The cloning and expression of a gene encoding haemolytic activity from the fish pathogen Renibacterium salmoninarum

A gene encoding haemolytic activity from Renibacterium salmoninarum (strain PPD) was cloned into Escherichia coli using the cosmid vector pHC79, and subsequently subcloned on a 1.6 kbp SAlI fragment into pBR328. Southern blot hybridisation revealed that a homologous sequence is found in other strains of R. salmoninarum.     

Mucosal epithelial cells and Langerhans cells are targets for infection by the immunosuppressive type D retrovirus simian AIDS retrovirus serotype 1

Simian acquired immune deficiency syndrome (SAIDS) caused by the type D retrovirus SRV-1 results in opportunistic infections and a spectrum of oral lesions similar to those seen in humans with AIDS. To better understand the pathogenesis of these oral lesions we have retrospectively examined the oral mucosa from ten rhesus monkeys that died with SAIDS and prospectively examined the oral mucosa of ten additional animals inoculated with SRV-1 to determine at what time, and in what cells SRV-1 infection of the oral mucosa occurs. Using single and double label immunohistologic techniques, and electron microscopy we detected SRV-1 in clusters of oral epithelial cells and rare Langerhans cells as early as 1 month postinoculation.     

In vitro activity of 15-azasterol (A25822B) against chalkbrood pathogen Ascosphaera apis in the honey bee

The antifungal agent 15-azasterol A25822B was examined for effects on the growth and development of Ascosphaera apis. The minimum inhibition concentration (MIC) of azasterol against A. apis was 1 m. Growth and development of A. apis was completely controlled at this concentration. At a concentration of 0.01 m growth of A. apis was retarded and although sporocysts were formed developing spores were not be able to reach maturation. A major effect of azasterol at this low concentration was the accumulation of lipid in the hyphae, sporocysts and immature spores. In addition it caused a conformational change in mitochondria and damage to the spore membrane structure. On the basis of these results, further investigations of azasterol for the treatment of chalkbrood disease in the honey bee are warranted.Work was performed during sabbatical leave at the University of California, Davis.     

Isolation of a pathogenic clone of mouse mammary tumor virus.

Exogenous mouse mammary tumor virus (MMTV) was cloned from a GR mammary tumor. Clone lambda GRT39 contained a full-length integrated MMTV(GR) provirus and both 5' and 3' host flanking DNA. The lambda GRT39 provirus had no apparent structural changes associated with cloning and retained the exogenous MMTV gag gene poison sequence. When introduced into rat mammary adenocarcinoma LA7 cells, the lambda GRT39 provirus was fully expressed. lambda GRT39-transfected LA7 cells made MMTV RNA, had gp52 SU protein on the cell surface, and produced B-type retrovirus particles characteristic of MMTV. Mammary tumors developed in hormone-stimulated BALB/c females injected with MMTV from lambda GRT39-transfected LA7 cells [MMTV (lambda GRT39)]. The tumors had new, clonally integrated copies of the MMTV(lambda GRT39) provirus and were expressing MMTV antigen. These data indicate that the lambda GRT39 provirus is biologically active and pathogenic.     

Role of erythrocytes in leukocyte-endothelial interactions: mathematical model and experimental validation.

The binding of circulating cells to the vascular wall is a central process in inflammation, metastasis, and therapeutic cell delivery. Previous in vitro studies have identified the adhesion molecules on various circulating cells and the endothelium that govern the process under static conditions. Other studies have attempted to simulate in vivo conditions by subjecting adherent cells to shear stress as they interact with the endothelial cells in vitro. These experiments are generally performed with the cells suspended in Newtonian solutions. However, in vivo conditions are more complex because of the non-Newtonian flow of blood, which is a suspension consisting of 20-40% erythrocytes by volume. The forces imparted by the erythrocytes in the flow can contribute to the process of cell adhesion. A number of experimental and theoretical studies have suggested that the rheology of blood can influence the binding of circulating leukocytes by increasing the normal and axial forces on leukocytes or the frequency of their collision with the vessel wall, but there have been no systematic investigations of these phenomena to date. The present study quantifies the contribution of red blood cells (RBCs) in cell capture and adhesion to endothelial monolayers using a combination of mathematical modeling and in vitro studies. Mathematical modeling of the flow experiments suggested a physical mechanism involving RBC-induced leukocyte dispersion and/or increased normal adhesive contact. Flow chamber studies performed with and without RBCs in the suspending medium showed increases in wall collision and binding frequencies, and a decrease in rolling velocity in the presence of erythrocytes. Increased fluid viscosity alone did not influence the binding frequency, and the differences could not be attributed to large near-wall excesses of the lymphocytes. The results indicate that RBCs aid in the transport and initial engagement of lymphocytes to the vascular wall, modifying the existing paradigm for immune cell surveillance of the vascular endothelium by adding the erythrocyte as an essential contributor to this process.     

Low codon bias and high rates of synonymous substitution in Drosophila hydei and D. melanogaster histone genes

We have evaluated codon usage bias in Drosophila histone genes and have obtained the nucleotide sequence of a 5,161-bp D. hydei histone gene repeat unit. This repeat contains genes for all five histone proteins (H1, H2a, H2b, H3, and H4) and differs from the previously reported one by a second EcoRI site. These D. hydei repeats have been aligned to each other and to the 5.0-kb (i.e., long) and 4.8-kb (i.e., short) histone repeat types from D. melanogaster. In each species, base composition at synonymous sites is similar to the average genomic composition and approaches that in the small intergenic spacers of the histone gene repeats. Accumulation of synonymous changes at synonymous sites after the species diverged is quite high. Both of these features are consistent with the relatively low codon usage bias observed in these genes when compared with other Drosophila genes. Thus, the generalization that abundantly expressed genes in Drosophila have high codon bias and low rates of silent substitution does not hold for the histone genes.      

Increased cell surface hydrophobicity associated with possession of an additional surface protein by Aeromonas salmonicida

Abstract The cell surface of strains of Aeromonas salmonicida possessing an additional surface protein (A-protein) was shown to be more hydrophobic than strains devoid of this protein, using the techniques of phase partitioning, agglutination in the presence of ammonium sulphate and hydrophobic interaction chromatography.     

The Impact of Global Change Factors on Redox Signaling Underpinning Stress Tolerance

The plant cuticle is an extracellular hydrophobic layer that covers the aerial epidermis of all land plants, providing protection against desiccation and external environmental stresses. The past decade has seen considerable progress in assembling models for the biosynthesis of its two major components, the polymer cutin and cuticular waxes. Most recently, two breakthroughs in the long-sought molecular bases of alkane formation and polyester synthesis have allowed construction of nearly complete biosynthetic pathways for both waxes and cutin. Concurrently, a complex regulatory network controlling the synthesis of the cuticle is emerging. It has also become clear that the physiological role of the cuticle extends well beyond its primary function as a transpiration barrier, playing important roles in processes ranging from development to interaction with microbes. Here, we review recent progress in the biochemistry and molecular biology of cuticle synthesis and function and highlight some of the major questions that will drive future research in this field.The first plant colonizers of land, approximately 450 million years ago in the mid-Paleozoic era, faced a daunting set of challenges associated with their new terrestrial environment, including desiccation, temperature extremes, gravity, and increased exposure to UV radiation (Waters, 2003; Leliaert et al., 2011). The transition from an exclusively aquatic to a terrestrial life style, therefore, would have necessitated the evolution of a toolbox of morphological and physiological features, some of which are apparent through studies of the fossil record or by examining extant plant lineages. For example, the development of architecturally complex cell walls for biomechanical support and structural protection, which typify modern land plants, can be traced back to divergence and radiation within the Charophycean green algae, their immediate ancestors (Sørensen et al., 2011). However, the most critical adaptive trait for survival during terrestrialization would have been the ability to retain water in increasingly dehydrating habitats. Consequently, the capacity to synthesize, deposit, and maintain a hydrophobic surface layer, or cuticle, over the surfaces of aerial organs was arguably one of the most important innovations in the history of plant evolution. This idea is borne out by both fossil evidence (Edwards, 1993) and the ubiquity of cuticles among all extant embryophytes, from bryophytes (Budke et al., 2012) to angiosperms.Armed with a protective skin, together with a range of adaptive strategies for acquiring and conserving water, as well as for avoiding or tolerating water stress, embryophytes now thrive in a wide range of desiccating environments (Ogburn and Edwards, 2010; Aroca et al., 2012; Delaux et al., 2012; Jones and Dolan, 2012; Obata and Fernie, 2012; Gaff and Oliver, 2013). Accordingly, cuticles from a broad range of species, and in various ecological and agricultural contexts, have been studied from the perspective of their role as the primary barrier to transpirational water loss. However, it is now clear that cuticles play numerous other roles in plant development, physiology, and interactions with the abiotic environment and other organisms. Indeed, in recent years, there have been many instances of unexpected associations between the cuticle and diverse aspects of plant biology. In parallel, the past decade has seen considerable progress in understanding the biosynthesis of the major cuticle components and the complex regulatory networks that control cuticle synthesis and assembly.This review summarizes recent progress in elucidating the biochemistry and molecular biology of cuticle synthesis and function and highlights some of the connections to other aspects of plant biology, including signaling, pathogen defense, and development. Given the broad scope and space limitation, not every aspect of cuticle biosynthesis is covered in depth, and recent specialized reviews focusing on cuticle biomechanical properties (Domínguez et al., 2011), defensive functions (Reina-Pinto and Yephremov, 2009), and transport barrier properties (Burghardt and Riederer, 2006) may be of further interest. In addition, key ongoing questions in the field are discussed, and potential future approaches to resolving those questions are suggested.