Characterization of the temperate phage vB_RleM_PPF1 and its site-specific integration into the Rhizobium leguminosarum F1 genome

Abstract

Bacteriophages may play an important role in regulating population size and diversity of the root nodule symbiont Rhizobium leguminosarum, as well as participating in horizontal gene transfer. Although phages that infect this species have been isolated in the past, our knowledge of their molecular biology, and especially of genome composition, is extremely limited, and this lack of information impacts on the ability to assess phage population dynamics and limits potential agricultural applications of rhizobiophages. To help address this deficit in available sequence and biological information, the complete genome sequence of the Myoviridae temperate phage PPF1 that infects R. leguminosarum biovar viciae strain F1 was determined. The genome is 54,506 bp in length with an average G+C content of 61.9 %. The genome contains 94 putative open reading frames (ORFs) and 74.5 % of these predicted ORFs share homology at the protein level with previously reported sequences in the database. However, putative functions could only be assigned to 25.5 % (24 ORFs) of the predicted genes. PPF1 was capable of efficiently lysogenizing its rhizobial host R. leguminosarum F1. 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 genome. Phage PPF1 is the first temperate phage infecting members of the genus Rhizobium for which a complete genome sequence, as well as other biological data such as the integration site, is available.

Ecological and Evolutionary Benefits of Temperate Phage: What Does or Doesn't Kill You Makes You Stronger

“…While temperate phages are most well-studied in the context of bacterial pathogens, they are widespread in environmental microbial communities [5,69] where www.advancedsciencenews.com www.bioessays-journal.com their role is yet to be fully explored. However this is an exciting time for the study of temperate phages.…”

“…Given the ecological importance of temperate phage-bacteria interactions in a wide range of environments and the contribution of temperate phages to bacterial genome evolution, there is clearly an urgent need to better understand the ecology and evolution of these relationships. While temperate phages are most well-studied in the context of bacterial pathogens, they are widespread in environmental microbial communities [5,69] where www.advancedsciencenews.com www.bioessays-journal.com their role is yet to be fully explored. However this is an exciting time for the study of temperate phages.…”

Molecular and Biological Characterization of Ralstonia Phage RsoM1USA, a New Species of P2virus, Isolated in the United States

“…(2) Seven ORFs in phage RsoM1USA annotated as hypothetical/other proteins have no corresponding ORFs in phage RSA1. (3) Myoviruses are known to integrate into bacterial genomes mediated by a portion of bacterial tRNA, an att site, as in the cases of phages RSA1 and RSY 1 of R. solanacearum (Fujiwara et al, 2008; Askora et al, 2017), phage 16-3 of Rhizobium meliloti (Blaha et al, 2004), and phage vB_RleM_PPF1 on Rhizobium leguminosarum (Halmillawewa et al, 2016). We confirmed experimentally that phage RsoM1USA uses the 3′ 45-base portion of the threonine tRNA (TGT) gene as its att site (Figure 5), which is different from RSA1 which uses the 3′ 45-base of the arginine tRNA (CCG) gene (Fujiwara et al, 2008).…”

Introducing a Novel, Broad Host Range Temperate Phage Family Infecting Rhizobium leguminosarum and Beyond

“…In R. leguminosarum strains which lack the prophage this 12 bp motif is present within a tRNA-Leu indicating that this is the site of prophage integration (attB; see Supplementary Figure 2 ). tRNA sequences are used by a range of other mobile genetic elements for site-specific integration ( Reiter et al, 1989 ), including other rhizophages ( Semsey et al, 2002 ; Halmillawewa et al, 2016 ), retrotransposons ( Mularoni et al, 2012 ), and cyanopodoviruses ( Sullivan et al, 2005 ). This includes closely related pelagibacter viruses in the HTVC019Pvirus genus ( Zhao et al, 2019 ).…”