an-optimized-enrichment-technique-for-isolation-arthrobacter

An Optimized Enrichment Technique for the Isolation of <em>Arthrobacter</em> Bacteriophage Species from Soil Sample Isolates

Bacteriophage isolation from environmental samples has been performed for decades using principles set forth by pioneers in microbiology. The isolation of ...

University of the Sciences

Mycobacteriophages represent a genetically diverse group of viruses that infect mycobacterial hosts. Although more than 80 genomes have been sequenced, these still poorly represent the likely diversity of the broader population of phages that can infect the host, Mycobacterium smegmatis mc(2)155. We describe here a newly discovered phage, Marvin, which is a singleton phage, having no previously identified close relatives. The 65,100-bp genome contains 107 predicted protein-coding genes arranged in a noncanonical genomic architecture in which a subset of the minor tail protein genes are displaced about 20 kbp from their typical location, situated among nonstructural genes anticipated to be expressed early in lytic growth. Marvin is not temperate, and stable lysogens cannot be recovered from infections, although the presence of a putative xis gene suggests that Marvin could be a relatively recent derivative of a temperate parent. The Marvin genome is replete with novel genes not present in other mycobacteriophage genomes, and although most are of unknown function, the presence of amidoligase and glutamine amidotransferase genes suggests intriguing possibilities for the interactions of Marvin with its mycobacterial hosts.

Mycobacteriophage Marvin: a new singleton phage with an unusual genome organization.

In this work, a method was developed to study the structural proteome of mycobacteriophage Marvin, a recent isolate from soil with 107 predicted coding sequences. This prototype method was applied for semi-quantitative analysis of the composition of this mycobacteriophage virion using ion mobility spectrometry and data-independent acquisition (MS(E)-IMS). MS(E)-IMS was compared to a more conventional proteomics technique employing mass spectrometry with a data-dependent acquisition strategy. MS(E)-IMS provided broad coverage of the virion proteome and high sequence coverage for individual proteins. This shotgun method does not depend on the limited sensitivity of visualization of protein bands by staining reagents inherent in gel-based methods. The method is comprehensive, provides high sequence coverage and is proposed as a particularly efficient method for the study of bacteriophage proteomes.

Data-independent acquisition (MSE) with ion mobility provides a systematic method for analysis of a bacteriophage structural proteome.

Mycobacteriophages - viruses of mycobacterial hosts - are genetically diverse but morphologically are all classified in the Caudovirales with double-stranded DNA and tails. We describe here a group of five closely related mycobacteriophages - Corndog, Catdawg, Dylan, Firecracker, and YungJamal - designated as Cluster O with long flexible tails but with unusual prolate capsids. Proteomic analysis of phage Corndog particles, Catdawg particles, and Corndog-infected cells confirms expression of half of the predicted gene products and indicates a non-canonical mechanism for translation of the Corndog tape measure protein. Bioinformatic analysis identifies 8-9 strongly predicted SigA promoters and all five Cluster O genomes contain more than 30 copies of a 17 bp repeat sequence with dyad symmetry located throughout the genomes. Comparison of the Cluster O phages provides insights into phage genome evolution including the processes of gene flux by horizontal genetic exchange.

Trevor Cross

Biology Department

University of the Sciences

An Optimized Enrichment Technique for the Isolation of Arthrobacter Bacteriophage Species from Soil Sample Isolates

Trevor Cross

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Biology Department

University of the Sciences

An Optimized Enrichment Technique for the Isolation of Arthrobacter Bacteriophage Species from Soil Sample Isolates

Biology Department