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Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China
Accepted Manuscript Title: Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China Authors: Juan Xu, Wenping Lin, Weixing Xi, Yingyu Yao PII: DOI: Reference:
S2213-7165(17)30065-6 http://dx.doi.org/doi:10.1016/j.jgar.2017.03.004 JGAR 390
To appear in: Received date: Revised date: Accepted date:
23-1-2017 23-3-2017 29-3-2017
Please cite this article as: Juan Xu, Wenping Lin, Weixing Xi, Yingyu Yao, Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China (2010), http://dx.doi.org/10.1016/j.jgar.2017.03.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China
Juan Xu a,1, Wenping Lin b,1, Weixing Xi c, Yingyu Yao d,*
a
Institute of Hygiene, Zhejiang Academy of Medical Sciences, Hangzhou, Zhejiang
310013, China b
Centers for Disease Control and Prevention of Ningbo, Ningbo, Zhejiang 315010,
China c
Department of Clinical Laboratory, Tongde Hospital of Zhejiang Province, Hangzhou,
Zhejiang 310012, China d
Department of Obstetrics, Tongde Hospital of Zhejiang Province, Hangzhou,
Zhejiang 310012, China
ARTICLE INFO Article history: Received 23 January 2017 Accepted 29 March 2017
* Corresponding author. Tel.: +86 571 8997 2343; fax: +86 571 8997 2343. E-mail address: [email protected] (Y. Yao).
1
These two authors contributed equally to this study.
ABSTRACT Acinetobacter baumannii has emerged worldwide as a dominant pathogen in nosocomial infections. In this study, we report the draft genome sequence of a clinical multidrug-resistant (MDR) A. baumannii ST191 (CC92) strain. Whole-genome sequencing of the isolate was performed using an Illumina HiSeqTM 2500 system, and bioinformatics analysis was also performed. The draft genome length was 4 259 210 bp, harbouring 14 gene sequences relevant to antibiotic resistance. Antimicrobial susceptibility testing revealed that the isolate was resistant to all of the tested antibiotics except for tigecycline and colistin. These data will facilitate further understanding of the genomic and resistance features of MDR A. baumannii.
Keywords: Acinetobacter baumannii blaOXA-23 Bioinformatics analysis
Acinetobacter baumannii has emerged as a dominant opportunistic Gram-negative bacterial pathogen causing a variety of infections. Owing to the rapid evolution of antimicrobial resistance, multidrug-resistant (MDR) A. baumannii isolates, particularly carbapenem-resistant isolates, are frequently disseminated in nosocomial infections,
leaving few therapeutic options [1]. Acquired resistance to carbapenems in A. baumannii is mainly caused by mechanisms associated with carbapenem-hydrolysing class D -lactamases (CHDLs), particularly blaOXA-23 [2].
In this study, A. baumannii isolate TDAB1 was recovered from a blood sample of a female patient hospitalised with septicaemia in Hangzhou, Zhejiang Province, China, in 2016. The strain was identified both according to VITEK®2 system (bioMérieux, Marcy-l’Étoile, France) and rpoB gene sequencing. Antimicrobial susceptibility testing was conducted by the Etest method, including cefotaxime, cefepime, imipenem, meropenem, trimethoprim/sulfamethoxazole, piperacillin/tazobactam, ciprofloxacin, amikacin, tigecycline and colistin. The isolate was resistant to all of the tested antimicrobials except for tigecycline and colistin. The minimum inhibitory concentrations (MICs) are presented in Supplementary Table S1.
Genomic DNA was extracted using a QIAamp DNA Mini Kit (QIAGEN, Valencia, CA) following the manufacturer’s instructions. A TruSeq DNA Sample Preparation Kit (Illumina Inc., San Diego, CA) was used to create libraries for Illumina paired-end sequencing. The genome of A. baumannii TDAB1 was sequenced using an Illumina HiSeqTM 2500 system (Illumina Inc.) following a paired-end 2 150 bp protocol. The draft genome sequence was assembled using CLC Genomics Workbench 9.0 software (QIAGEN) and was automatically annotated by the NCBI Prokaryotic
Genomes Annotation Pipeline (PGAP) server. Multilocus sequence typing (MLST) of the isolate was analysed by BacWGSTdb server [3].
Resistance-related genes were analysed using ResFinder 2.1 with a 90% threshold for identification of genes [4]. Further bioinformatics analysis, such as identification of genomic islands, insertion elements (IS), prophage sequences, clustered regularly interspaced short palindromic repeat (CRISPR) sequences and secondary metabolite gene clusters, were predicted by application of IslandViewer, ISfinder, PHASTER, CRISPRFinder and antiSMASH tools, respectively [5,6].
The draft genome sequence of A. baumannii TDAB1 consisted of 64 contigs comprising 4 259 210 bp, and the PGAP server predicted a total of 3721 protein-coding sequences. The overall G+C content amounted to 39.4%. In total, 63 tRNA genes and 3 rRNA operons were identified. According to MLST analysis (Oxford/Pasteur), A. baumannii TDAB1 belongs to sequence type (ST) ST191/ST2, corresponding to clonal complex (CC) CC92/CC2. The predicted resistance genes are presented in Table 1. The aminoglycoside resistance genes aadA1, aph(3′)-Ic, armA and aacA4, the lactam
resistance
genes
blaOXA-66,
blaADC-25,
blaOXA-23
and
blaTEM-1D,
the
fluoroquinolone and aminoglycoside resistance gene aac(6′)-Ib-cr, the macrolide– lincosamide–streptogramin B resistance gene msr(E), the macrolide resistance gene mph(E), the phenicol resistance gene catB8 and the sulphonamide resistance genes
sul1 and sul2 were identified. The genome also contains at least 14 genomic islands and several IS elements, with the majority belonging to the IS3 and IS4 families. One incomplete prophage sequence can be predicted in the genome, and four putative secondary metabolite gene clusters including aryl polyene, a siderophore, hserlactone and a non-ribosomal peptide synthetase (NRPS) can also be predicted.
In summary, we report the draft genome sequence of a clinical MDR A. baumannii ST191/ST2 (CC92/CC2) strain harbouring 14 antimicrobial resistance genes in China. Antimicrobial susceptibility testing revealed that the isolate was resistant to most antibiotics except for tigecycline and colistin. Further studies and comparative analysis of this and other MDR A. baumannii are currently underway, which may help us to understand the genomic diversity of this bacterial pathogen in China. These data will unveil the molecular mechanisms contributing to the rapid development of antimicrobial resistance and expand our understanding of the genomic features of MDR A. baumannii in China.
This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number MSRV00000000. The version described in this paper is the first version, MSRV01000000.
Funding: This study was supported by a research grant from Zhejiang Traditional Chinese Medical Science and Technology Plan [2017ZA146].
Competing interests: None declared.
Ethical approval: Not required.
References [1] Mahamat A, Bertrand X, Moreau B, Hommel D, Couppie P, Simonnet C, et al. Clinical epidemiology and resistance mechanisms of carbapenem-resistant Acinetobacter baumannii, French Guiana, 2008–2014. Int J Antimicrob Agents 2016;48:51–5. [2] Poirel L, Nordmann P. Carbapenem resistance in Acinetobacter baumannii: mechanisms and epidemiology. Clin Microbiol Infect 2006;12:826–36. [3] Ruan Z, Feng Y. BacWGSTdb, a database for genotyping and source tracking bacterial pathogens. Nucleic Acids Res 2016;44:D682–7. [4] Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, et al. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 2012;67:2640–4. [5] Dhillon BK, Laird MR, Shay JA, Winsor GL, Lo R, Nizam F, et al. IslandViewer 3: more flexible, interactive genomic island discovery, visualization and analysis. Nucleic Acids Res 2015;43:W104–8. [6] Siguier P, Perochon J, Lestrade L, Mahillon J, Chandler M. ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res 2006;34:D32–6.
Table 1 Resistance genes in Acinetobacter baumannii strain TDAB1 Resistan
%
ce gene
Identit length/qu y
HSP
Contig
Position in
Predicted
Accessio
contig
phenotype
n no.
2336..3127
Aminoglycosi
JQ41404
ery
Aminoglycosides aadA1
100
792/792
contig_7 5
de
1
resistance aph(3′)-
99.88
816/816
Ic
contig_1
70..885
00
Aminoglycosi
X62115
de resistance
armA
100
774/774
contig_7
7816..8589
5
Aminoglycosi de
AY2205 58
resistance aacA4
99.64
555/555
contig_7
999..1553
5
Aminoglycosi de
KM2781 99
resistance -Lactams blaOXA-
100
825/825
6
66
blaADC-
99.91
1152/115
100
822/822
1D
99 30..1181
99.88
861/861
-Lactam resistance -Lactam resistance
contig_5
5228..6049
6
23
blaTEM-
contig_8
23875..246
2
25
blaOXA-
contig_1
contig_8 5
-Lactam resistance
719..1579
-Lactam resistance
FJ36053 0 EF01635 5 HQ7003 58 AF18820 0
Fluoroquinolones aac(6′)-
99.23
519/519
Ib-cr
contig_7
1035..1553
5
Fluoroquinolo ne and
EF63646 1
aminoglycos ide resistance Macrolide–lincosamide–streptogramin B (MLSB) msr(E)
100
1476/147 6
mph(E)
100
885/885
contig_7 5 contig_7 5
10888..123 63 12419..133 03
MLSB resistance Macrolide resistance
EU2942 28 EU2942 28
Phenicols catB8
100
633/633
contig_7
1646..2278
5
Phenicol resistance
AF22750 6
Sulphonamides sul1
100
927/927
contig_7
3545..4471
5 sul2
100
816/816
contig_1 31
HSP, high-scoring segment pair.
Sulphonamid e resistance
647..1462
Sulphonamid e resistance
CP0021 51 GQ4214 66
S2213-7165(17)30065-6 http://dx.doi.org/doi:10.1016/j.jgar.2017.03.004 JGAR 390
To appear in: Received date: Revised date: Accepted date:
23-1-2017 23-3-2017 29-3-2017
Please cite this article as: Juan Xu, Wenping Lin, Weixing Xi, Yingyu Yao, Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China (2010), http://dx.doi.org/10.1016/j.jgar.2017.03.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Draft genome sequence of a multidrug-resistant OXA-23-producing Acinetobacter baumannii ST191 clinical isolate from China
Juan Xu a,1, Wenping Lin b,1, Weixing Xi c, Yingyu Yao d,*
a
Institute of Hygiene, Zhejiang Academy of Medical Sciences, Hangzhou, Zhejiang
310013, China b
Centers for Disease Control and Prevention of Ningbo, Ningbo, Zhejiang 315010,
China c
Department of Clinical Laboratory, Tongde Hospital of Zhejiang Province, Hangzhou,
Zhejiang 310012, China d
Department of Obstetrics, Tongde Hospital of Zhejiang Province, Hangzhou,
Zhejiang 310012, China
ARTICLE INFO Article history: Received 23 January 2017 Accepted 29 March 2017
* Corresponding author. Tel.: +86 571 8997 2343; fax: +86 571 8997 2343. E-mail address: [email protected] (Y. Yao).
1
These two authors contributed equally to this study.
ABSTRACT Acinetobacter baumannii has emerged worldwide as a dominant pathogen in nosocomial infections. In this study, we report the draft genome sequence of a clinical multidrug-resistant (MDR) A. baumannii ST191 (CC92) strain. Whole-genome sequencing of the isolate was performed using an Illumina HiSeqTM 2500 system, and bioinformatics analysis was also performed. The draft genome length was 4 259 210 bp, harbouring 14 gene sequences relevant to antibiotic resistance. Antimicrobial susceptibility testing revealed that the isolate was resistant to all of the tested antibiotics except for tigecycline and colistin. These data will facilitate further understanding of the genomic and resistance features of MDR A. baumannii.
Keywords: Acinetobacter baumannii blaOXA-23 Bioinformatics analysis
Acinetobacter baumannii has emerged as a dominant opportunistic Gram-negative bacterial pathogen causing a variety of infections. Owing to the rapid evolution of antimicrobial resistance, multidrug-resistant (MDR) A. baumannii isolates, particularly carbapenem-resistant isolates, are frequently disseminated in nosocomial infections,
leaving few therapeutic options [1]. Acquired resistance to carbapenems in A. baumannii is mainly caused by mechanisms associated with carbapenem-hydrolysing class D -lactamases (CHDLs), particularly blaOXA-23 [2].
In this study, A. baumannii isolate TDAB1 was recovered from a blood sample of a female patient hospitalised with septicaemia in Hangzhou, Zhejiang Province, China, in 2016. The strain was identified both according to VITEK®2 system (bioMérieux, Marcy-l’Étoile, France) and rpoB gene sequencing. Antimicrobial susceptibility testing was conducted by the Etest method, including cefotaxime, cefepime, imipenem, meropenem, trimethoprim/sulfamethoxazole, piperacillin/tazobactam, ciprofloxacin, amikacin, tigecycline and colistin. The isolate was resistant to all of the tested antimicrobials except for tigecycline and colistin. The minimum inhibitory concentrations (MICs) are presented in Supplementary Table S1.
Genomic DNA was extracted using a QIAamp DNA Mini Kit (QIAGEN, Valencia, CA) following the manufacturer’s instructions. A TruSeq DNA Sample Preparation Kit (Illumina Inc., San Diego, CA) was used to create libraries for Illumina paired-end sequencing. The genome of A. baumannii TDAB1 was sequenced using an Illumina HiSeqTM 2500 system (Illumina Inc.) following a paired-end 2 150 bp protocol. The draft genome sequence was assembled using CLC Genomics Workbench 9.0 software (QIAGEN) and was automatically annotated by the NCBI Prokaryotic
Genomes Annotation Pipeline (PGAP) server. Multilocus sequence typing (MLST) of the isolate was analysed by BacWGSTdb server [3].
Resistance-related genes were analysed using ResFinder 2.1 with a 90% threshold for identification of genes [4]. Further bioinformatics analysis, such as identification of genomic islands, insertion elements (IS), prophage sequences, clustered regularly interspaced short palindromic repeat (CRISPR) sequences and secondary metabolite gene clusters, were predicted by application of IslandViewer, ISfinder, PHASTER, CRISPRFinder and antiSMASH tools, respectively [5,6].
The draft genome sequence of A. baumannii TDAB1 consisted of 64 contigs comprising 4 259 210 bp, and the PGAP server predicted a total of 3721 protein-coding sequences. The overall G+C content amounted to 39.4%. In total, 63 tRNA genes and 3 rRNA operons were identified. According to MLST analysis (Oxford/Pasteur), A. baumannii TDAB1 belongs to sequence type (ST) ST191/ST2, corresponding to clonal complex (CC) CC92/CC2. The predicted resistance genes are presented in Table 1. The aminoglycoside resistance genes aadA1, aph(3′)-Ic, armA and aacA4, the lactam
resistance
genes
blaOXA-66,
blaADC-25,
blaOXA-23
and
blaTEM-1D,
the
fluoroquinolone and aminoglycoside resistance gene aac(6′)-Ib-cr, the macrolide– lincosamide–streptogramin B resistance gene msr(E), the macrolide resistance gene mph(E), the phenicol resistance gene catB8 and the sulphonamide resistance genes
sul1 and sul2 were identified. The genome also contains at least 14 genomic islands and several IS elements, with the majority belonging to the IS3 and IS4 families. One incomplete prophage sequence can be predicted in the genome, and four putative secondary metabolite gene clusters including aryl polyene, a siderophore, hserlactone and a non-ribosomal peptide synthetase (NRPS) can also be predicted.
In summary, we report the draft genome sequence of a clinical MDR A. baumannii ST191/ST2 (CC92/CC2) strain harbouring 14 antimicrobial resistance genes in China. Antimicrobial susceptibility testing revealed that the isolate was resistant to most antibiotics except for tigecycline and colistin. Further studies and comparative analysis of this and other MDR A. baumannii are currently underway, which may help us to understand the genomic diversity of this bacterial pathogen in China. These data will unveil the molecular mechanisms contributing to the rapid development of antimicrobial resistance and expand our understanding of the genomic features of MDR A. baumannii in China.
This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number MSRV00000000. The version described in this paper is the first version, MSRV01000000.
Funding: This study was supported by a research grant from Zhejiang Traditional Chinese Medical Science and Technology Plan [2017ZA146].
Competing interests: None declared.
Ethical approval: Not required.
References [1] Mahamat A, Bertrand X, Moreau B, Hommel D, Couppie P, Simonnet C, et al. Clinical epidemiology and resistance mechanisms of carbapenem-resistant Acinetobacter baumannii, French Guiana, 2008–2014. Int J Antimicrob Agents 2016;48:51–5. [2] Poirel L, Nordmann P. Carbapenem resistance in Acinetobacter baumannii: mechanisms and epidemiology. Clin Microbiol Infect 2006;12:826–36. [3] Ruan Z, Feng Y. BacWGSTdb, a database for genotyping and source tracking bacterial pathogens. Nucleic Acids Res 2016;44:D682–7. [4] Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, et al. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 2012;67:2640–4. [5] Dhillon BK, Laird MR, Shay JA, Winsor GL, Lo R, Nizam F, et al. IslandViewer 3: more flexible, interactive genomic island discovery, visualization and analysis. Nucleic Acids Res 2015;43:W104–8. [6] Siguier P, Perochon J, Lestrade L, Mahillon J, Chandler M. ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res 2006;34:D32–6.
Table 1 Resistance genes in Acinetobacter baumannii strain TDAB1 Resistan
%
ce gene
Identit length/qu y
HSP
Contig
Position in
Predicted
Accessio
contig
phenotype
n no.
2336..3127
Aminoglycosi
JQ41404
ery
Aminoglycosides aadA1
100
792/792
contig_7 5
de
1
resistance aph(3′)-
99.88
816/816
Ic
contig_1
70..885
00
Aminoglycosi
X62115
de resistance
armA
100
774/774
contig_7
7816..8589
5
Aminoglycosi de
AY2205 58
resistance aacA4
99.64
555/555
contig_7
999..1553
5
Aminoglycosi de
KM2781 99
resistance -Lactams blaOXA-
100
825/825
6
66
blaADC-
99.91
1152/115
100
822/822
1D
99 30..1181
99.88
861/861
-Lactam resistance -Lactam resistance
contig_5
5228..6049
6
23
blaTEM-
contig_8
23875..246
2
25
blaOXA-
contig_1
contig_8 5
-Lactam resistance
719..1579
-Lactam resistance
FJ36053 0 EF01635 5 HQ7003 58 AF18820 0
Fluoroquinolones aac(6′)-
99.23
519/519
Ib-cr
contig_7
1035..1553
5
Fluoroquinolo ne and
EF63646 1
aminoglycos ide resistance Macrolide–lincosamide–streptogramin B (MLSB) msr(E)
100
1476/147 6
mph(E)
100
885/885
contig_7 5 contig_7 5
10888..123 63 12419..133 03
MLSB resistance Macrolide resistance
EU2942 28 EU2942 28
Phenicols catB8
100
633/633
contig_7
1646..2278
5
Phenicol resistance
AF22750 6
Sulphonamides sul1
100
927/927
contig_7
3545..4471
5 sul2
100
816/816
contig_1 31
HSP, high-scoring segment pair.
Sulphonamid e resistance
647..1462
Sulphonamid e resistance
CP0021 51 GQ4214 66