Actinomycetes University of Udine, Mycology Department ISSN: 0732-0574 Vol. 7, Num. 1, 1996

ACTINOMYCETOLOGICA Vol. 9, No. 2, 1995

Published by the Society for Actinomycetes, Japan

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EMERGING CONCEPTS OF SECONDARY METABOLISM IN ACTINOMYCETES

A.L.Demain and A.Fang

Dept. Biology, Massachusetts Inst. Technology, Cambridge, MA 02139, USA

Actinomycetologica, 9: 98-117, 1995

Secondary metabolism in actinomycetes is brought on by exhaustion of a nutrient and/or by a growth rate decrease. These events generate signals which effect a cascade of regulatory events resulting in chemical differentiation (secondary metabolism) and morphological differentiation (morphogenesis). The signal is often a low molecular weight butyrolactone inducer (autoregulatory factor) which acts by negative control, i.e. by binding to a regulatory protein (repressor protein/recep- tor protein) which prevents secondary metabolism and morphogenesis during rapid growth and nutrient sufficiency. Nutrient/growth rate signals presumably activate a "master gene" which either acts at the level of translation by encoding a rare tRNA, or by encoding a positive transcription factor. Such master genes control both secondary metabolism and morphogenesis. At the second level of regulatory hierarchy are genes which control one branch of the cascade, i.e. either secondary metabolism or morphogenesis but not both. In the secondary metabolism branch, genes at the third level control formation of particular groups of secondary metabolites. At the fourth level are genes which control smaller groups, and finally, fifth level genes control individual biosynthetic pathways; these are usually positively acting but at least one acts negatively. These are also several levels of hierarchy on the morphogenesis branch. The second level includes genes which control aerial mycelium formation plus all the conidial genes lower in the cascade. Each third level locus controls a particular stage of conidiation (e.g. coiling, septation, wall thickening, spore matura- tion or spore pigmentation). Many of the these conidiation loci are complex, containing a number of genes; at least two code for sigma factors. Feedback regulation also plays a role in secondary metabolite control.

Authors' Abstract

THE STRINGENT RESPONSE AND THE INDUCTION OF NIKKOMYCIN PRODUCTION IN STREPTOMYCES TENDAE

U.Pfefferle^1, K.Ochi^2 and H.-P.Fiedler^1

^1 Universitat Tubingen, Biologisches Inst., Auf der Morgenstelle 28, D-72076 Tubingen, Germany and ^2 National Food Research Inst., 2-1-2 Kannondai, Tsukuba, Ibaraki 305, Japan

Actinomycetologica, 9: 118-123, 1995

Streptomyces tendae Tu 901/8c produces several secondary metabolites originating from different biosynthetic pathways, namely the nikkomycins, ketomycin, chlorothricin and the juglomycins. The significance of the stringent response in the induction of the nikkomycin production during different growth rates in continuous culture and batch fermentations was investigated. Merely, after nutritional shift down into chemically defined medium the strain exhibited a stringent response with two accompanying intracellular accumulations of ppGpp. This was found to be due to different consumption rates of some intracellular amino acids and dependent on relA functions. A relaxed mutant with a reduced ability to accumulate ppGpp, also with respect to the second accumulation observed, failed to produce nikkomycins.

Authors' Abstract

ISOLATION OF MUTANTS OF STREPTOMYCES GRISEUS THAT SPORULATE IN NUTRIENT RICH MEDIA: CLONING OF DNA FRAGMENTS THAT SUPPRESS THE MUTATIONS

S.Kawamoto^1 and J.C.Ensign^2

^1 National Food Research Inst., 2-1-2 Kannondai, Tsukuba City, 305, Japan and ^2 Dept. Bacteriology, 1550 Linden Drive, University of Wisconsin, Madison, WI 53706, USA

Actinomycetologica, 9: 124-135, 1995

Streptomyces griseus does not sporulate when grown in a nutrient rich medium. Two mutants that sporulate in such media were isolated and characterized. The mutants sporulate at the same time as the wildtype when grown in a nutrient poor medium. The mutants and wildtype exhibit the same commitment to sporulation following a nutritional shiftdown. Six genomic DNA fragments of wildtype cells when transformed into the mutants caused these mutants to acquire the wildtype phenotype of not sporulating in rich growth media. Restriction analyses show that the six cloned DNA fragments are different. The six clones sup- pressed sporulation when transformed into S.coelicolor and S.lividans and secondary metabolite production was sup- pressed in S.coelicolor.

Authors' Abstract

CLONING AND CHARACTERIZATION OF A GENE INVOLVED IN REGULATION OF SPORULATION AND CELL DIVISION OF STREPTOMYCES GRISEUS

S.Kawamoto^1 and J.C.Ensign^2

^1 National Food Research Inst., 2-1-2 Kannondai, Tsukuba City, 305, Japan and ^2 Dept. Bacteriology, 1550 Linden Drive, University of Wisconsin, Madison, WI 53706, USA

Actinomycetologica, 9: 136-151, 1995

A mutant of Streptomyces griseus, NY5, differs from the parent in sporulating when grown in a nutrient rich growth medium. Cloned genes of the wildtype organism in the multicopy plasmid pIJ702 transformed into the NY5 mutant cells resulted in converting the mutant phenotype of sporulating in rich medium to the parental genotype of not sporulating in rich medium. A 1.5 kb DNA fragment was sequenced and found to contain two open reading frame gene regions. Restriction deletion mapping and subcloning revealed a gene which when transformed in high copy number into S. griseus wildtype, mutant NY5 and S. lividans resulted in suppression of sporulation and fragmented cell growth. The gene, named ssgA (DDJB/EMBL/Gen Bank accession no. D50051), encodes a 145 amino acid protein with a calculated size of 15.8 kDa. The predicted protein has a strong negative charge and shows no significant sequence homology to known proteins. Southern analysis detected regions in S.coelicolor and S.lividans DNA homologous to ssgA.

Authors' Abstract

CHEMOTAXIS IN THE ZOOSPORIC ACTINOMYCETE CATENULOPLANES JAPONICUS

M.Hayakawa, M.Ariizumi, T.Yamazaki and H.Nonomura

Dept. Applied Chemistry and Biotechnology, Yamanashi University, Takeda-4, Kofu 400, Japan

Actinomycetologica, 9: 152-163, 1995

 The motility and chemotactic behavior of the soil actinomycete Catenuloplanes japonicus IFO 14176 was studied. The mature growth on the humic acid-vitamin agar medium exhibited the abundant formation of dichotomously branched aerial hyphae, from which motile, flagellated spores, or zoospores, were released upon immersion into a 10^-2M phosphate buffer (pH 7.0) containing 10% soil extract. A quantitative microcapillary assay revealed that the zoospores are significantly attracted to a variety of organic compounds common to the soil environments. Monosaccharides such as L-arabinose, D-xylose, D-glucose and D- galactose served as positive chemoattractants at considerably low concentrations; the maximum responses to the compounds occurred at 10^-5 to 10^-3 M, and the thresholds were 10^-6 to 10^-5 M. Amino acids (D-alanine, L-glutamate, and L-proline), aromatic compounds (vanillin, vanillate, protocatechuate, and myricetin), uronic acid (alpha-D-galacturonate) and amino sugars (D-glu- cosamine, and N-acetyl-D-glucosamine) all elicited concentration- dependent positive responses within the range of 10^-4 to 10^-1 M. Of the tested compounds, vanillin at 10^-l M provided the strongest response. The chemotactic activities of C.japonicus should tend to bring it toward the appropriate ecological locations where concentrated food sources are present. The Palleroni chemotactic method based on the 10^-l M-vanillin attraction achieved the selective isolation of Catenuloplanes spp. from natural soil samples.

 Authors' Abstract

TWO NEW SPECIES IN THE GENUS ACTINOMADURA: A.GLOMERATA SP. NOV. AND A.LONGICATENA SP. NOV.

T.Itoh, T.Kudo, H.Oyaizu^1 and A.Seino^2

Japan Collection of Microorganisms (JCM), Inst. Physical and Chemical Res. (RIKEN), 2-1 Hirosawa, Wako-shi, Saitama 351-01, ^1 Faculty of Agriculture, Tokyo University, 1-1- 1 Yayoi, Bunkyo-ku, Tokyo 113 and ^2 Research Center for Biological Function, The Kitasato Institute, 5-9-1, Shirokane, Minato-ku, Tokyo 108, Japan

Actinomycetologica, 9: 164-177, 1995

Taxonomy of two new actinomycete strains, which were isolated from a decayed leaf and a root of herbaceous plant, was studied. The organisms had branched substrate mycelia and chains of arthrospores on aerial mycelia. Cell walls of both strains contained meso-diaminopimelic acid and whole-cells contained madurose as a diagnostic sugar. Major menaquinone was MK-9(H6), and major fatty acids were hexadecanoic acid, 14-methylpentadeca- noic acid, octadecanoic acid, and 10-methyloctadecanoic acid. Diphosphatidylglycerol, phosphatidylinositol, and phos- phatidylethanolamine were detected as major phospholipid components. These morphological and chemotaxonomic features sug- gested that the isolates related to the genus Actinomadura, although most of Actinomadura species contain no major amount of phosphatidylethanolamine. Furthermore, comparison of the 16S rRNA genes allocates the two isolates to the genus Actinomadura. From their morphological, physiological, biochemical characteristics and DNA-DNA hy- bridisation data, the two isolates represented two new species in the genus Actinomadura, for which we propose A.glomerata sp. nov. (type strain, 1-226 = JCM 9376), and A.longicatena sp. nov. (type strain, I-497 = JCM 9377).

Authors' Abstract

No Abstracts were available for the following papers:

K.Ochi (National Food Research Inst., 2-1-2 Kannondai Tsukuba, Ibaraki 305, Japan). A phylogenetic study of the genus Arthrobacter by comparative ribosomal protein sequence analyses. Actinomycetologica, 9: 178-183

K.Ochi and Y.Inatsu (National Food Research Inst., 2-1-2 Kannondai, Tsukuba, Ibaraki 305, Japan). Comparative study of GTP content during growth of Streptomyces griseus and Streptomyces setae in relation to sporulation. Actinomycetologica, 9: 184-187, 1995

M.Kizuka, R.Enokita, N.Nakazawa and T.Okazaki (Tsukuba Research Labs, Sankyo Co., Ltd., 33, Miyukigaoka, Tsukuba-shi, Ibaraki, 305 Japan). Taxonomic studies of pentalenolactone- producing actinomycetes. Actinomycetologica, 9: 188-192, 1995

H.Seto (Inst.Molecular and Cellular Biosciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113, Japan). Biosynthesis of a natural C-P compound, bialaphos, produced by Streptomyces hygroscopicus. Actinomycetologica, 9: 193-205, 1995

T.Tamura (Inst. for Fermentation, Osaka, 2-17-85 Jusohonmachi, Yodogawa-ku, Osaka 532, Japan). Establishment of Kineococcus, Luteococcus, Catenuloplanes and Couchio- planes as new actinomycete genera including new species, and their relationships with related organisms. Actinomycetologica, 9: 206-215, 1995

S.Taguchi (Dept. Biological Science and Technology, Science University of Tokyo, 2641 Yamazaki, Noda-shi, Chiba 278, Japan). Communication between protease and protease inhibitor in the Streptomyces world. Actinomycetologica, 9: 216-227, 1995

Proceedings of SAJ Symposium on

Emerging Research Targets in Actinomycete Secondary Metabolism

(Tokyo, June 29, 1995)

T.Yamada, N.Ohara, N.Wada, S.Matsumoto, H.Yukitake, T.Matsuo, H.Kitaura, M.Takano, T.Nishiyama, H.Nomoguchi and M.Matsuo (Dept. Oral Microbiology, School of Dentistry, Nagasaki University, 1-7-1, Sakamoto, Nagasaki City, Nagasaki 852, Japan). Biochemical and immunological characterization of ribosomal fraction and culture filtrate from Mycobacterium. Actinomycetologica, 9: 228-235, 1995

A.Ishihama (Dept. Molecular Genetics, National Institute of Genetics, Shizuoka 411, Japan). Genetic strategies of bacteria for stationary-phase survival. Actinomycetologica, 9: 236-243, 1995

S.Horinouchi, T.Umeyama, H.Onaka, K.Ueda^1 and T.Beppu^1 (Dept. Biotechnology, Division of Agriculture and Life Sciences, Tokyo University, Bunkyo-ku, Tokyo 113 and ^1Dept. Ap- plied Biological Sciences, College of Agriculture and Veterinary Medicine, Nihon University, Fujisawa-shi, Kanagawa 352, Japan). Streptomyces species as boundary microorganisms: eucaryotic regulatory systems for secondary metabolism and morphological differentiation. Actinomycetologica, 9: 244-253, 1995

H.Osada (Antibiotics Lab., Inst. Physical and Chemical Research, RIKEN, Hirosawa 2-1, Wako-shi, Saitama 351-01, Japan). Fascinating bioactive compounds from actinomycetes. Actinomycetologica, 9: 254-262, 1995

SAJ REGULAR COLLOQUIA

13th SAJ Colloquium

Inst. of Microbial Chemistry, Tokyo, April 27, 1995

HOW NATURE SYNTHESIZES VITAMIN B12. A SURVEY OF THE LAST FOUR BILLION YEARS

A.I.Scott

Center for Biological NMR, Dept. of Chemistry, Texas A & M University, College Station, Texas 77843-3255, USA

Actinomycetologica, 9: S6, 1995

The acquisition and sequencing of the genes encoding the enzyme for vitamin B12 biosynthesis in Salmonella typhimurium and Pseudomonas denitrificans have dramatically altered the direction of research on the pathway from uro'gen III to the corrinoids. Using a combination of molecular biology, organic chemistry and NMR spectroscopy, logical progression along the sequence has been made. Recent work from our laboratory is focused on the discovery and specificities of the methyl trans- ferase connecting uro'gen III with cobyrinic acid, the temporal resolution of cobalt insertion and a comparison of the anaerobic and aerobic pathways in S.typhymurium and P.denitrificans respectively. The implication of two parallel routes to corrins in these bacteria will be discussed. The complete repertoire of gene products can be combined to reconstruct the biosynthetic pathway in vitro. Of great interest is the mechanism of ring contraction leading from porphyrinoids to corrins.

Author's Abstract

SITE-SPECIFIC RECOMBINATION OF THE ACTINOPHAGE R4

M.Shirai and M.Matsuura

Division of Biotechnology, School of Agriculture, Ibaraki University

Actinomycetologica, 9: S6-7, 1995

A temperate R4 phage isolated from soil was found to have a wide host range for Streptomyces species, and the viral DNA was linear, double-strand, and terminally redundant molecule of 53.7 kb. The R4 phage genome integrated site-specifically into the S.parvulus 2297 chromosome during the lysogenization process and integration occurred within 12-bp attachment core sequences (5'-GAAGCAGTGGTA3') on the phage genome and the host chromosome. To isolate a gene encoding a site-specific recombi- nase, the nucleotide sequence around the attP core region of the R4 phage was determined. An open reading frame (ORF469) was identified at 27bp downstream from the attP core site. The ORF469 is assumed to begin at ATG and terminate at TAG, and to encode a 469-amino acid polypeptide with a predicted molecular mass of 50.7 kDa. The predicted amino acid sequence of ORF469 showed a similarity to a resolvase of a Tn2501 and SpoVICA of Bacillus subtilis. To genetically confirm whether ORF469 is the structure gene for integration, the DNA fragments containing ORF469 and thiostrepton resistant gene were subcloned into pUC118. The resultant plasmids were transformed into S.parvulus, and the presence of the integration activity was tested by assaying thiostrepton resistance. The plasmids containing ORF469 were able to integrate, whereas plasmids without ORF469 were unable to. Furthermore, plasmids with site-directed mutations within ORF469 were unable to integrate. On the other hand, a gene disruption of ORF469 on prophage genome inhibited an excision of the pro- phage genome. These results suggest that the ORF469 encodes a site-specific recombinase essential for integration and excision of R4 phage genome.

Authors' Abstract

SEARCH FOR ANTITUMOR ANTIBIOTICS WITH SELECTIVE CYTOTOXICITY AGAINST TRANSFORMED CELLS

Y.Hayakawa

Inst. Molecular and Cellular Biosciences, University of Tokyo

Actinomycetologica, 9: S7, 1995

The retinoblastoma tumor suppressor protein (pRB) plays a central role in mammalian cell cycle control and is inactivated during the development of various human cancers. Human papillo- maviruses (HPV) encode E6 and E7 oncoproteins, which bind and inactivate the tumor suppressors pS3 and pRB, respectively. We established immortalized rat glia cell lines by transfection with HPV16 E6 and E7 genes. In the course of our screening of fermentation broths by using these cells, Streptomyces sp. R2827 was found to produce two new active substances. These metabolites were found to be new members of the leptomycin-an- guinomycin family by NMR spectral analysis and designated an- guinomycins C and D.

Anguinomycins A to D arrested the growth of normal glia cells without cytotoxicity and induced cell death against glia cells transformed with E7 or both E6 and E7 genes. Moreover, anguino- mycins induced growth arrest against normal rat 3Yl cells and ras-transformed cells, and caused cell death against 3Yl cells transformed with simian virus 40, adenovirus type 12 and its E1A gene. Cell lines highly sensitive to the killing effect of anguinomycins commonly express viral oncoproteins that can bind and inactivate pRB. Flow cytometric analysis revealed that anguinomycins arrested the cycle of 3Yl cells mainly in G1 phase.

Author's Abstract

14th Colloquium

National Institute of Health,

September 29, 1995

INDUCTION OF kinA EXPRESSION AND åH ACCUMULATION AT THE INITIATION OF SPORULATION IN B.SUBTILIS

F.Kawamura^1, K.Asai^2, T.Sebata^1, H.Yoshikawa^2 and H.Takahashi^2

^1 College of Science, Rikkyo University, ^2 Inst. Molecular and Cellular Biosciences, The University of Tokyo

Actinomycetologica, 9: S7, 1995

Induction of the Bacillus subtilis kinA gene, coding for a major kinase of the phosphorelay pathway, at the onset of sporulation required the spoOH gene but not the genes spoOA, spoOB and spoOF. Since spoOH gene codes for sH, which is necessary for the transcription of several early acting genes, including kinA gene, transcription of a spoOH-lacZ fu- sion and the levels of åH protein were examined in these spoO mutants. Whereas induction of spoOH transcrip- tion in spoOA, spoOB and spoOF mutants was appreciably inhibited, the level of åH protein in all of these mutants was increased at the onset of sporulation. These results indicate that the induction of spoOH transcription re- quires SpoOA-P and that accumulation of åH protein does not re- quire the products of spoOA, spoOB and spoOF. In addition, kinA expression was almost completely eliminated in a sporulation medium supplemented with 1% glucose and 0.1% glutamine, even though the usual stationary-phase associated in- crease in åH protein was observed under these conditions. It is thus most likely that the åH-dependent kinA in- duction is regulated by unknown factor(s) which may sense a subtle change of environmental conditions.

Authors' Abstract

NEUROKININ RECEPTOR ANTAGONISTS OF MICROBIAL ORIGIN

M.Nishikawa

Exploratory Research Labs., Fujisawa Pharmaceutical Co., Ltd.

Actinomycetologica, 9: S8, 1995

Increasing knowledge of the underlying pathophysiology of asthma has led to the hypothesis that axon reflex mechanisms are involved in the pathogenesis. Neurokinin such as substance P (SP) and neurokinin A (NKA) have been found in capsaicin sensitive C- fibre afferent to the lower airways of the respiratory system. SP and NKA released from C-fibre by way of axon reflex may result in bronchoconstriction, mucus hypersecretion and vascular leakage. Neurokinins may play an important role in the pathophys- iology of airway diseases, especially asthma. Neurokinin receptor antagonists may therefore be a useful targets for antiasthmatic drugs. In our search for activities inhibiting the binding of 3H- SP to guinea-pig lung membranes preparation, we found a fermentation product, WS 9326A, isolated from Streptomyces violaceusniger. In vitro, WS9326A selectively inhibited the binding of SP (NKl receptor) and NKA (NK2 receptor) to their re- ceptors. In vivo, the agent also inhibited both exogenous and indigenous neurokinininduced bronchoconstriction and vascular leakage in guinea pigs. We obtained tetrahydro WS9326A (FK224) which exhibits more potent biological activity than WS9326A. The catalytic hydrogenation of WS9326A produces FK 224. Pharmacological studies showed that FK224 was about ten times more active than WS9326A both in vitro and in vivo, and was also active as a dual NKl and NK2 neurokinin receptor antagonist. Recently, Takishima and coworkers have demonstrated that FK224 inhibited both bronchoconstriction and coughing in- duced by bradykinin in asthmatic patients. These findings suggest that FK 224 may be a useful tool for the study of the role of neurokinin in the pathology of asthma as well as a useful therapeutic agent.

Author's Abstract

A NEW POTENT ANGIOGENESIS INHIBITOR OF MICROBIAL ORIGIN

T.Shibata

Exploratory Research Labs., Fujisawa Pharmaceutical Co., Ltd.

Actinomycetologica, 9: S8-9, 1995

Angiogenesis is the process of new blood vessel formation by the endothelial cells and often associated with diseases, such as cancer, diabetic retinopathy, and rheumathoid arthritis. It is well known that the growth of solid tumors depends on angiogenesis and that angiogenesis plays an important role in tumor metastasis. Therefore, angiogenesis inhibitors might be beneficial drugs for treatment of cancer.

During the course of a screening for new angiogenesis inhibitors from microbial products, we found that a fungus Scolecobasidium arenarium F-2015 produced a new angiogenesis inhibitor, FR 111142. FR 111142 inhibited the growth of endothelial cells at low concentrations and displayed potent angiogenesis inhibitory activity in vivo on the chick em- bryo chorioallantoic membrane. FR 111142 also suppressed the murine solid tumors MethA and colon38, but not active against murine leukemia P388 in ascites form, the growth of which does not require angiogenesis. In addition, FR 111142 had a much weaker inhibitory activity against the growth of such tumor cells than endothelial cells in vitro. These results suggest that the potent antiangiogenic and antitumor activities of FR 111142 may be due to its antiangiogenic action, probably by the inhibition of endothelial cell growth.

This discovery of FR111142 led us to search for derivatives with more potent antiangiogenic activity and weaker side effect than the parent compound FR 111142. We chemically synthesized many derivatives of FR 111142 and tested for growth inhibitory activity against endothelial cells, and selected FR 118487. In the endothelial cell growth assay in vitro and the angiogenesis assay in vivo on the chick embryo chorioallantoic membrane, FR118487 had about 5-10 times more potent antiangiogenic activity than FR111142. In addition, FR118487 inhibited neovascularization in the rabbit comea induced by several angiogenic stimuli. FR118487 also suppressed the growth of a wide variety of solid tumors in mice. These facts suggest that FR 118487 would be a new type of drug for treatment of solid tumors.

Author's Abstract

THE MOLECULAR PHYLOGENY AND SYSTEMATICS OF THE ACTINOBACTERIA

A.Yokota

Inst. Molecular and Cellular Biosciences, The University of Tokyo

Actinomycetologica, 9: S9, 1995

Recent progress of the systematics of actinobacteria was reviewed. Taxonomic studies during recent three years on actinobacteria added many new taxa, including new genera of cocci such as Kineococcus, Luteococcus, Microluna- tus, and Pelczaria, and new genera of rods such as Dietzia, Propioniferax, Rathayibacter, Tricella, "Leucobacter", and "Oza- leobacterium", as well as 39 new species or new combinations in 14 genera.

Phylogenetic relationships based on 16S rRNA sequences revealed the phylogenetic intermixing among many taxa of actinobacteria created based mainly upon the phenotypic characteristics, es- pecially on the combinations of chemotaxonomic characteristics such as peptidoglycan types, menaquinone compositions, polar lipids, etc. As examples of such intermixing genera, species of the genera Arthrobacter and Micrococcus, and those of Aureobacterium and Microbacterium, were demonstrated, and the 16S rRNA studies strongly suggest necessity of taxonomic reconstruction of these genera. These examples illustrates that peptidoglycan composition is not a reliable indicator per se of phylogeny.

Author's Abstract

SOME MICROBIAL AND BIOCHEMICAL CHARACTERISTICS OF ACIDIC TEA FIELD SOIL

I.Nioh

Faculty of Agriculture, Shizuoka University

Actinomycetologica, 9: S9, 1995

Tea field soils are generally acidic due to the heavy applying of ammonium fertilizer. Tea plant absorbs ammonium nitrogen actively in its growing stage, and as a results in organic anions such as sulfate accumulates in the soil. These soils show peculiar biochemical and microbial characteristics. For instance, active nitrifying activity of ammonium ions found in these soils irrespective of general recognition that nitrification occurs mostly in neutral or slightly alkaline soils. An acid-tolerant autotrophic nitrifying bacterium has been isolated. Recently nitrous oxide, well known as a gas involving earth's warming and destroying ozone layer, was found to evolve from the strongly acidic tea field soils. We have isolated several kind of fungi as the agents of nitrous oxide production.

Determination of the number of microorganisms in a tea field soil from the experimental field of the National Institute of Vegetable, Ornamental Plants and Tea, Shizuoka prefecture, showed a relative abundance of actinomycetes. Eighty-five % of the actinomycete isolates were acidophilic, and 15% were acid tol- erant. Identification of these isolates by means of chemical analysis of the cell components together with morphological observation revealed that genus Streptomyces or its related genera were only 23%. Most of this group were acid-toler- ant. Other 77% had meso- and 3-OH type diaminopimelic acid and whole cell sugar type A and D. All of the actinomycetes of this group were acidophilic. Besides other groups of microorganisms, these results indicated that the acidic tea field soil sustains a characteristic actinomycete flora.

Author's Abstract

Copyright 1996 C.E.T.A., The International Centre for Theoretical and Applied Ecology, Gorizia