Summary. A container system was constructed to study the response of soybean roots to infection by mycorrhizal or pathogenic fungi. The system allows a rapid and synchronous inoculation by Glomus mosseae or Rhizoctonia solani. The phytoalexin glyceollin was measured in roots of inoculated and uninoculated plants for a period of 30 days. A significantly increased content of phytoalexin was found in R. solani-infected roots as compared to uninfected control roots. However, there was no difference in the glyceollin contents of the mycorrhizal and the control roots for up to 23 days after inoculation. The accumulation of glyceollin in R. solani-infected roots was not influenced by a subsequent inoculation with G. mosseae. Moreover glyceollin accumulated in mycorrhizal plants to the same extent as in control plants when they were inoculated with R. solani. The two fungi did not mutually influence the course of infection when they were inoculated together. Key words. VA-mycorrhiza; Glycine max; glyceollin; rhizosphere; rhizotron.The vesicular-arbuscular mycorrhiza (VAM) is a widespread symbiosis between plants and fungi. Although there are many studies on the established symbiosis, for instance concerning phosphorus uptake, carbon transport, heavy-metal toxicity or the influence of pesticides, little is known about the physiology of the interaction between plant and fungus at the beginning of infection 1. In particular it is poorly known whether the mutualistic character of the established symbiosis manifests itself already at the beginning of infection, and whether recognition differs from that involved in pathogenic interactions. One reason for the lack of data is the difficulty of handling mycorrhizas in the laboratory. A fast and homogeneous course of infection is needed. In the present work we have employed a modified version of the container system of Schfiepp et al. 2 which allows one to obtain such infections and therefore to investigate the first infection stages of mycorrhiza formation. A short description of our technique has already been published 3. We have used this system to compare the phytoalexin response of roots to infection by a VAM fungus and a pathogen. Phytoalexins are antimicrobial compounds of low molecular weight which accumulate in plants after exposure to microorganisms. Their production appears to result from de novo synthesis of the corresponding biosynthetic enzymes upon a challenge of the plant by a pathogen or an elicitor 4, s. There is evidence that accumulation of phytoalexins at the site of attempted infection is one of the mechanisms conferring resistance against infections in plants 6. The phytoalexin response to pathogens has been particularly well studied in soybeans which accumulate the pterocarpan glyceollin 5.Little is known about the accumulation of this phytoalexin in response to mycorrhizal fungi. An early report indicated glyceollin accumulation in the fully established VAM symbiosis 7. Using our container system, we have recently obtained first indications that the phytoa...
Wild-type soybean (Glycine max L. Merr. cv. Bragg) plants and two soybean mutants derived from cv. Bragg (nod 49 and nod 139) unable to form nodules with Bradyrhizobium japonicum were compared with regard to their reaction to the vesicular-arbuscular mycorrhizal fungus Glomus mosseae. The roots from wild-type and mutant plants entered equally well into vesicular-arbuscular mycorrhiza symbiosis. Polyadenylated RNA was isolated from nodule-free mycorrhizal and non-mycorrhizal roots of wild-type and mutant plants and translated in vitro. The translation products were subjected to immunoprecipitation using antisera reacting with soluble or membrane-bound nodulins. The antisera did not immunoprecipitate any of the translation products from non-mycorrhizal roots. However, they reacted with specific translation products from mycorrhizal roots of both wild-type and mutant plants: two polypeptides (MWs 135-140 and 18 kDa) were immunoprecipitated with the antiserum against soluble nodulins and three (MWs 21-28 kDa) with the antiserum against membrane-bound nodulins. These results indicate that symbiosis-specific polypeptides, possibly identical with nodulins, are induced in the mycorrhiza and therefore can be termed "mycorrhizins".
Mycorrhizal fungi are usually identified on the basis of the morphological characters shown by fruit bodies, spores, vegetative mycelia or symbiotic structures. The development of molecular techniques provides a valuable and alternative approach to identify mycorrhizal fungi, especially when it is difficult to gather a sufficient number of data on morphological features. Short arbitrary oligonucleotides were used as primers for the amplification of genomic DNA extracted from spores of arbuscular fungi. The RAPD fingerprints showed banding patterns which allowed us to distinguish between species and even isolates within Glomales. In order to identify mycorrhizal fungi during their symbiotic phase, a nonpolymorphic RAPD band identified as marker for some isolates of Glomus mosseae was purified from agarose gels and cloned in a bluescript vector. The fragment was sequenced and specific primers (PO-M3) were designed for the mycorrhizal fungus. They specifically and successfully amplified the DNA not only from G. mosseae spores, but also from roots of pea, clover, leek and onion plants when they were colonized by G. mosseae isolates.
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