Identification of Open Reading Frames Unique to a Select Agent: Ralstonia solanacearum Race 3 Biovar 2

Abstract

An 8x draft genome was obtained and annotated for Ralstonia solanacearum race 3 biovar 2 (R3B2) strain UW551, a United States Department of Agriculture Select Agent isolated from geranium. The draft UW551 genome consisted of 80,169 reads resulting in 582 contigs containing 5,925,491 base pairs, with an average 64.5% GC content. Annotation revealed a predicted 4,454 protein coding open reading frames (ORFs), 43 tRNAs, and 5 rRNAs; 2,793 (or 62%) of the ORFs had a functional assignment. The UW551 genome was compared with the published genome of R. solanacearum race 1 biovar 3 tropical tomato strain GMI1000. The two phylogenetically distinct strains were at least 71% syntenic in gene organization. Most genes encoding known pathogenicity determinants, including predicted type III secreted effectors, appeared to be common to both strains. A total of 402 unique UW551 ORFs were identified, none of which had a best hit or >45% amino acid sequence identity with any R. solanacearum predicted protein; 16 had strong (E < 10(-13)) best hits to ORFs found in other bacterial plant pathogens. Many of the 402 unique genes were clustered, including 5 found in the hrp region and 38 contiguous, potential prophage genes. Conservation of some UW551 unique genes among R3B2 strains was examined by polymerase chain reaction among a group of 58 strains from different races and biovars, resulting in the identification of genes that may be potentially useful for diagnostic detection and identification of R3B2 strains. One 22-kb region that appears to be present in GMI1000 as a result of horizontal gene transfer is absent from UW551 and encodes enzymes that likely are essential for utilization of the three sugar alcohols that distinguish biovars 3 and 4 from biovars 1 and 2.

Bacteriophages of Ralstonia solanacearum: Their Diversity and Utilization as Biocontrol Agents in Agriculture

“…NZ_AAKL00000000). This gene cluster was present in all race 3/biovar 2 (R3B2) strains tested, from a wide variety of geographical sources (Gabriel et al, 2006). As described above, RSM4 is also a filamentous prophage of the RSM group.…”

“…AY374414) and Korean strains of R. solanacearum (Murugaiyan et al, 2010). Of RSM1-related phages (B), RSM3, RSM4, RSM5, and RSM6 are prophages of strains MAFF730139, UW551 (race 3, biovar 2; Gabriel et al, 2006), IPO1609 (race 3, biovae 2; Remenant et al, 2010), and CMR15 (phylotype III, Remenant et al, 2010), respectively. A similar prophage found in Ralstonia pickettii 12J is also shown.…”

Ralstonia solanacearum requires F-box-like domain-containing type III effectors to promote disease on several host plants

“…Indeed all F-box domains (plant and other eukaryotes) have a conserved leucine at that position, which has been shown to be important for the interaction with the SKP1-like protein (19). Interestingly this F65 is reverted to the conserved leucine residue in the GALA4 orthologous gene in strains Molk2 (N.P., C. Boucher, and S.G., unpublished data) and UW551 (17). GALA6 Binds to Arabidopsis ASK1 and ASK2.…”

“…Recent draft sequences of two other R. solanacearum strains adapted to specific host plants revealed the presence of orthologous genes for six of the seven GALA genes in both strains (Molk2 banana-specific strain (N.P., C. Boucher, and S.G., unpublished data) and UW551, a potato-specific strain) (17). The conservation of these effectors among evolutionarily distant strains (18) suggests that these T3SS effectors are selected for and are, therefore, probably important for R. solanacearum pathogenicity.…”

Global regulation of pathogenicity mechanism of Ralstonia solanacearum

“…Genome sequence analysis was performed on GMI1000 (Salanoubat et al 2002) and R. solanacearum strain UW551 (Gabriel et al 2006), which are nonpathogenic to tobacco plants, and has been analyzed in another two strains (Boucher, personal communication). Based on genomic analysis, screening of type III effectors controlled by HrpB and genes induced in tomato during growth has been conducted using in vivo expression technology, resulting in isolation of many candidate type III effector genes Mukaihara et al 2004) and pathogenicity-related genes .…”