Surveillance of West Nile Virus in Tunisia: Evidence from Human and Entomological Investigation

Abstract

West Nile virus (WNV) is a mosquito-borne pathogen of the Flaviviridae family that poses recurring public health threats in Tunisia, where Culex pipiens is recognized as the primary vector. Identification of circulating strains in different mosquito species is essential for targeted prevention and control. Between November 2021 and October 2022, mosquitoes were collected at four high-risk sites, and human samples were obtained through the national meningitis surveillance program. Human serum, cerebrospinal fluid (CSF), and urine samples were tested for WNV-specific IgM and IgG antibodies using ELISA, and molecular diagnosis was performed using Real-time RT-PCR (RRT-PCR). Positive samples underwent sequencing for phylogenetic characterization. Serological investigation on human serum revealed the presence of IgM and/or IgG antibodies reactive to WNV antigens, which may indicate exposure to WNV or related flaviviruses. RNA of WNV was detected in 21 mosquito pools (10.19%) belonging to Culex pipiens, Cx. perexiguus, Aedes caspius, and Ae. detritus, as well as in three human cases. Phylogenetic analysis of positive human and mosquito samples showed that all detected WNV strains belonged to sublineage 1a. The concurrent detection of WNV in vectors and humans confirms active circulation in Tunisia and underscores the role of Culex spp. Mosquitoes in transmission. Sustained multidisciplinary surveillance integrating entomological and clinical data is critical for early detection, guiding control measures, and preventing future outbreaks in humans and animals.

Keywords: Aedes caspius; Culex perexiguus; Culex pipiens; Culex theileri; Tunisia; West Nile virus; sublineage 1a; vector surveillance.

Figure 1

Neuroinvasive outbreaks in humans in TUNISIA.

Figure 2

Geographical distribution of human and mosquito sampling sites in Tunisia, according to climatic and ecological conditions.

Figure 3

West Nile virus lineage 1 phylogenetic subtree: Phylogenetic tree was generated using MEGA v.X software. Tree topology was inferred using the Neighbor-Joining method with the Kimura 2-parameter model and 1000 bootstrap replicates. The Usutu virus (USUV, NC006551) was used as an outgroup. Human generated sequences are marked by a red diamond while the sequence originating from mosquito is marked in green. For clarity, only representative sequences of sub-lineage 1a are displayed. Phylogenetic branches of the Mediterranean subtype’s representative sequences are highlighted in red.