DNA Packaging and Genomics of the Salmonella 9NA-Like Phages

We present the genome sequences of tailed phages Sasha, Sergei, and Solent. These phages, along with phages 9NA, FSL_SP-062, and FSL_SP-069 and the more distantly related phage PmiS-Isfahan, have similarly sized genomes of between 52 and 57 kbp in length that are largely syntenic. Their genomes also show substantial genome mosaicism relative to one another, which is common within tailed phage clusters. Their gene content ranges from 80 to 99 predicted genes, of which 40 are common to all seven and form the core genome, which includes all identifiable virion assembly and DNA replication genes. The total number of gene types (pangenome) in the seven phages is 176, and 59 of these are unique to individual phages. Their core genomes are much more closely related to one another than to the genome of any other known phage, and they comprise a well-defined cluster within the family To begin to characterize this group of phages in more experimental detail, we identified the genes that encode the major virion proteins and examined the DNA packaging of the prototypic member, phage 9NA. We show that it uses a site-directed headful packaging mechanism that results in virion chromosomes that are circularly permuted and about 13% terminally redundant. We also show that its packaging series initiates with double-stranded DNA cleavages that are scattered across a 170-bp region and that its headful measuring device has a precision of ±1.8%. The 9NA-like phages are clearly highly related to each other but are not closely related to any other known phage type. This work describes the genomes of three new 9NA-like phages and the results of experimental analysis of the proteome of the 9NA virion and DNA packaging into the 9NA phage head. There is increasing interest in the biology of phages because of their potential for use as antibacterial agents and for their ecological roles in bacterial communities. 9NA-like phages that infect two bacterial genera have been identified to date, and related phages infecting additional Gram-negative bacterial hosts are likely to be found in the future. This work provides a foundation for the study of these phages, which will facilitate their study and potential use.