RESUMEN
Lassa virus infects hundreds of thousands of people each year across rural West Africa, resulting in a high number of cases of Lassa fever (LF), a febrile disease associated with high morbidity and significant mortality. The lack of approved treatments or interventions underscores the need for an effective vaccine. At least four viral lineages circulate in defined regions throughout West Africa with substantial interlineage nucleotide and amino acid diversity. An effective vaccine should be designed to elicit Lassa virus specific humoral and cell mediated immunity across all lineages. Most current vaccine candidates use only lineage IV antigens encoded by Lassa viruses circulating around Sierra Leone, Liberia, and Guinea but not Nigeria where lineages I-III are found. As previous infection is known to protect against disease from subsequent exposure, we sought to determine whether LF survivors from Nigeria and Sierra Leone harbor memory T cells that respond to lineage IV antigens. Our results indicate a high degree of cross-reactivity of CD8+ T cells from Nigerian LF survivors to lineage IV antigens. In addition, we identified regions within the Lassa virus glycoprotein complex and nucleoprotein that contributed to these responses while T cell epitopes were not widely conserved across our study group. These data are important for current efforts to design effective and efficient vaccine candidates that can elicit protective immunity across all Lassa virus lineages.
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Antígenos Virales/inmunología , Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/inmunología , Virus Lassa/inmunología , África Occidental , Reacciones Cruzadas , Femenino , Humanos , Masculino , Especificidad de la EspecieRESUMEN
During 2018, an unusual increase in Lassa fever cases occurred in Nigeria, raising concern among national and international public health agencies. We analyzed 220 Lassa virus genomes from infected patients, including 129 from the 2017-2018 transmission season, to understand the viral populations underpinning the increase. A total of 14 initial genomes from 2018 samples were generated at Redeemer's University in Nigeria, and the findings were shared with the Nigerian Center for Disease Control in real time. We found that the increase in cases was not attributable to a particular Lassa virus strain or sustained by human-to-human transmission. Instead, the data were consistent with ongoing cross-species transmission from local rodent populations. Phylogenetic analysis also revealed extensive viral diversity that was structured according to geography, with major rivers appearing to act as barriers to migration of the rodent reservoir.
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Genoma Viral , Fiebre de Lassa/virología , Virus Lassa/genética , ARN Viral/análisis , Adolescente , Adulto , Animales , Teorema de Bayes , Reservorios de Enfermedades , Femenino , Variación Genética , Humanos , Fiebre de Lassa/epidemiología , Fiebre de Lassa/transmisión , Masculino , Cadenas de Markov , Persona de Mediana Edad , Nigeria/epidemiología , Filogenia , Filogeografía , Roedores , Análisis de Secuencia de ARN , Zoonosis/transmisiónRESUMEN
Early and robust T cell responses have been associated with survival from Lassa fever (LF), but the Lassa virus-specific memory responses have not been well characterized. Regions within the virus surface glycoprotein (GPC) and nucleoprotein (NP) are the main targets of the Lassa virus-specific T cell responses, but, to date, only a few T cell epitopes within these proteins have been identified. We identified GPC and NP regions containing T cell epitopes and HLA haplotypes from LF survivors and used predictive HLA-binding algorithms to identify putative epitopes, which were then experimentally tested using autologous survivor samples. We identified 12 CD8-positive (CD8+) T cell epitopes, including epitopes common to both Nigerian and Sierra Leonean survivors. These data should be useful for the identification of dominant Lassa virus-specific T cell responses in Lassa fever survivors and vaccinated individuals as well as for designing vaccines that elicit cell-mediated immunity.IMPORTANCE The high morbidity and mortality associated with clinical cases of Lassa fever, together with the lack of licensed vaccines and limited and partially effective interventions, make Lassa virus (LASV) an important health concern in its regions of endemicity in West Africa. Previous infection with LASV protects from disease after subsequent exposure, providing a framework for designing vaccines to elicit similar protective immunity. Multiple major lineages of LASV circulate in West Africa, and therefore, ideal vaccine candidates should elicit immunity to all lineages. We therefore sought to identify common T cell epitopes between Lassa fever survivors from Sierra Leone and Nigeria, where distinct lineages circulate. We identified three such epitopes derived from highly conserved regions within LASV proteins. In this process, we also identified nine other T cell epitopes. These data should help in the design of an effective pan-LASV vaccine.
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Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/química , Fiebre de Lassa/inmunología , Virus Lassa/inmunología , Nucleoproteínas/inmunología , Proteínas del Envoltorio Viral/inmunología , Adolescente , Secuencia de Aminoácidos , Animales , Anticuerpos Antivirales/biosíntesis , Antígenos Virales/química , Antígenos Virales/genética , Antígenos Virales/inmunología , Linfocitos T CD8-positivos/virología , Niño , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Femenino , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/inmunología , Antígenos HLA-DQ/genética , Antígenos HLA-DQ/inmunología , Haplotipos , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Humanos , Sueros Inmunes/análisis , Memoria Inmunológica , Fiebre de Lassa/genética , Fiebre de Lassa/patología , Virus Lassa/patogenicidad , Masculino , Nigeria , Nucleoproteínas/genética , Sierra Leona , Sobrevivientes , Proteínas del Envoltorio Viral/genética , Adulto JovenRESUMEN
Lassa virus is genetically diverse with several lineages circulating in West Africa. This study aimed at describing the sequence variability of Lassa virus across Nigeria and inferring its spatiotemporal evolution. We sequenced and isolated 77 Lassa virus strains from 16 Nigerian states. The final data set, including previous works, comprised metadata and sequences of 219 unique strains sampled between 1969 and 2018 in 22 states. Most of this data originated from Lassa fever patients diagnosed at Irrua Specialist Teaching Hospital, Edo State, Nigeria. The majority of sequences clustered with the main Nigerian lineages II and III, while a few sequences formed a new cluster related to Lassa virus strains from Hylomyscus pamfi Within lineages II and III, seven and five sublineages, respectively, were distinguishable. Phylogeographic analysis suggests an origin of lineage II in the southeastern part of the country around Ebonyi State and a main vector of dispersal toward the west across the Niger River, through Anambra, Kogi, Delta, and Edo into Ondo State. The frontline of virus dispersal appears to be in Ondo. Minor vectors are directed northeast toward Taraba and Adamawa and south toward Imo and Rivers. Lineage III might have spread from northern Plateau State into Kaduna, Nasarawa, Federal Capital Territory, and Bauchi. One sublineage moved south and crossed the Benue River into Benue State. This study provides a geographic mapping of lineages and phylogenetic clusters in Nigeria at a higher resolution. In addition, we estimated the direction and time frame of virus dispersal in the country.IMPORTANCE Lassa virus is the causative agent of Lassa fever, a viral hemorrhagic fever with a case fatality rate of approximately 30% in Africa. Previous studies disclosed a geographical pattern in the distribution of Lassa virus strains and a westward movement of the virus across West Africa during evolution. Our study provides a deeper understanding of the geography of genetic lineages and sublineages of the virus in Nigeria. In addition, we modeled how the virus spread in the country. This knowledge allows us to predict into which geographical areas the virus might spread in the future and prioritize areas for Lassa fever surveillance. Our study not only aimed to generate Lassa virus sequences from across Nigeria but also to isolate and conserve the respective viruses for future research. Both isolates and sequences are important for the development and evaluation of medical countermeasures to treat and prevent Lassa fever, such as diagnostics, therapeutics, and vaccines.
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Fiebre de Lassa/virología , Virus Lassa/clasificación , Animales , Evolución Molecular , Variación Genética , Humanos , Fiebre de Lassa/epidemiología , Fiebre de Lassa/transmisión , Virus Lassa/genética , Murinae/virología , Nigeria/epidemiología , Filogenia , FilogeografíaAsunto(s)
Fiebre de Lassa/epidemiología , Complicaciones Infecciosas del Embarazo/epidemiología , Complicaciones Infecciosas del Embarazo/virología , Adolescente , Adulto , Manejo de la Enfermedad , Femenino , Edad Gestacional , Humanos , Fiebre de Lassa/diagnóstico , Fiebre de Lassa/tratamiento farmacológico , Virus Lassa , Nigeria/epidemiología , Embarazo , Complicaciones Infecciosas del Embarazo/tratamiento farmacológico , Resultado del Embarazo , Vigilancia en Salud Pública , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Evaluación de Síntomas , Adulto JovenRESUMEN
Infection with Lassa virus (LASV) can cause Lassa fever, a haemorrhagic illness with an estimated fatality rate of 29.7%, but causes no or mild symptoms in many individuals. Here, to investigate whether human genetic variation underlies the heterogeneity of LASV infection, we carried out genome-wide association studies (GWAS) as well as seroprevalence surveys, human leukocyte antigen typing and high-throughput variant functional characterization assays. We analysed Lassa fever susceptibility and fatal outcomes in 533 cases of Lassa fever and 1,986 population controls recruited over a 7 year period in Nigeria and Sierra Leone. We detected genome-wide significant variant associations with Lassa fever fatal outcomes near GRM7 and LIF in the Nigerian cohort. We also show that a haplotype bearing signatures of positive selection and overlapping LARGE1, a required LASV entry factor, is associated with decreased risk of Lassa fever in the Nigerian cohort but not in the Sierra Leone cohort. Overall, we identified variants and genes that may impact the risk of severe Lassa fever, demonstrating how GWAS can provide insight into viral pathogenesis.
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Fiebre de Lassa , Humanos , Fiebre de Lassa/genética , Fiebre de Lassa/diagnóstico , Fiebre de Lassa/epidemiología , Estudio de Asociación del Genoma Completo , Estudios Seroepidemiológicos , Virus Lassa/genética , Fiebre , Genética HumanaRESUMEN
Effective infectious disease surveillance in high-risk regions is critical for clinical care and pandemic preemption; however, few clinical diagnostics are available for the wide range of potential human pathogens. Here, we conduct unbiased metagenomic sequencing of 593 samples from febrile Nigerian patients collected in three settings: i) population-level surveillance of individuals presenting with symptoms consistent with Lassa Fever (LF); ii) real-time investigations of outbreaks with suspected infectious etiologies; and iii) undiagnosed clinically challenging cases. We identify 13 distinct viruses, including the second and third documented cases of human blood-associated dicistrovirus, and a highly divergent, unclassified dicistrovirus that we name human blood-associated dicistrovirus 2. We show that pegivirus C is a common co-infection in individuals with LF and is associated with lower Lassa viral loads and favorable outcomes. We help uncover the causes of three outbreaks as yellow fever virus, monkeypox virus, and a noninfectious cause, the latter ultimately determined to be pesticide poisoning. We demonstrate that a local, Nigerian-driven metagenomics response to complex public health scenarios generates accurate, real-time differential diagnoses, yielding insights that inform policy.
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Fiebre de Lassa , Virus , Humanos , Nigeria/epidemiología , Metagenómica , Fiebre de Lassa/diagnóstico , Fiebre de Lassa/epidemiología , Virus Lassa/genética , Virus/genéticaRESUMEN
OBJECTIVES: To assess the prevalence of acute kidney injury (AKI), and its impact on outcome in hospitalized pediatric patients with Lassa fever (LF). METHODS: We reviewed the presenting clinical and laboratory features and outcomes of 40 successive hospitalized children with PCR-confirmed LF. The diagnosis and staging of AKI was based on KDIGO criteria. We compared groups of patients using t- or χ2 tests as necessary, and took p-values <0.05 as indicative of the presence of significant differences. RESULTS: Sixteen (40%) children had AKI. Case fatality rate (CFR) was 9/16 (56%) in children with and 1/24 (4%) in those without AKI (OR [95% CI] of CFR associated with AKI = 29.57 [3.17, 275.7]). Presentation with abnormal bleeding (p = 0.008), encephalopathy (p = 0.004), hematuria plus proteinuria (p = 0.013), and elevated serum transaminase levels (p <0.02) were significantly associated with an increased prevalence of AKI. CONCLUSION: AKI prevalence in hospitalized pediatric patients with Lassa fever is high, and correlated with illness severity/CFR. The high prevalence underscores the need for access to hemodialysis, and clinical presentation and/or presence of hematuria plus proteinuria could serve as a ready prompt for referral for such specialized care.
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Lesión Renal Aguda/epidemiología , Fiebre de Lassa/complicaciones , Fiebre de Lassa/mortalidad , Diálisis Renal , Lesión Renal Aguda/diagnóstico , Lesión Renal Aguda/terapia , Preescolar , Femenino , Accesibilidad a los Servicios de Salud , Hematuria/complicaciones , Humanos , Lactante , Recién Nacido , Masculino , Nigeria/epidemiología , Prevalencia , Proteinuria/complicaciones , Índice de Severidad de la EnfermedadRESUMEN
Fifty patients with unexplained fever and poor outcomes presented at Irrua Specialist Teaching Hospital (ISTH) in Edo State, Nigeria, an area endemic for Lassa fever, between September 2018 - January 2019. After ruling out Lassa fever, plasma samples from these epidemiologically-linked cases were sent to the African Centre of Excellence for Genomics of Infectious Diseases (ACEGID), Redeemer's University, Ede, Osun State, Nigeria, where we carried out metagenomic sequencing which implicated yellow fever virus (YFV) as the etiology of this outbreak. Twenty-nine of the 50 samples were confirmed positive for YFV by reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR), 14 of which resulted in genome assembly. Maximum likelihood phylogenetic analysis revealed that these YFV sequences formed a tightly clustered clade more closely related to sequences from Senegal than sequences from earlier Nigerian isolates, suggesting that the YFV clade responsible for this outbreak in Edo State does not descend directly from the Nigerian YFV outbreaks of the last century, but instead reflects a broader diversity and dynamics of YFV in West Africa. Here we demonstrate the power of metagenomic sequencing for identifying ongoing outbreaks and their etiologies and informing real-time public health responses, resulting in accurate and prompt disease management and control.
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Sistemas de Computación , Brotes de Enfermedades , Metagenoma , Enfermedades no Diagnosticadas/epidemiología , Enfermedades no Diagnosticadas/genética , Fiebre Amarilla/epidemiología , Fiebre Amarilla/genética , Adolescente , Adulto , Niño , Preescolar , Femenino , Humanos , Funciones de Verosimilitud , Masculino , Persona de Mediana Edad , Nigeria/epidemiología , Filogenia , Enfermedades no Diagnosticadas/virología , Fiebre Amarilla/virología , Adulto JovenRESUMEN
Lassa virus (LASV) is the causative agent of Lassa fever (LF), an often-fatal hemorrhagic disease. LF is endemic in Nigeria, Sierra Leone and other West African countries. Diagnosis of LASV infection is challenged by the genetic diversity of the virus, which is greatest in Nigeria. The ReLASV Pan-Lassa Antigen Rapid Test (Pan-Lassa RDT) is a point-of-care, in vitro diagnostic test that utilizes a mixture of polyclonal antibodies raised against recombinant nucleoproteins of representative strains from the three most prevalent LASV lineages (II, III and IV). We compared the performance of the Pan-LASV RDT to available quantitative PCR (qPCR) assays during the 2018 LF outbreak in Nigeria. For patients with acute LF (RDT positive, IgG/IgM negative) during initial screening, RDT performance was 83.3% sensitivity and 92.8% specificity when compared to composite results of two qPCR assays. 100% of samples that gave Ct values below 22 on both qPCR assays were positive on the Pan-Lassa RDT. There were significantly elevated case fatality rates and elevated liver transaminase levels in subjects whose samples were RDT positive compared to RDT negative.
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Anticuerpos Antivirales/metabolismo , Pruebas Diagnósticas de Rutina/métodos , Fiebre de Lassa/diagnóstico , Virus Lassa/aislamiento & purificación , ARN Viral/genética , Adulto , Antígenos Virales/inmunología , Brotes de Enfermedades , Femenino , Humanos , Virus Lassa/genética , Virus Lassa/inmunología , Masculino , Persona de Mediana Edad , Nigeria , Sistemas de Atención de Punto , Sensibilidad y Especificidad , Análisis de Secuencia de ARN , Adulto JovenRESUMEN
Recent outbreaks of viral hemorrhagic fevers (VHFs), including Ebola virus disease (EVD) and Lassa fever (LF), highlight the urgent need for sensitive, deployable tests to diagnose these devastating human diseases. Here we develop CRISPR-Cas13a-based (SHERLOCK) diagnostics targeting Ebola virus (EBOV) and Lassa virus (LASV), with both fluorescent and lateral flow readouts. We demonstrate on laboratory and clinical samples the sensitivity of these assays and the capacity of the SHERLOCK platform to handle virus-specific diagnostic challenges. We perform safety testing to demonstrate the efficacy of our HUDSON protocol in heat-inactivating VHF viruses before SHERLOCK testing, eliminating the need for an extraction. We develop a user-friendly protocol and mobile application (HandLens) to report results, facilitating SHERLOCK's use in endemic regions. Finally, we successfully deploy our tests in Sierra Leone and Nigeria in response to recent outbreaks.
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Ebolavirus/patogenicidad , Fiebre Hemorrágica Ebola/diagnóstico , Fiebre de Lassa/diagnóstico , Virus Lassa/patogenicidad , Anticuerpos Antivirales , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Ebolavirus/genética , Fiebre Hemorrágica Ebola/virología , Fiebre de Lassa/virología , Virus Lassa/genéticaRESUMEN
Background: The general lack of comprehensive data on the trends of Lassa fever (LF) outbreaks contrasts with its widespread occurrence in West Africa and is an important constraint in the design of effective control measures. We reviewed the contribution of LF to admissions and mortality among hospitalized patients from 2001 to 2018 in the bid to address this gap. Methods: Observational study of LF caseload and mortality from 2001 to 18 in terms of the contribution of confirmed LF to admissions and deaths, and case fatality (CF) among patients with confirmed LF at a specialist center in Nigeria. The diagnosis of LF was confirmed using reverse transcription polymerase chain reaction (RT-PCR) test, and medians and frequencies were compared using Kruskal-Wallis, Mann-Whitney and χ2 tests, with p-values <0.05 taken as significant. Results: The contribution of confirmed LF to deaths (362/9057, 4.0%) was significantly higher than to admissions (1,298/185,707, 0.7%; OR [95% CI] = 5.9 [5.3, 6.7], p < 0.001). The average CF among patients with confirmed LF declined from 154/355 (43%) in 2001-09 to 183/867 (21.1%) (OR [95% CI] = 2.9 [2.2, 3.7], p < 0.001) in 2011-18. The annual CF declined from 94% in 2001 to 15% in 2018 whereas the caseload increased from 0.3 to 3.4%. The outbreaks were characterized by irregular cycles of high caseload in 2005-2007, 2012-2014, and 2016-2018, and progressive blurring of the seasonality. Conclusion: LF outbreaks in Nigeria have upgraded spatially and temporally, with the potential for cycles of increasing severity. The strategic establishment of LF surveillance and clinical case management centers could be a pragmatic and cost-effective approach to mitigating the outbreaks, particularly in reducing the associated CF. Urgent efforts are needed in reinvigorating extant control measures while the search for sustainable solutions continues.
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[This corrects the article DOI: 10.3389/fpubh.2019.00170.].
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Metagenomic sequencing has the potential to transform microbial detection and characterization, but new tools are needed to improve its sensitivity. Here we present CATCH, a computational method to enhance nucleic acid capture for enrichment of diverse microbial taxa. CATCH designs optimal probe sets, with a specified number of oligonucleotides, that achieve full coverage of, and scale well with, known sequence diversity. We focus on applying CATCH to capture viral genomes in complex metagenomic samples. We design, synthesize, and validate multiple probe sets, including one that targets the whole genomes of the 356 viral species known to infect humans. Capture with these probe sets enriches unique viral content on average 18-fold, allowing us to assemble genomes that could not be recovered without enrichment, and accurately preserves within-sample diversity. We also use these probe sets to recover genomes from the 2018 Lassa fever outbreak in Nigeria and to improve detection of uncharacterized viral infections in human and mosquito samples. The results demonstrate that CATCH enables more sensitive and cost-effective metagenomic sequencing.
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Biología Computacional/métodos , Genoma Viral , Metagenoma , Metagenómica , Animales , Culicidae/virología , Brotes de Enfermedades , Biblioteca de Genes , Variación Genética , Genómica , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Fiebre de Lassa/virología , Nigeria/epidemiología , Sondas de Oligonucleótidos , Oligonucleótidos/genética , Análisis de Secuencia de ADN , VirosisRESUMEN
It is rare both to have the central nervous system (CNS) as the main focus in the acute phase of Lassa fever infection without associated bleeding, and to find Lassa virus (LAV) in the cerebrospinal fluid (CSF) but not in the serum. We report the case of a 38-year-old Nigerian woman with mainly CNS manifestation of Lassa fever. She was admitted twice within 11 days because of persistent fever. A clinical diagnosis of acute LAV encephalitis was made because of a high index of suspicion and CNS involvement confirmed by positive reverse transcriptase polymerase chain reaction (RT-PCR) for LAV in the CSF, while her blood was repeatedly negative for LAV by RT-PCR test. She recovered fully following supportive care coupled with treatment with an 18-day course of ribavirin, and suffered no long-term neurological complication or relapse. Post-treatment CSF examination by RT-PCR did not detect LAV.
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BACKGROUND: Lassa fever is a viral haemorrhagic disease endemic to west Africa. No large-scale studies exist from Nigeria, where the Lassa virus (LASV) is most diverse. LASV diversity, coupled with host genetic and environmental factors, might cause differences in disease pathophysiology. Small-scale studies in Nigeria suggest that acute kidney injury is an important clinical feature and might be a determinant of survival. We aimed to establish the demographic, clinical, and laboratory factors associated with mortality in Nigerian patients with Lassa fever, and hypothesised that LASV was the direct cause of intrinsic renal damage for a subset of the patients with Lassa fever. METHODS: We did a retrospective, observational cohort study of consecutive patients in Nigeria with Lassa fever, who tested positive for LASV with RT-PCR, and were treated in Irrua Specialist Teaching Hospital. We did univariate and multivariate statistical analyses, including logistic regression, of all demographic, clinical, and laboratory variables available at presentation to identify the factors associated with patient mortality. FINDINGS: Of 291 patients treated in Irrua Specialist Teaching Hospital between Jan 3, 2011, and Dec 11, 2015, 284 (98%) had known outcomes (died or survived) and seven (2%) were discharged against medical advice. Overall case-fatality rate was 24% (68 of 284 patients), with a 1·4 times increase in mortality risk for each 10 years of age (p=0·00017), reaching 39% (22 of 57) for patients older than 50 years. Of 284 patients, 81 (28%) had acute kidney injury and 104 (37%) had CNS manifestations and thus both were considered important complications of acute Lassa fever in Nigeria. Acute kidney injury was strongly associated with poor outcome (case-fatality rate of 60% [49 of 81 patients]; odds ratio [OR] 15, p<0·00001). Compared with patients without acute kidney injury, those with acute kidney injury had higher incidence of proteinuria (32 [82%] of 39 patients) and haematuria (29 [76%] of 38) and higher mean serum potassium (4·63 [SD 1·04] mmol/L) and lower blood urea nitrogen to creatinine ratio (8·6 for patients without clinical history of fluid loss), suggesting intrinsic renal damage. Normalisation of creatinine concentration was associated with recovery. Elevated serum creatinine (OR 1·3; p=0·046), aspartate aminotransferase (OR 1·5; p=0·075), and potassium (OR 3·6; p=0·0024) were independent predictors of death. INTERPRETATION: Our study presents detailed clinical and laboratory data for Nigerian patients with Lassa fever and provides strong evidence for intrinsic renal dysfunction in acute Lassa fever. Early recognition and treatment of acute kidney injury might significantly reduce mortality. FUNDING: German Research Foundation, German Center for Infection Research, Howard Hughes Medical Institute, US National Institutes of Health, and World Bank.
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Fiebre de Lassa/patología , Fiebre de Lassa/terapia , Adulto , Estudios de Cohortes , Femenino , Humanos , Modelos Logísticos , Masculino , Análisis Multivariante , Nigeria/epidemiología , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
BACKGROUND: The classical method for detection of Lassa virus-specific antibodies is the immunofluorescence assay (IFA) using virus-infected cells as antigen. However, IFA requires laboratories of biosafety level 4 for assay production and an experienced investigator to interpret the fluorescence signals. Therefore, we aimed to establish and evaluate enzyme-linked immunosorbent assays (ELISA) using recombinant Lassa virus nucleoprotein (NP) as antigen. METHODOLOGY/PRINCIPAL FINDINGS: The IgM ELISA is based on capturing IgM antibodies using anti-IgM, and the IgG ELISA is based on capturing IgG antibody-antigen complexes using rheumatoid factor or Fc gamma receptor CD32a. Analytical and clinical evaluation was performed with 880 sera from Lassa fever endemic (Nigeria) and non-endemic (Ghana and Germany) areas. Using the IFA as reference method, we observed 91.5-94.3% analytical accuracy of the ELISAs in detecting Lassa virus-specific antibodies. Evaluation of the ELISAs for diagnosis of Lassa fever on admission to hospital in an endemic area revealed a clinical sensitivity for the stand-alone IgM ELISA of 31% (95% CI 25-37) and for combined IgM/IgG detection of 26% (95% CI 21-32) compared to RT-PCR. The specificity of IgM and IgG ELISA was estimated at 96% (95% CI 93-98) and 100% (95% CI 99-100), respectively, in non-Lassa fever patients from non-endemic areas. In patients who seroconverted during follow-up, Lassa virus-specific IgM and IgG developed simultaneously rather than sequentially. Consistent with this finding, isolated IgM reactivity, i.e. IgM in the absence of IgG, had no diagnostic value. CONCLUSIONS/SIGNIFICANCE: The ELISAs are not equivalent to RT-PCR for early diagnosis of Lassa fever; however, they are of value in diagnosing patients at later stage. The IgG ELISA may be useful for epidemiological studies and clinical trials due its high specificity, and the higher throughput rate and easier operation compared to IFA.
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Ensayo de Inmunoadsorción Enzimática/métodos , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Fiebre de Lassa/diagnóstico , Virus Lassa/inmunología , Nucleoproteínas/inmunología , Anticuerpos Antivirales/sangre , Técnica del Anticuerpo Fluorescente Indirecta , Alemania/epidemiología , Ghana/epidemiología , Humanos , Fiebre de Lassa/epidemiología , Fiebre de Lassa/inmunología , Virus Lassa/aislamiento & purificación , Nigeria/epidemiología , Nucleoproteínas/genética , ARN Viral/sangre , Sensibilidad y EspecificidadRESUMEN
OBJECTIVES: To determine the prevalence of malaria and anemia among pregnant women attending a traditional birth center as well as the effect of herbal remedies, gravidity, age, educational background and malaria prevention methods on their prevalence. METHODS: Blood specimens were collected from 119 pregnant women attending a Traditional Birth Home in Benin City, Nigeria. Malaria parasitemia was diagnosed by microscopy while anemia was defined as hemoglobin concentration <11 g/dL. RESULTS: The prevalence of malaria infection was (OR=4.35 95% CI=1.213, 15.600; p=0.016) higher among primigravidae (92.1%). Pregnant women (38.5%) with tertiary level of education had significantly lower prevalence of malaria infection (p=0.002). Malaria significantly affected the prevalence of anemia (p<0.05). Anemia was associated with consumption of herbal remedies (OR=2.973; 95% CI=1.206, 7.330; p=0.017). The prevalence of malaria parasitemia and anemia were not affected by malaria prevention methods used by the participants. CONCLUSION: The overall prevalence of malaria infection and anemia observed in this study were 78.9% and 46.2%, respectively. Higher prevalence of malaria infection was associated with primigravidae and lower prevalence with tertiary education of subjects. Anemia was associated with consumption of herbal remedies. There is urgent need to control the prevalence of malaria and anemia among pregnant women attending traditional birth homes.