Genomic and biological characterization of a new member of the genus Phikmvvirus infecting phytopathogenic Ralstonia bacteria

Abstract

Bacterial wilt caused by Ralstonia spp., soil-borne Gram-negative bacteria, is considered one of the most important plant diseases in tropical and subtropical regions of the world. A large number of bacteriophages capable of lysing or physiologically reprogramming cells of Ralstonia spp. have been reported in Asia. Despite the potential use of these organisms in the management of bacterial wilt, information on viruses that infect Ralstonia spp. is nonexistent in the Americas. We isolated a virus that infects Ralstonia spp. from a soil sample in the state of Minas Gerais, Brazil. Microscopy and genomic and phylogenetic analysis allowed us to classify the virus as a member of the family Podoviridae, genus Phikmvvirus. In spite of its relationship to Ralstonia virus RSB3, an Asian isolate, genomic and biological characteristics showed that the virus isolated in Brazil, tentatively named "Ralstonia virus phiAP1" (phiAP1), belongs to a new species. phiAP1 has EPS depolymerase activity and contains two putative virion-associated peptidoglycan hydrolases (VAPGHs), which reveals a robust mechanism of pathogenesis. Furthermore, phiAP1 specifically infects Ralstonia solanacearum, R. pseudosolanacearum and R. syzygii, causing cell lysis, but it was not able to infect thirteen other bacteria that were tested. Together, these characteristics highlight the biotechnological potential of this virus for the management of bacterial wilt.

Genomic Analysis of the First European Bacteriophages with Depolymerase Activity and Biocontrol Efficacy against the Phytopathogen Ralstonia solanacearum

“…Most of the endolysins reported so far are lytic transglycosylases [74]. Two putative virion-associated peptidoglycan hydrolases have been recently reported in the American R. solanacearum phiAP1 phage, being proposed to act as exolysins, which locally pierce peptidoglycan to eject the phage DNA into the host, in the first steps of the infection [48], as already reported in other phages of the Autographiviridae family, and like one of the lysins expressed by these European phages (ORF 47 and 48) but, with a poorly conserved region when compared with the phiAP1 phage. The two identified lysins in the three European phages will be further characterized to better determine their contribution to the biological control of the bacterial wilt disease.…”

“…Bioinformatic analysis of ORFs 48 and 49 coding for tail fiber proteins in the three European phages allowed the identification of the domain IPR011050, associated with pectin lyase activity, which is involved in the cleavage of glycoside bonds in capsular polysaccharides and essential for initiation of phage infection. A short tail fiber was recently annotated in the ORF 43 of the R. solanacearum phage phiAP1 with a RmLc-like cupin domain, present in sugar isomerases with EPS depolymerase activity but, no experimental validation was performed [48]. No significant similarity was found between this ORF and ORFs 48 and 49 of the European R. solanacearum phages.…”

“…Phylogenetic analysis was performed with the whole genomes of the three river water European phages and a selection of their 26 closest viruses belonging to the Autographiviridae family (formerly Podoviridae) able to infect either R. solanacearum (three African phages [46,47], one American [48] and one Asian (accession number: MF979559) or the closely related species R. pseudosolanacearum (21 phages from diseased plants, soil or irrigation water isolated from Africa (14) [49] and Asia (7) [50][51][52]).…”

Genomic Analysis of the First European Bacteriophages with Depolymerase Activity and Biocontrol Efficacy Against the Phytopathogen<em> Ralstonia solanacearum</em>

“…Bioinformatic analysis of ORFs 48 and 49 coding for tail fiber proteins in the three European phages allowed the identification of the domain IPR011050, associated with pectin lyase activity, which is involved in the cleavage of glycoside bonds in capsular polysaccharides and essential for initiation of phage infection. A short tail fiber was recently annotated in the ORF 43 of the R. solanacearum phage phiAP1 with a RmLclike cupin domain, present in sugar isomerases with EPS depolymerase activity but, no experimental validation was performed [48]. No significant similarity was found between this ORF and ORFs 48 and 49 of the European R. solanacearum phages.…”

“…2.5.3. Phylogenetic analysis Phylogenetic analysis was performed with the whole genomes of the three river water European phages and a selection of their 26 closest viruses belonging to Autographiviridae family (formerly Podoviridae) able to infect either R. solanacearum (three African phages [46,47], one American [48] and one Asian (accession number: MF979559) or the closely related species R. pseudosolanacearum (21 phages from diseased plants, soil or irrigation water isolated from Africa (14) [49] and Asia (7) [50][51][52]). A genome-based phylogeny was generated using VICTOR [53], intergenomic distances were calculated with the Genome BLAST Distance Phylogeny (GBDP) d0 formula for nucleotidic sequences.…”