Viability and Maturation of Aphanomyces cochlioides Oospores

Abstract

JSTOR is a not-for-profit service that helps scholars, researchers, and students discover, use, and build upon a wide range of content in a trusted digital archive. We use information technology and tools to increase productivity and facilitate new forms of scholarship. For more information about JSTOR, please contact support@jstor.org. This content downloaded from 155.247.166.234 on Fri, 01 Jan 2016 07:14:02 UTC All use subject to JSTOR Terms and Conditions Mycologia, 95(2), 2003, pp. 321-326.Abstract: Plasmolysis, tetrazolium bromide staining and microscopic appearance were tested for their usefulness in determining viability of oospores of Aphanomyces cochlioides. For comparison, three lethal treatments were employed to contrast the reaction of dead oospores and untreated, presumably viable oospores. Few oospores stained with tetrazolium bromide, even though plasmolysis and microscopic appearance indicated that 85% were viable. Cytoplasm of viable oospores was densely organized and uniformly granular (DOUG), whereas cytoplasm of oospores exposed to lethal treatments was loosely organized and non-uniformly granular (LONG). Doseresponse bioassay experiments were conducted with untreated oospores of varying inoculum densities or with mixtures of untreated DOUG and heat-treated LONG oospores in varying proportions. The number of DOUG oospores was correlated (R2 = 0.62, P < 0.001) with severity of damping-off of sugar beet seedlings caused by A. cochlioides. Thus, the granular appearance of cytoplasm offered a fast, easy and reliable indicator of viability of A. cochlioides oospores. Tests with newly formed oospores/oogonia showed that >80% harvested at 3-4 d after inoculation of hypocotyls stained with tetrazolium, but by 8-9 d <10% stained, apparently because of declining permeability of the spore wall to tetrazolium as oospores matured.

Homothallism in Pseudoperonospora humuli

“… Viability was determined by inspection of approximately 300 oospores from each of three replicate plants. DOUG is an acronym for densely organized and uniformly granular organization of oosporic cytoplasm (Dyer & Windels, ). …”

“…For direct observation, three replicates of approximately 100 oospores from each isolate or cross of isolates were placed in slide wells in 30 μ L water. Oospores were counted and the number of oospores with cytoplasm showing densely organized uniform granules was recorded, as this morphology is correlated with viability in other oomycetes (Dyer & Windels, ) and often assumed to be an indication of viability (Tommerup et al ., ; Morgan, ; Fig. ).…”

“…After inspection, oospores were allowed to dry overnight, resuspended in 30 μ L 4 m NaCl and incubated for 5 min to 1 h. Oospores showing cytoplasm shrinkage and detachment from the cell wall, indicative of cell membrane integrity, were counted as viable. Those that showed no change after an hour were considered nonviable (Jiang & Erwin, ; Dyer & Windels, ). In total, 10 different monosporic isolates and 13 crosses of two isolates were examined.…”

Testing of life history traits of a soilborne pathogen in vitro: Do characteristics of oospores change according the strains of Aphanomyces euteiches and the host plant infected by the pathogen?

“…The second explanation involves inconsistencies associated with the MTT staining method that have already been noted (Etxeberria et al, ) suggesting that the evaluation of oospore quality may be subjective. Finally, to validate the virulence of these oospores, it would be necessary to have a correlation between visual tests and the ability of oospores to induce disease (Dyer & Windels, ).…”

A reference genome sequence resource for the sugar beet root rot pathogen Aphanomyces cochlioides

“…Tissue from A. cochlioides oospores, mycelium, and infected sugar beet seedlings were collected, and RNA was extracted using a RNeasy Plant Mini Kit (Qiagen, Inc., Valencia, CA, USA). Mycelium was grown as described above, and oospores were grown on 2-week old sugar beet hypocotyls using established methods (Dyer & Windels, 2003). Also, 2-week old sugar beet seedlings of cultivar Crystal 093 RR were soil-drip inoculated with 5,000 zoospores per plant following established protocols, and diseased roots were removed from pots at 1, 3, 5, and 10 days post inoculation (dpi) for RNA extraction (Windels & Brantner, 2000).…”