A Food-borne Illness, at its broadest definition, can encompass any illness that is contracted from the consumption of, or exposure to, food. This could involve exposure to infective, toxic, or even carcinogenic agents that may or may not have a microbiological origin. For the purposes of this article, the definition will largely be restricted to microbiologically derived food-borne illness. However, there will be some discussion of other factors that cause food-borne illness and generally fall under the heading of natural toxins. Typically, when one thinks of a food-borne illness, it is in a patient with acute gastrointestinal signs and symptoms, typified by nausea, vomiting, diarrhea, and abdominal pain.
However, it is important to remember that some of the consequences of foodborne infections are systemic and may occur several days or weeks after the initial exposure to the foodborne pathogen. Examples include the association of Shiga toxin-producing E. coli with hemolytic uremic syndrome, Campylobacter and the development of Guillain-Barré syndrome, and the association of a number of enteric bacterial pathogens with the development of reactive arthritis. A variety of food-borne illnesses will be discussed, including those due to bacterial, viral, and protozoal infections, as well as some natural toxins. Some of the basic diagnostic microbiology will be addressed, as well as the clinical presentation and potential complications from the various infections.
I. FOOD-BORNE ILLNESS EPIDEMIOLOGY
The total burden of food-borne illnesses in the United States and other parts of the world is essentially unknown. Yet, death from food- or water-related exposure to pathogens worldwide is considered to be enormous. Several million children are thought to die each year from diarrheal disease, the majority of which is most likely due to contaminated food or water. In the United States, the number of deaths from food-borne disease is estimated to be between 1000 and 9000 per year. Until recently, these numbers were little more than extrapolations from small epidemiological studies in various parts of the country. The development of FoodNet by the Centers for Disease Control and Prevention has, for the first time, offered an opportunity for us to determine what the epidemiology of food-borne disease is in large numbers of people in the United States. The Food-borne Diseases Active Surveillance Network (FoodNet) is the main food-borne disease component of the CDC’s Emerging Infections Program (EIP) and is a collaborative venture with EIP program sites, the United States Department of Agriculture, and the United States Food and Drug Administration. FoodNet has undertaken population-based active surveillance for confirmed cases of Campylobacter, Escherichia coli O157:H7, Listeria, Salmonella, Shigella, Vibrio, and Yersinia infections. In 1997, they had active surveillance in Minnesota, Oregon, parts of California, Connecticut, and Georgia. The total population under study was 15.9 million. More sites came on-line in 1998 and the plan is to expand even further in the future. During 1997, which is the most recent year in which the data has been tabulated, FoodNet found a total of 8557 confirmed cases of infections caused by the pathogens listed above, as well as a number of Cryptosporidium and Cyclospora infections, as shown in Table 40.1.
There were seasonal variations, with the summer months (June, July, and August) being the peak isolation times for Vibrios (66%), E. coli O157:H7 (52%), Campylobacter (35%) and Salmonella (32%), thus illustrating the need for heightened awareness for food-borne pathogens during the warmer times of the year. Interestingly, there were large variations in the incidence rates of specific pathogens in the different sites. For example, the incidence rates for Campylobacter varied from 14/100,000 in Georgia to 50/100,000 in California. The incidence of S. enteritidis was highest in Connecticut. It is unclear what these geographic differences mean but they suggest that regional variations among food-borne pathogens do occur, and yet there do not appear to be any areas in which the incidence of these major pathogens is so low that they can be dismissed as unimportant. In terms of age and sex distribution of food-borne illnesses, there is a clear preponderance for the young and the elderly. The incidence rates were much higher in the FoodNet study in children under the age of one year, especially in relation to Salmonella infection. Not all food-borne pathogens are equal in terms of the clinical severity of the disease and this is well illustrated from the FoodNet data. Listeria infection was associated with the highest hospitalization rate (88%) and the highest fatality rate (20%). This was followed by E. coli O157:H7 (29%), Salmonella (21%), Yersinia (15%), Shigella (13%), Campylobacter (10%), and Vibrio (10%). FoodNet has offered an opportunity to take a glimpse at the causes of food-borne disease in those patients from whom a sample is obtained for microbiological analysis. It does not, however, address many other food-borne pathogens that were not part of the study. Nor does the FoodNet surveillance address what is going on in those patients who do not have a sample sent for testing. As part of the adjunctive studies reported by CDC, it was determined by phone consultations that, of 10,000 residents within the FoodNet sites, 11% reported a diarrheal illness during the previous month. This translates to 1.4 episodes of diarrhea per year, which, if multiplied roughly by the population of the United States, represents approximately 350 million cases per year. Of greater interest, only 8% of those that were ill sought medical care, and of those 8%, only 20% reported submitting a stool sample for culture. Thus, our best data on the causes of acute gastrointestinal disturbance are based on cultures of _2% of diarrheal episodes. There are still no clear data on the numbers of deaths from food-borne disease in the United States, but original estimates of around 9000 per year are probably too high and the real number may be in the 1000–2000 per year range. However, globally there are still over 3 million children dying each year from diarrheal diseases, many of which are probably related to contaminated food or water. Thus, on a global basis, food-borne illness remains a huge problem.
II. BACTERIAL FOOD-BORNE PATHOGENS
There are many different bacterial pathogens that have been associated with food-borne disease, as shown in Table 40.2. Despite this large number of bacterial pathogens, they generally cause disease by a limited number of mechanisms, such as via a preformed toxin, the production of a toxin in the intestine, invasion of the intestinal epithelial cells, or some other process. The various organisms are broken down into these mechanistic groups and will be described in more detail. A. Bacteria causing disease primarily mediated by a preformed toxin This group of organisms includes bacteria that usually make a toxin in food prior to ingestion of the food. This is in contrast to other toxin-producing bacteria (discussed in a later section), which produce a toxin after they have been ingested. 1. Clostridum botulinum C. botulinum is a gram-positive, spore-forming, toxinproducing, obligate anaerobe. Its natural habitat is soil and, therefore, its spores are frequently present on fresh fruits and vegetables. The disease associated with ingestion of the toxins from C. botulinum is known as botulism. There are seven antigenically distinct types of botulinus toxin, each of which is designated by a letter (A–G). Types A, B, E, F, and G are the ones that are associated with human disease, with type A accounting for about 25% of outbreaks and type B 8%, although some regionality is seen with the various types within the United States. Botulinus toxin is heat-labile (5 minutes of boiling will destroy the toxin) and is produced at any temperature in which bacterial growth occurs. The majority of outbreaks have been traced to home-produced foods, especially vegetables, fish, fruits, and condiments. Commercial food sources are occasionally incriminated. The key to preventing this disease is to destroy the C. botulinum spores. Food needs to be subjected to a temperature of 120_ for 30 minutes, usually with the aid of a pressure cooker in the home environment, to destroy spores. Any surviving spores may germinate in the anaerobic environment of the food and produce their toxins. Botulinus toxin is absorbed from the proximal intestine and spreads via the bloodstream to the peripheral cholinergic nerve synapses. The toxin then irreversibly blocks acetylcholine release, resulting in flaccid paralysis. The toxin does not cross the blood–brain barrier. Typically, symptoms occur 18–36hr after toxin ingestion, but incubation may be as short as a few hours or as long as 8 days. Cranial nerves are usually affected first but, subsequently, respiratory muscles may be affected, resulting in respiratory paralysis and death if left untreated. This food-borne infection does not typically cause any major gastrointestinal symptoms. Diagnostic confirmation is dependent on detection of the toxin or the organism in the patient or the implicated food. Samples, such as the food, vomitus, serum, gastrointestinal washings, and feces, are all reasonable specimens to test for the toxin. The toxin can be detected in specialist centers, using the mouse neutralization test. The organisms themselves or the spores can also be cultured, but this takes longer. Molecular methods, based on polymerase chain reaction (PCR) for the neurotoxin genes, are also available on a research basis. In terms of therapy, there are several approaches one can take. If the diagnosis is made quickly, emetics or gastric lavage may be helpful to remove unabsorbed toxin. A trivalent (ABE) horse serum antitoxin is available and should be used. However, supportive therapy is the mainstay of treatment. Botulism carries a significant mortality, with up to 25% of patients affected by the type A toxin dying. The longer the incubation period, the better the prognosis, since the shorter incubation periods are generally associated with a greater initial toxin load. Eventually, if patients survive the acute symptoms, the prognosis is good and most patients recover completely.
2. Staphylococcus aureus S. aureus has the capacity to produce a variety of toxins (A, B, C1–C3, D, E) that are all small proteins with similar biological activities. Other Staphylococcus species, such as S. hyicus and S. intermedius, as well as coagulase negative species, have, on rare occasion, been found to produce the enterotoxins also. Strains that produce type A toxin, either alone or in combination with type D, are the ones most frequently associated with outbreaks. It is these toxins that are responsible for the disease manifestations and ingestion of as little as 100–200 ng is considered to be enough to cause disease in humans. Typically, patients become symptomatic within 6 hr of ingesting these toxins, with nausea (73–90%), vomiting (82%), and abdominal cramps (64–74%). Diarrhea occurs in a significant proportion of patients (41–88%), but fever is rare. This disease is often associated with outbreaks that may be very large, with over 1000 individuals involved. Anumber of different foods have been associated with staphylococcal food poisoning, especially dairy-based products, such as cream-filled desserts and cakes. Other foods that are often incriminated include egg products, meat products, poultry, mayonnaise, and tuna. Typically, the course of events is that a foodhandler contaminates the food product with S. aureus from their fingers or nose. The organisms then grow in the food if it is held at room temperature for any length of time. Thus, disease is not often associated with commercially prepared food but is more frequently associated with food handled in a food-service establishment or in the home. The bacteria produce the toxin as they grow and bacterial levels in the region of 105 per gram will produce enough toxin in food to cause disease. The toxin is heat-stable and is not inactivated by heating or even boiling the food product. The toxin is also considered to be stable against pH extremes, proteases, and radiation. So, once formed in food, these toxins are almost impossible to remove. The pathogenetic mechanisms whereby these toxins cause their effects are not fully understood, but they are thought to act via stimulation of the autonomic nervous system.
The inflammatory response is also thought to play a role and elevation of certain cytokines may be a critical factor. S. aureus enterotoxins are also classified as superantigens and are potent T-cell activators. The toxins are not absorbed systemically; therefore, protective immunity is not induced following exposure to them. Culturing the implicated food, stools, or vomitus can make a definitive diagnosis. Phage typing is considered to be a useful epidemiological tool, with type III being the one most often associated with outbreaks. Latex agglutination and enzyme immunoassays are available for the direct detection of S. aureus enterotoxins in food in the ng/g food range. Treatment is supportive and dehydration may be a significant problem. Symptoms usually abate within 8 hr and there is no need to treat patients with antibiotics directed toward the S. aureus strain. The outcome is usually good. 3. Bacillus cereus B. cereus is a gram-positive, spore-forming aerobe that causes two distinct clinical syndromes. One is a short incubation period emetic syndrome and the other a longer incubation period diarrheal syndrome. The emetic syndrome has an incubation period and clinical presentation much like S. aureus-related foodborne disease, with nausea and vomiting being the principal features, occurring in a 1–6 hr incubation period following exposure. Occasionally, very short incubation periods (15–30 min) or longer ones (6–12 hr) have been reported. Fever is not associated with this disease and recovery is usual, although there has been at least one report of acute hepatic necrosis associated with exposure to B. cereus emetic toxin. The emetic toxin is a preformed toxin (5–10 kDa) that is typically produced when the organism is present in starchy foods—especially rice. The toxin is heat-stable and resists proteases and variations in pH. So reheating food in which the toxin has formed will not ensure its destruction and may lead to a false sense of security. B. cereus is present in soil and water and in most raw foods; even 10–40% of humans are colonized with this bacteria.
Diagnosis is dependent on finding the organism in the food or vomitus of the patient. The frequency of asymptomatic carriage in the stool makes stool cultures alone unhelpful. Heating food may destroy the organisms, thus making culture confirmation difficult, but, as already mentioned, heat will not destroy the toxin. The toxin associated with the diarrheal syndrome is heat-labile and trypsinsensitive and is thought to be made up of three components (designated B, L1, and L2, for proposed binding and cell lysis properties). There have also been hemolytic activities described in proteins from B. cereus and this may be part of the activity of the three-component toxin. Like the emetic toxin, the diagnosis requires culture confirmation, preferably from the incriminated food source. Recovery is usual and therapy is supportive. B. Bacteria causing disease by production of toxins within the intestine This group of organisms generally causes disease through the production of a toxin following intestinal colonization. Some of the bacteria in this group have other virulence attributes as well, and the disease process may be a combination of the toxin and some of these other factors. 1. Vibrio species Currently, there are over 30 Vibrio species, of which about one-third are known to cause disease in humans. Vibrios are gram-negative facultative aerobes and, in this section, a couple of the more important ones will be discussed. a. Vibrio cholerae O1 V. cholerae O1 is a gram-negative organism that is divided into two biotypes: classical and E1 Tor. There are various ways to differentiate between the two but they were initially differentiated following the discovery that the E1 Tor strains make hemolysins. From a food-borne illness perspective, the main factor that is involved in the spread of V. cholerae is contaminated water. There have been six previous pandemics of cholera and we are currently in the seventh, which began in 1961. Although previous pandemics have been due to classical biotypes, the current pandemic is due to E1 Tor. V. cholerae is a facultative anaerobe and thiosulfate–citrate bile salts–sucrose (TCBS) agar is frequently used for isolation because it inhibits the growth of most other enteric organisms.
V. cholerae is spread via contaminated water and food and cleaning up water supplies can have a major impact on the prevalence of this disease. Its survival in foods is affected by factors such as temperature, pH, and inoculum size, but it may survive for a couple of days on contaminated food. Foods such as seafood, pickled or dried fish, coconut milk, lettuce, and rice have all been associated with carriage of the organism. Clinically, the incubation period is usually 1–3 days but may be as short as a few hours or as long as 5 days. The infectious dose is around 106 organisms, but decreased gastric acidity appears to predispose individuals to infection with V. cholerae. The organisms principal virulence factor is the production of cholera toxin that is an activator of cyclic AMP in intestinal epithelial cells. This results in profound intestinal secretion of fluid, profuse watery diarrhea, and, if untreated, death secondary to dehydration. This is known clinically as cholera. The organisms can be cultured from stools of infected patients, but treatment in the form of rehydration, which can be life-saving, should not wait for culture confirmation. Antibiotics are not required but may shorten the duration of illness. b. Non-O1 V. cholerae In recent years, there has been clinical disease, predominantly in the Indian subcontinent, associated with V. cholerae O139. This is the first time a non- O1 strain has been associated with epidemic disease. It appears to be spread and to cause disease via the production of cholera toxin in the same way as do the O1 strains. Other non-O1 V. cholerae strains have also been associated with food-borne disease—especially in relation to consumption of raw oysters—but such disease is usually sporadic in nature. c. Vibrio parahemolyticus This organism requires NaCl for growth, and, like the other members of the Vibrio species, lives in water. It is frequently found in shellfish and consumption of shellfish is usually associated with clinical disease. In the past, this has been a major food-borne pathogen in Japan, but is less common in the United States. Clinically, symptoms may appear in as little as 4 hr, but are typically present 12–24 hr after exposure. Diarrhea, abdominal cramps, nausea, and vomiting are the principal features. Fever and chills may occur in about 25% of cases and there have been reports of a dysentery-like diarrhea associated with this organism. The major factor produced by V. parahemolyticus that is considered to be important in pathogenesis is a 23 kDa protein called thermostable direct hemolysin (TDH). Isolation from stool can be undertaken with TCBS agar and V. parahemolyticus is differentiated from V. cholerae by the fact that it will not grow on 0% NaCl. The disease is usually self-limiting and attention to fluid replacement is important. Antibiotics may be required if the intestinal symptoms become persistent. d. Vibrio vulnificus Like V. parahemolyticus, V. vulnificus is a free-living estuarine organism that is frequently isolated from shellfish. The majority of illnesses related to this organism are thought to be acquired by eating raw oysters. Individuals who are alcoholic or who have underlying illnesses, such as liver disease or hemochromatosis, are especially susceptible. The consumption of even small amounts of alcohol may increase the risk of becoming infected with this bacterium. V. vulnificus is the most common cause of serious Vibrio-related disease in the United States and, typically, presents as a bacteremia, bullous skin lesions, and hypotension.
The mortality rate in those with bacteremia is around 50% and rises to 90% in those with hypotension. Gastrointestinal symptoms may occur in about 25% of patients. Fully virulent V. vulnificus is encapsulated, and these encapsulated organisms are resistant to the bactericidal activity of normal human serum. They are also very sensitive to the amount of transferrin-bound iron in the host, which is almost certainly linked to the increased susceptibility in patients with hemochromatosis. A definitive diagnosis requires culture of the organism from blood or, occasionally, from bullous fluid. Blood agar or other nonselective media can be used for isolation from blood. The organism may also be isolated from stool in an infected patient, using TCBS agar. In view of the severity and the high mortality, it is important to initiate antibiotic therapy as early as possible; thus, a rapid clinical diagnosis may be life-saving. V. vulnificus is susceptible to a variety of antimicrobials, including tetracycline, ciprofloxacin, trimethoprim–sulfamethoxazle, chloramphenicol, and ampicillin. e. Other vibrios A number of other members of the Vibrio species have been associated with food-borne disease, including V. hollisae, V. furnissii, V. fluvialis, and V. mimicus. These share similar epidemiology with the Vibrios already discussed, and, typically, cause gastroenteritis. Some of these other Vibrios have occasionally been associated with bloody diarrhea. Biochemical differentiation is the best way to tell them apart. 2. Clostridum perfringens C. perfringens is an anaerobic, spore-forming, gram-positive rod that is associated with at least two types of food-borne disease. Of the various types of C. perfringens, type Ais the one predominantly associated with food-borne disease. Type Aresults in a noninflammatory diarrhea that is usually linked to the consumption of meat or poultry (typically, foods that are high in protein) that has been prepared and allowed to remain between 15_ and 60 _C for more than 2 hrs. During this period, clostridial spores germinate and begin vegetative growth. When around 105 vegetative cells are ingested, they are present in large enough numbers to transiently colonize portions of the intestine and produce enough toxin to cause disease. Ingestion of preformed toxin or nongerminated spores will not usually result in disease. C. perfringens enterotoxin (CPE) is a heatlabile 35-kDa protein encoded by the cpe gene. In fact, C. perfringens types A, C, and D all have this gene but, for some reason that is not fully understood, only type A is frequently associated with food-borne diseases. CPE has a complex mechanism of action and appears to insert itself into the host cell membrane to form a protein complex. This results in membrane permeability changes, loss of intracellular potassium, and other intracellular disruptions.
Clinically, typical symptoms include diarrhea and severe abdominal cramps developing 6–14hrs after exposure; vomiting and fever are less common. Making a definitive diagnosis is complicated, in view of the fact that C. perfringens is found in the normal bowel flora of many individuals. However, a number of tests have been developed to detect the toxins in stool, using either enzyme immunoassays or latex agglutination. Ideally, one should attempt to recover large numbers (_105/g) of organisms from the suspected food. C. perfringens type C is also a food-borne cause of disease but the clinical picture is very different from type A disease discussed above. C. perfringens type C causes a necrotizing enterocolitis that is seen in the context of poor nutrition and so is almost exclusively a disease of developing countries. The type C strains produce three toxins (an enterotoxin and an _ and _ toxin). It is the _ toxin that appears to be primarily responsible for the necrosis seen following infection with type C strains. _ toxin is usually inactivated by proteolytic enzymes in the intestine and so the disease is associated with conditions in which the enzymes are inadequate (e.g., in malnutrition) or in the presence of trypsin inhibitors, such as undercooked pork or sweet potatoes. 3. Shiga toxin-producing E. coli Shiga toxin-producing E. coli (STEC) are relative newcomers to the scene of food-borne pathogens. The first STEC to be associated with disease in humans was E. coli O157:H7, following two outbreaks of hemorrhagic colitis in 1982. Since then, it has been learned that there are, in fact, many different serotypes of STEC and at least 60 different types have been associated with clinical disease. Recent studies have suggested that around 1% of samples submitted to clinical microbiology laboratories in the United States contain STEC, of which around two-thirds are O157:H7, the remainder being non-O157. STEC are present in the gastrointestinal tracts of many mammalian species but appear to be especially common in ruminants (cattle, sheep, and goats). Therefore, the main source of STEC in our food supply is from bovine products. Recently, there have been an increasing number of reports associating STEC infection with fresh produce (lettuce, alfalfa sprouts, apple cider) and water. This is thought to be mainly due to contamination with fecal material from cattle pasture. Clinically, STEC cause a variety of diseases, ranging from diarrhea, which may or may not be bloody, hemorrhagic colitis, and the development of hemolytic uremic syndrome (HUS). HUS is a triad of renal failure, thrombocytopenia, and hemolytic anemia. Acutely, HUS has a mortality rate around 5% and up to 50% of HUS patients may have some degree of permanent renal insufficiency. The main virulence factor from STEC is the production of one or more bacteriophage-encoded Shiga toxins (Stx). Shiga toxins are of two main types, Stx1 and Stx2, and there are at least 4 subtypes of Stx2 (Stx2, 2c, 2d, and 2e). All members of the Stx family have been associated with human disease, although Stx2e mainly causes disease in pigs. Following ingestion of the bacteria, they colonize portions of the lower intestinal tract and produce the toxins. Stx is then thought to cross the intestinal epithelial cell barrier and damage distant target sites, especially the kidney and brain, by a direct effect on endothelial cells in the microvasculature. The infectious dose of STEC may be very low, in the region of 10–100 organisms, in some instances. Symptoms typically develop 2–4 days following ingestion but may occur in as little as 1 day or as long as 8 days. The diarrhea can be of variable type (bloody or nonbloody) and may contain leukocytes.
The type of diarrhea is not a reliable indication of who will go to develop HUS. E. coli O157:H7 has the unique biochemical characteristic of being a slow fermenter of sorbitol. This has been utilized as a means to diagnose O157:H7, using sorbitol MacConkey agar (SMAC). The disadvantage of this is that the other non-O157 STEC usually ferment sorbitol and so will be missed. An alternative strategy is to detect STEC in stool by looking either for Shiga toxins themselves directly or for the presence of Shiga toxin in broth cultures of stool samples. A positive result indicates the presence of STEC in the original sample. Enzyme immunoassays are now available to detect Shiga toxins under both conditions. The mainstay of treatment for STEC and its major complications is supportive. There is a degree of controversy over the use of antibiotics and a number of studies have suggested that certain antimicrobials (e.g., trimethoprim– sulfamethoxazole) actually increase the likelihood that a patient will go on to develop serious complications. In view of this, routine antimicrobial testing of STEC isolates is not recommended. 4. Enterotoxigenic E. coli Enterotoxigenic E. coli (known as ETEC) are a common cause of disease in developing countries and are frequently associated with travelers’ diarrhea. Like many other E. coli strains, they are transmitted by the consumption of contaminated water and food. It is not clear how frequently ETEC are a cause of sporadic disease in the United States because they are not routinely looked for. However, they have caused a number of large outbreaks due to the consumption of contaminated food. Incubation periods range from 12 hr to 2 days and, typically, symptoms are diarrheal (usually watery and without blood) with abdominal discomfort; fever is unusual. The diarrhea is not inflammatory and the illness is usually self-limiting within a few days. However, persistent diarrhea (defined as greater than 14 days) has been associated with ETEC infection. ETEC have two significant virulence characteristics: the ability to colonize the intestine and the capacity to produce toxins. A variety of colonization factor antigens (CFA) have been found in ETEC and at least four major groups CFAI–IV are described with several subgroups. ETEC produce two different types of toxin known as heat-stable toxins (ST) and heat-labile toxins (LT). The ST group is further subdivided into two major types and they have the characteristics of being small peptides that mediate their effects by increasing the intracellular concentration of cyclic GMP. The LT toxins are also part of a family of toxins but are structurally and functionally much like cholera toxin from V. Cholerae.
The diagnosis of ETEC is usually clinical because ETEC do not have unique microbiological or biochemical properties to differentiate them from other E. coli strains. While a number of E. coli O serogroups are more frequently associated with STEC, this is not an absolute association. Detection of the toxin genes, using PCR or DNA probes, is effective but impractical for routine diagnostics. A variety of immunological assays, using latex bead agglutination or reversed passive latex agglutination, have been described. Oral rehydration is the mainstay of treatment of ETEC and may be life-saving. Antibiotic therapy is not routinely required. C. Bacteria causing disease primarily by invading the intestinal epithelial cells This group of food-borne bacteria causes disease by a variety of mechanisms, with the common theme that invasion of the intestinal epithelial cell barrier is part of their pathogenic process. Some of them also produce toxins that may contribute to the disease process, but toxin production is not necessarily the primary mechanism by which this group of bacteria makes people sick. 1. Salmonella spp. Salmonella are one of the most common causes of food-borne illness in humans. There are many types of Salmonella but they can be divided into two broad categories: those that cause typhoid and those that do not. The typhoidal Salmonella, such as S. typhi and S. paratyphi, only colonize humans and are usually acquired by the consumption of food or water contaminated with human fecal material. The much broader group of nontyphoidal Salmonella are found in the intestines of other mammals and, therefore, are acquired from the consumption of food or water that has been contaminated with fecal material from a wide variety of animals and poultry. With a few exceptions (S. typhi and paratyphi C, which express a polysaccharide capsule called Vi), Salmonella are nonencapsulated gram-negative motile rods. They ferment glucose, not lactose, and generally produce H2S, that is seen as a black pigment in triple sugar–iron agar. Their somatic (O) antigens and flagella (H) antigens differentiate the more then 2300 Salmonella. Many of these different isolates have been named after the towns or by the individuals who first discovered them. Using newer DNA-based classification systems, the number of species has been reduced to two: S. bongori, containing nonhuman organisms, and S. enterica, consisting of six groups, including human pathogenic strains (Table 40.3). S. typhi and S. paratyphi are the causes of typhoid fever that continues to be a global health problem but is less of a problem in North America. Having said that, the most likely explanation for cases of typhoid in the United States are either from acquisition overseas or from food that has been contaminated by a chronic carrier. In contrast, the number of cases of nontyphoidal Salmonella has increased steadily over the last four decades. S. entertidis, particularly, has become a growing problem, especially in hen eggs. Currently, the estimate is that around 1 in 10,000 eggs is contaminated with Salmonella. It is now known that Salmonella can penetrate intact eggs lying in fecally contaminated material, and can also infect eggs transovarially during egg development, before the shell is formed.
Other than eggs, common sources of nontyphoidal salmonellosis are milk, foods containing raw eggs, meat and poultry, and fresh produce. Essentially, as with many of the other food-borne bacterial infections, Salmonellae are frequently transmitted through fecal contamination of food because of the large numbers of animals that carry the organism. The infectious dose of S. typhi is thought to be around 105 organisms. The infective dose of nontyphoidal Salmonella may vary from _100 to 106, depending on the host and on the actual type of Salmonella. Irrespective of the type of Salmonella, the most critical virulence determinant of these bacteria is their ability to invade the intestinal epithelium, following which they interact with underlying lymphoid tissue. Avariety of bacterial genes are involved in the invasion process, and much has now been determined about the ways in which various Salmonella proteins then subvert the intestinal epithelial cell biology, discussion of which is beyond the scope of this article. Clinically, Salmonella can cause a multitude of symptoms. Typhoid fever typically causes prolonged high fever, associated with bacteremia and, if untreated, a mortality rate as high as 30%. The incubation period is usually around 5–21 days, shorter if the inoculum is high and the patient is high risk. The disease may begin with gastroenteritis, which often will disappear by the time the fever begins—patients may even be constipated. If left untreated, intestinal perforation and hemorrhage are a concern. Health providers in developed countries are much more familiar with the clinical presentation of nontyphoidal Salmonella. This typically presents with gastroenteritis 24–48 hr after exposure to the organisms. There is usually nausea, vomiting, abdominal cramps, and diarrhea, which may be watery or, occasionally, bloody. It is not unusual for there to be associated symptoms of fever, chills, headache, and myalgia. Occasionally, there can be long-term consequences following Salmonella infection, such as reactive arthritis (especially in individuals who are HLA B27 positive), endocarditis, and localized infections, such as osteomyelitis, septic arthritis, and soft tissue infections. Diagnosing infection with Salmonella is dependent on culturing the organism, usually from either stool or blood cultures. In the case of nontyphoidal Salmonella, it is also worth trying to culture organisms from the incriminated food. Serological tests, such as the Widal test that measures agglutinating antibody titers to the O-antigen, are also available. They are simple and may be helpful in the diagnosis of typhoid fever, but are usually not specific or rapid enough to be of major value.
Microbiologically, most laboratories use MacConkey media (the vast majority of Salmonella do not ferment lactose but about 1% do), deoxycholate agar, salmonella shigella (SS), or Hektoen agar. Following primary isolation, either commercial identification systems are used or a screening media, such as triple sugar iron and lysine iron agar. In terms of therapy, gastroenteritis secondary to nontyphoidal Salmonella is usually self-limiting and rehydration is the most critical aspect of treatment. Antibiotic therapy is not routinely required for this aspect of Salmonella infection and has, in some instances, been thought to promote chronic carriage. When there is systemic invasion with the bacteria and in cases of enteric fever, antibiotic therapy is important. Third-generation cephalosporins and quinolones are most frequently used, although chloramphenicol has been the mainstay of treatment for typhoid fever for many years and is still used in many developing countries, but chloramphenicol does carry a risk of complications. Antibiotic resistance has become a major problem with many Salmonella serovars. Of particular concern is the recent emergence and spread of Salmonella DT104 that carries resistance to multiple antibiotics, including, in some instances, to the fluoroquinolones, such as ciprofloxacin. 2. Campylobacter spp. Food-borne disease due to Campylobacter, which was not recognized until the mid-1970s, is now one of the most frequently diagnosed causes of food-related gastroenteritis. In fact, Campylobacter and Salmonella probably account for 70% of diagnosed cases of bacterial food-borne disease in the United States. Campylobacters are gram-negative, spiral organisms. They are strictly microaerophilic and need oxygen for growth but the level of oxygen in air is too high for them, in view of their sensitivity to oxygen free radicals. An atmosphere of 5–10% O2 and 1–10% CO2 is ideal for their growth, which is better at 42–43 _C and gives additional selectivity for Campylobacter. There are two main species of Campylobacter responsible for most of the illness seen in humans. C. jejuni accounts for the vast majority (~90%) and C. coli accounts for the majority of the remainder. Other Campylobacter species that have been associated with gastroenteritis in humans include C. fetus, C. upsaliensis, C. hyointestinalis, and C. lari. Campylobacter are fragile organisms and tend to die in transport media; therefore, rapid plating of sample is an advantage. Enriched media is usually used that contains a variety of antibiotics to reduce competitive flora.
Campylobacter isolates can be typed using biotyping methods. Serotyping is probably the most widely used method currently and is based on the somatic lipopolysaccharide antigens or the heatlabile surface protein antigens. The infectious dose of Campylobacter can be very wide. Low numbers of Campylobacter may be all that is needed to cause disease and, in some instances in human volunteers, less than 100 organisms can cause disease. These organisms are more frequently associated with sporadic disease rather than outbreaks and person-to-person spread does not appear to be common, although it can occur in a family environment, for example. This sporadic epidemiology suggests that the infectious dose may actually be higher in many situations. C. jejuni and C. coli are commensals in the intestines of many animals and birds, including domestic pets. The main vehicles for infection are raw meats, especially poultry, milk, and water. Surface water is frequently contaminated with campylobacters and water-borne outbreaks have occurred. The pathogenesis of Campylobacter is dependent on its motility. In vitro, nonmotile strains are not capable of invading intestinal epithelial cells. It is assumed that invasion in the intestine is one of the principal pathogenic mechanisms of these bacteria, although the genes involved are not known. However, following infection with campylobacters there is typically an inflammatory response, with marked inflammatory infiltration of the lamina propria, resulting in leukocytes being present in the stool. Clinically, symptoms usually occur within 2–3 days following exposure; this may be as long as 7 days after exposure. Nongastrointestinal symptoms, such as fever (which may be high), headache, and myalgia may precede the onset of nausea, vomiting, and diarrhea. Diarrhea is usually the predominant symptom and it may be watery or bloody and is usually associated with severe cramping abdominal pain. Interestingly, the disease is occasionally biphasic, with an apparent settling of symptoms after 4–5 days, only to be followed by a recrudescence. A number of complications are associated with Campylobacter, including cholecystitis, hepatitis, acute appendicitis, pancreatitis, and focal extraintestinal infections. Longer-term complications include reactive arthritis, Reiter’s syndrome, uveitis, and Guillain-Barré syndrome; molecular mimicry may be involved in some of these complications. A definitive diagnosis requires culture using selective media as previously outlined; however, newer methodologies, such as enzyme immunoassays, are now available and are designed to detect specific campylobacter antigens. Such enzyme immunoassays have the advantage of being able to detect dead organisms, if the transport to the laboratory has been delayed. However, they do not allow typing or assessment of antibiotic sensitivity. The majority of infections are self-limiting and require supportive therapy only, especially in the form of rehydration. Antibiotic therapy is not routinely required; however, patients with prolonged or severe symptoms or who have other significant risk factors (AIDS, cirrhosis, diabetes, etc.) should be treated. Erythromycin is a reliable therapy, although thee is a general move toward using quinolone antibiotics. Despite the use of erythromycin for many years, the resistance levels have remained low. This is not the case with the quinolones and there are increasing reports of ciprofloxacin resistance, which may become more of a problem, in view of the expanded use of these antibiotics in agriculture. 3. Yersinia spp. Of the three members of the genus Yersinia, Y. enterocolitica, and Y. pseudotubeculosis are considered to be food-borne; Y. pestis is not. Y. enterocolitica and Y. pseudotuberculosis are nonlactose fermenting gramnegative organisms that are distinguished by biochemical and serological criteria. Y. enterocolitica is divided into biogroups according to biochemical properties. Serotyping based on the more than 50 described O-antigens is used to designate strains. Most human isolates are of serotypes O3, O8, or O9, although there are geographic variations in the serotype prevalence. Six serotypes and 4 subtypes of Y. pseudotuberculosis have been described. Serotype O1 is associated with about 80% of human cases. Yersinia are not very commonly found as causes of food-borne illness, compared with Salmonella or Campylobacter. However, they are clearly transmitted in food and can cause significant gastrointestinal illness.
The food most frequently associated with Yersiniosis is pork. Swine are a major reservoir of these organisms, and, although they have been found in many other animals (e.g., sheep, dogs, cats, cattle) consumption of undercooked pork is a common association. Milk is another frequently reported source and since Y. enterocolitica can survive and, indeed, multiply in milk at 4 _C small numbers of organisms can become a significant health threat, even if the milk is refrigerated. Infection with Y. pseudotuberculosis has been associated with consumption of contaminated water or unpasterurized milk. Clinically, Y. enterocolitica may cause a variety of symptoms. A gastrointestinal presentation is the most common with diarrhea, low-grade fever, and abdominal pain—usually in children. In about 25% of patients, the stools are bloody and leukocytes are present in about half the patients. Y. enterocolitica is a cause of “pseudo-appendicitis,” in which patients have fever, abdominal pain, and a leukocytosis. Such patients frequently end up having a laparotomy, in which the appendix is typically normal but there is inflammation in the terminal ileum. Y. pseudotuberculosis typically causes an acute mesenteric lymphadenitis, that also presents much like appendicitis. Symptoms in Y. enterocolitica infection can be prolonged, lasting several weeks or even longer. Most infections are, however, self-limiting, although complications may occur, such as ulceration and intestinal perforation. A classic long-term complication following yersiniosis is the development of reactive arthritis. As with other enteric pathogens, this is more likely in patients who are HLA B27 positive. Y. enterocolitica are invasive organisms and at least three bacterial proteins (Invasin, Ail (attachment invasion locus), and YadA) are involved in this process. Not all Y. enterocolitica strains are equally pathogenic and this may be related to levels of expression of some of the various virulence factors. Following invasion across the intestinal epithelial cell barrier (which is likely occurring via M cells), the bacteria localize in the Peyer’s patches and mesenteric lymph nodes. Y. enterocolitica may be isolated from stool or other body tissue on commonly used enteric media, such as MacConkey agar, on which it appears as lactose-negative colonies after 48 hrs of growth at 25–28 _C. Cold enrichment and selective agars have also been used but they may result in isolation of nonpathogenic strains. A variety of tests can be used to determine if a strain is pathogenic, including congo red absorption, salicin fermentation, and esculin hydrolis. Uncomplicated cases will settle spontaneously and antibiotic therapy is not routinely required. Many antimicrobials are effective, but ceftriaxone or fluoroquinolones are recommended for serious infection. 4. Listeria monocytogenes L. monocytogenes is a gram-positive motile rod that is one of the most frightening food-borne pathogens because of the high mortality rate associated with infection. Of the seven Listeria species, only L. monocytogenes is pathogenic for humans.
L. monocytogenes is a common environmental organism and is frequently present in soil and water, on plants, and in the intestinal tracts of many animals. It is has been found in 37 different types of mammals and at least 17 species of birds. Between 1% and 10% of people are carriers of L. monocytogenes. This organism is associated with both sporadic disease and outbreaks. Incriminated foods include milk, cheese, raw vegetables, undercooked meat, and foods prepared for instant use, such as hot dogs. The infectious dose is not really known; some studies have suggested that it may be very high (up to 109 organisms) and others that it may be as low as several hundred. In practice, the most critical aspect is probably individual susceptibility, rather than the infectious dose per se. Clinically, it usually begins with nonspecific symptoms, such as fever, myalgia, and gastrointestinal upset in the form of diarrhea and nausea. What makes L. monocytogenes exceptional as a foodborne pathogen is its very high mortality rate. Of the around 1800 cases per year that are estimated to occur in the United States, there are over 400 deaths. This gives a case fatality rate of over 20%. There are certain groups that are especially at risk of developing listeriosis and these include pregnant women, the elderly, and the immunocompromised. Transplacental transmission in pregnant women is a major concern, although it does not inevitably lead to major consequences. Spontaneous abortion, prematurity, neonatal sepsis, and meningitis are all complications of transplacental transmission. Although L. monocytogenes is readily killed by heat and cooking, the fact that it is so ubiquitous makes recontamination a real risk. This then poses a major health problem, because the organism will grow and multiply at standard refrigerator temperatures. Thus, even minor contamination of a product may, after storage, result in high levels of bacteria, even if the product has been adequately refrigerated. L. monocytogenes grows well on blood or other nutrient agar; it can be separated from other bacteria by growing at cold temperatures (cold enrichment). Selective media may also be used and is better than cold enrichment.
Following diagnosis, L. monocytogenes is readily treated by penicillins or aminoglycosides. 5. Shigella spp. There are four species of Shigella (S. dysenteriae, S. felxneri, S. boydii, and S. sonne). Group-specific polysaccharide antigens of LPS, biochemical properties, and phage or colicin susceptibility differentiate them. All closely resemble E. coli at the genetic level and will grow readily on many routine laboratory media, but selective media are usually used. MacConkey agar, deoxycholate or eosin–methylene blue, and salmonella– shigella media are all adequate, but xylose lysine– deoxycholate (XLD) or Hektoen agars are better for the recovery of Shigella. Shigella is typically spread via the fecal–oral route, through contamination of food or water, or by person-to-person spread. They are highly host-adapted and infect only humans and some nonhuman primates. Therefore, when food becomes contaminated with Shigella, it usually originates from some other infected human. The low infectious dose (of the order of 10–100 organisms, in some cases) makes person-to-person spread a real problem and has resulted in large outbreaks in institutions. Shigellosis is a major problem in developing countries and results in significant morbidity and mortality. Clinically, the disease usually begins with fever, fatigue, anorexia, and malaise. Soon thereafter, watery diarrhea develops, which may or may not become bloody with the development of dysentery in hours or days. The classic dysenteric stool consists of small amounts of blood and mucus, sometimes grossly purulent, and is typically associated with severe abdominal cramps and tenesmus. The severity of the disease varies with the type of Shigella, S. dysenteriae being the worst, followed by S. flexneri and S. sonne or S. boydii—the latter is unusual in most settings. S. dysenteriae type 1 carries the added clinical complication of hemolytic uremic syndrome (a triad of renal failure, thrombocytopenia, and hemolytic anemia), because it is able to express Shiga toxin. Shiga toxin from S. dysenteriae type 1 strains is almost identical to Shiga toxin 1 from E. coli, and the clinical diseases that the two toxins produce may be very similar. However, S. dysenteriae is rare in developed countries, where S. sonne or S. flexneri is more commonly seen. Other complications of Shigella infection include encephalopathy, reactive arthritis, Reiter’s syndrome, toxic megacolon, and perforation. The pathogenesis of Shigella relates largely to their capacity to invade intestinal epithelial cells and induce an inflammatory response. Many of the genes responsible for the invasion are on a virulence plasmid. Only S. dysenteriae type 1 strains are able to produce Shiga toxins that are chromosomally encoded and, unlike E. coli, are not transmitted on bacteriophages. The diagnosis can be confirmed microbiologically; however, Shigella of all types are fastidious organisms, and transporting fecal samples to the laboratory rapidly is important. As with all enteric infections, rehydration is the most critical therapeutic modality. In patients with S. flexneri or S. dysenteriae, antibiotic therapy is usually recommended. However, resistance is becoming a significant problem in many developing countries, but most strains are sensitive to quinolones and naladixic acid is frequently used in developing countries.
Alternatives include cefixime or ceftriaxone. S. sonne infection is usually mild and selflimiting and requires supportive therapy only. 6. Enteroinvasive E. coli Enteroinvasive E. coli (EIEC) are not frequently recognized as food-borne pathogens. However, they are not routinely diagnosed in patients and so little data are available. EIEC have been associated with several outbreaks linked to both water and other foods, such as cheese. A number of prominent serogroups found to be EIEC have been described, including O28, O112, O124, O136, O143, O144, O147, and O164. Clinically, they produce a disease much like shigellosis, described previously, with a watery diarrhea that may lead to dysentery. As with Shigella, EIEC are invasive; however, they do not produce Shiga toxins. Microbiologically, they are lactose-fermenting and indistinguishable from other E. coli. This makes the diagnosis difficult to confirm. Therefore, in the context of a patient with a dysentericlike clinical picture, in whom other invasive organisms have been ruled out, one should consider EIEC, especially if there are large numbers of leukocytes in the stool. Isolating and typing the strains may help, but the only way to confirm the diagnosis is to determine the presence of specific virulence genes using molecular techniques (PCR or DNA probes) or to show that the organisms are invasive in the Sereny test. The latter is a biological assay in which invasion of a guinea pig conjunctiva is demonstrated. D. Other bacterial causes of food-borne illness 1. Aeromonas spp. Aeromonads are gram-negative, facultatively anaerobic, motile, oxidase-positive bacilli. They are frequently present in soil and freshwater, and tend to peak during the summer months in water, which may lead to contamination of fresh produce, meat, and dairy products via contaminated water. Of the various Aeromonas spp., A. hydrophila, A. caviea, A. veonii, and A. jandaei are the ones that are most frequently associated with gastroenteritis and food-borne infections. They grow on most media used for fecal testing, including MacConkey, XLD, desoxycholate, and blood agar, and can be differentiated biochemically. Clinically, this group of organisms causes watery diarrhea that may become persistent. Fecal leukocytes and red cells are usually absent (although dysenteric-like symptoms have been associated with Aeromonas infection) and the patients usually have abdominal pain. Nausea, vomiting, and fever may occur in up to 50% of patients. Simply finding Aeromonas in the stool may not be enough to confirm the diagnosis since asymptomatic carriage has been reported. Therefore, immunological tests to confirm recent infection may be needed. In practice, however, infection with Aeromonas is usually self-limiting and previously healthy people are likely to recover fully, do not require antimicrobial therapy, and the diagnosis is often of academic interest only. The exception to this may be in a patient with persistent diarrhea in whom no other cause has been identified. 2. Plesiomonas shigelloides Plesiomonas shigelloides was placed in its own genus in 1962. It is primarily a freshwater organism and, like Aeromonas spp., the isolation rates increase in warmer months. Plesiomonads are gram-negative, motile, facultative anaerobes and will grow on a variety of media. About 30% ferment lactose and, if one is examining specifically for Plesiomonas, inositol– brilliant green–bile salts may be used. Once isolated, Plesiomonas can be differentiated from other Vibrionaceae and Enterobacteriaceae biochemically. As with the Aeromonas spp., contamination of a variety of food products with water that contain Plesiomonas may cause disease. Data that Plesiomonas are truly associated with enteric disease is variable. Some volunteer studies have not been very convincing that it is a true pathogen in humans.
However, treatment of infected patients with antibiotics has been associated with an improvement, suggesting that it is contributing to a disease process in some way. This is supported by outbreaks in Japan associated with contaminated oysters and water. Symptoms usually occur within 24–48 hrs of exposure and are thought to be linked to the production of an enterotoxin. Typically, infection with Plesiomonas is associated with abdominal cramping; nausea, vomiting, and fever are less common, but bloody stools have been reported. Therapeutically, rehydration is key and, if antibiotic therapy is required, most strains are sensitive to quinolones and trimethoprim–sulfamethoxazole. 3. Enteropathogenic E. coli Enteropathogenic E. coli (EPEC), like Shigella species, are transmitted mainly by the fecal–oral route from one infected individual to another. Therefore, they may be transmitted via food and water but are usually introduced into the food chain from another infected person. EPEC are not associated with an animal reservoir; therefore, meat, poultry, and fresh produce—which are the typical sources of food-borne pathogens—are not usually contaminated at the source, but may become contaminated during handling. EPEC are an important cause of enteric disease in developing countries, but their role in developed nations is questionable. They have caused major outbreaks in the past in various developed countries, but the lack of routine diagnostics for EPEC makes it difficult to know what their role in sporadic disease may be. Clinically, EPEC infection usually presents with a watery diarrhea in the absence of blood. Low-grade fever and vomiting are common, and in developing countries, the mortality rates may be very high, especially in infants, although the factors that are actually causing death in these children are not clear. Microbiologically, EPEC are like any other E. coli. However, there are certain serotypes that may be considered classical EPEC serotypes, including O55, O86, O111, O114, O119, O127, and O142, that are closely associated with EPEC virulence factors. Finding one of these serotypes does not mean that it is an EPEC; indeed, many of the same serotypes have been shown to carry Shiga toxins and to be associated with HUS. Confirmation, using molecular tools, is the only way to be sure an isolate is a true EPEC. Much is known about the pathogenesis of EPEC, and localized adherence, intimate attachment to intestinal epithelial cells followed by activation of a number of epithelial cell signal pathways, all seem to be important in EPEC virulence. Treatment of EPEC requires appropriate rehydration and supportive therapy. Antibiotics may be required, but EPEC are frequently resistant to many antimicrobials and, therefore, susceptibility testing is suggested. 4. Enteroaggregative E. coli Enteroaggregative E. coli (EAEC) get their name from the way in which they adhere to epithelial cells in an aggregative, or “stacked brick,” pattern. EAEC have been associated with persistent diarrhea in many developing countries and in immunocompromised patients in developed nations. There are no known animal reservoirs for EAEC, which is more a reflection of a lack of detailed studies than accurate epidemiological information. Fecal–oral spread from one person to another is considered to be the usual route of transmission. As with EPEC, contamination of food and water from infected individuals is probably important. Clinically, EAEC cause watery diarrhea without major associated symptoms, Microbiologically, they are like any other E. coli and can only be reliably differentiated by their characteristic aggregative phenotype. A number of virulence factors have been found in EAEC, including the production of fimbriae important in aggregation and the expression of a plasmid-encoded toxin. However, not all EAEC have these and it is not clear what the critical virulence determinants really are. At least one study has suggested that treating HIV-positive patients who have persistent diarrhea and are colonized with EAEC results in clearing of the organisms and improvement in symptoms, suggesting that they are true pathogens, but may be more opportunistic than some other food-borne bacteria.
III. VIRAL FOOD-BORNE PATHOGENS Viral agents are considered to be an increasingly important cause of food-borne illness. A number of different viral agents have been associated with foodborne disease and cause a variety of illnesses, varying from a simple gastroenteritis to major systemic upset, such as hepatitis. Food and water are vehicles for viruses, but viruses do not reproduce in food and nor do they produce toxins in food. Some viruses, such as Norwalk, cause large outbreaks; others seem to be more frequently associated with sporadic disease. Overall, the difficulty in diagnosing viral illness has precluded the development of large amounts of epidemiological data. However, the advent of rapid tests (e.g. enzyme immunoassays for the detection of rotavirus) is beginning to change this, and will eventually lead to a better understanding of the epidemiology and disease burden caused by the various food-borne viral pathogens. A. Hepatitis A virus Hepatitis A is an RNA virus belonging to the family Picornaviridae and is present throughout the world. Hepatitis A is usually spread by direct contact from one person to another via the fecal–oral route. There have been many examples of outbreaks of hepatitis A due to contaminated food or water. Some of the incriminated foods include raw shellfish, milk, potato salad, and orange juice. The incubation period following exposure is around 30 days, with a range of 15–50 days. This long incubation period can make tracing the source of infection very difficult. Clinically, hepatitis A may be mild or even asymptomatic, especially in children. In adults, it usually begins with flulike symptoms of headache, myalgia, anorexia, nausea, vomiting, and headache. This is then followed by the development of jaundice. In hepatitis A, the rule is that the disease is self-limiting and long-term consequences are unusual, but occasionally the hepatitis can become fulminant. While during the acute illness, it is usually possible to detect hepatitis A virus in the stool. The virus rapidly disappears after that. Patients usually excrete the virus for 3 weeks prior to the onset of illness and for about 1 week after. Therefore, there is a prolonged period when an individual is infectious and does not realize it. Diagnosis is usually based on the detection of antihepatitis AIgM serum antibodies. IgG antibodies usually appear after resolution of the clinical disease. B. Hepatitis E virus Hepatitis E virus was first described in 1977 and is a small RNA virus from the Caliciviridae family. It is usually transmitted in contaminated drinking water, and probably in food as well, but this has not been documented. Person-to-person spread also occurs. Hepatitis E has an incubation period of 2–9 weeks, although most people develop symptoms around 40 days postexposure. Clinically, the disease is very much like hepatitis A, with constitutional symptoms followed by jaundice. Patients usually recover but the mortality rate has been reported to be as high as 3% in some instances, especially in pregnant women. The diagnosis is made serologically. C. Rotavirus Rotaviruses have become a very common cause of gastrointestinal disease in both developed and developing nations. In the United States, an estimated 1 million cases per year occur and the numbers are much higher in other countries, with over half a million deaths associated with rotavirus infection. Rotavirus is primarily a disease of children and, like the other enteric food-borne viruses, it is transmitted by the fecal–oral route. In some animal models, as few as one viral particle is enough to cause disease. Rotaviruses have a double-stranded RNA genome and are classified by group, subgroup, and serotype. There are three main groups, designated A, B, and C, and it is group A that is the most common cause of human disease.
Groups B and C have also been linked to food-borne transmission, again by the fecal–oral route. Rotaviruses are lytic and cause diarrhea by invading and then destroying villous intestinal epithelial cells—the cells primarily involved in absorption of fluid. The incubation period is usually between 1 and 3 days and is followed by watery diarrhea, vomiting, and fever. The diarrhea may persist for a week or more, resulting in severe dehydration if untreated. Occasionally, the disease can be fulminant and, even with appropriate rehydration, lead to death of the infected infant. Currently, the most rapid way to diagnose the infection is by enzyme immunoassay, although genetic-based and electron microscopybased tests are also available. Supportive care is required for patients infected with rotavirus, especially rehydration, and the disease will usually be self-limiting with no long-term problems. D. Other food-borne viruses Norwalk virus is a small round structured virus (SRSV) and was the first virus to be clearly associated with gastroenteritis. Norwalk is a calicivirus and this group of viruses causes disease worldwide and they have been associated with some large outbreaks, often in confined environments, such as cruise ships. Outbreaks have been associated with contaminated drinking water, swimming water, consumption of undercooked shellfish, ice, and salads. As with the other enteric viruses, fecal contamination of food or water is usually found to be the ultimate source. The incubation time following exposure is around 48 hrs and the clinical illness usually consists of vomiting and diarrhea. The diarrhea is watery without red cells, leukocytes or mucus. The disease is usually selflimiting, settles in 24 hr, and requires no specific therapy. Specific diagnosis is difficult. Anumber of assays are available, including electron microscopy, enzyme immunoassays, and RT–PCR. A number of other viruses have also been associated with outbreaks of gastroenteritis and are, therefore, thought to be spread by the fecal–oral route. The list of potential food-borne viruses is long and includes the viruses previously discussed as well as enteric adenovirus, coronaviruses, toroviruses, reoviruses, and the smaller-sized viruses, such as caliciviruses, astroviruses, parvoviruses, and picobirnaviruses. All of these viruses cause a similar type of gastroenteritis, which typically consists of a noninflammatory watery diarrhea, which is self-limiting. Diagnosis is difficult and usually requires direct electron microscopy of stool samples.
IV. PROTOZOAL FOOD-BORNE PATHOGENS A. Cryptosporidium parvum C. parvum is an apicomplexan protozoan parasite, capable of causing disease in both immunocompetent and immunocompromised individuals. It has gained a reputation in recent years as being a major problem in AIDS patients, but it is important to realize that it can infect normal hosts as well. It is a cause of diarrhea in cattle and so water that is close to cattle pasture may be contaminated with this organism. Other domestic and wild animals are also reservoirs for C. parvum. Both water and food are sources of C. parvum and one of the largest outbreaks, infecting over 400,000 people, was due to contamination of a municipal water supply. Clinically, C. parvum causes watery diarrhea, which may be profuse, abdominal cramping, nausea, and vomiting. Fever is not seen and neither is intestinal bleeding. The organisms are ingested as cysts, which then excyst to release four sporozoites. The sporozoites then attach to and invade intestinal epithelial cells. The invasion remains superficial and C. parvum does not penetrate beyond the intestinal epithelial cell barrier. C. pravum is diagnosed by microscopy of stool to look for cysts or sporozoites, by immunofluorescene microscopy, or by enzyme immunoassay. In immunocompetent patients, the natural history of C. parvum infection is for the disease to be self-limiting and recovery is the rule after a week or two. The pattern is very different in immunocompromised hosts in which the infection is not cleared, and malabsorption becomes a significant and life-threatening problem. There is currently no treatment that is effective for the eradication of C. parvum. B. Giardia lamblia G. lamblia is probably the most frequently found enteric protozoan worldwide. This organism does not cause a dramatic enteric disease or systemic complications, yet infection with it can lead to profound malabsorption and misery in the patient. Like other enteric protozoa, it is found in fecally contaminated water and food and is yet another example of the fecal–oral transmission route. There are different Giardia types but only Lamblia are known to infect humans. The disease is initiated by ingestion of the cysts, and as few as 10–100 may be an infectious dose. Following ingestion, the cysts excyst in the proximal small intestine and release trophozoites. The trophozoites divide by binary fission and attach intimately to the intestinal epithelium, via a ventral disc on the trophozoite. Clinically, giardiasis may be extremely variable. At one extreme, there may be asymptomatic infection, and at the other, severe chronic diarrhea, leading to intestinal malabsorption. Acutely, infection usually results in watery diarrhea and abdominal discomfort. G. lamblia can be diagnosed by fecal microscopy, looking for either the cysts or the trophozoites. Currently, many laboratories look for the parasites using commercially available kits, that utilize either fluorescence microscopy with specific antibodies or enzyme immunoassays. In terms of therapy, metronidazole is the drug of choice. C. Entamoeba histolytica E. histolytica is one the leading causes of parasitic death in the world. It is usually spread by the fecal–oral route, either directly or via contamination of food, e.g., lettuce or water. The cyst is the infective form and cysts may survive weeks in an appropriate environment. Following ingestion, the cysts, excyst in the small bowel and form trophozoites, which then colonize the large bowel, and multiply or encyst, depending on local conditions in the intestine. There are various types of E. histolytica, some of which are pathogenic to humans and some of which are not.
They have been differentiated based on zymodeme analysis, which is a determination of the electrophoretic mobility of certain isoenzymes. E. histolytica causes amebiasis, which may have various clinical manifestations. The trophozoites have the capacity to invade the host from the intestinal lumen, which in the colon results in ulceration of the mucosa, causing amebic dysentery. When the trophozoites invade further, they gain access to the portal blood vessels and are transmitted to the liver. Once in the liver, they are then able to destroy the parenchyma, resulting in a hepatic amebic abscess (this occurs in about 1% of patients with intestinal amebiasis). Intestinal infection with E. histolytica is diagnosed by microscopic examination of the stool, either by wet mount or trichrome stain. Serology is useful in the diagnosis of patients with invasive amibiasis. Patients with E. histolytica need to be treated, and metronidazole is the drug of choice. Luminal drugs that are poorly absorbed are an alternative therapy for carriers of cysts. Drugs such as paromomycin or iodoquinol are used for this purpose. D. Cyclospora cayetanensis C. cayetanensis is a recently described apicomplexan parasite that has been found in food. Most recently, it has been responsible for a number of outbreaks in North America associated with consumption of imported raspberries. It has also been associated with undercooked meat and poultry, and contaminated drinking water and swimming water. Clinically, it causes a self-limiting diarrhea, with nausea, vomiting, and abdominal pain in immunocompetent patients but may lead to a more persistent diarrhea in immunocompromised individuals. C. cayetanenis is diagnosed by direct stool microscopy and oocyst autofluorescence, which appears blue by Epi-illumination and a 365-nm dichroic filter and green by a 450–490-nm dichroic filter. In the laboratory, oocysts may be induced to sporulate, even in the presence of potassium dichromate used to preserve the specimens. After 1–2 weeks, approximately 40% of oocysts contain two sporocysts with two sporozoites in each. Excystation requires a number of steps and occurs when oocysts are subjected to bile salts and sodium taurcholate plus mechanical pressure. Susceptible humans are infected by ingesting sporulated oocysts and, in view of the complexity of the process and the time required, direct person-toperson spread is considered unlikely. The infection can be successfully treated with trimethoprim– sulfamethoxazole. E. Other protozoan infections A number of other protozoa have been associated with food- and water-borne infections in humans. These include Microsporidium, that causes watery diarrhea and malabsorption, and are becoming an increasingly recognized problem in the immunocompromised. Various microsporida, including Enterocytozoon bieneusi and Septata intestinalis, cause human disease. Isospora belli, another apicomplexan protozoon, is an opportunistic pathogen in immunocompromised patients. Sarcocystosis is a rare zoonotic infection in humans that can, on occasion, cause necrotizing enteritis. Dientamoeba fragilis was originally thought to be a commensal but now appears to be associated with a variety of gastrointestinal symptoms, including abdominal pain, nausea, diarrhea, and anorexia. Balantidium coli is the only ciliate known to parasitize humans.
Although most infections are asymptomatic, the disease may present itself as dysentery. Blastocystis hominis is a strict anaerobic protozoon that infects both immunocompetent and immunocompromised hosts and results in a variety of gastrointestinal symptoms, including diarrhea, abdominal pain, nausea, vomiting, anorexia, and malaise. V. CESTODES AND WORMS A. Taenia saginata Taenia saginata, the beef tapeworm, is highly endemic in certain areas of the world, such as parts of South America, Africa, South Asia, and Japan. Humans are the definitive host for the adult tapeworm, which is one of the largest human parasites. They may live as long as 20 years and grow up to 25 meters in length. Consumption of undercooked or raw beef containing living larval forms is how it is acquired. Cattle are the intermediate hosts, in which the hexacanth embryos emerge from the eggs and pass by blood or lymph to muscle, subcutaneous tissue, or viscera. Then, when humans eat the undercooked animal tissue, the life cycle is completed. Clinically, the worms are remarkably quiescent and nausea or a feeling of fullness may be the only symptoms. Vomiting, nausea, and diarrhea may occur. Diagnosis depends on detecting the proglottids in stool and treatment with either praziquantel or albendazole should be curative. B. Taenia solium T. solium is the pork tapeworm, and, unlike the T. saginatum, the larval stage can invade humans and cause infections of the central nervous system. T. solium is distributed worldwide and is acquired by ingesting pork meat that is infected with cysticerci. The adult worm in humans sheds proglottids, which are then eaten by pigs, after which the hexacanth embryos emerge and penetrate the pig’s intestinal wall, where they migrate to muscle and other tissues. Humans then eat the larval forms, which completes the cycle. T. solium is usually smaller than T. saginatum so the clinical symptoms are even less remarkable. Infected humans may just notice the proglottids in stool. The diagnosis is dependent on identifying the proglottids and the treatment is with praziquantel or albendazole. C. Diphyllobothrium latum D. latum is a fish tapeworm and is most common in Northern Europe and Scandinavia. Eating raw fish is the greatest risk factor and the increased consumption of sushi in the United States has resulted in more cases than in the past. The life cycle is complex; after the eggs are passed in human feces, they must hatch and be eaten by copepods, which are freshwater crustaceans. They then develop into larval forms and subsequently are eaten by fish. The procecoids then invade the stomach wall of the fish and, finally, reside in the muscle of the fish. Humans then become infected by eating a fish that harbors a viable plerocercoid larva. Clinically, the infection is usually asymptomatic, but diarrhea, fatigue, and distal paresthesia are well described, as is pernicious anemia, since the tapeworm actively absorbs free vitamin B12. Diagnosis is made by identifying the ova or proglottids in stool. Treatment with praziquantel or niclosamide is effective. D. Hymenolepis nana H. nana, the dwarf tapeworm, is very common in humans and is transmitted via the fecal–oral route. This worm does not require an intermediate host. The majority of infections are asymptomatic but diarrhea, abdominal cramping, and anorexia may occur. The diagnosis is dependent on identifying the typical double lumen eggs in the stool and treatment with praziquantel should be curative. E. Ascariasis Ascaris is the most common intestinal helminth worldwide. Ascaris lumbricoides is specific for humans, who are infected by ingesting food containing the mature ova. The larvae are then released in the small intestine, enter the circulation, and then reach the pulmonary alveoli, where they develop. This pulmonary infestation may cause pneumonitis and allergic manifestations. Finally, larvae migrate up the bronchial tree and are swallowed. Humans are the definitive hosts, but soil is necessary for development of the eggs and also acts as a reservoir. Humans then ingest the developed eggs to complete the cycle. Thus, food or water that are contaminated are sources that infect humans. The diagnosis is made by finding adult worms, larvae, or eggs in the stool. Acarisis may be treated with mebendazole or pyrantel pamoate. F. Trichuriasis Trichuris trichiura is commonly known as whipworm and is frequently found in the same parts of the world as ascaris. Humans are the definitive host and eggs that are passed in stool mature in warm moist soil to become infective. They then contaminate food and are ingested by a new host.
Clinically, the worms remain associated with the intestine and may be either asymptomatic or result in chronic diarrhea. In the context of a heavy worm burden, dysentery may develop along with malnutrition. The diagnosis is made by finding adult worms or eggs in stool. Treatment with mebendazole is usually curative. G. Trichinella spiralis This nematode begins its infection in humans following the ingestion of the first-stage larvae and its nurse cell in striated skeletal muscle tissue. The larvae are released from tissue in the stomach and pass to the small intestine, where they infect epithelial cells. The larvae then develop into adult worms and are shed in the stool. Larvae also penetrate into lymph or blood vessels and then on into muscle cells, where a nurse cell begins to form. The principal mode of transmission to humans is through the consumption of undercooked meat, usually pork. The major clinical features of this disease relate to cellular destruction secondary to the parasitic penetration of cardiac or nervous tissue. Gastrointestinal symptoms are also common and include diarrhea and vomiting. The diagnosis is dependent on the histologic identification of nurse cells containing larvae within infected muscle tissue. Serological tests are also of value. The infection may be treated with thiabendazole.
VI. NATURAL TOXINS
There are a number of naturally occurring toxins that may be present in various types of food. Many of these are associated with consumption of seafood but others are related to different and specific foods. A. Ciguatera Ciguatera poisoning is due to the ingestion of a neurotoxin from fish. The toxin is produced in dinoflagellates (e.g., Gambierdiscus toxicus). It then accumulates in the flesh of the fish. This occurs mainly in tropical and subtropical marine fin fish, including mackerel, groupers, barracudas, snappers, amberjack, and triggerfish, although not all of these types of fish are infected all of the time. In humans, the incubation period ranges from as little as 5 min up to 30 hr (with a mean of about 5 hr). There are usually gastrointestinal and neurological symptoms, including nausea, vomiting, watery diarrhea, parasthesias, ataxia, vertigo, and blurred vision. Some patients may go on to develop cranial nerye palsies or even respiratory paralysis. The symptoms may last up to a week, but then usually resolve. The initial diagnosis is usually clinical and confirming the presence of the toxin is difficult. Toxin detection can be undertaken using a mouse bioassay and enzyme immunoassays for toxin detection are being developed. B. Scrombroid Scrombroid poisoning typically occurs after the ingestion of spoiled fish, especially tuna and mackerel. Excess levels of histamine in the flesh of the fish are thought to be the cause of this poisoning. Histamine and other amines are formed in food by the action of decarboxylases, produced by bacteria that act on the histidine or other amino acids. Scrombroid has been associated with a number of other foods, such as Swiss cheese. However, it is more frequently associated with fish, especially if the fish has not been frozen rapidly after being caught. Clinically, symptoms may begin within 10 min to 3 hr following ingestion. Nausea, vomiting, diarrhea, flushing, and headache may all occur. Respiratory distress is a rare complication. The natural history of this disease is that it will resolve in a few hours. The diagnosis is usually clinical and can be confirmed by detecting elevated levels of histamine in the suspected food. C. Shellfish poisoning Four main types of shellfish poisoning have been described, including paralytic shellfish poisoning, neurotoxic shellfish poisoning, diarrheic shellfish poisoning, and toxic–encephalopathic shellfish poisoning. Shellfish poising is due to toxins made by algae (usually dinoflagellates) that accumulate in the shellfish. Paralytic shellfish poisoning is due to saxitoxin, a sodium channel toxin. Clinically, symptoms occur usually within an hour and consist of nausea, vomiting, and paralysis that may be limited to the cranial nerves or, in more severe cases, involve the respiratory muscles. Neurotoxic shellfish poisoning is due to brevitoxin, which is a lipophilic, heat-stable toxin that stimulates postgaglionic cholinergic neurons. Symptoms usually occur within 3 hr of exposure and consist of nausea, vomiting, and parasthesisa. Paralysis does usually not occur. Diarrheic shellfish poisoning, as the name implies, causes a mainly gastrointestinal disturbance with nausea, vomiting, and diarrhea. Toxic–encephalopathic shellfish poisoning (also known as amnesic shellfish poisoning) has caused outbreaks of disease in association with consumption of mussels. Symptoms include nausea, vomiting, diarrhea, server headache, and, occasionally, memory loss. With all these types of poisoning, symptoms usually occur rapidly (within 2 hr) and will usually resolve spontaneously. The exception to this is toxic–encephalopathic poisoning, when the symptoms may not occur for 24–48 hr following exposure. The diagnosis in humans is clinical. However, it may be possible to detect the presence of the toxins using either mouse bioassays or by high performance liquid chromatography (HPLC). D. Tetrodotoxin Tetrodotoxin is present in certain organs within puffer fish and, if ingested, can cause rapid paralysis and death. Symptoms may occur in as little as 20 minutes or after several hours. Symptoms progress from a gastrointestinal disturbance to almost total paralysis, cardiac arrhythmias, and, finally, death within 4–6 hr, after ingestion of the toxin. The diagnosis is clinical and based on history of exposure. Mouse bioassays and HPLC have been used to detect these toxins in food. E. Mushroom toxins and aflatoxins There are a large variety of toxins from different mushrooms that cause a wide variety of diseases in humans.
These toxins can be divided into four general groups as follows: protoplasmic poisons (e.g., amatoxins, hydrazines, orellanine) that cause cellular damage and organ failure (e.g., hepatorenal syndrome); neurotoxins (e.g., muscarine, ilbotenic acid, muscimol, psilocybin) that cause coma, convulsions, and hallucinations, etc; gastrointestinal irritants that produce nausea, vomiting, and diarrhea; and disulfiramlike toxins that only cause a problem if the person ingesting the mushroom has had exposure to alcohol in the previous 48–72 hr. The diagnosis of mushroom poisoning is based largely on the clinical picture and the history of exposure. There is, however, a commerical radioimmunoassay available for the amanitins. Therapy is largely supportive but interventions to reduce toxin absorption from the intestine, such as lavage or administration of activated charcoal, may help. Plasmapheresis also helps to reduce the mortality rate. Aflatoxins are produced by certain strains of fungi, e.g., Aspergillus flavus and A. parasiticus, that grow on various types of food and produce toxins. Nuts, especially tree nuts (Brazil nuts, pecans, pistachio nuts, and walnuts), peanuts, and other oilseeds, including corn and cottonseed, have been implicated most often. There are various types of aflatoxins (B1, B2, G1, and G2), of which B1 is the most common and the most toxic. Clinically, these toxins cause liver damage that may be in the form of cirrhosis or hepatic malignancy. Occasionally, the ingested dose of aflatoxin is so high that an acute condition develops, known as aflatoxicosis, in which there is fever, jaundice, abdominal pain, and vomiting. Diagnosis in humans is clinical, but there are assays available for the detection of the toxins in food. F. Other natural toxins A number of other naturally occurring toxins have been reported. These include grayanotoxin, which is from eating honey made from rhododendrons. This usually causes nausea, vomiting, and weakness soon after the honey is ingested and, typically, is self-limiting in 24 hr. Akee fruit from Jamaica contains hypoglycin A, that causes hypoglycemia and vomiting in 4–10 hr. Curcurbitacin E, from bitter cucumber, can cause cramps and diarrhea within 1–2 hr of ingestion. Hydrogen cyanide may be present in lima beans or cassava root and can lead to death within minutes. Castor beans can contain a hemagglutinin that may cause nausea and vomiting. Red kidney beans also produce a hemagglutinin, known as phytohemagglutinin. This is associated with eating raw or undercooked red kidney beans and usually causes symptoms in 1–3 hr following exposure. Patients may develop severe nausea and vomiting that can be followed by diarrhea. The toxin is heat-sensitive but needs to reach a high enough temperature to be inactivated. A number of cases have been linked to beans cooked in slow cookers, in which the temperature does not get high enough. The outcome is usually good and supportive therapy is all that is needed. Many other agents that may occur in food, such as the pyrrolizidine alkaloids causing liver damage, and a variety of chemicals and heavy metals can be included in the long list of agents that cause food-borne illness; however, it is beyond the scope of this article to discuss them.
VII. SUMMARY
As time moves on, we are finding more and more microbes of multiple types that are associated with food-borne illness. The various bacteria, protozoa, viruses, and natural chemicals discussed in this chapter include the major ones—but there may be others that we do not even know about yet. It is disconcerting that the majority of cases of gastroenteritis that may be related to food and water go undiagnosed. The majority of our epidemiological data is based on less than 5% of cases, and many laboratories do not routinely look for many of the enteric viruses that are probably causing much of the disease. In recent years, food safety has become a major issue, following a number of highly publicized outbreaks involving a variety of enteric pathogens, including E. coli, Salmonella, Listeria, Cyclospora, and Hepatitis A. The food industry has made major efforts to improve the safety of foodprocessing and we are beginning to see the benefits of this with lower levels of bacterial pathogens in poultry. One of the key unanswered questions in relation to food-borne illness is a determination of the outcome, in most cases. We presume that, in the vast majority of food-borne disease, the outcome is good, with no longterm problems. However, we do not know this for a fact. Occasionally however, the outcome is disastrous, with the development of conditions such as HUS, Guillain-Barré, or reactive arthropathy following infection with enteric pathogens. The story is very different in developing countries, where sanitation may be poor and where fecal contamination of food and water is frequent. In such places, food-borne illness is probably killing hundreds of thousands of children each year and resulting in nutritional deficiency in many more. In developed nations, our immediate goals are to further our understanding of how these organisms cause disease, and how to reduce the load in the animals and fresh produce that provide our food supply. In developing countries, simply instituting basic sanitation to reduce fecal–oral spread can have a huge impact on morbidity and mortality. Maintaining good personal hygiene and sanitation will reduce the chance of fecal–oral spread. Similarly, paying attention to food handling, proper cooking, and the avoidance of temperature abuse will all go a long way to reduce the burden of food-borne illness.
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