Complete Genome Sequences of Nine Cluster A Mycobacteriophages: AgentM, Ajay, Aragog, Archetta, ForGetIt, Koko, Ph8s, Phlorence, and Wilkins

Background Bacteriophages are ubiquitous, highly diverse, and relatively understudied. Growing interest in phage therapy has underscored the importance of isolating and characterizing novel bacteriophages. The Science Education Alliance-Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program aims to address this need by involving undergraduates around the world in authentic research. Materials and Methods Nine novel mycobacteriophages - AgentM, Ajay, Aragog, Archetta, ForGetIt, Koko, Ph8s, Phlorence, and Wilkins - were isolated from soil samples in southern California using host Mycobacterium smegmatis mc2155. Each purified phage was characterized using transmission electron microscopy and genome sequencing and annotation. Results All nine bacteriophages were placed into mycobacteriophage Cluster A based on nucleotide similarity with other phages. The average genome length of all nine phages was 50,706 bp. On average, each phage had 87 total coding genes and GC content between 60-63%, consistent with that of other Cluster A phages. Transmission electron microscopy of phage particles revealed they had icosahedral heads and long, flexible tails, consistent with that of the Siphoviridae family. Plaque morphology and genome analysis confirms the nine novel phages are temperate as expected of Cluster A phages. Conclusions AgentM, Ajay, Aragog, Archetta, ForGetIt, Koko, Ph8s, Phlorence, and Wilkins are all mycobacteriophages that belong to the Siphoviridae family. Comparative genomic analyses revealed genetic mosaicism and diversity among these Cluster A phages. The discovery of these novel phages expands on the existing library of mycobacteriophage genomes.