Microbial stratification in low pH oxic and suboxic macroscopic growths along an acid mine drainage

Macroscopic growths at geographically separated acid mine drainages (AMDs) exhibit distinct populations. Yet, local heterogeneities are poorly understood. To gain novel mechanistic insights into this, we used OMICs tools to profile microbial populations coexisting in a single pyrite gallery AMD (pH ∼2) in three distinct compartments: two from a stratified streamer (uppermost oxic and lowermost anoxic sediment-attached strata) and one from a submerged anoxic non-stratified mat biofilm. The communities colonising pyrite and those in the mature formations appear to be populated by the greatest diversity of bacteria and archaea (including ‘ARMAN'' (archaeal Richmond Mine acidophilic nano-organisms)-related), as compared with the known AMD, with ∼44.9% unclassified sequences. We propose that the thick polymeric matrix may provide a safety shield against the prevailing extreme condition and also a massive carbon source, enabling non-typical acidophiles to develop more easily. Only 1 of 39 species were shared, suggesting a high metabolic heterogeneity in local microenvironments, defined by the O₂ concentration, spatial location and biofilm architecture. The suboxic mats, compositionally most similar to each other, are more diverse and active for S, CO₂, CH₄, fatty acid and lipopolysaccharide metabolism. The oxic stratum of the streamer, displaying a higher diversity of the so-called ‘ARMAN''-related Euryarchaeota, shows a higher expression level of proteins involved in signal transduction, cell growth and N, H₂, Fe, aromatic amino acids, sphingolipid and peptidoglycan metabolism. Our study is the first to highlight profound taxonomic and functional shifts in single AMD formations, as well as new microbial species and the importance of H₂ in acidic suboxic macroscopic growths.     

Pleomorphism in type I collagen fibrils produced by persistence of the procollagen N-propeptide

The assembly of type I collagen and type I pN-collagen was studied in vitro using a system for generating these molecules enzymatically from their immediate biosynthetic precursors. Collagen generated by C-proteinase digestion of pC-collagen formed D-periodically banded fibrils that were essentially cylindrical (i.e. circular in cross-section). In contrast, pN-collagen generated by C-proteinase digestion of procollagen formed thin, sheet-like structures that were axially D-periodic in longitudinal section, of varying lateral widths (up to several microns) and uniform in thickness (approximately 8 nm). Mixtures of collagen and pN-collagen assembled to form a variety of pleomorphic fibrils. With increasing pN-collagen content, fibril cross-sections were progressively distorted from circular to lobulated to thin and branched structures. Some of these structures were similar to fibrils observed in certain heritable disorders of connective tissue where N-terminal procollagen processing is defective. The observations are considered in terms of the hypothesis that the N-propeptides are preferentially located on the surface of a growing assembly. The implications for normal diameter control of collagen fibrils in vivo are discussed.     

Distinct roles of beta1 metal ion-dependent adhesion site (MIDAS), adjacent to MIDAS (ADMIDAS), and ligand-associated metal-binding site (LIMBS) cation-binding sites in ligand recognition by integrin alpha2beta1

Integrin-ligand interactions are regulated in a complex manner by divalent cations, and previous studies have identified ligand-competent, stimulatory, and inhibitory cation-binding sites. In collagen-binding integrins, such as alpha2beta1, ligand recognition takes place exclusively at the alpha subunit I domain. However, activation of the alphaI domain depends on its interaction with a structurally similar domain in the beta subunit known as the I-like or betaI domain. The top face of the betaI domain contains three cation-binding sites: the metal-ion dependent adhesion site (MIDAS), the ADMIDAS (adjacent to MIDAS), and LIMBS (ligand-associated metal-binding site). The role of these sites in controlling ligand binding to the alphaI domain has yet to be elucidated. Mutation of the MIDAS or LIMBS completely blocked collagen binding to alpha2beta1; in contrast mutation of the ADMIDAS reduced ligand recognition but this effect could be overcome by the activating monoclonal antibody TS2/16. Hence, the MIDAS and LIMBS appear to be essential for the interaction between alphaI and betaI, whereas occupancy of the ADMIDAS has an allosteric effect on the conformation of betaI. An activating mutation in the alpha2 I domain partially restored ligand binding to the MIDAS and LIMBS mutants. Analysis of the effects of Ca(2+), Mg(2+), and Mn(2+) on ligand binding to these mutants showed that the MIDAS is a ligand-competent site through which Mn(2+) stimulates ligand binding, whereas the LIMBS is a stimulatory Ca(2+)-binding site, occupancy of which increases the affinity of Mg(2+) for the MIDAS.     

Integrin αvβ1 Is a Receptor for Foot-and-Mouth Disease Virus

Infection by field strains of Foot-and-mouth disease virus (FMDV) is initiated by binding to certain species of arginine-glycine-aspartic acid (RGD)-dependent integrin including alphavbeta3 and the epithelial integrin alphavbeta6. In this report we show that the integrin alphavbeta1, when expressed as a human/hamster heterodimer on transfected CHOB2 cells, is a receptor for FMDV. Virus binding and infection mediated by alphavbeta1 was inefficient in the presence of physiological concentrations of calcium and magnesium but were significantly enhanced by reagents that activate the integrin and promote ligand binding. The ability of chimeric alpha5/alphav integrin subunits, in association with the beta1 chain, to bind FMDV and mediate infection matched the ligand binding specificity of alphavbeta1, not alpha5beta1, thus providing further evidence for the receptor role of alphavbeta1. In addition, data are presented suggesting that amino acid residues near the RGD motif may be important for differentiating between the binding specificities of alphavbeta1 and alphavbeta6.     

The effect of supplementing maize stover with cowpea ( Vigna unguiculata) haulms on the intake and growth performance of Ethiopian sheep

This study compared the effect of supplementing maize stover (MS) with cowpea (Vigna unguiculata) haulms or commercial concentrate (CC) on feed intake, nutrient digestibility, live weight gain and carcass yield of male Ethiopian Highland sheep. Two cowpea genotypes, 12688 (forage) and IT96D-774 (dual-purpose), were used. A randomised block design was applied with groups of eight sheep, blocked by weight, allocated to one of six treatments; MS ad libitum either unsupplemented or supplemented daily with 150 or 300 g dry matter (DM) of either cowpea or CC. MS contained more neutral detergent fibre (NDF), acid detergent fibre (ADF) and lignin than either cowpeas or CC. Crude protein (CP) content of the forage-type cowpeas was higher than either dual-purpose or CC, while MS had the lowest CP content. Relative to the negative control group, cowpea at either level significantly (P < 0.01) increased both MS intake and total NDF and lignin. Supplementation significantly (P < 0.01) increased nitrogen (N) intakes relative to the negative control, with N intake for CC and dual-purpose cowpea (high level) being similar to the intakes for cowpeas at 150 g. N intake with the forage-type cowpea offered at higher levels was significantly (P < 0.01) greater than the other groups. No significant differences (P > 0.01) in MS intake were identified between cowpeas at either level or CC and, although intake level of CC increased, it did not differ significantly from the negative control group. Supplementation significantly (P < 0.01) improved average daily gain, with the negative control group losing weight over the experimental period, and increased final live weight, carcass cold weight and dressing percentage. Supplementation significantly improved the apparent digestibility of DM, organic matter and NDF, with no significant difference found between cowpeas at either level. N retention was negative for sheep offered only MS, but positive with all supplements, with cowpeas improving N retention to a greater extent than CC. Interestingly, N retention/N intake was higher with cowpeas offered at the lower level suggesting an improvement in utilisation efficiency. The results indicate that the supplementation of MS with cowpea enhanced ruminant production through improvements in digestibility and intake. Further, as production improvements associated with the two levels of supplementation did not differ significantly, it is suggested that where limited quantities of cowpea are available, it may be of greater nutritional benefit to offer smaller quantities over an increased number of animal days.     

Plasma IP-10, apoptotic and angiogenic factors associated with fatal cerebral malaria in India

Background

Knowledge on insecticide resistance in target species is a basic requirement to guide insecticide use in malaria control programmes. Malaria transmission in the Mekong region is mainly concentrated in forested areas along the country borders, so that decisions on insecticide use should ideally be made at regional level. Consequently, cross-country monitoring of insecticide resistance is indispensable to acquire comparable baseline data on insecticide resistance.

Methods

A network for the monitoring of insecticide resistance, MALVECASIA, was set up in the Mekong region in order to assess the insecticide resistance status of the major malaria vectors in Cambodia, Laos, Thailand, and Vietnam. From 2003 till 2005, bioassays were performed on adult mosquitoes using the standard WHO susceptibility test with diagnostic concentrations of permethrin 0.75% and DDT 4%. Additional tests were done with pyrethroid insecticides applied by the different national malaria control programmes.

Results

Anopheles dirus s.s., the main vector in forested malaria foci, was susceptible to permethrin. However, in central Vietnam, it showed possible resistance to type II pyrethroids. In the Mekong delta, Anopheles epiroticus was highly resistant to all pyrethroid insecticides tested. It was susceptible to DDT, except near Ho Chi Minh City where it showed possible DDT resistance. In Vietnam, pyrethroid susceptible and tolerant Anopheles minimus s.l. populations were found, whereas An. minimus s.l. from Cambodia, Laos and Thailand were susceptible. Only two An. minimus s.l. populations showed DDT tolerance. Anopheles vagus was found resistant to DDT and to several pyrethroids in Vietnam and Cambodia.

Conclusion

This is the first large scale, cross-country survey of insecticide resistance in Anopheles species in the Mekong Region. A unique baseline data on insecticide resistance for the Mekong region is now available, which enables the follow-up of trends in susceptibility status in the region and which will serve as the basis for further resistance management. Large differences in insecticide resistance status were observed among species and countries. In Vietnam, insecticide resistance was mainly observed in low or transmission-free areas, hence an immediate change of malaria vector control strategy is not required. Though, resistance management is important because the risk of migration of mosquitoes carrying resistance genes from non-endemic to endemic areas. Moreover, trends in resistance status should be carefully monitored and the impact of existing vector control tools on resistant populations should be assessed.     

A Serological Survey of Ruminant Livestock in Kazakhstan During Post-Soviet Transitions in Farming and Disease Control

The results of a serological survey of livestock in Kazakhstan, carried out in 1997–1998, are reported. Serum samples from 958 animals (cattle, sheep and goats) were tested for antibodies to foot and mouth disease (FMD), bluetongue (BT), epizootic haemorrhagic disease (EHD), rinderpest (RP) and peste des petits ruminants (PPR) viruses, and to Brucella spp. We also investigated the vaccination status of livestock and related this to changes in veterinary provision since independence in 1991. For the 2 diseases under official surveillance (FMD and brucellosis) our results were similar to official data, although we found significantly higher brucellosis levels in 2 districts and widespread ignorance about FMD vaccination status. The seroprevalence for BT virus was 23%, and seropositive animals were widespread suggesting endemicity, despite the disease not having being previously reported. We found a few seropositives for EHDV and PPRV, which may suggest that these diseases are also present in Kazakhstan. An hierarchical model showed that seroprevalence to FMD and BT viruses were clustered at the farm/village level, rather than at a larger spatial scale. This was unexpected for FMD, which is subject to vaccination policies which vary at the raion (county) level.     

Synthesis and biological evaluation of chalcones as inhibitors of the voltage-gated potassium channel Kv1.3

Chalcone derivatives of the natural product khellinone were synthesised and screened for bioactivity against the voltage-gated potassium channel Kv1.3. X-ray crystallography was employed to investigate relationships between the structure and function of a selection of the reported chalcones.     

An Arabidopsis gene encoding a chloroplast-targeted beta-amylase

beta-Amylase is one of the most abundant starch degrading activities found in leaves and other plant organs. Despite its abundance, most if not all of this activity has been reported to be extrachloroplastic and for this reason, it has been assumed that beta-amylases are not involved in the metabolism of chloroplast-localized transitory leaf starch. However, we have identified a novel beta-amylase gene, designated ct-Bmy, which is located on chromosome IV of Arabidopsis thaliana. Ct-Bmy encodes a precursor protein which contains a typical N-terminal chloroplast import signal and is highly similar at the amino acid level to extrachloroplastic beta-amylases of higher plants. Expression of the ct-Bmy cDNA in E. coli confirmed that the encoded protein possesses beta-amylase activity. CT-BMY protein, synthesized in vitro, was efficiently imported by isolated pea chloroplasts and shown to be located in the stroma. In addition, fusions between the predicted CT-BMY transit peptide and jellyfish green fluorescent protein (GFP) or the entire CT-BMY protein and GFP showed accumulation in vivo in chloroplasts of Arabidopsis. Expression of the GUS gene fused to ct-Bmy promoter sequences was investigated in transgenic tobacco plants. GUS activity was most strongly expressed in the palisade cell layer in the leaf blade and in chlorenchyma cells associated with the vascular strands in petioles and stems. Histochemical staining of whole seedlings showed that GUS activity was largely confined to the cotyledons during the first 2 weeks of growth and appeared in the first true leaves at approximately 4 weeks.