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Lim, Cho, Kim, Kim, Kwon, Park, and Koo: The Genetic Characteristics of Multidrug-resistant Acinetobacter baumannii Coproducing 16S rRNA Methylase armA and Carbapenemase OXA-23
Original Article

J Bacteriol Virol 2013;43(1):27-36.

Published online: 9 February 2013

DOI: https://doi.org/10.4167/jbv.2013.43.1.27

The Genetic Characteristics of Multidrug-resistant Acinetobacter baumannii Coproducing 16S rRNA Methylase armA and Carbapenemase OXA-23

Jinsook Lim, Hye Hyun Cho, Semi Kim, Jimyung Kim, Kye Chul Kwon, Jong Woo Park, Sun Hoe Koo*

Department of Laboratory Medicine, College of Medicine, Chungnam National University, Daejeon, Korea

*Corresponding author: Sun Hoe Koo, M.D., Ph.D. Department of Laboratory Medicine, Chungnam National University Hospital, 33 Munhwa-ro, Joong-gu, Daejeon, 301-721, Korea. Phone: +82-42-280-7798, Fax: +82-42-257-5365, e-mail: shkoo@cnu.ac.kr

Received 28 December 201211 January 2013       Accepted 29 January 2013

Copyright © 2013 Journal of Bacteriology and Virology

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Acinetobacter baumannii is a gram-negative organism reported worldwide as a cause of health-care associated infections. Due to its increasing drug resistance, several studies on coproduction of armA and carbapenemase in South Korea and other parts of the world were reported, which can pose significant therapeutic threat. The aim of this study was to investigate genetic characteristics of multidrug-resistant A. baumannii coproducing armA and carbapenemase and its epidemiological relatedness. Forty-five multidrug resistant (MDR) A. baumannii clinical isolates were collected. Antimicrobial susceptibility was determined by agar dilution, Etest and VITEK 2 system. The presence of 16S rRNA methylase and carbapenemase were analyzed by polymerase chain reaction (PCR) and sequencing. Repetitive element palindromic (REP)-PCR was also performed for epidemiologic investigation. All of A. baumannii isolates harbored blaOXA-51-like gene and 10 isolates showed an upstream ISAba1. 36 isolates (80%) showed amplification of OXA-23, all of which except one had an upstream ISAba1. 16S rRNA methylase armA was found in 44 isolates with high level resistance to aminoglycosides. The rate of coproduction was found in 36 isolates (80%). All isolates showed dominant two patterns in REP-PCR profile. The prevalence of MDR A. baumannii coproducing OXA-23 and armA was high, which the rate of blaOXA-23 coproduction was also high.

Keywords: Acinetobacter baumannii, 16S rRNA methylase, Aminoglycoside resistance

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Figure 1.
Repetitive extragenic palindromic (REP)-PCR of genomic DNA from MDR A. baumannii clinical isolates. Most isolates showed A pattern, while some isolate 208 and 218 showed B type.
jbv-43-27f1.tif
Table 1.
Primers used in this study
Primer Nucleotide sequence (5′ → 3′) Reference
armA F AGGTTGTTTCCATTTCTGAG 14
armA R TCTCTTCCATTCCCTTCTCC
rmtA F CTA GCG TCC ATC CTT TCC TC 14
rmtA R TTT GCT TCC ATG CCC TTG CC
rmtB F CCC AAA CAG ACC GTA GAG GC 14
rmtB R CTC AAA CTC GGC GGG CAA GC
rmtC F CGA AGA AGT AAC AGC CAA AG 14
rmtC R ATC CCA ACA TCT CTC CCA CT
rmtD F ATG AGC GAA CTG AAG GAA AAA CTG C 14
rmtD R GCT CCA AAA GCG GCA GCA CCT TA
npmA F GGAGGGCTATCTAATGTGGT 11
npmA R GCCCAAAGAGAATTAAACTG
OXA-23-like F CTTGCTATGTGGTTGCTTCTC 23
OXA-23-like R ATCCATTGCCCAACCAGTC
OXA-51-like F ATGAACATTAAAGCACTC 23
OXA-51-like R CTATAAAATACCTAATTGTTC
OXA-24-like F GTACTAATCAAAGTTGTGAA 24
OXA-24-like R TTCCCCTAACATGAATTTGT
OXA-58-like F CGATCAGAATGTTCAAGCGC 24
OXA-58-like R ACGATTCTCCCTCTGCGC
IMP F CATGGTTTGGTGGTTCTTGT 24
IMF R ATAATTTGGCGGACTTTGGC
VIM F ATTGGTCTATTTGACCGCGTC 24
VIM R TGCTACTCAACGACTGAGCG
PW 166 (ISAba1) CCTATCAGGGTTCTGCCTTCT 23
REP 1 IIIGCGCCGICATCAGGC 25
REP 2 ACGTCTTATCAGGCCTAC
Table 2.
Minimal inhibitory concentrations of different antimicrobial agents
Isolates MIC (μg/ml)
CIP MN CS AN GEN TOB IMP MEM FEP P/T AZ
171 >64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 64
173 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 16 ≥256 64
177 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 16 ≥256 64
183 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 48 ≥256 32
191 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 64
193 64 2 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 8 ≥256 16
196 64 2 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 24 ≥256 16
197 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 ≥256
202 64 2 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 12 ≥256 16
207 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 16
208 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 48
211 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 48
216 64 4 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 24 ≥256 48
217 64 2 2 6 24 <256 1 1 24 ≥256 32
218 64 2 1 ≥1024 ≥1024 ≥1024 1 0.5 12 ≥256 16
219 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 24 < 0.16 16
223 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 24 ≥256 ≥256
225 64 2 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 64
226 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 24 ≥256 ≥256
227 64 2 0.5 ≥1024 ≥1024 ≥1024 ≥32 ≥32 8 ≥256 16
228 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 ≥256
229 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 ≥256
230 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 48 ≥256 ≥256
231 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 ≥256
232 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 48
233 32 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 ≥256
234 32 ≤1 1 ≥1024 ≥1024 ≥1024 2 0.75 8 ≥256 16
235 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 ≥256
236 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 64
237 > 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 64
238 64 ≤1 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥256 ≥256
241 > 64 ≤1 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥256 ≥256 16
242 64 2 1 ≥1024 ≥1024 ≥1024 4 8 16 ≥256 24
243 16 ≤1 2 512 ≥1024 <265 1 0.75 ≥64 ≥128 ≥64
61 ≥4 ≤1 ≤0.5 ≥1024 ≥1024 ≥1024 ≥32 ≥32 32 ≥128 32
68 ≥4 ≤1 ≤0.5 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
90 ≥4 8 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
93 ≥4 8 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
94 ≥4 16 1 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
96 ≥4 16 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
104 ≥4 8 ≤0.5 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64
108 ≥4 4 ≤0.5 512 128 128 ≥32 ≥32 ≥64 ≥128 ≥64
146 ≥4 16 ≤0.5 128 1024 128 ≥32 ≥32 ≥64 ≥128 ≥64
154 ≤0.25 ≤1 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 4 ≤4 32
159 ≥4 ≤1 2 ≥1024 ≥1024 ≥1024 ≥32 ≥32 ≥64 ≥128 ≥64

MICs were determined by Etest (bioMérieux, Marcy l'Etoile, France). But MICs of antimicrobials except Amikacin, gentamicin, tobramycin, imipenem and meropenem were determined by VITEK 2 system (bioMérieux, Marcy l'Etoile, France) in isolates in red color. Abbreviations: MIC, minimal inhibitory concentration; CIP, ciprofloxacin; MN, minocycline; CS, colistin; AN, amikacin; GEN, gentamicin; TOB, tobramycin; IMP, imipenem, MEM, meropenem, FEP, cefepime, P/T, piperacillin/tazobactam; AZ, azteronam.

Table 3.
Genetic characteristics of MDR A. bauamannii
Isolates Antimicrobial resistance determinants
OXA-51 ISAba1/OXA-51 OXA-23 ISAba1/OXA-23 armA REP-PCR
171 + − + + + A
173 + − + + + A
177 + − + + + A
183 + − + + + A
191 + − + + + A
193 + + − − + A
196 + − + + + A
197 + − + + + A
202 + + − − + A
207 + − + + + A
208 + − + + + B
211 + − + + + A
216 + − + + + A
217 + − − − − A
218 + + − − + B
219 + − + + + A
223 + − + + + A
225 + + + + + B
226 + − + + + A
227 + + − − + A
228 + − + + + B
229 + − + + + A
230 + + + + + B
231 + − + + + A
232 + − + + + A
233 + + − − + A
234 + + − − + B
235 + − + + + A
236 + − + + + A
237 + − + + + A
238 + − + + + C
241 + − + + + A
242 + + − − + A
243 + − − − + C
61 + − + + + A
68 + − + + + A
90 + − + + + A
93 + − + − + A
94 + − + + + A
96 + − + + + A
104 + + + + + A
108 + − + + + A
146 + − + + + A
154 + − + + + A
159 + − + + + A
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