History

Microorganisms have been observed in the stomachs of animals and humans for more than 100 years. The first sighting has been credited to Rappin in 1881 (86) who observed spiral-shaped bacteria in a dog's stomach. Bizzozero (6) confirmed Rappin's discovery in 1892 and noted that these bacteria were associated with parietal cells of a dog stomach. This was the first observation of bacteria being associated with distinct cells of the stomach. However, it was not until 1896 that the first comprehensive study of spiral bacteria inhabiting the gastric mucosa was undertaken (89). Hugo Salomon observed spiral-shaped bacteria in the stomachs of dogs, cats, and a wide variety of animals. He noted that there were many different types of spiral-shaped organisms, a number of which had a similar morphology to spirochetes. Salomon reported that these spiral organisms inhabited the gastric pits of the pyloric and fundic glands, with some of these bacteria closely associated with the acid-producing parietal cells and in some cases inside the canuliculi of these cells. Attempts to culture these spiral bacteria on artificial media were unsuccessful, but Salomon was able to propagate them by feeding gastric mucus from infected animals to uninfected white mice. Many of the in vivo culture techniques that were undertaken by Salomon are still widely used today, especially for the isolation and characterization of gastric helicobacters that have been unable to be cultivated in vitro, a classic example being "Helicobacter heilmannii" (19). Over the next few years other investigators reported similar spiral-shaped bacteria in a variety of animals, mostly dogs and cats. These sightings were not without their critics; in 1909 Carnot and Lelievre disagreed with the work of Regaud (in the same year), stating that what he and others had actually observed were secretions from parietal cells and not microorganisms (8). This disagreement continued until Regaud finally convinced the skeptics that what he had observed were actually spiral-shaped bacteria and not parietal cell secretions (87). Interestingly, we now know that these bacteria can enter the canaliculi of parietal cells so the confusion was understandable. Reports of spiral bacteria colonizing the stomach continued over the next few years. Balfour in 1906 reported spiral bacteria near ulcerated lesions (3). Kasai and Kobayashi in 1919 (52) were able to repeat the earlier work undertaken by Salomon. Lim in 1920 added that some of the spiral-shaped organisms could be seen in the duodenum, close to the pyloric sphincter as well as in the pylorus, fundus, and the cardia from a cat (68).

The first human sighting has been credited to Krienitz (55), but it was not until 1939, when an extensive study of these organisms in humans and the macacus rhesus monkey was carried out by Doenges (21). Doenges found that 43% of 242 human stomachs observed at postmortem contained spirochetes as did all 19 monkeys. Many of the spirochetes were associated with parietal cells but never with goblet or pepsin-producing chief cells. Doenges also reported that these organisms were never found in the intestine. Most of the areas of infection were devoid of distinctive clinical symptoms, but Doenges observed that some parietal cells had shrunken nuclei, vacuolated or granulated cytoplasm, which he thought were probably due to the bacterial infection of the cells. He also noticed that infection of parietal cells was patchy, with some parietal cells being full of organisms whereas other adjacent cells were uninfected. Throughout the 20th century there have been many sightings of spiral bacteria colonizing the gastric mucosa of animals and humans, some suggesting their role in gastric pathology. Despite these observations, the stomach was still considered a sterile environment until the discovery of H. pylori.

Since the isolation and characterization of H. pylori, renewed interest in gastric microbiology has led to the reappraisal of spiral mucus-associated bacteria and their possible links with disease. Some of these gastric spirals, e.g., Helicobacter mustelae and Helicobacter acinonychis, were discovered in response to a particular pathology, whereas others were discovered when trying to investigate the microecosystems of the gastric mucosa of different animals (26, 34, 40, 43, 49, 56, 62). New and more sophisticated techniques, especially in the field of molecular biology, have led to the discovery of many new organisms (of which a large proportion have now been classified as Helicobacter species) that inhabit the stomach, lower bowel, and liver. Many in the past were missed due to ineffective culturing techniques or were in low enough number that they were not observed during histopathological analysis.

"Helicobacter heilmannii" ("Gastrospirillum hominis"/"Candidatus Helicobacter suis")

Morphology

"Helicobacter heilmannii" is a tightly spiraled gram-negative bacterium. The organism varies from 4 to 8 turns and is between 4 and 10 μm in length and 0.5 to 1 μm wide (46, 71) (Fig. 1). The bacterium is vigorously motile by means of 12 to 20 sheathed bipolar flagella (65).

Figure 1

"Helicobacter heilmannii" in (A) the gastric mucosa of a mouse and (B and C) the canaliculi of a rat parietal cell. Arrows indicate the position of the bacterium.

Nomenclature

Many of the tightly spiraled bacteria (or spirochetes) that colonize the stomachs of animals (in particular dogs and cats) are now referred to as "H. heilmannii" in honor of the late German histopathologist Konrad Heilmannii (83), who compiled one of the most extensive studies of this bacterium's infection in humans. "H. heilmannii" was originally referred to as "Gastrospirillum hominis" by McNulty and coworkers in 1989 (71) when they found a tightly spiraled bacterium associated with six cases of type B gastritis in humans. This bacterium was later shown to be a Helicobacter species by 16s rRNA (93) and the name "Helicobacter heilmannii" was proposed. Mendes and coworkers in 1990 described a bacterium similar to that of McNulty, from the pig gastric mucosa, and referred to this bacterium as "Gastrospirillum suis" (72). In 1999 De Groote and coworkers proposed the name of the pig isolate as "Candidatus Helicobacter suis" (16, 17).

Solnick and coworkers suggested that there were two subgroups, one isolated fropm humans and the other isolated from animals. This group of bacteria has been recently split into two types. "H. heilmannii" type 1 includes isolates from humans, monkeys, and pigs (i.e., "Candidatus Helicobacter suis"). The 16 rRNA from all these isolates cluster closely together (82). "H. heilmannii" type 2 has been isolated from primarily cats and dogs, and their 16s rRNA cluster closely with other helicobacters such as H. felis, H. salomonis, and H. bizzozeronii (82).

Natural habitat

"H. heilmannii" has the widest host range of any gastric helicobacter. It has been observed colonizing the gastric mucosa of animals for more than 100 years. Many of the early sightings of gastric spirals undoubtedly were "H. heilmannii" (52, 68, 89). This organism has been shown to naturally colonize dogs, cats, monkeys, pigs, and humans. Experimentally this bacterium colonizes rodents (19) and has been used extensively as an animal model for H. pylori infection. "H. heilmannii" is well adapted to the gastric mucosa, able to colonize the length of the stomach from the cardia to the pylorus. It will preferentially colonize the length of the gastric pits. Like many of the other gastric helicobacters, it is free living and does not attach to the epithelial cell layer. One of its attributes is that it is extremely acid resistant, inferred from its ability to colonize the acid-secreting parietal cell (Fig. 1). Interestingly, this bacterium in the rodent will out-compete other gastric Helicobacter spp. such as H. pylori and H. felis (67).

Growth requirements

"H. heilmannii" is an extremely fastidious bacterium that has eluded successful culture. To date, this bacterium has been studied by passaging the organism in mice, a technique that was originally described by Salomon (89) and further developed by Dick et al. (19). It was this in vivo culture that enabled Solnick to characterize and provisionally name the organism "H. heilmannii" and for Mendes et al. to characterize their isolate "Candidatus Helicobacter suis" (74, 93). There has been one report by Holck et al. of successful in vitro culture of "H. heilmannii" (48). In a follow-up publication they reported on the growth conditions (1); however, it must be stressed that attempts to repeat in vitro culture using Holck's method have been unsuccessful. They mentioned the following: " `H. heilmannii" is a microaerophilic bacterium that has a growth range between 36 and 41°C when grown on 7% lysed, defibrinated horse blood agar plates. The small translucent colonies were apparent within 3 to 7 days. No growth occurred under anaerobic conditions. `H. heilmannii' is oxidase, catalase, nitrite, nitrate, and urease positive. It produces alkaline phosphatase and arginine arylamidase and is sensitive to cephalothin (30 μg disc), resistant to nalidixic acid (30 μg disc)."

Pathogenesis

A wide range of pathologies has been observed with "H. heilmannii" infection. In its natural hosts the inflammatory response is mostly mild, initiating a neutrophilic and mononuclear gastritis (47). There have been several reports of "H. heilmannii" ("Candidatus Helicobacter suis") association with ulcers of the pars esophagus in pigs. Reports indicate that a very high percentage of pigs that have ulcers in this region also have "H. heilmannii" present in the gastric mucosa (4, 73). To date, there has been only one study that has attempted to investigate "H. heilmannii" ulcerogenesis. Krakowka and coworkers' (53) findings indicate that "H. heilmannii" was not involved in ulcers of the pars esophagus. Gnotobiotic piglets that were fed a high-carbohydrate diet and then fed fermentative commensal bacteria (Lactobacillus spp. and Bacillus spp.) developed gastroesophageal ulcers, whereas piglets fed the same diet but inoculated with "H. heilmannii" did not. They suggested that gastroesophageal ulcers develop secondary to epithelial damage mediated by microbial-origin acids whose production is potentiated by high dietary carbohydrate and parietal cell-origin hydrochloric acid.

In humans, the incidence of pathogenic effects induced by this bacterium is low. Less than 1% of all upper gastric complaints are due to "H. heilmannii." Infection with the bacterium induces a gastritis, but the grade of this gastritis is statistically less than that seen with H. pylori infection and the rate of ulceration, erosions, and intestinal metaplasia as a consequence of this bacterium's presence is also low (46, 94). "H. heilmannii" has been implicated in low-grade mucosa-associated lymphoid tissue (MALT) lymphoma of humans (77). Interestingly, the prevalence of MALT lymphoma associated with "H. heilmannii" infection is much higher than with H. pylori. Morgner and associates diagnosed 543 patients with "H. heilmannii" infection between 1988 to 1998, 8 (1.47%) of whom had primary gastric MALT lymphoma. This is compared to 263,680 patients diagnosed with H. pylori (during the same period), of whom 1,745 (0.66%) had primary gastric MALT lymphoma (77). This same group also reported that antimicrobial therapy to cure the stomach of "H. heilmannii" led to complete remission of the MALT lymphoma (76). Most of the severe pathologies that have been reported to be due to "H. heilmannii" infection have occurred in experimental animals; these include neutrophilic and mononuclear gastritis, lymphofollicular gastritis (58), gastric ulcer (24), and MALT lymphoma (80).

Treatment

Data on the antibiotic susceptibility of "H. heilmannii" are limited. Andersen et al. reported that their isolate was sensitive to cephalothin (30 μg disc) and resistant to nalidixic acid (30 μg disc). In vivo, "H. heilmannii" is sensitive to triple therapy in the human; however, in the rodent, triple therapy (metronidazole, bismuth subcitrate, and tetracycline) is unable to clear the bacterium (14). Long-term eradication of "H. heilmannii" from naturally infected cats was also not very effective (79). Using different antimicrobial combinations of azithromycin, tinidazole, ranitidine, clarithromycin, metronidazole, and bismuth, Neiger and coworkers were only able to clear the bacterium initially. After 42 days posttreatment only 56% of animals remained clear.

Helicobacter felis

Morphology

A bacterium morphologically similar to "H. heilmannii" is H. felis. Lee and coworkers first cultured this bacterium in 1988 from the gastric mucosa of a cat (62). H. felis has since been cultured from the antrum of dogs and cats. Morphologically, this gram-negative bacterium is tightly coiled and ranges from 5 to 7.5 μm in length and is 0.4 μm in width (Fig. 2). It possesses bipolar tufts of 10 to 17 sheathed flagella that are positioned slightly off center (62, 84). Each flagellum has a basal body, similar to that of other gram-negative bacteria, that consists of three rings that correspond to the MS, P, and L ring (11). The other striking feature of H. felis (that is shared with a few other helicobacters, e.g., Helicobacter muridarum, Helicobacter bilis, and "Helicobacter rappini") is the possession of periplasmic fibrils that encase the bacterium (Fig. 2). These fibrils are located under the outer membrane, in the periplasm (62). Unlike the flagella, which have clear insertion points and a basal structure, it is unclear if similar structures are associated with the fibrils. Analysis of electron micrographs from freeze-dried, freeze-fractured, negatively stained, and sectioned specimens has yet to illustrate such a structure associated with the fibrils. Ultrastructurally these periplasmic fibrils are quite distinct from the flagella and measure 56 nm in diameter. Degradation with a detergent, Teepol (Shell Petroleum), has revealed that the fibrils have a multilayer structure. An outer sheath covers an outer core that is 21 nm in diameter; inside is an inner core that has a diameter of 7 nm. This inner core is not a single strand but a series of segments and could be involved in some contractile function (Danon, unpublished results). Interestingly, these fibrils are not complete strands that start at one pole of the bacterium and finish at the other. There are several strands that start and finish at different points along the body and appear as concentric helical ridges that can be arranged either singularly, in pairs, in threes, or as a quadruple (11).

Figure 2

Electron micrographs of H. felis. (A) Freeze-fracture preparation; arrow indicates the periplasmic fibrils (photo courtesy of S. Kouprach). (B) Negatively stained preparation; arrow indicates the periplasmic fibrils. (C) H. felis colonizing the gastric (more...)

Growth requirements

H. felis is a fastidious bacterium and ideally requires a complex agar or broth base plus blood or serum for growth. Amphotericin B should be included to prevent the growth of fungi since H. felis prefers conditions that have a high humidity. For optimum growth the organism requires a microaerophilic environment and a temperature of 37°C for 2 days (62). This bacterium grows as a thin film across the surface of solid media and rarely produces discrete colonies. Primary isolation from the stomach using media with Skirrow's supplement (vancomycin [100 μg ml⁻¹], polymyxin B [3.3 μg ml⁻¹], trimethoprim lactate [25 μg ml⁻¹], amphotericin B [50 μg ml⁻¹]) or Glaxo selective supplement (vancomycin [100 μg ml⁻¹], polymyxin B [3.3 μg ml⁻¹], trimethoprim lactate [25 μg ml⁻¹], amphotericin B [50 μg ml⁻¹], bacitracin [200 μg ml⁻¹], and nalidixic acid [10.1 μg ml⁻¹]) is advised to reduce the number of contaminating organisms. Agar plates should be incubated for 4 to 7 days initially. Any spreading growth should be recultured onto fresh media until a pure culture has been obtained. Coccoidal forms will predominate in older cultures. Similar to H. pylori, H. felis is urease, catalase, and oxidase positive and does not readily use carbohydrates (84).

Natural habitat and pathogenesis

H. felis will only colonize defined regions of the stomach, these being the glands of the antrum and the cardia. Unlike "H. heilmannii" and H. pylori, H. felis is very sensitive to the number of parietal cells in the gastric glands and, thus, acid. It cannot survive in gastric glands that have a full complement of parietal cells. This has been shown in germ-free laboratory animals in which the bacterium was only found in antrum and cardia (36, 60), and in a separate study in which H. felis was only observed in the body of the stomach after acid-suppressant therapy (15).

Most of the pathogenic effects due to the presence of H. felis have been observed in laboratory animals (88); however, in its natural host (cats and dogs) H. felis can induce a mild gastritis with lymphofollicular hyperplasia (91). Very few neutrophils are seen, which is in contrast to one of the few human cases that reported a large influx of neutrophils (101).

Transmission

As mentioned earlier, H. felis has been isolated from the gastric mucosa of cats and dogs, and there have been a few cases of this bacterium colonizing the human stomach. The incidence of human infection is low and thought to be a zoonosis, with patients becoming infected due to close contact with animals, most likely cats and dogs. The route of transmission has been suggested as oral-oral (59). Evidence to support this theory arises from experimental data in laboratory animals. Germ-free mice that were infected with H. felis by oral gavage did not pass on the infection to cohoused H. felis-negative animals, nor did they pass the bacterium to H. felis-negative animals that were placed in the same cage (59). This is in contrast to studies conducted in germ-free beagles (59). Experimentally infected germ-free beagle puppies did pass on H. felis to their littermates. The puppies probably passed on H. felis during playtime when they licked and bit each other. The difference between the puppies and mice is that mice cannot regurgitate nor do they have a gag reflex. The evidence against fecaloral transmission is that even though mice can coprophage, the bacterium was not transmitted even when infected germ-free animals were housed with uninfected littermates (59).

Treatment

H. felis is sensitive to a variety of antimicrobial agents. These include metronidazole, cephalothin, erythromycin, ampicillin, and the antimicrobial agents used in triple therapy (bismuth, metronidazole, and tetracycline) (84). Clearance of H. felis from mice (particularly C57BL/6 and BALB/c) with the triple therapy antimicrobial cocktail led to the development of an animal model that could be used for new antimicrobial agents and/or a combination of antimicrobial agents against H. pylori (20).

Helicobacter mustelae

Morphology

A few years after the discovery of H. pylori by Marshall and Warren in 1983 (70, 99), Fox and coworkers isolated an organism from a duodenal ulcer of a ferret (Mustela putorius furo) (34). Even though this gram-negative bacterium is genetically similar to other helicobacters, it was morphologically different. Instead of being spiral in shape, it is a small rod ranging from 2 to 5 μm in length and 0.5 to 1 μm wide (Fig. 3). H. mustelae is highly motile by means of bipolar and lateral sheathed flagella, which have a terminal bulb (39).

Figure 3

Electron micrographs of H. mustelae. (A) H. mustelae colonizing the gastric mucosa of the ferret. Arrow indicates organisms adhering to the epithelial cell layer and partial endocytosis. (B) Negatively stained preparation of H. mustelae. (Photos courtesy (more...)

Growth requirements

The growth requirements are similar to that of H. felis. A complex basal medium (e.g., blood agar base) supplemented with serum or blood is required for optimum growth. After 2 days of incubation at 37°C in a microaerophilic humid environment, small 1- to 2-mm translucent colonies or a spreading film is evident. The organism can tolerate 42°C. Older cultures, like other Helicobacter spp., form coccoidal forms. H. mustelae is urease and catalase positive (39).

Natural habitat

Similar to H. pylori, H. mustelae has a very specific host range. It is a natural colonizer of ferrets (Mustela putorius furo) in many countries, including the United States, Australia, the United Kingdom, and Canada. The only country where this organism has not been found in the ferret is New Zealand (35, 95). It should be noted that ferrets are not native to New Zealand.

Like the other gastric helicobacters, H. mustelae will only colonize the stomach. Unlike H. felis, which is sensitive to the number of parietal cells per gland, this organism can colonize the entire length of the stomach from the proximal duodenum, antrum, through the fundus and cardia. The organism colonizes preferentially the gastric glands of the proximal duodenum and antrum but can readily be seen in the fundus (35, 38, 81). H. mustelae has an unusual motion produced by its polar and lateral flagella. This motion allows the organism to navigate through mucus and attach to epithelial cells. Like H. pylori, H. mustelae adheres firmly to the gastric epithelium; indeed, most are seen attached to epithelial cells. However, unlike H. pylori, many H. mustelae organisms in the ferret stomach are seen to actually enter these epithelial cells via endocytosis (81) (Fig. 3). The bacterium is not degraded but instead is bound inside the epithelial cell by a membrane-bound inclusion (81).

Pathogenesis

The presence of this organism in ferrets results in gastritis consisting of neutrophils and mononuclear cells. Gastric and duodenal ulcers are commonly observed naturally and experimentally. It has been shown that infection occurs after weaning and the severity of infection increases as the animal ages (38). Koch's postulates have been fulfilled; inoculation with H. mustelae into uninfected ferrets results in a similar disease that occurs in naturally infected animals (38). This infection is characterized by a superficial gastritis, which develops in the oxyntic gastric mucosa, and a full-thickness gastritis, composed primarily of lymphocytes and plasma cells plus small numbers of neutrophils and eosinophils, in the antrum (32, 38). Clinical signs of infection include vomiting, melena, chronic weight loss, and low hematocrit; in some acute episodes gastric bleeding can also occur (85). H. mustelae infection of ferrets is the only model of Helicobacter-associated ulcer disease and thus is a very useful model (32). Natural infection with H. mustelae in ferrets can also lead to precancerous lesions, and focal glandular atrophy of the proximal antrum. H. mustelae has also been linked to gastric adenocarcinoma (33) and MALT lymphoma (29).

Treatment

The antimicrobial therapy that is required to clear H. mustelae infection from ferrets is similar to the therapy needed to eradicate H. pylori from humans. The therapy achieves clearance in ferrets in a 3- to 4-week course of amoxicillin (30 mg/kg of body weight), metronidazole (20 mg/kg), and bismuth sub-salicylate (17.5 mg/kg) given three times a day. Also, a combination of ranitidine bismuth citrate and clarithromycin was efficacious in eradicating H. mustelae infection from ferrets in a study undertaken by Marini and coworkers (69).

Helicobacter salomonis

H. salomonis was isolated by Jalava in 1997 (49) from the gastric mucosa of healthy pet dogs and experimental beagles. This gram-negative organism was named in honor of Hugo Salomon. The bacterium is a loose spiral that is 0.8 to 1.2 μm wide and 5 to 7 μm long with polar tufts of 10 to 23 sheathed flagella (Fig. 4B). In vitro it grows as a spreading film across moist agar plates but, like other helicobacters, forms coccoidal forms in old cultures. H. salomonis will not grow in media containing 1% bile, 1% glycine, or 1.5% NaCl or if the incubation temperature is not 37°C. H. salomonis is urease, catalase, and oxidase positive and resistant to nalidixic acid but sensitive to cephalin and cefoperazone (49).

Figure 4

Negatively stained preparations of (A) H. bizzozeronii, magnification × 18,000, and (B) H. salomonis, magnification × 18,000 (photos courtesy of K. Jalava).

Transmission of H. salomonis occurs early in life; puppies can acquire the organism from the dam during the lactation period or via contact with other already infected puppies (42). Antimicrobial therapy with metronidazole, amoxicillin, and bismuth subcitrate will suppress but not totally eradicate the organism from the stomach.

The Ecology of the Gastric Helicobacters

As mentioned at the beginning of this chapter, gastric spiral bacteria are a group of fastidious organisms that inhabit a very harsh environment. So what makes these microorganisms so special that they are able to survive where so many other bacteria fail? All of these bacteria have evolved specialized traits that enable them to survive in the stomach. These include (but are not limited to) a spiral-shaped body, the possession of a highly active urease enzyme, a highly efficient and, in some cases, unique form of motion, and the ability to survive in a low oxygen tension (61).

References

1.

Andersen L. P., Boye K., Blom J., Holck S., Norgaard A., Elsborg L. Characterization of a culturable "Gastrospirillum hominis" (Helicobacter heilmannii) strain isolated from human gastric mucosa. J. Clin. Microbiol. 1999;37:1069–1076. [PMC free article: PMC88651] [PubMed: 10074528]

2.

Andrutis K. A., Fox J. G., Schauer D. B., Marini R. P., Li X. T., Yan L. L., Josenhans C., Suerbaum S. Infection of the ferret stomach by isogenic flagellar mutant strains of Helicobacter mustelae. Infect. Immun. 1997;65:1962–1966. [PMC free article: PMC175254] [PubMed: 9125590]

3.

Balfour A. A haemogregarine of mammals and some notes on trypanosomiasis in the Anglo-Egyptian Sudan. J. Trop. Med. 1906;9:81–92.

4.

Barbosa A. J. A., Silva J. C. P., Nogueira A. M. M. F., Paulino E., Miranda C. R. Higher incidence of Gastrospirillum sp. in swine with gastric ulcer of the pars oesophagea. Vet. Pathol. 1995;32:134–139. [PubMed: 7771053]

5.

Berg H. C. Bacterial behaviour. Nature. 1975;254:389–392. [PubMed: 1090851]

6.

Bizzozero G. Ueber die schlauchfoermigen drusen des magendarmkanals und die beziehungen ihres epithels zu dem oberfachenepithel der schleimhaut. Arch. Mikrobial. Anat. 1892;42:82–152.

7.

Bronsdon M. A., Goodwin C. S., Sly L. I., Chilvers T., Schoenknecht F. D. Helicobacter nemestrinae sp. nov., a spiral bacterium found in the stomach of a pigtailed macaque (Macaca nemestrina). Int. J. Syst. Bacteriol. 1991;41:148–153. [PubMed: 1995031]

8.

Carnot P., Lelievre A. Morphologie du product d'excretion des cellules bordants. Comptes Rendus Soc. Biol. 1909;66:311–313.

9.

Cave D. R., King W. W., Hoffman J. S. Production of two chemically distinct and inhibitory factors produced by H. pylori. Eur. J. Gastroenterol. Hepatol. 1993;5(Suppl.):23–27.

10.

Cave D. R., Vargas M. Effect of a Campylobacter pylori protein on acid secretion by parietal cells. Lancet. 1989;ii:187–189. [PubMed: 2568521]

11.

Danon, S., A. Lee, J. O'Rourke, and S. Kouprach. 1989. The periplasmic fibrils of spiral bacteria that colonise intestinal mucus, p. 416–419. Campylobacter V: Proceedings of the 5th International Workshop on Campylobacter Infections, Puerto Vallarta, Mexico. Department of Infectious Diseases, Instituto Nacional de la Nutricion, Mexico.

12.

Danon S. J., Eaton K. A. The role of gastric Helicobacter and N^-methyl-N′-nitro-N-nitrosoguanidine in carcinogenesis of mice. Helicobacter. 1998;3:260–268. [PubMed: 9844067]

13.

Danon S. J., Fryklund J., Larsson H., Lee A. Acid inhibitory factors of Helicobacter pylori: fact or artefact? Gastroenterology. 1993;104:A688.

14.

Danon S. J., Moss N. D., Larsson H., Arvidsson S., Ottosson S., Dixon M. F., Lee A. Gastrin release and gastric acid secretion in the rat infected with either Helicobacter felis or Helicobacter heilmannii. J. Gastroenterol. Hepatol. 1998;13:95–103. [PubMed: 9737579]

15.

Danon S. J., O'Rourke J. L., Moss N. D., Lee A. The importance of local acid production in the distribution of Helicobacter felis in the mouse stomach. Gastroenterology. 1995;108:1386–1395. [PubMed: 7729630]

16.

De Groote D., Ducatelle R., van Doorn L. J., Tilmant K., Quint W. G. V., Verschuurn A., Haesebrouck F. Detection of "Candidatus Helicobacter suis" in gastric samples of pig by PCR: comparison with other invasive diagnostic techniques. J. Clin. Microbiol. 2000;38:1131–1135. [PMC free article: PMC86356] [PubMed: 10699008]

17.

De Groote D., van Doorn L. J., Ducatelle R., Verschuuren A., Tilmant K., Quint W. G. V., Haesebrouck F., Vandamme P. Phylogenetic characterization of `Candidatus Helicobacter bovis,' a new gastric helicobacter in cattle. Int. J. Syst. Bacteriol. 1999;49:1707–1715. [PubMed: 10555352]

18.

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]

19.

Dick E., Lee A., Watson G., O'Rourke J. Use of the mouse for the isolation and investigation of stomach-associated, spiral-helical shaped bacteria from man and other animals. J. Med. Microbiol. 1989;29:55–62. [PubMed: 2498521]

20.

Dick-Hegedus E., Lee A. Use of a mouse model to examine anti-Helicobacter pylori agents. Scand. J. Gastroenterol. 1991;26:909–915. [PubMed: 1947781]

21.

Doenges J. L. Spirochaetes in the gastric glands of Macacus rhesus and humans without definite history of related disease. Arch. Pathol. 1939;27:469–477.

22.

Eaton K. A., Dewhirst F. E., Radin M. J., Fox J. G., Paster B. J., Krakowka S., Morgan D. R. Helicobacter acinonyx sp. nov., isolated from cheetahs with gastritis. Int. J. Syst. Bacteriol. 1993;43:99–106. [PubMed: 8379970]

23.

Eaton K. A., Radin M. J., Fox J. G., Paster B., Dewhirst F., Krakowka S., Morgan D. R. Helicobacter acinonyx, a new species of Helicobacter isolated from cheetahs with gastritis. Microbial. Ecol. Health Dis. 1991;4(Suppl.):S104. [PubMed: 8379970]

24.

Eaton K. A., Radin M. J., Krakowka S. An animal model of gastric ulcer due to bacterial gastritis in mice. Vet. Pathol. 1995;32:489–497. [PubMed: 8578639]

25.

Eaton K. A., Radin M. J., Kramer L., Wack R., Sherding R., Krakowka S., Fox J. G., Morgan D. R. Epizootic gastritis associated with gastric spiral bacilli in cheetahs (Acinonyx jubatus). Vet. Pathol. 1993;30:55–63. [PubMed: 8442328]

26.

Eaton K. A., Radin M. J., Kramer L., Wack R., Sherding R., Krakowka S., Morgan D. R. Gastric spiral bacilli in captive cheetahs. Scand. J. Gastroenterol. 1991;26:38–42. [PubMed: 1866593]

27.

Eaton K. A., Suerbaum S., Josenhans C., Krakowka S. Colonization of gnotobiotic piglets by Helicobacter pylori deficient in two flagellin genes. Infect. Immun. 1996;64:2445–2448. [PMC free article: PMC174096] [PubMed: 8698465]

28.

Enno A., O'Rourke J. L., Howlett C. R., Jack A., Dixon M. F., Lee A. MALToma-like lesions in the murine gastric mucosa after long-term infection with Helicobacter felis—a mouse model of Helicobacter pylori-induced gastric lymphoma. Am. J. Pathol. 1995;147:217–222. [PMC free article: PMC1869885] [PubMed: 7604881]

29.

Erdman S. E., Correa P., Coleman L. A., Schrenzel M. D., Li X. T., Fox J. G. Helicobacter mustelae-associated gastric MALT lymphoma in ferrets. Am. J. Pathol. 1997;151:273–280. [PMC free article: PMC1857920] [PubMed: 9212752]

30.

Ferrero R. L., Hazell S. L., Lee A. The urease enzymes of Campylobacter pylori and a related bacterium. J. Med. Microbiol. 1988;27:33–40. [PubMed: 3172169]

31.

Fox J. G., Blanco M., Murphy J. C., Taylor N. S., Lee A., Kabok Z., Pappo J. Local and systemic immune responses in murine Helicobacter felis active chronic gastritis. Infect. Immun. 1993;61:2309–2315. [PMC free article: PMC280850] [PubMed: 8500873]

32.

Fox J. G., Correa P., Taylor N. S., Lee A., Otto G., Murphy J. C., Rose R. Helicobacter mustelae-associated gastritis in ferrets. An animal model of Helicobacter pylori gastritis in humans. Gastroenterology. 1990;99:352–361. [PubMed: 2365188]

33.

Fox J. G., Dangler C. A., Sager W., Borkowski R., Gliatto J. M. Helicobacter mustelae-associated gastric adenocarcinoma in ferrets (Mustela putorius furo). Vet. Pathol. 1997;34:225–229. [PubMed: 9163879]

34.

Fox J. G., Edrise B. M., Cabot E. B., Beaucage C., Murphy J. C., Prostak K. S. Campylobacter-like organisms isolated from gastric mucosa of ferrets. Am. J. Vet. Res. 1986;47:236–239. [PubMed: 3954197]

35.

Fox J. G., Lee A. The role of Helicobacter species in newly recognised gastrointestinal tract diseases of animals. Lab. Animal Sci. 1997;47:222–255. [PubMed: 9241625]

36.

Fox J. G., Lee A., Otto G., Taylor N. S., Murphy J. C. Helicobacter felis gastritis in gnotobiotic rats: an animal model of Helicobacter pyroli gastritis. Infect. Immun. 1991;59:785–791. [PMC free article: PMC258328] [PubMed: 1997430]

37.

Fox J. G., Li X. T., Cahill R. J., Andrutis K., Rustgi A. K., Odze R., Wang T. C. Hypertrophic gastropathy in Helicobacter felis-infected wild-type C57BL/6 mice and p53 hemizygous transgenic mice. Gastroenterology. 1996;110:155–166. [PubMed: 8536852]

38.

Fox J. G., Otto G., Taylor N. S., Rosenblad W., Murphy J. C. Helicobacter mustelae-induced gastritis and elevated gastric pH in the ferret (Mustela putorius furo). Infect. Immun. 1991;59:1875–1880. [PMC free article: PMC257936] [PubMed: 2037349]

39.

Fox J. G., Taylor N. S., Edmonds P., Brenner D. J. Campylobacter pylori subsp. mustelae subsp. nov. isolated from the gastric mucosa of ferrets (Mustelae putorius furo), and an emended description of Campylobacter pylori. Int. J. Syst. Bacteriol. 1988;38:367–370.

40.

Goto K., Ohashi H., Ebukuro S., Itoh K., Tohma Y., Takakura A., Wakana S., Ito M., Itoh T. Isolation and characterization of Helicobacter species from the stomach of the house musk shrew (Suncus murinus) with chronic gastritis. Curr. Microbiol. 1998;37:44–51. [PubMed: 9625789]

41.

Gunther H., Schulze F. Histologische untersuchungen zum vorkommen von Camplylobacter-ahnlich geformten keimen im labmagen von kalbern. Zentralbl. Vetmed B. 1992;39:737–745. [PubMed: 1492515]

42.

Hanninen M. L., Happonen I., Jalava K. Transmission of canine gastric Helicobacter salomonis infection from dam to offspring and between puppies. Vet. Microbiol. 1998;62:47–58. [PubMed: 9659691]

43.

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]

44.

Hanninen M. L., Hirvi U. Genetic diversity of canine gastric helicobacters, Helicobacter bizzozeronii and H. salomonis studied by pulsed-field gel electrophoresis. J. Med. Microbiol. 1999;48:341–347. [PubMed: 10509475]

45.

Haringsma P., Mouwen J. Mogelijke betekenis van spirilvormige bacterien bij het ontstaan van lebmaagzweren bij het volwassen rund. Tjdschr. Diergeneeskd. 1992;117:485–487. [PubMed: 1412361]

46.

Heilmann K. L., Borchard F. Gastritis due to spiral shaped bacteria other than Helicobacter pylori: clinical, histological, and ultrastructural findings. Gut. 1991;32:137–140. [PMC free article: PMC1378794] [PubMed: 1864530]

47.

Henry G. A., Long P. H., Burns J. L., Charbonneau D. L. Gastric spirillosis in beagles. Am. J. Vet. Res. 1987;48:831–836. [PubMed: 3592386]

48.

Holck S., Ingeholm P., Blom J., Norgaard A., Elsborg L., Adamsen S., Andersen L. P. The histopathology of human gastric mucosa inhabited by Helicobacter heilmannii-like (Gastrospirillum hominis) organisms, including the first culturable case. APMIS. 1997;105:746–756. [PubMed: 9368589]

49.

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. Bacteriol. 1997;47:975–982. [PubMed: 9336895]

50.

Johnson J. H., Wolf A. M., Jensen J. M., Fossum T., Rohn D., Green R. W., Willard M. Duodenal perforation in a cheetah (Acinonyx jubilatus). J. Zoo Wildlife Med. 1997;28:481–484. [PubMed: 9523644]

51.

Josenhans C., Ferrero R. L., Labigne A., Suerbaum S. Cloning and allelic exchange mutagenesis of two flagellin genes of Helicobacter felis. Mol. Microbiol. 1999;33:350–362. [PubMed: 10411751]

52.

Kasai K., Kobayashi R. The stomach spirochete occurring in mammals. J. Parasitol. 1919;6:1–11.

53.

Krakowka S., Eaton K. A., Rings D. M., Argenzio R. A. Production of gastroesophageal erosions and ulcers (GEU) in gnotobiotic swine monoinfected with fermentative commensal bacteria and fed high-carbohydrate diet. Vet. Pathol. 1998;35:274–282. [PubMed: 9684971]

54.

Kreig N. R. Biology of the echemoheterotrophic spirilla. Bacteriol. Rev. 1976;40:55–115. [PMC free article: PMC413939] [PubMed: 773367]

55.

Kreinitz W. Ueber das auftreten von spirochaeten verschiedener form im magenihalt bei carcinoma ventriculi. Dtsch. Med. Wochenschr. 1906;32:872.

56.

La Perle K. M. D., Wack R., Kaufman L., Blomme E. A. G. Systemic candidiasis in a cheetah (Acinonyx jubatus). J. Zoo Wildlife Med. 1998;29:479–483. [PubMed: 10065861]

57.

Lee A., Chen M., Coltro N. Long term infection of mice with gastric Helicobacters: development of gastric atrophy. Ir. J. Med. Sci. 1992;161:35.

58.

Lee A., Chen M. H., Coltro N., O'Rourke J., Hazell S., Hu P. J., Li Y. Y. Long term infection of the gastric mucosa with Helicobacter species does induce atrophic gastritis in an animal model of Helicobacter pylori infection. Zentralbl. Bakteriol. Int. J. Med. Microbiol. 1993;280:38–50. [PubMed: 8280955]

59.

Lee A., Fox J. G., Otto G., Dick E. H., Krakowka S. Transmission of Helicobacter spp.: a challenge to the dogma of faecal-oral spread. Epidemiol. Infect. 1991;107:99–109. [PMC free article: PMC2272035] [PubMed: 1831765]

60.

Lee A., Fox J. G., Otto G., Murphy J. A small animal model of human Helicobacter pylori active chronic gastritis. Gastroenterology. 1990;99:1315–1323. [PubMed: 2210240]

61.

Lee A., Hazell S. L. Campylobacter pylori in health and disease: an ecological perspective. Microbial. Ecol. Health Dis. 1988;1:1–16.

62.

Lee A., Hazell S. L., O'Rourke J., Kouprach S. Isolation of a spiral-shaped bacterium from the cat stomach. Infect. Immun. 1988;56:2843–2850. [PMC free article: PMC259659] [PubMed: 3169989]

63.

Lee A., Krakowka S., Fox J. G., Otto G., Eaton K. A., Murphy J. C. Role of Helicobacter felis in chronic canine gastritis. Vet. Pathol. 1992;29:487–494. [PubMed: 1448894]

64.

Lee A., O'Rourke J. Gastric bacteria other than Helicobacter pylori. Gastroenterol. Clinics North Am. 1993;22:21–42. [PubMed: 8449568]

65.

Lee, A., and J. L. O'Rourke. 1993. Ultrastructure of Helicobacter organisms and possible revelance for pathogenesis, p. 15–35. In C. S. Goodwin and B. W. Worsley (ed.), Helicobacter pylori Biology and Clinical Practice. CRC Press, Boca Raton, Fla.

66.

Lee, A., J. L. O'Rourke, and J. E. Kellow. 1995. Gastrospirillum hominis ("Helicobacter heilmannii") and other gastric infections in humans, p. 589–601. In M. J. Blaser, P. D. Smith, J. I. Ravdin, H. B. Greenberg, and R. L. Guerrant (ed.), Infections of the Gastrointestinal Tract. Raven Press, New York, N.Y.

67.

Leung V., van Zanten S. V., O'Rourke J. L., Lee A. Competitive exclusion of H. pylori from the rodent antrum by H. felis, insights into gastric colonisation. Gut. 1998;43(Suppl. 2):A115.

68.

Lim R. K. S. A parasitic spiral organism in the stomach of the cat. Parasitology. 1920;12:108–113.

69.

Marini R. P., Fox J. G., Taylor N. S., Yan L. L., McColm A. A., Williamson R. Ranitidine bismuth citrate and clarithromycin, alone or in combination, for eradication of Helicobacter mustelae infection in ferrets. Am. J. Vet. Res. 1999;60:1280–1286. [PubMed: 10791942]

70.

Marshall B. J. Unidentified curved bacillus on gastric epithelium in active chronic gastritis. Lancet. 1983;i:1273–1275. [PubMed: 6134060]

71.

McNulty C. A. M., Dent J. C., Curry A., Uff J. S., Ford G. A., Gear M. W., Wilkinson S. P. New spiral bacterium in gastric mucosa. J. Clin. Pathol. 1989;42:585–591. [PMC free article: PMC1141985] [PubMed: 2738164]

72.

Mendes E. N., Queiroz D. M., Rocha G. A., Moura S. B., Leite V. H., Fonseca M. E. Ultrastructure of a spiral micro-organism from pig gastric mucosa ("Gastrospirillum suis"). J. Med. Microbiol. 1990;33:61–66. [PubMed: 2231673]

73.

Mendes E. N., Queiroz D. M., Rocha G. A., Nogueira A. M., Carvalho A. C., Lage A. P., Barbosa A. J. Histopathological study of porcine gastric mucosa with and without a spiral bacterium ("Gastrospirillum suis"). J. Med. Microbiol. 1991;35:345–348. [PubMed: 1753392]

74.

Mendes E. N., Queiroz D. M. M., Coimbra R. S., Moura S. B., Barbosa A. J. A., Rocha G. A. Experimental infection of Wistar rats with Gastrospirillum suis. J. Med. Microbiol. 1996;44:105–109. [PubMed: 8642570]

75.

Mendz G. L., Hazell S. L. The urea cycle of Helicobacter pylori. Microbiology. 1996;142:2959–2967. [PubMed: 8885413]

76.

Morgner A., Bayerdorffer E., Neubauer A., Stolte M. Gastric MALT lymphoma and its relationship to Helicobacter pylori infection: management and pathogensis of the disease. Microsc. Res. Tech. 2000;48:349–356. [PubMed: 10738316]

77.

Morgner A., Lehn N., Andersen L. P., Thiede C., Bennedsen M., Trebesius K., Neubauer B., Neubauer A., Stolte M., Bayerdorffer E. Helicobacter heilmannii-associated primary gastric low-grade MALT lymphoma: complete remission after curing the infection. Gastroenterology. 2000;118:821–828. [PubMed: 10784580]

78.

Munson L., Nesbit J. W., Meltzer D. G. A., Colly L. P., Bolton L., Kriek N. P. J. Diseases of captive cheetahs (Acinonyx jubatus jubatus) in South Africa: a 20-year retrospective survey. J. Zoo Wildlife Med. 1999;30:342–347. [PubMed: 10572855]

79.

Neiger R., Seiler G., Schmassmann A. Use of a urea breath test to evaluate short-term treatments for cats naturally infected with Helicobacter heilmannii. Am. J. Vet. Res. 1999;60:880–883. [PubMed: 10407483]

80.

O'Rourke J. L., Enno A., Lee A., Dixon M. Gastric B-cell lymphomas induced in a single mouse strain by various isolates of Helicobacter heilmannii: similarities and differences. Gut. 1995;37(Suppl. 1):A7.

81.

O'Rourke J. L., Lee A., Fox J. G. An ultrastructural study of Helicobacter mustelae and evidence of a specific association with gastric mucosa. J. Med. Microbiol. 1992;36:420–427. [PubMed: 1613782]

82.

O'Rourke, J. L., B. A. Neilan, and A. Lee. 1999. Phylogenic relationship of Helicobacter heilmannii-like organisms originating from humans and animals. 10th International Workshop on Campylobacter, Helicobacter and Related Organisms, Baltimore, Md.

83.

O'Rourke J. L., Solnick J., Lee A., Tompkins L. S. Helicobacter heilmannii" (previously Gastrospirillum), a new species of helicobacter in humans and animals. Ir. J. Med. Sci. 1992;161(Suppl. 10):31.

84.

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]

85.

Perkins S. E., Fox J. G., Walsh J. H. Helicobacter mustelae-associated hypergastrinemia in ferrets (Mustela putorius furo). Am. J. Vet. Res. 1996;57:147–150. [PubMed: 8633798]

86.

Rappin, J. P. 1881. Contra l'etude de bacterium de la bouche a l'etat normal, quoted by, p. 68. In R. S. Breed, E. G. D. Murray, and A. P. Hitchens (ed.), Bergey's Manual of Determinative Bacteriology, 6th ed. Williams & Wilkins, Baltimore, Md.

87.

Regard C. Sur une curieuse lacalisation de spirilles parasites dans les canalisations glandulaires de la gastrique normale, chez le chien et le chat. Soc. Biol. 1909;66:229–231.

88.

Sakagami T., Dixon M., O'Rourke J., Howlett R., Alderuccio F., Vella J., Shimoyama T., Lee A. Atrophic gastric changes in both Helicobacter felis and Helicobacter pylori infected mice are host dependent and separate from antral gastritis. Gut. 1996;39:639–648. [PMC free article: PMC1383385] [PubMed: 9026476]

89.

Salomon H. Uber das spirillum des saugetiermagens und sein verhalten zu den belegzellen. Zentralbl. Bakteriol. 1896;19:433–441.

90.

Schreiber S., Stuben M., Josenhans C., Scheid P., Suerbaum S. In vivo distribution of Helicobacter felis in the gastric mucus of the mouse: experimental method and results. Infect. Immun. 1999;67:5151–5156. [PMC free article: PMC96864] [PubMed: 10496889]

91.

Simpson K. W., Strauss-Ayali D., Scanziani E., Straubinger R. K., McDonough P. L., Straubinger A. F., Chang Y. F., Domeneghini C., Arebi N., Calam J. Helicobacter felis infection is associated with lymphoid follicular hyperplasia and mild gastritis but normal gastric secretory function in cats. Infect. Immun. 2000;68:779–790. [PMC free article: PMC97205] [PubMed: 10639446]

92.

Skouloubris S., Thiberge J. M., Labigne A., Dereuse H. The Helicobacter pylori ureI protein is not involved in urease activity but is essential for bacterial survival in vivo. Infect. Immun. 1998;66:4517–4521. [PMC free article: PMC108549] [PubMed: 9712811]

93.

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]

94.

Stolte M., Kroher G., Meining A., Morgner A., Bayerdorffer E., Bethke B. Comparison of Helicobacter pylori and H. heilmannii gastritis—matched control study involving 404 patients. Scand. J. Gastroenterol. 1997;32:28–33. [PubMed: 9018763]

95.

Tompkins D. S., Wyatt J. I., Rathbone B. J., West A. P. The characterization and pathological significance of gastric Campylobacter-like organisms in the ferret: a model for chronic gastritis? Epidemiol. Infect. 1988;101:269–278. [PMC free article: PMC2249373] [PubMed: 3181311]

96.

Vargas M., Lee A., Fox J. G., Cave D. R. Inhibition of acid secretion from parietal cells by non-human-infecting Helicobacter species: a factor in colonization of gastric mucosa? Infect. Immun. 1991;59:3694–3699. [PMC free article: PMC258940] [PubMed: 1894369]

97.

Wack R. F., Eaton K. A., Kramer L. W. Treatment of gastritis in cheetahs (Acinonyx jubatus). J. Zoo Wildlife Med. 1997;28:260–266. [PubMed: 9365937]

98.

Wang T. C., Dangler C. A., Chen D., Goldenring J. R., Koh T., Raychowdhury R., Coffey R. J., Ito S., Varro A., Dockray G. J., Fox J. G. Synergistic interaction between hypergastrinemia and Helicobacter infection in a mouse model of gastric cancer. Gastroenterology. 2000;118:36–47. [PubMed: 10611152]

99.

Warren J. R. Unidentified curved bacilli on gastric epithelium in active chronic gastritis. Lancet. 1983;i:1273. [PubMed: 6134060]

100.

Weeks D. L., Eskandari S., Scott D. R., Sachs G. A H + -gated urea channel: the link between Helicobacter pylori urease and gastric colonization. Science. 2000;287:482–485. [PubMed: 10642549]

101.

Wegmann W., Aschwanden M., Schaub N., Aenishänslin W., Gyr K. Gastrospirillum-hominis-assoziierte gastritis—eine zoonose? Schweiz Med. Wochenschr. 1991;121:245–254. [PubMed: 2011719]