Louis Encephalitis Research Articles

Malaria mosquitoes, Plasmodium parasites and skin microbiota, towards control of malaria: A review

Vector-borne pathogens are transmitted by bites of infected arthropod species. The pathogens require two different host species to complete their lifecycle, usually a vertebrate and an invertebrate host, of which the latter transmits the pathogen between vertebrate hosts. Blood-feeding mosquitoes are important vectors of several human pathogens, including arboviruses namely: chikungunya, dengue fever, Rift Valley fever, yellow fever and Zika transmitted by Aedes mosquitoes amongst others. Culex mosquitoes on the other hand transmit Saint Louis and Japanese encephalitis, lymphatic filariasis, West Nile fever and Rift Valley Fever virus while Anopheles mosquitoes transmit malaria. The contact rate between vectors and their vertebrate hosts affects the intensity of pathogen transmission, and hence disease epidemiology. The ability of vectors to locate their blood hosts therefore is crucial in disease maintenance and perseverance. Host odour is known to play an important role in mosquito host-seeking behaviour and hence influences the contact rate between vectors and hosts. This literature review focuses on interactions between malaria parasites, humans, mosquitoes and how skin bacterial volatiles mediate host-seeking behaviour of mosquitoes.

WNV and SLEV coinfection in avian and mosquito hosts: impact on viremia, antibody responses, and vector competence.

West Nile virus (WNV) and St. Louis encephalitis virus (SLEV) are closely related flaviviruses that can cause encephalitis in humans and related diseases in animals. In nature, both are transmitted by Culex, with wild birds, including jays, sparrows, and robins, serving as vertebrate hosts. WNV and SLEV circulate in the same environments and have recently caused concurrent disease outbreaks in humans. The extent that coinfection of mosquitoes or birds may alter transmission dynamics, however, is not well characterized. We therefore sought to determine if coinfection alters infection kinetics and virus levels in birds and infection rates in mosquitoes. Accordingly, American robins (Turdus migratorius), two species of mosquitoes, and vertebrate and invertebrate cells were infected with WNV and/or SLEV to assess how simultaneous exposure may alter infection outcomes. There was variable impact of coinfection in vertebrate cells, with some evidence that SLEV can suppress WNV replication. However, robins had comparable viremia and antibody responses regardless of coinfection. Conversely, in Culex cells and mosquitoes, we saw a minimal impact of simultaneous exposure to both viruses on replication, with comparable infection, dissemination, and transmission rates in singly infected and coinfected mosquitoes. Importantly, while WNV and SLEV levels in coinfected mosquito midguts were positively correlated, we saw no correlation between them in salivary glands and saliva. These results reveal that while coinfection can occur in both avian and mosquito hosts, the viruses minimally impact one another. The potential for coinfection to alter virus population structure or the likelihood of rare genotypes emerging remains unknown.IMPORTANCEWest Nile virus (WNV) and St. Louis encephalitis virus (SLEV) are closely related viruses that are transmitted by the same mosquitoes and infect the same birds in nature. Both viruses circulate in the same regions and have caused concurrent outbreaks in humans. It is possible that mosquitoes, birds, and/or humans could be infected with both WNV and SLEV simultaneously, as has been observed with Zika, chikungunya, and dengue viruses. To study the impact of coinfection, we experimentally infected vertebrate and invertebrate cells, American robins, and two Culex species with WNV and/or SLEV. Robins were efficiently coinfected, with no impact of coinfection on virus levels or immune response. Similarly, in mosquitoes, coinfection did not impact infection rates, and mosquitoes could transmit both WNV and SLEV together. These results reveal that WNV and SLEV coinfection in birds and mosquitoes can occur in nature, which may impact public health and human disease risk.

Evaluation of the In2care Mosquito Station against Culex quinquefasciatus mosquitoes (Diptera: Culicidae)under semifield conditions.

Culex quinquefasciatus is an important mosquito vector responsible for the transmission of filarial worms, arthropod-borne viruses like Oropouche, St. Louis encephalitis, andWest Nile and protozoans that cause avian malaria. Due to insecticide resistance documented in Cx. quinquefasciatus populations worldwide, integrated vector management programs can benefit from new strategies to control this species. The In2Care Mosquito Station (In2Care station), a commercially available dissemination station containing pyriproxyfen (PPF) and Beauveria bassiana spores, has been shown to be effective against skip-ovipositing Aedes aegypti and Aedes albopictus in previously conducted semifield and field trials. To determine the potential of Cx. quinquefasciatus adult females to autodisseminate PPF and if the In2Care station could be used for Cx. quinquefasciatus control, we assessed its efficacy in a semifield setting against wild Cx. quinquefasciatus. We found that the In2Care station was attractive to gravid Cx. quinquefasciatus females, with a significantly higher percentage of egg rafts laid in the In2Care station compared to alternative ovipots. Adult females successfully autodisseminated PPF from the In2Care station to surrounding ovipots, leading to a significant increase in mosquito emergence inhibition. Additionally, adult Cx. quinquefasciatus exposure to B. bassiana spores significantly reduced mosquito survivorship. These results suggest that the In2Care station may be effective against Cx. quinquefasciatus in addition to Ae. aegypti and Ae. albopictus. Additional field evaluations are needed to assess impacts at the population level.

Suitable Mouse Model to Study Dynamics of West Nile Virus Infection in Culex quinquefasciatus Mosquitoes.

West Nile Virus (WNV) poses a significant global public health threat as a mosquito-borne pathogen. While laboratory mouse models have historically played a crucial role in understanding virus biology, recent research has focused on utilizing immunocompromised models to study arboviruses like dengue and Zika viruses, particularly their interactions with Aedes aegypti mosquitoes. However, there has been a shortage of suitable mouse models for investigating WNV and St. Louis encephalitis virus interactions with their primary vectors, Culex spp. mosquitoes. Here, we establish the AG129 mouse (IFN α/β/γ R-/-) as an effective vertebrate model for examining mosquito-WNV interactions. Following intraperitoneal injection, AG129 mice exhibited transient viremia lasting several days, peaking on the second or third day post-infection, which is sufficient to infect Culex quinquefasciatus mosquitoes during a blood meal. We also observed WNV replication in the midgut and dissemination to other tissues, including the fat body, in infected mosquitoes. Notably, infectious virions were present in the saliva of a viremic AG129 mouse 16 days post-exposure, indicating successful transmission capacity. These findings highlight the utility of AG129 mice for studying vector competence and WNV-mosquito interactions.

Wolbachia and mosquitoes: Exploring transmission modes and coevolutionary dynamics in Shandong Province, China.

Vector-borne diseases leave a large footprint on global health. Notable culprits include West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and Japanese encephalitis virus (JEV), all transmitted by Culex mosquitoes. Chemical insecticides have been widely used to reduce the spread of mosquito-borne diseases. Still, mosquitoes are becoming more and more resistant to most chemical insecticides which cause particular harm to the ecology. Wolbachia belongs to the family Ehrlichiaceae in the order Rickettsiales and is a matrilineally inherited endosymbiont present in 60% of insects in nature. Wolbachia is capable of inducing a wide range of reproductive abnormalities in its hosts, such as cytoplasmic incompatibility, and can alter mosquito resistance to pathogen infection. Wolbachia has been proposed as a biological alternative to chemical vector control, and specific research progress and effectiveness have been achieved. Despite the importance of Wolbachia, this strategy has not been tested in Culex pipiens pallens, the most prevalent mosquito species in Shandong Province, China. Little is known about how the mass release of Wolbachia-infected mosquitoes may impact the genetic structure of Culex pipiens pallens, and how the symbiotic bacterium Wolbachia interacts with mitochondria during host mosquito transmission. Based on the population genetic structure of Culex pipiens pallens in Shandong Province, this study investigated the infection rate and infection type of Wolbachia in Shandong Province and jointly analysed the evolutionary relationship between the host mosquito and the symbiotic bacterium Wolbachia. Our study showed that Wolbachia naturally infected by Culex pipiens pallens in Shandong Province was less homologous to Wolbachia infected by Aedes albopictus released from mosquito factory in Guangzhou. Our results also show that Culex pipiens pallens is undergoing demographic expansion in Shandong Province. The overall Wolbachia infection rate of Culex pipiens pallens was 92.8%, and a total of 15 WSP haplotypes were detected. We found that the genetic diversity of Wolbachia was low in Culex pipiens pallens from Shandong Province, and the mosquitoes were infected only with type B Wolbachia. Visualizing the relationship between Culex pipiens pallens and Wolbachia using a tanglegram revealed patterns of widespread associations. A specific coevolutionary relationship exists between the host mosquito and Wolbachia. Knowledge of this mosquito-Wolbachia relationship will provide essential scientific information required for Wolbachia-based vector control approaches in Shandong Province and will lead to a better understanding of the diversity and evolution of Wolbachia for its utility as a biocontrol agent.

Global diversity of Aedes species (Diptera: Culicidae) relevant to public health

Mosquito-borne arboviruses represent one of the main challenges for public health today. The Aedes genus includes more than 950 mosquito species, many of which cause discomfort through their bites and are disease vectors. However, little is known about this biodiversity and its role in the context of public health. This study aimed to list the worldwide diversity of mosquitoes belonging to the Aedes genus with the potential to transmit pathogens to humans. An integrative literature review was conducted. The articles were searched during July 2024, in the following databases: Lilacs, Scielo, Science Direct and PubMed. A total of 38 species of mosquitoes belonging to the Aedes genus with the potential to transmit pathogens to humans were listed. The literature consulted reports a total of 44 pathogens potentially transmitted by Aedes mosquitoes, including Dengue, Zika virus, Chikungunya, yellow fever, Rift Valley fever virus, West Nile virus, Ross River virus, Japanese encephalitis virus, Saint Louis encephalitis virus, and equine encephalitis virus. This study provides useful information to support further research and the development of actions to monitor and control mosquito species with vector potential.

Host-Feeding Behavior of Mosquitoes in the Florida Everglades.

Background: West Nile virus (WNV), Everglades virus (EVEV), and five species of Orthobunyavirus were isolated from mosquitoes collected in the Everglades in 2016-2017. Prior studies of blood meals of mosquitoes in southern Florida have related findings to acquisition and transmission of EVEV, St. Louis encephalitis virus, and WNV, but not the Orthobunyavirus viruses associated with the subgenus Melanoconion of the genus Culex. Materials and Methods: In the present study, blood-fed mosquitoes were collected in the Everglades in 2016, 2017, 2021, and 2022, and from an industrial site in Naples, FL in 2017. Blood meals were identified to host species by PCR assays using mitochondrial cytochrome b gene. Results: Blood meals were identified from Anopheles crucians complex and 11 mosquito species captured in the Florida Everglades and from 3 species collected from an industrial site. The largest numbers of blood-fed specimens were from Culex nigripalpus, Culex erraticus, Culex cedecei, and Aedes taeniorhynchus. Cx. erraticus fed on mammals, birds, and reptiles, particularly American alligator. This mosquito species could transmit WNV to American alligator in the wild. Cx. nigripalpus acquired blood meals primarily from birds and mammals and frequently fed on medium-sized mammals and white-tailed deer. Water and wading birds were the primary avian hosts for Cx. nigripalpus and Cx. erraticus in the Everglades. Wading birds are susceptible to WNV and could serve as reservoir hosts. Cx. cedecei fed on five species of rodents, particularly black and hispid cotton rats. EVEV and three different species of Orthobunyavirus have been isolated from the hispid cotton rat and Cx. cedecei in the Everglades. Cx. cedecei is likely acquiring and transmitting these viruses among hispid cotton rats and other rodents. The marsh rabbit was a frequent host for An. crucians complex. An. crucians complex, and other species could acquire Tensaw virus from rabbits. Conclusions: Our study contributes to a better understanding of the host and viral associations of mosquito species in southwestern Florida.

Projections of Aedes and Culex mosquitoes across North and South America in response to climate change

IntroductionClimate and land use change may cause the geographical range of mosquitoes to expand, shift, or contract, ultimately changing what communities are at risk for contracting mosquito-borne diseases. Across North and South America, mosquitoes from the Aedes and Culex genera are vectors for numerous diseases, including chikungunya, dengue, various equine encephalitis viruses, Saint Louis encephalitis virus, West Nile virus, yellow fever virus, and Zika virus. The goal of our study was to project the geographical distributions of important mosquito vectors across North and South America in response to climate change, which is important to inform public health planning. MethodsWe used an ecological niche model and future projections of climate and land use to project the geographical ranges of two Aedes species (Ae. aegypti, Ae. albopictus) and seven Culex species (Cx. erraticus, Cx. nigripalpus, Cx. pipiens, Cx. quinquefasciatus, Cx. restuans, Cx. salinarius, Cx. tarsalis) over North and South America in response to both a high (Shared Socioeconomic Pathway [SSP] 5, Representative Concentration Pathway [RCP] 8.5) and moderate (SSP2 RCP4.5) climate warming scenario through years 2050 and 2090. ResultsFor SSP5 RCP8.5, six species (Ae. aegypti, Ae. albopictus, Cx. erraticus, Cx. nigripalpus, Cx. quinquefasciatus, Cx. restuans) are projected to expand in geographical range, two species (Cx. pipiens, Cx. tarsalis) are projected to shift in geographical range, and one species (Cx. salinarius) is projected to nearly remain the same. Five species (Ae. aegypti, Ae. albopictus, Cx. erraticus, Cx. nigripalpus, Cx. quinquefasciatus) show the largest increase in high habitat suitability (>0.5 on a 0–1 scale) for SSP5 RCP8.5, three species (Cx. pipiens, Cx. restuans, Cx. tarsalis) show the largest increase for SSP2 RCP4.5, and one species (Cx. salinarius) shows a relatively small decrease in response to both scenarios. ConclusionsWe found that all nine species responded resiliently to climate change under both the high and moderate climate warming scenario, suggesting mosquito-borne disease is likely to be a continued threat in response to climate change. The projected geographical ranges can be used to inform disease risk analyses and mitigation strategies in response to climate change.

Japanese encephalitis virus NS1 and NS1' protein disrupts the blood-brain barrier through macrophage migration inhibitory factor-mediated autophagy.

Flaviviruses in the Japanese encephalitis virus (JEV) serogroup, such as JEV, West Nile virus, and St. Louis encephalitis virus, can cause severe neurological diseases. The nonstructural protein 1 (NS1) is a multifunctional protein of flavivirus that can be secreted by infected cells and circulate in the host bloodstream. NS1' is an additional form of NS1 protein with 52 amino acids extension at its carboxy-terminal and is produced exclusively by flaviviruses in the JEV serogroup. In this study, we demonstrated that the secreted form of both NS1 and NS1' can disrupt the blood-brain barrier (BBB) of mice, with NS1' exhibiting a stronger effect. Using the in vitro BBB model, we found that treatment of soluble recombinant JEV NS1 or NS1' protein increases the permeability of human brain microvascular endothelial cells (hBMECs) and leads to the degradation of tight junction proteins through the autophagy-lysosomal pathway. Consistently, NS1' protein exhibited a more pronounced effect compared to NS1 in these cellular processes. Further research revealed that the increased expression of macrophage migration inhibitory factor (MIF) is responsible for triggering autophagy after NS1 or NS1' treatment in hBMECs. In addition, TLR4 and NF-κB signaling was found to be involved in the activation of MIF transcription. Moreover, administering the MIF inhibitor has been shown to decrease viral loads and mitigate inflammation in the brains of mice infected with JEV. This research offers a novel perspective on the pathogenesis of JEV. In addition, the stronger effect of NS1' on disrupting the BBB compared to NS1 enhances our understanding of the mechanism by which flaviviruses in the JEV serogroup exhibit neurotropism.IMPORTANCEJapanese encephalitis (JE) is a significant viral encephalitis worldwide, caused by the JE virus (JEV). In some patients, the virus cannot be cleared in time, leading to the breach of the blood-brain barrier (BBB) and invasion of the central nervous system. This invasion may result in cognitive impairment, behavioral disturbances, and even death in both humans and animals. However, the mechanism by which JEV crosses the BBB remains unclear. Previous studies have shown that the flavivirus NS1 protein plays an important role in causing endothelial dysfunction. The NS1' protein is an elongated form of NS1 protein that is particularly produced by flaviviruses in the JEV serogroup. This study revealed that both the secreted NS1 and NS1' of JEV can disrupt the BBB by breaking down tight junction proteins through the autophagy-lysosomal pathway, and NS1' is found to have a stronger effect compared to NS1 in this process. In addition, JEV NS1 and NS1' can stimulate the expression of MIF, which triggers autophagy via the ERK signaling pathway, leading to damage to BBB. Our findings reveal a new function of JEV NS1 and NS1' in the disruption of BBB, thereby providing the potential therapeutic target for JE.

The Susceptibility of Chickens to Zika Virus: A Comprehensive Study on Age-Dependent Infection Dynamics and Host Responses.

Zika virus (ZIKV) remains a public health concern, with epidemics in endemic regions and sporadic outbreaks in new areas posing significant threats. Several mosquito-borne flaviviruses that can cause human illness, including West Nile, Usutu, and St. Louis encephalitis, have associations with birds. However, the susceptibility of chickens to ZIKV and their role in viral epidemiology is not currently known. We investigated the susceptibility of chickens to experimental ZIKV infection using chickens ranging from 1-day-old chicks to 6-week-old birds. ZIKV caused no clinical signs in chickens of all age groups tested. Viral RNA was detected in the blood and tissues during the first 5 days post-inoculation in 1-day and 4-day-old chicks inoculated with a high viral dose, but ZIKV was undetectable in 6-week-old birds at all timepoints. Minimal antibody responses were observed in 6-week-old birds, and while present in younger chicks, they waned by 28 days post-infection. Innate immune responses varied significantly between age groups. Robust type I interferon and inflammasome responses were measured in older chickens, while limited innate immune activation was observed in younger chicks. Signal transducer and activator of transcription 2 (STAT2) is a major driver of host restriction to ZIKV, and chicken STAT2 is distinct from human STAT2, potentially contributing to the observed resistance to ZIKV infection. The rapid clearance of the virus in older chickens coincided with an effective innate immune response, highlighting age-dependent susceptibility. Our study indicates that chickens are not susceptible to productive ZIKV infection and are unlikely to play a role in the ZIKV epidemiology.

CAPTURING TRENDS IN ARBOVIRAL SURVEILLANCE: COMPARING TRADITIONAL REVERSE TRANSCRIPTION PCR AND QUANTITATIVE REVERSE TRANSCRIPTION PCR ASSAYS

West Nile virus (WNV) and St. Louis encephalitis virus (SLEV) pose a significant public health threat in the United States. These viruses are known to adapt rapidly to new amplifying hosts and geographic environments, making effective surveillance critical for public health efforts. This study evaluated the effectiveness of traditional reverse transcription polymerase chain reaction (RT-PCR) for surveillance purposes compared to quantitative RT-PCR (RT-qPCR) in detecting WNV and SLEV in mosquito pools. Mosquito pools were collected and screened for WNV and SLEV over a 10-year period. This study found an increase in the number of flavivirus-positive yet WNV-/SLEV-negative mosquito pools during 2018 compared to previous years. Quantitative RT-PCR detected more positive WNV and SLEV pools compared to traditional RT-PCR, eliminating false negatives and identifying false positives. The findings underscore the importance of using RT-qPCR for arboviral surveillance to accurately detect circulating viruses and enable timely public health interventions. Changes in local trends in mosquito-borne viruses and vector populations have the potential to impact public health, emphasizing the need for proactive surveillance measures.

Serological survey reveals enzootic circulation of St. Louis encephalitis and West Nile viruses in semiarid Monte ecosystem of Argentina

St. Louis encephalitis virus (SLEV) and West Nile virus (WNV) are arboviruses transmitted by Culex mosquitoes and amplified in avian hosts. The present study aimed to investigate the presence and seasonal circulation of SLEV and WNV in La Rioja province, within the semiarid ecoregion of the Monte, Argentina. Over a two-year period, avian sera were collected and tested for neutralizing antibodies against SLEV and WNV. Our results reveal the enzootic activity of both viruses in this challenging environment. SLEV seroprevalence was 4.5% (35/778), with higher activity in spring (2016) and autumn (2017). WNV seroprevalence was 3.5% (27/778), peaking during the summer 2016–2017. Greater seroprevalence for SLEV in 2016 was detected for the Lark-like Brushrunner (Coryphistera alaudina) and the Short-billed Canastero (Asthenes baeri) and in 2017 for the Black-crested Finch (Lophospingus pusillus) and Lark-like Brushrunner, whereas for WNV greater seroprevalence in 2016 was detected for the Picui Ground Dove (Columbina picui) and in 2017 for the Lark-like Brushrunner and Band-tailed Seedeater (Catamenia analis). Additionally, five avian individuals experienced seroconversion during the sampling period, namely the Lark-like Brushrunner and White-fronted Woodpecker (Melanerpes cactorum) for SLEV, and the Lark-like Brushrunner, Greater Wagtail Tyrant (Stigmatura budytoides) and Many-colored Chaco Finch (Saltatricula multicolor) for WNV. The study highlights the persistence and circulation of these viruses in a semiarid ecosystem, raising questions about overwintering mechanisms and transmission dynamics. This research contributes to understanding arbovirus ecology in diverse environments. Further investigations are needed to assess the specific mechanisms facilitating virus persistence in the Monte ecoregion.

Haemovigilance survey and screening strategy for arthropod-borneviruses in blood donors from Argentina.

Arthropod-borne viruses (arboviruses) count among emerging infections, which represent a major challenge for transfusion safety worldwide. To assess the risk of arboviruses-transmission by transfusion (ATT), we performed a survey to evaluate the potential threat for transfusion safety. Samples were retrospectively and randomly collected from donors who donated during the peak of dengue incidence in Cordoba (years: 2016and 2019-2022). A cost-efficient strategy for molecular screening was implemented with a nucleic acid test (NAT) configured with Flavivirus and Alphavirus-universal degenerated primers targeting conserved gene regions. Besides, we evaluated the neutralizing antibody (NAb) prevalence by plaque reduction neutralization test (PRNT). A total of 1438 samples were collected. Among the NAT-screened samples, one resulted positive for Flavivirus detection. Subsequent sequencing of the PCR product revealed Saint Louis Encephalitis Virus (SLEV) infection (GeneBank accession number OR236721). NAb prevalence was 2.95% for anti-Dengue, 9.94% anti-SLEV, 1.09% anti-West Nile Virus, and 0% anti-Chikungunya. One of the NAb-positive samples also resulted positive for IgM against SLEV but negative by ARN detection. This is the first haemovigilance study developed in Argentina that evaluates the potential risk of ATT and the first research to determine the prevalence of NAb against Flavivirus through PNRT to avoid possible cross-reactions between Ab against Flavivirus. Herein, the finding of one SLEV-viremic donor and the detection of anti-SLEV IgM in a different donor demonstrated a potential threat for transfusion safety and emphasized the need for increased vigilance and proactive measures to ensure the safety of blood supplies.

Viral Interference between the Insect-Specific Virus Brejeira and the Saint Louis Encephalitis Virus In Vitro.

The Saint Louis encephalitis virus (SLEV) is an encephalitogenic arbovirus (Flaviviridae family) that has a wide geographical distribution in the western hemisphere, especially in the Americas. The negevirus Brejeira (BREV) was isolated for the first time in Brazil in 2005. This study aimed to verify the existence of a possible interfering effect of BREV on the course of SLEV infection and vice versa. We used clone C6/36 cells. Three combinations of MOIs were used (SLEV 0.1 × BREV 1; SLEV 1 × BREV 0.1; SLEV 1 × BREV 1) in the kinetics of up to 7 days and then the techniques of indirect immunofluorescence (IFA), a plaque assay on Vero cells, and RT-PCR were performed. Our results showed that the cytopathic effect (CPE) caused by BREV was more pronounced than the CPE caused by SLEV. Results of IFA, the plaque assay, and RT-PCR showed the suppression of SLEV replication in the co-infection condition in all the MOI combinations used. The SLEV suppression was dose-dependent. Therefore, the ISV Brejeira can suppress SLEV replication in Aedes albopictus cells, but SLEV does not negatively interfere with BREV replication.

Culex erraticus (Diptera: Culicidae) utilizes gopher tortoise burrows for overwintering in North Central Florida.

Mosquito-borne diseases represent a significant threat to human and animal health in the United States. Several viruses, including West Nile, Saint Louis encephalitis, and Eastern equine encephalitis are endemic. In humans, the disease is typically detected during the summer months, but not during the winter months. The ability of these viruses to reemerge year after year is still not fully understood, but typically involves persistence in a reservoir host or vector during periods of low transmission. Mosquito species are known to overwinter at different life stages (adults, larvae, or eggs) in manufactured or natural sites. Gopher tortoise burrows are known to serve as refuge for many vertebrate and invertebrate species in pine savannas. In this study, we surveyed the interior of gopher tortoise burrows for overwintering mosquitoes. We identified 4 species (Anopheles crucians s.l., Culex erraticus, Mansonia dyari, and Uranotaenia sapphirina). Cx. erraticus was the most abundant, and its presence and abundance increased in winter months, implying that this species utilized gopher tortoise burrows for overwintering. Bloodfed Cx. erraticus and An. crucians s.l. females were detected. While An. crucians s.l. fed exclusively on the white-tailed deer, Cx. erraticus had a more diverse host range but fed primarily on the gopher tortoise. Tortoises and other long-lived reptiles like the American alligator have been shown to sustain high viremia following West Nile virus (WNV) and Eastern equine encephalitis virus (EEEV) infection and therefore could play a role in the maintenance of these viruses. In addition, Cx. erraticus is naturally infected with WNV and is a known bridge vector for EEEV. As such, these overwintering sites may play a role in perpetuating over-winter arboviral activity in Florida.

Usutu virus, an emerging arbovirus with One Health importance.

Usutu virus (USUV, Flaviviridae) is an emerging arbovirus that has led to epizootic outbreaks in birds and numerous human neuroinvasive disease cases in Europe. It is maintained in an enzootic cycle with Culex mosquitoes and passerine birds, a transmission cycle that is shared by West Nile virus (WNV) and St. Louis encephalitis virus (SLEV), two flaviviruses that are endemic in the United States. USUV and WNV co-circulate in Africa and Europe, and SLEV and WNV co-circulate in North America. These three viruses are prime examples of One Health issues, in which the interactions between humans, animals, and the environments they reside in can have important health impacts. The three facets of One Health are interwoven throughout this article as we discuss the mechanisms of flavivirus transmission and emergence. We explore the possibility of USUV emergence in the United States by analyzing the shared characteristics among USUV, WNV, and SLEV, including the role that flavivirus co-infections and sequential exposures may play in viral emergence. Finally, we provide insights on the importance of integrated surveillance programs as One Health tools that can be used to mitigate USUV emergence and spread.

Phenotypical Screening of an MMV Open Box Library and Identification of Compounds with Antiviral Activity against St. Louis Encephalitis Virus.

St. Louis encephalitis virus (SLEV) is a neglected mosquito-borne Flavivirus that may cause severe neurological disease in humans and other animals. There are no specific treatments against SLEV infection or disease approved for human use, and drug repurposing may represent an opportunity to accelerate the development of treatments against SLEV. Here we present a scalable, medium-throughput phenotypic cell culture-based screening assay on Vero CCL81 cells to identify bioactive compounds that could be repurposed against SLEV infection. We screened eighty compounds from the Medicines for Malaria Venture (MMV) COVID Box library to identify nine (11%) compounds that protected cell cultures from SLEV-induced cytopathic effects, with low- to mid-micromolar potencies. We validated six hit compounds using viral plaque-forming assays to find that the compounds ABT-239, Amiodarone, Fluphenazine, Posaconazole, Triparanol, and Vidofludimus presented varied levels of antiviral activity and selectivity depending on the mammalian cell type used for testing. Importantly, we identified and validated the antiviral activity of the anti-flavivirus nucleoside analog 7DMA against SLEV. Triparanol and Fluphenazine reduced infectious viral loads in both Vero CCL81 and HBEC-5i cell cultures and, similar to the other validated compounds, are likely to exert antiviral activity through a molecular target in the host.

Emergence of genotype III St. Louis encephalitis virus in the western United States potentially linked to a wetland in Argentina

St. Louis encephalitis virus (SLEV) is endemic in the Americas and its transmission networks involve Culex mosquitoes and avian species. In 2015, a human encephalitis outbreak took place in Arizona and California, indicating the re-emergence of this pathogen in the US. Viral strains isolated in that outbreak belong to genotype III SLEV previously detected only in South America. In this study, genotype III SLEV was detected in mosquitoes collected in Mar Chiquita Lagoon (Córdoba, Argentina), an overwintering site for numerous migratory bird species. The genotype III SLEV sequence detected in this site shares the closest known ancestor with those introduced in Arizona in 2015. Our results highlight the potential significance of wetlands as key sites for arbovirus maintenance and emergence.

Effective population size of Culex quinquefasciatus under insecticide-based vector management and following Hurricane Harvey in Harris County, Texas.

Introduction: Culex quinquefasciatus is a mosquito species of significant public health importance due to its ability to transmit multiple pathogens that can cause mosquito-borne diseases, such as West Nile fever and St. Louis encephalitis. In Harris County, Texas, Cx. quinquefasciatus is a common vector species and is subjected to insecticide-based management by the Harris County Public Health Department. However, insecticide resistance in mosquitoes has increased rapidly worldwide and raises concerns about maintaining the effectiveness of vector control approaches. This concern is highly relevant in Texas, with its humid subtropical climate along the Gulf Coast that provides suitable habitat for Cx. quinquefasciatus and other mosquito species that are known disease vectors. Therefore, there is an urgent and ongoing need to monitor the effectiveness of current vector control programs. Methods: In this study, we evaluated the impact of vector control approaches by estimating the effective population size of Cx. quinquefasciatus in Harris County. We applied Approximate Bayesian Computation to microsatellite data to estimate effective population size. We collected Cx. quinquefasciatus samples from two mosquito control operation areas; 415 and 802, during routine vector monitoring in 2016 and 2017. No county mosquito control operations were applied at area 415 in 2016 and 2017, whereas extensive adulticide spraying operations were in effect at area 802 during the summer of 2016. We collected data for eighteen microsatellite markers for 713 and 723 mosquitoes at eight timepoints from 2016 to 2017 in areas 415 and 802, respectively. We also investigated the impact of Hurricane Harvey's landfall in the Houston area in August of 2017 on Cx. quinquefasciatus population fluctuation. Results: We found that the bottleneck scenario was the most probable historical scenario describing the impact of the winter season at area 415 and area 802, with the highest posterior probability of 0.9167 and 0.4966, respectively. We also detected an expansion event following Hurricane Harvey at area 802, showing a 3.03-fold increase in 2017. Discussion: Although we did not detect significant effects of vector control interventions, we found considerable influences of the winter season and a major hurricane on the effective population size of Cx. quinquefasciatus. The fluctuations in effective population size in both areas showed a significant seasonal pattern. Additionally, the significant population expansion following Hurricane Harvey in 2017 supports the necessity for post-hurricane vector-control interventions.

Molecular and Cellular Mechanisms Underlying Neurologic Manifestations of Mosquito-Borne Flavivirus Infections.

Flaviviruses are a family of enveloped viruses with a positive-sense RNA genome, transmitted by arthropod vectors. These viruses are known for their broad cellular tropism leading to infection of multiple body systems, which can include the central nervous system. Neurologic effects of flavivirus infection can arise during both acute and post-acute infectious periods; however, the molecular and cellular mechanisms underlying post-acute sequelae are not fully understood. Here, we review recent studies that have examined molecular and cellular mechanisms that may contribute to neurologic sequelae following infection with the West Nile virus, Japanese encephalitis virus, Zika virus, dengue virus, and St. Louis encephalitis virus. Neuronal death, either from direct infection or due to the resultant inflammatory response, is a common mechanism by which flavivirus infection can lead to neurologic impairment. Other types of cellular damage, such as oxidative stress and DNA damage, appear to be more specific to certain viruses. This article aims to highlight mechanisms of cellular damage that are common across several flavivirus members and mechanisms that are more unique to specific members. Our goal is to inspire further research to improve understanding of this area in the hope of identifying treatment options for flavivirus-associated neurologic changes.