Complete genome of the broad-host-range Erwinia amylovora phage phiEa21-4 and its relationship to Salmonella phage felix O1

Abstract

The first complete genome sequence for a myoviridal bacteriophage, PhiEa21-4, infecting Erwinia amylovora, Erwinia pyrifoliae, and Pantoea agglomerans strains has been determined. The unique sequence of this terminally redundant, circularly permuted genome is 84,576 bp. The PhiEa21-4 genome has a GC content of 43.8% and contains 117 putative protein-coding genes and 26 tRNA genes. PhiEa21-4 is the first phage in which a precisely conserved rho-independent terminator has been found dispersed throughout the genome, with 24 copies in all. Also notable in the PhiEa21-4 genome are the presence of tRNAs with six- and nine-base anticodon loops, the absence of a small packaging terminase subunit, and the presence of nadV, a principle component of the NAD(+) salvage pathway, which has been found in only a few phage genomes to date. PhiEa21-4 is the first reported Felix O1-like phage genome; 56% of the predicted PhiEa21-4 proteins share homology with those of the Salmonella phage. Apart from this similarity to Felix O1, the PhiEa21-4 genome appears to be substantially different, both globally and locally, from previously reported sequences. A total of 43 of the 117 genes are unique to PhiEa21-4, and 32 of the Felix O1-like genes do not appear in any phage genome sequences other than PhiEa21-4 and Felix O1. N-terminal sequencing and matrix-assisted laser desorption ionization-time of flight analysis resulted in the identification of five PhiEa21-4 genes coding for virion structural proteins, including the major capsid protein.

FIG. 1.

TEM image of E. amylovora phage ΦEa21-4 in the uncontracted (left) and contracted (right) states. Bar, 50 nm.

FIG. 2.

Alignment of the annotated ΦEa21-4 and Felix O1 genomes using Mauve. Both gene maps begin with the rIIA gene and are measured against a ruler in kilobases. Genes transcribed from the negative strand are displaced downward within each gene map. The degree of sequence similarity between aligned regions is indicated by the height of the similarity profile. The tRNA genes are indicated by the filled gene blocks. In ΦEa21-4 these span bases 21823 through 25229; the 22 Felix O1 tRNA genes are slightly dispersed, between bases 23699 and 30180.

FIG. 3.

Sequence logo for the ΦEa21-4 conserved terminator sequence.

FIG. 4.

ΦEa21-4 tRNA gene cluster. The tRNAScan_SE classifications and predicted tRNA anticodons are indicated. Predicted pseudogenes are marked with an asterisk. The tRNAs with unusual anticodon loops are labeled in red.