Date of Award

1980

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Abstract

Part I. Microcolonies of a filamentous, branched organism were observed by phase-contrast and interference-contrast microscopy in vascular extracts from internodes of sugarcane with ratoon stunting disease. The microcolonies were present in a matrix extracted from the xylem vessels of internodes by negative pressure. Matrices and microcolonies were not found in sugarcane known to be free of the disease. Electron microscopy revealed branched chains of cells with septa and mesosomes. The individual cells of the filaments were characteristic of the previously described coryneform bacterium associated with the disease. Vascular extracts from one- or two-node cuttings of diseased cane did not reveal matrices of microcolonies but did contain the individual cells or short branched chains of the organism. The filamentous spiderlike morphology of the microcolonies and true branching suggest a relation to the Actinomycetales. Part II. Freshly sectioned tissues of sugarcane with ratoon stunting disease were examined with the scanning electron microscope. The RSD-associated bacterium was recognized by its diameter, pleomorphism and xylem association. It commonly occupied the protoxylem and metaxylem vessels. The bacterium was observed in the gaps between secondary wall thickenings of xylem vessels and also adhering to the secondary wall. The lacunae, tracheids and xylem parenchyma cells also were invaded by the bacterium. The lumen of some infected xylem elements was filled with a dense network of bacterial cells. Remnants of a supporting matrix were observed in some of these lumina. Unicellular forms were most frequently observed; however, swollen forms, filamentous branching forms and microcolonies consisting of tightly aggregated bacterial cells which I have previously described from extracts of diseased tissue also were observed. Part III. The RSD-associated bacterium was extracted from diseased sugarcane and purified by differential and density gradient centrifugation. Cells were sonicated, DNA was extracted, and walls were prepared by enzymatic digestion. After cell wall hydrolysis, sugars and amino acids were analyzed. Fucose and rhamnose were the major sugar components. Lysine, ornithine, alanine, glutamic acid, and glycine were presumed to make up the peptidoglycan tetrapeptide and linkage. The Tm of the DNA was determined and the G + C content calculated at 60 moles %. These results are characteristic of the family Actinomycetaceae and more specifically the genus Actinomyces. Part IV. The transpiration rate of detached sugarcane shoots with ratoon stunting disease was significantly lower than that of detached healthy shoots. Apparent root pressure also decreased in diseased plants. Intact plant transpiration was measured by weight loss experiments from well watered pots contaning healthy or diseased canes which were unwatered for 7 days. Diffusive resistance of leaves to water vapor loss was measured daily. Diseased plants had a significantly lower transpiration rate than healthy plants; however, no significant differences in diffusive resistance between healthy and diseased plants was detected. This implied that the stomatal behavior of the plants was not affected by the disease, and that the lower transpiration rate in diseased plants appeared to be the consequence of the physical plugging of the xylem vessels by the RSD-associated bacterium and the gel matrices which were described previously.

Pages

81

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