Shedding of soluble glycoprotein 1 detected during acute Lassa virus infection in human subjects

<p>Abstract</p> <p>Background</p> <p>Lassa hemorrhagic fever (LHF) is a neglected tropical disease with significant impact on the health care system, society, and economy of Western and Central African nations where it is endemic. With a high rate of infection that may lead to morbidity and mortality, understanding how the virus interacts with the host's immune system is of great importance for generating vaccines and therapeutics. Previous work by our group identified a soluble isoform of the Lassa virus (LASV) GP1 (sGP1) <it>in vitro </it>resulting from the expression of the glycoprotein complex (GPC) gene <abbrgrp><abbr bid="B1">1</abbr><abbr bid="B2">2</abbr></abbrgrp>. Though no work has directly been done to demonstrate the function of this soluble isoform in arenaviral infections, evidence points to immunomodulatory effects against the host's immune system mediated by a secreted glycoprotein component in filoviruses, another class of hemorrhagic fever-causing viruses. A significant fraction of shed glycoprotein isoforms during viral infection and biogenesis may attenuate the host's inflammatory response, thereby enhancing viral replication and tissue damage. Such shed glycoprotein mediated effects were previously reported for Ebola virus (EBOV), a filovirus that also causes hemorrhagic fever with nearly 90% fatality rates <abbrgrp><abbr bid="B3">3</abbr><abbr bid="B4">4</abbr><abbr bid="B5">5</abbr></abbrgrp>. The identification of an analogous phenomenon <it>in vivo </it>could establish a new correlate of LHF infection leading to the development of sensitive diagnostics targeting the earliest molecular events of the disease. Additionally, the reversal of potentially untoward immunomodulatory functions mediated by sGP1 could potentiate the development of novel therapeutic intervention. To this end, we investigated the presence of sGP1 in the serum of suspected LASV patients admitted to the Kenema Government Hospital (KGH) Lassa Fever Ward (LFW), in Kenema, Sierra Leone that tested positive for viral antigen or displayed classical signs of Lassa fever.</p> <p>Results</p> <p>It is reasonable to expect that a narrow window exists for detection of sGP1 as the sole protein shed during early arenaviral biogenesis. This phenomenon was clearly distinguishable from virion-associated GP1 only prior to the emergence of <it>de novo </it>viral particles. Despite this restricted time frame, in 2/46 suspected cases in two studies performed in late 2009 and early 2010, soluble glycoprotein component shedding was identified. Differential detection of viral antigens GP1, GP2, and NP by western blot yielded five different scenarios: whole LASV virions (GP1, GP2, NP; i.e. active viremia), different combinations of these three proteins, sGP1 only, NP only, and absence of all three proteins. Four additional samples showed inconclusive evidence for sGP1 shedding due to lack of detection of GP2 and NP by western blot; however, a sensitive LASV NP antigen capture ELISA generated marginally positive signals</p> <p>Conclusions</p> <p>During a narrow window following active infection with LASV, soluble GP1 can be detected in patient sera. This phenomenon parallels other VHF infection profiles, with the actual role of a soluble viral glycoprotein component <it>in vivo </it>remaining largely speculative. The expenditure of energy and cellular resources toward secretion of a critical protein during viral biogenesis without apparent specific function requires further investigation. Future studies will be aimed at systematically identifying the role of LASV sGP1 in the infection process and outcome <it>in vitro </it>and <it>in vivo</it>.</p

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

Emerging trends in Lassa fever: redefining the role of immunoglobulin M and inflammation in diagnosing acute infection

Abstract Background Lassa fever (LF) is a devastating hemorrhagic viral disease that is endemic to West Africa and responsible for thousands of human deaths each year. Analysis of humoral immune responses (IgM and IgG) by antibody-capture ELISA (Ab-capture ELISA) and Lassa virus (LASV) viremia by antigen-capture ELISA (Ag-capture ELISA) in suspected patients admitted to the Kenema Government Hospital (KGH) Lassa Fever Ward (LFW) in Sierra Leone over the past five years is reshaping our understanding of acute LF. Results Analyses in LF survivors indicated that LASV-specific IgM persists for months to years after initial infection. Furthermore, exposure to LASV appeared to be more prevalent in historically non-endemic areas of West Africa with significant percentages of reportedly healthy donors IgM and IgG positive in LASV-specific Ab-capture ELISA. We found that LF patients who were Ag positive were more likely to die than suspected cases who were only IgM positive. Analysis of metabolic and immunological parameters in Ag positive LF patients revealed a strong correlation between survival and low levels of IL-6, -8, -10, CD40L, BUN, ALP, ALT, and AST. Despite presenting to the hospital with fever and in some instances other symptoms consistent with LF, the profiles of Ag negative IgM positive individuals were similar to those of normal donors and nonfatal (NF) LF cases, suggesting that IgM status cannot necessarily be considered a diagnostic marker of acute LF in suspected cases living in endemic areas of West Africa. Conclusion Only LASV viremia assessed by Ag-capture immunoassay, nucleic acid detection or virus isolation should be used to diagnose acute LASV infection in West Africans. LASV-specific IgM serostatus cannot be considered a diagnostic marker of acute LF in suspected cases living in endemic areas of West Africa. By applying these criteria, we identified a dysregulated metabolic and pro-inflammatory response profile conferring a poor prognosis in acute LF. In addition to suggesting that the current diagnostic paradigm for acute LF should be reconsidered, these studies present new opportunities for therapeutic interventions based on potential prognostic markers in LF.</p

Clinical Illness and Outcomes in Patients with Ebola in Sierra Leone

Background;Limited clinical and laboratory data are available on patients with Ebola virus disease (EVD). The Kenema Government Hospital in Sierra Leone, which had an existing infrastructure for research regarding viral hemorrhagic fever, has received and cared for patients with EVD since the beginning of the outbreak in Sierra Leone in May 2014. Methods;We reviewed available epidemiologic, clinical, and laboratory records of patients in whom EVD was diagnosed between May 25 and June 18, 2014. We used quantitative reverse-transcriptase–polymerase-chain-reaction assays to assess the load of Ebola virus (EBOV, Zaire species) in a subgroup of patients. Results;Of 106 patients in whom EVD was diagnosed, 87 had a known outcome, and 44 had detailed clinical information available. The incubation period was estimated to be 6 to 12 days, and the case fatality rate was 74%. Common findings at presentation included fever (in 89% of the patients), headache (in 80%), weakness (in 66%), dizziness(in 60%), diarrhea (in 51%), abdominal pain (in 40%), and vomiting (in 34%). Clinical and laboratory factors at presentation that were associated with a fatal outcome included fever, weakness, dizziness, diarrhea, and elevated levels of blood urea nitrogen, aspartate aminotransferase, and creatinine. Exploratory analyses indicated that patients under the age of 21 years had a lower case fatality rate than those over the age of 45 years (57% vs. 94%, P=0.03), and patients presenting with fewer than 100,000 EBOV copies per milliliter had a lower case fatality rate than those with 10 million EBOV copies per milliliter or more (33% vs. 94%, P=0.003). Bleeding occurred in only 1 patient.Conclusions; The incubation period and case fatality rate among patients with EVD in Sierra Leone are similar to those observed elsewhere in the 2014 outbreak and in previous outbreaks. Although bleeding was an infrequent finding, diarrhea and other gastrointestinal manifestations were common. (Funded by the National Institutes of Health and others.)Organismic and Evolutionary Biolog

CFRs in suspected LF cases presenting to the KGH Lassa Ward by serostatus, 2008–12.

<p>Panel A: CFR by serostatus. The presence of LASV Ag and anti-LASV IgM in serum of patients with verifiable outcomes was assessed by recombinant Ag− and IgM− capture ELISA, respectively. Panel B: Alternative calculation of CFRs. Ag+/IgM± plus Ag−/IgM+ compared to Ag−/IgM−. Statistical significance was determined using a logistic regression model predicting CFR (<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002748#pntd.0002748.s004" target="_blank">Table S3</a>). NS = not significant.</p