Ultrastructural studies of sporulation in Bacillus sphaericus.

Spore septum formation in Bacillus sphaericus 9602 occurs 2 h after the end of exponential growth at one end of the vegetative cell, which retains a uniform diameter. The apparently rigid spore septum contains an inner cell wall layer which disappears when the sporulation septum "bulges" into the mother cell cytoplasm. This process occurs simultaneously with terminal swelling at the end of the cell containing the spore septum. It is suggested that the inner cell wall layer is peptidoglycan and that its dissolution and the terminal swelling are consequences of a localized autolysis. Engulfment of the forespore by membrane proliferation results in the production of a forespore surrounded by two flexible, closely apposed membranes. These membranes appear to become more rigid as a peptidoglycan-like layer appears between them, concomitant with the condensation of the forespore nucleoid into a crescent-shaped structure. After nuclear condensation, visible development of distinct cortex, primordial cell wall, and spore coat layers begin, and the forespore cytoplasm assumes an appearance similar to that of a refractile spore. The spore coats consist of an amorphous inner layer, a lamellar midlayer, and a structured outer layer. As cortex synthesis and spore coat assembly continue, exosporium development commences close to that portion of the mother cell plasma membrane which surrounds the forespore. The exosporium is lamellar and in tangential section is seen to have a hexagonal arrangement of subunits. The timing of these morphological events has the expected correlation with the appearance of unique enzyme activites required for cortex synthesis.     

Microscopic observations of germination and septum formation in pycnidiospores of Botryodiplodia theobromae

A population of aseptate pycnidiospores of the fungus Botryodiplodia theobromae can be induced to germinate or to form septa delimiting two cells; this developmental process is dependent upon nutritional and environmental factors. Transmission electron microscope investigations indicate that during germination of the aseptate spore, a new inner wall layer is synthesized de novo at the site of germ tube emergence. Formation of the septum also involves the de novo synthesis of an inner wall layer which comprises the majority of the septum and completely surrounds the spore. The wall of the germ tube emerging from the septate spore is a direct extension of this inner layer deposited during the formation of the septum. Although the early stages of spore germination may involve localized enzymatic degradation of the internal layers of the spore wall, transmission and scanning electron micrographs of germinating spores show that the outer wall layers are physically fractured by the emerging germ tube. It is suggested that spore germination and septum formation are initially similar processes regarding cell wall genesis but that some mechanism responsive to environmental and nutritional conditions determines the course of development.     

Sporogenesis in Streptomyces melanochromogenes

The mode of spore differentiation in a strain of Streptomyces melanochromogenes was followed by analysis of ultrathin sections of sporulating aerial hyphae at various stages of sporogenesis. A special accent was laid on the formation of the sporulation septum and its alterations in the course of spore delimitation and separation. Distinct differences in formation and substructure have been observed between the cross walls of vegetative hyphae and the sporulation septa.Cross walls of vegetative hyphae are formed in a way typical for Gram-positive bacteria by a centripetal annular ingrowth of cytoplasmic membrane, on which wall material immediately is deposited. The development of the sporulation septa is characterized by the accumulation of amorphous material in addition to the newly synthesized wall layer inside the invaginating cytoplasmic membrane. This amorphous septal material will later be decomposed presumably by two lytic systems which cause the separation of the spores. The central region of the finished sporulation septum is perforated by microplasmodesmata. Spores are released by a break down of the surface sheath. The complete spores are enveloped by a twolayered cell wall and the spiny surface sheath.     

ELECTRON MICROSCOPY OF PHIALIDES AND CONIDIOGENESIS IN TRICHODERMA SATURNISPORUM

Each phialide had a thick-walled neck region located immediately below a light microscopically inconspicuous collarette. The thickened wall of the phialide neck was multilaminate, with layers of different electron transmission properties. A developmental stage in the formation of the first conidial initial was observed. Conidial initials blew out through the thickened neck region, increased in size, and were eventually delimited by centripetally developing septa. Mature, winged conidia had an electron-opaque outer wall layer and an electron-transparent inner wall layer. The wing was formed by separation of these outer and inner wall layers and buckling or wrinkling of the outer layer. As early as they could be discerned, conidial initials had developed the electron-opaque wall layer which characterized mature conidia. Each conidium-delimiting septum became bilayered; the upper layer formed part of the conidial base, and the lower layer became a portion of the wall of the next conidial initial. Phialides lacked an electron-opaque wall layer, and they possessed areas of abundant rough endoplasmic reticulum, as well as free ribosomes. Lipid globules were also abundant, especially in conidia. The distinction between phialides and annellides was questioned.     

PARTICIPATION OF THE CYTOPLASMIC MEMBRANE IN THE GROWTH AND SPORE FORMATION OF BACILLI

A polyester embedding technique was used to study the early stages of spore formation in members of the genus Bacillus in order to investigate further the origin and nature of the initial spore septum and the resulting forespore envelope. Whereas previously, with a methacrylate procedure, this layer had appeared to be continuous with the cell wall, this study reveals it as a double layer of cytoplasmic membrane. Perisporal, membranous organelles connected both to the developing forspore envelope and to the cytoplasmic membrane were encountered in the four species studied. Similar organelles were prominent during growth at the sites of transverse septa formation. These were connected to, or continuous with, the cytoplasmic membrane and often adherent to the chromatin bodies of the dividing bacilli.     

Cell division in baeocyte producing cyanobacteria

Summary An ultrastructural examination of cell division in two baeocyte producing cyanobacteria,Pleurocapsa minor andDermocarpa violaceae, reveals two distinct patterns of binary (transverse) fission. Septate binary fission, inPleurocapsa minor, involves centripetal synthesis and deposition of the mucopolymer cell wall layer (L 2). The ingrowth of the cytoplasmic membrane and L 1 cell wall layer, along with the synthesis of the L 2 cell wall layer, results in the formation of a prominent septum. Partitioning of the cell occurs by the constriction of the outer cell wall layers (L 3 and L 4) through the septum. InDermocarpa violaceae, constrictive binary fission occurs by the simultaneous ingrowth or constriction of the cytoplasmic membrane and all cell wall layers (L1, L2, L3, L4). Septate and constrictive binary fission may proceed symmetrically (medially) or asymmetrically (nonmedially). Multiple fission occurs regularly inDermocarpa violaceae and provides for a rapid means of reproduction when compared to binary fission. Successive radial and tangential divisions of the protoplast result in formation of many small daughter cells (baeocytes). The process of multiple fission is similar to septate binary fission with reduced septa being formed. However, constriction of the outer cell wall layers, through the septa, proceeds concurrently with septum formation.     

小麦条锈菌胞间菌丝的超微结构和细胞化学研究

本文应用电镜技术和细胞化学方法,对小麦条锈菌寄主胞间菌丝的超微结构进行了研究。观察发现:小麦条锈菌胞间菌丝有两种类型,即具隔膜菌丝和无隔膜菌丝。在胞间菌丝中,多核现象极为普遍。常规染色和细胞化学染色结果表明:胞间菌丝的细胞壁由四层组成,隔膜由三层构成,细胞壁的内层与隔膜的外层相连,细胞壁和隔膜中含有蛋白质和多糖物质。隔膜的发育可分三个阶段,即隔膜突的形成,隔膜壁的延伸和隔膜孔结构的形成。本研究中还观察到胞间菌丝间的融合现象。本文的研究结果表明:小麦条锈菌胞间菌丝的一些特征显然不同于其它锈菌。     

SEPTAL PORE STRUCTURE IN TRICHODERMA SATURNISPORUM

Electron microscopic observations of a previously undescribed ascomycetous septal pore structure are presented and discussed. Hyphal septa and septa at the base of phialides in the hyphomycete, Trichoderma saturnisporum Hammill, developed a membrane-bounded, electron-opaque septal pore body which was fine-structurally similar to Woronin bodies. Within a septal pore body, several electron-transparent layers appeared to develop centripetally from the septal pore rim. The number of layers observed varied from two to about five, with lower numbers being more frequently observed. The electron-transparent layers perhaps functioned as a vinculum, binding the septal pore body in place. Questions about the origin and function of septal pore bodies are discussed.     

柿树炭疽菌侵染寄主的细胞学研究*

超微结构研究表明,柿树炭疽菌(Colletotrichum gloeosporioides)侵染后在寄主细胞中形成初生菌丝和次生菌丝,寄主细胞膜外沉积了一层厚的电子不透明物质,初生菌丝与具有沉积物的寄主原生质膜之间有一层界面基质(interfacial matrix)。当初生菌丝扩张并侵染相邻细胞时, 围绕着初生菌丝层的界面基质消失,具有沉积物的原生质膜被逐步降解。初生菌丝在穿透寄主细胞壁过程中形成一个漏斗状的菌丝锥,然后穿透寄主细胞壁并迅速膨大, 然后形成厚壁的初生菌丝。初生菌丝在寄主细胞壁中收缩狭窄处产生一个隔膜,隔膜两边菌丝中细胞质的电子密度明显不同,菌丝锥中有浓密的电子密度。死体营养的次生菌丝在死的细胞中繁殖和扩展,并产生分枝。次生菌丝可直接穿透较薄的寄主细胞壁,无缢缩或任何变形现象,菌丝顶端部分未见隔膜产生;在穿透较厚的细胞壁时,靠近顶端处产生隔膜,顶端细胞膨大,使寄主细胞壁撕裂。接种90h后分生孢子盘在枝条表面形成。柿树炭疽菌其侵染过程有两个阶段,即初生菌丝的活体营养阶段和次生菌丝的死体营养阶段。     

Budding in the Dimorphic Fungus Phialophora dermatitidis

Ultrastructural comparisons of yeast and hyphal bud formation in Phialophora dermatitidis reveal that bud initiation is characterized by a blastic rupture of the outer portion of the yeast or hyphal wall and the emergence of a bud protuberance through the resulting opening. The wall of the emerging bud is continuous, with only an inner wall layer of the parental yeast or hypha. The outer, ruptured portion of the parental wall typically forms a collar around the constricted emergence region of the developing bud. The cytoplasm within the very young emerging bud invariably contains a small number of membrane-bound vesicles. The septum formed between the daughter bud and the parental yeast or hypha is a complete septum devoid of a septal pore, septal pore plug, or any associated Woronin bodies characteristic of simple septa of the moniliform or true hyphae. These observations suggest that yeast bud formation and lateral hyphal bud formation in the dimorphic fungus P. dermatitidis involve a growth process which occurs identically in both the yeast and mold phase of this human pathogenic organism.     

SEPTAL PORES IN THE HETEROBASIDIOMYCETIDAE,PUCCINIA GRAMINIS AND P. RECONDITA

The septal pores in uredial mycelium of Puccinia graminis and P. recondita lack the septal swelling and septal pore cap (dolipore-parenthosome configuration) typically associated with the pores of previously investigated Homobasidiomycetidae and the Tremellales among the Heterobasidiomycetidae. The pores in young hyphae of these two species of Puccinia are characterized by the presence of a cytoplasmic matrix which apparently occludes the pore and acts as a plug, thus preventing the migration of organelles from cell to cell. Large vesicles are typically present at the periphery of the pore matrix and the matrix may be very incompletely bounded by a membrane. Nuclei and other cytoplasmic structures migrate from cell to cell through an opening in the septum lateral to the pore. The available evidence indicates that this peripheral gap in the septum results from a breakdown of a portion of an initially complete septum rather than from incomplete septum formation. In addition to the centripetally formed septa, the hyphae of P. graminis and P. recondita are further compartmentalized by shallow infoldings of the lateral wall and limited unilateral septum formation. There is apparent free passage of cellular material between adjacent compartments.     

STRUCTURE AND DEVELOPMENT OF HYPHAL SEPTA IN ALLOMYCES

Development of hyphal septa (pseudosepta) in Allomyces macrogynus begins with the formation of five or more discontinuous pieces of wall material that project inward from the hyphal wall. Lateral fusion of these projections leaves a central pore in the septum that is later filled in by centripetal deposition of wall material. However, lateral fusion of the projections is not complete; peripheral pores remain in the rim of the mature septum. The position of cytoplasmic microtubules corresponds to the position of actively moving cellular particles and organelles. Allomyces reticulatus and A. arbuscula have similar septa.     

Formation of septum-like structures at locations remote from the budding sites in cytokinesis-defective mutants of Saccharomyces cerevisiae.

Cell wall structures that partition membrane-bound portions of cytoplasm were formed at sites along the peripheral wall when a cytokinesis-defective cell division cycle mutant (cdc3) of Saccharomyces cerevisiae was grown at a restrictive temperature. The appearance of these structures, as observed in electron micrographs, was similar to that of normal septa. Aberrant septa were also detected in cytokinesis mutants harboring mutations cdc10, cdc11, and cdc12, after growth at 37 degrees C. Formation of the abnormal septa was abolished by the introduction, in a cdc3-containing strain, of additional cell cycle mutations that precluded events leading to cytokinesis and cell division. These results showed that septum formation can occur in the absence of cytokinesis. Formation of the abnormal structures was controlled by the same sequences of cell cycle events as formation of normal septa but was not subject to the spatial controls that ensure association of the septum with the budding site.     

FORMATION AND STRUCTURE OF THE SPORE OF BACILLUS COAGULANS

Spore formation in Bacillus coagulans has been studied by electron microscopy using an epoxy resin (Araldite) embedding technique. The developmental stages from the origin of the initial spore septum to the mature spore were investigated. The two forespore membranes developed from the double layer of cytoplasmic membrane. The cortex was progressively deposited between these two membranes. The inner membrane finally became the spore protoplasmic membrane, and the outer membrane part of the inner spore coat or the outer spore coat itself. In the mature spore the completed integuments around the spore protoplasm consisted of the cortex, a laminated inner coat, and a dense outer coat. No exosporium was observed. The method of formation of the cortex and the spore coats is discussed.     

Pollen Wall Development in Gibasis (Commelinaceae)

The development of the one and-inline of the pollen wall aredescribed for Gibasis karwinsk yana and G. venustula. Duringthe tetrad stage the appearance of electron-opaque depositionsor tri-partite plates at discrete sites between the plasma membraneof the spore and the inward surface of the callose special wallare the first indications of exine development. The sulcus rapidlydifferentiates being composed of discrete exine granules ona thin foot layer. Probacula in non-apertural areas developin an electron-opaque granular layer situated between the plasmamembrane, which is highly convoluted, and the callose specialwall. A foot layer is formed from electron-opaque lamellae atthe plasma membrane. Exine pattern is clearly established withinthe tetrad. After release of the spores from the tetrad an intimate associationis rapidly developed between the plasma membrane of the periplasmodialtapetum and the newly-formed exine. Compacted electron-opaquematerial is found at the interface between membrane and theexine and vesicular material is added from the tapetum. Theincrease in volume that occurs in both spore and anther is accompaniedby considerable vacuolation. Intine development begins just prior to pollen grain mitosisand continues rapidly at the aperture. The thin foot layer becomesdiscontinuous. Further intine deposition takes place after mitosisand a bilayer is apparent in mature grains. The matrix of thislayer contains conspicuous electron-opaque platelets. The exineof the mature spore stains less intensely than in the youngspore and the interbacula spaces are filled with material fromthe degenerate tapetum. Gibasis karwinskyana, Gibasis venustula, Commelinaceae, exine, intine, tapetum, pollen wall, ultrastructure     

Fine Structural Studies on the Pycnidiospores of Botryodiplodia theobromae Pat.

The structure of the pycnidiospores of B. theobromae was investigatedby a combination of light microscope, a scanning electron microscopeand a transmission electron microscope observations. As observedwith the light microscope the hyaline non-septate pycnidiosporeis highly vesiculated while the pigmented septate one exhibitslongitudinal hyaline striations. Investigations with the scanningelectron microscope showed the hyaline non-septate pycnidiosporeto be smooth-walled without any form of ornamentation. The pigmentedseptate pycnidiospore on the other hand had a rough outer surfacewith a transverse groove, which probably indicated the positionof the septum. The longitudinal striations seen on such pycnidiosporeswith the light microscope were not observed under these conditions.Observations with the transmission electron microscope showedthat the outermost layer of the hyaline pycnidiospore was smootherthan that of the pigmented one. The pigment was on the outerlayer of the double-layered wall of the pigmented pycnidiospore.The inner non-pigmented wall layer was continuous with the septum.The septum tapered gradually to a thin layer in the centre whereit was perforated by a simple pore. The observations indicatedthat the pigmented spore was multinucleate while the hyalineone was uninucleate.     

The ultrastructure of cell wall formation and of gamma particles during encystment of Allomyces macrogynus zoospores

Structural changes during cell wall formation by populations of semisynchronously germinating zoospores were studied in the water mold Allomyces macrogynus. Fluorescence microscopy using Calcofluor white ST (which binds to -1,4-linked glycans) demonstrated that Calcofluor-specific material was deposited around most cells between 2–10 min after the induction of encystment (beginning when a wall-less zoospore retracts its flagellum and rounds up). During the first 15 min of encystment there was a progressive increase in fluorescence intensity. Ultrastructural analysis of encysting cells showed that within 2–10 min after the induction of encystment small vesicles 35–70 nm diameter were present near the spore surface, and some were in the process of fusing with the plasma membrane. The fusion of vesicles with the zoospore membrane was concomitant with the appearance of electron-opaque fibrillar material outside the plasma membrane. Vesicles similar to those near the spore surface were found within the gamma () particles of encysting cells. These particles had a crystalline inclusion within the electron-opaque matrix. During the period of initial cyst cell wall formation numerous vesicles appeared to arise at the crystal-matrix interface. Approximately 15–20 min was required for the cell wall to be formed. We suggest that the initial response of the zoospore to induction of encystment is the formation of a cell wall mediated by the fusion of cytoplasmic vesicles with the plasma membrane.Non-Standard Abbreviations GlcNac N-Acetylglucosamine - DS sterile dilute salts solution - PYG peptone-yeast extract-glucose broth     

Ultrastructure of the Cell Wall and Cell Division of Unicellular Blue-green Algae

The fine structure of the cell wall and the process of cell division were examined in thin sections of two unicellular blue-green algae grown under defined conditions. Unilateral invagination of the photosynthetic lamellae is the first sign of cell division in the rod-shaped organism, Anacystis nidulans. Symmetrical invagination of the cytoplasmic membrane and inner wall layers follows. One wall layer, which appears to be the mucopolymer layer, is then differentially synthesized to form the septum; the outer wall layers are not involved in septum formation. Centripetal splitting of the inner layer separates the two daughter cells. A second division, in a plane parallel to the first, usually occurs before the first daughter cells are separated. In the coccoid organism, Gleocapsa alpicola, the features of cell division are broadly similar; however, unilateral invagination of the lamellae is not observed and the second division takes place in a plane perpendicular to the plane of the previous division.     

The function of chitin synthases 2 and 3 in the Saccharomyces cerevisiae cell cycle

The morphology of three Saccharomyces cerevisiae strains, all lacking chitin synthase 1 (Chs1) and two of them deficient in either Chs3 (calR1 mutation) or Chs2 was observed by light and electron microscopy. Cells deficient in Chs2 showed clumpy growth and aberrant shape and size. Their septa were very thick; the primary septum was absent. Staining with WGA-gold complexes revealed a diffuse distribution of chitin in the septum, whereas chitin was normally located at the neck between mother cell and bud and in the wall of mother cells. Strains deficient in Chs3 exhibited minor abnormalities in budding pattern and shape. Their septa were thin and trilaminar. Staining for chitin revealed a thin line of the polysaccharide along the primary septum; no chitin was present elsewhere in the wall. Therefore, Chs2 is specific for primary septum formation, whereas Chs3 is responsible for chitin in the ring at bud emergence and in the cell wall. Chs3 is also required for chitin synthesized in the presence of alpha-pheromone or deposited in the cell wall of cdc mutants at nonpermissive temperature, and for chitosan in spore walls. Genetic evidence indicated that a mutant lacking all three chitin synthases was inviable; this was confirmed by constructing a triple mutant rescued by a plasmid carrying a CHS2 gene under control of a GAL1 promoter. Transfer of the mutant from galactose to glucose resulted in cell division arrest followed by cell death. We conclude that some chitin synthesis is essential for viability of yeast cells.     

ULTRASTRUCTURAL STUDY ON SPORE WALL MORPHOGENESIS IN LYCOPODIUM CLAVATUM (LYCOPODIACEAE)

Spore wall morphogenesis of Lycopodium clavatum was observed by transmission electron microscopy. The spore plasma membrane indicates the reticulate spore sculpture shortly after meiosis. The mature spore wall of this species consists of two layers, inner endospore and outer exospore. There is no perispore in the sporoderm of this species. The exospore formation begins during the tetrad stage; and this layer is divided into two distinct sublayers, an outer lamellar layer and an inner granular layer. The lamellar layer is formed on the sculptured spore plasma membrane. Additional lamellae attach to this layer in a centripetal direction. For that reason, this layer may be derived from spore cytoplasm. The granular layer is formed only in the proximal region following lamellar layer formation, and it also may be derived from spore cytoplasm. The endospore is formed lastly and seems to be derived from spore cytoplasm as well. Accordingly, the spore sculpture of this species may be under the genetic control of the spore nucleus.