West Nile virus

1. Structure of West Nile Virus

WNV is a positive-sense, single-stranded (SS) RNA virus. It is an enveloped virus with icosahedral symmetry ^[3] comprising three structural proteins that form the capsid (C), premembrane (PrM/M), envelope (E), and seven nonstructural proteins including NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5 ^[2].

The WNV possesses an about 11kb long, single-stranded, positive sensed RNA genome, which is translated into a polyprotein that is subsequently cleaved into 10 functional proteins by both host and viral proteases. The structural proteins assemble virion particles and mediate host receptor binding and viral entry into host cells. Meanwhile, nonstructural proteins participate in viral genomic RNA replication and the regulation of host immune responses ^[4].

3. West Nile Virus Life Cycle

WNV's life cycle is maintained through its transmission between birds and mosquitoes. Birds can act as a reservoir and amplifying host of WNV. When a mosquito bites an infected bird, the virus enters the mosquito's bloodstream and eventually moves into its salivary glands. Once being bitten, the infected mosquito inoculates the virus into the host's skin.

WNV, akin to other flaviviruses, enters host cells through clathrin-mediated endocytosis with the assistance of various host factors. Subsequently, it releases its genome from the endosome into the cytoplasm following acidification-induced membrane fusion.

The viral RNA genome is transported through the cytosol to the endoplasmic reticulum (ER) using the host cellular machinery. In the ER, the viral positive-sense RNA is transcribed into negative-sense RNA for the replication of the progeny viral genome RNA. The synthesized viral RNA is translated into structural and non-structural proteins for the assembly of progeny viruses.

The structural proteins construct the nucleocapsid on ER membranes and then bud into the cytoplasm through the Golgi network. The progeny virus is transported to the cell surface in an exocytic vesicle and undergoes maturation upon cleavage of the prM by cellular enzymes, leading to the release of mature virions ^[7].

WNV can replicate in various cells including neutrophils, macrophages, and keratinocytes. Following spread, the virus undergoes an amplification phase, disseminating to visceral organs, and potentially progressing to the neuroinvasive phase, affecting the central nervous system (CNS).

Figure 1. West Nile virus transmission cycle

4. West Nile Virus Transmission

WNV is transmitted to humans via the bite of an infected mosquito. Direct human-to-human transmission initially was believed to be caused only by occupational exposure, such as in a laboratory setting ^[5], or conjunctival exposure to infected blood ^[6]. The US outbreak identified additional transmission methods:

• Blood transfusions
• Organ transplants
• Pregnancy: An infected mother can infect her fetus, but the risk is very low.
• Breast-feeding: There is a very small chance of passing on the virus through breast milk. So that the Centers for Disease Control and Prevention (CDC) advise mothers to continue breast-feeding.
• Percutaneous and aerosol infections: These two infection ways have been reported in laboratory personnel.

5. Protective Immune Response to West Nile Virus

The protective immune response to WNV involves a coordinated effort among the innate and adaptive immune systems as well as the complement system, working together to recognize and eliminate the virus.

5.1 Innate Immunity

Recognition of WNV is initiated by pattern recognition receptors (PRRs) on the surface of cells, including TLRs. WNV can be recognized by TLR3 due to its ssRNA making dsRNA during genomic replication ^[8]. Infected cells produce interferons (IFNs) such as IFN-α and IFN-β as part of the antiviral response. IFNs signal neighboring cells to enter an antiviral state, inhibiting viral replication ^[9]. NK cells play a role in the early control of viral infections. They can directly kill infected cells and contribute to the modulation of the adaptive immune response.

5.2 Complement

The complement system, comprising interconnected proteins, defends against WNV through various pathways. Activation, involving classical, lectin, and alternative pathways, leads to the formation of the membrane attack complex (MAC). Genetic deficiencies in complement components increase susceptibility to WNV infection. Mannose-binding lectin (MBL) recognizes N-linked glycans on the structural proteins of WNV, contributing to neutralization through a C3- and C4-dependent mechanism that involves both the canonical and bypass lectin activation pathways ^[10]. WNV NS1 protein attenuates complement activation, and C1q enhances the potency of antibodies against WNV by regulating the stoichiometric requirements for neutralization ^[11].

5.3 Adaptive Immunity

Adaptive immunity against WNV involves B-cell and T-cell responses. B cells are responsible for producing antibodies that specifically recognize and neutralize WNV. These antibodies can inhibit viral entry and promote the clearance of circulating virus particles. Antibodies and B cells play a crucial role in defending against WNV infection, as evidenced by increased serum and CNS viral burdens and susceptibility to lethal infection in mice lacking B cells and antibodies ^[12]. The humoral immune response, mediated by antibodies, contributes to the prevention of WNV spread within the host. During WMV infection, early anti-WNV IgM limits viremia and propagation into the central nervous system. T cells, including both CD4⁺ helper T cells and CD8⁺ cytotoxic T cells, play a critical role in the cellular immune response against WNV. Helper T cells coordinate the immune response by assisting B cells in antibody production and activating other immune cells. Cytotoxic T cells directly target and kill virus-infected cells, limiting viral replication.