Basic Bacteriology and Culture

Lief Percival Andersen; Torkel Wadström

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Mobley HLT, Mendz GL, Hazell SL, editors. Helicobacter pylori: Physiology and Genetics. Washington (DC): ASM Press; 2001.

Helicobacter pylori: Physiology and Genetics.

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Chapter 4Basic Bacteriology and Culture

Lief Percival Andersen and Torkel Wadström.

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Campylobacteriaceae

In 1957 E. O. King (81) described human infections due to members of the Campylobacteriaceae, with high mortality in patients with septicemia and meningitis caused by Vibrio fetus and "related vibrios." Taxonomic studies in the early 1960s (133, 155) introduced a new genus, Campylobacter, that included microaerobic vibrios that did not resemble Vibrio cholerae. Vibrio fetus was renamed as Campylobacter fetus and the "related vibrios" included Campylobacter jejuni and Campylobacter coli. In the 1970s Butzler et al. isolated C. jejuni frequently from fecal cultures of children with diarrhea (23), and Skirrow (136) introduced a selective medium for stool cultures for isolating Campylobacter spp., which was a breakthrough in the research of thermophilic Campylobacter species. The family Campylobacteriaceae was proposed by Vandamme et al. in 1991 (154). This family includes the genera Campylobacter, Arcobacter, and Helicobacter as well as Thiovulum (87), Wollinella (164), "Flexispira" (7), two Bacteroides species (B. gracilis and B. ureolyticus) (119), and the closely related Anaerobiospirillum (22). Taxonomic studies (152, 153, 154) divided the genus Campylobacter into three new genera: (i) Campylobacter including B. gracilis (138, 152), (ii) Helicobacter including "Flexispira," and (iii) Arcobacter, Wollinella, Thiovulum, and Anaerobiospirillum were reported as separate genera.

The Genus Helicobacter

When a new slow-growing Campylobacter-like organism (CLO) was cultured by Marshall in 1982, it was classified as Campylobacter pyloridis and shortly after corrected to C. pylori (91, 93). New intestinal CLOs were discovered at the same time, and C. pylori was sometimes referred to as gastric CLO (GCLO) and GCLO-1 when another CLO (GCLO-2, C. jejuni subsp. doylei) was isolated from the human stomach (78, 152). It soon became clear that even though C. pylori resembles Campylobacter in many aspects, it differs in important features such as flagellum morphology, fatty acid content, and 16S rRNA sequence (53, 54, 92, 127). C. pylori was transferred to a new genus, Helicobacter, and named Helicobacter pylori in 1989, together with Campylobacter fennelliae and Campylobacter cinaedae (52).

During the last decade, the genus Helicobacter has expanded tremendously and new species are regularly included. The majority of these new Helicobacter species are found in the stomachs and intestines of different animals (7, 21, 35, 40, 46, 48, 49, 50, 51, 52, 58, 71, 89, 91, 93, 96, 144).

Cell Biology of H. pylori

Factors involved in colonization and adhesion

Several virulence factors for gastric colonization, tissue damage, and survival have been identified in H. pylori (Table 1). Flagella, urease, and adhesins are all essential factors for H. pylori to colonize the gastric mucosa. Mutants of H. pylori without flagella or without urease are unable to colonize the gastric mucosa in laboratory animals (41, 42).

Table 1

Virulence factors identified in H. pylori.

Flagella and motility

The curved morphology of H. pylori and the polar motility caused by flagella in one end cause screw-like movements, which may enable the organism to penetrate the mucin layer. The motility of H. pylori is increased when the viscosity of the media is increased in vitro and transverses a methyl glucose solution 10 times more efficiently than Escherichia coli (61), but the motility is pH dependent and impaired at a pH below 4 (97).

Urease, catalase, superoxide dismutase, and alkylhydroperoxidase reductase

Urease is one of the key enzymes in H. pylori pathogenesis. It has a molecular weight of 550 kDa and consists of three subunits of 26.5 kDa (Ure A), 61 kDa (Ure B), and 13 kDa (Ure C) (84, 100). Urease is necessary for H. pylori to maintain a pH-neutral microenvironment around the bacteria, necessary for survival in the acidic stomach (59, 123). Urease induces self-destruction of H. pylori in vitro in nonacidic media (110). Urease is strongly immunogenic and chemotaxic for phagocytes.

Superoxide dismutase has been isolated from H. pylori, which breaks down superoxide produced in polymorphonuclear leukocytes and macrophages and thereby prevents the killing of these organisms (143). Catalase protects H. pylori against the damaging effects of hydrogen peroxide released from phagocytes (60). Urease and catalase may be excreted from H. pylori to the surrounding environment and may protect this pathogen from the humoral immune response (59).

Outer membrane proteins, phospholipids, glycolipids, and other adhesins

H. pylori adheres to mucin and binds specifically to gastric mucosa epithelial cells both in vivo and in vitro (45, 64, 150). Different adhesion patterns, which are different in children and in adults, have been described (14). Several putative gastric tissue receptor structures have been described for H. pylori such as (i) sialoglycoconjugates in gastric mucins and on epithelial cells, phagocytes, and extra-cellular matrix (43, 69, 157), (ii) sulfated glycoconjugates such as heparan sulfate and other glycosaminoglycans (158), and (iii) sulfatides (9, 77, 130, 151) and various sialylated and nonsialylated glycolipids (97).

Binding of H. pylori to cell surface fucosylated blood group antigens, H1 and Lewis part of the blood group O in the ABO system, was first described by Borén and collaborators (18) and was shown to mediate adherence to human and rhesus monkey gastric tissue surface cells (19). More recently, Petersson and collaborators (124) identified the H. pylori blood group antigen-binding adhesins, Bab A and Bab B, purified the proteins, and cloned the babA and babB genes. Two closely related basic proteins of 78 kDa were characterized. The sialic acid-specific lectins of 19 and 23 kDa have also been purified and the corresponding genes cloned (T. Borén, personal communication). These proteins, unlike BabA and -B, do not belong to the family of proteins, mostly named helicobacter outer membrane proteins by a nomenclature introduced by T. J. Trust and associates (see chapter 7). Cell surface adhesins recognizing sulfatides were not identified, whereas heparan sulfate binding proteins (HeBPs) were purified and characterized by Utt et al. (151; unpublished). None of the glycolipid-binding surface proteins has been purified and characterized yet. Interestingly, no similar putative mucin and cell adhesin has been identified in H. felis or other newly identified species, except for H. mustelae (P. O'Toole and T. Trust, personal communication). In vitro affinity binding studies for the Lewis binding give high-affinity constants (K_a, − 2.5; 10 to 11 M) and also reveal high affinity for human mucin-binding glycoconjugates to hemagglutinating sialic acid-specific adhesins or lectins (SALs) (157). Recent studies by Syder et al. (139) in a transgene mouse model suggest that SALs may become key adhesins in inflamed gastric mucosa. It is noteworthy that these SALs are produced already in the exponential growth phase while the Bab A and B adhesins appear mainly in the stationary phase cells. SALs and HeBPs may be the key receptors on leukocytes and macrophages, and trigger lectinophagocytosis as for several other microbial pathogens (158).

Factors involved in tissue damage and survival factors

Enzymes: protease, etc

Conflicting results have been reported concerning proteolytic enzymes in H. pylori. It seems probable that H. pylori glycosulfatase degrades gastric mucin (135, 137). H. pylori possesses phospholipase A, which can digest phospholipids of cell membranes (86). Urease has a cytotoxic activity (61, 98). Recently, alcohol dehydrogenase has been described to contribute to gastric mucosal injury (128).

Toxins: vacuolating cytotoxin A, lipopolysaccharide

H. pylori contains a toxin, Vac A, which can produce vacuoles in gastric epithelial cells and has been related to peptic ulcer, severe gastritis (25, 31, 32, 131, 147), and mucosal integrity (47). Lipopolysaccharide (LPS) in H. pylori has a low biological activity as compared to LPS from other gram-negative bacteria (30, 112), which may be explained by the unusual composition of lipid A (95). LPS from H. pylori stimulates phenotypic transcription and functional changes in monocytes (112). Assays using gastric mucosal laminin (integrin) receptor binding to a laminin-coated surface have revealed a significant decrease in receptor binding occurring in the presence of H. pylori LPS (126). When the gastric epithelial barrier is weakened by disruption of the mucosal surface cells and the extracellular matrix, LPS is responsible for a marked increase in epithelial cell apoptosis (126). LPS from H. pylori has recently attracted interest because LPS from most strains mimic Lewis and/or Lewis blood group antigens (102). This mimicry may play a role in the regulation of the immune response and induce autoantibodies against the gastric proton pump.

Other putative virulence factors

Several heat shock proteins (Hsp) such as 58.2-kDa Gro-El (Hsp B), 13-kDa Gro-Es (Hsp A), and a 70-kDa Hsp have been identified in H. pylori (37, 44, 106). Hsp are produced by all cells and are involved in stabilizing and probably repairing proteins under harsh conditions that may be important to survive in the stomach.

H. pylori transforms into coccoid forms (15, 24, 39) under certain conditions such as nutrient starvation and media containing growth inhibitors (bismuth, proton pump inhibitor, or certain antibiotics). These coccoid forms have been reported to survive for several years in river water and have been proposed by some to be an important factor for transmission, by fecal excretion, and for therapy failure.

Microscopy and Growth of H. pylori

Specimens for culture of H. pylori

H. pylori is the microorganism most frequently found in the human gastric mucosa in association with gastric epithelial cells, but other curved bacteria have also been found in the human gastric mucosa: C. jejuni subsp. doylei (GCLO-2) (55, 78, 140), "Helicobacter heilmannii" ("Gastrospirillum hominis," "H. germanium") (34, 63, 66), and H. felis (120). In our hospital laboratory, Campylobacter sputorum, Campylobacter upsaliensis, and Selenomonas species have occasionally been cultured from human gastric biopsies (L. P. Andersen, unpublished data). Some of these microorganisms may be difficult to distinguish by routine laboratory methods. Apart from H. pylori, "H. heilmannii" is the most common bacterium in human gastric mucosa, with a prevalence of up to 0.5% in dyspeptic patients in western Europe (66). "H. heilmannii" is usually found in the foveolae associated with mild chronic gastritis whereas H. pylori is usually found on the surface epithelium associated with severe gastritis. The contact with epithelial cells is usually more superficial for "H. heilmannii" than for H. pylori (66). Occasionally, "H. heilmannii" and H. pylori are found simultaneously (34).

Gastric specimens

H. pylori is most regularly found in the antral part of human gastric mucosa of untreated persons. In persons treated with acid-suppressive drugs (proton pump inhibitors and H₂ antagonists) H. pylori may be present in higher numbers in the body of the stomach. H. pylori is more frequently found in gastric antrum than in duodenal biopsies even in persons with duodenitis and duodenal ulcer (about 50%). H. pylori can only be cultured from gastric juice in about 15% of persons with H. pylori cultured from gastric antrum and from less than 50% of esophageal biopsies from untreated persons with esophagitis, even though H. pylori can be cultured from gastric antrum (1, 3). Thus, it is always important to obtain antral biopsies as well as corpus biopsies from persons recently treated with acid-suppressive drugs, whereas duodenal and esophageal biopsies and gastric juice are of less importance for routine diagnostics but may be useful for PCR diagnostics (163) and special research purposes.

The number of biopsies necessary to diagnose H. pylori by culture has been estimated in a study where more than 95% of H. pylori was cultured from one antral biopsy. For optimal results at least four biopsies should be cultured (108). It is generally accepted, according to the modified Sydney classification of gastritis (36), that at least one biopsy from antrum and two biopsies from corpus should be taken for culture to ensure a sufficient diagnosis.

Extragastric specimens

H. pylori has occasionally been cultured from ectopic gastric mucosa in Meckel's diverticulum, esophagus, rectum, urinary bladder, dental plaque, and feces (12, 28, 38, 79, 122, 146, 148). Recently, H. pylori has also been detected by PCR in specimens from the gallbladder and liver (114). H. pylori has mainly been identified by PCR in dental plaque, liver specimens, and fecal specimens. Biopsies should be taken from sites with gastric metaplasia and dental plaque or gingival crest. No systematic studies have been carried out to recommend optimal sample sites for extragastric H. pylori infections. Several Helicobacter and Campylobacter species are harbored in the mouth and intestine. Culture-confirmed microbiological identification is preferable to ensure the bacteriological diagnosis of isolates from these sites, at least until molecular biological methods have been better evaluated than they are today.

Microbiological detection of H. pylori

Primary microscopy of helicobacter-like organisms

Microscopy of gram-stained smears or imprint of gastric biopsies reveals curved gram-negative rods resembling Helicobacter in 60 to 100% of the culture-positive biopsies (1, 3, 73, 101, 109, 111). "H. heilmannii" and H. felis are easy to distinguish from H. pylori by microscopy because of the long corkscrew shape of "H. heilmannii" and H. felis (5, 63, 66, 88). In smears, helicobacter-like organisms are usually seen unevenly distributed in clusters or rows along the epithelium cells. The presence of polymorphonuclear leukocytes is not always a dominant feature in smears. Phase-contrast microscopy (125) and other staining methods such as silver stain (118) and acridine orange have been described. The probability of detecting H. pylori by microscopy may be increased by using specific immunofluorescence or peroxidase conjugated antibodies to H. pylori.

The atmosphere for culture of H. pylori

In general, primary cultures of H. pylori have less oxygen tolerance than most Campylobacter species, with a growth maximum at 3 to 7% of O₂. H. pylori is usually grown in jars with gas-generation kits (4, 85, 94) or a standard microaerobic atmosphere, in CO₂ incubators or anaerobic chambers with a microaerobic atmosphere. Most studies with standardized atmospheres for culture of H. pylori have used 2 to 5% O₂, 5 to 10% (optimal closer to 10%) CO₂, and 0 to 10% H₂ (1–3, 5, 6, 73, 109, 111).

H. pylori strains are variable in their growth response to different culture conditions, but no systematic studies on the different atmospheres and growth systems have been published. In our hands, the atmospheres produced by gas-generation kits were unstable for reliable culture of H. pylori when the gas generator was changed every third day (L. P. Andersen, unpublished data). Other laboratories have used this technique successfully by changing the gas generator every day (F. Megraud, personal communication). No quantitative differences were found between atmospheres with (jars) and without (chambers) H₂ (L. P. Andersen, unpublished data). Subcultures of H. pylori can rapidly be adapted to grow anaerobically or in a standard CO₂ mixture (18% O₂, 10% CO₂) in an incubator even under aerobic conditions (149).

Nonselective and selective media for growing H. pylori

H. pylori can grow on different solid media containing blood or blood products (blood or lysed blood agar plates). Most studies have used Brucella agar or Columbia agar as the agar base. An amount of 7 to 10% blood improves the growth of H. pylori as compared with 5% blood. Horse blood may also improve the growth of H. pylori as compared to sheep blood (33, 83). Supplement of agar with cyclodextrin B can be used for blood-free culture media for H. pylori, but with large differences between different batches of cyclodextrin (116). Egg yolk emulsion agar has also been described as a blood-free medium for growth of H. pylori (161). In a small study, egg yolk emulsion as a liquid medium was compared with four other liquid media described in the literature and it was found superior with regard to growth rate of H. pylori, but it contained too many non-H. pylori proteins to be useful for antigen production (A. K. V. Jensen, unpublished data).

Often H. pylori grows poorly or not at all on selective media containing antibiotics. Skirrows and Dents selective media seem to be the best available commercial selective media and have been used in several studies (62, 76, 78, 85). There seem to be greater differences between horse and sheep blood agar, in favor of horse blood, than between horse blood agar with and without antibiotics (33, 83). By comparing agar plates containing 5% horse blood, 10% horse blood, 7% lysed horse blood, 7% lysed horse blood with trimethoprim, and selective Campylobacter plates (modified Skirrows medium) (all from SSI Diagnostica, Hillerød, Denmark), we found that H. pylori grew with more and larger colonies on 10% horse blood agar plates and 7% lysed horse blood agar plates than on the other media, but the numbers of H. pylori-positive patients were almost equal with all media (unpublished data).

Usually H. pylori grows slowly in liquid media, with formation of a high number of coccoid forms (161, 162). Contaminating microorganisms (staphylococci, yeasts, etc.) usually grow much faster than H. pylori and make liquid media useless for primary culture of biopsies. Because of the risk of contaminated samples, a selective medium is usually recommended in addition to the nonselective media for routine culture.

Transportation and handling of biopsies for culture of H. pylori

Some studies have shown that sufficient culture of H. pylori will be obtained after transportation of the biopsies in a medium such as Stuarts transport medium for up to 24 h at low temperature (about 4°C), whereas a higher temperature (about 20°C) decreases the number of positive cultures significantly (82, 129, 141). Our experience is, however, that there is a more than 95% concordance between culture and histological detection of H. pylori when the biopsies are inoculated on agar plates within 4 h after the biopsies are taken (1, 3). H. pylori was only cultured from about 80% of biopsies with helicobacter-like organisms detected in histological sections in a study with similar culture conditions but a transportation time for biopsies of up to 24 h (4). Thus, a decrease in culture rate of about 15% was found when biopsies were transported or stored overnight. A long transportation time decreases the number of H. pylori especially after antibiotic therapy, and if the number of bacteria is low, culture may become false negative. This yield can be improved by prolonged incubation, up to 12 days.

The above data are based on culture of unhomogenized biopsies by inoculating the biopsies or the agar plates and subsequently spreading the material stepwise using the conventional technique. Grinding the biopsies 10 to 15 s before culture has been proposed to increase the number of H. pylori colonies and improve H. pylori isolation. However, this method may not increase the number of positive biopsies (A. Hirschl, personal communication), and in our hands the growth of contaminations improved more than the growth of H. pylori (L. P. Andersen, unpublished data).

Identification of H. pylori

H. pylori colonies are small (0.5 to 2 mm), translucent to yellowish colonies on 7% lysed horse blood agar and with translucent to pale grayish colonies of 0.5 to 1 mm in size on blood agar. In very young cultures H. pylori may appear as almost straight rods on microscopy. After 3 to 5 days of incubation the bacteria look pleomorphic, with irregular curved rods, several being U shaped. In old cultures, H. pylori appears as degenerative coccoid forms that Gram stain poorly (unpublished data). Because of their small size, H. pylori colonies may be difficult to identify and isolate when there are few colonies and additional contaminating oral microbiota is present. Some contaminating microorganisms may grow as small colonies but differ usually from H. pylori in color.

H. pylori is biochemically closely related to Campylobacter, Arcobacter, and Wollinella species but also resembles Bacteroides, Thiovulum, and Selenomonas species. They are all characterized as being gram-negative rods that are able to grow microaerobically or anaerobically. The rods may be more or less curved, depending on the growth conditions. There are conflicting data in the literature about the nitrite and nitrate reaction of H. pylori. The urease reaction is a key reaction in identifying Helicobacter species, but some Campylobacter lari (UPTC) strains are urease positive and at least one urease-negative H. pylori strain has been isolated from a patient (F. Megraud, personal communication). Several Helicobacter species are gram-negative motile curved rods that are oxidase, catalase, and urease positive, and it may, therefore, be necessary to undertake protein profiles or genomic analysis to ensure the correct identification.

Detection of H. pylori from extragastric specimens by culture and genome methods

Several new species have been discovered during the past years, mainly isolated from animals. Only H. pylori and "H. heilmannii" have been recognized regularly in the human gastric mucosa. H. pylori has also been detected from several extragastric sites. Successful cultures have mainly been associated with findings of gastric metaplasia in esophagus (20, 90), Meckel's diverticulum (10, 104), and rectum (38), whereas in one case H. pylori could not be cultured from the gallbladder with gastric metaplasia (8). Occasionally, H. pylori has been cultured from dental plaque (28, 156) and fecal samples (79, 148). The culture methods used in these cases were similar to those described earlier in this chapter. Detection of extragastric H. pylori from dental plaque, fecal samples, atherosclerotic plaques, and liver was mainly achieved by genome methods and serology (13, 57, 107, 121, 132).

Transformation and survival of H. pylori

Factors responsible for the survival of cultured H. pylori

The conversion and morphological change of spiral-shaped H. pylori into coccoid forms are achieved in various ways: (i) by nutrient deprivation (39, 160), (ii) by exposure to anti-ulcer drugs and antibiotics (12, 17, 27, 70), (iii) by extended incubation (27, 39, 105), (iv) by pH adjustment (9, 50, 72), and (v) by attachment to the gastric epithelium (134). Changes in the morphology of H. pylori in culture on agar plates over time can be observed: after 3 days spiral forms dominated, after 6 days about half of the bacteria had converted into U-shaped or coccoid forms, and after 10 days only coccoid forms may be found (11, 17, 24, 26, 27, 29, 41, 75, 105, 113, 160). Morphological changes are induced faster with exposure to detrimental environmental circumstances (68, 160).

Transformation of H. pylori from rods to coccoids

The spiral form of H. pylori is a curved rod that is 2 to 4 μm long and 0.5 to 0.8 μm wide and possesses one to seven sheathed flagella originating from one pole, with a characteristic polar membrane (29, 75, 113). When spiral forms of H. pylori transform into coccoid forms in vitro, a C or U form is initiated by ingrowth of the periplasm on one site of the bacteria. Both in vitro and in vivo coccoid forms of H. pylori vary in size from 1 to 2 μm in diameter for organisms with dense cytoplasmic bodies up to 3 to 4 μm in diameter for organisms with less dense cytoplasm (17, 24, 29, 75, 113). The early stages of coccoid forms have preserved the characteristic polar membrane, flagella, and motility as seen in spiral forms (11, 17, 75). Three-month-old coccoid forms have a complete cell wall, cell membrane, and cytoplasm (56, 113).

Coccoid forms of H. pylori are able to maintain an oxidative metabolism at the same level as spiral forms for several months (56, 113). It remains to be established whether this metabolism of coccoids represents viability or a nonviable "bag" of enzymes (56). The preservation of metabolic activity is supported by a low but constant ATP level over a period of 1 month and the presence of polyphosphates as a phosphorus and energy source that allow a certain endogenous metabolism (16, 17, 113). The protein content only changes a little whereas the lipid content changes considerably after coccoid transformation (145). Coccoid forms of H. pylori grown on solid media for 4 weeks lost their urease activity and the metabolic activity of some enzymes (leucine arylamidase, naphthol-SA-1β-phosphohydrolase) was reduced, whereas the metabolic activities of other enzymes (alcaline and acid phosphatase) were unchanged (67). The genes coding for urease subunit C and a 26-kDa protein were detected unchanged by PCR even though the urease activity was lost, indicating the vitality of the coccoid forms (67). This is confirmed by the fact that coccoid forms of H. pylori can be detected by means of acridine orange staining (25). Newly synthesized DNA has been detected by bromodeoxyuridine incorporated into 3-month-old coccoid forms of H. pylori (17).

Viability and regrowth of H. pylori from coccoid forms

Coccoid forms of H. pylori are usually nonculturable and difficult to study (17, 56, 113). Attempts to document the biological significance and the viability of coccoid forms of H. pylori have been carried out in vitro in cell cultures and in vivo in animal models.

In cell cultures, the initial contact and attachment of H. pylori to human gastric epithelial cells occur rapidly, often within minutes, at the aflagellated end of the bacterium. Transformation into coccoid forms is common after attachment or internalization of the bacterium (134). H. pylori does not multiply intracellularly, and many of the coccoid forms that are found within a few hours after uptake are degenerative forms (134). Coccoid forms of H. pylori adhere to cultured Kato III cells and human gastric carcinoma cells and cause similar morphological changes as spiral forms (25, 134). Actin rearrangement occurs directly beneath the site of attachment of H. pylori, forming a fine condensed structure concentric to the bacterium. Coccoid H. pylori appears to induce an earlier and stronger cytoskeletal response than spiral forms (134). The coccoid forms of H. pylori adhere to the same components as the spiral forms and also contain cell-surface agglutinins and heparan sulfate-binding proteins (43, 65, 80, 103, 117).

In mice, coccoid forms of H. pylori colonize the stomach, induce inflammation, and cause humoral immune response similar to that of spiral forms (26, 74, 159). The inflammatory response might be similar to that of a passive immunization with dead organisms, but H. pylori could be cultured from the stomach of mice 1, 2, and 4 weeks after inoculation with the coccoid forms of H. pylori (26, 159). H. pylori grown in liquid media for 20 days formed coccoids that were nonculturable on solid media but could after inoculation in mice be isolated from the murine stomach (27). In gnotobiotic piglets, urease-negative mutants of H. pylori are unable to colonize the gastric mucosa, and nonmotile or weakly motile strains of H. pylori are less virulent and colonize the gastric mucosa to a lesser extent than motile strains (39, 41).

H. pylori can be found free in the mucin layer or attached to the gastric epithelial cells in the human stomach, where it induces changes in epithelial cell architecture: adhesion pedestals, indentation sites, abutting adhesion, membrane fusions or condensations, vacuolation, and internalization (14, 61, 64, 115, 142). Epithelial degeneration is present when more than 20% of the bacteria have formed adhesion sites (64). Mucosal epithelial cells have been found in increased numbers in the vicinity of the attachment of coccoid forms (11). Degeneration is recognized histologically by ragged cytoplasmic margins, a high nucleocytoplasmic ratio, and a heaping up of cells to form syncytium-like masses. In children, coccoid forms of H. pylori have been found to be closely associated with damaged mucous cells whereas spiral forms have been found in proximity to unchanged or less damaged cells (72). Coccoid forms of H. pylori are found more frequently and in larger numbers in the gastric mucosa of patients with gastric cancer than in patients with peptic ulcer disease (29, 126).

References

1.: Andersen L. P., Elsborg L., Justesen T. Campylobacter pylori in peptic ulcer disease: II. Endoscopic findings and cultivation of C. pylori. Scand. J. Gastroenterol. 1988;23:760–764. [PubMed: 3175537]
2.: Andersen L. P., Espersen F. Immunoglobulin G antibodies to Helicobacter pylori in patients with dyspeptic symptoms investigated by the Western immunoblot technique. J. Clin. Microbiol. 1992;30:1743–1751. [PMC free article: PMC265374] [PubMed: 1629330]
3.: Andersen L. P., Holck S., Povlsen C. O., Elsborg L., Justesen T. Campylobacter pyloridis in peptic ulcer disease: I. Gastric and duodenal infections caused by C. pyloridis. Histopathologic and microbiologic findings. Scand. J. Gastroenterol. 1987;22:219–224. [PubMed: 3576129]
4.: Andersen L. P., Kiilerick S., Pedersen G., Thoreson A. C., Jørgensen F., Rath J., Larsen N.-E., Børup O., Krogfelt K., Scheibel J., Rune S. An analysis of seven different methods to diagnose Helicobacter pylori infections. Scand. J. Gastroenterol. 1998;33:24–30. [PubMed: 9489904]
5.: Andersen L. P., Nørgaard A., Holck S., Blom J., Elsborg L. Isolation of a "Helicobacter heilmannii"-like organism from the human stomach. Eur. J. Clin. Microbiol. Infect. Dis. 1996;15:95–96. [PubMed: 8641315]
6.: Andersen L. P., Raskov H. H., Elsborg L., Holck S., Justesen T., Hansen B. F., Nielsen C. M., Gaarslev K. Prevalence of antibodies against heat-stable antigens from Helicobacter pylori in patients with dyspeptic symptoms and normal persons. APMIS. 1992;100:779–789. [PubMed: 1389098]
7.: Archer J. R., Romero S., Ritchie A. E., Hamacher M. E., Steiner B. M., Bryner J. H., Schell R. F. Characterization of an unclassified microaerophilic bacterium associated with gastroenteritis. J. Clin. Microbiol. 1988;26:101–105. [PMC free article: PMC266203] [PubMed: 3343302]
8.: Arnaout A. H., Abbas S. H., Shousha S. Helicobacter pylori is not identified in areas of gastric metaplasia of gall bladder. J. Pathol. 1990;160:333–334. [PubMed: 2358970]
9.: Ascensio F., Fransson L. Å., Wadström T. Affinity of the gastric pathogen Helicobacter pylori for the N-sulphated glycosaminoglycan heparan sulphate. J. Med. Microbiol. 1993;38:240–244. [PubMed: 7682621]
10.: Bemelman W. A., Bosma A., Wiersma P. H., Rauws E. A., Brummelkamp W. H. Role of Helicobacter pylori in the pathogenesis of complications of Meckel's diverticula. Eur. J. Surg. 1993;159:171–175. [PubMed: 8102893]
11.: Benaissa M., Babin P., Quellard N., Pezennec L., Cenatiempo Y., Fauchere J. L. Changes in Helicobacter pylori ultrastructure and antigens during conversion from the bacillary to the coccoid form. Infect. Immun. 1996;64:2331–2335. [PMC free article: PMC174074] [PubMed: 8675345]
12.: Berry V., Jennings K., Woodnutt G. Bactericidal and morphological effects of amoxicillin on H. pylori. Antimicrob. Agents Chemother. 1995;39:1859–1861. [PMC free article: PMC162840] [PubMed: 7486933]
13.: Bickley J., Owen R. J., Fraser A. G., Pounder R. E. Evaluation of the polymerase chain reaction for detecting the urease C gene of Helicobacter pylori in gastric biopsy samples and dental plaque. J. Med. Microbiol. 1993;39:338–344. [PubMed: 8246250]
14.: Blom J., Gernow A., Holck S., Wewer V., Nørgaard A., Graff L., Krasilnikoff P. A., Andersen L. P. Different patterns of Helicobacter pylori adherence to gastric mucosa cells. Scand. J. Gastroenterol. 2000;35:1033–1040. [PubMed: 11099055]
15.: Bode G., Malfertheiner P., Ströhle A., Mauch F., Nilius M., Ditschuneit H. Polymorphismus bei Helicobacter pylori—Schlüsselfunktion für Infektionsrezidive? Med. Klin. 1992;87:179–184. [PubMed: 1579096]
16.: Bode G., Mauch F., Ditschuneit H., Malfertheiner P. Identification of structures containing polyphosphate in Helicobacter pylori. J. Gen. Microbiol. 1993;139:3029–3033. [PubMed: 8126429]
17.: Bode G., Mauch F., Malfertheiner P. The coccoid forms of Helicobacter pylori. Criteria for their viability. Epidemiol. Infect. 1993;111:483–490. [PMC free article: PMC2271265] [PubMed: 8270008]
18.: Borén T., Falk P., Roth K. A., Larson G., Normark S. Attachment of Helicobacter pylori to human gastric epitelium mediated by blood group antigens. Science. 1993;262:1892–1895. [PubMed: 8018146]
19.: Borén, T., T. Wadström, S. Normark, J. I. Gordon, and P. Falk. 1997. Methods for the identification of H. pylori host receptors, p. 205–224. In C. L. Clayton and H. L. T. Mobley (ed.), Methods in Molecular Medicine. Helicobacter pylori Protocols. Humana Press Inc., Totowa, N.J. [PubMed: 21351035]
20.: Borhan-Manesh F., Farnum J. B. Study of Helicobacter pylori colonization of patches of heterotopic gastric mucosa (HGM) at the upper esophagus. Dig. Dis. Sci. 1993;38:142–146. [PubMed: 8420747]
21.: Bronsdon M. A., Goodwin C. S., Sly L. I., Chilvers T., Schoenknecht F. D. Helicobacter menestrinae sp. nov., a spiral bacterium found in the stomach of pigtailed macaque (Macaca nemestrina). Int. J. Syst. Bacteriol. 1991;41:148–153. [PubMed: 1995031]
22.: Bryant, M. P. 1984. Genus VII: Anaerobiospirillum Davis, Cleven, Brown and Balish 1976, 503AL, p. 645–646. In N. R. Krieg and J. G. Holt (ed.), Bergey's Manual of Systematic Bacteriology, vol 1. The Williams & Wilkins Co., Baltimore, Md.
23.: Butzler J.-P., Dekeyser P., Detrain M., Dehaen F. Related vibrio in stool. J. Pediatr. 1973;82:493–495. [PubMed: 4572934]
24.: Catrenich C. E., Makin K. M. Characterization of the morphologic conversion of Helicobacter pylori from bacillar to coccoid forms. Scand. J. astroenterol. 1991;26(Suppl. 181):58–64. [PubMed: 1866596]
25.: Catrenich C. E., Chestnut M. H. Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori. J. Med. Microbiol. 1992;37:389–395. [PubMed: 1460658]
26.: Cellini L., Allocati N., Angelucci D., Iezzi T., Di-Campli E., Marzio L., Dainelli B. Coccoid Helicobacter pylori not culturable in vitro reverts in mice. Microbiol. Immunol. 1994;38:843–850. [PubMed: 7898382]
27.: Cellini L., Allocati N., Di-Campli E., Dainelli B. Helicobacter pylori: a fickle germ. Microbiol. Immunol. 1994;38:25–30. [PubMed: 8052159]
28.: Cellini L., Allocati N., Piattelli A., Petrelli I., Fanci P., Dainelli B. Microbiological evidence of Helicobacter pylori from dental plaque in dyspeptic patients. New Microbiol. 1995;18:187–192. [PubMed: 7603346]
29.: Chan W. Y., Hui P. K., Leung K. M., Chow J., Kwok F., Ng C.-S. Coccoid forms of Helicobacter pylori in the human stomach. Am. J. Clin. Pathol. 1994;102:503–507. [PubMed: 7524304]
30.: Conrad R. S., Kist M., Schlecht S., Galanos C. Extraction and biochemical analyses of Helicobacter pylori lipopolysaccaride. Curr. Microbiol. 1992;24:165–169.
31.: Cover T. L., Blaser M. J. Purification and characterization of the vacuolating toxin from Helicobacter pylori. J. Biol. Chem. 267:10570–10575. [PubMed: 1587837]
32.: Cover T. L., Cao P., Lind C. D., Tham K. T., Blaser M. J. Correlation between vacuolating cytotoxin production by Helicobacter pylori isolates in vitro and in vivo. Infect. Immun. 1993;61:5008–5012. [PMC free article: PMC281276] [PubMed: 8225576]
33.: Dent J., McNulty C. A. M. Evaluation of a selective medium for Campylobacter pylori. Eur. J. Clin. Microbiol. Infect. Dis. 1988;7:555–568. [PubMed: 3141172]
34.: Dent J. C., McNulty C. A. M., Uff J. C., Wilkinson S. P., Gear M. W. L. Spiral organisms in the gastric antrum. Lancet. 1987;ii:96. [PubMed: 2885587]
35.: Dewhirst F. E., Seymour C., Fraser G. J., Paster B. J., Fox J. G. Phylogeny of Helicobacter isolates from bird and swine feces and description of Helicobacter pametensis sp. nov. Int. J. Syst. Bacteriol. 44:553–560. [PubMed: 7520743]
36.: Dixon M. F., Genta R. M., Yardley J. H., Correa P. Classification and grading of gastritis. The updated Sydney system. International Workshop on the Histopathology of Gastritis, Houston, 1994. Am. J. Surg. Pathol. 1996;20:1161–1181. [PubMed: 8827022]
37.: Dunn B. E., Roop R., Sung C. C., Sharma S. A., Perez-Perez G. I., Blaser M. J. Identification and purification of a cpn60 heat shock protein homolog from Helicobacter pylori. Infect. Immun. 1992;60:1946–1951. [PMC free article: PMC257099] [PubMed: 1563786]
38.: Dye K. R., Marshall B. J., Frierson H. F., Pambianco D. J., McCallum R. W. Campylobacter pylori colonizing heterotopic tissue in the rectum. Lancet. 1992;340:1194–1195. [PubMed: 2294694]
39.: Eaton K. A., Catrenich C. E., Makin K. M., Krakowka S. Virulence of coccoid and bacillary forms of Helicobacter pylori in gnotobiotic piglets. J. Infect. Dis. 1995;171:459–462. [PubMed: 7844390]
40.: Eaton K. A., Dewhirst F. E., Radin M. J., Fox J. G., Paster B. J., Krakowska S., Morgan D. R. Helicobacter acinonyx sp. nov., isolated from cheetahs with gastritis. Int. J. Syst. Bacteriol. 1993;43:99–106. [PubMed: 8379970]
41.: Eaton K. A., Morgan D. R., Brooks C. L., Krakowka S. Essential role of urease in the pathogenesis of gastritis induced by Helicobacter pylori in gnotobiotic piglets. Infect. Immun. 1991;59:2470–2475. [PMC free article: PMC258033] [PubMed: 2050411]
42.: Eaton K. A., Morgan D. R., Krakowka S. Motility as a factor in the colonisation of gnotobiotic piglets by Helicobacter pylori. J. Med. Microbiol. 1992;37:123–127. [PubMed: 1629897]
43.: Evans D. G., Evans D. J., Moulds J. J., Graham D. Y. N-acetylneuraminyllactose-binding fibrillar hemagglutinin of Campylobacter pylori: a putative colonization factor antigen. Infect. Immun. 1988;56:2896–2906. [PMC free article: PMC259668] [PubMed: 2459065]
44.: Evans, D. J. Jr, Evans D. G., Engstrand L., Graham D. Y. Urease-associated heat shock protein of Helicobacter pylori. Infect. Immun. 1992;60:2125–2127. [PMC free article: PMC257126] [PubMed: 1348725]
45.: Falk P., Roth K. A., Borén T., Westblom T. U., Gordon J. I. An in vitro adherence assay reveals that Helicobacter pylori exhibits cell lineage-specific tropism in the human gastric epithelium. Proc. Natl. Acad. Sci. USA. 1993;90:2035–2039. [PMC free article: PMC46015] [PubMed: 8383333]
46.: Fennell C. L., Totten P. A., Quinn T. C., Patton D. L., Holmes K. K., Stamm W. E. Characterization of Campylobacter-like organisms isolated from homosexual men. J. Infect. Dis. 1984;149:58–66. [PubMed: 6693790]
47.: Figura N. Helicobacter pylori exotoxins and gastroduodenal diseases associated with cytotoxic strain infection. Aliment. Pharmacol. Ther. 1996;10(Suppl. 1):79–96. [PubMed: 8730263]
48.: Fox J. G., Chilvers T., Goodwin C. S., Taylor N. S., Edmonds P., Sly L. I., Brenner D. J. Campylobacter mustelae, a new species resulting from the elevation of Campylobacter pylori subsp. mustelae to species status. Int. J. Syst. Bacteriol. 1989;39:301–303.
49.: Fox J. G., Dewhirst F. E., Tully J. G., Paster B. J., Yan L., Taylor N. S., Collins M. J., Gorelick P. L., Ward J. M. Helicobacter hepaticus sp. nov., a microaerophilic bacterium isolated from liver and intestinal mucosal scrapings from mice. J. Clin. Microbiol. 1994;32:1238–1245. [PMC free article: PMC263656] [PubMed: 8051250]
50.: Fox J. G., Taylor N. S., Edmonds P., Brenner D. J. Campylobacter pylori subsp. mustelae subsp. nov. isolated from the gastric mucosa of ferrets (Mustela putorius furo), and an emended description of Campylobacter pylori. Int. J. Syst. Bacteriol. 1988;38:367–370.
51.: Fox J. G., Yan L. L., Dewhirst F. E., Paster B. J., Shames B., Murphy J. C., Hayward A., Belcher J. C., Mendes E. N. Helicobacter bilis sp. nov., a novel Helicobacter species isolated from bile, livers, and intestine of aged, inbred mice. J. Clin. Microbiol. 1995;33:445–454. [PMC free article: PMC227964] [PubMed: 7536217]
52.: Goodwin C. S., Armstrong J. A., Chilvers T., Peters M., Collins D., Sly L., McConnell W., Harper W. S. Transfer of Campylobacter pylori and Campylobacter mustelae to Helicobacter gen. nov. as Helicobacter pylori comb. nov. and Helicobacter mustelae comb. nov., respectively. Int. J. Syst. Bacteriol. 1989;4:397–405.
53.: Goodwin C. S., Collins M. D., Blinkow E. The absence of thermoplasmaquinones in Campylobacter pyloridis, and its temperature and growth range. Microb. Lett. 1986;32:137–140.
54.: Goodwin C. S., McConnell W., McCullough R. K., McCullough C., Hill R., Bronsdon M. A., Kasper G. Cellular fatty acid composition of Campylobacter pylori from primates and ferrets compared with those of other campylobacters. J. Clin. Microbiol. 1989;27:938–943. [PMC free article: PMC267458] [PubMed: 2745703]
55.: Goodwin S., Blincow E., Armstrong J., McCulloch R. Campylobacter pyloridis is unique; GCLO-2 is an ordinary campylobacter. Lancet. 1985;ii:38–39. [PubMed: 2861477]
56.: Gribbon L. T., Barer M. R. Oxidative metabolism in nonculturable Helicobacter pylori and Vibrio vulnificus cells studied by substrate-enhanced tetrazolium reduction and digital image processing. Appl. Environ. Microbiol. 1995;61:3379–3384. [PMC free article: PMC167617] [PubMed: 7574647]
57.: Gubbins G. P., Moritz T. E., Marsano L. S., Talwalkar R., McClain C. J., Mendenhall C. L. Helicobacter pylori is a risk factor for hepatic encephalopathy in acute alcoholic hepatitis: the ammonia hypothesis revisited. The Veterans Administration Cooperative Study Group No. 275. Am. J. Gastroenterol. 1993;88:1906–1910. [PubMed: 8237940]
58.: Hanninen M. L., Happonen I., Saari S., Jalava K. Culture and characteristics of Helicobacter bizzozeronii, a new canine gastric Helicobacter sp. Int. J. Syst. Bacteriol. 1996;46:160–166. [PubMed: 8573490]
59.: Hawtin P. R., Stacey A. R., Newell D. G. Investigation of the structure and location of the urease of Helicobacter pylori using monoclonal antibodies. J. Gen. Microbiol. 1990;136:1995–2000. [PubMed: 2269872]
60.: Hazell S. L., Evans, Jr D. J., Graham D. Y. Helicobacter pylori catalase. J. Gen. Microbiol. 1991;137:57–61. [PubMed: 2045782]
61.: Hazell S. L., Lee A., Brady L., Hennessy W. Campylobacter pyloridis and gastritis: association with inter-cellular spaces and adaptation to an environment of mucus as important factors in colonization of the gastric epithelium. J. Infect. Dis. 1986;153:658–663. [PubMed: 3950447]
62.: Hazell S. L., Markesich D. C., Evans D. J., Evans D. G., Graham D. Y. Influence of media supplements on growth and survival of Campylobacter pylori. Eur. J. Clin. Microbiol. Infect. Dis. 1989;8:597–602. [PubMed: 2506019]
63.: Heilmann, K. L., and F. Borchard. 1991. Further observations on human spirobacteria, p. 63–73. In H. Menge, M. Gregor, G. N. I. Tytgat, B. I. Marshall, and C. A. M. McNulty (ed.), Helicobacter pylori 1990. Springer Verlag, Berlin, Germany.
64.: Hessay S. J., Spencer J., Wyatt J. I., Sobala G., Rathbone B. J., Axon A. T. R., Dixon M. F. Bacterial adhesion and disease activity in Helicobacter associated chronic gastritis. Gut. 1990;31:134–138. [PMC free article: PMC1378366] [PubMed: 2311970]
65.: Hirmo S., Litt M., Ringer M., Wadström T. Inhibition of heparan sulphate and other glycosaminoglycans binding to Helicobacter pylori by various polysulphated carbohydrates. FEMS Immunol. Med. Microbiol. 1995;10:301–306. [PubMed: 7539671]
66.: Holck S., Ingeholm P., Blom J., Nørgaard A., Elsborg L., Adamsen S., Andersen L. P. The histopathology of human gastric mucosa inhabited by Helicobacter heilmannii-like (Gastrospirillum hominis) organisms including one culturable case. APMIS. 1997;105:746–756. [PubMed: 9368589]
67.: Hua J., Ho B. Is the coccoid form of H. pylori viable? Microbios. 1996;87:103–112. [PubMed: 9032959]
68.: Hulten K., Han S. W., Enroth H., Klein P. D., Opekun A. R., Gilman R. H., Evans D. G., Engstrand L., Graham D. Y., El-Zaatari F. A. Helicobacter pylori in the drinking water in Peru. Gastroenterology. 1996;110:1031–1035. [PubMed: 8612990]
69.: Hynes S. O., Hirmo S., Wadstrom T., Moran A. P. Differentiation of Helicobacter pylori isolates based on lectin binding of cell extracts in an agglutination assay. J. Clin. Microbiol. 1999;37:1994–1998. [PMC free article: PMC85005] [PubMed: 10325361]
70.: Ikeda F., Yokota Y., Mine Y., Tatsuta M. Activity of cefixime against Helicobacter pylori and affinities for the penicillin-binding protein. Antimicrob. Agents Chemother. 1990;24:2426–2428. [PMC free article: PMC172075] [PubMed: 2088199]
71.: Jalava K., Kaartinen M., Utriainen M., Happonen I., Hanninen M. L. Helicobacter salomonis sp. nov., a canine gastric Helicobacter sp. related to Helicobacter felis and Helicobacter bizzozeronii. Int. J. Syst. Microbiol. 1997;47:975–982. [PubMed: 9336895]
72.: Janas B., Czkwianianc E., Bak-Romaniszyn L., Bartel H., Tosik D., Planeta-Malecka I. Electron microscopic study of association between coccoid forms of Helicobacter pylori and gastric epithelial cells. Am. J. Gastroenterol. 1995;90:1829–1833. [PubMed: 7572903]
73.: Jiang S. J., Liu W. Z., Zhang D. Z., Shi Y., Xiao S. D., Zhang Z. H., Lu D. Y. Campylobacter-like organisms in chronic gastritis, peptic ulcer, and gastric carcinoma. Scand. J. Gastroenterol. 1987;22:553–558. [PubMed: 3629180]
74.: Jiesong H., Megraud F. Evidence of the viability of nonculturable coccoidal forms of Helicobacter felis. Gut. 1995;37(Suppl. 1):376.
75.: Jones D. M., Curry A., Fox A. J. An ultrastructural study of the gastric Campylobacter-like organism "Campylobacter pyloridis.". J. Gen. Microbiol. 1985;131:2335–2341. [PubMed: 4067580]
76.: Jones D. M., Lessels A. M., Elridge J. Campylobacter-like organisms on the gastric mucosa: culture, histological, and serological studies. J. Clin. Pathol. 1984;37:1002–1006. [PMC free article: PMC498915] [PubMed: 6470176]
77.: Kamisago S., Iwamori M., Tai T., Mitamura K., Yazaki Y., Sugano K. Role of sulfatides in adhesion of Helicobacter pylori to gastric cancer cells. Infect. Immun. 1996;64:624–628. [PMC free article: PMC173811] [PubMed: 8550217]
78.: Kasper G., Dickgiesser N. Isolation from gastric epithelium of Campylobacter-like bacteria that are distinct from "Campylobacter pyloridis.". Lancet. 1985;i:111–112. [PubMed: 2857011]
79.: Kelly S. M., Pitcher M. C. L., Farmery S. M., Gibson G. R. Isolation of Helicobacter pylori from feces of patients with dyspepsia in the United Kingdom. Gastroenterology. 1994;107:1671–1674. [PubMed: 7958677]
80.: Khin M. M., Ringer M., Alejung P., Wadström T., Ho B. Binding of human plasminogen and lactoferrin by Helicobacter pylori coccoid forms. J. Med. Microbiol. 1996;45:433–439. [PubMed: 8958246]
81.: King E. O. Human infections with Vibrio fetus and a closely related vibrio. J. Infect. Dis. 1957;101:119–128. [PubMed: 13475869]
82.: Kjøller M., Fischer A., Justesen T. Transport conditions and number of biopsies necessary for culture of Helicobacter pylori. Eur. J. Clin. Microbiol. Infect. Dis. 1991;10:166–167. [PubMed: 2060517]
83.: Krajden S., Bohnen J., Anderson J., Kempston J., Fuksa M., Matlow A., Marcon N., Haber G., Kortan P., Karmali M., Corey P., Petrea C., Babida C., Hayman S. Comparison of selective and nonselective media for recovery of Campylobacter pylori from antral biopsies. J. Clin. Microbiol. 1987;25:1117–1118. [PMC free article: PMC269150] [PubMed: 3597756]
84.: Labigne A., Cussac V., Courcoux P. Shuttle cloning and nucleotide sequences of Helicobacter pylori genes responsible for urease activity. J. Bacteriol. 1991;173:1920–1931. [PMC free article: PMC207722] [PubMed: 2001995]
85.: Langenberg M. L., Tytgat G. N. J., Schipper M. E. I., Rietra P. J. G. M., Zanen H. C. Campylobacter-like organisms in the stomach of patients and individuals. Lancet. 1984;i:1348. [PubMed: 6145040]
86.: Langton S. R., Cesareo S. D. Helicobacter pylori associated phospholipase A2 activity: a factor in peptic ulcer production? J. Clin. Pathol. 1992;45:221–224. [PMC free article: PMC495476] [PubMed: 1556229]
87.: La Riviére, J. W. M., and J. G. Kuenen. 1989. Genus Thiovulum Hinze 1913, 195AL, p. 1841–1842. In M. Stalay, P. Bryant, N. Pfenning, and J. G. Holt (ed.), Bergey's Manual of Systematic Bacteriology, vol. 3. The Williams & Wilkins Co., Baltimore, Md.
88.: Lee A., O'Rourke J. Gastric bacteria other than Helicobacter pylori. Gastroenterol. Clin. North Am. 1993;22:21–42. [PubMed: 8449568]
89.: Lee A., Phillips M. W., O'Rourke J. L., Paster B. J., Dewhirst F. E., Fraser G. J., Fox J. G., Sly L. I., Romaniuk P. J., Trust T. J., Kouprach S. Helicobacter muridarum sp. nov., a microaerophilic helical bacterium with a novel ultrastructure isolated from the intestinal mucosa of rodents. Int. J. Syst. Bacteriol. 1992;42:27–36. [PubMed: 1736969]
90.: Loffeld R. J., Ten-Tije B. J., Arends J. W. Prevalence and significance of Helicobacter pylori in patients with Barrett's esophagus. Am. J. Gastroenterol. 1992;87:1598–1600. [PubMed: 1442681]
91.: Marshall B. J., Goodwin C. S. Revised nomenclature of Campylobacter pyloridis. Int. J. Syst. Bacteriol. 1987;37:68.
92.: Marshall B. J., McGechie D. B., Rogers P. A., Glancy R. J. Pyloric Campylobacter infection and gastroduodenal disease. Med. J. Australia. 1985;142:439–444. [PubMed: 3982346]
93.: Marshall B. J., Royce H., Annear D. I., Goodwin C. S., Pearman J. W., Warren J. R., Armstrong J. A. Original isolation of Campylobacter pyloridis from human gastric mucosa. Microbios Lett. 1984;25:83–88.
94.: Marshall B. J., Warren J. R. Unidentified curved bacilli in the stomach of patients with gastritis and peptic ulceration. Lancet. 1984;i:1311–1315. [PubMed: 6145023]
95.: Mattsby-Baitzer I. Z. Mielniczuk, Larson L., Lindgren K., Goodwin S. Lipid A in Helicobacter pylori. Infect. Immun. 1992;60:4383–4387. [PMC free article: PMC257475] [PubMed: 1398948]
96.: Mendes E. N., Queiroz D. M., Dewhirst F. E., Paster B. J., Moura S. B., Fox J. G. Helicobacter trogontum sp. nov., isolated from the rat intestine. Int. J. Syst. Bacteriol. 1996;46:916–921. [PubMed: 8863417]
97.: Miller-Podraza H. J. Bergstrom, Teneberg S., Milh M. A., Longard M., Olsson B. M., Uggla L., Karlsson K. A. Helicobacter pylori and neutrophils: sialic acid-dependent binding to various isolated glycoconjugates. Infect. Immun. 1999;67:6309–6313. [PMC free article: PMC97034] [PubMed: 10569742]
98.: Minocha A., Mokshagundam S., Gallo S. H., Rahal P. S. Alterations in upper gastrointestinal motility in Helicobacter pylori-positive nonulcer dyspepsia. Am. J. Gastroenterol. 1994;89:1797–1800. [PubMed: 7942670]
99.: Mizogushi H., Fujioka T., Kishi K., Nishizono A., Kodama R., Nasu M. Diversity in protein synthesis and viability of Helicobacter pylori coccoid forms in response to various stimuli. Infect. Immun. 1998;66:5555–5560. [PMC free article: PMC108699] [PubMed: 9784573]
100.: Mobley H. L. T., Cortesia M. J., Rosenthal L. E., Jones B. D. Characterization of urease from Campylobacter pylori. J. Clin. Microbiol. 1988;26:831–836. [PMC free article: PMC266469] [PubMed: 3384908]
101.: Montgomery E. A., Martin D. F., Peura D. A. Rapid diagnosis of Campylobacter pylori by Gram's stain. Am. J. Pathol. 1988;90:606–609. [PubMed: 2459961]
102.: Moran A. P., Appelmelk B. J., Aspinall G. O. Molecular mimicry of host structures by lipopolysaccharides of Campylobacter and Helicobacter spp.: implications in pathogenesis. J. Endotox. Res. 1996;3:521–531.
103.: Moran A. P., Kuusela P., Kosunen T. U. Binding of Helicobacter pylori to extracellular matrix proteins. Ital. J. Gastroenterol. 1991;23(Suppl. 2):7.
104.: Morris A., Nicholson G., Zwi J., Vanderwee M. Campylobacter pylori infection in Meckel's diverticula containing gastric mucosa. Gut. 1989;30:1233–1235. [PMC free article: PMC1434255] [PubMed: 2806990]
105.: Moshcowitz M., Corea A., Arber N., Konikoff F., Berger S., Gilat T. Morphological transformation of Helicobacter pylori during prolonged incubation: association with decreased acid resistance. J. Clin. Pathol. 1994;47:172–174. [PMC free article: PMC501837] [PubMed: 8132834]
106.: Moutiala A., Moutiala A., Helander I. M., Pyhälä L., Kosunen T. U., Moran A. P. Low biological activity of Helicobacter pylori lipopolysaccharide. Infect. Immun. 1992;60:1714–1716. [PMC free article: PMC257055] [PubMed: 1548097]
107.: Murray L. J., Bamford K. B., O'Reilly D. P., McCrum E. E., Evans A. E. Helicobacter pylori infection: relation with cardiovascular risk factors, ischaemic heart disease, and social class. Br. Heart J. 1995;74:497–501. [PMC free article: PMC484068] [PubMed: 8562233]
108.: Nederskov-Sørensen P., Aase S., Bjørneklett A., Fausa O., Bukholm G. Sampling efficiency in the diagnosis of Helicobacter pylori infection and chronic active gastritis. J. Clin. Microbiol. 1991;29:672–675. [PMC free article: PMC269851] [PubMed: 1890167]
109.: Nederskov-Sørensen P., Bjørneklett A., Fausa O., Bukholm G., Aase S., Jantzen E. Campylobacter pylori infection and its relation to chronic gastritis. An endoscopic, bacteriologic, and histomorphologic study. Scand. J. Gastroenterol. 1988;23:867–874. [PubMed: 3067331]
110.: Neithercut W. D., Greig M. A., Hossack M., McColl K. E. L. Suicidal destruction of Helicobacter pylori: metabolic consequence of intracellular accumulation of ammonia. J. Clin. Pathol. 1991;44:380–384. [PMC free article: PMC496867] [PubMed: 2045495]
111.: Nichols L., Sughayer M., DeGirolami P. C., Baloch K., Pleskow D., Eichelberger K., Santos M. Evaluation of diagnostic methods for Helicobacter pylori gastritis. Am. J. Pathol. 1991;95:769–773. [PubMed: 2042584]
112.: Nielsen H., Birkholz S., Andersen L. P., Moran A. Neutrophil activation by Helicobacter pylori lipopolysaccharides. J. Infect. Dis. 1994;170:135–139. [PubMed: 8014489]
113.: Nilsson, H. O., J. Blom, M. Stollenwerk, L. P. Andersen, and T. Wadström. Ultrastructure and ATP content of bacillary and coccoid forms of Helicobacter pylori. Submitted for publication.
114.: Nilsson H. O., Taneera J., Castedal M., Glatz E., Olsson R., Wadstrom T. Identification of Helicobacter pylori and other Helicobacter species by PCR, hybridization, and partial DNA sequencing in human liver samples from patients with primary sclerosing cholangitis or primary biliary cirrhosis. J. Clin. Microbiol. 2000;38:1072–1076. [PMC free article: PMC86342] [PubMed: 10698999]
115.: Noach L. A., Rolf T. M., Tytgat G. N. Electron microscopic study of association between Helicobacter pylori and gastric and duodenal mucosa. J. Clin. Pathol. 1994;47:699–704. [PMC free article: PMC502139] [PubMed: 7962619]
116.: Olivieri R., Bugnoli M., Armellini D., Bianciardi S., Rappuoli R., Bayeli P. F., Abate L., Esposito E., De Gregorio L., Aziz J., Basagni C., Figura N. Growth of Helicobacter pylori in media containing cyclodextrins. J. Clin. Microbiol. 1993;31:160–162. [PMC free article: PMC262644] [PubMed: 8417026]
117.: Osaki T., Yamaguchi H., Taguchi H., Kumada J., Ogata S., Kamiva S. Studies on the relationship between adhesive activity and hemagglutination by Helicobacter pylori. J. Med. Microbiol. 1997;46:117–121. [PubMed: 9060870]
118.: Parsonnet J., Welch K., Compton C., Strauss R., Wang T., Kelsey P., Ferraro M. J. Simple microbiological detection of Campylobacter pylori. J. Clin. Microbiol. 1988;26:948–949. [PMC free article: PMC266492] [PubMed: 3384915]
119.: Paster B. J., Dewhirst F. E. Phylogeny of campylobacters, wollinellas, Bacteroides gracilis, and Bacteroides urealyticus by 16S ribosomal ribonucleic acid sequensing. Int. J. Syst. Bacteriol. 1988;38:56–62.
120.: Paster B. J., Lee A., Fox J. G., Dewhirst F. E., Tordoff L. A., Fraser G. J., O'Rourke J. L., Taylor N. S., Ferrero R. Phylogeny of Helicobacter felis sp. nov., Helicobacter mustelae, and related bacteria. Int. J. Syst. Bacteriol. 1991;41:31–38. [PubMed: 1704791]
121.: Patel P., Mendall M. A., Carrington D., Strachan D. P., Leatham E., Molineaux N., Levy J., Blakeston C., Seymour C. A., Camm A. J., Northfield T. C. 1995Association of Helicobacter pylori and Chlamydia pneumoniae infections with coronary heart disease and cardiovascular risk factors Br. Med. J 311711–714.. (Erratum, 311:985.) [PMC free article: PMC2550716] [PubMed: 7549683]
122.: Paull G., Yardley J. H. Gastric and esophageal Campylobacter pylori in patients with Barrett's esophagus. Gastroenterology. 1988;95:216–218. [PubMed: 3371618]
123.: Perez-Perez G. I., Olivares A. Z., Cover T. L., Blaser M. J. Characteristics of Helicobacter pylori variants selected for urease deficiency. Infect. Immun. 1992;60:3658–3663. [PMC free article: PMC257374] [PubMed: 1500174]
124.: Petersson C., Larsson B., Mahdavi J., Boren T., Magnusson K. E. A new method to visualize the Helicobacter pylori-associated Lewis(b)-binding adhesin utilizing SDS-digested freeze-fracture replica labeling. J. Histochem. Cytochem. 2000;48:877–883. [PubMed: 10820161]
125.: Pinkard K. J., Harrison B., Capstick J. A., Medley G., Lambert J. R. Detection of Campylobacter pyloridis in gastric mucosa by phase contrast microscopy. J. Clin. Pathol. 1986;39:112–113. [PMC free article: PMC499625] [PubMed: 2419365]
126.: Piotrowski J., Piotrowski E., Skrodzka A., Slomiany A., Slomiany B. L. Induction of gastritis and epithelial apoptosis by Helicobacter pylori lipopolysaccharide. Scand. J. Gastroenterol. 1997;32:203–211. [PubMed: 9085455]
127.: Rieggs, S. J., B. E. Dunn, and M. J. Blaser. 1995. Microbiology and pathogenesis of Helicobacter pylori, p. 535–563. In M. J. Blaser, P. D. Smith, J. I. Ravdin, R. L. Greenberg, and R. L. Guerrant (ed.), Infection of the Gastrointestinal Tract. Raven Press, New York, N.Y.
128.: Roine R. P., Salmela K. S., Höök-Nikanne J., Kosunen T. U., Salaspuro M. Alcohol dehydrogenase mediated acetaldehyde production by Helicobacter pylori—a possible mechanism behind gastric injury. Life Sci. 1992;51:1333–1337. [PubMed: 1406052]
129.: Roosendaal R., Kuipers E. J., Peña A. S., de Graaff J. Recovery of Helicobacter pylori from biopsy specimens is not dependent on the transport medium used. J. Clin. Microbiol. 1995;33:2798–2800. [PMC free article: PMC228582] [PubMed: 8567932]
130.: Saitoh T., Natomi H., Zhao W. L., Okuzumi K., Sugano K., Iwamori M., Nagai Y. Identification of glycolipid receptors for Helicobacter pylori by TLC-immunostaining. FEBS Lett. 1991;282:385–387. [PubMed: 2037054]
131.: Schmitt W., Hass R. Genetic analysis of Helicobacter pylori vacuolating cytotoxin: structural similarities with the IgA protease type of exported protein. Mol. Microbiol. 1994;12:308–319. [PubMed: 8057855]
132.: Scragg R. K., Fraser A., Metcalf P. A. Helicobacter pylori seropositivity and cardiovascular risk factors in a multicultural workforce. J. Epidemiol. Community Health. 1996;50:578–579. [PMC free article: PMC1060354] [PubMed: 8944869]
133.: Sebald M., Véron M. Teneur en bases de l'ADN et classification des vibrions. Ann. Inst. Pasteur (Paris). 1963;105:897–910. [PubMed: 14098102]
134.: Segal E., Falkow S., Thompkins L. S. Helicobacter pylori attachment to gastric cells induces cytoskeletal rearrangement and tyrosine phosphorylation of host cell proteins. Proc. Natl. Acad. Sci. USA. 1996;93:1259–1264. [PMC free article: PMC40067] [PubMed: 8577751]
135.: Sidebotham R. L., Batten J. J., Karim Q. N., Spencer J., Baron J. H. Breakdown of gastric mucus in presence of Helicobacter pylori. J. Clin. Pathol. 1991;44:52–57. [PMC free article: PMC497015] [PubMed: 1997534]
136.: Skirrow M. B. Campylobacter enteritidis: a "new" disease. Br. Med. J. 1977;2:9–11. [PMC free article: PMC1631297] [PubMed: 871765]
137.: Slomiany B. L., Murty V. L. N., Piotrowski J., Liau Y. H., Sundaram P., Slomiany A. Glycosulfatase activity of Helicobacter pylori toward gastric mucin. Biochem. Biophys. Res. Comm. 1992;183:506–513. [PubMed: 1372502]
138.: Smith T., Taylor M. S. Some morphological and biochemical characters of the spirilla (Vibrio fetus n. sp.) associated with disease of the fetal membranes in cattle. J. Exp. Med. 1919;30:299–312. [PMC free article: PMC2126685] [PubMed: 19868360]
139.: Syder A. J., Guruge J. L., Li Q., Hu Y., Olcksiewicz C. M., Lorenz R. G., Karam S. M., Falk P. G., Gordon J. I. Helicobacter pylori attaches to NeuAc alpha 2,3Gal beta 1,4 glycoconjugates produced in the stomach of transgenic mice lacking parietal cells. Mol. Cell. 1999;3:263–274. [PubMed: 10198629]
140.: Solnick J. V., O'Rourke J., Lee A., Paster B. J., Dewhirst F. E., Tompkins L. S. An uncultured gastric spiral organism is a newly identified Helicobacter in humans. J. Infect. Dis. 1993;168:379–385. [PubMed: 8335974]
141.: Soltesz V., Zeeberg B., Wadström T. Optimal survival of Helicobacter pylori under various transport conditions. J. Clin. Microbiol. 1992;30:1453–1456. [PMC free article: PMC265309] [PubMed: 1624562]
142.: Sörberg M., Nilsson M., Hanberger H., Nilsson L. E. Morphologic conversion of Helicobacter pylori from baccilar to coccoid form. Eur. J. Clin. Microbiol. Infect. Dis. 1996;15:216–219. [PubMed: 8740856]
143.: Spigelhalder C., Gerstenecker B., Kersten A., Schiltz E., Kist M. Purification of Helicobacter pylori superoxide dismutase and cloning and sequencing of the gene. Infect. Immun. 1993;61:5315–5325. [PMC free article: PMC281317] [PubMed: 8225605]
144.: Stanley J., Linton D., Burnens A. P., Dewhirst F. E., Owen R. J., Porter A., On S. L. W., Costas M. Helicobacter canis sp. nov., a new species from dogs: an integrated study of phenotype and genotype. J. Gen. Microbiol. 1993;139:2495–2504. [PubMed: 8254320]
145.: Steinhilber W., Loeffler S., Siebler A., Rektorschek M., Melchers K., Schäfer K. P. Lipid composition is changed in coccoids from Helicobacter pylori and Helicobacter felis. Gut. 1996;39(Suppl. 2):61–62.
146.: Talley N. J., Cameron A. J., Shorter R. G., Zinsmeister A. R., Phillips S. F. Campylobacter pylori and Barrett's esophagus. Mayo Clin. Proc. 1988;63:1176–1180. [PubMed: 3199885]
147.: Telford J., Ghiara P., Dell'Orco M., Comanducci M., Burroni D., Bugnoli M., Tecce M. F., Censini S., Covacci A., Xiang Z., Papini E., Montecucco C., Parente L., Rappuoli R. Gene structure of the Helicobacter pylori cytotoxin and evidence of its key role in gastric disease. J. Exp. Med. 1994;179:1653–1658. [PMC free article: PMC2191472] [PubMed: 8163943]
148.: Thomas J. E., Gibson G. R., Darboe M. K., Dale A., Weaver L. T. Isolation of Helicobacter pylori from human faeces. Lancet. 1992;ii:1194–1195. [PubMed: 1359263]
149.: Tompkins D. S., Dave J., Mapstone N. P. Adaptation of Helicobacter pylori to aerobic growth. Eur. J. Clin. Microbiol. Infect. Dis. 1994;13:409–412. [PubMed: 8070455]
150.: Tzouvelekis L. S., Mentis A. F., Makris A. M., Spiliadis C., Blackwell C., Weir D. M. In vitro binding of Helicobacter pylori to human gastric mucin. Infect. Immun. 1991;59:4252–4254. [PMC free article: PMC259024] [PubMed: 1937781]
151.: Utt M., Wadström T. Identification of heparan sulphate binding surface proteins of Helicobacter pylori: inhibition of heparan sulphate binding with sulphated carbohydrate polymers. J. Med. Microbiol. 1997;46:541–546. [PubMed: 9236737]
152.: Vandamme P., Daneshvar M. I., Dewhirst F. E., Paster B. J., Kersters K., Goosens H., Moss C. W. Chemotaxonomic analyses of Bacteroides grasilis and Bacteroides ureolyticus and reclassification of B. gracilis as Campylobacter grasilis comb. nov. Int. J. Syst. Bacteriol. 1995;45:145–152. [PubMed: 7857794]
153.: Vandamme P., De Ley J. Proposal for a new family, Campylobacteriaceae. Int. J. Syst. Bacteriol. 1991;41:451–455.
154.: Vandamme P., Falsen E., Rossau R., Hoste B., Segers P., Tytgat R., DeLey J. Revision of Campylobacter, Helicobacter, and Wollinella taxonomy: emendatum of genetic descriptions and proposals of Arcobacter gen. nov. Int. J. Syst. Bacteriol. 1991;41:88–103. [PubMed: 1704793]
155.: Véron M., Chaterlain R. Taxonomic study of the genus Campylobacter Sebald and Véron and designation of the neotype strain for the type species Campylobacter fetus (Smith and Taylor) Sebald and Véron. Int. J. Syst. Bacteriol. 1973;23:122–134.
156.: Von-Recklinghausen G., Weischer T., Ansorg R., Mohr C. No cultural detection of Helicobacter pylori in dental plaque. Zentralbl. Bakteriol. 1994;281:102–106. [PubMed: 7803925]
157.: Wadstrom T., Hirmo S., Borén T. Biochemical aspects of Helicobacter pylori colonization of the human gastric mucosa. Aliment. Pharmacol. Ther. 1996;1:17–27. [PubMed: 8730256]
158.: Wadström T., Ljungh Å. Glycosaminoglycan-binding microbial proteins in tissue adhesion and invasion: key events in microbial pathogenicity. J. Med. Microbiol. 1999;48:223–233. [PubMed: 10334589]
159.: Wang X., Sturegård E., Rupar R., Nilsson H. O., Aleljung P. A., Carlen B., Willen R., Wadström T. Infection of BALB/cA mice by spiral and coccoid forms of Helicobacter pylori. J. Med. Microbiol. 1997;46:657–663. [PubMed: 9511813]
160.: West A. P., Millar M. R., Tompkins D. S. Survival of Helicobacter pylori in water and saline. J. Clin. Pathol. 1990;43:609. [PMC free article: PMC502595] [PubMed: 2199542]
161.: Westblom T. U., Madan E., Midkiff B. R. Egg yolk emusion agar, a new medium for the cultivation of Helicobacter pylori. J. Clin. Microbiol. 1991;29:819–821. [PMC free article: PMC269878] [PubMed: 1890184]
162.: Westblom T. U., Medan E., Midkiff B. R., Ackins V. W., Subik M. A. Failure of Campylobacter pylori to grow in commercial blood culture systems. J. Clin. Microbiol. 1988;26:1029–1030. [PMC free article: PMC266510] [PubMed: 3384893]
163.: Westblom T. U., Phadnis S., Yang P., Czinn S. J. Diagnosis of Helicobacter pylori infection by means of a polymerase chain reaction assay for gastric juice aspirates. Clin. Infect. Dis. 1993;16:367–371. [PubMed: 8452948]
164.: Wolin M. J., Wolin E. A., Jacobs N. J. Cytochrome-producing anaerobic vibrio, Vibrio succinogenes, sp. n. J. Bacteriol. 1961;81:911–917. [PMC free article: PMC314759] [PubMed: 13786398]

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