Sequence Variability and Geographic Distribution of Lassa Virus, Sierra Leone - Volume 21, Number 4—April 2015 - Emerging Infectious Disease journal - CDC

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Sequence Variability and Geographic Distribution of Lassa Virus, Sierra Leone - Volume 21, Number 4—April 2015 - Emerging Infectious Disease journal - CDC

Volume 21, Number 4—April 2015

Research

Sequence Variability and Geographic Distribution of Lassa Virus, Sierra Leone

On This Page

• Methods
• Results and Discussion
• Suggested Citation

Figures

• Figure 1
• Figure 2
• Figure 3
• Figure 4

Tables

• Table 1
• Table 2
• Table 3

Technical Appendicies

• Technical Appendix

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Tomasz A. Leski1 , Michael G. Stockelman1, Lina M. Moses, Matthew Park, David A. Stenger, Rashid Ansumana, Daniel G. Bausch, and Baochuan Lin

Author affiliations: Naval Research Laboratory, Washington, DC, USA (T.A. Leski, M.G. Stockelman, D.A. Stenger, B. Lin); Tulane University, New Orleans, Louisiana, USA (L.M. Moses); Tulane School of Public Health and Tropical Medicine, New Orleans (L.M. Moses, D.G Bausch), Thomas Jefferson High School, Alexandria, Virginia, USA (M. Park); Mercy Hospital Research Laboratory, Bo, Sierra Leone (R. Ansumana); Liverpool School of Tropical Medicine, Liverpool, UK (R. Ansumana); Njala University, Bo (R. Ansumana)

Suggested citation for this article

Abstract

Lassa virus (LASV) is endemic to parts of West Africa and causes highly fatal hemorrhagic fever. The multimammate rat (Mastomys natalensis) is the only known reservoir of LASV. Most human infections result from zoonotic transmission. The very diverse LASV genome has 4 major lineages associated with different geographic locations. We used reverse transcription PCR and resequencing microarrays to detect LASV in 41 of 214 samples from rodents captured at 8 locations in Sierra Leone. Phylogenetic analysis of partial sequences of nucleoprotein (NP), glycoprotein precursor (GPC), and polymerase (L) genes showed 5 separate clades within lineage IV of LASV in this country. The sequence diversity was higher than previously observed; mean diversity was 7.01% for nucleoprotein gene at the nucleotide level. These results may have major implications for designing diagnostic tests and therapeutic agents for LASV infections in Sierra Leone.

Lassa fever (LF) belongs to a group of viral hemorrhagic fevers characterized by a febrile syndrome and high case-fatality rates (1). LF differs from most viral hemorrhagic fevers in that it is endemic to a large geographic area of sub-Saharan Africa. Human cases of LF have been reported in (or imported from) Guinea, Sierra Leone, Liberia, Mali, Burkina Faso, and Nigeria; however, LF outbreaks seem to be restricted to Guinea, Sierra Leone, Liberia (the Mano River Union region), and Nigeria (2–4). In some areas of Sierra Leone and Guinea, more than half of the population has antibodies against Lassa virus (LASV; family Arenaviridae), the etiologic agent of LF (5,6). According to various estimates, 300,000–500,000 cases of LF result in 5,000–10,000 deaths annually in West Africa (6,7). An analysis based on seroepidemiologic data suggested that the number of cases might be much higher, reaching 3 million cases and 67,000 fatalities per year (8). Overall, the population at risk might include as many as 200 million persons living in a large swath of West Africa from Senegal to Nigeria and beyond (4).

LASV can cause infection in the multimammate rat (Mastomys natalensis), a natural host and reservoir of this pathogen (9,10). The multimammate rat is a commensal rodent ubiquitous in Africa (11,12). Although the routes of LASV infection are poorly characterized, humans probably get infected by eating contaminated food (13), by inhaling virus-contaminated aerosols (14), or while butchering infected rat meat (15). Person-to-person transmission of LASV is well documented, mostly in the form of nosocomial outbreaks (13).

Like other arenaviruses, LASV is an enveloped virus with a bisegmented single-stranded RNA genome encoding 4 proteins using an ambisense coding strategy (16). The small segment contains genes for the glycoprotein precursor (GPC) and nucleoprotein (NP), which serves as the main viral capsid protein. The large segment encodes the small zinc-binding protein (Z), which contains a RING motif, and another gene (L) containing the RNA-dependent RNA polymerase domain.

Complete genome sequences are available for several LASV strains, as are a considerable number of partial sequences from isolates originating from humans and rodents (17–20). Their analysis revealed the existence of high sequence diversity (up to 27% nt) and 4 major lineages of LASV, which correlate with geographic location (17). Lineages I, II, and III, and the greatest diversity of LASV strains, were found among isolates from Nigeria, whereas strains from Guinea, Sierra Leone, and Liberia seemed to be more closely related and belong exclusively to lineage IV. Sequence of the AV strain (21) and recently published sequences from rodent LASV isolates from Mali (18) suggest the existence of an additional clade (proposed as lineage V) (22). LASV sequences of isolates from humans and rodents are found interspersed throughout the phylogenetic tree, which is consistent with the notion that human cases typically result from transmission from rodents (17).

The high degree of sequence divergence of LASV genomes is a major problem affecting the development of molecular and immune-based diagnostic technologies, vaccines, and possibly antiviral drugs (13,16,17,23–25). Forty-seven unique partial LASV sequences from Sierra Leone were available in GenBank at the time of this analysis, which included fragments of NP (27 sequences), GPC (9 sequences), L (9 sequences), and Z (2 sequences) genes plus full sequences of small and large segments of 2 strains—Josiah and NL. Most of these sequences are from isolates collected >30 years ago; only 2 more recent sequences (GPC and L gene fragments) from strain SL06-2057 were isolated in 2006 (17,19).

To fill this gap, we investigated the sequence diversity of strains circulating among small rodents captured in peridomestic settings in Sierra Leone. In 2014, we screened 214 samples collected during 2009 from several species of rodents trapped in villages where LF was reported in humans. We used diagnostic reverse transcription PCR (RT-PCR) and high-density resequencing microarrays to detect LASV and amplify fragments of NP, GPC, and L genes. The obtained amplicons were sequenced and compared with previously published sequences from Sierra Leone to obtain a more complete and updated picture of the strains circulating in this country.

Dr. Leski is a research biologist at the Center for Bio/Molecular Science and Engineering at the Naval Research Laboratory. His research interests include the development and application of molecular diagnostics for pathogen detection and tracking the spread of antimicrobial resistance determinants in bacterial pathogens.

Acknowledgments

We thank Benjamin Kirkup and Zheng Wang for their critical evaluation of this manuscript.

Funding for this project was provided by the Office of Naval Research. M.P. was a Science and Engineering Apprenticeship Program (SEAP) summer intern supported by the American Society for Engineering Education as part of the Office of Naval Research, SEAP, at the Naval Research Laboratory.

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Figures

• Figure 1. A) Locations of origin for Lassa virus (LASV) nucleic acid sequences, Sierra Leone. B) Enlarged view of region from which rodent specimens were collected. Major roads (red) and waterways...
• Figure 2. Phylogenetic analysis of Lassa virus isolates from Sierra Leone based on partial nucleoprotein (NP) gene sequences. The homologous NP fragments of 621 nt were aligned. The isolate Z-158, which...
• Figure 3. Phylogenetic analysis of Lassa virus (LASV) isolates from Sierra Leone based on partial glycoprotein precursor (GPC) gene sequences. The homologous GPC fragments of 284 nt were aligned. The isolate...
• Figure 4. Phylogenetic analysis of Lassa virus (LASV) isolates from Sierra Leone based on partial polymerase (L) gene sequences. The homologous L fragments of 373 nt were aligned. The isolate Z-158,...

Tables

• Table 1. PCR and sequencing primers used in study of Lassa virus, Sierra Leone
• Table 2. Results of Lassa virus detection among rodent samples with >1 positive test result, Sierra Leone
• Table 3. Estimates of average evolutionary divergence of NP, GPC, and L gene fragments for Lassa virus strains, Sierra Leone

Technical Appendix

• Technical Appendix. Rodent trapping procedures; coordinates of sampling locations; and list of all analyzed samples.  126 KB

Suggested citation for this article: Leski TA, Stockelman MG, Moses LM, Park M, Stenger DA, Ansumana R, et al. Sequence variability and geographic distribution of Lassa virus, Sierra Leone. Emerg Infect Dis [Internet]. 2015 Apr [date cited]. http://dx.doi.org/10.3201/eid2104.141469

DOI: 10.3201/eid2104.141469

1These authors contributed equally to this article.