# Characterization of Culex pipiens cell lines: virus infection and RNAi response

## TL;DR

This paper introduces new Culex pipiens cell lines that can be used to study arboviruses, which are transmitted by mosquitoes and pose a global health threat.

## Contribution

The study characterizes two new Culex pipiens cell lines for virus susceptibility and RNAi response, addressing a research gap in Culex-borne arbovirus studies.

## Key findings

- Both CPE/LULS50 and CPL/LULS56 cell lines support replication of arboviruses from Flaviviridae, Peribunyaviridae, and Togaviridae families.
- Small RNA profiling showed virus-specific siRNA production in both cell lines for all tested viruses.
- Virus-specific piRNA was detected only for Peribunyaviridae in the cell lines.

## Abstract

Arboviruses transmitted by mosquitoes pose a global health threat, causing diseases ranging from mild fevers to severe encephalitis and hemorrhagic fevers. Despite their growing impact, arbovirus research is hindered by biosafety constraints and the need of specialized BSL-3 insectariums. To circumvent these challenges, mosquito-derived cell lines have become indispensable tools for investigating virus-vector interactions. However, most available cell lines originate from Aedes and Anopheles spp., creating a critical research gap for other key vectors such as Culex spp. Although a few cell lines were previously established, they did not represent primary transmitters of West Nile virus (WNV) and other emerging arboviruses in Europe, such as Culex pipiens.

To address this gap, the current study aimed to characterize two recently established Culex pipiens cell lines: CPE/LULS50 (Culex pipiens pipiens & molestus) and CPL/LULS56 (Culex pipiens molestus) in more detail including testing their virus susceptibility, antiviral RNAi response, and possible presence of insect-specific viruses.

The replication of arboviruses from three clinically relevant families (Flaviviridae, Peribunyaviridae, and Togaviridae), as well as insect-specific viruses, was observed in both CPE/LULS50 and CPL/LULS56 cell lines. Furthermore, small RNA profiling revealed production of virus-specific small interfering RNA (siRNA) in both cell lines for all tested viruses. Interestingly, virus-specific PIWI-interacting RNA (piRNA) was only detected for the Peribunyaviridae.

The current study demonstrates that the CPE/LULS50 and CPL/LULS56 cell lines are suitable candidates to facilitate research into Culex-specific virus-vector interactions, ultimately contributing to mitigation of the impact of Culex-borne arboviruses on public health.

The online version contains supplementary material available at 10.1186/s13071-026-07248-w.