Rhizobiophages are the group of bacteriophages that infect rhizobia. The rhizobia constitute a bacterial group that includes members of several different genera and are capable of nodulating legume plant roots, and providing a source of fixed nitrogen for the plant. Rhizobiophages can play a vital role in the Rhizobium-legume symbiosis by altering the population dynamics of the rhizobia present in the rhizosphere, which can be used in agriculture to improve the efficacy of commercial Rhizobium inoculants and mitigate the Rhizobium competition problem. As a prerequisite for the application of such technology, a thorough understanding of rhizobiophage biology is required. Isolation of rhizobiophages from soil samples obtained from Alberta, Saskatchewan, Ontario and British Columbia was performed using several different strains of rhizobia as trapping hosts. The isolated phages were characterized using host range, transmission electron microscopy, protein profiles and genomic characterization. All phage isolates were characterized as tailed phages belonging to the Order Caudovirales, while further classification allowed us to place them in families Siphoviridae, Myoviridae and Podoviridae. Depending on the host range, morphotype and trapping host, several phages were selected and detailed genome characterization was performed using whole genome sequencing. Five rhizobiophage genomes were sequenced to finished state (vB_RleM_P10VF, vB_RglS_P106B, vB_RleM_PPF1, vB_MloP_Lo5R7ANS, and vB_MloP_Cp1R7ANS-C2) and annotated. The complete genome sequences of vB_RglS_P106B (KF977490), vB_RleM_PPF1 (KJ746502), vB_RleM_P10VF (KM199770) and vB_MloP_Lo5R7ANS (KM199771) are publically available at the database of National Center of Biological Information (NCBI). The genome of vB_MloP_Cp1R7ANS-C2 will be submitted to the NCBI in the near future. The integration of temperate phage vB_RleM_PPF1 into its bacterial host R. leguminosarum
F1 was also studied. The site-specific recombination system of the phage targets an integration site that lies within a putative tRNA-Pro (CGG) gene in R. leguminosarum
F1. Upon integration, the phage is capable of restoring the disrupted tRNA gene, owing to the 50 bp homologous sequence (att core region) it shares with its rhizobial host. To
develop a phage-based inoculant technology, the phages of Rhizobium leguminosarum with broadest possible host range were selected and tested in nodulation competition assays. The presence of phages altered the nodule occupancy by phage-sensitive and phage-resistant strains of rhizobia, increasing the efficacy of nodulation by phageresistant
rhizobia under controlled environmental conditions.