Surprisingly high genetic divergence of the mitochondrial DNA barcode fragment (COI) within Central European woodlice species (Crustacea, Isopoda, Oniscidea).

DNA barcoding has become the most popular approach for species identification in recent years. As part of the German Barcode of Life project, the first DNA barcode library for terrestrial and freshwater isopods from Germany is presented. The analyzed barcode library included 38 terrestrial (78% of the documented species of Germany) and five freshwater (63%) species. A total of 513 new barcodes was generated and 518 DNA barcodes were analyzed. This analysis revealed surprisingly high intraspecific genetic distances for numerous species, with a maximum of 29.4% for Platyarthrushoffmannseggii Brandt, 1833. The number of BINs per species ranged from one (32 species, 68%) to a maximum of six for Trachelipusrathkii (Brandt, 1833). In spite of such high intraspecific variability, interspecific distances with values between 12.6% and 29.8% allowed a valid species assignment of all analyzed isopods. The observed high intraspecific distances presumably result from phylogeographic events, Wolbachia infections, atypical mitochondrial DNAs, heteroplasmy, or various combinations of these factors. Our study represents the first step in generating an extensive reference library of DNA barcodes for terrestrial and freshwater isopods for future molecular biodiversity assessment studies.

Maurieseuma, a new genus to accommodate Hylebainosoma nontronense Mauriès & Kime, 1999, and the revalidation of the genus Romanosoma Mauriès, 2015 (Diplopoda: Chordeumatida: Haaseidae).

A new genus, Maurieseuma gen. nov., is established to accommodate the species Hylebainosoma nontronense Mauriès & Kime, 1999. As result, the following new combination is proposed: Maurieseuma nontronense (Mauriès & Kime, 1999) comb. nov. The new genus is diagnosed and briefly described with some new details missing in the previous two descriptions of the species accommodated in it. In addition, we consider Romanosoma Mauriès, 2015 as a valid genus, leaving Hylebainosoma Verhoeff, 1899 with two species. The similarities and differences between the genera Maurieseuma gen. nov. and Romanosoma, as well as with the remaining three genera of Haaseidae are briefly discussed. An identification key is given to all five genera of the family Haaseidae.

The dark side of pseudoscorpion diversity: The German Barcode of Life campaign reveals high levels of undocumented diversity in European false scorpions.

DNA barcoding is particularly useful for identification and species delimitation in taxa with conserved morphology. Pseudoscorpions are arachnids with high prevalence of morphological crypsis. Here, we present the first comprehensive DNA barcode library for Central European Pseudoscorpiones, covering 70% of the German pseudoscorpion fauna (35 out of 50 species). For 21 species, we provide the first publicly available COI barcodes, including the rare Anthrenochernes stellae Lohmander, a species protected by the FFH Habitats Directive. The pattern of intraspecific COI variation and interspecific COI variation (i.e., presence of a barcode gap) generally allows application of the DNA barcoding approach, but revision of current taxonomic designations is indicated in several taxa. Sequences of 36 morphospecies were assigned to 74 BINs (barcode index numbers). This unusually high number of intraspecific BINs can be explained by the presence of overlooked cryptic species and by the accelerated substitution rate in the mitochondrial genome of pseudoscorpions, as known from previous studies. Therefore, BINs may not be an appropriate proxy for species numbers in pseudoscorpions, while partitions built with the ASAP algorithm (Assemble Species by Automatic Partitioning) correspond well with putative species. ASAP delineated 51 taxonomic units from our data, an increase of 42% compared with the present taxonomy. The Neobisium carcionoides complex, currently considered a polymorphic species, represents an outstanding example of cryptic diversity: 154 sequences from our dataset were allocated to 23 BINs and 12 ASAP units.

The subgenus Monotarsobius in the Iberian Peninsula with a description of a new pseudo-cryptic species from Northern Spain revealed by an integrative revision of Lithobius crassipes L. Koch, 1862 (Chilopoda, Lithobiomorpha, Lithobiidae).

The widespread European centipede species Lithobius (Monotarsobius) crassipes L. Koch, 1862 was revised using an integrative approach incorporating sequence data and morphology. The partial mitochondrial cytochrome c oxidase subunit I (COI) barcoding gene was amplified and sequenced for 21 individuals from northern Spain, France and Germany as well as for individuals of three other species of the subgenus Monotarsobius Verhoeff, 1905. The dataset was used for molecular phylogenetic analysis and genetic distance determination. In addition, Monotarsobius specimens from more than 100 localities in northern Spain, France, and Germany were morphologically investigated. Both morphological and molecular data indicate that specimens from the Navarre and Gipuzkoa provinces, northern Spain, represent a distinct pseudo-cryptic species, only differing in some minor characters from L. crassipes. The new species L. (Monotarsobius) crassipesoides sp. n. is described and compared to L. crassipes in detail using morphology and morphometric statistics for body, head, and antennae length, number of ocelli and coxal pores, as well as the starting leg for legpair spines Vmt and DaP. The Iberian and European records of L. crassipes are discussed. The subspecies L. crassipes morenoi Garcia Ruiz, 2014 from Southern Spain is elevated to species as L. morenoistat. n. A checklist, distribution map and key to all five species of Monotarsobius of the Iberian Peninsula are presented.

Hungarosoma bokori Verhoeff, 1928 (Diplopoda: Chordeumatida): new insights into its taxonomy, systematics, molecular genetics, biogeography and ecology.

Hungarosoma bokori Verhoeff, 1928 is a millipede species which was originally classified solely on the basis of a female specimen. Subsequently, a long history of field searching for and surmising about the systematic position of this small, enigmatic species followed. In April 2013, 85 years after its first description, a series of nine specimens were sampled in the type locality, the Abaliget Cave, in southern Hungary. An adult male was collected for the first time, along with females and juveniles. Descriptions of the gonopods and the female vulvae, both important for considerations of the systematic position of the species, are presented for the first time. Revision and re-designation of the type material was made.The cryptic life of the species is connected with its activity in winter, and its known fragmented distribution corresponds with its presence in undisturbed microhabitats having a specific microclimate, often in the soil at cave entrances.Molecular methods showed a positive detection of the intracellular prokaryotic parasite Wolbachia in H. bokori, reflecting its highly probable parthenogenetic character in the main part of its known area of occurrence. This is the first demonstration of Wolbachia in a millipede.The legitimacy of the family Hungarosomatidae Ceuca, 1974, as a separate taxon was analysed using morphological and molecular approaches. Results of both methods confirmed the existence of a distinct phyletic line. DNA barcoding has shown its closest position to Attemsiidae Verhoeff, 1899, or Neoatractosomatidae Verhoeff, 1901. Based on records from Austria, the Czech Republic, Hungary and Slovakia, the residual circum-pannonic distribution that the whole genus (family) probably represents is proposed.

Towards a DNA Barcode Reference Database for Spiders and Harvestmen of Germany.

As part of the German Barcode of Life campaign, over 3500 arachnid specimens have been collected and analyzed: ca. 3300 Araneae and 200 Opiliones, belonging to almost 600 species (median: 4 individuals/species). This covers about 60% of the spider fauna and more than 70% of the harvestmen fauna recorded for Germany. The overwhelming majority of species could be readily identified through DNA barcoding: median distances between closest species lay around 9% in spiders and 13% in harvestmen, while in 95% of the cases, intraspecific distances were below 2.5% and 8% respectively, with intraspecific medians at 0.3% and 0.2%. However, almost 20 spider species, most notably in the family Lycosidae, could not be separated through DNA barcoding (although many of them present discrete morphological differences). Conspicuously high interspecific distances were found in even more cases, hinting at cryptic species in some instances. A new program is presented: DiStats calculates the statistics needed to meet DNA barcode release criteria. Furthermore, new generic COI primers useful for a wide range of taxa (also other than arachnids) are introduced.

Barcoding of Central European Cryptops centipedes reveals large interspecific distances with ghost lineages and new species records from Germany and Austria (Chilopoda, Scolopendromorpha).

In order to evaluate the diversity of Central European Myriapoda species in the course of the German Barcode of Life project, 61 cytochrome c oxidase I sequences of the genus Cryptops Leach, 1815, a centipede genus of the order Scolopendromorpha, were successfully sequenced and analyzed. One sequence of Scolopendra cingulata Latreille, 1829 and one of Theatops erythrocephalus Koch, 1847 were utilized as outgroups. Instead of the expected three species (Cryptops parisi Brolemann, 1920; Cryptops anomalans Newport, 1844; Cryptops hortensis (Donovan, 1810)), analyzed samples included eight to ten species. Of the eight clearly distinguishable morphospecies of Cryptops, five (Cryptops parisi; Cryptops croaticus Verhoeff, 1931; Cryptops anomalans; Cryptops umbricus Verhoeff, 1931; Cryptops hortensis) could be tentatively determined to species level, while a further three remain undetermined (one each from Germany, Austria and Croatia, and Slovenia). Cryptops croaticus is recorded for the first time from Austria. A single specimen (previously suspected as being Cryptops anomalans), was redetermined as Cryptops umbricus Verhoeff, 1931, a first record for Germany. All analyzed Cryptops species are monophyletic and show large genetic distances from one another (p-distances of 13.7-22.2%). Clear barcoding gaps are present in lineages represented by >10 specimens, highlighting the usefulness of the barcoding method for evaluating species diversity in centipedes. German specimens formally assigned to Cryptops parisi are divided into three clades differing by 8.4-11.3% from one another; their intra-lineage genetic distance is much lower at 0-1.1%. The three clades are geographically separate, indicating that they might represent distinct species. Aside from Cryptops parisi, intraspecific distances of Cryptops spp. in Central Europe are low (<3.3%).

First results of the German Barcode of Life (GBOL) - Myriapoda project: Cryptic lineages in German Stenotaenialinearis (Koch, 1835) (Chilopoda, Geophilomorpha).

As part of the German Barcode of Life (GBOL) Myriapoda program, which aims to sequence the COI barcoding fragment for 2000 specimens of Germany's 200 myriapod species in the near future, 44 sequences of the centipede order Geophilomorpha are analyzed. The analyses are limited to the genera Geophilus Leach, 1814 and Stenotaenia Koch, 1847 and include a total of six species. A special focus is Stenotaenia, of which 19 specimens from southern, western and eastern Germany could be successfully sequenced. The Stenotaenia data shows the presence of three to four vastly different (13.7-16.7% p-distance) lineages of the genus in Germany. At least two of the three lineages show a wide distribution across Germany, only the lineage including topotypes of Stenotaenialinearis shows a more restricted distribution in southern Germany. In a maximum likelihood phylogenetic analysis the Italian species Stenotaenia 'sorrentina' (Attems, 1903) groups with the different German Stenotaenialinearis clades. The strongly different Stenotaenialinearis lineages within Germany, independent of geography, are a strong hint for the presence of additional, cryptic Stenotaenia species in Germany.

Barcoding Fauna Bavarica: Myriapoda - a contribution to DNA sequence-based identifications of centipedes and millipedes (Chilopoda, Diplopoda).

We give a first account of our ongoing barcoding activities on Bavarian myriapods in the framework of the Barcoding Fauna Bavarica project and IBOL, the International Barcode of Life. Having analyzed 126 taxa (including 122 species) belonging to all major German chilopod and diplopod lineages, often using four or more specimens each, at the moment our species stock includes 82% of the diplopods and 65% of the chilopods found in Bavaria, southern Germany. The partial COI sequences allow correct identification of more than 95% of the current set of Bavarian species. Moreover, most of the myriapod orders and families appear as distinct clades in neighbour-joining trees, although the phylogenetic relationships between them are not always depicted correctly. We give examples of (1) high interspecific sequence variability among closely related species; (2) low interspecific variability in some chordeumatidan genera, indicating that recent speciations cannot be resolved with certainty using COI DNA barcodes; (3) high intraspecific variation in some genera, suggesting the existence of cryptic lineages; and (4) the possible polyphyly of some taxa, i.e. the chordeumatidan genus Ochogona. This shows that, in addition to species identification, our data may be useful in various ways in the context of species delimitations, taxonomic revisions and analyses of ongoing speciation processes.