Cross-pathogenicity of Rhizoctonia solani strains on pasture legumes in pasture-crop rotations

In Australia, in the past, pasture legumes were rotated mainly with cereals, but increasingly these rotations now involve pasture legumes with a wider range of crops, including legumes. This increasing frequency of the leguminous host in the rotation system may be associated with increased root rots in legumes in the current pasture-crop rotations. The primary aim of this study was to see whether the pathogenicity on pasture legumes of strains of Rhizoctonia solani sourced from lupins and cereals (common crops in rotation with pastures) is associated with increased incidence of root rots in pasture legumes in the disease conducive sandy soils of the Mediterranean regions of southern Australia. The second aim was to determine sources of resistance among newly introduced pasture legumes to R. solani strains originating from rotational crops as this would reduce the impact of disease in the pasture phase. Fifteen pasture legume genotypes were assessed for their resistance/susceptibility to five different zymogram groups (ZG) of the root rot pathogen R. solani under glasshouse conditions. Of the R. solani groups tested, ZG1–5 and ZG1–4 (both known to be pathogenic on cereals and legumes) overall, caused the most severe root disease across the genotypes tested, significantly more than ZG6 (known to be pathogenic on legumes), in turn significantly >ZG4 (known to be pathogenic on legumes) which in turn was >ZG11 (known to be pathogenic on legumes including tropical species). Overall, Ornithopus sativus Brot. cvs Cadiz and Margurita, Trifolium michelianum Savi. cvs Paradana and Frontier and T. purpureum Loisel. cv. Electro showed a significant level of resistance to root rot caused by R. solani ZG11 (root disease scores ≤1.2 on a 1–3 scale where 3 = maximum disease severity) while O. sativus cvs Cadiz and Erica showed a significant level of resistance to root rot caused by R. solani ZG4 (scores ≤1.2). O. compressus L. cvs Charano and Frontier, O. sativus cv. Erica, and T. purpureum cv. Electro showed some useful resistance to root rot caused by R. solani ZG6 (scores ≤1.8). This is the first time that cvs Cadiz, Electro, Frontier, Margurita and Paradana have been recognised for their levels of resistance to root rot caused by R. solani ZG11; and similarly for cvs Cadiz and Erica against ZG4; and for cvs Charano, Erica, and Electro against ZG6. These genotypes with resistance may also serve as useful sources of resistance in pasture legume breeding programs and also could potentially be exploited directly into areas where other rotation crops are affected by these R. solani strains. None of the tested genotypes showed useful resistance to R. solani ZG1–4 (scores ≥2.0) or ZG1–5 (scores ≥2.5). This study demonstrates the relative potential of the various R. solani ZG strains, and particularly ZG1–4, ZG1–5, ZG4 and ZG6 to attack legume pastures and pose a significant threat to non-pasture crop species susceptible to these strains grown in rotation with these pasture legumes. Significantly, the cross-pathogenicity of these strains could result in the continuous build-up of inoculum of these strains that may seriously affect the productivity eventually of legumes in all rotations. In particular, when choosing pasture legumes as rotation crops, caution needs to be exercised so that the cultivars deployed are those with the best resistance to the R. solani ZGs most likely to be prevalent at the location.     

Cancer chemotherapy and somatic cell mutation

The occurrence of a second neoplasm is one of the major obstacles in cancer chemotherapy. The elucidation of the genotoxic effects induced by anti-cancer drugs is considered to be helpful in identifying the degree of cancer risk. Numerous investigations on cancer patients after chemotherapy have demonstrated: (i) an increase in the in vivo somatic cell mutant frequency (Mf) at three genetic loci, including hypoxanthine–guanine phosphoribosyl-transferase (hprt), glycophorin A (GPA), and the T-cell receptor (TCR), and (ii) alterations in the mutational spectra of hprt mutants. However, the time required for and the degree of such changes are quite variable among patients even if they have received the same chemotherapy, suggesting the existence of underlying genetic factor(s). Accordingly, some cancer patients prior to chemotherapy as well as patients with cancer-prone syndrome have been found to show an elevated Mf. Based on the information obtained from somatic cell mutation assays, an individualized chemotherapy should be considered in order to minimize the risk of a second neoplasm.     

Promotion of indole alkaloid production in Catharanthus roseus cell cultures by rare earth elements

Production of the indole alkaloids, ajmalicine or catharanthine, in cell suspension cultures of Catharanthus roseus was enhanced by cerium (CeO₂ and CeCl₃), yttrium (Y₂O₃) and neodymium (NdCl₃). The yield of ajmalicine in these treated-cultures reached 51 mg l^–1 (CeO₂), 40 mg l^–1 (CeCl₃), 41 mg l^–1 (Y₂O₃) and 49 mg l^–1 (NdCl₃) while catharanthine production reached to 36 mg l^–1 (CeO₂) and 31 mg l^–1 (CeCl₃). A major portion of increased alkaloids was released into medium in these treatments. But Sm₂O₃, SmCl₃, La₂O₃, LaCl₃, complex of chromium (III)-titanium (IV) and NaSeO₄ treatments had little effect on alkaloid production of C. roseus cell cultures.     

Cell surface energy,contact angles and phase partition III. Adhesion of bacterial cells to hydrophobic surfaces

The densities of adhesion of Staphylococcus epidermis, Staphylococcus aureus and Serratia marcescens to five types of plastics were studied in relation to interfacial free energies at the aqueous interfaces of both the bacteria and the plastics. The free energy of adhesion of bacteria to plastic in an aqueous medium is a linear function of partition of the bacteria between the solid surface and the liquid phase. These results show that the thermodynamics of the partitioning of a suspended particle between two immiscible liquid phases also apply to partitioning between a liquid and a solid phase.     

Analysis of Trunk Neural Crest Cell Migration using a Modified Zigmond Chamber Assay

Neural crest cells (NCCs) are a transient population of cells present in vertebrate development that emigrate from the dorsal neural tube (NT) after undergoing an epithelial-mesenchymal transition ^1,2. Following EMT, NCCs migrate large distances along stereotypic pathways until they reach their targets. NCCs differentiate into a vast array of cell types including neurons, glia, melanocytes, and chromaffin cells ^1-3. The ability of NCCs to reach and recognize their proper target locations is foundational for the appropriate formation of all structures containing trunk NCC-derived components ³. Elucidating the mechanisms of guidance for trunk NCC migration has therefore been a matter of great significance. Numerous molecules have been demonstrated to guide NCC migration ⁴. For instance, trunk NCCs are known to be repelled by negative guidance cues such as Semaphorin, Ephrin, and Slit ligands ^5-8. However, not until recently have any chemoattractants of trunk NCCs been identified ⁹. Conventional in vitro approaches to studying the chemotactic behavior of adherent cells work best with immortalized, homogenously distributed cells, but are more challenging to apply to certain primary stem cell cultures that initially lack a homogenous distribution and rapidly differentiate (such as NCCs). One approach to homogenize the distribution of trunk NCCs for chemotaxis studies is to isolate trunk NCCs from primary NT explant cultures, then lift and replate them to be almost 100% confluent. However, this plating approach requires substantial amounts of time and effort to explant enough cells, is harsh, and distributes trunk NCCs in a dissimilar manner to that found in in vivo conditions. Here, we report an in vitro approach that is able to evaluate chemotaxis and other migratory responses of trunk NCCs without requiring a homogenous cell distribution. This technique utilizes time-lapse imaging of primary, unperturbed trunk NCCs inside a modified Zigmond chamber (a standard Zigmond chamber is described elsewhere¹⁰). By exposing trunk NCCs at the periphery of the culture to a chemotactant gradient that is perpendicular to their predicted natural directionality, alterations in migratory polarity induced by the applied chemotactant gradient can be detected. This technique is inexpensive, requires the culturing of only two NT explants per replicate treatment, avoids harsh cell lifting (such as trypsinization), leaves trunk NCCs in a more similar distribution to in vivo conditions, cuts down the amount of time between explantation and experimentation (which likely reduces the risk of differentiation), and allows time-lapse evaluation of numerous migratory characteristics.     

Molecular dynamics simulations of alpha2 --> 8-linked disialoside: conformational analysis and implications for binding to proteins.

Computational methods have played a key role in elucidating the various three-dimensional structures of oligosaccharides. Such structural information, together with other experimental data, leads to a better understanding of the role of oligosaccharide in various biological processes. The disialoside Neu5Ac-alpha2-->8-Neu5Ac appears as the terminal glycan in glycoproteins and glycolipids, and is known to play an important role in various events of cellular communication. Neurotoxins such as botulinum and tetanus require Neu5Ac-alpha2 --> 8-Neu5Ac for infecting the host. Glycoconjugates containing this disialoside and the enzymes catalyzing their biosynthesis are also regulated during cell growth, development, and differentiation. Unlike other biologically relevant disaccharides that have only two linkage bonds, the alpha2-->8-linked disialoside has four: C2-O, O-C8', C8'-C7', and C7'-C6'. The present report describes the results from nine 1 ns MD simulations of alpha2-->8-linked disialoside (Neu5Ac-alpha2-->8-Neu5Ac); simulations were run using GROMOS96 by explicitly considering the solvent molecules. Conformations around the O-C8' bond are restricted to the +sc/+ap regions due to stereochemical reasons. In contrast, conformations around the C2-O and C8'-C7' bonds were found to be largely unrestricted and all the three staggered regions are accessible. The conformations around the C7'-C6' bond were found to be in either the -sc or the anti region. These results are in excellent agreement with the available NMR and potential energy calculation studies. Overall, the disaccharide is flexible and adopts mainly two ensembles of conformations differing in the conformation around the C7'-C6' bond. The flexibility associated with this disaccharide allows for better optimization of intermolecular contacts while binding to proteins and this may partially compensate for the loss of conformational entropy that may be incurred due to disaccharide's flexibility.     

Lactosylceramide-induced stimulation of liposome uptake by Kupffer cells in vivo

Incorporation of 8 mol percent lactosylceramide into small unilamellar vesicles consisting of cholesterol and sphingomyelin in an equimolar ratio and containing [³H]inulin as a marker resulted in an increase in total liver uptake and a drastic change in intrahepatic distribution of the liposomes after intravenous injection into rats. The control vesicles without glycolipid accumulated predominantly in the hepatocytes, but incorporation of the glycolipid resulted in a larger stimulation of Kupffer-cell uptake (3.2-fold) than of hepatocyte uptake (1.2-fold). Liposome preparations both with and without lactosylceramide in which part of the sphingomyelin was replaced by phosphatidylserine, resulting in a net negative charge of the vesicles, were cleared much more rapidly from the blood and taken up by the liver to higher extents. The negative charge had, however, no influence on the intrahepatic distributions. The fast hepatic uptake of the negatively charged liposomes allowed competition experiments with substrates for the galactose receptors on liver cells. Inhibition of blood clearance and liver uptake of lactosylceramide-containing liposomes by $\text{N-}\text{acetyl-}\text{d}\text{-galactosamine}$ indicated the involvement of specific recognition sites for the liposomal galactose residues. This inhibitory effect of $\text{N-}\text{acetyl-}\text{d}\text{-galactosamine}$ was shown to be mainly the result of a decreased liposome uptake by the Kupffer cells, compatible with the reported presence of a galactose specific receptor on this cell type (Kolb-Bachofen et al. (1982) Cell 29, 859–866). The difference between the results on sphingomyelin-based liposomes as described in this paper and those on phosphatidylcholine-based liposomes as published previously (Spanjer and Scherphof (1983) Biochim. Biophys. Acta 734, 40–47) are discussed.     

Immunocytochemical localization of tubulin with colloidal gold in cilia of rabbit tracheal epithelial cultures

Ciliated tracheal epithelia cell cultures were investigated immunocytochemically with anti-tubulin and colloidal gold. When rabbit tracheal cultures were fixed in paraformaldehyde, treated with acetone, anti-tubulin and a second antibody coupled to FITC, fluorescence was associated with cytoskeletal and axonemal microtubules. Cilia covering the apical surface of the ciliated tracheal cells fluoresced very brightly thus facilitating identification of this cell type. Electron microscopy of tracheal cultures fixed as above, treated with Triton-X 100 and incubated in anti-tubulin and protein A coupled to colloidal gold resulted in the highly specific localization of tubulin in ciliary axonemes and basal bodies. Omission of primary or secondary antibody resulted in extremely low levels of fluorescence while no colloidal gold particles could be detected in cultures at the electron microscopy level when rabbit anti-tubulin was omitted.     

Design and operation of a completely automated Beckman microsequencer

A unique, efficient, and inexpensive system has been designed and built for the automatic conversion of anilinothiazolinone derivatives extracted from a Beckman spinning-cup sequencer with subsequent on-line high-pressure liquid chromatography separation of the phenylthiohydantoin derivatives. The Auto Converter-Auto Sampler system is controlled by a tape programmer or microprocessor and operates by transfer of the sample from the conversion vial into an HPLC injection loop by nitrogen pressure. Incorporation of a minor programming change on the sequencer allows the introduction of nitrogen vapor into the spinning cup during phenylisothiocyanate coupling. These modifications have resulted in a completely automated subnanomole protein sequencer.