Cytoplasmic Granule Formation and Translational Inhibition of Nodaviral RNAs in the Absence of the Double-Stranded RNA Binding Protein B2

Flock House virus (FHV) is a positive-sense RNA insect virus with a bipartite genome. RNA1 encodes the RNA-dependent RNA polymerase, and RNA2 encodes the capsid protein. A third protein, B2, is translated from a subgenomic RNA3 derived from the 3′ end of RNA1. B2 is a double-stranded RNA (dsRNA) binding protein that inhibits RNA silencing, a major antiviral defense pathway in insects. FHV is conveniently propagated in Drosophila melanogaster cells but can also be grown in mammalian cells. It was previously reported that B2 is dispensable for FHV RNA replication in BHK21 cells; therefore, we chose this cell line to generate a viral mutant that lacked the ability to produce B2. Consistent with published results, we found that RNA replication was indeed vigorous but the yield of progeny virus was negligible. Closer inspection revealed that infected cells contained very small amounts of coat protein despite an abundance of RNA2. B2 mutants that had reduced affinity for dsRNA produced analogous results, suggesting that the dsRNA binding capacity of B2 somehow played a role in coat protein synthesis. Using fluorescence in situ hybridization of FHV RNAs, we discovered that RNA2 is recruited into large cytoplasmic granules in the absence of B2, whereas the distribution of RNA1 remains largely unaffected. We conclude that B2, by binding to double-stranded regions in progeny RNA2, prevents recruitment of RNA2 into cellular structures, where it is translationally silenced. This represents a novel function of B2 that further contributes to successful completion of the nodaviral life cycle.     

Virus Factories of Cauliflower Mosaic Virus Are Virion Reservoirs That Engage Actively in Vector Transmission

Cauliflower mosaic virus (CaMV) forms two types of inclusion bodies within infected plant cells: numerous virus factories, which are the sites for viral replication and virion assembly, and a single transmission body (TB), which is specialized for virus transmission by aphid vectors. The TB reacts within seconds to aphid feeding on the host plant by total disruption and redistribution of its principal component, the viral transmission helper protein P2, onto microtubules throughout the cell. At the same time, virions also associate with microtubules. This redistribution of P2 and virions facilitates transmission and is reversible; the TB reforms within minutes after vector departure. Although some virions are present in the TB before disruption, their subsequent massive accumulation on the microtubule network suggests that they also are released from virus factories. Using drug treatments, mutant viruses, and exogenous supply of viral components to infected protoplasts, we show that virions can rapidly exit virus factories and, once in the cytoplasm, accumulate together with the helper protein P2 on the microtubule network. Moreover, we show that during reversion of this phenomenon, virions from the microtubule network can either be incorporated into the reverted TB or return to the virus factories. Our results suggest that CaMV factories are dynamic structures that participate in vector transmission by controlled release and uptake of virions during TB reaction.     

Serious adverse reactions associated with ivermectin: A systematic pharmacovigilance study in sub-Saharan Africa and in the rest of the World

BackgroundIvermectin is known to cause severe encephalopathies in subjects infected with loiasis, an endemic parasite in Sub-Saharan Africa (SSA). In addition, case reports have described ivermectin-related serious adverse drug reactions (sADRs) such as toxidermias, hepatic and renal disorders. The aim of this study was to identify suspected sADRs reported after ivermectin administration in VigiBase, the World Health Organization’s global individual case safety reports database and analyze their frequency relative to the frequency of these events after other antinematodal drugs reported in SSA and other areas of the world (ROW).MethodsAll antinematodal-related sADRs were extracted from VigiBase. Disproportionality analyses were conducted to investigate nervous, cutaneous, psychiatric, respiratory, renal, hepatic and cardiac suspected sADRs reported after ivermectin and benzimidazole drug administration across the world, in SSA and RoW.Principal findings2041 post-ivermectin or post-benzimidazole suspected sADRs were identified including 667 after ivermectin exposure (208 in SSA and 459 in the RoW). We found an increased reporting for toxidermias, encephalopathies, confusional disorders after ivermectin compared to benzimidazole drug administration. Encephalopathies were not only reported from SSA but also from the RoW (adjusted reporting odds ratios [aROR] 6.30, 95% confidence interval: 2.68–14.8), highlighting the fact these types of sADR occur outside loiasis endemic regions.ConclusionWe described for the first time suspected sADRs associated with ivermectin exposure according to geographical origin. While our results do not put in question ivermectin’s excellent safety profile, they show that as for all drugs, appropriate pharmacovigilance for adverse reactions is indicated.     

Antitumoral cyclic peptide analogues of chlamydocin

A series of cyclic tetrapeptides bearing the bioactive alkylating group on an ε-amino-lysyl function have been examined for their antitumoral activity on L1210 and P388 murine leukemia cell lines. One analogue belonging to the chlamydocin family and bearing a β-chloroethylnitrosourea group was found to be potent at inhibiting L1210 cell proliferation and had a higher therapeutic index than the reference compound bis-β-chloroethylnitrosourea (BCNU) on the in vivo P388-induced leukemia model.     

Expression of serotonin and enkephalins in calanoid copepods (Crustacea): an immunohistochemical study

Copepods are the main metazoan component of zooplankton, interms of both the number of species and biomass. Thus, theyhave a key role in pelagic food webs; but curiously, nothingis known of the neuroendocrine substances involved in theirphysiological processes. In higher crustaceans, especially theDecapoda, the role of such molecules in different physiologicalprocesses (motility, feeding, reproduction, day-night migrationsand so on) has been well explored; so, we have mapped expressionsites of some of these substances to provide a better understandingof copepod physiology. Serotonin and, for the first time, leucine-and methionine-enkephalin were detected in copepods using immunofluorescencetechniques. Serotonin has mainly been identified in the centralnervous system acting probably as a neurotransmitter, notablyin the control of the escape reflex. In contrast, enkephalinsare only present in peripheral organs such as the naupliar eye,gut and shell glands. This localization suggests that opioidsare involved in visual function and reproductive and digestiveprocesses. In memory of Professor Jacques Mazza, an eminent copepodologist.     

Biologically active oxidized lipids (phytoprostanes) in the plant diet and parenteral lipid nutrition

Phytoprostanes (PP) are autoxidation products of alpha-linolenate that are present in all plant tissues. Several classes of PP with a prostaglandin (PG) F1-, E1-, A1- and B1-like structure were identified and quantified by gas chromatography-mass spectrometry in vegetable oils and parenteral nutrition (intralipid). High levels of PP (0.09 up to 99 mg/l) were found even in apparently fresh vegetable oils. After oral consumption of olive or soybean oil, PPF1 were absorbed, found to circulate in plasma in conjugated form and excreted in free form into urine. Evidence is emerging that certain PP, such as the PPE1, may modulate the function of immune cells in a PG-like fashion. Here, we show that PPA1- and deoxy-PPJ1 display potent anti-inflammatory and apoptosis inducing activities similar to PGA1 and deoxy-PGJ2. Results of this study indicate that PP are novel, biologically active lipids in plant nutrition.     

The endogenous neurotransmitter, serotonin, modifies neuronal nitric oxide synthase activities

Serotonin, an important neurotransmitter, is colocalized with neuronal nitric oxide synthase (nNOS), a homodimeric enzyme which catalyzes the production of nitric oxide (NO(.-)) and/or oxygen species. As many interactions have been reported between the nitrergic and serotoninergic systems, we studied the effect of serotonin on nNOS activities. Our results reveal that nNOS is activated by serotonin as both NADPH consumption and oxyhemoglobin (OxyHb) oxidation were enhanced. The generation of L-citrulline from L-arginine (L-Arg) was not affected by serotonin in the range of 0-200 microM, suggesting an additional production of oxygen-derived species. But 5-hydroxytryptamine (5HT) induced the formation of both O and H(2)O(2) by nNOS, as evidenced by electron paramagnetic resonance (EPR) and by using specific spin traps. Overall, these results demonstrate that serotonin is able to activate nNOS, leading to the generation of reactive oxygen species (ROS) in addition to the NO(.-) production. Such a property must be considered in vivo as various nNOS-derived products mediate different signaling pathways.     

Selectivity for D-lactate incorporation into the peptidoglycan precursors of Lactobacillus plantarum: role of Aad, a VanX-like D-alanyl-D-alanine dipeptidase

Lactobacillus plantarum produces peptidoglycan precursors ending in D-lactate instead of D-alanine, making the bacterium intrinsically resistant to vancomycin. The ligase Ddl of L. plantarum plays a central role in this specificity by synthesizing D-alanyl-D-lactate depsipeptides that are added to the precursor peptide chain by the enzyme MurF. Here we show that L. plantarum also encodes a D-Ala-D-Ala dipeptidase, Aad, which eliminates D-alanyl-D-alanine dipeptides that are produced by the Ddl ligase, thereby preventing their incorporation into the precursors. Although D-alanine-ended precursors can be incorporated into the cell wall, inactivation of Aad failed to suppress growth defects of L. plantarum mutants deficient in d-lactate-ended precursor synthesis.