Soil properties influence the toxicity and availability of Zn from ZnO nanoparticles to earthworms.

To develop models that support site-specific risk assessment for nanoparticles (NPs), a better understanding of how NP transformation processes, bioavailability and toxicity are influenced by soil properties is needed. In this study, the influence of differing soil properties on the bioavailability and toxicity of zinc oxide (ZnO) NPs and ionic Zn to the earthworm Eisenia fetida was investigated. Earthworms were exposed to ZnO_NPs and ionic Zn, between 100 and 4400 mg Zn/kg, in four different natural soils (organic matter content: 1.8-16.7%, soil pH: 5.4-8.3, representing sandy loam to calcareous soils). Survival and reproduction were assessed after 28 and 56 days, respectively. Zn concentrations in soil pore waters were measured while labile concentrations of Zn were measured using an in-situ dynamic speciation technique (diffusive gradient in thin films, DGT). Earthworm Zn tissue concentrations were also measured. Soil properties influenced earthworm reproduction between soil controls, with highest reproductive output in soils with pH values of 6-7. Toxicity was also influenced by soil properties, with EC50s based on total Zn in soil ranging from 694 to >2200 mg Zn/kg for ZnO_NP and 277-734 mg Zn/kg for ionic Zn. Soil pore water and DGT measurements showed good agreement in the relative amount of Zn extracted across the four soils. Earthworms exposed to ZnO_NPs survived higher Zn concentrations in the soils and had higher tissue concentrations compared with ionic Zn exposures, particularly in the high organic content calcareous soil. These higher tissue concentrations in ZnO_NP exposed earthworm could have consequences for the persistence and trophic mobility of Zn in terrestrial systems and need to be further investigated to elucidate if there any longer-term risks associated with sustained input of ZnO_NP to soil.

A Kinetic Approach for Assessing the Uptake of Ag from Pristine and Sulfidized Ag Nanomaterials to Plants.

Nanomaterials (NMs) are thermodynamically unstable by nature, and exposure of soil organisms to NMs in the terrestrial environment cannot be assumed constant. Thus, steady-state conditions may not apply to NMs, and bioaccumulation modeling for uptake should follow a dynamic approach. The one-compartment model allows the uptake and elimination of a chemical to be determined, while also permitting changes in exposure and growth to be taken into account. The aim of the present study was to investigate the accumulation of Ag from different Ag NM types (20 nm Ag0 NMs, 50 nm Ag0 NMs, and 25 nm Ag2 S NMs) in the crop plant wheat (Triticum aestivum). Seeds were emerged in contaminated soils (3 or 10 mg Ag/kg dry soil, nominal) and plants grown for up to 42 d postemergence. Plant roots and shoots were collected after 1, 7, 14, 21, and 42 d postemergence; and total Ag was measured. Soil porewater Ag concentrations were also measured at each sampling time. Using the plant growth rates in the different treatments and the changing porewater concentrations as parameters, the one-compartment model was used to estimate the uptake and elimination of Ag from the plant tissues. The best fit of the model to the data included growth rate and porewater concentration decline, while showing elimination of Ag to be close to zero. Uptake was highest for Ag0 NMs, and size did not influence their uptake rates. Accumulation of Ag from Ag2 S NMs was lower, as reflected by the lower porewater concentrations. Environ Toxicol Chem 2021;40:1861-1872. © 2021 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Transplacental transmission of Neospora caninum in moose (Alces alces).

A 5-years-old moose (Alces alces) cow kept in a zoo in the German Federal State of Brandenburg aborted a female foetus of 44cm crown rump length (CRL). Pathohistological analysis revealed several Neospora (N.) caninum infected cells and cysts, as well as multifocal gliosis, necrosis, haemorrhages, dystrophic mineralisation and haemosiderosis in the brain, predominantly in cerebrum and brainstem. In addition, mild lymphocytic meningitis was present. Together with the fresh foetus, a mummified foetus of 16cm CRL was expelled. Neither focal necrosis, nor inflammation was detected in the brain of the mummified foetus. By two polymerase chain reactions (PCR) targeting the pNc5 gene of N. caninum (i.e. an end point PCR and a real-time PCR), by two serological methods (immunofluorescence test and immunoblot), by histological and immunohistochemical analyses, transplacental N. caninum infection was confirmed in the fresh foetus and interpreted as possible cause of abortion. Infection with other agents causing abortion including Bovine Herpesvirus 1 (BHV1), Bluetongue Virus (BTV), Bovine Virus Diarrhoea Virus (BVDV), Brucella spp., Chlamydia spp., Coxiella burnetii and Toxoplasma gondii were excluded. Our findings show that control measures may be necessary to protect captive moose against accidental N. caninum infection. Further studies are needed to explore the importance of neosporosis in wild and captive moose.

Reduced magnetisation transfer ratio in cognitively impaired patients at the very early stage of multiple sclerosis: a prospective, multicenter, cross-sectional study.

OBJECTIVES: Cognitive impairment belongs to the core symptoms in multiple sclerosis (MS) and can already be present at the very early stages of the disease. The present study evaluated cognitive functioning after the first clinical presentation suggestive of MS and brain tissue damage in a non-lesion focused MRI approach by using magnetisation transfer imaging (MTI). SETTING AND PARTICIPANTS: 47 patients (15 men and 32 women; mean age: 31.17 years) after the first clinical event suggestive of MS were recruited in six different MS centres in Germany and underwent a neuropsychological test battery including tests for attention, memory and executive function as well as depression and fatigue. MTI and conventional MRI measures (T1/T2 lesion load) were assessed. In addition, Magnetisation Transfer Ratio (MTR) maps were calculated. Primary outcome measure was the investigation of cognitive dysfunction in very early MS in correlation to MRI data. RESULTS: 55.3% of patients with MS failed at least one test parameter. Specifically, 6% were reduced in working memory, 14.9% in focused attention, 25.5% in figural learning and up to 14.9% in executive function. When the sample was subdivided into cognitively impaired and preserved, MTR scores within the cognitively impaired subgroup were significantly lower compared with the preserved group (t(43)=2.346, p=0.02*). No significant differences between the two groups were found in T2-weighted and T1-weighted lesion volume. CONCLUSIONS: After the first MS-related clinical event, 55.3% of patients showed distinct cognitive deficits. Cognitively impaired patients had significantly lower whole brain MTR, but no differences in focal brain lesion volumes supporting the idea that early cognitive deficits may be related to diffuse loss of brain tissue integrity.

Use of forward genetic screens to identify genes required for RNA-directed DNA methylation in Arabidopsis thaliana.

RNA-directed DNA methylation is a small RNA-mediated epigenetic modification that contributes to transcriptional silencing of transposons and repetitive sequences in plants. We have conducted several forward genetic screens to identify factors required for RNA-directed DNA methylation and transcriptional gene silencing in Arabidopsis thaliana. Here, we review the findings from these screens and report on two new mutants, dms12 and dms13, that are defective in Pol V-specific subunits NRPE5 and NRPE9b. Cumulative results from genetic screens performed in our laboratory and those of other investigators have revealed that RNA-directed DNA methylation requires a complex transcriptional machinery comprising a number of plant-specific factors, many of which were functionally uncharacterized before being implicated in this pathway. Future challenges include unraveling the detailed mechanism and full range of functions of RNA-directed DNA methylation.

Evaluation of sampling tools for environmental sampling of bacterial endospores from porous and nonporous surfaces.

AIMS: Having and executing a well-defined and validated sampling protocol is critical following a purposeful release of a biological agent for response and recovery activities, for clinical and epidemiological analysis and for forensic purposes. The objective of this study was to address the need for validated sampling and analysis methods called out by the General Accounting Office and others to systematically compare the collection efficiency of various swabs and wipes for collection of bacterial endospores from five different surfaces, both porous and nonporous. This study was also designed to test the collection and extraction solutions used for endospore recovery from swabs and wipes. METHODS AND RESULTS: Eight collection tools, five swabs and three wipes, were used. Three collection/preservation solutions were evaluated: an ink jet aerosol generator was used to apply Bacillus subtilis endospores to five porous and nonporous surfaces. The collection efficiencies of the swabs and wipes were compared using a statistical multiple comparison analysis. CONCLUSIONS: The ScottPure wipe had the highest collection efficiency and phosphate-buffered saline (PBST) with 0.3% Tween was the best collection solution of those tested. SIGNIFICANCE AND IMPACT OF THE STUDY: Validated sampling for potential biological warfare is of significant importance and this study answered some relevant questions.

Effect of hormone replacement therapy on epidermal barrier lipids.

Current evidence of the effect of hormone replacement therapy (HRT) on skin lipids, of postmenopausal women is scanty and indirect. Here, we report the ultrastructural differences in epidermal lipids between postmenopausal subjects who were and were not on HRT and a comparison is made with younger subjects. Biopsies were obtained from arms and legs, in a blinded, no-treatment, study conducted on postmenopausal subjects who were and were not on HRT and younger subjects. The ultrastructure of skin lipids and the lipid coverage of underlying corneocytes were compared for biopsies obtained from different subjects. Qualitative assessment as well as quantitative estimation of lipid-covered regions of corneocytes shows that skin lipids do not cover corneocytes effectively in postmenopausal women who are not on HRT. However, women who are on HRT show significantly improved lipid coverage of corneocytes, which is comparable with the younger subjects. This implies that HRT should improve the lipid coverage and skin condition of postmenopausal women.

RNA-directed DNA methylation and Pol IVb in Arabidopsis.

Recent work in Arabidopsis has revealed a plant-specific RNA polymerase, pol IV, that is specialized for RNA interference (RNAi)-mediated, chromatin-based gene silencing. Two functionally diversified pol IV complexes have been identified: pol IVa is required to produce or amplify the small RNA trigger, whereas pol IVb, together with the plant-specific SWI/SNF-like chromatin remodeling factor DRD1, acts downstream from small RNA formation to induce de novo cytosine methylation of homologous DNA by an unknown mechanism. Retrotransposon long terminal repeats (LTRs) and other unannotated sequences that encode small RNAs are prime targets for DRD1/pol IVb-mediated cytosine methylation. In drd1 and pol IVb mutants, silent LTRs in euchromatin can be derepressed, resulting in enhanced transcription of adjacent genes or intergenic regions. In addition to mediating de novo methylation, some evidence suggests that DRD1 and pol IVb are also involved in a reciprocal process of active demethylation, perhaps in conjunction with DNA glycosylase domain-containing proteins such as ROS1. We speculate that DRD1/pol IV-dependent methylation/demethylation evolved in the plant kingdom as a means to facilitate rapid, reversible changes in gene expression, which might have adaptive significance for immobile plants growing in unpredictable environments.

Host defenses to transposable elements and the evolution of genomic imprinting.

Genomic imprinting is the differential expression of maternally and paternally inherited alleles of specific genes. Several organismic level hypotheses have been offered to explain the evolution of genomic imprinting. We argue that evolutionary explanations of the origin of imprinting that focus exclusively on the organismic level are incomplete. We propose that the complex molecular mechanisms that underlie genomic imprinting originally evolved as an adaptive response to the mutagenic potential of transposable elements (TEs). We also present a model of how these mechanisms may have been co-opted by natural selection to evolve molecular features characteristic of genomic imprinting.

Preferential elimination of repeated DNA sequences from the paternal, Nicotiana tomentosiformis genome donor of a synthetic, allotetraploid tobacco.

Nicotiana tabacum (tobacco, 2n = 4x = 48) is a natural allotetraploid combining two ancestral genomes closely related to modern Nicotiana sylvestris and Nicotiana tomentosiformis. Here we examine the immediate consequences of allopolyploidy on genome evolution using 20 S4-generation plants derived from a single synthetic, S0 plant made by Burk in 1973 (Th37). Using molecular and cytogenetic methods we analysed 14 middle and highly repetitive sequences that together total approximately 4% of the genome. Two repeats related to endogenous geminiviruses (GRD5) and pararetroviruses (NtoEPRV), and two classes of satellite repeats (NTRS, A1/A2) were partially or completely eliminated at variable frequency (25-60%). These sequences are all from the N. tomentosiformis parent. Genomic in situ hybridization revealed additivity in chromosome numbers in two plants (2n = 48), while a third was aneuploid for an N. tomentosiformis-origin chromosome (2n = 49). Two plants had homozygous translocations between chromosomes of the S- and T-genomes. * The data demonstrate that genetic changes in synthetic tobacco were fast, targeted to the paternal N. tomentosiformis-donated genome, and some of the changes showed concordance with changes that presumably occurred during evolution of natural tobacco.

Neutron spectrometry in mixed fields: superheated drop (bubble) detectors.

The BINS neutron threshold spectrometer permits the analysis of the main features of a neutron field for radiation protection purposes. The system offers a virtually complete photon discrimination and nested threshold responses to neutrons, which allow the use of very effective 'few-channel' unfolding procedures. To date, the practical operating energy range of a BINS is 0.1-10 MeV, over which a resolving power of 20-30% can be expected when the deconvolution is performed without explicit pre-information. Spectrum unfolding results in relatively high uncertainties on the differential fluence distributions, but due to negative correlations in adjacent energy groups the uncertainties on integral quantities such as dose equivalent are small and of the order of 5% to 10%, similar to the results of other active spectrometers. In comparison with most radiation detectors, the BINS is an extremely slow system due to the intrinsic duration of a bubble pulse and to the time associated with pulse analysis. For example, the maximum sustainable fluence rate of 1 MeV neutrons is about 10(4) cm(-2) s(-1), which is low for many neutron physics experiments. However, this rate corresponds to an ambient dose equivalent rate of about 1 mSv h(-1), making the active device adequate for radiation protection applications in the workplaces described in Section 1. There are ample margins for improvement of the spectrometer. In particular, in the low-energy region a thermal-epithermal neutron group may be added by using chlorine-bearing emulsions stabilised at suitable temperatures. In fact, the latest version of the system achieves this goal by using a single superheated emulsion of dichlorotetrafluoroethane (R-114) operated at temperatures up to 55 degrees C. This extends the range of the spectrometer and at the same time removes the undue enhancement of the UNFANA output in the low energy region. Above 10 MeV, the resolution can be improved by adding more thresholds, e.g. by starting from a lower initial temperature and using finer temperature increments. Based on neutron kinematics, the theoretical upper energy threshold which can be generated with superheated emulsions is greater than 100 MeV. However, this would most likely require refrigerating the detectors, while the current simpler approach is to operate the detectors at incremental temperature steps starting from the ambient temperature. A range that should be easily achieved in practice is from thermal energies to 20 MeV.

Endogenous viral sequences and their potential contribution to heritable virus resistance in plants.

Tobacco endogenous pararetroviruses (TEPRVs) represent the first virus-derived repetitive sequence family found in plants. The sequence conservation of TEPRVs and the lack of an exogenous form of the virus suggest that TEPRVs serve a beneficial function, perhaps by furnishing virus resistance via homologous sequence interactions. This hypothesis is supported by the observation that TEPRVs are methylated and negligibly transcribed. Moreover, transgenes driven by the TEPRV enhancer are silenced and methylated when introduced into tobacco, but remain active and unmethylated in non-host species devoid of sequences homologous to TEPRVs. In transgenic Arabidopsis, the TEPRV enhancer is active primarily in shoot meristems. This suggests that the virus giving rise to TEPRVs could infect germ cell precursors, a prerequisite for meiotically heritable insertions into host chromosomes. The copy number, organization and methylation of TEPRVs in tetraploid tobacco and one of its diploid ancestors, Nicotiana sylvestris, the presumed original host for the virus, have remained constant since polyploid formation. The remarkable conservation of these features in two independently evolving species further supports a role for TEPRVs in viral immunity.

Transposition of IS10 from the host Escherichia coli genome to a plasmid may lead to cloning artefacts.

During recloning of Nicotiana tabacum L. repetitive sequence R8.3 in Escherichia coli, a modified clone that differed from the original by the insertion of an IS10 sequence was unintentionally produced. The insert was flanked by a 9-bp direct repeat derived from the R8.3 sequence, the 9-bp duplication of acceptor DNA in the site of insertion being a characteristic of IS10 transposition events. A database search using the FASTA program showed IS10 and other prokaryotic IS elements inserted into numerous eukaryotic clones. Unexpectedly, the IS10, which is not a natural component of the E. coli genome, appeared to be by far the most frequent contaminant of DNA databases among several IS sequences tested. In the GenEMBL database, the IS10 query sequence yielded positive scores with more than 500 eukaryotic clones. Insertions of shortened IS10 sequences having only one intact terminal inverted repeat were commonly found. Most full-length IS10 insertions (32 out of 40 analyzed) were flanked by 9-bp direct repeats having the consensus 5'-NPuCNN-NGPyN-3' with a strong preference for 5'-TGCTNA-GNN-3'. One insertion was flanked by an inverted repeat of more than 400 bp in length. PCR amplification and Southern analysis revealed the presence of IS10 sequences in E. coli strains commonly used for DNA cloning, including some reported to be Tn10-free. No IS10-specific PCR product was obtained with N. tabacum or human DNA. Our data suggest that transposition of IS10 elements may accompany cloning steps, particularly into large BAC vectors. This might lead to the relatively frequent contamination of DNA databases by this bacterial sequence. It is estimated that one in approximately every thousand eukaryotic clone in the databases is contaminated by IS-derived sequences. We recommend checking submitted sequences for the presence of IS10 and other IS elements. In addition, DNA databases should be corrected by removing contaminating IS sequences.

A directional dose equivalent monitor for neutrons.

A directional dose equivalent monitor is introduced which consists of a 30 cm diameter spherical phantom hosting a superheated drop detector embedded at a depth of 10 mm. The device relies on the similarity between the fluence response of neutron superheated drop detectors based on halocarbon-12 and the quality-factor-weighted kerma factor. This implies that these detectors can be used for in-phantom dosimetry and provide a direct reading of dose equivalent at depth. The directional dose equivalent monitor was characterised experimentally with fast neutron calibrations and numerically with Monte Carlo simulations. The fluence response was determined at angles of 0, 45, 90, 135 and 180 degrees for thermal to 20 MeV neutrons. The response of the device is closely proportional to the fluence-to-directional dose equivalent conversion coefficient, h'phi (10; alpha, E). Therefore, our monitor is suitable for a direct measurement of neutron directional dose equivalent, H'(10), regardless of angle and energy distribution of the neutron fluence.

RNA: guiding gene silencing.

In diverse organisms, small RNAs derived from cleavage of double-stranded RNA can trigger epigenetic gene silencing in the cytoplasm and at the genome level. Small RNAs can guide posttranscriptional degradation of complementary messenger RNAs and, in plants, transcriptional gene silencing by methylation of homologous DNA sequences. RNA silencing is a potent means to counteract foreign sequences and could play an important role in plant and animal development.

Resistance of RNA-mediated TGS to HC-Pro, a viral suppressor of PTGS, suggests alternative pathways for dsRNA processing.

In plants, double-stranded (ds) RNA that is degraded to small (sm) RNAs that are approximately 23 nucleotides in length can trigger the degradation of homologous RNAs in the cytoplasm (posttranscriptional gene silencing or PTGS) and de novo methylation of homologous DNA in the nucleus [1]. PTGS is similar to quelling in fungi [2] and RNAi in animals [3]. RNA-directed DNA methylation (RdDM) can lead to transcriptional gene silencing (TGS) and the methylation of homologous target promoters if dsRNAs containing promoter sequences are involved [4]. HC-Pro is a plant viral suppressor of PTGS that acts by preventing the accumulation of smRNAs [5, 6] that provide the specificity determinant for homologous RNA degradation [7-10]. Here, we show that HC-Pro does not suppress TGS induced by promoter dsRNA. Moreover, the amount of promoter smRNAs is elevated 5-fold in the presence of HC-Pro, and target promoter methylation is slightly increased without a concomitant rise in the level of promoter dsRNA. The promoter dsRNA, which is not polyadenylated, failed to trigger substantial degradation of polyadenylated, single-stranded promoter RNA. The differential effects of HC-Pro on smRNA accumulation associated with dsRNA-mediated TGS and at least some cases of PTGS suggest that dsRNA processing can occur by alternative pathways, and they support the idea that RdDM is triggered by smRNAs.