Prevalence of extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae in an urban hospital in Dhaka, Bangladesh

International Journal of Antimicrobial Agents 24 (2004) 508–510

Short Communication

Prevalence of extended-spectrum ␤-lactamase-producing Escherichia coli and Klebsiella pneumoniae in an urban hospital in Dhaka, Bangladesh M. Mushfequr Rahmana , J. Ashraful Haqb,∗ , M. Akram Hossainc , Rosy Sultanaa , Faridul Islama , A.H.M. Shafiqul Islamb a

Department of Immunology and Microbiology, National Healthcare Network, 69/M, Panthopath, Dhaka1205, Bangladesh b Department of Microbiology, Bangladesh Institute of Research and Rehabilitation in Diabetes, Endocrine and Metabolic Disorders (BIRDEM), 122 Kazi Nazrul Islam Avenue, Dhaka 1000, Bangladesh c Department of Microbiology, Mymensingh Medical College, Mymensingh, Bangladesh Received 25 February 2004; accepted 23 May 2004

Abstract The prevalence of extended-spectrum ␤-lactamases (ESBL)-producing organisms in an urban hospital in Dhaka City was assessed over a 10-month period. A double disk test was performed to detect ESBL-producing Escherichia coli and Klebsiella pneumoniae. 43.2% and 39.5% of E. coli and K. pneumoniae had ESBL phenotypes, respectively. The combination of augmentin with ceftazidime detected the most ESBL-producing E. coli (39.5%) while augmentin with ceftriaxone was the best combination for the detection of ESBL (31.6%) in K. pneumoniae. © 2004 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. Keywords: ESBL; Extended-spectrum ␤-lactamase; Bangladesh; Escherichia coli; Klebsiella pneumoniae

1. Introduction Bacterial antibiotic resistance has become a major clinical concern worldwide including Bangladesh [1–3]. Recently, the use of second and third generation cephalosporins has led to the selection of Gram-negative organisms resistant to ␤-lactamase stable cephalosporins. This resistance is attributed to the production of extended-spectrum ␤-lactamases (ESBL) [4–8]. ESBL-producing organisms have been reported from many parts of the world [5–7] but so far no study has determined the prevalence of ESBL-producing bacteria in Bangladesh.

∗ Corresponding author. Tel.: + 880 2 861 6641-50x2248/2271; fax: +880 2 861 3004. E-mail address: [email protected] (J.A. Haq).

This study was designed to detect the prevalence of ESBL-producing Escherichia coli and Klebsiella pneumoniae in clinical specimens from an urban hospital in Dhaka city.

2. Materials and methods 2.1. Study population and types of specimen All the samples were collected from hospitalized patients of a 550-bed referral hospital of Dhaka City, Bangladesh. Each infected anatomical site was considered to be a separate infection regardless of whether the pathogen was the same or different and each patient was sampled once. Samples were collected over a 10-month period from both sexes and different age groups. The specimen types included in this study

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M.M. Rahman et al. / International Journal of Antimicrobial Agents 24 (2004) 508–510

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Table 1 Antibiogram of isolated E. coli and K. pneumoniae by a disk diffusion method Organisms

% Sensitive to

E. coli (n = 131) K.pneumoniae (n = 108)

Amp

Cepl

CXM

CAZ

CRO

CTX

Imp

Cip

Cot

Gen

Net

7.6 5.5

21.3 17.5

37.4 35.1

56.4 55.5

58.0 51.8

61.1 57.4

98.4 94.4

40.4 36.1

23.6 30.5

67.9 52.8

59.5 62.0

Note: Amp: ampicillin, Cepl: cephalexin, CXM: cefuroxime, CAZ: ceftazidime, CRO: ceftriaxone, CTX: cefotaxime, Imp: imipenem, Cip: ciprofloxacillin, Cot: cotrimoxazole, Gen: gentamicin and Net: netilmicin.

Table 2 Rate of isolation of ESBL-producing organisms by double disk diffusion technique using augmentin disk in combination with ceftriaxone, ceftazidime, cefotaxime or aztreonam disk Organism

E. coli (n = 81) K. pneumoniae (n = 76)

Total ESBL positive∗

ESBL positive by

No. (%)

CRO + Aug Pos

%

Pos

%

Pos

%

Pos

%

35 (43.2) 30 (39.5)

29 24

35.8 31.6

32 22

39.5 28.9

28 19

34.6 25.0

25 21

30.9 27.6

CAZ + Aug

CTX + Aug

ATM + Aug

Note: CRO: ceftriaxone (30 ␮g), CAZ: ceftazidime (30 ␮g), CTX: cefotaxime (30 ␮g), ATM: aztreonam (30 ␮g), Aug: augmentin (amoxycillin 20 ␮g + clavulanic acid 10 ␮g). ∗ An isolate was considered ESBL positive if there was an enhancement of zone of inhibition with any one of the four disks and the augmentin disk.

were urine, pus, sputum, blood, vaginal swab and throat swab. ESBL were only looked for in E. coli and K. pneumoniae as they were the most prevalent organisms isolated. 2.2. Microbiological methods All samples were routinely cultured on MacConkey and blood agar plates. Blood and sputum were cultured on chocolate agar plates. All suspected colonies were identified by Gram staining, colony characteristics, motility and biochemical reactions [9].

2.4. Detection of ESBL by double disk diffusion synergy method ESBL was detected by a double disk synergy technique in which an augmentin disk (amoxicillin 20 ␮g and clavulanic acid 10 ␮g) was placed in the centre of a plate and cefotaxime (30 ␮g), ceftazidime (30 ␮g), aztreonam (30 ␮g) and ceftriaxone (30 ␮g) disks were placed 30 mm (centre to centre) from the augmentin disk. The enhancement of the zone of inhibition of any one of the four drug disks towards the disk containing clavulanic acid suggested the presence of an extended-spectrum ␤-lactamases [12].

2.3. Antibiotic susceptibility testing 3. Results Antimicrobial susceptibility testing of the isolated organisms was done by a disk diffusion method using the Kirby–Bauer technique [10] and as per the recommendations of the NCCLS [11]. All disks were obtained from Oxoid Ltd., Basingstoke, Hampshire, UK. Antibiotic potency of the disks was standardized against the reference strain, E. coli ATCC 25922.

During the 10-month study period, a total of 393 E. coli and K. pneumoniae were isolated from various specimens; antibiograms and production of ESBLs were recorded. The majority of the specimens were urine (48.09%), sputum (22.39%) and pus (18.07%). The detail antibiotic sensitivity pattern of the E. coli and K. pneumoniae isolates is

Table 3 Rate of isolation of ESBL-producing E. coli (n = 81) and K. pneumoniae (n = 76) in different samples Specimen

Total number of E. coli

Number of ESBL positive E. coli

Total number of K. pneumoniae

Number of ESBL positive K. pneumoniae

Urine Sputum Pus Throat swab Others∗

70 4 4 1 2

30 2 1 0 2

27 11 25 9 4

13 6 8 2 1

Total

81

35

76

30

∗

Blood and vaginal swabs.

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M.M. Rahman et al. / International Journal of Antimicrobial Agents 24 (2004) 508–510

given in Table 1. About 45–66% isolated E. coli and K. pneumoniae were sensitive to third generation cephalosporin such as ceftriaxone, ceftazidime and cefotaxime. The first and second generation cephalosporins were less effective. However, most isolates were sensitive to imipenem (94–98%). Of 131 E. coli and 108 K. pneumoniae isolated, we serially tested the first 81 and 76 isolates respectively for ESBL phenotypes. The remaining isolates could not be tested for ESBL production due to logistic constraints. It was observed that 43.2% and 39.5% of E. coli and K. pneumoniae had ESBL phenotypes (Table 2). As shown in Table 2, ESBL-producing E. coli was best detected with an augmentin–ceftazidime combination (39.5%) while an aztreonam–augmentin combination was least sensitive (30.9%). The best combination for detection of ESBL-producing K. pneumoniae was augmentin–ceftriaxone (31.6%) while least sensitive was the augmentin–cefotaxime combination (25.0%). ESBL-producing E. coli were detected in all types of samples (Table 3). About 42.0% of E. coli isolated from urine were ESBL positive. The isolation rate of ESBL-producing K. pneumoniae was highest among pus (54.5%), followed by urine (32.0%), and sputum (17.1%). 4. Discussion It was observed that 43.2% and 39.5% of isolated E. coli and K. pneumoniae had ESBL phenotypes. ESBL-positive E. coli and K. pneumoniae were isolated from all types of specimen studied. This rate is higher than that of countries of the western Pacific region [4], North America or Europe and some South American nations [5,7]. Since no previous data is available about the prevalence of ESBL-positive E. coli and K. pneumoniae in Bangladesh, it is assumed that this high rate of ESBL-positive organisms is due to gradual accumulation in hospital environment and may be a potential source of spread in the community. Recently, an increasing trend of ESBL-producing E. coli (2.1–7.5%) has been reported in stool flora of patients attending outpatient departments in Spain, proving transmission in the community [13]. Also there is a potential for widespread dissemination of these strains due to clonal and horizontal spread [14]. We detected ESBL-producing organisms using several combinations of cephalosporin disks with an augmentin disk. Of the four drugs tested, ceftazidime was found to be the best ESBL detector for E. coli and ceftriaxone for K. pneumoniae in combination with augmentin. Therefore, use of

only one disk combination might fail to detect ESBL production resulting in under reporting of prevalence. Simultaneous use of four cephalosporin disks with an augmentin disk is recommended in screening for ESBL-producing organisms. The present study has, for the first time, revealed the extent of ESBL-producing organisms responsible for various infections in a large hospital of Bangladesh.

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