Field Validation of the ReEBOV Antigen Rapid Test for Point-of-Care Diagnosis of Ebola Virus Infection.

BACKGROUND: The 2013-2016 West African Ebola virus disease (EVD) epidemic is the largest recorded. Triage on the basis of clinical signs had limited success, and the time to diagnosis by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) could exceed 5 days. Here we describe the development and field validation of the ReEBOV Antigen Rapid Test (ReEBOV RDT) to aid triage of individuals with suspected EVD. METHODS: Samples from patients with suspected EVD were submitted to Kenema Government Hospital, Sierra Leone, for Lassa fever and EVD screening throughout 2014. Banked residual clinical samples were tested in November 2014 and January 2015 in a blinded field trial to estimate the clinical effectiveness of the ReEBOV RDT, compared with EBOV-specific qRT-PCR. RESULTS: Preliminary ReEBOV RDT performance demonstrated a positive percentage agreement (PPA) of 91.1% (195 of 214 results; 95% confidence interval [CI], 86.5%-94.6%) and a negative percentage agreement (NPA) of 90.2% (175 of 194; 95% CI, 85.1%-94.0%). The final estimates used by the Food and Drug Administration to determine whether to grant emergency use authorization for the test, which excluded a qRT-PCR reference method threshold cutoff, were a PPA of 62.1% (72 of 116 results; 95% CI, 52.6%-70.9%) and a NPA of 96.7% (58 of 60; 95% CI, 88.5%-99.6%), with a diagnostic likelihood of 18.6. A subsequent, independent evaluation by the World Health Organization generated results consistent with the preliminary performance estimates. CONCLUSIONS: The ReEBOV RDT demonstrated the potential to provide clinically effective rapid and accurate point-of-care test results and, thus, to be a powerful tool for increasing triage efficiency.

An Outbreak of Ebola Virus Disease in the Lassa Fever Zone.

BACKGROUND: Kenema Government Hospital (KGH) has developed an advanced clinical and laboratory research capacity to manage the threat of Lassa fever, a viral hemorrhagic fever (VHF). The 2013-2016 Ebola virus (EBOV) disease (EVD) outbreak is the first to have occurred in an area close to a facility with established clinical and laboratory capacity for study of VHFs. METHODS: Because of its proximity to the epicenter of the EVD outbreak, which began in Guinea in March 2014, the KGH Lassa fever Team mobilized to establish EBOV surveillance and diagnostic capabilities. RESULTS: Augustine Goba, director of the KGH Lassa laboratory, diagnosed the first documented case of EVD in Sierra Leone, on 25 May 2014. Thereafter, KGH received and cared for numbers of patients with EVD that quickly overwhelmed the capacity for safe management. Numerous healthcare workers contracted and lost their lives to EVD. The vast majority of subsequent EVD cases in West Africa can be traced back to a single transmission chain that includes this first diagnosed case. CONCLUSIONS: Responding to the challenges of confronting 2 hemorrhagic fever viruses will require continued investments in the development of countermeasures (vaccines, therapeutic agents, and diagnostic assays), infrastructure, and human resources.

Lassa Fever among Children in Eastern Province, Sierra Leone: A 7-year Retrospective Analysis (2012-2018).

Pediatric Lassa fever (LF) usually presents as a nonspecific febrile illness, similar to other endemic diseases in countries like Sierra Leone, where LF is considered to be hyperendemic. The nonspecificity of presentation and lack of research have made it difficult to fully understand best practices for pediatric management. We aim to describe clinical characteristics of hospitalized pediatric patients suspected or diagnosed with LF and assess factors associated with hospital outcomes among those with LF antigen-positive results. We conducted a 7-year retrospective cohort study using routine data for all children younger than 18 years admitted at the Kenema Government Hospital's LF ward. A total of 292 children with suspected or confirmed LF were analyzed. Overall, mortality was high (21%). Children with antigen-positive results had a high case fatality rate of 63% (P < 0.01). In univariate analyses, children who presented with unexplained bleeding (odds ratio [OR]: 3.58; 95% CI: 1.08-11.86; P = 0.040) and confusion (altered sensorium) (OR: 5.37; 95% CI: 1.34-21.48; P = 0.020) had increased odds of death. Abnormal serum levels of alanine aminotransferase (P = 0.001), creatinine (P = 0.004), and potassium (P = 0.003) were associated with increased likelihood of death in these children. Treatment with ribavirin was not significantly associated with survival (P = 0.916). Our findings provide insights into current pediatric LF clinical presentation and management. More evidence-based, high-quality research in creating predictive algorithms of antigen-positivity and hospital outcomes is needed in the management of pediatric LF.

Field validation of recombinant antigen immunoassays for diagnosis of Lassa fever.

Lassa fever, a hemorrhagic fever caused by Lassa virus (LASV), is endemic in West Africa. It is difficult to distinguish febrile illnesses that are common in West Africa from Lassa fever based solely on a patient's clinical presentation. The field performance of recombinant antigen-based Lassa fever immunoassays was compared to that of quantitative polymerase chain assays (qPCRs) using samples from subjects meeting the case definition of Lassa fever presenting to Kenema Government Hospital in Sierra Leone. The recombinant Lassa virus (ReLASV) enzyme-linked immunosorbant assay (ELISA) for detection of viral antigen in blood performed with 95% sensitivity and 97% specificity using a diagnostic standard that combined results of the immunoassays and qPCR. The ReLASV rapid diagnostic test (RDT), a lateral flow immunoassay based on paired monoclonal antibodies to the Josiah strain of LASV (lineage IV), performed with 90% sensitivity and 100% specificity. ReLASV immunoassays performed better than the most robust qPCR currently available, which had 82% sensitivity and 95% specificity. The performance characteristics of recombinant antigen-based Lassa virus immunoassays indicate that they can aid in the diagnosis of LASV Infection and inform the clinical management of Lassa fever patients.

Most neutralizing human monoclonal antibodies target novel epitopes requiring both Lassa virus glycoprotein subunits.

Lassa fever is a severe multisystem disease that often has haemorrhagic manifestations. The epitopes of the Lassa virus (LASV) surface glycoproteins recognized by naturally infected human hosts have not been identified or characterized. Here we have cloned 113 human monoclonal antibodies (mAbs) specific for LASV glycoproteins from memory B cells of Lassa fever survivors from West Africa. One-half bind the GP2 fusion subunit, one-fourth recognize the GP1 receptor-binding subunit and the remaining fourth are specific for the assembled glycoprotein complex, requiring both GP1 and GP2 subunits for recognition. Notably, of the 16 mAbs that neutralize LASV, 13 require the assembled glycoprotein complex for binding, while the remaining 3 require GP1 only. Compared with non-neutralizing mAbs, neutralizing mAbs have higher binding affinities and greater divergence from germline progenitors. Some mAbs potently neutralize all four LASV lineages. These insights from LASV human mAb characterization will guide strategies for immunotherapeutic development and vaccine design.

Lassa fever in post-conflict sierra leone.

BACKGROUND: Lassa fever (LF), an often-fatal hemorrhagic disease caused by Lassa virus (LASV), is a major public health threat in West Africa. When the violent civil conflict in Sierra Leone (1991 to 2002) ended, an international consortium assisted in restoration of the LF program at Kenema Government Hospital (KGH) in an area with the world's highest incidence of the disease. METHODOLOGY/PRINCIPAL FINDINGS: Clinical and laboratory records of patients presenting to the KGH Lassa Ward in the post-conflict period were organized electronically. Recombinant antigen-based LF immunoassays were used to assess LASV antigenemia and LASV-specific antibodies in patients who met criteria for suspected LF. KGH has been reestablished as a center for LF treatment and research, with over 500 suspected cases now presenting yearly. Higher case fatality rates (CFRs) in LF patients were observed compared to studies conducted prior to the civil conflict. Different criteria for defining LF stages and differences in sensitivity of assays likely account for these differences. The highest incidence of LF in Sierra Leone was observed during the dry season. LF cases were observed in ten of Sierra Leone's thirteen districts, with numerous cases from outside the traditional endemic zone. Deaths in patients presenting with LASV antigenemia were skewed towards individuals less than 29 years of age. Women self-reporting as pregnant were significantly overrepresented among LASV antigenemic patients. The CFR of ribavirin-treated patients presenting early in acute infection was lower than in untreated subjects. CONCLUSIONS/SIGNIFICANCE: Lassa fever remains a major public health threat in Sierra Leone. Outreach activities should expand because LF may be more widespread in Sierra Leone than previously recognized. Enhanced case finding to ensure rapid diagnosis and treatment is imperative to reduce mortality. Even with ribavirin treatment, there was a high rate of fatalities underscoring the need to develop more effective and/or supplemental treatments for LF.