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Land cover is recognized as an important determinant of mosquito community assemblages and pathogen transmission, but few studies have investigated the role of land cover in populations of La Crosse virus (LACV) vectors. La Crosse virus is a zoonotic disease primarily maintained by native Aedes triseriatus, with accessory transmission by invasive vectors Ae. albopictus and Ae. japonicus. The objective of this study was to investigate the association between land cover composition and the egg abundances of LACV vectors in a city with endemic LACV transmission. Oviposition cups were deployed at sites ranging from recreational forests to urban areas. Mixed-effects regression models were used to test the relationships between land cover composition and species abundances. Sites characterized by large percentages of impervious and built surfaces were dominated by Ae. albopictus, but Ae. triseriatus was the most abundant species in forested areas. Aedes japonicus was rare at all sites, but its presence was associated with higher percentages of forested land. These results indicate that forested areas may allow Ae. triseriatus to maintain large populations near urban areas that are dominated by Ae. albopictus. Further research should explore the potential for these land cover-dependent population dynamics to influence LACV transmission cycles. [ABSTRACT FROM AUTHOR]

The large Bunyavirales order includes several families of viruses with a segmented ambisense (−) RNA genome and a cytoplasmic life cycle that starts by synthesizing viral mRNA. The initiation of transcription, which is common to all members, relies on an endonuclease activity that is responsible for cap-snatching. In La Crosse virus, an orthobunyavirus, it has previously been shown that the cap-snatching endonuclease resides in the N-terminal domain of the L protein. Orthobunyaviruses are transmitted by arthropods and cause diseases in cattle. However, California encephalitis virus, La Crosse virus and Jamestown Canyon virus are North American species that can cause encephalitis in humans. No vaccines or antiviral drugs are available. In this study, three known Influenza virus endonuclease inhibitors (DPBA, L-742,001 and baloxavir) were repurposed on the La Crosse virus endonuclease. Their inhibition was evaluated by fluorescence resonance energy transfer and their mode of binding was then assessed by differential scanning fluorimetry and microscale thermophoresis. Finally, two crystallographic structures were obtained in complex with L-742,001 and baloxavir, providing access to the structural determinants of inhibition and offering key information for the further development of Bunyavirales endonuclease inhibitors.

Background: La Crosse virus is an important endemic public health concern in the North Carolina Appalachian Mountains; however, human incidence is not commonly noted in this region on the South Carolina side of the mountain range border. No relevant studies have been performed in South Carolina evaluating mosquito vector populations for La Crosse virus (LACV) infection; thus, a pilot mosquito surveillance study was executed in summer 2020. Material and Methods: Vector surveillance occurred at five South Carolina state parks bordering neighboring state endemic counties from May to August 2020. Collections were approved by the state park authority, as noted in Permit No. N-8-20. Results: All three competent mosquito vectors were collected during the study duration; however, these vectors were collected in low abundance: Aedes triseriatus (4.5% of all collected mosquitos); Aedes albopictus (2.0%); Aedes japonicus (1.4%). Principal mosquito vector specimens, Ae. triseriatus, were sent to Centers for Disease Control and Prevention for testing of LACV by real-time reverse transcription PCR—all were negative. Discussion: While entomologic evidence suggests low transmission risk for this arbovirus in the South Carolina Appalachian Mountain region, further eco-epidemiologic investigations are warranted to understand this endemicity variance within a relatively small geographic area. [ABSTRACT FROM AUTHOR]

Mosquito-borne La Crosse virus (LACV; family: Peribunyaviridae) is the leading cause of pediatric arboviral encephalitis in the United States, with clinical cases generally centered in the Midwest and Appalachian regions. Incidence of LACV cases in Appalachian states has increased, such that the region currently represents the majority of reported LACV cases in the USA. The amount of reported LACV cases from Virginia, however, is minimal compared to neighboring states such as North Carolina, West Virginia, and Tennessee, and non-Appalachian regions of Virginia are understudied. Here we examine the hypothesis that LACV is circulating widely in Virginia, despite a low clinical case report rate, and that the virus is circulating in areas not associated with LACV disease. In this study, we screened local mammalian wildlife in northwestern counties of Virginia using passive surveillance via patients submitted to wildlife rehabilitation centers. Blood sera (527 samples; 9 species, 8 genera) collected between October 2019 and December 2022 were screened for neutralizing antibodies against LACV, indicating prior exposure to the virus. We found an overall LACV seroprevalence of 1.90% among all wild mammals examined and reveal evidence of LACV exposure in several wild species not generally associated with LACV, including eastern cottontails and red foxes, along with established reservoirs, eastern gray squirrels, although there was no serological evidence in chipmunks. These data indicate the circulation of LACV in Virginia outside of Appalachian counties, however, at a lower rate than reported for endemic areas within the state and in other states.

The first step of any immune response is the recognition of foreign molecular structures inside the host organism. An important molecule that is generally foreign to eukaryotic cells is long double-stranded RNA (dsRNA), which can be generated during virus replication. The mechanisms of sensing viral dsRNA are well-studied in mammalian systems but are only poorly understood in insects, including disease vectors such as Culex quinquefasciatus mosquitoes. These mosquitoes are vectors for important arboviruses, such as West Nile virus, and Culex speciesmosquitoes are distributed across the globe in many temperate and tropical regions. The major antiviral response triggered by dsRNA in mosquitoes is RNA interference - a sequence-specific response which targets complementary viral RNA for degradation. However, here, we aimed to identify whether sequence-independent dsRNA sensing, mimicked by poly(I:C), can elicit an antiviral response. We observed a significant reduction in replication of La Crosse virus (LACV) in Cx. quinquefasciatus mosquito cells following poly(I:C) priming. We identified a number of antimicrobial peptides and Toll receptors that were upregulated at the transcript level by poly(I:C) stimulation. Notably, Toll6was upregulated andwe determined that a knockdown of Toll6 expression resulted also in increased LACV replication. Future efforts require genetic tools to validate whether the observed Toll6 antiviral activity is indeed linked to dsRNA sensing. However, large-scale functional genomic and proteomic approaches are also required to determine which downstream responses are part of the poly(I:C) elicited antiviral response. [ABSTRACT FROM AUTHOR]

ABSTRACT California serogroup viruses (CSGVs) of medical importance in the United States include La Crosse virus, Jamestown Canyon virus (JCV), California encephalitis virus, and snowshoe hare virus. Current diagnosis of CSGVs relies heavily on serologic techniques for detecting immunoglobulin M (IgM), an indication of a recent CSGV infection. However, human-positive control sera reactive to viruses in the serogroup are scarce because detection of recent infections is rare. Here, we describe the development of new murine monoclonal antibodies (MAbs) reactive to CSGVs and the engineering of a human-murine chimeric antibody by combining the variable regions of the broadly CSGV cross-reactive murine MAb, 3-3B6/2-3B2 and the constant region of the human IgM. MAb 3-3B6/2-3B2 recognizes a tertiary epitope on the Gn/Gc heterodimer, and epitopes important in JCV neutralization were mapped to the Gc glycoprotein. This engineered human IgM constitutively expressed in a HEK-293 stable cell line can replace human-positive control sera in diagnostic serological techniques such as IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). Compared to the parent murine MAbs, the human-chimeric IgM antibody had identical serological activity to CSGVs in ELISA and demonstrated equivalent reactivity compared to human immune sera in the MAC-ELISA. Importance Orthobunyaviruses in the California serogroup cause severe neurological disease in children and adults. While these viruses are known to circulate widely in North America, their occurrence is rare. Serological testing for CSGVs is hindered by the limited availability and volumes of human-positive specimens needed as controls in serologic assays. Here, we described the development of a murine monoclonal antibody cross-reactive to CSGVs engineered to contain the variable regions of the murine antibody on the backbone of human IgM. The chimeric IgM produced from the stably expressing HEK293 cell line was evaluated for use as a surrogate human-positive control in a serologic diagnostic test.

Mosquito-borne La Crosse virus (LACV; family: Peribunyaviridae) is the leading cause of pediatric arboviral encephalitis in the United States, with clinical cases generally centered in the Midwest and Appalachian regions. Incidence of LACV cases in Appalachian states has increased, such that the region currently represents the majority of reported LACV cases in the USA. The amount of reported LACV cases from Virginia, however, is minimal compared to neighboring states such as North Carolina, West Virginia, and Tennessee, and non-Appalachian regions of Virginia are understudied. Here we examine the hypothesis that LACV is circulating widely in Virginia, despite a low clinical case report rate, and that the virus is circulating in areas not associated with LACV disease. In this study, we screened local mammalian wildlife in northwestern counties of Virginia using passive surveillance via patients submitted to wildlife rehabilitation centers. Blood sera (527 samples; 9 species, 8 genera) collected between October 2019 and December 2022 were screened for neutralizing antibodies against LACV, indicating prior exposure to the virus. We found an overall LACV seroprevalence of 1.90% among all wild mammals examined and reveal evidence of LACV exposure in several wild species not generally associated with LACV, including eastern cottontails and red foxes, along with established reservoirs, eastern gray squirrels, although there was no serological evidence in chipmunks. These data indicate the circulation of LACV in Virginia outside of Appalachian counties, however, at a lower rate than reported for endemic areas within the state and in other states. [ABSTRACT FROM AUTHOR]

Simple Summary: We investigated the vertical transmission potential of the Orthobunyavirus La Crosse virus (LACV) in the mosquitoes Ae. albopictus and Ae. aegypti. In both mosquito species, productive midgut infection with the virus was a prerequisite for the following ovary infection. However, the midgut infection levels varied between both species, with Ae. albopictus being more susceptible to LACV infection than Ae. aegypti. In both mosquito species, the vertical transmission rates of LACV from mothers to offspring were below those levels typically described for transovarial transmission (TOT). The LACV infection patterns in the ovarian tissue of Ae. aegypti suggested the transovum transmission of the virus. In several Ae. albopictus samples, LACV antigen was detected in follicular tissue or in a few developing oocytes, indicating that the TOT of LACV could be potentially occurring in this mosquito species. Thus, TOT is not a general feature of LACV infections in mosquitoes. La Crosse virus (LACV) is circulating in the midwestern and southeastern states of the United States and can cause human encephalitis. The main vector of the virus is the eastern tree-hole mosquito, Aedes triseriatus. Ae. albopictus has been also described as a natural LACV vector, while Ae. aegypti has been infected with the virus under laboratory conditions. Here, we compare the vertical transmission potential of LACV in Ae. albopictus and Ae. aegypti, with emphasis given to the ovarian infection patterns that the virus generates in both species. Both mosquito species received artificial bloodmeals containing LACV. At defined time points post-infection/bloodmeal, midguts, head tissue, and ovaries were analyzed for the presence of virus. Viral infection patterns in the ovaries were visualized via immunofluorescence confocal microscopy and immunohistopathology assays using an LACV-specific monoclonal antibody. In Ae. aegypti, LACV was confronted with midgut infection and escape barriers, which were much less pronounced in Ae. albopictus, resulting in a significantly higher prevalence of infection in the latter. Following the ingestion of a single virus-containing bloodmeal, no progeny larvae were found to be virus-infected. Regardless, females of both species showed the presence of LACV antigen in their ovariole sheaths. Furthermore, in a single Ae. albopictus female, viral antigen was associated with the nurse cells inside the primary follicles. Following the ingestion of a second non-infectious bloodmeal at 7- or 10-days post-ingestion of an LACV-containing bloodmeal, more progeny larvae of Ae. albopictus than of Ae. aegypti were virus-infected. LACV antigen was detected in the egg chambers and ovariole sheaths of both mosquito species. Traces of viral antigen were also detected in a few oocytes from Ae. albopictus. The low level of vertical transmission and the majority of the ovarian infection patterns suggested the transovum rather than transovarial transmission (TOT) of the virus in both vector species. However, based on the detection of LACV antigen in follicular tissue and oocytes, there was the potential for TOT among several Ae. albopictus females. Thus, TOT is not a general feature of LACV infection in mosquitoes. Instead, the TOT of LACV seems to be dependent on its particular interaction with the reproductive tissues of a female. [ABSTRACT FROM AUTHOR]

Background La Crosse virus (LACV) is the most common neuroinvasive arboviral infection in children in the United States. However, data regarding predictors of disease severity and neurologic outcome are limited. Additionally, long-term neurologic and neurobehavioral outcomes remain relatively sparse. Methods This was a single-center, retrospective cohort study, followed by recruitment for a cross-sectional analysis of long-term neurobehavioral outcomes, among children aged 0–18 years with proven or probable LACV neuroinvasive disease (LACV-ND) between January 2009 and December 2018. Case ascertainment was assured by International Classification of Diseases , Ninth and Tenth Revision , Clinical Modification codes cross-referenced with laboratory results detecting LACV. Demographics, diagnostics, radiographs, and outcomes were evaluated. Recruitment of patients with prior diagnosis of LACV-ND occurred from January 2020 to March 2020, with assessment performed by validated pediatric questionnaires. Results One-hundred fifty-two children (83 males; median age, 8 years [interquartile range, 5–11.5 years]) were diagnosed with proven (n = 61 [47%]) and probable (n = 91 [60%]) LACV-ND. Sixty-five patients (43%) had severe disease. Altered mental status (AMS) (odds ratio [OR], 6.36 [95% confidence interval {CI}, 2.03–19.95]; P =.0002) and seizures at presentation (OR, 10.31 [95% CI, 3.45–30.86]; P =.0001) were independent predictors of severe disease. Epileptiform discharges on electroencephalogram (EEG) were independently associated with epilepsy diagnosis at follow-up (OR, 13.45 [95% CI, 1.4–128.77]; P =.024). Fifty-four patients were recruited for long-term neurobehavioral follow-up, with frequent abnormal assessments identified (19%–54%) irrespective of disease severity. Conclusions Severe disease was observed frequently among children with LACV-ND. Seizures and AMS at presentation were independent predictors of severe disease. EEG may help determine long-term epilepsy risk. Long-term neurobehavioral issues are frequent and likely underrecognized among children with LACV-ND. [ABSTRACT FROM AUTHOR]

La Crosse Virus (LACV) encephalitis patients are at risk for long-term deficits in cognitive function due to neuronal apoptosis following virus infection. However, the specific etiology underlying neuronal damage remains elusive. In this study, we examined how differentiation and mitotic inhibition of neuroblastoma cells influence their susceptibility to LACV infection and cell death. Treatment of SH-SY5Y cells with retinoic acid induced a neuronal cell phenotype which was similarly susceptible to LACV infection as untreated cells but had significantly delayed virus-induced cell death. Protein and RNA transcript analysis showed that retinoic acid-treated cells had decreased oxidative stress responses to LACV infection compared to untreated cells. Modulation of oxidative stress in untreated cells with specific compounds also delayed cell death, without substantially impacting virus production. Thus, the oxidative stress response of neurons to virus infection may be a key component of neuronal susceptibility to virus-induced cell death., Importance: Encephalitic viruses like La Crosse Virus (LACV) infect and kill neurons. Disease onset and progression is rapid meaning the time frame to treat patients before significant and long-lasting damage occurs is limited. Examining how neurons, the primary cells infected by LACV in the brain, resist virus-induced cell death can provide avenues for determining which pathways to target for effective treatments. In the current study, we studied how changing neuroblastoma growth and metabolism with retinoic acid treatment impacted their susceptibility to LACV-induced cell death. We utilized this information to test compounds for preventing death in these cells., Competing Interests: The authors declare no conflict of interest.

La Crosse virus (LACV) is a primary cause of pediatric arboviral encephalitis in the United States, particularly affecting children aged 16 years or younger. This age-related susceptibility extends to murine models, where weanling mice (3 weeks old) succumb to LACV infection, while adults (≥6 weeks old) demonstrate resistance. Despite its clinical relevance, the host immune response to LACV is not fully understood. In this study, we investigated the roles of neutralizing antibodies (nAbs), cytokines, and chemokines in weanling and adult mice following infection with 5 × 10 5 plaque-forming units (PFU) of LACV. Weanling mice demonstrated early disease onset with elevated peripheral viremia, but passive transfer of adult serum, confirmed to have nAbs, to naïve weanlings prior to infection completely rescued them from death. Moreover, adult mice had increased Th1 cytokines, Th9/Th17/Th22/Treg cytokines, and many chemokines. In contrast, weanlings had higher Th2 cytokines, correlating with symptoms onset. Flow cytometry and intracellular cytokine staining further demonstrated that weanling mice produced higher levels of IL-4 by CD4 + and CD8 + T cells compared to adults, regardless of infection status. Conversely, LACV-infected adult mice had increased IFN-γ production by CD8 + T cells compared to uninfected controls. Finally, the adoptive transfer of splenocytes from immune adult mice to naïve weanlings delayed neurological symptoms and improved survival. In conclusion, this study links nAbs and cytokine and chemokine responses to protective immunity in adult mice, contrasting with the pathogenesis seen in weanlings. These findings underscore the importance of further research into innate and adaptive immune mechanisms during LACV infection.IMPORTANCELa Crosse virus (LACV) is a primary cause of pediatric encephalitis in the United States, with an impact on children aged 16 years or younger. This age-related susceptibility is recapitulated in mouse models, where young mice succumb to LACV-induced disease, while adults demonstrate resistance. Our understanding of host responses to LACV remains underexplored. This study sheds light on the dynamics of neutralizing antibodies (nAbs), cytokines, and chemokines following LACV infection in both adult and weanling mice. Our study reveals age-specific variations in viremia, neutralizing antibody titers, survivability, and levels of cytokines and chemokines. Adult mice exhibit significantly elevated levels of Th1 cytokines, contrasting with elevated levels of Th2 cytokines observed in weanling mice, often coinciding with the onset of symptoms. These data reveal age-specific dynamics in cytokines and chemokines associated with protective versus pathogenic immunity, emphasizing the need for further studies on innate and adaptive immunity.

Several methods of mosquito collection are used for the surveillance of the primary La Crosse virus (LACV) vectors, Aedes triseriatus (Say, 1823), Ae. albopictus (Skuse, 1895), and Ae. japonicus (Theobald, 1901). However, little is known about how the choice of collection method may confound inferences made from LACV vector surveillance data. Therefore, the objective of this study was to investigate potential biases in the surveillance of LACV vectors using the Biogents BG-Sentinel 2 (BGS), CDC-Light Trap (CDC-LT), Biogents Gravid Aedes Trap (BG-GAT), and standard oviposition cup (ovicup). The traps were deployed simultaneously at 10 sites in Knovxille, Tennessee, USA for 20 consecutive weeks. Surveillance results differed widely among the traps, demonstrating a strong potential for trap biases in LACV vector surveillance. The BGS and CDC-LT were effective for collecting Ae. albopictus but were not sensitive to the presence of Ae. triseriatus or Ae. japonicus. The ovicup was the best trap for detecting Ae. triseriatus, while the BG-GAT was the only trap that regularly collected Ae. japonicus. Surveillance conducted with the CDC-LT or BGS indicated that Ae. albopictus was dominant at all sites, but the ovicup and BG-GAT suggested a much larger relative abundance of Ae. triseriatus and Ae. japonicus, respectively. Aedes albopictus and Ae. triseriatus collected in the BG-GAT were significantly larger than those collected from the BGS and CDC-LT, indicating that the traps sampled different sub-populations. A multi-method surveillance approach is recommended to reduce potential biases when conducting surveillance of LACV vectors., (© The Author(s) 2024. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Purpose of Review: La Crosse virus encephalitis (LACV-E) is among the most common neuroinvasive arthropod-borne viral infections of childhood in North America. Children are uniquely predisposed to symptomatic disease, whereas symptomatic adult infections remain uncommon. Infection results frequently in neurologic symptomatology including headaches, seizures, and altered mentation, often necessitating hospitalization and significant diagnostic evaluation. The purpose of this review is to provide a contemporary assessment of clinical, laboratory, and neurobehavioral outcomes of children with LACV-E., Recent Findings: Common clinical manifestations at presentation, specifically seizure activity and altered mental status, are independent predictors of disease severity. Epileptiform discharges on electroencephalogram (EEG) during hospitalization may predict long-term epilepsy diagnosis. Lastly, long-term neurologic sequelae from acute infection is persistent and likely under-recognized among children with LACV-E., Summary: As climate change alters the geographic distribution of mosquito borne illnesses, a possibility of regional expansion of La Crosse virus (LACV) endemicity exists. The above data highlight readily identifiable features and testing modalities for clinicians who may encounter this viral infection. Importantly, an emphasis on long term neurobehavioral follow up is necessary to better identify and provide support of affected individuals. Future research in identifying host immune responses to LACV infection, as well as therapeutic options, are needed., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

La Crosse virus (LACV) is circulating in the midwestern and southeastern states of the United States and can cause human encephalitis. The main vector of the virus is the eastern tree-hole mosquito, Aedes triseriatus. Ae. albopictus has been also described as a natural LACV vector, while Ae. aegypti has been infected with the virus under laboratory conditions. Here, we compare the vertical transmission potential of LACV in Ae. albopictus and Ae. aegypti, with emphasis given to the ovarian infection patterns that the virus generates in both species. Both mosquito species received artificial bloodmeals containing LACV. At defined time points post-infection/bloodmeal, midguts, head tissue, and ovaries were analyzed for the presence of virus. Viral infection patterns in the ovaries were visualized via immunofluorescence confocal microscopy and immunohistopathology assays using an LACV-specific monoclonal antibody. In Ae. aegypti, LACV was confronted with midgut infection and escape barriers, which were much less pronounced in Ae. albopictus, resulting in a significantly higher prevalence of infection in the latter. Following the ingestion of a single virus-containing bloodmeal, no progeny larvae were found to be virus-infected. Regardless, females of both species showed the presence of LACV antigen in their ovariole sheaths. Furthermore, in a single Ae. albopictus female, viral antigen was associated with the nurse cells inside the primary follicles. Following the ingestion of a second non-infectious bloodmeal at 7- or 10-days post-ingestion of an LACV-containing bloodmeal, more progeny larvae of Ae. albopictus than of Ae. aegypti were virus-infected. LACV antigen was detected in the egg chambers and ovariole sheaths of both mosquito species. Traces of viral antigen were also detected in a few oocytes from Ae. albopictus. The low level of vertical transmission and the majority of the ovarian infection patterns suggested the transovum rather than transovarial transmission (TOT) of the virus in both vector species. However, based on the detection of LACV antigen in follicular tissue and oocytes, there was the potential for TOT among several Ae. albopictus females. Thus, TOT is not a general feature of LACV infection in mosquitoes. Instead, the TOT of LACV seems to be dependent on its particular interaction with the reproductive tissues of a female.

La Crosse virus (LACV) is a major cause of pediatric encephalitis and aseptic meningitis in the Midwestern, Mid-Atlantic, and Southern United States, where it is an emerging pathogen. The LACV Gc glycoprotein plays a critical role in the neuropathogenesis of LACV encephalitis as the putative virus attachment protein. Previously, we identified and experimentally confirmed the location of the LACV fusion peptide within Gc and generated a panel of recombinant LACVs (rLACVs) containing mutations in the fusion peptide as well as the wild-type sequence. These rLACVs retained their ability to cause neuronal death in a primary embryonic rat neuronal culture system, despite decreased replication and fusion phenotypes. To test the role of the fusion peptide in vivo, we tested rLACVs in an age-dependent murine model of LACV encephalitis. When inoculated directly into the CNS of young adult mice (P28), the rLACV fusion peptide mutants were as neurovirulent as the rLACV engineered with a wild-type sequence, confirming the results obtained in tissue culture. In contrast, the fusion peptide mutant rLACVs were less neuroinvasive when suckling (P3) or weanling (P21) mice were inoculated peripherally, demonstrating that the LACV fusion peptide is a determinant of neuroinvasion, but not of neurovirulence. In a challenge experiment, we found that peripheral challenge of weanling (P21) mice with fusion peptide mutant rLACVs protected from a subsequent WT-LACV challenge, suggesting that mutations in the fusion peptide are an attractive target for generating live-attenuated virus vaccines. Importantly, the high degree of conservation of the fusion peptide amongst the Bunyavirales and, structurally, other arboviruses suggests that these findings are broadly applicable to viruses that use a class II fusion mechanism and cause neurologic disease. [ABSTRACT FROM AUTHOR]

Abstract Background A key factor in the development of viral encephalitis is a virus crossing the blood-brain barrier (BBB). We have previously shown that age-related susceptibility of mice to the La Crosse virus (LACV), the leading cause of pediatric arbovirus encephalitis in the USA, was associated with the ability of the virus to cross the BBB. LACV infection in weanling mice (aged around 3 weeks) results in vascular leakage in the olfactory bulb/tract (OB/OT) region of the brain, which is not observed in adult mice aged > 6–8 weeks. Thus, we studied age-specific differences in the response of brain capillary endothelial cells (BCECs) to LACV infection. Methods To examine mechanisms of LACV-induced BBB breakdown and infection of the CNS, we analyzed BCECs directly isolated from weanling and adult mice as well as established a model where these cells were infected in vitro and cultured for a short period to determine susceptibility to virus infection and cell death. Additionally, we utilized correlative light electron microscopy (CLEM) to examine whether changes in cell morphology and function were also observed in BCECs in vivo. Results BCECs from weanling, but not adult mice, had detectable infection after several days in culture when taken ex vivo from infected mice suggesting that these cells could be infected in vitro. Further analysis of BCECs from uninfected mice, infected in vitro, showed that weanling BCECs were more susceptible to virus infection than adult BCECs, with higher levels of infected cells, released virus as well as cytopathic effects (CPE) and cell death. Although direct LACV infection is not detected in the weanling BCECs, CLEM analysis of brain tissue from weanling mice indicated that LACV infection induced significant cerebrovascular damage which allowed virus-sized particles to enter the brain parenchyma. Conclusions These findings indicate that BCECs isolated from adult and weanling mice have differential viral load, infectivity, and susceptibility to LACV. These age-related differences in susceptibility may strongly influence LACV-induced BBB leakage and neurovascular damage allowing virus invasion of the CNS and the development of neurological disease.

The neuroinvasive disease caused by Jamestown Canyon virus (JCV) infection is rare. However, increasing incidence and widespread occurrence of the infection make JCV a growing public health concern. Presently, clinical diagnosis is achieved through serological testing, and mosquito pool surveillance requires virus isolation and identification. A rapid molecular detection test, such as real-time RT-PCR, for diagnosis and surveillance of JCV has not been widely utilized. To enhance testing and surveillance, here, we describe the development and validation of a real-time RT-PCR test for the detection of JCV RNA. Three primer and probe sets were evaluated for analytical sensitivity and specificity. One probe set, JCV132FAM, was found to be the most sensitive test detecting 7.2 genomic equivalents/µL. While less sensitive, a second probe set JCV231cFAM was the most specific test with limited detection of Keystone virus at high RNA loads. Taken together, these data indicate both probe sets can be utilized for a primary sensitive screening assay and a secondary specific confirmatory assay. While both primer and probe sets detected high viral loads of Keystone virus, these assays did not detect any virus in the California encephalitis virus clade, including negative detection of the medically important La Crosse virus (LACV) and snowshoe hare virus (SSHV). The real-time RT-PCR assay described herein could be utilized in diagnosis and surveillance in regions with co-circulation of JCV and LACV or SSHV to inform public health action. [ABSTRACT FROM AUTHOR]

Abstract Background La Crosse virus (LACV) (genus Orthobunyavirus, family Peribunyaviridae) is a mosquito-borne virus that causes pediatric encephalitis and accounts for 50–150 human cases annually in the USA. Human cases occur primarily in the Midwest and Appalachian regions whereas documented human cases occur very rarely in the northeastern USA. Methods Following detection of a LACV isolate from a field-collected mosquito in Connecticut during 2005, we evaluated the prevalence of LACV infection in local mosquito populations and genetically characterized virus isolates to determine whether the virus is maintained focally in this region. Results During 2018, we detected LACV in multiple species of mosquitoes, including those not previously associated with the virus. We also evaluated the phylogenetic relationship of LACV strains isolated from 2005–2018 in Connecticut and found that they formed a genetically homogeneous clade that was most similar to strains from New York State. Conclusion Our analysis argues for local isolation and long-term persistence of a genetically distinct lineage of LACV within this region. We highlight the need to determine more about the phenotypic behavior of these isolates, and whether this virus lineage poses a threat to public health.

The combined region of eastern Tennessee and western North Carolina has a persistently high risk of pediatric La Crosse virus neuroinvasive disease (LACV-ND). To guide public health intervention in this region, the objectives of this retrospective ecological study were to investigate the geographic clustering and predictors of pediatric LACV-ND risk at the ZIP code tabulation area (ZCTA) level. Data on pediatric cases of LACV-ND reported between 2003 and 2020 were obtained from Tennessee Department of Health and North Carolina Department of Health and Human Services. Purely spatial and space-time scan statistics were used to identify ZCTA-level clusters of confirmed and probable pediatric LACV-ND cases from 2003-2020, and a combination of global and local (i.e., geographically weighted) negative binomial regression models were used to investigate potential predictors of disease risk from 2015-2020. The cluster investigation revealed spatially persistent high-risk and low-risk clusters of LACV-ND, with most cases consistently reported from a few high-risk clusters throughout the entire study period. Temperature and precipitation had positive but antagonistic associations with disease risk from 2015-2020, but the strength of those relationships varied substantially across the study area. Because LACV-ND risk clustering in this region is focally persistent, retroactive case surveillance can be used to guide the implementation of targeted public health intervention to reduce the disease burden in high-risk areas. Additional research on the role of climate in LACV transmission is warranted to support the development of predictive transmission models to guide proactive public health interventions., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Day et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: La Crosse virus neuroinvasive disease (LACVND) is the most common cause of arboviral encephalitis in children within the United States; in North Carolina, it is the most prevalent endemic mosquito-borne disease in humans., Methods: Here we report a surveillance summary of confirmed and probable LACVND during 2000-2020 using North Carolina Electronic Disease Surveillance System data, and we describe associated demographic characteristics, spatiotemporal distribution, clinical features, and mortality rates., Results: A total of 355 cases (74.9% confirmed) were reported from 41 North Carolina counties; most cases (92%) occurred in 19 Western North Carolina counties. An average of 17 cases were reported annually with the majority (94%) of cases occurring between mid-June and early October (epiweeks 25-41). The median case age was nine years (range: 1-95 years), 79% were aged ≤ 18 years, and 56.6% were male. Cases commonly presented with headache (95.4%), fever (95.1%), and altered mental status (80.8%); encephalitis (82.0%) and meningitis (45.9%) were frequently diagnosed. Encephalitis was more common in children (87%) than adults (62%) ( P < .001). Similarly, seizures were more common in children (54%) than adults (27%) ( P < .01). The case fatality rate was 1.4%; however, differences in age-specific rates were observed., Limitations: Changes in case definitions and reporting requirements, missing data, different reporting sources, and the retrospective nature of this study are all important limitations of our study., Conclusion: Given the persistent endemicity of La Crosse virus in Western North Carolina, clinicians and public health providers should consider La Crosse virus disease in all individuals, especially children, with compatible symptoms and a travel history to endemic counties. Prevention measures and educational outreach/public health messaging should focus on caregivers and children in Western North Carolina counties and during times when transmission risk is higher (i.e., summer and early fall)., Competing Interests: Acknowledgments: The authors would like to thank Mary Nordgulen (Western Carolina University Mosquito and Vector-borne Disease Laboratory) for her assistance with Figure 1. Figure 1 was created using Biorender.com (Agreement Number: OC260BN5WH). Funding: This study was supported, in part, by the North Carolina Department of Health and Human Services, Communicable Disease Branch (Contract # 43152) as an Epidemiology and Laboratory Capacity for Prevention and Control of Emerging Infectious Diseases (NU50CK000530) subcontract (Centers for Disease Control and Prevention). Disclosure of interests: The authors have no relevant conflicts of interest relating to this study., (Copyright ©2024 by the North Carolina Institute of Medicine and The Duke Endowment. All rights reserved.)

Background: A key factor in the development of viral encephalitis is a virus crossing the blood-brain barrier (BBB). We have previously shown that age-related susceptibility of mice to the La Crosse virus (LACV), the leading cause of pediatric arbovirus encephalitis in the USA, was associated with the ability of the virus to cross the BBB. LACV infection in weanling mice (aged around 3 weeks) results in vascular leakage in the olfactory bulb/tract (OB/OT) region of the brain, which is not observed in adult mice aged > 6-8 weeks. Thus, we studied age-specific differences in the response of brain capillary endothelial cells (BCECs) to LACV infection.Methods: To examine mechanisms of LACV-induced BBB breakdown and infection of the CNS, we analyzed BCECs directly isolated from weanling and adult mice as well as established a model where these cells were infected in vitro and cultured for a short period to determine susceptibility to virus infection and cell death. Additionally, we utilized correlative light electron microscopy (CLEM) to examine whether changes in cell morphology and function were also observed in BCECs in vivo.Results: BCECs from weanling, but not adult mice, had detectable infection after several days in culture when taken ex vivo from infected mice suggesting that these cells could be infected in vitro. Further analysis of BCECs from uninfected mice, infected in vitro, showed that weanling BCECs were more susceptible to virus infection than adult BCECs, with higher levels of infected cells, released virus as well as cytopathic effects (CPE) and cell death. Although direct LACV infection is not detected in the weanling BCECs, CLEM analysis of brain tissue from weanling mice indicated that LACV infection induced significant cerebrovascular damage which allowed virus-sized particles to enter the brain parenchyma.Conclusions: These findings indicate that BCECs isolated from adult and weanling mice have differential viral load, infectivity, and susceptibility to LACV. These age-related differences in susceptibility may strongly influence LACV-induced BBB leakage and neurovascular damage allowing virus invasion of the CNS and the development of neurological disease. [ABSTRACT FROM AUTHOR]

Anthropogenic land-use change may affect the transmission risk for endemic vector-borne diseases such as La Crosse encephalitis. In this study, we applied a comparative ecological approach to evaluate differences in vector species abundance, gonotrophic status, and environmental variables among six residential habitats (historical case houses) and six paired adjacent forest patches in a La Crosse virus endemic area of North Carolina. A total of 93,158 container Aedes spp. eggs were obtained by ovistrips and 1,040 resting mosquito adults were collected by large-bore aspiration from 10 June through 8 October, 2010. At sites characterized by high densities of artificial containers, the totals of eggs and adult mosquitoes were higher in the peridomestic plots. At sites characterized by lower densities of artificial containers, the totals of eggs and adult mosquitoes were higher in the forested plots. Although Aedes triseriatus, the primary La Crosse virus vector, was more commonly found in forested habitat overall, in sites characterized by high density of artificial containers, it was found in higher abundance in the peridomestic habitat. Similarly, the proportion of gonotrophically active (parous and gravid) mosquitoes was higher in the peridomestic habitat for sites with highartificial container density. Our study suggests that La Crosse virus transmission risk may be higher at peridomestic habitats with higher densities of artificial containers and thus reinforces the importance of public health measures to improve source reduction efforts. [ABSTRACT FROM AUTHOR]

Abstract Background La Crosse virus (LACV) infection has been shown to manipulate the blood-feeding behaviors of its main vector, Aedes triseriatus. Here, we investigated the effects of virus infection on serotonin and dopamine and their potential roles in host-seeking. In mosquitoes, serotonin depletion has been shown to interfere with blood-feeding but not host-seeking. Dopamine depletion does not affect either blood-feeding or host-seeking; elevations of dopamine, however, has been shown to inhibit host-seeking. The purpose of this study was to determine the effects of LACV infection on the host-seeking behavior of and neurotransmitter levels in Ae. triseriatus. Methods Host-seeking behavior was evaluated using a uni-port olfactometer and a membrane feeder assay. Levels of serotonin and dopamine in infected and control mosquito heads were measured using HPLC-ED. Results Infection with LACV significantly inhibited the activation and attraction of Ae. triseriatus females to a host. A higher proportion of uninfected Ae. triseriatus females were activated by the presence of a host compared to infected mosquitoes and more uninfected mosquitoes were full responders (95.7%) compared to infected ones (91.1%). However, infection with LACV did not significantly affect the landing, probing, or blood-feeding rates of female mosquitoes. LACV-infected mosquitoes had lower serotonin levels than controls (104.5 vs 138.3 pg/head) while the dopamine levels were not affected by infection status (282.3 vs 237 pg/head). Conclusions Our work suggests that virus-induced reduction of serotonin is related to previously reported blood-feeding alterations in LACV-infected mosquitoes and could lead to enhanced transmission and increased vectorial capacity. In addition, some aspects of host-seeking were inhibited by virus infection.

La Crosse Virus (LACV) is an arbovirus found in Eastern Appalachia and can cause pediatric encephalitis in prepubescent children. To assess the risk and transmission of this disease, it is particularly important to understand the average population of Aedes mosquitoes, which are the vectors of this virus. We use a deterministic compartmental model to study the effects of environmental factors on the population dynamics of Aedes mosquitoes in the Knox County area. We use locally-collected mosquito population data to adjust our model outputs and find that model transitions are heavily dependent on the fluctuations of both temperature and accumulated precipitation. These findings should be considered for mosquito management in Southern Appalachia, as well as in other regions with slight modifications to our model.

La Crosse virus (LACV) is a major cause of pediatric encephalitis and aseptic meningitis in the Midwestern, Mid-Atlantic, and Southern United States, where it is an emerging pathogen. The LACV Gc glycoprotein plays a critical role in the neuropathogenesis of LACV encephalitis as the putative virus attachment protein. Previously, we identified and experimentally confirmed the location of the LACV fusion peptide within Gc and generated a panel of recombinant LACVs (rLACVs) containing mutations in the fusion peptide as well as the wild-type sequence. These rLACVs retained their ability to cause neuronal death in a primary embryonic rat neuronal culture system, despite decreased replication and fusion phenotypes. To test the role of the fusion peptide in vivo, we tested rLACVs in an age-dependent murine model of LACV encephalitis. When inoculated directly into the CNS of young adult mice (P28), the rLACV fusion peptide mutants were as neurovirulent as the rLACV engineered with a wild-type sequence, confirming the results obtained in tissue culture. In contrast, the fusion peptide mutant rLACVs were less neuroinvasive when suckling (P3) or weanling (P21) mice were inoculated peripherally, demonstrating that the LACV fusion peptide is a determinant of neuroinvasion, but not of neurovirulence. In a challenge experiment, we found that peripheral challenge of weanling (P21) mice with fusion peptide mutant rLACVs protected from a subsequent WT-LACV challenge, suggesting that mutations in the fusion peptide are an attractive target for generating live-attenuated virus vaccines. Importantly, the high degree of conservation of the fusion peptide amongst the Bunyavirales and, structurally, other arboviruses suggests that these findings are broadly applicable to viruses that use a class II fusion mechanism and cause neurologic disease.

The neuroinvasive disease caused by Jamestown Canyon virus (JCV) infection is rare. However, increasing incidence and widespread occurrence of the infection make JCV a growing public health concern. Presently, clinical diagnosis is achieved through serological testing, and mosquito pool surveillance requires virus isolation and identification. A rapid molecular detection test, such as real-time RT-PCR, for diagnosis and surveillance of JCV has not been widely utilized. To enhance testing and surveillance, here, we describe the development and validation of a real-time RT-PCR test for the detection of JCV RNA. Three primer and probe sets were evaluated for analytical sensitivity and specificity. One probe set, JCV132FAM, was found to be the most sensitive test detecting 7.2 genomic equivalents/µL. While less sensitive, a second probe set JCV231cFAM was the most specific test with limited detection of Keystone virus at high RNA loads. Taken together, these data indicate both probe sets can be utilized for a primary sensitive screening assay and a secondary specific confirmatory assay. While both primer and probe sets detected high viral loads of Keystone virus, these assays did not detect any virus in the California encephalitis virus clade, including negative detection of the medically important La Crosse virus (LACV) and snowshoe hare virus (SSHV). The real-time RT-PCR assay described herein could be utilized in diagnosis and surveillance in regions with co-circulation of JCV and LACV or SSHV to inform public health action.

The large Bunyavirales order includes several families of viruses with a segmented ambisense (-) RNA genome and a cytoplasmic life cycle that starts by synthesizing viral mRNA. The initiation of transcription, which is common to all members, relies on an endonuclease activity that is responsible for cap-snatching. In La Crosse virus, an orthobunyavirus, it has previously been shown that the cap-snatching endonuclease resides in the N-terminal domain of the L protein. Orthobunyaviruses are transmitted by arthropods and cause diseases in cattle. However, California encephalitis virus, La Crosse virus and Jamestown Canyon virus are North American species that can cause encephalitis in humans. No vaccines or antiviral drugs are available. In this study, three known Influenza virus endonuclease inhibitors (DPBA, L-742,001 and baloxavir) were repurposed on the La Crosse virus endonuclease. Their inhibition was evaluated by fluorescence resonance energy transfer and their mode of binding was then assessed by differential scanning fluorimetry and microscale thermophoresis. Finally, two crystallographic structures were obtained in complex with L-742,001 and baloxavir, providing access to the structural determinants of inhibition and offering key information for the further development of Bunyavirales endonuclease inhibitors., (open access.)

Abstract Background La Crosse virus (LACV) causes pediatric encephalitis in the USA. LACV induces severe inflammation in the central nervous system, but the recruitment of inflammatory cells is poorly understood. A deeper understanding of LACV-induced neural pathology is needed in order to develop treatment options. However, there is a severe limitation of relevant human neuronal cell models of LACV infection. Methods We utilized human neural stem cell (hNSC)-derived neuron/astrocyte co-cultures to study LACV infection in disease-relevant primary cells. hNSCs were differentiated into neurons and astrocytes and infected with LACV. To characterize susceptibility and responses to infection, we measured viral titers and levels of viral RNA, performed immunofluorescence analysis to determine the cell types infected, performed apoptosis and cytotoxicity assays, and evaluated cellular responses to infection using qRT-PCR and Bioplex assays. Results hNSC-derived neuron/astrocyte co-cultures were susceptible to LACV infection and displayed apoptotic responses as reported in previous in vitro and in vivo studies. Neurons and astrocytes are both targets of LACV infection, with neurons becoming the predominant target later in infection possibly due to astrocytic responses to IFN. Additionally, neuron/astrocyte co-cultures responded to LACV infection with strong proinflammatory cytokine, chemokine, as well as MMP-2, MMP-7, and TIMP-1 responses. Conclusions hNSC-derived neuron/astrocyte co-cultures reproduce key aspects of LACV infection in humans and mice and are useful models to study encephalitic viruses. Specifically, we show astrocytes to be susceptible to LACV infection and that neurons and astrocytes are important drivers of the inflammatory responses seen in LACV infection through the production of proinflammatory cytokines and chemokines.

Case Report : A 10-year-old male with T1DM and recent travel to North Carolina presented to an ED with 1 day of fever, vomiting, and headaches. He was discharged home with the presumptive diagnosis of viral gastroenteritis but returned nine hours later, agitated, and unable to speak. CSF showed pleocytosis. MRI brain was normal, and EEG showed intermittent seizures. He was started on antiepileptics. Antibiotics were discontinued after negative bacterial work-up. Repeat MRI brain one week later showed enhancement in the left cerebral cortex. IVIG was started due to concern for autoimmune encephalitis. Repeat lumbar puncture was positive for La Crosse virus IgM. Discussion: This is the first case of La Crosse encephalitis (LACe) reported in Rhode Island. 1 La Crosse virus (LACv) is a ssRNA Bunyavirus transmitted by the eastern tree-hole mosquito typically between July and September. LACv is endemic to the upper Midwestern US and Appalachia. In 2018, 81 of 86 total cases reported by the CDC were pediatric. Children are more likely to present with vomiting, seizures, and focal cortical inflammation or cerebral edema on brain imaging. IgM may be negative early in the disease course. Treatment is antiepileptics and supportive care. [ABSTRACT FROM AUTHOR]

Mosquito-borne La Crosse virus (LACV; genus Orthobunyavirus, family Peribunyaviridae, order Bunyavirales) causes up to 100 annual cases of severe meningoencephalitis in children and young adults in the United States. A major virulence factor of LACV is the nonstructural protein NSs, which inhibits host cell mRNA synthesis to prevent the induction of antiviral type I interferons (IFN-α/β). To achieve this host transcriptional shutoff, LACV NSs drives the proteasomal degradation of RPB1, the large subunit of mammalian RNA polymerase II. Here, we show that NSs acts in a surprisingly rapid manner, as RPB1 degradation was commencing already at 1 h postinfection. The RPB1 degradation was partially dependent on the cellular E3 ubiquitin ligase subunit Elongin C. Consequently, removal of Elongin C, but also of the subunits Elongin A or B by siRNA transfection partially rescued general RNAP II transcription and IFN-beta mRNA synthesis from the blockade by NSs. In line with these results, LACV NSs was found to trigger the redistribution of Elongin C out of nucleolar speckles, which, however, is an epiphenomenon rather than part of the NSs mechanism. Our study also shows that the molecular phenotype of LACV NSs is different from RNA polymerase II inhibitors like α-amanitin or Rift Valley fever virus NSs, indicating that LACV is unique in involving the Elongin complex to shut off host transcription and IFN response. [ABSTRACT FROM AUTHOR]

La Crosse Virus (LACV) is an arbovirus found in Eastern Appalachia and can cause pediatric encephalitis in prepubescent children. To assess the risk and transmission of this disease, it is particularly important to understand the average population of Aedes mosquitoes, which are the vectors of this virus. We use a deterministic compartmental model to study the effects of environmental factors on the population dynamics of Aedes mosquitoes in the Knox County area. We use locally-collected mosquito population data to adjust our model outputs and find that model transitions are heavily dependent on the fluctuations of both temperature and accumulated precipitation. These findings should be considered for mosquito management in Southern Appalachia, as well as in other regions with slight modifications to our model. [ABSTRACT FROM AUTHOR]

The first step in disease pathogenesis for arboviruses is the establishment of infection following vector transmission. For La Crosse virus (LACV), the leading cause of pediatric arboviral encephalitis in North America, and other orthobunyaviruses, the initial course of infection in the skin is not well understood. Using an intradermal (ID) model of LACV infection in mice, we find that the virus infects and replicates nearly exclusively within skin-associated muscle cells of the panniculus carnosus (PC) and not in epidermal or dermal cells like most other arbovirus families. LACV is widely myotropic, infecting distal muscle cells of the peritoneum and heart, with limited infection of draining lymph nodes. Surprisingly, muscle cells are resistant to virus-induced cell death, with long term low levels of virus release progressing through the Golgi apparatus. Thus, skin muscle may be a key cell type for the initial infection and spread of arboviral orthobunyaviruses., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Infection with La Crosse virus can cause meningoencephalitis, but it is not known to cause acute flaccid paralysis (AFP). During 2008–2014, nine confirmed or probable La Crosse virus disease cases with possible AFP were reported in Ohio, USA. After an epidemiologic and clinical investigation, we determined no patients truly had AFP.

La Crosse virus (LACV) is the leading cause of pediatric viral encephalitis in North America, and is an important public health pathogen. Historically, studies involving LACV pathogenesis have focused on lineage I strains, but no former work has explored the pathogenesis between or within lineages. Given the absence of LACV disease in endemic regions where a robust entomological risk exists, we hypothesize that some LACV strains are attenuated and demonstrate reduced neuroinvasiveness. Herein, we compared four viral strains representing all three lineages to determine differences in neurovirulence or neuroinvasiveness using three murine models. A representative strain from lineage I was shown to be the most lethal, causing >50% mortality in each of the three mouse studies. However, other strains only presented excessive mortality (>50%) within the suckling mouse neurovirulence model. Neurovirulence was comparable among strains, but viruses differed in their neuroinvasive capacities. Our studies also showed that viruses within lineage III vary in pathogenesis with contemporaneous strains, showing reduced neuroinvasiveness compared to an ancestral strain from the same U.S. state (i.e., Connecticut). These findings demonstrate that LACV strains differ markedly in pathogenesis, and that strain selection is important for assessing vaccine and therapeutic efficacies.

The California serogroup of orthobunyaviruses comprises a group of mosquitoborne viruses, including La Crosse (LACV), snowshoe hare (SSHV), Tahyna (TAHV), Jamestown Canyon (JCV), and Inkoo (INKV) viruses, that cause neurologic disease in humans of differing ages with varying incidences. To determine how the pathogenesis of these viruses differs, we compared their ability to induce disease in mice and replicate and induce cell death in vitro. In mice, LACV, TAHV, and SSHV induced neurologic disease after intraperitoneal and intranasal inoculation, and JCV induced disease only after intranasal inoculation. INKV rarely induced disease, which correlated with less viral antigen in the brain than the other viruses. In vitro, all viruses replicated to high titers; however, LACV, SSHV, and TAHV induced high cell death, whereas JCV and INKV did not. Results demonstrated that CSG viruses differ in neuropathogenesis in vitro and in vivo, which correlates with the differences in pathogenesis reported in humans. [ABSTRACT FROM AUTHOR]

Urbanisation and climate change are altering the pattern of California serogroup viruses in North America. As La Crosse virus (LACV) is the most pathogenic of the California serogroup, it is important to identify changes in distribution, transmission and pathogenesis. A scoping review (ScR) was prioritised to summarise the global evidence on LACV. A comprehensive search strategy was used, identified references were screened for relevance and relevant articles were characterised. Each step was conducted by two independent reviewers using pre-tested forms. Analysis identified areas of research saturation and gaps. The ScR included 481 research articles that were mostly journal articles (78.2%) conducted in North America (90.9%) from 1969 to 2016. Most evidence focused on epidemiology (44.9%), virus characteristics (25.8%), transmission conditions (18.7%) and pathogenesis of LACV in hosts (18.3%). Fewer studies evaluated the accuracy of diagnostic tests (8.7%), the efficacy of treatments (3.5%), prevention and control strategies (3.1%), the economic burden of infection (0.6%) and social impact (0.2%) of LACV. None of the literature predicted the impact of climate change on LACV, nor were any cases reported in Canada. These findings are intended to guide research to close knowledge gaps and inform evidence-based decisions surrounding activities for the prevention and control of LACV. [ABSTRACT FROM AUTHOR]

La Crosse encephalitis is a viral disease that has emerged in new locations across the Appalachian region of the United States. Conventional wisdom suggests that ongoing emergence of La Crosse virus (LACV) could stem from the invasive Asian tiger (Aedes albopictus) mosquito. Efforts to prove this, however, are complicated by the numerous transmission routes and species interactions involved in LACV dynamics. To analyze LACV transmission by Asian tiger mosquitoes, we constructed epidemiologic models. These models accurately predict empirical infection rates. They do not, however, support the hypothesis that Asian tiger mosquitoes are responsible for the recent emergence of LACV at new foci. Consequently, we conclude that other factors, including different invasive mosquitoes, changes in climate variables, or changes in wildlife densities, should be considered as alternative explanations for recent increases in La Crosse encephalitis.

Resident cells in the skin serve as the first innate line of defense against insect-borne pathogens, but the role of these cell types in promoting or limiting arbovirus replication is not completely understood. Here, we have examined the outcome of infection of cultured human keratinocyte cells with La Crosse virus (LACV), using a spontaneously transformed cell line, HaCaT. In single cycle infections, keratinocyte HaCaT cells supported rapid and high level LACV replication, resulting in high virus yields and extensive caspase-dependent cell death. By contrast, multi-cycle LACV replication in HaCaT cells was restricted by an antiviral response elicited by the production of both IFN-β and IFN-λ. During low multiplicity LACV infections, HaCaT cell death was seen in non-infected bystander cells. Media from LACV-infected cells induced caspase-dependent killing of naïve non-infected HaCaT cells, and this bystander cell death was relieved by IFN-β neutralizing antibodies or by an inhibitor of JAK-STAT signaling. Naïve HaCaT cells showed dose-dependent killing by treatment with exogenous IFN-β but not IFN-λ. Our data suggest a model whereby keratinocytes produce IFNs which limit virus spread through both antiviral signaling and by induction of bystander cell death of potential new target cells for infection.