Susceptibility and transmissibility of SARS-CoV-2 variants in transgenic mice expressing the cat angiotensin-converting enzyme 2 (ACE-2) receptor.

The emergence of SARS-CoV-2 in 2019 and its rapid spread throughout the world has caused the largest pandemic of our modern era. The zoonotic origin of this pathogen highlights the importance of the One Health concept and the need for a coordinated response to this kind of threats. Since its emergence, the virus has caused >7 million deaths worldwide. However, the animal source for human outbreaks remains unknown. The ability of the virus to jump between hosts is facilitated by the presence of the virus receptor, the highly conserved angiotensin-converting enzyme 2 (ACE2), found in various mammals. Positivity for SARS-CoV-2 has been reported in various species, including domestic animals and livestock, but their potential role in bridging viral transmission to humans is still unknown. Additionally, the virus has evolved over the pandemic, resulting in variants with different impacts on human health. Therefore, suitable animal models are crucial to evaluate the susceptibility of different mammalian species to this pathogen and the adaptability of different variants. In this work, we established a transgenic mouse model that expresses the feline ACE2 protein receptor (cACE2) under the human cytokeratin 18 (K18) gene promoter's control, enabling high expression in epithelial cells, which the virus targets. Using this model, we assessed the susceptibility, pathogenicity, and transmission of SARS-CoV-2 variants. Our results show that the sole expression of the cACE2 receptor in these mice makes them susceptible to SARS-CoV-2 variants from the initial pandemic wave but does not enhance susceptibility to omicron variants. Furthermore, we demonstrated efficient contact transmission of SARS-CoV-2 between transgenic mice that express either the feline or the human ACE2 receptor.

Glycolytic shift during West Nile virus infection provides new therapeutic opportunities.

BACKGROUND: Viral rewiring of host bioenergetics and immunometabolism may provide novel targets for therapeutic interventions against viral infections. Here, we have explored the effect on bioenergetics during the infection with the mosquito-borne flavivirus West Nile virus (WNV), a medically relevant neurotropic pathogen causing outbreaks of meningitis and encephalitis worldwide. RESULTS: A systematic literature search and meta-analysis pointed to a misbalance of glucose homeostasis in the central nervous system of WNV patients. Real-time bioenergetic analyses confirmed upregulation of aerobic glycolysis and a reduction of mitochondrial oxidative phosphorylation during viral replication in cultured cells. Transcriptomics analyses in neural tissues from experimentally infected mice unveiled a glycolytic shift including the upregulation of hexokinases 2 and 3 (Hk2 and Hk3) and pyruvate dehydrogenase kinase 4 (Pdk4). Treatment of infected mice with the Hk inhibitor, 2-deoxy-D-glucose, or the Pdk4 inhibitor, dichloroacetate, alleviated WNV-induced neuroinflammation. CONCLUSIONS: These results highlight the importance of host energetic metabolism and specifically glycolysis in WNV infection in vivo. This study provides proof of concept for the druggability of the glycolytic pathway for the future development of therapies to combat WNV pathology.

Pharmacological Elevation of Cellular Dihydrosphingomyelin Provides a Novel Antiviral Strategy against West Nile Virus Infection.

The flavivirus life cycle is strictly dependent on cellular lipid metabolism. Polyphenols like gallic acid and its derivatives are promising lead compounds for new therapeutic agents as they can exert multiple pharmacological activities, including the alteration of lipid metabolism. The evaluation of our collection of polyphenols against West Nile virus (WNV), a representative medically relevant flavivirus, led to the identification of N,N'-(dodecane-1,12-diyl)bis(3,4,5-trihydroxybenzamide) and its 2,3,4-trihydroxybenzamide regioisomer as selective antivirals with low cytotoxicity and high antiviral activity (half-maximal effective concentrations [EC50s] of 2.2 and 0.24 µM, respectively, in Vero cells; EC50s of 2.2 and 1.9 µM, respectively, in SH-SY5Y cells). These polyphenols also inhibited the multiplication of other flaviviruses, namely, Usutu, dengue, and Zika viruses, exhibiting lower antiviral or negligible antiviral activity against other RNA viruses. The mechanism underlying their antiviral activity against WNV involved the alteration of sphingolipid metabolism. These compounds inhibited ceramide desaturase (Des1), promoting the accumulation of dihydrosphingomyelin (dhSM), a minor component of cellular sphingolipids with important roles in membrane properties. The addition of exogenous dhSM or Des1 blockage by using the reference inhibitor GT-11 {N-[(1R,2S)-2-hydroxy-1-hydroxymethyl-2-(2-tridecyl-1-cyclopropenyl)ethyl]octanamide} confirmed the involvement of this pathway in WNV infection. These results unveil the potential of novel antiviral strategies based on the modulation of the cellular levels of dhSM and Des1 activity for the control of flavivirus infection.

Nanobodies Protecting From Lethal SARS-CoV-2 Infection Target Receptor Binding Epitopes Preserved in Virus Variants Other Than Omicron.

The emergence of SARS-CoV-2 variants that escape from immune neutralization are challenging vaccines and antibodies developed to stop the COVID-19 pandemic. Thus, it is important to establish therapeutics directed toward multiple or specific SARS-CoV-2 variants. The envelope spike (S) glycoprotein of SARS-CoV-2 is the key target of neutralizing antibodies (Abs). We selected a panel of nine nanobodies (Nbs) from dromedary camels immunized with the receptor-binding domain (RBD) of the S, and engineered Nb fusions as humanized heavy chain Abs (hcAbs). Nbs and derived hcAbs bound with subnanomolar or picomolar affinities to the S and its RBD, and S-binding cross-competition clustered them in two different groups. Most of the hcAbs hindered RBD binding to its human ACE2 (hACE2) receptor, blocked cell entry of viruses pseudotyped with the S protein and neutralized SARS-CoV-2 infection in cell cultures. Four potent neutralizing hcAbs prevented the progression to lethal SARS-CoV-2 infection in hACE2-transgenic mice, demonstrating their therapeutic potential. Cryo-electron microscopy identified Nb binding epitopes in and out the receptor binding motif (RBM), and showed different ways to prevent virus binding to its cell entry receptor. The Nb binding modes were consistent with its recognition of SARS-CoV-2 RBD variants; mono and bispecific hcAbs efficiently bound all variants of concern except omicron, which emphasized the immune escape capacity of this latest variant.

Low Immune Cross-Reactivity between West Nile Virus and a Zika Virus Vaccine Based on Modified Vaccinia Virus Ankara.

Zika virus (ZIKV) is a mosquito-borne flavivirus whose infection in pregnant women is associated with a spectrum of birth defects, which are together referred as Congenital Zika Syndrome. In addition, ZIKV can also induce Guillain-Barré syndrome, which is an autoimmune disease with neurological symptoms. The recent description of the first local infections of ZIKV in the European continent together with the expansion of one of its potential vectors, the Asian tiger mosquito (Aedes albopictus), invite us to be prepared for future outbreaks of ZIKV in this geographical region. However, the antigenic similarities of ZIKV with other flaviviruses can lead to an immune cross-reactivity with other circulating flaviviruses inducing, in some cases, flavivirus-disease exacerbation by antibody-dependent enhancement (ADE) of infection, which is a major concern for ZIKV vaccine development. Until now, West Nile virus (WNV) is the main medically relevant flavivirus circulating in the Mediterranean Basin. Therefore, anticipating the potential scenario of emergency vaccination against ZIKV in areas of Europe where WNV is endemic, in this investigation, we have evaluated the cross-reactivity between WNV and our previously developed ZIKV vaccine candidate based on modified vaccinia virus Ankara (MVA) vector expressing ZIKV structural proteins (MVA-ZIKV). To this end, mice were first immunized with MVA-ZIKV, subsequently challenged with WNV, and then, the ZIKV- and WNV-specific immune responses and protection against WNV were evaluated. Our results indicate low cross-reactivity between the MVA-ZIKV vaccine candidate and WNV and absence of ADE, supporting the safety of this ZIKV vaccine candidate in areas where the circulation of WNV is endemic.

Previous Usutu Virus Exposure Partially Protects Magpies (Pica pica) against West Nile Virus Disease But Does Not Prevent Horizontal Transmission.

The mosquito-borne flaviviruses USUV and WNV are known to co-circulate in large parts of Europe. Both are a public health concern, and USUV has been the cause of epizootics in both wild and domestic birds, and neurological cases in humans in Europe. Here, we explore the susceptibility of magpies to experimental USUV infection, and how previous exposure to USUV would affect infection with WNV. None of the magpies exposed to USUV showed clinical signs, viremia, or detectable neutralizing antibodies. After challenge with a neurovirulent WNV strain, neither viremia, viral titer of WNV in vascular feathers, nor neutralizing antibody titers of previously USUV-exposed magpies differed significantly with respect to magpies that had not previously been exposed to USUV. However, 75% (6/8) of the USUV-exposed birds survived, while only 22.2% (2/9) of those not previously exposed to USUV survived. WNV antigen labeling by immunohistochemistry in tissues was less evident and more restricted in magpies exposed to USUV prior to challenge with WNV. Our data indicate that previous exposure to USUV partially protects magpies against a lethal challenge with WNV, while it does not prevent viremia and direct transmission, although the mechanism is unclear. These results are relevant for flavivirus ecology and contention.

Akt Kinase Intervenes in Flavivirus Replication by Interacting with Viral Protein NS5.

Arthropod-borne flaviviruses, such as Zika virus (ZIKV), Usutu virus (USUV), and West Nile virus (WNV), are a growing cause of human illness and death around the world. Presently, no licensed antivirals to control them are available and, therefore, search for broad-spectrum antivirals, including host-directed compounds, is essential. The PI3K/Akt pathway controls essential cellular functions involved in cell metabolism and proliferation. Moreover, Akt has been found to participate in modulating replication in different viruses including the flaviviruses. In this work we studied the interaction of flavivirus NS5 polymerases with the cellular kinase Akt. In vitro NS5 phosphorylation experiments with Akt showed that flavivirus NS5 polymerases are phosphorylated and co-immunoprecipitate by Akt. Polymerase activity assays of Ala- and Glu-generated mutants for the Akt-phosphorylated residues also indicate that Glu mutants of ZIKV and USUV NS5s present a reduced primer-extension activity that was not observed in WNV mutants. Furthermore, treatment with Akt inhibitors (MK-2206, honokiol and ipatasertib) reduced USUV and ZIKV titers in cell culture but, except for honokiol, not WNV. All these findings suggest an important role for Akt in flavivirus replication although with specific differences among viruses and encourage further investigations to examine the PI3K/Akt/mTOR pathway as an antiviral potential target.

Novel Nonnucleoside Inhibitors of Zika Virus Polymerase Identified through the Screening of an Open Library of Antikinetoplastid Compounds.

Zika virus (ZIKV) is a mosquito-borne pathogen responsible for neurological disorders (Guillain-Barré syndrome) and congenital malformations (microcephaly). Its ability to cause explosive epidemics, such as that of 2015 to 2016, urges the identification of effective antiviral drugs. Viral polymerase inhibitors constitute one of the most successful fields in antiviral research. Accordingly, the RNA-dependent RNA polymerase activity of flavivirus nonstructural protein 5 (NS5) provides a unique target for the development of direct antivirals with high specificity and low toxicity. Here, we describe the discovery and characterization of two novel nonnucleoside inhibitors of ZIKV polymerase. These inhibitors, TCMDC-143406 (compound 6) and TCMDC-143215 (compound 15) were identified through the screening of an open-resource library of antikinetoplastid compounds using a fluorescence-based polymerization assay based on ZIKV NS5. The two compounds inhibited ZIKV NS5 polymerase activity in vitro and ZIKV multiplication in cell culture (half-maximal effective concentrations [EC50] values of 0.5 and 2.6 µM for compounds 6 and 15, respectively). Both compounds also inhibited the replication of other pathogenic flaviviruses, namely, West Nile virus (WNV; EC50 values of 4.3 and 4.6 µM for compounds 6 and 15, respectively) and dengue virus 2 (DENV-2; EC50 values of 3.4 and 9.6 µM for compounds 6 and 15, respectively). Enzymatic assays confirmed that the polymerase inhibition was produced by a noncompetitive mechanism. Combinatorial assays revealed an antagonistic effect between both compounds, suggesting that they would bind to the same region of ZIKV polymerase. The nonnucleoside inhibitors of ZIKV polymerase here described could constitute promising lead compounds for the development of anti-ZIKV therapies and, eventually, broad-spectrum antiflavivirus drugs.

Pathogenicity and virulence of West Nile virus revisited eight decades after its first isolation.

West Nile virus (WNV) is a flavivirus which transmission cycle is maintained between mosquitoes and birds, although it occasionally causes sporadic outbreaks in horses and humans that can result in serious diseases and even death. Since its first isolation in Africa in 1937, WNV had been considered a neglected pathogen until its recent spread throughout Europe and the colonization of America, regions where it continues to cause outbreaks with severe neurological consequences in humans and horses. Although our knowledge about the characteristics and consequences of the virus has increased enormously lately, many questions remain to be resolved. Here, we thoroughly update our knowledge of different aspects of the WNV life cycle: virology and molecular classification, host cell interactions, transmission dynamics, host range, epidemiology and surveillance, immune response, clinical presentations, pathogenesis, diagnosis, prophylaxis (antivirals and vaccines), and prevention, and we highlight those aspects that are still unknown and that undoubtedly require further investigation.

Dengue Virus Strikes Back: Increased Future Risk of Severe Dengue Disease in Humans as a Result of Previous Exposure to Zika Virus.

Mosquito-borne flaviviruses include medically important pathogens that are responsible for a variety of human diseases, such as dengue, Zika congenital syndrome, and West Nile fever [...].

Current Progress of Avian Vaccines Against West Nile Virus.

Birds are the main natural host of West Nile virus (WNV), the worldwide most distributed mosquito-borne flavivirus, but humans and equids can also be sporadic hosts. Many avian species have been reported as susceptible to WNV, particularly corvids. In the case that clinical disease develops in birds, this is due to virus invasion of different organs: liver, spleen, kidney, heart, and mainly the central nervous system, which can lead to death 24-48 h later. Nowadays, vaccines have only been licensed for use in equids; thus, the availability of avian vaccines would benefit bird populations, both domestic and wild ones. Such vaccines could be used in endangered species housed in rehabilitation and wildlife reserves, and in animals located at zoos and other recreational installations, but also in farm birds, and in those that are grown for hunting and restocking activities. Even more, controlling WNV infection in birds can also be useful to prevent its spread and limit outbreaks. So far, different commercial and experimental vaccines (inactivated, attenuated, and recombinant viruses, and subunits and DNA-based candidates) have been evaluated, with various regimens, both in domestic and wild avian species. However, there are still disadvantages that must be overcome before avian vaccination can be implemented, such as its cost-effectiveness for domestic birds since in many species the pathogenicity is low or zero, or the viability of being able to achieve collective immunity in wild birds in freedom. Here, a comprehensive review of what has been done until now in the field of avian vaccines against WNV is presented and discussed.

Lipid Metabolism as a Source of Druggable Targets for Antiviral Discovery against Zika and Other Flaviviruses.

The Zika virus (ZIKV) is a mosquito-borne flavivirus that can lead to birth defects (microcephaly), ocular lesions and neurological disorders (Guillain-Barré syndrome). There is no licensed vaccine or antiviral treatment against ZIKV infection. The effort to understand the complex interactions of ZIKV with cellular networks contributes to the identification of novel host-directed antiviral (HDA) candidates. Among the cellular pathways involved in infection, lipid metabolism gains attention. In ZIKV-infected cells lipid metabolism attributed to intracellular membrane remodeling, virion morphogenesis, autophagy modulation, innate immunity and inflammation. The key roles played by the cellular structures associated with lipid metabolism, such as peroxisomes and lipid droplets, are starting to be deciphered. Consequently, there is a wide variety of lipid-related antiviral strategies that are currently under consideration, which include an inhibition of sterol regulatory element-binding proteins (SREBP), the activation of adenosine-monophosphate activated kinase (AMPK), an inhibition of acetyl-Coenzyme A carboxylase (ACC), interference with sphingolipid metabolism, blockage of intracellular cholesterol trafficking, or a treatment with cholesterol derivatives. Remarkably, most of the HDAs identified in these studies are also effective against flaviviruses other than ZIKV (West Nile virus and dengue virus), supporting their broad-spectrum effect. Considering that lipid metabolism is one of the main cellular pathways suitable for pharmacological intervention, the idea of repositioning drugs targeting lipid metabolism as antiviral candidates is gaining force.

A Recombinant Subviral Particle-Based Vaccine Protects Magpie (Pica pica) Against West Nile Virus Infection.

The mosquito-borne West Nile virus (WNV) is a highly neurovirulent Flavivirus currently representing an emergent zoonotic concern. WNV cycles in nature between mosquito vectors and birds that act as amplifier hosts and play an essential role in virus ecology, being, thus, WNV a threat to many species. Availability of an efficient avian vaccine would benefit certain avian populations, both birds grown for hunting and restocking activities, as well as endangered species in captive breeding projects, wildlife reservations, and recreation installations, and would be useful to prevent and contain outbreaks. Avian vaccination would be also of interest to limit WNV spillover to humans or horses from susceptible bird species that live in urbanized landscapes, like magpies. Herein, we have addressed the efficacy of a single dose of a WNV recombinant subviral particle (RSP) vaccine in susceptible magpie (Pica pica). The protective capacity of the RSP-based vaccine was demonstrated upon challenge of magpies with 5 × 103 plaque forming units of a neurovirulent WNV strain. A significant improvement in survival rates of immunized birds was recorded when compared to vehicle-inoculated animals (71.4 vs. 22.2%, respectively). Viremia, which is directly related to the capacity of a host to be competent for virus transmission, was reduced in vaccinated animals, as was the presence of infectious virus in feather follicles. Bird-to-bird transmission was recorded in three of six unchallenged (contact) magpies housed with non-vaccinated WNV-infected birds, but not in contact animals housed with vaccinated WNV-infected magpies. These results demonstrate the protective efficacy of the RSP-based vaccine in susceptible birds against WNV infection and its value in controlling the spread of the virus.

Targeting host metabolism by inhibition of acetyl-Coenzyme A carboxylase reduces flavivirus infection in mouse models.

Flaviviruses are (re)-emerging RNA viruses strictly dependent on lipid metabolism for infection. In the search for host targeting antivirals, we explored the effect of pharmacological modulation of fatty acid metabolism during flavivirus infection. Considering the central role of acetyl-Coenzyme A carboxylase (ACC) on fatty acid metabolism, we analyzed the effect of three small-molecule ACC inhibitors (PF-05175157, PF-05206574, and PF-06256254) on the infection of medically relevant flaviviruses, namely West Nile virus (WNV), dengue virus, and Zika virus. Treatment with these compounds inhibited the multiplication of the three viruses in cultured cells. PF-05175157 induced a reduction of the viral load in serum and kidney in WNV-infected mice, unveiling its therapeutic potential for the treatment of chronic kidney disease associated with persistent WNV infection. This study constitutes a proof of concept of the reliability of ACC inhibitors to become viable antiviral candidates. These results support the repositioning of metabolic inhibitors as broad-spectrum antivirals.

A Vaccine Based on a Modified Vaccinia Virus Ankara Vector Expressing Zika Virus Structural Proteins Controls Zika Virus Replication in Mice.

Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus that affects humans and can cause severe neurological complications, including Guillain-Barré syndrome and microcephaly. Since 2007 there have been three large outbreaks; the last and larger spread in the Americas in 2015. Actually, ZIKV is circulating in the Americas, Southeast Asia, and the Pacific Islands, and represents a potential pandemic threat. Given the rapid ZIKV dissemination and the severe neurological and teratogenic sequelae associated with ZIKV infection, the development of a safe and efficacious vaccine is critical. In this study, we have developed and characterized the immunogenicity and efficacy of a novel ZIKV vaccine based on the highly attenuated poxvirus vector modified vaccinia virus Ankara (MVA) expressing the ZIKV prM and E structural genes (termed MVA-ZIKV). MVA-ZIKV expressed efficiently the ZIKV structural proteins, assembled in virus-like particles (VLPs) and was genetically stable upon nine passages in cell culture. Immunization of mice with MVA-ZIKV elicited antibodies that were able to neutralize ZIKV and induced potent and polyfunctional ZIKV-specific CD8+ T cell responses that were mainly of an effector memory phenotype. Moreover, a single dose of MVA-ZIKV reduced significantly the viremia in susceptible immunocompromised mice challenged with live ZIKV. These findings support the use of MVA-ZIKV as a potential vaccine against ZIKV.

Direct Activation of Adenosine Monophosphate-Activated Protein Kinase (AMPK) by PF-06409577 Inhibits Flavivirus Infection through Modification of Host Cell Lipid Metabolism.

Mosquito-borne flaviviruses are a group of RNA viruses that constitute global threats for human and animal health. Replication of these pathogens is strictly dependent on cellular lipid metabolism. We have evaluated the effect of the pharmacological activation of AMP-activated protein kinase (AMPK), a master regulator of lipid metabolism, on the infection of three medically relevant flaviviruses, namely, West Nile virus (WNV), Zika virus (ZIKV), and dengue virus (DENV). WNV is responsible for recurrent outbreaks of meningitis and encephalitis, affecting humans and horses worldwide. ZIKV has caused a recent pandemic associated with birth defects (microcephaly), reproductive disorders, and severe neurological complications (Guillain-Barré syndrome). DENV is the etiological agent of the most prevalent mosquito-borne viral disease, which can induce a potentially lethal complication called severe dengue. Our results showed, for the first time, that activation of AMPK using the specific small molecule activator PF-06409577 reduced WNV, ZIKV, and DENV infection. This antiviral effect was associated with an impairment of viral replication due to the modulation of host cell lipid metabolism exerted by the compound. These results support that the pharmacological activation of AMPK, which currently constitutes an important pharmacological target for human diseases, could also provide a feasible approach for broad-spectrum host-directed antiviral discovery.

High susceptibility of magpie (Pica pica) to experimental infection with lineage 1 and 2 West Nile virus.

West Nile virus (WNV), a zoonotic pathogen naturally transmitted by mosquitoes whose natural hosts are birds, has spread worldwide during the last few decades. Resident birds play an important role in flavivirus epidemiology, since they can serve as reservoirs and facilitate overwintering of the virus. Herein, we report the first experimental infection of magpie (Pica pica) with two strains of West Nile virus, lineages 1 (NY-99) and 2 (SRB Novi-Sad/12), which are currently circulating in Europe. Magpies were highly susceptible to WNV infection, with similar low survival rates (30% and 42.8%) for both lineages. All infected magpies developed viremia detectable at 3 days post-infection with titers above those necessary for successful transmission of WNV to a mosquito. Neutralizing antibodies were detected at all time points analyzed (from 7 to 17 days post-infection). WNV genome was detected in the brains and hearts of all magpies that succumbed to the infection, and, in some of the surviving birds. WNV-RNA was amplified from swabs (oral and cloacal) at 3, 6 and 7 days post-infection and feather pulps, from 3 to 17 days post-infection, of infected animals. Even more, infectious virus was recovered from swabs up to 7 days post-infection and from feather pulps up to 10 days post infection. Sham-infected control animals were negative for viremia, viral RNA, and antibodies. These results suggest that the magpie, which is one of the most abundant corvid species in Europe, could represent a source of WNV transmission for birds and humans. Our observations shed light on the pathogenesis, transmission, and ecology of WNV and can benefit the implementation of surveillance and control programs.

Zika virus infection confers protection against West Nile virus challenge in mice.

Flaviviruses are RNA viruses that constitute a worrisome threat to global human and animal health. Zika virus (ZIKV), which was initially reported to cause a mild disease, recently spread in the Americas, infecting millions of people. During this recent epidemic, ZIKV infection has been linked to serious neurological diseases and birth defects, specifically Guillain-Barrè syndrome (GBS) and microcephaly. Because information about ZIKV immunity remains scarce, we assessed the humoral response of immunocompetent mice to infection with three viral strains of diverse geographical origin (Africa, Asia and America). No infected animals showed any sign of disease or died after infection. However, specific neutralizing antibodies were elicited in all infected mice. Considering the rapid expansion of ZIKV throughout the American continent and its co-circulation with other medically relevant flaviviruses, such as West Nile virus (WNV), the induction of protective immunity between ZIKV and WNV was analyzed. Remarkably, protection after challenge with WNV was observed in mice previously infected with ZIKV, as survival rates were significantly higher than in control mice. Moreover, previous ZIKV infection enhanced the humoral immune response against WNV. These findings may be relevant in geographical areas where both ZIKV and WNV co-circulate, as well as for the future development of broad-spectrum flavivirus vaccines.

Pathogen-specific B-cell receptors drive chronic lymphocytic leukemia by light-chain-dependent cross-reaction with autoantigens.

Several lines of evidence indirectly suggest that antigenic stimulation through the B-cell receptor (BCR) supports chronic lymphocytic leukemia (CLL) development. In addition to self-antigens, a number of microbial antigens have been proposed to contribute to the selection of the immunoglobulins expressed in CLL. How pathogen-specific BCRs drive CLL development remains, however, largely unexplored. Here, we utilized mouse models of CLL pathogenesis to equip B cells with virus-specific BCRs and study the effect of antigen recognition on leukemia growth. Our results show that BCR engagement is absolutely required for CLL development. Unexpectedly, however, neither acute nor chronic exposure to virus-derived antigens influenced leukemia progression. Rather, CLL clones preferentially selected light chains that, when paired with virus-specific heavy chains, conferred B cells the ability to recognize a broad range of autoantigens. Taken together, our results suggest that pathogens may drive CLL pathogenesis by selecting and expanding pathogen-specific B cells that cross-react with one or more self-antigens.