Changes in leaf,stem, and root anatomy of Chrysanthemum cv. Lillian Hoek following paclobutrazol application

Plants ofChrysanthemum cv. Lillian Hoek were treated with a paclobutrazol (PBZ) soil drench and histologically examined after 3 months. PBZ application resulted in thicker leaves, reduced stem diameter, and roots with an increased diameter and an unusual segmented appearance. Increased leaf thickness was partly due to an additional layer of palisade mesophyll, although individual palisade cells were shorter, of smaller diameter, and more tightly packed. Spongy mesophyll depth was also greater and the individual cells were more rounded and the volume of intercellular space was reduced. The narrower stems had an increased development of secondary xylem, but had a marked reduction in the number of sclerenchyma bundle caps. Increased root diameter was due to an increase in the number of rows and diameter of cortical cells. In PBZ-treated plants, root cortical cell length was 50–70% less than in untreated plants, and this reduction appeared to be associated with the segmentation of the roots. PBZ inhibited secondary vascular development in the roots. This study is similar to other relevant studies in recording thicker leaves and roots with PBZ application; however, many of the underlying anatomical changes described above have not been previously reported.     

Fish size and habitat depth relationships in headwater streams

Summary Surveys of 262 pools in 3 small streams in eastern Tennessee demonstrated a strong positive relationship between pool depth and the size of the largest fish within a pool (P<0.001). Similarly, the largest colonizers of newly-created deep pools were larger than the colonizers of shallow pools. We explored the role of predation risk in contributing to the bigger fish — deeper habitat pattern, which has been noted by others, by conducting five manipulative field experiments in two streams. Three experiments used stoneroller minnows (Campostoma anomalum); one used creek chubs (Semotilus atromaculatus); and one used striped shiners (Notropis chrysocephalus). The stoneroller experiments showed that survival of fish approximately 100 mm in total length (TL) was much lower in shallow pools (10 cm deep) than in deep (40 cm maximum) pools (19% versus 80% survival over 12 d in one experiment) and added cover markedly increased stoneroller survival in shallow pools (from 49% to 96% in an 11-d experiment). The creek chub experiment showed that, as for stonerollers, pool depth markedly influenced survival: the chubs survived an average of 4.9 d in shallow pools and >10.8 d in deep pools. In the striped shiner experiment in shallow artificial streamside troughs, no individuals 75–100 mm TL survived as long as 13 d, where-as smaller (20–25 mm) fish had 100% survival over 13 d. The results of the experiments show that predation risk from wading/diving animals (e.g., herons and raccoons) is much higher for larger fishes in shallow water than for these fishes in deeper water or for smaller fish in shallow water. We discuss the role of predation risk from two sources (piscivorous fish, which are more effective in deeper habitats, and diving/wading predators, which are more effective in shallow habitats) in contributing to the bigger fish — deeper habitat pattern in streams.     

Use of a photolabeling technique to identify nonviable cells in fixed homologous or heterologous cell populations

Flow cytometric determination of viable versus nonviable cells in fixed samples can be accomplished by utilizing the irreversible binding of photoactivated ethidium monoazide (EMA). EMA is a positively charged molecule which is excluded by cells with intact membranes (viable cells), included by cells with damaged membranes, and can be photochemically crosslinked to nucleic acids using visible light. EMA fluorescence can be excited using a standard argon laser operating at 488 nm and is able to be distinguished from fluorescein and phycoerythrin. Fixation is important when analyzing cells from a potentially infectious origin. EMA is photochemically crosslinked and therefore unable to leak out of cells when removed from the extracellular media, unlike propidium iodide (PI) or other viability stains, which were heretofore commonly used. We demonstrate the usefulness of EMA in combination with fluoresceinated and phycoerythrin labeled monoclonal antibodies in immunophenotyping. The photoaffinity labeling technique allows for a quick and efficient means of identifying nonviable cells which cannot be distinguished on the basis of light-scattering properties.     

Prohibitin, an evolutionarily conserved intracellular protein that blocks DNA synthesis in normal fibroblasts and HeLa cells.

Genes that act inside the cell to negatively regulate proliferation are of great interest because of their implications for such processes as development and cancer, but these genes have been difficult to clone. This report details the cloning and analysis of cDNA for prohibitin, a novel mammalian antiproliferative protein. Microinjection of synthetic prohibitin mRNA blocks entry into S phase in both normal fibroblasts and HeLa cells. Microinjection of an antisense oligonucleotide stimulates entry into S phase. By sequence comparison, the prohibitin gene appears to be the mammalian analog of Cc, a Drosophila gene that is vital for normal development.     

Product inhibition of immobilized Escherichia coli arising from mass transfer limitation.

Mass transfer-limited removal of metabolic products led to product-inhibited growth of Escherichia coli that was immobilized in a model system. Comparison of the growth kinetics of immobilized and free-living cells revealed no further physiological differences between cells in these two modes of existence beyond those manifested in the local concentrations of substrate and product. Bacteria were retained on a microporous membrane in a dense, planar aggregate and were grown anaerobically on a glucose-based minimal medium. Radioisotope labeling of the immobilized cell mass with 35S was used to determine growth kinetic parameters. Growth rates in the immobilized cell layer were measured by an autoradiographic technique which allowed comparison of the size of the growing region with the rate of cell convection caused by growth. Immobilized cell growth rates and growth yields ranged from near maximal (0.56 h-1 and 39 g of dry cell weight/mol of glucose, respectively) to substantially reduced (0.15 h-1 and 15 g/mol). The depression of these kinetic parameters was attributed to product inhibition arising from mass transfer-limited removal of acidic waste products from the cell mass. A simple one-dimensional reaction-diffusion model, which incorporated data on the product-inhibited growth kinetics of free-living cells collected in a product-limited chemostat, satisfactorily predicted product inhibition of immobilized cell growth.     

Kinetic measurements of fusion of phosphatidylserine-containing vesicles by electron microscopy and fluorometry

Large unilamellar vesicles (REV) containing phosphatidylserine and phosphatidylethanolamine at a ratio of 1:3 were induced to fuse by adding calcium (4 mM). The kinetics of fusion was monitored by fluorometry using terbium or dipicolinic acid-containing vesicles. The morphology and the states of vesicle aggregation and fusion were examined at approx. 2, 30, 60, 150 and 900 s after calcium addition, by rapid quenching and freeze-fracture electron microscopy. The size and the state of aggregation of vesicles are quantitated from 4000 randomly selected vesicles. The aggregation and fusion kinetics as assayed by fluorescence volume mixing is very well simulated and predicted by the mass action model. The model essentially predicts the time course of the distribution of the aggregates and the increase in size of fused particles as measured by electron microscopy, although in some cases the predicted fusion rate exceeds that by morphometric measurement. No morphological features can be defined as fusion intermediates, although bead-like and rim-like materials may be attributed to the remnants of broken diaphragms between fusion partners.     

Synthesis and characterization of deoxy analogues of diphytanylglycerol phospholipids

Novel analogues of diphytanyl phospholipids, 2,3-diphytanyl sn-1-glycerol-1-phosphoryl-1'-(1',3'-propanediol) (dPG), 2,3-diphytanyl-sn-glycerol-1-phosphoryl-1'-propanol (ddPG) and 2,3-diphytanyl-sn-glycerol-1-phosphoryl-1'-(1',3'-propanediol-3'-p hosphate) (dPGP), were synthesized according to modifications of previously published procedures. The samples were TLC and analytically pure and were characterized by 13C- and 1H-NMR and negative FAB/MS. The pK values of dPGP in aqueous dispersions or in methanol/water (1:1, v/v) were determined by potentiometric titration and compared with those of 2,3-diphytanyl-sn-glycerol-1-phosphoryl-3'-sn-glycerol-1'-phosphat e (PGP). The dissociation constant of the third ionizable POH group of dPGP was more than 2 pK units higher than that of PGP, indicating that the free glycerol hydroxyl group plays an important role in headgroup conformation and stabilization, perhaps through hydrogen bonding with the phosphate group(s).     

Excimer fluorescence of equine platelet tropomyosin labeled with N-(1-pyrenyl)iodoacetamide

Tropomyosin from equine platelets was reacted with N-(1-pyrenyl)iodoacetamide, a sulfhydryl-specific fluorescent reagent, to give an average extent of incorporation of 1.12 pyrene (Py) groups per platelet tropomyosin (P-TM) chain. The predominant site of reaction on P-TM was the penultimate COOH-terminal residue, Cys-246. The high proportion of the total emission that is due to pyrene ecximers and the pretransition observed in thermal denaturation of Py-P-TM point to a rather loose structure for the COOH-terminal amino acid residues of P-TM. The label on Cys-246 also reports on end-to-end overlap interactions that occur between two different tropomyosin molecules. Additions to a Py-P-TM solution at low ionic strength of unlabeled P-TM, rabbit cardiac tropomyosin (C-TM), or a carboxypeptidase A treated, nonpolymerizable derivative of C-TM all reduce the extent of excimer fluorescence from the sample. Addition of salt greatly reduces the effects of the unlabeled TM species on the Py-P-TM emission spectrum. Circular dichroism measurements indicate Py-P-TM still to be greater than 95% helical. However, analysis of excimer fluorescence levels in samples that contained a constant protein concentration but different mole ratios of labeled to unlabeled P-TM suggests that the bulky pyrene group may diminish the tendency of Py-P-TM to polymerize in an end-to-end manner.     

A note on the control of sulphate-reducing bacteria in seawater by u.v. irradiation

Continuous and batch cultures of marine sulphate-reducing bacteria (SRB) in North Sea water were irradiated with 110000 to 329500 μWs/cm² of ultraviolet radiation (wavelength 253.7 nm) with a commercial u.v. sterilizing unit. A 100% kill was obtained with logarithmic cultures of Desulfovibrio desulfuricans NCIMB 8400 at population densities of 10–10⁴/ml. A >99.99% kill was obtained with a mixture (ca 10⁵/ml) of batch grown Desulfovibrio spp. and oilfield SRB enrichments. Ultraviolet irradiation was less effective against the indigenous heterotrophic bacteria in the seawater ( ca 90% kill).