Original Article

Prevalence and antibiotic resistance pattern of

extended‑spectrum beta‑lactamase‑producing
Escherichia coli in clinical specimens
Kiana Shirani1, Elahe Seydayi1, Kiarash Salimi Boroujeni2
1
Isfahan Infectious Diseases Research Center, Isfahan University of Medical Sciences, Isfahan, Iran, 2Faculty of Medicine, Isfahan University
of Medical Sciences, Isfahan, Iran

Background: Extended‑spectrum ß‑lactamase (ESBL)‑producing Enterobacteriaceae seem to have an extended antibiotic resistance, but
have different resistance patterns throughout different sites and regions. This study aimed to evaluate the antibiotic resistance pattern
of ESBL‑producing Escherichia coli. Materials and Methods: One hundred swab samples from patients hospitalized due to a clinical
suspicion of any kind of infection (with manifestations such as fever, leukocytosis, and an active urinalysis result) were processed in Alzahra
Microbiology Laboratory, Isfahan, Iran. Isolated E. coli were cultured on Mueller–Hinton agar and antibiotic susceptibility was tested by
Kirby–Bauer disk diffusion method following the Clinical and Laboratory Standard Institute 2017 guidelines. Results: ESBL‑producing
samples had higher antibiotic resistance rates than ESBL‑non‑producing samples: ceftriaxone (58.8% vs. 27.3%), cefotaxime (73.5% vs.
30.3%), ceftizoxime (76.5% vs. 33.3%), cefixime (79.4% vs. 40.9%), and cefpodoxime (73.5% vs. 53%), except for carbenicillin (29.4% vs. 48.5%).
Imipenem and meropenem were the least resisted antibiotics in ESBL‑producing samples (5.9% and 11.8%). Conclusion: ESBL‑producing
Enterobacteriaceae have a high resistance rate to third‑generation cephalosporins and high susceptibility to imipenem and meropenem.

Key words: Bacterial, beta‑lactamases, drug resistance, Escherichia coli

How to cite this article: Shirani K, Seydayi E, Boroujeni KS. Prevalence and antibiotic resistance pattern of extended‑spectrum beta‑lactamase‑producing
Escherichia coli in clinical specimens. J Res Med Sci 2019;24:103.

INTRODUCTION an ascending trend of growth in both community and

hospital infections in Iran.[6‑8]
One of the most important mechanisms of
bacteria against antibiotics is the production of Sufficient identification of ESBL‑producing strains
enzymes destroying β‑lactam ring in the antibiotics is essential to make an appropriate choice of
structure. Extended‑spectrum ß‑lactamase (ESBL) antimicrobial regimen and evaluation strategy. [9]
is an important group of β‑lactamases.[1] Escherichia Because no comprehensive studies in the territory of
coli is the most prevalent and hence the most ESBL‑producing E. coli in Iran are available, we aimed to
important multidrug‑resistant Gram‑negative evaluate the prevalence and antibiotic resistance pattern
infection, especially in patients with urinary tract of ESBL‑producing E. coli in clinical specimens.
infection (UTI). [2,3] Throughout the recent century,
ESBL‑producing Enterobacteriaceae have been introduced MATERIALS AND METHODS
in the literature.[4] ESBL‑producing E. coli has been
isolated in community and nosocomial settings as Study design and target group
well.[5] This might be a result of extensive antibiotic Throughout a cross‑sectional study, we evaluated
usage and can cause antibiotic resistance in human clinical specimens from hospitalized patients in
pathogens. Infection with ESBL‑producing E. coli has Isfahan Alzahra Hospital, Center of Iran, from August
to December 2015. Four milliliters of midstream
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10.4103/jrms.JRMS_634_18
For reprints contact: [email protected]

Address for correspondence: Mr. Kiarash Salimi Boroujeni, No 12, Sajjadieh 3, Artesh Street, Isfahan, Iran. E‑mail: [email protected]
Received: 02‑11‑2018; Revised: 26‑12‑2018; Accepted: 24-09-2019

1 © 2019 Journal of Research in Medical Sciences | Published by Wolters Kluwer - Medknow | 2019 |
 Shirani, et al.: Antibiotic resistance pattern in E. coli with ESBL

urine was collected from each patient into a sterile tube. acid (30 μg + 10 μg), cefotaxime (30 μg), ceftazidime–clavulanic
Samples were then transported to the hospital laboratory acid (30 μg + 10 μg), and ceftazidime (30 μg) combination
as soon as possible. Patients were instructed properly for disks. The tests were interpreted according to the most
the means of sampling.[10,11] Sources of the samples varied recent CLSI guidelines (2017), and a difference of 5 mm
throughout the patients, in accordance with their symptoms, between IZ of a single disk and in combination with
practitioners’ clinical suspicion, and the standard diagnostic clavulanic acid (inhibitor) was confirmed to be produced
guidelines (with blood [24%], urine [44%], abscess [3%], by an ESBL‑positive isolate.
CSF [4%], sputum [5%], rectal swab [3%], perianal
swab [3%], and skin swab [13%], differing based on the Data analysis
patients’ manifestations). Statistical analysis of data was performed using SPSS 22.0
software. To compare qualitative variables between groups,
Laboratory assessment and extended‑spectrum Chi‑square test was performed. The normal distribution
ß‑lactamase detection of all studied parameters was checked with Kolmogorov–
Two hours after the collection, 100 swab samples, isolated Smirnov test. Student’s t‑test was used for variables which
from urine specimen of patients hospitalized due to were distributed in a normal way, besides Mann–Whitney
various reasons with a clinical suspicion of any kind of and Wilcoxon tests were performed for variables that have
infection (fever, leucocytosis), were streaked directly not normal distribution. Two‑tailed P < 0.05 was considered
on eosin methylene blue agar, MacConkey agar, and statistically significant.
blood agar plates. Such plates were incubated at 37°C
aerobically, and after overnight incubation, they were RESULTS
assessed for E. coli growth. E. coli existence was proved by
their colony morphology, Gram staining characteristics, The results of the study showed that ESBL‑producing
biochemical tests of glucose fermentation, Voges–Proskauer E. coli was found in 34% of all samples (ergo 34 ESBL
reaction (acetyl methyl carbinol production from dextrose) screening‑positive samples). ESBL‑producing samples
on the Triple Sugar Iron agar, gas producing, lactose had higher antibiotic resistance rate to third‑generation
metabolism, production of indole from tryptophan, cephalosporins than ESBL‑non‑producing samples such as
sulfide‑indole‑motility, and methyl red Voges–Proskauer. ceftriaxone (58.8% vs. 27.3%, P < 0.001), cefotaxime (73.5%
vs. 30.3%, P < 0.001), ceftizoxime (76.5% vs. 33.3%, P < 0.001),
Isolated E. coli were cultured on Mueller–Hinton agar (MHA), cefixime (79.4% vs. 40.9%, P < 0.001), and cefpodoxime (73.5%
and antibiotic susceptibility was tested by Kirby–Bauer disk vs. 53%, P = 0.045). On the other hand, carbenicillin in
diffusion method after the Clinical and Laboratory Standard ESBL‑producing samples had lower antibiotic resistance
Institute (CLSI) guidelines, 2017.[12] Below is the list of drug rate than ESBL‑non‑producing samples (29.4% vs. 48.5%,
P = 0.031), which is a rather strange finding. Furthermore,
concentrations used for disc diffusion testing: ceftazidime (30
we found that imipenem and meropenem had the lowest
μg; inhibition zone (IZ) size equal or smaller than 22 mm);
antibiotic resistance rate in ESBL‑producing samples (5.9%
amikacin (30 μg), ampicillin (10 μg), piperacillin (100 μg),
and 11.8%) [Tables 1 and 2].
cefixime (5 μg), cefotaxime (30 μg; IZ ≤27 mm), amoxicillin/
clavulanic acid (30 µg), ceftriaxone (30 μg; IZ ≤ 25 mm),
Table 1: Demographic characteristics of patients and
ciprofloxacin (5 μg), cotrimoxazole (23.75 μg studied variables on account of extended‑spectrum
sulfamethoxazole/1.25 μg trimethoprim), ceftizoxime (30 µg), ß‑lactamase production
imipenem (10 μg), meropenem (30 µg), nalidixic acid (30 Variables ESBL P
μg), gentamicin (10 µg), carbenicillin (100 µg), and Positive (%) Negative (%)
cefpodoxime (30 μg; IZ ≤ 17 mm). Age (years) 46.35±12.97 45.93±11.8 0.873
Sex
Isolates showing IZs less than the values stated above were Male (53) 15 (44.1) 38 (57.6) 0.201
interpreted as screening positive for ESBL production. Only Female (47) 19 (55.9) 28 (42.4)
E. coli were screened for ESBL production. Clinic sample
Blood (24) 7 (20.6) 17 (25.8) 0.176
Urine (44) 14 (41.2) 31 (47)
For ESBL confirmation, 2–3 colonies of the organisms
Abscess (3) 0 3 (4.5)
were suspended in 0.5 ml of sterile broth and the
CSF (4) 2 (5.9) 2 (3)
turbidity matched to 0.5 McFarland. Using a sterile
Sputum (5) 1 (2.9) 4 (6.1)
cotton swab, the broth culture was uniformly swabbed Rectal swab (3) 2 (5.9) 1 (1.5)
on MHA. All the E. coli isolates resistant to at least Perianal swab (3) 0 3 (4.5)
ceftazidime, ceftriaxone, and/or cefotaxime were Skin swab (13) 8 (23.5) 5 (7.6)
tested for confirmation using cefotaxime–clavulanic ESBL=Extended‑spectrum ß‑lactamase

| 2019 | Journal of Research in Medical Sciences 2

 Shirani, et al.: Antibiotic resistance pattern in E. coli with ESBL

Table 2: Antibiotic susceptibility patterns on account of extended‑spectrum ß‑lactamase production

Antibiotics ESBL P
Positive (%) Negative (%)
Sensitive Intermediate Resistance Sensitive Intermediate Resistance
Antibiotic susceptibility
Ampicillin 1 (2.9) 8 (23.5) 25 (73.5) 2 (3) 21 (31.8) 43 (65.2) 0.683
Amikacin 1 (2.9) 16 (47.1) 17 (50) 1 (1.5) 24 (36.4) 41 (62.1) 0.487
Amoxicillin/clavulanic acid 7 (20.6) 13 (38.2) 14 (41.2) 10 (15.2) 28 (42.4) 28 (42.4) 0.781
Ceftriaxone 4 (11.8) 10 (29.4) 20 (58.8) 40 (60.6) 8 (12.8) 18 (27.3) <0.001
Cefotaxime 0 9 (26.5) 25 (73.5) 24 (36.4) 22 (33.3) 20 (30.3) <0.001
Ceftizoxime 0 8 (23.5) 26 (76.5) 34 (51.5) 10 (15.2) 22 (33.3) <0.001
Cefixime 0 7 (20.6) 27 (79.4) 18 (27.3) 21 (31.8) 27 (40.9) <0.001
Carbenicillin 9 (26.5) 15 (44.1) 10 (29.4) 21 (38.1) 13 (19.7) 32 (48.5) 0.031
Ciprofloxacin 2 (5.9) 10 (29.4) 22 (64.7) 7 (10.6) 29 (43.9) 30 (45.5) 0.185
Cefpodoxime 0 9 (26.5) 25 (73.5) 8 (12.1) 23 (34.8) 35 (53) 0.045
Trimethoprim 5 (14.7) 13 (38.2) 16 (47.1) 9 (13.6) 17 (25.8) 40 (60.6) 0.383
Imipenem 31 (91.2) 1 (2.9) 2 (5.9) 64 (97) 1 (1.5) 1 (1.5) 0.42
Meropenem 29 (85.3) 1 (2.9) 4 (11.8) 57 (86.4) 4 (6.1) 5 (7.6) 0.645
Sulfamethoxazole 9 (26.5) 13 (38.2) 12 (35.3) 21 (31.8) 17 (25.8) 28 (42.4) 0.435
Piperacillin 5 (14.7) 9 (26.5) 20 (58.8) 11 (16.7) 17 (25.8) 38 (57.6) 0.968
Nalidixic acid 15 (44.1) 6 (17.6) 13 (38.2) 26 (39.4) 14 (21.2) 26 (39.4) 0.873
Gentamicin 14 (41.2) 7 (20.6) 13 (38.2) 19 (28.8) 21 (31.8) 26 (39.4) 0.357
ESBL=Extended‑spectrum ß‑lactamase

DISCUSSION antibiogram in hospital‑admitted UTI patients and select

the best choice of antibiotics.
The present piece of research focused solely on the prevalence
and antibiotic resistance pattern of ESBL‑producing E. coli CONCLUSION
due to shortage of the project budget.
Our results showed high prevalence of ESBL in hospital
We found that the prevalence of ESBL‑producing bacteria in samples in Isfahan, Iran. Because Alzahra Hospital is a major
clinical samples of the hospital was 34 %. This is a completely and characteristic hospital laboratory dealing specifically
high amount for such a prevalent microorganism which with exceptional patients, the conduction of this study in
would be realy catastrophic in the treatment approaches. that specific laboratory setting in Isfahan should interest
This value has been reported in lower amounts in some of readers from clinical and epidemiological perspective. Our
the other studies,[13‑17] whereas other studies reported higher data confirmed that ESBL had high resistance rate to third
prevalence as compared to our results.[18,19] As reported in a generation of cephalosporins and high susceptibility to
cross‑sectional study by Mihankhah et al., E. coli is among imipenem and meropenem. These findings suggest further
the most prevalent Gram‑negative specimens obtained from studies in this field.
clinical samples of UTIs in Iran with 37.8% of the whole.[20]
Acknowledgments
ESBLs are enzymes destroying β‑lactam ring in the This article was carried out as a doctoral dissertation project
antibiotic structure, such as monobactams (e.g., aztreonam), under the supervision of Isfahan University of Medical
third‑generation cephalosporins (e.g., ceftriaxone, Sciences, with the ethical code: IR.MUI.REC.1394.3.719.
ceftazidime, and cefotaxime), and carbapenems (e.g.,
imipenem, meropenem, and ertapenem), but not the Financial support and sponsorship
Isfahan University of Medical Sciences.
cephamycins (e.g., cefoxitin and cefotetan).[21] Such enzymes
are sensitive to β‑lactamase inhibitors (clavulanic acid,
Conflicts for interest
sulbactam, and tazobactam).[22] Bacterial resistance has
There are no conflicts for interest.
increased during the recent decades.[23,24] As our statistical
data witness, although third‑generation cephalosporins
are strong and widely used antibiotics, there is a high rate
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