Cytoplasmic vesicles in germinating spores ofGilbertella persicaria

Summary Germ tube apices ofGilbertella persicaria contain cytoplasmic vesicles, similar to the secretory vesicles found at the tips of vegetative hyphae. The vesicles are present at all stages of development, from the time of germ tube initiation to the establishment of branched hyphae. In contrast to the abundant vesicles at tips of established hyphae, the germ tubes have only a few apical vesicles in a layer next to the plasma membrane. When germinated spores are treated by washing and centrifuging prior to fixation, the cytoplasm is often disrupted near the apex, and the clusters of apical vesicles disappear. The findings indicate the delicate nature of hyphal tips and the necessity of avoiding prefixation stresses in order to preserve the apical apparatus of growing hyphae.     

Ultrastructure of germination and mucilage production inHalosphaeria appendiculata (Halosphaeriaceae)

The germination of ascospores of the marine fungusHalosphaeria appendiculata was investigated with transmission electron microscopy. Prior to germination, settled ascospores became surrounded by a fibro-granular layer. Small, membrane-bounded vesicles and larger electron-dense membrane-bounded vesicles aggregated at the site of germ tube formation where the plasmalemma adjacent to the aggregation was convoluted. The vesicles appeared to fuse with the plasmalemma, releasing their contents. Enzymatic digestion of the spore wall probably occurred at the time of germ tube emergence. After the nucleus had migrated into the newly formed germ tube, a septum was formed to delimit the germ tube from the ascospore. The growing germ tube can be divided into 3 morphological regions, namely the apical, sub-apical and vacuolated regions, and is typical of other fungi. A mucilaginous sheath was associated with the older mycelium. The germ tube displaced the polar appendage, and the ascospore, germ tube and appendage were enclosed in a mucilaginous sheath. In ascospores which subtended old germ tubes, the nucleus and lipid body became irregular in shape and the cytoplasm was more vacuolated. Microbody-like structures remained associated with the lipid throughout development, and were present in old ascospores.     

Mitochondrial behaviour during the yeast-hypha transition of Candida albicans

Yeast cells of Candida albicans were brought to germ tube formation and hyphal growth in liquid synthetic medium. The behaviour of mitochondria and mitochondrial nucleoids (mt-nucleoids) during morphological conversion was examined by fluorescence staining with 2-(4-dimethylaminostyryl)-1-methylpyridinium iodide (DASPMI) and 4',6-diamidino-2-phenylindole (DAPI). Parent yeast cells possessed one or very few branched giant mitochondria which were stained intensely with DASPMI. When a germ tube emerged from the parent cell, one end of a giant mitochondrion extended into the germ tube and developed into the elongated form. In mycelia, apical hyphal cells contained giant mitochondria, whereas older hyphal compartments near the parent cells were vacuolated and possessed small, peripherally located mitochondria. The vacuolated hyphal compartments resynthesized cytoplasm before producing branches and contained giant mitochondria. The cytological model for germ tube formation and hyphal growth proposed by Gow and Gooday (1984) is discussed.     

Early development in an algal gametophyte: role of the cytoskeleton in germination and nuclear translocation

Summary Biflagellate zoospores from the giant kelpMacrocystis pyrifera underwent germination after adhering to a substrate and produced germ tubes that were approximately 13–15 m in length. Coincident with the germ tube elongation was organelle (other than the nucleus) translocation along the tube. Shortly after formation of the germ tube, the zoospore nucleus divided and one daughter nucleus translocated along the germ tube. The nucleus did not appear to undergo chromosomal condensation prior to division. The nuclear division and/or translocation of the daughter nucleus did not begin until well after germ tube elongation was complete, demonstrating that these are temporally distinct developmental events. The translocation of one daughter nucleus coincided with differentiation of the distal end of the germ tube into a bulbous structure. Following this, the first gametophytic cross wall was formed and, subsequently, the daughter nucleus remaining in the original zoospore body underwent repositioning, assuming a position in the germ tube near the cross wall. Cytochalasin D inhibited germ tube elongation suggesting that actin microfilaments are probably involved in this developmental process. In addition, when cytochalasin D was added to the culture after the germ tube elongation was complete, it did not affect either nuclear division or translocation, indicating that microfilaments were not directly involved in these nuclear events. Colchicine and the plant specific microtubule disrupting agent, amiprophos methyl blocked nuclear division and translocation without affecting germination or germ tube elongation. These data suggest that actin microfilaments are primarily responsible for complete germination, specifically germ tube elongation, while microtubules are involved in nuclear division and translocation. The present study demonstrates that germination (and germ tube elongation) and nuclear translocation inM. pyrifera gametophytes are temporally and mechanistically distinct developmental events.     

Induction and formation ofCochliobolus sativus appressoria

Summary GerminatingCochliobolus sativus spores were induced to form appressoria on a variety of artificial surfaces, including replicas of the barley leaf surface. Evidence was obtained for the involvement of chemical and topographic signals during induction of appressorium formation inC. sativus. Germ tube thigmotropism was also observed in vitro. Ultrastructure relevant to appressorium formation was observed, including the germ tube apex, apical swelling of the germ tube apex prior to appressorium formation, the appressorium with associated septation and the penetration peg. Cytochemical probes applied to germlings at the electron microscope level failed to detect -D-mannan, -D-glucan, -D-galactan, D-glcNAc or D-galNAc polymers in the extracellular mucilage associated with the fungal germlings. The ultrastructure of hyphal apices from germlings grown under different nutritional conditions differed with respect to Spitzenkörper morphology, apex shape and in the quantity of associated extracellular mucilage. Experimental findings are discussed relative to current understanding of appressorium induction in more extensively studied systems.Abbreviations PDA potato dextrose agar - DS dilute salts - Con A concanavalin A - RcA₁₂₀ Ricinus communis agglutinin₁₂₀ - WGA wheat germ agglutinin - HpA Helix pomatia agglutinin - DIC differential interference contrast - UV ultraviolet - TEM transmission electron microscopy - NNF National Nanofabrication Facility     

Effects of brefeldin A on the endomembrane system and germ tube formation of the tetraspore of Gelidium floridanum (Rhodophyta,Florideophyceae)

Gelidium floridanum W.R. Taylor tetraspores are units of dispersal and are responsible for substrate attachment. This study aimed to examine evidence of direct interaction between germ tube formation and Golgi activity during tetraspore germination of G. floridanum. After release, the tetraspores were incubated with brefeldin A (BFA) in concentrations of 4 and 8 μM over a 6 h period. The controls and treatments were analyzed with light, fluorescence (FM4‐64 dye) and transmission electron microscopy. In the control samples, the Golgi bodies were responsible for germ tube formation. In contrast, BFA‐treated samples were observed to inhibit spore adhesion and germ tube formation. These tetraspores also showed an increase in volume (≥30 μm width). BFA treatment also resulted in the disassembly of Golgi cisternae and the formation of vesiculated areas of the cytoplasm, blocking the secretion of protein and amorphous matrix polysaccharides. When stained with FM4‐64, the control samples showed fluorescence in the apical region of the germ tube, but the treated samples showed an intense fluorescence throughout the cytoplasm. From these results, we can conclude that the germ tube is formed by the incorporation of vesicles derived from Golgi. Thus, vesicle secretion and Golgi organization are basic processes and essential in adhesion and tube formation. By blocking the secretion of protein and amorphous matrix polysaccharides, BFA treatment precluded tetraspore germination.     

Activity of Azole Fungicides and ABC Transporter Modulators on Mycosphaerella graminicola

The antimicrobial activity of the azole fungicides cyproconazole and propiconazole as single active ingredients and in mixtures with the ATP-Binding Cassette (ABC) transporter modulators rhodamine 6G, quercetin, quinidine, and verapamil and the strobilurin kresoxim-methyl was assessed against the wheat pathogen Mycosphaerella graminicola . Interactions amongst these compounds were evaluated on germination and germ tube growth of pycnidiospores using the Colby and Wadley method. Water agar proved to be the best test medium since all pycnidiospores germinated within 24 h of incubation and apical germ tube growth dominated over bud formation by intermediate cells. Analysis with the Colby method revealed that interactions between the compounds in all mixtures tested on germination of pycnidiospores were additive. With regard to germ tube growth, mixtures of cyproconazole and verapamil or kresoxim-methyl displayed a synergistic interaction. Analysis of mixtures of cyproconazole and kresoxim-methyl with the Wadley method revealed that the interaction between the two compounds was purely additive. These results indicate that the Colby method overestimated the interaction between these two compounds in a mixture.     

Untersuchungen über den chemischen Aufbau von Sporen- und Keimschlauchwänden der Uredosporen des Weizenrostes (Puccinia graminis var. tritici)

Zusammenfassung Zellwände und Keimschläuche von Uredosporen des Weizenrostes (Puccinia graminis var. tritici) wurden isoliert, und ihre chemische Zusammensetzung wurde quantitativ untersucht. Als gemeinsame Bausteine enthalten Sporenwände und Keimschlauchwände Proteine, Lipide und die Neutralzucker Galaktose, Glucose und Mannose. Die einzelnen Komponenten liegen in unterschiedlicher Menge vor. Auch qualitativ unterscheiden sich die Sporenwände und die Keimschlauchwände: Melanin ist nur in den Sporenwänden vorhanden, in den Keimschlauchwänden dagegen nicht. Der polymer gebundene Aminozucker der Keimschlauchwände ist N-Acetylglucosamin, das mit großer Wahrscheinlichkeit als Chitin vorliegt. Die Sporenwände enthalten dagegen polymeres Glucosamin (vermutlich Chitosan).Sporenwände sind in 3% iger NaOH löslich. Aus diesem Extrakt läßt sich mit Fehlingscher Lösung ein Galaktoglucomannan fällen, das überwiegend aus Mannose besteht. Aus der entsprechenden Fraktion der Keimschlauchwände, in der ebenfalls Mannose überwiegt, kann mit Fehlingscher Lösung kein Mannan gewonnen werden. Der in NaOH unlösliche Satz der Keimschlauchwände ist zum größten Teil aus Glucose und N-Acetylglucosamin aufgebaut. Es gibt keine identischen Polysaccharidfraktionen von Sporen- und Keimschlauchwänden. Sie sind heteropolymer und setzen sich jeweils aus Galaktose, Glucose und Mannose zusammen.

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Investigations on the chemical composition of spore walls and germ tube walls of wheat rust (Puccinia graminis var. tritici) uredospores

Summary Spore walls and germ tube walls from uredospores of wheat stem rust (Puccinia graminis var. tritici) were isolated and their chemical compositions determined quantitatively. The spore and germ tube walls are commonly composed of proteins, lipids, and the neutral sugars mannose, glucose and galactose. Carbon and nitrogen content, total lipids, composition of bound amino acids, total glucosamine and chitin content, and neutral sugars of spore and germ tube walls were compared. While the carbon content of the germ tube walls is only slightly higher than that of the spore walls, the germ tube walls contain twice as much nitrogen and lipids as the spore walls. The higher nitrogen content of the germ tube walls is due to higher amounts of bound amino acids and hexosamine. The polymeric germ tube wall hexosamine is insoluble in 3% NaOH, while the bulk of the polymeric spore wall hexosamine will go into solution when treated with 3% NaOH. The polymeric amino sugar of the germ tube wall is N-acetylglucosamine, which in all probability is present as chitin. In comparison, spore walls contain polymeric glucosamine (probably chitosan).The predominant neutral sugar of the spore walls is polymeric mannose (90%) while the germ tube walls contain polymeric glucose and mannose in nearly equal amounts. Galactose occurs in both wall types as a minor constituent.From spore walls completely dissolved in 3% NaOH we were able to precipitate a galactoglucomannan with fehling's solution. This galactoglucomannan was composed mainly of mannose. The corresponding fraction of the germ tube wall gave no precipitate with Fehling's solution. An alkali insoluble fraction of the germ tube wall consists mainly of glucose and N-acetylglucosamine. There are no identical polysaccharide fractions in spore walls and germ tube walls. They are always heteropolymers. Melanine is found in spore walls but not in germ tube walls.

     

The arrangement of F-actin and microtubules during germination of Mucor rouxii sporangiospores

The distribution of F-actin microfilaments and microtubules was analyzed in germinating sporangiospores of Mucor rouxii by labeling with rhodamine-tagged phalloidin and by immunofluorescence microscopy. The transition from isodiametrical to apical growth was accompanied by a switch from uniform distribution of F-actin patches to a polarized accumulation of F-actin material at the germ tube tips. Immunoblotting of cell-free extracts of M. rouxii with a monoclonal anti-porcine -tubulin antibody (TU-01) disclosed two discrete bands of -tubulin suggesting the existence of two -tubulin genes in this fungus. Immunofluorescence microscopy of germinating cells stained with the same antibody revealed an elaborate network of cytoplasmic microtubules that persisted during the entire germination process and extended into the apex of the germ tube. Although their precise roles remain undetermined, the observed arrangement of cytoskeletal elements during germination is consistent with their presumed involvement in cell wall morphogenesis: the long axial microtubules serving as long-distance conveyors of wall-building vesicles to the apical region while the concentrated F-actin patches mark the participation of microfilaments in the zone of intense vesicle exocytosis at the hyphal apex.Abbreviations DAPI 4,6-diamidino-2-phenylindole - DTT dithiothreitol - EGTA Ethylene glycol-bis (beta-aminoethyl ether) - N,N,N,N tetraacetic acid - F-actin Filamentous actin - MES 2-(N0morpholino)-ethanesulfonic acid - PIPES Piperazine-N,N-bis-2-ethanesulfonic acid - PMSF Phenyl-methylsulphonyl fluoride - TBS Tris-buffered saline     

Morphogenesis in germinating Fusarium graminearum macroconidia

Fusarium graminearum (teleomorph Gibberella zeae) is a significant pathogen of wheat and corn. F. graminearum forms multicellular macroconidia that play an important role in dissemination of the disease. The spatial pattern of morphogenesis in germinating macroconidia is described. Germ tubes preferentially emerge from the apical cells in a bipolar pattern that appears to be common to filamentous fungi. Chitin deposition occurs at two locations: the spore apices and cortical regions of macroconidial cells that subsequently produce a germ tube. The spatial pattern of morphogenesis requires the presence of functional microtubules, which may be responsible for the transport of key polarity factors to specific sites. These observations suggest that F. graminearum possesses a regulatory system that marks germ tube emergence sites. Perturbation of this system may represent an effective approach for inhibiting colonization of host plant surfaces.     

The effects of antimicrotubule agents on organelle positioning in the cowpea rust fungus,Uromyces phaseoli var.vignae

Six antimicrotubule agents were investigated for their effects on organelle positioning in living germ tubes of the cowpea rust fungus. Three (colchicine, colcemid, and isopropyl-n-phenylcarbamate) had no marked or consistent effect, even at the highest concentration in which the urediospores would germinate. In contrast, the remainder (nocodazole, vincristine sulfate, and griseofulvin) elicited striking, and remarkably similar, changes in the relative positions of mitochondria, nuclei, and vacuoles, as well as inhibiting the saltatory movements of lipid bodies. Nevertheless, germ tubes continued to grow in a reasonably normal fashion, in contrast to the “balloon-like” growth elicited by cytochalasin B treatment. In ultrathin, serial sections, a normal distribution of cytoplasmic microtubules was observed in colcemid-treated germ tubes, but no microtubules were seen after vincristine sulfate treatment, even though other components of the cytoplasm appeared undamaged. These results suggest that microtubules, either directly or through being part of a more complex cytoskeletal system, are involved in the positioning of cytoplasmic components in the cowpea rust germ tube. However, no evidence was found to support the hypothesis that microtubules are involved in apical vesicle transport, and a similar distribution of apical vesicles was observed in vincristine sulfate-treated and untreated germ tubes.     

Biomechanical basis of diazepam-induced neural tube defects in early chick embryos: a morphometric study

The biomechanical basis of diazepam (Valium/Roche)-induced neural tube defects in the chick was investigated using a combination of electron microscopy and morphometry. Embryos at stage 8 (four-somite stage) of development were explanted and grown for 6 hr in nutrient medium containing 400 micrograms/ml diazepam. Nearly 80% of these embryos exhibited neural tube defects that were most pronounced in the forming midbrain region and typified by a "relaxation" or "collapse" of neural folds. The hindbrain and spinal cord regions were less affected. Electron microscopy revealed that neuroepithelial cells in diazepam-treated embryos had smoother apical surfaces and broader apical widths than did controls. Morphometric measurements supported this observation and further showed that these effects were focused at sites within the wall of the forming neural tube that typically exhibit the greatest degree of bending and apical constriction (i.e., the floor and midlateral walls). Overall results indicate that neural tube defects associated with exposure to diazepam are due largely to a general inhibition of the contractile activity of apical microfilament bundles in neuroepithelial cells. These findings 1) emphasize the important contribution of microfilament-mediated apical constriction of neuroepithelial cells in providing the driving forces for bending of the neuroepithelium during neural tube formation and 2) suggest that agents or conditions that impair their contractile activity could play a role in the pathogenesis of certain types of neural tube defects.     

Antagonistic effect of Erwinia herbicola on in vitro spore germination and germ tube elongation of Botrytis cinerea and Penicillium expansum

Interaction between Erwinia herbicola and Botrytis cinerea and Penicillium expansum was studied in liquid culture. The results show that the bacteria directly inhibited spore germination of both fungi, especially during the first hours of the paired cultivation. The distinct taxis of bacteria to spores and germ tubes was frequently followed by their lysis. It is likely that bacteria act also by competition for nutrients. The rate of antagonistic activity of the bacteria against both fungi depended on their concentration in the mixture. Formation of chlamydospore-like structures at the apical end of B. cinerea germ tube suggests induction of a defence mechanism of this fungus while in unfavourable conditions.     

Confocal microscopy of Spitzenkörper dynamics during growth and differentiation of rust fungi

Summary. The membrane-selective fluorescent dye FM4-64, N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl)pyridium dibromide, was used to stain the apical vesicle cluster within the specialized Spitzenkörper of the germ tube of the rust fungi Uromyces vignae and Puccinia graminis f. sp. tritici grown on glass surfaces. The Spitzenkörper stained within 15 min following addition of the dye. Optical sectioning by confocal microscopy of stained hyphal tips showed that the Spitzenkörper was asymmetrically positioned close to the cell–substratum interface during germ tube growth. The Spitzenkörper showed variations in shape and positioning over short (5 s) time intervals. The movement to a new location in the hyphal dome was followed by new growth in that region, consistent with the view that the Spitzenkörper supplies secretory vesicles for germ tube growth. A pronounced Spitzenkörper disappeared at the onset of appressorium differentiation during swelling of the germ tube. However, a stained structure, similar in appearance to a Spitzenkörper, was again observed during the formation of the highly polarized penetration peg.Correspondence and reprints: Centraalbureau voor Schimmelcultures, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.Received October 25, 2002; accepted February 26, 2003; published online August 26, 2003     

Electron microscopy of spore germination and cell wall formation in Mucor rouxii

Summary The fine structure of ungerminated and aerobically germinated sporangiospores of Mucor rouxii was compared. The germination process may be divided into two stages: I, spherical growth; II, emergence of a germ tube. In both stages, germination is growth in its strictest sense with overall increases in cell organelles; e.g., the increase in mitochondria is commensurate with the overall increase in protoplasmic mass. Noticeable changes occurring during germination are the disappearance of electron-dense lipoid bodies, formation of a large central vacuole and, most strikingly, formation of a new cell wall. Unlike many other fungi, M. rouxii does not germinate by converting the spore wall into a vegetative wall. Instead, as in other Mucorales, a vegetative wall is formed de novo under the spore wall during germination stage I. This new wall grows out, rupturing the spore wall, to become the germ tube wall. Associated with the apical wall of the germ tube is an apical corpuscle previously described. The vegetative wall exhibits a nonlayered, uniformly microfibrillar appearance in marked distinction to the spore wall which is triple-layered, with two thin electron dense outer layers, and a thick transparent inner stratum. The lack of continuity between the spore and vegetative walls is correlated with marked differences in wall chemistry previously reported. A separate new wall is also formed under the spore wall during anaerobic germination leading to yeast cell formation. On the other hand, in the development of one vegetative cell from another, such as in the formation of hyphae from yeast cells, the cell wall is structurally continuous. This continuity is correlated with a similarity in chemical composition of the cell wall reported earlier.     

Arabidopsis AIP1-1 regulates the organization of apical actin filaments by promoting their turnover in pollen tubes

Apical actin filaments are highly dynamic structures that are crucial for rapid pollen tube growth, but the mechanisms regulating their dynamics and spatial organization remain incompletely understood. We here identify that AtAIP1-1 is important for regulating the turnover and organization of apical actin filaments in pollen tubes. AtAIP1-1 is distributed uniformly in the pollen tube and loss of function of AtAIP1-1 affects the organization of the actin cytoskeleton in the pollen tube. Specifically, actin filaments became disorganized within the apical region of aip1-1 pollen tubes. Consistent with the role of apical actin filaments in spatially restricting vesicles in pollen tubes, the apical region occupied by vesicles becomes enlarged in aip1-1 pollen tubes compared to WT. Using ADF1 as a representative actin-depolymerizing factor, we demonstrate that AtAIP1-1 enhances ADF1-mediated actin depolymerization and filament severing in vitro, although AtAIP1-1 alone does not have an obvious effect on actin assembly and disassembly. The dynamics of apical actin filaments are reduced in aip1-1 pollen tubes compared to WT. Our study suggests that AtAIP1-1 works together with ADF to act as a module in regulating the dynamics of apical actin filaments to facilitate the construction of the unique "apical actin structure" in the pollen tube.     

The FERM protein Epb4.1l5 is required for organization of the neural plate and for the epithelial-mesenchymal transition at the primitive streak of the mouse embryo

During early mouse development, a single-layered epithelium is transformed into the three germ layers that are the basis of the embryonic body plan. Here we describe an ENU-induced mutation, limulus (lulu), which disrupts gastrulation and the organization of all three embryonic germ layers. Positional cloning and analysis of additional alleles show that lulu is a null allele of the FERM-domain gene erythrocyte protein band 4.1-like 5 (Epb4.1l5). During gastrulation, some cells in lulu mutants are trapped in the primitive streak at an intermediate stage of the epithelial-mesenchymal transition; as a result, the embryos have very little paraxial mesoderm. Epithelial layers of the later lulu embryo are also disrupted: definitive endoderm is specified but does not form a gut tube, and the neural plate is broad and forms ectopic folds rather than closing to make the neural tube. In contrast to zebrafish and Drosophila, in which orthologs of Epb4.1l5 control the apical localization and activity of Crumbs proteins, mouse Crumbs proteins are localized normally to the apical surface of the lulu mutant epiblast and neural plate. However, the defects in both the lulu primitive streak and neural plate are associated with disruption of the normal organization of the actin cytoskeleton. We propose that mouse Lulu (Epb4.1l5) helps anchor the actin-myosin contractile machinery to the membrane to allow the dynamic rearrangements of epithelia that mediate embryonic morphogenesis.     

Candida albicans VPS11 is required for vacuole biogenesis and germ tube formation

The Candida albicans vacuole has previously been observed to undergo rapid expansion during the emergence of a germ tube from a yeast cell, to occupy the majority of the parent yeast cell. Furthermore, the yeast-to-hypha switch has been implicated in the virulence of this organism. The class C vps (vacuolar protein sorting) mutants of Saccharomyces cerevisiae are defective in multiple protein delivery pathways to the vacuole and prevacuole compartment. In this study C. albicans homologues of the S. cerevisiae class C VPS genes have been identified. Deletion of a C. albicans VPS11 homologue resulted in a number of phenotypes that closely resemble those of the class C vps mutants of S. cerevisiae, including the absence of a vacuolar compartment. The C. albicans vps11Delta mutant also had much-reduced secreted lipase and aspartyl protease activities. Furthermore, vps11Delta strains were defective in yeast-hypha morphogenesis. Upon serum induction of filamentous growth, mutants showed delayed emergence of germ tubes, had a reduced apical extension rate compared to those of control strains, and were unable to form mature hyphae. These results suggest that Vps11p-mediated trafficking steps are necessary to support the rapid emergence and extension of the germ tube from the parent yeast cell.     

Induction of germ tube formation by N-acetyl-D-glucosamine in Candida albicans: uptake of inducer and germinative response

A number of strains of Candida albicans were tested for germ tube formation after induction by N-acetyl-D-glucosamine (GlcNAc) and other simple (proline, glucose plus glutamine) or complex (serum) compounds. A proportion of strains (high responders) were induced to form germ tubes evolving to true hyphae by GlcNAc alone or by proline or glucose plus glutamine mixture. The majority of strains were low responders because they could be induced only by serum or GlcNAc-serum medium. Two strains were found to be nonresponders: they grew as pseudohyphae in serum. Despite minor quantitative differences, all strains efficiently utilized GlcNAc for growth under the yeast form at 28 degrees C. They also had comparable active, inducible, and constitutive uptake systems for GlcNAc. During germ tube formation in GlcNAc, the inducible uptake system was modulated, as expected from induction and decay of GlcNAc kinase. Uranyl acetate, at a concentration of 0.01 mM, inhibited both GlcNAc uptake and germ tube formation and was reversed by phosphates. Germinating and nongerminating cells differed in the rapidity and extent of GlcNAc incorporation into acid-insoluble and alkali-acid-insoluble cell fractions. During germ tube formation induced by proline, GlcNAc was almost totally incorporated into the acid-insoluble fraction after 60 min. Moreover, hyphal development on induction by either GlcNAc or proline was characterized by an apparent "uncoupling" between protein and polysaccharide metabolism, the ratio between the two main cellular constituents falling from more than 1 to less than 0.5 after 270 min of development. The data suggest that utilization of the inducer for wall synthesis is a determinant of germ tube formation C. albicans but that the nature and extent of inducer uptake is not a key event for this phenomenon to occur.     

Fine Structure of Germinating Botrytis fabae Sardina Conidia

The fine structure of germinating Botrytis fabae conidia wasstudied using both chemically stained sections and freeze-etchedreplicas. Germinating conidia have fewer organelles than restingconidia, glycogen is absent, and prevacuoles have disappeared.Endoplasmic reticulum which occurs as small strands close tothe cell wall of resting conidia becomes, on germination, multiplesheets surrounding the nuclei. A cross wall is formed at thebase of the germ tube soon after germination commences. Thenew wall material which appears to be continuous with this septalwall is produced, at least partly, from a new wall layer laiddown in the centre of the old conidial wall. An apical corpuscleis present at the apex of young germ tubes. Freeze-etched preparationsshow the formation of lomasomes by the passage of vesicles throughthe plasmalemma of conidia and germ tubes. In young hyphae lomasomescontain a complex arrangement of branching tubules. Some ofthe particles on the outer plasmalemma of young hyphae are arrangedin a geometrical pattern.