Electrophysiological characteristics of a population of mushroom body neurons in Apis mellifera L. honey bee under conditions of kynurenines deficit

Neurophysiological characteristics of mushroom body neurons were examined by electrophysiological methods in mutants snow(laranja) and wild-type honey bees. Mutation snow(laranja) causes a drastic decrease in the activity of enzyme tryptophane oxygenase that results in deficiency of all kynurenines. It also modifies bioelectrical properties of neurons in the mushroom bodies. The duration of afterdepolarization in spikes recorded from calyx neurons and the amplitude of postsynaptic potentials in these neurons evoked by focal stimulation of antennal lobes were shown to be most dependent on the mutation and, consequently, on the content of endogenous kynurenines. A tendency to an increase in the frequency of spontaneous spikes was also observed. The effect of the mutation on neurophysiological characteristics under study was recessive, i.e. it was observed only in homozygous individuals.     

New temperate Pseudomonas aeruginosa phage, phi297: Specific features of genome structure

The genome structure and some specific features of temperate Pseudomonas aeruginosa phage phi297 are considered. Analysis of sequencing data and genome annotation suggest that the phi297 genome displays a mosaic structure, which has formed through combining gene blocks from bacteria of taxonomically remote groups and/or their phages. The results of a comparison of the phi297 DNA homology level and pattern with the genome sequences of the currently known related P. aeruginosa bacteriophages are interpreted from the perspective of assumed active migration of these phages between different bacterial species.     

Identification of the UDP-N-Acetylglucosamine 4-Epimerase Involved in Exosporium Protein Glycosylation in Bacillus anthracis

Spores of Bacillus anthracis, the causative agent of anthrax, are enclosed by a loosely fitting exosporium composed of a basal layer and an external hair-like nap. The filaments of the nap are formed by trimers of the collagen-like glycoprotein BclA. The side chains of BclA include multiple copies of two linear rhamnose-containing oligosaccharides, a trisaccharide and a pentasaccharide. The pentasaccharide terminates with the unusual deoxyamino sugar anthrose. Both oligosaccharide side chains are linked to the BclA protein backbone through an N-acetylgalactosamine (GalNAc) residue. To identify the gene encoding the epimerase required to produce GalNAc for BclA oligosaccharide biosynthesis, three annotated UDP-glucose 4-epimerase genes of B. anthracis were cloned and expressed in Escherichia coli. The candidate proteins were purified, and their enzymatic activities were assessed. Only two proteins, encoded by the BAS5114 and BAS5304 genes (B. anthracis Sterne designations), exhibited epimerase activity. Both proteins were able to convert UDP-glucose (Glc) to UDP-Gal, but only the BAS5304-encoded protein could convert UDP-GlcNAc to UDP-GalNAc, indicating that BAS5304 was the gene sought. Surprisingly, spores produced by a mutant strain lacking the BAS5304-encoded enzyme still contained normal levels of BclA-attached oligosaccharides. However, monosaccharide analysis of the oligosaccharides revealed that GlcNAc had replaced GalNAc. Thus, while GalNAc appears to be the preferred amino sugar for the linkage of oligosaccharides to the BclA protein backbone, in its absence, GlcNAc can serve as a substitute linker. Finally, we demonstrated that the expression of the BAS5304 gene occurred in a biphasic manner during both the early and late stages of sporulation.Bacillus anthracis, the causative agent of anthrax, is a gram-positive, rod-shaped soil bacterium that forms spores when deprived of essential nutrients (15). Spore formation begins with an asymmetric septation that divides the developing cell into a smaller forespore compartment and a larger mother cell compartment, each containing a copy of the genome. The mother cell then engulfs the forespore and surrounds it with three protective layers: a cortex composed of peptidoglycan, a closely apposed proteinaceous coat, and a loosely fitting exosporium (11). Mother cell lysis releases the mature spore, which is dormant and capable of surviving in harsh environments for many years (16). When spores encounter an aqueous environment containing nutrients, they can germinate and grow as vegetative cells (23).Recently, interest in B. anthracis spores has intensified in response to their use as agents of bioterrorism. Of particular interest has been the outermost exosporium layer, which serves as a semipermeable barrier excluding potentially harmful macromolecules (9, 26) and as a vital first point of contact with the immune system of an infected host (14, 18, 32). The exosporium of B. anthracis and closely related species such as Bacillus cereus and Bacillus thuringiensis is a prominent structure comprised of a paracrystalline basal layer and an external hair-like nap (2). The basal layer contains approximately 20 different proteins (22, 25), while the filaments of the nap are formed by trimers of a single collagen-like glycoprotein called BclA (4, 27). The central region of BclA contains a large number of GXX repeats, mostly GTP triplets, and this region varies in length in naturally occurring strains of B. anthracis, resulting in hair-like naps of differing lengths (24, 28). Multiple copies of two O-linked oligosaccharides, a trisaccharide and a pentasaccharide, are attached to the protein component of BclA. The pentasaccharide side chains appear to be attached to threonine residues within the central region, while the trisaccharide side chains are attached to presently undefined residues in the protein (7).The precise structure of the trisaccharide side chain has not been determined, but its sequence is 3-O-methyl-l-rhamnose-l-rhamnose-N-acetylgalactosamine (GalNAc) (7). Except for a single glycosidic linkage, the structure of the pentasaccharide is known. Its structure is 2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-β-d-glucopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→2)-l-rhamnopyranosyl-(1→?)-N-acetylgalactosamine (7). Both oligosaccharides are attached to the BclA protein backbone through GalNAc residues. The pentasaccharide sugar 2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-d-glucose, which was given the trivial name anthrose, has been found only in B. anthracis strains and a limited number of highly pathogenic strains of B. cereus and B. thuringiensis (7, 8). For that reason, anthrose has joined other exosporium components as targets for the detection of B. anthracis spores and as new targets for therapeutic intervention in anthrax (6, 26, 29).In view of the potential importance of the BclA oligosaccharides, especially the anthrose-containing pentasaccharide, we have undertaken a comprehensive study of their biosynthesis. This effort involves identifying the biosynthetic genes for the three component sugars, anthrose, rhamnose, and GalNAc, as well as the genes involved in assembling the oligosaccharides and attaching them to the protein backbone of BclA. We recently reported the identification of a four-gene anthrose biosynthetic operon (8). A four-gene rhamnose biosynthetic operon has also been identified (24). This paper describes the identification of the gene encoding the UDP-N-acetylglucosamine (GlcNAc) 4-epimerase necessary for GalNAc biosynthesis. It also describes a surprising alternative BclA oligosaccharide biosynthetic pathway, which is active only in the absence of the UDP-GlcNAc 4-epimerase. Finally, this paper reports a biphasic pattern of expression of the gene encoding this epimerase during sporulation.     

Spatial-typological differentiation of ecosystems of the west siberian plain. Communication IV: Terrestrial vertebrates

Differentiation of the vertebrate communities almost coincides with the differentiation of ecosystems in vegetation at the type level, as judged by formalized classifications of various blocks of ecosystems of West Siberia by geobotanical map units, and it differs significantly from that in the underground component due to the greater effect of waterlogging on the latter. In invertebrate communities, significant differences are observed in the boreal-subboreal part, where waterlogging is more significant and greater similarity is found among middle and southern-taiga communities than among subtaiga-steppe ones. Over the groups of map units, the heterogeneity of the vertebrate communities differs from that in all the examined blocks of ecosystems in greater differentiation in the tundra zone. In the pretundra northern taiga subzones and from the middle taiga to the steppe zone, the heterogeneity of vertebrate communities is somewhat lower than that of vegetation, and especially in the underground block of ecosystems. However, these differences relate only to the hierarchy of the division, and often occur at the level of taxa of the rank of subtype or class. All these data indicate the relative independence of changes in the distinguished blocks of ecosystems, which gives rise to noncoincidence of the boundaries and sizes of taxa within them, as well as the continuality of the ecosystems as a whole. A significant difference is observed between the traditional geobotanical and typological-chorological zones of terrestrial vertebrate communities.     

Antibodies to gp350/220 enhance the ability of Epstein-Barr virus to infect epithelial cells

Epstein-Barr virus (EBV) is a persistent, orally transmitted herpesvirus that replicates in B cells and epithelial cells and is associated with lymphoid and epithelial malignancies. The virus binds to CD21 on B cells via glycoprotein gp350/220 and infects efficiently. Infection of cultured epithelial cells has not typically been efficient but can occur in the absence of gp350/220 and CD21 and in vivo is thought to be important to the development of nasopharyngeal carcinoma. We report here that antibodies to gp350/220, which inhibit EBV infection of B cells, enhance infection of epithelial cells. The effect is not mediated by Fc receptor binding but is further enhanced by antibody cross-linking, which may patch gp350/220 in the virus envelope. Saliva from EBV-seropositive individuals has similar effects that can be reversed by depletion of antibody. The results are consistent with a model in which gp350/220 interferes with the access of other important players to the epithelial cell surface. The results may have implications for the development of nasopharyngeal carcinoma in high-risk populations in which elevated titers of antibody to EBV lytic cycle proteins are prognostic.     

Molecular genetic evidence for the role of SGT1 in the intramolecular complementation of Bs2 protein activity in Nicotiana benthamiana

Pepper plants (Capsicum annuum) containing the Bs2 resistance gene are resistant to strains of Xanthomonas campestris pv vesicatoria (Xcv) expressing the bacterial effector protein AvrBs2. AvrBs2 is delivered directly to the plant cell via the type III protein secretion system (TTSS) of Xcv. Upon recognition of AvrBs2 by plants expressing the Bs2 gene, a signal transduction cascade is activated leading to a bacterial disease resistance response. Here, we describe a novel pathosystem that consists of epitope-tagged Bs2-expressing transgenic Nicotiana benthamiana plants and engineered strains of Pseudomonas syringae pv tabaci that deliver the effector domain of the Xcv AvrBs2 protein via the TTSS of P. syringae. This pathosystem has allowed us to exploit N. benthamiana as a model host plant to use Agrobacterium tumefaciens-mediated transient protein expression in conjunction with virus-induced gene silencing to validate genes and to identify protein interactions required for the expression of plant host resistance. In this study, we demonstrate that two genes, NbSGT1 and NbNPK1, are required for the Bs2/AvrBs2-mediated resistance responses but that NbRAR1 is not. Protein localization studies in these plants indicate that full-length Bs2 is primarily localized in the plant cytoplasm. Three protein domains of Bs2 have been identified: the N terminus, a central nucleotide binding site, and a C-terminal Leu-rich repeat (LRR). Co-immunoprecipitation studies demonstrate that separate epitope-tagged Bs2 domain constructs interact in trans specifically in the plant cell. Co-immunoprecipitation studies also demonstrate that an NbSGT1-dependent intramolecular interaction is required for Bs2 function. Additionally, Bs2 has been shown to associate with SGT1 via the LRR domain of Bs2. These data suggest a role for SGT1 in the proper folding of Bs2 or the formation of a Bs2-SGT1-containing protein complex that is required for the expression of bacterial disease resistance.     

A two-step purification procedure for sheep liver 6-phosphogluconate dehydrogenase

A two-step procedure for the purification of 6-phosphogluconate dehydrogenase (EC 1.1.1.44; 6-PGDH) from sheep liver is described. The enzyme is directly bound to cellulose phosphate by batch extraction and eluted with a linear salt gradient. Purification is completed by affinity chromatography using NADP(+)-agarose. The result is 6-PGDH of high purity, greatly increased yield, and the highest specific activity yet achieved, with a significant reduction in the purification time.     

Temporal variation in the genetic structure of host-associated populations of the small ermine moth Yponomeuta padellus (Lepidoptera, Yponomeutidae)

Temporal changes in allele frequencies were studied in host-associated populations of the small ermine moth Yponomeuta padellus. At one site, populations from three host plants (Sorbus aucuparia, Amelanchier larnarckii , and Crataegus spp.) were sampled annually during a four-year-period and analysed with 20 polymorphic allozyme markers. At two other sites, allele frequencies at 5- 6 enzyme loci of Y. padellus populations from two different host plants were also tested for consistency over a 13-year-pcriod. Significant allele frequency changes occurred in the short-term analysis, whereas allele frequencies remained relatively stable through time in the long-term analyses. Furthermore, allele frequencies of Y. padellus populations from Crataegus spp. were relatively stable compared to the other host populations. The role of the agents responsible for the observed patterns is discussed.