• Microbiology and Biotechnology Letters
• /
• v.32 no.3
• /
• pp.282-285
• /
• 2004

The polyene antibiotics including nystatin, pimaricin, amphotericin and candicidin are a family of most promising antifungal polyketide compounds, typically produced by rare actinomycetes species. The biosynthetic gene clusters for these polyenes have been previously investigated, revealing the presence of highly homologous biosynthetic genes among polyene-producers such as polyketide synthase (PKS) and cytochrome P450 hydroxylase (CYP) genes. Based on amino acid sequence alignment among actinomycetes CYP genes, the highly-conserved regions specific for only polyene CYP genes were identified and chosen for degenerate PCR primers, followed by the PCR-screening with various actinomycetes genomic DNAs. Among tested several polyene non-producing actinomycetes strains, Pseudonorcardia autotrophica strain was selected based on the presence of PCR product with polyene-specific CYP gene primers, and then confirmed to contain a cryptic novel polyene hydroxylase gene in the chromosome. These results suggest that the polyene-specific hydroxylase gene PCR should be an efficient way of screening and isolating potentially-valuable cryptic polyene antibiotic biosynthetic genes from various microorganisms including rare actinomycetes.

• Journal of Microbiology
• /
• v.39 no.1
• /
• pp.17-23
• /
• 2001

Various pretreatment procedures and selective media were applied to assess the optimal conditions for the isolation of rare actinomycetes from soil. Pretreatment of wet-heating for 15 min at 70$^{\circ}C$ and phenol treatment of soil suspension were the most effective methods for the isolation of these microorganisms. Hair hydrolysate vitamin agar (HHVA) was the most suitable medium for the recovery of rare actinomycetes. Thirty-five rare actinomycete strains were chosen using selective isolation approaches, then morphological and chemical properties of the isolates were determined. The isolates belonged to one of the following genus, Micromonospora, Microbispora, Actinoplanes and Streptosporangium.

• Microbiology and Biotechnology Letters
• /
• v.28 no.3
• /
• pp.129-133
• /
• 2000

Different actinomycete strains were isolated from natural lime caves of Ondal Chemongok Hwanseon and Yongyeon which are located at Kangwon or chungcheongbook province in Korea and were identified to the genus level. Soil sam-ples were collected at 6 sites inside and 2 sites outside of each natural lime cave, As the result the strains belonging to genus Streptomyces and rare actinomycetes were isolated at the average of 2.1 and 3.4 strains per g soil on inside cave whereas which were isolated at the 6.0 and 1.8 strains per g soil on outside cave. How-ever the generic distribution of Streptomyces and rare actinomycetes isolated from outside cave was quite dif-ferent from that of inside cave. It was shown that rare actinomycetes at natural lime caves is generally highly abundant than Streptomyces.

• Microbiology and Biotechnology Letters
• /
• v.27 no.6
• /
• pp.515-517
• /
• 1999

From the soils collected seasonally at the 0.5~2cm and 50$\pm$1cm depths of riverside, different strains of actinomycetes were isolated and identified to the genus. At the 0.5~2cm depth, Streptomyces and rare actinomycetes were in total 73 and 62 strains, respectively. Streptomyces was approximately 2-fold more in spring and autumn than summer, and rare actinomycetes was at least 4-fold more in autumn and winter than spring. At the 50$\pm$1cm depth, Streptomyces and rare actinomycetes were in total isolated 53 and 41 strains, and these were at least 2-fold more diverse in autumn than spring, summer, and winter.

• Microbiology and Biotechnology Letters
• /
• v.22 no.2
• /
• pp.222-225
• /
• 1994

Three kinds of pretreatment methods were used for selectivel isolation of soil actinomy cetes. One hundred and six strains were isolated on Bennet's agar and 114 strains were on humic acid-vitamin agar from 5 domestic soil samples. All of these isolates were identified to the genus level based on morphological and physiological characheristics. Among three methods, dry heat was most effective to isolate many different strains including rare actinomycetes. On genus, Bennet's agar was effective for selective isolation of Nocardia but bumic acid-vitamin agar for another rare actinomycetes.

• Microbiology and Biotechnology Letters
• /
• v.27 no.4
• /
• pp.273-277
• /
• 1999

From each sediments collected at Chudong(static) and Hoenam(streaming) site of Lake Daechung on May 18(before rainy season) and on August 24 in 1998(after rainy season), different strains of Actinomycetes were isolated and identified to genus level. For comparison, the genus diversity of Actinomycetes in control soils such as grass land around lake, lake shore, and shallow(5-10cm) lake was also investigated. In consequence, the isolation of Streptomyces from the sediments of Lake Daechung was the most copious(41 strains) at the streaming site before rainy season and the least(27 strains) at the static site before rainy season, which were 2.4- and 1.6-fold larger than the average(17 strains) of control soils, respectively. In addition, the isolation of rare Actinomycetes from the sediments of Lake Daechung was the most copious(23 strains) at the streaming site before rainy season and the least(12 strains) at the static site after rainy season, which were 3.2-and 1.6-fold larger than the average(17 strain) of controls, respectively. Therefore, it is considered that lake sediments exhibit higher diversity of Streptomycete as well as rare Actinomycetes than general soils, and thereby can be utilized as useful sources to isolate diverse Actinomycetes.

• Microbiology and Biotechnology Letters
• /
• v.23 no.1
• /
• pp.36-42
• /
• 1995

Actinomycetes occur in a wide range of environments and many more actinomycetes remain to be detected in the natural environment. In isolation stages, selection of the environment as a source of useful isolates is important. Two hundred and eighteen strains were isolated on Bennet's agar and 346 strains were on humic acid-vitamin agar from five each paddy field, field, forest, grass land, riverside soil samples. These isolates were identified to the genus level based on morphological and physiological characteristics. Among them, 386 strains were Streptomyces, 49 strain were Nocardia, 35 strains were Microbispora and Micromonospora each, 15 strains were Nocardiopsis, 13 strains were Actinomadura, 10 strains were Streptosporangium, and the others were isolated rarely. According to soil type, Nocardia, Micromonospora, Microbispora and Streptosporangium were dominant in paddy field, Microbispora Nocardia, Nocardioides and Micromonospora were dominant in field, Nocardia, Micromonospora, Microbispora and Actinomadura were dominant in grass land, Nocardia, Micromonospora and Microbispora were dominant in forest, Nocardia, Microbispora and Micromonospora were dominant in riverside. Generally, Nocardia, Micromonospora, Microbispora and Actinomadura were isolated in all kinds of soils, Streptosporangium were paddy field, Dactylosporangium were forest, Nocardiopsis were field, forest and riverside.

• Journal of Microbiology and Biotechnology
• /
• v.24 no.9
• /
• pp.1226-1231
• /
• 2014

The rare actinomycete Pseudonocardia autotrophica was previously shown to produce a solubility-improved toxicity-reduced novel polyene compound named $\underline{N}ystatin$-like $\underline{P}seudonocardia$ $\underline{P}olyene$ (NPP). The low productivity of NPP in P. autotrophica implies that its biosynthetic pathway is tightly regulated. In this study, $wblA_{pau}$ was isolated and identified as a novel negative regulatory gene for NPP production in P. autotrophica, which showed approximately 49% amino acid identity with a global antibiotic down-regulatory gene, wblA, identified from various Streptomycetes species. Although no significant difference in NPP production was observed between P. autotrophica harboring empty vector and the S. coelicolor wblA under its native promoter, approximately 12% less NPP was produced in P. autotrophica expressing the wblA gene under the strong constitutive $ermE^*$ promoter. Furthermore, disruption of the $wblA_{pau}$ gene from P. autotrophica resulted in an approximately 80% increase in NPP productivity. These results strongly suggest that identification and inactivation of the global antibiotic down-regulatory gene wblA ortholog are a critical strategy for improving secondary metabolite overproduction in not only Streptomyces but also non-Streptomyces rare actinomycete species.

• 한국생물공학회:학술대회논문집
• /
• 2005.04a
• /
• pp.396-397
• /
• 2005

The polyene antibiotics are a family of most promising antifungal polyketide compounds, typically produced by actinomycetes species. Using the polyene CYP-specific PCR screening with served actinomycetes genomic DNAs, Pseudonocardia autotrophica strain was identified to contain a unique polyene-specific CYP gene. The genomic DNA library screening using the polyene-specific CYP gene probe revealed the positive cosmid clone containing an approximately 34.5 kb DNA fragment revealed a total of seven complete and two incomplete open reading frame (ORFs), which are highly homologous but unique to previously-known polyene biosynthetic genes. These results suggest that the polyene-specific screening approach should be an efficient way of isolating potectially-valuable cryptic polyene biosynthetic gene cluster from various rare actinomycetes.

• Journal of Microbiology and Biotechnology
• /
• v.15 no.1
• /
• pp.188-191
• /
• 2005

Abstract Cyclosporins are a family of clinically-important immunosuppressive cyclic peptides produced by Tolypocladium inflatum. The structural modification of cyclosporins via hydroxylation at various positions of N-methyl leucines in cyclosporin A leads to a dramatic change of their bioactive spectra. Among over 100 soil actinomycetes screened, two actinomycetes species, Sebekia benihana and Pseudonocardia autotrophica, were identified to contain superior cyclosporin A hydroxylation activities. A HPLC-based cyclosporin A hydroxylation assay revealed that each strain possesses distinctive hydroxylation specificity and regiospecificity; mono-hydroxylation at the 4th N-methyl leucine of cyclosporin A by S. benihana, and di-hydroxylations at both 4th and 9th N-methyl leucines of cyclosporin A by P. autotrophica. The conversion yields for cyclosporin A hydroxylation by both S. benihana and P. autotrophica were significantly improved from less than 10% and 18% up to 58% and 45%, respectively, in the optimized culture containing molybdenum with 0.05 g/l of cyclosporin A concentration. An ancymidol-specific inhibition of cyclosporin hydroxylation also suggested that the regiospecific cyclosporin hydroxylation might be catalyzed by a putative cytochrome P450 mono-oxygenase enzyme.