|Kimbrel, Jeffrey -|
|Givan, Scott -|
|Creason, Allison -|
|Mills, Dalice -|
|Armstrong, Donald -|
|Chang, Jeff -|
Submitted to: Biomed Central (BMC) Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 29, 2010
Publication Date: September 28, 2010
Citation: Kimbrel, J.A., Givan, S.A., Halgren, A.B., Creason, A.L., Mills, D.I., Banowetz, G.M., Armstrong, D.J., Chang, J.H. 2010. An Improved, High-Quality Draft Genome Sequence of the Germination-Arrest Factor-Producing Pseudomonas fluorescens WH6. Biomed Central (BMC) Genomics. 11:522-536. Interpretive Summary: WH6 is a strain of the soil bacterium Pseudomonas fluorescens that produces the bioherbicide called Germination Arrest Factor (GAF). In previous work we showed that GAF arrests the germination of many grassy weeds that impact the production of a variety of crops. This study sequenced the entire genome of WH6 and reported on two mutant strains that do not produce GAF. The mutations in these strains were identified and revealed new information on how GAF is produced by WH^ and secreted into the soil where GAF herbicidal activity occurs.
Technical Abstract: Pseudomonas fluorescens is a genetically and physiologically diverse species of bacteria present in many habitats and in association with plants. This species of bacteria produces a large array of secondary metabolites with potential as natural products. P. fluorescens isolate WH6 produces Germination-Arrest Factor (GAF), a predicted small peptide or amino acid analog with herbicidal activity that specifically inhibits germination of seeds of graminaceous species. We used a hybrid next-generation sequencing approach to develop a highquality draft genome sequence for P. fluorescens WH6. We employed automated, manual, and experimental methods to further improve the draft genome sequence. From this assembly of 6.27 megabases, we predicted 5876 genes, of which 3115 were core to P. fluorescens and 1567 were unique to WH6. Comparative genomic studies of WH6 revealed unexpected similarity in synteny and higher than expected orthology of genes with P. fluorescens SBW25. A phylogenomic study also placed WH6 in the same lineage as SBW25. In a previous non-saturating mutagenesis screen we identified two genes necessary for GAF activity in WH6. Mapping of their flanking sequences revealed genes that encode a candidate anti-sigma factor and an aminotransferase. Finally, we discovered several candidate virulence and host-association mechanisms, one of which appears to be a complete type III secretion system. The improved high-quality draft genome sequence of WH6 contributes towards resolving the P. fluorescens species, providing additional impetus to establishing two separate lineages in P. fluorescens. Despite higher than expected levels of orthology and synteny to SBW25, WH6 still had a substantial number of unique genes and represents another source for the discovery of genes with implications in affecting plant growth and health. Two genes are demonstrably necessary for GAF and further characterization of their proteins is important for developing natural products as control measure against grassy weeds. Finally, WH6 is the first isolate of P. fluorescens reported to encode a complete T3SS. This gives us the opportunity to explore the role of what has traditionally been thought of as a virulence mechanism for non-pathogenic interactions with plants.