The effects of a primary-treated municipal effluent on the immune system of rainbow trout (Oncorhynchus mykiss): exposure duration and contribution of suspended particles

Municipal sewage effluents are complex mixtures of contaminants known to disrupt both immune and endocrine functions in aquatic organisms. The present study sought to determine the impacts of municipal effluent on the immune systems of juvenile rainbow trout (Oncorhynchus mykiss), by exposing specimens to low concentrations (0.01%, 0.1%, 1% or 10%) of sewage effluent for periods of 28 or 90 days. The soluble and insoluble fractions of the effluent were also studied to assess the contribution of fractions rich in microorganisms and particles on fish immune systems. To this end, the trout were also exposed to soluble and insoluble fractions of the effluent for a period of 28 days. Immunocompetence was assessed by the following three parameters: phagocytosis, natural cytotoxic cells (NCC) and blastogenesis of lymphocytes under mitogen stimulation. Fish exposed to the 1% sewage effluent concentration for 28 days had enhanced phagocytic activity; at 90 days, phagocytic activity was reduced. T and B lymphocyte proliferation in fish from both groups was similarly stimulated. Phagocytosis and NCC activities were influenced more by the insoluble fraction than the soluble fraction of the effluent. Conversely, mitogen-stimulated T lymphocyte proliferation was enhanced in cells of fish exposed to the soluble fraction of the effluents, with a dampening effect on the insoluble (particulate) fraction of the effluent. In conclusion, the effects of the effluent and its fractions were higher at the cellular-mediated immunity level than at the acquired immunity level. Immunotoxicity data on the soluble fraction of the effluent were more closely associated to data on the unfractionated effluent, but the contribution of the particulate fraction could not be completely ignored for phagocytosis and B lymphocyte proliferation.     

Nitrogen catabolite repression modulates the production of aromatic thiols characteristic of Sauvignon Blanc at the level of precursor transport

The free thiols 3-mercapto-hexanol (3MH) and its acetate, practically absent from musts, are liberated by yeast during fermentation from a cysteinylated precursor [S-3-(hexan-1-ol)-l-cysteine (Cys-3MH)] present in the grape must and contribute favorably to the flavor of Sauvignon white wines. Production of 3MH is increased when urea is substituted for diammonium phosphate (DAP) as the sole nitrogen source on a synthetic medium. On grape must, complementation with DAP induces a decrease of 3MH production. This observation is reminiscent of nitrogen catabolite repression (NCR). The production of 3MH is significantly lower for a gap1Delta mutant compared with the wild type, during fermentation of a synthetic medium containing Cys-3MH as the precursor and urea as the sole nitrogen source. Mutants isolated from an enological strain with a relief of NCR on GAP1 produce significantly higher amounts of 3MH on synthetic medium than the parental strain. These phenotypes were not confirmed on grape must. It is concluded that on synthetic medium, Cys-3MH enters the cell through at least one identified transporter, GAP1p, whose activity is limiting the release of volatile thiols. On grape must, the uptake of the precursor through GAP1p is not confirmed, but the effect of addition of DAP, eventually prolonging NCR, is shown to decrease thiol production.     

Development of a high-performance liquid chromatographic method for the determination of a new potent radioiodinated melanoma imaging and therapeutic agent

N-(2-diethylaminoethyl)-6-iodoquinoxaline-2-carbamide (ICF 01012) is a new melanoma imaging agent showing promising properties for application in internal radionuclide therapy. We developed an analytical protocol for detection of ICF 01012 in biological samples using HPLC. The proposed method was first validated using standard of ICF 01012 and four potent metabolites of this compound and then applied to follow the metabolic fate of [(125)I]ICF 01012 after injection in melanoma-bearing mice. The results demonstrate that this method exhibits a good linearity (r(2)=0.9947), specificity and acceptable accuracy. This simple method appears convenient and sufficient for pharmacokinetic studies on [(125)I]ICF 01012.     

Strategies for improving production and purification of a recombinant protein: rP30 of Toxoplasma gondii expressed in the yeast Schizosaccharomyces pombe

Many problems concerned with the production and the purification of recombinant proteins must be addressed prior to launching an industrial production process. Among these problems, attention is focused on low-level expression that complicates the purification step and can jeopardise the process. The expression of a membrane protein, rP30, of Toxoplasma gondii in the yeast Schizosaccharomyces pombe led to a secretion of only 0.5 microg ml(-1). In order to obtain a sufficient quantity for biochemical characterization and evaluation in vitro diagnostic test development, strategies for both production and purification had to be optimized. First, the influence of four nitrogen sources (three peptones and yeast extract) on the growth rate, but also on the separation between the protein and the components of the fermentation broth was assessed. Second, batch and fed-batch fermentations were compared in terms of final biomass and rP30 concentrations. Third, three different protocols that included fixed and expanded bed ion exchange chromatography were compared for processing a large volume of feedstock. By using the most appropriate strategies, i.e. fed-batch fermentation, capture on EBA cation exchanger and affinity chromatography polishing, a purification factor of 1778 and a yield of 49% were achieved. These performances allowed a 12.5-fold increase for the overall rP30 process productivity.     

Surface expression of Hsp70B’ in response to proteasome inhibition in human colon cells

Hsp70B’ was expressed on the surface of HT-29 and CRL-1809 but not SW-480 human colon cell lines in response to proteasome inhibition as detected using flow cytometry. Surface expression was not detected under non-stress conditions nor was heat shock an inducer of surface expression in the three cell lines tested. Phylogenetic analysis indicated that the Hsp70B’ protein sequence was most closely related to another major inducible human Hsp70, Hsp72. Hsp70B’ appeared to be recently diverged, as homologs for Hsp70B’ have not been found in rodents. Hsp72 and Hsp70B’ shared 100% amino acid sequence identity in their predicted peptide-binding regions suggesting that they bind the same peptide substrates, perhaps in extracellular antigen presentation. Amino acid sequence differences were concentrated in the lid regions and the C-terminal domains raising the possibility that Hsp72 and Hsp70B’ bind different co-chaperones or cell surface receptors.     

Taxonomic distribution of large DNA viruses in the sea

Background  

Viruses are ubiquitous and the most abundant biological entities in marine environments. Metagenomics studies are increasingly revealing the huge genetic diversity of marine viruses. In this study, we used a new approach - 'phylogenetic mapping' - to obtain a comprehensive picture of the taxonomic distribution of large DNA viruses represented in the Sorcerer II Global Ocean Sampling Expedition metagenomic data set.     

Serologic survey for selected viral pathogens in free-ranging endangered European mink (Mustela lutreola) and other mustelids from south-western France

To investigate the possible role of selected pathogens in the decline of endangered European mink (Mustela lutreola) populations and the potential for these pathogens to affect mink survival, a serologic survey was conducted using serum samples collected from March 1996 to March 2003 in eight departments of south-western France. In total, 481 free-ranging individuals of five mustelid species (including the European mink) were tested. Sympatric mustelids can serve as sentinels to determine the presence of antibodies to viruses in the study area that could potentially infect mink. Antibodies to Canine distemper virus (CDV) were detected in all species; 9% of 127 European mink, 20% of 210 polecats (Mustela putorius), 5% of 112 American mink (Mustela vison), 33% of 21 stone marten (Martes foina) and 5% of 20 pine marten (Martes martes). Antibody prevalence was significantly higher in stone marten and polecats, possibly because their ranges overlap more closely with that of domestic species than that of the other species tested. Antibodies to Canine adenovirus were detected in all species but the pine marten; antibody prevalence estimates ranging from 2% to 10%. Antibodies to canine parainfluenza virus were detected in 1% of European mink, 1% of American mink and 5% of tested polecats but were not detected in Martes species. Antibodies to Rabies virus (RV) were detected in three animals, possibly because of interspecies transmission of bat lyssaviruses as the sampling area is considered to be free of RV, or to a lack of test specificity, as antibody titers were low. The high antibody prevalence to potentially lethal CDV suggests that this pathogen could have significant effects on the free-ranging populations and has implications for the conservation efforts for the endangered European mink.     

Diseases and reproductive success in a wild mammal: example in the alpine chamois

Density-dependent and climatic factors affect reproduction and dynamics of wild ungulates. Parasites can also decrease reproductive success through either a direct abortive effect or a negative impact on host growth and body condition. However, few studies have investigated the effect of parasitism on fecundity of ungulates in natural conditions. We studied three bacterial infections caused by Salmonella enterica serovar Abortusovis, Chlamydophila abortus and Coxiella burnetii. These bacteria are leading causes of reproductive failure in sheep, goat and cattle, which raises the question of their influence on population dynamics of wild ungulates. A long-term study of demography and epidemiology of an alpine chamois (Rupicapra rupicapra, L.) population (Les Bauges Reserve, France) and a generalized linear modeling approach were used to analyze the reproductive success of chamois according to population density, weather conditions and the prevalence of antibodies against the three bacteria in females. This approach enabled us to identify the confounding effect of weather and parasitism on fecundity in a natural population. After accounting for density, the prevalence of antibodies against the three bacteria explained 36% of the annual variation in reproductive success, and weather conditions explained an additional 31%. This study was, to our knowledge, the first to compare the decrease in fecundity due to bacterial infections and weather conditions in a population of wild mountain ungulates.