ESBL- and Carbapenemase-Producing Enterobacteriaceae in Patients with Bacteremia, Yangon, Myanmar, 2014

Abstract

Among 42 gram-negative bloodstream isolates from inpatients in 3 hospitals in Yangon, Myanmar, admitted during July–December 2014, 16 (38%) were extended-spectrum β-lactamase–producing Enterobacteriaceae and 6 (14%) produced carbapenemase. The high prevalence of multidrug-resistant gram-negative bacteria raises concerns about the empiric treatment of patients with sepsis in Yangon.

Infections with extended-spectrum β-lactamase (ESBL)–producing gram-negative bacteria and carbapenem-resistant Enterobacteriaceae (CRE) have been reported worldwide (1). Little is known about the occurrence of ESBL-producing and CRE bacteria in Yangon, Myanmar. Therefore, we characterized 42 gram-negative organisms isolated from routine blood cultures from adult inpatients in Yangon.

All bacteria had been isolated at the microbiology laboratories of 3 hospitals in Yangon during July–December 2014. During the study period, 592 blood cultures were processed, 536 from Yangon General Hospital (YGH) and 56 from 2 private hospitals. YGH is a 2,000-bed tertiary referral and teaching hospital in Yangon, providing free hospital care to civilians. The 2 private hospitals have 350 and 100 beds and provide secondary-level medical and surgical services to paying patients.

Of the 592 blood cultures, 42 (7.8%) yielded gram-negative bacteria, 28 (67%) from YGH and 14 (33%) from the 2 private hospitals. No clinical information was available about the patients from whom the cultures were taken. The identity and antimicrobial drug susceptibility of isolates were confirmed at Southern Community Laboratories (Dunedin, New Zealand) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (Microflex LT; Bruker Daltonics, Billerica, MA, USA), disc diffusion testing using the Clinical and Laboratory Standards Institute method (2), and the Phoenix Automated Microbiology System (Bruker Daltonics) (panel NMIC/ID-95).

We conducted phenotypic confirmation of ESBL production on cefpodoxime-resistant isolates using cefotaxime and ceftazidime with and without clavulanic acid and that of carbapenmase production on meropenem-resistant isolates by modified Hodge test according to Clinical and Laboratory Standards Institute criteria (2). We performed PCR for β-lactamase genes on all ESBL- and potential carbapenemase-producing organisms (3,4). We conducted bidirectional Sanger sequencing of amplicons and identified DNA sequences by BLAST (https://blast.ncbi.nlm.nih.gov/Blast.cgi) and by comparison with known β-lactamase gene sequences.

Of the 42 isolates, 34 (81%) were Enterobacteriaceae (20 Escherichia coli, 7 Klebsiella pneumoniae, 6 Salmonella enterica, and 1 Enterobacter cloacae) and 8 (19.0%) were nonfermenting gram-negative bacilli. Of the Enterobacteriaceae, 20 (59%) were multidrug resistant (MDR), with resistance to >3 classes of antimicrobial drugs, and 7 (21%) were extensively drug resistant, with susceptibility to <2 classes of antimicrobial drugs (5). All MDR Enterobacteriaceae were susceptible to polymyxin (Table). Phenotypic testing suggested the presence of an ESBL in 16 (38%) and a carbapenemase in 6 (14%) of all gram-negative isolates. Molecular analysis showed that 13 (81%) ESBL-producing isolates contained a group 1 CTX-M gene; all were confirmed as CTX-M-15 by sequencing. Carbapenemase-producing isolates, including 3 E. coli and 3 K. pneumoniae, contained the New Delhi metallo-β-lactamase (NDM) gene, sequenced as NDM-4 in 5 (83%) and NDM-7 in 1 (17%).

Table. Antimicrobial drug susceptibility of Enterobacteriaceae and Acinetobacter spp. isolated from blood cultures of inpatients from 3 hospitals in Yangon, Myanmar, 2014*.

	Organism, no. (%) susceptible
Agent	Escherichia coli, n = 20	Klebsiella pneumoniae, n = 7	Salmonella enterica, n = 6	Enterobacter cloacae, n = 1	Acinetobacter spp., n = 3
Ampicillin	2 (10)	0	6 (100)	0	NT
Amoxicillin/clavulanic acid	5 (25)	1 (14)	6 (100)	0	NT
Piperacillin/tazobactam	13 (65)	2 (29)	6 (100)	0	2 (67)
Cefuroxime	5 (25)	1 (14)	NT	0	NT
Ceftriaxone	6 (30)	1 (14)	6 (100)	0	1 (33)
Ceftazidime	6 (30)	1 (14)	6 (100)	0	0
Cefipime	7 (35)	1 (14)	6 (100)	0	2 (67)
Cefoxitin	11 (55)	4 (57)	NT	0	NT
Aztreonam	6 (30)	1 (14)	6 (100)	0	NT
Ertapenem	17 (85)	4 (57)	6 (100)	1 (100)	NT
Imipenem	17 (85)	4 (57)	6 (100)	1 (100)	3 (100)
Meropenem	17 (85)	4 (57)	6 (100)	1 (100)	3 (100)
Gentamicin	14 (70)	2 (29)	NT	1 (100)	2 (67)
Tobramycin	9 (45)	1 (14)	NT	0	3 (100)
Ciprofloxacin	1 (5)	1 (14)	3 (43)	0	2 (67)
Chloramphenicol	11 (55)	2 (29)	6 (100)	0	NT
Colistin†	20 (100)	7 (100)	6 (100)	1 (100)	3 (100)
Nitrofurantoin	16 (80)	1 (14)	NT	0	NT
Trimethoprim/ sulfamethoxazole	2 (10)	2 (29)	6 (100)	0	2 (67)

*NT, not tested.
†Because there are no Clinical and Laboratory Standards Institute for susceptibility testing criteria for colistin for Enterobaceriaceae, European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria were used (susceptible if MIC <2 mg/L, per EUCAST criteria version 6.0, 2016; http://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Breakpoint_tables/v_6.0_Breakpoint_table.pdf).

Our study revealed a high proportion of ESBL- and carbapenemase-producing organisms among gram-negative bloodstream isolates during the study period from hospital inpatients in Yangon. Half of E. coli isolates and 43% of K. pneumoniae isolates produced ESBLs. This finding is consistent with the high proportion of ESBL production reported in isolates from India (>80%), China (>60%), and other Asia and Southeast Asia countries (>30%) (6). Carbapenemase production (15% in E. coli and 43% in K. pneumoniae) in this study was comparable to those previously reported from clinical isolates in India (7).

CTX-M-15 ESBL and NDM carbapenemase were the most prevalent mechanisms of resistance to β-lactams in our study. This finding is consistent with the current global dissemination of CTX-M-15 among E. coli isolates (8). All CRE isolates were NDM-4 or NDM-7. Two previous case reports have indicated the presence of NDM-producing Enterobacteriaceae from travelers to Myanmar: 1 NDM-7 (9) and 1 NDM-4 (10).

Two thirds of all isolates included in this study originated from YGH, the largest public hospital in Myanmar. All CRE isolates and 14 (88%) of 16 ESBL producers were isolated from YGH; this may reflect a higher prevalence of colonization with MDR organisms among patients in YGH or they may be healthcare-associated infections. Of concern, at YGH 11 (73%) of 15 E. coli and 6 (100%) of 6 K. pneumoniae isolates produced either an ESBL or carbapenemase; among these, 3 (20%) of 15 E. coli and 3 (50%) of 6 K. pneumoniae isolates were NDM producers.

Whereas all 23 (100%) of the Enterobacteriaceae at YGH were susceptible to treatment with colistin, an empiric treatment regimen of meropenem plus gentamicin would have covered only 18 (78%) isolates. This finding highlights the difficulties with designing an effective empiric antimicrobial regimen for patients with suspected gram-negative sepsis in a setting of a high prevalence of antimicrobial resistance, without providing further selective pressure for the spread of CRE and the emergence of colistin resistance.

Our study has limitations. First, clinical data were not prospectively collected, and it was not possible to obtain data retrospectively because of poor recording systems. Second, we cannot be certain that study isolates represent the population of organisms causing gram-negative sepsis in Yangon. However, the high proportion of ESBL- and carbapenemase-producing gram-negative bacteria among bloodstream isolates from hospitalized patients in Yangon raises concern for the treatment of patients with gram-negative sepsis and suggests a need to reduce selective pressure and control the spread of resistant organisms.

Biography

Dr. Myat completed this work while she was working as a lecturer at the Department of Microbiology, University of Medicine 1 in Yangon, Myanmar. Currently, she is undertaking doctoral study from the University of Otago, New Zealand. Her research focuses on bacterial causes of febrile illness in Yangon.