Food and Drug Administration
Center for Food Safety and Applied Nutrition
U. S. Department of Agriculture
Food Safety and Inspection Service
May 13, 1999

Prepared by the Listeria monocytogenes Risk Assessment Team

Table of Contents

The Food and Drug Administration, Center for Food Safety and Applied Nutrition (FDA), in consultation with the Department of Agriculture, Food Safety and Inspection Service (FSIS), is conducting a risk assessment to determine the prevalence and extent of consumer exposure to foodborne Listeria monocytogenes and to assess the resulting public health impact of such exposure. The risk assessment team is collecting data in four areas: the presence of L. monocytogenes in foods, the consumption levels of these foods, information from epidemiological investigations, and data from experimentation that defines the dose-response relationship between this pathogen and populations with different immune conditions. This report describes the status of the risk assessment after development of the structure and culmination of the initial data collection. In addition to being a progress report, this document is an invitation for comments on the planned approach, assumptions and objectives for completion of the risk assessment and to request communication of any additional data that are relevant to the risk assessment.

Introduction

A series of outbreaks in the early 1980's in cole slaw, pasteurized milk and Mexican-style cheese impelled the recognition of this bacteria as an important foodborne pathogen. An intensive period of research revealed much about the microorganisms ecology, characteristics, presence in foods and public health effects. This research was summarized in several reviews and books (Farber et al., 1996; Farber and Peterkin, 1991:Miller et al., 1990; Ryser and Marth, 1991). The National Advisory Committee on Microbiological Criteria for Foods presented its analysis and recommendations to FSIS, FDA and other U.S. Government agencies in 1991 (NACMCF, 1991). Control strategies the committee recommended were to develop an effective surveillance system, target efforts on specific foods, and use a HACCP-based control program to cover foods from processing to consumption to minimize the presence, survival and multiplication of L. monocytogenes in foods.

Based upon the known characteristics of this microorganism and the disease, FDA maintains a policy of "zero tolerance" for L. monocytogenes in ready to eat foods. In other words, the detection of any L. monocytogenes in a 25 gram sample renders the food adulterated within the meaning of the Federal Food, Drug, and Cosmetic Act, 21 U.S.C. 342(a)(1). As recently as 1995, FDA affirmed this policy, as reflected in the decision in United States v. Union Cheese Co., 902 F. Supp. 778, 784, 786 (N.D. Ohio 1995). USDA’s Food Safety and Inspection Service, which regulates meat and poultry, likewise has historically had a zero tolerance policy for L. monocytogenes in ready-to-eat products.

FDA, in collaboration with FSIS, is conducting this risk assessment to gather scientific information, particularly recent additions to the literature, needed to review current programs relating to the regulation of L. monocytogenes contamination in foods to ensure that such programs provide maximum public health protection.

L. monocytogenes is a bacterium that occurs widely in both the agricultural (soil, plants, and water) and food processing environment. The bacterium is resistant to various environmental conditions such as high salt or acidity (Ryser and Marth, 1991). L. monocytogenes grows at low oxygen conditions and refrigeration temperatures, and survives for long periods in the environment, on foods, in the processing plant, and in the household refrigerator. Although frequently present in raw foods of both plant and animal origin, it also can be present in cooked foods due to post-processing contamination. L. monocytogenes has been isolated in such foods as raw and pasteurized fluid milk, cheeses (particularly soft-ripened varieties), ice cream, raw vegetables, fermented raw-meat sausages, raw and cooked poultry, raw meats (all types), and raw and smoked fish (Farber and Peterkin, 1991; FDA, 1999; Ryser and Marth, 1991). Even when L. monocytogenes is initially present at a low level in a contaminated food, the organism can multiply during storage, including storage at refrigeration temperatures. A survey of a wide variety of foods from the refrigerators of listeriosis patients in the United States found 11 percent of the samples contained L. monocytogenes (Pinner et al., 1992).

It is well established that ingestion of L. monocytogenes can cause serious human illness, listeriosis. (CAST, 1994; Rocourt and Cossart, 1997; Farber and Peterkin, 1991; Miller et al., 1990; Ryser and Marth, 1991). In 1997, the Centers for Disease Control and Prevention (CDC) Foodborne Diseases Active Surveillance Network (FoodNet) showed that of all food borne illnesses, the rate of hospitalization was highest for persons infected with L. monocytogenes (88 percent). Similarly, of all of the food borne pathogens tracked by CDC, L. monocytogenes had the highest case fatality rate in that 20 percent of persons hospitalized with listeriosis died. CDC also found that the incidence of listeriosis is 0.5 per 100,000 population, compared to a combined rate of 51.2 per 100,000 for all nine of the food borne illnesses surveyed (CDC, 1998). Thus, although serious, listeriosis is a relatively rare food borne illness. Most cases of listeriosis occur in pregnant women or individuals with a predisposing disease (such as alcoholism, diabetes, or cirrhosis of the liver) or an impaired immune system resulting from either a disease (such as AIDS) or immunosuppressive treatment for a malignancy or an organ transplant. (Rocourt and Cossart, 1997; Ryser and Marth, 1991).

Listeriosis has a long incubation time (up to 5 weeks) and a range of symptoms. Infection of a pregnant woman may result in flu-like symptoms with fever, muscular pain, or headache, or the listeriosis infection may be asymptomatic. Importantly, however, when a pregnant woman contracts listeriosis, the fetus or new borne infant is likely to suffer severe consequences from the maternal infection, including spontaneous abortion, fetal death, stillbirth, neonatal septicemia, or meningitis. In non-pregnant adults, septicemia and meningitis are the most common result of a listeriosis infection, although organ infections and mild gastroenteritis can also occur.

As noted, although the incidence of listeriosis is relatively low and the consequences of an infection may be severe, an estimated 2 to 6 percent of the healthy population harbors L. monocytogenes in their intestinal tract without signs of illness (Rocourt and Cossart, 1997; Ryser and Marth, 1991). Because the documented prevalence of L. monocytogenes in people and in commonly-eaten foods is much higher than the documented incidence of listeriosis, some experts believe that the ingestion of low levels of L. monocytogenes may not result in illness and thus, may not constitute a general public health hazard (Farber et al., 1996, ICMSF, 1994).

Other countries, including certain major trading partners of the United States, take a slightly different approach to L. monocytogenes contamination. Relying upon their interpretation of the existing scientific data, countries such as Canada and Denmark have a "non-zero tolerance" for L. monocytogenes for some classes of foods (ICMSF, 1994; IFST, 1995). For example, in Canada, ready-to-eat foods that have not been associated with an outbreak and do not allow any growth of L. monocytogenes during a 10 day period of refrigerated storage may contain up to 100 L. monocytogenes organisms per gram without being considered unlawful (Health Canada, 1994). Denmark has six classes of foods that have to meet various criteria for L. monocytogenes. In raw, ready to eat foods, for example, two of five samples can contain between 10 and 100 organisms per gram and no sample can exceed 100 organisms per gram. Although the course taken by other countries concerning L. monocytogenes contamination is not determinative of the U.S. approach, the policies of certain major trading partners provides further context to any reexamination of current U.S. policy.

Objectives of the Risk Assessment

As noted above, FDA and USDA/FSIS are jointly planning to conduct an assessment of the risk posed by L. monocytogenes to American consumers. A RA is a systematic and comprehensive collection of information and analysis of such information that promotes an understanding of the interactions of various factors in a complex situation and provides a basis for making decisions. The goal of this RA is to provide FDA and FSIS with the information needed to review current programs relating to the regulation of L. monocytogenes contamination in foods to assure that such programs provide maximum public health protection.

Risk Assessment Plan:
The RA will seek and analyze four types of information: information concerning the level of L. monocytogenes contamination of foods and consumption levels of such foods (i.e., an exposure assessment), information concerning the epidemiology of food borne listeriosis and information regarding the human health consequences of such exposure (i.e., a dose-response analysis). Because of the significant role that epidemiology plays in the dose-response relationship, dose-response and epidemiology will be addressed together under the heading of Public Health.

Exposure assessment, therefore, comprises prevalence/levels of L. monocytogenes in foods (#1) and the consumption of these foods (#2). The hazard assessment incorporates the epidemiology (#3) and experimental evidence (#4). The logical flow of the risk assessment is illustrated on Figure 1.

The RA will examine a number of issues, including which foods contribute most to the consumption of L. monocytogenes, what are the numbers of organisms when a food is contaminated, how frequently are foods heavily contaminated, are there common characteristics in the composition, processing or storage of contaminated foods, are some strains of L. monocytogenes more virulent that others, what is the extent of organism growth during storage (including storage at refrigeration temperatures), and what is the likelihood of illness to various subpopulations from consuming different numbers of L. monocytogenes. All assumptions and uncertainties in the RA will be identified and documented. The RA process will also include an evaluation of the adequacy of current scientific knowledge, data, and information. This will suggest where future research could be directed to reduce any uncertainty in the risk estimate that prevents a clear understanding of the causes and impact of listeriosis.

This risk assessment is concerned with the levels of L. monocytogenes present in the foods at the time of consumption. How those foods become contaminated at any particular level or the effects of a specified process, storage period or preparation practice is outside the immediate scope of this risk assessment. Upon completion of this assessment and the identification of specific foods which contribute the largest consumption of L. monocytogenes, it is anticipated that the risk managers in FDA would request a second phase of this risk assessment be developed to examine the causes of high levels in a food and anticipated effects of possible mitigations.

Table 1 lists the members of the risk assessment team. Risk management guidance is provided by senior FDA and FSIS officials, headed by Dr. R. L. Buchanan. The proposed time line for the project (Table 2) calls for this document to be presented in conjunction with a public meeting of the National Advisory Committee on Microbial Criteria for Foods (NACMCF). The risk assessment team will incorporate suggestions, recommendations and additional data from the May meeting and then begin the calculations and risk assessment phase. The target date to complete the risk assessment and present a draft report to the risk management team is September, 1999. Following internal review, presentations will be made and the risk assessment will be publicly available.

References

CAST. 1994. Foodborne Pathogens. Council for Agricultural Science and Technology, Task Force Report 122. Ames, IA.

CDC. 1998. Morbidity and Mortality Weekly Report., Incidence and Foodborne Illnesses-Foodnet, 1997. 47(37);782.

FDA. 1999. Bad Bug Book (Foodborne Pathogenic Microorganisms and Natural Toxins). Internet address: http://vm.cfsan.fda.gov/~mow/intro.html

Farber, J.M. and Peterkin, P.I. 1991. Listeria monocytogenes, a food-borne pathogen. Microbiol. Rev. 55:476-511.

Farber, J.M., Ross, W.H. and Harwig, J. 1996. Health risk assessment of Listeria monocytogenes in Canada. Int. J. Food Microbiol. 30:145-156.

FSIS. 1998. Salmonella Enteritidis risk assessment. Shell eggs and egg products. USDA, FSIS, Washington, DC.

Health Canada. 1994. Compliance Guide/Policy on Listeria Monocytogenes in Ready-to-Eat Foods.

ICMSF. 1994. Choice of sampling plan and criteria for Listeria monocytogenes. Int. J. Food Microbiol. 22:83-96.

IFST. 1995. Microbiological criteria for retail foods. Professional Food Microbiology Group, Inst. Food Science and Technology. Lett. Appl. Microbiol. 20:331-332.

Miller, A.L., Smith, J.L. and Somkuti, G.A. 1990. Foodborne Listeriosis. Soc. Indust. Microbiol., Elsevier, NY.

NACMCF. 1991. Listeria monocytogenes. Recommendations by the National Advisory Committee on Microbiological Criteria for Foods. International J. Food Microbiol. 14:185-246.

Pinner, R.W., Schuchat, A., Swaminathan, B., Hayes, P.S., Deaver, K.A., Weaver, R.E., Plikaytis, B.D., Reeves, M., Broome, C.V. and Wenger, J.D. 1992. Role of foods in sporadic listeriosis. 2. Microbiologic and epidemiologic investigation. JAMA 267:2046-2050.

Rocourt, J. and Cossart, P. 1997. Listeria monocytogenes. In Doyle, M.P., Beuchat. L.R. and Montville, T.J. eds. Food Microbiology, Fundamentals and Frontiers. ASM Press, Washington, DC.

Ryser, E.T. and Marth, E.H. 1991. Listeria, listeriosis, and food safety. Dekker, NY.

Introduction
The goal of this work-group is to collect the available Listeria monocytogenes food contamination data which will be used in conjunction with U. S. dietary intake data collected by the Food Consumption Work Group to determine how much of the viable pathogen is consumed in the USA.

Data collection

• General approach
  All possible data is being collected in order to see what is available. Subsequent data selection will depend on what kinds of data are available.

• Documentation Sources
  Any available data is being collected with preference for data published in the scientific literature and government documents. The preliminary list of data sources used so far is shown in the references. It mainly documents the larger multi-food studies.

• Geographical Origin
  Listeria occurrence data from all countries is being considered with preference for data of domestic and the imported foods. Listeria occurrence data from different countries do not seem to differ greatly overall.

• Organisms
  Data on food contamination by any Listeria species are being considered. The hope is that the amount of data about Listeria monocytogenes will be sufficient but, in cases where it is not, the other species can act as surrogates in providing data. Usually about 45% of the instances of Listeria occurrences are accounted for by L. monocytogenes.

• Serotypes
  Quantitative contaminant serotypes information will be considered as necessary. There do not appear to be major differences in the virulence of the various serotypes, despite differences in the frequencies of the clinical and foodborne occurrences. Information on the presence or frequencies of various serotypes in foodborne occurrences is not always available.

• Type of data
  Contaminant concentration data is preferable, but most data is presence and absence data. The latter type of data can be converted to concentration frequency data for a food group if at least some of the data is frequency data or frequency data from other food groups is appropriate.

• Foods
  The emphasis is on ready to eat foods, but this is not always easy to determine in dietary or microbial studies. Dietary intake data is often highly specific as to the food under consideration, e.g. smoked mussels. In contrast, contamination data is usually more generic, e.g. the frequency of contamination of shellfish. Thus, harmonization of the foods in dietary and microbiological data is a problem. A preliminary harmonized list of food groupings is given in Table 1. These are the categories that both the Food Contamination and the Food Consumption Modules will use.

• Time scale
  Data from 1980 to the present are being collected with special attention to the most recent data. It may be worthwhile to try comparing data from the eighties with that from the nineties to see if the increasing awareness of food-borne listeriosis affected contamination rates.

• Sample size
  The analytical portion size is generally 25 g. Usually it is taken from a non-composite sample. In the risk analysis phase these factors will have to be allowed for.

• Isolation method sensitivity
  Generally the methods used are well known and of equivalent sensitivity that is at least about 1 CFU/25 g (e.g., Hayes et al., 1992).

• Examples from the current database
  Table 2 contains the available data for sandwiches and it is illustrative of some of the above bullets.

Conclusion

The contamination data collection phase is still in progress. A comprehensive universal database may be useful to allow extrapolation of the values from one food group to another closely related one.

A. Foods in general

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Greenwood, M.H., Roberts, D., and Burden P. 1991. The occurrence of Listeria species in milk and dairy products; a national survey in England and Wales. Interntl. J. Food Microbiology 12:197-206

Klinger, I. And Rosenthal, I. 1997. Public health and the safety of milk and milk products from sheep and goats. Rev. Sci. Tech 16(2):482-484.

Loncarevic, S., Danielsson-Tham, M.L., and Tham, W. 1995. Occurrence of Listeria monocytogenes in soft and semi-soft cheeses in retail outlets in Sweden. International Journal of Food Microbiology 26:245-250.

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Epidemiological investigations of outbreaks and sporadic cases of listeriosis have clearly demonstrated that the consumption of contaminated food is responsible for a high proportion of human listeriosis cases (Rocourt, 1997; Tappero, 1995). Since the mid 80s, progress in microbiological detection and epidemiological investigations with increased use of case-control studies have demonstrated that all kinds of foods, at each step of the food chain, can transmit the disease (Rocourt, 1995). However, the importance of ingestion of specific contaminated foods in sporadic disease, which account for the majority of listeriosis cases, is unclear (Pinner, 1992; Schuchat, 1992). Because of the widespread nature of the organism, it has been nearly impossible to eliminate it from the food supply (Farber, 1991b; Shelef, 1989). The ability of L. monocytogenes to grow at refrigeration temperatures, coupled with appearance of the pathogen in raw and processed meats, as well as poultry, vegetables, fruit, dairy, and seafood products, makes this bacterium a serious threat to susceptible consumers and to the entire food industry (Ryser, 1989; Farber, 1991b; Schlech, 1991). In addition, a number of the foods associated with transmission of listeriosis have been highly processed, have extended shelf lives at refrigerated temperatures, are capable of supporting the growth of L. monocytogenes, and are consumed without further cooking (i.e., ready-to-eat) (McLauchlin, 1996). L. monocytogenes is also more heat resistant than most vegetative microbes (Farber, 1988), can be isolated in foods that have been adequately frozen (Tappero, 1995), and occurs in the home through cross-contamination with other foods (Schuchat, 1991).

The purpose of the food consumption module is to model the consumption of foods that have a high potential for L. monocytogenes contamination. These foods represent a subset of all foods that comprise an individual’s total diet. We will not include foods that have not been linked to L. monocytogenes contamination, such as grain products (e.g., bread, cookies, cakes), soft drinks, selected fruits (e.g., apples, bananas, avocados, lemons), cooked mixed dishes (e.g., lasagna, soups), and cooked vegetables. Once the levels of contamination of L.monocytogenes are identified for various foods in the Contamination Module, we will finalize food categories and provide the distributions of food consumption for those categories. With the food contamination data, we will then develop exposure estimates for not only the aggregate US population but also for different subpopulations that are particularly susceptible to listeriosis.

The first step in determining estimates of food consumption was to select appropriate food products to include in the risk assessment. In order to delineate foods that have a high potential for L. monocytogenes, we completed a comprehensive review of the literature. We concentrated on five subject areas that address the epidemiological and microbiological aspects of L. monocytogenes. Those subject areas with associated citations include:

All references used in the food consumption module and cited in the text will be included in the reference list, sorted by subject matter. Those general subjects are divided into background, listeriosis, and food groups (produce, dairy, seafood, and meat and poultry).

• Two 24-hour recalls of foods eaten

The second nationwide survey of food consumption is the National Health and Nutrition Examination Survey (NHANES III) (1988-94), conducted by the National Center for Health Statistics in the Center for Disease Control (CDC) of the Department of Health and Human Services. The survey consists of:

• One 24-hour recall of foods eaten

In order to provide estimates of exposure to L. monocytogenes via distributions of food consumption, we initially plan to use data from both the CSFII 94-96 and NHANES III data bases. We will first determine the amount of each food that is eaten (in grams) at an eating occasion, though we will most likely base the final distributions on the amount of the food eaten per person per day.

1. Underreporting of food consumption: We will make the assumption that the data bases are the best and most comprehensive available, and that under reporting will always be an issue without any concrete solution. Survey experts have not determined a factor to use to correct for under reporting.

2. Different weighting factors: Each data base is derived from a probability sample of respondents. All estimates of food consumption should be weighted to represent the US population, and weights differ for each data base. If there is no reasonable method for combining the two data sources, without compromising their integrity, we may need to use data for selected food groups from only one of the data bases. For small sample sizes, we may select data from both sources to provide a reasonable picture of consumption and will need to determine an appropriate method for combining the data.

3. Mixed dishes: It may be necessary to select individual ingredients from mixed dishes, such as sandwiches. For other mixed dishes, we will be able to use the intact food for estimation of consumption (e.g., vegetable salads). All foods do not have corresponding recipes by which we can calculate the proportion of the total product weight accounted for by an individual ingredient; however, others should be more straightforward (e.g., beef in cheeseburgers).

4. Varying sample sizes of food groups. This issue is related to the weighting issue. Some food groups will have a lot of eating occasions, but others will have a small sample. In some instances, we may have to calculate deciles and/or percentiles for very large groups of data and use those reduced data points. For those large data sets, we may need to make the decision to use data from only one data base. In other instances, where the data set is very small, we will have to determine how to merge weighted distributions.

5. Merging food consumption data with data from the L monocytogenes contamination level module. The final food category groupings will be determined after compilation of the contamination level data. In some instances, food consumption data for several foods or groupings may be merged in order to match the available contamination data.

Each of the food consumption data bases contains a food code to identify each food. We have reviewed over 7,200 food codes and have grouped the foods implicated with contamination of L. monocytogenes (outbreaks, sporadic cases, recalls, analytical testing) into food categories. The initial food groupings are listed in Table 1 of the Food Contamination Module, but may be modified based upon levels of L. monocytogenes identified in that module.

Reduction Of Uncertainty

1. Fluid milk: The 1995 and 1996 Behavioral Risk Factor Surveillance Systems (BRFSS) included 12 questions regarding food safety, which were developed by the CDC, FDA, and several state health departments. The set of questions was used to collect data about food-handing, preparation, and consumption behaviors that have been associated with foodborne illness in adults. The survey was administered in the states of Colorado, Florida, Missouri, New York, and Tennessee in 1995 and in Indiana and New Jersey in 1996. Two questions were included on the survey administered in South Dakota. The study included 19,356 completed questionnaires (Yang, 1998).

   Results indicate that, 1.4% of the respondents reported that they had drunk raw milk during the past 12 months. Although federal law requires milk in interstate commerce to be pasteurized, some states allow milk consumed within the state to be sold and drunk as raw. Several studies have shown that L. monocytogenes is frequently present in raw milk, but it is inactivated by standard pasteurization (Bradshaw, 1985).

   In order to estimate the distribution of fluid milk that is consumed, we plan to assume that 1.4% of the amount consumed is unpasteurized milk and use those data to estimate the consumption of fluid milk that is most at risk for L. monocytogenes.

   In addition, we will consider at a later date that L. monocytogenes can be cultured from approximately 5% of raw (unpasteurized) milk samples (CDC, 1988) and possibly include that correction factor.

2. Burgers containing ground beef: The 1995 and 1996 BRFSS indicate that 19.7% of the respondents reported eating pink hamburgers. In order to estimate the distribution of ground beef that is consumed in hamburgers and cheeseburgers, we plan to assume that 19.7% of the amount consumed is undercooked ground beef and use those data to estimate the consumption of ground beef that is most at risk for L. monocytogenes.
3. Cheese: We designed a schema to better describe cheeses that will be considered in the risk assessment (Johnson, 1990a, 1990b, 1990c; 21 CFR 133, Herbst, 1995; Ryser, 1999) Each cheese and associated food code will be linked to five other variables:

1. Type of Cheese
   1. soft
   2. semisoft
   3. hard
   4. processed
2. Category of Cheese
   1. Mandatory milk pasteurization (38% of cheese production)
   2. Utilized in pasteurization process varieties (27%)
   3. Pasteurization optional but frequently used (mild cheddar, colby, blue mold, brick, and limburger) (10%)
   4. Not defined (cold pack/other processed foods/food service) (10%)
   5. All other varieties (edam, gouda, port salut, havarti, brie, camembert, feta) + Mexican cheeses (2%)
   6. Milk heat treatment frequently utilized (sharp cheddar, hard Italian, Swiss) (13%)
3. Pasteurization Required
   1. Yes, pasteurized milk required or product is pasteurized
   2. No, all others
4. Surveillance and epidemiology of L. monocytogenes
   1. Recall(s)
   2. Outbreak(s)
   3. Sporadic Case(s)
5. Risk designation
   1. lower
   2. higher (if associated with a recall, cheeses normally having a lower risk are placed at higher risk)
   3. highest (if associated with sporadic case(s) or outbreak(s), cheeses normally having either a lower or higher risk are placed at the highest risk)

Example:

4.  Juice: The FDA economic analysis used for the juice HACCP regulation estimated the proportion of unpasteurized apple and orange juices consumed to be 1.7%. Because there were no data available to estimate the amount of unpasteurized vegetable juice consumed, they again used 1.7%. In order to estimate the distribution of unpasteurized juices that are consumed, we plan to assume that 1.7% of the fruit and vegetable juices consumed are unpasteurized and use those data to estimate the consumption of juice that is most at risk for L. monocytogenes. Also, it is not apparent that L monocytogenes will survive in citrus juices, although one study shows that acid-adapted cells showed greatly improved survival in low-pH foods (orange juice and salad dressing) (Gahan, 1996).

A. Background, Review Articles, Foods Linked To Listeriosis, Recalls

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Farber, J.M., J.Y. D’Aoust, M. Diotte, A. Sewell, and E. Daley. 1998b. Survival of Listeria spp. on raw whole chickens cooked in microwave ovens. J Food Prot 61(11):1465-1469.

Fenlon, D.R., J. Wilson, and W. Donachie. 1996. The incidence and level of Listeria monocytogenes contamination of food sources at primary production and initial processing. J Appl Bacteriol 81(6):641-650.

Gilbert, R.J., J. McLauchlin, and S.K. Velani. 1993. The contamination of pate by Listeria monocytogenes in England and Wales in 1989 and 1990. Epidemiol Infect 110(3):543-551.

Goulet, V., J. Rocourt, I. Rebiere, C. Jacquet, C. Moyse, P. Dehaumont, G. Salvat, and P. Veit. 1998. Listeriosis outbreak associated with the consumption of rillettes in France in 1993. J Infect Dis 177(1):155-160.

Houang, E., and R. Hurley. 1991. Isolation of Listeria species from precooked chilled foods. J Hosp Infect 19(4):231-238.

Jacquet, C., B. Catimel, R. Brosch, C. Buchrieser, P. Dehaumont, V. Goulet, A. Lepoutre, P. Veit, and J. Rocourt. 1995. Investigations related to the epidemic strain involved in the French listeriosis outbreak in 1992. Appl Environ Microbiol 61(6):2242-2246.

Johnson, J.L., M.P. Doyle, and R.G. Cassens. 1988. Survival of Listeria monocytogenes in ground beef. Int J Food Microbiol 6(3):243-247.

McLauchlin J., S.M. Hall, S.K. Velani, and R.J. Gilbert. 1991. Human listeriosis and pate: a possible association. BMJ 303(6805):773-775.

Morris, I.J., and C.D. Ribeiro. 1991. The occurrence of Listeria species in pate: the Cardiff experience 1989. Epidemiol Infect 107(1):111-117.

Newton, L, S.M. Hall, M. Pelerin, and J. McLauchlin. 1992. Listeriosis surveillance: 1991. Commun Dis Rep CDR Rev 2(12):R142-R144.

Nichols, G., J. McLauchlin, and J. de Louvois. 1998. The contamination of pate with Listeria monocytogenes—results from the 1994 European community-coordinated food control program for England and Wales. J Food Prot 61(10):1299-1304.

Salvat, G., M.T. Toquin, Y. Michel, and P. Colin. 1995. Control of Listeria monocytogenes in the delicatessen industries: the lessons of a listeriosis outbreak in France. Int J Food Microbiol 25(1):75-81.

Schwartz, B., C.A. Ciesielski, C.V. Broome, S. Gaventa, G.R. Brown, B.G. Gellin, A.W. Hightower, and L. Mascola. 1988. Association of sporadic listeriosis with consumption of uncooked hot dogs and undercooked chicken. Lancet 2(8614):779-782.

Trussel, M. 1989. The occurrence of Listeria in the production of processed meat, salami, and mettwurst. Schweiz Arch Tierheilkd 131(7):409-412, 417-421.

Wilson, I.G. 1996. Occurrence of Listeria species in prepacked retail sandwiches. Epidemiol Infect. 117(1):89-93.

Dose-response is the term used to relate the number of ingested organisms to the outcome of the ingestion event (illness, death, etc.) for a given population. Although microbiological dose-response models are in the nascent stages of development, data collected from epidemiological investigations, clinical trials, and animal studies still provide useful insights.

There are no known human clinical trials with Listeria monocytogenes. therefore, epidemiologic investigations provide the only opportunity to directly collect information on the susceptible human populations. The more complete investigations provide information on the pathogen levels in implicated foods and the number and immune status of ill and exposed-asymptomatic persons. Because L. monocytogenes is an intercellular pathogen that is mechanistically similar to other human disease, a large number of animal studies have been conducted for many years. This risk assessment will attempt to utilize the data from these animal studies, as described below, to provide information useful for dose-response modeling.

This Public Health Module will focus primarily on three parameters (or data inputs) to model L. monocytogenes dose-response. The parameters are consistent with the disease triangle that considers the environment (in this case the food matrix), the pathogen (virulence characteristics or factors), and the host (susceptibility or immune status factors). Data to feed into the parameters come from humans (outbreaks, case reports, case-controlled studies), animals (mice, rats, primates, and other species), and in vitro studies.

Listeriosis

L. monocytogenes is a microbial pathogen which has been found in low concentrations world-wide, in soil, water, vegetation, slaughterhouse waste, animal feed, and the gastrointestinal tract of over 50 different animals, including man. However, the ability of the organism to survive and grow in adverse environments and the severity of the illness it causes makes L. monocytogenes a dangerous foodborne pathogen. Thirteen serotypes of L. monocytogenes have been identified, and most human cases of listeriosis are caused by serotypes 1/2a, 1/2b, and 4b. (Ryser and Marth, 1998; Gray and Killinger, 1966; Schuchat et al., 1991)

Pathology
Recent studies have shown that L. monocytogenes is carried in the gut of about 5% of the general population and that the large intestine is the principle reservoir for the organism in humans. L. monocytogenes causes illness by penetrating the lining of the gastrointestinal tract. Once it has invaded the tissue, the organism can protect itself against phagocytosis, grow, and migrate throughout the host. A spectrum of illness severity is observed resulting from exposure to Listeria, ranging from asymptomatic carriers through non-invasive gastrointestinal disease to systemic or invasive conditions. A partial list of systemic illness and disease caused by L. monocytogenes includes bacteremia, bacterial meningitis, conjunctivitis, CNS infection, cutaneous infection, encephalitis, endocarditis, meningoencephalitis, miscarriage, neonatal disease, osteomyelitis, peritonitis, pleural infection, pneumonia, premature birth, prodromal illness in pregnant women, septicemia, and stillbirth. Milder symptoms associated with listeriosis, i.e., diarrhea, fever, headache, and myalgia, are often the result of high doses of L. monocytogenes in otherwise healthy individuals. The mortality rate of the severe form of listeriosis is about 20% of those hospitalized. (Slutsker and Schuchat, 1999; Schuchat et al., 1991; Farber and Peterkin, 1991; Ryser and Marth, 1998)

Epidemiology
Most cases of listeriosis occur sporadically. Mild cases may go unnoticed or unreported and in these cases the cause of the illness is usually not determined. However, there have been some sporadic cases and several large outbreaks where the vehicle of infection was determined or suspected to be food. (Schuchat et al., 1991; Ryser and Marth, 1998; Linnan et al., 1988)

Not everyone exposed to foodborne L. monocytogenes develops listeriosis. Persons who become ill must be susceptible and receive a sufficient dose of the virulent organism. Most human cases of listeriosis occur in persons that have suppressed T-cell-mediated immunity. Those most at risk of infection are pregnant women, neonates, the elderly, and the immunocompromised. The immunocompromised include persons taking immunosuppressive medications, and persons with chronic illness like cancer, diabetes and alcoholism. Healthy adults have a relative low risk of illness from L. monocytogenes. Listeriosis in children, adolescents, and young adults is rare but does occur superimposed upon other disorders. In older patients, listeriosis is usually superimposed on neoplastic disease, diabetes, or other severe illness. It's been suggested that susceptibility is enhanced if the host is simultaneously being infected by another bacteria or virus. (Schwatrz et al., 1989; Gray and Killinger, 1966; Schuchat et al., 1991; Linnan et al., 1988; Salamina et al., 1996; Gellin and Broome, 1989; Farber and Peterkin, 1991)

Virulence

Food Vehicle

This parameter assumes variability in the relationship between the physical/ chemical nature of a Listeria-contaminated food and the fate of the organism following ingestion. This variance would influence the number of organisms required to produce illness. This parameter includes data on the number of Listeria in particular food types associated with disease outbreaks and data on protective or enhancing effects, relative to infection, associated with particular food chemical and physical characteristics.

Susceptibility
This parameter assumes that human subpopulations exhibit varying degrees of susceptibility to listeriosis based on certain intrinsic biological characteristics. The variability in susceptibility would influence the number of organisms required to produce illness and the type of illness produced. Information on this parameter includes epidemiologic and case reports of conditions which predispose to infection, and studies with animal surrogates on the role of host defense components in susceptibility to Listeria infection.

Human studies include information on prevalence and severity of listeriosis in cancer patients, AIDS patients, the elderly and other immunosuppressed populations.

• Listeria infection in healthy adults is usually associated with asymptomatic carriage (Slutsker and Schuchat, 1999).
• Of 148 adult listeriosis cases not involving pregnancy, cancer was an underlying condition in 25% of meningitis cases and 33% of incidences of bacteremia (Nieman and Lorber 1980).
• Of 261 cases not involving pregnancy in Britain (1967-1985), 40% were associated with cancer. Of these, 32% had leukemia, 29% had lymphoma and 39% other cancers. (McLauchlin, 1980).
• In 84 cases from Australia, 39% of nonpregnant adults were over 60 years of age. 75% of these individuals were on prednisone, azathioprine or cyclosporin, and 20% had previously undergone cancer chemotherapy. (Paul et al., 1994)
• A review of 98 cases of non-pregnancy associated sporadic listeriosis in the U.S. revealed that 98% of persons involved had at least one underlying condition, most, but not all, of which were associated with probable immunosuppression (Schuchat et al., 1992).
• Use of antacids and laxatives may possibly increase the risk of listeriosis since Listeria is acid sensitive (Ho et al., 1986).
• In 94 cases of listeriosis reported in Los Angeles County, serotype 4b was more common in perinatal cases and serotype 1/2b in non-perinatal cases (Mascola et al., 1989).

Animal studies include information on morbidity and mortality (including surrogate endpoints), mouse strains with specific immune dysfunctions (knockouts) and pathogenic mechanisms.

• Mice with severe combined immunodeficiency (SCIDS) did not completely clear experimental Listeria infections as compared with normal mice at 12 days of infection , but did not develop a fatal infection ( Bancroft et al., 1986).
• Neutralization of Interleukin 12 (IL-12) or Tumor Necrosis Factor by monoclonal antibodies increases the number of Listeria in spleens of infected SCIDS mice (a strain mimicking increased susceptibility due to immunocompromise) by 1-3 logs (Tripp et al., 1994).
• SCIDS mice maintain a chronic but nonfatal state of infection in the absence of T cells, due to control of the infection by polymorphonuclear leukocytes (PMS) and to IL-12 production by NK cells (Unanue, 1997).
• IL-6 knockout mice (unable to produce Interleukin-6) show decreased resistance to Listeria infection as manifested by 500-fold increase in Listeria CFU/liver and a 10-fold increase in Listeria CFU/spleen. (Dalrymple et al, 1995).
• In mice infected intragastrically with L. monocytogenes and depleted of neutrophils, there was an approximate 4 log decrease in the number of Listeria required for a lethal dose as compared to controls (Czuprynski et al., 1996).

Farber, J. M. and Peterkin, P. I. 1991. Listeria monocytogenes, a foodborne pathogen. Microbiological Reviews, 55(3):476-511.

Gellin, B. G. and Broome, C. V. 1989. Listeriosis. JAMA, 261(9):1313-1320.

Gray, M. L. and Killinger, A. H. 1966. Listeria monocytogenes and listeric infections. Bacteriological Reviews, 30(2):309-382.

Linnan, M. J., Mascola, L., Leu, X. D., Goulet, V., May, S.,Salminen, C., D. Hird, D. W., Yonkura, M. L., Hayes, P., Weaver, R., Audurier, A., Plikaytis, B. D., Fannin, S. L., Kleks, A., and Broome, C. V. 1988. Epidemic listeriosis associated with Mexican-style cheese. N. Engl. J. Med. 319:823-828

Ryser, E. T. and Marth, E. H. 1999. Listeria, Listeriosis, and Food Safety, Second Edition. Marcel Dekker, Inc., New York.

Salamina, G., Dalle Donne, E., Niccolini, A., Poda, G., Cesaroni, D., Bucci, M., Fini, R., Maldini, M., Schuchat, A., Swaminathan, B., Bibb, W., Rocourt, J., Binkin, N., and Salmaso, S. 1996. A foodborne outbreak of gastroenteritis involving Listeria monocytogenes. Epidemiol. Infect., 117: 429-436.

Schuchat, A., Swaminathan, B., and Broome, C. V. 1991. Epidemiology of Human Listeriosis. Clinical Microbiology Reviews, 4(2):169-183.

Schwartz, B., Hexter, D., Broome, C. V., Hightower, A. W., Hirschhorn, R. B., Porter, J. D., Hayes, P. S., Bibb, W. F., Lorber, B., and Faris, D. G. 1989. Investigation of an outbreak of Listeriosis: New hyptothese for the etiology of epidemic Listeria monocytogenes infections. Journal of Infectious Diseases, 159(4):680-685.

Barnes, R., Archer, J., Strack, J., and Istre, G. R. 1989. Listeriosis associated with consumption of turkey franks. MMWR 38:267-68.

Conlan, J. W. and North, R. J. 1992 Early pathogenesis of infection in the liver with the facultative intracellular bacteria Listeria monocytogenes, Francisella tularensis,and Salmonella typhimurium involes lysi. Infection And Immunity 60(12):5164-5171.

Czuprinski, C. J., Theisen, C., and Brown, J. F. 1996. Treatment with the anti-granulocyte monoclonal antibody RB6-8C5 impairs resistance of mice to gastrointestinal infection with Listeria monocytogenes. Infect. Immun. 64:3946-3949.

Dabiri, G. A., Sanger, J. M., Portnoy, D. A., and Southwick, F. S. 1990. Listeria monocytogenes moves rapidly through the host-cell cytoplasm by inducing directional actin assembly. Proc. Natl. Acad. Sci. USA 87:6068-6072

Dalrymple, S. A., et al. 1995. Interleukin-6-deficient mice are highly susceptible to Listeria monocytogenes infection: Correlation with inefficient neutrophilia. Infection and Immunity 63(6):2262-2268.

Dalton, C. B., Austin, C. C., Sobel, J., Hayes, P. S., Bibb, W. F., Graves, L. M., Swaminathan, B., Proctor, M. E. and Griffin, P. M. 1997. An outbreak of gastroenteritis and fever due to Listeria monocytogenes in milk. N. Engl. J. Med. 336(2):100-105.

Farber, J. M. and Peterkin, P. I. 1991. Listeria monocytogenes, a foodborne pathogen. Microbiological Reviews, 55(3):476-511.

Gaillard, J. L., Berche, P., and Sansonetti, P. 1986. Transposon mutagenesis as a tool to study the role of hemolysin in the virulence of Listeria monocytogenes. Infec. Immun. 52:50-55.

Ho, J. L., Shands, K. N., Friedland, G., Eckind, P., and Fraser, D. W. 1986. An outbreak of type 4b Listeria monocytogenes infection involving patients from eight Boston hospitals. Arch. Intern. Med. 146:520-524.

Junttila, J. and Brander, B., 1989. Listeria monocytogenes septicemia associated with consumption of salted mushrooms. Scand. Journal of Infectious diseases, 21, 339-342.

Kocks, C., Gouin, E., Tabouret, M., Berche, P., Ohayon, H., and Cossart, P. 1992. Listeria monocytogenes-induced actin assembly requires the acta gene product, a surface protein. Cell 68:521-531

Kuhn, M. and Goebel, W. 1999. Pathogenesis of Listeria monocytogenes. pp. 97-130 in Ryser, E. T. and Marth, E. H. (eds.) Listeria, Listeriosis, and Food Safety, Second Edition. Marcel Dekker, Inc., New York. Pp. 97-130

Linnan, M. J., Mascola, L., Leu, X. D., Goulet, V., May, S., Salminen, C. D., Hird, D. W., Yonkura, M. L., Hayes, P., Weaver, R., Audurier, A., Plikaytis, B. D., Fannin, S. L., Kleks, A., and Broome, C. V. 1988. Epidemic listeriosis associated with Mexican-style cheese. N. Engl. J. Med. 319:823-828.

Mascola, L., Sorvillo, F., Neal, J., Iwakoshi, K., and Weaver, R. 1989. Surveillance of listeriosis in Los Angeles county, 1985-1986: a first year's report. Arch. Intern. Med., 149, 1569-1572.

McLauchlin, J. S. 1990. Human listeriosis in Britain, 1967-85: A summary of 722 cases: 2. Listeriosis in non-pregnant individuals, a changing pattern of infection and seasonal incidence. Epidemiol. Infect. 104:191-201.

McLauchlin, J., Audurier, A., and Taylor, A. G. 1986. Aspects of the epidemiology of human Listeria monocytogenes infections in Britian 1967-1984; the use of serotyping and phage typing. J. Med. Microbiol. 22:367-3767.

Nieman, R. E. and Lorber, B. 1980. Listeriosis in adults: A changing pattern; Report of eight cases and review of the literature. Reviews of Infectious Diseases 2(2):207-227.

Paul, M. L., Dwyer, D. E., Chow, C., Robson, J., Chambers, I., Eagles, G., and Ackerman, V. 1994. Listeriosis-a review of eighty-four cases. Med. J. Austral. 160:489-493.

Pinner, R. W., Schuchat, A., Swaminathan, B., Hayes, P. S., Deaver, K. A., Weaver, R. E., Plikaytis, B. D., Reeves, M., Broome, C. V., and Wenger, J. D. 1992. The role of foods in sporadic listeriosis: II. Microbiologic and epidemiologic investigation. JAMA 1992;267.2046-2050.

Portnoy, D., Jacks, P. S., and Hinrichs, D. J. 1988. Role of hemolysin for the intracellular growth of Listeria monocytogenes. J. Exp. Med. 167:1459-1471.

Raybourne, R. B. and Bunning, V. K. 1994. Bacteriun-host cell interactions at the cellular level: Fluorescent labeling of bacteria and analysis of short-term bacterium-phagocyte interaction by flow cytometry. Infect. Immun. 62: 665-672.

Riedo, F. X., Pinner, R. W., Tosca, M. L., Cartter, M. L., Graves, L. M., Reeves, M. W., Weaver, R. E., Plikaytis, B. D., and Broome, C.V. 994. A point-source foodborne listeriosis outbreak: documented incubation period and possible mild illness. J. Infect. Dis. 170:693-696.

Roll, J. T., and Czuprynski, C. J. 1990. Hemolysin is required for extraintestinal dissemination of Listeria monocytogenes in intragastrically inoculated mice. Infect Immun. 58:3147-3150.

Ryser, E. T. and Marth, E. H. 1999 Listeria, Listeriosis, and Food Safety, Second Edition. Marcel Dekker, Inc., New York.

Salamina, G., Dalle Donne, E., Niccolini, A., Poda, G., Cesaroni, D., Bucci, M., Fini, R., Maldini, M., Schuchat, A., Swaminathan, B., Bibb, W., Rocourt, J., Binkin, N., and Salmaso, S. 1996. A foodborne outbreak of gastroenteritis involving Listeria monocytogenes. Epidemiol. Infect. 117: 429-436.

Schlech, W. F., Lavigne, P. M., Bortulussi, R. A., Allen, A. C., Haldane, E. V., Wort, A. J., Hightower, A. W., Johnson, S. E., King, S. H., Nicholls, E. S., and Broome, C. V. 1983. Epidemic listeriosis: Evidence for transmission by food. N. Engl. J. Med. 308:203-206.

Schuchat, A., Deaver, K. , Wenger, J. D., Plikaytis, B. D., Mascola, L., Pinner, R. W., Reingold, A. L., Broome, C.V., and the Listeria Study Group. 1992. Role of foods in sporadic listeriosis. JAMA. 267(15):2041-2045.

Schwartz, B., Broome, C. V., Brown, G. R., Hightower, A. W., Ciesielski, C. A., Gaventa, S., Gellin, B. G., Mascola, L., and the Listeriosis Study Group. 1988. Association of sporadic listeriosis with consumption of uncooked hot dogs and undercooked chicken. Lancet 2:779-782.

Slutsker, L. and Schuchat, A. 1999. Listeriosis in humans. pp. 75-95 in Ryser, E. T. and Marth, E. H. (eds.) Listeria, Listeriosis, and Food Safety, Second Edition. Marcel Dekker, Inc., New York.

Tripp, C. S., Wolf, S. F., and Unanue, E. R. 1993. Interleukin 12 and tumor necrosis factor a are costimulators of interferon g production by natural killer cells in severe combined immunodeficiency mice with listeriosis. Proc. Natl. Acad. Sci. 90: 3725-3729.

Unanue, E. R. 1997. Studies in listeriosis show the strong symbiosis between the innate cellular system and the T-cell response. Immunological Reviews 158:11-25.