Helicobacter pylori enfeksiyonu tanısında tanısal yöntemler

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Introduction
Helicobacter pylori (H.pylori) is a spiral-shaped, gramnegative bacterium that is found in the gastric mucous
layer or adherent to the epithelial lining of the stomach.
Most persons who are infected by H.pylori never suffer
any symptoms related to the infection; however H.pylori
causes chronic gastritis (95%), gastroduodenal ulcers
(70-80%) and duodenal structural and functional
abnormalities. Infected persons have 2- to 6-fold increased risk of developing gastric cancer and mucosal-associated-lymphoid-type (MALT) lymphoma (1). It has
been classified as a Class I carcinogen. The majority of
patients having symptoms or signs of these diseases
were given long term medications such as H2
blockers
and proton pump inhibitors. These medications relieve
ulcer-related symptoms, heal gastric mucosal inflammation, and may heal the ulcer but they do not treat the
infection and there is no chance of permanent cure and
the majority of ulcers recur. Eradication of this infection with appropriate antibiotic therapy has been shown
to decrease ulcer recurrence <10% in one year versus a
60-100% one year recurrence with use of anti-ulcer
medications alone. Given the clinical importance of
H.pylori, there is a need for prompt and accurate diagnosis followed by eradication therapy and confirmation
of eradication after therapy.
H.pylori infection can be diagnosed by invasive and
non-invasive techniques (Table I) (2). Invasive techniques require endoscopy and mucosal biopsy and
specimens that will later be subjected to culture, rapid
urease test or histological examination. However
endoscopy is expensive, unpleasant for patients and carries a small but definite risk of complications. Thus, the
*Department of Nuclear Medicine, Ufuk University School of
Medicine
Reprint request: Dr. Emel Öztürk, Department of Nuclear Medicine,
Ufuk University School of Medicine, Konya Yolu No: 82-86 Balgat-
06520, Ankara
E-mail: eozturk@ufuk.edu.tr
Date submitted: April 12, 2007
Accepted: June 15, 2007
DERLEME/REVIEW ARTICLE Gülhane Týp Dergisi 2008; 50: 60-64
© Gülhane Askeri Týp Akademisi 2008
Summary
Helicobacter pylori is a spiral, gram negative bacterium and
found to be associated with gastritis, peptic ulcer disease,
gastric adenocarcinoma and MALT lymphoma. Thus there is
an increasing importance of the treatment of Helicobacter
pylori and thereby a great need for simple and accurate
diagnostic methods. Helicobacter pylori infection can be
diagnosed by invasive and non-invasive techniques. Invasive
diagnostic methods require mucosal biopsy during
endoscopy and then subjecting them to culture, rapid urease test, polymerase chain reaction or histologic analysis.
Non-invasive methods include antibody detection (serology), stool antigen and urea breath tests.
Key words: Helicobacter pylori, rapid urease test, urea
breath test
Özet
Helicobacter pylori enfeksiyonu tanýsýnda tanýsal yöntemler
Helicobacter pylori, spiral yapýsýnda gram negatif bir bakteridir ve gastrit, peptic ülser, gastrik adenokarsinoma ve
MALT lenfoma ile iliþkili olduðu gösterilmiþtir. Bu nedenle
tedavisi gittikçe önem kazanmaktadýr ve basit, doðru
tanýsal yöntemlere gereksinim vardýr. Helicobacter pylori
enfeksiyonu tanýsý invaziv ve noninvaziv yöntemler ile konulabilir. Ýnvaziv tanýsal yöntemler endoskopik iþlem sýrasýnda
biyopsi alýnmasýný ve bu doku parçalarýnýn daha sonra
kültür, hýzlý üreaz testi, polimeraz zincir reaksiyonu veya
histolojik analize tabi tutulmasýný gerektirir. Noninvaziv
yöntemler ise, serumda antikor saptanmasý (seroloji), gaitada antijen ve üre nefes testlerini içerir.
Anahtar kelimeler: Helicobacter pylori, hýzlý üreaz testi,
üre nefes testiCilt 50 · Sayý 1 · Gülhane TD Helicobacter pylori infection · 61
use of non-invasive tests to diagnose H.pylori infection
is becoming more frequent. Indeed, recent studies have
demonstrated that a strategy of 'test and treat' for
H.pylori in uninvestigated, young (<50 years), dyspeptic patients in primary care is safe and reduces the need
for endoscopy. Non-invasive methods include serology,
stool antigen test and urea breath tests (UBT). There
are two different types of non-invasive tests: direct and
indirect (Table I). The direct tests look for evidence of
the presence of H.pylori. The stool test assesses the presence of bacterial antigens in stool. Indirect tests assess
the presence of the infection by evaluating indirect evidence such as the presence of antibodies to H.pylori, or
the amount of labeled CO2
in the breath, an expression
of the urease activity of the bacterium.
Invasive techniques
Histology
Histological examination of biopsy samples taken
during endoscopy is usually considered 'the gold standard' for the diagnosis of H.pylori. But owing to the
patchy distribution of H.pylori in gastric mucosa, the
biopsy-based tests may suffer from sampling error (3,4).
Furthermore, histological examination is highly dependent on the experience of the pathologist, and high
inter-observer variation has been reported (4,5).
Rapid urease test (CLO test)
Biopsies of gastric mucosa are placed in a gel containing urea, and the subsequent ammonia production
causes a pH change, which is observed as a color
change. Besides suffering from biopsy sampling error,
the CLO test depends greatly on the pH of the media
and the amount of the urea in the medium. These factors may vary in different products and thereby influence the results obtained with other tests (3,6).
Culture
Culture is the most specific diagnostic method for
H.pylori infection but its sensitivity is low. The role of
culture for primary diagnosis is limited but it is an
important method as isolates for the traditional susceptibility testing are obtained. Although routine susceptibility testing for H.pylori is not recommended, increasing resistance rates to metronidazole and clarithromycin might make routine susceptibility more popular (7).
Non-invasive techniques
Serological tests
Serological tests are based on the detection of specific anti-H.pylori IgG antibodies in a patient's serum.
While serological tests are simple and easy to perform,
they are not reliable tests for the diagnosis of H.pylori
infection in elderly people because of poor antibody
production, or for determination of eradication of
H.pylori, since it remains positive for a long period
despite adequate treatment (6,8). Serological tests are
not able to distinguish between active infection and a
previous exposure to H.pylori. Different commercial
kits also have different levels of diagnostic accuracy
(range 68-82%) (9,10).
Stool antigen test
An enzyme immunoassay, which detects the presence of antigen in stool specimen, has recently become
available. This assay has undergone extensive testing for
the initial diagnosis of the H.pylori infection and in the
confirmation of eradication after treatment. Several
studies have suggested that polyclonal antibody test is
comparable to the ure breath test in the initial diagnosis
of H.pylori infection (sensitivity 93.2% and specificity
93.2%). It has been reported that stool antigen test is
less accurate than UBT in the post-treatment setting
(11). Recently it has been reported that monoclonal
technique has higher sensitivity than the polyclonal one
especially in the post-treatment setting (12).
Urea breath tests
H.pylori produces urease, an enzyme that splits urea
into ammonia and carbon dioxide. The production of
high amounts of urease by H.pylori has been used in the
development of urea breath tests (Figure 1). Patients
ingest urea labeled
13C or
14C. Hydrolysis of urea occur
within the mucous layer and results in the production
of ammonia and labeled CO2
. Labeled CO2
diffuses
into the blood vessel and can be detected in the breath
as a marker of infection (Figure 1). UBTs with either
13C or
14C-urea sample the whole stomach and reflect
the active infection. Both tests have been proved to be
very accurate, with reported sensitivities of 97-100%
and specificities of 95-100% for both diagnosis and
proof of eradication of H.pylori infection after therapy
(5,13-18), and it is even used as gold standard in some
studies. While the two isotopes seem to offer similar
diagnostic accuracy,
13C-UBT has the inconvenience of
requiring (a) more complex and expensive equipment
on site or else analysis off-site by an external laboratory
and (b) administration of a test meal and cold urea to
the patient. These are not necessary with
14C, and the
test is thus more simple, faster and cheaper. The diagTable I. Diagnostic tests for Helicobacter pylori infection
______________________________________________________
Noninvasive techniques
_______________________________
Direct Indirect
______________________________________________________
Histology Stool antigen test Urea breath test
Rapid urease test Serological tests
Culture
______________________________________________________
Invasive techniques62 · Mart 2008 · Gülhane TD Öztürk
nostic reproducibility of the
14C-UBT is very good
(19).
The routine test protocol of
14C-UBT requires
ingestion of
14C-urea, collection of breath samples at
frequent intervals using a liquid CO2
trapping medium
(usually hyamine), addition of a liquid scintillation
cocktail and counting with a ß-scintillation counter.
Several different methodological approaches have been
proposed to develop a more simple and quicker test.
Recently a new device for
14C-UBT has been described
(20). It uses a flat, dry breath card, which is able to
adsorb exhaled via chemical binding (Heliprobe
BreathCard). This card is then read using a small
analyser with a slot into which the card is put
(Heliprobe analyzer). In addition to the simple and easy
collection of breath, this system prevents accidental
ingestion of hazardous organic CO2
absorber solutions
during breath sampling. Clinical validation of this new
system showed high diagnostic accuracy for both diagnosis and proof of eradication of H.pylori infection after
therapy (20,21). Additional advantages of the Heliprobe
system are the shorter test time and the low cost. Breath
samples are analyzed with a ß-scintillation counter in
14C-UBT and with a mass spectrometer in
13C-UBT.
Because both items of equipment are expensive, analysis can be done in an external laboratory by mail order
and results are usually obtained a few days later. In contrast, with the Heliprobe system the results are obtained
in half an hour on-site and the analyzer is much cheaper than either a ß-scintillation counter or a mass spectrometer.
Various factors affect the results of the UBT and this
test has been extensively modified since its first description. The differences concern variations in the doses
and forms of
14C-urea, patient preparation before the
test, sampling time, number of breath samples and
modes of quantification.
The original
14C-UBT system used relatively high
activities (200-400 kBq) and multiple breath sampling.
Later studies showed that the diagnostic accuracy of
14C
UBT is maintained even with low doses such as 37 kBq
(1 µCi) and single breath sample (15,22).
The UBT indirectly detects gastric H.pylori by measuring urease activity. Sources of urease other than
H.pylori, such as bacterial overgrowth in the stomach or
upper intestine, may rarely cause false-positive test
results (22). Urease-producing bacteria are also present
in the oropharynx and may cause false-positive results,
especially in early breath samples. Late breath sampling
may result in false-negative results because of emptying
of urea from the stomach. Several procedures to avoid
contamination of breath by the oropharyngeal flora
have been suggested, including mouth washing, simultaneous meal to delay gastric emptying, and performance of multiple breath sampling. Another more simple and effective method is the use of
14C-urea in a
gelatin capsule, thus bypassing the oropharynx. Hamlet
et al. have reported that when the
14C-urea is supplied
in a capsule, a single 10-min breath sample is highly
accurate (100% sensitivity and specificity) for the diagnosis of H.pylori infection. They have compared the
capsule method with the urea drink method and found
the former to be more reliable because no overlapping
in activity has occurred between H.pylori -positive and -
negative patients; by contrast, conventional breath testing has showed overlapping during the whole 30-min
test period. Their study has also showed that a fatty test
meal lowers the
14CO2
excretion during the first 20
min and may adversely affect the accuracy of a rapid
UBT (16). Other advantages of the capsule form
include commercial availability, no risk of spills, shorter test duration and a lower radiation dose.
The expression of results of UBT varies between
investigators. Henze et al. and Veldhuyzen van Zanten
et al. have used CPM (23,24). Because CPM is affected
by chemical or color quenching, chemical changes of
the cocktail and methods of sample preparation, Pathak
et al. have strongly suggested the use of DPM counts
(25).
Some authors have used formulas to correct for
body weight or body surface to account for differences
in endogenous CO2
production, the results being
expressed as recovery standard units [(% of administered dose recovered/mmol CO2
trapped) × body
weight (kg)] (13,15). However, neither of these factors
has been proved to influence the results of the breath
test. Indeed, it has even been reported that uncorrected
Figure 1. The principle of urea breath testCilt 50 · Sayý 1 · Gülhane TD Helicobacter pylori infection · 63
counts result in better distinction between H.pylori -
positive and -negative patients (16,24,25).
Adequate patient preparation is important if accurate
results are to be obtained with
14C-UBT. Fasting state
of patients is important first step for the test (26). A
large number of investigators have reported that the
UBT becomes false negative during therapy with proton pump inhibitors, lansoprazole, bismuth compounds, antibiotics and ranitidine (27,28). Preliminary
reports indicate that addition of citric acid to the urea
solution/capsule may diminish the negative effect of
acid-inhibitory drugs on the accuracy of UBT (30). The
exact value of acidified urea needs further verification.
Carbon-13 is a non-radioactive isotope, but
13CUBT is more expensive because it requires mass spectrometry.
14C has a physical half-life of about 5,000
years, raising the question of the risks of radiation exposure. Because nearly the entire ingested isotope is rapidly excreted in urine or breath over the following 72 h
and only a small amount of isotope is used, the test
actually entails low radiation exposure (3 µSv) (31,32).
In fact, the dose is less than the natural background
radiation in one day; it is similar to the additional radiation received from cosmic rays during a 1-h jet flight.
In terms of other radiological investigations, the dose
from one
14C-UBT is equivalent to roughly one-seventh of that from a chest x-ray (20 µSv), or one-thousandth of that from a barium meal (3 mSv). For this reason, in 1997 the Nuclear Regulatory Commission permitted in vivo diagnostic use of capsules containing 1
µCi of
14C-urea without a license (33). This margin of
safety makes it acceptable to use the test for sequential
studies.
Conclusions
∗ Non-invasive tests are becoming more important in
the clinical management of dyspeptic patients.
∗ A number of clinical guidelines recommend noninvasive testing in dyspeptic patients, followed by treatment of H.pylori.
∗ European Helicobacter pylori study group has recommended the use of urea breath test or stool testing in
initial diagnosis of H.pylori infection and stool test may
be an alternative to UBT after treatment.
∗ Different serological tests have varying levels of
accuracy and are not appropriate for confirming eradication.
∗ The accuracy of low dose, single sample
14C-UBT is
high.
∗ A low dose capsule
14C-UBT gives less than the natural background radiation in one day.