Promotion of sustainable capture fisheries and aquaculture in Asian reservoirs and lakes

A collaborative international project funded by the European Union's INCO-DC programme is undertaking limnological, fish biological, environmental and socio-economic research in five tropical lakes and reservoirs in Sri Lanka, Thailand and the Philippines over the period 1998–2001. The aim is to determine their trophic structure and their capacity to sustain both their existing fisheries and present and future aquaculture. In some cases, these activities could potentially be expanded for the benefit of rural communities and of the local market within the bounds of social and environmental sustainability. This paper describes the concepts and methods involved in this innovative multidisciplinary project which aims to integrate limnological, fisheries and socio-economic issues in a comparative approach involving Asian and European research teams.Deceased.     

An analysis approach to identify specific functional sites in orthologous proteins using sequence and structural information: Application to neuroserpin reveals regions that differentially regulate inhibitory activity

The analysis of sequence conservation is commonly used to predict functionally important sites in proteins. We have developed an approach that first identifies highly conserved sites in a set of orthologous sequences using a weighted substitution‐matrix‐based conservation score and then filters these conserved sites based on the pattern of conservation present in a wider alignment of sequences from the same family and structural information to identify surface‐exposed sites. This allows us to detect specific functional sites in the target protein and exclude regions that are likely to be generally important for the structure or function of the wider protein family. We applied our method to two members of the serpin family of serine protease inhibitors. We first confirmed that our method successfully detected the known heparin binding site in antithrombin while excluding residues known to be generally important in the serpin family. We next applied our sequence analysis approach to neuroserpin and used our results to guide site‐directed polyalanine mutagenesis experiments. The majority of the mutant neuroserpin proteins were found to fold correctly and could still form inhibitory complexes with tissue plasminogen activator (tPA). Kinetic analysis of tPA inhibition, however, revealed altered inhibitory kinetics in several of the mutant proteins, with some mutants showing decreased association with tPA and others showing more rapid dissociation of the covalent complex. Altogether, these results confirm that our sequence analysis approach is a useful tool that can be used to guide mutagenesis experiments for the detection of specific functional sites in proteins. Proteins 2015; 83:135–152. © 2014 Wiley Periodicals, Inc.     

Characterization of Glycosaminoglycan (GAG) Sulfatases from the Human Gut Symbiont Bacteroides thetaiotaomicron Reveals the First GAG-specific Bacterial Endosulfatase

Despite the importance of the microbiota in human physiology, the molecular bases that govern the interactions between these commensal bacteria and their host remain poorly understood. We recently reported that sulfatases play a key role in the adaptation of a major human commensal bacterium, Bacteroides thetaiotaomicron, to its host (Benjdia, A., Martens, E. C., Gordon, J. I., and Berteau, O. (2011) J. Biol. Chem. 286, 25973–25982). We hypothesized that sulfatases are instrumental for this bacterium, and related Bacteroides species, to metabolize highly sulfated glycans (i.e. mucins and glycosaminoglycans (GAGs)) and to colonize the intestinal mucosal layer. Based on our previous study, we investigated 10 sulfatase genes induced in the presence of host glycans. Biochemical characterization of these potential sulfatases allowed the identification of GAG-specific sulfatases selective for the type of saccharide residue and the attachment position of the sulfate group. Although some GAG-specific bacterial sulfatase activities have been described in the literature, we report here for the first time the identity and the biochemical characterization of four GAG-specific sulfatases. Furthermore, contrary to the current paradigm, we discovered that B. thetaiotaomicron possesses an authentic GAG endosulfatase that is active at the polymer level. This type of sulfatase is the first one to be identified in a bacterium. Our study thus demonstrates that bacteria have evolved more sophisticated and diverse GAG sulfatases than anticipated and establishes how B. thetaiotaomicron, and other major human commensal bacteria, can metabolize and potentially tailor complex host glycans.     

Asymmetric lipid bilayer formation stabilized by DNA at the air/water interface

The lipid bis(guanidinium)-tren-cholesterol (BGTC) is a cationic cholesterol derivative bearing guanidinium polar headgroups used for gene transfection either alone or formulated as liposomes with the zwitterionic lipid 1,2-di-[cis-9-octadecenoyl]-sn-glycero-3-phosphoethanolamine (DOPE). Previous investigations have shown its ability to strongly interact with DNA and form asymmetric lipid bilayers at the air/water interface when mixed with DOPE. Here, with a view to further investigate its physicochemical behavior, we studied the interactions of mixtures of BGTC with another zwitterionic lipid, 1,2-Dimyristoyl-sn-Glycero-3-Phosphocholine, (DMPC), with DNA at the air/water interface by using the Langmuir monolayer technique coupled with Brewster Angle Microscopy (BAM) and Polarization Modulation Infra Red Reflexion Absorption (PMIRRAS) spectroscopy and we investigate DNA–BGTC/DMPC interactions. We demonstrate that when DNA is injected into the subphase in excess compared to the positive charges of BGTC, it adsorbs to BGTC/DMPC monolayers at 20 mN/m whatever the lipid monolayer composition (1/5, 2/3 or 3/2 BGTC/DMPC molar ratio) and forms an incomplete monolayer of either isotropic or anisotropic double strands depending on the BGTC content in the monolayer. Compression beyond the collapse of some mixed DNA–BGTC/DMPC (2/3 and 3/2 molar ratio) systems leads to the formation of DNA monolayers underneath asymmetric lipid bilayers characterized by a bottom layer of BGTC in contact with DNA and a top layer mainly constituted of DMPC.     

Fluorescent substrate analog for monitoring chain elongation by undecaprenyl pyrophosphate synthase in real time

Farnesyl pyrophosphate (FPP) is a common substrate for a variety of prenyltransferases for synthesizing isoprenoid compounds. In this study, (2E,6E)-8-O-(N-methyl-2-aminobenzoyl)-3,7-dimethyl-2,6-octandien-1-pyrophosphate (MANT-O-GPP), a fluorescent analog of FPP, was synthesized and demonstrated as a satisfactory substrate for Escherichia coli undecaprenyl pyrophosphate synthase (UPPS) with a K_m of 1.5 μM and a k_cat of 1.2 s⁻¹ based on [¹⁴C]IPP consumption. Interesting, we found that its emission fluorescence intensity at 420 nm increased remarkably during chain elongation, thereby useful for real-time monitoring kinetics of UPPS to yield a K_m of 1.1 μM and a k_cat of 1.0 s⁻¹, consistent with those measured using radiolabeled substrate. Using this assay, the IC₅₀ of a known UPPS inhibitor farnesyl thiopyrophosphate (FsPP) was confirmed. Our studies provide a convenient and environmentally friendly alternative for kinetics and inhibition studies on UPPS drug target.     

Novel Use of PIT Tags in Sea Cucumbers: Promising Results with the Commercial Species Cucumaria frondosa

The lack of a reliable and innocuous mark-recapture method has limited studies that would provide essential information for the management of commercial sea cucumbers. Tagging sea cucumbers is notoriously difficult because of their plastic nature and autolysis capacities. The markers that have so far been tested, mainly on or through the body wall, were either lost rapidly or had major drawbacks (e.g. suitable only for batch identification, requiring complex analysis, causing infections, necrosis, behavioural changes and mortality). The present study explored the efficacy of passive integrated transponder (PIT) tags for individually marking sea cucumbers by assessing retention rates and long-term side effects of tags inserted in previously unstudied tissues/organs. Individuals of the species Cucumaria frondosa were tagged in the body wall, aquapharyngeal bulb and at the base of the oral tentacles. They were monitored closely for evidence of stress, infection, change in feeding and spawning behaviour and tag retention rate. Implanting the tag in an oral tentacle to reach the hydrovascular system of the aquapharyngeal bulb achieved the best retention rates in full-size individuals: from a maximum of 92% after 30 days to 68% at the end of the experimental period (300 days). Efficacy was lower in smaller individuals (84% after 30 d and 42% after 300 d). Following a slight increase in cloacal movements for 15 h post tagging, no side effect was noted in sea cucumbers tagged in the aquapharyngeal bulb via the tentacles. Feeding and spawning behaviours were not affected and no signs of infections or abnormal cell development in the vicinity of the tags were observed. This study indicates that marking sea cucumbers with 8.2 mm long PIT tags implanted via the oral tentacle is an effective technique, yielding relatively high retention rates over long periods without any detectable physiological or behavioural effects.     

Identification of Mutations in TMEM5 and ISPD as a Cause of Severe Cobblestone Lissencephaly

Cobblestone lissencephaly is a peculiar brain malformation with characteristic radiological anomalies. It is defined as cortical dysplasia that results when neuroglial overmigration into the arachnoid space forms an extracortical layer that produces agyria and/or a “cobblestone” brain surface and ventricular enlargement. Cobblestone lissencephaly is pathognomonic of a continuum of autosomal-recessive diseases characterized by cerebral, ocular, and muscular deficits. These include Walker-Warburg syndrome, muscle-eye-brain disease, and Fukuyama muscular dystrophy. Mutations in POMT1, POMT2, POMGNT1, LARGE, FKTN, and FKRP identified these diseases as alpha-dystroglycanopathies. Our exhaustive screening of these six genes, in a cohort of 90 fetal cases, led to the identification of a mutation in only 53% of the families, suggesting that other genes might also be involved. We therefore decided to perform a genome-wide study in two multiplex families. This allowed us to identify two additional genes: TMEM5 and ISPD. Because TMEM has a glycosyltransferase domain and ISPD has an isoprenoid synthase domain characteristic of nucleotide diP-sugar transferases, these two proteins are thought to be involved in the glycosylation of dystroglycan. Further screening of 40 families with cobblestone lissencephaly identified nonsense and frameshift mutations in another four unrelated cases for each gene, increasing the mutational rate to 64% in our cohort. All these cases displayed a severe phenotype of cobblestone lissencephaly A. TMEM5 mutations were frequently associated with gonadal dysgenesis and neural tube defects, and ISPD mutations were frequently associated with brain vascular anomalies.     

Effects of soil warming and nitrogen foliar applications on bud burst of black spruce

Key message

In mature black spruce, bud burst process is anticipated by soil warming, while delayed by foliar applications of nitrogen; however, the effects depend on growth conditions at the site.

Abstract

The observation of phenological events can be used as biological indicator of environmental changes, especially from the perspective of climate change. In boreal forests, the onset of the bud burst is a key factor in the length of the growing season. With current climate change, the major factors limiting the growth of boreal trees (i.e., temperature and nitrogen availability) are changing and studies on mature trees are limited. The aim of this study was to investigate the effects of soil warming and increased nitrogen (N) deposition on bud burst of mature black spruce [Picea mariana (Mill.) BSP]. From 2008 onwards, an experimental manipulation of these environmental growth conditions was conducted in two stands (BER and SIM) at different altitudes in the boreal forest of Quebec, Canada. An increase in soil temperature (H treatment) and a canopy application of artificial rain enriched with nitrogen (N treatment) were performed. Observations of bud phenology were made during May–July 2012 and 2013. In BER, H treatment caused an anticipation (estimated as 1–3 days); while N treatment, a delay (estimated as 1–2 days but only in 2012) in bud burst. No treatments effect was significant in SIM. It has been demonstrated that soil temperature and N availability can play an important role in affecting bud burst in black spruce but the effects of these environmental factors on growth are closely linked with site conditions.