Title

Insights into the Molecular Mechanism of Streptomyces ipomoeae Pathogenesis: Thaxtomin C Biosynthesis and Regulation

Identifier

etd-04192011-184316

Degree

Doctor of Philosophy (PhD)

Department

Biological Sciences

Document Type

Dissertation

Abstract

Streptomyces ipomoeae is the causal agent of Streptomyces soil rot of sweetpotato, a disease marked by highly necrotic destruction of adventitious roots, including the development of necrotic lesions on the fleshy storage roots. To facilitate further genetic manipulation of this agriculturally important pathogen, an effective gene transfer system for S. ipomoeae was developed by using an optimized intergeneric conjugation procedure from Escherichia coli. Conjugation results varied greatly depending on the particular media used for S. ipomoeae spore preparation and conjugation. Transconjugant to recipient ratios as great as 4.1 x 10-5 were achieved when International Streptomyces Project Medium 4 was used for both sporulation and conjugation protocols. Both site-specifically integrating and autonomously replicating plasmids could be introduced and maintained in S. ipomoeae, and plasmids could be introduced with approximate equivalent frequencies from either methyl-proficient or methyl-deficient E. coli donors; the latter result indicates a likely absence of relevant methyl-specific restriction in S. ipomoeae. Streptomyces potato scab pathogens produce a phytotoxin (thaxtomin A) that appears to facilitate their entrance into host plants. S. ipomoeae produces a less-modified thaxtomin derivative (thaxtomin C) whose role in pathogenicity has not been examined. Here, we cloned and sequenced the thaxtomin C (txt) gene cluster of S. ipomoeae and I then constructed targeted txt mutants. The mutants were unable to penetrate intact adventitious roots but still caused necrosis on storage root tissue. These results, taken in context with previous histopathological study of S. ipomoeae infection, suggest that thaxtomin C plays an essential role in inter- and intracellular penetration of sweetpotato adventitious roots by S. ipomoeae. Once inside the plant host, the pathogen uses a yet-to-be-determined factor(s) to necrotize root tissue, including that of any storage roots it encounters. An AraC/XylS family transcriptional regulator TxtR encoded in the thaxtomin C gene cluster is shown here to be essential for S. ipomoeae pathogenicity, though the putative host-derived ligand that binds TxtR to activate thaxtomin C production remains to be identified. Another conserved gene txtH, which is also imbedded in the thaxtomin C gene cluster, was found to be not required for S. ipomoeae pathogenicity.

Date

2011

Document Availability at the Time of Submission

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Committee Chair

Pettis, Gregg S.

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