Intact ribosomal DNA arrays of Potentilla origin detected in Erythronium nucleus suggest recent eudicot-to-monocot horizontal transfer.

During our initial phylogenetic study of the monocot genus Erythronium (Liliaceae), we observed peculiar eudicot-type internal transcribed spacer (ITS) sequences in a dataset derived from genomic DNA of Erythronium dens-canis. This raised the possibility of horizontal transfer of a eudicot alien ribosomal DNA (rDNA) into the Erythronium genome. In this work we aimed to support this hypothesis by carrying out genomic, molecular, and cytogenetic analyses. Genome skimming coupled by PacBio HiFi sequencing of a bacterial artificial chromosome clone derived from flow-sorted nuclei was used to characterise the alien 45S rDNA. Integration of alien rDNA in the recipient genome was further proved by Southern blotting and fluorescence in situ hybridization using specific probes. Alien rDNA, nested among Potentilla species in phylogenetic analysis, likely entered the Erythronium lineage in the common ancestor of E. dens-canis and E. caucasicum. Transferred eudicot-type rDNA preserved its tandemly arrayed feature on a single chromosome and was found to be transcribed in the monocot host, albeit much less efficiently than the native counterpart. This study adds a new example to the rarely documented nuclear-to-nuclear jumps of DNA between eudicots and monocots while holding the scientific community continually in suspense about the mode of DNA transfer.

Integrative taxonomy by molecular species delimitation: multi-locus data corroborate a new species of Balkan Drusinae micro-endemics.

BACKGROUND: Taxonomy offers precise species identification and delimitation and thus provides basic information for biological research, e.g. through assessment of species richness. The importance of molecular taxonomy, i.e., the identification and delimitation of taxa based on molecular markers, has increased in the past decade. Recently developed exploratory tools now allow estimating species-level diversity in multi-locus molecular datasets. RESULTS: Here we use molecular species delimitation tools that either quantify differences in intra- and interspecific variability of loci, or divergence times within and between species, or perform coalescent species tree inference to estimate species-level entities in molecular genetic datasets. We benchmark results from these methods against 14 morphologically readily differentiable species of a well-defined subgroup of the diverse Drusinae subfamily (Trichoptera, Limnephilidae). Using a 3798 bp (6 loci) molecular data set we aim to corroborate a geographically isolated new species by integrating comparative morphological studies and molecular taxonomy. CONCLUSIONS: Our results indicate that only multi-locus species delimitation provides taxonomically relevant information. The data further corroborate the new species Drusus zivici sp. nov. We provide differential diagnostic characters and describe the male, female and larva of this new species and discuss diversity patterns of Drusinae in the Balkans. We further discuss potential and significance of molecular species delimitation. Finally we argue that enhancing collaborative integrative taxonomy will accelerate assessment of global diversity and completion of reference libraries for applied fields, e.g., conservation and biomonitoring.

A hairy case: The evolution of filtering carnivorous Drusinae (Limnephilidae, Trichoptera).

The caddisfly subfamily Drusinae BANKS comprises roughly 100 species inhabiting mountain ranges in Europe, Asia Minor and the Caucasus. A 3-gene phylogeny of the subfamily previously identified three major clades that were corroborated by larval morphology and feeding ecologies: scraping grazers, omnivorous shredders and filtering carnivores. Larvae of filtering carnivores exhibit unique head capsule complexities, unknown from other caddisfly larvae. Here we assess the species-level relationships within filtering carnivores, hypothesizing that head capsule complexity is derived from simple shapes observed in the other feeding groups. We summarize the current systematics and taxonomy of the group, clarify the systematic position of Cryptothrix nebulicola, and present a larval key to filtering carnivorous Drusinae. We infer relationships of all known filtering carnivorous Drusinae and 34 additional Drusinae species using Bayesian species tree analysis and concatenated Bayesian phylogenetic analysis of 3805bp of sequence data from six gene regions (mtCOI5-P, mtCOI3-P, 16S mrDNA, CADH, WG, 28S nrDNA), morphological cladistics from 308 characters, and a total evidence analysis. All analyses support monophyly of the three feeding ecology groups but fail to fully resolve internal relationships. Within filtering carnivores, variation in head setation and frontoclypeus structure may be associated with progressive niche adaptation, with less complex species recovered at a basal position. We propose that diversification of complex setation and frontoclypeus shape represents a recent evolutionary development, hypothetically enforcing speciation and niche specificity within filtering carnivorous Drusinae.

Non-monophyly of Siberian Erythronium (Liliaceae) leads to the recognition of the formerly neglected Erythronium sajanense.

Four Erythronium species have been traditionally recognised within Eurasia based on their disjunct distributions and the slight morphological divergence between them: E. dens-canis, E. caucasicum, E. sibiricum and E. japonicum. The range of E. sibiricum includes adjacent parts of southern Siberia, Kazakhstan, China and Mongolia in the Altai-Sayan mountain region. Despite several recently proposed taxa within the range of E. sibiricum (E. sajanense, E. sibiricum subsp. altaicum, E. sibiricum subsp. sulevii), this species has never been tested for genetic subdivisions. We here used nucleotide sequence variation in one nuclear (internal transcribed spacer) and two plastid (rpl32-trnL, rps15-ycf1) regions to test for genetic divisions within Siberian Erythronium and, in particular, to examine the phylogenetic position of E. sajanense. The plastid phylogeny revealed a basal polytomy among E. japonicum, E. sibiricum populations pertaining to E. sajanense and a third strongly supported lineage that includes E. dens-canis, E. caucasicum and the remainder of E. sibiricum, thus rendering Siberian Erythronium non-monophyletic. The nuclear topology agrees with the plastid one in recovering E. sajanense as a distinct lineage that is weakly supported as sister to E. japonicum. Topological incongruences exist between the plastid and nuclear phylogenies but these do not affect the taxonomic recognition of E. sajanense (endemic to the Western Sayan Mts.). This species is morphologically distinguishable on the basis of its subulate stamen filaments. Whereas nuclear phylogeny failed to resolve any genetic grouping within E. sibiricum s. str., plastid data recovered a deep (possibly phylogeographically meaningful) lineage from samples referred to as E. sibiricum subsp. altaicum.

Larval morphology and phylogenetic position of Drusus balcanicus, Drusus botosaneanui, Drusus serbicus and Drusus tenellus (Trichoptera: Limnephilidae: Drusinae).

In a recent 3-gene phylogeny of the Trichoptera subfamily Drusinae Banks, 1916 molecular data clearly correlated with the morphology and feeding ecology of larvae. The largest of three main groups, the Drusinae grazer clade, exhibits an unusual larval feeding ecology for Limnephilidae, and is the most diverse group. In this paper we describe four previously unknown Drusinae larvae from this clade: Drusus balcanicus Kumanski, 1973 (micro-endemic to Eastern Balkans); Drusus botosaneanui Kumanski, 1968 (Dinaric Western Balkans, Hellenic and Eastern Balkan, Asia Minor), Drusus serbicus Marinkovic-Gospodnetic, 1971a (micro-endemic to Dinaric Western Balkans); and Drusus tenellus (Klapálek, 1898) (Carpathians, Dinaric Eastern Balkans). Characteristically, the larvae of these species develop toothless mandibles typical for the Drusinae grazer clade. Larvae and adults were unambiguously associated by a phylogenetic approach based on two mitochondrial (mtCOI, mtLSU= 16S rDNA) and two nuclear genes (nuWG, nuCAD). In addition, information on the morphology of the larvae is given and the diagnostic features necessary for identification are illustrated.

The larvae of Drusus franzressli Malicky 1974 and Drusus spelaeus (Ulmer 1920) (Trichoptera: Limnephilidae: Drusinae) with notes on ecology and zoogeography.

Water quality monitoring is greatly dependent on identification tools for aquatic and semi-aquatic insects. Species-level identification improves resolution and precision of water quality assessment and requires comprehensive keys. With the aim of increasing the suitability of Drusinae for such applications, this paper gives a description of the hitherto unknown larvae of Drusus franzressli Malicky 1974 and Drusus spelaeus (Ulmer 1920). Information on the morphology of the larvae is given and the most important diagnostic features are illustrated. In the context of already available keys, the larvae of D. franzressli and D. spelaeus key together with Metanoea flavipennis (Pictet 1834), M. rhaetica Schmid 1956, D. improvisus McLachlan 1884, D. nigrescens Meyer-Dür 1875 and Ecclisopteryx malickyi Moretti 1991. These species are easily separated by differences in larval morphology (dorsal outline and sculpturing of pronotum, presence/absence of lateral gills at 2nd and 3rd abdominal segments, start of lateral fringe) and their distribution ranges. Drusus franzressli is endemic to the Hellenic western Balkans whereas D. spelaeus is endemic to the western Alps (Grenoble area). In addition, ecological characteristics are briefly discussed.

Taxonomic notes and key to the West Palearctic Antocha (Antocha) Osten Sacken, 1860 (Diptera, Limoniidae) with description of a new species from Morocco.

Background: The Mediterranean Region of the West Palearctic is one of the most species-rich biomes in the world, hosting a high level of endemism and relict species with important conservation value. The North Africa Atlas Mountains (spanning Morocco, Algeria and Tunisia) belong to a poorly-investigated region of the Mediterranean area, with overlooked aquatic biodiversity; hence, a number of species still remain to be discovered. New information: The subgenus Antocha (Antocha) Osten Sacken, 1860 is recorded for the first time from Africa, with a description of A. (A.) staryi Keresztes & Mabrouki sp. nov. from hilly regions of the Middle Atlas region, Morocco. The unique design of the male terminalia differentiates well the newly-discovered species from its closely-related and range-restricted A. (A.) phoenicia Thomas and Dia, 1982. This is in contrast with the high intraspecific and geographically poorly-defined variability of the widespread A. (A.) vitripennis (Meigen, 1830), for which morphological variability of male genital structures is discussed.Illustrations of male genital parts, distribution data and key to the species from the West Palearctic area are also provided.

Helen's twins in the Balkans: discovery of two new Paraptychoptera Tonnoir, 1919 species closely related to P.helena Peus, 1958, with systematic revision of the "lacustris" group (Diptera, Ptychopteridae).

Ptychopteracastor Keresztes & Kappert, sp. nov. and P.pollux Keresztes & Török, sp. nov. both belong to the subgenus Ptychoptera (Paraptychoptera)Tonnoir (1919) and are described from boggy headwaters in the south Balkan area. These new species are closely related to the range-restricted P.helena Peus, 1958, which is known only from Oiti village, Mount Oeta, Phthioitis region, Greece and, together with P.lacustris, forms a morphologically well-defined unit in the subgenus Paraptychoptera. Based on cladistic analyses of 53 different morphological characters using the male antenna, wing, and genital structures, a general revision of the "lacustris" group is proposed with a dichotomous key of Paraptychoptera species.

Ecology of West Nile Virus in the Danube Delta, Romania: Phylogeography, Xenosurveillance and Mosquito Host-Feeding Patterns.

The ecology of West Nile virus (WNV) in the Danube Delta Biosphere Reserve (Romania) was investigated by combining studies on the virus genetics, phylogeography, xenosurveillance and host-feeding patterns of mosquitoes. Between 2014 and 2016, 655,667 unfed and 3842 engorged mosquito females were collected from four sampling sites. Blood-fed mosquitoes were negative for WNV-RNA, but two pools of unfed Culex pipiens s.l./torrentium collected in 2014 were tested positive. Our results suggest that Romania experienced at least two separate WNV lineage 2 introductions: from Africa into Danube Delta and from Greece into south-eastern Romania in the 1990s and early 2000s, respectively. The genetic diversity of WNV in Romania is primarily shaped by in situ evolution. WNV-specific antibodies were detected for 19 blood-meals from dogs and horses, but not from birds or humans. The hosts of mosquitoes were dominated by non-human mammals (19 species), followed by human and birds (23 species). Thereby, the catholic host-feeding pattern of Culex pipiens s.l./torrentium with a relatively high proportion of birds indicates the species' importance as a potential bridge vector. The low virus prevalence in combination with WNV-specific antibodies indicate continuous, but low activity of WNV in the Danube Delta during the study period.

DNA barcode reference libraries for the monitoring of aquatic biota in Europe: Gap-analysis and recommendations for future work.

Effective identification of species using short DNA fragments (DNA barcoding and DNA metabarcoding) requires reliable sequence reference libraries of known taxa. Both taxonomically comprehensive coverage and content quality are important for sufficient accuracy. For aquatic ecosystems in Europe, reliable barcode reference libraries are particularly important if molecular identification tools are to be implemented in biomonitoring and reports in the context of the EU Water Framework Directive (WFD) and the Marine Strategy Framework Directive (MSFD). We analysed gaps in the two most important reference databases, Barcode of Life Data Systems (BOLD) and NCBI GenBank, with a focus on the taxa most frequently used in WFD and MSFD. Our analyses show that coverage varies strongly among taxonomic groups, and among geographic regions. In general, groups that were actively targeted in barcode projects (e.g. fish, true bugs, caddisflies and vascular plants) are well represented in the barcode libraries, while others have fewer records (e.g. marine molluscs, ascidians, and freshwater diatoms). We also found that species monitored in several countries often are represented by barcodes in reference libraries, while species monitored in a single country frequently lack sequence records. A large proportion of species (up to 50%) in several taxonomic groups are only represented by private data in BOLD. Our results have implications for the future strategy to fill existing gaps in barcode libraries, especially if DNA metabarcoding is to be used in the monitoring of European aquatic biota under the WFD and MSFD. For example, missing species relevant to monitoring in multiple countries should be prioritized for future collaborative programs. We also discuss why a strategy for quality control and quality assurance of barcode reference libraries is needed and recommend future steps to ensure full utilisation of metabarcoding in aquatic biomonitoring.

Description of a new species of Mediotipula from Albania, with consideration of the eastern Mediterranean as a diversity hotspot (Diptera, Tipulidae).

A new species of the TipulasubgenusMediotipula is described from the south-eastern part of Albania, south-eastern Europe. Morphologically, the new species is most similar to T. (M.) stigmatella Schummel, 1833, but differs mainly with respect to males, having a distinctly shaped posterior margin of tergite 9-10, a widened outer gonostylus and a series of details of the inner gonostylus (anterior end of the anterior arm, shape of the posterior arm), as well as having more bulbous and rounded hypogynal valves in the females. Further morphological differences of the male terminalia between allopatric populations of T. (M.) stigmatella in the Carpathians and Balkans, south-eastern Europe, are discussed.

Circulation of Dirofilaria immitis and Dirofilaria repens in the Danube Delta Biosphere Reserve, Romania.

BACKGROUND: Dirofilariosis is an emerging vector-borne parasitic disease in Europe. Monitoring of wild and domestic carnivores demonstrated circulation of Dirofilaria spp. in Romania in the past. For the implementation of control measures, knowledge on the native mosquito community responsible for Dirofilaria spp. transmission is required. METHODS: Mosquito samples originated from a longitudinal study previously performed in the Danube Delta Biosphere Reserve. Mosquito pools were screened for Dirofilaria immitis and Dirofilaria repens. The samples comprised 240,572 female mosquito specimens collected every ten days between April and September in 2014 at four different trapping sites. In addition, blood samples of 36 randomly selected dogs were collected in 2016 in each of the four mosquito sampling sites. A duplex real-time assay was used to detect the presence of one or both Dirofilaria species for each sample. This assay targets the cytochrome c oxidase subunit 1 and the 16S rRNA gene fragments to differentiate both parasites. RESULTS: Dirofilaria immitis and D. repens were detected in mosquito pools at all four trapping sites. In the 2118 mosquito pools tested, D. immitis was identified for eight and D. repens for six of the 14 screened mosquito taxa, with a higher prevalence of D. immitis (4.53% of analysed pools) compared to D. repens (1.09%). Dirofilaria spp. were also identified in dogs from the same sampling sites with a prevalence of 30.56%. For both Dirofilaria species, the highest estimated infection rates (EIRs) were found in Anopheles maculipennis (s.l.) (D. immitis: EIR = 0.206 per 100 specimens, D. repens: EIR = 0.066 per 100 specimens). In contrast, Coquillettidia richiardii and Anopheles hyrcanus as the most frequent taxa had infection rates which were significantly lower: Cq. richiardii (D. immitis: EIR = 0.021; D. repens: EIR = 0.004); An. hyrcanus (D. immitis: EIR = 0.028; D. repens: EIR = 0.006). The number of positive pools per calendar week was positively correlated with the number of screened pools per calendar week, suggesting constant Dirofilaria spp. transmission during the observation period. CONCLUSIONS: This study further confirms significant circulation of Dirofilaria spp. in eastern Europe, with high parasite prevalence in domestic canids and mosquitoes. Therefore, systematic monitoring studies are required to better understand the environmental risk factors for Dirofilaria transmission, allowing the implementation of effective surveillance and control measures.

Taxonomic revision of the Carpathian endemic Pedicia (Crunobia) staryi species-group (Diptera, Pediciidae) based on morphology and molecular data.

Three new species of the genus Pedicia, subgenus Crunobia (Diptera: Pediciidae) belonging to the staryi group are described on the basis of a combination of molecular and morphology datasets, and a key to discriminate between species of the subgenus Crunobia is added. Geographic projection of the identified taxa suggests insular-like distribution and shows the importance of the Carpathians as a genetic center which is home to an exceptionally high aquatic diversity in Europe.

Larval morphology of the Western Balkans endemic caddisflies Drusus krusniki Malicky 1981, D. vernonensis Malicky 1989, and D. vespertinus Marinkovic 1976 (Trichoptera, Limnephilidae, Drusinae).

Drusinae (Trichoptera, Limnephilidae) are highland caddisflies inhabiting high-gradient, turbulent running water and spring habitats. They are disjunctly distributed over the Eurasian mountain ranges, and the majority of species is endemic to particular mountain areas. The most diverse of three main groups of the Drusinae, the grazer clade, consists of species in which larvae feed on epiltihic biofilm and algae. In this paper we describe three previously unknown grazer-clade Drusinae larvae: Drusus krusniki Malicky 1981 (endemic to the Dinaric western Balkans), D. vernonensis Malicky 1989 (endemic to the Hellenic western Balkans), and D. vespertinus Marinkovic 1976 (endemic to the Dinaric western Balkans). The larvae of these species have toothless mandibles typical of the Drusinae grazer clade. Larvae and adults were unambiguously associated using molecular genetic data, i.e., the mitochondrial cytochrome oxidase I gene fragment (mtCOI3-P). Morphological characteristics of the larvae are described and the diagnostic features enabling species-level identification are illustrated. We further discuss the ecology and distribution of three Western Balkan endemic species.

Drusus sharrensis sp. n. (Trichoptera, Limnephilidae), a new species from Sharr National Park in Kosovo, with molecular and ecological notes.

In this paper we describe Drusus sharrensis sp. n., from the Sharr Mountains in Kosovo. Males of the new species are morphologically most similar to Drusus krusniki Malicky, 1981, Drusus kerek Oláh, 2011 and Drusus juliae Oláh, 2011 but differ mainly in exhibiting (1) a differently shaped spinose area on tergite VIII; (2) intermediate appendages anteriorly curved in lateral view with broad tips in dorsal view; (3) inferior appendages with a distinct dorsal protrusion in the proximal half. Females of the new species are morphologically most similar to Drusus krusniki, Drusus kerek, Drusus juliae, and Drusus plicatus Radovanovic, 1942 but mainly differ in (1) segment X that is longer than the supragenital plate with distinctly pointed tips; (2) supragenital plate quadrangular with a distinct round dorsal protrusion; (3) a vulvar scale with a small median lobe. Results of phylogenetic species delimitation support monophyly of Drusus sharrensis sp. n. and recover it as sister to a clade comprising (Drusus pelasgus Oláh, 2010 + Drusus juliae + Drusus arbanios Oláh, 2010 + Drusus plicatus + (Drusus dacothracus Oláh, 2010 + Drusus illyricus Oláh, 2010)). The new species is a micro-endemic of the Sharr Mountains, a main biodiversity hotspot in the Balkan Peninsula. Main threats to the aquatic ecosystems of this part of the Balkan Peninsula are discussed.

Pilot longitudinal mosquito surveillance study in the Danube Delta Biosphere Reserve and the first reports of Anopheles algeriensis Theobald, 1903 and Aedes hungaricus Mihályi, 1955 for Romania.

BACKGROUND: Mosquito-borne viruses (moboviruses) are of growing importance in many countries of Europe. In Romania and especially in the Danube Delta Biosphere Reserve (DDBR), mosquito and mobovirus surveillance are not performed on a regular basis. However, this type of study is crucially needed to evaluate the risk of pathogen transmission, to understand the ecology of emerging moboviruses, or to plan vector control programmes. METHODS: We initiated a longitudinal mosquito surveillance study with carbon dioxide-baited Heavy Duty Encephalitis Vector Survey traps at four sampling sites to analyse the spatio-temporal pattern of the (i) mosquito species composition and diversity, (ii) functional groups of mosquitoes (oviposition sites, overwintering stage, and number of generations), and (iii) the occurrence of potential West Nile virus (WNV) vectors. RESULTS: During 2014, a total of 240,546 female mosquitoes were collected. All species were identified using morphological characteristics and further confirmed by mitochondrial cytochrome c oxidase subunit I (COI) gene analysis of selected specimens. The two most common taxa were Coquilettidia richiardii (40.9 %) and Anopheles hyrcanus (34.1 %), followed by Culex pipiens (sensu lato) (s.l.)/Cx. torrentium (7.7 %), Aedes caspius (5.7 %), Cx. modestus (4.0 %), An. maculipennis (s.l.) (3.9 %), and Ae. vexans (3.0 %). A further seven species were less common in the area studied, including two new records for Romania: An. algeriensis and Ae. hungaricus. Phylogenetic analysis of COI gene demonstrated the evolutionary relatedness of most species with specimens of the same species collected in other European regions, except Ae. detritus and An. algeriensis, which exhibited high genetic diversity. Due to the dominance of Cq. richiardii and An. hyrcanus (75 % of all collected specimens), the overall phenology and temporal pattern of functional groups basically followed the phenology of both species. A huge proportion of the mosquito population in the course of the entire sampling period can be classified as potential WNV vectors. With 40 % of all collected specimens, the most frequent species Cq. richiardii is probably the most important vector of WNV in the DDBR. CONCLUSION: This is the first DNA-barcoding supported analysis of the mosquito fauna in the DDBR. The detection of two new species highlights the lack of knowledge about the mosquito fauna in Romania and in the DDBR in particular. The results provide detailed insights into the spatial-temporal mosquito species composition, which might lead to a better understanding of mobovirus activity in Romania and thus, can be used for the development of vector control programs.

A new species and new records of Molophilus Curtis, 1833 (Diptera: Limoniidae) from the Western Palaearctic Region.

BACKGROUND: Molophilus Curtis, 1833 is the most species-rich Limoniidae genus with a total number of 1006 species and subspecies, from which 97 are recorded in the Western-Palaearctic region so far. However new species are still expected from less investigated regions, like the Balkans or the Eastern Europe. NEW INFORMATION: In the present article, we desrcibe a new limonid crane fly species, Molophilus balcanicus Kolcsár sp. n. from the Central Balkan area (Bulgaria). This new taxa is closely related to M. serpentiger Edwards, 1938 and M. variispinus Starý​, 1971 based on the external male genital structures, but differs from its siblings mostly in the structure of the inner and outer gonostylus. Additionally, a number of species are reported for the first time from various European countries, like M. variispinus Starý, 1971 and M. occultus de Meijere, 1918 from Romania; M. crassipygus de Meijere 1918, M. obsoletus Lackschewitz, 1940 and M. medius de Meijere, 1918 from Greece; M. flavus Goetghebuer, 1920 from Andorra; M. cinereifrons de Meijere, 1920 from Bulgaria and M. corniger Meijere, 1920 from Spain.

New records for the Kosovo caddisfly fauna with the description of a new species, Drusus dardanicus sp. nov. (Trichoptera: Limnephilidae).

The Balkan Peninsula is one of the most important European hotspots of freshwater biodiversity. The region is, however, to a large extent insufficiently investigated. Here we present data on distribution of caddisflies in one particularly understudied area, the Republic of Kosovo. Our data include the first records of Adicella altandroconia Botosaneanu & Novak and Halesus tessellatus (Rambur) for the Kosovo caddisfly fauna, and a new locality for the recently described Ecclisopteryx keroveci Previsic, Graf, & Vitecek. Further, we describe the new caddisfly species Drusus dardanicus sp. nov. from the Kopaonik Mountains. The new species belongs to the D. discophorus Species Group and differs morphologically from its most similar congeners (D. discophorus Radovanovic, D. balcanicus Kumanski, and D. bureschi Kumanski) mainly in exhibiting (1) subtrianglar superior appendages; (2) a narrow, dorsal spinate area of tergite VIII; and (3) evenly rounded tips of intermediate appendages in caudal view. In phylogenetic analysis, D. dardanicus sp. nov. is well delineated and recovered as a sister taxon to D. osogovicus Kumanski, a species recorded from Bulgaria. The recent discovery of a new species and other rare or microendemic species presents important contributions to the knowledge on the rich freshwater biodiversity in Kosovo. These species face increasing anthropogenic pressure and threats to their conservation.

Description of a new species of Wormaldia from Sardinia and a new Drusus species from the Western Balkans (Trichoptera, Philopotamidae, Limnephilidae).

New species are described in the genera Wormaldia (Trichoptera, Philopotamidae) and Drusus (Trichoptera, Limnephilidae, Drusinae). Additionally, the larva of the new species Drususcrenophylax sp. n. is described, and a key provided to larval Drusus species of the bosnicus-group, in which the new species belongs. Observations on the threats to regional freshwater biodiversity and caddisfly endemism are discussed. The new species Wormaldiasarda sp. n. is an endemic of the Tyrrhenian island of Sardinia and differs most conspicuously from its congeners in the shape of segment X, which is trilobate in lateral view. The new species Drususcrenophylax sp. n. is a micro-endemic of the Western Balkans, and increases the endemism rate of Balkan Drusinae to 79% of 39 species. Compared to other Western Balkan Drusus, males of the new species are morphologically most similar to Drususdiscophorus Radovanovic and Drususvernonensis Malicky, but differ in the shape of superior and intermediate appendages. The females of Drususcrenophylax sp. n. are most similar to those of Drususvernonensis, but differ distinctly in the outline of segment X. Larvae of Drususcrenophylax sp. n. exhibit toothless mandibles, indicating a scraping grazing-feeding ecology.

Description of two new filtering carnivore Drusus species (Limnephilidae, Drusinae) from the Western Balkans.

Two new species of the genus Drusus (Trichoptera, Limnephilidae, Drusinae) from the Western Balkans are described. Additionally, observations on the biodiversity and threats to the region's endemic aquatic fauna are discussed. Drususkrpachi sp. n. is a micro-endemic of the Korab Mountains, Macedonia, and Drususmalickyi sp. n. is a micro-endemic of the Prokletije Mountains, Albania. Both new species are most similar to Drususmacedonicus but differ from the latter in the shape of segment IX, the shape of the tips of the intermediate appendages in lateral view, the shape of the inferior appendages, and the form and shape of the parameres. In addition, males of the European species of filtering carnivore Drusinae are diagnosed and illustrated, including Cryptothrixnebulicola McLachlan, Drususchrysotus Rambur, Drususdiscolor Rambur, Drususmacedonicus Schmid, Drususmeridionalis Kumanski, Drususmuelleri McLachlan, Drususromanicus Murgoci and Botosaneanu, and Drusussiveci Malicky. These additions to the Western Balkan fauna demonstrate the significance of this region for European biodiversity and further highlight the importance of faunistic studies in Europe.