Morphological and cultural characteristics, and levels of DNA-DNA relatedness of Streptomyces coelicolor A3(2) and S. lividans 66 were examined and compared with those of S. coelicolor Müller, S. lividans ISP 5434 and S. violaceoruber ISP 5049 to clarify their taxonomic status. Results reveal that S. coelicolor A3(2) is distinctly different from the type strain S. coelicolor Müller, but closely related to S. violaceoruber ISP 5049. In addition, S. lividans 66 closely resembles S. lividans ISP 5434 and S. violaceoruber ISP 5049 by the same taxonomic criteria. These results indicate that S. coelicolor A3(2), S. lividans 66 and S. lividans ISP 5434 are closely related to S. violaceoruber.
An acidic solution (pH 2.5∼2.6) with a high oxidation-reduction potential (ORP; about +1,170 mV) and an alkaline solution (pH 11.5∼11.7) with a low ORP (about −880mV) that resulted from electrolysis of 20 mM NaCl (dissolved in a pure water) were tested for their effect on the growth of Streptomyces spp. When spores (∼2×107) were exposed to the electrolyzed solutions (2 ml) for 1 minute, colony formation was totally inhibited by the acidic solution, but little by the alkaline solution although extending the exposure (10 minutes) resulted in a marked inhibition. The 1 minute exposure to their mixture (1:1, v/v) showed a strong inhibition (but weaker than that of the acidic solution). When the unexposed spores were streaked and incubated on ISP No. 4 (inorganic salts - starch medium) agar plate containing a cross density gradient of the acidic and alkaline solutions, a biased growth inhibition toward the acidic solution side was observed although the pH range of the acidic solution end of the plate was around 6.2. It seemed thus unlikely that low pH value contributed to the antimicrobial activity of the acidic solution. It was notable that S. griseus SS-1198 formed a unique morphology on the cross gradient plate. In addition, clear growth inhibition by the acidic solution was observed without direct contact with spores, probably because of chlorine gas release. Acidic solutions (pH 2.6∼2.7) resulting from the electrolysis of 20 mM of Na2SO4 show no significant antimicrobial activity when tested by the cross gradient plate method. It thus seemed likely that chlorine played a key role for the antimicrobial activity of the acidic electrolyzed NaCl solution.
Methods for the selective isolation of actinomycetes from pine litter layers were evaluated. Pretreatment of source material with a 0.05% of sodium dodecyl sulfate solution and the addition of 20 μg/ml of nalidixic acid to the agar media were effective in reducing bacterial growth. The addition of 200 μg/ml of benlate to the agar medium was effective in reducing fungal growth. By removing actinomycetes from the surface of pine needles with the surface sterilization method by 70% of ethanol and sodium hypochlorite solution containing 1% of chloride, we were able to isolate those actinomycetes which reside under the waxy layer of needles. Comparative studies of isolates obtained from pine litter layers showed both a qualitative and quantitative difference by layers.
The linearity of the chromosome has been observed in several Streptomyces species, and thus may be characteristic of the genus. The termini of the chromosomes contain inverted repeats, and carry covalently-bound proteins at the 5’ end, presumably the primer for replication. The terminal inverted repeats of the Streptomyces lividans chromosome are about 30 kb long, of which the extreme 16 kb is indistinguishable from that of the right end of the 50-kb linear plasmid SLP2. The terminal sequence of S. lividans exhibits abundant palindromes, resembling those of several linear Streptomyces plasmids for the first 12∼17 bp. In spite of this, the terminal DNA sequences among different species are not highly conserved over long distances. A functional oriC has been located at the center of the chromosomes of Streptomyces coelicolor and S. lividans. These chromosomes appear to replicate bi-directionally from the center and are presumably patched up at the ends by protein-primed replication. At least in S. lividans, the telomeres are dispensable and the chromosome could be circularized in viable cells by targeted recombination, which removed the telomeres and joined the two arms. The circularized chromosomes were unstable, undergoing further rearrangements, and resulted in relatively poor growth and sporulation. Long stretches of DNA on various Streptomyces chromosomes are prone to spontaneous deletions and amplifications. In S. lividans and S. coelicolor, the unstable regions correspond to the termini. Moreover, circularized chromosomes, containing deletions of several hundreds of kb in the terminal regions, were found in some spontaneous deletion mutants of S. lividans ZX7. These observations indicate the readiness of the linear chromosome to be circularized, and implicate the termini in the structural instability of Streptomyces chromosomes.
Streptomyces coelicolor A3(2) and S. lividans 66 were examined for their morphological and cultural characteristics as well as DNA-DNA relatedness in comparison with those of S. coelicolor Müller, S. lividans ISP 5434 and S. violaceoruber ISP 5049 in order to clarify their taxonomic status. It turned out that S. coelicolor A3(2) was distinctly different from the type strain S. coelicolor Müller, but closely related to S. violaceoruber ISP 5049. On the other hand, S. lividans 66 was found to have taxonomic characteristics that closely resemble those of S. lividans ISP 5434 and S. violaceoruber ISP 5049. These results indicated that S. coelicolor A3(2), S. lividans 66 and S. lividans ISP 5434 were all closely related to S. violaceoruber. For details, see the review on pp. 47-50 of this issue of ACTINOMYCETOLOGICA.
The nomenclature of Streptomyces kasugaensis that produces kasugamycin has not been approved taxonomically, although it was approved as one of the Streptomyces hosts in the Japanese guideline for recombinant DNA experiment. In order to establish its taxonomic status, we examined two strains, M338-M1 and MB273-C4, of S. kasugaensis for their taxonomic properties; not only conventional characterization such as morphology and physiology, but also analyses of cell wall type and DNA. Consequently, the following results were obtained. Both strains, M338-M1 and MB273-C4, developed spiral aerial mycelium which at maturity formed chains of 10∼50 spores with smooth surface. Aerial mass color of their surface growth on agar media was Gray series color. The type of 2,6-diaminopimelic acid in the cell wall was LL-type. These characteristics indicated that they were regarded as Streptomyces. Based on the data obtained, we searched for candidate species that have similar characteristics by the aid of a data base of actinomycetes. It turned out that S. xantholiticus was selected as the one with similarity to the strains of S. kasugaensis. Subsequently, these two strains were compared with S. xantholiticus IMC S-0620 (=ISP 5244) in terms of morphology, physiology, chemical components of cell wall, DNA homology etc. Although detailed data were not elaborated here, there was a big difference allowing to distinguish S. kasugaensis from S. xantholiticus. Furthermore, DNA homology was high (77% or higher) between S. kasugaensis strains, and low (21% or lower) between S. kasugaensis and S. xantholiticus. Another clear difference was observed in the RAPD (random amplified polymorphic DNA) profiles. Based on these, we propose that Streptomyces kasugaensis can be regarded as an independent or new species. Details of taxonomic characterization of S. kasugaensis strains will be described in the next volume of ACTINOMYCETOLOGICA.