Sunday A. Omilabu

Low Genetic Diversity despite Hyperendemicity of Hepatitis B Virus Genotype E throughout West Africa

Sub-Saharan Africa suffers from an excessively high endemicity of hepatitis B virus (HBV), but little is known about the prevalent genotypes. In this study, we investigated the PreS1/PreS2/S genes of 127 viruses obtained from 12 locations in Mali, Burkina Faso, Togo, Benin, Nigeria, Cameroon, and the Democratic Republic of Congo. Except for those obtained from the Cameroon HIV cohort (18/22 HBV genotype A), 96 of 105 sequences belonged to HBV genotype E (HBV/E), and viral DNA was very similar (1.67% diversity) throughout this vast HBV/E crescent, which spans 6000 km across Africa. The low diversity suggests that HBV/E may have a short evolutionary history. Considering a typical mutation rate of DNA viruses, it would take only 200 years for the strain diversity of HBV/E viruses to develop from a single introductory event. The relatively recent introduction of HBV/E into humans would also explain its conspicuous absence in the Americas, despite the forced immigration of slaves from west Africa, until the early 19th century. Infection during infancy is mostly associated with chronic carrier status, and this combination can account for the explosive spread of virtually identical viruses within a community, but whether other routes of long-range transmissions must be considered becomes an important question.

Nomenclature- and database-compatible names for the two Ebola virus variants that emerged in Guinea and the Democratic Republic of the Congo in 2014.

In 2014, Ebola virus (EBOV) was identified as the etiological agent of a large and still expanding outbreak of Ebola virus disease (EVD) in West Africa and a much more confined EVD outbreak in Middle Africa. Epidemiological and evolutionary analyses confirmed that all cases of both outbreaks are connected to a single introduction each of EBOV into human populations and that both outbreaks are not directly connected. Coding-complete genomic sequence analyses of isolates revealed that the two outbreaks were caused by two novel EBOV variants, and initial clinical observations suggest that neither of them should be considered strains. Here we present consensus decisions on naming for both variants (West Africa: “Makona”, Middle Africa: “Lomela”) and provide database-compatible full, shortened, and abbreviated names that are in line with recently established filovirus sub-species nomenclatures.

Improved Detection of Lassa Virus by Reverse Transcription-PCR Targeting the 5′ Region of S RNA

ABSTRACT The method of choice for the detection of Lassa virus is reverse transcription (RT)-PCR. However, the high degree of genetic variability of the virus poses a problem with the design of RT-PCR assays that will reliably detect all strains. Recently, we encountered difficulties in detecting some strains from Liberia and Nigeria in a commonly used glycoprotein precursor (GPC) gene-specific RT-PCR assay (A. H. Demby, J. Chamberlain, D. W. Brown, and C. S. Clegg, J. Clin. Microbiol. 32:2898-2903, 1994), which prompted us to revise the protocol. The design of the new assay, the GPC RT-PCR/2007 assay, took into account 62 S RNA sequences from all countries where Lassa fever is endemic, including 40 sequences generated from the strains in our collection. The analytical sensitivity of the new assay was determined with 11 strains from Sierra Leone, Liberia, Ivory Coast, and Nigeria by probit analysis; the viral loads detectable with a probability of 95% ranged from 342 to 2,560 S RNA copies/ml serum, which corresponds to 4 to 30 S RNA copies/assay. The GPC RT-PCR/2007 assay was validated with 77 serum samples and 1 cerebrospinal fluid sample from patients with laboratory-confirmed Lassa fever. The samples mainly originated from Liberia and Nigeria and included strains difficult to detect in the assay of 1994. The GPC RT-PCR/2007 assay detected virus in all clinical specimens (100% sensitivity). In conclusion, a new RT-PCR assay, based in part on the protocol developed by Demby et al. in 1994, for the detection of Lassa virus is described. Compared to the assay developed in 1994, the GPC RT-PCR/2007 assay offers improved sensitivity for the detection of Liberian and Nigerian Lassa virus strains.

Filovirus RefSeq Entries: Evaluation and Selection of Filovirus Type Variants, Type Sequences, and Names

Sequence determination of complete or coding-complete genomes of viruses is becoming common practice for supporting the work of epidemiologists, ecologists, virologists, and taxonomists. Sequencing duration and costs are rapidly decreasing, sequencing hardware is under modification for use by non-experts, and software is constantly being improved to simplify sequence data management and analysis. Thus, analysis of virus disease outbreaks on the molecular level is now feasible, including characterization of the evolution of individual virus populations in single patients over time. The increasing accumulation of sequencing data creates a management problem for the curators of commonly used sequence databases and an entry retrieval problem for end users. Therefore, utilizing the data to their fullest potential will require setting nomenclature and annotation standards for virus isolates and associated genomic sequences. The National Center for Biotechnology Information’s (NCBI’s) RefSeq is a non-redundant, curated database for reference (or type) nucleotide sequence records that supplies source data to numerous other databases. Building on recently proposed templates for filovirus variant naming [ ()////-], we report consensus decisions from a majority of past and currently active filovirus experts on the eight filovirus type variants and isolates to be represented in RefSeq, their final designations, and their associated sequences.

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.

Lassa Fever, Nigeria, 2005–2008

To the Editor: Lassa fever affects ≈100,000 persons per year in West Africa (1). The disease is caused by Lassa virus, an arenavirus, and is associated with bleeding and organ failure. The case-fatality rate in hospitalized patients is 10%–20%. The reservoir of the virus is multimammate mice (Mastomys natalensis). Investigations in the 1970s and 1980s pointed to the existence of 3 disease-endemic zones within Nigeria: the northeastern region around Lassa, the central region around Jos, and the southern region around Onitsha (2,3). The current epidemiologic situation is less clear because no surveillance system is in place. In 2003 and 2004, we conducted a hospital-based survey in Irrua, which demonstrated ongoing transmission of the virus in Edo State, Nigeria (4). Since then, laboratory capacity at the University of Lagos for diagnosing Lassa fever has been improved and used for small-scale passive surveillance in other parts of the country. Public health officials or hospital staff reported suspected cases. Blood samples were sent to Lagos, or staff from Lagos collected samples on site. Confirmatory testing, sequencing, and virus isolation were performed at the Bernhard Nocht Institute for Tropical Medicine in Hamburg, Germany. Primary testing was done by reverse transcription–PCR (RT-PCR) that targeted the glycoprotein (GP) gene (5,6). An RT-PCR that targeted the large (L) gene was used as a secondary test (7), and PCR products were sequenced. Serologic testing for Lassa virus–specific immunoglobulin (Ig) G and IgM was performed by immunofluorescent antibody test using Vero cells infected with Lassa virus. Virus isolation with Vero cells was conducted in the BioSafety Level 4 laboratory in Hamburg. From 2005 through 2008, 10 cases of Lassa fever were confirmed by virus detection (cases 3–10) or implicated by epidemiologic investigation and serologic testing (cases 1 and 2) (Appendix Table). Case-patients 1–4 were involved in a nosocomial outbreak that occurred in February 2005 at the Ebonyi State University Teaching Hospital (EBSUTH) in Abakaliki. Retrospective investigation suggests the following transmission chain. The presumed index case-patient was a male nurse living in Onitsha, who became ill on January 21, 2005, and traveled ≈200 km to EBSUTH for better medical treatment. The detection of Lassa virus–specific IgM during his convalescent phase indicates that he had Lassa fever. The second case-patient was a female nurse who had contact with the index case-patient on February 4. She was admitted on February 7 and died 6 days later. Her clinical features were compatible with Lassa fever, but laboratory confirmation is lacking because specimens were not collected. Two additional case-patients among hospital staff (case-patients 3 and 4) were seen on February 21; each had had contact with case-patient 2. Case-patient 3 took care of case-patient 2 and slept in the same room with her for 4 days. Lassa fever was confirmed in case-patients 3 and 4 by RT-PCR as well as by IgM and IgG seroconversion in the surviving patient (case-patient 3). Case-patient 4, a pregnant nurse, had a spontaneous abortion and died on day 9 of hospitalization. Sequencing the GP and L gene PCR fragments showed that case-patients 3 and 4 were infected with the same virus strain (100% identity). In March and April 2005, blood was collected from 50 hospital staff members (including those who had had contact with the case-patients) and screened for Lassa virus–specific IgM and IgG. No positive blood samples were found, which indicated that no additional staff members were involved in the outbreak. Case-patients 5 and 6 were admitted to EBSUTH in 2008 on January 17 and March 5, respectively. Both were medical doctors, one at a local hospital and the other at EBSUTH, and both died. Encephalopathy with generalized seizures and loss of consciousness preceded death in both cases. The source of infection is unknown, although it is likely that they became infected while they treated patients without knowing they had Lassa fever. In agreement with the epidemiology, the viruses from the 2 patients were similar, though not identical (89% and 87% identity in the GP and L genes, respectively). Cases 7 to 10 occurred in Abuja and Jos from December 2007 through March 2008. Healthcare workers appeared not to be involved, and no molecular epidemiologic evidence indicated that transmission occurred among the 3 case-patients from Jos (94–97% and 90–94% identity in the GP and L genes, respectively). In conjunction with our previous report (4), the cases presented here demonstrate current Lassa fever activity in the states of Edo, Ebonyi, Federal Capital Territory, and Plateau. These findings correspond to early reports on Lassa fever in southern and central parts of Nigeria. That healthcare workers are still at as high a risk of contracting and dying from the disease as they were 20 years ago (8) is alarming. A key to solving this problem would be the establishment of diagnostic facilities that can provide rapid molecular testing at referral centers in the disease-endemic zones. This testing would facilitate appropriate case and contact management, including early treatment and postexposure prophylaxis with ribavirin, and eventually raise awareness that Lassa fever should be considered in every severe febrile illness in these regions.

New Hosts of The Lassa Virus.

Lassa virus (LASV) causes a deadly haemorrhagic fever in humans, killing several thousand people in West Africa annually. For 40 years, the Natal multimammate rat, Mastomys natalensis, has been assumed to be the sole host of LASV. We found evidence that LASV is also hosted by other rodent species: the African wood mouse Hylomyscus pamfi in Nigeria, and the Guinea multimammate mouse Mastomys erythroleucus in both Nigeria and Guinea. Virus strains from these animals were isolated in the BSL-4 laboratory and fully sequenced. Phylogenetic analyses of viral genes coding for glycoprotein, nucleoprotein, polymerase and matrix protein show that Lassa strains detected in M. erythroleucus belong to lineages III and IV. The strain from H. pamfi clusters close to lineage I (for S gene) and between II & III (for L gene). Discovery of new rodent hosts has implications for LASV evolution and its spread into new areas within West Africa.

Hospital-based surveillance for Lassa fever in Edo State, Nigeria, 2005-2008

Objectives  To estimate the burden of Lassa fever in northern and central Edo, a state in south Nigeria where Lassa fever has been reported.

The West African ebola virus disease epidemic 2014–2015: A commissioned review

The first epidemic of Ebola haemorrhagic disease in West Africa is the largest and longest Ebola epidemic till date, where the outbreak notably involved three countries with distant spread to other countries. It has caused significant mortality, with reported case fatality rates of up to 70%. Data and relevant information were extracted from the review of majorly relevant publications/papers about the Ebola epidemic in West Africa and other previous outbreaks of Ebola virus (EBOV). As of 2016, with the epidemic under control, the World Health Organization has warned that flare-ups of the disease are likely to continue for some time as recently occurred in Sierra Leone and the on-going in Guinea. As this may not be the last outbreak of Ebola virus disease (EVD) in West Africa, there is a need to focus on diagnostic and research capacity required to curtail EVD with adequate measures for emergency preparedness and policies for innovative treatment strategies.

Ebola Virus Epidemiology and Evolution in Nigeria

Abstract Containment limited the 2014 Nigerian Ebola virus (EBOV) disease outbreak to 20 reported cases and 8 fatalities. We present here clinical data and contact information for at least 19 case patients, and full-length EBOV genome sequences for 12 of the 20. The detailed contact data permits nearly complete reconstruction of the transmission tree for the outbreak. The EBOV genomic data are consistent with that tree. It confirms that there was a single source for the Nigerian infections, shows that the Nigerian EBOV lineage nests within a lineage previously seen in Liberia but is genetically distinct from it, and supports the conclusion that transmission from Nigeria to elsewhere did not occur.