Animal Viruses Research Articles

Obituary

Obituary

Rabies in Europe in 2010-2019

The paper presents the epizootic and epidemiological situation of rabies in European countries during the last decade. The presented results indicate that the oral immunisation of fox anti-rabies (ORV), used in many European countries, significantly reduced the number of rabies cases found in ground mammals, but did not eliminate the virus at all. Currently, the largest reservoir of the virus are Eastern European countries where there are no immunisation activities or their effectiveness is low. Due to the absence of geographical barriers, the virus reappears in countries that have been described as free from rabies. As a rule, it is dragged into these areas along with the movement of companion animals and by people travelling to countries where the prevalence of the virus is common. It should be emphasised that due to the significant elimination of the virus in wild and domestic animals, it found quite quickly found a new reservoir in a specific group of mammals, having the ability to fly, like bats. Currently there is no possibility of carrying out any preventive measures in bats, so all the virus strains found in this group of animals are dangerous to humans, raising possibilities of epidemiological threat. It should be noted that despite the significant elimination of rabies in many European countries, given the almost unlimited possibilities of virus transmission to new areas, it still poses a serious threat to public health. Thus, it is necessary to constantly monitor the occurrence of the virus and possibly take preventive actions in terms of its elimination from the environment.

Alphavirus RNA replication in vertebrate cells.

Alphaviruses are positive-strand RNA viruses, typically transmitted by mosquitoes between vertebrate hosts. They encode four essential replication proteins, the non-structural proteins nsP1-4, which possess the enzymatic activities of RNA capping, RNA helicase, site-specific protease, ADP-ribosyl removal and RNA polymerase. Alphaviruses have been key models in the study of membrane-associated RNA replication, which is a conserved feature among the positive-strand RNA viruses of animals and plants. We review new structural and functional information on the nsPs and their interaction with host proteins and membranes, as well as with viral RNA sequences. The dodecameric ring structure of nsP1 is likely to be one of the evolutionary innovations that facilitated the success of the progenitors of current positive-strand RNA viruses.

Gain-of-Function and Pathogenic Viruses

Gain of function research is a little known and controversial area of study in virology. Though there is some disagreement in the exact definition of gain of function, the process involves modifying animal viruses to infect human cells for the purposes of advancing the understanding of the way viruses mutate. This article examines the controversy, with particular consideration of the COVID-19 lab leak hypothesis, exploring the danger of creating such modified viruses.

Second sialic acid-binding site of influenza A virus neuraminidase: binding receptors for efficient release.

Influenza A viruses (IAVs) are a major cause of human respiratory tract infections and cause significant disease and mortality. Human IAVs originate from animal viruses that breached the host species barrier. IAV particles contain sialoglycan receptor-binding hemagglutinin (HA) and receptor-destroying neuraminidase (NA) in their envelope. When IAV crosses the species barrier, the functional balance between HA and NA needs to be adjusted to the sialoglycan repertoire of the novel host species. Relatively little is known about the role of NA in host adaptation in contrast to the extensively studied HA. NA prevents virion aggregation and facilitates release of (newly assembled) virions from cell surfaces and from decoy receptors abundantly present in mucus and cell glycocalyx. In addition to a highly conserved catalytic site, NA carries a second sialic acid-binding site (2SBS). The 2SBS preferentially binds α2,3-linked sialic acids and enhances activity of the neighboring catalytic site by bringing/keeping multivalent substrates in close contact with this site. In this way, the 2SBS contributes to the HA-NA balance of virus particles and affects virus replication. The 2SBS is highly conserved in all NA subtypes of avian IAVs, with some notable exceptions associated with changes in the receptor-binding specificity of HA and host tropism. Conservation of the 2SBS is invariably lost in human (pandemic) viruses and in several other viruses adapted to mammalian host species. Preservation or loss of the 2SBS is likely to be an important factor of the viral host range.

Experimental infection of brown trout (Salmo trutta), zebrafi sh (Danio rerio), and swan mussel (Anodonta cygnea) with infectious pancreatic necrosis virus (IPVN)

Aim. Although more than 80 fi sh species and some invertebrate aquatic organisms are known to be susceptible to in- fectious pancreatic necrosis virus (IPNV) and aquatic birnaviruses, relatively little is known about its ability to infect brown trout (Salmo trutta), zebrafi sh (Danio rerio), and a mollusk, swan mussel (Anodonta cygnea). Therefore, the aim was to study in more detail the effects of IPNV on the said organisms. The information on the virulence of birnaviruses for aquatic organisms is important for better understanding of their transmission and for the identifi cation of suscep- tible or transient (symptomless) hosts. Methods. Cell culture method, using RTG-2 cells, was used for in vitro culti- vation of IPNV as well as virus titer assessment. The supernatants from infected RTG-2 cells were used for artifi cial inoculation to obtain experimental infection of the fi sh and mollusk. Fish and mollusk behaviour and mortality were monitored daily. The infectious titer of the virus in experimental animals was assessed by cell culture assay and reverse transcription PCR (RT-PCR). Results. This paper presents the results of IPNV experimental infection of brown trout (Salmo trutta), zebrafi sh (Danio rerio), and an aquatic invertebrate mollusk - swan mussel (Anodonta cygnea). The mortality of brown trout and zebrafi sh was 76.6 and 40 % respectively. Swan mussel was apparently resistant to the virus, but it apparently accumulated and maintained substantial levels (up to 10 -10 ID 50 /ml -1 ) of the virus for the dura- tion of the experiment (35 days). The highest infectious titer was found in brown trout (10 6 ID 50 /ml -1 ). In zebrafi sh and swan mussels, it was up to 10 4.5 ID 50 /ml -1 and 10 2 ID 50 /ml -1 respectively. Conclusions. At the end of the experiments the IPNV titers were higher than the initial viral doses in brown trout and zebrafi sh, indicating virus reproduction. In diseasesd fi sh the blackening, whirling and anorexia were observed. It was shown that swan mussel accumulates and maintains virus for at least 35 days under experimental conditions and that it may act as symptomless vector of IPNV

Virus-Mediated Cell-Cell Fusion.

Cell-cell fusion between eukaryotic cells is a general process involved in many physiological and pathological conditions, including infections by bacteria, parasites, and viruses. As obligate intracellular pathogens, viruses use intracellular machineries and pathways for efficient replication in their host target cells. Interestingly, certain viruses, and, more especially, enveloped viruses belonging to different viral families and including human pathogens, can mediate cell-cell fusion between infected cells and neighboring non-infected cells. Depending of the cellular environment and tissue organization, this virus-mediated cell-cell fusion leads to the merge of membrane and cytoplasm contents and formation of multinucleated cells, also called syncytia, that can express high amount of viral antigens in tissues and organs of infected hosts. This ability of some viruses to trigger cell-cell fusion between infected cells as virus-donor cells and surrounding non-infected target cells is mainly related to virus-encoded fusion proteins, known as viral fusogens displaying high fusogenic properties, and expressed at the cell surface of the virus-donor cells. Virus-induced cell-cell fusion is then mediated by interactions of these viral fusion proteins with surface molecules or receptors involved in virus entry and expressed on neighboring non-infected cells. Thus, the goal of this review is to give an overview of the different animal virus families, with a more special focus on human pathogens, that can trigger cell-cell fusion.

Detección y caracterización molecular del SARS-CoV- 2 en animales

El origen zoonótico del SARS-CoV-2 ha planteado la necesidad de realizar el monitoreo y control de esta virosis en animales domésticos y silvestres para determinar el posible rol de los animales como portadores, reservorios y/o amplificadores del virus. Los estudios para comprender mejor la susceptibilidad de diferentes especies animales al SARS-CoV-2 y evaluar la dinámica de la infección en estas especies, así como también la identificación de posibles reservorios y/o transmisores, son una herramienta fundamental para la vigilancia de la pandemia. El diseño de protocolos específicos para el relevamiento epidemiológico y la toma de muestras de animales pertenecientes a pacientes positivos a COVID-19, dieron inicio al trabajo.
 Los objetivos del proyecto involucran el diagnóstico y la vigilancia epidemiológica del SARS-CoV-2 en animales que conviven con personas infectadas, para analizar la infectividad potencial del virus y el rol que cumplen los animales en la pandemia. Estos estudios son una herramienta que nos permitirá conocer el comportamiento del virus en diferentes hospedadores y sus implicancias en la transmisión y perpetuación de la enfermedad.
 El proyecto comenzó con el diseño de protocolos para toma de muestras, definiendo caso sospechoso en animales, e indicando cómo debe proceder el Médico Veterinario particular al ser consultado por los propietarios. Dicho protocolo fue difundido por el Colegio de Veterinarios de la Provincia de Buenos Aires (Distrito II). Conjuntamente, se creó un consentimiento informado, aprobado por la Comisión Institucional para el Cuidado y Uso de Animales de Laboratorio (CICUAL) que es firmado por los responsables de los animales y del proyecto, acordando la toma de muestras. Por otro lado, el área de zoonosis dependiente del Ministerio de Salud de la Nación adoptó los protocolos y trabaja en la implementación de un sistema de vigilancia epidemiológica. Se realizó la difusión del proyecto en diversos medios de comunicación, permitiendo que aquellos pacientes diagnosticados positivos a SARS-CoV-2 se comunicaran con el grupo de trabajo para consultas y coordinación de la toma de muestras a sus mascotas.
 Hasta el 31 de agosto, se tomaron muestras de hisopados orofaríngeos y rectales de 15 caninos y 11 felinos, todos en estrecho contacto con personas diagnosticados positivos a SARS-CoV-2. Además, se colectaron muestras de un chimpancé del Ecoparque La Plata. Una vez en el laboratorio se realizó la extracción del ARN de las muestras utilizando un kit comercial, y el diagnóstico molecular por real-time RT-PCR adaptando kits comerciales. Las muestras analizadas fueron no reactivas a los targets específicos de SARS-CoV-2.

In silico mutagenesis of human ACE2 with S protein and translational efficiency explain SARS-CoV-2 infectivity in different species.

The coronavirus disease COVID-19 constitutes the most severe pandemic of the last decades having caused more than 1 million deaths worldwide. The SARS-CoV-2 virus recognizes the angiotensin converting enzyme 2 (ACE2) on the surface of human cells through its spike protein. It has been reported that the coronavirus can mildly infect cats, and ferrets, and perhaps dogs while not pigs, mice, chicken and ducks. Differences in viral infectivity among different species or individuals could be due to amino acid differences at key positions of the host proteins that interact with the virus, the immune response, expression levels of host proteins and translation efficiency of the viral proteins among other factors. Here, first we have addressed the importance that sequence variants of different animal species, human individuals and virus isolates have on the interaction between the RBD domain of the SARS-CoV-2 spike S protein and human angiotensin converting enzyme 2 (ACE2). Second, we have looked at viral translation efficiency by using the tRNA adaptation index. We find that integration of both interaction energy with ACE2 and translational efficiency explains animal infectivity. Humans are the top species in which SARS-CoV-2 is both efficiently translated as well as optimally interacting with ACE2. We have found some viral mutations that increase affinity for hACE and some hACE2 variants affecting ACE2 stability and virus binding. These variants suggest that different sensitivities to coronavirus infection in humans could arise in some cases from allelic variability affecting ACE2 stability and virus binding.

On the role of viruses in nature and what this means for the COVID-19 pandemic.

On the role of viruses in nature and what this means for the COVID-19 pandemic.

Preventing the Next Pandemic: Exploring the Origins and Spread of Animal Viruses

Preventing the Next Pandemic: Exploring the Origins and Spread of Animal Viruses

A retrospective survey of the seroprevalence of severe fever with thrombocytopenia syndrome virus in wild animals in Japan.

Severe fever with thrombocytopenia syndrome (SFTS) has a high fatality rate and is caused by SFTS virus (SFTSV). Currently, SFTS is endemic to some areas in western Japan, and wild animals are considered to play important roles in the circulation of SFTSV in the environment. Previous retrospective surveys using samples mainly obtained between 2006 and 2015 revealed serological evidence of SFTSV infection in wild animals; however, seroprevalence before 2006 remains unclear. In this study, we investigated the presence of anti‐SFTSV antibodies in a total of 521 serum samples from nine wild animal species collected from 11 prefectures in central and eastern Japan between 1980 and 2000. All samples yielded negative results for antibodies to SFTSV, suggesting that there had been few or no SFTSV infections before 2000 in the sampled areas.

Assessment of Viral Targeted Sequence Capture Using Nanopore Sequencing Directly from Clinical Samples.

Shotgun metagenomic sequencing (SMg) enables the simultaneous detection and characterization of viruses in human, animal and environmental samples. However, lack of sensitivity still poses a challenge and may lead to poor detection and data acquisition for detailed analysis. To improve sensitivity, we assessed a broad scope targeted sequence capture (TSC) panel (ViroCap) in both human and animal samples. Moreover, we adjusted TSC for the Oxford Nanopore MinION and compared the performance to an SMg approach. TSC on the Illumina NextSeq served as the gold standard. Overall, TSC increased the viral read count significantly in challenging human samples, with the highest genome coverage achieved using the TSC on the MinION. TSC also improved the genome coverage and sequencing depth in clinically relevant viruses in the animal samples, such as influenza A virus. However, SMg was shown to be adequate for characterizing a highly diverse animal virome. TSC on the MinION was comparable to the NextSeq and can provide a valuable alternative, offering longer reads, portability and lower initial cost. Developing new viral enrichment approaches to detect and characterize significant human and animal viruses is essential for the One Health Initiative.

Antiviral drugs targeting endosomal membrane proteins inhibit distant animal and human pathogenic viruses

The endocytic pathway is a common strategy that several highly pathogenic viruses use to enter into the cell. To demonstrate the usefulness of this pathway as a common target for the development of broad-spectrum antivirals, the inhibitory effect of drug compounds targeting endosomal membrane proteins were investigated. This study entailed direct comparison of drug effectiveness against animal and human pathogenic viruses, namely Ebola (EBOV), African swine fever virus (ASFV), and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).A panel of experimental and FDA-approved compounds targeting calcium channels and PIKfyve at the endosomal membrane caused potent reductions of entry up to 90% in SARS-CoV-2 S-protein pseudotyped retrovirus. Similar inhibition was observed against transduced EBOV glycoprotein pseudovirus and ASFV. SARS-CoV-2 infection was potently inhibited by selective estrogen receptor modulators in cells transduced with pseudovirus, among them Raloxifen inhibited ASFV with very low 50% inhibitory concentration. Finally, the mechanism of the inhibition caused by the latter in ASFV infection was analyzed.Overall, this work shows that cellular proteins related to the endocytic pathway can constitute suitable cellular targets for broad range antiviral compounds.

The Roles of Apoptosis in Swine Response to Viral Infection and Pathogenesis of Swine Enteropathogenic Coronaviruses.

Apoptosis is a tightly regulated mechanism of cell death that plays important roles in various biological processes including biological evolution, multiple system development, anticancer, and viral infections. Swine enteropathogenic coronaviruses invade and damage villous epithelial cells of the small intestine causing severe diarrhea with high mortality rate in suckling piglets. Transmissible gastroenteritis virus (TGEV), Porcine epidemic diarrhea virus (PEDV), Porcine deltacoronavirus (PDCoV), and Swine acute diarrhea syndrome coronavirus (SADS-CoV) are on the top list of commonly-seen swine coronaviruses with a feature of diarrhea, resulting in significant economic losses to the swine industry worldwide. Apoptosis has been shown to be involved in the pathogenesis process of animal virus infectious diseases. Understanding the roles of apoptosis in host responses against swine enteropathogenic coronaviruses infection contribute to disease prevention and control. Here we summarize the recent findings that focus on the apoptosis during swine coronaviruses infection, in particular, TGEV, PEDV, PDCoV, and SADS-CoV.

White Spot Syndrome Virus Benefits from Endosomal Trafficking, Substantially Facilitated by a Valosin-Containing Protein, To Escape Autophagic Elimination and Propagate in the Crustacean Cherax quadricarinatus.

As the most severely lethal viral pathogen for crustaceans in both brackish water and freshwater, white spot syndrome virus (WSSV) has a mechanism of infection that remains largely unknown, which profoundly limits the control of WSSV disease. By using a hematopoietic tissue (Hpt) stem cell culture from the red claw crayfish Cherax quadricarinatus suitable for WSSV propagation in vitro, the intracellular trafficking of live WSSV, in which the acidic-pH-dependent endosomal environment was a prerequisite for WSSV fusion, was determined for the first time via live-cell imaging. When the acidic pH within the endosome was alkalized by chemicals, the intracellular WSSV virions were detained in dysfunctional endosomes, resulting in appreciable blocking of the viral infection. Furthermore, disrupted valosin-containing protein (C. quadricarinatus VCP [CqVCP]) activity resulted in considerable aggregation of endocytic WSSV virions in the disordered endosomes, which subsequently recruited autophagosomes, likely by binding to CqGABARAP via CqVCP, to eliminate the aggregated virions within the dysfunctional endosomes. Importantly, both autophagic sorting and the degradation of intracellular WSSV virions were clearly enhanced in Hpt cells with increased autophagic activity, demonstrating that autophagy played a defensive role against WSSV infection. Intriguingly, most of the endocytic WSSV virions were directed to the endosomal delivery system facilitated by CqVCP activity so that they avoided autophagy degradation and successfully delivered the viral genome into Hpt cell nuclei, which was followed by the propagation of progeny virions. These findings will benefit anti-WSSV target design against the most severe viral disease currently affecting farmed crustaceans.IMPORTANCE White spot disease is currently the most devastating viral disease in farmed crustaceans, such as shrimp and crayfish, and has resulted in a severe ecological problem for both brackish water and freshwater aquaculture areas worldwide. Efficient antiviral control of WSSV disease is still lacking due to our limited knowledge of its pathogenesis. Importantly, research on the WSSV infection mechanism is also quite meaningful for the elucidation of viral pathogenesis and virus-host coevolution, as WSSV is one of the largest animal viruses, in terms of genome size, that infects only crustaceans. Here, we found that most of the endocytic WSSV virions were directed to the endosomal delivery system, strongly facilitated by CqVCP, so that they avoided autophagic degradation and successfully delivered the viral genome into the Hpt cell nucleus for propagation. Our data point to a virus-sorting model that might also explain the escape of other enveloped DNA viruses.

Here, the huge rainbow within the COVID-19 storm.

The real-life experience of the 2020 pandemic has shown us how totally unprepared we were to face such an event, albeit an event with several unique features. The inter-species jump by an animal virus to humans is, perhaps, one of its least unique characteristics as this has occurred several times in the past. The total absence of pre-existing immunity in the human host was the worst possible scenario in the face of rapid worldwide virus dissemination through international travel making COVID-19 effectively unstoppable.

Multi-targeted gene silencing strategies inhibit replication of Canine morbillivirus

BackgroundCanine morbilivirus (canine distemper virus, CDV) is a highly contagious pathogen associated with high morbidity and mortality in susceptible carnivores. Although there are CDV vaccines available, the disease poses a huge threat to dogs and wildlife hosts due to vaccine failures and lack of effective treatment. Thus, the development of therapeutics is an urgent need to achieve rapid outbreak control and reduce mortality in target species. Gene silencing by RNA interference has emerged as a promising therapeutic approach against different human and animal viruses. In this study, plasmid-based short hairpin RNAs (shRNAs) against three different regions in either CDV nucleoprotein (N), or large polymerase (L) genes and recombinant adenovirus-expressing N-specific multi-shRNAs were generated. Viral cytopathic effect, virus titration, plaque-forming unit reduction, and real-time quantitative RT-PCR analysis were used to check the efficiency of constructs against CDV.ResultsIn CDV-infected VerodogSLAM cells, shRNA-expressing plasmids targeting the N gene markedly inhibited the CDV replication in a dose-dependent manner, with viral genomes and titers being decreased by over 99%. Transfection of plasmid-based shRNAs against the L gene displayed weaker inhibition of viral RNA level and virus yield as compared to CDV N shRNAs. A combination of shRNAs targeting three sites in the N gene considerably reduced CDV RNA and viral titers, but their effect was not synergistic. Recombinant adenovirus-expressing multiple shRNAs against CDV N gene achieved a highly efficient knockdown of CDV N mRNAs and successful inhibition of CDV replication.ConclusionsWe found that this strategy had strong silencing effects on CDV replication in vitro. Our findings indicate that the delivery of shRNAs using plasmid or adenovirus vectors potently inhibits CDV replication and provides a basis for the development of therapeutic strategies for clinical trials.

Review on structural mechanism and mode of action of corona virus

Human coronaviruses, first reported in the 1960s, were the reason for a large number of children infected with the upper respiratory tract. SARS and four other coronaviruses have been reported since 2003, causing substantial morbidity and mortality. A new coronavirus was firstly reported from Hubei Province, Wuhan City in China, in December 2019 and named as novel Corona Virus. Later on, this virus is referred to as COVID-19, which can cause diseases similar to SARS and named for the spikes that protrude from their membranes, like the sun’s corona. The virus is believed to have been spread by animals sold in Wuhan from the seafood market and Health authorities in China reported fever, cough, breathing problems, and pneumonia in patients. The virus has a variety of physical and pathogenic features similar to the SARS-CoV-2 virus. Closer contact may cause the virus to be transmitted Initial flu-like symptoms, such as fever, cough, myalgia, and dyspnea in the 2-14 days of viral contact. People who are chronically ill are usually older or have medical co-morbidity. No treatments or virus vaccines are currently available. Seasonal Influenza has less mortality rate and less infectious than SARS-CoV-2. Many of the viruses remain to be identified as they will spread. Future directions for SARS-CoV work include greater knowledge of the replication process, tropism, and immune-response mechanisms, taking account of the possible functions of groups-specific proteins; development of animal and human virus vaccine strategies, and anti-viral therapies and very likely isolation and characterization of new human pathogenic coronaviruses. Emergencies for identifying and quarantining contaminated individuals early in the process of preventing their further spread are being made through global efforts

Hepatitis D virus in Bulgaria: virology, epidemiology and pathogenesis in chronic HBV carriers with liver dysfunction

Inroduction: Hepatitis D (HDV) is the most interesting and unique among animal viruses. It causes viral hepatitis D only in individuals already infected with HBV (hepatitis B). This dual infection leads to the most aggressive hepatic dysfunction of all human viral hepatitis. Aim: This study was made to outline the hepatitis D virus among patients with chronic liver disorders in northeastern Bulgaria, in the sight of virus epidemiology, pathogenicity and viral genotype. Materials and methods: This is a retrospective study conducted between 2013-2019 at St. Marina University Hospital,Varna, Bulgaria. We have analyzed 418 serum samples from 391 patients with chronic liver disease using ELISA, PCR and HDV sequencing and genotyping. Results and Discussion: From 391 patients with chronic liver abnormalities, 16.6% (95% CI: 15.9% - 23.8%, n = 65) had an etiological association with HDV in ELISA. We found HDV RNA positive results in 63 out of all 65 anti-HDV Ab (antibody) positive patients (96.9%). Twenty-four of them, or 38.1% (95% CI: 26.1% - 51.2%, n = 24), were on antiviral HBV/HDV therapy. For five of them, or 20.8% (95% CI: 7.1% - 42.2%, n = 5), HDV genotype I was found. Conclusion: HDV infection has still many mysteries to discover - in terms of pathogenesis, clinical outcome in chronic HBV/HDV-infected individuals, as well as genotype variations and their role in avoiding immune elimination of the virus. All these unanswered questions pose a great challenge to the scientific thought and efforts of humankind to reduce and gradually eliminate viral hepatitis D.