BackgroundHerpes simplex 1 (HSV-1) causes various human clinical manifestations, ranging from simple cold sores to encephalitis. Innate immune cells recognize pathogens through Toll-like receptors (TLRs), thus initiating the immune response. Previously, we demonstrated that the immune response against HSV-1 is dependent on TLR2 and TLR9 expression and on IFN gamma production in the trigeminal ganglia (TG) of infected mice. In this work, we further investigated the cells, molecules, and mechanisms of HSV-1 infection control, especially those that are TLR-dependent.MethodsC57BL/6 wild-type (WT), TLR2−/−, TLR9−/−, and TLR2/9−/− mice were intranasally infected with HSV-1. On the viral peak day, the TG and brains were collected from mice and TLR expression was measured in the TG and brain and inducible nitric oxide synthase (iNOS) expression was measured in the TG by real-time PCR. Immunofluorescence assays were performed in mice TG to detect iNOS production by F4/80+ cells. Intraperitoneal macrophages nitric oxide (NO) production was evaluated by the Griess assay. WT, CD8−/−, RAG−/−, and iNOS−/− mice were intranasally infected in a survival assay, and their cytokine expression was measured in the TG by real-time PCR.ResultsInfected WT mice exhibited significantly increased TLR expression, compared with their respective controls, in the TG but not in the brain. TLR-deficient mice had moderately increased TLR expression in the TG and brain in compare with the non-infected animals. iNOS expression in the WT infected mice TG was higher than in the other groups with increased production by macrophages in the WT infected mice, which did not occur in the TLR2/9−/− mice. Additionally, the intraperitoneal macrophages of the WT mice had a higher production of NO compared with those of the TLR-deficient mice. The CD8−/−, RAG−/−, and iNOS−/− mice had 100% mortality after the HSV-1 infection compared with 10% of the WT mice. Cytokines were overexpressed in the iNOS−/− infected mice, while the RAG−/− mice were nearly unresponsive to the virus.ConclusionTLRs efficiently orchestrate the innate immune cells, eliciting macrophage response (with NO production by the macrophages), thereby controlling the HSV-1 infection through the immune response in the TG of mice.

A non-transgenic approach based on RNA interference was employed to induce protection against tomato mosaic virus (ToMV) infection in tomato plants. dsRNA molecules targeting the cp gene of ToMV were topically applied on plants prior to virus inoculation. Protection was dose-dependent and sequence-specific. While no protection was achieved when 0–16 µg dsRNA were used, maximum rates of resistance (60 and 63%) were observed in doses of 200 and 400 µg/plant, respectively. Similar rates were also obtained against potato virus Y when targeting its cp gene. The protection was quickly activated upon dsRNA application and lasted for up to 4 days. In contrast, no detectable antiviral response was triggered by the dsRNA from a begomovirus genome, suggesting the method is not effective against phloem-limited DNA viruses. Deep sequencing was performed to analyze the biogenesis of siRNA populations. Although long-dsRNA remained in the treated leaves for at least 10 days, its systemic movement was not observed. Conversely, dsRNA-derived siRNA populations (mainly 21- and 22-nt) were detected in non-treated leaves, which indicates endogenous processing and transport through the plant. Altogether, this study provides critical information for the development of novel tools against plant viruses; strengths and limitations inherent to the systems are discussed.

BackgroundHerpes simplex virus type 1 (HSV-1) cause not only mild symptoms but also blindness and encephalitis. It was previously shown that the immune response against HSV-1 occurs mainly in the trigeminal ganglia (TG) and that Toll-like receptors 2 and 9 (TLR2/9) are important in mediating this response. It was also demonstrated that iNOS (nitric oxide synthase) and interleukin 1 beta (IL-1β) play an essential role in the defense against HSV-1 infection. Importantly, the present work aimed to identify the primary cells responsible for iNOS and IL-1β production and search for other important molecules and cells that might or might not depend on TLR2/9 receptors to mediate the immune response against HSV-1.MethodsC57BL/6 (wild type, WT) and TLR2/9−/− mice were infected by the intranasal route with HSV-1 (1 × 106 p.f.u.). Cells were obtained from the TG and spleen tissues and the profile of immune cells was determined by flow cytometry in infected and mock infected WT and knockout mice. The percentage of cells producing iNOS, IL-1β, granzyme B and perforin was also determined by flow cytometry. Chemokine monocyte chemoattractant protein-1 (MCP1) was measured by Cytometric Bead Array (CBA) in the TG, spleen and lung. Expression of type I interferons (IFNs), interleukins (IL) 5 and 10, IL-1β and granzyme B were quantified by real time PCR.ResultsThe results indicate that dendritic cells (DCs) and monocytes/macrophages (Mo/Mϕ) were the main sources of IL-1β and iNOS, respectively, which, together with type I IFNs, were essential for the immune response against HSV-1. Additionally, we showed that granzyme B produced by CD8+ T and NK lymphocytes and MCP-1 were also important for this immune response. Moreover, our data indicate that the robust production of MCP-1 and granzyme B is either TLR-independent or down regulated by TLRs and occurs in the TG of TLR2/9−/− infected mice.ConclusionTaken together, our data provide strong evidence that the responses mediated by DCs, Mo/Mϕ, NK and CD8+ T lymphocytes through IL-1β, iNOS and granzyme B production, respectively, together with the production of type I IFN early in the infection, are crucial to host defense against HSV-1.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-017-0692-x) contains supplementary material, which is available to authorized users.

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Tomato mottle virus (ToMoV) is a single-stranded DNA (ssDNA) begomovirus transmitted to solanaceous crops by the whitefly species complex (Bemisia tabaci), causing stunted growth, leaf mottling, and reduced yield. Using a genetic repertoire of seven genes, ToMoV pathogenesis includes the manipulation of multiple plant biological processes to circumvent antiviral defenses. To further understand the effects of whitefly feeding and whitefly-transmitted ToMoV infection on tomato plants (Solanum lycopersicum ‘Florida Lanai’), we generated comprehensive protein profiles of leaves subjected to feeding by either viruliferous whiteflies harboring ToMoV, or non-viruliferous whiteflies, or a no-feeding control. The effects of whitefly feeding and ToMoV infection were measured both locally and systemically by sampling either a mature leaf directly from the site of clip-cage confined whitefly feeding, or from a newly formed leaf 10 days post feeding (dpf). At 3 dpf, tomato’s response to ToMoV included proteins associated with translation initiation and elongation as well as plasmodesmata dynamics. In contrast, systemic impacts of ToMoV on younger leaves 10 dpf were more pronounced and included a virus-specific change in plant proteins associated with mRNA maturation and export, RNA-dependent DNA methylation, and other antiviral plant processes. Our analysis supports previous findings and provides novel insight into tomato’s local and systemic response to whitefly feeding and ToMoV infection.

Arracacha mottle virus (AMoV) is the only potyvirus reported to infect arracacha (Arracacia xanthorrhiza) in Brazil. Here, the complete genome sequence of an isolate of AMoV was determined to be 9,630 nucleotides in length, excluding the 3' poly-A tail, and encoding a polyprotein of 3,135 amino acids and a putative P3N-PIPO protein. Its genomic organization is typical of a member of the genus Potyvirus, containing all conserved motifs. Its full genome sequence shared 56.2 % nucleotide identity with sunflower chlorotic mottle virus and verbena virus Y, the most closely related viruses.

Brugmansia suaveolens mottle virus (BsMoV) was the first potyvirus isolated from "angel trumpet" (Brugmansia suaveolens), described in Brazil. In this study, the complete nucleotide (nt) genome sequence of BsMoV was determined, and the deduced amino acid (aa) sequence was analyzed. The BsMoV RNA genome consists of 9870 nt without a poly-A tail, encoding a putative typical potyviral polyprotein of 3090 aa. Pairwise comparisons of the complete BsMoV genome with those of the most closely related potyviruses revealed a maximum nucleotide identity of 63.7% with pepper mottle virus. These results and phylogenetic analyses based on the complete genome sequence of the most closely related potyviruses confirmed that BsMoV should be considered a member of a distinct species of the genus Potyvirus.

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