagnostic methods for important foodborne bacterial species allows for very accurate studies of natural infections in commercial and experimental herds of cattle. Sources of infection, dynamics of infection and effects of external influences (such as management, environment, etc.) are identified as potential control points to reduce prevalence of these bacteria in livestock, thereby, improving the safety of the U.S. food supply. This project supports National Program 108, Food Safety (100%). Determining how to reduce microbial pathogens in food products, throughout food operations from farm to fork, is the most urgent food safety problem today. A major goal of this program is to develop tests
that are precise and rapid enough to detect contamination in all foods prior to their entering into commerce. Equally important is the development of effective, reliable and cost-effective methods to control or eliminate pathogens in/on food producing animals throughout production and processing. Using epidemiological surveillance, a better definition of the economic impact of livestock diseases can be gained to better understand the ecology of emerging diseases and natural transmission cycles. This is needed to develop control strategies to prevent disease. The emergence of a new disease or introduction of an exotic disease into the U.S. could rapidly escalate into an epidemic due to the lack of resistance in host animals, absence of vaccines or effective drugs and limited resources to effectively manage the spread of these pathogens. This research addresses several goals of the National Program Action Plan for Food Safety including: 1.1.1 - Sampling, isolation, identification, and
quantification of pathogens in animal fluids and tissues, manure, and the environment, including feed, water, and wild animals. 1.2.1 - Ecology and assessment of risk factors of pathogens in food- producing animals, including those carrying antibiotic resistance, outside of the host animal. 1.3.1 - Identify sites and mechanisms of colonization in animals, and identify and characterize virulence attributes which play a role in the host-pathogen relationship. 2. List the milestones (indicators of progress) from your Project Plan. FY 2002: Sequencing of first sets of enterohemorrhagic E. coli (EHEC) O157 gene targets. Sampling of infected cattle for host immune response studies. Transmission parameters for EHEC O157 estimated. FY 2003: Sampling of cattle for reservoir and carrier studies. Host immune response analysis. Bacterial constructs for host gene expression studies. FY 2004: Host gene expression studies. Bacterial gene expression studies. FY 2005: Completion of confirmatory
studies on host gene expression. 3. Milestones: A. List the milestones that were scheduled to be addressed in FY 2004. How many milestones did you fully or substantially meet in FY 2004 and indicate which ones were not fully or substantially met, briefly explain why not, and your plans to do so. FY 2002: Sequencing of first sets of EHEC O157 gene targets. Completed, manuscript in preparation. Sampling of infected cattle for host immune response studies. Completed. Transmission parameters for EHEC O157 estimated. Completed, manuscript published. FY 2003: Sampling of cattle for reservoir and carrier studies. Reduction in funding prevented these studies. Host immune response analysis. Partially completed, emphasis shifted to innate immunity, data being analyzed. Bacterial constructs for gene expression studies. In progress. FY 2004: Host gene expression studies. Substantially completed, some gene targets being further characterized, gene expression under investigation in other cell
types. Bacterial gene expression studies. In progress. FY 2005: Completion of confirmatory studies on host gene expression. Substantially completed, one manuscript published. B. List the milestones that you expect to address over the next 3 years (FY 2005, 2006, & 2007). What do you expect to accomplish, year by year, over the next 3 years under each milestone? FY 2005: Completion of confirmatory studies on host gene expression. Characterization of host gene expression in other cell types in response to EHEC O157. Bacterial gene expression studies. Bacterial gene influences on infection-related phenotypes characterized. 4. What were the most significant accomplishments this past year? A. Single most significant accomplishment during FY 2004: As a result of deep resequencing through panels of E. coli isolates, genes representing highly specific diagnostic targets for E. coli O157:H7 were identified. Real-time polymerase chain reaction (RT-PCR) assays were developed for these gene
targets and when compared their specificity exceeds existing RT-PCR assays. Optimization of sample preparation for use in live cattle diagnostics is proceeding as is adaptation to field instrumentation for on-farm testing. B. Other significant accomplishment(s), if any: Examined expression of over 7,000 genes in bovine macrophages in response to E. coli O157:H7 lipopolysaccharide (LPS) with expression of 44 genes either increased or decreased significantly. Eighteen of the differentially expressed genes were previously not described in cattle. These experiments were performed using human gene chips and bovine cells, the first use of cross-species gene expression profiling using microarray hybridization. Results were confirmed using RT-PCR. Impact: These differentially expressed genes are potential candidates for EHEC infection resistance genes in cattle. C. Significant activities that support special target populations: None D. Progress Report: None 5. Describe the major
accomplishments over the life of the project, including their predicted or actual impact. There have been four major accomplishments over the life of this project. First, we have shown that there is a clear and significant relationship between E. coli O157:H7 infection of live cattle and contamination of beef carcasses. This relationship underpins the entire concept of preharvest food safety. Second, we have developed an unsurpassed panel of monoclonal antibodies for the detection and characterization of EHEC and Salmonella, including type, group and virotype specific antibodies. Several of these antibodies have been formatted for diagnostic tests, including one test which is widely used in the U.S. meat industry. Third, we have described the epidemiology of infection of cattle with EHEC and Salmonella typhimurium DT104 throughout the production cycle. These studies have shown the differences in progression of infection between these types of bacteria in cattle and have quantified the
transmission between infected and naive cattle, important parameters for designing preharvest control methods. Epidemiologic studies were made possible by the development of sensitive and specific diagnostic methods. Using these methods it was shown that E. coli O157:H7 prevalence in beef cattle was underestimated by nearly 10-fold. Fourth, we have evaluated several potential intervention strategies to reduce fecal shedding of E. coli O157:H7. Results of these experiments will reduce utilization of plausible, but ineffective, control strategies for E. coli O157:H7 and have indicated strategies worthy of further development. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? There has been great interest in the diagnostic techniques, reagents and
protocols among industry, research, academic, diagnostic and regulatory groups. Scientists from major meat packing companies, several universities, regulatory and other Federal researchers have been provided with culture protocols and/or been trained in the use of this technique in our laboratories. In addition, we have responded to many requests for information about cross-species microarray hybridization. Constraints on the development of rational preharvest control strategies still remain, particularly incomplete knowledge of the ecology and epidemiology of the relevant bacteria in a production setting. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. Use of DNA markers for traceback and identification of cattle, NCBA Annual Convention, Phoenix, AZ, January 2004. Probiotics for pathogen control in poultry and livestock, Reciprocal Meat Conference, Lexington, KY, June 2004. A real-time PCR assay
that detects E. coli O157:H7, FSIS/ARS Annual Research Workshop, February 2004. Popular press reports about this research appeared in Medical Devices & Surgical Technology Week, Medical Letter on the CDC & FDA, Drug Week, Genomics & Genetics Weekly, Health Business Week, Physician Law Weekly, Pharma Law Weekly, TB & Outbreaks Week, and Health & Medicine Week.
Impacts
(N/A)
Publications
- BONO, J.L., KEEN, J.E., MILLER, L.C., FOX, J.M., CHITKO MCKOWN, C.G., HEATON, M.P., LAEGREID, W.W. EVALUATION OF A REAL-TIME PCR KIT FOR DETECTING ESCHERICHIA COLI O157 IN BOVINE FECAL SAMPLES. APPLIED AND ENVIRONMENTAL MICROBIOLOGY. 2004. V. 70. P. 1855-1857.
- CHITKO MCKOWN, C.G., FOX, J.M., MILLER, L.C., HEATON, M.P., BONO, J.L., KEEN, J.E., GROSSE, W.M., LAEGREID, W.W. GENE EXPRESSION PROFILING OF BOVINE MACROPHAGES IN RESPONSE TO ESCHERICHIA COLI O157:H7 LIPOPOLYSACCHARIDE. DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY. 2004. V. 28. P. 635-645.
- Laegreid, W.W., Keen, J.E. 2004. Estimation of the basic reproduction ratio (ro) for shiga toxin-producing escherichia coli o157:h7 (stec o157) in beef calves. Epidemiology and Infection. 132(2):291-295.
- Bono, J.L., Keen, J.E., Clawson, M.L., Heaton, M.P., Fox, J.M., Chitko McKown, C.G., Laegreid, W.W. 2004. Discrimination of Escherichia coli O157:H7 from other bacteria by genotyping single nucleotide polymorphisms [abstract]. American Society for Microbiology. Paper No. C-311.
- CHITKO MCKOWN, C.G., FOX, J.M., MILLER, L.C., HEATON, M.P., BONO, J.L., KEEN, J.E., GROSSE, W.M., LAEGREID, W.W. GENE EXPRESSION PROFILING OF BOVINE MACROPHAGES IN RESPONSE TO ESCHERICHIA COLI O157:H7 LIPOPOLYSACCHARIDE. IMMUNOLOGY RESEARCH WORKSHOP. 2003. ABSTRACT NO. 23.
- CHITKO MCKOWN, C.G., FREKING, B.A., MACNEIL, M.D., SMITH, T.P. DEVELOPMENT OF ANIMAL CELL LINES FOR USE IN GENOMIC STUDIES. RESEARCH WORKERS IN ANIMAL DISEASES CONFERENCE PROCEEDINGS. 2003. ABSTRACT NO. 90P.
- LAEGREID, W.W., CLAWSON, M.L., HEATON, M.P., KEEN, J.E., CHITKO MCKOWN, C. G., FOX, J.M., SMITH, T.P., SNELLING, W.M., KEELE, J.W. GENETIC RISK FACTORS FOR FAILURE OF PASSIVE TRANSFER IN THE BOVINE NEONATAL FC RECEPTOR. PLANT AND ANIMAL GENOME CONFERENCE PROCEEDINGS. ABSTRACT NO. P642.
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Progress 10/01/02 to 09/30/03
Outputs
1. What major problem or issue is being resolved and how are you resolving it? Foodborne disease is a major public health concern in the U.S. The gastrointestinal tracts of livestock serve as the source of some of the more important bacteria which cause foodborne diseases, such as Escherichia coli O157:H7 and Salmonella typhimurium DT104. Thus, understanding the epidemiology and ecology of livestock infection with such bacteria, as well as the interaction of bacteria with livestock and other host species, is required for rational design of control strategies in the preharvest portion of the production cycle. Developing improved diagnostic methods for important foodborne bacterial species allows for very accurate studies of natural infections in commercial and experimental herds of cattle. Sources of infection, dynamics of infection and effects of external influences (such as management, environment, etc.) are identified as potential control points to reduce prevalence
of these bacteria in livestock, thereby, improving safety of the U.S. food supply. 2. How serious is the problem? Why does it matter? Foodborne disease is one of the most important issues in public health and animal agriculture. Changes in the production, processing, distribution and consumption of food have contributed to the emergence of new pathogens and to the increasing importance of old ones. This has markedly influenced both the domestic and international markets for meat and related products in the U.S. Prevention and control strategies for foodborne pathogens must be designed and implemented at all levels of the production cycle, including the preharvest period, to reduce the risk to consumers and economic impact on producers. 3. How does it relate to the National Program(s) and National Program Component(s) to which it has been assigned? National Program 108, Food Safety (100%) Determining how to reduce microbial pathogens in food products, throughout food operations from
farm-to-fork, is the most urgent food safety problem today. A major goal of this program is to develop tests that are precise and rapid enough to detect contamination in all foods prior to their entering into commerce. Equally important is the development of effective, reliable and cost- effective methods to control or eliminate pathogens in/on food-producing animals throughout production and processing. Using epidemiological surveillance, a better definition of the economic impact of livestock diseases can be gained to better understand the ecology of emerging diseases and natural transmission cycles. This is needed to develop control strategies to prevent disease. The emergence of a new disease or introduction of an exotic disease into the U.S. could rapidly escalate into an epidemic due to the lack of resistance in host animals, absence of vaccines or effective drugs and limited resources to effectively manage the spread of these pathogens. 4. What were the most significant
accomplishments this past year? A. Single most significant accomplishment during FY 2003: Quantitative estimation of E. coli O157 transmission within cattle populations is critical baseline information for evaluation of prevention and control strategies for this important foodborne pathogen. Determining the basic reproduction ratio (Ro, a parameter indicative of transmission between individuals) for E. coli O157 in beef cattle represents the culmination of several years of research at the U.S. Meat Animal Research Center, developing accurate serologic tests for antibodies to the O157 antigen, adapting complex statistical methodologies to this problem and obtaining and analyzing over 10,000 serologic samples from beef cattle in a typical production environment. In addition to quantifying transmission, Ro is used to determine how effective an intervention must be to significantly reduce or eradicate E. coli O157 from beef cattle herds and may be used in predictive risk assessment
models. This is the first estimation of Ro for E. coli O157 in any species. B. Other significant accomplishments: None C. Significant activities that support special target populations: None D. Progress report: None 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. There have been four major accomplishments over the life of this project. First, we have shown that there is a clear and significant relationship between E. coli O157:H7 infection of live cattle and contamination of beef carcasses. This relationship underpins the entire concept of preharvest food safety. Second, we have developed an unsurpassed panel of monoclonal antibodies for the detection and characterization of enterohemorrhagic E. coli and Salmonella, including: type, group and virotype-specific antibodies. Several of these antibodies have been formatted for diagnostic tests, including one test which is widely used in the U.S. meat industry. Third, we have
described the epidemiology of infection of cattle with enterohemorrhagic E. coli and Salmonella typhimurium DT104 throughout the production cycle. These studies have shown the differences in progression of infection between these types of bacteria in cattle and have quantified the transmission between infected and naive cattle--important parameters for designing preharvest control methods. Epidemiologic studies were made possible by the development of sensitive and specific diagnostic methods. Using these methods, it was shown that E. coli O157:H7 prevalence in beef cattle was underestimated by nearly 10-fold. Fourth, we have evaluated several potential intervention strategies to reduce fecal shedding of E. coli O157:H7. Results of these experiments will reduce utilization of plausible, but ineffective, control strategies for E. coli O157:H7 and have indicated strategies worthy of further development. 6. What do you expect to accomplish, year by year, over the next 3 years? During FY
2004, we will trace the flow of E. coli O157:H7 and related bacteria in the production environment. Further studies on host and bacterial responses to infection will be initiated. In FYs 2003 and 2004, herd sampling and epidemiologic studies of E. coli reservoirs and transmission within and between herds will be continued. 7. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Reducing the numbers of cattle infected with E. coli O157:H7 should significantly reduce the risk of contamination of the U.S. meat supply. Methods to reduce numbers of infected cattle should be evaluated in naturally infected animals under production conditions to minimize the cost of instituting ineffective methods and to estimate unintended outcomes such as adverse health effects.
These studies found that two plausible control strategies were either ineffective (high levels of pen sanitation) or associated with significant adverse health and economic effects. Furthermore, the inadvertent beneficial effect of transport was demonstrated. These results have been communicated to cattle-producer groups directly and through the lay press (Beef, September 1999; National Geographic, May 2002). Widespread use of culture techniques with greater sensitivity will significantly improve monitoring of infected livestock, evaluation of intervention strategies and detection of environmental contamination. As the recent outbreak of enterohemorrhagic E. coli O111 in Texas demonstrated, serotypes other than O157:H7 represent threats to the U.S. food supply. Serotypes O26, O111 and O103, among others, are relatively more common elsewhere in the world and our studies have shown them to be present in U.S. cattle. Use of monoclonal antibodies to these O-antigen types in rapid
diagnostic and screening tests will provide an estimate of the potential risk to the U.S. meat supply. There has been great interest in these culture techniques, reagents and protocols among industry, research, academic, diagnostic and regulatory groups. Scientists from major meat packing companies, several universities, NVSL, NAHMS and other ARS researchers have been provided with culture protocols and/or trained in the use of this technique in our laboratories. Constraints on the development of rational preharvest control strategies still remain, particularly incomplete knowledge of the ecology and epidemiology of the relevant bacteria in a production setting.
Impacts
(N/A)
Publications
- BONO, J.L., KEEN, J.E., LAEGREID, W.W. SINGLE NUCLEOTIDE POLYMORPHISMS IN THE RFB LOCUS OF ESCHERICHIA COLI O157 STEC AND NON-STEC ISOLATES. INTERNATIONAL SYMPOSIUM AND WORKSHOP ON SHIGA TOXIN ... ESCHERICHIA COLI. 2003. ABSTRACT P. 116.
- CHITKO MCKOWN, C.G., HEATON, M.P., GROSSE, W.M., KEEN, J.E., BONO, J.L., CLAWSON, M.L., FOX, J.M., MILLER, L.C., LAEGREID, W.W. ASSOCIATION OF NOS2A HAPLOTYPES AND BACTERICIDAL ACTIVITY OF BOVINE MONOCYTE-DERIVED MACROPHAGES. AMERICAN ASSOCIATION OF IMMUNOLOGISTS PROCEEDINGS. 2003. ABSTRACT NO. 150.1.
- LAEGREID, W.W., BONO, J.L., CLAWSON, M.L., KEEN, J.E. SEQUENCE DIVERSITY OF ESCHERICHIA COLI O157:H7 ISOLATES. INTERNATIONAL SYMPOSIUM AND WORKSHOP ON SHIGA TOXIN ... ESCHERICHIA COLI. 2003. ABSTRACT P. 137.
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Progress 10/01/01 to 09/30/02
Outputs
1. What major problem or issue is being resolved and how are you resolving it? Foodborne disease is a major public health concern in the U.S. The gastrointestinal tracts of livestock serve as the source of some of the more important bacteria which cause foodborne diseases, such as Escherichia coli O157:H7 and Salmonella typhimurium DT104. Thus, understanding the epidemiology and ecology of livestock infection with such bacteria, as well as the interaction of bacteria with livestock and other host species, is required for rational design of control strategies in the preharvest portion of the production cycle. Developing improved diagnostic methods for important foodborne bacterial species allows for very accurate studies of natural infections in commercial and experimental herds of cattle. Sources of infection, dynamics of infection and effects of external influences (such as management, environment, etc.) are identified as potential control points to reduce prevalence
of these bacteria in livestock, thereby, improving the safety of the U.S. food supply. 2. How serious is the problem? Why does it matter? Foodborne disease is one of the most important issues in public health and animal agriculture. Changes in the production, processing, distribution and consumption of food have contributed to the emergence of new pathogens and to the increasing importance of old ones. This has markedly influenced both the domestic and international markets for meat and related products in the U.S. Prevention and control strategies for foodborne pathogens must be designed and implemented at all levels of the production cycle, including the preharvest period, to reduce the risk to consumers and economic impacts on producers. 3. How does it relate to the national Program(s) and National Program Component(s) to which it has been assigned? National Program 108, Food Safety (100%). Determining how to reduce microbial pathogens in food products, throughout food operations
from farm to fork, is the most urgent food safety problem today. A major goal of this program is to develop tests that are precise and rapid enough to detect contamination in all foods prior to their entering into commerce. Equally important is the development of effective, reliable and cost-effective methods to control or eliminate pathogens in/on food producing animals throughout production and processing. Using epidemiological surveillance, a better definition of the economic impact of livestock diseases can be gained to better understand the ecology of emerging diseases and natural transmission cycles. This is needed to develop control strategies to prevent disease. The emergence of a new disease or introduction of an exotic disease into the U.S. could rapidly escalate into an epidemic due to the lack of resistance in host animals, absence of vaccines or effective drugs and limited resources to effectively manage the spread of these pathogens. 4. What was your most significant
accomplishment this past year? A. Single most significant accomplishment(s) during FY 2002: No highly significant accomplishments in FY 2002. B. Other significant accomplishment(s), if any: None C. Significant activities that support special target populations: N/A D. Progress report: N/A 5. Describe your major accomplishments over the life of the project, including their predicted or actual impact? There have been four major accomplishments over the life of this project. First, we have shown that there is a clear and significant relationship between infection of live cattle with E. coli O157:H7 and contamination of beef carcasses. This relationship underpins the entire concept of preharvest food safety. Second, we have developed an unsurpassed panel of monoclonal antibodies for the detection and characterization of enterohemorrhagic E. coli and Salmonella, including type, group and virotype specific antibodies. Several of these antibodies have been formatted for diagnostic tests,
including one test which is widely used in the U.S. meat industry. Third, we have described the epidemiology of infection of cattle with enterohemorrhagic E. coli and Salmonella typhimurium DT104 throughout the production cycle. These studies have shown the differences in progression of infection between these types of bacteria in cattle and have partially quantified the transmission between infected and naive cattle, important parameters for designing preharvest control methods. Fourth, we have evaluated several potential intervention strategies to reduce fecal shedding of E. coli O157:H7. Results of these experiments will reduce utilization of plausible, but ineffective, control strategies for E. coli O157:H7 and have indicated strategies worthy of further development. 6. What do you expect to accomplish, year by year, over the next 3 years? During FY 2003, we will continue to develop improved methods to trace the flow of E. coli O157:H7 and related bacteria in the production
environment. Further studies on host and bacterial responses to infection will be initiated. In FY 2003-2004, herd sampling and epidemiologic studies of E. coli reservoirs and transmission within and between herds will be continued. 7. What technologies have been transferred and to whom? When is the technology likely to become available to the end user (industry, farmer other scientist)? What are the constraints, if known, to the adoption durability of the technology? Reduction of numbers of cattle infected with E. coli O157:H7 should significantly reduce the risk of contamination of the U.S. meat supply. Methods to reduce numbers of infected cattle should be evaluated in naturally infected animals under production conditions to minimize costs of instituting ineffective methods and to estimate unintended outcomes such as adverse health effects. These studies found that two plausible control strategies were either ineffective (high levels of pen sanitation) or associated with
significant adverse health and economic effects. Furthermore, the inadvertent beneficial effect of transport was demonstrated. These results have been communicated to cattle producer groups directly (NCBA) and through the lay press (Beef, September 1999). Widespread use of culture techniques with greater sensitivity will significantly improve monitoring of infected livestock, evaluation of intervention strategies and detection of environmental contamination. As the recent outbreak of enterohemorrhagic E. coli O111 in Texas demonstrated, serotypes other than O157:H7 represent threats to the U.S. food supply. Serotypes O26, O111 and O103, among others, are relatively more common elsewhere in the world and our studies have shown them to be present in U.S. cattle. Use of monoclonal antibodies to these O antigen types in rapid diagnostic and screening tests will provide and estimate of the potential risk to the U.S. meat supply. There has been great interest in these culture
techniques, reagents and protocols among industry, research, academic, diagnostic and regulatory groups. Scientists from major meat packing companies, several universities, NVSL, NAHMS and other ARS researchers have been provided with culture protocols and/or been trained in the use of this technique in our laboratories. Constraints on the development of rational preharvest control strategies still remain, particularly incomplete knowledge of the ecology and epidemiology of the relevant bacteria in a production setting. 8. List your most important publications and presentations, and articles written about your work (NOTE: this does not replace your review publications which are listed below) Our work on the prevalence of E. coli O157:H7 in beef cattle and its relationship to contamination of meat products was described in the May 2002 issue of National Geographic Magazine.
Impacts
(N/A)
Publications
- Bono, J.L., Keele, J.W., Kutish, G.F., Keen, J.E., Laegreid, W.W. Genomic sample sequencing of three shiga-toxigenic E. coli (STEC) isolated from bovine feces. 2nd American Society of Microbiologist and the Institute for Genomic Research. 2002. Abstract p. 40.
- Bono, J.L., Keele, J.W., Kutish, G.F., Keen, J.E., Laegreid, W.W. Genomic sample sequencing of three bovine shiga-toxigene E. coli (STEC) O157, O111 and O26. Research Workers in Animal Diseases Conference. 2001. Abstract No. 6P.
- Bono, J.L., Keen, J.E., Laegreid, W.W. Detection of E. coli O157 in bovine feces using the R.A.P.I.D. system E. coli O157 detection kit. American Society for Microbiology. 2002. Abstract p. 130.
- Chitko-McKown, C.G., Laegreid, W.W., Heaton, M.P. Distribution of inducible nitric oxide synthase (NOS2) haplotypes in U.S. beef cattle. International Cytokine Society and the Society for Leukocyte Biology. 2001. Abstract No. 206.
- Uhlich, G.A., Keen, J.E., Elder, R.O. Variations in the csgD promoter of Escherichia coli O157:H7 associated with increased virulence in mice and increased invasion of HEp-2 cells. Infection and Immunity. 2002. v. 70. p. 395-399.
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Progress 10/01/00 to 09/30/01
Outputs
1. What major problem or issue is being resolved and how are you resolving it?
Foodborne disease is a major public health concern in the U.S. The gastrointestinal tracts of livestock serve as the source of some of the more important bacteria which cause foodborne diseases, such as Escherichia coli O157:H7 and Salmonella typhimurium DT104. Thus, understanding the epidemiology and ecology of livestock infection with such bacteria, as well as the interaction of bacteria with livestock and other host species, is required for rational design of control strategies in the preharvest portion of the production cycle. Developing improved diagnostic methods for important foodborne bacterial species allows for very accurate studies of natural infections in commercial and experimental herds of cattle. Sources of infection, dynamics of infection and effects of external influences (such as management, environment, etc.) are identified as potential control points to reduce prevalence of these bacteria in livestock, thereby, improving the safety of the U.S. food supply.
2. How serious is the problem? Why does it matter?
Foodborne disease is one of the most important issues in public health and animal agriculture. Changes in the production, processing, distribution and consumption of food have contributed to the emergence of new pathogens and to the increasing importance of old ones. This has markedly influenced both the domestic and international markets for meat and related products in the U.S. Prevention and control strategies for foodborne pathogens must be designed and implemented at all levels of the production cycle, including the preharvest period, to reduce the risk to consumers and economic impacts on producers.
3. How does it relate to the National Program(s) and National Component(s)?
National Program 108, Food Safety (60%), National Program 103, Animal Health (40%)Determining how to reduce microbial pathogens in food products, throughout food operations from farm to fork, is the most urgent food safety problem today. A major goal of this program is to develop tests that are precise and rapid enough to detect contamination in all foods prior to their entering into commerce. Equally important is the development of effective, reliable, and cost-effective methods to control or eliminate pathogens in/on food producing animals throughout production and processing. Using epidemiological surveillance, a better definition of the economic impact of livestock diseases can be gained to better understand the ecology of emerging diseases and natural transmission cycles. This is needed to develop control strategies to prevent disease. The emergence of a new disease or introduction of an exotic disease into the U.S. could rapidly escalate into an epidemic due to the lack of
resistance in host animals, absence of vaccines or effective drugs and limited resources to effectively manage the spread of these pathogens.
4. What were the most significant accomplishments this past year?
A. Single Most Significant Accomplishment during FY 2001 Year: B. Other Significant Accomplishments: C. Significant Accomplishments/Activities that Support Special Target Populations: D. Progress Report:
5. Describe the major accomplishments over the life of the project including their predicted or actual impact.
This CRIS project is a replacement for the 5438-42000-003-00D project which was scheduled to terminate on July 25, 1999. The Project Plan has been submitted and reviewed by the Panel and revisions are being made. There have been four major accomplishments over the life of this project. First, we have shown that there is a clear and significant relationship between infection of live cattle with E. coli O157:H7 and contamination of beef carcasses. This relationship underpins the entire concept of preharvest food safety. Second, we have developed an unsurpassed panel of monoclonal antibodies for the detection and characterization of enterohemorrhagic E. coli and Salmonella, including type, group and virotype-specific antibodies. Several of these antibodies have been formatted for diagnostic tests, including one test which is widely used in the U.S. meat industry. Third, we have described the epidemiology of infection of cattle with enterohemorrhagic E. coli and Salmonella typhimurium
DT104 throughout the production cycle. These studies have shown the differences in progression of infection between these types of bacteria in cattle and have partially quantified the transmission between infected and naive cattle, important parameters for designing preharvest control methods. Fourth, we have evaluated several potential intervention strategies to reduce fecal shedding of E. coli O157:H7. Results of these experiments will reduce utilization of plausible, but ineffective, control strategies for E. coli O157:H7 and have indicated strategies worthy of further development.
6. What do you expect to accomplish, year by year, over the next 3 years?
During FY 2002, we expect to trace the flow of E. coli O157:H7 and related bacteria in the production environment and identify immunologic correlates of bacterial clearance in cattle. Host and bacterial genetic response to infection will be examined. In FY 2001-2003, herd sampling and epidemiologic studies of E. coli reservoirs and transmission within and between herds will be continued.
7. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end user (industry, farmer, other scientists)? What are the constraints if known, to the adoption & durability of the technology product?
Reduction of numbers of cattle infected with E. coli O157:H7 should significantly reduce the risk of contamination of the U.S. meat supply. Methods to reduce numbers of infected cattle should be evaluated in naturally infected animals under production conditions to minimize costs of instituting ineffective methods and to estimate unintended outcomes such as adverse health effects. These studies found that two plausible control strategies were either ineffective (high levels of pen sanitation) or associated with significant adverse health and economic effects. Furthermore, the inadvertent beneficial effect of transport was demonstrated. These results have been communicated to cattle producer groups directly (NCBA) and through the lay press (Beef, September 1999). Improved detection methodology for E. coli O157:H7 was developed. There has been great interest in this methodology among industry, research, academic, diagnostic and regulatory groups. Scientists from major meat packing
companies, several universities, NVSL, NAHMS and other ARS researchers have been provided with the protocol and/or been trained in the use of this technique in our laboratories. Widespread use of culture techniques with greater sensitivity will significantly improve monitoring of infected livestock, evaluation of intervention strategies, and detection of environmental contamination. As the recent outbreak of enterohemorrhagic E. coli O111 in Texas demonstrated, serotypes other than O157:H7 represent threats to the U.S. food supply. Serotypes O26, O111 and O103, among others, are relatively more common elsewhere in the world and our studies have shown them to be present in U.S. cattle. Use of monoclonal antibodies to these O antigen types in rapid diagnostic and screening tests will provide and estimate of the potential risk to the U.S. meat supply. Constraints on the development of rational preharvest control strategies still remain, particularly incomplete knowledge of the ecology
and epidemiology of the relevant bacteria in a production setting.
8. List your most important publications in the popular press (no abstracts) and presentations to non-scientific organizations and articles written about your work (NOTE: this does not replace your peer-reviewed publications which are listed below)
Impacts
(N/A)
Publications
- Laegreid, W. W. Diagnosis of infection by Escherichia coli O157:H7 in beef cattle. National Conference on Animal Production Food Safety. 2000. Abstract p. 101-103.
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