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      Genetic characterization of Lassa virus strains isolated from 2012 to 2016 in southeastern Nigeria

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          Abstract

          Lassa virus (LASV) is endemic in parts of West Africa where it causes Lassa fever (LF), a viral hemorrhagic fever with frequent fatal outcomes. The diverse LASV strains are grouped into six major lineages based on the geographical location of the isolated strains. In this study, we have focused on the lineage II strains from southern Nigeria. We determined the viral sequences from positive cases of LF reported at tertiary hospitals in Ebonyi and Enugu between 2012 and 2016. Reverse transcription-polymerase chain reaction (RT-PCR) showed that 29 out of 123 suspected cases were positive for the virus among which 11 viral gene sequences were determined. Phylogenetic analysis of the complete coding sequences of the four viral proteins revealed that lineage II strains are broadly divided into two genetic clades that diverged from a common ancestor 195 years ago. One clade, consisting of strains from Ebonyi and Enugu, was more conserved than the other from Irrua, although the four viral proteins were evolving at similar rates in both clades. These results suggested that the viruses of these clades have been distinctively evolving in geographically separate parts of southern Nigeria. Furthermore, the epidemiological data of the 2014 outbreak highlighted the role of human-to-human transmission in this outbreak, which was supported by phylogenetic analysis showing that 13 of the 16 sequences clustered together. These results provide new insights into the evolution of LASV in southern Nigeria and have important implications for vaccine development, diagnostic assay design, and LF outbreak management.

          Author summary

          Lassa fever (LF) is a viral hemorrhagic fever caused by Lassa virus (LASV). The different LASV strains are grouped into lineages based on the geographical location of the isolated strains. The aim of our study was to characterize the lineage II strains in southern Nigeria. We sequenced LASV RNA genome from positive cases of LF between 2012 and 2016 which were reported at tertiary hospitals in Ebonyi and Enugu in southeastern Nigeria. Phylogenetic analysis of the viral proteins showed the division of lineage II strains into two genetic clades with one clade being more conserved than the other despite evolving at similar rates. Also, our phylogenetic analysis supported the role of human to human transmission in the 2014 outbreak, in keeping with the epidemiological data. These results provide additional information on the evolution of LASV in southern Nigeria and LF outbreak management.

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          Most cited references10

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          Lassa fever.

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            Improved detection of Lassa virus by reverse transcription-PCR targeting the 5' region of S RNA.

            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.
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              New Lineage of Lassa Virus, Togo, 2016

              We describe a strain of Lassa virus representing a putative new lineage that was isolated from a cluster of human infections with an epidemiologic link to Togo. This finding extends the known range of Lassa virus to Togo.
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                Author and article information

                Contributors
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: VisualizationRole: Writing – original draft
                Role: Project administrationRole: ResourcesRole: SupervisionRole: Writing – review & editing
                Role: Investigation
                Role: Investigation
                Role: Investigation
                Role: Investigation
                Role: Resources
                Role: Resources
                Role: Resources
                Role: Resources
                Role: Resources
                Role: Resources
                Role: MethodologyRole: ValidationRole: Writing – review & editing
                Role: ConceptualizationRole: MethodologyRole: ValidationRole: Writing – review & editing
                Role: ConceptualizationRole: MethodologyRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: MethodologyRole: Project administrationRole: SupervisionRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS Negl Trop Dis
                PLoS Negl Trop Dis
                plos
                plosntds
                PLoS Neglected Tropical Diseases
                Public Library of Science (San Francisco, CA USA )
                1935-2727
                1935-2735
                30 November 2018
                November 2018
                : 12
                : 11
                : e0006971
                Affiliations
                [1 ] Department of Emerging Infectious Diseases, Institute of Tropical Medicine (NEKKEN), Nagasaki University, Nagasaki, Japan
                [2 ] Graduate School of Biomedical Sciences and Program for Nurturing Global Leaders in Tropical and Emerging Communicable Diseases, Nagasaki University, Nagasaki, Japan
                [3 ] Department of Medicine, Federal Teaching Hospital Abakaliki, Abakaliki, Ebonyi, Nigeria
                [4 ] Department of Medicine, University of Nigeria Teaching Hospital, Ituku-Ozalla, Enugu, Nigeria
                [5 ] Department of Medicine, College of Medicine, University of Nigeria, Ituku-Ozalla, Enugu, Nigeria
                [6 ] Department of Microbiology, University of Nigeria Teaching Hospital, Ituku-Ozalla, Enugu, Nigeria
                [7 ] Department of Community Medicine, College of Medicine, University of Nigeria, Ituku-Ozalla, Enugu, Nigeria
                [8 ] National Research Center for the Control and Prevention of Infectious Diseases (CCPID), Nagasaki University, Nagasaki, Japan
                The University of Kansas, UNITED STATES
                Author notes

                The authors have declared that no competing interests exist.

                Article
                PNTD-D-18-01169
                10.1371/journal.pntd.0006971
                6267959
                30500827
                6e3e8660-132f-4014-84c4-bcde16382557
                © 2018 Oloniniyi et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 2 August 2018
                : 3 November 2018
                Page count
                Figures: 4, Tables: 3, Pages: 13
                Funding
                Funded by: Japan Agency for Medical Research and Development
                Award ID: 17fk0108101h0001
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001691, Japan Society for the Promotion of Science;
                Award ID: JP25305016
                Award Recipient :
                This work was supported by grants from Japan Agency for Medical Research and Development (17fk0108101h0001 to JY https://www.amed.go.jp/en/index.html) and Japan Society for Promotion of Science (JSPS) Grant-in-Aid for Scientific Research (JP25305016 to JY https://www.jsps.go.jp/english/index.html). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Biology and Life Sciences
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Computer and Information Sciences
                Data Management
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                People and Places
                Geographical Locations
                Africa
                Nigeria
                Research and Analysis Methods
                Database and Informatics Methods
                Bioinformatics
                Sequence Analysis
                Research and analysis methods
                Database and informatics methods
                Bioinformatics
                Sequence analysis
                DNA sequence analysis
                Biology and Life Sciences
                Molecular Biology
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Reverse Transcriptase-Polymerase Chain Reaction
                Research and Analysis Methods
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Reverse Transcriptase-Polymerase Chain Reaction
                Research and Analysis Methods
                Database and Informatics Methods
                Bioinformatics
                Sequence Analysis
                Sequence Alignment
                Medicine and Health Sciences
                Tropical Diseases
                Neglected Tropical Diseases
                Lassa Fever
                Medicine and Health Sciences
                Infectious Diseases
                Viral Diseases
                Lassa Fever
                Research and Analysis Methods
                Database and Informatics Methods
                Bioinformatics
                Sequence Analysis
                Amino Acid Sequence Analysis
                Custom metadata
                All relevant data are within the paper and its Supporting Information files.

                Infectious disease & Microbiology
                Infectious disease & Microbiology

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