Table of Contents

Abstract: Dengue has become the most important arboviral infection worldwide with more than 30 million cases of dengue fever estimated to occur each year. The need for a dengue vaccine is great and several live attenuated dengue candidate vaccines are proceeding through clinical evaluation. The need to induce a balanced immune response against all four DENV serotypes with a single vaccine has been a challenge for dengue vaccine developers. A live attenuated DENV chimeric vaccine produced by Sanofi Pasteur has recently entered Phase III evaluation in numerous dengue-endemic regions of the world. Viral interference between serotypes contained in live vaccines has required up to three doses of the vaccine be given over a 12-month period of time. For this reason, novel DENV candidate vaccines are being developed with the goal of achieving a protective immune response with an immunization schedule that can be given over the course of a few months. These next-generation candidates include DNA vaccines, recombinant adenovirus vectored vaccines, alphavirus replicons, and sub-unit protein vaccines. Several of these novel candidates will be discussed.

Abstract: Human T-cell leukemia virus type-1 (HTLV-1) is the etiological agent of adult T-cell leukemia (ATL), an aggressive and highly chemoresistant malignancy. Rho family GTPases regulate multiple signaling pathways in tumorigenesis: cytoskeletal organization, transcription, cell cycle progression, and cell proliferation. Geranylgeranylation of Rho family GTPases is essential for cell membrane localization and activation of these proteins. It is currently unknown whether HTLV-1-transformed cells are preferentially sensitive to geranylgeranylation inhibitors, such as GGTI-298. In this report, we demonstrate that GGTI-298 decreased cell viability and induced G₂/M phase accumulation of HTLV-1-transformed cells, independent of p53 reactivation. HTLV-1-LTR transcriptional activity was inhibited and Tax protein levels decreased following treatment with GGTI-298. Furthermore, GGTI-298 decreased activation of NF-κB, a downstream target of Rho family GTPases. These studies suggest that protein geranylgeranylation contributes to dysregulation of cell survival pathways in HTLV-1-transformed cells.

Abstract: Although no physical fossils of viruses have been found, retroviruses are known to leave their molecular fossils in the genomes of their hosts, the so-called endogenous retroviral elements. These have provided us with important information about retroviruses in the past and their co-evolution with their hosts. On the other hand, because non‑retroviral viruses were considered not to leave such fossils, even the existence of prehistoric non-retroviral viruses has been enigmatic. Recently, we discovered that elements derived from ancient bornaviruses, non-segmented, negative strand RNA viruses, are found in the genomes of several mammalian species, including humans. In addition, at approximately the same time, several endogenous elements of RNA viruses, DNA viruses and reverse-transcribing DNA viruses have been independently reported, which revealed that non-retroviral viruses have played significant roles in the evolution of their hosts and provided novel insights into virology and cell biology. Here we review non-retroviral virus-like elements in vertebrate genomes, non-retroviral integration and the knowledge obtained from these endogenous non-retroviral virus-like elements.

Abstract: Insects are commonly infected with multiple viruses including those that cause sublethal, asymptomatic, and latent infections. Traditional methods for virus isolation typically lack the sensitivity required for detection of such viruses that are present at low abundance. In this respect, next generation sequencing technologies have revolutionized methods for the discovery and identification of new viruses from insects. Here we review both traditional and modern methods for virus discovery, and outline analysis of transcriptome and small RNA data for identification of viral sequences. We will introduce methods for de novo assembly of viral sequences, identification of potential viral sequences from BLAST data, and bioinformatics for generating full-length or near full-length viral genome sequences. We will also discuss implications of the ubiquity of viruses in insects and in insect cell lines. All of the methods described in this article can also apply to the discovery of viruses in other organisms.

Abstract: One of the major obstacles that must be overcome in the design of effective lentiviral vaccines is the ability of lentiviruses to evolve in order to escape from neutralizing antibodies. The primary target for neutralizing antibodies is the highly variable viral envelope glycoprotein (Env), a glycoprotein that is essential for viral entry and comprises both variable and conserved regions. As a result of the complex trimeric nature of Env, there is steric hindrance of conserved epitopes required for receptor binding so that these are not accessible to antibodies. Instead, the humoral response is targeted towards decoy immunodominant epitopes on variable domains such as the third hypervariable loop (V3) of Env. For feline immunodeficiency virus (FIV), as well as the related human immunodeficiency virus-1 (HIV-1), little is known about the factors that lead to the development of broadly neutralizing antibodies. In cats infected with FIV and patients infected with HIV-1, only rarely are plasma samples found that contain antibodies capable of neutralizing isolates from other clades. In this review we examine the neutralizing response to FIV, comparing and contrasting with the response to HIV. We ask whether broadly neutralizing antibodies are induced by FIV infection and discuss the comparative value of studies of neutralizing antibodies in FIV infection for the development of more effective vaccine strategies against lentiviral infections in general, including HIV-1.

Abstract: We developed a feline model of lentiviral cross-species transmission using a puma lentivirus (PLV or FIV_Pco) which infects domestic cats but does not cause disease. Infection with PLV protects cats from CD4+ T-cell decline caused by subsequent infection with virulent feline immunodeficiency virus (FIV). Previous studies implicate innate immune and/or cellular restriction mechanisms for FIV disease attenuation in PLV-infected cats. In this study, we evaluated viral infection and cytokine mRNA transcription in 12 different tissue reservoirs approximately five months post infection. We quantitated tissue proviral load, viral mRNA load and relative transcription of IL-10, IL-12p40 and IFNγ from tissues of cats exposed to FIV, PLV or both viruses and analyzed these parameters using a multivariate statistical approach. The distribution and intensity of FIV infection and IFNγ transcription differed between single and co-infected cats, characterized by higher FIV proviral loads and IFNγ expression in co-infected cat tissues. Variability in FIV mRNA load and IFNγ was significantly more constrained in co-infected versus singly infected cat tissues. Single-infected:co-infected ratios of FIV mRNA load compared to FIV proviral load indicated that active viral transcription was apparently inhibited during co-infection. These results indicate that previous PLV infection increases activation of tissue innate immunity and constrains the ability of FIV to productively infect tissue reservoirs of infection for months, independent of FIV proviral load, supporting a model in which innate immunity and/or modulation of target cell susceptibility play a key role in PLV-induced protection from FIV disease.

Abstract: Despite the induction of effective immune responses, 80% of hepatitis C virus (HCV)-infected individuals progress from acute to chronic hepatitis. In contrast to the cellular immune response, the role of the humoral immune response in HCV clearance is still subject to debate. Indeed, HCV escapes neutralizing antibodies in chronically infected patients and reinfection has been described in human and chimpanzee. Studies of antibody-mediated HCV neutralization have long been hampered by the lack of cell-culture-derived virus and the absence of a small animal model. However, the development of surrogate models and recent progress in HCV propagation in vitro now enable robust neutralization assays to be performed. These advances are beginning to shed some light on the mechanisms of HCV neutralization. This review summarizes the current state of knowledge of the viral targets of anti-HCV-neutralizing antibodies and the mechanisms that enable HCV to evade the humoral immune response. The recent description of the HCV glycan shield that reduces the immunogenicity of envelope proteins and masks conserved neutralizing epitopes at their surface constitutes the major focus of this review.

Abstract: All life must survive their corresponding viruses. Thus antiviral systems are essential in all living organisms. Remnants of virus derived information are also found in all life forms but have historically been considered mostly as junk DNA. However, such virus derived information can strongly affect host susceptibility to viruses. In this review, I evaluate the role viruses have had in the origin and evolution of host antiviral systems. From Archaea through bacteria and from simple to complex eukaryotes I trace the viral components that became essential elements of antiviral immunity. I conclude with a reexamination of the ‘Big Bang’ theory for the emergence of the adaptive immune system in vertebrates by horizontal transfer and note how viruses could have and did provide crucial and coordinated features.

Abstract: Frog virus 3 (FV3) is the best characterized member of the family Iridoviridae. FV3 study has provided insights into the replication of other family members, and has served as a model of viral transcription, genome replication, and virus-mediated host-shutoff. Although the broad outlines of FV3 replication have been elucidated, the precise roles of most viral proteins remain unknown. Current studies using knock down (KD) mediated by antisense morpholino oligonucleotides (asMO) and small, interfering RNAs (siRNA), knock out (KO) following replacement of the targeted gene with a selectable marker by homologous recombination, ectopic viral gene expression, and recombinant viral proteins have enabled researchers to systematically ascertain replicative- and virulence-related gene functions. In addition, the application of molecular tools to ecological studies is providing novel ways for field biologists to identify potential pathogens, quantify infections, and trace the evolution of ecologically important viral species. In this review, we summarize current studies using not only FV3, but also other iridoviruses infecting ectotherms. As described below, general principles ascertained using FV3 served as a model for the family, and studies utilizing other ranaviruses and megalocytiviruses have confirmed and extended our understanding of iridovirus replication. Collectively, these and future efforts will elucidate molecular events in viral replication, intrinsic and extrinsic factors that contribute to disease outbreaks, and the role of the host immune system in protection from disease.

Abstract: Lentiviruses are known for their narrow cell- and species-tropisms, which are determined by cellular proteins whose absence or presence either support viral replication (dependency factors, cofactors) or inhibit viral replication (restriction factors). Similar to Human immunodeficiency virus type 1 (HIV-1), the cat lentivirus Feline immunodeficiency virus (FIV) is sensitive to recently discovered cellular restriction factors from non-host species that are able to stop viruses from replicating. Of particular importance are the cellular proteins APOBEC3, TRIM5α and tetherin/BST-2. In general, lentiviruses counteract or escape their species’ own variant of the restriction factor, but are targeted by the orthologous proteins of distantly related species. Most of the knowledge regarding lentiviral restriction factors has been obtained in the HIV-1 system; however, much less is known about their effects on other lentiviruses. We describe here the molecular mechanisms that explain how FIV maintains its replication in feline cells, but is largely prevented from cross-species infections by cellular restriction factors.

Abstract: Hepatitis C virus (HCV) is a Flavivirus with a positive-sense, single-stranded RNA genome of about 9,600 nucleotides. It is a major cause of liver disease, infecting almost 200 million people all over the world. Similarly to most RNA viruses, HCV displays very high levels of genetic diversity which have been used to differentiate six major genotypes and about 80 subtypes. Although the different genotypes and subtypes share basic biological and pathogenic features they differ in clinical outcomes, response to treatment and epidemiology. The first HCV recombinant strain, in which different genome segments derived from parentals of different genotypes, was described in St. Petersburg (Russia) in 2002. Since then, there have been only a few more than a dozen reports including descriptions of HCV recombinants at all levels: between genotypes, between subtypes of the same genotype and even between strains of the same subtype. Here, we review the literature considering the reasons underlying the difficulties for unequivocally establishing recombination in this virus along with the analytical methods necessary to do it. Finally, we analyze the potential consequences, especially in clinical practice, of HCV recombination in light of the coming new therapeutic approaches against this virus.