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Procurement_Operations_Manual_revised_1-6-6

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6elkin H Pope’*, Charles A Bowman’*, Daniel A Russell’*, Deborah Jacobs-Sera’, David J Asai7, Steven G Cresawn3, William R Jacobs Jr4, Roger w Hendrix’

, Jeffrey G Lawrence1, Graham F Hatfull’*, Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science, Phage Hunters Integra Procurement_Operations_Manual_revised_1-6-6

ting Research and Education, Mycobacterial Genetics Course‘Department of Biological Sciences, University of Pittsburgh, Pittsburgh, United States; 2Ho

Procurement_Operations_Manual_revised_1-6-6

ward Hughes Medical Institute, Chevy Chase, United States; department of Biology, James Madison University, Harrisonburg, United States; department of

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6ally to this workCompeting interests: The authors declare that noAbstract The bacteriophage population is large, dynamic, ancient, and genetically div

erse. Limited genomic information shows that phage genomes are mosaic, and the genetic architecture of phage populations remains ill-defined. To under Procurement_Operations_Manual_revised_1-6-6

stand the population structure of phages infecting a single host strain, we isolated, sequenced, and compared 627 phages of Mycobacterium smegmatrs. T

Procurement_Operations_Manual_revised_1-6-6

heir genetic diversity is considerable, and there are 28 distinct genomic types (dusters) with related nucleotide sequences. However, amino acid seque

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6ersity, albeit with uneven representation of different phages. Furthermore, rarefaction analysis shows that the mycobacteriophage population is not do

sed, and there is a constant influx of genes from other sources. Phage isolation and analysis was performed by a large consortium of academic institut Procurement_Operations_Manual_revised_1-6-6

ions, illustrating the substantial benefits of a disseminated, structured program involving large numbers of freshman undergraduates in scientific dis

Procurement_Operations_Manual_revised_1-6-6

covery.DOI: 10J5M/el.iicC6'J16(»1competing interests exist.Funding: See page 61Received: 11 January 2015 Accepted: 19 March 2015 Published: 28 April 2

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6eative Commons Attribution license, which permits unrestricted use and redistribution provided that the original author and source arc credited.Introd

uctionBacteriophages are the dark matter of the biological universe, forming a vast, ancient, dynamic, and genetically diverse population, replete wit Procurement_Operations_Manual_revised_1-6-6

h genes of unknown function (Pedulla Ct al., 2003). Phages are the most abundant organisms in the biosphere, and the ~10:” tailed phage particles part

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icipate in ~10ZJ infections per second on a global scale, with the entire population turning over every few days (Suttle, 2007). The population is not

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6sing that bacteriophages are genetically highly diverse, although their comparative genomics has lagged behind that of other microbes, largely due to

the lack of individual isolates for genomic analyses (Hatfull and Hendrix, 2011). To date, there are approximately 2000 completely sequenced bacteriop Procurement_Operations_Manual_revised_1-6-6

hage genomes in the GenBank database, a small number relative to the more than 30,000 sequenced prokaryotic genomes (http://www.ncbi.nlm.nih.gov/genom

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e/browse/), in spite of phage genomes being only 1-5% of the size of their host genomes.Double-stranded DNA tailed phages are proposed to have evolved

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6e06416. DO:-10 7554/el.ife 064161 of 65eLIFE Research articleGenomics and evolutionary biology I Microbiology and infectious diseaseeLife digest Virus

es are unable to replicate independently. To generate copies of itself, a virus must instead invade a target cell and commandeer that cell's replicati Procurement_Operations_Manual_revised_1-6-6

on machinery. Different viruses are able to invade different types of cell, and a group of viruses known as bacteriophages (or phages for short) repli

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cate within bacteria. The enormous number and diversity of phages in the world means that they play an important role in virtually every ecosystem.Des

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6ntain their genetic information in the form of strands of DNA. Using genetic sequencing to find out where and how different genes are encoded in the D

NA can reveal information about how different viruses are related to each other. These relationships are particularly complicated in phages, as they c Procurement_Operations_Manual_revised_1-6-6

an exchange genes with other viruses and microbes.Previous studies comparing the genomes—the complete DNA sequence—of reasonably small numbers of phag

Procurement_Operations_Manual_revised_1-6-6

es that infect the Mycobacterium group of bacteria have found that the phages can be sorted into ‘clusters' based on similarities in their genes and w

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6ed. Are the clusters separate, or do they form a 'continuum' with different genes and DNA sequences shared between different clusters?Here, Pope. Bowm

an, Russell et al. compare the individual genomes of 627 bacteriophages that infect the bacterial species Mycobacterium smegmatis. This is by far the Procurement_Operations_Manual_revised_1-6-6

largest number of phage genomes analyzed from a single host species. The large number of genomes analyzed allowed a much clearer understanding of the

Procurement_Operations_Manual_revised_1-6-6

complexity and diversity of these phages to be obtained. The isolation, sequencing and analysis of the hundreds of M. smegmatis bacteriophage genomes

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6 Science (SEA PHAGES) program. This enabled thousands of undergraduate students from different institutions to contribute to the phage discovery and s

equencing project, and co-author the report. SEA-PHAGES therefore shows that it is possible to successfully incorporate genuine scientific research in Procurement_Operations_Manual_revised_1-6-6

to an undergraduate course, and that doing so can benefit both the students and researchers involved.The results show that while the genomes could be

Procurement_Operations_Manual_revised_1-6-6

categorized into 28 clusters, the genomes are not completely unrelated. Instead, a spread of diversity is seen, as genes and groups of genes are share

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6 acquires genes from other microorganisms and viruses.DOI: 10.7554/ebfe 06416002Phage genomes are typified by their mosaic architectures generated by

gene loss and gain through horizontal genetic exchange; however, the parameters influencing access to the common gene pool are numerous and likely inc Procurement_Operations_Manual_revised_1-6-6

lude host range, genome size, replication mode, and life style (temperate vs lytic). Migration to new hosts is probably common, but is affected by loc

Procurement_Operations_Manual_revised_1-6-6

al host diversity and mutation rates, as well as resistance mechanisms such as receptor availability, restriction. CRISPRs, and abortive infection sys

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6particularly among virion structural genes—and by size limits of DNA packaging (Juhala ct al., 2000; Hatfull and Hendrix. 2011).We have previously des

cribed comparative analyses of modest numbers of mycobacteriophages and shown that they can be sorted by nucleotide sequence and gene content comparis Procurement_Operations_Manual_revised_1-6-6

ons into groups of closely related genomes referred to as 'dusters' (designated Cluster A, B, c, etc.); phages without any close relatives are referre

Procurement_Operations_Manual_revised_1-6-6

d to as ’singletons’. Some of the clusters can be further divided into subclusters (e g., Subcluster Al, A2, A3, etc.) according to nucleotide sequenc

9lifesciences.orgRESEARCH ARTICLEaWhole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversityWe

Procurement_Operations_Manual_revised_1-6-6 as assemblages of modules, many of which are single genes (Pedulla et al.. 2003). Each mycobacteriophage cluster has features particular to that dust

er (e.g., regulatory systems, repeated sequences, tRNA genes, etc. I Pope Ct al., 2011a, 2011b, 2013, 2014a, 2014b]), butPope et al. elife 2015;4:e064 Procurement_Operations_Manual_revised_1-6-6

16. DO: 10.7554/eUfe.064162 of 65eLIFE Research articleGenomics and evolutionary biology I Microbiology and infectious disease

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