RESUMO
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.
Assuntos
Interações Hospedeiro-Patógeno , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Vírus da Febre do Vale do Rift/fisiologia , Internalização do Vírus , Animais , Especificidade de Anticorpos/imunologia , Sequência de Bases , Encéfalo/patologia , Encéfalo/virologia , Sistemas CRISPR-Cas/genética , Membrana Celular/metabolismo , Células Cultivadas , Glicoproteínas/metabolismo , Glicosaminoglicanos/metabolismo , Glicosilação , Humanos , Proteína Associada a Proteínas Relacionadas a Receptor de LDL/metabolismo , Ligantes , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/deficiência , Glicoproteínas de Membrana/metabolismo , Camundongos , Ligação Proteica , Desnaturação Proteica , Febre do Vale de Rift/patologia , Febre do Vale de Rift/prevenção & controle , Febre do Vale de Rift/virologia , Vírus da Febre do Vale do Rift/imunologiaRESUMO
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.
Assuntos
Transmissão Vertical de Doenças Infecciosas , Placenta , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Vacinas Atenuadas , Vacinas Virais , Replicação Viral , Vírus da Febre do Vale do Rift/fisiologia , Vírus da Febre do Vale do Rift/imunologia , Animais , Feminino , Gravidez , Ovinos , Placenta/virologia , Humanos , Febre do Vale de Rift/virologia , Febre do Vale de Rift/transmissão , Vacinas Virais/imunologia , Doenças dos Ovinos/virologiaRESUMO
We analyzed hospital discharge records of patients with coccidioidomycosis-related codes from the International Classification of Diseases, 10th revision, Clinical Modification, to estimate the prevalence of hospital visits associated with the disease in Texas, USA. Using Texas Health Care Information Collection data for 2016-2021, we investigated the demographic characteristics and geographic distribution of the affected population, assessed prevalence of hospital visits for coccidioidomycosis, and examined how prevalence varied by demographic and geographic factors. In Texas, 709 coccidioidomycosis-related inpatient and outpatient hospital visits occurred in 2021; prevalence was 3.17 cases per 100,000 total hospital visits in 2020. Geographic location, patient sex, and race/ethnicity were associated with increases in coccidioidomycosis-related hospital visits; male, non-Hispanic Black, and Hispanic patients had the highest prevalence of coccidioidomycosis compared with other groups. Increased surveillance and healthcare provider education and outreach are needed to ensure timely and accurate diagnosis and treatment of coccidioidomycosis in Texas and elsewhere.
Assuntos
Coccidioidomicose , Coccidioidomicose/epidemiologia , Coccidioidomicose/diagnóstico , Humanos , Texas/epidemiologia , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Adolescente , Idoso , Adulto Jovem , Criança , Pré-Escolar , Lactente , Prevalência , Hospitalização/estatística & dados numéricos , História do Século XXI , Recém-Nascido , Idoso de 80 Anos ou maisRESUMO
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.
Assuntos
Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Ratos , Animais , Vírus da Febre do Vale do Rift/fisiologia , Citocinas , Encéfalo , Morte CelularRESUMO
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.
Assuntos
Vírus da Febre do Vale do Rift , Humanos , Bovinos , Ovinos/genética , Animais , Reação em Cadeia da Polimerase em Tempo Real/métodos , Transcrição Reversa , Laboratórios , RNA ViralRESUMO
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.
Assuntos
Hepatócitos , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Replicação Viral , Animais , Humanos , Camundongos , Hepatócitos/patologia , Hepatócitos/virologia , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Febre do Vale de Rift/virologia , Vírus da Febre do Vale do Rift/fisiologiaRESUMO
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.
Assuntos
Autofagia , Imunidade Inata , Nucleoproteínas , Vírus da Febre do Vale do Rift , Imunidade Inata/imunologia , Vírus da Febre do Vale do Rift/imunologia , Nucleoproteínas/imunologia , Nucleoproteínas/metabolismo , Autofagia/imunologia , Replicação Viral , Linhagem Celular , Febre do Vale de Rift/imunologia , Humanos , Animais , Macrófagos/virologiaRESUMO
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.
Assuntos
Homeostase , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Animais , Febre do Vale de Rift/patologia , Vírus da Febre do Vale do Rift/patogenicidade , Ovinos , Transcriptoma , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Fígado , Interações Hospedeiro-Patógeno , Interferons/metabolismoRESUMO
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.
Assuntos
Doenças Transmissíveis , Lipossomos , Nanopartículas , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Animais , Humanos , Vírus da Febre do Vale do Rift/genética , Febre do Vale de Rift/prevenção & controle , Pandemias , Anticorpos AntiviraisRESUMO
Coccidioides is an endemic fungus that causes infections ranging from mild respiratory illness to life-threatening disease, and immunocompromised hosts such as solid organ transplant recipients are at higher risk for disseminated infection and mortality. Our center administers fluconazole prophylaxis to kidney transplant recipients residing in geographic areas with higher incidences of coccidioidomycosis. However, because drug-drug interactions occur between triazoles and immunosuppressants used in transplant medicine, we undertook a study to ascertain whether fluconazole prophylaxis was associated with any important safety outcomes in kidney transplant recipients. This retrospective study evaluated patients who had undergone kidney transplantation between 2016 and 2019. Data on patient demographics, transplant-related clinical information, use of fluconazole prophylaxis (200 mg daily for 6-12 months post-transplant), and patient outcomes were obtained. The primary outcome was mean estimated glomerular filtration rate (eGFR) at 12 months, comparing those who received fluconazole prophylaxis to those who did not. Secondary outcomes included mean eGFR at 3 months, 6 months, and 9 months post-transplant, patient survival, biopsy-proven graft rejection, graft loss, or a new requirement for post-transplant dialysis, all within 12 months post-transplant. The mean eGFR at 12 months was similar between both groups, with 66.4 ml/min/1.73 m² in the fluconazole prophylaxis group vs. 64.3 ml/min/1.73 m² in the non-fluconazole prophylaxis group (P = 0.55). Secondary outcomes were similar across both groups. Multivariable linear regression found no significant association between fluconazole use and graft function. Fluconazole prophylaxis for prevention of coccidioidomycosis was not associated with adverse graft outcomes in kidney transplant recipients.
Solid organ transplant recipients can be highly immune suppressed, and infection with Coccidioides (valley fever) after transplant can lead to severe infections in these patients. Our study showed that fluconazole was safe and effective for preventing Coccidioides in kidney transplant recipients.
Assuntos
Coccidioidomicose , Transplante de Rim , Humanos , Fluconazol/efeitos adversos , Coccidioidomicose/epidemiologia , Coccidioidomicose/veterinária , Antifúngicos/efeitos adversos , Transplante de Rim/efeitos adversos , Transplante de Rim/veterinária , Estudos Retrospectivos , TransplantadosRESUMO
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.
Assuntos
Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Animais , Febre do Vale de Rift/diagnóstico , Camarões , Anticorpos Antivirais , Primatas , Estudos SoroepidemiológicosRESUMO
Rift Valley fever virus (RVFV) is a mosquito-borne RNA virus of the Phlebovirus genus in the phenuviridae family. Its genome is trisegmented with small (S), medium (M) and large (L) fragments. In nature, the virus exists as a single serotype that is responsible for outbreaks of Rift Valley fever (RVF), a zoonotic disease that often occurs in Africa and the Middle East. RVFV genomes are thought to undergo both recombination and reassortment and investigations of these events is important for monitoring the emergence of virulent strains and understanding the evolutionary characteristics of this virus. The aim of this study was to characterize the genomes of RVFV isolates from cattle, sheep, and goats collected during an interepidemic period in Kenya between June 2016 and November 2021. A total of 691 serum samples from cattle (n = 144), goats (n = 185) and sheep (n = 362) were analysed at the Central Veterinary Laboratories. The competitive IgM-capture ELISA, was used to screen the samples; 205 samples (29.67%) tested positive for RVFV. Of the 205 positive samples, 42 (20.5%) were from cattle, 57 (27.8%) from goats, and 106 (51.7%) from sheep. All the IgM-positive samples were further analyzed by qPCR, and 24 (11.71%) tested positive with Ct values ranging from 14.788 to 38.286. Two samples, 201808HABDVS from sheep and 201810CML3DVS from cattle, had Ct values of less than 20.0 and yielded whole genome sequences with 96.8 and 96.4 coverage, respectively. There was no statistically significant evidence of recombination in any of the three segments and also phylogenetic analysis showed no evidence of reassortment in the two isolated RVFV segments when compared with other isolates of different lineages from previous outbreaks whose genomes are deposited in the GenBank. No evidence of reassortment leaves room for other factors to be the most probable contributors of change in virulence, pathogenicity and emergence of highly virulent strains of the RVFV.
Assuntos
Doenças dos Bovinos , Genoma Viral , Doenças das Cabras , Cabras , Filogenia , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Doenças dos Ovinos , Animais , Cabras/virologia , Vírus da Febre do Vale do Rift/genética , Vírus da Febre do Vale do Rift/isolamento & purificação , Ovinos , Febre do Vale de Rift/virologia , Febre do Vale de Rift/epidemiologia , Bovinos , Quênia/epidemiologia , Doenças das Cabras/virologia , Doenças das Cabras/epidemiologia , Doenças dos Ovinos/virologia , Doenças dos Ovinos/epidemiologia , Doenças dos Bovinos/virologia , Doenças dos Bovinos/epidemiologia , Surtos de Doenças/veterináriaRESUMO
BACKGROUND: Coccidioidomycosis is a systemic fungal disease endemic to arid regions of the Western Hemisphere. In the south-western US, Coccidioides spp. may account for up to 20%-25% of all cases of community acquired pneumonia. Clinical manifestations vary widely, from asymptomatic infection to life-threatening disease, especially in immunocompromised hosts. OBJECTIVES: The primary objective of the study was to characterise cases of coccidioidomycosis in an area of the United States not considered traditionally endemic for the disease. METHODS: We performed a single-centre retrospective study of all cases of coccidioidomycosis from 1 January 2000 to 31 December 2020, in the University of Oklahoma Health Sciences Medical Center. RESULTS: A total of 26 patients were included for analysis. The central nervous system (CNS) and the lungs were the sites most frequently involved. Twenty (77%) had travelled to a coccidioidomycosis endemic region. Most were male (81%) with a median age of 42 years (range: 3-78 years). The majority (46%) were Caucasians, 19% were African American, 19% Hispanic, and 12% Native American. The most common comorbidities were diabetes mellitus and acquired immunodeficiency syndrome, identified in 27% and 23% of patients, respectively. Patients on immunosuppressive therapy accounted for 12% of all cases. CONCLUSION: Our study is one of the largest single-centre case series of coccidioidomycosis from a non-endemic area. Diabetes mellitus was the most frequent comorbidity. Compared to other case series of coccidioidomycosis, our patient population had higher rates of immunosuppression and had both a higher rate of disseminated disease and overall mortality.
Assuntos
Coccidioidomicose , Humanos , Coccidioidomicose/epidemiologia , Coccidioidomicose/microbiologia , Coccidioidomicose/tratamento farmacológico , Estudos Retrospectivos , Oklahoma/epidemiologia , Pessoa de Meia-Idade , Masculino , Feminino , Adulto , Idoso , Adolescente , Adulto Jovem , Criança , Pré-Escolar , Hospedeiro Imunocomprometido , Coccidioides/isolamento & purificação , ComorbidadeRESUMO
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.
Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Vírus da Febre do Vale do Rift/imunologia , Proteínas Virais de Fusão/imunologia , Animais , Anticorpos Monoclonais/imunologia , Linhagem Celular Tumoral , Células Cultivadas , Chlorocebus aethiops , Epitopos/química , Epitopos/imunologia , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Células Vero , Proteínas Virais de Fusão/químicaRESUMO
Coccidioidomycosis is a fungal infection with a range of clinical manifestations. Currently used antifungal agents exhibit variable efficacy and toxicity profiles that necessitate evaluation of additional therapeutic options. Improvement was observed in the majority of patients treated with isavuconazole, with clinical failures observed only in those with coccidioidal meningitis.
Assuntos
Coccidioidomicose , Humanos , Coccidioidomicose/tratamento farmacológico , Coccidioidomicose/microbiologia , Coccidioides , Triazóis/uso terapêutico , Antifúngicos/uso terapêuticoRESUMO
Coccidioides spp. are dimorphic fungi that are capable of infecting human and non-human mammals and can cause diverse manifestations of coccidioidomycosis or Valley fever (VF). In combination with clinical symptoms and radiographic findings, antibody-based diagnostic tests are often used to diagnose and monitor patients with VF. Chitinase 1 (CTS1) has previously been identified as the seroreactive antigen used in these diagnostic assays to detect anticoccidial IgG. Here, an indirect enzyme-linked immunosorbent assay to detect IgG to CTS1 demonstrated 165 of 178 (92.7%) patients with a positive result by immunodiffusion (ID) and/or complement fixation (CF) had antibodies to the single antigen CTS1. We then developed a rapid antibody lateral flow assay (LFA) to detect anti-CTS1 antibodies. Out of 143 samples tested, the LFA showed 92.9% positive percent agreement [95% confidence interval (CI), 84.3%-96.9%] and 97.7% negative percent agreement (95% CI, 87.9%-99.6%) with ID and CF assays. Serum or plasma from canines, macaques, and dolphins was also tested by the CTS1 LFA. Test line densities of the CTS1 LFA correlated in a linear manner with the reported CF and ID titers for human and non-human samples, respectively. This 10-min point-of-care test for the rapid detection of anti-coccidioidal antibodies could help to inform healthcare providers in real-time, potentially improving the efficiency of healthcare delivery.
Assuntos
Bioensaio , Coccidioidomicose , Humanos , Animais , Cães , Coccidioides , Coccidioidomicose/diagnóstico , Ensaio de Imunoadsorção Enzimática , Macaca , Imunoglobulina G , MamíferosRESUMO
Coccidioidomycosis is a fungal disease associated with soil exposure that frequently goes undiagnosed due at least in part to its nonspecific presentation and the lack of clinical suspicion by health care providers. Currently available diagnostics for coccidioidomycosis offer qualitative results that can suffer from low specificity, while semiquantitative assays are labor-intensive and complex and can require multiple days to complete. Furthermore, significant confusion exists regarding the optimal diagnostic algorithms and appropriate usage of available diagnostic tests. This review aims to inform clinical laboratorians and treating clinicians about the current diagnostic landscape, appropriate diagnostic strategies, and future diagnostic directions for coccidioidomycosis, which is expected to become more prevalent due to increased migration into areas of endemicity and climate changes.
Assuntos
Coccidioidomicose , Humanos , Coccidioidomicose/diagnóstico , Coccidioides , Anticorpos Antifúngicos , BioensaioRESUMO
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.
Assuntos
Técnicas Microbiológicas/métodos , Proteínas do Nucleocapsídeo/análise , Febre do Vale de Rift , Vírus da Febre do Vale do Rift , Idoso , Idoso de 80 Anos ou mais , Animais , Anticorpos Antivirais , Estudos Transversais , Feminino , Humanos , Imunoglobulina G , Gado , Masculino , Moçambique/epidemiologia , Febre do Vale de Rift/epidemiologia , Vírus da Febre do Vale do Rift/fisiologia , Estudos SoroepidemiológicosRESUMO
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.
Assuntos
Modelos Moleculares , Conformação Proteica , RNA Polimerase Dependente de RNA/química , Vírus da Febre do Vale do Rift/enzimologia , Motivos de Aminoácidos , Domínio Catalítico , Fenômenos Químicos , Sequência Conservada , Microscopia Crioeletrônica , Domínios e Motivos de Interação entre Proteínas , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Recombinantes de Fusão , Proteínas Virais/químicaRESUMO
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.