MSA - Cell and molecular biology/Physiology & Genetics
Bentivenga, Stephen P. , Kumar, T.K. Arun , Kumar, Leticia , Celio, Gail J. , Roberson, Robert W. , McLaughlin, David J. .
Cellular organization in germ tube tips of the Glomeromycota: the taming of Gigaspora.
The fine structure of fungal hyphal tips often is phylogenetically informative. In particular, morphology of the Spitzenkörper varies among higher taxa. To date, no one has thoroughly characterized the hyphal tips of members of the phylum Glomeromycota. This is partly due to difficulty growing and manipulating living hyphae of these obligate symbionts. We observed growing germ tubes of Gigaspora gigantea, G. margarita, and G. rosea, using a combination of light microscopy (LM) and transmission electron microscopy (TEM). We used a variety of optics for LM (phase contrast, differential interference contrast, and confocal laser scanning), using both transmitted and fluorescent light with the lipophilic dye FM4-64. For TEM, we used both chemical fixation and freeze-substitution. Spores were surface disinfested and germinated on cellophane sheets overlaying water agar prior to processing and viewing. We also germinated spores in agar in inverted cover glass culture chambers. To date, we have not detected evidence of an organized Spitzenkörper, either using LM or TEM techniques. Germ tubes of all species were extremely sensitive to manipulation. Healthy germ tubes often showed rapid bidirectional cytoplasmic streaming, whereas germ tubes that had been disturbed showed reduced or no cytoplasmic movement. Actively growing germ tubes contain a cluster of 10-20 large (0.5 - 1.0 µm dia.) spherical bodies approximately 3-8 µm behind the apex. The bodies, which we hypothesize are lipid bodies or vacuoles, move rapidly in healthy germ tubes. These bodies disappear immediately after any cellular perturbation. Freeze-substituted cells showed superior preservation of the two-layered cell wall when compared to cells that had been chemically fixed. Freeze-substitution also revealed at least three distinct types of putative vesicles near the tip. Our work emphasizes the ephemeral nature of cellular organization, and the need for as little manipulation as possible to observe germ tube structure accurately.
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1 - University of Wisconsin Oshkosh, Department of Biology and Microbiology, 800 Algoma Blvd., Oshkosh, WI, 54901, U.S.A.
2 - University of Minnesota, Department of Plant Biology, 250 Biological Science Center, 1445 Gortner Ave., St. Paul, MN, 55108, U.S.A.
3 - University of Minnesota, Imaging Center, College of Biological Sciences, 123 Snyder Hall, 1475 Gortner Ave., St. Paul, MN, 55108, U.S.A.
4 - Arizona State University, School of Life Sciences, P.O. Box 874601, Tempe, Arizona, 85287, USA
Presentation Type: Poster:Posters for Topics
Location: Event Tent/Cliff Lodge
Date: Tuesday, July 28th, 2009
Time: 5:30 PM