Lrp1 is a host entry factor for Rift Valley fever virus.

Rift Valley fever virus (RVFV) is a zoonotic pathogen with pandemic potential. RVFV entry is mediated by the viral glycoprotein (Gn), but host entry factors remain poorly defined. Our genome-wide CRISPR screen identified low-density lipoprotein receptor-related protein 1 (mouse Lrp1/human LRP1), heat shock protein (Grp94), and receptor-associated protein (RAP) as critical host factors for RVFV infection. RVFV Gn directly binds to specific Lrp1 clusters and is glycosylation independent. Exogenous addition of murine RAP domain 3 (mRAPD3) and anti-Lrp1 antibodies neutralizes RVFV infection in taxonomically diverse cell lines. Mice treated with mRAPD3 and infected with pathogenic RVFV are protected from disease and death. A mutant mRAPD3 that binds Lrp1 weakly failed to protect from RVFV infection. Together, these data support Lrp1 as a host entry factor for RVFV infection and define a new target to limit RVFV infections.

Acute Rift Valley fever virus infection induces inflammatory cytokines and cell death in ex vivo rat brain slice culture.

Rift Valley fever virus (RVFV) is an emerging arboviral disease with pandemic potential. While infection is often self-limiting, a subset of individuals may develop late-onset encephalitis, accounting for up to 20 % of severe cases. Importantly, individuals displaying neurologic disease have up to a 53 % case fatality rate, yet the neuropathogenesis of RVFV infection remains understudied. In this study, we evaluated whether ex vivo postnatal rat brain slice cultures (BSCs) could be used to evaluate RVFV infection in the central nervous system. BSCs mounted an inflammatory response after slicing, which resolved over time, and they were viable in culture for at least 12 days. Infection of rat BSCs with pathogenic RVFV strain ZH501 induced tissue damage and apoptosis over 48 h. Viral replication in BSCs reached up to 1×107 p.f.u. equivalents/ml, depending on inoculation dose. Confocal immunofluorescent microscopy of cleared slices confirmed direct infection of neurons as well as activation of microglia and astrocytes. Further, RVFV-infected rat BSCs produced antiviral cytokines and chemokines, including MCP-1 and GRO/KC. This study demonstrates that rat BSCs support replication of RVFV for ex vivo studies of neuropathogenesis. This allows for continued and complementary investigation into RVFV infection in an ex vivo postnatal brain slice culture format.

Rift Valley Fever virus M and L genome segment detection: a comparison of field-deployable reverse transcription insulated isothermal PCR (RT-iiPCR) and laboratory-based multiplex reverse transcription real-time PCR.

Rift Valley Fever phlebovirus (RVFV) is a mosquito-borne zoonotic pathogen that causes major agricultural and public health problems in Africa and the Arabian Peninsula. It is considered a potential agro-bioterrorism agent for which limited countermeasures are available. To address diagnostic needs, here we describe a rapid and sensitive molecular method immediately employable at sites of suspected outbreaks in animals that commonly precede outbreaks in humans. The strategy involves the concurrent detection of two of the three RVFV genome segments (large and medium) using reverse transcription insulated isothermal PCR (RT-iiPCR) performed on a portable, touch screen nucleic acid analyzer, POCKIT. The analytical sensitivity for both the RT-iiPCR and a laboratory-based L and M multiplex reverse transcription real-time PCR assay was estimated at approximately 0.1-3 copies/reaction using synthetic RNA or viral RNA. The diagnostic sensitivity and specificity of detection of RVFV on the POCKIT, determined using sera from sheep and cattle (n = 181) experimentally infected with two strains of RVFV (SA01 and Ken06), were 93.8% and 100% (kappa = 0.93), respectively. Testing of ruminant field sera (n = 193) in two locations in Africa demonstrated 100% diagnostic sensitivity and specificity. We conclude that the POCKIT dual-gene RVFV detection strategy can provide reliable, sensitive, and specific point-of-need viral RNA detection. Moreover, the field detection of RVFV in vectors or susceptible animal species can aid in the surveillance and epidemiological studies to better understand and control RVFV outbreaks. IMPORTANCE: The content of this manuscript is of interest to the diverse readership of the Journal of Clinical Microbiology, including research scientists, diagnosticians, healthcare professionals, and policymakers. Rift Valley Fever virus (RVFV) is a zoonotic mosquito-borne pathogen that causes major agricultural and public health problems. Current and most sensitive diagnostic approaches that are molecular-based are performed in highly specialized molecular diagnostic laboratories. To address diagnostic needs, we developed a novel, rapid, and sensitive molecular method using a portable PCR machine, POCKIT, capable of immediate deployment at sites of suspected outbreaks. Here, we demonstrate that field-deployable RVFV detection can provide reliable, sensitive, and specific point-of-need viral RNA detection that could be used for diagnostic investigations and epidemiological studies, and can be performed in the field.

Limiting viral replication in hepatocytes alters Rift Valley fever virus disease manifestations.

Rift Valley fever virus (RVFV) causes mild to severe disease in humans and livestock. Outbreaks of RVFV have been reported throughout Africa and have spread outside Africa since 2000, calling for urgent worldwide attention to this emerging virus. RVFV directly infects the liver, and elevated transaminases are a hallmark of severe RVFV infection. However, the specific contribution of viral replication in hepatocytes to pathogenesis of RVFV remains undefined. To address this, we generated a recombinant miRNA-targeted virus, RVFVmiR-122, to limit hepatocellular replication. MicroRNAs are evolutionarily conserved non-coding RNAs that regulate mRNA expression by targeting them for degradation. RVFVmiR-122 includes an insertion of four target sequences of the liver-specific miR-122. In contrast to control RVFVmiR-184, which contains four target sequences of mosquito-specific miR-184, RVFVmiR-122 has restricted replication in vitro in primary mouse hepatocytes. RVFVmiR-122-infected C57BL/6 mice survived acute hepatitis and instead developed late-onset encephalitis. This difference in clinical outcome was eliminated in Mir-122 KO mice, confirming the specificity of the finding. Interestingly, C57BL/6 mice infected with higher doses of RVFVmiR-122 had a higher survival rate which was correlated with faster clearance of virus from the liver, suggesting a role for activation of host immunity in the phenotype. Together, our data demonstrate that miR-122 can specifically restrict the replication of RVFVmiR-122 in liver tissue both in vitro and in vivo, and this restriction alters the clinical course of disease following RVFVmiR-122 infection. IMPORTANCE Rift Valley fever virus (RVFV) is a hemorrhagic fever virus that causes outbreaks in humans and livestock throughout Africa and has spread to continents outside Africa since 2000. However, no commercial vaccine or treatment is currently available for human use against RVFV. Although the liver has been demonstrated as a key target of RVFV, the contribution of viral replication in hepatocytes to overall RVFV pathogenesis is less well defined. In this study we addressed this question by using a recombinant miRNA-targeted virus with restricted replication in hepatocytes. We gained a better understanding of how this individual cell type contributes to the development of disease caused by RVFV. Techniques used in this study provide an innovative tool to the RVFV field that could be applied to study the consequences of limited RVFV replication in other target cells.

Transcriptomic Profiling Reveals Intense Host-Pathogen Dispute Compromising Homeostasis during Acute Rift Valley Fever Virus Infection.

Rift Valley fever virus (RVFV) (family Phenuiviridae) can cause severe disease, and outbreaks of this mosquito-borne pathogen pose a significant threat to public and animal health. Yet many molecular aspects of RVFV pathogenesis remain incompletely understood. Natural RVFV infections are acute, characterized by a rapid onset of peak viremia during the first days post-infection, followed by a rapid decline. Although in vitro studies identified a major role of interferon (IFN) responses in counteracting the infection, a comprehensive overview of the specific host factors that play a role in RVFV pathogenesis in vivo is still lacking. Here, the host in vivo transcriptional profiles in the liver and spleen tissues of lambs exposed to RVFV are studied using RNA sequencing (RNA-seq) technology. We validate that IFN-mediated pathways are robustly activated in response to infection. We also link the observed hepatocellular necrosis with severely compromised organ function, which is reflected as a marked downregulation of multiple metabolic enzymes essential for homeostasis. Furthermore, we associate the elevated basal expression of LRP1 in the liver with RVFV tissue tropism. Collectively, the results of this study deepen the knowledge of the in vivo host response during RVFV infection and reveal new insights into the gene regulation networks underlying pathogenesis in a natural host. IMPORTANCE Rift Valley fever virus (RVFV) is a mosquito-transmitted pathogen capable of causing severe disease in animals and humans. Outbreaks of RVFV pose a significant threat to public health and can result in substantial economic losses. Little is known about the molecular basis of RVFV pathogenesis in vivo, particularly in its natural hosts. We employed RNA-seq technology to investigate genome-wide host responses in the liver and spleen of lambs during acute RVFV infection. We show that RVFV infection drastically decreases the expression of metabolic enzymes, which impairs normal liver function. Moreover, we highlight that basal expression levels of the host factor LRP1 may be a determinant of RVFV tissue tropism. This study links the typical pathological phenotype induced by RVFV infection with tissue-specific gene expression profiles, thereby improving our understanding of RVFV pathogenesis.

Rift Valley Fever Virus Nucleoprotein Triggers Autophagy to Dampen Antiviral Innate Immune Responses.

Rift Valley fever virus (RVFV) is a mosquito-borne bunyavirus that causes severe and potentially fatal hemorrhagic fever in humans. Autophagy is a self-degradative process that can restrict viral replication at multiple infection steps. In this study, we evaluated the effects of RVFV-triggered autophagy on viral replication and immune responses. Our results showed that RVFV infection triggered autophagosome formation and induced complete autophagy. Impairing autophagy flux by depleting autophagy-related gene 5 (ATG5), ATG7, or sequestosome 1 (SQSTM1) or treatment with autophagy inhibitors markedly reduced viral RNA synthesis and progeny virus production. Mechanistically, our findings demonstrated that the RVFV nucleoprotein (NP) C-terminal domain interacts with the autophagy receptor SQSTM1 and promotes the SQSTM1-microtubule-associated protein 1 light chain 3 B (LC3B) interaction and autophagy. Deletion of the NP C-terminal domain impaired the interaction between NP and SQSTM1 and its ability to trigger autophagy. Notably, RVFV-triggered autophagy promoted viral infection in macrophages but not in other tested cell types, including Huh7 hepatocytes and human umbilical vein endothelial cells, suggesting cell type specificity of this mechanism. It was further revealed that RVFV NP-triggered autophagy dampens antiviral innate immune responses in infected macrophages to promote viral replication. These results provide novel insights into the mechanisms of RVFV-triggered autophagy and indicate the potential of targeting the autophagy pathway to develop antivirals against RVFV. IMPORTANCE We showed that RVFV infection induced the complete autophagy process. Depletion of the core autophagy genes ATG5, ATG7, or SQSTM1 or pharmacologic inhibition of autophagy in macrophages strongly suppressed RVFV replication. We further revealed that the RVFV NP C-terminal domain interacted with SQSTM1 and enhanced the SQSTM1/LC3B interaction to promote autophagy. RVFV NP-triggered autophagy strongly inhibited virus-induced expression of interferon-stimulated genes in infected macrophages but not in other tested cell types. Our study provides novel insights into the mechanisms of RVFV-triggered autophagy and highlights the potential of targeting autophagy flux to develop antivirals against this virus.

Pharmacokinetic Analyses of a Lipid Nanoparticle-Encapsulated mRNA-Encoded Antibody against Rift Valley Fever Virus.

Rift Valley fever virus (RVFV) could cause an emergency illness characterized by fever, muscle pain, and even death in humans or ruminants. However, there are no approved antiviral drugs that prevent or treat RVFV infection. While therapeutic antibodies have shown promising potential for prevention or treatment in several studies, many studies are ongoing, especially in the field of infectious diseases. Among these studies, the mRNA-LNP platform shows great potential for application, following the COVID-19 pandemic. Previously, we have obtained a neutralizing antibody against RVFV, which was named A38 protein and verified to have a high binding and neutralization ability. In this study, we aimed to identify an effectively optimized sequence and expressed the prioritized mRNA-encoded antibody in vitro. Notably, we effectively expressed mRNA-encoded protein and used the mRNA-LNP platform to generate A38-mRNA-LNP. Pharmacokinetic experiments were conducted in vivo and set up in two groups of mRNA-A38 group and A38 protein group, which were derived from mRNA-LNP and plasmid DNA-expressed proteins, respectively. A38-mRNA-LNPs were administrated by intramuscular injection, A38 proteins were administrated by intravenous administration, and their unique ability to maintain long-lasting protein concentrations by mRNA-encoded protein was demonstrated with the mRNA-encoded protein providing a longer circulating half-life compared to injection of the free A38 protein. These preclinical data on the mRNA-encoded antibody highlighted its potential to prevent infectious diseases in the future.

First detection of Rift Valley fever virus antibodies in non-human primates in Cameroon.

We tested for Rift Valley fever virus (RVFV) from at least 15 species of non-human primates. RVFV IgG/IgM antibodies were detected in 3.7% (2 out of 53) of chimpanzees (Pan troglodytes) and in 1.4% (1 out of 72) of unidentified non-human primate species. This study was the first investigation of RVFV in monkeys in Cameroon.

Potent neutralization of Rift Valley fever virus by human monoclonal antibodies through fusion inhibition.

Rift Valley fever virus (RVFV), an emerging arboviral and zoonotic bunyavirus, causes severe disease in livestock and humans. Here, we report the isolation of a panel of monoclonal antibodies (mAbs) from the B cells of immune individuals following natural infection in Kenya or immunization with MP-12 vaccine. The B cell responses of individuals who were vaccinated or naturally infected recognized similar epitopes on both Gc and Gn proteins. The Gn-specific mAbs and two mAbs that do not recognize either monomeric Gc or Gn alone but recognized the hetero-oligomer glycoprotein complex (Gc+Gn) when Gc and Gn were coexpressed exhibited potent neutralizing activities in vitro, while Gc-specific mAbs exhibited relatively lower neutralizing capacity. The two Gc+Gn-specific mAbs and the Gn domain A-specific mAbs inhibited RVFV fusion to cells, suggesting that mAbs can inhibit the exposure of the fusion loop in Gc, a class II fusion protein, and thus prevent fusion by an indirect mechanism without direct fusion loop contact. Competition-binding analysis with coexpressed Gc/Gn and mutagenesis library screening indicated that these mAbs recognize four major antigenic sites, with two sites of vulnerability for neutralization on Gn. In experimental models of infection in mice, representative mAbs recognizing three of the antigenic sites reduced morbidity and mortality when used at a low dose in both prophylactic and therapeutic settings. This study identifies multiple candidate mAbs that may be suitable for use in humans against RVFV infection and highlights fusion inhibition against bunyaviruses as a potential contributor to potent antibody-mediated neutralization.

Adaptation to a Multiplex Bead Assay and Seroprevalence to Rift Valley Fever N Protein: Nampula Province, Mozambique, 2013-2014.

Rift Valley fever virus (RVFV) is endemic in sub-Saharan Africa (SSA), with outbreaks reported in the Arabian Peninsula and throughout SSA. The natural reservoir for RVFV are ruminants, with livestock populations exceeding 50% exposure rates in some areas of SSA. Transmission to humans can occur through exposure to infected livestock products or multiple species of mosquito vectors. In 2013 and 2014, cross-sectional surveys occurred in two districts of Nacala-a-Velha and Mecubúri in northern Mozambique, and participants provided blood samples for later serological assays. IgG against the N protein of RVFV was detected through multiplex bead assay (MBA). Of the 2,278 persons enrolled between the two surveys and study sites, 181 (7.9%, 95% confidence interval (CI): 6.9%-9.1%) were found to be IgG seropositive with increasing seroprevalence with older age and significantly higher seroprevalence in Nacala-a-Velha (10.5%, 8.8%-12.5%) versus Mecubúri (5.7%, 4.5%-7.1%). Seroprevalence estimates were not significantly different between the 2013 and 2014 surveys. Significant spatial clustering of IgG positive persons were consistent among surveys and within the two districts, pointing toward the consistency of serology data for making population-level assumptions regarding RVFV seroprevalence. A subset of persons (n = 539) provided samples for both the 2013 and 2014 surveys, and a low percentage (0.81%) of these were found to seroconvert between these two surveys. Including the RVFV N protein in an MBA antigen panel could assist elucidate RVFV exposure in SSA. IMPORTANCE Due to sporadic transmission, human contact with Rift Valley Fever Virus (RVFV) is difficult to ascertain at a population level. Detection of antibodies against RVFV antigens assist in estimating exposure as antibodies remain in the host long after the virus has been cleared. In this study, we show that antibodies against RVFV N protein can be detected from dried blood spot (DBS) samples being assayed by multiplex bead assay. DBS from two districts in northern Mozambique were tested for IgG against the N protein, and 7.9% of all enrolled persons were seropositive. Older persons, males, and persons residing closer to the coast had higher RVFV N protein seroprevalence. Spatial clustering of IgG positive persons was noted in both districts. These results show low exposure rates to RVFV in these two northern districts in Mozambique, and the ability to perform serology for the RVFV N protein from dried blood samples.

Rift Valley Fever Virus Infects the Posterior Segment of the Eye and Induces Inflammation in a Rat Model of Ocular Disease.

People infected with the mosquito-borne Rift Valley fever virus (RVFV) can suffer from eye-related problems resulting in ongoing vision issues or even permanent blindness. Despite ocular disease being the most frequently reported severe outcome, it is vastly understudied compared to other disease outcomes caused by RVFV. Ocular manifestations of RVFV include blurred vision, uveitis, and retinitis. When an infected individual develops macular or paramacular lesions, there is a 50% chance of permanent vision loss in one or both eyes. The cause of blinding ocular pathology remains unknown in part due to the lack of a tractable animal model. Using 3 relevant exposure routes, both subcutaneous (SC) and aerosol inoculation of Sprague Dawley rats led to RVFV infection of the eye. Surprisingly, direct inoculation of the conjunctiva did not result in successful ocular infection. The posterior segment of the eye, including the optic nerve, choroid, ciliary body, and retina, were all positive for RVFV antigen in SC-infected rats, and live virus was isolated from the eyes. Proinflammatory cytokines and increased leukocyte counts were also found in the eyes of infected rats. Additionally, human ocular cell lines were permissive for Lrp1-dependent RVFV infection. This study experimentally defines viral tropism of RVFV in the posterior segment of the rat eye and characterizes virally-mediated ocular inflammation, providing a foundation for evaluation of vaccines and therapeutics to protect against adverse ocular outcomes. IMPORTANCE Rift Valley fever virus (RVFV) infection leads to eye damage in humans in up to 10% of reported cases. Permanent blindness occurs in 50% of individuals with significant retinal scarring. Despite the prevalence and severity of this outcome, very little is known about the mechanisms of pathogenesis. We addressed this gap by developing a rodent model of ocular disease. Subcutaneous infection of Sprague Dawley rats resulted in infection of the uvea, retina, and optic nerve along with the induction of inflammation within the posterior eye. Infection of human ocular cells induced inflammatory responses and required host entry factors for RVFV infection similar to rodents. This work provides evidence of how RVFV infects the eye, and this information can be applied to help mitigate the devastating outcomes of RVF ocular disease through vaccines or treatments.

Structure of Rift Valley Fever Virus RNA-Dependent RNA Polymerase.

Rift Valley fever virus (RVFV) belongs to the order Bunyavirales and is the type species of genus Phlebovirus, which accounts for over 50% of family Phenuiviridae species. RVFV is mosquito-borne and causes severe diseases in both humans and livestock, and consists of three segments (S, M, L) in the genome. The L segment encodes an RNA-dependent RNA polymerase (RdRp, L protein) that is responsible for facilitating the replication and transcription of the virus. It is essential for the virus and has multiple drug targets. Here, we established an expression system and purification procedures for full-length L protein, which is composed of an endonuclease domain, RdRp domain, and cap-binding domain. A cryo-EM L protein structure was reported at 3.6 Å resolution. In this first L protein structure of genus Phlebovirus, the priming loop of RVFV L protein is distinctly different from those of other L proteins and undergoes large movements related to its replication role. Structural and biochemical analyses indicate that a single template can induce initiation of RNA synthesis, which is notably enhanced by 5' viral RNA. These findings help advance our understanding of the mechanism of RNA synthesis and provide an important basis for developing antiviral inhibitors. IMPORTANCE The zoonosis RVF virus (RVFV) is one of the most serious arbovirus threats to both human and animal health. RNA-dependent RNA polymerase (RdRp) is a multifunctional enzyme catalyzing genome replication as well as viral transcription, so the RdRp is essential for studying the virus and has multiple drug targets. In our study, we report the structure of RVFV L protein at 3.6 Å resolution by cryo-EM. This is the first L protein structure of genus Phlebovirus. Strikingly, a single template can initiate RNA replication. The structure and assays provide a comprehensive and in-depth understanding of the catalytic and substrate recognition mechanism of RdRp.

Safety and immunogenicity of inactivated Rift Valley Fever Smithburn viral vaccine in sheep.

BACKGROUND: The live-attenuated Rift Valley Fever Smithburn (SB) vaccine is one of the oldest products widely used in ruminants for control of RVF infections. Vaccinations with RVF Smithburn result in residual pathogenic effect and is limited for use in non-pregnant animals. Commercially available RVFV inactivated vaccines are considered safer options to control the disease. These products are prepared from virulent RVFV isolates and present occupational safety concerns. This research study evaluates the ability of an inactivated SB vaccine strain to elicit neutralising antibody response in sheep. METHODS: The RVF Smithburn vaccine was inactivated with binary ethylenimine at 37 °C. Inactivated RVFV cultures were adjuvanted with Montande™ Gel-01 and aluminium hydroxide (Al (OH)3) gel for immunogenicity and safety determination in sheep. The commercial RVF inactivated vaccine and a placebo were included as positive and negative control groups, respectively. RESULTS: Inactivated RVFV vaccine formulations were safe with all animals showing no clinical signs of RVFV infection and temperature reactions following prime-boost injections. The aluminium hydroxide formulated vaccine induced an immune response as early as 14 days post primary vaccination with neutralising antibody titre of 1:20 and a peak antibody titre of 1:83 was reached on day 56. A similar trend was observed in the animal group vaccinated with the commercial inactivated RVF vaccine obtaining the highest antibody titre of 1:128 on day 56. The neutralizing antibody levels remained within a threshold for the duration of the study. Merino sheep vaccinated with Montanide™ Gel-01-Smithburn were characterised with overall lower immune response when compared to aluminium hydroxide vaccine emulsions. CONCLUSIONS: These finding suggests that the inactivated RVF Smithburn vaccine strain adjuvanted with aluminium-hydroxide can be used an alternative to the products prepared from virulent RVFV isolates for protection of ruminants against the disease. The vaccine can further be evaluated for safety in pregnant ewes.

Antiviral and protective effect of small interfering RNAs against rift valley fever virus in vitro.

BACKGROUND: Rift Valley Fever Virus (RVFV) is an arbovirus, a zoonotic disease that resurfaces as a potential hazard beyond geographic boundaries. Fever that can proceed to encephalitis, retinitis, hemorrhagic fever, and death is the main manifestation observed in human infections. RVFV has no authorized medication. The RNA interference (RNAi) gene silencing pathway is extremely well conserved. By targeting specific genes, small interfering RNA (siRNA) can be used to suppress viral replication. The aim of this study was to design specific siRNAs against RVFV and evaluate their prophylactic and antiviral effects on the Vero cells. METHODS AND RESULTS: Various siRNAs were designed using different bioinformatics tools. Three unique candidates were tested against an Egyptian sheep cell culture-adapted strain BSL-2 that suppressed RVFV N mRNA expression. SiRNAs were transfected a day before RVFV infection (pre-transfection), and 1 h after the viral infection (post-transfection), and were evaluated to detect the silencing activity and gene expression decrease using real-time PCR and a TCID50 endpoint test. The degree of N protein expression was determined by western blot 48 h after viral infection. D2 which targets the (488-506 nucleotides), the middle region of RVFV N mRNA was the most effective siRNA at 30 nM concentration, it almost eliminates N mRNA expression when utilized as antiviral or preventive therapy. siRNAs had a stronger antiviral silencing impact when they were post-transfected into Vero cells. CONCLUSION: Pre and post-transfection of siRNAs significantly reduced RVFV titer in cell lines, offering novel and potentially effective anti-RVFV epidemics and epizootics therapy.

Computational prediction of phytochemical inhibitors against the cap-binding domain of Rift Valley fever virus.

Rift Valley fever is a zoonotic disease that can spread through livestock and mosquitoes, and its symptoms include retinitis, photophobia, hemorrhagic fever and neurological effects. The World Health Organization has identified Rift Valley fever as one of the viral infections that has potential to cause a future epidemic. Hence, efforts are urgently needed toward development of therapeutics and vaccine against this infectious disease. Notably, the causative virus namely, the Rift Valley fever virus (RVFV), utilizes the cap-snatching mechanism for viral transcription, rendering its cap-binding domain (CBD) as an effective antiviral target. To date, there are no published studies towards identification of potential small molecule inhibitors for the CBD of RVFV. Here, we employ a virtual screening workflow comprising of molecular docking and molecular dynamics (MD) simulation, to identify 5 potential phytochemical inhibitors of the CBD of RVFV. These 5 phytochemical inhibitors can be sourced from Indian medicinal plants, Ferula assa-foetida, Glycyrrhiza glabra and Leucas cephalotes, used in traditional medicine. In sum, the 5 phytochemical inhibitors of the CBD of RVFV identified by this purely computational study are promising drug lead molecules which can be considered for detailed experimental validation against RVFV infection.

Pseudotyped Viruses for Phlebovirus.

Rift Valley fever virus (RVFV) is a member of the Phlebovirus genus, one of the 20 genera in the Phenuiviridae family. RVFV causes disease in animals and humans and is transmitted by sandflies or ticks. However, research into RVFV is limited by the requirement for biosafety level 3 (BSL-3) containment. Pseudotyped virus overcomes this limitation as it can be handled in a BSL-2 environment. Pseudotyped RVFV possesses an identical envelope protein structure to that of the authentic virus, simulating the same process of receptor binding and membrane fusion to host cells. Pseudotyped phleboviruses are therefore useful tools to study the infection mechanism of these viruses and for the screening of inhibitory drugs and the development of therapeutic monoclonal antibodies.

Estimation of Rift Valley fever virus spillover to humans during the Mayotte 2018-2019 epidemic.

Rift Valley fever (RVF) is an emerging, zoonotic, arboviral hemorrhagic fever threatening livestock and humans mainly in Africa. RVF is of global concern, having expanded its geographical range over the last decades. The impact of control measures on epidemic dynamics using empirical data has not been assessed. Here, we fitted a mathematical model to seroprevalence livestock and human RVF case data from the 2018-2019 epidemic in Mayotte to estimate viral transmission among livestock, and spillover from livestock to humans through both direct contact and vector-mediated routes. Model simulations were used to assess the impact of vaccination on reducing the epidemic size. The rate of spillover by direct contact was about twice as high as vector transmission. Assuming 30% of the population were farmers, each transmission route contributed to 45% and 55% of the number of human infections, respectively. Reactive vaccination immunizing 20% of the livestock population reduced the number of human cases by 30%. Vaccinating 1 mo later required using 50% more vaccine doses for a similar reduction. Vaccinating only farmers required 10 times as more vaccine doses for a similar reduction in human cases. Finally, with 52.0% (95% credible interval [CrI] [42.9-59.4]) of livestock immune at the end of the epidemic wave, viral reemergence in the next rainy season (2019-2020) is unlikely. Coordinated human and animal health surveillance, and timely livestock vaccination appear to be key to controlling RVF in this setting. We furthermore demonstrate the value of a One Health quantitative approach to surveillance and control of zoonotic infectious diseases.

Seroprevalence of Rift Valley Fever virus in one-humped camels (Camelus dromedaries) in Egypt.

Rift Valley fever (RVF) is a mosquito-borne viral disease that affects a variety of domestic animals, including cattle, sheep, goats, and camels, and has zoonotic potential. Although the rift valley fever virus (RVFV) is usually asymptomatic in camels, it can induce abortion in some pregnant animals. In the current study, a serosurvey was carried out to investigate the prevalence of RVFV antibodies and related risk factors in camels from four Egyptian governorates. A total of 400 serum samples were examined for anti-RVFV antibodies using a competitive enzyme-linked immunosorbent assay (c-ELISA). The results revealed that the overall prevalence of RVF among examined camels was 21.5% and the disease was more prevalent in Kafr ElSheikh governorate in Nile Delta of Egypt. In addition, the age group of camels with more than 5 years (OR=4.49, 95%CI: 1.39-14.49), the female sex (OR=3.38, 95%CI: 1.51-7.58), the emaciated animals (OR=1.52, 95%CI: 0.86-2.66), the summer season's infection (OR=5.98, 95%CI: 1.79-19.93), the presence of mosquitoes (OR= 2.88, 95%CI: 1.39-5.95), and the absence of mosquitoes control (OR=3.97, 95%CI: 2.09-7.57) were identified as risk factors for RVFV infection. The results of this study support knowledge on the risk factors for RVFV infection and demonstrate that camels raising in Egypt have RVFV antibodies. Quarantine measures or vaccination program should be implemented to reduce the likelihood of RVFV introduction, dissemination among susceptible animals, and ultimately transmission to humans.

Genomic surveillance of Rift Valley fever virus: from sequencing to lineage assignment.

Genetic evolution of Rift Valley fever virus (RVFV) in Africa has been shaped mainly by environmental changes such as abnormal rainfall patterns and climate change that has occurred over the last few decades. These gradual environmental changes are believed to have effected gene migration from macro (geographical) to micro (reassortment) levels. Presently, 15 lineages of RVFV have been identified to be circulating within the Sub-Saharan Africa. International trade in livestock and movement of mosquitoes are thought to be responsible for the outbreaks occurring outside endemic or enzootic regions. Virus spillover events contribute to outbreaks as was demonstrated by the largest epidemic of 1977 in Egypt. Genomic surveillance of the virus evolution is crucial in developing intervention strategies. Therefore, we have developed a computational tool for rapidly classifying and assigning lineages of the RVFV isolates. The computational method is presented both as a command line tool and a web application hosted at https://www.genomedetective.com/app/typingtool/rvfv/ . Validation of the tool has been performed on a large dataset using glycoprotein gene (Gn) and whole genome sequences of the Large (L), Medium (M) and Small (S) segments of the RVFV retrieved from the National Center for Biotechnology Information (NCBI) GenBank database. Using the Gn nucleotide sequences, the RVFV typing tool was able to correctly classify all 234 RVFV sequences at species level with 100% specificity, sensitivity and accuracy. All the sequences in lineages A (n = 10), B (n = 1), C (n = 88), D (n = 1), E (n = 3), F (n = 2), G (n = 2), H (n = 105), I (n = 2), J (n = 1), K (n = 4), L (n = 8), M (n = 1), N (n = 5) and O (n = 1) were also correctly classified at phylogenetic level. Lineage assignment using whole RVFV genome sequences (L, M and S-segments) did not achieve 100% specificity, sensitivity and accuracy for all the sequences analyzed. We further tested our tool using genomic data that we generated by sequencing 5 samples collected following a recent RVF outbreak in Kenya. All the 5 samples were assigned lineage C by both the partial (Gn) and whole genome sequence classifiers. The tool is useful in tracing the origin of outbreaks and supporting surveillance efforts.Availability: https://github.com/ajodeh-juma/rvfvtyping.

Rift Valley Fever Outbreak during COVID-19 Surge, Uganda, 2021.

Rift Valley fever, endemic or emerging throughout most of Africa, causes considerable risk to human and animal health. We report 7 confirmed Rift Valley fever cases, 1 fatal, in Kiruhura District, Uganda, during 2021. Our findings highlight the importance of continued viral hemorrhagic fever surveillance, despite challenges associated with the COVID-19 pandemic.