Introducing Casbah, Kronus, and MmasiCarm, Members of the Mycobacteriophage Subcluster B3

Abstract

Background: As part of a large science education effort, bacteriophages that lyse Mycobacterium smegmatis mc²155 continue to be discovered.

Materials and methods: Phages were isolated from soil samples from urban sites in the Northeastern United States. Their genomes were sequenced, assembled, and bioinformatically compared.

Results: Three lytic siphoviruses belonging to subcluster B3 with high similarity to each other and other B3 mycobacteriophages were isolated. These phages contain double-stranded DNA genomes (68,754 to 69,495 bp) with high GC content (67.4-67.5%) and 102-104 putative protein coding genes. Notable features include a HicA-like toxin and 33 genes exclusive to subcluster B3. One phage had an intein in its terminase sequence.

Conclusions: Genomic analyses of these phages provide insights into genome evolution and horizontal gene transfer (HGT). The networks for HGT are apparently vast and gene specific. Interestingly, a number of genes are found in both B3 and Gordonia DR phages.

Keywords: HicA-like toxin; comparative genomics; functional annotation; horizontal gene transfer; intein; mycobacteriophages.

FIG. 1.

Representative brightfield transmission electron microscopy image of Casbah stained with uranyl acetate solution. Images were taken at 50,000× magnification using a Hitachi 7800 transmission electron microscope.

FIG. 2.

Genome map of Kronus, chosen because it has the longest genome. The ruler shows genome length (in kilobases) with forward and reverse genes shown above and below the ruler, respectively. Function or putative functions are listed above genes. Map created using Phamerator. Gene boxes are colored based on pham membership, as determined by amino acid alignment using the PhaMMseqs pipeline.

FIG. 3.

Comparative genome maps of the B3 mycobacteriophages: Casbah, Kronus, and MmasiCarm. Gene boxes are colored based on pham membership, as determined by amino acid alignment using the PhaMMseqs pipeline. Map created using Phamerator. The color spectrum between two adjacent sequences reflects nucleotide sequence similarity as measured by pairwise local alignment using BLASTn “align two sequences”, with lavender being the most similar (E = 0) and red being the least similar (E = 10⁻⁵, set as the minimal threshold). A lack of shading indicates that there is the minimal similarity threshold that has not been met (E score greater than 10⁻⁵).