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Helicobacter ISSN 1523-5378

Diagnosis of Helicobacter pylori Infection


Lurdes Monteiro,* Mónica Oleastro,* Philippe Lehours à and Francis Mégraud à
*Departamento de Doenças Infecciosas, Instituto Nacional Saúde Dr Ricardo Jorge, Lisboa, Portugal,  INSERM U853, F 33076 Bordeaux, àUniversité
Victor Segalen Bordeaux 2, Laboratoire de Bactériologie, F33076 Bordeaux, France

Keywords Abstract
Urease test, histology, culture, stool antigen
The articles published this last year in the field of Helicobacter pylori diagnosis
test, urea breath test, molecular methods,
antimicrobial susceptibility testing.
reported the development of in vivo histology, small improvements in some
invasive methods (urease test, culture, and histology) and new kits for the
Reprint requests to: Francis Mégraud, Labora- stool antigen tests. They also contributed to increasing our knowledge, by
toire de Bactériologie, Université Victor further exploration into specific conditions for the urea breath test and into
Segalen Bordeaux 2, Bat. 2B RDC Zone Nord, the significance of cagA antibodies. The role of serum markers of atrophy
33076 Bordeaux cedex, France. was also confirmed. Molecular methods are still being developed for direct
E-mail: francis.megraud@chu-bordeaux.fr
genotyping, detection of H. pylori and its clarithromycin resistance, either by
polymerase chain reaction or fluorescent in-situ hybridization. For the first
time, there was a report on a possible interest of magnetic resonance
spectroscopy.

A variety of tests for detecting Helicobacter pylori infec- alternative method for the diagnosis of H. pylori infec-
tion since the discovery of this pathogen have been tion [2].
described. While there has been no recent break- The new methods of magnifying endoscopy currently
through in this topic, a number of original articles com- developed have an added value compared to standard
ing especially from emerging countries were published endoscopy, i.e. the possibility of performing in vivo his-
last year on the different molecular and nonmolecular tology. A prospective study on 129 patients performed
diagnostic tests for H. pylori. in Turkey confirmed that the new method of high reso-
lution magnifying endoscopy is superior to standard
endoscopy for the diagnosis of H. pylori gastritis, and
Non-molecular Methods
identification of specific histopathologic features, such
as atrophy and intestinal metaplasia seems possible [3].
Invasive Tests
In a review of confocal laser endomicroscopy, Kiess-
Graham et al. published a review article providing rec- lich et al. highlighted the possibility of virtual histology
ommendations regarding when endoscopic gastric and its role in diagnosing H. pylori gastritis and targeting
mucosa assessment must be preferred rather than non- biopsy specimens [4].
invasive methods [1]. Interestingly, Kim et al. studied the sites for perform-
ing biopsies to detect H. pylori in 194 patients with gas-
tric cancer. They found that the best site was the upper
Endoscopy
body greater curvature (sensitivity of histology: 95.1%;
To obtain biopsies, an upper digestive endoscopy must 95% CI: 89.6–98.2), probably due to the proportion of
be performed. Cho et al. proposed a new method of atrophy and intestinal metaplasia, which were signifi-
standard endoscopic diagnosis of H. pylori: the phenol cantly lower than in the antrum and in the upper body
red mucosal pH test. A 0.1% phenol red solution was lesser curvature which were also tested [5].
sprayed on the gastric mucosa. The extent of staining,
expressed as a staining score, was positively correlated
Histology
with the urea breath test (UBT) values and with
H. pylori density as measured by histology. The pH mea- An alternative to Giemsa staining has been proposed in
sured in this study with an antimony electrode was Thailand [6]. A mixture of carbol fuschin and alcian
significantly higher in H. pylori infected mucosa. blue staining was compared blindly to Giemsa and
Therefore, endoscopic phenol red staining may be an hematoxylin & eosin staining on 423 histologic

ª 2009 The Authors


8 Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14
Monteiro et al. H. pylori Diagnosis

preparations. They found the same rate of positive sam-


Non-invasive Tests
ples and highlighted the low cost and simplicity of the
combined staining and its value in identifying goblet Urea breath test
13
cells in intestinal metaplasia. C-UBT has been shown numerous times to be the
The impact of mixed H. pylori infections was studied most accurate H. pylori diagnostic test. Several remain-
in 30 patients by Sheu et al. [7]. They found that the ing questions were addressed this year. Buzas and
seven patients with mixed infections had marginally Szeles compared UBT values after first-, second- and
higher scores of chronic inflammation and H. pylori third-line treatments. They found as in previous studies
density in the corpus and higher rates of intestinal that higher pretreatment UBT values were associated
metaplasia in the antrum (p = .005) compared to the with lower eradication rates but interestingly, they also
23 patients with a single strain. showed a marked tendency to increase UBT values for
An article referred to the new staging system for atro- the patients who failed, i.e. from 13.2& (CI: 7.3–19.1)
phy (OLGA) and its application in diagnostic practice to 19.2& (CI: 13.4–25.0) after second-line therapy and
[8]. It was also used to assess atrophic gastritis in 63 to 25.8& (CI: 19.8–31.2) after third-line therapy, but
H. pylori positive patients with various gastric diseases. they could not explain this phenomenon [15]. In
They found the OLGA staging system useful for the another study, UBT values were also found to be higher
assessment of the severity of atrophic gastritis and sim- when clarithromycin resistant H. pylori were present,
ple to use [9]. In another study concerning different but not when other resistances occurred [16].
risks of gastric cancer in populations, the OLGA staging The possibility of false positive results due to urease
mirrored the gastric cancer incidence [10]. positive bacteria from the oral cavity in patients with
The histogenesis of gastric carcinomas was re-evalu- atrophic gastritis was highlighted by Osaki et al. indicat-
ated by Kakinoki et al. [11]. They compared H. pylori- ing that the histologic status of the stomach, i.e. pres-
negative and -positive cases and found that carcinoma ence or absence of atrophy, must be considered in
cells could occur independently of the intestinal meta- interpreting the results [17]. To avoid false positive
plasia. If this finding is confirmed, it would indicate results, the capsule UBT can be used [18].
that intestinal metaplasia is not a precancerous but a There is another situation where false positive results
paracancerous lesion. may occur, i.e. in children younger than 6 years of age.
The cause may not only be the effect of endogenous
CO2 production but also other unidentified factors [19].
Urease test
In contrast to the RUT, the UBT was found to be unreli-
As in previous studies, the sensitivity of the rapid ure- able in patients with Billroth II gastrectomy [20]. Test-
ase test (RUT) was shown to be reduced in patients ing for the eradication of H. pylori is an important
with bleeding ulcers but the short-term use of standard aspect of clinical trial design and is of critical impor-
dose proton-pump inhibitor did not have an impact tance in the evaluation of new therapies for this patho-
[12]. gen. From the evaluation of Vakil et al. a single UBT,
As a solution to the low sensitivity of the RUT, some 4 weeks after treatment was as effective as two serial
authors proposed to increase the number of biopsies breath tests in confirming H. pylori eradication and the
tested up to four. Comparing one biopsy to four, the posi- incremental cost of the second breath test was very
tive results increased from 52 to 96%, respectively [13]. high with no incremental clinical benefit [21]. A study
performed in Israel to evaluate the indications for UBT
used by primary care doctors and to examine the
Culture
appropriateness of these indications to the accepted
Because of the slow growth and the particular require- guidelines of the Maastricht Consensus Report revealed
ments of H. pylori with regard to culture conditions, this a substantial noncompliance with guidelines for
area still remains a particular challenge. Sainsus et al. H. pylori testing among primary care doctors in this
tried to develop a liquid culture medium for the rapid country [22].
isolation, identification, and subsequent antibiotic sus-
ceptibility testing of H. pylori from biopsy specimens.
Stool antigen test
They selected Ham’s F12 medium with 5% horse serum
with antibiotics which provided the most rapid and reli- Stool antigen detection kits for the diagnosis of H. pylori
able growth. The CIM medium seems a promising solu- infection have been widely used because of their full
tion to solve some of the current problems concerning noninvasive nature. Mohammadian et al. presented a
H. pylori culture in solid media [14]. simple, rapid, and cost-effective production of a

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Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14 9
H. pylori Diagnosis Monteiro et al.

polyclonal antibody against alkyl hydroperoxide reduc- (ELISA)-IgG assays showed low sensitivity: 79.2%
tase (AhpC) of H. pylori for stool-antigen enzyme (95% CI: 77.3–81.0) and high specificity: 92.4% (95%
immunoassay [23]. Nguyen et al. evaluated the sensi- CI: 91.6–93.3); (3) ELISA commercial tests varied
tivity and specificity of the new monoclonal antibody- widely in performance (test for heterogeneity,
based antigen-in-stool enzyme immunoassay (Premier p = .0001); and (4) in-house ELISA with whole-cell
Platinum HpSA PLUS; Meridian Bioscience, Cincinnati, antigen tests showed the highest overall performance:
OH, USA) for diagnosis of H. pylori infection in Viet- sensitivity: 94% (95% CI: 90.2–96.7), specificity: 96.4%
namese children. The sensitivity was 96.6% (95% CI: (95% CI: 94.2–97.9), LR+: 19.9 (95% CI: 7.9–49.8),
93.3–98.5) and the specificity 94.9% (95% CI: 88.5– LR-2: 0.08 (95% CI: 0.04–0.15), DOR: 292.8 (95% CI:
98.3) [24]. In addition, Kuloğlu et al. evaluated the 101.8–841.7) [27].
diagnostic accuracy of a rapid immunochromatographic The Pyloriset EIA-G (Orion Diagnostics, Espoo, Fin-
stool antigen test (Rapid HpSA; LI_ NEAR Chemical, Bar- land), which is considered to be one of the most reliable
celona, Spain) and a practical low-dose 14C-UBT (Helip- tests in Europe, was used to detect H. pylori infection in
robe; Kibion, Uppsala, Sweden) in children before two groups of peptic ulcer disease patients, one group
and after eradication therapy. The sensitivity of Rapid vagotomized and the other medically treated, as well as
HpSA and 14C-UBT was 65% and 92.5% (p = .0003), community controls. Using positive histology and ⁄ or
respectively; the specificity of Rapid HpSA and 14C-UBT culture as the gold standard, the sensitivity of the sero-
was 92.3 and 85.5% (p = .180), respectively. After logic test was good but its specificity poor in contrast to
eradication therapy, endoscopy, 14C-UBT and Rapid 14
C-UBT results. However, no data were available on
HpSA were repeated. Both tests had the same specificity possible antibiotic treatments for these patients [28].
(100%) while the sensitivity of Rapid HpSA and 14C- Mohammadi et al. evaluated an in-house ELISA based
UBT was 60 and 100%, respectively [25]. However, the on soluble antigenic fractions of H. pylori proteins in
most interesting study compared six tests and the Iran. The sensitivity, specificity, and accuracy were over
results are reported in Table 1 [26]. 90% as was the performance of Helico Blot 2.1 (Gene-
labs Diagnostics, Singapore) and two foreign commercial
ELISA kits (Trinity kit and IBL kit) while the BioHit kit
Antibody detection
(Helsinki, Finland) did not perform as well [29].
Numerous antibody-based tests have been developed Anti-CagA antibodies (detected either by a specific
over the last decades. Leal et al. carried out a systematic ELISA or immunoblot) have been described as long
review and meta-analysis to evaluate the performance standing antibodies of interest to confirm H. pylori
of the different antibody-based detection tests available etiology years after the disappearance of the bacterium.
for H. pylori infection in children by determining sensi- Recently, Veijola et al. confirmed that CagA antibodies
tivity and specificity as well as additional accuracy val- detected by immunoblot (Helico Blot 2.1) could still be
ues relevant to clinical practice (positive and negative detected in 87% of the patients 10 years or more after
likelihood ratios (LR+ and LR)) and the diagnostic odds a successful H. pylori eradication [30]. Studies were per-
ratio (DOR)). The results were as follows: (1) western formed in several countries with various results. In
blot (WB) tests showed high overall performance, sensi- Australia, the use of Helicoblot 2.1 detecting CagA anti-
tivity: 91.3% (95% CI: 88.9–93.3, specificity: 89% bodies improved the sensitivity of H. pylori detection in
(95% CI:85.7–91.9), LR+: 8.2 (95% CI: 5.1–13.3), LR-2: patients with noncardia gastric cancer from 79% with
0.06 (95% CI: 0.02–0.16), and DOR: 158.8 (95% CI: H. pylori ELISA to 94%. Interestingly, pepsinogen I lev-
57.8–435.8)); (2) enzyme-linked immunosorbent assay els showed the lowest median level to be in cases
which were negative by ELISA but positive by immu-
noblot [31]. In Mexico, CagA antibodies were associ-
Table 1 Summary of sensitivity and specificity of the stool antigen
tests cited by Blanco et al. [26] ated with young gastric cancer cases only, and were
also a risk factor for intestinal metaplasia [32]. In Esto-
Sensitivity Specificity nia, positivity for CagA antibodies was able to predict
the development of atrophy, particularly in the corpus
Immunodiagnostic ELISA 87.3 83.3 (OR: 7.0, 1.8–27.7). In addition, the prevalence of anti-
HpStAR 95 66.6 canalicular antibodies increased with the duration of
HpSA-EIA 92.5 72.2
the H. pylori gastritis (22–46% in 12 years) [33]. In
H. pylori Lihtest 83.6 66.6
contrast, in India, the prevalence of H. pylori detected
Immunoland HpSA 52.5 94.4
RAPID HpStAR 78.8 55.5 by H. pylori serology or CagA antibodies was compara-
ble both in gastric cancer patients and in controls [34].

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10 Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14
Monteiro et al. H. pylori Diagnosis

Serum biomarkers, especially those proposed by Bio- PCR and in situ hybridization, targeting a 76-bp region
Hit in the GastroPanel (pepsinogen I & II, gastrin 17, of H. pylori 16S rRNA, that is highly conserved in a
H. pylori Ab) have been tested in comparison to histol- large number of H. pylori strains and is specific to this
ogy. In the Kalixanda study (Northern Sweden), an bacterium. Both approaches were shown to be very
overall agreement of 96% was found. Sensitivity and sensitive and specific and combined together can be
specificity of the markers for atrophic gastritis were 71 used for specific detection, identification and quantifica-
and 98%, respectively [35]. The GastroPanel was also tion of H. pylori in biological samples, from humans,
able to differentiate Japanese patients into two groups animals, or environmental source [42]. In another
with ‘‘healthy’’ or ‘‘diseased’’ stomachs with a 94% study, real-time PCR targeting the 23S rRNA gene
accuracy. In total, 5% of the patients had advanced applied on aortic and left internal mammary artery
atrophic corpus gastritis [36]. biopsies, was used to demonstrate, for the first time,
With regard to cost effectiveness, a Markov model that acute coronary ischemia was significantly more pre-
was designed based on 237,900 Chinese males, aged valent in H. pylori-positive patients than in H. pylori-neg-
35–44 years and living in Singapore, who would be ative patients (p = .001), suggesting a pathogenic role of
screened and treated with eradication therapy. Serology this bacterium in atherosclerosis [43]. Finally, Gill et al.
appeared to be twice as cost effective as the UBT in developed a nanodiagnostic method using thermophilic
screening this population [37]. helicase-dependent isothermal amplication of ureC and
From two studies concerning the value of RAPIRUN gold nanoparticle probes for hybridization and colori-
test, a urinary antibody test. Nguyen et al. verified that metric detection of H. pylori DNA. This method allowed
in the Vietnamese population sensitivity, specificity, the detection of 10 CFU ⁄ mL within less than 1 hour and
and accuracy of the test were 79.5, 90.7, and 84.5%, provided a sensitivity of 92.5% and a specificity of
respectively [38]. These values are in accordance with 95.4%, with culture as the reference [44].
the overview presented by Yamamoto et al. concerning Sugimoto et al. identified 26 sets of primers used to
the diagnosis of H. pylori infection using the same detect H. pylori. They first tested their detection limit.
detection kit [39]. The development of a model for eco- Five of the 26 sets with a detection limit <100 CFU ⁄ mL
nomic evaluation related to the diagnosis accuracy of were then tested further. All produced some false posi-
near patient tests used in office laboratories, as opposed tive results. These results indicate that results of H.
to using hospital-based tests was presented by Fauli pylori detection by PCR outside of the stomach should
et al. [40]. be interpreted with caution. Identification based on the
presence of multiple specific genes could be the way
forward [45].
Molecular Methods

Detection Genotyping
Over the last year, novel molecular methods based Simultaneous genotyping of bacterial and host cells is
mainly on a real-time polymerase chain reaction (PCR) sometimes difficult due to the small amounts of sample.
have been described to improve the detection and char- To overcome this problem, Ryberg et al. used the multi-
acterization of H. pylori. A new multiplex fluorescence ple displacement amplification technique on minute
resonance energy transfer real-time PCR, for amplifi- amounts of gastric biopsy specimens. Then, the ampli-
cation of H. pylori ureA and human b-globin, was fied DNA was used for concurrent PCR-based genotyp-
developed, allowing quantification of the bacterial den- ing, of both H. pylori 16S rRNA, vacA, hsp60, ureI, sod,
sity by determination of the ureA ⁄ b-globin amount ureA and cagA genes, and human cytokine polymor-
ratio in gastric biopsies. Using this assay, a significantly phisms [46]. Puz et al. developed a novel noninvasive
increased bacterial density was found in macroscopic genotyping method, using stool specimens, based on
erosions when compared with the healthy portion of two H. pylori-specific biprobe real-time PCR assays tar-
the stomach (p < .01). This PCR was not able to detect geting the glmM and recA genes. Discrimination
the ureA gene in H. pylori-positive formalin-fixed paraf- between strains is made using the differences in the
fin-embedded biopsy sections, probably because DNA melting temperature of the amplicons. The sensitivity of
was broken and amplification of a 411-bp fragment was the assay on stool samples was 92.2% and the specific-
not possible [41]. To overcome the problem of exten- ity was 100%. A discriminatory capacity of 100% was
sive genetic polymorphism for precise PCR detection achieved when the sequence analysis of the glmM
of H. pylori, Liu et al. developed a novel molecular amplicon was performed. Due to its noninvasiveness
approach, based on real-time reverse transcriptase (RT)- and high accuracy in detection and discrimination of

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Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14 11
H. pylori Diagnosis Monteiro et al.

H. pylori strains, this genotyping assay has the potential satisfactory sensitivity on both fresh and paraffin-
of being used in large-scale studies, contributing to the embedded biopsies isolated from children (84.1 and
clarification of the transmission pathways of this organ- 80.7%, respectively), previously diagnosed as H. pylori-
ism [47]. Paraffin-embedded gastric biopsies were used positive by histology and RUT. When used to screen
for H. pylori genotyping with success (97% of the clarithromycin resistance genotypes, using probes tar-
cases). The genotyping enabled a comparison of the geting the 23S rRNA point mutations, the FISH test was
prevalence of cagA positive, vacA s1m1 genotypes in more sensitive in detecting mixed susceptible and resis-
high and low risk areas for gastric cancer in Colombia tant populations than did the agar-dilution method.
(84.3 and 60.5%, respectively) [48]. The authors emphasize that clarithromycin resistance
Kanada et al. evaluated the accuracy of immunohis- should be assessed in biopsies both from the antrum
tochemistry for genotyping the cagA East-Asian EPIYA and the corpus, as in one-third of children with mixed
motif. They used a monoclonal antibody against this infection the resistant strains were found in the fundus
motif and tested it on gastric biopsy samples from Japa- only [52]. Woo et al. developed a dual-priming oligo-
nese patients. Compared to sequencing of the cagA nucleotide (DPO)-based multiplex PCR to detect both
3¢-region, containing the EPIYA motifs, the new assay H. pylori infection and the most common point muta-
showed a sensitivity of 93.2% and a specificity of tions occurring in the 23S rRNA conferring resistance
72.7%, suggesting a further optimization to be useful as to clarithomycin (A2142G and A2143G), directly on
a cagA typing method [49]. gastric biopsy specimens. The DPO-based multiplex was
slightly less sensitive in identifying H. pylori-positive
cases than histology, but was able to identify more clar-
Antimicrobial Resistance
ithromycin-resistant strains than the phenotypic meth-
With the growing H. pylori resistance to antibiotics, ods. This assay proved to be fast, does not require
molecular diagnostic tests for an accurate and rapid expensive instrumentation, and can thus be valuable in
identification of these strains are an attractive alterna- countries with high prevalence of clarithromycin resis-
tive to the time consuming culture-based susceptibility tance [53]. Kawai et al. used fecal specimens to detect
testing. Thus, this is still an emergent field in H. pylori H. pylori and its resistance to clarithromycin using a
research, targeting in particular the clarithromycin nested PCR based on the 23S rRNA gene and sequenc-
resistance-associated gene mutations. A study employ- ing of the amplicons, prior to H. pylori treatment. They
ing Scorpion primers, which combine a primer and a obtained a 94.3% eradication rate in the tailored group
probe in a single molecule, was reported by Burucoa and 71.4% in the control group [54].
et al. to detect H. pylori and clarithromycin resistance Finally, Chisholm et al. assessed the potential benefits
directly on gastric biopsies. The Scorpion PCR was of the application to routine testing, of a novel algo-
highly sensitive and specific (98.3% and 92.5%) in rithm comprising a panel of three previously described
detecting H. pylori, using culture as the gold standard, PCR assays for detection and antibiotic susceptibility
and, as usual, it was more sensitive in detecting mixed testing of H. pylori. All culture-negative gastric biopsies
populations with susceptible and resistant strains than were first tested for H. pylori and Helicobacter heilmannii-
the E-test. Clarithromycin genotypes determined with like organisms by a multiplex PCR assay targeting vacA
Scorpion PCR were concordant with those obtained by and 16S rRNA genes, respectively. Then, in the positive
PCR-restriction fragment length polymorphism [50]. cases, antibiotic susceptibility to clarithromycin and tet-
Fluorescent in-situ hybridization (FISH) can be used to racycline was assessed by real-time PCR probe hybrid-
identify H. pylori and antibiotic resistance in biopsy ization and melting point analysis assays targeting the
specimens without PCR. Tajbakhsh et al. reported a 23S rRNA and 16S rRNA, respectively. The authors
FISH procedure targeting the H. pylori ribosomal RNA, demonstrated that PCR testing was particularly useful
and this assay showed a sensitivity and specificity of when H. pylori culture was unsuccessful, due to con-
97.9 and 100% respectively, for detection of H. pylori, tamination of the biopsy or when the specimen trans-
when compared to histology. However, the FISH assay, portation was delayed. Without this additional testing,
employing four different probes, one wild type and 16.9% of all patients examined could have been misdi-
three mutated, showed a weak sensitivity in detecting agnosed as H. pylori negative by culture only [55].
clarithromycin susceptibility-associated genotypes, as
only 19 of the 47 FISH-positive samples were recog- Finally, two articles were published using magnetic res-
nized with these probes [51]. Another study employing onance spectroscopy (MRS). The first study concerned
the FISH technology, performed using DNA ProbeMix gastric biopsies obtained from patients with various dis-
targeting the 16S rRNA H. pylori gene, showed a eases studied ex vivo by high resolution – magic angle

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12 Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14
Monteiro et al. H. pylori Diagnosis

spinning – MRS (HR-MAS-MRS) and compared to ul- urease test in bleeding peptic ulcers: a prospective case-control
trastructural data. Several amino acids, e.g. glycine, ala- study. J Clin Gastroenterol 2009;43:133–9.
13 Siddique I, Al-Mekhaizeem K, Alateeqi N, Memon A, Hasan F.
nine, choline, and triglycerides, were identified as
Diagnosis of Helicobacter pylori: improving the sensitivity of
possible markers of differentiation toward neoplastic CLOtest by increasing the number of gastric antral biopsies.
lesions. Such a technique could be applied in vivo [56]. J Clin Gastroenterol 2008;42:356–60.
In the second study, the metabolic profile of gerbils 14 Sainsus N, Cattori V, Lepadatu C, Hofmann-Lehmann R. Liquid
infected or not with H. pylori was studied on urine spec- culture medium for the rapid cultivation of Helicobacter pylori
from biopsy specimens. Eur J Clin Microbiol Infect Dis
imens. Results showed that H. pylori infection disturbs
2008;27:1209–17.
carbohydrate and amino acid metabolism and modifies 15 Buzas GM, Szeles I. Interpretation of the 13C-urea breath test
the gut microbiota [57]. This method should open a in the choice of second- and third-line eradication of Helicobact-
new field of exploration of H. pylori infection. er pylori infection. J Gastroenterol 2008;43:108–14.
16 Kawai T, Kawakami K, Kataoka M, Itoi T, Takei K, Moriyasu F,
et al. A study of the relationship between Helicobacter pylori
Conflicts of Interest microbial susceptibility, 13C-urea breath test values. Hepatogast-
roenterology 2008;83:786–90.
The authors have declared no conflicts of interest. 17 Osaki T, Mabe K, Hanawa T, Kamiya S. Urease-positive bacteria
in the stomach induce a false-positive reaction in a urea breath
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14 Journal compilation ª 2009 Blackwell Publishing Ltd, Helicobacter 14 (Suppl. 1): 8–14