title: Optimizing the design of a reproduction toxicity test with the pond snail Lymnaea stagnalis.
abstract: This paper presents the results from two ring-tests addressing the feasibility, robustness and reproducibility of a reproduction toxicity test with the freshwater gastropod Lymnaea stagnalis (RENILYS strain). Sixteen laboratories (from inexperienced to expert laboratories in mollusc testing) from nine countries participated in these ring-tests. Survival and reproduction were evaluated in L. stagnalis exposed to cadmium, tributyltin, prochloraz and trenbolone according to an OECD draft Test Guideline. In total, 49 datasets were analysed to assess the practicability of the proposed experimental protocol, and to estimate the between-laboratory reproducibility of toxicity endpoint values. The statistical analysis of count data (number of clutches or eggs per individual-day) leading to ECx estimation was specifically developed and automated through a free web-interface. Based on a complementary statistical analysis, the optimal test duration was established and the most sensitive and cost-effective reproduction toxicity endpoint was identified, to be used as the core endpoint. This validation process and the resulting optimized protocol were used to consolidate the OECD Test Guideline for the evaluation of reproductive effects of chemicals in L. stagnalis.
authors.affiliation: Univ Lyon, Université Lyon 1, UMR CNRS 5558, Laboratoire de Biométrie et Biologie Évolutive, F-69100 Villeurbanne, France. Electronic address: sandrine.charles@univ-lyon1.fr.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Charles
authors.affiliation: Institut National de la Recherche Agronomique (INRA), Centre de Recherche de Rennes, 65 rue de Saint-Brieuc, F-35042 Rennes, France; Bayer Aktiengesellschaft, Crop Science Division, BCS AG-R&D-D-EnSa-ETX-AQ, Alfred-Nobel Straße 50, D-40789 Monheim am Rhein, Germany.
authors.forename: Virginie
authors.initials: V
authors.lastname: Ducrot
authors.affiliation: Institut National de la Recherche Agronomique (INRA), Centre de Recherche de Rennes, 65 rue de Saint-Brieuc, F-35042 Rennes, France.
authors.forename: Didier
authors.initials: D
authors.lastname: Azam
authors.affiliation: The Food and Environment Research Agency (now Fera Science Ltd), Sand Hutton, York, YO41 1LZ, United Kingdom.
authors.forename: Rachel
authors.initials: R
authors.lastname: Benstead
authors.affiliation: Goethe University Frankfurt am Main, Department Aquatic Ecotoxicology, Max-von-Laue-Straße 13, D-60438 Frankfurt, Germany.
authors.forename: Denise
authors.initials: D
authors.lastname: Brettschneider
authors.affiliation: Laboratory of Environmental Toxicology and Aquatic Ecology, Faculty of Bioscience Engineering, Ghent University, Belgium.
authors.forename: Karel
authors.initials: K
authors.lastname: De Schamphelaere
authors.affiliation: Department of Biology & CESAM, Centre for Environmental and Marine Studies, University of Aveiro, 3810-193 Aveiro, Portugal.
authors.forename: Sandra
authors.initials: S
authors.lastname: Filipe Goncalves
authors.affiliation: DuPont, PO Box 60, 1090 Elkton Road, DuPont Stine-Haskell Research Center, S315/1369 Newark, Delaware, USA.
authors.forename: John W
authors.initials: JW
authors.lastname: Green
authors.affiliation: Department of Biology, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark.
authors.forename: Henrik
authors.initials: H
authors.lastname: Holbech
authors.affiliation: School of Biological Sciences, University of Plymouth, Plymouth, PL4 8AA, United Kingdom.
authors.forename: Thomas H
authors.initials: TH
authors.lastname: Hutchinson
authors.affiliation: Bayer Aktiengesellschaft, Crop Science Division, BCS AG-R&D-D-EnSa-ETX-AQ, Alfred-Nobel Straße 50, D-40789 Monheim am Rhein, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Faber
authors.affiliation: Departamento de Biologia, Universidade de Aveiro, 3810-193 Aveiro, Portugal.
authors.forename: Filipe
authors.initials: F
authors.lastname: Laranjeiro
authors.affiliation: Old School House, Brow Edge, Backbarrow, Ulverston, Cumbria LA128QX, United Kingdom.
authors.forename: Peter
authors.initials: P
authors.lastname: Matthiessen
authors.affiliation: Department of Pathology, Faculty of Veterinary Science, Swedish University of Agricultural Sciences, P.O. Box 7028, Uppsala, S-750 07, Sweden.
authors.forename: Leif
authors.initials: L
authors.lastname: Norrgren
authors.affiliation: Goethe University Frankfurt am Main, Department Aquatic Ecotoxicology, Max-von-Laue-Straße 13, D-60438 Frankfurt, Germany.
authors.forename: Jörg
authors.initials: J
authors.lastname: Oehlmann
authors.affiliation: Department of Environmental Toxicology, Texas Tech University, Lubbock, TX, USA.
authors.forename: Evelyn
authors.initials: E
authors.lastname: Reategui-Zirena
authors.affiliation: Ibacon GmbH, Arheilger Weg 17, 64380 Rossdorf, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Seeland-Fremer
authors.affiliation: Fraunhofer Institute for Molecular Biology and Applied Ecology, Department of Ecotoxicology, Auf dem Aberg 1, 57392 Schmallenberg, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Teigeler
authors.affiliation: University of Liège, Laboratory of Animal Ecology and Ecotoxicity (LEAE-CART), Allée du 6 Août, 11, Sart-Tilman, Belgium.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Thome
authors.affiliation: WIL Research, Department of In vitro and Environmental Toxicology, Ashland, United States.
authors.forename: Marysia
authors.initials: M
authors.lastname: Tobor Kaplon
authors.affiliation: BASF SE, Crop Protection - Ecotoxicology, Speyerer Straße 2, D-67117 Limburgerhof, Germany.
authors.forename: Lennart
authors.initials: L
authors.lastname: Weltje
authors.affiliation: Institut National de la Recherche Agronomique (INRA), Centre de Recherche de Rennes, 65 rue de Saint-Brieuc, F-35042 Rennes, France; Bayer Aktiengesellschaft, Crop Science Division, BCS AG-R&D-D-EnSa-ETX-AQ, Alfred-Nobel Straße 50, D-40789 Monheim am Rhein, Germany.
authors.forename: Laurent
authors.initials: L
authors.lastname: Lagadic
mesh_terms: D000818:Animals; D019256:Cadmium Chloride / Q000633:toxicity; D052284:Clutch Size / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005240:Feasibility Studies; D019983:Guideline Adherence; D017408:Guidelines as Topic; D007093:Imidazoles / Q000633:toxicity; D008195:Lymnaea / Q000187:drug effects*; D015233:Models, Statistical; D010063:Ovum / Q000187:drug effects; D012044:Regression Analysis; D015203:Reproducibility of Results; D012098:Reproduction / Q000187:drug effects; D012107:Research Design / Q000592:standards; D018570:Risk Assessment; D013997:Time Factors; D018675:Toxicity Tests / Q000379:methods* / Q000592:standards; D014204:Trenbolone Acetate / Q000633:toxicity; D014220:Trialkyltin Compounds / Q000633:toxicity; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D007093:Imidazoles; D014220:Trialkyltin Compounds; D014874:Water Pollutants, Chemical; C011559:tributyltin; C045362:prochloraz; D019256:Cadmium Chloride; D014204:Trenbolone Acetate
keywords: Count data; ECx; Fecundity; Mollusc; Test design optimization