Re-emergence of Oropouche virus between 2023 and 2024 in Brazil: an observational epidemiological study

Abstract

Background: Oropouche virus is an arthropod-borne virus that has caused outbreaks of Oropouche fever in central and South America since the 1950s. This study investigates virological factors contributing to the re-emergence of Oropouche fever in Brazil between 2023 and 2024.

Methods: In this observational epidemiological study, we combined multiple data sources for Oropouche virus infections in Brazil and conducted in-vitro and in-vivo characterisation. We collected serum samples obtained in Manaus City, Amazonas state, Brazil, from patients with acute febrile illnesses aged 18 years or older who tested negative for malaria and samples from people with previous Oropouche virus infection from Coari municipality, Amazonas state, Brazil. Basic clinical and demographic data were collected from the Brazilian Laboratory Environment Management System. We calculated the incidence of Oropouche fever cases with data from the Brazilian Ministry of Health and the 2022 Brazilian population census and conducted age-sex analyses. We used reverse transcription quantitative PCR to test for Oropouche virus RNA in samples and subsequently performed sequencing and phylogenetic analysis of viral isolates. We compared the phenotype of the 2023-24 epidemic isolate (AM0088) with the historical prototype strain BeAn19991 through assessment of titre, plaque number, and plaque size. We used a plaque reduction neutralisation test (PRNT₅₀) to assess the susceptibility of the novel isolate and BeAn19991 isolate to antibody neutralisation, both in serum samples from people previously infected with Oropouche virus and in blood collected from mice that were inoculated with either of the strains.

Findings: 8639 (81·8%) of 10 557 laboratory-confirmed Oropouche fever cases from Jan 4, 2015, to Aug 10, 2024, occurred in 2024, which is 58·8 times the annual median of 147 cases (IQR 73-325). Oropouche virus infections were reported in all 27 federal units, with 8182 (77·5%) of 10 557 infections occurring in North Brazil. We detected Oropouche virus RNA in ten (11%) of 93 patients with acute febrile illness between Jan 1 and Feb 4, 2024, in Amazonas state. AM0088 had a significantly higher replication at 12 h and 24 h after infection in mammalian cells than the prototype strain. AM0088 had a more virulent phenotype than the prototype in mammalian cells, characterised by earlier plaque formation, between 27% and 65% increase in plaque number, and plaques between 2·4-times and 2·6-times larger. Furthermore, serum collected on May 2 and May 20, 2016, from individuals previously infected with Oropouche virus showed at least a 32-fold reduction in neutralising capacity (ie, median PRNT₅₀ titre of 640 [IQR 320-640] for BeAn19991 vs <20 [ie, below the limit of detection] for AM0088) against the reassortant strain compared with the prototype.

Interpretation: These findings provide a comprehensive assessment of Oropouche fever in Brazil and contribute to an improved understanding of the 2023-24 Oropouche virus re-emergence. Our exploratory in-vitro data suggest that the increased incidence might be related to a higher replication efficiency of a new Oropouche virus reassortant for which previous immunity shows lower neutralising capacity.

Funding: São Paulo Research Foundation, Burroughs Wellcome Fund, Wellcome Trust, US National Institutes of Health, and Brazilian National Council for Scientific and Technological Development.

Translation: For the Portuguese translation of the abstract see Supplementary Materials section.

Figure 1

Spatial–temporal dynamics of Oropouche fever in Brazil, 2015–24 (A) Incidence of laboratory-confirmed Oropouche fever cases per epidemiological week in all 27 federal units (ie, 26 Brazilian states and the federal district), from epidemiological week 1 of 2015 (Jan 4–10) to epidemiological week 32 of 2024 (Aug 4–10). The dashed line indicates the implementation of Oropouche virus testing in 2024 across all central public health laboratories in Brazil by the Brazilian Ministry of Health. (B) Maps were coloured according to the incidence of laboratory-confirmed Oropouche fever cases by federal unit between January, 2015, and December, 2023, and between January and August, 2024. (C) Oropouche fever incidence based on the age–sex distribution of cases from 2015 to 2024. The dashed line indicates the national-level cumulative incidence. AC=Acre. AL=Alagoas. AM=Amazonas. AP=Amapá. BA=Bahia. CE=Ceará. DF=Distrito Federal (Federal District). ES=Espírito Santo. GO=Goiás. MA=Maranhão. MG=Minas Gerais. MS=Mato Grosso do Sul. MT=Mato Grosso. PA=Pará. PB=Paraíba. PE=Pernambuco. PI=Piauí. PR=Paraná. RJ=Rio de Janeiro. RN=Rio Grande do Norte. RO=Rondônia. RR=Roraima. RS=Rio Grande do Sul. SC=Santa Catarina. SE=Sergipe. SP=São Paulo. TO=Tocantins.

Figure 2

Phylogenetic analysis of Oropouche virus Maximum likelihood phylogenetic tree of 476 representative Oropouche virus genomes, including two new genomes generated in this study from serum samples of patients with febrile illnesses in Manaus City. Phylogenetic trees are shown for the S segment, M segment, and L segment. Tips are coloured according to the location country of each sample. Phylogenies were midpoint rooted for clarity of presentation. Scale bar indicates the evolutionary distance of substitutions per nucleotide site. Bootstrap values based on 1000 replicates are shown on principal nodes. The GenBank accession numbers of sequences used in this figure are shown in appendix 2 (pp 17–28). Detailed information on the collapsed clade with Oropouche virus reassortant strains circulating in 2023 and 2024 is provided in appendix 2 (pp 5–7). IQTV=Iquitos virus. JATV=Jatobal virus. MMDV=Madre de Dios virus. PDEV=Perdões virus.

Figure 3

Characterisation in vitro of 2024 Oropouche virus reassortment strain (A) Viral replication properties of Oropouche virus strain AM0088 and Oropouche virus strain BeAn19991 in Vero CCL81, Huh7, and U-251 cell lines. The median is presented by the dot, and the upper and lower limits represent the 75th and 25th percentiles (ie, whiskers). (B) Number of plaques formed by Oropouche virus strains AM0088 and BeAn19991 after infection in Vero CCL81 cells (n=12 wells). The median is presented by the bar, and the upper and lower limits represent the 75th and 25th percentiles (ie, whiskers). (C) Size of plaques formed by Oropouche virus strains AM0088 and BeAn19991 after infection in Vero CCL81 cells (n≥118 plaques). The median is presented by the middle line, and the upper and lower limits represent the 75th and 25th percentiles (ie, whiskers). FFU=focus-forming units.

Figure 4

Neutralisation of Oropouche virus strains BeAn19991 and AM0088 by PRNT₅₀ For relative plaque formation, each data point represents the mean of all serum samples for each group at each dilution level (shown as log₂ serum dilution) and error bars represent SD. Dashed lines indicate the lower limit of detection (PRNT₅₀=20) of the PRNT₅₀ assay for samples with low or absent virus neutralisation capacity. The median titre is presented by the middle line, and the upper and lower limits represent the 75th and 25th percentiles (ie, whiskers). (A) Serum samples from individuals previously infected with Oropouche virus in Coari, Amazonas State, Brazil (n=22). (B) Serum samples were collected from C57BL/6 mice 28 days after infection with Oropouche virus strain AM0088 (n=7). (C) Serum samples were collected from C57BL/6 mice 28 days after infection with Oropouche virus strain BeAn19991 (n=5).