New methods for the highly selective isolation of Streptosporangium and Dactylosporangium from soil

Abstract

Two methods for the isolation of the rare actinomycetes Streptosporangium and Dactylosporangium in soil, are described. The methods use the ability of both the sporangiospores of Streptosporangium and the globose bodies (aleuriospore) of Dactylosporangium to withstand dry heating and treatment with benzethonium chloride (BC). In addition, the differential antibiotic resistance of these actinomycetes is also utilized. (i) To isolate streptosporangia, an air-dried soil sample is first subjected to dry heat treatment (120°C, 1 h). A water suspension of the heated sample is then treated with 0.01% BC, diluted and cultured on HV agar supplemented with nalidixic acid and leucomycin. (ii) To isolate dactylosporangia, a water suspension of a soil sample given dry heat and 0.03% BC treatment is cultured on HV agar supplemented with nalidixic acid and tunicamycin. The dry heat and BC treatment drastically eliminated bacteria and unwanted actinomycetes contaminants, including streptomycetes, from the isolation plates, thereby facilitating the selective isolation of streptosporangia and dactylosporangia. Nalidixic acid in HV agar was useful to suppress the growth of dry heat- and BC-resistant bacteria. The use of leucomycin and tunicamycin increased the selectivity of HV agar for streptosporangia and dactylosporangia, respectively, through elimination of the growth of unwanted actinomycetes which remained after the employment of pretreatments. From various field soils (vegetable, orchard, corn, and rice paddy), the methods described above consistently achieved the highly selective isolation of streplosporangia and dactylosporangia, respectively, as each group accounted for more than 20% of the total number of colonies recovered.