In 1983 spiral Gram-negative rods resembling campylobacters were found in patients with type B gastritis (chronic inflammation of the stomach antrum [pyloric end]). The organisms were originally classified as Campylobacter but were subsequently reclassified as a new genus, Helicobacter. Helicobacters were subsequently subdivided into species that primarily colonize the stomach (gastric helicobacters) and those that colonize the intestines (enterohepatic helicobacters). The most important species is H. pylori, which is a gastric helicobacter associated with gastritis, peptic ulcers, gastric adenocarcinoma, and gastric mucosa-associated lymphoid tissue (MALT) B-cell lymphomas (Table 1). The most important enterohepatic helicobacters associated with gastroenteritis and bacteremia are H. cinaedi and H. fennelliae, which have been isolated most commonly in immunocompromised patients (e.g., homosexual males with human immunodeficiency virus [HIV] infections).
Table1. Helicobacter Species Associated With Human Disease
PHYSIOLOGY AND STRUCTURE
Helicobacter species are characterized according to sequence analysis of their 16 S rRNA genes, their cellular fatty acids, and the presence of polar flagella. Currently, more than 50 species have been characterized, but this taxonomy is changing rapidly. Helicobacters have a bacillary or spiral shape in young cultures (0.5–1.0 μm wide × 2–4 μm long) and, like campylobacters, can assume coccoid forms in older cultures (Fig. 1).
Fig1. Scanning electron micrograph of Helicobacter pylori in a 7-day culture. Bacillary and coccoid forms are bound to paramagnetic beads used in immunomagnetic separation. (Courtesy Dr. L. Engstrand, Uppsala, Sweden.)
All gastric helicobacters, including H. pylori, are highly motile (corkscrew motility) and produce an abundance of urease. These properties are believed to be important for survival in gastric acids and rapid movement through the viscous mucus layer toward a neutral pH environment. Most helicobacters are catalase- and oxidase-positive and do not ferment or oxidize carbohydrates, although they can metabolize amino acids by fermentative pathways. LPS, consisting of lipid A, core oligosaccharide, and an O side chain, is present in the outer membrane. H. pylori lipid A has low endotoxin activity compared with other Gram-negative bacteria, and the O side chain is antigenically similar to the Lewis blood group antigens that may protect the bacteria from immune clearance.
Growth of H. pylori and other helicobacters requires a complex medium supplemented with blood, serum, char coal, starch, or egg yolk in microaerophilic conditions (decreased oxygen and increased carbon dioxide) and in a temperature range between 30°C and 37°C. Because helicobacters are relatively difficult to isolate in culture and identify by biochemical testing, most diseases caused by H. pylori are confirmed by nonculture techniques.
PATHOGENESIS AND IMMUNITY
H. pylori is a remarkable bacterium in its ability to establish lifelong colonization in the stomach of untreated humans. Most research into the virulence factors in helicobacters has focused on H. pylori. Multiple factors contribute to the gastric colonization, inflammation, alteration of gastric acid production, and tissue destruction that are characteristic of H. pylori disease. Initial colonization is facilitated by (1) blockage of acid production by a bacterial acid–inhibitory protein and (2) neutralization of gastric acids with the ammonia produced by bacterial urease activity. The actively motile helicobacters can then pass through the gastric mucus and adhere to the gastric epithelial cells by multiple surface adhesion proteins. Surface proteins can also bind host proteins and help the bacteria evade the immune system. Localized tissue damage is mediated by urease by-products, mucinase, phospholipases, and the activity of vacuolating cytotoxin A (VacA), a protein that after penetration into epithelial cells damages the cells by producing vacuoles. Another important virulence fac tor of H. pylori is the cytotoxin-associated gene (cagA) that resides on a pathogenicity island containing approximately 30 genes. These genes encode a structure (type VI secretion system) that acts like a syringe to inject the CagA protein into the host epithelial cells, which interferes with the normal cytoskeletal structure of the epithelial cells. The cag phosphoribosylanthranilate isomerase (PAI) genes also induce interleukin (IL)-8 production, which attracts neutrophils. Release of proteases and reactive oxygen molecules by these neutrophils is believed to contribute to gastritis and gastric ulcers.
EPIDEMIOLOGY
An enormous amount of information about the prevalence of H. pylori has been collected since 1984 when the organ ism was first isolated in culture. The highest incidence of carriage is found in developing countries, in which 70% to 90% of the population is colonized, most before the age of 10 years. The prevalence of H. pylori in industrial countries such as the United States is less than 40% and is decreasing because of improved hygiene and active treatment of colonized individuals. These studies have also demonstrated that 70% to 100% of patients with gastritis, gastric ulcers, or duodenal ulcers are infected with H. pylori. Humans are the primary reservoir for H. pylori, and colonization is believed to persist for life unless the host is specifically treated. Transmission is most likely via the fecal-oral route.
An interesting observation about H. pylori colonization has been made. This organism is clearly associated with diseases such as gastritis, gastric ulcers, gastric adenocarcinoma, and gastric MALT lymphomas. It is anticipated that treatment of colonized or infected individuals will lead to a reduction of these diseases. However, colonization with H. pylori appears to offer protection from gastroesophageal reflux disease and adenocarcinomas of the lower esophagus and gastric cardia. Thus, it may be unwise to eliminate H. pylori in patients without symptomatic disease. Certainly, the complex relationship between H. pylori and its host continues to be defined.
CLINICAL DISEASES
Disease caused by helicobacters is directly related to their site of colonization. For example, H. pylori is associated with gastritis, whereas the enterohepatic species cause gastroenteritis. Colonization with H. pylori invariably leads to histologic evidence of gastritis (i.e., infiltration of neutrophils and mononuclear cells into the gastric mucosa). The acute phase of gastritis is characterized by a feeling of fullness, nausea, vomiting, and hypochlorhydria (decreased acid production in the stomach). This can evolve into chronic gastritis, with disease confined to the gastric antrum (in which few acid-secreting parietal cells are present) in individuals with normal acid secretion, or involve the entire stomach (pangastritis) if acid secretion is suppressed. Approximately 10% to 15% of patients with chronic gastritis will progress to develop peptic ulcers. The ulcers develop at the sites of intense inflammation, commonly involving the junction between the corpus and antrum (gastric ulcer) or the proximal duodenum (duodenal ulcer). H. pylori is responsible for 85% of the gastric ulcers and 95% of the duodenal ulcers. Recognition of the role of H. pylori has dramatically changed the treatment and prognosis of peptic ulcer disease (Clinical Case 1).
Clinical Case 1 . The Discovery of Helicobacter pylori
Chronic gastritis eventually leads to replacement of the normal gastric mucosa with fibrosis and proliferation of intestinal-type epithelium. This process increases the patient’s risk for gastric cancer by almost 100-fold. This risk is influenced by the strain of H. pylori and the host’s response (cagA-positive strains and high levels of IL-8 production are associated with a higher risk for cancer). Infection with H. pylori is also associated with infiltration of lymphoid tissue into the gastric mucosa. In a small number of patients, a monoclonal population of B cells may develop and evolve into a MALT lymphoma.
LABORATORY DIAGNOSIS
Microscopy
H. pylori is detected by histologic examination of gastric biopsy specimens. Although the organism can be seen in specimens stained with hematoxylin-eosin or Gram stain, the Warthin-Starry silver stain is the most sensitive. When an adequate specimen is collected and examined by an experienced microscopist, the test sensitivity and specificity approaches 100% and is considered diagnostic. Because this is an invasive test, alternative test procedures are preferred for routine diagnosis. The microscopic examination of stool specimens for helicobacters is not reliable because the organisms are difficult to see and Campylobacter and nonpathogenic helicobacters may be present.
Antigen Detection
Biopsy specimens can also be tested for the presence of bacterial urease activity. The abundance of urease produced by H. pylori permits detection of the alkaline by-product in less than 2 hours. The sensitivity of the direct test with biopsy specimens varies from 75% to 95%; however, the specificity approaches 100%. Thus, a positive reaction is compelling evidence of an active infection. As with micros copy, the limitation of this method is the requirement for a biopsy specimen. Noninvasive urease testing of human breath (urea breath test) after consumption of an isotopically labeled urea solution has excellent sensitivity and specificity. Unfortunately, this assay is relatively expensive because of the cost of the detection instruments.
A number of polyclonal and monoclonal immunoassays for H. pylori antigens excreted in stool have been developed and demonstrated to have sensitivities and specificities exceeding 95%. These tests are easy to perform, inexpensive, and able to be used on stool specimens rather than biopsies. These assays are now widely recommended for both detection of H. pylori infections and confirmation of cure after antibiotic treatment.
Nucleic Acid–Based Tests
Currently, nucleic acid–based amplification tests for H. pylori and enterohepatic helicobacters are restricted to research laboratories and not used in clinical laboratories.
Culture
H. pylori adheres to gastric mucosa and is not recovered in stool or blood specimens. The bacteria can be isolated in culture if the specimen is inoculated onto an enriched medium supplemented with blood, hemin, or charcoal and incubated in a microaerophilic atmosphere for up to 2 weeks. However, diagnosis of H. pylori infections is most commonly done by noninvasive methods (e.g., immunoassay), with culture reserved for antibiotic susceptibility tests.
Identification
Presumptive identification of isolates is based on their growth characteristics under selective conditions; typical microscopic morphologic findings; and detection of oxidase, catalase, and urease activity. MALDI mass spectrometry can be used for definitive species identification.
Antibody Detection
Serology is an important screening test for the diagnosis of H. pylori, with a variety of commercial tests available. Although IgM antibodies disappear rapidly, IgA and IgG antibodies can persist for months to years. Because the antibody titers persist for many years, the test cannot be used to discriminate between past and current infection. Furthermore, the titer of antibodies measured does not correlate with the severity of disease or the response to therapy.
However, the tests are useful for documenting exposure to the bacteria, either for epidemiologic studies or for the initial evaluation of a symptomatic patient.
TREATMENT, PREVENTION, AND CONTROL
Numerous antibiotic regimens have been evaluated for treating H. pylori infections. Use of a single antibiotic or an antibiotic combined with bismuth is ineffective. The greatest success in curing gastritis or peptic ulcer disease has been accomplished with the combination of a proton pump inhibitor (e.g., omeprazole), a macrolide (e.g., clarithromycin), a β-lactam (e.g., amoxicillin), and metronidazole with administration for 7 to 10 days initially. Treatment failure is most commonly associated with clarithromycin resistance. Susceptibility testing should be performed if the patient does not respond to therapy.
Infection with H. pylori stimulates a strong TH1 cell mediated inflammatory response. Use of H. pylori antigens in experimental vaccines that stimulate TH1 cells leads to enhanced inflammation. In contrast, use of antigens in combination with mucosal adjuvants that induce a TH2 cell response is protective in an animal model and can eradicate existing infections. The effectiveness of these vaccines in humans remains to be demonstrated.
