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Transcription-replication collisions (TRCs) are crucial determinants of genome instability. R-loops were linked to head-on TRCs and proposed to obstruct replication fork progression. The underlying mechanisms, however, remained elusive due to the lack of direct visualization and of non-ambiguous research tools. Here, we ascertained the stability of estrogen-induced R-loops on the human genome, visualized them directly by electron microscopy (EM), and measured R-loop frequency and size at the single-molecule level. Combining EM and immuno-labeling on locus-specific head-on TRCs in bacteria, we observed the frequent accumulation of DNA:RNA hybrids behind replication forks. These post-replicative structures are linked to fork slowing and reversal across conflict regions and are distinct from physiological DNA:RNA hybrids at Okazaki fragments. Comet assays on nascent DNA revealed a marked delay in nascent DNA maturation in multiple conditions previously linked to R-loop accumulation. Altogether, our findings suggest that TRC-associated replication interference entails transactions that follow initial R-loop bypass by the replication fork.

Tetra-ortho-fluoro-azobenzenes are a class of photoswitches useful for the construction of visible-light-controlled molecular systems. They can be used to achieve spatio-temporal control over the properties of a chosen bioactive molecule. However, the introduction of different substituents to the tetra-fluoro-azobenzene core can significantly affect the photochemical properties of the switch and compromise biocompatibility. Herein, we explored the effect of useful substituents, such as functionalization points, attachment handles, and water-solubilizing groups, on the photochemical properties of this photochromic system. In general, all the tested fluorinated azobenzenes exhibited favorable photochemical properties, such as high photostationary state distribution and long half-lives, both in organic solvents and in water. One of the azobenzene building blocks was functionalized with a trehalose group to enable the uptake of the photoswitch into mycobacteria. Following metabolic uptake and incorporation of the trehalose-based azobenzene in the mycobacterial cell wall, we demonstrated photoswitching of the azobenzene in the isolated total lipid extract.

Cluster of differentiation (CD155), formerly identified as poliovirus receptor (PVR) and later as immunoglobulin molecule, is involved in cell adhesion, proliferation, invasion and migration. It is a surface protein expressed mostly on normal and transformed malignant cells. The expression of the receptor varies based on the origin of tissue. The expression of the protein is determined by factors involved in the sonic hedgehog pathway, Ras-MEK-ERK pathway and during stressful conditions like DNA damage response. The protein uses an alternate splicing mechanism, producing four isoforms, two being soluble (CD155β and CD155γ) and two being transmembrane protein (CD155α and CD155δ). Apart from being a viral receptor, researchers have identified CD155 to play important roles in cancer research and the cell signaling field. The receptor is recognized as a biomarker for identifying cancerous tissue. The receptor interacts with molecules involved in the cells’ defense mechanism. The immunesurveillance role of CD155 is being deciphered to understand the mechanistic approach it utilizes as an onco-immunologic molecule. CD155 is a non-MHC-I ligand which helps in identifying non-self to NK cells via an inhibitory TIGIT ligand. The TIGIT–CD155 pathway is a novel MHC-I-independent education mechanism for cell tolerance and activation of NK cells. The receptor also has a role in metastasis of cancer and transendothelial mechanism. In this review, the authors discuss the virus-host interaction that occurs via a single transmembrane receptor, the poliovirus infection pathway, which is being exploited as a therapeutic pathway. The oncolytic virotherapy is now a promising modality for curing cancer.

The incidence of cutaneous malignant melanoma is increasing worldwide. While the treatment of initial stages of the disease is simple, the advanced disease frequently remains fatal despite novel therapeutic options . This requires identification of novel therapeutic targets in melanoma. Similarly to other types of tumours, the cancer microenvironment plays a prominent role and determines the biological properties of melanoma. Importantly, melanoma cell-produced exosomes represent an important tool of intercellular communication within this cancer ecosystem. We have focused on potential differences in the activity of exosomes produced by melanoma cells towards melanoma-associated fibroblasts and normal dermal fibroblasts. Cancer-associated fibroblasts were activated by the melanoma cell-produced exosomes significantly more than their normal counterparts, as assessed by increased transcription of genes for inflammation-supporting cytokines and chemokines, namely IL-6 or IL-8. We have observed that the response is dependent on the duration of the stimulus via exosomes and also on the quantity of exosomes. Our study demonstrates that melanoma-produced exosomes significantly stimulate the tumour-promoting proinflammatory activity of cancer-associated fibroblasts. This may represent a potential new target of oncologic therapy .

Background/aim Per literature, patients with epidermal growth factor receptor (EGFR) exon-20 insertions respond poorly to tyrosine kinase inhibitors (TKIs). This study analyzed real-world data to examine the prognostic and predictive value of these mutations. Patients and methods We conducted a retrospective cohort study using Czech TULUNG Registry data, with data on multiple mutation types, collected in 2011-2020. Results We analyzed 554 (95.85%) patients with EGFR exon-19 deletions or exon-21 L858R substitutions and 24 (4.15%) patients with exon-20 insertions who received first-line high-value therapies. We summarized clinical characteristics and outcomes in all patients and by cohort. The risk of progression was statistically significantly higher (86%) in the exon-20 insertion cohort compared to the cohort with other mutations. Although not statistically significant, the risk of death was 44% higher in patients with exon-20 insertions. Conclusion Advanced NSCLC patients with rare EGFR exon-20 insertions have a high risk of progression.

Elevated levels of reactive oxygen species (ROS) reduce replication fork velocity by causing dissociation of the TIMELESS-TIPIN complex from the replisome. Here, we show that ROS generated by exposure of human cells to the ribonucleotide reductase inhibitor hydroxyurea (HU) promote replication fork reversal in a manner dependent on active transcription and formation of co-transcriptional RNA:DNA hybrids (R-loops). The frequency of R-loop-dependent fork stalling events is also increased after TIMELESS depletion or a partial inhibition of replicative DNA polymerases by aphidicolin, suggesting that this phenomenon is due to a global replication slowdown. In contrast, replication arrest caused by HU-induced depletion of deoxynucleotides does not induce fork reversal but, if allowed to persist, leads to extensive R-loop-independent DNA breakage during S-phase. Our work reveals a link between oxidative stress and transcription-replication interference that causes genomic alterations recurrently found in human cancer.

Ectomycorrhiza is a mutualistic interaction between higher land plants and certain fungi. The fungal partner receives carbohydrates from its plant partner and in return provides usable nitrogen and other nutrients. Not much is known about the mechanisms of the formation of the mycorrhizal structure and which signals play a role in both partners recognising each other. However, a number of genes have been found to be differentially regulated depending on mycorrhization. RNAsequencing and microarray analyses in the research group of Nehls and others have shown that many genes in poplar roots are differentially regulated depending on mycorrhiza. The aim of this work was to identify mycorrhiza-dependent elements in the promoter sequences of two genes, Potrx6G1673 and Potrx6G0093. For this purpose, corresponding promoter fragments were amplified from genomic DNA and tested in transgenic poplars. Composite poplars were produced in which only the roots are transgenic, but not the shoots (Neb et al. 2017). In this work, different visual markers were tested for their suitability to visualise the corresponding promoter fragments in the roots in order to detect differential expression of these fragments. Fluorescent proteins were chosen as visual markers because they allow in vivo analyses. On the one hand, super yellow fluorescent protein (sYFP) was tested with different promoter fragments in poplar roots, but due to the small size of the fluorescent protein, the visualisation results were not satisfactory. These problems occurred with both sYFP with nuclear localisation sequence and sYFP with peroxisomal localisation sequence. Therefore, double green fluorescent protein (dGFP) was established as a fluorescent marker for visualising promoter fragment activity. In the course of the work, it became apparent that the originally selected binary vector for plant transformation, pGreen, was not suitable for answering the research question of this work, so the produced constructs were cloned into another binary plant transformation vector, pCXUN. This system of binary vector and dGFP as visual marker was established in various experiments and subsequently successfully used to study the differential expression of corresponding poplar promoter fragments. Initial results indicate that the two promoters investigated appear to be subject to different regulatory mechanisms.

Pets play a pivotal role as definitive or reservoir hosts for many zoonotic parasites. Dogs and cats without any clinical signs may be a carrier for the infection. In a one-year study, collected fecal samples of 257 dogs and 50 cats were examined coproscopically for endoparasite infections. Out of 307 investigated fecal samples, 107 (34.9%) were positive for the presence of the propagative stages of endoparasites In 257 dogs fecal samples, following 12 different species of endoparasites were detected: Giardia duodenalis, Cystoisospora spp., Sarcocystis spp., Hammondia/Neospora-like eggs, Angiostrongylus vasorum larvae, Capillaria aerophila, Crenosoma vulpis, Toxocara spp., Toxascaris leonina, Trichuris vulpis, Strongyloides stercoralis, and eggs from the family Ancylostomatidae. Only 4 different parasitic species were found in 50 domestic cat fecal samples - G. duodenalis cysts Cystoisospora spp., T. cati, and larvae of Aelurostrongylus abstrusus. It was confirmed that significant differences were found concerning age for G. duodenalis, T. canis, S. stercoralis, and family Ancylostomatidae. Close and frequent contact between pets and people increases the risks for the transmission of zoonotic diseases.

Lice are blood-sucking insects that are of medical and veterinary significance as parasites and vectors for various infectious agents. More than half of described blood-sucking lice species are found on rodents. Rodents are important hosts of several Bartonella and Rickettsia species, and some of these bacteria are characterized as human pathogens in Europe. Rodent ectoparasites, such as fleas and ticks, are important vectors of Bartonella spp. and Rickettsia spp., but knowledge about the presence of these bacteria in lice is limited. The aim of this study was to determine the prevalence of Bartonella and Rickettsia bacteria in lice collected from rodents in Slovakia. The ectoparasites were collected from small rodents captured from 2010 to 2015 at four different sites in eastern Slovakia. The presence of Bartonella and Rickettsia species in lice samples was screened by real-time PCR, targeting ssrA and gltA genes, respectively. The molecular characterization of the Bartonella strains was based on sequence analysis of partial rpoB and intergenic spacer (ITS) genes, and of the Rickettsia species on sequence analysis of the gltA gene. A total of 1074 lice of seven species were collected from six rodent species. Bartonella DNA was detected in Hoplopleura affinis (collected from Apodemus agrarius, Apodemus flavicollis, and Myodes glareolus), Polyplax serrata (from A. agrarius), and Hoplopleura sp. (from A. flavicollis). Sequence analysis revealed that the Bartonella strains belonged to the Bartonella coopersplainsensis, Bartonella tribocorum, and Bartonella taylorii genogroups. Rickettsia DNA was detected in H. affinis and P. serrata collected from A. agrarius. Sequence analysis revealed two Rickettsia species: Rickettsia helvetica and Rickettsia sp. The results of the study confirm the presence of Bartonella spp. and Rickettsia spp. in lice collected from rodents.

Background Data about the genotype-specific human papillomavirus (HPV) prevalence in the Czech Republic is limited. We aimed to evaluate the prevalence and concordance of genotype-specific HPV infection detected in semen samples, penile swabs and cervical swabs from non-vaccinated heterosexual couples without HPV-associated disease. Methods Semen samples and penile swabs were collected from male partners and cervical swabs were collected from female partners of heterosexual couples treated for infertility (n = 195). Presence of HPV DNA in semen samples and cervical swabs was analyzed using the cobas® HPV Test and PapilloCheck®. Only the PapilloCheck® test was used to detect HPV in penile swabs. The genotype-specific prevalence and concordance of HPV infection not targeted by vaccine were evaluated using Fisher exact test. Results Both partners were infected with any HPV type in 13.8% (27/195) of couples and, of these couples, 55.6% (15/27) harbored at least one mutual genotype. High-risk HPV (hrHPV) genotypes were detected in 12.3% (24/195) of semen samples, 31.3% (61/195) of penile swabs, and 19.5% (38/195) of cervical swabs (P P P = 0.055). Conclusions This study showed that the detection of HPV infection differ by anatomic site and gender. Regardless the anatomic site, high prevalence of HPV genital infection was found in both Czech men and women.

Background/Aim: We investigated efficacy differences for afatinib versus gefitinib in non-small-cell lung cancer (NSCLC) according to epidermal growth factor receptor (EGFR) mutations. Patients and Methods: We retrospectively analysed data for 343 patients with NSCLC with performance status 1 having EGFR mutations treated with gefitinib or afatinib. Overall response rate (ORR) was tested by Fisher & rsquo;s exact test. Overall (OS) and progression free (PFS) survival were estimated by Kaplan & ndash;Meier method. Results: ORR did not differ in any group or subgroup. Among all patients, we observed significantly longer PFS for those treated with afatinib vs. gefitinib (median 13.4 vs. 9.5 months, p=0.026), but only a nonsignificant trend was observed for OS. We showed nonsignificant trends of better PFS and OS using afatinib for exon 19 deletion and L858R subgroups. We observed no significant PFS differences for other EGFR mutations but a nonsignificant trend towards better OS for those treated with afatinib. Conclusion: Afatinib led to longer PFS for patients with common EGFR mutations but not for those with rare mutations.

Arachis hypogaea, L. is an oil seed crop with a worldwide importance and the seeds are eaten at one of several stages from immature to fully ripe, raw, or cooked. The aim of the study was to analyze specific iPBS fingerprints of twenty-one accessions of peanut that were collected in the places of their natural occurrence in Bolivia and describe the existing genetic polymorphism. For genomic imprinting, three different iPBS markers were chosen - 1882, 2079, 2274 and a PCR reactions were performed. Obtained iPBS fingerprints were evaluated for the presence/absence of individual amplified loci and scored in 1/0 matrices. A Jaccard coefficient of genetic similarity was applied in UPGMA analysis for dendrogram construction. Polymorphism level was achieved in the range from 48% up to the 75% per primer. None of the iPBS markers used in the study was considered to distinguish all of the analyzed peanut accessions, but combining them in the final analysis, the level of genomic polymorphism was sufficient to clear separating of iPBS fingerprints of the collected accessions and unique iPBS loci were recorded in genomes of some of them. We found that, by selecting the appropriate iPBS markers, it is possible to characterize the peanut genome in the individual level with a specific fingerprint.

Natural products have been attracting increasing attention in human diet, both due to the possible negative effects of synthetic food additives on human health and the increased consumer perception. Apricot seeds contain a wide variety of bioactive components and their consumption is associated with a reduced risk of chronic diseases. The objective of the present study was to evaluate the effect of consumption of bitter apricot seeds on blood lipid and endocrine profile in Slovak women (n = 18, 41.60 ± 11.28 years) of reproductive age. Volunteers consumed 60 mg.kg

The present study investigates the effects of Enterococcus durans ED26E/7 beneficial strain and its enterocin – durancin (Ent) ED26E/7 on selected parameters in rabbits: growth performance, caecal enzymatic activity, jejunal morphometry and meat physico-chemical characteristics. Seventytwo rabbits (aged five weeks, M91 meat line, both sexes) were divided into experimental groups E1 (E. durans ED26E/7 strain; dose 500 μL/animal/day, concentration 109 CFU/mL) and E2 (durancin EntED26E/7; dose 50 μL/animal/day, with activity 12 800 AU/mL) and control group (C). The additives were administered in drinking water for a period of 21 days. All animals remained in good health during the experiment. The highest body weight gain (increase by 1.5% compared to C) was noted in E1 group during ED26E/7 strain application (PE. durans ED26E/7 strain and its durancin ED26E/7. The diet supplementation with bacteriocinogenic and probiotic E. durans ED26E/7 strain and its EntED26E/7 may improve the growth performance, caecal enzymatic activity and jejunal morphometry of rabbits, without any negative effect on rabbit meat quality.

This study was conducted to estimate the apparent digestibility coefficients (ADC) of the nutrients and energy contained in banana peel, cassava peel, cocoa husk, copra waste, and sugarcane bagasse in GIFT Nile tilapia (initial body weight 22.38 ± 0.33 g). A reference diet was formulated to satisfy the nutrient requirement of the fish. Five test diets were formulated to contain 70% of the reference diet and 30% of each test ingredient. The experiment lasted 42 days. The ADCs of dry matter (DM), crude lipid (CL), ash, crude fiber (CF), total carbohydrate (TC), and gross energy (GE) were 16.19%, 6.56%, 11.40%, 38.98% and 15.68% for banana peel; the ADCs of DM, crude protein (CP), CL, ash, CF, TC, and GE were 69.11%, 31.61%, 61.07%, 68.88%, 73.14%, 71.24%, and 69.00% for cassava peel; the ADCs of DM, ash, CF, TC, and GE were 10.56%, 0.84%, 28.70%, 2.82%, and 8.459.69% for cocoa husk; the ADCs of DM, CP, CL, ash, CF, TC, and GE were 63.08%, 20.14%, 66.60%, 32.53%, 69.70%, 66.21%, and 64.06% for copra waste; and the ADCs of DM, CF, TC, and GE were 15.22%, 36.71%, 34.34%, and 15.55% for sugarcane bagasse in tilapia respectively. This experiment generated basic information for the use of these ingredients in diet formulation for tilapia. However, considering the relatively low ADCs of nutrients observed in the present experiment, further processing could be an interesting avenue in improving the nutritional value and digestibility of these ingredients.

Aging of the immune system, or immunosenescence, alters the viral immune response in the elderly, especially when frailty exists. Research findings have demonstrated an imbalance in pro- and anti-inflammatory mechanisms, reduced production and diversification of T lymphocytes, and an alteration in immunovigilance and antibody synthesis. In this context, nutrition has a role in combating sarcopenia and frailty. Some food components that contribute to immune-competence are protein, vitamin D, n-3 fatty acids, antioxidant vitamins (vitamins C and E), zinc, selenium and iron. In times of a pandemic, nutritional recommendations for immune-competence in the elderly should be based on clinical studies. In this article, immunosenescence and its relationship to nutrition are addressed, including interventions studied in the context of the COVID-19 pandemic.

Drug discovery is still behind in the race compared to vaccine discovery in fighting COVID-19. Recently, a few alkaloids from a traditional Indian medicinal plant, Vasaka (Justicia adhatoda), have been linked computationally to the main protease (Mpro) of SARS_CoV_2. To expand the knowledge and for further investigation, we have selected 41 quinazoline alkaloids from two natural product databases to create an adequate library and performed detailed computational studies against the main protease (Mpro) of SARS_CoV_2. The screening of the library was carried out through blending the rigid docking and pharmacokinetic analysis that resulted in nine alkaloids as initial leads against Mpro. These nine alkaloids were further subjected to advance flexible docking using first reference famotidine for the analysis of structure-based interactions. For further selection, a second screening was carried out based on binding energies and interaction profiles that yielded three alkaloids namely CNP0416047, 3-hydroxy anisotine and anisotine as hits. The stereo-electronic features of hit alkaloids were further investigated through additional structure-based E-pharmacophore mapping against a second reference, known X77 ligand. Additionally, the reactivity of hit alkaloids at the binding site of the protein was estimated by measuring the electron distribution on the frontier molecular orbitals and HOMO-LUMO band energies. Finally, the stabilities of complexes between hit alkaloids with the protein were accessed extensively using robust molecular dynamics simulation through RMSD, RMSF, Rg, and MM-PBSA calculation. Thus, this study identifies three natural quinazoline alkaloids as potential inhibitors of MPro through extensive computational analysis.

International audience; A longstanding research divide exists in plant ecology: either focusing on plant clonality, with no ambition to address nonclonal plants, or focusing on all plants, ignoring that many ecological processes can be affected by the fact that some plants are clonal while others are not. This gap cascades into a lack of distinction and knowledge about the similarities and differences between clonal and nonclonal plants. Here we aim to bridge this gap by identifying areas that would benefit from the incorporation of clonal growth into one integrated research platform: namely, response to productivity and disturbance, biotic interactions, and population dynamics. We are convinced that this will provide a roadmap to gain valuable insights into the ecoevolutionary dynamics relevant to all plants.

Transcription factors (TFs) must bind at specific genomic locations to accurately regulate gene expression. The ability of TFs to recognize specific DNA sequence motifs arises from the inherent preferences of their globular DNA-binding domains (DBDs). Yet, these preferences are insufficient to explain the in vivo TF binding site selection. TFs are enriched with intrinsically disordered regions (IDRs), most of which are poorly characterized. While not generally considered as determinants of TF binding specificity, IDRs guide protein-protein interactions within transcriptional condensates, and multiple examples exist in which short IDRs flanking the DBD contribute to binding specificity via direct contact with the DNA. We recently reported that long IDRs, present away from the DBD, act as major specificity determinants at the genomic scale. Here, we discuss mechanisms through which IDRs contribute to DNA binding specificity, highlighting the role of long IDRs in dictating the in vivo binding site selection.

Organ and tissue growth result from an integration of biophysical communication across biological scales, both in time and space. In this review, we highlight new insight into the dynamic connections between control mechanisms operating at different length scales. First, we consider how the dynamics of chemical and electrical signaling in the shape of gradients or waves affect spatiotemporal signal interpretation. Then, we discuss the mechanics underlying dynamic cell behavior during oriented tissue growth, followed by the connections between signaling at the tissue and organismal levels.

Lake Ontario boasts a diverse fish community comprised of native and introduced species that support vibrant recreational, commercial and Indigenous fisheries. The effective delivery of a program to assess and control the sea lamprey (Petromyzon marinus) is crucial to achievement of Lake Ontario Fish Community Objectives of rehabilitating native fish stocks while protecting and maintaining the abundance of introduced salmonines. During 2000–2019, the Great Lakes Fishery Commission (GLFC) and its control agents, Fisheries and Oceans Canada (DFO) and the U. S. Fish and Wildlife Service (USFWS), delivered a consistent program of sea lamprey assessment and control. Beginning in 2004, rising sea lamprey abundance and marking rates on lake trout (Salvelinus namaycush) in Lake Ontario coincided with a decline of large lake trout in gillnet surveys. Efforts were undertaken to identify and control important sources of juvenile sea lampreys, including larvae that survived treatment, inhabited deepwater areas, or colonised previously uninhabited stream reaches and tributaries. A renewed reliance on proven conventional controls, including lampricide treatments and barriers, has resulted in consistent suppression of sea lamprey abundance and lake trout marking to prescribed targets in Lake Ontario during 2014–2019. This achievement is unprecedented in the 49-year history of Sea Lamprey Control Program delivery in Lake Ontario, and is attributable to the collaborative efforts of the GLFC, the control agents, and federal, provincial, state, and Indigenous partners.

In this work, we investigated in vitro the antioxidant, cytotoxic and anti-leishmanial activities of a lignin extracted from the leaves of Morinda citrifolia. Initially, an analysis of the composition of the sheets was performed, then the lignin was obtained by alkaline delignification and characterized by different techniques: elemental analysis, FT-R, UV–vis, HSQC-NMR, thermal analysis, Py-GC/MS and by GPC. The results showed that the leaves had in their composition cellulose (31.29%), hemicellulose (25.01%), lignin (18.34%), extractives (14.39%) and ash (10.03%). The lignin extraction yield was 89.8%. The lignin obtained is of the GSH type with the following contents 79.39%, 13.58% and 7.03% respectively. Furthermore, it is low molecular weight and thermally stable. It had a phenolic content of 93.3 mg GAE/g and low antioxidant activity. In macrophage cytotoxicity assays, it presented a CC50 of 31.0 μg/mL, showing less toxicity than amphotericin B. In assays against the promastigote forms of Leishmania amazonensis, lignin presented an IC50 of 29.56 μg/mL, a less effective concentration than amphotericin B (IC50 = 0.14 μg/mL). However, it was able to promote inhibition of the parasites, a fact confirmed by structural changes. These findings reinforce that M. citrifolia lignin is a promising macromolecule for use as an antiparasitic and antioxidant agent.

The formation of stress granules (SGs), membrane-less organelles which are comprised of mainly messenger ribonucleoprotein assemblies, is the result of a conserved evolutionary strategy to cellular stress. During their formation which is triggered by robust environmental stress, SGs sequester translationally inactive mRNA molecules, which are either forwarded for further processing elsewhere or stored during a period of stress within SGs. Removal of mRNA molecules from active translation and their sequestration in SGs allows preferential translation of stress response transcripts. By affecting the specificity of mRNA translation, mRNA localization and stability, SGs are involved in the overall cellular reprogramming during periods of environmental stress and viral infection. Over the past two decades, we have learned which processes drive SGs assembly, how their composition varies under stress, and how they co-exist with other subcellular organelles. Yeast, as a model has been instrumental in our understanding of stress granule biology. Despite the specific differences between the stress granules of yeast and mammals, yeast have been shown to be a valuable tool to the study of SGs in translation-related stress response. This review summarizes the data surrounding SGs that are formed under different stress conditions in Saccharomyces cerevisiae and other yeast species. It offers a comprehensive and up-to-date view on these still somewhat mysterious entities.

Introduction Pre-analytical sample handling might affect the results of Alzheimer's disease blood-based biomarkers. We empirically tested variations of common blood collection and handling procedures. Methods We created sample sets that address the effect of blood collection tube type, and of ethylene diamine tetraacetic acid plasma delayed centrifugation, centrifugation temperature, aliquot volume, delayed storage, and freeze-thawing. We measured amyloid beta (Aβ)42 and 40 peptides with six assays, and Aβ oligomerization-tendency (OAβ), amyloid precursor protein (APP)699-711 , glial fibrillary acidic protein (GFAP), neurofilament light (NfL), total tau (t-tau), and phosphorylated tau181. Results Collection tube type resulted in different values of all assessed markers. Delayed plasma centrifugation and storage affected Aβ and t-tau; t-tau was additionally affected by centrifugation temperature. The other markers were resistant to handling variations. Discussion We constructed a standardized operating procedure for plasma handling, to facilitate introduction of blood-based biomarkers into the research and clinical settings.

Detached leaves of ivy (Hedera helix L.) were used as a model for the study of zinc uptake and transport in vascular plants. By the uptake via the surface of fully immersed leaves in 25 % Hoagland nutrient media (HM) spiked with 65ZnCl2 (50 μmol/dm3 ZnCl2), concentration in leaves 4.98 μg Zn/g (dry wt.), i. e. 2.6 μg Zn/dm2 leaf area after 7d exposition were obtained. By the uptake via immersed stalks of not immersed (transpiring) leaves concentrations up to 370 μg Zn/g (dry wt.) were obtained. When Zn enters into detached leaves via the surface of immersed leaf blades, zinc is uniformly distributed in leaf blades and leaf stalks. When zinc enters detached leaves via immersed stalks of non-immersed transpiring leaves, only small part of zinc is transported to leaf blades and the prevailing part remains in leaf stalks. Stalks act as a trap, able to prevent other leaf tissues against inhibitory effects of high Zn concentrations. Mineral nutrient salts in solutions mobilize Zn trapped in leaf stalks and facilitate Zn transport by transpiration stream to leaf blades, what means that Zn in stalks is bound in ion-exchageable forms.

Temperature and thermal variability are increasing worldwide, with well-known survival consequences. However, effects on other potentially more thermally sensitive reproductive traits are less understood, especially when considering thermal variation. Studying the consequences of male reproduction in the context of climate warming and ability to adapt is becoming increasingly relevant. Our goals were to test how exposure to different average temperatures that either fluctuated or remained constant impacts different male reproductive performance traits and to assess adaptive potential to future heat stress. We took advantage of a set of Drosophila melanogaster isogenic lines of different genotypes, exposing them to four different thermal conditions. These conditions represented a benign and a stressful mean temperature, applied either constantly or fluctuating around the mean and experienced during development when heat stress avoidance is hindered because of restricted mobility. We measured subsequent male reproductive performance for mating success, fertility, number of offspring produced and offspring sex ratio, and calculated the influence of thermal stress on estimated heritability and evolvability of these reproductive traits. Both costs and benefits to different thermal conditions on reproductive performance were found, with some responses varying between genotypes. Mating success improved under fluctuating benign temperature conditions and declined as temperature stress increased regardless of genotype. Fertility and productivity were severely reduced at fluctuating mean high temperature for all genotypes, but some genotypes were unaffected at constant high mean temperature. These more thermally robust genotypes showed a slight increase in productivity under the fluctuating benign condition compared to constant high temperature, despite both thermal conditions sharing the same temperature for 6 hr daily. Increasing thermal stress resulted in higher heritability and evolvability. Overall, the effects of temperature on reproductive performance depended on the trait and genotype; performance of some traits slightly increased when high temperatures were experienced for short periods but decreased substantially even when experiencing a benign temperature for a portion of each day. While thermal stress increased genetic variation that could provide adaptive potential against climate warming, this is unlikely to compensate for the overall severe negative effect on reproductive performance as mean temperature and variance increase.

For genotoxicity study simultaneous phytotoxicity and mutagenicity assay with Vicia sativa L. var. Klára was used. For phytotoxicity the following rank orders of growth inhibition can be arranged: for roots: Ni(II) > Cr(VI) > Cr(III); for shoots: Ni(II) > Cr(VI) ≥ Cr (III). For mutagenicity assay root tips of V. sativa were used and chromosome aberrations were determined at least in 500-anatelophases. All tested metals exerted in V. sativa a significant increase of chromosomal aberration rate in applied concentrations. Maximum of aberrations invoked Cr(VI) and the rank order of aberrations fall was: Cr(VI) > Ni(II) > Cr(III). Genotoxic effects of metals were determined by analysis of micronuclei frequency in the pollen tetrads of Tradescantia plants. None of tested metal significantly stimulated micronuclei frequency and genotoxic effect was decreased in order: Cr(VI) ≥ Ni(II) > Cr(III).

While seasonal trends in testosterone levels are known from cross-cohort studies, data on testosterone inter-annual individual repeatability in wild birds are rare. Also, our understanding of hormonal age-dependent changes in testosterone levels is limited. We assessed plasma testosterone levels in 105 samples originating from 49 repeatedly captured free-living great tits (Parus major) sampled during the nesting to investigate their relative long-term repeatability and within-individual changes. Furthermore, we examined the inter-annual repeatability of condition-related traits (carotenoid- and melanin-based plumage ornamentation, ptilochronological feather growth rate, body mass, and haematological heterophil/lymphocyte ratio) and their relationships to testosterone levels. We show that testosterone levels are inter-annually repeatable in females, with a non-significant pattern in males, both in absolute values and individual ranks (indicating the maintenance of relative status in a population). In males, we found a quadratic dependence of testosterone levels on age, with a peak in midlife. In contrast, female testosterone levels showed no age-dependent trends. The inter-annual repeatability of condition-related traits ranged from zero to moderate and was mostly unrelated to plasma testosterone concentrations. However, males with elevated testosterone had significantly higher carotenoid-pigmented yellow plumage brightness, a trait presumably involved in mating. Showing inter-annual repeatability in testosterone levels, this research opens the way to further understanding the causes of variation in condition-related traits. Based on a longitudinal dataset, this study demonstrates that male plasma testosterone undergoes age-related changes that may regulate resource allocation. Our results thus suggest that, unlike females, male birds undergo hormonal senescence similar to mammals.