On May 20, 2024, the NRL-IPK received samples from 89 patients with AFI of unknown etiology in Santiago de Cuba and Cienfuegos Provinces, Cuba, where an increase of similar cases had been reported, most negative for dengue IgM. Patients reported acute fever onset with headache and joint pain for 2–3 days and rapid recovery. Median patient age was 35 years (interquartile range [IQR] 19–50); 49.4% of patients were female and 51.6% male. Of the 89 serum samples received, 69 were from Santiago de Cuba (Boniato, n = 39; Songo la Maya, n = 30) and 20 from Cienfuegos (Abreu, n = 4; Cienfuegos, n = 11; and Rodas, n = 5). Urine (n = 6) and feces (n = 30) samples were also collected from the 89 patients.

We extracted RNA by using the commercial QIAamp Viral RNA Mini Kit (QIAGEN, https://www.qiagen.com) according to the manufacturer’s instructions. We processed extracted RNA for dengue, Zika and chikungunya viruses using by real-time PCR (7–9), and all serum samples were negative. Rapid test results for dengue nonstructural protein 1 (Bioline Dengue Duo; Abbott Laboratories, https://www.abbott.com) and chikungunya IgM (Kabla Diagnósticos, https://www.biodiagnosticos.com) were also negative. Fecal samples were negative for enterovirus (10).

We examined 89 serum and 6 urine samples for Oropouche and Mayaro viruses by using real-time PCR (11). All samples were negative for Mayaro virus, and 74 (83. 1%) serum samples (54 from Santiago de Cuba and 20 from Cienfuegos) and 5 (83.3%) urine samples were positive for OROV.

Figure 1

Figure 1. Molecular phylogenetic analysis of Oropouche viruses isolated in Cuba (red bracket) and reference sequences. The evolutionary history was inferred by using the maximum-likelihood method based on the Kimura 2-parameter model...

For the viral genetic characterization, we studied 9 serum samples (3 from Boniato, 3 from Songo La Maya, and 3 from Cienfuegos). We subjected extracted RNA to reverse transcription by PCR using the QIAGEN OneStep RT-PCR Kit system and primers NORO5 (5′-AAAGAGGATCCAATAATGTCAGAGTTCATTT-3′) and ORON3 (5′-AATTCGGAATTGGCATATAGTGGAATTCAC-3′) (12). We purified the 626-bp fragments obtained, which encode for the viral nucleocapsid (positions 85–718), by using the QIAquick PCR Purification Kit (QIAGEN), followed by sequencing with the Dye Terminator Cycle Sequencing Quick Start Kit (Analis, https://www.analis.com) and primers NORO5, ORON3, OROV-F (5′-TCCGGAGGCAGCATATGTG-3′), and OROV-R (5′-AAGTGCTCAATGCTGGTGTTGT-3′) (donated by the Pan American Health Organization). We purified the sequencing products and then separated the generated fragments on a CEQ 8800 Genetic Analysis System sequencer (Analis). We edited and assembled the electropherograms by using Sequencher version 4.10.1 software (Gene Codes Corporation, https://www.genecodes.com). As a reference, we used the complete sequence of the small segment of the Oropouche orthobunyavirus strain (GenBank accession no. OP244879.1, Oropouche orthobunyavirus isolate 0200178W, small segment, complete sequence). Phylogenetic analyses (MEGA version 6) (13) revealed that the isolated viruses clustered in a separate branch, closely related to those reported from Brazil in 2023 (Figure 1).

After we had identified OROV transmission, national surveillance for dengue and Oropouche fever was intensified (active AFI case search). In addition, vector control intervention (adulticide treatment, source reduction, environmental management, biolarvicide application on permanent breeding sites) was applied in the areas with confirmed transmission.

Figure 2

Figure 2. Confirmed Oropouche cases according to date of onset of signs/symptoms and provinces, Cuba, May 2024.

During May 28–June 3, 2024, we processed 31 serum samples collected from locations with increased AFI cases (San Nicolas de Bari in Mayabeque; Perico, Cienaga de Zapata, and Jovellanos in Matanzas; and Ranchuelo in Villa Clara). OROV infection was confirmed in 25 (80.6%) samples: 7 from Matanzas, 9 from Mayabeque, and 9 from Villa Clara Provinces (Table 1). Median patient age was 34.5 years (range 4–83 years), male:female ratio was 1.1. Signs and symptoms by order of frequency were fever, headache, general malaise, myalgia, and arthralgia (Table 2; Figure 2; Appendix Table). No serious or fatal cases were reported.

To investigate the vector involved with OROV transmission in Cuba, we collected 156 C. quinquefasciatus mosquitoes and 49 Ceratopogonidae spp. from suspected areas of transmission in 5 blocks of houses in Boniato. After potential vector identification, we grouped not visibly engorged female C. quinquefasciatus mosquitoes into 9 pools (10–20/pool) and Ceratopogonidae spp. specimens into 2 pools (7–19/pool). We extracted RNA and conducted real-time PCR. Positive samples were attributed to 5 (56%) pools of C. quinquefasciatus mosquitoes and 1 (50%) pool of Ceratopogonidae biting midges.

Among our study limitations was collection of signs/symptoms by using a standardized form for AFI or dengue cases; consequently, we might have missed less common presentations such as meningitis. In addition, the presence of viral RNA does not confirm the vector role in virus transmissibility. More work is needed to determine the primary vectors responsible for the current outbreak.

Oropouche fever is an emerging disease of concern in South and Central America. Transmission outside of the Amazon region is probably silent and not detected by public health surveillance systems. Our results confirm an outbreak of Oropouche fever in Cuba. Transmission was detected in semi-urban localities of 5 of 16 provinces located in the western, central, and eastern parts of the country. Dengue surveillance enabled us to identify cases with nondengue AFI. As of August 2024, OROV transmission had been confirmed in 7 provinces. The outbreak in Cuba alerts the Americas and the world of the need for integrated dynamic surveillance systems to detect the introduction and early transmission of OROV and consequently implement effective measures for its control.