|Year : 2014 | Volume
| Issue : 3 | Page : 602-605
Prevalence of extended-spectrum β-lactamase-producing Enterobacteriaceae in Menoufia University Hospitals
Amira A El-Hendi1, Ayman M El-Lehleh2, Abeer H El-Shalakany1, Sheriene M Moussa MBBCh 1
1 Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Menoufia, Egypt
2 Department of Tropical Medicine, Faculty of Medicine, Menoufia University, Menoufia, Egypt
|Date of Submission||01-Sep-2013|
|Date of Acceptance||30-Nov-2013|
|Date of Web Publication||26-Nov-2014|
Sheriene M Moussa
Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Yassin Abdel-Ghaffar Street, 4, Shebin El-Koom, 32511 Menoufia
Source of Support: None, Conflict of Interest: None
The aim of the study was to evaluate the prevalence of extended-spectrum β-lactamase (ESBLs) in Enterobacteriaceae among patients conducted at Menoufia University Hospitals.
Resistance to β-lactam antibiotics is an increasing problem, and β-lactamase production is the most common mechanism of drug resistance, especially in Enterobacteriaceae. The Clinical and Laboratory Standard Institute (CLSI) interpretive guidelines stated that Enterobacteriaceae that produce ESBLs are resistant to therapy with penicillin, cephalosporins, and aztreonam, despite apparent in-vitro susceptibility to some of these agents. Therefore, detection of ESBLs in Enterobacteriaceae is crucial for optimal treatment of patients and to control the spread of resistance.
Patients and methods
The isolated strains of Enterobacteriaceae are subjected to phenotype detection by screening for ESBLs by disk diffusion test then to confirmatory tests for ESBLs by double disk test.
A total of 97 of 160 (60.6%) isolates were found to be ESBL producers by the CLSI confirmatory method. Of 97 ESBL producers, 45 (46.4%) were Escherichia coli, 28 (28.9%) were Enterobacter spp., and 22 (22.7%) were Klebsiella spp. Of the various clinical samples, the most frequent ESBLs isolates were from urine samples (27.8%), followed by sputum samples (18.6%).
High prevalence of ESBL producers in our hospital (60.6%) calls for strict policies regarding antibiotic usage and their screening methods, and hospital-based clinical laboratories should screen isolates following hospitalization in patients in need for antibiotics to formulate effective antibiotic strategy and plan a proper hospital infection control strategy to prevent the spread of these ESBL strains.
Keywords: drug resistance, Enterobacteriaceae, extended-spectrum β-lactamase, prevalence
|How to cite this article:|
El-Hendi AA, El-Lehleh AM, El-Shalakany AH, Moussa SM. Prevalence of extended-spectrum β-lactamase-producing Enterobacteriaceae in Menoufia University Hospitals. Menoufia Med J 2014;27:602-5
|How to cite this URL:|
El-Hendi AA, El-Lehleh AM, El-Shalakany AH, Moussa SM. Prevalence of extended-spectrum β-lactamase-producing Enterobacteriaceae in Menoufia University Hospitals. Menoufia Med J [serial online] 2014 [cited 2020 Feb 24];27:602-5. Available from: http://www.mmj.eg.net/text.asp?2014/27/3/602/145525
| Introduction|| |
Resistance to β-lactam antibiotics is an increasing problem, and β-lactamase production is the most common mechanism of drug resistance, especially in Enterobacteriaceae  .
Extended-spectrum β-lactamase (ESBL)-producing strains are a major problem in many hospitals leading to increased morbidity, mortality, and healthcare costs. Cephalosporins are first-line drugs in treatment of infections caused by Enterobacteriaceae family; however, extensive use of third-generation cephalosporin has resulted in increased prevalence of ESBL among organisms  .
The most important mechanism of resistance to β-lactam antibiotics is the production of β-lactamase enzymes. The β-lactamases confer significant antibiotic resistance to their bacterial hosts by hydrolysis of the amide bond of the β-lactam ring. Classes A, C, and D include enzymes that hydrolyze their substrates by forming an acyl (penicilloic or cephalosporoic) enzyme through an active-site serine, whereas class B β-lactamases are metalloenzymes that utilize at least one active-site zinc ion. These enzymes are especially important in Gram-negative bacteria, as they constitute the major defense mechanism against β-lactam-based drugs  .
β-Lactamases are remarkably diversified due to their continuous mutation  .
The Clinical and Laboratory Standard Institute (CLSI) interpretive guidelines stated that Enterobacteriaceae that produce ESBLs are resistant to therapy with penicillin, cephalosporins, and aztreonam, despite apparent in-vitro susceptibility to some of these agents  . Third-generation cephalosporin resistance is often mediated by TEM-type and SHV-type l-lactamases in Enterobacteriaceae .
The aim of the study was to evaluate the prevalence of ESBLs in Enterobacteriaceae among patients conducted at Menoufia University Hospitals.
| Patients and methods|| |
Collection of samples
A total of 160 Enterobacteriaceae isolates from urine, sputum, stool, blood, pus, drains, and other samples were collected from patients who referred to Menoufia University Hospitals, a referral center offering tertiary care to about one-third of population inhabiting the Nile Delta at North of Egypt.
Isolation and identification
Colony isolates were obtained after culture of urine specimens on cysteine lactose electrolyte deficient, whereas pus and sputum specimens were cultured on blood agar and MacConkey's agar; thereafter, all specimens were incubated aerobically at 37°C for 24 h. Blood samples were inoculated into blood culture bottles, then subcultures were performed also on blood and MacConkey's agar. A single isolated colony was considered for further studies and identification was made using standard conventional, morphological, cultural, and biochemical tests  .
Detection of extended-spectrum β-lactamase
Screening for extended-spectrum 0β-lactamase producers
It was performed according to CLSI 2011 recommendations to identify potential ESBL-producing isolates using standard disk diffusion test.
Ceftazidime (30 μg), cefotaxime (30 μg), and ceftriaxone (30 μg) are used. If a zone diameter of less than 22 mm for ceftazidime, less than 27 mm for cefotaxime, and less than 25 mm for ceftriaxone was recorded, the strain was considered to be 'suspicious for ESBL production'. The use of more than one antimicrobial agent for screening improves the sensitivity of detection.
Phenotypic confirmatory test for extended-spectrum β-lactamase production by cephalosporin/clavulanate combination disks
All strains that were screened out for ESBL production were also subjected to confirmation using the PCDDT, as recommended by the CLSI 2011. The ceftazidime (30 μg) and cefotaxime (30 μg) disks alone and in combination with clavulanic acid (ceftazidime+clavulanic acid, 30/10 μg disks, cefotaxime+clavulanic acid, 30/10 μg) were applied onto a plate of Mueller-Hinton agar, which was inoculated with the test strain.
An increase of at least 5 mm in zone diameter for either antimicrobial agent tested in combination with clavulanic acid versus its zone when tested alone denoted ESBL-producing strains.
Reference strains used in this study were Escherichia coli ATCC 25922 as negative control and Klebsiella pneumoniae ATCC 700603 as positive control.
| Results|| |
The age of the studied patients ranged from 10 to 75 years with a mean ± SD of 55.72 ± 8.94 years and a median of 45 years.
A total of 160 clinical samples were cultured from various sample sites from patients admitted at Menoufia University Hospitals. The distribution of isolates was 46.8% E. coli, 30.0% Enterobacter spp., 21.8% Klebsiella spp., and 1.3% Serretia spp.
Most patients presented with fever (73%) as the chief complaint; others presented with persistent dysuria (15%), persistent wheezy chest (10%), and wound infection (2%).
Clinically, eight patients had ascites, 10 had jaundice, eight had shrunken cirrhotic liver, eight had splenomegaly, four had encephalopathy, 41 had pallor, 15 had abdominal pain, 20 had wheezy chest, three had clinically infected wounds and CVP lines, 18 had suprapubic pain, and nine had recurrent loin pain.
Overall, 97 (60.6%) isolates were detected as ESBL producers by phenotypic confirmatory test for ESBL production (cephalosporin/clavulanate combination disks) and included 45 (46.4%) E. coli, 28 (28.9%) Enterobacter spp., 22 (22.7%) Klebsiella spp., and two (2.1%) Serretia spp.
Among 60.6% ESBL isolates, there were 45.6% ESBL isolates from patients with previous antimicrobial intake, which were more than isolates from patients without previous antimicrobial intake 15.0%; ESBLs isolates are higher in male individuals (35.6%) than in female individuals (25.0%).
The most frequent ESBLs isolates were from urine samples (27.8%), followed by sputum samples (18.6%), stool (12.4%), and pus (9.3%), whereas the least frequent ESBLs isolates were from nasal swabs and CVP line samples.
| Discussion|| |
There is a problem that occurred among ESBL-producing Enterobacteriaceae, which have widely caused the spread of infections worldwide ,, . Although hundred variants of ESBLs have been described today , , the most common of them are derivatives of TEM or SHV enzymes. In addition, in recent years non-TEM and non-SHV ESBLs have been reported, mainly the CTX-M enzymes [8, 12, 13].
ESBL-producing strains are a major problem in many hospitals, leading to increased morbidity, mortality, and healthcare costs. Therefore, detection of ESBLs in Enterobacteriaceae is crucial for optimal treatment of patients and to control the spread of resistance  .
The uncontrolled use of antibiotics at hospital could be a leading contributory factor to the high ESBL prevalence because third-generation cephalosporins are usually first line against many severely infectious diseases. Justifiable use will be an effective means of controlling and decreasing spread of ESBLs strains. This study showed that the highest rate of ESBL-producing Enterobacteriaceae was found in patients with history of antibiotic administration (45.6%) than in other patients without antibiotic administration (15.0%). Similar results were reported by Graffunder et al.  in the USA who found a correlation between the selective pressures of antimicrobial agents identified as risk factors for ESBL-producing organisms and the presence of related resistance genes residing on the plasmid.
Suggested approaches to the management of patients with serious infections due to ESBL-producing Enterobacteriaceae are as follows:
(1) First-line therapy:
(b) Piperacillin-tazobactam (low inoculum).
(c) Fosfomycin (oral formulation for simple urinary tract infections).
(2) Second-line therapy:
(a) Tigecycline (not in urinary tract infections).
(c) Colistin 15.
In the present study, the prevalence of ESBL-producing Enterobacteriaceae isolates was 60.6%. Nearly same results were obtained by Olowe et al.  from Nigeria, who showed that the prevalence of ESBL-producing Enterobacteriaceae isolates was about 51.3%, and also by Dallal et al.  from Iran who found that 57.5% of isolates were ESBL-producing Enterobacteriaceae.
In several western studies, prevalence of ESBLs was less than that found in our study. The uncontrolled use of third-generation cephalosporins at our hospital could be a leading contributory factor to the high ESBL prevalence observed in this study.
In the present study, the maximum ESBL production was seen among the isolates of E. coli (46.4%), followed by Enterobacter spp. (28.9%), Klebsiella spp. (22.7%), and Serretia spp. (2.1%). Nearly similar results were observed by Balan  from India who reported that ESBLs were detected among the following isolates: 24 (52.2%) E. coli, 15 (32.6%) K. pneumoniae, three (6.5%) Proteus mirabilis, three (6.5%) Enterobacter spp., and one (2.2%) Citrobacter spp. Another study in Chennai reported that ESBL production was 47% among E. coli and 37% among K. pneumoniae in a tertiary care center  .
However, these results disagree with that of the study of Kaur and Aggarwal  , which showed that the maximum ESBL production was found among the isolates of K. pneumoniae (52.27%), followed by those of E. coli (46.43%) ([Figure 1] and [Table 1], [Table 2] and [Table 3]).
|Figure 1: Confirmatory test for detection of ESBL: >5 mm increase in zone diameter for cefotaxime and ceftazidime when tested in combination with clavulanic acid versus its zone when tested alone. C, clavulanic acid; CAZ, ceftazidime; CTX, cefotaxime; ESBL, extended-spectrum b-lactamase.|
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|Table 1: Distribution of Enterobacteriaceae isolates according to extended-spectrum b -lactamases production |
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|Table 2: Distribution of extended-spectrum b -lactamases-producing Enterobacteriaceae isolates according to the type of sample |
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|Table 3: Comparison of Enterobacteriaceae isolates according to extended-spectrum b -lactamase production and history of antimicrobial intake |
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| Conclusion|| |
There is high prevalence of ESBL producers among Enterobacteriaceae isolates in our hospital (60.6%). This calls for strict policies regarding antibiotic usage and their screening methods.
It is important for the clinical microbiology laboratory to implement one or more methods to detect ESBLs, and hospital-based clinical laboratories should screen isolates from the community-based patients before hospitalization to prevent the spread of ESBL-mediated resistance.
The use of cephalosporins should be discouraged for empirical therapy of infections and should only be used according to results of culture and sensitivity.
| Acknowledgements|| |
Conflicts of interest
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[Table 1], [Table 2], [Table 3]