Aureobasidium pullulans is a polymorphic microfungus that produces extracellular pullulan in culture media with a life-cycle involving blastospores, hyphae, chlamydospores and intermediate forms. The form(s) responsible for polysaccharide synthesis is still uncertain. In order to clarify the relation between pullulan synthesis and the different morphological forms, we carried out studies under different culture conditions, varying nutrients, incubation time, aeration and agitation. The extracellular polysaccharides were analysed by enzymic and acidic hydrolysis and thinlayer chromatography. The number of residues in repeating units was established. The results are reported in relation to the cell density and the percentage of the different morphological types. The best conditions for producing polysaccharides, particularly pullulan, were determined. The results suggest that swollen cells and chlamydospores are the forms that produce extracellular polysaccharides. When significant amounts of chlamydospores are present, authentic pullulan is always observed whatever the culture conditions. The swollen cells are responsible for the synthesis of another polysaccharide capable of being transformed into pullulan ; the hyphae do not play a role in extracellular polysaccharide biosynthesis.

To identify the cellular forms that are responsible for the synthesis of pullulan produced by Aureobasidium pullulans, we performed cytochemical and ultrastructural localizations of glucan in the cellular forms of this microorganism (blastospores and resting forms). Growth conditions, cell populations, and pullulan production were studied concurrently. Our results are consistent with a model in which the resting forms (swollen cells and chlamydospores) might be primarily involved in this extracellular polysaccharide elaboration. At the cellular level, pullulan production could be the result of three main stages: (i) cell wall thickening and extracellular polysaccharide synthesis by the swollen cell, (ii) fibrillar arrangement of this polysaccharide into pullulan along a capsular network around the chlamydospore, and (iii) subcellular hydrolysis separating the capsule from the periplasmic zone and consequently permitting the solubilization of pullulan in the culture medium. A melanization process in the outer layer of the cell wall and the capsule accompanies these patterns.Key words: Aureobasidium pullulans, capsule, cytochemistry, polysaccharide, pullulan, resting forms.

Pullulan is a well known extracellular polysaccharidic product of the hyphomycete Aureobasidium pullulans coming into more and more frequent use in commercial and industrial applications. Nevertheless, its cellular origin and biosynthesis pathways still remain uncertain. As pullulan synthesis is increasing while glycogen production is decreasing during growth, it should be possible that the kinetics of production of both polysaccharides, whose cellular locations differ, might be correlated. To check this hypothesis, we have performed biochemical analyses and microscopic studies of the biomass removed at regular intervals during growth. The ultrastructural data have shown that glycogen units were present in all the different cellular types (conidia, swollen cells, chlamydospores) and at all the stages of cellular development. Moreover, analytical studies have shown that glycogen level is time dependent, decreasing in the early exponential stage, whereas the extracellular pullulan content increases. The correlation coefficient (r) calculated between intracellular glycogen and extracellular pullulan levels by the chi2 method suggests that these products are inversely correlated. Ultrastructural and confocal fluorescence data indicate that glycolipids could be implicated in the pullulan biosynthetic pathway.Key words: Aureobasidium pullulans, pullulan, glycogen, metabolism, microscopy.

Substrate specificity tests were used to identify the presence of laccase in two strains of Leptosphaerulina briosiana (Poll.) Graham and Luttrell, an ascomycete which causes leaf spot in alfalfa. Cytochemical localization of monophenol monooxygenase (laccase) as well as the ultrastructures of the two strains were investigated. Laccase was observed in the outer layers of the cell walls of both strains. The ultrastructures of vegetative hyphae of both strains were typical of those found in most ascomycetes. mine the concentration necessary for complete inhibition of the enzyme. Various concentrations of sodium azide (e.g., 7.

The developmental pattern of Impatiens Balsamina L. gynoecium, established by morphological, anatomical, ontogenical, and teratological studies, leads us to an original conception of its organization. The syncarpous, pentamerous, pentalocular gynoecium is composed of a protecting wall consisting of five congenitally fused sterile carpels and an axial fertile part (placental column), whose vascularization is cauline. These different parts are initiated from distinct organogenic portions of the floral meristem. Carpellary primordia are initiated from lateral, meristematic portions (anneau initial floral) like gamophyllous foliar appendages; the placental column is initiated by dedifferentiation and reactivation of the apical central zone (which was previously inactive) and from cellular proliferation of the subjacent medullary meristem. This placental column represents the ultimate part of the floral axis. Ovarian walls are initiated on the periphery of the apical axial zone like common radiate intercarpellary protuberances.