Persistence and dissemination of the multiple-antibiotic-resistance plasmid pB10 in the microbial communities of wastewater sludge microcosms

Plasmid-mediated dissemination of antibiotic resistance genes is widely recognized to take place in many environmental compartments but remains difficult to study in a global perspective because of the complexity of the environmental matrices considered and the lack of exhaustive tools. In this report, we used a molecular approach based on quantitative PCR to monitor the fate of the antibiotic resistance plasmid pB10 and its donor host in microbial communities collected from various wastewater treatment plant (WWTP) sludges and maintained in microcosms under different conditions. In aerated activated sludge microcosms, pB10 did not persist because of an apparent loss of the donor bacteria. The persistence of the donor bacteria noticeably increased in non-aerated activated sludge microcosms or after amending antibiotics (sulfamethoxazole or amoxicillin) at sub-inhibitory concentrations, but the persistence of the donor bacteria did not stimulate the dissemination of pB10. The dissemination of the plasmid appeared as an increasing plasmid to donor ratio in microcosm setups with microbial communities collected in anaerobic digesters or the spatially organized communities from fixed biofilm reactors. As a whole, the data collected suggest that some WWTP processes, more than others, may sustain microbial communities that efficiently support the dissemination of the multiple-antibiotic-resistance plasmid pB10.     

Phages and HIV-1: From Display to Interplay

The complex hide-and-seek game between HIV-1 and the host immune system has impaired the development of an efficient vaccine. In addition, the high variability of the virus impedes the long-term control of viral replication by small antiviral drugs. For more than 20 years, phage display technology has been intensively used in the field of HIV-1 to explore the epitope landscape recognized by monoclonal and polyclonal HIV-1-specific antibodies, thereby providing precious data about immunodominant and neutralizing epitopes. In parallel, biopanning experiments with various combinatorial or antibody fragment libraries were conducted on viral targets as well as host receptors to identify HIV-1 inhibitors. Besides these applications, phage display technology has been applied to characterize the enzymatic specificity of the HIV-1 protease. Phage particles also represent valuable alternative carriers displaying various HIV-1 antigens to the immune system and eliciting antiviral responses. This review presents and summarizes the different studies conducted with regard to the nature of phage libraries, target display mode and biopanning procedures.     

Spatial interaction modelling

Abstract. Spatial interaction (SI) is the process whereby entities at different points in physical space make contacts, demand/supply decisions or locational choices. The entities can be individuals or firms and the choices can include housing, jobs, production quantities, exports, imports, face-to-face contacts, schools, retail centres and activity centres. The first SI models can be grouped under the generic heading gravity models. Their main characteristic is that they model the behaviour of demand or supply segments, rather than that of individuals and firms. This article traces the development of these models from their inception in the early part of the twentieth century to the present. The key advances include the replacement of the gravity analogy by the more general concepts of entropy or information theory, a statistical framework commonly used in physics. With the arrival of the regional science paradigm over 50 years ago, a key challenge has been to broaden these models compared to those arising in spatial economics, thus arriving at a more inclusive probabilistic framework. These efforts are discussed here, as well as inclusion of geographical advances, embracing activities as generators of travel, time-geography, recognition of spatial interdependencies, and use of neuro-computing principles.     

Real-time amperometric analysis of reactive oxygen and nitrogen species released by single immunostimulated macrophages

Macrophages are key cells of the immune system. Immunologically activated macrophages are known to release a cocktail of reactive oxygen and nitrogen species. In this work, RAW 264.7 macrophages were activated by interferon-gamma and lipopolysaccharide, and the reactive mixture released by single cells was analyzed, in real time, by amperometry at platinized carbon microelectrodes. In comparison with untreated macrophages, significant increases in amperometric responses were observed for activated macrophages. Nitric oxide (NO*), nitrite (NO2*-), and peroxynitrite (ONOO-) were the main reactive species detected. The amounts of these reactive species were quantified, and their average fluxes released by a single, activated macrophage were evaluated. The detection of ONOO- is of particular interest, as its role and implications in various physiological conditions have been widely debated. Herein, direct evidence for the formation of ONOO- in stimulated macrophages is presented. Finally, the presence of 1400W, a selective inducible nitric oxide synthase (iNOS) inhibitor, led to an almost complete attenuation of the amperometric response of activated RAW 264.7 cells. The majority of the reactive species released by a macrophage are thus likely to be derived from NO* and superoxide (O2*-) co-produced by iNOS.     

Monitoring in real time with a microelectrode the release of reactive oxygen and nitrogen species by a single macrophage stimulated by its membrane mechanical depolarization

Macrophages are key cells of the immune system. During phagocytosis, the macrophage engulfs a foreign bacterium, virus, or particle into a vacuole, the phagosome, wherein oxidants are produced to neutralize and decompose the threatening element. These oxidants derive from in situ production of superoxide and nitric oxide by specific enzymes. However, the chemical nature and sequence of release of these compounds is far from being completely determined. The aim of the present work was to study the fundamental mechanism of oxidant release by macrophages at the level of a single cell, in real time and quantitatively. The tip of a microelectrode was positioned at a micrometric distance from a macrophage in a culture to measure oxidative-burst release by the cell when it was submitted to physical stimulation. The ensuing release of electroactive reactive oxygen and nitrogen species was detected by amperometry and the exact nature of the compounds was characterized through comparison with in vitro electrochemical oxidation of H2O2, ONOO-, NO*, and NO2(-) solutions. These results enabled the calculation of time variations of emission flux for each species and the reconstruction of the original flux of production of primary species, O2*- and NO*, by the macrophage.     

The fluorimetric FAST method, a simple tool for the optimization of microwave pasteurization of milk

The effect of microwave pasteurization of cow's milk on its nutritional quality was examined by the FAST method (Fluorescence of Advanced Maillard products and Soluble Tryptophan). Raw milk samples were submitted to different laboratory scale in batch microwave treatments using a central composite experimental design based on specific power and treatment time. The FAST index and bacterial count were monitored to assess protein denaturation, modification by the Maillard reaction and pasteurization efficiency, respectively. High discrimination between samples indicated that the FAST method is a potent tool for estimating the deterioration of the milk quality during experimental microwave treatment. Thus, the FAST index can be effectively used as the continuous response in experimental designs set up and to maximize information economically. In short, the FAST method allows us to retain the rapidity of experimental design while providing the advantages of convenience and low cost.     

Prevention of volatile fatty acids production and limitation of odours from winery wastewaters by denitrification

The effect of the addition of nitrate to winery wastewaters to control the formation of VFA in order to prevent odours during storage and treatment was studied in batch bioreactors at different NO(3)/chemical oxygen demand (COD) ratios and at full scale in natural evaporation ponds (2 x 7000 m(2)) by measuring olfactory intensity. In the absence of nitrate, butyric acid (2304 mgL(-1)), acetic acid (1633 mgL(-1)), propionic acid (1558 mgL(-1)), caproic acid (499 mgL(-1)) and valeric acid (298 mgL(-1)) were produced from reconstituted winery wastewater. For a ratio of NO(3)/COD=0.4 gg(-1), caproic and valeric acids were not formed. The production of butyric and propionic acids was reduced by 93.3% and 72.5%, respectively, at a ratio of NO(3)/COD=0.8, and by 97.4% and 100% at a ratio of NO(3)/COD=1.2 gg(-1). Nitrate delayed and decreased butyric acid formation in relation to the oxidoreduction potential. Studies in ponds showed that the addition of concentrated calcium nitrate (NITCAL) to winery wastewaters (3526 m(3)) in a ratio of NO(3)/COD=0.8 inhibited VFA production, with COD elimination (94%) and total nitrate degradation, and no final nitrite accumulation. On the contrary, in ponds not treated with nitrate, malodorous VFA (from propionic to heptano?c acids) represented up to 60% of the COD. Olfactory intensity measurements in relation to the butanol scale of VFA solutions and the ponds revealed the pervasive role of VFA in the odour of the untreated pond as well as the clear decrease in the intensity and not unpleasant odour of the winery wastewater pond enriched in nitrates. The results obtained at full scale underscored the feasibility and safety of the calcium nitrate treatment as opposed to concentrated nitric acid.     

Influence of a stormwater sediment deposit on microbial and biogeochemical processes in infiltration porous media

The purpose of this study was to test the relative influence of organic matter quantity and quality and the pollutant content of a stormwater sediment deposit on mineralization processes, microbial characteristics, and the release of solutes in infiltration sediment systems. In microcosm experiments, two other natural sediment deposits (one low and one rich particulate organic matter deposits) were studied to compare their effects with those of the stormwater deposit. The results showed that the biogeochemical processes (aerobic respiration, denitrification, fermentative processes), the microbial metabolism (enzymatic activities), and the releases of several solutes (NH(4)(+) and DOC) were stimulated in presence of the stormwater deposit and the natural particulate organic matter (POM)-rich deposit because of the quantity of the POM in these deposits. In the stormwater deposit, the high availability of the POM (indicated by its low C/N ratio and its high P content) produced a higher stimulation of the microbial metabolism than in presence of the POM-rich deposit (with a high C/N ratio). Pollutant (hydrocarbon and heavy metal) contents of the stormwater deposit did not have a significant effect on microbial processes. Thus, main effects of the stormwater sedimentary deposit on infiltration system were due to its organic matter characteristics (quantity and quality). Such organic matter characteristics need to be considered in future studies to determine the contamination potential of stormwater management practices.