title: Integrative taxonomic approach to the cryptic diversity of Diplostomum spp. in lymnaeid snails from Europe with a focus on the 'Diplostomum mergi' species complex.
abstract: BACKGROUND
  Recent molecular studies have discovered substantial unrecognised diversity within the genus Diplostomum in fish populations in Europe and North America including three species complexes. However, data from the first intermediate host populations are virtually lacking. This study addresses the application of an integrative taxonomic approach to the cryptic species diversity of Diplostomum spp. in natural lymnaeid snail populations in Europe with a focus on the 'D. mergi' species complex.
  
  
  METHODS
  Totals of 1,909 Radix auricularia, 349 Radix peregra, 668 Stagnicola palustris and 245 Lymnaea stagnalis were sampled at five reservoirs of the Ruhr river system in Germany and screened for infections with Diplostomum spp. Cercariae were examined and identified alive, fixed and under scanning electron microscopy. Sequences from the barcode region of the cytochrome c oxidase subunit 1 (cox1) mitochondrial gene and from the internal transcribed spacer cluster (ITS1-5.8S-ITS2) of the rRNA gene were amplified for 51 and 13 isolates, respectively.
  
  
  RESULTS
  Detailed morphological and molecular analyses provided evidence for three named species (Diplostomum spathaceum, D. pseudospathaceum and D. parviventosum), and a further four species-level lineages ('D. mergi Lineages 2-4' and 'Diplostomum sp. Clade Q' in the lymnaeid snail populations from the Ruhr river basin. The paper provides the first descriptions of molecularly identified cercariae of D. spathaceum and of the cercariae of D. parviventosum, three lineages of the 'D. mergi' species complex and of 'Diplostomum sp. Clade Q'.
  
  
  CONCLUSIONS
  The integration of molecular and morphological evidence for Diplostomum spp. achieved in this study will serve as a baseline for species identification of these important parasites of snail and fish populations and thus advance further studies on the distribution of Diplostomum spp. in Europe.
authors.affiliation: Department of Aquatic Ecology and Centre for Water and Environmental Research (ZWU), University of Duisburg-Essen, Universitätsstraße 5, D-45141, Essen, Germany. christian.selbach@uni-due.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Selbach
authors.affiliation: Institute of Parasitology, Biology Centre of the Czech Academy of Sciences, Branišovská 31, 370 05, České Budějovice, Czech Republic. soldanova@paru.cas.cz.
authors.forename: Miroslava
authors.initials: M
authors.lastname: Soldánová
authors.affiliation: Institute of Parasitology, Biology Centre of the Czech Academy of Sciences, Branišovská 31, 370 05, České Budějovice, Czech Republic. georgieva@paru.cas.cz.
authors.forename: Simona
authors.initials: S
authors.lastname: Georgieva
authors.affiliation: Institute of Parasitology, Biology Centre of the Czech Academy of Sciences, Branišovská 31, 370 05, České Budějovice, Czech Republic. aneta.kostadinova@uv.es.
authors.forename: Aneta
authors.initials: A
authors.lastname: Kostadinova
authors.affiliation: Department of Aquatic Ecology and Centre for Water and Environmental Research (ZWU), University of Duisburg-Essen, Universitätsstraße 5, D-45141, Essen, Germany. bernd.sures@uni-due.de.|Department of Zoology, University of Johannesburg, Auckland Park 2006, PO Box 524, Johannesburg, South Africa. bernd.sures@uni-due.de.
authors.forename: Bernd
authors.initials: B
authors.lastname: Sures
mesh_terms: D000818:Animals; D044822:Biodiversity; D005260:Female; D005393:Fish Diseases / Q000469:parasitology*; D005399:Fishes / Q000145:classification / Q000469:parasitology*; D005858:Germany; D008297:Male; D008969:Molecular Sequence Data; D009656:North America; D010802:Phylogeny; D012908:Snails / Q000145:classification / Q000469:parasitology*; D014200:Trematoda / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D014201:Trematode Infections / Q000469:parasitology / Q000662:veterinary*