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The ability of tissue endosymbionts to degrade and detoxify agrochemicals is increasingly recognized as a mechanism supporting the survival of arthropods in agroecosystems. Therefore, tissue endosymbionts have the potential to drive insecticide resistance in agrobiont spiders, i.e., in major generalist predators and pest control agents within agroecosystems. We hypothesized that the abundance and diversity of the endosymbiotic bacteria of Philodromus cespitum, a philodromid spider dominating central European apple orchards, vary with regard to differences in predation capacity and drive host insecticide resistance. We provisioned P. cespitum with diets of varying protein and lipid content and topically exposed them to field-relevant doses of commonly used insecticides, namely Mospilan (acetamiprid), Movento (spirotetramat), Gondola (sulfoxaflor), Decis (deltamethrin), Coragen (chlorantraniliprole), and Benevia (cyantraniliprole). The analyses were based on 16S rDNA profiles from lysates of the cephalothorax and legs of the tested spiders. The application of Benevia, Mospilan, and Movento was partially lethal. The spiders that were resistant to the treatments with Benevia, Mospilan, or Movento were associated with the increased relative abundance of Mycoplasmatota by more than one order of magnitude. Additionally, the abundance of other bacteria differed in Mospilan-resistant and Mospilan-sensitive individuals. In contrast, the diet regimens were not associated with any major differences in the microbiome diversity nor the diversity of endosymbionts. Philodromus cespitum hosts assemblages with unexpectedly high beta diversity of endosymbionts. The OTU identified as the alpha proteobacterium endosymbiont of Coelostomidia zealandica was an obligate endosymbiont of the analyzed P. cespitum population. Wolbachia, Rickettsia, and Spiroplasma endosymbionts were also highly prevalent and differed in their responses to the applied treatments. In conclusion, differences in the abundance and diversity of endosymbiotic bacteria drove the resistance of the spider host to selected insecticides., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The two stork species that nest in Central Europe, Ciconia ciconia and Ciconia nigra , have been repeatedly shown to host the digenetic trematode Cathaemasia hians (Rudolphi, 1809) in their esophagus and muscular stomach. These host species differ in their habitat and food preferences, and the morphologic characters of C. hians isolates ex Ci. nigra and Ci. ciconia are not identical. These differences led to a previous proposal of two subspecies, C athaemasia hians longivitellata Macko, 1960, and Cathaemasia hians hians Macko, 1960. We hypothesize that the Cathaemasia hians isolates ex Ci. nigra and Ci. ciconia represent two independent species. Therefore, in the present study, we performed the first molecular analyses of C. hians individuals that were consistent with the diagnosis of C. hians hians (ex Ci. nigra ) and C. hians longivitellata (ex Ci. ciconia ). The combined molecular and comparative morphological analyses of the central European Cathaemasia individuals ex Ci. nigra and Ci. ciconia led to the proposal of a split of C. hians into C. hians sensu stricto (formerly C. hians hians ) and C. longivitellata sp. n. (formerly C. hians longivitellata ). Morphological analyses confirmed that the length of the vitellaria is the key identification feature of the two previously mentioned species. Both Cathaemasia spp. substantially differ at the molecular level and have strict host specificity, which might be related to differences in the habitat and food preferences of the two stork species.

Feather bacterial load affects key avian life-history traits such as plumage condition, innate immunity, and reproductive success. Investigating the interplay between life-history traits and feather microbial load is critical for understanding mechanisms of host-microbiome interactions. We hypothesize that spatiotemporal variation associated with migration and molting, body size affecting colonizable body surface area, and preening intensity could shape feather bacterial load. Integrating 16S rDNA-qPCR and flow cytometry, we examined total and viable bacterial loads in the feathers of 316 individuals of 24 Palearctic passerine species. We found that viable bacterial load in feathers was lower in larger species and higher in residents compared to migrants. In contrast, total bacterial load was not explained by any of the life-history traits but varied considerably among species, sampling sites, and years. By pinpointing main drivers of bacterial loads on avian body surfaces, we identify key mechanisms shaping host-microbiome interactions and open alternative research directions., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

This is the first study providing long-term data on the dynamics of bees and wasps and their parasitoids for the evidence-based management of reed beds. Ten years ago, we identified Lipara (Chloropidae) - induced galls on common reed (Phragmites australis, Poaceae) as a critically important resource for specialized bees and wasps (Hymenoptera: Aculeata). We found that they were surprisingly common in relatively newly formed anthropogenic habitats, which elicited questions about the dynamics of bees and wasps and their parasitoids in newly formed reed beds of anthropogenic origin. Therefore, in the winter and spring of 2022/23, we sampled reed galls from the same set of reed beds of anthropogenic and natural origin as those in 2012/13. At 10 sites, the number of sampled galls was similar in both time periods (80-122% of the value from 2012/13); 12 sites experienced a moderate decline (30-79% of the value from 2012/13), and the number of galls at six sampling sites was only 3-23% of their abundance in 2012/13. Spontaneous development was associated with increasing populations. After 10 years of spontaneous development, the populations of bees and wasps (including their parasitoids) bound to Lipara-induced reed galls increased in abundance and species richness or remained at their previous levels, which was dependent on the sampling site. The only identified threat consisted of reclamation efforts. The effects of habitat age were limited, and the assemblages in habitats of near-natural and anthropogenic origin largely overlapped. However, several species were consistently present at lower abundances in the anthropogenic habitats and vice versa. In conclusion, we provided evidence-based support for the establishment of oligotrophic reed beds of anthropogenic origin as management tools providing sustainable habitats for specialized reed gall-associated aculeate hymenopteran inquilines, including the threatened species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Aims: Autoantibodies against hexokinase 1 (HK1) were recently proposed to be associated with diabetic macular edema (DME). We hypothesized that anti-HK1 autoantibodies can be used as DME markers and to predict DME onset., Materials and Methods: Serum from patients with 1) DME, 2) diabetes mellitus (DM), 3) allergies or autoimmunities, and 4) control subjects was tested for anti-HK1 and anti-hexokinase 2 (HK2) autoantibodies by immunoblotting. Patients with DM were prospectively followed for up to nine years, and the association of anti-HK1 antibodies with new-onset DME was evaluated. The vitreous humor was also tested for autoantibodies., Results: Among patients with DME, 32 % were positive for anti-HK1 autoantibodies (42 % of those with underlying type 1 DM and 31 % of those with underlying type 2 DM), and 12 % were positive for anti-HK2 autoantibodies, with only partial overlap of these two groups of patients. Anti-HK1 positive were also 7 % of patients with DM, 6 % of patients with allergies and autoimmunities, and 3 % of control subjects. The latter three groups were anti-HK2 negative. Only one of seven patients with DM who were initially anti-HK1 positive developed DME., Conclusions: Anti-HK1 autoantibodies can be used as DME markers but fail to predict DME onset., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Anti-glutamic acid decarboxylase (GAD) autoantibodies are a hallmark of stiff-person syndrome (SPS) and insulin-dependent diabetes mellitus (IDDM). However, patients with concurrent IDDM and SPS often manifest insulin resistance, and SPS-associated IDDM probably has heterogeneous causes. Some patients manifest IDDM associated only with high titers of anti-GAD65 caused by SPS. By contrast, other patients develop IDDM only after being treated with high-dose corticosteroids or they progress to insulin dependency following their treatment with high-dose corticosteroids. The profile of autoantibodies differs markedly between type 1 diabetes mellitus (T1DM), late-onset diabetes mellitus, and SPS-associated IDDM. Therefore, as with new-onset diabetes after transplantation (NODAT), SPS-associated IDDM should be classified as a specific diabetes entity, the pathophysiology of which requires increased attention., Competing Interests: Declaration of interests No potential conflicts of interest relevant to this review were reported., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Protein tyrosine phosphatase, nonreceptor type 22 (PTPN22), is an archetypal non-HLA autoimmunity gene. It is one of the most prominent genetic contributors to type 1 diabetes mellitus outside the HLA region, and prevalence of its risk variants is subject to enormous geographic variability. Here, we address the genetic background of patients with type 1 diabetes mellitus of Armenian descent. Armenia has a population that has been genetically isolated for 3000 years. We hypothesized that two PTPN22 polymorphisms, rs2476601 and rs1310182, are associated with type 1 diabetes mellitus in persons of Armenian descent. In this association study, we genotyped the allelic frequencies of two risk-associated PTPN22 variants in 96 patients with type 1 diabetes mellitus and 100 controls of Armenian descent. We subsequently examined the associations of PTPN22 variants with the manifestation of type 1 diabetes mellitus and its clinical characteristics. We found that the rs2476601 minor allele (c.1858T) frequency in the control population was very low (q = 0.015), and the trend toward increased frequency of c.1858CT heterozygotes among patients with type 1 diabetes mellitus was not significant (OR 3.34, 95% CI 0.88-12.75; χ2 test p > 0.05). The control population had a high frequency of the minor allele of rs1310182 (q = 0.375). The frequency of c.2054-852TC heterozygotes was significantly higher among the patients with type 1 diabetes mellitus (OR 2.39, 95% CI 1.35-4.24; χ2 test p < 0.001), as was the frequency of the T allele (OR 4.82, 95% CI 2.38-9.76; χ2 test p < 0.001). The rs2476601 c.1858CT genotype and the T allele correlated negatively with the insulin dose needed three to six months after diagnosis. The rs1310182 c.2054-852CC genotype was positively associated with higher HbA1c at diagnosis and 12 months after diagnosis. We have provided the first information on diabetes-associated polymorphisms in PTPN22 in a genetically isolated Armenian population. We found only a limited contribution of the prototypic gain-of-function PTPN22 polymorphism rs2476601. In contrast, we found an unexpectedly close association of type 1 diabetes mellitus with rs1310182., Competing Interests: This study was supported by the Charles University projects COOPERATIO 39 and 260646/SVV/2023. The funders had no involvement in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decision to submit the article for publication. The authors declare no other competing financial interests. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2023 Žak et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The proper course and reproducibility of diagnostic techniques depend on narrowly defined reaction conditions, including the reaction pH. Nevertheless, numerous assays are affected by an inaccurately defined reaction pH. Buffers are sometimes suggested for use outside their useful pH ranges, which complicates the reproducibility of results because the buffering capacity is insufficient to retain the disclosed pH. Here, we focus on the comet assay lysis buffer. Comet assay is broadly used for quantifying DNA breaks in eukaryotic cells. The most widespread comet assay protocols employ lysis of the cells before electrophoresis in a buffer containing Triton X-100, a high concentration of NaCl, sodium sarcosinate, EDTA, and Tris, with some modifications. However, nearly all researchers report that they use Tris buffer at pH 10, and some report the pH of the Tris additive alone. Alternatively, others report the pH of the final lysis buffer. However, the lysis solution used in the comet assay is buffered at a pH outside the useful range of Tris. Tris-based buffers have a useful pH range of 7.0 - 9.0. The buffer composed of 10 mM Tris has pKa 8.10 at 25°C and 8.69 at 4°C. The cell lysis conditions used in nearly all modifications of comet assay protocols remain imprecise and uncritically employed. Despite the pH of the lysis buffer likely has negligible effect on the detection of DNA breaks, precise lysis conditions are highly important for the use of comet assay in the detection of base modifications, which are often unstable and sensitive to pH., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Significance: Cancer-associated tissue-specific lactic acidosis stimulates and mediates tumor invasion and metastasis and is druggable. Rarely, malignancy causes systemic lactic acidosis, the role of which is poorly understood. Recent Advances: The understanding of the role of lactate has shifted dramatically since its discovery. Long recognized as only a waste product, lactate has become known as an alternative metabolism substrate and a secreted nutrient that is exchanged between the tumor and the microenvironment. Tissue-specific lactic acidosis is targeted to improve the host body's anticancer defense and serves as a tool that allows the targeting of anticancer compounds. Systemic lactic acidosis is associated with poor survival. In patients with solid cancer, systemic lactic acidosis is associated with an extremely poor prognosis, as revealed by the analysis of 57 published cases in this study. Although it is considered a pathology worth treating, targeting systemic lactic acidosis in patients with solid cancer is usually inefficient. Critical Issues: Research gaps include simple questions, such as the unknown nuclear pH of the cancer cells and its effects on chemotherapy outcomes, pH sensitivity of glycosylation in cancer cells, in vivo mechanisms of response to acidosis in the absence of lactate, and overinterpretation of in vitro results that were obtained by using cells that were not preadapted to acidic environments. Future Directions: Numerous metabolism-targeting anticancer compounds induce lactatemia, lactic acidosis, or other types of acidosis. Their potential to induce acidic environments is largely overlooked, although the acidosis might contribute to a substantial portion of the observed clinical effects. Antioxid. Redox Signal . 37, 1130-1152.

Arbuscular mycorrhizal fungi (AMF) provide crucial support for the establishment of plants in novel environments. We hypothesized that the OTU/genus richness and diversity of soil- and root-associated AMF associated with alien plant species in their exotic ranges are lower than those in their native ranges. We examined the root-associated and soil-dwelling AMF of 11 invasive plant species in their native and exotic ranges in the United States and Europe by DNA sequencing of the ITS2 locus. Examined root-associated AMF assemblages were simplified, which manifested as the loss of several AMF genera in the exotic ranges of the plants. These fungal assemblages were also characterized by greater dominance and simplification of the fungal assemblages. The dominant fungal genera were present regardless of whether their host plants were in their native or exotic ranges. Interestingly, both the native and invaded soils hosted diverse local AMF assemblages. Therefore, alien plant invasions were not limited to soils with low AMF diversity. Some AMF taxa could be context-dependent passengers rather than drivers of alien plant invasions. Further studies should identify functions of AMF missing or less abundant in roots of plants growing in exotic ranges., (© 2022 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The mygalomorph spiders of the family Atypidae are among the most archaic spiders. The genus Atypus Latreille, 1804 occurs in Eurasia and northern Africa, with a single enigmatic species, Atypus snetsingeri Sarno, 1973, known only from a small area in southeastern Pennsylvania in eastern USA. A close relationship to European species could be assumed based on geographic proximity, but A. snetsingeri more closely resembled Asian species. This study was undertaken to learn more about the genetics of A. snetsingeri, its habitat requirements and natural history. Molecular markers (CO1 sequences) were compared to available data for other atypids and showed that A. snetsingeri is identical with A. karschi Dönitz, 1887 native to East Asia. Natural history parameters in Pennsylvania were also similar in every respect to A. karschi in Japan, therefore, we propose that the spider is an introduced species and the specific epithet snetsingeri is relegated to a junior synonym of A. karschi. Cytogenetic analysis showed an X0 sex chromosome system (42 chromosomes in females, 41 in males) and we also detected nucleolus organizing regions and heterochromatin, the latter for the first time in the Atypoidea. In Pennsylvania the spider is found in a variety of habitats, from forests to suburban shrubbery, where the above-ground webs are usually attached vertically to trees, shrubs, or walls, although other webs are oriented horizontally near the ground. Prey include millipedes, snails, woodlice, carabid beetles and earthworms. Atypus karschi is the first known case of an introduced purse-web spider. It is rarely noticed but well-established within its range in southeastern Pennsylvania., Competing Interests: The authors have declared that no competing interests exist.

As the main component of the Gram-negative bacterial cell wall, lipopolysaccharide (LPS) is well documented as an inducer of inflammation in bovine mammary cells. Lycium barbarum (goji) polysaccharides (LBP) have been used in nonruminants as prebiotics to improve growth performance, immune ability, and antioxidant capacity. We aimed to investigate the underlying effects of LBPs on proinflammatory responses in LPS-stimulated primary bovine mammary epithelial cells (bMECs). Cells were isolated from mammary tissue of three lactating Holstein cows without clinical disease (30.26 ± 3.1 kg/d of milk yield; 175 ± 6 DIM). For the pre-experimental treatment, bMECs were precultured with serum-free medium for 12 h. Treatments were as follows: pretreatment with culture medium devoid of LPS or LBP for 30 h (CON); CON for 24 h followed by challenge with 2 μg/mL LPS for 6 h (LPS); pretreatment with 100 or 300 μg/mL LBP for 24 h followed by LPS challenge (2 μg/mL) for 6 h (LBP(100)+LPS; LBP(300)+LPS). To further determine if the effect of LBP on immuneregulation is peroxisome proliferator-activated receptor-γ (PPARγ) activation dependent, an inhibitor of PPARγ, GW9662, at a concentration of 1 μM was used. Cells treated with LBP at 100, 300, and 500 μg/mL had upregulated protein abundance of PPARγ, while PGC1α had a higher expression only at 300 μg/mL of LBP treatment. Compared with CON, cells pretreated with LBP at 100 and 300 μg/mL had greater protein abundance of SCD1 and SREBP1. 5-Ethynyl-2'-deoxyuridine (EdU) staining and cell wound healing assays showed that the negative effect of LPS alone on cell proliferation was reversed by pretreatment with LBP at both 100 and 300 μg/mL. Upregulation of gene and protein abundance of proinflammatory factors and cytokines (COX-2, NLRP3, TNF-α, IL-1β, and IL-6) induced by LPS stimulation were alleviated by LBP pretreatment at 300 μg/mL (more than 2-fold decrease). Compared with LPS challenge alone, phosphorylation of proteins involved in NF-κB (IκBα and p65) and MAPK (p38, JNK, and ERK) pathways was downregulated following LBP treatment. Additionally, inhibition of PPARγ by GW9662 weakened the protective effect of LBP on LPS-induced protein abundance of phosphorylated p65, COX-2, IL-1β, and TNF-α. These results indicated that the protective effect of LBP on LPS-induced bMECs inflammatory responses is PPARγ activation-dependent. As such, this knowledge might help design strategies for intervening against the detrimental effects of bovine mastitis., Interpretive Summary: Current research examined Lycium barbarum polysaccharides (LBP) for combating LPS-induced inflammatory responses in primary bovine mammary epithelial cells. We uncovered a preventive role of LBP in reducing detrimental effects induced by LPS including inhibition of NF-κB and MAPK along with peroxisome proliferator-activated receptor-γ (PPARγ) activation. The decrease in cell proliferation due to LPS was curtailed by pretreatment with LBP. Moreover, the effect of LBP on regulation of inflammatory responses in bovine mammary epithelial cell was PPARγ dependent. Collectively, data suggest that LBP reverses LPS-induced inflammatory response via MAPK/NF-κB signaling in a PPARγ-activation-dependent manner. Thus, the study provides new insights into therapeutic strategies for combating mastitis using LBP and highlighted the link between PPARγ and regulation of mammary cell inflammation., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Azole fungicides (benzimidazoles, triazoles and imidazoles) are among the most widely used agrochemicals in the world. Unfortunately, azole fungicides are increasingly recognized for playing the role of endocrine disruptors in non-target organisms. Previously, the fecundity of ants with semi-claustral colony founding was found to be severely decreased in response to field-realistic concentrations of azole fungicides. However, during claustral colony founding, the ant queens do not feed and could therefore be protected against effects of agrochemicals applied during the colony founding. In the present study, we hypothesized that claustral colony founding is associated with a lower risk of oral exposure of ant queens to azole fungicides. We exposed queens of a common farmland ant species with claustral colony founding, Lasius niger, to four azole fungicides (epoxiconazole, flusilazole, prochloraz and thiophanate-methyl) that are commonly used in foliar applications and analyzed the differences in fecundity between fungicide-treated groups and the control water-treated group. We found that oral exposure to all four tested formulations of azole fungicides decreased the fecundity of L. niger queens. The decreases in fecundity ranged from 30.5% (epoxiconazole) to 40.3% (prochloraz), although the concentrations of fungicides used were several times lower than the minimum effective concentrations used to eliminate the target fungi by foliar applications of examined fungicides on various crops. Ants with both claustral and semi-claustral colony founding are highly vulnerable to field-realistic concentrations of azole fungicides that are sprayed in foliar applications. Azole fungicides substantially decrease the fitness of ant queens and may explain part of the recently observed decreases in farmland insect abundance and diversity., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Adult trematodes of the genera Conodiplostomum Dubois, 1937 and Neodiplostomum Railliet, 1919 (Trematoda: Diplostomidae) parasitize the intestines of birds of prey, owls and, rarely, passeriform birds. Although the family is taxonomically unsettled, molecular phylogenetics have not been applied to analyze Conodiplostomum and Neodiplostomum and the reference DNA sequences from adult Diplostomidae are scarce and limit studies of their indistinct larval forms. We analyze the Conodiplostomum and Neodiplostomum spp. found during the examination of Czech birds performed from 1962 to 2017, and we provide comparative measurements and host spectra, including prevalence and intensity; we also provide and analyze the sequences of four DNA loci from eight diplostomid species. Molecular phylogenetic analysis suggested that Conodiplostomum spathula (Creplin, 1829), the type species of this genus, is nested in Neodiplostomum. Thus, we suggest the rejection of Conodiplostomum spathula (Creplin, 1829) and the resurrection of Neodiplostomum spathula (Creplin, 1829) La Rue, 1926 and reclassification of all species of Conodiplostomum with the neodiplostomulum type of metacercariae to Neodiplostomum as well. Conodiplostomum canaliculatum (Nicoll, 1914) is reclassified as Neodiplostomum spathulaeforme (Brandes, 1888). The molecular analysis suggested that Conodiplostomum perlatum (Ciurea, 1911), the species with the neascus type of metacercariae, belongs to Crassiphialinae Sudarikov, 1960. We erect the genus Ciureatrema gen. nov. Heneberg & Sitko and reclassify Conodiplostomum perlatum (Ciurea, 1911) as Ciureatrema perlatum (Ciurea, 1911) and establish it as a type species of Ciureatrema gen. nov. Further research should focus on the evolution of the neascus and neodiplostomulum types of metacercariae, as well as the evolution of the genital cone and pseudosuckers in Diplostomidae., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Beauverolides (beauveriolides) are abundant, biologically active cyclodepsipeptides produced by many entomopathogenic fungi, including those that are used as biopesticides. Beauverolides act as cholesterol acyltransferase inhibitors in humans; thus, their mode of action has been the subject of pharmacological and clinical research. The cost-effective analytical methods are needed for fast, routine laboratory analysis of beauverolides. We isolated beauverolides from the fungal strain Isaria fumosorosea PFR 97-Apopka and opened the rings of the isolated beauverolides using a pyridine alkaline medium. We separated fractions of cyclic and linearized beauverolides by thin-layer chromatography, and found the chloroform-acetate (9:1, v/v) and chloroform-acetonitrile-acetate (8:1:1, v/v/v) mobile phases, respectively, to be the most efficient. We examined all the fractions by liquid chromatography-mass spectrometry using ion trap and Orbitrap high resolution mass spectrometry. For rapid screening of the contents of cyclic, and, particularly, linearized beauverolides, we developed a novel analytical method that consisted of using capillary electrophoresis coupled with contactless conductivity detection. Furthermore, we improved the separation of the peptides by applying capillary micellar electrokinetic chromatography with the N-cyclohexyl-2-aminoethanesulfonic acid:SDS:NaOH buffer, pH 9.8 as the background electrolyte. The described novel methods allow fast and cost-effective separation of chemically related groups of beauverolides., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Tarantula urticating setae are modified setae located on the abdomen or pedipalps, which represent an effective defensive mechanism against vertebrate or invertebrate predators and intruders. They are also useful taxonomic tools as morphological characters facilitating the classification of New World theraphosid spiders. In the present study, the morphology of urticating setae was studied on 144 taxa of New World theraphosids, including ontogenetic stages in chosen species, except for species with urticating setae of type VII. The typology of urticating setae was revised, and types I, III and IV were redescribed. The urticating setae in spiders with type I setae, which were originally among type III or were considered setae of intermediate morphology between types I and III, are newly considered to be ontogenetic derivatives of type I and are described as subtypes. Setae of intermediate morphology between that of body setae and type II urticating setae that were found in Iridopelma hirsutum and Antillena rickwesti may provide another evidence that type II urticating setae evolved from body setae. It is supposed that the fusion of barbs with the shaft may lead to the morphology of type II setae. As the type II setae of Aviculariinae evolved independently to the UrS of Theraphosinae and both subfamilies represent two non-sister groups, this should explain the differences in the morphology of body setae in Aviculariinae and Theraphosinae. The terminology of "barbs" and "reversed barbs" was revised and redefined, newly emphasizing the real direction of barbs., Competing Interests: The authors have declared that no competing interests exist.

Deep evolutionary relationships within raptorial niche have recently been challenged. Little is known as to whether birds of the raptorial niche share congruent or host-switching communities of parasites. Here, we analyzed the helminth component communities associated with birds of prey and owls. From 1962 to 2015, we examined 1731 birds of prey and owls in Czechia, and we provide a meta-analysis based on the available literature. Both the analysis of newly examined birds as well as the meta-analysis of previous studies suggested low similarities in the helminth component communities in Strigiformes relative to those in Accipitriformes (Sørensen similarity indices 0.380 in Czechia and 0.324 worldwide) or Falconiformes (0.341 and 0.328), as well as low similarities in the helminth component communities in Falconiformes to those in Accipitriformes (0.366 and 0.413). Globally, 59.6% of helminth species found in Accipitriformes, 39.5% of those in Falconiformes and 38.3% of those in Strigiformes were obligate specialists that were limited to a single examined bird order. Another 11.5%, 12.8% and 8.3% of species had core hosts in only a single order. Only five helminth species infected all three bird orders at a similar prevalence. The differences in prevalence cannot be explained by differences in food composition. We provide detailed information on the prevalence, seasonality, age- and sex-specificity, intensity and lethality of helminth infections. In conclusion, we provide the first systematically collected evidence on the congruence of the helminth distribution and phylogeny of the raptorial niche, which is consistent with its split into Australaves and Afroaves., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Neonicotinoids are very effective in controlling crop pests but have adverse effects on predators and pollinators. Spiders are less sensitive to neonicotinoids compared to insects because of the different structure of their acetylcholine receptors, the binding targets of neonicotinoids. We tested whether short-term exposure to neonicotinoids affected the predation rate in different densities of prey of spiders and led to their paralysis or eventual death. To examine these effects, we topically exposed dominant epigeic, epiphytic and sheet-weaving farmland spiders to four widely used neonicotinoids (imidacloprid, thiamethoxam, acetamiprid and thiacloprid). We applied the neonicotinoids at concentrations recommended by the manufacturers for spray application under field conditions. Short-term exposure to the formulations of all four tested neonicotinoids had adverse effects on the predation rate of spiders, with imidacloprid (Confidor) associated with the most severe effects on the predation rate and exhibiting partial acute lethality after one hour (15-32%). Acetamiprid also displayed strong sublethal effects, particularly when applied dorsally to Philodromus cespitum. Day-long exposure to dorsally applied acetamiprid or thiacloprid led to paralysis or death of multiple Linyphiidae spp., with the effects particularly prominent in males. To conclude, we provided multiple lines of evidence that short-term exposure to neonicotinoids, which were applied at recommended field concentrations, caused severe health effects or death in multiple families of spiders. Even acetamiprid caused strong effects, despite being subject to less strict regulations in the European Union, compared with those for imidacloprid because of claims of its negligible off-target toxicity.

Recent investigation of somatic variations of allosterically regulated proteins in cancer genomes suggested that variations in glucokinase (GCK) might play a role in tumorigenesis. We hypothesized that somatic cancer-associated GCK variations include in part those with activating and/or stabilizing effects. We analyzed the enzyme kinetics and thermostability of recombinant proteins possessing the likely activating variations and the variations present in the connecting loop I and provided the first experimental evidence of the effects of somatic cancer-associated GCK variations. Activating and/or stabilizing variations were common among the analyzed cancer-associated variations, which was in strong contrast to their low frequency among germinal variations. The activating and stabilizing variations displayed focal distribution with respect to the tertiary structure, and were present in the surroundings of the heterotropic allosteric activator site, including but not limited to the connecting loop I and in the active site region subject to extensive rearrangements upon glucose binding. Activating somatic cancer-associated variations induced a reduction of GCK's cooperativity and an increase in the affinity to glucose (a decline in the S 0.5 values). The hotspot-associated variations, which decreased cooperativity, also increased the half-maximal inhibitory concentrations of the competitive GCK inhibitor, N-acetylglucosamine. Concluded, we have provided the first convincing biochemical evidence establishing GCK as a previously unrecognized enzyme that contributes to the reprogramming of energy metabolism in cancer cells. Activating GCK variations substantially increase affinity of GCK to glucose, disrupt the otherwise characteristic sigmoidal response to glucose and/or prolong the enzyme half-life. This, combined, facilitates glucose phosphorylation, thus supporting glycolysis and associated pathways., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Significance: Hexokinases are key enzymes that are responsible for the first reaction of glycolysis, but they also moonlight other cellular processes, including mitochondrial redox signaling regulation. Modulation of hexokinase activity and spatiotemporal location by reactive oxygen and nitrogen species as well as other gasotransmitters serves as the basis for a unique, underexplored method of tight and flexible regulation of these fundamental enzymes. Recent Advances: Redox modifications of thiols serve as a molecular code that enables the precise and complex regulation of hexokinases. Redox regulation of hexokinases is also used by multiple parasites to cause widespread and severe diseases, including malaria, Chagas disease, and sleeping sickness. Redox-active molecules affect each other, and the moonlighting activity of hexokinases provides another feedback loop that affects the cellular redox status and is hijacked in malignantly transformed cells., Critical Issues: Several compounds affect the redox status of hexokinases in vivo. These include the dehydroascorbic acid (oxidized form of vitamin C), pyrrolidinium porrolidine-1-carbodithioate (contraceptive), peroxynitrite (product of ethanol metabolism), alloxan (a glucose analog), and isobenzothiazolinone ebselen. However, very limited information is available regarding which amino acid residues in hexokinases are affected by redox signaling. Except in cases of monogenic diabetes, direct evidence is absent for disease phenotypes that are associated with variations within motifs that are susceptible to redox signaling., Future Directions: Further studies should address the propensity of hexokinases and their disease-associated variants to participate in redox regulation. Robust and straightforward proteomic methods are needed to understand the context and consequences of hexokinase-mediated redox regulation in health and disease.

Background: CD16 was previously suggested to be a new marker of basophils that is subject to downregulation by FcεRI crosslinking. Certain compounds, including supraoptimal concentrations of the PKC inhibitors, bisindolylmaleimides, decouple the release of granules containing CD203c, CD63 and histamine, and may thus help to identify the mechanisms related to the CD16 externalization., Objective: We hypothesized that CD16 is differentially expressed on the surface of basophils in patients with birch pollen or insect venom allergy and is subject to a regulation in response to allergens. We also employed CD203c and CD63 externalization decoupling by bisindolylmaleimides., Methods: We performed a basophil activation test coupled with CD16 and histamine detection using cells isolated from patients with allergy to birch pollen or insect venom and negative controls. We employed two PKC inhibitors, bisindolylmaleimide II and Ro 31-8220 at their supraoptimal concentrations and, after difficulties reproducing previously published data, we analyzed the fluorescence of these inhibitors alone. We identified the CD16 isoforms by sequencing nested RT-PCR amplicons from flow cytometry sorted basophils and by cleaving the CD16b GPI anchor using a phospholipase C., Results: We provide the first evidence that CD16a is expressed as a surface antigen on a small subpopulation of human basophils in patients with respiratory and insect venom allergy, and this antigen shows increased surface expression following allergen challenge or FcεRI crosslinking. We rejected the apparent decoupling of the surface expression of basophil activation markers following the administration of bisindolylmaleimides., Conclusions & Clinical Relevance: The inclusion of αCD16 in negative selection cocktails selects against a subset of basophils that are CD16 + or CD16 dim . Using CD16 dim basophils and unstained leucocytes, we show that previous studies with supraoptimal concentrations of bisindolylmaleimides are likely flawed and are not associated with the differential expression of CD203c and CD63., (© 2018 John Wiley & Sons Ltd.)

Background: Testing for autoantibodies against the zinc transporter ZnT8 (ZnTA) is becoming routine in pediatric diabetes. However, available data are inconclusive when focusing on adult-onset diabetes, including autoimmune diabetes, which does not require insulin at diagnosis (LADA)., Basic Procedures: We examined the ZnTA prevalence and titers and matched them with the clinical phenotype and PTPN22 genotypes of Czech LADA patients who were positive for GADA and/or IA2A and had a fasting C-peptide level >200 pmol/L at diagnosis as well as HNF4A-, GCK- or HNF1A-MODY patients and healthy controls., Main Findings: Most LADA patients were negative for ZnTA, and the sensitivity of the assay was only 18-20% for patients with LADA-like progression to insulinotherapy compared to healthy controls. In LADA patients, there was no association between the ZnTA and PTPN22 risk genotypes. LADA patients positive for ZnTA had a lower BMI than those positive for other autoantibodies alone. Importantly, MODY patients were completely negative for ZnTA, and the levels of ZnTA in MODY patients were similar to those in healthy controls., Conclusions: ZnTA quantification did not improve LADA diagnosis. However, positivity for ZnTA can be used as a negative MODY pre-diagnostic criterion even in the region of Central and East Europe, where other islet cell autoantibodies are common in MODY patients., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Strigeidae Railliet, 1919 are digenean parasites of birds and mammals that are characteristic by their cup-shaped forebody and bilobed holdfast organ. Despite that the family is taxonomically unsettled, particularly due to a very limited number of visible autapomorphic identification features, molecular phylogenetics have never been applied to analyze the relationships among European members of Strigeidae except for the genus Ichthyocotylurus. Here, we analyze the Strigeidae found during the examination of Czech birds performed from 1962 to 2017, and we provide comparative measurements and host spectra, including prevalence and intensity; we also provide and analyze sequences of four DNA loci of 12 of the Strigeidae species. We suggest the reclassification of Parastrigea robusta Szidat, 1928 as Strigea robusta (Szidat, 1928) Heneberg and Sitko, 2018 comb. n. The genera Strigea Abildgaard, 1790 and Parastrigea Szidat, 1928 appear paraphyletic, and morphological diagnostic features of genera within Strigeini Dubois, 1936 are invalid. The mute swan Cygnus olor hosts two Cotylurus spp., Cotylurus syrius Dubois, 1934 and a second species with molecular identification features shared in part with Cotylurus cornutus (Rudolphi, 1808) and Cotylurus gallinulae Lutz, 1928. New host records are provided for seven species. Analyses of non-European genera of the Strigeidae are needed to provide an updated key to Strigeini., (Copyright © 2018 Elsevier B.V. All rights reserved.)

A sensitive capillary electrophoretic method with on-line sample preconcentration by large volume sample stacking has been developed for determination of the anti-microbial agent pentamidine. The separation is performed in a fused silica capillary coated with covalently bound hydroxypropyl cellulose, with an internal diameter of 50 μm and length of 31.5 cm; the background electrolyte was 100 mM acetic acid/Tris at pH 4.7. The stacking is tested using a model sample of 1 μM pentamidine dissolved in 25% infusion solution and 75% acidified acetonitrile. Stacking permits the injection of a sample zone with a length of 95% of the total capillary length to achieve an enhancing factor of 77 compared to low injection into 1.8% of the total capillary length, with simultaneous high separation efficiency of approximately 1 350 000 plates/m. Stacking is based on simultaneous application of a separation field and a hydrodynamic pressure to force the acetonitrile zone out of the capillary. This approach allows the determination of pentamidine in rat blood plasma using only 12.5 μL of plasma treated by the addition of acetonitrile in a ratio of 1:3 v/v. The attained LOD is 0.03 μM and the intra-day repeatability is 0.1% for the migration time and 1.0% for the peak area at the injection 28.3% of capillary length. The performed pharmacokinetic study with ten-second scanning of the blood reveals rapid dynamics of pentamidine in the arterial bloodstream, while the changes are much slower in the venous system., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Trematodes of the genus Philophthalmus Loos, 1899 are the eye parasites of birds and mammals, which use freshwater snails as their first intermediate hosts. Here we examined the presence of philophthalmids in a total of 1515 gulls (589 Larus fuscus and 926 Larus michahellis) admitted between January 2010 and October 2016 for rehabilitation at Olhão (Portugal), by the use of combined morphological and molecular analysis. We recorded the first infected L. fuscus and L. michahellis in July and November 2015, respectively. The philophthalmids were located in the conjunctival sac or under the nictitating membrane. Gulls infected with Philophthalmus lucipetus Rudolphi, 1819 presented no clinical signs, while those infected with Philophthalmus lacrymosus Braun, 1902 presented serious eye damage in the same host species. The prevalence of P. lucipetus reached 3.6% in L. fuscus and 0.8% in L. michahellis; the prevalence of P. lacrymosus reached 0.3% and 0.0%, respectively. The outbreak of P. lucipetus likely started in a narrowly defined area, since the first six cases, found between July and October 2015, originated from a single municipality, and only later more cases started to be retrieved from other municipalities of Portugal. These findings represent the first records of both philophthalmids in the Iberian Peninsula, their first records in L. michahellis and the first record of P. lacrymosus in L. fuscus. Further follow-up of the outbreak and the identification of intermediate hosts are needed., (Copyright © 2017. Published by Elsevier B.V.)

Background: A portion of adults with humoral immune changes have clinical diabetes that is initially not insulin-requiring (latent autoimmune diabetes of the adult, LADA). One of the genes strongly associated with autoimmune diabetes is PTPN22. We hypothesized that the manifestation and clinical features of LADA are linked to functional variants of PTPN22., Methods: We genotyped allelic frequencies of 1 protective and 3 risk-associated PTPN22 variants in 156 Czech LADA patients, 194 type 2 diabetes mellitus patients with LADA-like progression to insulinotherapy and 324 type 1 diabetes mellitus patients, and subsequently examined the associations of PTPN22 variants with the expression of autoantibodies and other clinical features of LADA., Results: We challenged the paradigm that stated that the PTPN22 c.1858T allele serves as a risk allele for LADA, although we confirmed its risk status in the geographically matched T1DM cohort. In contrast, the frequencies of other PTPN22 alleles (c.-1123C, c.788A and c.1970-852C) differed significantly from the healthy controls. We confirmed gender-related differences in the frequency of some PTPN22 polymorphisms (but not c.1858C>T) in LADA. The particular PTPN22 alleles and genotypes were associated with specific clinical features of the examined patients (autoantibodies, HbA1c and age at diagnosis of diabetes)., Conclusions: The variability in PTPN22 haplotypes suggests that the genetic signature of LADA is independent and should not be considered a hybrid form of T1DM and T2DM. Further studies should elucidate the associations with clinical characteristics of the LADA patients and focus on the newly emerging types of diabetes with the disease onset in early to mid-adulthood., (© 2018 S. Karger AG, Basel.)

The Pegosomum Ratz, 1903 are digenean parasites of piscivorous birds. They exhibit few morphological autapomorphies and some of their identification features (number of collar spines) can be altered before or during fixation. Several re-classifications within the genus were suggested, but they have never been supported by molecular analyses. We addressed the synonymization of species within Pegosomum asperum/saginatum complex suggested by Dubinin, Dubinina and Saidov. We analyzed one nuclear (ITS2) and two mitochondrial (CO1, ND1) loci of two central European species of Pegosomum, namely Pegosomum asperum (Wright, 1879) Ratz, 1903 and Pegosomum saginatum (Ratz, 1898) Ratz, 1903. Our combined molecular and comparative morphological analyses confirmed the validity of the two Pegosomum spp. Both species had highly similar morphology and occurred sympatrically in the gall bladder and bile duct of Ardea alba (Linnaeus, 1758). P. saginatum occurred more frequently in hosts infected concurrently with P. asperum. We also provided host-, age- and sex-specific prevalence and intensity of infections, and comparative measurements of the two Pegosomum spp. based on an extensive dataset collected from 1962 to 2016. These species can be morphologically distinguished based on the extent of vitellarium, which reached anteriorly to the pharynx in P. asperum but extended anteriorly to the midline of the esophagus in P. saginatum. The species also differed in the cirrus diameter (P. asperum≥447, P. saginatum≤447)., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Computational methods that allow predicting the effects of nonsynonymous substitutions are an integral part of exome studies. Here, we validated and improved their specificity by performing a comprehensive bioinformatics analysis combined with experimental and clinical data on a model of glucokinase (GCK): 8835 putative variations, including 515 disease-associated variations from 1596 families with diagnoses of monogenic diabetes (GCK-MODY) or persistent hyperinsulinemic hypoglycemia of infancy (PHHI), and 126 variations with available or newly reported (19 variations) data on enzyme kinetics. We also proved that high frequency of disease-associated variations found in patients is closely related to their evolutionary conservation. The default set prediction methods predicted correctly the effects of only a part of the GCK-MODY-associated variations and completely failed to predict the normoglycemic or PHHI-associated variations. Therefore, we calculated evidence-based thresholds that improved significantly the specificity of predictions (≤75%). The combined prediction analysis even allowed to distinguish activating from inactivating variations and identified a group of putatively highly pathogenic variations (EVmutation score <-7.5 and SNAP2 score >70), which were surprisingly underrepresented among MODY patients and thus under negative selection during molecular evolution. We suggested and validated the first robust evidence-based thresholds, which allow improved, highly specific predictions of disease-associated GCK variations.

Cyclocoelidae Stossich, 1902 are medium-sized to large digenean bird parasites. Although these parasites bear few visible autapomorphic signs, and their diagnostic characters are unstable in response to the pressure applied during preparation, the numerous hitherto suggested re-classifications within the family have not been supported by any molecular analysis. We analyse here cyclocoelids found during the extensive examination of central European birds performed from 1962 to 2016, provide comparative measurements, host spectra, prevalence and intensity, and provide and analyse sequences of four DNA loci of five of the cyclocoelid species. Cyclocoleum Brandes, 1892 appears paraphyletic; thus we suggest the re-classification of Cyclocoleum obscurum (Leidy, 1887) as Harrahium obscurum (Leidy, 1887) Sitko and Heneberg comb. n. Molecular phylogenetics questioned also the validity of Cyclocoelinae Stossich, 1902 and Hyptiasminae Dollfus, 1948, which formed a single clade, whereas Allopyge Johnston, 1913, Prohyptiasmus Witenberg, 1923 and Morishitium Witenberg, 1928 formed another clade. Haematotrephinae Dollfus, 1948 are newly characterized as having a pretesticular or intertesticular ovary that forms a triangle with the testes. Analyses of non-European genera of the Cyclocoelidae and an examination of the position of families within Echinostomata La Rue, 1926 are needed.

The ability of aculeate Hymenoptera to utilize wetlands is poorly understood, and descriptions of their nests and developmental stages are largely absent. Here we present results based on our survey of hymenopterans using galls induced by Lipara spp. flies on common reed Phragmites australis in the years 2015-2016. We studied 20,704 galls, of which 9,446 were longitudinally cut and the brood from them reared in the laboratory, while the remaining 11,258 galls reared in rearing bags also in laboratory conditions. We recorded eight species that were previously not known to nest in reed galls: cuckoo wasps Chrysis rutilans and Trichrysis pumilionis, solitary wasps Stenodynerus chevrieranus and Stenodynerus clypeopictus, and bees Pseudoanthidium tenellum, Stelis punctulatissima, Hylaeus communis and Hylaeus confusus. Forty five species of Hymenoptera: Aculeata are known to be associated with reed galls, of which 36 make their nests there, and the other are six parasitoids of the family Chrysididae and three cuckoo bees of the genus Stelis. Of these species, Pemphredon fabricii and in southern Europe also Heriades rubicola are very common in reed galls, followed by Hylaeus pectoralis and two species of the genus Trypoxylon. We also found new host-parasite associations: Chrysis angustula in nests of Pemphredon fabricii, Chrysis rutilans in nests of Stenodynerus clypeopictus, Trichrysis pumilionis in nests of Trypoxylon deceptorium, and Stelis breviuscula in nests of Heriades rubicola. We provide new descriptions of the nests of seven species nesting in reed galls and morphology of mature larvae of eight species nesting in reed galls and two parasitoids and one nest cleptoparasite. The larvae are usually very similar to those of related species but possess characteristics that make them easy to distinguish from related species. Our results show that common reeds are not only expansive and harmful, but very important for many insect species associated with habitats dominated by this plant species., Competing Interests: The authors have declared that no competing interests exist.

The Renicolidae are digenean parasites of piscivorous and molluscivorous birds. Although they exhibit few morphological autapomorphies and are highly variable, the numerous suggested re-classifications within the family have never been supported by any molecular analyses. We address the possible synonymization of species within the Renicola pinguis complex suggested previously by Odening. We provide and analyse sequences of two nuclear (ITS2, 28S rDNA) and two mitochondrial (CO1, ND1) DNA loci of central European species of the Renicolidae, namely Renicola lari, Renicola pinguis and Renicola sternae sp. n., and we also provide first sequences of Renicola sloanei. The combined molecular and comparative morphological analysis confirms the previously questioned validity of the three Renicola spp. of highly similar morphology, which display strict niche separation in terms of host specificity and selectivity. We identify two previously unreported clades within the genus Renicola; however, only one of them is supported by the analysis of adult worms. We also provide comparative measurements of the three examined closely related central European renicolids, and describe the newly proposed tern-specialized species Renicola sternae sp. n., which was previously repeatedly misidentified as Renicola paraquinta. Based on the extensive dataset collected in 1962-2015, we update the host spectrum of Renicolidae parasitizing central European birds (Renicola bretensis, R. lari, Renicola mediovitellata, R. pinguis, Renicola secunda and R. sternae sp. n.) and discuss their host-specific prevalence and intensity of infections.

Objective: AIDS-related mortality has changed dramatically with the onset of highly active antiretroviral therapy (HAART), which has even allowed compensated HIV-infected patients to withdraw from secondary therapy directed against opportunistic pathogens. However, in recently autopsied HIV-infected patients, we observed that associations with a broad spectrum of pathogens remain, although detailed analyses are lacking. Therefore, we focused on the possible frequency and spectrum shifts in pathogens associated with autopsied HIV-infected patients., Design: We hypothesized that the pathogens frequency and spectrum changes found in HIV-infected patients examined postmortem did not recapitulate the changes found previously in HIV-infected patients examined antemortem in both the pre- and post-HAART eras. Because this is the first comprehensive study originating from Central and Eastern Europe, we also compared our data with those obtained in the West and Southwest Europe, USA and Latin America., Methods: We performed autopsies on 124 HIV-infected patients who died from AIDS or other co-morbidities in the Czech Republic between 1985 and 2014. The pathological findings were retrieved from the full postmortem examinations and autopsy records., Results: We collected a total of 502 host-pathogen records covering 82 pathogen species, a spectrum that did not change according to patients' therapy or since the onset of the epidemics, which can probably be explained by the fact that even recently deceased patients were usually decompensated (in 95% of the cases, the last available CD4+ cell count was falling below 200 cells*μl-1) regardless of the treatment they received. The newly identified pathogen taxa in HIV-infected patients included Acinetobacter calcoaceticus, Aerococcus viridans and Escherichia hermannii. We observed a very limited overlap in both the spectra and frequencies of the pathogen species found postmortem in HIV-infected patients in Europe, the USA and Latin America., Conclusions: The shifts documented previously in compensated HIV-infected patients examined antemortem in the post-HAART era are not recapitulated in mostly decompensated HIV-infected patients examined postmortem., Competing Interests: The authors have declared that no competing interests exist.

Among Accipitriformes sensu stricto, only a few species have been reported to form hybrid zones; these include the red kite Milvus milvus and black kite Milvus migrans migrans. M. milvus is endemic to the western Palearctic and has an estimated total population of 20-24,000 breeding pairs. The species was in decline until the 1970s due to persecution and has declined again since the 1990s due to ingestion of rodenticide-treated baits, illegal poisoning and changes in agricultural practices, particularly in its core range. Whereas F1 M. milvus × M. migr. migrans hybrid offspring have been found, F2 and F3 hybrids have only rarely been reported, with low nesting success rates of F1 hybrids and partial hybrid sterility likely playing a role. Here, we analyzed the mitochondrial (CO1 and CytB) and nuclear (Myc) DNA loci of 184 M. milvus, 124 M. migr. migrans and 3 F1 hybrid individuals collected across central Europe. In agreement with previous studies, we found low heterozygosity in M. milvus regardless of locus. We found that populations of both examined species were characterized by a high gene flow within populations, with all of the major haplotypes distributed across the entire examined area. Few haplotypes displayed statistically significant aggregation in one region over another. We did not find mitochondrial DNA of one species in individuals with the plumage of the other species, except in F1 hybrids, which agrees with Haldane´s Rule. It remains to be investigated by genomic methods whether occasional gene flow occurs through the paternal line, as the examined Myc gene displayed only marginal divergence between M. milvus and M. migr. migrans. The central European population of M. milvus is clearly subject to free intraspecific gene flow, which has direct implications when considering the origin of individuals in M. milvus re-introduction programs.

Metorchis spp. are flukes (Platyhelminthes: Digenea) that infect vertebrates, including humans, dogs, cats, poultry and wild game, with cyprinid freshwater fish serving as typical second intermediate hosts. In their definitive hosts, the Metorchis spp. are difficult to identify to species. We provide and analyze sequences of two nuclear (18S rDNA and ITS2) and two mitochondrial (CO1 and ND1) DNA loci of four morphologically identified European species of the Metorchis, namely Metorchis albidus, Metorchis bilis, Metorchis crassiusculus and Metorchis xanthosomus, and of another opisthorchiid, Euamphimerus pancreaticus. DNA analysis suggests that the Metorchis specimens identified morphologically as M. albidus (from Lutra lutra), M. bilis (from Phalacrocorax carbo) and M. crassiusculus (from Aquila heliaca and Buteo rufinus) represent a single species. Thus, M. albidus (Braun, 1893) Loos, 1899 and M. crassiusculus (Rudolphi, 1809) Looss, 1899 are recognized as junior subjective synonyms of M. bilis (Braun, 1790) Odening, 1962. We also provide comparative measurements of the Central European Metorchis spp., and address their tissue specificity and prevalence based on the examination of extensive bird cohort from 1962 to 2015. M. bilis and M. xanthosomus can be morphologically diagnosed by measuring the extent of genitalia relative to body length and by the size ratio of their suckers. They also differ in their core definitive hosts, with ducks (Anas, Aythya) and coots (Fulica) hosting M. xanthosomus, and cormorants (Phalacrocorax), the birds of prey (Buteo, Aquila, etc.), piscivorous mammals (Lutra, Vulpes, Ursus, etc.) and humans hosting M. bilis. Previous reports on the Metorchis spp. contain numerous suspected misidentifications., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Bibliometric indicators increasingly affect careers, funding, and reputation of individuals, their institutions and journals themselves. In contrast to author self-citations, little is known about kinetics of journal self-citations. Here we hypothesized that they may show a generalizable pattern within particular research fields or across multiple fields. We thus analyzed self-cites to 60 journals from three research fields (multidisciplinary sciences, parasitology, and information science). We also hypothesized that the kinetics of journal self-citations and citations received from other journals of the same publisher may differ from foreign citations. We analyzed the journals published the American Association for the Advancement of Science, Nature Publishing Group, and Editura Academiei Române. We found that although the kinetics of journal self-cites is generally faster compared to foreign cites, it shows some field-specific characteristics. Particularly in information science journals, the initial increase in a share of journal self-citations during post-publication year 0 was completely absent. Self-promoting journal self-citations of top-tier journals have rather indirect but negligible direct effects on bibliometric indicators, affecting just the immediacy index and marginally increasing the impact factor itself as long as the affected journals are well established in their fields. In contrast, other forms of journal self-citations and citation stacking may severely affect the impact factor, or other citation-based indices. We identified here a network consisting of three Romanian physics journals Proceedings of the Romanian Academy, Series A, Romanian Journal of Physics, and Romanian Reports in Physics, which displayed low to moderate ratio of journal self-citations, but which multiplied recently their impact factors, and were mutually responsible for 55.9%, 64.7% and 63.3% of citations within the impact factor calculation window to the three journals, respectively. They did not receive nearly any network self-cites prior impact factor calculation window, and their network self-cites decreased sharply after the impact factor calculation window. Journal self-citations and citation stacking requires increased attention and elimination from citation indices.

The recent identification of Pseudamphistomum truncatum, (Rudolphi, 1819) (Trematoda: Opisthorchiidae) and Metorchis bilis (Braun, 1790) Odening, 1962 (synonymous with Metorchis albidus (Braun, 1893) Loos, 1899 and Metorchis crassiusculus (Rudolphi, 1809) Looss, 1899 (Trematoda: Opisthorchiidae)) in otters from Britain caused concern because of associated biliary damage, coupled with speculation over their alien status. Here, we investigate the presence, intensity and phylogeny of these trematodes in mustelids (principally otters) across Europe (Czech Republic, Denmark, France, Germany, Norway, Poland and Sweden and Britain). The trematodes were identified to species using the internal transcribed spacer II (ITS2) locus. Both parasites were found across Europe but at unequal frequency. In the German state of Saxony, eight out of eleven (73%) otters examined were infected with P. truncatum whilst this parasite was not found in either mink from Scotland (n=40) or otters from Norway (n=21). Differences in the phylogenies between the two species suggest divergent demographic histories possibly reflecting contrasting host diet or competitive exclusion, with M. bilis exhibiting greater mitochondrial diversity than P. truncatum. Shared haplotypes within the ranges of both parasite species probably reflect relatively unrestricted movements (both natural and anthropogenic) of intermediate and definitive hosts across Europe., (Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.)

The Brachylaimoidea are digenean parasites of vertebrates, including humans, domestic animals, poultry and wild game. Numerous Brachylaimoidea, particularly adults of Brachylaima and Leucochloridium, are difficult to identify to species. We provide and analyse sequences of two nuclear (18S rDNA, ITS2) and two mitochondrial (CO1, ND1) DNA loci of central European species of the Brachylaimoidea, namely Leucochloridium holostomum, Leucochloridium paradoxum, Leucochloridium perturbatum, Leucochloridium subtilis, Leucochloridium vogtianum, Urotocus rossitensis, Urogonimus macrostomus, Michajlovia migrata, Leucochloridiomorpha lutea, Brachylaima arcuatus, Brachylaima fuscata and Brachylaima mesostoma. We identified three clades in the genus Leucochloridium, which do not correspond to the previously suggested subgenera Neoleucochloridium, Papilloleucochloridium and Leucochloridium. We reject classification of Urotocus and Urogonimus in Leucochloridiinae, and, instead, re-establish the subfamilies Urotocinae and Urogoniminae. We synonymize the genus Renylaima with the genus Brachylaima. We reject M. migrata as a member of Leucochloridiinae sensu stricto or Brachylaimidae suggested by some previous authors. We found that the previously sequenced Glaphyrostomum sp. does not cluster with any hitherto sequenced Brachylaimidae. We also provide comparative measurements of the examined central European Brachylaimoidea, address the the specificity of their localization in the host and discuss their host-specific prevalence and intensity of infections based on the extensive dataset of birds examined in 1962-2015.

Monotypic stands of common reed and the reed-gall-associated insect assemblages are distributed worldwide. However, fungi associated with these assemblages have not been characterized in detail. Here we examined 5200 individuals (12 species) of immature aculeate hymenopterans or their parasitoids collected at 34 sampling sites in Central Europe. We noticed fungal outgrowth on exoskeletons of 83 (1.60%) larvae and pupae. The most common host was eudominant Pemphredon fabricii. However, the less abundant aculeate hymenopteran reed gall inquilines were infected at higher prevalence, these included Trypoxylon deceptorium, Trypoxylon minus, Hoplitis leucomelana and Hylaeus moricei (all considered new host records). We identified three fungal species, Penicillium buchwaldii (72% of cases), Aspergillus pseudoglaucus (22%) and Penicillium quebecense (6%). When multibrooded nests were affected, only a part of individuals was infected in 62% of cases. The sampling site-specific infection rate reached up to 13%, thus fungal infections should be considered an important variable driving the abundance of gall inquilines. Infections of generalist host species were more frequent than those of reed gall specialists, suggesting that suboptimal conditions decreased the immunocompetence of non-specialized species, which only occasionally nest in reed galls and feed in reed beds., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Growth stimuli in cancer growth resemble those exhibited in wound healing. However, the process of nemosis is absent in cancer-associated fibroblasts (CAFs), which remain constitutively active. CAFs are present in almost all solid tumors but are most abundant in breast, prostate and pancreatic cancers. TGF-β1, TGF-β2, PDGF, IL-6, bFGF, reactive oxide species and protein kinase C are considered the key players in tumor-induced transdifferentiation of surrounding fibroblasts. Full-extent transdifferentiation was obtained only when the medium contained TGF-β1 or TGF-β2 (with or without other factors), whereas PDGF, bFGF or IL-6 (each alone) induced only partial transdifferentiation. Recent evidence suggests that the fibroblasts associated with primary cancers differ from those associated with metastases. The metastases-associated fibroblasts are converted by a metastasis-specific spectrum of factors. A large portion of paracrine tumor signaling is mediated by cancer cell-derived vesicles termed exosomes and microvesicles. The cancer cell-derived exosomes contain abundant and diverse proteomes and a number of signaling factors (TGF-ß1, TGF-ß2, IL-6, MMP2 and MMP9), particularly under hypoxic conditions. In contrast to the traditional view, the clonal expansion and selection of neoplastic cells should not be viewed outside the host body context. It is vital for a neoplastic cell to achieve the ability to re-program host body cells into CAFs and by this influence to modulate its microenvironment and receive positive feedback for growth and drug resistance. Neoplastic cells, which fail to develop such capacity, do not pass critical barriers in tumorigenesis and remain dormant and benign., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Cancer arises in the context of an in vivo tumor microenvironment. This microenvironment is both a cause and consequence of tumorigenesis. Tumor and host cells co-evolve dynamically through indirect and direct cellular interactions, eliciting multiscale effects on many biological programs, including cellular proliferation, growth, and metabolism, as well as angiogenesis and hypoxia and innate and adaptive immunity. Here we highlight specific biological processes that could be exploited as targets for the prevention and therapy of cancer. Specifically, we describe how inhibition of targets such as cholesterol synthesis and metabolites, reactive oxygen species and hypoxia, macrophage activation and conversion, indoleamine 2,3-dioxygenase regulation of dendritic cells, vascular endothelial growth factor regulation of angiogenesis, fibrosis inhibition, endoglin, and Janus kinase signaling emerge as examples of important potential nexuses in the regulation of tumorigenesis and the tumor microenvironment that can be targeted. We have also identified therapeutic agents as approaches, in particular natural products such as berberine, resveratrol, onionin A, epigallocatechin gallate, genistein, curcumin, naringenin, desoxyrhapontigenin, piperine, and zerumbone, that may warrant further investigation to target the tumor microenvironment for the treatment and/or prevention of cancer., (Copyright © 2015 Elsevier Ltd. All rights reserved.)