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ORIGINAL ARTICLE

Carbon-14 urea breath test: does it work in patients with partial gastric resection?

Fuat Dede1,5 • Hüseyin Civen2 • Faysal Dane3 • Mehmet Aliustaoglu4 •

Serdar Turhal3 • Halil Turgut Turoglu1 • Sabahat Inanir1

Received: 3 March 2015 / Accepted: 8 July 2015 / Published online: 18 July 2015

� The Japanese Society of Nuclear Medicine 2015

Abstract

Objective The diagnostic value of Carbon-14 urea breath

test (C-14 UBT) in the detection of Helicobacter pylori (H.

pylori) infection in non-operated patients has been proved.

However, the efficacy of C-14 UBT in patients with partial

gastric resection (PGR) has not been evaluated yet. Herein,

the results of the C-14 UBT and H. pylori stool antigen test

(HpSAT) in this patient group were compared with the

endoscopic findings.

Methods Multi-breath samples C-14 UBT and HpSAT

were performed in all patients on the same day. Histology

was used as a gold standard for testing C-14 UBT and

HpSAT diagnostic efficacies.

Results 30 patients (mean age: 54.6 ± 11 year) with

PGR were included. The sensitivity and specificity of

standard C-14 UBT were 29 and 100 %, respectively.

When breath samples were collected at 20th min, and[35 CPM was selected as radioactivity threshold, the sensitivity

raised to 86 % without any loss of specificity. The

specificity and sensitivity of the HpSAT were 71 and 96 %,

respectively.

Conclusions The sensitivity of the standard C-14 UBT

was very poor for patients with PGR, and results of HpSAT

were superior in this population. Certain modifications are

needed if C-14 UBT is to be used in PGR patients.

Keywords Carbon-14 urea breath test � Stool antigen test � Partial gastrectomy � Gastric cancer � Helicobacter pylori

Introduction

After the discovery of Helicobacter pylori (H. pylori) in

1982, epidemiologic studies have revealed that it is a very

common pathogen in the society that has infected nearly

half of the world’s population [1, 2]. Geographic area, age,

race, and socioeconomic status determine the prevalence of

H. pylori infection [3]. H. pylori plays an important role in

the development of duodenal ulcer (responsible for

90–95 % of all duodenal ulcers), atrophic gastritis and

gastric cancer (represents nearly 5.5 % of all cancers and

25 % of all infection-related cancers) [4–6]. Because of

this, diagnosing this bacteria and thereafter starting multi-

drug eradication therapy is important. Invasive (endoscopy,

histology, rapid urease test, and culture) and non-invasive

[serology, urea breath test (UBT), and H. pylori stool

antigen test (HpSAT)] methods are used to diagnose this

microorganism [7].

UBT is a very successful method for both initial diag-

nosis of H. pylori and monitoring response to treatment [7].

The principles and mechanisms of the test are as follows:

The labeled urea [with either non-radioactive carbon-13

(C-13) or radioactive carbon-14 (C-14)] in the test material

& Fuat Dede [email protected]

1 Department of Nuclear Medicine, Marmara University

School of Medicine, Istanbul, Turkey

2 Nuclear Medicine Clinic, Kocaeli State Hospital, Kocaeli,

Turkey

3 Department of Medical Oncology, Marmara University

School of Medicine, Istanbul, Turkey

4 Internal Medicine Clinic, Kartal Dr. Lutfi Kirdar Research

and Training Hospital, Istanbul, Turkey

5 Nukleer Tip Anabilim Dali, S.B. Marmara Universitesi

Pendik Egitim ve Arastirma Hastanesi, -1 kat A1 Blok Fevzi

Cakmak Mahallesi Mimar Sinan Caddesi No:41 Ustkaynarca,

Pendik, Istanbul, Turkey

123

Ann Nucl Med (2015) 29:786–791

DOI 10.1007/s12149-015-1005-3

is degraded to carbon dioxide (CO2) and ammonia with the

presence of urease, an enzyme that is synthesized by H.

pylori. The labeled CO2 is then absorbed from gastric

mucosa and exhaled. The detection of the labeled CO2 in

the exhaled breath confirms the diagnosis of H. pylori.

C-14 UBT is a cheap and rapid test that does not require a

test meal [8]. On the other hand, the need for authorized

centers for handling radioactive material and transport

problems limit its usage. Since C-13 is a stable isotope, it

can be safely used in children and childbearing women.

The major disadvantage of C-13 UBT is its higher cost.

The risk of developing cancer in the residual stomach is

increased in patients who underwent partial gastric resec-

tion (PGR) due to benign (ulcer, etc.) or malignant causes

[9–11]. The incidence of gastric stump cancer reached up

to 2 % in this group [12]. As in non-operated patients,

atrophic gastritis, intestinal metaplasia, dysplasia, and

finally gastric cancer are closely related with the H. pylori

infection in PGR patients [1]. Therefore, these high-risk

cases should be screened for H. pylori, and eradication

therapy should be given when the infection is detected [9–

11].

The importance of UBT in patients without history of

gastric surgery is undebatable [13, 14]. However, studies

with C-13 UBT showed that with its relatively low sensi-

tivity and specificity rates (77 and 89 %, respectively), the

test failed to meet the expectations after PGR [15–18].

Decreased gastric volume, decreased bacterial load, rapid

gastric emptying, and increased gastric pH could be

responsible for the failure of the C-14 UBT in partially

gastrectomized patients [17, 19, 20]. Certain modifications

were recommended in order to suppress the effects of these

factors. On the other hand, the diagnostic performance of

C-14 UBT in this patient group has not been fully inves-

tigated. The aim of this study was to evaluate the perfor-

mance of the standard and modified C-14 UBT and to

compare results with HpSAT and endoscopy in patients

with PGR.

Materials and methods

Patients

A total of 30 gastric cancer patients (F/M = 8/22; mean

age = 54.6 ± 11 years; range = 30–74 years) with PGR

were included in this prospective study. Billroth II pro-

cedure was performed in 26 patients (87 %), Roux-en-Y

anastomosis in 3 patients (10 %), and wedge resection

was done in one case (3 %). The pathology was reported

as adenocarcinoma in all but two patients (94 %). Gas-

trointestinal stromal tumor was the diagnosis in one

patient (3 %) and maltoma in the other one (3 %).

Patients who received eradication therapy for H. pylori

after surgery and patients who were found to have taken

medication (bismuth, antibiotics, proton pump inhibitors,

H2 blockers, and antacids) 4 weeks before the diagnostic

tests were excluded from the study. This study was

approved by the institutional ethics committee and all

patients gave informed consent for participation in the

study.

Histopathological analysis

All patients underwent postoperative routine fiber-optic

esophagogastroscopy 15 days to 4 weeks (median

2.5 weeks) before the non-invasive tests (C-14 UBT and

HpSAT). During procedure, multiple mucosal biopsy

samples were obtained and stained with hematoxylin and

eosin and modified Giemsa. Specimens were examined for

the presence of H. pylori. Histology was used as a gold

standard for testing C-14 UBT and HpSAT diagnostic

efficacies.

Carbon-14 Urea breath test

After an overnight fast, C-14 UBT (Heliprobe � System,

Kibion AB, Uppsala, Sweden) was performed for all

enrolled patients. After ingestion of 37 kBq (1 lCi) C-14 Urea capsule with 50 mL water, the breath samples were

collected at 10th, 20th, and 30th min after ingestion.

During this 30-minute C-14 UBT urea reaction period,

patients lay on the left side horizontally. The results were

expressed as both counts per minute (CPM), and grading

[0–1 (negative for H. pylori infection, CPM B 50), and 2

(positive for H. pylori infection, CPM [ 50)] as suggested by the manufacturer.

H. pylori stool antigen test (HpSAT)

For all cases, HpSAT was performed simultaneously with 14 C-UBT on the same day. H. pylori antigens in feces were

investigated by monoclonal antibodies with one-step col-

ored immunochromatography (ImmunoCard STAT!

HpSA, Meridian Diagnostics Inc., Cincinnati, Ohio, USA)

technique according to the standard manufacturer

recommendations.

Statistics

Statistical analysis was performed by GraphPad InStat

Version 3.00 (GraphPad Software Inc, Sandiego, Califor-

nia, USA) and MedCalc Version 11.6.1.0 (MedCalc Soft-

ware, Mariakerke, Belgium). Based on analyzed data,

Kruskal–Wallis test (non-parametric ANOVA), the Mann–

Whitney U-test, Spearman non-parametric correlation,

Ann Nucl Med (2015) 29:786–791 787

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ROC curve analysis, and comparison of ROC curves were

used. A p value less than 0.05 was considered significant.

Results

The interval between the surgery and UBT ranged from

27 days to 21 years (mean = 27.7 ± 47 months; med-

ian = 14 months). H. pylori was detected in 7 patients

(23.3 %) at follow-up endoscopy. The mean ages of H.

pylori positive and negative patients were 53 ± 7 and

55 ± 23 years, respectively. No statistically significant

difference in terms of age was detected between these two

groups (Mann–Whitney U-test).

UBT results

With standard C-14 UBT criteria, H. pylori was detected in

2 patients at 10th min, 4 patients at 20th min, and 5 patients

at 30th min. The sensitivity rates for each time point were

29, 57, and 71 %, respectively, while the specificity was

100 % for all (Table 1).

UBT radioactivity counts (CPM)

The C-14 UBT radioactivity counts in non-operated H.

pylori (?) patients were reported to change between 69

CPM and 770 CPM (median and mean 269 and 300 CPM,

respectively) [21]. When the radioactivity counts in H.

pylori positive and negative patients were analyzed sepa-

rately, we found that the breath sampling time did not have

a statistically significant impact on radioactivity count rates

(Kruskal–Wallis test, P [ 0.05, Table 2). When endoscopy

was assumed to be the gold standard, the radioactivity

thresholds for 10th, 20th, and 30th min were found as

[23, [35, and [29 CPM, respectively, by ROC curve analysis (Fig. 1). The sensitivity, specificity, negative

predictive value (NPV), positive predictive value (PPV),

and accuracy are presented in Table 1. Although no sta-

tistically significant difference was found between three

ROC curves, the best performance was achieved with 20th

min breath sampling and radioactivity threshold[35 CPM.

HpSAT results

It gave true positive results in 5 patients and false-positive

result in one patient. The sensitivity and specificity of

HpSAT in our study population were 71.4 and 95.7 %,

respectively (Table 1).

Elapsed time after surgery

Although not statistically significant, inverse relation

between the prevalence of H. pylori and the elapsed time

after surgery was found in partially gastrectomized patients

(Spearman non-parametric correlation, r: -0.69, p [ 0.05, Fig. 2).

Discussion

Regardless of the type and cause of the surgery, patients

with PGR are prone to developing gastric cancer [9–11].

Enterogastric reflux and H. pylori colonization in the

residual stomach are the main risk factors for the occur-

rence of malignancy [16]. In this group of subjects,

Table 1 Results of C-14 UBT and HpSAT

t a

Test Sensitivity Specificity NPV PPV Accuracy

Threshold b

C-14 UBT

10 [50c 28.6 100 82.1 100 83.3 20 [50c 57.1 100 88.5 100 90 30 [50c 71.4 100 92 100 93 10 [23 85.7 86.9 95.2 66.7 87 20 [35 85.7 100 95.8 100 97 30 [29 71.4 100 92 100 93 HpSAT 71.4 95.7 91.7 83 90

C-14 UBT d and HpSAT 85.7 95.7 95.7 85.7 93.3

Bold values indicate better results

NPV negative predictive value, PPV positive predictive value, UBT urea breath test, HpSAT H. pylori stool

antigen test a breath sampling time (minute)

b counts per minute (CPM)

c standard radioactivity threshold

d Standard C-14 UBT

788 Ann Nucl Med (2015) 29:786–791

123

screening for H. pylori is important in order to start multi-

drug eradication therapy after diagnosis for preventing

undesirable consequences. The utility of UBT in normal

population has been proved. However, its usage in PGR is

controversial due to the reported relatively low sensitivity

and specificity rates (77 and 89 %, respectively) [15, 16].

Up to now, all of the studies investigating the performance

of UBT in partially gastrectomized patients have used C-13

non-radioactive isotope as screening tool. Although no

statistically significant difference in terms of sensitivity

and specificity was found between C-13 and C-14 UBT in

normal population, the exact results of C-14 UBT in this

patient group have not been revealed yet [22].

Herein, we studied multi-breath sample C-14 UBT in

patients who underwent distal subtotal gastrectomy due to

gastric malign tumors. Similar to previous results obtained

with C-13 UBT, very low sensitivity rate (28.6 %) was

found for standard C-14 UBT (10th min breath sample and

Table 2 Comparison of radioactivity counts at 10th,

20th, and 30th min in H. Pylori

(?) and (-) patients

10th min 20th min 30th min p:

H. pylori (?) (n:7) 65.1 ± 72 CPM 77 ± 66 CPM 72 ± 58 CPM 0.84*

H. pylori (-) (n:23) 15.3 ± 10 CPM 14.3 ± 9 CPM 15.4 ± 8 CPM 0.80*

p: 0.0011** <0.0001** 0.0174**

p values in bold and italics are statistically significant

CPM counts per minute

* Kruskal–Wallis test (non-parametric ANOVA)

** Mann–Whitney U-test

Fig. 1 ROC curves generated from radioactivity counts belonging to 10th, 20th, and 30th min breath samples

Fig. 2 Statistically insignificant inverse relation between the prevalence of H. pylori and the elapsed time after surgery (r: -0.69, p [ 0.05)

Ann Nucl Med (2015) 29:786–791 789

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radioactivity count threshold [50 CPM) [16–18]. Asking the patient to stay in left lateral decubitus or supine posi-

tion during urease reaction, giving citric acid or anti-

motility drugs, direct spraying of the C-13 urea over the

gastric mucosa endoscopically, collecting breath samples

in later periods and decreasing the DOB (‘‘delta over

baseline’’) threshold were recommended in order to

increase the performance of UBT [17, 19, 20, 23, 24]. In

this study, obtaining unsatisfactory results with standard

C-14 UBT despite placing the patient in left lateral decu-

bitus position in the first 10 minutes showed that the

patient position itself might not be enough for improving

the test performance. However, the extension of the reac-

tion period after oral intake of C-14 labeled urea (i.e.,

collecting breath samples at 20th or 30th min) gave rise to

more than a twofold increase in sensitivity (28.6 % at 10th

min vs. 57.1 % at 20th min and 71.4 % at 30th min) as

reported in previous studies with C-13 UBT. We noticed

that the sensitivity of the standard radioactivity threshold

([50 CPM) was very low for PGR patients. In accordance with the results of the C-13 UBT studies, decreasing this

threshold might improve the sensitivity even with 10th min

breath samples without deteriorating the specificity

significantly.

The liquid forms of the test were not preferred anymore

due to the false-positive results caused by the urease pos-

itive oral bacterial flora [25]. In our study, rapidly disin-

tegrating capsules containing labeled urea were used

instead and no false-positive result was encountered.

Although not statistically significant, this study also

revealed an inverse relation between the prevalence of H.

pylori and the elapsed time after surgery in partially gas-

trectomized patients. In early postoperative period, the

prevalence is as high as that in the normal population [1].

However, due to the improper microenvironment for bac-

terial colonization in the residual stomach, this declines by

the time [1].

Combined use of at least two diagnostic tests was rec-

ommended for the detection of H. pylori infection [26]. For

this reason, C-14 UBT and HpSAT were compared with

invasive reference test. The sensitivity of the standard C-14

UBT was significantly lower than that of the HpSAT (29

vs. 71 %). However, when C-14 UBT’s criteria were

modified, results were changed in favor of C-14 UBT. It is

noteworthy that HpSAT gave false-positive result in one

patient and false negative in another one in whom C-14

UBT was strongly positive even with standard diagnostic

criteria. As a qualitative diagnostic method, HpSAT is

advantageous since it does not require adjusting a thresh-

old. However, the two frequently experienced complaints

in this group of patients, diarrhea and constipation, can

cause false-negative results [27, 28]. Giving false-positive

results in the presence of gastrointestinal bleeding is the

another disadvantage of this method [27, 29]. None of

these factors are anticipated to affect the performance of

UBT.

Relatively small sample size was the main limitation of

the study. Additionally, all of the participants suffered from

gastric cancer and most of them underwent Billroth type II

reconstruction. Because of these we could not perform

subgroup analysis according to type and cause of the sur-

gery. The relatively long time interval (max. 4 weeks)

between UBT and endoscopy and the variable post-op

periods was the other potential limitations of our study.

In conclusion, the sensitivity of the standard C-14 UBT

was very poor for patients with PGR, and the results of

HpSAT were superior in this patient population. As in C-13

UBT, promising results could be obtained with collecting

breath samples at a later time point and decreasing the

radioactivity cut-off level.

Conflict of interest All the authors state that there were no conflicts of interests when the manuscript was written.

References

1. Lin YS, Chen MJ, Shih SC, Bair MJ, Fang CJ, Wang HY.

Management of helicobacter pylori infection after gas-

tric surgery. World J Gastroenterol. 2014;20:5274–82.

2. Wroblewski LE, Peek RM Jr, Wilson KT. Helicobacter

pylori and gastric cancer: factors that modulate disease risk.

Clin Microbiol Rev. 2010;23:713–39.

3. Brown LM. Helicobacter pylori: epidemiology and routes of

transmission. Epidemiol Rev. 2000;22:283–97.

4. Testerman TL, Morris J. Beyond the stomach: an updated view of

helicobacter pylori pathogenesis, diagnosis, and treatment. World

J Gastroenterol. 2014;20:12781–808.

5. Herrera V, Parsonnet J. Helicobacter pylori and gastric adeno-

carcinoma. Clin Microbiol Infect. 2009;15:971–6.

6. Ford AC, Forman D, Hunt RH, Yuan Y, Moayyedi P. Heli-

cobacter pylori eradication therapy to prevent gastric cancer in

healthy asymptomatic infected individuals: systematic review and

meta-analysis of randomised controlled trials. BMJ.

2014;348:g3174. doi:10.1136/bmj.g3174.

7. Lopes AI, Vale FF, Oleastro M. Helicobacter pylori infection—

recent developments in diagnosis. World J Gastroenterol.

2014;20:9299–313.

8. Savarino V, Vigneri S, Celle G. The 13C urea breath test in the

diagnosis of Helicobacter pylori infection. Gut. 1999;45(Suppl

1):I18–22.

9. Takeno S, Noguchi T, Kimura Y, Fujiwara S, Kubo N, Kawahara

K. Early and late gastric cancer arising in the remnant stomach

after distal gastrectomy. Eur J Surg Oncol. 2006;32:1191–4.

10. Kaminishi M, Shimizu N, Hashimoto M, Sakai S, Oohara T.

Different carcinogenesis in the gastric remnant after gastrectomy

for gasric cancer. Cancer. 1996;77:1646–53.

11. Miwa K, Kamata T, Miyazaki I, Hattori T. Kinetic changes and

experimental carcinogenesis after Billroth I and II gastrectomy.

Br J Surg. 1993;80:893–6.

12. Takeno S, Hashimoto T, Maki K, Shibata R, Shiwaku H, Yamana

I, et al. Gastric cancer arising from the remnant stom-

ach after distal gastrectomy: a review. World J Gastroenterol.

2014;20:13734–40.

790 Ann Nucl Med (2015) 29:786–791

123

13. Gisbert JP, Pajares JM. Review article: 13C-urea breath test in

the diagnosis of Helicobacter pylori infection—a critical review.

Aliment Pharmacol Ther. 2004;20:1001–17.

14. Mauro M, Radovic V, Wolfe M, Kamath M, Bercik P, Armstrong

D. 13C urea breath test for Helicobacter pylori: evaluation of

10-minute breath collection. Can J Gastroenterol. 2006;20:775–8.

15. Tian XY, Zhu H, Zhao J, She Q, Zhang GX. Diagnostic perfor-

mance of urea breath test, rapid urea test, and histology for

Helicobacter pylori infection in patients with partial gastrectomy:

a meta-analysis. J Clin Gastroenterol. 2012;46:285–92.

16. Kato S, Matsukura N, Matsuda N, Tsukada K, Naito Z, Tajiri T.

Helicobacter pylori eradication therapy modulates acidity and

interleukin-1b mRNA levels in un-operated stomach and in remnant stomach after gastrectomy in gastric cancer patients.

Aliment Pharmacol Ther. 2006;24:278–84.

17. Togashi A, Matsukura N, Kato S, Masuda G, Ohkawa K, Taku-

naga A, et al. Simple and accurate (13)C-urea breath test for

detection of Helicobacter pylori in the remnant stomach after

surgery. J Gastroenterol. 2006;41:127–32.

18. Rino Y, Takanashi Y, Hasuo K, Ashida A, Harada H, Kawamato

M, et al. 13C-urea breath test in patients having undergone total

gastrectomy. Hepatogastroenterology. 2007;54:1601–3.

19. Kubota K, Hiki N, Shimuzu N, Shimoyama S, Noguchi C, Tange

T, et al. Utility of 13C urea breath test for Helicobacter pylori

detection in partial gastrectomy patients. Dig Dis Sci.

2003;48:2135–8.

20. Kubota K, Shimoyama S, Shimizu N, Noguchi C, Mafune K,

Kaminishi M, et al. Studies of 13C-urea breath test for diagnosis

of Helicobacter pylori infection in patients after partial gastrec-

tomy. Digestion. 2002;65:82–6.

21. Oztürk E, Yeşilova Z, Ilgan S, Arslan N, Erdil A, Celasun B,

et al. A new, practical, low-dose 14C-urea breath test for the

diagnosis of Helicobacter pylori infection: clinical validation and

comparison with the standard method. Eur J Nucl Med Mol

Imaging. 2003;30:1457–62.

22. Chey WD, Wong BC, Practice Parameters Committee of the

American College of Gastroenterology. American College of

Gastroenterology guideline on the management of Helicobacter

pylori infection. Am J Gastroenterol. 2007;102:1808–25.

23. Lombardo L, Masoero G, Della Monica P, Crocella L, Ruggia O,

Ravarino N, et al. Multiple sampling 13C urea breath test:

improvement of diagnosis in postgastrectomy patients. Minerva

Gastroenterol Dietol. 2003;49:181–6.

24. Urita Y, Maeda T, Ishihara S, Sugimoto M, Watanebe T, Domon

K, et al. Endocopic 13C-urea breath test for detection of Heli-

cobacter pylori infection after partial gastrectomy. Hepatogas-

troenterology. 2007;54:1891–4.

25. Peng NJ, Lai KH, Liu RS, Lee SC, Tsay DG, Lo CC, et al.

Capsule 13C-urea breath test for the diagnosis of Helicobacter

pylori infection. World J Gastroenterol. 2005;11:1361–4.

26. Ramis IB, de Moraes EP, Fernandes MS, Mendoza-Sassi R,

Rodrigues O, Juliano CR, et al. Evaluation of diagnostic meth-

ods for the detection of Helicobacter pylori in gastric biopsy

specimens of dyspeptic patients. Braz J Microbiol.

2012;43:903–8.

27. Shimoyama T. Stool antigen tests for the management of Heli-

cobacter pylori infection. World J Gastroenterol.

2013;19:8188–91.

28. Mégraud F, Lehours P. Helicobacter pylori detection and

antimicrobial susceptibility testing. Clin Microbiol Rev.

2007;20:280–322.

29. van Leerdam ME, van der Ende A, ten kate FJ, Rauws EA, Tytgat

GN. Lack of accuracy of the noninvasive Helicobacter pylori

stool antigen test in patients with gastroduodenal ulcer bleeding.

Am J Gastroenterol. 2003;98:798–801.

Ann Nucl Med (2015) 29:786–791 791

123