Sister Position Research Articles

The complete mitochondrial genome of the leopard shark Triakis semifasciata (Triakidae)

ABSTRACT The leopard shark Triakis semifasciata (family Triakidae) is threatened by habitat loss and targeted by recreational and commercial fishermen in the Eastern Pacific Ocean, from Oregon, USA, to the Gulf of California, Mexico. Despite environmental issues, there are few genetic and genomic resources available for this species. This study assembled the complete mitochondrial genome of T. semifasciata from a skin metagenomic sample and described it in detail. The phylogenetic position of T. semifasciata amongst closely related species was also examined using mitochondrial protein-coding genes (PCGs). The mitochondrial genome of T. semifasciata is 16,613 bp in length and consists of 13 PCGs, two ribosomal RNA (rRNA) genes, and 22 transfer RNA (tRNA) genes. A 30 bp long region was identified as the origin of replication for the light strand (OL) between the trnN and trnC genes, and a 974 bp long putative control region (CR) contains the origin of replication for the heavy strand (OH). The gene order in T. semifasciata is identical to that of cofamilial species. An analysis of Ka/Ks ratios for all PCGs yielded values < 1, indicating that all PCGs experience strong purifying selection. All tRNAs exhibit a canonical ‘cloverleaf’ secondary structure except for tRNA-Ser1 which lacks the stem of the dihydrouridine (DHU) arm and in place possesses a simple loop. A phylomitogenomic maximum likelihood (ML) analysis did not support the monophyly of the family Trakidae and placed T. semifasciata in a sister position to Hemitriakis japanica. However, the aforementioned sister position was poorly supported by ML bootstrap values. This study represents a new genomic resource for this commercially and recreationally important species and confirms that mitochondrial genomes can be assembled from skin metagenomic samples as shown before in an unrelated shark species.

Evolutionary relationships in the African frog family Ptychadenidae, including the first molecular analysis, range extension, and distribution modelling of the monotypic genus Lanzarana

ABSTRACT The family Ptychadenidae contains three ecologically and morphologically distinct genera: the widespread, species-rich and above-ground dwelling Ptychadena, the less widespread and diverse, burrowing Hildebrandtia, and the little-known, burrowing, monotypic Lanzarana, endemic to Somalia. Previous morphological studies placed Lanzarana in a sister position to Hildebrandtia and these two genera as sister to Ptychadena. However, our analyses of mitochondrial and multilocus nuclear DNA data revealed Lanzarana as a sister lineage to Ptychadena (divergence ∼28 Mya), and Hildebrandtia sister to them (∼42 Mya). This suggests that the common ancestor of ptychadenids may have exhibited a burrowing lifestyle similar to Hildebrandtia and Lanzarana, rather than the more active, above-ground lifestyle of Ptychadena. In addition, based upon analysis of available mitochondrial DNA data, we have identified major clades within the genus Ptychadena and propose them as four species groups, with the P. mascareniensis group being the first diverging lineage (∼20 Mya). We also report four new localities of Lanzarana largeni from central Somaliland, south-western Puntland, southern Somalia and south-eastern Ethiopia, extending the presently known range of the species westward. This represents the first official record of L. largeni from Ethiopia. An ecological niche model of L. largeni for present climatic conditions identifies extensive climatically highly suitable areas in central Somalia and south-eastern Ethiopia. A model for future climatic conditions estimates a substantial extension of climatically suitable areas to northern Somalia and north-eastern Kenya, likely due to predicted more humid conditions accompanying the increase of temperatures in the western Indian Ocean.

Challenge accepted: Evolutionary lineages versus taxonomic classification of North American shrub willows (Salix).

The huge diversity of Salix subgenus Chamaetia/Vetrix clade in North America and the lack of phylogenetic resolution within this clade has presented a difficult but fascinating challenge for taxonomists to resolve. Here we tested the existing taxonomic classification with molecular tools. In this study, 132 samples representing 46 species from 22 described sections of shrub willows from the United States and Canada were analyzed and combined with 67 samples from Eurasia. The ploidy levels of the samples were determined using flow cytometry and nQuire. Sequences were produced using a RAD sequencing approach and subsequently analyzed with ipyrad, then used for phylogenetic reconstructions (RAxML, SplitsTree), dating analyses (BEAST, SNAPPER), and character evolution analyses of 14 selected morphological traits (Mesquite). The RAD sequencing approach allowed the production of a well-resolved phylogeny of shrub willows. The resulting tree showed an exclusively North American (NA) clade in sister position to a Eurasian clade, which included some North American endemics. The NA clade began to diversify in the Miocene. Polyploid species appeared in each observed clade. Character evolution analyses revealed that adaptive traits such as habit and adaxial nectaries evolved multiple times independently. The diversity in shrub willows was shaped by an evolutionary radiation in North America. Most species were monophyletic, but the existing sectional classification could not be supported by molecular data. Nevertheless, monophyletic lineages share several morphological characters, which might be useful in the revision of the taxonomic classification of shrub willows.

Genome reduction and horizontal gene transfer in the evolution of Endomicrobia-rise and fall of an intracellular symbiosis with termite gut flagellates.

Bacterial endosymbionts of eukaryotic hosts typically experience massive genome reduction, but the underlying evolutionary processes are often obscured by the lack of free-living relatives. Endomicrobia, a family-level lineage of host-associated bacteria in the phylum Elusimicrobiota that comprises both free-living representatives and endosymbionts of termite gut flagellates, are an excellent model to study evolution of intracellular symbionts. We reconstructed 67 metagenome-assembled genomes (MAGs) of Endomicrobiaceae among more than 1,700 MAGs from the gut microbiota of a wide range of termites. Phylogenomic analysis confirmed a sister position of representatives from termites and ruminants, and allowed to propose eight new genera in the radiation of Endomicrobiaceae. Comparative genome analysis documented progressive genome erosion in the new genus Endomicrobiellum, which comprises all flagellate endosymbionts characterized to date. Massive gene losses were accompanied by the acquisition of new functions by horizontal gene transfer, which led to a shift from a glucose-based energy metabolism to one based on sugar phosphates. The breakdown of glycolysis and many anabolic pathways for amino acids and cofactors in several subgroups was compensated by the independent acquisition of new uptake systems, including an ATP/ADP antiporter, from other gut microbiota. The putative donors are mostly flagellate endosymbionts from other bacterial phyla, including several, hitherto unknown lineages of uncultured Alphaproteobacteria, documenting the importance of horizontal gene transfer in the convergent evolution of these intracellular symbioses. The loss of almost all biosynthetic capacities in some lineages of Endomicrobiellum suggests that their originally mutualistic relationship with flagellates is on its decline.IMPORTANCEUnicellular eukaryotes are frequently colonized by bacterial and archaeal symbionts. A prominent example are the cellulolytic gut flagellates of termites, which harbor diverse but host-specific bacterial symbionts that occur exclusively in termite guts. One of these lineages, the so-called Endomicrobia, comprises both free-living and endosymbiotic representatives, which offers the unique opportunity to study the evolutionary processes underpinning the transition from a free-living to an intracellular lifestyle. Our results revealed a progressive gene loss in energy metabolism and biosynthetic pathways, compensated by the acquisition of new functions via horizontal gene transfer from other gut bacteria, and suggest the eventual breakdown of an initially mutualistic symbiosis. Evidence for convergent evolution of unrelated endosymbionts reflects adaptations to the intracellular environment of termite gut flagellates.

Convergent evolution of the sensory pits in and within flatworms

BackgroundUnlike most free-living platyhelminths, catenulids, the sister group to all remaining flatworms, do not have eyes. Instead, the most prominent sensory structures in their heads are statocysts or sensory pits. The latter, found in the family Stenostomidae, are concave depressions located laterally on the head that represent one of the taxonomically important traits of the family. In the past, the sensory pits of flatworms have been homologized with the cephalic organs of nemerteans, a clade that occupies a sister position to platyhelminths in some recent phylogenies. To test for this homology, we studied morphology and gene expression in the sensory pits of the catenulid Stenostomum brevipharyngium.ResultsWe used confocal and electron microscopy to investigate the detailed morphology of the sensory pits, as well as their formation during regeneration and asexual reproduction. The most prevalent cell type within the organ is epidermally-derived neuron-like cells that have cell bodies embedded deeply in the brain lobes and long neurite-like processes extending to the bottom of the pit. Those elongated processes are adorned with extensive microvillar projections that fill up the cavity of the pit, but cilia are not associated with the sensory pit. We also studied the expression patterns of some of the transcription factors expressed in the nemertean cephalic organs during the development of the pits. Only a single gene, pax4/6, is expressed in both the cerebral organs of nemerteans and sensory pits of S. brevipharyngium, challenging the idea of their deep homology.ConclusionsSince there is no morphological or molecular correspondence between the sensory pits of Stenostomum and the cerebral organs of nemerteans, we reject their homology. Interestingly, the major cell type contributing to the sensory pits of stenostomids shows ultrastructural similarities to the rhabdomeric photoreceptors of other flatworms and expresses ortholog of the gene pax4/6, the pan-bilaterian master regulator of eye development. We suggest that the sensory pits of stenostomids might have evolved from the ancestral rhabdomeric photoreceptors that lost their photosensitivity and evolved secondary function. The mapping of head sensory structures on plathelminth phylogeny indicates that sensory pit-like organs evolved many times independently in flatworms.

Erysiphe elevata causing powdery mildew on Eucalyptus urophylla × E. camaldulensis in China.

Eucalyptus urophylla × E. camaldulensis, named Chiwei eucalypt, is a hybrid species widely used in China. Many of its clones are cultivated for afforestation due to cold tolerance, high yield, high strength and disease resistance. Clone LH1 is planted extensively for its high stability and machinability in South China. In December 2021, severe powdery mildew signs were observed on clone LH1 in Zhanjiang, Guangdong (N28°8'29"; E110°17'5"). Whitish powder principally appeared on both abaxial and adaxial leaf surfaces. All plants were infected within about a week and above 90% leaves were diseased, which resulted in abnormal growth and shrinkage of leaves. Hyphae with single, lobed appressoria were hyaline, septate, branched, 3.3-6.8 µm (ave. 4.9 µm, n>50) wide. Conidiophores with a straight to flexuous foot-cell (14.7-46.1×5.4-9.7 µm, ave. 25.8×7.9 µm, n>30) were erect, hyaline, 2-septate, unbranched, 35.4-81.8 × 5.7-10.7 (ave. 56.7×8.7 µm, n>50). Conidia were solitary, hyaline, cylindrical to elliptical, 27.7-46.6 ×11.2-19.0 (ave. 35.7×16.6 µm, n>50). Chamothecia were not found on infected trees. The further identification was confirmed by partial sequences of internal transcribed spacer (ITS), large submit rRNA gene (LSU), Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), glutamine synthetase (GS), and RNA polymerase II second largest subunit (RPB2) gene. A very small amount of mycelia and spores from voucher specimens CCAS-ASBF-1 and CCAS-ASBF-2 were deposited in the herbarium of Guangdong Ocean University. Specimens were PCR amplified and sequenced with primer pairs ITS1/ITS4 (White et al 1990), LROR/LR7 (Moncalvo et al 1995), PMGAPDH1/PMGAPDH3R, GSPM2/GSPM3R and PmRpb2_4/ PMRpb2_6R (Bradshaw, et al. 2022), respectively. BLASTn results showed that ITS (OP270019 and OQ380937), LSU (OP270018 and OQ380938), GAPDH, GS and RPB2 (OQ414445- OQ414450) were above 99% identical with those of E. elevata on Catalpa bignonioides (ITS: AY587013) (Cook et al 2004), Plumeria rubra (ITS: MH985631) (Yeh et al 2019), Cerbera manghas (ITS: MZ379159; LSU: MZ379160) (Mukhtar et al 2022), and Eucalyptus camaldulensis (LSU: LC177375-6) (Meebon et al. 2017), and above 99% identical with those of Erysiphe vaccinii FH00941201 on Vaccinium corymbosum (ITS: ON073869; RPB2: ON119159; GS: ON075687) and FH00112205 on V. vacillans (ITS: ON073870; GAPDH: ON075646) (Bradshaw et al 2022). This is the first sequence data for non rDNA of E. elevata. In an ITS tree phylogenetic analysis with Maximum likelihood (ML) method showed the fungus clustered in a highly supported clade with E. elevata and E. vaccinii. In a multi-locus tree, E. elevata grouped in a sister position to E. vaccinii FH00941201. Thus, the pathogen was identified as E. elevata based on morphology, DNA BLASTn and phylogenetic analysis (Braun and Cook 2012). Pathogenicity tests were conducted on healthy leaves of 1-year-old potted plants. Ten leaves were cleaned with sterile water, inoculated by gently dusting conidia from single lesion on the naturally infected leaves, and then covered with plastic bags containing wet absorbent cotton. Non-inoculated leaves served as controls. Symptoms developed on all inoculated leaves 3 to 5 days after inoculation, and the fungus was identical to the original fungus on the infected leaves, whereas control plants remained symptomless. This is the first report of powdery mildew caused by E. elevata on Eucalyptus sp. from China. This finding is helpful for land managers to diagnose and control the disease.

The Unicellular, Parasitic Fungi, Sanchytriomycota, Possess a DNA Sequence Possibly Encoding a Long Tubulin Polymerization Promoting Protein (TPPP) but Not a Fungal-Type One.

The unicellular, parasitic fungi of the phylum Sanchytriomycota (sanchytrids) were discovered a few years ago. These unusual chytrid-like fungi parasitize algae. The zoospores of the species of the phylum contain an extremely long kinetosome composed of microtubular singlets or doublets and a non-motile pseudocilium (i.e., a reduced posterior flagellum). Fungi provide an ideal opportunity to test and confirm the correlation between the occurrence of flagellar proteins (the ciliome) and that of the eukaryotic cilium/flagellum since the flagellum occurs in the early-branching phyla and not in terrestrial fungi. Tubulin polymerization promoting protein (TPPP)-like proteins, which contain a p25alpha domain, were also suggested to belong to the ciliome and are present in flagellated fungi. Although sanchytrids have lost many of the flagellar proteins, here it is shown that they possess a DNA sequence possibly encoding long (animal-type) TPPP, but not the fungal-type one characteristic of chytrid fungi. Phylogenetic analysis of p25alpha domains placed sanchytrids into a sister position to Blastocladiomycota, similarly to species phylogeny, with maximal support.

Morphological and Phylogenetic Study of Protococcidians Sheds Light on the Evolution of Epicellular Parasitism in Sporozoa (Apicomplexa), with the Description of Eleutheroschizon planoratum sp. nov

The order Protococcidiida is one of the most poorly studied basal groups of Sporozoa (Apicomplexa sensu stricto). To date, the phylogenetic unity of protococcidians and their relationship with other sporozoans are understudied. Only the protococcidian Eleutheroschizon duboscqi has molecular evidence of a sister position to “true” coccidians (Eimeria, Sarcocystis, Toxoplasma). E. duboscqi is characterized by epicellular development in the so-called parasitophorous sac of the host cell origin. The unusual localization of Eleutheroschizon is comparable to that of Cryptosporidium. We describe a new species of the genus, E. planoratum ex Naineris quadricuspida polychaete from the White Sea, using light and electron microscopy. The morphology of attachment apparatus, phylogenetic analyses of concatenated DNA sequences of the nuclear ribosomal operon (SSU rDNA, ITS1, 5.8S rDNA, ITS2, and LSU rDNA), and compensatory base changes in ITS2 secondary structures of both protococcidians confirm the new species. The resulting phylogenies also confirm that Eleutheroschizon is sister to eimeriid coccidians, while Cryptosporidium tends to be grouped with gregarines. We discuss a new type of endoparasitism among sporozoans—the closed epicellular parasitism that evolved convergently in Eleutheroschizon and Cryptosporidium. The diagnosis of the new species and the emended diagnoses of the species E. duboscqi and the genus Eleutheroschizon are presented.

Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae), a gill parasite of Cirrhinus molitorella (Cyprinidae, Labeoninae) in South China.

Paradiplozoon cirrhini n. sp. (Monogenea, Diplozoidae) is described from the gills of mud carp, Cirrhinus molitorella (Valenciennes, 1844) (Cyprinidae, Labeoninae), collected in Wuzhou, Guangxi Province, and Conghua, Guangdong Province as part of an ongoing survey of the diplozoid fauna in the Pearl River basin of China. The new Paradiplozoon species is distinguished from congeners by the structure of median plate and its outgrowth sclerites. The ITS2 sequences of the new species differ from all known available diplozoid sequences by 22.04%-38.34%. The new species is the first diplozoid species parasitic on Labeoninae in China. Molecular phylogenetic analyses using rRNA ITS2 placed Paradiplozoon cirrhini n. sp. in a sister position to the other Chinese Paradiplozoon, implying that Labeoninae represents an early and potentially ancestral host group for China Paradiplozoon. We also provided ITS2 sequences for four other diplozoids species, namely P. megalobramae Khotenovsky, 1982, P. saurogobionis (Jiang, et al., 1985) Jiang, Wu & Wang, 1989, Sindiplozoon hunanensis Yao & Wang, 1997, and Sindiplozoon sp., and validated their phylogenetic position. The results confirm that all diplozoid species are spilt into two major clades and show monophyly of Sindiplozoon but paraphyly of Paradiplozoon.

Nearctic Dactylogyrus species (Platyhelminthes, Monogenea) parasitizing cypriniform fishes in the context of morphology and phylogeny, with descriptions of seven new species.

Ribosomal DNA sequences are currently available for 32 morphologically recognized species of Dactylogyrus parasitizing Nearctic cypriniforms, but only 16 of them are assigned to nominal species. Herein, morphological data on 28 of the 32 species are provided, together with comments on their phylogenetic relationships in the context of the morphology of taxonomically important structures. Seven new species of Dactylogyrus are described from five species of leuciscids and one species of catostomids, as follows: D. aduncus n. sp. from Campostoma spadiceum, D. cloutmani n. sp. from Luxilus chrysocephalus isolepis, D. cornifrons n. sp. from Cyprinella venusta, D. fimbratus n. sp. from Rhinichthys cataractae, D. mcallisteri n. sp. from Semotilus atromaculatus, and D. chieni n. sp. and D. haneki n. sp. from Hypentelium nigricans. Four previously described species of Dactylogyrus, D. atromaculatus from S. atromaculatus, D. eos from C. neogaeus, D. parvicirrus from Notemigonus crysoleucas and D. perlus from L. c. isolepis, are redescribed and/or figured. As for the remaining 17 species, only the male copulatory organs (MCOs) taken from the respective hologenophores are illustrated. On the basis of phylogenetic analyses, two main clades of Nearctic Dactylogyrus were recognized and supported by the different morphology of the MCO. The first one included 22 strictly Nearctic species sharing the same MCO type with Dactylogyrus spp. parasitizing cyprinids likely of Asian origin. The second clade encompassed Dactylogyrus spp. with diverse MCO morphology and was placed in the sister position to Dactylogyrus spp. parasitizing European leuciscids and North-West African cyprinids.

The oldest genus and species of the fulgoroidian lineage in hemipteran Fulgoroidea from the Lower Cretaceous Laiyang Formation of China

Laiyangella zhangi gen. et sp. nov., the first planthopper from the Lower Cretaceous Laiyang formation, Shandong Province of China, is described. Although exhibiting a cixioid venation that would seem to indicate its placement with or near the Cixiidae, the well-preserved impression of the genitalia of this female specimen is not of the orthopteroid type, but fulgoroid. Laiyangella gen. nov. is thus placed in a sister position to the base of the fulgoroidian lineage in Fulgoroidea; it represents the oldest fossil of this lineage. Laiyangella gen. nov. appears as a key taxon to better understand the early evolution of Fulgoroidea. It warns us the weakness of the taxonomic diagnoses currently used in the identification of major groups that have to be reassessed.

The new chytridiomycete Paradinomyces triforaminorum gen. et sp. nov. co-occurs with other parasitoids during a Kryptoperidinium foliaceum (Dinophyceae) bloom in the Baltic Sea

A new chytrid genus and species was isolated and cultured from samples obtained in the Baltic Sea during a dinoflagellate bloom event. This species is characterized by having a spherical sporangium without papillae and zoospores of 2–3 µm in diameter that are released through 3 discharge pores. Molecular phylogeny based on ribosomal operon showed its sister position to the Dinomyces cluster in Rhizophydiales. Zoospores lack fenestrated cisternae but contain a paracrystalline inclusion, found in a Rhizophydiales representative for the first time. Additionally, the kinetid features are uncommon for Rhizophydiales and only observed in Dinomyces representatives so far. These morphological features and its phylogenetic relationships justify the description of the new genus and speciesParadinomyces triforaminorum gen. nov. sp. nov. belonging to the family Dinomycetaceae. The chytrid was detected during a high-biomass bloom of the dinoflagellate Kryptoperidinium foliaceum. Laboratory experiments suggest this species is highly specific and demonstrate the impact it can have on HAB development. The chytrid co-occurred with three other parasites belonging to Chytridiomycota (Fungi) and Perkinsea (Alveolata), highlighting that parasitic interactions are common during HABs in brackish and marine systems, and these multiple parasites compete for similar hosts.

A Phylogenetic Re-Evaluation of the Stenakrine Opecoelids (Trematoda, Digenea: Opecoeloidea) with Some Taxonomic Novelties

The Opecoeloidea is a large group of xiphidiate digeneans parasitizing marine and freshwater fishes. According to the current taxonomic model, this superfamily contains only one family with numerous subfamilies. This study is devoted to the members of the Stenakrinae. Based on phylogenetic analysis of concatenated sequences of 18S and 28S rRNA genes of stenakrine opecoelids Caudotestis dobrovolski, C. cf. dobrovolski, Hexagrammia zhukovi, Stenakron vetustum, as well as the deep-sea xiphidiate digenean Zdzitowieckitrema incognitum, which so far has had an ambiguous phylogenetic status, we erect a new opecoeloid family, the Zdzitowieckitrematidae fam. nov. The genera Holsworthotrema and Scorpidotrema are removed from the Stenakrinae to the Scorpidotrematinae subfam. nov. within the Opecoelidae. We also remove the Stenakrinae from the Opecoelidae and recognize it as a separate family within the Opecoeloidea. The Stenakridae stat. nov. is a sister taxon to a well-supported Opecoelidae. The Zdzitowieckitrematidae occupies a sister position relative to the stenakrids and the opecoelids taken together. All three families are clearly phylogenetically distinct, however convincing morphological differences are revealed only between the Zdzitowieckitrematidae and the Stenakridae and between the Opecoelidae and the Stenakridae.

The complete chloroplast genome of dalmatian pyrethrum (Tanacetum cinerariifolium (Trevir.) Sch. Bip. (Asteraceae)), the source of the natural insecticide pyrethrin

Tanacetum cinerariifolium is an endemic species of the eastern Adriatic coast that synthesizes the natural insecticide pyrethrin. We have characterized the complete chloroplast genome of the species and analyzed its phylogeny within the Asteraceae family. The complete chloroplast genome of T. cinerariifolium has a size of 150,136 bp, including a large single-copy (LSC) region of 82,717 bp, a small single-copy (SSC) region of 18,411bp, and a pair of inverted repeats (IRs) of 24,504 bp. The chloroplast genome of T. cinerariifolium encodes 108 genes, including 77 protein-coding genes (PCGs), 27 tRNA genes, and 4 rRNA genes. Phylogenetic analyses based on the complete chloroplast genomes placed T. cinerariifolium in a sister position to species of the genera Artemisia and Chrysanthemum.

A cryptic powdery mildew (Golovinomyces hieraciorum sp. nov.) on Hieracium and Pilosella (Compositae)

The Golovinomyces cichoracearum complex is morphologically highly variable and causes powdery mildew on a wide range of Compositae, including Hieracium spp. and Lapsana communis. A comprehensive phylogenetic analysis of Golovinomyces, published in 2013, revealed that G. cichoracearum s. str. was confined to Scorzonera and Tragopogon spp. as hosts (Compositae, tribe Lactuceae, subtribe Scorzonerinae). To clarify the phylogeny and taxonomy of Golovinomyces on Hieracium spp. (Compositae, tribe Lactuceae, subtribe Hieraciinae), which is common in Asia and Europe, morphology and phylogenetic analyses of ITS + 28S rDNA sequences were conducted for 57 powdery mildew specimens of Hieracium spp. and Lapsana communis. Golovinomyces on Hieracium spp. in Asia and Europe, previously referred to as G. cichoracearum, is a previously undescribed, cryptic species, which is described here as Golovinomyces hieraciorum sp. nov.. Since a sequence retrieved from a powdery mildew on Lapsana communis in Switzerland clustered in sister position to the G. hieraciorum clade, numerous German specimens identified as G. cichoracearum were included in the present examinations. All the re-examined collections of “G. cichoracearum” on L. communis were misidentified, and were shown to belong to Neoërysiphe nevoi or Podosphaera ergerontis-canadensis.

Further evidence for the hybrid status of the Brazilian native fern Hypolepis ×paulistana (Dennstaedtiaceae).

Hypolepis ×paulistana was described in 2016 as a putative hybrid, known from a single gathering. The hybrid status of these plants was based solely on the intermediate morphology of the sporophyte, when compared to its presumed parent species. These were thought to be H. stolonifera and H. rugosula, but, H. rigescens (Kunze) T. Moore could not be explicitly ruled out either. In the present work, we tested the hybrid status of Hypolepis ×paulistana adding palynological evidence and by using chloroplast sequences to unambiguously identify the maternal progenitor of the species. We find that sporangia of Hypolepis ×paulistana contain both well-formed spores, as well as spores with morphological and developmental anomalies. The size of the regular spores and the abnormal spores suggest that H. ×paulistana is likely a diploid, and probably infertile hybrid. The ornamentation of the regular spores of H. ×paulistana is similar to that of H. stolonifera. The chloroplast sequences of H. ×paulistana are identical to those of H. stolonifera, as well as their sister position within the global phylogeny of the genus. Thus, we provide new evidence for the hybrid status of H. ×paulistana, and we corroborate the earlier finding that H. stolonifera is the maternal parent.

A new species of Sabuloglossum (Geoglossaceae, Ascomycota) from montane areas

The fungal genus Sabuloglossum (earth tongues) was originally described as a monotypic genus based on the species Geoglossum (Microglossum) arenarium. It typically occupies sandy coastal habitats and forms mycorrhizas with Empetrum nigrum (Ericaceae). While studying material from central Europe (Czech Republic, Slovakia, Austria, and Germany), ascomata morphologically very similar to S. arenarium were observed. They were ecologically limited to mid-altitudinal mountainous habitats and associated with Calluna vulgaris or Vaccinium sp. pl. (Ericaceae). The relationship to S. arenarium and related genera was evaluated using a combination of genetic (nrITS1–5.8S–ITS2 and the 28S nrLSU regions) and comparative morphological analysis. Macro- and micromorphological investigations revealed that mountain populations differ from typical S. arenarium in longer apical cells of paraphyses, more frequently septate ascospores, and the presence of inflated cells in the stipe squamules. In phylogenetic analyses, mountain populations appeared in sister position to S. arenarium in a strongly supported clade. According to these morphological and genetic analyses, in addition to ecological and host-type specificities, the mountain populations from central Europe are proposed here as a distinct species, S. monticola sp. nov. Our study indicates that the speciation of S. monticola may have been driven predominantly by a shift in ecology and mycorrhizal association.

Weak population‐genetic structure of a widely distributed nematode parasite of frogs in the western Palearctic

The genetic structure of parasite populations is affected by various factors such as host–parasite interactions, life-history strategies, and the evolutionary histories of both interacting organisms. In this study, we investigated the distribution, prevalence, and population-genetic structure of Icosiella neglecta (Spirurida, Onchocercidae), a nematode parasite found in Ranidae frogs. We reported this parasite from eight species of water frogs (genus Pelophylax) in Europe, the Middle East, and Central Asia. Its prevalence across investigated localities varied from 3.03% to 95.83%. Based on nucleotide variation in a 28S ribosomal RNA gene, all investigated I. neglecta sequences formed a well-supported phylogenetic clade and were placed in the sister position to the genus Ochoterenella. Despite the substantial genetic variability in a mitochondrial cytochrome c oxidase subunit I (COI) fragment (33 unique haplotypes recognized among 91 sequences), we found only weak population-genetic structure across the study area. There was no obvious association of COI haplotypes with geography, except haplotypes from eastern Turkey, Lebanon, and Iraq which formed a homogeneous, albeit only weakly differentiated group. The historical demographic analyses suggest that the species underwent a sudden and relatively recent population expansion. According to our results, we assume that the population-genetic structure of I. neglecta might be linked to the evolutionary history and dispersal of its dipteran vectors than water frog hosts.

Cortinarius anaunianus (Agaricales, Cortinariaceae): a new species of the Humolentes clade from Italy

Cortinarius anaunianus is described as a new species within the Humolentes clade of the section Calochroi s. lato from alpine silver fir forests (Abies alba) in northeastern Italy (Val di Non, province of Trento). Phylogeny based on the combined nrDNA ITS and LSU, rpb1, rpb2 and the coded gaps of the ITS region supports the independence of C. anaunianus from the closest taxa and its sister position to C. humolens from which it differs by (i) more vivid olivaceous tinges on pileus and lamellae when young, (ii) pileus surface strongly turning ocher-brown with age and exposure (iii) growing in silver fir forests rather than in association with broadleaved trees. Photos of fresh basidiomes in field and a full description of its main macro- and micromorphological features are provided and compared to those of the morphologically most similar species.

Multiple independent losses of cell mouth in phylogenetically distant endosymbiotic lineages of oligohymenophorean ciliates: A lesson from Clausilocola

The cell mouth is a property of the vast majority of free-living and endosymbiotic/epibiotic ciliates of the class Oligohymenophorea. Cytostome, however, naturally absents in the whole endosymbiotic subclass Astomatia and was naturally or experimentally lost in a few members of the subclass Hymenostomatia. This poses a question of how homoplastic might be the lack of oral structures in the oligohymenophorean evolution. To address this question, we used two mitochondrial genes, five nuclear markers, and detailed morphological data from an enigmatic mouthless ciliate, Clausilocola apostropha, which we re-discovered after more than half of a century. According to the present phylogenetic analyses, astomy evolved at least three times independently and in different time frames of the oligohymenophorean phylogeny, ranging from the Paleozoic to the Cenozoic period. Mouthless endosymbionts inhabiting mollusks (represented by Clausilocola), planarians (Haptophrya), and annelids (‘core’ astomes) never clustered together. Haptophrya grouped with the scuticociliate genus Conchophthirus, ‘core’ astomes were placed in a sister position to the scuticociliate orders Philasterida and Pleuronematida, and Clausilocola was robustly nested within the hymenostome family Tetrahymenidae. The tetrahymenid origin of Clausilocola is further corroborated by the existence of mouthless Tetrahymena mutants and the huge phenotypic plasticity in the cytostome size in tetrahymenids.