Potential of Marine Strains of Pseudoalteromonas to Improve Resistance of Juvenile Sea Bass to Pathogens and Limit Biofilm Development.

The European sea bass (Dicentrarchus labrax), one of the most produced marine fish species in Europe, is acutely vulnerable to multiple infectious hazards. In this study, we investigated the potential probiotic effect of some marine Pseudoalteromonas bacterial strains against two major pathogens of this species, Vibrio harveyi and the nervous necrosis virus (NNV), and examined their antibiofilm effect. Impregnation phase was done by repeated immersion of juvenile's sea bass during 8 to 12 weeks in seawater containing the probiotic candidates at a concentration of 106 CFU/mL. Four candidates were tested: (1) a combination of two strains producing antimicrobial compounds, hCg-42 and hOe-125; (2) strain 3J6, with known antibiofilm properties; (3) strain RA15, from the same genus, but with no identified probiotic effect; and (4) a control group without probiotics. At the end of the impregnation phase, fish underwent an infection challenge with V. harveyi or with a pathogenic strain of NNV and mortality was monitored. For the V. harveyi challenge, improved survival rates of 10 and 25% were obtained for the RA15 and the mix hCg-42 + hOe-125-impregnated groups, respectively. For the NNV challenge, no significant benefic effect of the probiotics on infection kinetics or cumulative mortality was observed. At the end of the impregnation phase, the maximal thickness of biofilm was significantly lower in the 3J6, double strain, and RA15 groups, compared with the non-impregnated control group. This study highlights the interesting probiotic potential of marine bacteria to limit mortalities induced by bacterial pathogens as well as biofilm development.

Evaluation of in vitro tests to assess the efficacy of formulations as topical skin protectants against organophosphorus compounds.

Prevention of exposure to the neurotoxic organophosphorus compounds (OP) is a major concern both for pesticide users and soldiers. Skin barrier creams are being developed to complement or replace uncomfortable chemical protective suits. Quick evaluation of formulations efficacy mainly relies on in vitro tests which lead to consistent, complementary and relevant results. The objectives of this work were to determine the consistency of results from in vitro tests and importance of the formulation composition in the skin protective efficacy. The efficacy of three formulations, i.e. oil-in-water and water-in-oil emulsions and perfluorinated compounds-based cream, was evaluated against the OP paraoxon in vitro. Our results indicated that the least effective formulations could be quickly identified by performing in vitro permeation tests with silicone membrane and by evaluating interfacial interactions between formulations and OP. Among the tested formulations, the perfluorinated compounds-based cream could have a broader spectrum of efficacy than emulsions against OP and other toxic chemicals.

In vitro selection and efficacy of topical skin protectants against the nerve agent VX.

Against highly toxic chemicals that are quickly absorbed in the skin, topical formulations could adequately complement specific protective suits and equipments. In this work, we evaluated in vitro and compared the skin protection efficacy against the nerve agent VX of four different topical formulations: oil-in-water and water-in-oil emulsions, a perfluorinated-based cream and a hydrogel. Semi-permeable silicone membrane, pig-ear and human abdominal split-thickness skin samples mounted in diffusion cells were compared as in vitro permeation tests. The results showed that silicone membrane could be used instead of skin samples to screen for potentially effective formulations. However, the results indicated that due to potentially significant interactions between formulations and skin, relevant ranking of formulations according to their protective efficacy could require tests with skin samples. The main phase of emulsions, water or oil, was not found to be critical for skin protective efficacy against VX. Instead, specific film-forming ingredients such as perfluorinated-based polymers and silicones could significantly affect the skin protective efficacy of formulations. We showed that a hydrogel containing specific hydrophilic polymers was by far the most effective of the formulations evaluated against VX skin permeation in vitro.

Percutaneous penetration and distribution of VX using in vitro pig or human excised skin validation of demeton-S-methyl as adequate simulant for VX skin permeation investigations.

The organophosphorus (OP) chemical warfare V agent O-ethyl-S-[2(di-isopropylamino)ethyl] methyl phosphonothioate (VX), is a highly toxic compound which mainly penetrates the body via percutaneous pathways. Hence, the following prerequisite: to ascertain compound absorption and percutaneous profile distribution with a view to further assessing the efficacy of topical skin protectants. We first selected the most appropriate receptor fluid to carry out in vitro VX absorption experiments, namely: Hanks's Balanced Salt Solution (HBSS). After a 24-h topical exposure time lapse, we measured altogether the percentage of applied dose unabsorbed and absorbed, penetration rate, lag time, permeability coefficient (K(p)), and dose of VXeq present in skin. To such an end, we used full-thickness and split-thickness pig-ear or human abdominal skin membranes. Further, we scrutinised the potential use of two specific molecules as suitable surrogates for VX percutaneous penetration analyses: thus, we compared the present VX toxicokinetic parameters to earlier findings from our research unit, with respect to OP insecticides demethon-S-methyl (DSM) and paraoxon (POX). Within the framework of our study, we wish to highlight the following evidence: (a) pig-ear skin proves a relevant model to predict in vitro human abdominal skin, taking into account a 2-fold higher skin permeability to VXeq; (b) both full or split-thickness skin membranes may be used indiscriminately to gauge penetration rate and absorbed dose; (c) DSM applied on full-thickness pig-ear skin is the most relevant model to mimic the in vitro VX absorption through full-thickness skin model.

In vitro percutaneous penetration of organophosphorus compounds using full-thickness and split-thickness pig and human skin.

Organophosphorus compounds (OPs), such as pesticides and chemical warfare agents like sarin (GB), soman (GD) and VX, are highly toxic compounds. The OP vapours and their liquid forms are readily absorbed through the skin, therefore, protecting the skin of people who are potentially exposed to these agents is crucial. The development of effective countermeasures relies on a better knowledge of the percutaneous penetration of such molecules. The purpose of this present study is to determine the in vitro percutaneous penetration parameters of two pesticides DSM and DFP, as potential simulants of V and G agents, respectively, using four in vitro systems: full-thickness and split-thickness human abdominal and pig-ear skin membranes mounted on static diffusion cells. Based on the toxicokinetic parameters of the percutaneous penetration of DSM and DFP, we demonstrated that (a) pig-ear skin is a relevant model to predict the in vitro human skin permeability taking into account a 2-fold difference between these two species (b) both full and split-thickness skin membranes could be used indiscriminately, (c) DSM and DFP would be appropriate surrogates for V and G agents to perform skin permeation studies.

Percutaneous penetration and absorption of parathion using human and pig skin models in vitro and human skin grafted onto nude mouse skin model in vivo.

This study determined and compared the percutaneous penetration and absorption of an organophosphorus (OP) pesticide, parathion (PA), using three experimental skin models: namely the human abdominal- and pig-ear skin in vitro models and the Human Skin grafted onto a nude mouse (HuSki) in vivo model. The percentage of topically applied dose absorbed and the doses present in the stratum corneum and skin were systematically determined at 24 h under similar experimental conditions. The three experimental skin models were first compared. Then, the advantages of the HuSki model for in vivo PA skin absorption studies were evaluated compared with the pig in vivo model previously used by others. Lastly, the relevance of each skin model to predict the permeability of human skin to PA in vivo was assessed by comparing our results with previously published in vivo human volunteer values. It was demonstrated that (a) pig-ear skin is relevant for predicting the in vitro human abdominal skin absorption taking into account a 2-3 times higher skin permeability to PA, (b) using ethanol as the vehicle, the absorption of PA was 4-5 times higher in the HuSki model than in the pig model but supports the usefulness of the HuSki model to easy mass balance studies, (c) both human in vitro and HuSki models closely predict the in vivo human volunteer absorption at 24 h when acetone is used as a vehicle but the HuSki model overcomes the known limitations of in vitro models for studying the fate of PA in the different skin layers after topical application.