Clinical Microbiology and Infection 24 (2018) 780.e5e780.

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Research note

Primary antibiotic resistance of Helicobacter pylori in Chinese patients:

a multiregion prospective 7-year study
D.-S. Liu 1, y, Y.-H. Wang 1, y, Z.-R. Zeng 2, Z.-Y. Zhang 3, H. Lu 4, J.-M. Xu 5, Y.-Q. Du 6, Y. Li 7,
J.-B. Wang 8, S.-P. Xu 9, Y. Chen 10, C.-H. Lan 11, H. Cheng 12, M.-D. Jiang 13, L.-X. Zhang 14,
L.-J. Huo 15, S.-Y. Chen 16, G.-X. Zhang 17, K.-C. Wu 18, X. Zhu 1, Y.-X. Chen 1, Y. Zhu 1,
X. Shu 1, Y. Xie 1, *, N.-H. Lu 1, *
1)
Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China
2)
Department of Gastroenterology, The First Affiliated Hospital of Sun Yat-sen University, Guangdong Province, China
3)
Department of Gastroenterology, Nanjing First Hospital, Jiangsu, China
4)
Department of Gastroenterology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
5)
Department of Gastroenterology, The First Affiliated Hospital of Anhui Medical University, Anhui, China
6)
Department of Gastroenterology, Changhai Hospital, Second Military Medical University, Shanghai, China
7)
Department of Gastroenterology, Shengjing Hospital Affiliated to China Medical University, Liaoning Province, China
8)
Department of Gastroenterology, China-Japan Union Hospital of Jilin University, Changchun, Jilin, China
9)
Department of Gastroenterology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, HuBei Province, China
10)
Department of Gastroenterology, Nanfang Hospital, Southern Medical University, Guangzhou, China
11)
Department of Gastroenterology, Daping Hospital and the Research Institute of Surgery of the Third Military Medical University, Chongqing, China
12)
Department of Gastroenterology, Peking University First Hospital, Beijing, China
13)
Department of Gastroenterology, General Hospital of Chengdu Military Region, Sichuan Province, China
14)
Department of Gastroenterology, Xi'an Central Hospital, Shaanxi Province, China
15)
Department of Gastroenterology, First Clinical Medical College, Shanxi Medical University, Shanxi Province, China
16)
Department of Gastroenterology, Zhongshan Hospital, Fudan University, Shanghai, China
17)
Department of Gastroenterology, First Affiliated Hospital of Nanjing Medical University, Nanjing, China
18)
Xijing Hospital of the Fourth Military Medical University, Xi'an, China

a r t i c l e i n f o a b s t r a c t

Article history:
Objectives: To explore the characteristics of Helicobacter pylori resistance in China and the association
Received 14 August 2017
between antibiotic resistance and several clinical factors.
Received in revised form
6 November 2017 Methods: H. pylori strains were collected from patients in 13 provinces or cities in China between 2010
Accepted 7 November 2017 and 2016. Demographic data including type of disease, geographic area, age, gender and isolation year
Available online 11 November 2017 were collected to analyse their association with antibiotic resistance. Antibiotic resistance was detected
using the Etest test and the Kirby-Bauer disc diffusion method.
Editor: M. Paul Results: H. pylori were successfully cultured from 1117 patients. The prevalence of metronidazole, clar-
ithromycin (CLA), azithromycin, levofloxacin (LEV), moxifloxacin, amoxicillin (AMO), tetracycline and
Keywords: rifampicin resistance was 78.2, 22.1, 23.3, 19.2, 17.2, 3.4, 1.9 and 1.5%, respectively. No resistance to
Antibiotic resistance furazolidone was observed. The resistance rates to LEV and moxifloxacin were higher in strains isolated
China
from patients with gastritis compared to those with duodenal ulcer and among women. Compared to
Helicobacter pylori
patients 40 years old, younger patients exhibited lower resistance rates to CLA, azithromycin, LEV and
Clinical factors
Multiregion moxifloxacin. The resistance rates to CLA and AMO were higher in strains isolated more recently, and we
also found that the prevalence of resistance to metronidazole, CLA, azithromycin and AMO were
significantly different among different regions of China.

* Corresponding authors.
E-mail addresses: xieyong_tfahoncu@163.com (Y. Xie), lunonghua@ncu.edu.cn (N.-H. Lu).
y
The first two authors contributed equally to this article, and both should be considered first author.

https://doi.org/10.1016/j.cmi.2017.11.010
1198-743X/© 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.
 D.-S. Liu et al. / Clinical Microbiology and Infection 24 (2018) 780.e5e780.e8
Conclusions: The resistance rates to metronidazole, CLA and LEV were high in China. Patient age, gender,
disease and location were associated with the resistance of H. pylori to some antibiotics. Furazolidone,
AMO and tetracycline are better choices for H. pylori treatment in China. D.-S. Liu, Clin Microbiol Infect
2018;24:780.e5e780.e8
© 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All
Introduction
Helicobacter pylori plays an important role in some gastroin-
testinal diseases, such as peptic ulcers, chronic gastritis and
gastric adenocarcinoma [1,2]. To achieve a higher eradication
rate, an empirical therapy recommended as primary treatment
must be established on the basis of local antibiotic resistance
characteristics [3].
China has a large H. pylorieinfected population that exhibits
complicated antibiotic resistance features [4]. Several studies have
reported antibiotic resistance in some regions of China. For
instance, Song et al. [5] examined the resistance patterns of H. pylori
isolated from patients in four cities and found high clarithromycin
(CLA), levofloxacin (LEV) and rifampicin (RIF) resistance between
2008 and 2012, and the resistance rate to metronidazole (MET) in
the southeast coastal province of China was 95.4% [6]. As a result of
the prevalence of high resistance rates for MET and CLA in most
area of China, bismuth-containing quadruple therapy was the rec-
ommended first-line treatment for H. pylori eradication in the latest
consensus report of H. pylori infection in China, and seven antibiotic
combinations used in bismuth-containing quadruple therapy are
also suggested in the consensus [7].
Because dynamic monitoring of resistance rates is important for
eradication treatment, we performed a survey of H. pylori antibiotic
resistance from 2010 to 2016 in 13 provinces or cities of China and
analysed H. pylori resistance characteristics. In addition, we inves-
tigated the correlation between antibiotic resistance and several
clinical factors.
Materials and methods
Patients and H. pylori strains
H. pylori strains were collected from patients undergoing upper
gastrointestinal endoscopy in 18 hospitals between 2010 and 2016.
Adult patients were included if they had not received treatment for
H. pylori infection previously. Patients receiving antibiotics, proton-
pump inhibitors, H2 receptor blockers or bismuth salts within
4 weeks of the endoscopy were excluded. The demographic data
included the type of disease, geographic area, patient age and
gender, and the year the sample was collected. Informed consent
was obtained from all participants. The protocol was approved by
the ethics committee of the First Affiliated Hospital of Nanchang
University (IRB2011002).
For H. pylori culture, biopsy samples were homogenized and
cultured on Campylobacter agar (Oxoid, Basingstoke, UK) supple-
mented with 5% sheep's blood (Bio-kont, Zhejiang, China) con-
taining vancomycin, trimethoprim, polymyxin B and amphotericin
B (Duly Biotech, Nanjing, China). The plates were incubated at 37 C
under microaerophilic conditions (10% CO2, 5% O2 and 85% N2) in a
humid atmosphere for 3 to 5 days.
780.e6
rights reserved.
Antibiotic susceptibility test
An Etest (bioMe rieux, Marcy l’Etoile, France) was used to
determine the minimum inhibitory concentrations (MIC) of
amoxicillin (AMO), MET, CLA, LEV, azithromycin (AZI), moxifloxacin
(MOX), RIF and tetracycline (TET). The Kirby-Bauer disc diffusion
method (Oxoid) was used to determine the inhibition zone for
furazolidone (FUR). A strain was considered resistant if the MIC
>1 mg/mL for AMO, 1 mg/mL for CLA and AZI, >2 mg/mL for TET,
>4 mg/mL for MET (http://www.biomerieux-usa.com/sites/
subsidiary_us/files/supplementary_inserts_-_16273_-_b_-_en_-_
eag_-_etest_application_guide-3.pdf), MIC >1 mg/mL for LEV, MOX
and RIF (http://www.eucast.org/fileadmin/src/media/PDFs/
EUCAST_files/Breakpoint_tables/Breakpoint_table_v_3.1.pdf) and
if the inhibition zone was <7 mm for FUR [8]. H. pylori strain ATCC
43504 was included as an antibiotic susceptibility testing quality
control. All antibiotic susceptibility tests were conducted at the
Institute of Gastroenterology and Hepatology, First Affiliated Hos-
pital of Nanchang University.
Statistical analysis
Data analysis was performed by SPSS 17.0 (IBM SPSS, Chicago, IL,
USA). Frequencies and percentages were used to describe the
antibiotic resistance rates of H. pylori isolates. The univariate as-
sociation between each factor was quantified by Fisher's exact test
and the chi-square test. A probability value of <0.05 from a two-
tailed test was considered statistically significant.
Results
Overall H. pylori antibiotic resistance
Of the 1117 H. pylori strains, 960 strains completed susceptibility
testing for nine antibiotics, whereas 157 strains completed sus-
ceptibility testing for eight antibiotics (except for FUR). The prev-
alence of H. pylori resistance is shown in Table 1. The resistance
rates to MET, CLA, AZI, LEV, MOX, AMO, TET, RIF and FUR were 78.2,
22.1, 23.3, 19.2, 17.2, 3.4, 1.9, 1.5 and 0%, respectively. On the other
hand, we also compared four antibiotic combinations among MET,
CLA, LEV and seven antibiotic combinations used in bismuth-
containing quadruple therapy recommend in the fifth Chinese na-
tional consensus report on the management of H. pylori infection. In
general, the antibiotic combinations consisting of MET, CLA and LEV
had higher combined resistance rate, and the antibiotic combina-
tions recommended in bismuth-containing quadruple therapy had
lower combined resistance rates. Results are depicted in Fig. 1.
Factors associated with antibiotic resistance
The analysis revealed that compared to men, women had a
significantly higher resistance rate to LEV (p 0.016) and MOX
780.e7 D.-S. Liu et al. / Clinical Microbiology and Infection 24 (2018) 780.e5e780.e8

Table 1
Factors associated with Helicobacter pylori resistance

Characteristic N MET CLA AZI LEV MOX AMO TCT RIF FUR

Age * * * *
<40 years 522 397 (76.1) 97 (18.6) 105 (20.1) 78 (14.9) 80 (15.3) 19 (3.6) 8 (1.5) 5 (1.0) 0
40 years 595 477 (80.2) 150 (25.2) 155 (26.1) 136 (22.9) 120 (20.2) 19 (3.2) 13 (2.2) 12 (2.0) 0
Gender * *
Male 676 517 (76.5) 148 (21.9) 154 (22.8) 114 (16.9) 97 (14.3) 21 (3.1) 12 (1.8) 12 (1.8) 0
Female 441 357 (81.0) 99 (22.4) 106 (24.0) 100 (22.7) 103 (23.4) 17 (3.9) 9 (2.0) 5 (1.1) 0
Year of isolation * *
2010e2012 355 276 (77.7) 65 (18.3) 70 (19.7) 72 (20.3) 73 (20.6) 4 (1.1) 8 (2.3) 7 (2.0) 0
2013e2016 762 598 (78.5) 182 (23.9) 190 (24.9) 142 (18.6) 127 (16.7) 34 (4.5) 13 (1.7) 10 (1.3) 0
Disease * *
DU 1093 855 (78.2) 240 (22.0) 251 (23.0) 204 (18.7) 191 (17.5) 38 (3.5) 19 (1.7) 17 (1.6) 0
Gastritis 24 19 (79.2) 7 (29.2) 9 (37.5) 10 (41.7) 9 (37.5) 0 2 (8.3) 0 0
Area * * * *
Northeast 66 59 (89.4) 30 (45.5) 34 (51.5) 14 (21.2) 14 (21.2) 6 (9.1) 2 (3.0) 0 0
East 312 233 (74.7) 75 (24.0) 71 (24.4) 63 (20.2) 48 (15.4) 14 (4.5) 3 (1.0) 2 (0.6) 0
Central 683 535 (78.3) 127 (18.6) 138 (19.5) 121 (17.7) 122 (17.9) 17 (2.5) 14 (2.0) 13 (1.9) 0
West 56 47 (83.9) 15 (26.8) 17 (30.4) 13 (23.2) 16 (28.6) 1 (1.8) 2 (3.6) 2 (3.6) 0
Overall 1117 874 (78.2) 247 (22.1) 260 (23.3) 214 (19.2) 200 (17.2) 38 (3.4) 21 (1.9) 17 (1.5) 0

Data are presented as n (%).

AMO, amoxicillin; AZI, azithromycin; CLA, clarithromycin; DU, duodenal ulcer; FUR, furazolidone; LEV, levofloxacin; MET, metronidazole; MOX, moxifloxacin; RIF, rifampicin;
TCT, tetracycline.
*p < 0.05.

(p < 0.001). Compared to adult patients (40 years old), younger
patients (<40 years old) showed lower resistance rates to CLA (p
0.008), AZI (p 0.019), LEV (p 0.001) and MOX (p 0.042). The resis-
tance rate for CLA (p 0.037) and AMO (p 0.014) were higher in
strains isolated in 2013e2016 than in those isolated in 2010e2012.
Resistance rates to LEV (p 0.009) and MOX (p 0.017) were higher in
strains isolated from gastritis patients than duodenal ulcer patients.
Furthermore, we also observed the prevalence of MET (p 0.041),
CLA (p < 0.001), AZI (p < 0.001) and AMO (p 0.025) resistance were
significantly different in different regions of China, and resistance
rates were high in the northeast area.
Discussion
This study included a large number of strains from 13 provinces
or cities in China collected between 2010 and 2016. Overall, the
results indicate that high rates of primary resistance to antibiotics
are still a challenge to the eradication of H. pylori in China. The
prevalence of MET and CLA resistance in China is significantly
higher than in previous decades [9], which may also induce low
eradication rates for sequential and concomitant therapies [3]. The
prevalence of AMO and CLA resistance has increased recently,
which suggests that the dynamic monitoring of AMO and CLA
susceptibility is essential. Our results also suggest that younger
patients have a lower resistance rate to CLA, AZI, LEV and MOX than
patients over 40, a phenomenon that may be related to the accu-
mulation of drug use with age as well as the fact that elderly pa-
tients are more likely experience respiratory tract and urinary
infections. In addition, the resistance rates to MET, CLA, AZI and
AMO were significantly different in different areas, indicating that
the use of these antibiotics should consider local antibiotic
resistance.
In China, antieH. pylori drugs such as bismuth, TET and FUR are
available. Therefore, quadruple therapy with proton pump in-
hibitors, bismuth and a combination of two antibioticsdspecifi-
cally FUR, TET or AMOdwould be more suitable for Chinese
patients. This treatment is also recommended in the Maastricht V/
Florence Consensus Report [3].

Conclusions

The resistance rates to MET, CLA and LEV were high in China.
Patient age, gender, disease and geographical location were asso-
ciated with the resistance of H. pylori to some antibiotics. Consid-
ering the antibiotic resistance profiles, FUR, AMO and TET are better
choices for H. pylori treatment.

Transparency Declaration

Funding was provided by grants from National Science and

Fig. 1. Characteristics of antibiotic resistance for Helicobacter pylori in China. M þ C,
metronidazole þ clarithromycin; M þ L, metronidazole þ levofloxacin; C þ L,
clarithromycin þ levofloxacin; A þ M, amoxicillin þ metronidazole; A þ C,
amoxicillin þ clarithromycin; A þ L, amoxicillin þ levofloxacin; A þ T,
amoxicillin þ tetracycline; A þ F, amoxicillin þ furazolidone; T þ M,
tetracycline þ metronidazole; T þ F, tetracycline þ furazolidone; M þ C þ Q,
metronidazole þ clarithromycin þ levofloxacin.
Technology Major Projects for ‘Major New Drugs Innovation and
Development’ of China (2011ZX09302-007-03), National Key
Research and Development Grogram of China (2016YFC1302201);
and the National Natural Science Foundation of China (81460115),
Science and Technology Projects of Jiangxi province
(2014BBG70019). All authors report no conflicts of interest rele-
vant to this article.
 D.-S. Liu et al. / Clinical Microbiology and Infection 24 (2018) 780.e5e780.e8 780.e8

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