Publications by authors named "Onikepe Folarin"

45 Publications

Polymorphisms in Plasmodium falciparum dihydropteroate synthetase and dihydrofolate reductase genes in Nigerian children with uncomplicated malaria using high-resolution melting technique.

Sci Rep 2021 Jan 12;11(1):471. Epub 2021 Jan 12.

African Centre of Excellence for Genomics of Infectious Diseases, Redeemer's University, Ede, Nigeria.

In 2005, the Nigerian Federal Ministry of Health revised the treatment policy for uncomplicated malaria with the introduction of artemisinin-based combination therapies (ACTs). This policy change discouraged the use of Sulphadoxine-pyrimethamine (SP) as the second-line treatment of uncomplicated falciparum malaria. However, SP is used as an intermittent preventive treatment of malaria in pregnancy (IPTp) and seasonal malaria chemoprevention (SMC) in children aged 3-59 months. There have been increasing reports of SP resistance especially in the non-pregnant population in Nigeria, thus, the need to continually monitor the efficacy of SP as IPTp and SMC by estimating polymorphisms in dihydropteroate synthetase (dhps) and dihydrofolate reductase (dhfr) genes associated with SP resistance. The high resolution-melting (HRM) assay was used to investigate polymorphisms in codons 51, 59, 108 and 164 of the dhfr gene and codons 437, 540, 581 and 613 of the dhps gene. DNA was extracted from 271 dried bloodspot filter paper samples obtained from children (< 5 years old) with uncomplicated malaria. The dhfr triple mutant IRN, dhps double mutant GG and quadruple dhfr IRN + dhps G mutant haplotypes were observed in 80.8%, 13.7% and 52.8% parasites, respectively. Although the quintuple dhfr IRN + dhps GE and sextuple dhfr IRN + dhps GEG mutant haplotypes linked with in-vivo and in-vitro SP resistance were not detected, constant surveillance of these haplotypes should be done in the country to detect any change in prevalence.
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http://dx.doi.org/10.1038/s41598-020-80017-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7803958PMC
January 2021

Polymorphisms in Plasmodium falciparum chloroquine resistance transporter (Pfcrt) and multidrug-resistant gene 1 (Pfmdr-1) in Nigerian children 10 years post-adoption of artemisinin-based combination treatments.

Int J Parasitol 2021 Mar 24;51(4):301-310. Epub 2020 Dec 24.

African Centre of Excellence for Genomics of Infectious Diseases, Redeemer's University, Ede, Nigeria; Department of Biological Sciences, Redeemer's University, Ede, Nigeria; Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA, USA. Electronic address:

The emergence and spread of Plasmodium falciparum parasites resistant to artemisinin derivatives and their partners in southeastern Asia threatens malaria control and elimination efforts, and heightens the need for an alternative therapy. We have explored the distribution of P. falciparum chloroquine resistance transporter (Pfcrt) and multidrug-resistant gene 1 (Pfmdr-1) haplotypes 10 years following adoption of artemisinin-based combination therapies in a bid to investigate the possible re-emergence of Chloroquine-sensitive parasites in Nigeria, and investigated the effect of these P. falciparum haplotypes on treatment outcomes of patients treated with artemisinin-based combination therapies. A total of 271 children aged <5 years with uncomplicated falciparum malaria were included in this study. Polymorphisms on codons 72-76 of the Pfcrt gene and codon 86 and 184 of Pfmdr-1 were determined using the high resolution melting assay. Of 240 (88.6%) samples successfully genotyped with HRM for Pfcrt, wildtype CMNK (42.9%) and mutant CIET (53.8%) were observed. Also, wildtype NY (62.9%) and mutant NF (21.1%), YY (6.4%), and YF (0.4%) haplotypes of Pfmdr-1 were observed. Measures of responsiveness to ACTs were similar in children infected with P. falciparum crt haplotypes (CIET and CMNK) and major mdr-1 haplotypes (NY, NF and YY). Despite a 10 year gap since the malaria treatment policy changed to ACTs, over 50% of the P. falciparum parasites investigated in this study harboured the Chloroquine-resistant CIET haplotype, however this did not compromise the efficacy of artemisinin-based combination therapies. Should complete artemisinin resistance emerge from or spread to Nigeria, chloroquine might not be a good alternative therapy.
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http://dx.doi.org/10.1016/j.ijpara.2020.10.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7940560PMC
March 2021

Antibodies from Sierra Leonean and Nigerian Lassa fever survivors cross-react with recombinant proteins representing Lassa viruses of divergent lineages.

Sci Rep 2020 09 29;10(1):16030. Epub 2020 Sep 29.

Ahmadu Bello University, Zaria, Nigeria.

Lassa virus (LASV) is the causative agent of Lassa fever, an often-fatal hemorrhagic disease that is endemic in West Africa. Seven genetically distinct LASV lineages have been identified. As part of CEPI's (Coalition for Epidemic Preparedness Innovations) Lassa vaccine development program, we assessed the potential of the human immune system to mount cross-reactive and cross-protective humoral immune responses to antigens from the most prevalent LASV lineages, which are lineages II and III in Nigeria and lineage IV in Sierra Leone. IgG and IgM present in the blood of Lassa fever survivors from Nigeria or Sierra Leone exhibited substantial cross-reactivity for binding to LASV nucleoprotein and two engineered (linked and prefusion) versions of the glycoproteins (GP) of lineages II-IV. There was less cross-reactivity for the Zinc protein. Serum or plasma from Nigerian Lassa fever survivors neutralized LASV pseudoviruses expressing lineage II GP better than they neutralized lineage III or IV GP expressing pseudoviruses. Sierra Leonean survivors did not exhibit a lineage bias. Neutralization titres determined using LASV pseudovirus assays showed significant correlation with titres determined by plaque reduction with infectious LASV. These studies provide guidance for comparison of humoral immunity to LASV of distinct lineages following natural infection or immunization.
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http://dx.doi.org/10.1038/s41598-020-72539-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7525497PMC
September 2020

Field evaluation of a Pan-Lassa rapid diagnostic test during the 2018 Nigerian Lassa fever outbreak.

Sci Rep 2020 05 26;10(1):8724. Epub 2020 May 26.

Viral Hemorrhagic Fever Program, Kenema Government Hospital, Kenema, Sierra Leone.

Lassa virus (LASV) is the causative agent of Lassa fever (LF), an often-fatal hemorrhagic disease. LF is endemic in Nigeria, Sierra Leone and other West African countries. Diagnosis of LASV infection is challenged by the genetic diversity of the virus, which is greatest in Nigeria. The ReLASV Pan-Lassa Antigen Rapid Test (Pan-Lassa RDT) is a point-of-care, in vitro diagnostic test that utilizes a mixture of polyclonal antibodies raised against recombinant nucleoproteins of representative strains from the three most prevalent LASV lineages (II, III and IV). We compared the performance of the Pan-LASV RDT to available quantitative PCR (qPCR) assays during the 2018 LF outbreak in Nigeria. For patients with acute LF (RDT positive, IgG/IgM negative) during initial screening, RDT performance was 83.3% sensitivity and 92.8% specificity when compared to composite results of two qPCR assays. 100% of samples that gave Ct values below 22 on both qPCR assays were positive on the Pan-Lassa RDT. There were significantly elevated case fatality rates and elevated liver transaminase levels in subjects whose samples were RDT positive compared to RDT negative.
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http://dx.doi.org/10.1038/s41598-020-65736-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7250850PMC
May 2020

Identification of Common CD8 T Cell Epitopes from Lassa Fever Survivors in Nigeria and Sierra Leone.

J Virol 2020 06 1;94(12). Epub 2020 Jun 1.

Viral Immunobiology Laboratory, Department of Immunology and Microbiology, The Scripps Research Institute, La Jolla, California, USA

Early and robust T cell responses have been associated with survival from Lassa fever (LF), but the Lassa virus-specific memory responses have not been well characterized. Regions within the virus surface glycoprotein (GPC) and nucleoprotein (NP) are the main targets of the Lassa virus-specific T cell responses, but, to date, only a few T cell epitopes within these proteins have been identified. We identified GPC and NP regions containing T cell epitopes and HLA haplotypes from LF survivors and used predictive HLA-binding algorithms to identify putative epitopes, which were then experimentally tested using autologous survivor samples. We identified 12 CD8-positive (CD8) T cell epitopes, including epitopes common to both Nigerian and Sierra Leonean survivors. These data should be useful for the identification of dominant Lassa virus-specific T cell responses in Lassa fever survivors and vaccinated individuals as well as for designing vaccines that elicit cell-mediated immunity. The high morbidity and mortality associated with clinical cases of Lassa fever, together with the lack of licensed vaccines and limited and partially effective interventions, make Lassa virus (LASV) an important health concern in its regions of endemicity in West Africa. Previous infection with LASV protects from disease after subsequent exposure, providing a framework for designing vaccines to elicit similar protective immunity. Multiple major lineages of LASV circulate in West Africa, and therefore, ideal vaccine candidates should elicit immunity to all lineages. We therefore sought to identify common T cell epitopes between Lassa fever survivors from Sierra Leone and Nigeria, where distinct lineages circulate. We identified three such epitopes derived from highly conserved regions within LASV proteins. In this process, we also identified nine other T cell epitopes. These data should help in the design of an effective pan-LASV vaccine.
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http://dx.doi.org/10.1128/JVI.00153-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7307091PMC
June 2020

High crossreactivity of human T cell responses between Lassa virus lineages.

PLoS Pathog 2020 03 6;16(3):e1008352. Epub 2020 Mar 6.

Viral-Immunobiology Laboratory, Department of Immunology and Microbiology, The Scripps Research Institute, La Jolla, California, United States of America.

Lassa virus infects hundreds of thousands of people each year across rural West Africa, resulting in a high number of cases of Lassa fever (LF), a febrile disease associated with high morbidity and significant mortality. The lack of approved treatments or interventions underscores the need for an effective vaccine. At least four viral lineages circulate in defined regions throughout West Africa with substantial interlineage nucleotide and amino acid diversity. An effective vaccine should be designed to elicit Lassa virus specific humoral and cell mediated immunity across all lineages. Most current vaccine candidates use only lineage IV antigens encoded by Lassa viruses circulating around Sierra Leone, Liberia, and Guinea but not Nigeria where lineages I-III are found. As previous infection is known to protect against disease from subsequent exposure, we sought to determine whether LF survivors from Nigeria and Sierra Leone harbor memory T cells that respond to lineage IV antigens. Our results indicate a high degree of cross-reactivity of CD8+ T cells from Nigerian LF survivors to lineage IV antigens. In addition, we identified regions within the Lassa virus glycoprotein complex and nucleoprotein that contributed to these responses while T cell epitopes were not widely conserved across our study group. These data are important for current efforts to design effective and efficient vaccine candidates that can elicit protective immunity across all Lassa virus lineages.
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http://dx.doi.org/10.1371/journal.ppat.1008352DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7080273PMC
March 2020

Real-time Metagenomic Analysis of Undiagnosed Fever Cases Unveils a Yellow Fever Outbreak in Edo State, Nigeria.

Sci Rep 2020 02 21;10(1):3180. Epub 2020 Feb 21.

African Centre of Excellence for Genomics of Infectious Diseases (ACEGID), Redeemer's University, Ede, Osun State, Nigeria.

Fifty patients with unexplained fever and poor outcomes presented at Irrua Specialist Teaching Hospital (ISTH) in Edo State, Nigeria, an area endemic for Lassa fever, between September 2018 - January 2019. After ruling out Lassa fever, plasma samples from these epidemiologically-linked cases were sent to the African Centre of Excellence for Genomics of Infectious Diseases (ACEGID), Redeemer's University, Ede, Osun State, Nigeria, where we carried out metagenomic sequencing which implicated yellow fever virus (YFV) as the etiology of this outbreak. Twenty-nine of the 50 samples were confirmed positive for YFV by reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR), 14 of which resulted in genome assembly. Maximum likelihood phylogenetic analysis revealed that these YFV sequences formed a tightly clustered clade more closely related to sequences from Senegal than sequences from earlier Nigerian isolates, suggesting that the YFV clade responsible for this outbreak in Edo State does not descend directly from the Nigerian YFV outbreaks of the last century, but instead reflects a broader diversity and dynamics of YFV in West Africa. Here we demonstrate the power of metagenomic sequencing for identifying ongoing outbreaks and their etiologies and informing real-time public health responses, resulting in accurate and prompt disease management and control.
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http://dx.doi.org/10.1038/s41598-020-59880-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7035389PMC
February 2020

Clinical illness and outcomes in Nigerian children with persistent early-appearing anaemia following initiation of artemisinin-based combination treatments of uncomplicated falciparum malaria.

Parasite 2019 13;26:56. Epub 2019 Sep 13.

Antimalarial Therapeutic Efficacy Monitoring Group, National Malaria Elimination Programme, The Federal Ministry of Health, Abuja 900211, Nigeria - Institute for Medical Research and Training, College of Medicine, University of Ibadan, Ibadan 200212, Nigeria - Department of Pharmacology and Therapeutics, College of Medicine, University of Ibadan, Ibadan 200284, Nigeria - Department of Clinical Pharmacology, University College Hospital, Ibadan 200212, Nigeria.

In non-anaemic children with malaria, early-appearing anaemia (EAA) is common following artemisinin-based combination treatments (ACTs) and it may become persistent (PEAA). The factors contributing to and kinetics of resolution of the deficit in haematocrit from baseline (DIHFB) characteristic of ACTs-related PEAA were evaluated in 540 consecutive children with malaria treated with artemether-lumefantrine, artesunate-amodiaquine or dihydroartemisinin-piperaquine. Asymptomatic PEAA occurred in 62 children. In a multiple logistic regression model, a duration of illness ≤3 days before presentation, haematocrit <35% before and <25% one day after treatment initiation, drug attributable fall in haematocrit ≥6%, and treatment with dihydroartemisinin-piperaquine independently predicted PEAA. Overall, mean DIHFB was 5.7% (95% CI 4.8-6.6) 7 days after treatment initiation and was similar for all treatments. Time to 90% reduction in DIHFB was significantly longer in artemether-lumefantrine-treated children compared with other treatments. In a one compartment model, declines in DIHFB were monoexponential with overall mean estimated half-time of 3.9 days (95% CI 2.6-5.1), Cmax of 7.6% (95% CI 6.7-8.4), and Vd of 0.17 L/kg (95% CI 0.04-0.95). In Bland-Altman analyses, overall mean anaemia recovery time (AnRT) of 17.4 days (95% CI 15.5-19.4) showed insignificant bias with 4, 5 or 6 multiples of half-time of DIHFB. Ten children after recovery from PEAA progressed to late-appearing anaemia (LAA). Progression was associated with female gender and artesunate-amodiaquine treatment. Asymptomatic PEAA is common following ACTs. PEAA or its progression to LAA may have implications for case and community management of anaemia and for anaemia control efforts in sub-Saharan Africa where ACTs have become first-line antimalarials. Trial registration: Pan Africa Clinical Trial Registration PACTR201709002064150, 1 March 2017 http://www.pactr.org.
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http://dx.doi.org/10.1051/parasite/2019058DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6743267PMC
March 2020

Declining responsiveness of childhood Plasmodium falciparum infections to artemisinin-based combination treatments ten years following deployment as first-line antimalarials in Nigeria.

Infect Dis Poverty 2019 Aug 6;8(1):69. Epub 2019 Aug 6.

Department of Pharmacology and Therapeutics, University of Ibadan, Ibadan, Nigeria.

Background: The development and spread of artemisinin-resistant Plasmodium falciparum malaria in Greater Mekong Subregion has created impetus for continuing global monitoring of efficacy of artemisinin-based combination therapies (ACTs). This post analyses is aimed to evaluate changes in early treatment response markers 10 years after the adoption of ACTs as first-line treatments of uncomplicated falciparum malaria in Nigeria.

Methods: At 14 sentinel sites in six geographical areas of Nigeria, we evaluated treatment responses in 1341 children under 5 years and in additional 360 children under 16 years with uncomplicated malaria enrolled in randomized trials of artemether-lumefantrine versus artesunate-amodiaquine at 5-year interval in 2009-2010 and 2014-2015 and at 2-year interval in 2009-2010 and 2012-2015, respectively after deployment in 2005.

Results: Asexual parasite positivity 1 day after treatment initiation (APPD1) rose from 54 to 62% and 2 days after treatment initiation from 5 to 26% in 2009-2010 to 2014-2015 (P = 0.002 and P <  0.0001, respectively). Parasite clearance time increased significantly from 1.6 days (95% confidence interval [CI]: 1.55-1.64) to 1.9 days (95% CI, 1.9-2.0) and geometric mean parasite reduction ratio 2 days after treatment initiation decreased significantly from 11 000 to 4700 within the same time period (P <  0.0001 for each). Enrolment parasitaemia > 75 000 μl, haematocrit > 27% 1 day post-treatment initiation, treatment with artemether-lumefantrine and enrolment in 2014-2015 independently predicted APPD1. In parallel, Kaplan-Meier estimated risk of recurrent infections by day 28 rose from 8 to 14% (P = 0.005) and from 9 to 15% (P = 0.02) with artemether-lumefantrine and artesunate-amodiaquine, respectively. Mean asexual parasitaemia half-life increased significantly from 1.1 h to 1.3 h within 2 years (P <  0.0001).

Conclusions: These data indicate declining parasitological responses through time to the two ACTs may be due to emergence of parasites with reduced susceptibility or decrease in immunity to the infections in these children.

Trial Registration: Pan African Clinical Trial Registration PACTR201508001188143 , 3 July 2015; PACTR201508001191898 , 7 July 2015 and PACTR201508001193368 , 8 July 2015 PACTR201510001189370 , 3 July 2015; PACTR201709002064150 , 1 March 2017; https://www.pactr.samrca.ac.za.
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http://dx.doi.org/10.1186/s40249-019-0577-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6683392PMC
August 2019

Capturing sequence diversity in metagenomes with comprehensive and scalable probe design.

Nat Biotechnol 2019 02 4;37(2):160-168. Epub 2019 Feb 4.

Division of Infectious Diseases and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA.

Metagenomic sequencing has the potential to transform microbial detection and characterization, but new tools are needed to improve its sensitivity. Here we present CATCH, a computational method to enhance nucleic acid capture for enrichment of diverse microbial taxa. CATCH designs optimal probe sets, with a specified number of oligonucleotides, that achieve full coverage of, and scale well with, known sequence diversity. We focus on applying CATCH to capture viral genomes in complex metagenomic samples. We design, synthesize, and validate multiple probe sets, including one that targets the whole genomes of the 356 viral species known to infect humans. Capture with these probe sets enriches unique viral content on average 18-fold, allowing us to assemble genomes that could not be recovered without enrichment, and accurately preserves within-sample diversity. We also use these probe sets to recover genomes from the 2018 Lassa fever outbreak in Nigeria and to improve detection of uncharacterized viral infections in human and mosquito samples. The results demonstrate that CATCH enables more sensitive and cost-effective metagenomic sequencing.
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http://dx.doi.org/10.1038/s41587-018-0006-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6587591PMC
February 2019

Characterization of Plasmodium falciparum structure in Nigeria with malaria SNPs barcode.

Malar J 2018 Dec 17;17(1):472. Epub 2018 Dec 17.

African Centre of Excellence for Genomics of Infectious Diseases, Redeemer's University, Ede, Osun State, Nigeria.

Background: Plasmodium falciparum malaria remains a major health challenge in Nigeria despite the global decline of its incidence and mortality rates. Although significant progress has been made in preventing the transmission of P. falciparum and controlling the spread of the infection, there is much to be done in the area of proper monitoring, surveillance of the parasite, investigating the population dynamics and drug resistance profiling of the parasite as these are important to its eventual eradication. Polymorphic loci of msp1, msp2 and/or glurp genes or microsatellites have been traditionally used to characterize P. falciparum population structure in various parts of Nigeria. The lack of standardization in the interpretation of results, as well as the inability of these methods to distinguish closely related parasites, remains a limitation of these techniques. Conversely, the recently developed 24 single nucleotide polymorphism (SNP)-based molecular barcode assay has the possibility of differentiating between closely related parasites and offer additional information in determining the population diversity of P. falciparum within and between parasite populations. This study is therefore aimed at defining the population diversity of P. falciparum in and between two localities in Nigeria using the SNPs barcode technique.

Methods: The 24-SNP high-resolution melt (HRM) barcode assay and msp2 genotyping was used to investigate both intra and inter population diversity of the parasite population in two urban cities of Nigeria.

Results: Based on SNP barcode analysis, polygenomic malaria infections were observed in 17.9% and 13.5% of population from Enugu and Ibadan, respectively, while msp2 analyses showed 21% and 19.4% polygenomic infections in Enugu and Ibadan, respectively. Low levels of genetic diversity (π) of 0.328 and 0.318 were observed in Enugu and Ibadan parasite populations, respectively, while the F value of 0.02 (p = 0.055) was obtained when the genetic divergence of both populations was considered.

Conclusions: The 24-SNP barcode assay was effective in analysing P. falciparum population diversity. This study also showed that P. falciparum populations in Enugu and Ibadan had a degree of intra-population diversity, but very low divergence between the population. A low degree of polygenomic infections were also observed in the two parasite populations unlike previous years. This maybe as a result of the effect of artemisinin-based combination therapy (ACT), long-lasting insecticide-treated nets (LLITNs) and intermittent preventive treatments in the study populations.
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http://dx.doi.org/10.1186/s12936-018-2623-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6296064PMC
December 2018

Parasite reduction ratio one day after initiation of artemisinin-based combination therapies and its relationship with parasite clearance time in acutely malarious children.

Infect Dis Poverty 2018 Dec 7;7(1):122. Epub 2018 Dec 7.

Department of Pharmacology and Therapeutics, University of Ibadan, Ibadan, Nigeria.

Background: In acute falciparum malaria, asexual parasite reduction ratio two days post-treatment initiation (PRRD2) ≥ 10 000 per cycle has been used as a measure of the rapid clearance of parasitaemia and efficacy of artemisinin derivatives. However, there is little evaluation of alternative measures; for example, parasite reduction ratio one day after treatment initiation (PRRD1) and its relationship with parasite clearance time (PCT) or PRRD2. This study evaluated the use of PRRD1 as a measure of responsiveness to antimalarial drugs.

Methods: In acutely malarious children treated with artesunate-amodiaquine (AA), artemether-lumefantrine (AL) or dihydroartemisinin-piperaquine (DHP), the relationships between PRRD1 or PRRD2 and PCT, and between PRRD1 and PRRD2 were evaluated using linear regression. Agreement between estimates of PCT using PRRD1 and PRRD2 linear regression equations was evaluated using the Bland-Altman analysis. Predictors of PRRD1 > 5000 per half cycle and PRRD2 ≥ 10 000 per cycle were evaluated using stepwise multiple logistic regression models. Using the linear regression equation of the relationship between PRRD1 and PCT previously generated in half of the DHP-treated children during the early study phase, PCT estimates were compared in a prospective blinded manner with PCTs determined by microscopy during the later study phase in the remaining half.

Results: In 919 malarious children, PRRD1 was significantly higher in DHP- and AA-treated compared with AL-treated children (P <  0.0001). PRRD1 or PRRD2 values correlated significantly negatively with PCT values (P <  0.0001 for each) and significantly positively with each other (P <  0.0001). PCT estimates from linear regression equations for PRRD1 and PRRD2 showed insignificant bias on the Bland-Altman plot (P = 0.7) indicating the estimates can be used interchangeably. At presentation, age > 15 months, parasitaemia > 10 000/μl and DHP treatment independently predicted PRRD1 > 5000 per half cycle, while age > 30 months, haematocrit ≥31%, body temperature > 37.4 °C, parasitaemia > 100 000/μl, PRRD1 value > 1000 and no gametocytaemia independently predicted PRRD2 ≥ 10 000 per cycle. Using the linear regression equation generated during the early phase in 166 DHP-treated children, PCT estimates and PCTs determined by microscopy in the 155 children in the later phase were similar in the same patients.

Conclusions: PRRD1 and estimates of PCT using PRRD1 linear regression equation of PRRD1 and PCT can be used in therapeutic efficacy studies.

Trial Registration: Pan African Clinical Trial Registration PACTR201709002064150, 1 March 2017, http://www.pactr.org.
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http://dx.doi.org/10.1186/s40249-018-0503-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6284283PMC
December 2018

Genomic Analysis of Lassa Virus during an Increase in Cases in Nigeria in 2018.

N Engl J Med 2018 11 17;379(18):1745-1753. Epub 2018 Oct 17.

From the Broad Institute of the Massachusetts Institute of Technology (MIT) and Harvard University (K.J.S., K.G.B., S.M., S.F.S., S.M.W., R.R.S., J.Q., S.W., P.B., S.Y., B.C., D.K., A.C., A.G.-Y., C.A.F., D.J.P., N.L.Y., B.L.M., P.C.S.), the Center for Systems Biology, Department of Organismic and Evolutionary Biology (K.J.S., K.G.B., S.F.S., S.W., B.C., D.K., C.A.F., N.L.Y., P.C.S.), and the Faculty of Arts and Sciences (M.N.), Harvard University, Harvard University Extension School (R.R.S.), and Harvard-MIT Health Sciences and Technology, MIT (S.Y., P.C.S.), Cambridge, and the Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health (K.G.B., S.F.S., B.L.M., P.C.S., C.T.H.), and Beth Israel Deaconess Medical Center, Division of Infectious Diseases (S.M.), Boston - all in Massachusetts; the African Center of Excellence for Genomics of Infectious Diseases (P.E., J.U.O., E.U., T.K., F.A., J.U., A.G., M.M., I.N., P.O., O.T., O.A.F., C.T.H.) and the Department of Biological Sciences, College of Natural Sciences (J.U.O., E.U., T.K., F.A., J.U., P.O., O.A.F., C.T.H.), Redeemer's University, Ede, the Institute of Lassa Fever Research and Control (I.O., C. Iruolagbe, J.A., E.U., P.A., G.O., O.O., B.O., E.B.M., M.A., R.E., B.E., E.O.-E., G.A., S.O., P.O.O., C.T.H.) and the Department of Medicine (P.O.O.), Irrua Specialist Teaching Hospital, Irrua, the Nigeria Center for Disease Control, Abuja (C. Ihekweazu), and the Department of Medicine, Faculty of Clinical Sciences, Ambrose Alli University, Ekpoma (P.O.O.) - all in Nigeria; the Laboratory of Parasitology/Mycology HALD, Cheikh Anta Diop University of Dakar, Dakar, Senegal (A.G.); Kenema Government Hospital, Kenema, Sierra Leone (M.M.); Tulane Health Sciences Center, Tulane University, New Orleans (R.F.G.); the Departments of Immunology and Microbial Science and Integrative Structural and Computational Biology, Scripps Research Institute, and the Scripps Translational Science Institute, La Jolla, CA (K.G.A.); and Howard Hughes Medical Institute, Chevy Chase, MD (P.C.S.).

During 2018, an unusual increase in Lassa fever cases occurred in Nigeria, raising concern among national and international public health agencies. We analyzed 220 Lassa virus genomes from infected patients, including 129 from the 2017-2018 transmission season, to understand the viral populations underpinning the increase. A total of 14 initial genomes from 2018 samples were generated at Redeemer's University in Nigeria, and the findings were shared with the Nigerian Center for Disease Control in real time. We found that the increase in cases was not attributable to a particular Lassa virus strain or sustained by human-to-human transmission. Instead, the data were consistent with ongoing cross-species transmission from local rodent populations. Phylogenetic analysis also revealed extensive viral diversity that was structured according to geography, with major rivers appearing to act as barriers to migration of the rodent reservoir.
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http://dx.doi.org/10.1056/NEJMoa1804498DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6181183PMC
November 2018

Efficacy of Artemisinin-Based Combination Treatments of Uncomplicated Falciparum Malaria in Under-Five-Year-Old Nigerian Children Ten Years Following Adoption as First-Line Antimalarials.

Am J Trop Med Hyg 2018 09 21;99(3):649-664. Epub 2018 Jun 21.

Institute for Medical Research and Training, University of Ibadan, Ibadan, Nigeria.

The efficacies of 3-day regimens of artemether-lumefantrine (AL), artesunate-amodiaquine (AA), and dihydroartemisinin-piperaquine (DHP) were evaluated in 910 children < 5 years old with uncomplicated malaria from six geographical areas of Nigeria. Parasite positivity 1 day and Kaplan-Meier estimated risk of persistent parasitemia 3 days after therapy initiation were both significantly higher, and geometric mean parasite reduction ratio 1 day after treatment initiation (PRRD1) was significantly lower in AL-treated children than in AA- and DHP-treated children. No history of fever, temperature > 38°C, enrollment parasitemia > 75,000 μL, and PRRD1 < 5,000 independently predicted persistent parasitemia 1 day after treatment initiation. Parasite clearance was significantly faster and risk of reappearance of asexual parasitemia after initial clearance was significantly lower in DHP-treated children. Overall, day 42 polymerase chain reaction-corrected efficacy was 98.3% (95% confidence interval [CI]: 96.1-100) and was similar for all treatments. In a non-compartment model, declines of parasitemias were monoexponential with mean terminal elimination half-life of 1.3 hours and unimodal frequency distribution of half-lives. All treatments were well tolerated. In summary, all three treatments evaluated remain efficacious treatments of uncomplicated malaria in young Nigerian children, but DHP appears more efficacious than AL or AA.
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http://dx.doi.org/10.4269/ajtmh.18-0115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6169162PMC
September 2018

A modified anthrax toxin-based enzyme-linked immunospot assay reveals robust T cell responses in symptomatic and asymptomatic Ebola virus exposed individuals.

PLoS Negl Trop Dis 2018 05 24;12(5):e0006530. Epub 2018 May 24.

Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA, United States of America.

Background: Ebola virus (EBOV) caused more than 11,000 deaths during the 2013-2016 epidemic in West Africa without approved vaccines or immunotherapeutics. Despite its high lethality in some individuals, EBOV infection can produce little to no symptoms in others. A better understanding of the immune responses in individuals who experienced minimally symptomatic and asymptomatic infection could aid the development of more effective vaccines and antivirals against EBOV and related filoviruses.

Methodology/principle Findings: Between August and November 2017, blood samples were collected from 19 study participants in Lagos, Nigeria, including 3 Ebola virus disease (EVD) survivors, 10 individuals with documented close contact with symptomatic EVD patients, and 6 control healthcare workers for a cross-sectional serosurvey and T cell analysis. The Lagos samples, as well as archived serum collected from healthy individuals living in surrounding areas of the 1976 Democratic Republic of Congo (DRC) epidemic, were tested for EBOV IgG using commercial enzyme-linked immunosorbent assays (ELISAs) and Western blots. We detected antibodies in 3 out of 3 Lagos survivors and identified 2 seropositive individuals not known to have ever been infected. Of the DRC samples tested, we detected antibodies in 9 out of 71 (12.7%). To characterize the T cell responses in the Lagos samples, we developed an anthrax toxin-based enzyme-linked immunospot (ELISPOT) assay. The seropositive asymptomatic individuals had T cell responses against EBOV nucleoprotein, matrix protein, and glycoprotein 1 that were stronger in magnitude compared to the survivors.

Conclusion/significance: Our data provide further evidence of EBOV exposure in individuals without EVD-like illness and, for the first time, demonstrate that these individuals have T cell responses that are stronger in magnitude compared to severe cases. These findings suggest that T cell immunity may protect against severe EVD, which has important implications for vaccine development.
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http://dx.doi.org/10.1371/journal.pntd.0006530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5991402PMC
May 2018

Field validation of recombinant antigen immunoassays for diagnosis of Lassa fever.

Sci Rep 2018 04 12;8(1):5939. Epub 2018 Apr 12.

Department of Biological Sciences, College of Natural Sciences, Redeemer's University, Ede, Osun State, Nigeria.

Lassa fever, a hemorrhagic fever caused by Lassa virus (LASV), is endemic in West Africa. It is difficult to distinguish febrile illnesses that are common in West Africa from Lassa fever based solely on a patient's clinical presentation. The field performance of recombinant antigen-based Lassa fever immunoassays was compared to that of quantitative polymerase chain assays (qPCRs) using samples from subjects meeting the case definition of Lassa fever presenting to Kenema Government Hospital in Sierra Leone. The recombinant Lassa virus (ReLASV) enzyme-linked immunosorbant assay (ELISA) for detection of viral antigen in blood performed with 95% sensitivity and 97% specificity using a diagnostic standard that combined results of the immunoassays and qPCR. The ReLASV rapid diagnostic test (RDT), a lateral flow immunoassay based on paired monoclonal antibodies to the Josiah strain of LASV (lineage IV), performed with 90% sensitivity and 100% specificity. ReLASV immunoassays performed better than the most robust qPCR currently available, which had 82% sensitivity and 95% specificity. The performance characteristics of recombinant antigen-based Lassa virus immunoassays indicate that they can aid in the diagnosis of LASV Infection and inform the clinical management of Lassa fever patients.
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http://dx.doi.org/10.1038/s41598-018-24246-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5897328PMC
April 2018

Factors contributing to anaemia after uncomplicated falciparum malaria in under five year-old Nigerian children ten years following adoption of artemisinin-based combination therapies as first-line antimalarials.

BMC Infect Dis 2017 12 19;17(1):781. Epub 2017 Dec 19.

Antimalarial Therapeutic Efficacy Monitoring Group, National Malaria Elimination Programme, The Federal Ministry of Health, Abuja, Nigeria.

Background: Artemisinin-based combination therapies (ACTs) have remained efficacious treatments of acute falciparum malaria in many endemic areas but there is little evaluation of factors contributing to the anaemia of acute falciparum malaria following long term adoption of ACTs as first-line antimalarials in African children.

Methods: Malarious <5 year-olds randomized to artemether-lumefantrine, artesunate-amodiaquine or dihydroartemisinin-piperaquine treatments were followed up clinically for 6 weeks. Anaemia was defined as haematocrit <30%; Malaria-attributable fall in haematocrit (MAFH) as the difference between haematocrit 28-42 days post- and pre-treatment; Total MAFH (TMAFH) as the difference between days 28-42 haematocrit and the lowest haematocrit recorded in the first week post-treatment initiation; Drug-attributable fall in haematocrit (DAFH) as the difference between MAFH and TMAFH; Early appearing anaemia (EAA) as haematocrit <30% occurring within 1 week in children with normal haematocrit pre-treatment. Predictors of anaemia pre-treatment, EAA, MAFH or DAFH >4% were evaluated by stepwise multiple logistic regression models. Survival analysis and kinetics of DAFH were evaluated by Kaplan-Meier estimator and non-compartment model, respectively.

Results: Pre-treatment, 355 of 959 children were anaemic. Duration of illness >2 days and parasitaemia ≤10,000 μL were independent predictors of anaemia pre-treatment. EAA occurred in 301 of 604 children. Predictors of EAA were age ≤ 15 months, history of fever pre-treatment and enrolment haematocrit ≤35%. The probabilities of progression from normal haematocrit to EAA were similar for all treatments. MAFH >4% occurred in 446 of 694 children; its predictors were anaemia pre-treatment, enrolment parasitaemia ≤50,000 μL, parasitaemia one day post-treatment initiation and gametocytaemia. DAFH >4% occurred in 334 of 719 children; its predictors were history of fever pre-and fever 1 day post-treatment initiation, haematocrit ≥37%, and parasitaemia >100,000 μL. In 432 children, declines in DAFH deficits were monoexponential with overall estimated half-time of 2.2d (95% CI 1.9-2.6). Area under curve of deficits in DAFH versus time and estimated half-time were significantly higher in non-anaemic children indicating greater loss of haematocrit in these children.

Conclusion: After ten years of adoption of ACTs, anaemia is common pre-and early post-treatment, falls in haematocrit attributable to a single infection is high, and DAFH >4% is common and significantly lower in anaemic compared to non-anaemic Nigerian children.

Trial Registration: Pan African Clinical Trial Registry (PACTR) [ PACTR201709002064150, 1 March 2017 ].
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http://dx.doi.org/10.1186/s12879-017-2876-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5738206PMC
December 2017

Anaemia following Artemisinin-Based Combination Treatments of Uncomplicated Plasmodium falciparum Malaria in Children: Temporal Patterns of Haematocrit and the Use of Uncomplicated Hyperparasitaemia as a Model for Evaluating Late-Appearing Anaemia.

Chemotherapy 2017 28;62(4):231-238. Epub 2017 Apr 28.

Antimalarial Therapeutic Efficacy Monitoring Group, National Malaria Elimination Programme, The Federal Ministry of Health, Abuja, Nigeria.

Background: In severe malaria, intravenous artesunate may cause delayed haemolytic anaemia but there has been little evaluation of the propensity of oral artemisinin-based combination treatments (ACTs) to cause late-appearing anaemia.

Methods: The frequency of anaemia (haematocrit <30%), and temporal changes in haematocrit were evaluated in 1,191 malarious children following ACTs. "Haematocrit conservation" was evaluated by using the fall in haematocrit/1,000 asexual parasites cleared from the peripheral blood (FIH/1,000 asexual parasites cpb), and the ratio of the average haematocrit (on the first 3 days of starting treatment):total parasitaemia cleared.

Results: The frequency of anaemia decreased significantly following treatment. FIH/1,000 asexual parasites cpb, average haematocrit:total parasitaemia cleared, and mean haematocrit 5 weeks after treatment began were significantly lower in hyperparasitaemic children than in children without hyperparasitaemia, suggesting haematocrit conservation during treatment followed later by a loss of haematocrit. Asymptomatic late-appearing anaemia occurred in 6% of the children.

Conclusion: Artesunate-amodiaquine and artemether-lumefantrine contribute to haematocrit conservation at high parasitaemias but may cause late-appearing anaemia.
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http://dx.doi.org/10.1159/000449366DOI Listing
September 2017

Therapeutic efficacy and effects of artemisinin-based combination treatments on uncomplicated Plasmodium falciparum malaria -associated anaemia in Nigerian children during seven years of adoption as first-line treatments.

Infect Dis Poverty 2017 Feb 7;6(1):36. Epub 2017 Feb 7.

Department of Biological Sciences, Redeemer's University, Ede, Osun State, Nigeria.

Background: Artemisinin-based combination treatments (ACTs) are the first-line treatments of uncomplicated Plasmodium falciparum malaria in many endemic areas but there are few evaluation of their efficacy in anaemic malarious children.

Methods: Therapeutic efficacy of 3-day regimens of artesunate-amodiaquine and artemether-lumefantrine was evaluated in 437 anaemic and 909 non-anaemic malarious children following treatment during a seven-year period (2008-2014). Patterns of temporal changes in haematocrit were classified based on haematocrit values <30% and ≥30%. Kinetics of the disposition of the deficit in haematocrit from 30% following treatment were evaluated using a non-compartment model.

Results: PCR-corrected parasitological efficacy 28 days after start of treatment was significantly higher in artesunate-amodiaquine- compared to artemether-lumefantrine-treated children [97% (95%CI: 92.8-100) versus 96.4% (95%CI: 91.3-99.4), P = 0.02], but it was similar in non-anaemic and anaemic children. Fall in haematocrit/1 000 asexual parasites cleared from peripheral blood was significantly greater at lower compared to higher parasitaemias (P < 0.0001), and in non-anaemic compared to anaemic children (P = 0.007). In anaemic children at presentation, mean anaemia recovery time (AnRT) was 15.4 days (95%CI: 13.3-17.4) and it did not change over the years. Declines in haematocrit deficits from 30% were monoexponential with mean estimated half-time of 1.4 days (95%CI: 1.2-1.6). Anaemia half-time (t) correlated positively with AnRT in the same patients (r = 0.69, P < 0.0001). Bland-Altman analysis of 10 multiples of t and AnRT showed narrow limit of agreement with insignificant bias (P = 0.07) suggesting both can be used interchangeably in the same patients.

Conclusions: Artesunate-amodiaquine and artemether-lumefantrine remain efficacious treatments of uncomplicated P. falciparum infections in non-anaemic and anaemic Nigerian children in the last 7 years of adoption as first-line treatments. These ACTs may also conserve haematocrit at high parasitaemias and in anaemic children.

Trials Registration: Pan African Clinical Trial Registry PACTR201508001188143 , 3 July 2015; PACTR201510001189370 , 3 July 2015; PACTR201508001191898 , 7 July 2015 and PACTR201508001193368 , 8 July 2015.
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http://dx.doi.org/10.1186/s40249-016-0217-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5294876PMC
February 2017

Early rising asexual parasitaemia in Nigerian children following a first dose of artemisinin-based combination treatments of falciparum malaria.

BMC Infect Dis 2017 01 31;17(1):110. Epub 2017 Jan 31.

Department of Biological Sciences, Redeemer's University, Ede, Nigeria.

Background: Early rising asexual parasitaemia (ERAP), initially defined as 'an increase in the parasite count over the baseline pre-treatment level during the first 24 h of treatment' of falciparum malaria with artemisinin derivatives is well documented, but there is no characterization of its risk factors, kinetics, molecular features or relationship to late-appearing anaemia (LAA) in acute falciparum malaria in African children following oral artemisinin-based combination therapies (ACTs).

Methods: ERAP was defined as ≥5% increase in pre-treatment parasitaemia within 8 h of initiating treatment. Parasitaemia was quantified pre-treatment and 1-2 hourly for 8 h, and less frequently thereafter for 6 weeks following randomized treatment of acutely malarious children with artesunate-amodiaquine, artemether-lumefantrine or dihydroartemisinin-piperaquine. Risk factors were determined by stepwise multiple logistic regression model. Kinetics of release into and of elimination of asexual parasites and DNA clones from peripheral blood were evaluated by method of residuals and non-compartment model, respectively. Parasite population changes were evaluated morphologically and by molecular genotyping.

Results: ERAP occurred in 205 of 416 children. A parasitaemia <100,000/μL and parasitaemia 1 day post-treatment initiation were independent predictors of ERAP. In children with ERAP: mean and peak time of increase in parasitaemia were 105.6% (95% CI 81-130.1) and 2.5 h (95% CI 2.2-2.7), respectively. Mean lag time, half-time and rate constant of release were 0.2 h (95% CI 0.2-0.3), 1 h (95% CI 0.9-1.1), and 0.9 h (95% CI 0.8-1), respectively. Schizonts and young gametocytes were seen only in peripheral blood of few children with ERAP. In age-, gender-, baseline parasitaemia- and treatment-matched children with and without ERAP, parasite DNA clearance time and area under curve of number of DNA clones versus time were significantly higher in children with ERAP indicating peripheral retention of released parasites followed by elimination. DNA clone elimination was monoexponential.

Conclusion: ERAP is common, occurs rapidly as first order process and may be due to mobilization of parasites from deep tissue following a first dose of ACTs of acute childhood falciparum malaria.

Trials Registration: Pan African Clinical Trial Registry PACTR201508001188143 , 3 July 2015; PACTR201510001189370, 3 July 2015; PACTR201508001191898, 7 July 2015 and PACTR201508001193368, 8 July 2015.
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http://dx.doi.org/10.1186/s12879-016-2173-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5286790PMC
January 2017

Ebola Virus Epidemiology and Evolution in Nigeria.

J Infect Dis 2016 10 4;214(suppl 3):S102-S109. Epub 2016 Jul 4.

Department of Biological Sciences African Center of Excellence for Genomics of Infectious Diseases, Redeemer's University, Ede, Osun State.

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.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5050462PMC
http://dx.doi.org/10.1093/infdis/jiw190DOI Listing
October 2016

Clinical Sequencing Uncovers Origins and Evolution of Lassa Virus.

Cell 2015 Aug;162(4):738-50

FAS Center for Systems Biology, Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, MA 02138, USA; Broad Institute, Cambridge, MA 02142, USA.

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.
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http://dx.doi.org/10.1016/j.cell.2015.07.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4537774PMC
August 2015

A survey of malaria and some arboviral infections among suspected febrile patients visiting a health centre in Simawa, Ogun State, Nigeria.

J Infect Public Health 2016 Jan-Feb;9(1):52-9. Epub 2015 Aug 7.

Department of Biological Sciences, Redeemers University, Ede, Osun State, Nigeria.

Most febrile patients are often misdiagnosed with malaria due to similar symptoms, such as fever shared by malaria and certain arboviral infections. This study surveyed the incidence of malaria, chikungunya and dengue infections among a number of suspected febrile patients visiting Simawa Health Centre, Ogun State, Nigeria. Venous blood samples were obtained from 60 febrile patients (age 3-70 years) visiting the centre between April and May 2014. The rapid diagnostic test (RDT) was used to detect the presence of chikungunya (CHK) antibodies (IgM), dengue (DEN) virus and antibodies (NS1, IgM and IgG) and malaria parasites (Plasmodium falciparum and Plasmodium vivax). Malarial confirmatory tests were by microscopy and nested polymerase chain reaction (PCR) using the polymorphic region of Glutamate-Rich Protein (GLURP) gene. The complexity of P. falciparum infection in the community also determined by the use of nested PCR. These three mosquito-borne infections were observed in 63% (38) of the patients. The prevalence of CHK, DEN and malarial infections singularly were 11%, 0% and 63%, respectively, whereas malaria with either CHK or DEN infections were 24% (9) and 3% (1), respectively. No subjects were positive for CHK and DEN co-infection. Malarial microscopic confirmation was in 94% (32) of the malaria RDT-positive samples, 50% (17) were successfully analysed by nested PCR and the mean multiplicity of infection was 1.6 (1-3 clones). One patient sample harboured both P. falciparum and P. vivax. The study reports the presence of some arboviral infections having similar symptoms with malaria at Simawa, Ogun State. The proper diagnosis of infectious diseases is important for controlling them.
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http://dx.doi.org/10.1016/j.jiph.2015.06.009DOI Listing
October 2016

Discovery of novel rhabdoviruses in the blood of healthy individuals from West Africa.

PLoS Negl Trop Dis 2015 Mar 17;9(3):e0003631. Epub 2015 Mar 17.

Institute of Lassa Fever Research and Control, Irrua Specialist Teaching Hospital, Irrua, Edo State, Nigeria; Department of Biological Sciences, College of Natural Sciences, Redeemer's University, Redemption City, Ogun State, Nigeria.

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.
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http://dx.doi.org/10.1371/journal.pntd.0003631DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4363514PMC
March 2015

Empowering African genomics for infectious disease control.

Genome Biol 2014 Nov 7;15(11):515. Epub 2014 Nov 7.

At present, African scientists can only participate minimally in the genomics revolution that is transforming the understanding, surveillance and clinical treatment of infectious diseases. We discuss new initiatives to equip African scientists with knowledge of cutting-edge genomics tools, and build a sustainable critical mass of well-trained African infectious diseases genomics scientists.
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http://dx.doi.org/10.1186/s13059-014-0515-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4282014PMC
November 2014

Enhanced methods for unbiased deep sequencing of Lassa and Ebola RNA viruses from clinical and biological samples.

Genome Biol 2014 ;15(11):519

We have developed a robust RNA sequencing method for generating complete de novo assemblies with intra-host variant calls of Lassa and Ebola virus genomes in clinical and biological samples. Our method uses targeted RNase H-based digestion to remove contaminating poly(rA) carrier and ribosomal RNA. This depletion step improves both the quality of data and quantity of informative reads in unbiased total RNA sequencing libraries. We have also developed a hybrid-selection protocol to further enrich the viral content of sequencing libraries. These protocols have enabled rapid deep sequencing of both Lassa and Ebola virus and are broadly applicable to other viral genomics studies.
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http://dx.doi.org/10.1186/PREACCEPT-1698056557139770DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4262991PMC
July 2015

Lassa fever in post-conflict sierra leone.

PLoS Negl Trop Dis 2014 Mar 20;8(3):e2748. Epub 2014 Mar 20.

FAS Center for Systems Biology, Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, Massachusetts, United States of America; Broad Institute, Cambridge, Massachusetts, United States of America.

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.
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http://dx.doi.org/10.1371/journal.pntd.0002748DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961205PMC
March 2014

Molecular diagnostics for lassa fever at Irrua specialist teaching hospital, Nigeria: lessons learnt from two years of laboratory operation.

PLoS Negl Trop Dis 2012 27;6(9):e1839. Epub 2012 Sep 27.

Institute of Lassa Fever Research and Control, Irrua Specialist Teaching Hospital, Edo State, Nigeria.

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.
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http://dx.doi.org/10.1371/journal.pntd.0001839DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3459880PMC
January 2013

In vitro-reduced susceptibility to artemether in P. falciparum and its association with polymorphisms on transporter genes.

J Infect Dis 2012 Aug 21;206(3):324-32. Epub 2012 May 21.

Laboratory of Molecular Infectology and Parasitology, Clementino Fraga Filho University Hospital, Federal University of Rio de Janeiro, Brazil.

Plasmodium falciparum with reduced sensitivity to artemisinin derivatives has been observed in endemic areas, but the molecular mechanisms for this reduced sensitivity remain unclear. We evaluated the association between in vitro susceptibility of P. falciparum isolates obtained from southwest Nigeria and polymorphisms in selected putative transporter genes (PFE0775C, PF13_0271, pfmrp1, pfcrt, and pfmdr1). Modified schizont inhibition assay was used to determine the in vitro parasite susceptibility to artemether (ATH). Polymorphisms in selected genes were detected by polymerase chain reaction followed by direct DNA sequencing. The half-maximal inhibitory concentration (IC(50)) geometric mean (GM) for all P. falciparum isolates was 1.78 nM (range, 0.03-10.43 nM). Polymorphisms at codons 241, 86, and 76 of PFE0775C, pfmdr1, and pfcrt genes, respectively, were associated with reduced susceptibility to ATH. A new S263P single-nucleotide polymorphism on the PFE0775C gene was also detected in 27% of the isolates. Patient isolates harboring V241L or S263P polymorphisms on the PFE0775C gene showed increased IC(50) (GM: 3.08 nM and 1.79 nM, respectively). Plasmodium falciparum isolates harboring mutant Y86 pfmdr1 and P263 PFE0775C alleles showed a 2.5-5.5-fold increase in ATH IC(50.) This study shows that polymorphisms on the PFE0775C and pfmdr1 genes are associated with reduced sensitivity to ATH in fresh isolates of P. falciparum from Nigeria.
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http://dx.doi.org/10.1093/infdis/jis359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3490696PMC
August 2012