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Protoplasma

, Volume 174, Issue 1–2, pp 62–68 | Cite as

Status of nuclear division in arbuscular mycorrhizal fungi during in vitro development

  • G. Bécard
  • P. E. Pfeffer
Article

Summary

The number of nuclei in spores and along hyphae of an arbuscular mycorrhizal fungiGigaspora margarita was measured in digital images of fluorescence arising from mithramycin stained cultures. Typical dormant spores (250 μm diameter) contained 2000 nuclei. Eight hundred nuclei were mobilized during the first 3 days of germination. The number of nuclei in the spores nearly returned to the initial number after 22 days of hyphal growth. The average relative DNA content in the nuclei of dormant spores and in the nuclei of spores incubated for 22 days was comparable, as judged from fluorescence intensity. Hyphal elongation occurred with 460 nuclei per cm under a special set of in vitro conditions that promote extensive hyphal growth of arbuscular mycorrhizal fungi. We found an average total of 26000 hyphal nuclei per germinating spore after 22 days. The specific DNA polymerase α inhibitor aphidicolin did not inhibit spore germination but it rapidly reduced the rate of hyphal growth and arrested growth after 4 days. No nuclei were produced de novo during this time. These results demonstrate thatG. margarita replicates nuclear DNA and undergoes nuclear division when grown in vitro even in the absence of a plant host.

Keywords

Charge-coupled device DNA fluorescence Gigaspora margarita Image processing In vitro culture Nuclear division 

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References

  1. Allan RA, Miller JJ (1980) Influence of S-adenosylmethionine on DAPI-induced fluorescence of polyphosphate in the yeast vacuole. Can J Microbiol 26: 912–920PubMedGoogle Scholar
  2. Bécard G, Fortin JA (1988) Early events of vesicular-arbuscular mycorrhiza formation on Ri T-DNA transformed roots. New Phytol 108: 211–218Google Scholar
  3. Bécard G, Piché Y (1989 a) New aspects on the acquisition of biotrophic status by a VAM fungus,Gigaspora margarita. New Phytol 112: 77–83Google Scholar
  4. — — (1989 b) Fungal growth stimulation by CO2 and root exudates in vesicular-arbuscular mycorrhizal symbiosis. Appl Environ Microbiol 55: 2320–2325Google Scholar
  5. — — (1992) Establishment of VA mycorrhizae in root organ culture: review and proposed methodology. In: Norris JR, Read DJ, Varma AK (eds) Methods in microbiology, vol 24. Academic Press, London, pp 89–108Google Scholar
  6. —, Douds DD, Pfeffer PE (1992) Extensive hyphal growth of vesicular-arbuscular mycorrhizal fungi in the presence of CO2 and flavonols. Appl Environ Microbiol 58: 821–825Google Scholar
  7. Beilby JP, Kidby DK (1982) The early synthesis of RNA, protein, and some associated metabolic events on germinating vesicular-arbuscular mycorrhizal fungal spores ofGlomus caledonius. Can J Microbiol 28: 623–628Google Scholar
  8. Burggraaf AJP, Beringer JE (1988) Nuclear division and VA-mycorrhizal in vitro culture. In: Sylvia DM, Hung LL, Graham JH (eds) Mycorrhizae in the next decade. University of Florida, Gainesville, p 190Google Scholar
  9. — — (1989) Absence of nuclear DNA synthesis in vesicular-arbuscular mycorrhizal fungi during in vitro development. New Phytol 111: 25–33Google Scholar
  10. Coleman AW, Maguire MJ, Coleman JR (1981) Mithramycin-and 4′-6-diamidino-2-phenylindole (DAPI)-DNA staining for fluorescence microspectrophotometric measurement of DNA in nuclei, plastids, and virus particles. J Histochem Cytochem 29: 959–968PubMedGoogle Scholar
  11. Cook JC, Gemma JN, Koske RE (1987) Observations of nuclei in vesicular-arbuscular mycorrhizal fungi. Mycologia 79: 331–333Google Scholar
  12. Gonzales RC, Wintz P (1987) Digital image processing. Addison-Wesley, Reading, MAGoogle Scholar
  13. Heath B (1987) Fluorescent staining of fungal nuclei. In: Fuller MS, Jaworski A (eds) Zoosporic fungi in teaching and research. Southeastern Publishing, Greenville, NC, pp 169–171Google Scholar
  14. Hepper CM (1979) Germination and growth ofGlomus caledonius spores: the effects of inhibitors and nutrients. Soil Biol Biochem 11: 269–277Google Scholar
  15. Hiraoka Y, Sedat JW, Agard DA (1987) The use of charge-coupled device for quantitative optical microscopy of biological structures. Science 238: 36–41PubMedGoogle Scholar
  16. Viera A, Glenn MG (1990) DNA content of vesicular-arbuscular mycorrhizal fungal spores. Mycologia 82: 263–267Google Scholar
  17. Williams PG (1991) Axenic culture of arbuscular mycorrhizal fungi. In: Norris JR, Read DJ, Varma AK (eds) Methods of microbiology, vol 24. Academic Press, London, pp 203–220Google Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • G. Bécard
    • 1
  • P. E. Pfeffer
    • 1
  1. 1.U. S. Department of Agriculture, Agricultural Research ServiceEastern Regional Research CenterPhiladelphiaUSA

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