Microscopic and molecular analyses showed the presence of endobacteria inside the mycelia of four out of twelve nitrous oxide (N 2 O)-producing fungal isolates identified as Mortierella elongata. The 16S rRNA gene was successfully amplified with DNA extracted directly from the endobacterium-containing fungal strains and all sequences were related to that of Candidatus Glomeribacter gigasporarum in the family Burkholderiaceae. Bacterial endotoxin was detected in the endobacterium-positive fungal strains but only trace levels were found in endobacterium-negative strains. No significant relationship was found between the fungal N 2 O-producing activity and the presence of endobacteria.Key words: endobacterium, N 2 O production, Mortierella elongata, soil fungusThe intracellular distribution of bacteria, or more specifically bacterial endosymbiosis, is known for numerous eukaryotic organisms including ciliate protozoa (25), various insects (11), a frenulata (marine invertebrate) (12), and wellknown legume roots. Concerning the association of bacteria with fungi, bacterial endosymbiosis has been reported in some strains of Glomeromycota [e.g. Geosiphon pyriform (22), and Gigaspora and Scutellospora species (4, 6, 7, 15)], Ascomycota [e.g. Tuber borchii (1)], Basidiomycota [e.g. Laccaria bicolor; (3)] and Zygomycota [e.g. Rhizopus species (10, 18)]. In our recent study, nitrous oxide (N 2 O)-producing fungal strains were isolated from upland soils and the intramycelial localization of bacterium (endobacterium) in the fungal isolates was found (Sato, Y., et al. 2008. Abstracts for the 12th International Symposium on Microbial Ecology, Cairns, Australia). The purpose of this study is to examine further the presence of endobacteria using fluorescence and electron microscopy, the amount of endotoxin, the 16S rRNA gene, and the relationship between the presence of endobacteria and fungal N 2 O-producing activity.Ten fungal strains used in this study were isolated from no-tilled, low-nitrogen input cropland soils at the Field Science Center, Ibaraki University College of Agriculture, located in the Kanto plains of Japan and were identified as phylum Zygomycota, Mortierella elongata from their morphological characteristics and their 18S-26S/28S internal transcribed spacer sequence (ITS). All of the M. elongata strains were examined for activity to produce N 2 O and found to have low activity levels during growth in cultures containing 10 mM nitrite but no activity in cultures with 10 mM nitrate (Y. Sato, T. Nishizawa, M. Umezu, K. Tsuruta, K. Narisawa, M. Komatsuzaki, N. Kaneko, H. Ohta, submitted for publication). Comparisons were made with herbarium materials, Mortierella elongata Linnemann NBRC 8570 and M. elongata MAFF 425591, obtained from the National Institute of Technology and Evaluation (NITE Biological Resource Center, Chiba, Japan), and National Institute of Agrobiological Sciences (NIAS Genebank, Ibaraki, Japan), respectively.The morphology of M. elongata strains was observed by fluorescence microscope. Briefly, ...
