Genomic and Biological Characterization of Ralstonia solanacearum Inovirus Brazil 1, an Inovirus that Alters the Pathogenicity of the Phytopathogen Ralstonia pseudosolanacearum.

Filamentous bacteriophages contain a single-stranded DNA genome and have a peculiar lifestyle, since they do not cause host cell lysis, but establish a persistent association with the host, often causing behavioral changes, with effects on bacterial ecology. Over the years, a gradual reduction in the incidence of bacterial wilt has been observed in some fields from Brazil. This event, which has been associated with the loss of pathogenicity of Rasltonia spp. isolates due to infection by filamentous viruses of the inovirus group, is widely reported for Ralstonia spp. Asian isolates infected by inoviruses. In an attempt to elucidate which factors are associated with the phenomenon reported in Brazil, we investigated one isolate of R. solanacearum (UB-2014), with unusual characteristics for R. solanacearum, obtained from eggplant with mild wilt symptoms. To verify if the presence of filamentous bacteriophage was related to this phenotype, we performed viral purification and nucleic acid extraction. The phage genome was sequenced, and phylogenetic analyses demonstrated that the virus belongs to the family Inoviridae and was named as Ralstonia solanacerarum inovirus Brazil 1 (RSIBR1). RSIBR1 was transmitted to R. pseudosolanacearum GMI1000, and the virus-infected GMI1000 (GMI1000 VI) isolate showed alterations in phenotypic characteristics, as well as loss of pathogenicity, similarly to that observed in R. solanacearum isolate UB-2014. The presence of virus-infected UB-2014 and GMI1000 VI plants without symptoms, after 3 months, confirms that the infected isolates can colonize the plant without causing disease, which demonstrates that the phage infection changed the behavior of these pathogens.

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

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.