Comparative ultrastructure of a mucoid strain of Pseudomonas aeruginosa isolated from cystic fibrosis patient and its spontaneous non-mucoid mutant

The ultrastructure of a mucoid strain of Pseudomonas aeruginosa of cystic fibrosis origin and its spontaneous non-mucoid variant was compared by transmission electron microscopy. Negatively-stained preparations of the mucoid strain obtained from plate cultures demonstrated dense, fibrous material projecting from the cell. No such material was observed in thin-sections or in negatively-strained preparations from liquid cultures. Thin-sections of ethanol-precipitated extracellular material from liquid cultures of the mucoid-strain revealed a cottony mesh of thin electron dense fibres. The non-mucoid strain did not produce such material. When prefixed with glutaraldehyde/malachite green mixture, cells of both strains demonstrated electron dense intracellular and extracellular malachite green-stainable structures. The internal complexes were frequently associated with the nucleoid or cell membrane and were replaced by electron transparent areas in cells prefixed with glutaraldehyde alone. Aeruginocins of the R-type were observed in mitomycin C induced cultures of both strains. Bacteriophages with 'claw-shaped' tail-tips were observed in the mucoid strain. Crystalline material was produced by the mucoid strain but only when plated on certain media.     

Cytochemical analysis of organelle degradation in phagosomes and apoptotic cells of the mucoid epithelium of mice.

The activity of mitochondrial cytochrome oxidase and peroxisomal catalase in the phagolysosomes and apoptotic bodies of mucoid epithelial cells was analysed. Tissue from 2-6 day old mice was used. The activity of acid phosphatase in lysosomes was also estimated. Cytochrome oxidase was demonstrated in well-preserved mitochondria inside phagosomes. Mitochondria in cells exhibiting apoptotic death also show activity of cytochrome oxidase. The enzyme activity in swollen mitochondria ceases before the membranes of the cristae disappear completely. Apoptotic bodies are phagocytosed by sister mucoid cells and, later on, they are digested inside the cell. Phagosomes which contain already degraded mitochondria show still active catalase in sequestered peroxisomes. The acid phosphatase involved in degradation of phagocytosed material originates from endocytosed lysosomes and primary and secondary lysosomes which fuse with the membranes of phagosomes.     

Cytochemical analysis of organelle degradation in phagosomes and apoptotic cells of the mucoid epithelium of mice

Summary The activity of mitochondrial cytochrome oxidase and peroxisomal catalase in the phagolysosomes and apoptotic bodies of mucoid epithelial cells was analysed. Tissue from 2–6 day old mice was used. The activity of acid phosphatase in lysosomes was also estimated. Cytochrome oxidase was demonstrated in well-preserved mitochondria inside phagosomes. Mitochondria in cells exhibiting apoptotic death also show activity of cytochrome oxidase. The enzyme activity in swollen mitochondria ceases before the membranes of the cristae disappear completely. Apoptotic bodies are phagocytosed by sister mucoid cells and, later on, they are digested inside the cell. Phagosomes which contain already degraded mitochondria show still active catalase in sequestered peroxisomes. The acid phosphatase involved in degradation of phagocytosed material originates from endocytosed lysosomes and primary and secondary lysosomes which fuse with the membranes of phagosomes.     

A Single Mutation in the Gene Responsible for the Mucoid Phenotype of Bifidobacterium animalis subsp. lactis Confers Surface and Functional Characteristics

Exopolysaccharides (EPS) are extracellular carbohydrate polymers synthesized by a large variety of bacteria. Their physiological functions have been extensively studied, but many of their roles have not yet been elucidated. We have sequenced the genomes of two isogenic strains of Bifidobacterium animalis subsp. lactis that differ in their EPS-producing phenotype. The original strain displays a nonmucoid appearance, and the mutant derived thereof has acquired a mucoid phenotype. The sequence analysis of their genomes revealed a nonsynonymous mutation in the gene Balat_1410, putatively involved in the elongation of the EPS chain. By comparing a strain from which this gene had been deleted with strains containing the wild-type and mutated genes, we were able to show that each strain displays different cell surface characteristics. The mucoid EPS synthesized by the strain harboring the mutation in Balat_1410 provided higher resistance to gastrointestinal conditions and increased the capability for adhesion to human enterocytes. In addition, the cytokine profiles of human peripheral blood mononuclear cells and ex vivo colon tissues suggest that the mucoid strain could have higher anti-inflammatory activity. Our findings provide relevant data on the function of Balat_1410 and reveal that the mucoid phenotype is able to alter some of the most relevant functional properties of the cells.     

Polyclonal B cell stimulation and interleukin 1 induction by the mucoid exopolysaccharide of Pseudomonas aeruginosa associated with cystic fibrosis

Mucoid exopolysaccharide isolated from Pseudomonas aeruginosa obtained from colonized cystic fibrosis patients was found to be a potent mitogen for mouse lymphocytes. The responding lymphocyte was a B cell, and we found no evidence that T cell could proliferate or synergize with B cells in response to the mucoid exopolysaccharide. Proliferation was not inhibitable by polymyxin B, which blocks lipopolysaccharide (LPS)-induced proliferation, indicating that a minor LPS contaminant in the purified exopolysaccharide was not the mitogenic component. Mucoid exopolysaccharide induced secretion of IgG, suggesting that it is polyclonal mitogen. It also induced splenic adherent cells (macrophages) to produce interleukin 1. We propose that mucoid exopolysaccharide produced by P. aeruginosa present in the lungs of cystic fibrosis patients may have potent in vivo consequences resulting in aberrant immunoregulation and inhibition of effective immune elimination of P. aeruginosa.     

Role of alginate lyase in cell detachment of Pseudomonas aeruginosa.

The exopolysaccharide alginate of Pseudomonas aeruginosa was shown to be important in determining the degree of cell detachment from an agar surface. Nonmucoid strain 8822 gave rise to 50-fold more sloughed cells than mucoid strains 8821 and 8830. Alginate anchors the bacteria to the agar surface, thereby influencing the extent of detachment. The role of the P. aeruginosa alginate lyase in the process of cell sloughing was investigated. Increased expression of the alginate lyase in mucoid strain 8830 led to alginate degradation and increased cell detachment. Similar effects were seen both when the alginate lyase was induced at the initial stage of cell inoculation and when it was induced at a later stage of growth. It appears that high-molecular-weight alginate polymers are required to efficiently retain the bacteria within the growth film. When expressed from a regulated promoter, the alginate lyase can induce enhanced sloughing of cells because of degradation of the alginate. This suggests a possible role for the lyase in the development of bacterial growth films.     

Histochemical studies on pseudocysts in adenoid cystic carcinoma of the human salivary gland

Summary Pseudocysts are unique structures found in adenoid cystic carcinomata of human salivary glands. They were studied in 13 such cases by histochemical and immunohistochemical means. The pseudocysts contained an abundance of mucoid materials which reacted strongly with both Alcian Blue and dialysed iron ferrocyanide. The mucoid material was digested with chondroitinase ABC and heparitinase, but was resistant toStreptomyces hyaluronidase. The inner surfaces of the pseudocysts were strongly reactive for laminin, whereas the interface between the tumour cell nests and the outer stromal area was intensely reactive for fibronectin. Numerous fibronectin-reactive fibrils and blood coagulation factor XIII (F-XIII)-positive cells were distributed extensively in the outer stromal area. The F-XIII-positive cells were also found within some pseudocysts. The results obtained in the present study have shown that the pseudocysts represent a peculiar structure consisting of basement membrane components; laminin, fibronectin, heparan sulphate and chondroitin sulphate.     

Morphological classification of pituitary adenomas and its value for clinical diagnosis]

Pituitary adenomas should be classified not only by their tinctorial affinities but also by their degree of differentiation. Then useful correlation to the clinical data can be obtained; On this principle our own collection of 299 tumors was classified in undifferentiated acidophilic, highly differentiated acidophilic GH cell-, highly differentiated acidophilic prolactin cell-adenomas, in undifferentiated mucoid cell-, highly differentiated mucoid ACTH cell-, highly differentiated mucoid TSH cell-adenomas, in chromophobic adenomas of small cell type and of large cell type, and in oncocytic adenomas. 95% of the cases with acromegaly based on undifferentiated acidophilic or highly differentiated GH cell adenomas. All patients with hypothalamic-hypophyseal Cushing's syndrome or Nelson's syndrome had undifferentiated mucoid cell adenomas or highly differentiated ACTH cell adenomas. In cases with hyperprolactinemia prolactin cell adenomas or chromophobic adenomas of large cell type with ultrastructurally demonstrated very highly developed rough endoplasmic reticulum or endocrinologically inactive chromophobic adenomas of small cell type were found. In the latter cases the prolactin is probably produced not by the tumor but by the surrounding tumor-free pituitary tissue.     

The carbohydrates of secretory granules and the glycocalyx in developing mucoid cells

Summary Complex carbohydrates in secretory granules and at the apical cell surface of mouse gastric mucoid cells were studied during embryogenesis and in the early postnatal period by various cytochemical methods; the periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) and tannic acid-uranyl acetate (TA-UA) procedures made neutral mucosubstances (NMS) visible, whereas the hexose residues of glycoconjugates were identified using WGA-, RCA II- and ConA-ferritin. The glycocalyx was stained with ruthenium red (RR). During differentiation of the embryonic mucoid cells the number of secretory granules increased in parallel to the increase in their carbohydrate component. NMS-stainable parts in secretory granules also had binding sites for the conjugates RCA II- and WGA-ferritin, but the binding of ConA could not be identified. The increasing quantity of NMS in secretory granules was correlated with the increased amount of PA-TCH-SP and TA-UA positive substances in the apical glycocalyx only in 14- and 18-day-old embryos. The observed uniform affinity for RR and lectin conjugates in all analysed developmental stages remains to be explained.     

金鱼、八目鳗和蟾蜍蝌蚪表皮Malpighian细胞超微结构的比较研究

本文对50只金鱼、5只八目鳗和10只蟾蜍蝌蚪的表皮做了电镜研究。在八目鳗和蝌蚪都观察到了典型的带微嵴的Malpighian细胞,而金鱼只在无鳞皮区观察到这种带微嵴的Malpighian细胞或称微嵴细胞。认为,微嵴细胞只见于无鳞皮,细胞表面的微嵴是一恒定性结构。并对其机能进行了讨论。     

The Functional Cytology of the Human Adenohypophysis

Using a combination of cytological, cytochemical and immunohistological techniques, an endeavour has been made to localize the sources of production of the seven protein and polypeptide hormones of the anterior pituitary gland. For growth hormone, TSH and ACTH, this has been done with reasonable certainty, and FSH and LH have been confirmed as coming from the mucoid cells. Although each of the hormones may be derived from a single cell type, present evidence has suggested that three or perhaps four cell types share the production of the seven known hormones and of any which remain to be discovered.     

Observations on the pituitary gland of some members of the suborder Suiformes (Mammalia: Artiodactyla)

The macroscopical and microscopical anatomy of the pituitary gland of the warthog (Phacochoerus aethiopicus) is described and compared with that of the Vietnam pig (a domestic race of Sus scrofa). The glands of the two animals differ considerably in shape but agree in anatomical details.
The morphological and histochemical characteristics of cells of the pars distalis are described in the warthog, bush pig (Potamochoerus porcus) , Vietnam pig (all members of the family Suidae) and hippopotamus (Hippopotamus amphibius: Hippopotamidae). Close agreement was found between five cell types of the warthog and bush pig and the corresponding cell types in the domestic pig. The mucoid cells of the less closely related hippopotamus differed from those of the domestic pig in some morphological details.     

Structure and histochemical characteristics of the mucosa of the Vater's ampulla in man (according to materials of aimed biopsies)]

Examination of 52 aimed duodenal biopsies has revealed that there are several variants of the structure of the Vater's papilla mucous membrane. In a number of cases it is identicalto the mucous membrane of the duodenum. In 20 cases the Vater's papilla was covered with mucoid epithelium which was histochemically similar to the epithelium lining the stomach, bile ducts and the gallbladder. The "intermedial" forms found in these cases appearedto be capable to change into mucoid or edging epithelium. The mucous cells occasionally found in the Vater's papilla epithelium might be a variation of goblet cells. It seems that they are secreting by holocrine type.     

Co-localization of TFF2 with gland mucous cell mucin in gastric mucous cells and in extracellular mucous gel adherent to normal and damaged gastric mucosa

Trefoil factor 2 (TFF2) is mucin associated peptide that has a mucosal barrier function in addition to participating in repair and healing. We examined the localization of TFF2 and gastric mucins in gastric mucous cells, the surface mucous gel layer (SMGL) adherent to normal gastric mucosa, and in the mucoid cap covering gastric erosions. Carnoy’s solution, or formalin/picric acid-fixed paraffin embedded materials from resected stomachs and formalin-fixed paraffin embedded gastric biopsy materials were used. Sections were immunostained for the TFF2 and histochemically stained for gastric mucins. In addition, thick sectioned gastric mucosa fixed in Carnoy’s solution were stained with FITC-labeled GSA-II lectin specific for gland mucous cell mucin and examined for three-dimensional images of the SMGL using a confocal laser scanning microscope. The TFF2 and gland mucous cell mucin were found intermixed together in the gastric gland mucous cells, in the SMGL in laminated layers, and in the mucoid cap. A laminated arrangement of continuous sheets of gland mucous cell mucin in the SMGL was demonstrated in the three-dimensional images. Co-localization of the TFF2 with gland mucous cell mucin suggests a physical interaction between the TFF2 and gland mucous cell mucin. The TFF2 trapped in the adherent mucins may be responsible for mucosal defense, healing, and repair.     

Alginate acetylation influences initial surface colonization by mucoid Pseudomonas aeruginosa

Mucoid strains of Pseudomonas aeruginosa overproduce the exopolysaccharide alginate, which is substituted with O-acetyl groups. Under non-growing conditions in phosphate buffer, a mucoid clinical strain formed microcolonies on steel surfaces, while an acetylation-defective mutant was unable to form cell clusters. Enzymatic degradation of alginate by alginate lyase prevented microcolony formation of the mucoid parent strain. In a continuous-culture flow-cell system, using gluconate minimal medium, the mucoid strain with acetylated alginate formed microcolonies and grew into heterogenous biofilms, whereas the acetylation-defective mutant produced a thinner and more homogeneous biofilm. A lowered viscosity of extracellular material from the acetylation-defective mutant indicated a weakening of exopolymer interactions by loss of acetyl groups. These results suggest that acetyl substituents are necessary for the function of high-molecular-mass alginate to mediate cell aggregation into microcolonies in the early stages of biofilm development by mucoid P. aeruginosa, thereby determining the architecture of the mature biofilm.     

The cytological study of the hypophysis pars anterior in the Indian grey mongoose, Herpestes edwardsi (Geoffroy)

The pituitary gland of the Herpestes edwardsi has been studies with special reference to the cytology of the pars anterior. In addition to the agranular chromophobic cells, six morphologically distinct chromophilic cell types have been observed in the pars anterior on the basis of selective histochemical and cytochemical staining techniques which can be classified into various groups such as STH, LTH, TSH, FSH, LH/ICSH and ACTH. The mucoid cells TSH, FSH and LH are much more concentrated in the centro-median region of the pars anterior. Highly concentrated colloid material appears in the acini or follicles which are scattered throughout the pars anterior. A relationship between the distribution of colloid material and any particular type of cell could not be established. The double intermedia layer surrounds almost completely the entire pars nervosa.     

Comparative morphology of the epidermis of seven species of polyclad flatworms (Platyhelminthes: Rhabditophora)

The ultrastructure of the epidermis of seven species of polyclad flatworms (Phaenocelis medvedica, Phaenocelis peleca, Pleioplana atomata, Boninia divae, Pericelis orbicularis, Enchiridium periommatum, and Cycloporus variegatus) representing six families is described. In all seven species, the epidermis consists of a single layer of columnar cells that rests on a bipartite basement membrane. Epithelial cell surfaces are covered by numerous microvilli and cilia. Cilia contain microtubules arranged in the 9 + 2 pattern, and from their basal bodies two striated rootlets arise, a rostrally directed one running parallel to the apical cell membrane, and a vertical one at a right angle to the first rootlet. Numerous epithelosomes and mitochondria occupy the apical parts of the cells. The basal part of the cells is highly folded, forming a cell web that connects to the basement membrane. The basement membrane consists of a thin basal lamina and a thick, multilayered reticular lamina. The number of layers in the reticular lamina varies among the different species and appears to be correlated with body size. Numerous canals containing either pigment granules or nervous processes perforate the basement membrane. We have identified four different types of glands: rhabdite glands, rhabdoid glands, mucoid glands containing vacuoles filled with flocculent material, and mucoid glands resembling thread cells of hagfish slime glands. The latter have been found only in P. orbicularis. Pigment cells were found in all species examined with the exception of C. variegatus, which takes its coloration from its ascidian prey. Our results further support the unique taxonomic status of Boniniidae.     

A histochemical study of the secretory gland cells of Cercaria shikokuensis and their role during development from cercaria to metacercaria.

The roles of secretory glands during the developmental process from an immature cercaria to a metacercaria in Cercaria shikokuensis were studied. Four types of secretory cells were identified in this species. On maturation of the cercaria in redia, the products of ventral gland cells and mucoid gland cells formed a thick surface coat on the mature cercaria, and the products of cephalic gland cells also formed a thin cover on the surface coat. In the process leading to the formation of a metacercaria, the surface coat constituted the outer layer of the cyst, mucoid gland cells secreted mucous substances inside the wall, and then cystogenous gland cells discharged their products to the inner wall. The cyst wall was composed of four layers, and it was thought that the outermost surface layer helped the cyst wall to adhere to the matrix and the intermediate layers helped to put together outer and inner walls.     

Identification, cloning, and characterization of rcsF, a new regulator gene for exopolysaccharide synthesis that suppresses the division mutation ftsZ84 in Escherichia coli K-12.

A new gene, designated rcsF, was located adjacent to drpA at the 5.2-min position of the genetic map of Escherichia coli. The deduced amino acid sequence encoded by the rcsF gene indicates a small protein of 133 amino acid residues with a calculated pI of 10.8 that is rich in proline, serine, alanine, and cysteine residues. When overexpressed as a result of its presence on a multicopy plasmid, rcsF confers a mucoid phenotype and restores colony formation to ftsZ84 mutant cells on L agar medium containing no added NaCl. These two phenotypes are not observed in rcsB mutant cells. Ion mutant cells harboring an rcsF mutation accumulate considerably lower levels of exopolysaccharides, whereas the presence of a multicopy rcsF plasmid not only increases capsule synthesis but also confers a mucoid phenotype at 37 degrees C, a temperature at which ion mutant cells are known not to form mucoid colonies. RcsF does not stimulate the expression of rcsB, indicating that it exerts its action through the RcsB protein, possibly by phosphorylation. It is also shown that RcsF stimulation of capsule synthesis is RcsA-dependent, whereas colony formation of ftsZ84 mutant cells can be restored by RcsF in the absence of RcsA.