During the life cycle of the streptomycetes, large numbers of hyphae die; the surviving ones undergo cellular differentiation and appear as chains of spores in the mature colony. Here we report that the hyphae of Streptomyces antibioticus die through an orderly process of internal cell dismantling that permits the doomed hyphae to be eliminated with minimum disruption of the colony architecture. Morphological and biochemical approaches revealed progressive disorganization of the nucleoid substructure, followed by degradation of DNA and cytoplasmic constituents with transient maintenance of plasma membrane integrity. Then the hyphae collapsed and appeared empty of cellular contents but retained an apparently intact cell wall. In addition, hyphal death occurred at specific regions and times during colony development. Analysis of DNA degradation carried out by gel electrophoresis and studies on the presence of dying hyphae within the mycelium carried out by electron microscopy revealed two rounds of hyphal death: in the substrate mycelium during emergence of the aerial hyphae, and in the aerial mycelium during formation of the spores. This suggests that hyphal death in S. antibioticus is somehow included in the developmental program of the organism.

Cellophane cultivation techniques have been proven to be useful for the study of colony growth in Streptomyces. Results obtained by this procedure indicate that, in S. antibioticus, substrate mycelium was a nutrient support for aerial mycelium growth. Oleandomycin synthesis starts before aerial mycelium formation and may play an important role during colony growth.

203During germination, Streptomyces antibioticus arthrospores passed through three stages : darkening, swelling and germ tube emergence. The first stage, darkening, whose main features were a decrease in absorbance and a loss of refractility, only required exogenous divalent cations (Ca2+, Mg2+ or Fe2+) and energy that can be obtained from the spore reserves. This stage was blocked by agents that inhibit ATP formation but not by antibiotics that inhibit macromolecular synthesis. The second stage, swelling, needed an excgenous carbon source and was not blocked by mitomycin C. In this stage, the spores exhibited the highest cytochrome oxidase and catalase activities and respiratory quotient. The last stage, germ tube emergence, required additional carbon and nitrogen sources. Ammonium compounds were superior to nitrate. Dry weight remained constant during the stages of darkening and swelling, with a rapid increase from the moment of germ tube emergence. Optimum pH and temperature for germination were 8.0 and 45 "C, respectively. Heat treatment (55 "C for 10 min) had no effect on germination.The fine structure of the spore underwent important changes during germination. The wall of the swollen spore became stratified and the inner layer was continuous with the germ tube wall.Macromolecular synthesis occurred in the sequence RNA, protein and then DNA. Rifampicin, streptomycin and mitomycin C prevented synthesis when added at the start of incubation. The same effect was obtained if the addition was made during germination, except with mitomycin C which inhibited DNA, but not RNA and protein synthesis.

This is the first study of sporogenesis in Streptomyces carried out on a relatively high number of species (seven) which allows us, using also previously published results, to establish a general picture of this process. In the sporogenesis of Streptomyces two basic stages can be considered: the sporulation septum synthesis and the arthrospore maturation. Our ultrastructural study of the sporulation septum formation suggests the existence within this genus of three basic types. Type I is distinguished because the septum is formed from the beginning by two separate cross walls. Within this type we include Streptomyces erythraeus, Streptomyces albus, and Streptomyces aureofaciens and also include Streptomyces venezuelae, Streptomyces griseus, and Streptomyces osteogriseus. Type II is distinguished because there is a deposit of material previous to the synthesis of the double annulus which completes the septum. This type can be divided into two subtypes. In the first the deposits are wedge-shaped and the double annulus is clearly visible, and to this group belong Streptomyces flaveolus, Streptomyces ambofaciens, and Streptomyces coelicolor. In the second the deposits, which have a different shape and are very well developed, constitute almost entirely the sporulation septum in which the double annulus is barely visible; Streptomyces antibioticus and also Streptomyces viridochromogenes belong to this group. Type III, represented by Streptomyces cinnamonensis, is distinguished because the septum is formed by a single cross wall.The genus Streptomyces belonging to the order Actinomycetales shows a comparatively complex development cycle in which from a branched vegetative mycelium submerged in the substrate is formed an aerial mycelium with hyphae of greater diameter. Through the sporulation process these aerial hyphae give rise to chains of arthrospores, which placed under favorable conditions germinate and repeat the cycle.The term arthrospores has been proposed by Cross (5) to refer to the asexual spores produced by Streptomyces that are formed by the annular ingrowth of specialized, regularly spaced cross walls in existing hyphae enclosed in a fibrQus sheath. The arthrospores are differentiated from bacterial endospores both by the sporogenesis process and by their properties.In recent years great interest has been shown in the study of sporulation processes in bacteria, because they constitute a simple, and therefore an attractive, model for the study of a process of cellular differentiation. Although most of these studies are performed on species of two genera Bacillus and Clostridium, several works have been published on the different processes of sporulation within the Actinomycetes, particularly in the genera Streptomyces and Thermoactinomyces (6).In this paper we present the results of the ultrastructural study of the sporulation in seven species of Streptomyces. We compare these results with those obtained previously by other authors and make a tentative classification of the genus Streptomyces according to ...

The intracytoplasmic accumulation of polysaccharides has been demonstrated in four species of Streptomyces. After extraction and purification, the structural properties of these polysaccharides were investigated by chemical, spectrophotometric, and enzymatic methods. The results clearly indicated that the streptomycete polysaccharides have a degree of branching and a macromolecular structure similar to that of the glycogens isolated from other bacteria and animal sources.