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Der mitteleuropäische Wasserfroschkomplex umfasst Pelophylax lessonae (Genotyp LL), Pelophylax ridibundus (RR) und deren Hybriden Pelophylax esculentus (LR), der sich hemiklonal durch Rückkreuzen mit LL in lessonae-esculentus (L-E)-Populationen oder RR in ridibundus-esculentus (R-E)-Populationen reproduziert. Außerdem können Hybriden in reine Hybridpopulationen (E) bilden, in denen triploide Individuen die Elternarten funktionell ersetzen. Bislang ist wenig über die molekularen Mechanismen bekannt, die der klonalen Gametenbildung in der Keimbahn der Hybridform zugrunde liegen. In dieser Studie wurden erstmalig 160 Gene der Elternarten untersucht, die an der Gametenbildung beteiligt sind. Zusätzlich wurden 131 SNPs von 52 dieser Gene von 652 Wasserfröschen aus 26 Populationen analysiert. Im Ergebnis wurden 14 SNPs von 10 Genen entdeckt, deren Frequenzen mit dem Populationssystem assoziiert waren. In Übereinstimmung mit ihren Funktionen könnten diese Gene im Zusammenheng mit den system-spezifischen hybridogenetischen Reproduktionsmodi stehen. Sowohl transkriptomische als auch SNP-Daten lieferten Hinweise auf genetische Introgression, d. h. einen Transfer lessonae-spezifischer Allelen in den ridibundus-Genpool oder umgekehrt. Außerdem wurde bei P. lessonae eine kryptische genetische Diversität beobachtet. SNP-Analysen ergaben auch, dass LR-Individuen aus E-Populationen eine höhere genetische Ähnlichkeit mit LR-Individuen aus R-E als aus L-E-Populationen aufweisen. Diese Ergebnisse werfen die Frage nach dem Ursprung der E-Populationen auf, von denen bisher angenommen wurde, dass sie aus L-E-Populationen hervorgegangen sind. Die neuen molekularen Daten stehen mit den hemiklonalen Fortpflanzungsmodi von P. esculentus im Einklang und unterstreichen, dass diese wahrscheinlich auf komplexen Wechselwirkungen zwischen verschiedenen Genen und Faktoren basieren., The Central European water frog complex comprises Pelophylax lessonae (genotype LL), Pelophylax ridibundus (RR), and their hybridogenetic hybrid Pelophylax esculentus (LR), which reproduces by backcrossing with LL in lessonae-esculentus (L-E) populations or RR in ridibundus-esculentus (R-E) populations. In addition, hybrids are able to reproduce in all-hybrid (E) populations in which triploid individuals functionally replace the parental species. To date, little is known about the molecular mechanisms underlying clonal gamete formation in the hybrid germline. In this study, 160 genes from P. lessonae and P. ridibundus known to be involved in gametogenesis were characterized for the first time. In addition, 131 single nucleotide polymorphisms (SNP) from 52 of these genes were analyzed, derived from 652 water frogs of 26 populations. As a result of logistic regressions, 14 SNPs of 10 genes were detected whose frequencies were associated with the population system. Consistent with their functions during gametogenesis, these genes can be considered as candidates associated with the system-specific hybridogenetic modes, i.e. clonal inheritance of the L and/or R genomes. Both transcriptomic and SNP data provided evidence for genetic introgression, i.e. a transfer of lessonae-specific alleles into the ridibundus gene pool or vice versa. In addition, cryptic genetic diversity was observed in P. lessonae. SNP analysis also revealed that LR individuals from E populations exhibit higher genetic similarity to LR individuals from R-E than L-E populations where triploid frogs do not occur. These findings raise the question of the origin of E populations, which were previously thought to have arisen from L-E populations after the emergence of triploid hybrids. The new molecular data are consistent with the reproductive modes of P. esculentus and emphasize that clonal inheritance is most likely caused by complex interactions of different genes and genetic factors.

Elektronisches Supplement zur Dissertation "Neue Einblicke in die molekularen Grundlagen der Gametogenese des hybridogenetischen Wasserfroschs Pelophylax esculentus". Die Dateien enthalten: e1: Probeninformationen. e2: Codierende Sequenzen gametogenetischer Gene. e3: Codierende Sequenzen respiratorischer Gene, e4: Referenzsequenz für das initiale Read-Mapping mit dem Programm Geneious. e5: Referenzsequenzen für Pelophylax lessonae. e6: Referenzsequenzen für P. ridibundus. e7: Single Nucleotid Polymorphismen (SNP), die für die Genotyp- und Ploidiebestimmung verwendet wurden. e8: Master-Tabelle aller SNPs und genotypisierten Individuen. e9: Charakteristika der untersuchten Gene., Electronic supplementary of the dissertation "New insights into the molecular basis of gametogenesis of the hybridogenetic water frog Pelophylax esculentus". The files contain: e1: Sample information. e2: Coding sequences of gametogenic genes. e3: Coding sequences of respiratory genes. e4: Reference sequence for initial read mapping with Geneious. e5: Reference sequences for Pelophylax lessonae. e6: Reference sequences for P. ridibundus. e7: Single nucleotide polymorphisms (SNP) used for genotype and ploidy determination. e8: Master table of all SNPs and individuals genotyped. e9: Characteristics of the genes studied.

Purpose The study aims to characterise and compare the helminth assemblages and helminth infracommunities in the marsh frog, Pelophylax ridibundus and the edible frog, P. esculentus collected in the northern part of Ukraine. Methods Occurrence and abundance of the helminths were analysed by calculating the prevalence, intensity, and mean abundance of infection; similarities between the infracommunities were estimated by the Bray–Curtis index and visualised using nMDS plots. Dissimilarities were estimated using the ANOSIM and SIMPER routines. Results In total, 27 helminth species were found in 143 frogs. Pelophylax ridibundus (n = 86) harboured 20 species of helminths, 24 species were found in P. esculentus (n = 57), and 17 species were shared by the two hosts. Oswaldocruzia bialata and larval Strigea sp. were absent in P. ridibundus, while they reached the prevalence of 30% and 10%, respectively, in P. esculentus. Cosmocerca ornata, Diplodiscus subclavatus, Opisthioglyphe ranae, and Codonocephalus urniger had significantly larger prevalence in P. ridibundus, whereas Haematoloechus asper was found to be more prevalent in P. esculentus. Acanthocephalus ranae, Icosiella neglecta, Haematoloechus variegatus, Pleurogenes claviger, Pleurogenoides medians, and Prosotocus confusus were equally common in both hosts. Helminth infracommunities in the two hosts had identical species richness (1–10 species, 4 on average); abundance was significantly higher in P. ridibundus. Conclusions Helminth assemblages of the two hosts in northern Ukraine are rather similar; however, small but significant differences were found in their species composition, parameters of infection in some species, and structure of helminth infracommunities

Purpose The study aims to characterise and compare the helminth assemblages and helminth infracommunities in the marsh frog, Pelophylax ridibundus and the edible frog, P. esculentus collected in the northern part of Ukraine. Methods Occurrence and abundance of the helminths were analysed by calculating the prevalence, intensity, and mean abundance of infection; similarities between the infracommunities were estimated by the Bray–Curtis index and visualised using nMDS plots. Dissimilarities were estimated using the ANOSIM and SIMPER routines. Results In total, 27 helminth species were found in 143 frogs. Pelophylax ridibundus (n = 86) harboured 20 species of helminths, 24 species were found in P. esculentus (n = 57), and 17 species were shared by the two hosts. Oswaldocruzia bialata and larval Strigea sp. were absent in P. ridibundus, while they reached the prevalence of 30% and 10%, respectively, in P. esculentus. Cosmocerca ornata, Diplodiscus subclavatus, Opisthioglyphe ranae, and Codonocephalus urniger had significantly larger prevalence in P. ridibundus, whereas Haematoloechus asper was found to be more prevalent in P. esculentus. Acanthocephalus ranae, Icosiella neglecta, Haematoloechus variegatus, Pleurogenes claviger, Pleurogenoides medians, and Prosotocus confusus were equally common in both hosts. Helminth infracommunities in the two hosts had identical species richness (1–10 species, 4 on average); abundance was significantly higher in P. ridibundus. Conclusions Helminth assemblages of the two hosts in northern Ukraine are rather similar; however, small but significant differences were found in their species composition, parameters of infection in some species, and structure of helminth infracommunities

The Major Histocompatibility Complex (MHC) is a genomic region encoding immune loci that are important and frequently used markers in studies of adaptive genetic variation and "disease resistance. Given the primary role of infectious diseases in contributing to global amphibian declines, we characterized the hypervariable exon 2 and flanking introns of the MHC Class II beta chain for 17 species of frogs in the Ranidae, a speciose and cosmopolitan family facing widespread pathogen infections and declines. We find high levels of genetic variation concentrated in the Peptide Binding Region (PBR) of the exon. Ten codons are under positive selection, nine of which are located in the mammal-defined PBR. We hypothesize that the tenth codon (residue 21) is an amphibian-specific PBR site that may be important in disease resistance. Trans-species and trans-generic polymorphisms are evident from exon-based genealogies, and co-phylogenetic analyses between intron, exon and mitochondrial based reconstructions reveal incongruent topologies, likely due to different locus histories. We developed two sets of barcoded adapters that reliably amplify a single and likely functional locus in all screened species using both 454 and Illumina based sequencing methods. These primers provide a resource for multiplexing and directly sequencing hundreds of samples in a single sequencing run, avoiding the labour and chimeric sequences associated with cloning, and enabling MHC population genetic analyses. Although the primers are currently limited to the 17 species we tested, these sequences and protocols provide a useful genetic resource and can serve as a starting point for future disease, adaptation and conservation studies across a range of anuran taxa.

We present the first comprehensive DNA barcoding study of German reptiles and amphibians representing likewise the first on the European herpetofauna. A total of 248 barcodes for all native species and subspecies in the country and a few additional taxa were obtained in the framework of the projects ‘Barcoding Fauna Bavarica’ (BFB) and ‘German Barcode of Life’ (GBOL). In contrast to many invertebrate groups, the success rate of the identification of mitochondrial lineages representing species via DNA barcode was almost 100% because no cases of Barcode Index Number (BIN) sharing were detected within German native reptiles and amphibians. However, as expected, a reliable identification of the hybridogenetic species complex in the frog genus Pelophylax was not possible. Deep conspecific lineages resulting in the identification of more than one BIN were found in Lissotriton vulgaris, Natrix natrix and the hybridogenetic Pelophylax complex. A high variety of lineages with different BINs was also found in the barcodes of wall lizards (Podarcis muralis), confirming the existence of many introduced lineages and the frequent occurrence of multiple introductions. Besides the reliable species identification of all life stages and even of tissue remains, our study highlights other potential applications of DNA barcoding concerning German amphibians and reptiles, such as the detection of allochthonous lineages, monitoring of gene flow and also noninvasive sampling via environmental DNA. DNA barcoding based on COI has now proven to be a reliable and efficient tool for studying most amphibians and reptiles as it is already for many other organism groups in zoology.

Water frogs inhabiting Cyprus represent a distinct evolutionary species of Messinian origin that is formally described in this paper. The systematic status of Cypriot frogs is evidenced by specific characters in their mitochondrial (mt) and nuclear (nu) DNA sequences, and the fact that they form a well supported monophyletic clade in both mtDNA and nuDNA phylogenies. While genetic data revealed clear and reproducible differences between this new taxon and all other western Palearctic water frog species including Pelophylax bedriagae in the Levant and two Anatolian water frogs lineages (P. cf. bedriagae -1 and P. cf. bedriagae -2), there is no diagnostic morphological or morphometric character that allows a clear discrimination between Cyprus frogs and frogs from the adjacent mainland. If several morphometric indices are combined as predictor variables in a discriminant analysis, however, both females and males of Cypriot water frogs are correctly distinguished from the other eastern Mediterranean lineages. While phylogenies based on concatenated sequences of two mitochondrial genes (ND2 + ND3) suggest a sister group relationship of Cypriot and Anatolian water frog lineages, our nuclear data hypothesize a sister group relationship between Cypriot frogs (sp. n.) and Crete frogs (P. cretensis), thus speaking for the same isolation time of both island populations.