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Draft Genome Sequence of a Multidrug-Resistant Klebsiella pneumoniae Carbapenemase-Producing Acinetobacter baumannii Sequence Type 2 Isolate from Puerto Rico Teresa Martínez,a Alexander J. Ropelewski,b Ricardo González-Mendez,c Guillermo J. Vázquez,a Iraida E. Robledoa Department of Microbiology and Medical Zoology, University of Puerto Rico, School of Medicine, San Juan, Puerto Ricoa; Pittsburgh Supercomputing Center, Carnegie Mellon University, Pittsburgh, Pennsylvania, USAb; Department of Radiological Sciences, University of Puerto Rico, School of Medicine, San Juan, Puerto Ricoc
We report here the draft genome sequence of Acinetobacter baumannii strain M3AC14-8, sequence type 2 (ST2), carrying a chromosomally carried blaKPC-2 gene. The draft genome consists of a total length of 4.11 Mbp and a GⴙC content of 39.25%. Received 6 June 2016 Accepted 23 June 2016 Published 18 August 2016 Citation Martínez T, Ropelewski AJ, González-Mendez R, Vázquez GJ, Robledo IE. 2016. Draft genome sequence of a multidrug-resistant Klebsiella pneumoniae carbapenemaseproducing Acinetobacter baumannii sequence type 2 isolate from Puerto Rico. Genome Announc 4(4):e00758-16. doi:10.1128/genomeA.00758-16. Copyright © 2016 Martínez et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Iraida E. Robledo, [email protected].
A
cinetobacter baumannii is associated with severe infections that have high morbidity and mortality. Treatment of infections caused by this organism is hampered by the emergence of antibiotic-resistant isolates. The mechanisms of low-level resistance to carbapenems include reduced antibiotic uptake due to loss of porins combined with derepression of the chromosomal AmpC -lactamase gene and/or the overexpression of an efflux pump system (1). -Lactamases are the most common mechanism of resistance to the -lactam antibiotic. They are frequently located on mobile genetic elements, which facilitates their inter- and intraspecies dissemination. Carbapenems are commonly the last-resort antibiotics for the treatment of infections caused by multidrug-resistant Gram-negative bacilli. The Klebsiella pneumoniae carbapenemase (KPC) enzyme hydrolyzes all known -lactam antibiotics, including the carbapenems (2). The KPC-producing A. baumannii strain M3AC14-8 was isolated from the skin of a 62-year-old female patient hospitalized in the intensive care unit. The strain belongs to the globally disseminated sequence type 2 (ST2). Next-generation sequencing was performed using an Illumina MiSeq platform in a 2 ⫻ 150-bp paired-end (PE) configuration by Genewiz, Inc. The Pittsburgh Supercomputing Center Blacklight supercomputer system (3) was used to performed de novo assembly and preliminary genome annotation. De novo assembly was done using a combination of Velvet (version 1.2.10) (4), with additional scaffolding provided through SSPACE Basic 2.0 (5). Additional PCR and sequencing were performed to join the resulting scaffolds. The draft genome of A. baumannii strain M3AC14-8 consists of 30 scaffolds, with a total length of 4,117,354 bp, an average contig length of 146,869 bp, a maximum contig length of 521,258 bp, and two complete and circularized plasmids of 5,441 and 18,043 bp. The G⫹C content was determined to be 39.25%. Genome annotation was performed using the National Center for Biotechnology Information (NCBI) Prokaryotic Genome Annotation Pipeline and was corroborated using the complete proteome of A. baumannii strain MDR-ZJ06 (accession no.
July/August 2016 Volume 4 Issue 4 e00758-16
CP001937), and 3,746 ORFs with E values of ⱕ1e-5 were common to the two genomes. The Pfam database gave 2,285 unique protein families (6). A total of 65 tRNA genes and nine rRNA were identified using tRNAscan-SE (7) and RNAmmer (8), respectively. In addition to the chromosomally carried KPC-2 gene, ResFinder-2.1 at the Center for Genomic Epidemiology server (9) identified the presence of multiple resistance genes against aminoglycoside, -lactam, fluoroquinolone, macrolide, chloramphenicol, sulfonamide, and tetracycline antibiotics in the genome. No resistance genes were associated with the two plasmids identified. Further analysis identified the presence of AdeIJK and AdeABC efflux pumps with amino acid mutations in the regulatory genes (adeRS) as well as amino acid mutations within the gyrA and parC genes. Accession number(s). This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession no. LDDY00000000. The version described in this paper is version LDDY01000000. ACKNOWLEDGMENTS This work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation grant no. OCI-1053575. Specifically, it used the Blacklight supercomputer system at the Pittsburgh Supercomputing Center (PSC). This work was also supported by the PSC’s National Institutes of Health Minority Access to Research Careers (MARC) grant T36-GM-095335 and the University of Puerto Rico Medical Sciences Campus grants MBRS/RISE (R25GM061838-14), RCMI/NIH (8G12- MD007600), Proyecto Adopte un Gen, and Associate Deanship for Biomedical Sciences Graduate Program School of Medicine, Medical Sciences Campus, University of Puerto Rico. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. Any opinions, findings and conclusions or recommendations expressed are those of the authors and do not necessarily reflect the views of the NSF. This research project constitutes a partial fulfillment of the doctoral thesis dissertation of T.M.
Genome Announcements
genomea.asm.org 1
Martínez et al.
FUNDING INFORMATION This work, including the efforts of Teresa Martínez, was funded by Research Center for Minority Institutions (RCMI) (8G12MD007600). This work, including the efforts of Teresa Martínez, was funded by Minority Biomedical Research Support Research Initiative for Scientific Enhancement (MBRS-RISE) (R25GM061838-14). This work, including the efforts of Alexander J. Ropelewski, was funded by HHS | National Institutes of Health (NIH) (T36-GM-095335). This work, including the efforts of Alexander J. Ropelewski, was funded by National Science Foundation (NSF) (OCI-1053575).
REFERENCES
5. 6.
7.
1. Manchanda V, Sanchaita S, Singh N. 2010. Multidrug resistant Acinetobacter. J Glob Infect Dis 2:291–304. http://dx.doi.org/10.4103/0974 -777X.68538. 2. Yigit H, Queenan AM, Anderson GJ, Domenech-Sanchez A, Biddle JW, Steward CD, Alberti S, Bush K, Tenover FC. 2001. Novel carbapenemhydrolyzing beta-lactamase, KPC-1, from a carbapenem-resistant strain of Klebsiella pneumoniae. Antimicrob Agents Chemother 45:1151–1161. http://dx.doi.org/10.1128/AAC.45.4.1151-1161.2001. 3. Towns J, Cockerill T, Dahan M, Foster I, Gaither K, Grimshaw A, Hazlewood V, Lathrop S, Lifka D, Peterson GD, Roskies R, Scott JR,
2 genomea.asm.org
4.
8.
9.
Wilkens-Diehr N. 2014. XSEDE: accelerating scientific discovery. Comput Sci Eng 16:62–74. http://dx.doi.org/10.1109/MCSE.2014.80. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821– 829. http:// dx.doi.org/10.1101/gr.074492.107. Boetzer M, Henkel CV, Jansen HJ, Butler D, Pirovano W. 2011. Scaffolding pre-assembled contigs using SSPACE. Bioinformatics 27:578 –579. http://dx.doi.org/10.1093/bioinformatics/btq683. Finn RD, Mistry J, Schuster-Böckler B, Griffiths-Jones S, Hollich V, Lassmann T, Moxon S, Marshall M, Khanna A, Durbin R, Eddy SR, Sonnhammer EL, Bateman A. 2006. Pfam: clans, Web tools and services. Nucleic Acids Res 34:D247–D251. http://dx.doi.org/10.1093/nar/gkj149. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955. Lagesen K, Hallin P, Rødland EA, Staerfeldt H-H, Rognes T, Ussery DW. 2007. RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res 35:3100 –3108. http://dx.doi.org/10.1093/nar/ gkm160. Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, Aarestrup FM, Larsen MV. 2012. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 67:2640 –2644. http:// dx.doi.org/10.1093/jac/dks261.
Genome Announcements
July/August 2016 Volume 4 Issue 4 e00758-16
Draft Genome Sequence of a Multidrug-Resistant Klebsiella pneumoniae Carbapenemase-Producing Acinetobacter baumannii Sequence Type 2 Isolate from Puerto Rico Teresa Martínez,a Alexander J. Ropelewski,b Ricardo González-Mendez,c Guillermo J. Vázquez,a Iraida E. Robledoa Department of Microbiology and Medical Zoology, University of Puerto Rico, School of Medicine, San Juan, Puerto Ricoa; Pittsburgh Supercomputing Center, Carnegie Mellon University, Pittsburgh, Pennsylvania, USAb; Department of Radiological Sciences, University of Puerto Rico, School of Medicine, San Juan, Puerto Ricoc
We report here the draft genome sequence of Acinetobacter baumannii strain M3AC14-8, sequence type 2 (ST2), carrying a chromosomally carried blaKPC-2 gene. The draft genome consists of a total length of 4.11 Mbp and a GⴙC content of 39.25%. Received 6 June 2016 Accepted 23 June 2016 Published 18 August 2016 Citation Martínez T, Ropelewski AJ, González-Mendez R, Vázquez GJ, Robledo IE. 2016. Draft genome sequence of a multidrug-resistant Klebsiella pneumoniae carbapenemaseproducing Acinetobacter baumannii sequence type 2 isolate from Puerto Rico. Genome Announc 4(4):e00758-16. doi:10.1128/genomeA.00758-16. Copyright © 2016 Martínez et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Iraida E. Robledo, [email protected].
A
cinetobacter baumannii is associated with severe infections that have high morbidity and mortality. Treatment of infections caused by this organism is hampered by the emergence of antibiotic-resistant isolates. The mechanisms of low-level resistance to carbapenems include reduced antibiotic uptake due to loss of porins combined with derepression of the chromosomal AmpC -lactamase gene and/or the overexpression of an efflux pump system (1). -Lactamases are the most common mechanism of resistance to the -lactam antibiotic. They are frequently located on mobile genetic elements, which facilitates their inter- and intraspecies dissemination. Carbapenems are commonly the last-resort antibiotics for the treatment of infections caused by multidrug-resistant Gram-negative bacilli. The Klebsiella pneumoniae carbapenemase (KPC) enzyme hydrolyzes all known -lactam antibiotics, including the carbapenems (2). The KPC-producing A. baumannii strain M3AC14-8 was isolated from the skin of a 62-year-old female patient hospitalized in the intensive care unit. The strain belongs to the globally disseminated sequence type 2 (ST2). Next-generation sequencing was performed using an Illumina MiSeq platform in a 2 ⫻ 150-bp paired-end (PE) configuration by Genewiz, Inc. The Pittsburgh Supercomputing Center Blacklight supercomputer system (3) was used to performed de novo assembly and preliminary genome annotation. De novo assembly was done using a combination of Velvet (version 1.2.10) (4), with additional scaffolding provided through SSPACE Basic 2.0 (5). Additional PCR and sequencing were performed to join the resulting scaffolds. The draft genome of A. baumannii strain M3AC14-8 consists of 30 scaffolds, with a total length of 4,117,354 bp, an average contig length of 146,869 bp, a maximum contig length of 521,258 bp, and two complete and circularized plasmids of 5,441 and 18,043 bp. The G⫹C content was determined to be 39.25%. Genome annotation was performed using the National Center for Biotechnology Information (NCBI) Prokaryotic Genome Annotation Pipeline and was corroborated using the complete proteome of A. baumannii strain MDR-ZJ06 (accession no.
July/August 2016 Volume 4 Issue 4 e00758-16
CP001937), and 3,746 ORFs with E values of ⱕ1e-5 were common to the two genomes. The Pfam database gave 2,285 unique protein families (6). A total of 65 tRNA genes and nine rRNA were identified using tRNAscan-SE (7) and RNAmmer (8), respectively. In addition to the chromosomally carried KPC-2 gene, ResFinder-2.1 at the Center for Genomic Epidemiology server (9) identified the presence of multiple resistance genes against aminoglycoside, -lactam, fluoroquinolone, macrolide, chloramphenicol, sulfonamide, and tetracycline antibiotics in the genome. No resistance genes were associated with the two plasmids identified. Further analysis identified the presence of AdeIJK and AdeABC efflux pumps with amino acid mutations in the regulatory genes (adeRS) as well as amino acid mutations within the gyrA and parC genes. Accession number(s). This whole-genome shotgun project has been deposited at DDBJ/ENA/GenBank under the accession no. LDDY00000000. The version described in this paper is version LDDY01000000. ACKNOWLEDGMENTS This work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation grant no. OCI-1053575. Specifically, it used the Blacklight supercomputer system at the Pittsburgh Supercomputing Center (PSC). This work was also supported by the PSC’s National Institutes of Health Minority Access to Research Careers (MARC) grant T36-GM-095335 and the University of Puerto Rico Medical Sciences Campus grants MBRS/RISE (R25GM061838-14), RCMI/NIH (8G12- MD007600), Proyecto Adopte un Gen, and Associate Deanship for Biomedical Sciences Graduate Program School of Medicine, Medical Sciences Campus, University of Puerto Rico. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. Any opinions, findings and conclusions or recommendations expressed are those of the authors and do not necessarily reflect the views of the NSF. This research project constitutes a partial fulfillment of the doctoral thesis dissertation of T.M.
Genome Announcements
genomea.asm.org 1
Martínez et al.
FUNDING INFORMATION This work, including the efforts of Teresa Martínez, was funded by Research Center for Minority Institutions (RCMI) (8G12MD007600). This work, including the efforts of Teresa Martínez, was funded by Minority Biomedical Research Support Research Initiative for Scientific Enhancement (MBRS-RISE) (R25GM061838-14). This work, including the efforts of Alexander J. Ropelewski, was funded by HHS | National Institutes of Health (NIH) (T36-GM-095335). This work, including the efforts of Alexander J. Ropelewski, was funded by National Science Foundation (NSF) (OCI-1053575).
REFERENCES
5. 6.
7.
1. Manchanda V, Sanchaita S, Singh N. 2010. Multidrug resistant Acinetobacter. J Glob Infect Dis 2:291–304. http://dx.doi.org/10.4103/0974 -777X.68538. 2. Yigit H, Queenan AM, Anderson GJ, Domenech-Sanchez A, Biddle JW, Steward CD, Alberti S, Bush K, Tenover FC. 2001. Novel carbapenemhydrolyzing beta-lactamase, KPC-1, from a carbapenem-resistant strain of Klebsiella pneumoniae. Antimicrob Agents Chemother 45:1151–1161. http://dx.doi.org/10.1128/AAC.45.4.1151-1161.2001. 3. Towns J, Cockerill T, Dahan M, Foster I, Gaither K, Grimshaw A, Hazlewood V, Lathrop S, Lifka D, Peterson GD, Roskies R, Scott JR,
2 genomea.asm.org
4.
8.
9.
Wilkens-Diehr N. 2014. XSEDE: accelerating scientific discovery. Comput Sci Eng 16:62–74. http://dx.doi.org/10.1109/MCSE.2014.80. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821– 829. http:// dx.doi.org/10.1101/gr.074492.107. Boetzer M, Henkel CV, Jansen HJ, Butler D, Pirovano W. 2011. Scaffolding pre-assembled contigs using SSPACE. Bioinformatics 27:578 –579. http://dx.doi.org/10.1093/bioinformatics/btq683. Finn RD, Mistry J, Schuster-Böckler B, Griffiths-Jones S, Hollich V, Lassmann T, Moxon S, Marshall M, Khanna A, Durbin R, Eddy SR, Sonnhammer EL, Bateman A. 2006. Pfam: clans, Web tools and services. Nucleic Acids Res 34:D247–D251. http://dx.doi.org/10.1093/nar/gkj149. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955. Lagesen K, Hallin P, Rødland EA, Staerfeldt H-H, Rognes T, Ussery DW. 2007. RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res 35:3100 –3108. http://dx.doi.org/10.1093/nar/ gkm160. Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, Aarestrup FM, Larsen MV. 2012. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 67:2640 –2644. http:// dx.doi.org/10.1093/jac/dks261.
Genome Announcements
July/August 2016 Volume 4 Issue 4 e00758-16
