The Enterobacteriaceae is a large family of gram-negative rods found primarily in the colon of humans and other animals, many as part of the normal flora. These organisms are the major facultative anaerobes in the large intestine but are present in relatively small numbers compared with anaerobes such as Bacteroides. Although the members of the Enterobacteriaceae are classified together taxonomically, they cause a variety of dis eases with different pathogenetic mechanisms. The organisms and some of the diseases they cause are listed in Table 1.
Table1. Diseases Caused by Members of the Enterobacteriaceae
Features common to all members of this heterogeneous family are their anatomic location and the following four metabolic processes: (1) they are all facultative anaerobes; (2) they all ferment glucose (fermentation of other sugars varies); (3) none have cytochrome oxidase (i.e., they are oxidase-negative); and (4) they reduce nitrates to nitrites as part of their energy generating processes.
These four reactions can be used to distinguish the Enterobacteriaceae from another medically significant group of organisms—the nonfermenting gram-negative rods, the most important of which is Pseudomonas aeruginosa.
P. aeruginosa, a significant cause of infection for those who have been hospitalized or have had significant prior antibiotic exposure, does not ferment glucose or reduce nitrates and is oxidase positive. In contrast to the Enterobacteriaceae, it is a strict aerobe and derives its energy from oxidation, not fermentation.
Pathogenesis
All members of the Enterobacteriaceae contain endotoxin in their cell walls. In addition, several exotoxins are produced (e.g., E. coli and Vibrio cholerae secrete exotoxins, called enterotoxins, that activate adenylate cyclase within the cells of the small intestine, causing diarrhea). In addition, E. coli O157 produces Shiga toxin that causes hemolytic-uremic syndrome.
Antigens
The antigens of several members of the Enterobacteriaceae, especially Salmonella and Shigella, are important; they are used for identification purposes both in the clinical laboratory and in epidemiologic investigations. The three surface antigens are as follows:
(1) The cell wall antigen (also known as the somatic, or O, antigen) is the outer polysaccharide portion of the lipopolysaccharide. The O antigen, which is composed of repeating oligosaccharides consisting of three or four sugars repeated 15 or 20 times, is the basis for the serologic typing of many enteric rods. The number of different O antigens is very large (e.g., there are approximately 1500 types of Salmonella and 150 types of E. coli).
(2) The H antigen is on the flagellar protein. Only flagellated organisms, such as Escherichia and Salmonella, have H antigens, whereas the nonmotile ones, such as Klebsiella and Shigella, do not. The H antigens of certain Salmonella species are unusual because the organisms can reversibly alternate between two types of H antigens called phase 1 and phase 2. The organ isms may use this change in antigenicity to evade the immune response.
(3) The capsular or K polysaccharide antigen is particularly prominent in heavily encapsulated organisms such as Klebsiella. The K antigen is identified by the quellung (capsular swelling) reaction in the presence of specific antisera and is used to serotype E. coli and Salmonella typhi for epidemiologic purposes. In S. typhi, the cause of typhoid fever, it is called the Vi (or virulence) antigen.
Laboratory Diagnosis
Specimens suspected of containing members of the Enterobacteriaceae and related organisms are usually inoculated onto two media, a blood agar plate and a selective differential medium such as MacConkey’s agar or eosin-methylene blue (EMB) agar. The differential ability of these latter media is based on lactose fermentation, which is the most important metabolic criterion used in the identification of these organisms (Table 2). On these media, the nonlactose fermenters (e.g., Salmonella and Shigella) form colorless colonies, whereas the lactose fermenters (e.g., E. coli) form colored colonies. On EMB agar, E. coli colonies have a characteristic green sheen. The selective effect of the media in suppressing unwanted gram-positive organisms is exerted by bile salts or bacteriostatic dyes in the agar.
Table2. Lactose Fermentation by Members of the Enterobacteriaceae and Related Organisms
An additional set of screening tests, consisting of triple sugar iron (TSI) agar and urea agar, is performed prior to the definitive identification procedures. The rationale for the use of these media and the reactions of several important organisms are presented in the box titled “Agar Media for Enteric Gram Negative Rods” and in Table 3. The results of the screening process are often sufficient to identify the genus of an organism; however, an array of 20 or more biochemical tests is required to identify the species.
Table3. Triple Sugar Iron (TSI) Agar Reactions
Another valuable piece of information used to identify some of these organisms is their motility, which is dependent on the presence of flagella. Proteus species are very motile and characteristically swarm over the blood agar plate, obscuring the colonies of other organisms. Motility is also an important diagnostic criterion in the differentiation of Enterobacter cloacae, which is motile, from Klebsiella pneumoniae, which is nonmotile.
If the results of the screening tests suggest the presence of a Salmonella or Shigella strain, an agglutination test can be used to identify the genus of the organism and to determine whether it is a member of group A, B, C, or D.
Coliforms & Public Health
Contamination of the public water supply system by sewage is detected by the presence of coliforms in the water. In a general sense, the term coliform includes not only E. coli but also other inhabitants of the colon such as Enterobacter and Klebsiella. However, because only E. coli is exclusively a large intestine organism, whereas the others are found in the environment also, it is used as the indicator of fecal contamination.
In water quality testing, E. coli is identified by its ability to ferment lactose with the production of acid and gas, its ability to grow at 44.5°C, and its characteristic colony type on EMB agar. An E. coli colony count above 4/dL in municipal drinking water is indicative of unacceptable fecal contamination. Because E. coli and the enteric pathogens are killed by chlorination of the drinking water, there is rarely a problem with meeting this standard. Disinfection of the public water supply is one of the most important advances of public health in the twentieth century.
Antibiotic Therapy
The appropriate treatment for infections caused by members of the Enterobacteriaceae and related organisms must be individually tailored to the antibiotic sensitivity of the organism.
Generally speaking, a wide range of antimicrobial agents are potentially effective (e.g., some penicillins and cephalosporins, aminoglycosides, tetracyclines, quinolones, and sulfonamides). The specific choice usually depends on the results of antibiotic sensitivity tests.
Note that some isolates of these enteric gram-negative rods are highly antibiotic-resistant because of the production of β-lactamases, including extended-spectrum β-lactamases (ESBL) and other drug-modifying enzymes. These organisms undergo conjugation frequently, at which time they acquire plasmids (R factors) that mediate multiple drug resistance. For example, plasmid-encoded New Delhi metallo β-lactamase causes resistance to penicillins, cephalosporins, monobactams, and carbapenems.
