One of the crucial steps during trials for Zika and other vaccines is to recruit participants and to understand how participants' attitudes and sociodemographic characteristics affect willingness to participate (WTP). This study was conducted to assess WTP, its explanatory variables, and the impact of financial compensation on WTP in Indonesia. A health facility-based cross-sectional study was conducted in eleven regencies in the Aceh and West Sumatra provinces of Indonesia. Participants were recruited via a convenience sampling method and were interviewed. The associations between explanatory variables and WTP were assessed using a two-step logistic regression analysis. A total of 1,102 parents were approached, and of these 956 (86.8%) completed the interview and were included in analysis. Of those, 144 (15.1%) were willing to participate in a Zika vaccine trial without a financial compensation. In the multivariate analysis, WTP was tied to an age of more than 50 years old, compared to 20⁻29 years (odds ratio (OR): 5.0; 95% confidence interval (CI): 2.37⁻10.53), to being female (OR: 2.20; 95% CI: 1.11⁻4.37), and to having heard about Zika (OR: 2.41; 95% CI: 1.59⁻3.65). Participants' WTP increased gradually with higher financial compensation. The rate of WTP increased to 62.3% at the highest offer (US$ 350.4), and those who were still unwilling to participate (37.7%) had a poorer attitude towards childhood vaccination. This study highlights that pre-existing knowledge about Zika and attitudes towards childhood vaccination are important in determining community members being willing to participate in a vaccine trial. Financial incentives are still an important factor to enhance participant recruitment during a vaccine trial.
Micro RNAs (miRNAs) are a class of small non-coding single-stranded RNA, which play an important role in modulating host-Influenza A virus (IAV) crosstalk. The interplay between influenza and miRNA interaction is defined by a plethora of complex mechanisms, which are not fully understood yet. Here, we demonstrate that in IAV infected A549 cells, a synchronous increase was observed in the expression of mTOR up to 24 hpi and significant downregulation at 48 hpi. Additionally, NP of IAV interacts with mTOR and modulates the levels of mTOR mRNA and protein, thus regulating the translation of host cell. RNA sequencing and qPCR analysis of IAV-infected A549 cells and NP transfected cells revealed that miR-101 downregulates mTOR transcripts at later stages of infection. Ectopic expression of miR-101 mimic led to a decrease in expression of NP, a reduction in IAV titer and replication. Moreover, treatment of the cells with Everolimus, a potent inhibitor of mTOR, resulted in an increase of miR-101 transcript levels, which further suppressed the viral protein synthesis. Collectively, the data suggest a novel mechanism that IAV stimulates mTOR pathway at early stages of infection; however, at a later time-point, positive regulation of miR-101 restrains the mTOR expression, and hence, the viral propagation.
Nipah disease is a highly fatal zoonosis which is caused by the Nipah virus. The Nipah virus is a BSL-4 virus with fruit bats being its natural host. It is mainly prevalent in Southeast Asia. The virus was first discovered in 1997 in Negeri Sembilan, Malaysia. Currently, it is mainly harmful to pigs and humans with a high mortality rate. This study describes the route of transmission of the Nipah virus in different countries and analyzes the possibility of the primary disease being in China and the method of its transmission to China. The risk factors are analyzed for different susceptible populations to Nipah disease. The aim is to improve people's risk awareness and prevention and control of the disease and reduce its risk of occurring and spreading in China.
To establish a productive infection in host cells, viruses often use one or multiple host membrane glycoproteins as their receptors. For Influenza A virus (IAV) such a glycoprotein receptor has not been described, to date. Here we show that IAV is using the host membrane glycoprotein CD66c as a receptor for entry into human epithelial lung cells. Neuraminidase (NA), a viral spike protein, binds to CD66c on the cell surface during IAV entry into the host cells. Lung cells overexpressing CD66c showed an increase in virus binding and subsequent entry into the cell. Upon comparison, CD66c demonstrated higher binding capacity than other membrane glycoproteins (EGFR and DC-SIGN) reported earlier to facilitate IAV entry into host cells. siRNA mediated knockdown of CD66c from lung cells inhibited virus binding on cell surface and entry into cells. Blocking CD66c by antibody on the cell surface resulted in decreased virus entry. We found that CD66c is a specific glycoprotein receptor for influenza A virus that did not affect entry of non-IAV RNA virus (Hepatitis C virus). Finally, IAV pre-incubated with recombinant CD66c protein when administered intranasally in mice showed decreased cytopathic effects in mice lungs. This publication is the first to report CD66c (Carcinoembryonic cell adhesion molecule 6 or CEACAM6) as a glycoprotein receptor for Influenza A virus.
The Enterovirus genus of the Picornaviridae family comprises many important human pathogens, including polioviruses, rhinovirus, enterovirus A71, and enterovirus D68. They cause a wide variety of diseases, ranging from mild to severe life-threatening diseases. Currently, no effective vaccine is available against enteroviruses except for poliovirus. Enteroviruses subvert the autophagic machinery to benefit their assembly, maturation, and exit from host. Some enteroviruses spread between cells via a process described as autophagosome-mediated exit without lysis (AWOL). The early and late phases of autophagy are regulated through various lipids and their metabolizing enzymes. Some of these lipids and enzymes are specifically regulated by enteroviruses. In the present review, we summarize the current understanding of the regulation of autophagic machinery by enteroviruses, and provide updates on recent developments in this field.
Dengue virus (DENV) presents a significant threat to global public health with more than 500,000 hospitalizations and 25,000 deaths annually. Currently, there is no clinically approved antiviral drug to treat DENV infection. The envelope (E) glycoprotein of DENV is a promising target for drug discovery as the E protein is important for viral attachment and fusion. Understanding the structure and function of DENV E protein has led to the exploration of structure-based drug discovery of antiviral compounds and peptides against DENV infections. This review summarizes the structural information of the DENV E protein with regards to DENV attachment and fusion. The information enables the development of antiviral agents through structure-based approaches. In addition, this review compares the potency of antivirals targeting the E protein with the antivirals targeting DENV multifunctional enzymes, repurposed drugs and clinically approved antiviral drugs. None of the current DENV antiviral candidates possess potency similar to the approved antiviral drugs which indicates that more efforts and resources must be invested before an effective DENV drug materializes.
Mammalian retroviruses cause a variety of diseases in their hosts, including hematological and immunodeficiency disorders. Both human T-cell leukemia (HTLV) and human immunodeficiency (HIV) viruses originated from several independent zoonotic transmissions, indicating that cross-species transmissions from animal to humans may still occur. Thus, as the risk for retroviral transmissions from animals to humans increase, we investigated whether mammalian retroviruses are involved in selected pediatric idiopathic diseases whose symptoms evoke retroviral infections. Blood samples, sera, and synovial fluids, or bone marrow cells were collected from pediatric patients under 18 years of age with different autoimmune idiopathic diseases. Overall, we screened clinical samples from 110 children using sensitive nested and semi-nested PCR strategies targeting env genes, and a C-type retrovirus reverse transcriptase (RT) activity kit. All clinical samples were free of retroviral signatures, indicating the unlikelihood of an etiological role of the retroviruses we assessed in the pediatric diseases we tested.
The SARS-CoV-2 virus is responsible for the rapid global spread of the COVID-19 disease. As a result, it is critical to understand and collect primary data on the virus, infection epidemiology, and treatment. Despite the speed with which the virus was detected, studies of its cell biology and architecture at the ultrastructural level are still in their infancy. Therefore, we investigated and analyzed the viral morphometry of SARS-CoV-2 to extract important key points of the virus's characteristics. Then, we proposed a prediction model to identify the real virus levels based on the optimization of a full recurrent neural network (RNN) using transmission electron microscopy (TEM) images. Consequently, identification of virus levels depends on the size of the morphometry of the area (width, height, circularity, roundness, aspect ratio, and solidity). The results of our model were an error score of training network performance 3.216 × 10-11 at 639 epoch, regression of -1.6 × 10-9, momentum gain (Mu) 1 × 10-9, and gradient value of 9.6852 × 10-8, which represent a network with a high ability to predict virus levels. The fully automated system enables virologists to take a high-accuracy approach to virus diagnosis, prevention of mutations, and life cycle and improvement of diagnostic reagents and drugs, adding a point of view to the advancement of medical virology.
The increasing number of HIV-infected people who are receiving ART, including those with low adherence, is causing the spread of HIV drug resistance (DR). A total of 1396 plasma samples obtained from treatment-experienced patients from the Volga federal district (VFD), Russia, were examined to investigate HIV DR occurrence. The time periods 2008−2015 and 2016−2019 were compared. Fragmentary Sanger sequencing was employed to identify HIV resistance to reverse transcriptase inhibitors (RTIs) and protease inhibitors (PIs) using an ABI 3500XL genetic analyzer, a ViroSeq™ HIV-1 genotyping system (Alameda, CA, USA) and AmpliSense HIV-Resist-Seq reagent kits (Moscow, Russia). In 2016−2019, HIV DR was detected significantly more often than in 2008−2015 (p < 0.01). Mutations to RTIs retained leading positions in the structure of DR. Frequencies of resistance mutations to nucleoside and non-nucleoside RTIs (NRTIs and NNRTIs) in the spectra of detected mutations show no significant differences. Resistance to NRTIs after 2016 began to be registered more often as a part of multidrug resistance (MDR), as opposed to resistance to a single class of antiretrovirals. The frequency of DR mutations to PIs was low, both before and after 2016 (7.9% and 6.1% in the spectrum, respectively, p > 0.05). MDR registration rate became significantly higher from 2008 to 2019 (17.1% to 72.7% of patients, respectively, p < 0.01). M184V was the dominant replacement in all the years of study. A significant increase in the frequency of K65R replacement was revealed. The prevalence of integrase strand transfer inhibitor (INSTI) resistance mutations remains to be investigated.
Bat-borne viruses can pose a serious threat to human health, with examples including Nipah virus (NiV) in Bangladesh and Malaysia, and Marburg virus (MARV) in Africa. To date, significant human outbreaks of such viruses have not been reported in the European Union (EU). However, EU countries have strong historical links with many of the countries where NiV and MARV are present and a corresponding high volume of commercial trade and human travel, which poses a potential risk of introduction of these viruses into the EU. In assessing the risks of introduction of these bat-borne zoonotic viruses to the EU, it is important to consider the location and range of bat species known to be susceptible to infection, together with the virus prevalence, seasonality of viral pulses, duration of infection and titre of virus in different bat tissues. In this paper, we review the current scientific knowledge of all these factors, in relation to the introduction of NiV and MARV into the EU.
Nipah virus (NiV) is a paramyxovirus responsible for a high mortality rate zoonosis. As a result, it has been included in the list of Blueprint priority pathogens. Bats are the main reservoirs of the virus, and different clinical courses have been described in humans. The Bangladesh strain (NiV-B) is often associated with severe respiratory disease, whereas the Malaysian strain (NiV-M) is often associated with severe encephalitis. An early diagnosis of NiV infection is crucial to limit the outbreak and to provide appropriate care to the patient. Due to high specificity and sensitivity, qRT-PCR is currently considered to be the optimum method in acute NiV infection assessment. Nasal swabs, cerebrospinal fluid, urine, and blood are used for RT-PCR testing. N gene represents the main target used in molecular assays. Different sensitivities have been observed depending on the platform used: real-time PCR showed a sensitivity of about 103 equivalent copies/reaction, SYBRGREEN technology's sensitivity was about 20 equivalent copies/reaction, and in multiple pathogen card arrays, the lowest limit of detection (LOD) was estimated to be 54 equivalent copies/reaction. An international standard for NiV is yet to be established, making it difficult to compare the sensitivity of the different methods. Serological assays are for the most part used in seroprevalence studies owing to their lower sensitivity in acute infection. Due to the high epidemic and pandemic potential of this virus, the diagnosis of NiV should be included in a more global One Health approach to improve surveillance and preparedness for the benefit of public health. Some steps need to be conducted in the diagnostic field in order to become more efficient in epidemic management, such as development of point-of-care (PoC) assays for the rapid diagnosis of NiV.
Dengue is a major global health threat causing 390 million dengue infections and 25,000 deaths annually. The lack of efficacy of the licensed Dengvaxia vaccine and the absence of a clinically approved antiviral against dengue virus (DENV) drive the urgent demand for the development of novel anti-DENV therapeutics. Various antiviral agents have been developed and investigated for their anti-DENV activities. This review discusses the mechanisms of action employed by various antiviral agents against DENV. The development of host-directed antivirals targeting host receptors and direct-acting antivirals targeting DENV structural and non-structural proteins are reviewed. In addition, the development of antivirals that target different stages during post-infection such as viral replication, viral maturation, and viral assembly are reviewed. Antiviral agents designed based on these molecular mechanisms of action could lead to the discovery and development of novel anti-DENV therapeutics for the treatment of dengue infections. Evaluations of combinations of antiviral drugs with different mechanisms of action could also lead to the development of synergistic drug combinations for the treatment of dengue at any stage of the infection.
Maize dwarf mosaic virus (MDMV) is a serious maize pathogen, epidemic worldwide, and one of the most common virus diseases for monocotyledonous plants, causing up to 70% loss in corn yield globally since 1960. MDMV belongs to the genus Potyvirus (Potyviridae) and was first identified in 1964 in Illinois in corn and Johnsongrass. MDMV is a single stranded positive sense RNA virus and is transmitted in a non-persistent manner by several aphid species. MDMV is amongst the most important virus diseases in maize worldwide. This review will discuss its genome, transmission, symptomatology, diagnosis and management. Particular emphasis will be given to the current state of knowledge on the diagnosis and control of MDMV, due to its importance in reducing the impact of maize dwarf mosaic disease, to produce an enhanced quality and quantity of maize.
Positive-strand RNA virus evolution is partly attributed to the process of recombination. Although common between closely genetically related viruses, such as within species of the Enterovirus genus of the Picornaviridae family, inter-species recombination is rarely observed in nature. Recent studies have shown recombination is a ubiquitous process, resulting in a wide range of recombinant genomes and progeny viruses. While not all recombinant genomes yield infectious progeny virus, their existence and continued evolution during replication have critical implications for the evolution of the virus population. In this study, we utilised an in vitro recombination assay to demonstrate inter-species recombination events between viruses from four enterovirus species, A-D. We show that inter-species recombinant genomes are generated in vitro with polymerase template-switching events occurring within the virus polyprotein coding region. However, these genomes did not yield infectious progeny virus. Analysis and attempted recovery of a constructed recombinant cDNA revealed a restriction in positive-strand but not negative-strand RNA synthesis, indicating a significant block in replication. This study demonstrates the propensity for inter-species recombination at the genome level but suggests that significant sequence plasticity would be required in order to overcome blocks in the virus life cycle and allow for the production of infectious viruses.
We are in the midst of a pandemic where the infective agent has been identified, but how it causes mild disease in some and fatally severe disease in other infected individuals remains a mystery [...].
Among emerging zoonotic pathogens, mosquito-borne viruses (MBVs) circulate between vertebrate animals and mosquitoes and represent a serious threat to humans via spillover from enzootic cycles to the human community. Active surveillance of MBVs in their vectors is therefore essential to better understand and prevent spillover and emergence, especially at the human-animal interface. In this study, we assessed the presence of MBVs using molecular and phylogenetic methods in mosquitoes collected along an ecological gradient ranging from rural urbanized areas to highland forest areas in northern Thailand. We have detected the presence of insect specific flaviviruses in our samples, and the presence of the emerging zoonotic Tembusu virus (TMUV). Reported for the first time in 1955 in Malaysia, TMUV remained for a long time in the shadow of other flaviviruses such as dengue virus or the Japanese encephalitis virus. In this study, we identified two new TMUV strains belonging to cluster 3, which seems to be endemic in rural areas of Thailand and highlighted the genetic specificities of this Thai cluster. Our results show the active circulation of this emerging flavivirus in Thailand and the need for continuous investigation on this poorly known but threatening virus in Asia.
Influenza still remains one of the most challenging diseases, posing a significant threat to public health. Host lipid rafts play a critical role in influenza A virus (IAV) assembly and budding, however, their role in polyvalent IAV host binding and endocytosis had remained elusive until now. In the present study, we observed co-localization of IAV with a lipid raft marker ganglioside, GM1, on the host surface. Further, we isolated the lipid raft micro-domains from IAV infected cells and detected IAV protein in the raft fraction. Finally, raft disruption using Methyl-β-Cyclodextrin revealed significant reduction in IAV host binding, suggesting utilization of host rafts for polyvalent binding on the host cell surface. In addition to this, cyclodextrin mediated inhibition of raft-dependent endocytosis showed significantly reduced IAV internalization. Interestingly, exposure of cells to cyclodextrin two hours post-IAV binding showed no such reduction in IAV entry, indicating use of raft-dependent endocytosis for host entry. In summary, this study demonstrates that host lipid rafts are selected by IAV as a host attachment factors for multivalent binding, and IAV utilizes these micro-domains to exploit raft-dependent endocytosis for host internalization, a virus entry route previously unknown for IAV.
Zika virus (ZIKV) causes no-to-mild symptoms or severe neurological disorders. To investigate the importance of viral and host genetic variations in determining ZIKV infection outcomes, we created three full-length infectious cDNA clones as bacterial artificial chromosomes for each of three spatiotemporally distinct and genetically divergent ZIKVs: MR-766 (Uganda, 1947), P6-740 (Malaysia, 1966), and PRVABC-59 (Puerto Rico, 2015). Using the three molecularly cloned ZIKVs, together with 13 ZIKV region-specific polyclonal antibodies covering nearly the entire viral protein-coding region, we made three conceptual advances: (i) We created a comprehensive genome-wide portrait of ZIKV gene products and their related species, with several previously undescribed gene products identified in the case of all three molecularly cloned ZIKVs. (ii) We found that ZIKV has a broad cell tropism in vitro, being capable of establishing productive infection in 16 of 17 animal cell lines from 12 different species, although its growth kinetics varied depending on both the specific virus strain and host cell line. More importantly, we identified one ZIKV-non-susceptible bovine cell line that has a block in viral entry but fully supports the subsequent post-entry steps. (iii) We showed that in mice, the three molecularly cloned ZIKVs differ in their neuropathogenicity, depending on the particular combination of viral and host genetic backgrounds, as well as in the presence or absence of type I/II interferon signaling. Overall, our findings demonstrate the impact of viral and host genetic variations on the replication kinetics and neuropathogenicity of ZIKV and provide multiple avenues for developing and testing medical countermeasures against ZIKV.
Flavonoids are natural biomolecules that are known to be effective antivirals. These biomolecules can act at different stages of viral infection, particularly at the molecular level to inhibit viral growth. Enterovirus A71 (EV-A71), a non-enveloped RNA virus, is one of the causative agents of hand, foot and mouth disease (HFMD), which is prevalent in Asia. Despite much effort, no clinically approved antiviral treatment is available for children suffering from HFMD. Flavonoids from plants serve as a vast reservoir of therapeutically active constituents that have been explored as potential antiviral candidates against RNA and DNA viruses. Here, we reviewed flavonoids as evidence-based natural sources of antivirals against non-picornaviruses and picornaviruses. The detailed molecular mechanisms involved in the inhibition of EV-A71 infections are discussed.
The recent emergence of Zika virus (ZIKV) in Brazil was associated with an increased number of fetal brain infections that resulted in a spectrum of congenital neurological complications known as congenital Zika syndrome (CZS). Herein, we generated de novo from sequence data an early Asian lineage ZIKV isolate (ZIKV-MY; Malaysia, 1966) not associated with microcephaly and compared the in vitro replication kinetics and fetal brain infection in interferon α/β receptor 1 knockout (IFNAR1-/-) dams of this isolate and of a Brazilian isolate (ZIKV-Natal; Natal, 2015) unequivocally associated with microcephaly. The replication efficiencies of ZIKV-MY and ZIKV-Natal in A549 and Vero cells were similar, while ZIKV-MY replicated more efficiently in wild-type (WT) and IFNAR-/- mouse embryonic fibroblasts. Viremias in IFNAR1-/- dams were similar after infection with ZIKV-MY or ZIKV-Natal, and importantly, infection of fetal brains was also not significantly different. Thus, fetal brain infection does not appear to be a unique feature of Brazilian ZIKV isolates.