General Concepts

Introduction

Salmonellae are ubiquitous human and animal pathogens, and salmonellosis, a disease that affects an estimated 2 million Americans each year, is common throughout the world. Salmonellosis in humans usually takes the form of a self-limiting food poisoning (gastroenteritis), but occasionally manifests as a serious systemic infection (enteric fever) which requires prompt antibiotic treatment. In addition, salmonellosis causes substantial losses of livestock.

Clinical Manifestations

Some infectious disease texts recognize three clinical forms of salmonellosis: (1) gastroenteritis, (2) septicemia, and (3) enteric fevers. This chapter focuses on the two extremes of the clinical spectrum—gastroenteritis and enteric fever. The septicemic form of salmonella infection can be an intermediate stage of infection in which the patient is not experiencing intestinal symptoms and the bacteria cannot be isolated from fecal specimens. The severity of the infection and whether it remains localized in the intestine or disseminates to the bloodstream may depend on the resistance of the patient and the virulence of the Salmonella isolate.

The incubation period for Salmonella gastroenteritis (food poisoning) depends on the dose of bacteria. Symptoms usually begin 6 to 48 hours after ingestion of contaminated food or water and usually take the form of nausea, vomiting, diarrhea, and abdominal pain. Myalgia and headache are common; however, the cardinal manifestation is diarrhea. Fever (38°C to 39°C) and chills are also common. At least two-thirds of patients complain of abdominal cramps. The duration of fever and diarrhea varies, but is usually 2 to 7 days.

Enteric fevers are severe systemic forms of salmonellosis. The best studied enteric fever is typhoid fever, the form caused by S typhi, but any species of Salmonella may cause this type of disease. The symptoms begin after an incubation period of 10 to 14 days. Enteric fevers may be preceded by gastroenteritis, which usually resolves before the onset of systemic disease. The symptoms of enteric fevers are nonspecific and include fever, anorexia, headache, myalgias, and constipation. Enteric fevers are severe infections and may be fatal if antibiotics are not promptly administered.

Structure, Classification, and Antigenic Types

Salmonellae are Gram-negative, flagellated, facultatively anaerobic bacilli possessing three major antigens: H or flagellar antigen; O or somatic antigen; and Vi antigen (possessed by only a few serovars). H antigen may occur in either or both of two forms, called phase 1 and phase 2. The organisms tend to change from one phase to the other. O antigens occur on the surface of the outer membrane and are determined by specific sugar sequences on the cell surface. Vi antigen is a superficial antigen overlying the O antigen; it is present in a few serovars, the most important being S typhi.

Antigenic analysis of salmonellae by using specific antisera offers clinical and epidemiological advantages. Determination of antigenic structure permits one to identify the organisms clinically and assign them to one of nine serogroups (A-I), each containing many serovars (Table 1). H antigen also provides a useful epidemiologic tool with which to determine the source of infection and its mode of spread.

Table 21-1

Ecologic Classification of Salmonellae.

As with other Gram-negative bacilli, the cell envelope of salmonellae contains a complex lipopolysaccharide (LPS) structure that is liberated on lysis of the cell and, to some extent, during culture. The lipopolysaccharide moiety may function as an endotoxin, and may be important in determining virulence of the organisms. This macromolecular endotoxin complex consists of three components, an outer O-polysaccharide coat, a middle portion (the R core), and an inner lipid A coat. Lipopolysaccharide structure is important for several reasons. First, the nature of the repeating sugar units in the outer O-polysaccharide chains is responsible for O antigen specificity; it may also help determine the virulence of the organism. Salmonellae lacking the complete sequence of O-sugar repeat units are called rough because of the rough appearance of the colonies; they are usually avirulent or less virulent than the smooth strains which possess a full complement of O-sugar repeat units. Second, antibodies directed against the R core (common enterobacterial antigen) may protect against infection by a wide variety of Gram-negative bacteria sharing a common core structure or may moderate their lethal effects. Third, the endotoxin component of the cell wall may play an important role in the pathogenesis of many clinical manifestations of Gram-negative infections. Endotoxins evoke fever, activate the serum complement, kinin, and clotting systems, depress myocardial function, and alter lymphocyte function. Circulating endotoxin may be responsible in part for many of the manifestations of septic shock that can occur in systemic infections.

Pathogenesis

Salmonellosis includes several syndromes (gastroenteritis, enteric fevers, septicemia, focal infections, and an asymptomatic carrier state) (Fig. 1). Particular serovars show a strong propensity to produce a particular syndrome (S typhi, S paratyphi-A, and S schottmuelleri produce enteric fever; S choleraesuis produces septicemia or focal infections; S typhimurium and S enteritidis produce gastroenteritis); however, on occasion, any serotype can produce any of the syndromes. In general, more serious infections occur in infants, in adults over the age of 50, and in subjects with debilitating illnesses.

Figure 21-1

Pathogenesis of salmonellosis.

Most non-typhoidal salmonellae enter the body when contaminated food is ingested (Fig. 2). Person-to-person spread of salmonellae also occurs. To be fully pathogenic, salmonellae must possess a variety of attributes called virulence factors. These include (1) the ability to invade cells, (2) a complete lipopolysaccharide coat, (3) the ability to replicate intracellularly, and (4) possibly the elaboration of toxin(s). After ingestion, the organisms colonize the ileum and colon, invade the intestinal epithelium, and proliferate within the epithelium and lymphoid follicles. The mechanism by which salmonellae invade the epithelium is partially understood and involves an initial binding to specific receptors on the epithelial cell surface followed by invasion. Invasion occurs by the organism inducing the enterocyte membrane to undergo “ruffling” and thereby to stimulate pinocytosis of the organisms (Fig. 3). Invasion is dependent on rearrangement of the cell cytoskeleton and probably involves increases in cellular inositol phosphate and calcium. Attachment and invasion are under distinct genetic control and involve multiple genes in both chromosomes and plasmids.

Figure 21-2

Scheme of the Pathogenesis of Salmonella enterocolitis and diarrhea.

Figure 21-3

Invasion of intestinal mucosa by Salmonella.

After invading the epithelium, the organisms multiply intracellularly and then spread to mesenteric lymph nodes and throughout the body via the systemic circulation; they are taken up by the reticuloendothelial cells. The reticuloendothelial system confines and controls spread of the organism. However, depending on the serotype and the effectiveness of the host defenses against that serotype, some organisms may infect the liver, spleen, gallbladder, bones, meninges, and other organs (Fig. 1). Fortunately, most serovars are killed promptly in extraintestinal sites, and the most common human Salmonella infection, gastroenteritis, remains confined to the intestine.

After invading the intestine, most salmonellae induce an acute inflammatory response, which can cause ulceration. They may elaborate cytotoxins that inhibit protein synthesis. Whether these cytotoxins contribute to the inflammatory response or to ulceration is not known. However, invasion of the mucosa causes the epithelial cells to synthesize and release various proinflammatory cytokines, including: IL-1, IL-6, IL-8, TNF-2, IFN-U, MCP-1, and GM-CSF. These evoke an acute inflammatory response and may also be responsible for damage to the intestine. Because of the intestinal inflammatory reaction, symptoms of inflammation such as fever, chills, abdominal pain, leukocytosis, and diarrhea are common. The stools may contain polymorphonuclear leukocytes, blood, and mucus.

Much is now known about the mechanisms of Salmonella gastroenteritis and diarrhea. Figures 2 and 3 summarize the pathogenesis of Salmonella enterocolitis and diarrhea. Only strains that penetrate the intestinal mucosa are associated with the appearance of an acute inflammatory reaction and diarrhea (Fig. 4); the diarrhea is due to secretion of fluid and electrolytes by the small and large intestines. The mechanisms of secretion are unclear, but the secretion is not merely a manifestation of tissue destruction and ulceration. Salmonella penetrate the intestinal epithelial cells but, unlike Shigella and invasive E. coli, do not escape the phagosome. Thus, the extent of intercellular spread and ulceration of the epithelium is minimal. Salmonella escape from the basal side of epithelial cells into the lamina propria. Systemic spread of the organisms can occur, giving rise to enteric fever. Invasion of the intestinal mucosa is followed by activation of mucosal adenylate cyclase; the resultant increase in cyclic AMP induces secretion. The mechanism by which adenylate cyclase is stimulated is not understood; it may involve local production of prostaglandins or other components of the inflammatory reaction. In addition, Salmonella strains elaborate one or more enterotoxin-like substances which may stimulate intestinal secretion. However, the precise role of these toxins in the pathogenesis of Salmonella enterocolitis and diarrhea has not been established.

Figure 21-4

Electron photomicrograph demonstrating invasion of guinea pig ileal epithelial cells by Salmonella typhimurium. Arrows point to invading Salmonella organisms. (Courtesy Akio Takeuchi, Walter Reed Army Institute of Research, Washington, D.C.).

Host Defenses

Various host defenses are important in resisting intestinal colonization and invasion by Salmonella (Table 2). Normal gastric acidity (pH < 3.5) is lethal to salmonellae. In healthy individuals, the number of ingested salmonellae is reduced in the stomach, so that fewer or no organisms enter the intestine. Normal small intestinal motility also protects the bowel by sweeping ingested salmonellae through quickly. The normal intestinal microflora protects against salmonellae, probably through anaerobes, which liberate short-chain fatty acids that are thought to be toxic to salmonellae. Alteration of the anaerobic intestinal flora by antibiotics renders the host more susceptible to salmonellosis. Secretory or mucosal antibodies also protect the intestine against salmonellae. Animal strains genetically resistant to intestinal invasion by salmonellae have been described. When these host defenses are absent or blunted, the host becomes more susceptible to salmonellosis; factors that render the host more susceptible to salmonellosis are listed in Table 3. For example, in AIDS, Salmonella infection is common, frequently persistent and bacteremic, and often resistant to even prolonged antibiotic treatment. Relapses are common. The role of host defenses in salmonellosis is extremely important, and much remains to be learned.

Table 21-2

Host Defenses Against Salmonellae.

Table 21-3

Factors Increasing Susceptibility to Salmonellosis.

Epidemiology

Contaminated food is the major mode of transmission for non-typhoidal salmonellae because salmonellosis is a zoonosis and has an enormous animal reservoir. The most common animal reservoirs are chickens, turkeys, pigs, and cows; dozens of other domestic and wild animals also harbor these organisms. Because of the ability of salmonellae to survive in meats and animal products that are not thoroughly cooked, animal products are the main vehicle of transmission. The magnitude of the problem is demonstrated by the following recent yields of salmonellae: 41% of turkeys examined in California, 50% of chickens cultured in Massachusetts, and 21% of commercial frozen egg whites examined in Spokane, WA.

The epidemiology of typhoid fever and other enteric fevers primarily involves person-to-person spread because these organisms lack a significant animal reservoir. Contamination with human feces is the major mode of spread, and the usual vehicle is contaminated water. Occasionally, contaminated food (usually handled by an individual who harbors S typhi) may be the vehicle. Plasmid DNA fingerprinting and bacteria phage lysotyping of Salmonella isolates are powerful epidemiologic tools for studying outbreaks of salmonellosis and tracing the spread of the organisms in the environment.

In typhoid fever and non-typhoidal salmonellosis, two other factors have epidemiologic significance. First, an asymptomatic human carrier state exists for the agents of either form of the disease. Approximately 3% of persons infected with S typhi and 0.1% of those infected with non-typhoidal salmonellae become chronic carriers. The carrier state may last from many weeks to years. Thus, human as well as animal reservoirs exist. Interestingly, children rarely become chronic typhoid carriers. Second, use of antibiotics in animal feeds and indiscriminant use of antibiotics in humans increase antibiotic resistance in salmonellae by promoting transfer of R factors.

Salmonellosis is a major public health problem because of its large and varied animal reservoir, the existence of human and animal carrier states, and the lack of a concerted nationwide program to control salmonellae.

Diagnosis

The diagnosis of salmonellosis requires bacteriologic isolation of the organisms from appropriate clinical specimens. Laboratory identification of the genus Salmonella is done by biochemical tests; the serologic type is confirmed by serologic testing. Feces, blood, or other specimens should be plated on several nonselective and selective agar media (blood, MacConkey, eosin-methylene blue, bismuth sulfite, Salmonella-Shigella, and brilliant green agars) as well as intoenrichment broth such as selenite or tetrathionate. Any growth in enrichment broth is subsequently subcultured onto the various agars. The biochemical reactions of suspicious colonies are then determined on triple sugar iron agar and lysine-iron agar, and a presumptive identification is made. Biochemical identification of salmonellae has been simplified by systems that permit the rapid testing of 10–20 different biochemical parameters simultaneously. The presumptive biochemical identification of Salmonella then can be confirmed by antigenic analysis of O and H antigens using polyvalent and specific antisera. Fortunately, approximately 95% of all clinical isolates can be identified with the available group A-E typing antisera. Salmonella isolates then should be sent to a central or reference laboratory for more comprehensive serologic testing and confirmation.

Control

Salmonellae are difficult to eradicate from the environment. However, because the major reservoir for human infection is poultry and livestock, reducing the number of salmonellae harbored in these animals would significantly reduce human exposure. In Denmark, for example, all animal feeds are treated to kill salmonellae before distribution, resulting in a marked reduction in salmonellosis. Other helpful measures include changing animal slaughtering practices to reduce cross-contamination of animal carcasses; protecting processed foods from contamination; providing training in hygienic practices for all food-handling personnel in slaughterhouses, food processing plants, and restaurants; cooking and refrigerating foods adequately in food processing plants, restaurants, and homes; and expanding of governmental enteric disease surveillance programs.

Recently, The U.S. Department of Agriculture has approved the radiation of poultry to reduce contamination by pathogenic bacteria, e.g. salmonella and campylobacter. Unfortunately, radiation pasteurization has not yet been widely accepted in the U.S. Adoption and implementation of this technology would greatly reduce the magnitude of the salmonella problem.

Vaccines are available for typhoid fever and are partially effective, especially in children. No vaccines are available for non-typhoidal salmonellosis. Continued research in this area and increased understanding of the mechanisms of immunity to enteric infections are of great importance.

General salmonellosis treatment measures include replacing fluid loss by oral and intravenous routes, and controlling pain, nausea, and vomiting. Specific therapy consists of antibiotic administration. Typhoid fever and enteric fevers should be treated with antibiotics. Antibiotic therapy of non-typhoidal salmonellosis should be reserved for the septicemic, enteric fever, and focal infection syndromes. Antibiotics are not recommended for uncomplicated Salmonella gastroenteritis because they do not shorten the illness and they significantly prolong the fecal excretion of the organisms and increase the number of antibiotic-resistant strains.

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