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Arthropod-borne viruses (arboviruses) such as dengue, Zika, and chikungunya viruses cause morbidity and mortality among human populations living in the tropical regions of the world. Conventional mosquito control efforts based on insecticide treatments and/or the use of bednets and window curtains are currently insufficient to reduce arbovirus prevalence in affected regions. Novel, genetic strategies that are being developed involve the genetic manipulation of mosquitoes for population reduction and population replacement purposes. Population replacement aims at replacing arbovirus-susceptible wild-type mosquitoes in a target region with those that carry a laboratory-engineered antiviral effector to interrupt arboviral transmission in the field. The strategy has been primarily developed for Aedes aegypti(L.), the most important urban arbovirus vector. Antiviral effectors based on long dsRNAs, miRNAs, or ribozymes destroy viral RNA genomes and need to be linked to a robust gene drive to ensure their fixation in the target population. Synthetic gene-drive concepts are based on toxin/antidote, genetic incompatibility, and selfish genetic element principles. The CRISPR/Cas9 gene editing system can be configurated as a homing endonuclease gene (HEG) and HEG-based drives became the preferred choice for mosquitoes. HEGs are highly allele and nucleotide sequence-specific and therefore sensitive to single-nucleotide polymorphisms/resistant allele formation. Current research efforts test new HEG-based gene-drive designs that promise to be less sensitive to resistant allele formation. Safety aspects in conjunction with gene drives are being addressed by developing procedures that would allow a recall or overwriting of gene-drive transgenes once they have been released.

The mosquito-borne chikungunya virus (CHIKV) poses a threat to human health in tropical countries throughout the world. The molecular interactions of CHIKV with its mosquito vector Aedes aegyptiare not fully understood. Following oral acquisition of CHIKV via salinemeals, we analyzed changes in the proteome of Ae. aegyptiin 12 h intervals by label-free quantification using a timsTOF Pro mass spectrometer. For each of the seven time points, between 2647 and 3167 proteins were identified among CHIKV-infected and noninfected mosquito samples, and fewer than 6% of those identified proteins were affected by the virus. Functional enrichment analysis revealed that the three pathways, Endocytosis, Oxidative phosphorylation, and Ribosome biogenesis, were enriched during CHIKV infection. On the other hand, three pathways of the cellular RNA machinery and five metabolism related pathways were significantly attenuated in the CHIKV-infected samples. Furthermore, proteins associated with cytoskeleton and vesicular transport, as well as various serine-type endopeptidases and metallo-proteinases, were modulated in the presence of CHIKV. Our study reveals biological pathways and novel proteins interacting with CHIKV in the mosquito. Overall, CHIKV infection caused minor changes to the mosquito proteome demonstrating a high level of adaption between the vector and the virus, essentially coexisting in a nonpathogenic relationship. The mass spectrometry data have been deposited to the MassIVE repository (https://massive.ucsd.edu/ProteoSAFe/dataset.jsp?task=abfd14f7015243c69854731998d55df1) with the data set identifier MSV000085115.

Automotive manufacturing requires high productive manufacturing technologies to ensure a high-volume efficient production. Joining metal sheets in an overlap joint configuration of zinc-coated steel is an important task for body-in-white production. This paper describes the influence of different spot sizes regarding process stability in laser beam welding. Therefore, welds were performed in an overlap joint configuration with fully penetrated welds. Relevant material combinations for automotive industry were investigated. A broad-based parameter study and the analysis of interaction between spot- and keyhole diameter state a higher process stability. Welds with a smaller spot diameter lead to a sufficient increase of joint quality by using a zero gap overlap configuration. It is shown that the amount of joint instabilities such as pores, pinholes, spatter and further defects can be minimized by using smaller spot diameters. This can be explained by different keyhole regimes. Moreover, the change of isothermal interaction in the interlayer zone caused by the adjusted spot diameter leads to high-quality joints.

Advantages of tourniquet use in TKA include benefits for surgeons and patients, varying from a bloodless operation site to a reduced intervention time. The time under ischemia and the reperfusion period are crucial phases for affected soft-tissue, most commonly the extensor mechanism.

•DENV2 resistance in transgenic mosquitoes overexpressing an antiviral IR effector.•Design of a hammerhead ribozyme that efficiently blocks DENV2 in cell culture.•Trans-splicing Group I Intron blocking all four DENV serotypes in cell culture.

The self‐assembly of chiral depsipeptide dendrons 4, which contain a cyanuric acid building block at their focal point, with the homotritopic Hamilton receptor 1is reported. The 1:3 compositions of the resulting chiral supramolecular dendrimers, the association constants Kn, and the cooperativity of binding expressed by Scatchard plots and the Hill coefficients nHwas determined by NMR titration experiments. The most pronounced positive cooperativity was found for the complexes 1L3with L being the second‐generation dendrons 4 c–e. The least stable complexes are formed with the bulky third‐generation dendrons 4 f–h. Similar results are obtained by the corresponding complexation of the achiral Frechét‐type first‐ to third‐generation dendrons 3with 1. Chiroptical investigations of 1:3 complexes of 1and 4reveal chirality transfer from the dendron to the Hamilton receptor as demonstrated by the appearance of new CD absorption bands at 310 nm.

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ABSTRACTCucumber mosaic virus (CMV), the type member of the genus Cucumovirus(family Bromoviridae), is transmitted by aphids in a nonpersistent manner. Mutagenesis experiments identified the ßH-ßI loop of the capsid subunit as a potential key motif responsible for interactions with the insect vector. To further examine the functional characteristics of this motif, we generated monoclonal antibodies that bound to native virions but not to ßH-ßI mutants. Fab fragments from these antibodies were complexed with wild-type CMV and the virus-Fab structure was determined to 12-A° resolution by using electron cryomicroscopy and image reconstruction techniques. The electron density attributed to the bound antibody has a turret-like appearance and protrudes from each of the 12 fivefold axes of the icosahedral virus. Thus, the antibody binds only to the pentameric clusters (pentons) of A subunits of the T=3 quasisymmetric virus and does not appear to bind to any of the B and C subunits that occur as hexameric clusters (hexons) at the threefold (quasi-sixfold) axes. Modeling and electron density comparisons were used to analyze the paratope-epitope interface and demonstrated that the antibody binds to three ßH-ßI loops in three adjacent A subunits in each penton. This antibody can discriminate between A and B/C subunits even though the ßH-ßI loop adopts the same structure in all 180 capsid subunits and is therefore recognizing differences in subunit arrangements. Antibodies with such character have potential use as probes of viral assembly. Our results may provide an additional rationale for designing synthetic vaccines by using symmetrical viral particles.

The influence of dental amalgam and heavy metal cations on interleukin1β IL1β expression by peripheral blood mononuclear cells from healthy donors was studied. A marked decrease in the production of IL1β was caused by freshly prepared amalgam or amalgamconditioned culture medium, but not by amalgam aged for 6 weeks. When metal cations were added as salts, Cu2, Hg2, and Agat high concentrations 33.3 and 333.3 μM were highly inhibitory. Among other heavy metal cations, Au3, Pt4, Ni2, Pd2, but not Ga3or Sn2, inhibited IL1β production in a concentrationdependent manner. Flow cytometry studies indicated that Hg2and Agstrongly reduced the percentage of CD14cells containing IL1β intracellularly. As shown by Northern blot analysis, Hg2inhibited the level of IL1βspecific mRNA by 28 at 3.3 μMand completely at 33.3 μM. Only slight inhibitory effects were induced by Cu2at 33.3 μM. Interestingly, Agat a concentration of 3.3 μMincreased twofold the amount of IL1βspecific mRNA. Our data show that IL1β production is altered at protein and mRNA levels by components released from fresh amalgam and by other heavy metal cations, suggesting a role of these cations in changes in the cell phenotype and IL1mediated cell functions. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res, 51, 88–95, 2000.

Purified faba bean necrotic yellows virus (FBNYV; genus Nanovirus) alone is not transmissible by its aphid vector, Acyrthosiphon pisum, regardless of whether it is acquired from artificial diets or directly microinjected into the aphid's hemocoel. The purified virus contains all of the genetic information required for its infection cycle as it readily replicated in cowpea protoplasts and systemically infected Vicia fabaseedlings that were biolistically inoculated using gold particles coated with intact virions or viral DNA. The bombarded plants not only developed the typical disease syndrome, thus indicating that FBNYV is the sole causal agent of the disease, but also served as a source from which the virus was readily acquired and transmitted by A. pisum. The defect of the purified virus in aphid transmissibility suggests that FBNYV requires a helper factor (HF) for its vector transmission that is either nonfunctional or absent in purified virus suspensions. The requirement for an HF was confirmed in complementation experiments using two distinct isolates of the virus. These experiments revealed that aphids transmitted the purified virus isolate from artificial diets only when they had fed previously on plants infected with the other FBNYV isolate. Also, microinjected FBNYV, which persisted to the same extent in A. pisumas naturally acquired virus, was transmissible when aphids had acquired the HF from infected plants. This suggests that one of the functions of the HF in the transmission process is to facilitate virus transport across the hemocoel–salivary gland interface.

The effects of dental amalgam on cytokine production by human peripheral blood mononuclear cells (PBMC) from healthy donors were analyzed. To induce cytokine production, PBMC were stimulated with lipopolysaccharide, phytohemagglutinin, or staphylococcal enterotoxin A and cultured for 48 h in the presence of either freshly prepared amalgam, aged amalgam, or amalgam-conditioned culture medium (ACCM). The concentrations of several cytokines were measured in PBMC supernatants by enzyme-amplified sensitivity immunoassays (EASIAs). Freshly prepared amalgam as well as ACCM induced a decrease in the production of interferon-? (IFN-?) and interleukin-10 (IL-10), and an increase in the concentrations of tumor necrosis factor-a (TNF-a). Both fresh amalgam and ACCM showed no effects on IL-2, IL-6, or granulocyte-macrophage colony-stimulating factor levels. Amalgam aged for 6 weeks did not affect the concentration of any of the above cytokines. To investigate which heavy metal cations released from amalgam caused the observed immunomodulatory effects, Cu2+, Hg2+, and Sn2+, which were detected in amalgam supernatants by inductively coupled plasma atomic spectrophotometry, were added as salts to the cultures. Cu2+and Hg2+induced a decrease in IFN-? and IL-10 levels, and Hg2+an increase in TNF-a concentrations. Cytokine production was not significantly modulated by Sn2+. Under these experimental conditions, release of Ag+into culture medium was not detectable. However, Ag+markedly suppressed the production of IFN-?, IL-10, and TNF-a. In summary, our results show that fresh amalgam, but not amalgam aged for 6 weeks, causes changes in the cytokine pattern of PBMC in vitro, and that these effects are due to the release of Cu2+and Hg2+. © 1998 John Wiley & Sons, Inc. J Biomed Mater Res, 42, 76–84, 1998.