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Draft Genome Sequence of a Clinically Isolated Extensively DrugResistant Pseudomonas aeruginosa Strain Bhavani Manivannan,a Niranjana Mahalingam,b Sudhir Jadhao,c Amrita Mishra,c Pravin Nilawe,d
Bulagonda Eswarappa Pradeepa
Department of Biosciences, Sri Sathya Sai Institute of Higher Learning, Prasanthi Nilayam, Indiaa; Department of Microbiology, Sri Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, Indiab; Bionivid Technology Pvt. Ltd., Bengaluru, Indiac; Thermo Fisher Scientific, Mumbai, Indiad
We present the draft genome assembly of an extensively drug-resistant (XDR) Pseudomonas aeruginosa strain isolated from a patient with a history of genito urinary tuberculosis. The draft genome is 7,022,546 bp with a GⴙC content of 65.48%. It carries 7 phage genomes, genes for quorum sensing, biofilm formation, virulence, and antibiotic resistance. Received 3 February 2016 Accepted 6 February 2016 Published 24 March 2016 Citation Manivannan B, Mahalingam N, Jadhao S, Mishra A, Nilawe P, Pradeep BE. 2016. Draft genome sequence of a clinically isolated extensively drug-resistant Pseudomonas aeruginosa strain. Genome Announc 4(2):e00162-16. doi:10.1128/genomeA.00162-16. Copyright © 2016 Manivannan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Bulagonda Eswarappa Pradeep, [email protected].
P
seudomonas aeruginosa is an opportunistic human pathogen associated with several life-threatening infections including pneumonia, bacteremia, meningitis, cystic fibrosis, and urinary tract and wound infections. Although several genome sequences of P. aeruginosa are available in the GenBank database (1, 2), the enormous versatility of this pathogen to adapt and thrive in various ecological niches developing enhanced resistance to most of the available antibiotics necessitates sequencing of additional clinical isolates. In this report, we present the draft genome sequence of an extensively drug-resistant (XDR) P. aeruginosa recovered from the urine sample of a 33-year old female patient with a history of genito urinary tuberculosis at Sri Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, India. She had undergone a right ureteric reimplant for right lower ureteric stricture with proximal hydroureteronephrosis. This pathogen was found to be resistant to penicillins, second and third generation cephalosporins, aminoglycosides, carbapenems, tetracyclines, quinolones, and trimethoprim-sulfamethoxazole and sensitive to only colistin when tested with Vitek-2. Total DNA was extracted from an overnight culture of a single colony in Luria broth (LB) medium using a genomic DNA extraction kit (Macherey-Nagel, Germany). Sequencing on Ion Torrent single-end technology produced a total of 1,022,829 bp single-end reads with an average read length of 271.50 ⫾ 99.16 bp. Assembly by SPADES v3.5.0 (3) revealed that the draft genome consists of 190 scaffolds with an average length of 36,960.77 bp, G⫹C content of 65.68%, and N50 size of 65,43,920 bp constituting a total of 7,022,546 bp. Genome annotation by RAST (4) and NCBI PGAAP servers revealed that the draft genome has 7,803 protein coding sequences (CDSs), 3 rRNAs, 49 tRNAs, and 1 noncoding RNA (ncRNA) genes. No plasmid was identified when analyzed using Webcutter (v2.0) and Plasmid Finder (v1.3) (5). The contigs of the draft genome share (93.10%) identity with those of the P. aeruginosa 19BR genome having polymyxin-B adaptation (GenBank Accession no. AFXJ00000000) (6). RAST annotation revealed the presence of several genes re-
March/April 2016 Volume 4 Issue 2 e00162-16
sponsible for quorum sensing, biofilm formation, and virulence. Analysis by Resfinder 2.1 (7) indicates that the draft genome has genes encoding for resistance to aminoglycosides, fosfomycin, -lactams, sulfonamides, and trimethoprim. Further, 7 putative prophage sequences were identified through the PHAST server (8). Taxonomy identification of the draft genome by EzTaxon (9) and MEGA6 inferred that Pseudomonas aeruginosa is the putative species (as per sequence homology) while exhibiting closest homology to Pseudomonas otitidis. A more detailed report of this strain will be included in a future publication. Nucleotide sequence accession numbers. The whole-genome shotgun project of this strain has been deposited at DDBJ/EMBL/ GenBank under the accession no. LQBU00000000. The version described in this paper is LQBU00000000.1. ACKNOWLEDGMENTS This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors. We thank the Department of Microbiology, Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, India, for providing the strain and clinical information.
FUNDING INFORMATION This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.
REFERENCES 1. Stover CK, Pham XQ, Erwin AL, Mizoguchi SD, Warrener P, Hickey MJ, Brinkman FS, Hufnagle WO, Kowalik DJ, Lagrou M, Garber RL, Goltry L, Tolentino E, Westbrock-Wadman S, Yuan Y, Brody LL, Coulter SN, Folger KR, Kas A, Larbig K, Lim R, Smith K, Spencer D, Wong GK, Wu Z, Paulsen IT, Reizer J, Saier MH, Hancock RE, Lory S, Olson MV. 2000. Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen. Nature 406:959 –964. http://dx.doi.org/10.1038/ 35023079. 2. Benson DA, Clark K, Karsch-Mizrachi I, Lipman DJ, Ostell J, Sayers EW. 2015. GenBank. Nucleic Acids Res 43:D30 –D35. http://dx.doi.org/ 10.1093/nar/gku1216. 3. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin
Genome Announcements
genomea.asm.org 1
Manivannan et al.
AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA. 2012. SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19:455– 477. http://dx.doi.org/10.1089/cmb.2012.0021. 4. Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M, Vonstein V, Wattam AR, Xia F, Stevens R. 2014. The SEED and the Rapid Annotation of microbial genomes using Subsystems Technology (RAST). Nucleic Acids Res 42: D206 –D214. http://dx.doi.org/10.1093/nar/gkt1226. 5. Carattoli A, Zankari E, García-Fernández A, Voldby Larsen M, Lund O, Villa L, Møller Aarestrup F, Hasman H. 2014. In silico detection and typing of plasmids using plasmidFinder and plasmid multilocus sequence typing. Antimicrob Agents Chemother 58:3895–3903. http://dx.doi.org/ 10.1128/AAC.02412-14. 6. Boyle B, Fernandez L, Laroche J, Kukavica-Ibrulj I, Mendes CM, Hancock RW, Levesque RC. 2012. Complete genome sequences of three Pseu-
2 genomea.asm.org
domonas aeruginosa isolates with phenotypes of polymyxin B adaptation and inducible resistance. J Bacteriol 194:529 –530. http://dx.doi.org/ 10.1128/JB.06246-11. 7. 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. 8. Zhou Y, Liang Y, Lynch KH, Dennis JJ, Wishart DS. 2011. PHAST: a fast phage search tool. Nucleic Acids Res 39:W347–W352. http://dx.doi.org/ 10.1093/nar/gkr485. 9. Kim OS, Cho YJ, Lee K, Yoon SH, Kim M, Na H, Park SC, Jeon YS, Lee JH, Yi H, Won S, Chun J. 2012. Introducing EzTaxon-e: a prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species. Int J Syst Evol Microbiol 62:716 –721. http://dx.doi.org/10.1099/ ijs.0.038075-0.
Genome Announcements
March/April 2016 Volume 4 Issue 2 e00162-16
Draft Genome Sequence of a Clinically Isolated Extensively DrugResistant Pseudomonas aeruginosa Strain Bhavani Manivannan,a Niranjana Mahalingam,b Sudhir Jadhao,c Amrita Mishra,c Pravin Nilawe,d
Bulagonda Eswarappa Pradeepa
Department of Biosciences, Sri Sathya Sai Institute of Higher Learning, Prasanthi Nilayam, Indiaa; Department of Microbiology, Sri Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, Indiab; Bionivid Technology Pvt. Ltd., Bengaluru, Indiac; Thermo Fisher Scientific, Mumbai, Indiad
We present the draft genome assembly of an extensively drug-resistant (XDR) Pseudomonas aeruginosa strain isolated from a patient with a history of genito urinary tuberculosis. The draft genome is 7,022,546 bp with a GⴙC content of 65.48%. It carries 7 phage genomes, genes for quorum sensing, biofilm formation, virulence, and antibiotic resistance. Received 3 February 2016 Accepted 6 February 2016 Published 24 March 2016 Citation Manivannan B, Mahalingam N, Jadhao S, Mishra A, Nilawe P, Pradeep BE. 2016. Draft genome sequence of a clinically isolated extensively drug-resistant Pseudomonas aeruginosa strain. Genome Announc 4(2):e00162-16. doi:10.1128/genomeA.00162-16. Copyright © 2016 Manivannan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Bulagonda Eswarappa Pradeep, [email protected].
P
seudomonas aeruginosa is an opportunistic human pathogen associated with several life-threatening infections including pneumonia, bacteremia, meningitis, cystic fibrosis, and urinary tract and wound infections. Although several genome sequences of P. aeruginosa are available in the GenBank database (1, 2), the enormous versatility of this pathogen to adapt and thrive in various ecological niches developing enhanced resistance to most of the available antibiotics necessitates sequencing of additional clinical isolates. In this report, we present the draft genome sequence of an extensively drug-resistant (XDR) P. aeruginosa recovered from the urine sample of a 33-year old female patient with a history of genito urinary tuberculosis at Sri Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, India. She had undergone a right ureteric reimplant for right lower ureteric stricture with proximal hydroureteronephrosis. This pathogen was found to be resistant to penicillins, second and third generation cephalosporins, aminoglycosides, carbapenems, tetracyclines, quinolones, and trimethoprim-sulfamethoxazole and sensitive to only colistin when tested with Vitek-2. Total DNA was extracted from an overnight culture of a single colony in Luria broth (LB) medium using a genomic DNA extraction kit (Macherey-Nagel, Germany). Sequencing on Ion Torrent single-end technology produced a total of 1,022,829 bp single-end reads with an average read length of 271.50 ⫾ 99.16 bp. Assembly by SPADES v3.5.0 (3) revealed that the draft genome consists of 190 scaffolds with an average length of 36,960.77 bp, G⫹C content of 65.68%, and N50 size of 65,43,920 bp constituting a total of 7,022,546 bp. Genome annotation by RAST (4) and NCBI PGAAP servers revealed that the draft genome has 7,803 protein coding sequences (CDSs), 3 rRNAs, 49 tRNAs, and 1 noncoding RNA (ncRNA) genes. No plasmid was identified when analyzed using Webcutter (v2.0) and Plasmid Finder (v1.3) (5). The contigs of the draft genome share (93.10%) identity with those of the P. aeruginosa 19BR genome having polymyxin-B adaptation (GenBank Accession no. AFXJ00000000) (6). RAST annotation revealed the presence of several genes re-
March/April 2016 Volume 4 Issue 2 e00162-16
sponsible for quorum sensing, biofilm formation, and virulence. Analysis by Resfinder 2.1 (7) indicates that the draft genome has genes encoding for resistance to aminoglycosides, fosfomycin, -lactams, sulfonamides, and trimethoprim. Further, 7 putative prophage sequences were identified through the PHAST server (8). Taxonomy identification of the draft genome by EzTaxon (9) and MEGA6 inferred that Pseudomonas aeruginosa is the putative species (as per sequence homology) while exhibiting closest homology to Pseudomonas otitidis. A more detailed report of this strain will be included in a future publication. Nucleotide sequence accession numbers. The whole-genome shotgun project of this strain has been deposited at DDBJ/EMBL/ GenBank under the accession no. LQBU00000000. The version described in this paper is LQBU00000000.1. ACKNOWLEDGMENTS This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors. We thank the Department of Microbiology, Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, India, for providing the strain and clinical information.
FUNDING INFORMATION This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.
REFERENCES 1. Stover CK, Pham XQ, Erwin AL, Mizoguchi SD, Warrener P, Hickey MJ, Brinkman FS, Hufnagle WO, Kowalik DJ, Lagrou M, Garber RL, Goltry L, Tolentino E, Westbrock-Wadman S, Yuan Y, Brody LL, Coulter SN, Folger KR, Kas A, Larbig K, Lim R, Smith K, Spencer D, Wong GK, Wu Z, Paulsen IT, Reizer J, Saier MH, Hancock RE, Lory S, Olson MV. 2000. Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen. Nature 406:959 –964. http://dx.doi.org/10.1038/ 35023079. 2. Benson DA, Clark K, Karsch-Mizrachi I, Lipman DJ, Ostell J, Sayers EW. 2015. GenBank. Nucleic Acids Res 43:D30 –D35. http://dx.doi.org/ 10.1093/nar/gku1216. 3. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin
Genome Announcements
genomea.asm.org 1
Manivannan et al.
AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA. 2012. SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19:455– 477. http://dx.doi.org/10.1089/cmb.2012.0021. 4. Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M, Vonstein V, Wattam AR, Xia F, Stevens R. 2014. The SEED and the Rapid Annotation of microbial genomes using Subsystems Technology (RAST). Nucleic Acids Res 42: D206 –D214. http://dx.doi.org/10.1093/nar/gkt1226. 5. Carattoli A, Zankari E, García-Fernández A, Voldby Larsen M, Lund O, Villa L, Møller Aarestrup F, Hasman H. 2014. In silico detection and typing of plasmids using plasmidFinder and plasmid multilocus sequence typing. Antimicrob Agents Chemother 58:3895–3903. http://dx.doi.org/ 10.1128/AAC.02412-14. 6. Boyle B, Fernandez L, Laroche J, Kukavica-Ibrulj I, Mendes CM, Hancock RW, Levesque RC. 2012. Complete genome sequences of three Pseu-
2 genomea.asm.org
domonas aeruginosa isolates with phenotypes of polymyxin B adaptation and inducible resistance. J Bacteriol 194:529 –530. http://dx.doi.org/ 10.1128/JB.06246-11. 7. 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. 8. Zhou Y, Liang Y, Lynch KH, Dennis JJ, Wishart DS. 2011. PHAST: a fast phage search tool. Nucleic Acids Res 39:W347–W352. http://dx.doi.org/ 10.1093/nar/gkr485. 9. Kim OS, Cho YJ, Lee K, Yoon SH, Kim M, Na H, Park SC, Jeon YS, Lee JH, Yi H, Won S, Chun J. 2012. Introducing EzTaxon-e: a prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species. Int J Syst Evol Microbiol 62:716 –721. http://dx.doi.org/10.1099/ ijs.0.038075-0.
Genome Announcements
March/April 2016 Volume 4 Issue 2 e00162-16
