The gene (spaP) coding for the Streptococcus mutans major surface protein antigen P1 (or IIH) has been cloned into Escherichia coli (

The aim of this study was to elucidate the effects of povidone-iodine (PVP-I) on cell ultrastructure by electron microscopy and to monitor changes in enzyme activity and nucleotide efflux. Staphylococcus aureus, Escherichia coli and Candida albicans, medically relevant gram-positive, gram-negative and yeast microorganisms, served as models. In the presence of PVP-I, rapid partitioning of the cytoplasm and pronounced coagulation of nuclear material was noted. E. coli and S. aureus showed no major structural wall damage. C. albicans exhibited a rapid, dose-dependent ‘loosening’ of the cell wall; cells remained intact without lysis, rupture or wall breakage. Changes in β-galactosidase and nucleotide concentrations were measured in E. coli. A rapid and dose-dependent loss of cellular β-galactosidase activity was found, with no increase in the supernatant; loss of cellular nucleotides corresponded with an increase in the supernatant. Electron-microscopic and biochemical observations support the conclusion that PVP-I interacts with cell walls of microorganisms causing pore formation or generating solid-liquid interfaces at the lipid membrane level which lead to loss of cytosol material, in addition to enzyme denaturation.

The spore coat forms as a rigid extracellular wall around each spore cell during culmination. Coats purified from germinated spores contain multiple protein species and an approximately equal mass of polysaccharide, consisting mostly of cellulose and a galactose/N-acetylgalactosamine polysaccharide (GPS). All but the cellulose are prepackaged during prespore cell differentiation in a regulated secretory compartment, the prespore vesicle. The morphology of this compartment resembles an anastomosing, tubular network rather than a spherical vesicle. The molecules of the prespore vesicles are not uniformly mixed but are segregated into partially overlapping domains. Although lysosomal enzymes have been found in the prespore vesicle, this compartment does not function as a lysosome because it is not acidic, and a common antigen associated with acid hydrolases is found in another, acidic vesicle population. All the prespore vesicle profiles disappear at the time of appearance of their contents outside of the cell; this constitutes an early stage in spore coat formation, which can be detected both by microscopy and flow cytometry. As an electron-dense layer, the future outer layer of the coat, condenses, cellulose can be found and is located immediately beneath this outer layer. Certain proteins and the GPS become associated with either the outer or inner layers surrounding this middle cellulose layer. Assembly of the inner and outer layers occurs in part from a pool of glycoproteins that is shared between spores, and unincorporated molecules loosely reside in the interspore matrix, a location from which they can be easily washed away. When the glycosylation of several major protein species is disrupted by mutation, the coat is assembled, but differences are found in its porosity and the extractibility of certain proteins. In addition, the retention or loss of proteolytic fragments in the mutants indicates regions of spore coat proteins that are required for association with the coat. Comparative examination of the macrocyst demonstrates that patterns of molecular distributions are not conserved between the macrocyst and spore coats. Thus spore coat assembly is characterized by highly specific intermolecular interactions, leading to saturable associations of individual glycoproteins with specific layers and the exclusion of excess copies to the interspore space.

The objectives of the present study were to develop an antibody probe to the porcine estrogen-dependent oviductal glycoproteins and to determine, by use of immunogold electron microscopy, whether these glycoproteins become associated with oviductal and uterine oocytes and early embryos. Polyclonal antibody, prepared using the M(r) 75,000-85,000 glycoprotein, separated from other proteins by two-dimensional SDS-PAGE, specifically recognized all three estrogen-dependent glycoproteins (acidic 75,000-85,000 M(r); acidic 100,000 M(r); basic 100,000 M(r)). In ampullary tissue collected from ovariectomized and estrogen-treated gilts and from gilts at Day 1 of estrus, gold particles were clustered over putative secretory granules restricted to the apical region of secretory epithelial cells. While follicular oocytes did not react with immunoreactive colloidal gold, oviductal and uterine unfertilized oocytes were found to be densely and uniformly labeled by colloidal gold throughout the zona pellucida, associated with flocculent material in the perivitelline space, and associated with microvilli and vitelline membrane. Similarly, in oviductal (1-4-cell) and unhatched uterine (4-cell/blastocyst) embryos, colloidal gold particles were distributed throughout the zona pellucida, heavily associated with flocculent material in the perivitelline space, and associated with the plasma membrane of the blastomeres. Immunoreactive colloidal gold remained detectable within Day 7 hatched uterine embryos, but not with embryos from later days. These results further support the proposal that porcine estrogen-dependent oviductal glycoproteins are released into the oviductal lumen, become associated with oviductal and uterine oocytes and early embryos, and are retained by oocytes and early embryos in the uterus.

Strains of Dictyostelium discoideum were paired in various combinations. Certain pairings resulted in the production of macrocysts, which are thought to be the sexual stage in the cellular slime molds. One selfcompatible strain was found. Other strains produced macrocysts only when paired with an alternate mating type. Two strains were found to form macrocysts when paired with either mating type but not alone or with each other. Several strains did not form macrocysts under any circumstances. Strains were graded according to their mating competency based on the number of macrocysts produced relative to the aggregated myxamoebae observed in the pairings.Macrocysts were first described by Blaskovics and Raper (1) as heavy-walled structures of multicellular origin that developed after cell aggregation in some isolates of Dictyostelium mucoroides and D. minutum and represented a morphogenetic phase alternative to sorocarp formation. Whether formed singly or in clusters, they noted that development began at the center of each incipient macrocyst and proceeded outward, the constituent myxamoebae seemingly being transformed into cells with semirigid walls, termed endocytes. They speculated that the myxamoebae became walled since they had strongly refractile boundaries. Later, these endocytes disappeared and the contents of the cyst became homogeneous.Subsequent ultrastructural studies have shown that a large cell, termed a cytophagic cell (2) or a giant cell (3), arises in the population and progressively engulfs the surrounding myxamoebae, which, once enclosed in vacuoles, represent the endocytes. Before this, the associated myxamoebae form a loose, fibrillar wall (primary wall) around the nascent cyst similar to the slime sheath that surrounds the pseudoplasmodium and the developing sorocarp. After engulfment of the myxamoebae is complete, the giant cell secretes a rigid, cellulosic wall (secondary wall) and later a thicker, more flexible, trilaminar wall. Coincident with the formation of this tertiary wall the endocytes are reduced to small fragments and ultimately disappear, giving the cyst a homogeneous appearance. After a variable period of dormancy, depending upon the species, the single large protoplast of the macrocyst undergoes progressive cleavage to give rise to trophic amoebae that emerge upon germination (4).Factors affecting the formation and germination of macrocysts have been studied (5, 6). In general, macrocyst formation is favored by darkness, very wet cultural conditions, temperature at the higher range of tolerance of the organism, and the absence of phosphate in the medium. The opposite conditions favor sorocarp production. Macrocysts that have undergone the appropriate period of dormancy can be induced to germinate by placing them in the light at low temperature (15-200).There is strong evidence to indicate that the macrocyst represents the true sexual stage in the cellular slime molds. In Polysphondylium violaceum (3), the giant cell has been shown to be at first binucleate but to be un...