RESUMEN
BACKGROUND: Rift Valley fever virus (RVFV) is a zoonotic mosquito-borne virus with serious implications for livestock health, human health, and the economy in Africa, and is suspected to be endemic in north-eastern KwaZulu-Natal (KZN), South Africa. The vectors of RVFV in this area are poorly known, although several species, such as Aedes (Neomelaniconion) mcintoshi, Aedes (Neomelaniconion) circumluteolus, Aedes (Aedimorphus) durbanensis, and Culex (Lasioconops) poicilipes may be involved. The aim of the study was to determine the vertebrate blood meal sources of potential RVFV mosquito vectors in north-eastern KZN and to characterize the host-biting network. METHODS: Blood-fed mosquitoes were collected monthly from November 2019 to February 2023 using a backpack aspirator, CO2-baited Centers for Disease Control and Prevention (CDC) miniature light traps and tent traps, in the vicinity of water bodies and livestock farming households. The mosquitoes were morphologically identified. DNA was extracted from individual mosquitoes and used as templates to amplify the vertebrate cytochrome c oxidase I (COI) and cytochrome b (cytb) genes using conventional polymerase chain reaction (PCR). Amplicons were sequenced and queried in GenBank and the Barcode of Life Data systems to identify the vertebrate blood meal sources and confirm mosquito identifications. All mosquitoes were screened for RVFV using real time reverse transcription (RT)-PCR. RESULTS: We identified the mammalian (88.8%) and avian (11.3%) blood meal sources from 409 blood-fed mosquitoes. Aedes circumluteolus (n = 128) made up the largest proportion of collected mosquitoes. Cattle (n = 195) and nyala (n = 61) were the most frequent domestic and wild hosts, respectively. Bipartite network analysis showed that the rural network consisted of more host-biting interactions than the reserve network. All mosquitoes tested negative for RVFV. CONCLUSIONS: Several mosquito species, including Ae. circumluteolus, and vertebrate host species, including cattle and nyala, could play a central role in RVFV transmission. Future research in this region should focus on these species to better understand RVFV amplification.
Asunto(s)
Aedes , Mosquitos Vectores , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Animales , Sudáfrica , Mosquitos Vectores/virología , Mosquitos Vectores/fisiología , Virus de la Fiebre del Valle del Rift/genética , Virus de la Fiebre del Valle del Rift/aislamiento & purificación , Virus de la Fiebre del Valle del Rift/fisiología , Fiebre del Valle del Rift/transmisión , Fiebre del Valle del Rift/virología , Fiebre del Valle del Rift/epidemiología , Aedes/virología , Aedes/fisiología , Aedes/genética , Aedes/clasificación , Humanos , Conducta Alimentaria , Culex/virología , Culex/fisiología , Mordeduras y Picaduras de Insectos , Femenino , Culicidae/virología , Culicidae/fisiología , Culicidae/clasificaciónRESUMEN
Rift Valley fever virus (RVFV) infection causes abortions in ruminant livestock and is associated with an increased likelihood of miscarriages in women. Using sheep and human placenta explant cultures, we sought to identify tissues at the maternal-fetal interface targeted by RVFV. Sheep villi and fetal membranes were highly permissive to RVFV infection resulting in markedly higher virus titers than human cultures. Sheep cultures were most permissive to wild-type RVFV and ΔNSm infection, while live-attenuated RVFV vaccines (LAVs; MP-12, ΔNSs, and ΔNSs/ΔNSm) exhibited reduced replication. The human fetal membrane restricted wild-type and LAV replication, and when infection occurred, it was prominent on the maternal-facing side. Type I and type III interferons were induced in human villi exposed to LAVs lacking the NSs protein. This study supports the use of sheep and human placenta explants to understand vertical transmission of RVFV in mammals and whether LAVs are attenuated at the maternal-fetal interface.IMPORTANCEA direct comparison of replication of Rift Valley fever virus (RVFV) in sheep and human placental explants reveals comparative efficiencies and permissivity to infection and replication. Vaccine strains of RVFV demonstrated reduced infection and replication capacity in the mammalian placenta. This study represents the first direct cross-host comparison of the vertical transmission capacity of this high-priority emerging mosquito-transmitted virus.
Asunto(s)
Transmisión Vertical de Enfermedad Infecciosa , Placenta , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Vacunas Atenuadas , Vacunas Virales , Replicación Viral , Virus de la Fiebre del Valle del Rift/fisiología , Virus de la Fiebre del Valle del Rift/inmunología , Animales , Femenino , Embarazo , Ovinos , Placenta/virología , Humanos , Fiebre del Valle del Rift/virología , Fiebre del Valle del Rift/transmisión , Vacunas Virales/inmunología , Enfermedades de las Ovejas/virologíaRESUMEN
Rift Valley fever (RVF) is a re-emerging vector-borne zoonosis with a high public health and veterinary impact. In West Africa, many lineages were previously detected, but since 2020, lineage H from South Africa has been the main cause of the outbreaks. In this study, clinical samples collected through national surveillance were screened for RVF virus (RVFV) acute infection by RT-PCR and IgM ELISA tests. Sequencing, genome mapping and in vitro phenotypic characterization in mammal cells were performed on RT-PCR positive samples in comparison with other epidemic lineages (G and C). Four RVFV human cases were detected in Senegal and the sequence analyses revealed that the strains belonged to lineage H. The in vitro kinetics and genome mapping showed different replication efficiency profiles for the tested RVFV lineages and non-conservative mutations, which were more common to lineage G or specific to lineage H. Our findings showed the re-emergence of lineage H in Senegal in 2022, its high viral replication efficiency in vitro and support the findings that genetic diversity affects viral replication. This study gives new insights into the biological properties of lineage H and calls for deeper studies to better assess its potential to cause a future threat in Senegal.
Asunto(s)
Genoma Viral , Filogenia , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Replicación Viral , Virus de la Fiebre del Valle del Rift/genética , Virus de la Fiebre del Valle del Rift/aislamiento & purificación , Virus de la Fiebre del Valle del Rift/clasificación , Virus de la Fiebre del Valle del Rift/fisiología , Fiebre del Valle del Rift/virología , Fiebre del Valle del Rift/epidemiología , Fiebre del Valle del Rift/transmisión , Senegal/epidemiología , Humanos , Animales , Enfermedades Transmisibles Emergentes/virología , Enfermedades Transmisibles Emergentes/epidemiología , Enfermedades Transmisibles Emergentes/veterinaria , Brotes de Enfermedades , África Occidental/epidemiología , Variación Genética , MutaciónRESUMEN
Vector competence assays allow to measure, in the laboratory, the ability of a mosquito to get infected and then retransmit an arbovirus while mimicking natural vector infection route. Aedes aegypti is a major vector of arboviruses worldwide and thus a reference species used in vector competence assays. Rift Valley fever virus (RVFV) is a major public health threat, mostly in Africa, that infects humans and animals through the bite of mosquito vectors. Here, we describe vector competence assay of Aedes aegypti mosquitoes for RVFV, from mosquito exposure to the virus through an infectious artificial blood meal to the measurement of virus prevalence in the mosquito's body, head, and saliva.
Asunto(s)
Aedes , Mosquitos Vectores , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Animales , Aedes/virología , Virus de la Fiebre del Valle del Rift/fisiología , Virus de la Fiebre del Valle del Rift/aislamiento & purificación , Mosquitos Vectores/virología , Fiebre del Valle del Rift/transmisión , Fiebre del Valle del Rift/virología , Saliva/virología , HumanosRESUMEN
RNAseq is a valuable tool that can aid researchers in uncovering the transcriptional changes that occur when a viral pathogen infects a host cell. Viral infection will invariably cause differential expression of many genes, from transcription of mRNA to alternative splicing and degradation. This change in gene expression can be a result of immune activation or a direct activity of the virus to alter the host cell's environment to make it more favorable for viral replication. Studying the innate immune response to a pathogen can reveal which cellular pathways are active, indicating the steps that the host takes to halt viral infection, and detecting virus-mediated mRNA expression changes can help with identifying the pathways which may be exploited by the virus. Gene expression changes-both cell-caused and virus-caused-can be studied through RNAseq, helping to provide a clearer picture of the cellular changes that occur during viral infection. In this protocol, we outline methods to carry out mRNA sequencing in Rift Valley fever virus-infected cell cultures, from infection to library prep and analysis.
Asunto(s)
Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift/genética , Virus de la Fiebre del Valle del Rift/fisiología , Humanos , Fiebre del Valle del Rift/virología , Fiebre del Valle del Rift/genética , Interacciones Huésped-Patógeno/genética , Análisis de Secuencia de ARN/métodos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Animales , Replicación Viral/genética , Empalme Alternativo , Empalme del ARN , Transcripción Genética , Línea CelularRESUMEN
Rift Valley fever virus (RVFV) is an arbovirus in the Phenuiviridae family identified initially by the large 'abortion storms' observed among ruminants; RVFV can also infect humans. In humans, there is a wide variation of clinical symptoms ranging from subclinical to mild febrile illness to hepatitis, retinitis, delayed-onset encephalitis, or even hemorrhagic fever. The RVFV is a tri-segmented negative-sense RNA virus consisting of S, M, and L segments. The L segment encodes the RNA-dependent RNA polymerase (RdRp), termed the L protein, which is responsible for both viral mRNA synthesis and genome replication. Phosphorylation of viral RdRps is known to regulate viral replication. This study shows that RVFV L protein is serine phosphorylated and identified Casein Kinase 1 alpha (CK1α) and protein phosphatase 1 alpha (PP1α) as L protein binding partners. Inhibition of CK1 and PP1 through small molecule inhibitor treatment, D4476 and 1E7-03, respectively, caused a change in the phosphorylated status of the L protein. Inhibition of PP1α resulted in increased L protein phosphorylation whereas inhibition of CK1α decreased L protein phosphorylation. It was also found that in RVFV infected cells, PP1α localized to the cytoplasmic compartment. Treatment of RVFV infected cells with CK1 inhibitors reduced virus production in both mammalian and mosquito cells. Lastly, inhibition of either CK1 or PP1 reduced viral genomic RNA levels. These data indicate that L protein is phosphorylated and that CK1 and PP1 play a crucial role in regulating the L protein phosphorylation cycle, which is critical to viral RNA production and viral replication.
Asunto(s)
Proteína Fosfatasa 1 , Virus de la Fiebre del Valle del Rift , Replicación Viral , Virus de la Fiebre del Valle del Rift/fisiología , Virus de la Fiebre del Valle del Rift/genética , Fosforilación , Humanos , Animales , Proteína Fosfatasa 1/metabolismo , Proteína Fosfatasa 1/genética , Genoma Viral , Proteínas Virales/metabolismo , Proteínas Virales/genética , Caseína Quinasa Ialfa/metabolismo , Caseína Quinasa Ialfa/genética , Chlorocebus aethiops , Línea Celular , ARN Polimerasa Dependiente del ARN/metabolismo , ARN Polimerasa Dependiente del ARN/genética , Células Vero , ARN Viral/genética , ARN Viral/metabolismo , Fiebre del Valle del Rift/virologíaRESUMEN
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 20â% of severe cases. Importantly, individuals displaying neurologic disease have up to a 53â% 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.
Asunto(s)
Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Ratas , Animales , Virus de la Fiebre del Valle del Rift/fisiología , Citocinas , Encéfalo , Muerte CelularRESUMEN
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.
Asunto(s)
Hepatocitos , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Replicación Viral , Animales , Humanos , Ratones , Hepatocitos/patología , Hepatocitos/virología , Ratones Endogámicos C57BL , MicroARNs/genética , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/fisiologíaRESUMEN
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.
Asunto(s)
Oftalmopatías , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Ratas , Humanos , Animales , Virus de la Fiebre del Valle del Rift/fisiología , Ratas Sprague-Dawley , Inflamación , Citocinas , Aerosoles , CegueraRESUMEN
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.
Asunto(s)
Técnicas Microbiológicas/métodos , Proteínas de la Nucleocápside/análisis , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Anciano , Anciano de 80 o más Años , Animales , Anticuerpos Antivirales , Estudios Transversales , Femenino , Humanos , Inmunoglobulina G , Ganado , Masculino , Mozambique/epidemiología , Fiebre del Valle del Rift/epidemiología , Virus de la Fiebre del Valle del Rift/fisiología , Estudios SeroepidemiológicosRESUMEN
Rift Valley fever (RVF) is a zoonotic disease caused by RVF Phlebovirus (RVFV). The RVFV MP-12 vaccine strain is known to exhibit residual virulence in the case of a deficient interferon type 1 response. The hypothesis of this study is that virus replication and severity of lesions induced by the MP-12 strain in immunocompromised mice depend on the specific function of the disturbed pathway. Therefore, 10 strains of mice with deficient innate immunity (B6-IFNARtmAgt, C.129S7(B6)-Ifngtm1Ts/J, B6-TLR3tm1Flv, B6-TLR7tm1Aki, NOD/ShiLtJ), helper T-cell- (CD4tm1Mak), cytotoxic T-cell- (CD8atm1Mak), B-cell- (Igh-Jtm1DhuN?+N2), combined T- and B-cell- (NU/J) and combined T-, B-, natural killer (NK) cell- and macrophage-mediated immunity (NOD.Cg-PrkdcscidIl2rgtm1WjI/SzJ (NSG) mice) were subcutaneously infected with RVFV MP-12. B6-IFNARtmAgt mice were the only strain to develop fatal disease due to RVFV-induced severe hepatocellular necrosis and apoptosis. Notably, no clinical disease and only mild multifocal hepatocellular necrosis and apoptosis were observed in NSG mice, while immunohistochemistry detected the RVFV antigen in the liver and the brain. No or low virus expression and no lesions were observed in the other mouse strains. Conclusively, the interferon type 1 response is essential for early control of RVFV replication and disease, whereas functional NK cells, macrophages and lymphocytes are essential for virus clearance.
Asunto(s)
Inmunidad Adaptativa , Inmunidad Innata , Fiebre del Valle del Rift/inmunología , Virus de la Fiebre del Valle del Rift/fisiología , Animales , Apoptosis , Femenino , Humanos , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/virología , Hígado/inmunología , Hígado/virología , Macrófagos/inmunología , Macrófagos/virología , Masculino , Ratones , Ratones Endogámicos NOD , Fiebre del Valle del Rift/genética , Fiebre del Valle del Rift/fisiopatología , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/genética , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/virología , Linfocitos T Colaboradores-Inductores/inmunología , Linfocitos T Colaboradores-Inductores/virologíaRESUMEN
Rift Valley fever virus (RVFV) is a negative-sense, tripartite RNA virus that is endemic to Africa and the Arabian Peninsula. It can cause severe disease and mortality in humans and domestic livestock and is a concern for its potential to spread more globally. RVFV's nucleocapsid protein (N) is an RNA-binding protein that is necessary for viral transcription, replication, and the production of nascent viral particles. We have conducted crosslinking, immunoprecipitation, and sequencing (CLIP-seq) to characterize N interactions with host and viral RNAs during infection. In parallel, to precisely measure intracellular N levels, we employed multiple reaction monitoring mass spectrometry (MRM-MS). Our results show that N binds mostly to host RNAs at early stages of infection, yielding nascent virus particles of reduced infectivity. The expression of N plateaus 10 h post-infection, whereas the intracellular viral RNA concentration continues to increase. Moreover, the virions produced later in infection have higher infectivity. Taken together, the detailed examination of these N-RNA interactions provides insight into how the regulated expression of N and viral RNA produces both infectious and incomplete, noninfectious particles.
Asunto(s)
Cápside/metabolismo , Proteínas de la Nucleocápside/metabolismo , ARN Viral/metabolismo , Virus de la Fiebre del Valle del Rift/fisiología , Empaquetamiento del Genoma Viral , Animales , Chlorocebus aethiops , Células HEK293 , Humanos , Proteínas de la Nucleocápside/genética , Unión Proteica , ARN/metabolismo , ARN sin Sentido/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/metabolismo , Células Vero , Virión/metabolismoRESUMEN
Rift Valley fever (RVF) is a major emerging arboviral disease with a complex epidemiological cycle. RVF virus (RVFV) is transmitted by mosquito vectors to ruminants, causing epizootics, and then from animals to humans, triggering epidemics. During its cycle, RVFV infects a wide range of hosts, but the associated pathogenesis has yet to be elucidated. RVFV displays a predominant hepatic tropism, but also has a multicellular tropism inducing physiopathological effects in several tissues. However, there is variability between species in terms of physiopathology : a common clinical picture is found (severe hepatitis, hemorrhages, leukopenia), but certain forms are mainly found in humans (neurological and ocular damage) or in ruminant herds (waves of abortions). Although the molecular mechanisms involved are still poorly understood, it seems that early inflammatory response is related to the severity of the pathology. A better understanding of the pathogenesis of RVFV seems essential, especially since no specific treatment exists to date.
Asunto(s)
Epidemias , Fiebre del Valle del Rift , Virus de la Fiebre del Valle del Rift , Animales , Humanos , Mosquitos Vectores , Fiebre del Valle del Rift/epidemiología , Virus de la Fiebre del Valle del Rift/fisiología , RumiantesRESUMEN
Rift Valley fever virus (RVFV) causes morbidity and mortality in humans and domestic ungulates in sub-Saharan Africa, Egypt, and the Arabian Peninsula. Mosquito vectors transmit RVFV between vertebrates by bite, and also vertically to produce infectious progeny. Arrival of RVFV into the United States by infected mosquitoes or humans could result in significant impacts on food security, human health, and wildlife health. Elucidation of the vectors involved in the post-introduction RVFV ecology is paramount to rapid implementation of vector control. We performed vector competence experiments in which field-collected mosquitoes were orally exposed to an epidemic strain of RVFV via infectious blood meals. We targeted floodwater Aedes species known to feed on cattle, and/or deer species (Aedes melanimon Dyar, Aedes increpitus Dyar, Aedes vexans [Meigen]). Two permanent-water-breeding species were targeted as well: Culiseta inornata (Williston) of unknown competence considering United States populations, and Culex tarsalis Coquillett as a control species for which transmission efficiency is known. We tested the potential for midgut infection, midgut escape (dissemination), ovarian infection (vertical transmission), and transmission by bite (infectious saliva). Tissues were assayed by plaque assay and RT-qPCR, to quantify infectious virus and confirm virus identity. Tissue infection data were analyzed using a within-host model under a Bayesian framework to determine the probabilities of infection outcomes (midgut-limited infection, disseminated infection, etc.) while estimating barriers to infection between tissues. Permanent-water-breeding mosquitoes (Cx. tarsalis and Cs. inornata) exhibited more efficient horizontal transmission, as well as potential for vertical transmission, which is contrary to the current assumptions of RVFV ecology. Barrier estimates trended higher for Aedes spp., suggesting systemic factors in the differences between these species and Cx. tarsalis and Cs. inornata. These data indicate higher potential for vertical transmission than previously appreciated, and support the consensus of RVFV transmission including a broad range of potential vectors.
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Aedes/virología , Culex/virología , Mosquitos Vectores/virología , Fiebre del Valle del Rift/transmisión , Virus de la Fiebre del Valle del Rift/fisiología , Aedes/genética , Aedes/fisiología , Animales , Bovinos/virología , Colorado , Culex/fisiología , Ciervos/virología , Mosquitos Vectores/clasificación , Mosquitos Vectores/fisiología , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/genética , Virus de la Fiebre del Valle del Rift/aislamiento & purificación , Saliva/virologíaRESUMEN
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.
Asunto(s)
Interacciones Huésped-Patógeno , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Virus de la Fiebre del Valle del Rift/fisiología , Internalización del Virus , Animales , Especificidad de Anticuerpos/inmunología , Secuencia de Bases , Encéfalo/patología , Encéfalo/virología , Sistemas CRISPR-Cas/genética , Membrana Celular/metabolismo , Células Cultivadas , Glicoproteínas/metabolismo , Glicosaminoglicanos/metabolismo , Glicosilación , Humanos , Proteína Asociada a Proteínas Relacionadas con Receptor de LDL/metabolismo , Ligandos , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad/deficiencia , Glicoproteínas de Membrana/metabolismo , Ratones , Unión Proteica , Desnaturalización Proteica , Fiebre del Valle del Rift/patología , Fiebre del Valle del Rift/prevención & control , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/inmunologíaRESUMEN
Rift Valley fever virus (RVFV) is an arbovirus found throughout Africa. It causes disease that is typically mild and self-limiting; however, some infected individuals experience severe manifestations, including hepatitis, encephalitis, or even death. Reports of RVFV encephalitis are notable among immunosuppressed individuals, suggesting a role for adaptive immunity in preventing this severe complication. This phenomenon has been modeled in C57BL/6 mice depleted of CD4 T cells prior to infection with DelNSs RVFV (RVFV containing a deletion of nonstructural protein NSs), resulting in late-onset encephalitis accompanied by high levels of viral RNA in the brain in 30% of animals. In this study, we sought to define the specific type(s) of CD4 T cells that mediate protection from RVFV encephalitis. The viral epitopes targeted by CD4 and CD8 T cells were defined in C57BL/6 mice, and tetramers for both CD4 and CD8 T cells were generated. RVFV-specific CD8 T cells were expanded and of a cytotoxic and proliferating phenotype in the liver following infection. RVFV-specific CD4 T cells were identified in the liver and spleen following infection and phenotyped as largely Th1 or Tfh subtypes. Knockout mice lacking various aspects of pathways important in Th1 and Tfh development and function were used to demonstrate that T-bet, CD40, CD40L, and major histocompatibility complex class II (MHC-II) mediated protection from RVFV encephalitis, while gamma interferon (IFN-γ) and interleukin-12 (IL-12) were dispensable. Virus-specific antibody responses correlated with protection from encephalitis in all mouse strains, suggesting that Tfh/B cell interactions modulate clinical outcome in this model. IMPORTANCE The prevention of RVFV encephalitis requires intact adaptive immunity. In this study, we developed reagents to detect RVFV-specific T cells and provide evidence for Tfh cells and CD40/CD40L interactions as critical mediators of this protection.
Asunto(s)
Antígenos CD40 , Ligando de CD40 , Encefalitis Viral/prevención & control , Fiebre del Valle del Rift/inmunología , Virus de la Fiebre del Valle del Rift/inmunología , Virus de la Fiebre del Valle del Rift/fisiología , Linfocitos T/inmunología , África , Animales , Formación de Anticuerpos , Linfocitos B/inmunología , Encéfalo/virología , Linfocitos T CD8-positivos/inmunología , Modelos Animales de Enfermedad , Encefalitis Viral/inmunología , Encefalitis Viral/virología , Epítopos , Femenino , Hígado/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
The persistence mechanisms of Rift Valley fever (RVF), a zoonotic arboviral haemorrhagic fever, at both local and broader geographical scales have yet to be fully understood and rigorously quantified. We developed a mathematical metapopulation model describing RVF virus transmission in livestock across the four islands of the Comoros archipelago, accounting for island-specific environments and inter-island animal movements. By fitting our model in a Bayesian framework to 2004-2015 surveillance data, we estimated the importance of environmental drivers and animal movements on disease persistence, and tested the impact of different control scenarios on reducing disease burden throughout the archipelago. Here we report that (i) the archipelago network was able to sustain viral transmission in the absence of explicit disease introduction events after early 2007, (ii) repeated outbreaks during 2004-2020 may have gone under-detected by local surveillance, and (iii) co-ordinated within-island control measures are more effective than between-island animal movement restrictions.
Asunto(s)
Modelos Teóricos , Fiebre del Valle del Rift/prevención & control , Fiebre del Valle del Rift/transmisión , Virus de la Fiebre del Valle del Rift/fisiología , Animales , Comoras/epidemiología , Ganado/virología , Fiebre del Valle del Rift/epidemiología , Estudios Seroepidemiológicos , Zoonosis/epidemiología , Zoonosis/prevención & control , Zoonosis/transmisiónRESUMEN
Rift Valley fever virus (RVFV) is a mosquito-borne bunyavirus with a wide host range including ruminants and humans. RVFV outbreaks have had devastating effects on public health and the livestock industry in African countries. However, there is no approved RVFV vaccine for human use in non-endemic countries and no FDA-approved antiviral drug for RVFV treatment. The RVFV 78kDa protein (P78), which is a membrane glycoprotein, plays a role in virus dissemination in the mosquito host, but its biological role in mammalian hosts remains unknown. We generated an attenuated RVFV MP-12 strain-derived P78-High virus and a virulent ZH501 strain-derived ZH501-P78-High virus, both of which expressed a higher level of P78 and carried higher levels of P78 in the virion compared to their parental viruses. We also generated another MP-12-derived mutant virus (P78-KO virus) that does not express P78. MP-12 and P78-KO virus replicated to similar levels in fibroblast cell lines and Huh7 cells, while P78-High virus replicated better than MP-12 in Vero E6 cells, fibroblast cell lines, and Huh7 cells. Notably, P78-High virus and P78-KO virus replicated less efficiently and more efficiently, respectively, than MP-12 in macrophage cell lines. ZH501-P78-High virus also replicated poorly in macrophage cell lines. Our data further suggest that inefficient binding of P78-High virus to the cells led to inefficient virus internalization, low virus infectivity and reduced virus replication in a macrophage cell line. P78-High virus and P78-KO virus showed lower and higher virulence than MP-12, respectively, in young mice. ZH501-P78-High virus also exhibited lower virulence than ZH501 in mice. These data suggest that high levels of P78 expression attenuate RVFV virulence by preventing efficient virus replication in macrophages. Genetic alteration leading to increased P78 expression may serve as a novel strategy for the attenuation of RVFV virulence and generation of safe RVFV vaccines.
Asunto(s)
Macrófagos/virología , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/fisiología , Proteínas del Envoltorio Viral/metabolismo , Replicación Viral/fisiología , Animales , Ratones , Virus de la Fiebre del Valle del Rift/patogenicidad , Proteínas del Envoltorio Viral/genética , VirulenciaRESUMEN
The Government of Rwanda reported an outbreak of Rift Valley fever (RVF) in the Eastern Province in 2018. To respond to the outbreak, vaccination and education campaigns about the disease were carried out. Because RVF cases continue to be detected in Rwanda and the disease impacts livelihoods and health, accurate knowledge and communication are imperative. The objectives of this study were to evaluate knowledge and risk perceptions of RVF transmission among livestock farmers in Nyagatare District, Eastern Province, Rwanda, and to compare RVF knowledge, risk perceptions, and farming practices between male and female livestock farmers. This cross-sectional, quantitative study was conducted in selected sectors of Nyagatare District in the Eastern Province of Rwanda in June 2019. A 34-question survey was used to ask about demographics, livestock ownership, risk perceptions about zoonotic diseases and livestock management, RVF knowledge, preferred communication sources and information sharing strategies, and protective strategies for RVF mitigation while working with livestock. Livestock farmers were interviewed at three milk collection centers, two village meeting points, a farm cooperative meeting, and during door-to-door visits in villages. In total, 123 livestock farmers were interviewed. The survey found that most livestock farmers lacked knowledge about epizootic and zoonotic transmission of RVF, more male livestock farmers were familiar with RVF and risk mitigation strategies, and female livestock farmers are not viewed as reliable sources of information. Additionally, most livestock farmers had not vaccinated their animals against RVF despite past vaccination campaigns. Radio was the most popular communication channel. These findings show that RVF knowledge and information sharing are inadequate among livestock farmers in Eastern Province. Therefore, vaccination and education campaigns may need to be reevaluated within the context of these trends in order to prepare for future RVF outbreaks.
Asunto(s)
Agricultores/psicología , Conocimientos, Actitudes y Práctica en Salud , Fiebre del Valle del Rift/psicología , Zoonosis Virales/psicología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Comunicación , Estudios Transversales , Femenino , Humanos , Difusión de la Información , Ganado/crecimiento & desarrollo , Masculino , Persona de Mediana Edad , Percepción , Fiebre del Valle del Rift/virología , Virus de la Fiebre del Valle del Rift/fisiología , Rwanda , Zoonosis Virales/virología , Adulto JovenRESUMEN
Viral interference is a common occurrence that has been reported in cell culture in many cases. In the present study, viral interference between two capripox viruses (sheeppox SPPV and lumpy skin disease virus LSDV in cattle) with Rift Valley fever virus (RVFV) was investigated in vitro and in their natural hosts, sheep and cattle. A combination of SPPV/RVFV and LSDV/RVFV was used to co-infect susceptible cells and animals to detect potential competition. In-vitro interference was evaluated by estimating viral infectivity and copies of viral RNA by a qPCR during three serial passages in cell cultures, whereas in-vivo interference was assessed through antibody responses to vaccination. When lamb testis primary cells were infected with the mixture of capripox and RVFV, the replication of both SPPV and LSDV was inhibited by RVFV. In animals, SPPV/RVFV or LSDV/RVFV combinations inhibited the replication SPPV and LSDV and the antibody response following vaccination. The combined SPPV/RVFV did not protect sheep after challenging with the virulent strain of SPPV and the LSDV/RVFV did not induce interferon Gamma to LSDV, while immunological response to RVFV remain unaffected. Our goal was to assess this interference response to RVFV/capripoxviruses' coinfection in order to develop effective combined live-attenuated vaccines as a control strategy for RVF and SPP/LSD diseases. Our findings indicated that this approach was not suitable for developing a combined SPPV/LSDV/RVFV vaccine candidate because of interference of replication and the immune response among these viruses.