Novel Arthrobacter sulfureus phage Emotion shares greatest structural gene similarity to Arthrobacter AZ cluster phages

Pearl Doan, Conner Kidd, Asefeh Tarighat, Lisa Yan, Michelle Zorawik, Dominic Garza, Juliet Stephenson, Krisanavane Reddi, Amanda C Freise

In spite of existing guidelines for phage clustering, some phages are left temporarily unclustered and may require more in-depth examination. For example, novel A. sulfureus phage Emotion remained unclustered for some time before the final decision was made to place it in Cluster AZ, which made us curious about its relationship to other phages, especially since it was isolated on a different host than any other Cluster AZ phage. We decided to examine overall genomic similarity and structural and non-structural gene similarity between novel phage Emotion and other phages, since structural proteins are involved in the recognition and attachment of bacteriophages to host bacteria and are both highly diverse and host-specific.

Phages chosen for comparison to Emotion included phages isolated on host A. sulfureus; phages with low BLASTp e-values when Emotion’s genome was used as a query; and member phages of phams (gene families of related sequences) with <50 members. The phages chosen were from Arthrobacter clusters AZ and FP and Microbacterium cluster EH. Phamerator maps, dot plots, and gene content similarity (GCS) analyses were conducted to quantitatively and qualitatively evaluate the similarities and differences between Emotion and these other phages. As expected based on its cluster designation, Emotion was determined to be most similar to Cluster AZ phages based on an average pairwise GCS (~50%) above the 35% GCS clustering threshold. Similarly, Emotion had higher amino acid sequence identity to Cluster AZ compared to Clusters EH and FP based on whole-genome amino acid dotplots. Phylogenetic trees were used to compare several of Emotion’s structural and non-structural genes to those found in Cluster AZ, EH, and FP phages, showing that structural genes shared hypothetical evolutionary relationships between Emotion and Cluster AZ phages.

These results of high GCS and shared genes corroborate the clustering of Emotion in AZ. Notably, FP phages share some non-structural genes with Emotion with greater sequence conservation than AZ phages. However, findings could be obscured by Emotion’s large number of uncharacterized genes. Overall, these findings provide a model for analyzing the complex relationships of unclustered phages through similarity in specific structural and non-structural genes.