Metagenomic surveillance uncovers diverse and novel viral taxa in febrile patients from Nigeria.

Effective infectious disease surveillance in high-risk regions is critical for clinical care and pandemic preemption; however, few clinical diagnostics are available for the wide range of potential human pathogens. Here, we conduct unbiased metagenomic sequencing of 593 samples from febrile Nigerian patients collected in three settings: i) population-level surveillance of individuals presenting with symptoms consistent with Lassa Fever (LF); ii) real-time investigations of outbreaks with suspected infectious etiologies; and iii) undiagnosed clinically challenging cases. We identify 13 distinct viruses, including the second and third documented cases of human blood-associated dicistrovirus, and a highly divergent, unclassified dicistrovirus that we name human blood-associated dicistrovirus 2. We show that pegivirus C is a common co-infection in individuals with LF and is associated with lower Lassa viral loads and favorable outcomes. We help uncover the causes of three outbreaks as yellow fever virus, monkeypox virus, and a noninfectious cause, the latter ultimately determined to be pesticide poisoning. We demonstrate that a local, Nigerian-driven metagenomics response to complex public health scenarios generates accurate, real-time differential diagnoses, yielding insights that inform policy.

Prevalence and outcome of Lassa fever among hospitalized patients in Ebonyi State, Nigeria, 2018-2019.

Lassa fever (LF) is a viral hemorrhagic illness endemic in West Africa. Annually, about 300,000-500,000 people are being infected, with about 5000 deaths. Symptoms of LF include high grade fever, headache, malaise, abdominal pain, vomiting, diarrhea, or sore throat. Terminal features may include bleeding from all orifices (mouth, nose, ear, anus and vagina), facial and neck oedema or pleural effusion. People of all ages, gender, and occupations were included in this study. A total of 440 patients' samples and Bio data were used for this study. The samples were analyzed for Lassa fever virus RNA using Real Time Reverse Transcriptase Polymerase Chain Reaction. The data obtained were analyzed using SPSS 20.0 and version 7 of Epi-Info statistical software. Analysis of these samples showed LASV prevalence of 25.7%. Chi-square analysis (p ≤ 0.05) showed that LASV infection does not depend on age, gender, or occupation. Our research re-emphasized the fact that LASV is a serious cause of fatality in humans. Our data showed that among 327 negative patients, 19 died. On the contrary, 113 LASV confirmed positive cases had 42 deaths. This result is highly significant. More so, Lassa fever disease outcome was compared across gender. There was no significant difference between the two genders. Death or recovery from LF infection does not depend on sex. However, recovery from LF significantly depends on age of the patient. Fatal outcome is significantly higher among adults/elderly. We aim to raise awareness to the recurrence of LASV in Ebonyi State and urgent need for other medical interventions, including other therapeutic measures, and possible vaccine production, considering the impact of this virus.

Development of an RT-LAMP assay for the detection of Lassa viruses in southeast and south-central Nigeria.

Lassa virus (LASV) causes Lassa fever (LF), a viral hemorrhagic fever endemic in West Africa. LASV strains are clustered into six lineages according to their geographic location. To confirm a diagnosis of LF, a laboratory test is required. Here, a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay using a portable device for the detection of LASV in southeast and south-central Nigeria using three primer sets specific for strains clustered in lineage II was developed. The assay detected in vitro transcribed LASV RNAs within 23 min and was further evaluated for detection in 73 plasma collected from suspected LF patients admitted into two health settings in southern Nigeria. The clinical evaluation using the conventional RT-PCR as the reference test revealed a sensitivity of 50% in general with 100% for samples with a viral titer of 9500 genome equivalent copies (geq)/mL and higher. The detection limit was estimated to be 4214 geq/mL. The assay showed 98% specificity with no cross-reactivity to other viruses which cause similar symptoms. These results suggest that this RT-LAMP assay is a useful molecular diagnostic test for LF during the acute phase, contributing to early patient management, while using a convenient device for field deployment and in resource-poor settings.

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

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.