Viruses — Open Access Journal

Rabies is a fatal zoonotic disease transmitted by the bite of a rabid animal. More than 95% of the human rabies cases in India are attributed to exposure to rabid dogs. This study evaluated the utility of a lateral flow immunochromatographic assay (LFA) (Anigen Rapid Rabies Ag Test Kit, Bionote, Hwaseong-si, Korea) for rapid post mortem diagnosis of rabies in dogs. Brain tissue was collected from 202 animals that were screened through the Government of Goa rabies surveillance system. Thebrain tissue samples were obtained from 188 dogs, nine cats, three bovines, one jackal and one monkey. In addition, 10 dogs that died due to trauma from road accidents were included as negative controls for the study. The diagnostic performance of LFA was evaluated using results from direct fluorescence antibody test (dFT); the current gold standard post mortem test for rabies infection. Three samples were removed from the analysis as they were autolysed and not fit for testing by dFT. Of the 209 samples tested, 117 tested positive by LFA and 92 tested negative, while 121 tested positive by dFT and 88 tested negative. Estimates of LFA sensitivity and specificity were 0.96 (95% CI 0.91–0.99) and 0.99 (95% CI 0.94–1.00), respectively. The LFA is a simple and low-cost assay that aids in the rapid diagnosis of rabies in the field without the need for expensive laboratory equipment or technical expertise. This study found that Bionote LFA has potential as a screening tool in rabies endemic countries. Full article

To understand the mechanism of replication used by baculoviruses, it is essential to describe all the factors involved, including virus and host proteins and the sequences where DNA synthesis starts. A lot of work on this topic has been done, but there is still confusion in defining what sequence/s act in such functions, and the mechanism of replication is not very well understood. In this work, we performed an AgMNPV replication kinetics into the susceptible UFL-Ag-286 cells to estimate viral genome synthesis rates. We found that the viral DNA exponentially increases in two different phases that are temporally separated by an interval of 5 h, probably suggesting the occurrence of two different mechanisms of replication. Then, we prepared a plasmid library containing virus fragments (0.5–2 kbp), which were transfected and infected with AgMNPV in UFL-Ag-286 cells. We identified 12 virus fragments which acted as origins of replication (ORI). Those fragments are in close proximity to core genes. This association to the core genome would ensure vertical transmission of ORIs. We also predict the presence of common structures on those fragments that probably recruit the replication machinery, a structure also present in previously reported ORIs in baculoviruses. Full article

Knowledge on haemostatic changes in humans infected with Ebola virus is limited due to safety concerns and access to patient samples. Ethical approval was obtained to collect plasma samples from patients in Sierra Leone infected with Ebola virus over time and samples were analysed for clotting time, fibrinogen, and D-dimer levels. Plasma from healthy volunteers was also collected by two methods to determine effect of centrifugation on test results as blood collected in Sierra Leone was not centrifuged. Collecting plasma without centrifugation only affected D-dimer values. Patients with Ebola virus disease had higher PT and APTT and D-dimer values than healthy humans with plasma collected in the same manner. Fibrinogen levels in patients with Ebola virus disease were normal or lower than values measured in healthy people. Clotting times and D-dimer levels were elevated during infection with Ebola virus but return to normal over time in patients that survived and therefore could be considered prognostic. Informative data can be obtained from plasma collected without centrifugation which could improve patient monitoring in hazardous environments. Full article

Avian orthoavulavirus13 (AOAV-13), also named avian paramyxovirus 13 (APMV-13), has been found sporadically in wild birds around the world ever since the discovery of AOAV-13 (AOAV-13/wild goose/Shimane/67/2000) in a wild goose from Japan in 2000. However, there are no reports of AOAV-13 in China. In the present study, a novel AOAV-13 virus (AOAV-13/wild goose/China/Hubei/V93-1/2015), isolated from a wild migratory waterfowl in a wetland of Hubei province of China, during active surveillance from 2013 to 2018, was biologically and genetically characterized. Phylogenetic analyses demonstrated a very close genetic relationship among all AOAV-13 strains, as revealed by very few genetic variations. Moreover, pathogenicity tests indicated that the V93-1 strain is a low virulent virus for chickens. However, the genome of the V93-1 virus was found to be 16,158 nucleotides (nt) in length, which is 12 nt or 162 nt longer than the other AOAV-13 strains that have been reported to date. The length difference of 12 nt in strain V93-1 is due to the existence of three repeats of the conserved sequence, “AAAAAT”, in the 5′-end trailer of the genome. Moreover, the HN gene of the V93-1 virus is 2,070 nt in size, encoding 610 aa, which is the same size as the AOAV-13 strain from Japan, whereas that of two strains from Ukraine and Kazakhstan are 2,080 nt in length, encoding 579 aa. We describe a novel AOAV-13 in migratory waterfowl in China, which suggests that diversified trailer region sequences and HN gene lengths exist within serotype AOAV-13, and highlight the need for its constant surveillance in poultry from live animal markets, and especially migratory birds. Full article

Andes virus (ANDV) and Sin Nombre virus (SNV) are the main causative agents responsible for hantavirus cardiopulmonary syndrome (HCPS) in the Americas. HCPS is a severe respiratory disease with a high fatality rate for which there are no approved therapeutics or vaccines available. Some vaccine approaches for HCPS have been tested in preclinical models, but none have been tested in infectious models in regard to their ability to protect against multiple species of HCPS-causing viruses. Here, we utilize recombinant vesicular stomatitis virus-based (VSV) vaccines for Andes virus (ANDV) and Sin Nombre virus (SNV) and assess their ability to provide cross-protection in infectious challenge models. We show that, while both rVSVΔG/ANDVGPC and rVSVΔG/SNVGPC display attenuated growth as compared to wild type VSV, each vaccine is able to induce a cross-reactive antibody response. Both vaccines protected against both homologous and heterologous challenge with ANDV and SNV and prevented HCPS in a lethal ANDV challenge model. This study provides evidence that the development of a single vaccine against HCPS-causing hantaviruses could provide protection against multiple agents. Full article

The begomoviruses (BGVs) are plant pathogens that evolved in the Old World during the Cretaceous and arrived to the New World (NW) in the Cenozoic era. A subgroup of NW BGVs, the “Squash leaf curl virus (SLCV) lineage” (S-Lin), includes viruses with unique characteristics. To get clues on the evolutionary origin of this lineage, a search for divergent members was undertaken. Four novel BGVs were characterized, including one that is basal to the group. Comparative analyses led to discover a ~670 bp genome module that is nearly exclusive of this lineage, encompassing the replication origin, the AC4 gene, and 480 bp of the Rep gene. A similar DNA module was found in two curtoviruses, hence suggesting that the S-Lin ancestor acquired its distinctive genomic segment by recombination with a curtovirus. This hypothesis was definitely disproved by an in-depth sequence analysis. The search for homologs of S-Lin Rep uncover the common origin of Rep proteins encoded by diverse Geminiviridae genera and viral “fossils” integrated at plant genomes. In contrast, no homolog of S-Lin Rep was found in public databases. Consequently, it was concluded that the SLCV clade ancestor evolved by a recombination event between a primitive NW BGV and a virus from a hitherto unknown lineage. Full article

Arthropod-borne viruses (arboviruses) of vertebrates including dengue, zika, chikungunya, Rift Valley fever, and blue tongue viruses cause extensive morbidity and mortality in humans, agricultural animals, and wildlife across the globe. As obligate intercellular pathogens, arboviruses must be well adapted to the cellular and molecular environment of both their arthropod (invertebrate) and vertebrate hosts, which are vastly different due to hundreds of millions of years of separate evolution. Here we discuss the comparative pressures on arbovirus RNA genomes as a result of a dual host life cycle, focusing on pressures that do not alter amino acids. We summarize what is currently known about arboviral genetic composition, such as dinucleotide and codon usage, and how cyclical infection of vertebrate and invertebrate hosts results in different genetic profiles compared with single-host viruses. To serve as a comparison, we compile what is known about arthropod tRNA, dinucleotide, and codon usages and compare this with vertebrates. Additionally, we discuss the potential roles of genetic robustness in arboviral evolution and how it may vary from other viruses. Overall, both arthropod and vertebrate hosts influence the resulting genetic composition of arboviruses, but a great deal remains to be investigated. Full article

Hantaviruses are transmitted by rodents producing the hantavirus pulmonary syndrome (HPS) in the Americas. Today, no human cases of HPS have been reported in Mexico in spite of similar environmental conditions with Central America and the USA where several cases have occurred. To understand the current situation of hantaviruses in Mexico and the public health risk, a systematic review of studies was conducted reporting hantaviruses in rodents to known state seroprevalence and hantavirus genotypes. Simultaneously, this study identified the potential hantaviruses based on the phylogenetic diversity (PD) of hantaviruses reported in the Americas in hosts with the distribution in Mexico. A total 3862 rodents belonging to 82 species have been tested since 1999 to 2017. Overall, 392 individuals representing 43 rodent species were seropositive, and the seroprevalence ranged from 0 to 69.22%. Seven hantaviruses genotypes have been described in Mexico and three are zoonotic. Four host species of rodents are widely distributed in Mexico harboring the highest PD of viruses. According to the hosts distribution, 16 genotypes could be circulating in Mexico and some of these represent a potential risk for public health. This study proposed multidisciplinary and interinstitutional collaborations to implement systematic surveillance in rodents. Full article

A recent study reported the discovery of an endogenous reptilian foamy virus (FV), termed ERV-Spuma-Spu, found in the genome of tuatara. Here, we report two novel reptilian foamy viruses also identified as endogenous FVs (EFVs) in the genomes of panther gecko (ERV-Spuma-Ppi) and Schlegel’s Japanese gecko (ERV-Spuma-Gja). Their presence indicates that FVs are capable of infecting reptiles in addition to mammals, amphibians, and fish. Numerous copies of full length ERV-Spuma-Spu elements were found in the tuatara genome littered with in-frame stop codons and transposable elements, suggesting that they are indeed endogenous and are not functional. ERV-Spuma-Ppi and ERV-Spuma-Gja, on the other hand, consist solely of a foamy virus-like env gene. Examination of host flanking sequences revealed that they are orthologous, and despite being more than 96 million years old, their env reading frames are fully coding competent with evidence for strong purifying selection to maintain expression and for them likely being transcriptionally active. These make them the oldest EFVs discovered thus far and the first documented EFVs that may have been co-opted for potential cellular functions. Phylogenetic analyses revealed a complex virus–host co-evolutionary history and cross-species transmission routes of ancient FVs. Full article

The Usutu virus (USUV) is a flavivirus that is drawing increasing attention because of its potential for emergence. First isolated in Africa, it was introduced into Europe where it caused significant outbreaks in birds, such as in Austria in 2001. Since then, its geographical distribution has rapidly expanded, with increased circulation, especially in the last few years. Similar to West Nile virus (WNV), the USUV enzootic transmission cycle involves Culex mosquitoes as vectors, and birds as amplifying reservoir hosts, with humans and other mammals likely being dead-end hosts. A similarity in the ecology of these two viruses, which co-circulate in several European countries, highlights USUV’s potential to become an important human pathogen. While USUV has had a severe impact on the blackbird population, the number of human cases remains low, with most infections being asymptomatic. However, some rare cases of neurological disease have been described, both in healthy and immuno-compromised patients. Here, we will discuss the transmission dynamics and the current state of USUV circulation in Europe. Full article

Monitoring West Nile virus (WNV) and Usutu virus (USUV) activity now has the highest priority among mosquito-borne pathogenic viruses circulating in the European Union. This study documents a first time detection and the co-circulation of WNV lineage-2 (with the minimal prevalence of 0.46%) and USUV clade Europe 2 (with the minimal prevalence of 0.25%) in mosquitoes from the same habitat of south-western Slovakia and underlines necessity to perform rigorous surveillance in birds, mosquitoes, horses and humans in that country. Full article

Noroviruses, in the genus Norovirus, are a significant cause of viral gastroenteritis in humans and animals. For almost 50 years, the lack of a cultivation system for human noroviruses (HuNoVs) was a major barrier to understanding virus biology and the development of effective antiviral strategies. This review presents a historical perspective of the development of a cultivation system for HuNoVs in human intestinal epithelial cell cultures. Successful cultivation was based on the discovery of genetically-encoded host factors required for infection, knowledge of the site of infection in humans, and advances in the cultivation of human intestinal epithelial cells achieved by developmental and stem cell biologists. The human stem cell-derived enteroid cultivation system recapitulates the multicellular, physiologically active human intestinal epithelium, and allows studies of virus-specific replication requirements, evaluation of human host-pathogen interactions, and supports the pre-clinical assessment of methods to prevent and treat HuNoV infections. Full article

Orthohantaviruses are tightly linked to the ecology and evolutionary history of their mammalian hosts. We hypothesized that in regions with dramatic climate shifts throughout the Quaternary, orthohantavirus diversity and evolution are shaped by dynamic host responses to environmental change through processes such as host isolation, host switching, and reassortment. Jemez Springs virus (JMSV), an orthohantavirus harbored by the dusky shrew (Sorex monticola) and five close relatives distributed widely in western North America, was used to test this hypothesis. Total RNAs, extracted from liver or lung tissue from 164 shrews collected from western North America during 1983–2007, were analyzed for orthohantavirus RNA by reverse transcription polymerase chain reaction (RT-PCR). Phylogenies inferred from the L-, M-, and S-segment sequences of 30 JMSV strains were compared with host mitochondrial cytochrome b. Viral clades largely corresponded to host clades, which were primarily structured by geography and were consistent with hypothesized post-glacial expansion. Despite an overall congruence between host and viral gene phylogenies at deeper scales, phylogenetic signals were recovered that also suggested a complex pattern of host switching and at least one reassortment event in the evolutionary history of JMSV. A fundamental understanding of how orthohantaviruses respond to periods of host population expansion, contraction, and secondary host contact is the key to establishing a framework for both more comprehensive understanding of orthohantavirus evolutionary dynamics and broader insights into host–pathogen systems. Full article

Measles vaccines have been in use since the 1960s with excellent safety and effectiveness profiles. Limited data are available on detection of measles vaccine virus (MeVV) RNA in human subjects following vaccination. Available evidence suggests MeVV RNA can be identified up to 14 days after vaccination, with detection beyond this rare. In routine diagnostic testing, we used two real-time reverse transcription-polymerase chain reaction (RT-rPCR) assays targeting M and F genes to identify measles virus (MeV) and MeVV RNA. Confirmatory testing was performed with an N gene RT-rPCR, followed by sequence confirmation of RT-rPCR positives by semi-nested conventional RT-PCR assays targeting portions of the N, H, and L genes. We report detection and confirmation of MeVV RNA from the respiratory tract of 11 children between 100 and 800 days after most recent receipt of measles-containing vaccine. These novel findings emphasize the importance of genotyping all MeV detections and highlight the need for further work to assess whether persistent MeVV RNA represents viable virus and if transmission to close contacts can occur. Full article

Infectious laryngotracheitis (ILT) is an acute respiratory disease of poultry caused by infectious laryngotracheitis virus (ILTV). Control of the disease with live attenuated vaccines administered via eye drop build upon immune responses generated by the eye-associated lymphoid tissues. The aim of this study was to assess cytokine and lymphocyte changes in the conjunctiva-associated lymphoid tissues (CALT) and Harderian gland (HG) stimulated by the ocular inoculation of the ILTV chicken embryo origin (CEO) vaccine strain and virulent strain 63140. This study offers strong evidence to support the roles that the CALT and HG play in the development of protective ILTV immune responses. It supports the premise that ILTV-mediated immunomodulation favors the B cell response over those of T cells. Further, it provides evidence that expansions of CD8α⁺ cells, with the concomitant expression of the Granzyme A gene, are key to reducing viral genomes in the CALT and halting ILTV cytolytic replication in the conjunctiva. Ultimately, this study revealed that the early upregulation of interleukin (IL)-12p40 and Interferon (IFN)-γ cytokine genes, which shape the antigen-specific cell-mediated immune responses, retarded the decline of virus replication, and enhanced the development of lesions in the conjunctiva epithelium. Full article

We previously generated a recombinant reporter Akabane virus expressing enhanced green fluorescence protein (eGFP-AKAV), with an artificial S genome encoding eGFP in the ambisense RNA. Although the eGFP-AKAV was able to detect infected cells in in vivo histopathological study, its fluorescent signal was too weak to apply to in vivo imaging study. Here, we successfully generated a modified reporter, eGFP/38-AKAV, with 38-nucleotide deletion of the internal region of the 5′ untranslated region of S RNA. The eGFP/38-AKAV expressed higher intensity of eGFP fluorescence both in vitro and in vivo than the original eGFP-AKAV did. In addition, eGFP/38-AKAV was pathogenic in mice at a comparable level to that in wild-type AKAV. In the mice infected with eGFP/38-AKAV, the fluorescent signals, i.e., the virus-infected cells, were detected in the central nervous system using the whole-organ imaging. Our findings indicate that eGFP/38-AKAV could be used as a powerful tool to help elucidate the dynamics of AKAV in vivo. Full article

Reassortment contributes to the evolution of RNA viruses with segmented genomes, including Bluetongue virus (BTV). Recently, co-circulation of natural and vaccine BTV variants in Europe, and their ensuing reassortment, were proposed to promote appearance of novel European BTV strains, with potential implications for pathogenicity, spread and vaccination policies. Similarly, the geographical features of the Mediterranean basin, which spans over portions of three continents, may facilitate the appearance of clinically relevant reassortants via co-circulation of BTV strains of African, Asian and European origins. In August–October 2017, BTV serotype 6 (BTV-6) was identified in young animals exhibiting classical clinical signs of Bluetongue (BT) at Israeli sheep and cattle farms. Sequencing and pairwise analysis of this Israeli BTV-6 isolate revealed the closest sequence homology of its serotype-defining Segment 2 was with that of South African reference BTV-6 strain 5011 (93.88% identity). In contrast, the other viral segments showed highest homology (97.0%–99.47% identity) with BTV-3, -4 and -9 of Mediterranean and African origins. Specifically, four viral segments were nearly identical (99.13%–99.47%), with Tunisian and Italian BTV-3 strains (TUN2016 and SAD2018, correspondingly). Together, our data suggest that Mediterranean co-circulation and reassortment of BTV-3 and BTV-6 drove the emergence of a novel and virulent BTV-6 strain Full article

Viroporins are a family of small hydrophobic proteins found in many enveloped viruses that are capable of ion transport. Building upon the ability to inhibit influenza by blocking its archetypical M2 H⁺ channel, as a family, viroporins may represent a viable target to curb viral infectivity. To this end, using three bacterial assays we analyzed six small hydrophobic proteins from biomedically important viruses as potential viroporin candidates. Our results indicate that Eastern equine encephalitis virus 6k, West Nile virus MgM, Dengue virus 2k, Dengue virus P1, Variola virus gp170, and Variola virus gp151 proteins all exhibit channel activity in the bacterial assays, and as such may be considered viroporin candidates. It is clear that more studies, such as patch clamping, will be needed to characterize the ionic conductivities of these proteins. However, our approach presents a rapid procedure to analyze open reading frames in other viruses, yielding new viroporin candidates for future detailed investigation. Finally, if conductivity is proven vital to their cognate viruses, the bio-assays presented herein afford a simple approach to screen for new channel blockers. Full article

With the availability of an increasing number of 3D structures of bacteriophage components, combined with powerful in silico predictive tools, it has become possible to decipher the structural assembly and functionality of phage adhesion devices. In the current study, we examined 113 members of the 936 group of lactococcal siphophages, and identified a number of Carbohydrate Binding Modules (CBMs) in the neck passage structure and major tail protein, on top of evolved Dit proteins, as recently reported by us. The binding ability of such CBM-containing proteins was assessed through the construction of green fluorescent protein fusion proteins and subsequent binding assays. Two CBMs, one from the phage tail and another from the neck, demonstrated definite binding to their phage-specific host. Bioinformatic analysis of the structural proteins of 936 phages reveals that they incorporate binding modules which exhibit structural homology to those found in other lactococcal phage groups and beyond, indicating that phages utilize common structural “bricks” to enhance host binding capabilities. The omnipresence of CBMs in Siphophages supports their beneficial role in the infection process, as they can be combined in various ways to form appendages with different shapes and functionalities, ensuring their success in host detection in their respective ecological niches. Full article