Distribution of Neoparamoeba sp. in sediments around marine finfish farming sites in Tasmania

Marine sediment samples collected from various sites at 2 Atlantic salmon farms in Tasmania were analysed for the presence of Neoparamoeba sp., an amoeba associated with amoebic gill disease (AGD) in farmed Atlantic salmon. Environmental variables of the sediment layer at each site, including redox potential and sulphide concentration, were measured and the general biological condition assessed by video observation. Sediments and environmental data were collected on 4 occasions at each site over a 12 mo period. Neoparamoeba sp. was detected in populations of amoebae recovered by culture from all sites and in 50% of all sediment samples taken. There was evidence of a seasonal influence on the presence of the amoeba, but this was different at each farm. No Neoparamoeba sp. was recovered from any sites at Farm 1 during the winter of 2002 whereas at Farm 2 this was the case for the summer of 2003. There appeared to be no relationship between the presence of Neoparamoeba sp., salmon farming activities and environmental parameters.     

Reinforcement of reproductive isolation between adjacent populations in the Park Grass Experiment

It has been debated, ever since Charles Darwin and Alfred Russell Wallace disagreed about the matter, whether natural selection plays a role in reinforcing reproductive isolation during the earliest stages of speciation. Recent theory suggests that it can do so, but until now the empirical evidence has conspicuously lacked a case in which reinforcement has actually been observed to split a population. We show that this has occurred at least once in populations of the grass Anthoxanthum odoratum growing in the Park Grass Experiment where flowering time has shifted at the boundaries between plots. As a consequence, gene flow via pollen has been severely limited and adjacent populations that had a common origin at the start of the experiment in 1856 have now diverged at neutral marker loci.     

Commensal Bacteria Lipoteichoic Acid Increases Skin Mast Cell Antimicrobial Activity against Vaccinia Viruses

Mast cells (MCs) are considered sentinels in the skin and mucosa. Their ability to release antimicrobial peptides, such as cathelicidin, protects against bacterial infections when the epithelial barrier is breached. We recently described that MCs defend against bacterial and viral infections through the release of cathelicidin during degranulation. In this study, we hypothesize that cathelicidin expression is induced in MCs by the activation of TLR2 from bacterial products (lipoteichoic acid) produced by commensal bacteria at the epithelial surface. Our research shows that signaling through TLR2 increases the production and expression of cathelicidin in mast cells, thereby enhancing their capacity to fight vaccinia virus. MCs deficient in cathelicidin were less efficient in killing vaccinia virus after lipoteichoic acid stimulation than wild-type cells. Moreover, the activation of TLR2 increases the MC recruitment at the skin barrier interface. Taken together, our findings reveal that the expression and control of antimicrobial peptides and TLR signaling on MCs are key in fighting viral infection. Our findings also provide new insights into the pathogenesis of skin infections and suggest potential roles for MCs and TLR2 ligands in antiviral therapy.     

Mutational analysis of conserved glycine residues 142, 143 and 146 reveals Gly(142) is critical for tetramerization of CTP synthase from Escherichia coli

CTPS (cytidine 5'-triphosphate synthase) catalyses the ATP-dependent formation of CTP from UTP using either ammonia or L-glutamine as the nitrogen source. Binding of the substrates ATP and UTP, or the product CTP, promotes oligomerization of CTPS from inactive dimers to active tetramers. In the present study, site-directed mutagenesis was used to replace the fully conserved glycine residues 142 and 143 within the UTP-binding site and 146 within the CTP-binding site of Escherchia coli CTPS. CD spectral analyses of wild-type CTPS and the glycine mutants showed a slight reduction of approximately 15% in alpha-helical content for G142A and G143A relative to G146A and wild-type CTPS, suggesting some local alterations in structure. Relative to wild-type CTPS, the values of k(cat)/K(m) for ammonia-dependent and glutamine-dependent CTP formation catalysed by G143A were reduced 22- and 16-fold respectively, whereas the corresponding values for G146A were reduced only 1.4- and 1.8-fold respectively. The glutaminase activity (k(cat)) of G146A was similar to that exhibited by the wild-type enzyme, whereas that of G143A was reduced 7.5-fold. G146A exhibited substrate inhibition at high concentrations of ammonia and a partial uncoupling of glutamine hydrolysis from CTP production. Although the apparent affinity (1/[S](0.5)) of G143A and G146A for UTP was reduced approximately 4-fold, G146A exhibited increased co-operativity with respect to UTP. Thus mutations in the CTP-binding site can affect UTP-dependent activity. Surprisingly, G142A was inactive with both ammonia and glutamine as substrates. Gel-filtration HPLC experiments revealed that both G143A and G146A were able to form active tetramers in the presence of ATP and UTP; however, nucleotide-dependent tetramerization of G142A was significantly impaired. Our observations highlight the sensitivity of the structure of CTPS to mutations in the UTP- and CTP-binding sites, with Gly(142) being critical for nucleotide-dependent oligomerization of CTPS to active tetramers. This 'structural sensitivity' may limit the number and/or types of mutations that could be selected for during the development of resistance to cytotoxic pyrimidine nucleotide analogues.     

Effect of Compost and Manure Soil Amendments on Nematodes and on Yields of Potato and Barley: A 7-Year Study

A 7-year study located in Prince Edward Island, Canada, examined the influence of compost and manure on crop yield and nematode populations. The compost used in this study consisted of cull waste potatoes, sawdust, and beef manure in a 3:3:1 ratio, respectively. No plant-parasitic nematodes were detected in samples collected from windrow compost piles at 5- and 30-cm depths prior to application on field plots. Low population densities of bacterial-feeding nematodes were recovered from compost windrows at the 5-cm depth. Field plots of potato (Solanum tuberosum cv. Kennebec) received compost applied at 16 metric tonnes per hectare, or beef manure applied at 12 metric tonnes per hectare. An adjacent trial with barley (Hordeum vulgare cv. Mic Mac) received only the compost treatment. In both trials the experimental design was a complete randomized block with four replicates. Data averaged over seven growing seasons indicated that population levels of root-lesion nematodes (primarily Pratylenchus penetrans) were higher in root-zone soil in potato plots treated with either compost or manure compared to the untreated control plots. The soil amendments did not affect root-knot nematode (Meloidogyne hapla) population densities in the potato plots, but clover-cyst nematodes (Heterodera trifolii) were more numerous in the root-zone soils of barley treated with compost compared to the untreated plots. Numbers of bacterial-feeding nematodes (primarily Diplogaster lheritieri) were greater in soil in potato plots treated with manure and in soil around barley roots than in untreated plots. Total yields of potato tubers averaged over seven growing seasons increased by 27% in the plots treated with either compost or manure. Grain yields of barley also were increased by 12% when compost was applied. These results indicated that organic amendments increased crop yields, but the impacts on different nematode species varied and usually increased soil population levels.     

Studies on an antineoplastic fraction from human urine. Characterization of the major protein in this fraction.

A fraction has been isolated from human urine which exhibits antiproliferative activity against human tumour cell lines without affecting the growth of several normal diploid cell lines or tumour cells of mouse or hamster origin. The major protein present in this fraction has been characterized and tentatively designated antineoplastic urinary protein (ANUP). An S020,W value of 3.69 S was obtained by sedimentation velocity analysis, and a subunit molecular mass of 16 300 Da was obtained by sedimentation equilibrium and by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Centrifugation data also indicated that the protein self-associates. The amino acid analysis of ANUP was consistent with its low pI (4.2) as determined by chromatofocusing analysis. Furthermore, the amino acid composition exhibited some features similar to collagen, as shown by high levels of proline and glycine, the absence of cysteine, and the presence of low levels of hydroxyproline.