Complete Genome Sequences of Six BI Cluster Streptomyces Bacteriophages, HotFries, Moozy, Rainydai, RavenPuff, Scap1, and SenditCS

Abstract

Six double-stranded DNA Streptomyces bacteriophages, HotFries, Moozy, RavenPuff, Scap1, Rainydai, and SenditCS, were isolated using the phytopathogen Streptomyces scabiei as a host. These phages have been identified as Siphoviridae and members of cluster BI by genomic analysis.

A Novel Genus of Actinobacterial Tectiviridae

“…Both phages displayed a fairly limited host range among the tested strains, being able to infect only S. mirabilis NRRL B-2400 at an efficiency-of-plating (EOP) of approximately 20, in addition to the isolation host S. scabiei RL-34 (Table S2). Phages infecting only S. mirabilis and S. scabiei among this group of tested strains have been reported previously for Siphoviridae phages infecting S. scabiei [21], suggesting that S. mirabilis and S. scabiei share important properties for phage infection that are absent in the closely related S. diastatochromogenes [59].…”

“…Putative WheeHeim and Forthebois lysogens were isolated by spotting diluted crude lysate on NA + overlaid with 250 µL of a 48 h culture of S. scabiei RL-34 in trypticase soy soft agar incubated for 96 h at 30 • C. Cells were isolated from a zone of clearing and streak purified three times. A resulting colony was then used to inoculate 3 mL NB + with 0.05% polyethylene glycol (PEG 8000) and incubated for 48 h at 30 • C. To test for the presence of released phage, 500 µL of the culture was centrifuged for 1 min at 11,700× g, and the supernatant was serially diluted and 3 µL spotted on NA + overlaid with 250 µL of a 48 h culture of S. scabiei RL-34 in trypticase soy soft agar incubated for 48 h at 30 • C. To test for superinfection immunity, crude lysate stocks of Streptomyces phages Scap1 [21], Forthebois, WheeHeim, and S. scabiei (Forthebois) lysogen supernatant were serially diluted and 3 µL spotted on NA + overlaid with 250 µL of a culture of S. scabiei RL-34 (Forthebois) lysogens in trypticase soy soft agar incubated for 96 h at 30 • C with daily examination.…”

“…Our results show that, after purification, S. scabiei cells isolated from apparent colonies within turbid plaques were able to induce plaque formation on a S. scabiei RL-34 lawn, indicating that they are releasing phage (Figures 3a and S2). Furthermore, lawns of these putative S. scabiei lysogens could be infected by Streptomyces phage Scap1 (a Siphoviridae infecting S. scabiei [21]), but not by WheeHeim or Forthebois, indicating that lysogens exhibited superinfection exclusion (Figure 3b). Bacillus-infecting tectiviruses have been reported to use the host's LexA repressor to activate the lytic cycle by binding at multiple promoters [15,74], but no putative LexA-binding sites could be detected on genome-wide scans of Streptomyces phages WheeHeim and Forthebois genome sequences.…”

“…Rhodococcus phage Toil, a phage infecting Rhodococcus opacus, was recently characterized and reported as the first tectivirus capable of infecting Actinobacteria [17]. Here we report the isolation, characterization and genome sequencing of two new tectiviruses as part of an ongoing effort to characterize phages infecting the causative agent of potato scab, Streptomyces scabiei, within the Science Education Alliance-Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) and STEM BUILD at UMBC programs [18][19][20][21]. S. scabiei is best known for its worldwide economic impact on potato crops [22,23], but it has a broad host range that encompasses carrot and radish taproot crops, as well as many other dicots and monocots [22].…”

“…Because there is no known mechanism for effective disease suppression after initial infestation, multiple management strategies and approaches have been explored for the control of S. scabiei [22], including the effective use of virulent bacteriophages [25]. The novel tectiviruses reported in this work (Streptomyces phage WheeHeim and Streptomyces phage Forthebois) were isolated on S. scabiei and expand the existing collection of Caudovirales phages active against this important pathogen [21,[25][26][27][28]. These two phages share morphological features, genomic structure and moderate sequence similarity with Rhodococcus phage Toil.…”