George O. Akpede

Clinical Sequencing Uncovers Origins and Evolution of Lassa Virus

The 2013-2015 West African epidemic of Ebola virus disease (EVD) reminds us of how little is known about biosafety level 4 viruses. Like Ebola virus, Lassa virus (LASV) can cause hemorrhagic fever with high case fatality rates. We generated a genomic catalog of almost 200 LASV sequences from clinical and rodent reservoir samples. We show that whereas the 2013-2015 EVD epidemic is fueled by human-to-human transmissions, LASV infections mainly result from reservoir-to-human infections. We elucidated the spread of LASV across West Africa and show that this migration was accompanied by changes in LASV genome abundance, fatality rates, codon adaptation, and translational efficiency. By investigating intrahost evolution, we found that mutations accumulate in epitopes of viral surface proteins, suggesting selection for immune escape. This catalog will serve as a foundation for the development of vaccines and diagnostics. VIDEO ABSTRACT.

Molecular Diagnostics for Lassa Fever at Irrua Specialist Teaching Hospital, Nigeria: Lessons Learnt from Two Years of Laboratory Operation

Background Lassa fever is a viral hemorrhagic fever endemic in West Africa. However, none of the hospitals in the endemic areas of Nigeria has the capacity to perform Lassa virus diagnostics. Case identification and management solely relies on non-specific clinical criteria. The Irrua Specialist Teaching Hospital (ISTH) in the central senatorial district of Edo State struggled with this challenge for many years. Methodology/Principal Findings A laboratory for molecular diagnosis of Lassa fever, complying with basic standards of diagnostic PCR facilities, was established at ISTH in 2008. During 2009 through 2010, samples of 1,650 suspected cases were processed, of which 198 (12%) tested positive by Lassa virus RT-PCR. No remarkable demographic differences were observed between PCR-positive and negative patients. The case fatality rate for Lassa fever was 31%. Nearly two thirds of confirmed cases attended the emergency departments of ISTH. The time window for therapeutic intervention was extremely short, as 50% of the fatal cases died within 2 days of hospitalization—often before ribavirin treatment could be commenced. Fatal Lassa fever cases were older (p = 0.005), had lower body temperature (p<0.0001), and had higher creatinine (p<0.0001) and blood urea levels (p<0.0001) than survivors. Lassa fever incidence in the hospital followed a seasonal pattern with a peak between November and March. Lassa virus sequences obtained from the patients originating from Edo State formed—within lineage II—a separate clade that could be further subdivided into three clusters. Conclusions/Significance Lassa fever case management was improved at a tertiary health institution in Nigeria through establishment of a laboratory for routine diagnostics of Lassa virus. Data collected in two years of operation demonstrate that Lassa fever is a serious public health problem in Edo State and reveal new insights into the disease in hospitalized patients.

Management of Children With Prolonged Fever of Unknown Origin and Difficulties in the Management of Fever of Unknown Origin in Children in Developing Countries

This is Part II of a 2-part paper on fever of unknown origin (FUO) in children.It examines the aetiology and management of prolonged FUO in children and the difficulties in the management of FUO in children in developing countries. Part I of this paper discussed acute FUO in children and was published in the March 2001 issue of Paediatric Drugs.Prolonged FUO is documented fever of more than 7 to 10 days which has no apparent source and no apparent diagnosis after 1 week of clinical investigations. About 34% of cases of prolonged FUO are caused by infections, with bacterial meningitis and urinary tract infection accounting for about 6.5 and 11.4%, respectively, of cases attributable to infections. Chronic infections, particularly tuberculosis and ‘old’ disorders such as Kawasaki disease, cat-scratch disease and Epstein-Barr virus infection presenting with ‘new’ manifestations, collagen-vascular diseases and neoplastic disorders are the other issues of major concern in prolonged FUO. Overall, however, there is a trend towards an increased number of undiagnosed cases. This is due to advancements in diagnostic techniques, such that illnesses which were previously common among the causes of prolonged FUO are now diagnosed earlier, before the presentation becomes that of prolonged FUO.Clinical examination supplemented with laboratory tests to screen for serious bacterial infections should be the mainstay of initial evaluation of children with prolonged FUO. Use of scanning techniques (such as computerised tomography and ultrasound) as additional supplements to this clinical examination may allow for the earlier diagnosis of causes of prolonged FUO in children such as ‘occult’ abdominal tumours. A common error in management of children with prolonged FUO is the failure to perform a complete history and physical examination; repeated clinical examination and continued observation are of paramount importance in the diagnosis of difficult cases.Major difficulties in the management of FUO in children in developing countries include constraints in the availability and reliability of laboratory tests, cost, misuse of antibiotics and difficulties encountered in the diagnosis of malaria and typhoid fever. Malaria and typhoid fever are major aetiological considerations in both acute and prolonged FUO in children in developing countries. The newer quinolones may hold great promise for the treatment of serious bacterial infections, including meningitis, which are associated with prolonged FUO in developing countries.

Current Molecular Epidemiology of Lassa Virus in Nigeria

ABSTRACT Recent Lassa virus strains from Nigeria were completely or partially sequenced. Phylogenetic analysis revealed the predominance of lineage II and III strains, the existence of a previously undescribed (sub)lineage in Nigeria, and the directional spread of virus in the southern part of the country. The Bayesian analysis also provided estimates for divergence times within the Lassa virus clade.

Discovery of Novel Rhabdoviruses in the Blood of Healthy Individuals from West Africa

Next-generation sequencing (NGS) has the potential to transform the discovery of viruses causing unexplained acute febrile illness (UAFI) because it does not depend on culturing the pathogen or a priori knowledge of the pathogen’s nucleic acid sequence. More generally, it has the potential to elucidate the complete human virome, including viruses that cause no overt symptoms of disease, but may have unrecognized immunological or developmental consequences. We have used NGS to identify RNA viruses in the blood of 195 patients with UAFI and compared them with those found in 328 apparently healthy (i.e., no overt signs of illness) control individuals, all from communities in southeastern Nigeria. Among UAFI patients, we identified the presence of nucleic acids from several well-characterized pathogenic viruses, such as HIV-1, hepatitis, and Lassa virus. In our cohort of healthy individuals, however, we detected the nucleic acids of two novel rhabdoviruses. These viruses, which we call Ekpoma virus-1 (EKV-1) and Ekpoma virus-2 (EKV-2), are highly divergent, with little identity to each other or other known viruses. The most closely related rhabdoviruses are members of the genus Tibrovirus and Bas-Congo virus (BASV), which was recently identified in an individual with symptoms resembling hemorrhagic fever. Furthermore, by conducting a serosurvey of our study cohort, we find evidence for remarkably high exposure rates to the identified rhabdoviruses. The recent discoveries of novel rhabdoviruses by multiple research groups suggest that human infection with rhabdoviruses might be common. While the prevalence and clinical significance of these viruses are currently unknown, these viruses could have previously unrecognized impacts on human health; further research to understand the immunological and developmental impact of these viruses should be explored. More generally, the identification of similar novel viruses in individuals with and without overt symptoms of disease highlights the need for a broader understanding of the human virome as efforts for viral detection and discovery advance.

Sensorineural hearing loss in Lassa fever: two case reports

IntroductionLassa fever is an acute arena viral haemorrhagic fever with varied neurological sequelae. Sensorineural hearing loss is one of the rare complications which occur usually during the convalescent stage of the infection.Case presentationThe cases of two female patients aged 19 and 43 years old, respectively, with clinical features suggestive of Lassa fever and confirmed by immunoserological/Lassa-virus-specific reverse transcriptase polymerase chain reaction are presented. Both patients developed severe sensorineural hearing loss at acute phases of the infections.ConclusionSensorineural hearing loss from Lassa fever infections can occur in both acute and convalescent stages and is probably induced by an immune response.

Aetiology and Management of Children With Acute Fever of Unknown Origin

This is part I of a 2-part paper on fever of unknown origin (FUO) in children. FUO is best defined as fever without obvious source on initial clinical examination and then classified into acute (illness of ≤1 week’s duration) and prolonged (>7 to 10 days’ duration). Aetiologically, there is a marked overlap between acute and prolonged FUO, and infections are major players in both. Age, climate, local epidemiology and host factors are the major aetiological factors that should be considered in the choice of definitive tests. Depending on age, serious bacterial infections (including bacteraemia, meningitis and urinary tract infection) occur in 3 to 20% of cases of acute FUO. Prevention of mortality and sequelae from these infections, particularly bacteraemia and meningitis, is of particular concern in acute FUO. An individualised approach, based on clinical evaluation supplemented with screening and definitive laboratory tests to determine the need for empiric antibiotic therapy and hospitalisation, seems to be the best approach to acute FUO (although this may be less applicable to neonates and infants younger than 90 days, particularly those aged 0 to 7 days). The place of laboratory tests, empiric antibiotic therapy and hospitalisation are important issues that are likely to remain so for some time.

Lassa fever presenting as acute abdomen: a case series

Lassa fever, an endemic zoonotic viral infection in West Africa, presents with varied symptoms including fever, vomiting, retrosternal pain, abdominal pain, sore-throat, mucosal bleeding, seizures and coma. When fever and abdominal pain are the main presenting symptoms, and a diagnosis of acute abdomen is entertained, Lassa fever is rarely considered in the differential diagnosis, even in endemic areas. Rather the diagnosis of Lassa fever is suspected only after surgical intervention. Therefore, such patients often undergo unnecessary surgery with resultant delay in the commencement of ribavirin therapy. This increases morbidity and mortality and the risk of nosocomial transmission to hospital staff.We report 7 patients aged between 17 months and 40 years who had operative intervention for suspected appendicitis, perforated typhoid ileitis, intussuception and ruptured ectopic pregnancy after routine investigations. All seven were post-operatively confirmed as Lassa fever cases. Four patients died postoperatively, most before commencement of ribavirin, while the other three patients eventually recovered with appropriate antibiotic treatment including intravenous ribavirin.Surgeons working in West Africa should include Lassa fever in the differential diagnosis of acute abdomen, especially appendicitis. The presence of high grade fever, proteinuria and thrombocytopenia in patients with acute abdomen should heighten the suspicion of Lassa fever. Prolonged intra-operative bleeding should not only raise suspicion of the disease but also serve to initiate precautions to prevent nosocomial transmission.

Clinical and laboratory predictors of Lassa fever outcome in a dedicated treatment facility in Nigeria: a retrospective, observational cohort study

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.

Prevalence of Lassa Virus Disease (LVD) in Nigerian children with fever or fever and convulsions in an endemic area

Background Convulsions with fever in children are a common neurologic emergency in the tropics, and determining the contribution of endemic viral infections can be challenging. In particular, there is a dearth of data on the prevalence and clinical differentiation of Lassa virus disease (LVD) in febrile children in endemic areas of Nigeria, which has multiple lineages of the virus. The aim of this study was to determine the prevalence and presentation of LVD in febrile children with and without convulsions. Methodology/Principal findings This was a prospective study of consecutive febrile children aged ≥1 month– 15 years admitted to the Children’s Emergency Room of Irrua Specialist Teaching Hospital over a period of 1 year. Febrile children with convulsions (Cases) were compared with those without convulsions (Controls). LVD was defined by the presence of a positive Lassa virus RT-PCR test. Rates were compared between groups using χ2 or Fisher’s exact tests and p <0.05 taken as significant. 373 (40.9%) of 913 admissions had fever. Of these, 108/373 (29%) presented with convulsions. The overall prevalence of LVD was 13/373 (3.5%; 95% CI = 1.9%, 5.7%) in febrile admissions, 3/108 (2.8%) in Cases and 10/265 (3.8%) in Controls [(Odds Ratio (95% Confidence Interval) (OR (95% CI)) of LVD in Cases versus Controls = 0.73 (0.2, 2.7)]. Only vomiting (OR (95% CI) = 0.09 (0.01, 0.70)) and bleeding (OR (95% CI) = 39.56 (8.52, 183.7)) were significantly associated with an increased prevalence of LVD. Conclusions/Significance LVD is an important cause of fever, including undifferentiated fever in children in endemic areas, but it is not significantly associated with convulsions associated with fever. Its prevalence, and lack of clinical differentiation on presentation, underscores the importance of a high index of suspicion in diagnosis. Screening of febrile children with undifferentiated fever in endemic areas for LVD could be an important medical and public health control measure.