Seroprevalence of West Nile Virus in Blood Donors in Mainland Portugal

The genus Orthoflavivirus includes several mosquito-borne pathogenic viruses, notably West Nile virus (WNV), which is endemic to the Mediterranean region. In Portugal, WNV circulation has been documented in equines, birds and mosquitoes, however human cases remain rare and no recent human seroprevalence studies have been conducted. This study aimed to estimate the national and regional seroprevalence of WNV among blood donors in mainland Portugal and explore associations with sociodemographic factors. A cross-sectional study conducted in 2022 included 3593 blood donors from across mainland Portugal. Serum samples were tested for WNV immunoglobulin G (IgG) by enzyme-linked immunosorbent assay (ELISA) and positive and borderline samples were confirmed using a virus neutralization test. Sociodemographic data were collected through a structured paper questionnaire. Statistical analyses, including multivariate logistic regression, identified factors associated with seropositivity. A total of 55 samples (1.5%) tested positive, and 21 samples (0.6%) were classified as borderline for WNV antibodies by ELISA. Of these, 47 were confirmed by viral neutralization, giving an estimated national seroprevalence of 1.4%. Significant regional variation was noted, with higher seroprevalence observed in the Beira Baixa, Grande Lisboa and Médio Tejo regions. Some seropositive individuals were identified in northern coastal regions such as Ave, Cávado and Área Metropolitana do Porto. In multivariate analysis, geographical area of residence was the only factor associated with seropositivity. This study highlights regions at potential higher risk for human WNV exposure, primarily in the southern half of Portugal. Continued and integrated surveillance is crucial to inform public health strategies to mitigate WNV transmission risks in these regions, as well as in other regions where WNV may be emerging as a relevant One Health concern. Implementing preventive measures for both animals and humans is critical to minimizing exposure and infection, and further confirmatory studies using virus neutralization tests will be important for refining these estimates.