Prevalence, development, and molecular mechanisms of bacteriocin resistance in Campylobacter

Bacteriocins (BCNs) are antimicrobial peptides produced by bacteria with narrow or broad spectra of antimicrobial activity. Recently, several unique anti-Campylobacter BCNs have been identified from commensal bacteria isolated from chicken intestines. These BCNs dramatically reduced C. jejuni colonization in poultry and are being directed toward on-farm control of Campylobacter. However, no information concerning prevalence, development, and mechanisms of BCN resistance in Campylobacter exists. In this study, susceptibilities of 137 C. jejuni isolates and 20 C. coli isolates to the anti-Campylobacter BCNs OR-7 and E-760 were examined. Only one C. coli strain displayed resistance to the BCNs (MIC, 64 μg/ml), while others were susceptible, with MICs ranging from 0.25 to 4 μg/ml. The C. coli mutants resistant to BCN OR-7 also were obtained by in vitro selection, but all displayed only low-level resistance to OR-7 (MIC, 8 to 16 μg/ml). The acquired BCN resistance in C. coli could be transferred at intra- and interspecies levels among Campylobacter strains by biphasic natural transformation. Genomic examination of the OR-7-resistant mutants by using DNA microarray and random transposon mutagenesis revealed that the multidrug efflux pump CmeABC contributes to both intrinsic resistance and acquired resistance to the BCNs. Altogether, this study represents the first report of and a major step forward in understanding BCN resistance in Campylobacter, which will facilitate the development of effective BCN-based strategies to reduce the Campylobacter loads in poultry.     

Microtubules, motors, and mRNA localization mechanisms: watching fluorescent messages move.

Proper spatial and temporal localization of specific mRNAs is pivotal in the early stages of development. To dissect the mechanisms of localization, several groups are employing advanced fluorescence microscopy to track RNA movements in live oocytes and embryos.     

A biological interpretation of transient anomalous subdiffusion. I. Qualitative model

Anomalous subdiffusion has been reported for two-dimensional diffusion in the plasma membrane and three-dimensional diffusion in the nucleus and cytoplasm. If a particle diffuses in a suitable infinite hierarchy of binding sites, diffusion is well known to be anomalous at all times. But if the hierarchy is finite, diffusion is anomalous at short times and normal at long times. For a prescribed set of binding sites, Monte Carlo calculations yield the anomalous diffusion exponent and the average time over which diffusion is anomalous. If even a single binding site is present, there is a very short, almost artifactual, period of anomalous subdiffusion, but a hierarchy of binding sites extends the anomalous regime considerably. As is well known, an essential requirement for anomalous subdiffusion due to binding is that the diffusing particle cannot be in thermal equilibrium with the binding sites; an equilibrated particle diffuses normally at all times. Anomalous subdiffusion due to barriers, however, still occurs at thermal equilibrium, and anomalous subdiffusion due to a combination of binding sites and barriers is reduced but not eliminated on equilibration. This physical model is translated directly into a plausible biological model testable by single-particle tracking.     

Caenorhabditis elegans HOPS and CCZ-1 mediate trafficking to lysosome-related organelles independently of RAB-7 and SAND-1

As early endosomes mature, the SAND-1/CCZ-1 complex acts as a guanine nucleotide exchange factor (GEF) for RAB-7 to promote the activity of its effector, HOPS, which facilitates late endosome–lysosome fusion and the consumption of AP-3–containing vesicles. We show that CCZ-1 and the HOPS complex are essential for the biogenesis of gut granules, cell type–specific, lysosome-related organelles (LROs) that coexist with conventional lysosomes in Caenorhabditis elegans intestinal cells. The HOPS subunit VPS-18 promotes the trafficking of gut granule proteins away from lysosomes and functions downstream of or in parallel to the AP-3 adaptor. CCZ-1 also acts independently of AP-3, and ccz-1 mutants mistraffic gut granule proteins. Our results indicate that SAND-1 does not participate in the formation of gut granules. In the absence of RAB-7 activity, gut granules are generated; however, their size and protein composition are subtly altered. These observations suggest that CCZ-1 acts in partnership with a protein other than SAND-1 as a GEF for an alternate Rab to promote gut granule biogenesis. Point mutations in GLO-1, a Rab32/38-related protein, predicted to increase spontaneous guanine nucleotide exchange, specifically suppress the loss of gut granules by ccz-1 and glo-3 mutants. GLO-3 is known to be required for gut granule formation and has homology to SAND-1/Mon1–related proteins, suggesting that CCZ-1 functions with GLO-3 upstream of the GLO-1 Rab, possibly as a GLO-1 GEF. These results support LRO formation occurring via processes similar to conventional lysosome biogenesis, albeit with key molecular differences.     

Intracellular motility: A special delivery service

Recent studies have identified a delivery service that operates in specialised cell appendages: two motor proteins and a novel protein organelle use axonemal microtubules as tracks to shuttle essential components to the tips of flagella and the dendrites of sensory neurons.     

Transcriptional responses of Arabidopsis thaliana plants to As (V) stress

Background

Plants encode a large number of leucine-rich repeat receptor-like kinases. Legumes encode several LRR-RLK linked to the process of root nodule formation, the ligands of which are unknown. To identify ligands for these receptors, we used a combination of profile hidden Markov models and position-specific iterative BLAST, allowing us to detect new members of the CLV3/ESR (CLE) protein family from publicly available sequence databases.

Results

We identified 114 new members of the CLE protein family from various plant species, as well as five protein sequences containing multiple CLE domains. We were able to cluster the CLE domain proteins into 13 distinct groups based on their pairwise similarities in the primary CLE motif. In addition, we identified secondary motifs that coincide with our sequence clusters. The groupings based on the CLE motifs correlate with known biological functions of CLE signaling peptides and are analogous to groupings based on phylogenetic analysis and ectopic overexpression studies. We tested the biological function of two of the predicted CLE signaling peptides in the legume Medicago truncatula. These peptides inhibit the activity of the root apical and lateral root meristems in a manner consistent with our functional predictions based on other CLE signaling peptides clustering in the same groups.

Conclusion

Our analysis provides an identification and classification of a large number of novel potential CLE signaling peptides. The additional motifs we found could lead to future discovery of recognition sites for processing peptidases as well as predictions for receptor binding specificity.     

Survival of <Emphasis Type="Italic">Listeria monocytogenes</Emphasis> in Wilted and Additive-Treated Grass Silage

Grass was field-dried to 3 different dry matter (DM) levels (200, 430 and 540 g/kg) and inoculated with 10⁶–10⁷ cfu/g of a Listeria monocytogenes strain sharing a phagovar occasionally involved in food-borne outbreaks of listeriosis. Formic acid (3 ml/kg) or lactic acid bacteria (8·10⁵/g) with cellulolytic enzymes were applied only to forages with low and intermediate DM levels. Forages were ensiled in laboratory silos (1700 ml) and were stored at 25°C for 30 or 90 days. After 90 days of storage, L. monocytogenes could not be detected in any silo, except one with the high dry matter grass without additive. After 30 days of storage, between 10² and 10⁶ cfu L. monocytogenes/g silage were isolated from the untreated silages. Increasing the DM content from 200 to 540 g/kg did not reduce listeria counts possibly because of the lower production of fermentation acids (higher pH). In silages treated with additives, counts of L. monocytogenes were always lower than in silages without additive. In wet silages (DM 200 g/kg) both additives were effective, but in the wilted silages (DM 430 g/kg) only the bacterial additive reduced listeria counts below detection level. Listeria counts were highly correlated to silage pH (r = 0.92), the concentration of lactic acid (r = -0.80) and the pooled amount of undissociated acids (r = -0.83).     

Autoantibodies directed to novel components of the PM/Scl complex, the human exosome

The autoantigenic polymyositis/scleroderma (PM/Scl) complex was recently shown to be the human homologue of the yeast exosome, which is an RNA-processing complex. Our aim was to assess whether, in addition to targeting the known autoantigens PM/Scl-100 and PM/Scl-75, autoantibodies also target recently identified components of the PM/Scl complex. The prevalence of autoantibodies directed to six novel human exosome components (hRrp4p, hRrp40p, hRrp41p, hRrp42p, hRrp46p, hCsl4p) was determined in sera from patients with idiopathic inflammatory myopathy (n = 48), scleroderma (n = 11), or the PM/Scl overlap syndrome (n = 10). The sera were analyzed by enzyme-linked immunosorbent assays and western blotting using the affinity-purified recombinant proteins. Our results show that each human exosome component is recognized by autoantibodies. The hRrp4p and hRrp42p components were most frequently targeted. The presence of autoantibodies directed to the novel components of the human exosome was correlated with the presence of the anti-PM/Scl-100 autoantibody in the sera of patients with idiopathic inflammatory myopathy (IIM), as was previously found for the anti-PM/Scl-75 autoantibody. Other clear associations between autoantibody activities were not found. These results further support the conception that the autoimmune response may initially be directed to PM/Scl-100, whereas intermolecular epitope spreading may have caused the autoantibody response directed to the associated components.     

Diversity and Characterization of Sulfate-Reducing Bacteria in Groundwater at a Uranium Mill Tailings Site

Microbially mediated reduction and immobilization of U(VI) to U(IV) plays a role in both natural attenuation and accelerated bioremediation of uranium-contaminated sites. To realize bioremediation potential and accurately predict natural attenuation, it is important to first understand the microbial diversity of such sites. In this paper, the distribution of sulfate-reducing bacteria (SRB) in contaminated groundwater associated with a uranium mill tailings disposal site at Shiprock, N.Mex., was investigated. Two culture-independent analyses were employed: sequencing of clone libraries of PCR-amplified dissimilatory sulfite reductase (DSR) gene fragments and phospholipid fatty acid (PLFA) biomarker analysis. A remarkable diversity among the DSR sequences was revealed, including sequences from δ-Proteobacteria, gram-positive organisms, and the Nitrospira division. PLFA analysis detected at least 52 different mid-chain-branched saturate PLFA and included a high proportion of 10me16:0. Desulfotomaculum and Desulfotomaculum-like sequences were the most dominant DSR genes detected. Those belonging to SRB within δ-Proteobacteria were mainly recovered from low-uranium (≤302 ppb) samples. One Desulfotomaculum-like sequence cluster overwhelmingly dominated high-U (>1,500 ppb) sites. Logistic regression showed a significant influence of uranium concentration over the dominance of this cluster of sequences (P = 0.0001). This strong association indicates that Desulfotomaculum has remarkable tolerance and adaptation to high levels of uranium and suggests the organism's possible involvement in natural attenuation of uranium. The in situ activity level of Desulfotomaculum in uranium-contaminated environments and its comparison to the activities of other SRB and other functional groups should be an important area for future research.     

Supramolecular structures of peptide assemblies in membranes by neutron off-plane scattering: method of analysis

In a previous paper (Yang et al., Biophys. J. 75:641-645, 1998), we showed a simple, efficient method of recording the diffraction patterns of supramolecular peptide assemblies in membranes where the samples were prepared in the form of oriented multilayers. Here we develop a method of analysis based on the diffraction theory of two-dimensional liquids. Gramicidin was used as a prototype model because its pore structure in membrane in known. At full hydration, the diffraction patterns of alamethicin and magainin are similar to gramicidin except in the scale of q (the momentum transfer of scattering), clearly indicating that both alamethicin and magainin form pores in membranes but of different sizes. When the hydration of the multilayer samples was decreased while the bilayers were still fluid, the in-plane positions of the membrane pores became correlated from one bilayer to the next. We believe that this is a new manifestation of the hydration force. The effect is most prominent in magainin patterns, which are used to demonstrate the method of analysis. When magainin samples were further dehydrated or cooled, the liquid-like diffraction turned into crystal-like patterns. This discovery points to the possibility of investigating the supramolecular structures with high-order diffraction.