Vincent M. Russo

Effect on germination and post-germination development of Colletotrichum dematium var circinans due to light and dark incubation and coverslip placement

Light regime was not a major factor in germination of spores of Colletotrichum dematium var circinans. Number of germ-tubes and appressoria formed were affected by incubation regime and coverslip placement. Most germ-tubes and appressoria were produced on spores under shimmed coverslips. Light incubation favored germ-tube production and dark incubation favored appressoria production. Predicted time to 100% germination was affected by treatments. Shortest times were predicted for light incubated spores under shimmed coverslips. Longest times were predicted for non-coverslipped spores. A significant number of appressoria were produced sessile on spores. Most sessile appressoria were produced on dark incubated spores. Approximately 56 % of all spores germinated, and approximately 60% of all germ-tubes and appressoria were produced on these spores which were situated closest to coverslip or droplet edges. Placement of germ-tubes on spores was not affected by treatments. However, percentages of germ-tubes, initiated from various areas of spores, were significantly different. This variation appears to be due to factors internal to spores.

Germination and post-germination development of Colletotrichum dematium f. circinans on Allium cepa

Spores of Colletotrichum dematium f. circinans were inoculated onto the abaxial epidermis of washed and unwashed pieces of leaf-base tissue taken from the first thru fourth layers of white, yellow and red onions. Washing tissue dramatically improved germination, less than 0.1% for unwashed versus 83% for washed. Germination capacity, and the number of germination types (16) found on washed tissue was not affected by onion bulb color or leaf-base layer inoculated. Spores with one sessile appressorium, type 001, or with one germ tube without an appressorium, type 100, or with one germ tube with an appressorium produced at the tip, type 110, made up approximately 93% of the types for those spores which germinated.

Protoplast responses in the epidermis of Allium cepa induced by penetration by Botrytis allii

Penetration of Allium cepa epidermal cells (white, yellow, and red varieties) by Botrytis allii induced a response by host protoplasts in normal tissue which was not observed when penetrations were made in protoplast-free host cell walls. Callose and auto-fluorescing substances (possibly phenolic compounds) were located at the penetration sites only in normal host cells containing protoplasts. Lignin tests were negative. Halos were clearly visible in both types of tissue. Autofluorescence was observed at penetration sites in normal cells of all cultivars but general wall background autofluorescence was not observed in white onions. Autofluorescence was generally yellow green and when treated with ammonium hydroxide became green. Treatment with sodium hydroxide abolished autofluorescence. No attempt was made to isolate the autofluorescing material.

Degenerative changes in the epidermal cell wall of Allium cepa caused by Botrytis allii: Loss of dry mass, pectin, and cellulose in halos

Halos detected using interference microscopy (even- and fringle-field modes with monoand poly-chromatic light) around penetration sites of Botrytis allii in cell walls of normal and protoplast-free outer epidermal tissue of white, yellow, and red onions were alike. Halos in protoplast-free cell walls contained 33% less dry mass than areas of these walls adjacent to halos (quantitative interference miscroscopy with 546 nm light in the even-field mode). Halos were significantly larger in the white onion than in the yellow and red varieties. The loss of cell wall dry mass during the production of halos involved the loss of pectin and cellulose. We infer that this is caused by enzymes released from the pathogen. Cuticle degradation at penetration sites was not observed.

Effect of prism generated discreet wavelengths of light on germination and post-germination development of Colletotrichum dematium var circinans (Berk.) v. Arx

Spores, harvested from 8 da old colonies ofColletotrichum dematium varcircinans (Berk.) v. Arx, were placed on slides under shimmed coverslips and subjected to 10 min irradiation with wavelengths of light ranging from 400 to 750 nm. Controls consisted of spores exposed to 10 min of fluorescent or tungsten light, or to continuous dark. Germination, and production of sessile and non-sessile appressoria, and germ-tubes was monitored. In addition position of germ-tubes on spores was noted and germination types were classified by numbers of germ-tubes and sessile and non-sessile appressoria produced. When compared to dark controls certain wavelengths, 400, 450, 600 and 650 nm, appeared to be inhibitory to germination and post-germination development, while others, 500, 550 and 750 nm, appeared to be stimulatory. Spores irradiated with some of the wavelengths (550, 700, 750 nm) that affected other responses produced the smallest percentage of sessile appressoria (approx. 10%). Placement of germ-tubes was not affected by treatment. More germ-tubes were produced from spore bottoms (53%) than tops (32%), and tips (15%). The fewest number of germ-tubes was produced from spore centers and from one tip. Thirteen germination types were noted. About 53% of spores germinated with one germ-tube, one non-sessile appressoria, and no sessile appressoria. The remaining 47% were distributed throughout the remaining 12 types, and ranged from 21% to less than 1% of the total. Treatments did not affect this distribution.

Spore age and coverslip placement affects germination and post-germination development of Colletotrichum dematium var circinans

Spore suspensions from young (10–14 da; young spores) and old (4 mo; old spores) colonies of PColletotrichum dematium var circinans were placed on slides. Coverslips were left off, placed on in the normal manner, or supported on shims. Slides were placed in moist chambers and incubated in light or dark for up to 48 hrs. Germination and post-germination development were studied. Shimming had some beneficial effect on germination, especially for old spores in dark. In general, more germ-tubes and appressoria were produced on spores under shims than spores with other coverslip treatments. By 48 hrs more old spores under shims germinated, and greater numbers of germ-tubes and appressoria were produced than on other old spores under different coverslip treatments. However, numbers produced were lower than those predicted for comparably treated young spores. Spore age, incubation regime, and placement of coverslips did not affect germ-tube initiation. For all treatments more germ-tubes were initiated from spore tops than bottoms or tips. Fewer germ-tubes were initiated from spore centers than other locations on tops and bottoms, and from both tips than one tip. Approximately 26 % of all appressoria were produced sessile. A higher percentage of sessile appressoria were produced on old spores (80 %) than on young spores (20%).