From unique killers to a jumbo genome - isolation and characterization of phages that infect the plant pathogen, Agrobacterium tumefaciens

Bacteriophages and their lytic peptides can protect plants from phytopathogens such as Agrobacterium tumefaciens. To better understand mechanisms of phagemediated host killing, we isolated and characterized five lytic bacteriophages with activity against A. tumefaciens C58. These phages come in different shapes and sizes--from T7-like phages with podoviral morphology and isometric heads to T4-like phages with myoviral morphology and a contractile tail--and exhibit varying host ranges and killing efficiencies. The smallest Agrobacterium phages are phiKMVlike phages in the T7 superfamily that are efficient at killing their hosts. Their lethality can be attributed to their expression of a unique endolysin, called Phage Peptidoglycan Hydrolase (PPH). The atypical domain structure of PPH, along with the absence of obvious accessory proteins, suggest PPH may function independently to mediate host cell lysis. Contrary to the narrow host range of the phage, expression of PPH from an inducible promoter inhibits cell growth and blocks cell division in a broad range of bacteria including Agrobacterium, Sinorhizobium, and Escherichia strains. Another member of the Podoviridae family, Atu_ph08, carries remnants of a lysogen and shares 60.2% identity with Agrobacterium genomospecies 3. The T4-like phages in our collection are not potent killers. Atu_ph04 is unique T4-like phage that is similar to a group of rhizophages. The largest Agrobacterium phage, Atu_ph07, has a head diameter of 146 nm, an extended tail length of 136 nm, and a genome of 490 kbp. Our results indicate a high degree of morphological and genomic diversity and also suggest novel mechanisms of host cell killing remain to be uncovered.