Virus-Host Interactions

Dalam beberapa dekade terakhir, vaksinasi menjadi istilah umum untuk pemaparan antigen terhadap manusia atau binatang dalam membangkitkan respon kekebalan. Vaksin potensial merupakan syarat utama untuk tujuan ini sehingga dapat mengontrol penyakit secara efektif. Kebanyakan vaksin virus yang digunakan saat ini merupakan sel utuh yang telah dilemahkan atau dimatikan. Keuntungan vaksin ini pada umumnya mampu menghasilkan imunitas cukup lama dan merangsang seluruh reaksi kekebalan pada host yaitu Humoral antibody dan Cell-Mediated. Cara konvensional ini mampu mengeliminasi berbagai penyakit, namun masih banyak penyakit infeksi dan parasit yang belum dapat diatasi dengan vaksinasi, karena belum dapat dibuat vaksinnya.
Berkembangnya bioteknologi terutama rekombinasi DNA akhir-akhir ini telah membuka harapan-harapan baru. Teknologi ini memungkinkan memproduksi vaksin yang saat ini belum dapat dibuat. Selain itu teknologi ini juga dapat dimanfaatkan untuk memperbaiki vaksin yang sudah ada, sehingga didapatkan vaksin yang lebih aman dan efektif. Inovasi teknologi ini memungkinkan pembuatan vaksin sub unit dan vaksin idiotipe (vaksin terapi).
Oleh karena itu, kami menggagas konsep Nyamuk sebagai Vaccine Delivery Host : Sebuah Pemikiran Futuristik sebagai upaya perlindungan pencegahan dan pengendalian yang efektif sebagai solusi menghadapi krisis wabah penyakit infeksi dan parasit yang terjadi di Indonesia. Konsep ini memanfaatkan vektor nyamuk untuk produksi antigen melalui teknik DNA rekombinan. Dimana antigen dari mikrobia parasit tertentu diklon dalam plasmid bakteri yang direkayasa untuk meningkatkan ekspresi gen yang diinsersikan di dalam sel-sel tubuh nyamuk vektor, sehingga pada saat menggigit nyamuk tidak menularkan penyakit tetapi justru meningkatkan respon kekebalan terhadap suatu penyakit.
Tujuan konsep ini adalah selain sebagai upaya perlindungan dan pengendalian terhadap wabah penyakit infeksi dan parasit di daerah endemis penyakit, hal ini juga menjawab respon atas krisis kesehatan pada kondisi kontemporer.

The ongoing pandemic of severe acute respiratory syndrome (SARS), caused by the SARS- CoV-2 human coronavirus (HCoV), has brought the international scientific community before a state of emergency that needs to be addressed with intensive research for the discovery of pharmacological agents with antiviral activity. Potential antiviral natural products (NPs) have been discovered from plants of the global biodiversity, including extracts, compounds and categories of compounds with activity against several viruses of the respiratory tract such as HCoVs. However, the scarcity of natural products (NPs) and small-molecules (SMs) used as antiviral agents, especially for HCoVs, is notable. This is a review of 203 publications, which were selected using PubMed/MEDLINE, Web of Science, Scopus, and Google Scholar, evaluates the available literature since the discovery of the first human coronavirus in the 1960s; it summarizes important aspects of structure, function, and therapeutic targeting of HCoVs as well as NPs (19 total plant extracts and 204 isolated or semi- synthesized pure compounds) with anti-HCoV activity targeting viral and non-viral proteins, while focusing on the advances on the discovery of NPs with anti-SARS-CoV-2 activity, and providing a critical perspective.

Okra leaf curl disease (OLCD) is an important disease responsible for yield losses in okra in all okra growing regions in the world. Effective management of the OLCD is very important in order to improve yield. The study was conducted to evaluate the effectiveness of different botanicals in the management of the OLCD. The treatments applied were 10% (w/v) crude extract each of neem leaf, garlic, mahogany bark, chili pepper fruit, pawpaw dried leaf, bougainvillea leaf and the control (water), in a randomised complete block design with four replications. The plant extracts significantly reduced the population of whitefly vector, as well as the incidences and severities of OLCD infections and increased fruit yield compared to the control (P< 0.05). Neem leaf extract followed by garlic extract significantly (P< 0.05) reduced the populations of whitefly and the incidences and severities of OLCD but significantly increased ((P < 0.05) fruit yield compared to crude extracts of mahogany, bougainvillea, chili pepper, and pawpaw leaves. The population of whitefly was positively correlated with severity of OLCD (r = 0.679; P> 0.05). Fruit yield was also negatively correlated with the severity of OLCD (r = -0.857; P< 0.05) and whitefly population (r = -0.750; P< 0.05). Farmers can manage OLCD in their okra fields by spraying with neem leaf or garlic extracts in order to improve fruit yields.

The COVID-19 pandemic has strengthened the interest in the biological mechanisms underlying the complex interplay between infectious agents and the human host. The spectrum of phenotypes associated with the SARS-CoV-2 infection, ranging from the absence of symptoms to severe systemic complications, raised the question as to what extent the variable response to coronaviruses (CoVs) is influenced by the variability of the hosts' genetic background. To explore the current knowledge about this question, we designed a systematic review encompassing the scientific

The oral cavity is a persistent reservoir for Epstein-Barr virus (EBV) with lifelong infection of resident epithelial and B cells. Infection of these cell types results in distinct EBV gene expression patterns regulated by epigenetic modifications involving DNA methylation and chromatin structure. Regulation of EBV gene expression relies on viral manipulation of the host epigenetic machinery that may result in long-lasting host epigenetic reprogramming. To identify epigenetic events following EBV infection, a transient infection model was established to map epigenetic changes in telomerase-immortalized oral keratinocytes. EBV-infected oral keratinocytes exhibited a predominantly latent viral gene expression program with some lytic or abortive replication. Calcium and methylcellulose-induced differentiation was delayed in EBV-positive clones and in clones that lost EBV compared to uninfected controls, indicating a functional consequence of EBV epigenetic modifications. Analysis of global cellular DNA methylation identified over 13,000 differentially methylated CpG residues in cells exposed to EBV compared to uninfected controls, with CpG island hypermethylation observed at several cellular genes. Although the vast majority of the DNA methylation changes were silent, 65 cellular genes that acquired CpG methylation showed altered transcript levels. Genes with increased transcript levels frequently acquired DNA methylation within the gene body while those with decreased transcript levels acquired DNA methylation near the transcription start site. Treatment with the DNA methyltransferase inhibitor, decitabine, restored expression of some hypermethylated genes in EBV-infected and EBV-negative transiently infected clones. Overall, these observations suggested that EBV infection of keratinocytes leaves a lasting epigenetic imprint that can enhance the tumorigenic phenotype of infected cells. IMPORTANCE Here, we show that EBV infection of oral keratinocytes led to CpG island hypermethylation as an epigenetic scar of prior EBV infection that was retained after loss of the virus. Such EBV-induced epigenetic modification recapitulated the hypermethylated CpG island methylator phenotype (CIMP) observed in EBV-associated carcinomas. These epigenetic alterations not only im-pacted gene expression but also resulted in delayed calcium and methylcellulose-induced keratinocyte differentiation. Importantly , these epigenetic changes occurred in cells that were not as genetically unstable as carcinoma cells, indicating that EBV infection induced an epigenetic mutator phenotype. The impact of this work is that we have provided a mechanistic framework for how a tumor virus using the epigenetic machinery can act in a " hit-and-run " fashion, with retention of epigenetic alterations after loss of the virus. Unlike genetic alterations, these virally induced epigenetic changes can be reversed pharmacologically, providing therapeutic interventions to EBV-associated malignancies.

In Kaposi’s sarcoma-associated herpesvirus (KSHV) oncogenesis, both latency and reactivation are hypothesized to potentiate tumor growth. The KSHV Rta protein is the lytic switch for reactivation. Rta transactivates essential genes via interactions with cofactors such as the cellular RBP-Jk and Oct-1 proteins, and the viral Mta protein. Given that robust viral reactivation would facilitate antiviral responses and culminate in host cell lysis, regulation of Rta’s expression and function is a major determinant of the latent-lytic balance and the fate of infected cells. Our lab recently showed that Rta transactivation requires the cellular peptidyl-prolyl cis/trans isomerase Pin1. Our data suggest that proline‑directed phosphorylation regulates Rta by licensing binding to Pin1. Despite Pin1’s ability to stimulate Rta transactivation, unchecked Pin1 activity inhibited virus production. Dysregulation of Pin1 is implicated in human cancers, and KSHV is the latest virus known to co-opt Pin1 function. We propose that Pin1 is a molecular timer that can regulate the balance between viral lytic gene expression and host cell lysis. Intriguing scenarios for Pin1’s underlying activities, and the potential broader significance for isomerization of Rta and reactivation, are highlighted.

Ich präsentiere knapp und verständlich neuere Ergebnisse der Evolutionsforschung zu Symbiogenese und lateralem Gentransfer und schlage eine neue Metapher für die Netz-Struktur der Evolution vor. Ich betrachte kurz einige wissenschaftsgeschichtliche und epistemologische Aspekte der Evolutionstheorie und öffne Perspektiven hin auf ein neues, umfassendes Szenario der Evolution, das u.a. eine  symbiogenetische, polyphyletische Abstammung der Metazoa in Betracht zieht. Dies ist ein populärwissenschaftlich geschriebener Artikel, der eigene wissenschaftstheoretische Konzepte anreisst und weitere Veröffentlichungen vorbereitet.

Viruses produce nucleic acids during their replication, either during genomic replication or transcription. These nucleic acids are present in the cytoplasm or endosome of an infected cell, or in the extracellular space to be sensed by neighboring cells during lytic infections. Cells have mechanisms of sensing virus-generated nucleic acids; these nucleic acids act as flags to the cell, indicating an infection requiring defense mechanisms. The viral nucleic acids are called pathogen-associated molecular patterns (PAMPs) and the sensors that bind them are called pattern recognition receptors (PRRs). This review article focuses on the most recent findings regarding nucleic acids PRRs in fish, including: Toll-like receptors (TLRs), RIG-I-like receptors (RLRs), cytoplasmic DNA sensors (CDSs) and class A scavenger receptors (SR-As). It also discusses what is currently known of the downstream signaling molecules for each PRR family and the resulting antiviral response, either type I interferons (IFNs) or pro-inflammatory cytokine production. The review highlights what is known but also defines what still requires elucidation in this economically important animal. Understanding innate immune systems to virus infections will aid in the development of better antiviral therapies and vaccines for the future.

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Persistent infection with cancer risk-related viruses leads to molecular, cellular and immune response changes in host organisms that in some cases direct cellular transformation. Alternative splicing is a conserved cellular process that increases the coding complexity of genomes at the pre-mRNA processing stage. Human and other animal tumour viruses use alternative splicing as a process to maximize their transcriptomes and proteomes. Medical therapeutics to clear persistent viral infections are still limited. However, specific lessons learned in some viruses [e.g. HIV and HCV (hepatitis C virus)] suggest that drug-directed inhibition of alternative splicing could be useful for this purpose. The present review describes the basic mechanisms of constitutive and alternative splicing in a cellular context and known splicing patterns and the mechanisms by which these might be achieved for the major human infective tumour viruses. The roles of splicing-related proteins expressed by these viruses in cellular and viral gene regulation are explored. Moreover, we discuss some currently available drugs targeting SR (serine/arginine-rich) proteins that are the main regulators of constitutive and alternative splicing, and their potential use in treatment for so-called persistent viral infections.

Lytic infection with the two human gammaherpesviruses, Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV), leads to significant depletion of the cellular transcriptome. This host shutoff phenotype is driven by the conserved herpesviral alkaline exonuclease, termed SOX in KSHV and BGLF5 in EBV, which in gammaherpesviruses has evolved the genetically separable ability to target cellular mRNA. We now show that host shutoff is also a prominent consequence of murine gammaherpesvirus 68 (MHV68) infection, which is widely used as a model system to study pathogenesis of these viruses in vivo. The effector of MHV68-induced host shutoff is its SOX homolog, here termed muSOX. There is remarkable functional conservation of muSOX host shutoff activities with those of KSHV SOX, including the recently described ability of SOX to induce mRNA hyperadenylation in the nucleus as well as cause nuclear relocalization of the poly(A) binding protein. SOX and muSOX localize to both the nucleus and cytoplasm of infected cells. Using spatially restricted variants of these proteins, we go on to demonstrate that all known host shutoff-related activities of SOX and muSOX are orchestrated exclusively from the cytoplasm. These results have important mechanistic implications for how SOX and muSOX target nascent cellular transcripts in the nucleus. Furthermore, our findings establish MHV68 as a new, genetically tractable model to study host shutoff.

ABSTRACT: The present study compares 13 physiological and immunological variables between
a group of healthy Panulirus argus lobsters and a group of lobsters naturally infected with Panulirus
argus Virus 1 (PaV1). Viral infection was determined through histopathology and PCR. Ten
of the 13 variables differed significantly between the 2 groups. Using these variables, a principal
component analysis yielded 2 separate clusters: one corresponding to the healthy group and the
other corresponding to the infected group. In particular, infected lobsters exhibited significantly
lower levels of osmotic pressure, total hemocyte counts, plasmatic proteins, and total phenoloxidase
(PO) activity in plasma, as well as significantly higher levels of cholesterol and acylglycerides.
These features are consistent with metabolic wasting, hyperlipidemia, and presumed
immune suppression. Infection with PaV1 appears to increase the susceptibility of lobsters to some
other opportunistic pathogens, as 61.1% of infected lobsters presented infestations of ciliate
epibionts (Epystilis and Zoothamniun) in the gill chamber compared with 11.5% lobsters in the
healthy group. Infected lobsters also showed significantly higher levels of total PO activity in
degranulated hemocytes and trypsin inhibitor activity, potentially indicating activation of immune
response by the PO system during the systemic infection with PaV1.

the upper respiratory tract of pigs (H. Kida et al., J. Gen. Virol. 75:2183-2188, 1994). It was shown that A/chicken/Yamaguchi/7/2004 (H5N1) [Ck/Yamaguchi/04 (H5N1)] did not replicate in pigs (N. Isoda et al., Arch. Virol. 151:1267-1279, 2006).

The dynamics of a general in-host model with intracellular delay is studied. The model can describe in vivo infections of HIV-I, HCV, and HBV. It can also be considered as a model for HTLV-I infection. We derive the basic reproduction number R 0 for the viral infection, and establish that the global dynamics are completely determined by the values of R 0 . If R 0 ≤ 1, the infection-free equilibrium is globally asymptotically stable, and the virus are cleared. If R 0 > 1, then the infection persists and the chronic-infection equilibrium is locally asymptotically stable. Furthermore, using the method of Lyapunov functional, we prove that the chronic-infection equilibrium is globally asymptotically stable when R 0 > 1. Our results shows that for intercellular delays to generate sustained oscillations in in-host models it is necessary have a logistic mitosis term in target-cell compartments.

Imperata yellow mottle caused by Imperata yellow mottle virus (IYMV) of the genus Sobemovirus was first characterized on Imperata cylindrica and Zea mays in Burkina Faso. The disease has been reported in several localities of the country but its ecology and epidemiology are poorly known. In particular, only I. cylindrica and maize have been reported within IYMV host range. The aim of this study was to investigate the experimental host range of the virus. Mechanical inoculation of a mixture of four IYMV isolates to 18 plant species, including four crops (maize, rice, sorghum and pearl millet) and 14 wild grasses showed clear mottle symptoms in maize, sorghum and pearl millet and two wild grass species (Setaria verticillata and Rottboellia exaltata). Symptom development was confirmed by Enzyme-linked immunosorbent assay and reverse transcription–polymerase chain reaction (RT-PCR). Infection of crop species by IYMV depended on cultivars tested. Therefore, proportions of positive cultivars were 16/36 in maize, 4/10 in sorghum and 4/9 in pearl millet , respectively. Studies on virus-host interactions using individual virus isolates showed two pathogenic patterns. Three out of the four isolates tested infected all plant species and cultivars. In contrast, the fourth IYMV isolate could infect only one maize cultivar. These results expand the previously known host range of IYMV from two to five species, indicating a narrow host range. Among the new characterized host species, sorghum and pearl millet are important cereal crops. Therefore, Imperata yellow motte disease is a potential threat for the cereal crop production and its ecology and epidemiology should be thoroughly investigated.

We have shown that Macrobrachium rosenbergii nodavirus (MrNV) was able to infect Sf9 cells and that MrNV virus-like particles (MrNV-VLPs) were capable nanocontainers for delivering nucleic acid-based materials. Here, we demonstrated that chymotryptic removal of a C-terminal peptide and its truncated variant (F344-MrNV-VLPs) exhibited a drastically reduced ability to interact and internalize into Sf9 cells. Electron microscopic observations revealed that the loss of C-terminal domain either from enzyme hydrolysis or genetic truncation did not affect the generated MrNV-VLPs' icosahedral conformation, but did drastically affect the VLPs' internalization ability into Sf9 cells. Homology-based modelling of the MrNV capsid with other icosahedral capsid models revealed that this chymotrypsin-sensitive C-terminal domain was not only exposed on the capsid surface, but also constituted the core of the viral capsid protrusion. These results therefore suggest the importance of the C-terminal domain as a structure for targeted cell interaction which is presumably localized at the protruding domain. This work thus provided the functional insights into the role of the MrNV C-terminal domain in viral entry into Sf9 cells and lead to the development of strategies in combatting MrNV infection in susceptible cells.

Filoviruses are highly lethal in humans and nonhuman primates, likely due to potent antagonism of host interferon (IFN) responses early in infection. Filoviral protein VP35 is implicated as the major IFN induction antagonist, while Ebola virus (EBOV) VP24 or Marburg virus (MARV) VP40 are known to block downstream IFN signaling. Despite progress elucidating EBOV and MARV antagonist function, those for most other filoviruses, including Reston (RESTV), Sudan (SUDV), Taï Forest (TAFV), Bundibugyo (BDBV) and Ravn (RAVV) viruses, remain largely neglected. Thus, using standardized vectors and reporter assays, we characterized activities by each IFN antagonist from all known ebolavirus and marburgvirus species side-by-side. We uncover noncanonical suppression of IFN induction by ebolavirus VP24, differing potencies by MARV and RAVV proteins, and intriguingly, weaker antagonism by VP24 of RESTV. These underlying molecular explanations for differential virulence in humans could guide future investigations of more-neglected filoviruses as well as treatment and vaccine studies.

We describe a novel spike pseudoparticle neutralisation
assay (ppNT) for seroepidemiological studies on
Middle East respiratory syndrome coronavirus (MERSCoV)
and apply this assay together with conventional
microneutralisation (MN) tests to investigate 1,343
human and 625 animal sera. The sera were collected
in Egypt as a region adjacent to areas where MERS has
been described, and in Hong Kong, China as a control
region. Sera from dromedary camels had a high prevalence
of antibody reactive to MERS-CoV by MERS NT
(93.6%) and MERS ppNT (98.2%) assay. The antibody
titres ranged up to 1,280 and higher in MN assays
and 10,240 and higher in ppNT assays. No other
investigated species had any antibody reactivity to
MERS-CoV. While seropositivity does not exclude the
possibility of infection with a closely related virus, our
data highlight the need to attempt detection of MERSCoV
or related coronaviruses in dromedary camels. The
data show excellent correlation between the conventional
MN assay and the novel ppNT assay. The newly
developed ppNT assay does not require Biosafety Level
3 containment and is thus a relatively high-throughput
assay, well suited for large-scale seroepidemiology
studies which are needed to better understand the
ecology and epidemiology of MERS-CoV.

Terza guerra mondiale È oramai evidente: siamo nel bel mezzo della terza guerra mondiale. Tutto il pianeta è investito dall'ondata di virus che sta sconvolgendo il precedente ritmo di vita nel quale, più o meno male (parlare di bene sarebbe un insulto alla quasi totalità dell'umanità) eravamo, se non abituati, comunque immersi. Gli effetti devastanti che osserviamo sono esattamente quelli che potevano essere descritti in una simulazione di "guerra batteriologica". Prendere atto che siamo in guerra, indipendentemente dalla sua causa, è un punto di partenza necessario. Occorre averne piena consapevolezza. Solo così, con lucidità e approccio razionale, possiamo valutare modi e tempi sul come affrontarla....
Non rinunciare a vivere qui ed ora, al meglio di noi stessi, pregando e indirizzando energie positive a coloro che sono in prima linea.
È ciò che ha fatto un Maestro, Ananda Bhavanani, un Guru indiano, dedicando espressamente questo messaggio: Mahamrityunjaya contemplation in the times of COVID19
https://www.youtube.com/watch?v=juXsEh2QAxo&feature=youtu.be

In this study we demonstrated that Macrobrachium rosenbergii nodavirus (MrNV) was able to internalize and replicate in Sf9 insect cells, with levels of infection altered by substances affecting the caveolin-(CAV) mediated endocytosis pathway. The use of Sf9 cells for efficient MrNV replication and propagation was demonstrated by confocal microscopy and PCR amplification, through which early viral binding and internalization were initially detectable at 30 min post-infection; whereas at 72 h, the distinguishable sign of late-MrNV infection was observable as the gradual accumulation of a cytopathic effect (CPE) in the cells, ultimately resulting in cellular disruption. Moreover, during the early period of infection, the MrNV signals were highly co-localized with CAV1 signals of the CAV-mediated endocytosis pathway. The use of genistein as an inhibitor of the CAV-mediated endocytosis pathway significantly reduced MrNV and CAV1 co-localization, and also reduced the levels of MrNV infection in Sf9 cells as shown by PCR and ELISA. Moreover, the addition of the pathway agonist okadaic acid not only recovered but also augmented both the levels of MrNV co-localization with CAV1 and of Sf9 infection in the presence of genistein inhibition; therefore demonstrating that MrNV infection in Sf9 cells was associated with the CAV-mediated endocytosis pathway machinery.

Alcelaphine herpesvirus–1 (AlHV-1), a causative agent of malignant catarrhal fever in cattle, was detected in wildebeest (Connochaetes taurinus) placenta tissue for the first time. Although viral load was low, the finding of viral DNA in over 50% of 94 samples tested lends support to the possibility that placental tissue could play a role in disease transmission and that wildebeest calves are infected in utero. Two viral loci were sequenced to examine variation among virus samples obtained from wildebeest and cattle: the ORF50 gene, encoding the lytic cycle transactivator protein, and the A9.5 gene, encoding a novel polymorphic viral glyco-protein. ORF50 was well conserved with six newly discovered alleles differing at only one or two base positions. In contrast, while only three new A9.5 alleles were discovered, these differed by up to 13% at the nucleotide level and up to 20% at the amino acid level. Structural ho-mology searching performed with the additional A9.5 sequences determined in this study adds power to recent analysis identifying the four-helix bundle cytokine interleukin-4 (IL4) as the major homologue. The majority of MCF virus samples obtained from Tanzanian cattle and wildebeest encoded A9.5 polypeptides identical to the previously characterized A9.5 allele present in the laboratory maintained AlHV-1 C500 strain. This supports the view that AlHV-1 C500 is suitable for the development of a vaccine for wildebeest-associated MCF.

Hantaviruses are primarily rodent-borne pathogens which have received considerable attention recently due to their high mortality rates in humans. In order to find the causes of rapid transmission and emergence of hantavirus-associated diseases anthropogenic changes are a priority. These include deforestation, urbanization, noise pollution, light pollution and electromagnetic fields, all of which have been shown to profoundly affect rodent physiology and immunology. Moreover, anthropogenic events promote human-rodent co-habitation and thereby provide a driver to increase rates of transmission and, by extrapolation, levels of infection in humans. Such environmental disruption acts as a chronic stressor to rodents and causes elevated concentrations of glucocorticoids, which are a major class of immunosuppressive hormone. Glucocorticoids are responsible for altering the immune tolerance of rodents, thereby rendering them susceptible to infection. Glucocorticoids induce regulatory T lymphocytes to reduce inflammatory and antiviral responses and to activate regulatory responses, principally through production of the cytokines interleukin-10 and transforming growth factor-β to support viral persistence. In order to develop a low-cost intervention strategy for hantavirus infection consideration should be given to a systemic approach to therapy. This would both aim to achieve a reduction of anthropogenic stressors and to gain a greater understanding of host-pathogen interactions.

Viruses are extremely heterogeneous entities; the size and the nature of their genetic information, as well as the strategies employed to amplify and propagate their genomes, are highly variable. However, as obligatory intracellular parasites, replication of all viruses relies on the host cell. Having co-evolved with their host for several million years, viruses have developed very sophisticated strategies to hijack cellular factors that promote virus uptake, replication, and spread. Identification of host cell factors (HCFs) required for these processes is a major challenge for researchers, but it enables the identification of new, highly selective targets for anti viral therapeutics. To this end, the establishment of platforms enabling genome-wide high-throughput RNA interference (HT-RNAi) screens has led to the identification of several key factors involved in the viral life cycle. A number of genome-wide HT-RNAi screens have been performed for major human pathogens. These studies enable first inter-viral comparisons related to HCF requirements. Although several cellular functions appear to be uniformly required for the life cycle of most viruses tested (such as the proteasome and the Golgi-mediated secretory pathways), some factors, like the lipid kinase Phosphatidylinositol 4-kinase IIIα in the case of hepatitis C virus, are selectively required for individual viruses. However, despite the amount of data available, we are still far away from a comprehensive understanding of the interplay between viruses and host factors. Major limitations towards this goal are the low sensitivity and specificity of such screens, resulting in limited overlap between different screens performed with the same virus. This review focuses on how statistical and bioinformatic analysis methods applied to HT-RNAi screens can help overcoming these issues thus increasing the reliability and impact of such studies.

The process of conducting cell-based phenotypic screens can result in data sets from small libraries or portions of large libraries, making accurate hit picking from multiple data sets important for efficient drug discovery. Here, we describe a screen design and data analysis approach that allow for normalization not only between quadrants and plates but also between screens or batches in a robust, quantitative fashion, enabling hit selection from multiple data sets. We independently screened the MicroSource Spectrum and NCI Diversity Set II libraries using a cell-based phenotypic highthroughput screening (HTS) assay that uses an interferon-stimulated response element (ISRE)-driven luciferase-reporter assay to identify interferon (IFN) signal enhancers. Inclusion of a per-plate, per-quadrant IFN dose-response standard curve enabled conversion of ISRE activity to effective IFN concentrations. We identified 45 hits based on a combined z score ≥2.5 from the two libraries, and 25 of 35 available hits were validated in a compound concentration-response assay when tested using fresh compound. The results provide a basis for further analysis of chemical structure in relation to biological function. Together, the results establish an HTS method that can be extended to screening for any class of compounds that influence a quantifiable biological response for which a standard is available.

Japanese Encephalitis Virus (JEV), a globally important pathogen, belongs to the family Flaviviridae, is transmitted between vertebrate hosts by mosquitoes, principally by Culex tritaeniorhynchus. The E-glycoprotein of the virus mediates its attachment to the host cell receptors. In this study, we cloned and purified JEV E-glycoprotein in pET28a vector using E. coli BL21 (DE3) cells. A pull down assay was performed using plasma membrane fraction of BALB/c mouse brain and E-glycoprotein as a bait protein. 2-Dimensional Gel Electrophoresis based separation of the interacting proteins was analyzed by mass spectrometry. Among all the identified partners of E-glycoprotein, PLVAP (Plasmalemma vesicle associated protein) and GKN3 (Gastrokine3) showed significant up-regulation in both JEV infected mouse brain and neuro2a cells. In-silico studies also predicted significant interaction of these receptors with E-glycoprotein. Additionally, overexperssion and silencing of these receptors result...

The severe acute respiratory syndrome (SARS) epidemic started in late 2002 and swiftly spread across 5 continents with a mortality rate of around 10%. Although the epidemic was eventually controlled through the implementation of strict quarantine measures, there continues a need to investigate the SARS coronavirus (SARS-CoV) and develop interventions should it re-emerge. Numerous studies have shown that neutralizing antibodies against the virus can be found in patients infected with SARS-CoV within days upon the onset of illness and lasting up to several months. In contrast, there is little data on the kinetics of T cell responses during SARS-CoV infection and little is known about their role in the recovery process. However, recent studies in mice suggest the importance of T cells in viral clearance during SARS-CoV infection. Moreover, a growing number of studies have investigated the memory T cell responses in recovered SARS patients. This review covers the available literature on the emerging importance of T cell responses in SARS-CoV infection, particularly on the mapping of cytotoxic T lymphocyte (CTL) epitopes, longevity, polyfunctionality and human leukocyte antigen (HLA) association as well as their potential implications on treatment and vaccine development.

Background: The newly emerged Middle East respiratory syndrome coronavirus (MERS-CoV) that first appeared in Saudi Arabia during the summer of 2012 has to date (20th September 2013) caused 58 human deaths. MERS-CoV utilizes the dipeptidyl peptidase 4 (DPP4) host cell receptor, and analysis of the long-term interaction between virus and receptor provides key information on the evolutionary events that lead to the viral emergence. Findings: We show that bat DPP4 genes have been subject to significant adaptive evolution, suggestive of a long-term arms-race between bats and MERS related CoVs. In particular, we identify three positively selected residues in DPP4 that directly interact with the viral surface glycoprotein. Conclusions: Our study suggests that the evolutionary lineage leading to MERS-CoV may have circulated in bats for a substantial time period.

Research on the pathogenesis of asthma has concentrated on initial stimuli, genetic susceptibilities, adaptive immune responses, and end-organ alterations (particularly in airway mucous cells and smooth muscle) as critical steps leading to disease. Recent evidence indicates that the innate immune cell response to respiratory viruses also contributes to the development of inflammatory airway disease. We further develop this concept by raising the issue that the interaction between host airway epithelial cells and respiratory viruses is another aspect of innate immunity that is also a critical determinant of asthma. We also introduce a rationale for how antiviral performance at the epithelial cell level might be improved to prevent acute infectious illness and chronic inflammatory disease caused by respiratory viruses.

Background: Feline immunodeficiency virus (FIV) is a global pathogen of Felidae species and a model system for Human immunodeficiency virus (HIV)‑induced AIDS. In felids such as the domestic cat (Felis catus), APOBEC3 (A3) genes encode for single‑domain A3Z2s, A3Z3 and double‑domain A3Z2Z3 anti‑viral cytidine deaminases. The feline A3Z2Z3 is expressed following read‑through transcription and alternative splicing, introducing a previously untrans‑ lated exon in frame, encoding a domain insertion called linker. Only A3Z3 and A3Z2Z3 inhibit Vif‑deficient FIV. Feline A3s also are restriction factors for HIV and Simian immunodeficiency viruses (SIV). Surprisingly, HIV‑2/SIV Vifs can counteract feline A3Z2Z3. Results: To identify residues in feline A3s that Vifs need for interaction and degradation, chimeric human–feline A3s were tested. Here we describe the molecular direct interaction of feline A3s with Vif proteins from cat FIV and present the first structural A3 model locating these interaction regions. In the Z3 domain we have identified residues involved in binding of FIV Vif, and their mutation blocked Vif‑induced A3Z3 degradation. We further identified additional essen‑ tial residues for FIV Vif interaction in the A3Z2 domain, allowing the generation of FIV Vif resistant A3Z2Z3. Mutated feline A3s also showed resistance to the Vif of a lion‑specific FIV, indicating an evolutionary conserved Vif–A3 binding. Comparative modelling of feline A3Z2Z3 suggests that the residues interacting with FIV Vif have, unlike Vif‑interacting residues in human A3s, a unique location at the domain interface of Z2 and Z3 and that the linker forms a homeobox‑ like domain protruding of the Z2Z3 core. HIV‑2/SIV Vifs efficiently degrade feline A3Z2Z3 by possible targeting the linker stretch connecting both Z‑domains. Conclusions: Here we identified in feline A3s residues important for binding of FIV Vif and a unique protein domain insertion (linker). To understand Vif evolution, a structural model of the feline A3 was developed. Our results show that HIV Vif binds human A3s differently than FIV Vif feline A3s. The linker insertion is suggested to form a homeo‑box domain, which is unique to A3s of cats and related species, and not found in human and mouse A3s. Together, these findings indicate a specific and different A3 evolution in cats and human.

Self-replicating genetic material presumably provided the architecture necessary for generating the last universal ancestor of all nucleic-acid-based life. As biological complexity increased in the billions of years that followed, the same genetic material also morphed into a wide spectrum of viruses and other parasitic genetic elements. The resulting struggle for existence drove the evolution of host defenses, giving rise to a perpetual arms race. This Perspective summarizes the antiviral mechanisms evident across the tree of life, discussing each in their evolutionary context to postulate how the coevolution of host and pathogen shaped the cellular antiviral defenses we know today. Introduction The combination of time and adaptability has bestowed extraordinary diversity into all life. The molecular understanding of cellular processes, particularly with the recent advent of high throughput sequencing and computational biology, has provided us the means to use conservation to infer the evolutionary past. This technique has given us profound insight with regards to the origins of life and has led to significant advances in our understanding of the processes that drive its diversity. Arguably, one of the most powerful drivers of evolution stems from genetic parasites such as transposable elements and viruses (Koonin and Dolja, 2013). Their capacity to change, duplicate , edit, and/or transfer genetic information is often credited for the diversity of life. Despite the value provided by mobile genetic information, if left unchecked, this process would be catastrophic. It therefore follows that in addition to natural attenua-tion, the ability to successfully block expansion and spread of genetic pathogens is an essential attribute of host evolutionary survival. From prokaryotes to archaea to eukaryotes, every living species on the planet has evolved elaborate measures to recognize and react to viruses and/or transposable elements in some capacity. These defenses are composed of diverse strategies that often reflect the evolutionary trajectory that shaped each species, with some general themes proving to be timeless. Despite the mechanistic overlap of some antiviral defenses across both kingdoms and species, there is immense diversity in how they evolved and function (Figure 1). Moreover, specific evolutionary branches demonstrate the emergence of defense systems that are exclusive to a given lineage. Here the various antiviral pathways are summarized in their evolutionary context to highlight both common and unique features of each strategy as a means of postulating how they may have emerged. Although necessarily speculative, the evolutionary trajectory for each specific defense system is based on observed conservation and the principle of evolutionary parsimony. This concept states that the number of assumptions needed to generate a hypothesis should be inversely correlated to its likelihood. When applied to evolutionary biology, maximum parsimony equates to the idea that any biological component that can be found in all three domains of life (bacteria, archaea, and eukaryotes), is likely to have existed in some form in our last universal ancestor

Here, we address the question of why the influenza A virus hemagglutinin (HA) does not escape immunity by hyperglycosylation. Uniquely among dozens of monoclonal antibodies specific for A/Puerto Rico/8/34, escape from H28-A2 neutralization requires substitutions introducing N-linked glycosylation at residue 131 or 144 in the globular domain. This escape decreases viral binding to cellular receptors, which must be compensated for by additional substitutions in HA or neuraminidase that enable viral replication. Sequence analysis of circulating H1 influenza viruses confirms the in vivo relevance of our findings: natural occurrence of glycosylation at residue 131 is always accompanied by a compensatory mutation known to increase HA receptor avidity. In vaccinated mice challenged with WT vs. H28-A2 escape mutants, the selective advantage conferred by glycan-mediated global reduction in antigenicity is trumped by the costs of diminished receptor avidity. These findings show that, although...