Comparative ultrastructural and functional studies of Helicobacter pylori and Helicobacter mustelae flagellin mutants: both flagellin subunits, FlaA and FlaB, are necessary for full motility in Helicobacter species.

Helicobacter mustelae causes chronic gastritis and ulcer disease in ferrets. It is therefore considered an important animal model of human Helicobacter pylori infection. High motility even in a viscous environment is one of the common virulence determinants of Helicobacter species. Their sheathed flagella contain a complex filament that is composed of two distinctly different flagellin subunits, FlaA and FlaB, that are coexpressed in different amounts. Here, we report the cloning and sequence determination of the flaA gene of H. mustelae NCTC12032 from a PCR amplification product. The FlaA protein has a calculated molecular mass of 53 kDa and is 73% homologous to the H. pylori FlaA subunit. Isogenic flaA and flaB mutants of H. mustelae F1 were constructed by means of reverse genetics. A method was established to generate double mutants (flaA flaB) of H. mustelae F1 as well as H. pylori N6. Genotypes, motility properties, and morphologies of the H. mustelae flagellin mutants were determined and compared with those of the H. pylori flaA and flaB mutants described previously. The flagellar organizations of the two Helicobacter species proved to be highly similar. When the flaB genes were disrupted, motility decreased by 30 to 40%. flaA mutants retained weak motility by comparison with strains that were devoid of both flagellin subunits. Weakly positive motility tests of the flaA mutants correlated with the existence of short truncated flagella. In H. mustelae, lateral as well as polar flagella were present in the truncated form. flaA flaB double mutants were completely nonmotile and lacked any form of flagella. These results show that the presence of both flagellin subunits is necessary for complete motility of Helicobacter species. The importance of this flagellar organization for the ability of the bacteria to colonize the gastric mucosa and to persist in the gastric mucus remains to be proven.     

comH, a novel gene essential for natural transformation of Helicobacter pylori

Helicobacter pylori is naturally competent for transformation, but the DNA uptake system of this bacterium is only partially characterized, and nothing is known about the regulation of competence in H. pylori. To identify other components involved in transformation or competence regulation in this species, we screened a mutant library for competence-deficient mutants. This resulted in the identification of a novel, Helicobacter-specific competence gene (comH) whose function is essential for transformation of H. pylori with chromosomal DNA fragments as well as with plasmids. Complementation of comH mutants in trans completely restored competence. Unlike other transformation genes of H. pylori, comH does not belong to a known family of orthologous genes. Moreover, no significant homologs of comH were identified in currently available databases of bacterial genome sequences. The comH gene codes for a protein with an N-terminal leader sequence and is present in both highly competent and less-efficient transforming H. pylori strains. A comH homolog was found in Helicobacter acinonychis but not in Helicobacter felis and Helicobacter mustelae.     

Characterization and Therapy for Experimental Infection by Helicobacter mustelae in Ferrets

Background. Numerous clinical trials evaluating the efficacy of various antimicrobial compounds against Helicobacter pylori infection have been performed in humans. A convenient animal model for Helicobacter infection would facilitate the evaluation of novel therapies. These experiments were performed to evaluate the use of ferrets as a model of Helicobacter infection.
Materials and Methods. Ferrets were infected experimentally with Helicobacter mustelae and subsequently treated with bismuth subsalicylate (BSS) triple therapy (BSS, metronidazole, and amoxicillin), or left untreated. The status of infection and serology was assessed during treatment and for 8 weeks posttreatment. Seven ferrets successfully treated with triple therapy were challenged with H. mustelae and monitored for infection for an additional 5 weeks.
Results. Infection of ferrets by H. mustelae was accompanied by gastritis and a specific antibody response. Treatment of H. mustelae -infected ferrets with BSS suppressed bacterial growth in four of nine animals but did not eradicate infection. Triple therapy eradicated infection in all nine ferrets with a reduction in gastric inflammation. No relapse of infection occurred up to 8 weeks posttherapy. Challenge with H. mustelae of ferrets successfully treated with triple therapy resulted in a 100% rate of reinfection.
Conclusions. H. mustelae infection can be eliminated by triple therapy, but this does not result in protective immunity against reinfection by H. mustelae. This model, using a strain of Helicobacter indigenous to the host, may be useful for assessing therapeutic efficacy of novel therapies for the treatment of human infection by H. pylori.     

Construction of isogenic urease-negative mutants of Helicobacter pylori by allelic exchange.

Isogenic urease-negative mutants of Helicobacter pylori were constructed by allelic replacement. A region of cloned H. pylori DNA containing the structural urease genes (ureA and ureB) was disrupted by insertion of a mini-Tn3-Km transposon. Electrotransformation of H. pylori cells with kanamycin-ureB-disrupted derivative plasmids resulted in isolation of kanamycin-resistant H. pylori transformants. Competence for electrotransformation appeared to be restricted to certain wild-type H. pylori isolates; only 1 isolate (of 10 tested) was consistently transformed. Two of the kanamycin-resistant H. pylori transformants were further studied and shown to be urease negative. Southern hybridization analyses demonstrated that the urease-negative mutants had been constructed by allelic exchange involving simultaneous replacement of the ureB gene with the kanamycin-ureB-disrupted copy and loss of the vector. Immunoblot studies of whole-cell extracts of the isogenic ureB mutants with anti-H. pylori sera indicated the absence of a polypeptide with an apparent molecular mass of 61 kDa; thus, the mutants no longer synthesized the UreB product. Generation of stable, genetically engineered urease mutants of H. pylori will be useful for addressing the role of urease in the pathogenesis of H. pylori infection.     

Global transposon mutagenesis and essential gene analysis of Helicobacter pylori

We have constructed a genome-saturating mutant library of the human gastric pathogen Helicobacter pylori. Microarray tracking of transposon mutants (MATT) allowed us to map the position of 5,363 transposon mutants in our library. While we generally found insertions well distributed throughout the genome, 344 genes had no detectable transposon insertions, and this list is predicted to be highly enriched for essential genes. Comparison to the essential gene set of other bacteria revealed a surprisingly limited overlap with all organisms tested (11%), while 55% were essential in some organisms but not others. We independently verified the essentiality of several gene products, including an HtrA family serine protease, a hypothetical protein with putative phospholipase D activity, and a riboflavin specific deaminase. A limited screen for motility mutants allowed us to estimate that 4.5% of the genome is dedicated to this virulence-associated phenotype.     

Loss of urease activity in Helicobacter mustelae

Urease-negative variants of Helicobacter mustelae were isolated after spontaneous loss of activity during sub-culture. The whole-cell protein patterns showed that the loss of urease activity was linked to the absence of two polypeptides of 29·1 and 65·4 kDa. Restriction endonuclease analysis of chromosomal DNA indicated no substantial differences between the urease negative and positive variants. It is likely that a change in the expression of the gene for urease was responsible for these observations.     

Cloning and allelic exchange mutagenesis of two flagellin genes of Helicobacter felis

Helicobacter felis has been used extensively in animal model studies of gastric Helicobacter infections. Attempts to manipulate H. felis genetically have, however, been unsuccessful and, consequently, little is known about the pathogenic mechanisms of this bacterium. In common with other Helicobacter spp., H. felis is a highly motile organism. To characterize the flagellar structures responsible for this motility, we cloned and sequenced the two flagellin-encoding genes, flaA and flaB, from H. felis. These genes encode two flagellin proteins that are expressed simultaneously under the control of putative sigma28 and sigma54 promoters respectively. Isogenic mutants of H. felis in flaA and flaB were generated by electroporation-mediated allelic disruption and replacement, showing for the first time that H. felis could be manipulated genetically. Both types of H. felis flagellin mutants exhibited truncated flagella and were poorly motile. H. felis flaA mutants were unable to colonize the gastric mucosa in a mouse infection model.     

Motility of urease-deficient derivatives of Helicobacter pylori

Early studies of a ureB mutant derivative of Helicobacter pylori had suggested that urease is needed for motility and that urease action helps energize flagellar rotation. Here we report experiments showing that motility is unaffected by deletion of ureA and ureB (urease genes) or by inactivation of ureB alone, especially if H. pylori strains used as recipients for transformation with mutant alleles are preselected for motility. This result was obtained with the strain used in the early studies (CPY3401) and also with 15 other strains, 3 of which can colonize mice. We conclude that urease is not needed for H. pylori motility.     

Helicobacter pylori ABC transporter: effect of allelic exchange mutagenesis on urease activity.

Helicobacter pylori urease requires nickel ions in the enzyme active site for catalytic activity. Nickel ions must, therefore, be actively acquired by the bacterium. NixA (high-affinity nickel transport protein)-deficient mutants of H. pylori retain significant urease activity, suggesting the presence of alternate nickel transporters. Analysis of the nucleotide sequence of the H. pylori genome revealed a homolog of NikD, a component of an ATP-dependent nickel transport system in Escherichia coli. Based on this sequence, a 378-bp DNA fragment was PCR amplified from H. pylori genomic DNA and used as a probe to identify an H. pylori lambda ZAPII genomic library clone that carried these sequences. Four open reading frames of 621, 273, 984, and 642 bp (abcABCD) were revealed by sequencing and predicted polypeptides of 22.7, 9.9, 36.6, and 22.8 kDa, respectively. The 36.6-kDa polypeptide (AbcC) has significant homology (56% amino acid sequence identity) to an E. coli ATP-binding protein component of an ABC transport system, while none of the other putative proteins are significantly homologous to polypeptides in the available databases. To determine the possible contribution of these genes to urease activity, abcC and abcD were each insertionally inactivated with a kanamycin resistance (aphA) cassette and allelic exchange mutants of each gene were constructed in H. pylori UMAB41. Mutation of abcD resulted in an 88% decrease in urease activity to 27 +/- 31 mumol of NH3/min/mg of protein (P < 0.0001), and a double mutant of nixA and abcC resulted in the near abolishment of urease activity (1.1 +/- 1.4 mumol of NH3/min/mg of protein in the double mutant versus 228 +/- 92 mumol of NH3/min/mg of protein in the parent [P < 0.0001]). Synthesis of urease apoenzyme, however, was unaffected by mutations in any of the abc genes. We conclude that the abc gene cluster, in addition to nixA, is involved in production of a catalytically active urease.     

Differentiation between isolates of Helicobacter pylori by PCR-RFLP analysis of urease A and B genes and comparison with ribosomal RNA gene patterns

Abstract Genetic diversity amongst 21 human gastric isolates of Helicobacter pylori was investigated by polymerase chain reaction amplification and Hae III digest (restriction fragment length polymorphism) analysis of an internal 2.4-kb segment of the urease A and urease B genes. H. pylori from 11 independent individuals yielded nine distinct restriction fragment patterns but only one pattern was common to H. pylori from two individuals. By contrast, multiple isolate sets of H. pylori from two patients each had common urease gene patterns. Most strains with the same urease gene patterns were distinguishable in their ribosomal RNA gene patterns. The study demonstrated diversity amongst H. pylori and established that PCR analysis of urease genes provided a novel method of identifying isolates. The profiles were reproducible and convenient to obtain and analyse, and were almost as discriminatory as Hae III ribopatterns.     

Urease-mediated destruction of bacteria is specific for Helicobacter urease and results in total cellular disruption

Abstract The survival of Helicobacter mustelae, Proteus mirabilis, Escherichia coli and Campylobacter jejuni in the presence of urea and citrate at pH 6.0 was examined. H. mustelae , which has urease activity similar to H. pylori , had a markedly reduced survival, median 2.5% (0–78%) ( P <0.001) when incubated nder these conditions. Only 7% of the ammonia produced by H. mutelae urease activity was recovered from the buffer, a similar percentage to that previously reported with H. pylori . None of the other organisms, all of which had lower urease activity, had impaired survival under these conditions. Electron microscopical studies demonstrated extensive structural damage to H. pylori following exposure to urea and citrate at pH 6.0. This structural damage to the organisms makes it unlikely that the low recovery of ammonia was due to retention of ammonia within the bacteria and suggests that the ammonia may have been incorporated into glutamate or other amino acids. Incorporation of ammonia into these compounds would deplete the cell of the key metabolic intermediate α-ketoglutarate and could thus explain the mechanism of the urease-dependent destruction of the organism.     

Comparative chemical and biological characterization of the lipopolysaccharides of gastric and enterohepatic helicobacters

BACKGROUND: The lipopolysaccharide of Helicobacter pylori plays an important role in colonization and pathogenicity. The present study sought to compare structural and biological features of lipopolysaccharides from gastric and enterohepatic Helicobacter spp. not previously characterized. MATERIALS AND METHODS: Purified lipopolysaccharides from four gastric Helicobacter spp. (H. pylori, Helicobacter felis, Helicobacter bizzozeronii and Helicobacter mustelae) and four enterohepatic Helicobacter spp. (Helicobacter hepaticus, Helicobacter bilis, 'Helicobacter sp. flexispira' and Helicobacter pullorum) were structurally characterized using electrophoretic, serological and chemical methods. RESULTS: Structural insights into all three moieties of the lipopolysaccharides, i.e. lipid A, core and O-polysaccharide chains, were gained. All species expressed lipopolysaccharides bearing an O-polysaccharide chain, but H. mustelae and H. hepaticus produced truncated semirough lipopolysaccharides. However, in contrast to lipopolysaccharides of H. pylori and H. mustelae, no blood group mimicry was detected in the other Helicobacter spp. examined. Intra-species, but not interspecies, fatty acid profiles of lipopolysaccharides were identical within the genus. Although shared lipopolysaccharide-core epitopes with H. pylori occurred, differing structural characteristics were noted in this lipopolysaccharide region of some Helicobacter spp. The lipopolysaccharides of the gastric helicobacters, H. bizzozeronii and H. mustelae, had relative Limulus amoebocyte lysate activities which clustered around that of H. pylori lipopolysaccharide, whereas H. bilis, 'Helicobacter sp. flexispira' and H. hepaticus formed a cluster with approximately 1000-10,000-fold lower activities. H. pullorum lipopolysaccharide had the highest relative Limulus amoebocyte lysate activity of all the helicobacter lipopolysaccharides (10-fold higher than that of H. pylori lipopolysaccharide), and all the lipopolysaccharides of enterohepatic Helicobacter spp. were capable of inducing nuclear factor-Kappa B(NF-kappaB) activation. CONCLUSIONS: The collective results demonstrate the structural heterogeneity and pathogenic potential of lipopolysaccharides of the Helicobacter genus as a group and these differences in lipopolysaccharides may be indicative of adaptation of the bacteria to different ecological niches.     

Isolation of Helicobacter pylori genes that modulate urease activity

Helicobacter pylori urease, a nickel-requiring metalloenzyme, hydrolyzes urea to NH3 and CO2. We sought to identify H. pylori genes that modulate urease activity by constructing pHP8080, a plasmid which encodes both H. pylori urease and the NixA nickel transporter. Escherichia coli SE5000 and DH5alpha transformed with pHP8080 resulted in a high-level urease producer and a low-level urease producer, respectively. An H. pylori DNA library was cotransformed into SE5000 (pHP8080) and DH5alpha (pHP8080) and was screened for cotransformants expressing either lowered or heightened urease activity, respectively. Among the clones carrying urease-enhancing factors, 21 of 23 contained hp0548, a gene that potentially encodes a DNA helicase found within the cag pathogenicity island, and hp0511, a gene that potentially encodes a lipoprotein. Each of these genes, when subcloned, conferred a urease-enhancing activity in E. coli (pHP8080) compared with the vector control. Among clones carrying urease-decreasing factors, 11 of 13 clones contained the flbA (also known as flhA) flagellar biosynthesis/regulatory gene (hp1041), an lcrD homolog. The LcrD protein family is involved in type III secretion and flagellar secretion in pathogenic bacteria. Almost no urease activity was detected in E. coli (pHP8080) containing the subcloned flbA gene. Furthermore, there was significantly reduced synthesis of the urease structural subunits in E. coli (pHP8080) containing the flbA gene, as determined by Western blot analysis with UreA and UreB antiserum. Thus, flagellar biosynthesis and urease activity may be linked in H. pylori. These results suggest that H. pylori genes may modulate urease activity.     

Essential role of Helicobacter pylori gamma-glutamyltranspeptidase for the colonization of the gastric mucosa of mice

Constitutive expression of gamma-glutamyltranspeptidase (GGT) activity is common to all Helicobacter pylori strains, and is used as a marker for identifying H. pylori isolates. Helicobacter pylori GGT was purified from sonicated extracts of H. pylori strain 85P by anion exchange chromatography. The N-terminal amino acid sequences of two of the generated endo-proteolysed peptides were determined, allowing the cloning and sequencing of the corresponding gene from a genomic H. pylori library. The H. pylori ggt gene consists of a 1681 basepair (bp) open reading frame encoding a protein with a signal sequence and a calculated molecular mass of 61 kDa. Escherichia coli clones harbouring the H. pylori ggt gene exhibited GGT activity at 37 degrees C, in contrast to E. coli host cells (MC1061, HB101), which were GGT negative at 37 degrees C. GGT activity was found to be constitutively expressed by similar genes in Helicobacter felis, Helicobacter canis, Helicobacter bilis, Helicobacter hepaticus and Helicobacter mustelae. Western immunoblots using rabbit antibodies raised against a His-tagged-GGT recombinant protein demonstrated that H. pylori GGT is synthesized in both H. pylori and E. coli as a pro-GGT that is processed into a large and a small subunit. Deletion of a 700 bp fragment within the GGT-encoding gene of a mouse-adapted H. pylori strain (SS1) resulted in mutants that were GGT negative yet grew normally in vitro. These mutants, however, were unable to colonize the gastric mucosa of mice when orally administered alone or together (co-infection) with the parental strain. These results demonstrate that H. pylori GGT activity has an essential role for the establishment of the infection in the mouse model, demonstrating for the first time a physiological role for a bacterial GGT enzyme.     

Conservation and Diversity of the Helicobacter pylori Copper-Transporting ATPase Gene (copA) Sequence Among Helicobacter Species and Campylobacter Species Detected by PCR and RFLP

Background Helicobacter pylori is a causative pathogen of such human stomach diseases as chronic type B gastritis, ulcer, and possibly gastric carcinoma. As a co-factor in various redox enzymes and an essential trace metal required for the synthesis of metalloproteins, copper might play a role in the pathogenesis of H. pylori. A gene, copA , associated with copper transport, has been isolated from H. pylori UA802. In this study, conservation and diversity of this gene were analyzed among some Helicobacter and Campylobacter species.
Materials and Methods. Twenty-one clinical isolates and strains of helicobacters and campylobacters were used in this study. Methods including polymerase chain reaction (PCR) amplification, restriction fragment-length polymorphisms (RFLPs), and hybridization were employed to carry out this work.
Results. The copA gene was highly conserved in all the H. pylori isolates tested ( Helicobacter nemestrinae and Helicobacter felis but not in Helicobacter mustelae and the Campylobacter species), whereas the sequence downstream of the copA appears to diverge among H. pylori isolates. In addition, two restriction patterns of the PCR-amplified copA fragments from seven H. pylori isolates and H. nemestrinae were identified, and the RFLP of H. nemestrinae was identical to that of one of the H. pylori isolate group.
Conclusions. The adenosine triposphatase-derived copper-transporting mechanism is employed by various H. pylori strains, H. nemestrinae, H. felis , and perhaps by other Helicobacter species. The nucleotide mutations have risen in the copA gene. It appears that there is a genetic relatedness of the copA gene to H. pylori and H. nemestrinae.     

Dependence of Helicobacter pylori urease activity on the nickel-sequestering ability of the UreE accessory protein

The Helicobacter pylori ureE gene product was previously shown to be required for urease expression, but its characteristics and role have not been determined. The UreE protein has now been overexpressed in Escherichia coli, purified, and characterized, and three altered versions were expressed to address a nickel-sequestering role of UreE. Purified UreE formed a dimer in solution and was capable of binding one nickel ion per dimer. Introduction of an extra copy of ureE into the chromosome of mutants carrying mutations in the Ni maturation proteins HypA and HypB resulted in partial restoration of urease activity (up to 24% of the wild-type levels). Fusion proteins of UreE with increased ability to bind nickel were constructed by adding histidine-rich sequences (His-6 or His-10 to the C terminus and His-10 as a sandwich fusion) to the UreE protein. Each fusion protein was overexpressed in E. coli and purified, and its nickel-binding capacity and affinity were determined. Each construct was also expressed in wild-type H. pylori and in hypA and hypB mutant strains for determining in vivo urease activities. The urease activity was increased by introduction of all the engineered versions, with the greatest Ni-sequestering version (the His-6 version) also conferring the greatest urease activity on both the hypA and hypB mutants. The differences in urease activities were not due to differences in the amounts of urease peptides. Addition of His-6 to another expressed protein (triose phosphate isomerase) did not result in stimulation of urease, so urease activation is not related to the level of nonspecific protein-bound nickel. The results indicate a correlation between H. pylori urease activity and the nickel-sequestering ability of the UreE accessory protein.     

Steryl glycosides: a characteristic feature of the Helicobacter spp.?

The lipids of different species of Helicobacter (H.felis, H. muridarum, H. mustelae, H. fennelliae, and H. cinaedi) were studied. Different types of cholesteryl glucosides were found in all of the species studied except H. cinaedi. The total amount of cholesteryl glucosides varied from 14.8% of total lipids in H. mustelae to 33.1% of total lipids in H. felis. The different types of cholesteryl glucosides and their species distribution are cholesteryl-6-O-acyl-alpha-D-glucopyranoside (cholesteryl-6-O-tetradecanoyl-alpha-D-glucopyranoside in H. felis and cholesteryl-6-O-dodecanoyl-alpha-D-glucopyranoside in H. muridarum), cholesteryl-alpha-D-glucopyranoside (H. felis, H. muridarum, H. mustelae, and H. fennelliae), and cholesteryl-6-O-phosphatidyl-alpha-D-glucopyranoside (H. fennelliae). The neutral lipid fractions showed a high percentage of cholesterol, with selective accumulation of free cholesterol. The study thus shows that the characteristic presence of steryl glycosides in Helicobacter spp. may be an important chemotaxonomic marker for many of the species, and the helicobacters show a selective accumulation of free cholesterol from the media.     

Helicobacter pylori rocF is required for arginase activity and acid protection in vitro but is not essential for colonization of mice or for urease activity

Arginase of the Helicobacter pylori urea cycle hydrolyzes L-arginine to L-ornithine and urea. H. pylori urease hydrolyzes urea to carbon dioxide and ammonium, which neutralizes acid. Both enzymes are involved in H. pylori nitrogen metabolism. The roles of arginase in the physiology of H. pylori were investigated in vitro and in vivo, since arginase in H. pylori is metabolically upstream of urease and urease is known to be required for colonization of animal models by the bacterium. The H. pylori gene hp1399, which is orthologous to the Bacillus subtilis rocF gene encoding arginase, was cloned, and isogenic allelic exchange mutants of three H. pylori strains were made by using two different constructs: 236-2 and rocF::aphA3. In contrast to wild-type (WT) strains, all rocF mutants were devoid of arginase activity and had diminished serine dehydratase activity, an enzyme activity which generates ammonium. Compared with WT strain 26695 of H. pylori, the rocF::aphA3 mutant was approximately 1, 000-fold more sensitive to acid exposure. The acid sensitivity of the rocF::aphA3 mutant was not reversed by the addition of L-arginine, in contrast to the WT, and yielded a approximately 10, 000-fold difference in viability. Urease activity was similar in both strains and both survived acid exposure equally well when exogenous urea was added, indicating that rocF is not required for urease activity in vitro. Finally, H. pylori mouse-adapted strain SS1 and the 236-2 rocF isogenic mutant colonized mice equally well: 8 of 9 versus 9 of 11 mice, respectively. However, the rocF::aphA3 mutant of strain SS1 had moderately reduced colonization (4 of 10 mice). The geometric mean levels of H. pylori recovered from these mice (in log(10) CFU) were 6.1, 5.5, and 4.1, respectively. Thus, H. pylori rocF is required for arginase activity and is crucial for acid protection in vitro but is not essential for in vivo colonization of mice or for urease activity.     

Natural and Experimental Helicobacter mustelae Reinfection Following Successful Antimicrobial Eradication in Ferrets

Background. Recrudescence or reinfection may occur after eradication of Helicobacter pylori in humans.
Materials and Methods. We used the ferret Helicobacter mustelae model to investigate the effect of prior infection and eradication on reinfection by experimental and natural routes. Two groups of ferrets with naturally acquired H. mustelae infection were treated with an eradication protocol using amoxicillin, metronidazole, and bismuth subsalicylate. The ferrets were monitored for recrudescence by repeated cultures of endoscopic gastric mucosal biopsies. The ferrets were challenged at 17 months (group I) and 6 months (group II) after eradication with a strain of H. mustelae having a distinctive restriction endonuclease analysis pattern. The eradication protocol was repeated to eliminate the infection produced by experimental challenge. The ferrets were then cohoused intermittently with naturally infected ferrets.
Results. The original H. mustelae infection was successfully eliminated by the eradication protocol. No recrudescence was observed in group I for 12 months nor for 3 months in group II after eradication. All ferrets became persistently reinfected with the challenge strain. The infection from the challenge strain was eradicated successfully. No ferrets in group I and all ferrets in group II became infected through cohousing.
Conclusions. These results suggest that though prior infection with H. mustelae may confer some protection against reinfection, such protection is not universal in all circumstances; that susceptibility to reinfection by contact with infected animals varies between individuals; and that age may be a factor in this individual variability. These results are applicable to studies of reinfection after eradication of H. pylori in humans.