A reproducible and reliable protoplasting and regeneration condition for the genus Actinomadura strains, which is extraordinarily resistant to lysozyme, has been studied. Protoplasts were prepared from all strains examined by use of both lysozyme and N-acetylmuramidase only when they were grown in the presence of sucrose (10∼30%) and glycine (0.3%). The optimum concentration of sucrose was strain-dependent. Protoplasts could regenerate on the modified R5 medium when the sucrose concentration was the same as that of culture medium used. Thus established regeneration medium provided us with regeneration ratios of 0.4 to 30%.
We isolated a kasugamycin-producing actinomycete, strain MF861-C4, which had spiny spore surface. This strain was different from Streptomyces kasugaensis but rather close to Streptomyces albulus with regard to morphological, cultural, and physiological characteristics, DNA homology and partial base sequences of 16S ribosomal RNA. The strain MF861-C4 was identified as S. albulus.
Effect of some inorganic salts, carbon and nitrogen sources on panose production in the former medium1) by strain GT-32 was investigated. In the medium when 0.3% MnSO4·4H2O was added after removing NH4Cl and NaCl, the strain gave a higher panose production (relative productivity; around 610%) than that of the former medium studied. With the optimum fermentation conditions for panose production, the highest production titer (9.8 mg/ml) was given in 5 days cultured broth.
The 16S rRNA gene sequences of 18 strains of Streptomyces lavendulae and Streptomyces virginiae were determined. Fifteen strains including the type strains of S. lavendulae and S. virginiae had 1,514 nucleotides in positions 10 to 1,541 of the Escherichia coli numbering system, and their similarities were more than 98.1%. On the other hand the sequences of S. lavendulae subsp. lavendulae IFO 3361, S. lavendulae subsp. lavendulae IFO 12341 and S. virginiae ATCC 13013 consisted of 1,516 nucleotides. The differences in the number of nucleotides were observed between 3 strains containing 1,516 nucleotides and 15 strains containing 1,514 nucleotides in positions 181 to 219,1,133 to 1,141 and 1,446 to 1,456 (E. coli numbering system) and various sequence lengths of these areas were corresponded to the similarity level of the 16S rRNA gene sequence. The homology values of DNA-DNA hybridization between these three strains and their type strains were very low (15 to 22%) and these three strains were located in a cluster far from the other strains in the cluster analysis based on the taxonomic characteristics (Anzai, Y.; T. Okuda & J. Watanabe, J. Antibiot. 47: 183-193,1994). Therefore, we conclude that S. lavendulae subsp. lavendulae IFO 3361 and IFO 12341, and S. virginiae ATCC 13013 should not be assigned to S. lavendulae or S. virginiae.
Screening of known and unidentified actinomycetes revealed three strains which can hydroxylate compactin to pravastatin. They showed taxonomic properties which are consistent with those of Actinomadura sp., Streptomyces tanashiensis and Streptomyces anulatus. Actinomadura sp. strain 2966 effected a 60% bioconversion of added compactin.