Phytophthora and Pythium Associated with Feeder Root Rot of Citrus in the Transvaal Province of South Africa

Mild to extensive feeder root rot was present in all 23 orchards, with trees showing symptoms of citrus decline from nine areas in the Transvaal Province of South Africa. Phytophthora nicotianae and Pythium spp. were isolated from diseased roots and rhizosphere soils in all areas sampled. Isolations from diseased feeder roots showed P. nicotianae present in 26% of orchards during Spring and 61% of orchards during Autumn, while Pythium spp. were present in 56% of orchards during Spring and 65% of orchards during Autumn. In isolations from baited rhizosphere soils, P. nicotianae was present in 56% of orchards during Spring and 52% of orchards during Autumn, while Pythium spp. were present in 69% of orchards during Spring and 82% of orchards during Autumn. In rhizosphere soils, the mean population density of Pythium spp. was higher than that of P. nicotianae throughout the season. Only P. nicotianae was consistently isolated during thesurvey. Different Pythium spp. were isolated of which two were tentatively identified as P. paroecandrum and Pythium‘Gp.G’.     

A systematic study of labelling an α-helix in a protein with a lanthanide using IDA-SH or NTA-SH tags

The previously published IDA-SH and NTA-SH tags are small synthetic lanthanide-binding tags derived from cysteine, which afford site-specific lanthanide labelling by disulfide-bond formation with a cysteine residue of the target protein. Following attachment to a single cysteine in an α-helix, sizeable pseudocontact shifts (PCS) can be observed, if the lanthanide is immobilized by additional coordination to a negatively charged amino-acid side chain that is located in a neighboring turn of the helix. To identify the best labelling strategy for PCS measurements, we performed a systematic study, where IDA-SH or NTA-SH tags were ligated to a cysteine residue in position i of an α-helix, and aspartate or glutamate residues were placed in the positions i ? 4 or i + 4. The largest anisotropy components of the magnetic susceptibility tensor were observed for an NTA-SH tag in position i with a glutamate residue in position i ? 4. While the NTA-SH tag produced sizeable PCSs regardless of the presence of nearby carboxyl groups of the protein, the IDA-SH tag generated a good lanthanide binding site only if an aspartate was placed in position i + 4. The findings provide a firm basis for the design of site-directed mutants that are suitable for the reliable generation of PCSs in proteins with paramagnetic lanthanides.     

Vulnerability of Pathogenic Biofilms to Micavibrio aeruginosavorus

The host specificity of the gram-negative exoparasitic predatory bacterium Micavibrio aeruginosavorus was examined. M. aeruginosavorus preyed on Pseudomonas aeruginosa, as previously reported, as well as Burkholderia cepacia, Klebsiella pneumoniae, and numerous clinical isolates of these species. In a static assay, a reduction in biofilm biomass was observed as early as 3 hours after exposure to M. aeruginosavorus, and an ~100-fold reduction in biofilm cell viability was detected following a 24-h exposure to the predator. We observed that an initial titer of Micavibrio as low as 10 PFU/well or a time of exposure to the predator as short as 30 min was sufficient to reduce a P. aeruginosa biofilm. The ability of Micavibrio to reduce an existing biofilm was confirmed by scanning electron microscopy. In static and flow cell experiments, M. aeruginosavorus was able to modify the overall P. aeruginosa biofilm structure and markedly decreased the viability of P. aeruginosa. The altered biofilm structure was likely caused by an increase in cell-cell interactions brought about by the presence of the predator or active predation. We also conducted a screen to identify genes important for P. aeruginosa-Micavibrio interaction, but no candidates were isolated among the ~10,000 mutants tested.     

Precise vicinal coupling constants3JHNα in proteins from nonlinear fits of J-modulated [15N,1H]-COSY experiments

Summary Improved experimental schemes for the recently introduced J-modulated [¹⁵N,¹H]-correlation experiment for measurements of the homonuclear amide proton-C proton vicinal coupling constants.³J_HN, in uniformly¹⁵N-labeled proteins are described, and a nonlinear fit procedure is presented for quantitative evaluation of³J_HN. The method was first tested with the N-terminal DNA-binding domain of the 434 repressor (M=7.3 kDa), where at 13 C precise values of³J_HN in the range 2.0–9.5 Hz were obtained for all residues with resolved¹⁵N-¹H cross peaks. It was then applied to theAntennapedia homeodomain complexed to a synthetic 14-base pair DNA fragment (molecular weight of the complex 18 kDa). The³J_HN values measured were found to be in excellent agreement with those predicted from the secondary structure of this protein in the complex.Abbreviations and symbols NOE nuclear Overhauser effect - COSY two-dimensional correlated spectroscopy - ³J_HN or J homonuclear vicinal amide proton-C proton coupling constant - 434 repressor(1–69) N-terminal DNA-binding domain of the 434 repressor comprising 69 residues     

Protein kinase C plays a role in the induction of tyrosine phosphorylation of lymphoid microtubule-associated protein-2 kinase. Evidence for a CD3-associated cascade that includes pp56lck and that is defective in HPB-ALL.

Ligation of the CD3 receptor induces multiple signal transduction events that modify the activation state of the T cell. We have compared two lines that express biologically active CD3 receptors but differ in their biochemical activation pathways during ligation of this receptor. Jurkat cells respond to anti-CD3 with Ca2+ mobilization, PKC activation, induction of protein tyrosine phosphorylation, and activation of newly characterized lymphoid microtubule associated protein-2 kinase (MAP-2K). MAP-2K itself is a 43-kDa phosphoprotein that requires tyrosine phosphorylation for activation. Although ligation of the CD3 receptor in HPB-ALL could stimulate tyrosine phosphorylation of a 59- kDa substrate, there was no associated induction of [Ca2+]i flux, PKC, or MAP-2K activation. A specific PKC agonist, PMA, which bypasses the CD3 receptor, could, however, activate MAP-2K in HPB-ALL cells. This implies that defective stimulation of PKC by the CD3 receptor is responsible for its failure to activate MAP-2K in HPB-ALL. The defect in PKC activation is likely distal to the CD3 receptor as A1F14- failed to activate MAP-2K in HPB-ALL but was effective in Jurkat cells. The stimulatory effect of PMA on MAP-2K activity in HPB-ALL was accompanied by tyrosine phosphorylation of this kinase which implies that PKC may, in some way, regulate tyrosine phosphorylation of MAP-2K. A candidate for this role is pp56lck which underwent posttranslational modification (seen as mobility change on SDS-PAGE) during anti-CD3 and PMA stimulation in Jurkat or PMA treatment in HPB-ALL. There was, in fact, exact coincidence between induction of PKC activity, posttranslational modification of lck and tyrosine phosphorylation/activation of MAP-2K. Lck kinase activity in an immune complex kinase assay was unchanged during PMA treatment. An alternative explanation is that modification of lck may alter its substrate profile. We therefore looked at the previously documented ability of PKC to dissociate lck from the CD4 receptor and found that PMA could reduce the stoichiometry of the lck interaction with CD4 in HPB-ALL and to a lesser extent in Jurkat cells. These results imply the existence of a kinase cascade that is initiated by PKC and, in the course of which, lck and MAP-2K may interact.     

The complete amino acid sequence of low molecular mass urokinase from human urine

The sequence of all 253 amino acids of the heavy (B-) chain of human urinary urokinase was determined. The fragmentation strategy employed included cyanogen bromide cleavage of S-carboxymethylated B-chain at Met and/or Trp residues, cleavage of acid-labile Asp-Pro bonds, and the use of the specific endoproteinases Lys-C and Arg-C for generation of overlapping fragments. For sequence determination automated solid- or liquid-phase techniques of Edman degradation were used. The amino acid sequence obtained substantiates the serine protease character of the B-chain of urokinase: a considerable homology with other serine proteinases, especially with the B-chain of human plasmin, was proved. The pertinent active site amino acids were localized: His-46, Asp-97, and Ser-198. A carbohydrate side chain, containing at least 4 glucosamine and 2 galactosamine residues, was demonstrated to be fixed at asparagine in position 144. The sequence data presented, together with the sequence of the second (A1-) chain of low molecular mass urokinase which was reported by us in an earlier communication, complete the knowledge of the whole primary structure of an active form of human urinary urokinase.     

New Palaeomacromiid Dragonflies from the Upper Paleocene of Argentina

A new genus of palaeomacromiid dragonflies, Curviarculia , based on Curviarculia delicata sp. nov. and Curviarculia lamasi sp. nov., is erected from the upper Paleocene Maíz Gordo Formation of north–western Argentina. Phylogenetic relationships within Palaeomacromiidae are discussed, leading to a new family diagnosis.     

Removal of Waterborne Human Enteric Viruses and Coliphages with Oxidized Coal

Human enteric viruses and coliphages have been detected in water that has undergone what is generally considered adequate treatment, including chlorination. Because small numbers of virus particles are needed for the initiation of a productive virus infection, the presence of any number of virus particles in water resources will always be of concern. In this investigation the ability of oxidized coal to remove viruses from water was investigated. The oxicoal product was found to be able to remove not only coliphages, but also various pathogenic human viruses from seeded water sources. Removal was dependent upon the type of virus, the period of exposure, and the concentration of oxidized coal. Received: 20 August 1997 / Accepted: 22 January 1998     

Allometric extension

Constants of allometric growth are commonly estimated by the first eigenvector of the covariance matrix of log measurements. Hills (1982, in Encyclopedia of Statistical Sciences, 48-54) defines a model of allometric extension for two related species by the conditions that (a) the constants of allometric growth are identical for both species and (b) the vector of mean differences is proportional to the common first eigenvector of both covariance matrices. We give a test for allometric extensionand discuss estimation of the parameters of the allometric extension model, including standard errors.