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Announcement
27 November 2013

Genome Sequences of Five B1 Subcluster Mycobacteriophages

ABSTRACT

Mycobacteriophages infect members of the Mycobacterium genus in the phylum Actinobacteria and exhibit remarkable diversity. Genome analysis groups the thousands of known mycobacteriophages into clusters, of which the B1 subcluster is currently the third most populous. We report the complete genome sequences of five additional members of the B1 subcluster.

GENOME ANNOUNCEMENT

More than 3,600 mycobacteriophages have been isolated to date, and >298 of their genome sequences are accessible in GenBank (see http://www.phagesdb.org). These numbers reflect the significant biological diversity among bacteriophages, especially because all of these phages infect a single host, Mycobacterium smegmatis strain mc2155 (1). Dot-plot analyses, average nucleotide identities, gene content analyses, and pairwise genome analyses of mycobacteriophage genomes grouped the phages into 17 clusters and 39 subclusters (2). Of these, the B1 subcluster of mycobacteriophages is one of the most populous, accounting for approximately 10% of the mycobacteriophage genome sequences found in GenBank. We report here the genome sequences of five novel B1 subcluster phages.
The phages were isolated and characterized by students in the Freshman Research Initiative course at Brigham Young University (BYU). The program is part of the Howard Hughes Medical Institute Science Education Alliance's Phage Hunters Advancing Genomics and Evolutionary Sciences Program (SEA-PHAGES). In the course, each student isolated and plaque purified a single mycobacteriophage using M. smegmatis mc2155 as the host. After three rounds of plaque purification, followed by amplification to a high titer, the genomic DNA was isolated (Wizard DNA Clean-Up System, Promega Corp., Madison, WI) and submitted for 454 pyrosequencing (Roche) at the BYU DNA Sequencing Center. Students in the program named the phages and annotated the genome sequences. The teaching assistants and faculty checked the annotated genome sequences prior to their submission to GenBank.
Each genome had at least 65-fold coverage (average, 119.8-fold). Assembly was conducted using GS de novo assembler 2.6 (Newbler; Roche) and verified with Consed version 19 (3). Base one was called by alignment with the other B1 phages. The genome sequences were annotated using GeneMark coding potential maps with M. tuberculosis CDC 1551 as a reference strain (4) and DNA Master software (J. G. Lawrence lab [http://cobamide2.bio.pitt.edu]), which integrates Glimmer 3.02, GeneMark, BLAST, Shine-Dalgarno (SD) position-weighted scores, and general annotation tools.
The sizes (average, 68,800 bp) and organization of the genomes (approximately 100 open reading frames) are consistent with those of other B1 subcluster mycobacteriophages catalogued in GenBank to date. As with other B1 subcluster phages, all five phages reported here are Siphoviridae with double-stranded DNA (dsDNA) circularly permuted genomes. The analysis of an increasing data set of phage genomes provides insights not only into the diversity of bacteriophages and phage-host interactions but also into their evolution. Additional information on these phage and their genomes is available at http://www.phagesdb.org. Subsequent reports will describe the specific attributes and comparative analyses of the phages announced herein.

Nucleotide sequence accession numbers.

The GenBank accession numbers and pertinent traits of the five genome sequences are reported in Table 1.
TABLE 1
TABLE 1 Data for five new B1 subcluster mycobacteriophage genome sequences
PhageGenBank accession no.Length (bp)% G+CNo. of ORFsYear isolated
AlexJX64910068,91066.521052009
GyaradJX64909968,00466.491002009
NachoJX64909869,32166.561042011
PigletJX64909768,99266.541012011
SerpentineJX64909668,88466.511042010

ACKNOWLEDGMENTS

We recognize the contributions of the students from each cohort of the BYU Phage Hunters program. We thank Graham F. Hatfull, Lucia P. Barker, Kevin W. Bradley, Deborah Jacobs-Sera, David J. Asai, Tuajuanda C. Jordan, Welkin H. Pope, Daniel A. Russell, and Steven G. Cresawn for their technical support and program administration.
Funding was provided by the Howard Hughes Medical Institute and by the Department of Microbiology and Molecular Biology and the College of Life Sciences at Brigham Young University.

REFERENCES

1.
Jacobs-Sera D, Marinelli LJ, Bowman C, Broussard GW, Guerrero Bustamante C, Boyle MM, Petrova ZO, Dedrick RM, Pope WH, Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) Program, Modlin RL, Hendrix RW, and Hatfull GF. 2012. On the nature of mycobacteriophage diversity and host preference. Virology 434:187–201.
2.
Hatfull GF, Jacobs-Sera D, Lawrence JG, Pope WH, Russell DA, Ko CC, Weber RJ, Patel MC, Germane KL, Edgar RH, Hoyte NN, Bowman CA, Tantoco AT, Paladin EC, Myers MS, Smith AL, Grace MS, Pham TT, O'Brien MB, Vogelsberger AM, Hryckowian AJ, Wynalek JL, Donis-Keller H, Bogel MW, Peebles CL, Cresawn SG, and Hendrix RW. 2010. Comparative genomic analysis of 60 mycobacteriophage genomes: genome clustering, gene acquisition, and gene size. J. Mol. Biol 397:119–143.
3.
Gordon D. 2003. Viewing and editing assembled sequences using Consed. Curr. Protoc. Bioinformatics 2:11.2.1–11.2.43.
4.
Lukashin AV and Borodovsky M. 1998. GeneMark.hmm: new solutions for gene finding. Nucleic Acids Res. 26:1107–1115.

Information & Contributors

Information

Published In

cover image Genome Announcements
Genome Announcements
Volume 1Number 626 December 2013
eLocator: 10.1128/genomea.00968-13

History

Received: 15 October 2013
Accepted: 27 October 2013
Published online: 27 November 2013

Contributors

Authors

Donald P. Breakwell
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
E. Zane Barrus
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Alex B. Benedict
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Alicia K. Brighton
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Joshua N. B. Fisher
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Adam V. Gardner
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Brittany J. Kartchner
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Kara C. Ladle
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Bryce L. Lunt
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Present address: Bryce L. Lunt, Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Bryan D. Merrill
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
John D. Morrell
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Sandra H. Burnett
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA
Julianne H. Grose
Department of Microbiology and Molecular Biology, Brigham Young University, Provo, Utah, USA

Notes

Address correspondence to Donald P. Breakwell, [email protected], or Julianne H. Grose, [email protected].

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